Settlement by modern humans in Ukraine and its vicinity dates back to 32,000 BCE, with evidence of the Gravettian culture in the Crimean Mountains. By 4,500 BCE, the Neolithic Cucuteni–Trypillia culture was flourishing in wide areas of modern Ukraine, including Trypillia and the entire Dnieper-Dniester region. Ukraine is also considered to be the likely location of the first domestication of the horse. During the Iron Age, the land was inhabited by Cimmerians, Scythians, and Sarmatians. Between 700 BCE and 200 BCE it was part of the Scythian kingdom.

From the 6th century BCE, Greek, Roman, and Byzantine colonies were established on the north-eastern shore of the Black Sea, such as at Tyras, Olbia, and Chersonesus. These thrived into the 6th century CE. The Goths stayed in the area, but came under the sway of the Huns from the 370s. In the 7th century, the territory that is now eastern Ukraine was the centre of Old Great Bulgaria. At the end of the century, the majority of Bulgar tribes migrated in different directions, and the Khazars took over much of the land.

In the 5th and 6th centuries, the Early Slavic, Antes people lived in Ukraine. The Antes were the ancestors of Ukrainians: White Croats, Severians, Eastern Polans, Drevlyans, Dulebes, Ulichians, and Tiverians. Migrations from the territories of present-day Ukraine throughout the Balkans established many South Slavic nations. Northern migrations, reaching almost to Lake Ilmen, led to the emergence of the Ilmen Slavs, Krivichs, and Radimichs, the groups ancestral to the Russians. Following an Avar raid in 602 and the collapse of the Antes Union, most of these peoples survived as separate tribes until the beginning of the second millennium.