Background and Objectives: Formetanate hydrochloride is an effective insecticide and acaricide, which is frequently used in citrus gardens. Given its high toxicity, it is necessary to evaluate effects of this insecticide on living factors. In this study, we investigate effects of formetanate hydrochloride on human hemoglobin using spectroscopic analysis and molecular docking modeling. Methods: Effects of formetanate hydrochloride on human hemoglobin were investigated using molecular docking technique and thermodynamic and spectrophotometric methods such as normal and synchronous fluorescence spectroscopy, UV-Vis absorption spectroscopy and circular dichroism. Results: Decrement of the soret band without any significant shift proved that the insecticide could penetrate into the heme pocket and directly interact with the porphyrin ring. Results obtained from the intrinsic fluorescence spectroscopy and heme degradation study at 321 and 460 nm indicated that formetanate hydrochloride could potentially destroy the heme prosthetic group and eliminate its filtering effect, which results in fluorescence enhancement. The fluorescence study demonstrated a competitive behavior between hydrophobic probe ANS and formetanate hydrochloride for binding to the hemoglobin hydrophobic patches. Results of circular dichroism and synchronous fluorescence spectroscopy confirmed the structural change and polarity alteration around aromatic fluorophores. Conclusion: Our findings elucidate that the binding of formetanate hydrochloride to hemoglobin through the heme pocket is energetically much more favorable compared to binding to the hydrophobic central cavity. In addition, formetanate hydrochloride can cause conformational changes in the human hemoglobin and the competitive behavior of this insecticide for the oxygen site can subsequently reduce the oxy form of hemoglobin.
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