This standard specifies the method for determining the total number of bacterial colonies in foods. This standard is applicable to the determination of the total number of bacterial colonies in various types of foods. GB/T 4789.2-2003 Food hygiene microbiological examination Determination of total number of bacterial colonies GB/T4789.2-2003 Standard download decompression password: www.bzxz.net

ICS07.100.30
National Standard of the People's Republic of China
GB/T 4789.2--2003
Replaces GB/T4789.2-1994
Microbiological examination of food hygiene-Detection of aerobic bacterial countPromulgated on August 11, 2003
Ministry of Health of the People's Republic of China
Standardization Administration of the People's Republic of China
Implementation on January 1, 2004
GB/T4789.2-2003
This standard revise GB/T4789.2-1994 "Microbiological examination of food hygiene-Determination of aerobic bacterial count".
Compared with GB/T4789.2-1994, this standard is mainly modified as follows: The format and text of the standard text are modified in accordance with GB/T1.1-2000. The "equipment and materials" in the original standard are modified and standardized. 1. The unit of expression of total colony count is changed to: cfu/g (ml.). From the date of implementation of this standard, GB/T4789.2-1994 will be abolished at the same time. This standard is proposed and managed by the Ministry of Health of the People's Republic of China. The drafting unit of this standard: Nutrition and Food Safety Institute of China Center for Disease Control and Prevention. The main drafters of this standard: Liu Hongdao, Ji Rong, Fu Ping, Yang Baolan, Yao Jinghui. This standard was first issued in 1984, revised for the first time in 1994, and this is the second revision. 1 Scope
Microbiological examination of food hygiene
Determination of total colony count
This standard specifies the method for determining the total colony count in food. This standard is applicable to the determination of the total colony count in various types of food. 2 Normative references
GB/T4789.2—2003
The clauses in the following documents become the clauses of this standard through reference in this standard. For all dated references, all subsequent amendments (excluding errata) or revisions are not applicable to this standard. However, parties to an agreement based on this standard are encouraged to study whether the latest versions of these documents can be used. For all undated references, the latest versions are applicable to this standard. GB/T4789.28—2003 Food hygiene microbiological examination staining methods, culture media and reagents 3 Terms and definitions
The following terms and definitions apply to this standard. 3.1
Aerobic bacterial count The total number of colonies contained in 1 mL (g) of the food sample after it has been treated and cultured under certain conditions (such as culture medium composition, culture temperature and time, pH, aerobic properties, etc.). The results obtained under the culture conditions specified in this method only include the total number of a group of mesophilic aerobic colonies growing on nutrient agar. The total number of colonies is mainly used as a sign to determine the degree of food contamination. This method can also be used to observe the dynamics of bacterial reproduction in food, so as to provide a basis for the hygienic evaluation of the sample being tested. 4 Equipment and materials
4.1 Refrigerator: 0℃~4℃.
4.2 Constant temperature incubator: 36℃±1℃.
4.3 Constant temperature water bath: 46℃±1℃.
4.4 Homogenizer or sterile mortar.
4.5 Rack-plate medicine balance: 0g~500g, accurate to 0.5g4.6 Colony counter.
Magnifying glass 4×.
Sterile pipette: 1mL (with 0.01mL scale), 10mL (with 0.1mL scale). Sterile conical flask: 500mL.
Sterile glass beads: about 5mm in diameter.
Sterile culture III: 90mm in diameter.
Sterile test tube: 16mm×160mm.
Sterile knife, scissors, tweezers, etc.
Culture medium and reagents
5.1 Nutrient agar culture medium: in accordance with 4.7 of GB/T4789.28-2003. 9
GB/T 4789.2-2003
5.2 Phosphate buffer in accordance with 3.22 of GB/T4789.28-2003. 5.30.85% sterile saline.
5.475% ethanol.
6 Inspection procedure
The inspection procedure for the total number of colonies is shown in Figure 1.
Make several dilutions with appropriate multiples
Choose 2~3 appropriate dilutions
Take 1mL of each and add to sterile culture medium
Add appropriate amount of nutrient agar to each medium
48h±2h
Count colonies
7 Operation steps
7.1 Dilution and culture of test samples
7.1.1 Aseptically cut 25g (mL) of the test sample into pieces and place them in a sterile glass bottle containing 225mL of sterile physiological saline or other diluent (with appropriate number of glass beads pre-placed in the bottle) or a sterile mortar, and shake or grind them thoroughly to make a 1:10 uniform dilution. After adding the dilution, it is best to place the solid test sample in a homogenizer at a speed of 8000r/min~10000r/min for 1min to make a 1:10 uniform dilution.
7.1.2 Use a 1mL sterile pipette to draw 1mL of the 110 dilution solution, and slowly inject it into a test tube containing 9mL of sterile physiological saline or other diluent along the tube wall (be careful not to let the tip of the pipette touch the diluent in the tube), shake the test tube, mix evenly, and make a 1:100 dilution solution. 7.1.3 Take another 1mL sterile pipette and make 10-fold incremental dilutions according to the above operation method. After each incremental dilution, use a 1mL sterile pipette.
7.1.4 According to the requirements of food hygiene standards or the estimation of specimen contamination, select 2 to 3 appropriate dilutions. While making 10-fold incremental dilutions, use the pipette that draws the dilution to transfer 1mL of the dilution solution into a sterile culture dish. Make two culture dishes for each dilution.
7.1.5 After the diluent is transferred to the culture dish, about 15 mL of the nutrient agar medium cooled to 46°C (can be placed in a 46°C ± 1°C water bath for insulation) should be injected into the culture dish in a timely manner, and the culture dish should be rotated to mix evenly. At the same time, the nutrient agar medium is poured into the sterilized culture III with 1 mL of diluent as a blank control. Www.bzxZ.net
7.1.6 After the agar solidifies, turn the plate over and place it in a 36°C ± 1°C incubator for 48h ± 2h. 7.2 Colony counting method
GB/T4789.2--2003
When counting the colonies on the plate, you can observe with the naked eye and use a magnifying glass when necessary to prevent omissions. After recording the number of colonies on each plate, calculate the average total number of colonies on each plate with the same dilution. 7.3 Report of Colony Counts
7.3.1 Selection of Plate Colony Counts
Select plates with colony counts between 30 and 300 as the standard for total colony count determination. When two plates are used for one dilution, the average of the two plates should be used. If one plate has large flake colonies, it should not be used. Instead, the plate without flake colonies should be used as the colony count for that dilution. If flake colonies are less than half of the plate, and the colonies in the remaining half are evenly distributed, half of the plate can be counted and then multiplied by 2 to represent the total III colony count. If there are chain colonies growing on the plate (no obvious boundaries between colonies), if there is only one chain, it can be regarded as one colony; if there are several chains from different sources, each chain should be counted as one colony. 7.3.2 Selection of Dilutions
7.3.2.1 A dilution with an average colony count between 30 and 300 should be selected and reported by multiplying it by the dilution multiple (see Example 1 in Table 1). 7.3.2.2 If there are two dilutions, and the number of colonies grown is between 30 and 300, the ratio of the two dilutions shall determine the number of colonies. If the ratio is less than or equal to 2, the average shall be reported; if it is greater than 2, the smaller number shall be reported (see Example 2 and Example 3 in Table 1). 7.3.2.3 If the average colony count of all dilutions is greater than 300, the average colony count of the highest dilution multiplied by the dilution factor shall be reported (see Example 4 in Table 1).
7.3.2.4 If the average colony count of all dilutions is less than 30, the average colony count of the lowest dilution multiplied by the dilution factor shall be reported (see Example 5 in Table 1).
7.3.2.5 If no colonies grow at any dilution, report it as less than 1 multiplied by the lowest dilution factor (see Example 6 in Table 1). 7.3.2.6 If the average colony counts of all dilutions are not between 30 and 300, and some of them are greater than 300 or less than 30, then the average colony count closest to 30 or 300 multiplied by the dilution factor shall be reported (see Example 7 in Table 1). 7.3.3 Report of colony counts
When the colony count is less than 100, it shall be reported as the actual number. When it is greater than 100, two significant figures shall be used. The value after the two significant figures shall be rounded up. In order to shorten the number of zeros after the number, it can also be expressed as an exponent of 10 (see Table 1). Table 1 Selection of dilution and reporting method of colony counts Dilution and colony counts
Too many to count
Too many to count
Too many to count
Too many to count
Too many to count
Too many to count
Too many to count
Ratio of two dilutions
Total number of colonies/
[efu/g(mL)]
313000||tt ||<1×10
Reporting method/[cfu/g(mL）]
16000 or 1.6×10
38000 or 3.8×10
27000 or 2.7×10
310000 or 3.1×105
270 or 2.7×102
31000 or 3.1×10
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