Isolation and structural characterization of mushroom polysaccharides in relationship with their antioxidant activities

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Isolation and structural characterization of mushroom polysaccharides in relationship with their antioxidant activities

 

Author: Siu, Ka Chai
Title: Isolation and structural characterization of mushroom polysaccharides in relationship with their antioxidant activities
Degree: Ph.D.
Year: 2015
Subject: Edible fungi.
Polysaccharides.
Antioxidants.
Hong Kong Polytechnic University -- Dissertations
Department: Dept. of Applied Biology and Chemical Technology
Pages: xxi, 182 leaves : illustrations (some color) ; 30 cm
Language: English
InnoPac Record: http://library.polyu.edu.hk/record=b2798181
URI: http://theses.lib.polyu.edu.hk/handle/200/7896
Abstract: Edible fungi (mushrooms) are well-known for their notable nutraceutical functions and medicinal properties. Polysaccharides (PS) including polysaccharide-protein complexes (PSPs) are major bioactive constituents of edible and medicinal fungi. In addition to their well-recognized antitumor and immunomodulatory properties, significant antioxidant activities have also been reported in the crude PS (CPS) extracted from many edible fungi. However, the complex and unclear chemical composition of the CPS fractions, which are mostly crude mixtures of PS, proteins, and their complexes in a wide molecular weight (MW) range, poses a major challenge in evaluating and understanding their bioactivities. This research project aims at the isolation and purification of PS from important edible/medicinal fungi, both wild and cultivated, and the investigation of their chemical composition, molecular properties and relationship to antioxidant activities. Fourteen species of wild edible mushrooms were collected from the Lesser Khingan Range in northeast China and CPS were isolated from their water extracts by ethanol (EtOH) precipitation, and partially purified by deproteinization and dialysis. The antioxidant activities varied with both the species, of which the three species showing the highest activities were from Handkea utriformis (535.8 μmol Trolox/g), Macrolepiota mastoidea (378.6) and Armillaria ostoyae (329.1) in radical scavenging, H. utriformis (5.94 mmol Fe2+/g), Lepista nuda (4.65) and Armillaria ostoyae (4.42) in reducing power, and Armillaria cepistipes (484.6 μmol Trolox/g), H. utriformis (274.8) and M. mastoidea (202.5) in ferrous ion chelating. For most of the mushroom species under investigation, this is the first report on their PS contents and bioactivities. These wild mushrooms provide a good source of PS as potential antioxidants. Among the wild mushrooms from the forests in Lesser Khingan Range, A. ostoyae is one of the most well-known and the largest living organisms in the world. The CPS isolated from hot water (HWE), aqueous ammonium oxalate or aqueous alkaline (AKE) extraction were composed mainly of carbohydrates (43-85%) and proteins (16-30%) plus low content of phenolic pigments (< 2.5%). To isolate CPS of different MWs and evaluate the MW-activity relationship, we performed gradient precipitation using different volume ratios of EtOH so that the CPS with a higher MW was precipitated at a lower EtOH ratio. In addition to MW, the total phenolic content of CPS was also found to increase with increase in EtOH ratio. All CPS fractions from HWE, AKE and acid extraction (ACE) showed moderate antioxidant activities. In particular, both radical scavenging and reducing power showed a negative correlation to intrinsic viscosity (or MW) (TEAC, R2= 0.65, p<0.05; FRAP, R2=0.61, p<0.05), radical scavenging activity showed a positive correlation to protein content (R2= 0.91, p<0.05), and reducing power showed a positive correlation to total phenolic content (R2=0.83, p<0.05). The results suggested that the antioxidant activities of CPS were attributable to the protein and total phenolic contents, and the activities of CPS were mostly higher with lower MW.
The CPS isolated from the AKE extract of A. ostoyae by precipitation with 1 volume of EtOH was further purified by ion exchange chromatography (IEC) and size exclusion chromatography (SEC), yielding two PS fractions, AKPS1V-1 (66.6 kDa) and AKPS1V-2 (15.3 kDa), both of which composed mainly of glucose AKPS1V-1 (100%) and AKPS1V-2 (>86%). AKPS1V-2 exhibited a more significant antioxidant potential than AKPS1V-1 due probably to its lower MW. The linkage and structural characteristics of AKPS1V-2 were further identified, consisting of (1→6)-β -D-glucose and (1→3)-β-D-glucose and (1→3)-α-D-galactose residues in a ratio of 3:1:1 in the main chain branched with (1→3)-β-D-glucose residue. Although PS from many mushrooms have been reported to have significant antioxidant activities, these PS are mostly crude or partially purified PS fractions with proteins and low-MW pigments. To identify the constituents actually responsible for the antioxidant activities of CPS from mushrooms clearly and quantitatively, we further tested the CPS from three important medicinal mushrooms, Lentinus edodes, Grifola frondosa and Trametes versicolor. CPS was isolated from the hot water extract of each mushroom by EtOH precipitation contained 15-30% (w/w) total proteins and 2-5% phenolics. The CPS were fractionated by IEC and the neutral and slightly ionic PS fractions (eluted with 0 and 0.1 M NaCl) contained 70-80% carbohydrate with low protein and phenolic contents, while the ionic PS fractions (eluted with 0.3 and 0.7 NaCl) had higher contents of protein (20-70%) and phenolics (2-13%). The antioxidant activities of all PS fractions showed significant correlations to the total phenolic content and protein contents but not to the carbohydrate content. Purified PS free of phenolics and proteins had no significant activities. Therefore, phenolic and protein components instead of carbohydrates were mainly responsible for the antioxidant activities of mushroom CPS. This thesis presents new research results and findings which will make valuable contributions to the development and understanding of the processes and conditions for effective extraction, isolation and purification of PS from edible and medicinal mushrooms and their influences on the properties and bioactivities of PS and the property-bioactivity

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