The aim of this study was to identify the metabolic changes that have occurred in Holstein-Friesian cows in Japan over the past 2 decades, based on long-term metabolic profiles. From 1987 to 2004, metabolic profile tests were conducted in 1,700 dairy herds containing ~50,000 cows. The cows were divided into 5 lactation stage groups: early (calving to 49 d in milk, DIM), peak (50–109 DIM), mid (110–209 DIM), and late (210 to dry-off) lactation and the dry period. Principal component analysis was applied to the metabolic profiles at each lactation stage separately to investigate changes in metabolic profiles across the multiyear periods 1987–1992, 1993–1998, and 1999–2004. We determined that cows have probably experienced increasing negative energy balance, energy deficiency, and liver dysfunction during the past 2 decades in Japan.