Periodontal ligament (PDL) is mainly composed of collagen fiber bundles, and the content of collagen fiber is an important factor affecting the mechanical properties of PDL. Based on this, the purpose of this study is to explore the effect of the PDL collagen fiber content on its viscoelastic mechanical behavior. Transverse and longitudinal samples of different regions of PDL were obtained from the human maxilla. The fiber content at different regions of human PDL was quantitatively measured using image processing software, and a new viscoelastic constitutive model was constructed based on the fiber content. The nano-indentation experiment was carried out with a loading rate of 0.5 mN·s−1, a peak load of 3 mN, and a holding time of 200 s, and the model parameters were obtained through the experiment data. The results showed that with the increase of fiber content, the deformation resistance of PDL also increased, and compared with the neck and middle region, the compressive strain in the apical region of PDL was the largest. The range of reduced elastic modulus of human PDL was calculated to be 0.39~5.08 MPa. The results of the experimental data and the viscoelastic constitutive model fit well, indicating that the model can well describe the viscoelastic behavior of human PDL.