Stability of porphyrins and porphyrin precursors in urine and plasma samples: implications for sample handling and storage.

The porphyrias are rare disorders of haem biosynthesis. Diagnosis requires demonstrating increased porphyrins or porphyrin precursors in blood, urine and faeces. Patients may only be investigated once, and therefore, understanding the preanalytical factors affecting the reliability of results is crucial. Guidance for sample handling exists, but published evidence regarding the stability of porphyrins and their precursors is limited. The aim of this study was to evaluate the effect of light exposure and different storage temperatures on analyte stability for measurement of urinary aminolaevulinic acid and porphobilinogen, total urine porphyrin and plasma porphyrin. Our results confirm that all samples should be protected from light. Results from samples exposed to light for greater than 4 hours should be interpreted with caution and repeat samples requested. If transported to a specialist laboratory, samples should be stored at 4°C before transport. Transit time at ambient temperatures should be less than 24 hours.

Supramolecular Complexation in Biological Media: NMR Study on Inclusion of an Anionic Tetraarylporphyrin into a Per-O-Methylated ß-Cyclodextrin Cavity in Serum, Blood, and Urine.

The supramolecular complexation of 5,10,15,20-tetrakis(4-sulfonatophenyl)porphyrin (TPPS) with heptakis(2,3,6-tri-O-methyl)-ß-cyclodextrin (TMCD) has been known to be highly specific in aqueous media. In this study, we have used NMR spectroscopy to reveal that this supramolecular system also works even in biologically crowded media such as serum, blood, and urine. A 13 C-labeled heptakis(2,3,6-tri-O-methyl-13 C)-ß-cyclodextrin (13 C-TMCD) was synthesized and studied using one-dimensional (1D) HMQC spectroscopy in serum and blood. The 1D HMQC spectrum of 13 C-TMCD showed clear signals due to the 2-, 3-, and 6-O13 CH3 groups, whose chemical shifts changed upon addition of TPPS due to quantitative formation of the 13 C-TMCD/TPPS=2/1 inclusion complex in such biological media. The 1 H NMR signals of non-isotope-labeled TPPS included by 13 C-TMCD were detected using the 13 C-filtered ROESY technique. A pharmacokinetic study of 13 C-TMCD and its complex with TPPS was carried out in mice using the 1D HMQC method. The results indicated that (1) 1D HMQC is an effective technique for monitoring the inclusion phenomena of 13 C-labeled cyclodextrin in biological media and (2) the intermolecular interaction between 13 C-TMCD and TPPS is highly selective even in contaminated media like blood, serum, and urine.

Experience in management of porphyria cutanea tarda in a tertiary referral Brazilian hospital from 2002 to 2017.

BACKGROUND: Porphyria cutanea tarda (PCT) is the most common porphyria worldwide. The known acquired precipitating factors that induce PCT include alcoholism, hepatitis C virus infection, human immunodeficiency virus infection, and estrogen intake. Hereditary hemochromatosis is considered an inherited risk factor. The aim of this study was to describe and analyze precipitating factors and family history, with emphasis on PCT management. METHODS: A retrospective study of 87 patients with PCT was conducted between January 2002 and December 2017. RESULTS: A male predominance of 1.8 : 1 was found. The median age at diagnosis was 49 years (range 18-71). Family history of PCT was observed in 19.5% of patients. Two or more acquired precipitating factors were present in 42.5%. Patients were treated with antimalarial monotherapy (72.4%), antimalarial combined with phlebotomy (22.9%), and only with phlebotomy (4.6%). Acquired precipitating factors and inherited factors were not associated with treatment group. There was a difference in 24 h-UP normalization rate between treatment groups; combined therapy takes longer than antimalarial monotherapy, 38 months versus 15 months, respectively (CI 95%, 6.5-63.5 vs. 12.9-17) (log-rank test, P = 0.004). CONCLUSION: Precipitating factors did not seem to be associated with treatment choice; however, all acquired and inherited precipitating factors should be investigated, and the choice between phlebotomy and/or antimalarials should be individualized. All dermatologists treating PCT patients should observe transferrin saturation and ferritin levels to search for underlying hereditary hemochromatosis.

Chlorin e6-Coated Superparamagnetic Iron Oxide Nanoparticle (SPION) Nanoclusters as a Theranostic Agent for Dual-Mode Imaging and Photodynamic Therapy.

Photodynamic therapy (PDT) is an approved modality for the treatment of various types of maligancies and diseased states. However, most of the available photosensitizers (PS) are highly hydrophobic, which limits their solubility and dispersion in biological fluids and can lead to self-quenching and sub-optimal therapeutic efficacy. In this study, chlorin e6 (Ce6)-coated superparamagnetic iron oxide nanoparticle (SPION) nanoclusters (Ce6-SCs) were prepared via an oil-in-water emulsion. The physical-chemical properties of the Ce6-SCs were systematically evaluated. Dual-mode imaging and PDT was subsequently performed in tumor-bearing mice. Chlorin e6 is capable of solubilizing hydrophobic SPION into stable, water-soluble nanoclusters without the use of any additional amphiphiles or carriers. The method is reproducible and the Ce6-SCs are highly stable under physiological conditions. The Ce6-SCs have an average diameter of 92 nm and low polydispersity (average PDI < 0.2). Encapsulation efficiency of both Ce6 and SPION is ≈100%, and the total Ce6 payload can be as high as 56% of the total weight (Ce6 + Fe). The Ce6-SCs localize within tumors via enhanced permeability and retention and are detectable by magnetic resonance (MR) and optical imaging. With PDT, Ce6-SCs demonstrate high singlet oxygen generation and produce a significant delay in tumor growth in mice.

Plasma metabonomics investigation reveals involvement of fatty acid oxidation in hematotoxicity in Chinese benzene-exposed workers with low white blood cell count.

Benzene is an environmental and occupational contaminant. Health hazards associated with occupational benzene exposure is a major public health problem in China. In this study, we analyzed metabolite profiles among plasma samples collected from benzene-exposed workers with low white blood cell count (BLWs) and healthy controls using high-performance liquid chromatography-time-of-flight mass spectrometry. To screen potential benzene hematotoxicity biomarkers and metabolic pathways, principal component analysis was used to examine metabolite profile changes in plasma samples. The alterations in fatty acid oxidation (FAO) pathway were consistent with our previous findings in a mouse model; hence, two key genes were selected and verified in WBC samples. A total of nine identified metabolites were significantly changed in BLWs, which were involved in glutathione metabolism, porphyrin metabolism, lipid metabolism pathway, and FAO metabolism. Furthermore, compared with healthy controls, the mRNA expressions of carnitine acyltransferase (CRAT) and ACADVL were significantly increased in BLWs. Particularly, WBC counts was negatively correlated with the expression of AVADVL in BLWs. These aberrant metabolites could act as potential biomarkers for benzene hematotoxicity. In addition, fatty acid oxidation pathway may play a critical role in the development of hematotoxicity caused by benzene.

Phototherapy-induced blistering reaction and eruptive melanocytic nevi in a child with transient neonatal porphyrinemia.

Neonatal blue-light phototherapy induced a blistering reaction followed by eruption of melanocytic nevi on the exposed skin surface of a child with transient neonatal porphyrinemia. New nevi are still developing 4 years after the triggering event. The role of phototoxicity-induced epidermal injury, that of porphyrins and the influence of neonatal blue-light therapy, in this unique phenomenon are discussed.

A study for the detection of kidney cancer using fluorescence emission spectra and synchronous fluorescence excitation spectra of blood and urine.

In this study, we compared different types of biomolecular markers in kidney cancer patients and in normal healthy controls, using fluorescence emission spectra and synchronous fluorescence excitation spectra. We were able to provide an accurate classification of the spectral features of kidney cancer patients relative to that of normal controls, in terms of the concentration ratios of biomolecules (viz., tryptophan, NADH, FAD, basic porphyrin, and acidic porphyrin) based on the intensity of their spectral peaks. The specificity and sensitivity of the method were 90%. The rationale of our current approach is to evolve an innovative protocol for the spectral characterization of in vitro optical analyses suitable for both small clinics and hospitals.

Acquired erythropoietic uroporphyria secondary to myelodysplastic syndrome with chromosome 3 alterations: a case report.

Congenital erythropoietic porphyria is a rare autosomal recessive disease caused by a deficiency of uroporphyrinogen III synthase, owing to mutations in UROS in chromosome 10. Occasionally, patients show a mild, late-onset disease, without germline UROS mutations, associated with haematological malignancies. We report a 65-year-old patient with photosensitivity, overexcretion of porphyrins and thrombocytopenia. Bone marrow analysis gave a diagnosis of myelodysplastic syndrome (MDS) with the presence of a derivative chromosome 3, possibly due to an inversion including 3q21 and 3q26 break points. After allogeneic stem-cell transplantation, complete remission of MDS and uroporphyria was achieved. To our knowledge, this is the first reported case of acquired erythropoietic uroporphyria associated with MDS, with chromosome 3 alterations.

[Clinical and metabolic features of nonalcoholic fatty liver disease in men and women]. / Klinicheskie i metabolicheskie osobennosti nealkogol'noi zhirovoi bolezni pecheni u muzhchin i zhenshchin.

AIM: To evaluate clinical features and metabolic disorders in men and women with non-alcoholic fatty liver disease (NAFLD).

Best practice guidelines on first-line laboratory testing for porphyria.

The porphyrias are disorders of haem biosynthesis which present with acute neurovisceral attacks or disorders of sun-exposed skin. Acute attacks occur mainly in adults and comprise severe abdominal pain, nausea, vomiting, autonomic disturbance, central nervous system involvement and peripheral motor neuropathy. Cutaneous porphyrias can be acute or chronic presenting at various ages. Timely diagnosis depends on clinical suspicion leading to referral of appropriate samples for screening by reliable biochemical methods. All samples should be protected from light. Investigation for an acute attack: • Porphobilinogen (PBG) quantitation in a random urine sample collected during symptoms. Urine concentration must be assessed by measuring creatinine, and a repeat requested if urine creatinine <2 mmol/L. • Urgent porphobilinogen testing should be available within 24 h of sample receipt at the local laboratory. Urine porphyrin excretion (TUP) should subsequently be measured on this urine. • Urine porphobilinogen should be measured using a validated quantitative ion-exchange resin-based method or LC-MS. • Increased urine porphobilinogen excretion requires confirmatory testing and clinical advice from the National Acute Porphyria Service. • Identification of individual acute porphyrias requires analysis of urine, plasma and faecal porphyrins. Investigation for cutaneous porphyria: • An EDTA blood sample for plasma porphyrin fluorescence emission spectroscopy and random urine sample for TUP. • Whole blood for porphyrin analysis is essential to identify protoporphyria. • Faeces need only be collected, if first-line tests are positive or if clinical symptoms persist. Investigation for latent porphyria or family history: • Contact a specialist porphyria laboratory for advice. Clinical, family details are usually required.

Transient Porphyrinemia in a Neonate: A Case Report.

We describe a neonate with anemia, thrombocytopenia, and hyperbilirubinemia secondary to hemolytic disease of the newborn. After phototherapy for hyperbilirubinemia, the neonate developed a photodistributed eruption with high serum and urine porphyrin levels. This transient porphyrinemia resolved at 1 month.

pH-Dependent Changes in the Mechanisms of Transport of Chlorine e6 and Its Derivatives in the Blood.

We studied the effects of medium pH on steady-state distribution of chlorine e6 and its derivatives between the main transport proteins of human blood plasma. The decrease in medium pH from weakly alkaline (pH 7.4) to acid (pH 5.0) was followed by an increase in relative affinity of chlorines to lipoproteins and reduced their affinity to serum albumin. pH-Dependent changes in the parameters of distribution of photosensitizers between the plasma and blood cells was revealed. We discussed the role of charge and polarity degree of photosensitizer molecule in the mechanism of binding to serum albumin. A possible role of changes in hydrogen ion activity in the processes of selective accumulation of chlorines by tumor cells is discussed.

Liver failure after Hydroxycut™ use in a patient with undiagnosed hereditary coproporphyria.

We report the case of a young male presenting with cholestatic liver failure. After an extensive workup, the etiology of the liver failure was determined to be due to hereditary coprophorphyria (HCP). The inciting event was the use of Hydroxycut™, an over-the-counter supplement to promote weight loss that has been reported to cause oxidative liver injury in vulnerable populations. Although HCP is a rare cause of cholestatic liver failure, it is treatable if diagnosed correctly and in a timely manner. In this clinical vignette, we discuss a case that highlights the genetic susceptibility to disease that can be unmasked by environmental exposures. We also review the relevant literature on Hydroxycut™ and how it can affect hepatic function.

Pseudoporphyria following self-medication with chlorophyll.

Two cases of pseudoporphyria are described in which the clinical features of porphyria cutanea tarda occurred in the absence of abnormalities in porphyrin metabolism. Both patients presented with skin fragility and bullae on the dorsal aspect of the hands. The patients consumed a commercial liquid chlorophyll drink in which we detected fluorescent compounds with characteristics typical of previously described chlorophyll derived photosensitisers.