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[Conditional inhibition of hemozoin formation by chloroquine in vitro]
Jun Sun1, Bo Chen, Yan-Yu Long
1Institute for Infectious Diseases and Vaccine Development, School of Medicine, Tongji University, Shanghai 200092, China. swksj@tongji.edu.cn
Objective:
To study the characteristics of inhibition on hemozoin formation by chloroquine under in vitro condition.
Methods:
Under different concentrations (0.5-2 mol/L) of sodium acetate (NaAc) and at the pH range of 4.0-5.0, chloroquine was tested for inhibition of beta-hematin (hemozion) formation by using the HPIA (heme polymerization inhibitory activity) assay. The morphology of beta-hematin crystals was determined by light microscopy. Ultraviolet spectrophotometry was employed to measure beta-hematin content, and the size of beta-hematin crystal was analyzed by X-ray diffraction (XRD).
Results:
Chloroquine exhibited varied effect on beta-hematin formation, depending on pH value and Na+ concentration. When the NaAc concentration increased from 0.5 mol/L (pH 4.2) to 2 mol/L (pH 4.8), the chloroquine inhibitory effect also increased. Results suggested that there exists a threshold pH, below which the beta-hematin formation escalates and chloroquine inhibition declines, and at or above which chloroquine exerts a stronger inhibitory effect on beta-hematin formation. With the increase of pH from 4.4 to 4.8, the crystallinity and the size of crystal changed from 6.93% and 357 angstrom to 6.32% and 264 angstrom, respectively. When pH reached to 5, no more beta-hematin formed. Chloroquine could reduce the crystallinity and crystal size of beta-hematin at same pH value. Morphology analysis on the samples was consistent with the above results.
Conclusion:
Chloroquine inhibits hemozoin formation only when the pH value is at or above threshold pH.
