Indiana University-Purdue University Indianapolis (IUPUI) / Vitamin D deficiency has been linked to chronic pain and increased opioid use. Realworld data such as electronic medical records and administrative claim data contain large amounts of clinical data and present an opportunity to study the relationship of serum 25- hydroxyvitamin D [25(OH)D] with pain intensity and opioid use. The first study assessed the association between serum 25(OH)D) levels and pain intensity. Compared to patients with normal 25(OH)D levels, those who had insufficient or deficient levels were more likely to experience moderate or severe pain, with multivariable-adjusted odds ratios (95% confidence intervals) of 1.19 (1.05-1.36) and 1.51 (1.28-1.79), respectively. Similar findings were obtained using propensity scores in the matched analyses. In the second study, we investigated the association between serum 25(OH.)D levels and opioid use among opioid-naïve patients. We revealed that those who had insufficient or deficient levels of 25(OH)D were more likely to receive an opioid prescription, with multivariableadjusted odds ratios of 1.10 (1.02-1.17) and 1.18 (1.09-1.28), respectively, compared to patients with normal 25(OH)D levels. Vitamin D deficiency was also associated with a longer duration of opioid use. In the third study, we performed machine learning to identify patient characteristics associated with persistent moderate-to-severe pain (PMSP), explicitly investigating if low serum 25(OH)D levels were a risk factor for heightened pain intensity among obese patients. Low levels of 25(OH)D were consistently identified as a key predictor from a large number of candidate variables in the machine learning models. We detected a significant positive association between serum 25(OH)D levels and PMSP in the logistic regression analysis. Compared to patients with normal levels of 25(OH)D, those who had insufficient or deficient levels of 25(OH)D were more likely to report PMSP, with multivariable-adjusted OR (95% CIs) of 1.15 (1.10-1.21) and 1.28 (1.21-1.35) respectively. We replicated the findings in the first study in a different cohort that showed that low serum 25(OH)D levels might play a role in pain perception. This research contributes to an improved understanding of the role of vitamin D on pain, and opioid use. Individuals who experience pain and need opioid therapy may benefit from optimizing their serum 25(OH)D levels.