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 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 establish a quality assessment method for lichens of Thamnolia subuliformis (Ehrh.) W. Culb. based on HPLC fingerprint, and qualitative analysis of the chemical constituents by ultra-high performance liquid chromatography coupled with quadrupole/electrostatic field orbital trap high resolution mass spectrometry (UHPLC-Q Exactive Focus MS/MS).

Agilent TC-C₁₈(250 mm×4.6 mm, 5 μm) chromatographic column was used, the mobile phase was methol -0.1% phosphoric acid with gradient elution at the flow rate of 1.0 mL·min^-1, the column temperature was 30 ℃, the detection wavelength was 254 nm, and the injection volume was 10 μL. HPLC fingerprints of lichens of Thamnolia subuliformis was established. The software of similarity calculation for traditional Chinese medicines fingerprints (version 2012) was used to establish the fingerprinting of lichens of Thamnolia subuliformis. The chemical constituents were analyzed by UHPLC-Q Exactive Focus MS/MS, The chromatographic separation was performed on a Waters Acquity UPLC BEH C₁₈ (50 mm×2.1 mm, 1.7 μm) column with acetonitrile(A) -0.1% formic acid aqueous solution(B) as mobile phase for gradient elution, the flow rate was 0.3 mL·min^-1 and the column temperature was 30 ℃.Mass spectrometry was performed using an electrospray ionization and data was collected in negative ion modes in the range of m/z 80-1 200.

Eight common peaks were identified from fingerprints of 10 batches of samples. The RSD values of relative retention time of 8 common peaks of chromotograms of samples were all below 0.5% and the similarities were above 0.9. 4 of the identified peaks were further confirmed by UHPLC-Q Exactive Focus MS/MS as squamatic acid, baeomycesic acid, thamnoliadepsides B, barbatinic acid.

The established method of fingerprint is stable, reliable, and specific, which can be used for quality evaluation of lichens of Thamnolia subuliformis.

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 develop an HPLC method for the separation of perindopril tert-butylamine and its epimer [(±)-1”-epi-perindopril tert-butylamine] and the determination of the epimer.

Perindopril and (±)-1”-epi-perindopril were separated on an Agilent Poroshell CS-C₁₈ column (100 mm×3.0 mm, 2.7 μm) maintained at 50 ℃ with the mobile phase containing a mixture of 0.15% sodium heptanesulfonate solution (adjusted to pH 2.0 with phosphoric acid) and acetonitrile-pentanol(217∶3)(82∶18, V/V) at 0.8 mL·min^-1, and the detection wavelength was set at 215 nm. The injection volume was 2 μL.

(±)-1”-epi-perindopril and perindopril were separated successfully in 25 min with peak to valley ratio more than 3.0 or a resolution factor of 1.7. Good linear relationships were established between the peak response and the concentration in the range of 2-2 000 μg·mL^-1 for the epimer and perindopril tert-butylamine(r>0.999). The quantitative limits(S/N= 10) were both about 1.0 μg·mL^-1, and the detection limits(S/N=3) were both 0.3 μg·mL^-1. The spiked recovery of the epimerer was 97.2% (RSD=1.8%, n=9). The content of (±)-1”-epi-perindopril tert-butylamine in 10 batches of samples ranged from 0.025% to 0.078%.

The proposed method enhances the resolution efficiency, shows high accuracy, repeatability and stability. It can be effectively employed for the quality control of perindopril tert-butylamine．

To establish a screening method for the detection of adulteration of Draconis Sanguis in Zhenghonghua oil.

To establish a thin layer chromatography (TLC) method with 7, 4’-dihydroxyflavone as an index for fast screen of the adulteration in Zhenghonghua oil. The suspected samples were screened by TLC and the quantification was performed by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The chromatographic column was SuperLu C₁₈ (2)(100 mm×2.1 mm, 1.8 μm), the mobile phase was acetonitrile-water (25∶75) at a flow rate of 0.3 mL·min^-1 and the column temperature was 25 ℃. The detection ion pairs for 7, 4’-dihydroxyflavone were m/z 253.1→117.1 (quantitative) and m/z 253.1→91.0 (qualitative).

TLC method was specific and durable. The linear range of 7, 4’-dihydroxyflavone was 0.470-37.58 ng·mL^-1 (r=0.999 7) by UPLC-MS/MS. The average recovery rate(n=6) was 96.1% and RSD was 3.2%. Thirty batches of Zhenghonghua oil were screened by TLC, and 24 batches were suspected to have 7, 4’-dihydroxyflavone. The suspected samples were verified by UPLC-MS/MS, 24 batches of Zhenghonghua oil exceeded the proposed limit, and the verification results were consistent with TLC method.

This method is rapid, accurate and can be used for the screening and analysis of adulterated Zhenghonghua oil.

To establish a method for simultaneous determination of aluminum trioxide, silicon dioxide and magnesium oxide in montmorillonite and its preparations.

The method was optimized and established for simultaneous determination of aluminum (²⁷Al), silicon (²⁸Si), and magnesium (²⁴Mg) in montmorillonite using rhadium (¹⁰³Rh) as the internal standard, a dilute nitric acid hydrofluoric acid saturated boric acid solution with secondary microwave digestion and inductively coupled plasma mass spectrometry(ICP-MS). The method was compared and analyzed with the EDTA volumetric method for the determination of aluminum trioxide, the muffle ignition combustion method for the determination of silicon dioxide and the AAS method for the determination of magnesium oxide after melting samples of lithium metaborate in the USP.

The linear of aluminum (Al) and silicon (Si) were in the range of 25-300 ng·mL^-1. The linear of magnesium (Mg) was in the range of 20-240 ng·mL^-1, r≥0.999. The detection limits of Al, Si, and Mg were 0.49, 1.30, 0.71 ng·mL^-1. The average recoveries were 99.7%-100.6%, 99.9%-101.2%, 99.8%-101.2%, and the average repeatability RSDs (n=6) were 0.3%-1.3%, 0.4%-1.2%, and 0.4%-1.0%. The contents of aluminum trioxide, silicon dioxide and magnesium oxide in montmorillonite, montmorillonite powder, and montmorillonite dispersion tablets were determined using the newly established method and the original standard method, and the measurement results were basically consistent. Eight common peaks were identified from fingerprints of 10 batches of samples. The RSD values of relative retention time of 8 common peaks of chromotograms of samples were all below 0.5% and the similarities were above 0.9. The contents of aluminum trioxide in montmorillonite were determined using the newly established method and the original standard method, which were 2.3% to 2.4%, 2.0% to 2.2%, 58.7% to 61.5%, 55.3% to 59.4%, and 15.9% to 20.6%, 14.6% to 19.4%, respectively. The results of the measurements were basically consistent for the determination of aluminum trioxide, silicon dioxide, and magnesium oxide content in montmorillonite raw materials and formulations.

The newly established content determination method uses a combination of secondary microwave digestion and ICP-MS technology to determine the content of aluminum oxide, silicon dioxide, and magnesium oxide in montmorillonite samples. It is simple, fast, sensitive, and accurate, and can be used.

To develop an HPLC method for the determination of related substances and assay of levamisole hydrochloride tablets.

The chromatographic separation of related substances was performed on a Thermo Hypsil C₁₈ column(100 mm×4.6 mm, 3 μm). A gradient elution was applied with a mobile phase composed of 0.75% monobasic ammonium phosphate solution(adjusted at pH 6.5 with diiopropylamine) and acetonitrile. The assay analytical column was packed with IntertSustain C₁₈ column(150 mm×4.6 mm, 5 μm). The mobile phase was 70 volumes of 0.75% monobasic ammonium phosphate solution(adjusted at pH 7.0 with diiopropylamine) with 30 volumes of acetonitrile. The wavelength detection was set at 215 nm, the injection volumn was 10 μL, and the flow rate was 1.0 mL·min^-1.

Levamisole and its impurities were separated well by related substances HPLC method above. Impurities A, B, C, D, E showed the good linearities in the concentration ranges of 10.87-25.37 μg·mL^-1, 11.62-27.10 μg·mL^-1, 12.38-28.90 μg·mL^-1, 30.89-72.07 μg·mL^-1, 12.41-28.95 μg·mL^-1 (r>0.999). The average correction factors of impurities A, B, C, D, E determined by three columns and liquid chromatographies were 1.6, 1.4, 2.6, 1.2, 2.4, respectively, the recovery rates(n=9) were 98.1%, 99.0%, 98.6%, 100.1%, 99.9% and the RSDs were 0.63%, 0.79%, 0.92%, 0.96%, 0.33%. The separation of levamisole and impurity C was good by assay HPLC method. Levamisole showed the good linearity in the concentration range of 0.10 14-0.304 3 mg·mL^-1 (r=0.999 9). The average recovery rate(n=9) of levamisole was 100.2%, RSD=0.52%. Ten batches levamisole hydrochloride tablets from five domestic pharmaceutical enterprises were determined by the above related substances and assay HPLC method, the total impurities mass were 0.05%-0.62%, and the assays were 99.5%-104.3%.

The established method is high selection and accurate. It is suitable applied for determination of related substances and assay of levamiole hydrochloride tablets.

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.