Study On Quality Standard Of Xin Wei Nai An(XWNA) Tablet

ObjectiveThe prescription of Wei Nai An(WNA)capsules is composed of Astragalus,panax notoginseng,Red ginseng,Pearl layer powder,Calculus bovis factitius.Guided by the result of the pharmacological experiments,the XWNA tablet was made,which was kept original prescription of WNA the same,engaged modem techniques such as the macroporous resin and membrane separation.Mix water extraction was used to the process with Astragalus,panax notoginseng and Red ginseng.And the Pearl layer powder and Calculus bovis factitius were micronized.In this study,XWNA tablet's quality standards and stability were studied,with the new method of modem quality control,which was the fundamental research for XWNA tablet's quality control.Methods1.IdentificationMicroscopic identification was used to identify the Pearl layer powder.And TLC method was adopted to simultaneously identify four isoflavones of Astragalus including formononetin,calycosin,ononin and calycosin-7-O-?-D-glucopyranoside in a thin-layer plate,and four saponins including astragaloside,ginsenoside Rg1,ginsenoside Rb1 and notoginsenoside R1 from Astragalus,panax notoginseng and Red ginseng in a thin-layer plate,and the cholic acid and hyodesoxycholic acid from Calculus bovis factitius in a thin-layer plate.In addition,the active ingredients of XWNA tablet can scavenge DPPH free radicals were selected by TLC-bioautography.2.Quality inspectionThe weight variation,disintegration time,hardness,microbial limit and water content were checked by the relevant provisions of tablet in Chinese Pharmacopeia 2010.3.AssayThe determination employed high performance liquid chromatography coupled with a photo diode array detector and an evaporative light scattering detector(HPLC-PDA-ELSD).Calycosin-7-O-?-D-glucopyranoside,ononin,calycosin and formononetin were detected with the wavelength of260nm by PDA,and notoginsenoside R1,ginsenoside Rg1,ginsenoside Re,ginsenoside Rb1,cholic acid,hyodesoxycholic acid were detected by ELSD.Calycosin-7-O-?-D-glucopyranoside was used as the internal reference substance.The relative correlation factors(RCF)of ononin,calycosin and formononetin to calycosin-7-O-?-D-glucopyranoside were calculated and evaluated.The contents of these four isoflavones were determined by the multi-components quantitive by one marker(QAMS)and external standard method,respectively.Rationality,feasibility of the QAMS method was verified by comparing the results obtained from the two different methods.Astragaloside was determined by HPLC-ELSD.4.Specific chromatogramHPLC-PDA-ELSD was employed to establish a specific chromatogram method for XWNA tablet.The chromatogram was indicated and classified by the negative controls of XWNA tablet,and the reference substances.5.Stability studyAccording to the quality standard,the accelerated stability(at temperature40? ± 2?,relative humidity 75%±5%)and long term stability(at temperature25?±2?,relative humidity 60%±10%)were used to evaluate the stability of XWNA tablet by the appearance,identification,inspection,astragaloside contents and specific chromatogram in twelve months.Results1.IdentificationPearl layer powder's microscopic characteristic was clear.The TLC spots showed the same colors to the reference substances in the corresponding position.And the negative control showed no interference.TLC-bioautography indicated that calycosin and calycosin-7-O-?-D-glucopyranoside have the ability for DPPH free radical scavenge.2.Quality inspectionThe weight variation,disintegration time,hardness,and microbial limit tests of 3 batches of samples were all up to the standards.And the water content were in the range of 3.1%-3.4%.3.AssayThe HPLC-PDA-ELSD analysis methods were established to determine ten components,which showed good linearity(r>0.999)in the range of the test concentration,and the average recoveries were between 94.1%-106.2%(RSD?2.9).It has been no significant difference between the calculated contents(QAMS)and determined contents(external standard method)for the four isoflavones.The linear range of the astragaloside determination was 1.539-15.39?g(r=0.9998),and the average recoverie(n=6)was 97.0%(RSD=1.5%).The medicinal materials transfer rate of astragaloside in the samples(3 batches)were 61.7%,58.2%,and 65.4%,respectively.4.Specific chromatogramThe specific chromatogram of HPLC-PDA and HPLC-ELSD were built up.The chromatogram of PDA has four common peaks,and calycosin-7-O-?-D-glucopyranoside was used as reference compound.Ginsenoside Rb,was used as reference compound,and confirmed six characteristic peaks in HPLC-ELSD chromatogram.Meanwhile,the relative retention time of characteristic peaks were confired.The RSD of relative retention time of samples were less than or equal to 0.1%.5.Stability studyIn twelve months,the accelerated stability and long term stability of the XWNA tablet comformed with the quality requirements.ConclusionThe quality standard of XWNA Tablet was built up by using multi-target ingredient determination and the finger printing analysis technologies,which could control the ten compounds from Astragalus,Red ginseng,panax notoginseng and Calculus bovis factitius.These chemicals were astragaloside,calycosin-7-O-?-D-glucopyranoside,ononin,calycosin,and formononetin,ginsenoside Rg1,ginsenoside Re,ginsenoside Rb1 and notoginsenoside R1,cholic acid and hyodesoxycholic acid.In this research,TLC-bioautography was used to screen active ingredients of XWNA tablet which have radical scavenging and antioxidant activities.And HPLC-PDA-ELSD was introduced to analyze the chemincals with or without ultraviolet absorption-ingredient.Moreover,a quantitative assay of multi-components for the determination of four Isoflavones in XWNA tablet has been established.In twelve months,XWNA tablet is stable for the quality requirements.The results showed that the quality standard of XWNA Tablet is stable,specific,can be used as the evaluation method of the products.