Sensitivity Intensified Ninhydrin-Based Chromogenic System by Ethanol-Ethyl Acetate: Application to Relative Quantitation of GABA

Abstract

Gamma-aminobutyric acid (GABA) is a functional metabolite in various organisms. Herein, a sensitivity intensified ninhydrin-based chromogenic system (SINICS), achieved by ethanol and ethyl acetate, is described for the reliable relative quantitation of GABA. A 2.9 mL SINICS kit comprises 1% ninhydrin, 40% ethanol, 25% ethyl acetate, and 35 μL 0.2 M sodium acetate buffer (pH 5.0). In practice, following the addition of a 0.1 mL sample to the kit, the chromogenic reaction is completed by heating at 70 °C for 30 min. The kit increased the color development sensitivity of L-glutamic acid and GABA, with the detection limits being reduced from 20 mM and 200 mM to 5 mM and 20 mM, respectively. The chromophore was stable for at least 2 h at room temperature, which was sufficient for a routine colorimetric analysis. The absorbance at 570 nm with the deduction of background directly represents the content of amino acid. For a proof-of-concept, the SINICS was adopted to optimize the GABA fermentation process of Levilactobacillus brevis CD0817. The results demonstrated that SINICS is an attractive alternative to the available ninhydrin-based colorimetric methods.