The effects of pH and total dissolved inorganic carbon (DIC) availability on the growth and biochemical composition of Phaeodactylum tricornutum Bohlin acclimated to restricted and saturated light conditions have been investigated. Diluted, unbuffered and nutrient enriched batch cultures were grown at several pHs (different [CO 2( aq ) ] for the same DIC) and various DIC conditions (different [CO 2( aq ) ] for an equal pH) under saturating (150 μmol m −2 s −1 ) and subsaturating (30 μmol m −2 s −1 ) light levels. The growth rate was unaffected by pH treatments at saturating light. In contrast, under subsaturating light conditions, the growth rate was significantly reduced at pHs over 8.5 and also at low DIC levels to a similar extent. Biochemical composition revealed that the growth rate was limited by different resources. At natural DIC (DIC = 2.1 mM) and low light conditions, there was an increase of the C:N ratio under high pH (low [CO 2( aq ) ]) associated with a reduction of nitrogen, chlorophyll a and total protein content, resembling the biochemical composition of a diatom cultured under nitrogen limitation. On the contrary, under low light and low DIC conditions, in which both [CO 2( aq ) ] and [HCO 3 − ] are reduced, cellular carbon content decreased for a relatively constant nitrogen content, decreasing the C:N ratio. The results are discussed in terms of competition between carbon and nitrogen metabolism for energy, and showed that the imposition of different [CO 2( aq ) ] by changing pH or DIC levels of seawater affected in different ways the final biochemical composition of Phaeodactylum tricornutum .