HPLC-UVによる穀類中の遊離アスパラギン分析法の開発

摘要

アクリルアミドの前駆体である遊離アスパラギン（Asn) の穀類中の分析法を開発した.試料中から5% (w/v)トリ クロロ酢酸溶液でAsnを抽出し，ポリマー系逆相固相 カートリッジを用いて精製した後，ダンシル誘導体化を行 い,HPLC-UVで測定した.分析カラムはODSを用い， 0.01 mol/L酢酸アンモニウム溶液とアセトニトリルのダラ ジェント溶出により行った.検量線は，0.5〜1OO μg/mL の範囲で良好な直線性を示した.片栗粉，うるち米粉，小 麦全粒粉の3試料を用いて添加回収実験を行った結果，平 均回収率は97.7〜102.6%，併行精度（RSDr)は0.8〜 2.0%,室内再現精度（RSDwr)は1.4〜6.2%の良好な結 果が得られた.本法による定量限界は，片栗粉で13mg/ kg,うるち米粉で4mg/kgであった.片栗粉，うるち米 粉.小麦全粒粉を含む15試料について，本法とアミノ酸 自動分析計による定量値の比較を行ったところ，いずれの 試料においても定量値は同等であった.%A novel analytical method was developed for the determination of free asparagine (Asn) ， which is a precursor of acrylamide, in grains. Asn was extracted from a sample with 5% (w/v) aqueous trichloroacetic acid solution, and the crude extract was cleaned up using a reversed-phase solid-phase cartridge. The cleaned extract was derivatized with dansyl chloride and analyzed by HPLC-UV. HPLC separation was performed by gradient elution on a ODS column using 0.01 mol/L ammonium acetate and acetonitrile mixture as the mobile phase. The calibration curve was linear in the range of 0.5—100 μg/mL. The mean recoveries from potato starch, non-glutinous rice flour and whole wheat flour ranged from 97.7 to 102.6%, the repeatability (RSDs) ranged from 0.8 to 2.0%, and the within-laboratory reproducibility (RSDwr) ranged from 1.4 to 6.2%. Limits of quantitation (LOQs) were 13 mg/kg for potato starch and 4 mg/kg for non-glutinous rice flour. The quantitative values obtained for about 15 kinds of grains using this developed method were approximately equal to those obtained with an automatic amino acid analyzer.