The only available ethnobotanical information on Pteronia incana has been recorded by the Montagu Museum in the Western Cape Province of South Africa. It was reported that the plant is used to treat influenza, fever, kidney ailments and backache. In common with other species of Pteronia, the plant contains an essential oil reminiscent of pine turpentine oil with b-pinene, limonene, 1,8-cineole, myrcene, spathulenol, p-cymene and methyleugenol as main compounds present in all or most of the samples, with smaller amounts of α-pinene, sabinene, g-terpinene, terpinen-4-ol, biclogermacrene, globulol and α-bisabolol in some of the distillates. We investigated the oil composition of 11 individual plants collected at three geographically distant localities but found limited variation, both within and between populations. Leaf sections of P. incana showed that it is anatomically similar to P. divaricata in the presence of a secretory duct associated with the main vascular bundle (and often other bundles as well), in addition to glandular and non-glandular trichomes on both leaf surfaces. One yeast (Cryptococcus neoformans), two Gram-negative bacteria (Moraxella catarrhalis and Klebsiella pneumoniae) and one Gram-positive bacterium (Mycobacterium smegmatis) were selected for antimicrobial activity studies using the minimum inhibitory concentration (MIC) microtitre plate method. The results showed that methanol:dichloromethane (MeOH:CH2Cl2) extracts were active against M. smegmatis (lowest MIC values of 0.5–0.8 mg/ml) and C. neoformans (lowest MIC values of 0.5–2.0 mg/ml). The essential oil was most active against C. neoformans (lowest MIC value of 0.3 mg/ml). These results provide a scientific rationale for the use of Pteronia incana in Cape herbal medicine.