Analysis of tumour-localizing haematoporphyrin derivative by high-performance liquid chromatography and fast-atom bombardment mass spectrometry.

Reversed-phase chromatography using a MOS-Hypersil (C8) column with methanol-1 M ammonium acetate buffer (pH 4.6) (60:40) as mobile phase has been developed for the isolation of tumour-localizing haematoporphyrin derivative (HPD). The system effectively resolved the diastereoisomers of haematoporphyrin and its acetyl derivatives. The chromatography peaks were identified by fast-atom bombardment mass spectrometry and were confirmed by chemical synthesis. The main components of HPD before alkaline hydrolysis were diacetylhaematoporphyrin, 8-(1-acetoxyethyl)haematoporphyrin and 3-(1-acetoxyethyl)haematoporphyrin with small amounts of haematoporphyrin, 8-(1-hydroxyethyl)-3-vinyldeuteroporphyrin, 3-(1-hydroxyethyl)-8-vinyldeuteroporphyrin, 8-(1-acetoxyethyl)-3-vinyldeuteroporphyrin, 3-(1-acetoxyethyl)-8-vinyldeuteroporphyrin and protoporphyrin. After hydrolysis with 0.1 M sodium hydroxide, the main components were haematoporphyrin, hydroxyethylvinyldeuteroporphyrins, and protoporphyrin.