1110 Biological tests
1101 Sterility test
Sterility test is a method for determining
whether sterile drugs, biological products,
medical instruments, raw materials, accessories
and other varieties that are required to be sterile
in the pharmacopoeia are sterile. If the sample
complies with the requirements of sterility test, it
only indicates that no microbial contamination is
found under the test condition.
The sterility test should be conducted under
the sterile condition, the test environment must
meet the requirements of sterility test, and the
whole testing process should strictly follow the
sterile procedures to prevent microbial
contamination, and the measures to prevent
contamination should not affect the detection of
microorganisms in the sample.
Medium
Thioglycollate fluid medium is mainly used
for culturing anaerobic bacteria and it can also
be used for culturing aerobic bacteria; trypsin
liquid medium is used for culturing fungi and
aerobic bacteria.
Preparation and culturing conditions of
medium
The medium can be prepared according to
the following prescription, and dehydrated
medium or prepared medium which is produced
according to the prescription and meets the
specification can also be used. After preparation,
a qualified sterilization procedure should be
conducted for the sterilization. The finished
medium should be stored in a dark place with the
temperature of 2°C to 25°C, if stored in an
unsealed container, generally it should be used
within 3 weeks; and if stored in an airtight
container, it can be used within a year.
- Thioglycollate fluid medium
Casein tryptone 15.0g Sodium chloride 2.5g
Bacto-extract 5.0g
Newly prepared 0.1% resazurin solution 1.0ml
Anhydrous glucose 5.0g
L-cystine 0.5g Agar 0.75g
Sodium thiosulfate 0.5g Water 1000ml
(Or thioglycollic acid) (0.3ml)
Except for glucose and resazurin solution,
mix the above ingredients, dissolve them at a
low temperature, adjust pH to alkalescence,
boiling and filtering. Add glucose and resazurin
solution, shake them well and adjust pH to 7.1
土 0.2 at 25°C after sterilization. Then divide it
into suitable containers, and the proportion of
the quantity and the height of the container
should be in accordance with that the oxide
layer (pink) of the medium after culturing is not
exceed 1/2 of its depth. Sterilizing. Before
inoculation, the height of the oxide layer of the
medium should not exceed 1/5 of the depth of
the medium, otherwise it has to be heated in
water bath at 100°C until the pink disappears
(not more than 20 minutes), and cools rapidly, it
should be heated only once and be protected
from contamination.
Unless otherwise specified, the
thioglycollate fluid medium should be cultured
between 30°C and 35°C. - Trypsin liquid medium
Casein tryptone 17.0g Sodium chloride 5.0g
Soybean papain hydrolysate 3.0g
Dipotassium hydrogen phosphate 2.5g
Glucose / Anhydrous glucose 2.5g / 2.3g
Water 1000ml
Except for glucose, mix the above
ingredients, dissolve them at a low temperature,
filtering, adjust pH to 7.3 土 0.2 at 25°C after
sterilization, add glucose, subpackaging,
sterilizing.
Trypsin liquid medium should be cultured
between 20°C and 25°C. - Neutralizer or inactivated medium
According to the prescription and
preparation method of the thioglycollate fluid
medium or trypsin liquid medium, appropriate
neutralizer, inactivator or surfactant can be added
before the medium is sterilized or used, the
dosage can refer to the method applicability test. - 0.5% broth medium (used for sterility
test of antibiotics like streptomycin sulfate)
Peptone 10g Sodium chloride 5.0g
Beef extract 3.0g Water 1000ml
Glucose 5.0g
Except for glucose, mix the above
ingredients, dissolve them at a low temperature,
adjust pH to alkalescence, boiling, add glucose
and after the dissolution, shake them well,
filtering, adjust pH to 7.2 土 0.2 at 25°C after
sterilization, subpackaging, sterilizing. - Trypsin soy agar medium
Casein tryptone 15.0g Agar 15.0g
Soybean papain hydrolysate 5.0g
Water 1000ml
Glucose 5.0g
Except for agar, mix the above ingredients,
dissolve them at a low temperature, adjust pH to
7.3 土 0.2 at 25°C after sterilization, add agar
and after heating and melting, shake them well,
subpackaging, sterilizing. - Sabouraud glucose liquid medium
Equivalent mixture of animal tissue pepsin
hydrolysate and casein tryptone 10.0g
Water 1000ml
Glucose 20.0g
Except for glucose, mix the above
ingredients, dissolve them at a low temperature,
adjust pH to 5.6 土 0.2 at 25°C after
sterilization, add glucose, shake them well,
subpackaging, sterilizing. - Sabouraud glucose agar medium
Equivalent mixture of animal tissue pepsin
hydrolysate and casein tryptone 10.0g
Agar 15.0g Water 1000ml
Glucose 40.0g
Except for glucose and agar, mix the above
ingredients, dissolve them at a low temperature,
adjust pH to 5.6 土 0.2 at 25°C after
sterilization, add agar and after heating and
melting, add glucose, shake them well,
subpackaging, sterilizing.
Applicability test for the medium
The thioglycollate fluid medium and trypsin
liquid medium that are used for sterility test
should conform to the requirements of sterility
test and sensitivity test of the medium.
Sterility test To randomly select no less
than 5 medium (bottles) from each batch, and
culture them at the specified temperature for 14
days with sterility
