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ObjectiveTo establish microbial limit test methods for 44 pediatric drugs. MethodsAccording to the general guidelines in Chinese Pharmacopoeia （2015 and 2020 edition， volume Ⅳ），a suitability test of the methods for 44 drugs was carried out by pour-plate method， neutralization method or dilution method. ResultsTotal aerobic microbial count： chemical oral liquid samples can be tested by 1∶10 plate method；traditional Chinese medicine need to be neutralized firstly. Then oral liquids could be tested by 1∶10 plate method and 1∶100 plate method was used for granules. Total count of molds and yeasts： all the samples can be tested by the 1∶10 plate method. The recoveries of five test strains were between 0.5 and 2.0. The specified microorganisms were all detected in the test group， while not found in the negative control group. ConclusionThe microbial limit test methods for the 44 pediatric drugs are established and the results are reliable and can be used in the quality control．
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OBJECTIVE: To establish a method to determine the genotoxic impurity, N-nitroso-N-methyl-4-aminobytyric acid, in losartan potassium using high performance liquid chromatography triple quadrupole mass spectrometry (HPLC-MS/MS). METHODS: The method was developed by using Shimadzu Shim-pack XR-ODS II column (2.0 mm×150 mm, 2.2 μm). Time program was conducted with mobile phase consisting of water (0.1% formic acid, A) and methanol (B). The flow rate was 0.3 mL•min-1, and the column oven temperature was maintained at 40 ℃. The samples were ionized by electrospray ionization (ESI) with multi reaction monitoring (MRM) data acquisition mode. The collision energies were -11, -13, and -13 V, CID gas was argon with pressure of 270 kPa.3 pairs of precursor, and product ions (m/z) of NMBA were 147.15→117.10, 147.15→87.10, and 147.15→44.10, respectively. RESULTS: The genotoxic impurity NMBA showed linearity between 1 and 100 ng•mL-1 with correlation coefficient of 0.999 9. The intra-day and inter-day repeatability was examined by relative standard deviations (RSDs) of retention time and peak area (RSD<1.10%, n=6 for intra-day repeatability and n=18 for inter-day repeatability). The accuracy was examined by percent recovery at three concentration levels, and the average percent recovery was between 94.40% and 98.04%. CONCLUSION: The established LC-MS/MS method is efficient for limit test and quantitation of NMBA in losartan potassium bulk drug.
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Objective To establish limit test method of camphor in Niuhuang qingxin pills (prescription of the bureau) . Methods The separation was performed on an HP-INNOWAX capillary column (0.25 mm×30 m, 0.25 μm) and the column temperature was 110 ℃ . The temperatures of the inlet and the FID detector were 200 and 230 ℃ , respectively. The flow rate of the carrier gas was 1.8 mL·min-1 and the split ratio was 10:1. The injection volume was 1 μL. Results Good linearity was obtained for camphor within the range of 1.65 to 165 μg·mL-1, and the average recovery of low, medium, high concentration were 99.12％, 99.56％ and 99.56％, respectively,with RSD were 1.01％,0.69％ and 0.38％, respectively. Trace camphor was found in 7 samples out of 38 samples from 9 manufactures. Conclusion The proposed method was accurate, sensitive and simple, and was suitable for quality evaluation and safety control of Niuhuang qingxin pills (prescription of the bureau) .
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Objective:To investigate the microbial contamination status of Chinese herbal pieces from Guangxi regions and provide reference for the microbial limit standard. Methods:According to Chinese Pharmacopoeia (2015 edition), the herbal pieces were analyzed for the total number of aerobe microbial, yeasts, molds, thermophilic bacteria and bile-resistant gram-negative bacteria,and salmonella was also detected. Results:In 90 batches of Chinese herbal pieces,the lg average value of the total number of aerobe microbial was 3.7,and that of the total yeasts and molds was 1.7,that of thermophilic bacteria was 0.9 and that of bile-resistant gram-negative bacteria was 1.1,and Salmonella was not detected out. There were statistically significant differences (P < 0.05) in the microbial contamination levels of herbal pieces with different processing techniques and packaging materials. Conclusion:According to different processing methods and administration, microbial limit standard should be established for classification in order to guide the standardized production of Chinese herbal pieces and improve their quality gradually.
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Objective: To establish the quality standard for Fuzheng capsules. Methods: TLC was adopted to identify Astragali Radix and Glycyrrhiza uralensis. The method validation for Fuzheng capsules was conducted by microbial limit test as described in the appendixes of Chinese Pharmacopeia (2015 edition). The content of epimedii in Epimedium brevicornu was determined by HPLC. The chromatographic separation was carried out on an Agilent TC-C18(2) (250 mm×4. 6 mm, 5 μm) column. The mobile phase consis-ted of acetonitrile-water( 30: 70) with gradient elution at a flow rate of 1. 0 ml·min-1,and the injection volume was 5 μl. The detec-tion wavelength was 270 nm. Results: The spots in TLC were clear without any interference. The methods of plating and direct inocu-lation could be used for the microbial limit test. The linear range was 0.101-1.008 μg (r =0.999 7). The average recovery was 99. 36% with the RSD of 0. 81% (n=6). Conclusion: The method is simple with high specificity and good repeatability, which can be used as the quality control method for Fuzheng capsules.
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Objective:To eliminate the interference of matrix in urea cream on the microbial limit test. Methods: A certain a-mount of magnesium sulfate was added to the test solution followed by stirring and placing sometime to make the cream layering, and the water layer was used for the microbial limit detection. Results:After magnesium sulfate was added to urea cream, microbial limit test was not affected by the matrix. The average recovery of Escherichia oli, Staphylococcus aureus, Bacillus subtilis, Candida albicans and Aspergillus niger was 96. 00%, 97. 00%, 100. 00%, 99. 00% and 101. 00%, respectively. The negative and positive results of Psudomonas aeruginosa and Staphylococcus auress were reliable. Conclusion:In the detection of microbial limit for urea cream, mag-nesium sulfate can be used as a good auxiliary agent to make the detection method more convenient and reliable.
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Objective:To investigate the neutralization of lecithin and polysorbate-80 in antibacterial activity of oral solid tradition-al Chinese medicine preparations without Chinese medicinal herb powder. Methods:According to the requirements in Chinese Pharma-copeia (2015 edition), the applicability of microbial counting method was tested in the diluent containing lecithin and polysorbate-80 and the conventional diluent, respectively, and the results were compared. Results: The recovery results of applicability of microbial counting method using neutralizers (lecithin and polysorbate-80) conformed to the standard of Chinese pharmacopoeia (2015 edition). Conclusion:The combination of lecithin and polysorbate-80 has notable neutralization in antibacterial activity of oral solid traditional Chinese medicine preparations without Chinese medicinal herb powder.
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Objective:To establish the microbial limit test methods for three preparations containing chlorhexidine acetate. Meth-ods:The validation of microbiological limit test for three preparations including chlorhexidine acetate solution, chlorhexidine acetate ointment and chlorhexidine acetate cream was carried out respectively using the conventional method,dilution method,membrane filtra-tion method and neutralization method. Results:The recovery rates of five strains in the verification of counting method by membirane filtration and nearailzation method were more than 50%. Every positive test micrcorganism could be detected while negative control bacteria could not grow in the examination of control bacteria.Conclusion:The membrane filtration method can be used for the deter-mination of control bacteria for chlorhexidine acetate solution and chlorhexidine acetate ointment,and the neutralization method can be applied in the determination of control bacteria for chlorhexidine acetate cream.
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Objective To establish a method for the microbial limit test of zinc oxide and talcum powder lotion in Chinese Pharmacopoeia 2015.Methods Microbial enumeration test and specified microorganisms test with instructions were conducted from Chinese Pharmacopoeia 2015,which involved 10 batches zinc oxide and talcum powder lotion and five species of bacteria in all.The samples were treated by centrifugation and membrane filtration.Microbial enumeration test:the total number of aerobic bacteria using TSA medium to examine,strains were Staphylococcus aureus,Pseudomonas aeruginosa,Bacillus subtilis,Candida albicans,and Aspergillus niger;The total number of molds and yeasts using SDA culture medium to examine,strains were Candida albicans and Aspergillus niger.Each strain was divided into two groups:Adding bacteria test group and bacteria control group.Samplecontrol group and negative control group of each culture medium was prepared respectively.Count the colonies and calculate the rate of recovery.Specified microorganisms test:Pseudomonas aeruginosa and Staphylococcus aureus were applicated and set the bacteria validation group,sample control group,and negative control group.The culture medium of each group was crossed on the corresponding medium plate,and Microflex LT mass spectrometer was used to identify the bacteria.Results The recoveries of all kinds of strains in microbial enumeration test,total yeast and mold count were 0.75-1.16 in all batches.In the validations of Staphylococcus aureus and Pseudomonas aeruginosa,all kinds of strains were respectively detected in bacteria validation group,and there were no bacterial growth in sample control group and negative control group.Conclusion The microbiological examination methods for zinc oxide and talcum powder lotion can meet the requirements of Chinese Pharmacopoeia 2015.
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Objective: To establish a microbial limit test method for compound benzocaine gel.Methods: According to the general principles of Chinese Pharmacopoeia (2015 edition), method applicable experiments were performed respectively for the routine method, neutralization method and dilution & neutralization method.Using the recovery ratio of test bacteria as the index, the medium of dilution & neutralization method was adopted in the total aerobic microbial count and total combined yeasts and molds count, and the neutralization method was used for the control bacteria detection.The neutralizing agents were polysorbate 80 and lecithin.Results: The method of dilution & neutralization could eliminate the bacteriostasis of the drug, and the recovery ratio of each test bacteria was within the range of 0.5-2.0, which was in line with the requirements of Chinese Pharmacopoeia.Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus could be detected out in the positive control bacterial test,and bacteria were not detected out in the negative test.Conclusion: The method can be used for the microbial limit test for compound benzocaine gel.
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OBJECTIVE:To establish a method of microbial limit test for liquid preparation of Iron sucrose injection before filtration and sterilization.METHODS:According to the microbial limit test in the 1005 and general rules 1006 of 2015 edition of Chinese Pharmacopoeia (vol.Ⅴ),plate method and membrane filtration method were used to measure total number of aerobic bacteria (Staphylococcus aureus,Pseudomonas aeruginosa,Bacillus subtilis,Candida albicans,Aspergillus niger) and total number of molds and yeasts (C.albicans,A.niger).The optimal test method was obtained by comparing the bacterial recoveries.RESULTS:By plate method,the recoveries of P aeruginosa and B.subtilis were 2％ and 5％.The test sample was diluted 10 times with pH 7.0 sodium chloride-peptone buffer solution,and the bacterial recoveries were in the range of 88％ to 96％;but he medium was dark in color.By membrane filtration method,without rinse solution,the bacterial recoveries in the range of 88％ to 95％.Add rinse solution,the bacterial recoveries were in the range of 91％ to 103％.After validated,the recoveries of menbrane filtration method with tlushing fluid ranged 50％-200 ％,which was in line with the requnements.CONCLUSIONS:The membrane filtration method established in this experiment has higher bacterial recovery rate than the plate method.The bacterial recoveries rate were higher after adding rinse solution,and no dark substance in the surface of filter membrane affect the accotmt.It can be used as the microbial limit test method for preparation liquid of Iron sucrose injection before filtration and sterilization.
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Objective To establish the methods of microbial limit test for three kinds of preparations with heavy metals, such asDingxian Pills,Pizhi Lotion andJiawei Huangqin Ointment.Methods According to Chinese Pharmacopoeia 2010, the recovery rates of bacteria, fungus and yeast treated by three preparations were detected. And the methods of testing control bacteria were also validated.Results Culture medium dilution method was proved to be applicable forDingxian Pills. Culture medium dilution method combined with pre-filtration method was proper for Pizhi Lotion. And the extraction method was adopted forJiawei Huangqin Ointment. The recovery rates of these five validation strains reached 70% by appropriate methods. And the same methods were used for validation of the control bacteria.Conclusion The methods of microbial limit test for these three different preparations were established through this study.
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Objective To establish a method of microbial limit tests for compound Hongjingtian oral liquid and to per-form the validation of the method.Methods The validation on the microbial limit test method was conducted according to the counting method of bacteria,mycetes and yeasts and the control bacteria test method stated in the appendix of China pharmaco-poeia (2010 edition).Results The recoveries were over 70% when culture diluting method was used for staphylococcus aureus and routine test method was used for Escherichiacoli,Bacillus subtilis,Candida albicans and Aspergillus niger.In the test of positive control bacteria,the growth was detected,while the negative control was no growth detected.Conclusion The micro-bial limit test for compound Hongjingtian oral liquid is validated in this study.The amount of staphylococcus aureus can be de-termined using diluted culture method ;while the amounts of Escherichiacoli,Bacillus subtilis,Candida albicans,Aspergillus niger can be determined by conventional method.
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Objective:To establish a method for the microbial limit test of Jingzhi Guanxin tablets. Methods: According to the methods in the 2010 edition and 2015 edition of Chinese Pharmacopoeia, the test was performed respectively. Results:Jingzhi Guanxin tablets showed obvious inhibitory effect on Bacillus subtilis. The bacteria count could be carried out by the culture medium diluting methods in the 2010 edition. The total amount of aerobe could be detected by the membrane filtration method in the 2015 edition. The total combined molds and yeasts count could be performed by the plate count method and the specified microorganism could be tested with the routine method. Conclusion:The above methods can be used for the microbial limit test of Jingzhi Guanxin tablets.
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OBJECTIVE:To establish a method of microbial limit test for Ofloxacin gel. METHODS:Test sample solutions were prepared by MnSO4,CaCl2 and MgCl2 gelout with different concentrations,the numbers of bacteria and control bacteria were detected by membrane filtration method,and the numbers of molds and yeasts were determined by conventional method. RE-SULTS:The conventional method showed the recoveries of Candida albicaus and Aspergillus niger were more than 70% after not treated by gel breaker;the membrane filtration method showed the recoveries of Escherichia coli,Staphylococcus aurus and Baci-lus subtilis were more than 70% after treated by 5% MnSO4,3% CaCl2 and 1% MgCl2;the membrane filtration method could de-tect the control bacteria after treated by gel breaker. CONCLUSIONS:Gel breaker with appropriate concentrations can effectively eliminate the antibacterial activity of Ofloxacin gel. The established method is suitable for its microbial limit test.
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OBJECTIVE: To carry out standardization research on pre-filtration method for microbial test solutions of drugs and establish a new double-membrane filtration method in microbial tests. METHODS: New filters with double-membranes of different materials or pore-sizes were designed and used as pre-filters for microbial tests. The effects of test solution pre-filtration on several wild-type strains were evaluated and compared with the methods described by Chinese Pharmacopoeia. RESULTS: No statistical difference was found between the double-membrane filtration method and the pharmacopoeia method. However, filter-clogging was significantly reduced by using the double-membrane filtration method, and the double-membrane filtration method could also eliminate the antibacterial activity of the samples, thus reducing the damage or loss of microorganisms in special samples. CONCLUSION: The double-membrane filtration method meets the requirements of the Chinese Pharmacopoeia; the test results are valid and the method can be used for sterility test and microbial limit test for drug products.
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Objective:To establish the method validation of microbial limit tests for hospital paste preparations, including com-pound salicylic acid paste, zinc oxide paste and compound pine tar paste. Methods:According to the microbial limit test described in China pharmacopoeia 2010 edition, the method validation of count of bacteria, fungi and yeasts and tests for specified microorganisms was established. Results:Medium dilution method could be used in bacteria, fungi and yeasts count and specified microorganisms ex-amination for compound salicylic acid paste and zinc oxide paste. For compound pine tar paste, bacteria, fungi and yeasts count and the pseudomonas aeruginosa examination could use medium dilution method, while the staphylococcus aureus examination should employ membrane filtration method. Conclusion:The methods of microbial limit tests for the three hospital paste preparations are established, which can be used to control the quality of hospital preparations effectively.
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Objective:To establish the microbial limit test methods for thirteen kinds of ointments. Methods:The microbial limit of 13 kinds of ointments was respectively determined by the routine method, culture medium dilution method and membrane filtration method. Results:The recovery of the tested bacteria in the samples was above 70% by the different methods. Conclusion:The micro-bial limit test methods for thirteen kinds of ointments are stablished, which may be used in the quality control.
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Objective To establish a method for microbial limit test of fast vertical liquid hand disinfectant gel .Methods The membrane filtration method, in accordance with the Chinese Pharmacopoeia 2010 Edition (part two) relevant requirements of microbial limit test method for validation of microbial limit test for fast vertical liquid hand disinfectant gel .Bacillus subtilis, Staphylococcus aureus, Escherichia coli, Candida a lbicans, Aspergillus niger were taken as test bacteria , Pseudomonas aeruginosa and Staphylococcus aureus were taken as control bacteria .Verification test was carried out in parallel test for 3 independent , recoveries and every trail rate were calculat-ed.Results In bacteria, molds and yeasts count method validation test , recovery test strain rate , the control bacteria test results met the requirements.Conclusion The membrane filtration method was feasible and effective , which could be used for microbial limit test of fast vertical liquid hand disinfectant gel .
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OBJECTIVE: To provide a method for the microbial limit test of Anshenning oral liquid and carry out validation of the mothed. METHODS: The validation on the microbial limit test method was conducted in accordance with the counting method of bacteria, mycetes and yeasts and the control bacteria test method stated in the Appendix of China Pharmacopoeia(2010 edition). Culture dilution method was used in bacterium count and total combined molds and yeasts count; while routine method was used in the control bacteria test. RESULTS: The recoveries of test organisms in the control group 3 preparations by routine method were more than 70%. The recoveries of the test group were more than 70%. In the 3 control bacteria tests the negative control bacterias, was not found in the negative control group, while the control bacteria was found in the test group. CONCLUSION: The established method substances can eliminate the antimicrobial in Anshenning oral liquid is accurate, and comprehensive, and can be used for the microbial limit test of Anshenning oral liquid.