The application of metabolic engineering principles to the rational design of microbial production processes crucially depends on the ability to make quantitative descriptions of the systemic ability of the central carbon metabolism to redirect fluxes to the product-forming pathways. The aim of this work was to further our understanding of the steps controlling the biotransformation of trimethylammonium compounds into l-carnitine by Escherichia coli. Despite the importance of l-carnitine production processes, development of a model of the central carbon metabolism linked to the secondary carnitine metabolism of E. coli has been severely hampered by the lack of stoichiometric information on the metabolic reactions taking place in the carnitine metabolism. Here we present the design and experimental validation of a model which, for the first time, links the carnitine metabolism with the reactions of glycolysis, the tricarboxylic acid cycle and the pentose-phosphate pathway. The results demonstrate a need for a high production rate of ATP to be devoted to the biotransformation process. The results demonstrate that ATP is used up in a futile cycle, since both trimethylammonium compound carriers CaiT and ProU operate simultaneously. To improve the biotransformation process, resting processes as well as CaiT or ProU knock out mutants would yield a more efficient system for producing l-carnitine from crotonobetaine or d-carnitine.