CN113621535A - Composite protective agent for improving survival rate of probiotic freeze-dried powder and preparation method thereof - Google Patents
Composite protective agent for improving survival rate of probiotic freeze-dried powder and preparation method thereof Download PDFInfo
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- CN113621535A CN113621535A CN202110792603.XA CN202110792603A CN113621535A CN 113621535 A CN113621535 A CN 113621535A CN 202110792603 A CN202110792603 A CN 202110792603A CN 113621535 A CN113621535 A CN 113621535A
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- 239000006041 probiotic Substances 0.000 title claims abstract description 69
- 235000018291 probiotics Nutrition 0.000 title claims abstract description 69
- 239000003223 protective agent Substances 0.000 title claims abstract description 56
- 239000000843 powder Substances 0.000 title claims abstract description 50
- 230000000529 probiotic effect Effects 0.000 title claims abstract description 50
- 239000002131 composite material Substances 0.000 title claims abstract description 28
- 230000004083 survival effect Effects 0.000 title claims abstract description 27
- 238000002360 preparation method Methods 0.000 title claims abstract description 10
- 244000005700 microbiome Species 0.000 claims abstract description 61
- 150000004676 glycans Chemical class 0.000 claims abstract description 57
- 229920001282 polysaccharide Polymers 0.000 claims abstract description 57
- 239000005017 polysaccharide Substances 0.000 claims abstract description 57
- 241000756042 Polygonatum Species 0.000 claims abstract description 46
- 239000000725 suspension Substances 0.000 claims abstract description 41
- 230000001580 bacterial effect Effects 0.000 claims abstract description 32
- 238000007710 freezing Methods 0.000 claims abstract description 24
- 239000001963 growth medium Substances 0.000 claims abstract description 24
- 230000008014 freezing Effects 0.000 claims abstract description 22
- 239000010802 sludge Substances 0.000 claims abstract description 20
- 238000002156 mixing Methods 0.000 claims abstract description 16
- 230000004913 activation Effects 0.000 claims abstract description 11
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 claims abstract description 10
- 108010010803 Gelatin Proteins 0.000 claims abstract description 10
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims abstract description 10
- 229930006000 Sucrose Natural products 0.000 claims abstract description 10
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims abstract description 10
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 claims abstract description 10
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 claims abstract description 10
- 229920000159 gelatin Polymers 0.000 claims abstract description 10
- 239000008273 gelatin Substances 0.000 claims abstract description 10
- 235000019322 gelatine Nutrition 0.000 claims abstract description 10
- 235000011852 gelatine desserts Nutrition 0.000 claims abstract description 10
- 239000008103 glucose Substances 0.000 claims abstract description 10
- 235000020183 skimmed milk Nutrition 0.000 claims abstract description 10
- 239000005720 sucrose Substances 0.000 claims abstract description 10
- 238000000855 fermentation Methods 0.000 claims abstract description 9
- 230000004151 fermentation Effects 0.000 claims abstract description 9
- 239000006228 supernatant Substances 0.000 claims abstract description 9
- 230000000813 microbial effect Effects 0.000 claims abstract description 8
- 238000009629 microbiological culture Methods 0.000 claims abstract description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 32
- OAKJQQAXSVQMHS-UHFFFAOYSA-N Hydrazine Chemical compound NN OAKJQQAXSVQMHS-UHFFFAOYSA-N 0.000 claims description 16
- 239000000284 extract Substances 0.000 claims description 8
- 238000000605 extraction Methods 0.000 claims description 8
- 238000001914 filtration Methods 0.000 claims description 8
- 238000000643 oven drying Methods 0.000 claims description 8
- 238000010298 pulverizing process Methods 0.000 claims description 8
- 238000007873 sieving Methods 0.000 claims description 8
- 238000005119 centrifugation Methods 0.000 claims description 4
- 239000002994 raw material Substances 0.000 claims description 3
- 238000001035 drying Methods 0.000 claims description 2
- 239000000047 product Substances 0.000 abstract description 8
- 230000000694 effects Effects 0.000 abstract description 5
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- 239000003814 drug Substances 0.000 abstract description 2
- 150000001875 compounds Chemical class 0.000 description 17
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- 210000004027 cell Anatomy 0.000 description 7
- 241000894006 Bacteria Species 0.000 description 6
- 238000013329 compounding Methods 0.000 description 6
- 238000009630 liquid culture Methods 0.000 description 6
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- 238000009777 vacuum freeze-drying Methods 0.000 description 6
- 210000002390 cell membrane structure Anatomy 0.000 description 2
- 230000003204 osmotic effect Effects 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 241000186015 Bifidobacterium longum subsp. infantis Species 0.000 description 1
- 241000186660 Lactobacillus Species 0.000 description 1
- 235000013960 Lactobacillus bulgaricus Nutrition 0.000 description 1
- 244000199885 Lactobacillus bulgaricus Species 0.000 description 1
- 241001468157 Lactobacillus johnsonii Species 0.000 description 1
- 240000006024 Lactobacillus plantarum Species 0.000 description 1
- 235000013965 Lactobacillus plantarum Nutrition 0.000 description 1
- 241000194035 Lactococcus lactis Species 0.000 description 1
- 241000037831 Polygonatum sibiricum Species 0.000 description 1
- 235000014897 Streptococcus lactis Nutrition 0.000 description 1
- 241000194020 Streptococcus thermophilus Species 0.000 description 1
- 239000006286 aqueous extract Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229940004120 bifidobacterium infantis Drugs 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 229940039696 lactobacillus Drugs 0.000 description 1
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- 229940072205 lactobacillus plantarum Drugs 0.000 description 1
- 239000008176 lyophilized powder Substances 0.000 description 1
- 230000010534 mechanism of action Effects 0.000 description 1
- 229940126680 traditional chinese medicines Drugs 0.000 description 1
- 235000013618 yogurt Nutrition 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/04—Preserving or maintaining viable microorganisms
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Abstract
The invention discloses a composite protective agent for improving the survival rate of probiotic freeze-dried powder and a preparation method thereof, wherein the composite protective agent comprises the following components: 1-10 parts of skimmed milk powder, 1-10 parts of trehalose, 1-10 parts of sucrose, 1-10 parts of glucose, 1-10 parts of gelatin and 1-10 parts of polygonatum polysaccharide; the preparation method of the probiotic freeze-dried powder comprises the following steps: s1: inoculating probiotic strains into a microbial culture medium, putting the culture medium into a shaking table for fermentation, culture and activation, and collecting activated microbial suspension; s2: centrifuging the microorganism suspension in the S1, removing supernatant, and collecting bacterial sludge; s3: adding a composite protective agent into the bacterial sludge, and uniformly mixing the probiotic suspension; s4: and pre-freezing the microbial suspension in the S3, and then performing vacuum freezing to obtain the probiotic freeze-dried powder. The invention improves the survival rate of the probiotic freeze-dried powder, and simultaneously utilizes the tonifying effect of the rhizoma polygonati which is a dual-purpose resource of medicine and food to increase the nourishing effect of the probiotic freeze-dried powder product.
Description
Technical Field
The invention relates to the technical field of traditional Chinese medicines, in particular to a composite protective agent for improving the survival rate of probiotic freeze-dried powder and a preparation method thereof.
Background
For the existing probiotic products, the problem of low survival rate of probiotics exists, so that the effectiveness of the probiotic products is seriously inhibited, and the technical problem to be solved by the invention is how to improve the survival rate of the probiotics in the probiotic products.
Disclosure of Invention
Based on the technical problems in the background art, the invention provides the composite protective agent for improving the survival rate of the probiotic freeze-dried powder and the preparation method thereof, and the composite protective agent improves the survival rate of the probiotic freeze-dried powder, and simultaneously utilizes the tonifying effect of the polygonatum polysaccharide which is a dual-purpose resource of medicine and food, so as to improve the nourishing effect of the probiotic freeze-dried powder product.
The invention provides a composite protective agent for improving the survival rate of probiotic freeze-dried powder, which comprises the following raw materials in parts by weight: 1-10 parts of skimmed milk powder, 1-10 parts of trehalose, 1-10 parts of sucrose, 1-10 parts of glucose, 1-10 parts of gelatin and 1-10 parts of polygonatum polysaccharide.
Preferably, the polygonatum polysaccharide is an aqueous extract, and the preparation method comprises the following steps: oven drying rhizoma Polygonati, pulverizing, sieving, mixing with water, extracting at high temperature, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution.
Preferably, the drying conditions are as follows: the temperature is 50-70 ℃ and the time is 2-6 h.
Preferably, the conditions of the high-temperature extraction are as follows: the mass volume ratio of rhizoma Polygonati to water is 1:5-15, the temperature is 90-125 deg.C, and the time is 0.5-2 h.
The invention provides application of a composite protective agent for improving the survival rate of probiotic freeze-dried powder in the probiotic freeze-dried powder.
Preferably, the preparation method of the probiotic freeze-dried powder comprises the following steps:
s1: inoculating probiotic strains into a microbial culture medium, putting the culture medium into a shaking table for fermentation, culture and activation, and collecting activated microbial suspension;
s2: centrifuging the microorganism suspension in the S1, removing supernatant, and collecting bacterial sludge;
s3: adding a composite protective agent into the bacterial sludge, and uniformly mixing the probiotic suspension;
s4: and pre-freezing the microbial suspension in the S3, and then performing vacuum freezing to obtain the probiotic freeze-dried powder.
Preferably, the conditions for activation in S1 are: the temperature is 30-38 ℃, and the time is 12-48 h.
Preferably, the centrifugation time in S2 is 1-10min, and the rotation speed during centrifugation is 8000-12000 rpm.
Preferably, the mass ratio of the bacterial sludge in the S3 to the composite protective agent is 1: 1-5.
Preferably, the pre-freezing conditions in S4 are: the temperature is-20 to-4 ℃, and the time is 10 to 24 hours; the vacuum freezing conditions are as follows: the vacuum degree is 0.1-20Pa, the temperature of the cold hydrazine is-180 to-45 ℃, and the time is 12-24 h.
Mechanism of action
The protective agent prepared by taking polygonatum polysaccharide as a raw material is mixed with the probiotic bacteria liquid, wherein the polygonatum polysaccharide can be combined with probiotic bacteria cells to protect the cell membrane structure, maintain stable cell osmotic pressure and protect the protein structure in the probiotic bacteria cells to be stable, so that the survival rate of the probiotic bacteria in the probiotic bacteria product is improved.
Compared with the prior art, the invention has the beneficial technical effects
The compound protective agent comprises polygonatum polysaccharide which can be combined with probiotic cells, so that the survival rate of probiotics is improved, and compared with probiotics without polygonatum polysaccharide, the survival rate of probiotics is improved by more than 10%.
Drawings
Fig. 1 is a microscopic image of probiotics from example 1 (left) and its control group (right) as proposed by the present invention;
fig. 2 is a microscopic image of probiotics from example 2 (left) and its control group (right) proposed by the present invention;
fig. 3 is a microscopic image of probiotics from example 3 (left) and its control group (right) as proposed by the present invention;
fig. 4 is a microscopic image of probiotics from example 4 (left) and its control group (right) as proposed by the present invention;
fig. 5 is a microscopic image of probiotics from example 5 (left) and its control group (right) as proposed by the present invention;
fig. 6 is a microscopic image of probiotics from example 6 (left) and its control group (right) proposed by the present invention;
Detailed Description
The skim milk powder of the present invention was purchased from Australian Yoghurt (China) Co., Ltd, trehalose was purchased from Nippon Co., Ltd, sucrose was purchased from Shanghai-derived leaf Biotech Co., Ltd, glucose was purchased from Kangyao Chemicals Co., Ltd, and gelatin was purchased from Biotech (Shanghai) Co., Ltd.
The above products are all commercially available, and are used directly without any treatment.
Example 1
The method for compounding the compound protective agent and the lactobacillus plantarum provided by the invention comprises the following steps:
(1) inoculating microorganisms into a microorganism liquid culture medium according to the volume ratio of the bacterial liquid to the microorganism culture medium of 1:100, then placing the microorganism culture medium containing the bacterial liquid in a shaking table for fermentation culture and activation at 30 ℃, and collecting microorganism suspension after 12 h.
(2) Centrifuging the microorganism suspension in a high-speed centrifuge with 12000rpm for 1min, discarding supernatant, and collecting bacterial sludge.
(3) Adding the compound protective agent and the bacterial sludge according to the volume ratio of 1:1, wherein the formula and the parts by weight of the compound protective agent are 1 part of skimmed milk powder, 1 part of trehalose, 1 part of sucrose, 1 part of glucose, 1 part of gelatin and 1 part of polygonatum polysaccharide, and uniformly mixing to obtain the microorganism suspension containing the polygonatum polysaccharide protective agent.
(4) Pre-freezing the microorganism suspension containing the polygonatum polysaccharide and the protective agent at-4 ℃ for-12 h, then placing the microorganism suspension in a vacuum freeze-drying machine for freezing for 24h to obtain the microorganism freeze-dried powder, wherein the vacuum freezing conditions are as follows: the vacuum degree is 20Pa, the temperature of the cold hydrazine is-180 ℃, and the time is 24 h.
The extraction method of the polygonatum polysaccharide comprises the following steps:
oven drying rhizoma Polygonati at 50 deg.C for 2 hr, pulverizing, sieving, mixing with water, extracting at 90 deg.C for 0.5 hr, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution. Wherein the mass volume ratio of rhizoma polygonati to water is 1: 5. Concentrating the obtained water extract to make the concentration of rhizoma Polygonati polysaccharide 0.5 g/L.
Example 2
The method for compounding the composite protective agent and the lactobacillus bulgaricus comprises the following steps:
(1) inoculating microorganisms into a microorganism liquid culture medium according to the volume ratio of the bacterial liquid to the microorganism culture medium of 1:100, then placing the microorganism culture medium containing the bacterial liquid in a shaking table for fermentation culture and activation at 37 ℃, and collecting microorganism suspension after 24 hours.
(2) Centrifuging the microorganism suspension in a 8000rpm high-speed centrifuge for 5min, discarding the supernatant, and collecting bacterial sludge.
(3) Adding the compound protective agent and the bacterial sludge according to the volume ratio of 1:3, wherein the formula and the parts by weight of the compound protective agent are 3 parts of skimmed milk powder, 3 parts of trehalose, 1 part of sucrose, 5 parts of glucose, 3 parts of gelatin and 5 parts of polygonatum polysaccharide, and uniformly mixing to obtain the microorganism suspension containing the polygonatum polysaccharide protective agent.
(4) Prefreezing the microorganism suspension containing the polygonatum polysaccharide protective agent at-4 ℃ for-12 h, and then putting the prefreezed microorganism suspension into a vacuum freeze-drying machine for freezing for 24h to obtain polygonatum microorganism freeze-dried powder, wherein the vacuum freezing conditions are as follows: the vacuum degree is 0.1Pa, the temperature of the cold hydrazine is-45 ℃ and the time is 12 h.
The extraction method of the polygonatum polysaccharide comprises the following steps:
oven drying rhizoma Polygonati at 60 deg.C for 4 hr, pulverizing, sieving, mixing with water, extracting at 108 deg.C for 1 hr, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution. Wherein the mass volume ratio of rhizoma polygonati to water is 1: 10. Concentrating the obtained water extract to make the concentration of rhizoma Polygonati polysaccharide 0.5 g/L.
Example 3
The method for compounding the compound protective agent and the lactobacillus johnsonii comprises the following steps:
(1) inoculating microorganisms into a microorganism liquid culture medium according to the volume ratio of the bacterial liquid to the microorganism culture medium of 1:100, then placing the microorganism culture medium containing the bacterial liquid in a shaking table for fermentation culture and activation at 37 ℃, and collecting microorganism suspension after 48 hours.
(2) Centrifuging the microorganism suspension in a high-speed centrifuge with 120000rpm for 1min, discarding the supernatant, and collecting bacterial sludge.
(3) The microbial suspension containing the polygonatum polysaccharide protective agent is prepared by adding the compound protective agent and the bacterial sludge according to the volume ratio of 1:1, wherein the formula and the parts by weight of the compound protective agent are 5 parts of skimmed milk powder, 10 parts of trehalose, 10 parts of sucrose, 2 parts of glucose, 3 parts of gelatin and 10 parts of polygonatum polysaccharide which are uniformly mixed.
(4) Pre-freezing microorganism suspension containing rhizoma Polygonati polysaccharide and protective agent at-4 deg.C for-12 h, and freezing in vacuum freeze-drying machine for 24h to obtain rhizoma Polygonati microorganism lyophilized powder, wherein the vacuum freezing conditions are as follows: 10Pa, the temperature of cold hydrazine is-90 ℃ and the time is 18 h.
The extraction method of the polygonatum polysaccharide comprises the following steps:
oven drying rhizoma Polygonati at 70 deg.C for 6 hr, pulverizing, sieving, mixing with water, extracting at 125 deg.C for 2 hr, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution. Wherein the mass volume ratio of rhizoma polygonati to water is 1: 15. Concentrating the obtained water extract to make the concentration of rhizoma Polygonati polysaccharide 0.5 g/L.
Example 4
The method for compounding the composite protective agent and the lactococcus lactis provided by the invention comprises the following steps:
(1) inoculating microorganisms into a microorganism liquid culture medium according to the volume ratio of the bacterial liquid to the microorganism culture medium of 1:100, then placing the microorganism culture medium containing the bacterial liquid in a shaking table for fermentation culture and activation at 37 ℃, and collecting microorganism suspension after 24 hours.
(2) Centrifuging the microorganism suspension in a 8000rpm high-speed centrifuge for 5min, discarding the supernatant, and collecting bacterial sludge.
(3) Adding the compound protective agent and the bacterial sludge according to the volume ratio of 1:3, wherein the formula and the parts by weight of the compound protective agent are 3 parts of skimmed milk powder, 3 parts of trehalose, 1 part of sucrose, 5 parts of glucose, 3 parts of gelatin and 5 parts of polygonatum polysaccharide, and uniformly mixing to obtain the microorganism suspension containing the polygonatum polysaccharide protective agent.
(4) Prefreezing the microorganism suspension containing the polygonatum polysaccharide protective agent at-4 ℃ for-12 h, and then putting the prefreezed microorganism suspension into a vacuum freeze-drying machine for freezing for 24h to obtain polygonatum microorganism freeze-dried powder, wherein the vacuum freezing conditions are as follows: the vacuum degree is 5Pa, the temperature of the cold hydrazine is-60 ℃, and the time is 18 h.
The extraction method of the polygonatum polysaccharide comprises the following steps:
oven drying rhizoma Polygonati at 50 deg.C for 4 hr, pulverizing, sieving, mixing with water, extracting at 98 deg.C for 1.5 hr, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution. Wherein the mass volume ratio of rhizoma polygonati to water is 1: 8. Concentrating the obtained water extract to make the concentration of rhizoma Polygonati polysaccharide 0.5 g/L.
Example 5
The method for compounding the compound protective agent and the bifidobacterium infantis comprises the following steps:
(1) inoculating microorganisms into a microorganism liquid culture medium according to the volume ratio of the bacterial liquid to the microorganism culture medium of 1:100, then placing the microorganism culture medium containing the bacterial liquid in a shaking table for fermentation culture and activation at 37 ℃, and collecting microorganism suspension after 24 hours.
(2) Centrifuging the microorganism suspension in a 8000rpm high-speed centrifuge for 5min, discarding the supernatant, and collecting bacterial sludge.
(3) Adding the compound protective agent and the bacterial sludge according to the volume ratio of 1:3, wherein the formula and the parts by weight of the compound protective agent are 3 parts of skimmed milk powder, 3 parts of trehalose, 1 part of sucrose, 5 parts of glucose, 3 parts of gelatin and 5 parts of polygonatum polysaccharide, and uniformly mixing to obtain the microorganism suspension containing the polygonatum polysaccharide protective agent.
(4) Prefreezing the microorganism suspension containing the polygonatum polysaccharide protective agent at-4 ℃ for-12 h, and then putting the prefreezed microorganism suspension into a vacuum freeze-drying machine for freezing for 24h to obtain polygonatum microorganism freeze-dried powder, wherein the vacuum freezing conditions are as follows: the vacuum degree is 0.1Pa, the temperature of the cold hydrazine is-45 ℃ and the time is 12 h.
The extraction method of the polygonatum polysaccharide comprises the following steps:
oven drying rhizoma Polygonati at 62 deg.C for 5 hr, pulverizing, sieving, mixing with water, extracting at 115 deg.C for 1.5 hr, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution. Wherein the mass volume ratio of rhizoma polygonati to water is 1: 12. Concentrating the obtained water extract to make the concentration of rhizoma Polygonati polysaccharide 0.5 g/L.
Example 6
The method for compounding the compound protective agent and the streptococcus thermophilus provided by the invention comprises the following steps:
(1) inoculating microorganisms into a microorganism liquid culture medium according to the volume ratio of the bacterial liquid to the microorganism culture medium of 1:100, then placing the microorganism culture medium containing the bacterial liquid in a shaking table for fermentation culture and activation at 37 ℃, and collecting microorganism suspension after 24 hours.
(2) Centrifuging the microorganism suspension in a 8000rpm high-speed centrifuge for 5min, discarding the supernatant, and collecting bacterial sludge.
(3) Adding the compound protective agent and the bacterial sludge according to the volume ratio of 1:3, wherein the formula and the parts by weight of the compound protective agent are 3 parts of skimmed milk powder, 3 parts of trehalose, 1 part of sucrose, 5 parts of glucose, 3 parts of gelatin and 5 parts of polygonatum polysaccharide, and uniformly mixing to obtain the microorganism suspension containing the polygonatum polysaccharide protective agent.
(4) Prefreezing the microorganism suspension containing the polygonatum polysaccharide protective agent at-4 ℃ for-12 h, and then putting the prefreezed microorganism suspension into a vacuum freeze-drying machine for freezing for 24h to obtain polygonatum microorganism freeze-dried powder, wherein the vacuum freezing conditions are as follows: the vacuum degree is 0.1Pa, the temperature of the cold hydrazine is-45 ℃ and the time is 12 h.
The extraction method of the polygonatum polysaccharide comprises the following steps:
oven drying rhizoma Polygonati at 60 deg.C for 5 hr, pulverizing, sieving, mixing with water, extracting at 120 deg.C for 1.5 hr, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution. Wherein the mass volume ratio of rhizoma polygonati to water is 1: 10. Concentrating the obtained water extract to make the concentration of rhizoma Polygonati polysaccharide 0.5 g/L.
The viable count of the probiotics in the polygonatum sibiricum microbial freeze-dried powder prepared in the examples 1 to 6 is detected and microscopically observed according to the food safety national standard food microbiology test lactobacillus (GB 4789.35-2010), and meanwhile, no polygonatum polysaccharide is added as a control group for each experimental group, and the conditions of the control group and the corresponding experimental group are different in that no polygonatum polysaccharide is added, and the other conditions are the same as the experimental group, and the results are shown in table 1.
TABLE 1 detection results of viable count of probiotic bacteria
As can be seen from table 1, the survival rate of the probiotics added with the polygonatum polysaccharide composite protective agent is increased by more than 10% compared with the survival rate of the probiotics without the polygonatum polysaccharide composite protective agent, because the polygonatum polysaccharide can be combined with the probiotics cells to protect the cell membrane structure, maintain stable cell osmotic pressure, and protect the protein structure in the probiotics cells to be stable, thereby increasing the survival rate of the probiotics in the probiotics product. In addition, for protecting probiotics by using polygonatum polysaccharides, the polygonatum polysaccharide water extract can be used alone, and polygonatum polysaccharides in other forms can also be used, and the scheme is also in the protection scope of the invention.
Claims (10)
1. The composite protective agent for improving the survival rate of the probiotic freeze-dried powder is characterized by comprising the following raw materials in parts by weight: 1-10 parts of skimmed milk powder, 1-10 parts of trehalose, 1-10 parts of sucrose, 1-10 parts of glucose, 1-10 parts of gelatin and 1-10 parts of polygonatum polysaccharide.
2. The composite protective agent for improving the survival rate of probiotic freeze-dried powder according to claim 1, wherein the polygonatum polysaccharide is polygonatum polysaccharide water extract, and the preparation method comprises the following steps: oven drying rhizoma Polygonati, pulverizing, sieving, mixing with water, extracting at high temperature, and filtering to obtain rhizoma Polygonati polysaccharide water extractive solution.
3. The composite protective agent for improving the survival rate of probiotic freeze-dried powder according to claim 2, wherein the drying conditions are as follows: the temperature is 50-70 ℃ and the time is 2-6 h.
4. The composite protective agent for improving the survival rate of probiotic freeze-dried powder according to claim 2, characterized in that the conditions of high-temperature extraction are as follows: the mass volume ratio of rhizoma Polygonati to water is 1:5-15, the temperature is 90-125 deg.C, and the time is 0.5-2 h.
5. The use of the composite protectant for improving the survival rate of probiotic freeze-dried powder according to any one of claims 1 to 4 in probiotic freeze-dried powder.
6. The application of the composite protective agent for improving the survival rate of the probiotic freeze-dried powder in the probiotic freeze-dried powder according to claim 5, wherein the preparation method of the probiotic freeze-dried powder comprises the following steps:
s1: inoculating probiotic strains into a microbial culture medium, putting the culture medium into a shaking table for fermentation, culture and activation, and collecting activated microbial suspension;
s2: centrifuging the microorganism suspension in the S1, removing supernatant, and collecting bacterial sludge;
s3: adding a composite protective agent into the bacterial sludge, and uniformly mixing to obtain a probiotic suspension;
s4: and pre-freezing the microbial suspension in the S3, and then performing vacuum freezing to obtain the probiotic freeze-dried powder.
7. The application of the composite protective agent for improving the survival rate of probiotic freeze-dried powder in the probiotic freeze-dried powder according to claim 6, wherein the activation condition in S1 is as follows: the temperature is 30-38 ℃, and the time is 12-48 h.
8. The use of the composite protectant for improving the survival rate of probiotic freeze-dried powder in probiotic freeze-dried powder according to claim 6, wherein the centrifugation time in S2 is 1-10min, and the rotation speed during centrifugation is 8000-12000 rpm.
9. The application of the composite protective agent for improving the survival rate of probiotic freeze-dried powder in the probiotic freeze-dried powder according to claim 6 is characterized in that the mass ratio of the bacterial sludge in S3 to the composite protective agent is 1: 1-5.
10. The application of the composite protective agent for improving the survival rate of probiotic freeze-dried powder in the probiotic freeze-dried powder according to claim 6, wherein the pre-freezing condition in S4 is as follows: the temperature is-20 to-4 ℃, and the time is 10 to 24 hours; the vacuum freezing conditions are as follows: the vacuum degree is 0.1-20Pa, the temperature of the cold hydrazine is-180 to-45 ℃, and the time is 12-24 h.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN202110792603.XA CN113621535B (en) | 2021-07-14 | 2021-07-14 | Composite protective agent for improving survival rate of probiotics freeze-dried powder and preparation method thereof |
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