Pre-clinical MRI/MRS routinely uses anesthesia, which alters hemodynamics and metabolism. Here, we used hyperpolarised [1-13C]pyruvate to compare brain metabolism under isoflurane or urethane anesthesia and in awake rats. Spectroscopic [1-13C]pyruvate time courses were measured in sequence in awake, isoflurane and urethane anesthetized rats. Bicarbonate- and lactate-to-total carbon ratios decreased in order from awake animals to urethane to isoflurane anesthetized animals. No change was observed in bicarbonate-to-lactate ratio between the groups, however. The study shows dDNP experiments can be performed in awake rats thus avoiding issues related to anesthesia. However, the ratios between intracellular metabolites did not alter in awake rats.