Lessonia nigrescens used to be an abundant kelp species along the Chilean coast, but recent molecular studies revealed the existence of a L. nigrescens species complex, which includes the two cryptic species Lessonia berteroana and Lessonia spicata. Since these species have different distributions (16°S–30°S for L. berteroana and 29°S–42°S for L. spicata), they experience differences in environmental conditions, such as solar irradiance, seawater temperature and air exposure during low tide. This study tested to what extent the genetic distinctness of each of the two species [identified by a mitochondrial marker (atp8/trnS)] is reflected by ecophysiological traits (total lipids, fatty acid composition, phlorotannins, pigments and variable chlorophyll a fluorescence of PSII) in response to the respective environmental conditions, prevailing along the latitudinal gradient. We studied algal individuals from eight populations (27°S–32°S, including the species overlapping zone). Phlorotannins, pigments and Chl a fluorescence of PSII were most crucial for species-specific adaptations at the respective growth sites, whereas changes in total lipids and fatty acid compositions were negligible. Hence, species differentiation within the L. nigrescens complex is also manifested at the ecophysiological level. These findings may help to predict kelp responses towards future environmental changes.