Soil carbon sequestration was analysed in the topsoil (0–0.25 m) of putting greens of different ages (5, 9, 20, 30, 40 years) in a golf course in Palmerston North, New Zealand. The soil texture was the same for all putting greens and the intensive management guaranteed that the carbon (C) inputs to the soil were very similar for all ages. Significant and linear soil C sequestration rates occurred for 40 years. The soil C sequestration rate in 0–0.25 m depth was 69 ± 8 g/m2.year over a 40-year period totalling 28 t/ha over 40 years. The relative microbial activity (dehydrogenase activity/total soil C content) representing the bioavailability of soil C decreased by about 50% over 40 years. The C sequestration and decrease of bioavailability of soil C was much more pronounced in 0.1–0.25 m depth than in the top 0.1 m. In the top 0.1 m, very little C sequestration occurred, most probably due to the intensive soil management in this depth. We concluded that the C sequestration was mainly caused by the increasing humification of C in the undisturbed part of the soil (0.1–0.25 m depth) as was indicated by a significant decrease in the relative microbial activity. Turfgrass systems such as putting greens are well suited to sequester C in urban areas.