Thiol (SH) containing additives improve the mechanical properties of rigid, glassy gluten materials. However, the underlying molecular mechanism is still unclear. In particular, the importance of the preceding gluten-additive mixing conditions remains to be investigated. Here, different additives containing either only SH, only disulfide or both SH and disulfide functionalities were synthesized and their impact on the gluten network using different mixing conditions prior to subsequent molding were assessed. All SH containing additives decreased the gluten molecular weight (MW) during mixing to a degree depending on the conditions. Additives with only disulfide functionality did not significantly affect protein size during mixing irrespective of the conditions used. Only when mixing induced sufficient MW reduction did the strength and failure strain of rigid gluten materials increase. This shows that factors other than the degree of cross-linking affect the strength of rigid gluten materials. These results support our hypothesis that altered molecular conformations and improved molecular entanglements also contribute to material strength. The extent to which such conformational changes occur may depend on the additive and the way of mixing.