CN116814718A - Oyster polysaccharide extract and preparation method thereof - Google Patents
Oyster polysaccharide extract and preparation method thereof Download PDFInfo
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- CN116814718A CN116814718A CN202310949178.XA CN202310949178A CN116814718A CN 116814718 A CN116814718 A CN 116814718A CN 202310949178 A CN202310949178 A CN 202310949178A CN 116814718 A CN116814718 A CN 116814718A
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Abstract
The application relates to oyster polysaccharide extract and a preparation method thereof. The preparation method of oyster polysaccharide extract comprises the following steps: mixing the primarily processed oyster meat with water, pulping, and preparing oyster pulp; adding amylase and compound protease into oyster pulp for enzymolysis, and inactivating enzyme to prepare an enzymolysis solution; inoculating lactobacillus into the enzymolysis liquid to perform first fermentation to prepare a first fermentation product, and collecting fermentation liquid from the first fermentation product to prepare a first supernatant; inoculating Saccharomyces cerevisiae embryo into the first supernatant for secondary fermentation to prepare a secondary fermentation product, and collecting fermentation liquor from the secondary fermentation product to prepare oyster polysaccharide extract. In the enzymolysis process, the protein in the oyster is decomposed into amino acid to the maximum extent, then the amino acid is converted into acetic acid through lactobacillus fermentation, and then the saccharomyces cerevisiae is used for fermentation, so that the acetic acid is further converted into substances such as glucose, polysaccharide and the like, and the extraction degree of the polysaccharide in the oyster is greatly improved.
Description
Technical Field
The application relates to the technical field of oyster processing, in particular to an oyster polysaccharide extract and a preparation method thereof.
Background
Polysaccharide is a natural product, widely exists in organic matters such as plants, microorganisms, animals and the like, and has wide application in clinic, food industry, fermentation industry and petroleum industry. The oyster contains various active substances peculiar to marine organisms, and polysaccharide contained in the oyster has biological activities of regulating organism immunity, inhibiting tumor, delaying aging, reducing blood sugar, reducing blood fat and the like.
At present, most oyster polysaccharide is extracted from the existing glycogen of fresh oyster, the original oyster raw material is required to have higher sugar content, and the oyster polysaccharide with low sugar content or oyster polysaccharide with reduced sugar content after primary processing has poor extraction effect.
Disclosure of Invention
Based on this, it is necessary to provide a method for producing oyster polysaccharide extract, which can be suitably used for polysaccharide extraction of oyster having a low sugar content.
Further, it is also necessary to provide an oyster polysaccharide extract.
An embodiment of the application provides a preparation method of oyster polysaccharide extract, comprising the following steps:
mixing the primarily processed oyster meat with water, pulping, and preparing oyster pulp;
adding amylase and compound protease into the oyster pulp for enzymolysis, and inactivating enzyme to prepare an enzymolysis solution;
inoculating lactobacillus into the enzymolysis liquid to perform first fermentation to prepare a first fermentation product, and collecting fermentation liquid from the first fermentation product to prepare a first supernatant;
inoculating saccharomyces cerevisiae embryo into the first supernatant to perform second fermentation to prepare a second fermentation product, and collecting fermentation liquor from the second fermentation product to prepare oyster polysaccharide extract.
In one embodiment, the conditions for enzymolysis include:
the pH value is 7.0-8.0, the enzymolysis temperature is 50-60 ℃, and the enzymolysis time is 8-12 h.
In one embodiment, the amylase is used in an amount of 2 g-5 g per kg oyster slurry calculated by 60000U/g enzyme activity.
In one embodiment, the amount of the compound protease is 2 g-10 g per kg oyster slurry calculated by 600000U/g enzyme activity.
In one embodiment, the complex protease comprises papain and alkaline protease in a mass ratio of 1 (1-3).
In one embodiment, each mL of the oyster sauce is inoculated with 2x10 6 CFU～4x10 6 CFU of said lactic acid bacteria.
In one embodiment, the conditions of the first fermentation include: the fermentation temperature is 35-39 ℃ and the fermentation time is 24-48 h.
In one embodiment, the Saccharomyces cerevisiae embryo is prepared by:
and activating Saccharomyces cerevisiae by taking glucose water as an activating solution.
In one embodiment, the mass percentage concentration of glucose in the glucose water is 10% -15%.
In one embodiment, each mL of the oyster sauce is inoculated with 2x10 8 CFU～5x10 8 CFU said s.cerevisiae embryo.
In one embodiment, the conditions of the second fermentation include: the fermentation temperature is 43-47 ℃ and the fermentation time is 24-36 h.
In one embodiment, the means for collecting fermentation broth from the first fermentation product comprises centrifugation.
In one embodiment, the conditions of centrifugation include: the rotating speed is 6000 rpm-8000 rpm, and the time is 10 min-15 min.
In one embodiment, the means for collecting fermentation broth from the second fermentation product comprises centrifugation.
In one embodiment, the conditions of centrifugation include: the rotating speed is 6000 rpm-8000 rpm, and the time is 5 min-8 min.
In one embodiment, after the step of collecting fermentation broth from the second fermentation product, the steps of:
adjusting the pH of the fermentation broth collected from the second fermentation product to a target isoelectric point, filtering, collecting the filtrate and drying.
In one embodiment, the pH of the target isoelectric point is between 7.0 and 7.5.
An embodiment of the present application further provides an oyster polysaccharide extract, which is prepared by the method for preparing an oyster polysaccharide extract according to any one of the embodiments.
According to the preparation method of the oyster polysaccharide extract, amylase and compound protease are adopted to carry out enzymolysis on oyster pulp, then lactic acid bacteria and saccharomyces cerevisiae are sequentially adopted to carry out fermentation on the obtained enzymolysis liquid, in the enzymolysis process, protein in oyster is decomposed into amino acid to the maximum extent, then the amino acid is converted into acetic acid through lactic acid bacteria fermentation, and then the saccharomyces cerevisiae is utilized to carry out fermentation, and the acetic acid is further converted into substances such as glucose, polysaccharide and the like, so that the extraction degree of polysaccharide in oyster is greatly improved, and the oyster polysaccharide extract has a good polysaccharide extraction effect even on oyster raw materials with low sugar content.
Drawings
FIG. 1 is a glucose standard curve for colorimetrically determining polysaccharide extraction.
Detailed Description
In order that the application may be readily understood, a more particular description of the application will be rendered by reference to specific embodiments that are illustrated in the appended drawings. This application may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The terminology used herein in the description of the application is for the purpose of describing particular embodiments only and is not intended to be limiting of the application. The term "and/or" as used herein includes any and all combinations of one or more of the associated listed items.
In the application, the technical characteristics described in an open mode comprise a closed technical scheme composed of the listed characteristics and also comprise an open technical scheme comprising the listed characteristics.
An embodiment of the application provides a preparation method of oyster polysaccharide extract, comprising the following steps:
mixing the primarily processed oyster meat with water, pulping, and preparing oyster pulp;
adding amylase and compound protease into oyster pulp for enzymolysis, and inactivating enzyme to prepare an enzymolysis solution;
inoculating lactobacillus into the enzymolysis liquid to perform first fermentation to prepare a first fermentation product, and collecting fermentation liquid from the first fermentation product to prepare a first supernatant;
inoculating Saccharomyces cerevisiae embryo into the first supernatant for secondary fermentation to prepare a secondary fermentation product, and collecting fermentation liquor from the secondary fermentation product to prepare oyster polysaccharide extract.
The primary processed oyster meat can be, for example, but not limited to, oyster raw materials subjected to primary processing such as blanching, water filtering and the like, and the sugar content of the primary processed oyster meat is significantly lower than that of fresh open oyster meat. According to the preparation method of the oyster polysaccharide extract, amylase and compound protease are adopted to carry out enzymolysis on oyster pulp, then lactic acid bacteria and saccharomyces cerevisiae are sequentially adopted to carry out fermentation on the obtained enzymolysis liquid, in the enzymolysis process, protein in oyster is decomposed into amino acid to the maximum extent, then the amino acid is converted into acetic acid through lactic acid bacteria fermentation, and then the saccharomyces cerevisiae is utilized to carry out fermentation, and the acetic acid is further converted into substances such as glucose, polysaccharide and the like, so that the extraction degree of polysaccharide in oyster is greatly improved, and the oyster polysaccharide extract has a good polysaccharide extraction effect even on oyster raw materials with low sugar content.
In one embodiment, the mass ratio of the raw oyster meat to the water is 3 (1.5-2.5). The ratio of the feed liquid to the water is adopted for the primary processing oyster meat, so that the primary processing oyster meat has a good rehydration effect and is favorable for making oyster paste. It will be appreciated that the mass ratio of the as-processed oyster meat to water may be, for example, but not limited to, 3:1.5, 3:2, 3:2.5, etc. It can be appreciated that in the process of preparing oyster pulp, in order to make oyster meat finer and more uniform in pulping, the pre-processed oyster meat can also be subjected to cutting treatment.
In one embodiment, the conditions for enzymolysis include: the pH value is 7.0-8.0, the enzymolysis temperature is 50-60 ℃, and the enzymolysis time is 8-12 h. Before amylase and compound protease are added, the pH value of oyster pulp is adjusted to 7.0-8.0, so that the enzyme preparation can perform enzymolysis in the optimal environment, and the enzymolysis efficiency is improved; meanwhile, the enzymolysis conditions are controlled to be carried out at a proper temperature and time, so that the better enzymolysis effect can be ensured, the protein in the oyster can be decomposed maximally, and the microbial pollution caused in the enzymolysis process can be reduced as much as possible. It will be appreciated that the pH may be adjusted to, for example, but not limited to, 7.0, 7.2, 7.5, 7.7, 7.9, 8.0, etc. It will be appreciated that the enzymatic hydrolysis temperature may be, for example, but not limited to, 50 ℃, 53 ℃, 55 ℃, 58 ℃, 60 ℃, and the like. It is understood that the enzymolysis time may be, for example, but not limited to, 8h, 9h, 10h, 11h, 12h, etc.
Further, the step of adjusting the pH of the oyster sauce comprises the following steps: naOH is added to the oyster slurry. Preferably, naOH is food grade NaOH.
In one example, the amylase is used in an amount of 2g to 5g per kg of oyster syrup calculated as 60000U/g of enzyme activity. It will be appreciated that the amount of amylase used is 600000U/g of enzyme activity, and the corresponding amount of addition per kg of oyster syrup may be, for example, but not limited to, 2g, 3g, 4g, 5g, etc.
In one embodiment, the amount of the compound protease is 2 g-10 g per kg oyster sauce calculated by 600000U/g enzyme activity. The amount of the compound protease to be used is 600000U/g of enzyme activity, and the amount of the compound protease to be added per kg of oyster syrup may be, for example, but not limited to, 2g, 4g, 6g, 8g, 10g, etc.
In one embodiment, the complex protease comprises papain and alkaline protease.
Further, the mass ratio of the papain to the alkaline protease is 1 (1-3). It will be appreciated that the mass ratio of papain to alkaline protease may be, for example, but not limited to, 1:1, 1:1.5, 1:2, 1:2.5, 1:3, etc.
In one embodiment, the step of inactivating the enzyme comprises: adding edible salt, and heating in boiling water bath.
Further, the mass of the edible salt is 13-17% of the mass of the oyster sauce. It will be appreciated that the mass of the edible salt may be, for example, but not limited to, 13%, 14%, 15%, 16%, 17% of the mass of the oyster sauce, etc.
Further, the boiling water bath is heated for 20 min-40 min. It is understood that the time of boiling water bath heating may be, for example, but not limited to, 20 minutes, 25 minutes, 30 minutes, 35 minutes, 40 minutes, and the like.
In one embodiment, 2X10 inoculations are performed per mL of oyster sauce 6 CFU～5x10 6 Lactic acid bacteria of CFU. The inoculation amount of the lactobacillus is controlled within a specific range, so that the sufficient dosage of the lactobacillus can be ensured, amino acid in the enzymolysis liquid can be converted into acetic acid as much as possible, and meanwhile, the influence of insufficient nutritional ingredients on fermentation efficiency caused by excessive dosage can be avoided. It will be appreciated that the inoculum size of lactic acid bacteria inoculated per ml oyster sauce can be, for example, but not limited to, 2X10 6 CFU、3x10 6 CFU、4x10 6 CFU、5x10 6 CFU, etc.
In one embodiment, the conditions of the first fermentation include: the fermentation temperature is 35-39 ℃ and the fermentation time is 24-48 h. The first fermentation is carried out at a proper temperature for a proper time, so that the fermentation effect is good, the lactobacillus is promoted to convert amino acid in the enzymolysis liquid into acetic acid, and the time for expanding cultivation can be shortened. It is understood that the fermentation temperature of the first fermentation may be, for example, but not limited to, 35 ℃, 36 ℃, 37 ℃, 38 ℃, 39 ℃, and the like. It is understood that the fermentation time of the first fermentation may be, for example, but not limited to, 24h, 28h, 30h, 33h, 36h, 40h, 44h, 46h, 48h, etc.
In one embodiment, the means for collecting fermentation broth from the first fermentation product comprises centrifugation.
Further, the conditions of centrifugation include: the rotating speed is 6000 rpm-8000 rpm, and the time is 10 min-15 min. The first fermentation product is sticky, can have a good centrifugal separation effect by adopting a higher rotating speed and a longer time for separation, and can remove substances which are not utilized in the enzymolysis liquid, such as fat and the like, by carrying out centrifugal separation on the first fermentation product, thereby being beneficial to improving the fermentation efficiency in the subsequent second fermentation process. It will be appreciated that the centrifugation speed may be, for example, but not limited to, 6000rpm, 7000rpm, 8000rpm, etc., and the time may be, for example, but not limited to, 10min, 11min, 12min, 13min, 14min, 15min, etc.
In one embodiment, the Saccharomyces cerevisiae embryo is prepared by: and activating Saccharomyces cerevisiae by taking glucose water as an activating solution. The glucose water is adopted to activate the saccharomyces cerevisiae, so that the saccharomyces cerevisiae embryo can generate fermentation more quickly after being added into the first supernatant.
Further, the mass percentage concentration of glucose in glucose water is 10% -15%. By adopting glucose water with proper concentration as the activating solution, the saccharomyces cerevisiae has better activating effect, and can avoid adverse effect on the activity of the yeast caused by too high osmotic pressure due to too high concentration of the glucose water, and further, avoid the problem that the polysaccharide content in the final oyster polysaccharide extract cannot be accurately detected due to the fact that the oyster is introduced or exogenous is introduced due to the fact that too much glucose is introduced.
In one embodiment, 2X10 inoculations are performed per mL of oyster sauce 8 CFU～5x10 8 CFU s.cerevisiae embryos. The inoculation amount of the saccharomyces cerevisiae embryo is controlled in a proper range, so that acetic acid obtained by decomposition in the first supernatant can be further and fully converted into glucose, and the oyster polysaccharide can be effectively extracted. It will be appreciated that the amount of inoculated Saccharomyces cerevisiae embryos per ml of oyster slurry may be, for example, but not limited to, 2X10 per kg of oyster slurry 8 CFU、4x10 8 CFU、5x10 8 CFU, etc.
In one embodiment, the conditions of the second fermentation include: the fermentation temperature is 43-47 ℃ and the fermentation time is 24-36 h. The second fermentation is carried out at a proper temperature for a proper time, so that the problem that the polysaccharide content in the extract is reduced due to further consumption of sugar generated by the original fermentation due to insufficient nutrition substrate caused by overlong fermentation time is avoided while good fermentation effect is ensured. It will be appreciated that the fermentation temperature of the second fermentation may be, for example, but not limited to, 43 ℃, 44 ℃, 45 ℃, 46 ℃, 47 ℃, etc., and the fermentation time may be, for example, but not limited to, 24h, 28h, 30h, 32h, 34h, 36h, etc.
In one embodiment, the means for collecting fermentation broth from the second fermentation product comprises centrifugation.
Further, the conditions of centrifugation include: the rotating speed is 6000 rpm-8000 rpm, and the time is 5 min-8 min. By centrifuging the second fermentation product, insoluble fermentation residues can be separated for subsequent purification of oyster polysaccharide extract. It will be appreciated that the second fermentation product may be subjected to a centrifugation speed of, for example, but not limited to, 6000rpm, 7000rpm, 8000rpm, etc., and a time of, for example, but not limited to, 5min, 6min, 7min, 8min, etc.
In one embodiment, the step of collecting fermentation broth from the second fermentation product further comprises the steps of:
adjusting the pH of the fermentation broth collected from the second fermentation product to a target isoelectric point, filtering, collecting the filtrate and drying. The fermentation broth is adjusted to the target isoelectric point, and insoluble fermentation residues are more easily separated.
Further, the pH of the target isoelectric point is 7.0 to 7.5. It is understood that the pH of the target isoelectric point may be, for example, but not limited to, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, and the like.
An embodiment of the present application further provides an oyster polysaccharide extract, which is prepared by the method for preparing an oyster polysaccharide extract in any one of the above embodiments.
The oyster polysaccharide extract provided by the application maximally utilizes protein in oyster, and amino acid obtained by decomposing protein in enzymolysis liquid is further gradually converted into polysaccharide by adopting a mode of fermenting and decomposing microorganisms by adopting lactobacillus and saccharomycetes in sequence, so that the extraction effect of oyster polysaccharide is improved.
An embodiment of the present application further provides an application of the preparation method of the oyster polysaccharide extract in any one of the above embodiments in preparation of oyster juice products. According to the method provided by the application, even if oyster polysaccharide extracts are not independently used as industrial products, the method can be applied to improve the polysaccharide content in oyster juice and other products, so that the sweet and fragrant taste of the oyster juice products can be improved.
The oyster polysaccharide extract according to the present application and the method for preparing the same are described in further detail below by way of specific examples. The following embodiments are more specific, and it is understood that in other embodiments, this is not limiting. In the following examples, the instruments, reagents and materials involved, unless otherwise specified, are conventional instruments, reagents and materials already known in the art and are commercially available. The experimental methods, detection methods, and the like in the examples described below are conventional experimental methods and detection methods known in the prior art unless otherwise specified.
Example 1
A preparation method of oyster polysaccharide extract comprises the following steps:
(1) The mass ratio of the primary processed oyster meat to water is 3:2, mixing and pulping according to the proportion to prepare oyster pulp;
(2) Regulating the pH of oyster pulp to 7.5, adding amylase, papain and alkaline protease into the oyster pulp, wherein the mass of the amylase, the papain and the alkaline protease is calculated by 600000U/g enzyme activity, the corresponding addition amount of each kg of oyster pulp is 2g, 2g and 6g respectively, carrying out enzymolysis for 10 hours at the temperature of 55 ℃, adding edible salt into the product after the enzymolysis is finished, heating the product in boiling water bath for 30 minutes to carry out enzyme deactivation treatment, and preparing an enzymolysis liquid;
(4) Inoculating lactobacillus into the enzymolysis solution, wherein the inoculation amount of lactobacillus is 2x10 per mL oyster pulp 6 CFU, culturing for 24 hours at 37 ℃ in a shaking table to perform first fermentation, and preparing a first fermentation product;
(5) Centrifuging the first fermentation product at 6000rpm for 15min, and filtering to collect fermentation liquor to obtain a first supernatant;
(6) Activating Saccharomyces cerevisiae with glucose water with glucose concentration of 10% by mass, preparing Saccharomyces cerevisiae embryo, inoculating Saccharomyces cerevisiae embryo into the first supernatant, and adding 2×10 of Saccharomyces cerevisiae embryo per mL oyster slurry 8 CFU, culturing for 36h under 45 ℃ shaking table to perform second fermentation, and preparing a second fermentation product;
(7) Centrifuging the second fermentation product at 6000rpm for 8min, collecting the upper fermentation liquid, adjusting pH of the fermentation liquid to 7.0, filtering with 0.22 μm filter membrane to remove impurities, collecting filtrate, and drying to obtain oyster polysaccharide extract.
Example 2
A preparation method of oyster polysaccharide extract comprises the following steps:
(1) The mass ratio of the primary processed oyster meat to water is 3:2, mixing and pulping according to the proportion to prepare oyster pulp;
(2) Regulating the pH of oyster pulp to 7.5, adding amylase, papain and alkaline protease into the oyster pulp, wherein the mass of the amylase, the papain and the alkaline protease is calculated by 600000U/g enzyme activity, the corresponding addition amount of each kg of oyster pulp is 2g, 2g and 6g respectively, carrying out enzymolysis for 12 hours at 50 ℃, adding edible salt into the product after the enzymolysis is finished, heating the product in boiling water bath for 30min for enzyme deactivation, and preparing an enzymolysis liquid;
(4) Inoculating lactobacillus into the enzymolysis solution, and adding 4×10 lactobacillus into each mL oyster slurry 6 CFU, culturing for 48h at 37 ℃ in a shaking table to perform first fermentation, and preparing a first fermentation product;
(5) Centrifuging the first fermentation product at 6000rpm for 15min, and filtering to collect fermentation liquor to obtain a first supernatant;
(6) Activating Saccharomyces cerevisiae with glucose water with 15% of glucose mass percentage concentration to prepare Saccharomyces cerevisiae embryo, inoculating Saccharomyces cerevisiae embryo into the first supernatant, adding 2x10 of the inoculating amount of Saccharomyces cerevisiae embryo into each mL oyster slurry 8 CFU, culturing for 36h under 45 ℃ shaking table to perform second fermentation, and preparing a second fermentation product;
(7) Centrifuging the second fermentation product at 8000rpm for 5min, collecting fermentation liquor, adjusting pH of the fermentation liquor to 7.5, filtering with 0.22 μm filter membrane to remove impurities, collecting filtrate, and drying to obtain oyster polysaccharide extract.
Example 3
A preparation method of oyster polysaccharide extract comprises the following steps:
(1) The mass ratio of the primary processed oyster meat to water is 3:2, mixing and pulping according to the proportion to prepare oyster pulp;
(2) Adjusting the pH of oyster pulp to 8.0, adding amylase, papain and alkaline protease into the oyster pulp, wherein the mass of the amylase, the papain and the alkaline protease is calculated by 600000U/g enzyme activity, the corresponding addition amount of each kg of oyster pulp is 5g, 2g and 3g respectively, carrying out enzymolysis for 8 hours at the temperature of 60 ℃, adding edible salt into the product after the enzymolysis is finished, heating the product in boiling water bath for 30min for enzyme deactivation, and preparing an enzymolysis liquid;
(4) Inoculating lactobacillus into the enzymolysis solution, wherein the inoculation amount of lactobacillus is 3x10 per mL oyster pulp 6 CFU, culturing for 36h under a shaking table at 37 ℃ to perform first fermentation, and preparing a first fermentation product;
(5) Centrifuging the first fermentation product at 8000rpm for 10min, and filtering to collect fermentation liquor to obtain a first supernatant;
(6) Activating Saccharomyces cerevisiae with glucose water with 12% glucose mass percentage concentration to prepare Saccharomyces cerevisiae embryo, inoculating Saccharomyces cerevisiae embryo into the first supernatant, adding 5x10 per mL oyster slurry into the Saccharomyces cerevisiae embryo 8 CFU, culturing for 24 hours at 45 ℃ in a shaking table to perform second fermentation, and preparing a second fermentation product;
(7) Centrifuging the second fermentation product at 7000rpm for 8min, collecting the upper fermentation broth, adjusting pH of the fermentation broth to 7.2, filtering with 0.22 μm filter membrane to remove impurities, collecting filtrate, and drying to obtain oyster polysaccharide extract.
Example 4
A preparation method of oyster polysaccharide extract comprises the following steps:
(1) The mass ratio of the primary processed oyster meat to water is 3:2, mixing and pulping according to the proportion to prepare oyster pulp;
(2) Regulating the pH of oyster pulp to 7.0, adding amylase, papain and alkaline protease into the oyster pulp, wherein the mass of the amylase, the papain and the alkaline protease is calculated by 600000U/g enzyme activity, the corresponding addition amount of each kg of oyster pulp is 3g, 5g and 5g respectively, carrying out enzymolysis for 12 hours at 50 ℃, adding edible salt into the product after the enzymolysis is finished, heating the product in boiling water bath for 30min for enzyme deactivation, and preparing an enzymolysis liquid;
(4) Inoculating lactobacillus into the enzymolysis solution, wherein the inoculation amount of lactobacillus is 4x10 per mL oyster pulp 6 CFU, culturing for 48h at 37 ℃ in a shaking table to perform first fermentation, and preparing a first fermentation product;
(5) Centrifuging the first fermentation product at 6000rpm for 15min, and filtering to collect fermentation liquor to obtain a first supernatant;
(6) Activating Saccharomyces cerevisiae with glucose water with 15% of glucose mass percentage concentration to prepare Saccharomyces cerevisiae embryo, inoculating Saccharomyces cerevisiae embryo into the first supernatant, adding 2x10 of the inoculating amount of Saccharomyces cerevisiae embryo into each mL oyster slurry 8 CFU, culturing for 36h under 45 ℃ shaking table to perform second fermentation, and preparing a second fermentation product;
(7) Centrifuging the second fermentation product at 8000rpm for 5min, collecting the upper fermentation broth, adjusting pH of the fermentation broth to 7.5, filtering with 0.22 μm filter membrane to remove impurities, collecting filtrate, and drying to obtain oyster polysaccharide extract.
Example 5
A preparation method of oyster polysaccharide extract comprises the following steps:
(1) The mass ratio of the primary processed oyster meat to water is 3:2, mixing and pulping according to the proportion to prepare oyster pulp;
(2) Adjusting the pH of oyster pulp to 7.5, adding amylase, papain and alkaline protease into the oyster pulp, wherein the mass of the amylase, the papain and the alkaline protease is calculated by 600000U/g enzyme activity, the corresponding addition amount of each kg of oyster pulp is 5g, 1g and 2g respectively, carrying out enzymolysis for 12 hours at 50 ℃, adding edible salt into the product after the enzymolysis is finished, heating the product in boiling water bath for 30min for enzyme deactivation, and preparing an enzymolysis liquid;
(4) Inoculating lactobacillus into the enzymolysis solution, wherein the inoculation amount of lactobacillus is 4x10 per mL oyster pulp 6 CFU, culturing for 48h at 37 ℃ in a shaking table to perform first fermentation, and preparing a first fermentation product;
(5) Centrifuging the first fermentation product at 6000rpm for 15min, and filtering to collect fermentation liquor to obtain a first supernatant;
(6) Activating Saccharomyces cerevisiae with glucose water with 15% of glucose mass percentage concentration to prepare Saccharomyces cerevisiae embryo, inoculating Saccharomyces cerevisiae embryo into the first supernatant, adding 2x10 of the inoculating amount of Saccharomyces cerevisiae embryo into each mL oyster slurry 8 CFU, culturing at 45deg.C for 36h for the second timeFermenting to prepare a second fermentation product;
(7) Centrifuging the second fermentation product at 8000rpm for 5min, collecting the upper fermentation broth, adjusting pH of the fermentation broth to 7.5, filtering with 0.22 μm filter membrane to remove impurities, collecting filtrate, and drying to obtain oyster polysaccharide extract.
Comparative example 1
Substantially the same as in example 2, except that: the pH is not adjusted before enzymolysis.
Comparative example 2:
substantially the same as in example 2, except that: no amylase was added.
Comparative example 3:
substantially the same as in example 2, except that: the first fermentation was performed without inoculation with lactic acid bacteria. Specifically, the preparation steps are adjusted as follows:
(1) And (3): the same as in (1) to (3) of example 2;
(4) Centrifuging the enzymolysis liquid at 6000rpm for 15min, and filtering and collecting the supernatant to obtain a first supernatant;
(5) And (7): the same as in (6) to (8) of example 2.
Comparative example 4:
substantially the same as in example 2, except that: the second fermentation was performed without inoculating Saccharomyces cerevisiae embryos. Specifically, the preparation steps are adjusted as follows:
(1) And (4): the same as in (1) to (4) of example 2;
(5) Centrifuging the first fermentation product at 8000rpm for 5min, and collecting fermentation liquor to obtain a first supernatant;
(6) The pH of the first supernatant was adjusted to 7.5, and the oyster polysaccharide extract was prepared by filtering and removing impurities with a 0.22 μm filter membrane, collecting the filtrate and drying.
Comparative example 5
Substantially the same as in example 2, except that: (4) conditions under which the first fermentation is performed are different; (6) conditions under which the second fermentation is carried out are different. Specifically, steps (4) and (6) are adjusted to:
(4) Inoculating lactobacillus into the enzymolysis liquid, wherein the inoculation amount of the lactobacillus is that1x10 of oyster paste is added into each mL 6 CFU, culturing for 60h under a shaking table at 37 ℃ to perform first fermentation, and preparing a first fermentation product;
(6) Activating Saccharomyces cerevisiae with glucose water with glucose concentration of 20% by mass, preparing Saccharomyces cerevisiae embryo, inoculating Saccharomyces cerevisiae embryo into the first supernatant, and adding 6×10 of Saccharomyces cerevisiae embryo into each mL oyster slurry 8 CFU, cultured for 50h at 45 ℃ in a shaker to perform a second fermentation, to prepare a second fermentation product.
Polysaccharide extraction rate tests were performed on the raw oyster meat materials and the oyster polysaccharide extracts prepared in examples 1 to 5 and comparative examples 1 to 5, by using a colorimetric method, and by using a glucose standard curve for test as shown in fig. 1, and the test results are shown in table 1 below.
TABLE 1 polysaccharide extraction ratio of raw oyster meat material to oyster polysaccharide extract
As can be seen from Table 1, the polysaccharide extraction rates of the oyster polysaccharide extracts prepared in examples 1 to 5 were 11.2% as low as the initial oyster meat material, and were significantly higher than those of the oyster polysaccharide extracts of comparative examples 1 to 5.
The technical features of the above-described embodiments may be arbitrarily combined, and all possible combinations of the technical features in the above-described embodiments are not described for brevity of description, however, as long as there is no contradiction between the combinations of the technical features, they should be considered as the scope of the description.
The above examples illustrate only a few embodiments of the application, which are described in detail and are not to be construed as limiting the scope of the application. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the application, which are all within the scope of the application. Accordingly, the scope of protection of the present application is to be determined by the appended claims.
Claims (10)
1. A preparation method of oyster polysaccharide extract is characterized by comprising the following steps:
mixing the primarily processed oyster meat with water, pulping, and preparing oyster pulp;
adding amylase and compound protease into the oyster pulp for enzymolysis, and inactivating enzyme to prepare an enzymolysis solution;
inoculating lactobacillus into the enzymolysis liquid to perform first fermentation to prepare a first fermentation product, and collecting fermentation liquid from the first fermentation product to prepare a first supernatant;
inoculating saccharomyces cerevisiae embryo into the first supernatant to perform second fermentation to prepare a second fermentation product, and collecting fermentation liquor from the second fermentation product to prepare oyster polysaccharide extract.
2. The method for preparing oyster polysaccharide extract according to claim 1, wherein the conditions of enzymolysis include:
the pH value is 7.0-8.0, the enzymolysis temperature is 50-60 ℃, and the enzymolysis time is 8-12 h;
optionally, the usage amount of the amylase is calculated by 60000U/g enzyme activity, and the corresponding addition amount of each kg of oyster slurry is 2 g-5 g;
optionally, the usage amount of the compound protease is calculated by 600000U/g enzyme activity, and the corresponding addition amount of each kg of oyster slurry is 2 g-10 g;
optionally, the compound protease comprises papain and alkaline protease with a mass ratio of (1-3).
3. The method for preparing oyster polysaccharide extract according to claim 1, wherein each mL of oyster paste is inoculated with 2x10 6 CFU～4x10 6 CFU of said lactic acid bacteria.
4. The method for preparing oyster polysaccharide extract according to claim 1, wherein the conditions of the first fermentation include: the fermentation temperature is 35-39 ℃ and the fermentation time is 24-48 h.
5. The method for preparing oyster polysaccharide extract according to claim 1, wherein the saccharomyces cerevisiae embryo is prepared by the following method:
activating Saccharomyces cerevisiae by taking glucose water as an activating solution;
optionally, the mass percentage concentration of glucose in the glucose water is 10% -15%.
6. The method for preparing oyster polysaccharide extract according to claim 1, wherein each mL of oyster paste is inoculated with 2x10 8 CFU～5x10 8 CFU said s.cerevisiae embryo.
7. The method for preparing oyster polysaccharide extract according to claim 1, wherein the conditions of the second fermentation include: the fermentation temperature is 43-47 ℃ and the fermentation time is 24-36 h.
8. The method for preparing oyster polysaccharide extract according to any one of claims 1 to 7, wherein the means for collecting fermentation broth from the first fermentation product comprises centrifugation; optionally, the conditions of centrifugation include: the rotating speed is 6000 rpm-8000 rpm, and the time is 10 min-15 min;
or/and the combination of the two,
means for collecting fermentation broth from the second fermentation product include centrifugation; optionally, the conditions of centrifugation include: the rotating speed is 6000 rpm-8000 rpm, and the time is 5 min-8 min.
9. The method for preparing oyster polysaccharide extract according to any one of claims 1 to 7, further comprising the steps of, after the step of collecting the fermentation broth from the second fermentation product:
adjusting the pH of the fermentation broth collected from the second fermentation product to a target isoelectric point, filtering, collecting filtrate and drying;
optionally, the pH of the target isoelectric point is 7.0 to 7.5.
10. An oyster polysaccharide extract, characterized in that it is prepared by the method for preparing an oyster polysaccharide extract according to any one of claims 1 to 9.
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