In this study, we investigated the effect of aging on biochemical markers in serum obtained from Thoroughbred horses (n = 30) and ponies (n = 15). Aging was observed to have an effect on albumin, total bilirubin, thyroxine, and iron levels. The triglyceride concentration in ponies was found to be significantly higher than in Thoroughbreds. Furthermore, an age-related increase in triglycerides was only observed in ponies. There was no aging effect on diacron-reactive oxygen metabolites and biological antioxidant potential concentration. A significant negative correlation between age and bilirubin, known as a strong antioxidative substance, was obtained in both the Thoroughbred and pony groups. However, the distribution area of data was different due to higher bilirubin concentrations in Thoroughbreds. The serum iron levels were correlated with serum triglyceride levels in ponies. We propose that the metabolism of ponies differs from that of Thoroughbreds and that careful breeding and appropriate exercise are required for aging ponies.