Biotechnological approaches for non-enzymatic oxidative stress biomarkers in aquatic monitoring

Non-enzymatic oxidative stress biomarkers such as thiobarbituric acid reactive substances (TBARS), protein carbonyls (PC), and reduced glutathione (GSH) are among the most widely applied endpoints in aquatic biomonitoring and ecotoxicology, with relevance for assessing pollution across coastal, estuarine, and marine ecosystems. Despite decades of use, however, their translation into robust environmental monitoring tools remains constrained by analytical and interpretative bottlenecks, including poor interlaboratory comparability, lack of harmonized sampling and storage procedures, matrix interferences, inconsistent normalization strategies, and insufficient quality assurance frameworks. This review critically examines the use of non-enzymatic oxidative stress biomarkers in aquatic systems, shifting the focus from biomarker relevance toward methodological and technological barriers that currently limit operational deployment. We synthesize evidence on methodological variability, analytical artefacts, and confounding factors affecting biomarker responses across biological matrices and environmental contexts. Our analysis highlights that TBARS, PC, and GSH are highly sensitive indicators of oxidative damage associated with exposure to diverse contaminant classes, including metals, organic pollutants, and complex contaminant mixtures. However, substantial heterogeneity in analytical approaches and experimental designs across studies continues to hinder interstudy comparability and the establishment of ecologically meaningful baseline values. Finally, we propose a technology-oriented roadmap for improving biomarker standardization and field applicability, including minimum reporting requirements, validation priorities, and scalable analytical strategies such as miniaturized assays, microfluidic platforms, biosensing technologies, and smartphone-based detection systems. Overall, the integration of these approaches may facilitate the transition of non-enzymatic oxidative biomarkers from research endpoints to standardized and operational tools for environmental monitoring and decision-making in aquatic ecosystems.