To establish a quality evaluation method for Chaihu Yujinxiang granules based on fingerprint, multi-component content determination, and chemical pattern recognition, providing important basis for its quality control.

The “Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Edition)” was used to establish HPLC fingerprints of 10 batches of Chaihu Yujinxiang granules, identify common peaks, and perform similarity evaluation. HPLC method for determining the content of four active ingredients, namely liquiritin apioside, liquiritin, liquiritigenin, and isoliquiritigenin was established. Cluster analysis and orthogonal partial least squares discriminant analysis were conducted on Chaihu Yujinxiang granules using SPSS 26.0 and SIMCA 14.1 software. Differential components affecting the quality of Chaihu Yujinxiang granules were screened based on the criterion of variable importance projection (VIP) value>1.0.

The established fingerprint and multi-component content determination method achieved satisfactory results after methodological investigation. The similarity of the fingerprint of 10 batches of Chaihu Yujinxiang granules ranges from 0.953 to 0.977, with a total of 11 common peaks, 4 of which were identified. The average contents of active ingredients such as liquiritin apioside, liquiritin, liquiritigenin, and isoliquiritigenin in 10 batches of Chaihu Yujinxiang granules were 2.71 mg·g^-1, 10.17 mg·g^-1, 2.47 mg·g^-1, and 1.86 mg·g^–1, respectively. The results of cluster analysis and principal component analysis indicates that 10 batches of Chaihu Yujinxiang granules could be clustered into two categories, with S3, S6, and S8 in one category and the rest in another category. The VIP values of peaks 2, 9, 7, and 3 were above 1.0.

The established fingerprint and content determination method are stable and reliable. Combined with chemical pattern recognition technology, they can be used to evaluate the overall quality of Chaihu Yujinxiang granules. Peaks 2, 9, 7, and 3 are differential components that affect the quality of Chaihu Yujinxiang granules.

To establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for determining the concentration of cyclic guanosine monophosphate (cGMP) in lysates of human colon adenocarcinoma lung metastasis cells (T84 cells) after co-incubation with linaclotide.

A Kromasil 100-5-C₁₈ column (150 mm×2.1 mm, 5 μm) was used with a mobile phase consisting of 0.1% formic acid in water and 0.1% formic acid in methanol, employing a gradient elution at a column temperature of 50 ℃. The detection was performed using an electrospray ionization (ESI^-) source and multiple reaction monitoring (MRM) mode, with the monitored ion transitions for cGMP and the internal standard 8-Br-cGMP being m/z 344.20 → 150.00 and m/z 423.90 → 230.00, respectively.

The linear ranges for cGMP were 1 to 500 ng·mL^-1 (r≥ 0.999). The method demonstrated precision, accuracy, matrix effects, and extraction recovery rates that met analytical requirements. The method was successfully applied to accurately detect cGMP levels in cells after administration of two types of linaclotide capsule formulations. A significant concentration-dependent change in cGMP levels was observed after co-incubation of linaclotide with T84 cells for 30 min, with EC₅₀ values of 167.6 and 112.1 nmol·L^-1 for the reference and test formulations, respectively.

The method established in this study demonstrates excellent selectivity and accuracy, effectively quantifying cGMP levels in lysates of human colon adenocarcinoma lung metastasis cells, providing a reliable analytical tool for related pharmacological research.

To establish a method combining size exclusion chromatography (SEC) with an optimized Infogest in vitro static digestion model for determining the relative molecular mass distribution of short-peptide-based enteral nutrition before and after digestion. To evaluate and assess the quality of three different products, to explore product quality characteristics and digestive stability, and to provide guidance for quality control and clinical individualized application of short-peptide-based enteral nutrition.

The SEC method was performed using two ECOSIL SEC G 2000 columns (300 mm×7.8 mm, 5 μm) in series, with acetonitrile-water-trifluoroacetic acid(15：8 5：0 .1) as the mobile phase, a flow rate of 0.7 mL·min^-1, the column temperature 30 ℃, the detection wavelength 215 nm, and the sample volume 20 μL. The in vitro simulated digestion process referred to and optimized the Infogest in vitro static digestion model, divided into simulated gastric digestion, simulated intestinal digestion, and simulated gastro-intestinal total digestion stages. The established SEC method was used to compare changes in the relative molecular mass distribution before and after digestion to evaluate in vitro digestive stability.

Within the range of relative molecular mass from 165.19 to 12 327, the logarithms of the relative molecular mass of the 8 reference standards showed good linear relationships with retention time. Specificity tests indicated that blank solvent and the simulated gastric-intestinal digestion solution had minimal impact on the measurement of relative molecular mass distribution. The RSDs for precision, repeatability, and 24 h stability tests were all less than 1.0%. The relative molecular mass of various short peptide enteral nutrition products was mainly concentrated between 150 and 1 000, but there were significant differences in the distribution ratios of relative molecular mass among different products. The hydrolysis of peptide components primarily occurred during the production stage, and after in vitro simulated digestion, their relative molecular mass distribution characteristics remained largely unchanged, with the degree of change mainly related to the original hydrolysis level of the product.

The SEC method combined with the optimized Infogest model is suitable for studying the molecular mass distribution and digestive stability of short-peptide-based enteral nutrition. The method is straightforward and rapid, facilitating a comprehensive assessment of the products quality characteristics and nutritional value.

To establish an LC-MS/MS method for determining the concentrations of imipenem and cilastatin in human plasma, for monitoring clinical therapeutic drug concentrations, and to investigate the effects of adding stabilizers during the sample pretreatment on mass spectrometry signal intensity.

After protein precipitation, the sample was subjected to gradient elution using an Agilent TC-C₁₈ (2) (150 mm×4.6 mm, 5 µm) column with a mobile phase system of 0.15% formic acid in water and methanol. The electrospray ionization (ESI) mass spectrometer was operated in positive ion mode using multiple reaction monitoring (MRM): m/z 300.1 → 141.9 (imipenem),m/z 359.7 → 97.0 (cilastatin) and m/z 384.1 → 141.1 (meropenem, internal standard). The samples containing and without 3-(N-morpholino) propane sulfonic acid (MOPS) as stabilizers were pretreated and continuously analyzed to compare the changes in mass spectrometry signal intensity.

Both imipenem and cilastatin showed good linearities in the concentration ranges of 0.1-100.0 μg·mL^-1 (r>0.99). The intra-day and inter-day accuracy ranges from 95.3% to 108.5%, the precision (RSDs) were less than 9.3%, the extraction recovery rate ranges from 77.4% to 84.3%, and the matrix effect ranges from 97.1% to 111.2%. Imipenem in plasma samples was stable at room temperature for 3 h, at 4 ℃ for 6 h, and at -80 ℃ for 12 d, while it was significantly degraded at -20 ℃ for 12 d. Cilastatin was stable under a variety of conditions. The method was robust to changing conditions of column temperature ±5 ℃, flow rate ±0.1 mL·min^-1, formic acid concentration in the aqueous phase ±0.025%, and ion source temperature ±50 ℃. The samples containing stabilizers exhibited significant ion inhibition on mass spectrometry after continuous injection, while samples without stabilizers had no significant effect on the signal intensity of mass spectrometry.

The method is simple and accurate and can be used for clinical drug monitoring of imipenem and cilastatin. Nonvolatile salt stabilizers such as MOPS can reduce mass spectrometry sensitivity, and the absence of such stabilizers is more suitable for long-term analysis by LC-MS/MS.

To systematically analyze the pharmacokinetic parameters, oral bioavailability and in vivo metabolites of trilobatin in Sprague-Dawley (SD) rats using liquid chromatography - triple quadrupole mass spectrometry (LC-MS/MS).

The chromatographic conditions were performed on an ACQUITY UPLC BEH C₁₈ column (50 mm×2.1 mm, 1.7 µm) with 0.1% formic acid (mobile phase A) and acetonitrile (mobile phase B) as the mobile phase. A gradient elution program was carried out with an accompanying flow rate of 0.3 mL·min^-1, a column temperature of 40 ℃, and an injection volume of 2 μL. The mass spectrometry conditions comprised an electrospray ion source in conjunction with negative ionization mode, with an ionogenic temperature of 150 ℃, a capillary voltage of -3.0 kV, and a desolvation-gas flow temperature of 500 ℃. The desolvation-gas flow rate was set at 750 L·h^-1, and the conical pore gas volumetric flow rate was fixed at 150 L·h^-1. The analysis was conducted in multiple reaction monitoring mode. Trilobatin was given to rats via gavage and intravenous injection, respectively. Plasma, urine and fecal samples were collected, and the drug concentration was determined after methanol precipitation of proteins. Pharmacokinetic parameters and metabolites were analyzed by pharmacokinetic software and metabolite analysis and identification software.

Following the administration of trilobatin to SD rats at a dose of 100 mg·kg^-1 via gavage and intravenous injection, respectively. The area under the curve (AUC_0-t) was found to be (423.98 ± 295.42) ng·h·mL^-1 and (90 894.75 ± 25 472.44) ng·h·mL^-1, respectively. The oral bioavailability was determined to be 0.46%; C_max was (203.83±25.88) ng·mL^-1 and (181 814.90±113 461.60) ng·mL^-1, respectively. The oral half-life was 1.65 h, while the intravenous half-life was 3.82 h. Trilobatin was metabolized to phloretin in the intestine and underwent further biotransformation in vivo through deglycosylation, methylation, deoxygenation and hydrolysis.

The pilot study represents a preliminary investigation into the in vivo pharmacokinetics and metabolism of trilobatin in rats, providing a foundation for further pharmacodynamics research and subsequent formulation development.

To establish a principal component external standard method with HPLC and calibration factors for the determination of 11 impurities in flurbiprofen axetil injection, and to explore its detection results and limit values.

The Thermo BDS Hypersil C₁₈ (250×4.6 mm, 5 μm) was selected for gradient elution, with water-0.15% acetic acid and acetonitrile-0.15% acetic acid as the mobile phase at a flow rate of 1.0 mL·min^-1. The column temperature was 40 ℃, the detection wavelength was 254 nm and the injection volume was 10 μL.

Flurbiprofen axetil and 11 impurities were well separated by the method. Good linearity was obtained with correlation coefficients of 1.000 for the 3-fluoro-4-phenylphenol (4-OHB), 1-acetoxyethyl-2-(2-fluoro-4-biphenylyl)-2-hydroxypropionate (2-OHP), 4-acetyl-2-fluorobiphenyl (4-ACB), flurbiprofen ethyl ester, allyl -(2-fluoro-4-biphenyl) propionate (ALE), ChP impurityⅠ , impurity B, impurity C and impurity E in the range of 0.10-20 μg·mL^-1. The average recovery rates was from 96.6% to 103.7% and the relative standard deviations(RSDs) were lower than 1.4%. The correction factors of flurbiprofen axetil related substances 4-OHB, 2-OHP,4-ACB, flurbiprofen, flurbiprofen ethyl ester, ALE, ChP impurity Ⅰ, impurity B, impurity C and impurity E were 0.55, 1.05, 1.01, 0.76, 0.95, 0.86, 0.55, 0.93, 0.76 and 0.81, respectively. Notablely, desfluoro fiurbiprofen axetil of detected was around the prescribed limit 0.1%.

The method above is rapid, simple, accurate, and reliable, and can be applied for the determination and quality control of related substances in flurbiprofen axetil injection.

To identify the authenticity of four batches of “Ophiopogonis Radix” in the market by means of multiple means, and to explore the reasons for exceeding the limit of its phloem bundles, so as to provide evidence for its inspection and detection.

Based on the relevant provisions of Ophiopogon japonicus (L. f.) Ker-Gawl., a variety in the 2020 edition of Chinese Pharmacopoeia, comparing the common confused products of Ophiopogon japonicus (L. f.) Ker-Gawl., Combined with traditional identification methods (character identification, microscopic identification) and modern analysis techniques (molecular biology ITS 2 sequence), the data of genuine products with mixed products and substandard phloem bundle samples were campared and analyzed.

The differences between Liriope spicata (Thunb.) Lour. and Ophiopogon japonicus (L. f.) Ker-Gawl. were the surface color, the depth of vertical wrinkles and the thickness of the middle column, and the differences in microscopic cross-sections lay in the number of phloem bundles and whether the inner cortex cells were uniformly thickened. The four batches of “Ophiopogonis Radix” in the market all complied with the relevant regulations under Ophiopogon japonicus (L. f.) Ker-Gawl.. The number of unqualified phloem bundled into microscopic cross-sections. was more than 40%, but the inner cortex cells in the samples showed a comprehensive thickening phenomenon. The results of molecular biology study showed that the four batches of “Ophiopogonis Radix” on the market and the Ophiopogon japonicus (L. f.) Ker-Gawl. were clustered into one, the Liriope spicata (Thunb.) Lour.var. prolifera Y. T. Ma and the Liriope muscari (Decne.) Baily were clustered into one, and showed obvious bar code spacing. The number of phloem bundles was obvious positively correlated with the diameter of wood core by correlation analysis of the number of phloem bundles and the quantitative indexes related to traits.

These four batches of “Ophiopogonis Radix” on the market are the original of genuine products. Combined with the experimental research, it is speculated that the cause of the unqualified number of phloem bundles may be related to the growth years, and the middle column thickness can roughly predict whether it can meet the requirements of pharmacopoeia. To determine whether Ophiopogonis Radix is an accurate medicinal material based on the original, we should not only make a conclusion based on a certain feature of a certain identification method, but also should combine multiple methods to determine accurately.