Construction of multienzyme-hydrogel sensor with smartphone detector for on-site monitoring of organophosphorus pesticide

•A target-responsive hydrogel-portable kit was proposed to detect pesticide.•Smartphone-based detector combined with test kit for on-site monitoring of paraoxon.•Sensitive and high-throughput detection of OPs with a detection limit of 0.5 ng mL-1 and detection eleven samples simultaneously.•The test kit had been used in real samples with acceptable recovery. Despite the tremendous achievements of existing point-of-care testing (POCT) techniques, accurate on-site detection of pesticide remains a thorny issue. Herein, the concept of multi-enzyme cascade system-based hydrogel kit integrated with a smartphone detector for in-field screening of pesticide was proposed. In this work, we constructed a target-responsive hydrogel (TRhg)-based kit by embedding MnO2 nanoflakes (NFs) into sodium alginate hydrogel. Based on remarkable mimic oxidase activity, MnO2 nanozyme induced color reaction via introducing the sensing probe 3,3′,5,5′-tetramethylbenzidine (TMB). Acetylcholinesterase (AChE) and choline oxidase (ChO) catalyzed the hydrolysis of acetylcholine (ACh) to produce H2O2, which triggered MnO2 NFs decomposition, further blocked the oxidation of TMB. Pesticide (paraoxon) as inhibitor of AChE suppressed the generation of H2O2, which reduced the decomposition of MnO2 NFs, resulting in the color response of kit. Interestingly, the images of kit that captured through smartphone could be analyzed via self-made application program. The benchtop-size biosensor was capable of screening pesticide within 65 min in a sensitive manner (detection limit of 0.5 ng mL-1). More importantly, the obtained results were consistent with those of traditional laboratory microplate reader. Such a miniaturized sensor exhibited high reliability, excellent long-term stability and ease of operation for real-time measurement of pesticide.