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.
Rossi
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Clinical Biochemistry Department, Queen Elizabeth II Medical Centre, Nedlands, Western Australia.
Costin
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Garcia-Webb
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D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005227:Fatty Acids / Q000378:metabolism; D005294:Ferrochelatase / Q000097:blood*; D006801:Humans; D008099:Liver / Q000201:enzymology; D008107:Liver Diseases / Q000201:enzymology; D008190:Lyases / Q000097:blood*; D008214:Lymphocytes / Q000201:enzymology*; D008652:Mesoporphyrins; D009504:Neutrophils / Q000201:enzymology; D011164:Porphyrias / Q000201:enzymology; D051381:Rats; D015032:Zinc
D005227:Fatty Acids; D008652:Mesoporphyrins; C031468:mesoporphyrin IX; D008190:Lyases; D005294:Ferrochelatase; D015032:Zinc