| Doxorubicin is an antitumor drug which is widely used in the treatment with remarkable effect of leukemia,lymphoma etc. It can cause many adverse reactions,such as myelosuppressio,cardiac toxicity.Delisheng injection is composed of ginseng,radix astragali,cantharis and venenum bufonis. It is used to strengthen the treatment effect and weaken the myelosuppression induced by irradiation in patients with cancer. Both of them can be used together to achieve purpose of enhancing efficacy and reducing adverse reactions. However,the detection method of Resibufogenin in Delisheng has not yet established in serum. And the pharmacokinetics of two drug used together has not yet been reported at home and abroad. The following reports were presented for the application study of reversed phase-high performance liquid chromatographic (RP-HPLC) method for the determination of Resibufogenin and Doxorubicin in rats'serum. By the RP-HPLC method,the effect of Delisheng injection on pharmacokinetics of Doxorubicin injection was studied.Objective:To establish a RP-HPLC method for the determination of Doxorubicin injection and Delisheng injection in rats'serum, and study the pharmacokinetic interaction of them in rats.Methods:Using Daunorubi-einas internal standard(I.S),Doxorubicin in serum was determined by HPLC with liquid-liquid extraction of the compound from serum and was separated by the column of Phenomenex ODS C18(250mm×4.6mm,5μm) at 35℃. The mobile phase consisted of a mixture solution of methanol-0.01 mol·L-1 NH4H2PO4 (adjusted to PH4.65 with phosphoric acid)-acetic acid (30:20:0.1) pumped at a flow rate of 1.0 ml·min-1.A UV detection wavelength was 233nm. Using Norethindrone internal standard(I.S),Resibufogenin in serum was determined by HPLC with liquid-liquid extraction of the compound from serum and was separated by the column of Phenomenex ODS C18 (250mm×4.6mm,5μm) at 35℃. The mobile phase consisted of a mixture solution of acetonitrile-liquid (55:45) pumped at a flow rate of 1.0 ml·min-1. A UV detection wavelength was 296nm. The RP-HPLC method measuring Doxorubicin in concentration in serum was used to measure and compare the pharmacokinetic partners among two groups. Which was administrated with the Doxorubicin, Doxorubicin and Delisheng injection respectively. Blood samples were drawn respectively as follows:NO I:0.083,0.167,0.5,1,3,6,12,24h;NOⅡ:0.083,0.167,0.5,1,3,6,12,24h; (Delisheng injection given 30 min prior to Doxorubicin). Results:The drug-free serum did not interfere with the determination of Doxorubicin and I.S.There were good linear relationships (1 weighted,r2=0.9987)between peak high ratio of Doxorubicin to I.S. and C within the range of 0.206～20.6mg·L-1. The lower limit of quantineation was 3.5ng. The drug-free serum did not interfere with the determination of Resibufogenin and I.S. There were good linear relationships (1 weighted, r2=0.9997) between peak high ratio of Resibufogenin to I.S. and C within the range of 55～8800μg·L-1. The lower limit of quantineation was 1.3ng. The precision of inter-day and intra-day was less than 10%. Doxorubicin and Resibufogenin in serum was stable for at least 14 days and three freeze-thaw cycles. The fitting of concentration-time data were performed with DAS2.1.1 program.The concentration time curves of Doxorubicin injection used either alone or in combination fitted two-compartment model. The pharmacokineties parameters were calculated as followed:t1/2αwere 0.258±0.029,0.266±0.003h of the two group above respectively;t1/2βwere 30.213±5.259,35.243±2.59h;CL were 0.082±0.027,0.143±0.044L·kg-1·h-1;AUC(0-t) were 37.684±3.872,23.252±3.651mg·h·L-1; AUC(0-∞) were 81.867±29.656,46.791±18.063mg·h·L-1;V were 0.346±0.032,0.573±0.084L·kg-1; Cmax were 15.36±1.503,9.593±1.495 mg·L-1. The t1/2β,Cmax,AUC(0-t) and CL were significantly difference between two groups;The t1/2α,AUC(0-∞) and V were no significantly difference between two groups. So it is necessarily adjust the drug dosage and interval time.Conclusion:The method is simple,rapid,accurate,sensitive and reproducible.It can be applied to determine the concentrations of Doxorubicin injection and Resibufogenin of Delisheng injection in serum rapidly and can be suitable for use in studying pharmacokinetics of them. There was possible pharmacokinetic effect of Delisheng injection on Doxorubicin when Doxorubicin and Delisheng were co-administered, and the concentration of the Doxorubicin serum should be monitored when they were used in combination. But we should have more animal and clinical laboratory to get the conclusion above. |
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