caa a22 4500 445312742 CHVBK 20180317142635.0 cr unu---uuuuu 170323e20110301xx s 000 0 eng 10.1007/s11095-010-0300-x doi (NATIONALLICENCE)springer-10.1007/s11095-010-0300-x The Use of Disaccharides in Inhibiting Enzymatic Activity Loss and Secondary Structure Changes in Freeze-Dried β-Galactosidase during Storage [Elektronische Daten] [Ville Heljo, Kirsi Jouppila, Timo Hatanpää, Anne Juppo] ABSTRACT: Purpose: The purpose of this study is to show how disaccharides differ in their ability to protect lyophilized β-galactosidase from enzymatic activity loss and secondary structure changes during storage. Methods: β-galactosidase was lyophilized with trehalose, sucrose, cellobiose or melibiose at 2:1, 20:1 and 40:1 excipient/protein weight ratios, and stored up to 90days at 45°C. Protein enzymatic activity was studied using o-nitrophenyl-β-d-galactopyranoside cleavage test, and its secondary structure in lyophilizates analyzed using Fourier transform infrared spectroscopy. The crystallization tendencies, glass transition temperatures and water contents of lyophilizates were evaluated using x-ray powder diffractometry, differential scanning calorimetry and thermogravimetry, respectively. Results: The enzymatic activity of β-galactosidase decreased more slowly in lyophilizates containing trehalose or melibiose at 2:1 excipient/protein weight ratio when compared to those containing sucrose or cellobiose. Similar behavior was observed when analyzing the protein's secondary structure in lyophilizates. In 20:1 and 40:1 excipient/protein weight ratio lyophilizates the decrease of enzymatic activity was less dependent on the excipient, but activity was always amongst the highest in melibiose lyophilizates. Conclusions: Melibiose was shown to be effective in protecting lyophilized β-galactosidase during storage. The protein secondary structure was shown to change at comparable rate in lyophilizates as its enzymatic activity after rehydration. Springer Science+Business Media, LLC, 2010 β-galactosidase nationallicence disaccharides nationallicence lyophilization nationallicence protein secondary structure nationallicence storage stability nationallicence Heljo Ville Faculty of Pharmacy, Industrial Pharmacy, University of Helsinki, Helsinki, Finland aut Jouppila Kirsi Department of Food Technology, Faculty of Agriculture and Forestry, University of Helsinki, Helsinki, Finland aut Hatanpää Timo Department of Chemistry, Laboratory of Inorganic Chemistry, University of Helsinki, Helsinki, Finland aut Juppo Anne Faculty of Pharmacy, Industrial Pharmacy, University of Helsinki, Helsinki, Finland aut Pharmaceutical Research Springer US; http://www.springer-ny.com 28/3(2011-03-01), 540-552 0724-8741 28:3<540 2011 28 11095 https://doi.org/10.1007/s11095-010-0300-x text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s11095-010-0300-x text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Heljo Ville Faculty of Pharmacy, Industrial Pharmacy, University of Helsinki, Helsinki, Finland aut NATIONALLICENCE 700 1- Jouppila Kirsi Department of Food Technology, Faculty of Agriculture and Forestry, University of Helsinki, Helsinki, Finland aut NATIONALLICENCE 700 1- Hatanpää Timo Department of Chemistry, Laboratory of Inorganic Chemistry, University of Helsinki, Helsinki, Finland aut NATIONALLICENCE 700 1- Juppo Anne Faculty of Pharmacy, Industrial Pharmacy, University of Helsinki, Helsinki, Finland aut NATIONALLICENCE 773 0- Pharmaceutical Research Springer US; http://www.springer-ny.com 28/3(2011-03-01), 540-552 0724-8741 28:3<540 2011 28 11095 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer