CN105349613A - Detection method of high-temperature-resistant aerogenesis acidophil in table vinegar - Google Patents
Detection method of high-temperature-resistant aerogenesis acidophil in table vinegar Download PDFInfo
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- CN105349613A CN105349613A CN201510929508.4A CN201510929508A CN105349613A CN 105349613 A CN105349613 A CN 105349613A CN 201510929508 A CN201510929508 A CN 201510929508A CN 105349613 A CN105349613 A CN 105349613A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
- C12Q1/06—Quantitative determination
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
- C12Q1/10—Enterobacteria
Abstract
The invention provides a detection method of high-temperature-resistant aerogenesis acidophil in table vinegar. The detection method specifically comprises the following steps: (1) diluting and uniformly shaking a to-be-detected sample; (2) continuously performing gradient dilution on a diluted solution and applying a diluent sample to an experiment on microbiology; (3) performing static cultivation on a flora, dyeing and observing by using a microscope to determine the type of contamination bacteria, and finally detecting physicochemical indexes of the contamination; the detection method can detect a strain which generates an aerogenesis acidophilic phenomenon during table vinegar brewing through a simplest and effective way and is applicable to the detection on yeast, raw material and adjunct for brewing table vinegar as well as a bottle and a cover required by canning sealing, and is beneficial for the judgment on pollution sources and pollution condition, and is suitable for the requirement of large-scale industrial production; the pollution range is decreased and the loss of enterprises is reduced.
Description
Technical field
The present invention relates to making vinegar detection technique field, the detection method of high temperature resistant aerogenesis acidophilic bacteria in especially a kind of vinegar.
Background technology
Vinegar take grain as raw material, by the techniques such as microorganism koji, saccharification, zymamsis, acetic fermentation and each seed amino acid, zymin in conjunction with brew.The existence of microorganism is had, such as: the bacterium of mould, yeast, some amount and actinomycetes in finished product vinegar production process.Genus also its local flavor comprehensive of seasoning food is belonged to according to vinegar, its sterilising conditions is: 60-80 DEG C of constant temperature 20-30min or vinegar liquid are warmed to 100 DEG C and maintain 5min, if without special contamination, the bacterium of vinegar in brewing process can kill by above-mentioned condition, and the Microbiological requirements that the hygienic standard GB2719-2003 meeting nowadays making vinegar formulates: coliform is less than 3MPN/100mL, total number of bacterial colony must not more than 10000cfu/mL, must not pathogenic bacterium be detected, after filling and sealing, there will not be aerogenesis addicted to sour phenomenon.If it occurs that aerogenesis is addicted to sour phenomenon, illustrate by other fungi pollutions, above-mentioned sterilising conditions then can not be killed, then can not meet the requirement of coliform in making vinegar hygienic standard and total number of bacterial colony (refer under certain condition (as aerobic situation, nutritional condition, pH, culture temperature and time etc.) every gram of (every milliliter) sample the total number of bacterial colonies that grows out).Therefore, simple and rapid detection go out to cause making vinegar aerogenesis addicted to the bacterial classification class of sour phenomenon and position particularly crucial.
Microbiological detection of foods index has three: coliform, total number of bacterial colony and pathogenic bacterium.Common microbiological detection method has: cultivate embrane method, spiral planar counting process, filter membrane method, ATP bioluminescence method, electrical impedance method, colour-change method, low cytometric analysis and laser scanner technique etc., these technical costss are high, and the company being unsuitable for suitability for industrialized production carries out microorganism detection.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind of composition with antihypertensive effect.
Another technical problem to be solved by this invention is to provide the above-mentioned preparation method with the composition of antihypertensive effect.
For solving the problems of the technologies described above, technical scheme of the present invention is:
A detection method for high temperature resistant aerogenesis acidophilic bacteria in vinegar, concrete steps are as follows:
(1) volume ratio g/ml or count by volume with the dilution proportion of 1:10, testing sample is shaken up that (when sample is solid, 10g sample is put into containing 100mL distilled water and the triangular flask of sterilizing together by weight; When sample is liquid, 10mL sample is put into containing 100mL distilled water and the triangular flask of sterilizing together);
(2) after getting dilution, solution proceeds gradient dilution, and the diluent sample of diluting soln in step (1) and each gradient is respectively got 6mL for Experiment on Microbiology, wherein, each test tube 1mL, totally two test tubes are for detecting coliform; The each 1mL of each flat board, totally four flat boards---two dull and stereotyped detection total number of bacterial colony, two other dull and stereotyped detection mould;
(3) flora static gas wave refrigerator, wherein,
Coliform 37 DEG C of quiescent culture 24h ± 2h;
Total number of bacterial colony 37 DEG C of quiescent culture 48h ± 2h;
Mould and yeast 28 DEG C of quiescent culture 5-7d ± 2h, after counting is observed, utilize Yihong methylene blue substratum detection to have and the bacterium of aerogenesis addicted to sour phenomenon can occur, carry out choosing bacterium line, rule rear 37 DEG C of static gas wave refrigerator 24h ± 2h, after cultivation, carries out dyeing and with microscopic examination determination microbiological contamination.
For microbiological contamination kind and finally detection is carried out to its physical and chemical index and can utilize BIOLOG identification systems.
Preferably, the detection method of high temperature resistant aerogenesis acidophilic bacteria in above-mentioned vinegar, in described step (2), the method for gradient dilution is that 1mL diluent adds in test tube containing 9mL distilled water and sterilized together, once dilutes 2 times.
Preferably, the detection method of high temperature resistant aerogenesis acidophilic bacteria in above-mentioned vinegar, use Yihong methylene blue substratum to detect causing aerogenesis acidophilic bacteria in described step (3), the prescription of described Yihong methylene blue substratum is in often liter of substratum: peptone 10g; Lactose 10g; Dipotassium hydrogen phosphate 2g; Agar 20-30g; 2% Yihong aqueous solution 20ml; 0.5% methylene blue aqueous solution 13ml; All the other are water (distilled water or deionized water); Be stirred to and dissolve completely, 121 DEG C of autoclaving 15min and get final product.
Principle: when intestinal bacteria reduce lactose produce acid time bacterium is positively charged is dyed to redness, then be combined with methylene blue and form atropurpureus bacterium colony, and with green metal gloss.Gas bacillus is then formed in brown macrocolony; The bacterium that produces acid of not reducing lactose in alkaline environment is not painted, and Yihong and methylene blue can not be in conjunction with, therefore Salmonellas etc. are colourless or amber translucent colony.S. aureus L-forms does not grow on this substratum.
Preferably, the detection method of high temperature resistant aerogenesis acidophilic bacteria in above-mentioned vinegar, in described step (3):
The detection of coliform, use lactose bile salt medium, the prescription of described lactose bile salt medium is in often liter of substratum: peptone 20.0g; Lactose 5.0g; Sodium-chlor 5.0g; Dipotassium hydrogen phosphate 4.0g; Potassium primary phosphate 1.3g; Sodium deoxycholate/bovine bile 0.5g (sodium deoxycholate)/2.0g (bovine bile); All the other are water (distilled water or deionized water); Final pH=7.4 ± 0.2;
Using method: take above-mentioned each component and add distilled water or deionized water in beaker, stirring heating is boiled to dissolving completely, 115 DEG C of sterilizing 15min are after being chilled to normal temperature, for subsequent use;
Principle: peptone provides carbon nitrogen source; Lactose is fermentable carbohydrate; Sodium-chlor maintains balanced osmotic pressure; Sodium phosphate dibasic and potassium primary phosphate are buffer reagent; Bovine bile and sodium deoxycholate are selectivity fungistat;
The detection of total number of bacterial colony, use plate count substratum, the prescription of described plate count substratum is in often liter of substratum: Tryptones: 5.0g; Yeast leaching powder: 2.5g; Glucose: 1.0g; Agar: 15.0g; All the other are water (distilled water or deionized water); PH7.0 scholar 0.2;
Using method: take above-mentioned each component and add distilled water or deionized water in beaker, be stirred to and dissolve completely, 121 DEG C of autoclaving 15min, for subsequent use;
Principle: Tryptones provides Carbon and nitrogen sources; Yeast extract paste powder provides vitamin B group; Glucose provides the energy; Agar is the peptizer of substratum;
Mould and saccharomycetic detection, use rose bengal medium, the prescription of described rose bengal medium is in often liter of substratum: peptone 5g; Glucose 10g; Potassium primary phosphate 1g; Magnesium sulfate 0.5g; Agar 15g; Rose-bengal 0.03g; Paraxin 0.1g; All the other are water (distilled water or deionized water); Final pH=7.2 ± 0.2;
Using method: take above-mentioned each component and add distilled water or deionized water in beaker, be stirred to and dissolve completely, 121 DEG C of autoclaving 20min, for subsequent use;
Principle: peptone provides Carbon and nitrogen sources; Glucose provides the energy; Potassium primary phosphate is buffer reagent; Magnesium sulfate provides necessary trace element; Agar is the peptizer of substratum; Paraxin can the growth of anti-bacteria; Rose-bengal can the growth of anti-bacteria as selectivity fungistat, and can slow down some mould because growing too fast and causing bacterium colony to stretch growth.
Beneficial effect of the present invention:
The detection method of high temperature resistant aerogenesis acidophilic bacteria in above-mentioned vinegar; the bacterial classification of making vinegar generation aerogenesis addicted to sour phenomenon is detected by most simple and efficient way; and be applicable to the detection of the Daqu of making vinegar, supplementary material and Bottle and closure needed for canning sealing; be conducive to the judgement of source of pollution and pollution condition; pollution range reduced and reduces the loss of enterprise, being applicable to the demand that large-scale industrial is produced.
Embodiment
Below in conjunction with specific embodiment, technical scheme of the present invention is further described.
Embodiment 1
A detection method for high temperature resistant aerogenesis acidophilic bacteria in vinegar, concrete steps are as follows:
(1) by liquid vinegar by volume 1:10 dilution proportion and shake up, namely 10mL sample is put into containing 100mL distilled water and the triangular flask of sterilizing together;
(2) after getting dilution, solution proceeds gradient dilution, the method of gradient dilution is that 1mL diluent adds in test tube containing 9mL distilled water and sterilized together, once dilute 2 times, the diluent sample of each gradient in diluting soln in step (1) and step (2) is respectively got 6mL for Experiment on Microbiology, wherein, each test tube 1mL, totally two test tubes are for detecting coliform; The each 1mL of each flat board, totally four flat boards---two dull and stereotyped detection total number of bacterial colony, two other dull and stereotyped detection mould;
(3) flora static gas wave refrigerator, wherein,
Coliform 37 DEG C of quiescent culture 24h;
Total number of bacterial colony 37 DEG C of quiescent culture 48h;
Mould and yeast 28 DEG C of quiescent culture 7d, after counting is observed, the detection of coliform, uses lactose bile salt medium (see above-mentioned specification sheets technical scheme part); The detection of total number of bacterial colony, uses plate count substratum (see above-mentioned specification sheets technical scheme part); Mould and saccharomycetic detection, use rose bengal medium (see above-mentioned specification sheets technical scheme part); Utilize Yihong methylene blue substratum (see above-mentioned specification sheets technical scheme part) detection to have and the bacterium of aerogenesis addicted to sour phenomenon can occur, carry out choosing bacterium line, rule rear 37 DEG C of static gas wave refrigerator 24h, after cultivation, carries out dyeing and with microscopic examination determination microbiological contamination.
By the bacterial classification of method separation and purification described in embodiment 1 after dyeing, by microscopical observation, there is Gram-positive and there is gemma characteristic, confirming that there is its feature through Physicochemical test.
Vinegar detection is carried out by method described in embodiment 1, for finding there is Gram-positive by the method and the microbiological contamination with gemma characteristic, the sterilizing under 135 DEG C of 8s conditions of a vinegar part will be detected, embodiment 1 detection method is repeated after sterilizing, do not find to there is Gram-positive and the microbiological contamination with gemma characteristic again, and aerogenesis does not occur addicted to sour phenomenon after sterilizing; Another part detects vinegar sterilizing under 121 DEG C of 30min and 130 DEG C 8s condition respectively, embodiment 1 detection method is repeated after sterilizing, all again find have Gram-positive and have the microbiological contamination of gemma characteristic, and the former quantity being more than the latter, still there is aerogenesis addicted to sour phenomenon in vinegar.
To sum up, the vinegar that described in employing embodiment 1, method carries out detecting is not when reaching its fatal temperature and time (135 DEG C of 8s), pollution bacterial classification can be detected, vinegar generation aerogenesis is addicted to sour phenomenon, when more than 135 DEG C of 8s, do not detect that this pollutes bacterial classification, and aerogenesis does not occur addicted to sour phenomenon.
Meanwhile, detect that utilizing method described in embodiment 1 after causing the bacterium of vinegar aerogenesis addicted to sour phenomenon, after utilizing the sterilizing of high pressure instantaneous sterilizing method, finished product vinegar aerogenesis does not occur addicted to sour phenomenon.And for when not by the fungi pollution of this initiation vinegar aerogenesis addicted to sour phenomenon, the sterilising conditions of finished product vinegar is 90 DEG C of sterilizing 20min, if there are intestinal bacteria, it is comparatively strong with thermotolerance compared with other bacillus, but its 20min at 121 DEG C can be killed substantially; If there is general sporeformer, it also can be killed under 121 DEG C of 15min, and other bacteriums, mould and yeast can be killed at the same temperature, and described in embodiment 1, method detects it, and result is consistent with existing detection method result.
Above-mentioned detailed description of carrying out with reference to the detection method of embodiment to aerogenesis acidophilic bacteria high temperature resistant in this kind of vinegar; illustrative instead of determinate; several embodiments can be listed according to institute's limited range; therefore in the change do not departed under general plotting of the present invention and amendment, should belong within protection scope of the present invention.
Claims (4)
1. the detection method of high temperature resistant aerogenesis acidophilic bacteria in vinegar, is characterized in that: concrete steps are as follows:
(1) testing sample shakes up with the dilution proportion of 1:10 by volume ratio g/ml or count by volume by weight;
(2) after getting dilution, solution proceeds gradient dilution, and the diluent sample of diluting soln in step (1) and each gradient is respectively got 6mL for Experiment on Microbiology, wherein, each test tube 1mL, totally two test tubes are for detecting coliform; The each 1mL of each flat board, totally four flat boards---two dull and stereotyped detection total number of bacterial colony, two other dull and stereotyped detection mould;
(3) flora static gas wave refrigerator, wherein,
Coliform 37 DEG C of quiescent culture 24h ± 2h;
Total number of bacterial colony 37 DEG C of quiescent culture 48h ± 2h;
Mould and yeast 28 DEG C of quiescent culture 5-7d ± 2h, after counting is observed, utilize Yihong methylene blue substratum detection to have and the bacterium of aerogenesis addicted to sour phenomenon can occur, carry out choosing bacterium line, rule rear 37 DEG C of static gas wave refrigerator 24h ± 2h, after cultivation, carries out dyeing and with microscopic examination determination microbiological contamination.
2. the detection method of high temperature resistant aerogenesis acidophilic bacteria in vinegar according to claim 1, it is characterized in that: in described step (2), the method for gradient dilution is that 1mL diluent adds in test tube containing 9mL distilled water and sterilized together, once dilute 2 times.
3. the detection method of high temperature resistant aerogenesis acidophilic bacteria in vinegar according to claim 1, it is characterized in that: use Yihong methylene blue substratum to detect causing aerogenesis acidophilic bacteria in described step (3), the prescription of described Yihong methylene blue substratum is in often liter of substratum: peptone 10g; Lactose 10g; Dipotassium hydrogen phosphate 2g; Agar 20-30g; 2% Yihong aqueous solution 20ml; 0.5% methylene blue aqueous solution 13ml; All the other are water.
4. the detection method of high temperature resistant aerogenesis acidophilic bacteria in vinegar according to claim 1, is characterized in that: in described step (3):
The detection of coliform, use lactose bile salt medium, the prescription of described lactose bile salt medium is in often liter of substratum: peptone 20.0g; Lactose 5.0g; Sodium-chlor 5.0g; Dipotassium hydrogen phosphate 4.0g; Potassium primary phosphate 1.3g; Sodium deoxycholate/bovine bile 0.5g/2.0g; All the other are water; Final pH=7.4 ± 0.2;
The detection of total number of bacterial colony, use plate count substratum, the prescription of described plate count substratum is in often liter of substratum: Tryptones: 5.0g; Yeast leaching powder: 2.5g; Glucose: 1.0g; Agar: 15.0g; All the other are water; PH7.0 scholar 0.2;
Mould and saccharomycetic detection, use rose bengal medium, the prescription of described rose bengal medium is in often liter of substratum: peptone 5g; Glucose 10g; Potassium primary phosphate 1g; Magnesium sulfate 0.5g; Agar 15g; Rose-bengal 0.03g; Paraxin 0.1g; All the other are water; Final pH=7.2 ± 0.2.
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Cited By (3)
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| CN109371100A (en) * | 2018-11-20 | 2019-02-22 | 四川农业大学 | A kind of culture medium and its method for the detection of vinegar aerogenic bacteria |
| CN111440842A (en) * | 2020-05-28 | 2020-07-24 | 江南大学 | Method for detecting microorganisms in rice dumplings |
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Cited By (5)
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| CN105624262A (en) * | 2016-03-17 | 2016-06-01 | 内蒙古蒙牛乳业（集团）股份有限公司 | Method for measuring content of non-lab (non-lactic acid bacteria) in dairy product |
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| CN109371100B (en) * | 2018-11-20 | 2023-09-22 | 四川农业大学 | Culture medium for detecting vinegar gas-producing bacteria and method thereof |
| CN111440842A (en) * | 2020-05-28 | 2020-07-24 | 江南大学 | Method for detecting microorganisms in rice dumplings |
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