Coproporphyrinogen III oxidase assay.

An assay for coproporphyrinogen oxidase activity is described which uses a [14C]-coproporphyrinogen substrate with product isolation by methylation, extraction, and thin layer chromatography. This method affords high sensitivity, since a high specific activity of the substrate and good reproducibility due to the incorporation of an internal standard can be obtained. The activity in rat liver and human lymphocytes was found to be 140 nmol protoporphyrin/h/g of liver and 483 pmol protoporphyrin/h/mg of lymphocyte protein, respectively.

Formation and disposition of newly synthesized heme in adult rat hepatocytes in primary culture.

Studies with the intact liver have suggested that newly synthesized heme exists transiently in a small pool before its incorporation into tissue heme proteins. The same or a closely related pool may regulate synthesis of heme and serve as the precursor of "early peak" bilirubin. To delineate this postulated pool by a direct approach, we have utilized primary cultures of adult rat hepatocytes.

Studies of porphyrin synthesis in fibroblasts of patients with congenital erythropoietic porphyria and one patient with homozygous coproporphyria.

Partial deficiencies in enzymes activity of the heme biosynthesis pathway have been demonstrated in cultured skin fibroblasts and other tissues from patients suffering from congenital erythropoietic porphyria and hereditary coproporphyria. Using a new fluorimetric method, we have assessed quantitatively porphyrin biosynthesis from added delta-aminolevulinic acid in cultured fibroblasts of two congenital erythropoietic porphyria patients and one homozygous case of hereditary corproporphyria. The results were compared with those of the patients' parents and those of normal controls.

Prenatal exclusion of congenital erythropoietic porphyria (Günther's disease) in a fetus at risk.

Amniotic fluid porphyrins, biosynthesis of porphyrins by amniotic cells, and uroporphyrinogen III cosynthetase were studied after the 17th week of a pregnancy at risk for congenital erythropoietic porphyria (CEP). Only coproporphyrin was found in amniotic fluid. A diagnosis of CEP was ruled out by the demonstration of normal cosynthetase activity; biosynthesis of porphyrins was identical, not only in the porpositus and in control amniotic cells, but also in patients with CEP and in control skin fibroblasts.

Some kinetic properties of human red cell uroporphyrinogen decarboxylase.

Several kinetic properties of uroporphyrinogen decarboxylase (uroporphyrinogen-III carboxy-lyase, EC 4.1.1.

The mitochondrial localization of coproporphyrinogen III oxidase.

The location of coproporphyrinogen III oxidase in mitochondria was studied in rat liver by using the digitonin method or hypo-osmotic media for fractionation. The enzyme was found in the intermembrane space with a fraction loosely bound to the inner membrane. This fraction was released by washing the inner-membrane-matrix complex with alkaline solutions or solutions of high ionic strength.

[Demonstration of hereditary enzyme defect in coproporphyria].

We measured lymphocytes Coproporphyrinogen III Oxidase activity in 17 subjects with hereditary coproporphyria. The mean activity was about 50% of that in lymphocytes from normal subjects. This finding suggests that decreased coproporphyrinogen III oxidase activity reflects the primary genetic defect in Hereditary Coproporphyria.

The spectrophotometric determination of uroporphyrinogen I synthetase activity.

A simple spectrophotometric method for uroporphyrinogen I synthetase in erythrocytes is described. Results obtained on intermittent acute porphyria patients and carriers are similar to the results obtained with fluorimetric methods. Reproducibility, relationship between enzyme activity and enzyme concentration, and effect of time on enzymatic activity are described.

[Acute intermittent porphyria. Detection of asymptomatic carriers of the genetic defect].

Measurement of uroporphyrinogen I synthetase activity in red blood cells indicates a 40-50 p.cent decrease in patients with intermittent acute porphyria (30 +/- 6 units) when compared to normal control subjects (50 +/- 8 units). This measurement makes relatively easy the detection of asymptomatic carriers of the genetic defect, as early as the first day of life.