The method of Three-Dimensional Excitation Emission Matrix Fluorescence Spectrophotometer was used to identify the interaction mechanism between humic acid (HA) and antibiotics. The effects of antibiotic concentration, temperature, and pH on the bonding strength between HA and antibiotics were investigated. The results showed that with the increase of antibiotics concentration, the quenching effect on HA was enhanced. The quenching of HA by both oxytetracycline (OTC) and sulfadiazine (SD) is a single static quenching of the complexes, and the interaction forces are mainly a hydrogen bond and the van der Waals force. The quenching constant KsvOTC > KsvSD and the binding constant KbOTC > KbSD, indicates that HA has a more obvious quenching effect on OTC. The Kb of HA by OTC ranged from 3.223 × 10³ to 12.784 × 10³ L·mol−1 in the range of 298 K to 313 K, while the Kb of HA by SD ranged from 2.051 × 10³ to 5.533 × 10³ L·mol−1. With the increase of temperature, the quenching constant Ksv and binding constant Kb of both OTC and SD by HA gradually decrease, and the low temperature is beneficial to the interaction. The composite of OTC and HA is more stable than SD. Under neutral alkaline conditions, both OTC and SD had the strongest quenching effect on HA, and the resulting complex was the most stable. However, the Ksv and Kb of HA by OTC were greater than SD in the pH range of the experiment and the pH effect on quenching of HA by OTC (Ksv) was greater than that of SD.
Keywords: antibiotic; binding constant; fluorescence quenching; humic acid; interaction.