α-Tocopherol (Toc) is the tocopherol isoform with the highest vitamin E activity. Since soybean seeds contain a low percentage of α-Toc, increasing the α-Toc content to improve vitamin E activity is a breeding objective. Little is known about the genetic variability of the tocopherol content and composition in soybeans. In the present study, we analyzed the tocopherol composition of 1,109 germplasm accessions of cultivated and wild soybean by high performance liquid chromatography (HPLC) and identified three varieties, Dobrogeance, Dobruoza 14 Pancevo and Keszthelyi Aproszemu Sarga with a high α-Toc content. Dobrogeance showed a similar Toc composition to that of Dobruoza 14 Pancevo with the α-Toc content being four to seven times higher than that of standard varieties and almost the same as that of total Toc. Keszthelyi Aproszemu Sarga showed a markedly increased content of α-Toc, but a significantly lower total Toc content.