Organic-nitrogen-source-free culture medium for producing 1, 3-propylene glycol through clostridium butyricum fermentation and application fermentation method of organic-nitrogen-source-free culture medium

Abstract

The invention belongs to the technical field of bioengineering, and particularly discloses an organic nitrogen source-free culture medium for producing 1, 3-propylene glycol through clostridium butyricum fermentation and an application fermentation method of the organic nitrogen source-free culture medium. A mixed vitamin solution is added into a culture medium for producing 1, 3-propylene glycol through clostridium butyricum fermentation, the mixed vitamin solution contains vitamin B1 and vitamin B7, the concentration of the vitamin B1 is 20-837 mg / L, the concentration of the vitamin B7 is 4-56.21 mg / L, the addition amount of the mixed vitamin solution is 1 mL / L, and the pH value of the mixed vitamin solution is 3.0-7.5. The invention also discloses a method for producing 1, 3-propylene glycol through clostridium butyricum fermentation by using the culture medium, sugar and glycerol are used as co-substrates for fermentation, the conversion rate of glycerol is improved, and the cost of raw materials is reduced. According to the method, the fermentation cost of the 1, 3-propylene glycol can be reduced, and the separation and decolorization cost of the 1, 3-propylene glycol in the later period can be reduced, so that the economic benefit is improved, and the method has a good industrial application prospect.

Description

Technical Field

[0001] The invention belongs to the technical field of bioengineering, and specifically discloses an organic nitrogen source-free culture medium for producing 1,3-propanediol by fermentation of Clostridium butyricum and an application fermentation method thereof. Background Art

[0002] 1,3-propylene glycol (1,3-PDO) is an important chemical raw material. It can be used as a solvent, antifreeze agent, protective agent, etc. It can also be used as a monomer to synthesize polyester, polyether, polyurethane, etc. Its most important use is as a monomer to synthesize polytrimethylene terephthalate (PTT). Compared with traditional fiber materials, PTT has many characteristics such as superior UV resistance, low static electricity, low surface tension, strong anti-fouling ability, good resilience, biodegradability and room temperature dyeing. It has huge market prospects in the fields of clothing, carpets, interior decoration, etc., so the market demand for 1,3-propylene glycol continues to rise.

[0003] In the industrial production process of 1,3-propylene glycol, cost is a key factor in industrialization. In the process of industrial microbial fermentation of glycerol to produce 1,3-propylene glycol, the commonly used culture medium mainly uses yeast extract as an organic nitrogen source to provide nutrients such as proteins, peptides, amino acids, nucleotides, vitamins, trace elements, etc. for microbial growth and metabolism, and the addition amount is usually 2-4g / L. Yeast extract is a relatively expensive organic nitrogen source. For example, the price of Angel yeast extract FM888 is about 60 yuan / kg. The use of yeast extract as an organic nitrogen source increases the cost of auxiliary materials for microbial fermentation to produce 1,3-propylene glycol. The raw material cost of fermentation to produce 1,3-propylene glycol is mainly the substrate glycerol, followed by the organic nitrogen source yeast extract. In addition, the color of the fermentation broth with yeast extract is usually yellow, and 1,3-propylene glycol needs to be decolorized in the later stage, which increases the separation cost.

[0004] Therefore, a low-cost organic nitrogen source-free culture medium and a method for applying it to the fermentation of sugar and glycerol as co-substrates to produce 1,3-propylene glycol are developed to further improve the economic efficiency of industrial microbial fermentation to produce 1,3-propylene glycol, reduce the color of the fermentation broth after fermentation, and reduce the difficulty of separating and decolorizing 1,3-propylene glycol in the later stage, thereby reducing the fermentation and separation costs of 1,3-propylene glycol, which is of great practical significance for the industrial production of 1,3-propylene glycol. Summary of the invention

[0005] The purpose of the present invention is to overcome the shortcomings of the prior art and provide an improved fermentation medium and a co-substrate fermentation method for producing 1,3-propanediol by fermenting carbon sources with Clostridium butyricum based on the improved medium, so that Clostridium butyricum can produce 1,3-propanediol that is easy to separate and decolorize in the later stage with higher economy and efficiency.

[0006] In order to achieve the above object, the present invention adopts the following technical solutions:

[0007] A culture medium without an organic nitrogen source for producing 1,3-propanediol by fermentation of Clostridium butyricum, wherein a mixed vitamin solution is added to the culture medium for producing 1,3-propanediol by fermentation of carbon source of Clostridium butyricum to replace the organic nitrogen source, wherein the mixed vitamin solution contains vitamin B1 and vitamin B7, wherein each liter of the mixed vitamin solution contains: 20-837 mg of vitamin B1 and 4-56.21 mg of vitamin B7, the addition amount of the mixed vitamin solution is 1 mL / L, and the pH of the mixed vitamin solution is 3.0-7.5;

[0008] The butyric clostridium is butyric clostridium DL07, and its preservation number is CGMCC NO:17934.

[0009] Furthermore, the mixed vitamin solution also contains at least one of vitamin B5, vitamin B6 and vitamin B9.

[0010] Furthermore, each liter of the mixed vitamin solution contains: vitamin B5 3-768 mg, vitamin B6 1-300 mg or vitamin B9 1-60 mg.

[0011] Furthermore, the culture medium contains a carbon source, which is glycerol or a mixture of glycerol and sugars, with a glycerol concentration of 20-80 g / L and a sugar concentration of 0-80 g / L.

[0012] Furthermore, the sugar is any one of glucose, fructose, sucrose, maltose, galactose, lactose, xylose, mannose, starch hydrolysate, and lignocellulose hydrolysate.

[0013] A method for producing 1,3-propylene glycol by fermenting Clostridium butyricum, wherein the fermentation is carried out under anaerobic conditions, Clostridium butyricum is used as the fermentation strain, and is inoculated into the culture medium according to any one of claims 1 to 5 for batch or batch fed-batch fermentation, the fermentation culture temperature is 34-39°C, the stirring speed is 150-300 r / min, and the pH of the fermentation liquid is controlled to be 6.3-7.5.

[0014] Furthermore, when the fermentation method is batch feeding fermentation, the feed is glycerol, and the glycerol concentration in the fermentation broth is maintained at 10-40 g / L.

[0015] Furthermore, when the fermentation method is batch feeding fermentation, the feed is a mixture of glycerol and sugar, and the ratio of glycerol to sugar is (20-1):1 (g / g).

[0016] Compared with the prior art, the present invention has the following beneficial effects:

[0017] 1. The culture medium components are simple, easy to obtain and inexpensive. Low-cost vitamin solution is used to replace the expensive organic nitrogen source yeast extract powder in the culture medium. When the conversion rate is similar, the cost of the improved culture medium without organic nitrogen source is reduced by nearly 99% compared with the traditional culture medium containing organic nitrogen source yeast extract powder. At the same time, relatively cheap sugar solution is used to partially replace glycerol to improve the conversion rate of glycerol, which can significantly reduce the fermentation cost of industrial microbial fermentation to produce 1,3-propylene glycol.

[0018] 2. Using vitamin solution to replace the expensive organic nitrogen source yeast extract powder in the culture medium to ferment Clostridium butyricum to produce 1,3-propylene glycol can obtain a lighter-colored 1,3-propylene glycol fermentation broth, thereby reducing the difficulty and cost of subsequent 1,3-propylene glycol separation. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 Fermentation broth of 1,3-propanediol produced by Clostridium butyricum using the improved culture medium of Example 4;

[0020] Figure 2 The fermentation broth of Clostridium butyricum producing 1,3-propanediol using organic nitrogen source yeast extract fermentation medium; DETAILED DESCRIPTION

[0021] The present invention is further described in detail below in conjunction with the examples and drawings, but the embodiments of the present invention are not limited thereto. Where specific conditions are not specified in the examples, the experimental methods used are all conventional methods, and the reagents or instruments used can be conventional products purchased from the market.

[0022] The strain used in the fermentation is Clostridium butyricum DL07, and its preservation number is CGMCC NO:17934.

[0023] Fermentation culture method:

[0024] Batch feeding fermentation uses a 5L fully automatic fermentation tank with a liquid volume of 2L and sterilized at 121℃ for 30min. The inoculation amount is 10% (v / v), the temperature is 34-39℃, the rotation speed is 150-300r / min, and the pH is controlled at 6.3-7.5 with a pH regulator during the fermentation process. 0.05vvm ordinary nitrogen is introduced within 1h before and after inoculation to create an anaerobic environment in the tank.

[0025] Batch fermentation was carried out in a 250 mL anaerobic bottle with a liquid volume of 100 mL. 0.005 vvm nitrogen was introduced into the anaerobic bottle containing the culture medium for 3 minutes and sterilized at 121°C for 30 minutes. The inoculation amount was 10% (v / v), the temperature was 34-39°C, the rotation speed was 150-300 r / min, and the pH of the fermentation liquid was controlled at 6.3-7.5.

[0026] Glycerol composition (w / w): 80% glycerol, 15-17% water, 0.87% ash, <0.1% free fatty acids, pH 6.91.

[0027] The biomass determination method adopted in the present invention is:

[0028] The biomass was measured by turbidimetry. An appropriate amount of culture was diluted to a certain multiple, and its absorbance was measured at 600 nm using a spectrophotometer (the absorbance value range was 0.2 to 0.8). The absorbance reading was multiplied by the dilution multiple to obtain the OD 600 value of the culture solution.

[0029] The determination method of glycerol and 1,3-propylene glycol used in the present invention is:

[0030] The concentrations of glycerol and 1,3-propylene glycol were determined by high performance liquid chromatography (HPLC). The liquid chromatograph was Thermo Fisher Ultimate 3000, the detector was Shodex RI-101, the chromatographic column was American Bio-Rad AminexR HPX-87H organic acid and alcohol analysis column, the column temperature was 55°C, the mobile phase was 5mM sulfuric acid, the flow rate was 0.6mL / min, and the injection volume was 20μL.

[0031] Drawing of standard curve: Analytically pure samples (glycerol and 1,3-propylene glycol) were prepared into solutions with concentrations of 0.05 g / L, 0.1 g / L, 0.2 g / L, 0.4 g / L, 0.6 g / L, 0.8 g / L, and 1 g / L, respectively. Chromatographic analysis was performed together with the samples in each test to draw a concentration-peak area standard curve.

[0032] Sample treatment: Take 1 mL of fermentation broth, centrifuge at 12000 rpm for 6 min, take the supernatant and dilute it with the mobile phase by a corresponding multiple, filter it with a 13 mm, 0.2 μm sterile syringe filter to remove impurities, and use liquid chromatography to determine the substrate and product content in the fermentation broth.

[0033] The B vitamins detection method used in the present invention is:

[0034] Sample pretreatment: At the beginning and end of fermentation, take 500 mL of fermentation liquid respectively, filter with a 0.22 μm water filter membrane, place the filtrate in a 500 mL anaerobic bottle, seal it, and send it to Dalian Pony Testing Co., Ltd. for B vitamin determination. Sample testing is conducted by Pony Testing Co., Ltd., using an IE-287 UV spectrophotometer for detection. The detection methods are as follows: thiamine (VB1) uses GB 5009.84-2016; riboflavin (VB2) uses GB 5009.85-2006; niacin (VB3) uses GB5009.89-2016; pantothenic acid (VB5) uses GB 5009.210-2016; biotin (VB7) uses GB5009.259-2016; folic acid (VB9) uses GB 5009.211-2014; VB 12 Adopt GB 5413.14-2010.

[0035] Example 1

[0036] Fermentation medium composition (g / L): glycerol 20-80, KH2PO4 1.36, (NH4)2SO4 6.61, MgCl2·6H2O0.26, CaCl2 0.29, citric acid 0.42, in addition, add 5 mL of mixed trace element solution, 1 mL of mixed vitamin solution and 10 mL of saturated hydrochloric acid solution.

[0037] Among them, the composition of the mixed trace element solution (g / L): ZnCl2 0.68, MnCl2·4H2O 0.17, H3BO3 0.06, CuCl2·2H2O 0.47, NaMoO4·2H2O 0.005, FeCl2·4H2O 3.97, CoCl2·6H2O 0.47.

[0038] The composition of the mixed vitamin solution: vitamin B1 837 mg / L, vitamin B2 10 mg / L, vitamin B5 768 mg / L, vitamin B6 300 mg / L, vitamin B7 20 mg / L and vitamin B9 60 mg / L.

[0039] The mixed vitamin solution must be filtered through a 0.2 μm filter membrane before being added to the sterilized fermentation medium.

[0040] Example 2

[0041] The components of the fermentation medium were the same as those in Example 1, except that the composition of the mixed vitamin solution was changed to include 500 mg / L vitamin B1, 4 mg / L vitamin B7 and 15 mg / L vitamin B5.

[0042] Example 3

[0043] The components of the fermentation medium were the same as those in Example 1, except that the composition of the mixed vitamin solution was changed to include 100 mg / L vitamin B1, 20 mg / L vitamin B7 and 3 mg / L vitamin B5.

[0044] Example 4

[0045] The components of the fermentation medium were the same as those in Example 1, except that the composition of the mixed vitamin solution was changed to include 100 mg / L vitamin B1 and 50 mg / L vitamin B7.

[0046] Example 5

[0047] The components of the fermentation medium were the same as those in Example 1, except that the composition of the mixed vitamin solution was changed to 20 mg / L vitamin B1 and 35 mg / L vitamin B7.

[0048] Example 6

[0049] The components of the fermentation medium were the same as those in Example 1, except for the composition of the mixed vitamin solution: 500 mg / L vitamin B1 and 50 mg / L vitamin B7.

[0050] Example 7

[0051] The components of the fermentation medium were the same as those in Example 1, except that the composition of the mixed vitamin solution was changed to include 300 mg / L vitamin B1 and 56.21 mg / L vitamin B7.

[0052] Example 8

[0053] The components of the fermentation medium were the same as those in Example 1, except that the composition of the mixed vitamin solution was changed to include 280 mg / L vitamin B1 and 32 mg / L vitamin B7.

[0054] Comparative Example 1

[0055] Fermentation medium containing organic nitrogen source (g / L): glycerol 20-80, KH2PO4 1.36, (NH4)2SO4 6.61, MgCl2·6H2O 0.26, CaCl2 0.29, citric acid 0.42, yeast extract powder (Angel Yeast FM888) 2, and 5 mL of mixed trace element solution and 10 mL of saturated hydrochloric acid solution.

[0056] Among them, the composition of the mixed trace element solution (g / L): ZnCl2 0.68, MnCl2·4H2O 0.17, H3BO3 0.06, CuCl2·2H2O 0.47, NaMoO4·2H2O 0.005, FeCl2·4H2O 3.97, CoCl2·6H2O 0.47.

[0057] Example 9

[0058] The culture medium of Example 1 was used for batch feed fermentation to produce 1,3-propylene glycol. The activated Clostridium butyricum DL07 was inoculated into a fermenter containing the culture medium of Example 1 at a 10% (v / v) inoculation rate. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation, and 5 mol / L sodium hydroxide was used as a pH regulator. The culture conditions were: pH 6.5, temperature 37°C, and stirring speed 250 r / min. The initial glycerol concentration of the fermentation culture was 40 g / L. When the glycerol concentration dropped to 10 g / L, 80% glycerol was started to be fed to maintain the glycerol concentration at 20 g / L. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 87.28 g / L, and the conversion rate was 0.521 g / g glycerol.

[0059] The content of B vitamins in the fermented liquid before and after the fermentation of the embodiment is measured, and it is found that VB1 is fully consumed when the fermentation ends, which shows that VB1 may be involved in the fermentation of Clostridium butyricum DL07. In addition, there are also 4 kinds of B vitamins (VB5, VB6, VB7 and VB9) whose concentrations decrease after fermentation, and wherein VB5 consumes the most, and may be involved in cell metabolism to a certain extent. VB7 is the auxiliary factor of carboxylase, and usually catalyzes the carboxylation reaction that carbon dioxide participates in, and is closely related to cell growth and energy metabolism.

[0060] Example 10

[0061] The culture medium of Example 2 was used for batch fermentation to produce 1,3-propylene glycol. During fermentation, 5 mol / L sodium hydroxide was used as a pH regulator. The activated Clostridium butyricum DL07 was inoculated at a rate of 10% (v / v) into a fermenter containing the culture medium of Example 2. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation. The culture conditions were: pH 7.0, temperature 39°C, stirring speed 250 r / min, and initial glycerol concentration of 40 g / L. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 19.75 g / L, and the conversion rate was 0.490 g / g glycerol.

[0062] Embodiment 11

[0063] The culture medium of Example 3 was used for batch fermentation to produce 1,3-propylene glycol. The activated Clostridium butyricum DL07 was inoculated into a fermenter containing the culture medium of Example 3 at a 10% (v / v) inoculation rate. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation. The culture conditions were: pH 6.3, temperature 34°C, stirring speed 300 r / min, and initial glycerol concentration of 40 g / L. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 20.59 g / L, and the conversion rate was 0.511 g / g glycerol.

[0064] Example 12

[0065] The culture medium of Example 4 was used for batch feed fermentation to produce 1,3-propylene glycol. The activated Clostridium butyricum DL07 was inoculated into a fermenter containing the culture medium of Example 4 at a 10% (v / v) inoculation rate. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation, and 5 mol / L sodium hydroxide was used as a pH regulator. The culture conditions were: pH 6.5, temperature 37°C, and stirring speed 150 r / min. The initial glycerol concentration of the fermentation culture was 20 g / L. When the glycerol concentration dropped to 10 g / L, 80% glycerol was started to be fed to maintain the glycerol concentration at 20 g / L. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 72.85 g / L, and the conversion rate was 0.516 g / g glycerol.

[0066] In addition, the color of the fermentation liquid after fermentation is Figure 1 As shown, it is a very light yellow color. Compared with traditional organic nitrogen source culture medium, the lighter color facilitates the subsequent purification of 1,3-propanediol, especially alleviating the decolorization burden.

[0067] Embodiment 13

[0068] The culture medium of Example 5 was used for batch feed fermentation to produce 1,3-propylene glycol. The activated Clostridium butyricum DL07 was inoculated into a fermenter containing the culture medium of Example 5 at a 10% (v / v) inoculation rate. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation, and 5 mol / L sodium hydroxide was used as a pH regulator. The culture conditions were: pH 6.5, temperature 37°C, and stirring speed 200 r / min. The initial glycerol concentration of the fermentation culture was 60 g / L. When the glycerol concentration was reduced to 20 g / L, 80% glycerol and glucose mixture was started to be fed, wherein the mass ratio of glycerol to sugar was 5, and the glycerol concentration was maintained at 40 g / L. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 55.94 g / L, and the conversion rate was 0.532 g / g glycerol.

[0069] Embodiment 14

[0070] The culture medium of Example 6 was used for batch feed fermentation to produce 1,3-propylene glycol. The activated Clostridium butyricum DL07 was inoculated into a fermenter containing the culture medium of Example 6 at a 10% (v / v) inoculation rate. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation, and 5 mol / L sodium hydroxide was used as a pH regulator. The culture conditions were: pH 7, temperature 37°C, and stirring speed 250 r / min. The initial glycerol concentration of the fermentation culture was 40 g / L, and the fructose concentration was 5 g / L. When the glycerol concentration was reduced to 10 g / L, 80% glycerol was added to maintain the glycerol concentration at 20 g / L. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 72.31 g / L, and the conversion rate was 0.511 g / g glycerol.

[0071] Embodiment 15

[0072] The culture medium of Example 7 was used for batch feed fermentation to produce 1,3-propylene glycol. The activated Clostridium butyricum DL07 was inoculated into a fermenter containing the culture medium of Example 7 at a 10% (v / v) inoculation rate. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation, and 5 mol / L sodium hydroxide was used as a pH regulator. The culture conditions were: pH 6.5, temperature 37°C, and stirring speed 200 r / min. The initial glycerol concentration of the fermentation culture was 80 g / L. When the glycerol concentration dropped to 10 g / L, 80% glycerol was started to be fed to maintain the glycerol concentration at 30 g / L. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 75.29 g / L, and the conversion rate was 0.518 g / g glycerol.

[0073] Example 16

[0074] The culture medium of Example 8 was used for batch feed fermentation to produce 1,3-propylene glycol. During fermentation, 5 mol / L sodium hydroxide was used as a pH regulator. The activated Clostridium butyricum DL07 was inoculated at 10% (v / v) into a fermenter containing the culture medium of Example 8. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation. The culture conditions were: pH 6.5, temperature 37°C, stirring speed 250 r / min, initial glycerol concentration of 35 g / L, and glycerol was fed. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 83.74 g / L, and the conversion rate was 0.51 g / g glycerol.

[0075] Embodiment 17

[0076] The culture medium of Example 8 was used for batch feed fermentation to produce 1,3-propylene glycol. During fermentation, 5 mol / L sodium hydroxide was used as a pH regulator. The activated Clostridium butyricum DL07 was inoculated at a rate of 10% (v / v) into a fermenter containing the culture medium of Example 8. Nitrogen was introduced at a rate of 0.05 vvm 1 h before and after inoculation. The culture conditions were: pH 6.5, temperature 37°C, stirring speed 250 r / min, initial glycerol concentration of 40 g / L and glucose concentration of 20 g / L, and glycerol was fed. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 69.59 g / L, and the conversion rate was 0.56 g / g glycerol.

[0077] Comparative Example 2

[0078] Clostridium butyricum fermented 1,3-propanediol in an organic nitrogen source yeast extract fermentation medium. The fermentation medium was the medium of control example 1, and 5 mol / L sodium hydroxide was used as a pH regulator during fermentation. The activated Clostridium butyricum DL07 was inoculated with a 10% (v / v) inoculum into a fermenter containing an organic nitrogen source yeast extract medium. Nitrogen was introduced at a rate of 0.05 vvm 1h before and after inoculation, and 5 mol / L sodium hydroxide was used as a pH regulator. The culture conditions were: pH 7.0, temperature 37 ° C, and stirring speed 250r / min. The initial glycerol concentration of the fermentation culture was 40g / L, and batch feeding fermentation was adopted. When the glycerol concentration dropped to 20g / L, 80% glycerol was added to maintain the glycerol concentration at 20g / L. The fermentation results are shown in Table 1. The final 1,3-propanediol concentration was 84.86g / L, and the conversion rate was 0.520g / g glycerol.

[0079] From the analysis of Table 1, it can be seen that the cost of organic nitrogen source yeast extract powder for producing 1 ton of 1,3-propylene glycol is 1,500 yuan (the price of Angel yeast extract powder FM888 is 60 yuan / kg). The cost of mixed vitamins for producing 1 ton of 1,3-propylene glycol is 3.4-8.2 yuan (the price of vitamin B1 is 250 yuan / kg, the price of vitamin B2 is 300 yuan / kg, the price of vitamin B5 is 350 yuan / kg, the price of vitamin B6 is 300 yuan / kg, the price of vitamin B7 is 3000 yuan / kg, and the price of vitamin B9 is 400 yuan / kg). Under similar conversion rates, compared with the traditional culture medium containing organic nitrogen source yeast extract powder, the cost of the improved culture medium without organic nitrogen source is reduced by nearly 99%. It can be seen that the improved culture medium reduces the fermentation cost.

[0080] In addition, the color of the fermentation liquid after fermentation using the culture medium of Control Example 1 is as follows: Figure 2 As shown, the yellow color is relatively heavy, which brings difficulties to the subsequent purification of 1,3-propanediol.

[0081] Table 1 Summary of 1,3-propanediol production by Clostridium butyricum in different fermentation media

[0082]

[0083]

[0084]

Claims

1. An organic nitrogen source-free culture medium for producing 1,3-propanediol by fermentation of Clostridium butyricum, characterized in that: Adding a mixed vitamin solution to replace the organic nitrogen source in the culture medium for producing 1,3-propanediol by fermenting carbon sources with Clostridium butyricum, wherein the mixed vitamin solution contains vitamin B1 and vitamin B7, wherein each liter of the mixed vitamin solution contains: 20-837 mg of vitamin B1 and 4-56.21 mg of vitamin B7, the addition amount of the mixed vitamin solution is 1 mL / L, and the pH of the mixed vitamin solution is 3.0-7.5; The butyric clostridium is butyric clostridium DL07, and its preservation number is CGMCC NO:17934.

2. The organic nitrogen source-free culture medium for producing 1,3-propanediol by fermentation of Clostridium butyricum according to claim 1, characterized in that: The mixed vitamin solution also contains at least one of vitamin B5, vitamin B6 and vitamin B9.

3. The organic nitrogen source-free culture medium for producing 1,3-propanediol by fermentation of Clostridium butyricum according to claim 2, characterized in that: Each liter of mixed vitamin solution contains: vitamin B5 3-768 mg, vitamin B6 1-300 mg or vitamin B9 1-60 mg.

4. The organic nitrogen source-free culture medium for producing 1,3-propanediol by fermentation with Clostridium butyricum according to claim 3, characterized in that: The culture medium contains a carbon source, which is glycerol or a mixture of glycerol and sugars, with a glycerol concentration of 20-80 g / L and a sugar concentration of 0-80 g / L.

5. The organic nitrogen source-free culture medium for producing 1,3-propanediol by fermentation with Clostridium butyricum according to claim 4, characterized in that: The sugar is any one of glucose, fructose, sucrose, maltose, galactose, lactose, xylose, mannose, starch hydrolysate, and lignocellulose hydrolysate.

6. A fermentation method for producing 1,3-propanediol by fermentation with Clostridium butyricum, characterized in that: The fermentation is carried out under anaerobic conditions, Clostridium butyricum is used as the fermentation strain, which is inoculated into the culture medium according to any one of claims 1 to 5 for batch or batch fed-batch fermentation, the fermentation culture temperature is 34-39° C., the stirring speed is 150-300 r / min, and the pH of the fermentation liquid is controlled to be 6.3-7.

5.

7. The method for producing 1,3-propylene glycol by fermentation with Clostridium butyricum according to claim 6, characterized in that: When the fermentation method is batch feeding fermentation, the feed is glycerol, and the glycerol concentration in the fermentation liquid is maintained at 10-40g / L.

8. The method for producing 1,3-propylene glycol by fermentation with Clostridium butyricum according to claim 6, characterized in that: When the fermentation method is batch feeding fermentation, the feed is a mixture of glycerol and sugar, and the ratio of glycerol to sugar is (20-1):1 (g / g).

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