Phenolics characterization in propolis using HR-NMR and chemometrics

Propolis is a resinous substance produced by bees collecting resinous substances from exudates of different plants now widely used in the preparation of supplements and nutraceuticals, which is rich in well-known health-relevant phenolic compounds such as flavonoids and phenolic acids. Propolis extracts are very complex matrices difficult to study. Different analytical methods are usable to analyse propolis extracts and to obtain chemical fingerprint but at our knowledge NMR was not previously used for this purpose. This study aims to demonstrate that it is possible to use the 1H NMR for the simultaneous identification of phenolic compounds in propolis extracts, using appropriate tools for spectra pre-treatment and analysis. In this work as first step, twelve typical phenolic propolis compounds (apigenin, chrysin, galangin, kaempferol, quercetin, naringenin, pinocembrin, pinostrobin, caffeic acid, cinnamic acid, p-coumaric acid, ferulic acid) were considered as reference compounds and their presence in samples were verified by HPLC-MS, then NMR coupled with chemometric techniques were applied on the same samples. A simple 1H NMR sequence was used to obtain spectra of samples. Spectra were pre-treated by using an appropriate tool for spectra alignment and analysed by using a software for the study of spectra originated from complex matrices aiming to the recognition of the selected substances from their principal NMR signals. Sixty-five propolis samples were used to test the proposed identification procedure. Ten out of twelve considered compounds were identified as statistically significant in most of the samples. This work suggests that it is possible to efficiently use the 1H NMR, coupled with appropriate spectral analytical tools, for the simultaneous detection of phenolic compounds in complex matrices.