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Saliva is one of the most significant biological liquids for the development of a simple, rapid, and non-invasive biosensor for training load diagnostics. There is an opinion that enzymatic bioassays are more relevant in terms of biology. The present paper is aimed at investigating the effects of saliva samples, upon altering the lactate content, on the activity of a multi-enzyme, namely lactate dehydrogenase + NAD(P)H:FMN-oxidoreductase + luciferase (LDH + Red + Luc). Optimal enzymes and their substrate composition of the proposed multi-enzyme system were chosen. During the tests of the lactate dependence, the enzymatic bioassay showed good linearity to lactate in the range from 0.05 mM to 0.25 mM. The activity of the LDH + Red + Luc enzyme system was tested in the presence of 20 saliva samples taken from students whose lactate levels were compared by the Barker and Summerson colorimetric method. The results showed a good correlation. The proposed LDH + Red + Luc enzyme system could be a useful, competitive, and non-invasive tool for correct and rapid monitoring of lactate in saliva. This enzyme-based bioassay is easy to use, rapid, and has the potential to deliver point-of-care diagnostics in a cost-effective manner.

Bioluminescent-Triple-Enzyme-Based Biosensor with Lactate Dehydrogenase for Non-Invasive Training Load Monitoring