Carbodiimide Induced Cross-Linking, Ligand Addition, and Degradation in Gelatin

Abstract

The water-soluble carbodiimide, 1-ethyl-3-(3-(dimethylaminopropyl)-carbodiimide (EDC) is widely used in protein chemistry. We used EDC-induced gelatin cross-linking as a model for amide bond formation to resolve reaction ambiguities with common variables of buffers, gelatin concentration, and pH. Percentage changes in SEC high molecular weight peak areas were used to follow the reactions. Differences in reaction rate and extent were observed with four commonly used buffers, while differences in extent were observed for commonly used concentrations and pH. We also investigated an anhydride mechanism for aqueous EDC-induced amide bond formation that has received little attention since its proposal in 1995. Gelatin carboxyl groups had a synergistic role during the addition of hydrazine to corroborate the anhydride formation between carboxyl groups. EDC-induced degradation of gelatin was investigated using percentage changes in SEC low molecular weight peak areas. The degradation occurred in excess EDC at neutral to alkaline pH and was enhanced substantially when reacting amino groups were not available. A mechanism of EDC-induced gelatin degradation is proposed and designated the extended Khorana mechanism. This EDC side reaction has the potential to occur in peptides and proteins under similar conditions.

A Biomimetic Silk Fibroin/Sodium Alginate Composite Scaffold for Soft Tissue Engineering

“…These scaffolds seemed to be white sponges, which can be resistant to the water. According to the previous reports3536, reaction mechanism among EDC, SF and SA was hypothesized as shown in Fig. 1(b).…”

Development of tannin-inspired antimicrobial bioadhesives

“…Typically, the modification of gelatin for adhesives employs 1-Ethyl-3-(3-dimethylamino -propyl)carbodiimide (EDC) coupling or vinyl groups (e.g. methacrylated gelatins), but former method can self-crosslink gelatin [36–39], while the latter requires a large excess of methacrylic anhydride and subsequent dialysis [40]. Here, we demonstrate that gelatin modification can be performed with tannic acid (TA) through a one-step Michael addition reaction that is low-cost, convenient, and scalable.…”

Insulin released from titanium discs with insulin coatings-Kinetics and biological activity

“…Other factors that may play a role is to use fresh immobilization chemicals, where especially EDC/NHS is prone to spontaneous degradation in aqueous solutions. Also, too high EDC concentrations may participate in protein degradation and removal from surfaces . Both the above mechanisms could explain the occasional observation of decreased coating thicknesses upon a large number of EDC/NHS and insulin incubations.…”