To evaluate the application of metagenomic sequencing in identification and tracking of drug-contaminating bacteria.

Both metagenomic sequencing and isolation culturing method were employed to identify and genotype bacteria in two batches of possibly contaminated gel medicines as well as the environmental samples from testing laboratory. Bacteria isolated by culturing method were identified through Gram’s stain，biochemical reaction，MALIDI-TOF-MS，16srDNA sequencing and Bacillus specific PCR. The detected Bacillus cereus were typed by multilocus sequence typing (MLST) .The metagenomic sequencing results were analyzed at the strain level using MetaPhlan software.

Both MLST and MetaPhlan analysis revealed a substantial contamination of Bacillus cereus in gel medicines as well as environmental samples，with the presence of the same Bacillus cereus group. Both metagenomic sequencing and culturing methods detected Paenibacillus，Brevibacillus and Neobacillus in gel medicines，but amplicon sequencing also detected these bacteria in environment which might be the source of contamination.

These findings indicate that metagenomic sequencing technology is reliable and has the advantage over isolation culturing in comprehensively detecting potentially hazardous bacterial species and accurately performing strain-level traceability analysis. It holds significant application value in pharmaceutical contamination identification and traceability.

To classify the microbiological examination methods of live microecological products according to the results of suitability test for the enumeration method，and establish the decision-making method for microbiological examination of live microecological products.

The suitability test of the microbiological examination method of live microecological products was performed according to the requirements of the General Chapter 1105 in the Chinese Pharmacopoeia Vol Ⅳ. Escherichia coli，Staphylococcus aureus，Bacillus subtilis，Pseudomonas aeruginosa，Candida albicans and Aspergillus niger were used as the test strains. Tryptic Soy Agar，Sabouraud Dextrose Agar，Rose Bengal Agar and Rose Bengal Agar Ⅱ were used as medium.

For live microecological products composed of anaerobic bacteria or microorganisms with growth characteristics significantly different from the test strains，the suitability test for the method could be performed according to General Chapter 1105. For live microecological products containing Bacillus and enterococcus，the suitability test for the fungal enumeration method could be performed according to General Chapter 1105. Live microecological products could not be tested for suitability of the method due to the influence of the component microorganisms，gram-negative bacteria could be controlled using selective medium. In addition，some potentially harmful microorganisms could be controlled based on risk assessment according to the route of administration and the population of drug users.

This study provides a basis for the selection of the microbiological examination methods for live microecological products，and further complements and improves the quality standard system of live microecological products.

To establish a new HPLC method for the determination of the related substances in tranexamic acid injection by screening new type of HPLC columns，which can avoid the use of ion pair reagents in current methods and reduce salt concentration.

The method was carried out using a Thermo Mixed-mode WCX column (150 mm×4.6 mm，5 μm)，the mobile phase consisted of 10 mmol·L^-1 sodium dihydrogen phosphate solution (adjust the pH value to 5.2±0.05 with sodium hydroxide solution)-water-acetonitrile （50:5:45），the flow rate was 1.0 mL·min^-1，the column temperature was 25 ℃，the detection wavelength was 210 nm and the injection volume was 20 μL.

The chromatographic peaks of tranexamic acid and its four impurities B，C，D and E were all separated. Good linear relationship was shown between the concentration of all the five compounds and their corresponding peak areas (r≥0.999). The LODs were 0.34，0.50，0.005 6，0.002 1，0.12 μg·mL^-1. The average recoveries (n=9) were 97.4%，100.5%，98.4% and 96.6% with RSDs of 3.9%，0.24%，0.52% and 1.4%，respectively. The test solution and standard solution were all stable within 22 h. The detection results of 5 batches of tranexamic acid injection showed that the number of unspecified impurities and the total impurities content by using the new method were better than the current ChP method.

The established method is specified and sensitive. Good separation can be achieved with low concentration of phosphate. Its applicable to the determination of related substances in tranexamic acid injection.

To establish an inductively coupled plasma mass spectrometry (ICP-MS) method for the determination of migration Co，As，Br，Cd，Sb，Ba and Pb in recombinant human coagulation factor Ⅷ-Fc fusion protein for injection，and to quantitatively analyze the compatibility of long-term stored samples.

After adding 0.75 mL of water to the sample for 4 h，the purified water was filled to 5 mL，and the contents of 7 elements in the solution were determined by standard addition method. The working parameters of ICP-MS：radio frequency (RF) power of 1 550 W，sampling depth of 10.0 mm，carrier gas flow rate of 0.50 L·min^-1，atomizer pump speed of 0.30 r·s^-1，atomization chamber temperature of 2 ℃，dilution gas flow rate of 0.74 L·min^-1，repeated sampling times of 3 times，analysis mode of full quantitative.

The linear relationship of each element was good in the concentration range of the detected mass，and the recoveries of the element added were 92.0%-101.8%. When applied to the determination of the samples at each accelerated placement point，the increased migration amounts were 43.2，68.7 and 66.8 ng·mL^-1，respectively，except the element content of Br increased slightly. The contents of the remaining 6 elements were far below the prescribed limit.

The method is reliable and can be used for the determination of migration of 7 elements in recombinant human coagulation factor Ⅷ-Fc fusion protein for injection.

To establish an HPLC method for determination of the impurity profile in furosemide and its injection. At the same time，applying this method to detect and analyze products from many domestic and foreign enterprises，to evaluate the status quo of impurity control in API and injection，and the correlation of major degraded impurities in injection with prescription and packaging materials.

A YMC Hydrosphere C₁₈ column（250 mm×4.6 mm，3 μm）was used，and the mobile phase was 0.05% TFA solution (pH 2.23)-methanol-acetonitrile at the flow rate of 1.0 mL·min^-1，gradient elution. Detection wavelength was set at 238 nm and 277 nm and column temperature was 30 ℃. Impurity reference was used for localization and the relative retention time of each impurity was calculated. The known impurities were calculated using the principal component self-control method with correction factor，the unknown impurities were determined by principal component without correction factor.

The method for the determination of 11 kinds of known impurities，the potential genotoxic impurity in furosemide，its injection was established，the separation degree of all impurities met the requirements，and the sources of the impurities were identified by forced degradation test. The detected quantity of degraded impurity C and degraded impurity G in the injections produced by 2 to 3 companies exceeded the identification limit in ICH by 0.2%，the amount of potential genotoxic impurity furfural detected in the injection of 5 enterprises was much higher than that of the reference preparation，other impurities were the same as the reference preparation.

The established HPLC method can be used for rapid detection and analysis of impurities in furosemide and its injection. The impurity control level of injection produced by only 2 domestic enterprises is basically consistent with the reference preparation，others need to optimize prescription and packaging materials. This study can provide reference for improving the safety of furosemide injection and evaluating the quality consistency of generic drugs.

To establish a microbial limit test method and quality standard conforms to the USP for compound Kushen extract and its related components.

Comparative analysis was conducted on the standards for traditional Chinese medicine preparations and decoction pieces in the USP and the Chinese Pharmacopoeia 2020 Edition to determine the microbial limit standards for compound Kushen extract and its related components. Then，guided by the General Principles of the USP<61> and <62>，a microbial limit test method suitable for compound Kushen extract and its related components was developed. The plate pouring method was used to perform a methodological applicability test on the total aerobic bacterial count，mold and yeast count of compound Kushen extract and its two components Sophorae Flavescentis Radix and Heterosmilacis Rhizoma. The method applicability test was conducted on the control bacteria using the direct inoculation method. Finally，the MALDI/TOF MS microbial identification method and bacterial 16S rRNA gene sequence alignment analysis method were comprehensively used to identify the bacterial species of the contaminated microorganisms in the sample，and further analysis was conducted on the distribution and harm of the contaminated microbial population.

As a result，microbial limit standards for compound Kushen extract and its related components were established in accordance with the requirements of the USP. The recoveries of the counting methods for compound Kushen extract and its two components，Sophorae Flavescentis Radix and Heterosmilacis Rhizoma，were between 0.5 and 2.0. The positive control strains in the control bacterial method suitability test were able to grow，meeting the requirements of the USP. According to the identification results，40 contaminated microorganisms involved 13 species and 5 genera，mainly covering different types of Bacillus genus (87.5%)，Enterobacterium genus (10%)，and Pseudomonas genus (2.5%). Polluting microorganisms were all less harmful conditional pathogenic bacteria.

Established a microbial limit test method and quality standard conforms to the USP for compound Kushen extract and its related components. Defined the distribution and harm analysis of the pollution microorganism poulation of the above products. Improved the microbial quality control of the Chinese medicine preparation and its components.

To establish a fingerprint of Gualou Guizhi decoction and a method for determination of multi-component to clarify the transfer rule of quantities and quality from decoction pieces and substance benchmarks.

Fifteen batches of Gualou Guizhi decoction substance benchmarks were prepared and analyzed by the method of HPLC. Traditional Chinese medicine (TCM) Chromatographic Fingerprint Similarity Evaluation Software (2012) and hierarchical cluster analysis (HCA), principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) was used to evaluate the quality differences between batches of substance benchmarks, and screen the main chemical compositions that were led to quality discrepancy. The main differential components were determined by HPLC. And their contents, yields and transfer rates were analyzed for transfer rule of quantities and quality.

HPLC fingerprint of Gualou Guizhi decoction was established. A total of 30 common peaks in the fingerprint of Gualou Guizhi decoction substance benchmarks were assigned. Compared with the reference standards, 9 components were identified as gallic acid, albiflorin, paeoniflorin, liquiritin, ononin, liquiritigenin, cinnamic acid, isoliquiritigenin and glycyrrhizic acid. The similarities of 15 batches of Gualou Guizhi decoction substance benchmarks samples were all above 0.920. All of them were divided into two categories by three chemical recognition modes for 5 major differential components, including glycyrrhizic acid, liquiritigenin, paeoniflorin, iquiritin, and ononin. The HPLC was also applied to determine the contents of multi-components and the method validation results were good. The average transfer rates of five index components were 51.65%, 40.46%, 74.74%, 60.06%, and 34.54%, respectively and their average extraction rate was 14.20%.

The critical quality properties of Gualou Guizhi decoction can be stably transferred from decoction pieces to substance benchmarks. The method of fingerprint and content determination is accurate and reliable, which is able to provide technique for quality control of Gualou Guizhi decoction formulation.

To establish the RP-UPLC-PDA method for simultaneous determination of triptolide，triptonide, triptophenolide, wilforine, wilforlide A and celastrol in Tripterygii Radix.

Tripterygii Radix were extracted with ethyl acetate and the extracts were dissolved and separated by methanol. The six components were determined by RP-UPLC-PDA method. The chromatographic column was Acquity^TM UPLC BEH C₁₈ column (100 mm×2.1 mm, 1.7 μm), the column temperature was 30 ℃, the injection volumn was 2 μL, the flow rate was 0.4 mL·min^-1 and the mobile phase was acetonitrile (A)-0.1% formic acid (B).

The linear relationship of six components was good (0.999 2≤r≤0.999 7) in the concentration ranges. The average recoveries were 99.2%-103.1% and RSDs were 1.2%-2.9%. The contents in 10 batches of Tripterygii Radix from different habitat were determined. The results showed that the contents of Tripterygii Radix in prepared pieces from different producing areas were different. The highest content of triptolide was 140.2 μg·g^-1, and the lowest content was 103.2 μg·g^-1. The highest content of triptonide was 224.7 μg·g^-1 and the lowest content was 112.2 μg·g^-1. The highest content of triptophenolide was 306.7 μg·g^-1 and the lowest content was 189.6 μg·g^-1. The highest and lowest contents of wilforine were 283.2 μg·g^-1 and 211.2 μg·g^-1. The highest content of wilfhabitat was 31.2 μg·g^-1 and the lowest content was 16.8 μg·g^-1. The highest content of celastrol was 87.6 μg·g^-1, and the lowest content was 52.1 μg·g^-1.

The RP-UPLC-PDA method can simultaneously determine six components in Tripterygii Radix. The method is reliable and stable, which is suitable for quantitative analysis and determination of Tripterygii Radix.