To study and establish a method based on gas chromatography and chemometrics techniques for distinguishing Artemisiae Argyi Folium and its adulterants Artemisiae Mongolica Folium.

Gas chromatography method was established with Agilent HP-5 19091J (30 m×0.32 mm, 0.25 μm) as chromatographic column, and hydrogen flame ion detector (FID) as detector. After the chemical composition of 21 chromatographic peaks in the chromatogram were identified, and the peak area data of the 21 chromatographic peaks in 29 batches of samples were determined. Similarity analysis, correlation analysis, cluster analysis, principal component analysis and orthogonal partial least squares-discriminant analysis were applied to analyze the chromatographic data.

The results of chemometric analysis indicated that tpeak 20 (chamazulene), peak 3 (1, 8-cineole) and peak 19((1S, 8aα)-decahydro-1, 4aβ-dimethyl-7β-isopropenyl-1-naphthol) were the differential characteristic chromatographic peaks between Artemisiae Argyi Folium and its adulterants Artemisiae Mongolica Folium. The ratios of the peak areas of peak 3 to peak 20 were in the ranges of 54.50-348.39 and 0.16-0.87 respectively, and the ratios of the peak areas of peak 19 to peak 20 were in the ranges of 18.55-128.46 and 0.01-0.14 respectively. These significant differences could be used for the identification of Artemisiae Argyi Folium and its adulterant Artemisiae Mongolica Folium.

The research findings can be used for the identification of Artemisiae Argyi Folium and its adulterant Artemisiae Mongolica Folium, and these have certain reference significance for the research and analysis of Artemisiae Argyi Folium and related drugs.

To establish a method for quality evaluation of Artemisiae Scopariae Herba [Artemisia capillaris Thunb. (Mianyinchen)] dispensing granules by combining characteristic chromatogram, quantitative analysis of multi-components by single marker (QAMS) and chemical pattern recognition analysis.

The high performance liquid chromatography (HPLC) characteristic chromatogram was established by 15 batches of Artemisiae Scopariae Herba [Artemisia capillaris Thunb. (Mianyinchen)] standard decoctions and 3 batches of dispensing granules. The contents of 6 components were determined by QAMS. The chromatographic separation was achieved on a Acclaim^TM RSLC 120 C₁₈ column (100 mm×2.1 mm, 2.2 μm), with the mobile phase comprising of acetonitrile -0.05% phosphoric acid flowing at 0.4 mL·min^-1 in a gradient elution manner. And the detection wavelength was set at 327 nm. The similarity evaluation system of fingerprint of traditional Chinese medicine was used to determine the common peak for similarity evaluation. Hierarchical cluster analysis (HCA) and principal component analysis (PCA) were applied for chemical pattern recognition. The transfer rates of the 6 components from decoction pieces to standard decoctions and dispensing granules were calculated.

The similarities of characteristic chromatograms of 15 batches of Artemisiae Scopariae Herba [Artemisia capillaris Thunb. (Mianyinchen)] standard decoctions and 3 batches of dispensing granules were all above 0.85. And 8 common characteristic peaks were identified. The results of HCA and PCA indicated the similarity of ingredients in formula granules to those in standard decoctions. The contents of neochlorogenic acid, chlorogenic acid, cryptochlorogenic, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C in standard decoctions were 1.87-5.23, 7.44-15.26, 2.85-8.18, 3.05-6.14, 0.99-3.93 and 3.23-10.38 mg·g^-1 and the transfer rates of these components from decoction pieces to standard decoction were 23.85%-37.28%, 19.57%-31.93%, 28.15%-45.88%, 22.34%-36.59%, 16.64%-28.36% and 21.81%-39.19%, respectively. The contents and transfer rates of these 6 compounds in dispensing granules were close to that of standard decoctions.

The characteristic chromatogram and QAMS method established can be used for quality control and process research of Artemisiae Scopariae Herba [Artemisia capillaris Thunb. (Mianyinchen)] dispensing granules.

To evaluate the chemical composition difference of Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves by high-resolution mass spectrometry and omics analysis, so as to establish the detection method of Cassia auriculata L. leaves adulterated in Cassia angustifolia Vahl leaves.

The positive and negative MS^E data of 13 batches of Cassia angustifolia Vahl leaves and 9 batches of Cassia auriculata L. leaves were collected by ultra performance liquid chromatography coupled with electrospray ionization quadrupole time of flight mass spectrometry(UPLC-Q TOF MS). The mobile phase was acetonitrile(A)-1% acetic acid(B). The column temperature was 30 ℃. The flow rate was 0.3 mL·min^-1. Injection volume was 1 μL. The mass range was m/z 50-1 200. The chemical composition difference of Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves were processed by the omics analysis QI software based on the orthogonal partial least-squares discrimination analysis(OPLS-DA) after the negative MS^E data were obtained. One out of seven specific components from Cassia auriculata L. leaves was separated and identified. A method with the specific component as the reference substance for the detection of adulterated Cassia auriculata L. leaves in Cassia angustifolia Vahl leaves was established by UPLC, and it was used in 27 batches of Cassia angustifolia Vahl leaves samples and 3 batches of laboratory-made positive samples.

Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves were significantly different from each other. The specific component separated from Cassia auriculata L. leaves was identified as kaempferol 3-O-(2”-O-apiofuranosyl) rutinoside. In the method for the detection of adulterated Cassia auriculata L. leaves in Cassia angustifolia Vahl leaves by UPLC, the precision(RSD=1.3%), repeatability(RSD=1.3%) and stability(RSD=0.58%) met the requirements. No kaempferol 3-O-(2”-O-apiofuranosyl) rutinoside was detected in 23 out of 27 batches of Cassia angustifolia Vahl leaves samples, but it was detected in 4 batches of Cassia angustifolia Vahl leaves samples and 3 batches of positive samples made in the laboratory.

In this study, the difference of Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves is distinguished clearly based on the technology of UPLC-Q TOF MS and OPLS-DA. The specific component of Cassia auriculata L. leaves is separated and identified. A method for the detection of adulterated Cassia auriculata L. leaves in Cassia angustifolia Vahl leaves is established by UPLC, and it provides the basis for quality control and quality standard improvement of Cassia angustifolia Vahl leaves. The technology is helpful to solve the problem of adulteration identification of traditional Chinese medicine.

To perform a comprehensive analysis of extractables in rubber by pseudotargeted metabolomics.

The quality control(QC) solutions and test solutions were prepared by extracting the six kinds of rubbers with water, pH 2.6 phosphate buffer, pH 9.18 phosphate buffer, and 15% ethanol solution, respectively. The QC solutions were detected by LC-30A ultra performance liquid chromatography-Q TOF-9030 mass spectrometer in ESI⁺/ESI^- mode. The mobile phases were 0.1% formic acid-water and 0.1% formic acid-acetonitrile/0.05% ammonium-water and 0.05% ammonium -5% water-methanol. The separation chromatographic column was Waters Acquity BEH C8/HSS T3. TOF MS and MS/MS DDA scanning modes were used to detect the QC solutions. The scan results of QC solution were processed with MSConvert and MRM-Ion_Pair_Finder software to obtain MRM parameters. The extracts were detected by ExionLC UPLC-Qtrap6500+ mass spectrometer in ESI⁺/ESI^- mode with the same mobile phase and column as before, and Scheduled-MRM scan mode was applied to detect the extracts.

The 1 399 unknown compounds and 116 known compounds were analyzed. By extraction solution, it was found that pH 9.18 phosphate buffer and 15% ethanol solution could extract more compounds, and 27 compounds with large differences in different extraction solutions were identified. By rubbers, the affinities of different rubbers were determined by 25 differences.

In this study, the results of the extraction of six rubbers under four extracts are analyzed. There are differences between extracts under different extraction conditions. Analytical methods can be specifically developed for these key variables for quality control follow-up studies to ensure the stability and consistency of drug quality. The variables in different rubbers are also analyzed, which can be used to make a general class determination of unknown rubber samples.

To explore the correlations between opalescence and molecular size, purity or aggregation tendency in the antibody formulation.

Studied on the opalescence phenomena of various antibodies, including the influence of different concentrations, pH/buffer, excipients, as well as the influence of environmental factors such as temperature, light, freeze-thaw and agitation, and the correlations had been analyzed between opalescence and molecular size, purity (aggregation), K_D or T_agg.

The degree of antibody opalescence was positively correlated with the molecular size, the deepening of the opalescence indicated that the molecular size was increased and the more tendency of molecular aggregation. For example, the opalescence of C molecule obviously enhanced under high temperature, and the result of turbidity changed from 3.3 NTU to 13.6 NTU, the molecular size changed from 13.1 nm to 40.6 nm, and the K_D changed from positive to negative. The T_agg changed from 59.6 ℃ to below 50 ℃.

The opalescence can be affected by buffer system, pH, excipients and ambient temperature, the degree of opalescence increased, indicating the stability become worse and the molecules prone to aggregate. This study provides important reference for the analysis and evaluation of opalescence in the antibody formulation development.

To establish a rapid and accurate ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the detection of dextromethorphan and one metabolite dextrorphan in hair.

The hair sample containing dextromethorphan and dextrorphan were extracted with methanol containing internal standard proadifen hydrochloride(SKF_525A). The extract was filtered with 0.22 μm organic filter membrane and detected by UPLC-MS/MS. All components were separated by an ACQUITY UPLC HSS T3 column (100 mm×2.1 mm, 1.8 μm), using a gradient elution procedure consisting of 0.2% formic acid (10 mmol·L^-1 ammonium formate) and acetonitrile, at a flow rate of 0.3 mL·min^-1, and the column temperature was room temperature. Positive electrospray ionization was performed using multiple reaction monitoring mode (MRM).

The linear relationships of dextromethorphan and dextrorphan were good in the range of 1-100 ng·mL^-1. The linear equations were Y=1.349 49X-0.020 80 (r=0.998 8) and Y=0.775 10X-0.013 87 (r=0.999 1), respectively. The detection limit and quantitation limit were 0.010 ng·mL^-1 and 0.025 ng·mL^-1 respectively. Their recoveries ranged from 97.0%-104.8%. The intra-day RSD and inter-day RSD were 1.5%-3.9% and 2.1%-5.5%, respectively. The method was applied to cases, and the results showed that dextromethorphan and dextrorphan were detected in the hair of 6 abusers.

This method is simple and sensitive enough to be applied to detect dextromethorphan and one metabolite dextrorphan in hair.

To determine the contents of Cd and As in Isatidis Radix and to explore the application of target-organ toxicity dose(TTD) modification of hazard index (HI) method in assessing the risk of combined exposure to heavy metals and harmful elements in traditional Chinese medicines (TCMs).

According to the monitoring data of Cd and As in Isatidis Radix, the exposure doses were calculated. The health risk caused by combined exposure to Cd and As was preliminary screened by HI method. Moreover, a more accurate TTD method was used.

The qualified rates of Cd in 29 batches of Isatidis Radix were 100%. However, 5 batches of As contents exceeded the limit standard. The HQ values of As in 5 batches of Isatidis Radix were more than 1. The results of TTD method showed that for the end points of cardiovascular system, blood, nervous system, the HI values of 5 batches of Isatidis Radix were more than 1, and the health risk was not acceptable.

The cumulative risk assessment of heavy metals in TCMs is proposed in this study, which offers novel ideas for the development of risk assessment methods of exogenous harmful residues in TCMs, and provides technical support for formulating scientific limit standards.

To establish the first national standard for oncolytic activity assay of herpes simplex virus type 1(HSV-1).

According to the requirements in Chinese Pharmacopoeia(Volume Ⅲ, 2020 edition), the liquid and freeze-dried standard for oncolytic activity of HSV-1 were prepared and tested, of which the stability were evaluated by thermal acceleration test. The oncolytic activity of the standard was calibrated collaboratively by U-2 OS cells/CCK-8 method in 3 laboratories. The liquid standard was compared with the freeze-dried standard, and the more suitable one was selected as the national standard.

The prepared standard substance was all qualified, among which the moisture content of the freeze-dried standard was 1.09% and the dispensing accuracy was 0.15%. The results of stability test were calculated by Arrhenius formula. It was preliminarily predicted that it would take 7.7 years for the oncolytic activity of liquid standard to decrease by 10% at -70 ℃, and it would take 6.1×10⁵ years for the oncolytic activity of lyophilized standard to decrease by 10% at -70 ℃. Compared with liquid standard, the stability of lyophilized standard was greatly improved. Twenty-one times of collaborative calibration tests by 3 laboratories showed that the oncolytic activity liquid standard was 7.08×10⁴ U·mL^-1 and the oncolytic activity liquid standard was 1.82×10⁴ U·vial^-1. After lyophilized, the oncolytic activity of the bulk of HSV-1 standard decreased from 7.08×10⁴ U·mL^-1 to 3.03×10⁴ U·mL^-1. However, the good S-shaped dose-response curve still appeared after 100 times of pre-dilution, which did not affect the requirements of its use as a standard. When the liquid standard was used as the sample and the freeze-dried standard was used as the standard for calibration, the geometric coefficient of variation (GCV) of the results of the 3 laboratories decreased from 64.4% to 29.2%, and the precision of the experiment was greatly improved.

The batch of freeze-dried HSV-1 standards for oncolytic activity assay meets the relavant requirements, and is more suitable for use as a national standard than liquid standards. Its oncolytic activity is assigned a value of 1.82×10⁴ U·vial^-1.

To determine seven impurities in oxytocin for injection and investigate the limit values.

HPLC and principal component self-control with correction factor were adopted. The determination was performed on a Waters Xbridge C₁₈ column(150 mm×4.6 mm, 5 μm). The mobile phase consisted of 0.1 mol·L^-1 dihydrogen phosphate solution (adjusted to pH 5.4)-acetonitrile (90∶10, phase A), and acetonitrile (phase B) with gradient elution at a flow rate of 1.5 mL·min^-1. The column temperature was maintained at 32 ℃, and the detection wavelength was set at 220 nm. The injection volume was 100 μL. The linear equations of oxytocin, impurities Ac-Oxy, Oxy[Glu⁴], Oxy[+Gly¹⁰], Oxy[-NH₂], Oxy[trisulfide], Oxy[cis-dimer] and Oxy[trans-dimer] were drawn. The correction factors of each impurity related to oxytocin were calculated by slope. The contents of impurities in 3 batches of oxytocin for injection were determined and compared with the results of impurity reference method.

The limits of quantification for seven impurities were 2.75-5.66 ng, while the detection limits were 1.38-2.83 ng. The linear ranges of seven impurities were 0.03-3.40 μg·mL^-1 with good linearity(r>0.999). The correction factors of Ac-Oxy, Oxy[Glu⁴] and Oxy[-NH₂] were 1.1, while the correction factors of Oxy[+Gly¹⁰] and Oxy[trisulfide] were 1.2 and 0.9, respectively. The correction factors of Oxy[cis-dimer] and Oxy[trans-dimer] were both 1.3. The seven impurities were determined in 3 batches of samples by principal component self-control with correction factor. The contents of impurity Ac-Oxy were 0.96%, 0.93% and 1.01%, respectively. The contents of impurity Oxy[Glu⁴] were 0.07%, 0.06% and 0.08%, respectively. The contents of impurity Oxy[+Gly¹⁰] were 0.07%, 0.04% and 0.04%, respectively. The contents of impurity Oxy[-NH₂] were 0.09%, 0.05% and 0.07%, respectively. The contents of impurity Oxy[trans-dimer] were 0.27%, 0.18% and 0.22%, respectively. The maximum single impurity contents were 0.18%-0.19%, while the total impurity contents were 1.88%-2.06%. Compared the results measured by principal component self-control with correct factor method and the impurity reference method, there was no significant difference between two methods (p>0.05).

The method is proved to be simple, repeatable and accurate for the content determination of related substances in oxytocin for injection.