Methods for the co-production of organic acids and lignin from lignocellulose

Abstract

This invention provides a method for the co-production of organic acids and lignin from lignocellulose, comprising the following steps: Step 1, pretreating the lignocellulose raw material to obtain a pretreated material; Step 2, enzymatically hydrolyzing the pretreated material to obtain an enzymatic hydrolysate containing pentoses, hexoses, and lignin; Step 3, performing solid-liquid separation on the enzymatic hydrolysate to obtain an enzymatic hydrolysate containing pentoses and hexoses and a solid phase containing lignin; Step 4, subjecting the enzymatic hydrolysate containing pentoses and hexoses to acid-producing fermentation to obtain a fermentation broth containing a mixture of organic acids; Step 5, extracting lignin from the lignin-containing solid phase to obtain lignin. The method provided by this invention can efficiently separate lignin and obtain various organic acids through a simple process.

Description

Technical Field

[0001] This invention relates to the technical field of biochemical production, and more specifically, to a method for the co-production of organic acids and lignin from lignocellulose. Background Technology

[0002] Lignocellulose is mainly composed of lignin, hemicellulose, and cellulose. Lignin is a cross-linked phenolic polymer, hemicellulose is a short-chain heteropolysaccharide, and cellulose is a polymer of the monosaccharide glucose covalently linked by beta-1,4 bonds. In recent years, researchers have been continuously exploring and studying methods for separating lignin, hemicellulose, and cellulose, while also exploring methods for converting these polymers into other important intermediate raw materials.

[0003] Organic acids, containing a carboxyl group (-COOH) in their chemical structure, are acidic organic compounds and important raw materials for organic synthesis, industrial and agricultural production, and the pharmaceutical industry. Organic acids include natural and synthetic organic acids. Natural organic acids are widely found in the fruits, roots, and leaves of plants and can be directly extracted and separated. Synthetic organic acids are obtained through chemical synthesis, enzymatic fermentation, and microbial fermentation. Lignocellulose is an abundant and renewable source, and in recent years, various organic acids have been prepared using it as a raw material. For example, patent CN1934249A provides a method for degrading lignocellulose materials. In the presence of a surfactant, the enzymatic hydrolysis and saccharification of lignocellulose are promoted, and the saccharified lignocellulose is fermented to obtain organic acids. However, this method has a complex reaction system and difficult product extraction, which is not conducive to industrial application. Summary of the Invention

[0004] To address the above problems, this invention provides a method for the co-production of organic acids and lignin from lignocellulose, which can efficiently separate lignin and obtain a variety of organic acids through a simple process.

[0005] The purpose of this invention is to provide a method for the co-production of organic acids and lignin from lignocellulose, comprising the following steps:

[0006] Step 1: Pre-treat the lignocellulose raw material to obtain a pre-treated material containing cellulose, xylose and lignin;

[0007] Step 2: Enzymatically hydrolyze the pretreated material obtained in Step 1 to obtain an enzymatic hydrolysate containing pentose, hexose, and lignin;

[0008] Step 3: Perform solid-liquid separation on the enzymatic hydrolysate obtained in Step 2 to obtain an enzymatic hydrolysate containing pentose and hexose and a solid phase containing lignin;

[0009] Step 4: Perform acid-producing fermentation on the enzymatic hydrolysate containing pentose and hexose obtained in Step 3 to obtain a fermentation broth containing a mixture of organic acids;

[0010] Step 5: Extract lignin from the lignin-containing solid phase obtained in Step 3 to obtain lignin.

[0011] Preferably, the lignin cellulose raw material in this invention can be selected from one or more of corn stalks, rice stalks, wheat stalks, reeds, corn cobs, and soybean stalks.

[0012] Preferably, in step one, the method for pretreating the lignocellulose raw material is as follows: after the lignocellulose raw material is crushed and impurities are removed, it is subjected to acid treatment, alkali treatment, and steam explosion treatment to obtain the pretreated material.

[0013] It is worth mentioning that the pulverization and impurity removal of the lignocellulose raw material of the present invention is achieved by using a complete set of pulverization and impurity removal equipment, specifically including coarse pulverization, impurity and iron removal, fine pulverization and dust removal treatment sections.

[0014] Preferably, in step two, the enzyme used for enzymatic hydrolysis of the pretreated material is cellulase, wherein the mass ratio of the added cellulase to the mass of the lignocellulosic raw material is 0.01 to 0.3:1.

[0015] It is worth mentioning that after the pretreated material is enzymatically hydrolyzed, a solid-liquid mixed enzymatic hydrolysate containing pentose, hexose and lignin is obtained. In step three, in order to separate the solid and liquid, plate and frame filtration or centrifugation can be used to separate the solid-liquid mixed enzymatic hydrolysate into an enzymatic hydrolysate containing pentose and hexose and a solid phase containing lignin.

[0016] Preferably, in step four, the conditions for acid-producing fermentation are: batch fermentation, semi-continuous fermentation, or continuous fermentation under anaerobic conditions; pH of acid-producing fermentation = 5-10, preferably pH = 5-7; temperature of acid-producing fermentation = 30-58℃, preferably 35-40℃; C / N ratio = 20-60, preferably 30-40; and fermentation time = 2-50 days, preferably 7-14 days.

[0017] Preferably, the nitrogen source is one or a combination of NH4Cl, corn steep liquor, sludge, potassium nitrate, and urea.

[0018] Preferably, the acid-producing fermentation of the present invention uses an acid-producing microbial community; more preferably, it uses an anaerobic fermentation acid-producing microbial community enriched by the method provided in patent application (application number 2023113103664).

[0019] Specifically, the enrichment method for anaerobic fermentation acid-producing microbial communities includes stages F1 to F3. The reactor's operation can be divided into three stages: F1, F2, and F3. The organic loading of straw is gradually increased in each stage until stable operation is achieved in stage F3. Stage F1 is the reactor start-up stage, with low solids content and loading. Increasing the solids content and loading leads to stage F2. After acid production stabilizes in F2, the proportion of straw in the material is increased, raising the C / N ratio, leading to stage F3, where the microbial community adapts to the high C / N ratio straw material.

[0020] Specifically as follows:

[0021] F1 Stage:

[0022] The F1 stage takes 80 to 120 days, preferably 100 to 110 days;

[0023] Add the inoculum to the reactor, purge with nitrogen to remove air, and then seal the reactor. Do not feed or discharge any material for two days after starting the reactor. After that, supply the substrate under an organic load of 0.4–0.6 g VTS / L / d. The TS value of the substrate is 4.8–5.4%, the VTS value is 4.0–4.4%, and the C / N ratio is 18–22. Feed and discharge can be performed once every two days. The hydraulic retention time of the reactor is 90–110 days. The physicochemical parameters of the reactor can be monitored every four days.

[0024] Specifically, regarding the source of vaccination:

[0025] The solid content of the inoculum used in this invention is ≤5%.

[0026] Furthermore, to inhibit the activity of methanogens, it is preferable to pretreat the inoculum source. The pretreatment method involves heat-treating the inoculum source at 75–85°C for 0.5–1.2 hours and adding a methane inhibitor at a final concentration of 4–6 mM. Preferably, the methane inhibitor is 2-bromoethanosulfonic acid (BES).

[0027] Furthermore, the inoculum source can include inoculum from different sources; specifically, the inoculum from different sources can be selected from a mixture of anaerobic digestion sludge from grain and sugar mills, anaerobic digestion sludge from kitchen waste, anaerobic digestion sludge from cellulose, anaerobic digestion sludge from glucose, and sludge from anaerobic fermentation and acid production reactors from fruit waste. Among these, the cellulose anaerobic digestion sludge is preferably either thermophilic or mesophilic cellulose anaerobic digestion sludge; the glucose anaerobic digestion sludge is preferably either thermophilic or mesophilic glucose anaerobic digestion sludge. The appropriate sludge type can be selected based on different reaction temperatures.

[0028] Furthermore, the inoculum source comprises inoculum from different sources mixed according to total solids (TS), with each type of sludge accounting for more than 10% of the total solids (TS), and more preferably, multiple inoculum sludges from different sources are mixed in equal proportions according to the total solids (TS).

[0029] In practice, the inoculum source can be crushed with a mixer and then filtered (for example, filtered with 2-3 layers of gauze) to ensure that the solid content of the sludge is below 5% when the reactor is started.

[0030] Specifically, regarding the substrate:

[0031] The substrate used in this invention is a mixture containing straw, kitchen waste and nitrogen source substrate; in the F1 stage, the preferred VTS ratio of straw, kitchen waste and nitrogen source substrate in the substrate is (0.8~1.2):(0.8~1.2):(0.8~1.2).

[0032] Furthermore, the source substrate can be any substrate commonly used in the field, specifically wastewater treatment plant sludge. Currently, a large amount of wastewater is generated during the activated sludge process in urban wastewater treatment plants in my country. However, the sludge treatment technology is not yet mature. Using wastewater treatment plant sludge as a nitrogen source substrate can reuse the sludge, which is environmentally friendly and cost-effective.

[0033] Furthermore, the straw may be selected from at least one or a combination of corn straw, rice straw, wheat straw, reeds or bean straw.

[0034] Furthermore, the straw can be steam-exploded straw. Steam-exploded straw can be straw that has undergone conventional steam explosion treatment in this field, or it can be prepared by the following method: adding straw material and water to a sealed container, maintaining it for a period of time (which can be a few minutes), and then suddenly reducing the pressure to steam-explode the straw material, thereby destroying the hemicellulose and lignin bonding layer, exposing more active groups of cellulose, which is more conducive to degradation.

[0035] Furthermore, food waste can be regular food waste or homemade waste, such as wet waste containing crushed fruits, vegetables, and grains.

[0036] Preferably, the substrate can be stored at low temperature (e.g., 4°C) and diluted to the target solids content before use.

[0037] Specifically, regarding the reactor:

[0038] The reactor used in this invention can be a high-temperature reactor or a medium-temperature reactor. Conventional reactors in the art can be selected, such as conventional mechanically stirred tanks. The stirring speed of the reactor can be 0–150 ppm, sufficient to ensure uniform mixing of the materials.

[0039] Preferably, the pH value of the reactor is 5.5 to 6.0; the operating temperature of the high-temperature reactor is 50 to 55°C; and the operating temperature of the medium-temperature reactor is 35 to 40°C.

[0040] Furthermore, when the reactor is a high-temperature reactor, the cellulose anaerobic digestion sludge can be selected from cellulose high-temperature anaerobic digestion sludge, and the glucose anaerobic digestion sludge can be selected from glucose high-temperature anaerobic digestion sludge.

[0041] Furthermore, when the reactor is a mesophilic reactor, the cellulose anaerobic digestion sludge can be selected from cellulose mesophilic anaerobic digestion sludge, and the glucose anaerobic digestion sludge can be selected from glucose mesophilic anaerobic digestion sludge.

[0042] In practice, a semi-continuous fully mixed-flow anaerobic fermentation acid-producing reactor system can be constructed, using either a high-temperature reactor or a mesophilic reactor to enrich high-temperature and mesophilic bacterial communities, respectively.

[0043] F2 phase:

[0044] The operating time of the F2 stage is 50–100 days, preferably 75–90 days; the reactor pH is 5.5–6.0; the feed solids content is adjusted to 8–12%, and the organic load is increased to 0.8–1.2 g VTS / L / d for substrate supply, with substrate TS of 9.8–10.5% and substrate VTS of 8.0–8.5%, and the VTS ratio of straw, kitchen waste and sludge in the substrate is (0.8–1.2):(0.8–1.2):(0.8–1.2); the substrate C / N ratio is 18–22; the reactor can be fed and discharged once every two days; the hydraulic retention time of the reactor can be 90–110 days; and it should be continuously operated until it stabilizes.

[0045] F3 stage:

[0046] The operating time for stage F3 is 120–200 days, preferably 165–185 days; the reactor pH is 5.5–6.0; the feed solids content remains constant during this stage (8–12%), while the organic loading is increased to 1.8–2.2 g. Substrate was supplied at VTS / L / d, and the hydraulic retention time was adjusted to 45–55 days. The VTS ratio of straw, kitchen waste, and sludge in the feed was changed from (0.8–1.2):(0.8–1.2):(0.8–1.2) to (2.8–3.2):(1.8–2.2):(0.8–1.2). The substrate C / N ratio was increased from 18–22 to 38–42. The substrate TS was 9.5–10%, and the substrate VTS was 8.0–8.6%. Feeding and discharging were carried out every two days. The operation continued until stable. Microbial communities in the reactor were collected during the stable operation of the F3 stage.

[0047] It is worth mentioning that the microbial community of the reactor during the stable operation of the F3 stage was collected by taking the fermentation broth, centrifuging it at 4°C for 10 minutes, removing the supernatant, and the precipitate being the microbial community. The enriched microbial community was used for the acid-producing fermentation of the enzymatic hydrolysate of this invention.

[0048] Preferably, the enzymatic hydrolysate containing pentose and hexose is subjected to acid-producing fermentation by an enriched microbial community to obtain a fermentation broth containing a mixture of organic acids. The organic acids in the fermentation broth are combinations of lactic acid, acetic acid, propionic acid, isobutyric acid, butyric acid, valeric acid, and isovaleric acid. This invention can adjust the composition and content of acids in the mixed organic acids by controlling the conditions of the acid-producing fermentation.

[0049] Preferably, in step five, lignin can be extracted by alkali dissolution, solid-liquid separation, acid precipitation, water washing, and air drying.

[0050] Compared with the prior art, the beneficial effects of the present invention are as follows: The present invention provides a method for co-producing mixed organic acids and lignin by using lignocellulose as a raw material to replace grain raw materials, and by controlling the acid-producing fermentation conditions and adjusting the composition and content of acids in the mixed organic acids, a high concentration of organic acids is obtained in the fermentation broth of the mixed organic acids. Attached Figure Description

[0051] Figure 1 This is a flowchart illustrating the co-production of organic acids and lignin from lignocellulose using this invention. Detailed Implementation

[0052] The present invention will now be described in detail with reference to the accompanying drawings and embodiments. It should be noted that the following embodiments are only used to further illustrate the present invention and should not be construed as limiting the scope of protection of the present invention. Some non-essential improvements and adjustments made by those skilled in the art based on the content of the present invention are still within the scope of protection of the present invention.

[0053] Unless otherwise specified, the raw materials used in the embodiments are all publicly available in the prior art, such as those that can be directly purchased or prepared according to the preparation methods disclosed in the prior art.

[0054] The sources of sludge, kitchen waste, and straw used in the embodiments of the present invention are as follows:

[0055] Anaerobic digestion sludge from grain and sugar mills: COFCO Biochemical (Chengdu) Co., Ltd.;

[0056] Anaerobic digestion sludge from food waste: An anaerobic methanogenic reactor that operates stably at the Environmental Biotechnology Research Center of the School of Architecture and Environment, Sichuan University. The food waste is taken from the student canteen of Sichuan University.

[0057] Cellulose high-temperature anaerobic digestion sludge: The high-temperature anaerobic methanogenic reactor, stably operated by the Environmental Biotechnology Research Center of the School of Architecture and Environment, Sichuan University, uses commercially available carboxymethyl cellulose as a carbon source.

[0058] Cellulose mesophilic anaerobic digestion sludge: A mesophilic anaerobic methanogenic reactor stably operating at the Environmental Biotechnology Research Center of the School of Architecture and Environment, Sichuan University, using commercially available carboxymethyl cellulose as a carbon source.

[0059] High-temperature anaerobic digestion of glucose sludge: A high-temperature anaerobic methanogenic reactor that is operating stably at the Environmental Biotechnology Research Center of the School of Architecture and Environment, Sichuan University, using commercially available glucose as a carbon source.

[0060] Mesophilic anaerobic digestion of glucose sludge: A stable-running mesophilic anaerobic methanogenic reactor at the Environmental Biotechnology Research Center of the School of Architecture and Environment, Sichuan University, using commercially available glucose as a carbon source.

[0061] Sludge from the anaerobic fermentation acid-producing reactor for fruit waste: The anaerobic fermentation acid-producing reactor, which is stably operating at the Environmental Biotechnology Research Center of the School of Architecture and Environment, Sichuan University, uses commercially available fruits (a mixture of bananas, watermelons, and oranges in a weight ratio of 1:1:1) as raw materials.

[0062] The wastewater treatment plant's residual sludge is dewatered sludge from a wastewater treatment plant in Chengdu.

[0063] Artificially prepared food waste is made by mixing pulverized bananas, oranges, cabbage, and rice by wet weight;

[0064] Steam-exploded straw refers to straw that has been blasted by steam. The steam blasting process includes: adding corn stalks and water to a sealed container, maintaining a pressure of 1.7 MPa for 2 minutes, and then suddenly reducing the pressure to blast the straw material with steam.

[0065] Table 1 shows the TS and VTS data for inoculum and substrate from different sources.

[0066] Table 1

[0067]

[0068] In the example, one high-temperature reactor (working volume 4L) and one mesophilic reactor (working volume 2.4L) were prepared to enrich microbial communities suitable for high temperatures and mesophilic communities, respectively.

[0069] Example 1

[0070] This embodiment illustrates a method for enriching microbial communities.

[0071] 1. Reactor Construction

[0072] Preparation of inoculum source: In order to enrich the anaerobic fermentation acid-producing microbial community, anaerobic digestion sludge from grain and sugar mills, anaerobic digestion sludge from kitchen waste, high-temperature anaerobic digestion sludge from cellulose, high-temperature anaerobic digestion sludge from glucose, and sludge from anaerobic fermentation acid-producing reactors from fruit waste were mixed as inoculum source; inoculum from different sources were mixed in equal proportions (i.e., 1:1:1:1:1) according to the total solids (TS) concentration (TS and VTS of inoculum from different sources are shown in Table 1).

[0073] Substrate preparation: Steam-exploded straw, artificially prepared kitchen waste (a mixture of crushed bananas, oranges, cabbage, and rice, mixed by wet weight), and wastewater treatment plant sludge were used as substrates. Different substrates were mixed in equal proportions according to their volatile total solids (VTS) concentration (TS and VTS for each substrate are shown in Table 1). The mixed substrates were stored at 4°C and diluted to the target solids content before use.

[0074] Prepare one high-temperature reactor (working volume 4L, named H).

[0075] 2. Reactor Start-up and Operation

[0076] This invention starts a high-temperature reactor (H).

[0077] The reactor's entire operation process is divided into three stages: F1, F2, and F3. The organic load of straw is gradually increased in each stage until stable operation is achieved in stage F3. The operating conditions of the reactor in each stage are shown in Table 2.

[0078] Days 0 to 105 constitute the F1 stage.

[0079] The high-temperature reactor was started and operated at 53℃, with a stirring speed of 100 rpm and a pH value controlled between 5.5 and 6.0. The inoculum source, obtained by mixing inoculating sludge from various sources in equal proportions according to the TS (Standard Test), was crushed using a mixer and filtered through 2-3 layers of gauze to ensure the solid content of the reactor start-up sludge was below 5%. To inhibit methanogenic bacteria activity, the start-up sludge required pretreatment. The start-up sludge was heat-treated at 80℃ for 1 hour, and a methane inhibitor, 2-bromoethanosulfophate (BES), was added to a final concentration of 5 mM. The pretreated mixed inoculum was injected into the reactor's working volume, nitrogen was introduced to purge air to achieve an anaerobic state, and the reactor was then sealed. For the first two days after reactor startup, no feed or discharge was performed. Afterward, a substrate consisting of straw, kitchen waste, and sludge (TS: 5.2%; VTS: 4.1%; C / N ratio: 20) was supplied under an organic loading of 0.5 g VTS / L / d. The VTS ratio of straw, kitchen waste, and sludge in the substrate was 1:1:1. Feed and discharge were performed every two days. The hydraulic retention time of the reactor was 100 days. The reactor's physicochemical parameters were monitored every four days.

[0080] Days 105 to 189 are the F2 phase.

[0081] During this stage, the hydraulic retention time is maintained at 100 days, the feed solids content is adjusted to 10%, the substrate TS is 10.3%, the substrate VTS is 8.2%, the VTS ratio of straw, kitchen waste and sludge in the substrate is 1:1:1, the substrate C / N ratio is 20, the organic load is increased to 1g VTS / L / d, and the operation continues until it stabilizes.

[0082] Days 189 to 368 are the F3 phase.

[0083] During this phase, the feed solids content was maintained at 10%, the organic loading rate was increased to 2 g VTS / L / d, the hydraulic retention time was adjusted to 50 days, the substrate TS was 9.7%, the substrate VTS was 8.3%, and the VTS ratio of straw, kitchen waste, and sludge in the feed was changed from 1:1:1 to 3:2:1, increasing the proportion of straw and decreasing the proportion of sludge, thus raising the feed C / N ratio from 20 to 40. This process continued until stability was achieved.

[0084] The microbial community of the reactor during the stable operation of the F3 stage was collected by taking the fermentation broth, centrifuging it at 4°C for 10 minutes, removing the supernatant, and the precipitate was the microbial community.

[0085] Example 2

[0086] This embodiment illustrates a method for enriching microbial communities.

[0087] 1. Reactor Construction

[0088] Preparation of inoculum sources: In order to enrich the anaerobic fermentation acid-producing microbial community, anaerobic digestion sludge from grain and sugar mills, anaerobic digestion sludge from kitchen waste, mesophilic anaerobic digestion sludge from cellulose, mesophilic anaerobic digestion sludge from glucose, and sludge from anaerobic fermentation acid-producing reactors from fruit waste were mixed as inoculum sources; inoculum sources from different sources were mixed in equal proportions according to total solids (TS) concentration (TS and VTS of inoculum sources from different sources are shown in Table 1).

[0089] Substrate preparation: Steam-exploded straw, artificially prepared kitchen waste (a mixture of crushed bananas, oranges, cabbage, and rice, mixed by wet weight), and wastewater treatment plant sludge were used as substrates. Different substrates were mixed in equal proportions according to their volatile total solids (VTS) concentration (TS and VTS for each substrate are shown in Table 1). The mixed substrates were stored at 4°C and diluted to the target solids content before use.

[0090] Prepare one medium-temperature reactor (working volume 2.4L).

[0091] 2. Reactor Start-up and Operation

[0092] This invention starts a medium-temperature reactor.

[0093] The reactor's entire operation process is divided into three stages: F1, F2, and F3. The organic load of straw is gradually increased in each stage until stable operation is achieved in stage F3. The operating conditions of the reactor in each stage are shown in Table 2.

[0094] Days 0 to 105 constitute the F1 stage.

[0095] The mesophilic reactor was started and operated at 37°C with a stirring speed of 100 rpm and a pH value controlled between 5.5 and 6.0. The inoculum source, obtained by mixing sludge from various sources in equal proportions according to the TS (Standard Test), was crushed using a mixer and filtered through 2-3 layers of gauze to ensure the solid content of the reactor start-up sludge was below 5%. To inhibit methanogenic bacteria activity, the start-up sludge required pretreatment. The start-up sludge was heat-treated at 80°C for 1 hour, and a methane inhibitor, 2-bromoethanosulfophate (BES), was added to a final concentration of 5 mM. The pretreated mixed inoculum was injected into the reactor's working volume, nitrogen was introduced to purge air to achieve an anaerobic state, and the reactor was then sealed. For the first two days after reactor startup, no feed or discharge was performed. Afterward, a substrate consisting of straw, kitchen waste, and sludge (TS: 5.2%; VTS: 4.1%; C / N ratio: 20) was supplied under an organic loading of 0.5 g VTS / L / d. The VTS ratio of straw, kitchen waste, and sludge in the substrate was 1:1:1. Feed and discharge were performed every two days. The hydraulic retention time of the reactor was 100 days. The reactor's physicochemical parameters were monitored every four days.

[0096] Days 105 to 189 are the F2 phase.

[0097] During this stage, the hydraulic retention time is maintained at 100 days, the feed solids content is adjusted to 10%, the substrate TS is 10.3%, the substrate VTS is 8.2%, the VTS ratio of straw, kitchen waste and sludge in the substrate is 1:1:1, the substrate C / N ratio is 20, and the organic load is increased to 1g VTS / L / d, and the operation continues until it stabilizes.

[0098] Days 189 to 368 are the F3 phase.

[0099] During this phase, the feed solids content was maintained at 10%, the organic loading rate was increased to 2 g VTS / L / d, the hydraulic retention time was adjusted to 50 days, the substrate TS was 9.7%, the substrate VTS was 8.3%, and the VTS ratio of straw, kitchen waste, and sludge in the feed was changed from 1:1:1 to 3:2:1, increasing the proportion of straw and decreasing the proportion of sludge, thus raising the feed C / N ratio from 20 to 40. This process continued until stability was achieved.

[0100] To collect the reactor microbial community during the stable operation of the F3 stage, the specific collection method is as follows: take the fermentation broth, centrifuge at 4℃ for 10 minutes, remove the supernatant, and the precipitate is the microbial community.

[0101] Table 2 shows the operating conditions data for different stages of the reactor.

[0102] Table 2

[0103] F1 F2 F3 Substrate TS (%) 5.2 10.3 9.7 Substrate VTS (%) 4.1 8.2 8.3 The proportion of straw, kitchen waste and sludge VTS in the substrate 1:1:1 1:1:1 3:2:1 substrate C / N ratio 20 20 40 Organic loading (gVTS / L / d) 0.5 1 2 Duration of stay (d) 100 100 50

[0104] Example 3

[0105] This embodiment illustrates a method for obtaining an organic acid mixture and lignin from lignocellulose (see detailed process flow). Figure 1 The process includes the following steps:

[0106] Step 1: Crush and remove impurities from the corn stalks, and impregnate the corn stalk raw material with 2wt% dilute sulfuric acid. The amount of dilute sulfuric acid used is 5 times that of the dry corn stalks. Impregnate at room temperature for 20 minutes and treat at a pressure of 0.9 MPaG for 30 minutes to obtain pretreated material containing cellulose, xylose and lignin.

[0107] Step 2: Enzymatic hydrolysis of the pretreated material containing cellulose, xylose and lignin was performed. The amount of cellulase used was 0.18 g / g cellulose, the concentration of the enzymatic hydrolysate dry matter was 10 wt%, the pH of the enzymatic hydrolysis was 5, and the enzymatic hydrolysis time was 96 h, to obtain a solid-liquid mixed enzymatic hydrolysate containing pentose, hexose and lignin.

[0108] Step 3: Perform solid-liquid centrifugation on the solid-liquid mixed enzymatic hydrolysate containing pentose, hexose and lignin to obtain the enzymatic hydrolysate containing pentose and hexose and the solid phase containing lignin;

[0109] Step four involves acid-producing fermentation of the enzymatic hydrolysate containing pentose and hexose. The microbial community used is the one enriched in Example 2. The inoculum size for acid-producing fermentation is 2%, and the conditions are 37°C, pH=6, nitrogen source NH4Cl, and C / N ratio of 40. Fermentation is carried out for 8 days, yielding an organic acid mixture of 33.68 g / L with a carbon conversion rate of 88%. High-performance liquid chromatography (HPLC) analysis revealed lactic acid 16.9 g / L, acetic acid 5.43 g / L, propionic acid 2.7 g / L, isovaleric acid 8.34 g / L, and other volatile fatty acids such as butyric acid, isobutyric acid, and valeric acid 0.31 g / L.

[0110] Step 5: Extract lignin from the lignin-containing solid phase. The specific steps are as follows:

[0111] (1) Alkali dissolution: Sodium hydroxide is used to dissolve the lignin in the solid phase. The amount of sodium hydroxide is 11% of the dry solid phase, and the mass of water added is 7 times the mass of the dry solid phase. The treatment time is 2 hours to obtain an alkaline solution of lignin.

[0112] (2) Solid-liquid separation: The lignin alkaline solution is centrifuged to obtain a solution containing lignin;

[0113] (3) Acid precipitation: Add sulfuric acid to the lignin solution and treat at 80°C for 1 hour to precipitate the lignin. The mass of sulfuric acid used is 1.15 times the mass of sodium hydroxide used. Then, centrifuge to separate the wet lignin.

[0114] (4) Washing: The wet lignin is washed with water, the amount of water being 8 times that of the dry wet lignin, and then centrifuged to obtain the washed wet lignin.

[0115] (5) After air drying, lignin is obtained.

[0116] Example 4

[0117] This embodiment illustrates a method for obtaining an organic acid mixture and lignin from lignocellulose (see detailed process flow). Figure 1 The process includes the following steps:

[0118] Step 1: Crush and remove impurities from the corn stalks, and impregnate the corn stalk raw material with 2wt% dilute sulfuric acid. The amount of dilute sulfuric acid used is 5 times that of the dry corn stalks. Impregnate at room temperature for 20 minutes, and then treat under a pressure of 0.9 MPaG for 30 minutes to obtain a pretreated material containing cellulose, xylose and lignin.

[0119] Step 2: The pretreated material containing cellulose, xylose and lignin is subjected to enzymatic hydrolysis. The amount of cellulase used is 0.18 g / g cellulose, the concentration of the enzymatic hydrolysate dry matter is 10 wt%, the pH of the enzymatic hydrolysis is 5, and the enzymatic hydrolysis time is 96 h, to obtain a solid-liquid mixed enzymatic hydrolysate containing pentose, hexose and lignin.

[0120] Step 3: Perform solid-liquid centrifugation on the solid-liquid mixed enzymatic hydrolysate containing pentose, hexose and lignin to obtain the enzymatic hydrolysate containing pentose and hexose and the solid phase containing lignin;

[0121] Step four involves acid-producing fermentation of the enzymatic hydrolysate containing pentose and hexose. The microbial community used is the one enriched in Example 1. The inoculum size for acid-producing fermentation is 2%, and the conditions are 53°C, pH=5, nitrogen source NH4Cl, and C / N ratio of 40. Fermentation is carried out for 14 days, yielding an organic acid mixture of 21.72 g / L with a carbon conversion rate of 46%. High-performance liquid chromatography (HPLC) analysis revealed lactic acid 13.18 g / L, acetic acid 2.83 g / L, propionic acid 2.69 g / L, isovaleric acid 2.68 g / L, and other volatile fatty acids such as butyric acid, isobutyric acid, and valeric acid 0.33 g / L.

[0122] Step 5: Extract lignin from the lignin-containing solid phase. The specific steps are as follows:

[0123] (1) Alkali dissolution: Sodium hydroxide is used to dissolve the lignin in the solid phase. The amount of sodium hydroxide is 11% of the dry solid phase, and the mass of water added is 7 times the mass of the dry solid phase. The treatment time is 2 hours to obtain an alkaline solution of lignin.

[0124] (2) Solid-liquid separation: The lignin alkaline solution is centrifuged to obtain a solution containing lignin;

[0125] (3) Acid precipitation: Add sulfuric acid to the lignin solution and treat at 80°C for 1 hour to precipitate the lignin. The mass of sulfuric acid used is 1.15 times the mass of sodium hydroxide used. Then, centrifuge to separate the wet lignin.

[0126] (4) Washing: The wet lignin is washed with water, the amount of water being 8 times that of the dry wet lignin, and then centrifuged to obtain the washed wet lignin.

[0127] (5) After air drying, lignin is obtained.

Claims

1. A method for the co-production of organic acids and lignin from lignocellulose, characterized in that, The method comprises the following steps: Step one, pretreating the lignocellulose raw material to obtain pretreated material containing cellulose, xylose and lignin; Step two, enzymatic hydrolysis of the pretreated material obtained in step one to obtain enzymatic hydrolysis liquid containing pentose, hexose and lignin; Step three, solid-liquid separation of the enzymatic hydrolysis liquid obtained in step two to obtain enzymatic hydrolysis liquid containing pentose and hexose and solid phase containing lignin; Step four, acid production fermentation of the enzymatic hydrolysis liquid containing pentose and hexose obtained in step three to obtain fermentation liquid containing organic acid mixture; Step five, lignin extraction of the solid phase containing lignin obtained in step three to obtain lignin; The acid production fermentation uses an anaerobic fermentation acid-producing microbial community enriched by the following F1 stage-F3 stage method; F1 stage: Inoculum is added to the reactor, nitrogen is introduced to discharge air, and then the reactor is closed; the solid content of the inoculum is ≤5%; no material is fed or discharged for two days after starting the reactor, and then the substrate is supplied under the condition of 0.4-0.6 g VTS / L / d organic load; the C / N ratio is 18-22; wherein VTS is volatile solid concentration; F2 stage: The solid content of the feed is adjusted to 8-12%, the organic load is increased to 0.8-1.2 g VTS / L / d, and the substrate C / N ratio is 18-22; F3 stage: The solid content of the feed is kept at 8-12%, the organic load is increased to 1.8-2.2 g VTS / L / d, and the substrate C / N ratio is increased from 18-22 to 38-42; the microbial flora in the F3 stage reactor is collected.

2. The method of lignocellulose co-production of organic acids and lignin according to claim 1, characterized in that, In the step one, the lignocellulose raw material is derived from one or more of corn stalks, rice straw, wheat straw, reed, corn cob and bean straw.

3. The method of lignocellulose co-production of organic acids and lignin according to claim 1, characterized in that, In the step one, the method for pretreating the lignocellulose raw material is as follows: after the lignocellulose raw material is crushed and impurities are removed, it is subjected to acid treatment, alkali treatment or steam explosion treatment to obtain the pretreated material.

4. The method of co-producing organic acids and lignin from a lignocellulosic feedstock according to claim 1, wherein, In the step two, the enzyme used for enzymatic hydrolysis is cellulase.

5. The method of lignocellulose co-production of organic acids and lignin according to claim 1, characterized in that, In the step four, the conditions for acid production fermentation are as follows: Batch fermentation, semi-continuous fermentation or continuous fermentation under anaerobic conditions; and / or, The pH of acid production fermentation is 5-10; The temperature of acid production fermentation is 30-58℃; The C / N ratio is 20-60; The fermentation time is 2-50 days.

6. The method of lignocellulose co-production of organic acids and lignin according to claim 5, characterized in that, In the step four, the conditions for acid production fermentation are as follows: The pH of acid production fermentation is 5-7; The temperature of acid production fermentation is 35-40℃; The C / N ratio is 30-40; The fermentation time is 7-14 days.

7. The method of lignocellulose co-production of organic acids and lignin according to claim 5, characterized in that, The nitrogen source is one or a combination of NH4Cl, corn syrup, sludge, potassium nitrate and urea.

8. The method for co-producing organic acid and lignin from lignocellulose according to claim 1, wherein F1 stage: The running time of the F1 stage is 80-120 days; The TS value of the substrate is 4.8-5.4%; the VTS value is 4.0-4.4%; the substrate comprises a mixture of straw, kitchen waste and nitrogen source substrate; the material is fed and discharged once every two days; the hydraulic retention time of the reactor is 90-110 days; wherein TS is total solid concentration; F2 stage: The running time of the F2 stage is 50-100 days; the substrate TS is 9.8-10.5%, and the substrate VTS is 8.0-8.5%; the hydraulic retention time of the reactor is 90-110 days; The F3 stage: The running time of the F3 stage is 120-200 days; the hydraulic retention time of the reactor is adjusted to 45-55 days, the substrate TS is 9.5-10%, and the substrate VTS is 8.0-8.6%.

9. The method for co-producing organic acid and lignin from lignocellulose according to claim 8, characterized in that, The F1 stage: The VTS ratio of the straw, kitchen waste and nitrogen source substrate in the substrate is (0.8-1.2):(0.8-1.2):(0.8-1.2); the nitrogen source substrate is residual sludge from a sewage plant; and the straw is steam explosion straw; The F2 stage: The VTS ratio of the straw, kitchen waste and nitrogen source substrate in the substrate is (0.8-1.2):(0.8-1.2):(0.8-1.2); The F3 stage: The VTS ratio of the straw, kitchen waste and nitrogen source substrate in the substrate is changed from (0.8-1.2):(0.8-1.2):(0.8-1.2) to (2.8-3.2):(1.8-2.2):(0.8-1.2).

10. The method of lignocellulose co-production of organic acids and lignin according to claim 8, characterized in that, The inoculum source is pretreated by heating at 75-85℃ for 0.5-1.2 hours and adding a methane inhibitor with a final concentration of 4-6 mM; the methane inhibitor is selected from 2-bromoethanesulfonic acid.

11. The method for co-producing organic acid and lignin from lignocellulose according to claim 8, characterized in that, The inoculum source comprises inocula of different sources; the inocula of different sources are selected from a mixture of anaerobic digestion sludge from a grain sugar factory, anaerobic digestion sludge from kitchen waste, cellulose anaerobic digestion sludge, glucose anaerobic digestion sludge and acid production reactor sludge from anaerobic fermentation of fruit waste.

12. The method for co-producing organic acid and lignin from lignocellulose according to claim 11, characterized in that, The cellulose anaerobic digestion sludge is selected from cellulose high-temperature anaerobic digestion sludge or cellulose mesophilic anaerobic digestion sludge; and / or, The glucose anaerobic digestion sludge is selected from glucose high-temperature anaerobic digestion sludge or glucose mesophilic anaerobic digestion sludge.

13. The method of lignocellulose co-production of organic acids and lignin according to claim 11, characterized in that, The inoculum source comprises the inocula of different sources mixed according to the total solid concentration, and each sludge accounts for more than 10% of the total solid concentration.

14. The method of lignocellulose co-production of organic acids and lignin according to claim 13, characterized in that, Each sludge is mixed according to the total solid concentration in equal proportions.

15. The method of lignocellulose co-production of organic acids and lignin according to claim 11, characterized in that, The reactor is a high-temperature reactor or a mesophilic reactor.

16. The method of lignocellulose co-production of organic acids and lignin according to claim 15, characterized in that, The pH value of the reactor is 5.5-6.0; and / or, the working temperature of the high-temperature reactor is 50-55℃; and / or, the working temperature of the mesophilic reactor is 35-40℃.

17. The method for co-producing organic acid and lignin from lignocellulose according to claim 15, characterized in that, When the reactor is a high-temperature reactor, the cellulose anaerobic digestion sludge is selected from cellulose high-temperature anaerobic digestion sludge, and the glucose anaerobic digestion sludge is selected from glucose high-temperature anaerobic digestion sludge. When the reactor is a mesophilic reactor, the cellulose anaerobic digestion sludge is selected from a cellulose mesophilic anaerobic digestion sludge and the glucose anaerobic digestion sludge is selected from a glucose mesophilic anaerobic digestion sludge.

18. The method of lignocellulose co-production of organic acids and lignin according to claim 1, characterized in that, The organic acid is a plurality of combinations of lactic acid, acetic acid, propionic acid, isobutyric acid, butyric acid, valeric acid, and isovaleric acid.

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