CN113355254B - Formula and application of screwdriver protective agent - Google Patents
Formula and application of screwdriver protective agent Download PDFInfo
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- CN113355254B CN113355254B CN202010311191.9A CN202010311191A CN113355254B CN 113355254 B CN113355254 B CN 113355254B CN 202010311191 A CN202010311191 A CN 202010311191A CN 113355254 B CN113355254 B CN 113355254B
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- trehalose
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- 239000003223 protective agent Substances 0.000 title description 9
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims abstract description 45
- 239000007858 starting material Substances 0.000 claims abstract description 41
- 239000000203 mixture Substances 0.000 claims abstract description 29
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 claims abstract description 28
- 238000009472 formulation Methods 0.000 claims abstract description 27
- 239000000843 powder Substances 0.000 claims abstract description 25
- 235000013923 monosodium glutamate Nutrition 0.000 claims abstract description 18
- 235000020183 skimmed milk Nutrition 0.000 claims abstract description 18
- 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 17
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 claims abstract description 17
- 229930195725 Mannitol Natural products 0.000 claims abstract description 17
- 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 17
- 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 17
- 239000000594 mannitol Substances 0.000 claims abstract description 17
- 235000010355 mannitol Nutrition 0.000 claims abstract description 17
- 229940073490 sodium glutamate Drugs 0.000 claims abstract description 15
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 claims abstract description 14
- 229930003268 Vitamin C Natural products 0.000 claims abstract description 14
- 235000019154 vitamin C Nutrition 0.000 claims abstract description 14
- 239000011718 vitamin C Substances 0.000 claims abstract description 14
- 230000001012 protector Effects 0.000 claims abstract description 11
- PXEDJBXQKAGXNJ-QTNFYWBSSA-L disodium L-glutamate Chemical compound [Na+].[Na+].[O-]C(=O)[C@@H](N)CCC([O-])=O PXEDJBXQKAGXNJ-QTNFYWBSSA-L 0.000 claims abstract 12
- 241000589776 Pseudomonas putida Species 0.000 claims description 21
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims description 19
- 239000012535 impurity Substances 0.000 claims 1
- 238000000746 purification Methods 0.000 claims 1
- 230000001681 protective effect Effects 0.000 description 23
- 241000894006 Bacteria Species 0.000 description 22
- 239000007788 liquid Substances 0.000 description 21
- 230000004083 survival effect Effects 0.000 description 17
- 238000012258 culturing Methods 0.000 description 14
- 230000000052 comparative effect Effects 0.000 description 12
- 230000001580 bacterial effect Effects 0.000 description 10
- 238000011534 incubation Methods 0.000 description 7
- 238000002360 preparation method Methods 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 6
- 238000004108 freeze drying Methods 0.000 description 6
- LPUQAYUQRXPFSQ-DFWYDOINSA-M monosodium L-glutamate Chemical compound [Na+].[O-]C(=O)[C@@H](N)CCC(O)=O LPUQAYUQRXPFSQ-DFWYDOINSA-M 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 240000006024 Lactobacillus plantarum Species 0.000 description 5
- 235000013965 Lactobacillus plantarum Nutrition 0.000 description 5
- 229940072205 lactobacillus plantarum Drugs 0.000 description 5
- 239000010802 sludge Substances 0.000 description 5
- 239000007864 aqueous solution Substances 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- 239000004471 Glycine Substances 0.000 description 3
- 241000589516 Pseudomonas Species 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 239000003124 biologic agent Substances 0.000 description 3
- 238000000855 fermentation Methods 0.000 description 3
- 230000004151 fermentation Effects 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 238000004321 preservation Methods 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 238000013207 serial dilution Methods 0.000 description 3
- 238000003756 stirring Methods 0.000 description 3
- 229920001817 Agar Polymers 0.000 description 2
- 229920002261 Corn starch Polymers 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- 229910019142 PO4 Inorganic materials 0.000 description 2
- 229930006000 Sucrose Natural products 0.000 description 2
- 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 description 2
- 230000002378 acidificating effect Effects 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- 239000008120 corn starch Substances 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 239000004223 monosodium glutamate Substances 0.000 description 2
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 2
- 235000019799 monosodium phosphate Nutrition 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 2
- 239000010452 phosphate Substances 0.000 description 2
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 2
- CIJQGPVMMRXSQW-UHFFFAOYSA-M sodium;2-aminoacetic acid;hydroxide Chemical compound O.[Na+].NCC([O-])=O CIJQGPVMMRXSQW-UHFFFAOYSA-M 0.000 description 2
- 239000005720 sucrose Substances 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 230000035899 viability Effects 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- FXXMDJFRMDVSCF-RXSVEWSESA-N (2r)-2-[(1s)-1,2-dihydroxyethyl]-3,4-dihydroxy-2h-furan-5-one;hydrate Chemical compound O.OC[C@H](O)[C@H]1OC(=O)C(O)=C1O FXXMDJFRMDVSCF-RXSVEWSESA-N 0.000 description 1
- 102000011632 Caseins Human genes 0.000 description 1
- 108010076119 Caseins Proteins 0.000 description 1
- 241000235342 Saccharomycetes Species 0.000 description 1
- HVUMOYIDDBPOLL-XWVZOOPGSA-N Sorbitan monostearate Chemical compound CCCCCCCCCCCCCCCCCC(=O)OC[C@@H](O)[C@H]1OC[C@H](O)[C@H]1O HVUMOYIDDBPOLL-XWVZOOPGSA-N 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- HVUMOYIDDBPOLL-XGKPLOKHSA-N [2-[(2r,3r,4s)-3,4-dihydroxyoxolan-2-yl]-2-hydroxyethyl] octadecanoate Chemical compound CCCCCCCCCCCCCCCCCC(=O)OCC(O)[C@H]1OC[C@H](O)[C@H]1O HVUMOYIDDBPOLL-XGKPLOKHSA-N 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 235000013405 beer Nutrition 0.000 description 1
- 235000008429 bread Nutrition 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 239000012526 feed medium Substances 0.000 description 1
- 235000021001 fermented dairy product Nutrition 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 238000007602 hot air drying Methods 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
- 235000013379 molasses Nutrition 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 210000000496 pancreas Anatomy 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 238000013112 stability test Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 239000012137 tryptone Substances 0.000 description 1
- 235000013618 yogurt Nutrition 0.000 description 1
Classifications
-
- 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
-
- 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
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Wood Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Tropical Medicine & Parasitology (AREA)
- Mycology (AREA)
- Botany (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention relates to a starter protector formula and application thereof. A starter protectant formulation comprising: 1.125+ -10% to 4.5+ -10% trehalose by weight; 2.25+ -10% to 4.5+ -10% of skimmed milk powder; mannitol in an amount of 1.125.+ -. 10% to 2.25.+ -. 10%; 0.0225 + -10% to 0.1125 + -10% by weight of vitamin C;0.225 + -10% to 1.125 + -10% by weight of sodium glutamate; and 0.225.+ -. 10% to 1.125.+ -. 10% by weight of glycerol.
Description
Technical Field
The present invention relates to a starter protector formulation, and more particularly to a starter protector formulation suitable for both gram positive and gram negative bacteria.
Background
The number of bacteria of natural microorganisms is usually not as large as that required for practical use, and it is also very difficult to preserve them, and they are often subjected to environmental stimulus or mutation after multiple generations of culture, so they are often used and circulated in the form of "starter" after hot air drying or freeze drying. In the case of the food industry, the use of natural bacteria cannot be directly carried out when fermentation using microorganisms is required in the process because of the large operating volume. Often, the domesticated screwdriver is used for expanding cultivation in advance, and the subsequent process is performed after the bacterial count is enough. Practical industrial applications include industries where yeast is required as a raw material, such as: fermentation of beer and baking of bread. In addition, if applied to the biological agent industry of agriculture, the produced biological agent product also has a high requirement for stable maintenance of viable bacteria count, so that the biological agent product is also suitable for prior expanding culture by a starter. Therefore, after the development of the potential strain, a starter formulation is needed which can achieve the purpose of high stability and high viable count, thereby increasing the flexibility of application in the process. Meanwhile, the thalli can be sold in a bottle opener mode, so that the product innovation speed and the diversity are increased.
CN106244459a discloses a pseudomonas dry powder and a preparation method thereof, the pseudomonas dry powder comprises pseudomonas and a protective agent, wherein the protective agent comprises skimmed milk powder, mannitol, sodium glutamate or glycine, corn starch and trehalose.
CN107828683a discloses a lactobacillus plantarum freeze-dried powder for prolonging the shelf life of yogurt and a preparation method, which prepares the lactobacillus plantarum freeze-dried powder by mixing lactobacillus plantarum strains with an emulsion protecting agent and freeze-drying, wherein the lactobacillus plantarum strains are preserved in a China general microbiological culture collection center (CGMCC), and the preservation number is CGMCC No. 10453. The preparation method comprises the steps of strain activation, strain propagation, fermentation culture and freeze-drying. The lactobacillus plantarum freeze-dried powder has an inhibiting effect on main spoilage moulds and yeasts in the fermented dairy product.
The existing starter protectant formula has the defect of low viable count during culture, and particularly when the starter protectant formula is used after a period of time after preparation is completed, the viable count is greatly reduced.
Disclosure of Invention
The main object of the present invention is to provide a starter protector formulation which can improve the disadvantages of low viable count and survival rate, in particular a starter protector formulation with improved stability which after a period of storage still has a comparable viable count as compared to the as-prepared starter protector formulation.
A starter protector formulation according to the present invention comprises:
1.125.+ -. 10% trehalose by weight;
2.25+/-10% of skimmed milk powder in parts by weight;
1.125.+ -. 10% mannitol by weight;
0.0225 + -10% by weight of vitamin C;
0.225.+ -. 10% sodium glutamate; and
0.225.+ -. 10% By weight of glycerol.
Other preferred embodiments of the invention include, but are not limited to, those described in the following claims.
The starter protectant formulation of the present invention has a feature of improved viable count and survival rate, whether used for the cultivation of gram positive or gram negative bacteria.
Drawings
FIG. 1 shows the number of viable bacteria after culturing Pseudomonas putida starter prepared with each of the protective liquid groups in Table 1 of example 1 of the present invention at 30℃for 72 hours.
Fig. 2 to 4 show the viable count of yeasts 21849, 20405, 22745 each prepared by the respective protective liquid groups in table 1 of example 1 of the present invention after culturing the starter at 25℃for 72 hours.
FIG. 5 shows the number of viable bacteria after culturing the Pseudomonas putida starter prepared in each of the protective liquid groups in Table 2 and Table 3 of comparative example 1 at 30℃for 72 hours.
FIG. 6 shows the viable count of yeasts 21849, 20405, 22745, which were prepared in each of the protective liquid groups in Table 2 and Table 3 of comparative example 1, after culturing at 25℃for 72 hours.
FIG. 7 shows the survival rate of Pseudomonas putida drivers prepared with each of the protective liquid groups in Table 1 of example 1 of the present invention after incubation at 30℃for 72 hours.
Fig. 8 to 10 show the survival rates of yeasts 21849, 20405, 22745 starter prepared in each of the protective liquid groups in table 1 of example 1 of the present invention, respectively, after 72 hours of incubation at 25 ℃.
Fig. 11 shows the survival rate of pseudomonas putida starter prepared with each of the protective liquid groups in table 2 and table 3 of comparative example 1 after culturing at 30 ℃ for 72 hours.
FIG. 12 shows the survival rate of yeasts 21849, 20405, 22745, which were prepared in the respective protective liquid groups of Table 2 and Table 3 of comparative example 1, after culturing at 25℃for 72 hours.
FIG. 13 shows the number of viable bacteria of Pseudomonas putida starter prepared with each of the protective liquid groups in Table 1 of example 1 of the present invention and after storage at 25℃and 54℃for 14 days and incubation at 30℃for 72 hours.
Detailed Description
The present invention and its improvements will be further understood by the following examples and comparative examples. The following materials and instruments were used in the following examples and comparative examples.
Material
| Skimmed milk powder | MERCK |
| Casein protein | MERCK |
| Glycerol | SIGMA |
| Vitamin C | SIGMA |
| Trehalose | SIGMA |
| Sucrose | SIGMA |
| Sodium glutamate (monosodium glutamate) | Taste whole |
| Span 60 | SIGMA |
| YM Broth | Gibico |
Pseu F agar culture medium formula
| Tryptone 1% | MERCK |
| Pancreas protein 1% | MERCK |
| Dipotassium hydrogen phosphate (K 2HPO4) 0.15% | SIGMA |
| Magnesium sulfate (MgSO 4) 0.15% | SIGMA |
| Agar (Agar) | SIGMA |
Instrument for measuring and controlling the intensity of light
Pseudomonas putida culture mode
Pseudomonas putida YLW01 (Pseudomonas putida) was selected as the experimental strain. YLW01 was cultured in medium (50 mL medium/250 mL Erlenmeyer flask) containing 1% Molasses, 1% MSG, 1.5 g/L K 2HPO4 and 1.5 g/L MgSO 4 .7H2 O and 42 hr was cultured at 30℃in 150 rpm. After the incubation was completed, centrifugation was performed at 8000 rpm for 10 minutes at 4℃and the supernatant was discarded, washed with water and centrifuged again, and this washing was repeated twice. And taking out the centrifuged bacterial sludge and placing the bacterial sludge in a refrigerator for preservation.
Yeast culture mode
Yeast 21849, 20405, 22745 were selected as experimental strains. Yeast 21849, 20405, 22745 were cultured in YM broth (Becton Dickinson and Company) medium (50 mL medium/250 mL Erlenmeyer flask) and 48 hr were cultured at 25℃and 250 rpm. After the incubation was completed, centrifugation was performed at 8000 rpm for 10 minutes at 4℃and the supernatant was discarded, washed with water and centrifuged again, and this washing was repeated twice. And taking out the centrifuged bacterial sludge and placing the bacterial sludge in a refrigerator for preservation.
Formulation of protective agent formulation/protective liquid
The protective solution was prepared by dissolving the protective agent formulation of table 1 in 22.5 mL of water and stirring for 30 minutes until the solution was uniform, and then pH was adjusted to 7 or 8 with the following phosphate solution (alkaline to acidic) or glycine-sodium hydroxide buffer (acidic to alkaline).
Table 1: protective agent formula (g)/protective solution (g/vol%)
Glycine-sodium hydroxide buffer solution:
50 The mL glycine aqueous solution (0.2M) was mixed with 8.8 mL sodium hydroxide aqueous solution (0.2M) and quantified to 200 mL.
Phosphate solution preparation:
87.7 The mL of the sodium dihydrogen phosphate aqueous solution (0.2M) and the 12.3 mL sodium dihydrogen phosphate aqueous solution (0.2M) were mixed uniformly and then quantified to 200 mL.
Example 1
Preparation of screwdriver
Taking out the prepared bacterial mud from the refrigerator, respectively mixing 7.5 g bacterial mud with each protection liquid in table 1, stirring for 30 minutes until the bacterial mud is uniformly mixed, pre-freezing at-20 ℃, and freeze-drying to obtain the starter.
Taking a freeze-dried bottle opener which is just prepared, dissolving the bottle opener 1g back into 10mL water, carrying out serial dilution with proper multiplying power, taking 100 mu L bottle opener diluent to be smeared on a Pseu F culture medium, placing the bottle opener on the 30 ℃ for 72 hours, and carrying out colony counting.
Comparative example 1
The formulation of the drivers was carried out with the formulations of CN106244459 and CN107828683 described above. The protection solutions 22.5 mL in Table 2 and Table 3 were mixed with the bacterial sludge 7.5 g prepared as described above, and after stirring for 30 minutes until the mixture was uniform, the mixture was pre-frozen at-20℃and freeze-dried to obtain a starter of comparative example 1.
The freeze-dried bottle opener which is just prepared is taken out, 1g bottle opener is back dissolved into 10mL water, serial dilution with proper multiplying power is carried out, 100 mu L bottle opener diluent is smeared on Pseu F culture medium, and the bottle opener is placed at 30 ℃ for 72 hours for culturing, and then bacterial colony counting is carried out.
TABLE 2 CN106244459 protective liquid (g/vol%)
| Group of | Trehalose (%) | Skimmed milk powder (%) | Mannitol (%) | Yeast powder (%) | Monosodium glutamate (%) | Span60(%) | Glycine (%) | Corn starch (%) |
| CN106244459-1 | 1 | 25 | 0.5 | 1 | 3 | 2 | 3 | 0.1 |
| CN106244459-2 | 5 | 12 | 5 | 3 | 0 | 1 | 15 | 2 |
TABLE 3 NC 107828683 protective solution (g/vol%)
| Group of | Trehalose (%) | Skimmed milk powder (%) | Glycerol (%) | Mannitol (%) | Sucrose (%) | Ascorbic acid | Water and its preparation method |
| CN107828683 | 6 | 8 | 1 | 2 | 4 | 0.5 | 78.5 |
Results
FIG. 1 shows the number of viable bacteria after culturing Pseudomonas putida starter prepared with each of the protective liquid groups in Table 1 of example 1 at 30℃for 72 hours. In FIG. 1, the number of the living Pseudomonas putida is maintained above 10 9 CFU/g, which shows that the protective agent of the invention has good protective effect. Wherein the highest viable count of the group 2 reaches 6.3x10 9 CFU/g.
Fig. 2 to 4 show the viable count of yeasts 21849, 20405, 22745 each prepared in the respective protective liquid groups of table 1 of example 1 after culturing at 25℃for 72 hours. The highest viable count of yeasts 21849, 20405, 22745 were 3.44x10 9 CFU/g (FIG. 2, panel 5), 3.75x10 9 CFU/g (FIG. 3, panel 7) and 5x10 9 CFU/g (FIG. 4, panel 3), respectively.
FIG. 5 shows the number of viable bacteria after culturing the Pseudomonas putida starter prepared in each of the protective liquid groups in Table 2 and Table 3 of comparative example 1 at 30℃for 72 hours. The viable count of Pseudomonas putida was found to be 5.8X10 7、8.8x108 and 2.6X10 8 CFU/g in FIG. 5, respectively.
FIG. 6 shows the viable count of yeasts 21849, 20405, 22745, which were prepared in each of the protective liquid groups in Table 2 and Table 3 of comparative example 1, after culturing at 25℃for 72 hours. The culture effect is poor, most of viable bacteria count is 10 8 CFU/g, the group of the highest viable bacteria count is the formula of CN106244459-2 used by saccharomycete 22745, and the number reaches 1.99x10 9 CFU/g.
Fig. 7 shows the survival rate of pseudomonas putida starter prepared with each of the protective liquid groups in table 1 of example 1 after incubation at 30 ℃ for 72 hours. The survival rate calculation method is as follows:
Survival rate (%) = [ viable count after lyophilization (CFU)/number of added bacteria before lyophilization (CFU) ] ×100%
In FIG. 7, it was found that the number of Pseudomonas putida viable bacteria was maintained above 15%, with the highest survival rate of about 78% for group 2. The viability of each group in FIG. 7 is similar to that of FIG. 1.
Fig. 8 to 10 show the survival rates of yeasts 21849, 20405, 22745 starter prepared in each of the protection liquid groups of table 1 of example 1, respectively, after culturing at 25 ℃ for 72 hours. The survival rates of yeasts 21849, 20405, 22745 were up to 53.22% (FIG. 8, panel 3), 47.2% (FIG. 9, panel 3) and 47.41% (FIG. 10, panel 3), respectively.
Fig. 11 shows the survival rate of pseudomonas putida starter prepared with each of the protective liquid groups in table 2 and table 3 of comparative example 1 after culturing at 30 ℃ for 72 hours. The survival rates of Pseudomonas putida were found to be 0.07%, 1.02% and 0.3%, respectively, in FIG. 11.
FIG. 12 shows the survival rate of yeasts 21849, 20405, 22745, which were prepared in the respective protective liquid groups of Table 2 and Table 3 of comparative example 1, after culturing at 25℃for 72 hours. FIG. 12 shows the same trend in viability as the viable count of FIG. 6, around 2%, with the highest group being CN106244459-1 cultured on yeast 20405.
From the above figures 1 to 12, it can be seen that the number of viable bacteria and the survival rate of the formulation of the present invention were significantly improved compared to the formulations CN106244459-1, CN106244459-2 and CN107828683 by testing the formulations of the present invention and the formulations of the previous cases with pseudomonas putida and 3 yeasts (21849, 20405, 22745). The invention has been shown to have unexpected boosting efficacy.
Example 2: freeze-dried starter stability test
The freeze-dried starter prepared in example 1 was stored at 25℃and 54℃for 14 days, then 1 g starter was dissolved back into 10 mL water, serial dilutions were made at appropriate magnification, 100. Mu.L of Pseudomonas putida starter dilution was smeared on Pseu F medium, and after incubation at 30℃for 72 hours, colony counts were performed.
The results are shown in FIG. 13, in which the viable count of the freeze-dried starter just prepared is shown at the same time. FIG. 13 shows that the number of viable bacteria in most formulation groups was significantly reduced and the maximum reduction in group 2 was observed after the Pseudomonas putida starter prepared with each of the protective liquid groups of Table 1 of example 1 of the present invention was stored at 25℃for 14 days and then incubated at 30℃for 72 hours, but the number of viable bacteria was maintained at 2X 10 8 CFU/g. The decrease in the number of viable bacteria of group 4 was minimal, and the number of viable bacteria reached 1.2X10- 9 CFU/g, and the survival rate was maintained at 87% or more (not shown). As a result of the culture at 54℃for 14 days in FIG. 13, it was found that the number of viable bacteria was greatly reduced, the number of viable bacteria in most groups was reduced to 10 7 CFU/g or less, and only groups 3, 4 and 7 remained at 10 7 CFU/g or more, with the highest number of viable bacteria in group 7 reaching 6.2X 10 7 CFU/g. The freeze-dried bottle opener has good stability, and can maintain the viable count of more than 2 x 10 8 CFU/g after being stored for 14 days at 25 ℃, and the viable count is lower than that of the freeze-dried bottle opener which is just prepared by about one logarithmic value.
Claims (8)
1. A starter protectant formulation, which consists of the following components:
1.125+ -10% to 4.5+ -10% trehalose by weight;
2.25+ -10% to 4.5+ -10% of skimmed milk powder;
mannitol in an amount of 1.125.+ -. 10% to 2.25.+ -. 10%;
0.0225 + -10% to 0.1125 + -10% by weight of vitamin C;
0.225 + -10% to 1.125 + -10% by weight of sodium glutamate; and
0.225.+ -. 10% To 1.125.+ -. 10% by weight of glycerol.
2. The starter protectant formulation of claim 1, consisting of the following components: 1.125+ -5% to 4.5+ -5% trehalose by weight;
2.25+ -5% to 4.5+ -5% of skimmed milk powder;
mannitol in an amount of 1.125+ -5% to 2.25+ -5%;
0.0225+ -5% to 0.1125+ -5% by weight of vitamin C;
0.225 + -5% to 1.125 + -5% by weight of sodium glutamate; and
0.225.+ -. 5% To 1.125.+ -. 5% by weight of glycerol.
3. A starter protector formulation according to claim 2 selected from the group consisting of:
Group 1:
1.125 parts by weight of trehalose;
2.25 parts by weight of skim milk powder;
1.125 parts by weight of mannitol;
0.0225 parts by weight of vitamin C;
0.225 parts by weight of sodium glutamate; and
0.225 Parts by weight of glycerol;
group 2:
1.125 parts by weight of trehalose;
2.25 parts by weight of skim milk powder;
2.25 parts by weight of mannitol;
0.1125 parts by weight of vitamin C;
1.125 parts by weight of sodium glutamate; and
1.125 Parts by weight of glycerol;
Group 3:
4.5 parts by weight of trehalose;
4.5 parts by weight of skim milk powder;
1.125 parts by weight of mannitol;
0.0225 parts by weight of vitamin C;1.125 parts by weight of sodium glutamate; and
1.125 Parts by weight of glycerol;
Group 4:
4.5 parts by weight of trehalose;
4.5 parts by weight of skim milk powder;
2.25 parts by weight of mannitol;
0.1125 parts by weight of vitamin C;0.225 parts by weight of sodium glutamate; and
0.225 Parts by weight of glycerol;
Group 5:
1.125 parts by weight of trehalose;
4.5 parts by weight of skim milk powder;
1.125 parts by weight of mannitol;
0.1125 parts by weight of vitamin C;0.225 parts by weight of sodium glutamate; and
1.125 Parts by weight of glycerol;
Group 6:
1.125 parts by weight of trehalose;
4.5 parts by weight of skim milk powder;
2.25 parts by weight of mannitol;
0.0225 parts by weight of vitamin C;1.125 parts by weight of sodium glutamate; and
0.225 Parts by weight of glycerol;
group 7:
4.5 parts by weight of trehalose;
2.25 parts by weight of skim milk powder;
1.125 parts by weight of mannitol;
0.1125 parts by weight of vitamin C;
1.125 parts by weight of sodium glutamate; and
0.225 Parts by weight of glycerol;
Group 8:
4.5 parts by weight of trehalose;
2.25 parts by weight of skim milk powder;
2.25 parts by weight of mannitol;
0.0225 parts by weight of vitamin C;
0.225 parts by weight of sodium glutamate; and
1.125 Parts by weight of glycerol.
4. The starter protector formulation of claim 1, which is in powder form.
5. The starter protectant formulation of claim 1, consisting of the following components:
1.125+ -10% to 4.5+ -10% trehalose by weight;
2.25+ -10% to 4.5+ -10% of skimmed milk powder;
mannitol in an amount of 1.125.+ -. 10% to 2.25.+ -. 10%;
0.0225 + -10% to 0.1125 + -10% by weight of vitamin C;
0.225 + -10% to 1.125 + -10% by weight of sodium glutamate;
0.225.+ -. 10% to 1.125.+ -. 10% by weight of glycerol; and
Unavoidable impurities contained in the above components after purification.
6. A starter protectant formulation as claimed in any one of claims 1-5, wherein the parts by weight of the skimmed milk powder is less than 4.5 parts by weight and the parts by weight of mannitol is less than 2.25.
7. Use of a starter protector formulation according to claim 1 in the cultivation of yeasts.
8. Use of a starter protector formulation according to claim 1 in the cultivation of pseudomonas putida (Pseudomonas putida).
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| TW109107489A TWI737210B (en) | 2020-03-06 | 2020-03-06 | Starter culture protection formula and use thereof |
| TW109107489 | 2020-03-06 |
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| TW202134423A (en) | 2021-09-16 |
| TWI737210B (en) | 2021-08-21 |
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