In this contribution, we have compared the stabilizing effect of sucrose on overall as well as on domain-II of human serum albumin (HSA) against unfolding by different denaturating agents. HSA was denatured thermally by raising the temperature and also chemically by guanidine hydrochloride (GnHCl) and urea. Circular dichroism spectroscopy was used to monitor the change in the overall structure of HSA, whereas tryptophan fluorescence has been used to investigate the local structural alteration within the domain-II of HSA. The degree of folding using different combinations of GnHCl–sucrose, urea–sucrose, and temperature-sucrose was investigated to calculate the relative stability of the native, intermediate, and denatured states of HSA with increasing sucrose concentration. In the presence of sucrose, the intermediate state, formed during GnHCl-induced denaturation, populated at higher GnHCl concentration (1.5 M for overall denaturation and 2.5 M for domain-II) compared to that in the absence of sucrose (1 M for overall denaturation and 2 M for domain-II). A similar effect has also been observed for urea. These signify that sucrose stabilizes the native state of the protein. Extent of thermal unfolding is also found to be minimized in the presence of sucrose. In a nut shell, sucrose stabilizes different parts of HSA differently and also the net stabilizing effect toward different denaturation profiles is different.