Nmr suite professional

Initial peak assignments relied on database
searches,^{28 (link)−30 (link)} including the human metabolome database (HMDB)^{28 (link)} and the biological magnetic resonance data bank
(BMRB).^{29 (link)} However, unambiguous identification
of the coenzymes
necessitated the development of a new chemical shift database consisting
of the coenzymes and other compounds in solutions at concentrations
similar to their levels in tissue (Table 1). Spectral peaks for all the coenzymes were
identified using this database along with peak multiplicity, J coupling measurements, and the comprehensive analyses
of 2D DQF-COSY and TOCSY spectra. The coenzymes thus identified were
further confirmed by spiking experiments using authentic compounds.
Chenomx NMR Suite Professional (version 5.1; Chenomx Inc., Edmonton,
Alberta, Canada) was used to quantify the coenzyme peaks. Chenomx
allows fitting spectral lines using the standard metabolite library
for 800 MHz ¹H NMR spectra, and in particular, the determination
of concentrations. Since the proximity of chemical shift values for
signals from multiple compounds resulted in the software providing
multiple library hits for the same peak, the correct identification
of coenzymes’ peaks relied on the newly established peak assignments.
Peak fitting with reference to the internal TSP signal enabled the
determination of absolute concentrations of the coenzymes.