Assessment of cassava toxicity in patients with tropical chronic pancreatitis.

BACKGROUND AND AIM: There have been conflicting reports on the role of cassava ingestion in tropical pancreatitis (TCP). In this study we aimed to estimate cyanogens detoxifying enzyme rhodanese, thiocyanate and sulfur containing amino acids in cassava consumer as well as cassava non-consumer TCP patients and healthy controls and compare the same. METHODS: Eighty-six TCP patients and 90 healthy controls were recruited. Serum rhodanese, thiocyanate, plasma amino acids, urinary inorganic sulfate/creatinine were measured. RESULTS: There was significant reduction in serum rhodanese activity in both cassava consumer- and non-consumer TCP patients as compared to controls but no significant difference between cassava consumer- and non-consumer TCP patients was observed. Serum thiocyanate was significantly lower in cassava consumer TCP patients as compared to cassava consumer controls but not significantly different from cassava non-consumer TCP patients. Plasma methionine, cysteine and urinary inorganic sulfate / creatinine ratio was significantly lower in both cassava consumer and non-consumer TCP patients as compared to controls but were comparable among cassava consumers and non-consumers. CONCLUSIONS: Significant reduction in rhodanese activity with concomitant decrease in sulfur containing amino acids and antioxidants such as glutathione suggests that TCP patients are at higher risk of defective detoxification of cyanogens. However there was no difference between cassava consumers and non-consumers. Low levels of sulfur amino acids may contribute to the development of pancreatitis.

Rhodanese, but not cystathionine-gamma-lyase, is associated with dextran sulfate sodium-evoked colitis in mice: a sign of impaired colonic sulfide detoxification?

Clinical studies suggest that colonic luminal hydrogen sulfide (H(2)S), produced by sulfate-reducing bacteria or through other pathways, might be involved in the pathogenesis of inflammatory bowel disease (IBD). Nonetheless, this hypothesis has been poorly investigated by basic studies using laboratory animals. We thus focused on two enzymes, cystathionine-gamma-lyase (CSE) that generates H(2)S from l-cysteine, and rhodanese that directly or indirectly detoxifies H(2)S, particularly in relation to the colitis induced by dextran sulfate sodium (DSS) in mice. CSE was a major H(2)S-forming enzyme in colonic and renal homogenates from mice and rats, and the rhodanese activity was also detectable in both tissues. Colitis-related symptoms including decreased body weight gain, diarrhea, hematochezia and shortening of colon length were observed in the mice drinking DSS. Those symptoms were not or only slightly attenuated by repeated administration of a CSE inhibitor. CSE activity and protein levels in the colonic tissue did not notably change in the mice with colitis. In contrast, the activity and protein/mRNA levels of rhodanese in the colon, but not kidney, significantly decreased nearly in parallel with the development of colitis, followed by elevation of rhodanese activity in red blood cells (RBCs). These data show that rhodanese, but not CSE, is associated with DSS-induced colitis in mice, leading to a hypothesis that impaired detoxification of H(2)S due to down-regulation or suppression of colonic rhodanese is involved in IBD. The delayed enhancement of rhodanese activity in RBCs, a possible compensative event, might be available as a disease marker for IBD.

Effect of glucose-cysteine adduct on cysteine desulfuration in guinea pig tissues.

Effect of intraperitoneal administration (12 mmol/kg of body weight) of glucose-cysteine adduct 2-(D-gluco-pentahydroxypentyl)-thiazolidine-4-carboxylate, (glc-cys) on the rhodanese, gamma-cystathionase and 3-mercaptopyruvate sulfurtransferase (MPST) activity levels in guinea pig tissues was studied. The rhodanese activity value in liver increased by 41%, 3-mercaptopyruvate sulfurtransferase by 24%, and gamma-cystathionase by 12% after three successive days of the administration. In the kidney, on the contrary, glc-cys administration resulted in about 18% decrease in the gamma-cystathionase activity value, whereas no changes in MPST and rhodanese activity values were observed. In the case of the brain, rhodanese and gamma-cystathionase did not change their activity but the activity of MPST decreased by 21%. MPST level did not change substantially in whole blood after glc-cys treatment. The results seem to indicate that in guinea pig liver but not in kidney and brain, glc-cys has a potential to activate the desulfuration pathway of L-cysteine metabolism.

Changes in arginase, aminotransferases and rhodanese in sera of domestic animals with experimentally induced liver necrosis.

Changes in the serum concentrations of aspartic aminotransferase (AST), alanine aminotransferase (ALT), rhodanese and arginase were measured in dogs, sheep and cattle with hepatic necrosis induced by the oral administration of carbon tetrachloride. A new method for arginase assay was based on the determination of remaining arginine (after its conversion to urea and ornithine) by its reaction with p-nitrophenyl glyoxal (PNPG). In all species studied the serum arginase increased 6-12 h after liver damage, reached a peak value in 48 h and returned to normal thereafter. Rhodanese activity did not change in dogs but rose significantly in sheep and, to a lesser extent, in cattle. AST increased strikingly in sheep as compared with dogs and cattle and remained high for > 5 days. In dogs ALT rose sharply and remained elevated for > 10 days. No change in ALT was seen in sheep or cattle. The determination of arginase by a simple procedure such as the PNPG method, in conjunction with AST or ALT assay, may be of value in assessing the stage of liver necrosis.

Improved method for measurement of rhodanese activity using methanethiosulfonate as sulfur donor substrate and its application to human serum.

We have developed a sensitive method for the measurement of rhodanese activity in human serum which is based on the colorimetric method for the determination of thiocyanate produced from methanethiosulfonate and cyanide as substrates. Thiocyanate gives a red complex with ferric ion in an acidic condition. The present method is about 70-fold more sensitive than the conventional method using cyanide and thiosulfate as substrates and correlates well (r = 0.997) with the conventional method in bovine liver rhodanese. Within-run precision of the method is 0.91% for 420 units/l serum and the calibration curve is linear up to 1850 units/l. The normal value for human serum, determined by the present method on 31 healthy persons, was 20.9 +/- 20.0 units/l (mean +/- 2S.D.). Rhodanese activity was clearly elevated in some serum samples which were observed at abnormal values in some biochemical diagnostic tests and showed significant positive correlations with guanase activity (r = 0.728, p less than 0.01) and glutamic-oxalacetic transaminase activity (r = 0.625, p less than 0.01).

An experimental assessment of cyanide challenge dose-response, time-course and recovery of indicative toxicity parameters.

1. Several sub-lethal doses of cyanide were assayed with the aim of obtaining significant differences in the parameters studied. A dose of 4 mg/kg s.c. was selected. 2. Present studies were carried out to determine the time dependence of the effects produced by s.c. administration of a single dose of potassium cyanide as well as the recovery time of some of the toxicity indicative parameters. 3. It was found that cyanide effects continued for at least 3 days; after the parameters had recovered normal values.

Serum rhodanese in goitre and calcific pancreatitis of tropics.

Rhodanese is one of the enzymes concerned in the detoxification of cyanide. Cassava intake and consequent cyanide toxicity are incriminated in the pathogenesis of goitre and calcific pancreatitis of tropics. So we studied the activity of rhodanese in these patients. 14 controls, 13 patients with pancreatitis and 12 with goitre were studied. The median (and range) of rhodanese in these groups were 82 (50-144), 110 (64-180) and 71 (22-160) units respectively. The serum rhodanese was significantly higher (P less than 0.05) in patients with pancreatitis when compared to the other groups. There was no significant difference between the serum rhodanese in patients with goitre and the controls. The presence of adequate amounts of rhodanese indicates that goitre and chronic pancreatitis are not produced by impaired cyanide detoxification.

Pattern of enzyme changes in rabbits administered linamarin or potassium cyanide.

Diseases like tropical ataxic neuropathy and endemic goitre have been reported to have definite correlation with a chronic ingestion of cassava (Manihot esculenta Crantz). The toxicity of cassava has been attributed to its two cyanogenic glycosides, linamarin and lotaustralin. In this study, an attempt has been made to understand the pattern of changes in certain clinically significant enzymes brought about by the chronic administration of sublethal doses of linamarin to rabbits. The profound elevation in rhodanese activity observed in the linamarin and cyanide treated rabbits indicated the attempt of the tissues to detoxify cyanide. That intact linamarin could be hydrolysed in vivo was a significant finding from the study. The mode of toxicity of linamarin was similar to that of cyanide by producing a gradual shift from aerobic to anaerobic metabolism.

Interspecies differences in rhodanese (thiosulfate sulfurtransferase, EC 2.8.1.1) activity in liver, kidney and plasma.

Rhodanese levels have been measured in liver, kidney and plasma from a number of species. Liver activity was low in marmosets, pigeons and beagle bitches. Levels were high in rats and somewhat lower in hamsters and guinea pigs while levels in two strains of rabbits were intermediate between guinea pigs and marmosets. The relationship between hepatic and plasma rhodanese and cyanide sensitivity is discussed.

Red blood cell rhodanese: its possible role in modulating delta-aminolaevulinate synthetase activity in mammals.

The optimum conditions for measuring rhodanese activity in human erythrocytes were established. The mean control values for males (112 nmol SCN/30 min/mg protein) and females (127 nmol SCN/30 min/mg protein) were determined. Rhodanese activity was measured in different porphyric patients. The activity was diminished in porphyria cutanea tarda (PCT), acute intermittent porphyria (AIP), variegate porphyria (VP) and lead intoxication (Pb), remaining normal in erythropoietic protoporphyria (EPP). delta-Aminolaevulinate synthetase (ALA-S) activity was increased in PCT, AIP, VP and Pb showing no changes in EPP. It is suggested that a similar scheme, to that proposed for the control of ALA-S in Rhodopseudomonas spheroides and soybean callus, is also operating in animals.

Encapsulation of thiosulfate: cyanide sulfurtransferase by mouse erythrocytes.

Murine carrier erythrocytes, prepared by hypotonic dialysis, were employed in the encapsulation of several compounds including [14C]sucrose, [3H]inulin, and bovine thiosulfate:cyanide sulfurtransferase (rhodanese), a mitochondrial enzyme which converts cyanide to thiocyanate. Approximately 30% of the added [14C]sucrose, [3H]inulin, and rhodanese was encapsulated by predialyzed erythrocytes, and a decrease in the mean corpuscular volume and mean corpuscular hemoglobin was observed. In the encapsulation of rhodanese a recovery of 95% of the erythrocytes was achieved and an 85% equilibrium was established. The addition of potassium cyanide (50 mM) to intact, rhodanese-loaded erythrocytes containing sodium thiosulfate resulted in its metabolism to thiocyanate. These results establish the potential use of erythrocytes as biodegradable drug carrier in drug antagonism.

Genetic polymorphism of rhodanese from human erythrocytes.

Rhodanese of human erythrocytes was monomorphic in most populations tested, but was polymorphic in Ahtna and Upper Tanana Indians. This enzyme is under the control of a single autosomal locus.