THE EFFECT OF PH ON CHEMILUMINESCENCE OF DIFFERENT PROBES EXPOSED TO SUPEROXIDE AND SINGLET OXYGEN GENERATORS

The compromised optima for high intensity chemiluminescence (CL), usin g superoxide generators, were all above pH 9.0 for the CL probes lumin ol and lucigenin. With luminol the optima were at pH 9.0 and 9.4 for t he generators KO2 and hypoxanthine/xanthine oxidase (HX/XO), respectiv ely. Lucigenin, with the same generators, produced optima at pH 9.5 an d 10.0, respectively. The probe methyl-Cypridina-luciferin analogue (M CLA) produced optima closer to neutral pH, which is preferred for phys iological assessments. MCLA had optima at pH 6.0, 8.7 and 9.5 with KO2 and with HX/XO optima at pH 4.8, 6.0, 7.0 and 8.7. When CL was assess ed at physiological pH, MCLA observed superoxide radicals with a sensi tivity of 100- and 330-fold more than luminol or luicigenin respective ly. For singlet oxygen, the sensitivity of MCLA at this pH was 45- and 5465-fold more than for the said probes respectively. H2O2 did not el icit CL between pH 4 and 9.5 with any of the probes and did not influe nce the production of superoxide or singlet oxygen when co-assessed. T herefore CL could only be obtained when enzymes were used as converter s. The optima for the enzyme-conversion system horseradish peroxidase (HRP)/H2O2, and luminol, were at pH 8.0 and 9.2. Lucigenin and HRP/H2O 2 also had a biphasic CL profile with optima at pH 7.4 and 9.6. MCLA a nd HRP/H2O2 had five optima, with the major ones at pH 6.1 and beyond 10. The optima for the myeloperoxidase/H2O system were at 8.6 and beyo nd 10.0 when luminol and 0.15 mol/L NaBr were used. (C) 1997 John Wile y & Sons, Ltd.