There are two main types of mineral waters used in the Staraya Russa spa: (1) the saline waters of the Middle and Upper Devonian Arukjula–Shventoy aquifer with isotopic composition of δ¹⁸O from − 10.7 to − 11.3‰ and δ²H from − 81.3 to − 85‰ and (2) the brackish waters of the Upper Devonian Sargayev–Daugava aquifer with isotopic composition of δ¹⁸O from − 12.5 to − 12.9‰ and δ²H from − 93.0 to − 94.1‰. Hydrogeological conditions within the spa and its vicinity are complex due to hydraulic contacts between artesian aquifers and the saline waters ascension into shallower fresh water aquifers. The studies revealed that saline waters of the Arukjula–Shventoy aquifer are mixtures of approximately equal fractions (0.5) of snow melt water of δ¹⁸O around − 21.7‰ and seawater-like end-member of isotopic composition similar to modern ocean water (i.e. VSMOW). The brackish waters of the Sargayev–Daugava aquifer are result of mixing between infiltration waters and saline waters ascending from deeper Arukjula–Shventoy aquifer, the fraction of saline water may reach from around 0.2 to 0.4. The sulfur (δ³⁴S) and oxygen (δ¹⁸O) isotopic composition of sulfates dissolved in mineral waters indicate that their source is connected with marine sulfates. The ionic ratios of major chemical compounds corroborate the presence of the connate seawater-like component in the hydrogeological system, which is responsible for the observed chemical composition of mineral waters extracted in Staraya Russa spa.