A fluorescence quenching method was used to determine the equilibrium binding constants for the association of acridine, acridine orange, and acridine yellow G to humic acid. The fluorescence of each polycyclic aromatic nitrogen heterocycle (PANH) was monitored as aliquots of humic acid were added, and a Stern-Volmer plot was produced in which the slope is the equilibrium constant of the binding reaction. The quenching experiments were performed at temperatures of 30, 35, 40, and 45 °C. A van't Hoff plot generated from the equilibrium binding constants as a function of temperature for a given PANH resulted in a linear plot. Calculation of the ?<i>H</i><sub>binding</sub>, ?<i>G</i><sub>binding</sub>, and ?<i>S</i><sub>binding</sub> for each PANH leads to the conclusion that the equilibrium binding constant, and ?<i>G</i><sub>binding</sub>, may be predictors of bioavailability. The other thermodynamic quantities, ?<i>H</i><sub>binding</sub> and ?S<sub>binding</sub>, are helpful in understanding the relative binding of the compounds. For example, acridine yellow G appears to be the least bioavailable of the three PANHs studied because of its strong ?<i>H</i><sub>binding</sub> = ?29.8 kJ/mol, which leads to ?<i>G</i><sub>binding</sub> = ?0.71 kJ/mol. While acridine orange and acridine have similar ?<i>H</i><sub>binding</sub> values, acridine orange is more likely to bind to humic acid because the ?<i>S</i><sub>binding</sub> for the process is less negative. Thermodynamic values and equilibrium binding constants for all three compounds are reported.
Vol. 8, Iss. 8 Virtual Journal for Biomedical Optics
Jake T. Herb and Bruce D. Anderson, "Investigating the Binding of Acridine, Acridine Orange, and Acridine Yellow G to Humic Acid Through Fluorescence Quenching," Appl. Spectrosc. 67, 752-757 (2013)