A method for preparing a composite membrane material using fermented bacterial cells as raw materials

Abstract

The present invention belongs to the fields of fermentation engineering and chemical engineering, and specifically relates to the resource utilization of fermented bacteria after the end of fermentation to prepare membrane products. In the present invention, fermented bacteria are used to release macromolecules of the bacteria under the action of lytic enzymes or chemical lysis agents. The lysed bacterial solution contains components such as proteins, nucleic acids, chitosan, and peptidoglycan. A composite membrane is prepared by blending the macromolecules in the bacterial lysate with polyvinyl alcohol, realizing the product application of waste fermented bacteria. The composite membrane prepared by the present invention uses the bacterial lysate, which will increase the toughness and elasticity, improve the hydrophilic and moisture permeability of the composite membrane, and can release nutrients such as proteins for green plants to absorb during the heat preservation process of green plants.

Description

Technical Field:

[0001] The present invention belongs to the fields of fermentation engineering and chemical engineering, and particularly relates to a membrane product prepared by resource utilization of fermented bacteria after the end of fermentation production. Background Art:

[0002] Taking the fermentation of Corynebacterium glutamicum to produce glutamic acid and refined monosodium glutamate as an example, calculated according to the fact that 200 kg of dry bacteria can be extracted from the fermentation broth for every 1 ton of monosodium glutamate produced, the annual production of bacterial protein in the whole country is about 22×10 4 tons (Zhou Dawei, Xiao Dongguang, Guo Xuewu, Lv Hongyan. Food Research and Development, 2012). And more than 10,000 tons of L-tryptophan are produced by Escherichia coli in China every year. The production of 1 ton of L-tryptophan is accompanied by the production of 1.3 - 1.5 tons of waste bacterial cells (WBC) (Qingyang Xu, Fang Bai, Ning Chen, and Gang Bai, Bioengineered, 2019). When lactic acid is fermented by Rhizopus oryzae, 84.5 kg of Rhizopus oryzae cells are produced for every 1 ton of lactic acid obtained (Yang Lei, Li Xin, Yu Shiyuan. Chemistry and Industry of Forest Products, 2015).

[0003] The existing fermentation bacterial treatment processes include preparing a bacterial protein powder with comprehensive nutritional components from waste bacterial protein to replace yeast extract and reduce the cost of the fermentation medium (Chinese Patent Application CN 106086093A; Chinese Patent Application CN 110092528 A); preparing peptide products and feed products with higher purity to avoid waste of bacterial raw materials (Chinese Patent Application CN107011409A; Chinese Patent Application CN107418897A); preparing liquid organic fertilizer from waste bacteria in amino acid fermentation (Chinese Patent Application CN106116773A), and producing bacterial protein and polypeptide organic fertilizer from waste in vitamin C fermentation (Chinese Patent Application CN111995467A).

[0004] Polyvinyl alcohol (PVA) is a synthetic, non-ionic water-soluble, biodegradable and biocompatible linear macromolecular polymer with good film-forming ability. Polyvinyl alcohol membranes have excellent flexibility, transparency, non-toxicity and barrier properties, and have been widely used in the packaging industry (He Binbin, Yu Huirong, Zhang Li, Zhao Zhenghe, Ran Ruimin, Chen Saiyan. Plastics Industry. 2022). Blending the bacterial lysate with PVA to prepare a composite membrane can not only reduce costs, but also improve the comprehensive performance of the film. The composite membrane product prepared by the present invention can be applied to the directions of anti-fouling and self-cleaning or green plant heat preservation, and can also provide nutrients such as protein for green plants.

[0005] The existing treatment methods for fermented bacteria usually have problems such as high treatment costs and great environmental protection pressure, and most of the fermented bacteria have not been "turned waste into treasure". The present invention fully utilizes the bacteria and nutrients such as proteins released after lysis, and blends them with biodegradable polymers such as polyvinyl alcohol to prepare new composite membrane materials applicable to fields such as anti-fouling and self-cleaning, green plant heat preservation, and fertilizer slow release. Compared with the film prepared by using polyvinyl alcohol alone, the bacteria lysate will increase the toughness and elasticity of the composite membrane, improve the hydrophilicity and moisture permeability of the composite membrane, and can release nutrients such as proteins for green plants to absorb during the process of heat preservation for green plants. Summary of the Invention:

[0006] The object of the present invention is to release bacterial macromolecules under the action of lytic enzymes or chemical lysing agents from fermented bacteria. The lysed bacterial liquid contains components such as proteins, nucleic acids, and peptidoglycans. A composite membrane is prepared by blending the macromolecules in the bacterial lysate with polyvinyl alcohol, realizing the product application of waste fermented bacteria.

[0007] One of the technical solutions provided by the present invention is a method for preparing a membrane material using waste bacteria, including the following steps:

[0008] A method for preparing a membrane material using fermented bacteria as raw materials, including the following steps:

[0009] (1) Before or after extracting the target product from the microbial fermentation broth, collect the bacteria.

[0010] Furthermore, add a flocculant to the fermentation broth to collect the bacteria.

[0011] Furthermore, the flocculant is chitosan, and the addition amount is 0.01% - 1% (w / v) of the fermentation broth.

[0012] Furthermore, the flocculation conditions are: stirring speed 100 - 300 r / min, flocculation temperature 45 - 85 °C, stir for 5 - 20 min and then adjust the pH to 6 - 10.

[0013] Even further, the chitosan addition amount is 0.06% - 0.3%, and after reacting for 5 - 20 min, adjust the pH to 7 - 8.5 with calcium hydroxide.

[0014] Furthermore, collect the bacteria by plate and frame filtration, membrane filtration or centrifugation.

[0015] Furthermore, the fermentation broth is a lactic acid fermentation broth using Escherichia coli as the fermentation strain.

[0016] (2) After washing the collected bacteria, prepare a bacterial lysate by treating with lytic enzymes and / or chemical lysing agents.

[0017] Furthermore, the cell disruption rate in the cell lysate is 62% - 99.5%, and the protein content in the lysate ranges from 37 to 239 mg / g;

[0018] Furthermore, the collected cells are washed twice and resuspended with water at a ratio of 1:4 (w / v), and then lysozyme and / or chemical lysing agent are added;

[0019] Even further, HCl is added during each washing of the cells to adjust the pH of the cell suspension to 5.0 - 5.5;

[0020] Furthermore, the lysozyme is at least one of lysozyme, snailase, cellulase or lysostaphin;

[0021] Furthermore, the chemical lysing agent includes but is not limited to NaOH, KOH, SDS, Triton X - 100, etc.;

[0022] Furthermore, the lysis conditions are: 20°C - 100°C, for 10 - 40 min;

[0023] Even further, it is lysed at 50°C - 100°C, pH 7 - 14 for 15 - 30 min;

[0024] Furthermore, after washing and resuspending the collected cells, sodium hydroxide with a final concentration of 0.5 - 3 M and SDS with a concentration of 0.25 - 1.0% (w / v) are added as the lysing agent;

[0025] Even further, after washing and resuspending the collected cells, sodium hydroxide with a final concentration of 0.5 - 1.5 M and SDS with a concentration of 0.25 - 1% (w / v) are added as the lysing agent;

[0026] (3) The cell lysate is blended with polyvinyl alcohol and a plasticizer and a cross - linker are added to prepare a blend solution. The blend solution is spread on a PVC plate, and the solution is spread evenly with a film applicator and dried to form a film;

[0027] Furthermore, the concentration of the polyvinyl alcohol is 5% - 20% (w / v);

[0028] Furthermore, the proportion of the cell lysate in the mixture with the polyvinyl alcohol solution is 10% - 90% (w / w);

[0029] Even further, the proportion of the cell lysate in the mixture with the polyvinyl alcohol solution is 40% - 70%.

[0030] Furthermore, it is dried to form a film in an oven at 60°C;

[0031] Furthermore, the plasticizer is glycerol, sorbitol or propylene glycol, etc.;

[0032] Preferably, the plasticizer is glycerol with a final concentration of 1% - 15% (w / w);

[0033] Furthermore, the crosslinking agent is citric acid, polyethylene glycol, ammonium persulfate, etc.;

[0034] Preferably, the crosslinking agent is citric acid with a final concentration of 0.1% - 12% (w / w).

[0035] The second technical solution provided by the present invention is the membrane material prepared by the above method.

[0036] The third technical solution provided by the present invention is the application of the above membrane material, especially in the fields of anti-fouling and self-cleaning, green plant heat preservation, and fertilizer slow release. And during the green plant heat preservation process, it can release nutrients such as proteins for the green plants to absorb.

[0037] The beneficial effects achieved by the present invention:

[0038] The present invention can efficiently collect the bacteria in the fermentation broth, moderately lyse them and use them to prepare a composite membrane with PVA, realizing the synchronous production of the fermentation product and the composite membrane derived from the fermentation bacteria, and completely solving the formation of fermentation bacterial waste in the existing industrial production system. Thereby, it can also significantly improve the economic efficiency and environmental benefits of fermentation production and significantly reduce the comprehensive production cost.

[0039] The present invention can also be applied to the waste bacteria generated in the production of other organic acids such as citric acid, malic acid, succinic acid, etc., or amino acids such as lysine, glutamic acid, threonine, alanine, etc.

[0040] When collecting bacteria at the end of lactic acid fermentation in the present invention, using chitosan as a biological flocculant and maintaining a certain temperature and pH, the flocculant will be completely collected together with the bacteria without affecting the subsequent separation and purification of lactic acid. And the collected bacteria can be simply lysed and then blended with the polyvinyl alcohol solution to obtain a composite membrane with better hydrophilic and moisture permeability properties.

[0041] The membrane product prepared by the present invention can be applied to flower potted plants, anti-fouling and self-cleaning, etc.

[0042] The present invention fully utilizes the bacteria and macromolecular nutrients such as proteins released after lysis, and crosslinks them with biodegradable polymers such as polyvinyl alcohol to prepare a new composite membrane material suitable for the fields of anti-fouling and self-cleaning, green plant heat preservation, and fertilizer slow release. Compared with the film prepared by using polyvinyl alcohol alone, the bacterial lysate will increase the toughness and elasticity of the composite membrane, improve the hydrophilic and moisture permeability properties of the composite membrane, and during the green plant heat preservation process, it can release nutrients such as proteins for the green plants to absorb. Description of the drawings:

[0043] To make the content of the present invention easier to be clearly understood, the following further details the present invention according to specific embodiments of the present invention and in conjunction with the accompanying drawings, wherein:

[0044] Figure 1 The finished film material prepared by the present invention.

[0045] Figure 2 Infrared spectrum analysis of the thallus / PVA composite film.

[0046] Figure 3 The water contact angle of the thallus / PVA composite film. Specific embodiments:

[0047] To make the objectives, technical solutions and advantages of this patent clearer and more understandable, the following further details this patent in conjunction with specific embodiments. It should be understood that the specific embodiments described herein are only used to explain this patent and are not used to limit the present invention.

[0048] Taking the fermentation broth produced by the calcium salt method for lactic acid as an example, after the lactic acid fermentation is completed, the fermentation broth is heated to 45°C to 85°C, the biological flocculant chitosan is added and stirred to maintain the reaction for 5 - 20 minutes, and then solid-liquid separation is carried out by filtration. The solid part collected is the thallus, and the liquid part collected is the free lactic acid solution containing lactic acid monomers. The obtained thallus can be used for the subsequent preparation of the film material. The obtained free lactic acid solution is filtered, concentrated, etc., and is the crude lactic acid product, which can be used for subsequent refining such as nanofiltration, decolorization, ion exchange, etc. to obtain high-purity lactic acid monomers.

[0049] The main experimental methods adopted in the present invention are as follows:

[0050] (1) Preparation of Fermentation Medium - Collection of Bacterial Cells (Flocculation). The preparation of calcium salt method lactic acid fermentation broth was carried out according to the method of the invention patent (Wang Zhengxiang et al., ZL201580000781.7). The fermentation strains were CGMCC 11059 or CGMCC11060, among which the strain CGMCC 11059 was used for the fermentation production of D-lactic acid, and the strain CGMCC11060 was used for the fermentation production of L-lactic acid (Wang Zhengxiang et al., ZL201580000781.7). At the initial stage of fermentation, glucose was added to the basic fermentation medium to a final concentration of 10 - 50 g / L, and the cultivation was carried out at 30°C - 37°C, pH 5.5 - 7.5, ventilation of 0.1 - 2.0 vvm, and stirring at 100 - 1000 r / min; the cultivation time was 5 - 15 h, and the cell mass reached 10 - 50 OD; the ventilation was turned off, the stirring speed was reduced to 0 - 300 r / min, the fermentation temperature was increased to 37°C - 50°C, a glucose solution with a final concentration of 16% - 25% was supplemented, the flow rate was controlled at 3 g / (L h) - 25 g / (Lh), and 5% - 35% calcium hydroxide was simultaneously added dropwise to control the fermentation pH between 5.0 and 8.0. Flocculation was carried out by adding 0.1 - 10 g / L of the biological flocculant chitosan to the fermentation broth, with a stirring time of 5 - 20 min, a stirring speed of 100 - 300 r / min, and a flocculation temperature of 45 - 85°C. The pH of the fermentation broth was adjusted to 7.0 - 8.5 with calcium hydroxide, and the bacterial cells were collected by filtration.

[0051] (2) Lysis of Bacterial Cells. Escherichia coli was lysed using NaOH solution and SDS solution, with the final concentration of NaOH being 0.5 - 3 M, the final concentration of SDS being 0.25 - 1.5% (w / v), the temperature being 20°C - 100°C, pH 7 - 14, and the time being 10 - 40 min.

[0052] (3) Preparation of Blend Solution. The lysed bacterial cell lysate obtained above was mixed with PVA solution to prepare a blend solution. The concentration of the PVA solution was 5% - 20%, and the mixing ratio was 1:9 - 9:1; during the mixing process, glycerol with a final concentration of 1 - 15% was added as a plasticizer, and citric acid with a final concentration of 0.1 - 12% was added as a crosslinking agent.

[0053] (4) Preparation of Finished Product (Membrane Material). The lysed bacterial cell lysate was mixed with PVA solution according to the above method, and the mixed blend solution was spread evenly on a PVC plate, and the solution was spread out with a film applicator and dried into a film in an oven at 60°C.

[0054] The present invention will be further explained and illustrated through specific examples below.

[0055] Example 1 A Method for Fermentation of D-Lactic Acid

[0056] The glycerol stock of the D-lactic acid-producing strain CGMCC NO. 11059 was inoculated into 50 mL of LB liquid medium and cultured in a shaker at 37 °C and 200 r / min for 12 h to obtain the primary seed culture. The primary seed culture was inoculated into 5 L of M9 liquid medium with glucose as the carbon source, with an initial sugar concentration of 0.5%, and cultured in a shaker at 37 °C and 200 r / min for 10 h to obtain the secondary seed culture. The secondary seed culture was inoculated into a fermenter containing M9 liquid medium at an inoculation amount with an initial OD value of 0.3. After inoculation, 5 m 3 The initial volume of the fermenter was 2.5 m 3 , the initial addition amount of the converted syrup was 3%, and the fermentation production of lactic acid monomer was started. The initial fermentation temperature was controlled at 37 °C, ammonia water was used to maintain the pH at 6.5, the aeration rate was adjusted to 1.5 vvm during the cell growth process, and the stirring speed was 600 r / min. When the cell concentration reached OD 600 30, the ventilation was turned off, the fermentation temperature was controlled at 40 °C, the stirring speed was adjusted to 200 r / min, 25% calcium hydroxide suspension was added dropwise to maintain the pH at 7.0, a total of 6.0 kg of glucose was added, and the fermentation was ended when the residual sugar concentration was lower than 0.5 g / L.

[0057] Composition of M9 medium: 0.5 g / L of NaCl, 1 g / L of NH4Cl, 3 g / L of KH2PO4, 15.1 g / L of Na2HPO4·12H2O, and the rest was water.

[0058] Using the obtained lactic acid fermentation broth as the raw material, it was heated to 60 °C in a reaction kettle and maintained at this temperature. 2 g / L of chitosan solution was added under stirring at 100 r / min. After all the chitosan solution was added, the reaction was maintained for 15 min, and then 20% calcium hydroxide was added to adjust the pH of the fermentation broth to 7.5; after the reaction ended, the solid-liquid separation of the reaction solution was carried out using a plate and frame filter device with a filtration pore size of 6 μm, and the solid part collected was the cells.

[0059] Example 2 Removal of interfering factors - Lactic acid and miscellaneous acids

[0060] The main fermentation product lactic acid and by-products such as formic acid, acetic acid, pyruvic acid, and succinic acid are all soluble in water and will remain in the cells. The method of resuspending and washing the cells can be used to remove various acids in the cells.

[0061] The cells obtained in Example 1 and deionized water were resuspended after washing 2 times at a ratio of 1:4 (w / v). When washing, 1 M HCl was added to adjust the pH of the system to 5.0 (this was the experimental group; control group 1 was only resuspended without washing and without adding HCl to adjust the pH; control group 2 was resuspended after washing with water 2 times without adding HCl to adjust the pH), and treated at 2 M NaOH, 0.5% SDS, and 90 °C for 25 min to obtain the cell lysate.

[0062] The obtained cell lysate was miscible with 10% PVA solution at a ratio of 1:1 for 60 min, and glycerol with a final concentration of 4% was added as a plasticizer and citric acid with 3% as a crosslinking agent. The miscible blend solution was spread on a PVC plate, and the solution was spread evenly with a film applicator and dried into a film in an oven at 60 °C. Observe the morphology and appearance of the blend solution and the composite film.

[0063] Table 1

[0064]

[0065] Example 3: Film formation of cell lysate under different conditions

[0066] The cells obtained in Example 1 and water were resuspended at a ratio of 1:4 (w / v) after washing twice (adjust the pH of the system to 5.0 with 1 M HCl during washing), and a certain final concentration of NaOH and SDS solutions were added for cell wall breaking treatment at a certain temperature for a certain time. The components of each lysate in the lysate after lysis are shown in the following table. And the cell lysate was miscible with 10% PVA solution at a ratio of 5:1 for 60 min, and glycerol with a final concentration of 4% was added as a plasticizer and citric acid with 3% as a crosslinking agent. The miscible blend solution was spread on a PVC plate, and the solution was spread evenly with a film applicator and dried in an oven at 60 °C.

[0067] Table 2 Data corresponding to different lysis conditions

[0068]

[0069] Through the above experiments, it was determined that the cell breakage rate and protein concentration are important factors affecting film formation. On the basis of the above experiments, the inventor verified through a large number of experiments and finally determined that the cell breakage rate in the cell lysate is 62% - 99.5%, and at the same time, the protein content range in the lysate is 37 - 239 mg / g, which are the better conditions for finally forming the composite film.

[0070] Example 4: Preparation of composite film by mixing cells and PVA

[0071] The cells in Example 1 and water were resuspended at a ratio of 1:4 (w / v) after washing twice (adjust the pH of the system to 5.0 with 1 M HCl during washing), and lysed under the conditions of 0.5 M NaOH, 0.75% SDS, 30 °C, and 25 min. The obtained cell lysate was miscible with 5% PVA solution at a ratio of 1:9 for 60 min, and glycerol with a final concentration of 1% was added as a plasticizer and citric acid with 0.1% as a crosslinking agent. The miscible blend solution was spread on a PVC plate, and the solution was spread evenly with a film applicator and dried into a film in an oven at 60 °C.

[0072] Example 5: Preparation of Composite Membrane by Mixing Bacterial Cells with PVA

[0073] The bacterial cells in Example 1 and water were resuspended at a ratio of 1:4 (w / v) after washing twice (the pH of the system was adjusted to 5.0 with 1M HCl during washing). The bacterial cell lysate obtained by lysing at 50°C for 10 min under the conditions of 1M NaOH, 0.75% SDS was mixed with 20% PVA solution at a ratio of 9:1. The mixing time was 60 min, and glycerol with a final concentration of 15% was added as a plasticizer, and citric acid with a final concentration of 12% was added as a crosslinking agent. The mixed blend solution was spread on a PVC plate, and the solution was evenly spread with a film applicator and dried into a film in an oven at 60°C.

[0074] Example 6: Preparation of Composite Membrane by Mixing Bacterial Cells with PVA

[0075] The bacterial cells in Example 1 and water were resuspended at a ratio of 1:4 (w / v) after washing twice (the pH of the system was adjusted to 5.0 with 1M HCl during washing). The bacterial cell lysate obtained by lysing at 90°C for 25 min under the conditions of 2M NaOH, 0.5% SDS was mixed with 10% PVA solution at a ratio of 1:1. The mixing time was 60 min, and glycerol with a final concentration of 4% was added as a plasticizer, and citric acid with a final concentration of 3% was added as a crosslinking agent. The mixed blend solution was spread on a PVC plate, and the solution was evenly spread with a film applicator and dried into a film in an oven at 60°C.

[0076] Example 7: Preparation of Composite Membrane by Mixing Bacterial Cells with PVA

[0077] The bacterial cell lysate in Example 6 was mixed with 10% PVA solution at a ratio of 1:1. The mixing time was 60 min, and no plasticizer or crosslinking agent was added. The mixed blend solution was spread on a PVC plate, and the solution was evenly spread with a film applicator and dried in an oven at 60°C. Finally, due to the lack of plasticizer and crosslinking agent, a composite membrane could not be formed.

[0078] Example 8: Mechanical Property Parameters of the Composite Membrane of the Present Invention

[0079] For the membrane products obtained in the examples of the present invention, the corresponding mechanical property data are as follows in the table:

[0080] Table 3 Mechanical Property Parameters of the Composite Membrane

[0081]

[0082] Compared with the pure PVA membrane, the tensile property and rigidity of the composite membrane of the present invention decrease, but the elongation at break increases significantly. The longer the length that can be extended after being stressed, the better the toughness, and it is not easy to break when applied as a coating material.

[0083] Preparation of PVA film: Weigh 10 g of polyvinyl alcohol in a 250 mL container, add deionized water, and stir to dissolve it in a water bath at 90 °C. After complete dissolution, add deionized water to make the volume up to 100 mL to obtain a 10% PVA (w / v) solution. Spread it evenly on a PVC plate, use a film applicator to spread the solution evenly, and dry it into a film in an oven at 60 °C.

[0084] Preparation of PVA + bovine serum albumin composite film: Weigh 1.56 g of bovine serum albumin in a 25 mL container, add 8.44 g of deionized water (final concentration of bovine serum albumin is 156 mg / g), and stir with a magnetic stirrer at 90 °C until the bovine serum albumin is dissolved into a transparent and clear solution. Mix it with the 10% PVA solution in a 1:1 ratio, with a mixing time of 60 min, and add glycerol with a final concentration of 4% as a plasticizer and citric acid with a final concentration of 3% as a crosslinking agent. Spread the mixed blend solution evenly on a PVC plate, use a film applicator to spread the solution evenly, and dry it into a film in an oven at 60 °C.

[0085] Preparation of PVA + bovine serum albumin / chitosan composite film: Weigh 8.44 g of a 0.25% chitosan solution in a 25 mL container, add 1.56 g of bovine serum albumin (final concentration of bovine serum albumin is 156 mg / g) to it, and stir with a magnetic stirrer at 90 °C until it is dissolved into a transparent and clear solution. Mix it with the 10% PVA solution in a 1:1 ratio, with a mixing time of 60 min, and add glycerol with a final concentration of 4% as a plasticizer and citric acid with a final concentration of 3% as a crosslinking agent. Spread the mixed blend solution evenly on a PVC plate, use a film applicator to spread the solution evenly, and dry it into a film in an oven at 60 °C.

[0086] Example 9: Infrared spectrum analysis of the composite film of the present invention

[0087] Mix the cell lysate obtained in Example 6 with the 10% PVA solution in different ratios, with a mixing time of 60 min, and add glycerol with a final concentration of 4% as a plasticizer and citric acid with a final concentration of 3% as a crosslinking agent. Spread the mixed blend solution evenly on a PVC plate, use a film applicator to spread the solution evenly, and dry it into a film in an oven at 60 °C. Perform Fourier transform infrared spectroscopy analysis, and the results are as Figure 2 shown.

[0088] PVA has a strong absorption peak at 3418 cm -1 , which proves the existence of a large number of hydroxyl groups in PVA. The absorption peak at 2930 cm -1 should be caused by the absorption vibration of C-H. 1728 cm -1 is the characteristic stretching vibration peak of C=O, 1438 cm -1 is the bending vibration of O-H, and 1248 cm -1For N-H bending vibration and C-N stretching vibration, the peak of amide III band, and the peak at 1092 cm -1 The peak at is the C-O stretching vibration in PVA, and the peak at 1036 cm -1 is the stretching vibration of S-O, and the peak at 845 cm -1 is the C-C stretching vibration of PVA (Posati S, Tamara R, Giuri C, et al. European Polymer Journal. 2018). Compared with the PVA film, the composite film has an additional peak at 1590 cm -1 , and the other peaks are the same as those of PVA. The peak at 1590 cm -1 is the amide II band, representing the presence of protein. Therefore, it can be seen that during the preparation of the composite film, cross-linking is formed between the cell lysate and PVA, resulting in a new strong absorption peak of the amide II band in the composite film on the basis of the original PVA film.

[0089] Example 10: Moisture permeability and water contact angle test of the composite film of the present invention

[0090] The water vapor transmission coefficient can characterize the moisture permeability of the film. The pure PVA film prepared in Example 8, the PVA + bovine serum albumin composite film, and the composite film used in Example 9 were used as test samples.

[0091] 1. Detect the moisture permeability through a water vapor transmission rate tester.

[0092] The results are shown in Table 4. The water vapor transmission coefficient of the pure PVA film is 0.41×10 -4 . After blending PVA with the cells, its water vapor transmission coefficient increases. For the blended film with 60% of the cell lysate, the water vapor transmission coefficient increases to 0.96×10 -4 , indicating an improvement in its moisture permeability.

[0093] Moreover, compared with the composite film prepared from pure protein and PVA, the composite film prepared from the cell lysate and PVA has a higher water vapor transmission coefficient, which increases from 0.54×10 -4 to 0.97×10 -4 . This shows that the composite film prepared by replacing pure protein with the cell lysate is more suitable for applications with higher moisture permeability requirements.

[0094] Table 4 Water vapor transmission coefficient of the cell / PVA composite film

[0095]

[0096] 2. Use a JY-159 type contact angle measuring instrument to measure the static water contact angle of the sample.

[0097] Figure 3is the water contact angle of the cell / PVA composite film. As can be seen from the figure, the water contact angles of the pure PVA film, PVA + bovine serum albumin, PVA + bovine serum albumin / chitosan, and the blend films with the proportion of cell lysate ranging from 10% to 60% are 78.05°, 56.35°, 56.14°, 64.45°, 58.70°, 57.75°, 54.37°, 49.5°, and 49.3° respectively. Since a large number of exposed hydroxyl groups in the PVA molecular chain exhibit strong hydrophilicity, the PVA film presents as a hydrophilic film. The cell / PVA blend film is also hydrophilic, and from the perspective of the water contact angle, the hydrophilicity of the blend film is better than that of the pure PVA film.

[0098] As a protective film for potted flower transplantation, when the flower is transplanted into the flower pot, the better the moisture permeability and hydrophilicity are, the more conducive it is to ensuring the dissolution and transportation of water molecules. Due to its good hydrophilic property, when watering the flower, the composite film combines with water molecules, absorbs water and swells and then falls off, and the nutrients in the composite film are released for the flower to absorb and grow.

[0099] Example 11: Application of the composite film of the present invention

[0100] When the composite film of the present invention is applied during the transplantation of potted flowers, it can be used as a soil protective film. When the flower is transplanted into the pot, when watering, the composite film absorbs water and falls off while releasing nutrients such as proteins for the flower to absorb.

[0101] The composite films prepared in Examples 4, 5, and 6 of the present invention, the PVA film prepared in Example 8, the PVA + bovine serum albumin, and the PVA + bovine serum albumin + chitosan composite film are applied to potted flowers as follows:

[0102] Wrap the composite film around the outer layer of the soil of the flower. Before the flower is transplanted into the flower pot, the composite film provides heat preservation and protection performance. After the flower is transplanted into the flower pot, when watering the flower, the composite film will quickly absorb water and fall off while releasing nutrients such as proteins in the cell lysate for the flower to absorb, improving the survival rate of the flower.

[0103] Table 5 shows the number of flowers that can be wrapped per square meter of the film during the wrapping process of the composite film, and the survival rate of jasmine potted plants is observed after one month of cultivation with the PVA film, PVA + bovine serum albumin, PVA + bovine serum albumin + chitosan composite film, and the composite film of the present invention wrapped around the jasmine potted soil respectively. During this period, the potted plants are watered once a day, and no composite film is added as a control:

[0104] Table 5

[0105]

[0106] The results show that the composite film prepared from the bacterial cell lysate of the present invention has better ductility due to its better elongation at break, so more flowers can be wrapped per square meter of the film. Moreover, since the film contains richer nutrients, it is more conducive to the survival of flowers and improves the survival rate.

[0107] The composite film of the present invention can also be used in the field of anti-fouling and self-cleaning. When the composite film prepared by the present invention is coated on the surface of a glass curtain wall, a window, a building decoration material or any object that needs anti-fouling, water spreads fully on the film surface to form a uniform water film. On the one hand, it can eliminate the diffuse reflection caused by water droplets and achieve an anti-fog effect; on the other hand, during the cleaning process, when water spreads fully, it completely wets the coating and pollutants, so that the dust and dirt on the object surface float up and slide down under the action of stress and gravity, realizing the self-cleaning function.

[0108] The above are only the preferred embodiments of the present invention, and all equivalent changes and modifications made according to the scope of the patent application of the present invention shall fall within the scope covered by the present invention.

Claims

1. A method for preparing a membrane material using fermented bacterial cells as a raw material, characterized in that, It includes the following steps: (1) Before or after extracting the target product from the microbial fermentation broth, chitosan is added as a flocculant to collect the bacterial cells; (2) After washing the collected bacterial cells, the cell lysate is prepared by treatment with lytic enzymes and / or chemical lysing agents; (3) The cell lysate is blended with polyvinyl alcohol and a plasticizer and a crosslinking agent are added to prepare a blended solution. The blended solution is spread on a PVC plate, and the solution is spread evenly with a film applicator and dried to form a film; In step (2), the collected bacterial cells are washed with water and resuspended, and then lytic enzymes and / or chemical lysing agents are added; when washing the bacterial cells each time, HCl is added to adjust the pH of the bacterial suspension to 5.0 - 5.5; In the cell lysate described in step (2), the cell disruption rate is 62% - 99.5%, and at the same time, the protein content in the lysate ranges from 37 to 239 mg / g.

2. The method for preparing a membrane material using fermented bacterial cells as a raw material according to claim 1, characterized in that, The flocculation conditions in step (1) are: the stirring speed is 100 - 300 r / min, the flocculation temperature is 45 - 85°C, after stirring for 5 - 20 min, the pH is adjusted to 6 - 10; the addition amount of the flocculant chitosan is 0.01% - 1% w / v of the fermentation broth.

3. The method for preparing a membrane material from fermented bacterial cells as claimed in claim 1, wherein, In step (2), the lytic enzyme is at least one of lysozyme, snail enzyme, cellulase or lysing enzyme; the chemical lysing agents include NaOH, KOH, SDS, Triton X - 100.

4. The method for preparing a membrane material using fermented bacterial cells as raw materials according to claim 1, characterized in that, In step (2), the lysis conditions are: after washing and resuspending the bacterial cells, sodium hydroxide with a final concentration of 0.5 - 3 M and SDS with a concentration of 0.25 - 1.0% w / v are added as lysis agents; at 20°C - 100°C, lysis is carried out for 10 - 40 min.

5. A method for preparing a membrane material from fermented bacteria cells as claimed in claim 1, characterized in that, In step (3), the concentration of polyvinyl alcohol is 5% - 20% w / v; the proportion of the cell lysate in the mixture with the polyvinyl alcohol solution is 10% - 90% w / w.

6. The method for preparing a membrane material using fermented bacteria cells as raw materials according to claim 1, characterized in that, In step (3), the plasticizer is glycerol, sorbitol or propylene glycol; the crosslinking agent is citric acid, polyethylene glycol or ammonium persulfate.

7. The membrane material prepared by the method according to any one of claims 1 - 6.

8. The application of the membrane material according to claim 7 in the fields of anti - fouling and self - cleaning, green plant heat preservation, and fertilizer slow - release.

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