Living organisms depend on precisely calibrated biological mechanisms that are governed by enzymes. Scientists have always been captivated by the complexity of these systems and have endeavoured to replicate them by constructing cascade reactions and chemical reaction networks catalysed by enzymes. Enzymes play a crucial role in numerous applications as they possess the ability to function as sensors. Biosensors utilize the reaction's end product to generate a signal applicable in various fields, including biomedicine, the food and pharmaceutical industry, environmental pollution, and forensic science. Enzyme biosensors also have the potential for signal amplification, enhancing their sensitivity towards analytes present in low concentrations by augmenting the concentration of output molecules in response to input signals.
Our focus in this study is the utilization of enzymes in particle biosensors and reactions that display amplification, such as the urease reaction which is autocatalytic in pH. We also examine the role of transport phenomena, including diffusion, in sensing processes for enzyme-loaded micro and nanoparticles. The urease reaction has already been utilized in sensing applications, but its feedback and ultrasensitivity capabilities have not yet been fully exploited. When dealing with an autocatalytic reaction, positive feedback can be utilized to fine-tune sensor properties. In this particular study, we focused on optimizing parameters in enzyme particle biosensors to reduce response time, using experiments and computer simulations of the urea-urease reaction. We discovered that entrapment of the enzyme in thiol-acrylate polymer particles resulted in an optimum particle size for minimum response time, taking into account the diffusion of both substrate and product. These findings will aid in the design of enzyme-particle biosensors with autocatalysis.
Immobilized enzymes offer reusability, which makes them ideal for applications such as biosensors, industrial monitoring and transformations, and water quality examination. However, creating a suitable combination of enzyme and supportive material can be challenging as it may lack stability, especially due to enzyme denaturation over time or storage conditions. To address this issue, researchers have suggested a sustainable, process-free, and robust enzyme source such as the enzymes present in watermelon seeds (WMS) as an alternative to standard commercial enzymes. These enzymes are shielded from the external environment by a lipid layer known as 'protein microbodies', making them reusable and reproducible. We designed a system with WMS in various hydrogel particles that detected urea down to 0.4 mM in just one hour and were stored at room temperature for over a month.
Our research investigated the utilization of enzyme combinations in constructing chemical reaction networks to facilitate more complex sensing and chemo-mechanical processes. Enzyme networks that can modify pH can serve various purposes, such as pulsatile drug delivery. Nonetheless, acquiring enzymes from different sources can prove costly and result in incompatible or unstable enzymes. Our objective was to combine diverse enzyme types like urease, catalase, glucose oxidase, and beta-glucosidase to program pH shifts. We found that watermelon seeds yielded numerous enzymes that were not only easy to use, without requiring extensive processing, but also affordable. By combining these enzymes and building reaction networks, we were able to enhance the reaction rate of certain processes and accomplish significant amplitude pH changes that would have been otherwise difficult to achieve.
The integration of chemical stimuli with a mechanical response is an innovative concept that scientists find particularly intriguing. Smart hydrogels, which can transform by shrinking and swelling in response to changes in external conditions, are becoming increasingly popular in applications such as drug delivery, active sensors, and artificial tissues. This work has produced a pH-responsive gel equipped with multiple enzymes and a colourimetric indicator capable of producing a chemo-mechanical response when exposed to analytes, such as urea and glucose, that generate a basic or acidic environment. We investigated the reusability of particles with different enzyme sources and also obtained an enhanced mechanical response as a result of combining the effects of enzymes in watermelon seed, including catalase. This type of device has the potential to be used as a sensor that can regulate the release of a substance in time, such as drugs or chemicals for neutralising toxins.
Overall, these findings will aid in the enhancement of enzyme-particle biosensors, benefitting a wide range of industries from biomedicine to environmental pollution monitoring.
