The polar microalga Chlamydomonas sp. RCC2488 (malina) exhibits high PUFA production under phototrophic conditions, but there is no evidence of heterotrophic or mixotrophic growth. Hence, the grow and transmembrane carbon (glucose) transport was assessed under heterotrophic and mixotrophic conditions. The microalga transported carbon at a rate of 0.015 μmol/g/min, achieving 0.4 g/L of biomass with high PUFA content (122.5 mg/g) under mixotrophic conditions with glucose. No heterotrophic growth and transport were observed. The use of an alternative carbon source such as potato peel waste was evaluated. This material was subjected to an acid-hydrothermal pretreatment with 1% sulfuric acid at 121 °C for 120 min followed by enzymatic hydrolysis with α-amylase and amyloglucosidase. The resulting hydrolysates (PPH1 and PPH3) released up to 46.3 g/L of glucose with a reducing sugar conversion of 46%. But these hydrolysates also presented up to 4.7 and 8.5 g/L of furfural and hydroxymethylfurfural (HMF), respectively, which decreased the microalgal growth to 0.07 g/L and promoted the accumulation of 74.7 g/g of TAG. A water-washing extra step (PPH2) removed these toxic compounds achieving biomass and PUFA concentrations similar to the ones obtained with pure glucose. Therefore, pretreatment and hydrolysis optimization including toxic by-product removal are necessary to use potato peel as an efficient carbon source for microalgae cultivation. Mixotrophic cultivation of Chlamydomonas sp. RCC2488 (malina) was possible but not ideal since higher growth is attained under phototrophic conditions.