Determination of Free Amino Acid Compositon in Phrynium rheedei by Pre-column Derivatization HPLC and Flavor Quality Evaluation
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摘要: 建立柱前衍生化HPLC法测定柊叶游离氨基酸的方法，对其营养价值和风味进行评价。柱前衍生化HPLC法以异硫氰酸苯酯为衍生试剂，色谱条件为：C18色谱柱（4.6 mm×250 mm，5μm），醋酸钠缓冲液-乙腈（93：7）和乙腈-水（4：1）为流动相，进行梯度洗脱，流速为1.0 mL/min，柱温40℃，检测波长为254 nm。结果表明，18种氨基酸均呈现良好的线性关系（R2>0.9990），柊叶游离氨基酸的平均回收率分别为98.3%~105.6%（RSD为0.63%~3.11%，n=6）。所建立的方法稳定、灵敏、重现性好，可用于柊叶游离氨基酸的定量分析。柊叶含有18种氨基酸及8种人体必需氨基酸，必需氨基酸总量为38.85%，接近于WHO/FAO模式值（40.0%），各种必需氨基酸组分达到WHO/FAO氨基酸成人模式的需求；鲜味氨基酸和甜味氨基酸的含量分别为10.73和5.20 mg/g，占呈味氨基酸总含量的22.58%和46.59%；鲜味氨基酸Asp和甜味氨基酸Thr的含量较高，分别为3.75 mg/g和3.33 mg/g，鲜味氨基酸Asp和芳香族氨基酸Cys的RCT值分别为125.00和20.50，对风味贡献最高。总之柊叶氨基酸种类齐全，比例合理，呈味氨基酸含量丰富，具有极大开发价值。Abstract: The method for the determination of free amino acids in Phrynium rheedei was established,and its nutritional value and flavor was evaluated. HPLC with phenylisothiocyanateas derivatization reagent,chromatographic conditions were as follows: Using C18 column(4.6 mm×250 mm,5 μm),with sodium acetate buffer-acetonitrile(93∶7)and acetonitrile-water(4∶1)as mobile phrase in gradient elution,flow rate of 1.0 mL/min,column temperature of 40 ℃,the detection wavelength was set at 254 nm.The results showed that the calibration curves of 18 kinds of amino acids showed good linearity within their test ranges(R2>0.9990),and the average recovery rates of the free amino acids in Phrynium rheedei ranged from 98.3%~105.6%(RSD0.63%~3.11%,n=6). The method was stable,sebsitive,good reproducibility,and could be used to determine the content of free amino acids in Phrynium rheedei. The tested results showed that there were 18 amino acids and 8 essential amino acids in Phrynium rheedei. The total content of essential amino acids was 38.85%,which closed significantly the WHO/FAO model value(40.0%),the components of essential amino acids met the requirements of WHO/FAO amino acid model of adult. The contents of delicious amino acids and sweet amino acids were 10.73 mg/g and 5.20 mg/g,which accounted for 22.58% and 46.59% of the total respectively. The contents of Asp and Thr were more,were 3.75 mg/g and 3.33 mg/g respectively. The RCT values of Asp and Cys were 125.00 and 20.50 respectively,had more the contribution to flavour. In a word,Phrynium rheedei contains variety of amino acid,which is reasonable proportion,the content of its flavor amino acids is rich,it has broad development prospects.
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