Two‐dimensional correlated spectroscopy (COSY) of intact frog muscle: Spectral pattern characterization and lactate quantitation

Abstract

Two-dimensional scalar-correlated 1H spectra (COSY) were recorded from a model solution containing varying amounts of lactate and a fixed concentration of carnosine as the reference compound. The integrated volume of the lactate methyl to methine cross peak in the magnitude-calculated spectra showed a linear increase with respect to lactate concentration. Accordingly, the COSY cross-peak integral volume should be a useful parameter for the quantitative study of metabolism in intact tissue. We have tested this assumption in excised gastrocnemius frog muscle. Lactate, carnosine, and creatine/phosphocreatine cross peaks have been assigned in the spectrum of summer frog muscle. The change in concentration of lactate and phosphorylated compounds upon anaerobic storage of excised frog muscle has been followed by interleaving one-dimensional (1D) 1H and 31P spectra with 2D homonuclear 1H COSY spectra. The lactate concentration in the excised muscle derived from COSY agreed well with that determined from muscle extracts. Qualitative changes in the COSY pattern of the excised muscles under caffeine-induced contracture were detected. Furthermore, seasonal variations were observed in the COSY spectral pattern: winter frog muscle was found to differ from summer frog muscle by the appearance of cross peaks from phosphodiesters and mobile fatty acids.

in vivo identification and monitoring of changes in rat brain glucose by two‐dimensional shift‐correlated 1h nmr spectroscopy

“…A&s et al ( 11 ) have shown that the volume of the cross peaks in COSY spectra measured under homogeneous B1 field was proportional to the concentration of the product. To verify the reliability of this relationship in inhomogeneous B 1 field conditions we have constructed a calibration curve relating cross-peak volume ratios of glucose H,-H2 to total creatine verses a range of glucose concentrations ( 1, 5, 10, 15, and 20 mM) from phantom's solutions containing 50 m M KC1 and 10 m M total creatine.…”

“…The time course of the quantitative changes in intracerebral glucose level was measured on the 2D spectra from the changes in the volume of the HI-H2 glucose cross peaks. The results (means k SEM) are expressed as the ratio of the volume of the correlation signal to the reference intensity of the Cr-PCr resonance, measured on the diagonal at 3.04 ppm (11). For each metabolite, all data were normalized to the basal level mean value taken as 100% (at time t = 0).…”

“…This report describes the use of 2D COSY NMR as a technique to resolve and identify the cerebral glucose resonance under basal conditions and to follow the changes in brain glucose levels of rats after an intravenous injection of glucose. Information on the concentration of cerebral glucose can be obtained by volume integration of its cross peak ( 11 ). In 2D COSY-like spectra, the hydrogens contained in molecules give a set of cross peaks that shows scalar couplings characteristic of the structure of the molecules.…”

Two‐dimensional spectra of intact tissue: Homonuclear Hartmann‐Hahn spectroscopy provides increased sensitivity and information content as compared to COSY

“…The model solution, perchloric acid (PCA) extract, and tissue samples were prepared as described in ( 19). Grass frogs ( R a m pipiens leopard) were purchased from Charles D. Sullivan (Nashville, TN).…”

“…In this communication we compare the performance and information content of 2D homonuclear Hartmann-Hahn spectroscopy ( HOHAHA) with 2D doublequantum-filtered COSY (DQFCOSY) spectroscopy (19) in studies of metabolites in excised frog gastrocnemius muscle in the resting state. The results show the advantages of HOHAHA spectroscopy for the detection of metabolite patterns in intact tissue.…”