Enzymatic biosensor based on entrapment of d-amino acid oxidase on gold nanofilm/MWCNTs nanocomposite modified glassy carbon electrode by sol-gel network: Analytical applications for d-alanine in human serum.

Sensing and determination of d-alanine is studied by using an enzymatic biosensor which was constructed on the basis of d-amino acid oxidase (DAAO) immobilization by sol-gel film onto glassy carbon electrode surface modified with nanocomposite of gold nanofilm (Au-NF) and multiwalled carbon nanotubes (MWCNTs). The Au-NF/MWCNT nanocomposite was prepared by applying the potentiostatic technique for electrodeposition of Au-NF on the MWCNT immobilized on glassy carbon electrode surface. The modified electrode is investigated by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), linear sweep voltammetry (LSV) and cyclic voltammetry(CV) techniques. The linear sweep voltammetry was used for determination of d-alanine and the results showed an excellent linear relationship between biosensor response and d-alanine concentration ranging from 0.25μM to 4.5μM with correction coefficient of 0.999 (n=20). Detection limit for the fabricated sensor was calculated about 20nM (for S/N=3) and sensitivity was about 56.1μAμM-1cm-2. The developed biosensor exhibited rapid and accurate response to d-alanine, a good stability (4 weeks) and an average recovery of 98.9% in human serum samples.