Dry microorganism culture medium as well as preparation method and application thereof

Abstract

The invention provides a dry microorganism culture medium as well as a preparation method and application thereof. The method comprises the following steps: dissolving 7.5-15 parts of a nutritional agent, 2-3 parts of glycerol, 10-15 parts of gel and 0.75-1.2 parts of potassium chloride in deionized water to obtain a mixed solution; the pH value of the mixed solution is adjusted to 7.0-7.2, the mixed solution is poured into a flat plate after high-temperature sterilization, the flat plate is placed at the temperature of 0-20 DEG C for gelatinization treatment for 30-60 min, and a gel product is obtained; and drying the gel product under the sterile condition of 30-40 DEG C for 6-12 hours to obtain the dried microbial culture medium. According to the method, normal-temperature gelatinization treatment is performed firstly, and then drying is performed under the condition of being lower than the sol temperature, so that the gel property and rehydration capacity of the dried culture medium are effectively improved, dependence of a traditional culture medium on expensive materials such as agar is reduced, and the method has the characteristics of simple preparation process, low cost and excellent effect, is suitable for large-scale production, transportation and storage, and has wide application prospects. Wide application prospects are realized.

Description

Technical Field

[0001] The invention relates to the field of microbial culture, and in particular to a dry microbial culture medium and a preparation method and application thereof. Background Art

[0002] With the development of microbiology and biotechnology, the application of microbial culture media has become more and more extensive, covering many fields such as food, medicine, and environmental monitoring. The quality of microbial culture media directly affects the growth and metabolic process of microorganisms, but its preparation process is cumbersome and difficult to transport and store, which makes it difficult to control the quality of microbial culture media. Therefore, it is particularly important to develop culture media that are efficient, economical, and easy to transport and store.

[0003] At present, traditional microbial culture media are usually prepared on-site using ingredients such as agar and tryptone through multiple steps, which has technical problems such as time-consuming preparation and difficult quality control. Although traditional disposable culture media solve the problems of time-consuming preparation and poor quality control, they have a short storage period and are easily damaged during transportation. Therefore, it is necessary to develop a convenient, efficient and long-term stable quality microbial culture medium preparation method.

[0004] Agar is the most commonly used gelling agent when preparing culture medium. It can be prepared into disposable dry culture medium after being heated, dissolved and dried, but its rehydration ability is poor and it is impossible to provide sufficient water for microbial culture. In addition, the practicality of disposable dry culture medium is not only related to the preparation material, but also to the conditions of drying and rehydration, and there are certain technical difficulties in the preparation process. Therefore, it is of great practical significance to develop a dry microbial culture medium that is efficient, convenient for storage and transportation, and has good rehydration ability. To this end, the present invention provides an innovative preparation method, which overcomes the defects in the prior art by proposing a new culture medium preparation method and optimizing the formula and process, and meets the growing market demand for culture medium. Summary of the invention

[0005] In order to solve the problems mentioned in the background technology, the present application provides a dry microbial culture medium and a preparation method and application thereof.

[0006] In a first aspect of the present invention, a method for preparing a dry microbial culture medium is provided, the method comprising:

[0007] S1, dissolving 7.5-15 parts of a nutrient, 2-3 parts of glycerin, 10-15 parts of a gelling agent, and 0.75-1.2 parts of potassium chloride in 500-1500 parts of deionized water to obtain a mixed solution;

[0008] S2, adjusting the pH value of the mixed solution to 7.0-7.2, sterilizing it at high temperature, pouring it into a plate, placing it at 0-20°C for gelation treatment for 30-60 minutes, and obtaining a gel product;

[0009] S3, drying the gel product under sterile conditions at 30-40° C. and a relative humidity of 15-50% for 6-12 hours to obtain a dry microbial culture medium.

[0010] The above method effectively improves the gelation and rehydration capacity of the culture medium by first performing a gelation treatment at room temperature and then drying it below the sol temperature. It not only reduces the dependence of traditional culture media on expensive materials such as agar, but also has the characteristics of simple preparation process, low cost and excellent effect. It is suitable for large-scale production, transportation and storage, and has broad application prospects.

[0011] Furthermore, the nutrient agent includes one or more of 5-10 parts of tryptone, bovine bone peptone and fish peptone, and one or more of 2.5-5 parts of yeast extract powder and beef extract powder. The combination of multiple nutrient agents improves the nutritional value of the culture medium. It not only provides a rich nitrogen source and vitamins for the microorganisms to promote their growth and reproduction, but also can be flexibly prepared according to the needs of different microorganisms, thereby enhancing the applicability of the culture medium.

[0012] Furthermore, the gelling agent comprises 60-100% carrageenan, 0-40% agar and 0-40% konjac flour. The appropriate gelling agent ratio can effectively improve the rehydration capacity of the dry culture medium, and at the same time ensure that the culture medium has good water retention and gel strength after rehydration.

[0013] Furthermore, after adjusting the pH value of the mixed solution to 7.0-7.2 in step S2, the mixed solution is placed at 100-150°C and sterilized for 15 minutes. A reasonable pH value ensures the growth environment of microorganisms and helps maintain the stability of the culture medium components. High temperature sterilization can effectively remove miscellaneous bacteria, ensure the sterility of the culture medium, and improve the reliability of microbial experiments.

[0014] Furthermore, in step S1, a supplement is added, and the supplement includes one or more of a nitrogen source, an organic acid, a carbon source, a salt containing metal ions, and a growth factor. The addition of the supplement can optimize the growth environment of the microorganisms and enhance the functionality of the culture medium. By providing a variety of nutrients to meet the growth requirements of different microorganisms, the applicability and flexibility of the culture medium can be improved.

[0015] Furthermore, the nitrogen source includes one or more of amino acids, ammonium sulfate and zinc sulfate, the carbon source includes one or more of organic acids, monosaccharides, oligosaccharides and polysaccharides, and the growth factor includes one or more of thiamine, riboflavin and pantothenic acid. The selection of diverse nitrogen and carbon sources can significantly increase the growth rate and metabolic activity of microorganisms, while the addition of growth factors promotes the reproduction and product production of microorganisms, and overall improves the efficiency and biological activity of the culture medium.

[0016] In a second aspect of the present invention, a dry microbial culture medium is provided, wherein the culture medium is prepared by the preparation method described above.

[0017] The third aspect of the present invention provides an application of a dry microbial culture medium, wherein the dry microbial culture medium prepared by the preparation method described in the first aspect is used for microbial culture.

[0018] Further, a sterilizing solution with a pH value of 7.0-7.2 is added to the dry microbial culture medium, and the medium is rehydrated for 30-60 minutes to obtain a dry film rehydration culture medium.

[0019] Furthermore, the sterilization solution is a 0-0.9% sodium chloride solution. Selecting a sodium chloride solution of appropriate concentration can not only maintain the osmotic pressure of the culture medium and promote the rehydration of microorganisms, but also provide a suitable ion environment for the microorganisms, which helps to improve their growth and metabolic efficiency.

[0020] Compared with the prior art, the beneficial results of the present invention are:

[0021] (1) The method for preparing the dry microbial culture medium provided by the present invention has a simple process and low cost, and can effectively improve the growth efficiency of microorganisms.

[0022] (2) The dry microbial culture medium provided by the present invention has good rehydration properties and biocompatibility, and can quickly restore its biological activity after rehydration, providing a good growth environment for microorganisms to ensure the effectiveness and reliability of the experimental process.

[0023] (3) The dry microbial culture medium provided by the present invention significantly improves the convenience and efficiency of experimental operations. The quality of the culture medium is controllable and can effectively improve the stability of the culture medium. Compared with traditional culture media, the preparation time and cost of the culture medium are greatly reduced, and it is suitable for large-scale production, transportation and storage, and can meet the wide range of needs in scientific research and industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] The accompanying drawings provide a further understanding of the embodiments and are incorporated into and constitute a part of this specification. The accompanying drawings illustrate embodiments and together with the description are used to explain the principles of the present invention, and it will be easy to recognize other embodiments and many expected advantages of the embodiments as they become better understood by reference to the following detailed description. The elements of the drawings are not necessarily to scale with each other. The same reference numerals refer to corresponding similar parts.

[0025] Figure 1 is a flow chart of a method for preparing a dry microbial culture medium according to one embodiment of the present invention;

[0026] Figure 2a is a graph showing the results of streaking experiments on five common microorganisms using a traditional agar medium and a dry film rehydrated medium according to an embodiment of the present invention;

[0027] Figure 2 b is a graph showing the results of a coating experiment on five common microorganisms using a traditional agar medium and a dry film rehydrated medium according to an embodiment of the present invention;

[0028] Figure 3 are SEM images and TEM images of the upper surface, lower surface and cross-section of a carrageenan film prepared at different gelation temperatures and drying temperatures according to one embodiment of the present invention;

[0029] Figure 4a is a surface morphology of a carrageenan film at -20°C, 0°C gelling temperature and 40°C drying temperature according to one embodiment of the present invention;

[0030] Figure 4b is a surface morphology of a carrageenan film at a gelling temperature of 10° C., 20° C., and a drying temperature of 40° C. according to an embodiment of the present invention;

[0031] Figure 4c is a surface morphology of a carrageenan film at drying temperatures of 60° C. and 80° C. according to an embodiment of the present invention;

[0032] Figure 5 This is an experimental photograph of a carrageenan film according to an embodiment of the present invention;

[0033] Figure 6 is an XRD spectrum of carrageenan films prepared at different gelation temperatures and drying temperatures according to one embodiment of the present invention;

[0034] Figure 7 a is a line graph showing the effect of different gelation temperatures and drying temperatures on the weight change rate of carrageenan film according to an embodiment of the present invention;

[0035] Figure 7 b is a line graph showing the effect of different gelation temperatures and drying temperatures on the thickness change rate of the carrageenan film according to an embodiment of the present invention. DETAILED DESCRIPTION

[0036] The technical solutions in the embodiments will be described clearly and completely below in conjunction with the drawings in the present invention. Similar component numbers in the drawings represent similar components. Obviously, the embodiments described below are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0037] It should be understood that when used in this specification and the appended claims, the terms "include" and "comprises" indicate the presence of described features, integers, steps, operations, elements and / or components, but do not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components and / or combinations thereof.

[0038] It should also be understood that the terms used in this specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. As used in the specification of the present invention and the appended claims, unless the context clearly indicates otherwise, the singular forms "a", "an" and "the" are intended to include plural forms.

[0039] In the first aspect, the present invention provides a flow chart of a method for preparing a dry microbial culture medium, as shown in FIG. Figure 1 As shown, the method includes:

[0040] S101, dissolving 7.5-15 parts of a nutrient, 2-3 parts of glycerin, 10-15 parts of a gelling agent, and 0.75-1.2 parts of potassium chloride in 500-1500 parts of deionized water to obtain a mixed solution;

[0041] S102, adjusting the pH value of the mixed solution to 7.0-7.2, sterilizing it at high temperature, pouring it into a plate, placing it at 0-20°C for gelation treatment for 30-60 minutes, and obtaining a gel product;

[0042] S103, drying the gel product under sterile conditions at 30-40°C and a relative humidity of 15-50% for 6-12 hours to obtain a dry microbial culture medium.

[0043] In some specific embodiments, 5-10 parts of tryptone, 2.5-5 parts of yeast extract powder, 2-3 parts of glycerol, 10-15 parts of gelling agent and 0.75-1.20 parts of potassium chloride are dissolved in 500-1500 parts of deionized water to obtain a mixed solution. The pH value of the mixed solution is adjusted to the range of 7.0-7.2, and then sterilized at a temperature of 100-150°C for 12-20 minutes. The sterilized mixed solution is injected into a disposable plate and gelled at a temperature of 0-20°C for 30-60 minutes. Drying is performed in a sterile drying room at a temperature of 30-40°C and a relative humidity of 15-50% for 6-12 hours, thereby obtaining a disposable dry microbial culture medium. Among them, tryptone can be replaced by bovine bone peptone or fish peptone, which can provide nitrogen source, amino acids and vitamins; yeast extract powder can be replaced by beef extract powder, which can provide nitrogen source, carbon source, vitamins and growth factors; glycerol, as a plasticizer for dry microbial culture medium, can prevent the culture medium from cracking after drying, and can also serve as a carbon source; the gelling agent contains 60%-100% carrageenan, 0-40% agar and 0-40% konjac flour. Carrageenan is the main material of dry microbial culture medium and has excellent rehydration ability. The combination of agar and carrageenan can improve the water retention of the culture medium, and the combination of konjac flour and carrageenan can improve the gel strength of the culture medium; potassium chloride can promote the dissolution of carrageenan, improve the gel strength of the culture medium, and provide suitable ionic strength for microbial growth.

[0044] In some specific embodiments, amino acids, ammonium sulfate, zinc sulfate and other substances can be added to the dry culture medium as nitrogen sources according to needs; organic acids (such as lactic acid, citric acid and fumaric acid), monosaccharides (such as glucose and fructose), oligosaccharides (such as sucrose and maltose) and polysaccharides (such as starch and cellulose) can be added as carbon sources. In addition, salts of metal ions such as phosphorus, sulfur, iron and magnesium, as well as growth factors such as thiamine, riboflavin and pantothenic acid can also be added.

[0045] In a second aspect, an embodiment of the present invention provides a dry microbial culture medium, which is prepared by the preparation method described in the first aspect.

[0046] In a third aspect, an embodiment of the present invention proposes an application of a dry microbial culture medium, wherein 20-40 times the weight of a 0-0.9% sodium chloride sterilization solution (pH=7-7.2) is added to the dry microbial culture medium prepared by the preparation method described in the first aspect, and after rehydration for 30-60 minutes, a disposable dry film rehydrated culture medium is obtained for microbial culture.

[0047] Example 1

[0048] Take 5 parts of tryptone, 2.5 parts of yeast extract powder, 8 parts of carrageenan, 1 part of agar, 1 part of konjac flour and 3 parts of glycerol, and dissolve them in 1000 parts of 10mM potassium chloride solution. Adjust the pH value of the solution to 7, and then sterilize it at a temperature of 121°C for 15 minutes. Pour the sterilized solution into a disposable plate and gelate it at 10°C for 60 minutes. Place the obtained gel in a clean drying room at a temperature of 40±0.5°C and a relative humidity of 15±5% for drying for 8 hours to obtain a dry microbial culture medium.

[0049] The dry microbial culture medium is rehydrated with a 0.45% sodium chloride sterilization solution (pH=7) 30 times the weight of the dry microbial culture medium for 60 minutes to obtain a disposable dry film rehydrated culture medium, which can be used for microbial culture.

[0050] Example 2

[0051] Take 4 parts of pancreatic bovine bone peptone, 3.5 parts of beef extract powder, 4 parts of carrageenan, 2 parts of agar, 2 parts of konjac flour and 3 parts of glycerol, and dissolve them in 1000 parts of 10mM potassium chloride solution. Adjust the pH value of the solution to 7.2, and then sterilize it at a temperature of 121°C for 15 minutes. Pour the sterilized solution into a disposable plate and gelate it at 15°C for 60 minutes. Place the obtained gel in a clean drying room at a temperature of 36±0.5°C and a relative humidity of 20±5% for drying for 8 hours to obtain a dry microbial culture medium.

[0052] The dry microbial culture medium is rehydrated with a 0.65% sodium chloride sterilization solution (pH=7) of 40 times the weight of the dry microbial culture medium for 60 minutes to obtain a disposable dry film rehydrated culture medium, which can be used for microbial culture.

[0053] Control group 1

[0054] 18 g of nutrient broth (containing 9 g of tryptone, 4.5 g of yeast extract and 4.5 g of sodium chloride) and 20 g of agar were dissolved in 1 L of deionized water, sterilized at 121° C. for 15 min, and then poured into disposable plates to prepare traditional agar medium for microbial culture.

[0055] The disposable dry film rehydration medium prepared according to Example 1 and the traditional agar medium prepared according to the control group were used for microbial culture experiments. Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), Proteus vulgaris (P. vulgaris), Bacillus cereus (B. cereus) and Bacillus subtilis (B. subtilis) were streaked and spread cultured, and the results were as follows: Figure 2 As shown. Compared with traditional agar medium, the dry film rehydrated medium has higher transparency and higher recognition of single colonies. The five bacteria cultured by streaking on the dry film rehydrated medium can all be isolated into single colonies, and the colony size is larger than that of the traditional medium under the same conditions. In addition, the five bacteria cultured by spreading on the dry film rehydrated medium can all grow normally, and their colonies are easier to count than those on the traditional agar medium.

[0056] Compared with the traditional culture medium that is prepared and used immediately, the disposable culture medium is more convenient in preparation, and the quality is easy to control, which can effectively improve the stability of the culture medium. Compared with the traditional disposable culture medium, the disposable dry culture medium has significant advantages in transportation and storage. It can be obtained by rapid rehydration before use, avoiding the problem of culture medium rupture during transportation and quality degradation caused by long-term storage. The physical and chemical properties of the carrier of the disposable dry culture medium of this application, namely the carrageenan film, were further determined.

[0057] Example 3

[0058] 10g carrageenan and 3g glycerol were dissolved in 1L potassium chloride solution with a concentration of 10mM (corresponding to 0.745g / L), and heated at 90°C for 1h to obtain a film-forming solution. The film-forming solution was poured into a disposable plate, and gelled at -20°C, 0°C, 10°C and 20°C for 60min, and then dried at 40±0.5°C and 15±5% relative humidity for 8h to obtain a gel group carrageenan film.

[0059] Control group 2

[0060] The difference from Example 3 is that the film-forming liquid is poured into a disposable plate and dried for 4 hours at a temperature of 60° C. or 80° C. and a relative humidity of 15±5% to obtain a direct drying group carrageenan film.

[0061] The gel group carrageenan film and the dry group carrageenan film prepared in Example 3 and Control Group 2 were subjected to microstructure, surface morphology, XRD, mechanical properties, swelling characteristics and mechanical properties of the gel film after rehydration. The experimental results are referenced to Figure 3-7 and Table 1-2.

[0062] Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to conduct microstructure experiments on carrageenan films. The carrageenan films were cut into 3mm×3mm rectangles to prepare SEM samples for surface observation. At the same time, the films were broken with liquid nitrogen to prepare SEM samples for cross-section observation. The SEM samples were placed in silica gel for dehydration for two weeks. Before observing the samples, they were fixed on the sample holder with conductive glue and sprayed with gold, and then SEM images of the film surface and cross-section were taken at an accelerating voltage of 10kV.

[0063] In the TEM experiment, 0.1g carrageenan and 0.03g glycerol were dissolved in 100mL of 1mM potassium chloride solution and heated at 90℃ for 1h to obtain a sample solution. 5μL of the sample solution was added to a 200-mesh carbon support film and gelled and dried using the temperature and humidity conditions for preparing the carrageenan film, and then negatively stained with 1% phosphotungstic acid to obtain a transmission electron microscope sample. The TEM image was taken using an accelerating voltage of 120kV.

[0064] Figure 3 The SEM images and TEM images of the upper surface, lower surface and cross section of the carrageenan film treated with different gelation temperatures and drying temperatures prepared according to the present application are shown, as shown in FIG. Figure 3As shown in the figure, the upper and lower surfaces and cross-sections of the carrageenan films obtained by gelation at -20 and 0°C and drying at 40°C all have wrinkled structures, and there are cracks and pores in the cross-sections. The wrinkled structure of the -20°C gel group is the most obvious. The upper and lower surfaces and cross-sections of the carrageenan films obtained by gelation at 10 and 20°C and drying at 40°C are flat and dense, while the upper and lower surfaces of the carrageenan films directly dried at 60 and 80°C all have salt particles precipitated, and there are obvious cracks in the cross-sections. SEM results show that gelation at a temperature not higher than zero degrees will induce carrageenan aggregation and form a wrinkled structure after drying. Normal temperature gelation helps to form a membrane with a dense network structure, while high-temperature drying will cause salt to precipitate from the inside of the membrane. TEM results show that the carrageenan film gelled at -20℃ has obvious aggregates and obvious gaps in the network structure, while the network structure of the carrageenan film gelled at 0, 10 and 20℃ is neat and orderly, among which the network structure of the 10℃ gel group is the most orderly, followed by the 20℃ gel group, and there are a large number of pores in their network structures, and their structures are similar to porous sponges. There are a large number of aggregates in the carrageenan film directly dried at 60 and 80℃, and it appears as a piled cloud state, and its structure is similar to rice cake. The results show that 10℃ gel is conducive to the formation of an orderly porous network structure, while sub-zero temperature gelation and high temperature drying will destroy the structure of the film.

[0065] In the surface morphology observation experiment, the carrageenan film was cut into a 10 mm × 10 mm rectangle and fixed on a silicon wafer with double-sided tape. The atomic force microscope was used for scanning in tapping mode, with a scanning size of 50 μm × 50 μm and a scanning rate of 0.996 Hz. NanoScope Analysis V1.10 was used to process and analyze the images.

[0066] The experimental results refer to Figures 4 and Figure 5 , Figure 4 and Figure 5 The surface morphology of the carrageenan film prepared according to the present application and the experimental photos are shown respectively. Figure 4a-4c As shown in the figure, the surface of the carrageenan film in the -20 and 0℃ gel groups has obvious wrinkles and protrusions. This is because the formation of ice crystals during the -20℃ gelation process leads to a reduction in solvent, which causes the aggregation of carrageenan. On the other hand, the surface protrusion height of the carrageenan film in the -20℃ gel group is the highest, followed by the 0℃ gel group, 80 and 60℃ drying group carrageenan films, while the 10 and 20℃ gel group carrageenan films have the lowest surface protrusion height. Among them, the protrusions on the surface of the carrageenan film directly dried at high temperature can be attributed to the precipitation of salt. R of carrageenan film a and R qThe value can reflect the degree of surface roughness, and its order is consistent with the height of surface protrusions, which are low-temperature gel group (-20 and 0℃ gelation) > high-temperature drying group (60 and 80℃ drying) > room-temperature gel group (10 and 20℃ gelation). Compared with the upper surface, the surface protrusion height and roughness of the lower surface are smaller, indicating that the upper surface in contact with air during the process of inducing gelation and drying film formation is more likely to form wrinkles and protrusions, and salt will also migrate to the upper surface with water to form salt particles. The effects of gelation temperature and drying temperature on the surface morphology can be intuitively observed through the photos of carrageenan films. The surface of the carrageenan film obtained by low-temperature gelation and drying has a tree-like structure. This is because the ice crystals formed during the low-temperature process promote the aggregation of carrageenan, and the lower the gelation temperature, the more obvious the tree-like structure. The surface of the carrageenan film obtained by drying after gelation at 10 and 20℃ is smooth and dense, while the surface of the carrageenan film directly dried at 60 and 80℃ has a corrugated structure, and the higher the drying temperature, the more obvious the corrugated structure (reference Figure 5 ).

[0067] In the XRD experiment, an X-ray diffractometer equipped with Cu-Kα was used to measure the XRD pattern of the carrageenan film at 40 kV and 30 mA. The diffraction pattern of the carrageenan film was collected in the range of 5-50° (2θ) at a rate of 5° / min.

[0068] Figure 6 The XRD pattern of the carrageenan film prepared according to the present application is shown in FIG. Figure 6 As shown in the figure, the 2θ angle of the amorphous peak of the carrageenan film in the 80℃ drying group and -20℃ gel group is significantly higher than that of the carrageenan film in the 60℃ drying group and 0℃ gel group, while the 2θ angle of the carrageenan film in the 10℃ and 20℃ gel groups is the smallest. In addition, the characteristic crystallization peaks of potassium chloride at 2θ = 28.22° and 40.38° can be observed in the XRD patterns of the carrageenan films in the 60℃ and 80℃ drying groups. According to the Bragg equation, the movement of the peak to a higher angle represents a decrease in the interlayer spacing of the polymer, that is, an increase in the degree of aggregation, which is consistent with the observation of the network structure by TEM and the surface structure by SEM, that is, high temperature will cause the structure to collapse and salt crystallization to precipitate, while the gel under conditions not higher than zero degrees may promote the aggregation of carrageenan by forming ice crystals.

[0069] In the mechanical properties experiment, the film samples were cut into 20mm×45mm rectangles and the thickness was measured. The tensile test was performed using a texture analyzer, and the initial distance and tensile speed were set to 30mm and 60mm / min, respectively. The tensile strength (TS) and elongation at break (EAB) of the film were calculated according to the formula. Where TS (MPa) = F m / A, where Fm is the maximum tension required for the membrane to break (N), and A is the cross-sectional area of ​​the membrane (m 2); EAB (%) = (L / 30) × 100, where L is the increased length when the film breaks (mm), and 30 is the initial distance between the clamps (mm).

[0070] Table 1 Effects of gelation and drying temperature on the mechanical properties of carrageenan films

[0071]

[0072] Note: Different lowercase letters indicate significant differences among the groups, P < 0.05.

[0073] Table 1 shows the mechanical properties data of carrageenan films treated with different gelation temperatures and drying temperatures prepared according to the present application. As can be seen from Table 1, the carrageenan films in the 10 and 20°C gel groups have the highest tensile strength (TS). As the gelation temperature decreases or the drying temperature increases, the TS of the carrageenan films gradually decreases, and the carrageenan films in the 80°C drying group have the lowest TS. Low-temperature gelation treatment promotes the formation of ice crystals, thereby causing local aggregation of carrageenan and destroying its network structure, resulting in a decrease in the TS of the carrageenan film. During the high-temperature drying process, carrageenan is not easy to form double helices and helical aggregates, and ultimately a collapsed rice cake-like structure ( Figure 3 On the other hand, the decrease of gelation temperature leads to a decrease in the elongation at break (EAB) of the carrageenan film, which can be attributed to the destruction of the film network structure by ice crystals, while high temperature drying leads to an increase in the EAB of the carrageenan film, which is due to the physical entanglement between chains, which increases the flexibility of the carrageenan film.

[0074] In the experiment of testing the swelling properties of carrageenan film, the film samples were cut into 20mm×45mm rectangles and placed in deionized water for rehydration. The weight and thickness of the dry film and the rehydrated gel film were measured. The dry film was placed in a vacuum drying oven to dry before testing. Before measuring the weight and thickness of the rehydrated gel film, the excess water on the surface was wiped off with filter paper, and the weight and thickness change rates were calculated. Among them, weight change rate (%) = (W h -W d )×100 / W d , where W h is the weight of the rehydrated gel, W d is the weight of dry film; thickness change rate (%) = (T h -T d )×100 / T d , where T h is the weight of the rehydrated gel, T d is the weight of the dry film.

[0075] Figure 7The following is a line graph showing the weight change rate (a) and thickness change rate (b) of the carrageenan film prepared according to the present invention and treated with different gelation temperatures and drying temperatures during swelling. Figure 7 As shown in the figure, gelation temperature and drying temperature have similar effects on the weight change rate and thickness change rate of carrageenan film. The results show that the carrageenan film in the 10 and 20℃ gel groups has the highest weight change rate and thickness change rate, indicating that it has the strongest water absorption capacity, while -20℃ gelation treatment or high temperature drying will significantly reduce the water absorption capacity of the carrageenan film, among which the water absorption capacity of the carrageenan film in the 80℃ drying group is the worst. This result shows that the ice crystals formed by low-temperature gelation will destroy the dense network structure of the carrageenan film, resulting in partial rupture of the interchain interactions in the sponge-like structure, thereby reducing the water absorption capacity of the sponge-like structure carrageenan film, while the rice cake-like structure carrageenan film obtained by high temperature drying does not have enough space to accommodate water molecules, so its water absorption capacity is much lower than that of the carrageenan film prepared by drying at 40℃.

[0076] In the mechanical properties test experiment of the rehydrated gel film, the film sample was cut into a dumbbell shape of 8mm×75mm and soaked in deionized water for 1h to measure the mechanical properties of the rehydrated gel film. The initial distance and tensile speed were set to 50mm and 60mm / min respectively. The calculation formulas of TS and EAB of the rehydrated gel film are consistent with those of the carrageenan film.

[0077] Table 2 Effects of gelation and drying temperature on the mechanical properties of rehydrated gel films

[0078]

[0079] Note: Different lowercase letters indicate significant differences among the groups, P < 0.05.

[0080] Table 2 shows the data of the mechanical properties of the gel after rehydration of the carrageenan film prepared according to the present application. As shown in Table 2, the gel obtained by rehydrating the carrageenan film of the -20°C gel group has the lowest TS and EAB, followed by the gel obtained by rehydrating the carrageenan film of the 0, 10 and 20°C gel group, while the gel obtained by rehydrating the carrageenan film of the 80°C drying group has the highest TS and EAB. The increase in the water content and the destruction of the structure of the gel will both lead to a decrease in TS. Therefore, the high water content and the destruction of the structure of the gel prepared by the -20°C gel group carrageenan film in this study led to its lowest TS, while the low water content of the gel prepared by the 60 and 80°C drying group carrageenan film led to its higher TS. On the other hand, the destruction of the structure will lead to a decrease in the EAB of the gel, while the interchain physical entanglement structure of the carrageenan film of the 60 and 80°C drying group can provide the gel with higher flexibility after absorbing water.

[0081] Obviously, those skilled in the art can make various modifications and changes to the embodiments of the present invention without departing from the spirit and scope of the present invention. In this way, if these modifications and changes are within the scope of the claims of the present invention and their equivalents, the present invention is also intended to cover these modifications and changes. The word "comprising" does not exclude the presence of other elements or steps not listed in the claims. The simple fact that certain measures are recorded in mutually different dependent claims does not indicate that the combination of these measures cannot be used to profit. Any reference numerals in the claims should not be considered to limit the scope.

Claims

1. A method for preparing a dry microbial culture medium, characterized in that: The method comprises: S1, dissolving 7.5-15 parts of a nutrient, 2-3 parts of glycerin, 10-15 parts of a gelling agent, and 0.75-1.2 parts of potassium chloride in 500-1500 parts of deionized water to obtain a mixed solution; S2, adjusting the pH value of the mixed solution to 7.0-7.2, sterilizing it at high temperature, pouring it into a plate, placing it at 0-20° C. for gelation treatment for 30-60 minutes, and obtaining a gel product; S3, drying the gel product under sterile conditions at 30-40° C. and a relative humidity of 15-50% for 6-12 hours to obtain the dried microbial culture medium.

2. The method for preparing a dry microbial culture medium according to claim 1, characterized in that: The nutrient comprises 5-10 parts of one or more of tryptone, bovine bone peptone and fish peptone, and 2.5-5 parts of one or more of yeast extract powder and beef extract powder.

3. The method for preparing a dry microbial culture medium according to claim 1, characterized in that: The gelling agent comprises 60-100% carrageenan, 0-40% agar and 0-40% konjac flour.

4. The method for preparing a dry microbial culture medium according to claim 1, characterized in that: In the step S2, after adjusting the pH value of the mixed solution to 7.0-7.2, the mixed solution is placed at a temperature of 100-150° C. and sterilized for 12-20 minutes.

5. The method for preparing a dry microbial culture medium according to claim 1, characterized in that: In the step S1, a supplement is added, and the supplement includes one or more of a nitrogen source, an organic acid, a carbon source, a salt containing metal ions, and a growth factor.

6. The method for preparing a dry microbial culture medium according to claim 5, characterized in that: The nitrogen source includes one or more of amino acids, ammonium sulfate and zinc sulfate, the carbon source includes one or more of organic acids, monosaccharides, oligosaccharides and polysaccharides, and the growth factor includes one or more of thiamine, riboflavin and pantothenic acid.

7. A dried microbial culture medium prepared according to the preparation method according to any one of claims 1 to 6.

8. An application of a dry microbial culture medium, characterized in that: The dried microbial culture medium prepared according to the preparation method according to any one of claims 1 to 6 is used for microbial culture.

9. The use of the dry microbial culture medium according to claim 8, characterized in that: A sterilizing solution with a pH value of 7.0-7.2 is added to the dry microbial culture medium, and the culture medium is rehydrated for 30-60 minutes to obtain a dry film rehydration culture medium.

10. The use of the dry microbial culture medium according to claim 9, characterized in that: The sterilizing solution is a 0-0.9% sodium chloride solution.

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