The mixed enzymes-assisted mechanical bio-extraction method was first used to extract polysaccharides from Dendrobium officinale. Different parameters including the ratio of enzyme, the amount of enzyme, the grinding time, the extraction time and the solid/liquid ratio were investigated by single factor experiments and multifactorial experiments. Through the response surface methodology the optimal extraction conditions were obtained with the ratio of cellulase to pectinase was 2: 1 and total amount of enzyme was 0.23 mg, the grinding time of 11.48 min, the extraction time of 5.99 min. The obtained polysaccharide extracts were hydrolyzed and derivatized and then injected into high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF/MS) for monosaccharide composition analysis. After optimization of the chromatographic conditions (including mobile phase and column), twelve monosaccharides were successfully determined within 20 min. The proposed method provided satisfactory linearity with the correlation coefficients higher than 0.99, suitable recoveries (81.46-114.92 %), acceptable reproducibility ranging from 0.06 % to 4.77 %, low limits of detection (0.70-45.45 ng/mL). Compared with other methods, this method makes the extraction efficiency much higher and has the advantages of simple operation, environmental friendliness and mild extraction conditions. Therefore, this method can be used for the extraction of polysaccharides from plants and the determination of monosaccharides and has the potential to be used in more areas.
Keywords: Dendrobium officinale; Enzymes-assisted mechanical bio-extraction; High performance liquid chromatography-quadrupole time-of-flight mass spectrometry; Monosaccharide; Polysaccharide.
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