Tricase Angelo, Catacchio Michele, Marchianò Verdiana, Macchia Eleonora, Bollella Paolo, Torsi Luisa
Department of Pharmacy-Pharmaceutical Sciences, University of Bari Aldo Moro, Via E. Orabona, 4-70125 Bari, Italy.
Centre for Colloid and Surface Science, University of Bari Aldo Moro, Via E. Orabona, 4-70125 Bari, Italy.
Nanoscale. 2025 Apr 10;17(15):9197-9204. doi: 10.1039/d4nr04857a.
Herein, we describe the design and implementation of an ultrasensitive enzyme inhibition-based biosensor for 2,4-dichlorophenoxyacetic acid (2,4-D) detection. The biosensor utilizes alkaline phosphatase (AlP), immobilized on a photo-crosslinked polymer matrix of poly(vinyl alcohol) functionalized with -methyl-4(4'-formylstyryl)pyridinium (PVA-SbQ), supported by electrodes coated with highly porous gold nanocorals (hPGNCs). After preliminary electrochemical and morphological characterization, the PVA-SbQ/AlP/hPGNC electrode was tested for inhibition studies employing ascorbate 2-phosphate (A2P) as the initial substrate. The biosensor preparation/sensing time from electrode preparation to final results is approximately 45 minutes, which enables the possibility to easily scale up the electrode production process on a daily basis with a reliable analytical result in only 5 minutes of amperometric measurement. Following the initial kinetic studies and evaluation of analytical performance, the PVA-SbQ/AlP/hPGNC platform demonstrated a linear detection range from 0.002 to 22 ppt, with a sensitivity of 0.121 ± 0.006 ppt (RSD = 4.9%, = 0.996, and = 6) and a limit of detection (LoD) of 0.7 ppq. This sensitivity is 7-8 orders of magnitude below the regulatory thresholds in Europe and the USA. Furthermore, the biosensor was validated using 19 homogenized wheat leaf sample extracts, prepared in line with European Food Safety Authority (EFSA) guidelines, achieving average recoveries exceeding 96% and RSD values under 9.8%. The biosensor also exhibited robust operational and storage stability, maintaining 84% (30 hours of continuous operation) and 94% (120 days) of its initial response, respectively. These results highlight the potential of the PVA-SbQ/AlP/hPGNC biosensor for on-site 2,4-D monitoring in agricultural crops and its feasibility for integration with artificial intelligence for advanced diagnostics.
在此，我们描述了一种基于超灵敏酶抑制的生物传感器的设计与实现，用于检测2,4-二氯苯氧乙酸（2,4-D）。该生物传感器利用固定在由甲基-4(4'-甲酰基苯乙烯基)吡啶鎓（PVA-SbQ）功能化的聚乙烯醇光交联聚合物基质上的碱性磷酸酶（AlP），并由涂覆有高度多孔金纳米珊瑚（hPGNCs）的电极支撑。经过初步的电化学和形态学表征后，使用抗坏血酸2-磷酸酯（A2P）作为初始底物，对PVA-SbQ/AlP/hPGNC电极进行抑制研究测试。从电极制备到最终结果的生物传感器制备/传感时间约为45分钟，这使得能够轻松地按日常规模扩大电极生产过程，只需5分钟的安培测量即可获得可靠的分析结果。在进行初步动力学研究和分析性能评估后，PVA-SbQ/AlP/hPGNC平台的线性检测范围为0.002至22 ppt，灵敏度为0.121±0.006 ppt（相对标准偏差=4.9%，相关系数=0.996，n=6），检测限（LoD）为0.7 ppq。该灵敏度比欧洲和美国的监管阈值低7-8个数量级。此外，使用按照欧洲食品安全局（EFSA）指南制备的19种均质小麦叶样品提取物对该生物传感器进行了验证，平均回收率超过96%，相对标准偏差值低于9.8%。该生物传感器还表现出强大的操作和储存稳定性，分别保持其初始响应的84%（连续运行30小时）和94%（120天）。这些结果突出了PVA-SbQ/AlP/hPGNC生物传感器在农作物现场2,4-D监测方面的潜力及其与人工智能集成用于先进诊断的可行性。
