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In Vitro Antiplasmodial Activity of Cochlospermum planchonii and its
Phytomedicines
A. Lamien-Meda1, K. König1, M. Mischka1, R. Meda2, M. Compaoré3, M. Kiendrebeogo3, H.P. Fuehrer4, H. Nödl5, J. Novak1
1 Institute of Animal Nutrition and Functional Plant Compounds, University of Veterinary Medicine Vienna, Austria 2 UFR/ST, Université Polytechnique de Bobo-Dioulasso, Burkina Faso
3 Laboratory of Biochemistry and Chemistry (LABIOCA), UFR/SVT, University of Ouagadougou, Burkina Faso 4 Institute of Parasitology, University of Veterinary Medicine Vienna, Veterinaerplatz 1, Austria
5 Institute of Specific Prophylaxis and Tropical Medicine, Medical University of Vienna, Vienna, Austria
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Research background • MEAMP Project • Focus on the endangered
antimalarial medicinal plants from Burkina Faso: − Malaria is leading cause of infantile death − Malaria constitutes a public health
preoccupation for the whole population
Source: WHO Malaria Report 2011/2013 http://events.ucsf.edu/downloads/malaria/map.html
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Recent Antimalarial Natural Products from Plants
1993-2003 127 alkaloids 18 quassinoids 23 sesquiterpenes 27 triterpenoids 21 flavonoids/xanthones 9 quinones 25 miscellaneous compounds
Saxena et al. (2003)
2008 31 indole alkaloids (with high antiplasmodial activity)
Frederich et al. (2008)
1998-2008 266 antiplasmodial natural products: Alkaloids, terpenes, quassinoids, flavonoids, limonoids, chalcones, peptides, xanthones, quinones, coumarins, miscellaneous compounds
Kaur et al. (2009)
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Cochlospermum planchonii Hook. f. ex Planch. (Bixaceae)
• Rhizome traditionally used for the treatment of fevers, jaundice and malaria in West African countries
• The phytomedicine N’Dribala (rhizome of C. planchonii):
•
• The active compounds of the plant still remain unknown
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Why C. planchonii? Survey results (APPEAR project-75)
Cochlospermum planchonii Hook. f. ex Planch. (Bixaceae)
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Phytomedicines
C.planchonii
N’dribala
Saye
C.Planchonii root
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Extraction, Partition, Chromatography
MeOH Ethanol Aceton CH2Cl2 Water
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Continuous P. falciparum Culture
• Sterile hood (laminar flow); incubator, 37°C; gas mixture (5% O2, 5% CO2, 90% N2); liquid nitrogen tank; Microscope with oil immersion objective
• Cell culture flasks
• RPMI 1640 Medium
• Erythrocyte concentrate, preferably 0-
• Human serum, preferably A+, AB+ or corresponding to RBCs
• Giemsa stain
• Parasite used for testing: chloroquine sensitive 3D7
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Development of Quality Control System
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C. planchonii HPLC (High Pressure Liquid Chromatography) Profile
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C. planchonii HPLC (High Pressure Liquid Chromatography) Profile
Cochloxanthine
Dihydrocochloxanthine
TLC:EtOAC/MeOH/H2O (77/13/10)
Anisaldehyde/visible
Fractions of Cochlospermum plant material from MPLC (Aceton/Hexane 3/7; Acetic acid 0.1%)
Dihydrocochloxanthine
Cochloxanthine
HPLC and structure elucidation by Nuclear Magnetic Resonance (NMR) spectroscopy
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C. planchonii HPLC (High Pressure Liquid Chromatography) Profile
Cochloxanthine
Dihydrocochloxanthine
TLC:EtOAC/MeOH/H2O (77/13/10)
Anisaldehyde/visible
Fractions of Cochlospermum plant material from MPLC (Aceton/Hexane 3/7; Acetic acid 0.1%)
Dihydrocochloxanthine
Cochloxanthine
HPLC and structure elucidation by Nuclear Magnetic Resonance (NMR) spectroscopy
Are the two compounds active against malaria?
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Cochloxanthine
Dihydrocochloxanthine
C. planchonii HPLC (High Pressure Liquid Chromatography) Profile
TLC:EtOAC/MeOH/H2O (77/13/10)
Anisaldehyde/visible
Fractions of Cochlospermum plant material from MPLC (Aceton/Hexane 3/7; Acetic acid 0.1%)
Dihydrocochloxanthine
Cochloxanthine
HPLC and structure elucidation by Nuclear Magnetic Resonance (NMR) spectroscopy
IC50 (µg/ml)
Cochloxanthine 6.3
Dihydrocochloxanthine 6.6 C. planchonii rhizome
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0
2000000
4000000
6000000
8000000
10000000
12000000
14000000
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29
MeOH: DH Cochloxanthine/Cochloxanthine
C. planchonii Quality control
C. planchonii
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0
2000000
4000000
6000000
8000000
10000000
12000000
14000000
16000000
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29
Ethanol: DH Cochloxanthine/Cochloxanthine
C. planchonii
C. planchonii Quality control
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0
2000000
4000000
6000000
8000000
10000000
12000000
14000000
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 19 20 21 23 24 25 26 27 28 29
Acetone: DH Cochloxanthine/ Cochloxanthine
C. planchonii
C. planchonii Quality control
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0
1000000
2000000
3000000
4000000
5000000
6000000
7000000
8000000
9000000
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29
CH2Cl2: DH Cochloxanthine/Cochloxanthine
C. planchonii
C. planchonii Quality control
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0
200000
400000
600000
800000
1000000
1200000
1400000
1600000
1800000
1 2 3 4 5 6 7 8 9 12 13 14 15 18 24 25 28 29
H2O: DH Cochloxanthine/Cochloxanthine
C. planchonii
C. planchonii Quality control
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Why the decoction is active?
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Cochlospermum: Antiplasmodial Activity C. planchonii
0 10 20 30 40 50 60
H2O H2O H2O MeOH EtOH CH2Cl2 Aceton
Saye N'dribala CP2
IC50 (µg/ml)
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Cochlospermum: Antiplasmodial Activity C. planchonii
0 10 20 30 40 50 60
H2O H2O H2O MeOH EtOH CH2Cl2 Aceton
Saye N'dribala CP2
IC50 (µg/ml)
MeOH EtOH CH2Cl2 Aceton
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Cochlospermum: Antiplasmodial Activity C. planchonii
MeOH EtOH CH2Cl2 Aceton
27
39
57
43
51 50
21
1 11 7 6 0
10
20
30
40
50
60
H2O H2O H2O MeOH EtOH CH2Cl2 Aceton DCM EAF DH cochlox.
cochlox.
Saye N'dribala CP2 CP
IC50 (µg/ml)
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Cochlospermum: Antiplasmodial Activity C. planchonii
MeOH EtOH CH2Cl2 Aceton
26,9
39,2
56,8
43,3
50,9 50,3
20,8
0,8 10,9 6,6 6,4 0,0
10,0
20,0
30,0
40,0
50,0
60,0
H2O H2O H2O MeOH EtOH CH2Cl2 Aceton DCM EAF DH cochlox.
cochlox.
Saye N'dribala CP2 CP
IC50 (µg/ml)
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Cochlospermum: Antiplasmodial Activity C. planchonii
0
20
40
60
80
100
120
H2O H2O H2O MeOH EtOH CH2Cl2 Aceton DCM EAF DH cochlox.
cochlox.
Saye N'dribala CP2 CP
IC50, IC90, IC99 (µg/ml)
IC50 IC90 IC99
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C. planchonii, N‘dribala, Saye
Cochloxanthine & Dihydrocochloxanthine
Marker Compounds for Quality Control
UV spectra
Cochloxanthine
Dihydrocochloxanthine
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Research Perspectives
Further investigation on the antiplasmodial activity of C. planchonii: Fractions and isolated compounds from active fractions
Investigation of the mode of action of the fractions on the parasite life stages.
Cytotoxity of the the active fractions and/ or compounds
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Dr. Zephirin Dakuyo
Acknowledgement: Austrian Partnership Programme in Higher Education and
Research for Development – APPEAR project-75 (2012-2014)
Aline Lamien Meda Johannes Novak
Students: Martina Mischka Anna Karnthaler Johannes Klinger
Karin König (BOKU) Associate Partner:
Hans Peter Fuehrer Thomas Pacher
Tamara Kolodziej Markus Bacher (BOKU)
Associate Partners: University of Pretoria (Prof. K. ELOFF), University of Graz (Prof. Dr. R. BAUER),
University of Lausanne (Dr. B. GRAZ), University of Oxford (Dr. M. WILLCOX),
ULB Belgium (Prof. P. DUEZ)
Prof. Harald Noedl
Martin Kiendrebeogo Moussa Compaoré
Roland Meda Student:
Orokia Traoré Issa Karama
Associate Partner: Odile G. Nacoulma
Jeanne Millogo Patrice Zerbo Lassina Sanou
Souleymane Sanou Marc Olivier
DI Elke Stinnig Programme Manager Appear
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Vielen Dank für Ihre Aufmerksamkeit