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A High-Performance Liquid Chromatographic Method for Simultaneous Determination of 21 Free Amino Acids in Tea

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Abstract

Free amino acids are closely related to the savory taste and beneficial effects of tea, and high-performance liquid chromatography (HPLC) is the most widespread analytical approach for simultaneous determination of free amino acids in tea. However, the reported HPLC methods have drawbacks such as long run times, low resolution, or poor efficiency. In this study, a special amino acid analysis column was used to separate and verify 21 amino acids including l-theanine, the predominant amino acid in tea. The mobile phases, including the sodium acetate and acetic acid concentration in buffer B, and the pH and concentration of sodium acetate in buffer A were optimized. The elution gradients were optimized using DryLab 2000 Plus software. In this way, an online o-phthaldialdehyde precolumn derivatization HPLC-fluorescence detection method was developed for simultaneous determination of 21 amino acids. Comparison to other HPLC methods for simultaneous determination of free amino acids in tea showed that our method is easy (automated derivatization), quick (30 min), inexpensive, and green (using a minimum of solution). It has good resolution (≥1.8) and high selectivity (interpark time ≥ 0.5 min). Free amino acids in six tea samples were analyzed. This work provides an HPLC method to simultaneously measure 21 amino acids in tea and potential in other food products.

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Abbreviations

ACN:

Acetonitrile

Ala:

Alanine

Arg:

Arginine

Asn:

Asparagines

Asp:

Aspartic acid

Cys:

Cystine

DAD:

Diode array detector

EDTA:

Ethylenediaminetetraacetic acid disodium salt dihydrate

FLD:

Fluorescence detection

GABA:

γ-Amino butyric acid

Gln:

Glutamine

Glu:

Glutamic acid

Gly:

Glycine

HPLC:

High-performance liquid chromatography

His:

Histidine

Ile:

Isoleucine

Leu:

Leucine

LOD:

Limit of detection

LOQ:

Limit of quantification

Lys:

Lysine

MeOH:

Methanol

Met:

Methionine

OPA:

o-Phthaldialdehyde

PA:

Peak area

Phe:

Phenylalanine

Pro:

Proline

RSD:

Relative standard deviation

Rs:

Resolution

RT:

Retention time

Ser:

Serine

NaAc:

Sodium acetate

THF:

Tetrahydrofuran

Thr:

Threonine

TEA:

Triethylamine

Trp:

Tryptophan

Tyr:

Tyrosine

Val:

Valine

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Acknowledgments

This work was supported by grants from The National Natural Science Foundation of China (Grant No. 31160174), the Key Program for Applied Basic Researches of Yunnan Province, China (Grant No. 2009CC005), and the Natural Science Foundation of Yunnan Province, China (Grant No. 2010ZC084).

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Authors and Affiliations

  1. College of Pu-erh Tea, Yunnan Agricultural University, Kunming, Yunnan, 650201, China

    Ming Zhao, Yan Ma, Lei-li Dai, Dong-lian Zhang, Jia-hua Li, Wen-xia Yuan, Ya-li Li & Hong-jie Zhou

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  1. Ming Zhao
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  2. Yan Ma
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  3. Lei-li Dai
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  4. Dong-lian Zhang
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  5. Jia-hua Li
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  7. Ya-li Li
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  8. Hong-jie Zhou
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Correspondence to Ming Zhao or Hong-jie Zhou.

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Zhao, M., Ma, Y., Dai, Ll. et al. A High-Performance Liquid Chromatographic Method for Simultaneous Determination of 21 Free Amino Acids in Tea. Food Anal. Methods 6, 69–75 (2013). https://doi.org/10.1007/s12161-012-9408-4

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