Fluorescence quenching techniques complemented with optical rotation and viscosity measurements were employed to study the effects of urea and sodium dodecylsulfate on the formation of hydrophobic microdomains and its relationship to the extent of helical structuration of gelatin chains in aqueous solution. The effects were examined for both additives acting independently of each other as well as for their joint action. Addition of urea (0.2-2 M) to a 0.5% (w/v) gelatin solution decreases the fraction of the fluorescence of pyrene protected against quenching by acrylamide in a fashion directly related to the extent of triple helix structuration of the polypeptide chains. The decrease in the amount of structured microdomains elicited by urea addition is accompanied by a substantial decrease in the viscosity of the gelatin solution. Upon addition of 1 mM SDS to the gelatin solution at 0.5% (w/v), the fluorescence of pyrene is almost completely protected against quenching by acrylamide, the triple helix content slightly decreases, and the viscosity of the solution increases. The joint incorporation of both additives produces an increase in the viscosity of the solution when SDS is added to a gelatin solution containing urea but, at any SDS concentration, the presence of urea decreases the viscosity of the solution. Similarly, addition of SDS to a highly disorganized gelatin solution in the presence of urea increases the fraction of triple helix, but urea added to the gelatin solution in the presence of SDS decreases it. The effects observed are interpreted in terms of the formation of different types of microdomains when urea or SDS are added independently as well as when they are simultaneously present in the solution.