Quantitative Analysis Of Vam3 Concentrations In Rat Plasma And Tissue Using LC-MS/MS:Application To Pharmacokinetic And Tissue Distribution Study

Vam3 (Amurensin H), a resveratrol dimer, was isolated from ethanol extracts of Vitis amurensis Rupr. The levels of Vam3 insides most plants are low. Some mature synthetic routes for Vam3 have been achieved. The previous in vivo and in vitro studies have found that Vam3 had anti-asthmatic effects and anti-COPD effects. Chronic obstructive pulmonary disease (COPD) is characterized by progressive, partially reversible airflow obstruction associated with abnormal airway inflammation and oxidant/antioxidant imbalance, and has be predicted to become the third leading cause of mortality by 2030. Vam3 can inhibit autophagy in cigarette smoke condensate (CS) -treated human bronchial epithelial cells and CS-exposed mice lungs by preventing mitochondrial dysfunction and restoring the levels of Sirtl and FoxO3a, which are implicated in pulmonary emphysema in COPD. Some studies have shown that Vam3 was more effective than resveratrol at inhibiting autophagy induced by CS. There is no pharmacokinetic report can be found at present.A liquid chromatography mass spectrometry (LC-MS/MS) method to determine Vam3 in rat plasma and tissue was developed. The analyte and internal standard (IS), hesperetin, were extracted from plasma and tissue with ethyl acetate and chromatographed on a C18 short column with 0.1% formic acid acetonitrile:0.1% formic acid in water as mobile phase by gradient elution. Mass spectrometric detection was performed by electrospray ionization in negative ion multiple reaction monitoring modes. This method monitored the transitions m/z 451.0â†'345.0 and m/z 301.0â†'164.0 for Vam3 and IS, respectively. The calibration curve was linear over a concentration range of 1.64-1000 ng/mL for plasma and tissue homogenates. The inter-day precision (R.S.D.) and intra-day precision were less than 12.8%, while the inter-day accuracy (R.E.) and intra-day accuracy were ranged from —10.60 to 9.08% in plasma and tissue homogenates. The extraction recoveries were ranged from 52.5% to 78.2%, RSD were less than 14.7%. This rapid, simple, and sensitive method was successfully applied to investigate the pharmacokinetic and tissue distribution study of Vam3 in rats.Adopt the validated analytical method to determine the contents of Vam3 in rat plasma and tissue. The results were as follows:The pharmacokinetic parameters were calculated by DAS software. After oral administration Vam3 at 70 mg/kg, the time to peak concentration (Tmax) were observed at about 2.33 ± 0.13 h, and peak plasma concentrations (Cmax) were found at the low level for only 197.33 ±41.37 μg/L. The mean area under the plasma concentration time curve from time zero to the last measurable plasma concentration point (AUC0-t) and the mean area under the plasma concentration time curve from time zero to time infinity (AUC0-∞) values were 565.14 ± 148.20 and 569.97 ± 150.29 μg/L h, respectively. The apparent volume of distribution (â…¤) value was 269.58 ± 100.84 L/kg for oral group, suggesting that this compound could extensively distribute into organs and tissues. The result indicated Vam3 could be quickly elimination from blood circulatory system. Low oral absolute bioavailability (0.79%) of Vam3 was calculated after intravenous and oral administration.The distribution of Vam3 in the various tissues results indicated that Vam3 was widely and quickly distributed in the tissues. The maximum concentration was observed at 2.5 h in all tissues after oral administration. The highest tissue concentration of Vam3 was observed in the small intestine, followed by the liver, the kidney, lung and heart. The data indicated that small intestine might be mainly absorption sites of Vam3. Dense blood-vessel network with high blood flow may have relevance for the high distribution of Vam3 in the liver and kidney. The concertration of Vam3 in tissue decreased gradually with time. Vam3 had a relatively long terminal elimination half-life in lung tissue.Plasma and tissue samples were analyzed by LC-MS/MS to find Vam3 metabolites. The result showed the glucuronide conjugate of Vam3 was identified by comparing the ion fragmentation of product ion scan and was detected in both plasma and tissues. Therefore, we can deduce that the phase â…¡ metabolism was possible metabolic pathway of Vam3 in vivo.The developed method and the pharmacokinetic data can provide a basis for further studies on bioactivity of Vam3.