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Effects of Enhanced Ultraviolet B Radiation on Physiological Indices and Camptothecin Content in Camptotheca acuminata Decne

增强UV B辐射对喜树生理指标及喜树碱含量的影响



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,2016,36(5):0979-0986
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犈犳犳犲犮狋狊狅犳犈狀犺犪狀犮犲犱犝犾狋狉犪狏犻狅犾犲狋犅犚犪犱犻犪狋犻狅狀狅狀犘犺狔狊犻狅犾狅犵犻犮犪犾犐狀犱犻犮犲狊犪狀犱
犆犪犿狆狋狅狋犺犲犮犻狀犆狅狀狋犲狀狋犻狀犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪犇犲犮狀犲
WANGLingli1,2,ZHOUXiaojun2,3,LIU Wenzhe2
(1DepartmentofLifeScience,YunchengUniversity,Yuncheng,Shanxi044000,China;2ColegeofLifeScience,Northwest
University,Xian,710069,China;3SchoolofLifeSciences,LuoyangNormalUniversity,Luoyang,Henan471022,China)
犃犫狊狋狉犪犮狋:犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪 Decne.(Nyssaceae)isa majorsourceofanticancercamptothecin
(CPT).ItisimperativetoinduceCPTaccumulationinordertodevelopCPTproductionstrategiestosatis
fyclinicalusesofCPT.Inthisstudy,twoyearold犆.犪犮狌犿犻狀犪狋犪weredealt8heachdaywithUVBradi
ationfor12days,andoneyearold犆.犪犮狌犿犻狀犪狋犪wererespectivelyarrangedtoradiate2h,4h,6hand8h
withUVBradiationeachdayfor12days.Thecontentsofchlorophyl,MDAandfreeproline(Fpro),the
activityofSODinleafandCPTcontentinyoungleaves,youngshootsandrootwereseparatelymeasured
afterUVBtreatment.InordertorevealthatcamptothecinisthedefenseproductofUVBstress,the
effectsofUVBradiationonthephysiologicalindicesandsecondarymetaboliteswereanalyzed.Theresults
showedthat:(1)intwoyearold犆.犪犮狌犿犻狀犪狋犪,chlorophylcontentwassignificantlydecreased,MDA,
FproandCPTcontentsweresignificantlyincreasedafter8hUVBtreatmentdaily.Itindicatedthat8h
UVBradiationcausedgreatstressinfluencesontwoyearold犆.犪犮狌犿犻狀犪狋犪.(2)Inoneyearold犆.犪犮狌
犿犻狀犪狋犪,withthetimeofUVBradiationincreasing,chlorophylcontentwasgradualydecreased;Fpro
contentwassignificantlyincreased;MDAcontenthadnosignificantlydifferencebetween2hand6hUV
Bradiation,butsignificantlyincreasedafter8hradiationcomparedwiththecontrol;SODactivityde
creasedfirstly,thenincreased,lastlydecreasedwiththetimeofUVBradiationprolongingeveryday.It
showedthat8hUVBradiationalsocausedstressinfluencesononeyearold犆.犪犮狌犿犻狀犪狋犪.(3)CPTcon
tentsinvegetativeorgansofoneyearold犆.犪犮狌犿犻狀犪狋犪weregradualyincreasedwiththetimeofUVBra
diationprolonging,andthecontentswerethehighestafter8hUVBradiationeachday.Moreover,CPT
contentincreasedmoreobviouslyinyoungleavesandyoungshootsthanthoseinroots.Itconfirmedthat
enhancedUVBradiationcausedcertaindamageto犆.犪犮狌犿犻狀犪狋犪,and犆.犪犮狌犿犻狀犪狋犪respondedtothis
stressby not only changing physiologicalindices,but also changing secondary metabolism to
accumulateCPT.
犓犲狔狑狅狉犱狊:犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪;ultravioletB(UVB)radiation;camptothecin(CPT)content;physi
ologicalindex
  犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪Decne.belongstothe
Nyssaceaefamilyandisaperennialdeciduousplant
thatisuniquetoChina,andismainlydistributed
alongtheYangtzeRiverandSouthwestprovinces
ofChina[1].Theplantisknowntobotanyand
medicinebecauseitsvariousorganscontaintheal
kaloidcamptothecin (CPT)anditsderivatives
whichhaveimportantbiologicalactivities.
CPT,apentacyclicquinolinealkaloid,isan
effectivemedicineincancertreatment,whichwas
firstisolatedfromthestemof犆.犪犮狌犿犻狀犪狋犪,and
thestructuresofthisalkaloidweredeterminedby
Walandhiscolaborators[2].CPTisknownforits
remarkableanticanceractivitytoinhibittheeu
karyoticDNAtopoisomeraseI[3].Italsoinhibits
retrovirusessuchasthehumanimmunodeficiency
virus(HIV)andtheequineinfectiousanemiavi
rus[4].CPTisavaluablecompoundasachemicalpre
cursoroftopotecanandirinotecanwhichwereapproved
bytheUSFoodandDrugAdministrationin1996for
thetreatmentofovarianandcolorectalcancers[5].
Althoughmuchisknownaboutmanyfactors
whichaffecttheaccumulationofCPTin犆.犪犮狌犿犻
狀犪狋犪,stilonlylittleisknownabouttheeffectof
UVBradiationonCPTaccumulation.Becauseof
thepromisingclinicalusesofCPT,itisimportant
toinvestigatethefactorsaffectingCPTyieldin
plantmaterial.Inthepaststudy,droughtcanin
creasedCPTcontentinleavesof犆.犪犮狌犿犻狀犪狋犪[6].
Methyljasmonicacidandthetreatmentsofyeast
extractonleafdiscspunchedfrom犆.犪犮狌犿犻狀犪狋犪
seedlingspromotedthemRNAexpressionoftryp
tophandecarboxylase (TDC),akeyenzymein
volvedinCPTbiosynthesis[7].Similarly,methyl
jasmonicacidandyeastextracttreatmentson犆.
犪犮狌犿犻狀犪狋犪celsuspensionculturesincreasedCPT
accumulation[8].ItcanenhanceCPTproductionby
ethanoladditioninthesuspensioncultureofthe
endophyte,犉狌狊犪狉犻狌犿狊狅犾犪狀犻[9].Wang犲狋犪犾[10]re
portedCPTcontentdecreasedafter10daysUVB
radiation,andincreasedafter40daysUVBradia
tion.ButtheydidnotstudythechangeofCPT
contentbetween10days UVBradiation.This
studyinvestigatedtheeffectsofUVBradiation
between12daysontheaccumulationofCPTin犆.
犪犮狌犿犻狀犪狋犪,whichwouldprovideabasisformaxi
mizingCPTyieldanddesigninganaffectiveCPT
productionsystem.
089 ! " # $ % &                   36ñ
1 Materialsandmethods
1.1 犘犾犪狀狋犿犪狋犲狉犻犪犾狊
Oneyearoldandtwoyearold犆.犪犮狌犿犻狀犪狋犪
plantsweregrownfromseeds.Seedsof犆.犪犮狌犿犻
狀犪狋犪wereselectedtogatherfromBotanicalGarden
ofXi’aninNovemberof2012and2013,thenre
spectivelyembeddedinwetsandat25℃inMarch
thenextyear.Aftertheseedsgeminated,theuni
formseedlingswereselectedandtransplantedinto
flowerpotsinthebotanicalgardenofNorthwestU
niversity(Xi’an,China).
1.2 犝犞犅狉犪犱犻犪狋犻狅狀
SupplementalUVBradiationwasprovidedby
filteredGucunbrand(GucunInstrumentFactory,
Shanghai,China)30 W sunlamps.Lampswere
suspendedaboveandperpendiculartotheplanted
rowsandfilteredwitheither0.13mmthickcelu
losediacetate(transmissiondownto290nm)for
UVBirradianceor0.13mmpolyesterplasticfilms
(absorbsalradiationbelow320nm)asacontrol.
Thedesiredirradiationwasobtainedbychanging
thedistancebetweenthelampsandtheplants.
Thespectralirradiancefromthelampswasdeter
minedwithanOptronicsModel742(OptronicsLa
boratories,Orlando,FL,USA)spectroradiome
ter.Thespectralirradiancewasweightedwitha
generalizedplantresponsespectrumandnormal
izedat300nmtoobtainthedesiredlevelofbiolog
icalyeffectiveUVBradiation.Thelampheighta
bovetheplantswasadjustedtomaintainadistance
of0.15mbetweenthelampsandthetopofthe
plantsandprovidedsupplementalirradiancesof
2.1effectiveuw·cm-2.Twoyearoldplantswere
irradiatedfor12daysand8hdaily.Inorderto
furtherilustratetheeffectsofsupplementalUVB
irradiance,oneyearoldplantswereirradiatedfor
12daysandrespectivelyarranged2h,4h,6hand
8hradiationdaily.FilteredUVBradiationwas
regardedasthecontrol(CK).
1.3 犇犲狋犲狉犿犻狀犪狋犻狅狀狅犳狆犺狔狊犻狅犾狅犵犻犮犪犾狆犪狉犪犿犲狋犲狉狊
Physiologicalparametersweredeterminatedin
leavesof犆.犪犮狌犿犻狀犪狋犪.Chlorophylwasextracted
with96% alcohol,andchlorophyl contentwas
measuredaccordingtothemethodofZhang[11].
Malonaldehyde(MDA)contentwasdetermined
accordingtothemethodofHeathandPacker[12].
Fprocontentwasextractedfromleavesin3%
aqueoussulphosalicylicacidandestimatedusing
ninhydrinreagent[13].Superoxidedismutase (SOD)
activitywasdeterminedaccordingtothemethodof
GiannopolitisandRies[14].
1.4 犆犘犜犮狅狀狋犲狀狋犪狀犪犾狔狊犻狊
1.4.1 犎犘犔犆犪狀犪犾狔狊犻狊 TheHPLCsystemconsis
tedofaHPLCpump (LC10ATvp),areversed
phasecolumn(VPODS,150mm×4.6mm,5μm)
andaUVVISdetector(SPD10Avp)forthede
tectionofCPTat254nm[15].Sampleinjectionvol
umewas10μLaccordingtothepresumablealka
loidcontent.Theflowratewas1.0mL· min-1.
Themobilephaseusedwasmethanol/water(62/
38,V/V).Columntemperaturewas25℃.Under
thiscondition,the HPLCchromatogramsofthe
standardandsamplesolutionswereshowedinFig.1.
CPTstandardwaskindlysuppliedbyDr.H.Bischof
ofBoehringerIngelheimPharmaKG.inGermany.
Fig.1 TheHPLCchromatogramsofthestandard
andsample(peak1:camptothecin)
1.4.2 犛狋犪狀犱犪狉犱犮狌狉狏犲 CPTstandard2.5mgwas
putin50mLvolumetricflask,suitableamountof
chromatographicmethanolwasadded,andmetered
volumeaftertheultrasonichelpingdissolve.The
solutionwasshakedandfilteredwith0.45μmmi
croporousmembrane,then0.05mg·mL-1stand
ardsolutionwasgot.2,3,5,8and10mLofCPT
standardsolutionwerepreciselymeasuredandput
in10mLvolumetricflask,dilutedwithchromato
graphicmethanol,thenrespectivelymeteredvol
1895ò          ./0,ó:¾ÃUVB~€\]8‡¥¦Ÿ\]g›œhª«(ôF)
ume.ThestandardcurveforCPTwasconstructed
byseparateinjectionof10μLoftheabovemen
tionedstandardsolutionaccordingtotheabove
chromatographyconditions.Theregressionequa
tionbetweenpeakarea犢andtheconcentrationof
camptothecin犡 (μg·mL
-1)was:犢=16686犡
6577.5,犚2=0.9993.
1.4.3 犇犲狋犲狉犿犻狀犪狋犻狅狀狅犳犆犘犜犮狅狀狋犲狀狋 Thesam
plesweredriedintheshadeandgroundedina
mortar.100mgofthesampleswastransferredtoa
centrifugetubeand4mLmethanolwasadded.Af
terextractedwithsonicationfor10minatroom
temperature,30 mL waterand40 mL dichlo
romethanewereaddedandthiswasmixedvigor
ouslyfor5minonamagneticalstirrer[16],centrif
ugationfor10minat2000r·min-1yieldedtwo
phases.Thedichloromethanephase which was
provedtocontainCPT,wasrecoveredandevapo
ratedtodrynessinvacuumusingarotavapor.The
remainingresiduewasredissolvedinHPLCgrade
methanol(1mL),filteredwith0.45μm micro
porousmembranethengotsamplesolutionwhich
wasusedforthedeterminationofCPTcontent.
Samplepeaks withthesameretentiontimeto
standardswereverifiedbyspectralscananalysis.
2 Results
2.1 犈犳犳犲犮狋狊狅犳犝犞犅狉犪犱犻犪狋犻狅狀狅狀狋狑狅狔犲犪狉狅犾犱
犆.犪犮狌犿犻狀犪狋犪
  Thecontentsofchlorophyl,MDAandFpro
aswelasSODactivityweremeasuredaftertwo
yearoldplantswereirradiatedwithUVBradia
tion.TheresultsshowedinTable1,chlorophyla
andbcontentswerereduced,chlorophyla/bratio
hadalesserextentincreaseunderenhancedUVB.
MDAandFprocontentsincreased.Theactivityof
SODwasreduced.CPTcontentsinyoungleaves,
youngshootsandrootof犆.犪犮狌犿犻狀犪狋犪hadobvi
ouslyincreasedafter UVBtreatmentcompared
withthecontrol,CPTcontentinroothasnosig
nificantdifferencecomparedwiththecontrol.
2.2 犝犞犅狉犪犱犻犪狋犻狅狀犲犳犳犲犮狋狊狅狀犮犺犾狅狉狅狆犺狔犾犮狅狀狋犲狀狋狊
犻狀狅狀犲狔犲犪狉狅犾犱犆.犪犮狌犿犻狀犪狋犪
  TheeffectsofenhancedUVBonchlorophyl
犜犪犫犾犲1 犈犳犳犲犮狋狊狅犳犝犞犅狉犪犱犻犪狋犻狅狀狅狀狆犺狔狊犻狅犾狅犵犻犮犪犾犻狀犱犻犮犲狊
犪狀犱犆犘犜犮狅狀狋犲狀狋犻狀狋狑狅狔犲犪狉狅犾犱犆.犪犮狌犿犻狀犪狋犪
Index CK UVB(8h)
Chla/(mg·g-1) 1.507±0.125 1.193±0.266
Chlb/(mg·g-1) 0.646±0.102 0.505±0.142
Chla+Chlb/(mg·g-1) 2.152±0.165 1.698±0.075
Chla/Chlb 2.333±0.212 2.364±0.179
MDA/(nmol·g-1) 1.104±0.083 1.170±0.133
Fpro/(μg·g-1) 1.987±0.137 3.046±0.153
SOD/(U·g-1) 231.9±10.11 222.4±12.18
CPT/%
Youngleaves 0.219±0.017 0.313±0.019
Youngshoots 0.163±0.007 0.274±0.004
Root 0.045±0.008 0.067±0.013
  Note:Eachassaywasrepeatedthreetimesfromthreeinde
pendentexperiments.Thedataarethemeans±SEMofthreerepli
cates.Asterisk()indicatesstatisticalysignificantdifference(犘<
0.05;ANOVA,Tukeytest).
contentsinoneyearold犆.犪犮狌犿犻狀犪狋犪wereshown
inFig.2.Thesametotwoyearoldplant,chloro
phylcontentsdecreasedafterUVBradiation,and
withthetreatmenttimeprolonging,thechloro
phylaandchlorophyl bcontentsgradualyde
creased.Thetotalchlorophylcontentof2h,4h,
6hand8hUVBtreatmentdailywasrespectively
reducedby9.4%,10.8%,15.7%,26.8%than
thecontrol.While8h UVBtreatmentdaily,
chlorophylaandbcontentsdecreased26.4%and
27.8%,respectively.Chlorophyla/bratiohada
littlechangecomparedwiththecontrol,itissug
gestedthatthedifferenceofdestroyedextentabout
chlorophylaandbisnotobvious.
2.3 犝犞犅狉犪犱犻犪狋犻狅狀犲犳犳犲犮狋狊狅狀犕犇犃,犉狆狉狅犮狅狀
狋犲狀狋狊犪狀犱犛犗犇犪犮狋犻狏犻狋狔犻狀狅狀犲狔犲犪狉狅犾犱犆.犪犮狌犿犻狀犪狋犪
  UVBradiationhadobviouseffectsonMDA,
FprocontentandSODactivityin犆.犪犮狌犿犻狀犪狋犪.
WiththetimeofUVBradiationincreasing,Fpro
content gradualyincreased;SOD activity de
creasedfirstly,thenincreased,lastlydecreased;
MDAcontentkeptincreasingwiththetreatment
timeprolonging,however,therewerenosignifi
cantdifferencebetween6hUVBradiation,MDA
contentwassignificantlyincreasedafter8hUVB
treatment(Fig.3).UVBradiationofshorttime
inducedtheproductionofreactiveoxygenspecies
(ROS),suchasO2-· ,H2O2,andreducedSODac
tivity.Meanwhile,Fprocontentwassignificantly
increasedinordertodefenseUVBstress.When
289 ! " # $ % &                   36ñ
Eachassaywasrepeatedthreetimesfromthreeindependent
experiments.Thedataarethemeans±SEMof
threereplicates.Differentlettersindicatestatisticaly
significantdifferences(犘<0.05;ANOVA,Tukeytest).
Thesameasbelow
Fig.2 EffectofUVBradiationonchlorophylcontent
inoneyearold犆.犪犮狌犿犻狀犪狋犪
Fig.3 EffectsofUVBradiationonMDA,Fprocontents
andSODactivityinoneyearold犆.犪犮狌犿犻狀犪狋犪
Fig.4 EffectofUVBradiationonCPTcontent
inoneyearold犆.犪犮狌犿犻狀犪狋犪
thetimeofUVBtreatmentreachedto6hdaily,
SODactivityincreasedinordertoeliminateover
muchreactiveoxygenspeciesandFprocontent
keptinarelativebalanceatthesametime.When
thetimeofUVBtreatmentreachedto8hdaily,
SODactivitydecreasedandFprocontentsignifi
cantlyincreased.
2.4 犝犞犅狉犪犱犻犪狋犻狅狀犲犳犳犲犮狋狊狅狀犆犘犜犮狅狀狋犲狀狋狊犻狀狅狀犲
狔犲犪狉狅犾犱犆.犪犮狌犿犻狀犪狋犪
  UVBradiationalsohadobviouseffectson
CPTcontentsinyoungleaves,youngshootsand
rootsofoneyearold犆.犪犮狌犿犻狀犪狋犪(Fig.4).When
thetimeofenhancedUVBtreatmentreachedto8
hdaily,theCPTcontentsweresignificantlyin
creasedcomparedwiththecontrol.Withthetime
ofUVBradiationprolonging,CPTcontentsin
youngleavesandshootsincreasedfasterthanthat
intheroot,andCPTcontentofyoungleavesin
creasedto0.230%from0.106% ofthecontrol
groupafter8hradiationdaily,youngstemin
creasedfrom0.064%to0.158%,andtheincrease
ofCPTcontentinroothadalittlechange,in
creasedfrom0.036%to0.065%.
3 Discussion
  UVBradiationhasmanyeffectsonplants.
Althoughtherearesomesignificanteffectsabout
UVBradiationonplantgrowthanddevelopment
incertainspeciesandecosystems,itturnedout
thattheoveraldamagingeffectsofaboveambient
UVBaremodestordifficulttodetectundernatu
ralconditions[17].Anexaminationofmorethan
200plantspeciesrevealsthatroughly20%aresen
sitive,50% aremildlysensitiveortolerantand
30% arecompletelyinsensitiveto UVBradia
tion[18].UVBradiationactsasakindofenviron
mentalstress,andtheplantwouldhavetoadaptto
UVBradiationtominimumthedamagewhenthe
stresslevelreachtoalimitation.Withashortpe
riodofUVBradiationin犃狉犪犫犻犱狅狆狊犻狊狋犺犪犾犻犪狀犪
seedlings,rootswereelongatedandchlorophyl
andsolubleproteincontentwereincreasedinleaf,
butprolongedUVBradiationinhibitedtheelonga
tedrootlength,causedleafchlorophylcontent,
solubleproteincontentdecrease[19].Theeffectsof
enhanced UVB radiation on plantphysiology,
morphology,growthandbiomasshavebeeninves
tigatedextensively.EnhancedUVBradiationmay
exertanadverseinfluenceonthephysiologicaland
biochemicalprocessesofplants.Inthisstudy,we
reportedtheresponsesof犆.犪犮狌犿犻狀犪狋犪plantto
3895ò          ./0,ó:¾ÃUVB~€\]8‡¥¦Ÿ\]g›œhª«(ôF)
enhancedUVBradiation.Thisissupportedbythe
earlierfindingsofintraspecificresponsesinfla
vonoid metabolismin犆狌犮狌犿犻狊狊犪狋犻狏狌狊[20],soy
bean[21]and犃.狋犺犪犾犻犪狀犪[22]andinflavonoidcon
tentandchlorophylcontentdecreasedinrice[23].
UVBradiationhadobviouseffectsonchloro
phylcontentsof犆.犪犮狌犿犻狀犪狋犪.TheUVBradia
tionmechanismofdecreasingchlorophylcontents
wasstilnotclear.Changesinchlorophylcontents
haveoftenbeenusedasanindextoassessthede
greesofUVBradiationsensitivity.Inthisstudy,
chlorophylcontentsin犆.犪犮狌犿犻狀犪狋犪leaveswere
alsosensitivetoenhancedUVBradiation.UVB
radiationsignificantlydecreasedchlorophyl con
tents,primarilybecauseUVBradiationdestroyed
thestructureofthechloroplast,inhibitedthesyn
thesisofnewchlorophylanddestroyedthestruc
tureofchloroplastenvelope membraneandin
creasedthedegradationofchlorophyl.Yang犲狋
犪犾.[24]reportedthatUVBradiationreducedthe
photosyntheticpigmentinleaves (includingthe
contentsofchlorophylandcarotenoid,especialy
thecontentofchlorophyla).Zhangindicatedthat
UVBradiationofdifferentintensitycanmakethe
growth,chlorophylcontentsdecreasein犞犻犮犻犪犳犪
犫犪seeding[25].Theseresultsshowedthatenhanced
UVBradiationmadetheplantchlorophyl dam
age,changedtheproportionofchlorophylaand
chlorophylb,affectedtheformationofphotosyn
theticproteincomplexes,andinhibitedtheforma
tionoforganeles.
LittlewasknownoftheresponsesofSODac
tivityandMDAcontentstoenhancedUVBradia
tioninthepast.Theprimaryexplanationofcel
membrane’sdamageinducedbyUVBwasthefree
radicaltheory.UVBradiationmayinducethepro
ductionofreactiveoxygenspecies(ROS),suchas
O2-· ,H2O2,andchangetheSODactivity,result
inmembranelipidperoxidation,whichfurtherlead
tothechangesofmembranestructureandfinaly
altermembranepermeability.Asaresult,thecels
ofplantareinjuredandMDAcontentincreased.
SODisoneoftheprotectiveenzymestodefense
theinjuryofROS.Innormalcircumstances,the
generationandeliminationofROSinplantsarein
astateofdynamicbalance.Inourstudy,SODac
tivityfirstlydecreased,thenincreased,lastlyde
creased,whichisconsistentwiththechangeof
SODactivityunderlowtemperaturestresscondi
tion[26].ThestressofUVBinducedtheproduc
tionofacertainamountofROSunderthetreat
mentof4hdaily,inordertoeliminateROStime
ly,theactivityofSODthatisintrinsicalin犆.犪犮狌
犿犻狀犪狋犪decreased.Meanwhile,Fprocontentsig
nificantlyincreasedbetween4hradiationdaily
comparedwiththecontrol,whichindicatedthein
creasedFproimprovedtheresistancetotheUVB
stress.AcertainamountofUVBradiationmay
increasethecontentofROSsuchasO2-·inplant,
andSODcancatalyzeO2-· disproportionationto
generateH2O2andO2,enhancetheactivityofan
tioxidantenzymesystem[27].Feng犲狋犪犾.[28]alsore
portedthatenhancedUVBradiationinlowdose
orshorttermtreatmentcanbrieflystimulatethe
activityofSODandPODtoincrease.In犆.犪犮狌犿犻
狀犪狋犪,whenthetimeofUVBtreatmentwas4-6
h,alargeamountofROSwasgenerated,SODac
tivitysignificantlyincreasedtoeliminatesupera
bundantROS,meanwhileFproandMDAcontents
hadnosignificantlydifferences.Thissuggested
thathigherSODactivitycaninhibitmembranelip
idoxidation,andkeepMDAcontentinabalance.
WhenthetimeofUVBtreatmentreachedto8h,
asUVBstressdeepened,SODactivitydecreased,
ROSwasaccumulatedandthelipidperoxidationof
membrane system was enhanced,accordingly
MDAandFprocontentsignificantlyincreased.We
cametoaconclusionthatUVBstressof8hdailyde
creasedtheactivityofprotectiveenzyme,causedacer
taindamageofbiomembrane.Itcanbefurthertestified
thattheleavesof犆.犪犮狌犿犻狀犪狋犪becamewitheringafter
wehadstudied10hUVBradiationdaily.
Freeprolineaccumulationhasbeenobserved
inresponsetoawiderangeofabioticandbiotic
stressesinplants.Prolineisconsideredtobeone
ofthefirstmetabolicresponsestostress,andis
perhapsasecond messenger[29].Environmental
factorsincludingwaterdeprivation,salinization,
489 ! " # $ % &                   36ñ
highandlowtemperature,heavymetaltoxicity,
pathogeninfection,nutrientdeficiency,atmos
phericpolutionandUVradiationinducetheeleva
tionoftheprolinelevelinplants.In犆.犪犮狌犿犻狀犪狋犪
plant,enhancedUVBradiationmadeFprocontent
increase,itmaybeanadaptiveresponsetoresist
UVBradiationstress.Anegativecorrelationbe
tweenSODactivityandFprocontentwasfound
underUVBstressconditionin犆.犪犮狌犿犻狀犪狋犪.
MDAistheproductofmembranelipidperoxi
dation whentheplanttissuesufferedoxidative
stress,whichreflectsthedegreeofcel membrane
lipidperoxidationandtheplanttorespondto
stress.Inthispaper,MDAcontentshavenosig
nificantdifferencebetween6h UVBradiation
comparedwiththecontrol,however,MDAcon
tentwassignificantlyincreasedafter8h UVB
treatment.AlowdoseofUVBstressinducedthe
productionofacertainamountofROS,whilein
herentSODin犆.犪犮狌犿犻狀犪狋犪rapidlyeliminated
theseROS,andresultedinthedecreaseofSODac
tivity.WhenthetimeofUVBtreatmentreached
to8hdaily,alargenumberofROSweregenera
tedandcaused membranelipidperoxidation,so
MDAcontentwassignificantlyincreased.Incon
clusion,thereisacertaincorrelationamongthe
changeofSODactivity,Fprocontentand MDA
contentunderUVBstress,thedecreaseofSOD
activityisaccompaniedbytheincreaseofMDAor/
andFprocontent.
Undertheconditionofadversity,inaddition
totheplantphysiologicalindicescanbeinduced
somechanges,thesecondarymetabolismofplant
wilalsobechanged.Plantsinteractwiththeiren
vironmentbyproducingadiversearrayofseconda
rymetabolites,oneofwhichisalkaloid.Bioticand
abioticenvironmenthaveimportantrolesinthe
secondarymetabolizingofplant.Asasecondary
metabolite,CPTmayplayacrucialroleduringbi
oticandabioticstresses,whichposeagreatimpact
onalkaloidbiosynthesisandaccumulation[30].The
increaseofalkaloidaccumulationduringseedlings
developmentwasobservedandthisshowedthat
CPTmayplayadefensivefunctionfortheplants
duringthisvulnerablestageoftheirlifecycle.UV
lightresponsiveregionsinthepromoterofthe
tryptophandecarboxylase(狋犱犮)genein犆犪狋犺犪狋狉犪狀
狋犺狌狊狉狅狊犲狌狊hadbeenidentified[31].Inourstudy,
UVBradiationcanobviouslyenhanceCPTcontent
in犆.犪犮狌犿犻狀犪狋犪,wehypothesizethatUVBmight
stimulatetheexpressionof狋犱犮geneandincrease
CPTaccumulationin犆.犪犮狌犿犻狀犪狋犪.CPTaccumu
lationinducedby UVBradiationdemonstrated
thatCPT wasinvolvedinplantdefenseagainst
UVBradiation.When犆.犪犮狌犿犻狀犪狋犪sufferedUV
Bradiationinalowdose(2h,4h),CPTcontent
wasincreasedslowly;withthetimeofUVBradi
ationincreasing(6h,8h),CPTcontentwasin
creasedrapidly,whichilustratedthatthelong
timeofUVBradiationcausedcertaindamageto
犆.犪犮狌犿犻狀犪狋犪,and犆.犪犮狌犿犻狀犪狋犪defensedprima
rilybyincreasingthesecondarymetabolitesCPT.
Moreover,CPT contentsin youngleavesand
shootswereincreasedmoreobviouslythanthosein
root,itindicatedthatenhanced UVBradiation
couldpriorityimproveCPTaccumulationofaerial
organin犆.犪犮狌犿犻狀犪狋犪,becauseaerialorganmain
lysufferedUVBstress.
EnhancedUVBradiationcausedcertaindam
ageto犆.犪犮狌犿犻狀犪狋犪,notonlyaffectsthemorphol
ogyof犆.犪犮狌犿犻狀犪狋犪[10],butalsoaffectsthephysi
ologicalandbiochemicalmetabolism.Underthe
stressofUVB,犆.犪犮狌犿犻狀犪狋犪itselfdefensedthis
stressbychangingphysiologicalindices (i.e.,
SODactivity,MDAandFprocontent)andsecond
ary metabolismtoaccumulateCPT.Therefore,
thechangesofphysiologicalindicesandCPTaccu
mulationarealsotheadaptivemechanismtore
sponsetothestressofenhancedUVBradiationin
犆.犪犮狌犿犻狀犪狋犪.Furthermore,intheindustryplant
ingof犆.犪犮狌犿犻狀犪狋犪,themethodofsupplementing
8hUVBradiationdailytopromotetheincreaseof
CPTcontentisfeasible.
5895ò          ./0,ó:¾ÃUVB~€\]8‡¥¦Ÿ\]g›œhª«(ôF)
犚犲犳犲狉犲狀犮犲狊:
[1] £+%Q£#$õö÷øù,£#$ú[M]."û:+%
üýþ
,1983.52(2):144147.
[2] WALLME,WANIMC,COOKECE,犲狋犪犾.Plantantitu
moragents,theisolationandstructureofcamptothecin,ano
velalkaliodalleukaemiaandtumorinhibitorfrom犆犪犿狆狋狅狋犺犲犮犪
犪犮狌犿犻狀犪狋犪 [J].犑狅狌狉狀犪犾狅犳 犃犿犲狉犻犮犪狀 犆犺犲犿犻犮犪犾犛狅犮犻犲狋狔,
1966,88(16):38883890.
[3] KJELDSENE,SVEJSTRUPJQ,GROMOVAII,犲狋犪犾.
Camptothecininhibitsboththecleavageandrelegationreac
tionsofeukaryoticDNAtopoisomeraseI[J].犑狅狌狉狀犪犾狅犳犕狅
犾犲犮狌犾犪狉犅犻狅犾狅犵狔,1992,228(4):10251030.
[4] PRIELE,SHOWWALTERSD,ROBERTSM,犲狋犪犾.The
topoisomeraseIinhibitor,camptothecin,inhibitsequineinfec
tionsanemiavirusreplicationinchronicalyinfectedCF2Th
cels[J].犑狅狌狉狀犪犾狅犳犞犻狉狅犾狅犵狔,1991,65(8):41374141.
[5] GREEMERSJP,DESPASR,FAVALLIG,犲狋犪犾.Topote
can,anactivedruginthesecondlinetreatmentofepithelialo
variancancer:resultsofalargeEuropeanphaseIIstudy[J].
犑狅狌狉狀犪犾狅犳犆犾犻狀犻犮犪犾犗狀犮狅犾狅犵狔,1996,14(12):30563061.
[6] LIUZJ.Droughtinducedin狏犻狏狅synthesisofcamptothecinin
犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪seedlings[J].犘犺狔狊犻狅犾狅犵犻犪犘犾犪狀狋犪
狉狌犿,2000,110(4):483488.
[7] L?PEZMEYERM,NESSLERCL.Tryptophandecarboxylaseis
encodedbytwoantonomouslyregulatedgenesin犆犪犿狆狋狅狋犺犲犮犪犪犮狌
犿犻狀犪狋犪whicharediferentialyexpressedduringdevelopmentand
stress[J].犜犺犲犘犾犪狀狋犑狅狌狉狀犪犾,1997,11(6):11671175.
[8] SONGSH,BYUNSY.Characterizationofcelgrowthand
camptothecinproductionincelculturesof犆犪犿狆狋狅狋犺犲犮犪犪犮狌
犿犻狀犪狋犪[J].犑狅狌狉狀犪犾狅犳 犕犻犮狉狅犫犻狅犾狅犵狔犪狀犱 犅犻狅狋犲犮犺狀狅犾狅犵狔,
1998,8(6):631638.
[9] VENUGOPALANA,SRIVASTAVAS.Enhancedcamptoth
ecinproductionbyethanoladditioninthesuspensioncultureof
theendophyte,犉狌狊犪狉犻狌犿狊狅犾犪狀犻 [J].犅犻狅狉犲狊狅狌狉犮犲犜犲犮犺狀狅犾狅
犵狔,2015,188:251257.
[10] .ÿ!,EFG.¾ÃUVB~€\] "8$œŒ#$8
$g›œhª«
[J].#$+%%&,2011,29(6):712717.
WANGHX,LIU WZ,EffectsofenhancedUVBradiation
onbiomassandcontentsofcamptothecinand10hydroxy
camptothecinin犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪 [J].犘犾犪狀狋犛犮犻犲狀犮犲
犑狅狌狉狀犪犾,2011,29(6):712717.
[11] %&.#$8‡%æç()[M].Õÿ:Õÿ+%*+üý
þ
,1985:134135.
[12] HEATH RI,PACKERL.Photoperoxidationinisolated
chloroplasts:I.Kineticsandstoichiometryoffattyacidper
oxidation[J].犃狉犮犺犻狏犲狊狅犳犅犻狅犮犺犲犿犻狊狋狉狔犪狀犱犅犻狅狆犺狔狊犻犮狊,
1968,125(1):189198.
[13] BATESLS,WALDRONRP,TEAREID.Rapiddetermi
nationoffreeprolineforwaterstressstudies[J].犘犾犪狀狋犪狀犱
犛狅犻犾,1973,39(1):205208.
[14] GIANNOPOLITISCN,RIESSK.Superoxidedismutases.I.
Purificationandquantitativerelationshipwithwatersolublepro
teininseedlings[J].犘犾犪狀狋犘犺狔狊犻狅犾狅犵狔,1977,59(2):315318.
[15] ./0.\],-./h?@Ÿ\]g›œ0NÞßh67
[D].!V:!"U%,2006.
[16] VANHENGELAJ,HARKESMP,WICHERSHJ,犲狋犪犾.
Characterizationofcalusformationandcamptothecinproduc
tionbycellinesof犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪[J].犘犾犪狀狋犆犲犾犾,
犜犻狊狊狌狉犲犪狀犱.犗狉犵犪狀犆狌犾狋狌狉犲,1992,28(1):1118.
[17] SEARLESPS,FLINTSD,CALDWELLM M.Ametaa
nalysisofplantfieldstudiessimulatingstratosphericozone
depletion[J].犗犲犮狅犾狅犵犻犪,2001,127(1):110.
[18] TERAMURAAH.EffectsofultravioletBradiationonthe
growthandyieldofcropplants[J].犘犺狔狊犻狅犾狅犵犻犪犘犾犪狀狋犪狉狌犿,
1983,58(3):415427.
[19] HAN W,HANR.EffectofdifferenttimesofUVBradia
tiononseedlinggrowthof犃狉犪犫犻犱狅狆狊犻狊狋犺犪犾犻犪狀犪[J].犆犺犻狀犲狊犲
犅狌犾犾犲狋犻狀狅犳犅狅狋犪狀狔,2015,50(1):4046.
[20] MURALINS,TERAMURAAH.Intraspecificdifferences
in犆狌犮狌犿犻狊狊犪狋犻狏狌狊sensitivitytoultravioletBradiation[J].
犘犺狔狊犻狅犾狅犵犻犪犘犾犪狀狋犪狉狌犿,1986,68(4):673677.
[21] D’SURNEYSJ,TSCHAPLINSKITJ,EDWARDSNT,犲狋
犪犾.Biologicalresponsesoftwosoybeancultivarsexposedto
enhancedUVBradiation[J].犈狀狏犻狉狅狀犿犲狀狋犪犾犪狀犱犈狓狆犲狉犻
犿犲狀狋犪犾犅狅狋犪狀狔,1993,33(3):347356.
[22] FISCUSEL,PHILBECKR,犲狋犪犾.Growthof犃狉犪犫犻犱狅狆狊犻狊fla
vonoidmutantsundersolarradiationandUVfilters[J].犈狀狏犻狉狅狀
犿犲狀狋犪犾犪狀犱犈狓狆犲狉犻犿犲狀狋犅狅狋犪狀狔,1999,41(3):231245.
[23] TERAMURAAH,ZISKALH,SZTEINAE.Changesin
growthandphotosyntheticcapacityofricewithincreasedUVB
radiation[J].犘犺狔狊犻狅犾狅犵犻犪犘犾犪狀狋犪狉狌犿,1991,83(3):373380.
[24] 123,4 5,.67.¾ÃUVB~€89“:;jABA
Œ–—˜™šhª«
[J].8N%&,2000,20(1):3942.
YANGJH,CHENT,WANG XL.EfectofenhancedUVB
radiationonendogenousABAandfreeprolinecontentsinwheat
leaves[J].犃犮狋犪犈犮狅犾狅犵犻犮犪犛犻狀犻犮犪,2000,20(1):3942.
[25] %è,?0@,ó.ÌAÃBUVB~ÄŀCD
 "8ڟ“”•EFGžhª«
[J].![:YU%%&

)*+%ý
),2010,35(1):105110.
ZHANGHX,WUNB,HULT,犲狋犪犾.Theeffectsofdif
ferenttreatmentswith UVBradiationstressonseedling
growthandcharacterofchlorophylfluorescenceof犞犻犮犻犪犳犪
犫犪L..[J].犑狅狌狉狀犪犾狅犳犛狅狌狋犺狑犲狊狋犆犺犻狀犪犖狅狉犿犪犾犝狀犻狏犲狉狊犻狋狔
(NaturalScienceEdition),2010,35(1):105110.
[26] HIJ,%KL,1…M.»NÄŀ\] "SODž、MDA
Œ˜™š›œhª«
[J].OP+%67,2002,15(2):197202.
FENGJC,ZHANGYJ,YANGTZ.Effectoflowtempera
turestressonthemembranelipidperoxidationandthecon
centrationoffreeprolinein犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪seedling
[J].犉狅狉犲狊狋犚犲狊犲犪狉犮犺,2002,15(2):197202.
[27] 4 5,Qª«
[J].U±+%%&,1999,19(4):453455.
CHENT,REN H X,WANGXL.Influenceofenhanced
UVBradiationonantioxidantsystemsinwheatleaves[J].
犃犮狋犪犛犮犻犲狀狋犻犪犲犆犻狉犮狌犿狊狋犪狀狋犻犪犲,1999,19(4):453455.
[28] HV,VWG,HXY,ó.¾ÃUVB~€ZD[\]¨
©hª«
[J].#$%&,1999,41(8):833836.
FENGGN,ANLZ,FENGHY,犲狋犪犾.Effectsofenhanced
UVBradiationonproteinmetabolismofbeanleaves[J].犃犮
狋犪犅狅狋犪狀犻犮犪犛犻狀犻犮犪,1999,41(8):833836.
[29] HAREPD,CRESSW A,Metabolicimplicationsofstress
inducedprolineaccumulationinplants[J].犘犾犪狀狋犌狉狅狑狋犺
犚犲犵狌犾犪狋犻狅狀,1997,21(2):79102.
[30] ^Y_,` £a,` 2a,ó.\] "£\]gŒ10b,]g€cdh«¯GeŒfg
[J].#$%&,2003,45(7):
809814.
ZUYG,TANGZH,YUJH,犲狋犪犾.Differentresponsesof
camptothecinand10hydroxycamptothecintoheatshockin
犆犪犿狆狋狅狋犺犲犮犪犪犮狌犿犻狀犪狋犪seedlings[J].犃犮狋犪犅狅狋犪狀犻犮犪犛犻狀犻犮犪,
2003,45(7):809814.
[31] OUWERKERKPBF,HALLARDD,VERPOORTER,犲狋犪犾.
IdentificationofUVBlightresponsiveregionsinthepromoterof
thetryptophandecarboxylasegenefrom犆犪狋犺犪狉犪狀狋犺狌狊狉狅狊犲狌狊[J].
犘犾犪狀狋犕狅犾犲犮狌犾犪狉犅犻狅犾狅犵狔,1999,41(4):491503.

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