The ability of three species of Xanthobacter to metabolize cyclohexane and its derivatives has been compared. Xanthobacter flavus was unable to utilize any of the cycloalkanes under investigation. X. autotrophicus was unable to utilize cyclohexane but was able to grow with a limited range of substituted cycloalkanes, including cyclohexanol and cyclohexanone. Comparison of a previously isolated cyclohexane growing Xanthobacter sp. with X. flavus and X. autotrophicus indicated it to be closely related to X. autotrophicus. Studies with cell-free extracts have indicated that the route of metabolism for cyclohexanol by X. autotrophicus is the same as that shown for the cyclohexane growing Xanthobacter sp., proceeding via cyclohexanol-4cyclohexanone-* ecaprolactone-*-* adipic acid. A comparison of the cyclohexanol dehydrogenase found in X. autotrophicus with that found in the cyclohexane-growing Xanthobacter sp. indicated these enzymes to be distinctly different from one another on the basis of substrate specificity, molecular weight, and pH optima. The cyclohexanone monooxygenase enzymes found in the two bacteria were also found to be different when the pH optima and cofactor specificity of the two enzymes were compared. Preliminary genetic studies on the cyclohexane-growing Xanthobacter sp. have indicated that there are no plasmids present in this bacterium. The presence of RP4 in the Xanthobacter sp. can be detected following its conjugation with an RP4-carrying Escherichia coli strain.