w w w. s b f g n o s i a . o r g . b r / r e v i s t a
Review
On
the
use
of
nanotechnology-based
strategies
for
association
of
complex
matrices
from
plant
extracts
Giovanni
Konat
Zorzi
a,
Edison
Luis
Santana
Carvalho
b,
Gilsane
Lino
von
Poser
a,
Helder
Ferreira
Teixeira
a,∗aProgramadePós-graduac¸ãoemCiênciasFarmacêuticas,UniversidadeFederaldoRioGrandedoSul,PortoAlegre,RS,Brazil
bProgramadePós-graduac¸ãoemProdutosBioativoseBiociências,UniversidadeFederaldoRiodeJaneiro,CampusMacaé,RiodeJaneiro,RJ,Brazil
a
r
t
i
c
l
e
i
n
f
o
Articlehistory: Received6June2015 Accepted22July2015 Availableonline20August2015
Keywords: Essentialoil Liposomes Nanoemulsions Nanoparticles Nanotechnology Plantextract
a
b
s
t
r
a
c
t
Dependingon themethodof extraction,plantextracts cancontainanenormousvarietyof active molecules,suchasphenoliccompounds,essentialoils,alkaloids,amongothers.Inmanycases,from apharmacologicalpointofview,itisinterestingtoworkwithcrudeextractorfractionsinsteadofa singleisolatedcompound.Thiscouldbeduetomulti-targetingeffectoftheextract,lackofknowledgeof theactivecompounds,synergisticeffectoftheextractcompounds,amongothers.Inanycase,inorder toachieveafinalproductsomeissuesmustbeovercome,includingpoorstability,solventtoxicity,and lowsolubilityofthebioactivecompound.Recentlymanynanotechnology-basedstrategieshavebeen proposedasanalternativetosolvetheseproblems,especiallyliposomes,nanoemulsionsand nanopar-ticles.Inthissense,thepresentworkaimstoreviewthemainnanotechnologicalapproachesusedfor associationofdifferentplantextractsandthemainachievementsfromusingthesetechnologies.
©2015SociedadeBrasileiradeFarmacognosia.PublishedbyElsevierEditoraLtda.Allrightsreserved.
Introduction
Mankindalwayshadacloserelationshipwithplants,usingthem forthousandsofyearsassourceofpotentialmedicinestorelieve bothphysicalandspiritualpain.Throughoutthecenturies,the ther-apieshave evolvedand theuseofcomplexes mixturesasplant extractshasbeensystematicallyreplacedbytherapiesusinga sin-gleisolatedcompound.Nevertheless,theplantsarestillamajor sourceofbioactivecompoundsinmodernmedicine(Rates,2001). Ideallyasingle,pureandisolatedcompoundshouldbeusedin thedevelopmentofaformulation.Nevertheless,thisisnotalways themostviableorsuccessfulapproach.Amongthereasonsofusing complexmixtures,emphasiscouldbegiventothesynergismwith differentcompounds,thelossofactivityafterisolation,chemical instability,difficultyinthepurificationandthepossibilitytoact inmultipletargetsatonce(Gertsch,2011;Kimetal.,2008;Rando etal.,2009).However,seldomaplantextractitselfcanbeusedasa finalproduct.Asforpharmaceuticaldosageforms,some technolog-icalprocessmustbeperformedinordertoobtainafinalmedicine thatcanassurequality,safetyandefficacy(Pristaetal.,2011).
∗ Correspondingauthor.
E-mail:helder.teixeira@ufrgs.br(H.F.Teixeira).
Inthepastyears, thenumberofpublications relating tothe useofnanotechnology-basedsystemsassociationplantextracthas beengrowing.Thereisalreadyawell-documentedliteratureabout thebenefitsofassociatingisolatedcompoundsto nanotechnolog-icaldrugdeliverysystemsand extendingthesestudiestomore complexesmatrices,suchasplantextracts,wasjusta matterof time.Thereisacleardivisionamongtheworksrelatingtoplant extractandnanotechnologyandtwowelldefinedgroupscanbe distinguished.Thefirstconsiderstheuseofplantextractforthe for-mationofmetallicnanoparticles(Mittaletal.,2013)andthesecond, theuseofnanotechnology-basedsystemstoimprove biopharma-ceuticalandtechnologicalpropertiesofplantextracts(Ajazuddin andSaraf,2010;Bonifacioetal.,2014).Thelaterwillbethefocus ofthisreview.
Many nanostructures have been proposed for drug deliv-ery,eachonehavingtheirownadvantagesanddrawbacks.This manuscriptrevisestheuseoflipid-andpolymer-based nanostruc-turesintheassociationofdifferentextract,establishingarelation betweenthetypeofnanosystemsanditspreparationmethodto thedifferentplantextractsandmostabundantcompounds.
Nanosystems:characterizationandpreparationtechniques
So far,the association of plant extracts has been described forliposomes, nanoemulsionsand nanoparticles(eitherlipidor
http://dx.doi.org/10.1016/j.bjp.2015.07.015
Nanoparticles
Nanosphere Nanocapsule
Nanostructured lipid carrier Solid lipid
nanoparticle
Liposome
(o/w)
Nanoemulsion
Fig.1.Differentarchitecturesofnanosystemsusedtoimprovebiopharmaceutical propertiesofplantextracts.
polymer-based nanoparticles) (Fig. 1). Factors such as polarity ofactive compound, solubility,presenceof organicsolventand volatility must be taken into consideration when selectingthe nanosystemand its preparationtechnique. For example, essen-tialoilsarerichinlipophiliccompoundsandextremely volatile molecules.Inthissense,lipid-basedsystemspreparedatlow tem-peraturesandwithoutsolventevaporationaftertheadditionof theoils,suchasliposomeornanoemulsionaremorelikelytobe successful.
Regardlessthetype ofnanosystemorpreparationtechnique, thereare somebasiccharacterizationsofthesystem.Thebasic physicochemicalparametersareaveragesize,surfacechargeand associationefficiency.
Determinationofthesizeofcolloidalstructurescanbedone byseveraltechniquessuchastransmissionelectronmicroscopy, laserdiffraction,andsmallangleX-ray.Amongthem,dynamiclight scatteringoffersseveraladvantagessuchaswiderange of mea-surement,simplicityandreliability(BerneandPecora,2003).Itis anon-destructivetechniquethatrequiresasmallamountof sam-ple(normallydiluted).Nevertheless,forpolydispersedsamplesthe fluctuationsof angle-dependencyof scatteredlightmayleadto unrealisticresults.Asalternative,theuseofCONTINalgorithmfor dynamiclightscatteringornanoparticletrackinganalysisis recom-mended,beingthelateralsousedtodeterminetheconcentration ofnanoparticles(Filipeetal.,2010).Reductionofsizeresultsin theincreaseofsurfacefreeenergyand,inthisway,the prepara-tionmethodmustbechosenwisely(Shaw,1992).Thereduction ofinterfacialtensionalsodecreasestheamountofenergyrequired forloweringtheaveragesize,asdescribebytheLaplaceequation (Buttetal.,2003).Inthenanometricworld,someforcesthatplay amajorroleinthemacroscopicworldareminimized.The Brown-ianmotion,forexample,ismoresignificantthanthegravity.Ina similarway,thereductionofsizecanaltersome physicochemi-calpropertiesofbulkmaterial(Hiemenz,1997).Perhapsthemost emblematicistheincreaseinthesolubility.Duetothehighsurface areaofnanostructured colloids,thesolubilityoflipophilic com-poundisincreased,asdictatedbytheKelvinequation(Buttetal., 2003;Cosgrove,2010).Ultimately,thiscanresult inlossofthe activecompoundassociatedandincreaseinsizeduetotheOstwald ripening(Buttetal.,2003;Myers,1999).
Size,however,seldomcomesasasinglevaluebutrathera rep-resentativevalueofadistributionthatcanbenarrow,moderate
Table1
Amountofextract(dryweight)usuallyassociatedtoeachnanotechnology-based system.
System Amountofextract(%)
Nanoemulsion 0.5–20
Nanoparticle 0.5–2.0
Liposome 0.1–2.0
orbroad.Inthissense,thepolydispersityindexisamathematical approachtomeasurethewidthoftheparticlesizedistribution, beingthesquareof thedivisionof standarddeviationbymean diameterfordynamiclightscattering.Thispropertyvariesfrom0 to1,wherelowervaluesindicatenarrowerdistribution(Berneand Pecora,2003).Manyfactorsaffectthepolydispesityofasystem. Forfreshlypreparedformulations,themanufacturingtechniqueas wellastypeandamountofconstituentsplaysamajorrole.Change ofpolydispersitythroughtimeisoftenassociatedtostabilityissues (Washington,1992).
Estimationofthesurfacechargeisimportanttopredictcolloidal stabilityandadequabilityofrouteofadministration.Accordingto theDLVOtheory,colloidalstabilityisbasicallydependentonsteric hindranceandthebalanceofattractive/repulsiveforces(Hunter, 1989,2001).Inthisway,keepingthesurfacechargefarfrom neu-trality isa way tofavor thesystemstability. In thesame way, themoduleand intensityofsurfacecharge mayresultin unde-siredinteractionswithbodyfluidsandtissues(Peetersetal.,2005; Rabinovich-Guilattet al.,2004).For example,dependingonthe charge,moleculesassociatedtonanosystemmaypermeatemore orless intheskinlayers (Gillet etal.,2011; Ogisoetal.,2001; VenugantiandPerumal,2009).So,agivenchargemayberequired dependingonrouteofadministration.Thesurfacechargeiseasily estimatedbasedonthequantificationofthezetapotential,the dif-ferenceofpotentialontheslippingplaneofthecolloid(Birdi,2009; Buttetal.,2003).Thisdeterminationofzetapotentialisdoneby electrophoreticandelectroacousticmethods,speciallyevaluating theelectrophoreticmobilityoftheparticles(Hunter,1989).
Thus,zetapotentialcanbemodulatedbyajudiciousselection of materials that would be on the interface of nanostructure-continuousphase,suchassurfactants,andcompoundsthatwould beadsorbedonthesurfaceofthestructures(e.g.polymersor com-poundsfoundintheplantextract)(Zorzietal.,2011,2015).
Anotherimportantfactor isthedetermination ofthe associ-ation efficiency, i.e. theamountof active compound associated withthecolloidalstructure.Thisisatrickytask,however,because mostextractsarenotfullycharacterizedand/oramarkerisused torepresentthetotalcontent.Thebestwaytoproceedis deter-miningthe dry residue and quantifytheamount ofmarker (s) in respect to the dry residue, even if the extract is not used in its dried from. A judicious selection of the marker must be doneand themethodshouldbepreviously validated.Recently, Nemitzetal.presentedavalidatedmethodtoquantifyisoflavone agliconefractionfromGlycinemaxethanolicextractthatwas incor-poratedintonanoemulsion(Nemitzetal.,2015).Threerelevant isoflavones,daidzein,glycitein,and genistein, couldbe simulta-neously quantified and the method was validated to different matrices(nanoparticles,porcineskin,andesophagicmucosa). Sim-ilarly,Diasetal.havevalidatedahead-spaceGC–MSmethodfor thequantificationofCopaiferamultijugaessentialoilin nanoemul-sions (Dias et al., 2012). HPLC-UV validation of Achyrocline satureioideshydroethanolicextractsassociationtonanoemulsions wasalsorecentlyreported.Threeflavonoids withreported bio-logical activity couldbe simultaneouslydetermined (quercetin, 3-O-methylquercetin,andluteolin)(Bidoneetal.,2014a).
thesystemsthat canvery oftenincorporatehigher amounts of extract(especiallywhentheactiveoilisusedastheoilcoreof nanoemulsion),butmayexhibitimportant instabilityoncethey arenotthermodynamicallystablesystems.Nanoparticlescanbe madeusingdifferentmaterial,yieldingdifferentresponses. Poly-mericnanoparticlesaremadeusingpolymers,lipidnanoparticle andnanostructured lipidcarriers are madeusinglipid whereas nanocapsulesaremadeusingacompositionofpolymerandlipids.
Liposomes
Liposomesarevesicular structurescomposed ofamphiphilic molecules, suchas phospholipids. Their size can range from a few nanometers to several micrometers, being both prepara-tiontechnique and componentsimportant to definetheirfinal physicochemicalpropertiessuchsurfacecharge,size,andstability (Gregoriadis,2007b).Dependingonthesizeandnumberoflayers, theycanbeclassicallydividedintofourgroups:smallunilamellar vesicles(SUV), largeunilamellarvesicles (LUV), giant unilamel-larvesicles(GUV),andmultilamellarvesicles(MLV)(Gregoriadis, 2007a).Thepolargroupsofphospholips(orothercomponent)are linedtowardtheexteriorandinnercavity,wherehydrophilic com-poundscanbeadsorbed.Ontheotherhand,lipophilicdrugscanbe solubilizedwithinthebilayeramongthehydrophobicalkylchains (ChoiandMaibach,2005;Kauretal.,2004).Liposomesareuseful vehiclesfortopicaldrugdeliveryoncetheycanprovideprolonged andcontrolledactionafterapplicationandalsopreventenzymatic degradation(Fenskeetal.,2008;Mainardesetal.,2005).Themost commonand simplest preparation technique is film hydration, whereathinlipidfilmishydratedwiththedesiredsolution. Alter-nativelythereareethanolinjectionandreversephaseevaporation, thelaterrecognizedashavingsuperiorentrapmentofhydrophilic substancesinthevesiclecavity.Regardlessthetechniqueofchoice, asecondstepofsizingdowntheliposomesanddecreasethesize distributionmayberequired,suchasmembraneextrusion, ultra-sound,high pressure homogenization, freeze and thaw, among others.
Nanoemulsions
Nanoemulsionsaredispersionsoftwoimmiscibleliquids stabi-lizedbyasurfactantorsurfactantsystem.Theirphysicalstability canbe substantially improved by selectingsuitable emulsifiers that arecapable of surroundingthedispersed droplets in such wayastoreduceinterfacialtensionorincrease droplet-droplet repulsion(Solansetal.,2005;Tadrosetal.,2004).Dependingon theconcentrations of oil/water/emulsifierand the efficiency of theemulsificationequipmentused toobtaina reduced droplet size,thefinalemulsionmayhavedifferentcharacteristics.Inthis context,themoststudiednanoemulsionsystemsfor pharmacolog-icalapplicationsareoil-in-water(o/w)nanoemulsion(Tamilvanan, 2004).Spontaneousemulsificationandhighpressure homogeniza-tionarethemostusedtechniqueswhenassociatingplantextracts tonanoemulsion,butothermayalsobefoundsuchasphase inver-siontemperature.
Nanoparticles
Nanoparticlesarestructureswithsizebetween1nmand1m, mostlyofpolymericorlipidicnatureand,dependingontheendof use,mayormaynotincludeanactivemolecule(RaoandGeckeler, 2011;Sahooetal.,2007).In thismanuscriptthetermpolymeric nanoparticlewillbeusedtodescribeamatricialstructurewherethe activemoleculecanbeassociatedintothematrixand/oradsorbed onthesurface.Ontheotherhand,thetermnanocapsuleswill rep-resentstructureswithanoilcore(thatmayornotbetheactive
compound)surroundedbyapolymericshell.Inthisway,theactive compoundcanbeentrappedinthepolymer/oilcoreorbeadsorbed onthesurfaceofthenanocapsule.Therearemanytechniquesthat canbeusedtopreparepolymericnanoparticlesandnanocapsules dependingonthenatureofthepolymer.Ionicgelationand nano-precipitationare suitable for hydrophilic polymerswhereas for hydrophobicpolymersthetechniqueofchoiceissolvent displace-mentfollowed bynanoprecipitationofthepre-formedpolymer usingwatermiscibleorganicsolvents(e.g.ethanolandacetone).
Lipid-basednanoparticlescanbedividedintwogroups:solid lipid nanoparticles and nanostructured lipid carrier. Solid lipid nanoparticlesarenanostructurescomposedofsolidlipidsatroom andbodytemperature(Soutoetal.,2007;SoutoandMueller,2008). Dependingonthefabricationtechnique employed,theuseof a surfactantorco-surfactantcan berequired inorder tostabilize thesystem(Muchowetal.,2008).Eventhoughaveryattractive system,solid lipidnanoparticlesmayexhibitseveralissuesthat willlimittheirapplication.Problemssuchaslowloadingcapacity andthepotentialexpulsionofthedrugduringstorageareknown drawbacks.Evenlipophilic moleculescan beexpelledfromthe nanoparticlestructure due torearrangementoflipids(Pardeike etal.,2009).Thiscanbeinpartovercomebytheincorporation ofliquidlipids(ormixtureofdifferentlipids)tocreateamatrix that isas disorganized aspossible.These systemsare so-called
nanostructuredlipidcarriers(Fangetal.,2013).
Plantextracts
Forthesakeofsimplicity,inthisreview“plantextract”willbe consideredasacomplexmatrixobtainedfromaplantthatmay ormaynotcontainresidualsolvent.Thereisa vastsetof tech-niquessuitableforplantextractioninvolvingdifferentpartsofthe plantandmaintargetedcompounds.Dependingonthetechnique andprotocol,theextractionmayfavoragivengroupofsecondary metabolitesoveranother.Fractioningoftheextractmayoccur get-tingridofundesiredcompounds.Nevertheless,this reviewwill onlytakeintoconsiderationextractsassociatedwithnanosystems thatarestillacomplexmatrix(i.e.twoormorecompounds).In thisway,dependingonthelipophilicityandthenanostructureof choice,theextractcompoundscanbebasicallyfoundinthree dif-ferent places:(i)solubilized intheexternal aqueousphase; (ii) adsorbedontothesurfaceofthecarrier,and(iii)entrappedinto thecarrier.
Aqueousextracts
Aqueousextracts are those in which only water is used as extractionsolvent.Thereareseveralreasons whytochoosean aqueousextractionincludingprice,easinessinhandlingthe pro-cess, necessity of avoid a given class of substances, safety and environmentalconcerns.Themajorityofthecompoundextracted, suchasaminoacids,sugars,mucilages,heterosides,saponins,and alkaloidsinthesaltform,willhaveahydrophilicnature(Simões et al.,2001).Box 1shows someof therelevant applicationsof nanosystemforaqueousextracts.
Box1:Representativeexamplesofaqueousextractsassociatedtodifferentnanosystems.
Plant Partused Activecompound Activity System Reference
Whitetea Leaves Phenoliccompounds(catechins) Antioxidant Nanoparticle(PECL, alginate,Pluronic®
F-127)
Sannaetal.(2015)
Phoenixdactylifera Pit Phenoliccompounds – Nanocapsule(whey protein)
Bagherietal.(2013)
Centellaasiatica Dryleaves Asiaticoside Madecassoside Asiaticacid Madecassicacid
Antioxidant Nanoparticle(gelatin) Kwonetal.(2012)
Camelliasinensis Leaves Phenoliccompounds (epigallocatechin, epigallocatechin-3-gallate, epicatechin,epicatechin-3-gallate)
Antioxidant Nanoemulsion(lecithin, palmorsunfloweroil, Tween®20or80)
GadkariandBalaraman (2015)
Hibiscussabdariffa Flowers Phenoliccompounds (anthocyanins)
Antioxidant Liposome (lecithin)
Gibisetal.(2014)
Pluronic®F-68,polyoxyethylene-polyoxypropyleneblockcopolymer;PECL,poly(epsilon-caprolactone);Pluronic®F-127,polyoxyethylene-polyoxypropyleneblock
copoly-mer;Tween®20,polyoxyethylenesorbitanmonolaurate;Tween®80,polyoxyethylenesorbitanmonooleate.
cationofanthocyaninswiththenegativephophospholipidspresent inthelecithin.Lecithinisacomplexmixtureoflipidsanddueto thepresenceofacidphospholipids(suchasphosphatidylserineand phosphatidylinositol)caninteractwithpositivelycharged com-pounds.Inthisway,theauthorsproceedwithachitosancoatingfor theseliposomes.Chitosanisaheteropolymerderivedfromchitin deacetylation,andconsistsofseveralunitsofN-acetylglucosamine andN-glucosamine(Aranazetal.,2009;Dashetal.,2011; Garcia-FuentesandAlonso,2012).Duetothepresenceofseveralprimary aminegroupsitcanalsointeractwithlecithin-basedliposomes (Agnihotri etal.,2004;Menchicchietal.,2014).Asecondcover withpectincouldalsobeaddedinordertohaveafinalnegativezeta potential.Overall,theratiobetweenlecithinandextractprovedto becriticaltoavoidaggregation.Toimprovestabilityasecond poly-mericlayer,inthiscaseofpectin,canbeaddedtoavoidaggregation andincreaseinsize.
Gelatin nanoparticles associating Centella asiatica aqueous extractwasproposedasskin-protectiveagent,duetothepresence ofasiaticoside, madecassoside,asiaticacidand madecassic acid (Kwonetal.,2012).Nanoparticleshadsuperiorprotectiveeffecton UVA-inducedmatrixmetalloproteinase(MMP-1)and tyrosinase-inhibitoryactivitywithrespecttofreeextract.Oncetheauthors didnotuseanycrosslinkerordenaturation,itisreasonabletothink thatgelatinactuallyactsstabilizingtheextract.Thespontaneous formationofnanoparticlesfromaqueousextracts,especiallyupon cooling,hasalreadybeenreportedforblackteaextract,for exam-ple(Gröning and Breitkreutz,1994; Gröning etal.,2002).Afar lessusualmaterialforpharmaceuticalapplication,wheyprotein nanocapsulescouldassociatedatepalmaqueousextract(Phoenix dactylifera)withanencapsulationefficiencyof70–78%(Bagheri et al., 2013).The nanocapsules were prepared by coacervation (usingNaClandethanol)andstabilitybyheat,yieldingaformation around90%withsizerangingfrom100to160nm.The encapsu-lationofthisphenoliccompounds-richextractmayfoundinfood industryaninterestingapplication.
Green tea (Camellia sinensis) aqueous extract is rich in catechins,especiallyepigallocatechin-3-gallate, epigallocatechin, epicatechin-3-gallate, and epicatechin. Generally recognized by itsantioxidant activity,thegreen teaextract wasassociatedto alginateand poly(epsilon-caprolactone)(PECL)nanoparticles.As expected,lowentrapmentoftheactivecompounds,expressedby termsofepigallocatechin-3-gallateandepicatechin-3-gallate,was found(around30%).Nevertheless,theassociationofcatechinsto thepolymericmatrixcouldmodulatetheirreleasewhere25%was foundafter2hin comparison tonearly100% release fromfree extract(Sannaetal.,2015).Controlofkineticreleaseofantioxidant
isgenerallyrecognizedasthemainreasonforthesuperioractivity ofnanoencapsulatedantioxidants(Bidoneetal.,2014b;Coradini etal.,2014).Nevertheless,notalwaysanincreaseinthe antioxi-dantactivitycanbeobserved.GadkariandBalaraman(2015)used nanoemulsiontoassociateaqueousextractsofgreenteaandno differenceintheantioxidantactivity(ferricreducingantioxidant potential)wasobservedbetweenthefreeextractandthe formu-lations.Even thoughthe authorsdidnot provide thevalues of association efficiencyof the activecompounds (catechins), this resultisnotfarfromexpected.Duetothehighlipophilicityofthe nanoemulsionoilcore,alowassociationofcatechinsisexpected, especiallybecausetheextractusedwasaqueous.
Alcoholicextracts
Ethanolis oneof themostusedorganicsolvents forextract preparation. The ability to solubilize a wide variety of sub-stances, low price,relatively low toxicity and thefact of being anenvironment-friendlysolventare amongthemost attractive properties that make ethanolic extractspopular. Depending on the preparation method, there is no need of previousremoval of ethanol.Taking solventdisplacement techniqueas an exam-ple,theethanolicextractcanbeaddedtotheorganicphaseand thesolventremovedattheendofthenanoparticle/nanoemulsion formation. Box 2 exemplifies somerelevant formulationsusing alcoholicextracts.
Aspreviouslystated,thebiologicalactivityofantioxidant-rich extractscanbeimproveduponassociationtonanostructures. Phe-nolic compounds of Orthosiphon stamineus (Aisha et al., 2014),
Fraxinusangustifolia(Moulaouietal.,2015),Curcumalonga(Kaur andSaraf,2011),Cuscutachinensis(Yenetal.,2008)andVitisvinifera
(Spignoetal.,2013)wereassociatedtoliposomes,nanoemulsions andnanoparticles.
Liposomal formulation significantly improved the solubility of Orthosiphonstamineusethanolic extracts(Aisha et al.,2014). Entrapmentefficiencyofthemarkersrosmarinicacid, 3-hydroxy-5,6,7,4-tetramethoxyflavone,sinensetinandeupatorinvariesfrom 30to55%.Releasestudiesfromliposomesshowedimprovement in DPPH scavenging effect (around50%) in comparison to free extract.Ethosomes(liposomeswithpermeationenhancers,suchas ethanol)favorthewoundhealingpropertiesofFraxinusangustifolia
Box2:Representativeexamplesofethanolicextractsassociatedtodifferentnanosystems.
Plant Partused Activecompound Activity System Reference
Curcuma longa Dryrhizomes Phenoliccompounds (curcumin)
Antioxidant Liposome(PC,cholesterol) Ethosome(PC)
Tranferosome (lecithin:sodium deoxycholate)
KaurandSaraf (2011)
Fraxinusangustifolia Leavesandbarks Phenoliccompounds (quercetin,catechin,rutin andtannicacid)
Antioxidant Ethosome(lecithin) Moulaouietal. (2015)
Orthosiphonstamineus Leaves Phenoliccompounds (rosmarinicacid,eupatorin)
Antioxidant Liposome(lecithin) Aishaetal.(2014)
Phytolaccadecandra Roots Triterpenes Antineoplasic Nanoparticle(PLGA) Dasetal.(2012)
Barberared-grape(Vitisvinifera) Fruits Phenoliccompounds Antioxidant Nanoemulsion(lecithin, peanutorsunfloweroil, maltodextrins)
Spignoetal.(2013)
Cuscutachinensis Seeds Phenoliccompounds (quercetin,kaempferol)
Hepatotoxicity Nanoparticle(Pluronic®
F-68)
Yenetal.(2008)
PC,phosphatydilcholine;PLGA,poly(lactic-co-glycolicacid).
Liposomes,ethosomes andtransferosomes (aspecial type of liposomewitha membrane softening agent)wereprepared for loadingCurcuma longa (Kaur and Saraf,2011). The size varied withthe extract loading(0.5–2.0%) and theextract association efficiencyand hydrationlevelofskinwasasfollowed: transfer-osome>ethosomes>liposomes.
Stabilization of nanoparticles formed form Cuscuta chinen-sis ethanolic extract was performed in order to improve the antioxidant activity (Yen et al., 2008). This extract is rich in quercetin, kaempferol and their glycosides and was proposed for acetaminophen hepatotoxicity management. The authors concluded that after oral administration in Wistar rats, the nanoparticlesrequiredadose5-timeslowerthanthefreeextract toachievethesameeffect.
Phytolaccadecandraextract-loadedpolymericnanoparticlesof poly(lactic-co-glycolicacid(PLGA)werestudiedasanalternative intheinterventionagainstinducedlungadenocarcinomainmice
andonA549cells (Dasetal.,2012).Themajorcomponent was identifiedasaderivativeofbetulinicacidandhadentrapment effi-ciencyaround80%.Nanoencapsulationshowedpreventiveeffects concerningtoxicitybiomarkers(reactiveoxygenspecies genera-tion,CYP1A1,caspase3amongothers)andDNAfragmentationin higherextendedincomparisontofreeextract.
Hydroalcoholicextracts
Hydroethanolicmixtures are ableto extract more lipophilic compoundsthanaqueousoneduetothelowerdielectricconstant ofethanol.In thisway,higheramountsofextract,expressedin termsofdryweight,canbeincorporatedwithbetterentrapment. InBox3,manyapplicationsofhydroalcoholicextractsassociated toliposomes,nanoemulsionsandnanoparticlescanbeobserved.
As mentioned before, nanoencapsulation can bring several advantagesfortheadministrationofaplantextract.Amongthem
Box3:Listofsomerelevanthydroalcoholicextractsincorporatedtodifferentnanosystems.
Plant Solvent Partused Activecompound Activity Systems Reference
Panaxquinquefolius 0–90%ethanol Dryroots Saponin(Ginsenoside) Antioxidant Liposome (DSPE-PEG2000)
Tsaietal.(2012)
Polygonumaviculare 50%ethanol – Phenoliccompounds (quercetinandmyricetin)
Antiaging (antioxidant)
Liposome(DOPCand cholesterol)
Kwonetal.(2015)
Phyllanthusurinaria 30%ethanol – Phenoliccompounds Skinantiaging Nanoemulsion (Tween®80,Span®80)
Mahdietal.(2011)
Vellozia squamata 70%ethanol Leaves Phenoliccompounds Cosmetic Nanoemulsion(Babac¸u oil,Span®80
PEG-40)
Quintãoetal.(2013)
Glycinemax 60%ethanol Beans Isoflavoneaglycones Phytoestrogenic Nanoemulsion(lecithin, Tween®80,MCT)
Nemitzetal.(2015)
Achyroclinesatureioides 80%ethanol Flowers Phenoliccompound (3-O-methylquercetin)
Antiviral Nanoemulsion (lecithin,MCT)
Bidoneetal.(2014a,b)
Phenoliccompound (quercetin)
Topical antioxidant
Nanoemulsion (lecithin,ODD) Nanoparticle(PECL, Span®80,Pluronic®
F-68)
Zorzi(2007)
Carvalhoetal.(2008)
Ziziphusjujuba 40–100%ethanol Fruit Phenoliccompound (unspecific)
Antioxidant Nanoparticle(chitosan) Hanetal.(2015)
Salviaofficinalis 60%ethanol Leaf Phenoliccompounds Antioxidant (mercury poisoning)
Nanoparticle(PLGA, Pluronic®F-68)
KahilandSalvia(2013)
Picrorhizakurrooa 80%methanol Rootand rhyzome
Iridoids(PicrosideIandII) Hepatoprotection Nanoparticle(PLA, Pluronic®
F-68)
Jiaetal.(2015)
DOPC,1,2-dioleoyl-sn-glycero-3-phosphocholine;Pluronic®F-68,polyoxyethylene-polyoxypropyleneblockcopolymer;MCT,mediumchaintriglycerides;ODD,octyl
dodecanol;PECL,poly(epsilon-caprolactone);PEG,polyethyleneglycol;PLA,poly(lacticacid);PLGA,poly(lactic-co-glycolicacid);Span®
80,sorbitanmonooleate;Tween®
thereisthepossibilitytoprovidedrugsustainedreleaseinawayto decreasesideeffectanddecreasedosage.Ziziphusjujubafruitshave alowshelf-lifeduetothefastoxidationofitsphenoliccompounds (Han et al., 2015).In this way,the encapsulationof previously optimizedhydroethanolicextract(40–100%ethanol)inchitosan nanoparticlesincreasedthestabilityofthephenoliccompoundsin respecttofreeextract(DPPHantioxidantactivity).Regardlessthe plantpart,seedorpulp,highentrapmentefficiencywasobserved (80–90%).
PLGA nanoparticleswere developed to associate methanolic extractofPicrorhizakurrooa,aHimalayan-nativeplant(Jiaetal., 2015).ThisextractisrichintheiridoidspicrosideIandII, hepato-protectivecompoundswithpooraqueoussolubilityandintestinal absorption.Extract-loaded nanoparticleshad upto67% associa-tionefficiencyoftheactivecompound andwereabletosustain therelease upto200h. Theintermediate associationefficiency foundmaybeattributedtotheuseofmethanol,whichhasahigher dielectricconstantthanethanol,withconsequenthigh hydrophilic-ity of the extract. Thiswas ultimately resulting in low affinity tothehydrophobicPLGA(Roweetal.,2003).Dependingonthe amountofethanol,hydroalcoholicextractsofSalviaofficinalismay berichinphenoliccompoundssuchasrosmarinic,caffeicand sal-vianolicacids.Inthisway,60%ethanolextractwasassociatedto PLGAnanoparticlesinordertoincrease theantioxidantactivity againstmercury-inducedneurotoxicity(Kahil andSalvia,2013). Thenanoparticleswereabletoreducetheformationof oxygen reactivespeciesandthealterationsincortexofalbinoratsafter neurotoxicityinducedbymethylmercury.Thiseffectwassuperior tonon-treatedcontrol andnonencapsulatedextract. Nanoparti-cleswerealsoabletoimprovepharmacokineticparameters,such ashalf-life,whichwasincreasedupto10-foldafterintravenous administration.Liposomeswereusedascarriers forcommercial ginsengextract(Panaxquinquefolius)inordertoenhancethe intra-cellularantioxidant activity(Tsaiet al., 2012).These liposomes wereabletoincreasemembranepotentialafterH2O2damageand
decreaseapoptosis.The usefor foodproductis alsointeresting oncegrapemarcextract,forexample,wassuccessfullyassociated tonanoemulsionandhelpedtoreducetheoxidationofhazelnut paste(Spignoetal.,2013).
Oil-in-water nanoemulsions are also a good alternative for hydroalcoholicextracts.Bidoneet al.and Zorziet al.have pro-posedwaterinoilnanoemulsiontoassociateethanolicextractfrom
Achyroclinesatureioides(Bidoneetal.,2014b;Zorzi,2007).Thefirst studydescribedthedevelopmentoftopicalnanoemulsionforthe deliveryoftheantioxidantquercetinwhereasthesecondaddressed thetopical delivery of 3-O-methylquercetinas topical antiviral againstHSV-1(Bidoneetal.,2015).Associationwiththe nanoemul-sionimprovedtheantioxidanteffectofthefreeextract,probably duetothecontrolledreleaseofflavonoids(Bidoneetal.,2014b; Zorzi,2007).However,thehigheractivityoftheextract nanoemul-sionincomparisontoquercetinnanoemulsionwasattributedto thehigherskinretentionoftheflavonoidusingtheformulation withextract(Zorzi,2007).Theuseofnanoemulsionsubstantially increased3-O-methylquercetinpermeation(almost5-fold). Previ-oustechnologicaltreatmentoftheextract,suchasfreezedrying andspray-dryinginfluencedinthetotalamountofflavonoidsand theirreleasefromnanoemulsions(Bidoneetal.,2014b).Afewother reportsconcerningtheuseofnanoemulsiontoencapsulateVellozia squamataandPhyllanthusurinariawereproof-of-conceptanddid notbringresultsregardingthebiologicalactivityofthe formula-tions(Mahdietal.,2011;Quintãoetal.,2013).
Essentialoils
Essentialoilsareamixofseverallipophiliccompounds char-acterizedbyhighvaporpressure.Thecompositionvariesbutthe
majorityofessentialoilsarecomposedofterpenoidsand phenyl-propanoids.Especialattentionmustbegiventothepreparation techniquewhenassociatingessentialoilstonanostructures.As pre-viouslydescribed,manypreparationtechniquesrequiretheuseof hightemperatureororganicsolvents(thatmustbeeliminated). Duetothehighvaporpressureoftheessentialoilstheycanalso beremovedfromtheformulationinthisstep.Tocircumventthis problemafewsolutionscanbemadesuchasaddingamisciblefixed oilorusingnanosystems/techniquesthatinvolvelowtemperature andnoevaporationoforganicsolventafteradditionoftheessential oils.
Many essential oils present antimicro-bial/antiparasitic/insecticidal effect. In this sense, many formulations will have this objective. As can be observed in
Box4,liposomes,nanoparticles,nanoemulsionsandnanocapsules have beenproposedtoincorporateessential, each onewithits ownadvantagesanddrawbacks.
Nanoemulsions can associate thehighest amountof oil (up to 20%).Melaleucaalternifolia essential oilshowed lower mini-muminhibitory concentration against Saccharomyces cerevisiae,
Escherichiacoli,andLactobacillusdelbrueckii,afterincorporationto nanoemulsion(5%).Incombinationtofruitjuice,theproductshelf life couldbeextended due tothelow proliferationof microor-ganismandmaintenanceoforganolepticproperties(Donsìetal., 2011).Laterthissystemswereoptimizedbut,insteadof associat-ingthewholeessentialoil,isolatedvolatilecompoundswereused (carvacrol,limoneneandcinnamaldehyde),wherethesurfactants hadsignificanteffectoverthephysicochemicalpropertiesofthe nanoemulsionanditsantimicrobialactivity(Donsìetal.,2012). Similarly,associationofbasiloil(Ocimumbasilicum–88%estragole) intonanoemulsionhadadose-dependenteffectoverE.coli viabil-ity,wherenanoemulsion(0.6%ofoil)couldkill100%ofbacteria after45min(Ghoshetal.,2013).
Anotherimportantapplicationofessentialoils,especiallythose incorporatedtonanoemulsion,istheeffectoverinsects. Nanoemul-sionsofeucalyptoil(6%ofeucalyptoilwith17.73%eucalyptol) weretestedfor larvicidalactivityagainstCulexquinquefasciatus. Eucalyptoilhaditslarvicidalactivityagainstmosquitosignificantly improvedwhenusedasnanoemulsion.Theoilwasstabilizedwith polysorbate80andexhibit98%mortalityafter4hincubationwith 250ppmofoil.Forthesameconcentration,classicemulsionsonly achievedthisvalueafter24hofexposure.Evenaconcentration aslowas100ppmcouldresultinexpressivemortalityoflarves after6hincubation(around85%,versus20%foremulsionatsame concentrationandincubationtime)(Sugumaretal.,2014).Similar formulationsweretestedagainsttheredflourbeetle(Tribolium cas-taneum)howeverwithhighercontentofeucalyptolintheextract (66%)(Pantetal.,2014).Thisformulationwasabletokill85%of insectusing900ppm.Nevertheless,thiseffectdecreased25%after twomonths.Inordertoovercomethisdrawback,theauthors asso-ciatedtotheformulationtheaqueousextract ofJatrophacurcas
andPongamiaglabra,whichmaincompoundsarephorbolesters andkarinjin,respectively.Thisnotonlyincreasedtheinsecticidal effect(95%vs.85%)buttheeffectwasstablethroughtime.Inthis way,aftertheuseoftheaqueousextracttheIC50ofthemixtureof
extractsdroppedfrom5.4872mg/lto0.1646mg/l.Sakulkuetal. (2009) alsostudies theeffectof nanoencapsulated essential oil over insects.In thiscase, themosquitorepellentactivityof cit-ronellaoil(Cymbopogonnardus)wasevaluated.Thisterpenerichoil (41%d-limonene,40%citronellal)couldhaveacontrolledreleaseof
40–80%after48hwheninnanoemulsions.Therepellenteffectwas proportionaltotheviscosityoftheformulation,showingalower kineticreleaseparameter.
Box4:Differentessentialoilassociatedtonanosystems.HD,hydrodistillation;SC,supercritical.
Plant Obtention Partused Activecompound Activity System Reference
Origanum vulgare –a Leaves – – Nanoparticle(chitosan) Hosseinietal. (2013)
Lippiasidoides –a – Terpenes(thymol) – Nanoparticle(chitosan,
cashewgum)
Abreuetal.(2012)
Nanoparticle(alginate, cashewgum)
deOliveiraetal. (2014)
Eucalyptus camaldulensis
HD Leaves Terpenes(eucalyptol) Microbicide Liposomea
(phosphatidyl-choline, cholesterol)
Moghimipouretal. (2012)
Artemisia arborescens
HD Aerialparts Terpenes(camphor,
-thujoneand chamazulene)
Antiviral Liposomea
(phosphatidyl-choline, cholesterol,stearylamine)
Sinicoetal.(2005)
Atractylodes macrocephala
SCCO2 Rhizomes Sesquiterpenes
(Atractylone)
– Liposomea(lecithinand
cholesterol)
Wenetal.(2010)
Schinusmolle HD Woods Essentialoil(notspecified) Trypanocidal activity
Nanoemulsion(Tween®80,
Span®80)
Baldisseraetal. (2013)
Carapaguianensis –a Seeds Essentialoil(notspecified) Trypanocidal
activity
Nanoemulsion(Tween®80,
Span®80)
Baldisseraetal. (2013)
Eucalyptus globulus
HDa – Eucalyptol Insecticidal Nanoemulsion(Tween®80) Sugumaretal. (2014)
Ocimumbasilicum –a Leaves Phenylpropene(estragole) Microbicide Nanoemulsion(Tween®80) Ghoshetal.(2013)
Eucalyptus globulus
+
Jatrophacurcas
+
Pongamiaglabra
HDa
+ Aqueous + Aqueous
– Eucalyptol
Phorbolesters
Karingin
Insecticidal Nanoemulsion(Tween®80) Pantetal.(2014)
Stenachaenium megapotamicum
HD Flowersand leaves
Terpenes(fokienol,thymol andothers)
Microbicide Nanoemulsion(Tween®80) Daniellietal.(2013)
Copaiferamultijuga Direct Trunks Diterpenicacidsand sequiterpenes (-caryophyllene)
Topical anti-inflammatory
Nanoemulsion(Tween®80,
Span®80)
Diasetal.(2012) Diasetal.(2014)
Cymbopogon nardus
–a Leaves Terpenes(limoneand
citonellal)
Repelent (insect)
Nanoemulsion(glycerol, cetearylalcohol/cocoyl glucoside)
Sakulkuetal.(2009)
Melaleuca alternifolia
–a Leaves Terpenes(unspecified) Microbicide Nanoemulsion(lecithin,
palmorsunfloweroil)
Donsìetal.(2011)
HDa Leaves Terpenes(terpinen-4-ol,
andterpinenes)
– Nanoemulsion Nanocapsule (lecithin,Tween®
80, Span®®80,PECL)
Floresetal.(2011)
Jasmine –a –a Terpenes(linalool) – Nanocapsule(gelatin,
arabicgum,Tween®80,
Span®80)
Lvetal.(2014)
aCommercialproduct.Span®80,sorbitanmonooleate;Tween®80,polyoxyethylenesorbitanmonooleate.
essentialoilsassociatedtonanoemulsionswastestedusingpureoil (Baldisseraetal.,2013).Adose-dependentreductioninthenumber ofparasiteswasobservedafter1h,regardlesstheassociationtothe nanoemulsion.Nevertheless,themortalitywasmorepronounced fornanoemulsionthatcouldevenkill100%oftrypanosomesafter 1hwhereasfreeoilrequiredupto6hforthesameeffect.
Eventhoughhighpressure homogenizationand ultrasonica-tionarethemostdiffusedmethodsforessentialoilnanoemulsion preparation,thereareafewreportsconcerningspontaneous emul-sificationandphaseinversiontemperature.Diasetal.(2012,2014)
comparedhighpressurehomogenizationandspontaneous emul-sificationinordertopreparecobaibaoilnanoemulsion.Copaiba oil(Copaiferamultijuga)isanoleoresinthatcanbedirectedcollect fromthetreetrunk.Theoilisacomplexmixtureofterpenes,being
-caryophyllene themost abundant one, and chemical marker fortheoil.Spontaneousemulsificationrequiresastepofsolvent removalby reducedpressure that caneventuallyremove some components of the volatile essential oil. Because of its partial resinousnature,thelossduringpreparationwasminimal,beingthe finaloilcontenthigherthan90%.Nevertheless,thisisnotalwaysthe case,asobservedforStenachaeniummegapotamicumessentialoil nanoemulsion(Daniellietal.,2013).Thisfokienolandthymol-rich oilwasassociatedtonanoemulsionsintendedtobeusedagainst
Box5:Differentorganicorunspecifiedextractsassociatedtonanosystems.
Plant Solvent Partused Activecompound Activity System Reference
Diospyroskaki Ethylacetate Leaves Phenoliccompound (quercetin)
Cardiacandvascular conditions
Nanoemulsion
(Cremophor®EL,Labrafil®
M1944CSandTranscutol®
P)
Lietal.(2011)
Manilkara subsericea
Hexanefraction ofethanolic
Fruits ␣-and-amyrinacetate Insecticidal Nanoemulsion(Tween®80,
Span®80,ODM)
Fernandesetal.(2014)
Pterocaulon balansae
n-Hexane Leaves Coumarins(esculin) Antifungal Nanoemulsion(lecithin, MCT)
Viannaetal.(2011)
Curcumacomosa n-Hexane Rhizomes Diarylheptanoid Phytoestrogenic Nanoemulsion(Tween®®
60,PEG1000,oliveoil)
Suetal.(2013)
Passiflora serratodigitata
Fractionof hydroethanolic (60%ethanol)
Leaves – Antiulcerogenic Nanocapsule(PECL,MCT, Span®80)
Strasseretal.(2014)
Ginkgobiloba –a Leaves Terpenes(ginkgolides
A,B,Candbilobalide)
– Nanoparticle (mPEG–PLGA–mPEG)
Hanetal.(2012)
Greentea –a Leaves Catechins Microbicide Nanostructuredlipidcarrier
(cetylpalmitate,glyceryl stearate,grapeseed/St. John’swort/seabuckthorn oil.
Maneaetal.(2014)
Hypocholesterolemic Nanoemulsion(cholesterol, cetylphosphateMCT)
Kimetal.(2012)
aCommercialextract.
Cremophor®EL;polyethoxylatedcastoroil;Labrafil®M1944CS,oleoylmacrogol-6glyceride;MCT,mediumchaintriglycerides;ODM,octyldodecylmyristate;PECL,
poly(epsilon-caprolactone);PEG,polyethyleneglycol;PLGA,poly(lactic-co-glycolicacid);Span®80,sorbitanmonooleate;Transcutol®P,diethyleneglycolmonoethyl
ether;Tween®
,polyoxyethylenesorbitanmonostearate(60)ormonooleate(80).
oilintoliposomes prepared byrapid expansionof supercritical solutions(Wenetal.,2010).Thekeyfactorsfoundwerepressure, temperature and amount of co-solvent (ethanol).Nevertheless, evenwiththeprocessoptimization,asignificantlossofessential oilwasobservedovertime.
Duetothehighvolatility,theamountofessentialoilcan even-tually bereduced over time. For that, nanocapsules canbe an alternativetoavoidoillossthoughtime.Afterincubationat37◦C
theoilcontentofMelaleucaalternifoliareduced60%in nanoemul-sionand40%innanocapsules(Floresetal.,2011).Inasimilarway, Lvetal.haveproposedgelatin/arabicgumnanocapsulesasa heat-resistantsystemforessentialoilassociation(jasmineessentialoil) (Lvetal.,2014).Inthecaseofindol;however,areductionof95%in thenanocapsulescouldbeobservedafter5hathightemperature (80◦C).
Therearefewreportsconcerningtheassociationofessentialoil topolymericnanoparticles.In thissense,chitosan nanoparticles wereused,asproposedbyHosseinietal.(2013)toassociate Orig-anumvulgareessentialoil.However,thisapproachresultedinlow associationefficiency(5–25%)andloadingcontentaround1.3–2%, asdeterminedbyUV–VISspectroscopy.Inasimilarway,the asso-ciationefficiencyofchitosan/cashewgum(Abreuetal.,2012)and alginate/cashewgumnanoparticles(deOliveiraetal.,2014)was notveryhigh,beingaround40–70%and21–48%,respectively.
Otherextracts
Extractspreparedwithsolventswithlowerdielectricconstant arealsocommonandpresentawidevarietyofsecondary metabo-lites.Ethyletheranddichloromethaneareusedforextractionof lipophilicsubstancessuchaslipid,essentialoils,waxesand alka-loid in free base form. Also, mixture of different solvent may allowanoptimalextractionoftheactivecompounds.Currently ourresearchgroupstudiestheapplicationofthecoumarinrich
n-hexaneextractofPterocaulonbalansae,especiallyforantifungal andantiparasiticapplications.Thislipophilicextractisassociated withnanoemulsioninordertoincreasethesolubilityand activ-ityofsuchcoumarins(Panatieri,2015;Viannaetal.,2011).Box5
showsdifferentsolvent,mainlywithhigherlipophilicity,andtheir incorporationintonanosystems.
Similartoessentialoils,hexaneextractsshowahigh lipophilic-itymakingthemsuitabletobeassociatedintosystemswithhigh oilcontent. Nanoemulsionsallow highassociation efficiencyof theactive compound. Curcuma comosan-hexaneextractis rich in diarylheptanoid phytoestrogens. The use of nanoemulsions showedimportantassociationefficiencyoftheactivecompound (around75%)andanincreasein10-foldintheintestinalabsorption inWistarrats(Suetal.,2013).
Ethyl acetate fractions from Passiflora serratodigitata
hydroethanolic extract (60% ethanol), as well as the hydroal-coholic extract itself, were associated to PECL nanocapsules in ordertoimprovetheantiulcerativeeffect(Strasseretal.,2014). Eventhoughpromising,thelackofproperexperimentalcontrol andstatisticalanalysismakesdifficulttoconcludetherealeffects ofencapsulation.Furtherfractioningshowedthattheethylacetate fraction was the most active. Unfortunately, the authors did notmake acharacterizationoftheextractanditsfraction,only expressingthetotalflavonoidcontentusingquercetinasstandard. Oncetheflavonoidcontentoforiginalhydroethanolicextractand ethylacetatewerethesame,theexplanationforthisactivityisyet tobebetterunderstood.
Hexane-soluble fraction from ethanolic crude extract from
Manilkarasubsericeafruitswasformulatedinnanoemulsionaimed againstcottonpest(Dysdercusperuvianus)(Fernandesetal.,2014). Thisapolarfractionwasrichintriterpenes,mainly␣-and-amyrin acetate.Theformulationwasabletoinducemortalityin25%ofD. peruvianus.Eventhoughtheinsecticidalactivitywaslow,noeffect against acetylcholinesteraseormortalityin miceinduced high-lightingtoapossiblelowtoxicity. Nocomparisonwiththefree extractwasmade.
efficiency around 80%. In this way, a controlledrelease of the fourcomponentswasobservedbothinvivoandinvitro, show-ingacumulativereleaseof60%after24h.Concerningcommercial greenteaextract,bothhypocholesterolemic(Kimetal.,2012)and antibactericidal(Maneaetal.,2014)weretestedusing nanoemul-sionand nanostructured lipid carriers, respectively (Kim et al., 2012). For its hypercholesteremic activity, only LDL receptor expression was higher for the nanosystems in comparison to freeextract.Antimicrobialactivitysuggeststhatgreenteaextract nanoemulsioncouldbeutilizednotonlyasantioxidantbutalsoas avaluablenaturalsourceofantimicrobialagent.
Finalconsiderations
Theuseofnanonotechnology-basedsystemshasbeen grow-inginthepastyears andisrelativelynew,asconfirmedbythe recentliterature.Basedontheresultspresentedhere,thereare clearlytwomajoradvantages ofusingnanocarrierstoassociate complexextracts,basedonthebiologicalresponses:(i)increasing theantioxidanteffectduetothecontrolledkineticreleaseof antiox-idant,and(ii)increasetheactivityagainstmicroorganism,parasites andinsects.Theuseofsystemssuchasliposomes,nanoparticles andnanoemulsionscanbringotherbenefitsandstillhasabright pathahead.Nevertheless, thispath islongand dependsonthe advancesofknowledgeonthephytochemicalcompositionand bio-logicalactivityofthemedicinalplantextracts,aswellasonthe physicochemicalcharacterizationofthenanostructureddelivery systemscontainingthesecomplexesmatrices.
Authorcontributions
GZsearchedtheliteratureindifferentdatabases,collectedthe main data, and draftedthe first version of the manuscript. All authorssuggestedtheoutlineofthearticle,andparticipatedinthe selectionofthearticlesusedinthisreview.Allauthorsparticipated inthewriting,editing,andrevisingthefinalversionofthearticle.
Conflictsofinterest
Theauthorsdeclarenoconflictsofinterest.
Acknowledgements
TheauthorswanttothankBrazilianFederalAgencyforthe Sup-portandEvaluationofGraduateEducation,andNationalCouncil forScientificandTechnologicalDevelopmentforfinancialsupport. GZwishestothanktheNationalCouncilforScientificand Tech-nologicalDevelopment(ProgramaJovensTalentos–grantnumber 028/2012)forhispostdoctoralgrant.
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