This study describes the relationship between varying ascorbate intake, periodontal status, and subgingival microflora as part of a multidisciplinary investigation of ascorbic acid (AA) metabolism in young men housed for 13 weeks in a nutrition suite that provided controlled periods of ascorbic acid depletion and repletion. Twelve medically healthy non-smoking men, aged 25 to 43 years, ate a rotating four-day diet adequate in all nutrients except ascorbic acid. Following an initial baseline period during which the subjects received 250 mg AA/day, the subjects received 5 mg AA/day for a 32-day depletion period. Eight of the 12 subjects participated in a subsequent 56-day repletion period designed to replace the reduced body AA pool slowly. Plasma and leukocyte ascorbate levels, Plaque Index, Gingival Index, probing depths, and attachment level were monitored at the beginning and end of the depletion and repletion periods. Subgingival plaque samples were obtained and examined for selected organisms by indirect immunofluorescence microscopy. A uniform oral hygiene program was reinforced after each examination. Ascorbate concentrations in plasma and leukocytes responded rapidly to changes in vitamin C intake. There were no significant changes in plaque accumulation, probing pocket depth, or attachment level during the study. In contrast, gingival bleeding increased significantly after the period of AA depletion and returned to baseline values after the period of AA repletion. However, no relationship could be demonstrated between either the presence or proportion of target periodontal micro-organisms and measures of bleeding or ascorbate levels.