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Spaceflight (SF) conditions have a significant impact on the functioning of the human cardiovascular system. The endothelium plays an important role in the process of adaptation to SF factors. Therefore, the detection of biomarkers of endothelial dysfunction is necessary for understanding the molecular mechanisms involved in changes caused by SFs. For this purpose, the blood plasma proteins of 18 Russian cosmonauts were used for quantitative analysis by liquid chromatography with a UPLC 1290 Infinity chromatograph coupled to an Agilent 6490 triple quadrupole mass spectrometer. It was found that a decrease in the circulating plasma volume during the flight followed by the activation of fluid retention at the final stage of flight contributed to the changes in plasma protein concentrations on the first day after landing. We observed a significant increase in the concentration of the S100A9 protein that plays an important role in endothelium functioning and angiogenesis and can serve as a marker of inflammatory reactions. On the first day after landing, the complement system and acute-phase protein concentrations tended to increase, which can adversely affect the functioning of the endothelium.