2,2',3',4,4'5,5'-Hepatachlorobiphenyl (PCB 180)--on its toxicokinetics, biotransformation and porphyrinogenic action in female rats.

The toxicokinetics and biotransformation of 2,2',3',4,4',5,5'-heptachlorobiphenyl, as well as its influence on the activity of microsomal and cytosolic enzymes and on the porphyrin pathway in the liver were studied in female rats following oral treatment with 7 mg/kg every other day for 3 months. One day after cessation of treatment the concentration of the compound in liver, spleen, CNS and blood was 100-500 times and in the trachea it was only 5 times less than in the adipose tissue. The daily excretion with the feces and urine amounted to 35 and 1.5 micrograms, respectively. In both excreta, heptachlorobiphenylol was identified as a metabolite. The biotransformation rate was estimated to be about 5%. Investigations of the liver revealed increases in the relative liver weight, total cytochrome P-450 content, O-deethylation of 7-ethoxycoumarin and in the activity of glutathione S-transferases. Disturbances of the hepatic porphyrin pathway were not detected. Only at the end of a post-dosing period of 12 months did the hepatic uroporphyrinogen decarboxylase show diminished activity. Only one of these animals with diminished enzyme activity showed drastically elevated porphyrins. In these animals, the fecal and urinary porphyrins did not differ from controls. At no time did heptachlorobiphenyl influence the urinary excretion of delta-aminolevulinic acid and porphobilinogen. The results indicate 1) that this congener shows expected toxicokinetics with the exception of being accumulated in the trachea and 2) that this congener induces disturbances of the hepatic porphyrin pathway several months after cessation of treatment.

Pteridine modulation of lead inhibition of uroporphyrinogen synthesis in erythroid precursor cells.

The role of nutritional factors on heme synthesis and their influence on the development of anemia in the bone marrow during lead exposure is unclear. Previous reports suggested that pteridines could regulate the formation of uroporphyrinogen, a step midway along the heme synthetic pathway. Studies were undertaken to determine if pteridines could modulate lead inhibition of uroporphyrinogen synthesis in erythroid precursor cells. Pteroylpolyglutamates of various glutamate chain lengths were tested for the ability to protect against lead inhibition of uroporphyrinogen I (URO) synthase prepared from murine erythroleukemia cells (MELC). Pteroylpentaglutamate, the major endogenous polyglutamate form by chain length found to be present in MELC, afforded rapid and specific protection of URO synthase against lead inhibition. MELC are expected to be a useful in vitro model for studying the role of endogenous folates on uroporphyrinogen synthesis and heme formation in erythroid precursor cells following lead exposure.

[Definition and classification of porphyrias]. / Definition und Klassifikation der Porphyrien.

According to the newest recognitions a disease entity is to be ascribed to every deficient enzyme of the chain of haemobiosynthesis, so that the subdivisions of the porphyrin diseases, having often changed since several decades, may be substituted by classifications on ascertained enzymatic basis. An establishment of the well-known porphyrias is given, taking into consideration the first describers, the hereditary factors and first individual of homozygous diseases.

Reversal of sulfamerazine inhibition of rat hepatic uroporphyrinogen synthesis by folic acid.

The ability of rat hepatic uroporphyrinogen cosynthase to direct formation of uroporphyrinogen III and the synthesis of uroporphyrinogen in vitro was impaired by sulfamerazine. Inhibition was reversed by the addition of folic acid. Administration of a single, oral dose (1 g/kg) of sulfamerazine to rats was associated with elevated levels of hepatic uroporphyrin I isomer. These results suggest that sulfonamides may interfere with the biosynthesis of uroporphyrinogen III.

Inhibition of the biosynthesis of uroporphyrinogen and heme in rat liver during obstructive jaundice produced by bile duct ligation.

Altered hepatic microsomal drug metabolism has been reported to occur in afflicted with hyperbilirubinemia. Similarities of the chemical structures of hydroxymethylbilane, an intermediate in the biosynthesis of uroporphyrinogen, to bilirubin prompted investigations of the effect of bilirubin on the activity of uroporphyrinogen I synthase (porphobilinogen deaminase, EC 4.3.1.8) and the biosynthesis of heme. Bilirubin was found to be a reversible, noncompetitive inhibitor of uroporphyrinogen I synthase. The inhibition constant (Ki) for bilirubin was 1.5 microM. Bile acids had no effect on rat hepatic uroporphyrinogen I synthase activity. Hyperbilirubinemia was achieved in rats by biliary ligation in order to investigate whether elevated levels of bilirubin impair the biosynthesis of hepatic heme in vivo. The relative rate of heme biosynthesis, as measured by the rate of incorporation of delta-[4-14C]aminolevulinic acid into heme, was decreased 59% 24 h after biliary obstruction. The levels of hepatic microsomal heme and cytochrome P-450 were decreased by 43 and 40%, respectively, 72 h after biliary obstruction. The activities of hepatic delta-aminolevulinic acid synthase and uroporphyrinogen I synthase were increased by 39 and 46%, respectively, 72 h after biliary obstruction. During the 48- to 72-h period following biliary obstruction, the urinary excretion of porphobilinogen and uroporphyrin was increased 3.0- and 3.5-fold, respectively, whereas, the urinary excretion of delta-aminolevulinic acid was not altered. During this 48-to 72-h time interval following biliary obstruction, 100% of the uroporphyrin was excreted as isomer I. These results indicate that bilirubin is capable of depressing the biosynthesis of rat hepatic heme and thus cytochrome P-450-mediated drug metabolism by inhibition of the formation of uroporphyrinogen. These findings are a plausible mechanism for reports of impaired clearance of various drugs in patients afflicted with hyperbilirubinemic disease states.

Erythrocyte uroporphyrinogen synthase activity as a possible diagnostic aid in the diagnosis of lymphoproliferative diseases.

Patients with active lymphoproliferative diseases were shown to have high activity of erythrocyte uroporphyrinogen synthetase (URO-S), the enzyme which converts porphobilinogen to uroporphyrinogen. In a few patients examined the lymphocyte URO-S was markedly increased. No correlation was found between the high URO-S activity and the degree of anemia, reticulocytosis, or the presence of hemolysis. Patients with epithelial malignancies and with some common viral diseases had normal erythrocyte URO-S values. Three patients with nonalcoholic cirrhosis also had high erythrocyte URO-S activities. The determination of erythrocyte and lymphocyte URO-S activity may be of aid in the diagnosis of lymphoproliferative diseases. It may also indicate whether remission has been achieved and whether treatment should be continued or reinstituted. These preliminary observations justify the investigation of a larger patient and control material.

Toxicity of PBBs with special reference to porphyrinogenic action and spectral interaction with hepatic cytochrome P-450.

Some of the polyhalogenated aromatic compounds (PHAs) which are able to produce porphyria are presently known as environmental contaminants. Chronic exposure to PHAs causes hepatic porphyria in different species. Qualitatively PBBs act comparable to PHAs. An increase in accumulation of porphyrins caused by PHAs is not simply related to an increase of delta-ALAS activity in liver. Heme cannot exert a feedback when porphyria develops. Induction of P-450 mediated drug enzymes is needed. The PHAs interact with P-450 in vitro. The PHAs are converted into a reactive intermediate, not a known metabolite, which depletes liver GSH and then becomes reactive to tissue structures. Mitochondria are damaged; fluorescence of porphyrins is detected in the region of central veins where degenerative change in hepatocytes is most marked. A possible pathological change in the cell membrane permeability is assumed too. In this porphyric stage uropophyrinogen decarboxylase (urogen decarboxylase) is inhibited. The proportion of steroid hormones product by ovaries and testes compared to each other are possibly involved in sensitivity to porphyrinogenic compounds.

Biosynthesis of uroporphyrinogens. Interaction among 2-aminomethyltripyrranes and the enzymatic system.

Many hypotheses on uroporphyrinogen biosynthesis advanced the possibility that 2-aminomethyltripyrranes formed by porphobilinogen deaminase are further substrates or uroporphyrinogen III co-synthase in the presence of porphobilinogen. These proposals were put to test by employing synthetic 2-aminomethyltripyrranes formally derived from porphobilinogen. None of them was found to be by itself a substrate of deaminase or of co-synthase in the presence of porphobilinogen. The tripyrranes chemically formed uroporphyrinogens by dimerization reactions, and the latter had to be deducted in control runs during the enzymatic studies. Two of the tripyrranes examined, the 2-aminomethyltripyrrane 7 and the 2-aminomethyltripyrrane 8, were found to be incorporated into enzymatically formed uroporphyrinogen III in the presence of porphobilinogen and of the deaminase-co-synthase system. While the former gave only a slight incorporation, the latter was incorporated in about 16%. No incorporation of 8 into uroporphyrinogen I was detected. On the basis of these results, and of the previous results obtained with 2-aminomethyldipyrrylmethanes, an outline of the most likely pathway of uroporphyrinogen III biosynthesis from porphobilinogen is given.

The discovery of nature's biosynthetic pathways.

Uro'gen-III is a key intermediate on the biosynthetic pathways to the vitally important natural pigments haem, chlorophyll and the cytochromes. How the unexpected structure of uro'gen-III is synthesized by living things has long been a major puzzle. Studies based on 13C-labelling are described which show a) that a single intramolecular rearrangement occurs and b) that this step occurs after the open-chain linear tetrapyrrole system has been built. A second study involves stereospecific labelling with deuterium and tritium to elucidate the absolute stereochemistry of the enzymic reaction sequence which produces the vinyl groups of haem. The third and last section of the lecture is focussed on the biosynthetic intermediates lying between uro'gen-III and cobyrinic acid on the pathway to vitamin B12. An octacarboxylic isobacteriochlorin is isolated from a vitamin B12-producing organism and this is shown to be identical with sirohydrochlorin, previously obtained by Kamin and Siegel as the metal-free prosthetic group of certain sulphite-reducing bacteria. The structure and absolute stereochemistry of sirohydrochlorin are studied and comment is made on the evolutionary interest of these findings.

Studies on porphyrin biosynthesis in lead-intoxicated rabbits.

The present report deals with studies on porphyrins and porphyrinogen carboxy-lyase of red cells and urinary porphyrins from lead-intoxicated rabbits. It was shown that the free erythrocyte porphyrins are mixture of protoporphyrin 9, the main component, and minor proportions of Coproporphyrin, Uroporphyrin III and Phyriaporphyrin. Analysis of the urinary porphyrins deomonstrates the presence of Coproporphyrins III as the major component, together with 15-20% of other porphyrins: 10-14% 5-COOH, 1-2% 6-COOH, 2-3% 7-COOH porphyrin and 1-2% Uroporphyrin III. We have not been able to detect an increase of Uroporphyrin I. Assays of porphyrinogen carboxy-lyase activity in hemolysate supernatant using Uroporphyrinogen III and Phyriaporphyrinogen (Phyria'gen) III as substrates, showed the existence of a slight decrease of both decarboxylase activities, being more affected during the second stage, the Phyria'gen decarboxylation. A possible regulation mechanism responsible for the porphyrin picture is discussed.

Biosynthesis of uroporphyrinogens from porphobilinogen: mechanism and the nature of the process.

The enzymic self-polymerization of prophobilinogen gives rise to the cyclic tetrapyrroles uroporphyrinogen III and uroporphyrinogen I. The former is the precursor of all the natural porphyrins and chlorins. The formation of uroporphyrinogen III is catalysed by a dual enzymic system, porphobilinogen deaminase and uroporphyrinogen III cosynthase. Deaminase polymerizes four porphobilinogen units on the enzymic surface, without liberation of free intermediates into the reaction medium, and forms uroporphyrinogen I. Cosynthase enters into association with the deaminase, and acts as a 'specifier protein' of the latter, changing the mode of porphobilinogen condensation on the enzymic surface. The association is independent of the presence of substrate. While deaminase catalyses the head-to-tail condensation of the porphobilinogen units, the association deaminase-cosynthase catalyses the head-to-head condensation of the same units. As a result different enzyme-bound dipyrrylmethanes are formed form the beginning of the process, and this can be demonstrated by using synthetic dipyrrylmethanes and tripyrranes.

Synthesis of probable and improbable precursors for porphyrin biosynthesis.

Insights into details of the biomechanism by which porphobilinogen (1) cyclotetramerizes to uroporphyrinogen III (2) as well as promising synthetic applications are provided by investigation of this reaction in vitro. The cyclotetramerization of newly prepared norporphobilinogen (5) proved to be extemely specific due to strong conformation control. Advantage was taken of this finding by preparing a N,N,N,N-tetramethyl-porphyrinogen (13a) for the first time. Protected derivatives of the linear tetramer of porphobilinogen (20c) which is regarded as an intermediate of the cyclotetramerization were gained by total synthesis and their transformations investigated.