Gas sparging was found to be a useful technique to reduce hydrogen partial pressure in the liquid phase to enhance the hydrogen yields of strictly anaerobically fermentative bacteria. The effect of nitrogen (N(2)) sparging on hydrogen yield was investigated in sterile and non-sterile conditions using a pure strain of the hyperthermophilic eubacteria, Thermotoga neapolitana with glucose or xylose as a carbon source. The maximum hydrogen accumulations reached 41% of the gaseous mixtures after 30-40 h. Two applications of N(2) sparging after the H(2) content in the headspace reached the maximum levels gave an increase of H(2) production by 78% from 1.82 to 3.24 mol H(2)/mol glucose and by 56% from 1.41 to 2.20 mol H(2)/mol xylose. This result suggested that the removal of the produced H(2) from the gas headspace of the limited-volume, closed culture vial when it achieves the maximum level of H(2) tolerance of the bacterium is a necessary technique to improve its H(2) yield.