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Detection of bacterial and fungal contamination is of extreme importance in the fields of medical products or food packaging. Here a diagnostic tool based on pectinase and cellulase triggered release of a dye from a crosslinked polysaccharide matrix was developed. The hydrogel-based bioresponsive matrix consisted of carboxymethylcellulose (CMC) as a substrate for cellulases and polygalacturonate (PGA) as substrate for pectinases. To improve the stability of the hydrogels, methacrylic groups were inserted as crosslinking molecules. For polymerisation, two different methods were used, namely UV and thermal crosslinking. Controlled release triggered by extracellular enzymes of potentially pathogenic/contaminating microorganisms was investigated by the incorporation of Alizarin into the hydrogels. UV polymerised hydrogels turned out to be more suitable than thermal crosslinked polymers. Integration of such polymer based bioresponsive systems in medical surfaces or package systems could therefore act as an in situ monitoring system for detecting the presence of bacteria or fungi like e.g. Aspergillus species .

Bioresponsive polymers (BRPs) allow the detection of potentially pathogenic microorganisms. Here, peptidoglycan and cellulose based hydrogels were constructed with potential for diagnosis of wound infection or, for example, Aspergillosis, respectively. These systems respond to extracellular enzymes from microbes or enzymes secreted from the human immune system in case of infection. Laccases as ‘enhanzymes’ were incorporated into these devices for signal and stability enhancement when compared to simple dye release based systems. To retain the enhanzymes within the BRPs, they were either PEGylated laccase (Laccase_PEG) to increase size or methacrylated laccase (Laccase_MA) to allow covalent attachment to the polysaccharide matrices. PEGylation of Trametes hirsuta laccase led to a fivefold increase in size to 270 kDa according to size exclusion chromatography (SEC). Likewise, successful methacrylation of the laccase was demonstrated by using reversed phase chromatography while SEC analysis proved covalent attachment of the enzyme to the methacrylated polysaccharide matrix. Upon incubation of peptidoglycan based BRPs with fluid from infected wounds, the difference to controls was four times higher for Laccase_PEG based signalling when compared to simple dye release. Similarly, the control signals (i.e. leaching) were considerably reduced in case of Laccase_MA incorporated in crosslinked peptidoglycan (PG) and carboxymethylcellulose (CMC) hydrogels for signalling. In addition, Laccase_MA catalysed colour formation enhanced the signal dramatically with factors between 100- and 600-fold. Laccase_MA was demonstrated to oxidise silica gel immobilised ferulic acid incorporated into the BRP with clearly visible colour changes of 4.5 Δ E units according the CIELab concept upon incubation by trigger enzymes as well as infected wound fluids.