Identification and Quantification of Soluble Free, Soluble Conjugated, and Insoluble Bound Phenolic Acids in Durum Wheat (Triticum turgidum L. var. durum) and Derived Products by RP-HPLC on a Semimicro Separation Scale

Abstract

A straightforward semimicro separation scale RP-HPLC method was developed for the identification and quantification of phenolic acids (PAs) occurring as soluble free, soluble conjugated, and insoluble bound compounds, which were independently extracted from wholemeal of durum wheat and from its derived products coarse bran, semolina, and dried pasta. A narrow bore column and a semimicro photodiode array detector (PDA) cell, in conjunction with a single quadrupole mass spectrometer, equipped with an electrospray ionization source (ESI-MS), were employed. The method was validated in terms of linearity of calibration graphs, limits of detection, limits of quantification, repeatability, and accuracy, which was evaluated by a recovery study. In each sample (wholemeal, coarse bran, semolina, and dried pasta), the total amounts of the three different forms of PAs were in the order bound > conjugated > free, with bound PAs accounting for 61.0-83.6% of the total PAs. Ferulic acid was the most abundant PA in both soluble free and insoluble bound forms, whereas sinapic acid predominated in the conjugated ones. The highest PA content, calculated as the sum of total PAs quantified in the three forms, was found in coarse bran, followed by wholemeal, semolina, and dried pasta.

Influence of hot water blanching process on phenolic profile and antioxidant activity of Cinnamomum porrectum herbal tea

“…Free and conjugated phenolics are both soluble, whereas bound phenolics are insoluble. The major difference between free and conjugated phenolics is that the conjugated ones are usually esterified to sugars or other compounds with low molecular mass [33,34], while bound phenolics are covalently bound with cell wall structural components, such as cellulose and proteins [34,35]. TPC of un-treated herbal tea (57.07±1.45 mg GAE/ g DB) was lower than that of blanched herbal tea (66.25± 0.89 mg GAE/ g DB).…”

Variation of total antioxidant activity and of phenolic acid, total phenolics and yellow coloured pigments in durum wheat (Triticum turgidum L. var. durum) as a function of genotype, crop year and growing area

“…The 3 different forms of PAs occurring in durum wheat were individually extracted by a two step extraction method previously described (Nicoletti et al, 2013), using either 250 or 100 mg wholemeal for extracting the less abundant soluble free and conjugated forms and the more abundant insoluble bound PAs, respectively. Briefly, a common extraction step was performed using 80/20 (v/v) ethanol-water solution, then soluble free PAs were extracted twice with ethyl acetate, whereas soluble conjugated PAs were extracted with ethyl acetate after alkaline hydrolysis of the combined supernatants from the extractions with the ethanolwater solution.…”

“…The ethyl acetate extractions were preceded by acidification to pH 2.0 to suppress deprotonation and facilitate the solubilisation of PAs into the organic solvent. The identification and quantization of PAs was performed by reversed phase high performance liquid chromatography (RP-HPLC) employing a Polaris C18-A column (150 Â 2.0 mm I.D., 5 mm; Varian Inc. Lake Forest, CA, USA) with a C18 (30 Â 2 mm I.D., 5 mm) guard cartridge using instruments and experimental conditions previously described (Nicoletti et al, 2013). All standard and sample solutions were analysed in triplicate, and the concentrations of individual PAs in real samples were expressed in milligrams per kilogram of dry matter (mg/kg dm).…”

Development of stable isotope dilution assays for the quantitation of free phenolic acids in wheat and barley and malts produced thereof

“…For separation and quantitation, mainly high-performance liquid chromatography (HPLC) combined with different detectors, e.g., diode array detector [3,14,[16][17][18] or CoulArray detector [19], was applied. In recent studies, a diode array detector in combination with a mass spectrometer operating in the single ion monitoring (SIM) mode was used [20,21]. However, to date, results of the influence of different solvents and different extraction times on the efficacy of the extraction process are scarce.…”