Enzymatic method for measuring the absolute value of oxygen concentration

Area of Science:

• Biochemistry
• Analytical Chemistry

Background:

• Accurate dissolved oxygen measurement is critical in biological and chemical research.
• Existing methods may have limitations in precision or applicability to closed systems.

Purpose of the Study:

• To develop and validate a novel enzymatic assay for determining absolute oxygen concentration in aqueous solutions.
• To utilize 4-hydroxybenzoate 3-monooxygenase and glucose oxidase for sensitive oxygen detection.

Main Methods:

• Employing 4-hydroxybenzoate 3-monooxygenase for oxygen-dependent oxidation of 4-hydroxybenzoate and NADPH in a closed system.
• Utilizing glucose oxidase to consume atmospheric oxygen, enabling spectrophotometric measurement of NADPH consumption in an open system.
• Validating the assay by measuring oxygen in saturated and deoxygenated water samples.

Main Results:

• The enzymatic method accurately quantifies oxygen concentrations ranging from 8 µM to 1.1 mM.
• Oxygen-saturated water measured 1.10 mM and air-saturated water measured 0.23 mM at 25°C.
• Argon-bubbled water showed negligible oxygen levels, confirming assay specificity.

Conclusions:

• The described enzymatic method provides a reliable and sensitive approach for absolute oxygen concentration measurement.
• This assay is suitable for both open and closed systems and can verify anaerobicity.
• The dual-enzyme system offers a robust tool for dissolved oxygen analysis in diverse aqueous environments.