This study investigated whether Euphorbia subgenus Chamaesyce subsection Acutae contains C(3)-C(4) intermediate species utilizing C(2) photosynthesis, the process where photorespired CO(2) is concentrated into bundle sheath cells. Euphorbia species in subgenus Chamaesyce are generally C(4), but three species in subsection Acutae (E. acuta, E. angusta, and E. johnstonii) have C(3) isotopic ratios. Phylogenetically, subsection Acutae branches between basal C(3) clades within Euphorbia and the C(4) clade in subgenus Chamaesyce. Euphorbia angusta is C(3), as indicated by a photosynthetic CO(2) compensation point (Г) of 69 μmol mol(-1) at 30 °C, a lack of Kranz anatomy, and the occurrence of glycine decarboxylase in mesophyll tissues. Euphorbia acuta utilizes C(2) photosynthesis, as indicated by a Г of 33 μmol mol(-1) at 30 °C, Kranz-like anatomy with mitochondria restricted to the centripetal (inner) wall of the bundle sheath cells, and localization of glycine decarboxlyase to bundle sheath mitochondria. Low activities of PEP carboxylase, NADP malic enzyme, and NAD malic enzyme demonstrated no C(4) cycle activity occurs in E. acuta thereby classifying it as a Type I C(3)-C(4) intermediate. Kranz-like anatomy in E. johnstonii indicates it also utilizes C(2) photosynthesis. Given the phylogenetically intermediate position of E. acuta and E. johnstonii, these results support the hypothesis that C(2) photosynthesis is an evolutionary intermediate condition between C(3) and C(4) photosynthesis.