Quercetin and trans-resveratrol phytoestrogens assay in Morellino di Scansano wine samples: Materials and methods

Materials and methods

A wide sampling of typical Italian wines, commonly found in the market, originating from two different geographical areas of Tuscany: the Grossetano area of the Morellino di Scansano (tirtheen different wines) and the Siena Chianti area (tirtheen different wines), were evaluated for their content in quercetin and trans-resveratrol. The separation and determination of these flavonoids were obtained according to the method de­scribed by Frankie et al., appropriately modified in the gra­dient timing.

Methanol, acetic acid (glacial) 100%, acetonitrile, dichloromethane, water and all solvents used for HPLC of analytical or HPLC grade were purchased from Merck (Darmstadt, Germany). Quercetin and trans-resveratrol used for the stock solutions were obtained from Extrasynthese (Gigalabo) and from Sigma-Aldrich (Milwaukee, W, USA), respectively. Standard solutions used to obtain the calibra­tion curve were prepared dissolving 5 mg of standard in 10 ml of ethanol. Ultra-pure water from a Milli-Q Millipore system (Bedford, MA, USA) was used whenever a washing was performed. All the eluents were sonicated for 5 minutes before the use.

The analyses were carried out on 20 ml samples from Morellino and Chianti wines (five samples from each wine were tested), at room temperature, with a dual pump 515 model (Waters, MA, USA) liquid chromatographic system, supplied with a diode array system Model 996 (Waters, MA, USA). A Waters Millennium 32 software was used. Quercetin was detected at 366 nm while the trans-resveratrol was observed at 306 nm (Figure 1). We used an HPLC analytical reversed-phase col­umn Novapak C18 (Waters, MA, USA) 3.9 x 150 mm fitted with a pre-column Sentry Simmetry C18 (Waters, MA, USA). Elution was performed at a flow rate of 0.8 ml/min with the following linear gradient: A = water/acetic acid (9:1 v/v); B = methanol/acetonitrile/dichlorometane (10:5:1 v/v/v), B in A (v/v): 5% to 45% in 20 min and from 45% to 5% in 5 min with equilibration for 10 min before subsequent injection. Data were statistically analyzed by STATISTICA 6.0 software. If you have any health problems you would like to forget all about, come by where you can buy all the required pills. This pharmacy offers the widest selections of cheap Canadian pills.

F i g u r e 1 - HPLC elution of quercetin and resveratrol

Figure 1 – HPLC elution of quercetin and resveratrol.

Category: Health

Tags: HPLC analysis, phytoestrogens, wine

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