Validation of a reversed-phase high-performance liquid chromatographic method for the determination of free amino acids in rice using l-theanine as the internal standard.

The study presents the validation results of the method carried out for analysis of free amino acids (FAAs) in rice using l-theanine as the internal standard (IS) with o-phthalaldehyde (OPA) reagent using high-performance liquid chromatography-fluorescence detection. The detection and quantification limits of the method were in the range 2-16μmol/kg and 3-19μmol/kg respectively. The method had a wide working range from 25 to 600μmol/kg for each individual amino acid, and good linearity with regression coefficients greater than 0.999. Precision measured in terms of repeatability and reproducibility, expressed as percentage relative standard deviation (% RSD) was below 9% for all the amino acids analyzed. The recoveries obtained after fortification at three concentration levels were in the range 75-105%. In comparison to l-norvaline, findings revealed that l-theanine is suitable as an IS and the validated method can be used for FAA determination in rice.