Nuclear magnetic resonance spectroscopic and principal components analysis investigations into biochemical effects of three model hepatotoxins.
1H NMR spectroscopy of urine combined with pattern recognition (PR) methods of data analysis has been used to investigate the time-related biochemical changes induced in Sprague-Dawley rats by three model hepatotoxins: alpha-naphthyl isothiocyanate (ANIT), d-(+)-galactosamine (GalN), and butylated hydroxytoluene (BHT). The development of hepatic lesions was monitored by conventional plasma analysis and liver histopathology. Urine was collected continuously postdosing up to 144 h and analyzed by 600-MHz 1H NMR spectroscopy. NMR spectra of the urine samples showed a number of time-dependent perturbations of endogenous metabolite levels that were characteristic for each hepatotoxin. Biochemical changes common to all three hepatotoxins included a reduction in the urinary excretion of citrate and 2-oxoglutarate and an increased excretion 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 excretion of bile acids, while glycosuria was evident in BHT- and ANIT-treated rats. Data reduction of the NMR spectra into 256 integrated regions was used to further analyze the data. Mean values of each integrated region were analyzed by principal components analysis (PCA). Each toxin gave a unique time-related metabolic trajectory that could be visualized 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 metabolic trajectories changed direction and moved back toward the control region of the PR map during the postdose recovery phase. The combination of urinary metabolites which were significantly altered at various time points allowed for differentiation between biliary and parenchymal injury. This NMR-PR approach to the noninvasive detection of liver lesions will be of value in furthering the understanding of hepatotoxic mechanisms and assisting in the discovery of novel biomarkers of hepatotoxicity.
Beckwith-Hall
B M
BM
Department of Chemistry, Birkbeck College, University of London, Gordon House, 29 Gordon Square, London WC1H 0PP, U.K.
Nicholson
J K
JK
Nicholls
A W
AW
Foxall
P J
PJ
Lindon
J C
JC
Connor
S C
SC
Abdi
M
M
Connelly
J
J
Holmes
E
E
D015058:1-Naphthylisothiocyanate / Q000737:chemistry* / Q000633:toxicity; D000818:Animals; D002084:Butylated Hydroxytoluene / Q000737:chemistry* / Q000633:toxicity; D005688:Galactosamine / Q000737:chemistry* / Q000633:toxicity; D008099:Liver / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D051381:Rats; D013329:Structure-Activity Relationship
D002084:Butylated Hydroxytoluene; D015058:1-Naphthylisothiocyanate; D005688:Galactosamine