New preparative methods that provide a reliable source for stable-isotope labeled amino acids suitable for NMR study of membrane proteins are needed for further advances in this field. Phase I demonstrated the synthetic steps leading to optically pure, 13C-labeled leucine, tyrosine and threonine as proposed. The synthesis involved modifications of existing methods to obtain >99% diastereomeric excess and eliminate the need for further chromatographic purification. Subsequent hydrolytic steps were effected without detectable racemization. The chiral auxiliaries were satisfactorily recovered. The results suggest the selected approach as a viable commercial production method. Phase II aims to continue developing the preparative methods for all 21 major proteinogenic amino acids each 13C-labeled at every other position. Synthesis of the requisite labeled side chains will be facilitated by the availability of some new labeled precursors recently developed in our lab. In addition, a new simple extractive technique for separating and recovering the expensive chiral auxiliary. which should lower the production costs considerably. will be pursued. The amino acids will have a 99% enantiomeric excess minimum and 99% 13C-enrichment at all intended positions.
In addition to NMR, the labeled amino acids are also useful for metabolic study and as internal standards for GC-MS quantitative analysis. Other potential spin-offs are promising. The commercial potentials appear quite considerable.
