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Background: The clinical features of porphyria cutanea tarda (PCT) are usually distinctive and include blistering on sun-exposed areas, fragile skin, hypertrichosis and hyperpigmentation. Sclerodermatous changes are much less common, and may either reveal PCT or else appear later. We carried out a retrospective study of the files of six female patients presenting such lesions., Patients and Methods: Six women (age: 45 to 72 years) were referred for sclerodermatous lesions on sun-exposed areas of the upper body. In four patients, these lesions revealed PCT and in the remaining two patients they were indicative of previously treated but relapsing PCT. Four had sclerodermatous skin changes mimicking morphea of the neck and neckline, the top of the back and the face, while one presented more diffuse facial and cervical sclerosis. Associated alopecia was seen in three patients. The last patient presented isolated sclerodermiform alopecia. Associated malar hypertrichosis was seen in five cases and facial hyperpigmentation was noted in three cases. Four exhibited no blisters, cutaneous fragility, milia or photosensitivity. Histological findings were consistent with morphea or scleroderma in all cases. All patients presented abnormal liver tests: cirrhosis was present in four cases (primitive biliary cirrhosis, alcoholic cirrhosis and hepatitis C) and fatty liver in two cases. In four cases, there was excessive alcohol intake. Uroporphyrin levels were above the normal range in all cases. Local corticosteroid therapy associated with phlebotomy and/or low-dose hydroxychloroquine resulted in complete normalisation of porphyrin levels in four patients, with complete resolution of the cutaneous lesions in two patients and partial improvement in the other two., Discussion: Sclerodermatous changes are uncommon in PCT. They are not always late and secondary to the process of healing of blisters but can in fact constitute the first cutaneous symptom of the disease while revealing the underlying liver disease. Even in the absence of blisters, photosensitivity or cutaneous fragility, a diagnosis of PCT must be suspected in a setting of sclerodermatous changes distributed on the neck and face, or the neckline, or scarring alopecia, if associated with abnormal liver tests. Skin biopsy to confirm the diagnosis of scleroderma may delay the diagnosis, which is in fact based on porphyrin level. Normalization of the latter parameter under treatment allows regression of lesions., (Copyright © 2013 Elsevier Masson SAS. All rights reserved.)

Two Thai women who are siblings presented with a history of recurrent pruritic vesicles on dorsum of both hands and extensor surface of forearms where the sun-exposed areas are. The excoriated vesicles were healed with depressed scars. They had no previous history of intense abdominal pain, seizure, or psychiatric disorder Urinary porphyrins were analyzed by High Performance Liquid Chromatography (HPLC). The level of coproporphyrin III was detected to be higher than the uroporphyrin level. Fluorescence emission scanning of both patients' plasma was performed and demonstrated typical emission peak at 626 nm, that confirmed the diagnosis of variegate porphyria.

The dysfunction of hepatic heme synthesis by 2,3,7,8-tetrachlordibenzo- p-dioxin (TCDD) in mice, enhanced by iron, leads to accumulation of uroporphyrins I and III (uroporphyria) and resembles the human disorder porphyria cutanea tarda (PCT) precipitated by alcohol and estrogenic drugs. Although consequences of TCDD are considered entirely dependent on the aryl hydrocarbon receptor (AHR), this is not proven for uroporphyria. Administration of TCDD (75 microg/kg) caused uroporphyria in susceptible C57BL/6J mice with high-affinity AHR after 5 weeks (>600-fold increase in hepatic uroporphyrins). Transcriptomics showed significant modified gene expressions for intermediary, heme, and iron metabolism as well as for oxidative stress and cell injury. Resistant low-affinity AHR DBA/2 mice (no increase in porphyrins) showed far fewer changes. At this dose of TCDD, persistent up-regulation of some traditional AH battery genes occurred in both strains. Essentiality of AHR was demonstrated with C57BL/6 Ahr knockout mice. Elevation of hepatic uroporphyrins was 964-fold in Ahr (+/+) mice, lower in Ahr (+/-) (60-fold), but undetectable with Ahr (-/-) . Consistent with an oxidative mechanism, iron overload enhanced porphyria as well as general liver injury in Ahr (+/+) and Ahr (+/-) mice but had no interactive effect in Ahr (-/-) . In contrast, when iron-treated mice received, instead of TCDD, the heme precursor 5-aminolevulinic acid (ALA), causing uroporphyia in Ahr (+/+) mice (242-fold rise in uroporphyrins), elevation of uroporphyrins I and III (42-fold) also occurred in Ahr (-/-) mice and was seemingly associated with AHR-independent expression of Cyp1a2. The findings prove that AHR is a key factor in porphyria induced in mice by TCDD. However, in other models of human PCT, participation of AHR may not be an essential requirement.

Unlabelled: Porphyria cutanea tarda is a liver disease characterized by elevated hepatic iron and excessive production of uroporphyrin (URO). Phlebotomy is an effective treatment that probably acts by reducing hepatic iron. Here we used Hfe(-/-) mice to compare the effects on hepatic URO accumulation of two different methods of hepatic iron depletion: iron chelation using deferiprone (L1) versus iron-deficient diets. Hfe(-/-) mice in a 129S6/SvEvTac background were fed 5-aminolevulinic acid (ALA), which results in hepatic URO accumulation, and increasing doses of L1 in the drinking water. Hepatic URO accumulation was completely prevented at low L1 doses, which partially depleted hepatic nonheme iron. By histological assessment, the decrease in hepatic URO accumulation was associated with greater depletion of iron from hepatocytes than from Kupffer cells. The L1 treatment had no effect on levels of hepatic cytochrome P4501A2 (CYP1A2). L1 also effectively decreased hepatic URO accumulation in C57BL/6 Hfe(-/-) mice treated with ALA and a CYP1A2 inducer. ALA-treated mice maintained on defined iron-deficient diets, rather than chow diets, did not develop uroporphyria, even when the animals were iron-supplemented either directly in the diet or by iron dextran injection., Conclusion: The results suggest that dietary factors other than iron are involved in the development of uroporphyria and that a modest depletion of hepatocyte iron by L1 is sufficient to prevent URO accumulation.

Hydroxylated uroporphyrin I and urochlorin I derivatives formed by photochemical oxidation of uroporphyrinogen I were separated by high-performance liquid chromatography and fully characterized by electrospray ionization tandem mass spectrometry. The porphyrins and chlorins were identified by analysis of their product ion spectra with each hydroxylated derivative giving a characteristic collision-induced dissociation fragmentation pattern. The porphyrins and chlorins characterized were meso-hydroxyuroporphyrin I, alpha-hydroxypropionic acid uroporphyrin I, beta-hydroxypropionic acid uroporphyrin I, hydroxyacetic acid uroporphyrin I, trans-7-hydroxy-8-spirolactoneurochlorin I, cis-7-hydroxy-8-spirolactoneurochlorin I and trans- and cis-7,8-dihydroxyurochlorins I., ((c) 2006 John Wiley & Sons, Ltd.)

Background: Identification of porphyrias relies on the measurement of different porphyrins in urine, feces and plasma. Separation of porphyrin isomers is essential for the differential diagnosis of some porphyrias., Method: Separation of naturally occurring porphyrins was achieved on a Chromolith RP-18 column with fluorimetric detection using a methanol/ammonium acetate gradient mobile phase. Fecal and plasma porphyrins were extracted with acetonitrile and water at different pH values., Results: Eight porphyrins including protoporphyrin eluted within 20 min with good resolution of each of the I and III positional isomer pairs for standards, urine and plasma, and within 50 min for feces. Improvement of the extraction method for fecal and plasmatic porphyrins resulted in high recovery (up to 89%) and reliable quantification of protoporphyrin., Conclusions: The present RP-HPLC method is specific and efficient for routine analysis of porphyrins in human urine, feces and plasma.

Porphyrias are metabolic disorders of heme biosynthesis, which encompass a broad range of symptoms and signs, neurologic, cutaneous or mixed. Because of lack of specificity and polymorphous clinical picture, porphyrias can mimic either neuropsychiatric, dermatologic, or gastrointestinal diseases. We present the case of a 58 years old man to whom clinical presentation suspicious of Addison's disease (melanoderma, fatigue, weight loss, intermittent abdominal pain) was the disguise of porphyria cutanea tarda. A general background of porphyrias and differential diagnosis with other forms of hepatic porphyria, as well as other causes of hyperpigmentation, are given. The clinician should be aware of the protean manifestations of porphyrias and include them in clinical judgment in various situations.

Hepatic uroporphyria can be readily induced by a variety of treatments in mice of the C57BL strains, whereas DBA/2 mice are almost completely resistant. However, feeding of the protoporphyrinogen oxidase-inhibiting herbicide fomesafen (0.25% in the diet for 18 weeks) induced hepatic uroporphyria in male DBA/2N mice (liver porphyrin content up to 150 nmol/g, control animals 1 nmol/g), whereas fomesafen-treated male C57BL/6N mice displayed only a slight elevation of liver porphyrins (approximately 5 nmol/g). The profile of accumulated hepatic porphyrins in fomesafen-treated DBA/2N mice resembled the well-characterised uroporphyria induced by polyhalogenated aromatic hydrocarbons, while histological examination confirmed the presence of uroporphyria-specific cytoplasmic inclusions in the hepatocytes. Uroporphyrinogen decarboxylase activity decreased to about 30% of control values in fomesafen-treated DBA/2N mice; microsomal methoxyresorufin O-dealkylase activity was slightly reduced. The amount of CYP1A1 and CYP1A2 mRNA, as determined by real-time PCR, was not significantly changed; mRNA encoding the housekeeping 5-aminolevulinic acid synthase was elevated 10-fold. Total liver iron was slightly increased. A similar uroporphyria was induced by the herbicide formulation Blazer, containing a structurally related herbicide acifluorfen, when fed to DBA/2N mice at a dose corresponding to 0.25% of acifluorfen in the diet. Since DBA/2 mice are almost completely resistant to all well-characterised porphyrogenic chemicals, the results suggest the possible existence of a yet unknown mechanism of uroporphyria induction, to which the DBA/2 mouse strain is more sensitive than the C57BL strain.

Congenital erythropoietic porphyria (CEP) or Günther's disease is the rarest form of the porphyrias. The disease is usually diagnosed at birth or during early infancy, but rarely in utero. We describe here the first two cases of very early prenatal expression of CEP with cystic hygroma diagnosed at 14 weeks in the first fetus and at 19 weeks in the second. Both fetuses presented with severe nonimmune hydrops fetalis as early as 19 and 22 weeks, associated with intrauterine growth retardation, hyperechogenic kidneys and bones. Amniotic fluid was dark brown and uro- and coproporphyrin I was dramatically increased. Molecular screening of the CEP gene detected heterozygous C73R mutation in both fetuses, the other parental mutation being as yet unknown., (Copyright 2002 John Wiley & Sons, Ltd.)

Vertebrate ALAD is a cytosolic enzyme that catalyzes the second step of porphyrin formation, and it is expressed abundantly in liver and erythroid tissues as well as in other tissues. This unit describes a colorimetric assay for measuring ALAD activity in mammalian blood or tissues and the extent of reactivation of the enzyme in lead-inhibited samples.

Objectives: To describe the biochemical and clinical features in hereditary coproporphyria (HCP)., Design and Method: Within the last 20 years, we investigated 53 patients (male:female = 1:2.5; age = 8-86 years) suffering from HCP. We describe the characteristic levels of urine, and fecal porphyrins and their precursors in hereditary coproporphyria and present the clinical features. Especially, we measured the coproporphyrin isomers I and III., Results and Conclusion: The group of hereditary coproporphyria patients exhibited a significantly higher (p<0.0001) excretion of urinary porphyrin precursors, delta-aminolevulinic acid (median = 84 micromol/24 h) and porphobilinogen (median = 39 micromol/24 h), as compared to controls (delta-aminolevulinic acid: 22 micromol/24 h, porphobilinogen: 3 micromol/24 h; median, n = 20). The median of coproporphyrin in urine (1315 nmol/24 h) and feces (1855 nmol/g) were enhanced 12- and 168-fold, as compared to healthy subjects (urinary coproporphyrin: 106 nmol/24 h, fecal coproporphyrin: 11 nmol/g; median, n = 20). During therapy on one female patient, with IV application of heme arginate, a considerable decline of porphyrin precursors and porphyrin excretion was observed. The examination of urinary and fecal coproporphyrin isomers I and III revealed an excessive elevation of the coproporphyrin isomer III of 87% in urine and 94% in feces, respectively (normal: urinary isomer III = 69-83% and fecal isomer III = 25-40%). In feces the increase of isomer III caused an inversion of the physiologic coproporphyrin isomer III:I ratio that could be recognized in all various stages in hereditary coproporphyria and in children. Acute attacks of hereditary coproporphyria are accompanied by an acute polysymptomatic clinical syndrome, and this is associated with high levels of urinary porphyrin precursors. On review of our patients, the highest percentage had abdominal pain (89%), followed by neurologic (33%), psychiatric (28%), cardiovascular (25%), and skin symptoms (14%).

Arsenite (As[III]) effects on the intermediate steps of heme biosynthesis were studied in adult rat hepatocytes seeded on a feeder layer of 3T3 cells (3T3-hepatocytes) and maintained for 2 weeks with culture medium non-supplemented or supplemented with 150 microM 5-aminolevulinic acid (ALA). The activities of the intracellular enzymes porphobilinogen deaminase (PBG-D), uroporphyrinogen III synthase (UROIII-S), and uroporphyrinogen III decarboxylase (URO-D), and the intermediary uroporphyrins (URO), coproporphyrins (COPRO) and protoporphyrin IX (PROTO) were determined in these cultures. The 3T3-hepatocytes maintained the activities of PBG-D, UROIII-S and URO-D during 2 weeks and ALA addition to the culture medium increased PBG-D (2-3-fold) and UROIII-S (50%) activities and porphyrin production, which accumulated as PROTO. Exposure to 3.9 microM As(III) inhibited UROIII-S activity (down to 34%), and PBG-D and URO-D activities to a lower extent; these effects were magnified by ALA supplementation. As(III) also produced an intracellular accumulation and a decreased excretion of PROTO, and a 31% reduction of the COPRO/URO ratio in the culture medium. Additionally, As(III) caused cytoplasmic vacuolization and lipid accumulation. Our results show that As(III) exposure selectively inhibits several intermediary enzymes of heme metabolism and affects the intra- and extracellular content of porphyrins and their ratio in the culture medium. They also confirm that 3T3-hepatocytes are a suitable in vitro model to study hepatic heme metabolism and its alterations by hepatotoxic chemicals.

Congenital erythropoietic porphyria is a rare autosomal-recessive disorder of the porphyrin metabolism caused by the homozygous defect of uroporphyrinogen III cosynthase. High amounts of uroporphyrin I accumulate in all cells and tissues, reflected by an increased erythrocyte porphyrin concentration and excretion of high porphyrin amounts in urine and feces. Dermal deposits of uroporphyrin frequently induce a dramatic phototoxic oxygen-dependent skin damage with extensive ulcerations and mutilations. Splenomegaly and hemolytic anemia are typical internal symptoms. Skeletal changes such as osteolysis and calcifications are frequent. To date 130 cases of congenital erythropoietic porphyria have been published and are summarized here. Splenectomy, erythrocyte transfusions, and bone marrow transplantation have shown some beneficial effect. The best therapy is the avoidance of sunlight. In the two patients with congenital erythropoietic porphyria described here, oral administration of the oxygen quenchers ascorbic acid and alpha-tocopherol resulted in an improvement in the reduced hemoglobin and erythrocyte concentrations.

A spectrophotometric method for porphobilinogen deaminase assay in erythrocytes is described. This test is determinant for the definite diagnosis of acute intermittent porphyria. In the method described, delta-aminolevulinic acid is used as substrate. Mercaptoethanol and zinc ions are introduced to maintain delta-aminolevulinic acid dehydratase in optimal conditions and to guarantee the in vitro production of porphobilinogen. An incubation temperature of 45 degrees C leads to the production of uroporphyrins, which are measured spectrophotometrically at 405 nm, giving reproducible results. The assay can be performed easily in any clinical laboratory and is valuable for detecting both patients and carriers of acute intermittent porphyria.

Soret-excited resonance Raman (RR) spectra are reported for the sirohemes in the oxidized and Cr11(EDTA)-reduced forms of both desulforubidin from D. baculatus (DSR) and the low molecular weight sulfite reductase from D. vulgaris (1SIR) and for sirohydrochlorin in the oxidized form of desulfoviridin from D. gigas (DSV). Several patterns in the RR spectra of these enzymes can be utilized as signatures for the siroheme/sirohydrochlorin moiety. The active site for DSR and 1SIR consists of a siroheme exchange-coupled to a [4Fe-4S]2+ cluster. Upon addition of Cr11(EDTA), the active center of DSR and 1SIR undergoes a one-electron and two-electron reduction, respectively. The RR spectra of DSR suggest that the siroheme iron is high spin and 5-coordinate in the oxidized enzyme and probably remains high spin and 5-coordinate upon reduction. The iron in the siroheme of oxidized 1SIR changes from a low spin and probably 6-coordinate configuration to a high spin, 5-coordinate complex upon two-electron reduction of the active site. Close similarities between the RR spectral features of the two-electron-reduced assimilatory sulfite reductases from E. coli and from D. vulgaris (1SIR) are discussed.

Porphyria cutanea tarda (PCT) and experimental porphyria are characterized by a decreased activity of the enzyme uroporphyrinogen decarboxylase, and accumulation of uroporphyrins and heptacarboxylporphyrins in the liver. Iron (Fe) plays an important role in PCT and experimental porphyria. Biochemically and electron microscopically, we examined the relationship between Fe and porphyrins in liver tissue of C57BL/10 mice made porphyric by administration of iron dextran as Imferon (IMF), and in liver biopsies of patients with symptomatic PCT. Accumulation of uroporphyrins and heptacarboxylporphyrins, and an increased amount of Fe were observed in livers of mice treated with IMF and in liver biopsies of patients with PCT. In mice treated with IMF, the activity of uroporphyrinogen decarboxylase was decreased. Both in livers of mice treated with IMF and in livers of patients with PCT, needle-like structures, representing uroporphyrin crystals, were observed by electron microscopy. Uroporphyrin crystals and Fe (as ferritin) were observed in the same hepatocyte. Moreover, there was a striking morphological correlation between uroporphyrin crystals and ferritin-Fe, suggesting a role for (ferritin-)Fe in the pathogenesis of porphyria.

A simple method for the preparation and reversed-phase high-performance liquid chromatographic separation of hexacarboxylic porphyrinogen isomers is described. Uroporphyrin I or III was partially decarboxylated in 0.5 M hydrochloric acid at 150 degrees C. Unreacted uroporphyrin and the hepta-, hexa- and pentacarboxylic porphyrins formed were esterified and then group-separated by thin-layer chromatography. After hydrolysis, the porphyrins were reduced to the corresponding porphyrinogens with 3% (w/w) sodium amalgam. The hexacarboxylic porphyrinogens were separated on an ODS-Hypersil column by elution with acetonitrile-methanol-1 M ammonium acetate, pH 5.16 (8:12:80, v/v/v) as mobile phase. Isomers were identified by high-performance liquid chromatography of the characteristic mixture of two pentacarboxylic porphyrins formed after partial decarboxylation of individual isomers. Except for the two type I isomers, resolution of the hexacarboxylic porphyrinogens was superior to that of the corresponding porphyrins.

Four women, aged 23 to 26 years, fell ill with cutaneous hepatic porphyria. All of them had taken oestrogen-contaning oral contraceptives three to eight years before the illness. In one instance there was a time relationship with a progestogen preparation. The clinical and biochemical signs were those of porphyria cutanea tarda, but contrary to this disease there was a shift towards heptacarboxyporphyrin in the uroporphyrin/heptacarboxyporphyrin ratio (thin-layer chromatography, classified according to Doss). This difference and the onset at an early age suggest that the described disease may be called a hormone-induced premature cutaneous porphyria.

14C-labelled coproporphyrin I and III and uroporphyrin I were injected intravenously into healthy human subjects. Two experiments were performed with each porphyrin. Counting of blood, urine and faeces was made as a function of time. The coproporphyrin data were simulated by means of an analogy computer by using a four-compartment model with a delay pool. The physiological counterparts to the compartments were considered to be liver, blood and gut. From the blood radioactivity disappearance curve of uroporphyrin three exponential components could be worked out. The least square method was used for the solution of the plasma and urine radioactivity curves. A three-compartment model was constructed to illustrate the kinetics of uroporphyrin I. Coproporphyrin I and III behaved essentially similarly, whilst the kinetic behaviour of uroporphyrin I was completely different from that of coproporphyrins. A rapid hepatic uptake, preferential faecal excretion and a significant enterohepatic circulation were features of coproporphyrin kinetics. Over 70 per cent of the tracer activity of uroporphyrin I was found to accumulate to a rather large pool (volume 16--28 times the plasma volume) from where the back flow to plasma was rather slow. Practically all the uroporphyrin was excreted via urine.

Magnetic circular dichroism is a suitable technique for semiquantitative determination of total porphyrins in urine and feces, being rapid, reliable, and involving little sample preparation. The sensitivity of the assay suffices to distinguish normal from above-normal concentrations of total porphyrins in urine and feces. Results for urine compare well with those obtained by thin-layer chromatography of porphyrin esters (n = 42, r = 0.92) or ion-exchange column chromatography of porphyrin acids (n = 19, r = 0.80).

The porphyrin status of 38 renal transplant patients was investigated using a sensitive thin-layer chromatographic assay. Abnormalities of porphyrin synthesis observed in patients on hemodialysis were generally reversed on receipt of a transplant with sustained good renal function. Urinary coproporphyrin levels and the creatinine clearance appeared directly related in both individual cases and generally, substantiating the theory that coproporphyrin is of renal origin. The pre- and postoperative porphyrin status of a unique patient with variegate porphyria who received a successful renal transplant implies that the excess urinary porphyrins and precursors excreted in the acute porphyrias are also of mainly renal, as opposed to hepatic origin. Furthermore, the posttransplant overproduction of porphyrins by a kidney from a normal donor suggests the existence of an endogenous porphyrogenic agent in the circulation of patients with acute porphyria.

An open trial with canthaxanthin, a photoprotective agent, was carried out on 7 patients with erythrohepatic protoporphyria (EHP), 5 patients with variegate porphyria (VP), and 2 patients with symptomatic porphyria (SP). In 4 of the 7 patients with EHP, sun tolerance was significantly but only moderately increased. In 2 children sun tolerance was greatly increased. Only 2 of the 5 patients with VP and 1 of the 2 patients with SP showed any improvement.

We recorded fluorescence excitation and emission spectra of uro- and coproporphyrin under different experimental conditions, to see how these conditions influence quantifications based on measurement of fluorescence intensity. We found that, for bands alpha and beta of the emission spectra and the main peak of the excitation spectra, fluorescence depends on pH and is minimal near pH 5 and near pH 7-7.5 for copro- and uroporphyrin, respectively. For band gamma of the emission spectra there was a constant decrease of fluorescence with increasing alkalinity of the solution. The intensity of porphyrin fluorescence also depends on ionic strength, reaching sharp maxima at 0.1 mol/L (for uroporphyrin) and 1 mol/L (for coproporphyrin). The organic mixture ethyl acetate:acetic acid (4:1 by vol), commonly used to extract porphyrins from biological samples, markedly diminishes the fluorescence of both porphyrins as compared with the same concentration of each porphyrin in aqueous acidic solvent. Furthermore, when we measured different ratios of uro:copro mixture at three distinct pHs and buffers, we found that at pH 10.5 (in carbonate buffer) the measured units of fluorescence depend only on total porphyrin concentration and not on the composition of the mixture.

The patterns of urinary porphyria excretion and hepatic porphyrin accumulation in hexachlorobenzene-treated rats is similar to that observed in overt porphyria cutanea tarda and may be attributed to a decrease in activity of hepatic uroporphyrinogen (UROGEN) decarboxylase. The 3.5 diethoxycarbonyl-1,4-dihydrocollidine (DDC)-treated rat has been evaluated as a model to test the porphyrinogenicity of drugs.

The activity of uroporphyrinogen III cosynthetase may be determined in a direct assay system in which preuroporphyrinogen is generated from porphobilinogen by the action of porphobilinogen deaminase. The basis of the assay relies on the much faster enzymatic conversion of preuroporphyrinogen into uroporphyrinogen III by cosynthetase compared with the non-enzymic decomposition of preuroporphyrinogen into uroporphyrinogen I. The amount of porphyrinogen (measured as porphyrins) formed from porphobilinogen in the presence of the cosynthetase, after subtraction of the porphyrin formed with deaminase alone, gives a direct assessment of the uroporphyrinogen III and therefore of the cosynthetase activity. The assay may be utilized for the determination of pure cosynthetases and for cosynthetases in crude systems such as whole blood haemolysates or liver homogenates. The assay can be accomplished in a fraction of the time required for previous methods.

A boy exhibited severe bullous skin disease a few days after birth, followed by increased fragility of the exposed skin in spring and summer. Examination at 2 1/2 years of age led to characteristic biochemical findings: increased excretion of fecal porphyrins (coproporphyrin 121 to 131 and protoporphyrin 467 to 576 nmol/g dry weight), and increased erythrocyte protoporphyrin concentration (3643 to 4840 nmol/l). Lymphocyte protoporphyrinogen oxidase activity was very low in the patient (0.4 nmol/mg protein/h) and half-normal (2.7 and 2.3 nmol/mg protein/h) in the parents, suggesting that the patient had homozygous variegate porphyria. Severe skin symptoms and a high concentration of red cell protoporphyrin concentration in an infant should prompt suspicion of homozygous acute hepatic porphyria.

The routine clinical analysis of the porphyrin precursors of heme requires an internal standard to determine the efficiency of the analytical procedure used. 2-Vinyl-4-hydroxymethyldeuteroporphyrin IX has been prepared as an internal standard. Its application to porphyrin analysis has been demonstrated using high-performance liquid chromatographic resolution of the uro- to protoporphyrins in normal and porphyric urine.

Thirty patients suffering from chronic kidney disease were examined for urinary, fecal and total plasma porphyrins. Twenty patients had nitrogen retention whereas the remaining ten patients displayed normal values. Urinary coproporphyrin excretion and total plasma porphyrin levels were comparatively less pronounced in patients with nitrogen retention. Such changes in porphyrin metabolism could be explained by a diminished capacity for kidney excretion. It is concluded that the total amount of porphyrins synthesized in the body is decreased as a result of nitrogen retention.

Altogether 98 patients have been examined, suffering from latent porphyria (n = 18) and manifest porphyria cutanea tarda (n = 80). 90 (91.8%) examinees abused alcohol. Clinical manifestation of the disease is associated with an essential rise of the blood plasma and urine uroporphyrin. Three types of porphyrinemia have been detected in the patients with manifest porphyria cutanea tarda, these types related to the degree of uroporphyrin level increase in the blood plasma and to the length of the disease: (I) uroporphyrinemia, (II) urocoproporphyrinemia, (III) urocoproprotoporphyrinemia. Porphyrinemia types are estimated as successive stages in the development of porphyrin metabolism disorders and are regarded among the criteria determining the clinical pattern of manifest porphyria cutanea tarda.

Delta-aminolevulinic acid (DALA), porphobilinogen (PBG) and porphyrins in urine and feces were determined in 40 healthy controls (20 males and 20 females) and in 60 patients with acute leukosis (27 males and 33 females), with chronic myeloleukosis - 23 (10 males and 13 females), and 25 patients with chronic lympholeukosis (II males and 14 females). Another 15 patients (7 males and 8 females) were studied at the initial stage, recurrence, recidivation of acute leukosis (AL). DALA excreted with the urine, manifested no significant discrepancy as compared with the controls of the three groups examined patients and in all the three stage of AL, whereas PBG was moderately reduced in the AL patients - initial stage and recurrence and was within the normal limits in the patients with chronic myeloleukosis (CML) and chronic lympholeukosis (CLL). The urine excreted coproporhyrin was increased, to various degrees, in the patients with CML and CLL and at the initial stage, recurrence and recidivation of AL, whereas uroporhyrin was within the norm. Coproporphyrin, excreted in the feces, was increased in all three groups of patients with leukosis and in the three stages of AL, whereas the other fractions showed no significant difference as compared with the controls. It could be concluded, from the results obtained, that porphyrins metabolism is disturbed in the patients with leukosis.

Mass spectrometric studies were performed on selected specimens from my collection of over 700 crystalline porphyrin esters prepared over 20 years as well as on similar preparations from other laboratories. Mass spectrometry was performed by J Moller (Department of Chemistry, Odense University) and Elfinn Larsen (Department of Chemistry, The Experimental Division, Risoe, of the Danish Atomic Energy Commission). Nearly all typical crystalline porphyrins contained Cu- and Ni- and minor amounts of Zn- and Fe2-porphyrins, most often above one molar per cent, sometimes 5--10% and occasionally more, up to 35%. There was no close correlation between the metal content and the melting point. A method is proposed for preparation of metal-free ester preparations: Dissolution of either free or esterified porphyrin in conc. H2SO4; mixing of the solution with 19 volumes of methanol; after at least 12 hours adding of 2 vol. CHCl3 and 2 vol. H2O, and washing the CHCl3, phase with 1/2 vol. distilled water until washwater becomes neutral (8--10 times). Preparations made in this way showed below 0.5% metalloporphyrin contamination. The method does not work with protoporphyrin, which is oxidized by conc. H2SO4. The acid extraction leaves some porphyrin partially esterified, which can be removed by preparative TLC. A TLC-method for detection of metalloporphyrin in porphyrin esters is described.

Porphyria variegata, one of the rarer forms of hepatic porphyria, is brought to the attention through presentation of one case. Its clinical and chemical laboratory characteristics are compared with other hepatic porphyrias and pathogenesis, hereditary transmission and therapy are gone into.

A novel reversed-phase system is described for the simultaneous separation of type I and type III isomers of uro-, heptacarboxylic-, hexacarboxylic -, pentacarboxylic - and copro -porphyrins and the dicarboxylic meso- and proto-porphyrins. The porphyrins were resolved on a Hypersil -SAS column eluted with 10% (v/v) acetonitrile in 1 mol/l ammonium acetate, pH 5.16 (solvent A) and 10% (v/v) acetonitrile in methanol (solvent B) with a linear gradient from 100% A (0%B) to 35% A (65%B) in 30 min followed by isocratic elution at 65% B for a further 10 min. The method is simple and reproducible and has been applied to the analysis of porphyrins in urine and faeces from patients with acute intermittent, variegate, hereditary copro , congenital erythropoietic, erythrohepatic proto and symptomatic porphyrias.