Anti-inflammatory, anti-hyperalgesic, antiplatelet and antiulcer activities of Byrsonima japurensis A. Juss. (Malpighiaceae)

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ContentslistsavailableatSciVerseScienceDirect

Journal

of

Ethnopharmacology

j ou rn a l h o m e pa g e:w w w . e l s e v i e r . c o m / l o c a t e / j e t h p h a r m

Anti-inﬂammatory,

anti-hyperalgesic,

antiplatelet

and

antiulcer

activities

of

Byrsonima

japurensis

A. Juss.

(Malpighiaceae)

Fernanda

Guilhon-Simplicio

a

_,

_Carlos

_Cleomir

_de

_Souza

_Pinheiro

b

_,

_Gabrielly

_Galdino

_Conrado

b

_,

Gleyce

dos

Santos

Barbosa

a

_,

_Pierre

_Alexandre

_dos

_Santos

a

_,

_Maria

_de

_Meneses

_Pereira

a

_,

Emerson

Silva

Lima

a,∗

a_Faculdade_de_Ciências_{Farmacêuticas,}_Universidade_Federal_do_Amazonas,_Rua_Alexandre_Amorim,_330,_Aparecida,_69910-300,_Manaus,_Amazonas,_Brazil

b_Coordenac¸_ão_de_Pesquisas_em_Produtos_Naturais,_Instituto_Nacional_de_Pesquisas_da_Amazônia,_Avenida_André_Araújo,_2936,_Aleixo,_69060-001,_Manaus,_Amazonas,_Brazil

a

r

t

i

c

l

e

i

n

f

o

Articlehistory: Received21July2011

Receivedinrevisedform12January2012 Accepted13January2012

Available online 21 January 2012 Keywords: Byrsonimajapurensis Antioxidant Anti-inﬂammatory Toxicity Amazonian

a

b

s

t

r

a

c

t

Ethnopharmacologicalrelevance:DecoctionsorinfusionsofthestembarkofByrsonimajapurensisA.Juss. (Malpighiaceae)arewidelyusedasananti-inﬂammatorydruginfolkmedicineofAmazonasState(Brazil). Aimofthestudy:Toevaluatethepharmacologicalpotentialofanaqueousextractofthestembarkof Byrsonimajapurensis(BJEA)toscientiﬁcallyverifyofitstraditionaluse.

Materialsandmethods:Anti-inﬂammatory,antihyperalgesicandantiulceractivitieswereevaluatedin Wistarrats,aHippocraticscreeningwasperformedinSwissmicetoevaluatethetoxiceffects,and antiplateletevaluationwasperformedinhumanplateletrichplasmaassay.Additionally,antioxidant activitywasevaluatedbysuperoxideradicalscavengingmethodand␤-carotenebleachingtest. Results:Anti-inﬂammatory,antihyperalgesicandgastroprotectiveactivitieswereobservedinratstreated orallywithdifferentdosesofBJEA.Whilesignalsoftoxicitywereobservedinthemicetreatedwitha veryhighdoseofextract(5000mg/kg),nodeathoccurred.BJEAalsoshowedexpressiveantiplateletand antioxidantactivitiesinvitro.

Conclusion:Accordingtoourresults,itwasconcludedthatstembarkofByrsonimajapurensishas signiﬁ-cantandsafeanti-inﬂammatoryactivity,whichiscloselyrelatedwiththeirpotentantioxidantactivity, supportingthefolkmedicinaluseofthisspecies.

1. Introduction

Inflammationisanonspecificresponseofthemicrocirculation totissueinjurycausedbyphysical,chemicalorbiologicalstimuli, or somecombination of these. However,when there is loss of homeostaticcontrolofthisprocessofdefense,inflammationplays adamagingrolethatcontributestotheappearanceandworsening ofdiseases(Nathan,2002).Thenon-steroidalanti-inflammatory drugs(NSAIDs)utilizedintreatmentofinflammationareonethe mostwidelyusedclassesofdrugsthroughouttheworld,buttheir undesirablesideeffectsongastricmucosaandcardiovascular sys-temarewellknow(WallaceandVong,2008).Therefore,thesearch formoreeffectiveanti-inflammatorydrugsisstillveryrelevant.

Theuseofplantsasmedicinedatesbacktoearlyman.Oftenthis folkmedicineistheonlysourceofaccessforcertainpopulationsfor treatingtheirdiseases(Rates,2001).However,thetraditionaluse ofplantsisnotsufﬁcientforethicalvalidationoftheirtherapeutic

∗ Correspondingauthor.Tel.:+559233055000;fax:+559233055000. E-mailaddress:eslima@ufam.edu.br(E.S.Lima).

effects,requiringscientiﬁcstudiestoverifytheirreal pharmaco-logicalpotential(McChesneyetal.,2007).

The Byrsonima genus(Malpighiaceae) hasabout150species with remarkableneotropical distribution.In various regions of Brazil,severalspeciesofthisgenusarewidelyusedinthe treat-mentofgastrointestinalcomplications.Variousstudiesconfirmed differentbiologicalactivitiesintheseplants,especiallyantioxidant, antimicrobial,includingactionagainstbacteria,fungiandprotozoa andtopicalandsystemicanti-inflammatoryactivities. Phytochem-ically, this genus has being characterized by the presence of flavonoidsandtriterpenoids(Guilhon-SimplicioandPereira,2011). Byrsonima japurensis A.Juss. (Malpighiaceae) isa tree found inlowlandareasoftheAmazonianregionusedinfolkmedicine in ruralareas of AmazonasState(Brazil), where itis popularly knownas“saratudo”andconsideredapotentanti-inflammatory, beingusedagainstpathologiesofgastrointestinaland genitouri-narytracts. Normally,half aglass ofteapreparedby decoction or infusion done with about 5.0% of fragmented stem bark is consumed once a day until the reduction of the symptoms of inflammation,and,inmoreseverecases,twice.Alcoholicbeverages arealsousedinthepreparationoffolkmedicinesbymacerationof

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theplant.Forbothcases,therawmaterialispreviouslydriedin theshadeanditsouterlayerisscrapedforremovalofimpurities. However,thereisnopresentscientiﬁcdatatosupporttheseuses. Therefore this study intends to investigate pharmacological activitiesofByrsonima japurensis, tocontributetothe scientiﬁc validationof itspopularuse. Forthis, weevaluatethe antiede-matogenic,antihyperalgesic,antiplatelet,antiulcerandantioxidant activitiesofanaqueousextractofstembarkofthisplant.In addi-tion,acutetoxicpotentialwasevaluatedin animalexperiments inordertoverifythesafetyoforaladministrationfortherapeutic purposes.

2. Materialsandmethods

2.1. Plantmaterial

The collection of samples wasauthorized by the Managing CounciloftheGeneticPatrimony(CGEN),chairedbytheMinistryof Environment(Brazil),undertheregistration034/2008.Stembarkof ByrsonimajapurensiswascollectedinCareiro-Castanho,Amazonas, Brazil,onMarch2,2007.Theplantmaterial wastaxonomically identiﬁed and authenticatedby José Lima of the Herbarium of theInstitutoNacionaldePesquisasdaAmazônia,whereavoucher specimen(224415)wasdeposited.

2.2. Samplesofhumanblood

Thecollectionofsamplesofhumanbloodwasauthorizedbythe EthicalCommitteeonResearchwithHumansoftheUniversidade FederaldoAmazonas,undertheregistration038/2006.The sam-pleswereobtainedfromhealthyadultvolunteersofbothgenders.

2.3. Animals

Thetestswithexperimentalanimalswereperformedin accor-dancewiththeBrazilianNationalCouncilfortheControlofAnimal Experimentation.FemaleWistarratsweighing100–220gandmale albinoBalb/cmiceweighing20–30gwereusedinthisstudy.The animalswerehousedinpolypropylenecagesat25±2◦Cwith12h ofalight-and-darkcycleandacclimatizedintheexperimental envi-ronmentforatleast24hbeforethetests.Waterandabalanceddiet werecontinuallyprovidedadlibitum,butfoodwaswithdrawn12h beforethetests.

2.4. Drugsandchemicals

Arachidonicacid,adenosinediphosphate,epinephrineand ␭-carrageenanwaspurchasedfromFlukaChemicalCompany(Buchs, Switzerland);nitrobluetetrazolium(NBT),nicotinamideadenine dinucleotide (NADH) and phenazine methylsulfate (PMS) were purchasedfrom Chrono-LogCorporation(Havertown,PA,USA); butylatedhydroxytoluene(BHT),linoleicacidand␤-carotenewas purchasedfromSigma–Aldrich(St.Louis,MO,USA).Otherreagents werepurchasedlocally.

2.5. Preparationofextract

Inthisstudy,theaimwastoreplicatethefolkmedicineusage ofthespecieswhenobtainingthecrude extract.Thestembark wasdriedat40◦Cinastove(WHO,2003),andthenscrapedand pulverizedusingamechanicalgrinder.Theextractwasobtained byinfusionwithdistilledwaterat5%ofplantmaterialfor15min andlyophilizedforfurtheruse.Theyieldofextractionwas10.5%. TheextractobtainedisnamedBJAEinthiswork.

2.6. Hippocraticscreening

Themaximaldoseof5000mg/kgofBJAEdissolvedinnormal salinewereadministeredorallytoagroupofﬁvemice,observing abnormalmorphologicalandbehaviorsignsoftoxicityduringthe ﬁrst4hafteradministrationofextractandatevery24hfor14days (MaloneandRobichaud,1962).

2.7. Anti-inﬂammatoryactivity

Dosesof100,200and400mg/kgofBJAEinnormalsalinewere administeredorallyindifferentgroupsof6rats.Twoothergroups receivedindomethacinat10mg/kg(positivecontrol)andnormal saline(negativecontrol).After1h,asolutionof␭-carrageenan1%in normalsalinewasinjectedintherighthindpawofallanimals,and thepawedemaprogressionwasmeasuredusingadigital plethys-mometer(LE-7500,Panlab,Barcelona,Spain),inhourlyintervals, for5h(Winteretal.,1962).

2.8. Anti-hyperalgesicactivity

Differentgroupsof6ratsreceivedwere100,200and400mg/kg ofBJAEinnormalsalineandindomethacinat10mg/kg(positive control)andnormalsaline(negativecontrol)utilizedineach anal-ysis.After1h,asolutionof␭-carrageenan1%innormalsalinewas injectedintherighthindpawofallanimalsandpressurestimuli wereappliedontheinﬂamedpawathourlyintervalsfor4h,with themaximalloadbornebyeachanimalregisteredbyadigital anal-gesimeter(EFF301,Insight,RibeirãoPreto,Brazil),ingrams(Lapa etal.,2008).

2.9. Antiulceractivity

Ratswereorganizedingroupsofsixanimals.Waterwasused as a vehicle and negative control and ranitidine (50mg/kg, by oralroute)wasusedasapositivecontrol.Dosesof100,200and 400mg/kg ofBJAEwereadministeredin threedifferentgroups. After1h,indomethacin(10mg/kg)wassubcutaneously adminis-teredtoalltheanimals.After3h,alltheratsweresacriﬁcedwith excessofanestheticetherandtheirstomachwascutopenalong thelessercurvature,clearedofresidualmatterwithsalineandthe innersurfacewasexaminedforulcerationandtheulcerindexwas calculated(Djahanguiri,1969).

2.10. Antiplateletactivity

Thisanalysiswasperformedinaccordancewithmethodologyof

Born(1962),wheredifferentconcentrationsofBJAEareincubated withplateletrichplasmaobtainedfromhumanbloodat37◦Cfor 2min.Arachidonicacid,adenosinediphosphateandepinephrine wereusedasplateletinductorsofplateletaggregation. Acetylsali-cylicacidwasusedasapositivecontrol.Thereadingofresultswas doneinaBornaggregometer(PA-04,Qualiterm,SãoPaulo,Brazil).

2.11. Antioxidantactivity

2.11.1. Superoxideradicalscavengingtest

DifferentconcentrationsofBJAEdissolvedin50%ethanolwere incubatedfor5minindarknesswithNBT,NADHandPMSina 96-wellplate,intriplicate.Ascorbicacid,gallicacidand␣-tocopherol wereusedaspositivecontrols.Ethanolat50%wasusedasa neg-ativecontrol.The absorbancewasrecordedat560nmtocheck thebleachingofmixture,indicatingtheradicalscavenging(Ozturk etal.,2007).

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2.11.2. ˇ-Carotenebleachingtest

DifferentconcentrationsofBJAEdissolvedindimethyl sulfox-ide(DMSO)wereincubatedwithanemulsionoflinoleicacidand ␤-caroteneinwatersaturatedwithoxygen,for15minat50◦Cin a96-wellplate,intriplicate.BHTwasusedaspositivecontroland DMSOwasusedasanegativecontrol.Theabsorbancewasrecorded at492nmevery15mintocheckthebleachingofb-carotene, indi-catingperoxidation(Miller,1971).

2.12. Statisticalanalysis

The results are presented as mean_±standard deviation, when applicable. Results of groups in antiedematogenic, anti-hyperalgesic and antiulcer tests were compared with a paired Student–Newman–Keuls test,consideringvalues of p<0.05 sta-tisticallysigniﬁcant.Themedianinhibitoryconcentrations(IC50)

oftheantioxidantand antiplateletactivitieswerecalculatedby linearregressionmethod,andthestatisticalsigniﬁcanceoftheir differenceswasmeasuredwithStudent’st-test.

3. Results

3.1. Hippocraticscreening

Thefollowingsymptomswereobservedinallanimalsduring HippocraticscreeningofBJAE:decreaseinmotoractivity,increase in respiratory andcardiac frequency, dyspnea,palpebralptosis, pilomotorerection,cyanosis,anddecreaseindefecation,urination andappetite.However,nodeathsoccurredduringtheobservation periodof14daysandtheanimalsrecoveredthenormalparameters within48–72h.

3.2. Anti-inﬂammatoryandantihyperalgesicactivities

BJAEpresentedsignificantantiedematogenicactivitywith100, 200and400mg/kg,inearlyphase(firsthour)andlatephase(third hour)ofinflammatoryresponseinducedbycarrageenan,butthe doseof400mg/kgofextractpresentedsignificantlyhigher antiede-matogenicactivity thanthestandard drugin thesecond phase of carrageenan-induced inflammatory response (Fig. 1). In the analysisofanti-hyperalgesicactivity,onlythedoseof400mg/kg showedactivityinbothphasesofcarrageenan-induced inflamma-toryresponse,withgreateractivitythanthein positivecontrol (Fig.2).

3.3. Anti-ulceractivity

The results of our test for evaluation of anti-ulcer activity showedthatdosesof100,200and400mg/kgofBJAEhave signiﬁ-cantgastroprotectiveactionagainstindomethacin-inducedgastric lesion.However,therewerenodifferenceswithindoses. Further-more, ranitidine 50mg/kg tested in same conditions presented betterprotectiveeffectonthegastricmucosa(Fig.3).

3.4. Antiplateletactivity

BJAEpresentedantiplateletactivityagainstdifferentinductors ofaggregation,butitsactivitywasnotgreaterthanacetylsalicylic acid(AAS)testedinthesameconditions.TheIC50ofBJAEwhen

arachidonicacid, adenosine diphosphate and epinephrinewere usedasinductorsofaggregationwas178.5±1.6,117.7±4.5and 97.5±7.9␮g/mL,respectively.ForAAS,theresultswere32.5±0.2, 21.5±1.4and11.7±0.5,respectively.Thedifferencebetweenthe resultsisstatisticallysigniﬁcant,obtainingp-valuessmallerthan 0.01inallthecases.

Edema volume (mL) 0,0 0,1 0,2 0,3 0,4 0,5 Negative control Indomethacin 10 mg/kg BJAE 100 mg/kg BJAE 200 mg/kg BJAE 400 mg/kg ** ** * ♦

A

Edema volume (mL) 0,0 0,1 0,2 0,3 0,4 0,5 0,6 0,7 Negative control Indomethacin 10 mg/kg BJAE 100 mg/kg BJAE 200 mg/kg BJAE 400 mg/kg *** *** *** ♦♦ ***

B

Fig.1. Anti-inﬂammatoryactivityofByrsonimajapurensis.(A)Inearlyphaseof carrageenan-inducedpawedemaand(B)inlatephaseofcarrageenan-induced pawedema;p-valuesversusnegativecontrol:*p<0.05,**p<0.01,***p<0.001,and p-valuesversuspositivecontrol:_p_<_0.05,_p_<_0.01.

3.5. Antioxidantactivity

BJAEpresentsigniﬁcantantioxidantactivityinthesuperoxide radicalscavengingtest.Inthistest,theirIC50was42.5±1.3␮g/mL,

signiﬁcantly lower than ␣-tocopherol (180.0±14.2␮g/mL) and ascorbic acid (32.9±1.6␮g/mL) and higher than Gallic acid (7.8±1.2␮g/mL).Itsinhibitionoflipidperoxidationwasgreater thanBHTtestedinthesameconditions,inwhichIC50was45.7±3.1

and 81.6±7.0␮g/mL, respectively. The difference between the resultsisalsostatisticallysigniﬁcant,obtainingp-valuessmaller than0.01inallthecases.

4. Discussion

Theinflammatoryresponseproducedbycarrageenanisdivided intotwoparts:thefirstphaseoccurswithinfirsthourafter polysac-charideinjectionandismainlyrelatedtothereleaseofhistamine, serotonin,bradykinin,plateletactivatingfactorandleukotrienes; thesecondphaseoccurringwithinthefirstandthirdhours(peakof theinflammatoryresponse)andisduetothereleaseofprostanoids, especiallyprostaglandinsandprostacyclins,bytheactionof1and 2-cyclooxygenases(COX-1andCOX-2)(Vinegaretal.,1969). Dur-inganyinflammatoryresponse,usuallyinnocuousstimuliproduce

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Maximum charge in grams 0 20 40 60 80 100 120 BJAE 100 mg/kg Negative control Indomethacin 10 mg/kg BJAE 200 mg/kg BJAE 400 mg/kg * ♦

A

Maximum charge in grams

0 20 40 60 80 100 BJAE 100 mg/kg Negative control Indomethacin 10 mg/kg BJAE 200 mg/kg BJAE 400 mg/kg * ♦

B

Fig.2.Anti-hyperalgesicactivitiesofByrsonimajapurensis.(A)Inearlyphaseof carrageenaninducingpawedemaand(B)Inlatephaseofcarrageenaninducing pawedema;p-valuesversusnegativecontrol:*p<0.05,andp-valuesversuspositive control:_p_<_0.05.

Index of gastric lesion

0 1 2 3 4 5 6 7 BJAE 100 mg/kg Negative control Ranitidine 50 mg/kg BJAE 200 mg/kg BJAE 400 mg/kg

***

♦

Fig.3. Effectof extracton theformation ofgastriclesions by actionof the indomethacin;p-valuesversusnegativecontrol:***p<0.001,andp-valuesversus positivecontrol:_p_<_0.05.

pain,whichiscausedbythestimulationofafferentnervousﬁbers byactionofkinins,serotonin,histamine,prostanoids,protonsand reactivespecies of oxygen(ROS) releasedduring inﬂammation (ScholzandWoolf,2002).

Superoxideisthefirstreductionproductofmolecularoxygen, anditisanimportantsourceofhydroperoxidesanddeleterious freeradicals(ChauhanandChauhan,2006).ThisROSisinvolved indegenerativediseasesofaging,includingAlzheimer’sdisease, cancerand in theworsening ofinflammatory diseases, suchas rheumatoidarthritis andatherosclerosis,throughDNAand pro-teindamageorlipidperoxidation(Amesetal.,1993;Wisemanand Halliwell,1996).Ithasbeenshownthatlipidperoxidationisrelated totheaggravationandacuteandchronicinflammatoryresponses. Invivo,thisphenomenonisduemainlytotheformationof per-oxynitritefromthecombinationofsuperoxideradicalsandnitric oxidereleasedduringtheinflammatoryresponse(Salveminietal., 2006).

In humans and other species, the COX-1 is expressed con-stitutively throughoutthe gastrointestinaltract(GI), wherethe productionofprostaglandinE2(PGE2)andprostacyclin(PGI2)has

cytoprotectiveeffectsonthemucosabyreducinggastricacid secre-tionbyparietalcellsinthestomach,increasingmucosalbloodﬂow, andstimulatingthereleaseofviscousmucus(RaoandKnaus,2008). Gastricorduodenalulcersdevelopin15–30%ofpatientswho reg-ularlytakeanon-steroidalanti-inﬂammatorydrug(NSAID),which inhibittheactionofCOX-1(BorerandSimon,2005).

The selective inhibitors of COX-2, the inducible isoform of cyclooxygenase,wereintendedtobeasaferalternativeofNSAID togastrointestinaltract.However,theseagentshavebeen associ-atedwithundesirableeffectsonthecardiovascularsystem,such asheartattacksandstrokes,sincethattheydonotinhibit throm-bosis associated with release of thromboxane A2, a powerful

inductorofplateletaggregation,producedbyCOX-1constitutively presentinallplatelets,afterplateletactivationbycollagen, adeno-sine diphosphate (ADP), epinephrine, platelet activating factor, arachidonicacidmetabolites, polycations,serotoninand throm-bin,amongothersubstances involvedininﬂammatoryresponse (Weyrichetal.,2003;SohnandKrötz,2006).

Theresultsoftestsofanti-inﬂammatoryandanti-hyperalgesic activitiesshowedthattheeffectsofBJAEhaveapatternsimilarto theeffectofindomethacin,anNSAIDnotselectiveforCOX-1or COX-2,whichalsopresentedreductionofedemaandpaininboth phasesofcarrageenan-inducedinﬂammatoryresponse,suggesting acommonmechanismofactionbetweenthedrugs.This observa-tionisreinforcedbytheresultsfromtheantiplatelettest,inwhich BJAEwasshowntohaveagoodinhibitoryeffectagainstdifferent plateletinductors,suggestingthattheextractisactingbya mech-anismofinhibitioncommontoalltheinductors,suchasinhibition ofCOX-1(MeadowsandBhatt,2007).

The gastroprotective action against indomethacin-induced gastric lesions suggests that the anti-inﬂammatory and anti-hyperalgesicactivitiespresentedbyBJAEarenotonlyrelatedto theinhibitionofcyclooxygenases,sincethegastriclesionsinduced bynon-steroidalanti-inﬂammatorydrugs(NSAIDs)dependofthe inhibitionofbothCOX-1andCOX-2(Wallaceetal.,2000;Wallace andVong,2008).

Therefore,theactionofaqueousextractasaradicalscavenger ofsuperoxidemayalsocontributetoits anti-inflammatoryand anti-hyperalgesiceffect.Somestudiesshowedthattheremovalof superoxidehassignificantanti-inflammatoryeffects,both preclin-icallyandclinicallyanditisagoodstrategytoinhibitperipheral andcentralsensitizationassociatedwithseveralpainstates(Wang etal.,2004;Salveminietal.,2006).Indeed,itisknownthat superox-idesplayacriticalroleinmediatingtheformationofperoxynitrite inbothearlyandlatephasesofthecarrageenan-induced inflam-matoryresponse(Salveminietal.,1996;Cuzzocreaetal.,1998).

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Thesignalsoftoxicityand/orpharmacologicalactivityobserved inHippocraticscreeningofBJAEareverysimilartothosepresented byByrsonimacrassifoliaintwoprevioussimilaranalyses( Guilhon-SimplicioandPereira,2011).Intherespectivepapers,theauthors suggestthatthespecieshasaneffectonthecentralnervous sys-tem(CNS).Thishypothesisshouldbefurtherinvestigated,sincethe criticalroleofnicotinicacetylcholinereceptorsintheregulationof inﬂammationhasbeenestablished(Wangetal.,2003).

Theresultsobserved inthisstudycanbepartiallyexplained bythepresenceofdifferentclassofflavonoidsthatwedetected inthespeciesbyphytochemicalscreening,highperformance liq-uid chromatography, nuclear magnetic resonance and infrared spectrometry fingerprint (unpublished results). The capacity of these substances to inactivate enzymes involved in inflamma-toryresponse,suchphospholipasesandcyclooxygenasesandtheir expressiveradicalscavengingactivityiswellknown(Middleton et al.,2000).The increase in respiratory and cardiac frequency observedinHippocraticscreeningcanbeexplainedbythe pres-enceofcardioactiveheterosideswhichwasalsodetectedinthe samestudy.

5. Conclusion

Thisworkmetitsgoalofscientificallyverifyingthepopularuse ofstembarkofByrsonimajapurensis,whoseteaiswidelyconsumed infolkmedicineofAmazonasState(Brazil)asananti-inflammatory drug.Theaqueousextractobtainedbyinfusionat5%ofthisvegetal drugshowedgoodantiedematogenicandanti-hyperalgesic poten-tialafteroraladministrationinanimalexperiments.Indeed,this extractalsoshowedgoodantiplateletandanti-ulceractivitiesthat mayhavefewersideeffectscomparedtotraditionalNSAIDs.The expressiveantioxidantactionofextract,inadditiontothealready well-knownbenefitstohealth,maybecontributingtoits mecha-nismofanti-inflammatoryeffect.Itisimportanttohighlightthat nodeathoccurredafterexposuretoveryhighdosesofBJAE,which confirmedthesafetyofitsusefortherapeuticpurposesandfor advancementofstudiesinvivowiththisrawmaterial.Therefore, thisspeciesisapromisingobjectforfurtherstudiesaimingto sup-portitsmechanismofpharmacologicalactivitiespresentedinthis work.

Acknowledgements

TheauthorsaregratefultotheConselhoNacionalde Desenvolvi-mentoCientíficoeTecnológico(CNPq)andFundaçãodeAmparo àPesquisado Estadodo Amazonas(FAPEAM)forfinancial sup-portofthisresearchandtoCoordenaçãodeApefeiçoamentode Pessoalde NívelSuperior (CAPES) for theprovision of scholar-shipstothefirstauthor.E.S.L.ismembersoftheINCTdeProcessos RedoxemBiomedicina-Redoxoma(MCT/CNPq). AcademicEnglish-Solutions.comrevisedtheEnglish.

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