Walnut (Juglans regia L.) is one of the most common nuts in the world. There are a wide walnut variety of genotypes that differ in forestry, physical and chemical properties. Most of the walnut genotypes, which are natural resources have been evaluated as promising and germplasm sources for breeding. Walnuts are high in many beneficial biochemical compounds, so they have a useful profile for many people in relation to disease risk. Therefore, the demand for production and consumption of walnut is increasing day by day. In the present study, fatty acid and tocopherol profiles of 156 F1 genotypes belonging to Chandler × Kaplan-86 walnut population and their parents were investigated. Fatty acids and tocopherol isomers (α, β, γ) were determined by GC/FID (gas chromatography/ flame lonizing detector) and HPLC (high-pressure liquid chromatography) techniques, respectively. The fatty acid content of F1 walnut genotypes and their parents were found to be 0.00%–3.34% for myristic acid, 5.86%–10.05% for palmitic acid, 0.01%–4.46% for stearic acid, 0.00%–0.89% for palmitoleic acid, 10.00%–20.89% for oleic acid, 45.36%–66.20% for linoleic acid and 9.04%–20.55% for α-linolenic acid. The content of α-tocopherol, β-tocopherol and γ-tocopherol from the walnut population were determined to be 1.61–23.65 µg/g, 1.90–12.57 µg/g, 90.22–394.96 µg/g, respectively. The Chandler × Kaplan-86 F1 hybrid walnut population showed a significant variation in terms of the fatty acids and tocopherol contents. This research contains up-to-date scientific data on very wide walnut genetic resources and composition. Our data show that 156 F1 walnut genotypes have a potential source of monounsaturated fatty acids and tocopherols and the results are very important for future breeding studies on walnut.