Micro-HPLC and transfer techniques for directly measuring drug (indomethacin) transport across the isolated perfused renal proximal straight tubule.
We wished to study the active transport of indomethacin across isolated segments of renal proximal tubules. To do so, we developed a microbore HPLC technique capable of directly measuring pharmacologic concentrations of indomethacin in nanoliter sample volumes. The technique also required development of methods for transferring of samples, introduction of internal standard and identification of an oil, under which the sample could be collected without interference with the assay itself or the compounds of interest. We found that gamma-terpinene could be used as the oil, for it did not impede the assay, did not dissolve indomethacin or inulin, and indomethacin and inulin remained stable in the water phase. Transfer of sample was effected with a glass tube containing 640 nL of water plus internal standard into which the indomethacin sample pipette (110 nL) could be introduced. This transfer tube was then capped with PE-60 tubing, which could later be used for withdrawing sample into an injection syringe for HPLC assay. This method proved feasible for measuring indomethacin transport across isolated perfused proximal tubule segments and should be applicable for similar studies of other drugs.
de Zeeuw
D
D
University of Groningen, Netherlands.
Brater
D C
DC
Jacobson
H R
HR
D000818:Animals; D001693:Biological Transport, Active; D002851:Chromatography, High Pressure Liquid; D066298:In Vitro Techniques; D007213:Indomethacin / Q000493:pharmacokinetics*; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D011817:Rabbits; D013056:Spectrophotometry, Ultraviolet
D007213:Indomethacin