Isolation And Analysis Of Anthraquinones From Herba Hedyotis Diffusa And Its Pharmacokinetics

Hedyotis diffusa Willd.belongs to rubiaceous family,possessing activities that clear away the heat-evil,expel superficical evils,activate blood circulation to dissipate blood stasis, promote urination and clear intumesce.Now it is widely used to cure maliglant tumor, appendicitis,icteric hepatitis,enteritis,pneumonia and so on.With the development of modern pharmacology and activity study,Hedyotis diffusa Willd.increasingly attracts the attention of pharmaceutical researchers all over the world.In this paper,four kinds of components including two kinds of anthraquinones,scopoletine and ursolic acid were isolated from Hedyotis diffusa Willd.and identified by physicochemical property and spectroscopy analysis;HPLC and GC methods were developed and compared for simultaneously quantitative determination of two kinds of athraquinones in raw materials of Hedyotis diffusa Willd.from different sources;HPLC-UV method has been developed for the determination of 2-methyl-3-hydroxyanthraquinone in rat plasma and applied to a pharmacokinetic study in rats after intraperitoneal injection of 2-methyl-3-hydroxyanthraquinone.1.The preparation and identification of four kinds of chemical components in Hedyotis diffusa Willd.2-methyl-3-hydroxy-4-methoxyanthraquinone,2-methyt-3-hydroxyanthraquinone, scopoletine and ursolic acid were isolated from Hedyotis diffusa willd,by silica gel column chromatography and TLC methods.The structure elucidations were reported in the paper.2.Simultaneous HPLC and GC quantitation of two kinds of anthraquinones in Hedyotis diffusa Willd.of different sources.After the optimization of extraction condition using orthogonal experimental design based on extracting under reflux with alcohol,an HPLC method was developed.The analytical column was Diamonsil C₁₈with MeOH-H₂O（62:38,v/v）as mobile phase,at flow rate of 1.0 mL·min^-1,peaks were detected at 267 nm,column temperature was room temperature.A GC method was developed for the determination of two kinds of anthraquinones in Hedyotis diffusa from different sources for the first time.The separation was performed on the HP-5 elastic crystal Capillary column（30 m×0.32 mm×0.25μm）equipped with FID detector at split of 20:1 and the column temperature was maintained at 228℃.The carrier gas is Nitrogen and flow rate is 50 mL/min.The methods are accurate,reproducible for determination of two kinds of anthraquinones in Hedyotis diffusa Willd.Compared to HPLC,the sample treatment is easier,the analysis time is decreased in GC method.3.The pharmacokinetics of 2-methyl-3-hydroxyanthraquinone in rat plasmaA rapid and simple HPLC method has been established to determine 2-methyl-3-hydroxyanthraquinone in rat plasma.The analytes were separated on a Diamonsil C18 column（200 mm×4.6 mm,5μm）with methanol-water（78:22,v/v）as mobile phase and detected at 267 nm at room temperature.The calibration curves were linear over the concentration range 0.1～4.0μg·mL^-1.The lower limit of quantification of 2-methyl-3-hydroxyanthraquinone in rat plasma was 0.1μg·mL^-1.The internal standard was Ethinylestradiol.Following intraperitoneal injection in rats（6 mg/kg）,2-methyl-3-hydroxyanthraquinone was absorbed quickly and the individual variation is notable.The primary phamacokinetics parameters of 2-methyt-3-hydroxyanthraquinone in plasma were as follows:mean T_1/2is 59.9 min,AUC_0-tis 159μg min·mL^-1.