A rapid and direct method for the quantitative determination of tryptophan in the intact protein

Abstract

1. A method is given for the quantitative determination of free tryptophan or tryptophan in the intact protein by treating with ninhydrin in a mixture of formic acid and hydrochloric acid (reagent b), for 10min at 100°C. Glycyltryptophan was used as a standard for the determination of tryptophan in the intact protein. The extinction at 390nm was linear in the range 0.05–0.5μmol for free tryptophan (∈7120) and 0.05–0.30μmol for glycyltryptophan (∈15400). 2. Free tryptophan in the presence of protein may be determined by treating with ninhydrin in a mixture of acetic acid and 0.6m-phosphoric acid (reagent a) for 10min at 100°C, the extinction being linear for tryptophan in the range 0.05–0.9μmol. N-Terminal tryptophan peptides also give the typical yellow product on treatment with reagent a. 3. Tryptophan content of several pure intact proteins when treated with the above method gave values in good agreement with those reported by others. A mean tryptophan content of 11.25 (s.e.m. ±0.08) μmol/100mg of protein was found in rat brain during development from 1 to 82 days after birth.