title: Integrity of crystalline lysozyme exceeds that of a spray-dried form.
abstract: The development of proteins as therapeutic agents is challenging partly due to their inherent instabilities. Consequently, crystallisation and spray drying techniques were assessed to determine their effects on protein integrity using lysozyme as a model protein. Unprocessed, crystallised and spray-dried lysozyme were characterised by: thermal analysis using hot stage microscopy (HSM), differential scanning calorimetry (DSC), high sensitivity differential scanning calorimetry (HSDSC) and thermogravimetry (TGA); and spectroscopic analysis employing Fourier transform Raman (FT-Raman). Moisture contents were determined by TGA and Karl Fisher titration (KFT). Enzymatic assay measured biological activity. HSM showed no changes in crystals until complete melting. TGA and KFT indicated that spray-dried lysozyme contained a lower moisture content than crystals, hence the higher apparent thermal stability was shown by DSC. HSDSC revealed that crystallisation and spray drying did not affect the denaturation temperature of lysozyme in solution when compared with unprocessed material. However, in the solid state, FT-Raman spectra showed perturbation of the conformational structure of spray-dried sample, whereas crystal conformation remained intact. Enzymatic assay revealed increased activity retention of crystals compared with spray-dried powder. Hence, crystals maintained the conformational integrity and activity of lysozyme in solution.
authors.affiliation: Drug Delivery Group, School of Pharmacy, University of Bradford, Bradford BD7 1DP, UK.
authors.forename: Amal A
authors.initials: AA
authors.lastname: Elkordy
authors.forename: Robert T
authors.initials: RT
authors.lastname: Forbes
authors.forename: Brian W
authors.initials: BW
authors.lastname: Barry
mesh_terms: D003460:Crystallization / Q000379:methods; D004795:Enzyme Stability; D006358:Hot Temperature; D009113:Muramidase / Q000737:chemistry*; D013817:Thermography / Q000379:methods
chemical_list: D009113:Muramidase