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 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 investigate the metabolites of Periploca forrestii in plasma, urine, and feces of normal and adjuvant arthritis（AA） rats, and explore the effect of rheumatoid arthritis（RA） on the metabolism of active components of P. forrestii.

AA rat model was made by means of Freund’s complete adjuvant. Plasma, urine and feces of normal and AA rats were analyzed by UPLC-Q-TOF-MS^E method using ACQUITY UPLC BEH C₁₈ (100 mm×2.1 mm, 1.7 μm) column with 0.01% formic acid water -0.01% formic acid acetonitrile as mobile phase gradient elution at a flow rate of 0.25 mL·min^-1, and electrospray ion source under negative ion mode.

Two and three prototype components, 32 and 35 metabolites were detected in normal rats and AA model rats. The metabolic pathways mainly include monocaffeoylquinic acid reduction, methylation, ring-opening cleavage, glucuronidation, dicaffeoylquinic acid reduction, methylation, acetylation, isomerization, sulfation glucuronidation, etc.

The metabolites in plasma and urine of AA model rats are more diverse than those in normal rats, and the disease state of RA may affect the metabolic pathway of effective components of P. forrestii in the body.

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 investigate and use the different constituents in Bofonis Corium distinguishing from Bufonis Venenum based on ultra-performance liquid chromatography-quadrupole-electrostatic field orbitrap high-resolution mass spectrometry (UPLC-Q-Exactive Orbitrap MS) coupled with chemometrics.

UPLC analysis was performed on a Waters Atlantis T3 (150 mm×2.1 mm, 3 μm) column with gradient elution of acetonitrile and 0.1% formic acid at a flow rate of 0.3 mL·min^-1. The column temperature was set at 30 ℃. UPLC-Q-Exactive Orbitrap MS was chosen for data collection in both positive and negative ion modes. The potential differential ions were obtained by principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA). The Xcalibur 3.0 data processing system was utilized to extract the secondary fragmentation details of the differential ions. Subsequently, the specificity of differential ions was confirmed through the multiple reaction monitoring (MRM) mode of AB SCIEX 6500⁺ triple quadrupole mass spectrometry. Identification of adulterated Bofonis Corium in Bufonis Venenum was performed by applying the ion pairs with good specificity.

After the research and verification, a total of five distinction pairs (m/z: 377.5→243.2, 172.1；330.4→170.9, 127.0；313.4→201.2, 171.0；452.4→280.2, 298.2 and 614.7→332.4, 281.3) were discovered, and a method for the examination of Bofonis Corium in Bufonis Venenum was established and methodologically investigated by applying the information of the different ions. The constructed method had reasonable specificity and durability. Each ion pair showed an excellent linear relationship within a specific range, with correlation coefficients above 0.996 7. The RSDs of the precision test were in the range of 1.3%-4.1%. The RSDs of the repeatability test were in the range of 1.3%-3.2%. Five batches of market samples of Bufonis Venenum were determined according to the above process, and two batches of samples showed the presence of Bofonis Corium components but did not exceed the proposed 5% adulteration limit.

In this study, a rapid method for detecting discrepancy information based on UPLC-Q-Exactive Orbitrap MS combined with chemometrics is presented, and the discrepant information is applied to establish an identification method for adulterated Bofonis Corium in Bufonis Venenum.

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 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 establish an UPLC-MS/MS method for simultaneous determination of 11 components(caffeic acid, afzelin, gallic acid, neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, salidroside, eleutheroside B, ginsenoside Re, ginsenoside Rg₁, ginsenoside Rd) in Hongwushen capsules.

A Shim-pack GIST C₁₈ chromatographic column(100 mm×2.1 mm, 2 μm) was used. The mobile phase was composed of acetonitrile -0.1% acetic acid -1 mmol·L^-1 ammonium acetate aqueous solution and the gradient elution was applied. The flow rate was 0.3 mL·min^-1, the column temperature was 30 ℃, and the injection volume was 1 μL. Electrospray ionization (ESI) source and multiple reaction monitoring (MRM) mode in both positive and negative were used for ion detections.

The linear relationship of 11 components was good in the concentration range, and the linear correlation coefficients were all above 0.999 0. The RSD values of precision were less than 3%. The repeatability and stability were good, and the RSD values were less than 5%. The average recoveries were in the range of 97.1%-101.5% with RSDs≤3.7%. The contents of caffeic acid, afzelin, gallic acid, neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, salidroside, eleutheroside B, ginsenoside Re, ginsenoside Rg₁ and ginsenoside Rd in 10 batches of Hongwushen capsules were 0.010-0.013 mg·g^-1, 0.000 7-0.001 4 mg·g^-1, 0.035-0.038 mg·g^-1, 0.312-0.315 mg·g^-1, 0.413-0.417 mg·g^-1, 0.411-0.416 mg·g^-1, 4.355-4.358 mg·g^-1, 0.030-0.032 mg·g^-1, 0.993-0.999 mg·g^-1, 1.120-1.124 mg·g^-1 and 2.536-2.538 mg·g^-1, respectively.

The method is accurate, sensitive, stable and reproducible, and can be used for the quality control of Hongwushen capsules.

To establish a method for the rapid identification of the authenticity of Gastrodiae Rhizoma herbs based on enzymatic recombinase amplification (ERA) technique.

Following the principle of ERA primer design, Oligo 7.0 software was applied to screen and optimize the ERA-specific primers of Tianma based on the ITS2 genome sequences of Gastrodia elata and its common artifacts. Primer Premier 5.0 software was applied to design the specific PCR primers for the identification of Gastrodia elata, and through the optimization of ERA and PCR reaction system, the optimal reaction time for ERA was finally determined to be 17 min, the optimal reaction temperature was 40 ℃, the optimal annealing temperature for PCR was 57 ℃, and the cycle was 32 times, and the established method was verified for sensitivity and specificity, and the samples of asparagus available in the traditional Chinese medicine The sensitivity and specificity of the established method were verified, and the commercially available asparagus samples in the market were selected for testing.

The designed primers for the specific identification of ERA in Gastrodia elata did not cross-react with its common forgeries, and the specificity was good, a repeatability results showed that the three repeatability tests were consistent, with no false positives or false negatives；the sensitivity of this method for Gastrodia elata genomic DNA was 1 pg·μL^-1, which was higher than that of conventional PCR；the ERA technique can be used for the rapid identification of commercially available samples of Gastrodia elata and the results of the assay are the same as those of the PCR method.

The established detection method is simple, rapid, with high specificity and sensitivity, and provides a new means for the authentication of the Gastrodiae Rhizoma.