Immobilization of Carbonic Anhydrase in Glass Micropipettes and Glass Fiber Filters for Flow-Through Reactor...

There are various ways of immobilizing carbonic anhydrase (CA) on solid materials. One of the final aims is to apply immobilized CA for the catalytic hydration of carbon dioxide (CO₂) as a first step in the conversion of gaseous CO₂ into solid products. The immobilization method investigated allows a straightforward, stable, and quantifiable immobilization of bovine erythrocyte carbonic anhydrase (BCA) on silicate surfaces. The method is based on the use of a water-soluble, polycationic second-generation dendronized polymer with on average 1000 repeating units, abbreviated as de-PG2₁₀₀₀. Several copies of BCA were first covalently linked to de-PG2₁₀₀₀ through stable bisaryl hydrazone (BAH) bonds. Then, the de-PG2₁₀₀₀-BAH-BCA conjugates obtained were adsorbed noncovalently either on microscopy glass coverslips, inside glass micropipettes, or in porous glass fiber filters. The apparent density of the immobilized BCA on the glass surfaces was about 8-10 pmol/cm². In all three cases, the immobilized enzyme was highly active and stable when tested with p-nitrophenyl acetate as a model enzyme substrate at room temperature. The micropipettes and the glass fiber filters were applied as flow-through systems for continuous operation at room temperature. In the case of the glass fiber filters, the filters were placed inside a homemade flow-through filter holder which allows flow-through runs with more than one filter connected in series. This offers the opportunity of increasing the substrate conversion by increasing the number of BCA-containing filters.