Objective: To establish a dual derivatization method combined with tandem mass spectrometry for the simultaneous determination of 18 different steroid hormones in human serum. Methods: Serum samples were treated with hydroxylamine and 1, 2-dimethylimidazole-5-sulfonyl chloride for derivatization, and the resulting compounds were analyzed by liquid chromatography-tandem mass spectrometry in positive ion selected reaction monitoring (SRM) mode. The Kinetex^®C₈ column (100 mm×2.1 mm, 2.6 μm) was used for the separation. Mobile phase A (0.1% acetic acid in water) and mobile phase B (0.1% acetic acid in methanol) with gradient elution（0-0.5 min, 35% B; 0.5-5 min, 35% B→100% B; 5.0-5.1 min, 100% B→35% B; 5.1-7 min, 35% B）at the flow rate of 0.4 mL·min^-1 were applied. Column temperature was set at 40 ℃. Injection volume was 20 μL and collection time was 6 min. Results: The linearity correlation coefficients for all 18 steroid hormones were greater than 0.99, the recovery rates ranged from 85% to 115%, and the precision RSD was less than 15%. This method was successfully applied to the analysis of serum samples from 156 healthy subjects (75 males and 61 females), and reference intervals were established. Conclusion: This method can be used to simultaneously determine 18 types of steroid hormones, such as pregnenolone, 17-hydroxypregnenolone, progesterone, 17-hydroxyprogesterone, corticosterone, cortisol, 11-deoxycorticosterone, 21-deoxycorticosterone, aldosterone, testosterone, androstanedione, dehydroepiandrosterone, sulfated dehydroepiandrosterone, adrenocorticotropin, 18-hydroxicorticotropin, estrone, estradiol, and estriol using non-solid-phase extraction.

Objective: To develop a method to assess bioactivity of trasfer factor (TF) based on their protective effcets on Jurkat E6-1 cells. Methods: Proliferative effects of different concentrations of TF on 6-mercaptopurine（6-MP） treated Jurkat E6-1 cells detected by CCK-8 assay and the precision was validated. Results: TF exhibited protective activity to change the inhibitory concentration (IC₅₀) value of 6-MP from（0.46±0.10）μg·mL^-1 to （1.11±0.30）μg·mL^-1 when treated with 20 μg·mL^-1 TF. When cells were treated with 6-MP, there is a good linear relationship between the concentration of TF and cell survival rate, r>0.95. At each concentration level of the dose-response curve, the RSD was less than 20%. The median effective concentration (EC₅₀) of TF was （28.49±9.60）μg·mL^-1, and the confidence limit was less than 20%. Conclusions: TF significantly improved Jurkat E6-1 cell survival rate in dose dependent manner when challenged with 6-MP. It may be suitable for evaluate bioactivity of TF.

Objective: To establish an authentication method for Eupolyphaga sinensis based on loop-mediated isothermal amplification (LAMP). Methods: Two pairs of primers （outer primer DB-F3, DB-B3 and inner primer DB-FIP, DB-BIP） were designed according to Eupolyphaga sinensis COⅠspecific loci. The reaction system containing template, primers, Bst DNA polymerase and methyl red-phenol red indicator amplified at 63 ℃ for 90 min. The LAMP results were determined by visual observation and compared with DNA barcoding and polymerase chain reaction (PCR) methods to investigate the specificity and sensitivity. Results: All 11 batches of the authentic Eupolyphaga sinensis sample tubes colour changed from purple red to orange yellow, while sample tubes of 1 batch of Steleophaga plancyi, 9 batches Opisthoplatia orientalis and 1 batch of the adulterant, Cybister tripunctatus orientalis, remained purple red after the reaction. The LAMP results were consistent with those of DNA barcoding and PCR. The detection limit of LAMP was 0.984 ng·mL^-1. Conclusion: The established LAMP method is accurate, sensitive and easy to operate, low equipment required, and can be applied to the rapid screening of the authentication of Eupolyphaga sinensis.

Objective: To study the difference of chemical composition between ancient and modern clinical prescriptions of Baihe Dihuang decoction by multi-component analysis. Methods: The polysaccharide extract of Baihe Dihuang decoction was determined by anthrone-sulfuric acid-ultraviolet spectrophotometer under 580 nm ultraviolet light. Six monosaccharides and oligosaccharides in the ancient and modern Baihe Dihuang decoction were determined by UPLC-CAD. An Agilent InfinityLab Poroshell 120 HILIC-Z (3.0×100 mm, 2.7 μm) column was used. The mobile phase was consisted of A 0.2% ammonia in acetonitrile and 0.2% ammonia in water with gradient elution. The flow rate was 0.8 mL·min^-1, the column temperature 35 ℃ and the CAD detector nebulizer temperature was 70 ℃. Data acquisition frequency was 10 Hz and the filter was 1.05 s. The injection volumn was 2 μL. The non-polar components were qualitatively and quantitatively analyzed by high performance liquid-quadrupole tandem time-of-flight mass spectrometry (UPLC-Q TOF MS/MS) and high performance liquid-phase tandem triple quadrupole mass spectrometry (UPLC-QQQ MS), respectively. The differential components of ancient and modern Baihe Dihuang Decoction were analyzed and 9 components were determined. Results: The linear relationship of 6 mono-oligosaccharides/polysaccharides and 9 differential components was good (r>0.999). The average recoveries ranged from 97.7% to 104.9% with RSDs≤3.1%. Precision, repeatability and stability met the requirements. The results showed that the conents of all the analytes in ancient and modern Baihe Dihuang decoction were significantly different (P<0.05). Conclusion: The methods used in this experiment are accurate, sensitive, stable and reproducible, which can provide reference for the quality control and pharmacodynamic material basic research of Baihe Dihuang Decoction.

Objective: To establish the HPLC fingerprint of Suhexiang pills and determine the contents of gallic acid, piperine, costunolide, eugenol and cinnamic acid simultaneously, and to provide reference for the quality control method of Suhexiang pills. Methods: The separation was performed on a Swell Chromplus C₁₈ column (250 mm×4.6 mm, 5 μm) with methanol (A)-0.05% phosphoric acid solution (B) as the mobile phase at a flow rate of 1.0 mL·min^-1. The detection wavelength was 220 nm and the column temperature was 30 ℃. Fifteen batches of Suhexiang pills were analyzed. The “Traditional Chinese Medicine Fingerprint Similarity Evalution System” was used to establish the reference fingerprint and the similarity analysis was evaluated in combination with cluster analysis and principal component analysis. Results: The fingerprint of Suhexiang pills was established，and the similarities of 15 batches of samples were above 0.9. Thirteen common peaks were identified，and five components including galliic acid，cinnamic acid，eugenol，piperine and costunolide were identified. The contents of the five components were 2.506-3.652 mg·g^-1，0.666-0.876 mg·g^-1，3.834-5.140 mg·g^-1，0.884-1.306 mg·g^-1 and 19.908-55.704 mg·g^-1，respectively. Using cluster analysis，15 batches of Suhexiang pills were divided into four categories. Three principal components were selected，and the cumulative variance contribution rate was 76.102%，indicating that the principal components could contain most of the information of the original data. Conclusion: The fingerprint and multi-component content determination method of Suhexiang pills are stable and reliable，which can provide reference for the quality control and clinical application of Suhexiang pills.

Objective: To establish an HPLC method for the simultaneous determination of six components (uridine, adenine, adenosine (R, S)-goitrine, guanosine, clemastanin B) in Radix Isatidis, and to investigate the linear calibration with two reference substances (LCTRS) method for the qualitative analysis of multiple components in Radix Isatidis. Methods: HPLC method was used, with methanol as mobile phase A and water as mobile phase B. Gradient elution (0-3 min, 3% A; 3-18 min, 3% A→14% A; 18-25 min, 14% A→26% A; 25-34 min, 26% A; 34-40 min, 26% A→46% A; 40-60 min, 46% A→90% A) was performed at a flow rate of 0.8 mL·min^-1. The column temperature was 30 ℃, the detection wavelengths were 254 nm (0-32 min) and 230 nm (32-60 min). The injection volume was 10 μL. The actual retention time of 6 components in Radix Isatidis was determined on 13 C₁₈ chromatographic columns of different brands and models. Guanosine and clemastanin B were used as double reference compounds, and LCTRS method was used to locate the chromatographic peak of each component. Three unknown chromatographic columns were used for method validation. Using guanosine as a reference substance, the relative retention time method was used to predict the retention time of the other 5 components. The predictive accuracy and column coincidence of these two methods were compared. Results: The LCTRS method could effectively predict and qualitatively analyze the retention time of six indicator components. Compared with the relative retention time method, the LCTRS method had higher accuracy in predicting results and better column universality. Conclusion: The LCTRS method for simultaneous determination of multiple components in Radix Isatidis is feasible and accurate, with simple operation and good durability, and has promotional value.

Objective: To establish a multi-component content determination and characteristic chromatogram method for Jieyu Anshen granules, combined with chemometrics, to comprehensively evaluate the quality of Jieyu Anshen granules. Methods: The separation was performed on a CAPCELL PAK MG Ⅱ C₁₈(250 mm×4.6 mm, 5 μm) column using acetonitrile (A) and 0.1% phosphoric acid aqueous solution (B) as the mobile phases with gradient elution at a flow rate of 1.0 mL·min^-1. The column temperature was 30 ℃ and the detection wavelengths were set at 237 nm and 335 nm. The analysis method was validated. Based on the multi-component content determination method, characteristic chromatogram was established. The samples were analyzed and evaluated through chemometrics to identify the main factors affecting their quality. Results: A multi-component content determination method was established for Jieyu Anshen granules. The five components had good linear relationships within their respective ranges, with recovery rates ranging from 95.6% to 100.0%. And 145 batches of samples were tested, and a total of 54 batches were found to be unqualified, with a total unqualified rate of 37.2%. Establishing a characteristic chromatogram of Anshen granules, 12 characteristic peaks were attributed to 6 medicinal herbs. Through chemometric analysis, it was found that there were certain differences in the quality of samples from 10 manufacturers. The quality of fried gardenia and stir-baked licorice was the key factor. Conclusion: The established multi-component content determination method and characteristic chromatogram method for Jieyu Anshen granules are convenient and reliable, and can be used for comprehensive evaluation of the quality of Jieyu Anshen granules.

Objective: To establish a method for determination of aconite alkaloids in Fengshi Gutong tablets. Methods: Analysis was performed on a WatersXSelect^® CSH C₁₈（250 nm×4.6 mm, 5 μm） column with mobile phase consisting of 0.1% acetic acid aqueous solution, acetonitrile and methanol with gradient elution at a flow rate of 1.0 mL·min^-1. The detection wavelength was set at 245nm and the column temperature was 30 ℃. Results: Twelve aconite alkaloids could be separated well. When the injection amounts of ranaconitine, benzoylmesaconine, benzoylmesaconine, benzoylmesaconine, acoforestinine and beiwutine, mesaconitine, hypaconitine, indaconitine, aconitine, yunaconitine, bulleyaconitine A were 0.022 8-0.136 6 μg, 0.041 5-0.249 0 μg, 0.033 5-0.200 8 μg, 0.033 9-0.203 1 μg, 0.033 1-0.198 6 μg, 0.040 3-0.241 6 μg, 0.030 2-0.181 4 μg, 0.028 6-0.171 6 μg, 0.033 6-0.201 6 μg, 0.030 3-0.181 9 μg, 0.063 4-0.381 5 μg and 0.034 0-0.204 2 μg, respectively, the peak area showed a good linear relationship with the injection amounts. The average recovery rates of 12 aconite alkaloid components ranged from 93.6% to 101.5%, and the RSD ranged from 0.55% to 2.6%. The average contents of ranaconitine, benzoylmesaconine, benzoylmesaconine, benzoylmesaconine, acoforestinine and beiwutine, mesaconitine, hypaconitine, indaconitine and yunaconitine in three batches of Fengshi Gutong tablets (batch No. 211227, 220823 and 230425) were 0.492 μg, 65.78 μg, 7.319 μg, 10.164 μg, 1.068 μg, 5.583 μg, 2.573 μg, 5.865 μg, 2.021 μg, 2.050 μg, respectively. And aconitine and bulleyaconitine A were not detected. Conclusions: The established method is accurate and reliable, and can be used for the determination of 12 aconite alkaloids compounds in Fengshi Gutong tablets to achieve comprehensive quality control of preparations.

Objective: To establish the HPLC fingerprints for Zilian Shengji gel and to determine the contents of gallic acid, tableberberine, coptisine, palmatine, berberine and shikonin. Methods: The establishment of fingerprints was performed on a 25 ℃ thermostatic Agilent C₁₈-WR（250 mm×4.6 mm, 5 μm）column with the mobile phase comprising of acetonitrile -0.15% phosphoric acid water at the flowing rate of 0.8mL·min^-1 in a gradient elution manner, and the detection wavelength were set at 270 nm, 345 nm and 516 nm. Cluster analysis, principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) were adopted in chemical pattern recognition. The six components identified were quantitatively determined. Results: The HPLC fingerprint of Zilian Shengji gel was constructed with twenty-six common chromatographic peaks, eleven batches of samples with the similarities of 0.828-0.997, six chromatographic peaks of gallic acid, tableberberine, coptisine, palmatine, berberine and shikonin were identified by reference substance comparison. Six constituents showed good linear relationships within their own ranges (r≥0.999 5) whose average recoveries were 98.7%-99.3%, with the RSDs of 1.3%-1.8%. The content ranges of the above mentioned six components in eleven batches of samples were 827.74-1 513.50 μg·g^-1, 137.52-296.05 μg·g^-1, 381.83-884.73 μg·g^-1, 2 023.81-4 051.66 μg·g^-1, 524.15-986.13 μg·g^-1 and 47.65-549.46 μg·g^-1. Conclusion: This simple, stable, reliable and reproducible method can be used for the quality control of Zilian Shengji gel.

Objective: To establish an HPLC method for simultaneous determination of loganic acid, gentiopicroside, hesperidin, paeonol, aloe-emodin, rhein, emodin, chrysophanol and physcion in Tongshu Koushuang tablets (Rhei Radix et Rhizoma, Aurantii Fructus Immaturus and Gentianae Macrophyllae Radix, etc.). Methods: The samples were extracted by refluxing with methanol and the separation was performed on a Thermo Acclaim-C₁₈(250 mm×4.6 mm, 5 μm) column at 30 ℃, with mobile phase consisting of 0.15% phosphoric acid solution and acetonitrile with a gradient elution at a flow rate of 1.0 mL·min^-1. The injection volume was 10 μL, and the detection wavelength was set at 236 nm (detecting loganic acid), 280 nm (detecting gentiopicroside, hesperidin and paeonol) and 254 nm (detecting aloe-emodin, rhein, emodin, chrysophanol and physcion). Results: Nine constituents (loganic acid, gentiopicroside, hesperidin, paeonol, aloe-emodin, rhein, emodin, chrysophanol and physcion) were in good linearity within their own ranges (r were 0.999 1-0.999 9), and the average recoveries were 99.6% (RSD=1.9%), 101.1% (RSD=2.0%), 103.0% (RSD=1.5%), 101.6% (RSD=0.51%), 99.8% (RSD=1.4%), 100.9% (RSD=1.4%), 100.8% (RSD=1.8%), 102.7% (RSD=1.2%) and 102.9% (RSD=0.86%), respectively. The content ranges of nine constituents were 1.871-3.517 mg·g^-1, 6.274-12.55 mg·g^-1, 2.140-3.946 mg·g^-1, 1.393-2.018 mg·g^-1, 0.394 8-0.807 2 mg·g^-1, 0.657 5-1.246 mg·g^-1, 0.484 7-0.915 8 mg·g^-1, 0.839 8-1.429 mg·g^-1 and 0.327 3-0.547 9 mg·g^-1, respectively. Conclusion: This simple and accurate method can be used for the quality control and evaluation of Tongshu Koushuang tablets.