Hydrogels are made from natural or synthetic polymers and, currently, they have many biomedical applications. In this work, the conditions for obtaining a hydrogel with similar physicochemical characteristics to the vitreous humor were defined using polyvinyl alcohol (PVA) and glutaraldehyde (GLUT) as cross-linker. The concentration of PVA and GLUT were modified, and their effect was analyzed in terms of the refractive index, density, and dynamic viscosity. The hydrogel which was obtained using 3.98% (w/V) of PVA, 3.13 mL (1.57 g) of GLUT in 100 mL, and the initial pH of 7.2 showed similar characteristics to the vitreous humor (density = 1.0174 ± 0.0050 g mL , dynamic viscosity = 3.7425 ± 0.1800 mPa s and refractive index = 1.3410 ± 0.0010). The hydrogels were further investigated by rheological measurements, infrared spectroscopy, differential scanning calorimetry, X-ray diffraction and determination of swelling degree. The reticulation with GLUT promoted an increase in viscosity and glass transition temperature. On the other hand, it stimulated a decrease in the swelling degree, crystallinity, melting temperature, and intensity of the band related to the -OH bond, compared with the PVA without reticulation. The reticulated hydrogel displayed Newtonian behavior and a higher apparent viscosity than the PVA. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1558-1566, 2018.