Polygalaceae Absorption And Metabolism Of The Active Ingredient

Polygala tenuifolia(Yuan Zhi) is one of the famous herbs to improve learning and memory in traditional Chinese medicine and has often been incorprated into many traditional compound formulas intended for the treatment of amnesia and dementia. However,due to its chemical complexity,the active ingredients of Yuan Zhi responsible for learning and memory improvement are largely unknown and uninvestigated.Therefore,in the present study,interdisciplinary technologies in chemistry,pharmacokinetics and pharmacology will be applied and aim to characterise and elucidate the chemical ingredients and metabolites in rat plasma after Yuan Zhi extract is intragastric administered,and to systemically investigate the effects of the identified compounds on learning and memory improvement in vitro. Our working hypothesis is that effects of oral extracts is elicited only by the ingredients and metabolites present in the plasma,and the identified pharmacologically active compounds may be useful as chemical markers for quality control of Yuan Zhi,together as lead compounds for the development of novel therapeutics.Firstly,chemical ingredients and metabolites in the rat plasma after the Yuan Zhi extract is orally administered are to be identified using HPLC,LC-MS techniques based on our previous phytochemical data,which have determined the mass fragment profiles of various chemicals in the Yuan Zhi extract.Following identification,these bio-available components will be isolated and purified using various chromatographic techniques,and subjected to in vitro screening assay. Neuronprotective effects of bioavailable components should be evaluated by in vitro cell models using hydrogen peroxide,Aβ_25-35 and glutamic acid-induced SH-SY5Y cells.3,6'-disinapoylsucrose(DSS) was demonstrated to be the bioavailble active components of Yuanzhi.The developed LC-MS/MS method has been applied to a pharmacokinetic study for DSS in rats.The results indicated that DSS was absorbed rapidly into the body with peak time averaged 30 min after i.g.administration.However,its plasma concentration levels were very low with peak concentration averaged 16.22 ng/mL even given with the relative high dose of 50 mg/kg.The data from i.v.administration shows that DSS was eliminated rapidly from the body with half-life approximate 27 min.The absolute bioavailability of DSS after i.g.administration was found only 0.5%.After i.v.administration to rats,DSS was mainly excreted from bile 36.69%in 3 hours.Therefore,its fate in gastric intestinal tract serve as further investigation. Stability of DSS by peptidases was investigated by incubation of the drug with the contents of stomach,small intestine and colon at 37℃.Results from the experiments show the content of DSS decreased 50%at four hours in colon juice.These results indicate that the stability of DSS in GI can be affected by the Microorganism and enzyme in colon.In the caco-2 cell model,the absorption transport of DSS was fairly poor with Papp values of 1.1×10^-7 cm/s.Kinetic permeability parameters were provided by bidirectional Caco-2 experiment and pH and concentracion dependency measurements.The transport route and putative transporters involved in DSS were studied using EDTA-2Na and several inhibitors.The bidirectional studies were performed under non-gradient conditions at a concentration of 5～33μM.Opening up the paracellular tight junctions by EDTA-2Na and sodium caprate did increase the apparent permeability coefficients(Papp) of DSS.Transport of DSS was not reduced by others inhibitions.In the study of DSS in situ rat intestinal perfusion model,the Papp(blood) of DSS was 3.97×10^-5 cm/s for perfusing of DSS,which did significantly differ from that obtained for perfusion of DSS with EDTA-2Na (2.03×10^-4 cm/s) and sodium caprate(1.46×10^-4 cm/s).So,transport route of DSS was the passive paracellular.Finally,LC-MS was used to determine DSS and its metabolites in rat plasma and bile samples.DSS and three and six metabilites were detected and identified in biles collected after i.v.and oral administration by using MRM and IDA model.