NUCLEAR-MAGNETIC-RESONANCE SPECTROSCOPIC AND PRINCIPAL COMPONENTS-ANALYSIS INVESTIGATIONS INTO BIOCHEMICAL EFFECTS OF 3 MODEL HEPATOTOXINS

H-1 NMR spectroscopy of urine combined with pattern recognition (PR) m ethods of data analysis has been used to investigate the time-related biochemical changes induced in Sprague-Dawley rats by three model hepa totoxins: alpha-naphthyl isothiocyanate (ANIT), D-(+)-galactosamine (G alN), and butylated hydroxytoluene (BHT). The development of hepatic l esions was monitored by conventional plasma analysis and liver histopa thology. Urine was collected continuously postdosing up to 144 h and a nalyzed by 600-MHz H-1 NMR spectroscopy. NMR spectra of the urine samp les showed a number of time-dependent perturbations of endogenous meta bolite levels that were characteristic for each hepatotoxin. Biochemic al changes common to all three hepatotoxins included a reduction in th e urinary excretion of citrate and 2-oxoglutarate and an increased exc retion of taurine and creatine. Increased urinary excretion of betaine , urocanic acid, tyrosine, threonine, and glutamate was characteristic of GalN toxicity. Both GalN and ANIT caused increased urinary excreti on of bile acids, while glycosuria was evident in BHT-and ANIT-treated rats. Data reduction of the NMR spectra into 256 integrated regions w as used to further analyze the data. Mean values of each integrated re gion were analyzed by principal components analysis (PCA). Each toxin gave a unique time-related metabolic trajectory that could be visualiz ed in two-dimensional PCA maps and in which the maximum distance from the control point corresponded to the time of greatest cellular injury (confirmed by conventional toxicological tests). Thereafter, the meta bolic trajectories changed direction and moved back toward the control region of the PR map during the postdose recovery phase. The combinat ion of urinary metabolites which were significantly altered at various time points allowed for differentiation between biliary and parenchym al injury. This NMR-PR approach to the noninvasive detection of liver lesions will be of value in furthering the understanding of hepatotoxi c mechanisms and assisting in the discovery of novel biomarkers of hep atotoxicity.