Solidspec 3700duv

The cell wall polysaccharides were prepared and fractionated as described by Yoshioka et al. (2011) [20] , using the method based on that of Huber and O'Donoghue (1993) [21] (link). Briefly, 2 g of frozen flesh was homogenized in a 50-mL centrifuge tube with 8 mL of ice cold 0-°C ethanol using a homogenizer AHG-160A (AS ONE Corporation, Osaka, Japan) and centrifuged at 1600 g for 10 min. The insoluble pellet was washed twice with 10 mL cold 80 % ethanol, treated with 2 mL Tris-buffered (pH 8.0) phenol for 1 h at room temperature, and precipitated with 10 mL of ethanol at −30 °C. The ethanol precipitate was extracted with 10 mL of chloroform/methanol (1:1, v/v) for 30 min at room temperature, and washed once with 10 mL acetone. The precipitate was collected at each step by centrifugation at 1600 g for 10 min. The alcohol-insoluble solid (AIS) was incubated with 10 mL water at room temperature for 12 h. The supernatants were used for the water-soluble extract. The content of galacturonic acid (GalUA) in the extract was determined using the m-phenylphenol colorimetric method [22] (link). The absorbance of the solution at 520 nm was measured with SolidSpec-3700DUV (Shimadzu Corporation, Kyoto, Japan).