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Response surface methodology for optimization of the ultrasonic extraction of flavonoids from Lotus corniculatus L.

Lotus corniculatus L., Fabaceae (bird's-foot trefoil), is a perennial herbaceous plant species, wide spread in temperate regions of Europe, Asia and America [1]. In this work L. corniculatus was investigated as a cheap and widely available source of flavonoids and other phenols with potential use as food supplements. For efficient extraction of the desired compounds, ultrasonic-assisted extraction procedure was optimized using Box–Behnken design. The independent variables were: ethanol concentration (X1), temperature (X2), and extraction solvent pH (X3), while the responses were yield of the extraction (Y), DPPH radical scavenging activity of the extracts (RSA IC50), content of total phenols (TP), total flavonoids (TF) and total phenolic acids (TPA) which were analysed specrophotometrically. Concentration of individual flavonoids in the obtained extracts was determined using HPLC-DAD. Response-surface methodology was used to find the relationship between dependent and independent variables. The selected quadratic modelshadr2 values over 0.9. The most important extraction variable was X1which had a significant (P< 0.05) effect on Y, TP and TF, both as a linear and quadratic term. Furthermore, it influenced TPA but only as quadratic term. RSA IC50, on the other hand, was influenced by pH, temperature as well as their interaction. Taking into consideration only the statistically significant extraction variables it was found that TP and TPA were best extracted using 45 % ethanol, whereas 95 % ethanol was more appropriate for TF extraction. Lowest RSA IC50 was achieved at 60 C and pH 7. The main flavonoids in the extracts were glycosides of flavonols quercetin and kaempferol, as well as glycosides of isoflavone genistein. However, even though phenolic compounds are reputed antioxidants [2], the lack of correlation between RSA IC50 and the other investigated responses indicates that the other components play crucial role in the observed antiradical activity.

Lotus corniculatus ; Fabaceae ; polyphenols ; genistein ; quercetin ; DPPH ; kaempferol ; response surface methodology

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