In all sampling dates and years, 10 berries were collected from each vine for the analysis of °Brix, pH, and titratable acidity. Contemporarily, 20 berries per vine were collected for anthocyanin analysis via HPLC, following the method described in Mattivi et al.42 (link) and using a Waters 1525 HPLC (Waters, Milford, MA) equipped with a diode array detector (DAD) and a Phenomenex (Castel Maggiore, BO, Italy) reversed-phase column (RP18, 250 mm × 4 mm, 5 μM). Anthocyanins were quantified at 520 nm using an external calibration curve with malvidin-3-glucoside chloride as the standard (Sigma-Aldrich).
Rp 18
Manufactured by Phenomenex
Sourced in Italy, United Kingdom, United States
RP-18 is a reversed-phase liquid chromatography column. It is designed for the separation and purification of a wide range of organic compounds. The column packing material consists of silica particles with chemically bonded octadecyl (C18) functional groups.
Automatically generated - may contain errors
5 protocols using rp 18
1
Grape Anthocyanin Profiling Protocol
2
Quantitative UHPLC-PDA Analysis of Compounds
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The samples were analyzed by injecting 10 μL of the chromatographed fraction solutions (2 mg/mL in methanol) into a UHPLC connected to a photodiode array detector (Dionex Ultimate 3000) with a reverse-phase column (Phenomenex RP-18, 150 × 4.0 mm, 3 μm). The mobile phase consisted of (A) DDW with 0.1% formic acid and (B) acetonitrile containing 1% formic acid. The gradient started with 0% B for 5 min, then increased to 98% B for 30 min, and maintained at 98% B for another 5 min at a flow rate of 1 mL/min.
3
UHPLC-PDA Analysis of Fractions
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The chromatographed fractions were analyzed by injecting 20 µL of the fraction solution dissolved in methanol (2 mg mL−1) into a UHPLC connected to a photodiode array detector (Dionex Ultimate 3000) with a reverse-phase column (Phenomenex RP-18, 150 4.0 mm, 3 μm). The mobile phase consisted of (A) DDW with 0.1% formic acid and (B) acetonitrile containing 0.1% formic acid, run at a gradient starting from 40% B, then increased to 90% B for 22 min, and kept at 90% B for another 8 min, at a flow rate of 1 mL min−1.
4
HPLC Analysis of Phytochemicals
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The extracts were analysed on Shimadzu Class-VPV 5.03 (Kyoto, Japan) equipped with Shimadzu UV-Vis detector (SPD-10Avp) at 330 nm, LC-16ADVP binary pump, DCou-14 A degasser, and Phenomenex RP-18 (UK; 250 × 4.00 mm, 5 µ) column according to Kim et al. (2006 ). The solvent gradient used in this study was formed through a solvent (A methanol and B 0.03% phosphoric acid in water). The linear gradient was used for chromatographic separation: 0 min, 70% B; 6 min, 55% B; 20 min, 45% B. The run time was 30 min. The injection volume was 5 µL. The solvent flow rate was maintained at 1.0 mL/min. All analyses were carried out at 30 °C. The spectra were recorded at 330 nm. The method of the external standard was used for quantification.
5
Reversed-Phase HPLC for Oligonucleotide Crosslinking
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All oligonucleotide-crosslinker reactions were monitored by RP-HPLC measurements (Shimadzu, Korneuburg, Austria) on a reversed phase column for oligonucleotide analysis (Clarity 5 µ RP18 250 x 4.60 mm, Phenomenex, Torrance, USA). For all runs, a 60 min linear gradient of 8-44% acetonitrile in triethylammonium acetate (TEAA) buffer in nuclease-free water, adjusted to pH 7.0 was performed at a flow rate of 1 mL/min. Before RP-HPLC measurement, 25 mM dithiothreitol (DTT) was added to SPDP-crosslinked siRNA sample for disulfide cleavage. All siRNA samples were diluted to a 50 µM solution with TEAA buffer for measurements.
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