We have previously described the isolation of the new bacterial species, Ralstonia/Burkholderia sp. strain DSM 6920, which grows with 6-methylnicotinate and regioselectively hydroxylates this substrate in the C2 position by the action of 6-methylnicotinate-2-oxidoreductase to yield 2-hydroxy-6-methylnicotinate (Tinschert et al. 1997). In the present study we show that this enzymatic activity can be used for the preparation of a series of hydroxylated heterocyclic carboxylic acid derivatives. The following products were obtained from the unhydroxylated educts by biotransformation using resting cells: 2-hydroxynicotinic acid, 2-hydroxy-6-methylnicotinic acid, 2-hydroxy-6-chloronicotinic acid, 2-hydroxy-5,6-dichloronicotinic acid, 3-hydroxypyrazine-2-carboxylic acid, 3-hydroxy-5-methylpyrazine-2-carboxylic acid and 3-hydroxy-5-chloropyrazine-2-carboxylic acid. Thus the respective educts were all regioselectively mono-hydroxylated at the carbon atom between the ring-nitrogen and the ring-carbon atom carrying the carboxyl group. In contrast to its relatively broad biotransformation abilities, the strain shows a limited heterocyclic nutritional spectrum. It could grow only with three of the seven transformed educts: 6-methylnicotinate, 2-hydroxy-6-methylnicotinate and 5-methylpyrazine-2-carboxylate. 2-Hydroxynicotinate, 2-hydroxy-6-chloronicotinate, 2-hydroxy-5,6-dichloronicotinate, 3-hydroxypyrazine-2-carboxylate and 3-hydroxy-5-chloropyrazine-2-carboxylate were not degraded by the strain. Therefore, unlike 6-methylnicotinate-2-oxidoreductase, which has a broad substrate spectrum, the second enzyme of the 6-methylnicotinate pathway seems to have a much more limited substrate range. Among 28 aromatic heterocyclic compounds tested as the sole source of carbon and energy, only pyridine-2,5-dicarboxylate was found as a further growth substrate, and this was degraded by a pathway which did not involve 6-methylnicotinate-2-oxidoreductase. To the best of our knowledge the microbial production of 2-hydroxy-6-chloronicotinic acid, 2-hydroxy-5,6-dichloronicotinic acid and 3-hydroxy-5-methylpyrazine-2-carboxylic acid have not been reported before. Strain DSM 6920 is so far the only known strain which allows the microbial production of both these compounds and 3-hydroxypyrazine-2-carboxylic acid and 3-hydroxy-5-chloroypyrazine-2-carboxylic acid.