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

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.

A noninvasive and qualitative bioluminescent assay for express diagnostics of athletes' responses to physical exertion.

Upcoming professional sports authorities seek rapid noninvasive biosensing tools for regular monitoring of athletes' physiological states. The analysis of saliva through luminescence-based biosensors has been perceived as a suitable candidate for such purposes. The present study reports a qualitative bioluminescence assay based on a coupled enzyme system that consists of bacterial luciferase (BLuc) and nicotinamide adenine dinucleotide (NADH):flavin mononucleotide (FMN) oxidoreductase (Red), BLuc-Red, for the express diagnostics of athletes' stress levels before and after physical exertion. The volunteers who participated in the study were grouped as freestyle wrestlers and students who adapted to different levels of physical activities. Under physical exertion modelling conditions, the influence of participant saliva on BLuc-Red catalyzed light emission was investigated. Results showed a significant increase in residual luminescence (Iexp , mean maximum bioluminescence intensity of the experimental measurement (Iexp ); Ic , luminescence intensity in control; Iexp /Ic , %) values for participants in the wrestler group while a decrease in the student group (P < 0.05). Such contrasting residual luminescence values in both groups were found to be dependent on the catalase activity of saliva. The proposed bioluminescence assay can be utilized as a potential nonspecific biosensing tool for determining the physical state of athletes under high loads.