高效液相色谱法同时测定咖啡中6种绿原酸
目的 建立咖啡中同时测定5-咖啡酰奎宁酸、3-咖啡酰奎宁酸、4-咖啡酰奎宁酸、3,4-二咖啡酰奎宁酸、4,5-二咖啡酰奎宁酸、3,5-二咖啡酰奎宁酸6 种绿原酸的高效液相色谱(high performance liquid chromatography, HPLC)检测方法, 以及测定产自云南普洱的铁皮卡品种咖啡在不同烘焙风味下的绿原酸含量规律。方法 样品经纯水超声提取2 次, 过滤后供HPLC 测定, 采用甲醇(A)和0.1%磷酸(B)作为流动相进行梯度洗脱采用C18 柱进行分离, 光电二极管阵列(photo-diode array, PDA)检测器最终检测。结果 本方法在1 h内完成了6 种绿原酸的分离分析。6 种绿原酸在0.10~100.0 mg/L 范围内具有良好的线性, 相关系数(r)≥0.999,检出限为0.05~0.10 mg/kg, 定量限为0.18~0.29 mg/kg。在2000、5000、10000 mg/kg 添加水平的加标回收率为96.3%~104.8%, 相对标准偏差(relative standard deviation, RSD)为0.58%~2.10%之间(n=6)。在生咖啡豆中总绿原酸含量最高, 意式烘焙(23 min, 250 ℃)最低, 5-咖啡酰奎宁酸、4-咖啡酰奎宁酸和4,5-二咖啡酰奎宁酸这3种绿原酸在生咖啡豆到极轻度烘焙(12 min, 230 ℃)这个阶段, 含量逐渐上升并达到含量最高, 由极轻度烘焙(12 min, 230 ℃)到意式烘焙(23min, 250 ℃)含量逐渐降低, 而3-咖啡酰奎宁酸、3,4-二咖啡酰奎宁酸和3,5-二咖啡酰奎宁酸3 种绿原酸从生咖啡豆至意式烘焙(23 min, 250 ℃)都呈下降趋势。结论 该方法快速、准确、灵敏, 适合测定绿原酸含量。
Objective To establish a method for determination of 5-caffeoylquinic acid, 3-caffeoylquinic acid,4-caffeoylquinic acid, 3,4-dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid, and 3,5-dicaffeoylquat sour acid in coffee by high performance liquid chromatography (HPLC) and detect chlorogenic acid content in ferric pickled coffee produced from Yunnan Pu'er in different roasting flavors. Methods The samples were extracted by pure water for 2 times. After filtration, the samples were determined by HPLC. Methanol (A) and 0.1% phosphoric acid (B) were used as mobile phase for gradient elution and samples were separated by C18 column, and photo-diode array (PDA)detector was used for final detection. Results This method had completed the separation and analysis of 6 kinds of chlorogenic acids within 1 h. The linear ranges of the six kinds of chlorogenic acids were 0.10~100.0 mg/L, with the correlation coefficients (r) ≥0.999. The limits of detection were 0.05~0.10 mg/kg and the limits of quantization were 0.18~0.29 mg/kg. The recoveries were between 96.3% and 104.8% with relative standard deviations (RSDs) of 0.58%~2.10% (n=6) at the spiked levels of 2000, 5000 and 10000 mg/kg. The total content of total chlorogenic acid in raw coffee beans was the highest, with the lowest in Italian roast (23 min, 250 ℃). The content of 5-caffeoylquinic acid, 4-caffeoylquinic acid and 4,5-dicaffeoylquinic acid increased gradually and reached the highest content at the stage of very mild roasting (12 min, 230 ℃), while decreased from very mild roast (12 min, 230 ℃) to Italian roast (23 min, 250 ℃). The content of 3-caffeoylquinic acid, 3,4-dicaffeoylquinic acid and 3,5-dicaffeoylquinic acid showed a downward trend from raw coffee beans to Italian roasted (23 min, 250 ℃). Conclusion The method is rapid, accurate and sensitive, which is suitable for the determination of chlorogenic acid content.
标题：高效液相色谱法同时测定咖啡中6种绿原酸
英文标题：Determination of 6 kinds of chlorogenic acid by high performance liquid chromatography
作者：
胡皓 昆明医科大学药学院
宋红坤 昆明医科大学药学院
王继良 昆明医科大学药学院
林涛 云南省农业科学院质量标准与检测技术研究所
刘兴勇 云南省农业科学院质量标准与检测技术研究所
邵金良 云南省农业科学院质量标准与检测技术研究所
中文关键词:绿原酸,高效液相色谱,烘焙,咖啡,
英文关键词:chlorogenic acid,high performance liquid chromatography,baking,coffee,
发表日期：2017-12-28
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