Although it has been well established that periconceptional use of multivitamins containing folic acid (FA) reduces the risk for neural tube defects, two recent U.S. studies have shown an increased risk for multiple congenital anomalies (MCAs) associated with periconceptional use of vitamins containing FA. This study assessed the association between the periconceptional use of vitamins containing FA and MCAs in a third U.S. population. Mothers of infants with MCAs and a random sample of live births (control infants) born in Iowa during 1993–1995 were eligible to participate in the Birth Defects Risk Factor Surveillance case-control study. During a telephone interview, participants reported on exposure to FA through vitamins, cereal, and food supplements. There was no association between taking vitamins containing FA during the periconceptional period (3 months before conception through the first trimester) and MCAs in the crude estimates or after adjusting for maternal race or ethnicity, education, gravidity, smoking, or alcohol use in the first trimester, or body mass index prior to pregnancy [adjusted odds ratio (aOR) = 1.12, 95% confidence interval (CI) 0.75–1.69]. There was also no association between vitamin exposure beginning in the first trimester and MCAs outcome (aOR = 1.05, 95% CI 0.59–1.87). In contrast to the two recently published reports, there was no association between periconceptional vitamin exposure and MCAs in the Iowa population. Published 2007 Wiley-Liss, Inc.