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  • Identification and quantification of phytochemicals in nutraceutical products from green tea by UHPLC-Orbitrap-MS.

Identification and quantification of phytochemicals in nutraceutical products from green tea by UHPLC-Orbitrap-MS.

Food chemistry (2014-12-04)
Noelia López-Gutiérrez, Roberto Romero-González, Patricia Plaza-Bolaños, José Luis Martínez Vidal, Antonia Garrido Frenich
ABSTRACT

A method has been developed and validated for the simultaneous detection and quantification of phytochemicals in nutraceutical products obtained from green tea. For that purpose, ultra-high performance liquid chromatography coupled to single-stage Orbitrap high resolution mass spectrometry (UHPLC-Orbitrap-MS) has been used. A database containing 37 compounds has been used for the detection and identification of the target compounds. The developed methodology was based on solid-liquid extraction, using a mixture of methanol:H2O (80:20, v/v, pH 4), followed by dilution (10 times) with a mixture of ammonium acetate:methanol (50:50, v/v). Chromatographic conditions were optimised and full scan accurate mass data acquisition using electrospray ionisation in positive and negative ion mode was used. Moreover, all-ion fragmentation mode was used to get information of fragment ions, and they were used for identification purposes. The developed method was validated, obtaining repeatability (intra-day) and inter-day precision values (expressed as relative standard deviation, RSD) lower than 16% and 20%, respectively. Lower limits were also evaluated and limits of detection (LODs) ranged from 1 to 50 μg L(-1), while limits of quantification (LOQs) ranged from 2 to 150 μg L(-1). Recovery was performed at five levels and it ranged from 70% to 109%. Finally, this method was used to evaluate the phytochemical content in 10 samples (tablets or capsules), showing concentrations of (+)-catechin, (-)-epicatechin, gallic acid, (-)-gallocatechin and quercetin-3-O-rutinoside, ranging from 258 (C6) to 10,729 (C6) mg kg(-1).

MATERIALS
Product Number
Brand
Product Description

Supelco
Sodium dodecyl sulfate, dust-free pellets, suitable for electrophoresis, for molecular biology, ≥99.0% (GC)
Epigallocatechin, primary reference standard
Supelco
(−)-Gallocatechin gallate, analytical standard
Caffeine for system suitability, European Pharmacopoeia (EP) Reference Standard
Supelco
(−)-Gallocatechin, analytical standard
Supelco
Melting point standard 235-237°C, analytical standard
Chlorogenic acid, primary reference standard
Sigma-Aldrich
Methanol, NMR reference standard
Sigma-Aldrich
Quercetin, ≥95% (HPLC), solid
Supelco
Genistein, analytical standard
Supelco
Apigenin, analytical standard
Sigma-Aldrich
Sodium dodecyl sulfate, ReagentPlus®, ≥98.5% (GC)
Sigma-Aldrich
Sodium dodecyl sulfate, 92.5-100.5% based on total alkyl sulfate content basis
Sigma-Aldrich
Genistein, from Glycine max (soybean), ~98% (HPLC)
Sigma-Aldrich
(−)-Gallocatechin, ≥98% (HPLC)
Sigma-Aldrich
Taurocholic acid sodium salt hydrate, ≥95% (HPLC)
Sigma-Aldrich
Taurocholic acid sodium salt hydrate, BioXtra, ≥95% (HPLC)
Sigma-Aldrich
Sodium dodecyl sulfate, BioReagent, suitable for electrophoresis, for molecular biology, ≥98.5% (GC)
Sigma-Aldrich
(−)-Gallocatechin gallate, from green tea, ≥98% (HPLC)
Sigma-Aldrich
Sodium dodecyl sulfate, BioXtra, ≥99.0% (GC)
Sigma-Aldrich
(−)-Epigallocatechin, ≥95% (HPLC), from green tea
Sigma-Aldrich
Sodium dodecyl sulfate, ≥98.0% (GC)
Supelco
Acetic acid, analytical standard
Supelco
Methanol, analytical standard
Supelco
Sodium dodecyl sulfate, suitable for ion pair chromatography, LiChropur, ≥99.0%
Sigma-Aldrich
Apigenin, ≥95.0% (HPLC)
Sigma-Aldrich
Acetic acid, for luminescence, BioUltra, ≥99.5% (GC)
Sigma-Aldrich
Methanol, anhydrous, 99.8%
Sigma-Aldrich
Sodium dodecyl sulfate, ≥98.0% (GC)
Sigma-Aldrich
Acetic acid, natural, ≥99.5%, FG