In the presence of glycerol and PEG 4000, the loss of enzyme
activity was greater than 80.0% for both cases, indicating that they
were not effective at cryopreserving bromelain. Several investigators
have proposed that PEGs function as chaperones, which are
molecules that favor the folding of polypeptide chains into their
native structures. It has been assumed that this was only possible
to perform when protein molecules were in direct contact with the
PEG molecules [38,40]. In this specific case, bromelain could not be
interacting with PEG-4000 during the lyophilization process,
which explains the loss in enzymatic activity. In a solution containing
glycerol, when temperatures reach below 0 C, an increase in
viscosity occurs that allows more water to remain unfrozen than
in a solution without glycerol [41]. As the enzyme will not freeze,
it will remain active to exert its catalytic properties, which explains,
once again, the loss of enzyme activity during the lyophilization
process.
3.3. Fluorescence and CD measurements