The water-dependence of the catalytic activity of bilirubin oxidase suspensions in low-water systems

✍ Scribed by Mark J. Alston; Robert B. Freedman

• Publisher: John Wiley and Sons
• Year: 2002
• Tongue: English
• Weight: 122 KB
• Volume: 77
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/bit.10103

✦ Synopsis

Abstract

We investigated the enzymic activity of bilirubin oxidase when it is suspended as a lyophilized powder in a low‐water system. The enzyme required buffer salts and a source of water to show activity. This study investigated the complete range of water thermodynamic activity (a~w~) by combining the use of salt hydrates and two‐phase systems with concentrated solutes in the aqueous phase. When free water was added, activity reached a maximum at a defined water content, but this maximum increased with buffer content, suggesting that there was competition for water with the buffer salts from which the enzyme was lyophilized. Alternatively, a range of salt hydrates was used, each able to fix the water activity (a~w~) at a different value. By providing water to the organic solvent phase in this way, the dependency of enzyme activity upon a~w~ was investigated and shown to be independent of buffer concentration. However, the optimum a~w~ was uncertain because the available a~w~ range for salt hydrates is ≤0.90. Investigation of the remaining water activity range was made possible by using an a~w~ depressor (sorbitol) to lower the a~w~ of a two‐phase system. The optimum a~w~ for the bilirubin oxidase activity in this two‐phase system was a~w~ = 0.936, independent of buffer concentration. The study therefore confirmed the need to control the water `available' to low‐water systems and the dependence of enzyme activity on water thermodynamic activity (a~w~) not water content. © 2002 John Wiley & Sons, Inc. Biotechnol Bioeng 77: 651–657, 2002; DOI 10.1002/bit.10103