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SN/T 4684-2016 English PDF (SNT4684-2016)

SN/T 4684-2016 English PDF (SNT4684-2016)

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SN/T 4684-2016: Determination of Burkholderia cepacia in cosmetics for import and export

This standard specifies the detection method of Burkholderia cepacia in cosmetics for import and export. This standard applies to the qualitative detection of Burkholderia cepacia in cosmetics for import and export.
SN/T 4684-2016
SN
ENTRY AND EXIT INSPECTION AND QUARANTINE INDUSTRY
STANDARD OF THE PEOPLE REPUBLIC OF CHINA
Determination of Burkholderia cepacia in cosmetics for
import and export
ISSUED ON: DECEMBER 12, 2016
IMPLEMENTED ON: JULY 01, 2017
Issued by: General Administration of Quality Inspection and Quarantine of PRC Table of Contents
Foreword ... 3
1 Scope ... 4
2 Normative references ... 4
3 Equipment and materials ... 4
4 Media and reagents ... 5
5 Inspection procedures ... 5
6 Operation procedures ... 6
7 Report of results ... 8
8 Waste disposal ... 8
Appendix A (Normative) Media and reagents ... 9
Determination of Burkholderia cepacia in cosmetics for
import and export
1 Scope
This standard specifies the detection method of Burkholderia cepacia in cosmetics for import and export.
This standard applies to the qualitative detection of Burkholderia cepacia in cosmetics for import and export.
2 Normative references
The following documents are essential to the application of this document. For the dated documents, only the versions with the dates indicated are applicable to this document; for the undated documents, only the latest version (including all the amendments) is applicable to this standard.
GB 7918.1 Standard methods of microbiological examination for cosmetics - General rules
GB 19489 Laboratories - General requirements for biosafety
3 Equipment and materials
In addition to the routine sterilization and culture equipment in the microbiology laboratory, other equipment and materials are as follows.
3.1 Constant temperature incubator: 30 ??C ?? 1 ??C, 32 ??C ?? 1 ??C, 36 ??C ?? 1 ??C. 3.2 Homogenizer.
3.3 Oscillators.
3.4 Optical microscope.
3.5 Balance: Sensitivity 0.1 g.
3.6 Sterile pipette: 1 mL, 10 mL (with 0.1 mL graduation) or micropipette and tip. 3.7 Sterile petri dish: 90 mm in diameter (or other commercial disposable sterile petri dish).
3.8 Inoculation loop: 3 mm diameter.
3.9 Homogenizing bag or homogenizing bottle.
3.10 Automatic microbial biochemical identification system.
4 Media and reagents
4.1 SCDLP enrichment solution containing polymyxin B: See A.1.
4.2 BCSA enrichment solution containing gentamicin: See A.2.
4.3 BCSA agar containing polymyxin B and gentamicin: See A.3.
4.4 MacConkey agar containing polymyxin B and gentamicin (without crystal violet): See A.4.
4.5 Gram stain: See A.5.
4.6 Reagent for oxidase test: See A.6.
4.7 Glucose oxidation/fermentation tube: See A.7.
4.8 Sucrose oxidation medium: See A.8.
4.9 Simon's citrate medium: See A.9.
4.10 Esculin medium: See A.10.
4.11 Biochemical identification kit.
5 Inspection procedures
See Figure 1 for the inspection procedure of Burkholderia cepacia.
6.3.1 Initial screening
Pick more than 5 typical or suspicious colonies from the selective agar plate. Inoculate the same strain into two glucose oxidation/fermentation tubes, one of which is sealed with sterilized liquid paraffin, whilst the other is not sealed. Culture it at 36 ??C ?? 1 ??C for 24 h. If both of them turns yellow, it is the glucose fermentative type. However, if the tubes without paraffin turns yellow but the tubes with paraffin does not change color, it is the glucose oxidative type. At the same time, mark a line and purify it on nutrient agar plate. Incubate at 36 ??C ?? 1 ??C for 24 h. Select the pure culture of the oxidized form of glucose (it does not turn yellow when adding paraffin tube; it turns yellow when not adding paraffin tube), to continue the identification.
6.3.2 Gram staining
Pick smears of suspicious colonies, to perform Gram staining. When observed microscopically, Burkholderia cepacia is a Gram-negative non-bacillus.
6.3.3 Oxidase test
Use a glass rod or a disposable inoculation needle to pick a single characteristic colony. Spread it on a filter paper plate, which is wetted with oxidase reagent. Burkholderia cepacia is positive for retarded oxidase (purple, purple or dark blue); the reaction time is about 2 min ~ 10 min.
Note: Never use nickel/chromium materials in the experiment.
6.3.4 Catalase test
Use a glass rod or a disposable inoculation needle to pick a single characteristic colony. Place it in a clean test tube. Add 2 mL of 3% hydrogen peroxide solution (prepared just before use) dropwise. Observe the generation of bubbles. Burkholderia cepacia produces gas slowly; bubbles appears slowly after 1 second.
6.3.5 Oxidation of sucrose
Pick suspicious colonies and inoculate them in sucrose oxidation medium. Incubate at 36 ??C ?? 1 ??C for 24 h. If the culture turns yellow, it is positive; if it turns blue, it is negative. Burkholderia cepacia is positive.
6.3.6 Utilization of citrate
Pick suspicious colonies and inoculate them in Simon's citrate medium. Incubate at 36 ??C ?? 1 ??C for 24 hours. If the medium turns blue, it is positive; if it is light green, it is negative. Burkholderia cepacia is positive.
6.3.7 Utilization of esculin
Pick suspicious colonies and inoculate them in esculin medium. Incubate at 30 ??C ??

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