1. Introduction
The development of sensitive, selective, and robust biosensors for detecting biomolecules in complex sample matrices has become a significant focus in analytical chemistry and biotechnology. L-arginine (L-arg), an essential amino acid involved in various physiological processes, demands precise and reliable detection, especially in real samples like food or biological fluids. Conductometric biosensors, which rely on changes in conductivity to indicate analyte concentration, present a promising approach. In the context of L-arg determination, an innovative biosensor combining enzymatic activity with ion-sensitive materials offers a new pathway for enhanced accuracy and stability.
2. Biosensor Design and Enzyme Immobilization Strategy
The biosensor was fabricated by co-immobilizing two key enzymes—arginase and urease—alongside an ion-selective material, zeolite clinoptilolite (Clt), on gold interdigitated electrodes. The arrangement of these components on the sensor surface was critical to achieving optimal performance. Different configurations were tested to assess their influence on sensitivity, stability, and response characteristics. The most effective design involved the primary deposition of Clt, followed by the secondary co-immobilization of the enzymes. This structure provided a favorable microenvironment for enzymatic reactions and ion exchange, enhancing the biosensor’s responsiveness.
3. Analytical Performance of the Developed Biosensor
Among all tested configurations, the biosensor with Clt as the base layer and enzyme mixture on top exhibited superior analytical performance. It achieved a high sensitivity of 9.61 ± 0.01 μS/mM and a low detection limit of 5 μM, making it suitable for detecting trace levels of L-arg. The linear detection range of 0–280 μM and a broad dynamic range of up to 15 mM further demonstrate its applicability across various sample concentrations. These attributes underscore its potential for accurate L-arg quantification in complex samples.
4. Stability and Reproducibility of the Biosensor
Long-term usability is vital for practical applications of biosensors. The developed L-arg biosensor showed excellent operational and storage stability, retaining consistent performance over extended periods. Stability tests confirmed minimal signal degradation and high reproducibility of results across multiple uses. This robustness makes the biosensor suitable for routine analysis in laboratories or on-site food testing environments, reducing the need for frequent recalibration or replacement.
5. Influence of Solution Parameters on Sensor Response
The effect of environmental and solution parameters—such as pH, ionic strength, and buffer capacity—on biosensor sensitivity was systematically investigated. These factors significantly influence enzyme activity and ion exchange dynamics at the biosensor surface. The sensor's design effectively minimized fluctuations in performance under varied conditions, indicating its adaptability to real-world samples with differing chemical compositions. This adaptability is a crucial feature for sensors used in diverse applications, including food quality control and clinical diagnostics.
6. Application in Real Sample Analysis and Method Validation
To validate its real-world applicability, the biosensor was employed to quantify L-arg in food samples with complex matrices. The results were benchmarked against a reference method—ion chromatography. The close agreement between the biosensor and ion chromatography data (correlation coefficient R = 0.96) confirmed the accuracy and reliability of the biosensor. This high correlation proves its potential as a fast, user-friendly alternative to conventional laboratory techniques for L-arg analysis in food and possibly biological fluids.
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