Flow-injection photometric biosensing of cobalt(II) ions based on an apoenzyme reactivation method

Flow-injection photometric microdetermination of cobalt(II) ions based on an apoenzyme reactivation method is proposed. Alkaline phosphatase immobilized on oxirane-acrylic beads (Eupergit-C) is packed into a small polymer column and then used as the recognition element for cobalt(II) ions. The biosensing system is assembled with the immobilized enzyme column and a flow-through cell housed in a double-beam spectrophotometer for monitoring the enzymatic activity. Cobalt(II) ions are photometrically determined in 1.0 to 200 mu M through activation of the immobilized metal-free alkaline phosphatase(apoenzyme). The activity is assessed by injecting a 0.1 mL of 0.5 mM p-nitrophenyl phosphate solution as the substrate and thereby the change in absorbance at 405 nm due to the formation of p-nitrophenol resulting from the enzymic hydrolysis is measured. The column is generated by pumping a 40 mM 2,6-pyridine dicarboxylate solution (pH 6.0) between successive samples.<>