Method for preparing enzyme by fermenting fruits with edible mushrooms

Abstract

The invention belongs to the technical field of food processing, and particularly relates to a method for preparing enzyme by fermenting fruits with edible mushrooms. The invention provides a method for preparing enzyme by directly fermenting fruits by using a fungus liquid of edible fungi difficult to cultivate. The invention further provides a method for preparing the enzyme through secondary fermentation of the fruits with the fungus liquid of the edible fungi difficult to cultivate and the microbial strains. According to the method, on one hand, the problem that the difficulty of artificial cultivation of precious edible mushrooms difficult to cultivate is large is solved, meanwhile, nutritional ingredients of the fermented precious edible mushrooms can be directly used as enzymes, and the nutritional value of the final enzymes is improved; in the aspect of operation, the technological process is simplified, and the potential of large-scale production is achieved.

Description

Technical Field

[0001] The invention belongs to the technical field of food processing, and in particular relates to a method for preparing enzyme by fermenting fruit with edible fungi. Background Art

[0002] Enzymes contain a large number of biologically active ingredients and have become one of the research hotspots in this field. Conventional enzyme fermentation mostly uses lactic acid bacteria, acetic acid bacteria or yeast for fermentation. There are no reports of using fresh edible fungi as strains to ferment enzymes. Edible fungi are a kind of large fungi that can be used. At present, the research on edible fungi mainly focuses on the cultivation of the strains themselves, including the cultivation of fruiting bodies and the cultivation of high-density mycelium. Another application is edible fungi cultivation technology, that is, mushroom cultivation.

[0003] Raspberry, as a berry plant of the genus Rubus in the Rosaceae family, has a good market prospect and research value. However, this type of fruit, represented by raspberry, generally has the problem of extremely short shelf life and high storage difficulty. In order to improve the utilization rate of difficult-to-preserve fruits such as raspberries, they need to be deeply processed, for example, they are made into enzymes. The patent with publication number CN105707627A and name "a method for preparing a raspberry enzyme beverage" discloses the preparation of raspberry enzyme using raspberries as raw materials, inoculating spore-bearing Hansen yeast, Hansen Debary yeast, wine-leafed candidiasis, and acetic acid bacteria. The patented method uses common microbial strains for fermentation, and the nutritional components of the enzyme prepared are limited and the taste is relatively simple. The patent with publication number CN11955721A and name "a formula of a gastrodia elata edible fungus flavored polyphenol liquid plant enzyme and its preparation method" discloses a method for preparing enzymes using gastrodia elata and edible fungus powder as raw materials and adding probiotics. The patented method still uses common microbial strains for fermentation. Although edible fungi are used in the patent, the role of edible fungi is as a flavor or nutritional additive, not as a fermentation strain. However, preparing edible fungi into powder will inevitably lose some nutrients or bioactive ingredients, especially if rare strains are involved, the cost is often higher. Based on this, it is necessary to propose new methods and strategies. Summary of the invention

[0004] In view of this, the purpose of the present invention is to provide a method for preparing fruit enzymes by fermentation using edible fungi as fermentation strains. The specific technical scheme is as follows.

[0005] One aspect of the present invention provides a method for single fermentation of enzymes using edible fungi.

[0006] A method for preparing fruit enzyme by fermenting fruit with edible fungi, wherein the edible fungi are difficult to cultivate edible fungi, and the fruit includes berries or blackberries; the steps are as follows:

[0007] S01: taking fruits, drying them before pretreatment, and then ultra-fine grinding them to micron level to make fruit powder; or taking fresh fruits, beating them after pretreatment for later use;

[0008] S02: adding water to dissolve the fruit powder (obtaining fruit powder liquid) for enzymatic hydrolysis, and then adding complex sugar to prepare a raw material solution; or, taking pulped fresh fruit pulp for enzymatic hydrolysis, and then adding complex sugar to prepare a raw material solution;

[0009] S03: performing mild bacteriostasis on the raw material solution, wherein the mild bacteriostasis condition is a short-time treatment at a temperature not exceeding 85° C.;

[0010] The "short-time treatment" mentioned in the present invention means that the treatment time does not exceed 1 hour;

[0011] S04: pre-cultivating a difficult-to-cultivate edible fungus mass, then diluting the difficult-to-cultivate edible fungus mass with sterile water to obtain a difficult-to-cultivate edible fungus inoculum solution for fermenting the raw material solution; adding 3%-4% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry to obtain a culture solution in a sterile environment, further adjusting the pH value of the culture solution to no more than 5.5, setting the shaking table speed to 100-150r / min, and performing dynamic aerobic fermentation at room temperature for no more than 30 days to obtain a fermentation solution;

[0012] In some preferred embodiments, the dynamic aerobic fermentation temperature is 25°C;

[0013] S05: breaking the cell wall of the fermentation liquid, filtering it, and then performing mild bacteriostasis, wherein the mild bacteriostasis condition is a short-time treatment at an environment not exceeding 85° C.; obtaining a fruit enzyme.

[0014] It is understandable that the pre-treatment includes washing and drying the fruits to be treated.

[0015] Furthermore, the difficult-to-cultivate edible fungi include Grifola frondosa, Pseudomonas aeruginosa, Russula russula, Chanterelle or Schizophyllum.

[0016] Furthermore, the enzymatic hydrolysis uses a complex enzyme of cellulase and pectinase, the mass ratio of the cellulase to the pectinase is 1:1; the complex sugar includes glucose, brown sugar and white sugar; the mass ratio of the glucose, brown sugar and white sugar is 1:1:1.

[0017] It is to be understood that the types of complex sugars are merely examples and not limitations. A person skilled in the art can prepare complex sugars according to the taste requirements of the enzyme.

[0018] Furthermore, the mild bacteriostasis is low-temperature pasteurization, including sterilization at 60°C for 30 minutes or sterilization at 85°C for 15 minutes.

[0019] The preferred mild antibacterial condition is sterilization at 85°C for 15 minutes.

[0020] Another aspect of the present invention provides a method for secondary fermentation of enzymes using edible fungi and lactic acid bacteria.

[0021] A method for preparing fruit enzyme by fermenting fruit with edible fungi and lactic acid bacteria, characterized in that the edible fungi are difficult to cultivate edible fungi, and the fruit includes berries or blackberries; the steps are as follows:

[0022] S01: taking fruits, drying them before pretreatment, and then ultra-fine grinding them to micron level to make fruit powder; or taking fresh fruits, beating them after pretreatment for later use;

[0023] S02: adding water to dissolve the fruit powder (obtaining fruit powder liquid) for enzymatic hydrolysis, and then adding complex sugar to prepare a raw material solution; or, taking pulped fresh fruit pulp for enzymatic hydrolysis, and then adding complex sugar to prepare a raw material solution;

[0024] S03: performing mild bacteriostasis on the raw material solution, wherein the mild bacteriostasis condition is a short-time treatment at a temperature not exceeding 85° C.;

[0025] S04: pre-cultivating a difficult-to-cultivate edible fungus mass, then diluting the difficult-to-cultivate edible fungus mass with sterile water to obtain a difficult-to-cultivate edible fungus inoculum solution for fermenting the raw material solution; adding 3%-4% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry to the difficult-to-cultivate edible fungus inoculum solution in a sterile environment to obtain a culture solution, further adjusting the pH value of the culture solution to no more than 5.5, setting a first fermentation temperature for dynamic aerobic fermentation, and fermenting for no more than 30 days to obtain a fermentation solution;

[0026] S05: breaking the cell wall of the fermentation broth and filtering it, and then performing mild bacteriostasis, wherein the mild bacteriostasis condition is a short-time treatment at a temperature not exceeding 85° C.; obtaining a first enzyme;

[0027] The "short-time treatment" mentioned in the present invention means that the treatment time does not exceed 1 hour;

[0028] S06: adding 6%-8% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry to the first enzyme, and then adding microbial strains for fermentation; the added microbial strains are mixed lactic acid bacteria with a volume of 0.1%-0.3% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry, setting a second fermentation temperature for static anaerobic fermentation, and the fermentation time is no more than 7 days to obtain a second enzyme; the second fermentation temperature is higher than the first fermentation temperature;

[0029] S07: performing mild antibacterial treatment on the second enzyme, wherein the mild antibacterial condition is a short-time treatment at a temperature not exceeding 85° C. to obtain a finished fruit enzyme product.

[0030] Furthermore, the difficult-to-cultivate edible fungi include Grifola frondosa, Pseudomonas aeruginosa, Russula russula, Chanterelle or Schizophyllum.

[0031] Furthermore, the enzymatic hydrolysis uses a complex enzyme of cellulase and pectinase, the mass ratio of the cellulase to the pectinase is 1:1; the complex sugar includes glucose, brown sugar and white sugar; the mass ratio of the glucose, brown sugar and white sugar is 1:1:1.

[0032] Furthermore, the mixed lactic acid bacteria include one or more selected from Lactobacillus plantarum, Lactobacillus casei, Lactobacillus delbrueckii, Lactobacillus fermentum and / or Lactobacillus bulgaricus.

[0033] Furthermore, the first fermentation temperature is 25°C; the second fermentation temperature is 37°C; and the mild bacteriostasis is low-temperature pasteurization, including sterilization at 60°C for 30 minutes or sterilization at 85°C for 15 minutes.

[0034] Furthermore, the shaking parameter of the dynamic fermentation is 100-150r / min.

[0035] Beneficial technical effects

[0036] (1) The present invention provides a method for producing fruit enzymes by fermenting precious and difficult-to-cultivate edible fungi as fermentation strains; no separation or purification is required during the fermentation process and the antibacterial conditions are mild. The entire fermentation system can be used as a natural culture medium for difficult-to-cultivate edible fungi, and the fermentation product (enzyme) obtained in the end can be directly drunk. On the one hand, the method of the present invention alleviates the problem of the difficulty of artificially cultivating precious and difficult-to-cultivate edible fungi (including long cultivation time period or high cultivation cost). At the same time, the nutrients of the fermented precious edible fungi can be directly used as enzymes, thereby improving the nutritional value (richer content of polysaccharides and antioxidants) and taste level of the final enzyme. From the operational point of view, the process flow is also simplified, and it has the potential for large-scale production. The method of the present invention also alleviates the problem of preserving berries or berries that are difficult to store or difficult to store.

[0037] (2) The method of the present invention using difficult-to-cultivate edible fungi as fermentation strains for fermentation belongs to a closed-loop biological production system. On the one hand, the secondary metabolites produced by edible fungi during the growth process have certain health effects, which can give the enzyme additional value. At the same time, the edible fungi strains will produce substances that inhibit harmful bacteria during the growth process, making the enzyme less likely to be contaminated. Therefore, the method of the present invention only needs to use mild antibacterial conditions to achieve antibacterial and maintain the activity of the ingredients. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings required for use in the embodiments or the prior art descriptions are briefly introduced below. In all the drawings, similar elements or parts are generally identified by similar reference numerals. In the drawings, each element or part is not necessarily drawn according to the actual scale. Obviously, the drawings described below are some embodiments of the present invention. For those of ordinary skill in the art, other drawings can also be obtained based on these drawings without paying creative labor.

[0039] Figure 1 This is a process flow chart of preparing enzymes by fermenting fruits with edible fungi in one embodiment of the present invention;

[0040] Figure 2 This is a process flow chart of preparing enzymes by fermenting fruits with edible fungi and lactic acid bacteria in one embodiment of the present invention;

[0041] Figure 3 This is the effect of the edible fungus inoculation amount on the fermentation mycelium amount verified in one of the embodiments of the present invention. DETAILED DESCRIPTION

[0042] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0043] Herein "and / or" includes any and all combinations of one or more of the associated listed items.

[0044] Herein, "plurality" means two or more than two, ie, it includes two, three, four, five, etc.

[0045] It should be noted that, in this article, the terms "include", "comprises" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or device. In the absence of further restrictions, an element defined by the sentence "comprises a ..." does not exclude the existence of other identical elements in the process, method, article or device including the element.

[0046] As used in this specification, the term "about" typically means + / - 5% of the stated value, more typically + / - 4% of the stated value, more typically + / - 3% of the stated value, more typically + / - 2% of the stated value, even more typically + / - 1% of the stated value, and even more typically + / - 0.5% of the stated value.

[0047] In this specification, some embodiments may be disclosed in a format of being within a certain range. It should be understood that such description of "being within a certain range" is only for convenience and brevity, and should not be interpreted as a rigid limitation on the disclosed range. Therefore, the description of the range should be considered to have specifically disclosed all possible sub-ranges and independent numerical values ​​within this range. For example, the range The description of should be considered to have specifically disclosed sub-ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within this range, for example, 1, 2, 3, 4, 5, and 6. The above rules apply regardless of the breadth of the range.

[0048] Glossary

[0049] The "difficult-to-cultivate edible fungi" of the present invention refers to edible fungi that are difficult to separate manually, difficult to cultivate, have low yields or high cultivation costs, for example, including the Grifola frondosa, Bacillus cerevisiae, Russula aeruginosa, Russula russula, Chanterelle or Schizophyllum glabra of the present invention.

[0050] The "berries or berries" mentioned in the present invention refer to fruits with a short shelf life, which are not easy to store or are difficult to store, for example, raspberries, strawberries, blueberries, blackberries, mulberries, grapes, cherries, kiwis, papayas, guavas, carambolas, figs, and sapodillas.

[0051] Example 1

[0052] This embodiment provides an example of preparing enzymes by fermenting fruits with edible fungi. This method is also called a primary fermentation method. The specific steps of the method are as follows.

[0053] (1) Raw material selection: Select wild raspberries that are fresh in appearance, have firm stems, full grains, and are uniform in size.

[0054] (2) Sorting and cleaning: Soak wild raspberries in clean water to remove dust and impurities on the surface of the raspberries. During the cleaning process, try to avoid damaging the raspberries to prevent loss of nutrients.

[0055] (3) Ultrafine grinding: Use appropriate ultrafine grinding conditions to grind the raspberries into micron size.

[0056] (4) Low temperature drying: The ultrafinely crushed raspberries are freeze-dried at a relatively low temperature to form raspberry powder.

[0057] (5) Adding water: Take a certain amount of raspberry powder in a fermentation bottle, and add purified water 7 times the mass of the raspberry powder to fully dissolve the raspberry powder and place it in a transparent bottle container.

[0058] (6) Enzymatic hydrolysis: Add 2.2% of the mass of raspberry powder as a composite enzyme, adjust the pH value of the raw material solution to 5 (analytical grade sodium hydroxide can be used), and then perform enzymatic hydrolysis in a 62° C. water bath environment for 140 min; the composite enzyme is a complex of cellulase and pectinase, and the mass ratio thereof is 1:1.

[0059] (7) Adding sugar: Optionally, a composite sugar of 5% of the weight of raspberry powder is added to the raspberry liquid in step (6) to increase the flavor; the composite sugar is a mixture of glucose, brown sugar and white sugar, and the mass ratio is 1:1:1. It is understood that the composite sugar in this embodiment is only an example and not a limitation. Those skilled in the art can prepare composite sugars or even a single type of sugar solution according to the taste requirements of the enzyme.

[0060] (8) Mild antibacterial: After adding sugar solution, sterilize the raspberry juice in an 85°C water bath for 15 minutes (pasteurization).

[0061] (9) Grifola frondosa fermentation: Using potato dextrose broth medium, Grifola frondosa fungi were cultured at 25°C and a shaking speed of 150 r / min. Then, a Grifola frondosa inoculum was prepared under sterile conditions with sterile water at a ratio of 1 g:10 mL. Then, 3% of the volume of raspberry liquid was added to the Grifola frondosa inoculum to obtain a culture solution, and the pH value of the culture solution was further adjusted to 4.5. Then, the shaking speed was set to 120 r / min on a constant temperature shaking table at room temperature (preferably 25°C) and fermented for 22 days to obtain a fermented solution.

[0062] (10) Filtration: In a sterile environment, the fermentation liquid is placed in a wall-breaking machine to break the mycelium of Grifola frondosa, allowing the nutrients to flow out. The fermentation liquid is then filtered through gauze, and the filtrate is taken to obtain raspberry enzyme.

[0063] (11) Mild antibacterial: The obtained raspberry enzyme is sterilized in a water bath at 85°C for 15 minutes (pasteurization).

[0064] (12) Packaging: Take the raspberry enzyme under sterile conditions, package and seal it to obtain the finished raspberry enzyme.

[0065] The process flow chart of this embodiment is shown in Figure 1 .

[0066] The functional activity of the raspberry enzyme obtained by fermentation in this example was measured below, and the results are shown in Table 1.

[0067] Table 1 Determination results of relevant indicators of raspberry enzyme fermented by Grifola frondosa

[0068] This example further explores the inoculation amount of Grifola frondosa species.

[0069] The experiment found that as the inoculation amount of the strain increased, the amount of mycelium increased first, reaching the maximum at 3%. It began to decrease when the inoculation amount of the strain was 4%. The possible reason was that the nutrients were consumed too quickly, the strains showed self-melting, and the amount of mycelium decreased. When the inoculation amount of the strain was 1%-2%, the strain content was small and the amount of mycelium was low. Therefore, the inoculation amount of 3%-5% was selected. Related results can be found in Figure 3 .

[0070] Example 2

[0071] This embodiment provides an example of preparing enzymes by fermenting fruits with edible fungi and lactic acid bacteria. This method is also called a secondary fermentation method. The specific steps of the method are as follows.

[0072] (1) Raw material selection: Select wild raspberries that are fresh in appearance, have firm stems, full grains, and are uniform in size.

[0073] (2) Sorting and cleaning: Soak wild raspberries in clean water to remove dust and impurities on the surface of the raspberries. During the cleaning process, try to avoid damaging the raspberries to prevent loss of nutrients.

[0074] (3) Ultrafine grinding: Use appropriate ultrafine grinding conditions to grind the raspberries into micron size.

[0075] (4) Low temperature drying: The ultrafinely crushed raspberries are freeze-dried at a relatively low temperature to form raspberry powder.

[0076] (5) Adding water: Take a certain amount of raspberry powder in a fermentation bottle, and add purified water 7 times the mass of the raspberry powder to fully dissolve the raspberry powder and place it in a transparent bottle container.

[0077] (6) Enzymatic hydrolysis: Add 2.2% of the mass of raspberry powder as a composite enzyme, adjust the pH value of the raw material solution to 5 (analytical grade sodium hydroxide can be used), and then perform enzymatic hydrolysis in a 62° C. water bath environment for 140 min; the composite enzyme is a complex of cellulase and pectinase, and the mass ratio thereof is 1:1.

[0078] (7) Adding sugar: Optionally, a composite sugar of 5% of the weight of raspberry powder is added to the raspberry liquid in step (6) to increase the flavor; the composite sugar is a mixture of glucose, brown sugar and white sugar, and the mass ratio is 1:1:1. It is understood that the composite sugar in this embodiment is only an example and not a limitation. Those skilled in the art can prepare composite sugars or even a single type of sugar solution according to the taste requirements of the enzyme.

[0079] (8) Mild antibacterial: Sterilize the raspberry juice after adding sugar in an 85℃ water bath for 15 minutes.

[0080] (9) Grifola frondosa fermentation: Using potato dextrose broth medium, Grifola frondosa mass was cultured at 25°C and a shaking speed of 150 r / min. Then, sterile water was used to prepare an inoculum at a ratio of 1 g:10 mL under sterile conditions. Then, 3% of the volume of raspberry liquid was added to the Grifola frondosa inoculum to obtain a culture solution in a sterile environment, and the pH value of the culture solution was further adjusted to 4.5. Then, the shaking speed was set to 120 r / min on a constant temperature shaking table at room temperature (preferably 25°C), and fermented for 22 days to obtain a fermentation solution.

[0081] (10) Filtration: In a sterile environment, the fermentation liquid is placed in a wall breaking machine to break the mycelium of Grifola frondosa in the fermentation liquid, so that the nutrients flow out, and the fermentation liquid is filtered through gauze to obtain the filtrate to obtain the first enzyme.

[0082] (11) Lactic acid bacteria fermentation: In a sterile environment, 7.1% of the mass of raspberry powder in complex sugar and 0.24% of the mass of raspberry powder in mixed lactic acid bacteria fermentation agent are added to the filtrate, and fermented at 37°C for 7 days to obtain the second enzyme. The preferred mixed lactic acid bacteria fermentation agent is the probiotic fruit and vegetable enzyme fermentation agent produced by Angel Yeast.

[0083] (12) Mild antibacterial: The fermented second enzyme is sterilized in a water bath at 85°C for 15 minutes (pasteurization).

[0084] (13) Packaging: Take the raspberry enzyme under sterile conditions, package and seal it to obtain the finished raspberry enzyme.

[0085] The process flow chart of this embodiment is shown in Figure 2 .

[0086] The shelf life of the raspberry enzyme obtained by fermentation in this example was predicted below. The enzyme was stored at 4°C, 25°C, and 37°C, and samples were taken every 3 days to detect the SOD enzyme activity and total colony count of the raspberry enzyme. The results are shown in Table 2.

[0087] Table 2 Results of SOD enzyme activity and total colony count of raspberry enzyme

[0088] Since the total number of colonies was not detected, the SOD enzyme activity value was used as the evaluation index for the shelf life of raspberry enzyme. It was calculated that the shelf life of raspberry enzyme at 25°C was 116 days.

[0089] Example 3

[0090] This embodiment provides another example of preparing enzyme by fermenting fruit with edible fungi. The specific steps of the method are as follows.

[0091] (1) Raw material selection: Select wild raspberries that are fresh in appearance, have firm stems, full grains, and are uniform in size.

[0092] (2) Sorting and cleaning: Soak wild raspberries in clean water to remove dust and impurities on the surface of the raspberries. During the cleaning process, try to avoid damaging the raspberries to prevent loss of nutrients.

[0093] (3) Ultrafine grinding: Use appropriate ultrafine grinding conditions to grind the raspberries into micron size.

[0094] (4) Low temperature drying: The ultrafinely crushed raspberries are freeze-dried at a relatively low temperature to form raspberry powder.

[0095] (5) Adding water: Take a certain amount of raspberry powder in a fermentation bottle, and add purified water 7 times the mass of the raspberry powder to fully dissolve the raspberry powder and place it in a transparent bottle container.

[0096] (6) Enzymatic hydrolysis: Add 2.2% of the mass of raspberry powder as a composite enzyme, adjust the pH value of the raw material solution to 5 (analytical grade sodium hydroxide can be used), and then perform enzymatic hydrolysis in a 62° C. water bath environment for 140 min; the composite enzyme is a complex of cellulase and pectinase, and the mass ratio thereof is 1:1.

[0097] (7) Adding sugar: Optionally, a complex sugar with a mass ratio of 6% of raspberry powder is added to the raspberry juice in step (6) to increase the flavor; the complex sugar is a mixture of glucose, brown sugar and white sugar, and the mass ratio thereof is 1:1:1.

[0098] (8) Mild antibacterial: Sterilize the raspberry juice after adding sugar in an 85℃ water bath for 15 minutes.

[0099] (9) Schizophyllum fermentation: Using potato dextrose broth medium, Schizophyllum bacterial mass was cultured at 25°C and a shaking speed of 150 r / min. Then, sterile water was used under sterile conditions at a ratio of 1 g:10 mL to obtain an inoculum. Then, 3.5% of the volume of raspberry liquid was added to the Schizophyllum inoculum to obtain a culture solution in a sterile environment, and the pH value of the culture solution was further adjusted to 5.5. Then, on a constant temperature shaking table at room temperature (25°C), the shaking speed was set to 120 r / min, and fermented for 20 days to obtain a fermentation solution.

[0100] (10) Filtration: In a sterile environment, the fermentation liquid is placed in a wall-breaking machine to break the mycelium of Schizophyllum in the fermentation liquid, so that the nutrients flow out, and then filtered with gauze to obtain the filtrate.

[0101] (11) Mild antibacterial: Sterilize the fermented raspberry enzyme in an 85°C water bath for 15 minutes.

[0102] (12) Packaging: Take the raspberry enzyme under sterile conditions, package and seal it to obtain the finished raspberry enzyme.

[0103] Example 4

[0104] This embodiment provides another example of preparing enzyme by secondary fermentation of fruit with edible fungi and lactic acid bacteria. The specific steps of the method are as follows.

[0105] (1) Raw material selection: Select wild raspberries that are fresh in appearance, have firm stems, full grains, and are uniform in size.

[0106] (2) Sorting and cleaning: Soak wild raspberries in clean water to remove dust and impurities on the surface of the raspberries. During the cleaning process, try to avoid damaging the raspberries to prevent loss of nutrients.

[0107] (3) Ultrafine grinding: Use appropriate ultrafine grinding conditions to grind the raspberries into micron size.

[0108] (4) Low temperature drying: The ultrafinely crushed raspberries are freeze-dried at a relatively low temperature to form raspberry powder.

[0109] (5) Adding water: Take a certain amount of raspberry powder in a fermentation bottle, and add purified water 7 times the mass of the raspberry powder to fully dissolve the raspberry powder and place it in a transparent bottle container.

[0110] (6) Enzymatic hydrolysis: Add 2.2% of the mass of raspberry powder as a composite enzyme, adjust the pH value of the raw material solution to 5 (analytical grade sodium hydroxide can be used), and then perform enzymatic hydrolysis in a 62° C. water bath environment for 140 min; the composite enzyme is a complex of cellulase and pectinase, and the mass ratio thereof is 1:1.

[0111] (7) Adding sugar: Optionally, a complex sugar with a mass ratio of 5% of the mass of raspberry powder is added to the raspberry juice in step (6) to increase the flavor; the complex sugar is a mixture of glucose, brown sugar and white sugar, and the mass ratio thereof is 1:1:1.

[0112] (8) Mild antibacterial: Sterilize the raspberry juice after adding sugar in an 85℃ water bath for 15 minutes.

[0113] (9) Schizophyllum fermentation: Using potato dextrose broth medium, Schizophyllum bacterial mass was cultured at 25°C and a shaking speed of 150 r / min. Then, sterile water was used to prepare an inoculum at a ratio of 1 g:10 mL under sterile conditions. Then, 3.5% of the volume of raspberry liquid was added to the Schizophyllum inoculum to obtain a culture solution in a sterile environment, and the pH value of the culture solution was further adjusted to 5.5. Then, the shaking speed was set to 120 r / min on a constant temperature shaking table at room temperature (25°C) and fermented for 20 days to obtain a fermentation solution.

[0114] (10) Filtration: In a sterile environment, the fermentation liquid is placed in a wall-breaking machine to break the mycelium of Schizophyllum in the fermentation liquid, so that the nutrients flow out, and the liquid is filtered through gauze to obtain the filtrate to obtain the first enzyme.

[0115] (11) Lactic acid bacteria fermentation: In a sterile environment, 7.1% of the mass of raspberry powder in complex sugar and 0.24% of the mass of raspberry powder in mixed lactic acid bacteria fermentation agent are added to the filtrate, and fermented at 37°C for 7 days to obtain the second enzyme. The preferred mixed lactic acid bacteria fermentation agent is the probiotic fruit and vegetable enzyme fermentation agent produced by Angel Yeast.

[0116] (12) Mild antibacterial: Sterilize the fermented second enzyme, raspberry enzyme, in a water bath at 85°C for 15 minutes.

[0117] (13) Packaging: Take the raspberry enzyme under sterile conditions, package and seal it to obtain the finished raspberry enzyme.

[0118] The embodiments of the present invention are described above in conjunction with the accompanying drawings, but the present invention is not limited to the above-mentioned specific implementation modes, which are merely illustrative rather than restrictive. Under the guidance of the present invention, ordinary technicians in this field can also make many forms without departing from the scope of protection of the present invention and the claims, all of which are within the protection of the present invention.

Claims

1. A method for preparing fruit enzyme by fermenting fruit with edible fungi, characterized in that: The edible fungi are difficult to cultivate edible fungi, and the fruits include berries or blackberries; the steps are as follows: S01: The fruit is dried and then ultra-finely crushed to micron level to make fruit powder; Alternatively, take fresh fruit and pulp it after pre-treatment for later use; S02: adding water to dissolve the fruit powder for enzymolysis, and then adding complex sugar to prepare a raw material solution; or taking pulped fresh fruit pulp for enzymolysis, and then adding complex sugar to prepare a raw material solution; S03: performing mild antibacterial treatment on the raw material solution, wherein the mild antibacterial condition is a short-time treatment at a temperature not exceeding 85° C.; S04: pre-cultivating a difficult-to-cultivate edible fungus mass, then diluting the difficult-to-cultivate edible fungus mass with sterile water to obtain a difficult-to-cultivate edible fungus inoculum solution for fermenting the raw material solution; adding 3%-4% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry to obtain a culture solution in a sterile environment, further adjusting the pH value of the culture solution to no more than 5.5, setting the shaking table speed to 100-150r / min, and performing dynamic aerobic fermentation at room temperature for no more than 30 days to obtain a fermentation solution; S05: The fermentation liquid is subjected to cell wall breaking and then filtered, and then subjected to mild bacteriostasis, wherein the mild bacteriostasis condition is a short-time treatment at an environment not exceeding 85° C.; and fruit enzyme is obtained.

2. The method according to claim 1, characterized in that The difficult-to-cultivate edible fungi include Grifola frondosa, Pseudomonas aeruginosa, Russula aurea, Russula ruber, Chanterelle or Schizophyllum.

3. The method according to claim 1, characterized in that The enzymatic hydrolysis uses a composite enzyme of cellulase and pectinase, the mass ratio of the cellulase to the pectinase is 1:1; the composite sugar includes glucose, brown sugar and white sugar; the mass ratio of the glucose, brown sugar and white sugar is 1:1:

1.

4. The method according to claim 1, characterized in that The mild bacteriostasis is low-temperature pasteurization, including sterilization at 60°C for 30 minutes or sterilization at 85°C for 15 minutes.

5. A method for preparing fruit enzymes by fermenting fruits with edible fungi and lactic acid bacteria, characterized in that: The edible fungi are difficult to cultivate edible fungi, and the fruits include berries or blackberries; the steps are as follows: S01: The fruit is dried and then ultra-finely crushed to micron level to make fruit powder; Alternatively, take fresh fruit and pulp it after pre-treatment for later use; S02: adding water to dissolve the fruit powder for enzymolysis, and then adding complex sugar to prepare a raw material solution; or taking pulped fresh fruit pulp for enzymolysis, and then adding complex sugar to prepare a raw material solution; S03: performing mild bacteriostasis on the raw material solution, wherein the mild bacteriostasis condition is a short-time treatment at a temperature not exceeding 85° C.; S04: pre-cultivating a difficult-to-cultivate edible fungus mass, then diluting the difficult-to-cultivate edible fungus mass with sterile water to obtain a difficult-to-cultivate edible fungus inoculum solution for fermenting the raw material solution; adding 3%-4% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry to the difficult-to-cultivate edible fungus inoculum solution in a sterile environment to obtain a culture solution, further adjusting the pH value of the culture solution to no more than 5.5, setting a first fermentation temperature for dynamic aerobic fermentation, and fermenting for no more than 30 days to obtain a fermentation solution; S05: breaking the cell wall of the fermentation broth and filtering it, and then performing mild bacteriostasis, wherein the mild bacteriostasis condition is a short-time treatment at a temperature not exceeding 85° C.; obtaining a first enzyme; S06: adding 6%-8% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry to the first enzyme, and then adding microbial strains for fermentation; the added microbial strains are mixed lactic acid bacteria with a volume of 0.1%-0.3% of the volume of the fruit powder liquid or the volume of the fresh fruit slurry, setting a second fermentation temperature for static anaerobic fermentation, and the fermentation time is no more than 7 days to obtain a second enzyme; the second fermentation temperature is higher than the first fermentation temperature; S07: performing mild antibacterial treatment on the second enzyme, wherein the mild antibacterial condition is a short-time treatment at a temperature not exceeding 85° C. to obtain a finished fruit enzyme product.

6. The method according to claim 5, characterized in that The difficult-to-cultivate edible fungi include Grifola frondosa, Pseudomonas aeruginosa, Russula aurea, Russula ruber, Chanterelle or Schizophyllum.

7. The method according to claim 5, characterized in that The enzymatic hydrolysis uses a composite enzyme of cellulase and pectinase, the mass ratio of the cellulase to the pectinase is 1:1; the composite sugar includes glucose, brown sugar and white sugar; the mass ratio of the glucose, brown sugar and white sugar is 1:1:

1.

8. The method according to claim 5, characterized in that The mixed lactic acid bacteria include one or more selected from Lactobacillus plantarum, Lactobacillus casei, Lactobacillus delbrueckii, Lactobacillus fermentum and / or Lactobacillus bulgaricus.

9. The method according to claim 5, characterized in that The first fermentation temperature is 25°C; the second fermentation temperature is 37°C; the mild bacteriostasis is low-temperature pasteurization, including sterilization at 60°C for 30 minutes or sterilization at 85°C for 15 minutes.

10. The method according to claim 5, characterized in that The shaking parameter of the dynamic fermentation is 100-150r / min.

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