Abstract
To evaluate the state of ferriprotoporphyrin IX (FP) in malaria pigment, mouse erythrocytes infected with Plasmodium berghei NYU-2 parasites were lysed by hypotonic shock, and hemoglobin and other soluble material were removed by extensive washing. The amount of FP recovered in the insoluble pellet was 2.1 mumol/ml of packed infected erythrocytes, of which approximately 1% was attributable to hemoglobin contamination. This crude preparation then was digested with a nonspecific protease from Streptomyces griseus and extracted with chloroform/methanol. The residue of insoluble dark brown material had the spectral and solubility properties characteristic of the FP of malaria pigment, and various different preparations contained from 82 to 99% of FP by weight. By elemental analysis, highly purified preparations contained no chlorine and had an oxygen content consistent with 1 mol of hydroxyl/mol of FP (oxygen content: calculated, 12.6%; found, 12.5%). In comparison to hematin purchased from Sigma, which had a measured oxygen content of 14.7%, the low oxygen form of hematin purified from malaria pigment was remarkably less soluble in ethanol, 3% sodium bicarbonate, and chloroform.
Highlights
To evaluate the state of ferriprotoporphyrinIX (FP) the putative role of FP in the mode of action of chloroquine, in malaria pigment, mouse erythrocytes infected with we proposed that the toxic form of FP, which binds chloro
The purpose of found, 12.5%).Incomparison to hematin purchased this paper is to demonstrate that FPin fact is sequestered in from Sigma, which had a measured oxygen content of malaria pigment as a peculiarly insoluble aggregate of hema
Theextent of contamination of malaria pigment with hemoglobin was evaluated by using [14C]chloroquineasa reporter molecule to detect FP released by proteolysis
Summary
The toxic form to accumulate to a critical concentration would be amount ofFP recovered in the insoluble pellet was 2.1 susceptible to chloroquine whereas parasites that keep the gmol/ml ofpacked infected erythrocytes, of which approximately 1%was attributable to hemoglobin contamination This crude preparation was digested with a nonspecific protease from Streptomyces griseus and extracted with chloroform/methanol.The residue of insoluble dark brown material had the spectral and solubility propertiescharacteristic of the FP of malaria pigment, and various different preparations contained toxic form of FP below the critical concentration would be resistant to chloroquine. From this perspective, it seems important to describe the stateof FP in malaria pigment which renders it nontoxic and unable to bind chloroquine. Malaria pigment was remarkably less soluble in ethanol, 3%sodium bicarbonate, and chloroform
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