Development of a sensitive bioanalytical method for determination of PNU-83757 in rat, monkey and human plasma: from LC-UV to LC-MS/MS.
To support pre-clinical pharmacokinetic/toxicokinetic (PK/TK) evaluation, a sensitive bioanalytical method for determination of N-cyano-N'-(tert-pentyl)-N"-(3-pyridinyl) guanidine (PNU-83757), in rat and monkey plasma was required. Although the UV response of PNU-83757 was quite decent and the extracts using solid phase extraction (SPE) were very selective and concentrated, the best limit of quantitation (LOQ) achieved was 0.4 ng ml(-1) using 0.5 ml plasma for extraction and 2 ng ml(-1) using 0.1 ml plasma for extraction, which was insufficient for PK/TK evaluation at lower doses. When using liquid chromatography with atmospheric pressure chemical ionization tandem mass spectrometric detection (LC-APCI-MS/MS, positive ions) and SPE, a LOQ of 0.045 ng ml(-1) for PNU-83757 was reached. Quantitation was accomplished using the precursor --> product ion combinations of m/z 232 --> 162 for PNU-83757 and m/z 236 --> 166 for the internal standard, [2H(4)]PNU-83757, in the multiple reaction monitoring mode. This method has been successfully utilized for PK/TK evaluation in pre-clinical studies and proved to have sufficient sensitivity to determine plasma concentrations for a dose level as low as 1 microg kg(-1) day(-1) in the rat and monkey. Further improvement of this method by using electrospray mass spectrometric detection (LC-ESI-MS/MS, positive ions) and automated membrane SPE, gave an LOQ of 0.008 ng ml(-1), and allowed analysis of large numbers of samples to support clinical PK studies in microg dose levels.
D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D019791:Guanidine; D000882:Haplorhini; D006801:Humans; D051381:Rats; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet / Q000379:methods; D014665:Vasodilator Agents / Q000097:blood* / Q000737:chemistry
C456499:N-cyano-N'-(tert-pentyl)-N''-(3-pyridinyl) guanidine; D014665:Vasodilator Agents; D019791:Guanidine