Development and validation of a novel approach based on ultrasonic-assisted enzymatic extraction and RP-UHPLC for simultaneous determination of astaxanthin isomers and alpha-tocopherol in Haematococcuspluvialis derived supplements

摘要

A rapid, effective andgreenmethod was developed for simultaneous determination of total (free and esterified) astaxanthin (AX) isomers (all-E, 9Z and 13Z) and alpha-tocopherol(AT) in Haematococcuspluvialis derived supplements. The new method employed a highly efficient ultrasonic-assisted enzymatic extraction (UAEE) techniqueto perform deesterification with Cholesterol esterase from Pseudomonas fluorescenspermitting the concurrent detection. The subsequent RP-UHPLC method for separating and measuring was performed on a simple C18 column within 10.5 min by using methanol and ammonium acetate as mobile phase with a gradient elution. The proposed method was validated according to international guidelines and itproved to be highly accurate and robust. The optimized UHPLC method allowed easy transfer to HPLC, and allowed rapid analysis of active ingredients profiling in H. derived supplements.To our knowledge, this is the first quantification approach describing the rapid simultaneous analysis of the functional lipophilic substances including AX isomers in H. derived supplements using UAEE technique combined with RP-UHPLC.Moreover, this holistic approachcan be used to identify whether AX products are of natural origin or chemical synthesis, and may find more applications in new forms of H. derived productswith complexbiological matrix for more research on the bioavailability of AXisomersfrom natural source. (C) 2020 Elsevier B.V. All rights reserved.