..- r 1r ,,:per ir. the if.; e 1.:utells.J.
1. UIL: F Tel <. I?1 F h l;'1 TED ti TH~E C:f Ls tIT L .s.IE t
100 10lP I:F FHI LOIPH,'I~
U']IVETIT I if FLC' C arL
P lI. IJOULE IGE n l lTS
The authjr IxrerZSies his since-*** rhaniks to- [rr. T. E. Hu.nIphre.ls
forT hir 41uidncz throughohut thr. graduote Iprogi'ri m and fo~r hi5 red ice,
pa~tiencei and help ;n conducting e...perimecnts and Tpjrept.rin the r-a~nu-
ECr i p.. It is j pleasure to ....*ork in h~iE IdjTlabor Tor bTnd ch;are h
e~quiprrien t. The hEp oF 1.* rs. D. S. IAnthcr,, F., L. 5irat h ;ind r.. H.
b~cggs a roalitta.;r~ i,,chllers ;s jlEC ajppriciated. The. Forjn= D department[
FABLE OF CONTENTS
ACKNOWLEDGEMENTS .. .. . .. .. ... . ii
LIST OF TABLES .. .. .. .. .. .. . iv
LIST OF FIGURES .. .. .. .. .. .. .. v
ABSTRACT ,,. . . .. .. .. . . .. vi
LITERATURE REVIEW . .... .. .. .. .. .. .. ..
METH1ODS AND MATERIALS . .. .. .. . . 5
Preparation of Scutellum Slices .. .... 25
Analysis of Sugars .. ... .. .. ... 25
Manomectry . . . . . . . . . . 26
Metal Analysis ....... ... ... 27
RESULTS . ... .. .. . ... . .... . 2?
Kinetics ... .. .. .. .. .. ... 29
Sucrose Uptake . ...... . . S
Ga: EvChange .. .. ... .. .. . .. 2
LIST OF Trs3LE J
I, lehod of Cajlculjting Data ,, ,. .. . .. 31
2. Gluicrt UIptijke jS e'ifated b, Ulrjn*,l
loa n thieBatlhing Solution ....,....,, LS
3.Fermrentct onl in Water And Suigar
:olutions .. .. ... ... .. .. .. S
4. Thc E ffec t o f Uran,1 Iri t -ae Pre rea t-
me~nto~n ~uci'c w~ptjCe ............. 56
5. InhibitiOn of lugar Uprake b, Phlcridrin ,. j3
6. The linhibition b, 00lP of Uptake fromi
0.0111 Eugar SoluIon1 .. ... .. .. .. 59
7. ljltose Uptne . . , . . . . .. I61
?. Thez EiFfcts of' Turaneose and flanni rol
ojn Sacro; e U~prLke . .. ... ... .. 03
1. Suc~Tros TiiSue Lee I and CSuirise Uptake~ ,, . O'r
10, Sucrsse Uptake a Affected h, Various
Cations ... .. .. . ..... . .. . 701
LIST OF FIGURES
1. Sucrose Uptake Vs Time .. ... .. .. ... 30
2, Sucrose Uptake Vs Time .. .. .. .. . ... .. 35
3. Sucrose Uptake Vs Time . ,. .. .. . ... 36
4. Sucrose Uptake Vs Concentration . ... .. .. 37
5. Sucrose Uptake at Constant Sucrose
Concentration .. .. . .. .. .. .. ... 40
6. Glucose Uptake Vs Time .. .. ... .... 41
7. Glucose Uptake Vs Tinie .. ... .. .. .. .. 42
8. Rates of Fructose and Glucose Uptake
Vs Concentration ,. ... .. .. .. . .. 43
9. Glucose and Fructose Uptake as Affected
by Uranyl lon Pretreatment ,. .. .. . 44
10. Uranyl lon Pretreatment ard Glucose
Uptakte . ... .. .. .. ... ... . 48
11, Cluroze Uptakle at Constant rCo~ncntraton .. .. 50
12, Fi rre n rjt i on i n r Ij e r and Su; r ose .. .. .. 6b
13. Gas E chjlnge in r'jucr snd 0. 111 Surore ,. . .. 70
IL. Cjf Echangc in riata ,-ndj 0.lrIGlucee .. .. .. 7l
IB. Ecccr.ercatson ir. Eve:lar :Llurion .. .. .. .. 72
Ii. r~crsl tsin.~lr.:i ..............,.... 75
II, Effeci of Urari,I I;n Pretrtr c rncir ConrcLntra-
tiojn or. Supr Uptake, .. .. .. .. . .. :7
15. Fletal Einding Fo~llrow~ig ar. Acd P~ctreateentr . .. 78
Abstrjct of' Dircertation; Presenred to the
Graduate Council of the University of FloridJ in Parr;al iulfillImen~t
of the P~equirements for the Degree oi Doctor of Phillosophi
SUZG'.f. TP. USPCIFT lu, THE 1141:;E SCUTELLUM;
JoscFph Henr, Wn~tosell
Chairman: Dr. Thioma~S E. H~umphre-:s
Major Deparrreeent: Ecn
Characteristics of the uptaki- of sucrose, gluicoe and frucloto b,
maze scutellum slices are pre~sented. CSugarj Iere taken up at alm~oi!
a conrstnt raite until thej bjthing jclullo~n wal depie.edl e.cn at co~ncen-
[rations+ u:11 belou those whlich saturated the uptar.0 rmechanirms. Th
effect of DUP1Y, phinf'idrir, uran, I ion, jnd anor.io twas to ir.h~ibit r e
uptake of ucrorr e appro-.ii satel mic a~iE s bmuch as the UpEra ke ot h;hl5se.i.
Malrics wasr talen up wi; hoiur hydroli sis. TurCAnoSe I-.i1 not taken up
but slighri, inhiibiced the uptal.r of jucirose. The following conclulsions
are c~r awn. (a: Eucrore- is taken~ up -jCrivcl, aithiout in.erjion. (b)
Hesxose; are taken up bi 1'0 pr3 ~ocesce operating Simultadneou5i,, dirffu-
sion and cci'..*e transport. (c) The actise uitake mechianisms for sucrose
and the he.:-.oses arre locate at the plasmalerr~ain (dj The acti.; uptar."
mechasnisrms for bothi sucrose and thL he>;ose5 are driven by glicrlysiC.
(2) hotel binding chiarac reristics of the scutellum art different from
those of ;east in that binding is not specific to the urptaker rtes and
bound ralttal ionis aire apparently not released during :ugaJr uptalke.
Much research has been conducted on the movement of sugars in the
cells of animals and microorganisms. Stein (1) reviews this work in a
p~ljnE?, jrn.; ,r the Probt le.. is I:jhtrj l II0 [hi L..II r~: L rldin3 ilf plinE
ph,.CIMAt least it.. rcsprcr ;c. h;lhcr :r-3.n;:r~:I;i th.. re ..Trt orr
;) up :.-.1 c.-nos acc.:1.ride 12 Sucrose 'i 1, Fir the ireson .ii:Crver
st...I ib .~i~lr i i, rr- ,.10 -t d .. plan i [" IId IL.
rea.o :I ~ i:: ,*5. 0 .;r uni lil I: IIIln:.n-s of ~Ch.-- [rITI:.s.5 of 00 e.-:
su r: pro 1 .se .3irth iU.. Lhd th Lr I:E.i: ?nh:...~T~ rt ::in ii( i reser
Fr~lia c r ~ ~ ldl, 'Ir *:ced:,; fru l- .-cor and.lP:.ter partE of lj ants-~ J h)L are-
I NTRODUICT ION
neeir;al to supprrt juch concepts ir 50.11 grear.1, inedcquar-_. ILc iljo
5ta!Cs that the verry exists.*nce of~ c`rri rs has not: tie1 pro;.cn.
studir?5 or BiideSki (:) orn the~ acculrsulation of~ sugars b, plhlolm~ L1ssue
jdd support to th( CO[Lncentin that an acrlt., mostment or su~croSE ;s
probabl, a part of phlopm transport.
The rrork reported herein wa~s underrtake:n rrO 3-in a better ulnder-
sranding or thea process ofi sugjr uptake;,, espc~i;Jlly t."r.;t of ucrTe.
Several .h-racterscists makeZ the c..rn ;s~cullum well F.II;.:Jl for Lhe.
sHudy of iugar uptake.: 1,.3) Ths uptak3 of sucars is real oilorwing
in rhe- Form of sucrose rather rhan~ itarch: alucsse :s not -accur~Iulated:
(c) there ij nro aJeinJ phenomenon durirag which 4n adjustnerr~t in thr-
reSpirtory rT3[ ccurs;~j jnd (0) mOrphologicall,, the scutellun i:
leaf rissua which functions ir. cha mosement. Gi sugar r'r001 Ehe end~~iperm
to .he Je.loping seedling.
Several systems have been described in which sucrose is taken up
without prior inversion.
Weatherley (6) measured sucrose uptake by floating leaf disks of
Atropa belladonna on 10% sucrose solutions and taking dry weights be-
fore and after uptake. At times the amount of hydrolysis was very lower
and the amount of hydrolyzed sucrose in the bathing solution varied
independently of uptake. Whereas pH affected the amount of hydrolysis
it had very little effect on uptake. The amount of hydrolysis was
greater whien older leaves were used. Washing disks prior to treatnment
decreased the amount of hydrolysis. He concluded that sucrose was
absorbed as such. He found (7) that the uptake of glucose and sucrose
on a per mole basis was nearly identical.
Iweatherley (8) also found that the uptake of sucrose wras revers-
ibly inhibited about 75% under nitrogen. He followed the loss and up-
take of water as well as dry weight and included a discussion of the
osmotic situation prevailing in the tissues. He concluded that thle
uptake of sucrose was probably an active process.
Experiments by Pennell and Weatherley (9) showed that sucro:- up-
take was in~hibite-d about 5070 by 2,4-dinitrophenal (DNP) and not at all
by phloridzin (both at 1 mM). In this paper the amount of dry , 2.ht
increase was shown to be caused partly by an increase in s ar;se.
glucose and fructose. The amount of increase not due to th:.:- siy r
a 5.1 jssur.. *J t h. .: E .(. st -ar th ferrat ioi:.- s= .t s g F r..
aenou~nr of .insight inre.=.*~d rr:. jlas due ;o starh ;t w93 iArluLed thatL no
"uphalll" mrjl~ncnt. of sucrore need Occur,
Pojrter a4nd liev, I' 1) wo~rrked rwi h to~bsico leaf disks~ jndi measures
uptake, accumulation and Lgaj errhange in SE solurio~ni of juccose~ andl
in.*err sugaJr. Wrhen ;nterE 5uqrr was5 supplied, ?luicse dis3F~appeard a
3 faster rate thiar. Fruirtose. InvrrE sugjr cjiusedj ~a re rapid a.:cumu-
latio~n of starch than did juCrjse. The totajl uptake~ ojF 5ugar vaS [he
samie wrhethe'r indfrt 5Jgar Or SUCTCIse w35s Suppl;ied symual
late led sucrose could be r~co*.red fromn the le~af disks jfrer bei~ng
jpplied in the~ bathing solution ;hroling that juirose could be taken up
without ;ns\erjion. Thla! around ,n PQ? oF 0.74 du~ring ixubatiojn of tijjue
in varer, but this ;Increased to aboute 1.1 when [he Lilsue' was incubated
in ?ucrose' or he.*ose. The speCi fic ac t i it*I Of th;- CO2 c.o=.ed was5
about Ihe jiame aS thjt Of thC applied iugar leading thrm ra belireve
rhac rhe Obser~~td rate of gss l.changz .-jos noi the result of a hiigh
rare of formn.tnacior soverimposed on the 9.3' exrchange" i rraterl. They
ruggelead~ rather a crhif[ in subjrrarste LO~ Suyr win j,...prs wrCe jddedd
to the bathin3 solcrion.
Viccer: and fIerrcer (II) studied the urt'l:.? o' jurrose by bean
lea tee.-.The bjrhir.g Iolution contajined1 e-l; Sucro-e andj a !rjce
of fractateC jf t incubation, in';:aring that ex.trartellulj r .r,drclasi s
..Ja= ;mall or no~neiictnt. Wrhen tijjue sampFles Nere ant.lued .roit of
rhe Sugary woSS Sucrose I*;th small Snounts of reducing sugars present.
Gas e.*.changel was5 measured jnd1 rh-. P.2 increased fromi about nS to I.0)
r.hln sucrosee was Addd to .he barhing solution, The notiiced a liner
phase2 of jucrJSe Iip(? e wirth lime2 despite large~ concentration diffe-r-
ences in the bjfthJr jcj].jo;in. Tht ste.ao l ract ofi jsaro~se uotae Iwaj
about 9 umoles/g fr wt hr from 1% (0.028M) sucrose. An inhibition of
sucrose uptake of 55% was noted wi th 2.4 x 10dMW DNP. Uptake was mnea-
sured over periods of 8 to 26 hr.
Vickery and Mercer (12) reported increases in 02 consumption upon
the addition of surcrose that were of short duration and independent of
concentration. They stated that the rate of C02 production showed no
correlation with the concentration of sucrose in the external solution
(and hence, in the free space) but was strongly correlated with concen-
tration of sucrose in the apparent osmotic volume. This is used as an
argument that the sites of carbohydrate metabolism are included in the
asmotic volume for sucrose and that part, at least, of the cytoplasm
is included within the membranes involved in sucrose transport. Su-
crose accumulated against a gradient and the initial rates of sucrose
uptake followed a concentration vs uptake curve adaptive to Michaelis
and Menten kinetics, After several hr of uptake the rate decreased.
This decrease was attributed to inhibition of further uptake by sucrose
inside the cell. They argued against the possibili ty that sucrose
pumps occur only at the tonoplast.
Hardy and Norton (13) studied the uptake and utilization of 14C-
labeled sucrose, glucose and fructose by slices of potato tubers.
Glucose was taken up faster than fr-uctose. All three sugars were found
when untreated tissue was analyzed for sugars. It was suggest--I o,n tha
basis of the labelina of various intermediates that sucrose wa- asre.
unchanged and transported to storage where part of i t was hydro~l,:-c
resulting in storage of all three sugars.
Sacher (14), using bean pod tissue, found an extracellules ;I
o~uter p;ci ..-,.:rr a~i c ..n e- ricd .cas~n jl I n i ts activi ty. Hot-rei .-c
suiroie uptak~e ass njLe depenldent on in~tertsiF act..i C a; Snealrn b, the
obser.ation rhat :Jiose wajs taken up in rth abse~nce of ourc~r :pace
irler tose a;t; vi ttl. ClucO-;e uptate <--.;; thre times as5 Ifat J: Frur. CoSi
uptakte from 0.030l solutionl.. There L4-Sa no aiitct o: 10~ t o 10 'tI
uranyl nitrate on uptral.e ofi 0.0?11 glucosa or sucrjas. The Fact that
Sucrose~ wajS tken as as 5suich.j jlii als~OhST[ deosr trd bySholi~ng thjt the
giveme/frucT~i tose rljdoiour i .It rtjio kzs l iti ct:hajngd b:; Iiptjl.e a~han
us ii n, fruc tose -labr led SuCrose, Uni forlily labeler d sacrose relmaincd
unifo-ml, ljbeled ic .n in the presence of unlabe led fructose or glucose
in the~ ha .hing solution. DlrfP (: n 10- 11) inhtibied uprakei from 0.0003Hl
jucrosc 6b ,and ircm 0.0!n suc~ros e 881, In Freshl; cut !;jssue the
endogenous Sugun5 cnisted4( lajrgell of glucose a-nd :ruicts. dnd only
trjici jrounts of suiTrOe. Upon incubation in sugqar silurionn juc.-esee
Wji storedJ, apparentl i ini ths .acule, and the suloC.C/reducingnl jllljr
wrLd the sucrose .-as hyorollzed and the r~tic rjreiltl decreased.
jacher jrouei on cnhe bjsjisl ofSe.eral linesj of I::;jrdence thb[ L:-a rate
l im' rin1 5 tp in hcrose uprake lilj i n Ehd Formjtioi of ;uCTC;2 ind
that the cit~plicam iS frae space to the ta.(ese.
In the gterminating~ :astor bcan, suirose 15 syntrhsi.zed In the; iidospirsT
jt '.h e apensi of ijt. Thr suicree is ca'-.en uoj b, the- ijt**lldon5, and
It ii transplorted- fromr thean into the de.elc~oing ieadlling ar;s. In the
lui... Isest of thr .ugar~ is in the iorn of ;Lucrose with ..irtu-lll ro free
Iietuies present. h;heds ner.pa-o oreriht b
th~e rotyledoins and' suc:rose .>a accurT;ullated aginer olCentl. tion
gradient. DNP partially inhibited the uptake of sucrose. Sucrose was
shown to be taken up without inversion by several lines of evidence
including the retention of asymmetry of labeled sucrose applied in the
bathing solution. In this tissue sucrose was taken up at a higher rate
than either hexose and, more unusual, the r-ate of fructose uptake was
considerably greater than that of glucose uptake.
Kriedemann (17), on the basis of microautoradiographs of castor
bean cotyledons exposed to labeled sucrose for 20 min, suggests that
the cell walls and intercellular spaces provide a diffusion pathway
by which solutes can gain access to the vascular system from an external
Kursanov (3) refers to work in which it was shown that the fibro-
vascular bundles from sugar beet petioles took up sucrose, glucose and
fructose but exhibited a much higher affinity for sucrose.
Grant (18) studied the JptakC of glucose, fructose, and several
other monosaccharides by carrot and corn root tissue. Some of his
results were as follows. Carrot root tissue exhibited a )ag of several
br before uptake began. The uptake of glucose under N2 was less thouI
2E~o tha in ir.He sho.--ad that Jlucor entcred the ice II re jccu-
v~ulated as thc l'ree; sugjr jgjinst .j conecnrtrjrion gradient~. t l-
i'Cas co-~ernicr.11;Cr in the carrOI. ti5Clle oC~iEedd 1!.0!5tt j51C.rir~3 Squjl
d i r ribu! io~ r; aLh.;r [th ire: h us5 i ght o:f the t i .u-. Uptated of glueseI.
rroceededj at j conr rant ratec from~~ 0.0010II sclutionn urst" the batting
solution ~ ~ l ase-as e.H how' e Ehjt th~e ur Eal e OF 3luiiGSGI )nrJ
:e*,e~ril nwhrr csuaors folion- uFptake .: Concen[ trat~C1Tio curse accrdi
t~ liichi eiI ar..i rfierer r, I.iraticl.. 110: did n:t ferii rc. be co~n.; rni d
w~ith rth facr that! a coni cant irat c.1 uputjle t*Jih tiii6, in Fpite OF i
declining sugar' cor..:"'retra on, i r incon sir E::n t .-ai h reactio n~ ratci. ;a
pred~i ted t., rlchaeli5 anid hcntrn k;itt i cs. The mar i nes. ra r-i GI
surgjr uprjl.e b*; carror d-isc; werer frorc 3. to 10 unnolclc*'g b~r and frGD=~
corn roots sure 1ro.0? to 24 umolc.'g br (1 l. I t wa ElIcO:.n th;;t in
carrotl t;Isue the repIredT~ CO., wasr der;.*d preferrentiall*, fror, thi
e nte Pran I sjiuga r L.
Apee..s jnd Eee~ers (203) userng iarro[ and pOtato slices mieasured
0-, consulptionl 3nel C0-. ev~Il[olutio ;i r. o (0.5 umoles m~i) cor~c.-nrati;. ns~
of glui'ose, Thle T.Q did noj :..ar,* Li r.irr fiicanti ,l fromn ..aci: jnd the
addiction of glutose did rootL induci iign~tf cjnt changes~~ ;Ir 02 upruse or
PE i nihold and i~nhar i 'I dtlror.5:ra~ed ;in jci rie uptak~e me~chan nis
i n car rot root tIssu wC\hic was capal of~j atCI CCrIITmulat in rl -o-meth-,*gl('1]u-
cose. TIhe rate of uptake~ of ,-o-r..ath*,dglu.:ose -.-.ried uith con~ce.-.ra-
Si on approxima tEly a; pirtdicedL b iiichje i njyd fIern itn k~i netfi cs A .
a period of uptal.E the rirnue \ia rinse~d for !0 min ;rnd Lther plactJ in
uater and a concentrationi ratio of ;5: 1 ras mjirtair.=d britsmEn theI Can-
centrations in the ticsue and a-.ele. Chron.oOrlgraphy and jnd lis oif
C02 i ndi cj ed rhat r -r -w thlg iacose rra not ~~ rnetaolIized.
Harley prnd Jeranirage (22) Studied the uptake; of jugari by. becch
mycorrh; :as. They fournd that the~ rate c*1 uptak~e *. concentration
curses for glucsejs and fructo'e formed rectailgular h**pertolat butr the
shapes o~f thi curses wcre different, the m.iir:mum rcat for fruCrGle
bi i ng h;9ghr thar the me..imium rate for gluicesc. In rlhi; rssu ch re-c
is considerable hydrolysis :Ihen sucrose is supplied. iPmrd,
i nh ibi td the abjOrp [i on oi: he.-.oi-.i. When lnoitures of glu~cose2 and
Sr uc toid~ s-tre ;u~p pi ed g lu~c~te w~s prefeiren t;i I ly absorbe-d. The
addition of sugarr raused a re:.pirjrar, siuaie n rmeuooa
concentrations the stimulation caused by glucose was considerably
greater than that caused by sucrose in spite of the fact that a roughly
equal amount of sugar on a weight basis was taken up during the measure-
Mlorgan and Street: (23), studying the carbohydrate nutrition of
excised tomato roots, found an RQ of about 0.75 in water and about 1.0
in sucrose. The root segments had been starved prior to measurement
and the uptake of 02 was stimulated by the addition of sugars. The RQ
of root tips supplied with sucrose, dextrose, galactose, or reffinose
was within the range 0.90-0.96. The endogenous respiration had an RQ
as low as 0.70 and in mannose as low as 0.60.
Thomas and Weir (24) measured the uptake of sugar by tomato root
segments from solutions of 0.05M glucose and 0.025M sucrose. It was
found thjr mord~ Suqjr Co i rriighl basgi2 Irj Ejiin up arh;r suirOlt \ ii
zupp~lied! a co~mpar~ di to ucO-;t. Sucrose ii mortedI; SUpCTrO [r t
gluci.Zd ini 5urpoT[r tJ ng gro[h :I deCi fed [.:rnl.m l roo:[ .
Whenr radi Ih rec.([ ilicGE firr IFICu~baE.1 ir. Euiroie thereT if i COF.i-
-ider~abl. jmmount of :.
a hoerh~ or nojr 3-, zucrczC is t aler. up a 1 ho~ut ;n.trilon. .ac~rose at
0.029tri and 0.i05rf 1 hroh rirnulated the i-.luciion ofl CO, und i t rw-- rug-
cie_ d F.MEr this vjs duc to~ j ;1Uturade..i ofi respirT~jOTrZ, enz*tn
BIald:Ci (5) areasurtd the upukel; 01i iuiirot b eciterd .cEcular
buindle: Or phI~eile thf~ue: iriii a *jiC.rit* ilf plards.. II. appFl- phlomi
and celtr, .Jc'uljr bundleCS abou~ ;C.~ .0l [r. of L6. th ptk rom~ 1 1 suirose
could bei fourid ir. the- til.ue ii [he form of :u.:rose. ureupa,
irom 0.0031 andj 0.011: tolution, proiedLedd at 6 priogressi-.el, skwarr
race until rhe e-.:er nal sollut;ion contaiind abou[ 10: r of h- original
amoT~unt iof Iucre't.
Sucrose waSj takeni up, jgainst icncsntration gr..Jlents of the jrdcr
of 103. Thi ,rates o accumuljtionn t. .aScular LbundlL'; cr phlocle ti;s be
weire muCh higher th~an rates b; pjreracn,ma froml the samie plant. Vsua
cissue jccu-.aul3Led~ Microse at ratej of J to- 16 orales.i'g El- vt br from
P-i.haldJ aid Ellsion (2.; addresled. the qlucstion as to *.*hrether thr:r
wajs j difiusion barrier bet.-seen irternal jubstrjt: ~nd i i~es of resps-
rur.The ,* applied SurnilOw-r hrpojCot,*I segme~nts usi r labeled :3lucose
o~r Iluitamric aci.1 lrn the preihhnce or absence oi 00N ar~d liCO3Ured thC
total jnmoint of i.12 r.ol.id and l'd spec Ii I ac ti ,i t,. The,* .uibj ected
the data tr lrerlce arsalysis and concludedj that !here .*:35 110[ an affe~c-
ti-.c d; ffus~an barr;er beltween thel eaternal subs~rite and! thrIe '.s t
ritich jubjrrates Lre res.pircd. n lt ra a ie I re-a n
JU-'t possiible, ho.ae;er, that sorch j mechanism L(act;i:e rr.miip:.rC) d~f
oi"erate in Use- ;bsen.ce of EdIIP, buc that in its presence !he irrojecules
Euga~r car~e Ic probat.1,~ the ii.ost .tudJied of higher plints ,n
regrdsto uga naecets.Biolass.. (27, p ?i00) state the uptake.
prole a fales:"...IL was~ (cun.1 ihat d.:iI:. i f ju,3di jltr [ I IUC
plj:.d i7 acrarted de cillEd .vate, lus. **er, lircle of their enjogenousi
sug1-r to- the *,ater. Tniis e-ither the~ toniopiast ir retr.:mel.iprm.l
to :ula.* c~.oge..*:r. 01 rhare I Le aC'ri acumuiLatin trach~?ljm in the tell
which a~;...i.el oppose:; the Cui r.Trr' Jlrrusi;nal mn*.ement of iu3dr.
The firot is per~haps r!h. jlapJ.p r Cplanatiucn, but ra~ier the irotica.
o: r-plaJi n i ng not. the rougar c r ignl, be,~ II; came arcumu~ilated behi;nd thle
Sugar cane exhibits a large, rapid (1-br duration, 8-min half
time), apparent-free-space uptake followed by a slow uptakte which can?
occur against a gradient and which results in sugar accumulation. The
accumulation uptake will1 proceed over a period of 72 hr. In comparing
rates of uptake of various sugars he found that glucose uptake was mtore
than double that of sucrose uptake on a molar basis. Uptake of fructose
was similar to that of glucose.
Bieleski measured respiration during sugar uptake and found an
increased 02 uptake upon the addition of sugar to the bathing solution.
He does not mention any change in RQ associated with sugar uptake.
Bieleski (28) found that sugar accumulation was completely inhib-
ited by 10-SM DNP, Phloridzin at 2 x 10-3H caused from 10 to 80%/ inhi-
bition of the uptake of glucose. Wlhen tissue was prewashed in 2 x 10-3M
magnesium chloride it caused a 0-20%/ increase in the amount of glucose
accumulated. Double reciprocal plots of sucrose, glucose and fructose
uptake rates vs concentration yielded straight lines (29). The Vmax
reported for sucrose was 0.7 umoles/g hr.
Glaszlou (30, 31) suggested that the outer space consists of the
cell walls and cytoplasm and is in diffusion equilibrium with the ex-
ternal solut ion. "Hence the cytoplasm is part of the outer space
where outer space is defined as the tissue volume which comes to rapid
diffusion equilibrium with sugars in the external solution (the outer
and inner space for this tissue may be quite different for solutes
other than sugars)" (31, p. 178). Tracer studies showed that the
hexoses in the inner space came from hydrolysis of stored sucrose,
Hatch etal. (32) reported on some of the enzynies involved. TI- ,
report characteristics of sucrose synthetase in the direction of su-
rose 5,n the~s;s. / i ~cti titi n th- rie-. rse J Ireci Ion coulId notr beL de-
Eccred becauISi. 3' thE presence or i phosphcetsse which r pidl, hl;Jrol; ic
UDP to UJMP. Stidence For the prrtsence of SUCrG~c--P 5,ntheLr.i 10 wras pre-
3enred, and acdJ jnd jl kaline in.ir Ljces ...reT dcr~CCTbed. The;, couIld not
f indr Sucratre phO ~hor* I as,] En_-jmes for the C,ntheSi-,, ir.te~.rce~n.*ersion
nnd breakdiot-en of hccoce phosphst tes erre identified. The Jamounts 3f
scld and ilk.alineL in'.ls*F.35i S .jry wri h the growth ruC and~ Zhe SUCrose~
scorarge rare (.33) suggesting a 1.0,? role- for insecrt.ise in rejulating rhe~
meo.esnen[ and utiliijtion of uCriioje. Sjcher ct al, (34) p~resen~t a
schemer for' the sugar jccurnuljtion C,cle in immirure 3ugar ceni. Acid
in.ertate occurs both in thce outer CpaCE andj rhlo jtorjgC iC~impa1rtii l'-nt,
Sucrose is h ydrol,-- fd prior to up[JIkE. and, glu~coJ i:. [al.en iip SC..ral.~
times~ us Last as fructose. Sucrose is re-leased tro.T. ;toljgge .;a
ht~drol,sis 0..3 dif'fu'.lon of the booses~t Outt of 3iOrage.
Ilatchi (35) dc--imontra'ted rhe pretence oi iucrose-P syntheta~se in
bjoth lefi andl storage tissui of 'ug-r cane. H!e also showedJ thL; 5,r-
thesis; of Suirose-P by tisslue supplied .4twil luccee. Scoea
r tored nmore ri-pdl, fewii -uirose tha~n froma suerose-P an~d Irere rjpid ,
froml fructose~ th;n r m :r3~ iucrose-P.. Thi5 ic CongiStrTCent~ wih t prop-
ojitionn thjt sugar phaseshares do not penerrat iembra.ws~ as eaiil, as
do non~-phas~osphor~lat suga,-s. Age~nietr, of labeled Jucrcse -a~s lo~st
during storage. Imrile onl/ :Inj.ll qu~anLt S of Sucrose~ --.er Ctored
wrhin iuirosc-P \*.J: supplied t~he s-mmetrr of labell wa: large lly na-in-
acie.Thi-; w-as coil. is tenet w*i h a sc 6.10 or, a-hichi ;ucro ,e-P is
formedc b\ thei actioli Of rucroseC-P c,ithetse ;Ind 5uiroseC i5 stoed
jgainst a sucrose :orcentrraicsl gridiene *.ia the hiJrolesi; of suciase-P
to ;ield Sltored iucrose.
in further support of such a scheme Hawker and Hatch (36) demon-
strated the presence of a specific sucrose phosphatase in sugar cane,
carrot roots, etiolated barley, oat, and pea seedlings, parsnip root
and potato tuber. The enzyme was associated with particles which be-
haved l ike mi tochondri a duri ng di fferent ial centri fugat ion. Mend ic ino
(37) had earlier described enzymes in wheat germ and green leaves that
included sucrose synthetase, sucrose-P synthetase, and a nonspecific
Hawker and Hlatch (38) present a scheme for the mechani sm of sugar
storage in mature sugar cane tissue. Evidence was presented to show
that the hydrolysis of sucrose is a prerequisite to storage and a rate
limiting step. Mature cane tissue contains an acid, wall-bound inver-
tase and a neutral invertase apparently located in the cytoplasm. The
storage compartment invertase found in immature tissue is absent in
mature tissue. Sucrose storage takes place more rapidly from hexoses
than from sucrose. Uptake of both glucose and sucrose as a function
of the concentration of the bathing solution had the kinetic properties
of an enzyme-catalyzed reaction. In studies on the localization of
ctlr, fir; ii e-r-.. foun j rlh.-lit .mos if n~o[ all of thef Iucr oser s,*nthat-.jse
s.=ar loca ted ir. rhe condulcr ing tiissue, .srd i r i.' poiin ted out th3t i t
Hatchj an ljridou IJ7) prtltented direct evidenic th3e sucrote i.
thet prudJo~nin=.at mornner~lrrs of traniloCatc d pjhow~iiTrthjre ilc. :ugarj care.
71.- jsi.nrncr ofr Ilabelrd Iucrore Ius. .ma~intained through theC siculjr
tis~c v th la. chath and.ter..Pardonizjtion did occ~ur duiring
3iche-r (40)) p~c~rll.cotd -.. .-ar!.-0.nln for e: [TractioPlelrnic I1ucro..
Unthesis in the bean endocarp. He supplied UDPG and labteled~ ructose
ind obtlained sucroise ;n wrhcich the labl *r:- predomin~atcly ;n the iruic-
lose moler,. Exper-imennts alrso indicated thE CprcencCe Of UDrPG pyropho;-
ph~or,ldse in the ex.tra3c;[ioplasmic spaci. \lthn latealed fructose nias
sulpplied and suicrol synthsisi occurred i r. he citopls asm~ ;he 1 glu-
case/114C fruitose ratio was apprsroimatell one; thi; sucrose remin~ired
in the tissue e'.Cn aftlcr eve~lnsi-.e w~aching.
B, usinS aceton-extracted chloropljsst from ugjr cjne-, Ha,, and
Ha~Sid (4I) wlere able 10~ ShOu the srnthesi5 OF Sucrose-P fromi UDPC. and
fructose-P and the synthesis of su~crose fromi~ UE=FG jnd fructosr:. The
preparjtion contained phc.5phatajses that h;Jrolyzedd sucrose-F jnd frac-
Schoolar and Edeliia.n (L2) mieasur d stcrete~d iLugar, CO., fi;xicn,
ficated oni \ar~ious solutions. The amount of sucrose rccreted intLo the
bathing solution wasr ineirsease by 10 ft s~odium lodoactate (10-'.
About one-third of the E0E81 Eucrose synthlsizcd during .j 4-d.i, period
ras sre re ted. The ;nh~ibi ter caused no change in rhe- amiount of soluble
sugar \,ithin th-e dist.s .nd i t caused an inrcrsi in the amount of total
soluble: phorns,~n:hate produlcrd. Faspi ration leasurcd in the J.ir).
shoved an FQ of considerable less than I.0 snd this wajs reduced even
furherby 0A.Otherr respiratory inhibitors did not ilicir similar
Mlany investigations hJ.te been made ol various cn;~mes in~ol:ed in
sugar translorm tionsr~. Onl,; : few will e ment~ioned h~ere. In h-is
re,ica arricle on jugar transfori..ations in plants, Hajssid (Iy,) jis-
cussed the chjracteristics of sucrose SlnthPLjSE anid -ucrose-P c/n-
thetase, the two enzymes most likely to be involved in the synthesis
of sucrose fromn glucose and fructose or from either hexase alone.
Putman and Hassid (44) studied the transformation of sugars in
vacuumn-infiltrated diskts of Canna leaves. When labeled fructose or
glucose was provided, labeled sucrose was recovered which was labeled
in both hexoses; however, no free la'oeled glucose could be found when
labeled fructose was provided and vice versa, an indication that sucrose
was formed via phosphorylated hexose intermediates. When sucrose was
provided in the bathing solution there was rapid inversion of the su-
crose with the appearance of hexoses in the bathing solution followed
by a resynthesis of sucrose within the tissue.
Cardini et al. (45) point out that the equilibrium constant of
sucrose phosphorylase lies in the direction of sucrose hydrolysis and
that sucrose phosphorylase has not been found in higher plants, A
study of the characteristics of sucrose synth~etase from a variety of
plant tissues is reported. The equilibrium constant, K = (sucrose x
UDP)/(UDPG x fructose), varied from 2 to 8 at 370 and pH 7.14 in dif-
Leloir and Cardini (46) studied the properties of sucrose-P syn-
thetase but point out the difficulties caused by the presence of in-
ber.tween sucrose conrent jnd jcid ins~ertise actritic,. cdinets
ac Cini; L: .,j high .~iur In3 cr~nas of hi h sugar usage snd 1-.r. dur ingl times
o~f hiigh -ucrosez sto;rage. The, sulggest that~ high i~ver~tueS aCti*ity
pl~re.n~i suirojSe -.Lorage and1 that during periods of 10=;1 hc ost: dema-nd
hydroljjts is due toj ~)alkalin ;ra=ertaic br-;hch ;s not, assoC Dscd **;ith
the ..acuille but locrated in rlhe c*;top~Fleam. ;he; jcuggenr rhat rhe -acidj
Inerrrl se ;j located ir. [hS rrll and~ ;1; the tienopl jgt.
YKurianl. ct al. (4j) ;omosied the locali4liat~innd p~r...erl.ies of'
herok~injje .rithl upatak chjrscterisre c~i ofco~~~cndutn rirsus ircAln sugar
bee-t. TIhis lissues takes; up glucos~' e rust!sr th:lrl irructjSF andc ihe: boIXi-
kinese- jssociated vi ch the; sru~ctural 2lements~ of the rells h:1s a highe-r
afillnit, for gluCcje thjn iiDir frlctose. On rhis basis rhei ;lugge~st
that. he.,so.irase on1 [h-- ilmemrin mre m be part of the uprej'.e process.
The uprjke o; sugars :nd~ the Inrsetl binding choairacteristcs r
;ee,[ I-avre been Srud;'ed ;ntensi.ely.
P.0thst-;n (5Sr' ~resents~ s.eral l~ines of~ e.idenTce LO sh;eu that[
uranl icr. aifecits the up[;Ike ojf glo-ICose I0, east due to: ;ts bihinfrg
to-. rhe zurface and nor io an uotake~ into the iatoplarSm of the Ce 11.
Rathsrtin and Mcier 151) describe the com:pedE~c on fr urBan,I ;or, b;-
rrran thre :east cor~plrrming Ir;:; .arrd rar'ous~ c;.-le;raq agents 3dde*Ul ro
the baithiig solution. On1 rne basis of rhis ..,rk [he*; cond.uded th::E
the" banding sires on the suresee 01the east cag[ Cllj rere pal~pr ophaaces,
UJ~;.-aii.. blrck.l jbour 40~.; jf the uotie of glucose in ,part I50).
Se-.eral other iorions, iincluding io2 ng Ci *,nd win ,trdt
rhe s1 'aice of y/fdit ells butr ujrarl ;or. fOrms ? much l.cori stable-
romp le- 152). Data ..ere presenr-. to Shiou r'lat .hreas~l thC other
cations rere bound to the ina ;rne sic hjt b~nrd ur n~il lon, rhG,- did
nMr Inhibit the upr!tak of 31ucose.;
Data showing the amount of various ions bound to the surface of
yeast cells as a function of the ion concentration are also presented
by Van~teveninck and Booij (53). They showed that in the case of
N2+ or Co2+ when glucose was added to the cells the metal was dis-
placed from the surface of the cells and appeared free in solution.
When the glucose had been taken up by the cells the metals were again
bound. If cells were first poisoned with 10A and then supplied with
glucose a small amount of glucose uptake occurred but was complete in
15 min. The amount of glucose uptake by the poisoned cells was the
same (on a umole basis) as the amount of uranyl ion bound (on a uequiv-
alent basis) by nonpoisoned cells. It was possible by adjusting the
growth medium to vary the amount of phosphorus per yeast cell without
causing irreversible damage to the cells, The amount of uranyl ion
bound and the amount of glucose taken up after poisoning were both
reduced in phosphorus deficient yeast. 'There was a good correlation
between the amount of uranyl ion bound and th~e amount of glucose taken
up by poisoned cells. When yeast was poisoned and then provided with
glucose the uranyl binding capacity disappeared. The addition of 10A
alone caused a 50%0 inhibition of cation binding which could be reversed
by washing the cells in water.
VanSteveninck and Rothstein (54) present an argument to show that
in yeast, sugar uptake can proceed by facilitated diffusion or by an
active uptake mechanism. The faci 1itated diffusion systerc-c r, bei
demonstrated wi th gilj; rosec upr jke~ by uni nduced cel ls and e i h g luii:e
thnl Cneryj re please b, !l,CC I,si. i r. sire of~ [he f at that gluc65i
,s~stms are diff~erent rrlth rellpect to I[Jln bind~na, iclFcr o fi. on;:
upE.-rke, concentration-, of uran,l ion requirid to ir~hitbit uprjake, k~inetic
psramerers, and pjtterns of pecificiti.
Fothui r n Andl Vanirtevennind : () sulnrnarized wrork. donc on, uIprake.
jnd mental b~ilding~ t, ,east cells. It ~as p~ointed out tha. r .e ;nisii~i-
Ctr, CfifcI' of ur?... I on jnd NliL jrl: n1ot du to diTFpClicefler of: i
requi red car for.. 1he corac lusion is r TCjh- d that II.5 phoCphOTr, I i tes
to which uran, I o,i binds, are used continuoulsl, il. gluicoi- tra..sp~ort
and are; rc'ciinerTa td cont i nuouslyI b; j, glCol ,''C. In the ,east s,rtem
a close corlciaticn is pictulred b~r~etwen gl~ccolsis and uptak~e ;rnd
gl~oi*, tic ATP ;E arsumetd to be the: encrg, source for dr;.ingJ uptake.i
Cjrrier jnd gl,Col,tic reactions are thought to be in close geegreph~ic
prox~imi t,. In their Chscre~ to Explain the trsr.5portt of sugars the
cjrriers for fac;l;tated d;i."usion and3 for icti.e Ltrnjport jre ion-
silered to be the s.-c. WIhen act;.e tranlsport occuri the jmouunt of
carr ier jmai l jble for fac il1itra ed difuior I' s rO Edu1ced.
Wheeler and Hancher, (561 p~laced oat roots into n. I and 1.0 mM~
uranil acerate for ..arling periods of time and then made- ilcctron
reiierogrbphs in rwh ch cristjls, apparenli, comlposed oi a ur;anium~ complex,
could e abl, be iecn. Afer A 30-mir. treaciment fo~llorwed b, a :0-min
desorpt~ion the u~ran.71 complex wasi sharply; local;zed its Ceii rrillk
irsteircllular spaci-t jnd secretor, products ir. dirctr contact 4.ith c~il
w~alls. Wlith lonij.r L~-tretment times, up to 50 br anid the iowetr concenl-
tration, thec uran*,i comTple. Crystlalb could be: found in .e~;cls in rlhe
cICt6rl~ater3a in the .j-uole, Othcrs-sise the ccleli \ere nomasl w~i t' no
uran,l ion free ;n Lth c~toplsmi or in ccall orgajnillei, Lrn o
applrent;, caused a deiinite dilatic~oreo the~ mmbranes~ from a normal
width of 90 A to a width of from 150 to 200 A. This effect could be
seen on the plasmalemma of treated cells and in vesicles which con-
tained uranium. They concluded that few, if any, free uranyl ions
passed through the protoplast and that uranyl ion in addition to being
bound to the plasmalemma is bound to cell walls and to secretary prod-
ucts along its surface.
Roseman (57) has reviewed the literature on a bacterial phospho-
transferase system that is thought to be responsible for the uptake of
sugars. The system as it operates in Escherichia coli consists of
three protein fractions: Enizyme 1, Enzyme II, and a low molecular
weight protein designated HPr. Phosphoenolpyruvate (PEP) is the phos-
phate donor, and a variety of sugars including some disaccharides can
serve as acceptors.
Enzyme I and HPr are found in the cytoplasm and Enzyme 11 is
associated with the membrane. Enzyme I and HPr are constitutive where-
as Enzyme 11 is constitutive with respect to glucose. Most Enzymes 11
are inducible. The specific sugar requirements of the system are due
to Enzyme II. Enzyme I and HPr are common to all sugars phosphorylated
by the system. Enzyme I catalyzes the transfer of phosphate from PEPP
to HPr which serves as a phosphate carrier. Thi specific Enzyme 11
then~ cu tl onesr~ 1~1-. Errinster ced" pl-.cr Fphate f'ri.T ph~or phate-li-frr to th
exagernO.Ou iuqri -nteir thli cell ias Fugar Fho~Sphates ar.J this i-. d-
proceiC ; ij.Cll-3 jcti'.e traiport.
Ctlp loocu Jureu'- jcTcullated SUCrose1-P when ;ncuba~ted in
sucrose anJ it i: thought ;Ihat the phi'SphotranSferaS' Sjiem is opera-
Si ve 'T thv uptuike aInd FhospihoriIh[ion (5.0).
Edeslmjn et al. !.ES) wo~rl.e~l rwith scutlle,; roots and shots of CoLs,
rpie, whea~r, jand borln:,, Thiey ShiJwed that the scutellum Contrained a
higher rjtior of salcr350 to heicose thAn did the~ rOOt or 5hoort. HomGe
absorcplion wasd ;r.hibltedd by about h~alf wheln e.-.perimieints Ivre run under
reitrogen. :ubstantial sucro~se form~ation rcok place ii, the scuttiliuni
under rlitroge~~r., whreos incorporation~ into jmino-acidr, Dr..Ides, mralii
jcid, jnd sugar Iph:-sphaeS waS co~nsiderabl, reduicd. In t1.850 li.Lu?5
frucrosP is ib~srbedl ;t aIbo~t half the rare of glucoir. ur o-
phates, sucrose, gluci;e, rruc tase glutarnic and asparrtic acids and
their ;miic~e,, mnalii acid, i02, jnd pol,*sacchairide wreT found to contain
label after appl,ing trrccr bmounit5 of labc led f'ructose or glucoje.
The scurlclumT C.js :hour.ll to contin I;Ichl lowei(r lE.elS of~ h~Cdroi;tc
enz,.ne= than th~e r~orl or Shoots. All of the en-,*nles necessal,; for
the fouraition of so.:rosi fror, herose wercr found ;n [be Siutellum and.
ir jchenP is presen~ted to Ehowr (hC path of SUCr.oSe 5,nthe-5iS wh;Ch in-
.oldes5 thi entimi sac3rose-P S,nthe tase.
Hurophre;s and Carrard hia~e publiihcd a series of paperj dealing
w~i h 110i uptake productLion, s toraqe and lealkage of Sugari b,. the
corn scute lIlure Thci dlemronstiritd Ehat glvcose Iiprsae pro~ceeded jt a
ionstant rare e.ea through glucose in [be bat~hing SolutiGO was Iargeli
deple red of gl~ucose ai ar risult of uptjlke (60). The racir of givCose
urptake ,ras shown to war,- depending7 on the conditions anid length of the
prior incubation of the L;lsue. Changes in the tissue content of
various csugarS and -.urjr phosphatss jfrer ,erving periods of tilie in
water were presented, and it was shown that mannose inhibited the up-
take of glucose. Data were presented to show that the corn scutellum
accumulates carbohydrate mostly in the form of sucrose, the content of
starch and hexose being low,
Experiments concerning the glucose-free space of the scutellum,
which involved measuring the amount of glucose in the tissue after in-
cubation in various concentrations of glucose in the presence and
absence of DNP and mannose and the measurement of glucose exi t fol lowd-
ing transfer into water, indicated that the space was intracellular
and that a carrier was not involved. Fructose and mannose occupied
a space of similar size (61).
When incubated in high concentrations of fructose (0.1-0.9M),
scutellum slices synthesized sucrose, some of which was stored and
some of which leaked into the bathing solution (62). The leakage of
sucrose was reduced in th-e presence of Mg2+, r,2+ or Ca2+ and EDTA
increased the leakage from the synthesis compartment (63).
When sucrose storage was measured after incubation in fructose
im. lurLITe .hit in =0und~i EMEi the O lufj IjCe 2n ut ation .T.@J urn un. .. re~
abo t a lo, dcre se in cor d ucrole air.-c Iruics c r jr .5 l;-. e-o-enousi
Irlltj ; el :1he-n Iu.:rilj e .ij -dIrJd rio the barh-,n.) ;;lu~ti j.. .:onr:.ni n..)
Ops.I.Tau' .. il''unts .JI' he-,it. ThIs *:.tul dj not look. beer, Ilhe iase if jn.cros
.i.-ru twi~n.; ~...Jr Cl ..=3 ,r:sr- r : I p tal~e .:.r .b rin3j [l process-i r upt-si'-e.
The loiSs from~ st.ragi e was mecasured b, loading thie Stora'ge CompF~rtmen[
uith~ o Euirose andj then ;ncubating the tissues at different pH \jlves
and ri th- ,nd wi thout "cold" su.:rose. Ilore sucrose wras lost jt the
higher pH Lalues and th~e loss was greater in the presence of "cold"
sucrose than in wraEtr ind;cating an erch .n7gi betueecn errornal iind
c tored sucroe.
The addition of Fructose or Jlucose to s~~caeilum sl~cice (L)
resulttd ;n a strong, aErObic Fermenration and the conccmmicant pro-
duction oF ethanol. Increase jucrose i,nthesis~ upo~n ;rcubation ;n
fruCtoseE accompaniid an inlcreasee In gl.iol.sis i cholrt an increase ;rl
0., uptake~. This sujpiorted- the ;de3 that tli,col,tic C.TP 11.ugh1t be re-
sponiible for sucrose- synthesir. ihen ;ncubated in rnrir thef Fi: ior
intact scutElla wras about ), while that for slices wras near Lnit,*
It rwrs cone lud-.d, on th~E bisis of the lete It of ad;Ous phosphO-
fructokinese regulators during d~iffrent rates of gl,col;sis, th~at
control ofi glcolsis ;n rhs sctiure 11.1 a c -crtetd through thea ;avil-
abi lit, of SUbstrteC a-ld th~e distritu;uton o~f aden:n- nuleoti~es and
Prcticatmncit s.'ith tr; S!hldro> .nethil I)aminlOnethane (trisi pjretenltd
the StorGTe~ of exog~nous su!crose bult the inhibition could be reserred
b. h1,drogen ion or L,- fit Mn2+, and to d lesser Ea tent tig andl (O
Sucrose storage from fructose was5 little affected b, the pretresatiment
rl th tris (66).
Pretreatruent wri h uran*,I ni trate (67) <--eS Simi lar in i rs effects
to pre[treament with triS in that storage of: eogenous sacirC-a was
inhibi ted and the inihhibitin could be rev-ersed by. H, HI+ and~ lo Soile
er tent Mrl21- Uran,l ion pretre~arment only slightl,- inhibi ted sucrose
synthesis from hexase and the inhibition was not thought to act through
the uptake of hexose.
It is possible, by incubating slices in high concentrations of
fructose, to build up considerable concentrations of sucrose in the
synthesis compartment. That this is sucrose and not sucrose-P has
been demons tra ted. Upon reducing thle external sugar concentration and
inhibiting leakage, the free sucrose in the synthesis compartment will
be transferred to the storage compartment. Experiments with mannose
(68) indicated that whereas mannose inhibited the storage of exogenous
sucrose, it did not affect the storage of sucrose which had accumulated
in the synthesis compartment. The suggestion was offered that the
storage of synthesis compartment sucrose involved a non-nucleotide
phosphate doner such as occurs in the bacterial phosphotransferase
Pretreatment of scutellum slices with HCI (0.01M) did not inhibit
the storage of exogenous sucrose or the synthesis and storage of sucrose
from fructose (69),
Recently Humphreys and Garrard (70) suggested that leakage is
from the sieve tubes and is the end result of a series of events which
include intercellular sucrose transport, vein leading and phloemn
transport, Several compounds, all of which can either displace or
form complexes with Ca2+ and Mg2+, protect the leakage process which
is labile at 300 in water.
Since some confusion exists in the us;. of v~ri;,ii-..o!, cjrcerning
uptake studies, definitions of several ter...s ui-j n the,? -r. -e:ntrn
of data and the discussion are given -.ere.
Ilpr~lak--Th 5 tcr-in i uJ d ijr the disapp..:ranc- e of a iet -.ace~
from the b3thing solution and ;r not nearnt to ;mpl, a parricular neech-
anirm.. 50oe duthors should uie abrorprionr.
Diffiujion--TheP net nso?,ement of molecules as a res~ui t 01' rhC i f
thermal nl.tior. Fromi a region of higher LO One of 10,.r0( concentrration,
Wrhzre a miembtrane 15 r ro ssd rE:istance mayI be due to the l im; ted number
and rize of PoreS in thE miembrane~ or tc the rolubilIi ,* C harac teri stics
of the jolutE in the Intmbrane.
Faciliti ated di fFusion--Thi s is a procerr in which a co~nitr ntratio
grajdient is the dr;.ing forrce as in diffusion and the proics s.lads to
a disapipearan~ce of thec gradien~t. The process is thought LC in.ol~e j
membrane conntitucrnt (carrier) locj;ed o~n or in the Is..embranrr Hhich
"fiac i li tatLs diffu srLi on. Faclitated diffusion of a soluito jiroir J
membrane; requircs ne input of' energ, other than that nee~de to na;ntain
rtructure. Itr me, showr a high1 degree of spC~Cifcicit and Ikinaticf are
likely,* to :hhou rjcurjtion thus n~ot conForl..ung to Fick'=. law. of d~iff-
Actrive- rransport--This process in*~.ol.S the csz of metabolic cncrg,*
3as dr;.;ng force. Ir. ls capatie of bringing about the accumulat~ion
of a SubStance. agjir=1 its concentration gradient. It ij ?enerally
characterize b; a hi:7h degree of specific~t, and sj~rationo kin-tics.
METHODS AND MATERIALS
Preparation of Scutellum; Slices
Corn grains (Zea mays L., cy. Funks G-76) were soaked in runninJ
tap water for 24 hr and then placed on moist filter paper in the dari
at 24-250 for 72 br. The scutella were excised and cut transverse 1,
with a razor blade into slices 0.5 mm or less in thickness. The 1i~ceJ
were washed in distilled water until the wash water remained clear snd'
then were blotted on filter paper and weighed into groups of from :*,1
to 1.0 g depending on the type of experiment.
During IrFpr jarain,, th- Slier vetre [hroutlhl nli)."*I C0 thjt gjch
groups o f Ille r .*6j: a r indon~ r I, le i onl fromn S :- 10 0 Icu rt Ill Thli
resulted i nr. i cI IeI 3cr.Earemnt rwhcn m~easurem~: nS Oi uptakei .:r &c.;u-
mu~~lationl wrITI .m..,dei~ o. upl~ijca rgrups~ 01 Eslices fr~~ro.. OeJa,'s prcp..-
ration. e uLts werET notl as. cons i i-tnt w.ht n dup lct .e are compaFredl
Urnlu:r cirherrwise nored, iniiubsticon r.Evre carried Out vi th 1,CI r
EralnsuicL icienrif it (r..npar,, rNew truns .ick, II. J.i rotating jt jpp.rorr
imael 10 e*amnThe :LOiu.e oi Soluti~on was~ Uujulli ]Td *il
iC m leS tare i nc la e ri(chi"J '' .r hr .;. i re w rIa inte tas p' "oI ~I I ! lr LO)E
rrucrose jrnd reducinlg disacchairides Here jnalyzed jccording to
thle rielson-So,~c?~ogCi cpper reduction michod I:71, 7-?) as reported h,-
5pio 13).The alternate copper Ieagent suggested b:, C~omog3 I ra
L'hin samipling the: barhi~ng solutio;lnS for 511gers, amorunts[ beCs**eer
0.1 ain 2.0 mi wrer tak~en depending on rl~e ioncenrr. ti~on, and appro-
prjiat d~lutions wrerc madr so thar betueer. O jnd 140 ug of gluio:e or
fructose erecr used fo~r an1al,sis. T.Jrice this jmount wasr usd for disac
ch~arider other tha~n rucrose Ab~srban~c e wa rejd Or, a F~.letLL~t-Summeror,
flaoel 5.005 photoelctrirrc colrirrreter.
Ti5Elue 5ucro"e r:.-s etr[a[ted b; pouring 20 ml of boiling 20,:
ethajnal over thr cliCes and cOntInuin~g the boiling fo~r ;0 sec. iThC
slice: nrre usrepdo in thc jilcohol folr .0 nain, the allohol 1-35 dectntifd
anid thes pracedure repeated.j Thi 5 li cs r -*6re ther. rinsed three I ithn
rr; h 5 mi portlan of Citahol. The com~binecd C.*.LraC~ing solut ons wrecr
evjporated almnort to drincrs oil a steam bath. \.'ater vra-: addcd to a
voilumc of 50) mrl jnd 10 darops of 0.1.* Ii Ia0 added ro adjust thes yn. Th~e
Treulting ago~us solu=:ion~ was~ frozen, Afrtr rthandiig rh 50 uri~l n U.9
ccntrifuged for 10 min ir. j clinical icntrifuge. for the de-tereda;ritio-
of' CucrsE~, 0.1 fr1 olf this jolutio~n wa~ u~sed.
Experlments welre carried out r. I warbur? DEspiremlCetr ati j30
Thle direct method~ for CO, uWE used 174). The amount of rissue added
to the flasks~ <.as either 100 or 500 iig. when the sli~cs werrp prcpared
the,; sriE placed <-.i thout weighing~ ;nto 25 mi Eriennme,er fla;ks in 10 mi
of rater and incubiated for 1 br at j00. The wrater incubatiorn remo.en
leakatalr sucrose (64). Foliclw:i ng the i ncubtjlor, he slIi ces re re blot ted
and weighed into Warburg flasks. The sugar solutions were either added
at the time the slices were placed in the flasks or added from the side
arm during the course of gas exchange measurements.
Uranyl ion was determi ned by the method described by Rulfs et al.
(75). Absorbance was read at 400 nm as suggested by SilIverman et al.
(76). Aluminum was determined by the method of Gentry and Sherrington
(77) as reported under procedure A by Sandell (78). The extraction
was made at pH 5. The purpurate method of Wi lli ams and Moser (73) as
described by Sandell under procedure A (78) was used for the determina-
tion of calcium. Magnesium was determined by the Eriochrome Black T
Method (80). The permanganate method of Nydahl (81) reported by Sandell
(78) was used for the determination of manganese. Cobalt was determined
by a modification of the Nitrosc-R salt method of Marston and Dewey
(82) as reported under procedure B by Sandell (78).
The methods used for uranyl ion and aluminum are not specific;
however, since samples from untreated controls showed zero values
interfering ions were not present. The methods used for manganese
and cobalt are specific. Magnesium, in amounts that would be present
in the solutions analyzed in these experiments, is reported not to
interfer-e wi th the purpur-ate method for calcium. Copper, iron, and
manganese do interfere to some extent. The value obtained for calcium
in the control samples was low but not zero; however, some calcium
would be expected to leak from the control slices. The method for
magnesium is not specific, but the amounts of interfering metals in
these experiments were too low to cause significant error. The aRouTilsl
of calcium and miagnesiumi flounj b, u:ir.3 these methods agree very cloel,-
\rith tho~e found b, ato~mic absorption FpecCtreacep, in tearler wlork b,
ilumchr-r s r~d C..rrjrd 1.70).
In the first section kinetic data will be presented to show the
rates of uptake of sucrose, glucose, and fructose with time and the
variation of uptake rate with concentration of the bathing solution,
it being assumed thiat sucrose uptake occurs without inversion. In the
second section the assumption that sucrose is taken up as such wrill be
justified and the effects of several inhibi tors on7 sugar uptake will be
presented, In the third section gas exchange data will be presented
and it wi ll be shown that fermentation accompanies sugar uptake. The
fourth section presents the results of a study of metal binding charac-
teristics of the scutelluml slices and the effects of several cations
on the uptake of sugars.
Figure I shows the cumulative uptake of sucrose from two concentra-
tions of sucrose over a 2 br period. In these experiment lth fiirs
sample was taken I min after adding the sugar solution =.0 rte iliice.
When the bathing solutions were analyzed for sucrjic .j s..all
amount of glucose was usually found except in the sample. I; Latr, atI
min after adding the solution. This glucose could hav; cvnli friom the
extracellular inversion of sucrose cr from glucose difiuilrng cut of
the tissue following Intracellular inversion of sucrose. TbeIso-
the data from which the upper two curves in Figure 1 were calcull us.li
The rol;d lirr in FiguLre 1 repf858Dts datj cjlcu~latedd (froni- clui~nr.~,
:Ti rise red n
Figur L urs pceV i\e. Oego lcsntue
inf eah la n T lice ee icb td frIb
in a-arergiven I rin e, an then 10 mi f m g r
th as 0 min ha er eue o 80n
~I_~ __ __
Method of Calculating Data"
Time Without With
mi n invertase invertase
Values shown in Figure I were derived by multiplying the figures in
column 5 (dottedj line) and column 6 (solid line) by the appropriate
Tab le II 'a; i gnor ing, th.e non- iniert.1-.e -It reted complei hic.
i t i s asumrd rhjt glucose i s comin.) frm~ exrale li11lar ire.ir:ion Jnd
thel ~Lu:rose` inverUtFP but notI taken.T up i i notL counteI as i5urtiak~. The~
dotrzed line repres*:.-nt. the actual acreasej in :ucrose of rhe =olurio~n
(ca~~ljculJe froml column ',, Tjble 1.. Th~e two Is thods J0 no: result in
large J;iffrences in calculated uprake. Table I is presented~ to deI--
unstrzte t~cime method of m.eaiuring jucrose uptae. he..1.3n-r
'uLroSe uptak.e data p~risented~a ..a alculated t., thi .nethoJ r:Cr~:esened
bi rhe solid lines,
'Ihen thie rinet- ursrr unsc rrichd~rjaun bi' ucrior, i.Tun~edi-sal b.3fore
.adding rlhe sucrose, an uinme-surra s~nou.e o~f .rater adheircd to :he slice:
andl EG :he sides o~f the F lass and caused a dilation ofi trl; Ijdded jugar
solution.. There Is probabit also j r'ree npjce .coluT.. in t~e slice
which causes di Hlrion Co rhjt jnal,*Sis of the r'irst sanples i ndiclted
3 sucrose concenirtratin cornsirc~rjbl lo--aer than that vbi~ch .-,a in:Elalls
a3ddd. Thiis decrcr.s-- ;in con~centlratin can be Jccounted for by dilucinn.
In Ihe i-rfpejriment Of Figure 1, rle iconc~tientrno I mirs after adding rhr:
sucrjse <.asr 0.00fa.0,I whe-n 0.00j':O00 iucroce was jddedj s.]n 0.0132--#.1 Hhen
0.lr)150M1 S..acTros was5 adae.. These c<..rn~rentration canr be accesi~ired sor
case of 0.002500; :..**ro~se. The E p~ro fojr 0.0050%SJC sucrose :would be Imr
ai~l, f l eif frt wis in~de ro ~e~ter~rnine thet amount: o~F vlatr ,nd Eree
space. Thet problems was gnored byl r..e~asuring upr.--'tl *"rr~m melc time [he
fisrs ssample rwas r.ake. Thie ij.[3 Fare prasanted to Ihr.r that !ther is
Iict a large, rapid: pooJr Jf uptal-e -men :uCro.e i. FirSI added t, rth
The~ rat1e oiir u tal..e ;ocrease= iifter C:Ie firTC 3 niiri. 6.t hellr Lhe
rate gradually increases over the first 30 min or there is a delay be-
fore uptake begins. In subsequent experiments the First sample was
taken 15 min after adding the solution.
Notice that the rate of uptake was constant with time in spite of
the fact that the concentration of the external solution was continually
reduced. In the upper curve the concentration at the beginning of the
last 30-min period was 70%/ of that at the beginning of the first 30-min
period. In this same series of experiments a constant rate of uptake
was obtained with 0.001M sucrose (data not presented) in spite of the
fact that the concentration of the external solution was reduced to
40% of that added.
More experiments were run to determine the rate of sucrose uptake
with time and the erffct of sugar concentration on uptakte rate. The
rate of sucrose uptak~e with time is shown in Figur~es 2 and 3. The
curves are more unifcrm at the lower concentrations and as the concen-
trations increase the curves become more erratic owing to the difficult,
of detecting small changes in concentrated sugar solutions. A water
control showed that the leakage of sucrose amounted to no more than
0.2 umoles per g fr wt over the period of time during which uptake was
Thi. betl app~rOii.ne~tic-r. C\f .ll GIj [hL cur~i; 3 ;-earn to: bet j trlraighr
F; 'ur 4 sho**C (ht ffe;,[ .:.t iucri*(.F i~cC~=ntrjtion Or,n iarote UpI-
perijd j rr rrich uptake~ <..az re:asur ed. Ais seen t, the twior ctical
CUrse,~ the djja~.: cTI r, <...II ll;h the tyIp;Cjl rliihis*-li and~ herntcin
h*pertsolic subotrjte concentrats.-n cur.<- foir which h thF iOnItants werTe
deri.ed r'rcurs j L~ncij..co r .an Burk, pl.:t of the data,
F i ure 2. Sacrros? Uprtake Vs Tarne. In thtse emptiiirianen I -4 of r I ce-,
vwjs incusjted ;n water for hr. This r1as I:licIEed b;
2-13 c.! rinsr; and then the~ upralke rDlution v*LL add~d to rh-
lis.Tcn mi or' uptak~e C0'uian~ wasj idded sild th: arriot~nt
of sucrose remo3 0.0 in tii 5olvpling was taksn intou jaccoune
<*hen r:. 1 cu la t ;nr the~ du e Th i e e r.p : >= '.'.n
Ii~n jF ter jdd; tion oF th-e upta's solurion I.ir..s _. rr- on r1.<
?I jph).Sml;0"05n wr e !r S.i. Each
L.u..E rieprscnts .jr.a f roJ.1 1 d:*,' cx.
T i mie mi n
lii3 0. IIt
5 0 351
45 mO 1n .
Q 0 9
m -- a
Jud jon.1 .].]d. O...r._.-
The erp-rim~ent or' F;igur 5 ShlorS Sucrose5 uptaker; wh~an the btjhing
solution is rmain:oined at a constant sucrose concentration. In thi:
ezper ir..ant th .olumae of the t~athing Folution was reduced to II ml fo.~
greajter accuracy. At the end of i.ach 15-m~in upt:4l.; periodj th- solution
\as rzinoved snd fresh 0.0018I sucrose wras a~dded to, the slices. Th~e rate
of uptake cai be seen to increase v,;th time c.er a period of hr.
Figures 6 and i shot** the rate of rjlucose uptake \th ti ime. As
w~ith suirose, the glccosr uptak~e ca~r.es roughly, represent straight l~ine
rwith possibi, a little miore tenden, for the rjtes to iecre~asL a~ r;t
tIme. In thet case of 0.005ti glucose thc concentrratin of the sugar
solution <*as reduced b, about half during th-? course of th~E erocriienen,
,ct the A~te olf uptae c.rr the lajst three: periods Was about thie rjise~
Vler; simil~ ar data uere collutedd usin3 fructose but~ rher-.. are nlor pre-
Figure 8 -hou~s the iptalke .5 concentration datj for glucoji and
fr uc tose. n trne iase of gluiose and fructose thle data do not fi t jt
all urhen an attempts ;s ma-de '.0 find the constants Vr..a, and 1:rui bi clort-
ting the data according to a Linewcaserr an~ Burk plort, fleither di the
data agree Ilith whatr would be expirCte if difiusiOn wepre the drivings
force for glu~cose uptt.lli-
Uranyl ion has been jhown to inhibi;t sugar uptalke 1,67) jnd to~ act
at the cell surface (5.5). Uranyl ion caused a partial inl-ibition of
glucose anld fructoSe uptak~e. This was true both e-henn uran,I ion ass
added 10 the uptake solution and wvhen the .lices werre rr-alted r:;th
uran;1l ion prior ri. the uptak~e period. Th. dotted lIn;:s inl Figure 4
show tht uptakLe vi concentrrtior data for glucose and fructose a-.hen
uptak~e -;at Ireasured fo~l loving a pre-treate~ren rri th urar,l ion. Tat..e 2
Figure 5. Sucrose Uptake at Constant Sucrose Concentration. One g of
slices was incubated in water for I hr followed by 2-10 ml
rinses, 4,2 ml of 0.001M sucrose was then added and 1 min
later a 0.2 ml sample was taken. A second sample was taken
15 min after the first. Then the entire solution was removed
and 4.2 nl of a fresh 0.001M sucrose solution added. This
procedure wars continued throughout the experiment,
30 6090 F2
Fiue6 lcs ptk sTm. Tedaafo hc hs
cuvswr rpae oefo tesm yeo
exprimntsas hon i Fiure 2and: v thth
exepio thtguoewsth ua o r
/ 0. In n
Ti mir .1-
Figure 7. Glucose Uptake Vs Time. The data from -thich these
cur:cs werie p~reparedJ comrie f'rom the senser r.*pe of
periments"[ ac sh~or-an in Figures: 2 and 3 \-si th the
BKCeptican th5t glucrOSC \*JBS thi sugar Lak~en up.
, I ~.. I..L. ... .c _
.s t,': 0 n d :o H
I a d
.- a .
r OOC 00
cau a. . .-
C LI o ~
a .-- ai
L U i O L 5
E P u- >C
n A- ~ .- C
Cr a-c .,
-t L. C -
O~ - 130
c j .-
o r le OE-r.
4.- i D *
I -' C~ G. L E
<[ *** d d
(1~ c-- -- O
. 00 O
La O 1 T I
d~ 1. 3~
-- C D
C ~ ii .0 -
concentration Uptake period Inhibition
M hr %
.001 1 58
.005 1 54
.005 2 53
.01 2 44
.01 3 45
.1 3 34
Glucose Uptake as Affected by Uranyl
lon in the Bathing Solution
jhumsj the effect or' urjin,l ;ion on gluccst uptake'r when urinl;, ion ,.aj
adde~l to ther ulptal e solution, Ilocuce that in tooth pretrreatmorst anid
crearrentr during upcjlke, th-e inhib;ition ras greater at low-.
[;ons3 of Ilucase. The in~hititioni vas5 nreSte when the slices <-.ere pre-
created it urAnI,I ioni. Th is ii perhip5 .I result or' j long~ trm cr'fect
or LranI ion iinic in rhe prelrtreament apesrimentsit uran.l ;oni waj first
applied to~ the jii.:es 105 iil.. prior to the uptjlke per;od.
The glucose uptakle rs concentrjtion cur=.e sitir uranyl ion prc-
trejtmenti rcjsembled, at least at lo- concensrurioni,, a diffusion curse
(Figur~e i). Since glucose is niot dccumulatred b; th.e ..issuc bL ;s
rapidl, used for sucrose J rrlhesiis or ;s (frmanted (60. 65) thi question
arises as towehrtekntc rsne re tose of uptake i.c.
th-ar .glu~osa is. limliTed 'u, diiFUsionl or thojd 01 [h2 hb.-okinese: res~:CiOn.
Jone.. 1,83 studied r1.0 propertiesj of he:
and .rports che Kr. for glucose as 6.5 .s 10-61'.At. o hsmantd
indlcarcs rhat difr'usion and not rhe hiok~inaser reactionr i. linic~ng,
I~f dFfuionl ij rhs. dr;.*ng rorce for uptake and ;f :t ;r s Mr~uwl thje
[Ihe inte~rnal FlucileC cr.cencratio.-, remdins constant anI .er,* 10.? Ehecn
the~ u ata.e should be j srra;ight lIne function ofi thr 3lucate con;'rn~raj-
tion, ** straight line rai, ;1 fact, obtained jt cojn:cntralioior: of 0.OiMi
Und belour. The de.*i~atin frcal lIn:.;rit, at nigi- e* Oncentrat~on mTayi
be due1 to hiviedr intri'ral conclent."3Eijan s 1 i-i jIos- adl: EO- a Si~Uration
of the ?lucose ucilize don procejj .at thet '-iiger concentrations.
To cli=:R this hlpo~lchS;S furhthr CRe rjte of u,.ca~l.e ;nto urran,l
treated ard untreated -lices' *;os follou~ed ith [inic (F;~luit 10),. ).1
the care at glucost alont the uptake Proiced~ J a alm~asr ;1 .:.~rstent
rate u~nt I the 9;oc31e in t",u ''Ctake- 'olutio~i had ec~ 1 depil-!c.! co a
Figure 10. Uranyl lon Pretreatment and Glucose Uptake. The pretreatment
consisted of I hr in either water or 0.003M uranyl nitrate.
The slices were then given 2 rinses, 30 min in water, .ure
rinses, and then the 0.01M glucose solution was added.Th
first sample was taken 8 min after the solution was ;dded
and samples were taken every 30 min thereafter for a Ftri;d
of 210 min. The curves are an average of 2 determinations.
The volume of the uptake solution was 6 ml so that lbarne
differences in Klett readings were noted over short thi-e
/L tr pre tr a r -a t
12 r J0
P ~~rPi U~rlln ior re lr t.nin
60 120 I80
T i c..e mi;n
level below that of the detection system used for glucose analysis.
This curve cannot be explained on the basis of diffusion alone. It
might be argued that the rate of utilization of glucose by the tissue
is constant and that the curve simply represents the glucose utiliza-
tion rate. HowEever, as shown elsewhere by both uptakce and accumulation
data (Figure 7, Table 9), the uptake mechanisms of the tissue are not
saturated even at mnuch higher hexose concentrations. In contrast the
curve after uranyl nitrate pretreatment is typical of a diffusion curve.
If an arbitrary constant is multiplied by the concentration at the
beginning of each period of uptake, the curve represented by the dotted
line is obtained (Figure 10).
It is postulated that glucose uptake is the total of two processes,
one consisting of simple diffusion soon after uranyl ion treatment and
one an active process which is subject to inhibition by uranyl ion
If' this is true it means that the active component as represented
by the difference in the two curves in Figure 10 is increasing with
time (1.e., since the external concentration ii d creasing, diffus ;or
is decreasing and the active component must b~ inireaijing in ordirr [G;
maintain a steady rate). With this in mind jn cxpiriment -a run ;n
which thle concentration of glucose was kept c.rnety~t b, inreneir~3 the
solution after each sample as was done with fucr;se (Foure 5). The
results of this experiment failed to support the b..othts isFigene II).
Glucose uptake remained constant with time ir. sFit ofc~ the fact t h jt
the concentration was kept Ecentur.. The up tale C.r aj per~iod iof 2 b
was very close to the value obtained with~ delinlr., ?lucose ianicntrh-
tion. When uranyl nitrate was added to the uptale ;;lut~ar. ir. thiE
Uran, .o, ded ,-
~ ~ ~ ~ ~ ~ s~ ..iad~- rnlin d iezr
Tier ,i ot minl
Fiul I. C uc se U tal a C n ta E(o ce tato is u \/
sub cte to I h nw tr inc ,ad te h
0.0Er upJ:slto s. Te fr tCopew s rle
I i ftr ad ng te sl/o Te.L 'ue ae
a ecndsapl ws akn.. n h ptk outo -/
use~Ilrn,I nitra jte. The uptke o lm a mi
experiment the rate of uptake declined with time. This may be due Ic.
a long term effect of uranyl ion. The amount of inhibition caused by
uranyl ion was the same whether it was added at the beginning of the
uptake sampl ing or added after the tissue had been taking up glucose
for 80 min.
An experiment was run to determine whether or not the inhibition
of glucose uptake by uranyl ion was parallelled by an inhibition in
the amount of sucrose gained by the tissue. The slices were placed in
water or 0.0038 uranyl nitrate for I hr, rinsed, placed in water for
0~r~dr. rin rd, and JLIaCedI ir. .hIl fructose or 9lucse forJ ?O bri star
whi.::1 ~ ~ ~ ~ ~ ~ ~ F~b th sne:vreritd n klldfo .r5 i lj 01 Einu ;nucrose
i [Cd j'. .3~.-.d 4L.: .
le. Order toj 5Fud sucrc.:e: ult=Le ir should ter urlatbl;; hed !hj
iuicrose :s .uch if thC iulphr b9;nll !Bakeni Up 6.rl ie f .dj
v~ill be preiSenced FO ih~ru thaE suirose ; rla tnkr upF witho:ut in.trsio:n t,
rugar uptake- willI alo be promoted- in thii scioLCTi n.
The Sii.LIutI of ;r..ersio of suerC.:el in [I.6 .-r erma~l zo~luiusl i j
Jiinal jI n.:.32ured t., the spearancer 01 riluco'se ;n the So~lutior~.,
ir.itarli c it .. I.ucIro: e cors.:eat~ji ion l i. 0.0011 [hl r.6,,irlurel .j,.:OUnL 01
gluc;.e noted in the iolut~ion 0.1 0.000i')l. The amount .Bried~ o C-r the
upI3Ce Ieriodl frossI Ehli ..alue LO .30 Samun t [.10 th 1 G~it 01 dtEDrrso
folr [lle daunt oF wlr~plee used, Thi; ..ould mean a endsamliumil I-evote con-
cenrration of (3.U!Jlll Jnd an uiptake scordi, eg o F;gur- .: cl' appro .-
rought~ one~-r~-nth of the rate o~f sucr~ojs uptake o~bsei.rved (njule 41.
This jrigumernt jip3it S :IheL jSSUmptiion,. justifitd by Figure i, thit chrt
t*-so heroses ;Ire t il.-n uIp .t jboutI thJ iameI rtj* hi rutC.l a
beenr piaison ced fore b i Humpnre ys jnd Garr ard (6~4). The possijbili t?
- ..;-.s that In.ersion [jlkis plac: in J posi rion iuch Lhai he:-oses mJe a
prefertnri-all/ toward the point of uptake,.
Ki;ntilc djta on tlF: uproae oi sucrosi glucose and fructose rwere
pr1jnted in che first rition, TheJ on.erjll s~hapes of thr, upl.JI race
. j .ncent ration curies~ is co~ns i drab ig different. \lnesreas herose
uptal..e iicrt as es ..irh co~ne..lrrttrati ove.r the ranger sh.:.L.n. 3uslojse uptake
a~pproaches a ma.ximumi. The conrsiderable dirfferenc in !he ef'iett -of ion-
ccntrationn on up~Ale if an ;ndi-ation that. rv) different pro~cesses ire
B, concering the data in Figures 4 and E. it can; De sen thjt the
total amou~nr. cf carbt~n rjlkn up fr.1,i 0,1 11 solution i 3 highe r when her.-
oser .re suppl ieJ. In e.per;i.Tsent iiearuring c'jssue s*.crnse It <,as
notred tlhar rih amounllt of sucrose ga~ined b,' rhe rissue was thi janme or
highel.r hcn th lices j U.:5I.-eF Inrcubased T.~ :Utrose as iJompare]J to ;rcuFba-
r i on in hthXose. ir. Ordr Eoi shrrk this effec: Onl ?he sarre grculp oF
lI;cesj an I..pel'rine~! r-t wa run in; .hichi uptake wa~s alloued~ to proceed
Dual;cact flasks ..;ere rur, UpF~a!..o waS5 mejSuu c o.er i hr of :hi .Jpcjle
proThe amount ofr caJrbon taken up fr'rn. giis-acese sltir on~~ was 1 O
of rhat taken up from sucr'ose solution although th-e snount of ucrose
gained by the slices was the same in both cases, The slices were in-
cubated in water following the uptake period and then rinsed, killed
and analyzed for sucrose content.
When gas exchange studies were carried out using 300 mg of tissue
in the Warburg flasks it was found t'nat the addition of sucrose, glucose,
or fructose to the solution caused high rates of fermentation to occur.
It was assumed that the amount of 02 consumption represented sugars be-
ing completely r-espired wrhereas the amount of C02 evolved in excess
of the amount of 02 consumption represented sugars being fermented,
It has been demonstrated previously that under similar conditions
ethanol is produced in amounts equal to the excess CO2 (65).
The data in Table 3 show the amount of fermentation over a 2 hr
period caused by the various sugar solutions. In the fermentation
experiments the slices were incubated in water for I hr in the water
bath, blotted, and weighed into Warburg flasks which contained the
sugar solutions. The flasks were then attached to the manomneters,
equlil brated for 15 min and t:he readings begun. No effort was made to
the begin.1ir.) of the readil~ing
Th-- r.Ee of resp;rtitoo~~.i :: rc.3:crabi cor..irrntr regar3J1. of the
h o fa combelnricn rri r. Th r i i '-.jz ;n .uicro~.-- n t r so ..o e u l
pe.r bl;C ..:omp.re;~j [o -~ j; rate of 5 u...elesf for tHlc:re.- Up~t-li- of1 3lucos
a.j-..i ;.u TC-se~ fr~oli 0.010 Iiolutionl b, th.C jjir;.06 :i~iices. ga he
followr i lg ri. ults; uro~t 17 uml es/hi~h r (c~r j4I ,,,,c-lc: h; o. -}c, ?lucose
21uale/H.Il' the jToun[ oI' termecnrjtion is a ful'.:rior. I:r the coc.-
RespFi ratei in Fri Tnen ta ron
clmo les u~nle: I No r. ofi
Eol ut i on li.oe/rher~se,' nr prmn
(Djta j.erjgzd frolll se.evrrl dai'i preparjtio~n;)
W.'aer F.8 (0.4): 4.j (0.E) 5
0.01M suc roe 4.5 (0.41 1 Lt (1 .9) 5
0. 11 fo.to e4. g (0. ) 120 (17
0. 00511 fr uctore
r. 0.000M gllucos 53 0.1 13.8 ( i.2)
0. Irr Jucroje 5, 2.5
0. I1 fructase 5.0 (0. 1) 36.F (LIE.) 2
0.0 Ir lucose 4.6 40.5I
''Nor..ber s in parentheses'r indicate the j.l.erjag draiation. See 10/[, read
materialr and Int~atod;, for er.rprilrental detl I,.
Fcrmentraticn ir. Water ar~d Cusjr folul~non
centration of hexose in the fermentation compartment and if sucrose is
being inverted, even in the process of uptake, the rate of fermenta-
tion in sucrose should be higher than it is. The sane argument can be
made in the case of 0.1M sugar, since the amount of fermentation in
hexose solution is proportionately greater than the amount of hexose up-
Further evidence that sucrose is taken up wi thout inversion comes
from the effects of uranyl ion on sugar uptake. The effect of uraniyl
nitrate pretreatment on the uptake of hexoses has already been giveni.
Table 4 shows the effect of uranyl ion on sucrose uptake. Uranyl ion
more completely inhibits the uptake of sucrose. UHith hexvose the effect
is to cut the uptake roughly in half. If sucrose were being inverted
prior to uptake, the expected effect of uranyl ion on the basis of
the hexose uptake after uranyl treatment curves would be to cut the
uptake of sucrose in half. It might be argued that uranyl ion is in-
hi biting the inversion of sucrose but the inversion of sucrose as
measured by the amount of glucose found in solution is higher after
uran~yl ion pretreatmer~t than it is after water pretreatment. In mea-
suring sucrose, It was consistently noted that the non-invertase-treated
sample gave slightly higher Klett readings in :he cases where the slices
had been pretreated with uranyl ion.
The effects of anerobic conditions on the uptake of sucrose glo-
cose and f-ructose were determined by incubating slices in waei~r fo.r
Ihr and then transferring the slices to sugar solutions and ir.iutnling
in air or under nitrogen. Nitrogen was continuously bubbled clhrcshh
the solution. ;LucroJ Il ..pacE in Ng2 was only 33% of that in olr r.
wi th gl IcoF 3ndJ frucae roi .r. rates cl u'~ptak~e In 11, were 65% of~ U.l:ez
0.001 4.1 0.i
The Effecr of U~ron I rritrjte Fretreatmen.. sr. Sucrose Uptak~e
Ulptale uiinnle: hr ~
Uran,l I ntrate
Slices- (1,0 3 Tr ut) ~ere pla~ced ir. iither us[Er or liran,l
n~itrate for I br. The sli~ice were J;:rn T' 0 ter r;n5Ss p~~lace ;[
rater for 30 Irlin, gi'en r more rwater r~rirnse oind f;nall, .laed Ir. the
batLhi ni 50i utlon. T.*j sampilrs we~re tsken. rlhe firal 15 Pin sc rrr the
bathinglr so;utionT \las addedJ jri thj SecOnd .11 the end of the uIptl.F
pe~riod. Uptakel~ was measured c..er a piriod of ii0 mTn .ri tr. I.00111~ eirtd
0.00)511 scrosc 90! min .-al th 0.001l iucrose .-nd IIO min url h 0.05 ;nd
0.iM CuiTcee. PeteS of Upts~.e in the first thrte are o.:\rajse of the
rcsults 01 two~ e.periments, [hC las~t t1o are Iro~m o~ne rgriment.~n
Table 5 shows the inhibition of7 sugar uptake by phlorldzin. At
lx 10-3M this inhibitor caused aboui twice the amount of inhibition
of sucrose uptake as i t did wi th glucose uptake. Witeh nei the~r sugar
was uptake inhibited strongly by the low concentrations used with
Dinitrophenal (DN`P) also inhibited sugar uptake (Table 6). The
inhibition was considerably greater with sucrose than with glucose;
the sucrose inhibition approached 100% whereas the inhibition of glucose
uptake approached 50%/. sucrose uptake in the presence of lx~ 10-4M1
DNP was l inear with time for at least hr.
The uptake of sucrose was measured in the presence of several di-
saccharides to determine if the uptake process was subject to inhibition
by molecules of similar structure. Several of the disaccharides caused
an increase inl thle non-invertase-treated samples. This caused confusion
in interpreting the results since it was not known wh-ether the increased
amount of glucose in solution was coming from hydrolysis of the disac-
charide or from an increased hydrolysis of sucrose caused by the pres-
ence of the disacchalride. Unless otherwise stated these experiments were
run by incubating first for I hr in water and then placing the slices in
0.01M sucrose plus the various disaccharides in concentrations of 0.005,
0.01, and 0.05M. The uptake period' was- Ir.
Lactose was not taken up by the tissue as determined by measuring
the lactose concentration before and after I br incubations by the
reducing sugar method. When solutions were tested for glace-se the
amount was found to be insignificant. When sucrose uptakl.r- ,, menure.1
in thie presence of lactose there wras an inhibition of 33% i. rh c;:e
of 0.0EM lactosee ir i t is assumed that the higher glucose cone~F: ir.?m
Concntrto Inh it i [ i o
ti Clucose Su. rose
I. 10 1 I 25
2 10 -47
twrice, and placed in a solution of 0.0l1M rugar an~d phlorld.Tin as in-
dicated. F-esulis jre based on a non-inhlbited conrro~l, UpcJake as~
ns:-asured c~ar j L br period.
Inhib; itin of Sug~ar Uptak.e t., Phl;oridzln
The Inhibition by DNP of Uptake from 0.01M Sugar Solution
of DNP Glucose Sucrose
1x 10-51 2 31
3 x 10-58 14 51
1x 10-4MI 35 89
3 x10-4 M 43 94
The schedule was the same as wi th phloridzin. The uptake per iod
was 90 mi n,
the increased h,dial islr of ucrose in the iresence of lac rvte. Ir.
O~thJI woirds. ther rame~ jGrwunt of SucTrofe \iaS iou.10 it thi endC of Lthe
uptaike period vrith or rrithoIut lac toSC. There was mOre qluCOLE in SO u-
tion at thi colo of the vp[ake period w~hen using sucro~Se and lacrose
than ~there unas in the cas6 oi lacrk*Le or sucrose ljnS. If this glucoSC
came from h,drolesis of sucr'ose then u~ptake wars in~hibitedi. This is a
reasonablL assu~,ption s inlce no h,droly iis was i ound i n the cjre ofi
lactose obere asr ar iFLcrease in sacroseS hldrol'.Cis in the pI.E:CnCe of
\ar;ous inhibtlor5 tas commonJrlJ found,
50croji u~ptake Iva; not irnhlblted br m?Iib;ose. rehjrose or cm 1-
When rnaltose wasj suppliied to the ti'sue- there hlas con~siderablt
h~drolysis. As sho.rn in Tible 7 the glucose iconcentrationr incireased
fromi the beginning of the uptake. periodj to the end, w~hilr the co~nccntrs-
ricn of maltorEe declined. The daount of uptake~ is explressed ,-, t,.rn,
of umnOles of 91ucome. The con~centration of oluose i..nsufficient to
account for the uptake (Flgure b) indicating t~hat som~e maltose \rjs taken
up ri t hou t hi droly s ri This experiment jhows firstl*," [h.7 th" discjcha-
ride is being hydrol,red and second, that iGilowr~ina hrrolsis the
roonortc~hande0 cn to detiectej in th~e e.-:rernal so~lut~ion. It miight be
pointed out that maltose would be an expected product of the rreakdo:.-n
of starch in the endojperm.
The app~aranic of gl ease in solution coming from both maltons jnd
sucrose maks i t di fficul t to dcetemine th~e effect of maltosr on jiucrose
:he r. urajnOSe was added to the slices na glucGse appeared in solu-
tiorn. Whin turanose and ucrrose toglether w-erii added to the slices thec
Concentration Change in Change in Uptake
of added glucose mal1tose umoles
mal1tose umoless/flask umoles/flask qlucose/hr
0.005M 6 to 24 39 to 23 14
0.011 10 to 40 80 to 52 26
0.05M 22 to 78 346 to 274 88
Slices (1.0 g fr wt) were incubated in water for 1 hr, rinsed
twice, and then placed in the various concentrations of maltose, 15
min after adding the maltose the first 2 ml sample was teken. The
second sample was taken Ihr after the first, Glucose was measured by
using glucostat, Reducing sugars were run on the same solutions and
maltose determined by difference. Reducing sugar standard curves were
obtained for bothi sugars. The results are averages of 3 determinations
on 2 days' slices.
alu..oi 1 a redings ncars littll or nu higher [han uhien 5ruccose ras .dd~dc
alne uran~or = jt j.0511 ijurd jppro ;ijlcatel, a jU01 irhibi t on of;
ucroe ut..,.e(Tale ). he uptale of Eurannse alone was rhecked t.,
using t~E reduicin? sugar rnalr[ed and was found not to~ be :uken up LI., [he
slices.. That the- iel'ect of tbraraoit ;Is not a.ji ur;~ntic iffacr wras derlr-~in
strat-1 t., nleasurinig the ulptakE Oi suifo58 jl00, rucrose In thA pre.-
ence' of ruranost jnd ucrosei in rni piresence of Irlann; tol,
S*:.eral *::per;brants wrei disigrned to ihot-. the effect of tissur
sllcroser le.*l onr the rate- of jucrose~ upt.j:sk*. The c...periinr~nts jre
sual-i~arizied in Tabic 4. Generall, t.er~e is jn ice~irse: correljtionn be-
rr,.*een thE a~llroviL OI SaICrOfS in rre ririui and the- lc'tE of uprE~.ake Thel
tissue ..as subjected ro:~r~l.. rrious trec,ar. -Lquue~Cr- ce SO jn \ntr,' the
amo~unt of ruirose in theF [iSsuei prior to brjeauringl uptake;. I r.hosi
thra.. case~ narkid 6, ar. azterisk the equncrres were th: sjrne C>r.iept
fo-r the cinoncentrato oa' rugjr during !he firit I hr and in t:=.:h casei
;ucriJse uprake u3r IreaSUred during the fif h hr f rarri, th.- ime the irper; -
me1nt rarted. In thise ijses there ;is jn true-llent ;!ir.*;r*. correlation
t..-twEen [h= aBolount ofi Lissue~ SUCcroe C.'tored jnd rthe rjri ofi uptal.e.
Th~ir is tor t~e e-~pected since- thcre umust be~ somer limr~i to the: jlr~ount ofi
sucrose that cjr. t~e accuuilatedi b; the, slices and one would c.>.pect thi
rare- of uprake to to r-educed js thij liln; t is Tejihid.
11h1n yas5 t)change exprCTmnts[ areT carried our usi-ng 3.00 nap of
tiiSFu ir, the \~U~atJur FjLarSk the aliounL Of O, rjiten up tt the clssue
was limritd b, thr rate jt which 1? difluses into3 uner. Thsi h l
by thr: fo~llo\sing: (a) 7).@ rat-s or' 02 co~nsuti~ption noted in thiese experi-
mentrs we~ri cqurl rO the 11rinits of the rate at sshich 32 di fuser into
Uptake umoles/q br
0.01M1 sucrose 0.01M sucrose
Experi ment 0.01Ml sucrose 0.05M turanose 0. 05M manni tol
1 15.2 10.1
2 15.5 11.7
3 16.1 11.4 15.6
17.1 14.6 16.6
Slices (1.0 g fr wt) were incubated for I br in water, rinsed
twice, and the indicated solutions were added. The first sample was
taken 15 min after addition of the uptake solution. Uptake was
The Effects of Turanose and Mannitol on Sucrose Uptake
Trretnen: sequence umOlFIS.g umoleS'J hr
\lAter 3 Fr, rinse, waerIT br,
ricscoeIhr 50 25::
\ljrcr I r, rini:0, sucrose I hr 68g 17
0.020i Iru~ct:Le 7 hr, rlnje, water~
I br rir.Ee. S crose I r 81 Il
0.III fract;se 3hl. rinse, ueter
3 hr, rinSC, suciJte I hr 914 i
0,lIn fructoje ; he, rinrce, raterT
Ihr. rin:e. scrose I he 125 15;
0.111; fructose : hr. rinse ,
cuirjse I br 133 11
The results. from 3 cropsC of seedling s re riported hire. In each
case the uptakle period ejas I bir, the sucrose fronll r-st..c*, up:3l-e vas
nriasuiied ~as 0.0!11 and~ there was a 15 miir, delal L.etueen ocdainq hth upF-
tjLe scilution ond! taking ther first sanple. Dupl Icat rs amp les ,;re
5ubjiated to the requen~ce as listed and at the beginn~ing of the uiptake
Period onez group rjs k~illed for sucros~e anal,sis rrhile rne other .*.ai
usedto- rmeas~ure c_gar uptale.r
Sucro;e Ticsue leverl ilrnd Sucrose Ulptake
water (74). (b) W~hen DNP was added under these conditions the amount
of CO2 evolution was increased; however, the amount of 02 consumption
was not. (c) When experiments were run using 100 mg of tissue in water
the 02 consumption increased on a per weight basis and the RQ decreased.
The figures from two typical experiments were: wi th 300 mg tissue, 33
umoles 02/g br, RQ 1.3; with 100 mg tissue, 53 umoles 02/g hr, RQ 0.8.
Since when using 100 mg of tissue, the rate of 02 consumption is well
below the limit and since the RQ is quite low it was assumed that 02
was not limiting under these conditions.
It was not known whether or not 02 consumption was limited under
conditions which prevailed in the water bathi. An experiment in which
sucrose uptake from 0.005M sucrose was measured as a function of the
surface area of the liquid per weight of tissue showed that uptakte was
not l limited by the amount of surf ace area. In this experiment uptake
by 0.2, 0.4, and 1.0 g of tissue was measured. The volume to tissue
ratio was kept the same by starting with 2, 4, and 10 ml of sucrose
solution, respectively, and removing relatively the same amount of
sample for sucrose analysis. The rates of uptake were 11.9, 11.4, and
11..3 .arreiles/9C br t, ?.2, I?.1 4, an 1.0 J oF t;ue. Theic. differences
.uirrj; upt.3|e ase r.or I1I~r..rd L. 0 i b li .
where I fl3Cd, a Luj Seid ..nd 13:i umoles/31 alhlre 4 rwer: used~. !Th so~
t'igures 3re not ns! sucrocse .iccumulated jince thi amountnt Of: tijsu8
suCroSe at Ih: beginning o r ne upraki- period nas not been iiubtracred.)
\Jork byi Hus..phreys arid florrird (701) jnj by thi author hj.e shon,,
that rhe sulcrore content of rcu~telluslr. SliCes is Treducd rher,~ jliiCe
are inrubated in war-r ;ndicating? that rEugar is the jiubscrjite for rre-
pi ratio:n. his; waJ jlso rioted ..hen 0, 25 g of t i 3;ui per f l iSk: ras u~ed.
Hlowvesr, gas eYchdnge StudieS using 100 mg ofi Lis:u+ h e e~l idcjted jr.
P..) ir. water subjstntiall, less than I.0 indicatinq a subjtr. to other
than sugar. This 5 nconj s t ency weiight be a~r la I ned L,< the Con.irsion
of ilcrosee to olrganic 3i;ds.
Figure 12 shows: the anoun[ of Fernientation in \Iater snd in twoo
conccntrrtions oi iucrojsl when j.00 myg O ri.isue u15 used. It i, acjsumed
in present;ng thes fiiures rIha, for tachi uno's af CO2 e .ol\ed in ex~cs~s
of' 0) take..I up orle-h.11f uiinole) of Choxas .-sa; tbeir fe.-,i:nted, Figure 178
repiesents tht 33rs.* e~Perinent laut rhe aiouint of Forilentaririo in the
rwater control haJs been sbtcracted fromn that caus:d b, surose,
fnh the b-si2 Of 0: ptake iexperiments it r -,as clculated that thr
solurions in rheie rI asks w would be dEpleted of surcrose after j b~r in
the~ case of 0.001J:I an~d 4 ;rr ;n the: casei of 0).)035r slucroii.. In the~ case
of 0.0010r >Icro~ r th suga3r-csu jed feri-antation tr .e rh ar
is j dded, incr;ehie in rare, A~id then decrease1 at Frnc (10.9r then the
Jugrl" should all Lakes been tj!.en uo, T.;i sinountr of iiearment t ica caused
b,' 0.0051 juirore. ij about thrE e i~ t r'?j (MC cus:J br ;j.001ft iucrose
andJ as rhow.n eairlierT in Fi3url: 4 the~ raCL of upta're ;s about tlTree
time as rea.' WIh 0;.000.1 juirote thep agreemenr ;n timing bc ..:een
uptakec and ferl-enit.&tn i; not aj c lose, ho..-eST .,lrco care do-.5 begln
ro dzcline at a time~ where1 uptakCe ihouldJ. be coFplem.
Figure 12. Fermentation in later and Sucrose. Each flask contained
300 mg of tissue. The sucrose was placed in the side arm
and added at 30 min to give the concentration indicated on
the gr-aph. Three sets of flasks wrere used, one each for
the 2 concentrations of sucrose and one containing water.
The values were obtained from; CO2 -02/2 (in umoles), in
B the values found with water have been subtracted from the
values found wi th sucrose.
3 urs jOdddedrs~or
Sucrese odr'd edC
1 0O -q-q-twM-~
a r~S~:;% ?C~0. 00111 SuCroS C
.d2 e-I Ir C
Figure 13 shows the gas exchange in 0.1M sucrose as compared to
that in water. These experiments were run under conditions in which 02
was not limiting. With water the RQ is less than 1.0 and both 02 uptake
and CO2 evolution proceed at constant rates throughout the experiment.
When sucrose is added there is some depression in the rate of 02 con-
sumption. The rate of evolution of CO2 on the other hand continues to
increase throughout the experiment. Over the period of this experiment
the uptake of: sucrose would proceed at a constant r-ate (Figures 2 and
3). When 0.01M sucrose was added in the same type of experiment (data
not presented) the patter-n of gas exchange was the same; howJever: the
02 consumptioni was not depressed as much and the rate of CO2 evolution
was not as high.
Figure is shr.:si the~ sjare ie pa f U~Ita fo'r 0.1[1 glu~cos. r.ar
resolution of~ C0-. ; grtsll, nrcreased t., [Ihe adds F.(an 0i 1Urj glc li to
sincei. Ibe. F.I during Ith l ear. pr ..d mat-urcd ..jr z7; 115 .,,,.0?. acr..
0. lr.01/1 glu cs !data nres ireset ed)j caused ro:uJh.1, I'te rsame p L~ir r, -,
If Ith .assUmIPLtio is need tha thel d~;liffrtence ir. C02 t\olution ir.
vetrj E It r..u5 thol in sugar ii lui Er.11-I t r e tai;(.S' r r t
is ol~ib~lC to CalCUlateC Ihe reek.-:*r ofi umle!'~C i hCrSid tbeir.9 Fer.T:nited.
Figuret IF shar~~ ~-e thC.- J res lt of uh a icalcula tin. Th sr.lt. r
frir, thC sar~e e~p rir-nts .a Itpo~rted ;rn rigure: 1.. and IL*,
Rate: of~ IIIrrilireatio~ ~U suggested bt [lithe .-.t. d31 t.- ar-conl..ustrar
As ioir-tedJ out crl~ie, 1:F.6 rjte of ftri.,ntation~ ;r higher under icndi-
.p iO. -" arose
1 O, ae
<, 0 ,. i cr'
Sl ,-o~ O,,, water
C.. ..... .... -L 1.. IJ _1..1. ..L_ .
30 60 90 I20 I50 180 210
Ti rne~, ai n
\:ere run rri lhi 100 cl] L 5ue per i last.
olC'uri.Ior (LOM() was- ajdde f ruin thE Side
of ter re-dilngsr were tregun~.
jrmr 30 rainr
0 C2, glucose 1
O 02, gl ucose
y 02, we te r
-- .. . L e .
Tij no r
Figu e 1 L 1.ch ~ g. iri'a e a,< 0 11C' ...e cn iE o e
tr e It : i .. F a u e i
C, .0i r1 gluccose
ri* 0. I t avc ros5e
L L --. ...
i0 60 90 120 1',0 1 0 210
Figure IS. FcrTer~!a tior, in 5u3ar .elation. odio. n
Figures 13 and IN, F'e ull: \*ere ;alcu;lct d s LO,
accumulated is lower under conditions of limited 02 in the water bath.
The rate of glucose uptake from 0.1M glucose was about 87 umnoles/g hr.
The rate of fermentation as indicated in Figure, 15 was 28 umoles hexose/
g hr. This would leave about 60 umoles of hexose available for accumu-
lation, or enough to accumulate sucrose at a rate of 30 umoles/g br.
An experiment was run in which slices were incubated in 0.IM glucose
for 3 hr. Each flask contained 250 mg of slices in 2.5 ml glucose
solution. Sucrose accumulated at a rate of 35 umoles sucrose/g br.
Experiments were designed to determine the amount of uranyl ion
that would bind to the slices, the effect of other cations on sugar
uptake, and the relationship between the binding of some other cations
and the binding of uraniyl ion.
Table 10 shows the results of two experiments on the effect of
cations on sucrose uptake. In the first experiment the cations were
added to the uptake solutions. In the second experiment the slices
were first treated wi th 0.01N HC! to remove cations attached to the
slices, then treated with metal cations, rinsed, and placed in su-rose.
Uranyl ion reduces sucros~e uptake when it is used as a pretreat-
ment or when it is present in the uptake solution. Cations other than
uranyl ion have little effect on uptake. Notice that the acre~rol,
third column of Table 10, was treated with acid showing tnue rth acidl
treatment does niot ser-iously impair uptake.
The next series of experiments were designed to determine: _Th? quani-
ti ties of the various ions that would bind to the surface- of .nec laces.
Figure 16 shows the amounts of the various Ions that cjn to reno.ed~
..im rhr.01t ~i HI after p~re real~nirr witrh the various metalIs. ea
Upte..ce in urooles 'q br
Treatment Ul; th upt~l:e Siolution Pr~ e r lle et L
Suc-rrose Ilpraic as k.ifcceed b,' Variojus, (io~ns
UO2 3 ?
The d)(a~ ill trd riCond Co~lumn sf6 ffrnm ,n experilifnt ir nhi ch
slices were incu!,..ted ;rn warter for I br, rinsed. llith \tearC.r adTI: pl.=sCii
inr 30o1lclons containling 0.005M sucrose anj ii.i0311 cation solution.
Th~e t h Ird iCo lilr., repFrese n 1 a 1 5 -min i ncuta tion in 0].011 H C.I fo lkI..,e.1
by 2 rinses, 50 min in 0.003tt :ar.;or, solution, another rinre jnd i,-
nally the (>.00*,11 suirosE sojlution from r-th~ih uptak~e wass i~eneasred. In
btlUh e~.*periai~nlnt :he uptal:- period wa~s 2' hr b. ig 0 f slices. per flail:
--.I--I-------------- ----- ---- --I----------~-----l"r
0 M 2
(7 ] 022
W Ca 2+
Salt solution, N x 103
Figure 16. Metal Finding. The schedule for these e-peria-ncnr
involvedl the following: 1 br in the car~~ .: llr. olud
(or water control), 3 rinses, I hr in !*,<, 3 r~orar
r:i-. ---.' I.r analysiss of th~e respective c nores, tail
i1.0 c..com...ed 1 9 of slices.
points .;an be made. ,iTein..r on ftiecyr eit
searij' rinces and a I-br .r~cubation in eY->rer. (b) A- concentrjttion of
0.0020 .:ation rolution r,:ided enoujh ions5 Eo saturjte the rltes ;rn
all Cases. \c) ApPrCTiiabe quantities o~f the ;oni at 10.-, concentrJ--
rtoi rcr- ur boundl to the slice. In the case ofi ;Jbal: (0.000)11), 1.6
un~clers or the 3 umeles ;I.ji lble wrere bo~und b: rne jlices. (d) fljgne-
jium jppersr to be th-: ;on rhat, normally occupie; the sites. The data
from Fiqure e 1 hou thlE the slici-; incubjtedj in \-d~ear insread oi
mea~l iarion solurionj released3 6.5 umolej ofl 1122'+ aInd 0.?S uni~ls of
Cs- i. (I) G;-earer quait~itiesj of uran,*I ;c7, jrE boul~nd the. ,an OF the
O~lher ions [GE L.d evcept[ r1ig'
In theI caset oi I~renyl ion, twro T.-pZrimenri~ :,rTT de-~;i. d to see- i
rh: cffect in sugjr uotrake paralle~l !he deglre- of catio~n bin~in3 to
the ti res. i re i h a- h re u o h s p ri nt .nd t
can bt: seen that maximum ilhibtr ion i; reached jC concentratacni of
urinel ion tha~t jecrur.te the binding ;irej. TI-e inhibirlor. of glucoC'e
uptcake in this expcrinent was.s greater th~an ruju' !c'.g.s ce Flgu e 9).
IF the- slI;er we~rc first treated --with lICI uni thien incubear~il in
melitpl iolu~~Tions, thF jmouLn*, oir coacTn bihJndin \1as Consideraldl riduced.l.
This is shot-*n in Fi lur- e !2 fo." ursn,*, ;on, 11nl and (C'f. a.1n i
[he jcid treatmcnl I'endered i.oLE of !h's sitri und.alajlblz ror C3Eion
bind ing. M~e..avr, thi EliC; L oo'* up sucroje as ;hanr in Tabls 13 .i Ih
or rithojur replacing th; catinsh. Of TCoorCe the por::bilit C.:.its
thjt the~ sites in.crl ed if. sugarI 'JJ~rj e werre fear-TCapied~ b.' TJa;r; on rom~ri
.cin he 51maliZr I~auanic i. n Urlnyl( "on [I at bind5 after acid
[Ltrel:tmnt vas surficien[ 10 irhibit uitake s jS s sho.an ir. Took 10.
Wlork ,r;th~ 'ICast In..icvtsLda* ll et nll le i ir displaced f..On the binri-
20 .~ 9 Glucose
S2 3 4
Pretreatment uranyl ion concentration, M x 103
Figure 17. Effect of Urany) lon Pretreatment Concentration
on Sugar Uptake. In the two experiments reported
here the schedules were as follows: 1hr in the
designated concentration of uranyl nitrate, 2
rinses, 30 min in later, 2 more riioses, thE- ..pcIIe
solution. The uptakte solution was 10 ml or Ul.0111
sugar in both cases. With sucrose the ayrj;. pi'ned
was 90 min; with glucose the uptake period j; I br.
(Whereas the uptake period mnay affect the Jag~ree Ji
inhibition, in this case the experiment wa; ~sl i 5cja
to emphasize inhibition as a function of J on,I .00
binding and not the amount of inhibition.
I~ j r- f -n
U j ] ji ~' '
.1 so u io 111 1
ure I.T fla Bi~ ig r le na cd P
'ue:Le rr Tre sl cs'. r st
.:rc ?uj ce o th o lwn
!,r u.: nce s mn i.002 ~ ,I
ir ih 7 .5 l to. ".e o
to- PI h 2 a e r n r is ..0 0
HO re I3 slu,2 '-e c-. eoe
..or ar:1 ,* i .1 c e c t o
ing sites during sugar uptake and are again bound after the sugar has
been taken up (53). Sever-a; experiments which involved pre treatment
with Co2+ or M92+ fai led to detect this phenomena ri th slices of the
As described in the methods section, corn was normally grown in
tap wa ter, In one experiment corn was grown in distilled water, Su-
crose uptake by slices from this group of seedlings was about the same
as with slices from tap-water-grown seedlings (i.e.,17 umoles/g hr
from 0.01M su~crose). Prtetetwt at fC2+, Mg2+, or KC had
little effect on sucrose uptake into slices from seedlings grown in
From thie ri-rults of this 'tudy the follorinj conclusion jlre
drjwn. (5) SucroseC ;S telken up jcti\ ely~ without in\erdion. (b) Hezasej
3rt tak~en u~P byi tlvo prTO'LCe.Ss operating S;imulter.eo-iUtI,, dillu5;5r 5:rd
3Ct;ic Ironsport. (c) The acLi~ LCu [31 .511( 1801810$ fn $UjrorLe 8"r( 10
hc.srs ari locjted at the plj51male~i m~ma (d) The actise uc;:.i:L mechaj-
r~i ms for b~oth jucross and the hr:..G.5rcEre dlriken toi .0,iol,siia. ()
The charjiracr i st ic o m rE [al b~ndinj 35 relate [d to sugar uptle areCd
quLi re d~i ffrent in thei corln jcur-l lumT .-then ~cmji lre 1 EO (ho~re i*1 ,cjs t
Several line- of evidence hose- IJllCdHunhreis >..3 Garrard (115) to
thr -::niclus~ion tIat jaCrCote iC. taknC~ up vithoUt i I~crsior. b,. th3 Corn
9CUrtlyl. r;SU: 1' cut btained ir. this c-Jork. sul'po t that co~nclusion.
(j) The amount ofi ..xtracellular inversion <*: insufiident~ t, sLpport
u~ptak' it ;he ob C; -ed rate. (b) The tircat~ics of hC:.r~os uptakei d.e
different fr-om the~ ~inedeis ol sucrose uptake (ci. Figure 4 and Ficiure
8). (c) The rate cr ferarranjtition in sucro;F was less than would reo
expcted (Tiable 1) i f inve~rsioi occurred ci ther pr;or- to or during urp-
talke. (d) if inter;on prcu;lsd uprjke, thie attee.: o uranil ;on on
sucrose should I..G simllar to the rFfect on herose rvbich is nrio the
case (ofi. Tablel 4 .and Figure Y). (e) Tjhr rat;O ofr jcCL~Ul3[- d Su~ClO5C
LOr Suga~Tr ken up it ; g. hir Irhenl Cu;Troe iS iUppliced thaln obetn 3lucuse
is cuPplisd (p. 51).
The mj tor.: draa (Table 7) showJ thj; malic.Le is alsor trJ-i-n upF b,
thF LISSue w~itho1~ut hydrob)li .
That sucrose uptake is an active process is indicated by the fact
that it is taken up against concentration gradients. Slices that con-
tained 133 umoles sucrose/g (0.13M~ sucrose) will take up sucrose from
0.01M solution (Table 9). Slices incubated in water for I hr contained
68 ucoles sucrose/g (0.068M sucrose) and will take up sucrose from
0.001M sucrose (Figuire 2), The tissue sucrose concentrations given
above are minimum values since an equal distribution of sucrose through-
out the tissue water is assumed; compartmentation of sucrose would in-
crease the ratios. Ihe scutellum is composed of mesophyll parenchyma,
an epithelial layer and vascular tissue. It is assumed that most up-
take occurs in the parenchyma cells since these cells appear to make
up about 80-90%~ of the scutellum.
'The inhibition of sucrose uptake by DNP is consistent ,i th the
idea of an active, metabolic energy-requiring process.
The inhibition of sucrose uptake by turanose (and probably by
lactose) fulfills one of the criteria given for a facilitated diffusion
or an active transport process (I).
The data in Table :0 clearly show that a 15-m~In rinse in 0.01N
HCI does not disrupt the normal functioning of the cells insofar as
uptak~e is concerned. Since this treatment removes uranyl ion from the
cells anid restores the ab iity of the cells to take up sugar (67) it
appears that the effect of uranyl ion is at the cell surfjce. I: ii
concluded that the mechanism for sucrose uptake is located ji the plat-
malemulal and the same argument can be applied to the: activI j.-r."al jr:
glucose uptake (see below). :n contrast to :has..c results, ~is:M-r :ik)
found that uraniyl ion had no effect on sugar uptake. Hum h r: ; I pt, o 1l
suCiroe. i n con tra t prE t,-ca rirtet of i 5ic:1 ; a 1.0.4 aucrosei cauSed
the: slices to 3Cppearj il-cc;d and ienderedi IhemT IsocapillleS ~r :ath~esz-
ingJ sue Isse = hen pi ;_ed i n I!. 1. I Fruc re,L TheSe obse at~t io .5:~ :'uPF:rt
the~ col~tntentio [l,<. the~ .warm~ral R.'mbrjrnc is pFcret~eur. c cc herose. t..a
not [.' ruirose.
ilso0 has an acri.: i ose' transport miechanistll.. The constur nt rtei IJF
?luaSi uptake until ;'u bathi.ng solution h.15 been .Jeacre.)o- (Fi |13)
;;annor be- izploin.1.1 :. .lirfurion alone and ,cr ,i..an darj had bien
aCcumullated to ShlOu rhat Ihi- Cl!Oprl35m is .'ric SFpice U.j C00.~.~. Thi.
combinati o~ CI' diffusion and .in 3cti.C ICr.isport[ m-...aniser ce., i .plaIinr
rthese resulrs. Th: h:t.-use- uptjke j cancincrrcioron cur-:0s mighte ;'so
Is ar~p~aldiffs cuse.It i; ctruc that this curse co:ul.:1 jlro
h the result ofi i ..r;ocss -h~ic follor-0 enz,.r~e hirleic-: prOviGed that
rE.: I'ni is hign in iconsp~rljo n to the concenl rraton.Hss*er hesb
strteL c..ncnttrrration ia,a O.0111 glucoser which -souldJ require quit.-. 3 Yigh
blocking ~.he2 4ctivl porT[; on of gluoseJ Irptake. iht upat.E rtte s Con-
ce~n;..Mrir *:-.cj 'Or thd becoses~ (Figh~ F) jlnd ') do no7. t re-emble
paic-ISS r:-.xpr JFter LT.tret--irt ri te vlrar.,I ;inn in which :;:1 -s losr J
g Ilucc:O ;caus, Irlcai.~L :n I :3. dif ;'? i n 5e al.- re plr f5Cnt rf 0. .3
form sucrose or is catabolized. It is assumed that the glucose which
diffuses into the tissue is phosphorylated via hexokinase whi le that
glucose taken up actively is phosphorylated at the plasmalemma. This
is active transport in the sense that uptake is being energetically
driven at thle membrane but it is not active transport in the sense that
glucose is being accumulated against a gradient. The active process
described h~ere would be called group translocation by Roseman (57).
A combination of active transport and facilitated diffusion is
thought to be i involved i n the uptake of glucose by yeast (55). Rei nhold
and Ellam (2.6), working with sunflower hypocotyl, suggested that active
transport operated in the a'osence of DNP but that in i ts presence
sugars diffused into the cells.
The idea that all of sucrose uptake is active whereas half of
glucose uptake is active and half passive is supported by the following
results. (a) At concentrations of 0.01M sugar and 1 x 10~3M phloridzin
the inhibition of sucrose uptake was twice that of glucose uptake
(Table 5), (b) Sucrose uptake wlas inhibited twice as much by DNP as
was glucose uptake. (c) The effect of anerobic conditions (p. 'fl was
10~ irnhib~l ea'ur.:.' e parate [,.ICr i. iiuch "us ?luccee uptall._- h:.-Tae,
_uc.s, inh Ibi io jr; ri ecrose u.rsI; a rt. co l t .
Th: lac: tr t icrmorstjltion ie d-sected c.;i~r. hen 0, ii not lir..iring is
ar. ind ; c'io i on !h-at bL.jn lr[ rati o .Jri.,i; uptke iregrdicr s of' :h~e a jll-
o~F CO-. !o 5 ji!Luration of the le3;pirjtor*, C71 risT. sno h
case rirh J1:te~llum~ 5lic5 inci "hie cojnsumnption ,f On ra; relducId ;n
rlhB pr7eseceii o1: 0.It Sugar.
E.ide~nc;- hjs ben presented to show* thjr ierimenl.Jion dr;t.< suear
upjltae, t: c.-jinr~e hjs beein et~tjirn:J to 3hoes thJt i iPecific. 9l;roI l; i.
Cst'F is responsble, howet~er, !he data ar cjnsist~entr ..ri II- j,: r,.n
5uch as th: phof phot -jnorjes e Ie 5,seiT i n b3,reria w he-:re TE i s thr
enerjv sOurCe for up two Ga3rrard ;rnd Huimphrc,*s (.LS) U~.u-i,ingl iontrol
of gl*,colysse in the minjzi liurcllum foundl no diffriirnces ,n th? r\TP
leittr1 if the pr'eSenic jrid jbienii of fructOse, TIhe ~ir.ount of frucrore-
Pwih:riianslates prsphoffrTuar.lki nrte doubled i n the~ jlresicac of
fruiroie bJt [h.5 rrt; cOnsjiderd iniadequale to, aiccount fiir a ior-fc IJ
in~crea-,2 in th-e rjte of .,liCol,-sis The usr of PEP .0? the suar/~ uprC!;
proces ar ~;ii h tr i gge.r t rm n tajt ion.
Th: .fate ;n Figure & whiic h)\ shu TucO:G upE~he j+ 9 function jF
the concentrrat!Goi in chu bathing silutiorn ilojc!, fit flhes d
;enren cuirse. Tis j .pe of dataJ i of ten pr..:-r crt i,. su!;orr of j
iur-.t.: th:., rhC, J lInes* upiakle ri; h '.ice in .5 bjth of dlecrea~sing icar.-
centrj lion (Fip. 1, i, and 3) are nioc j ..II r,p~ical of rnz,m.. !:;..ic>.
Peg.3rdians of the n~cchainn~ir~a o uptjl:e, La it dififusion or .3 carricr-
ac.0 sted 3c :I.C p'rocu, the rate would~ 'Je pa ~tted rr ~...crejse js the
jug.- cojnirntri atn in th': ,jluti n waj rCduC' 2.5 3 result of upsti-c.
Ait a ;.2n c.)nce~r[ Trade. the ;on4tanL Uptak.e rTeI r.]ht be 61-~
plainrd. js 2. Saturi tion ,f th :Iptae sic..hdn;ism for sucrevez Zut this
tIr`c'lborlio is japparnT~LI na.! Cjaturad unti[l .1a5n--antration o~f aboar
0,4M (64) which does not explain the constant rate at 0.005M (Fi;. :1.
It appears that whereas the substr'ate is not the limiting factor
(Fig. 2) the substrate concentration has something to do wi th the rate
of uptake (Fig. 14). When the sugar concentration is maintained at a
constant level sucrose uptake increases with time~ whereas glucose up-
take does not (c-f. Figs. 5 and 11), A possible explanation for these
phenomena follow. The rate of sucrose uptake is governed by at least
two factors. the external concentration of sucrose and the internal con-
centration of the phosphate donor. When sucrose uptake begins, the
w~hir bi si-Z.:-. .s i.-.ci-:::ir ei rare orf r02 0 .it, H e .c e
Lei r..3 coure.l:; :J t, .r. I;~r..3:re ir ;n[r-t r..1 .isj .. Icl? :s aIC I:;rn`; 1 su
;U..rC;c upro;l e.
L-r.relir s .i., ~.i.por jr .1:nt d jlerine: 1-.e rate: oF CrJ:tr r:p r i r
Th~us :s: the C;co--st.itio dec~linE; inS Sci e proie~s. .: I~j j ijlal.. ..
car..:-c.rr aro ica of L.Y.:.:Phale~ doreT .r : rer~i~l UKl prC e::.; Tne r jIi
re~ul[ ; isi I.0 up..rla- CCate rJij i; almostJ 3 r.ornstinll uniil Ilh' Bla~ocs In
thec 3nmoun: Of e~xt:rnest gliro.,r ; i ConStant t murofd ui
;.COnSrant andl thusl rhe O.srall r.ite GIi uptal.. Irenlj;.5 cons .-.nt.
.4; comisinlar tion of sct;is are) paSrs: e jlutoi C~~~.: uptake netchani e. (ir
conc: ntrati[on .*,jljh bic rto i.he i ulllu r: lj 10, the 5rst..m is capable
oi' rL1soIn.)~l~ all of cihe j.J;I-ble glu.cose~ quickly; h3~e..cr w~hcn thi
--ndu',pern l IjllCUm: Is blig'1 the 'cultellun cji.. (;1;C up !lucorei ur. exceI
of the cjp~ci, of th:r ;J=rtl, procrss.
.i; juga- ul tal e~ i5-st~rldr. eri :.,- giJ,col, .;s .lgso fIts Che 1ole 3.
the Lo r~ ur.. Ther x,.lli, to d~i. .- uprjl.*= undEri I~jnited i, 5uPpl .
:lould be of ob..iau. ;.1.tintag toj j s CJ under 3oil canii~r.Jin. sr.r
and- riump;irc rj I.; ')as me..asun-d aj iQ of i wi ch ..holel .calleIcle ;n Air
indij cacingr t1\3 LhE sCu C. 11ucr.~ itseli meight ;imp.'i e conder5n on S o Ilsi r red
I' is ot;iouis 1..1t [hM mu l;l t..iiders. Chara4 cLer St.(. ofl Ir:utL ur..
sli :-sJifer rer, o e of ea=v. TheC tindinj s uran,*l I sn to ;er st
cills -.esonsr to e :u: r. spc~ir'ic to the uprjlid sit. r.53'. ;rt h sc i-
rillUil ;ICi.. noC li.. .,ct of' Ih' Ma~n.' urs.. ion is Csu if lVOL.~1 .3
thei sp[Ske i.r~re~SS aS sh7'~ran 'i! 'he sectucedl bur rill effecc:; .. as.,Irul
J:u ndJ .jfter acidi EreC reent-I (ci. F17r. 16 anld I I, sold T~t.Il: lIC). I
chdl ..randl ion ..a. t~ilun :I L, onr root~ ..issue, -t ....n i o b r s
thei m-~.mbraini surf ac-= If' the 'e~lTaJ F Oi bound icanOn; OiCcurs dancyi
segor uptak.e to sCU[Cll,' n Slice. th-1 as-sun =. 1- Eco,3 Cmil II~ 0. L aleC[':
rith th*- mci;J1 d;S53. pl'O.CJ1ri* uis ...
1. Ll. D. Stein, The Monvement _lof Molecue Across Cell Membaranes
Academic Press, New York (1967).
2. R. K. Crane, in Comprehepsive Biochemist~ry, ed. by Ri. D. Stotz.
Elsevier Publishiniy Company, Amsterdam, Vol. 17 (1969).
3. A. L. Kursanov, in Advances in Botanical Research ed, by R. D.
Preston, Academic Press, NewJ York, Vol. I (15163).
4. C. E. Hartt, HI. P. Kortschak, A. J. Forbes and G. 0. Burr, Plant
Phvsiol., 33, 305 (1963).
5. R. L. Bleleski, Plant ?111121., 41 Li47 (1966).
6. P. c:, \feathe~rley, New Phetol.i, 52, 76 (!953).
7. Libi., 53, 20:1 (locL}
8. Ibid.. Sk, 13 (19551.
12. lbid. 2.0, 565 (19< 1
13. P.' J, Hardy and C. .!:.rtc~r 'I i. ? .:.1. .. 1 I .6
17, P. Krie~demann, Pl r _:. . I
I G r r h . r n i .I
L, F.-int..:.id and 1. E-ht..r, Phon Ph,001,, 0,, 1021 (1). ).
J. L. H:3rh, 11.3 11. 11. lennar..ji, Pioc, P.o. SOC., (,nl. 0, 11,
' 05 I I.-iki,3.
D. P. 't.,rgan ina H. E. Stracr, rnn. Col. fl. .. 23, .9 151.1.
G. 1.. Its....nas and fl. P. \* ir, th'.. Ph tol., 6(. 125 1196 1.
A. E. rou*.;s fl. E. 'r-:.ur.is, and ll. 4. G=-cr. Pla..t C. 11 sird.
10, '.75 (1969).
L. Econr.-alry ar..1 .. Liam, J E-c. B.:,l... IC 29" (1990).
G. t.. P.;.Elesti, Auir. ). Biol. S.:s.. 11, 10?. (19.30).
bid., II, 221 (19001.
Ited., I'". 429 (196 1.1
V. T. Glas.viou, Plant c... ;,al., LE, 895 (1)(0).
It.id., LA, ],tb (1101),
n, 0 tare:I., J. A. Sacher, and I. T. GI..s.:.ou. Pl r.t FI.-ser.,
n. F.I. Hasrb e.1J (.. T C.12nior... PI-.,, lb Col., 3.:. 1' *1963)
J. .A. Sadler, ti Co Hatch and b'. T. Glastacu, Hart Ph, it..I..
MS (1915 ).
fl. G. Flar.h. 6Ioch.-n. J., at C.'I 11960).
J. 5. 11 .,1.. r and :1 0. Hatch, inchem .I.. 99. 101 I1966).
;. 11cad =:ino, I. Beol. Cher... 1.5. ?J. ?"9601.
.;. J. Ms-.1.-r vid II. G. Hatch. th.5 ..*1. II .1,, 10 4' s il96=,).
11. 1. Hare!. -.0 F. T. Glas ion, PI.r... Ph. ol., 19. 180 il9.50.r.
J. A. 5.1 '.er, *:fe Cg;,-,.,-, to 194
2.. H=q aC J. 2. .has i-, F ar.c F.", .il., O 591 (1905'.
it. I IC cohis arid J Ed. non. .'. Ey B-:.E. 2I, E. (197").
. 2. Pa od, Alar.. Fe., 11.,0 Ph ?.el., IE, 15:. 106.1.
F,.*,al. .ard U. ... 1-,s 0, ,j, ra, ,,.. 20', 585 (13 4).
45. C. E. Cardini, L. F. Leloir, aUnd J. Chiclboga, J. Biol, Chem.,
214, 149 (1955).
46, L. F. Leloir and C. E. Cardini, J. Biol Chem., 214, 157 (1955).
47, J. S. Hawker, Biochem. J., 105, 943 (1967).
48. C, P. P. Ricardo and T. ApRees, Phytoc'inem., 9, 239 (1970).
49. A. L. Kurssnov, 5. M. Sok~olova and M. V. Turkina, J. Esp Bot.
21, 30 (1969).
50. A, Rothstein, fi~eRL295&xP 15Biol., 8, 165 (1954).
51. A. Rothstein and R. C. Meler, J. Cell, Comp. Physiol,, 38, 245
52, A. Rothstein and A. D. Hayes, Arch. Biochem. Biophys., 63, 87
53. J. Vansteveninck and H. L. Booij J. Gen. Phvsiol., 48, 43 (1964).
54, J. VanSteveninck and A, Rothstein, J. Cen. Physiol., 49, 235
Si. A. Rothstein and, J. VanSteveninck, Ann. N. Y. Acad. Sci,, 137,
56. H. Wheeler and P. Hanchey, Science, 1ll, 68 (1971),
57. S, Roscman, J, Gen, JIysio, 54, 138 (1968).
58. W. Hengstenberg, J. B. Egan and M. L. Morse, J.Biol~I. Ce, 2113,
59, J. Edelimn, L. I. Shibko and A. J. Keys, J._ Expg Bot., 10, 178
60, T. E. H-umphreys and L. A. Garrard, Phggac-hem., 3, 6ir7 (1964).
61. L. A\. Gar-rard a~nd T. E. Humphreys, Nhatulre, 207, 1095 (119--?)
62. T. E. Hlumphreys and L. A. Garrard, Phytocjaem,, 5, 653 (19*4)!.
63. IbIdJ., 6, 1085; (1967).
Gk bid., 7, 701 (1968).
65. L. A. Garrard and T. E. Humphreys, Phytochem., 7, 1949 ~~i I.
G6. T. E. Humlphreys and L. A. Garrard, Phyt~chemp., 8, 1055 Il?69)i.
67. slbid., 9, 1715 '1l? 0).
70,. 1. C. Memph r, s ,,nd L. Ai. C.irrord, Q gto--hem.'~ , in press (IS'l ),
71. t, la s n o U =m 1 3 7 14 )
:j, r', i., ".ire, in Crl~prgx Larb~oh~drhrre:, ed, b.' E. F. IJessfeld and~
I!, Ginsbulrg. Aa.ii rs ,ue se k(9'6 ,p
/4, 1.. U,. Urn'carcit, r.. Hl. PRurris and J. F. Erjuffer, IV~norneeric Tch-
r l~~, Ru~rge. s, 11i one urspe lls i!1?, 97, p. 28,
17, 110 I19 5)
79. C, H.F.C n r ad LC h r igo ,An g 1 i I .
79, E ,kk.,r l rm t i e er i.to fT a e 1c a i 30.
81. i, I Joihl, And.U Chem c ts, 3..-st 140 f (1943).jn~-j.;l
Joseph Henry Whitesell was born August 11, 1936, at Clearwater,
Florida. In June, 1954, he was graduated from Large High School at
Largo, Florida. In June, 1958, ne received the degree of Bachelor
of Science writh a major in Ornamental Horticultulre from Auburn Univer-
sity, Auburn, Alabama. From 1958 until 1960 he served in the Uniited
States Nlavy and was stationied aboard thec USS Cutternut at Long Beach,
Ca lifornia. Following his active duty with~ the Nasvy, he w~orkedi for a
yea"r for the Countiy' of Los Ange-les as a1 marke-t inspector. In 1961
he returned to Florida wh~ere he w~orked as an Assistant County Agent in
Collier County, anld during that time he resided in N~aples, Florida.
In 196j he 2nro~lle in the Graduate School of the University of Florida,
Hea woirkedl as a graduate assistant in the Department of Botany and as
a r~ebeach 5Ssistant at the Pesticid~e Research Labor jor a .l e.
ber, :968, when he received the degren of Mlaster- cai "..:.n- e t ..r~ a1
m~aj or in Botdany. Fromt December, 1968, until the pre star nhna
puirsued his wrork toward the degree of: Occtor of Phi~... -il
Joseph Henry h'h~i teseli is malrried to the former !:.e.n-: 11, ..
Stens-on~ and is the father: of three children.
al..u i.ses n. ...sit .s c ~la di s r ia i r or te de r e 0
Profe i or o~f 150tin,
i c.rti., IhjI I h~a.i rE."d this itudrr and that ;ir rt, 1.pinQon .1
onf?-m r r.: acces EDLI~i srandarrdi Ofi shoi jrl prest~ni ot or, ari is f ul l,
i crCif; .r. L E l hs;.:- r:0CS Lhis stu d-, ur. thjt ir, n, opinion ;t
ior f:.i....r.5 to .:p.-ep tlc r.:ji.J-,d~ rds of h y :. pr snI o id fu
adrl' c quite -in* C e jnj q~ujl :v,, ar .; dlsctl Lijnn For r'.* de rs 01
DoctorL'Z C. f~leoh, -
P ychard C, mirn /h
ssiiit .t n. l fes -r f Bc a
:l. .I atJn.:. ..- Src.*e end .uolit,, as 2. dis oraI on[C for~ !Ie JEg:C-a of
Doctor ui *r'd:0(.h,.
Pc.L~ci H-. 8.ggs / /
Frofe nori l of F-rui~ r rr,
Thiis .J;~cirtaion ars sub~mirred to th~e Dejll of the L.ollege of Agricullalne
AIIJ to thei Gradve-ll: Council, andl was- acctepte ;I partial l ulfillmentr of
the requ~iem~in:i for the degree 01' i'c~ctr of Ph losoh,.
Juni, I 1;
Deac., College of Agr icul ture
ean IGraduatre ;ichoo