The saliva of athletes participating in anaerobic (track and field athletes) or aerobic (swimmers) physical activity was studied. Saliva sample was collected before (control) and after the workout. It was found that physical activity significantly increased the salivary total protein concentration and pH values in the athletes. The activity of catalase in saliva was significantly increased only after aerobic, but not anaerobic exercise performance. The bioluminescent indicator of athletes’ saliva is significantly higher during aerobic workout than anaerobic exercise. Salivary biomarkers such as lactate concentration, catalase activity, total protein concentration, calcium, potassium, magnesium ions, and the amide IV group may have a significant effect on the enhancement of light emission in a bioluminescent assay based on a system of coupled enzymatic reaction. The amide IV group and the presence of the salivary oligosaccharides, polysaccharides, phosphatase, and phospholipids in athletes during anaerobic physical activity can eliminate bioluminescence more efficiently than the concentration of triene conjugates and the amide II group in the saliva of athletes involved in aerobic exercises. Thus, significant biomarkers for the measurement of the physiological status of athletes who engaged in aerobic or anaerobic physical activity have been identified in saliva, these biomarkers can influence the integrated response of the bioluminescent enzymatic biotest. The change in the integrated bioluminescent indicator of athletes' saliva depending on the type of physical exertion can be used in sports medicine to prevent physical overload.