High-performance liquid chromatography of coproporphyrin isomers

Wright, D. J.; Rideout, J.M.; Lim, Chang Kee

doi:10.1042/bj2090553

Cited by 41 publications

(18 citation statements)

References 12 publications

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“…The methodology previously used for the estimation of urinary porphyrin isomer ratios allowed only the determination of isomers I and III. However, two independent HPLC techniques are available for the simultaneous Separation of all four coproporphyrin isomers, namely the hydrophobic interaction technique (14) and the ion-pair technique (2,15). Application of these advanced Separation methods to the pH-related analysis of urinary coproporphyrins from patients with abnormal isomer distribution rnight be helpful for further elucidation of the underlying excretion mechanisms.…”

Section: Preparation Of Urine Samples For Isomer Analysismentioning

confidence: 99%

The Isomer Ratios of Urinary Coproporphyrins I — IV are pH-Dependent

Jacob

Egeler²,

Hennel³

et al. 1991

Clinical Chemistry and Laboratory Medicine

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Summary:The percentage of porphyrinogens äs related to total porphyrin excretion was determined in the ! urine of healthy subjects. Acidic urines (pH 5.0 to 5.9) contained 62.9 + 10.7% (x ± s, N = 11) porphyrinogens, | whereas in neutral urines (pH 6.0 to 7.2) a somewhat lower percentage (51.2 + 15.3%, N = 11) was detected.However, there was no significant difference between the mean porphyrinogen Contents of acidic and neutral urines.Evidence was found for a previously unreported pH-dependent influence on the isomer ratios of urinary coproporphyrins I and III. Acidic urines (N = 18) from healthy subjects showed significantly higher percentages , of isomer I (27.1 + 6.4%), isomer II (2.7 ± 1.1%), and isomer IV (5.0 ± 1.3%) äs compared to respective values from neutral urines (22.2 + 5.1% isomer I, 0.6 ± 0.6% isomer II, and 1.5 + 1.3% isomer IV; N = 16, p < 0.001). Conversely, the percentage of isomer III was markedly lower in acidic urines than in neutral urines (65.1 + 7.9% vs. 75.9 + 5.4%; p < 0.001). The same relationship was confirmed in an individual subject by analysis of a series of urines (N = 13) with pH values ranging from 5.4 to 7.3. These results point to the possibiiity that the atypical coproporphyrin isomers II and IV are predominantly formed by an increased isomerization rate of coproporphyrinogens under acidic intravesical conditions.In the course of systematic investigations on the forCoproporphyrins of the series I and III are the prin-mation of the atypical coproporphyrins II and IV by cipal isomers to be excreted in human urine, whereas means of isocratic ion-pair high-performance liquid the atypical isomers II and IV are only present in chromatography (HPLC), we previously observed a small amounts (1-3). It has been suggested that stringent influence of urinary pH on the respective formation of the latter results from non-enzymatic isomer ratios (8). self-isomerization of the naturally occurring copro-Here we report our subsequent studies on the influporphyrinogens (3). Porphyrinogens are the metabol-ence of urinary pH on porphyrinogen excretion and ically active intermediates in the biosynthesis of haem. on the isomer ratios of the individual coproporphyrins They are easily oxidized to the corresponding por-I -IV. phyrins in vivo and in vitro. Both porphyrinogens and porphyrins are excreted in urine (4). The proportion of porphyrins and their reduced precursors has Materials and Methods been determined by various methods in urine of bpth individuals investigated porphyric patients (5-7) and healthy controls (7)..Spontaneous urine samples of apparently healthy persons of the laboratory staff were-used in the study. One of us (P. L.) took voluntarily part in the enforced pH shifting experiment. Acidic urines were obtained by a high intake of ascorbic acid (up to 5 g per day), whüe neutral and alkaline urines were ! ) Presented in part at the Conference "Biochemische Analytik produced after Ingestion of large doses of sodium-potassium-90", Munich, Germany. hydrogen citrate (up to 10 g per day).

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Section: Preparation Of Urine Samples For Isomer Analysismentioning

confidence: 99%

The Isomer Ratios of Urinary Coproporphyrins I — IV are pH-Dependent

Jacob

Egeler²,

Hennel³

et al. 1991

Clinical Chemistry and Laboratory Medicine

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“…Cultures were centrifuged, and the supernatant was diluted 200-fold in 0.1 M HCl. High-performance liquid chromatography (HPLC) analysis was as described by Wright and colleagues (47). A Symmetry C 18 analytical column (particle size, 5 m; 250 mm by 3 mm) (Waters) was used for separation.…”

Section: Vol 192 2010 Pleiotropic Effects Of Sam Deficit 1239mentioning

confidence: 99%

Coproporphyrin Excretion and Low Thiol Levels Caused by Point Mutation in the Rhodobacter sphaeroides S -Adenosylmethionine Synthetase Gene

et al. 2010

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A spontaneous mutant of Rhodobacter sphaeroides f. sp. denitrificans IL-106 was found to excrete a large amount of a red compound identified as coproporphyrin III, an intermediate in bacteriochlorophyll and heme synthesis. The mutant, named PORF, is able to grow under phototrophic conditions but has low levels of intracellular cysteine and glutathione and overexpresses the cysteine synthase CysK. The expression of molybdoenzymes such as dimethyl sulfoxide (DMSO) and nitrate reductases is also affected under certain growth conditions. Excretion of coproporphyrin and overexpression of CysK are not directly related but were both found to be consequences of a diminished synthesis of the key metabolite S-adenosylmethionine (SAM). The wild-type phenotype is restored when the gene metK encoding SAM synthetase is supplied in trans. The metK gene in the mutant strain has a mutation leading to a single amino acid change (H145Y) in the encoded protein. This point mutation is responsible for a 70% decrease in intracellular SAM content which probably affects the activities of numerous SAM-dependent enzymes such as coproporphyrinogen oxidase (HemN); uroporphyrinogen III methyltransferase (CobA), which is involved in siroheme synthesis; and molybdenum cofactor biosynthesis protein A (MoaA). We propose a model showing that the attenuation of the activities of SAM-dependent enzymes in the mutant could be responsible for the coproporphyrin excretion, the low cysteine and glutathione contents, and the decrease in DMSO and nitrate reductase activities.

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“…The UIIIS assay is performed as described above with the addition of 2 g of purified recombinant UIIIS (rUIIIS). The porphyrin isomers were resolved by a method derived from those described previously (35,39). An HPLC system that consisted of a Waters 2795 separations module (Alliance HT), a Waters 474 scanning fluorescence detector, a Waters 2996 photodiode array detector and a Phenomenex Gemini C 18 column (250 mm by 4.6 mm by 5 m) was used.…”

Section: In Vitro Assays Of Porphobilinogen Deaminase (Pbgd) Uroporpmentioning

confidence: 99%

Sampangine Inhibits Heme Biosynthesis in both Yeast and Human

Huang

Chen

et al. 2011

Eukaryot Cell

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The azaoxoaporphine alkaloid sampangine exhibits strong antiproliferation activity in various organisms. Previous studies suggested that it somehow affects heme metabolism and stimulates production of reactive oxygen species (ROS). In this study, we show that inhibition of heme biosynthesis is the primary mechanism of action by sampangine and that increases in the levels of reactive oxygen species are secondary to heme deficiency. We directly demonstrate that sampangine inhibits heme synthesis in the yeast Saccharomyces cerevisiae. It also causes accumulation of uroporphyrinogen and its decarboxylated derivatives, intermediate products of the heme biosynthesis pathway. Our results also suggest that sampangine likely works through an unusual mechanism-by hyperactivating uroporhyrinogen III synthase-to inhibit heme biosynthesis. We also show that the inhibitory effect of sampangine on heme synthesis is conserved in human cells. This study also reveals a surprising essential role for the interaction between the mitochondrial ATP synthase and the electron transport chain.

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High-performance liquid chromatography of coproporphyrin isomers

Abstract: A reversed-phase system is described for the simultaneous isocratic separation of coproporphyrin I, II, III and IV isomers. The retention behaviour of coproporphyrin I and III is studied in detail. The method is suitable for both analytical and semi-preparative separation.

Cited by 41 publications

References 12 publications

The Isomer Ratios of Urinary Coproporphyrins I — IV are pH-Dependent

The Isomer Ratios of Urinary Coproporphyrins I — IV are pH-Dependent

Coproporphyrin Excretion and Low Thiol Levels Caused by Point Mutation in the Rhodobacter sphaeroides S -Adenosylmethionine Synthetase Gene

Sampangine Inhibits Heme Biosynthesis in both Yeast and Human

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