Modified PSYCHE NMR - the possibility for the direct semi-quantitation of components in mixtures

The analysis of natural products using non-chromatographic methods is a domain of great interest, since the complexity of plant extract mixtures and the issue of co-eluting impurities, are not easily addressed by High-Performance Liquid Chromatography (HPLC). In contrast to HPLC, quantitative NMR spectroscopy supports both accurate purity determination of compound constituents, in addition to the identification and quantification of impurities. However, with a limited spectral range, a 1H NMR spectrum may be complicated by the many overlapping signals. The latter can be alleviated in PSYCHE NMR which suppresses the effects of homonuclear coupling, turning multiplet resonance signals into singlet peaks. In conjunction with qNMR, a new in-house homo-decoupled NMR spectroscopic technique, based on Pure Shift Yielded by Chirp Excitation (PSYCHE), proved to be a beneficial tool in natural product mixture analysis and crucially, component quantitation. As part of a toolkit involving the 2D techniques, COSY, HSQC and in some cases HMBC, our optimised PSYCHE NMR parameter set was able to improve the process of identification and quantification of components found in plant extracts. The PSYCHE NMR allowed clearer identification of components which normally would have overlapping multiplets, and importantly allowed their (semi)quantitation through the simplification of the 1D spectrum.

Indeed, a series of commonly occurring biologically relevant compounds were analysed using internally referenced PSYCHE NMR with Excitation Sculpting (ES), and the accuracy/precision compared favourably with that achieved using V Abstract conventional 1H qNMR. In the work described herein, the effect of chemical shift, coupling constants, swept frequency pulse angle, excitation sculpting and the impact of signal overlap in mixtures was evaluated in terms of statistically significant variation between the two techniques. It is shown that suitably optimised PSYCHE NMR with ES represents a potentially reliable method for the semi-quantitation of mixtures of compounds, whose spectroscopic signals overlap in conventional qNMR analysis, rendering accurate quantitation difficult with that technique. The outcome of this work is of particular relevance for complex mixtures of natural products. Of particular note is the effect of the excitation sculpting block introduced to supress baseline artefacts and the technical implementation of the Saltire chirp pulse.