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 ( 1 D) 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 caffeineinduced 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.