Determination of Amino Acids in Tunisian Animal Feedstuffs by HPLC-FLD

Abstract

The amino acids supplemented to animal feedstuffs cannot be determined directly. Thus,
we need to optimize the sample preparation step as well as the analytical method. In this
study, two hydrolysis mode were performed, one using 6 mol.L-1 hydrochloric acid at
105°C for 24 h and the other based on microwave radiation-induced hydrolysis.
Moreover, a chromatographic method was developed for simultaneous determination of
methionine and lysine in feedstuffs, using conventional High Performance Liquid
Chromatography equipment equipped with an automated precolumn derivatization
system using ortho-phthalaldehyde (OPA) / mercapto-2-ethanol (ME) reagents and the
fluorescence detector. Reversed-phase chromatography appears to be the preferred
method, with a C18 stationary phase and a gradient elution. Doehlert matrix was used to
get the optimal separation for 18 amino acids. System Suitability Test (SST), Linearity,
Limit of Detection (LOD), Limit of Quantification (LOQ), Repeatability and Specificity of
the method were studied. The LOD calculated were 93 and 9.4 pmol.μL-1 and the LOQ
were 173.5 and 36.2 pmol.µL-1 for methionine and lysine, respectively.