The acute-phase response (APR) is the most frequent side effect after the first dose of intravenous nitrogen-containing bisphosphonates (N-BPs). It has been demonstrated in vitro that N-BPs stimulate γδ T-cell proliferation and production of cytokines and that vitamin D is able to modulate them. Therefore, we have studied the relationship between bone metabolism parameters, particularly for 25-hydroxyvitamin D [25(OH)D], and APR in patients treated with 5 mg zoledronic acid intravenously. Ninety N-BP-naive osteoporotic women (63.7 ± 10.6 years of age) were stratified for the occurrence of APR (APR+) or not (APR–) and quantified by body temperature and C-reactive protein (CRP). The APR+ women had significantly lower 25(OH)D levels than the APR– women. Levels of 25(OH)D were normal (>30 ng/mL) in 31% of APR+ women and in 76% of APR– women. The odds ratio (OR) to have APR in 25(OH)D-depleted women was 5.8 [95% confidence interval (CI) 5.30–6.29; p < .0002] unadjusted and 2.38 (95% CI 1.85–2.81; p < .028) after multiple adjustments (for age, body mass index, CRP, calcium, parathyroid hormone, and C-telopeptide of type I collagen). Levels of 25(OH)D were negatively correlated with postdose body temperature (r = −0.64, p < .0001) and CRP (r = −0.79, p < .001). An exponential increase in fever and CRP has been found with 25(OH)D levels lower than 30 ng/mL and body temperature less than 37 °C, whereas normal CRP was associated with 25(OH)D levels above 40 ng/mL. The association between post-N-BPs APR and 25(OH)D suggests an interesting interplay among N-BPs, 25(OH)D, and the immune system, but a causal role of 25(OH)D in APR has to be proven by a randomized, controlled trial. However, if confirmed, it should have some practical implications in preventing APR. © 2010 American Society for Bone and Mineral Research.