The distinctive and visually striking wooden masks associated with the Bwa culture in Burkina Faso, West Africa, are carved from a soft wood into different shapes and display various geometrical patterns and symbols according to their purpose. One of their characteristic features is the use of the colors black, red, and white, which evoke the three major rivers crossing the country: the Black, Red, and White Voltas. According to published accounts of scholars who have worked directly with the artists, the materials used to obtain these colors include reptile excrement for the white, iron-rich stones powdered and mixed with egg or plant gums for the red, and boiled Acacia seed pods for the black, as well as modern materials such as enamel paint in some cases. A group of four Bwa masks in the Arts of Africa collection of the Art Institute of Chicago was investigated using a complement of analytical techniques including Fourier transform infrared spectroscopy, scanning electron microscopy with energy dispersive spectroscopy, pyrolysis gas chromatography–mass spectrometry, and mass spectrometry-based proteomics to characterize their painting materials. The results obtained corroborate the published accounts, while also providing new insights into the nature of the coloring materials and the selection and substitution of pigments and binders. These findings highlight the complementary value of scientific research, in combination with fieldwork and artists’ accounts, to generate a fuller understanding and appreciation of this traditional artistic practice.