Added volume of concentrated sulphuric acid was varied; 0.5, 1, 2, 3, 4, 5, 6, 7 and 8 mL, while amounts of reactants were kept constant at: 0.25 mL glyoxylic acid 0.028 M solution, 0.6 mL tryptophan 0.016 M solution and 2 mL ferric chloride 0.025M solution. The final volume of liquid in tubes was corrected with distilled water. Test procedure was described in the experimental work section. Figure shows an increase of optical absorbance up to 5 mL sulphuric acid addition. Which means that sufficient amount of concentrated sulphuric acid must be present in order to absorb reaction-and hydration water. Otherwise the aldehyde group of glyoxylic acid may be hydrated to germinal diol (K=300 at 25 o C), and eventually further dimerized to hemiacetal, thus making it not be available for reaction with tryptophan.