Sustainable extraction of phytoestrogens from soybean and okara using green solvents

Soy extract waste, okara, is a rich source of bioactive compounds such as isoflavones, which are phytoestrogens with potential health benefits. To develop a green approach to recovering these compounds and valorizing okara, a study was developed to screen variables for the extraction of isoflavones from okara and soybean (for comparison) using Deep Eutectic Solvents (DES) composed with choline chloride ([Ch]Cl) and acetic acid (AA) ([Ch]Cl: AA, 1:2). A fractional design (2^4-1) was used to evaluate variables in the extraction of isoflavones, followed by a Central Composite Rotatable Design (CCRD). The variables analyzed included temperature, % water, solid-liquid ratio, and stirring speed. Furthermore, cytotoxicity tests were carried out using Caco-2 cells from the extracts obtained under best conditions, and the solid matrices were evaluated by scanning electron microscopy (SEM). This study showed that 1194.56 µg TIC/g of dried soybeans were extracted using [Ch]Cl: AA with 70 % water added at low temperature (25 °C), 500 rpm, S/L ratio of 10 mg/mL and extraction time of 2 h. For okara, 450.9 µg TIC/g of dry okara were extracted at 40 °C, 500 rpm, 61.5 % water in DES, S/L ratio of 10 mg/mL, and extraction time of 1 h. In the cytotoxicity tests, it was observed that the neutralization step of the extracts is necessary to reduce the cytotoxic effects caused by the high acidity of the solvents. In summary, this work has shown that aqueous mixtures of DES can be used as sustainable alternative techniques for recovering isoflavones from soybean residue and can replace toxic conventional solvents.