Proton Nuclear Magnetic Resonance (1H NMR) Metabolic Profiles Discriminate Two Monovarietal Extra Virgin Olive Oils, Cultivars Arbequina and Koroneiki, with Different Geographical Origin

For NMR analysis, olive oil (~140 mg) was dissolved in deuterated chloroform (CDCl₃) containing tetramethylsilane (TMS) as an internal standard, in the ratio of oil:CDCL₃ of 13.5:86.5 w/w (standard Bruker methodology), and vortexed for 15 s. From the obtained mixture, a volume of 600 μL was transferred into a 5 mm NMR tube. The ¹H NMR spectra were acquired on a Bruker Avance III spectrometer operating at 400.13 MHz, T 300.0 K, equipped with a BBI 5 mm inverse detection probe incorporating a z-axis gradient coil. NMR acquisitions were performed under full automation for the entire process after loading each sample on a Bruker Automatic Samples Changer interfaced with IconNMR (Bruker) software. Automated tuning and matching, locking and shimming, and calibration of the 90° hard pulse P(90°) were performed for individual samples using standard Bruker routines ATMA, LOCK, TOPSHIM, and PLSECAL in order to optimize NMR conditions. The ¹H NMR spectra were recorded with the following acquisition parameters: zg Bruker pulse program, 64 k time domain (TD), spectral width (SW) of 20.5524 ppm, a receiver gain (RG) of 4 and 16 repetitions. Moreover, to enhance signals of the minor components, a Bruker multi-suppressed ¹H noesygpps NMR (noesygpps1d.comp2 pulse program) experiments was performed, using the following conditions: 32 k time domain, spectral amplitude 20.5555 (8223.685 Hz), p1 12.63 µs, pl1 −1.00 dB, 32 repetitions. All ¹H spectra were obtained by the Fourier Transformation (FT) of the Free Induction Decay (FID), applying an exponential multiplication with a line broadening factor of 0.3 Hz, automatically phased, and baseline corrected. Chemical shifts were reported with respect to TMS signals set as 0.0 ppm, obtaining peak alignment. The olive oil profile and phenolic molecules assignments were performed based on 1D NMR spectral analysis and by comparison with published data [35].