Structures, and immunomodulating and anti-ulcer activities of polysaccharides from Malian ...

88 Pages·2013·5.17 MB·English

Structures, and immunomodulating and
anti-ulcer activities of polysaccharides
from Malian medicinal plants
Ingvild Austarheim
Thesis submitted for the degree of Ph.D.
DepartmentofPharmacy
UniversityofOslo
November2012
© Ingvild Austarheim, 2013

Series of dissertations submitted to the
Faculty of Mathematics and Natural Sciences, University of Oslo
No. 1323

ISSN 1501-7710

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Cover: Inger Sandved Anfinsen.
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Contents
Acknowledgments v
Abstract vii
Listofpapers ix
Listofabbreviationsandsymbols xiii
1 Introduction 1
1.1 Plantpolysaccharides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
1.2 Pectinsasimmunomodulators . . . . . . . . . . . . . . . . . . . . . . . . . . 3
1.2.1 Complementsystem . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
1.2.2 Macrophagestimulation . . . . . . . . . . . . . . . . . . . . . . . . . 6
1.3 Internalandexternalwounds . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
1.3.1 GastriculcerandH.pyloriinfection . . . . . . . . . . . . . . . . . . . 7
1.3.2 Antiulceractivityininvivoexperimentalmodels . . . . . . . . . . . . 9
1.3.3 Anti-adhesiveactivitytowardsH.pylori . . . . . . . . . . . . . . . . . 10
1.3.4 Woundhealing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
1.4 Colacordifolia(Cav.)R.Br(Malvaceae) . . . . . . . . . . . . . . . . . . . . . 11
1.5 VernoniakotschyanaSch.Bip.exWalp(Asteraceae) . . . . . . . . . . . . . . 12
2 Aimsofthestudy 13
3 Summaryofpapers 15
4 Resultsanddiscussion 19
4.1 Ethnopharmacologicalresearch . . . . . . . . . . . . . . . . . . . . . . . . . . 19
4.2 Isolationandpurificationofthepecticpolymers . . . . . . . . . . . . . . . . . 20
4.2.1 StructuresofpectinsfromthebarkofC.cordifolia . . . . . . . . . . . 23
4.2.2 StructuresofpectinsfromleavesofC.cordifolia . . . . . . . . . . . . 30
4.2.3 ComparingpectinsfrombarkandleavesofC.cordifolia . . . . . . . . 31
iii
4.3 Immunomodulatingpropertiesandstructuralrequirements . . . . . . . . . . . 32
4.3.1 Complementfixationactivity . . . . . . . . . . . . . . . . . . . . . . . 32
4.3.2 Macrophageinduction . . . . . . . . . . . . . . . . . . . . . . . . . . 34
4.4 Anti-ulcer activity of polysaccharide rich extracts from V. kotschyana and C.
cordifolia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
4.5 GastricailmentsanddiagnosingH.pylori . . . . . . . . . . . . . . . . . . . . 37
4.6 Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37
References 45
Papers 47
PaperI.Chemicalandbiologicalcharacterizationofpectin-likepolysaccharidesfrom
thebarkoftheMalianmedicinaltreeColacordifolia . . . . . . . . . . . . . . 51
Paper II. Chemical and biological characterizations of pectins from Cola cordifolia
leaves . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63
PaperIII.Anti-ulcerpolysaccharidesfromColacordifoliabarkandleaves . . . . . . 87
PaperIV.Inulin-richfractionsfromVernoniakotschyanarootshaveanti-ulceractivity 97
PaperV.ChromatographicimmunoassaysforH.pyloridetection–aretheyreliable
enoughinMali(WestAfrica)? . . . . . . . . . . . . . . . . . . . . . . . . . . 103
iv
Acknowledgments
ThestudiespresentedinthisthesiswereperformedatDepartmentofPharmaceuticalChemistry,
School of Pharmacy, University of Oslo, from January 2008 to November 2012. Field work
and H. pylori testing of volunteers were performed in Mali in November 2008. In vivo anti-
ulcerexperimentswerecarriedoutinMali,October2010. Toxicityexperimentsonfibroblasts
were carried out in Israel, summer 2010. Financial support was provided by the following:
NorwegianPharmaceuticalsociety,EMBIOandUNIFORwhicharegratefullyacknowledged.
Foremost,IwouldliketoexpressmysinceregratitudetomyadvisorProfessorBeritSmestad
Paulsen, for the continuous support of my research, for her patience, motivation, enthusiasm,
andimmenseknowledge.Myco-supervisorDrissaDialloformakingmytripstoMalipossible
andpleasant.
IwanttothankallmycolleaguesattheDepartmentofPharmaceuticalChemistryforcreating
a nice and friendly atmosphere in addition to assistance in my work. Especially I would like
to thank Kari Inngjerdingen for all her kind support, my roomie, Anh Thu Pham, for sharing
goodandbadmomentsinourmutualgoal,andthecoolestprofessoremeritusalive,KarlEgil
Malterud,forsimplybeinghimself.
I would also like to thank my dear friends in Bamako, the twins Labass and Hamed Dahfalo
Koné, who treated me like family and my friends Adiaratou Togola, Mamady Mougare, and
N’GoloBallo.YoumademefeellikeBamakowasmysecondhome.
Finally,IwouldliketothankmybelovedpartnerOdinGramstad,forhisencouragementsand
motivation and help with computer problems and frustrations, and in general making my life
easytolive.IfitwasnotforyouIwouldprobablynotenjoymyPhDperiodasmuchasIdid.
Oslo,November2012
IngvildAustarheim
v

Abstract
ThemainpurposeofthisthesiswastoevaluatethepotentialofpectinsfromtheMalianmedici-
naltreeColacordifoliausedinthetreatmentofgastriculcerandwounds.Thisthesisisasmall
contribution to the ultimate goal of providing efficient none-toxic and inexpensive medicines
fortheMalianpopulation.
The Department of Traditional Medicine, our collaborating partner in Bamako, wants to pro-
mote the use of renewable plant parts to guarantee a sustainable supply of medicinal plants.
The structures and biological activities of the bark and leaf pectins were therefore compared
inordertomakerecommendationonplantpartsubstitutionasdebarkingcandamageoreven
killthetree. Wefoundthatthepectinsfromthebarkandleavesarestructurallyrelated. How-
ever,theleafpectinsaremorepolydisperseandheterogeneouscomparedtothepectinsfound
inbark. Pectinsfromthebarkweregenerallymoreactiveinthecomplementfixationtestand
the macrophage assay. Comparing the 50°C water extracts from bark and leaf in an experi-
mentalanti-ulcermodelshowedcomparableanddosedependentinhibitionofulcerformation.
However,aclinicaltrialisneededtoevaluatetheefficacyoftheplantparts.
PowderedrootsofVernoniakotschyanaarehighlyvaluedastheimprovedtraditionalmedicine
“Gastrosedal”. The anti-ulcer activity of the medicine has previously been attributed to the
presenceofsaponins. Inthisthesis,theanti-ulcerpotentialof50°Cand100°Cwaterextracts
depleted of saponins, but high in inulin, 98% and 83% respectively, were evaluated in an ex-
perimental mouse model. The tested dose corresponded to the recommended daily intake of
“Gastrosedal” and this dose showed a good inhibition of ulcer formation. We therefore con-
cludedthatinulincanalsoberesponsibleimportantfortheanti-ulceractivityof“Gastrosedal”.
InMali,gastricailmentsarerathercommonandcontributehighlytomorbidityinthecountry.
Inapreviousstudy,Helicobacterpyloriwasfoundtobepresentin95%ofMalianpatientswith
gastriculcer. Forfutureinvestigationsandclinicaltrials,itwasofinteresttofindareliableand
simplemethodforH.pyloridetection. Onestoolandoneserologicalbasedimmunochromato-
graphicmethodweretested,andtheresultsshowedthatthesensitivityofthesetestsistoolow
intheMalianpopulation. Thelowsensitivitywasprobablyduetostrainvariability,inaddition
tohighuseofanti-malariadrugs,whichmighteradicateorlowerthebio-burdenofH.pylori.
vii

List of papers
PaperI IngvildAustarheim,BjørnE.Christensen,IdaK.Hegna,BentO.Petersen,
JensO.Duus,RagnarBye,TerjeE.Michaelsen,DrissaDiallo,MaritInng-
jerdingen&BeritS.Paulsen
Chemicalandbiologicalcharacterizationofpectin-likepolysaccharidesfromthe
barkoftheMalianmedicinaltreeColacordifolia
CarbohydratePolymers89(2012),259–268.
PaperII Ingvild Austarheim, Bjørn E. Christensen, Christian Thöle, Drissa Diallo,
MaritInngjerdingen&BeritS.Paulsen
ChemicalandbiologicalcharacterizationsofpectinsfromColacordifolialeaves
Manuscript
PaperIII IngvildAustarheim,HaidaraMahamane,RokiaSanogo,AdiaratouTogola,
Mehdi Khaledabadi, Anne C. Vestrheim, Kari T. Inngjerdingen, Terje E.
Michaelsen,DrissaDiallo&BeritS.Paulsen
Anti-ulcerpolysaccharidesfromColacordifoliabarkandleaves
JournalofEthnopharmacology143(2012),221–227.
PaperIV Ingvild Austarheim, Cecilie S. Nergard, Rokia Sanogo, Drissa Diallo &
BeritS.Paulsen
Inulin-richfractionsfromVernoniakotschyanarootshaveanti-ulceractivity
JournalofEthnopharmacology144(2012),82–85.
PaperV IngvildAustarheim,KariT.Inngjerdingen,AdiaratouTogola,DrissaDiallo
&BeritS.Paulsen
Chromatographic immunoassays for H. pylori detection – are they reliable
enoughinMali(WestAfrica)?
AcceptedforpublicationinPANAfricanMedicalJournal(2013)
ix
Relevant co-authored papers
PaperVI KariT.Inngjerdingen,SelmaMeskini,IngvildAustarheim,N’GoloBallo,
Marit Inngjerdingen, Terje E. Michaelsen, Drissa Diallo & Berit S.
Paulsen
Chemical and biological characterization of polysaccharides from wild and
cultivatedrootsofVernoniakotschyana
JournalofEthnopharmacology139(2012),350–358.
PaperVII KariT.Inngjerdingen,BeateK.Langerud,HenrikRasmussen,TrudeK.
Olsen, IngvildAustarheim, TomE.Grønhaug, IngerS.Aaberget, Drissa
Diallo,BeritS.Paulsen,&TerjeE.Michaelsen
PecticpolysaccharidesisolatedfromMalianmedicinalplantsprotectsagainst
Streptococcuspneumoniaeinamousepneumococcalinfectionmodel
SubmittedtoScandinavianJournalofImmunology(2012)
PaperVIII Adiaratou Togola, Ingvild Austarheim, Annette Theis, Drissa Diallo &
BeritS.Paulsen
EthnopharmacologicalusesofErythrinasenegalensis: acomparisonofthree
areasinMali,andalinkbetweentraditionalknowledgeandmodernbiological
science
JournalofEthnobiologyandEthnomedicine4:6(2008)
x