Supplemental Figure 1.Representative HPLC–UV profiles for the standard solutions used to determine stilbene derivatives in extracts from grape Vitis amurensisbefore (a) and after 2 hours (b) sunlight exposure. The characteristic UV spectra of cis isomers detected in the extracts (and those of trans isomersfor comparison), recorded from the peak topswith a DAD detector, are shown with the imposition.The peaks are numbered as shown in Fig.1.
Supplemental Figure 2.The typicalMS/MS2spectra of piceid and viniferinisomersobtained in the negative ion mode using the ion trap mass spectrometer andrecorded from the peak tops.
Supplemental Figure 3.Segment of HPLC–UV profile,recorded at 310 nm, for theextractfrom grape Vitis amurensisleaves collected in October, demonstrates the presence of three major compounds: Q1, Q2 and Q3. The characteristic UV spectra of Q1, Q2 and Q3, recorded from the peak tops with a DAD detector, are shown with the imposition.
Supplemental Figure 4.The MSand MS2spectra of Q1, Q2 and Q3 obtained in the negative ion mode using the ion trap mass spectrometer and recorded from the peak tops.
Analytical chromatography
All solvents were of high performance liquid chromatography (HPLC) grade. Analytical standards: t-resveratrol, t-piceid, andpterostilbenewere obtained from Sigma-Aldrich (St. Louis, MO, USA);ε-viniferin was obtained from Panreac AppliChem (GmbH, Darmstadt, Germany).
Dried leaves, petioles, seeds, and berry skins of V.amurensis collected in June and October 2015 were analyzed for the presence of stilbenes at the Instrumental Centre of Biotechnology and Gene Engineering of IBSS FEB RAS. HPLC with MS and UV detection (HPLC-MS-UV) for identification andquantification of all components was performed using an 1260 Infinity analytical HPLC system (Agilent Technologies, Santa Clara, California, USA), equipped with a G1315D photodiode array detector, G1311C quaternary pump, G1316A column oven and G1329B auto sampler. The HPLC system was connected with a Bruker HCT ultra PTM Discovery System (Bruker Daltonik GmbH, Bremen, Germany), equipped with ion trap mass spectrometer. The extracts were separated on Zorbax C18 column (150 mm, 2.1-mm i.d., 3.5-μm part size, Agilent Technologies, USA); the column temperature was 40°C. The mobile phase consisted of a gradient elution of 0.1% aqueous acetic acid (A) and acetonitrile (B). The gradient profile with a flow rate of 0.2 mL/min was: 0 min 0% B; 35 min 40% B; 40 min 50% B. 50 min 100% B and then eluent B until 60 min. The injected volume was 1-5 μL. The MS analysis was carried out with electrospray ionization (ESI) and negative ions detection. The following settings were used: the range of m/z detection was 100-1,000, the drying gas (N2) flow rate was 8 L/min, the nebulizer gas (N2) pressure was 175 kPa, the ion source potential was -4.0 kV and the drying gas temperature was 325°C. Tandem mass spectra were acquired in Auto-MSn mode (smart fragmentation) using a ramping of the collision energy. The fragmentation amplitude was set to 1 V. If necessary, MS2experiments were performed only for the precursor ions of the monitoring compounds. UV spectra were recorded in the 200 – 400 nm range and chromatograms for quantification were acquired at 310 nm and 285 nm for trans- and cis- isomers respectively. Each quantification measurement was performed in duplicate.
Identification and quantification
Cis isomers of each stilbene were obtained under sunlight exposure of the respective standard solution containing the trans isomer as reported earlier (Romero-Pérez et al. 1999; Pezet et al. 2003). Almost 90% isomerisation was obtained after 3 h of exposure. Cis isomers of stilbenes are not available commercially. In order to define cis-piceid in our samples, the standard solutions of it were prepared and determined by comparison of the UV and MS data with information previously published (Vian et al. 2005; Huang and Mazza 2011). The MS2 and MS3 fragmentation patterns of cis-piceid were identical with those of trans-isomer obtained under the same MS conditions. The concentrations of cis-piceid were calculated from the difference between the concentrations before and after sunlight exposure of the trans-isomer.
In order to define cis-piceid and cis-ε-viniferin in our samples, the standard solutions of them were prepared and cis isomers were determined by comparison of the UV and MS data (Supplementary materials) with information previously published (Pezet et al. 2003; Vian et al. 2005; Mulinacci et al. 2010; Huang and Mazza 2011). Multistage MS analysis (MS2 and MS3, if necessary) for all components of the resulting mixture were realized, target fragmentation spectra were obtained in the negative ion mode using the ion trap mass spectrometer. The fragmentation patterns of cis isomers were similar with those of trans isomers obtained under the same MS conditions. The concentrations of cis isomers were calculated from the difference between the concentrations before and after sunlight exposure of the trans-isomers.
All determined components of the extracts (Fig.1) were identified as we described recently (Aleynova et al. 2016) on the base of UV spectra, recorded with a DAD detector, mass spectral dataandchromatographic separation with reference to the values of their respective standards. The contents of each component were determined using by external standard method using the four-point regression calibration curves built with the reference standards.
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