The rapid determination of chemically reactive lysine in the presence of carbohydrates by a modified trinitrobenzenesulphonic acid procedure

The trinitrobenzene sulphonic acid (TNBS) method has been shown by previous workers to give low results for chemically reactive lysine if carbohydrates are present in the sample. The major factor responsible has now been identified as loss of ϵ-trinitrophenyl (ϵ-TNP) lysine during the hydrolysis stage, while adsorption of ϵ-TNP lysine by humins formed from carbohydrates during hydrolysis has been shown to be insignificant. The loss can be minimised by a reduction of the hydrolysis time without affecting the necessary degree of protein hydrolysis. A modified procedure is suggested which is shown to give very similar results to Carpenter's method (incorporating the Booth modification) for pure proteins and to the Silcock method in the presence of carbohydrates. ϵ-TNP lysine hydrolysed for 90 min or D,L-lysine taken through the trinitrophenylation procedure followed by hydrolysis for 120 min are the most suitable standards. The procedure can be completed in under 6 h and does not require the use of expensive equipment or hazardous chemicals.