Visual colorimetric 'turn-off' biosensor for ascorbic acid detection based on hypochlorite-3,3',5,5',-Tetramethylbenzidine system.

An 'turn-off' approach for the detection of ascorbic acid (AA) using 3,3',5,5',-tetramethylbenzidine (TMB) as a colorimetric probe is developed. The proposed method is based on the fact that hypochlorite (ClO-) could oxidize TMB to oxidized TMB (oxTMB) along with a blue color and an absorption peak centered at 652 nm. However, the introduction of AA could cause the reduction of oxTMB, which results in a blue color fading and a decrease of the absorbance at 652 nm. Based on this finding, we proposed a sensitive colorimetric assay for quantitative determination of AA by UV-vis spectroscopy. Under the optimal conditions, a good linear response between the absorbance and AA concentration was achieved within the dynamic working range from 1 to 70 μM with a detection limit of 0.58 μM. More importantly, the proposed method offered advantages of simple, low-cost instruments and rapid assay without the utilization of nanomaterials and has also been successfully applied for the determination of AA in orange juice with satisfying recoveries over 97.0%. Furthermore, the obtained results were consistent with those measured by high performance liquid chromatography with the average relative error of 3.3%.