A HIGH-THROUGHPUT UPLC-MS/MS FOR THE SIMULTANEOUS ANALYSIS OF SIX PHYTOESTROGENS FROM GENISTA TINCTORIA EXTRACTS

Abstract

& A new rapid, sensitive UPLC method coupled with ESI-MS=MS detection was developed and validated for the simultaneous quantification of six phytoestrogens (daidzin, genistin, genistein, daidzein, formononetin, and coumestrol) from Genista tinctoria L. extracts. An adequate chromatographic separation was achieved with gradient elution and a mobile phase consisting of a mixture of 0.1% acetic acid=methanol (70=30, v=v). The total run time was of 5.5 min. The detection of target compounds was performed in multiple reaction monitoring mode. The method was validated and proved to be linear, accurate, and precise over the range of 10-800 ng=mL. The LLOQ values estimated based on the signal to noise ratio were of 5 ng=mL for each analyte, except for daidzein (LLOQ ¼ 10.78 ng=mL). The elaborated assay shows shorter run time, lower solvent consumption, and higher sensitivity than most of the existing LC methods described in the literature. The method was used to quantify the selected phytoestrogens in Genista tinctoria herba or flos extracts. The major compounds found were genistein and its glycosidic form genistin. The method proved to be adequate for the characterization of plant extracts from the point of view of their phytoestrogen content, a group of active compounds with great therapeutic potential.

Isoflavones: Vegetable Sources, Biological Activity, and Analytical Methods for Their Assessment

“…It is used in phytotherapy for its antianemic, antihemorrhagic, and remineralization properties [26] and for its hypocholesterolemic, antimicrobial, hypolipidemic, antioxidant, antiulcer, neuroprotective, and estrogenic properties [27]. Species of Genista (G. tinctoria L., G. sagittalis L.) contain essentially genistin and genistein [19,20,28,29]. They are known for their hypoglycemic [30], anti-inflammatory, antiulcer, spasmolytic, antioxidant, and estrogenic properties [31].…”

“…In recent years, due to the health benefits provided by isoflavones, higher attention has been paid to the analytical methods that allow identification and quantification of isoflavones from different types of samples: (a) food, for dietary intake assessing [15,70]; (b) food supplements, for standardization of nutraceuticals [5,71]; (c) vegetable products, for phytotherapeutic evaluation [19,20,28]; and (d) human biological samples (plasma, urine) [5]. These analytical methods are commonly used for isoflavone bioavailability assessing and in pharmacokinetic or pharmacological studies.…”

“…Hydroalcoholic extracts or tinctures can be prepared from either fresh or dry and pulverized vegetable materials. The hydroalcoholic extracts can be made in 70% ethanol or methanol, by refluxing and filtration; by cold maceration, pressing, and filtration [20]; by percolation [28]; or using modern methods, such as ultrasound-assisted extraction in 50% ethanol [19]. The extracts can be analyzed directly by LC-MS/MS, after an adequate dilution [20], or they can be subjected to an acid hydrolysis [19] in order to release aglycones.…”

“…For isoflavone identification, the following chromatographic methods are used: gas chromatography coupled with mass spectrometry (GC-MS) [5,15], high-performance liquid chromatography (HPLC) with UV detector (photodiode array, PDA) [28,70,71], fluorescence detector (FLD), electrochemical detector (ECD) or mass spectrometer detector (MS) [20,74,75], and, less often, capillary electrophoresis (CE).…”

GC‐MS Analysis of Various Phytoestrogens in Health Functional Foods

“…[41][42][43][44][45][46] Analysis of a wide range of 21 phytoestrogen contents in HFFs has not yet been reported. Intake of KAE, mainly found in fruits, broccoli, and many berries, is known to correlate with a low incidence of ovarian cancer in postmenopausal women, and KAE has been shown to sensitize cancer cells to anti-cancer agents.…”