Effects of hydroxylpropyl-β-cyclodextrin on in vitro insulin stability

Liefeng Zhang, Wenjie Zhu, Lingling Song, Yifan Wang, Hui Jiang, Suyun Xian, Yong Ren, Liefeng Zhang, Wenjie Zhu, Lingling Song, Yifan Wang, Hui Jiang, Suyun Xian, Yong Ren

Abstract

The objective of this study was to elucidate the effects of hydroxylpropyl-beta-cyclodextrin (HP-beta-CD) on the in vitro stability of insulin. It was found that HP-beta-CD had positive effects on the stability of insulin in acid and base and under high temperature conditions. Furthermore, use of HP-beta-CD could also increase the stability of disulfide bonds which are important to the conformation of insulin. Through (1)H-NMR experiments it was found that the protective effect of HP-beta-CD was due to complexation with insulin. The results suggest that the presence of HP-beta-CD could improve the stability of insulin in different environments.

Keywords: Protection; cyclodextrin; insulin; stability.

Figures

Figure 1.
Figure 1.
The remaining insulin in acid solution. ▴-▴ indicates the concentration change of insulin without HP-β-CD. ▪—▪ indicates the concentration change of insulin with the existence of HP-β-CD. n = 3, mean ± SD.
Figure 2.
Figure 2.
The remaining insulin in base solution. ▴-▴ indicates the concentration change of insulin without HP-β-CD. ▪—▪ indicates the concentration change of insulin with the existence of HP-β-CD. n = 3, mean ± SD.
Figure 3.
Figure 3.
The remaining insulin in a 55 °C water bath. ▴-▴ indicates the concentration change of insulin without HP-β-CD. ▪—▪ indicates the concentration change of insulin with the existence of HP-β-CD. N = 3, mean ± SD.
Figure 4.
Figure 4.
Effect of HP-β-CD on the stability of disulfide bonds. A and B are the PAGE results. The upper graph (A) was insulin without HP-β-CD, the lower graph (B) was insulin and HP-β-CD. Lanes 1-4 in (A) and (B) indicated insulin incubated with different GSH/GSSG (mM/mM) solutions with 0, 20/1, 40/1, 60/1, GSH/GSSG solution, respectively. C is the result of HPLC. ▪—▪ indicates the remaining insulin with HP-β-CD, ♦-♦ indicates the remaining insulin without HP-β-CD.
Figure 5.
Figure 5.
1H-NMR spectrum of aromatic region of insulin (4 mM) in the absence of HP-β-CD (A) and in the presence of HP-β-CD (120 mM) (B) in D2O containing 30% CD3COOD at room temperature.

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