Nipple aspirates of breast fluid (NAF) occur with different colorations (colorless, white, pale yellow, dark yellow, brown, green, and black). Increasing concentrations of cholesterol, cholesterol 5,6-epoxides, estrogens, and fluorescent products of lipid peroxidation have been positively associated with the dark colorations (dark yellow, brown, green, and black). Because of the absence of data on these variations in breast fluid coloration, we made an exploratory study of their possible associations with age, ethnicity, clinical breast status, and breast cancer risk factors. Dark NAF colorations increased with age among white women from 22.5% at 20-29 years to 49.2% at 50-59 years. Among Chinese and Japanese women, the overall proportion of dark breast fluids was significantly lower (highest proportion 23.5%). A positive association of dark NAF coloration was found with current cigarette smoking (odds ratio = 1.64 [1.04-2.59]). A dose response between amount smoked and dark coloration was found in women less than 50 years of age, with women who smoked more than one pack per day having an odds ratio of 2.31 (1.30-4.67). No significant association of dark NAF was found with the major breast cancer risk factors or with actual benign or malignant breast disease. The dark colorations may represent pigmented products of apocrine gland secretion, lipofuscin complexes of peroxidated lipoprotein, breakdown products of hemoglobin, and possibly, diet-related secretory products.