CN117646040A - Method for converting xylose mother liquor into xylitol through biological fermentation - Google Patents

Method for converting xylose mother liquor into xylitol through biological fermentation Download PDF

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CN117646040A
CN117646040A CN202311681748.8A CN202311681748A CN117646040A CN 117646040 A CN117646040 A CN 117646040A CN 202311681748 A CN202311681748 A CN 202311681748A CN 117646040 A CN117646040 A CN 117646040A
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xylitol
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李欢欢
张春燕
苏玉庆
刁功科
张文魁
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Henan Yuxin Sugar Alcohol Co ltd
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Abstract

The invention belongs to the technical field of xylitol preparation and biological fermentation, and particularly relates to a method for converting xylose mother liquor into xylitol through biological fermentation. According to the method for converting xylose mother liquor into xylitol through biological fermentation, kluyveromyces marxianus is used as a fermentation strain, and nitrosoguanidine is used for carrying out mutagenesis on the strain to obtain a base mutant strain; then, a TC staining method is adopted for screening to obtain strains with vigorous respiratory enzyme activity, so that blindness of common screening is reduced; and finally, according to the way of converting xylose into xylitol, special nutrition additives are added, so that the conversion of xylose into xylitol can be further promoted. The method not only omits the xylose purification step, but also simplifies the xylitol separation step, can effectively reduce the production cost of the xylitol, and has high content and purity of the xylitol in the obtained product, thereby having good application prospects in the fields of reuse of xylose mother liquor and production of the xylitol.

Description

Method for converting xylose mother liquor into xylitol through biological fermentation
Technical Field
The invention belongs to the technical field of xylitol preparation and biological fermentation, and particularly relates to a method for converting xylose mother liquor into xylitol through biological fermentation.
Background
Xylitol is a polyhydric alcohol containing five carbon atoms and five hydroxyl groups, and has wide application in industries such as food, medicine, paper making, chemical industry and the like. The current industrial method for preparing xylitol mainly comprises the following steps: xylose is obtained by acid hydrolysis of xylan, and xylitol is prepared by a chemical catalytic hydrogenation method. In the xylose extraction process, a great amount of xylose mother liquor byproducts are generated. The xylose mother liquor is brown yellow, contains more impurities, has low purity and high viscosity, and contains glucose, galactose, arabinose and other carbon sources with certain concentration and a small amount of furfural, tetrahydrofuran and other substances besides xylose.
In order to avoid resource waste and realize the reutilization of xylose mother liquor, many researchers have studied on the utilization and the reextraction technology of xylose mother liquor. The most common existing purification process is to decompress and evaporate xylose mother liquor, and then filter out evaporated xylose crystals. However, this method has problems of high energy consumption, unstable quality, and high water content of the crystals obtained. In addition, a scholars sequentially propose a process method for separating and purifying the xylose mother liquor by using resin and a process method for separating active ingredients in the xylitol mother liquor by using chromatography. The separation method needs to carry out procedures such as concentration and separation on xylose in mother liquor, and then carries out a series of operations such as filtration and exchange again to obtain xylitol, and has the problems of high equipment requirement and high operation difficulty. Later, some researchers propose a method for fermenting xylose mother liquor by using strains, which adopts the strains to directly ferment xylose in the xylose mother liquor into xylitol, but the problems of longer fermentation period and lower conversion rate still exist, so that the further application of the method is limited.
Disclosure of Invention
In order to solve the problems in the prior art, the invention aims to provide a method for converting xylose mother liquor into xylitol through biological fermentation, which comprehensively improves the conditions and the formula from strain breeding to fermentation, and can greatly improve the yield and the purity of the xylitol
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
a method for converting xylose mother liquor to xylitol by biological fermentation, comprising the steps of:
(1) Carrying out chemical mutagenesis treatment on Kluyveromyces marxianus by adopting nitrosoguanidine to obtain a mutagenized strain;
(2) Dyeing the strain subjected to mutagenesis in the step (1) by adopting 2,3, 5-triphenyltetrazolium chloride, and screening according to a dyeing result to obtain a colony with vigorous respiratory enzyme activity;
(3) Respectively inoculating the colonies with vigorous respiratory enzyme activity obtained in the step (2) into a screening culture medium containing xylose mother liquor for culture, measuring the xylitol content and purity in the obtained fermentation liquor after fermentation, and screening according to the content and purity measurement results to obtain the optimal strain;
(4) And (3) inoculating the optimal strain obtained by screening in the step (3) into a fermentation culture medium containing xylose mother liquor for fermentation culture, and thus, converting the xylose mother liquor into xylitol.
As a further preferable scheme, in the step (1), the chemical mutagenesis treatment is to add 0.8-1.2 mg/mL nitrosoguanidine solution into the yeast suspension of Kluyveromyces marxianus and treat the yeast suspension in a water bath at 35-40 ℃ for 20-40 min. Nitrosoguanidine mutagenesis mainly causes GC- & gtAT conversion in the replication region of the DNA strand, and causes high-frequency mutation in microorganisms.
As a further preferable mode, in the step (1), the kluyveromyces marxianus is subjected to an activation treatment in advance; the activation treatment is to inoculate Kluyveromyces marxianus into an activation culture medium, and perform activation culture for 10-15 h at the temperature of 28-32 ℃ at 200-300 rpm; the composition of the activation medium is as follows: glucose 9-12 g, yeast extract 0.3-0.7 g, peptone 0.3-0.5 g, and water to constant volume to 50mL.
As a further preferable scheme, in the step (2), the dyeing treatment is performed after mixing the 2,3, 5-triphenyltetrazolium chloride aqueous solution with the bacterial suspension of the mutagenized strain; the dyeing treatment time is 1-3 d; the mass concentration of the 2,3, 5-triphenyl tetrazolium chloride aqueous solution is 0.8% -1.2%. In the invention, 2,3, 5-Triphenyl Tetrazolium Chloride (TTC) developer is utilized for dyeing reaction, which is called TC dyeing method, is generally used for identifying the activity of plant seeds, and can also generate color reaction on the metabolic products of yeast, and the activity of respiratory enzymes in the yeast is judged according to the color development degree. The invention uses nitrosoguanidine to carry out chemical mutagenesis treatment on the Kluyveromyces marxianus strain, and then screens the Kluyveromyces marxianus strain by TC staining method, so that the xylitol production capacity of the yeast can be distinguished, and blind screening is avoided.
As a further preferred scheme, in the step (3), the screening culture medium consists of the following components in parts by mass: 16 to 20 percent of xylose mother liquor, 0.3 to 0.8 percent of corn steep liquor dry powder, 0.03 to 0.08 percent of monopotassium phosphate and water as solvent. When the screening culture medium is prepared, 16-20 g of xylose mother liquor, 0.3-0.8 g of corn steep liquor dry powder and 0.03-0.08 g of monopotassium phosphate are taken, and water is added to fix the volume to 100mL.
As a further preferable embodiment, in step (3), the culturing conditions are: culturing at 200-300 rpm and 28-33 deg.c for 70-80 hr.
As a further preferred embodiment, in step (4), the fermentation medium consists of a basal medium and a nutritional supplement; the basic culture medium consists of the following components in parts by mass: 23 to 28 percent of xylose mother liquor, 0.3 to 0.8 percent of corn steep liquor dry powder, 0.4 to 0.6 percent of urea, 0.02 to 0.05 percent of monopotassium phosphate and water as solvent; the composition of the nutritional supplement, based on the final concentration in the basal medium, is: amino acid mixed solution 4-6 mL/L, VB 2 8～12mg/L、VB 3 4～6mg/L。
Based on the consideration of further improving the yield and purity of xylitol, as a more preferable scheme, the basal medium consists of the following components in parts by mass: 25% of xylose mother liquor, 0.5% of corn steep liquor dry powder, 0.5% of urea and 0.025% of monopotassium phosphate, and the solvent is waterThe method comprises the steps of carrying out a first treatment on the surface of the The composition of the nutritional supplement, based on the final concentration in the basal medium, is: amino acid mixture 5mL/L, VB 2 10mg/L、VB 3 5mg/L。
As a further preferable mode, in the step (4), the conditions of the fermentation culture are: fermenting and culturing at 200-300 rpm, 28-33 deg.c and pH of 5.5-6.5 for 70-80 hr.
As a further preferable mode, in the step (4), the inoculation amount of the optimal strain is 13-17% V/V during the fermentation culture.
The technical scheme of the invention has the beneficial effects that:
according to the method for converting xylose mother liquor into xylitol through biological fermentation, kluyveromyces marxianus is used as a fermentation strain, and nitrosoguanidine is used for carrying out mutagenesis on the strain to obtain a base mutant strain; then, a TC staining method is adopted for screening to obtain strains with vigorous respiratory enzyme activity, so that blindness of common screening is reduced; finally, according to the way of converting xylose into xylitol, the amino acid mixture and vitamins are added, so that the conversion of xylose into xylitol is further promoted.
According to the method disclosed by the invention, the xylitol is produced by fermenting the rough xylose mother liquor serving as a substrate by utilizing the reductase in the microorganism, so that not only is the xylose purification step omitted, but also the xylitol separation step is simplified, and the production cost of the xylitol can be effectively reduced. In addition, the xylitol is produced by adopting a biotechnology, xylose is reduced into xylitol under the catalysis of xylose reductase in microbial cells, hydrogen is derived from hydrogen atoms in reduced coenzyme in the cells and hydrogen ions in water, independent hydrogen production is not needed, and no additional hydrogen source is needed, so that the production process is simple, safe and energy-saving, and the method is a very promising production method and has good application prospects in the fields of reuse of xylose mother liquor and production of xylitol.
Detailed Description
The technical scheme of the invention will be clearly and completely described below with reference to specific examples and test examples. It should be understood by those skilled in the art that the examples are only for illustrating the technical scheme of the present invention and should not be construed as limiting the scope of the present invention. All other embodiments, such as modified, modified or simply substituted embodiments, which would be apparent to one of ordinary skill in the art without undue effort based on the examples described below, are intended to be within the scope of the present invention.
In the following examples, kluyveromyces marxianus (Kluyveromyces marxianus) was used from Shanghai pacifying industries, inc.
In the following examples, the amino acid mixtures are conventional commercial products having the following composition: proline 0.6%, threonine 0.6%, methionine 0.7%, isoleucine 0.7%, alanine 0.4%, valine 0.6%, leucine 0.7%, cysteine 1.2%, phenylalanine 0.8%, p-aminobenzoic acid 0.2%, aspartic acid 0.7%, glycine 0.4%, histidine 0.8%, lysine 0.7%, serine 0.5%, tryptophan 1%, tyrosine 0.9%, glutamic acid 0.9%, arginine 0.2%.
Example 1
This example provides a method for converting xylose mother liquor to xylitol by biological fermentation, comprising the steps of:
(1) Carrying out chemical mutagenesis treatment on Kluyveromyces marxianus by adopting nitrosoguanidine to obtain a mutagenized strain;
the specific operation process of the steps is as follows: (1) 50mL of an activation medium (glucose 10g, yeast extract 0.5g, peptone 0.4g, and water was added to a volume of 50 mL) was prepared, and the mixture was placed in a 500mL shaking flask, and sterilized at 115℃for 25 minutes. (2) After the activated medium is cooled, a proper amount of thalli (Kluyveromyces marxianus) is taken and inoculated into a shake flask for culturing for 12 hours at 30 ℃ and 220 r. (3) After the culture is finished, respectively pouring the bacterial liquid into a centrifuge tube after the sterilization treatment, centrifuging for 10min at 12000r, and pouring out the supernatant; then 1mL of sterile water is added into the centrifuge tube with the bacterial precipitation, the mixture is fully mixed and washed, the mixture is centrifuged for 10min at 12000r, the supernatant is poured off, and the washing is repeated twice. (4) Adding 1mL of phosphate buffer solution (pH=6) into the washed thalli, and uniformly mixing to prepare bacterial suspension; then 100uL of nitrosoguanidine solution at 1mg/ml was added to the bacterial suspension, mixed well, and subjected to chemical mutagenesis treatment in a water bath at 37℃for 30min (1 time every 5 minutes).
(2) Performing TC staining treatment on the strain subjected to mutagenesis in the step (1) by adopting 2,3, 5-triphenyltetrazolium chloride, and screening according to a staining result to obtain a colony with vigorous respiratory enzyme activity;
the specific operation process of the steps is as follows: (1) preparing a 2,3, 5-Triphenyl Tetrazolium Chloride (TTC) aqueous solution with the mass concentration of 1%, filtering and sterilizing. 100mL of solid culture medium (15 g of glucose, 0.5g of yeast extract powder and 18g of agar) was prepared, water was added to a volume of 100mL, and the mixture was sterilized at 115℃for 25min. (2) 100uL of TTC aqueous solution is added when the solid medium is cooled to 50 ℃, and after gentle mixing, 4 plates are poured. (3) Gradient dilution of nitrosoguanidine chemically mutagenized bacterial suspension to 10 0 、10 -1 、10 -2 、10 -3 And (4) respectively sucking 200uL of the plates coated with the treatment, culturing for 2 days at the temperature of 30 ℃ in an inverted mode, and selecting a colony with redder color according to the dyeing result to serve as a colony with vigorous respiratory enzyme activity.
(3) Respectively inoculating the bacterial colonies with vigorous respiratory enzyme activity obtained in the step (2) into a screening culture medium containing xylose mother liquor for fermentation culture, measuring the xylitol content and purity in the obtained fermentation liquor after fermentation, and screening according to the content and purity measurement results to obtain the optimal bacterial strain;
the specific operation process of the steps is as follows: preparing solid culture medium (glucose 15g, yeast extract powder 0.5g, agar 18g, adding water and fixing solute 100 mL), subpackaging into test tubes, sterilizing at 115deg.C for 25min, and obliquely standing until the culture medium solidifies. And (3) selecting 20 single colonies which are redder after dyeing in the step (2), respectively named as M-1 to M-20, and culturing the single colonies on a test tube inclined plane in a streak mode for 2 days at the temperature of 30 ℃. Screening by shaking table after culturing, subpackaging screening culture medium (18% xylose mother liquor, 0.5% corn steep liquor dry powder, 0.05% potassium dihydrogen phosphate, 100mL of water-added fixed solute) into triangular flasks with edges, sterilizing each flask at 115 ℃ for 25min, inoculating the M-1-M-20 strain (the inoculation amount is one loop of an inoculating loop) respectively, culturing for 72h at 220r and 30 ℃, and detecting the purity and content of xylitol.
Wherein, the purity test of xylitol adopts HPLC method, HPLC instrument is provided with RID detector, mobile phase is pure water, flow rate is 0.6mL/min, and external standard method is used for quantification. And measuring the content of xylitol, namely measuring the mass content (unit g/100 mL) of dry matters in the solution obtained by fermentation by adopting a refractometer, and multiplying the mass content and the purity of the dry matters to obtain the content of xylitol. For example: 28g/100mL of dry matter of the fermented solution, and the purity of the xylitol is 60%, and the xylitol content in the solution is 28 x 60% = 16.8g/100mL.
The original strain of Kluyveromyces marxianus obtained by purchase was used as an experimental strain (control group), and the measurement results of the content and purity of xylitol obtained after fermentation of each strain are shown in Table 1.
TABLE 1 determination of xylitol content and purity by fermentation of different strains
Sequence number Experimental strains M1 M2 M3 M4 M5 M6
Content g/100mL 6.99 6.47 6.4 6.75 7.35 4.71 5.48
Xylitol purity% 58.29 51.77 51.21 55.31 61.78 36.2 42.8
Sequence number M7 M8 M9 M10 M11 M12 M13
Content g/100mL 7.72 6.85 6.68 5.93 4.66 7.22 6.83
Xylitol purity% 64.29 57.08 55.61 49.39 33.26 60.13 56.89
Sequence number M14 M15 M16 M17 M18 M19 M20
Content g/100mL 6.54 7.20 6.68 5.64 5.80 6.00 6.53
Xylitol purity% 54.47 60.04 55.64 47.04 48.37 50.02 57.29
As can be seen from Table 1, the xylitol has a purity of 58.29% and a content of 6.99g/100mL after fermentation for 72 hours in the experimental strain of the control group; the mutant strain M7 is fermented for 72 hours, the purity of xylitol is 64.29%, and the content of xylitol is 7.72g/100mL, which is obviously improved compared with a control group, so that the mutant strain M7 is used as an optimal strain for subsequent experiments.
(4) And (3) inoculating the optimal strain obtained by screening in the step (3) into a fermentation culture medium containing xylose mother liquor for fermentation culture, and thus, converting the xylose mother liquor into xylitol.
The selection and optimization of the culture of the fermentation medium in step (4) and the determination of the fermentation process parameters are specifically described below.
The fermentation medium is a carrier required by microorganism growth and propagation and biosynthesis of various metabolites, and is mainly a mixture of various nutrients prepared according to a certain proportion. The culture medium suitable for large-scale fermentation can meet the requirements of the most economical synthesis of the product, fewer byproducts, proper raw materials, low cost, stable performance, convenient purchase and transportation and suitability for large-scale storage.
1. Optimization of nitrogen source and nutrient additives selection
The nitrogen source is a nutrient source capable of providing nitrogen elements required by the growth and propagation of microorganisms. As xylitol belongs to a product without N, in theory, the N source only needs to meet the nutrition components required by the growth of thalli, so the invention takes the corn steep liquor dry powder with favorable price as the nitrogen source.
The pathway for the conversion of xylose to xylitol is the isomerisation of xylose to xylulose by the action of Xylose Isomerase (XI) followed by the production of xylitol by the action of xylitol reductase. Vitamin B2 is a prosthetic group of a flavinase that plays a role in hydrogen transfer during the biological redox process. Vitamin B3 is also known as niacin, vitamin B3 is together with tryptophan, which are (NAD + ) Coenzyme I, (NADP) + ) Coenzyme II and its respective reduced form (NAD (P) H) dehydrogenases are also useful for delivering hydrogen ions. The invention aims to strengthen the metabolic pathway of xylitol by adding proper amount of vitamins and amino acids to strengthen the nutritional factors.
The specific tests are shown in Table 2. Wherein, the fermentation medium comprises basic medium and nutrition additive, and basic medium formula is: 20% of xylose mother liquor, 0.5% of corn steep liquor dry powder, 0.5% of urea and 0.025% of monopotassium phosphate, and the solvent is water. Preparing basic culture medium, taking corn steep liquor dry powder 0.5g and urea0.5g, 0.025g of monopotassium phosphate, and 20% xylose mother liquor (dry matter 20g/100mL solution) were added to a volume of 100mL. The primarily selected nutritional additives mainly comprise amino acid mixed solution, choline and VB 2 、VB 3 The specific composition of the fermentation medium is shown in Table 2. When the strain is fermented, the mutagenic strain M7 is inoculated into a fermentation culture medium for fermentation culture, and the M7 seed liquid OD is used for inoculation 600 At 10-15, post-inoculation OD 600 Fermenting and culturing at 220r and 30deg.C under 0.7-1.0. The effect of the above-described nutritional supplements on xylitol content and purity is shown in Table 2. Wherein "/" means that the component is added and "/" means that the component is absent.
TABLE 2 optimization of fermentation Medium formulation results
As is clear from Table 2, 5ml/L of the amino acid mixture and 10mg/L of VB were added to the formulation of the fermentation medium 2 VB at 5mg/L 3 (corresponding to experiment group 14) it was possible to effectively shorten the fermentation period and increase the xylitol content, so that the optimal nutritional supplement composition was determined as follows: amino acid mixture 5mL/L, VB 2 10mg/L、VB 3 5mg/L. Furthermore, it was found by experiment that the addition of choline resulted in a significantly longer fermentation period, and therefore no further choline was added to the nutritional supplement.
2. Initial xylose mother liquor concentration optimization
And respectively preparing xylose mother liquor with concentration of 10%, 15%, 20%, 25% and 30%, performing comparative test, performing shake flask fermentation under the same fermentation conditions, and examining the influence of the xylose mother liquor concentration on xylitol yield. The results are shown in Table 3.
TABLE 3 influence of mother liquor concentration on fermentation effect
The results in Table 3 show that at 25% xylose mother liquor concentration, xylitol content and conversion were higher and overall cost was lowest.
3. Fermentation process optimization
3.1 control of fermentation temperature
The experiment examined the effect of fermentation temperature on xylitol production: the same fermentation medium is adopted, other fermentation conditions are the same, shake flask fermentation is respectively carried out at 28 ℃,30 ℃, 33 ℃ and 35 ℃, the influence of the fermentation temperature on the xylitol yield is examined, and the result shows that the fermentation temperature has the best effect at 30 ℃.
3.2 control of fermentation broth pH
The experiment examined the effect of the initial pH of the fermentation medium on xylitol production: the same fermentation medium is adopted, other fermentation conditions are kept the same, initial pH values are respectively adjusted to 5.0, 5.5, 6.0, 6.5 and 7.0, shake flask fermentation is carried out, the influence of different initial pH values on xylitol yield is examined, and the result shows that the fermentation pH values are basically consistent in effect and superior to 5.0 and 7.0 at 5.5, so that the preferable fermentation pH value range is 5.5-6.5.
3.3 Effect of inoculum size on fermentation
The effect of inoculum size on xylitol production was examined in this experiment: the same fermentation medium is adopted, other fermentation conditions are the same, liquid seeds of M7 mutagenesis strains are respectively inoculated according to inoculum sizes of 5%, 10% and 15% (V/V), the influence of different inoculum sizes on xylitol yield is examined, and the result shows that the fermentation result is the best when the inoculum size is 15%.
Therefore, in the optimized fermentation medium, the basic medium consists of the following components in parts by mass: 25% of xylose mother liquor, 0.5% of corn steep liquor dry powder, 0.5% of urea and 0.025% of monopotassium phosphate, and the solvent is water; the composition of the nutritional supplement, based on the final concentration in the basal medium, is: amino acid mixture 5mL/L, VB 2 10mg/L、VB 3 5mg/L. The conditions of fermentation culture are as follows: 200-300 rpm, 28-33 DEG CAnd (3) performing activation culture for 70-80 h under the pH condition of 5.5-6.5. The inoculation amount of the optimal strain is 15% V/V during fermentation culture.
In summary, the method for converting xylose mother liquor into xylitol through biological fermentation provided by the invention uses Kluyveromyces marxianus as a fermentation strain, and firstly uses nitrosoguanidine to mutagenize the strain to obtain a base mutant strain; then, a TC staining method is adopted for screening to obtain strains with vigorous respiratory enzyme activity, so that blindness of common screening is reduced; finally, according to the way of converting xylose into xylitol, the conversion of xylose into xylitol can be further promoted by adding the amino acid mixture and the vitamins. The method not only omits the xylose purification step, but also simplifies the xylitol separation step, can effectively reduce the production cost of the xylitol, and has the advantages of higher content and purity of the xylitol in the obtained product, simple, safe and energy-saving production process, and good application prospect in the fields of reuse of xylose mother liquor and production of the xylitol.

Claims (10)

1. A method for converting xylose mother liquor to xylitol by biological fermentation, comprising the steps of:
(1) Carrying out chemical mutagenesis treatment on Kluyveromyces marxianus by adopting nitrosoguanidine to obtain a mutagenized strain;
(2) Dyeing the strain subjected to mutagenesis in the step (1) by adopting 2,3, 5-triphenyltetrazolium chloride, and screening according to a dyeing result to obtain a colony with vigorous respiratory enzyme activity;
(3) Respectively inoculating the colonies with vigorous respiratory enzyme activity obtained in the step (2) into a screening culture medium containing xylose mother liquor for culture, measuring the xylitol content and purity in the obtained fermentation liquor after fermentation, and screening according to the content and purity measurement results to obtain the optimal strain;
(4) And (3) inoculating the optimal strain obtained by screening in the step (3) into a fermentation culture medium containing xylose mother liquor for fermentation culture, and thus, converting the xylose mother liquor into xylitol.
2. The method for converting xylose mother liquor into xylitol by biological fermentation according to claim 1, wherein in step (1), the chemical mutagenesis treatment is to add 0.8-1.2 mg/mL nitrosoguanidine solution into the bacterial suspension of kluyveromyces marxianus, and treat in water bath at 35-40 ℃ for 20-40 min.
3. The method for converting xylose mother liquor into xylitol by biological fermentation according to claim 1 or 2, wherein in step (1), the kluyveromyces marxianus is previously subjected to an activation treatment; the activation treatment is to inoculate Kluyveromyces marxianus into an activation culture medium, and perform activation culture for 10-15 h at the temperature of 28-32 ℃ at 200-300 rpm; the composition of the activation medium is as follows: glucose 9-12 g, yeast extract 0.3-0.7 g, peptone 0.3-0.5 g, and water to constant volume to 50mL.
4. The method for converting xylose mother liquor into xylitol by biological fermentation according to claim 1, wherein in step (2), the dyeing treatment is carried out after mixing an aqueous solution of 2,3, 5-triphenyltetrazolium chloride with a bacterial suspension of the strain after mutagenesis; the dyeing treatment time is 1-3 d; the mass concentration of the 2,3, 5-triphenyl tetrazolium chloride aqueous solution is 0.8% -1.2%.
5. The method for converting xylose mother liquor into xylitol by biological fermentation according to claim 1, wherein in step (3), the screening medium consists of the following components in parts by mass: 16 to 20 percent of xylose mother liquor, 0.3 to 0.8 percent of corn steep liquor dry powder, 0.03 to 0.08 percent of monopotassium phosphate and water as solvent.
6. The method for converting xylose mother liquor to xylitol by biological fermentation according to claim 1, wherein in step (3), the conditions of said cultivation are: culturing at 200-300 rpm and 28-33 deg.c for 70-80 hr.
7. The passliving being according to claim 1A method for converting xylose mother liquor into xylitol by fermentation, which is characterized in that in the step (4), the fermentation medium consists of a basic medium and nutrition additives; the basic culture medium consists of the following components in parts by mass: 23 to 28 percent of xylose mother liquor, 0.3 to 0.8 percent of corn steep liquor dry powder, 0.4 to 0.6 percent of urea, 0.02 to 0.05 percent of monopotassium phosphate and water as solvent; the composition of the nutritional supplement, based on the final concentration in the basal medium, is: amino acid mixed solution 4-6 mL/L, VB 2 8～12mg/L、VB 3 4～6mg/L。
8. The method for converting xylose mother liquor into xylitol by biological fermentation according to claim 7, wherein the basic culture medium consists of the following components in parts by mass: 25% of xylose mother liquor, 0.5% of corn steep liquor dry powder, 0.5% of urea and 0.025% of monopotassium phosphate, and the solvent is water; the composition of the nutritional supplement, based on the final concentration in the basal medium, is: amino acid mixture 5mL/L, VB 2 10mg/L、VB 3 5mg/L。
9. The method for converting xylose mother liquor into xylitol by biological fermentation according to claim 1, wherein in step (4), the conditions of the fermentation culture are: fermenting and culturing at 200-300 rpm, 28-33 deg.c and pH of 5.5-6.5 for 70-80 hr.
10. The method for converting xylose mother liquor into xylitol by biological fermentation according to claim 1, wherein in step (4), the inoculation amount of the optimal strain is 13% -17% V/V during the fermentation culture.
CN202311681748.8A 2023-12-08 2023-12-08 Method for converting xylose mother liquor into xylitol through biological fermentation Pending CN117646040A (en)

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