The reaction was first reported by Frederick Gowland Hopkins and in 1901, as part of their work on the first isolation of tryptophan itself. Nitrites, chlorates, nitrates and excess chlorides prevent the reaction from occurring. A purple ring appears between the two layers if the test is positive for tryptophan. Concentrated sulfuric acid is slowly added to form two layers. A protein solution is mixed with Hopkins Cole reagent, which consists of glyoxylic acid.
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