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食品安全国家标准 食品微生物学检验 菌落总数测定
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GB 4789.2-2022
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标准编号: GB 4789.2-2022 (GB4789.2-2022)
中文名称: 食品安全国家标准 食品微生物学检验 菌落总数测定
英文名称: National food safety standard - Microbiological examination of food: Aerobic plate count
行业: 国家标准
中标分类: X09
字数估计: 10,190
发布日期: 2022-06-30
实施日期: 2022-12-30
旧标准 (被替代): GB 4789.2-2016
归口单位: 国家健康卫生委员会
发布机构: 国家市场监督管理总局
GB 4789.2-2022: 食品安全国家标准 食品微生物学检验 菌落总数测定
GB 4789.2-2022 英文名称: National food safety standard - Microbiological examination of food: Aerobic plate count
书中华人民共和国国家标准
GB ４７８９．２-２０２２
１ 范围
本标准规定了食品中菌落总数（Ａｅｒｏｂｉｃｐｌａｔｅｃｏｕｎｔ）的测定方法。本标准适用于食品中菌落总数的测定。
５ 检验程序
菌落总数的检验程序见图１。
６ 操作步骤
６．１ 样品的稀释
６．１．１ 固体和半固体样品：称取２５ｇ样品置于盛有２２５ｍＬ无菌磷酸盐缓冲液或无菌生理盐水的无菌
均质杯内，８０００ｒ／ｍｉｎ～１００００ｒ／ｍｉｎ均质１ｍｉｎ～２ｍｉｎ，或放入盛有２２５ｍＬ稀释液的无菌均质袋
中，用拍击式均质器拍打１ｍｉｎ～２ｍｉｎ，制成１∶１０的样品匀液。
６．１．２ 液体样品：以无菌吸管吸取２５ｍＬ样品置于盛有２２５ｍＬ无菌磷酸盐缓冲液或无菌生理盐水的
无菌锥形瓶（瓶内可预置适当数量的无菌玻璃珠）中，充分混匀，或放入盛有２２５ｍＬ稀释液的无菌均质
袋中，用拍击式均质器拍打１ｍｉｎ～２ｍｉｎ，制成１∶１０的样品匀液。当结果要求为每ｇ样品中菌落总数时，按６．１．１操作。
６．１．３ 用１ｍＬ无菌吸管或微量移液器吸取１∶１０样品匀液１ｍＬ，沿管壁缓慢注于盛有９ｍＬ稀释液
的无菌试管中（注意吸管或吸头尖端不要触及稀释液面），在振荡器上振荡混匀，制成１∶１００的样品匀液。
６．１．４ 按６．１．３操作，制备１０倍系列稀释样品匀液。每递增稀释一次，换用１次１ｍＬ无菌吸管或吸头。
６．１．５ 根据对样品污染状况的估计，选择１个～３个适宜稀释度的样品匀液（液体样品可包括原液），吸
取１ｍＬ样品匀液于无菌培养皿内，每个稀释度做两个培养皿。同时，分别吸取１ｍＬ空白稀释液加入
两个无菌培养皿内作空白对照。
６．１．６ 及时将１５ｍＬ～２０ｍＬ冷却至４６℃～５０℃的平板计数琼脂培养基（可放置于４８℃±２℃恒温
装置中保温）倾注培养皿，并转动培养皿使其混合均匀。
６．２ 培养
６．２．１ 水平放置待琼脂凝固后，将平板翻转，３６℃±１℃培养４８ｈ±２ｈ。水产品３０℃±１℃培养
７２ｈ±３ｈ。如果样品中可能含有在琼脂培养基表面蔓延生长的菌落，可在凝固后的琼脂培养基表面覆
盖一薄层平板计数琼脂培养基（约４ｍＬ），凝固后翻转平板，进行培养。
６．２．２ 如使用菌落总数测试片，应按照测试片所提供的相关技术规程操作。
６．３ 菌落计数
６．３．１ 可用肉眼观察，必要时用放大镜或菌落计数器，记录稀释倍数和相应的菌落数量。菌落计数以
菌落形成单位（ｃｏｌｏｎｙｆｏｒｍｉｎｇｕｎｉｔ，ＣＦＵ）表示。
６．３．２ 选取菌落数在３０ＣＦＵ～３００ＣＦＵ之间、无蔓延菌落生长的平板计数菌落总数。低于３０ＣＦＵ
的平板记录具体菌落数，大于３００ＣＦＵ的可记录为多不可计。
６．３．３ 其中一个平板有较大片状菌落生长时，则不宜采用，而应以无较大片状菌落生长的平板作为该
稀释度的菌落数；若片状菌落不到平板的一半，而其余一半中菌落分布又很均匀，可计算半个平板后乘
以２，代表一个平板菌落数。
６．３．４ 当平板上出现菌落间无明显界线的链状生长时，则将每条单链作为一个菌落计数。
７ 结果与报告
７．１ 菌落总数的计算方法
７．１．１ 若只有一个稀释度平板上的菌落数在适宜计数范围内，计算两个平板菌落数的平均值，再将平
均值乘以相应稀释倍数，作为每ｇ（ｍＬ）样品中菌落总数结果，示例见Ｂ．１。
７．１．２ 若有两个连续稀释度的平板菌落数在适宜计数范围内时，按式（１）计算，示例见Ｂ．２。
７．１．３ 若所有稀释度的平板上菌落数均大于３００ＣＦＵ，则对稀释度最高的平板进行计数，其他平板可
记录为多不可计，结果按平均菌落数乘以最高稀释倍数计算，示例见Ｂ．３。
７．１．４ 若所有稀释度的平板菌落数均小于３０ＣＦＵ，则应按稀释度最低的平均菌落数乘以稀释倍数计算，示例见Ｂ．４。
７．１．５ 若所有稀释度（包括液体样品原液）平板均无菌落生长，则以小于１乘以最低稀释倍数计算，示例见Ｂ．５。
７．１．６ 若所有稀释度的平板菌落数均不在３０ＣＦＵ～３００ＣＦＵ之间，其中一部分小于３０ＣＦＵ或大于
３００ＣＦＵ时，则以最接近３０ＣＦＵ或３００ＣＦＵ的平均菌落数乘以稀释倍数计算，示例见Ｂ．６。
７．２ 菌落总数的报告
７．２．１ 菌落总数小于１００ＣＦＵ时，按“四舍五入”原则修约，以整数报告。
７．２．２ 菌落总数大于或等于１００ＣＦＵ时，第三位数字采用“四舍五入”原则修约后，采用两位有效数字，
后面用０代替位数；也可用１０的指数形式来表示，按“四舍五入”原则修约后，采用两位有效数字。
７．２．３ 若空白对照上有菌落生长，则此次检验结果无效。
７．２．４ 称重取样以ＣＦＵ／ｇ为单位报告，体积取样以ＣＦＵ／ｍＬ为单位报告。
GB 4789.2-2022
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National food safety standard - Microbiological examination
of food: Aerobic plate count
ISSUED ON: JUNE 30, 2022
IMPLEMENTED ON: DECEMBER 30, 2022
Issued by: National Health Commission of the PRC;
State Administration for Market Regulation.
Table of Contents
Foreword ... 3
1 Scope ... 4
2 Terms and definitions ... 4
3 Equipment and materials ... 4
4 Medium and reagents ... 5
5 Examination procedure ... 5
6 Operation steps ... 6
7 Result and report ... 8
Appendix A Medium and reagents ... 10
Appendix B Examples ... 12
National food safety standard - Microbiological examination
of food: Aerobic plate count
1 Scope
This Standard specifies the method for the determination of aerobic plate count in food.
This Standard applies to the determination of aerobic plate count in food.
2 Terms and definitions
2.1
Aerobic plate count
The total number of microbiological colonies formed in per g (mL) of test sample,
which is obtained after the food sample under test is processed and cultured under
certain conditions (such as culture medium, culture temperature, and incubation time,
etc.).
3 Equipment and materials
In addition to the routine sterilization and culture equipment in the microbiology
laboratory, other equipment and materials are as follows:
a) Constant-temperature incubator: 36 ℃±1 ℃, 30 ℃±1 ℃.
b) Refrigerator: 2 ℃~5 ℃.
c) Thermostat: 48 ℃±2 ℃.
d) Balance: Sensitivity is 0.1 g.
e) Homogenizer.
f) Oscillator.
g) Sterile straw: 1 mL (with 0.01 mL scale), 10 mL (with 0.1 mL scale) or
micropipette and tip.
h) Sterile conical flask: Capacity is 250 mL and 500 mL.
6.1.2 Liquid sample: Use a sterile straw to pipette 25 mL of sample; place it in a sterile
conical flask containing 225 mL of sterile phosphate buffer or sterile normal saline (an
appropriate number of sterile glass beads can be preset in the bottle); and mix
thoroughly. Or put it into a sterile homogeneous bag containing 225 mL of diluent. Use
a slap-type homogenizer to beat for 1 min~2 min, to make a 1:10 sample homogenate.
When the result is required to be the aerobic plate count per g of sample, operate
according to 6.1.1.
6.1.3 Use a 1 mL sterile straw or micropipette to draw 1 mL of the 1:10 sample
homogenate; along the tube wall, slowly inject it into a sterile test tube containing 9 mL
of diluent (be careful not to touch the surface of the diluent with the pipette or the tip).
Oscillate and mix on an oscillator, to make a 1:100 sample homogenate.
6.1.4 According to the operation in 6.1.3, prepare a 10-fold serial dilution of the sample
homogenate. For each incremental dilution, use a new 1 mL sterile straw or tip.
6.1.5 According to the estimation of the contamination status of sample, select 1 to 3
sample homogenates with appropriate dilution (liquid samples can include stock
solution). Pipette 1 mL of the sample homogenate into a sterile petri dish; make two
petri dishes for each degree of dilution. At the same time, respectively pipette 1 mL of
blank diluent; add it to two sterile petri dishes as blank control.
6.1.6 In a timely manner, pour 15 mL~20 mL of plate count agar medium cooled to
46 °C~50 °C (which can be kept in a thermostat at 48 °C±2 °C) into a petri dish; rotate
the petri dish to make it evenly mixed.
6.2 Culture
6.2.1 Place horizontally until the agar solidifies; turn the plate over; incubate at
36 °C±1 °C for 48 h±2 h. Aquatic products are cultured at 30 °C±1 °C for 72 h±3 h. If
the sample may contain colonies that spread and grow on the surface of the agar
medium, it is possible to cover the surface of the solidified agar medium with a thin
and culture.
6.2.2 If the test piece of aerobic plate count is used, it shall be operated in accordance
with the relevant technical regulations provided for the test piece.
6.3 Colony count
6.3.1 It is possible to use the naked eye to observe. If necessary, use a magnifying glass
or a colony counter, to record the dilution ratio and the corresponding number of
colonies. Colony counts are expressed in colony forming unit (CFU).
6.3.2 Select the plate with the colony number between 30 CFU and 300 CFU and no
spreading colony growth, to count the aerobic plate count. For plates with less than 30
recorded as incalculable.
6.3.3 When one of the plates has larger flaky colonies, it is not suitable to use it. The
plate without larger flaky colonies shall be used as the colony count of the degree of
dilution. If the flaky colonies are less than half of the plate, and the colonies in the
remaining half are evenly distributed, it is possible to calculate the number of the half
plate and multiply it by 2, to represent the number of colonies on one plate.
6.3.4 When there is a chain growth with no clear boundary between the colonies on the
plate, count each single chain as a colony.
7 Result and report
7.1.1 If the number of colonies on only one dilution plate is within the appropriate count
range, calculate the average of the number of colonies on the two plates; then multiply
the average by the corresponding dilution factor as the aerobic plate count per g (mL)
of the sample. Example is given in B.1.
7.1.2 If the number of colonies on the plate with two serial dilutions is within the
appropriate count range, calculate according to formula (1). For an example, see B.2.
Where:
N - The number of colonies in the sample;
ΣC - The sum of the colony counts on the plates (containing plates with appropriate
n1 - Number of plates at the first dilution (low dilution ratio);
n2 - Number of plates at the second dilution (high dilution ratio);
d - Dilution factor (first dilution).
7.1.3 If the number of colonies on all dilution plates is greater than 300 CFU, the plate
with the highest dilution shall be counted. The other plates can be recorded as
uncountable. The result shall be calculated by multiplying the average number of
colonies by the highest dilution ratio. For an example, see B.3.
7.1.4 If the plate colony count of all dilutions is less than 30 CFU, it shall be calculated
Appendix A
A.1 Plate count agar (PCA) medium
A.1.1 Ingredients
Tryptone (main nutrient): 5.0 g
Yeast extract (main nutrient): 2.5 g
Glucose (main nutrient): 1.0 g
Agar: 15.0 g
Distilled water: 1000 mL
A.1.2 Preparation method
Add the above ingredients to distilled water; boil to dissolve; adjust the pH to 7.0±0.2.
A.2 Sterile phosphate buffer
A.2.1 Ingredients
Potassium dihydrogen phosphate (KH2PO4): 34.0 g
Distilled water: 500 mL
A.2.2 Preparation method
Stock solution: Weigh 34.0 g of potassium dihydrogen phosphate and dissolve it in 500
mL of distilled water; use about 175 mL of 1 mol/L sodium hydroxide solution to adjust
the pH to 7.2; use distilled water to dilute to 1000 mL; store in the refrigerator.
Diluent: Take 1.25 mL of the stock solution; use distilled water to dilute to 1000 mL;
A.3 Sterile normal saline
A.3.1 Ingredients
Sodium chloride: 8.5 g
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