Biological and physical controls regulating variation of seasonal and interannual ecosystem carbon exchange in a scrub oak ecosystem in central Florida were determined by measuring net ecosystem exchange of CO2 (NEE) between the atmosphere and vegetation using the eddy covariance technique continuously for 6 years (April 2000–March 2006). Total net ecosystem production (NEP) was nearly 20 t C m−2 during the 6-year study and annual NEP for each phenological year (April–March) increased annually from 107 to 467 g C m−2 year−1. Although this ecosystem was productive during all parts of each year, greatest absolute values of daytime NEE (NEEday max) were −12.0 to −16.0 μmol CO2 m−2 s−1 in the summertime when leaf area, air temperature and soil moisture peaked. NEEday max decreased in magnitude to its lowest point in March when leaf area was lowest, and ranged between −7.6 and −9.8 μmol CO2 m−2 s−1. Mean monthly nighttime NEE (NEEnight) was between 2.0 and 6.9 μmol CO2 m−2 s−1 in the winter and summer, respectively, and was controlled primarily by temperature. Variation in seasonal NEP in this ecosystem occurred in three distinct phases. The first phase occurred in April through May with the emergence of new leaves and when soil respiration was low; daily carbon assimilation (1.0–2.4 g C m−2 day−1) was greatest during this 2-month period during which an average 38% of annual C assimilation occurred. During the second phase, June–September, the rate of carbon uptake (0.8–1.6 g C m−2 day−1) was dependent mainly on variation in temperature, precipitation and VPD; on average, only 12% of annual carbon assimilation occurred during these 4 months and in dry years (2000 and 2001), the ecosystem was a carbon source during this period. During the third phase, October–March, daily carbon assimilation (0.3–1.1 g C m−2 day−1) was intermediate between the first two phases, and accounted for nearly 50% of annual carbon accumulated during these 6 months. While mean daily NEP was highest in spring, NEEday reached its greatest intensity in summer. In order of importance, PPFD, temperature, SWC, LAI, and VPD regulated NEEday. Temperature and SWC were the main environmental variables controlling soil respiration, which was more than 85% of ecosystem respiration. Scrub oak represents a unique ecosystem in the southeastern US and within the Ameriflux network. These results are unique in establishing the role scrub oak plays in landscape and regional carbon budgets of this subtropical, evergreen, fire dependent and highly active ecosystem.