Ultra high performance liquid chromatography method for the determination of pentamidine and analog in rat biological fluids.

Pentamidine isethionate (PTMD) is an antiprotozoal agent used in different parasitic diseases as Human African Trypanosomiasis or Pneumocystis pneumonia. Given its side effects, numerous analogs are still under development worldwide. PTMD has been recently described having a potential activity in myotonic dystrophy (type 1). Here we present an UPLC method coupled to fluo or PDA detection for PTMD and one analog determination in rat plasma or urine. The chromatographic separation was achieved on a Acquity UPLC® HSS T3 analytical column using a mobile phase combining formic acid 0.1% (v/v) and acetonitrile (ACN) at a constant flow rate of 0.4 mL/min. Preliminary, an innovative μSPE (solid phase extraction) procedure using Oasis® WCX sorbent was processed and gave satisfying and reproducible results in terms of extraction yields. Additionally, the methods were successfully validated using the accuracy profiles approach (β=95% and acceptance limits=15%) over the ranges 2.88-287.52 ng/mL and from 143.76 ng/mL to 1.72 μg/mL in rat plasma and urine for PTMD and for EBAB, from 4.23 to 423.39 ng/mL and from 211.69 ng/mL to 2.54 μg/mL for plasma and urine, respectively. The validated protocols were applied to a pharmacokinetic (PK) study on rats and permitted to point out some relevant PK parameters on PTMD and its studied analog.