A method for co-producing 1,3-propylene glycol, micro-nano calcium carbonate and hydrogen using microorganisms

Abstract

The present invention discloses a method for the co-production of 1,3-propylene glycol, micro-nano calcium carbonate and hydrogen by microorganisms, comprising: inoculating the strain into a fermentation medium, fermenting with glycerol as a substrate, and using calcium hydroxide or ammonia water to couple calcium hydroxide as a pH regulator to adjust the pH value of the fermentation liquid during the fermentation process. The pH in the fermentation process is controlled by ammonia water and calcium hydroxide, the CO2 in the fermentation tail gas is fixed, the hydrogen concentration and the production intensity of 1,3-propylene glycol are increased, the electrical conductivity of the fermentation liquid is reduced, and 1,3-propylene glycol, micro-nano calcium carbonate and hydrogen are co-produced. In the present invention, calcium hydroxide reacts with carbon dioxide to form insoluble calcium carbonate, which can be separated by precipitation or low-speed centrifugation to obtain micro-nano calcium carbonate, and the supernatant is directly evaporated to obtain 1,3-propylene glycol, and the fermentation tail gas is collected to obtain hydrogen. The separation step is simple and easy, the production cost is low, and there is good industrial application prospect.

Description

Technical Field

[0001] The invention belongs to the technical field of bioengineering, and in particular relates to a method for co-producing 1,3-propylene glycol, micro-nano calcium carbonate and hydrogen by microorganisms. Background Art

[0002] 1,3-Propanediol (1,3-PDO) is an important chemical and pharmaceutical intermediate, used as a solvent, antifreeze agent, protective agent, and monomer in the synthesis of polyesters and polyurethanes. Polytrimethylene terephthalate (PTT), synthesized from 1,3-PDO, offers superior properties compared to traditional fiber materials, including UV resistance, low static electricity, low surface tension, strong stain resistance, good resilience, biodegradability, and room-temperature dyeability. This material holds significant market potential in the carpet and textile industries, driving a continuously increasing demand for 1,3-PDO.

[0003] Currently, there are two methods for the industrial production of 1,3-propylene glycol (1,3-PD)—chemical and biological. Chemical methods have harsh synthesis conditions, produce numerous byproducts, and are difficult to separate. Compared to chemical methods, biological methods for producing 1,3-PD offer advantages such as renewable raw materials, fewer byproducts, mild and safe reaction conditions, and minimal environmental pollution. These advantages have led to increasing research, but they pose challenges for industrial production due to their low substrate utilization, complex separation steps, and high separation costs, resulting in high production costs. During the bioconversion of glycerol to 1,3-PD, in addition to approximately 50% of the glycerol being converted into the main product, 1,3-PD, half of the substrate is converted into microbial cells, byproducts, and carbon dioxide and hydrogen in the exhaust gas. The proportion of carbon dioxide in fermentation exhaust gas exceeds 90%, and direct emission during large-scale production would have an impact on the local environment. In the separation process of 1,3-propylene glycol, unit operations such as membrane filtration (microfiltration and ultrafiltration), vacuum evaporation concentration, and distillation are usually used. Among them, membrane filtration has problems such as large equipment investment, regular renewal of membrane components, high power consumption, dilution of fermentation liquid, and wastewater generated by membrane cleaning. The concentration operation will increase the chloride concentration in the traditional fermentation culture medium. Long-term operation will cause corrosion to industrial production equipment, which is bound to increase equipment investment.

[0004] In the process of microbial fermentation to produce 1,3-propylene glycol, sodium hydroxide or potassium hydroxide is usually used to neutralize the organic acids in the fermentation broth. These soluble alkalis can react with the carbon dioxide partially dissolved in the fermentation broth to form soluble sodium carbonate or potassium carbonate, which significantly increases the salt concentration during the subsequent concentration operation, salting out proteins and polysaccharides, making the subsequent separation process difficult to proceed smoothly, and requiring the addition of desalting or deproteinization steps.

[0005] Nano-calcium carbonate is an important inorganic chemical material, widely used in industries such as rubber, plastics, papermaking, coatings, paints, inks, printing, food, medicine, cosmetics, and feed. Hydrogen is a clean, environmentally friendly fuel, characterized by its pollution-free, low-cost, and renewable nature, and has attracted widespread attention as a fossil fuel alternative. From an economic maximization perspective, the co-production of 1,3-propylene glycol (PD), micro-nano-calcium carbonate, and hydrogen will undoubtedly improve substrate atom economy and make solid-liquid separation and concentration operations easier and more cost-effective, thereby reducing production costs. This has important practical implications for the industrial production of 1,3-PD. Summary of the Invention

[0006] To address the problems of low production intensity, complex separation steps, and high separation costs in the production of 1,3-propylene glycol by microbial fermentation, which result in high production costs and low economic benefits, the present invention provides a method for the co-production of 1,3-propylene glycol, micro-nano calcium carbonate, and hydrogen.

[0007] The technical solutions of the present invention are as follows:

[0008] A method for co-producing 1,3-propylene glycol, micro-nano calcium carbonate and hydrogen by microorganisms comprises the steps of inoculating bacterial strains into a fermentation medium and fermenting with glycerol as a substrate. The method is characterized in that during the fermentation process, calcium hydroxide or ammonia water coupled with calcium hydroxide is used as a pH regulator to adjust the pH value of the fermentation liquid to 6-7, preferably 7.

[0009] Furthermore, in the above technical solution, the bacterial species employed is Clostridium butyricum or Klebsiella. Preferably, 1,3-propylene glycol is produced by batch fed-batch fermentation using pure or crude glycerol as a substrate. The Clostridium butyricum or Klebsiella is not specifically limited; any Clostridium butyricum or Klebsiella known in the art for fermentation production of 1,3-propylene glycol can be used.

[0010] Furthermore, in the above technical solution, the crude glycerol is a by-product of biodiesel glycerol or a product of oil hydrolysis with a glycerol content of 75-95%.

[0011] Furthermore, in the above technical solution, the fermentation uses calcium hydroxide or ammonia water coupled with calcium hydroxide to adjust the pH, and the culture medium does not need to be added with an inorganic nitrogen source.

[0012] Furthermore, in the above technical solution, when the pH regulator is ammonia water coupled with calcium hydroxide, an inorganic nitrogen source is added or not added to the fermentation medium, and during the fermentation process, ammonia water with a concentration of 22-28% and calcium hydroxide with a concentration of 1.0-6.0 mol / L are used alternately to adjust the pH value of the fermentation liquid, wherein the pH value of the fermentation liquid is adjusted with ammonia water between 0-5 hours after the start of fermentation, preferably 3 hours, and the amount of ammonia water added is 5-13.5 g / L of fermentation liquid, preferably 6.5 g / L.

[0013] Furthermore, in the above technical solution, when the pH regulator is calcium hydroxide, the concentration of the calcium hydroxide is 1.0-6.0 mol / L, preferably 5 mol / L.

[0014] Furthermore, in the above technical solution, after calcium carbonate and bacteria are separated from the fermentation liquid, 1,3-propylene glycol is obtained by evaporation, and the fermentation tail gas is collected to obtain hydrogen.

[0015] Furthermore, in the above technical solution, the fermentation broth is subjected to precipitation and / or centrifugation to separate calcium carbonate and bacterial cells. The specific method is as follows: the fermentation broth is allowed to stand for 5 to 60 minutes to allow precipitation, filtered to separate the precipitated calcium carbonate, and the fermentation broth from which calcium carbonate has been removed is subjected to low-speed centrifugation to separate the bacterial cells, wherein the low-speed centrifugation conditions are: a relative centrifugal force of 1000-3000g and a centrifugation time of 3-30 minutes; or the fermentation broth is centrifuged to separate the bacterial cells or a mixture of bacterial cells and calcium carbonate, wherein the centrifugation conditions are: a relative centrifugal force of 1000-3000g and a centrifugation time of 3-30 minutes. The fermentation broth from which calcium carbonate and bacterial cells have been removed is evaporated to obtain concentrated 1,3-propylene glycol, with a concentration factor of up to 5-7 times. In addition, the gases generated during the fermentation process are collected through the exhaust outlet of the fermentor and further separated to obtain hydrogen.

[0016] Furthermore, in the above technical solution, the hydrogen content in the fermentation tail gas is 10-40% (v / v).

[0017] Furthermore, in the above technical solution, the calcium carbonate obtained by precipitation and / or centrifugation of the fermentation broth is micro-nanoscale calcium carbonate crystals, which are in the form of flakes or spindles and have a size of 300 nm-50 μm.

[0018] Furthermore, in the above technical solution, the bacterial species is Clostridium butyricum or Klebsiella, and the fermentation method adopts batch feeding fermentation. During the fermentation process, the stirring speed is 100-350r / min, the culture temperature is 30-38°C, the pH is 6-7, and nitrogen is introduced into the fermentation process at 0.1vvm throughout the process or 1h before and after inoculation. Specifically, the fermentation tank is filled with a sterilized fermentation medium with a volume fraction of 30-80%, and 0.1vvm nitrogen is introduced within 1h before and after inoculation or throughout the fermentation process to create an anaerobic environment in the tank. The bacterial seed liquid is connected to the fermentation tank at an inoculum size of 1-10% for continuous fed-batch fermentation. The fermentation temperature is 30-38°C, preferably 37°C, the rotation speed is 100-350r / min, the fermentation pH is controlled to 6-7, preferably 7, by a pH regulator solution, and the culture time is 13-23h.

[0019] Furthermore, in the above technical solution, the inorganic salts in the fermentation medium are replaced by sulfates as chloride salts. Similarly, the inorganic salts in the seed culture medium used for bacterial activation are replaced by sulfates as chloride salts. That is, the inorganic salts added as chloride salts to the traditional fermentation medium or seed culture medium are replaced with sulfates to obtain the improved fermentation medium and improved seed culture medium. Furthermore, the components of the improved fermentation medium and improved seed culture medium include sulfuric acid instead of hydrochloric acid in the traditional culture medium.

[0020] Beneficial effects of the present invention:

[0021] (1) Utilizing alkaline calcium salts as pH regulators for fermentation medium, calcium hydroxide reacts with carbon dioxide produced during the fermentation process to form valuable insoluble calcium carbonate, which can fix CO2 in the fermentation tail gas, thereby increasing the hydrogen concentration in the fermentation tail gas and the production intensity of 1,3-propylene glycol. Simultaneously, due to precipitation in the form of insoluble salts, the conductivity of the fermentation liquid can be reduced. In addition, calcium ions can not only react with sulfate, phosphate, citrate, etc. in the culture medium to form insoluble salts, thereby reducing the salt concentration in the fermentation liquid, but also have a flocculation and sedimentation effect on proteins and cells, thereby facilitating the subsequent separation of 1,3-propylene glycol. The present invention can provide a highly economical fermentation model for the production of 1,3-propylene glycol by microorganisms.

[0022] (2) The present invention provides an improved culture medium formula, in which the inorganic salts in the improved culture medium are replaced by sulfates, which are highly corrosive to equipment in traditional culture media, without affecting the production of fermentation products, thereby solving the problem of corrosion of industrial production equipment by traditional culture media.

[0023] (3) The present invention provides a simple, low-cost product separation method, which can directly collect and obtain hydrogen. The fermentation broth is separated by precipitation and / or centrifugation to separate calcium carbonate and bacteria. The supernatant can be directly evaporated to concentrate to 5-7 times, and further separated to obtain 1,3-propylene glycol. This separation process can simplify the separation steps of products in the fermentation broth, reduce production costs, and facilitate industrial scale-up application. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The graph shows the change curve of 1,3-propylene glycol concentration in the fermentation broth of Examples 1, 7, 9, and 10.

[0025] Figure 2 The conductivity change curves of the fermentation broths of Examples 1, 3, 4, and 8 are shown.

[0026] Figure 3 A scanning electron microscope image showing the calcium carbonate crystals generated in Example 6.

[0027] Figure 4 This is a scanning electron microscope image of the calcium carbonate crystals generated in Example 9.

[0028] Figure 5 This is a scanning electron microscope image of the calcium carbonate crystals generated in Example 10.

[0029] Figure 6 It shows the evaporation phenomenon of the fermentation liquid of Example 1.

[0030] Figure 7 It shows the evaporation phenomenon of the fermentation liquid of Example 4. DETAILED DESCRIPTION

[0031] In the following examples, unless otherwise specified, the experimental methods used are conventional methods, and the reagents used can be purchased from chemical or biological reagent companies. In addition, it will be apparent to those skilled in the art that various changes or modifications to the components and amounts of the culture media in these embodiments, without departing from the spirit and scope of the present invention, are also within the scope of protection of the present invention.

[0032] The specific implementation of the present invention is described in detail below in conjunction with the technical solution.

[0033] 1. Culture medium used in the following examples:

[0034] Traditional seed culture medium (g / L): glycerol 20-45, KH2PO4·3H2O 1.3, K2HPO4 4.454, (NH4)2SO42, MgSO4·7H2O 0.2, yeast powder 1, 0.5g / L L-cysteine ​​hydrochloride, and 2mL trace element A solution, 1mL Fe 2+ solution and 1 mL of Ca 2+The composition of trace element A solution (mg / L): CuCl2·2H2O 20, ZnCl2 70, MnCl2·4H2O 100, H3BO3 60, CoCl2·6H2O 200, NiCl2·6H2O 25, NaMoO4·2H2O 35, saturated hydrochloric acid 9mL. 2+ The solution is 5g / L FeSO4·7H2O prepared with 4mL saturated hydrochloric acid. 2+ The solution is 20g / L CaCl2 solution. Add 0.2g CaCO3 per 100mL of anaerobic bottle.

[0035] Traditional fermentation medium (g / L): 35-80% glycerol, 1.36% KH2PO4·3H2O, 6.61% (NH4)2SO4, 0.26% MgCl2·6H2O, 0.29% CaCl2, 0.42% citric acid, 2% yeast powder, and 5 mL of trace element B solution was added, where the composition of trace element B solution (g / L) was: 0.68% ZnCl2, 0.17% MnCl2·4H2O, 0.06% H3BO3, 0.47% CuCl2·2H2O, 0.005% NaMoO4·2H2O, 3.97% FeCl2·4H2O, 0.47% CoCl2·6H2O, and 10 mL of saturated hydrochloric acid.

[0036] The composition of trace element A solution in the improved seed culture medium (mg / L) is: CuSO4 22, ZnSO4·7H2O 147.68, MnSO4·H2O 85.40, H3BO3 60, CoSO4·7H2O 236.29, NiSO4·6H2O 27.64, NaMoO4·2H2O 35. Fe 2+ The solution is 5g / L FeSO4·7H2O prepared with 4mL of dilute sulfuric acid. 2+ The solution is 13.33 g / L Ca(OH)2 solution.

[0037] Improved fermentation medium (g / L): glycerol 35-80, KH2PO4·3H2O 1.36, (NH4)2SO4 6.61, MgSO4·7H2O 0.63, citric acid 0.42, yeast powder 2, and 5 mL of trace element B solution was added, where the composition of trace element B solution (g / L): ZnSO4·7H2O 1.43, MnSO4·H2O 0.14, H3BO30.06, CuSO4 0.517, NaMoO4·2H2O0.005, FeSO4·7H2O 5.55, CoSO4·7H2O0.555.

[0038] 2. Seed Activation Culture Conditions: Anaerobic fermentation. Anaerobic seed activation was performed using 250 mL vials containing 100 mL of liquid. After the seed culture medium was added, nitrogen was passed through each vial for 3 minutes to deoxygenate the vial, and then capped with a butyl rubber stopper. During the experiment, inoculation and sampling were performed using a disposable sterile syringe, with an inoculum size of 1-10% (v / v). The incubation temperature was 37°C, the shaker speed was 200 rpm, and the incubation time was 15-24 hours.

[0039] 3. Fermentation: Use a 5L fully automatic fermenter with a 2L liquid volume and an inoculum size of 1-8% (v / v). Maintain the fermentation temperature at 37°C and a rotation speed of 250 r / min. Maintain the pH at 7.0 using a pH regulator. Maintain an anaerobic environment within the tank by introducing 0.15 v / vm of ordinary nitrogen gas within 1 hour before and after inoculation.

[0040] 4. During the fermentation process, the gases produced during the fermentation process were collected through the tail gas port of the fermentation tank, and the volume content (v / v) of each gas therein was determined.

[0041] 5. The strain used in the fermentation was Clostridium butyricum DL07, with a preservation number of CGMCC NO: 17934.

[0042] 6. Centrifuge the fermentation broth under the given centrifugal conditions. The centrifugal effect is determined by measuring the absorbance of the fermentation broth at 650nm, determining the density of the bacteria in the fermentation broth before and after centrifugation, and calculating the clarification rate of the fermentation broth after centrifugation:

[0043]

[0044] 7. Crude glycerol: A product of oil hydrolysis, its composition (w / w): 80% glycerol, 15-17% water, 0.87% ash, <0.1% free fatty acids, pH 6.91.

[0045] 8. Continuous feeding: When the initial substrate concentration drops to about 20g / L, start continuous flow addition of substrate, with 1h as the feeding segmentation point. Glycerol mass A (g) needs to be fed at each segmentation point. The fermentation tank feeding pump feeds 0.5g of glycerol each time it runs. n = 3600s / (A / 0.5) is the feeding cycle for each segmentation point. Therefore, continuous feeding only needs to set the feeding cycle of each feeding segmentation point in the fermentation process to 1 time / n seconds to maintain the substrate concentration at 15-35g / L.

[0046] Example 1 Traditional fermentation

[0047] Both the seed culture medium and the fermentation culture medium were traditional culture media, and 5 mol / L sodium hydroxide was used as a pH regulator during fermentation. Clostridium butyricum DL07 stored in a glycerol bottle at low temperature was inoculated with 4% (v / v) into an anaerobic bottle containing traditional seed culture medium. After culturing for 12 hours, it was transferred to a new traditional seed culture medium at a 2% (v / v) inoculation and cultured for 12 hours. After two rounds of activation, it was inoculated with 10% (v / v) into a fermentation tank containing traditional fermentation culture medium. Nitrogen was introduced at a rate of 0.1 vvm for 1 hour 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 rotation speed 250 r / min. The initial crude glycerol concentration of the fermentation culture was 40 g / L, and the fermentation time was 27 hours. The results are shown in Table 1. The final 1,3-propylene glycol concentration was 85.08 g / L ( Figure 1 ), the production intensity is 3.15g / (L·h), the content of hydrogen is 6.22% (v / v), carbon dioxide is 92.06% (v / v), and the rest is nitrogen. At the end of fermentation, the conductivity of the fermentation liquid is 42770μs / cm( Figure 2 ), the protein concentration in the fermentation broth was 2.72 g / L.

[0048] Example 2 Fermentation using improved culture medium

[0049] Inoculation and fermentation were carried out as in Example 1, except that both the seed culture medium and the fermentation medium were modified culture media. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 88.98 g / L, the production rate was 3.29 g / (L·h), the hydrogen content was 4.80% (v / v), the carbon dioxide content was 92.71% (v / v), and the remainder was nitrogen.

[0050] Example 3 Conventional fermentation using NaOH coupled with ammonia as pH regulator

[0051] The inoculation and fermentation of the strain were carried out according to Example 1, except that concentrated ammonia water (containing 25% ammonia) was used to adjust the initial pH of the fermentation medium to 7.0 before inoculation, sodium hydroxide was used to adjust the pH after the fermentation started, concentrated ammonia water was used to adjust the pH when the fermentation was carried out for 3 hours, and sodium hydroxide was used to adjust the pH when the amount of concentrated ammonia water added reached 6.8g / L fermentation liquid. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 85.07g / L, the production intensity was 4.05g / (L·h), the hydrogen content was 4.04% (v / v), the carbon dioxide content was 94.10% (v / v), and the rest was nitrogen. At the end of the fermentation, the conductivity of the fermentation liquid was 38220μs / cm( Figure 2 ).

[0052] Example 4 Traditional fermentation with 5 mol / L Ca(OH)2 as pH regulator

[0053] The inoculation and fermentation of the strain were carried out according to Example 1, except that 5 mol / L Ca(OH)2 was used as the pH regulator in the fermentation process, and the fermentation time was 16 h. The results are shown in Table 1. The final 1,3-propylene glycol concentration was 88.28 g / L, the production intensity was 5.52 g / (L·h), the hydrogen content was 17.22% (v / v), the collected hydrogen yield was 15.87 mmol / L fermentation liquid, carbon dioxide was 1.75% (v / v), and the rest was nitrogen. After the fermentation liquid settled for 30 minutes, the mass of calcium carbonate was 76.75 g / L, and the shapes were irregular spherical and rod-shaped, with the sizes mostly distributed in the range of 10-15 μm. At the end of fermentation, the conductivity of the fermentation liquid was 20500 μs / cm( Figure 2 ), which is 48% of the pH adjuster sodium hydroxide, which facilitates the subsequent separation of 1,3-PDO. The protein concentration in the fermentation broth is 1.52g / L, which is 56% of the pH adjuster sodium hydroxide, which facilitates the subsequent separation of 1,3-PDO.

[0054] Example 5 Traditional fermentation with 5 mol / L Ca(OH)2 as pH regulator

[0055] Inoculation and fermentation were carried out as in Example 4, except that nitrogen was flowed throughout the fermentation process (0.1 vvm) after inoculation. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 84.38 g / L, and the production rate was 5.27 g / (L·h). After the fermentation broth settled for 25 minutes, the mass of calcium carbonate was 14.37 g / L, and the shapes were irregular spheres and flakes.

[0056] Example 6 Conventional fermentation with 2.5 mol / L Ca(OH)2 as pH regulator

[0057] The inoculation and fermentation of the strain were carried out according to Example 4, except that 2.5 mol / L Ca(OH)2 was used as the pH regulator during the fermentation process. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 82.24 g / L, the production intensity was 5.14 g / (L·h), the hydrogen content was 18.62% (v / v), the collected hydrogen yield was 13.65 mmol / L fermentation liquid, carbon dioxide was 0.65% (v / v), and the remainder was nitrogen. After the fermentation liquid settled for 45 minutes, the mass of calcium carbonate was 74.09 g / L, and the shape was irregular spherical, with most of the size distributed around 10 μm ( Figure 3 ).

[0058] Example 7 Traditional fermentation with 2.5 mol / L Ca(OH)2 as pH regulator

[0059] The inoculation and fermentation of the strain were carried out according to Example 6, except that the fermentation tank was stirred at a speed of 350 r / min. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 84.14 g / L ( Figure 1 ), with a production rate of 5.25 g / L·h, a hydrogen content of 20.12% (v / v), and a collected hydrogen yield of 14.06 mmol / L of fermentation broth. Carbon dioxide accounted for 0.85% (v / v), with the remainder being nitrogen. After 60 minutes of fermentation broth settling, the mass of calcium carbonate was 87.21 g / L, with an irregular spherical shape and a size distribution of approximately 5-10 μm.

[0060] Example 8 Traditional fermentation with 5 mol / L Ca(OH)2 and ammonia as pH regulator

[0061] The inoculation and fermentation of the strain were carried out according to Example 4, except that concentrated ammonia water (containing 25% ammonia) was used to adjust the initial pH to 7.0 before inoculation, 5M calcium hydroxide was used to adjust the pH at the beginning of fermentation, concentrated ammonia water was used to adjust the pH when the fermentation was carried out for 3 hours, and calcium hydroxide was used to adjust the pH when the amount of concentrated ammonia water added reached 6.8g / L fermentation liquid. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 81.80g / L, the production intensity was 5.11g / (L·h), the hydrogen content was 18.01% (v / v), the collected hydrogen yield was 14.15mmol / L fermentation liquid, carbon dioxide was 0.97% (v / v), and the rest was nitrogen. After the fermentation liquid settled for 30min, the mass of calcium carbonate was 78.20g / L. At the end of the fermentation, the electrical conductivity of the fermentation liquid was 19300μs / cm( Figure 2 ), which is 50% of ammonia coupled with sodium hydroxide as a pH adjuster. The reduction of salt concentration will help the subsequent separation of 1,3-PDO.

[0062] Example 9 Fermentation using a modified culture medium with 5 mol / L Ca(OH)2 and ammonia as pH regulators

[0063] The inoculation and fermentation of the strain were carried out according to Example 8, except that the seeds and the fermentation medium were modified medium. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 82.02 g / L ( Figure 1 ), the production intensity was 5.13 g / (L·h), the hydrogen content was 18.11% (v / v), the collected hydrogen yield was 14.27 mmol / L fermentation liquid, carbon dioxide was 0.99% (v / v), and the rest was nitrogen. After the fermentation liquid settled for 30 minutes, the mass of calcium carbonate was 80.15 g / L, the shape was irregular spherical, and the size distribution was about 2-20 μm ( Figure 4 ).

[0064] Example 10 Conventional fermentation with 1.5 mol / L Ca(OH)2 and ammonia as pH regulators

[0065] The inoculation and fermentation of the strain were carried out according to Example 8, except that the concentration of Ca(OH)2 was changed from 5 mol / L to 1.5 mol / L, the stirring speed of the fermenter was adjusted to 350 r / min, and the fermentation time was 19 h. The fermentation results are shown in Table 1. The final 1,3-propylene glycol concentration was 77.63 g / L ( Figure 1 ), the production intensity was 4.08g / (L·h), the hydrogen content was 20.17% (v / v), the collected hydrogen yield was 10.63mmol / L fermentation liquid, carbon dioxide was 0.77% (v / v), and the rest was nitrogen. After the fermentation liquid was centrifuged at 3000g for 10 minutes, the mass of the bacteria and calcium carbonate was 83.03g / L. The calcium carbonate was in the form of flakes, with a size distribution of 300-800nm ​​( Figure 5 ).

[0066] Example 11 Centrifugal Removal of Bacteria

[0067] The fermentation broths of Examples 1 and 4 were centrifuged to remove the bacteria respectively. The results are shown in Table 2. It can be seen that when the centrifugal force is 1000g, the fermentation broth is centrifuged for 5 minutes, and the clarification rate of the calcium hydroxide fermentation broth reaches 88.3%, while the clarification rate of the sodium hydroxide fermentation broth is only 55.3%; the centrifugation time is 10 minutes, and the clarification rate of the calcium hydroxide fermentation broth is 89.0%, while the clarification rate of the sodium hydroxide fermentation broth is only 76.8%. The centrifugation time is 15 minutes, and the clarification rate of the calcium hydroxide fermentation broth is 96.2%, while the clarification rate of the sodium hydroxide fermentation broth is only 89.0%. It can be seen from the table that the clarification rate of the calcium hydroxide fermentation broth is significantly higher than that of the sodium hydroxide during low-speed centrifugation. With a centrifugal force of 1000g and a centrifugation time of 15 minutes, the clarification rate of the fermentation broth can reach 96.2%. In industrial production, low-speed centrifugation bacteria separation can be achieved by a horizontal screw centrifuge. The fermentation broths of the two were further evaporated. A large amount of foam appeared at the beginning of the evaporation of the sodium hydroxide fermentation broth (Example 1) ( Figure 6 ), so that evaporation cannot continue. However, the calcium hydroxide fermentation broth (Example 4) has basically no foam and can be smoothly evaporated and concentrated to 7 times ( Figure 7 The protein and salt concentrations in the calcium hydroxide fermentation broth were significantly reduced, which greatly promoted the separation of 1,3-propylene glycol in the later stage and reduced the separation cost.

[0068] Table 1. Fermentation product formation under different operating conditions

[0069]

[0070] In Table 1, “-” means no calcium carbonate was produced; “ / ” means not measured.

[0071] Table 2. Comparison of low-speed centrifugation bacterial separation of fermentation broth in Examples 1 and 4

[0072]

[0073]

[0074] The above description of the embodiments is intended to facilitate understanding and application of the present invention by those skilled in the art. It will be apparent that those skilled in the art can readily make various modifications to these embodiments and apply the general principles described herein to other embodiments without requiring inventive effort. Therefore, the present invention is not limited to the embodiments described herein. Improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the present invention should be within the scope of protection of the present invention.

Claims

1. A method for co-producing 1,3-propylene glycol, calcium carbonate and hydrogen by microorganisms, comprising: The step of inoculating Clostridium butyricum with a deposit number of CGMCC NO: 17934 into a fermentation medium and performing anaerobic fermentation with glycerol as a substrate is characterized in that during the fermentation process, calcium hydroxide or ammonia water coupled with calcium hydroxide is used as a pH regulator to adjust the pH value of the fermentation liquid to 6-7; The fermentation liquid is subjected to static sedimentation and / or low-speed centrifugation to separate calcium carbonate and bacteria, and then concentrated by evaporation to 5 to 7 times to obtain 1,3-propylene glycol concentrate, and the fermentation tail gas is collected to obtain hydrogen; The concentration of the calcium hydroxide is 1.0-6.0 mol / L; The calcium carbonate is a micro-nanoscale calcium carbonate crystal, the crystal form is flake or spindle-shaped, and the size is 300nm-50μm; During the fermentation process, nitrogen was introduced 1 h before and after inoculation at 0.1 vvm.

2. The method according to claim 1, characterized in that When the pH regulator is ammonia water coupled with calcium hydroxide, an inorganic nitrogen source is added or not added to the fermentation medium, and during the fermentation process, ammonia water and calcium hydroxide with a concentration of 22-28% are alternately used to adjust the pH value of the fermentation liquid. Ammonia water is used to adjust the pH value of the fermentation liquid between 0 and 5 hours after the start of fermentation, and the amount of ammonia water added is 5-13.5 g / L of fermentation liquid.

3. The method according to claim 1, characterized in that The fermentation liquid is allowed to stand for 5 to 60 minutes for precipitation, and the precipitated calcium carbonate is separated. The fermentation liquid from which the calcium carbonate has been removed is centrifuged to separate the bacterial cells. Alternatively, the fermentation liquid is centrifuged at a low speed to obtain the bacterial cells or a mixture of the bacterial cells and calcium carbonate, wherein the low-speed centrifugation conditions are: a relative centrifugal force of 1000-3000 g and a centrifugation time of 3-30 minutes.

4. The method according to claim 1, wherein The hydrogen content in the fermentation tail gas is 10-40% (v / v).

5. The method according to claim 1, wherein The fermentation method adopts batch feeding fermentation. During the fermentation process, the stirring speed is 100-350r / min, the culture temperature is 30-38℃, and the pH is 6-7.

6. The method according to any one of claims 1 to 5, characterized in that: Among the components in the fermentation medium, the inorganic salt is replaced by sulfate instead of chloride.

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