| Abstract: | Abstract An electrode method has been developed for the determination of flavin adenine dinucleotide (FAD) using a potentiometric gas sensor and commercially available enzyme preparations. The construction of the FAD-sensitive electrode is based on immobilizing alkaline phosphatase (E.C. 3.1.3.1) and adenosine deaminase (E.C. 3.5.4.4) on the sensing tip of an ammonia gas sensor. Alkaline phosphatase enzymatically catalyzes the hydrolysis of FAD to adenosine which is subsequently converted to ammonia by adenosine deaminase. The response of the dual-enzyme electrode is linear between 8 × 10?5 M and 4 × 10?3 M FAD with a slope of 43 mV/decade concentration at pH 8.5 and 37ddot]C. The optimum buffer system is 0.5 M diethanolamine, 1 × 10?3 M Tris-HCl and 1 × 10?3 M MgCl2. Electrodes constructed with enzymes immobilized by cross-linking with glutaraldehyde and bovine serum albumin showed longer life times than electrodes using enzymes entrapped by a dialysis membrane. The electrode is highly selective over riboflavin and flavin mononucleotide, but it does respond to other nucleotides. |
