Wound healing plants as targets for identification of pectins with immunomodulatory effects
Pr Drissa Diallo, Department of Traditional Medicine Bamako Mali
Wounds in Traditional Medicine
zIn TM, a wound is defined and classified according to:
– Its origin,
– Localization,
– and whether the wound is new or old
zThe treatment differs according to these conditions
Wounds in Traditional Medicine
z Classification: – New wound: injury with a sharp object
– Chronic wound: “Kélébé” • More than one year old
• provoked by a curse in TM concept
z Treatment:
– New wound: Plants having haemostatic properties
– Chronic wounds: plants having immunomodulatory and antiinflammatory properties
Identification of wound healing plants
z Ethnopharmacological survey (Diallo et al., 2002):
– 123 plants species identified
– Belonging to 50 botanical family z Wound healing plants studied
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Vernonia kotschyana
Trichilia emetica
Opilia celtidifolia
Biophytum petersianum Cola cordifolia
Glinus oppositifolius
Etc
Wound healing remedies
z Plants part used:
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V. kotschyana: root
E. africana: root, stem bark
T. emetica: leaves
O. celtidifolia: leaves
B. petersianum: leaves C. cordifolia: leaves, stem bark
C. tinctorium: root
z Mode of preparation:
– Decoction of plant material (wash the wound)
– Powdered plant material applied as ointment
– Carbonized plant material applied
Pectins from wound healing plants
z Extraction processes
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Removal of low molecular weight compounds
Water extraction at 50 and 100°C
Centrifugation of extracts
Dialysis z Fractionation
– Gel filtration (Biogel, Sephacryl)
– Ion exchange chromatography (DEAE Sepharose) Pectins from wound healing plants
z Determination of monosaccharides contents: – Methanolysis
– Gas Chromatography
z Structure determination:
– Reduction, methylation, hydrolysis, acetylation – GC‐MS – NMR
z Biological activity
– Immunomodulatory effects: stimulation of one or several components of the immune system
Pectins from V. Kotschyana
zStructures: (Cecilie et al., 2004, 2005)
– Neutral fractions contained inulin, β (1‐2) linked fructosyl units with glucosyl unit in the reducing end (fructose polymer)
– Acidic fractions contained:
• Arabinogalactan type II structures (AG‐II)
• Linked to pectic polysaccharides (Pectins)
Pectins from V. Kotschyana
zBiological activities
– Inulin: no activity
– Pectin: • complement fixation activity
• Mitogenic activity
• Both at a dose‐dependant manner Pectins from V. Kotschyana
zComplement fixation activities
Pectins from V. Kotschyana
zMitogenic activity on Spleen Cells
Vk50A2 Vk100A2 Positive controls
Pectins from V. Kotschyana
z B cell proliferation in Spleen Cell suspension
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Vk50A2 Vk100A2 BR2IIc
Pectins from O. celtidifolia
z Primary structure of polysaccharides of water 50 degree extract:
– Arabinogalactan Type II – Arabinogalactan Type I
– Pectic type polysaccharides
z Refine structure elucidation is ongoing…
Pectins from O. celtidifolia
zComplement fixing activity of A1 A2 and sub fractions
ICH50 Values (μg/ml)
ICH50
PMII
A1
A1d1
A1d2
A2
A2d1
A2d2
70.5
0.9
0.5
4.4
0.5
0.2
1.4
Pectins from O. celtidifolia
z T cells proliferation activity of A1 A2 and sub fractions
Medium
2.5%
LPS
25.6%
T cells proliferation activity
Oc50A1
13.9%
Oc50A2
24%
Pectins from O. celtidifolia
z B cells proliferation activity of A1 A2 and sub fractions
Moderate B cells proliferation activity approx. 20% of proliferation with A1 and A2 at 100µg/ml
Pectins from O. celtidifolia
z Macrophage stimulations activity to release NO of A1 A2 and sub fractions
Oc50A1
Oc50A2
--A1d1
--A1d2
--A2d1
--A2d2
Nitrite (µM)
LPS
A1 and A2 (100μg/ml) induced the release of ≈ 7 μM of NO
LPS at 200 ηg/ml induce ≈ 8 μM of NO
Pectins from T. emetica
zPrimary structure: – The acidic 1 of the 50 and the 100°C water extracts seemed to be an arabinogalactan
– The acidic 2 and acidic 3 of the 100°C were pectins with a galacturonan backbone (RGI)
Pectins from T. emetica
zComplement fixing ability:
– The 50 °C water extract showed an effect on the complement system with an ICH50 of 45 µg/ml. – The 100 °C water extract had complement fixing activity with an ICH50 of 35 µg/ml
Pectins from T. emetica
z Complement fixing activity of fractions
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Samples ICH50 (µg/ml) Orig. Hydr.
Te50 crude extract 45 Te 50 acidic 1
31,25 137
Te 100 crude extract 35 Te 100 neutral 52 Te 100 acidic 1 196 Te 100 acidic 2 155 500
Te 100 acidic 3 62,5 233
Te 100 acidic 4 15 90
PM II 33,2
Pectins from B. petersianum
zComplement fixing activity:
– Water extract 100 degree (Bp100) Ic50 < 7µg/ml
– Gel filtration of Bp100 gave Bp100III Ic50 < 9µg/ml
– Enzyme degradation of Bp1003 gave Bp100III.1 Ic50 < 2µg/ml, Bp100III.2 Ic50 = 1.53 µg/m
Pectins from B. petersianum
z Structural studies:
BP 100 polysaccharides are of:
– Pectic type with features of rhamnogalacturonan type I
– With arabinogalactan type II side chains
– and terminal xylose units linked to the some galacturonic acid residues Pectins from G. oppositifolius
z Structure of GoA1 :
– obtained after ion excnange chromatography of 50°C water extract of aerial part of G. oppositifolius
– contains an RGI type polysaccharide – with complex AGI and AGII side chains
– The side chains play a key role in the immunomodulating activities
Pectins from G. oppositifolius
z Biological activities GoA1
– Complement fixing activity: IC50 =34 µg/ml
– 44% of B cells proliferation at 500µg/ml
– Marked up‐regulation (100 fold) in mRNA for the proinflammatory cytokine IL‐1β in rat macrophage cell line R2‐MΦ
– Potent intestinal immune system modulating activity by stimulation of Peyer´s patch
Concluding remark
zThe selected wound healing plants all have immunomodulatory activity
zWound healing is then an indication for choosing plant having immunomodulatory properties
zPolysaccharides are amongst other one of the compounds responsible of the activities
Concluding remark
z The pectin structure is the common feature among the polysaccharide that have immunomodulatory activities
z The active site seem to be located in the heavily branched part of the molecules
z The activities are seen at low concentration which indicated that small dosages of traditional remedies are enough to cure patients. References
Diallo, et al., (2001) Journal of Ethnopharmacology, 74, 159-171.
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Togola, et al., (2008). Journal of Ethnopharmacology ;Volum 115. s. 423431, UiO
z Togola, et al., (2008) Carbohydrate Polymers ;Volum 73. s. 280-288, UiO
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