Solid-phase microextraction and on-fiber derivatization for assessment of mammalian and vegetable milks with emphasis on the content of major phytoestrogens

Pietro Cotugno,Antonella Aresta,Carlo Zambonin

doi:10.1038/s41598-019-42883-7

Pietro Cotugno, Antonella Aresta + Show 1 more

Open Access

https://doi.org/10.1038/s41598-019-42883-7

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Abstract

A new solvent-free method for the simultaneous determination of some major phytoestrogens (equol, enterodiol, daidzein, genistein, glycitein) in different commercial milks (cow, goat and soy-rice) was developed. After solid phase microextraction, performed by direct immersion of a 65 μm-polydimethylsiloxane–divinylbenzene fiber in diluted (1:100 with 0.2% formic acid - 30% sodium chloride) milk samples (18 °C for 20 min under stirring), a direct on-fiber silylation with N,O-bis (trimethylsilyl)trifluoroacetamide) containing 1% trimethylchlorosilane (70 °C for 20 min) was performed prior to gas chromatography–mass spectrometry analysis. Since the target compounds were determined as aglycones, the hydrolytic removal of the aglycone from the glycosides was performed. The method permitted the determination of the target analytes in all the considered milk samples as well as the detection of some major amphipathic fats indicating that the approach could potentially be applied in the future for further applications, such as milk profiling.

Highlights

Milk is the primary source of nutrition for young mammals
As far as the derivatization of the analytes is concerned, three different procedures can be used in combination with solid-phase microextraction (SPME): derivatization in the GC injection port, in-situ derivatization and on-fiber derivatization[23,24,25]
The in-situ derivatization involves the addition of the derivatizing agent to the sample matrix and the subsequent SPME of the derivatized analytes from the medium, even if the presence of the agent in the sample can negatively affect the extraction

Summary

Introduction

Milk is the primary source of nutrition for young mammals. It contains important nutrients which are essential for the growth of infant, such as lactose, fats, proteins, vitamins, and salts[1]. Genistin and glycitin are major isoflavones found in a wide variety of plant, in particular soybean and clover[5,6,7] They are metabolized in the intestine to their aglycone forms, daidzein, glycitein and genistein, respectively, that possess higher estrogenic power[3,8]. Equol is a lignan metabolized from daidzein that possess longer half-life, higher antioxidant properties and affinity for the estrogen receptors than its precursor[10]. It has protective effects against UV-induced DNA damages[11]. Enterodiol is a mammalian lignan with antitumor activity mainly produced in the colon by the action of gut bacteria on its glycosides matairesinol[14]

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