Investigating the binding interaction of azur A with hyaluronic acid via spectrophotometry and its analytical application.

The interaction between azur A (AA) and hyaluronic acid (HA) at AA concentrations from 3.430 x 10(-5) to 8.575 x 10(-5) M and sodium chloride concentrations from 0 to 0.01 M was investigated spectrophotometrically at 620 nm at temperatures from 0 to 50 degrees C. AA was shown to be a useful spectroscopic probe for detecting carboxyl groups in HA macromolecules. The interaction between AA and HA was temperature sensitive and little AA-HA interaction was observed at temperatures higher than 30 degrees C. The interaction of HA with AA was seen to be electrostatic in nature. The maximum binding number decreased with decreasing NaCl concentration, and the absorbance sensitivity decreased with increasing NaCl concentration in aqueous solution. Self-interference from the AA in the AA-HA interaction caused an overestimate of the molar mass of hyaluronic acid. An improved method was proposed to estimate the molar mass of HA, and a molar mass of 1.219 x 10(6) Da was obtained with this improved method for HA.