Urinary signature of pig carcasses with boar taint by liquid chromatography-high-resolution mass spectrometry.

Boar taint is an offensive odour that can occur while cooking pork or pork products and is identified in some uncastrated male pigs that have reached puberty. It is widely held that boar taint is the result of the accumulation in back fat of two malodorous compounds: androstenone and skatole. The purpose of this study is to assess a mass spectrometry-based metabolomics strategy to investigate the metabolic profile of urine samples from pig carcasses presenting low (untainted) and high (tainted) levels of androstenone and skatole in back fat. Urine samples were analysed by LC-ESI(+)-HRMS. Discrimination between tainted and untainted animals was observed by the application of multivariate statistical analysis, which allowed candidate urinary biomarkers to be highlighted. These urinary metabolites were positively correlated to androstenone and skatole levels in back fat. Therefore, the study suggests that the measurement of these urinary metabolites might provide information with regard to androstenone and skatole levels in live pigs.

Identification of beta-glucuronidase-resistant diastereomeric glucuronides of 3-hydroxy-3-methyloxindole formed during 3-methylindole metabolism in goats.

Goats were jugularly infused with the pneumotoxin 3-methylindole (3MI; 15 mg/kg, 0.5 microCi/kg) dissolved in cremophor-EL to characterize the urinary metabolites of 3MI in a ruminant specie. Urine was collected for 36 hr after the beginning of a 2-hr infusion period, and 3MI metabolites were purified using reversed-phase HPLC. Goats excreted 3MI as at least 11 distinct metabolites. Metabolites were characterized using a combination of UV spectroscopy, 1H- and 13C-NMR spectroscopy, and negative-ion FAB/MS. Two of the metabolites (E1 and E2), representing approximately 30% of the urinary radioactivity, were unambiguously identified as diastereomeric glucuronides of 3-hydroxy-3-methyloxindole [HMOI; 3-(beta-D-glucosiduronic acid)-3-methyloxindole]. Glucuronide conjugates were investigated using enzymatic and chemical hydrolysis. These ethereal glucuronides were unique in that they were not readily hydrolyzable with bovine beta-glucuronidase, although one of the diastereomers was hydrolyzed sparingly by beta-glucuronidase from Helix pomatia. Treatment of the glucuronides with 6 M HCI for a 2-hr period liberated unconjugated HMOI. Treatment of each diastereomer with dilute acid (pH 3) or dilute alkali (pH 10) was ineffective at hydrolyzing the conjugates. Goats form HMOI from 3MI and extensively glucuronidate the metabolite before excreting it, as opposed to mice that do not conjugate HMOI before excretion. These ethereal glucuronic acid conjugates seem to be unique in that they are essentially resistant to beta-glucuronidase-catalyzed hydrolysis.

Identification of goat and mouse urinary metabolites of the pneumotoxin, 3-methylindole.

1. Urine from goats dosed i.v. with 3-methylindole (3MI; 15 mg/kg) or [methyl-14C] 3MI (15 mg/kg, 0.5 microCi/kg) contained at least 11 metabolites of 3MI. 2. Goat metabolized 3MI to sulfate conjugates of 4- or 7-hydroxy-3-methyloxindole, 5- or 6-hydroxy-3-methyloxindole, and 3,5- or 6-dihydroxy-3-methyloxindole; glucuronic acid conjugates of indole-3-carboxylic acid and 4- or 7-hydroxy-3-methyloxindole; and unconjugated 3-hydroxy-3-methyloxindole. Diastereoisomeric glucuronic acid conjugates of 3-hydroxy-3-methyloxindole were also identified in goat urine. 3. Urine from mice dosed i.p. with 3MI (400 mg/kg) or [ring-UL-14C] 3MI (400 mg/kg, 125 microCi/kg) contained at least six metabolites of 3MI. 4. Mice metabolized 3MI to glucuronic acid conjugates of 3,5- or 6-dihydroxy-3-methyloxindole, 5- or 6-hydroxy-3-methyloxindole, and indole-3-carboxylic acid; and unconjugated indole-3-carboxylic acid. Unconjugated 3-hydroxy-3-methyloxindole was identified in mouse urine in a previous report. 5. Both goats and mice metabolized 3MI to a mercapturate, 3-[(N-acetyl-L-cystine-S-yl)methyl]indole, which has been previously identified and was confirmed in this study. 6. 3-Methyloxindole was not identified in the urine of either goats or mice. 7. The major pathways of 3MI biotransformation in goats and mice is the formation of mono- and dihydroxy-3-methyloxindoles and their subsequent conjugation with glucuronic acid or sulfate. 8. There are no apparent qualitative differences in the biotransformation of 3MI between goats and mice that can account for their different sensitivities to 3MI-induced lung injury.