Simultaneous extraction and purification of polysaccharides from Gentiana scabra Bunge by microwave-assisted ethanol-salt aqueous two-phase system

Abstract

An effective method of concurrent extraction and purification for polysaccharides from Gentiana scabra Bunge stems was developed by microwave-assisted aqueous two-phase extraction (MA-ATPE). Ethanol/sodium dihydrogen phosphate was employed to form aqueous two-phase system (ATPS) by evaluating the extraction properties of five combinations of ethanol/salt systems. The important factors, namely, the composition of ATPS, extraction temperature, solvent-to-material ratio, extraction time and the particle size were investigated by single factor experiment and response surface methodology (RSM) combined with Box–Behnken Design (BBD). The final extraction conditions were concluded as follows: ethanol concentration 21.73% (w/w) and NaH2PO4 concentration 23.27% (w/w) for the ATPS, extraction temperature 95°C, extraction time 5.8min, solvent-to-material ratio 21:1 (mL/g) and particle size 80mesh. Under the optimum conditions, the extraction yield of polysaccharides were 15.97±0.32% for the top phase, 0.58±0.26% for the bottom phase and 16.55±0.13% for total polysaccharides, and the purity of water-soluble polysaccharides was 78.19±0.42%. Compared with smashing tissue extraction (STE), microwave assisted-extraction (MAE), ultrasonic-assisted extraction (UAE) and heat reflux extraction (HRE), microwave-assisted aqueous two-phase extraction not only improved the yield and purity of water-soluble polysaccharides, but also obtained a variety of polysaccharides. In addition, the chemical characterization of polysaccahrides from both the two phases were carried out by Fourier transforms infrared spectrum (FT-IR) and High performance liquid chromatography derivatization with 1-phenyl-3-methy-5-pyrazolone (HPLC-PMP). The extraction mechanism was also discussed by scanning electron microscope (SEM). Therefore, as an efficient and promising technique, microwave-assisted aqueous two-phase extraction may be a viable option for the simultaneous extraction and purification of polysaccharides from plants.