Distinguishing Vaccinium Species by Chemical Fingerprinting Based on NMR Spectra, Validated with Spectra Collected in Different Laboratories

Abstract

A method was developed to distinguish Vaccinium species based on leaf extracts using nuclear magnetic resonance spectroscopy. Reference spectra were measured on leaf extracts from several species, including lowbush blueberry (Vaccinium angustifolium), oval leaf huckleberry (Vaccinium ovalifolium), and cranberry (Vaccinium macrocarpon). Using principal component analysis, these leaf extracts were resolved in the scores plot. Analysis of variance statistical tests demonstrated that the three groups differ significantly on PC2, establishing that the three species can be distinguished by nuclear magnetic resonance. Soft independent modeling of class analogies models for each species also showed discrimination between species. To demonstrate the robustness of nuclear magnetic resonance spectroscopy for botanical identification, spectra of a sample of lowbush blueberry leaf extract were measured at five different sites, with different field strengths (600 versus 700 MHz), different probe types (cryogenic versus room temperature probes), different sample diameters (1.7 mm versus 5 mm), and different consoles (Avance I versus Avance III). Each laboratory independently demonstrated the linearity of their NMR measurements by acquiring a standard curve for chlorogenic acid (R2 = 0.9782 to 0.9998). Spectra acquired on different spectrometers at different sites classifed into the expected group for the Vaccinium spp., confirming the utility of the method to distinguish Vaccinium species and demonstrating nuclear magnetic resonance fingerprinting for material validation of a natural health product.

Evolution of Quantitative Measures in NMR: Quantum Mechanical qHNMR Advances Chemical Standardization of a Red Clover (Trifolium pratense) Extract

“…Despite the 10-year consistency of the findings, it is important to realize that repetition of any LC-based analysis requires revalidation due to its dependence on numerous instrument-related factors, particularly those related to (non)linearity of the detectors (UV, MS), the injector system, and the characteristics of the columns. This does not apply to qHNMR methods: once established, they can be rerun anytime later, provided the NMR instrument is validated per se, 39 and the qHNMR acquisition parameters are identical or demonstrated to be congruent between the different hardware and magnetic fields used. Another unique advantage of qHNMR is that additional analytes can be included even years later when peak assignments have become available, to study the same sample or the original FID of the qHNMR measurement.…”

Selection and characterization of botanical natural products for research studies: a NaPDI center recommended approach

“…Quantitative NMR (qNMR) is another technique which provides chemical ngerprinting capable of distinguishing species and authenticating botanical origins. 67 The relative intensities, retention times, and spectroscopic data obtained by chromatographic analysis can be used to putatively identify a sample. Small molecules are more stable than DNA, and they typically survive the processing involved with preparing botanical natural products.…”

“…Methods can be validated both within a single laboratory (intralaboratory validation) 95,194 or across multiple laboratories (interlaboratory validation). 67,90 The International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) provides general guidelines for validation of analytical methods, 195 and more specic guidelines and denitions related to validation of constituents of botanicals have been draed by a subcommittee of AOAC INTERNATIONAL. 13,193 When selecting a method for analysis of botanicals, we recommend reviewing multiple published methods (if available), with particular attention to those that have been validated.…”

Vaccinium angustifolium (lowbush blueberry) leaf extract increases extravillous trophoblast cell migration and invasion in vitro

“…Vaccinium extracts, including those from V. angustifolium, are a rich source of phenolic compounds (Harris et al, 2007;Hicks et al, 2012;Markus et al, 2014;McIntyre et al, 2009) and have been shown to influence cell migration and invasion using in vitro models of cancer metastasis (Adams et al, 2010;Matchett, MacKinnon, Sweeney, Gottschall-Pass, & Hurta, 2005), in vitro models of endothelial cell migration and invasion (Tulio et al, 2012), and animal models of wound healing (Nayak et al, 2010). In this study, the .…”