This study investigated human diets during the nineteenth and twentieth century in Copenhagen through stable isotope analysis of carbon and nitrogen according to sex, age, socio-economic status, and period (of death). Stable isotope analysis was conducted on bone collagen (n = 114) and hair keratin (n = 21) recovered from individuals buried at the Assistens Cemetery. Animal samples (n = 40) from eighteenth and nineteenth century deposits in Copenhagen were also analyzed. Significant differences in collagen δ13C signals were found between males and females, while the differences in collagen δ15N values were not significant. When analyzed temporally, the male–female difference in δ13C values is significant during the twentieth century, but not during the nineteenth century. Significant differences were found in collagen δ15N values between males of different socio-economic status, while female diets showed no wealth dependence. Diet was not correlated with age; however, bone–hair analysis indicated change in nutritional intake or change in health status months prior to death. For some individuals, this may have been associated with disease and/or ill health. The impact of manuring in elevating baseline δ15N values was not detected. Overall isotopic results indicate a diet rich in protein from brackish fish and terrestrial C3-based animal products with a larger dietary diversity among males during the twentieth century. Male diet may have been more affected by economical means than female diet.