Since its discovery, Hopkins-Cole test has been neither easy nor reliable for tryptophan quantitative measurements due to slow rate of product formation, various color evolutions and low stability of the product. This pushed copious researchers to suggest many improvements as in . In auxiliary materials such as copper sulphate were added that gave dark crimson-violet color product. was the pioneer in the description of the crimson-violet color absorption at 560 nm. Other researchers studied the compounds and stoichiometry that can form between organic aldehydes and tryptophan. Quantitative determination of tryptophan in proteins succeeded by addition of mild oxidants such as persulfate and Fe 3+ ions, which accelerated chromogen appearance and increased its stability .