Background: Physical exercise may have some effect on cognition in patients with Alzheimer disease (AD). However, the underlying biochemical effects are unclear. Animal studies have shown that amyloid beta (Aβ), one of the pathological hallmarks of AD, can be altered with high levels of physical activity. Aim: The objective of this study was to elucidate the effect of 16 weeks of moderate- to high-intensity physical exercise on the biomarkers of AD, with special emphasis on the amyloidogenic pathway. Methods: From a total of 53 patients with AD participating in the Preserving Cognition, Quality of Life, Physical Health and Functional Ability in Alzheimer's Disease: The Effect of Physical Exercise (ADEX) study we analyzed cerebrospinal fluid samples for Aβ species, total tau (t-tau), phosphorylated tau (p-tau) and soluble amyloid precursor protein (sAPP) species. We also assessed the patients for apolipoprotein E ε4 (ApoE ε4) genotype. Results: We found no effect of 16 weeks of physical exercise on the selected biomarkers, and no effect of ApoE ε4 genotype. Conclusion: Our findings suggest that the possible effect of physical exercise on cognition in patients with AD is not due to modulation of Aβ, t-tau, p-tau and sAPP species.