Anti-inflammatory effects of Samsoeum, a Korean medicine for health insurance, on chronic bronchitis caused by lipopolysaccharide in rats.

BACKGROUND: Samsoeum (SSE), a Korean medicine, has been used to treat upper respiratory infection including residual coughs after catching a cold, and colds in patients with gastrointestinal disorder. In this study, we investigated the inhibitory effect of SSE against lipopolysaccharide (LPS)-induced bronchitis and characterized its optimal dosing range based on the improvement of SSE concentrations. MATERIALS AND METHODS: Male Sprague Dawley rats were intra-nasally administered LPS on day 0, 3 and 6. 2 g kg-1 dose of SSE for rat was determined by the human equivalent dose formula and orally administered once a day from day 3 to day 6. To clarify the optimal administration dose of SSE, various doses including 0.5 (1/4 fold), 1 (1/2 fold), 6 (3 fold), 12 (6 fold), 24 (12 fold) and 36 g kg-1 (18 fold) were also orally administered. In addition, the molecular mechanism of SSE in mucin hyperproduction was investigated in LPS-sensitized A549 cells. RESULTS: Oral administration of SSE ameliorated alveolar wall thickening and inflammatory cell infiltration of lung tissues in LPS-induced bronchitis at doses of 1/4 fold, 1/2 fold and 1 fold. The total cell and neutrophil numbers in bronchoalveolar lavage fluid (BALF) were reduced in the SSE-treated groups compared with the LPS group. In addition, 0.5, 1 and 2 g kg-1 of SSE suppressed LPS-induced mucin glycoprotein 5AC (MUC5AC) production in BALF. Furthermore, SSE treatment significantly inhibited the pro-inflammatory cytokines, resulting in the decrease of MUC5AC production by the JAK1/STAT6 signaling pathway. CONCLUSIONS: 1, 2 and 6 g kg-1 of SSE ameliorated chronic bronchitis by inhibiting LPS-induced neutrophil infiltration and MUC5AC release in BALF. These findings suggested that SSE with 0.5-3-fold of general daily intake dose would be a therapeutic agent for chronic bronchitis.

A 90-Day Repeated Oral Dose Toxicity Study of Alismatis Rhizoma Aqueous Extract in Rats.

Alismatis rhizoma (AR), the dried rhizome of Alisma orientale (Sam.) Juzep, is a well-known, traditional medicine that is used for the various biological activities including as a diuretic, to lower cholesterol and as an anti-inflammatory agent. The present study was carried out to investigate the potential toxicity of the Alismatis rhizoma aqueous extract (ARAE) following 90-day repeated oral administration to Sprague-Dawley rats. ARAE was administered orally to male and female rats for 90 days at 0 (control), 500, 1,000 and 2,000 mg/kg/day (n = 10 for male and female rats for each dose). Additional recovery groups from the control group and high dose group were observed for a 28-day recovery period. Chromatograms of ARAE detected main compounds with four peaks. Treatment-related effects including an increase in the red blood cells, hemoglobin, hematocrit, albumin, total protein, and urine volume were observed in males of the 2,000 mg/kg/day group (p < 0.05). However, the diuretic effect of ARAE was considered, a major cause of hematological and serum biochemical changes. The oral no-observed-adverse-effect level (NOAEL) of the ARAE was > 2,000 mg/kg/day in both genders, and no target organs were identified.

Simultaneous UPLC-MS/MS determination of four components of Socheongryong-tang tablet in human plasma: Application to pharmacokinetic study.

The purpose of this study was to develop a method for simultaneous analysis of schizandrin, ephedrine, paeoniflorin, and cinnamic acid as constituents of Socheongryong-tang tablet in human plasma using UPLC-MS/MS. These four components were separated using water containing 0.01% formic acid and methanol as a mobile phase by gradient elution at a flow rate of 0.3 mL/min with a HALO-C18 column (2.1 mm × 100 mm, 2.7 µm particle size). Quantitation was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique operated in multiple reaction monitoring mode. Mass transitions were m/z 432.9 → 384.1 for schizandrin, 165.8 → 148.1 for ephedrine, 525.0 → 449.2 for paeoniflorin, 146.8 → 102.9 for cinnamic acid, and 340.0 → 324.0 for papaverine as internal standard. Liquid-liquid extraction and protein precipitation with ethyl acetate-methanol (1:2, v/v) were used to obtain these four components. Chromatograms showed high resolution, sensitivity, and selectivity without interference by plasma constituents. Calibration curves of schizandrin, ephedrine, paeoniflorin, and cinnamic acid in human plasma ranged from 0.02 to 8 ng/mL, 0.5 to 200 ng/mL, 0.2 to 80 ng/mL, and 1 to 400 ng/mL, respectively. Calibration curves of each analyte displayed excellent linearity, with correlation coefficients > 0.99. For all four components, both intra- and inter-day precisions (CV%) were <5.99%. The accuracy was 99.35-103.30% for schizandrin, 98.48-104.38% for ephedrine, 97.06-103.34% for paeoniflorin, and 99.97-104.36% for cinnamic acid. This analytical method developed in this study satisfied the criteria of international guidance. It could be successfully applied to pharmacokinetic studies of schizandrin, ephedrine, paeoniflorin, and cinnamic acid after oral administration of Socheongryong-tang tablet to humans.

Development of Simultaneous Analysis of Thirteen Bioactive Compounds in So-Cheong-Ryong-Tang Using UPLC-DAD.

So-Cheong-Ryong-Tang, which is a standardized Korean medicine of the National Health Insurance, is a traditional prescription for the treatment of allergic rhinitis, bronchitis, and bronchial asthma. Simultaneous analysis and development of SCRT is essential for its stability, efficacy, and risk management. In this study, a simple, reliable, and accurate method using ultrahigh-performance liquid chromatography (UPLC) fingerprinting with a diode array detector (DAD) was developed for the simultaneous analysis. The chromatographic separation of the analytes was performed by an ACQUITY UPLC BEH C18 column (1.7 µM, 2.1 × 100 mm, Waters) with a mobile phase of water containing 0.01% (v/v) phosphoric acid and acetonitrile containing 0.01% (v/v) phosphoric acid. The flow rate and detection wavelength were set at 0.4 mL/min and 215, 230, 254, and 280 nm. All calibration curves of the thirteen components showed good linearity (R2 > 0.999). The limit of detection and limit of quantification ranged 0.001-0.360 and 0.004-1.200 µg/mL, respectively. The relative standard deviation (RSD) of intra- and interday was less than 2.60%, and the recoveries were within the range 76.08-103.79% with an RSD value of 0.03-1.50%. The results showed that the developed method was simple, reliable, accurate, sensitive, and precise for the quantification of bioactive components of SCRT.

Simultaneous Determination of Decursin, Decursinol Angelate, Nodakenin, and Decursinol of Angelica gigas Nakai in Human Plasma by UHPLC-MS/MS: Application to Pharmacokinetic Study.

Coumarins in Cham-dang-gwi, the dried root of Angelica gigas Nakai (AGN), possess pharmacological effects on anemia, pain, infection, and articular rheumatism. The AGN root containes decursin (D), decursinol angelate (DA), nodakenin, and decursinol (DOH), a major metabolite of D and DA. The aim of this study was to develop a simultaneous determination method for these four coumarins in human plasma using ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Chromatographic separation was performed on dual columns (Kinetex® C18 column and Capcell core C18 column) with mobile phase consisting of water and acetonitrile at a flow rate of 0.3 mL/min using gradient elution. Multiple reaction monitoring was operated in positive ion mode with precursors to product ion transition values of m/z 328.9→228.8, 328.9→228.9, 409.4→248.8, and 246.8→212.9 to measure D, DA, nodakenin, and DOH, respectively. Linear calibration curves were fitted over concentration range of 0.05⁻50 ng/mL for these four components, with correlation coefficient greater than 0.995. Inter- and intra-day accuracies were between 90.60% and 108.24%. These precisions were within 11.19% for all components. The established method was then applied to a pharmacokinetic study for the four coumarins after usual dosing in Korean subjects.

A sensitive UHPLC-MS/MS method for the simultaneous quantification of three lignans in human plasma and its application to a pharmacokinetic study.

The aim of this study was to develop an analytical method to simultaneously analyze schizandrin, schizandrol B, and gomisin N lignans in human plasma using ultra high performance liquid chromatography with tandem mass spectrometry. The three lignans were separated using a mobile phase of water and acetonitrile containing 0.02% acetic acid equipped with a Kinetex C18 column (2.1 mm × 50 mm, 1.7 µm). This analysis was achieved by multiple reaction monitoring mode in an electrospray interface. The mass transitions were m/z 433.1→384.0 for schizandrin, 398.8→367.8 for schizandrol B, and 400.6→299.8 for gomisin N. Liquid-liquid extraction with methyl tert-butyl ether was used to obtain the three lignans. The chromatograms showed high resolution, sensitivity, and selectivity with no interference with plasma constituents. The calibration curves for the three lignans in human plasma were 0.05-50 ng/mL and displayed excellent linearity with correlation coefficients greater than 0.99. Precision for all three lignans was within 11.23%. The accuracy was 88.3-99.0% for schizandrin, 90.6-103.4% for schizandrol B, and 90.2-103.5% for gomisin N. The developed simultaneous analytical method satisfied the criteria of international guidance and could be successfully applied to the pharmacokinetic study of three lignans after oral administration of Schisandrae Fructus extract powder to humans.

Simultaneous determination of imperatorin and its metabolite xanthotoxol in rat plasma and urine by LC-MS/MS and its application to pharmacokinetic studies.

An accurate, precise, selective, and sensitive liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) method was developed for the simultaneous determination of imperatorin (IMP) and its metabolite, xanthotoxol (XAN), in rat plasma and urine samples. The analytes, along with psoralen as an internal standard, were determined by multiple reaction monitoring (MRM) operated in the positive electrospray ionization (ESI) mode. Chromatographic separation was performed on an Acquity UPLC BEH C18 column (50mm×2.1mm, 1.7µm) with a mobile phase consisting of 0.1% formic acid solution and 0.1% formic acid in methanol at a flow rate of 0.3mL/min. The run time was 6min per sample and the injection volume was 5µL. The method had a lower limit of quantification (LLOQ) of 0.25ng/mL for IMP in plasma and urine, and 1ng/mL for XAN in urine. The linear calibration curves were fitted over the range of 0.25-1000ng/mL for IMP in plasma, 0.25-1000ng/mL for IMP in urine, and 1-1000ng/mL for XAN in urine, with correlation coefficients greater than 0.995. The inter- and intra-day accuracies (relative error, RE%) were between -8.5% and 3.5%, and the precisions (relative standard deviation, RSD%) were less than 10.0% for all quality control samples (QCs). The analytes were extracted from rat plasma and urine samples using a liquid-liquid extraction method with the extraction recovery in the range of 60.3-79.1%. A good stability of the analytes was observed in all the analysis procedures. The method was successfully validated and applied to determine the pharmacokinetics of IMP in rat plasma and, for the first time, the metabolite kinetics of IMP to XAN in rat urine after IMP administration.

Simultaneous determination of puerarin and its active metabolite in human plasma by UPLC-MS/MS: application to a pharmacokinetic study.

A rapid, selective and sensitive ultra-performance liquid chromatography (UPLC)-tandem mass spectrometry method about the simultaneous determination of puerarin and its major active metabolite, daidzein, in human plasma was developed and validated in order to investigate the pharmacokinetics (PKs) of Gegen after the usual oral dose administration to human. Chromatography was carried out on a Kinetex C18 column (2.1mm×50mm, 1.7µm) using 0.05% acetic acid in water and 0.05% acetic acid in methanol as mobile phase with a gradient elution. Liquid-liquid extraction with ethyl acetate in acidic condition could remove the interference and minimize the matrix effect of human plasma. The lower limit of quantification in human plasma was 0.2ng/mL for both of compounds, puerarin and daidzein. The calibration curves for puerarin and daidzein in human plasma were linear over all the concentration range of 0.2-100ng/mL with correlation coefficients greater than 0.998. This assay procedure was successfully applied to the PKs of puerarin and daidzein, after the usual oral dose of Gegen extract powder (2.56g, containing 9.984mg puerarin) in human subjects.