Microalgae are able to metabolize exogenous organic substrates. Differences in sets of available sources of energy and carbon cause various metabolic and physiological alterations. To characterize those in Chlamydomonas reinhardtii, we compared cells fed with acetate in batch cultures of CC-124 (wt-light) and CC-981 (mutant deficient in phosphoribulokinase, incapable to fix CO ₂), growing under illumination (prk-light) or in the darkness (prk-dark). The growth intensity of the wt-light culture was higher only at the beginning of the culture cycle. Its final density only slightly exceeded such of prk-light ones. The growth rate and final density of the prk-dark culture were several times lower. The GC-MS analysis revealed clear metabolomic differences in cultures with different trophic status. Cells of prk-light were characterized by higher content of fatty acids, sterols, glycolate, malate, and putrescine. Cells of prk-dark accumulated, in particular, relatively higher levels of arginine, ornithine, AMP, some other nitrogen-containing compounds, glycerol, citrate, and sucrose. Higher levels of some amino acids and threonic acid were detected in wt-light cells. Simultaneously, we tested expression of genes encoding key enzymes of plant metabolism. The expression of RBCS1 (Rubisco small subunit), CIS1, 2 (citrate synthase), AAA1 (chloroplast ATP/ADP antiporter), APE2 (triose phosphate translocator), OMT1 (oxoglutarate/malate antiporter) was analyzed. The comparison revealed that RBCS1 was drastically upregulated in prk-light cultures. The expression level of AAA1 in prk-dark cells slightly exceeded such in wt-light algae, but was almost zero in prk-light cells. The level of CIS1, 2, APE2 and OMT1 was maximal in wt-light cells.