超高效液相色谱-串联质谱法测定茶叶中游离氨基酸成分
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摘要
建立了使用超高效液相色谱-串联质谱(UHPLC-MS/MS)高效、快速直接测定茶叶中游离氨基酸的方法。通过对质谱、色谱条件及氨基酸提取条件的优化,以含0.2%(体积分数)甲酸的5 mmol/L乙酸铵水溶液和甲醇为流动相进行梯度洗脱,在电喷雾离子(ESI)源正离子扫描模式下检测,通过UHPLC-MS/MS测定,共解析了茶叶中的20种氨基酸。结果表明,茶氨酸(Thea)、Arg、Asn和Asp在50～500μg/L范围内线性关系良好,其他氨基酸在10～250μg/L范围内线性关系良好,相关系数均大于0.99;加标回收率为92.3%～109.2%,相对标准偏差为2.00%～9.88%,检出限为0.001～0.011 mg/L,定量限为0.010～0.053 mg/L。该方法灵敏、准确,具有良好的重复性和稳定性,可有效检测出茶叶中的20种氨基酸及氨基类成分。
To efficiently and quickly detect free amino acid components in tea, a method using ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) was developed. With the optimization of mass spectrometry, chromatographic conditions, and amino-acid extraction conditions, a total of 20 free amino acids were identified using 5 mmol/L ammonium acetate aqueous solution containing 0.2%(v/v) formic acid and methanol as mobile phases for gradient elution and detected by electrospray ionization(ESI) and positive-ion scanning. The results showed that all calibration curves expressed good linearities. Theanine(Thea), Arg, Asn, and Asp were in the range of 50-500 μg/L. The other amino acids were in the range of 10-250 μg/L with all correlation coefficients≥0.99. The average recoveries were between 92.3% and 109.2%. The relative standard deviation(RSDs) were between 2.00% and 9.88%. The limits of detection(LODs) were 0.001-0.011 mg/L, and the limits of quantification(LOQs) were 0.010-0.053 mg/L. The method is sensitive, accurate, and has good repeatability and stability. The method can effectively detect 20 types of amino acids and amino components in tea leaves from the samples of green tea, white tea, and black tea.
To efficiently and quickly detect free amino acid components in tea, a method using ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) was developed. With the optimization of mass spectrometry, chromatographic conditions, and amino-acid extraction conditions, a total of 20 free amino acids were identified using 5 mmol/L ammonium acetate aqueous solution containing 0.2%(v/v) formic acid and methanol as mobile phases for gradient elution and detected by electrospray ionization(ESI) and positive-ion scanning. The results showed that all calibration curves expressed good linearities. Theanine(Thea), Arg, Asn, and Asp were in the range of 50-500 μg/L. The other amino acids were in the range of 10-250 μg/L with all correlation coefficients≥0.99. The average recoveries were between 92.3% and 109.2%. The relative standard deviation(RSDs) were between 2.00% and 9.88%. The limits of detection(LODs) were 0.001-0.011 mg/L, and the limits of quantification(LOQs) were 0.010-0.053 mg/L. The method is sensitive, accurate, and has good repeatability and stability. The method can effectively detect 20 types of amino acids and amino components in tea leaves from the samples of green tea, white tea, and black tea.
引文
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[21] GB 8303-2013
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[25] GB/T 23193-2017
[2] Zhao H T.Amino Acid and Biotic Resources,1990(3):47 赵和涛.氨基酸杂志,1990(3):47
[3] Wan X C.Tea Biochemistry.3rd ed.Beijing:Agriculture Press of China.2003:32 宛晓春.茶叶生物化学.3版.北京:中国农业出版社,2003:32
[4] GB 8314-2013
[5] Liu J F,Gao J J,Tian A L,et al.Chinese Journal of Tropical Crops,2017,38(2):283 刘建福,高俊杰,田奥磊,等.热带作物学报,2017,38(2):283
[6] Zhao X,Li X S,Han H,et al.Amino Acids and Biotic Resources,2016,38(4):17 赵璇,李新生,韩豪,等.氨基酸和生物资源,2016,38(4):17
[7] Qi G N,Tian H,Liu A L,et al.Journal of Tea Science,2004,24(4):266 齐桂年,田鸿,刘爱玲,等.茶叶科学,2004,24(4):266
[8] Chen D,Ye X H,Yu Y,et al.Tea in Fujian,2014(4):24 陈丹,叶小辉,余滢,等.福建茶叶,2014(4):24
[9] Jiang T,Feng Y J,He X C,et al.Chemical Research and Application,2012,24(7):1159 姜涛,冯永建,何学超,等.化学研究与应用,2012,24(7):1159
[10] Yu H,Mou S F.Chinese Journal of Analytical Chemistry,2005,33(3):398 于泓,牟世芬.分析化学,2005,33(3):398
[11] Wang F H.Food Research and Development,2018,39(1):141 王富花.食品研究与开发,2018,39(1):141
[12] Li N N,Liu Y,Zhao Y,et al.Food Anal Method,2016,9 (5):1307
[13] Tu Y F,Yang X F,Zhang S K,et al.Chinese Journal of Chromatography,2012,30(2):184 涂云飞,杨秀芳,张士康,等.色谱,2012,30(2):184
[14] Huang Y F,Hu J.Chinese Journal of Chromatography,2010,28(6):615 黄翼飞,胡静.色谱,2010,28(6):615
[15] Deng Q X,Su M L,Huang C Z,et al.Journal of Analytical Science,2014,30(2):263 邓其鑫,苏明亮,黄朝章,等.分析科学学报,2014,30(2):263
[16] Wang X,Chi Y M,Kang A.Chinese Journal of Chromatography,2014,32(12):1326 王星,池玉梅,康安.色谱,2014,32(12):1326
[17] Zheng Z,Sun Q,Shi Y W,et al.Chinese Journal of Chromatography,2015,33(3):309 郑重,孙琦,石永伟,等.色谱,2015,33(3):309
[18] Li P F,Tao B B,Zhang X D,et al.Chinese Journal of Analytical Chemistry,2013,41(9):1347 李鹏飞,陶蓓蓓,张绪得,等.分析化学,2013,41(9):1347
[19] Sun X.[Master Dissertation].Nanchang:Nanchang University,2015 孙玄.[硕士学位论文].南昌:南昌大学,2015
[20] Zhang L,Zeng Z D,Ye G Z,et al.Chinese Journal of Chromatography,2014,32(8):804 张磊,曾仲大,叶国柱,等.色谱,2014,32(8):804
[21] GB 8303-2013
[22] Chen Y B,Ma Z J,Li W D,et al.Amino Acids & Biotic Resources,2011,33(4):42 陈永波,马作江,李卫东,等.氨基酸和生物资源,2011,33(4):42
[23] Du Y Y,Liang Y R,Wang H,et al.J Hortic Sci Biotechnol,2006,81(5):809
[24] Feng L.[Master Dissertation].Hefei:Anhui Agricultural University,2014 冯琳.[硕士学位论文].合肥:安徽农业大学,2014
[25] GB/T 23193-2017