Pharmacokinetics and excretion study of bergenin and its phase II metabolite in rats by liquid chromatography tandem mass spectrometry.

A sensitive and selective liquid chromatographic-tandem mass spectrometric (LC-MS/MS) method for the determination of bergenin and its phase II metabolite in rat plasma, bile and urine has been developed. Biological samples were pretreated with protein precipitation extraction procedure and enzymatic hydrolysis method was used for converting glucuronide metabolite to its free form bergenin. Detection and quantitation were performed by MS/MS using electrospray ionization and multiple reaction monitoring. Negative electrospray ionization was employed as the ionization source. Sulfamethoxazole was used as the internal standard. The separation was performed on a reverse-phase C18 (250 × 4.6 mm, 5 µm) column with gradient elution consisting of methanol and 0.5% aqueous formic acid. The concentrations of bergenin in all biological samples were in accordance with the requirements of validation of the method. After oral administration of 12 mg/kg of the prototype drug, bergenin and its glucuronide metabolite were determined in plasma, bile and urine. Bergenin in bile was completely excreted in 24 h, and the main excreted amount of bergenin was 97.67% in the first 12 h. The drug recovery in bile within 24 h was 8.97%. In urine, the main excreted amount of bergenin was 95.69% in the first 24 h, and the drug recovery within 24 h was <22.34%. Total recovery of bergenin and its glucuronide metabolite was about 52.51% (20.31% in bile within 24 h, 32.20% in urine within 48 h). The validated method was successfully applied to pharmacokinetic and excretion studies of bergenin.

A systematic data acquisition and mining strategy for chemical profiling of Aster tataricus rhizoma (Ziwan) by UHPLC-Q-TOF-MS and the corresponding anti-depressive activity screening.

In this study, a systematic data acquisition and mining strategy aimed at the traditional Chinese medicine (TCM) complex system based on ultra high-performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UHPLC-Q-TOF-MS) was reported. The workflow of this strategy is as follows: First, the high resolution mass data are acquired by both data-dependent acquisition mode (DDA) and data-independent acquisition mode (DIA). Then a global data mining that combined targeted and non-targeted compound finding is applied to analyze mass spectral data. Furthermore, some assistant tools, such as key product ions (KPIs), are employed for compound hunting and identification. The TCM Ziwan (ZW, Aster tataricus rhizoma) was used to illustrate this strategy for the first time. In this research, total 131 compounds including organic acids, peptides, terpenes, steroids, flavonoids, coumarins, anthraquinones and aldehydes were identified or tentatively characterized in ZW based on accurate mass measurements within ±5 ppm error, and 50 of them were unambiguously confirmed by comparing standard compounds. Afterwards, based on the traditional Chinese medical theory and the key determinants of firing patterns of ventral tegmental area (VTA) dopamine (DA) neurons in the development of depression, the confirmed compounds were subsequently evaluated the pharmacological effect of activity of VTA DA neurons and anti-depressive efficacy. This research provided not only a chemical profiling for further in vivo study of ZW, but also an efficient data acquisition and mining strategy to profile the chemical constituents and find new bioactive substances for other TCM complex system.

Effects of m-nisoldipine on the activity and mRNA expression of four CYP isozymes in rats.

1. For the first time, a systemic in vivo investigation was employed to evaluate the potential effects of m-nisoldipine on activities of rat cytochrome P450 enzymes (CYP1A2, CYP2C11, CYP2D1 and CYP3A1) by both cocktail probe drugs and the quantitative real-time reverse-transcription polymerase chain reaction (RT-qPCR). 2. m-Nisoldipine-treated and blank control groups were respectively administered m-nisoldipine at the dosage of 2.5, 5 and 12.5 mg/kg and CMC-Na solution for 15 days consecutively, then they were given the probe drugs of caffeine, diclofenac, dextromethorphan and midazolam (all probes were 5 mg/kg) by p.o. The blood samples were collected at different times for liquid chromatography coupled with mass spectrometry (LC-MS) analysis. The corresponding pharmacokinetic parameters were applied to evaluate the effects of m-nisoldipine on the four CYP isoforms in vivo. In addition, RT-qPCR was performed to determine the effects of m-nisoldipine on the mRNA expression of CYPs in rat liver. Results indicated that high dose and middle dose of m-nisoldipine showed significant effects on all four CYPs and CYP2C11, respectively. Moreover, for CYP2D1 and CYP1A2, there were no significant effects found at either low or middle dose of m-nisoldipine. 3. This study could provide not only experimental evidence for potential clinical application of m-nisoldipine but also a practical strategy for assessing CYP-mediated drug-drug interactions.

UHPLC-Q-TOF-MS/MS-based screening and characterization of metabolites of cnidilin in human liver microsomes.

Cnidilin is an active natural furocoumarin ingredient originating from well-known traditional Chinese medicine Radix Angelicae Dahuricae. In the present study, an efficient approach was developed for the screening and identification of cnidilin metabolites using ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry. In this approach, an on-line data acquisition method multiple mass defect filter combined with dynamic background subtraction was developed to trace all probable metabolites. Based on this analytical strategy, a total of 24 metabolites of cnidilin were detected in human liver microsomal incubation samples and the metabolic pathways were proposed. The results indicated that oxidation was the main biotransformation route for cnidilin in human liver microsomes. In addition, the specific cytochrome P450 (CYP) enzymes involved in the metabolism of cnidilin were identified using chemical inhibition and CYP recombinant enzymes. The results showed that CYP1A2 and CYP3A4 might be the major enzymes involved in the metabolism of cnidilin in human liver microsomes. The relationship between cnidilin and the CYP450 enzymes could provide us a theoretical basis of the pharmacological mechanism.

Simultaneous Determination of Five Components in Aster tataricus by Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry.

A novel quantitative method using high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry was developed for simultaneous determination of the important active constituents including shionone, luteolin, quercetin, kaempferol and ferulic acid in Aster tataricus from different habitats. The separation was performed on a C18 column with acidified aqueous acetonitrile gradients. Quantification of the analytes was achieved by the use of a hybrid quadrupole spectrometer. Multiple reaction monitoring scanning was employed with positive and negative modes at the same time in a single run. The validated results of the method indicated that the method was simple, rapid, specific and reliable. The results demonstrated that the quantitative difference in content of five active compounds was useful not only for chemotaxonomy of numerous samples from different sources but also for the standardization and differentiation of several similar samples. It was the first time to report a UPLC-ESI-MS-MS method for determination of five components in A. tataricus extract. Simultaneous quantification of bioactive components by UPLC-ESI-MS could be a well-acceptable strategy to control the quality of A. tataricus extract comprehensively.