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We have previously reported an association of impaired S-oxidation with primary biliary cirrhosis. In order to confirm and further define this relationship, we retested S-oxidation capacity via three metabolic pathways and sulphation capacity via a fourth pathway. Metabolism of S-carboxymethyl-L-cysteine is polymorphic -20% of healthy individuals being poor S-oxidisers. We found 26% with primary biliary cirrhosis were poor S-oxidisers, compared with 36% with other liver disease and 25% of healthy controls. Differences were not statistically significant. S-oxidation of ranitidine is dependent upon flavin mono-oxygenases. We showed a non-significant trend toward less S-oxide in primary biliary cirrhosis and other liver disease, compared with healthy controls, with no significant difference between disease groups. Conversion of cysteine to sulphate depends predominantly on cysteine dioxygenase. Impaired activity may be reflected by decreased plasma sulphate and elevated cysteine. We found that the plasma cysteine: sulphate ratio was significantly elevated not only in primary biliary cirrhosis (p < 0.0001), but also in other liver disease (p < 0.0001), compared with healthy individuals. Sulphation capacity was studied by analysing paracetamol metabolism. Paracetamol sulphate and sulphate: glucuronide ratio were reduced in primary biliary cirrhosis compared with normal individuals, (p < 0.05). A trend towards less sulphate in primary biliary cirrhosis compared other liver disease was not significant (p = 0.42). We conclude that although sulphation and some sulphoxidation pathways are impaired in primary biliary cirrhosis, we can currently find no evidence to substantiate the hypothesis that primary biliary cirrhosis is a disease specifically associated with poor S-oxidation, as assessed via these metabolic pathways.

Cysteine and its metabolites cysteine sulphinic acid (CSA) and taurine (TA) were shown to be toxic to human and rat neuronal cell lines. However, the mechanisms of action of CSA and TA appeared to be different. As MND/ALS patients have high plasma and CSF cysteine levels, they may have increased levels of neurotoxins in vivo, since cysteine was more toxic than CSA in the assay system used. Cysteic acid, homocysteic acid, BMAA and BOAA were also toxic to the cell lines used.

1. Fasting levels of plasma cysteine, plasma sulphate and the plasma cysteine/sulphate ratio were measured in patients with primary biliary cirrhosis and compared with those in patients with other liver disease, general intensive therapy unit patients and healthy subjects. 2. Plasma cysteine was significantly elevated in patients with primary biliary cirrhosis (median 0.364 nmol/mg of protein, P < 0.0001) and patients with other liver disease (median 0.445 nmol/mg of protein, P < 0.0001), compared with healthy control subjects (median 0.125 nmol/mg of protein) and increased progressively with the severity of liver disease. Plasma cysteine was also elevated in intensive therapy unit patients (median 1.564 nmol/mg of protein) compared with healthy control subjects (P < 0.0001) and patients with other liver disease (P < 0.0001). 3. Plasma sulphate was reduced significantly only in patients with primary biliary cirrhosis (median 0.822 nmol/mg of protein) compared with healthy control subjects (median 1.37 nmol/mg of protein, P < 0.05). There was no significant difference in plasma sulphate between disease groups. 4. The plasma cysteine/sulphate ratio was significantly elevated in patients with primary biliary cirrhosis (median 0.448, P < 0.0001) and patients with other liver diseases (median 0.394, P < 0.0001) compared with healthy control subjects (median 0.095). The ratio was also elevated in intensive therapy unit patients (median 1.650, P < 0.0001) compared with healthy control subjects and liver disease groups (P < 0.0001). 5. In conclusion, plasma cysteine rises in primary biliary cirrhosis and other forms of liver disease. This effect is not specific to liver disease, since cysteine is elevated in an heterogeneous group receiving intensive care. Impairment of trans-methylation and trans-sulphuration pathways does not explain the finding of increased plasma cysteine. Since cysteine is elevated in non-hepatic disease, it may reflect the effect of muscle breakdown and the catabolic state. Impaired activity of cysteine dioxygenase and impaired mitochondrial function may be contributory, but this requires further study. These metabolic changes may reflect progressively diminished detoxification capacity within the liver and other tissues.

Analysis of plasma from MND/ALS patients has shown no significant differences in metabolism of cysteine derivatives, although a sub-set of the population has raised glutamate values. Cysteine dioxygenase was found to have reduced activity in vitro, consistent with previous findings of a high plasma cysteine/sulphate ratio.

The wide individual variations seen in all metabolic pathways which have been examined largely explain differences in drug dose response and toxicity. In some cases, distinct sub-sets occur where the enzyme activity is markedly different from that in the rest of the population (genetic polymorphism). Studies on isolated enzymes and use of restriction fragment length polymorphisms have in some cases helped to elucidate the relevant molecular biology. However, the lack of certain concordance between genotype and phenotype has shown that many questions remain unanswered. As more information becomes available at the molecular level, it may be easier to gain an understanding of the endogenous biochemistry which underlies the human condition. Either genotyping or phenotyping may then be used to predict potential drug side-effects or clinical outcomes.

Introduction As one of the possible causes of inflammatory bowel disease (IBD), the microbiota in the human gut has been widely studied. Sulphate-reducing bacteria (SRB) have long been considered as candidate pathogens because their metabolic product, sulphide, through its toxic properties, damages the colonic epithelium. However, no specific SRB has been identified as a pathogen to IBD, and even their abundance in the human gut remains controversial. This project compares the amount of faecal SRB in healthy subjects with that in patients either with Crohn's disease (CD) or ulcerative colitis (UC), and traces the change, if any, after treatment in each patient. Methods Faecal samples were collected from 18 healthy subjects, 18 CD patients and 14 UC patients. CD patients were treated with elemental diet and UC patients received conventional treatment. A suspension of the faecal matter from each patient both before and after treatment was inoculated into a sulphate-rich growth medium, Postgate B. The abundance of SRB was gauged according to the amount of black precipitate (ferrous sulphide) formed by SRB. Results Contrary to expectation 88% of healthy subjects carry SRB that produce high levels of sulphide in vitro, despite its cytotoxicity in vivo. In contrast, this carriage rate is low (45%) in CD patients at their initial presentation. After taking elemental diet, the carriage rate of active SRB did not change despite a significant fall in the total amount of gut bacteria. In UC patients, who are often characterised with mucus depletion, the carriage rate of active SRB is significantly lower (31%, p

OBJECTIVE: To determine whether genetic factors associated with established rheumatoid arthritis could, in combination with rheumatoid factor, predict the development of radiological erosions in patients with early symmetrical (rheumatoid-like) arthritis. DESIGN: Prospective study. SETTING: Teaching hospital, early arthritis clinic. SUBJECTS: Forty nine patients with symmetrical polyarthritis attending the early arthritis clinic. MAIN OUTCOME MEASURES: Conserved sequence of DR beta third allelic hypervariable region, sulphoxidation capacity, rheumatoid factor, and development of radiologically determined bone erosions. RESULTS: None of the 49 patients had radiological erosions at presentation but 25 developed these by four years. Patients with the conserved class II major histocompatibility complex (third allelic hypervariable of DR beta 1) genes associated with rheumatoid arthritis had a relative risk for the development of erosions of 1.9 (95% confidence interval 0.8 to 4.5). For poor sulphoxidation the risk was 2.5 (1.1 to 5.6) and for the presence of rheumatoid factor 1.8 (0.9 to 3.7). Of the 33 patients who had two or three of these risk factors, 24 developed erosions, with a relative risk of 11.6 (1.7 to 78.5) compared with only one of the 16 individuals with no or one risk factor. CONCLUSIONS: This preliminary study shows that by using these stable markers it is possible to make clinically useful predictions of outcome in patients with early symmetrical inflammatory arthritis.

A minority of individuals have an impaired ability to oxidize sulphur. An increased prevalence of this poor S-oxidation capacity has been observed in patients with established rheumatoid arthritis (RA). The role of this enzyme system in the development of disease was studied by testing the S-oxidation capacity of 54 patients with recent onset symmetrical poly-arthritis, who were followed up at 1 and 4 years. Those patients with persistent disease at 1 year had a prevalence of poor S-oxidation of 69%. At 4 years 74% of those with a diagnosis of RA were poor S-oxidizers compared to 31% of those who were asymptomatic (P less than 0.01). A defective ability to oxidize sulphur appears to predispose to persistent clinical disease.

The hepatotoxicity induced by administration of ethionamide and thionicotinamide (TNA) was shown to be decreased by pre-administration of methimazole (MMI). Pre-administration of MMI was also shown to decrease the levels of excretion of TNA S-oxide. This indicates that thioamide S-oxidation, mediated by the flavin-containing mono-oxygenase, may be linked to the initiation of hepatotoxicity induced by these thioamides. SKF-525-A, the cytochrome P-450 inhibitor, did not affect either thioamide-induced toxicity or levels of excretion of TNA S-oxide; however, the role of the P-450 isoenzymes cannot be totally ruled out.

Xenobiotic enzymic systems have been studied in Parkinson9s disease. Platelet monoamine oxidage-B activity is increased by the use of phenylethylamine but decreased by the use of dopamine as substrate. Enzymes involved with sulfur oxidation and methylation are underactive.

1. Low dose (500 mg) paracetamol (acetaminophen) was administered to patients with rheumatoid arthritis (RA) and to age-matched healthy controls and to hospital controls. 2. At this dose level, patients with RA excreted decreased amounts of paracetamol sulphate (controls means 11.3 +/- 5.1, 10.6 +/- 5.9; RA mean 3.02 +/- 3.7). This difference is statistically significant (P less than 0.001). 3. The mean ratio of excretion of paracetamol sulphate/paracetamol glucuronide was 5.6 +/- 12.1, 5.3 +/- 10.7 in controls but 2.1 +/- 2.7 in RA patients (P less than 0.001). 4. Patients with RA appear to have less capacity for excreting paracetamol as non-toxic conjugates and may be more susceptible to paracetamol toxicity, especially on chronic dosage.

The fate of the three isomeric benzoic acid-substituted monochlorobenzanilides has been studied in the rat after their oral administration. The excretory profile of all three isomers was similar with over half of the dose being excreted in the 0-24 hour urine and between 83-87% being recovered during the three days following administration. The most prominent route of metabolism was that of 4'-hydroxylation on the unsubstituted aniline moiety followed by glucuronic acid conjugation.

The metabolism of the thioamide, thionicotinamide, in the male Wistar rat is described. Thionicotinamide was shown to be metabolised to inorganic sulphate, thionicotinamide S-oxide and also N1-methylnicotinamide, which arises from N-methylation of nicotinamide via desulphuration of the parent thioamide. At 24 hours from dosing about 35% of the dose had been excreted as inorganic sulphate and 20.5% as the S-oxide metabolite. Three metabolites, detected by their fluorescence under UV light, have not been positively identified but are postulated to be pyridones, resulting from ring oxidation, by analogy with other thioamides. S-Oxidation is an important metabolic pathway as it may give rise to toxic intermediates.

Using 5 methods, we assessed the ability of patients with a clinical diagnosis of Alzheimer9s disease (AD) to handle xenobiotics. Patients with AD, compared with controls, have reduced sulfoxidation of the probe drug S -carboxymethyl-l-cysteine; they also form less of the sulfate conjugate of acetaminophen. In addition, they have lower activity of the enzyme thiolmethyltransferase. In contrast, the capacity to oxidize debrisoquin and to acetylate sulfamethazine was normal. These findings suggest that a major risk factor for the development of AD is a skewed capacity for xenobiotic metabolism especially of compounds containing sulfur.

Penicillamine shows some structural similarities to carbocysteine. The ability to oxidize carbocysteine, i.e., the sulfoxidation status, shows a bimodal distribution in the general population. In this study, sulfoxidation status was determined in 50 of 60 rheumatoid arthritis patients receiving penicillamine. We found that poor sulfoxidation status, compared with good sulfoxidation status, was associated with a 3.9 times higher incidence of toxicity.

S-carboxy-L-methylcysteine was used to assess the activity of the S-oxidation pathway of sulphur metabolism in 35 patients with systemic lupus erythematosus (SLE); 25 (71%) showed impaired sulphoxidation and 21 (60%) produced virtually no sulphoxides, compared with 17 (36%) and 2 (4%), respectively, of 47 healthy controls. The substrate/product ratio of cysteine oxygenase (plasma cysteine/sulphate) was significantly higher in SLE patients than in controls (median [interquartile range] 362 [224-588] vs 65 [44-111]; p less than 0.00001). The alternative pathway of sulphur metabolism, S-methylation, catalysed by thiolmethyltransferase, was not impaired in the SLE patients. There is a biochemical difference in sulphur metabolism between SLE and rheumatoid arthritis, since both pathways are impaired in the latter disorder.

The association between sulphoxidation capacity and sodium aurothiomalate toxicity was investigated in 65 patients undergoing treatment for rheumatoid arthritis. Of those showing side-effects, 30/37 (81%) were also poor sulphoxidizers compared with only 9/28 (32%) in the group not displaying adverse reactions. A patient with poor sulphoxidation had a nine-fold greater risk of developing toxicity. The prior measurement of sulphoxidation capacity may help to identify those patients most susceptible to the deleterious effects of this drug.

The elimination of 2-halogenobenzanilides has been studied in the rat, over 75% of the oral dose being excreted in 3 days. The higher the molecular weight of the compound, the more important was the faecal elimination route. No such association was seen for biliary secretion.

The metabolism of cis-2,5-dimethylpyrrolidine-1-carboxanilide was studied in rats, rabbits, guinea-pigs and mice. The major metabolite in all species was cis-4′-hydroxy-2,5-dimethylpyrrolidine-1-carboxanilide, both free and as glucuronide and sulphate conjugates. About 95% of the compound was absorbed from the gut; over a 3-day period, about 50% of the administered dose was excreted in urine and about 27% in faeces.

1. The pungent urinary odour produced by certain individuals within a few hours of eating asparagus has been shown to be due to a combination of up to six sulphur-containing alkyl compounds identified as methanethiol, dimethyl sulphide, dimethyl disulphide, bis-(methylthio)methane, dimethyl sulphoxide and dimethyl sulphone. 2. The possible roles of S-methylmethionine and asparagusic acid as precursors of these odorous substances are discussed in relation to the known chemistry of the vegetable.

The metabolism of an oral dose of S-carboxymethylcysteine (SCMC) has been studied in man; the principal urinary component after 24 h was the unchanged compound (29.4-83.7%, 20 subjects). Markers variation was found between individuals; SCMC sulphoxide, S-methylcysteine, N-acetyl-S-carboxymethylcysteine, N-acetyl-S-methylcysteine with the corresponding sulphoxides and dicarboxymethylsulphide were also found as metabolites.

1. The excretion and metabolism of 35S-D-penicillamine (200 mg orally) was studied in four healthy male volunteers. 2. The major route of excretion was the urine with 82% of the administered dose being voided over three days (73% during the first 24 h); the remainder was excreted in the faeces. 3. Metabolism occurred via the pathways of S-methylation, N-acetylation and disulphide formation. Extensive degradation of the molecule was observed with the production of large amounts of inorganic sulphate.

The metabolism of benodanil (2-iodobenzanilide) was studied in rats following an oral dose of 150 mg benodanil kg−1 body weight. The major 24-h urinary metabolite was found to be the 4′-hydroxy derivative, both free (≈ 5%) and as the glucuronide (≈ 4%) and sulphate (≈ 4%) conjugates. Over a 6-day period, about 16% of the administered dose was excreted in the urine and about 80% in the faeces. After dosing with [14C]- benodanil, blood radioactivity levels were highest 30 min after dosing, with small broader peaks at 4 and 7 h, while biliary activity levels rose slowly to a maximum about 10–12 h after the dose, some 16% being excreted in 24 h as the glucuronide conjugate of the 4′-hydroxy derivative.

An impaired sulfoxidation pathway has been implicated in the pathogenesis of chlorpromazine-induced hepatotoxicity. Since some patients with chronic chlorpromazine-induced cholestasis may have features of primary biliary cirrhosis, we studied the ability to sulfoxidate the amino acid analogue S-carboxymethyl-cysteine in 44 patients with primary biliary cirrhosis and in two control groups--one without liver disease and one with a variety of liver diseases other than primary biliary cirrhosis. Poor sulfoxidation was observed in 84 percent of the patients with primary biliary cirrhosis, as compared with 24 percent of patients with other liver diseases and 22 percent of normal controls (P less than 0.0005 for both comparisons). Poor sulfoxidation did not correlate with the degree of hyperbilirubinemia or histologic severity of liver disease in any of the groups studied. There was an inverse correlation with age only in the patients with primary biliary cirrhosis (r = -0.44, P less than 0.001). Liver transplantation was performed in six of the patients and improved sulfoxidation in five; in the four with primary biliary cirrhosis, sulfoxidation improved from poor to good or intermediate. We conclude that poor sulfoxidation is closely associated with primary biliary cirrhosis but not with the other liver diseases we studied.

1. After administration of 2,4-diaminotoluene to rabbits, rats and guinea-pigs, phenolic metabolites are excreted, the major component being 5-hydroxy-2,4-diaminotoluene in all species investigated. Acetylated aminophenols and glucuronide conjugates were also found. 2. Rabbits, which produced the largest amounts of free amino-phenols, also had the highest levels of methaemoglobin in vivo.

Debrisoquine, carbocysteine, and paracetamol were selected as safe drugs to investigate the ability of the liver's microsomal system to oxidise carbon, oxidise sulphur, and conjugate sulphate in patients with motor neuron disease (MND), other hospital patients, and healthy volunteers. Subjects with poor sulphur-oxidising and sulphur-conjugating activities were heavily over-represented in the MND group.

A conjugate isolated from urine of human volunteers after an oral dose of S-carboxymethyl-L-cysteine was characterized as a carboxylic acid ester glucuronide. Of the 166 volunteers investigated, 61 gave no detectable drug glucuronide (less than 0.5% administered dose); the remaining 105 showed a unimodal distribution of drug glucuronide excretion accounting for 0.5-11.5% (mean 4.1%, median 3.4%) of the total dose recovered in the 0-8 h urine. No significant variation in subject age, sex or urinary pH was observed between the two groups, but those not excreting urinary glucuronide had significantly higher 0-8 h urine volumes (P less than 0.001), although notable exceptions occurred.

1. The metabolism of the pesticide, 3,5-dibromosalicylanilide, has been studied following oral administration to rats. 2. This compound is more toxic than other brominated salicylanilides, some fatalities being recorded at an oral dose of 100 mg/kg. 3. 3,5-Dibromosalicylanilide is well absorbed, and excreted in urine and bile as sulphate and glucuronide conjugates of the parent compound and of several phenolic metabolites. Radioactivity was present in tissues (blood, kidney, liver, fat) six days after dosing with 3,5-dibromosalicyl[14C]anilide. 4. Sex-differences occur, both in excretion route and in the proportions of urinary metabolites.

The major soluble metabolite of carboxin (2,3-dihydro-6-methyl-5-phenylcarbamoyi-1,4-oxathiin) in barley seedlings and mature plants was identified as the-hydroxy phenyl derivative. This phenol was also bound to the lignin fraction. The distribution of carboxin and its metabolites in seedlings and mature plants grown from treated seed was studied; maximum residues were found in the three oldest internodes, with traces in the roots and flowering heads. Treated plants had altered chlorophyll/carotenoid ratios, and raised levels of some free amino acids.

1. The metabolism of two pesticides, 3,4',5-tribromosalicylanilide (TBS) and 4',5-dibromosalicylanilide (DBS) has been studied after oral administration to rats. 2. Approximately 65% of the dose of TBS is absorbed, and then excreted as glucuronide and sulphate conjugates of two hydroxylated metabolites. One of these has been identified as 4'-hydroxy-3,5-dibromosalicylanilide, while evidence suggests that the other metabolite is 5-hydroxy-3,4'-dibromosalicylanilide. 3. In contrast, only 11% of the dose of DBS is absorbed; it is then excreted mainly as glucuronide and sulphate conjugates of the parent molecule.

Radiolabelled [35S]-D-penicillamine was administered orally to DA-strain rats. After 72 hours approximately 65% of the dose was excreted with no significant differences between male and female animals. The major urinary product was inorganic sulphate with small amounts of D-penicillamine. Penicillamine disulphide, penicillamine-cysteine, S-methyl penicillamine and N-acetyl penicillamine were also found as metabolites. The female rat excreted significantly less sulphate (P less than 0.1) and more penicillamine disulphide (P less than 0.01) than the male. The residual radioactivity was found in the carcass, with slight concentration in the gut, skin, kidney, bladder and liver.

The excretion and metabolism of radiolabelled S-methyl-L-cysteine (150 mg orally) was studied in three male volunteers. The major route of excretion was the urine with 55.9% of the administered radioactivity being voided over three days (33.3% during the first 24 h). Faecal excretion was relatively unimportant (c. 1.4%). The remaining radioactivity was either slowly excreted in the urine over the next 21 d (35S-labelled compound) or exhaled (14C-labelled compounds). Metabolism occurred via the pathways of S-oxidation, N-acetylation and deamination. Extensive degradation of the molecule was observed with the production of large amounts of inorganic sulphate and CO2. The options for the sequence of metabolic degradation of the amino acid derivative are discussed.