Coat colour is one of the most important qualitative traits of fur animals. Determining melanin pigments forming the basics of visible coat colour may contribute to a better understanding of the process of creating different coat colour variations in fur-bearing animals. This study aimed to (i) isolate pigment cells from the hair of American mink of 11 colour variations (standard brown, silverblue, palomino, black, wild type, sapphire, black cross, pearl, palomino cross, glow, and amber) using acid and alkali; and (ii) characterise the melanin pigments obtained. The purified pigment cells were observed under a light microscope and verified by spectrophotometry scanning and nuclear magnetic resonance spectroscopy. The method allowed for obtaining pure melanin specimens. Using acid and alkali to extract eumelanosomes did not affect their shape and structure; it also allowed for obtaining pheomelanin from the hair. The results have proven that the hair colour of the American mink is based on all types of melanin, and that its variations differ in terms of how much eumelanin and pheomelanin the hair contains.