The potential of individual glycolipid classes from lecithins (soybean, rapeseed, and sunflower) in breadmaking was determined in comparison to classical surfactants such as diacetyltartaric acid esters of mono- and diacylglycerides (DATEM), monoacylglycerides, sodium stearoyl-2-lactylate (SSL), and two synthetic glycolipids by means of rheological and baking tests on a microscale. A highly glycolipid-enriched sample containing the entire glycolipid moiety of the lecithin was obtained using an optimized batch procedure with silica gel. This sample was subsequently used to gain individual glycolipid classes through column chromatography on silica gel. The major glycolipid classes in the lecithins, digalactosyl diacylglycerides (1), sterol glucosides (2), acylated sterol glucosides (3), and cerebrosides (4), were identified and characterized. All isolated glycolipid classes displayed excellent baking performance. A better baking activity than that of the classical surfactants was displayed by 1, 3, and 4 and an equivalent baking activity by 2. The same glycolipid classes, except 3, of different lecithin origin showed only slight differences in their baking activities, due to different fatty acid compositions. Furthermore, the glycolipid classes influenced the crumb structure significantly by improving the crumb softness and grain. Interestingly, none of the glycolipid classes showed significant antistaling effect. A direct effect on the overall rheological behavior of the dough was only found for the commercial surfactants. However, the rheological effect seen on gluten isolated from surfactant-containing dough revealed that the surfactants could be divided into two main groups, one of them directly forming and stabilizing liquid film lamellae through adsorption to interfaces and the other indirectly increasing the surface activity of the endogenous lipids in the flour. The results suggest that in wheat dough, glycolipids seem to have an impact on the dough liquor rather than on the gluten-starch matrix.