Characterization of diamine oxidase-based biosensors for biogenic amines detection assembled onto functionalized SiO₂ substrates using aliphatic and aromatic di-aldehydes as crosslinkers

Abstract

Development of two biosensors to detect biogenic amines was carried out using immobilized diamine oxidase from Pisum sativum on 3-APTMS-functionalized SiO2 substrates with either glutaraldehyde or terephthalaldehyde as crosslinkers. The assembly process was validated by FT-IR, SEM-EDX, and AFM. Biosensor responses to biogenic amines were c.a. 3-fold lower compared to the free enzyme kinetics as assessed by UV/VIS spectrophotometry. The calculated initial velocities (ΔAbs·min-1) for glutaraldehyde and terephthalaldehyde biosensors in the given order were (3.2 ± 0.68) × 10–2, (0.48 ± 0.09) × 10–2 for putrescine; (4.6 ± 1.2) × 10–2, (0.35 ± 0.47) × 10–2 for cadaverine; (5.8 ± 2.4) × 10–3, (19.0 ± 155) × 10–3 for spermidine, and (3.0 ± 2.0) × 10–4, (0.0 ± 7) × 10–4for histamine. The 2-way t-tests indicated that the crosslinker type affected the catalytic activity of immobilized diamine oxidase as the responses of glutaraldehyde biosensors to putrescine and cadaverine were significantly higher than those of the terephthalaldehyde. Molecular Docking simulations showed that glutaraldehyde has a stronger affinity for DAO’s lysine and arginine residues while terephthalaldehyde to phenylalanine.