AbstractCalorimetry and other on‐line techniques are used for the first time as complement to the traditional off‐line methods in order to follow the growth of the green Chlorella vulgaris microalgae. A 2‐L photo‐bio‐reactor was adapted from a commercial calorimeter used previously to study heterotrophic microbial growth. An external source of light was added to favor the photosynthesis of the autotrophic cells. Heterotrophic growth was also tested with external glucose in the broth. A third mode, mixotrophic, allowed faster autotrophic plus heterotrophic growth. Calorimetric measurements were performed considering the corresponding calibrations in order to consider only the energy involved during the microalgal growth. The three different modes of Chlorella cultures were energetically characterized. Besides calorimetry, the weight of diluted nitric acid added to maintain the pH of the culture was correlated with the cellular growth and the nitrogen composition of the algae. Additionally, the on‐line infrared spectroscopy proved to be an efficient technique to follow the composition of the broth in glucose, nitrates, and phosphates. These results were compared and complemented with some classic off‐line techniques used to track this kind of cultures. Biotechnol. Bioeng. 2007;96:757–767. © 2006 Wiley Periodicals, Inc.