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In contrast with conventional drugs, biopharmaceuticals are highly complex molecules with remarkable heterogeneity. Protein glycosylation is an inherent source of this heterogeneity and also affects the safety, efficacy, and serum half-life of therapeutic glycoproteins. Therefore analysis of the glycan pattern is an important issue for characterization and quality control in the biopharmaceutical industry. In this publication we describe a complete workflow for the analysis of protein N-glycans. The sample-preparation procedure, consisting of the release of the N-glycans by PNGase-F, followed by fluorescence labeling with 2-aminobenzamide and removal of excess label, was optimized to avoid alteration of the glycan sample. Subsequently, labeled glycans were analyzed by hydrophilic-interaction liquid chromatography (HILIC) with fluorescence detection. The developed method was validated for analysis of antibody N-glycans. To demonstrate the accuracy of the method an antibody sample was additionally analyzed by an orthogonal method. The antibody was digested with lysyl endopeptidase and the (glyco-)peptides were analyzed by RP-HPLC-MS. The consistency of the results between these two methods demonstrates the reliability of the glycan analysis method introduced herein.

A method for the selective modification of tryptophan residues based on the reaction of malondialdehyde with the indole nitrogen of the tryptophan side chain at acidic conditions is presented. The condensation reaction is quantitative and leads to a substituted acrolein moiety with a remaining reactive aldehyde group. As is shown, this group can be further converted to a hydrazone using hydrazide compounds, but if hydrazine or phenylhydrazine are used, release of the free indole group is observed upon cleavage of the substitution. Alternatively, secondary amines such as pyrrolidine may also act as cleavage reagents. This general reaction scheme has been adapted and optimized for the derivatization of tryptophan-containing peptides and small N-heterocyclic compounds. It serves as the basis of a reversible tagging scheme for Trp-peptides or molecules of interest carrying indole structures as it allows the specific attachment and removal of a reactive group that may be used for a variety of purposes such as affinity tagging.

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The investigation of the adhesion layer between rubber and brass-coated steel wires is a challenging task due to its strong bonding. We explore the possibilities of olefin-metathesis as a method to degrade the cross-linked rubber network without destroying the adhesion layer. Using a ruthenium catalyst and 1-octene as a co-reactant, different types of rubber—natural rubber, acrylonitrile–butadiene rubber, and styrene–butadiene rubber—can be degraded into soluble fragments. The uncovered adhesion layers can be subsequently analyzed with common analytical methods such as optical microscopy, focusvariation microscopy, and scanning electron microscopy. The revealed surface structures are discussed considering the observed pull-out forces. In a second series, the influence of common additives—cobalt salt, silica, and a resin system—on the metathesis reaction is investigated.