The objectives of this study were to evaluate the effect of a single vitamin D3 intramuscular injection on calcium metabolism and bone markers in close-up cows, which fed a diet with negative DCAD (dietary cation anion difference). Twenty-four Holstein cows were randomly assigned to the treatment/control groups 21 days before parturition: twelve cows received a single 8,000,000 IU intra-muscular vitamin D3 injection as the treatment group and the other group received placebo injection (controls). Jugular blood was collected on days − 7, − 5, 1, 2, 4, 7, and 14 relating calving. Following Vit. D3 injection, serum 25(OH)D levels increased. Treated cows had significantly lower parathyroid hormone (PTH) than the control group at days 2 and 7. However, the serum ionized Ca (ICa) concentration was statistically lower in the treatment group. Furthermore, treated cows had lower crosslaps (CrL) than the controls and osteocalcin (OsC) concentrations exhibited a significant decrease at day 7 after calving in the treatment group. This study showed that a single IM injection of Vit. D3 pre-partum in dairy cows resulted in lower PTH, lower CrL, and decreased ICa post-partum. There was no benefit of a single intramuscular injection of vitamin D3 in close-up cows fed negative DCAD diets on Ca metabolism.

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