NMR spectroscopic studies on the haemolymph of the tobacco hornworm, Manduca sexta: assignment of 1H and 13C NMR spectra

High resolution 1 H nuclear magnetic resonance (NMR) spectroscopy has been used to investigate the low molecular weight metabolite composition of the whole haemolymph of 5th instar larvae of the Tobacco Hornworm, Manduca sexta . This technique provides a rapid, multicomponent profile of a wide range of intermediary metabolites. Assignment of the 1 H NMR resonances in the 1-dimensional spectrum has been facilitated by the use of high frequency 2-dimensional NMR techniques, including J-resolved spectroscopy, 1 H– 1 H total correlation spectroscopy and 1 H– 13 C heteronuclear correlation spectroscopy. Amongst the biochemically important metabolic intermediates that were detected in whole haemolymph were trehalose, glucose, alanine, lactate, choline and betaine. In total, it was possible to simultaneously detect and potentially quantify 19 endogenous metabolites. These studies indicate that NMR spectroscopy of the whole larval haemolymph may offer the potential for evaluation of whole organism biochemical status in relation to physiological or environmental stress.