Objective: The risk of primary ovarian insufficiency (POI) increases in association with autoimmune conditions. Adequate intake of vitamin D (vit D) and trace elements is required for the immune system to function efficiently. The aim of this study was to evaluate vit D, zinc, and copper blood levels in women with POI who had given birth to at least one child and in women with normal menstrual cycles. Methods: This was a cross-sectional, case-control study involving 63 participants divided into two groups: the study group, which is composed of 35 women with POI, and the control group, which is composed of 28 women with normal menstrual cycles. Serum concentrations of zinc, vit D, and copper were determined for each participant. Results: Women with POI had significantly higher serum copper levels and copper-to-zinc ratio but significantly lower serum vit D and zinc levels when compared with the healthy control group. Serum follicle-stimulating hormone levels were inversely correlated with zinc and vit D levels and positively correlated with the copper-to-zinc ratio and copper levels. Vit D levels were inversely correlated with follicle-stimulating hormone levels, copper-to-zinc ratio, and copper levels and positively correlated with zinc levels. Conclusions: Most women with POI are deficient in vit D. Zinc, copper, and vit D seem to correlate with hormonal status in the participants. The present study may generate hypotheses for future studies that will investigate the possible mechanisms behind alterations in trace elements and vit D deficiency in women with POI and whether these changes could be used to screen for the risk of developing POI. © 2013 by The North American Menopause Society.