title: Effects of cooling rate in microscale and pilot scale freeze-drying - Variations in excipient polymorphs and protein secondary structure.
abstract: Microscale freeze-drying makes rapid process cycles possible for early-stage formulation development. To investigate the effects of equipment scale and cooling rate on the solid state properties and the protein's secondary structure of a sample, three binary formulations of catalase were prepared and freeze-dried with sucrose, mannitol, or (2-hydroxypropyl)-β-cyclodextrin (HP-β-CD). The protein's secondary structure was assessed using attenuated total reflection Fourier transform infrared spectroscopy (FTIR-ATR). The solid state properties were assessed using differential scanning calorimetry (DSC) and X-ray diffraction (XRD). The results were interpreted with respect to the biological activity of catalase after its reconstitution. According to the results of both the protein secondary structure and the reconstituted biological activity, scale-up could be achieved with the sucrose-catalase formulation when it was prepared at a high cooling rate and with the mannitol-catalase formulation when prepared at a low cooling rate. However, differences in the polymorph composition of crystalline mannitol were noted. No cooling rate influence was found with the HP-β-CD formulation. The results clearly indicate that the effects of the cooling rate should be closely examined during microscale formulation development and scale-up of the freeze-drying process.
authors.affiliation: University of Eastern Finland, School of Pharmacy, Kuopio, Finland. Electronic address: bjorn.peters@uef.fi.
authors.forename: Björn-Hendrik
authors.initials: BH
authors.lastname: Peters
authors.affiliation: Ghent University, Ghent, Belgium.
authors.forename: Lotte
authors.initials: L
authors.lastname: Staels
authors.affiliation: University of Eastern Finland, Department of Applied Physics, Kuopio, Finland.
authors.forename: Jimi
authors.initials: J
authors.lastname: Rantanen
authors.affiliation: University of Eastern Finland, School of Pharmacy, Kuopio, Finland.
authors.forename: Ferdinand
authors.initials: F
authors.lastname: Molnár
authors.affiliation: Ghent University, Ghent, Belgium.
authors.forename: Thomas
authors.initials: T
authors.lastname: De Beer
authors.affiliation: University of Eastern Finland, Department of Applied Physics, Kuopio, Finland.
authors.forename: Vesa-Pekka
authors.initials: VP
authors.lastname: Lehto
authors.affiliation: University of Eastern Finland, School of Pharmacy, Kuopio, Finland.
authors.forename: Jarkko
authors.initials: J
authors.lastname: Ketolainen
mesh_terms: D003080:Cold Temperature; D004339:Drug Compounding; D005079:Excipients / Q000032:analysis / Q000737:chemistry*; D005612:Freeze Drying / Q000379:methods* / Q000639:trends; D017433:Protein Structure, Secondary; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods / Q000639:trends; D014961:X-Ray Diffraction / Q000379:methods / Q000639:trends
chemical_list: D005079:Excipients
keywords: Calorimetry (DSC); FTIR-ATR; Freeze-drying; Multivariate analysis; Protein formulation; Scale-up; X-ray diffraction (XRD)