Phytic acid dynamics during seed development and it’s composition in yellow and black Indian soybean (Glycine max L.) genotypes through a modified extraction and HPLC method

Abstract

Identifying or designing low phytic acid soybean demands an improved understanding of its dynamics and a need for a reliable and accurate phytate quantification method and hence an improved quantitative technique for accurate and rapid quantification of phytic acid (PA) using high-performance liquid chromatography is proposed in this paper. A rapid PA extraction method utilizing sonication of sample in 0.78 M HCl for 3 min followed by mechanical agitation and separation using strong anion exchange column in a vacuum manifold was optimized. The elution of PA was performed using a RP-C₁₈ column with an isocratic mobile phase [Acetonitrile, 35 mM formic acid and tetrabutylammonium hydroxide (4.8: 5.1: 0.1, v/v/v)]. The modified method was rapid, accurate, precise, and reproducible with relative standard deviation of 1.80 and 3.01 % (n = 10, for 1 mg ml⁻¹) for within and between days respectively with linearity (R² = 0.999, P < 0.05), low limit of detection (LOD = 7.8 μg ml⁻¹) and limit of quantification (LOQ = 31.25 μg ml⁻¹). PA dynamics was found increased in a linear trend from initial to later developing stages until maturity in both yellow (DS-9814) and black (DS-MM-64) genotypes. The PA content ranged from 2.38–4.72 g 100g⁻¹ in the 20 soybean genotypes screened and the variability in PA content was more in black genotypes (P < 0.0001). The black soybean genotypes JS 76-205 (2.38 g 100g⁻¹), Kalitur (2.50 g 100g⁻¹) and UPSL 652 (2.54 g 100g⁻¹), inherently rich in anthocyanin, contains the lowest PA content and hence can serve as potential genotypes with nutraceutical benefits.