Analytical Methods for Sake Characterization

Sake meter value, sake acidity, and amino acid levels were measured as previously described⁶ . Organic acids were quantified using HPLC with a Shim-pack SCR-102H column (Shimadzu, Kyoto, Japan). Aroma components were quantified using headspace GC with a J & W DB-WAX capillary column (30 m × 0.32 mm internal diameter × 0.50 μm film thickness; Agilent Technologies, Santa Clara, California, USA). Ethanol concentration was measured using an ethanol analyser (RIKEN KEIKI, Tokyo, Japan). Sugars were determined by monitoring post-column derivative reducing sugars separated using a Prominence reducing-sugar HPLC analytical system (Shimadzu) equipped with a fluorescence detector. Sake fermented with MC87-46 and K901 was separated on a Shim-pack 4.0 × 250-mm ISA-07/S2504 column (Shimadzu) with a linear gradient of 0.1 M potassium borate buffer (pH 8.0) and 0.4 M potassium borate buffer (pH 9.0) for 140 min at a flow rate of 0.6 mL/min^{37 (link),38 (link)}. Sugars were also lyophilized, trimethylsilylated, and analysed using a GCMS-QP2010 (Shimadzu) equipped with a J & W DB-5MS capillary column (30 m × 0.25 mm internal diameter × 0.25 μm film thickness; Agilent Technologies)^{39 (link)}.