Optimal conditions and sample storage for the determination of H2O2 in marine waters by the scopoletin-horseradish peroxidase fluorometric method

The conditions presently in use for the fluorometric determination of H₂O₂ in marine waters, by reacting H₂O₂ with scopoletin in the presence of horseradish peroxidase (HRP) and measuring the quenching of the fluorescence intensity of scopoletin, are not the optimal conditions. Under the optimized conditions of a pH of 8.5–9.5, an excitation wavelength of 390 nm and an emission wavelength of 460, the sensitivity of the method can be increased significantly, by up to more than a factor of 3 and the variations in the sensitivity from sample to sample can be significantly reduced. Furthermore, the samples need not be analyzed immediately after sample collection as presently prescribed. After scopoletin and HRP have been added to a sample immediately after sample collection, the sample may be stored at room temperature in the dark for up to four days before the quenched fluorescence intensity of scopoletin is read.