Precise control of cross-linking parameters such as reaction time and temperature is crucial in hydrogel synthesis. This control directly governs how the polymer network forms, providing a cost-effective way to optimize the entire process. In this paper, we examine the influence of heat treatment time on chitosan (CS)/oxidized pullulan (APUL) hydrogel to obtain biomaterials with tunable cross-linked density, viscoelastic, and physical properties for specific applications. In order to determine the structure of the hydrogels, analyses based on scanning electron microscopy (SEM) and Fourier-transform infrared spectroscopy (FTIR) were performed. Furthermore, the results of heat treatment time were assessed by gel fraction, swelling ratio, thermal analysis, and rheological testing. The results show that the gel fraction and elastic strength of the CS/APUL hydrogels increased from 61.7% ± 4.4% to 70.0% ± 2.8% and 795.7 ± 111.9 to 1836.0 ± 82.3 Pa respectively, as the heat treatment time increased due to increase in cross-linking density within the hydrogel matrix. In addition, the swelling ratio of the hydrogels decreased as the incubation time increased from 0.5 to 4 h, due to the higher cross-linking density of the network structure, which restricted water uptake. The variation of heat treatment time is a crucial parameter to obtain chitosan/oxidized pullulan hydrogels with interesting and tunable properties. © 2026 Wiley Periodicals LLC.