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PEGylation is an established method for improving the pharmacokinetic properties and pharmacodynamic effects of therapeutic proteins. Amino conjugation by N-hydroxysuccinimide activated polyethylene glycol (PEG-NHS) represents the most common modification of the target proteins. In this study, we compared the reaction velocities of pH-dependent preparations of bovine lactoferrin modified with branched PEG-NHS conducted at pH 7.4 to pH 9.0. PEGylation reaction rates were dependent on pH value, but not on PEG molecular mass. At pH 7.4, reactions advanced gradually and reached steady state by 2 h, whereas at pH 9.0, reactions advanced very quickly and reached steady state within 10 min. Hydrolysis half-life of PEG-NHS exceeded 120 min at pH 7.4, but was below 9 min at pH 9.0. Thus, the pH value is one of the most important factors to obtain the optimal condition for PEGylation by NHS esters.

The pH-Dependent Formation of PEGylated Bovine Lactoferrin by Branched Polyethylene Glycol (PEG)-N-Hydroxysuccinimide (NHS) Active Esters