Sensitive Amperometric Oxalate Biosensor for Food Analysis

Stjepan Milardović¹*, Zorana Grabarić¹ and Božidar S. Grabarić<sup>2

1</sup> University of Zagreb, Faculty of Chemical Engineering and Technology, Marulićev trg 19, HR-10000 Zagreb, Croatia
²University of Zagreb, Faculty of Food Technology and Biotechnology, Pierottijeva 6, HR-10000 Zagreb, Croatia

Article history:
Received March 1, 2000
Accepted May 17, 2000

Key words:
amperometric biosensor, graphite electrode modified with nickel, iron, ruthenium, hexacyanometallate, oxalate oxidase, determination of oxalate, food samples

Summary:
An oxalate biosensor developed by covalent immobilisation of oxalate oxidase enzyme on the top of ruthenium, nickel and iron hexacyanometallate (Ru-FeNiHCM) modified graphite electrode is described. It was noticed that the addition of ruthenium to FeNiHCM layer results in better catalytic efficiency for reduction of hydrogen peroxide at pH=2. To obtain the optimal response of biosensor the influence of several experimental parameters in batch mode, such as working potential, pH and linear concentration range, was examined. Enzymatically produced H₂O₂ was measured at –50 mV vs. Hg
Hg₂Cl₂
3 M KCl electrode in succinic buffer (pH=3.6). The linear concentration range was up to 100 µM with squared correlation coefficient R²=0.9985. Sensitivity of biosensor in this range was 43.2 nA/(µM cm²). Oxalate was determined in real samples of spinach after extraction in water, EDTA and HCl and in samples of different brands of beer. During 11 weeks biosensor sensitivity decreased only 15 % of the initial sensitivity, which means that the biosensor lifetime is much longer than three months.  

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