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 270kDa 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 ΔE units according the CIELab concept upon incubation by trigger
enzymes as well as infected
wound fluids.