Studies On The Chemical Constituents And Content Determinaion Of Folium Isatidis

This thesis included two parts:Studies on the Chemical Constituents of Folium isatidis and Content Determination of Folium isatidis.Folium isatidis is dried the leaves of Isatis indigotica Fort.This medicinal mater ial is now included in the 2020 edition of Chinese Pharmacopoeia.It has the effects of clearing heat and detoxifying,cooling blood and eliminatingspots.It used for war m-disease with high fever,infatuation,spot with rash,mumps,laryngeal obstruction,erysipelas,carbuncle.Pharmacological studies have proved that it has antibacterial,anti-endotoxin,anti-inflammatory,anti-cancerand other pharmacological activities.I n this paper,the chemical constituents of Folium isatidis were studied.We took 18 K g of the herbs of Folium isatidis,crushed by reflux extraction with 75%ethanol,the extracts were isolated and purified by silica gel,RP-C₁₈ and dextran gel Sephadex L H-20.The structures of the compounds were identified by NMR,MS and other spect ral techniques.Results:22 compounds were isolated from Folium isatidis and identif ied as 1H-indole-3-acetonitrile（1）,tryptanthrin（2）,Rutalexin（3）,isovitexin（4）,isovit exin-3″-O-glucopy ranoside（5）,isoscoparin（6）,liguadenosine B（7）,indican（8）,conif erin（9）,syringin（10）,apigenin 7-O-β-glucoside（11）,luteolin 7-O-β-glucoside-3′-met hyl ether（12）,1,2,2′-tri-O-E-sinapoyl-β-gentiobiose（13）,1,2,6′-tri-O-E-sinapoylgenti obiose（14）,larisiresinol 4′-O-β-D-glucopyranoside（15）,larisiresinol 9-O-β-D-glucop yranoside（16）,o-aminobenzoic acid-7-O-β-D-glucopyranosyl ester（17）,indole-3-ace tonitrile-6-O-β-D-glucopyranoside（18）,adenosine（19）,rutin（20）,chrysoeriol 6-C-β-D-glucopyranosyl-（1→3）-β-glucopyranoside（21）,wrightiadione（22）.Among them,compounds 1,3,7 to 9,11 to 18,21,22 were isolated from Folium isatidis for the fir st time.To establish an HPLC method for the simultaneous determination of fourchemi cal components from Folium isatidis,including 1,2,2′-tri-O-E-sinapoyl-β-gentiobios e and 1,2,6′-tri-O-E-sinapoylgentiobiose.Methods:The Yilite C₁₈column（250×4.6mm,5μm）was used;acetonitrile（A）-0.1%phosphoric acid aqueous solution（B）wa s used as the mobile phase and gradient sequential elution;gradient elution at a flow rate of 1.0 m L.min^-1.328 nm is used as the detection wavelength;and the column te mperature was at 30℃.Results:The injection volumes of 1,2,2′-tri-O-E-sinapoyl-β-gentiobiose and 1,2,6′-tri-O-E-sinapoylgentiobiose were in the ranges of 0.0202～1.0100μg（R²=0.9998）and 0.0281～1.4050μg（R²=0.9999）showed good linearity with peak area;good precision（RSD<2%）,stability（RSD<2%）and reproducibility（RSD<2%）;The average recoveries（n=6）were 99.05%（RSD=0.34%）and 99.60%（RSD=0.65%）,respectively;the ranges of two chemical compositions in the nine batches were 0.3007～3.0863 and 0.1135～2.6766 mg.g^-1,respectively.To establish an HPLC method for the simultaneous determination of fourchemi cal components from Folium isatidis,including isovitexin,isovitexin-3″-O-glucopyr anoside,isoscoparin and chrysoeriol 6-C-β-D-glucopyranosyl-（1→3）-β-glucopyrano side.Methods:The ACE Excel 5 C₁₈-AR column（250 mm×4.6mm,5μm）was use d;acetonitrile-0.1%phosphoric acid aqueous solution（13.5:86.5）was used as them obile phase and isometric elution;The flow rate was1.0m L·min-1;the detection wav elength was 269 nm;and the column temperaturewas 30℃.Results:The injection v olumes of isovitexin,isovitexin-3″-O-glucopyranoside,isoscoparin and chrysoeriol6-C-β-D-glucopyranosyl-（1→3）-β-glucopyranoside were in the ranges of 0.0438～1.0950μg（R²=1.0000）,0.0414～1.0350μg（R²=1.0000）,0.0288～0.7200μg（R²=1.0000）and 0.0520～1.300μg（R²=1.0000）showed good linearity with peak area;good precis ion（RSD<2%）,stability（RSD<2%）and reproducibility（RSD<2%）;The average re coveries（n=6）were 98.1%（RSD=1.25%）,97.0%（RSD=0.73%）,100.3%（RSD=1.24%）and 98.5%（RSD=1.06%）,respectively;the ranges of four chemical compositio ns in the nine batches were 1.0053～2.0666、0.1338～1.8760、0.5449～1.5192 and 0.1669～2.5872 mg.g^-1,respectively.