Process for preparing lactic acid by mechanically enhanced ammonia process pretreatment of straw biomass

Abstract

The present invention discloses a process for preparing lactic acid by mechanically enhanced ammonia method from straw-like biomass. The enzymatic hydrolysate of solid fibers obtained by the mechanically enhanced ammonia method from straw-like biomass is used as the carbon source of the fermentation medium in the process of fermenting lactic acid. The preparation of the carbon source in the fermentation medium comprises the following steps: S1: treating straw-like lignocellulosic biomass with a pretreatment liquid containing ammonia water and inorganic ammonium salts in a twin-screw device; S2: subjecting the obtained material to a heat preservation reaction, and after solid-liquid separation, obtaining solid fibers containing holocellulose; S3: enzymatically hydrolyzing the obtained solid fibers with Trichoderma reesei cellulase and xylanase, and performing solid-liquid separation, and the obtained supernatant is used as the carbon source in the fermentation medium for fermenting L-lactic acid. The present invention uses the sugar liquid obtained by enzymatic hydrolysis and saccharification of pretreated holocellulose, inoculates Bacillus coagulans to ferment L-lactic acid after supplementing nitrogen source and inorganic salts, and can greatly improve the concentration and conversion rate of L-lactic acid.

Description

Technical Field

[0001] The present invention relates to the fields of biochemistry and bioenergy, and particularly relates to a process for the mechanical enhanced ammonia pretreatment of straw biomass, and a process for preparing lactic acid from straw biomass by mechanical enhanced ammonia pretreatment. Background Art

[0002] With the exhaustion of global fossil energy, and the use of fossil energy will release greenhouse gases, toxic substances and standard pollutants, thus leading to environmental pollution and adverse climate change impacts. Therefore, it is urgent to develop renewable and sustainable green resources.

[0003] As the most abundant renewable resource on the earth, lignocellulosic biomass is mainly composed of cellulose, hemicellulose, lignin and a small amount of other minor components. Among them, cellulose and hemicellulose can be degraded into monosaccharides to provide a carbon source for lactic acid fermentation, effectively reducing the environmental pollution problems caused by the manufacture of petroleum-derived chemicals, and reducing the costs and food competition problems brought about by the fermentation of starch in food crops (such as rice, cassava or sorghum). However, lignin, which is composed of guaiacyl, syringyl and p-hydroxyphenyl subunits, is connected by C-O, C-C and H bonds as well as van der Waals forces to form a stubborn, crystalline three-dimensional structure that resists microbial digestion. Through pretreatment technology, the complex and stubborn structure of lignocellulose itself can be effectively destroyed, so as to realize the efficient utilization of lignocellulose.

[0004] The technologies for pretreating lignocellulosic biomass mainly include physical, chemical and biological pretreatments, etc. For example, Chinese Invention Application CN105296329A discloses a device and method for continuously pretreating lignocellulosic materials by microwave-dilute acid steam explosion. However, this method has high temperature and pressure during the pretreatment process, increasing the energy consumption cost, and at the same time having high requirements for the acid and pressure resistance of the equipment; Chinese Invention Application CN110760552A discloses a pretreatment method for improving the saccharification efficiency of lignocellulose. However, this method has a long pretreatment period, high production cost and complex process; Chinese Invention Application CN103993053A discloses a method for coupling water and ammonia to pretreat biomass. However, this method has high energy consumption and discontinuous process; Chinese Invention Application CN103255659A discloses a method for treating lignocellulosic biomass by ammonia combined with dilute alkali under normal pressure. However, this method uses a large amount of chemical reagents, and at the same time generates a large amount of high-salt wastewater, and does not recover the removed lignin components, which is contrary to the concept of green chemistry. The mechanical enhanced ammonia pretreatment can effectively remove the lignin components, get rid of the traditional batch reaction system, and reduce the energy consumption during the pretreatment process. At the same time, under normal temperature and pressure conditions, the generation of by-products during the pretreatment process can be effectively reduced and the equipment requirements can be reduced.

[0005] Therefore, the development of economic, efficient, green, and continuous pretreatment methods remains a key challenge for lignocellulose conversion technologies. Summary of the Invention

[0006] Object of the Invention: The technical problem to be solved by the present invention is to provide a twin-screw pretreatment process for straw-like lignocellulosic biomass in view of the deficiencies of the prior art, so as to solve the problems that sugar is degraded into by-products during the previous pretreatment process, affecting the saccharification yield and producing microbial fermentation inhibitors, as well as the polymerization problem of lignin under harsh conditions of high temperature and high pressure, and to provide a good biological substrate for subsequent enzymatic hydrolysis and fermentation to produce L-lactic acid.

[0007] Another technical problem to be solved by the present invention is to provide a process for preparing lactic acid by mechanically enhanced ammonia pretreatment of straw-like biomass.

[0008] To solve the above first technical problem, the present invention discloses a process for mechanically enhanced ammonia pretreatment of straw-like biomass, comprising the following steps:

[0009] S1: Treating straw-like lignocellulosic biomass with a pretreatment liquid containing ammonia water and inorganic ammonium salts in a twin-screw device;

[0010] S2: Subjecting the material obtained in step S1 to a heat preservation reaction, and after solid-liquid separation, obtaining solid fibers containing holocellulose and black liquor containing aminated lignin.

[0011] In step S1, the straw-like lignocellulosic biomass includes, but is not limited to, any one or several combinations of wheat straw, corn straw, rice straw, sorghum straw, bagasse, corn cob, switchgrass, reed, and rapeseed.

[0012] In step S1, the particle size of the straw-like lignocellulosic biomass is 3 - 5 cm.

[0013] In step S1, the pretreatment liquid is an aqueous solution containing 5% - 30% w / w ammonia water and 0.1% - 2% w / w inorganic ammonium salts, preferably an aqueous solution containing 20% - 30% w / w ammonia water and 0.5% - 1.5% w / w inorganic ammonium salts, and further preferably an aqueous solution containing 25% w / w ammonia water and 1% w / w inorganic ammonium salts.

[0014] In step S1, the inorganic ammonium salt is any one or several combinations of sodium ammonium hydrogen phosphate, ammonium carbonate, ammonium bicarbonate, ammonium sulfate, and ammonium chloride, preferably sodium ammonium hydrogen phosphate.

[0015] In step S1, the dosage of the pretreatment liquid is 2 - 10 times the mass of the dry straw-like lignocellulosic biomass.

[0016] In step S1, the processing parameters of the twin-screw equipment are as follows: the screw rotation speed is 100 - 300 r / min, preferably 200 r / min; the feeding speed is 15 - 25 kg / h, preferably 20 kg / h.

[0017] In step S2, the temperature of the heat preservation reaction is 80 - 140 °C.

[0018] In step S2, the time of the heat preservation reaction is 0.5 - 5 h.

[0019] Among them, through the above process, the retention rate of cellulose in the fixed component is more than 65%, the retention rate of hemicellulose in the fixed component is more than 65%, and the content of lignin in the black liquor is more than 50%.

[0020] In order to solve the above second technical problem, the present invention discloses a process for preparing lactic acid by mechanical enhanced ammonia pretreatment of straw-like biomass, using the enzymatic hydrolysate of solid fibers obtained by the process of mechanical enhanced ammonia pretreatment of straw-like lignocellulosic biomass as the carbon source of the fermentation medium in the process of fermenting lactic acid.

[0021] Among them, the preparation method of the carbon source in the fermentation medium includes the following steps:

[0022] S1: Treat straw-like lignocellulosic biomass with a pretreatment solution containing ammonia water and inorganic ammonium salts in a twin-screw equipment;

[0023] S2: Carry out a heat preservation reaction on the material obtained in step S1, after solid-liquid separation, obtain solid fibers containing holocellulose and black liquor containing aminated lignin;

[0024] S3: In a buffer solution, enzymatically hydrolyze the solid fibers obtained in step S2 with Trichoderma reesei cellulase and xylanase, carry out solid-liquid separation, and use the obtained supernatant as the carbon source in the fermentation medium for fermenting L-lactic acid.

[0025] In step S1, the straw-like lignocellulosic biomass includes but is not limited to any one or several combinations of wheat straw, corn straw, rice straw, sorghum straw, sugarcane bagasse, corn cob, switchgrass, reed, and rapeseed.

[0026] In step S1, the particle size of the straw-like lignocellulosic biomass is 3 - 5 cm.

[0027] In step S1, the pretreatment solution is an aqueous solution containing 5% - 30% w / w ammonia water and 1% - 20% w / w inorganic ammonium salts, preferably an aqueous solution containing 20% - 30% w / w ammonia water and 5% - 15% w / w inorganic ammonium salts, and further preferably an aqueous solution containing 25% w / w ammonia water and 10% w / w inorganic ammonium salts.

[0028] In step S1, the inorganic ammonium salt is any one or a combination of several of sodium ammonium hydrogen phosphate, ammonium carbonate, ammonium bicarbonate, ammonium sulfate, and ammonium chloride, preferably sodium ammonium hydrogen phosphate.

[0029] In step S1, the dosage of the pretreatment liquid is 2 to 10 times the mass of the dried straw-like lignocellulosic biomass.

[0030] In step S1, the processing parameters of the twin-screw equipment are: screw rotation speed 100 - 300 r / min, preferably 200 r / min; feeding speed 15 - 25 kg / h, preferably 20 kg / h.

[0031] In step S2, the temperature of the heat preservation reaction is 80 - 140 °C.

[0032] In step S2, the time of the heat preservation reaction is 0.5 - 5 h.

[0033] In step S3, the buffer solution is 5 mM sodium citrate buffer solution.

[0034] In step S3, the addition amount of Trichoderma reesei cellulase is 10 - 30 FPU / g solid fiber, preferably 20 FPU / g solid fiber.

[0035] In step S3, the addition amount of xylanase is 40 - 60 IU / g solid fiber, preferably 50 IU / g solid fiber.

[0036] In step S3, the dosage ratio of the solid fiber to the buffer solution is 1:(5 - 15) g / L, preferably 1:10 g / L.

[0037] In step S3, the temperature of the enzymatic hydrolysis is 40 - 60 °C, preferably 50 °C; the time of the enzymatic hydrolysis is 36 - 60 h, preferably 40 - 56 h, and further preferably 48 h.

[0038] Among them, the fermentation medium further includes a nitrogen source; the nitrogen source is yeast extract and / or corn steep liquor dry powder, preferably yeast extract and corn steep liquor dry powder; preferably, yeast extract and corn steep liquor dry powder are added so that their concentrations are 1 / 14 - 1 / 34 and 1 / 40 - 1 / 60 of the glucose concentration in the supernatant respectively; preferably, yeast extract and corn steep liquor dry powder are added so that their concentrations are 1 / 18 - 1 / 25 and 1 / 48 - 1 / 52 of the glucose concentration in the supernatant respectively.

[0039] Among them, the fermentation medium further includes inorganic salts; the inorganic salts are ammonium sulfate, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, magnesium sulfate monohydrate, and ferrous sulfate heptahydrate; preferably, the concentrations of each component are as follows: ammonium sulfate 2-4 g / L, potassium dihydrogen phosphate 0.12-0.32 g / L, magnesium sulfate heptahydrate 0.3-0.5 g / L, manganese sulfate monohydrate 0.01-0.05 g / L, ferrous sulfate heptahydrate 0.01-0.05 g / L; preferably, the concentrations of each component are as follows: ammonium sulfate 3 g / L, potassium dihydrogen phosphate 0.22 g / L, magnesium sulfate heptahydrate 0.4 g / L, manganese sulfate monohydrate 0.03 g / L, ferrous sulfate heptahydrate 0.03 g / L.

[0040] Among them, the method for fermenting L-lactic acid is to inoculate the Bacillus coagulans bacterial solution into the above fermentation medium and ferment to obtain a fermentation broth containing L-lactic acid.

[0041] Among them, the Bacillus coagulans is Bacillus coagulans ACCC 10229, and this bacterium is an existing bacterium.

[0042] Among them, the Bacillus coagulans bacterial solution is inoculated into the above fermentation medium at a volume ratio of 5%-20%, preferably 10% by volume.

[0043] Among them, the temperature of the fermentation is 40-60 °C, preferably 50 °C; the rotation speed of the fermentation is 100-200 rpm, preferably 150 rpm; the fermentation time is 62-82 h, preferably 72 h.

[0044] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0045] 1. The pretreatment liquid of the present invention is composed of a compound of ammonia water and inorganic ammonium salts. During the pretreatment operation, the physical chemical bond between lignin and holocellulose is effectively cut, the removal rate of lignin under mild conditions is improved, and the condensation of lignin under conventional high-temperature and high-pressure cooking conditions is avoided, and the lignin component with a complete structure is extracted; at the same time, the retention rates of cellulose and hemicellulose are also relatively high.

[0046] 2. The present invention uses the sugar liquid obtained by enzymatic hydrolysis and saccharification of pretreated holocellulose, supplements the nitrogen source and inorganic salts, and then inoculates Bacillus coagulans to ferment L-lactic acid, which can greatly increase the concentration and conversion rate of L-lactic acid.

[0047] 3. The present invention realizes a continuous treatment process, reduces the water consumption in the high solid-liquid ratio process, and reduces the external heating energy consumption, thereby reducing the production cost in many aspects.

[0048] 4. The present invention realizes the continuous operation of the pretreatment system, gets rid of the traditional batch reaction system, and improves the production efficiency.

[0049] 5. The continuous pretreatment process of the present invention enables the efficient recovery of holocellulose in straw for use in the L-lactic acid fermentation reaction after further enzymatic hydrolysis and saccharification. At the same time, lignin components with intact structures can be efficiently recovered, realizing the efficient utilization of each component of lignocellulose. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] The following further detailed description of the present invention will be made in conjunction with the accompanying drawings and specific embodiments, and the above and / or other advantages of the present invention will become clearer.

[0051] Figure 1 It is the two-dimensional NMR of lignin.

[0052] Figure 2 It is the micrograph of holocellulose. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0053] In the following embodiments, the experimental methods are all conventional methods unless otherwise specified; the reagents and materials can be obtained from commercial sources unless otherwise specified.

[0054] In the following embodiments, the detection method for the chemical composition analysis of biomass adopts the method of the National Renewable Energy Laboratory (NREL), and the detection steps are as follows:

[0055] 1) Add 300 ± 10 mg to 3.00 ± 0.01 mL (or 4.92 ± 0.01 g) of 72% sulfuric acid, mix well and place in a water bath shaker at 30 ± 3 for 60 ± 5 min;

[0056] 2) Add 84.00 ± 0.04 mL of deionized water to dilute the acid to a concentration of 4%. Place it in an autoclave and treat it at 121 °C for 1 h. After the autoclave cycle is completed, let the hydrolysate cool slowly to near room temperature, and then remove the lid;

[0057] 3) Vacuum filter the autoclave hydrolysis solution through a pre-weighed filter crucible, collect the filtrate for the determination of acid-soluble lignin and carbohydrates;

[0058] 4) Rinse the solid with at least 50 mL of fresh deionized water, dry the crucible and acid-insoluble residue at 105 ± 3 °C until a constant weight is reached. Take the sample out of the oven and cool it in a desiccator. Then place the crucible and residue in a muffle furnace at 575 ± 25 °C for 24 ± 6 h.

[0059] In the following embodiments, the lignocellulose raw material used is wheat straw, and the cellulose content in wheat straw is 34.56 wt%, the hemicellulose content is 21.46 wt%, and the lignin content is 23.13 wt%.

[0060] In the following examples, the enzyme activities of Trichoderma reesei cellulase and xylanase added are 260 FPU / ml and 50 IU / ml, respectively.

[0061] The activity of Trichoderma reesei cellulase is defined as: the amount of enzyme required to release 1 μmol of reducing sugar per minute, expressed in FPU / mL;

[0062] The activity of xylanase is defined as: the number of μmol of reducing sugar produced per minute by the crude enzyme used per milliliter, expressed in IU / mL.

[0063] The preparation of Bacillus coagulans ACCC 10229 bacterial liquid in the following examples is as follows:

[0064] 1) Take the glycerol strain of Bacillus coagulans ACCC 10229 stored at -80 °C, add a liquid seed medium 20 times the volume, and no calcium carbonate is added to this medium. Culture at 50 °C and 150 rpm for 12 h, denoted as bacteria 1;

[0065] 2) Streak plate inoculate bacteria 1 cultured at 50 °C and 150 rpm for 12 h onto a solid seed medium containing calcium carbonate, and the calcium carbonate concentration is 1 / 2 of the glucose concentration. Culture at 50 °C and 150 rpm for 12 - 24 h, denoted as bacteria 2;

[0066] 3) Inoculate the single colony grown from bacteria 2 into a liquid seed medium containing calcium carbonate, and the calcium carbonate concentration is 1 / 2 of the glucose concentration. Culture at 50 °C and 150 rpm for 12 h to obtain the Bacillus coagulans ACCC 10229 bacterial liquid.

[0067] The formula of the liquid seed medium is as follows: glucose 20 g / L, yeast extract 2 g / L, corn steep liquor dry powder 2.5 g / L, ammonium chloride 1 g / L, anhydrous magnesium sulfate 0.2 g / L, adjust the pH to 7.0, sterilize at 115 °C for 15 min.

[0068] The formula of the solid seed medium is as follows: glucose 20 g / L, yeast extract 2 g / L, corn steep liquor dry powder 2.5 g / L, ammonium chloride 1 g / L, anhydrous magnesium sulfate 0.2 g / L, agar powder 16 g / L, adjust the pH to 7.0, sterilize at 115 °C for 15 min.

[0069] The calculation of the yield of L-lactic acid in the following examples is as follows:

[0070]

[0071] C L-LA —— The concentration of L-lactic acid at the end of fermentation

[0072] C G0 —— The initial glucose concentration of fermentation

[0073] C G72 —— Glucose concentration 72 h after fermentation end

[0074] 1.04 - Correction coefficient of L-lactic acid.

[0075] The processing parameters of the twin-screw in the following examples are: screw rotation speed 200 r / min, feeding speed 20 kg / h.

[0076] Example 1

[0077] Take the dry wheat straw raw material and mechanically crush it into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnation liquid used is tap water, and it is heated to 90 °C. Start the twin-screw device, and continuously transport the heated impregnation liquid and wheat straw segments to the twin-screw through the feeding hopper at the same time. During this period, pump in a pretreatment liquid 7 times the mass of the dry material, which contains 25% W / W ammonia water and 1.0% W / W sodium hydrogen phosphate. Keep the wheat straw at 130 °C in the screw for 1 h by adjusting the frequency of the twin-screw motor. Transport the material after the heat preservation reaction to a single-screw extruder for solid-liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0078] After calculation, the retention rate of the obtained solid cellulose after pretreatment is 91.03%, the retention rate of hemicellulose is 71.30%, and the yield of lignin in the black liquor is 67.49%.

[0079] Figure 1 is the NMR characterization diagram of lignin in the obtained alkaline black liquor. Among them, feruloylamide (FA6, δC / δH 123.1 / 7.20) is identified in the lignin, which indicates that ferulic acid connected by an ester bond has grafted nitrogen. Figure 2 is the micrograph of the obtained holocellulose. From Figure 1 it can be seen that the lignin component structure obtained by pretreatment with twin-screw and ammonia water is intact. From Figure 2 it can be seen that the holocellulose components obtained by pretreatment with twin-screw and ammonia water have a fluffy structure, showing a complete fibril-like structure and no obvious fragmentation state.

[0080] Preparation of L-lactic acid from straw:

[0081] (a) Mix the above-obtained holocellulose with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and carry out enzymatic hydrolysis and saccharification at 50 °C for 48 h, and then carry out solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysate sugar solution.

[0082] (b) The conversion rate of cellulose in the above enzymatic hydrolysate was detected by HPLC to be 92.50%, and the glucose concentration was 59.09 g / L. Based on the glucose concentration, 2.46 g / L of yeast extract and 1.18 g / L of corn steep liquor dry powder were added to the enzymatic hydrolysate as nitrogen sources for the fermentation medium, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0083] (c) The above-prepared mixture was sterilized at 115 °C for 15 min.

[0084] (d) The Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm was inoculated into the above-prepared mixture at a volume ratio of 10%, and fermented at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0085] The above lactic acid fermentation broth was detected by HPLC to obtain L-lactic acid produced by fermenting straw-type lignocellulose as the raw material. The concentration of L-lactic acid produced in the sugar solution was detected by HPLC to be 53.01 g / L, and the conversion rate of sugar fermentation to L-lactic acid was 98.01%.

[0086] Example 2

[0087] Take the dry wheat straw raw material and mechanically crush it into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin-screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin-screw through the feed hopper at the same time. During this period, 7 times the mass of the dry material of the pretreatment solution is pumped in, which contains 25% W / W ammonia water and 1.0% W / W sodium hydrogen phosphate. By adjusting the frequency of the twin-screw motor, the wheat straw is kept at 90 °C in the screw for a reaction for 3 h. The material after the heat preservation reaction is transported to a single-screw extrusion machine for solid-liquid separation, and then holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin are obtained.

[0088] After calculation, the retention rate of the obtained solid cellulose after pretreatment is 86.44%, the retention rate of hemicellulose is 67.41%, and the yield of lignin in the black liquor is 58.31%.

[0089] Preparation of straw lactic acid fermentation:

[0090] (a) Mix the above-obtained holocellulose with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and carry out enzymatic hydrolysis and saccharification at 50 °C for 48 h. Then, perform solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysis sugar solution.

[0091] (b) The conversion rate of cellulose in the above enzymatic hydrolysis sugar solution detected by HPLC is 88.31%, and the concentration of glucose is 56.49 g / L. Based on the glucose concentration, add 2.35 g / L of yeast extract and 1.13 g / L of corn steep liquor dry powder as the nitrogen source of the fermentation medium to the enzymatic hydrolysis sugar solution, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0092] (c) Sterilize the above compounded mixture at 115 °C for 15 min.

[0093] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm into the above compounded mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0094] Perform HPLC detection on the above lactic acid fermentation broth to obtain L-lactic acid produced by fermenting straw-type lignocellulose as raw material. The concentration of L-lactic acid produced in the sugar solution detected by HPLC is 48.75 g / L, and the conversion rate of sugar fermentation to L-lactic acid is 89.47%.

[0095] Example 3

[0096] Take the dried wheat straw raw material and mechanically crush it into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin-screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin-screw through the feeding hopper at the same time. During this period, pump in 3 times the mass of the dry material of the pretreatment solution, which contains 25% W / W ammonia water and 1.0% W / W sodium hydrogen phosphate. Adjust the frequency of the twin-screw motor to keep the wheat straw at 130 °C in the screw for 3 h. Transport the material after the heat preservation reaction to a single-screw extruder for solid-liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0097] After calculation, the retention rate of the obtained solid cellulose after pretreatment is 86.58%, the retention rate of hemicellulose is 71.57%, and the yield of lignin in the black liquor is 57.85%.

[0098] Preparation of straw lactic acid fermentation:

[0099] (a) Mix the above-mentioned holocellulose with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g of holocellulose and 50 IU / g of holocellulose respectively, and carry out enzymatic hydrolysis and saccharification at 50 °C for 48 h, then carry out solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysate sugar solution.

[0100] (b) The conversion rate of cellulose in the above enzymatic hydrolysate sugar solution detected by HPLC is 90.09%, and the concentration of glucose is 54.83 g / L. Based on the glucose concentration, add 2.28 g / L of yeast extract and 1.10 g / L of corn steep liquor dry powder as the nitrogen source of the fermentation medium to the enzymatic hydrolysate sugar solution, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0101] (c) Sterilize the above-mentioned compounded mixture at 115 °C for 15 min.

[0102] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm into the above-mentioned compounded mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0103] Perform HPLC detection on the above lactic acid fermentation broth to obtain L-lactic acid produced by fermenting straw-type lignocellulose as raw material. The concentration of L-lactic acid produced in the sugar solution detected by HPLC is 50.79 g / L, and the conversion rate of sugar fermentation to L-lactic acid is 96.13%.

[0104] Example 4

[0105] Take dry wheat straw raw materials and mechanically crush them into 3-5 cm segments as lignocellulose raw materials for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin-screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin-screw through the feed hopper at the same time. During this period, pump in 3 times the mass of the dry material of the pretreatment solution, which contains 25% W / W ammonia water and 1.5% W / W sodium hydrogen phosphate. Adjust the frequency of the twin-screw motor to keep the wheat straw at 90 °C in the screw for 1 h. Transport the material after the heat preservation reaction to a single-screw extruder for solid-liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0106] After calculation, the retention rate of the solid cellulose obtained after pretreatment is 85.94%, the retention rate of hemicellulose is 65.92%, and the yield of lignin in the black liquor is 51.13%.

[0107] Preparation of straw lactic acid fermentation:

[0108] (a) Mix the above-obtained holocellulose with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and enzymatically hydrolyze and saccharify at 50 °C for 48 h, then perform solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatically hydrolyzed sugar solution.

[0109] (b) HPLC detection shows that the conversion rate of cellulose in the above enzymatically hydrolyzed sugar solution is 88.35%, the concentration of glucose is 54.93 g / L. Based on the glucose concentration, add 2.39 g / L of yeast extract and 1.10 g / L of corn steep liquor dry powder as the nitrogen source of the fermentation medium to the enzymatically hydrolyzed sugar solution, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0110] (c) Sterilize the above-prepared mixture at 115 °C for 15 min.

[0111] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm into the above-prepared mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0112] Perform HPLC detection on the above lactic acid fermentation broth to obtain L-lactic acid produced by fermenting straw-type lignocellulose as the raw material. The concentration of L-lactic acid produced in the sugar solution detected by HPLC is 44.59 g / L, and the conversion rate of sugar fermentation to L-lactic acid is 84.25%.

[0113] Example 5

[0114] Take the dry wheat straw raw material and mechanically crush it into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin - screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin - screw through the feeding hopper simultaneously. During this period, pump in the pretreatment solution which is 3 times the mass of the dry material, containing 15% W / W ammonia water and 1.5 W / W sodium hydrogen phosphate. Adjust the frequency of the twin - screw motor to keep the wheat straw at 90 °C in the screw for a reaction of 1 h. Transport the material after the heat - preservation reaction to a single - screw extruder for solid - liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0115] After calculation, the retention rate of the obtained solid cellulose after pretreatment is 90.11%, the retention rate of hemicellulose is 67.72%, and the yield of lignin in the black liquor is 56.22%.

[0116] Preparation of straw lactic acid fermentation:

[0117] (a) Mix the above - obtained holocellulose with 5 mM sodium citrate buffer at a solid - to - liquid ratio of 1:10 g / L. Add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and carry out enzymatic hydrolysis and saccharification at 50 °C for 48 h. Then carry out solid - liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysis sugar solution.

[0118] (b) HPLC detects that the conversion rate of cellulose in the above - mentioned enzymatic hydrolysis sugar solution is 86.29%, and the concentration of glucose is 56.57 g / L. Based on the glucose concentration, add 2.36 g / L of yeast extract and 1.13 g / L of corn steep liquor dry powder as the nitrogen source of the fermentation medium, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0119] (c) Sterilize the above - compounded mixture at 115 °C for 15 min.

[0120] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm into the above - compounded mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L - lactic acid.

[0121] Perform HPLC detection on the above - mentioned lactic acid fermentation broth to obtain L - lactic acid fermented from straw - type lignocellulose as the raw material. The concentration of L - lactic acid produced in the sugar solution detected by HPLC is 47.46 g / L, and the conversion rate of sugar fermentation to L - lactic acid is 86.97%.

[0122] Example 6

[0123] Take the dry wheat straw raw material and mechanically crush it into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin - screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin - screw through the feeding hopper simultaneously. During this period, pump in a pretreatment solution that is 3 times the mass of the dry material, containing 15% W / W ammonia water and 1.0 W / W sodium hydrogen phosphate. Adjust the frequency of the twin - screw motor to keep the wheat straw at 130 °C in the screw for 1 h for reaction. Transport the material after the heat - preservation reaction to a single - screw extruder for solid - liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0124] After calculation, the retention rate of the solid cellulose obtained after pretreatment is 87.31%, the retention rate of hemicellulose is 66.65%, and the yield of lignin in the black liquor is 53.43%.

[0125] Preparation of straw lactic acid fermentation:

[0126] (a) Mix the above - obtained holocellulose with 5 mM sodium citrate buffer at a solid - to - liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and carry out enzymatic hydrolysis and saccharification at 50 °C for 48 h. Then carry out solid - liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysis sugar solution.

[0127] (b) HPLC detects that the conversion rate of cellulose in the above - mentioned enzymatic hydrolysis sugar solution is 90.21%, and the concentration of glucose is 58.43 g / L. Based on the glucose concentration, add 2.43 g / L of yeast extract and 1.17 g / L of corn steep liquor dry powder as the nitrogen source of the fermentation medium, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0128] (c) Sterilize the above - compounded mixture at 115 °C for 15 min.

[0129] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm into the above - compounded mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L - lactic acid.

[0130] The above lactic acid fermentation broth was detected by HPLC, and L-lactic acid produced by fermenting straw-type lignocellulose as raw material was obtained. The concentration of L-lactic acid produced in the sugar solution detected by HPLC was 49.12 g / L, and the conversion rate of sugar fermentation to L-lactic acid was 87.05%.

[0131] Example 7

[0132] The dry wheat straw raw material was mechanically crushed into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnating solution used was tap water, and it was heated to 90 °C. The twin-screw device was started, and the heated impregnating solution and wheat straw segments were continuously transported to the twin-screw through the feed hopper simultaneously. During this period, the pretreatment solution, which was 7 times the mass of the dry material, was pumped in, containing 15% W / W ammonia water and 1.5 W / W sodium hydrogen phosphate. By adjusting the frequency of the twin-screw motor, the wheat straw was kept at 130 °C in the screw for a reaction for 3 h. The material after the heat preservation reaction was transported to a single-screw extruder for solid-liquid separation, and thus holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin were obtained.

[0133] After calculation, the retention rate of the solid cellulose obtained after pretreatment was 82.42%, the retention rate of hemicellulose was 65.96%, and the yield of lignin in the black liquor was 60.03%.

[0134] Preparation of straw lactic acid fermentation:

[0135] (a) The above-obtained holocellulose was mixed with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, and Trichoderma reesei cellulase and xylanase were added at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and enzymatically hydrolyzed and saccharified at 50 °C for 48 h, and then solid-liquid separation was carried out at 10000 rpm for 5 min. The obtained supernatant was the enzymatically hydrolyzed sugar solution.

[0136] (b) The conversion rate of cellulose in the above enzymatically hydrolyzed sugar solution detected by HPLC was 86.80%, the concentration of glucose was 58.15 g / L. Based on the glucose concentration, 2.42 g / L of yeast extract and 1.16 g / L of corn steep liquor dry powder were added to the enzymatically hydrolyzed sugar solution as the nitrogen source of the fermentation medium, and 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate were added as inorganic salts.

[0137] (c) The above-prepared mixture was sterilized at 115 °C for 15 min.

[0138] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial liquid with an absorbance of 1.0 at OD600nm into the above compound mixture at a volume ratio of 10%, and ferment at 50°C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0139] Perform HPLC detection on the above lactic acid fermentation broth to obtain L-lactic acid fermented and produced using straw-type lignocellulose as the raw material. The concentration of L-lactic acid produced in the sugar solution detected by HPLC is 43.92 g / L, and the conversion rate of sugar fermentation to L-lactic acid is 78.22%.

[0140] Example 8

[0141] Take the dry wheat straw raw material and mechanically crush it into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnating solution used is tap water, and it is heated to 90°C. Start the twin-screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin-screw through the feed hopper at the same time. During this period, pump in 7 times the mass of the dry material of the pretreatment solution, which contains 15% W / W ammonia water and 1.5 W / W sodium hydrogen phosphate. Adjust the frequency of the twin-screw motor to keep the wheat straw at 90°C in the screw for a reaction for 3 h. Transport the material after the heat preservation reaction to a single-screw pulp extruder for solid-liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0142] After calculation, the retention rate of the obtained solid cellulose after pretreatment is 84.00%, the retention rate of hemicellulose is 65.52%, and the yield of lignin in the black liquor is 61.08%.

[0143] Preparation of straw lactic acid fermentation:

[0144] (a) Mix the above-obtained holocellulose with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and perform enzymatic hydrolysis and saccharification at 50°C for 48 h, and then perform solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysis sugar solution.

[0145] (b) The conversion rate of cellulose in the above enzymatic hydrolysis sugar solution detected by HPLC is 84.95%, and the concentration of glucose is 52.93 g / L. Based on the glucose concentration, add 2.21 g / L of yeast extract and 1.06 g / L of corn steep liquor dry powder as the nitrogen source of the fermentation medium to the enzymatic hydrolysis sugar solution, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0146] (c) Sterilize the above-mentioned compounded mixture at 115 °C for 15 min.

[0147] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial liquid with an absorbance of 1.0 at OD600nm into the above-mentioned compounded mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0148] Perform HPLC detection on the above-mentioned lactic acid fermentation broth to obtain L-lactic acid produced by fermenting straw-type lignocellulose as raw material. The concentration of L-lactic acid produced in the sugar solution detected by HPLC is 41.16 g / L, and the conversion rate of sugar fermentation to L-lactic acid is 80.00%.

[0149] Example 9

[0150] Take dry wheat straw raw materials and mechanically crush them into 3-5 cm segments as lignocellulose raw materials for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin-screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin-screw through the feed hopper at the same time. During this period, pump in a pretreatment solution 7 times the mass of the dry material, including 15% W / W ammonia water and 0.5 W / W sodium hydrogen phosphate. Adjust the frequency of the twin-screw motor to keep the wheat straw at 130 °C in the screw for 1 h. Transport the material after the heat preservation reaction to a single-screw pulp extruder for solid-liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0151] After calculation, the retention rate of the obtained solid cellulose after pretreatment is 87.35%, the retention rate of hemicellulose is 67.69%, and the yield of lignin in the black liquor is 56.85%.

[0152] Preparation of straw lactic acid fermentation:

[0153] (a) Mix the above-obtained holocellulose with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and perform enzymatic hydrolysis and saccharification at 50 °C for 48 h, and then perform solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysis sugar solution.

[0154] (b) The conversion rate of cellulose in the above enzymatic hydrolysate was detected by HPLC to be 82.98%, and the concentration of glucose was 54.52 g / L. Based on the glucose concentration, 2.27 g / L of yeast extract and 1.09 g / L of corn steep liquor powder were added to the enzymatic hydrolysate as nitrogen sources for the fermentation medium, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0155] (c) The above-prepared mixture was sterilized at 115 °C for 15 min.

[0156] (d) The Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm was inoculated into the above-prepared mixture at a volume ratio of 10%, and fermented at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0157] The above lactic acid fermentation broth was detected by HPLC to obtain L-lactic acid produced by fermenting straw-type lignocellulose as a raw material. The concentration of L-lactic acid produced in the sugar solution was detected by HPLC to be 42.76 g / L, and the conversion rate of sugar fermentation to L-lactic acid was 81.42%.

[0158] Example 10

[0159] Take dry wheat straw raw materials and mechanically crush them into 3 - 5 cm segments as lignocellulose raw materials for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin-screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin-screw through the feeding hopper at the same time. During this period, 3 times the mass of the dry material of the pretreatment solution is pumped in, including 15 W / W ammonia water and 0.5 W / W sodium hydrogen phosphate. By adjusting the frequency of the twin-screw motor, the wheat straw is kept at 90 °C in the screw for a reaction for 3 h. The material after the heat preservation reaction is transported into a single-screw extruder for solid-liquid separation, and thus holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin are obtained.

[0160] After calculation, the retention rate of the solid cellulose obtained after pretreatment is 85.66%, the retention rate of hemicellulose is 68.62%, and the yield of lignin in the black liquor is 53.07%.

[0161] Preparation of straw lactic acid fermentation:

[0162] (a) Mix the holocellulose obtained above with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g holocellulose and 50 IU / g holocellulose respectively, and carry out enzymatic hydrolysis and saccharification at 50 °C for 48 h. Then, perform solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysate sugar solution.

[0163] (b) The conversion rate of cellulose in the above enzymatic hydrolysate sugar solution detected by HPLC is 80.39%, and the concentration of glucose is 53.66 g / L. Based on the glucose concentration, add 2.24 g / L of yeast extract and 1.07 g / L of corn steep liquor dry powder as the nitrogen source of the fermentation medium to the enzymatic hydrolysate sugar solution, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0164] (c) Sterilize the above compounded mixture at 115 °C for 15 min.

[0165] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm into the above compounded mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0166] Perform HPLC detection on the above lactic acid fermentation broth to obtain L-lactic acid produced by fermenting straw-type lignocellulose as the raw material. The concentration of L-lactic acid produced in the sugar solution detected by HPLC is 38.53 g / L, and the conversion rate of sugar fermentation to L-lactic acid is 74.58%.

[0167] Example 11

[0168] Take the dried wheat straw raw material and mechanically crush it into 3 - 5 cm segments as the lignocellulose raw material for pretreatment. The impregnating solution used is tap water, and it is heated to 90 °C. Start the twin-screw device, and continuously transport the heated impregnating solution and wheat straw segments to the twin-screw through the feeding hopper at the same time. During this period, pump in 7 times the mass of the dry material of the pretreatment solution, which contains 25% W / W ammonia water and 0.5 W / W sodium hydrogen phosphate. Adjust the frequency of the twin-screw motor to keep the wheat straw at 90 °C in the screw for 1 h for reaction. Transport the material after the heat preservation reaction to a single-screw extrusion machine for solid-liquid separation, and then obtain holocellulose rich in cellulose and hemicellulose components and alkaline black liquor containing hydrolyzed lignin.

[0169] After calculation, the retention rate of the solid cellulose obtained after pretreatment is 87.93%, the retention rate of hemicellulose is 67.44%, and the yield of lignin in the black liquor is 56.31%.

[0170] Preparation of straw lactic acid fermentation:

[0171] (a) Mix the above-obtained holocellulose with 5 mM sodium citrate buffer at a solid-liquid ratio of 1:10 g / L, add Trichoderma reesei cellulase and xylanase at 20 FPU / g of holocellulose and 50 IU / g of holocellulose respectively, and carry out enzymatic hydrolysis and saccharification at 50 °C for 48 h, and then perform solid-liquid separation at 10000 rpm for 5 min. The obtained supernatant is the enzymatic hydrolysis sugar solution.

[0172] (b) The conversion rate of cellulose in the above enzymatic hydrolysis sugar solution was detected by HPLC to be 89.83%, and the concentration of glucose was 55.35 g / L. Based on the glucose concentration, 2.31 g / L of yeast extract and 1.11 g / L of corn steep liquor dry powder were added to the enzymatic hydrolysis sugar solution as nitrogen sources for the fermentation medium, as well as 3 g / L of ammonium sulfate, 0.22 g / L of potassium dihydrogen phosphate, 0.4 g / L of magnesium sulfate heptahydrate, 0.03 g / L of manganese sulfate monohydrate, and 0.03 g / L of ferrous sulfate heptahydrate as inorganic salts.

[0173] (c) Sterilize the above compounded mixture at 115 °C for 15 min.

[0174] (d) Inoculate the Bacillus coagulans ACCC 10229 bacterial solution with an absorbance of 1.0 at OD600nm into the above compounded mixture at a volume ratio of 10%, and ferment at 50 °C and 150 rpm for 72 h to obtain a fermentation broth containing L-lactic acid.

[0175] The above lactic acid fermentation broth was detected by HPLC to obtain L-lactic acid fermented from straw-type lignocellulose as raw material. The concentration of L-lactic acid produced in the sugar solution detected by HPLC was 46.47 g / L, and the conversion rate of sugar fermentation to L-lactic acid was 87.10%.

[0176] Example 12

[0177] Same as Example 1, only change sodium hydrogen phosphate to other different inorganic ammonium salts, and the corresponding results are shown in Table 1.

[0178] Table 1

[0179]

[0180] Regarding this Example 1

[0181] Same as Example 1, except that the pretreatment solution is replaced with 25% W / W ammonia water alone.

[0182] After calculation, the retention rate of solid cellulose obtained after pretreatment was 58.89%, the retention rate of hemicellulose was 43.28%, and the yield of lignin in the black liquor was 38.69%.

[0183] The enzymatically hydrolyzed and saccharified lactic acid fermentation broth was detected by HPLC. The cellulose conversion rate was 72.41%, the glucose concentration was 45.03 g / L, and after fermentation, L-lactic acid was obtained with a concentration of 29.61 g / L. The conversion rate of sugar fermentation to L-lactic acid was 65%.

[0184] Comparative Example 2

[0185] Same as Example 1, except that the pretreatment solution was replaced with 1.0 W / W sodium hydrogen phosphate alone.

[0186] After calculation, the retention rate of solid cellulose obtained after pretreatment was 58.66%, the retention rate of hemicellulose was 45.59%, and the yield of lignin in the black liquor was 34.27%.

[0187] The enzymatically hydrolyzed and saccharified lactic acid fermentation broth was detected by HPLC. The cellulose conversion rate was 75.37%, the glucose concentration was 47.25 g / L, and after fermentation, L-lactic acid was obtained with a concentration of 25.82 g / L. The conversion rate of sugar fermentation to L-lactic acid was 56%.

[0188] Comparative Example 3

[0189] Same as Example 1, except that the pretreatment method was a high-temperature and high-pressure reaction kettle under the same conditions.

[0190] After calculation, the retention rate of solid cellulose obtained after pretreatment was 55.13%, the retention rate of hemicellulose was 48.66%, and the yield of lignin in the black liquor was 25.33%.

[0191] The enzymatically hydrolyzed and saccharified lactic acid fermentation broth was detected by HPLC. The cellulose conversion rate was 75.60%, the glucose concentration was 49.82 g / L, and after fermentation, L-lactic acid was obtained with a concentration of 36.92 g / L. The conversion rate of sugar fermentation to L-lactic acid was 60.05%.

[0192] Comparative Example 4

[0193] Same as Example 1, except that the pretreatment solution was an organic base 30% w / w methanolamine solution.

[0194] After calculation, the retention rate of solid cellulose obtained after pretreatment was 57.33%, the retention rate of hemicellulose was 45.37%, and the yield of lignin in the black liquor was 35.26%.

[0195] The enzymatically hydrolyzed and saccharified lactic acid fermentation broth was detected by HPLC. The cellulose conversion rate was 76.88%, the glucose concentration was 48.57 g / L, and after fermentation, L-lactic acid was obtained with a concentration of 46.74 g / L. The conversion rate of sugar fermentation to L-lactic acid was 75.86%.

[0196] Through comparative analysis, it can be seen that the pretreatment liquid and pretreatment method of the present invention can better promote the decomposition of straw, obtain a complete lignin structure with high purity and a holocellulose component with a high retention rate. Under the same pretreatment conditions, the retention rates of cellulose and hemicellulose are increased by 25% and 15% respectively. At the same time, the obtained holocellulose, as a biobased material, can be efficiently converted into L-lactic acid through enzymatic saccharification and can be further applied to multiple fields such as food, pharmacy, and chemical industry.

[0197] The present invention prepares a pretreatment liquid by compounding sodium hydrogen phosphate and ammonia water solution, and cooperates with a continuous pretreatment device, a twin-screw extruder, to efficiently separate holocellulose and lignin under mild conditions. While ensuring the cellulose retention rate, a lignin component with a complete structure retention is obtained. The retention rates of cellulose and hemicellulose are both maintained above 85% and 55%, and the yield of lignin is maintained above 65%. And no external heating is required during the pretreatment process, and the obtained fibers are not fragmented and have an obvious fibril structure. This pretreatment system has the feasibility of industrial scale-up. The holocellulose obtained after pretreatment is enzymatically saccharified and then efficiently converted into L-lactic acid through fermentation.

[0198] Lignocelluloses such as corn straw, rice straw, sorghum straw, sugarcane bagasse, corn cob, switchgrass, reed, and rapeseed are similar to wheat straw and can be treated and recycled using the same or similar methods.

[0199] The present invention provides an idea and method for a process of preparing lactic acid by mechanically enhanced ammonia method pretreatment of straw biomass. There are many specific methods and ways to implement this technical solution. The above description is only the preferred embodiment of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and refinements can be made, and these improvements and refinements should also be regarded as the protection scope of the present invention. Each component not clearly defined in this embodiment can be realized by existing technologies.

Claims

1. A process for preparing lactic acid by mechanically enhanced ammonia process pretreatment of straw biomass, characterized in that, The enzymatic hydrolysate of solid fibers obtained from straw-like biomass by mechanical enhanced ammonia pretreatment process is used as the carbon source of the fermentation medium in the process of lactic acid fermentation; The preparation of the carbon source in the fermentation medium includes the following steps: S1: Treat straw-like lignocellulosic biomass with a pretreatment solution containing ammonia water and inorganic ammonium salt in a twin-screw device; S2: Carry out a heat preservation reaction on the material obtained in step S1, and after solid-liquid separation, obtain solid fibers containing holocellulose; S3: In a buffer solution, enzymatically hydrolyze the solid fibers obtained in step S2 with Trichoderma reesei cellulase and xylanase, and after solid-liquid separation, the obtained supernatant is used as the carbon source in the fermentation medium for lactic acid fermentation; In step S1, the pretreatment solution is an aqueous solution containing 20% - 30% w / w ammonia water and 0.5% - 1.5% w / w inorganic ammonium salt; the inorganic ammonium salt is sodium hydrogen phosphate ammonium; the dosage of the pretreatment solution is 2 - 10 times the mass of the straw-like lignocellulosic biomass; In step S2, the temperature of the heat preservation reaction is 80 - 140 °C; the time of the heat preservation reaction is 0.5 - 5 h; In step S3, the addition amount of Trichoderma reesei cellulase is 10 - 30 FPU / g solid fiber; the addition amount of xylanase is 40 - 60 IU / g solid fiber.

2. The process according to claim 1, characterized in that, In step S3, the dosage ratio of the solid fiber to the buffer solution is 1:(5 - 15) g / L.

3. The process according to claim 1, wherein In step S3, the temperature of the enzymatic hydrolysis is 40 - 60 °C.

4. The process according to claim 1, wherein The fermentation medium also includes a nitrogen source and inorganic salts; the nitrogen source is yeast extract and / or corn steep liquor dry powder; the inorganic salts are ammonium sulfate, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, manganese sulfate monohydrate, and ferrous sulfate heptahydrate.

5. The process according to claim 4, wherein The concentration of inorganic salts in the fermentation medium is as follows: ammonium sulfate 2 - 4 g / L, potassium dihydrogen phosphate 0.12 - 0.32 g / L, magnesium sulfate heptahydrate 0.3 - 0.5 g / L, manganese sulfate monohydrate 0.01 - 0.05 g / L, ferrous sulfate heptahydrate 0.01 - 0.05 g / L.

6. The process according to claim 1, characterized in that, Inoculate the Bacillus coagulans bacterial solution into the fermentation medium containing the carbon source for fermentation, and a fermentation broth containing lactic acid is obtained.

7. The process according to claim 6, characterized in that, The Bacillus coagulans bacterial solution is inoculated into the fermentation medium at a volume ratio of 5% - 20%.

8. The process according to claim 6, characterized in that, The temperature of the fermentation is 40 - 60 °C; the rotation speed of the fermentation is 100 - 200 rpm; the time of the fermentation is 62 - 82 h.

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