Effects of types of sugar on the stabilization of protein in the dried state

Koreyoshi Imamura, Tomohiro Ogawa, Takaharu Sakiyama, Kazuhiro Nakanishi

Research output: Contribution to journalArticle

67 Citations (Scopus)

Abstract

The effects of various sugars on the structural stabilization of protein during freeze-drying were investigated. The degree of native structure of protein that was freeze-dried and rehumidified at constant relative humidities (RHs) was evaluated by measurement of the α-helix content by Fourier-transform infrared spectroscopy. Bovine serum albumin (BSA) and several types of sugars, including sucrose, trehalose, and dextrans, were used as a model protein and sugars, respectively. The glass transition temperature, Tg, for the amorphous sugar samples was measured by differential scanning calorimetry (DSC) to characterize the structural stability of sugars. The dependence of the α-helix content (Cα-helix) of BSA on the sugar content (Csugar) could, in most cases, be represented by a Langmuir-type equation: Cα-helix =K x (Cα-helixmax -Cα-helix0) x Csugar/(1 + K x csugar)+ Cα-helix0, where K is a constant, indicating the ability of amorphous sugar matrix to embed protein, and Cα-helix0 and Cα-helixmax indicate the α-helix content in the absence of sugar and saturating levels of sugar, respectively. The preservation effects of the sugars could be characterized by K and Cα-helixmax. Both K and Cα-helixmax values tended to be higher with decreasing Tg values for the amorphous sugar, probably because an amorphous sugar matrix with lower Tg values is structurally more flexible. The rehumidification of protein that was freeze-dried in the presence of sugar induced the refolding of protein structure, whereas the protein dried alone did not show any recovery of its native structure.

Original languageEnglish
Pages (from-to)266-274
Number of pages9
JournalJournal of Pharmaceutical Sciences
Volume92
Issue number2
DOIs
Publication statusPublished - Feb 1 2003

Keywords

  • Amorphous sugar
  • Freeze-dried protein
  • Glass transition temperature
  • Secondary structure
  • Stabilization

ASJC Scopus subject areas

  • Pharmaceutical Science

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