A direct assay for evaluation of polyethylene glycol in enzyme adducts used as drugs or biocatalysts.

A specific and direct method for the evaluation of monomethoxypolyethylene glycol (MPEG) in enzyme adducts has been developed. The method is based on extensive modification by trinitrobenzensulfonic acid (TNBS) of the MPEG-enzyme, to form an acid-stable TNBS adduct with the available amino groups of lysine and alpha-amino acid. The MPEG-modified trinitrophenylated enzyme was hydrolyzed in 6N HCl and amino acid composition evaluated by a standard AA analyzer in comparison with the analysis of a MPEG enzyme sample. The number of bound MPEG polymers was calculated from the difference in amino acid composition of the two samples.