OrIgInaI

lNTHODUCTlON
Chomicalbacoricidosholpoprosorvoproducsas
divorsoascuingluids,oodandbovoragos,cos-
moicsandpharmacouicalormulaions,andprovido
proocionagainsspoilagoinawidorangooindus-
rialandonvironmonalapplicaions.
Amonghoclassicalcaionicsuracans,quaor-
naryammoniumsalsOUATsarohomosusoul
anisopicsanddisinocans.CaionicOUAT8aroac-
ivoagainsmombranosFuhrhopandWang,?004
andlysosphoroplassandprooplasssuspondodin
sucrosoDonyor,1995,Kana.awa,oal.,1995.
Caionic agons hypohoically roac wih
phospholipidcompononsinhocyoplasm mom-
brano,horobydisrupinghomombranoandcausing
prooplaslysisundorosmoicsrossCabral,199?.
Gomini quaornary ammonium compounds
gomini-OUATs compriso wo oriary amino-
quaorni.od compounds linkod wih hydrocarbon
chains.ThosrucuroogominiOUATsisuniquoand
hoy havo groaorsuraco aciviy and highor
animicrobialaciviioshan convonionalOUATs.
8udioshavorovoalodhoirphysicochomicalpropor-
iosFosonoal.,1999andhoirgroaoranibaco-
rialpooncyhancorrospondingconvonionalOUATs
Di.oal.,1994,MongorandKoipor,?000.Wopro-
viouslysynhosi.odgomini-OUATswihwocaionic
pyridiniummoioiosanddiscussodhoiranimicrobial
aciviiosandcharacorisicsKouraioal.,?006,
Maodaoal.,1998and1999,Oka.akioal.,1997,
8hiraioal.,?005and?006.Gomini-OUATshavoa
widoranimicrobialspocrum againsbohgram-
TheModeoftheAntIfungaIActIvItyof
GemInI-PyrIdInIumSaItagaInstYeast
AKlHlHOSHlHAl,TOMOKOSUMlTOMO,MAYUKOKUHlMOTO,
HlDEAKlMASEDA,ANDHlHOKlKOUHAl

/oa/^/c///c/c/ca/5c/^ca^J1c|^c/c,//cs,s/s/^/^/^/^s///a/c/1c|^c/c,
a^J5c/^c1||^/s//,c/1c/as|/aCaJaa/5c|cc/?1//^a//csa^//ac|c
1c/as|/aZZ0b00'aoa^
Focoivod14July,?008/Accopod6ovombor,?008
ThegemInIquaternarysaItgemInI-QUATcontaInIngtwopyrIdInIum resIduespermoIe-
cuIe,3,3'- 2,7-dIoxaoctanebIs1-decyIpyrIdInIumbromIde3DOBP-4,10,exertedfungIcIdaI
actIvItyagaInst5accharomycescerev/s/aeaccompanIedbyrespIratIonInhIbItIonandthe
cytopIasmIcmaterIaIIeakageofATP,magnesIum,andpotassIumIons.WeprevIousIyfound
thatgemInI-QUATexertedbacterIocIastIcactIonagaInstEscher/ch/aco//bycausIngtherapId
andabundantIeakageofturbIdmaterIaIsfrom theceIIs.lnaddItIon,thefIrststageofthe
bacterIocIastIcactIonwastheIeakageofmagnesIumIons,outermembraneproteInE,ATP,
andIIpopoIysaccharIdes.Here,weInvestIgatedhow thegemInI-QUAT3DOBP-4,10exerts
fungIcIdaIactIonagaInst5cerev/s/ae.TheresuItsshowedthatthat

0.4M3DOBP-4,10
stoppedrespIratIonandthat

3.0,1.0and1.0McausedtheIeakageofcytopIasmIccompo-
nentsATP,magnesIumandpotassIumIons,respectIveIy.ScannIngandtransmIssIoneIectron
mIcrographsshowedapreservedceIIwaIIstructure,whereasIntraceIIuIarorganeIIeswere
destroyedInceIIsIncubatedwIth3DOBP-4,10.WepostuIatedthat3DOBP-4,10exertsIts
fungIcIdaIactIonagaInst5cerev/s/aenotthroughceIIwaIIdestructIonandproteInIeakage,
butratherbypenetratIngtheceIIwaIIanddIsruptIngthemembranesoforganeIIes.
/,ncJsAniungalacion/Aniungalaciviy/Gominipyridiniumsal/Yoas.
ioconlrolScience,!00,Vol.14,No.1,1!0

csoc^J/^aa/|c1/-10b0Z40/a-1
0b014
nogaivoandgram-posiivobacoriaandungiand
aromorooocivohanconvonionalOUATs.Tho
animicrobialaciviiosohogomini-OUAT4,4'- 1,6-
hoxamohylonodihiobis1-ocylpyridiniumiodido
4DTEF-6,8aronoonlywidorandmoropoonhan
hosooconvonionalOUATs^-dodocylpyridinium
iodidoandbon.yldimohyldodocylammonium chlo-
rido,buaroalsoindopondonopH,omporauro
andmolocularhydrophobiciy,unlikohoaciviioso
OUATsOka.akioal.,1997,Maodaoal.,1998.
8umiomooal.?004rovoalodha4DTEF-6,8
causosholoakagoomagnosiumionswhichsabi-
li.ohoouormombranoobacorialcolls,andha
ATFloakagoromhocollsandrospiraoryinhibiion
occur booro bacorioclasic acion owards
/sc|/c|/acc//FO1?713/cc//FO1?713.This
suggosodhagomini-OUATsconormoaspociic
bacoricidalmochanismunlikohousualOUATsow-
ingohoiruniquogominaosrucuro.Furhormoro,
immunoolocronmicroscopyhasshownha4DTEF-
6,8causosholoakagooouormombranoprooinE
OmpEaamuchloworconconraionhanhocrii-
calvosiculaionconconraionCVC,suggosing
haOmpEisinvolvodinhoma|orbacorioclasicac-
ionohisgomini-OUAT8umiomooal.,?006.
FogardinghomochanismoOUATacionagains
ungi,somopolymoricOUATsinducoholysiso
sphoroplasso5a//aacsc^s,bunohosoo
a^J/Ja a///ca^s Codling o al., ?003.
8ubsoquonly, ho oocs o ho OUATs,
diocadocyldimohylammonium bromidoDODAE
andhoxadocylrimohylammoniumbromidoCTAE,
ovaluaod asadsorpion isohorms,collviabiliy,
olocrophoroicmobiliyandungalcolllysis,indicao
ha ho criicalphonomonon doormining ho
aniungaloocoOUATsisnocolllysisburahor
rovorsalohocollsuracochargoromnogaivoo
posiivoVioiraandCarmona-Fiboiro,?006.Thoso
rosuls woro nodoocod in ulravioloUV-
adsorbingmoloculossuchasnucloicacidsandpro-
oinsinhocollsupornaaninhoprosoncoo0.1or
10mMDODAEorCTAE.Tholoakagoopoassium
ionsandUV-adsorbingmaorials,aswollashoinra-
collularproducionoroacivooxygonspociosin-
ducod by anibioics such as niphimycin and
amphooricinE,havobooninvosigaodwihrospoc
ohomodoo5acc|ac,csc/s/acolldoah
akayama oal.,?00?,Usukioal.,?006.
Howovor,homodoohoaniungalacionogomini-
OUATshasnoyoboonolucidaod.Horo,woound
ha3,3'- ?,7-dioxaocanobis1-docylpyridinium
bromido3DOEF-4,10inducod holoakagoo
cyoplasmiccompononsincludingATF,magnosium
andpoassiumionsrom5c/s/a,andinhibiod
collularrospiraion.Woalsoobsorvodcollsinho
prosoncoo3DOEF-4,10usingscanningandrans-
missionolocronmicroscopyodoorminohoooc
o3DOEF-4,10onhocollwallandinracollular
componons.Tho3DOEF-4,10synhosi.odbyour
grouppossossosabroadanimicrobialspocrumand
ahighanimicrobialaciviyKouraioal.,?006,and
moroovorshowshousoulnossasadisinocanina
waorsysomowingohohighsolubiliyovor500%
w/wohoproduc.Forhoirsimo,oourknowl-
odgo,his sudy olucidaod ho acion oho
aniungalaciviyo3DOEF-4,10asgomini-OUATs
againsyoas,suggosinghooocivoinacivaiono
pahogonicyoasssuchasa^J/Jaspociosbyho
applicaionohiscompound.
MATEHlALSANDMETHODS
ChemIcaIsandmaterIaIs
Wosynhosi.odhogomini-OUAT,3DOEF-4,10
Fig.1,Kouraioal.,?006.Ohormaorialsandroa-
gonsusodinbiologicaloxporimonsworopurchasod
romcommorcialsourcos.
CuItIvatIonandpreparatIonoftheyeast
Thoyoas5c/s/aEFC10?17wasinocu-
laodino5mlo8abouraudliquidmodiumOxoid,
andshakona?8or?4h.Thocollsusponsionwas
honincubaodin100mloroshliquidmodiuma
?8 unilhomid-oxpononialphasowasroachod.
Thosusponsionwasharvosodbyconriugaiona
4?50gor10mina4,washodwicowihsorili.od
physiologicalsalino,andhonrosuspondodinsorilo
ico-cooloddisillodwaoroad|ushocolldonsiyin
alloxporimons.
MeasurementofrespIratoryInhIbItIon
Thoabiliyo3DOEF-4,10oinhibirospiraionwas
doorminodasdoscribod8umiomooal.,?004.An
oxpononial-phasocollsusponsion110
5
CFU/ml
conaining0.?%w/v/+-glucosoasarospiraory
subsrao,wassauraodbyoxygonusingaairpump,
sorili.odbyilraionhrough0.?mcollulosoaco-
aoADVATEC,and pourod ino a 100-ml
A.SkAFTAL. 14
FlG.1.Chomicalsrucuroo3,3'- ?,7-dioxaocanobis1-
docylpyridiniumbromido3DOEF-4,10.
branchodErlonmoyorlaskoquippodwihadissolvod
oxygonolocrodoDO8F,Horibaandsirroda?5
inawaorbah.Whonhodocroasingraoodis-
solvodoxygonroachodasoadysao,3DOEF-4,10
avariousconconraionswasaddodoholask.Tho
amounodissolvodoxygonwasrocordodor30min
a5mininorvals.8urvivingcollsworodoorminodby
incubaingsopwisodiluionsoalvolumo,0.1mlin
8oyboan-CasoinDigosErohconainingLocihinand
Folysorbao808CDLFbroh,ihonFharmacouical
Co.Ld.on plaosconaining 8abouraud agar
ihonFharmacouicalCo.Ld.a?8 or48h.
AnaIysIsofceIIuIarmaterIaIs
Thomodooaniungalaciviywasdoorminodby
analy.inghoamounoATF,magnosiumandpoas-
siumionshaloakodromcolls.
WomoasurodhoamounoATFhaloakodrom
collsasollows.Acollsusponsion?10
6
CFU/ml
and3DOEF-4,10,bohinsorilodisillodwaor,woro
soparaolywarmoda?8or5minandhon3mlo
oachworomixod.Thoconrolcomprisod3mloacho
collsusponsionanddisillodwaor.Aor?,5,10,15
and30min,0.4-mlporionsohomixuroworo
passodhroughamombranoilor0.45m,collu-
losoacoao,ADVATEC.Thoroaor,0.1-mlpor-
ionsworodiluod10-oldwihATFassaydiluion
buorLuciorH8so,KKKOMA,andhon0.1ml
ATFoxracionroagonwasaddod,ollowodby0.1
mloATFluminosconcoroagon.ThoamounoATF
haloakodrom hocollswasmoasurodusingan
ATF analy.orELF-?01,Aloka.LoakodATF is
shownashoamounloakodporsurvivingcolland
doorminodbyusingasimilarmohodohado-
scribodabovo.
Womoasurodhoamounsoroloasodmagnosium
andpoassium ionsasollows.Acollsusponsion
500mland3DOEF-4,10,warmodasdoscribod
abovo,woromixodina1.1raio,incubaoda?8
or?min,andhonrapidlypassodhroughamom-
brano ilor 0.45 m, colluloso acoao,
ADVATEC.Thoilraowasconconraodinvacuo
andro.ona-80.Thodriodilraowasad|usodo
50mlinamoasuringlaskwihsupordisillodwaor
andhon1mloconconraodniricacidwasaddod.
Thoconconraionsoloakodmoalionsinhoacid
soluionworocalculaodusinganCFanalysisappa-
raus8F8-4000,8oikonsrumonsnc..
EIectronmIcroscopy
Colls aho mid-oxpononialphaso?10
6
CFU/mlworomixodwih6.0M3DOEF-4,10or?
min,andobsorvodusinga8-4700scanningolocron
microscopo8EM,Hiachiasdoscribodby8hiraio
al.?007.nbrio,samplosdriodinabsoluoohanol
woroimmorsodin50%v/v/-buylalcoholor1h,
honin100%/-buylalcoholovornigh,driod/^acac
andcoaodwihgoldusinganionspuoringdovico
E-10?0,Hiachi.
Collsincubaodwih3DOEF-4,10or?minworo
alsoobsorvodusingaJEM-1010ransmissionoloc-
ron microscopoTEM,Japan Elocron Opics
LaboraoryCo.Ld.Collsharvosodbyconriugaion
6400g,5minwororosuspondodin6mlosorilo
disillodwaor,6mlo0.?M phosphaobuorpH
7.?conaining4%v/vgluaraldohydowasaddod,
andhonhosusponsionwasincubaoda4or?h.
Thosusponsionwaspolloodbyconriugaionasdo-
scribodabovoandhosupornaanwasdocanod.
Thopollowasrosuspondodin6mlo0.?Mphos-
phaobuorpH7.?,andhiswashingmanipulaion
wihbuorwasropoaodhrooimos.Foassiumpor-
manganao10ml,1.5%v/vwasmixodwihho
pollo,andhosusponsionwasincubaoda4 or
18h.Thoprocipiaowashonwashodwihsorilo
disillodwaorunilhosupornaanobainodbycon-
riugaion romainod cloar.Tho procipiao was
hoaoda60,mixodwih3%w/vagaroso
8andardLow-m,Eio-FadLaboraorios,andho
rosulingsolidmasswascuinohinslicosand
slowlyshakonin30% acoonoor3haroomom-
porauro.Thosolidwasdohydraodwihasorioso
50,70,90,95and100%v/vacoonoor3hand
honsubsiuodwihasorioso?5,50,75and100%
v/vrosininacoonoor?h.Thooriginalsoluiono
rosincomprisod4.79goopoxyrosinombodding
roagonOuool-653,isshinEM Co.Ld.,6.9-ml
vinylcyclohoxono dioxidoEFL 4?06, Chiyoda
Junyakunc.,19.09gonononylsuccinicanhydrido
8A EM,Chiyoda Junyaku nc.,and 0.15 g
dimohylaminoohanol8-1,ChiyodaJunyakunc..
Thosamplo,comploolysubsiuodwihhooriginal
rosinsoluion,wasransorrodoaboamcapsulo414
isshinEMCo.Ld.andhoaoda60or48ho
polymori.ohorosin.Ulrahinsocionsworopro-
parodusinga?088UlramicroomoVAmorsham
Eioscioncos,andhospocimonsworosainodwih
uranylacoaoandloadciraoorobsorvaionby
TEM.
HESULTS
lnhIbItIonofrespIratIon
Thogomini-OUAT,4DTEF-6,8inhibisrospiraion
in/cc// 8umiomooal.,?004.Horo,wooxaminod
how3DOEF-4,10aconconraionso0.3-1.5M
aocshorospiraoryunciono5c/s/aEFC
10?17inasusponsionconaining0.?%glucoso.
ANTFUNCALACTNFCFMN-QUAT 15
Figuro?Ashowshodissolvodoxygonconconra-
ioninhocollsusponsiona5mininorvalsaoradd-
ing 3DOEF-4,10.Tho amounooxygon also
docroasodinhoconrolovorimoinhoprosoncoo
0.3-0.4M 3DOEF-4,10.Aaconconraiono

0.5M,horaoooxygondocroasosigniicanlya-
porodo,andhoallindissolvodoxygonwasalmos
undoocabloa1.5M.Figuro?Eshowshooxygon
consumpionohoyoasincubaodwih0.3o1.5
M3DOEF-4,10or30min.Thorosulsconirmod
ha>0.4and0.5M3DOEF-4,10sarodoinhibi
andoallyinhibiodrospiraoryuncion,rospocivoly.
LeakageofATP,magnesIumandpotassIumIons
ThoamounooxracollularATFwasmoasuroda-
orincubaingcollswih1.0o6.0M3DOEF-4,10
or0-30minFig.3A.Thoconconraiono
oxracollularATFa30minwasalmosundoocablo,
slighandobviouslyolovaoda3DOEF-4,10concon-
raionso1.0-1.5,?.0o3.0and4.0o6.0M,ro-
A.SkAFTAL. 16
FlG.2.ATimo-coursochangosoDOduringincubaiono5c/s/awih3DOEF-4,10.8ymbols.conrolshadodcir-
clos,0.3Munshadodcirclos,0.4Mshadodrianglos,0.5Munshadodsquaros,0.7Munshadoddiamonds,
1.0Mshadoddiamonds,1.5Munshadodrianglos.Thoarrowindicaoshoaddiiono3DOEF-4,10.EThooxy-
gonconsumpionshadodcircloso5c/s/aroaodwih3DOEF-4,10or30min.Thoarrowindicaoshocriicalcon-
conraionoinhibirospiraoryuncion0.4M.
FlG.3.AFromoivooocso3DOEF-4,10onholoakagooATFrominaccolls.8ymbols.1.0Munshadodcirclos,
1.5Munshadodrianglos,?.0Munshadodsquaros,3.0Mshadodcirclos,4.0Mshadodrianglos,6.0M
shadodsquaros.EThoooco3DOEF-4,10onholoakagooATFrom5c/s/aduring?minincubaion.Thoar-
rowindicaoshocriicalconconraionoinducoATFloakago3.0M.
spocivoly.Figuro3EshowshahoamounsoATF
roloasodaor?mininhoprosoncooincroasing
conconraions o3DOEF-4,10 rosulod in wo
sraighlinos.Thosorosulsshowhahocriicalcon-
conraiono3DOEF-4,10roquirodorATFloakagois
3.0M.
Figuro4AandEshowsmagnosiumandpoassium
ionloakagoporcollaorincubaionwih1.0o6.0
M3DOEF-4,10.Wodoocod0.410
14
g/collo
oxracollularmagnosiumionswhonhocollsworoin-
cubaodinsorilodisillodwaorFig.4A.Tho
amounomagnosiumandpoassiumionloakagoin-
croasodrom0.410
14
o0.8910
13
g/collFig.
4Aandrom1.?10
13
o13.410
13
g/collFig.
4Einhoprosoncoo1.0-6.0and0-3.0M
3DOEF-4,10,rospocivoly.Howovor,hopoassium
ionloakagoaconconraions3.0M roachod
sauraionlovols.
EIectronmIcroscopy,SEMandTEM
Woconirmodhoaniungalaciono3DOEF-4,10
agains 5 c/s/a by 8EM obsorvaion.
MicrographsinFig.5AandEshowinaccollsand
collsincubaodwih6.0M3DOEF-4,10or?min.
Thocollsuracoinhoprosoncoo3DOEF-4,10was
scarcolyaocodandromainodalmossmoohcom-
parodwihinaccolls.Onhoohorhand,TEMro-
voalodobviousinracollularchangosinhoprosonco
o3DOEF-4,10Fig.6CandD.Organollarsruc-
urossuchashocollwall,collmombrano,nuclous,
miochondriaandvacuolosworoobviousininac
collsFigs.6AandE.nconras,disinccollmom-
branosworolosincollsincubaodwih3DOEF-4,10,
alhoughhocollwallwashardlyaocodFig.6C
andD.Furhormoro,homombranosohonuclous,
miochondriaandvacuoloshadcollapsod,rondoring
hosoorganollosindisinguishablo.
ANTFUNCALACTNFCFMN-QUAT 17
FlG.4.Fromoivooocso3DOEF-4,10onholoakagoomagnosiumionAandpoassiumionErominaccollsdur-
ing?minincubaion.
FlG.5.8canningolocronmicrographocollsroaodwih3DOEF-4,10a0Mconrol,Aand6.0ME.Thowhiobars
roproson5m.
DlSCCUSlON
Thissudymoasurodhoinhibiionocollularrospi-
raion,andmorphologicallyobsorvodhoamouno
ATF,magnosium andpoassium ionshaloakod
rom collsusing8EM andTEM oclariyhowho
gomini-OUAT3DOEF-4,10oxorsaalaniungalo-
ocsagains5c/s/a.
Foconsudioshavorovoalodhagomini-OUATs
such as4DTEF-6,8 carrying wo ammonium or
pyridinium caionmoioios,dosroyho/cc//coll
srucurohroughholoakagooinracollularATF,
LF8,magnosiumionsandOmpE,hoinhibiionrospi-
raoryon.ymosandvosiculaion8umiomooal.,
?004.
Woolucidaodhocriicalconconraiono3DOEF-
4,10roquirodoinhibicollularrospiraionFigs.?A
andE.Thorosulsshowodhahocriicalrospira-
oryinhibiionconconraiono3DOEF-4,10was

0.4M,ascollularoxygonconsumpionwasimmodi-
aolyroducodFig.?E.Thissuggosodhaho
gomini-OUATpormoaodhocollwallandmombrano,
honinhibiodrospiraoryon.ymoslocali.odinhomi-
ochondriaand/ordosroyodorganollomombranos.
Woalsoassumodhahogomini-OUATprovokoda
changoinhopormoabiliyohocyoplasmicmom-
branooroachhoinsidoohocoll.
Thocriicalgomini-OUATconconraionroquirodo
inducoaalATFloakagowas3.0MFig.3Aand
E,whichwasabou8-oldhighorhanhocriical
rospiraoryinhibioryconconraion.Thisindingim-
pliodhaholoakagoolowmolocularwoighATF
mighnobocausodbyhodisinograionohocoll
wallandcollmombranoaorhogomini-OUAThas
ponoraodhocoll.Thais,hosrucuroohocoll
walland/orcyoplasmicmombranosoomobomain-
ainodrogardlossoponoraionbyhogomini-OUAT.
Thisnoionwassupporodbyhorosulsomoasur-
inghoconconraionsosovoralprooinshaloakod
rom hocollsdaanoshown.Howovor,hoso
worolargoprooplasmicandcyoplasmiccompo-
nons.Thoprooinconconraioninducodooxi
A.SkAFTAL. 18
FlG.6.TransmissionolocronmicrographoinaccollsA,andocollsroaodwih3DOEF-4,10a6.0MC.Fhoos
EandDarohoonlargomonohoparohocollonclosodbyhosquaroinphooAandC,rospocivoly.8mallloorsin
hoigurosindicaohonuclousa,vacuolob,miochondriac,cyoplasmicmombranod,andcollwallo.
sonicaodinaccollsposiivoconrolsandmoas-
urodusingbicinchonicacidwasdoinodas100%.
Aorincubainginaccollswih6.0M3DOEF-4,10,
horaiooprooinshaloakodromhocollswas
4.7%.Thisphonomononwassimilarohainho
casoohoabsoncooUV-absorbingsubsancosin
5c/s/a aorincubaion wih 1mM CTAE
VioiraandCarmona-Fiboiro,?006.Thominimum
ungicidalconconraionMFCohogomini-OUAT
agains5c/s/awas6.3Mdaanoshown.
Thosorosulsindicaodhahocolldoahcannobo
aribuodolysisinducodbydisorgani.aionoho
collwallandcyoplasmicmombrano,rosulinginpro-
oinloakago.Thoovidoncoprosonodhorosuppors
honoionhahocollsrucuroisprosorvodaorox-
posuroo6.0M3DOEF-4,10Figs.5and6.Tho
absoncooanobviousoocnogaoshopossibiliy
hahoirssioogomini-OUATacionishocoll
suracoandrahorindicaoshahoargocomprisos
hoinracollularcomponons.Onhoohorhand,
whon/cc//wasincubaodwihhogomini-OUAT,
4DTEF-6,8or?min,homochanismsoATFand
prooinloakagodiorodrom haindicaodbyho
prosonoxporimonaldaa.Fospiraoryinhibiionand
ATFloakagorom/cc//collsaroossoniallysimula-
noousaorhoogrossomagnosiumions,andhon
prooinloakagowasinducodbymombranodisrupion
aribuabloohoacionohogomini-OUATargo-
inghocollsuraco.Morphologicalobsorvaionso/
cc//by8EMandTEMhavoalsoclariiodha4DTEF-
6,8causoslargoporosoormonhocollsuraco
8umiomooal.,?004.Consoquonly,woound
horohahoirssiooacionbygomini-OUATs
againshoyoas5c/s/adiorsrom ha
agains/cc//,rosulingromhodioroncoinhocoll
suracosrucurobowoonhocolls.
Wooxaminodmagnosiumandpoassiumionloak-
agorom5c/s/acollsincubaodwihagomini-
OUAT.Gonorally,divalonionssuchasmagnosium
arosigniicanlyinvolvodwihhomoabolicpahway,
andaroakonupandsorodinvacuolosbyacivo
ranspor.Tho prosonsudyshowod ho doso-
dopondonloakagoomagnosium ionswihanin-
croasinggomini-OUATconconraionFig.4A.This
indingindicaodhahogomini-OUATinvadodho
cyoplasmwhoroidosroyodhovacuolomombrano
andproducodachangoinplasmamombranopormo-
abiliy.Woalsoassayodholoakagoohoabundan
cyoplasmiccaionpoassiumGhannoumandFico,
1999whichconribuosohomainonancooinra-
collularosmoicprossuroandohomoabolicpah-
way.Gomini-OUATconconraionso1.0and

3.0
Mlodohoincroasoinandhoroachingosaura-
ionlovolsopoassium loakago,rospocivoly.Tho
anibioicsniphimycinandamphooricinEaconcon-
raionsrangingrom0o10Mincroasohoamoun
opoassium ion loakago rom 5c/s/a
akayamaoal.,?00?,Usukioal.,?006.naddi-
ion,amphooricinEislossoocivoinpromoingho
loakagooUV-adsorbingmaorialssuchasnucloo-
idosovonaconconraionshighonoughoinduco
colldoahakayamaoal.,?00?.Thosoropors
domonsraodhahopormoabiliyohocollmom-
branowaschangodbyhogomini-OUAT,orhaho
uncionoprooins,whichacasranspororsonu-
rionsormoabolios,washamporod,causingsmall
ionsoloakhoughhocollmombranoandwall.Tho
prosonsudyshowodhabohsmallATFandsov-
orallargoinracollularcompononshardlyloakod
romcollsinhoprosoncoolowconconraionso
3DOEF-4,10

3.0M.ThoTEM micrographs
Figs.6CandDrovoalodobviousdisrupionoho
cyoplasmic,vacuolarandmiochondrialmombranos,
buascarcolyimpairodcollwall.Figuro7showsho
rolaionshipbowoonhoconconraiono3DOEF-
4,10anddosrucionocollularsrucuros.Furhorin-
vosigaioniscurronlyundorwayoundorsandho
ungicidalmochanismogomini-OUATsinmorodo-
ail.
HEFEHENCES
Cabral,J.F.8.199?Modooanibacorialaciono
dodino dodocylguanidino monoacoao in
FsaJcc^ass,/^aa^'//cc//c/.,38,115-1?3.
Codling,C.E.,Maillard,J.Y.,andFussoll,A.D.?003
Aspocsohoanimicrobialmochanismsoacionoa
polyquaornium and an amidoamino.'^^///cc/
|c/|.,51,1153-1158.
Donyor,8.F.1995Mochanismsoacionoanibacorial
biocidos./^///cJ//c//cJaJ.,36,??7-?45.
Di.,M.,Manrosa,A.,Fina.o,A.,Erra,F.,andnano,M.F.
1994 8ynhosis, suraco acivo proporios and
animicrobialaciviyonowbis-quaornaryammonium
compounds.'|5cc/F//^?[,1871-1876.
Fuhrhop,A.H.,andWang,T.Y.?004Eolaamphiphilos.
|/.,104,?901-?937.
Ghannoum,M.A.,andFico,L.E.1999Aniungalagons.
modooacion,mochanismsorosisanco,andcorrola-
ionohosomochanismswihbacorialrosisanco.//^
ANTFUNCALACTNFCFMN-QUAT 19
FlG.7.8chomaicroprosonaionohosopwisoaniungal
aciono3DOEF-4,10agains5c/s/a
//cc//a//.,12,501-517.
Kana.awa,A.,koda,T.,andEndo,T.1995Anovol-
approachomodooacionocaionicbiocidos.morpho-
logicalooconanibacorialaciviy.'^oo//ac//c/.,
78,55-60.
Kourai,H.,Yabuhara,T.,8hirai,A.,Maoda,T.,and
agamuno,H.?0068ynhososandanimicrobialac-
iviiosoasoriosonowbis-quaornaryammoniumcom-
pounds./a'/J|.,41,437-444.
Maoda,T.,Oka.aki,K.,agamuno,H.,Manabo,Y.,and
Kourai,H.1998 Eacoricidalacion o4,4'- 1,6-
polymohylonodihiobis1-alkylpyridiniumiodidos.//c/
F|a./a//1c/,c,21,1057-1061.
Maoda,T.,Yoshida,M.,Manabo,Y.,Oka.aki,K.,
agamuno,H.,andKourai,H.19998ynhosisand
animicrobial characorisics o 5,5'-|?,?'-
oramohylonodicarbonyldioxydiohyl]bis3-alkyl-4-
mohylhia.oliumiodidos.//ccc^/c/5c/.,4,1-7.
Mongor,F.M.,andKoipor,J.8.?000Gominisuracans.
^^n|/^//J/^/.,39,1906-19?0.
akayama,K.,Yamaguchi,T.,Doi,T.,Usuki,Y.,Tanicuchi,
M.,andTanaka,T.?00?8ynorgisiccombinaionodi-
rocplasmamombranodamagoandoxidaivosrossasa
causooaniungalaciviyopolyolmacrolidoanibioic
niphimycin.'//csc///c^.,94,?07-?11.
Oka.aki,K.,Maoda,T.,agamuno,H.,Manabo,Y.,and
Kourai,H.19978ynhosisandanimicrobialcharacor-
isics o 4,4'- 1,6-polymohylonodihiobis1-alkylpyri-
diniumiodidos.|F|a/a//1c/,c,45,1970-
1974.
Foson,M.J.,Mahias,J.H.,andDavonpor,L.1999
Aborranaggrogaionbohaviorincaionicgominisurac-
ansinvosigaodbysuracoonsion,inoracialonsion,
andluorosconcomohods./a^a/,15,7340-7346.
8hirai,A.,Maoda,T.,agamuno,H.,Masuki,H.,
Kanoshina,8.,andKourai,H.?005Eiologicaland
physicochomicalproporiosogominiquaornaryammo-
niumcompoundsinwhichhoposiionsoacross-linking
sulurinhospacordior./a'/J|.,40,113-
1?3.
8hirai,A.,Maoda,T.,Ohkia,M.,agamuno,H.,andKourai,
H.?007Anibacorialaciviyandcharacorisicso
modiiodorriopowdorcoaodwihagominipyridinium
salmoloculo.//ccc^/c/5c/.,12,83-91.
8hirai,A.,8umiomo,T.,Yoshida,M.,Kaimura,T.,
agamuno,H.,Maoda,T.,and Kourai,H.?006
8ynhosisandbiologicalproporiosogominiquaornary
ammonium compounds,5,5'-|?,?'- 1,6-polymohylnodi-
carbonyldioxydiohyl]bis3-alkyl-4-mohylhia.oliumio-
dido and 5,5'-|?,?'- o-phonylonodicarbonyldioxy
diohyl]bis3-alkyl-4-mohylhia.olium bromido.|
F|a/a//1c/,c,54,639-645.
8umiomo,T.,Maoda,T.,agamuno,H.,andKourai,H.
?004Eacorioclasicacionoabis-quaornaryammo-
niumcompoundagains/sc|/c|/acc////ccc^/c/5c/.,
9,1-9.
8umiomo,T.,agamuno,H.,Maoda,T.,andKourai,H.
?006Corrolaionbowoonhobacorioclasicaciono
abis-quaornaryammoniumcompoundandouormom-
branoprooins.//ccc^/c/5c/.,11,115-1?4.
Usuki,Y.,Masumoo,K.,nouo,T.,Yoshida,K.,io,H.,and
Tanaka,T.?0068rucuro-aciviyrolaionshipsudios
onniphimycin,aguanidylpolyolmacrolidoanibioic.Far
1.Thorolooho^-mohyl-^

-alkylguanidiniummoioy.
//cc/J|///.,16,1553-1556.
Vioira,D.E.,andCarmona-Fiboiro,A.M.?006Caionic
lipidsandsuracansasaniungalagons.modooac-
ion.'^^///cc/|c/|.,58,760-767.
A.SkAFTAL. 20