Tentative Exploration On Tripterygium Wilfordii Compatibility For Attenuation In Qingluo Tongbi Decoction Based On The Intestinal Absorption Characteristics And Metabolomics

Tripterygium wilfordii Hook.f.is widely used to treat rheumatic disease in clinic.However,its toxicity involving digestive systems,urogenital,blood,cardiovascular system limit its clinical application.Qinluo Tongbi prescription(QLTB),composed of Tripterygium wilfordii(TWHF),Panax notoginseng(PG),Caulis Sinomenii(QFT),Rehmannia glutinosa and Bombyx batryticatus,is an effective formula to treat rheumatoid arthritis and found to reduce toxicity and enhance efficacy of TWHF.Preliminary studies based on the chemical substances base and pharmacokinetics study of QLTB indicated that PG and QFT were the main components related to TWHF compatibility for attenuation in QLTB and the attenuation might be concerned with the in vivo process.This paper is investigated to compare the intestinal absorption characteristics and metabolic pathways of the four major active and toxic components from TWHF between TWHF,TWHF-PG,TWHF-QFT and QLTB groups to explore the attenuation mechanism of compatibility of TWHF-PG,TWHF-QFT and in QLTB based on the intestinal absorption characteristics and metabolitic pathways in vivo.The research work was carried out from two aspects:the intestinal absorption characteristics of the four major active and toxic ingredients from TWHF and the metabolomics while TWHF was combined with PG.1.Study on chemical basis of TWHF compatibility for attenuation in QLTB:According to the preliminary study of the chemical basis,we determined the triptolide in TWHF,TWHF-PG,TWHF-QFT and QLTB decoction to investigate the regularity of triptolide dissolution under the same dose of TWHF.Results showed that the concentrations of triptolide were in the order:TWHF>TWHF-PG>TWHF-QFT>QLTB.The statistics analysis showed that PG,QFT significantly reduced the dissolving action of triptolide(P<0.05)as well as in QLTB group.2.Study on the intestinal absorption characteristics of the four major active and toxic component in TWHF,TWHF-PG,TWHF-QFT and QLTB groups.(1)Firstly,we established the LC-MS detection method for the determination the four major active and toxic component(triptolide,triptonide,wilforlide A and tripterine)in perfusion samples.Method validation results showed that the specificity,linearity,extraction recovery,intra-and inter-day precision and stability requirements were in line with the analysis of biological samples,so the method was reliable and feasible.(2)The in situ perfusion method in rats was chose to analysis the intestinal absorption characteristics of triptolide,triptonide,wilforlide A and tripterine.Results showed that the four components could be well absorbed in general intestinal tract without specific absorption site.The absorption trend in different intestinal tract of the four components were in the following trend:Triptolide:ileum>duodenum>colon>jejunum;Triptonide:duodenum>colon>ileum>jejunum;Wilforlide A:colon>ileum>jejunum>duodenum;Tripterine:ileum>colon>jejunum>duodenum.Meanwhile,tripterine had a poor absorption about 6%among the four compound while the 10 cm%ABS of triptolide,wilforlide A,triptonide were in the range of 29%to 51%.(3)A comparative intestinal absorption study of triptolide,triptonide,wilforlide A and tripterine between TWHF,TWHF-PG TWHF-QFT and QLTB groups was investigated by the means of the in situ perfusion method in rats.Results showed that the absorption characteristic of triptolide,triptonide and tripterine in TWHF,TWHF-PG,TWHF-QFT and QLTB groups were the same as that in monomer perfusion while the absorption characteristic of wilforlide A in intestinal tract changed in the TWHF-PG TWHF-QFT group but without consistency.The absorption paramaters including Peff,10cm%ABS and Ka of triptolide,triptonide,wilforlide A decreased significantly in TWHF-PG,TWHF-QFT and QLTB groups compared to TWHF group while QLTB dropped most.It indicated that the intestinal absorption of the three major toxic component from TWHF were reduced and slower.However,the absorption paramaters of tripterine tends to be decreased in the order of QLTB,TWHF-PG,TWHF and TWHF-QFT.It indicated that the intestinal absorption of tripterine increased in TWHF-PG and QLTB group while reduced in TWHF-QFT group.That is to say,the poor oral absorption of tripterine could be appropriately improved in TWHF-PG and QLTB which might be related to the toxicity reducing and efficacy enhancing when TWHF was combined with PG and in QLTB decoction.3.Metabolomics study on the TWHF-PG compatibility for attenuation(1)Study on TWHF induced liver and kidney toxicty:A comparative study on the metabolites in urine and blood of rats after 7 day continuously administration of triptolide and TWHF was conducted by the means of metabolomics as well as the histopathological examination of liver and kidney and serum liver funcion test to find the possible marker related to the TWHF induced liver and kidney toxicty.We have initially determined 21 markers(11 in urine and 10 in blood)related to the toxicty.Among them,taurine,L-methionine,acetyl citrate,L-malic acid,fumaric acid,PC(o-22:0/22:3,18:1),LPC(14:1,20:4)and phosphoric acid were associated with TWHF induced liver injury while hippuric acid,creatine and L-methionine were related to renal injury.The metabolic pathways changed by TWHF and related to toxicty probably involved in protein synthesis,the krebs cycle,urea cycle,phospholipid metabolism,energy metabolism and lipid metabolism.(2)Study on the TWHF-PG compatibility for attenuation:A comparative study on the metabolites in urine and blood of rats after 7 day continuously administration of TWHF and TWHF-PG was conducted by the means of metabolomics as well as the histopathological examination of liver and kidney and serum liver funcion test to find the possible marker related to the TWHF-PG compatibility for attenuation.We have initially determined 17 markers related to the attenuation,including taurine,L-methionine,acetyl citrate,fumaric acid,methylsuccinic acid,cis-4-octenedioic acid,L-cysteine,D-threitol,tiglylglycine and PC(o-22:0/22:3,22:0),LPC(15:0,18:1,18:2,22:5,22:6)and phosphoric acid.The metabolic pathways changed by TWHF-PG probably involved in protein synthesis,the krebs cycle,urea cycle,phospholipid metabolism,energy metabolism,lipid metabolism and carbohydrate metabolism.These metabolic pathways might be changed due to TWHF-induced liver and renal injury and they could be called back when TWHF was combined with PG.It is likely to explain the machanism related to the TWHF-PG compatibility for attenuation.