Synthesis of specifically labelled L‐phenylalanines using phenylalanine ammonia lyase activity

Specifically labelled L‐phenylalanines have been prepared using a variety of classical synthetic methods in combination with phenylalanine ammonia lyase (PAL) enzyme activity of the yeast Rhodosporidium toruloides ATCC 10788 or Rhodotorula glutinis IFO 0559, respectively. Thus, L‐[2‐ 2 H]phenyl‐[2‐ 2 H]alanine ( 5 ) was formed from (E)‐[2,2′‐ 2 H 2 ]cinnamic acid and ammonia in 46% yield, whereas L‐phenyl‐[2‐ 13 C, 15 N]alanine ( 9 ) was obtained from (E)‐[2‐ 13 C]cinnamic acid in 45% overall yield using an 8.3‐fold excess of 15 NH 4 Cl and repeating the reaction after recovery of unchanged labelled material. Generally, labelled cinnamic acids were recovered in pure form from the reaction mixture, with a loss of 6‐8%. Likewise, unchanged 15 NH 3 was reisolated as 15 NH 4 Cl after steam distillation with overall losses of less than 4%. Labelled cinnamic acids were prepared by Knoevenagel condensations between appropriately labelled benzaldehydes and malonic acids. [2‐ 2 H]Benzaldehyde was obtained from 2‐bromotoluene by decomposition of the corresponding Grignard reagent with 2 H 2 O and subsequent oxidation. Since simple molecules, most of them commercially available in labelled form or otherwise easily accessible, may serve as starting material, and due to its defined stereochemistry, the reaction catalyzed by PAL opens a short and attractive route to specifically labelled L‐phenylalanines.