Ferrochelatase activity in human lymphocytes, as quantified by a new high-performance liquid-chromatographic method

We describe a new rapid, sensitive high-performance liquid-chromatographic (HPLC) method for determining ferrochelatase (EC 4.99.1.1) activity in lymphocytes. Zinc and mesoporphyrin are incubated aerobically with sonicated lymphocytes and the zinc mesoporphyrin formed is extracted with dimethyl sulfoxide-methanol-EDTA for quantification by HPLC. Incubation conditions, including the concentration of the palmitic acid activator, were optimized. The Michaelis constant (Km) was 2.1 mumol/L for mesoporphyrin, 22.2 mumol/L for zinc. The mean ferrochelatase activity (expressed as zinc mesoporphyrin formed per hour per milligram of lymphocyte protein) for a reference population was 3.25 (SD 0.43) nmol.h-1.mg-1. For three patients with erythrohepatic protoporphyria (EHP), activities were 1.11, 1.30, and 1.35. Neutrophils contain negligible ferrochelatase activity.