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NY/T 1331-2007: Enumeration of Colony of Psychrotrophic Microorganisms, Total Aerobic Bacterial Spores and Thermophilic Aerobic Bacterial Spores in Milk and Dairy Products
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ICS 67.100.10 X 16 PROFESSIONAL STANDARD Enumeration of Colony of Psychrotrophic Microorganisms, Total Aerobic Bacterial Spores and Thermophilic Aerobic Bacterial Spores in Milk and Dairy Products (ISO 6730.1992, NEN 6813.1999, NEN 6809.1999, NEQ) Issued on. APRIL 17, 2007 Implemented on. JULY 01, 2007 Issued by. The Ministry of Agriculture of the People's Republic of China
Foreword
This Standard is drafted by reference to "Milk - Enumeration of Colony-forming Units of Psychrotrophic Micro-organisms - Colony-Count Technique at 6.5℃" (ISO 6730.1992, English version), "Milk and Dairy Products-Determination of the Content of Spores of Mesophilic Aerobic Sporeforming Bacteria" (NEN 6813.1999, Dutch version) and "Milk and Dairy Products - Determination of the Content of Thermophilic Aerobic Sporeforming Bacteria" (NEN 6809.1999, Dutch version). This Standard is not equivalent to ISO 6730. 1992, NEN 6813.1999, NEN 6809.1999. This Standard was proposed by the Ministry of Agriculture of the People's Republic of China. This Standard is under the jurisdiction of Animal Husbandry of Standardization Administration of China. Drafting organization of this Standard. Ministry of Agriculture Dairy Quality Supervision and Testing Center Chief drafting staffs of this Standard. Li Qiying, Ma Wenhong, Zhang Hua and Wang Jinhua.Table of Contents
Part I. Enumeration of Colony of Psychrotrophic Microorganisms in Milk... 4 1 Scope... 4 2 Normative References... 4 3 Terms and Definitions... 4 4 Principle... 4 5 Diluent and Culture Medium... 4 6 Equipment and Materials... 5 7 Sampling... 5 8 Test Procedures... 5 9 Operational Steps... 6 10 Result Calculation... 7 11 Repeatability... 7 Part II. Enumeration of Colony of Aerobic Bacterial Spores... 9 1 Scope... 9 2 Normative References... 9 3 Terms and Definitions... 9 4 Principle... 9 5 Diluent and Culture Medium... 9 6 Equipment and Materials... 10 7 Sampling... 10 8 Test Steps... 10 9 Operation Procedures... 11 10 Result Calculation... 12 Part III. Enumeration of Colony of Thermophilic Aerobic Bacterial Spores... 14 1 Scope... 14 2 Normative References... 14 3 Terms and Definitions... 14 4 Principle... 14 5 Diluent and Culture Medium... 14 6 Equipment and Materials... 15 7 Sampling... 15 8 Test Procedures... 15 9 Operational Steps... 16 10 Result Calculation... 17 References and Original Chinese Documents... 19 Enumeration of Colony of Psychrotrophic Microorganisms, Total Aerobic Bacterial Spores and Thermophilic Aerobic Bacterial Spores in Milk and Dairy Products Part I. Enumeration of Colony of Psychrotrophic Microorganisms in Milk1 Scope
This Part specifies the test method for the enumeration of colony of psychrotrophic microorganisms in milk. This Part is applicable to the enumeration of colony of psychrotrophic microorganisms in raw and fresh milk and pasteurized milk.2 Normative References
The following normative documents contain provisions which, through reference by this Part, constitute the provisions of this Part. For dated reference, subsequent amendments (excluding the corrected contents) or revisions do not apply to this Part. However, parties who enter into agreements based on this Standard are encouraged to investigate the possibility of applying the most recent editions of the relevant standards indicated below. For any undated references, the latest edition of the document applies to this Part. GB/T 4789.1 Microbiological Examination of Food Hygiene - General Principles GB/T 4789.28-2003 Microbiological Examination of Food Hygiene - Staining Methods, Culture Mediums and Reagents3 Terms and Definitions
The following terms and definitions are applicable to this Part. 3.1 Psychrotrophic microorganisms Under condition of 6.5℃, with aerobic incubation of 10d, the bacteria, yeasts and mildews forming countable colonies on MPC agar plate.4 Principle
Incubate the sample in the milk plate count agar, at 6.5℃ for 10d, then count the number of psychrotrophic microorganisms per milliliter through colony counting.5 Diluent and Culture Medium
5.1 Peptone salt solution 5.1.1 Components Enzyme hydrolyzed casein peptone 1.0g 5.1.2 Preparation Method Dissolve the enzyme hydrolyzed casein peptone and sodium chloride into distilled water. Calculate the pH value to 7.0±0.1 with sodium hydroxide solution with mass fraction of 15%. Pour into test tubes and flasks, 9mL per tube, 90mL or 225mL per flask, relative error less than or equal to±2%. After sealed, 121℃±1℃ autoclave for 15min. 5.3 Milk plate count agar culture medium 5.3.2 Preparation Method Dissolve all components (excluding agar) specified in Article 5.3.1 into distilled water, and calibrate the pH value to 7.0±0.1 with sodium hydroxide solution with mass fraction of 15%. Add agar, heat up until boiling to melt the agar. If necessary, filtered by two-layer gauze filled with absorbent cotton.6 Equipment and Materials
6.1 Common equipment and materials of microbiological laboratory 6.2 Biochemical incubator. 6.5℃±0.5℃. 6.3 Water bath boiler. 45℃±1℃.7 Sampling
Sampling shall be carried out in accordance with GB/T 4789.1.8 Test Procedures
See Figure 1 for test procedures for the number of psychrotrophic microorganisms. Figure 19 Operational Steps
9.1 Sample treatment 9.1.1 Turn over the sample containers rapidly for 25 times to uniformly mix the sample and to avoid forming of foam. Then unwrap the sample containers with sterile operation. 9.2 Inoculation and incubation 9.2.1 According to the estimation of psychrotrophic microorganism contamination situation of the samples, select three suitable dilutions. While the 10-times dilution is being carried out, simultaneously use the sterile pipette to suck and transfer 1mL such diluent into the sterile petri dishes (see Section 6.4). Each dilution inoculates two dishes. 9.2.4 Conduct a blank test with 1mL diluent while inoculating. 9.2.5 After the agar is solidified, pile and reversely place the petri dishes in the incubator (see Section 6.2), at 6.5℃±0.5℃ for 10d. The piled petri dishes shall not exceed 6 pieces. 9.3 Colony counting 9.3.1 Use visual inspection for plate colony counting. Very tiny colonies shall also be counted. Magnifier shall be used if required. NY/T 1331-2007 ICS 67.100.10 X 16 PROFESSIONAL STANDARD Enumeration of Colony of Psychrotrophic Microorganisms, Total Aerobic Bacterial Spores and Thermophilic Aerobic Bacterial Spores in Milk and Dairy Products (ISO 6730.1992, NEN 6813.1999, NEN 6809.1999, NEQ) Issued on. APRIL 17, 2007 Implemented on. JULY 01, 2007 Issued by. The Ministry of Agriculture of the People's Republic of ChinaForeword
This Standard is drafted by reference to "Milk - Enumeration of Colony-forming Units of Psychrotrophic Micro-organisms - Colony-Count Technique at 6.5℃" (ISO 6730.1992, English version), "Milk and Dairy Products-Determination of the Content of Spores of Mesophilic Aerobic Sporeforming Bacteria" (NEN 6813.1999, Dutch version) and "Milk and Dairy Products - Determination of the Content of Thermophilic Aerobic Sporeforming Bacteria" (NEN 6809.1999, Dutch version). This Standard is not equivalent to ISO 6730. 1992, NEN 6813.1999, NEN 6809.1999. This Standard was proposed by the Ministry of Agriculture of the People's Republic of China. This Standard is under the jurisdiction of Animal Husbandry of Standardization Administration of China. Drafting organization of this Standard. Ministry of Agriculture Dairy Quality Supervision and Testing Center Chief drafting staffs of this Standard. Li Qiying, Ma Wenhong, Zhang Hua and Wang Jinhua.Table of Contents
Part I. Enumeration of Colony of Psychrotrophic Microorganisms in Milk... 4 1 Scope... 4 2 Normative References... 4 3 Terms and Definitions... 4 4 Principle... 4 5 Diluent and Culture Medium... 4 6 Equipment and Materials... 5 7 Sampling... 5 8 Test Procedures... 5 9 Operational Steps... 6 10 Result Calculation... 7 11 Repeatability... 7 Part II. Enumeration of Colony of Aerobic Bacterial Spores... 9 1 Scope... 9 2 Normative References... 9 3 Terms and Definitions... 9 4 Principle... 9 5 Diluent and Culture Medium... 9 6 Equipment and Materials... 10 7 Sampling... 10 8 Test Steps... 10 9 Operation Procedures... 11 10 Result Calculation... 12 Part III. Enumeration of Colony of Thermophilic Aerobic Bacterial Spores... 14 1 Scope... 14 2 Normative References... 14 3 Terms and Definitions... 14 4 Principle... 14 5 Diluent and Culture Medium... 14 6 Equipment and Materials... 15 7 Sampling... 15 8 Test Procedures... 15 9 Operational Steps... 16 10 Result Calculation... 17 References and Original Chinese Documents... 19 Enumeration of Colony of Psychrotrophic Microorganisms, Total Aerobic Bacterial Spores and Thermophilic Aerobic Bacterial Spores in Milk and Dairy Products Part I. Enumeration of Colony of Psychrotrophic Microorganisms in Milk1 Scope
This Part specifies the test method for the enumeration of colony of psychrotrophic microorganisms in milk. This Part is applicable to the enumeration of colony of psychrotrophic microorganisms in raw and fresh milk and pasteurized milk.2 Normative References
The following normative documents contain provisions which, through reference by this Part, constitute the provisions of this Part. For dated reference, subsequent amendments (excluding the corrected contents) or revisions do not apply to this Part. However, parties who enter into agreements based on this Standard are encouraged to investigate the possibility of applying the most recent editions of the relevant standards indicated below. For any undated references, the latest edition of the document applies to this Part. GB/T 4789.1 Microbiological Examination of Food Hygiene - General Principles GB/T 4789.28-2003 Microbiological Examination of Food Hygiene - Staining Methods, Culture Mediums and Reagents3 Terms and Definitions
The following terms and definitions are applicable to this Part. 3.1 Psychrotrophic microorganisms Under condition of 6.5℃, with aerobic incubation of 10d, the bacteria, yeasts and mildews forming countable colonies on MPC agar plate.4 Principle
Incubate the sample in the milk plate count agar, at 6.5℃ for 10d, then count the number of psychrotrophic microorganisms per milliliter through colony counting.5 Diluent and Culture Medium
5.1 Peptone salt solution 5.1.1 Components Enzyme hydrolyzed casein peptone 1.0g 5.1.2 Preparation Method Dissolve the enzyme hydrolyzed casein peptone and sodium chloride into distilled water. Calculate the pH value to 7.0±0.1 with sodium hydroxide solution with mass fraction of 15%. Pour into test tubes and flasks, 9mL per tube, 90mL or 225mL per flask, relative error less than or equal to±2%. After sealed, 121℃±1℃ autoclave for 15min. 5.3 Milk plate count agar culture medium 5.3.2 Preparation Method Dissolve all components (excluding agar) specified in Article 5.3.1 into distilled water, and calibrate the pH value to 7.0±0.1 with sodium hydroxide solution with mass fraction of 15%. Add agar, heat up until boiling to melt the agar. If necessary, filtered by two-layer gauze filled with absorbent cotton.6 Equipment and Materials
6.1 Common equipment and materials of microbiological laboratory 6.2 Biochemical incubator. 6.5℃±0.5℃. 6.3 Water bath boiler. 45℃±1℃.7 Sampling
Sampling shall be carried out in accordance with GB/T 4789.1.8 Test Procedures
See Figure 1 for test procedures for the number of psychrotrophic microorganisms. Figure 19 Operational Steps
9.1 Sample treatment 9.1.1 Turn over the sample containers rapidly for 25 times to uniformly mix the sample and to avoid forming of foam. Then unwrap the sample containers with sterile operation. 9.2 Inoculation and incubation 9.2.1 According to the estimation of psychrotrophic microorganism contamination situation of the samples, select three suitable dilutions. While the 10-times dilution is being carried out, simultaneously use the sterile pipette to suck and transfer 1mL such diluent into the sterile petri dishes (see Section 6.4). Each dilution inoculates two dishes. 9.2.4 Conduct a blank test with 1mL diluent while inoculating. 9.2.5 After the agar is solidified, pile and reversely place the petri dishes in the incubator (see Section 6.2), at 6.5℃±0.5℃ for 10d. The piled petri dishes shall not exceed 6 pieces. 9.3 Colony counting 9.3.1 Use visual inspection for plate colony counting. Very tiny colonies shall also be counted. Magnifier shall be used if required. ......Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al.
