Excipient-assisted method to enhance yield of genistein and daidzein from Trifolium pratensis L. flowers

Abstract

Introduction. Red clover (Trifolium pratensis L.) is a wild plant belonging to the legume family and is often used to relieve symptoms of menopause, high cholesterol, osteoporosis and conditions related to aging. This is because of their high. concentrations of phytoestrogens, specifically isoflavones (genistein, daidzein), which are thought to be especially healthful [1,4]. Nevertheless, isoflavones absorption varies considerably among individuals. This may be related to differences in the intestinal microbial population in each individual, as natural isoflavones are conjugated as glycosides that have to be degraded for absorption to occur [5]. Glycosides (daidzin and genistin) are inactive compounds but become active in aglycone (daidzein and genistein) form when the glucose molecule is removed from the structure. Hydrolysis of glycosides is an important step to obtain biologically active and easily absorbed forms of isoflavones [2]. To obtain high isoflavones aglycones yields from raw red clover material hydrolysis must be combined with extraction method. Excipients also can be used for improving extraction and changing environmental conditions – surfactants, emulsifiers, salts that change ion voltage and pH-adjusting substances [3]. Therefore, the aim of this study was to use excipient- assisted (croscarmellose sodium, sodium carboxymethyl starch) method to enhance yield of genistein and daidzein from Trifolium pratensis L. flowers. Materials and methods. Extractions were done using 0.3 ± 0.001 g of dried and milled flower heads and 10 mL of distilled water. Ultrasound-assisted extraction (frequency 38 kHz) was performed using 250 W power, temperature 40 °C, extraction time 10 and 30 min. After ultrasound processing samples were refluxed in the sand bath at 100 °C for 1 hour. Excipient sodium carboxymethyl starch or croscarmellose sodium was added to the extraction media before ultrasound processing. The excipient amount was based on solvent quantity. Control samples were prepared using the same conditions without excipient. Extraction samples for quantification of isoflavones aglycones (genistein, daidzein) were investigated using HPLC. [...].