Qiongfeng Liao

Simultaneous determination of berberine, palmatine and jatrorrhizine by liquid chromatography–tandem mass spectrometry in rat plasma and its application in a pharmacokinetic study after oral administration of coptis–evodia herb couple

A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed to simultaneously determine berberine, palmatine and jatrorrhizine in rat plasma. After mixing with the internal standard (IS) tetrahydropalmatine, plasma samples were pretreated by protein precipitation with acetonitrile-methanol (1:2, v/v). Chromatographic separation was carried out on a C18 column using a mixture of water (containing 0.1% formic acid) and acetonitrile (30:70, v/v) as mobile phase. The detection was performed by selected reaction monitoring (SRM) mode via electrospray ionization (ESI) source operating in the positive ionization mode. The method was linear over the concentration range of 1.0-250.0 ng/mL for all components. The intra- and inter-day precision values were less than 14.6% and the deviations were within +/-4.0%. The fully validated LC-MS/MS method has been successfully applied to the pharmacokinetic study of berberine, palmatine and jatrorrhizine in rat plasma after oral administration of coptis-evodia herb couple. Three peaks were observed in both individual and mean plasma-concentration curves of berberine, palmatine and jatrorrhizine, which may be attributed to distribution re-absorption and enterohepatic circulation.

Integrated pharmacokinetics of five protoberberine-type alkaloids in normal and insomnic rats after single and multiple oral administration of Jiao-Tai-Wan.

ETHNOPHARMACOLOGICAL RELEVANCE Jiao-Tai-Wan (JTW), an important herbal formula consists of Rhizoma coptidis and Cortex cinnamomi powder, is a famous prescription which has been used for centuries to treat insomnia in Traditional Chinese Medicine. The purpose of this study is to compare the pharmacokinetic properties of five protoberberine-type alkaloids (i.e. berberine, palmatine, coptisine, epiberberine and jatrorrhizine), the main bioactive constituents in JTW, between normal and insomnic rats. We also investigate the differences between single-dose and multiple-dose pharmacokinetics of five protoberberine-type alkaloids. MATERIALS AND METHODS The insomnic rat models were induced by intraperitoneal injection of one-dose para-chlorophenylalanine acid (PCPA). Quantification of five protoberberine-type alkaloids in rat plasma was achieved by using a rapid LC-MS/MS method. Plasma samples were collected at different time points to construct pharmacokinetic profiles by plotting drug concentration versus time and estimate pharmacokinetic parameters. An unpaired Student׳s t test was used for comparisons with SPSS 17.0. RESULTS The five protoberberine-type alkaloids of single-dose normal groups had slow absorption and low bioavailability, as well as a delay of peak time. In the single-dose oral administration, the Cmax and Tmax of five ingredients in insomnic rats had significant differences compared with those of normal rats. In the multiple-dose oral administration, the pharmacokinetic parameters of five protoberberine-type alkaloids varied greatly in insomnic rats. In the normal rats, there were significant differences (P<0.05) in the principal pharmacokinetic parameters such as Cmax and Tmax between single-dose and multiple-dose oral administration. In the insomnic rats, the five ingredients of multiple-dose groups showed better absorption than the single-dose groups. Particularly, three peaks were observed in multiple-dose model group of plasma-concentration curves. CONCLUSIONS The pharmacokinetic behavior of five protoberberine-type alkaloids was described in this paper. In both normal groups and model groups, the pharmacokinetic behavior of multiple-dose had significant differences comparing with the single-dose; either single-dose or multiple-dose, the pharmacokinetic behavior of insomnic rats had significant differences comparing the normal rats. Multiple dosing may improve the absorption of JTW in insomnic rats, which will increase the bioavailability and bring into active role in therapeutical effect.

Isolation, Identification and Activities of Natural Antioxidants from Callicarpa kwangtungensis Chun

Reactive oxygen species leads to some diseases associated with oxidative stress. Callicarpa kwangtungensis Chun (CK) is a common remedy in traditional Chinese medicine and possesses diverse biological activities involving antioxidant properties; its main compounds phenylethanoid glycosides (PG) and flavonoids are always reported as antioxidants. In order to develop CK as a safe and activated antioxidant, our investigation was performed to validate antioxidant properties and assess which types of compounds (similar polarity or similar structure), even which compounds, played the role of antioxidants. The extracted compounds of CK were analyzed qualitatively and quantitatively by HPLC-DAD-ESI-Trap MS and UV for their contents and antioxidant activities. The correlations between antioxidant activities and known contents were respectively counted and a semi-quantitative experiment was designed to screen antioxidant compounds of CK with HPLC-UV. The n-butanol fraction (BF) showed the highest total phenolic and flavonoid contents (TPC, TFC), and three PG (forsythiaside B, poliumoside and acteoside) contents. BF showed the significantly best (P<0.05) activities in most assays. There were significant correlations (P<0.05) between DPPH•, ABTS+•, •O2 − scavenging, Cu2+-chelating, anti-lipidperoxidation activities and TPC. BF also has significant antioxidant activities on CCl4-induced acute liver injury Mice and TBHP-reduced HepG2 cells. Nine PG (forsythiaside B, poliumoside, acteoside, alyssonoside, brandioside and their derivatives) and one flavone (rhamnazin) were screened out as antioxidants. BF in CK contained abundant polyphenolic, which reflected some definite antioxidant properties. The antioxidant compounds consisted at the least of nine PG and one flavone.

Analysis of Essential Oils from Cassia Bark and Cassia Twig Samples by GC-MS Combined with Multivariate Data Analysis

Cinnamomum cassia is one of several species of Cinnamomum which has been widely used as a spice. In this study, the chemical profiles of its bark and twig were characterized by gas chromatography-mass spectrometry (GC-MS) and multivariate data analysis. Principle component analysis (PCA) and orthogonal projection to latent structures discriminate analysis (OPLS-DA) of GC-MS data provided a clear separation between those samples. The result obviously showed that the metabolome of cinnamon bark (CB) and cinnamon twig (CT) is different, and the corresponding loading S-plots revealed that the differential metabolites between the essential oils of CB and CT were trans-cinnamaldehyde, trans-anethole, α-cubebene, γ-muurolene, γ-amorphene, δ-cadinene, (−)-calamenene, and o-methoxycinnamaldehyde. Our study demonstrates that the combination of GC-MS spectrum and multivariate data analysis can be applied to identify essential oils from different parts of C. cassia.

Rapid simultaneous determination of codeine and morphine in plasma using LC‐ESI‐MS/MS: Application to a clinical pharmacokinetic study

A rapid and sensitive high-performance LC-MS/MS method was developed and validated for the simultaneous quantification of codeine and its metabolite morphine in human plasma using donepezil as an internal standard (IS). Following a single liquid-liquid extraction with ethyl acetate, the analytes were separated using an isocratic mobile phase on a C(18 )column and analyzed by MS/MS in the selected reaction monitoring mode using the respective [M+H](+ )ions, mass-to-charge ratio (m/z) 300/165 for codeine, m/z 286/165 for morphine and m/z 380/91 for IS. The method exhibited a linear dynamic range of 0.2-100/0.5-250 ng/mL for codeine/morphine in human plasma, respectively. The lower LOQs were 0.2 and 0.5 ng/mL for codeine and its metabolite morphine using 0.5 mL of human plasma. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.0 min for each sample made it possible to analyze more than 300 human plasma samples per day. The validated LC-MS/MS method was applied to a pharmacokinetic study in which healthy Chinese volunteers each received a single oral dose of 30 mg codeine phosphate.

A strategy for the targeted metabolomics analysis of 11 gut microbiota-host co-metabolites in rat serum, urine and feces by ultra high performance liquid chromatography–tandem mass spectrometry

Microbiota-host co-metabolites are well-known to play important physiological roles, and their dysregulation has been found to be closely related to various diseases, including but not limited to inflammatory disorders. We developed herein an original and feasible method using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The method developed enables rapid quantification of 11 key gut microbiota-host co-metabolites spanning the succinate, phenylacetylglutamine, hippurate and trimethylamine metabolic pathways within 10 min. With this method, we were able to simultaneously monitor inflammation-induced alterations of these metabolites in rat serum, urine and feces matrices. The measured levels for this panel of endogenous metabolites ranged from 0.001 to 172.8 μg m L(-1). The intra- and inter-day precision of three analytes was less than 13.1% and the accuracy was between -13.0 to 11.2% for all QC levels. The extraction recoveries in serum ranged from 85.4 to 103.2%, while the RSD was 9.0% or less for all recoveries. In addition, extraction recoveries of 11 analytes in urine and feces samples were between 85.7% and 102.0% and RSD was less than 9.5%. The method developed here has been successfully applied to the analysis of real samples from 2,4,6-trinitrobenzenesulfonic acid-induced Crohns disease in rats. All of these results suggest that the presently developed method is sufficiently sensitive and robust to simultaneously monitor co-metabolites with diverse properties and a range of different concentrations. Therefore, this method will be expected to be useful for comprehensive studies of the pathophysiological roles and mechanisms of these key microbiota-host co-metabolites, which reflect the function of the intestine, consequently offering novel opportunities for evaluating the occurrence, development and therapeutic effects of diseases related to microbiota disturbances.

Modulation of Gut Microbiome Composition and Function in Experimental Colitis Treated with Sulfasalazine

Inflammatory bowel disease (IBD) results from alterations in intestinal flora and the immune system. Sulfasalazine (SASP) is a sulfa antimicrobial used to treat IBD in clinic for years. However, how SASP affects gut microbes and its potential functions remains unclear. To investigate the relationships of SASP, IBD, and gut microbiome, we used 2,4,6-trinitrobenzenesulfonic acid (TNBS) to induce experimental colitis in rats, and analyzed the microbiota in the fecal samples, which come from the control group (treated with ethanol + saline), the model group (treated with TNBS-ethanol + saline) and the SASP group (treated with TNBS-ethanol + SASP), with 16S gene sequencing and followed up a subset sample using shotgun sequencing. The study found that SASP treatment could not only restore the TNBS-induced gut dysbiosis, which was proved by the increasing amount of SCFAs-producing bacteria and lactic acid-producing bacteria as well as the decreasing amount of Proteobacteria, but also modulate the dysregulated function of the TNBS-induced colitis to resemble that of the control group, including an increased capacity for basic metabolism (carbohydrate metabolism, citrate cycle) and a decrease in the oxidative stress (riboflavin, sulfur, cysteine) as well as bacterial pathogenesis (cell motility and secretion, bacterial motility proteins, flagellar assembly). Moreover, a higher proportion of Mycoplasma was observed in the SASP group, which may associate with infertility. In all, the study provides insight into specific microbial clades and pathways linked with SASP treatment to elaborate the mechanism for treatment of IBD.

Simultaneous determination of atropine, scopolamine, and anisodamine from Hyoscyamus niger L. in rat plasma by high‐performance liquid chromatography with tandem mass spectrometry and its application to a pharmacokinetics study

In order to investigate the pharmacokinetics of tropane alkaloids in Hyoscyamus niger L., a sensitive and specific high-performance liquid chromatography with tandem mass spectrometry method for the simultaneous determination of atropine, scopolamine, and anisodamine in rat plasma is developed and fully validated, using homatropine as an internal standard. The separation of the four compounds was carried out on a BDS Hypersil C18 column using a mobile phase consisting of acetonitrile and water (containing 10 mmol ammonium acetate). Calibration curves were linear from 0.2 to 40 ng/mL for atropine, scopolamine, and from 0.08 to 20 ng/mL for anisodamine. The precision of three analytes was <5.89% and the accuracy was between -1.04 to 2.94%. This method is successfully applied to rat pharmacokinetics analysis of the three tropane alkaloids after oral administration of H. niger extract. The maximum concentration of these three tropane alkaloids was reached within 15 min, and the maximum concentrations were 31.36 ± 7.35 ng/mL for atropine, 49.94 ± 2.67 ng/mL for scopolamine, and 2.83 ± 1.49 ng/mL for anisodamine. The pharmacokinetic parameters revealed areas under the curve of 22.76 ± 5.80, 16.80 ± 3.08, and 4.31 ± 1.21 ng/h mL and mean residence times of 2.08 ± 0.55, 1.19 ± 0.45, and 3.28 ± 0.78 h for atropine, scopolamine, and anisodamine, respectively.

Simultaneous determination of borneol and its metabolite in rat plasma by GC–MS and its application to pharmacokinetic study

A gas chromatography mass spectrometry (GC–MS) method has been developed and fully validated for the simultaneous determination of natural borneol (NB) and its metabolite, camphor, in rat plasma. Following a single liquid–liquid extraction, the analytes were separated using an HP-5MS capillary column (0.25 mm×30 m×0.25 μm) and analyzed by MS in the selected ion monitoring mode. Selected ion monitor (m/z) of borneol, camphor and internal standard was 95, 95 and 128, respectively. Linearity, accuracy, precision and extraction recovery of the analytes were all satisfactory. The method was successfully applied to pharmacokinetic studies of NB after oral administration to Wistar rats.

Simultaneous determination of three phenylethanoid glycosides from Callicarpae Caulis et Folium in rat plasma by LC-MS/MS and its application to PK study

BACKGROUND Callicarpae Caulis et Folium (CCF) is a traditional Chinese medicine usually used for hemostasis in clinics. In this study, a novel LC-MS/MS method was developed and validated for the simultaneous quantification of three phenylethanoid glycosides in rat plasma (verbascoside, forsythoside B and poliumoside), which are the major bioactive compounds of CCF; MS was operated in negative mode. RESULTS This method was linear between 5.2 and 1010 ng/ml for poliumoside, 7.0 and 420 ng/ml for forsythoside B and 2.60 and 260.0 ng/ml for verbascoside. The MS/MS ion transitions monitored were m/z 769.4→160.5, m/z 755.3→593.3, m/z 623.1→160.5 and m/z 179.0→133.6 for poliumoside, forsythoside B, verbascoside and caffeic acid (IS), respectively. Linearity, accuracy, precision and extraction recovery of three analytes were all satisfactory. CONCLUSION The method developed was sensitive, specific and rapid. It has been successfully applied in a PK study of three phenylethanoid glycosides after a single oral administration of CCF extract to rats.