A MOF-stabilized enzyme-based colorimetric biosensor integrated with pH test strips for portable detection of D-limonene in agricultural applications

Volatile organic compounds (VOCs) emitted by plants provide essential insights into plant health, quality, and environmental stressors. Among these, D -limonene emissions fluctuate with agricultural stress and fruit quality and have emerged as a potential biomarker for monitoring plant condition. However, developing a portable, low-power, and user-friendly sensor for D -limonene detection in field conditions remains challenging. In this study, we report a novel colorimetric biosensor that employs a mild one-pot synthesis method to immobilize acetylcholinesterase (AChE) on a zinc-based metal–organic framework (AChE@Zn-MOF) and utilizes phenolphthalein as a pH indicator for highly sensitive D -limonene detection. The biosensor retains enzymatic activity comparable to free AChE while exhibiting enhanced stability across a range of environmental conditions. D -limonene inhibits AChE activity, disrupting acetylcholine hydrolysis and inducing a detectable pH shift, resulting in a visual color change. To enhance field practicality, the biosensor was integrated into commercial pH test strips coated with an agarose gel membrane to improve moisture retention and gas absorption. Coupled with smartphone-assisted image processing, the system enables detection of D -limonene at concentrations as low as 1 ppm and allows visualization of its release during mango post-harvest storage and citrus pest infestations. This approach establishes a foundation for portable, colorimetric smart sensing technologies in agriculture, facilitating real-time monitoring of VOCs like D -limonene in the field.