Chapter 5 Conclusion
To sum up, from this study we draw conclusions as follows:
Hemin, product of hemoglobin degradation in vacuole food of
parasite, interacted both with quinoline and artemsinin compounds. The major
advantage of water-propylene glycol and water-DMSO mixture is that hemin still
monomeric and, under these conditions, interpretation of results is not
complicated. Both quinoline and artemisinin drugs- hemin complexes exhibited
1:1 stoechiometry. It was found that water-propylene glycol is suitable for
hemin-quinolines interactions whereas aqueous DMSO solution is suitable
studying hemin-artemisnin interaction. For quinoline based drugs, the results
indicated that hemin complexed more strongly with quinidine than with
chloroquine and quinine, and the binding constants were pH-dependent. Because
of lack of quantitative data, about the bonding of hemin with endoperoxide
lactone based antimalarial drugs, we focused our investigation in it.
Artemisinin and derivates were studied by UV and HPLC/DAD/MS analysis for their
reactivity with hemin. The reported results showed that hemin and endoperoxide
lactone derived antimalarials slowly react to give rise to several
stereoisomers supramolecular adducts (three for artesunate, seven for
artemisinin and eight isomers for dihydroartemisinin) while many studies
certified that metal (III) porphyrins react poorly with those drugs. Generally,
hemin (FeIII) is firstly reduced to heme (FeII) in the
presence of reducing agent like glutathione (in high concentration in
erythrocytes)[Robert et al., 2002], secondly the resulting heme will react with
artemisinin drug. Thermodynamic data supported too our results and showed that
artesunate and dihydroartemisinin interacted more strongly with Fe (III) PPIX
that artemisinin did. Then, it must be considered too that Fe (III) PPIX should
be a potentiel target of artemisinin derived drugs.
It is already known that active endoperoxides react with
porphyrins while inactive ones do not, suggesting that this reaction may be
important in parasite. In the light of preview researches, we suggest that in
vivo, artemisinin and derivatives diffuses into the food vacuole, where its
reacts with newly formed monomere heme (released from digested hemoglobin) or
hemin to generate whether a covalent heme-artemisinin or whether a covalent
hemin adduct. This long-lived intermediate may ultimately participe, via
C4-centred alkylating radical, in the oxidative damage of membrane, which is
lethal for the malaria parasite. The full characterization of a covalent
artemisinin-hemin adduct is a key in the understanding of the mode of action of
this antimalarial drug, the lead molecule for the rational design of cheap and
highly efficient endoperoxide-containing molecules against the
chloroquine-resistant strains. Thus due to the high sensitivity, simplicity and
feasibility of the two analytical methods should be considered a rapid and
inexpensive approach in the search for new lead compounds having an
antimalarial activity similar to that of artemisinin. These methods can be
considered as widespread analysis techniques in the search of false
artemisinin, artesunate and dihydroartemisinin drugs on the market.
Recommandations:
Since DMSO solutions are a suitable medium of
hemin-artemisinin adduct formation, we recommend to produce this adduct in high
yield under optimal experimental conditions. Then, the artemisinin adduct
products will be separated by reverse HPLC, isolated and his different isomers
characterized for future parasitologic essay.
The structure of those isomers can be elucidated by R-X
diffraction, or by NMR spectroscopy. But because of paramagnetic character of
artemisinin-hemin adduct with Fe (III), some precautions will be taken, like
the demetallisation of those adducts.
As physiological medium is aqueous medium, it is suitable to
find the experimental optimal conditions in aqueous medium will permit both to
control the dimerization of hemin and take account of solubility of
antimalarial drugs. In the future, we suggest also:
-To study the artemisinin-drug interaction at food vacuole
pH of plasmodium in the used medium.
-To investigate the bonding of hemin with artemisinin based
drugs in presence of glutathione (reducing agent present in high concentration
in erythrocytes).
-To identify exactly the structure and the nature of the ART
290.
-To investigate by two-dimensional infrared correlation
spectroscopy the nature of interaction between artemisinin of drug and hemin.
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