Wen-Xin Decoction ameliorates vascular endothelium dysfunction via the PI3K/AKT/eNOS pathway in experimental atherosclerosis in rats.

BACKGROUND: Nitric oxide (NO) is the most powerful vasodilator that inhibits leukocyte adhesion, platelet aggregation, and vascular smooth muscle cell proliferation. However, excessive NO can cause lipid peroxidation and direct endothelial cell damage. Therefore, investigation of the role of NO in artherosclerosis development is important. Wen-Xin Decoction (WXD) has been shown to relieve myocardial ischemia reperfusion injury and prevent leukocyte adhesion and invasion; in addition, it can accelerate angiogenesis and prevent platelet activation and aggregation. In this study, we focused on the NO pathway to further clarify the protective effects of WXD on the vascular endothelium in rat models of artherosclerosis. METHODS: Wistar rats were randomly divided into a normal group (n = 10) and a model group (n = 75). Rat models of atherosclerosis were generated by intraperitoneal vitamin D3 (3 months) injections and administration of a high-fat diet (3 months with vitamin D3 and 2 months alone). The model rats were randomly divided into five groups (n = 15 each): model (saline), atorvastatin (4.8 mg/kg/d atorvastatin), high-dose WXD (9 g/kg/d), medium-dose WXD (4.5 g/kg/d), and low-dose WXD (2.25 g/kg/d) groups. Each group received continuous drug or saline administration (suspended liquid gavage) for 30 days, following which all animals were sacrificed. The ultrastructure and histopathological changes of vascular endothelial cells and the expression of PI3K/AKT/eNOS and iNOS in the thoracic aorta tissue were analyzed. RESULTS: WXD increased NO levels, modulated the NO/ET-1 ratio, and promoted repair of the injured vascular endothelium in a dose-dependent manner. At a high dose, WXD regulated the NO/ET-1 ratio as effectively as atorvastatin; furthermore, it increased NO levels within the physiological range to prevent endothelial damage caused by excessive NO expression. Real-time polymerase chain reaction and Western blot analysis showed that WXD significantly upregulated the mRNA and protein expressions of PI3K, AKT, and eNOS mRNA and significantly increased AKT and eNOS phosphorylation. CONCLUSIONS: Our results suggest that WXD protects and maintains the integrity of the vascular endothelium by activating the PI3K/AKT/eNOS pathway, decreasing iNOS expression, and promoting the release of physiological NO levels.

The application of metabolomics in traditional Chinese medicine opens up a dialogue between Chinese and Western medicine.

Metabolomics provides an opportunity to develop the systematic analysis of the metabolites and has been applied to discovering biomarkers and perturbed pathways which can clarify the action mechanism of traditional Chinese medicines (TCM). TCM is a comprehensive system of medical practice that has been used to diagnose, treat and prevent illnesses more than 3000 years. Metabolomics represents a powerful approach that provides a dynamic picture of the phenotype of biosystems through the study of endogenous metabolites, and its methods resemble those of TCM. Recently, metabolomics tools have been used for facilitating interactional effects of both Western medicine and TCM. We describe a protocol for investigating how metabolomics can be used to open up 'dialogue' between Chinese and Western medicine, and facilitate lead compound discovery and development from TCM. Metabolomics will bridge the cultural gap between TCM and Western medicine and improve development of integrative medicine, and maximally benefiting the human.

Ultra-performance liquid chromatography tandem mass spectrometry combined with automated MetaboLynx analysis approach to screen the bioactive components and their metabolites in Wen-Xin-Formula.

Wen-Xin-Formula (WXF), a famous traditional prescription, has been widely used to treat myocardial ischemia syndrome for thousands of years. However, the constituents absorbed into blood after oral administration of WXF remain unknown. Here, an integrative ultra performance liquid chromatography coupled with electrospray ionization/quadrupole-time-of-flight mass spectrometry (UPLC-ESI-Q-TOF-MS) combined with the MetaboLynx approach was established to investigate the absorbed constituents in rats after oral administration of WXF. A hyphenated electrospray ionization and quadrupole-time-of-flight analyzer was used for the determination of accurate mass of the molecule and fragment ions. With this rapid and automated analysis method, a total of 32 peaks were tentatively characterized in vivo based on MS and MS/MS data and comparison with available databasess, 26 of which were parent components and six metabolites. These components mainly were ginsenosides, paeoniflorin, galloyl glucose, berberis alkaloids, phenolic, phenolic glycosides and unsaturated fatty acids, glucuronide products of original berberis alkaloids. The present study demonstrates that integrative UPLC-ESI-Q-TOF-MS technique and MetaboLynx data processing method were successfully applied for the rapid discovery of potentially bioactive components and metabolites from WXF, and proved that the established method could help to explore the effective substances for further research into WXF.

Antiviral activity of Jinchai capsule against influenza virus.

OBJECTIVE: To evaluate the effect on influenza virus of Jinchai, a capsule made of Traditional Chinese Medicine. METHODS: Madin-darby canine kidney (MDCK) cells were infected with the FM1 strain of influenza virus A (subtype H1N1) in vitro. They were used to explore how Jinchai affected cell adsorption, cell membrane fusion, transcription and replication of the influenza virus. Hemagglutinin (HA) protein, intracellular pH, and influenza virus protein acid (PA) polymerase subunit were detected with confocal microscopy and real-time fluorescent quantitative polymerase chain reaction. RESULTS: Jinchai significantly reduced the expression of HA and PA polymerase subunit mRNA in infected MDCK cells. Jinchai also significantly decreased intracellular pH in infected cells. CONCLUSIONS: Jinchai had strong anti-influenza activity against the influenza virus. It weakened the ability of the influenza virus to adsorb to cell wall and fuse with cell membranes in the early infection stage, and inhibited the transcription and replication of the virus.

Metabolomics study of intervention effects of Wen-Xin-Formula using ultra high-performance liquid chromatography/mass spectrometry coupled with pattern recognition approach.

Metabolomics is a new approach based on the systematic study of the full complement of small molecular metabolites in a biological sample. It could map the perturbations of early biochemical changes on diseases and hence provides an opportunity to develop predictive biomarkers that can result in earlier intervention and possess valuable insights into the mechanisms of diseases. Given the poor diagnosis of myocardial ischemia syndrome (heart-qi deficiency, HQD), biomarkers of great significance are urgently needed. Fortunately, metabolomics may offer the possibility of identifying marker metabolites and pathways activated in HQD. This paper was designed to explore globally metabolomics characters of the HQD and the therapeutic effects of traditional Chinese medicine Wen-Xin-Formula (WXF). Serum biochemical analysis and histopathological examinations were simultaneously performed. Global metabolic profiling with UHPLC/MS (ultra high-performance liquid chromatography-mass spectrometry), multivariate analysis and database searching were performed to discover differentiating metabolites. Seventeen biomarkers were identified and pathway analysis tools suggest that the glycolysis or gluconeogenesis metabolism, biosynthesis of unsaturated fatty acids metabolism, fatty acid biosynthesis and purine metabolism networks were acutely perturbed by HQD. Of note, WXF has potential pharmacological effect through regulating multiple perturbed pathways to normal state, correlates well to the assessment of biochemistry and histopathological assay. Overall, this study successfully demonstrated that the power of metabolomics in unraveling protective effects of WXF and these findings may help better understand the mechanisms of disease, and the underlying pathophysiologic processes.

Expert consensus on the treatment of rheumatoid arthritis with Chinese patent medicines.

BACKGROUND: Chinese patent medicines (CPMs) are widely used for the treatment of rheumatoid arthritis (RA) in China and especially by Western biomedical doctors who are not well trained in TCM. Thus, it is important to create a guideline or an expert consensus so that the CPMs are used correctly. METHODS: The Delphi technique was used to generate the expert consensus. Twenty-eight (28) integrative medicine rheumatologists joined the consensus. A questionnaire regarding the general therapeutic principles, the categories, and the indications for the specific CPMs used for RA treatment was devised for this study. RESULTS: More than 80% of the experts agreed on the following therapeutic principles: CPM could be used to treat all patients with RA (82.10%), CPM should be used under the guidance of Traditional Chinese Medicine (TCM) (100%), and CPM could be used for active RA treatment in combination with Western medicine (WM) (85.71%), but could be used alone (92.86%) for the treatment of inactive RA. In addition, CPM and WM should be taken separately (82.14%), although the CPM could reduce the side-effects of the WMs if used in combination with CPM (96.43%). For the treatment of active RA, the CPMs were recommended by more than 50% of the experts included the Leigongteng Duogan tablets (85.7%), Zhengqing Dengtongning tablets (64.3%), and Simiao pills (53.6%). Alternatively, for the treatment of inactive RA, Duhuo Jisheng mixture (71.4%), Yishen Juanbi pills (53.6%), and Wangbi electuary (50.0%) were recommended. Total paeonia glucoside capsules were recommended for the active (50.0%) and inactive RA (64.3%) treatment. The indications of each CPM were specified according to the symptoms related to the TCM pattern classification. CONCLUSIONS: This expert consensus regarding the treatment of RA with various CPMs was formed to aid WM doctors in the correct use of CPMs.

[Effects of jin chai antiviral capsule on IFITM3 expression in mice].

This study is to investigate the treatment of Jin Chai antiviral capsule for influenza virus FM1/47 (H1N1) infection. The model of pneumonia was established by dropping influenza virus into the nose of normal mice, real-time PCR and Western blot technique were used to detect the virus load and the interferoninducible transmembrane protein3 (IFITM3) in lung of mice at the 1st day, 3rd day, 5th day and 7th day after affected. The results showed that Jin Chai antiviral capsule in large, middle, small dose groups can decrease virus load significantly at each time point, after being affected (P<0.05, P<0.01), Jin Chai antiviral capsule can increase the interferoninducible transmembrane protein3 in lung of mice, large dose groups are significantly higher in expression of IFITM3 compared with model group at each time point (P<0.05, P<0.01). Middle dose groups are significantly higher in expression of IFITM3 compared with model group at the 3th day and the 5th day (P<0.05), small dose groups are significantly higher in expression of IFITM3 compared with model group at the 3th day (P<0.05). It can be concluded that Jin Chai antiviral capsule exerts antiviral effects against influenzavirus by raised expression of IFITM3.

To compare the efficacy of two kinds of Zhizhu pills in the treatment of functional dyspepsia of spleen-deficiency and qi-stagnation syndrome: a randomized group sequential comparative trial.

BACKGROUND: In Traditional Chinese Medicine (TCM) theory, functional dyspepsia (FD) can be divided into different syndromes according to different clinical symptoms and signs, and the most common one is spleen-deficiency and qi-stagnation syndrome that can be treated by Chinese traditional patent medicine--two kinds of Zhizhu pills, between which the primary difference in ingredients is that one contains immature orange fruit of Citrus aurantium L.(IFCA) and the other contains that of Citrus sinensis Osbeck (IFCS). The trial's objective was to compare the efficacy of two kinds of Zhizhu pills on symptom changes in patients with FD of spleen-deficiency and qi-stagnation syndrome. METHODS: A randomized, group sequential, double-blinded, multicenter trial was conducted in patients with FD of spleen-deficiency and qi-stagnation syndrome at 3 hospitals in Beijing between June 2003 and May 2005. Participants were randomly allocated into two groups (IFCA group and IFCS group) in a 1:1 ratio, and respectively took one of the two kinds of Zhizhu pills orally, 6 g each time, 3 times a day, for 4 weeks. Statistical analysis was performed with use of a group sequential method, the triangular test (TT). RESULTS: A total of 163 patients were randomized, and 3 patients were excluded from analysis because of early dropouts, leaving 160 patients (IFCA group: n = 82; IFCS group: n = 78) for statistical analysis. Three interim analyses were done after 62, 116, and 160 patients had completed their 4-week treatment, respectively. At the third interim analysis, the sample path crossed the upper boundary and the trial was stopped, the cure-markedly effective rates were 45% for IFCS group and 67% for IFCA group, respectively, the one-sided p-value was 0.0036, the median unbiased estimate of the odds ratio (OR) for the benefit of IFCA relative to IFCS was 2.91 with 95%CI: 1.40 to 6.06.No adverse events were observed in the two groups. CONCLUSIONS: Zhizhu pills containing IFCA was superior to Zhizhu pills containing IFCS in the treatment of FD of spleen-deficiency and qi-stagnation syndrome. The application of group sequential analysis in clinical trials of TCM may offer some financial and ethical benefits. TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR): ChiCTR-TRC-00000485.

The role of Chinese medicine in the treatment of chronic diseases in China.

Chinese medicine (CM) has a long history of experience and proven successful treatment for chronic diseases and has also played an important role in the provision of health care in China. Patients with chronic diseases are happy to accept CM and physicians are willing to use CM to relieve patients suffering from chronic illnesses. The Chinese health authorities encourage CM development to meet the requirements for the treatment of chronic diseases. CM products are an essential part of medications that have a predominant role in the prevention and treatment of chronic diseases in China. A large number of CM clinical studies, including a substantial number of available randomized controlled trials and systematic reviews, have shown that CM is effective and safe in the treatment of chronic diseases. Although the efficacies of some evaluated CM therapies remain uncertain, it is worth assessing them by using CM pattern (Zheng or syndrome) differentiation to verify treatment outcomes. CM is considered to have a better safety profile compared to pharmaceutical chemicals, but inappropriate applications of CM also makes the safety issues a hot discussed subject. As a medical system, CM should be able to provide worldwide contribution for the patients who are suffering from chronic diseases. The application of CM pattern classification in diagnosis with corresponding prescribed treatment using herbal formulae in the relief of chronic diseases can be linked with modern biomedical parameters (biomarkers) as treatment outcomes. These outcome parameters, together with the patients' reported quality of life assessment, can provide innovative approaches for evidence-based estimation of the efficacy of CM treatment in chronic diseases.

Gualou xiebai banxia decoction inhibits NF-kappa B-dependent inflammation in myocardial ischemia-reperfusion injury in rats.

OBJECTIVE: To evaluate the myocardial protective effect of Gualou Xiebai Banxia decoction GXBD) and explore the mechanisms of inhibition of NF-kappa B activation and blockade of inflammatory responses induced by ischemia-reperfusion in rats. METHODS: Twenty-four Sprague Dawley (SD) rats were randomly divided into three groups. Rats in the treatment group received GXBD (13 g crude drug/kg) for three weeks, while rats in the model control and normal control groups received equal volumes of distilled water. On the 22nd day, rats in the ischemia-reperfusion (I/R) control and GXBD-treated groups underwent 30 min occlusion of the left anterior descending (LAD) coronary artery, followed by 120 min reperfusion. Electrocardiogram was recorded, and the activities of cardiac enzymes, cytokines, and NF-kappaB were assessed after I/R. RESULTS: Compared with the I/R control group, GXBD treatment restored the activity of the specific myocardial-injury marker creatine kinase (CK) and lactate dehydrogenase (LDH), and inhibited the inflammatory response involving the nuclear factor-kappaB (NF-KB) pathway, including down-regulation of interleukin (IL)-1beta and IL-6, and up-regulation of IL-10 gene expression. CONCLUSION: GXBD strongly reduced myocardial impairment in our I/R model, including inhibition of NF-kappaB activation and inflammatory cytokine responses.

[Effect of Pingtang recipe containing drug-serum on INS-1 pancreatic beta cells lipoapoptosis].

OBJECTIVE: To observe the effect and mechanism of Pingtang Recipe containing drug-serum (DS-PTR) in improving INS-1 beta pancreatic cells lipoapoptosis. METHODS: Experimental INS-1 beta cells were divided into 5 groups (6 pools for each group), namely, the blank control group treated with rat's serum (C), the other 4 model groups induced into lipoapoptosis by palmitic acid and treated respectively by rat's serum (M), high, middle and low dose DS-PTR (DSh, DSm and DSI). Cell apoptosis was detected by TUNEL staining; Caspase-3 activity of cells was measured by chemiluminescence method; intracellular production of reactive oxygen species (ROS) was detected by DCHF-DA incorporation, and expressions of uncoupling protein-2 (UCP-2) was determined by RT-PCR. RESULTS: INS-1 beta cell apoptosis in Group M was significantly higher than that in Group C (P < 0.01), while that showed a decreased trend in the three DS-PTR treated groups. Caspase-3 activity was enhanced in Group M, it decreased significantly in Group DSm (P < 0.05). The over-produced ROS in cells after modeling was inhibited in Groups DSm and DSI (P < 0.05), meantime, expression of UCP-2 excited by PA (2.244 +/- 0.421) was reduced significantly in Group DSI and Group DSm to 1.286 +/- 0.373 (P < 0.01) and 1.627 +/- 0.348 (P < 0.05) respectively. CONCLUSION: DS-PTR shows a protective effect on INS-1 beta pancrentic cells against lipoapoptosis, which is possibly play its mechanism through regulating ROS and UCP-2.

Clinical studies with traditional Chinese medicine in the past decade and future research and development.

Traditional Chinese medicine (TCM) is currently considered a complementary or alternative medical system in most Western countries and has been increasingly accepted worldwide. More and more clinical trials on TCM have been conducted internationally, and scientists worldwide are becoming increasingly interested in the evaluation of clinical efficacy of TCM based on clinical trials. This paper reviews the situation of clinical trials on TCM in the past decade, including systematic reviews about clinical trials either focusing on the treatment of disease with TCM approaches or focusing on one herbal product, conduction of clinical trials on TCM either with randomization and controlled methods or general observation. Some general issues on the conduct of clinical trials on TCM, such as randomization, control, quality of life (QOL), patient reported outcomes (PROs) and biomarkers, quality control, safety evaluation and case studies, are discussed, and accordingly some suggestions are proposed.

Pharmacokinetics-based elucidation on disparity in clinical effectiveness between varieties of Zhi Zhu Wan, a Traditional Chinese Medical formula.

AIM: Zhi Zhu Wan (ZZW) is a classical Chinese medical formulation used for the treatment of functional dyspepsia that attributed to Spleen-deficiency Syndrome. ZZW contains Atractylodes Rhizome and Fructus Citrus Immaturus, the later originates from both Citrus aurantium L. (BZZW) and Citrus sinensis Osbeck (RZZW). The present study is designed to elucidate disparities in the clinical efficacy of two ZZW varieties based on the pharmacokinetics of naringenin and hesperetin. MEHTOD: After oral administration of ZZWs, blood sample was collected from healthy volunteers at designed time points. Naringenin and hesperetin were detected in plasma by RP-HPLC, pharmacokinetic parameters were processed using mode-independent methods with WINNONLIN. RESULTS: After oral administration of BZZW, both naringenin and hesperetin were detected in plasma, and demonstrated similar pharmacokinetic parameters. Ka was 0.384+/-0.165 and 0.401+/-0.159, T(1/2(ke))(h) was 5.491+/-3.926 and 5.824+/-3.067, the AUC (mg/Lh) was 34.886+/-22.199 and 39.407+/-19.535 for naringenin and hesperetin, respectively. However, in the case of RZZW, only hesperetin was found in plasma, but the pharmacokinetic properties for hesperetin in RZZW was different from that in BZZW. T(max) for hesperetin in RZZW is about 8.515h, and its C(max) is much larger than that of BZZW. Moreover, it was eliminated slowly as it possessed a much larger AUC value. CONCLUSION: The distinct therapeutic orientations of the Chinese medical formula ZZWs with different Fructus Citrus Immaturus could be elucidated based on the pharmacokinetic parameters of constituents after oral administration.

Comparative study on the protective effects of Yinchenhao Decoction against liver injury induced by alpha-naphthylisothiocyanate and carbon tetrachloride.

OBJECTIVE: To optimize the animal model of liver injury that can properly represent the pathological characteristics of dampness-heat jaundice syndrome of traditional Chinese medicine. METHODS: The liver injury in the model rat was induced by alpha-naphthylisothiocyanate (ANIT) and carbon tetrachloride (CCl(4) ) respectively, and the effects of Yinchenhao Decoction (, YCHD), a proved effective Chinese medical formula for treating the dampness-heat jaundice syndrome in clinic, on the two liver injury models were evaluated by analyzing the serum level of alanine aminotransferase (ALT), asparate aminotransferase (AST), alkaline phosphatase (ALP), malondialchehyche (MDA), total bilirubin (T-BIL), superoxide dismutase (SOD), glutathione peroxidase (GSH-PX) as well as the ratio of liver weight to body weight. The experimental data were analyzed by principal component analytical method of pattern recognition. RESULTS: The ratio of liver weight to body weight was significantly elevated in the ANIT and CCl(4) groups when compared with that in the normal control (P<0.01). The contents of ALT and T-BIL were significantly higher in the ANIT group than in the normal control (P<0.05,P<0.01), and the levels of AST, ALT and ALP were significantly elevated in CCl(4) group relative to those in the normal control P<0.01). In the YCHD group, the increase in AST, ALT and ALP levels was significantly reduced (P<0.05, P<0.01), but with no significant increase in serum T-BIL. In the CCl(4) intoxicated group, the MDA content was significantly increased and SOD, GSH-PX activities decreased significantly compared with those in the normal control group, respectively (P<0.01). The increase in MDA induced by CCl(4) was significantly reduced by YCHD P<0.05). CONCLUSION: YCHD showed significant effects on preventing liver injury progression induced by CCl(4), and the closest or most suitable animal model for damp-heat jaundice syndrome may be the one induced by CCl(4).

Pharmacokinetics of cimifugin in rat plasma after oral administration of the extract of Saposhnikovia divaricatae root. Determination of cimifugin by high performance liquid chromatography coupled with solid phase extraction.

High performance liquid chromatography (HPLC) coupled with the solid phase extraction method was developed for determining cimifugin (a coumarin derivative; one of Saposhnikovia divaricatae's constituents) in rat plasma after oral administration of Saposhnikovia divaricatae extract (SDE), and the pharmacokinetics of cimifugin either in SDE or as a single compound was investigated. The HPLC analysis was performed on a commercially available column (4.6 mm x 200 mm, 5 pm) with the isocratic elution of solvent A (Methanol) and solvent B (Water) (A:B=60:40) and the detection wavelength was set at 250 nm. The calibration curve was linear over the range of 0.100-10.040 microg/mL. The limit of detection was 30 ng/mL. At the rat plasma concentrations of 0.402, 4.016, 10.040 microg/mL, the intra-day precision was 6.21%, 3.98%, and 2.23%; the inter-day precision was 7.59%, 4.26%, and 2.09%, respectively. The absolute recovery was 76.58%, 76.61%, and 77.67%, respectively. When the dosage of SDE was equal to the pure compound calculated by the amount of cimifugin, it was found to have two maximum peaks while the pure compound only showed one peak in the plasma concentration-time curve. The pharmacokinetic characteristics of SDE showed the superiority of the extract and the properties of traditional Chinese medicine.

Hydrolysis of flavanone glycosides and degradation of the corresponding aglycones from dried immature Citrus fruit by human fecal flora in vitro.

The hydrolysis of the flavanone glycosides contained in dried immature Citrus fruit, originating from Citrus aurantium L (Family Rutaceae), and degradation of their aglycones in human fecal flora have been analyzed. High-performance liquid chromatography was used to determine the flavanone glycosides and their corresponding aglycones in human fecal flora. The separation of compounds was performed with an ODS column by isocratic and stepwise gradient elution with 0.5 % (v/v) acetic acid-acetonitrile. As a result, the hydrolysis rate of hesperidin and narirutin (flavanone rutinoside) was faster than that of naringin and neohesperidin (flavanone neohesperioside). When the half-life time of each flavanone glycoside was carefully calculated (under the mixed conditions with the human fecal flora), hydrolysis of the flavanone rutinoside turned out to be approximately two times faster than of flavanone neohesperioside. The observed degradation rates of both aglycones was found not to be different. Therefore, it seems that the hydrolysis rate of flavanone glycosides in dried immature citrus fruit with human fecal flora is closely related to the steric hindrance of the sugar. This finding might be effectively used for further pharmacokinetics research on the flavanone glycosides of dried immature Citrus fruit.

Metabolic urinary profiling of alcohol hepatotoxicity and intervention effects of Yin Chen Hao Tang in rats using ultra-performance liquid chromatography/electrospray ionization quadruple time-of-flight mass spectrometry.

This paper was designed to study metabonomic characters of the hepatotoxicity induced by alcohol and the intervention effects of Yin Chen Hao Tang (YCHT), a classic traditional Chinese medicine formula for treatment of jaundice and liver disorders in China. Urinary samples from control, alcohol- and YCHT-treated rats were analyzed by ultra-performance liquid chromatography/electrospray ionization quadruple time-of-flight mass spectrometry (UPLC/ESI-QTOF-MS) in positive ionization mode. The total ion chromatograms obtained from the control, alcohol- and YCHT-treated rats were easily distinguishable using a multivariate statistical analysis method such as the principal components analysis (PCA). The greatest difference in metabolic profiling was observed from alcohol-treated rats compared with the control and YCHT-treated rats. The positive ions m/z 664.3126 (9.00 min) was elevated in urine of alcohol-treated rats, whereas, ions m/z 155.3547 (10.96 min) and 708.2932 (9.01 min) were at a lower concentration compared with that in urine of control rats, however, these ions did not indicate a statistical difference between control rats and YCHT-treated rats. The ion m/z 664.3126 was found to correspond to ceramide (d18:1/25:0), providing further support for an involvement of the sphingomyelin signaling pathway in alcohol hepatotoxicity and the intervention effects of YCHT.

Simultaneous determination of 6,7-dimethylesculetin and geniposide in rat plasma and its application to pharmacokinetic studies of Yin Chen Hao Tang preparation.

A method for the rapid and simultaneous determination of 6,7-dimethylesculetin (CAS 120-08-1) and geniposide (CAS 24512-63-8) in rat plasma has been developed, using validated high performance liquid chromatography (HPLC) with solid phase extraction (SPE). The HPLC analysis was performed on a commercially available column (200 mm x 4.6 mm, 5 microm) with acetonitrile-methanol-0.1% aqueous formic acid as mobile phase and the UV detection at 343 nm and 238 nm for 6,7-dimethylesculetin and geniposide, respectively. The calibration curves for 6,7-dimethylesculetin and geniposide were linear over the range 0.4-25.6 microg/mL and 1.12-71.68 microg/mL, respectively. The lower limits of quantitation were 0.40 microg/ mL and 1.12 microg/mL, and the lower limits of detection were 0.06 microg/mL and 0.09 microg/ mL, respectively. The intra-day and inter-day precision for 6,7-dimethylesculetin and geniposide were < 5%, whereas the absolute recovery percentages were > 74%. A successful application of the developed HPLC analysis was demonstrated for the pharmacokinetic study of a Traditional Chinese Medicine formula of Yin Chen Hao Tang preparation.

Simultaneous determination by UPLC-ESI-MS of scoparone, capillarisin, rhein, and emodin in rat urine after oral administration of Yin Chen Hao Tang preparation.

A simple, sensitive, and validated method was developed for simultaneous determination of scoparone, capillarisin, rhein, and emodin in rat urine by ultra-performance liquid chromatography/electrospray ionization quadruple time-of-flight mass spectrometry (UPLC-MS). The urinary samples were analyzed on an Acquity UPLC BEH C18 1.7 microm 2.1x50 mm column. Scoparone, capillarisin, rhein, and emodin in rat urine were simultaneously analyzed with good separation. The lower limits of detection were 6.0, 9.0, 7.0, and 3.0 ng/mL, and the lower limits of quantification were 20.0, 33.0, 24.0, and 12.0 ng/mL for scoparone, capillarisin, rhein, and emodin, respectively. The intra- and inter-day precisions (RSD) were less than 9%. The intra- and inter-accuracies were found to be in the range of 94.14-104.54% for scoparone, 101.72-107.34% for capillarisin, 95.24-103.59% for rhein, and 101.32-107.82% for emodin at three concentration levels. The absolute recoveries for scoparone, capillarisin, rhein, and emodin were not less than 77.0%. The developed method has been applied to determine scoparone, capillarisin, rhein, and emodin in rat urine after oral administration of Yin Chen Hao Tang preparation, a traditional Chinese medicine formulation widely used in China for treatment of jaundice and liver disorders.