It is essential to properly equilibrate carriers with a buffer solution before presenting enzymes to the surface. The buffer should maintain a pH that promotes enzyme solubility throughout the immobilization interaction and in the following days, otherwise the final biocatalyst may not perform well. Since the ionic capacity of many enzyme carriers, especially anion exchange carriers, is higher than normal biological buffer systems it is important to equilibrate the carriers to the desired pH before adding enzymes to the system. At large scale this can be accomplished by mixing the carrier with dilute buffer while dosing caustic or acid into the tank with a pump. Here in the lab we use small quantities of carrier, so we opt to add a small volume of buffer, wait until the pH stops changing, then remove continue with immobilization.

    Equal amounts of carriers (adjusted for dry solids content) of forty carriers and empirically determined the number of buffer washes necessary to bring each one into a relatively neutral pH range of 6.5 – 7.5. This experiment (Figure 1) illustrates one of the easiest mistakes to make when preparing a carrier which is checking the pH too early. If the buffer pH is measured on the same day it is mixed with the carrier it might appear similar to the starting pH and therefore ready to accept enzymes. However, overnight the pH will continue to change. Using such a carrier without longer equilibration could interfere with enzyme immobilization and even the starting activity of a sample. Sixteen of the forty carriers tested required an overnight equilibration, and most of those sixteen ended up needing multiple days to equilibrate. In the future, equilibration will be sped up by using larger buffer:carrier ratios to reduce the number of washes needed.