High-performance liquid chromatography analysis of naturally occurring d-amino acids in sake

We measured all of the d- and l-amino acids in 141 bottles of sakes using HPLC. We used two precolumn derivatization methods of amino acid enantiomer detection with o-phthalaldehyde and N-acetyl-l-cysteine, as well as (+)-1-(9-fluorenyl)ethyl chloroformate/1-aminoadamantane and one postcolumn derivatization method with o-phthalaldehyde and N-acetyl-l-cysteine. We found that the sakes contained the d-amino acids forms of Ala, Asn, Asp, Arg, Glu, Gln, His, Ile, Leu, Lys, Ser, Tyr, Val, Phe, and Pro. We were not able to detect d-Met, d-Thr d-Trp in any of the sakes analyzed. The most abundant d-Ala, d-Asp, and d-Glu ranged from 66.9 to 524.3 μM corresponding to relative 34.4, 12.0, and 14.6% d-enantiomer. The basic parameters that generally determine the taste of sake such as the sake meter value (SMV; “Nihonshudo”), acidity (“Sando”), amino acid value (“Aminosando”), alcohol content by volume, and rice species of raw material show no significant relationship to the d-amino acid content of sake. The brewing water (“Shikomimizu”) and brewing process had effects on the d-amino acid content of the sakes: the d-amino acid contents of the sakes brewed with deep-sea water “Kaiyoushinosousui”, “Kimoto yeast starter”, “Yamahaimoto”, and the long aging process “Choukijukusei” are high compared with those of other sakes analyzed. Additionally, the d-amino acid content of sakes that were brewed with the adenine auxotroph of sake yeast (“Sekishoku seishu kobo”, Saccharomyces cerevisiae) without pasteurization (“Hiire”) increased after storage at 25 °C for three months.