To establish the research method for the sealing integrity of the injection bottle packaging system for human albumin injection.

For the injection bottle packaging system of human albumin injection, positive control samples with pore diameter of 1 μm were prepared using a glass micropipette, positive control samples with pore diameter of 2, 5 and 10 μm were prepared using laser drilling. Two deterministic sealing integrity testing methods, vacuum decay method and high-voltage leak detection method were developed and tested.

The vacuum decay method could not effectively detect the leakage of the packaging system for the human albumin drug preparation due to the blockage of the leakage hole. In contrast, the high voltage leak detection method effectively avoided undetected leakage caused by the blockage of the leakage hole by the liquid medicine. The test voltage was set at 9 kV, with a threshold of 15 W. Method validation demonstrated that the high voltage leak detection method exhibited good repeatability, intermediate precision, accuracy, and durability, with a leakage detection limit of 1 μm.

The high-voltage leak detection method can serve as an effective means of inspecting the sealing integrity of the injection bottle packaging system for human albumin injection. The procedure is straightforward, and the results are both accurate and reliable, while also being non-destructive to the packaging. This method is well-suited for the sealing integrity inspection of commercial product packaging systems.

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 establish a high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (HPLC-Q TOF MS) method for the simultaneous analysis of illegal 4 anti-obesity small molecule drugs and 4 glucagon-like peptide-1 (GLP-1) peptide additives.

The samples were extracted by ultrasound using 50% acetonitrile as the extraction solvent. After centrifugation, the supernatant was taken and separated using an Agilent EC-_C18 chromatographic column (150 mm×3.0 mm, 2.7 μm). Acetonitrile (0.1% formic acid) and water (0.1% formic acid) were used as mobile phases, with gradient elution at a flow rate of 0.3 mL·min^-1 and column temperature of 40 ℃. Adopting positive ion full scanning and target ion secondary fragment scanning methods, with a fragmentation voltage of 150 V. The scanning range of the primary mass spectrometry was m/z 100-3 200, and the scanning range of the secondary mass spectrometry was m/z 50-3 200, with a scanning speed of 1 mass spectrum per second. Establish a data spectral library based on the chromatographic retention time, primary mass spectrometry, and secondary mass spectrometry information of the reference standard, and confirmed the structure through database comparison.

The screening detection limit for peptides was 0.5 µg·mL^-1, while small molecular drugs was 0.05 µg·mL^-1. The recoveries were in the range of 79.4% to 115.8%，with the relative standard deviations of 0.21% to 9.7%. Using this method, 20 batches of anti-obesity drugs were tested, in which semaglutide was identified in 4 samples and sibutratmine was identified in 1 batche.

Compared with the complementary method No. 2012005 by the China Food and Drug Administration, the method established in this study can simultaneously analyze small and large molecules (the relative molecular mass<5 000), featuring high efficiency and accuracy.

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.

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 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 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 discuss the influence of the pharmaceutical excipients on the nitrosamine formation in drugs by studying the nitrate and nitrite in native starch and modified starch.

A methodologyfor the determination of nitrate and nitrite in starch was developed by ion chromatography with a suppressed conductivity detector. A Dionex IonPac anion-exchange column (AS11-HC, 4 mm, 2×250 mm) at a temperature of 30 ℃ was utilized. An electrolytically generated potassium hydroxide solution with a concentration of 0.015 mol·L^-1 was delivered at a rate of 1.0 mL·min^-1. The injection volume was 25 µL. The method was validated in terms of specificity, linearity, LOD & LOQ, precision, accuracy and robustness.

119 samples from 9 different categories of native starch and modified starch were analyzed. Nitrate was detected in all 38 samples of native starch in a range of 1 - 70 µg·g^-1, and identified in 55 out of 81 samples of modified starch in a range of 2 - 1 110 µg·g^-1, Among the modified starch samples, dextrin had the maximum concentration of nitrate. Similarly, nitrite was detected in 33 out of 38 samples of native starch in a range of 0.3 - 2.5 µg·g^-1, and 17 out of 81 samples of modified starch in a range of 1.1 - 13.0 µg·g^-1.

This method has been proven to be suitable for determining the nitrate and nitrite in starch based pharmaceutical excipients. The controlling of nitrate and nitrite in the pharmaceutical excipients is an important part of the nitrosamine risk control in the drugs and should be given sufficient attention.

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.