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We report the first data on 25-hydroxyvitamin D plasma levels in natural populations of three species of land iguana endemic to the Galápagos Islands ( Conolophus marthae , C. subcristatus , and C. pallidus ). The pigment is present throughout the whole body in the skin of C. subcristatus and C. pallidus . On the contrary, pigment is not present in the skin of an extended part of the body in C. marthae . The only existing population of C. marthae is syntopic with a population of C. subcristatus , and the two species are closely related. These circumstances would suggest that, under the assumption that the species show a similar basking behavior and in the absence of compensatory mechanisms, lighter pigmentation should favor higher vitamin D levels. Thus, C. marthae , compared with C. subcristatus in Wolf Volcano, could show higher levels of 25(OH)D plasma levels, or equal, if compensatory mechanisms exist. The three species showed levels in the range of average values for healthy iguanas. However, contrary to the expectation, C. marthae consistently exhibited the lowest 25(OH)D plasma levels. We discuss possible factors affecting vitamin concentration and hypothesize that C. marthae may use the habitat to limit exposure to the high UVB irradiation at Wolf Volcano., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Cristina Di Giacomo et al.)