TY - JOUR
T1 - Plasmodium berghei
T2 - High pressure liquid chromatographic analysis of nucleotides from erythrocyte-free malarial parasites
AU - Van Dyke, Knox
AU - Wilson, Mark
AU - Trush, Michael
AU - Taylor, Michael
AU - Stealey, Patricia
N1 - Funding Information:
This work was supported financially by the World Health Organization. We would particularly like to extend thanks to Dr. W. Wernsdorfer for his continuing support. Partial support was provided by U.S. National Institutes of Health Grant No. GM 07039.
Copyright:
Copyright 2014 Elsevier B.V., All rights reserved.
PY - 1977/8
Y1 - 1977/8
N2 - Isolation and analysis of the nucleotides from the malarial parasite, Plasmodium berghei, are described. This was accomplished by reducing contaminating leukocytes and platelets by settling in 6% dextran-Krebs glucose. Contaminating red cells (either parasitized or nonparasitized) were lysed by a technique using a dilute saponin solution in a Krebs-glucose medium. The parasites were collected by centrifugation and destroyed by addition of 10% trichloroacetic acid. The acid was extracted with Alamine-Freon TF and the nucleotides analyzed using high pressure liquid chromatography with microparticle anion exchange columns. The following compounds were separated and quantitated: ATP, GTP, CTP, UTP, ADP, GDP, CDP, UDP, AMP, GMP, CMP, UMP, and NAD. In nanomoles of nucleoside triphosphate isolated, the order of highest to lowest concentration was: ATP > UTP > GTP > CTP, while the order of diphosphates was: ADP > CDP > UDP > GDP, and the monophosphates: NAD > AMP > CMP > UMP > GMP. The order in amount of nucleotides present was: nucleoside triphosphates > nucleoside monophosphates > nucleoside diphosphates. The statistical analysis of the nucleoside triphosphates showed little deviation from sample to sample. A comparison of the sample from the host mouse red blood cell shows significant concentrations only of ATP, ADP, and NAD, indicating that contamination from host blood cells was unlikely.
AB - Isolation and analysis of the nucleotides from the malarial parasite, Plasmodium berghei, are described. This was accomplished by reducing contaminating leukocytes and platelets by settling in 6% dextran-Krebs glucose. Contaminating red cells (either parasitized or nonparasitized) were lysed by a technique using a dilute saponin solution in a Krebs-glucose medium. The parasites were collected by centrifugation and destroyed by addition of 10% trichloroacetic acid. The acid was extracted with Alamine-Freon TF and the nucleotides analyzed using high pressure liquid chromatography with microparticle anion exchange columns. The following compounds were separated and quantitated: ATP, GTP, CTP, UTP, ADP, GDP, CDP, UDP, AMP, GMP, CMP, UMP, and NAD. In nanomoles of nucleoside triphosphate isolated, the order of highest to lowest concentration was: ATP > UTP > GTP > CTP, while the order of diphosphates was: ADP > CDP > UDP > GDP, and the monophosphates: NAD > AMP > CMP > UMP > GMP. The order in amount of nucleotides present was: nucleoside triphosphates > nucleoside monophosphates > nucleoside diphosphates. The statistical analysis of the nucleoside triphosphates showed little deviation from sample to sample. A comparison of the sample from the host mouse red blood cell shows significant concentrations only of ATP, ADP, and NAD, indicating that contamination from host blood cells was unlikely.
KW - High pressure liquid chromatography
KW - Malaria
KW - Mouse
KW - Nucleotides
KW - Parasitic protozoa
KW - Plasmodium berghei
KW - Purine/pyrimidine metabolism
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U2 - 10.1016/0014-4894(77)90084-4
DO - 10.1016/0014-4894(77)90084-4
M3 - Article
C2 - 195832
AN - SCOPUS:0017375607
SN - 0014-4894
VL - 42
SP - 274
EP - 281
JO - Experimental Parasitology
JF - Experimental Parasitology
IS - 2
ER -