An amperometric enzyme sensor for determination of triglycerides (TGs) was constructed by co-immobilization of lipase (E.C.22.214.171.124), glycerol kinase (GK, E.C.126.96.36.199), and glycerol-3-phosphate oxidase (GPO, E.C.188.8.131.52) on Au/PEDOTPSS nanocomposite modified on screen-printed carbon electrodes (SPEs), then connecting it to potentiostat along with Ag/AgCl reference electrode and Pt auxiliary electrode. In this study, the experiments were divided into two parts. Firstly, the effects of precursor and EDOT concentration of Au/PEDOT-PSS polymerization on electrochemical response were studied. It revealed that Au nanoparticle from precursor enhanced the electrochemical response. Optimal conditions of modified SPEs for TGs detection were fabricated from 0.015 M EDOT and 6.25 mM precursor. Secondly, effects of printed layer of Au/PEDOT-PSS nanocomposite on electrochemical response. It was found that the optimum of printed layer was fabricated at 5 layers.
The biosensor measures the electrons generated from H₂O₂ under a potential of 0.4 V which in turn formed from triolein/TGs by co-immobilized lipase, GK and GPO. The concentration of triolein/TG was directly proportional to the current measured. The enzyme electrode showed optimum response when operated at 30±2°C in 0.1 M sodium phosphate buffer, pH 7.0. Linear range of this biosensor was found in the range from 0.1-6.0 mM. The sensitivity of this biosensor was 2.66μA/mM and the detection limit of 0.09 mM triolein was obtained. The response time of this biosensor was found within 30 s. However, the current response of this biosensor decreased to 50% of the initial current after 30 days of storage at 4°C. Moreover, the R.S.D. of this biosensor reproducibility was 9.75%