Study On Active Compound To Restrain Agglutination Of Blood Platelet In Angelica Injection And Immobilization Of Glutathione S-transferases

It has been reported that ferulic acid is an important active compound in Angelica Injection. Ferulic acid can restrain agglutination of blood platelet induced by ADP whether in vitro or vivo. We use ethyl acetate extract Angelica injection at different pH and get two components. Restrain agglutination of blood platelet in vitro and high performance liquid chromatograph indicate that the component without ferulic acid also can restrain agglutination of blood platelet. Ethyl acetate extract of Angelica injection was analyzed with GC/MS, the gas chromatographic conditions was advanced as follows: All tech AT^TM-5 (30 m×0.2mm×0.2μm) column, high purity helium was used as carries gas with flow rate at lmL.min^-1;Injector tempertature:220 ℃; Column temperature:70℃ keeping for 1 min, from 70℃ to 200℃(10℃.min^-1), keeping for 1 min, then from 200℃ to 270℃( 20℃.min^-1) keeping for 5 min; MS detected with total ion monitoring. The MS of retention time of 8.06min on GC is matched with referenced standard HMF. It was demonstrated that HMF existed in Angelica Injection through GC/MS and HPLC methods with reference standard HMF. The condition of HPLC as follows: Agilent Zorbax C₁8 column(150mm×2.1mm×5μm),the mobile phase: methanol(B), l%acetic acid(A),10%B90%A-90%A10%B (25min) ,flow rate:1mL.min^-1, detective wave length:320nm, column temperature: 30℃ and the HPLC method for quantitation of HMF was established.Glutathione S transferase is one of the most important enzymes for detoxifcation and play important role in drug metabolization. We use E. coli BL21(DE3) to express GST of Schistosoma japonicum, and the crude extract of GST was immobilized on Chitosan and cross-linked with gluaraldehyde. Optimal pH, temperature, affinity to substrate l-chloro-2,4-dinnitrobenzen(CDNB) and glutathione(GSH) and thermal stability of immobilized GST were investigated and compared with the free enzyme. The activity recovery of immobilized GST reached 41.6%, optimal pH 6.5⁷.0, optimal temperature 37℃ and the thermal stability of immobilized GST wasenhanced. Immobilized GST showed high efficacy for detoxification in vitro.