Study On Pharmacokinetics Of ～3H-IN In Tumor-bearing Mice's Blood And Tumor

PrefaceIndomethacin is used as a kind of NASID anti - inflammation and anti - immune medicine for a long time in clinic, Recently there are so many experience and clinical observings show that IN has the anti - tumor function, To further study the metabolizability of IN in tumor and mechanisms of its anti - tumor function, In our experiment we use ³H - IN as a tracer and give it to the C₅₇ BL/ 6 mice bearing tumor, Then we use HPLC and LSC techniques to observe the pharmacokinetics of ³H - IN ,further discuss the anti - tumor mechanisms of IN.MaterialsMale C₅₇BL/6 mice bearing lewis lung cancerThe Hydrogen — 3 Labelled indomethacinScintillation LiquidBeckmen autoradiometeLC - 10AT attached with SPD - 10A violet detective instrumentChromotogram pole(C 184.6 mm ×250 mm)MethodsAnimal groups: After number all the tumor - bearing mice, divide 66 of them into eleven groups according to the time of administrating IN. It is 0.5 h,1 h,2 h,3 h,4 h,6 h,9 h,12 h,24 h,36 h and 48 h respectively.The Liquid Scintillation Measurement: Every tumor - bearing mouse was administrated ³H - IN Liquid 0. 2ml containing 90μg IN, radioactivity 180kbq. Then treat them at the different time point above. Firstly,get the mouses blood,collect about 5 drops blood on one piece of filter paper whose weight was already known. Then measure its cpm . After killing the mouse, take its tumor tissue then weigh and homogenate each one,then detect its cpm.HPLC analysis;As the same time as LSCT, Collecting lml blood by hepa-rinized taper tube,The sample operated by means of centrifugal force at 3500r/ min for 10min,take the upper plasma and put it in the -20X1 refrigerator to be detected;The tumor tissue above was blotted with filter paper,then weighed and was put into rubbing ware to triturate, added to Nacl (1:3), centrifuged at 3500 r/min for 10 min too, take the upper liquid and put it in the - 20TI refrigerator to be detected. Preparation of IN standard curve,The measured data of concentrations of IN in blood and tumor were substituted to regression equation.All the quantitative data expressed as x ± s, compare two samples' mean with t - test, size of test P = 0.01, P < 0.05 stand for significant;P < 0. 01 stand for extremely significant, pharmacokinetics parameters were calculated by DAS verl.O software.Results3H - IN had rapid blood clearance and accumulated in tumor tissue with higher levels than in blood,At 12 h the quantity of radioactivity of tumor is significantly higher than that of blood (P <0.01) , At 6 h and 9 h P <0. 05. At 3 h tumor -to blood ratios > 1. and with the time past it reached the peak at 12 h. There are still little radioactivity in tumor tissues at 36 h or 48 h. HPLC analysis results showed that the concentration of IN in tumor tissue reached the peak of 9. 77 fig/ml at 4 h ,and then clear rapidly,3.06 jjig/ml at 6 h,l. 72 |xg/ml and 1. 30 |xg/ml at 9 hN12 h. until 36 h or 48 h the concentration of IN is hardly detected, the data of INs concentrations were processed with DAS verl. 0 software.DiscussionThe radioactive isotope trace techniques has high sensitivity , it is widely used in the study of drug s pharmacokinetics, distrubition, metabolism and excre-tion. HPLC technique appeared in the last period of 1960s is a momentous renovation. It is well applied to the study of pharmacokinetics ,drug curation and inspect of drug curative effect.There are lots of reports about INs pharmacokinetics in human's plasma or tissues and mechanisms of its anti - tumor function, but that in tumor - bearing mice's plasma and tumor tissue has not reported yet in our country or foreign countries.This experiment leaded H - IN into tumor - bearing mice and observed its potential concentrating to the tumor tissues and the pharmacokinetics of IN prototype in tumor — bearing mices plasma and tumor tissues. The Liquid Scintillation Measurement research prove that 3H - IN has higher levels of radioactivity in tumor tissues and it keeps for a long time. It proves that 3H - IN has good affinity to the tumor tissue and stays in it for a long time well fit to take anti — tumor action.HPLC results proved that IN prototype reached the peak concentration at 4h in tumor tissue,and then clear rapidly,during the time of 6 - 12 h the concentrations were descending and had extremely statistic significant compared with that in blood (P <0.01). Combined with LSC results ,we can conclude that the major part of INs anti - tumor function is not its prototype ,but probably is IN& metabolized product.ConclusionThis experiment indicated that H - IN has good affinity to the tumor tissue and successfully detected the pharmacokinetics of IN prototype. The results showed that the major active part of anti - tumor is not IN s prototype.