RESEARCH ARTICLE Important issues in plant tissues analyses by HR‐MAS NMR Igor S. Flores 1 | Bruno C.B. Martinelli 2 | Vinicius S. Pinto 2 | Luiz H.K. Queiroz Jr 2 | Luciano M. Lião 2 1 Instituto Federal de Goiás, Luziânia, GO, Brazil 2 Instituto de Química, Universidade Federal de Goiás, Goiânia, GO, Brazil Correspondence Luciano M. Lião, Instituto de Química, Universidade Federal de Goiás, 74690‐900, Goiânia, GO, Brazil. Email: lucianoliao@ufg.br Abstract Introduction: High‐resolution magic‐angle spinning nuclear magnetic resonance (HR‐MAS NMR) spectroscopy enables the analysis of the metabolic profile of plant and animal tissues under close to natural conditions, as well as of other heteroge- neous natural or synthetic materials. Neither sample pretreatment is required after fragmentation nor powdering of the sample before insertion into the rotor. However, the efficiency of the method depends strongly on the sample preparation, rotor inser- tion procedure, and analysis conditions. Objective: To identify some of the variables that affect the spectral data and to pro- pose solutions that minimise their impact on the quality of the analyses and results. Methods: Dried plant tissues were powdered, weighed, and homogenised in a 50 μL rotor with an optimised volume of deuterated solvent and sample in order to prevent material from escaping during spacer insertion, avoiding variations in mag- netic susceptibility. Factors affecting the quality of HR‐MAS NMR analysis such as particle size, sample and solvent amounts, solvent polarity, swelling time, rotor manip- ulation and pulse sequence setting were evaluated. Results: A strong correlation was observed between the signal area and the particle size of the powdered sample. The spectral profile varied depending on the deuterated solvent used. An incubation period was necessary to achieve adequate swelling of the sample and to ensure good data reproducibility. Proper sealing of the rotor, number of cycles and τ time on cpmgpr1d pulse sequence were found to affect the signal areas. Conclusion: The study highlights the need for standardised sample preparation and instrumental setup protocols in order to achieve high reproducibility and obtain reliable data from HR‐MAS NMR analyses. KEYWORDS analyses protocols, HR‐MAS NMR, phytochemical analysis, phytotherapeutic 1 | INTRODUCTION High‐resolution magic‐angle spinning nuclear magnetic resonance (HR‐MAS NMR) spectroscopy is a useful tool used as a fast and easy way that permits direct chemical insight of the metabolites in hetero- geneous samples (i.e. swollen samples), such as vegetable and animal tissues. It combines the typical advantages of solid (magic angle spinning) and solution‐state (low power amplicators, lock channel and pulsed field gradients) NMR techniques. 1 It is based on the molec- ular mobility in the swollen samples, associated with fast spinning, from a rate of a few hundred hertz up to several kilohertz, at the magic angle (54.74°). This leads to drastically reduced line broadening effects due to dipolar couplings and chemical shift anisotropy, thus resulting in HR‐NMR spectra very similar to those obtained in solution. 2,3 Moreover, HR‐MAS NMR allows the acquisition of quantitative NMR (qNMR) data for multiple components in a unique sampling Received: 15 May 2017 Revised: 21 June 2018 Accepted: 1 July 2018 DOI: 10.1002/pca.2785 Phytochemical Analysis. 2018;1–9. © 2018 John Wiley & Sons, Ltd. wileyonlinelibrary.com/journal/pca 1