Process and installation for the enzymatic synthesis of biodiesel from used lipids

Abstract

The present invention relates to a process for synthesizing a biodiesel composed of a mixture of ethyl and / or methyl esters from used lipids originating from a used non-acidic vegetable oil, an acidic used oil and / or a used animal fat. The invention also relates to a plant for implementing the process of the invention [Fig. 1]

Description

Title of the invention: Method and installation for the enzymatic synthesis of biodiesel from used lipids Subject of the invention

[0001] The present invention relates to the field of enzymatic synthesis and concerns a process for synthesizing a biodiesel composed of a mixture of ethyl and / or methyl esters from used lipids originating from a used non-acidic vegetable oil, an acidic used oil and / or a used animal fat. The invention also concerns an installation for implementing the process of the invention. State of the art

[0002] Currently, only vegetable oil methyl esters (VEMEs) and animal fats are authorized in Europe and used as biofuel, in particular as biodiesel, by mixing them with diesel at a maximum of 30%. However, these vegetable oil methyl esters, like VMEs from used vegetable oils, must comply with the DIN EN14214 standard which governs all the physicochemical parameters of a biodiesel, except for the quantities of residual alcohol, which is only defined for methanol.

[0003] But with legislation evolving rapidly, vegetable oil ethyl esters (VEEOs) are in the process of being authorized by customs for sale and use if a declaration is made to this effect. However, DIN EN14214 is not suitable for measuring quantities of used vegetable oil ethyl esters due to the standard used. Aims of the invention

[0004] The present invention aims to obtain an eco-clean, simple, efficient, inexpensive and applicable design process from locally available lipid sources to produce biodiesel from the lipids of normal used vegetable oil, acidic used oil and / or used animal fat.

[0005] Furthermore, the invention aims to obtain such a process which does not include any step for chemically pretreating the raw material and for washing the biodiesel obtained in order to simplify the process and the apparatus and reduce costs. Summary of the invention

[0006] A first aspect of the invention relates to a method and an installation for obtaining a biodiesel consisting of a mixture of ethyl esters or methyl esters, by an enzymatic transformation of lipids, chosen from the group consisting of used vegetable oil, acidic vegetable oil and / or used animal fat, in which said enzymatic transformation preferably by an enzyme of the type triacylglycerol acyl hydrolase and which comprises an enzymatic transesterification and an enzymatic hydroesterification an addition of alcohol selected from the group consisting of ethanol, methanol or a mixture of these alcohols and an addition of water or a base. In the process according to the invention, the enzymatic transformation also comprises an addition of Glycerol.

[0007] Preferably, the method of the invention also has one or more of the following characteristics:

[0008] The enzyme is preferably selected from the group consisting of the enzymes Novozym ® 40116, Novozym ® 40086, Novozym ® 435, Lipozyme ® RM IM, Lipozyme ® TL IM, CalB Sigma ®, CalB Immoplus ®, NovoLhne ®, TL Sigma ®, Callera TM, Eversa ®, Pallatase ®, Eversa ® Transform 2.0 free in liquid form from Thermomyces lanuginosus, or a mixture of two or more of these enzymes.

[0009] The lipids are subjected, prior to the enzymatic transformation into biodiesel, to one or more stage(s) of filtration of the solid particles present in the lipids.

[0010] Lipids consisting of used palm oil or used animal fat are heated to a temperature of 40°C or more, preferably 45°C or more before the filtration step(s).

[0011] The lipids are subjected, prior to the enzymatic transformation into biodiesel, to one or more water removal steps, and to obtain lipids containing a water content of less than 1% by weight relative to the weight of the lipids.

[0012] The lipids of the used vegetable oil and / or the acidic vegetable oil are subjected, prior to the enzymatic transformation into biodiesel, to one or more stages of filtration and collection of a vegetable oil having a turbidity or cloudiness in the liquid close to 0.

[0013] Lipids are animal fats which are subjected, prior to enzymatic transformation into biodiesel, to a decantation step at room temperature for used oils and at 45°C or more for animal fats for a period of between 48 hours and 72 hours.

[0014] The alcohol is added at the molar ratio (alcohol:oil) ranging from (3:1) to (16:1), the lipids, water or base are added at the percentage of minus 5% and the glycerol is added at the percentage ranging from 1% to 10% and in which this medium is brought into contact with the enzyme for a period of between 30 minutes and 2 hours to form a reaction medium.

[0015] The alcohol is added to the reaction medium sequentially from 1 to 4 times, after a period of between 15 minutes and 4 hours, preferably after a period of between 30 minutes and two hours.

[0016] The method comprises a step of recycling the used enzyme, preferably with a step of reusing the free enzyme, a step of immobilizing the used enzyme, a step of collecting the used immobilized enzyme, a step of adding hexane to the collected enzyme and a step of drying the collected enzyme.

[0017] The process comprises the enzymatic transformation of used lipids into biodiesel, a step of refining the reaction medium including the separation of the alcohol, glycerol or glycerin formed during the enzymatic transformation, and the aqueous part of the reaction medium from the biodiesel obtained.

[0018] The process includes a refining step which comprises evaporation of the alcohol from the reaction medium, and optionally collection by condensation of the evaporated alcohol and / or the addition of the recycled alcohol in the process for obtaining biodiesel according to the invention.

[0019] The refining step comprises decantation of the glycerol and / or the aqueous part of the reaction medium.

[0020] The aqueous part of the reaction medium incorporating the enzyme is added with alcohol and used lipids to obtain the biodiesel according to the invention.

[0021] The method comprises after the refining step, a step of collecting the refined biodiesel, followed by a step of adding an antioxidant, preferably butylated hydroxytoluene, to the refined and collected biodiesel, and / or a step of mixing the collected refined biodiesel with another fuel to obtain a final 30% biodiesel (B30).

[0022] The installation for obtaining biodiesel by implementing the method according to the invention comprises:

[0023] a lipid pretreatment module comprising means for collecting and filtering lipids from a non-acidic used vegetable oil, an acidic used vegetable oil and / or a used animal fat, said module preferably consists of one or more filter grids whose opening size is between 3 mm and 1 mm, and possibly a de-oiler, capable of collecting pre-treated lipids, said lipid pretreatment module being connected to

[0024] a reaction module comprising a reactor, a device for heating the reactor, a device for stirring the reagents present in the reactor and means for adding pretreated lipids and other reagents to the reactor, in particular means for adding and possibly recycling enzymes, means for adding water and / or a base and means for adding one or more alcohols, said reactor being connected to

[0025] a module for collecting and refining a mixture of the biodiesel obtained and the reactants present in the reaction medium from the reactor, said collection module and refining comprising means for evaporating the surplus alcohol and possibly means for recycling said alcohol and the enzymes present in the reaction medium; means for collecting by decantation the refined biodiesel, the glycerol and the aqueous part of the reaction medium and possibly means for recycling the glycerol and the aqueous part of the reaction medium, said collection and refining module being possibly connected to

[0026] a module for processing and collecting the refined biodiesel obtained comprising a tank and means for adding an antioxidant to the tank, this tank being connected to a tank for mixing the refined biodiesel obtained with another fuel, preferably diesel.

[0027] The present invention will be described in detail in the examples below with reference to the appended figures and presented by way of illustration of a preferred and non-limiting embodiment of the invention. Brief description of the figures

[0028] [Fig.l] represents the steps of enzymatic treatment of used lipids according to the invention. Detailed description of the invention

[0029] The present invention relates to a process and an installation for synthesizing a biodiesel, from so-called used lipids, this biodiesel being defined as a mixture of ethyl or methyl esters originating from used vegetable oils (EEHVU and EMHVU), acid oils and / or used animal fats.

[0030] The method and installation of the invention essentially comprise several of the modules or steps described below:

[0031] l. A first module or a first step, called pretreatment of lipids (forming the incoming materials to be treated) by filtration and / or by decantation, corresponds to module or step 1, followed by

[0032] 2. a second module or a second step called reaction corresponds to the module or to step 2, this reaction comprises via suitable means present in the module to obtain a trans-esterification (by alcoholysis) and a hydroesterification (by hydrolysis and by esterification) of the lipids, with conditions adapted to each type of oil (used vegetable oils, acid vegetable oils and / or animal fats), followed (optionally) by

[0033] 3. a third module or a third step called enzyme recovery used in the second step, correspond to the module or step 3, followed by

[0034] 4. a fourth module or a fourth step called refining of a biodiesel obtained in module or in step 2 or 3, corresponds to module or in step 4, followed (optionally) by

[0035] 5. a fifth module or a fifth stage called mixing and collection refined biodiesel obtained in the fourth step with another fuel (diesel) corresponds to module or step 5.

[0036] In the method and installation of the invention, three types of so-called used lipids (used as input materials) can be treated for the preparation of the biodiesel of the invention.

[0037] A first type of lipid is obtained from a used vegetable oil (UVO) which corresponds to an oil resulting from cooking in oil of vegetable or camel foods, such as potatoes (fries), bananas, frying batters known as doughnut and churro batters or meats, preferably a used oil chosen from the group consisting of palm oil, sunflower oil and / or rapeseed oil. A used vegetable oil has undergone deterioration due to cooking causing the appearance of polar compounds, that is to say compounds not present in the initial oil and which are therefore degraded products and / or different polymers such as starch, as well as an increase in the level of free fatty acids, this level however not exceeding 5%. Additionally, as the cooking temperature induces the polymerization reaction, the viscosity of the vegetable oil increases due to the polymers formed, compared to unheated vegetable oil.

[0038] A second type of lipid is that of an acidic vegetable oil which comes from industrial sites or wastewater treatment plants. The main characteristic of an acidic oil is that it contains a high level of free fatty acids, this level varying from 5% to 95% for a very degraded acidic vegetable oil. Their very heterogeneous origin makes them a complex mixture to treat, particularly given the presence of water, solid residues and polluting compounds.

[0039] A third type of lipid is obtained from used animal fat which is used in catering for frying or cooking vegetable or meat foods, in particular potatoes (fries) or meats. Used animal fat has a different profile from used vegetable oil, distinguished in particular by the presence of sterols of animal origin such as cholesterol. In addition, like palm oil, animal fat is solid at room temperature, but the level of free fatty acids does not exceed 5%. The module and preprocessing step 1

[0040] Module or step 1 comprises the pretreatment of the lipids forming the input materials of the process of the invention. This pretreatment allows the elimination in the lipids of solid residues or particles, polymeric compounds and the majority of the residual water in the used vegetable oil, acidic vegetable oil and / or used fat, to arrive at a level of water present in the lipids of between 300 ppm and 1000 ppm.

[0041] In this first module and this first pretreatment step 1 which precedes the enzymatic treatment, the used lipids are all filtered at room temperature. Since palm oil and used food fats are solid at room temperature, they must be heated to 40°C or more to remain liquid during filtration and before being subjected to the same conditions as those described below in the pretreatment and the enzymatic treatment process. The decantation phase is carried out without additives at a temperature between 40°C and 50°C, preferably at approximately 45°C, for a period of between one day and three days, and the material transfers (by pumping) are carried out at the same temperature. The installation will include suitable heating means to obtain and maintain this temperature.

[0042] The filtration pretreatment step may be carried out by passing the melted waste oils or animal fats over one or more grids. Preferably, the diameter of the openings or size of the openings of the grids is largest for the first grid and decreases with each subsequent grid. A first grid has openings, such as holes, having a diameter of between 2 mm and 4 mm, preferably about 3 mm. Preferably, a second grid with openings, such as holes, will have a diameter or size of the openings of between 0.5 mm and 2 mm, preferably about 1 mm.

[0043] This preliminary filtration makes it possible to eliminate particles consisting of the largest residues (food and non-food), such as pieces of plants, meat, fish bones or pieces of paper.

[0044] Next, the oil obtained preferably passes through a de-oiler, which plays an inverse role and allows the elimination of water and a settling of the fats from the treated lipids. During this operation, the oil passes through the multi-compartment de-oiler at a flow rate preferably between 2 m3 / hour and 5 m3 / hour. The water is regularly removed either by manual valves or by an automatic valve which detects excess water. The reduced water content, following this operation, of the treated oil is preferably less than 1%.

[0045] The resulting water-reduced oil or animal fat is conveyed, preferably via several filters 12 having openings of 100 μm to 5 μm, to a storage tank 13 in order to collect water-reduced animal fat or oil having a turbidity close to 0. Turbidity is the measure of the cloudiness in a liquid measured by a Metler Toledo ® InFit turbidimeter. The reaction module or step 2

[0046] In the method and installation of the invention, the used lipids present in the storage tank are optionally subjected to the first decantation before being added to a reactor to undergo an enzymatic transformation. The reactor includes means for adding different reagents (used lipids, enzymes, water, base, alcohol, hexane, etc.) useful for the reaction and possibly for collecting the products obtained or recycled products (alcohol, glycerol, biofuel, enzymes, etc.) to treatment tanks or modules.

[0047] A. 1. Operating conditions for the treatment of a non-acidic used vegetable oil

[0048] The transformation of a non-acidic used vegetable oil into biodiesel consists of hydrolyzing-esterifying / trans-esterifying by enzymatic reaction the different glycerides (triglycerides, diglycerides and monoglycerides) present in the used non-acidic vegetable oil, via enzymatic catalysis. In the process of the invention, the efficiency of the enzyme was increased by preparing a homogeneous medium between the used oil and the added water and then reacting the enzyme with the used oil before adding the alcohol and triggering the transesterification. These two steps improved the efficiency of the enzyme which made it possible to reduce the amount of enzyme required to complete the reaction.The efficiency of the enzyme is also improved by adding a low concentration base and / or glycerol which could reduce the amount of enzyme needed to carry out this process and therefore the reaction time. In addition, the preferred enzyme in this process is present in liquid form, is not expensive and could advantageously be reused twice and which does not pose problems related to the type of mechanical agitation like immobilized enzymes.

[0049] The enzymatic reaction was optimized with the addition of alcohol, preferably ethanol or methanol).

[0050] A.2. Addition of the enzyme

[0051] In order to carry out the enzymatic transformation of used vegetable oil into biodiesel, the enzymes used are lipases, with their scientific name triacylglycerol acyl hydrolase (EC3.1.1.3) of bacterial origin or extracted from fungi.

[0052] Several commercial lipases were tested and the reaction proved effective with several of them such as lipases from the strains Thermomyces lanuginosus, Pseudomonas cepacia, Pseudomonas aeruginosa, Candida Antarctica, Candida Rugosa, Rhizomucor miehei or Aspergilus niger.

[0053] Enzymes can be used in three different states: immobilized, non-immobilized or lyophilized. Immobilizing the enzyme on a solid support allows for easier recovery of the enzyme and improved stability and thermal resistance, but generally increases the cost of the latter, offset by the reuse of the enzyme, as well as the possibility of damaging the support of these enzymes by mechanical agitation.

[0054] On the other hand, if the free enzymes are not very contagious and their recovery is difficult and their stability low, but it is also possible to use again in the same reactor the said enzyme present in the residual aqueous portion of the reaction.

[0055] The three states were tested for different enzymes. In particular, the three states were tested for enzymes from Thermomyces lanuginosus, Candida rugosa, Candida antartica, and Rhizmucor miehei.

[0056] Some tests were also carried out with non-immobilized enzymes, including Eversa ®, Callera TM Trans L, Novozym @40116, Lipozyme ® TL IM or Pallatase ®. Some tests were also carried out with lyophilized enzymes (in particular the enzyme from Pseudomonas cepacian and Porcine pancreas). Among the enzymes that showed high efficiency in the transformation of normal or acidic waste oil and animal fat into biodiesel, the commercial enzyme "Eversa ® Transform 2.0" achieved the best yield. Using this enzyme, the percentage of esters in the biodiesel reached more than 96%. Eversa ® Transform 2.0 used is free in liquid form from Thermomyces lanuginosus.

[0057] The amount of enzyme has a very important influence on the efficiency of the reaction. If too little is used, the reaction will be too slow and will not take place on a profitable time scale. If too much enzyme is used, the enzyme will have self-inhibited itself by capturing too many reactants without continuing the reaction.

[0058] Advantageously, an optimal quantity combined with all the other parameters allows the reaction to be carried out over a period of less than 24 hours.

[0059] The test range for which the experiments were feasible extends from (approximately) 1% (v / v) to (approximately) 10% (v / v) for free enzymes, from 0.5% (w / v) to 5% (w / v) for immobilized and freeze-dried enzymes. By optimizing the amount of water, the amount of alcohol, the addition of additive (sodium carbonate or sodium hydroxide) and the method of preparing the reaction medium before adding the enzyme... the amount of enzyme added can be reduced to 1% of free enzyme and 0.5% of immobilized enzyme with a yield of approximately 97% of esters in the biodiesel.

[0060] A.3. Addition of alcohol

[0061] Alcohol serves as a reagent for hydroesterification / transesterification (acyl acceptor) and also as a solvent to improve the solubility of the oil and thus reduce its viscosity.

[0062] The reaction between oil and alcohol in the presence of the enzyme is explained by the following diagram:

[0063] [Chem.l] Enzyme * ÇH^OH ÇH-OH GHrOH Glycerol R5COOCH3 + R2COOCH3 R^COOCHj Methyl esters ÇHrCOOR5 ÇH-COOR2 + CHj-COOR3 Triglyceride 3GHjOH Methanol CH,-COOR! CH^-OH RfCOOC,H, ÔH-COOR2 + 3C2HsOH Enzyme ÇH-OH + R2COOC2Hs ÔH2-COOR3 Çh2-oh r3cooc2h5 Triglyceride Ethanoi Glycerol Ethyl esters

[0064] The addition of several short carbon chain alcohols (methanol, ethanol and propanol) was tested.

[0065] Ethanol is chosen to obtain an optimal transformation of oils into biodiesel, because this alcohol is not very toxic and can be obtained via a process of treating a renewable biological material, for example via biomass fermentation. Even if biodiesel from ethyl esters is not yet authorized like biodiesel from methyl esters, the process has also been tested in the presence of ethanol, because ethanol can also be obtained via a process of treating a renewable biological material.

[0066] Different ratios (alcohol / oil) from (about) 3:1 (mol / mol) to (about) 16:1 (mol / mol) were tested. Since triglyceride contains three fatty acids, the amount of alcohol must be three times higher than that of triglyceride. Therefore, the molar ratio (3:1) represents the minimum ratio to carry out this operation. The excess of alcohol is essential to direct the reaction towards the synthesis of esters provided that this excess does not cause inhibition of the enzyme. In this study, the molar ratio (16:1) represents the maximum ratio that does not cause any enzyme inhibition. Consequently, the molar ratio range from (3:1) to (16:1) achieved a percentage of esters in biodiesel between 80% and 97%.

[0067] Like other chemical reagents, some alcohols can distort the normal three-dimensional conformation of enzyme proteins, which will lose their activity. This effect depends on the nature and state of the enzyme. To avoid this effect, the alcohol was preferably added sequentially during the reaction, preferably before adding the enzyme, to reduce direct contact between a high concentration of the alcohol and the enzyme. For this, the addition of alcohol was tested from a single addition to four or more additions after different time intervals. ranging from about 30 minutes to about 2 or 3 hours. Indeed, up to the molar ratio (6:1) (ethanol:oil), ethanol is added in one go at the beginning of the reaction before adding the enzyme without any enzymatic inhibition while ethanol is added in several times from the molar ratio (7:1) to avoid enzymatic inhibition, because at this ratio, the enzyme has lost about 15% of its activity, when ethanol is added in one go. For methanol, the addition is carried out in several times from the molar ratio (3:1), because methanol is more inhibiting than ethanol for the enzyme, its carbon chain being shorter, it can block the active site of enzyme.

[0068] A.4. Addition of water

[0069] The water present ensures the stability of the enzyme activity and the course of the reaction, because the aqueous layer around the enzyme helps to maintain the normal three-dimensional conformation of the enzyme proteins. The activity of water is specific to each enzyme (type, nature). Here, like other parameters, the amount of water has been optimized to achieve the best enzymatic activity and the best yield. Even without water, the reaction takes place, but it has extremely slow kinetics. A large amount of water leads the reaction towards hydrolysis, in turn reducing the synthesis of esters and therefore the final yield of biodiesel. Beyond a certain amount of water, the reaction no longer takes place, the water inhibiting the synthesis reaction and leading it 100% towards hydrolysis.The optimal reaction takes place for water contents between 0% (v / v) without water (but without water with preferably an addition of glycerol) with an ester percentage of about 65% benefiting from the water generated by the esterification reaction and 5% (v / v) water with an ester percentage of about 55% due to the hydrolysis induced by the high amount of water in the reaction medium. Between 0% and 5% water, a maximum percentage of esters reached 97%. To avoid hydrolysis caused by the added water and the generated water, glycerol is added at the beginning of the treatment, preferably before the addition of the enzyme in the reaction medium to reduce the amount of water added and create hydrophilic microdroplets where the enzyme works. The percentage of glycerol ranging from 1% to 20% is tested in the absence and presence of small amounts of water.In general, the addition of glycerol improves the yield for the free enzyme by reducing the possibility of hydrolysis, especially at moderate concentrations around 5%. In addition, the addition of a base at a low concentration ranging from 1% to 4% instead of water was able to improve the enzyme activity and therefore reduce the amount of enzyme by about 25% to 40% and the reaction time by about 8h compared to the reaction without added base, because the enzyme catalyst shows better activity in basic medium (at a value above 7, preferably at a pH of 8).

[0070] A.5.Temperature Change

[0071] Increasing the temperature improves the reaction and will allow the cost of producing the resulting biodiesel to be estimated. A very high temperature causes irreversible thermal denaturation of the enzyme protein, distorting the normal three-dimensional conformation of the enzyme protein. Even with a temperature below the optimal temperature, the reaction takes place but has extremely slow kinetics. The optimal temperature is specific to each enzyme and is independent of the manipulator's choices. In the case of the Novozym ® 40086 enzyme, the optimal reaction temperature is 30 °C. Some enzymes have a higher tolerance to temperature variations, such as Novozym ® 435, whose optimal temperature is located at 40 °C but which still has high activity between (approximately) 30 °C and (approximately) 60 °C.In general, the reaction temperature used in the process of the invention is between 30°C and 70°C, preferably between 35°C and 55°C, preferably between 40°C and 50°C.

[0072] A.6. Agitation in the reaction medium

[0073] This operational parameter ensures the equitable distribution of enzymes and reagents in a homogeneous manner in the reaction medium and forms a homogeneous emulsion which facilitates the function of the enzyme which acts at the interface of an organic / aqueous medium (here oil / alcohol). Indeed, the enzymes adopt an active open form when passing from the aqueous medium to the organic medium. Weak agitation leads to a poor distribution of enzymes and reagents which slows down the reaction rate and therefore lengthens the time required to complete the reaction, as well as an inhibition of the enzyme in certain places where the alcohol is in high concentration. Strong agitation can deform the enzymes, particularly immobilized ones, by damaging the support or by detaching the enzyme from the support, making the enzyme unusable and irrecoverable.For this, the reaction takes place at stirring speeds between 200 rpm and 1000 rpm, preferably between 300 rpm and 900 rpm. In this range, the percentage of esters in the biodiesel was found to be between 70% and 97% and the best yield was found at moderate stirring.

[0074] The shape of the reactor of module 2 of the invention is that of a cylindrical vessel with a conical bottom and a double jacket with a propeller-shaped blade. The double jacket of the reactor will ensure a stable and optimal temperature during the reaction. In addition, the propeller-shaped blade will ensure the even stirring of the reaction contents to obtain a homogeneous emulsion.

[0075] At the laboratory scale, the optimization of the operational parameters was carried out in batch-shaped glass reactors with a volume ranging from 100 ml to 10 liters. At the pilot scale, an optimal reaction was tested at a volume of 500 liters in a metal reactor. At the industrial scale, the production of biodiesel was carried out in a metal reactor with a volume of 5000 liters.

[0076] A.7. Preparation of the reaction medium

[0077] Since the enzyme (lipase) acts at the interface of an organic / aqueous medium, a preparation to form a homogeneous emulsion before adding the enzyme is carried out, which facilitates the efficient activity of the enzyme and ensures an optimal and homogeneous temperature throughout the reaction medium. Before adding the enzyme, a mixture of vegetable oil and water (or with a base) is stirred for a period of between 15 minutes and 2 hours, preferably between 30 minutes and 1 hour.

[0078] To increase the exchange surface between the enzyme (aqueous phase) and the triglycerides (organic phase) and to reduce the possibility of hydrolysis by water, glycerol is added. Then, the lipase is added into the reaction medium for a certain time to adapt with the medium before adding the alcohol and at the same time to hydrolyze a certain amount of triglycerides which accelerates the reaction rate by bringing about the esterification reaction which is faster than the trans-esterification reaction. This step improves the efficiency of the enzyme which in turn increases the reaction and reduces the time required to transform the triglycerides from lipids into biodiesel. Finally, the alcohol is added in several times to trigger the trans-esterification of the triglycerides.

[0079] B. Operating conditions for the treatment of an acidic vegetable oil

[0080] A vegetable oil can have a high level of free fatty acids. The objective is to recover all the glycerides and fatty acids present in the used acidic vegetable oil via two reactions: an esterification and a transesterification, both catalyzed by enzymes. Esterification is a reaction allowing acids to be transformed into esters by making them condense with an alcohol.

[0081] Three methods can be used to carry out this reaction. Either the reaction takes place in two stages. During the first stage, the pre-treated used acidic vegetable oil is reacted with ethanol or methanol and a first enzyme to esterify the free fatty acids into ethyl or methyl esters. This mixture is then recovered and then reacted with a second enzyme to trans-esterify the remaining glycerides. All the compounds in the used acidic vegetable oil have thus become ethyl or methyl esters. Esterification, unlike trans-esterification, will generate water.

[0082] A second method is to react the ethanol or methanol / acidic vegetable oil mixture with a single enzyme which carries out the esterification and transesterification.

[0083] A third method is to transform the acids in used acidic vegetable oil into triglycerides by esterification with glycerol at high temperature and then transforming these triglycerides into biodiesel in the presence of ethanol or methanol by enzymatic catalysis.

[0084] Several immobilized commercial enzymes such as Lipozyme ® TL IM, Novozym ® 40086, Lipozyme ® RM IM, CalB Sigma ®, CalB Immoplus ®, NovoLime ®, Novozym ® 435, as well as free commercial enzymes in liquid form such as TL Sigma ®, Callera TM, Eversa ®, Novozym @40116, RM liquid, Pallatase ® from Candida rugosa have been tested.

[0085] These three methods depend mainly on the nature of the enzyme. Some enzymes are 1,3-specific, 3-specific or have no specificity. Similarly, some enzymes are more affined for esterification, transesterification or both.

[0086] For example, Novozym ® 435 has no specificity and can therefore react with free fatty acids and glycerides and thus carry out esterification and transesterification at the same time. This, however, modifies the reaction kinetics of each reaction.

[0087] Lipozyme ® RM IM is a very efficient enzyme in esterification. Lipozyme ® TL IM is very efficient in trans-esterification and it is possible to react acid oil and ethanol first with Lipozyme ® RM IM, then with Lipozyme ® TL IM.

[0088] The immobilized enzyme Novozym ® 435 can convert acid oil with 75% free acids into biodiesel containing 95% ethyl esters, after 24 hours in the presence of ethanol as acyl acceptor.

[0089] C. Operating conditions for the treatment of used animal fat (beef fat)

[0090] For the treatment of used animal fat, like vegetable oil, an enzymatic reaction is optimized.

[0091] Two methods of transesterification were used:

[0092] transesterification in an organic solvent: used animal fat is reacted with ethanol or methanol and a commercial enzyme in the presence of hexane allowing better dissolution of the fats.

[0093] transesterification without organic solvent: used animal fat is reacted directly with ethanol or methanol and a commercial enzyme at a temperature that allows the fat to be kept in liquid form. This method requires much stronger stirring, especially at the start of the reaction and has slower kinetics. A second way to optimize the kinetics of this reaction is to carry out the reaction at a higher temperature (between approximately 40°C and approximately 50°C) with enzymes adapted or resistant to these temperatures. For example, the activity of Lipozyme ® TL IM is greatly reduced above 35°C. Conversely, certain enzymes such as Novozym ® 40086 have much larger activity ranges relative to the reaction temperature.

[0094] Three types of enzymes were tested: immobilized (TL-IM, Novozyme ® 40086, Lipozyme ® RM IM, CalB Sigma ®, CalB Immoplus ®, NovoLime ®, Novozym ® 435), free in solution such as TL Sigma ®, Callera TM, Eversa ®, Novozym ® 40116, RM liquid, Lipozyme ® TL IM, Pallatase ®, Candida rugosa) and lyophilized (Candida rugosa).

[0095] The free enzyme Novozym ® 40116 was able to transform beef fat into 80% ethyl ester biodiesel at 40°C after 24 hours. The enzyme recycling module or step 3

[0096] Advantageously and in order to improve the production of biodiesel in economic and ecological terms, the immobilized enzymes used can be collected from the reaction medium and cleaned in order to recycle and reuse them, preferably in the process of the invention. For this, the enzyme is washed in a closed circuit in module 3 with absolute hexane and is then dried overnight at room temperature. Using this washing method, the enzyme was reused several times while retaining almost 95% of its activity. This module and this step 3 is only useful for immobilized enzymes. For example, by applying this recycling method, Novozym ® 435 was reused 10 times for a period of one month while retaining almost 95% of its initial activity.

[0097] For free enzymes present in liquid form, recycling the enzyme is complex, because it is difficult to recover from the reaction medium. However, the Eversa ® enzyme could be reused twice by leaving it in the reaction medium and adding reagents (oils and alcohol) again without adding water.

[0098] The module or step 4 of refining the biodiesel obtained

[0099] To avoid generating waste and losing potential reagents or valuable by-products, the process of refining the crude biodiesel obtained is carried out in two sub-stages: a stage of evaporation of the residual alcohol in the mixture and a stage of natural decantation of the glycerol and water generated from the biodiesel obtained during the reaction.

[0100] The evaporation step has been optimized in order to eliminate the majority, preferably all of the excess alcohol and to recover this excess alcohol to re-react it in the process of the invention.

[0101] Evaporation takes place in module 4, i.e. a tank maintained under a residual pressure of approximately 150 mbar and a mixture heating temperature of approximately 120°C. These conditions are only slightly adjustable, except when working under lower residual pressures of up to approximately 30 mbar, thus making it possible to reduce the evaporation time using a falling film evaporator.

[0102] The liquid arrives in the upper part of the evaporation column and flows along the heated tubes. The apparatus is put under depression (at approximately 150 mbar) and the alcohol then evaporates to be condensed in a condenser. The collected ethanol (99% pure) is then reused in the transesterification reaction.

[0103] After evaporating all the alcohol, the mixture is transferred to a settling tank in the module where the glycerol and biodiesel separate naturally. Three phases are recovered: glycerol, a mixture of glycerol and refined biodiesel obtained consisting of pure biodiesel and pure biodiesel (B 100). The last two first products are kept in other 1000 liter tanks.

[0104] The glycerol obtained can be used in the food or cosmetics markets, the glycerol-biodiesel mixture being reintroduced into the refining module. The B100 is, for its part, transferred, via a pump, into the biodiesel treatment module described below.

[0105] The module or step 5 of treatment of refined biodiesel

[0106] Module 5 for processing refined biodiesel obtained from module or step 4 breaks down into two parts. In a tank 50, butylated hydroxytoluene (antioxidant) is added to the refined biodiesel obtained at a rate of 1500 ppm and this refined biodiesel obtained can then be transferred to another tank 51 and added with diesel at a rate of 70% in order to obtain a B30 biodiesel.

[0107] The most important advantage of the process and installation of the invention is the possibility of using the same enzymatic catalyst to trigger the transformation of so-called normal or acidic used oils and animal fats into biodiesel. This is a so-called "eco-clean" process, because unlike chemical processes using acidic or basic catalysts, such a process will only give biodiesel (as the main product) and glycerol (as a co-product), without side reactions (saponification process), thanks to the specificity and selectivity of the enzyme.

[0108] In the process of the invention, the efficiency of the enzyme used is also increased by preparing a homogeneous medium between the used oil and the added water and then reacting the enzyme with the used oil before adding the alcohol and triggering the transesterification. These two steps therefore made it possible to advantageously reduce the amount of enzyme required to carry out the reaction.

[0109] The efficiency of the enzyme could also be improved by adding a base (20% of a NaOH solution or 4% of NAeCO3) at low concentration and / or 5% glycerol which can reduce the necessary quantity of the enzyme to carry out this process and therefore the reaction time. In addition, the most effective enzyme in this process is present in liquid form, is not expensive and could be reused at least twice, without pose problems related to the type of mechanical agitation, unlike immobilized enzymes.

[0110] This process can be advantageously combined with local collection of lipids from used oils and used animal fats and local use of this biodiesel for vehicle traffic. Examples [YES] Example 1: Transformation of a sample of a vegetable oil into biodiesel

[0112] 100 ml of a vegetable oil obtained is poured into a 400 ml reactor 20 and then 3% (v / v) of water or base as well as glycerol (5%) are mixed with the oil for 30 minutes 2% (v / v) of the Eversa ® Transform 2.0 enzyme in liquid form is added to the reaction medium which is left for 1 hour without alcohol and therefore without triggering the reaction. This step makes the enzyme react with the oil to generate fatty acids and to accelerate the reaction. Then, ethanol at the molar ratio (4:1) to the vegetable oil is added in three times at time intervals of 0, 2 and 4 hours. After about 22 hours, the separation of the organic phase and the aqueous phase was carried out by centrifugation at 4700 rpm for 15 minutes.

[0113] For the production of biodiesel from methyl esters, the same process for ethyl esters as described above was applied, except that the molar ratio (Methanol: oil) is 6:1.

[0114] The content of the obtained refined biodiesel was studied by analyzing the samples by chromatographic methods (HPLC and GC).

[0115] Example 2: Transformation of a sample of an acidic vegetable oil into biodiesel

[0116] In the present invention, the transformation of acidic vegetable oils into biodiesel was catalyzed by the same enzyme that was used for the transformation of normal used vegetable oils into biodiesel. During this reaction, the step of thoroughly mixing the oil with water or a base was omitted, because the reaction took place without added water. The amount of free fatty acids is very high, which will induce the esterification reaction in the presence of the alcohol. Since the latter type of reaction produces the ester and water, the amount of water generated in the reaction medium will be sufficient to trigger the transesterification reaction between the triglycerides and the alcohol.Thus, the step to generate fatty acids between the enzyme and the alcohol-free oil was eliminated, because the oil used in this process is already acidic and contains a high amount of free fatty acids. Glycerol was added into the reaction medium which can create a site on which the enzyme works and decrease the hydrolysis and therefore decrease the amount of free fatty acids.

[0117] 100 ml of an acidic vegetable oil is poured into a 400 ml reactor 20 and 0.5% NaOH and 10% glycerol were added and then 1% (v / v) of the enzyme Eversa ® Transform 2.0 in liquid form was added. Finally, ethanol at the molar ratio (4:1) to acidic vegetable oil was added in three times at time intervals of 0, 2 and 4 hours to avoid enzymatic inhibition by alcohol. After 22 hours, the separation of the organic phase and the aqueous phase was carried out by centrifugation at 4700 rpm for 15 minutes. To study the content of the obtained refined biodiesel, samples were analyzed by chromatographic methods (HPLC and GC). For the production of biodiesel) from methyl esters, the same procedure for ethyl esters as described above was applied, except that the molar ratio (Methanol: oil) of 6:1 was used.

[0118] Example 3: Transformation of an animal fat sample into biodiesel

[0119] In the present invention, three enzymes have shown satisfactory results: Lipozyme ® TL IM, Novozym ® 40086 and Eversa ® Tranform 2.0. Therefore, the transformation of animal fats can be carried out by the same enzymatic catalyst used to transform normal or acid waste oils into biodiesel. As the objective of this invention is to find an eco-clean process, the transformation of an animal fat into biodiesel has been optimized without the addition of organic solvent.

[0120] 100 g of animal fat was weighed into a 400 ml reactor 20 and then 2% (w / v) of water or base was mixed with the oil for 30 minutes at 45°C to melt the fat well and mix it well with water or base. 4% (v / v) of the Eversa ® Transform 2.0 enzyme in liquid form was then added to the reaction medium which was left for 1 hour, without the addition of alcohol. Finally, ethanol at the molar ratio (5:1) to animal fat was added in three times at 0, 2 and 4 hours to avoid enzyme inhibition by alcohol.

[0121] After 22 hours, the separation of the organic phase and the aqueous phase was carried out by centrifugation at 4700 rpm for 15 minutes. To study the content of the obtained refined biodiesel, samples were analyzed by chromatographic methods (HPLC and GC). For the production of biodiesel from methyl esters, the same method for ethyl esters as described above was applied, except that the molar ratio (Methanol: fat) is 6:1.

Claims

Claims

1. A process for obtaining a biodiesel consisting of a mixture of ethyl esters or methyl esters by enzymatic transformation of lipids selected from the group consisting of used vegetable oil, acidic vegetable oil and / or used animal fat, wherein said enzymatic transformation comprises enzymatic transesterification and enzymatic hydroesterification, addition of alcohol, preferably an alcohol selected from the group consisting of ethanol, methanol or a mixture of these alcohols, addition of water or an aqueous base and optionally addition of glycerol, characterized in that prior to the addition of alcohol the lipids, the water or the aqueous base and optionally the glycerol form a homogeneous medium and in which this medium is then brought into contact with the enzyme to form a reaction medium

2. The method of claim 1, wherein the enzyme is a triacylglycerol acyl hydrolase enzyme.

3. Method according to any one of the preceding claims, in which the lipids are subjected, prior to the enzymatic transformation into biodiesel, to one or more step(s) of filtration of the solid particles present in the lipids, and obtaining lipids containing a water content of less than 1% by weight relative to the weight of the lipids.

4. Process according to claim 3, comprising, prior to the enzymatic transformation into biodiesel, one or more step(s) of water removal.

5. A method according to claim 3, wherein the lipids consist of used palm oil or used animal fat which are heated to a temperature of 40°C or more, preferably 45°C or more, before the filtration step(s).

6. Process according to claim 3, in which the lipids are animal fats which are subjected, prior to the enzymatic transformation into biodiesel, to a decantation step at room temperature for used oils and at 45°C or more for animal fats for a period of between 48 hours and 72 hours.

7. A method according to any preceding claim, wherein the homogeneous medium is contacted with the enzyme for a period of between 30 minutes and 2 hours to form a reaction medium.

8. A method according to any one of the preceding claims, wherein the alcohol is added to said reaction medium sequentially from 1 to 4 times, after a period of between 15 minutes and 4 hours, preferably after a period of between 30 minutes and two hours.

9. A method according to any one of the preceding claims and comprising a step of recycling the used enzyme, said recycling step comprising a step of reusing the free enzyme, a step of immobilizing the used enzyme, a step of collecting the used immobilized enzyme, a step of adding hexane to the collected enzyme and a step of drying the collected enzyme.

10. A method according to any one of the preceding claims, comprising after the enzymatic transformation of the used lipids into biodiesel, - a step of refining the reaction medium including a separation of the alcohol, the glycerol formed during the enzymatic transformation, and an aqueous part of the reaction medium from the biodiesel obtained which comprises an evaporation of the alcohol from the reaction medium, and optionally a collection by condensation of the evaporated alcohol and / or an addition of the alcohol recycled in the process for obtaining the biodiesel according to any one of the preceding claims and a decantation of the glycerol and / or the aqueous part of the reaction medium and - a step of collecting the refined biodiesel, followed by a - step of adding an antioxidant, preferably butylated hydroxytoluene, to the refined and collected biodiesel,and / or a step of blending the collected refined biodiesel with another fuel to obtain a final 30% biodiesel (B30).,

11. A method according to claim 10, wherein an aqueous portion of the reaction medium incorporating the enzyme is added with alcohol and used lipids to obtain the biodiesel according to any one of the preceding claims 1 to 10.

12. A method according to claim 10 or 11, wherein the antioxidant is butylated hydroxytoluene.

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