Purpose: To examine trends of population-level semen quality over a 20-year period. Materials and Methods: We performed a retrospective review of data from the andrology lab of a high volume tertiary hospital. All men with semen samples between 2000 and 2019 were included and men with azoospermia were excluded. Semen parameters were reported using the World Health Organization (WHO) 4th edition. The primary outcome of interest was changes in semen parameters over time. Generalized least squares (GLS) with restricted cubic splines were used to estimate average-monthly measurements, adjusting for age and abstinence period. Contrasts of the estimated averages based on GLS between the first and last months of collection were calculated. Results: A total of 8,990 semen samples from subfertile non-azoospermic men were included in our study. Semen volume decreased over time and estimate average at the beginning and end were statistically different (p<0.001). Similarly sperm morphology decreased over time, with a statistically significant difference between estimated averages from start to finish (p<0.001). Semen pH appeared to be increasing over time, but this difference was not significant over time (p=0.060). Sperm concentration and count displayed an increase around 2003 to 2005, but otherwise remained fairly constant over time (p=0.100 and p=0.054, respectively). Sperm motility appeared to decrease over time (p<0.001). Conclusions In a large sample of patients presenting to a single institution for fertility assessment, some aspects of semen quality declined across more than two decades. An understanding of the etiologies and driving forces of changing semen parameters over time is warranted.