The development of an optical biosensor based on immobilization of 3-methyl-2-benzothiazolinone hydrazone (MBTH) in hybrid nafion/sol-gel silicate film and tyrosinase in chitosan film for the detection of phenolic compounds has been described. Tyrosinase was immobilized in chitosan film deposited on the hybrid nafion/sol-gel silicate film containing MBTH. The enzymatic oxidation product of phenolic compounds were stabilized through formation of adduct with MBTH to produce a maroon color adduct. The color intensity of adduct was found to increase proportionally with the increase of the substrate concentrations after 5min exposure. The linearity of the biosensor towards phenol, catechol and m-cresol were in the respective concentration range of 0.5-7.0, 0.5-10.0 and 1.0-13.0mg/L with detection limit of 0.18, 0.23 and 0.43mg/L, respectively. The biosensor shows a good stability for at least 3 months.
The stacked-film immobilization of 3-methyl-2-benzothiazolinone hydrazone (MBTH) in hybrid nafion/sol-gel silicate film and horseradish peroxidase (HRP) in chitosan, performed in order to allow the determination of phenolic compounds, was investigated via an optical method. The stacked films were deposited onto a microscope glass slide by a spin-coating technique. The quinone or free radical product formed by the enzymatic reactions of phenolic compounds interacts with MBTH to form azo-dye products, which can be measured spectrophotometrically at a wavelength of 500 nm. The color intensity of the product was found to increase in proportion to the phenolic concentration after 5 min of exposure. The response of the biosensor was linear over concentration ranges of 0.025-0.500, 0.010-0.070 and 0.050-0.300 mM for guaiacol, resorcinol and o-cresol, respectively, and gave detection limits of 0.010, 0.005 and 0.012 mM. The sensor exhibited good sensitivity and stability for at least two months.
The fabrication of an optical biosensor by using stacked films where 3-methyl-2-benzothiazolinone hydrazone (MBTH) was immobilized in a hybrid nafion/sol-gelsilicate film and laccase in a chitosan film for the detection of phenolic compounds wasdescribed. Quinone and/or phenoxy radical product from the enzymatic oxidation ofphenolic compounds was allowed to couple with MBTH to form a colored azo-dye productfor spectrophometric detection. The biosensor demonstrated a linear response to catecholconcentration range of 0.5-8.0 mM with detection limit of 0.33 mM and response time of10 min. The reproducibility of the fabricated biosensor was good with RSD value of 5.3 %(n = 8) and stable for at least 2 months. The use of the hybrid materials of nafion/sol-gelsilicate to immobilize laccase has altered the selectivity of the enzyme to various phenoliccompounds such as catechol, guaicol, o-cresol and m-cresol when compared to the non-immobilized enzyme. When immobilized in this hybrid film, the biosensor response onlyto catechol and not other phenolic compounds investigated. Immobilization in this hybridmaterial has enable the biosensor to be more selective to catechol compared with the non-immobilized enzyme. This shows that by a careful selection of different immobilizationmatrices, the selectivity of an enzyme can be modified to yield a biosensor with goodselectivity towards certain targeted analytes.
An optical biosensor based on glutamate dehydrogenase (GLDH) immobilized in a chitosan film for the determination of ammonium in water samples is described. The biosensor film was deposited on a glass slide via a spin-coating method. The ammonium was measured based on beta-nicotinamide adenine dinucleotide (NADH) oxidation in the presence of alpha-ketoglutaric acid at a wavelength of 340 nm. The biosensor showed optimum activity at pH 8. The optimum chitosan concentrations and enzyme loading were found to be at 2% (w/v) and 0.08 mg, respectively. Optimum concentrations of NADH and alpha-ketoglutaric acid both were obtained at 0.15 mM. A linear response of the biosensor was obtained in the ammonium concentration range of 0.005 to 0.5 mM with a detection limit of 0.005 mM. The reproducibility of the biosensor was good, with an observed relative standard deviation of 5.9% (n=8). The biosensor was found to be stable for at least 1 month when stored dry at 4 degrees C.