O phthaldialdehyde reagent

Accumulated GABA in the culture medium was measured as follows. First, the culture broth was separated from cells by centrifugation (10000 × g for 10 min at 4°C), and the supernatant was filtered using a 0.2 μm membrane filter (Pall Acrodisc^®Syringe Filters, USA). Then, GABA concentrations of the supernatant were determined by pre-column derivatization with o-phthaldialdehyde reagent (97% purity, HPLC grade, Sigma, USA) [39 (link)]. Separations were performed by HPLC (Agilent, 1100 Series, USA) in conjunction with a Hypersil ODS2 C₁₈ column (Dalian Elite Analytical Instruments Co. Ltd., China) at 30°C, and the wavelength of the UV detector was 338 nm. The mobile phase was a mixture of sodium acetate (0.05 M, 500 mL), methanol (490 mL, chromatographic grade, Tianjin Guangfu Fine Chemical Co., Ltd., China) and tetrahydrofuran (10 mL chromatographic grade, Tianjin Fuyu Fine Chemical Co., Ltd., China), then the mixture solution was filtered using a 0.2 μm membrane filter and degassed by sonication [12 (link),40 (link)]. The flow rate was 1 ml min^-1. Calibration curves were obtained based on several concentrations of a GABA standard (Sigma, USA), and the coefficient of determination (r²) was greater than 0.999.

Triton X-100, Tetraethyl orthosilicate (TEOS), (3-Aminopropyl) triethoxysilane (APTS), sodium alginate, chlorhexidine diacetate, sodium acetate trihydrate, piperazine, gentamicin sulphate, phosphate buffer solution (PBS) tablets, o-phthaldialdehyde reagent were purchased from Sigma-Aldrich, UK.
Cyclohexane, 1-hexanol, ammonium hydroxide 35 %, acetonitrile, ethanol, methanol, glacial acetic acid and 1-propanol were purchased from Fishers, UK. All reagents were stored according to manufacturer's guidelines and used as received.