A metabonomic study of biochemical changes characteristic of genetically hypertensive rats based on (1)H NMR spectroscopic urinalysis.

Spontaneously hypertensive rats (SHR) provide a simple model for studying essential hypertension. Their genetic and metabolic features are of great interest because they may provide insights into the pathophysiological processes underlying essential hypertension. We have thus investigated the metabolic characteristics of SHR at various ages, covering the prehypertensive stage and the developmental phase of hypertension, using a (1)H nuclear magnetic resonance (NMR)-based metabonomic approach. Twenty-four-hour urine samples from the SHR and their age-matched normotensive control, Wistar-Kyoto rats, were analyzed using (1)H NMR spectroscopy, and the spectral data were subjected to principal components analysis (PCA) to find metabolic differences between the two strains. Consequently, it was possible to separate the urine samples between the two strains at any age ranging from 4 to 20 weeks in the principal component scores plots. The major spectral regions and signals (metabolites) contributing to the separation were picked up based on the loadings. Subsequently, the urinary excreted levels of metabolites highlighted by the PCA were compared based on the signal intensities corrected by urine volume and body weight. These investigations revealed the major metabolic changes characteristic of the SHR, which included differences in citrate, α-ketoglutarate, succinate, hippurate, phenylacetylglycine, p-cresol glucuronide, creatine, taurine, medium-chain dicarboxylates (tentative), unknown (δ 3.11), and the regions at 3.60, 3.64, 3.68 and 3.88 p.p.m. The results supported the occurrence of metabolic acidosis in the SHR in the period of prehypertension as well as rapidly rising blood pressure. In addition, the intestinal microfloral populations in the SHR were suggested to be altered in the developmental phase of hypertension.

LC-NMR identification of a novel taurine-related metabolite observed in (1)H NMR-based metabonomics of genetically hypertensive rats.

This paper describes the LC-NMR spectroscopic identification of a novel urinary endogenous metabolite responsible for the signals, which were found as major contributors to the separation between genetically hypertensive rats (SHRSP) and normotensive control rats (WKY) in previous NMR-based metabonomic studies. Urine samples from 26-week-old normotensive rats were analyzed by an LC-NMR system equipped with a reversed-phase column having high retention ability for polar compounds. (1)H NMR spectra were continuously obtained in the on-flow mode, and the retention times of the unassigned signals in question were determined. Various two-dimensional spectra were subsequently measured for the fraction containing the unassigned signals under the stop-flow mode, which enables for a long accumulation resulting in the enhancement of signal-to-noise ratios. The candidate compound obtained from these LC-NMR data was synthesized, and the NMR and mass spectra were compared with those of the LC-NMR fraction. The unknown metabolite was identified as succinyltaurine from these experiments together with standard addition experiments. This novel metabolite, which is characteristic of the normotensive rats, is very interesting because it is structurally related to hypotensive taurine, and not substantially detected in the genetically hypertensive rats, which excreted more taurine than the normotensive rats. The biological and pathophysiological significances of succinyltaurine remain to be investigated.

1H NMR-based metabonomic analysis of urine from young spontaneously hypertensive rats.

Spontaneously hypertensive rats (SHR) and their substrains are a useful model for studying essential hypertension which is a complex, polygenic, and multifactorial disorder. Their genetic and metabolic features are of great interest because they may provide insights into the mechanism of blood pressure regulation. We have compared urinary metabolic profiles of young SHR with those of their age-matched normotensive controls, Wistar Kyoto rats, using (1)H NMR-based metabonomics. Principal components analysis was applied to the NMR spectral data after data-reduced and normalized by the total integral or the creatinine integral. Consequently, a clear separation of urine samples between the two strains was observed in the principal components scores plot. The loadings plot from the data normalized by the creatinine integral showed that many metabolites such as citrate, alpha-ketoglutarate, and hippurate contributed to the separation, and the urinary levels of most metabolites used in this study, including these three, were lower in SHR than in Wistar Kyoto rats. These metabolic changes may be concerned with blood pressure regulation in SHR, although a relation to other strain differences cannot be ruled out. The present study suggests the usefulness of a (1)H NMR-based metabonomic approach using SHR in the field of hypertension research.

Investigations into biochemical changes of genetic hypertensive rats using 1H nuclear magnetic resonance-based metabonomics.

Genetically hypertensive rats provide a simple and accessible model for studying essential hypertension, which is a polygenic, heterogenous and multifactorial disease. Their genetic and metabolic features are of great interest because they may provide insight into the pathophysiological processes underlying essential hypertension. We have investigated the genetic influence on metabolic balance and metabolite excretion patterns in stroke-prone spontaneously hypertensive rats (SHRSP) with established hypertension using 1H NMR-based metabonomics. Urinary metabolite profiles for SHRSP and their age-matched normotensive controls, Wistar Kyoto rats, were acquired using 1H NMR spectroscopy. Principal components analysis was applied to these complex NMR data to facilitate differentiation and determine metabolic differences between urine samples collected from the hypertensive and normotensive rats. Consequently, it was possible to distinguish urine samples between the two strains in the principal components scores plot. The loadings plot showed that taurine, creatine and some unidentified metabolites resonating at around delta 2.48, 3.10 and 3.58 predominantly contributed to the separation. In SHRSP, the urinary levels of taurine and creatine were found to be higher and the intensities of the unknown signals much lower than those in the Wistar Kyoto rats. Although the pathophysiological significance of these components remains to be elucidated, this study suggests that 1H NMR-based metabonomics is a promising approach to provide new information on metabolic changes related to the pathophysiological processes of the genetically hypertensive rats.

Himeic acid A: a new ubiquitin-activating enzyme inhibitor isolated from a marine-derived fungus, Aspergillus sp.

A new ubiquitin-activating enzyme (E1) inhibitor, himeic acid A, was isolated from a culture of marine-derived fungus, Aspergillus sp. The structure was determined by spectroscopic analysis. The formation of an E1-ubiquitin (Ub) intermediate was 65% inhibited by himeic acid A at the concentration of 50 microM, while two new related compounds, himeic acids B and C, showed little inhibitory activity even at 100 microM.

Tetrapetalone A, a novel lipoxygenase inhibitor from Streptomyces sp.

A simple new assay was designed for lipoxygenase inhibitors. This assay was used to find the novel lipoxygenase inhibitor, tetrapetalone A (1). Tetrapetalone A (1), C26H33NO7, was isolated from Streptomyces sp. USF-4727 strain. Its planar structure was determined by spectroscopic evidence and by methylating with diazomethane to show the presence of a novel tetracyclic skeleton and a beta-D-rhodinosyl moiety. The stereochemistry of 1 was investigated by the coupling constant in the 1H-NMR spectrum, NOE correlations, modified Mosher's method and derivation. We have reported the structural elucidation of 1 in our previous paper. However, further investigation gave another structure for 1, which is described in this paper. Tetrapetalone A showed similar inhibitory activity against soybean lipoxygenase to the two well-known lipoxygenase inhibitors, kojic acid and NDGA, while methylated tetrapetalone A (2) showed little inhibitory activity, even at a concentration of 1 mM.