UPLC–ESI-Q-TOF/MS based metabolic profiling of protosappanin B in rat plasma, bile, feces, urine and intestinal bacteria samples

Background: Caesalpinia sappan L. is a traditional medicinal plant that is used to promote blood circulation and treat stroke in China. Protosappanin B (PTB) is a unique homoisoflavone compound isolated from Sappan Lignum (the heartwood of Caesalpinia sappan L). In a previous study, the metabolic fate of PTB remained unknown. Objective: To explore whether PTB is extensively metabolized, the metabolites of PTB in bile, plasma, urine, feces, and intestinal bacteria samples in rats were investigated. Method: The biosamples were investigated by ultraperformance liquid chromatography combined with time-of-flight mass spectrometry (UPLC-TOF-MS/MS) with MetabolitePilot software. Result: 28 metabolites were identified in the biosamples: 18 metabolites in rat bile, 8 in plasma, 20 in feces, 7 in urine and 2 in intestinal bacteria samples. Both phase I and phase II metabolites were observed. Metabolite conversion occurred via 9 proposed pathways: sulfate conjugation, glucuronide conjugation, bis-glucuronide conjugation, glucose conjugation, dehydration, oxidation, hydrolysis, methylation and hydroxymethylene loss. The metabolic pathways differed among biosamples and exhibited different distributions. Among these pathways, the most important were sulfate and glucuronide conjugation. Conclusion: The results showed that the small intestinal and biliary routes play an important role in the clearance and excretion of PTB. The main sites of metabolism in the PTB chemical structure were the phenolic hydroxyl and the side-chains on the eight-element ring.

Simultaneous detection of phillyrin and its three metabolites in rat bile, excrement and urine via HPLC-ESI-MS/MS quantitative method

Introduction: Phillyrin, the main pharmacological component of Forsythia suspensa, exhibits a wide variability of therapeutic activities, such as anti-oxidative stress, free radical-clearing, antibacterial activity, hepatic protection, restoration of endothelial glycocalyx damage, prevention of bone loss, attenuation of inflammatory responses, and so on. Previous research has found that phillyrin is not easily absorbed by the body and is rarely excreted into bile, excrement and urine, suggesting that phillyrin circulates primarily in the form of metabolites. Materials and Methods: In the present study, HPLC-ESI-MS/MS method was used for the simultaneous detection of phillyrin and its three metabolites in rat bile, excrement and urine samples. Liquid-liquid extraction with ethyl acetate was carried out for the pretreatment of bile and urine samples, while excrement samples were subjected to ultrasonic pretreatment with acetone. Chromatographic separation was performed on a C18 column with gradient elution. A tandem mass system coupled with a TurboIonSpray interface operating in simultaneous positive and negative ion multiple reaction monitoring modes was employed for the simultaneous detection of the analytes. Results: The proposed method demonstrated excellent accuracy and repeatability. Conclusion: This method was successfully applied for the pharmacokinetic evaluation of phillyrin and its three metabolites simultaneously.

Pharmacokinetics and Intestinal Metabolism of Compound K in Rats and Mice

We aimed to investigate the plasma concentration, tissue distribution, and elimination of compound K following the intravenous administration of compound K (2 mg/kg) in rats and mice. The plasma concentrations of compound K in mice were much higher (about five-fold) than those in rats. In both rats and mice, compound K was mainly distributed in the liver and underwent biliary excretion. There was 28.4% fecal recovery of compound K in mice and 13.8% in rats, whereas its renal recovery was less than 0.1% in both rats and mice. Relative quantification of compound K and its metabolite protopanaxadiol (PPD) in rat bile and intestinal feces indicated that the metabolism from compound K into PPD occurred in the intestine but not in the plasma. Therefore, PPD detected in the plasma samples could have been absorbed from the intestine after metabolism in control rats, while PPD could not be detected in the plasma samples from bile duct cannulated rats. In conclusion, mice and rats shared common features such as exclusive liver distribution, major excretion pathway via biliary route, and intestinal metabolism to PPD. However, there were significant differences between rats and mice in the plasma concentrations of compound K and the fecal recovery of compound K and PPD.

Metabolism Analysis of Alantolactone and Isoalantolactone in Rats by Oral Administration

Alantolactone and isoalantolactone are the major active ingredients of Inulae Radix. Their metabolism in vivo and in vitro was investigated by UPLC-Q-TOF-MS for the first time. As a result, nine metabolites in vivo including cysteine conjugates, oxidates, dehydrogenates, and hydrates were detected in rat bile after oral administration. The metabolites produced in vitro by incubation with rat liver microsomes were found to be substantially identical to those detected in vivo. However, no metabolites were detected in the samples of plasma, feces, and urine or in the incubates of gastric juice, intestinal juice, and intestinal bacteria. These results reveal that the liver is the main metabolic organ for alantolactone and isoalantolactone, and the first pass effect of the liver appears to be the reason for the low oral bioavailability of the two lactones.