A wet extraction process improved by acidic and chelating conditions

Abstract

Disclosed herein are processes for extracting natural products from a biomass. The processes include forming an acidic feedstock stream having a pH of at most 4.0 and containing a biomass and an aqueous salt solution; contacting the acidic feedstock stream with an extraction solvent to form a dispersion; separating the dispersion into multiple layers, the layers including: a solvent extract layer containing at least one hydrophobic natural product and at least a portion of the extraction solvent, a raffinate layer containing at least a portion of the aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer. Systems configured to perform the processes are also disclosed.

Description

A WET EXTRACTION PROCESS IMPROVED BY ACIDIC AND CHELATING CONDITIONSTECHNICAL FIELD

[0001] The present disclosure relates to processes and systems for extracting natural products from a biomass. Also, the present disclosure relates to uses of the systems to extract natural products from a biomass.BACKGROUND

[0002] There is an increasing interest in using biomass as a key intermediate for a plethora of sustainable products, such as a source of renewable energy, as a mode to safely and efficiently capture carbon dioxide from the atmosphere for carbon sequestration, as a source of natural carotenoids and as a renewable source of chemical intermediates.

[0003] KR 102387853 discloses sulfuric acid catalyzed biodiesel production processes for microalgae.

[0004] US 2020253238 discloses methods for recovering material from a biomass that involve solvent-free fractionation of lipids and proteins with the aid of concentrated organic acid. Here, a large amount of acid is used to force lipid and protein-containing fractions out from a biomass. The acid concentration used in these fractionations can range from 0.1 M to 0.5M or higher. These processes also involve the dewatering of the biomass before extraction and no experimental examples involving an algal biomass are disclosed. The processes disclosed in US 2020253238 also produce protein fractions low in protein content (e.g., 20-45%).

[0005] US 8,906,236 discloses methods and processes for the recovery of nutrients from non-organic phases produced during recovery of oleaginous compounds from biomass. These methods and processes involve a harsh heat and acid treatment prior to wet extraction with or without solvent. Here, dewatering is done with the aid of strong heat (e.g., 200 to 500°C) and no solvent is used during the dewatering. Also, the salinity of the biomass is reduced to a minimum prior to strong acid treatment (e.g., sulfuric acid), and nomild organic acids are mentioned. Protein recovery and metal chelation are not addressed in this patent.

[0006] US 10,342,772 discloses solvent-free processes for obtaining microbial oil from microbial cells. There is no disclosure of using these processes to extract proteins from microbial cells. Acidic hydrolysis and metal chelation is not demonstrated or discussed in this patent.

[0007] US 4,341 ,038 discloses processes wherein D. salina are thickened by raising the pH to above 10 prior to performing a wet extraction process. These processes do not involve an acidification prior to extraction.

[0008] US 4,680,314 discloses processes that use a flocculating agent to harvest algae and then the pH is adjusted to decomplex the flocculant. These processes run the risk of damaging the algal biomass due to the inclusion of a flocculating agent during harvesting.

[0009] US 2010233761 A1 discloses processes for fractionating biomass that include permeating walls of biomass cells, liberating cell products from the cells, and fractionating and recovering the liberated cell products and derivatives. There is no disclosure of salinity or neutral salts (e.g., NaCI) in the systems used in the extraction processes. The processes also heat the biomass feed to temperatures ranging from 25°C to 200°C before performing an extraction.Metal chelation is not disclosed, and the processes do not teach dewatering the biomass prior to fractionation (to minimize volumes prior to separation).

[0010] US 2011256594 A1 discloses processes that involve the removal of nitrogen from a chlorophyll or pheophytin containing biomass (e.g., algae). There is no mention of salinity or neutral salts being present in the biomass and there is no disclosure of using a multi-product approach to recover natural products from the biomass. There is also no disclosure of using metal chelation.

[0011] One drawback from using algal biomass as a renewable energy source is that the biomass is wet. Because algae grow in water, biomass harvested from algae includes water which can lead to complicated extraction processes.

[0012] To avoid these complicated extraction processes, algal biomass is usually dried before extraction of valuable products. A drawback of drying the algal biomass before extraction is that the drying of an algal biomass is an energy-intensive process, thus requiring large amounts of heat and resources.Bypassing this drying step could make processes of using algal biomass as a renewable energy source significantly more feasible.

[0013] The challenge with using wet biomass in extraction processes comes from the water acting as an “extra” phase between an extraction solvent and the biomass. The presence of this “extra” phase can lead to many disadvantages during extraction, such as: decreased contact surface area between the biomass and the extraction solvent, reduced decantation kinetics, high biomass loss into the water phase, reduced extraction of natural products into the extraction solvent, increased levels of metals in the extract phase, increased soluble protein loss in the water phase, increased microbial contamination and increased viscosity of the algal biomass.

[0014] The following disclosure provides more economical and efficient processes and systems for extracting natural products from a wet algal biomass as a solution to the foregoing issues.SUMMARY

[0015] Disclosed herein is a process for extracting natural products from a biomass, the process including (i.e., comprising) at least one or more of: forming an acidic feedstock stream having a pH of at most 4.0 and containing a biomass and an aqueous salt solution; contacting the acidic feedstock stream with an extraction solvent to form a dispersion; separating the dispersion into multiple layers, the layers including: a solvent extract layer containing at least one hydrophobic natural product and at least a portion of the extraction solvent, a raffinate layer containing at least a portion of the aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer.

[0016] Disclosed herein is also a process for extracting natural products from a biomass, the process including at least one or more of: forming an acidic feedstock stream having a pH of at most 4.0 and containing a biomass and an aqueous salt solution; contacting the acidic feedstock stream with a first solvent in a first contacting unit to form a first dispersion; allowing the first dispersion to phase separate into a first separated liquid phase and a first remaining dispersion; removing at least a portion of the first separated liquid phase fromthe first remaining dispersion; contacting the first remaining dispersion with a second solvent in a second contacting unit to form a second dispersion; allowing the second dispersion to phase separate into a biomass-rich phase, a heavy separated liquid phase and a light separated liquid phase; and recovering at least one or more of the biomass-rich phase, the heavy separated liquid phase, and / or the light separated liquid phase for output as products.

[0017] Disclosed herein is also a process for extracting natural products from a biomass, the process including at least one or more of: obtaining a feedstock stream containing the biomass and an aqueous salt solution, the feedstock stream having a pH of more than 4.0 or at least 5.0; transferring the feedstock stream to an extraction zone; introducing an additive stream containing an extraction solvent and at least one acidic additive and / or one buffering agent into the extraction zone to form a dispersion, the least one acidic additive being either a Bronsted acid, an acidic chelating agent or a combination thereof, wherein the dispersion contains an aqueous phase having a pH of at most 4.0; separating the dispersion into multiple layers, the layers including: a solvent extract layer containing at least one hydrophobic natural product and at least a portion of the extraction solvent, a raffinate layer containing at least a portion of the aqueous salt solution, and a rag layer containing at least a portion of a lipid- depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer.

[0018] Disclosed herein is a system for extracting natural products from a biomass, the system including at least one or more of: a mixing zone configured to mix a feedstock stream with an additive stream, and contain an acidic feedstock stream having a pH of at most 4.0; an extraction zone configured to mix an acidic feedstock stream having a pH of at most 4.0 with an extraction solvent, and contain a dispersion having an aqueous phase with a pH of at most 4.0; and a separation zone configured to separate a dispersion having an aqueous phase with a pH of at most 4.0 into a solvent extract layer containing at least one hydrophobic natural product and at least a portion of an extraction solvent, a raffinate layer containing at least a portion of an aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass.

[0019] Disclosed herein is also a system for extracting natural products from a biomass, the system including at least one or more of: an acidic feedstock stream import line; a first contacting unit in communication with the acidic feedstock stream import line; a first solvent import line in communication with the first contacting unit; a first effluent export line in communication with the first contacting unit; a second contacting unit in communication with the first contacting unit; a second solvent import line in communication with the second contacting unit; and a first extraction line in communication with the second contacting unit.

[0020] Disclosed herein is also a use of the systems or processes of the present disclosure for extracting natural products from a biomass and / or for separating a dispersion into multiple layers, the layers including: a solvent extract layer, a raffinate layer and a rag layer.BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Other features and advantages of the methods and systems disclosed herein will be apparent to those skilled in the art reading the following detailed description in conjugation with the exemplary embodiments illustrated in the drawings, wherein:

[0022] FIG. 1 shows a flow diagram of an exemplary embodiment of an extracting process for the isolation of natural products from a biomass, the process involving forming an acidic feedstock stream in a mixing zone, forming a dispersion from the acidic feedstock stream in an extraction zone and separating the dispersion into multiple layers in a separation zone.

[0023] FIG. 2 shows a flow diagram of an exemplary embodiment of an extracting process for the isolation of natural products from a biomass, the process involving forming an acidic feedstock stream, contacting the acidic feedstock stream with a first solvent in a first contacting unit to form a first dispersion; separating a liquid phase from the first dispersion to form a first remaining dispersion; contacting the first remaining dispersion with a second solvent in a second contacting unit to form a second dispersion and allowing the second dispersion to phase separate into multiple layers.

[0024] FIG. 3 shows a flow diagram of an exemplary embodiment of an extracting process for the isolation or recovery of natural products from a biomass, the process involving forming an acidic feedstock stream in an extraction zone, forming a dispersion from the acidic feedstock stream in the extraction zone and separating the dispersion into multiple layers in a separation zone.

[0025] FIG. 4 shows a flow diagram of an exemplary embodiment of an extracting process for the isolation of natural products from a biomass, the process involving forming a dispersion from a feedstock stream and an additive stream containing an extraction solvent in an extraction zone, and separating the dispersion into multiple layers in a separation zone.

[0026] FIG. 5 shows a flow diagram of the exemplary process used in Examples 1-3 to extract natural products from an algal feedstock.

[0027] FIG. 6 shows a flow diagram of the exemplary process used in Examples 4-8 to extract natural products from a feedstock stream.

[0028] FIG. 7 shows a flow diagram of exemplary embodiments of extracting processes used in Examples 9-11 for the isolation of natural products from a biomass.

[0029] DETAILED DESCRIPTION

[0030] In extraction processes involving wet biomass, several parameters play a key role in the efficiency of the extraction. These factors include, but are not limited to, solvent, solvent to biomass ratio, extraction medium, temperature, pressure, agitation type and rate, and the type of extractor used. In addition to addressing the physical parameters that influence the efficacy of these liquid- liquid-solid extraction process, addressing the chemical parameters is also important.

[0031] One chemical parameter that has been discovered to influence the efficacy of extraction processes on wet biomass is acidity / basicity, or simply pH. Algae predominately grow in algal growth medium having a pH of 7-8 (Kumar K. et al. Renewable and Sustainable Energy Reviews 51 (2015) 875-885.). If algae originating from these growth conditions are subjected to extraction processes, then those extraction processes would occur at near neutral pH.Performing extraction processes at these pH levels on wet biomass has been found to increase the negative effects that the water phase has on the extraction efficiency.

[0032] After conducting a series of experiments and extensive research, it has surprisingly been discovered that an improved extraction process of natural products from a wet biomass stream can occur if the pH of the wet biomass stream is adjusted to a pH of at most 4.0 before extraction of the natural products.

[0033] When the pH of a wet biomass stream is adjusted to be at most 4.0, mass transfer during extraction and phase separation kinetics during decantation are improved. While not wishing to be bound by any particular theory, it is considered that this improvement occurs in part due to the change in polarity and charges of compounds in the aqueous medium at these pH levels. Additionally, because pH plays a key role in chemical reactions such as hydrolysis, these improvements can occur in part due to the hydrolysis of compounds present in the wet biomass stream (e.g., hydrolysis of glycolipids that are chemically bound to the biomass matrix via ester linkages).

[0034] By carefully selecting the acidic additives that the wet biomass stream comes in contact with before extraction, it has been discovered that one can both adjust the pH of the wet biomass stream to a pH of at most 4.0 and utilize a hydrophilic acidic chelating agent to trap at least a portion of unwanted metals in the wet biomass stream in a raffinate phase formed from the extraction process, thereby reducing the metal content in the extracted natural products. Furthermore, because biomass is as a biological system affected by changes in pH, adjusting the pH of the wet biomass stream to a pH of at most 4.0 can not only have a direct impact on the extraction and / or decantation outcomes, it can also improve the shelf life and processability of the biomass, before and after extraction.

[0035] In view of the surprising discoveries discussed above, the present disclosure provides processes and systems that take advantage of the unexpected benefits of employing acidic and chelating conditions in wet biomass extraction processes. These benefits can include, but are not limited to, an increase in the phase separation rate, especially regarding the aqueousphase (i.e., raffinate) formed from the extraction process; an increase in the separation kinetics (the organic phase (i.e., extract) separates more efficiently than at higher, near neutral or basic pH levels); increased repulsion of the acidic aqueous environment by the biomass (can result in reduced biomass loss into the raffinate and higher biomass recovery); reduced emulsion formation and increased hydrolysis (can improve the extraction efficiency and provide higher carotenoid and oil recovery); increased trapping of metal ions in the raffinate (when an acidic chelating agent is contacted with the wet biomass, the chelating agent traps them and holds them in the raffinate); a decrease in the soluble protein content in the raffinate (a higher amount of precipitated proteins can be recovered, thus improving the overall protein recovery of the extraction process); an increase in biomass shelf life (due to the low pH inhibiting microbial growth); and a decrease in viscosity of the rag layer (due to reduced emulsion at low pH, the biomass fraction formed during decantation has lower viscosity, thus making the material transfer less energy intensive).

[0036] As used herein, “shelf life” refers to the microbiological stability of the feed, products and / or intermediate products produced or used in the processes disclosed herein.

[0037] As used herein “microbiological stability” refers to the degree of control or inhibition of unwanted microbiological growth. Unwanted microbiological organisms can include bacteria and fungi species

[0038] Where wet extraction of a solid is applied, there is no disclosure in the art that mentions, for example, of carrying out the extraction at a low pH, under mild conditions (e.g., T < 100°C) and high salinity to obtain at least an oil and protein rich fraction. Moreover, the chelating of metals to improve the quality of oil extracted from an algal biomass has not been disclosed in the art. Furthermore, the use of low pH to (i) improve the processability of the wet biomass and / or the natural products isolated from the wet biomass and (ii) increase shelf life of the biomass has not been disclosed.

[0039] Accordingly, one aspect of the present disclosure is a process for extracting natural products from a biomass, the process including at least one or more of: forming an acidic feedstock stream having a pH of at most 4.0 and containing a biomass and an aqueous salt solution; contacting the acidicfeedstock stream with an extraction solvent to form a dispersion; separating the dispersion into multiple layers, the layers including: a solvent extract layer containing at least one hydrophobic natural product and at least a portion of the extraction solvent, a raffinate layer containing at least a portion of the aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer.

[0040] As used herein, the term “recovering” includes processes that separate the layers and / or phases disclosed herein into a form wherein the layers and / or phases can be independently processed.

[0041] As used herein, the terms “allowing the dispersion to phase separate” may be understood in this disclosure as “separating the dispersion into multiple layers”.

[0042] In exemplary embodiments, the forming of the acidic feedstock stream includes at least one or more of: obtaining a feedstock stream containing the biomass and the aqueous salt solution, the feedstock stream having a pH of more than 4.0 or at least 5.0; and contacting the feedstock stream with an additive stream containing at least one buffering agent and / or at least one acidic additive selected from, but not limited to, a Bronsted acid, an acidic chelating agent and a combination thereof.

[0043] In exemplary embodiments, the contacting of the feedstock stream with the additive stream occurs in a mixing zone.

[0044] The mixing zone can include at least one or more of a static mixer, an in-line mixer, an eductor, a transpiring wall, a diffuser, a mixing-T piece, a turbulent flow, an agitated vessel, an extraction column, or a combination thereof.

[0045] FIG. 1 shows an exemplary embodiment of the extraction process wherein a feedstock stream is contacted with an additive stream in a mixing zone. In this embodiment, a feedstock stream (100) is transferred into a mixing zone (104). An additive stream (102) is also transferred into the mixing zone (104), where it contacts the feedstock stream (100) to form an acidic feedstock stream (106).

[0046] Additionally, or alternatively, the system of the present disclosure comprises a mixing zone and extraction zone, wherein the mixing zone is in communication with the extraction zone. The terms “the mixing zone is in communication with the extraction zone” may be understood as the that the mixing zone and the extraction zone” are in connection or in fluid connection with each other, via the use of any structural component capable of connecting them to each other (e.g., a tube, a pipe, or a canal).

[0047] Additionally, or alternatively, the mixing zone is separate of the extraction zone. Additionally, or alternatively, the mixing zone precedes the extraction zone.

[0048] The expression “feedstock stream” refers to a stream originating from a biomass culture, the stream containing a biomass solution or suspension that includes natural products (e.g., algal oils, proteins, lipids, carotenoids, hydrophobic products), growth medium (e.g., an aqueous growth medium), and a biomass.

[0049] In exemplary embodiments, the feedstock stream originates from at least one or a combination of plant, algae, micro-organism, bacteria, or microalgae feedstock sources. Suitable algae or microalgae feedstock sources can be derived from algal aquaculture ponds. These algal aquaculture ponds can be operated in batch, continuous, semi-continuous or other modes known in the art to produce the desired biomass in the feedstock stream.

[0050] The algal aquaculture pond can be any type of algal aquaculture pond used to grow algae including, but not limited to, an open pond, a closed pond, a raceway pond, an enclosed or partly enclosed photobioreactor, a tubular reactor, a flat panel (i.e., flat-plate) reactor, a column reactor, an enclosed raceway, a covered pond, an open raceway pond, an earthen pond, a pond in a greenhouse, a fermenter, a naturally occurring body of water, a solar salt pond, or any combinations thereof.

[0051] Open algal aquaculture ponds can be configured either with or without agitation or liners. When present, suitable liner materials include plastic, cement or clay. Plastic algal aquaculture pond liners can be formed from polyethylene, polypropylene, or polyvinyl chloride. Different types of these basic polymers can be used, for example, linear low-density polyethylene liners are used for algaecultivation on a large scale. These liners can also include additives, such as carbon black, to provide resistance to ultraviolet radiation. These liners can also include Nylon or other fibers to provide additional structural integrity. Raven Industries (South Dakota) provides a full line of suitable liners that include one or more layers of materials. Suitable clay liners can include bentonite clay. However, when algal aquaculture ponds are flooded, components in the water, especially saline, can often form a barrier that seals the algal aquaculture pond. It can be desirable to include liners in just a portion of the algal aquaculture pond. For example, liners can be utilized to protect earthen borders where the hydraulic flow can be elevated.

[0052] In exemplary embodiments, additional aqueous medium is periodically or continuously provided to the feedstock source to replace water that evaporates from the feedstock source and / or increase the liquid depth of the feedstock source, i.e., the depth of the algal growth medium. The additional aqueous medium can be combined with recycled growth medium to form an aqueous medium that enters a feedstock source optionally via a pretreatment unit to form an aquaculture medium. If the growth medium in the feedstock source contains mineral salt, the salinity of the additional aqueous medium fed to the feedstock source can be less than the highest salinity experienced by the biomass in the feedstock source.

[0053] The amount of the additional aqueous medium possibly added each day can depend upon several factors: salinity targets in the feedstock source, the wind speed and the relative humidity. Daily evaporation rates can be a major factor in the determination of daily water needs for the feedstock source. If evaporation rates are high, then the salinity of the feedstock source will increase, and water will be required to make up for the losses and to maintain the salinity target for each feedstock source. Wind tends to accelerate evaporation rates, and thus the higher the wind speed, the higher the evaporation rates for a given salinity. The amount of moisture in the atmosphere is termed the relative humidity, and the lower this value the higher the evaporation rates when all other factors are constant. Consequently, the rate and amount of evaporation directly influences the daily demand for additional aqueous medium.

[0054] In exemplary embodiments, the feedstock stream can contain a biomass concentration greater than about 10 ppm, 100 ppm, 0.1wt%, 1wt%, 10wt%, 20wt%, 30wt%, 40wt% or 50wt%.

[0055] As used herein, the term "about" refers to a value that is ± 5% of the stated value. In addition, it is understood that reference to a range of a first value to a second value includes the range of the stated values, e.g., a range of about 1 to about 5 also includes the more precise range of 1 to 5. It is also understood that the ranges disclosed herein include any selected subrange within the stated range, e.g., a subrange of about 50 to about 60 is contemplated in a disclosed range of about 1 to about 100.

[0056] In exemplary embodiments, the feedstock stream has a suitable viscosity so that it can be pumped. A suitable viscosity for pumping can be less than 10,000 cp, less than about 1 ,000 cp and / or less than 500 cp.

[0057] The feedstock stream can undergo various harvesting processes before advancing to the extraction processes disclosed herein. These harvesting processes can include, but are not limited to, fracking, adsorptive bubble separation, filtration, deep bed filtration, belt pressing, screw pressing, centrifugation, adsorption, sedimentation, mechanical floatation, froth flotation, flocculation and combinations thereof. Examples of these and other harvesting processes and equipment which can be used to condition the feedstock stream before being subjected to the extraction processes disclosed herein can be found in U.S. Pat. No. 5,541 ,056; U. S. Pat. No. 4,554,390; U.S. Pat. No.4,115,949; U.S. Pat. No. 5,951 ,875; U.S. Pat. No. 4,680,314; U.S. Pat. No.6,524,486; U.S. Pat. No. 6,405,948; U.S. Pat. No. 5,776,349; U.S. Pat. No.6,000,551 ; U.S. Pat. No. 8,512,998; U.S. Pat. No. 4,397,741 ; U.S. Pat. No.4,938,865; U.S. Pat. No. 5,188,726; U.S. Pat. No. 5,332,100; WO2008 / 156,795; WO 2008 / 156,835; U.S. Pat. No. 4,981 ,582; U.S. Pat. No.5,167,798; all the contents of which are incorporated herein by reference in their entireties.

[0058] In exemplary embodiments, prior to an adsorptive bubble separation process, the biomass in the feedstock stream can be flotation conditioned by a number of processes. Suitable flotation conditioning processes that can be used prior to the adsorptive bubble separation unit include, but are not limited to,adding a flotation aid, adding a frother, adding a collector, adding an activator, adding a depressor, and combinations thereof.

[0059] Collectors selectively render one or more of the species of particles in the feedstock stream hydrophobic, thereby assisting in the process of collection by gas bubbles. Activators aid the adsorption of the collector to certain particles increasing the number of those particles which become hydrophobic.Depressors inhibit the adsorption of the collector to certain undesired particles decreasing the number of those particles which become hydrophobic. Also, frothing agents and frothers can be added to the feedstock stream to assist in the formation of a stable froth on the surface of a liquid.

[0060] Sedimentation processes can include the addition of alum to and / or lack of agitation of the feedstock stream. The addition of ferric chloride can also be included in sedimentation processes to cause flocculation. Any polymer or ions that cause flocculation can also be used during sedimentation processes. Cyclones can also be used to accelerate the rate of sedimentation. Any sedimentation equipment known in the art can be used to separate the flocculated natural products from an aqueous salt solution prior to an adsorptive bubble separation process.

[0061] Adsorption can be used as a conditioning process to reduce the volumetric flow of the feedstock stream to an adsorptive bubble separation unit. Some feedstock, for example Dunaliella salina, can be concentrated by adsorbing the algae onto a hydrophobic surface, and then desorbing the algae with another fluid. Thus, adsorption can be used to preconcentrate the feedstock stream.

[0062] Deep bed filtration can be used to further concentrate the feedstock stream in addition to an adsorptive bubble separation process. Deep bed filtration relies upon a bed of granular media, such as sand, through which the feedstock stream containing natural products flows downward under gravity. The natural products are deposited in the pores of the granular media and in the interstitial spaces between the grains of media. Deep bed filtration should not be confused with straining filtration. Straining takes place on the surface of a mesh or fabric, and is only suitable to further concentrate the feedstock stream with natural products that will not blind the filtration equipment.

[0063] Adsorptive bubble processes can include a step of rendering material or natural products within the feedstock stream hydrophobic by treating particle surfaces with chemicals, or other techniques that selectively modify the material or natural products to be separated. In some cases, the particles or natural products are not initially hydrophobic, and need to be rendered hydrophobic to be separated or harvested from the feedstock stream.

[0064] A flocculating agent can be utilized during adsorptive bubble separation processes to cause accumulations of biomass or natural products to float out during adsorptive bubble processes.

[0065] The feedstock stream or the biomass included in the feedstock stream can also be subjected to a cell rupturing process before proceeding to the extraction process. The feedstock stream can include cellular material which contains natural products. In these instances, rupturing the cell wall and / or cell membrane of the cellular material can release natural products that can be purified in later processes. Cell rupturing can be achieved by a number of methods which include, but are not limited to, chemical, physical or mechanical methods. Chemical methods can include enzymatic digestion, detergent solubilization, lipid dissolution with a solvent, and alkali treatment (lipid saponification). Physical methods can include osmotic shock, decompression, sonication, heat treatment, and freeze-thawing. Mechanical methods can include grinding, high shear homogenization, passing the algal concentrate across a pressure drop, and pressure extrusion.

[0066] Other cell disruption processes which can be used include pumping the feedstock stream at high pressures through a restricted orifice valve. An equipment which can perform this disruption method is, as an example, the MICROFLUIDIZER™ cell disruption equipment of Microfluidics, Newton, MA, US, which utilizes pressures of about 5,000 to 40,000 psig (345 - 2760 bar).

[0067] A mill, such as a vibratory mill, can also be used to rupture cellular material in the feedstock stream.

[0068] In exemplary embodiments wherein the feedstock stream contains algae or microalgae, fracking processes can be performed on the feedstock stream before the extraction process. The partial rupturing of algae is referred to as fracking. Fracked algae can be advantageous over completely rupturedalgae due to the difference in size of the resulting particles. Particles resulting from fracking algae are larger than the particles resulting from the complete rupturing of algae and thus adsorptive bubble separation processes can be more effective when larger particles are present. Fracking the algae or microalgae can produce fracked cells possessing hydrophobic components while still retaining a significant portion of the intracellular material within the cellular membrane. This can result in increased recovery of the intracellular material. Fracking can take place in any device known in the art in which algae or microalgae can be partially ruptured including, but not limited to, a vibratory mill, a French press, a pump, an agitated vessel, or combinations thereof.

[0069] As used herein, the term “algae” refers to unicellular and multicellular eukaryotic algae, microalgae, diatoms, dinoflagellates, coccolithophores, cyanobacteria and combinations thereof.

[0070] The algae or microalgae which can be present in the feedstock stream and / or the biomass can include, but is not limited to, algae from the divisions of Bacillariophyta, Chlorophycophyta, Chrysophycophyta, Cyanophycophyta, Cryptophycophyta, Phaeophycophyta, Pyrrhophycophyta, Rhodophycophyta and combinations thereof. The algae or microalgae present in the feedstock stream and / or biomass can include, but is not limited to, species from the following genera: Acutodesmus, Achnahtes, Amphipora, Amphora, Anabaena, Ankistrodesmus, Arthrospira (also known as Spirulina), Asteromonas, Asterionella, Boekelovia, Borodinella, Botryococcus, Bracteacoccus, Carteria, Chaetoceros, Chlamydomonas, Chlorella, Chlorococcum, Chlorogonium, Chloromonas, Chroomonas, Chrysophaera, Ceratium, Closterium, Coccolithus, Coelastrella, Coscinodiscus, Cosmarium, Cricosphaera, Crocosphaera, Crypthecodinium, Cryptomonas, Cyanocystis, Cyanospira, Cyclotella, Desmodesmus, Ditylum, Dunaliella, Ellipsoidon, Emiliania, Eremosphaera, Euglena, Fragilaria, Franceia, Galdieria, Gracilaria, Graesiella, Guinardia, Haematococcus, Halocafeteria, Halospirulina, Hantzschia, Hymenomonas, Isochrysis, Lepocinclis, Limnothrix, Micractinium, Microactinium, Microcystis, Monochrysis, Monodus, Monoraphidium, Muriellopsis, Nannochloris, Nannochloropsis, Navicula, Neochloris, Neospongiococcum, Nephrochloris, Nephroselmis, Nitzschia, Nodularia, Nostoc, Ochromonas, Oedogonium,Oocystis, Oscillatoria, Ostreococcus, Parachlorella, Pavlova, Peridinium, Phaeodactylum, Picochlorum, Platymonas, Pleurochrysis, Pleurococcus, Porphyra, Porphyridium, Prochlorococcus, Prototheca, Prymnesium, Pseudanabaena, Pseudochlorella, Pseudochoricystis, Pseudoneochloris, Pyramimonas, Pyrobotrys, Rhodomonas, Scenedesmus, Schizochytrium, Scytonema, Skeletonema, Spirogyra, Stichococcus, Synechococcus, Tetrachlorella, Tetradesmus, Tetraselmis, Thalassiosira, Tisochrysis, Tolypothrix, Tribonema, Trichodesmium, Ulothrix, Vaucheria, Viridiella, Volvox, and genetically-engineered varieties or combinations (mixtures, mixed cultures, co-cultures or synthetic co-cultures) thereof. In exemplary embodiments, the algae or microalgae is selected from the group includingDunaliella sp., Dunaliella bardawil, Dunaliella salina, Dunaliella kone, Dunaliella tertiolecta, Dunaliella parva and Dunaliella viridis, and any combination thereof. In exemplary embodiments, the algae or microalgae is Dunaliella salina.

[0071] The algae or microalgae which can be present in the feedstock stream and / or biomass can also include any microalgal species (including diatoms, coccolithophorids and dinoflagellates) selected from, but not limited to, Amphora sp., Ankistrodesmus sp., Arthrospira (Spirulina) plantesis, Botryococcus braunii, Chlamydomonas sp., Chlamydomonas reinhardtii, Chlorella protothecoides, Chlorella sp., Closterium sp., Cosmarium sp., Crypthecoddinium cohnii, Cyclotella sp., Dunaliella salina, Dunaliella tertiolecta, Haematococcus pluvialis, Hantzschia sp., Nannochloris sp., Nannochloropsis sp., Navicula sp., Neochloris oleoabundans, Nitzschia sp., Phaeodactylum tricornutum, Scenedesmus sp., Schiochytrium limacinum, Stichococcus sp., Tetraselmis suecica, and Thalassiosira pseudonana, and genetically- engineered varieties or combinations (mixtures, or mixed cultures) of these microalgal species.

[0072] The algae or microalgae which can be present in the feedstock stream and / or the biomass can also include algae with flagella, cilia and / or eyespots. Flagella are a tail-like projection that protrudes from the cell body of certain algae and functions in locomotion. Cilia are an adaptation that allows independent cellular creatures, like algae, to move around in search of food. Photosensitive eyespots are found in some free-swimming unicellular algae.Photosensitive eyespots are sensitive to light. They enable the algae to move in relation to a light source. Such algae have the capability of independent motion, phototaxis, and can move towards the surface during daylight. Phototaxis is the movement of microalgae in response to light. Certain algae (e.g., Dunaliella) can perceive light by means of a sensitive eyespot and move to regions of higher light concentration to enhance photosynthesis.

[0073] The algae or microalgae which can be present in the feedstock stream and / or biomass also include marine algae that thrive at salt concentrations above that found in seawater. Suitable marine algae can be selected from, but are not limited to, Amphora sp. (diatom), Arthrospira sp., Arthrospira (Spirulina) obliquus, Arthrospira (Spirulina) platensis, Chlorella sp., Chlorella fusca, Chlorella protothecoides, Chlorella pyrenoidosa, Chlorella stigmataphora, Chlorella vulgaris, Chlorella zofingiensis, Dunaliella sp., Dunaliella bardawil, Dunaliella salina, Dunaliella tertiolecta, Dunaliella viridis, Isochrysis galbana, Microcystis sp., Nannochloropsis sp., Nannochloropsis salina, Navicula sp. (diatom), Navicula saprophila (diatom), Nitzschia laevis (diatom), Nitzschia alba (diatom), Nitzschia communis (diatom), Nitzschia paleacea (diatom), Nitzschia closterium (diatom), Nitzschia palea, (diatom), and genetically-engineered varieties or combinations (mixtures, or mixed cultures) of these algal species.

[0074] In exemplary embodiments, the algae is microalgae. In other exemplary embodiments, the algae or microalgae have not been genetically modified or do not originate from genetically engineered algae or microalgae.

[0075] In exemplary embodiments, the algae or microalgae which can be present in the feedstock stream and / or biomass include a group of algae or microalgae that has not been genetically modified or does not originate from genetically engineered algae or microalgae. This group of algae or microalgae can include, but is not limited to, Dunaliella sp., Dunaliella bardawil, Dunaliella kone, Dunaliella salina, Dunaliella bioculata, Dunaliella granulata, Dunaliella maritima, Dunaliella minuta, Dunaliella parva, Dunaliella peircei, Dunaliella polymorpha, Dunaliella primolecta, Dunaliella pseudosalina, Dunaliella quartolecta, Dunaliella terricola, Dunaliella tertiolecta, and Dunaliella viridis.

[0076] All of the possible algae and microalgae which can be included in the feedstock stream can also be included within any biomass present in the feedstock stream.

[0077] The expression “natural products” refers to products which are naturally produced or found within an environment or a living organism. Natural products can include those which are hydrophobic, hydrophilic or amphipathic.

[0078] In exemplary embodiments, the natural products are those which are naturally produced by a plant, a microbe, an algae or microalgae species which can be included within the feedstock stream or the biomass. These natural products can include one or more of lipids, algal lipids, carotenoids, fatty acids, algal fatty acids, triacylglycerols, diacylglycerols, monoacylglycerols, oils, algal oils, chlorophyll, glycerol, phospholipids, carbohydrates, fibers, and proteins.

[0079] In exemplary embodiments, the feedstock stream contains an aqueous salt solution.

[0080] The expression “aqueous salt solution” refers to a solution containing water and at least one salt. The salt can be any one or combination of salts found in sea water, terminal lakes, or aquifers. In exemplary embodiments the aqueous salt solution is or includes culture medium from the feedstock stream’s source.

[0081] The aqueous salt solution can include combinations of ions found in seawater and / or the biomass.

[0082] The aqueous salt solution can contain concentrations of salts which range from trace amounts to saturating amounts. Suitable terms to describe the salinity or salt concentration of the aqueous salt solution range from fresh water, brackish water, salt water, brine, and saturated brine, respectively, as the salt concentration in the aqueous salt solution increase. The desired concentration of salt in the aqueous salt solution will depend on the type of feedstock (e.g., algae species) present in the feedstock stream.

[0083] The expression “salinity” refers to the total amount of dissolved salts in the aqueous solution. Salts which can be dissolved and found in the aqueous solution include, but are not limited to, those found in natural waters such as sodium chloride, magnesium chloride, calcium and magnesium sulfates, bicarbonates, and carbonates. It is a standard practice to express salinity asparts per thousand (%o), which is not a true percent but an approximation of the milligrams of salt per gram of water. In more general terms, salinity is indicated by the water source, such as a freshwater, a brackish water, a saline water, and a brine. Ranges of salinity are associated with these general terms and these ranges are defined as < 0.05 wt% for freshwater, 0.05 - 3 wt% for brackish water, 3 - 5 wt% for saline water, and > 5 wt% for a brine.

[0084] Various combinations of ions found in seawater can be included in the aqueous salt solution. Suitable ion combinations can be derived from one or more of the following sources including: water derived from streams, lakes, rivers, or other sources associated with fresh water; water derived from underground aquifers that can include various ion concentrations; water derived from industrial, agricultural, or municipal sources that can or cannot have received treatment; or water derived from brackish sources where fresh water is combined with sea water or ocean water in various proportions; sea water or ocean water that can be derived from the various seas and oceans located around the globe; water derived from terminal lakes; or combinations thereof. The combination of ions for the aqueous salt solution can be derived directly from these sources, or can be derived by evaporating the desired amount of water from any of these sources to leave the desired ion-rich solution for use as the aqueous salt solution. An example of an ion combination source is disclosed in U.S. Pat. No. 6,986,323, the contents of which are included herein by reference. Other examples include the evaporation of ancient sea waters that form terminal lakes, such as the Great Salt Lake in Utah, and that form various aquifers. The combination of ions can result up to and include crystallizers wherein sodium chloride ions are precipitated.

[0085] The aqueous salt solution and / or the feedstock stream can have a salinity that is lower than 5 wt%, about 5 wt%, greater than 5 wt%, about 6 wt% or greater than 6 wt%, about 7 wt% or greater than 7 wt%, for example at least about 8 wt%, at least about 9 wt%, at least about 10 wt%, at least about 11 wt%, at least about 12 wt%, at least about 13 wt%, at least about 14 wt%, at least about 15 wt%, at least about 16 wt%, at least about 17 wt%, at least about 18 wt%, at least about 19 wt%, at least about 20 wt%, at least about 21 wt%, at least about 22 wt%, at least about 23 wt%, at least about 24 wt%, or at leastabout 25 wt%. In exemplary embodiments, the aqueous salt solution and / or the feedstock stream is saturated with salt. In other exemplary embodiments, the aqueous salt solution and / or the feedstock stream can have a salinity that is about 5 wt% to about saturation, from about 10 wt% to saturation, from about 20 wt% to saturation, from about 5 wt% to about 20 wt%, from about 10 wt% to about 20 wt%, from about 5 wt% to about 15 wt%, from about 10 wt% to about 15 wt%, or from about 5 wt% to about 10 wt%. In exemplary embodiments, the aqueous salt solution and / or the feedstock stream has a salinity greater than 5 wt%, greater than 10 wt%, greater than 15 wt% or greater than 20 wt%.

[0086] As used herein, wt% refers to a dry mass of a component in a solution in grams divided by 100 grams of the solution.

[0087] At least one salt can either be present in the aqueous salt solution or added to the feedstock stream to increase its salinity. The presence of salt, and specifically elevated salt compositions provides several advantages. First, the presence of salt in the feedstock stream can reduce solvent solubility in a raffinate layer, and thus reduce either potential solvent loss or solvent recovery costs. Second, the presence of salt in the feedstock stream can increase the density of a raffinate layer, thus enhancing phase separation rates that lead to reduced decanter sizes. Third, the presence of salt in the aqueous salt solution and / or the feedstock stream can retard spoilage of the biomass in the processing step. All of these advantages, alone or in combination, adds significant value to the processes disclosed herein.

[0088] The at least one salt can be selected from, but not limited to, a sea salt, an underground salt, a salt of aquifer water, a salt of a terminal lake, sodium chloride, and / or any combination thereof.

[0089] The biomass in the aqueous salt solution and / or the feedstock stream can include or be a plant biomass, a microbial biomass, an algal biomass or any combination thereof.

[0090] All of the possible plants and / or microbes which can be included in the feedstock stream can also be included within the biomass in the aqueous salt solution.

[0091] All of the possible plant or microbial biomass which can be included in the feedstock stream can include any plant or microbial biomass.

[0092] The biomass can also include or contain some or all of the natural products within the feedstock stream.

[0093] The biomass content in the feedstock stream can be as low as about 0.05 wt%, greater than about 0.5 wt% and even greater than 1 wt%. The maximum biomass content in the feedstock stream is limited by the maximum amount of biomass that allows the feedstock stream to flow, and this can be less than about 20 wt%, or less than about 10 wt%.

[0094] The feedstock stream can contain a water content before being subjected to the extraction processes disclosed herein. The water content can be about 0.1 wt% to about 5 wt%, about 5 wt% to about 10 wt%, about 10 wt% to about 15 wt%, about 15 wt% to about 20 wt%, about 20 wt% to about 30 wt%, about 30 wt% to about 40 wt%, about 40 wt% to about 50 wt%, or about any range within 0.1 wt% to 50 wt% of the total weight of the feedstock stream. In exemplary embodiments, the feedstock stream contains a water content before the extraction process that is greater than 50 wt% or about any range within 50 wt% to 99 wt%.

[0095] The biomass can include or be a conditioned biomass. As used herein “a conditioned biomass” refers to a biomass that has been treated with one or more conditioning processes before being subjected to the extraction processes disclosed herein. Suitable conditioning processes can include, but are not limited to, fracking, adsorptive bubble separation, filtration, deep bed filtration, belt pressing, screw pressing, centrifugation, adsorption, sedimentation, mechanical floatation, froth flotation, flocculation and combinations thereof.

[0096] The feedstock stream can have a pH above 4.0, of at least 5.0, a pH within the range of 4.1 - 12.0, a pH within the range of 4.5 - 12.0, a pH within the range of 5.0 to 12.0, a pH within the range of 5.0 to 11 .0, a pH within the range of 5.0 to 10.0, a pH within the range of 5.0 to 9.0, a pH within the range of 5.0 to 8.0, a pH within the range of 5.0 to 7.0, a pH within the range of 5.0 to 6.0, a pH within the range of 6.0 to 12.0, a pH within the range of 6.0 to 11 .0, a pH within the range of 6.0 to 10.0, a pH within the range of 6.0 to 9.0, a pH within the range of 6.0 to 8.0, a pH within the range of 6.0 to 7.0, a pH within the range of 7.0 to 12.0, a pH within the range of 7.0 to 11 .0, a pH within the range of 7.0 to 10.0, a pH within the range of 7.0 to 9.0, a pH within the range of 7.0 to8.0, a pH within the range of 8.0 to 12.0, a pH within the range of 8.0 to 11 .0, a pH within the range of 8.0 to 10.0, a pH within the range of 8.0 to 9.0 or any pH above 9.0.

[0097] The expression “additive stream” refers to a stream containing at least one acidic additive and / or at least one additive such as a buffering agent.

[0098] In exemplary embodiments, the at least one acidic additive in the additive stream is a Bronsted acid selected from, but not limited to, hydrochloric acid, phosphoric acid, nitric acid, sulfuric acid, formic acid, propionic acid, glyoxylic acid, acetic acid and mono- and polyprotic acids including, but not limited to, citric acid, phosphoric acid, glycolic acid, 2-ethylhexanoic acid, malic acid, malonic acid, 2,3-dihydroxybenzoic acid, gluconic acid, tartaric acid, carnosic acid, ethylenediaminetetraacetic acid, 2-ethylhexanoic acid and oxalic acid.

[0099] In exemplary embodiments, the at least one acidic additive in the additive stream is an acidic chelating agent selected from mono- and polyprotic acids including, but not limited to, citric acid, phosphoric acid, glycolic acid, 2- ethylhexanoic acid, malic acid, malonic acid, 2,3-dihydroxybenzoic acid, gluconic acid, tartaric acid, carnosic acid, ethylenediaminetetraacetic acid, 2- ethylhexanoic acid, and oxalic acid.

[0100] As used herein, the term “acidic chelating agent” means acidic compounds that are capable of binding to metal ions.

[0101] The exemplary listed acidic chelating agents can, in instances, act as a Bronsted acid. The conditions where this function occurs can be determined by those skilled in the art. Those skilled in the art will also appreciate that there are instances where one of the exemplary listed Bronsted acids act as an acidic chelating. The conditions where this function occurs can also be determined by those skilled in the art. Those skilled in the art will also appreciate that not all of the exemplary listed Bronsted acids can act as an acidic chelating agent.

[0102] In exemplary embodiments, the at least one additive in the additive stream is a buffering agent selected from, but not limited to, ammonium chloride, ammonium acetate, ammonium phosphate, ammonium sulfate, ammonium citrate, sodium chloride, sodium acetate, sodium phosphate, sodium sulfate, sodium citrate, sodium tartrate, sodium malonate, sodium gluconate,sodium formate, sodium propionate, potassium chloride, potassium acetate, potassium phosphate, potassium sulfate, potassium citrate, potassium tartrate, potassium malonate, potassium gluconate, potassium formate, potassium propionate, or any equivalent magnesium or calcium salts.

[0103] In exemplary embodiments, the additive stream contains at least one Bronsted acid, at least one acidic chelating agent, or at least one buffering agent.

[0104] In exemplary embodiments, the additive stream contains a combination of any one or more Bronsted acid, acidic chelating agent and / or buffering agent (e.g., at least one Bronsted acid and at least one acidic chelating agent; e.g., at least one acidic chelating agent and at least one buffering agent; e.g., at least one Bronsted acid and at least one buffering agent).

[0105] In exemplary embodiments, the volumetric flow of the additive stream is lower than the volumetric flow of the feedstock stream. An advantage of using a lower volumetric flow for the additive stream is that it can minimize the size and costs of the extraction process equipment. The volumetric ratio of the additive stream to feedstock stream can be less than 0.5, less than 0.4, less than 0.3, less than 0.2, or less than 0.1 .

[0106] To determine whether sufficient mixing has occurred between the additive stream and the feedstock stream, the pH of the mixture formed after contact of these two streams can be measured over a sufficient period of time and compared with the pH of the feedstock stream before the contact with the additive stream. pH measurements can be determined by collecting samples of the feedstock stream and the mixture formed after contact and subjecting them to a pH probe (or using in-line measurement). Sufficient mixing has occurred when the pH of the mixture resulting from the contact of the feedstock stream and the additive stream is more acidic (e.g., at most 4.0) than the pH of the feedstock stream and optionally when further mixing is not affecting the pH significantly (e.g., less than 0.1 pH units per minute).

[0107] The expression “acidic feedstock stream” refers to a stream containing a mixture resulting from contact of the feedstock stream and the additive stream.

[0108] The acidic feedstock stream can have an acidic additive concentration in the range of 0.1 to 50 mM (i.e., 0.0001 to 0.05 M).

[0109] The acidic feedstock stream can contain a biomass solution or suspension that includes natural products, a biomass and an acidic aqueous solution.

[0110] The acidic aqueous solution can contain any of the ions and / or salts that the aqueous salt solution can contain. The acidic aqueous solution can also have any salinity that the aqueous salt solution can possess.

[0111] The acidic feedstock stream can have a pH of at most 4.0, a pH within the range of 0 to 4.0, a pH within the range of 0 to 3.5, a pH within the range of 0 to 3.0, a pH within the range of 0 to 2.5, a pH within the range of 0 to 2.0, a pH within the range of 0 to 1 .5, a pH within the range of 0 to 1 .0, a pH within the range of 0 to 0.5, a pH within the range of 1 .0 to 4.0, a pH within the range of 1 .0 to 3.5, a pH within the range of 1 .0 to 3.0, a pH within the range of 1 .0 to 2.5, a pH within the range of 1 .0 to 2.0, a pH within the range of 2.0 to 4.0, a pH within the range of 2.0 to 3.5, a pH within the range of 2.0 to 3.0, a pH within the range of 3.0 to 4.0 or any pH below 4.0.

[0112] The exemplary process in FIG. 1 also depicts the acidic feedstock stream (106) being transferred to an extraction zone (108). In the extraction zone (108), the acidic feedstock stream (106) undergoes an extraction process that involves contacting the acidic feedstock stream (106) with an extraction solvent (110) to form a dispersion (112). The dispersion (112) is then transferred to a separation zone (114). In the separation zone (114), the dispersion (112) separates into three different distinct layers: a solvent extract layer (116), a rag layer (118) and a raffinate layer (120). At least a portion of each of these layers is removed from the separation zone (114) and subjected to further downstream processing.

[0113] In exemplary embodiments, the extraction process conducted on the acidic feedstock stream is a liquid-liquid-solid extraction process including at least one or more of the following: forming a dispersion by contacting (e.g., by intimately contacting) the acidic feedstock stream with an extraction solvent in an extraction zone; passing the dispersion to a separation zone; separating the dispersion into multiple layers, the layers including: a solvent extract layercontaining at least one hydrophobic natural product and an extraction solvent, a raffinate layer containing an aqueous salt solution, and a rag layer containing a lipid-depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer.

[0114] The expression “liquid-liquid-solid extraction” refers to a process wherein an acidic feedstock stream containing a biomass including one or more natural products is (e.g., intimately) contacted with an extraction solvent capable of extracting one or more of the hydrophobic natural products from the biomass.

[0115] The term “dispersion” relates to a heterogeneous mixture containing an aqueous salt solution, a biomass, and an extraction solvent. The dispersion can exist as an emulsion.

[0116] The extraction solvent forms a second liquid phase or layer with the acidic feedstock stream in the extraction zone. Suitable extraction solvents include, but are not limited to a polar solvent, a non-polar solvent, a non-polar organic solvent, a dense gas solvent, an aqueous two-phase solvent, a deep eutectic solvent (DES), a natural deep eutectic solvent (NADES) (such as choline chloride, glucose, lactic acid, malic acid, and / or any combination thereof), an ionic liquid, any combination thereof and / or any combination of miscible solvents.

[0117] In an exemplary embodiment, the extraction solvent is or includes at least one or more of a non-polar solvent, a polar solvent, a non-polar organic solvent, a dense gas solvent, an aqueous two-phase solvent, a deep eutectic solvent (DES), a natural deep eutectic solvent (NADES), an ionic liquid, or any combination thereof.

[0118] In another exemplary embodiment, the extraction solvent is or includes at least one or more of a non-polar solvent, a non-polar organic solvent, a dense gas solvent, an aqueous two-phase solvent, a deep eutectic solvent (DES), a natural deep eutectic solvent (NADES), or any combination thereof.

[0119] In another exemplary embodiment, the extraction solvent is or includes at least one or more of a non-polar solvent, a non-polar organic solvent, a dense gas solvent, an aqueous two-phase solvent, or any combination thereof.

[0120] In another exemplary embodiment, the extraction solvent is or includes at least one or more of a non-polar solvent, a non-polar organic solvent, a dense gas solvent, or any combination thereof.

[0121] The extraction solvent can be chosen such that its polarity is appropriate to extract the desired natural products. Thus, the optimal extraction solvent for the liquid-liquid-solid extraction process can depend on which natural products are desired to be extracted.

[0122] Any solvent system that forms a second immiscible liquid phase or layer with the acidic feedstock stream can be used as the extraction solvent. These solvent systems should not adversely impact the quality or quantity of the natural products. These solvent systems can include, but are not limited to, synthetic and / or natural flavorants, edible oils, petrochemicals, dense gases, and combinations of these so long as the mixture of the solvent system and the acidic feedstock stream forms two immiscible liquid phases at a desired extraction zone temperature. Some of these solvents are more desirable than others for various reasons as discussed below and the results obtained are not necessarily equivalent.

[0123] The solvent system can include petrochemical solvents due to their low viscosity and favorable solute molecular diffusivity. Natural oils are soluble in petrochemical solvents and concentrated extracts are possible. Suitable petrochemical solvents can include those that are disclosed in "Organic Solvents: Physical Properties and Methods of Purification", edited by J. A. Riddick et al, Volume 2, Fourth Edition, ISBN Number 0-471- 08467-0, such as 2-methyl oxolane. In exemplary embodiments, the petrochemical solvents can include, but are not limited to, aliphatic hydrocarbons (such as pentane, hexane, heptane, octane, nonane, decane, dodecane, cyclohexane, petroleum ether, their isomers, and mixtures thereof), aromatic hydrocarbons (including but not limited to benzene, toluene, xylene), alcohols (including, but not limited to butanol, pentanol, hexanol, octanol, dodecanol, cyclohexanol, benzyl alcohol, their isomers, and combinations thereof), ketones (including, but not limited to methyl isobutyl ketone, hexanone, heptanone, octanone, their isomers, and combinations thereof), esters (including, but not limited to methyl acetate, ethyl acetate, propyl acetate, butyl acetate, pentyl acetate, hexyl acetate, ethylpropionate, ethyl butyrate, ethyl valerate, their isomers, and combinations thereof), and / or ethers (including but not limited to 2-methyltetrahydrofuran, diisopropyl ether, methyl tert-butyl ether, cyclopentyl methyl ether, and combinations thereof). Combinations of petrochemical solvents can also be used if desired.

[0124] The petrochemical solvents can also contain one or more co-solvents to improve extractability of solutes. Examples of these co-solvents include methanol, ethanol, 1 -propanol, 2-propanol, 1 -hexanol, 2-methoxy ethanol, acetone, tetrahydrofuran, 1 ,4-dioxane, acetonitrile, dichloromethane, chloroform, dimethyl sulfoxide, formic acid, carbon disulfide, methylene chloride, amines, chelating agents, phase transfer catalysts and combinations thereof. The co-solvents can also be added to the acidic feedstock stream to enhance recovery of a solute or hydrophobic natural product in the extraction solvent.

[0125] The edible oils which can be included within the solvent system can be chosen from those obtained from plant or animal sources, such as fish oils. Edible vegetable oil solvents include, but are not limited to, those derived from corn, olive, algae, soybean, flax, safflower, sunflower, palm, jatropha, coconut, other oils known in the art, and combinations thereof. Compared to petrochemical solvents, edible oils can be more viscous, and the solute molecular diffusivity is lower.

[0126] The solvent system can also include synthetic and natural flavorants. These flavorants can be more desirable than petrochemical solvents and edible oils if the natural products are to be used for human or animal consumption. Naturally derived flavorants have appeal in nutritional supplements. Flavorants classified by the Flavor and Extract Manufacturers Association, or FEMA, as Generally Recognized As Safe, or GRAS, do not have the drawbacks of petrochemical solvents in association with nutritional supplements. The presence of residual flavorant solvents in nutritional supplements is generally acceptable in comparison with petrochemical solvents, which reduces downstream purification and recovery costs. The flavorants can be chosen from those which have boiling points, viscosities, and molecular diffusivity properties comparable to petrochemical solvents. Examples of such flavorants include, but are not limited to, methyl-, ethyl-, propyl-, butyl-, isobutyl-, benzyl-, and octyl-esters with the carboxylic acid component of the ester including acetate, ethanoate, propionate, butyrate, hexanoate, caproate, heptanoate, octanoate, decanoate, cinnamate, and isovalerate. Other examples of flavorants which can be used include, but are not limited to, benzaldehyde, other aldehydes, limonene, and other terpenes. Combinations of flavorants can also be used.

[0127] Suitable dense gases which can be used as the extraction solvent include, but are not limited to, carbon dioxide, ethane, propane, butane, chlorofluorocarbons, and mixtures thereof. The dense gas extraction can be operated in any manner known in the art including leaching, batch extraction, and continuous counter-current extraction as described in U.S. Pat. No.6,106,720 and U.S. Pat. No. 5,932,101 , the contents of which are incorporated herein by reference in their entirety. Additional suitable dense gases can be methane, isobutane, dimethyl ether, sulfur hexafluoride, ammonia, fluorocarbons, and mixtures thereof. Any combination of the above dense gases can also be used.

[0128] The dense gases can also contain one or more co-solvents to improve extractability of solutes. Examples of these co-solvents include methanol, ethanol, 1 -propanol, 2-propanol, 1 -hexanol, 2-methoxy ethanol, acetone, tetrahydrofuran, 1 ,4-dioxane, acetonitrile, dichloromethane, chloroform, dimethyl sulfoxide, formic acid, carbon disulfide, methylene chloride, amines, chelating agents, phase transfer catalysts and combinations thereof. Other examples of dense gases and co-solvents are listed in U.S. Pat. Nos. 4,345,976 and 5,490,884, the contents of which are incorporated herein by reference in their entirety. The co-solvents can also be added to the acidic feedstock stream to enhance recovery of a solute or hydrophobic natural product in the extraction solvent.

[0129] The solvent system can also include an ionic liquid. Suitable ionic liquids include, but are not limited to, solvent systems that are in the liquid phase at the extraction temperature, those that include a cation and an anion, and those that are immiscible with a water-rich algal concentrate phase.

[0130] In exemplary embodiments, the extraction solvent is selected such that the selectivity of the extraction solvent for specific non-polar compounds versus specific salts is greater than unity (i.e., greater than 1). The selectivity iscalculated by dividing the distribution coefficient for a specific non-polar compound of interest to the distribution coefficient for a specific salt of interest. The distribution coefficients are defined as the concentration of the nonpolar compound in the extract phase or solvent extract layer divided by the concentration of the nonpolar compound in the raffinate phase or raffinate layer.

[0131] In exemplary embodiments, forming the dispersion by contacting the acidic feedstock stream with the extraction solvent involves using an acidic feedstock stream to extraction solvent ratio (e.g., a volumetric ratio of the acidic feedstock stream to extraction solvent) of from about 20 to about 0.05, 8 to about 0.1 , from about 5 to about 0.2, or 1 (e.g., a 1 to 1 volumetric ratio).

[0132] The acidic feedstock stream can contact the extraction solvent for about 1 minute to 5 hours, or for about 2 minutes to 5 hours, and the contact time can differ based on the type of contactor used in the extraction zone. When a centrifugal extractor is used for the extraction zone, the contact time can range from about 0.5 to 10 minutes, or less than 2 minutes. When an agitated vessel is used for the extraction zone, the contact time can range from about 1 minute to 5 hours, or between 2 and 120 minutes. The contact time can be 5 to 60 minutes. The acidic feedstock stream can contact the extraction solvent for about 2 to 180 minutes, about 5 to 180 minutes or about 10 to 60 minutes in a counter-current extraction column. In exemplary embodiments, the dispersion is retained in a counter-current extraction column for a residence time of about 2 minutes to about 2 hours.

[0133] The extraction zone can include a mixer, a static mixer, a settler, a cocurrent extraction column, a counter-current extraction column, an extractor (e.g., a centrifugal extractor), an emulsion phase contactor, or any combination thereof known in the art.

[0134] Suitable mixers for the extraction zone include agitated vessels where a mechanical agitator is used to intimately contact the acidic feedstock stream and the extraction solvent. The mechanical agitator can include one or more impellers on a rotating shaft. Suitable impellers include, but are not limited to Rushton Turbines, flat-blade turbines, pitch-blade turbines, marine propellers, hydrofoils, impellers that are sold by Chemineer (Dayton Ohio), or SPX / Lightnin (Rochester, New York). Regardless of the type of impeller used, the degree ofagitation required is important for efficient mass transfer of the solute. The degree of agitation required can be calculated by the minimum impeller speed to completely disperse one immiscible liquid in another, as defined by Skelland and Ramsay [1987 l&EC Res. 26, 1 , 77-81], Skelland and Moeti [1989, l&EC Res. 28, 1 , 122-127] and Skelland and Kanel [1993, l&EC Res. 29, 7, 1300- 1306]. Static mixers of any design can also be used as the extraction zone. Suitable static mixers include, but are not limited to, those produced by Chemineer in their Kenics line.

[0135] The extraction zone can be followed by a separation zone, and mass transfer can continue to occur while the dispersion is separating in the separation zone. In an exemplary embodiment, there is a balance between the amount of shear energy added in the extraction zone to generate an acceptable drop size distribution in the liquid-liquid dispersion in order to achieve acceptable mass transfer kinetics in the extraction zone, and reasonable decantation kinetics in the separation zone. Since the separation zone can be larger in volume than the extraction zone, it is advantageous not to add too much shear that generates a small drop size distribution that would increase the size of the decanter.

[0136] Suitable extraction columns which can be used as the extraction zone include, but are not limited to, those that are mechanically agitated and those that have stationary internals. The latter is preferred when the extraction solvent is a dense gas and / or the operating pressure of the extractor is elevated so that more expensive mechanical seals are needed. Suitable extraction columns with stationary internals can include, but are not limited to, packed, perforated plate, baffle tray, and combinations thereof. Suitable packings include structured or random packings that are known to those skilled in the art. Suitable mechanically agitated extraction columns can include, but are not limited to, the Karr reciprocating plate column, the York Scheibel column, and the rotating disc column, all made by Koch Modular Process Technology Corporation, which is located in Paramus, N.J., the Kuhni column, which is sold by Sulzer in Switzerland, the asymmetric rotating disc column, pulsed columns, and combinations thereof.

[0137] The separation zone can include or be a decanter which is configured to perform at least one or more of gravity settling, centrifugal settling, and / or combinations thereof to separate the dispersion into the multiple layers. In exemplary embodiments, the separation zone can include one or more fixed or moving separation aids like mesh pad coalescers, wire pad coalescers, structured packing, inclined plates, perforated plates, baffles, ultrasonic waves, acoustic waves, and / or combinations thereof.

[0138] In exemplary embodiments, the separation zone can include at least one or more of a decanter, a tricanter, a coalescer, a centrifuge or an electric field coalescer.

[0139] In an exemplary embodiment, the extraction zone includes a mixer and / or a counter-current extraction column, and / or the separation zone includes a decanter.

[0140] In exemplary embodiments, the extraction zone and separation zone are combined in a counter-current extraction column. In these exemplary embodiments, the raffinate layer can exit the column at one end while the solvent extract layer can exit the column at the opposite end. The rag layer can be removed from the counter-current extraction column with either the raffinate layer or the solvent extract layer, or alternatively removed from the extraction column as a sidedraw. Any of the types of extraction columns described above can be used for counter-current extraction. The algal concentrate can contact the extraction solvent for about 5 to 180 minutes, about 5 to 180 minutes or about 10 to 60 minutes in a counter-current extraction column.

[0141] Suitable centrifugal extractors that can be used to provide both the extraction zone and the separation zone include, but are not limited to those produced by CINC, Alfa Lavel, Podbielniak, Robatel, Westfalia, and combinations of these centrifugal extractors.

[0142] Suitable emulsion phase contactors that can be used to provide both the extraction zone and the separation zone include, but are not limited to, those produced by Schlumberger termed the NATCO dual frequency electrostatic treater.

[0143] The separation of the dispersion into multiple layers can be carried out or performed under a gravitational field or by decanting.

[0144] Separating the dispersion into multiple layers can occur in as little as 5 minutes to about 24 hours, at least 20 minutes to 12 hours, at least 30 minutes to 6 hours, or 40 minutes to 3 hours.

[0145] Separating the dispersion into multiple layers can occur or can be performed at a pressure ranging from atmospheric to supercritical conditions for the extraction solvent.

[0146] Separating the dispersion into multiple layers can occur or can be performed at a temperature of about 100°C or less than about 100°C, about 10°C to about 100°C, about 10°C to about 90°C, about 20°C to about 90°C, about 30°C to about 90°C, about 40°C to about 90°C, about 50°C to about 90°C, about 60°C to about 90°C, about 10°C to about 80°C, about 20°C to about 80°C, about 30°C to about 80°C, about 35°C to about 80°C, about 40°C to about 80°C, about 50°C to about 80°C, about 60°C to about 80°C, about 10°C to about 70°C, about 20°C to about 70°C, about 30°C to about 70°C, about 35°C to about 70°C or about 40°C to about 70°C.

[0147] The solvent extract layer can include at least one hydrophobic natural product present within the dispersion. These hydrophobic natural products can include one or more selected from the group including lipids, algal lipids, carotenoids, fatty acids, algal fatty acids, triacylglycerols, diacylglycerols, monoacylglycerols, oils, algal oils and combinations thereof.

[0148] The carotenoids can include beta-carotene, alpha-carotene, lutein, zeaxanthin, beta-cryptoxanthin, astaxanthin, phytoene, phytofluene, lycopene, and / or combinations thereof.

[0149] The solvent extract layer can include extraction solvent, algal oil, carotenoids, trace amounts of water and salt. The solvent extract layer can contain extraction solvent in amounts of more than 50 wt%, such as above 60 wt% or above 70 wt% of its total weight; algal oil in amounts than less 30 wt%, such as less than 20 wt% or less than 10 wt% of its total weight; carotenoids in amounts less than 5 wt%, such as less than 3 wt% or less than 1 wt% of its total weight; water in amounts less than 10 wt%, such as less than 5 wt% or less than 2 wt% of its total weight; and / or salt in amounts less than 3 wt%, such as less than 2 wt% or less than 1 wt% of its total weight.

[0150] The solvent extract layer can include limited amounts of lipid-depleted biomass and an aqueous salt solution.

[0151] The raffinate layer can include an aqueous salt solution depleted of hydrophobic natural products. The raffinate layer can possess a salt concentration of above 5 wt%, above 7 wt%, above 10 wt% above 15 wt%, above 18 wt% up to saturation. In an exemplary embodiment, the raffinate layer has a salinity of about 5 wt% or greater, about 10 wt% or greater, about 15 wt% or greater, or about 20 wt% or greater. In exemplary embodiments a salinity of the raffinate layer is about 5 wt% or greater than 5 wt%, about 6 wt% or greater than 6 wt%, about 7 wt% or greater than 7 wt%, for example at least about 8 wt%, at least about 9 wt%, at least about 10 wt%, at least about 11 wt%, at least about 12 wt%, at least about 13 wt%, at least about 14 wt%, at least about 15 wt%, at least about 16 wt%, at least about 17 wt%, at least about 18 wt%, at least about 19 wt%, at least about 20 wt%, at least about 21 wt%, at least about 22 wt%, at least about 23 wt%, at least about 24 wt%, or at least about 25 wt%. In other exemplary embodiments the raffinate layer is saturated with salt. In more exemplary embodiments, the raffinate layer can have a salinity that is about 5 wt% to about saturation, from about 10 wt% to saturation, from about 20 wt% to saturation, from about 5 wt% to about 20 wt%, from about 10 wt% to about 20 wt%, from about 5 wt% to about 15 wt%, from about 10 wt% to about 15 wt%, or from about 5 wt% to about 10 wt%.

[0152] The raffinate layer can contain algal growth medium that is depleted in algal biomass and / or algal oils.

[0153] The raffinate layer can include water, salts, trace amounts of extraction solvent, trace amounts of algal biomass (such as less than 5 wt% or less than 1 wt% of its total weight) and / or trace amounts of algal oil (such as less than 5 wt% or less than 1 wt% of its total weight). The raffinate layer can contain water above 50 wt% (such as from 60 wt% to 90 wt%, 70 wt% to 85 wt% or 80 wt% to 90 wt%) or at about 50 wt% of its total weight, and / or the raffinate layer can contain salt from about 0.1 wt% to about 50 wt%, about 5 wt% to about 40 wt%, about 10 wt% to 30 wt%, and / or about 15 wt% to 26 wt% of its total weight. The raffinate layer can also contain trace amounts of the extraction solvent in both soluble and / or insoluble forms. The solubility of the extraction solvent in theraffinate layer influences the amount, or concentration, of the extraction solvent in the raffinate layer. The soluble amount of extraction solvent can be experimentally determined by measuring the solubility of the extraction solvent in the raffinate layer as a function of temperature, pressure, pH, salinity, and / or other factors known to those skilled in the art. The insoluble amount of the extraction solvent in the raffinate layer can be determined by the amount of entrainment of both extraction solvent droplets and the amount of extraction solvent that is associated with entrained algal biomass in the raffinate layer. Trace amounts of algal oil in the raffinate layer can be present in the solvent of the raffinate layer and / or in the biomass as unextracted oil.

[0154] The rag layer can form at any location between, above or below the solvent extract and raffinate layers. In one embodiment, the rag layer can be located between the solvent extract and raffinate layers. The rag layer can include a lipid-depleted biomass. The lipid-depleted biomass can include at least one or more of, chlorophyll, glycerol, phospholipids, proteins, carbohydrates, fibers, and limited amounts of lipids, carotenoids, raffinate relative to the dispersion and / or any combination thereof. In exemplary embodiments, the rag layer contains at most about 45 wt% solvent extract layer, about 45 wt% raffinate layer and about 10 wt% lipid-depleted biomass. In an exemplary embodiment, the rag layer contains at most about 60 wt% solvent extract layer (such as from 30 wt% to 60 wt%, e.g., 40 wt% to 50 wt% or about 45 wt%), at most about 60 wt% raffinate layer (such as from 30 wt% to 60 wt%, e.g., 40 wt% to 50 wt% or about 45 wt%) and / or about from 2 wt% to 20 wt% lipid-depleted biomass (e.g., from 5 wt% to 10 wt%).

[0155] The rag layer can contain a majority of the biomass. Reducing the volume of the rag layer can be desirable to minimize the cost of further processing the rag layer to recover any entrained biomass.

[0156] The rag layer can include water, salt, extraction solvent, algae oil, and / or biomass. In exemplary embodiments, the rag layer is a mixture of three layers: 1 ) a layer containing biomass that has been depleted of non-polar compounds, 2) at least a portion of the solvent extract layer, and 3) at least a portion of the raffinate layer. The ratio of these three layers can vary depending on the conditions used to separate the dispersion into multiple layers. The layercontaining the biomass that has been depleted of non-polar compounds can vary from about 0.01 wt% of the total weight of the rag layer to about 20 wt%. The solvent extract layer and the raffinate layer can vary from about 1 wt% to about 99 wt% of the total weight of the rag layer. In exemplary embodiments, the layer containing biomass that has been depleted of non-polar compounds is between about 1 and 10 wt% of the total weight of the rag layer, and the solvent extract layer and the raffinate layer range from about 10 wt% to 90 wt% of the total weight of the rag layer.

[0157] In exemplary embodiments, the rag layer includes a biomass having a particle distribution size from about 0.1 microns to about 1000 microns, about 0.2 microns to about 100 microns, from about 0.4 microns to about 25 microns, from about 0.4 microns to about 20 microns or at least 20 microns.

[0158] The rag layer can have an intermediate density between the solvent extract layer and the raffinate layer, so in decantation and / or centrifugation processes, it is located between the solvent extract layer and the raffinate layer.

[0159] In exemplary embodiments, the rag layer does not exist as a true thermodynamic phase and exists as a mixture of solid biomass, raffinate layer and solvent extract layer. The raffinate layer and the solvent extract layer can exist as true thermodynamic phases.

[0160] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream includes performing a heat exchange before the acidic feedstock stream enters the extraction zone, before the extraction solvent enters the extraction zone, and / or before the dispersion enters the separation zone.

[0161] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream includes recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer. The solvent extract layer can be e.g., overflowed or pumped out of the separation zone, the rag layer can be pumped out of the separation zone and / or the raffinate layer can be removed e.g., from the bottom of the separation zone.

[0162] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream includes contacting the isolated solvent extract layer with an aqueous phase to remove any residual salt concentrations that can be present within the solvent extract layer.

[0163] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream includes using a coalescer after formation of the dispersion.

[0164] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream includes filtering the solvent extract layer after isolation to remove any entrained biomass and / or filtering the raffinate layer after isolation to remove any entrained biomass.

[0165] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream includes evaporating the extraction solvent from the solvent extract layer after isolation of the solvent extract layer.

[0166] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream is performed at a temperature of about 100°C or less, about 90°C (e.g., 90 ± 1-10°C) or less, about 80°C or less, about 70°C or less, about 60°C or less, or e.g., within a temperature range from 10°C to 90°C, from 20°C to 100°C, from 20°C to 90°C, from 25°C to 90°C, from 30°C to 90°C, from 40°C to 90°C, from 50°C to 90°C, from 55°C to 90°C, from 60°C to 90°C, from 10°C to 80°C, from 20°C to 80°C, from 30°C to 80°C, from 40°C to 80°C, from 50°C to 80°C, from 60°C to 80°C, about 10°C to about 100°C, about 10°C to about 70°C, about 20°C to about 70°C, about 30°C to about 70°C, about 35°C to about 70°C or about 40°C to about 70°C. It can be advantageous to operate the extraction at a temperature below 100°C to preserve the algal oils and carotenoids.

[0167] In exemplary embodiments, forming the dispersion by contacting the acidic feedstock stream with the extraction solvent in the extraction zone and separating the dispersion into multiple layers in the separation zone can be performed at the same or different temperatures.

[0168] In exemplary embodiments, the separating of the dispersion into multiple layers occurs or is performed at a temperature of about 100°C or less, of about 95°C or less, about 90°C or less, about 85°C or less, about 80°C orless, about 75°C or less, about 70°C or less, about 65°C or less, about 60°C or less, or within a temperature range from 10°C to 90°C, from 20°C to 90°C, from 30°C to 90°C, from 35°C to 90°C, from 40°C to 90°C, from 45°C to 90°C, from 50°C to 90°C, from 55°C to 90°C, from 60°C to 90°C, from 10°C to 80°C, from 20°C to 80°C, from 30°C to 80°C, from 35°C to 80°C, from 40°C to 80°C, from 45°C to 80°C, from 50°C to 80°C, from 55°C to 80°C, or from 60°C to 80°C, from 10°C to 70°C, from 20°C to 70°C, from 30°C to 70°C, from 35°C to 70°C, from 40°C to 70°C, from 45°C to 70°C, from 50°C to 70°C, from 55°C to 70°C, or from 60°C to 70°C.

[0169] In exemplary embodiments, the separating of the dispersion into multiple layers occurs or is performed at a temperature of about 10°C to 90°C, 20°C to 90°C, 30°C to 90°C, 40°C to 90°C, 50°C to 90°C, 60°C to 90°C, from 20°C to 80°C, from 30°C to 80°C, 35°C to 80°C, 35°C to 70°C or 40°C to 70°C.

[0170] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream can be performed without addition of salt during the forming of the dispersion and during the separating of the dispersion into the multiple layers. For some exemplary embodiments, the extraction process includes performing the forming, passing, separating and recovering steps without an addition of salt. In other exemplary embodiments, the biomass or acidic feedstock stream containing the biomass in an aqueous salt solution does not include added salt.

[0171] In exemplary embodiments, the liquid-liquid-solid extraction process performed on the acidic feedstock stream is a continuous process. The liquid- liquid-solid extraction process can be configured as a continuous process wherein the forming, passing, separating and recovering steps are performed sequentially. Continuous operation can allow for the production of biofuels and / or other hydrophobic natural products with reduced capital and operating costs. In exemplary embodiments, the acidic feedstock stream and the extraction solvent are contacted so that the solvent receives the hydrophobic natural products. In other exemplary embodiments, the hydrophobic natural products are either pressed from the biomass or extracted with additional algal (or vegetable) oil. The resulting raffinate phase and the extract phases areseparated so that the hydrophobic natural products can be further processed into desirable products.

[0172] A variety of extraction equipment components can be used for continuous extraction including, but not limited to, mixers and settlers, countercurrent extraction columns, centrifugal extractors, and other classes of extractors known in the art as described by Pratt et al., Selection, Design, PilotTesting, and Scale-Up of Extraction Equipment, Chapter 8, in Science and Practice of LiquidLiquid Extraction, Volume 1 , Clarendon Press, Oxford, 1992, the contents of which are incorporated herein by reference. The acidic feedstock stream and the extraction solvent can be contacted in a countercurrent or co-current flow.

[0173] Suitable centrifugal extractors can include, but are not limited to, those manufactured by GEA Westfalia Separator GmbH, which is headquartered in Oelde, Germany; Alfa Laval, with a location in Richmond, Virginia; Robatel, which is located in Pittsfield, Massachusetts; and Podbelniak, which is manufactured by Baker Perkins of Saginaw, Michigan.

[0174] Suitable other extraction equipment includes, but is not limited to, hollow fiber membrane extractors and other novel extractor designs known in the art. In some cases, hollow fiber membrane extractors are used since they obviate the need to separate the solvent from the algal biomass.

[0175] Gravity settling is useful in a continuous extraction process. Separation of the multiple layers can be achieved in a centrifugal or gravitational force field, but gravity settling can be of lower cost. A coalescer can be added to assist in the decantation. The raffinate layer can be coalesced to recover any additional extraction solvent that is entrained before being recycled to a bioreactor or returned to a pond, depending on the type of aquaculture practiced. A coalescer, liquid / liquid / solid centrifuge, flotation cell, and / or liquid / liquid cyclone can be used to recover solvent from the aqueous salt solution, or the aqueous salt solution can be recycled to a flotation device for cleanup.

[0176] Suitable materials for the construction of the mixer, decanter, and / or extraction equipment include, but are not limited to, carbon steel, stainless steel, super duplex steel, Hastelloy, and non-ferrous materials, glass, Teflon, plastics, fiberglass, fiberglass reinforced plastic such as fiberglass reinforcedHDPE, concrete and combinations thereof. Non-ferrous materials are advantageous due to the salt content of the acidic feedstock stream and the raffinate layer in the extraction process. The salinity of these components could cause stress corrosion cracking in ferrous materials, greatly increasing the maintenance required on the mixer, decanter, and extraction equipment. Plastic and fiberglass equipment is resistant to the effects of the elevated salinity and can be less expensive than equipment constructed of ferrous material.

[0177] The solvent extract layer, the lipid-depleted biomass, the rag layer, the raffinate layer and / or a combination thereof can be stabilized against degradation by any means known in the art including, but not limited to, one or more of the following methods: the addition of antioxidants, storage of the material in the absence of light exposure, storage under an inert environment such as nitrogen, argon, or carbon dioxide, chilling, and subjecting the material to a thermal cycle to destroy bacteria. Suitable antioxidants include, but are not limited to carotenoids, tertiary butyl hydroquinone (TBHQ), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), vitamin E, vitamin C, rosemary extracts, sodium citrate, and combinations thereof.

[0178] Exemplary advantages of performing a liquid-liquid-solid extraction process disclosed herein include, but are not limited to, not drying the biomass prior to the hydrophobic natural products being extracted, traditional liquid-liquid extraction equipment can be used instead of expensive leaching equipment, salt does not need to be removed prior to the extraction process, and / or washing of the solvent extract layer can be accomplished in traditional liquidliquid extraction equipment.

[0179] Another aspect of the present disclosure is a process for extracting natural products from a biomass, the process including at least one or more of: forming an acidic feedstock stream having a pH of at most 4.0 and containing a biomass and an aqueous salt solution; contacting the acidic feedstock stream with a first solvent in a first contacting unit to form a first dispersion; allowing the first dispersion to phase separate into a first separated liquid phase and a first remaining dispersion; removing at least a portion of the first separated liquid phase from the first remaining dispersion; contacting the first remaining dispersion with a second solvent in a second contacting unit to form a seconddispersion; allowing the second dispersion to phase separate into a biomassrich phase, a heavy separated liquid phase and a light separated liquid phase; and recovering at least one or more of the biomass-rich phase, the heavy separated liquid phase, and / or the light separated liquid phase for output as recovered products.

[0180] In exemplary embodiments, the process can include measuring decantation curves that form upon and / or during contact in the first contacting unit; selecting a phase with the shortest time to reach 90% of a dimensionless height asymptote and removing a desired amount of said phase from the first contacting unit before passing the first remaining dispersion to the second contacting unit; determining decantation curves formed upon and / or during contact in the second contacting unit; and selecting operating parameters for the second contacting unit to optimize the overall extraction costs of the process.

[0181] FIG. 2 shows an exemplary embodiment of the extraction process wherein natural products are extracted from an acidic feedstock stream in two separate contacting units. In this embodiment, a feedstock stream (200) is transferred into a mixing zone (204). An additive stream (202) is also transferred into the mixing zone (204), where it contacts the feedstock stream (200) to form an acidic feedstock stream (206).

[0182] The acidic feedstock stream (206) in the exemplary process depicted in FIG. 2 is transferred to a first contacting unit (208), wherein it contacts a first solvent (210) to form a first dispersion (212).

[0183] The first dispersion (212) is then transferred to a first separating unit (214), wherein the first dispersion (212) is allowed to separate into a first remaining dispersion (218) and a first separated liquid phase (216). The first separated liquid phase (216) is removed from the first separating unit (214).

[0184] The first remaining dispersion (218) is transferred to a second contacting unit (220), wherein it comes in contact with a second solvent (222) to form a second dispersion (224).

[0185] The second dispersion (224) is then transferred to a second separating unit (226) wherein it is allowed to separate into a light separated liquid phase (228), a biomass-rich phase (230) and a heavy separated liquid phase (232).These three phases are then removed or recovered from the second separating unit (226) for further processing.

[0186] The first contacting unit can be, but is not limited to, a mixer-settler, a counter-current extraction column, a centrifugal extractor, a membrane extractor, an emulsion phase contactor and any extractor that employs nonstandard contact methods (e.g., extractors that use electrical fields, ultrasonic waves, and / or microwave waves to aid in the extraction of materials) and combinations thereof. Suitable mixer-settlers include a mixing section (or contacting unit) and a settling section (or separating unit). Suitable mixers, or contacting units include, but are not limited to static mixers, in-line mixers, agitated vessels, eductors, extraction columns operated in co-current mode, and other means known in the art, and combinations thereof. Suitable settlers or separating units include, but are not limited to gravity decanters, coalescers, electrically enhanced coalescers, cyclones, centrifuges, and other means known in the art, and combinations thereof. Many different designs of mixersettlers have been developed for mining and minerals applications, and these are described by T.C. Lo et al., (1991 ) [Handbook of Solvent Extraction, ISBN 0-89464-546-3],

[0187] Suitable counter-current extraction columns include, but are not limited to, those that are mechanically agitated and those that have stationary internals. The latter is preferred when the first and / or second solvent is a dense gas and / or the operating pressure of the extractor is elevated so that more expensive mechanical seals are needed. Suitable extraction columns with stationary internals can include, but are not limited to, those that contain either structured or random packing, perforated plates, baffle trays, sieve trays, spray columns, and combinations thereof. Suitable packings include structured or random packings that are known to those skilled in the art. Suitable mechanically agitated extraction columns can include, but are not limited to, the Karr reciprocating plate column, the York Scheibel column, and the rotating disc column, all made by Koch Modular Process Technology Corporation, which is located in Parasippany, N.Y., the Kuhni column, which is sold by Sulzer in Switzerland, the asymmetric rotating disc column, the rotating disc column, pulsed columns, and combinations thereof.

[0188] In exemplary embodiments, the counter-current columns have a length- to-width ratio from about 2 to 150, about 2 to 100, from about 2 to 50, from about 2 to 30, about 2 to about 20, from about 2 to about 15, or greater than 2.

[0189] The counter-current columns can operate at nearly atmospheric pressure or under a slight vacuum that is about up to about 30% below atmospheric pressure. In exemplary embodiments, the counter-current columns operate under a slight amount of pressure wherein the solvent or other components in the system vaporize below the column operating pressure (e.g., about 2-10 bar). In other exemplary embodiments, the operating pressure of the counter-current columns is greater than the vapor pressure of the lowest boiling solvent or component in the system so that gas bubbles of solvent are not generated in the counter-current columns.

[0190] The operating temperature in the counter-current columns can range from about 0-100°C, 10-100°C, 20-80°C, or 30-70°C. The minimum temperature that the counter-current column can operate at is based on the viscosity and freezing point of the components in the acidic feedstock stream or any dispersion formed from the acidic feedstock stream. The maximum temperature that the counter-current column can operate at is based on the degradation temperature of the components in the system.

[0191] The counter-current columns can operate at a pH between 1-7, 2-6 or 3-5. The minimum pH can be set to allow preservation of the components in the system when acids are added to stabilize the components. The maximum pH can be set to that which occurs naturally in aquaculture systems during daylight respiration.

[0192] The counter-current columns can operate at about 50% to about 95% of a measured flooding flux. Flux is defined as the volumetric flow rate of the combined liquid phase inputs (e.g., acidic feedstock stream plus the extraction solvent) divided by the cross-sectional area of the counter-current columns. The cross-sectional area of the counter-current columns is taken in the active section, where droplets are flowing in a counter-current manner. For a given counter-current column, the flux can vary considerably depending on the physical properties, including but not limited to the densities and viscosities of each phase along with the interfacial tension, the presence of surfactants, andthe direction of mass transfer. The flux can be increased to the point where the counter-current column floods. In exemplary embodiments, the counter-current column operates close to its flooding point. Thus, a desired flux in the countercurrent column can be found by experimenting with specific systems and operating conditions and measuring the flux at which the counter-current column floods (flooding flux). The counter-current column can be operated so that the chosen operating flux minimizes the overall cost of the counter-current column based on its diameter and height.

[0193] As used herein, the phrase “active section of the counter-current column” refers to the section or area of the counter-current column where at least two liquid phases contact one another and mix with each other.

[0194] Suitable centrifugal extractors that can be used as the first contacting unit include, but are not limited to those produced by CINC, Alfa Lavel, Podbielniak, Robatel, Westfalia, and combinations of these centrifugal extractors. Other suitable centrifugal extractors include but are not limited to those manufactured by GEA Westfalia Separator GmbH, which is headquartered in Oelde, Germany; Alfa Laval, with a location in Richmond, Virginia; Robatel, which is located in Pittsfield, Massachusetts; and Podbielniak, which is manufactured by Baker Perkins of Saginaw, Michigan.

[0195] Suitable emulsion phase contactors that can be used as the first contacting unit include, but are not limited to, those produced by Schlumberger termed the NATCO dual frequency electrostatic treater.

[0196] The first solvent can be any solvent that, upon contacting and / or during contacting with the acidic feedstock stream, is capable of separating the first dispersion into at least a two-layer extraction system, wherein at least one layer rapidly settles.

[0197] A layer that “rapidly settles” is one that reaches 90% of a dimensionless height asymptote within about 30 minutes or less on a decantation curve and has a dimensionless height 90 (DH90) ratio of the fast separating layer to the slow separating layer of less than 0.8. A DH90 ratio is the ratio of the settling time for the rapidly settling layer at DH90 to the settling time for the slowest settling layer at DH90.

[0198] The first solvent can be an extraction solvent including, but not limited to, a non-polar solvent, a non-polar organic solvent, a dense gas solvent, an aqueous two-phase solvent, an ionic liquid, a light solvent, a heavy solvent, or a combination thereof. The first solvent can also be a mixture of miscible solvents. The first solvent is chosen such that it can form an immiscible liquid phase with the acidic feedstock stream and has a density that differs from the acidic feedstock stream. Thus, an optimal extraction solvent can depend on which natural products are desired to be extracted from the acidic feedstock stream.

[0199] The expression “light solvent” refers to a solvent that forms a liquid phase that floats on top of the other liquid phases during the phase separations and is in contact with the vapor phase. The expression “heavy solvent” refers to a solvent that forms a heavy liquid phase that collects at the bottom of a settler and can be in contact with the bottom of the tank or vessel, and / or is just above an even heavier solid phase.

[0200] In exemplary embodiments, the first solvent is a solvent system that forms a two-layer extraction system with the acidic feedstock stream. These solvent systems should not adversely impact the quality or quantity of the natural products. These solvent systems can include any one or more of the non-polar solvents, non-polar organic solvents, aqueous two-phase solvents, deep eutectic solvents, natural deep eutectic solvents, synthetic and / or natural flavorants, edible oils, petrochemicals, bio-based chemicals, dense gases, ionic liquids and combinations of these disclosed herein so long as the mixture of the solvent system and the acidic feedstock stream form two immiscible phases. The solvent system can also include any one or more of the petrochemical solvents disclosed herein due to their low viscosity and favorable solute molecular diffusivity. Natural oils solubilized in petrochemical solvents and concentrated extracts are also possible.

[0201] The first solvent can be a hydrocarbon, ester, ketone, acetate, dense gas, and other solvents identified by J. A. Riddick et al., (1986) [Organic Solvents: Physical Properties and Methods of Purification, 4th Edition, ISBN 0- 471-08467-0],

[0202] In exemplary embodiments, the contacting of the acidic feedstock stream and the first solvent in the first contacting unit occurs under a firstsolvent to acidic feedstock stream ratio (e.g., a volumetric ratio) from about 9 to about 0.1 , from about 5 to about 0.2, or of 1 (e.g., a 1 to 1 volumetric ratio).

[0203] In exemplary embodiments, the contacting of the acidic feedstock stream and the first solvent in the first contacting unit occurs for about 1 minute to 30 minutes, for about 5 minutes to 10 minutes, or any amount of time under 15 minutes. The contacting time can differ based on the type of first contacting unit used and the extraction kinetics to extract the desired solute from the solid matrix. For example, when the first contacting unit is a centrifugal extractor, the contact time can range from about 0.5 to 10 minutes, or less than 2 minutes. When an agitated vessel is used for the first contacting unit, the contact time can range from about 1 minute to 10 minutes. The acidic feedstock stream can contact the first solvent for about 2 to 15 minutes, about 5 to 10 minutes or about 10 to 15 minutes in a counter-current extraction column. Also, for example, when the extraction kinetics of the solute from the solid matrix is rapid relative to the interfacial mass transfer kinetics, then the contact time can range in the various contacting equipment as just described. However, if the extraction kinetics are slow relative to the interfacial mass transfer kinetics, then the required contact time can be longer.

[0204] The term “first dispersion” relates to a heterogeneous mixture containing at least a portion of the acidic feedstock stream and at least a portion of the first solvent along with the biomass.

[0205] In exemplary embodiments, the separation of the first dispersion into the first separated liquid phase and the first remaining dispersion occurs in the first contacting unit. In other exemplary embodiments, the separation of the first dispersion into the first separated liquid phase and the first remaining dispersion does not occur in the first contacting unit and instead occurs within a first separating unit.

[0206] The first separating unit can include, but is not limited to, a decanter, a coalescer, a centrifuge, an electrically enhanced decanter, a settler, a hydroclone or combinations thereof.

[0207] The first contacting unit and the first separating unit can be combined in, for example, a mixer-settler, a counter-current extraction column, acentrifugal extractor, and other methods known in the art, and combinations thereof.

[0208] In embodiments, both the contacting and separating can take place in the same or different units. Contacting can be carried out in a contacting unit and separating can be carried out in a separating unit. Alternatively, both contacting and separating can be carried out in one unit (a combined contacting and separating unit) comprising one or more extraction zones and one or more separation zones, or in one unit (a combined contacting and separating unit) comprising one or more mixing zones, one or more extraction zones and one or more separation zones.

[0209] Additionally, or alternatively, the system or process of the present invention can comprise one or more contacting units and one or more separating units.

[0210] In embodiments, a contacting unit (or any contacting unit, two or more contacting units, or all contacting units) comprises an extraction zone (or two or more extraction zones) and optionally a mixing zone (or two or more mixing zones). In other words, a mixing zone, if present in the system or process of the present invention, can be located in a contacting unit or can be separate from a contacting unit. In embodiments, the mixing zone or a mixing unit comprising the mixing zone is located before the contacting unit. In embodiments, the mixing zone is located before the extraction zone.

[0211] In embodiments, a contacting unit (or any contacting unit, two or more contacting units, or all contacting units) comprises one or more (such as two, three, four, five, six, etc.) extraction zones. Additionally, or alternatively, several contacting units (such as two, three, four, five, six etc.) or several extraction zones (such as two, three, four, five, six etc.) can be placed or used sequentially in the system or process of the present invention.

[0212] In embodiments, a separating unit (or any separating unit, two or more separating units, or all contacting units) comprises one or more (such as two, three, four, five six, etc.) separation zones. Additionally, or alternatively, several separating units (such as two, three, four, five, six etc.) or several separation zones (such as two, three, four, five, six etc.) can be placed or used sequentially in the system or process of the present invention.

[0213] In exemplary embodiments, the first separating unit includes or is a decanter which is configured to perform at least one or more of gravity settling, centrifugal settling, and / or combinations thereof to separate the first dispersion into the first separated liquid phase and the first remaining dispersion. In exemplary embodiments, the first separating unit includes one or more fixed or moving separation aids, for example, mesh pad coalescers, wire pad coalescers, plastic pad coalescers, glass pad coalescers, structured packing, inclined plates, perforated plates, baffles, ultrasonic waves, acoustic waves, electric fields, gravitational fields and / or combinations thereof.

[0214] Additionally, or alternatively, the second separating unit, the third separating unit, and the fourth separating unit are centrifuges suitable for separation, wherein the centrifuges suitable for separation are selected from centrifuges suitable for extraction, such as, but not limited to, centrifugal extractors, decanter centrifuges, and disc-stack centrifuges. Centrifugal extractors are a type of extractors where one or more solutes are transferred between two immiscible liquid phases and phase separation zone by applying a centrifugal force field as the phases flow countercurrent to each other through perforated cylinders to effect contact. Suitable types of centrifugal extractors include, but are not limited to, differential extractors, stage-wise extractors that operate in either multi-stage extractor or single-stage extractors and miscellaneous extractors.

[0215] Additionally, or alternatively, the centrifugal extractors are each independently selected from differential extractors, stage-wise extractors that operate in either multi-stage extractor or single-stage extractors and miscellaneous extractors.

[0216] Examples of differential extractors include, but are not limited to, Podbielniak centrifugal extractor, B&P Littleford ® (Baker Perkins Chemical Machinery Inc.), Quadronic, Sanders Equipment Company, Inc., Florida, USA, De Laval, Tumba, Sweden, Sharpies Super Centrifuge, Albertson, NY, USA, Coil Planet Centrifuge, Nagaoka-cho, Kyoto, Japan, and Rotating packed bed contactors (HiGee), ANDRITZ AG, Graz, Austria.

[0217] Examples of stage-wise extractors include, but are not limited to, multistage extractor e.g., Rousselet-Robatel LX (Annonay, France) and stage-wiseextractors include, but are not limited to, Rousselet-Robatel ® BXP (Annonay, France), Luwesta / Luvesta (Lurgi-Westfalia) contactor, Westfalia Separator AG, Oelde, West Germany, Spinning disc / rotating disc contactor (RDC), KMPS, Paramus, USA, Annular centrifugal contactors for example but are not limited to Argonne National Laboratory (ANL), U.S., CINC ® (Costner Industries Nevada Corporation, Inc.) e.g., Annular centrifugal contactor (ACC), Annular centrifugal extractor (ACE), Centrifugal contactor separator (CCS), Continuous centrifugal contactor separator (CCCS), H-centrifuges, MEAB ®, Sweden, Dorr Oliver, FLSmidth A / S, Copenhagen, Denmark and Bird centrifuge, Bird Machine Company, Inc, Mass., USA.

[0218] The allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion can occur for about 1 minute to 30 minutes, 5 minutes to 15 minutes, 1 minute to 5 minutes, 5 minutes to 10 minutes, or any amount of time under 15 minutes.

[0219] In exemplary embodiments, the first separated liquid phase and the first remaining dispersion do not phase separate in less than 30 minutes under gravitational acceleration.

[0220] The allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion can be performed at ambient conditions so that the temperature can range from 10-100°C and the pressure can be atmospheric. In exemplary embodiments, the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion is performed at room temperature. In other exemplary embodiments, the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion is performed at about 70°C.

[0221] In exemplary embodiments, the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion includes retaining the first dispersion in the first separating unit for a set period of time until the first separated liquid phase and the first remaining dispersion form. The set period of time can be any time from 1 minute to 30 minutes, 2 minutes to 15 minutes, 5 minutes to 15 minutes, 1 minute to 5 minutes, 5 minutes to 10 minutes, or any amount of time under 15 minutes.

[0222] In exemplary embodiments, at least 50 wt%, at least 60 wt%, at least 70 wt%, at least 80 wt%, at least 90 wt% or any amount from 50 wt% to 99 wt% of the total first separated liquid phase is removed from the first separating unit.

[0223] In exemplary embodiments, the first separated liquid phase contains a higher amount of the aqueous solution originating from the acidic feedstock stream than the first remaining dispersion. The first separated liquid phase can contain up to 50 wt%, up to 60 wt%, up to 70 wt%, up to 80 wt%, up to 90 wt% or any amount from 50 wt% to 99 wt% of the aqueous solution relative to the total wt% of the aqueous solution originating in the acidic feedstock stream .

[0224] In exemplary embodiments of the phase separation kinetics, the first separated liquid phase reaches 90% of a dimensionless height asymptote within about 30 minutes ((settle) < 30 min) during the forming of the first separated liquid phase and the first remaining dispersion from the first dispersion.

[0225] In exemplary embodiments, the first separated liquid phase and the first remaining dispersion have a settling time ratio at DH90 of about 0.8 or less.

[0226] In exemplary embodiments, the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion includes maintaining a first solvent to acidic feedstock ratio of about 0.05 to 20, to about 0.1 to 10, and / or a flux of about 10 m3 / h / m2to 80 m3 / h / m2.

[0227] In an exemplary embodiment, the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion includes maintaining a first solvent to acidic feedstock stream ratio of about 0.05 to 20 or about 0.1 to 10.

[0228] In exemplary embodiments, the removing of the first separated liquid phase from the first remaining dispersion can include transferring the first separated liquid phase from the first separating unit and / or the first contacting unit to a first separated liquid phase processing unit. The first separated liquid phase processing unit can include, but is not limited to, crystallization ponds, polishing ponds, aquaculture ponds and / or combinations thereof. In these embodiments, at least a portion of the first remaining dispersion remains in the first separating unit and / or the first contacting unit.

[0229] In exemplary embodiments, the first remaining dispersion is transferred to at least one auxiliary contacting unit and / or separating unit before the first remaining dispersion is transferred to the second contacting unit. For example, the first remaining dispersion can be transferred to at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten or more auxiliary contacting units and / or separating units before being transferred to the second contacting unit.

[0230] In exemplary embodiments, the first remaining dispersion is contacted with the first solvent in one or more auxiliary contacting units. In exemplary embodiments, the first solvent and the first remaining dispersion flow in cocurrent, crossflow, or counter-current mode.

[0231] The second contacting unit can include, but is not limited to, any of the mixer-settlers, counter-current extraction columns, centrifugal extractors, membrane extractors, extractors that employs non-standard contact methods (e.g., extractors that use electrical fields, ultrasonic waves, and / or microwave waves to aid in the extraction of materials) and combinations thereof that the first contacting unit can include. In exemplary embodiments, the second contacting unit contains at least one different extraction equipment from the first contacting unit.

[0232] The second solvent can be any solvent that can form the second dispersion upon and / or during contact with the first remaining dispersion. Alternatively, the second solvent can be any solvent that can form the third dispersion upon and / or during contact with the second remaining dispersion. In exemplary embodiments, the second solvent is the same as the first solvent to reduce the complexity of the solvent recovery system. In other exemplary embodiments, the second solvent differs from the first solvent in order to facilitate better phase separation of the remaining layers. In other exemplary embodiments, the second solvent differs from the first solvent in order to facilitate recovery of a different solute from the system. The second solvent can be any solvent or solvent system of the first solvent.

[0233] In exemplary embodiments, the second solvent is a solvent that has a density that is at least about 0.05 g / cm3less than the density of the first separated liquid phase.

[0234] In exemplary embodiments, the second solvent is a solvent that has a viscosity that is at least about 1000 mPa / s less than the viscosity of the first separated liquid phase.

[0235] In exemplary embodiments, the second solvent is added to the first remaining dispersion in a ratio of about 0.05 to 20 or about 0.1 to 10.

[0236] The term “second dispersion” relates to the heterogeneous mixture containing at least a portion of the first remaining dispersion and at least a portion of the second solvent. The second dispersion can have a higher concentration of biomass than the first dispersion. Therefore, different methods of handling the second dispersion than those deployed to handle the first dispersion can be necessary.

[0237] The term “third dispersion” relates to the heterogeneous mixture containing at least a portion of the second remaining dispersion and at least a portion of the second solvent. The third dispersion can have a higher concentration of biomass than the first dispersion and / or the first remaining dispersion and / or the second remaining dispersion. Therefore, different methods of handling the third dispersion than those deployed to handle the first dispersion and / or the first remaining dispersion and / or the second remaining dispersion can be necessary.

[0238] The term “fourth dispersion” relates to the heterogeneous mixture containing at least a portion of the third remaining dispersion and at least a portion of the third solvent. The fourth dispersion can have a higher concentration of biomass than the first dispersion and / or the first remaining dispersion and / or the second remaining dispersion and / or the third remaining dispersion. Therefore, different methods of handling the third dispersion than those deployed to handle the first dispersion and / or the first remaining dispersion and / or the second remaining dispersion can be necessary and / or the third remaining dispersion.

[0239] In exemplary embodiments, the contacting between the first remaining dispersion and the second solvent in the second contacting unit can occur from 1 minute to about 60 minutes, but additional time can be required if the leaching kinetics are slow relative to the interfacial mass transfer kinetics.

[0240] In exemplary embodiments, the contacting between the second and / or the third remaining dispersion and the second or the third solvent in the second or third contacting unit, respectively, can occur from 1 minute to about 60 minutes, but additional time can be required if the leaching kinetics are slow relative to the interfacial mass transfer kinetics.

[0241] In exemplary embodiments, the separation of the second dispersion into the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase occurs within the second contacting unit. In other exemplary embodiments, the separation of the second dispersion into the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase does not occur in the second contacting unit and instead occurs within a second separating unit.

[0242] In exemplary embodiments, the separation of the fourth dispersion into (i.e., the allowing the fourth dispersion to phase separate into) the biomass-rich phase, the second heavy separated liquid phase and the third light separated liquid phase occurs within the third contacting unit. In other exemplary embodiments, the separation of the fourth dispersion into the separation of the fourth dispersion into (i.e., the allowing the fourth dispersion to phase separate into) the biomass-rich phase, the second heavy separated liquid phase and the third light separated liquid phase does not occur in the third contacting unit and instead occurs within the fourth separating unit.

[0243] The operating conditions of the second contacting and second separating unit can differ significantly from the operating conditions of the first contacting and first separating unit. In exemplary embodiments, the temperature of the first and second contacting and separating units differ. In other exemplary embodiments, the pH and or the salinity of the first and second contacting and separating units differ. In other exemplary embodiments, the pressure of the first and second contacting and separating units differ. In other exemplary embodiments, the equipment types used for the first and second contacting and separating units differ. Other exemplary embodiments, the gravitational acceleration used in the first and second contacting and separating units differ. These differences between the first and second contacting and separating units can result from the removal of the first separated liquid phase.

[0244] In exemplary embodiments, the second dispersion is transferred to at least one other auxiliary contacting unit and / or separating units before the second dispersion is transferred to the second separating unit. For example, the second dispersion can be transferred to at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten or more auxiliary contacting units and / or separating units before being transferred to the second separating unit. In exemplary embodiments, the first remaining dispersion is transferred to at least one other auxiliary contacting unit and / or separating units before contacting in the second contacting unit. For example, the first remaining dispersion can be transferred to at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten or more auxiliary contacting units and / or separating units before being transferred to the second contacting unit. In specific embodiments, the first remaining dispersion can be transferred to at least one, at least two, or at least three auxiliary separating units before being transferred to the second contacting unit. In exemplary embodiments, the process comprises allowing the dispersion to phase separate in two consecutive steps, in two consecutive separating units. In exemplary embodiments, the system comprises two consecutive separating units.Therefore, in specific embodiments, the first separating unit being in communication with the first contacting unit and the second separating unit, wherein the second separating unit being in communication with the second contacting unit.

[0245] In exemplary embodiments, the second dispersion is contacted with the second solvent in one or more auxiliary contacting units. In exemplary embodiments, the second solvent and the first remaining dispersion flow in cocurrent, crossflow, or counter-current mode.

[0246] In exemplary embodiments, the third solvent is the same as the first solvent and / or the second solvent. In exemplary embodiments, the third solvent is different from the first solvent and / or the second solvent. The third solvent can be any solvent or solvent system of the first solvent and / or the second solvent. It is to be understood that the first, the second, and the third solvent may be an extraction solvent as disclosed in the present disclosure.

[0247] In exemplary embodiments, the method includes adding the first solvent, the second solvent, the third solvent and / or any other solvent to any of the auxiliary contacting and / or separating units.

[0248] In exemplary embodiments, the phase separation of the second dispersion into the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase occurs in the second contacting unit. In these embodiments, the second separating unit can be optional.

[0249] In the second contacting unit or the second separating unit, the second dispersion can be contacted with the second solvent. The hydrophobic natural products or algal oils in the second dispersion are transferred from the second dispersion into the extraction solvent (i.e., the second solvent). The second dispersion then separates into multiple phases. One phase can be depleted of hydrophobic algal components and contain the aqueous solution originating from the acidic feedstock stream (i.e., the heavy separated liquid phase).Biomass can collect at a fluid-fluid interface, and this material is a component of the biomass-rich layer (biomass-rich phase). The biomass-rich layer (biomassrich phase) can also include salt-laden lipid-depleted biomass as well as limited amounts of the heavy separated liquid phase. Thus, two immiscible fluid phases, the heavy separated liquid phase and the light separated liquid phase, and a biomass-rich layer (biomass-rich phase) can be formed. Either of the two immiscible fluid phases can be made the continuous phase. The biomass-rich layer (biomass-rich phase) can be formed between the light separated liquid phase, which includes lipids, carotenoids, and the extraction solvent, and the heavy separated liquid phase, which is rich in water and salts and can contain trace amounts of lipids and carotenoids. The biomass of the biomass-rich layer (biomass-rich phase) can be rich in chlorophyll, glycerol, phospholipids and proteins.

[0250] The phase separation of the second dispersion and / or the fourth dispersion can be performed at ambient conditions so that the temperature and pressure can range from 10-100°C and the pressure is atmospheric. The phase separation can be performed at much higher temperatures. Bloch et al. in U.S. Pat. No. 4,341 ,038 teach that phase separation processes can be operated at temperatures up to 300°C, to obtain certain results. The phase separation canoperate at a temperature below 100°C to preserve the algal oils and carotenoids. Likewise, the pressure can be increased for the use of supercritical fluids. U.S. Pat. No. 6,106,720 teaches the advantages of high-pressure phase separations.

[0251] The second separating unit, third separating unit, and / or the fourth separating unit can include, but is not limited to, a decanter, a coalescer, a centrifuge, an electrically enhanced decanter, a hydrocyclone or combinations thereof.

[0252] The second contacting unit and the second separating unit, the second contacting unit and the third separating unit and / or the third contacting unit and the fourth separating unit can be combined in, for example, a batch-operating mixer-settler, counter-current extraction column, a centrifugal extractor, and other methods known in the art, and combinations thereof.

[0253] In exemplary embodiments, the second separating unit includes or is a decanter which is configured to perform at least one or more of gravity settling, centrifugal settling, and / or combinations thereof to separate the second dispersion into the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase. In exemplary embodiments, the second separating unit includes one or more fixed or moving separation aids, for example, mesh pad coalescers, wire pad coalescers, structured packing, inclined plates, perforated plates, baffles, ultrasonic waves, acoustic waves, and / or combinations thereof.

[0254] In exemplary embodiments, the process includes reducing the total extraction volume of the first, second, third, fourth and / or auxiliary contacting units and separating units. In exemplary embodiments, the total extraction volume is reduced by more than 20%, or by more than 40%, when compared to the total extraction volume of a single contacting and separating unit.

[0255] In exemplary embodiments, the second dispersion or the fourth dispersion is allowed a specified period of time to phase separate into the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase. The specified period of time can be from at least 5 minutes to about 24 hours, at least 20 minutes to 12 hours, at least 30 minutes to 6 hours, or 40 minutes to 3 hours.

[0256] The term “biomass-rich phase” and “biomass-rich layer” relates to a fluid layer or phase that includes a majority of the biomass originating from the acidic feedstock stream and can also include at least a portion of the heavy separated liquid phase and at least a portion of the light separated liquid phase.

[0257] The biomass-rich phase or layer can form at any location between, above or below the heavy separated liquid phase and the light separated liquid phase. In exemplary embodiments, the biomass-rich phase forms between the heavy separated liquid phase and the light separated liquid phase.

[0258] In exemplary embodiments, the biomass-rich phase forms below the light separated liquid phase.

[0259] The biomass-rich phase can include a lipid-depleted biomass. The lipid-depleted biomass can include at least one or more of, chlorophyll, glycerol, phospholipids, proteins, carbohydrates, fibers, and limited amounts of lipids, carotenoids and / or salts relative to the second dispersion or any combination thereof.

[0260] In exemplary embodiments, the biomass-rich phase composition contains about 10 wt% biomass, about 45 wt% of light separated liquid phase and about 45 wt% of the heavy separated liquid phase.

[0261] In exemplary embodiments, the biomass-rich phase contains at least 50 wt%, at least 60 wt%, at least 70 wt%, at least 80 wt%, at least 90 wt% or any amount from 50 wt% to 99 wt% of biomass. The biomass present in the biomass-rich phase can include lipid-depleted biomass. The amount of lipid- depleted biomass in the biomass-rich phase can be at most 10 wt%, at most 20 wt%, at most 30 wt%, at most 40 wt%, at most 50 wt%, at most 60 wt%, at most 70 wt%, at most 80 wt%, at most 90 wt% or any amount from 50 wt% to 99 wt% of the total weight of the biomass-rich phase.

[0262] The term “heavy separated liquid phase” relates to a phase or layer that is composed mostly of water or saltwater and can contain at least a portion of the biomass-rich layer (biomass-rich phase) and at least a portion of the second solvent used to create the second dispersion. In exemplary embodiments, less than 10%, 20%, 30% of the biomass-rich layer (biomass-rich phase) is entrained in the heavy separated liquid phase.

[0263] The heavy separated liquid phase can include an aqueous salt solution depleted of hydrophobic natural products.

[0264] The heavy separated liquid phase and / or the first separated liquid phase can possess a salt concentration of above 5 wt%, above 7 wt%, above 10 wt% above 15 wt%, above 18 wt% up to saturation. In exemplary embodiments a salinity of the heavy separated liquid phase layer and / or the first separated liquid phase is about 5 wt% or greater than 5 wt%, about 6 wt% or greater than 6 wt%, about 7 wt% or greater than 7 wt%, for example at least about 8 wt%, at least about 9 wt%, at least about 10 wt%, at least about 11 wt%, at least about 12 wt%, at least about 13 wt%, at least about 14 wt%, at least about 15 wt%, at least about 16 wt%, at least about 17 wt%, at least about 18 wt%, at least about 19 wt%, at least about 20 wt%, at least about 21 wt%, at least about 22 wt%, at least about 23 wt%, at least about 24 wt%, or at least about 25 wt%. In other exemplary embodiments, the heavy separated liquid phase and / or the first separated liquid phase is saturated with salt, e.g., contains a salt concentration of at least 26.5 wt%. In exemplary embodiments, the heavy separated liquid phase and / or the first separated liquid phase can have a salinity that is about 5 wt% to about saturation, from about 10 wt% to saturation, from about 20 wt% to saturation, from about 5 wt% to about 20 wt%, from about 10 wt% to about 20 wt%, from about 5 wt% to about 15 wt%, from about 10 wt% to about 15 wt%, or from about 5 wt% to about 10 wt%.

[0265] In exemplary embodiments, the heavy separated liquid phase includes an aqueous salt solution and a polar organic solvent, for example, methanol, dimethyl sulfoxide, or dimethylformamide.

[0266] The term “light separated liquid phase” relates to a phase or layer that is composed mostly of a solvent and contains at least one hydrophobic natural product originating from the biomass of the acidic feedstock stream.

[0267] The first solvent can include or is the same solvent that is used as the second solvent. The second solvent can also include a portion of the first solvent. The first solvent can include or is the same solvent that is used as the third solvent. The third solvent can also include a portion of the first solvent.

[0268] The light separated liquid phase can also contain at least a portion of the biomass-rich layer (biomass-rich phase).

[0269] The light separated liquid phase can include at least one hydrophobic natural product. These hydrophobic natural products can include, but are not limited to, lipids, algal lipids, carotenoids, fatty acids, algal fatty acids, triacylglycerols, diacylglycerols, monoacylglycerols, oils, algal oils and combinations thereof.

[0270] The carotenoids can include beta-carotene, alpha-carotene, lutein, zeaxanthin, beta-cryptoxanthin, astaxanthin, phytoene, phytofluene, lycopene, and / or combinations thereof.

[0271] The light separated liquid phase can also include limited amounts of lipid-depleted biomass and the aqueous salt solution.

[0272] In exemplary embodiments, the separating of at least one or more of the biomass-rich phase, the heavy separated liquid phase, and / or the light separated liquid phase for output as recovered products includes removing at least a portion of one or more of the biomass-rich phase, the heavy separated liquid phase, and / or the light separated liquid phase from the second contacting unit and / or the second separating unit.

[0273] The separation of the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase can be carried out or performed under one or more of a gravitational, centrifugal, or electrical field. The second separating unit can include, but is not limited to, a decanter, a coalescer, a centrifuge, an electrically enhanced decanter, a hydroclone or combinations thereof.

[0274] The separation of the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase can occur or can be performed at a pressure ranging from atmospheric to supercritical conditions for the solvent.

[0275] In exemplary embodiments, the extraction process can include performing a temperature and / or pressure adjustment before the acidic feedstock stream contacts the first solvent in the first contacting unit, before the first solvent enters the first contacting unit, during the contacting of the acidic feedstock stream with the first solvent in the first contacting unit and / or any combination thereof.

[0276] In exemplary embodiments, the extraction process can include a multiple extraction step process. The multiple extraction step process can beone wherein at least the first contacting unit, the first separating unit, the second contacting unit, the second separating unit and / or any combination thereof is at least duplicated to achieve the extraction process.

[0277] In exemplary embodiments, at least a portion of the light separated liquid phase is overflowed, removed, or pumped out of the second contacting unit or second separating unit, the biomass-rich phase is removed or pumped out of the second contacting unit or second separating unit and / or the heavy separated liquid phase is removed from the bottom of the second contacting unit or second separating unit.

[0278] In exemplary embodiments, the extraction process is configured as a continuous process wherein the forming of the acidic feedstock stream, the contacting of the acidic feedstock stream with the first solvent in the first contacting unit to form the first dispersion, the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion, the removing of at least a portion of the first separated liquid phase from the first remaining dispersion, the contacting of the first remaining dispersion with the second solvent in the second contacting unit to form the second dispersion, the allowing of the second dispersion to phase separate into the biomass-rich phase, the heavy separated liquid phase and the light separated liquid phase, and the separating of the at least one or more biomassrich phase, heavy separated liquid phase, and / or light separated liquid phase are performed sequentially. Continuous operation can allow for the production of biofuels and / or other hydrophobic natural products with reduced capital and operating costs.

[0279] A variety of extraction equipment components can be used for continuous extraction including, but not limited to, the mixers and settlers, the counter-current extraction columns, the centrifugal extractors, and the other classes of extractors disclosed herein.

[0280] The light separated liquid phase, the heavy separated liquid phase, the biomass-rich phase or a combination thereof can be stabilized against degradation by any of the following means including, but not limited to, with the addition of antioxidants, storage of the material in the absence of light exposure, storage under an inert environment such as nitrogen, argon, or carbon dioxide,and subjecting the material to a thermal cycle to destroy bacteria. Suitable antioxidants include, but are not limited to carotenoids, tertiary butyl hydroquinone (TBHQ), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), vitamin E, vitamin C, rosemary extracts, sodium citrate, and combinations thereof.

[0281] In exemplary embodiments, the first contacting unit and the second contacting unit are counter-current extraction columns, wherein the contacting of the acidic feedstock stream with the first solvent to form the first dispersion and the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion occur in the first contacting unit, and wherein the contacting of the first remaining dispersion with the second solvent to form the second dispersion and the allowing of the second dispersion to phase separate into the biomass-rich phase, the heavy separated liquid phase and the separated light separated liquid phase occur in the second contacting unit.

[0282] In exemplary embodiments, the extraction process includes repeating the contacting of the acidic feedstock stream with the first solvent, or another solvent, in the first contacting unit to form the first dispersion and / or repeating the allowing of the first dispersion to phase separate into the first separated liquid phase and the first remaining dispersion in the first separating unit before the first remaining dispersion is transferred to the second contacting unit and / or any auxiliary contacting or separating unit.

[0283] In exemplary embodiments, the formation of the acidic feedstock stream does not involve contacting the feedstock stream with the additive stream in a mixing zone.

[0284] In exemplary embodiments, the process includes contacting the feedstock stream with the additive stream in an extraction zone to form the acidic feedstock stream.

[0285] An exemplary process wherein the extraction process includes forming the acidic feedstock stream in an extraction zone is shown in FIG. 3. In this embodiment, a feedstock stream (300) is transferred to an extraction zone (304). An additive stream (302) is also transferred into the extraction zone(304), wherein it contacts the feedstock stream (300) to form an acidic feedstock stream (306).

[0286] The acidic feedstock stream (306), while in the extraction zone (304), contacts an extraction solvent (308) to form a dispersion (310). The dispersion (310) is transferred to a separation zone (312), wherein it is allowed to separate into a solvent extract layer (314), a rag layer (316) and a raffinate layer (312). The multiple layers formed in the separation zone (312) are then isolated for further processing.

[0287] Mixing of the feedstock stream and the additive stream can occur in the extraction zone and, depending on the equipment included in the extraction zone, the introduction point of these two streams into the extraction zone can be selected to provide optimal mixing of the two streams. In this case, the feedstock stream and the additive stream could be added into the extraction zone at a point where they would be efficiently mixed. This can be accomplished, for example, in the following modes, but other modes known to one ordinarily skilled in the art can also be deployed, these modes including, but not limited to, adding the two streams in the impeller region of an extraction zone, adding the two streams in an active section of a counter-current extraction column, and adding the two streams in an active section of a countercurrent column where the feedstock stream is continuously fed into the column. In other exemplary embodiments, the additive stream is added above the feedstock stream as in a fractional extractor so that the additive stream and the solvent stream flow counter-currently and are mixed in the extraction zone.

[0288] In exemplary embodiments, the extraction process includes contacting the acidic feedstock stream with the extraction solvent in the extraction zone, the extraction zone being a counter-current extraction column, and the process optionally including retaining the dispersion in the counter-current extraction column for a residence time of about 2 minutes to about 2 hours.

[0289] In exemplary embodiments, the separating of the dispersion into multiple layers includes at least one or more of gravity settling, centrifugal settling, and / or combinations thereof.

[0290] In exemplary embodiments, the extraction process includes performing the separating of the dispersion into multiple layers by gravity settling under a gravitational field.

[0291] In exemplary embodiments, the extraction process includes performing the separating of the dispersion into multiple layers by decanting.

[0292] In exemplary embodiments, the at least one hydrophobic natural product is or includes at least one lipid, algal lipid, carotenoid, fatty acid, algal fatty acid, triacylglycerol, diacylglycerol, monoacylglycerol, oil, algal oil, sterol, wax or any combination thereof.

[0293] In exemplary embodiments, the aqueous salt solution has a salinity greater than 0.1wt%, greater than 5 wt%, greater than 10 wt%, greater than 15 wt% or greater than 20 wt%.

[0294] In exemplary embodiments, the aqueous salt solution is or includes the culture medium of the biomass in the feedstock source or biomass culture.

[0295] In exemplary embodiments, the lipid-depleted biomass includes at least one or more of chlorophyll, glycerol, phospholipids, carbohydrates, fibers, proteins or combinations thereof.

[0296] In exemplary embodiments, the extraction process includes performing the process as a continuous process, wherein the forming of the acidic feedstock stream, the contacting of the acidic feedstock stream with the extraction solvent, the separating of the dispersion into multiple layers, and the recovering of at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer are performed sequentially.

[0297] In exemplary embodiments, the process includes recovering at least a portion of the solvent extract layer.

[0298] In exemplary embodiments, the separating of the dispersion includes using a coalescer to assist in the separating of the dispersion into multiple layers.

[0299] In exemplary embodiments, the extraction process includes at least one or more of the following: filtering the solvent extract layer after isolation to remove any entrained biomass; filtering the raffinate layer after isolation to remove any entrained biomass.

[0300] In exemplary embodiments, the extraction process includes evaporating the extraction solvent from the solvent extract layer after isolation of the solvent extract layer.

[0301] In exemplary embodiments, the extraction process includes allowing the acidic feedstock stream to contact the extraction solvent for about 1 minute to about 5 hours.

[0302] In exemplary embodiments, the extraction process includes performing the separating of the dispersion for at least 5 minutes to about 24 hours.

[0303] In exemplary embodiments, the extraction process includes performing the separating of the dispersion at a pressure ranging from atmospheric to supercritical conditions for the extraction solvent.

[0304] In exemplary embodiments, the extraction process includes recovering the rag layer and recovering the lipid-depleted biomass from the rag layer.

[0305] In exemplary embodiments, the extraction process includes removing any residual extraction solvent, salts and / or clay from the isolated lipid-depleted biomass.

[0306] In exemplary embodiments, the extraction process includes pelletizing the isolated lipid-depleted biomass.

[0307] In exemplary embodiments, the extraction process includes recovering the raffinate layer and removing any entrained biomass, any entrained extraction solvent and / or any soluble solvent from the isolated raffinate layer.

[0308] In exemplary embodiments, the acidic feedstock stream contains a water content before extracting.

[0309] In exemplary embodiments, the extraction process includes performing a heat exchange before the acidic feedstock stream contacts the extraction solvent and / or before the separating of the dispersion.

[0310] In exemplary embodiments, the biomass is or includes conditioned biomass.

[0311] In exemplary embodiments, the rag layer is formed between the solvent extract layer and the raffinate layer.

[0312] In exemplary embodiments, the separating of the dispersion into multiple layers occurs in a temperature regulated environment.

[0313] In exemplary embodiments, the raffinate layer has a salinity of about 0.1 wt% or greater, about 5 wt% or greater, about 10 wt% or greater, about 15 wt% or greater, or about 20 wt% or greater.

[0314] In exemplary embodiments, the rag layer contains at most about 60 wt% solvent extract layer (such as from 30 wt% to 60 wt%, e.g. 40 wt% to 50 wt% or about 45 wt%), at most about 60 wt% raffinate layer (such as from 30 wt% to 60 wt%, e.g. 40 wt% to 50 wt% or about 45 wt%) and / or about from 2 wt% to 20 wt% lipid-depleted biomass (e.g. from 5 wt% to 10 wt%).

[0315] In exemplary embodiments, the raffinate layer contains salt from about 0.1 wt% to about 50 wt%, from about 5 wt% to about 40 wt%, from about 10 wt% to 30 wt%, or from about 15 wt% to 26 wt% of the total weight of the raffinate layer.

[0316] In exemplary embodiments, the separating of the dispersion into multiple layers and / or the contacting of the acidic feedstock stream with the extraction solvent occurs at a temperature of about 100°C, less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°C to 90°C, 30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C or 40°C to 70°C. Additionally, or alternatively, the allowing the first dispersion, the first remaining dispersion, the second remaining dispersion, the third dispersion, the third remaining dispersion, and / or the fourth dispersion to phase separate and / or the contacting the (acidic) feedstock stream, the first remaining dispersion, the second remaining dispersion, and / or the third remaining dispersion with a first solvent, a second solvent, or a third solvent occurs at a temperature of about 100°C, less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°C to 90°C, 30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C or 40°C to 70°C. Additionally, or alternatively, the (acidic) feedstock stream, the first dispersion, the first remaining dispersion, the second remaining dispersion, the third dispersion, the third remaining dispersion, and / or the fourth dispersion are each independently heated at a temperature of at least one or more of about 100°C, less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°C to 90°C,30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C and / or 40°C to 70°C.

[0317] Another aspect of the present disclosure is a process for extracting natural products from a biomass, the process including at least one or more of: obtaining a feedstock stream containing the biomass and an aqueous salt solution, the feedstock stream having a pH of more than 4.0 or at least 5.0; transferring the feedstock stream to an extraction zone; introducing an additive stream containing an extraction solvent and at least one acidic additive and / or one buffering agent into the extraction zone to form a dispersion, the least one acidic additive being either a Bronsted acid, an acidic chelating agent or a combination thereof, wherein the dispersion contains an aqueous phase having a pH of at most 4.0; separating the dispersion into multiple layers, the layers including: a solvent extract layer containing at least one hydrophobic natural product and at least a portion of the extraction solvent, a raffinate layer containing at least a portion of the aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer.

[0318] The extraction solvent can be mixed with the additive stream provided that the acidic additive in the additive stream is soluble in the extraction solvent.

[0319] An exemplary process wherein the dispersion is formed from contact of a feedstock stream and an additive stream (e.g., containing at least one acidic additive) and an extraction solvent is depicted in FIG. 4. In this exemplary embodiment, a feedstock stream (400) is transferred to an extraction zone (404). An additive stream (402) containing an extraction solvent and e.g., at least one acidic additive is also transferred to the extraction zone (404). The additive stream (402) and feedstock stream (400) contact one another along with the extraction solvent in the extraction zone (404) and produce a dispersion (406).

[0320] The dispersion (406) is transferred to a separation zone (408), wherein it is allowed to separate into a solvent extract layer (410), a rag layer (412) and a raffinate layer (414). The multiple layers are then removed from the separation zone (408) for further processing.

[0321] In exemplary embodiments, the processes disclosed herein include culturing a feedstock source in an algal aquaculture pond to form a preharvested feedstock stream; transferring the pre-harvested feedstock stream to a harvesting zone; and performing at least one harvesting process on the preharvested feedstock stream to form the feedstock stream.

[0322] Additionally, or alternatively, the process comprises: contacting the acidic feedstock stream with an extraction solvent in a contacting unit to form a dispersion; and separating the dispersion into multiple layers in a separating unit.

[0323] Additionally, or alternatively, both the contacting the acidic feedstock stream with an extraction solvent to form a dispersion, and the separating the dispersion into multiple layers are carried out in a contacting unit or a separating unit.

[0324] Additionally, or alternatively, the process comprises: providing an acidic feedstock stream having a pH of at most 4.0, such as a pH in the range from 3.0 to 4.0, and containing a biomass and an aqueous salt solution, optionally further containing an additive stream containing at least one buffering agent and / or at least one acidic additive selected from a group consisting of a Bronsted acid, an acidic chelating agent and a combination thereof; contacting the acidic feedstock stream with a first solvent in a first contacting unit, such as a vessel, to form a first dispersion; allowing the first dispersion to phase separate in a first separating unit into a first separated liquid phase, such as a first separated (heavy) liquid phase, and a first remaining dispersion; removing at least a portion of the first separated liquid phase, such as the first separated (heavy) liquid phase; allowing the first remaining dispersion to phase separate into a first light separated liquid phase, and a second remaining dispersion in a second separating unit, such as a centrifuge; removing at least a portion of the first light separated liquid phase; contacting the second remaining dispersion with a second solvent in a second contacting unit to form a third dispersion; allowing the third dispersion to phase separate into a second light separated liquid phase, and a third remaining dispersion in a third separating unit, such as a centrifuge; removing at least a portion of the second light separated liquid phase; contacting the third remaining dispersion with a third solvent in a thirdcontacting unit to form a fourth dispersion; allowing the fourth dispersion to phase separate into a biomass-rich phase, a second heavy separated liquid phase and a third light separated liquid phase in a fourth separating unit; and recovering at least one or more of at least a part of the biomass-rich phase, at least a part of one or more of the first and the second separated (heavy) liquid phase, and / or at least a part of one or more of the first, the second, and the third light separated liquid phase for output as products.

[0325] Additionally, or alternatively, the contacting the acidic feedstock stream with a first solvent in a first contacting unit to form a first dispersion comprises agitating the first dispersion in a vessel comprising an impeller, preferably the impeller speed of the impeller is a predetermined impeller value being 110 — 140% of the minimum impeller speed (Nmin) of the impeller, such as 120% or 140% of the minimum impeller speed (Nmin) of the impeller, to disperse two immiscible liquid phases, and heating the first dispersion at a temperature of at least one or more of about 100°C, less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°C to 90°C, 30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C and / or 40°C to 70°C.

[0326] Additionally, or alternatively, the allowing the first remaining dispersion to phase separate into a first light separated liquid phase, and a second remaining dispersion in a second separating unit is performed in a centrifuge at 1500 - 15000 G, such as at 2000 G, optionally for 0.5 - 10 min, such as for 5 minutes, at 5 - 90 °C, or at 35 - 60 °C, such as at 45 - 50 °C.

[0327] Additionally, or alternatively, the extraction solvent, the first solvent, the second solvent, and / or the third solvent is / are a hydrocarbon, such as hexane, heptane, or octane, or any combination thereof. Additionally, or alternatively, the extraction solvent, the first solvent, the second solvent, and the third solvent are heptane.

[0328] Additionally, or alternatively, the dispersion, the first dispersion, the first remaining dispersion, the second remaining dispersion, the third dispersion, the third remaining dispersion, and / or the fourth dispersion are each independently heated at a temperature of at least one or more of about 100°C, less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°Cto 90°C, 30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C and / or 40°C to 70°C.

[0329] Additionally, or alternatively, recovering one or more of the at least part of the solvent extract layer, the at least part of the first light separated liquid phase, the at least part of the second light separated liquid phase, and the at least part of the third light separated liquid phase for output as products; combining one or more of the recovered at least part of the solvent extract layer, at least part of the first light separated liquid phase, at least part of the second light separated liquid phase, and at least part of the third light separated liquid phase to form a combined phase of light separated liquid phases; filtering the combined phase of light separated liquid phases; evaporating the filtered combined phase of light separated liquid phases; and recovering algal oil for output as product.

[0330] Another aspect of the present disclosure is a system for extracting natural products from a biomass, the system including at least one or more of: a mixing zone configured to mix a feedstock stream with an additive stream, and contain an acidic feedstock stream having a pH of at most 4.0; an extraction zone configured to mix an acidic feedstock stream having a pH of at most 4.0 with an extraction solvent, and contain a dispersion having an aqueous phase with a pH of at most 4.0; and a separation zone configured to separate a dispersion having an aqueous phase with a pH of at most 4.0 into a solvent extract layer containing at least one hydrophobic natural product and at least a portion of an extraction solvent, a raffinate layer containing at least a portion of an aqueous salt solution, and a rag layer containing at least a portion of a lipid- depleted biomass.

[0331] Another aspect of the present disclosure is a system for extracting natural products from a biomass, the system including at least one or more of: an acidic feedstock stream import line; a first contacting unit in communication with the acidic feedstock stream import line; a first solvent import line in communication with the first contacting unit; a first effluent export line in communication with the first contacting unit; a second contacting unit in communication with the first contacting unit; a second solvent import line incommunication with the second contacting unit; and a first extraction line in communication with the second contacting unit.

[0332] The different components of the systems disclosed herein can be in communication with each other via the use of any structural component capable of connecting one component of the system to another (e.g., a tube, a pipe, or a canal).

[0333] In exemplary embodiments, the system is designed to where its total extraction volume is reduced by more than 20%, or more than 40%, by adding the second contacting and separating units at the same decantation efficiency of the light and / or heavy phases compared to a single stage system.

[0334] The expression “light phase” refers to the liquid phase that floats on top of the other liquid phases during the phase separations and is in contact with the vapor phase. The expression “heavy phase” refers to the heavy liquid phase that collects at the bottom of a settler and is in contact with the bottom of the tank or vessel, and / or is just above an even heavier solid phase.

[0335] The expression “total extraction volume” means the combined volume of contacting unit(s) and separating unit(s).

[0336] The expression “decantation efficiency” refers to the percentage of approach to a dimensionless height asymptote.

[0337] In exemplary embodiments, the first contacting unit and / or the second contacting unit and / or the third contacting unit include at least one or more of a mixer-settler unit, a counter-current extraction column, a co-current extraction column, a centrifugal extractor, membrane extractors, an emulsion phase contactor, any extractor that relies upon non-standard contact methods and / or any combination thereof.

[0338] In exemplary embodiments, the acidic feedstock stream import line, the first solvent import line, the first effluent export line, the second solvent import line and / or the first extraction line is a pipe, a tube or a canal.

[0339] In exemplary embodiments, the system includes a second extraction line in communication with the second contacting unit; and a third extraction line in communication with the second contacting unit. In exemplary embodiments, any of the acidic feedstock stream import line, the first solvent import line, the first effluent export line, the second solvent import line, the second effluentexport line, the third solvent import line, the third effluent export line, the first extraction line, the second extraction and / or the third extraction line is a pipe, a tube or a canal.

[0340] In exemplary embodiments, the first extraction line is configured to remove a light separated liquid phase from the second contacting unit, the second extraction line is configured to remove a biomass-rich phase from the second contacting unit, and the third extraction line is configured to remove a heavy separated liquid phase from the second contacting unit. Additionally, or alternatively, the first separating unit further comprises an effluent line configured to remove a light separated liquid phase from the first separating unit.

[0341] Additionally, or alternatively, a system for extracting natural products from a biomass, the system comprising: an acidic feedstock stream import line; three contacting units consisting of a first, a second, and a third contacting unit; three solvent import lines consisting of a first, a second, and a third solvent import line; three effluent export lines consisting of a first, a second, and a third effluent export line; four separating units consisting of a first, a second, a third, and a fourth separating unit; and three extraction lines consisting of a first, a second, and a third extraction line; wherein the first contacting unit being in communication with the acidic feedstock stream import line, the first solvent import line, and the first separating unit; the first separating unit being in communication with the first effluent export line, and the second separating unit, the second separating unit being in communication with the second effluent export line, and the second contacting unit, the second contacting unit being in communication with the second solvent import line, and the third separating unit; the third separating unit being in communication with the third effluent export line, and the third contacting unit, the third contacting unit being in communication with the third solvent import line, and the fourth separating unit; and the fourth separating unit being in communication with the first, the second, and the third extraction line.

[0342] Additionally, or alternatively, the first effluent export line is configured to remove a first separated (heavy) liquid phase from the first separating unit, the second effluent export line is configured to remove a first light separated liquidphase from the second separating unit, the third effluent export line is configured to remove a second light separated liquid phase from the third separating unit, the first extraction line is configured to remove a third light separated liquid phase from the fourth separating unit; the second extraction line is configured to remove a biomass-rich phase from the fourth separating unit; and the third extraction line is configured to remove a second heavy separated liquid phase from the fourth separating unit.

[0343] Additionally, or alternatively, the system further comprises: a mixing zone in communication with the first contacting unit; a feedstock stream import line in communication with the mixing zone; and an additive stream import line in communication with the mixing zone.

[0344] Additionally, or alternatively, the system further comprises six transport lines consisting of a first, a second, a third, a fourth, a fifth, and a sixth transport line, wherein the first transport line being in communication with the first contacting unit and the first separating unit; the second transport line being in communication with the first separating unit and the second separating unit; the third transport line being in communication with the second separating unit and the second contacting unit; the fourth transport line being in communication with the second contacting unit and the third separating unit; the fifth transport line being in communication with the third separating unit and the third contacting unit; and the sixth transport line being in communication with the third contacting unit and the fourth separating unit.

[0345] Additionally, or alternatively, the transport lines are each independently selected from the group consisting of a pipe, a tube, and a canal.

[0346] Additionally, or alternatively, the first remaining dispersion comprises a pure emulsion or two (partly) separated phases.

[0347] Additionally, or alternatively, recycling at least a part of the third light separated liquid phase as second solvent. Additionally, or alternatively, recycling at least part of the third light separated liquid phase as second solvent to the second contacting unit. Additionally, or alternatively, the fourth separating unit is configured to recycle at least part of the third light separated liquid phase as the second solvent to the second contacting unit. Additionally, or alternatively, the second contacting unit is configured to receive at least a partof the third light separated liquid phase as second solvent. Additionally, or alternatively, the fourth separating unit is in communication with the second solvent import line configured to introduce the second solvent and / or the at least part of the third light separated liquid phase into the second contacting unit.

[0348] Additionally, or alternatively, the fourth separating unit comprises a recycling extraction line for recycling at least a part of the third light separated liquid phase as second solvent and the recycling extraction line is in communication to the second contacting unit.

[0349] In exemplary embodiments, the first contacting unit and the second contacting unit and / or the third contacting unit are counter-current extraction columns.

[0350] In exemplary embodiments, the system includes a first separating unit in communication with the first contacting unit and the second contacting unit. In exemplary embodiments, the system includes a first transport line in communication with the first contacting unit and the first separating unit. The first transport line can be a pipe, a tube or a canal.

[0351] In exemplary embodiments, the system includes a second separating unit in communication with the second contacting unit and at least one or more of the first extraction line, the second extraction line, the third extraction line and / or any combination thereof. In exemplary embodiments, the system includes a second transport line in communication with the second contacting unit and the second separating unit. The second transport line can be a pipe, a tube or a canal.

[0352] In exemplary embodiments, the first contacting unit and the first separating unit are part of a single extraction unit, e.g., a counter-current extraction column.

[0353] In exemplary embodiments, the second contacting unit and the second separating unit are part of a single extraction unit, e.g., a counter-current extraction column. In exemplary embodiments, the system further includes a third separating unit, a third contacting unit, and a fourth separating unit, wherein the second contacting unit and the third separating unit are part of a single extraction unit, e.g., a counter-current extraction column, and the third contacting unit and the fourth separating unit are part of a single extraction unit,e.g., a counter-current extraction column, and wherein the second separating unit being in communication with the first separating unit and the second contacting unit.

[0354] In exemplary embodiments, the system includes more than two contacting units and / or separating units. For example, the system can include at least three, four, five, six, seven, eight, nine, ten or more auxiliary contacting units; and at least three, four, five, six, seven, eight, nine, ten or more auxiliary separating units. These additional auxiliary contacting and separating units can include any of the extraction equipment that the first contacting unit, the first separating unit, the second contacting unit and the second separating unit can have. These additional auxiliary contacting and separating units can be separate units from each other and / or can be part of a single extraction unit, e.g., a counter-current extraction column.

[0355] In exemplary embodiments, the first extraction line is configured to remove a light separated liquid phase from the second separating unit, the second extraction line is configured to remove a biomass-rich phase from the second separating unit, and the third extraction line is configured to remove a heavy separated liquid phase from the second separating unit.

[0356] In exemplary embodiments, the first effluent export line is configured to remove a first separated (heavy) liquid phase from the first separating unit; the second effluent export line is configured to remove a first light separated liquid phase from the second separating unit; the third effluent export line is configured to remove a second light separated liquid phase from the third separating unit; the first extraction line is configured to remove a third light separated liquid phase from the fourth separating unit; the second extraction line is configured to remove a biomass-rich phase from the fourth separating unit; and the third extraction line is configured to remove a second heavy separated liquid phase from the fourth separating unit.

[0357] Additionally, or alternatively, the first separating unit is configured to separate the first dispersion into a first separated liquid phase, such as a first separated (heavy) liquid phase, a light separated liquid phase, and a first remaining dispersion.

[0358] Additionally, or alternatively, the first separating unit, such as a settler or a decanter, further comprises an outlet for removing a light separated liquid phase.

[0359] Additionally, or alternatively, the allowing the first dispersion to phase separate in a first separating unit is allowing the first dispersion to phase separate in a first separating unit into a first separated liquid phase, such as a first separated (heavy) liquid phase, a light separated liquid phase, and a first remaining dispersion. Additionally, or alternatively, the process further comprises, before or after removing at least a portion of the first separated liquid phase, removing at least a portion of the light separated liquid phase.

[0360] In exemplary embodiments, the systems are configured for recovering products from an acidic feedstock stream containing biomass in an aqueous salt solution and can include one or more of the following components: an acidic feedstock stream import line configured for delivering an acidic feedstock stream to a first contacting unit, wherein the first contacting unit is configured for receiving an acidic feedstock stream from the acidic feedstock stream import line and a first solvent from a first solvent import line, the first contacting unit including: a first means configured for separating an acidic feedstock stream into a first separated liquid phase and a first remaining dispersion, a second means configured for removing at least a portion of a first separated liquid phase from a first remaining dispersion, and a third means configured for transferring a first remaining dispersion to a second contacting unit, wherein the second contacting unit includes: a fourth means configured for separating a first remaining dispersion into a biomass-rich phase, a heavy separated liquid phase and a light separated liquid phase and a fifth means configured for removing at least one or more of a biomass-rich phase, a heavy separated liquid phase and a light separated liquid phase from the second contacting unit.

[0361] In exemplary embodiments, the first means and / or the fourth means is at least one or more of a mixer-settler unit, a counter-current extraction column, a co-current extraction column, a centrifugal extractor, membrane extractors, an emulsion phase contactor, any extractor that relies upon non-standard contact methods and / or any combination thereof.

[0362] In exemplary embodiments, the first means and / or the fourth means include any separating unit disclosed herein.

[0363] In exemplary embodiments, the second means is a first effluent export line in communication with the first contacting unit.

[0364] In exemplary embodiments, the third means is a first stream forward line in communication with the first contacting unit and the second contacting unit.

[0365] In exemplary embodiments, the fifth means is at least one extraction line in communication with the second contacting unit.

[0366] In exemplary embodiments, the fifth means includes at least one or more of a first extraction line configured to remove a light separated liquid phase from the second contacting unit, a second extraction line configured to remove a biomass-rich phase from the second contacting unit, and a third extraction line configured to remove a heavy separated liquid phase from the second contacting unit.

[0367] In exemplary embodiments, the fourth means is in communication with a second solvent import line configured to introduce a second solvent into the second contacting unit.

[0368] In exemplary embodiments, the fourth means includes a second separating unit, and the fifth means includes at least one or more of a first extraction line configured to remove a light separated liquid phase from the second separating unit, a second extraction line configured to remove a biomass-rich phase from the second separating unit, and a third extraction line configured to remove a heavy separated liquid phase from the second separating unit.

[0369] In exemplary embodiments, the acidic feedstock stream import line, the first solvent import line, the second means, the third means, the fifth means and / or any combination thereof is a pipe, a tube or a canal.

[0370] In exemplary embodiments, the system includes more than two contacting units configured to perform any extraction process described herein. For example, the system can include at least three, four, five, six, seven, eight, nine, ten or more auxiliary contacting units possessing any of the first means, second means, third means and / or fourth means discussed herein.

[0371] In exemplary embodiments, the systems disclosed herein include at least one harvesting zone configured to operatively communicate with an extraction zone, a contacting unit and / or a mixing zone; and at least one algal aquaculture zone configured to operatively communicate with the at least one harvesting zone and / or the mixing zone. In exemplary embodiments, the at least one harvesting zone is configured to perform an adsorptive bubble separation process on an algal concentrate.

[0372] In exemplary embodiments, the systems of the present disclosure are for carrying out the processes of the present disclosure.

[0373] The natural products recovered with the systems and processes disclosed herein can include at least one or more of lipids, algal lipids, carotenoids, fatty acids, algal fatty acids, triacylglycerols, diacylglycerols, monoacylglycerols, oils, algal oils or any combination thereof.

[0374] The natural products and / or the biomass recovered from the systems and processes disclosed herein can be used to produce various products including, but not limited to, biofuels, animal feed ingredients, renewable plastics, renewable polymers, renewable chemicals, nutraceuticals, cosmaceuticals, soaps or components of a soap or detergent compositions, and cosmetic ingredients (e.g., carotenoids, omega fatty acids, and other lipids).

[0375] Biofuels that can be produced from high temperature processing of the recovered natural products and / or biomass include, but are not limited to, biodiesel, green diesel, renewable diesel, methane, alcohols, and dried algal biomass. Algal biodiesel is produced via any transesterification process known in the art, including those which utilize two immiscible liquid phases, and those that utilize a solid acid catalyst. Green diesel can be produced by hydrogenation, cracking, or a combination thereof of algal oil or any derivative thereof in order to produce hydrocarbons that can be used directly in the existing diesel distribution system. Methane and / or hydrogen can be produced from the recovered natural products and / or biomass by any anaerobic process known in the art. Fermentation of the recovered natural products and / or biomass can occur through any process known in the art to produce methanol, ethanol, butanol, n-butanol, i-butanol, other alcohols, and combinations thereof. The recovered natural products and / or biomass can be torrified for theproduction of a soil builder or for use in combination with coal for power or steam generation. The recovered natural products and / or biomass can be gasified or combusted either by itself or in combination with coal or biomass.

[0376] Suitable animal feeds include, but are not limited to, feeds for shrimp, fish, shellfish, brine shrimp, chickens, poultry, cows, ducks, dogs, pigs, sheep, goats, and combinations thereof.

[0377] Suitable dietary supplements include, but are not limited to alpha carotene, betacarotene, lutein, zeaxanthin, cryptoxanthin, phytoene, phytofluene, and the various cis- and trans-isomers and the various alpha, beta, gamma, delta isomers of the various carotenoids, and combinations thereof.

[0378] Suitable methods of carbon storage include, but are not limited to, burying the recovered natural products and / or biomass, sinking them, torifying them and using them as a soil builder, or combinations thereof.

[0379] Suitable methods for water and wastewater treatment include, but are not limited to, removal of BOD (biological oxygen demand), and or TOC (total organic carbon) from a water stream. This can be useful for municipal wastewater treatment processes, and it can be important for the treatment of brines being used for the production of sodium chloride salt and other salts via evaporation.

[0380] Suitable methods to process the recovered natural hydrophobic products and / or biomass into useful compounds include, but are not limited to, torrefaction, gasification, liquefaction, fermentation, drying, combustion, burial, and combinations thereof. Suitable applications of the torrified natural products and / or biomass include, but are not limited to, use as a soil builder and a material to be combined with coal, wood, or other combustible material for power generation. Suitable applications of gasified natural products and / or biomass include, but are not limited to, the production of the entire suite of products that can be produced via syngas chemistry, as described by the Gasification Technologies Council. Suitable products from syngas include, but are not limited to, chemicals, fertilizers, power generation, substitute natural gas, hydrogen, and transportation fuels. Suitable chemicals include, but are not limited to, hydrogen, carbon monoxide, methanol, dimethyl ether, acetic acid, propionic acid, butyric acid, acetic anhydride, methyl acetate, ethylene,propylene, olefins, and combinations thereof. Suitable fertilizers that can be produced from the syngas include, but are not limited to ammonia, ammonium nitrate, urea, and others known in the art. Suitable substitute natural gas can be generated from the syngas produced by gasifying the recovered natural products and / or biomass, and this includes methane. Suitable liquid fuels include gasoline, diesel fuel, jet fuels, and combinations thereof. All of the chemicals that are produced by Eastman Chemicals and by Sasol via their gasification processes can also be produced by the gasification of the recovered natural products and / or biomass. Products produced by the utilization of syngas can also be produced by gasification of the recovered natural products and / or biomass. Illustrative processes are described in U. S. Pat. No. 6,310,260, the contents of which are incorporated herein by reference in their entirety, include, for example, hydroformylation, hydroacylation (intramolecular and intermolecular), hydrocyanation, hydroamidation, hydroesterification, aminolysis, alcoholysis, hydrocarbonylation, reductive hydroformylation, hydrogenation, olefin oligomerization, hydroxycarbonylation, carbonylation, olefin isomerization, transfer hydrogenation and the like. Other processes involve the reaction of organic compounds with carbon monoxide, or with carbon monoxide and a third reactant, e.g., hydrogen, or with hydrogen cyanide, in the presence of a catalytic amount of a metal-organophosphorus ligand complex catalyst. More advantageous processes include hydroformylation, hydrocyanation, hydrocarbonylation, hydroxycarbonylation and carbonylation.Examples

[0381] The present disclosure will be described in more detail with reference to the following Examples, which shows exemplary embodiments in accordance with the present disclosure. The present disclosure is not limited to these exemplary embodiments.

[0382] General Procedures for Examples 1-3

[0383] Three liquid-liquid-solid algae biomass extractions were carried out in conical 50 mL test tubes at varying pH levels, while other conditions were kept the same. FIG. 5 illustrates the extraction and decantation process performed for these examples step by step.

[0384] Brine (1844 g) was added to harvested induced Dunaliella salina biomass (158 g) to make up a mixture with desired algae biomass concentration and salinity. The resulting 2 wt% dispersion of fracked Dunaliella salina biomass in aqueous brine including 20 wt% sodium chloride salt was then mixed. Parameters for Examples 1-3 are shown in Table 1 .Table 1 : Parameters for Examples 1-3‘Extractions were carried out in a 50 mL tube

[0385] Table 1 Column Notations: A. Example number; B. Weight-% sodium chloride in the Feed; C. Weight-% biomass in the Feed; D. Additive (acid or base) used to adjust the pH of the feed; E. pH of the feed after the pH adjustment; F. Extraction Solvent; G. Solvent:Feed volumetric ratio; H. Agitation time in minutes; I. Vortex mixer speed in revolutions per minute; J. Extraction and Decantation Temperature in °C.

[0386] Example 1

[0387] An aliquot of fracked algal feed (20 g) was treated with an aqueous 20 wt% citric acid solution (0.20 g) to lower the pH from 5.4 to 3.0. Next, 10 mL of this feed (pH 3.0) was charged to a 50 mL tube followed by careful addition of heptane (5 mL) onto the feed. The tube was sealed with a cap and the multiphase mixture was then heated to 60°C, after which it was vortex-mixed for 2 min with a mixing speed of 1600 rpm. Once mixing was stopped, decantation of the bottom layer at 60°C was recorded over time until equilibrium was reached.

[0388] Example 2

[0389] A 10 mL aliquot of fracked algal feed was used as such without pH adjustment. The experiment was carried out otherwise the same as Example 1 .

[0390] Example 3

[0391] A 20 g aliquot of the fracked algal feed was treated with an aqueous 2 wt% NaOH solution (0.20 g) to raise the pH from 5.4 to 7.0. The experiment was carried out otherwise the same as Example 1 .

[0392] Table 2 summarizes the results of Examples 1-3 after decantation. The initial rate of the separation was highest at pH 3 (0.015 depth / s) while for pH 5.4 and 7 the initial rate was the same (0.011 depth / s). Also, after 60 min decantation time, the biomass concentration in raffinate was lowest at pH 3. Table 2: Results for Examples 1-3 after Extraction and Decantation

[0393] Table 2 Column Notations: A. Example number; B. pH of the feed (after the pH adjustments for Examples 1 and 3); C. Initial rate of bottom layer separation (dimensionless depth / seconds); D. Relative initial rate of separation compared to Example 2. The relative initial rate of separation was calculated by dividing the initial rate of separation in mm / second by the initial rate of separation for the non-modified case. The higher the number, the faster the separation rate; E. Measured biomass concentration in raffinate after 60 min decantation time.

[0394] General Procedure for Examples 4-8

[0395] The extraction process performed in Examples 4-8 is shown in FIG. 6. The counter-current extraction column used in Examples 4-8 was a Karrreciprocating plate extractor. Parameters and results for Examples 4-8 are presented in Table 3 and Table 4, respectively. Here, essentially comparable are Examples 4 and 5 (effect of pH), Examples 6 and 7 (effect of pH), and Examples 7 and 8 (temperature effect). Table 3: Parameters for Examples 4-8*top of the column, heptane as solvent, agitation rate was kept approximately the same (within 15% error margin), flux was kept constant and was approximately 80% of the flood point.

[0396] Table 3 Column Notations: A. Example number; B. Biomass cultivated in different conditions (non-induced refers to unstressed while induced refers to moderately stressed); C. Weight-% biomass in the Feed to the extractor; D. pH of the feed (after possible pH adjustment by citric acid); E. Solvent:Feed mass ratio; F. Solvent:Feed volumetric ratio; G. Extraction and Decantation Temperature in °C.

[0397] Example 4

[0398] The objective of this example was to separate the aqueous phase and algal biomass from each other and extract algal oil and carotenoids from algal biomass.

[0399] A feedstock stream (600) having a pH of 6 and including water, salt, fracked non-induced Dunaliella salina biomass, carotenoids, and algal oil was fed to the top of an extraction column (604), and it was preheated to a temperature of 75°C.

[0400] An extraction solvent stream (602) including heptane was fed to the bottom of the extraction column (604), and this stream was preheated to 70°C. The temperature of the extraction mixture was maintained at 75°C. The aqueous phase was maintained as the continuous phase.

[0401] The first raffinate stream (Raffinate 1 , 606) included primarily water and salt. The ratio of the algal biomass concentration in the first raffinate stream divided by the feed stream was reduced to 0.31 (biomass loss in raffinate: 18 wt%).

[0402] The remainder of the algal biomass along with the solvent phase including heptane, algal oil and carotenoids were removed as an overflow stream (608) and fed to a tricanter (610).

[0403] In the external tricanter (610), three phases, or layers, were formed. These phases, or layers, were: (i) a top phase (i.e., a solvent extract layer or Extract 1 , 612) including heptane, algal oil and carotenoids; (ii) a middle phase (i.e., a rag layer, 614) including heptane, water, salt, algal biomass, carotenoids and algal oil; and a bottom phase (i.e., a raffinate layer or Raffinate 2, 616) including mainly water and salt.

[0404] A sample from the middle phase (614) was transferred to a centrifuge (618) and centrifuged at 2500 G for 10 min at 25°C. Three phases were formed: (i) a top phase (i.e., an extract phase or Extract 2, 620), (ii) a middle phase (i.e., a biomass phase, 622) and (iii) a bottom phase (i.e., a raffinate phase, 624).The top phase (620) including heptane, algal oil and carotenoids, was removed.

[0405] The solvent extract layer (612) and extract phase (620) streams were combined. The resulting extract stream was analyzed for carotenoids.Carotenoid yield was 28 wt%. After removing the solvent from the extract, the resulting oil was analyzed for total fatty acids and metals. Total fatty acid yieldwas 27 wt%. The content of common chelatable metals (Mg, Ca, Fe) in oil was 1385 mg / kg.

[0406] Example 5

[0407] The process in this example was carried out as in Example 4 with the following exceptions and data:

[0408] The objective of this example was to study the effect of low pH on: 1 ) the extraction of algal oil and carotenoids from algal biomass and 2) the composition of the extracted oil. This example is essentially comparable with Example 4.

[0409] Citric acid was added to the feed stream to adjust the pH of the feed stream from 6 to 3.

[0410] The first raffinate stream (aqueous phase, Raffinate 1 ) included primarily water, salt, and citric acid. The ratio of the algal biomass concentration in the first raffinate stream divided by the feed stream was reduced to 0.18 (biomass loss in raffinate: 11 wt%).

[0411] In a tricanter, three phases were formed. Top phase (Extract 1 ) included heptane, algal oil and carotenoids. Middle phase (Rag layer) included heptane, water, salt, algal biomass, carotenoids and algal oil. Bottom phase (Raffinate 2) included mainly water, salt and citric acid.

[0412] By citric acid addition, the interphase in the tricanter between top and middle phase was clear, thus allowing better interphase control. Less strong emulsion with lower viscosity was formed.

[0413] Carotenoid yield was 77 wt%. Total fatty acid yield was 54 wt%. The content of common chelatable metals (Mg, Ca, Fe) in oil was 307 mg / kg.

[0414] Compared to Example 4, the presence of citric acid as a hydrophilic acidic chelating agent increased the carotenoid yield from 28 to 77 wt% and the total fatty acid yield from 27 to 54 wt%, while the chelatable metal content in the oil decreased from 1385 to 307 mg / kg. Additionally, the free fatty acid (FFA) content in oil increased from 22 to 35 %, thus supporting the role of acidic hydrolysis during the algae extraction process.

[0415] Example 6

[0416] The objective of this example was to separate the aqueous phase and algal biomass from each other and extract algal oil and carotenoids from algal biomass.

[0417] A feed stream including water, salt, fracked induced algal biomass, carotenoids, and algal oil was fed to the top of an extraction column, and it was preheated to a temperature of 75°C.

[0418] A solvent stream including heptane was fed to the bottom of the extraction column, and this stream was preheated to 70°C. The solvent to feed mass ratio was maintained at 0.4. The temperature of the extraction mixture was maintained at 75°C. The aqueous phase was maintained as the continuous phase.

[0419] The first raffinate stream (aqueous phase, Raffinate 1 ) included primarily water and salt. The ratio of the algal biomass concentration in the first raffinate stream divided by the feed stream was reduced to 0.14 (biomass loss in raffinate: 11 wt%).

[0420] The remainder of the algal biomass, along with the solvent phase including heptane, algal oil and carotenoids, were removed as the overflow and fed to the tricanter.

[0421] In an external tricanter, three phases were formed. Top phase (Extract 1 ) including heptane, algal oil and carotenoids. Middle phase (Rag layer) including heptane, water, salt, algal biomass, carotenoids and algal oil. Bottom phase (Raffinate 2) including mainly water and salt.

[0422] A sample from the middle phase was centrifuged (2500 G, 10 min, 25°C) and three layers were formed. The top phase (Extract 2), including heptane, algal oil and carotenoids, was removed.

[0423] Extract 1 and Extract 2 streams were combined. The resulting extract was analyzed for carotenoids. Carotenoid yield was 91 wt%. After removing the solvent from the extract, the resulting oil was analyzed for total fatty acids and metals. Total fatty acid yield was 38 wt%. The content of common chelatable metals (Mg, Ca, Fe) in oil was 990 mg / kg.

[0424] Example 7

[0425] The objective of this example was to study the effect of low pH on the extraction of algal oil and carotenoids from algal biomass and the composition of the extracted oil. This example is essentially comparable with Example 6.

[0426] Citric acid was added to the feed stream to adjust the pH of the feed stream from 6 to 3.

[0427] The first raffinate stream (aqueous phase, Raffinate 1 ) included primarily water, salt, and citric acid. The ratio of the algal biomass concentration in the first raffinate stream divided by the feed stream was reduced to 0.16 (biomass loss in raffinate: 12 wt%).

[0428] In an external tricanter, three phases were formed. Top phase (Extract 1 ) including heptane, algal oil and carotenoids. Middle phase (Rag layer) including heptane, water, salt, algal biomass, carotenoids and algal oil. Bottom phase (Raffinate 2) including mainly water, salt and citric acid.

[0429] By citric acid addition, the interphase in the tricanter between the top and middle phase was clear, thus allowing better interphase control. Less strong emulsion with lower viscosity was formed.

[0430] Carotenoid yield was 91 wt%. Total fatty acid yield was 62 wt%. The content of common chelatable metals (Mg, Ca, Fe) in oil was 376 mg / kg.

[0431] Compared to Example 6, the presence of citric acid as a hydrophilic acidic chelating agent increased the carotenoid yield from 69 to 91 wt% and the total fatty acid yield from 38 to 62 wt%, while the chelatable metal content in the oil decreased from 990 to 376 mg / kg. Additionally, the free fatty acid (FFA) content in oil increased from 8 to 23 % thus supporting the role of acidic hydrolysis during the algae extraction process.

[0432] Example 8

[0433] The process in this example was carried out as Example 7 with the following exceptions and data:

[0434] The objective of this example was to study the effect of low pH and temperature on the extraction of algal oil and carotenoids from algal biomass and the composition of the extracted oil. This example is essentially comparable with Example 7.

[0435] A solvent stream including heptane was fed to the bottom of the extraction column, and this stream was preheated to 50°C. The temperature of the extraction mixture was maintained at 50°C.

[0436] The ratio of the algal biomass concentration in the first raffinate stream divided by the feed stream was reduced to 0.29 (biomass loss in raffinate: 27 wt%).

[0437] Carotenoid yield was 91 wt%. Total fatty acid yield was 51 wt%. The content of common chelatable metals (Mg, Ca, Fe) in oil was 318 mg / kg.

[0438] Compared to Example 7, the lower temperature did not affect carotenoid yield, but total fatty acid yield decreased from 62 to 51 wt% while the chelatable metal content in the oil decreased slightly from 376 to 318 mg / kg.Table 4: Results for Examples 4-8 after Extraction and Separation

[0439] Table 4 Column Notations: A. Example number; B. Approximate biomass loss in Raffinate 1 (wt%); C. Carotenoid yield (wt%) to Extract; D. Total fatty acid yield (wt%) to extracted algae oil (measured after hydrolysis of fatty acid esters); E. Approximate free fatty acid (FFA) content in extracted algae oil (%); F. Common chelatable metal (Mg, Ca, Fe) content in total extracted algae oil (mg / kg).

[0440] Total soluble nitrogen was measured to estimate the soluble protein (and amino acid) content in the aqueous phase (raffinate) for Examples 6-8. The results are shown in Table 5 below and demonstrate the benefit of lowering the pH to minimize soluble protein losses in the raffinate.Table 5: Total soluble nitrogen results for Examples 6-8

[0441] Table 5 Column Notations: A. Example number; B. pH of the first raffinate after the extraction and decantation; C. Extraction temperature; D. Total soluble nitrogen content in the first raffinate (measured at 25°C after filtration).

[0442] General information for the Mixer-Settler process (Examples 9-11 )

[0443] The following experimental equipment and methods were used for Examples 9-11 . The liquid-liquid-solid extraction process was performed in a mixer-settler unit (a combined contacting and separating unit), for the isolation of natural products from algal biomass, where the cell membrane is predominantly disrupted, in an aqueous brine solution with a solvent. The mixing and settling were performed in the same vessel with the following geometrical parameters: 1 ) liquid depth in vessel to vessel diameter ratio = 1 ; 2) baffle width = 0.1 X vessel diameter, with four equally spaced baffles that extended from the gas-liquid interface to the start of the bottom dish; 3) impeller diameter = 0.5 X vessel diameter; 4) 3-blade HE-3 impeller from Chemineer (Dayton, Ohio) was centrally mounted in the vessel, 5) a variable speed drive was used to control and set the agitation rate, 6) a condenser was used to retain the solvent while allowing the system to be swept with an inert gas, 7) the jacketed agitated vessel was fitted with sight glasses so that the mixing and decantation steps could be visually observed, and 8) the vessel was cylindrical with a shallow dish at the bottom with a bottom drain valve that is typical of agitated vessels used commercially. This agitated vessel served as both themixer (first contacting unit) (708) and settler (first separating unit) (714) in these examples (see FIG. 7). Thus, the impeller was operated during the mixing step, and it was turned off to initiate the settling step. The algal feed comprised algal biomass and brine (22.5 wt% NaCI, to adjust the salinity of the feed to 20 wt% NaCI) that was initially charged to the agitated vessel. The heat transfer fluid in the jacket on the agitated vessel was set to the desired temperature for the run, and agitation was started at a rate to facilitate proper heat transfer. When the algal feed has reached the desired temperature for the run, the agitation rate was then increased to the desired impeller speed for the experiment, and the flow patterns were established with the algal feed before the first solvent (710) was rapidly added to the agitated vessel at time zero for mixing. After the prescribed mixing time was reached, the impeller operation was stopped at time zero for decantation. The elevation of the interface between the first separated liquid phase (716) and the first remaining dispersion (718) was visually determined and recorded as a function of time. Likewise, the possible elevation in the vessel of the interface between the first light separated liquid phase (722), if any separated, and the biomass rich phase (748), if any separated, was visually determined and recorded as a function of time during the decantation process. The interface elevations were converted to dimensionless depths by dividing the measured elevation above the bottom of the vessel by the total liquid elevation.

[0444] General procedure for examples 9-11

[0445] The steps of the extraction process performed in Examples 9-11 are shown in FIG. 7. Parameters and results for the examples 9-11 are presented in Table 6 and Table 7, respectively. Here, essentially comparable are examples 9 and 10 (effect of biomass type at pH 3), and examples 10 and 11 (effect of pH using the same biomass).

[0446] Example 9

[0447] The objective of this example was to study 1 ) the extraction of algal oil from algal biomass, 2) the decantation efficiency of the first separating bottom layer (raffinate), and 3) the composition of the extracted oil, when an acidic feedstock stream consisting of non-induced biomass and high salt concentration is used. The terms “high salt concentration” and “high salinity” asused herein and hereafter refers to salt concentrations that are at least 10 wt%, at least 15 wt%, at least 20 wt%, or 10 - 24 wt%, such as 20 wt%.

[0448] About 5 wt% dispersion ((acidic) feedstock stream) (706) of noninduced Dunaliella salina biomass in aqueous brine comprising 20 wt% sodium chloride salt, and 0.34 wt% phosphoric acid to adjust the pH of the dispersion from 6.0 to 3.0, was charged to the agitated vessel (a first contacting unit) (708) as the algal feed. Agitation and heating commenced to allow the algal feed to reach 70 °C. The impeller speed was increased to the desired impeller speed setpoint, and the first solvent (710), heptane, was rapidly added to achieve a solvent to feed volumetric ratio of 0.33. The impeller speed setpoint was 140% of the minimum impeller speed [Nmin] to completely disperse two immiscible liquid phases as defined by Skelland and Ramsay (Ind. Eng. Chem. Res. 1987, 26, 1 , 77-81 ). The first liquid-liquid-solid dispersion (first dispersion) (712) was agitated for 20 minutes to facilitate mass transfer, and then the agitation ceased and decantation commenced in the settler (first separating unit) (714). The bottom layer separation took about 30 minutes to achieve 95% of the equilibrium elevation. Once the phase equilibrium was reached (bottom layer interface did not elevate anymore), approximately 58 wt% of the algal feed charged to the mixer (first contacting unit) (708) was removed as the first separated (heavy) liquid phase (716) via the bottom drain valve of the settler (first separating unit) (714) with a loss of about 3.4 wt% of the biomass charged.

[0449] The first remaining dispersion (718) consisting of algal biomass, salt, water, and solvent phase comprising heptane and algal oil (natural products) was fed to a centrifuge (second separating unit) (720) that was operated at 2000 G for 5 minutes at 45-50 °C, and three layers were formed. About 50% of the first light separated liquid phase (722) comprising heptane and algal oil was removed. The second remaining dispersion (724) comprising algal biomass, salt, water, and a solvent phase comprising heptane and algal oil was then placed into the second contacting unit (726). The second solvent (728), in this example heptane, was added to the second contacting unit (726) at a 1 :1 mass ratio with the second remaining dispersion. The resulting dispersion was heated to 45 °C and agitated for 2 minutes in the second contacting unit (726) to intimately mix the phases of the resulting dispersion.

[0450] The resulting third dispersion (730) was fed to a centrifuge (third separating unit) (732) that was operating at 4600 G for 5 minutes and 45-50 °C and three layers were formed: a second light separated liquid phase (734), a biomass-rich phase, and a heavy liquid phase. Essentially all of the second light separated liquid phase (734) comprising heptane and algal oil was removed. The third remaining dispersion (736), containing the biomass-rich phase and the heavy liquid phase, was fed to the third contacting unit (738). The third solvent (740), in this example heptane, was added to the third contacting unit (738) at a 1 :1 mass ratio with the third remaining dispersion (736). The resulting mixture was heated to 45 °C and agitated for 2 minutes in the third contacting unit (738) to fully mix the phases.

[0451] The resulting fourth dispersion (742) was fed to a centrifuge (fourth separating unit) (744) that was operated at 4600 G for 5 minutes at 45-50 °C, and three layers were formed. Essentially all of the third light separated liquid phase (746) comprising heptane and algal oil was removed. The biomass rich phase (748) and / or the second heavy separated liquid phase (750) was / were also separated.

[0452] All of the separated light phases (722, 734, 746) (i.e., the first, the second, and the third light separated liquid phases) were combined, filtered and evaporated to recover algal oil. The total fatty acid yield of the algal oil was 63 wt% based on the total weight of the fatty acids in the algal feed (i.e., the ((acidic) feedstock stream) (706)) charged to the process. The content of common chelatable metals (Mg, Ca, Fe) of the recovered algal oil was 548 mg / kg. Examples 10-11 were performed in the identical manner with minor exceptions mentioned below, and the parameters and the results are shown in Table 6 and Table 7, respectively.

[0453] Example 10

[0454] The objective of this example was to study the effect of biomass type on 1) the extraction of algal oil from algal biomass, 2) the decantation efficiency of the first separating bottom layer (raffinate), and 3) the composition of the extracted oil, when an acidic feedstock stream consisting of induced biomass and high salt concentration is used. This example is essentially comparable with Example 9.

[0455] The algae was induced biomass and the pH of the feed was adjusted with 0.19 wt% phosphoric acid from about 5.6 to 3.0. The impeller speed was 120% of the minimum impeller speed [Nmin], After the agitation was completed, the bottom layer separation took about 20 minutes to achieve 95% of the equilibrium elevation. After the phase equilibrium was reached (bottom layer interface did not elevate anymore), approximately 63 wt% of the algal feed charged to the mixer (first contacting unit) (708) was removed as the first separated (heavy) liquid phase (716) via the bottom drain valve of the settler (first separating unit) (714) with a loss of about 5.0 wt% of the biomass charged. The total fatty acid yield of the algal oil was 80 wt% based on the total weight of the fatty acids in the algal feed (i.e., the ((acidic) feedstock stream) charged to the process. The content of common chelatable metals (Mg, Ca, Fe) of the recovered algal oil was 88 mg / kg.

[0456] Example 11

[0457] The objective of this example was to study the effect of pH on 1 ) the extraction of algal oil from algal biomass, 2) the decantation efficiency of the first separating bottom layer (raffinate), and 3) the composition of the extracted oil, when a non-acidic feedstock stream consisting of induced biomass and high salt concentration is used. This example is comparable with Example 10.

[0458] The algae was induced biomass and the pH of the feed was kept as is (ca. pH 5.6). The impeller speed was 120% of the minimum impeller speed [Nmin], After the agitation was completed, the bottom layer separation took about 20 minutes to achieve 95% of the equilibrium elevation. After the phase equilibrium was reached (bottom layer interface did not elevate anymore), approximately 57 wt% of the algal feed charged to the mixer (first contacting unit) (708) was removed as the first separated (heavy) liquid phase (716) via the bottom drain valve of the settler (first separating unit) (714) with a loss of about 5.1 wt% of the biomass charged. The total fatty acid yield of the algal oil was 61 wt% based on the total weight of the fatty acids in the algal feed (i.e., the ((acidic) feedstock stream) charged to the process. The content of common chelatable metals (Mg, Ca, Fe) of the recovered algal oil was 137 mg / kg.

[0459] Table 6. Parameters for the examples 9-11 *‘General conditions: Feed (F) contains 20 wt% sodium chloride, Solvent (S) is heptane and the S / F mass ratio is 0.2, extraction (708) and decantation (714) temperature was 70 °C, agitation time was 20 min, impeller was HE-3 and located in the middle of the total liquid level (at 0.5 height). Table 6 Column Notations: A = Example number; B = Biomass cultivated in different conditions (non-induced refers to unstressed while induced refers to moderately stressed); C = Weight-% biomass in the Feed to the extractor; D = pH of the feed (after possible pH adjustment by phosphoric acid); E = First contacting unit (708) and first separation unit (714), Temperature in °C; F = Agitation rate in revolutions per minute, rpm; G = Agitation rate relative to the minimum impeller speed of a corresponding pure liquid-liquid system.

[0460] Table 7. Results for the examples 9-11 after the mixer-settler extraction and separation process.‘Comparative experiment. Table 7 Column Notations; A = Example number; B = Time elapsed for 95% bottom layer separation towards phase equilibrium in first separating unit (714); C = Relative Time elapsed for 95% bottom layer separation towards phase equilibrium in first separating unit (714); D = Removed bottom layer (wt% of feed); E = Approximate biomass loss in removed bottom layer (wt%) F = Total fatty acid yield (wt%) to extracted algal oil (measured after hydrolysis of fatty acid esters); G = Approximate Free fatty acid (FFA) content in extracted algal oil (%); H = Common chelatable metal (Mg, Ca, Fe) content in extracted algal oil (mg / kg).

[0461] It will be appreciated by those skilled in the art that the present disclosure can be embodied in other specific forms without departing from the spirit or essential characteristics thereof. The presently disclosed embodiments are therefore considered in all respects to be illustrative and not restricted. The scope of the disclosure is indicated by the appended claims rather than the foregoing description and all changes that come within the meaning and range and equivalence thereof are intended to be embraced therein.

Claims

CLAIMS1 . A process for extracting natural products from a biomass, the process comprising: forming an acidic feedstock stream having a pH of at most 4.0 and containing a biomass and an aqueous salt solution; contacting the acidic feedstock stream with an extraction solvent to form a dispersion; separating the dispersion into multiple layers, the layers including: a solvent extract layer containing at least one hydrophobic natural product and at least a portion of the extraction solvent, a raffinate layer containing at least a portion of the aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer.

2. The process of claim 1 , the forming of the acidic feedstock stream comprising: obtaining a feedstock stream containing the biomass and the aqueous salt solution, the feedstock stream having a pH of more than 4.0 or at least 5.0; and contacting the feedstock stream with an additive stream containing at least one buffering agent and / or at least one acidic additive selected from a group consisting of a Bronsted acid, an acidic chelating agent and a combination thereof.

3. The process of claim 2, comprising: contacting the feedstock stream with the additive stream in a mixing zone, the mixing zone optionally including at least one of a static mixer, an in-line mixer, an eductor, a transpiring wall, a diffuser, a mixing-T piece, a turbulent flow, an agitated vessel, an extraction column, or a combination thereof.

4. The process of claim 2, comprising: contacting the feedstock stream with the additive stream in an extraction zone.

5. The process of any one of claims 2 to 4, wherein the additive stream comprises: at least one Bronsted acid selected from a group consisting of hydrochloric acid, nitric acid, sulfuric acid, formic acid, propionic acid, glyoxylic acid, acetic acid and mono- and polyprotic acids including but not limited to citric acid, phosphoric acid, glycolic acid, 2-ethylhexanoic acid, malic acid, malonic acid, 2,3-dihydroxybenzoic acid, gluconic acid, tartaric acid, carnosic acid, ethylenediaminetetraacetic acid, 2-ethylhexanoic acid and oxalic acid.

6. The process of any one of claims 2 to 5, wherein the additive stream comprises: at least one acidic chelating agent selected from a group consisting of citric acid, phosphoric acid, glycolic acid, 2-ethylhexanoic acid, malic acid, malonic acid, 2,3-dihydroxybenzoic acid, gluconic acid, tartaric acid,carnosic acid, ethylenediaminetetraacetic acid, 2-ethylhexanoic acid and oxalic acid.

7. The process of any one of claims 2 to 6, wherein the additive stream comprises: at least one buffering agent selected from a group consisting of ammonium chloride, ammonium acetate, ammonium phosphate, ammonium sulfate, ammonium citrate, sodium chloride, sodium acetate, sodium phosphate, sodium sulfate, sodium citrate, sodium tartrate, sodium malonate, sodium gluconate, sodium formate, sodium propionate, potassium chloride, potassium acetate, potassium phosphate, potassium sulfate, potassium citrate, potassium tartrate, potassium malonate, potassium gluconate, potassium formate, potassium propionate, and any equivalent magnesium or calcium salts.

8. The process of any one of the previous claims, wherein the acidic feedstock stream has a pH of at most 3.0.

9. The process of any one of the previous claims, wherein the biomass is or comprises: at least one or more of a plant biomass, a microbial biomass, an algal biomass and / or any combination thereof.

10. The process of any one of the previous claims, wherein the extraction solvent is or comprises: at least one or more of a polar solvent, a non-polar solvent, a nonpolar organic solvent, a dense gas solvent, an aqueous two-phase solvent, adeep eutectic solvent (DES), a natural deep eutectic solvent (NADES), an ionic liquid, or any combination thereof.11 . The process of any one of claims 4 to 10, wherein the extraction zone comprises: at least one or more of a mixer, a static mixer, a settler, a cocurrent extraction column, a counter-current extraction column, a centrifugal extractor, an emulsion phase contactor, or any combination thereof.

12. The process of any one of claims 1 to 11 , wherein the contacting of the acidic feedstock stream with the extraction solvent occurs in an / the extraction zone, the extraction zone being a counter-current extraction column, and the process optionally comprising: retaining the dispersion in the counter-current extraction column for a residence time of about 2 minutes to about 2 hours.

13. The process of any one of the previous claims, wherein the separating of the dispersion into multiple layers comprises: at least one or more of gravity settling, centrifugal settling, and / or combinations thereof.

14. The process of any one of the previous claims, comprising: performing the separating of the dispersion into multiple layers by gravity settling under a gravitational field.

15. The process of any one of the previous claims, comprising: performing the separating of the dispersion into multiple layers by decanting.

16. The process of any one of the previous claims, wherein the at least one hydrophobic natural product is or comprises: at least one lipid, algal lipid, carotenoid, fatty acid, algal fatty acid, triacylglycerol, diacylglycerol, monoacylglycerol, oil, algal oil, sterol, wax or any combination thereof.

17. The process of any one of the previous claims, wherein the aqueous salt solution has a salinity of at least one or more of greater than 0.1 wt%, greater than 5 wt%, greater than 10 wt%, greater than 15 wt% and / or greater than 20 wt%.

18. The process of any one of the previous claims, wherein the aqueous salt solution is or comprises: culture medium of the biomass.

19. The process of any one of the previous claims, wherein the lipid- depleted biomass comprises: at least one or more of chlorophyll, glycerol, phospholipids, carbohydrates, fibers, proteins or combinations thereof.

20. The process of any one of the previous claims, comprising:performing the process as a continuous process, wherein the forming of the acidic feedstock stream, the contacting of the acidic feedstock stream with the extraction solvent, the separating of the dispersion into multiple layers, and the recovering of at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer are performed sequentially.21 . The process of any one of the previous claims, comprising: recovering at least a portion of the solvent extract layer.

22. The process of any one of the previous claims, the separating of the dispersion comprising: using a coalescer to assist in the separating of the dispersion into multiple layers.

23. The process of any one of the previous claims, comprising: filtering the recovered solvent extract layer after isolation to remove any entrained biomass; filtering the recovered raffinate layer after isolation to remove any entrained biomass; and / or filtering the recovered rag layer after isolation to remove any entrained solvent or any entrained aqueous salt solution.

24. The process of any one of the previous claims, comprising:evaporating the extraction solvent from the recovered solvent extract layer after recovery of the solvent extract layer.

25. The process of any one of the previous claims, comprising: allowing the acidic feedstock stream to contact the extraction solvent for about 1 minute to about 5 hours.

26. The process of any one of the previous claims, comprising: performing the separating of the dispersion for at least 5 minutes to about 24 hours.

27. The process of any one of the previous claims, comprising: performing the separating of the dispersion at a pressure ranging from atmospheric to supercritical conditions for the extraction solvent.

28. The process of any one of the previous claims, comprising: recovering the rag layer and recovering the lipid-depleted biomass from the rag layer.

29. The process of claim 28, comprising: removing any residual extraction solvent, salts and / or clay from the isolated lipid-depleted biomass.

30. The process of claim 28 or 29, comprising: pelletizing the isolated lipid-depleted biomass.31 . The process of any one of the previous claims, comprising: recovering the raffinate layer; and removing any entrained biomass, any entrained extraction solvent and / or any soluble solvent from the recovered raffinate layer.

32. The process of any one of the previous claims, wherein the acidic feedstock stream contains a water content before extracting.

33. The process any one of the previous claims, comprising: performing a heat exchange before the acidic feedstock stream contacts the extraction solvent and / or before the separating of the dispersion.

34. The process of any one of the previous claims, wherein the biomass is or comprises: conditioned biomass.

35. The process of any one of the previous claims, wherein the rag layer is formed between the solvent extract layer and the raffinate layer.

36. The process of any one of the previous claims, wherein the separating of the dispersion into multiple layers occurs in a temperature regulated environment.

37. The process of any one of the previous claims, wherein the raffinate layer has a salinity of at least one or more of about 0.1 wt% or greater, about 5 wt% or greater, about 10 wt% or greater, about 15 wt% or greater, and / or about 20 wt% or greater.

38. The process of any one of the previous claims, wherein the rag layer contains at most about 60 wt% solvent extract layer (such as from 30 wt% to 60 wt%, e.g. 40 wt% to 50 wt% or about 45 wt%), at most about 60 wt% raffinate layer (such as from 30 wt% to 60 wt%, e.g. 40 wt% to 50 wt% or about 45 wt%) and / or about from 2 wt% to 20 wt% lipid-depleted biomass (e.g. from 5 wt% to 10 wt%).

39. The process of any one of the previous claims, wherein the raffinate layer contains salt from at least one or more of about 0.1 wt% to about 50 wt%, from about 5 wt% to about 40 wt%, from about 10 wt% to 30 wt%, and / or from about 15 wt% to 26 wt% of the total weight of the raffinate layer.

40. The process of any one of the previous claims, wherein the separating of the dispersion into multiple layers and / or the contacting of the acidic feedstock stream with the extraction solvent occurs at a temperature of at least one or more of about 100°C, less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°C to 90°C, 30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C and / or 40°C to 70°C.41 . A process for extracting natural products from a biomass, the process comprising: obtaining a feedstock stream containing the biomass and an aqueous salt solution, the feedstock stream having a pH of more than 4.0 or at least 5.0; transferring the feedstock stream to an extraction zone; introducing an additive stream containing an extraction solvent and at least one acidic additive and / or a buffering agent into the extraction zone to form a dispersion, the least one acidic additive being selected from a group consisting of a Bronsted acid, an acidic chelating agent and a combination thereof, wherein the dispersion contains an aqueous phase having a pH of at most 4.0; separating the dispersion into multiple layers, the layers including: a solvent extract layer containing at least one hydrophobic natural product and at least a portion of the extraction solvent, a raffinate layer containing at least a portion of the aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass; and recovering at least part of the solvent extract layer, at least part of the raffinate layer and / or at least part of the rag layer.

42. A system for extracting natural products from a biomass, the system comprising: a mixing zone configured to mix a feedstock stream with an additive stream, and contain an acidic feedstock stream having a pH of at most4.0;an extraction zone configured to mix an acidic feedstock stream having a pH of at most 4.0 with an extraction solvent, and contain a dispersion having an aqueous phase with a pH of at most 4.0; and a separation zone configured to separate a dispersion having an aqueous phase with a pH of at most 4.0 into a solvent extract layer containing at least one hydrophobic natural product and at least a portion of an extraction solvent, a raffinate layer containing at least a portion of an aqueous salt solution, and a rag layer containing at least a portion of a lipid-depleted biomass.

43. Use of the system of claim 42 for extracting natural products from a biomass and / or for separating a dispersion into multiple layers, the layers including: a solvent extract layer, a raffinate layer and a rag layer.

44. A process for extracting natural products from a biomass, the process comprising: forming an acidic feedstock stream having a pH of at most 4.0 and containing a biomass and an aqueous salt solution; contacting the acidic feedstock stream with a first solvent in a first contacting unit to form a first dispersion; allowing the first dispersion to phase separate into a first separated liquid phase and a first remaining dispersion; removing at least a portion of the first separated liquid phase; contacting the first remaining dispersion with a second solvent in a second contacting unit to form a second dispersion;allowing the second dispersion to phase separate into a biomassrich phase, a heavy separated liquid phase and a light separated liquid phase; and recovering at least one or more of the biomass-rich phase, the heavy separated liquid phase, and / or the light separated liquid phase for output as products.

45. A system for extracting natural products from a biomass, the system comprising: an acidic feedstock stream import line; a first contacting unit in communication with the acidic feedstock stream import line; a first solvent import line in communication with the first contacting unit; a first effluent export line in communication with the first contacting unit; a second contacting unit in communication with the first contacting unit; a second solvent import line in communication with the second contacting unit; and a first extraction line in communication with the second contacting unit.

46. The process for extracting natural products from a biomass as claimed in any of claims 1 - 41 and 44, wherein the process comprises:contacting the acidic feedstock stream with an extraction solvent in a contacting unit to form a dispersion; and separating the dispersion into multiple layers in a separating unit.

47. The process for extracting natural products from a biomass as claimed in any of claims 1 - 41 , and 44, wherein both the contacting the acidic feedstock stream with an extraction solvent to form a dispersion, and the separating the dispersion into multiple layers are carried out in a contacting unit or a separating unit.

48. The process for extracting natural products from a biomass as claimed in any of claims 1 - 41 , 44, 46, and 47, wherein the process comprises: providing an acidic feedstock stream having a pH of at most 4.0, such as a pH in the range from 3.0 to 4.0, and containing a biomass and an aqueous salt solution, optionally further containing an additive stream containing at least one buffering agent and / or at least one acidic additive selected from a group consisting of a Bronsted acid, an acidic chelating agent and a combination thereof; contacting the acidic feedstock stream with a first solvent in a first contacting unit, to form a first dispersion; allowing the first dispersion to phase separate in a first separating unit into a first separated liquid phase, such as a first separated (heavy) liquid phase, and a first remaining dispersion; removing at least a portion of the first separated liquid phase, such as the first separated (heavy) liquid phase;allowing the first remaining dispersion to phase separate into a first light separated liquid phase, and a second remaining dispersion in a second separating unit; removing at least a portion of the first light separated liquid phase; contacting the second remaining dispersion with a second solvent in a second contacting unit to form a third dispersion; allowing the third dispersion to phase separate into a second light separated liquid phase, and a third remaining dispersion in a third separating unit; removing at least a portion of the second light separated liquid phase; contacting the third remaining dispersion with a third solvent in a third contacting unit to form a fourth dispersion; allowing the fourth dispersion to phase separate into a biomass-rich phase, a second heavy separated liquid phase and a third light separated liquid phase in a fourth separating unit; and recovering at least one or more of at least a part of the biomass-rich phase, at least a part of one or more of the first and the second separated (heavy) liquid phase, and / or at least a part of one or more of the first, the second, and the third light separated liquid phase for output as products.

49. The process for extracting natural products from a biomass as claimed in claim 48, wherein the contacting the acidic feedstock stream with a first solvent in a first contacting unit to form a first dispersion comprises agitating the first dispersion in a vessel comprising an impeller, preferably the impeller speed of the impeller is a predetermined impeller value being 110 - 140% of theminimum impeller speed (Nmin) of the impeller, such as 120% or 140% of the minimum impeller speed (Nmin) of the impeller, to disperse two immiscible liquid phases, and heating the first dispersion at a temperature of at least one or more of about 100°C, less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°C to 90°C, 30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C and / or 40°C to 70°C.

50. The process for extracting natural products from a biomass as claimed in any of claims 48 - 49, wherein the allowing the first remaining dispersion to phase separate into a first light separated liquid phase, and a second remaining dispersion in a second separating unit is performed in a centrifuge at 1500 - 15000 G, such as at 2000 G, optionally for 0.5 - 10 min, such as at 5 minutes, at 5 - 90 °C, or at 35 - 60 °C, such as at 45 - 50 °C.51 . The process for extracting natural products from a biomass as claimed in any of claims 1 - 50, wherein the extraction solvent, the first solvent, the second solvent, and / or the third solvent is / are a hydrocarbon such as hexane, heptane, or octane or any combination thereof.

52. The process for extracting natural products from a biomass as claimed in any of claims 1 - 51 , wherein the dispersion, the first dispersion, the first remaining dispersion, the second remaining dispersion, the third dispersion, the third remaining dispersion, and / or the fourth dispersion are each independently heated at a temperature of at least one or more of about 100°C,less than about 100°C, of about 10°C to 90°C, 10°C to 80°C, 20°C to 90°C, 20°C to 80°C, 30°C to 90°C, 30°C to 80°C, 40°C to 90°C, 40°C to 80°C, 50°C to 90°C, 50°C to 80°C, 60°C to 90°C, 60°C to 80°C, 35°C to 80°C, 35°C to 70°C and / or 40°C to 70°C.

53. The process for extracting natural products from a biomass as claimed in any of claims 1 - 52, wherein recovering one or more of the at least part of the solvent extract layer, the at least part of the first light separated liquid phase, the at least part of the second light separated liquid phase, and the at least part of the third light separated liquid phase for output as products; combining one or more of the recovered at least part of the solvent extract layer, at least part of the first light separated liquid phase, at least part of the second light separated liquid phase, and at least part of the third light separated liquid phase to form a combined phase of light separated liquid phases; filtering the combined phase of light separated liquid phases; evaporating the filtered combined phase of light separated liquid phases; and recovering algal oil for output as product.

54. A system for extracting natural products from a biomass, the system comprising: an acidic feedstock stream import line; three contacting units consisting of a first, a second, and a third contacting unit;three solvent import lines consisting of a first, a second, and a third solvent import line; three effluent export lines consisting of a first, a second, and a third effluent export line; four separating units consisting of a first, a second, a third, and a fourth separating unit; and three extraction lines consisting of a first, a second, and a third extraction line; wherein the first contacting unit being in communication with the acidic feedstock stream import line, the first solvent import line, and the first separating unit; the first separating unit being in communication with the first effluent export line, and the second separating unit, the second separating unit being in communication with the second effluent export line, and the second contacting unit, the second contacting unit being in communication with the second solvent import line, and the third separating unit; the third separating unit being in communication with the third effluent export line, and the third contacting unit, the third contacting unit being in communication with the third solvent import line, and the fourth separating unit; and the fourth separating unit being in communication with the first, the second, and the third extraction line.

55. The system as claimed in claim 54, wherein the first effluent export line is configured to remove a first separated (heavy) liquid phase from the first separating unit, the second effluent export line is configured to remove a first light separated liquid phase from the second separating unit, the third effluent export line is configured to remove a second light separated liquid phase from the third separating unit, the first extraction line is configured to remove a third light separated liquid phase from the fourth separating unit; the second extraction line is configured to remove a biomass-rich phase from the fourth separating unit; and the third extraction line is configured to remove a second heavy separated liquid phase from the fourth separating unit.

56. The system as claimed in any of claims 54 - 55, wherein the system further comprises: a mixing zone in communication with the first contacting unit; a feedstock stream import line in communication with the mixing zone; and an additive stream import line in communication with the mixing zone.

57. Use of the system of any of claims 45 and 54 - 56 for extracting natural products from a biomass and / or for separating a dispersion into multiple layers, the layers including: a solvent extract layer, a raffinate layer and a rag layer.

Images