Understanding the global climate change from greenhouse to icehouse conditions at the Eocene–Oligocene Transition (EOT) 34 million years ago requires climatic records from oceanic as well as continental realms of the key Late Eocene “doubthouse” period preceding this switch. Here, we report integrated stratigraphic results from well-dated Late Eocene continental mudflat to saline lake paleoenvironments of the Xining Basin (northeastern Tibetan Plateau, western China) recording regional and global change. Cyclostratigraphic analysis strongly suggests continuous dominance of the 41-kyr obliquity cycle in the whole late Eocene interval down to the base of polarity chron C18n.2n at 39 Ma with additional input of the ~ 100-kyr eccentricity cycle up to the base of chron C13r at ~ 34.7 Ma. This might imply that high-latitude climates dominated the area long before the EOT, probably related to incipient ice-volume fluctuations. Furthermore, our results reveal two paleoenvironmental deterioration steps preceding the Eocene–Oligocene Transition. The first step occurs in the top of chron C17n.1n at ~ 36.6 Ma. This age closely corresponds to (1) the high-altitude pollen appearance in chron C16.2r at ~ 36.4 Ma in the same section, (2) the recently dated final retreat of the Tarim Sea in western China, and (3) a shift from precession to obliquity dominance in the Atlantic Ocean. This near co-occurrence suggests global change at this time. We hypothesize this change is related to an increase in incipient ice sheet volume leading to passing threshold conditions for the high-altitude pollen appearance and Tarim Sea retreat, finally leading to decreased moisture availability in the Xining Basin. At the second step, in the base of chron C13r at ~ 34.7 Ma, a substantial increase in clastic sedimentation rates is observed. This might relate to increased climate variability preceding the greenhouse to icehouse transition at the EOT that prevented landscapes to attain equilibrium configurations.