We have studied in vitro the effects of altered physicochemical properties of thyroglobulin molecules in solution and of the solution itself on iodination kinetics and hormone synthesis. Any change in hydrodynamic properties had a far greater effect in compartmentalized systems, obtained by coating the test tubes with peroxidase, than in conventional homogeneously mixed systems. Increasing thyroglobulin concentration in a range still far below that existing in vivo greatly retarded iodination and hormone synthesis. In contrast, a number of physiological and non-physiological changes of thyroglobulin structure, such as desialylation, preiodination, oxidation and denaturation, strikingly accelerated iodination. The highly variable physical-chemical state of thyroglobulin molecules appears to be a main determinant of protein diffusion within the colloid and, thereby, of iodination kinetics and rate of hormone synthesis. Moreover, alterations of the physicochemical state of thyroglobulin molecules may explain some hitherto ill-understood diffusion phenomena in live follicles.