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GB 4789.41-2016 PDF English

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GB 4789.41-2016: National food safety standard - Microbiological Examination of Food Hygiene - Examination of Enterobacteriaceae
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GB 4789.41-201685 Add to Cart Auto, 9 seconds. National food safety standard - Microbiological Examination of Food Hygiene - Examination of Enterobacteriaceae Valid

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GB 4789.41-2016: National food safety standard - Microbiological Examination of Food Hygiene - Examination of Enterobacteriaceae


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GB NATIONAL STANDARD OF THE PEOPLE’S REPUBLIC OF CHINA National Food Safety Standard - Microbiological Examination of Food Hygiene - Examination of Enterobacteriaceae Issued on. AUGUST 31, 2016 Implemented on. MARCH 1, 2017 Issued by. National Health and Family Planning Commission of the People’s Republic of China

Table of Contents

1 Application Scope... 3 2 Terms and Definitions... 3 3 Apparatus and Materials... 3 4 Culture Media and Reagents... 4 5 Test Procedures... 5 6 Operating Procedures... 5 7 Report... 9 8 Test Procedures... 9 9 Operating Procedures... 11 10 Report... 11 Annex A Examples for Calculation of Results... 12 Annex B Culture Media and Reagents... 14 Annex C Retrieval Table of Enterobacteriaceae Most Probable Numbers (MPNs)... 18 National Food Safety Standard - Microbiological Examination of Food Hygiene - Examination of Enterobacteriaceae

1 Application Scope

This Standard specifies the method for the test of Enterobacteriaceae in foods. The first method of this Standard applies to the bacterial counts of Enterobacteriaceae in the foods which has a high content of Enterobacteriaceae; and the second method applies to the bacterial counts of Enterobacteriaceae in the foods which has a low content of Enterobacteriaceae.

2 Terms and Definitions

2.1 Enterobacteriaceae The oxidase-negative aerobic or facultative anaerobic Gram-negative non-spore- bearing bacillus, which ferments glucose producing acid under given conditions. 2.3 most probable number; MPN An indirect enumeration method based on Poisson's distribution.

3 Apparatus and Materials

In addition to the conventional sterilization and culture apparatus in the microbiological laboratories, the other apparatus and materials are as follows. 3.1 Thermostatic incubator. 36°C ± 1°C. 3.2 Refrigerator. 2°C ~ 5°C. 3.3 Water bath. 46°C ± 1°C. 3.6 Homogenizer. 3.7 Shaker. 3.8 Sterile pipettes. 1 ml (having graduates of 0.01 ml), 10 ml (having 0.1 ml) or micropipettors and tips. 3.9 Sterile conical flasks or equivalent vessels. capacities 150 ml and 500 ml. 3.12 pH meters or pH colorimetric tubes or precise pH test paper.

4 Culture Media and Reagents

4.1 Buffered peptone water (BPW). see B.1. 4.2 Buffered glucose brilliant green bile broth (EE broth). see B.2. 4.5 Glucose agar. see B.5. 4.6 Gram stain solution. see B.6. 4.7 Oxidase reagent. see B.7. 4.8 Sterile 1 mol/l NaOH. see B.8.

5 Test Procedures

See Figure 1 for the test procedures for the Enterobacteriaceae plate count method.

6 Operating Procedures

6.1 Specimen dilution Test specimen 6.1.1 Solid and semi-solid specimens. take 25 g of specimen to place into a sterile homogeneous cup containing 225 ml of BPW, homogenize for 1 min ~ 2 min at 8 000 r/min ~ 10 000 r/min, or place into a homogeneous bag containing 225 ml of BPW, 6.1.2 Liquid specimens. use a sterile pipette to absorb 25 ml of specimen to pour into a sterile conical flask (in which an appropriate quantity of sterile glass beads are provided in the flask in advance) containing 225 ml of BPW, mix up, and make homogeneous specimen solutions of 1.10. 6.2 Pout plate and culture 6.2.1 In accordance with the estimation of the contamination of specimens and relevant limit requirements, select 2 ~ 3 homogeneous specimen solutions (liquid specimens may be raw solutions) of appropriate continuous dilution and inoculate the solutions of each dilution degree to 2 sterile plates. And Meanwhile, absorb 1 ml of BPW each to add to two sterile plates as blank control. 6.2.4 Turn over the plate after the upper-layer agar solidifies on the VRBGA plate and culture for 18 h ~ 24 h at 36°C ± 1°C. 6.3 Enumeration and confirmation of typical colonies 6.3.1 The typical colonies of Enterobacteriaceae are pink to red or violet colonies with or without precipitation rings. Select the plates with typical colony count between 15 CFU ~ 150 CFU and without spreading growth of colonies and only enumerate the typical colonies. The colony counting is expressed by the colony-forming unit, CFU. 6.3.5 Confirmation of typical colonies. 6.4 Calculation of results 6.4.1 General principles If the plate counts of typical colonies of two continuous dilution degrees are within the appropriate counting range, calculate in accordance with Formula (1). See A.1, A.2, A.3 and A.4 for the examples. 6.4.2 Small colony counts If all the typical colony counts of the plate of minimum dilution degree (including the raw solution of the liquid specimen) are less than 15 CFU and there are confirmed Enterobacteriaceae colonies, then multiply the confirmed colony count by the minimum dilution factor. 6.4.3 Special circumstances If all the typical colony counts on the plate of the first dilution degree are greater than 150 CFU and there are confirmed Enterobacteriaceae colonies; and if there is no confirmed Enterobacteriaceae colony on the plate of the second dilution degree or the typical colony counts are not between 15 CFU ~ 150 CFU, then multiply the confirmed colony count by the first dilution factor. See A.5 for the example.

7 Report

7.1 When the colony count is less than 100, it shall be reported with integers. 7.2 When the colony count is greater or equal to 100, take the first two digits after rounding off the third digit, and use 0 to replace the places after it; 7.3 The numbers shall be rounded off to the nearest whole numbers. 7.4 If it is impossible to count because of spreading colonies on all the plates, then report the colony spreading.

8 Test Procedures

See Figure 2 for the test procedures for the Enterobacteriaceae most probable number method.

9 Operating Procedures

9.1 Specimen dilution As in 6.1. 9.2 Inoculation and culture 9.2.1 Nonselective pre-enrichment In accordance with the estimation of the contamination of specimens and relevant limit requirements, select 3 homogeneous specimen solution of appropriate continuous dilution degrees, 3 test tubes for each dilution degree and altogether 9 test tubes of BPW, and culture for 18 h ± 2 h at 36°C ± 1°C. 9.3 Confirmation of typical colonies See 6.3 for the typical colonies.

10 Report

Enterobacteriaceae is Gram-negative non-spore-bearing bacillus, which ferments glucose producing acid and is oxidase negative. Once one colony is confirmed Enterobacteriaceae, the EE tube represented by it is Enterobacteriaceae positive.

Annex A

Examples for Calculation of Results A.1 Both of two continuous dilution degrees include appropriate typical colony counting plates and contain confirmed Enterobacteriaceae colonies. See Table A.1 for the data. A.2 If both of two continuous dilution degrees include appropriate typical colony counting plates, including one plate colony count of some dilution degree not within the appropriate range or one plate colony count on each of two dilution degrees not within the appropriate range, then the corresponding plates shall not be used as counting plates. See Table A.2 for the data. A.3 If both of two continuous dilution degrees include appropriate colony counting plates, including one plate on which there is not confirmed Enterobacteriaceae colony, the corresponding plates are not used as counting plates. See Table A.3 for the data. A.5 If the colony counts on the plate of the first dilution degree are greater than 150 CFU and there are confirmed Enterobacteriaceae colonies; and if there is no confirmed Enterobacteriaceae colony on the plate of the second dilution degree or the typical colony counts are not between 15 CFU ~ 150 CFU, then multiply the confirmed colony count by the first dilution factor. See A.5 for the example.

Annex B

Culture Media and Reagents B.1 Buffered peptone water (BPW) B.2 Buffered glucose brilliant green bile broth (EE broth) B.3 Crystal violet neutral red bile salt glucose agar (VRBGA) B.4 Nutrient agar (NA) B.5 Glucose agar B.6 Gram stain solutions B.6.1 Crystal violet stain solution

Annex C

Retrieval Table of Enterobacteriaceae Most Probable Numbers (MPNs) ......
Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al.
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