AbstractBiotransformation of (±)‐threo‐7,8‐dihydroxy(7,8‐2H2)tetradecanoic acids (threo‐(7,8‐2H2)‐3) in Saccharomyces cerevisiae afforded 5,6‐dihydroxy(5,6‐2H2)dodecanoic acids (threo‐(5,6‐2H2)‐4), which were converted to (5S,6S)‐6‐hydroxy(5,6‐2H2)dodecano‐5‐lactone ((5S,6S)‐(5,6‐2H2)‐7) with 80% e.e. and (5S,6S)‐5‐hydroxy(5,6‐2H2)dodecano‐6‐lactone ((5S,6S)‐5,6‐2H2)‐8). Further β‐oxidation of threo‐(5,6‐2H2)‐4 yielded 3,4‐dihydroxy(3,4‐2H2)decanoic acids (threo‐(3,4‐2H2)‐5), which were converted to (3R,4R)‐3‐hydroxy(3,4‐2H2)decano‐4‐lactone ((3R,4R)‐9) with 44% e.e. and converted to 2H‐labeled decano‐4‐lactones ((4R)‐(3‐2H1)‐ and (4R)‐(2,3‐2H2)‐6) with 96% e.e. These results were confirmed by experiments in which (±)‐threo‐3,4‐dihydroxy(3,4‐2H2)decanoic acids (threo‐(3,4‐2H2)‐5) were incubated with yeast. From incubations of methyl (5S,6S)‐ and (5R,6R)‐5,6‐dihydroxy(5,6‐2H2)dodecanoates ((5S,6S)‐ and (5R,6R)‐(5,6‐2H2)‐4a), the (5S,6S)‐enantiomer was identified as the precursor of (4R)‐(3‐2H1)‐ and (2,3‐2H2)‐6). Therefore, (4R)‐6 is synthesized from (3S,4S)‐5 by an oxidation/keto acid reduction pathway involving hydrogen transfer from C(4) to C(2).In an analogous experiment, methyl (9S,10S)‐9,10‐dihydroxyoctadecanoate ((9S,10S)‐10a) was metabolized to (3S,4S)‐3,4‐dihydroxydodecanoic acid ((3S,4S)‐15) and converted to (4R)‐dodecano‐4‐lactone ((4R)‐18).