Pharmaceutical Research

, Volume 16, Issue 5, pp 702–708

A Central Composite Design to Investigate the Thermal Stabilization of Lysozyme

  • Sébastien Branchu
  • Robert T. Forbes
  • Peter York
  • Håkan Nyqvist
Article

DOI: 10.1023/A:1018876625126

Cite this article as:
Branchu, S., Forbes, R.T., York, P. et al. Pharm Res (1999) 16: 702. doi:10.1023/A:1018876625126

Abstract

Purpose. The formulation and processing of protein drugs requires the stabilization of the native, biologically active structure. Our aim was to investigate the thermal stability of a model protein, lysozyme, in the presence of two model excipients, sucrose and hydroxypropyl-β-cyclodextrin (HP-β-CD).

Methods. We used high sensitivity differential scanning calorimetry (HSDSC) in combination with a central composite design (CCD). As indicators of protein thermal stability, the measured responses were the unfolding transition temperature (Tm), the onset temperature of the denaturation (T0), and the extrapolated onset temperature (To,e).

Results. A highly significant (F probability <0.001) statistical model resulted from analysis of the data. The largest effect was due to pH (over the range 3.2-7.2), and the pH value that maximized Tm was 4.8. Several minor but significant effects were detected that were useful for mechanistic understanding. In particular, the effects of protein concentration and cyclodextrin concentration on Tm and To,e were found to be pH-dependent. This was indicative of the partially hydrophilic nature of protein-protein interactions and protein-cyclodextrin interactions, respectively.

Conclusions. Response surface methodology (RSM) proved efficient for the modeling and optimization of lysozyme thermal stability as well as for the physical understanding of the protein-sugar-cyclodextrin system in aqueous solution.

protein stabilizationhigh-sensitivity differential scanning calorimetrycentral composite design

Copyright information

© Plenum Publishing Corporation 1999

Authors and Affiliations

  • Sébastien Branchu
    • 1
  • Robert T. Forbes
    • 1
  • Peter York
    • 1
  • Håkan Nyqvist
    • 2
  1. 1.Drug Delivery Group, Postgraduate Studies in Pharmaceutical Technology, The School of PharmacyUniversity of BradfordBradfordUK
  2. 2.Licensing DepartmentAstra Arcus ABSödertäljeSweden