The biosensor for monitoring asparagine levels in Fruit juices was developed in present research work. Lasparaginase (biocomponent) has been extracted from Withania somnifera (L) Dunal, and immobilized into TEOS hydrosol gel-chitosan matrix. Kinetic characterization of both native and immobilized enzyme was compared. Response time studies had been carried out by Phenol Red indicator coimmobilized with asparaginase and colour visualization strategy has been optimized for a range of asparagine concentrations. The detection limit of asparagine achieved was 10-10–10-1 M. Furthermore, the developed biosensor was applied on different fruit juices for detection of asparagine levels. Biocomponent was found to be stable for 32 days in Na borate buffer (pH 8.5) at 4°C and it is reliable for fruit juices.