Studies on Selective Extraction and Purification of Bioactive Compounds from Kokum (Garcinia Indica) Fruits using Alcohol Based Aqueous Two-Phase Systems

Abstract

The bioactive compounds namely natural pigments and antioxidants are gaining high demand in the global market because of the increasing public awareness on the positive health benefits of these compounds due to their greater functional and bioactive properties and their potential applications in food, nutraceuticals, and cosmetic industries. The difficulty in the recovery from the complex natural sources and the instability due to the denaturation during the extraction and purification were inhibiting the application of these molecules in various industries. The substantial effort has been made to develop new technologies to extract and purify these bioactive compounds by retaining their native characteristics and stability. Kokum fruits (Garcinia indica) contain the important bioactive compounds like anthocyanin (ACNs), hydroxycitric acid (HCA), garcinol (GL), and isogarcinol (IGL) at a significant quantity. The simultaneous extraction of all these bioactive components in a crude extract is achieved using the aqueous mixtures of 1-propanol and ethanol as solvents. The aqueous two-phase extraction (ATPE) is explored for the first time to simultaneously partitioning the four bioactive compounds from the crude extract of kokum rinds. Alcohol-based aqueous two-phase systems (ATPSs) were proved to be suitable for the simultaneous partitioning during screening studies. The ethanol-ammonium sulphate and 1-propanol-ammonium sulphate/magnesium sulphate ATPSs are proved to be better for differential partitioning of GL and IGL into alcohol rich top phase and ACNs and HCA into the salt rich bottom phase. The effect of phase compositions and Tie Line Length (TLL) on the differential partitioning was investigated in terms of partitioning coefficient and extraction efficiency. The Response Surface Methodology (RSM) was adopted to optimize the process variables through desirability based multi-response optimization by considering the partitioning coefficients (K) and extraction efficiency (EE) of all the four bioactive compounds as responses. The ATPS consisting of 15.202 % (w/w) 1-propanol, 10.242 % (w/w) ammonium sulphate having the TLL of 28.505 % (w/w) at a crude load of 25 % (w/w) able to partition 97.39 % GL (K=370.770) and 92.38 % IGL (K=120.581) in the top phase and 99.19% ACNs (K=0.080) and 99.83% HCA (K=0.016) in the bottom phase with a purity higher than 99% by implementing the second stage ATPS. An attempt was further made to extract the bioactive compounds in the continuous extractor, Rotating Disc Contactor (RDC) with 1-propanol- ammonium sulphate ATPS. The efficacy of RDC column was analysed by studying the dispersed phase holdup, mass transfer coefficient, and recovery and separation efficiency of the bioactive compounds at different operating conditions.