Summary: Internal solutes in a halotolerant Brevibacterium sp. (strain JCM 6894) were analysed by 1H NMR spectroscopy. At the lower osmolarity of complex medium, the main compatible solutes in the cells were glutamate, glycine betaine and hydroxyproline. At higher external salt concentrations, ectoine (1,4,5,6-tetrahydro-2-methyl-4-pyrimidinecarboxylic acid) levels increased and this became the predominant compatible solute at 1.5 M NaCl, both ectoine and hydroxyectoine became dominant, as was also the case in complex medium. Additional compatible solutes detected were γ-aminobutyrate and proline, neither of which were detected in cells grown in complex medium. Regardless of the medium composition and incubation time, ectoine levels were high at high osmolarity, indicating that the halotolerant nature of Brevibacterium sp. is closely related to its accumulation of ectoine. From its accumulation of compatible solutes, we suggest that Brevibacterium sp. JCM 6894 is a different species from other members of the genus Brevibacterium.