Abstract
The preparation and high-performance liquid chromatography (HPLC) separation of meso-hydroxyuroporphyrinogen I, hydroxyacetic acid uroporphyrinogen I and β-hydroxypropionic acid uroporphyrinogen I is described. meso-Hydroxyuroporphyrin I, hydroxyacetic acid uroporphyrin I and β-hydroxypropionic acid uroporphyrin I were isolated from the urine of a patient with congenital erythropoietic porphyria. The porphyrins were reduced to the corresponding porphyrinogens with 3% (w/w) Na/Hg amalgam. The hydroxy porphyrinogens were separated on a Hypersil ODS column with 4% (v/v) acetonitrile in 1 M ammonium acetate buffer, pH 5.16, containing EDTA (0.27 m M) as the mobile phase, and detected electrochemically. Reduction of meso-hydroxyuroporphyrin I and hydroxyacetic acid uroporphyrin I, followed by HPLC analysis, showed that, in addition to the expected formation of meso-hydroxyuroporphyrinogen I and hydroxyacetic uroporphyrinogen I, respectively, uroporphyrinogen I was also produced. Reduction of β-hydroxypropionic acid uroporphyrin I, however, gave β-hydroxypropionic acid uroporphyrinogen I, acrylic acid uroporphyrinogen I and uroporphyrinogen I as the products. The peaks were identified by conversion into the porphyrin methyl esters and analysed by liquid secondary-ion mass spectrometry.
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