The assessment of differential left and right lung function is important for patients under consideration for lung resection procedures such as single lung transplantation. We developed an automated, knowledge-based segmentation algorithm for purposes of deriving functional information from dynamic computed tomography (CT) image data. Median lung attenuation (HU) and area measurements were automatically calculated for each lung from thoracic CT images acquired during a forced expiratory maneuver as indicators of the amount and rate of airflow. The accuracy of these derived measures from fully automated segmentation was validated against those from segmentation using manual editing by an expert observer. A total of 1313 axial images were analyzed from 49 patients. The images were segmented using our knowledge-based system that identifies the chest wall, mediastinum, trachea, large airways and lung parenchyma on CT images. The key components of the system are an anatomical model, an inference engine and image processing routines, and segmentation involves matching objects extracted from the image to anatomical objects described in the model. The segmentation results from all images were inspected by the expert observer. Manual editing was required to correct 183 (13.94%) of the images, and the sensitivity, specificity, and accuracy of the knowledge-based segmentation were greater than 98.55% in classifying pixels as lung or nonlung. There was no significant difference between median lung attenuation or area values from automated and edited segmentations (p > 0.70). Using the knowledge-based segmentation method we can automatically derive indirect quantitative measures of single lung function that cannot be obtained using conventional pulmonary function tests.