Determination of 5,10,15,20-tetra-(m-hydroxyphenyl)chlorin in human plasma by high performance liquid chromatography

A high performance liquid chromatographic method for the determination of the photodynamic chemotherapeutic agent 5,10,15,20-tetra(m-hydroxyphenyl)chlorin (m-THPC) in human plasma following intravenous infusion is described. The procedure involves extraction of the drug in plasma with methanol/dimethyl sulphoxide (4:1 v/v) containing 5,10,15,20-tetra(p-hydroxyphenyl)chlorin as the internal standard and separation on a C18 reversed phase column with acetonitrile:0.1% trifluoroacetic acid (77:23 v/v) as the mobile phase. The drug was detected specifically and sensitively at its absorption maximum of 423 nm with a detection limit of 15 ng/mL (signal-to-noise ratio of 5). The intra- and inter-assay coefficients of variation (CV) on analysis of a plasma spiked with m-THPC (1 micrograms/mL) were 2.3 and 3.4% (n = 6), respectively.