Simultaneous determination of 20 amino acids in Sichuan black tea by the method of online pre-column OPA derivatization HPLC
To establish a method of online pre-column derivatization HPLC for simultaneous determina-tion of 20 amino acids in different varieties of Sichuan black tea,and to evaluate their correlation with taste quality,Ortho phthalaldehyde(OPA)was used as a derivative reagent,and derivatized in the automatic sampler of liquid chromatograph.The derivatives of amino acids were separated by gradient elution with phosphate buffer(0.016 mol/L,pH=8.2)and methanol-acetonitrile-water(4.5∶4.5∶1,V∶V∶V)under the conditions of 1 mL/min and column temperature 50 ℃ in Hypersil BDS C18 chroma-tographic column(150 mm ×4.6 mm,3 µm),and were detected in a fluorescence detector with exci-tation wavelength 340 nm and emission wavelength 450 nm,then the correlation between amino acid contents of different flavor attributes and the taste of black tea was compared through correlation analy-sis.Results showed that:The 20 amino acids showed a good linear relationship with their respective peak areas from 0.195 to 2.584 μg/mL with correlation coefficients ranging from 0.995 4 to 0.999 9,their detection limit was between 0.006～0.036 µg/mL,the quantification limit was 0.020～0.184μg/mL,and the spiked recovery rate ranged from 90.40％to 99.13％.With the exception of histi-dine that was not detected in black teas prepared with Sichuan populations tea plant and Meizhan tea plant,other amino acids were dectected in 5 black teas with a content of 27.33～65.29 mg/g,and the highest content of variety was Huangjinya black tea.Among the amino acid components,the high-est content was theanine,followed by aspartic acid and glutamic acid.Through relevant analysis,it showed that there was a significant or extremely significant positive correlation between the content of fresh amino acids,bitter amino acids,sweet amino acids,total amino acids to the scored of freshness,sweetness and overall taste,and they played an important role in freshness and intensity of the taste of black tea.The pre-column OPA derivatization HPLC has high precision and recovery rate in detecting amino acid components,and has the characteristics of low cost,fast and accurate.It can be used for the rapid determination of 20 amino acid components in black tea.
Ortho phthalaldehydepre-column derivatizationSichuan black teaamino acid
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