In Hispanics, acculturation may lead to negative health outcomes. This study used a cross-sectional design to investigate the psychosocial and biological risks in acculturating pregnant women of Hispanic origin (n = 470). Psychosocial risks-depressive symptoms, anxiety, and stress-were assessed by self-report, whereas biological measures included stress-related and reproductive hormones. Mental health deteriorated across generations, with worsening depression, anxiety, and stress with successive generations. Stress and reproductive hormone levels decreased across generations, whereas body mass index and number of sexual partners increased. These data provide potential biobehavioral explanations of the relationship between acculturation and declining health among Hispanic women in the United States.