Fecal Porphyrin Excretion Research Articles

Swiss patients with variegate porphyria have unique mutations.

Variegate porphyria (VP), also known as South African porphyria, is a low-penetrance, autosomal dominant disorder as the result of a partial deficiency of protoporphyrinogen oxidase (PPOX). Clinically, VP is characterised by photosensitivity and neurovisceral attacks whereby the two symptoms can appear separately or together in patients. VP is little known in Switzerland. In this study, we report a clinical, biochemical and mutational study of eight Swiss VP patients and their families. Six of the eight index patients presented with only skin symptoms, and one with only neurological symptoms. Another patient had both skin and neurological symptoms. Faecal porphyrin excretion was elevated in all patients thus enabling diagnosis. Four different mutations including three novel mutations (G11D, 1041-1042 ins T and 1262-1263 ins 22bp) were identified in this cohort. Mutation 1082-1083 ins C, which had been reported in the French VP population, was shared by five apparently unrelated patients of this study. The novel PPOX gene mutations are apparently unique to the Swiss population. Both clinical and biochemical presentations varied considerably even among those patients who carried an identical mutation, which does not favour the existence of a genotype-phenotype correlation in VP.

Swiss patients with variegate porphyria have unique mutations.

Variegate porphyria (VP), also known as South African porphyria, is a low-penetrance, autosomal dominant disorder as the result of a partial deficiency of protoporphyrinogen oxidase (PPOX). Clinically, VP is characterised by photosensitivity and neurovisceral attacks whereby the two symptoms can appear separately or together in patients. VP is little known in Switzerland. In this study, we report a clinical, biochemical and mutational study of eight Swiss VP patients and their families. Six of the eight index patients presented with only skin symptoms, and one with only neurological symptoms. Another patient had both skin and neurological symptoms. Faecal porphyrin excretion was elevated in all patients thus enabling diagnosis. Four different mutations including three novel mutations (G11D, 1041-1042 ins T and 1262-1263 ins 22bp) were identified in this cohort. Mutation 1082-1083 ins C, which had been reported in the French VP population, was shared by five apparently unrelated patients of this study. The novel PPOX gene mutations are apparently unique to the Swiss population. Both clinical and biochemical presentations varied considerably even among those patients who carried an identical mutation, which does not favour the existence of a genotype-phenotype correlation in VP.

Molecular and Biochemical Studies of Acute Intermittent Porphyria in 196 Patients and Their Families

Acute intermittent porphyria (AIP) is a metabolic disease with clinical manifestations that mimic other abdominal, neurologic, or mental crises. We studied the diagnostic accuracy of current laboratory tests during an acute attack and in remission. Since 1966, we have studied all known Finnish AIP patients (n = 196) and their families (n = 45) and identified the porphobilinogen deaminase (PBGD) mutation in each family. Diagnoses or exclusions of AIP were based on clinical data (including family history), biochemical tests, and in 239 cases, mutation testing. We retrospectively evaluated the diagnostic accuracy of erythrocyte PBGD activity, urinary excretion of porphobilinogen (PBG) and delta-aminolevulinic acid, and urinary and fecal excretion of porphyrins in these patients. Measurement of urinary PBG identified all 35 AIP patients studied during an acute attack. The mean excretion of PBG was 50-fold above the reference interval, although the intraindividual increases were modest (1.6- to 4.0-fold). In the mutation-screened population, urinary PBG analysis identified only 85% of 81 AIP patients studied during remission, but by ROC curve analysis it was nonetheless the best of the biochemical tests. It was increased <or=2-fold in 29% of healthy relatives. Erythrocyte PBGD activity was decreased in only 84% of AIP patients, with results within the reference interval mainly in the variant form of AIP; it was decreased in 23% of healthy relatives. Measurement of urinary PBG is the best biochemical test for AIP, although it is unspecific and does not distinguish AIP from other acute porphyrias. Because the acute increase in PBG is often modest, the medical history, signs, and symptoms must be evaluated carefully during an acute attack. In addition, because biochemical analyses often remain indeterminate in remission, mutation analysis is needed to exclude or confirm the diagnosis of AIP.

Novel Mutation and Polymorphisms of the HMBS Gene Detected by Denaturing HPLC

Acute intermittent porphyria (AIP) is an autosomal dominant, inborn error of the metabolism of heme biosynthesis caused by partial deficiency of hydroxymethylbilane synthase (HMBS). This enzyme catalyzes the condensation of four molecules of porphobilinogen to a tetrapyrrole, hydroxymethylbilane. AIP is characterized by acute attacks of a neurological disorder manifesting as abdominal pain, hypertension, tachycardia, peripheral neuropathy, and mental dysfunction. Biochemical diagnosis of AIP relies on increased urinary porphobilinogen and normal fecal porphyrin excretion (1). Identification of presymptomatic AIP carriers in families with affected individuals is of clinical importance because avoidance of precipitating agents (e.g., drugs and alcohol) can prevent the occurrence of the first porphyric attack, which may be life-threatening. However, there is a significant overlap between the enzyme activities of healthy individuals and patients with AIP (2). In addition, there is a variant of AIP, in which the red blood cell (RBC) HMBS activity is normal (3). DNA-based diagnosis of presymptomatic AIP has been more reliable than diagnosis by RBC HMBS activity, and, thus, direct detection of mutations in presymptomatic AIP is now the definitive approach. The human HMBS gene spans ∼10 kb of DNA on chromosome 11q24.1-24.2 and contains 15 exons (4). Extensive allelic heterogeneity has been demonstrated in this gene, and 159 mutations have been identified in the HMBS gene (5). Most of these mutations are found only in individual families, except those found in Dutch (R116W) (6) and Swedish (W198X) (7) AIP families. Although DNA sequencing can identify all the mutations, this approach is both labor-intensive and time-consuming (8). Several methods that accelerate mutational screening before sequencing have been developed, such as single-strand conformation polymorphism analysis (9), heteroduplex gel analysis (10), and denaturing gradient gel electrophoresis (11)(12). Recently, denaturing HPLC (DHPLC) appears to be more sensitive than other methods in mutation detection, as exemplified …

Novel Single Nucleotide Polymorphism (9678G→A) for Linkage Analysis of Acute Intermittent Porphyria

Acute intermittent porphyria (AIP) is an autosomal dominant inborn error of metabolism caused by a partial deficiency of the third enzyme of the heme biosynthetic pathway, hydroxymethylbilane synthase (EC 4.3.1.8.; HMBS). This enzyme catalyzes the condensation of four molecules of porphobilinogen to a tetrapyrrole hydroxymethylbilane. Clinically, AIP is characterized by acute attacks of neurological disorders manifesting as abdominal pain, hypertension, tachycardia, peripheral neuropathy, and mental dysfunction. Biochemical diagnosis of AIP relies on increased urinary porphobilinogen and fecal porphyrin excretion within the health-related reference interval. The definitive diagnosis requires the assay of red blood cell (RBC) HMBS activity (1). AIP is a genetic disease characterized by poor penetrance; therefore, identification of presymptomatic AIP carriers in families with manifesting individuals is of utmost clinical importance because avoidance of precipitating agents, e.g., drugs and alcohol, can prevent the occurrence of the first porphyric attacks, which may be life-threatening. However, the identification of AIP carriers by assaying the RBC HMBS activity is problematic because there is substantial overlap between the enzyme activities of healthy individuals and patients with AIP (2). In addition, in one variant of AIP, the decreased enzyme activity is confined to …

Analysis of urinary and faecal porphyrin excretion patterns in human porphyrias by fast atom bombardment mass spectrometry

We report a new method for obtaining urinary and faecal porphyrin excretion patterns in human porphyrias based on fast atom bombardment mass spectrometry (FAB-MS). Porphyrins were esterified and extracted from urine or faeces as their methyl esters for analysis by FAB-MS. The protonated pseudo-molecular ion [M + H] + observed for each porphyrin is characteristic of that porphyrin, thus allowing a mixture of porphyrins to be analysed without the need for chromatographic separation. By using tandem MS, identification and characterisation of unknown porphyrins can be achieved. The urinary and faecal porphyrin excretion patterns from various porphyric patients obtained by FAB-MS are in good agreement with those analysed by TLC or HPLC methods.

Effects of chloroquine in hematoporphyrin-treated animals

Porphyrins and related compounds are useful in photodynamic therapy but can cause cutaneous photosensitivity. We determined whether chloroquine, which is effective in treating porphyria cutanea tarda, would mobilize an administered porphyrin from tissues and enhance its excretion. Hematoporphyrin with and without chloroquine was administered to chick embryos, mice, and rats. Tissue and plasma porphyrin levels were markedly increased after hematoporphyrin dosing. Porphyrin concentrations in liver, spleen, and kidney were not significantly affected by chloroquine. Total urinary and fecal porphyrin excretion in rats treated with hematoporphyrin (50 mg/kg, i.p.) was not influenced by chloroquine treatment (100 mg/kg, s.c.). Excretion of heptacarboxylporphyrin, normally a minor fraction of urinary porphyrins, was significantly increased in chloroquine-treated rats. These results suggest that chloroquine is unlikely to be useful after photodynamic therapy for mobilizing exogenous porphyrins from tissues such as liver, spleen, and kidney. Increased urinary excretion of heptacarboxylporphyrin may contribute to the beneficial effect of chloroquine in porphyria cutanea tarda.

Dual porphyria of coexisting variegata and cutanea tarda.

While porphyria cutanea tarda and porphyria variegata are independent diseases, we report on seven rare cases with a coincidence of these two different porphyrias in one individuum. The mutual clinical symptom was a cutaneous photosensitivity, which is a major symptom in porphyria cutanea tarda and a facultative one in porphyria variegata. Additionally, five patients had also experienced episodes of acute abdominal pain, which were in three cases accompanied by neurological symptoms, thus offering evidence for an acute hepatic porphyria, such as porphyria variegata. Determination of urinary porphyrin metabolites revealed a porphyria cutanea tarda-like excretion pattern with an elevation of uroporphyrin (mean 1134 nmol/24 h, range 563-4052, normal < or = 30) and heptacarboxyporphyrin (mean 389 nmol/24 h, range 64-830, normal < or = 4). In all patients, however, urinary coproporphyrin was also increased, reaching levels too high for porphyria cutanea tarda but typical for porphyria variegata (mean 1788 nmol/24 h, range 142-4168, normal < or = 120). Fecal porphyrin excretion also resembled the variegate-type with a high concentration especially of protoporphyrin (mean 628 nmol/g dry weight, range 401-1018, normal < or = 151), accompanied by an increase of coproporphyrin (mean 194 nmol/g dry weight, range 75-409, normal < or = 37). The urinary porphyrin precursors 5-aminolaevulinic acid and porphobilinogen were markedly elevated only in one patient, who was in an acute porphyric phase at the time of investigation. The activity of uroporphyrinogen decarboxylase in erythrocytes was considerably decreased in six of our cases (33-64%) and slightly diminished in the other one (83% of normal activity).(ABSTRACT TRUNCATED AT 250 WORDS)

Prototype Application of a Robot in the Clinical Laboratory Enabling Fully Automated Quantification of Fecal Porphyrins

Unpleasant specimens, sensitive analytes, and a lengthy chromatographic procedure were the main reasons we implemented fecal porphyrin analysis with a laboratory robot. We describe the system in detail and compare it with the same technique performed manually. The day-to-day variation of assays of standards was lower with the robot than with the manual operation: 8% (CV) for coproporphyrin I and 11% for protoporphyrin IX. We repeatedly analyzed a specimen from a healthy volunteer and determined that the specimen contained (in nmol/g dry wt) 7.1 (SD 0.7) for coproporphyrin I, 3.0 (SD 0.4) for coproporphyrin III, and 44.4 (SD 4.3) for protoporphyrin IX. Upper reference limits as measured in 20 healthy volunteers were 20 nmol/g dry wt for coproporphyrin I, 12 nmol/g for coproporphyrin III, and 80 nmol/g for protoporphyrin IX. We also present characteristic chromatograms for samples from various different porphyrias that exhibit abnormal fecal porphyrin excretion. Calculations of return on investment show that the robot, working at full capacity, is a profitable tool.

Skin changes in variegate porphyria

The skin of 20 patients with variegate porphyria (VP) was studied using light, fluorescent, and electron microscopy. Twelve patients had skin symptoms and markedly increased fecal protoporphyrin excretion. Their sun-exposed skin was characterized by homogeneous PAS-positive thickening and IgG deposition in the vessel walls. The basic ultrastructural change was thickening of the vascular walls caused by reduplication of the basal lamina and perivascular deposition of amorphous material. Qualitatively similar but less prominent histopathological changes occurred in sun-protected skin in some of the patients. Six patients had no skin symptoms but an increased porphyrin excretion. The light microscopical changes were comparable to those in the patients with skin symptoms, but the ultrastructural changes were less severe. No abnormal histopathological changes occurred in two symptomless patients who had low lymphocyte protoporphyrinogen oxidase activity but normal fecal porphyrin excretion. These results show that the primary site of skin damage in VP is the vessel wall, and that histopathological changes of the skin also occur in porphyric patients who have never had skin symptoms. Factors determining the occurrence of skin symptoms in VP are discussed.

Effect of ingestion of hemoproteins on fecal excretion of hemes and porphyrins.

Stools from asymptomatic volunteers on diets containing red meat, whole blood, or high fiber were analyzed for their content of hemes and dicarboxylic (heme-derived) porphyrins by the "HemoQuant" assay, the "Hemoccult" test, and "high-performance" liquid chromatography (HPLC). In 49 subjects, ingestion of red meat increased HemoQuant-determined combined fecal heme plus dicarboxylic porphyrins by an average 375%; the contribution of heme-derived porphyrins to total fecal porphyrins increased from 37% to 78%. Of subjects on a red-meat diet, 27% passed stools with a porphyrin content suggestive of a porphyria, compared with only 4% on a red-meat-free diet. These increases were due largely to protoporphyrin and its derivatives pemptoporphyrin and deuteroporphyrin, all of which were present in feces as iron-free porphyrins and iron-ligated (heme) forms. Ingestion of blood had an effect similar to that of red meat, but ingestion of fiber had no effect. These effects of dietary and endogenous hemoproteins must be considered when such methods are used to test feces for occult blood or to test for excess fecal porphyrins as an indicator of a porphyria.

Renal and enteral elimination of coproporphyrin isomers in Rotor's syndrome. A family study

Urinary and fecal total, isomer I, and isomer III coproporphyrin excretion of a Rotor's syndrome patient and his family were determined. The propositus showed increased urinary total coproporphyrin excretion (248 micrograms/24 h) and a shift of the coproporphyrin isomer I/III relation (70%/30%). The propositus's father and two siblings also had elevated renal excretion of coproporphyrin I. Total coproporphyrin excretion was enhanced only in the propositus's father and one sibling, while being normal in another sibling. All family members that could be investigated showed considerably decreased fecal porphyrin excretion. In Rotor's syndrome porphyrin excretion is mainly renal. The coproporphyrin isomer I/III relation is shifted towards isomer I. Phenotypically normal relatives with normal bilirubin plasma levels may have alterations in both their renal and enteral coproporphyrin excretion.

Biochemical study of fecal porphyrin in porphyria cutanea tarda.

AbstractFecal, urinary and erythrocyte porphyrin analyses using the solvent extraction method were performed in 26 patients with porphyria cutanea tarda (PCT) and 144 normal controls. The levels of fecal uroporphyrin (UP) and coproporphyrin (CP) were markedly increased in the PCT group, especially in comparison with the protoporphyrin (PP) level. The values of the UP/PP and CP/PP ratios in the feces were also elevated over those in the control group. It appears that fecal porphyrin excretion is basically similar to urinary porphyrin excretion in PCT. The analysis of fecal porphyrins and the observation of fecal UP/PP and CP/PP ratios may be helpful in the biochemical diagnosis of PCT. In particular, the elevation of CP/PP ratio is characteristic of PCT.

Quantitative and qualitative porphyrin excretion in normal subjects

1. 1. Normal males ( n = 103) excreted a greater amount of urinary porphyrins with a higher percentage of tetracarboxylic coproporphyrin and with a smaller percentage of octocarboxylic uroporphyrin than normal females ( n = 116). 2. 2. These differences were mainly observed in age groups from 21 to 40 yr and from 41 to 60 yr. 3. 3. Quantitative and qualitative faecal porphyrin excretion were similar in both sexes.

Fecal Porphyrin Abnormalities in a Features of Rotor's Patient with Featu Syndrome

Detailed porphyrin analysis and additional ultrastructural abnormalities of liver cells are described in a patient with features of Rotor's syndrome. Normal values of porphyrin precursors, delta-aminolevulinic acid and porphobilinogen, and porphyrins were found in plasma, urine, and erythrocytes. However, fecal porphyrin excretion was markedly increased to 406 micrograms/g dry stool weight (normal range less than 100 micrograms/g) constituted predominantly of protoporphyrin 225 micrograms/g (normal range less than 76 micrograms/g),. with a smaller increase in coproporphyrin III of 56 micrograms/g (normal range less than 20 micrograms/g). Increased fecal porphyrin excretion in this patient appears to reflect abnormal hepatic porphyrin excretion. Electron microscopic examination of liver biopsy tissue showed new abnormalities of peroxisomes and mitochondria. The predominant abnormality of mitochondria was "bleb" deformation of the outer membrane and these changes may reflect abnormal hepatic mitochondrial porphyrin metabolism. The significance of abnormalities involving peroxisomes is uncertain. Increased fecal porphyrin excretion may be a feature of Rotor's syndrome that requires further study.

The effect of bleeding and iron administration on the development of hexachlorobenzene-induced rat porphyria

1. 1. Iron overload and iron depletion respectively potentiated and diminished the porphyrinogenic effect of hexachlorobenzene in rats, as assessed by hepatic, renal and colonie activity of uroporphyrinogen decarboxylase and by urinary and faecal porphyrin excretion. 2. 2. In the early phase of hexachlorobenzene intoxication the rat undergoes a definite porphyric phase different from the porphyria cutanea tarda-like phase which occurs later. 3. 3. Preliminary indications are that faecal and urinary uroporphyrin excretion can be correlated with colonie and renal uroporphyrinogen decarboxylase activity respectively.

Variegate porphyria.

In temperate and cold climates the most usual presenting symptom of variegate porphyria is an acute porphyric attack, indistinguishable from that seen in acute intermittent porphyria. Increased fragility of the skin in sun-exposed areas occurs in only half of such patients, and even then is usually mild and easily overlooked. The diagnosis depends on fecal excretion of porphyrins, which is greatly increased in variegate porphyria and consists predominantly of protoporphyrin. Urinary excretion of porphobilinogen and delta-aminolevulinic acid increases only during acute attacks. There are reasons for thinking that variegate porphyria is commoner than hitherto supposed. During an acute attack a patient without skin symptoms may well be misdiagnosed as having acute intermittent porphyria, because of identical symptoms and excretion of porphobilinogen in the urine. Thus, for a correct diagnosis, every patient presenting with symptoms of acute porphyria requires a fecal analysis.

Faecal porphyrin excretion in various types of porphyria

A new method of thin layer chromatography was used for the study of faecal porphyrins in 31 porphyric patients (20 cases of porphyria cutanea tarda, 5 cases of porphyria variegata, 2 cases of hereditary coproporphyria, 1 case of acute intermittent porphyria and 3 cases of erythropoietic protoporphyria), 14 of their clinically normal relatives and 5 controls. The pattern obtained was characteristic of each type of porphyria and compared to previously published data.

Relationships between acute hepatic porphyrias due to genetic variability of primary enzyme defects and limiting function of uroporphyrinogen synthase

1. 1. Clinical and biochemical analogies and progressions exist among the hereditary acute hepatic porphyrias (AHP), acute intermittent porphyria (AIP). variegate porphyria (PV) and hereditary coproporphyria (HCP). 2. 2. The AHP are essentially regulatory disorders at the molecular level, and have several features in common: drug idiosyncracy, enhanced excretion of porphyrins and their precursors (ALA and PBG) in typical constellations in the urine, elevated porphyrin excretion in the stool (in PV and HCP, and occasionally in AIP). enhanced inducibility of hepatic ALA synthase due to partial enzyme defects in the biosynthetic sequence favorable response to treatment with glucose, propranolol, and hematin, and development in a four-stage progression: phase of the genetic defect→ latency phase (compensated → decompensated) → clinical manifestation. 3. 3. Cutaneous signs can be fully lacking not only in AIP, but also in HCP and PV. 4. 4. The absolute extent of the enzyme defect can vary considerably (overlapping with normal). 5. 5. Gradual transitions between AIP and PV, and probably among AIP, HCP and PV as well. are suggested by several observations: PV can occur without cutaneous signs, thus imitating AIP: in AIP, elevated excretion of porphyrins in the stool is occasionally seen in both the manifest and latent phases, and coproporphyrin can be the dominant urinary porphyrin in AIP: both of these findings can cause confusion of AIP with PV. 6. 6. In HCP fecal excretion of protoporphyrin can reach the same level as that of coproporphyrin. If in AIP patients fecal porphyrin excretion is increased, one obtains the first complete overlapping between AIP and PV. 7. 7. In both AIP and in normals PBG in elevated concentrations can not enzymatically be converted to porphyrinogens in equimolar amounts by liver tissue (“physiologic URO'gen synthase defect”). Thus the step catalysed by URO'gen synthase apparently is the bottleneck or limiting step in porphyrin formation. 8. 8. Similar excretion profiles of porphyrin precursors and porphyrins (COPRO > URO >PENTA or URO >COPRO >PENTA) in AIP indicate limitation of uroporphyrinogen synthesis which due to the kinetics of URO'gen synthase (very low activity in human liver) becomes apparent when there is a massive oversupply of PBG (e.g. oral loading with ALA in normals or induction of ALA synthase in AHP). 9. 9. The increased porphyrin synthesis in AIP can therefore be interpreted as part of a compensatory mechanism or regulatory adaptive process to insure formation of adequate amounts of heme. 10. 10. The elevated substrate supply is due to counterregulation. and compensates the partial defect: elevated excretion of metabolites distal to URO'gen synthase masks the deficiency of this enzyme; elevated porphyrin excretion in a clinical-biochemical sign of compensation in AIP. 11. 11. The enzyme defect in the red cells has no regulatory consequence for hemoglobin synthesis.

PORPHYRIA CUTANEA TARDA—A GENETIC DISEASE?

Abstract. The siblings of patients with clinically manifest porphyria cutanea tarda (PCT) have been investigated by chemical analysis of urinary and faecal porphyrin excretion and by fluorescence microscopical examination of smears from fine needle biopsy specimens of the liver. Four families and one alcoholic and one non‐alcoholic control group were studied. In two families red porphyrin fluorescence was demonstrated in 5 out of 15 asymptomatic siblings, and one sibling who refused liver biopsy was found to have a clinically manifest PCT. In the two other families no porphyrin abnormality could be demonstrated in the 7 asymptomatic siblings. In the control group comprising 36 non‐alcoholic subjects no porphyrin fluorescence could be demonstrated, but in the alcoholic control group of 59 subjects liver porphyrin fluorescence was found in two. Alcoholic abuse was common in the two families in which several members had porphyrin abnormalities indicating latent PCT. In the group of alcoholics liver porphyrin fluorescence was uncommon. Hence the present results indicate that PCT might be a genetic disease.