Persistent organic pollutants (POPs) are toxic chemical compounds that can bioaccumulate, adhere to lipid matrices, and affect human health. This study aimed to investigate the association between maternal pre-pregnancy overweight/obesity or dietary intake during pregnancy and POPs concentrations in the human milk of Brazilian women. A total of 77 women were followed from the third trimester of pregnancy until 119 days postpartum. Pregnancy dietary intake, which was estimated using a semi-quantitative food frequency questionnaire, and pre-pregnancy body mass index were collected at baseline. Human milk samples were collected between 2 and 119 days postpartum, and POPs concentrations were analysed using gas chromatography-triple quadrupole mass spectrometry. Multivariate logistic regression and linear regression models were used to investigate the association between pre-pregnancy overweight/obesity, dietary intake during pregnancy, and POPs concentrations in human milk. The analysis was adjusted for the maternal age and schooling, total cholesterol serum levels, and time postpartum and corrected for multiple comparisons using the Benjamini–Hochberg test. Significant associations were observed in the adjusted models between pre-pregnancy overweight/obesity and dichlorodiphenyldichloroethane (ppDDE), polychlorinated biphenyl (PCB)74, PCB138, PCB153, PCB170, PCB180, total PCBs, total 4PCBs, total 2 organochlorine pesticides (OCPs), and total POPs concentrations; and higher daily lipid intake during pregnancy increased human milk hexachlorobenzene (HCB). Collectively, this study showed that pre-pregnancy overweight/obesity and total lipid intake during pregnancy were associated with POPs concentrations in the milk of Brazilian women. To promote adequate nutritional status since preconception and surveillance and control of POP in the environment could be essential to ensure binomial mother-infant health.