| Summary: | This paper describes the design, development and characterisation of an electrochemical biosensor for the measurement of linoleic and α-linolenic acid, as representative free polyunsaturated fatty acids (PUFAs), that may be implicated in food safety and food quality. Initial cyclic voltammetric studies were performed with solutions that contained enzyme-generated hydroperoxides of the two PUFAs. These were examined with plain screen-printed carbon electrodes (SPCEs) and screen-printed carbon electrodes containing the electrocatalyst cobalt phthalocyanine (CoPC). The electrocatalytic oxidation peaks obtained with the latter occurred at potentials about 300 mV lower than the those obtained by direct oxidation with the plain SPCEs and were better defined; as these attributes would lead to better selectivity and sensitivity for fatty acid determinations, the CoPC-SPCEs were used in the fabrication of amperometric biosensors. The enzyme lipoxygenase (LOX) was immobilised on the surface of these devices using the crosslinking agent glutaraldehyde. These biosensors were optimised for the measurement of linoleic and α-linolenic acid using amperometry in stirred solution; the optimum conditions were deduced by studying the effect of enzyme loading, pH and temperature on the amperometric responses. These responses were examined over the concentration range 2.0 to 20 µM and the results indicated that the following conditions were optimal: LOX loading 15 units; pH 8.0; temperature 37 °C. Low concentration calibration studies were performed with the two PUFAs and it was shown that the steady state currents were linear between 0.2 and 10 µM for linoleic acid and 0.2 and 10 µM for α-linolenic acid; the detection limits were 24 and 100 nM, respectively. The precision (coefficient of variation, <i>n</i> = 6) was 5.3% for α-linoleic acid and 3.3% for linoleic acid, which were calculated from the steady state current following additions (<i>n</i> = 6) of 0.2 µM PUFA. These results demonstrate that the novel amperometric biosensor holds promise for determining whether foods contain acceptable levels of free fatty acids.
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