TWI526540B - Method of producing lactic acid from high-lignin fibrous material source - Google Patents

Description

Method for producing lactic acid by using high lignin fiber source

The present invention relates to a process for the lactic acid process for high lignin fiber sources, and more particularly to a method for lactic acid fermentation using a high lignocellulosic material source.

At present, the production of lactic acid fermentation mainly uses grain (such as corn) as its main source, and the bulk production raw material is corn starch, which makes the current lactic acid process unable to effectively reduce the cost of raw materials.

In view of this, agricultural and forestry waste and the lignocellulose rich in it are a low-cost source that can be effectively developed. The use of lignocellulosic raw materials to manufacture chemicals generally includes procedures such as raw material pretreatment, enzyme hydrolysis and fermentation, and product purification. In these procedures, there are still many technical difficulties in the fermentation of lignocellulose as a fermentation substrate because of the problems in process development. However, there are few researches and applications in the production of lactic acid by fermentation, and in the international literature. The disclosed production method has the problems of adding a nutrient source, high production cost and low yield. Therefore, the present invention specifically develops a process for high lignin cellulose source to improve the production process of lignocellulose to produce lactic acid.

And the well-known commercial production of lactic acid mostly uses food crops as raw materials for production, so that the price of lactic acid is linked with the price of food, and there is doubts about food competition with the people; and the current development of wood fiber lactic acid technology is scarce, the published literature Most of them are based on laboratory grades. The production of lactic acid is mostly between 30 and 60 g/L, which is still some distance from the commercial threshold of lactic acid production.

In order to solve the above-mentioned shortcomings of the prior art, the object of the present invention is to provide a high-efficiency production, high concentration and high optical purity of the fiber lactic acid process, and at the same time, specifically for the high lignin fiber source, specialized hydrolysis and fermentation, obtained by the method The level of lactic acid production can reach commercial indicators and has the value of commercial development.

In order to achieve the above and other objects, the present invention provides a method for producing lactic acid by using a high lignin fiber source, which comprises: a strain cultivation step: the selected strain is cultured in a suitable medium for the culture step, and The strain is lactic acid bacteria; a dilute acid pretreatment step: pretreating the high lignin fiber source with dilute acid; an enzyme hydrolysis step: adding the source obtained by the above dilute acid pretreatment step to the polymer polymerization a substance or a surfactant, and a hydrolyzed enzyme for enzymatic hydrolysis; a lactate fermentation step: adding the product obtained by the enzyme hydrolysis step to the bacterial body and the nitrogen source obtained by the cultivation step of the inoculum to carry out lactic acid fermentation.

The method as described above, wherein the dilute acid used in the dilute acid pretreatment step is 0.5-3.0% dilute sulfuric acid, and the dilute acid pretreatment step is carried out at 160-200 °C.

The method as described above, wherein the polymer or surfactant used in the enzyme hydrolysis step is added in an amount of 0.1 to 2% of a polymer or a surfactant according to the lignin content and the operation volume. .

The method as described above, wherein the high molecular polymer or surfactant is polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60), dodecane Trimethylammonium chloride (DTC), polyoxyethylene sorbitan monopalmitate (Tween 40), polyoxyethylene stearate-30 (SE-30), polyoxyethylene stearate-40 ( SE-40), polyoxyethylene dehydrated sorbus Alcohol monolaurate (Tween 20), polyethylene glycol (PEG 6000), polyethylene glycol (PEG 4000) or polyethylene glycol (PEG 1500).

The method as described above, wherein the high lignin fiber material source is rice straw, bagasse, wood material or pennisetum.

The lactic acid production of the high lignin fiber source is carried out by the method for producing lactic acid by using the high lignin fiber source provided by the invention, wherein the high lignin fiber source can be rice straw, bagasse, wood material or pennisetum, and The lactic acid yield obtained by the invention method is more than 80 g/L, the lactic acid efficiency of glucose conversion is more than 95%, the optical purity of lactic acid is over 97%, and can be applied to a scale above a pilot plant, and the high wood provided by the present invention is utilized. The commercial production of lactic acid from the source of quality fiber can achieve the effect of energy saving and carbon reduction, and can reduce the dispute with the people.

S0‧‧‧ breeding step

S1‧‧‧ dilute acid pretreatment steps

S2‧‧‧ Enzyme hydrolysis step

S3‧‧‧Lactic acid fermentation step

Figure 1 is a flow diagram of a method of producing lactic acid using a high lignin fiber source.

Figure 2 is a graph showing the results of lactic acid production after hydrolysis and fermentation with or without additives.

Figure 3 is a graph showing the results of lactic acid production after fermentation of different lactic acid strains from high lignin fiber sources.

Figure 4 is a graph showing the results of lactic acid production after fermentation of lactic acid bacteria with different high lignin sources.

In order to fully understand the objects, features and advantages of the present invention, the present invention will be described in detail with reference to the accompanying drawings. A flow chart of a method for producing lactic acid using a high lignin fiber source. As shown in Fig. 1, the present invention provides a method for producing lactic acid by using a high lignin fiber source, first performing a strain cultivation step S0, and then using a high lignin fiber source as a dilute acid for dilute acid. In the pretreatment step S1, an additive and an enzyme are further added to carry out the enzyme hydrolysis step S2, and then a nitrogen source and a lactic acid strain (the strain obtained from the strain culture step S0) are added to carry out a lactic acid fermentation step S3 to produce lactic acid by fermentation. The lactic acid production of the high lignin fiber source is carried out by the method for producing lactic acid by using the high lignin fiber source provided by the invention, the lactic acid production is above 80 g/L, the lactic acid efficiency of glucose conversion is over 95%, and the optical purity of lactic acid is over 97%. And can be applied to the scale above the pilot plant.

First, in the inoculum culture step S0, the lactic acid bacteria culture medium (see Table 1 below) prepared according to the present invention is adjusted to a pH of 5.5 to 7.0, and autoclaved at 121 ° C for 15 to 20 minutes as a medium. Then, inoculate lactic acid strains, such as Enterococcus INER A, Lactobacillu s INER B, Lactobacillu s INER C or commonly used lactic acid fermentation strains, and adjust the culture pH with lye (NaOH, NH ₄ OH), culture 4~12 The inoculum culture step is completed in an hour.

Furthermore, in the dilute acid pretreatment step S1, the high lignin fiber source (lignin content 20-60%, as shown in Table 2 below) is used to cut the high lignin fiber source to 0.5-2 cm by using a chopper, and is added. 0.5-3% dilute sulfuric acid, heated to 160-200 ° C, and maintained for 0.5-3 minutes, after heating, the pressure difference is formed by the pressure in the tank to structurally destroy the high lignin fiber source, thereby improving the hydrolysis efficiency of the enzyme.

Table 2. Composition of high lignin fiber source

Next, the enzyme hydrolysis step S2 of the high lignin fiber source is used, and the high lignin dilute acid pretreatment source is configured to have a solid-liquid ratio of 10 to 30%, and the polymer is added in an amount of 0.1 to 2% according to the lignin content and the operation volume. Or surfactant (see Table 3), adjust the pH to 4.8~5.2, and add fibrin to hydrolyze. The enzyme is Novozymes Cellic® CTec3, and its enzyme activity is 5~30FPU per gram of cellulose, and at temperature 50. After the temperature is °C, the pH is 5.2, and the hydrolysis is carried out for 48 to 72 hours, the enzyme hydrolysis step S2 is completed.

The product obtained by the above enzyme hydrolysis step is subjected to solid-liquid separation, and the obtained liquid is added to the cells obtained by the above-mentioned inoculating step, the inoculum amount is 0.25 to 1 g/L, and the liquid obtained after solid-liquid separation is added to 0.2 to 2%. Conventional nitrogen source, at 35~42 °C, pH 6~8, fermentation for 24~72 hours, milk The acid is fermented in step S3 to obtain the complete fermented lactic acid; the fermented sample obtained by fermenting the lactic acid is analyzed by high performance liquid chromatography.

The present invention uses a high lignin source (wood waste) via an inoculum culture step S0, a dilute acid pretreatment step S1, an enzyme hydrolysis step S2, and a lactate fermentation step S3. In one embodiment, the present invention uses Enterococcus INER A as an experimental culture fermentation strain, in this embodiment a 20% high lignocellulosic acid pretreatment source (treatment conditions: 1.2% dilute sulfuric acid, treatment temperature 200 ° C , reaction time 5 min, material of the source: wood waste), and 15FPU enzyme was added for hydrolysis, and then inoculated with Enterococcus INER A at a dose of 0.4 g / L. Each experimental group was added 0.5% in the enzyme hydrolysis step. High molecular weight polymer (PEG6000), 1% surfactant (Tween80) and no added control group. The experimental results are shown in Fig. 2. From the experimental results in Fig. 2, it was found that 0.5% polymer was added to the program. (PEG6000) and 1% surfactant (Tween80) had lactic acid yield of 88.3g/L and 85.51g/L, respectively, while in the unadded control group, the lactic acid yield was 67.11g/L, although the sugar of the three The conversion lactic acid efficiency was higher than 95%, but the lactic acid yield was significantly different. In the additive group, the lactic acid production was higher than that of the unadded group by 20~25%, and the lactic acid yield was about 20g/L.

In another embodiment, the source is pretreated with a 20% high lignocellulosic acid (treatment conditions: 1.2% dilute sulfuric acid, treatment temperature 200 ° C, reaction time 5 min, material source is wood waste), and 15 FPU is added. The enzyme and 1% surfactant (Tween80) were hydrolyzed, and the strains of Enterococcus INER A, Lactobacillu s INER B and Lactobacillu s INER C were inoculated at a dose of 0.4 g/L, respectively. The experimental results are shown in Fig. 3. From the experimental results in Figure 3, it can be found that these strains can produce sugar in the hydrolyzed liquid after hydrolysis, and the conversion rate of glucose to lactic acid is higher than 95%, and the lactic acid yield is higher than 80g/L; High performance liquid chromatography (HPLC) analysis, Enterococcus INER A. produced L-type lactic acid optical purity of 99%; Lactobacillu s INER B. produced L-type lactic acid optical purity of 99%; and Lactobacillu s INER C produced D The lactic acid optical purity is also up to 99%.

In still another embodiment, the 20% high wood fiber dilute acid pretreatment material source (treatment conditions are: 0.5 to 1.5% dilute sulfuric acid, treatment temperature 185 to 200 ° C, reaction time 1 to 5 min, high lignin fiber source respectively For rice straw, bagasse, wood chips (wood material) or pennisetum, and then 15FPU enzyme and 1% surfactant (Tween80) for hydrolysis, followed by Enterococcus INER A. inoculated with 0.4 g / L of bacteria The experimental results are shown in Fig. 4. From the experimental results in Fig. 4, it can be found that Enterococcus INER A can produce lactic acid in the hydrolyzed liquid after hydrolysis of different seed sources, and the conversion ratio of glucose-transformed lactic acid is higher than 95%. The lactic acid production can be higher than 70g/L depending on the source characteristics. The experimental results show that the method for producing lactic acid by using the high lignin fiber source can be applied to different sources of lactic acid. Fermentation, and the production of different sources of lactic acid by the method provided by the present invention can reach commercial standards.

Through the above experiments, it is proved that the method for producing lactic acid by using a high lignin fiber source provides the lactic acid production of a high lignin fiber source, wherein the high lignin fiber source can be rice straw, bagasse, wood chips (wood materials). Or pennisetum, can obtain high yield of lactic acid higher than 70g / L; and the lactic acid yield obtained by the method of the invention can also reach 80g / L or more, glucose conversion lactic acid efficiency of more than 95%, the obtained lactic acid optical purity Up to 97% or more, and can be applied to the scale of the pilot plant. Therefore, the commercial production of lactic acid by using the high lignin fiber source provided by the present invention can achieve the effect of energy saving and carbon reduction, and can reduce the number of people Controversy over food.

The invention has been described above in terms of the preferred embodiments, and it should be understood by those skilled in the art that the present invention is not intended to limit the scope of the invention. It should be noted that variations and permutations equivalent to those of the embodiments are intended to be included within the scope of the present invention. Therefore, the scope of protection of the present invention is defined by the scope of the patent application.

S0‧‧‧ breeding step

S1‧‧‧ dilute acid pretreatment steps

S2‧‧‧ Enzyme hydrolysis step

S3‧‧‧Lactic acid fermentation step

Claims (5)

The invention relates to a method for producing lactic acid by using a high lignin fiber source, which comprises: a strain cultivation step: the selected strain is cultured in a suitable culture medium, and the strain is a lactic acid bacteria; a dilute acid Pretreatment step: pretreating the high lignin fiber source with dilute acid; an enzyme hydrolysis step: adding the source obtained by the above dilute acid pretreatment step to the polymer or surfactant, and hydrolyzing the enzyme to carry out Enzymatic hydrolysis; a lactic acid fermentation step: adding the product obtained by the enzyme hydrolysis step to the bacterial cells and nitrogen sources obtained in the culture step of the bacteria to carry out lactic acid fermentation. The method of claim 1, wherein the dilute acid used in the dilute acid pretreatment step is 0.5-3.0% dilute sulfuric acid, and the dilute acid pretreatment step is carried out at 160-200 °C. The method of claim 1, wherein the polymer or surfactant used in the enzyme hydrolysis step is added in an amount of 0.1 to 2% based on the lignin content and the operation volume. Or surfactant. The method of claim 3, wherein the high molecular polymer or surfactant is polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60) ), dodecyltrimethylammonium chloride (DTC), polyoxyethylene sorbitan monopalmitate (Tween 40), polyoxyethylene stearate-30 (SE-30), polyoxyethylene stearin Acid ester-40 (SE-40), polyoxyethylene sorbitan monolaurate (Tween 20), polyethylene glycol (PEG 6000), polyethylene glycol (PEG 4000) or polyethylene glycol (PEG 1500). The method of claim 1, wherein the high lignin fiber material source is rice straw, bagasse, wood material or pennisetum.