Applying gas chromatography to monitor extracellular free amino acids content in cultivation medium during lactic acid fermentation

Abstract

The aim of this work was the adaptation of a Gas Chromatographic-Flame Ionization Detector (GC-FID) method for detection and quantification of extracellular free amino acids in demineralized water, De Mann Rogosa Sharpe (MRS) medium and corn grits (CG) withdrawn during lactic acid fermentation. In order to analyze free amino acids by the GC-FID method it was necessary to convert free amino acids to volatile compounds. This was accomplished by derivatization of free amino acids with ethylchlor formate in aqueous medium followed by extraction of volatile free amino acid esters with chloroform. It was proven that the combination of derivatization and extraction procedure with developed GC-FID method gave accurate, reproducible and sensitive analytical results. Quantification of 15 (Ala, Gly, Val, Leu, Ile, Phe, Tyr, Trp, Asn, Met, Pro, Lys, His, Asp and Glu) out of 20 ethoxycarbonyl-ethyl esters of free amino acids in demineralized water and MRS medium was achieved by established methods. In corn grits medium all of the above mentioned 15 amino acids, except His, were quantified with this GC-FID method. The established method was efficiently verified in monitoring of extracellular free amino acid concentration during lactic acid production with Lactobacillus rhamnosus DSM 20021 T in MRS medium and Lactobacillus amylovorus DSM 20531 T in corn grits medium.