There is increasing interest in the hydroponic technology to produce leafy vegetables for ready-to-eat salads. Optimisation of the growing system can lead to higher yield and/or improved nutritional value of the product. Selenium (Se) is an essential element for animal and humans, with quite a narrow range between deficiency and toxicity, whereas it is assumed beneficial for plants. In the present study, two cultivars (Gala and Baron) of corn salad (Valerianella locusta (L.) Laterr.) were used to test the possibility to increase Se content in the edible parts (leaves). Effects on yield, nitrate content, and accumulation of sulfur (S) and S-containing amino acids and Se and Se-containing amino acids were studied. Results showed that corn salad tolerates selenate (Na₂SeO₄) concentrations ranging from 10 to 40 µm in the nutrient solution, with plants accumulating Se at levels compatible with the need in human diets at 10 µm selenate. Se-treated plants showed some benefits with respect to a decrease of nitrate concentration and increase of pigment contents (chlorophylls and carotenoids). At 10 µm selenate, Se-cysteine and Se-methionine were produced, without affecting non-protein thiols or cysteine and methionine contents. At the higher Se supply, sulfate accumulated in the leaves with a parallel decrease in the amount of S-amino acids and a rise in the relative amount of Se-amino acids. Based on the chemical analyses, cv. Gala showed better tolerance than cv. Baron to moderate selenate supply (40 µm).