Exploring microbiological transformation of glutamate to γ-aminobutyric acid unleashed by a potential Lactobacillus spp. isolated from commercial cheese samples

The present study investigated the microbiological transformation of glutamate to γ-aminobutyric acid (GABA) using bacterial strains isolated from waste cheese. The waste cheese samples were collected from various sources, including local dairy shops and pizza outlets in Lahore, as well as from the industrial area of Sahiwal, Pakistan. From a total of 40 bacterial isolates, 14 were screened for their ability to produce GABA. Among them, isolates ISL-7 (32.7 mM) and ISL-15 (29.4 mM) exhibited the highest GABA production under submerged fermentation. ISL-7 was identified as Lacticaseibacillus casei through 16S rDNA gene amplification, while ISL-15 was confirmed as bacillus shaped cheese isolate via scanning electron microscopy. To improve GABA yield, various cultural conditions were optimized, including glutamate concentration (0.5 %), initial pH (6.5), incubation period (48 h), and inoculum size (8 %). The thermophilic behavior of the isolates and the influence of stimulatory compounds such as casamino acid, thiamine HCl, n-butanol, and glycerol on glutamate decarboxylase (GAD) activity were investigated. Maximum GAD activity was recorded at 35 °C, with casamino acid serving as the most effective stimulator, reaching 113.1 U/ml in ISL-7 and 100.4 U/ml in ISL-15. The effects of micro minerals (MgSO₄·7H₂O, MnCl₂, K₂Cr₂O₇, KI) and macro minerals (NH₄NO₃, NaNO₃, CaCl₂, KH₂PO₄) on GABA production were also evaluated. The optimized addition of MgSO₄·7H₂O (4 mM), KI (0.5 mM), CaCl₂ (0.4 %), and KH₂PO₄ (0.3 % for ISL-7 and 0.2 % for ISL-15) enhanced GABA production to 244 ± 1.25 mM (25.16 ± 0.13 g/L) and 232 ± 1.27 mM (23.91 ± 0.13 g/L) respectively. Overall, GABA production increased 7.46-fold in ISL-7 and 7.89-fold in ISL-15 (p ≤ 0.05). ANN was employed on the GABA production to establish a valid correlation between the experimental and predicted results by the superior isolate of Lactobacillus casei. Gas chromatography–mass spectrometry (GC–MS) analysis validated the presence of GABA in the culture broth.