TER VERKRIJGING VAN DEN GRAAD VAN DOCTOR
IN DE WIS- EN NATUURKUNDE AAN DE RIJKS-
UNIVERSITEIT TE UTRECHT, OP GEZAG VAN DEN
RECTOR-MAGNIEICUS Dr. W. E. RINGER, HOOG-
LEERAAR IN DE FACULTEIT DER GENEESKUNDE,
VOLGENS BESLUIT VAN DEN SENAAT DER
UNIVERSITEIT TE VERDEDIGEN TEGEN DE BE-
DENKINGEN VAN DE FACULTEIT DER WIS- EN
NATUURKUNDE OP MAANDAG 28 JUNI 1937
DES NAMIDDAGS TE 4 UUR

Het verschijnen van dit proefschrift biedt mij de welkome
gelegenheid, op deze plaats allen te danken, die tot mijn vor-
ming hebben bijgedragen.

Hierbij gedenk ik mijn leermeester, professor Went. Aan hem
zal ik steeds een dankbare herinnering blijven bewaren. U,
mevrouw Went, dank ik voor de grote gastvrijheid welke ik
steeds in Uw huis mocht ondervinden.

U, Hoogleeraren in de faculteit der Wis- en Natuurkunde,
vooral U, Hooggeleerde Jordan, Nierstrasz, Pulle, Rutten en
Westerdijk, dank ik voor wat Gij mij in colleges en practica,
op excursies en in particuliere gesprekken leerde.

Hooggeleerde Koningsberger, Hooggeachte Promotor, U dank
ik voor Uw 'hulp en kritiek bij het bewerken van dit proef-
schrift. Daarnaast dank ik, ook namens mijn vrouw, mevrouw
Koningsberger en U voor de gastvrijheid en steun, die wij van
U beiden mochten ondervinden.

Zeergeleerde Offerijns, aan U dank ik de keuze van dit
studievak, waarin ik zoveel bevrediging vind.

Het personeel van het Botanisch Laboratorium dank ik voor
de aangename samenwerking gedurende de afgelopen jaren; mijn
collega's assistenten tevens voor het vele werk, dat zij van mij
overnamen, teneinde mij in de gelegenheid te stellen dit proef-
schrift te bewerken.

Chapter IV. The Relation between Glucose Content, Res-
piration and Auxin Production of the Root Tip 305

2.nbsp;The Influence of Phosphate Buffer Solu-
tions on the Production of Auxin and on

Boysen Jensen discovered in 1933 that cut root tips put on
agar containing 10% of glucose delivered more auxin than those
put on plain agar. This statement of Boysen Jensen was chosen
as a starting point for the research reported in this paper.

Generally speaking one may say that auxin mostly is present
in young organs and not, or in distinctly smaller quantities, in
adult tissues. In some cases auxin is regularly spread all over
the organ concerned, e.g. in the hypocotyl of Lupinus (Dijkman,
1934) and in the epicotyl of Vicia Faba (Van der Laan, 1934).
In other cases a part of the organ is richer in auxin content

than the other parts; the auxin diffuses from this „auxin-cen-
trumquot; towards the other cells of the young organ. The best
known case of this is the eoleoptile of the Gramineae, where
auxin is continually produced in the tip and is transported from
the tip to the basal regions.

The question may rise, why the auxin content is so much
higher in the cells of the tip of the eoleoptile than in the other
cells. One may explain the lower auxin content in the region
of growth by assuming that the auxin is consumed in the growth
process (Bonner and Thimann, 1935). This, however, does not hold
good for the cells below this region, which have stopped growing.
The fact, that the auxin content of these cells is also lower than
that of cells of the tip, indicates that their metabolism, in its
broadest sense, is different.

Which metabolic processes do determine the auxin content
of the cell? By which processes the auxin is produced and —
apart from transport to elsewhere — by which processes does
auxin disappear?

This is not only a problem in the eoleoptile of the Gramineae,
but it presents itself everywhere auxin occurs. It can only be
solved by comparing the metabolism of two cells with different
auxin contents. Of course those cells, which differ only in their
auxin content and as far as possible not in other characters,
would be most suitable for such a comparison. In that case the
probability would be greatest that differences eventually stated
in the metabolism of these cells really may correlate with a
difference in auxin content.

This equality in other characters will be realized as good as
possible in that cases, where the auxin content of one and the
same cell is rapidly changed under the influence of an external
factor. Such cases, however, rarely occur.

An example of it is the so called regeneration of the physio-
logical tip, as discovered by Rothert (1896). Dolk (1926) has
investigated this phenomenon more thoroughly. He cut off the
upper zone of decapitated coleoptiles and determined the amount
of auxin, delivered by this small cylinder. He found that the
upper cylinder produced no or only little auxin, if the coleoptiles
were decapitated shortly before. If, however, the decapitation
preceded a considerable time before, the upper cylinders proved
to deliver auxin in appreciable amounts. This really is a case,
in which the auxin content of certain cells increases rather
rapidly and that, therefore, is suitable for a research into the
problems mentioned above. This phenomenon thus has been

investigated by several authors. Soding (1929) e.g. stated that
the amount of auxin produced by the regenerated tip was in-
dependent from the length of the decapitated end, which varied
from 1 to 5-6 mm. Tsi Tsung Li (1931) studied the influence of
temperature upon the velocity of the regeneration of the phy-
siological tip. Skoog (1937) succeeded in suppressing the re-
generation of the physiological tip by removing one day before
the endosperm from the seedlings. Such „deseededquot; plants were
used as test plants in the following experiment. Cylinders of
normal coleoptiles were placed with their apical cut surface
upon agar slides for some time. Blocks of this agar then were
placed unilaterally upon deseeded coleoptiles. In the begirming
these plants behaved as if no auxin was present in the agar.
This is in full agreement with Van der Wey's (1932) experiments,
who stated that no auxin is tranported in apical direction. There-
fore no auxin could diffuse into the agar from an apical cut
surface. Two hours after placing the agar blocks on the deseeded
coleoptiles, however, a curvature started, which gradually in-
creased. The agar thus proved to contain a substance, which
gradually is transformed into auxin. In this way Skoog showed
that auxin is produced from a „precursorquot;, which is transported
from the seed to the apex. These investigations on the regenera-
tion of the physiological tip have yielded important data on the
origin of auxin in the coleoptile.

Another case in which cells rather quickly change their auxin
content is the effect of glucose upon the auxin content of root
tips mentioned above. Research on this subject is still scanty

Boysen Jensen (1933a), discovering this phenomenon, supposed
in the beginning that the normal root tip was too poor in plastic
material to be able to produce auxin. Stating later on, that
mannite had the same effect as glucose, he gave up this idea
since mannite probably could nôt be converted by the cells of
the root tip. He then ascribed the increase of the auxin delivery
to some physical influence, e.g. an improved contact between
root tip and agar, if the latter contained some hygroscopic sub-
stance like glucose or mannite.

Also Thimann (1934) ascribes the influence of glucose on the
auxin production to some „physic (osmotic) action.quot;

Cholodny (1934) on the other hand, believes that glucose acts
as a nutrient, since root tips with glucose continue to deliver
auxin for a longer time than without.

The mechanism of the glucose effect on the auxin production
has only been studied occasionally by these three authors.

Although Boysen Jensen's discovery has been of great im-
portance for a better understanding of the auxin function in
the root, till now it has not been applied to elucidate the pro-
cesses, which determine the auxin content of the cells. That was
the aim when starting the investigations reported in the present
paper.

It seems appropriate to resume briefly what was known about
auxin in the root before Boysen Jensen's discovery. For a more
detailed survey of the literature must be referred to Gorter
(1932) and Boysen Jensen (1935).

The literature on growth and tropisms of roots of the last ten
years — since 1926 — is reigned by discussions on Cholodnys'
theory (1924; 1926). According to this theory the positive geo-
tropism of the root is explained as follows. In the root tip the
same hormone would be produced as in the tip of the aerial
organs. This hormon accelerating the growth of the cells in the
latter, would retard the growth in the root. By placing the root
horizontally, the hormon, as in the stem, would shift to the
lower side, which consequently would be more retarded in
growth than the upper side. This would result into a positive
curvature.

The presence of auxin in aerial organs and the inequal distri-
bution of this substance under the unilateral influence of gravity
has been proved by the work of Went (1928) and Dolk (1930).
The proof of their experiments is chiefly based on the method
devised by Went to test quantitatively the amount of auxin in
tissues. To this purpose the tissue to be tested is placed for a
certain time on a slice of agar, which afterwards is divided up
into small agar blocks, which are placed unilaterally upon a
decapitated coleoptile of Avena. This curves negatively under
the influence of the auxin present in the agar. The magnitude
of the curvature, is, within certain limits, a measure for the
amount of auxin in the agar block. By means of this method
the presence of auxin in any organ can be stated directly. Before
Went applied this method already theories were formulated,
explaining tropistic movements by means of chemical substances
(e.g. Boysen Jensen (1911); Pabl (1919). The data endorsing
these theories, however, were obtained only indirectly and were
therefore more liable to criticism than the experiments of Went
and Dolk, who really isolated the auxin from the coleoptile of
Avena.

It was obvious to try to judge the correctness of Cholodnys'
theory on the action of auxin in the root by means of the same
direct method. This proved, however, to meet with great diffi-
culties. Hawker (1932) succeeded in recovering auxin in this
way from root tips. In her experiments geotropieally stimulated
root tips of Vicia Faha were split and the two halves were put
on gelatine for some time. Then the blocks of gelatin were placed
unilaterally upon the stumps of decapitated roots. A curvature
resulted in the direction of the gelatin block, the curvature being
larger with gelatin blocks, on which the lower half of the root
tips had been placed, than with those, on which the upper halves
had stood.

Contrasting to these experiments are those of Gorter (1932).
She tried to isolate auxin from root tips of Pisum and Zea by
placing pieces of roots on agar or sand. In the latter case the
sand was washed with water and ether; this extract was mixed
with agar and its auxin content tested with Avena coleoptiles.
Both methods failed to show the presence of auxin in roots.

The results of these affords to isolate auxin from roots are
contradictory. The same applies for the data obtained with in-
direct methods.

Cholodny himself (1924, 1926) based his theory on the following
experimental results. Decapitated roots of Lupinus did not curve
geotropieally; the ability to curve was restored by placing a tip.
of a eoleoptile of corn on the cut surface. Decapitated roots of
corn grew under water and in humid air markedly quicker than
intact roots. By sticking a eoleoptile tip on the cut surface the
growth rate was slackened again. A new proof was given in
1928: a decapitated root of corn was placed horizontally. A tip
of a eoleoptile of corn was fixed on it by means of gelatin in the
normal (vertical) position. Although the eoleoptile tip was not
stimulated geotropieally, the root curved positively geotropieally.
This proves in the mean time that the curvature is not to be
ascribed to specific geotropic stimulating agents. Coleoptiles of
Avena were decapitated and the tips replaced by those of roots
of corn. The phototropic and geotropic responses proved to be
much larger than without these root tips.

Snow (1923) also had discovered already that decapitated roots
of Vicia Faba regained their geotropic sensitivity by replacing
their tips.

biinning (1928) found in a number of plants that after deca-
pitation the root shows a retardation of growth or even a con-

traction first, but that the rate of growth afterwards increases
to a higher value than that of the normal root. He ascribes these
phenomena to wound growth reactions and does not believe
that the root tip produces growth accelerating or -retarding
substances. On the other hand he also states that the rate of
growth of decapitated roots of Lupinus is decreased by replacing
their tips on them. The same effect, however, is produced by
placing, in stead of tips, any other section of a root on the cut
surface.

Cholodny (1929) repeats BtInning's experiments, but he finds
that tips of roots placed on decapitated roots decrease the growth
rate and that cylinders from other sections of the root have no
effect upon the growth rate. BiiNNiNc's results are due by Cho-
lodny to deficiency of his material.

Gorter (1932) did not find that decapitation affects the rate
of growth of roots of Pisum. Further she stated that the geotropic
response of roots decapitated at only 1 mm from the root tip
equalled that of normal roots. Root tips of Pisum, unilaterally
placed on decapitated coleoptiles of Avena did not produce a
curvature. The geotropic reactivity of decapitated coleoptiles
was not increased either by placing root tips on them.

Also on this criticism Cholodny replied (1933). He deduced
from Gorter's growth curves that after decapitation indeed an
acceleration of the growth of the roots did occur. Moreover in
her experiments the growth rate showed a constant decrease,
even in not decapitated roots, a fact, which Cholodny ascribes
to lack of humidity in the milieu. Also the fact, that root tips
failed to produce curvatures in decapitated and horizontally
placed coleoptiles, is ascribed by Cholodny to too dry experi-
mental conditions. It is rather amusing that on the contrary
Gorter explains the positive result of Cholodny in the latter
experiment to exsiccation of the decapitated coleoptiles, on which
no root tip was placed, so that they could not curve.

From these discussions it will be clear, how difficult it is to
prove the presence of auxin in the root by indirect methods.
Boysen Jensen's discovery (1933 a), mentioned above, however,
took away all uncertainty on this point. He placed root tips of
corn on agar to which 10% of glucose was added. The tips
delivered considerable amounts of auxin to this agar. The results
were still better with agar 10% of glucose 0,1% of Ca(N03)2
0,025% of K2HPO4 0,025% of MgS04 a trace of FeClg.
A root tip of corn delivered more auxin to this agar than a tip
of a coleoptile of Avena.

Boysen Jensen (1933 b) also with the aid of glucose agar in-
vestigated the influence of gravity on the distribution of auxin
in horizontally placed root tips. To this purpose a geotropically
stimulated root tip was placed on two separated agar blocks.
In one block the auxin from the upper half, in the other that
of the lower half of the root tip was received. He found that the
lower half delivered more auxin than the upper half.

By these experiments of Boysen Jensen the presence of auxin
in the root tip as well as the unequal distribution by geotropic
stimulus was proved. Since Kögl, Haagen Smit and Erxleben
(1934) had shown that auxin inhibits the growth of roots,
Cholodny's theory on the function of auxin in the root was
completely confirmed. Undecided, however, remained the ques-
tion, whether auxin really is produced in the tip of the root.
The literature on this point will be discussed in chapter III.

For the estimation of the auxin content of the agar blocks the
„Avenaquot; test was used as described by Went (1928), later on
improved in some details by Van der Wey (1931). For the test
the pure line of oats, „Siegeshaferquot; was used from Svalöv, kindly
supplied by Prof. Dr. A. Akerman, ass. director of the Experi-
ment Station of the „Svensk Utsädes Föreningquot;.

The coleoptiles were decapitated three times, with intervals
of ly^ hours. Immediately after the third decapitation the agar
blocks were placed upon the stumps. These agar blocks were
obtained by deviding up an agar slice of 8 X 6 X 0,9 nam into
12 exactly equal parts. In this paper an agar slice means a layer
of agar of the given dimensions, an agar block is 1/12 part of it
and therefore has a volume of 3.6 mm'.

The test plants were photographed 1 h 50 min after placing
the agar blocks upon the stumps. The curvatures were measured
by means of a protractor. In the tables the auxin quantities
always are expressed by the curvatures (in degrees) of the test
plants. These figures are the averages of 10-20 plants, the mean
error being calculated from the formula

The root tips, used in my experiments, were all of Vicia Faba
L. The variety used was „Origineele Mansholt's Wierboonenquot;

The seeds were soaked in water for 24 hours and than planted
in saw dust. For the experiments the root tips of 5 days old
plants were used. The beans were taken from the saw dust and
thoroughly washed under the tap; then the tips were cut at a
length of 5 mm. To this purpose the roots were placed over a
glass slide and a piece of calibrated (in mm) paper. A razor
blade was used to cut off the tips.

1)nbsp;A number of root tips (mostly 7) were placed for some time
on an agar slice; then the auxin content of the agar was deter-
mined. When the influence of glucose on the delivery of auxin
was investigated glucose Ph. Ned. Ed. 5 was used. In order to
obtain a certain concentration of glucose, in the beginning the
amount of glucose wanted was added when preparing the agar
(3%). Later on the agar slices simply were soaked in solutions
of glucose of the desired concentration. The latter procedure also
was followed, when the influence of salts on the delivery of
auxin was studied. In this way it is not certain that the total
concentration in the agar slice is the same as that in the solution,
but it is probable that the free fluid, not adsorbed to the col-
loidal agar particles, indeed has the same concentration.

2)nbsp;A number of root tips were extracted with ether, according
to prescriptions by Dolk and Thimann (1932), Kögl, Haagen Smit
and Erxleben (1933) and Thimann (1934). Briefly the method
may be described as follows: the root tips are ground with a
little acid and ether, free from peroxides. When the tips had
been suspended in a fluid as was the case in the experiments
on respiration, also the fluid was acidified and washed with
ether. The ether was decanted and the extraction repeated twice
with new ether. Finally all the ether was evaporated and the
résidu solved in 0,2 cmquot; of a buffer solution. Two agar slices
remained over night in the refrigerator in this solution. On the
next day their auxin content was determined.

The ether was freed from peroxide by destilling it shortly
before the extraction over FeS04 and CaO.

For extraction the pulp of the root tips was acidified in order
to liberate the auxin, which eventually was bound to alkali. In
the experiments of chapter III this was done by adding a few
drops of hydrochloric acid. Then the extract was placed over
night in the refrigerator over ignited Na2S04. This method
proved to be not entirely reliable; some times the auxin had

disappeared from the extract. To eUminate this objection in the
other experiments a few drops of 0,5 n sulphuric acid was used
instead of hydrochloric acid Since this acid is not volatile,
it could not be removed from the ether by evaporation. For
that reason the extract was washed twice with water, acidified
with sulphuric acid to the conversion point of congo red.
Although in this way most of the sulphuric acid was removed
from the ether, still the danger remained that a last trace of
this acid would destroy the auxin after the evaporation of the
ether.

In order to prevent this, the method of evaporation was
changed a little. The extract was not entirely evaporated, but
only so far that the volume was about 5 cm\ This rest was put
into a small tube and 0,2 cmquot; of the buffer solution was added.
This tube was kept in a beaker with warm water and the
evaporating ether was blown away by means of an air current
In this way the auxin was dissolved in the buffer solution without
evaporation of the extract to dry. In order to remove the water
insoluble substances as far as possible, the preparation was
washed with petrolether, which afterwards was decanted (ac-
cording to Kögl, Haagen Smit and Erxleben (1933), auxin is
insoluble in petrolether).

The buffer, in which the auxin was solved, served to eliminate
the eventual last traces of acid. It was a diluted buffer after
McIlvaine, a solution of 0,04 mol citric acid and 0,02 mol
Na2HP04; its pH was ± 5,4.

I often succeeded in extracting by means of the described
methods a substance from root tips, which shows auxin activity.
From investigations of recent years it is, however, known, that
a number of chemically widely different substances show a
positive result in the Avena test (e.g. Haagen Smit and Went,
1935). Therefore it is not quite certain that the growth sub-
stance from the root really is identical with auxin. This is,
however, made highly probable by the investigations by Heyn
(1935), who estimated the molecular weight of the growth sub-
stance from the root by means of its diffusion rate through agar.
He found as an average for the diffusion coefficient the value
D = 0,391, which corresponds to a molecular weight of i 330.

1)nbsp;Perhaps it would be advisable to add in future in experiments of this
kind diluted acetic acid.

2)nbsp;The blowing off of the ether vapour by means of an air current and
the application of petrolether were suggested by Dr. J. MacPherson
Robertson, to whom I feel much indebted for his valuable advice.

§ 1. The influence of different glucose concentrations on the
delivery of auxin by root tips.

Boysen Jensen (1933 a), when discovering that the delivery
of auxin by root tips could be increased by the addition of glu-
cose to the agar, applied only one concentration of glucose, i.e.
of 10%. He did not mention whether this really is the optimal
concentration.

In an earlier paper some results of experiments on this sub-
ject have been published (Van Raalte, 1936); fig. 1 represents

these results once more. In these experiments root tips were
placed on agar slices containing different concentrations of glu-
cose. Ten tips stood for two hours on each slice; then the auxin
content of the agar was determined. In the graph the glucose
content of the agar is plotted on the abcissa, the amount of
auxin on the ordinates (in degrees of the curvature of the test
plants). The optimal concentration proves to be 10% of glucose.
The fact, that agar with 15% of glucose induces smaller curva-
tures in the test plants, does not necessarily indicate that this
agar contains less auxin than that with 10% of glucose. This
smaller curvature may be caused also by an adverse influence
of the high glucose concentration on the test plants. Such an
influence was apparent in control tests. Unilateral application
of agar blocks, containing 15% of glucose but no auxin, often
induced positive curvatures in the test plants. Such curvatures
would partly counterbalance eventual curvatures induced by
auxin.

In order to isolate a well measurable amount of auxin from
the root tip of Vicia, one must add glucose to the agar. In this
regard the root tip behaves differently from other organs, e.g.
from the tip of the coleoptile of the Gramineae; the latter also
delivers auxin to the agar without addition of glucose. The ques-
tion rises what may be the reason of this difference.

Possibly the synthesis of auxin in the coleoptile proceeds in
quite a different way than in the root, the presence of glucose
not being necessary in the former. Another possibility is, that
glucose (or another reducing sugar) is required for the synthesis
of auxin in the coleoptile as well as in the root tip, the coleoptile,
however, containing a sufficient amount of these substances of
itself, the addition of extra quantities being superfluous.

A simple Feelings' test already shows the different sugar con-
tents of the coleoptile and the root tip. When one crushes some
tips of Auena-coleoptiles on a slide, adds a few drops of Feelings'
solution and heats, a strongly positive reaction appears. The same
experiment with root tips of Vicia gives a much weaker reaction;
it often is questionable, whether any reaction occurs. Apparently
root tips contain considerably less reducing sugars than tips of
coleoptiles.

It still remains open, whether the difference in sugar content
is large enough to eliminate the influence of an extra supply of

sugar on the dehvery of auxin by coleoptile tips. The next exper-
iment had to discriminate this question. Equal numbers of
coleoptile tips of Avena stood for four hours on plain agar and
on agar, containing 10% of glucose. Then the auxin content of
the agar was determined (table 1).

TABLE 1. (16-l-'34). Amount of auxin delivered by a coleoptile tip of
Avena during 4 hours.

The table shows that no favourable effect of glucose on the
delivery of auxin occurs. The result points to the contrary; it
has, however, not been investigated, why less auxin had been
delivered to the glucose agar.

In any case the former experiment shows that in the coleoptile
tip the content of reducing sugars is not a limiting factor in the
proceeded as follows. Two days after laying out the seeds for
tried to reduce this content to such a degree, that the production
of auxin would decrease. If this could be obtained, the addition
of glucose should increase the delivery of auxin.

In order to obtain plants with a lower content of nutrients, I
proceeded as follows. Two days after laying out the seeds for
germination the endosperm was removed from the seedling. The
latter was planted into saw dust. The plants treated in this way
grew well; the coleoptiles, however, being thinner than those of
normal plants; moreover they often remained a little shorter
than those. At an age of 5 days the coleoptile tips were cut. Part
of them were put on plain agar, part on agar in which certain
substances were solved. After a certain time the tips were
removed and the auxin content of the agar determined (tables
2, 3, 4 and 5).

TABLE 2. (30-l-'34). Effect of glucose on the amount of auxin delivered
by tips of deseeded *) Avena coleoptiles.
Amount of auxin delivered by 10 tips to one agar slice during:

*) These „deseedbdquot; plants are not to be confused with the deseeded
plants of Skoog (1937), as the seed was taken away in a different stage
of their development.

delivered by tips of deseeded Avena coleoptiles.
Amount of auxin delivered by 15 tips to one agar slice, during 4 hours.

TABLE 4. (26-l-'34). Effect of glucose and fructose on the amount of auxin
delivered by tips of deseeded Avena coleoptiles.
Amount of auxin delivered by 12 tips to one agar slice, during:

by tips of deseeded Avena coleoptiles.
Amount of auxin delivered by 15 tips to one agar slice during 3 hours.

When looking through the tables it is evident, that the amount
of auxin, delivered to the plain agar is, as a rule, considerably
less than the amount delivered by normal tips. Under the con-
ditions prevailing in these experiments, a eoleoptile tip delivers,
as an average, an amount of auxin agreeing with a curvature of
10° in the test plants.

When comparing the delivery of auxin by tips from agar con-
taining glucose or fructose with that by tips from plain agar,
never a distinct effect of the sugar can be stated. We therefore
did not succeed in getting the sugar content of the tip limiting
factor for the auxin production.

After the experiments described above, some experiments were
done in which also peptone or asparagine was added to the agar.
Tables 6 and 7 give the results.

It is clear that also the combination of peptone with glucose
does not exert a distinct influence upon the delivery of auxin
by the tips. The peptone containing agar always was rich in
auxin, but also the control blocks proved to exert growth-

TABLE 6. (7-2-'34). Effect of glucose peptone and glucose asparagine
on the amount of auxin delivered by tips of deseeded Avena coleoptiles.
Amount of auxin delivered by 15 tips to one agar slice during 4 hours

the amount of auxin delivered by tips of deseeded Avena, coleoptUes.
Amount of auxin delivered by 15 tips to one agar slice during 3 hours

substance activity. This is not surprising, since experiments by
Thimann (1935) showed that peptone may contain hetero-auxin,
that can originate from oxidative changes of tryptophane. Trypto-
phane itself uses to be an impurity in peptone.

In order to eliminate many possible impurities, an experiment
M^as done with peptone washed in ether and dried afterwards
(table 8).

of auxin delivered by tips of deseeded Avena coleoptiles.
Amount of auxin delivered by 12 tips to one agar slice during 4 hours

Purified peptone proved indeed to exert only a weak growth
substance effect; the agar in controls, on which no root tips had

stood, giving a mean curvature of only 1°,0 ± 0°,6. The dif-
ference in the amounts of auxin delivered to the different agar
preparations, however, has now practically disappeared. Neither
glucose or glucose combined to peptone proved therefore to be
able to increase the auxin production in the coleoptile tips.

§ 3. The influence of glucose on the delivery of auxin by coty-
ledons of Raphanus.

Another object in which was tried to detect a glucose effect
were the cotyledons of Raphanus. Van Overbeek (1933) found
that the cotyledons of seedlings of Raphanus, when kept in the
dark for some time, delivered less auxin to agar blocks on which
they were placed than those of seedlings normally grown in
the light. If the „darkquot; plants were brought back into the light,
the auxin content of the cotyledons increased again. Van Over-
beek could thus show that auxin is only formed in the cotyledons
in the light. The decrease in auxin content, however, did not
occur in the cotyledons of very young plants, kept in the dark.
In these the cotyledons had a reserve stock of auxin, stored
already in the seed. Another case, in which auxin is formed
only in the light has been described by Avery (1935) for tabacco
leaves.

Nothing is still known about the way in which light affects
the synthesis of auxin. It is an obvious hypothesis that auxin
would be formed as an accessory product in photosynthesis.

Another possibility, however, is that in the cited cases the
formation of auxin is dependent upon the content of reducing
sugars as is the case in root tips. In long lasting dark periods
the carbohydrate content of the green parts decreases, since there
is no photosynthesis. This decrease in carbohydrate content could
possibly check the synthesis of auxin. If the latter hypothesis
holds true, it should be possible to increase the auxin content
of plants, kept in the dark, by the supply of glucose. In order
to investigate this next experiment was taken.

A number of seedlings of Raphanus was kept for two days in
the dark; an equal number remained in the green house. The
seedlings were 9 days old, the age at which, according to Van
Overbeek (I.e. p. 576), the cotyledons are free from reserve auxin.
At the end of the second day (of darkness) the cotyledons of
both series were cut and on the cut surface of each cotyledon
an agar block was placed in the way described by Van Overbeek
(I.e. p. 566). One half of the cotyledons of each series got a block
of plain agar, the other half a block of agar containing 5% of
glucose. The results are given in table 9.

Glucose seems to increase perhaps the delivery of auxin a
little as well in normal as in etiolated cotyledons. The addition
of glucose, however, entirely fails to balance the deficiency of
light. Since another experiment yielded the same result, we may
conclude, that the auxin formation in the light is not caused by
an increase in concentration of reducing sugars.

In recent years a continuously increasing number of cases has
been reported in literature, in which very small amounts of
certain organic compounds would affect the growth rate. Apart
from auxin and physiologically related substances, the compounds
of the „biosquot;-complex have to be mentioned, which in very small
quantities strongly accelerate the growth of yeast and other
fungi. It could be possible that such a substance with oligodyna-
mical activity would occur as an impurity in glucose and would
affect the metabolism in the root cells in such a way, that the
latter would produce more auxin.

In order to check this possibility, the effect of the usually
applied glucose „purissimumquot; was compared with that of care-
fully purified glucose. To this purpose glucose was solved in
water, washed with ether and then recrystallized from an alco-
holic solution. The experiment was done as follows: an equal
number of root tips was placed for two hours a) on a slice of
plain agar, b) on a slice of agar containing ordinary glucose
and c) on a slice containing purified glucose. After removal of
the tips, the auxin content of the agar was determined (table 10).

TABLE 10. (27-9-'35). Effect of purified and ordinary glucose.
Amount of auxin delivered by the same number of root tips to one agar

The action of purified glucose proves to equal that of ordinary-
glucose. This proves that eventual impurities of ordinary glucose,
soluble in ether and alcohol, do not affect the auxin delivery by
root tips.

Another effect of glucose, which one could suggest, is the pro-
tection of the auxin against inactivation by enzjones. When pas-
sing from the root tip into the agar, the auxin has to travel
through the cut surface, where a great number of dead and
dying cells occurs. Now, it is a well known fact, that no or only
very little auxin can be recovered from tissues, ground in water
(Went, 1928; Thimann, 1934). Probably the auxin is destroyed
by liberated intracellular enzymes. Recently such auxin destroying
substances proceeding from dead or dying tissues actually have
been detected. Van Overbeek (1935, 1936) put tips of coleoptiles,
of which the auxin production had ceased, and eoleoptile cylin-
ders on agar blocks, containing known amounts of auxin. After
some time a part of the- auxin proved to have disappeared. He
made it probable, that this disappearance of auxin is to be due
to a destruction of auxin by oxidative enzymes, diffusing from
the tips and cylinders into the agar blocks. Larsen (1936) in a
similar way could detect an auxin inactivating substance in
wounded stems and ground tissues of Phaseolus. The next ex-
periment shows that such auxin inactivating substances also
may arise in root cells of Vicia Faba. A number of root tips is
ground to a pulpy mass. In this pulp slices of agar were soaked,
they remamed there over night at 4° C. Next day 12 tips of
Auena-coleoptiles were placed for two hours on each slice. As
a blanc the same number of tips was placed on slices of plain
agar for two hours. After removing the eoleoptile tips, the auxin
content of the agar slices was determined (table 11).

It is evident that the agar slices from the root tip pulp con-
tained some auxin inactivating substance. The character of this
substance has not been investigated further. The substance
detected by Larsen in Phaseolus is thermolabile, the inactivation
irreversible and therefore destructive. This points to an action
of destroying enzymes, as was the case in Van Overbeeks' in-
vestigations. There is some evidence, that the inactivation, stated
here, is also due to a similar enzymatic action. In cut root tips
these enzymes chiefly will be liberated in the cut surface and
here partly destroy the passing auxin.

TABLE 11. Auxin inactivating substances in ground root tips.
Amount of auxin delivered by 12 tips of Avena coleoptiles during 2 hours

Expt. 1 (17-6-'35)

Expt. 2 (18-6-'35)

Now, it might be possible that the glucose effect actually
means that the sugar prevents this destruction. If this would
hold true, this action chiefly would be confined to the neigh-
bourhood of the cut surface, i.e. there, where the tip and the
agar block are in contact with each other. In that case the glu-
cose effect should realize itself as soon as this contact is made,
which can be checked in a simple way. To this purpose the root
tips have only to stand on the agar as short a time as is required
to get a detectable amount of auxin. If the analysis shows that
even then the auxin content of glucose agar is higher, i.e. that
the glucose effect is realized in such a short time, there would
be given evidence, that the glucose acts in the neighbourhood
of the boundary surface. If however, the sugar has to travel
along a certain distance in the root tip in order to exert its in-
fluence, a certain lapse of time will be wanted, until the sugar
has covered that distance. In the latter case during a short time
after placing the root tips on the agar no or only little effect
of the glucose will be noticeable. Table 12 gives the results of
an experiment on this subject. In this experiment 10 root tips
were placed on each agar slice. After one hour already these
were removed and the auxin content of the agar tested.

TABLE 12. (13-4-'34). Amovmt of auxin delivered by 10 root tips to one
agar slice during the first hour after decapitation.

It is clear from table 12 that the glucose has only little effect
during the first hour. This is still endorsed by determining the
amount of auxin delivered to the agar during the second hour.
In an earlier paper (Van Raalte, 1936, p. 264) the results of
such an experiment have been reported; they are reproduced in
fig. 2. In this graph the glucose concentrations are plotted on

the abcissa, the amount of auxin on the ordinates. Comparison
of the curves for the first and the second hour teaches that the
effect of glucose mainly demonstrates itself in the second hour.

These results indicate that diffusion should proceed for some
time, before the glucose reacts. One can imagine that first a
sufficient amount of glucose has to travel upw^ard in the root
tip, but also that first a sufficient amount of substances (pre-
cursor?) has to diffuse from the tip into the agar, where they
would have to be transformed into auxin. The latter idea,
however, does not agree with the facts, as could be shown in
the following way.

A number of root tips of Vicia stood for two hours on a slice
of pure agar. Then they were removed and for another two hours
the agar slice was covered by a second agar slice, containing
10% of glucose. All kinds of substances from the root tip were
present in the first slice. If these substances could react with
the glucose from the second slice, auxin had to be formed in

the agar. The analysis, however, showed that this is not the case.
This proves that the glucose effect is not realized in the agar.
It seems therefore probable that the glucose has to travel up-
ward in the root tip to exert its effect upon the auxin content
of it.

This view is endorsed to some degree by the fact that one
also can make diffuse the glucose previously into the root tip
and catch the auxin afterwards in a slice of plain agar. This
experiment was taken as follows: 12 root tips were soaked for
two hours in a 10% solution of glucose, then they stood for
another two hours on a slice of plain agar. For comparison
another set of 12 root tips was placed immediately after cutting
on plain agar, also for two hours, a third set for the same time
on agar containing 10% of glucose (table 13).

TABLE 13. (8-12-'33).Amount of auxin delivered to plain agar by root tips
to which glucose had been applied previously.

Resuming we may conclude that probably the glucose does
not exert its influence upon the delivery of auxin in the agar
and neither in the boundary between agar and root tip.

In the preceding chapters the terms „productionquot; and „deli-
veryquot; of auxin have been intermingled. In fact this was not cor-
rect. If one discusses the fate of auxin in the root tip, it is
necessary to discriminate between the symptoms observed and
the supposed internal processes.

Above the delivery of auxin meant the transit of auxin from
root tips into the agar. The glucose effect showed itself in an
increased delivery: the agar slices with glucose received more
auxin from the root tips than those without glucose.

What can be the cause of this increased delivery? The answer
depends upon the question whether or not auxin can be synthe-
tisized in the root tip from other substances. This process, the
synthesis of auxin from other substances, further will be called
auxin production.

increase. In that case probably there would be also one or more
processes, which make the auxin concentration decrease. Pro-
cesses of this kind will be indicated as auxin inactivation. It
will, however, not be tried to discriminate whether 1) auxin is
really inactivated or destructed, or 2) it is consumed in some
growth process, or 3) the reaction precursor auxin is accom-
panied by an opposite reaction: auxin precursor.

Now, the glucose effect can be due to different possible causes.
These possibilities are surveyed below:

This survey shows, that it is of evident importance to know,
whether the root tip can produce auxin or not. In the latter case
the factors II 3 and II 4 can be excluded and only the factors
1 and 2 remain open to further research.

The possibility of auxin production in the root tip for years
has been a much discussed problem. A priori there is no reason
to exclude this possibility. Since the root tip, however, does
give off very little, if any auxin to plain agar, it was impossible
to demonstrate auxin in the root directly, until Boysen Jensen
(1933 a) showed, that the auxin delivery is increased by glucose.
The indirect methods, earlier applied, yielded contradictory-
results.

Two theories were given on the topographic origin of auxin,
eventually present in the root tip. The older one by Cholodny
(1926), who stated that a decapitated root grows faster than a
normal one. This increase in growth rate was stopped by placing
a tip of a coleoptile of com on the cut surface. Cholodny con-
cluded, that the root tip produces auxin in the same way as the
tip of the coleoptile does. This hormone, causing an increase of

1.nbsp;translocation of auxin from
the tip to the glucose agar,

2.nbsp;a decrease of auxin inacti-
vation under the influence
of glucose.

the growth in the coleoptile, would, however, decrease the growth
rate in the root. In a root, declining from the vertical position,
the lower half would show a higher auxin concentration than
the upper half. The growth in the lower half would be retarded
more than in the upper half, which would result into a down-
ward curvature of the root tip. Cholodny thus assumes auxin
production in the root tip.

Went (1932), however, gives quite a different theory on the
origin of auxin in the root. He concluded from the literature
and from own experiments, that there would exist in the plant
an electric potential difference between the tip of the stem and
the tip of the roots, the potential of the root tip being positive,
that of the stem tip negative. Under the influence of this P.D.
positive and negative ions would travel cataphoretically in op-
posite directions. The kations would move towards the region
of growth in the stem, the anions towards that of the roots.
Auxin bearing an acid character, Went assumes, that the (nega-
tive) auxin ions move continuously from the upper parts of the
stem to the zone of growth in the root. According to this con-
ception no auxin is formed in the root tip and, if once present
there, the auxin cannot move away.

This theory seemed to be endorsed by investigations of Thi-
mann (1934). This author extracted root tips by means of chloro-
form. The quantities of auxin, obtained in this way, were com-
pared with those delivered by the same number of root tips to
glucose agar. The quantities delivered to the agar never exceeded
those obtained by extraction with chloroform. From this Thimann
concluded, that auxin is not produced in the root tip, but that
it diffuses from the seed and accumulates in the tip. The glucose
effect was explained by the osmotic force of the glucose, which
would — as has been mentioned above — extract the auxin
from the root tip against the original polar direction of its
transport.

Thimann suggests still another possible explanation of his ex-
periments. One can imagine, that auxin is formed in the root
tip, but that the stock of „precursorquot; is so small, that it is con-
sumed quickly after cutting off the tip. So Thimann actually
considers the possibility of auxin production in the root tip. He
stresses, however, his first hypothesis so much, that this one
is often cited in literature as the conception of Thimann.

périment is the most conclusive one: roots of corn are detipped
and then a cylinder of 8 mm long is cut from the apical end.
These cylinders are placed horizontally. A tip of a corn coleoptile
is placed either on the apical or on the basal cut surface, an
agar block on the opposite surface. After some time the agar
blocks on the basal surface do contain auxin, those on the apical
surface do not. This proves that in the root of corn auxin is
only transported from the tip to the base, that means: the pola-
rity is just opposite to that postulated by Went's theory. No
auxin can reach the root tip from without; it can only be syn-
thetisized from other substances on the spot.

Boysen Jensen (1936) repeats Thimann's experiments on Avena
roots with roots of Vicia. He states, that the quantity of auxin
delivered by a root tip during 20 hours is 21 times as large as
that, obtained by extraction with chloroform. Also in this case
auxin must be produced in the root tip.

Nagao (1935) isolated from roots of Vicia small cylinders, 2—6
mm from the tip. Those cylinders are placed on glucose agar,
either with the basal or with the apical cut surface. Much more
auxin diffuses from the basal surface than from the apical one.
Also in Vicia the polarity of auxin transport runs from the tip
to the base of the root.

Nagao also repeats Thimann's experiments with Avena roots.
His results with Avena are quite different from those of Thimann.
He states that the quantity of auxin, delivered by the root tip
in one hour, is twice as large as that extracted with chloroform.
Moreover a tip on agar continued for at least 6 hours to deliver
auxin to the agar. Being obtained with the same object, this
result directly contradicts Thimanns'.

Cholodnys' theory is opposed by a recent paper of Fiedler
(1936), who cultivated excised roots in pure culture. The roots
developed very well, they also showed a geotropic response, but
the extraction of auxin by means of Thimanns' chloroform method
failed. It was proved that in corn and pea the auxin disappeared
from the roots within 24 hours after their excision. Fiedler ex-
plains this disappearance by a diffusion of the auxin to the cut
surface, where it is destructed by oxydases. The fact, that no
auxin is regenerated, proves, according to Fiedler, that the root
tip has no ability to produce auxin.

It is clear, that the data in the literature on the problem of
auxin production in the root tip are not conclusive. Therefore
all possibilities, mentioned on p. 299, must be considered for
an explanation of the glucose effect. For that reason the first

possibility was investigated: is the glucose effect caused by a
translocation of auxin from the tip to the agar, without any
change in the total amount of auxin? (i.e. Thimanns' prevailing
explanation of the glucose effect; see p. 300).

This being true, root tip and agar slice may be compared with
communicating vessels: an increase of the auxin content of the
slice can only coincide with a decrease in the tip. Since more
auxin is delivered to glucose agar than to plain agar, the auxin
content of root tips from glucose agar should be smaller than
that of root tips from plain agar.

It has been investigated, in the following experiments whether
this holds true or not. Different numbers of root tips were
placed during a certain time on slices of plain agar or of agar
containing 10% of glucose. Afterwards thfe tips were ground and
extracted with ether and the obtained quantity of auxin tested.
Table 14 gives a survey of the data obtained.

TABLE 14. Amount of auxin present in root tips from glucose containing
agar and from plain agar.

The last column of table 14 gives the ratios of the mean auxin
content of one root tip from glucose agar to that of one tip from
plain agar (the tips stood on the agar during the same time).
Not always the same numbers of tips were extracted. This is
due to the fact, that only limited concentrations of auxin can be
determined quantatively in the Avena test. The curvature of the
test plants, caused by this concentration, was called by Went

the „Hmit-anglequot; („Grenzwinkelquot;). The size of this angle varies
from day to day and cannot be predicted. By extracting different
numbers of root tips almost always a cj^uantity of auxin could
be obtained, which yielded a smaller curvature than the limit
angle. For this reason it was not always feasible to compare
equal numbers of tips from glucose agar and from plain agar.

Column 5 of table 14 shows, that root tips from glucose agar
contain 2—4,5 times as much auxin as those from plain agar.
The glucose effect therefore is not restricted to a higher delivery
of auxin to the glucose agar, but it also increases the auxin
concentration in the tips themselves. It cannot be a mere trans-
location of auxin from the tip to the agar.

In the preceding experiment, however, the higher auxin con-
tent of tips from glucose agar was only relative, in comparison
with that of tips from plain agar. It remains open, whether this
must be ascribed to a real increase of the quantity of auxin
on glucose agar, or to an inactivation of auxin on plain agar,
which would be prevented by glucose. In order to investigate
this question, next experiment was made. 60 root tips were ex-
tracted with ether immediately after cutting. Another set of 60
tips first remained during two hours on agar, containing 10%
glucose, and then were extracted with ether, together with the
agar on which they had stood. (Table 15).

From this experiment can be concluded, that the glucose effect
consists in an increase of the quantity of auxin in the root tip.
This increase of the quantity of auxin in a root tip, isolated from
the plant, can only be explained by the assumption, that in the
root tip production of auxin, i.e. synthesis from other substances,
has taken place.

This result completely agrees with the experiments of Cho-
lodny, Boysen Jensen and Nagao. It is hard to explain Thimann's
different results. Du Buy and Nuernbergk (1935, p. 347) suppose,
that Thimann did not change the agar slices, on which the roots
were standing, frequently enough. This could have caused such
a high auxin concentration in the agar, that the auxin production

was hampered, the equihbrium precursor ^ auxin being shifted
to the left. With greater evidence Nagao suggested, that the
auxin would be consumed or inactivated during the transport
in the root tips, which, in Thimanns' case, were 10 mm long. In
fact, this author did not succeed either in isolating auxin by
diffusion into agar from root tips of 10 mm length, but it was
readily isolated from 2 mm tips.

Thimann's results being probably to be ascribed to his method,
this does not hold for Fiedler's conception. He also believes that
no auxin can be produced in the root tip and says literally:
„Durch die dargelegten Versuche ist wohl bewiesen, dass die
Wurzelspitze als ein ständig Wuchsstoff produzierendes Organ
nicht in Frage kommen kannquot;. (I.e. p. 426). I do not believe that
this conclusion from Fiedler's experiments is justified. As all
other organs, which do not contain chlorophyll (the coleoptile
of the Gramineae included), the root depends for the production
of its organic material upon the substances supplied by the seed
or by the green parts. After an interruption of this supply at
some time the stock of these substances in the root must become
exhausted and the normal production of organic material will be
checked. This holds as well for auxin as for other substances,
e.g. cellulose. The statement „dass der Wurzel nicht als ein
ständig Zellulose produzierendes Organ in Frage kommtquot; can
be considered as superfluous. The problem is not, whether the
root is able to produce auxin, cellulose etc. without the aid of
the green organs or the seed. The question in consideration is,
whether these substances are delivered as such to the root, or
synthetisized in the root from other substances, supplied by dif-
ferent parts of the plant to the root. In the latter case it would
be allowed to say, that auxin, cellulose etc. are produced in the
root.

In Fiedler's experiments, the only organic substance supplied
to the root is glucose. In his case no auxin could be recovered
from roots 24 hours after their excision. That only means that
the root with glucose as its only carbon source failed to produce
auxin so quickly, that the production surpassed the consumption
or inactivation. It seems improbable that only glucose would be
transported from the seed or from the green parts to the roots;
the transport of a large number of substances is probable. The
possibility that the synthesis of auxin proceeds quicker from
one of the other substances than from glucose is great; accor-
ding to Thimann this substance can be indicated as the „pre-
cursorquot;.

According to this view, the production of auxin in the root
is not fundamentally different from that of other organic sub-
stances. This also holds if glucose is a more suitable precursor
of many other materials than of auxin. Also the fact, that ap-
parently the stock of the precursor of auxin is quickly ex-
hausted, cannot be an objection to the term „auxin productionquot;
in the root tip. Also excised tips of Avena coleoptiles rather
soonly stop to deliver auxin (Van Overbeek, 1935).

The next experiment shows, that the delivery of auxin by the
root tips is continued indeed for several hours after cutting:
two sets of 8 root tips were placed on an agar slice, containing
5% of glucose. From time to time the agar slice was replaced by
a fresh one, the former being kept in the refrigerator. At the
end of the experiment the agar slices were tested on their auxin
content (table 16).

Table 16 shows, that the maximum of the delivery of auxin
is reached in the third hour; later on it decreases, but it con-
tinues during 7 hours after cutting the tips.

1)nbsp;the glucose effect is not only due to a translocation of
auxin from the tip to the agar,

The Relation Between Glucose Content, Respiration and Auxin
Production of the Root Tip.

In the preceding chapter has been shown, that auxin is pro-
duced in the tip of the roots of Vicia Faha and that the auxin
content increases under the influence of glucose. Considering
the possibilities, resumed on p. 299, it is clear that this influence
of glucose may be caused by 1) an increased production of

It is very probable that both of these two processes depend
upon other metabolic processes in the root tip. It therefore
seemed reasonable to investigate, whether also some other meta-
bolic process is influenced by adding glucose.

For this purpose the respiration has been chosen, being the
best known and most accessible process. In the experiments,
referred in this chapter, the following questions have been in-
vestigated:

2)nbsp;whether the same effect on the respiration could be pro-
duced by some other agent than glucose,

The respiration of root tips was estimated by means of the
manometrical method after Warburg (1928 b).

Open manometers were used, one of them acting as a blanc control for
temperature and barometer alterations. Brodie's fluid was used for the
manometer readings. The volxmies of the vessels, determined by wheighing
their mercury capacity, varied from 16,89 to 22,18 cm».

During the experiments the vessels were placed in a waterbath of con-
stant temperature, the fluctuations in temperature as a rule not exceeding
0,03° C. Most of the experiments were made at 23° C.

The manometers were shaken at a high rate (200 times per minute),
the amplitude of the oscillation, however, being small.

For the determination of the oxygen consumption 0,2 cm' of a 20% KOH
solution was put in a small central well in the vessel for the absorption
of the carbon dioxide developed.

The carbon dioxide was estimated by expelling it by means of sulphuric
acid. Three vessels containing equal numbers of root tips were treated as
follows. Two of the vessels contained 0,2 cm^ 0,5 n sulphuric acid in their
side bulb, the third one KOH in its central well. Immediately after the
first manometer reading the amount of carbon dioxide, present in the first
vessel at the start of the experiment was determined in the usual way.
At the end of the experiment the carbon dioxide in the second vessel was
estimated. The consumed quantity of oxygen is known from the alteration
in pressure in the third vessel.

The manometer readings were estimated to an accuracy of 0,1 mm. The
error in the readings cannot exceed 0,3 mm, that means an error in the
estimation of the volumes of 0,6 mm^ at most.

Equal numbers of root tips were used in each experiment;
they were suspended in 2 cm® of fluid. This fluid was either
destilled water (destilled from glass over glass) or a solution of
the substance, the influence on the respiration of which was

tested. Buffer solutions soonly proved to affect the auxin pro-
duction; for that reason the root tips could not be suspended
in such a solution.

In order to determine the auxin content of the tips, they were
taken from the vessels at the end of the experiment, ground
and extracted with ether. The fluid from the vessels was also
washed with ether.

The respiration figures were not calculated upon units of
fresh or dry weight. In the tables the total respiration is given
in mm® per hour of the total mass of root tips, present in each
vessel. The root tips were cut as accurately as possible over milli-
meter paper at a length of 5 mm from the tips. Since it was
impossible to apply in each experiment root tips in exactly the
same stage of development, the intensities of respiration of the
different sets of experiments cannot be compared with each other.

It was still questionable, whether a number of root tips of the
same age and from the same culture shows the same respiration.
This depends on the question, whether the number of root tips
is large enough to eliminate the influence of individual variabi-
lity. In order to estimate the influence of this variability some
experiments were done, in which the respiration of equal num-
bers of root tips was compared (tables 17, 18 and 19).

TABLE 19. Oxygen consumption of 5 lots of 10 tips. Liquid medium
2 cc. phosphate buffer, concentration N, p.H. 6,8.

total respiration in 2^ hours, mm^ 177,3 176,0 186,6 197,8 186,5
largest difference: 197,8 — 176,0 = 21,8 = 12,5%.

The tables show that the values obtained for vessels, containing
equal numbers of root tips, differ rather much from each other.
For that reason always, if possible, 30 tips per vessel were used.
With this number the error due to variability can be estimated
on 10%. If only the oxygen consumption and not the carbon
dioxide production had to be determined, the determination.s
could run in two or three parallel readings, the average figures
thus obtained having a greater probability and a smaller error
than 10%.

A serious impediment in these experiments is the relatively
large diameter of the root tips (± 2 mm at the base), by which
the gaseous exchange in the central cells is seriously hampered.
Warburg (1928 b, p. 105) especially emphasized the objections
against experiments on respiration with objects of considerable
diameter. The centrally located cells are subjected to oxygen
deficiency and respire in a different way from the cells in the
periphery. If in such a case changes in the respiration are stated
one cannot discriminate, whether the respiration of all cells has
been changed or whether the number of normally respiring
cells has been changed. This objection could be eliminated by
slicing the root tips into thin slices. In our case, however, this
would be inadequate, since probably the auxin production
seriously would be affected by the large number of deteriorating
cells. The objection of a hampered diffusion could therefore in
our case not be eliminated. This will be taken into consideration,
when discussing the results of the experiments.

Another possible source of error was the influence of bacteria.
The seeds were not desinfected and also the saw dust, in which
the roots grew, was not sterile. The possibility of the develop-
ment of a considerable quantity of bacteria in the vessels during
the experiment is not to be excluded, so that their metabolism
could interfere with the respiration of the root tips. In order to
investigate this possible influence, 20 root tips were put in each
vessel with 2 cm' of water or of a 7% glucose solution. During
two hours the respiration of the tips was measured in the ther-
mostat, then the tips were removed and the vessels with the
residual fluid were placed again in the thermostat. It then was

controlled whether still oxygen consumption or carbon dioxide
production occurred, which had to be ascribed to bacteria,
developed in the fluid. The result was negative. Since most of
the experiments did not last for a longer time than 2]/^ hours,
one may conclude that no disturbing effect of bacteria occurred.

§ 2. The influence of glucose on respiration and auxin produc-
tion of the root tips.

It has already been shown (p. 294), that the glucose effect is
not due to impurities present in the glucose. Still the glucose
powder, used in the experiments described in this chapter, was
treated with alcohol and ether to get it free from adhering
impurities.

Table 20 represents the results of three experiments, in which
the effect of different glucose concentrations on the oxygen
consumption of root tips was determined. They show that glucose
gives a depression of the respiration.

It is clear that the root tips, during the estimation of their
respiration, are conditioned quite differently from those on agar
during the delivery of auxin. In the latter case the tips are
almost all-round surrounded by the air, but during the experi-
ments on respiration they float in a fluid. By shaking the ap-
paratus the fluid may be kept air saturated, but it remains pos-
sible that the auxin production proceeds in a way, different
from that of root tips on agar. Therefore it was necessary to
determine also the auxin content of the same tips of which the
respiration had been estimated. The experiment on respiration
being finished, the root tips with the surrounding liquid were
removed from the vessels and extracted with ether. The results
of these experiments are given in table 21.

As in the former experiments table 21 shows the depressing
effect of glucose on the respiration of root tips. The last column
shows that glucose increases the auxin content of root tips, also
under these totally different conditions. Both phenomena, how-

TABLE 21. Effect of glucose on the oxygen consumption and the auxin
production of root tips.

The figures marked by an * have been determined with a diluted solution;
the curvatures which where really obtained remained within the limit angle.

ever, do apparently not correlate in a simple quantitative way.
The depression of the respiration in all four experiments of
table 21 being of the same order of magnitude, the increase in
auxin content is widely different (from 100 to 400%).

In the preceding experiments the respiration was estimated as
oxygen consumption. The question rises whether the carbon
dioxide production is shifted in the same way by glucose, i.e.
whether the respiratory quotient remains unchanged. Table 22
gives the results of some experiments on this subject.

The second column shows that, in agreement with the results
of the former experiments, the oxygen consumption of root tips
is lower in glucose than in water. Column 3, however, shows
that the carbon dioxide production decreases much less than the
oxygen consumption; as an average the production of carbon
dioxide remains about the same. Consequently the respiratory
quotient increases.

In the literature only a few cases are mentioned, in which the
respiration was reduced by higher concentrations of the sub-
strate. In most cases the respiration is increased by the addition
of glucose. A good survey of the literature is given by Geiger—

Huber (1935). Palladin and Komleff (1902) determined the
carbon dioxide delivery by pieces of leaves of Vicia Faba, floating
on sugar solutions of different concentrations. The respiration
proved to be at its maximum at a concentration of 5%; at higher
concentrations (up to 50%) the respiration was lower.

Hopkins (1924) studied the influence of low temperatures upon
the delivery of carbon dioxide by potatoes. He found, that at
0° C the respiration increased, then being greater than at 4,5° C.
This increase continued till a maximum was reached, from this
point the respiration decreased again. Hopkins ascribes this
decrease to the sugar concentration, which increases continually
under the influence of the low quot;temperature. This suggestion of
Hopkins, however, is opposed to by Barker (1933). Barker be-
lieves, that the curve, representing for the potatoe the relation
between sugar concentration and respiration, has the shape of
a rectangular hyperbola: the respiration first would rapidly
increase at increasing concentrations of sugar, at higher concen-
trations, however, further increase in sugar concentrations would
not affect the respiration any more. Barker, however, finds a
lower respiration than could be expected from the relation sup-
posed by himself. He explains this by accepting a factor, that
would accumulate within the cells and inhibit respiration. This
factor would exert its influence mainly at higher temperatures.

of seedlings of Vicia and embryos of Phaseolus at increasing
sugar concentrations till an optimum was reached. At higher
concentrations a decrease set in.

Meyerhof (1925) stated that the respiration rate of yeast was
greater in solutions of 0.3 and 0.5% of glucose than in 10 X
higher concentrations. This was confirmed by Geiger—Huber
(1935), who also found a lower respiration at higher sugar con-
centrations (3%).

The reason of this inhibiting effect of higher sugar concen-
trations upon respiration is not known. It seems obvious to
ascribe this phenomenon to the dehydratation by concentrated
solutions. In the literature also data are found on the decrease
of the respiration under the influence of high salt concentrations.
Kosinsky (1902) found that the respiration of Aspergillus niger
was decreased for 13%, if the fungus was cultivated in a nutrient
solution with 8% of NaCl. Inman (1921) stated that the respiration
of Laminaria Agardhii was reduced in concentrations higher as
well as lower than that of sea water. These data, however, are
too scanty to be conclusive; the reason of the glucose effect on
the respiration must be left undefined.

The decrease in respiration in the experiments of table 22 was
accompanied by an increase of the R.Q. That means that the
root tips produce a quantity of carbon dioxide in excess to that
due to respiration. It is already known from the investigations
by Pasteur (1872) and Pfeffer (1878, 1885) that, like in yeast,
also in higher plants in certain cases the fermentation increases
proportionately if the respiration decreases. In the absolute
absence of oxygen this aerobic fermentation changes into an
anaerobic one. Reversely the aerobic fermentation decreases
again if the respiration increases.

The inhibition of fermentation was called „pasteur-reactionquot;
by Warburg (1926). Also in our case the increase of the R.Q.
should be ascribed to an increase of the aerobic fermentation.

We therefore may conclude upon two effects of the addition
of glucose to root tips: 1) the auxin content is increased, 2) the
respiration is reduced while the aerobic fermentation is increased.
The next question to be investigated is whether these two pheno-
mena depend upon each other.

If a correlation can be found, this may be either: 1) the higher
auxin content causes a lower respiration or 2) — reversely —
the decrease in respiration causes a higher auxin content. The
first possibility is not very probable. Investigations by van Huls-
sen (1936) and Bonner (1936) have shown that auxin has no

effect upon the respiration of coleoptiles of Avena. There is no
reason to postulate such an effect in the root tips of Vicia.

There are two procedures to investigate the second possibility.
One can try to induce the same effect on respiration by an agent
other than glucose and see afterwards, whether also in that case
the auxin content is increased. It is also possible to proceed in
the reverse way: by inducing an increase in the respiration the
auxin content should be decreased. A few experiments on this
subject are described in the next paragraphs.

Especially Warburg's work has focussed the attention upon
the specific inhibition of the respiration by HCN. By this agent
respiration is inhibited and fermentation is brought about. The
relation between HCN concentration and inhibition of respiration
recently has been thoroughly investigated by Hoogerheide (1935).
Working with yeast, he found that at increasing concentrations
of cyanic acid the fermentation is increased up to an HCN

concentration of ^^ ^^^^ (= 51 X lOquot;® mol/1). At this concen-
tration the respiration was decreased to about 22% of the
original value. At still higher concentrations of HCN the respi-
ration sank to an extreme low value, but also the fermentation

The effect of HCN on the respiration of green plants has e.g.
been investigated for Chlorella. Warburg (1919) found that HCN
has only little influence upon the respiration of this alga. Emerson
(1927), however, stated that this holds only partially: the respi-
ration of autotrophically living Chlorella not being affected by
HCN. When cultivated in a medium containing 1% of glucose,
however, the respiration of the algae was much higher. If in the
latter case HCN was added, the respiration was decreased and
brought on the same level as that of Chlorella without the
addition of glucose. GéNévois (1928) found the same for etiolated
seedlings of Lathyrus. Also in this case the respiration was
increased by the addition of glucose and the increase could be
suppressed by means of HCN. Always a residual respiration
remained, however, which could not be checked by HCN. The
decrease of the respiration by HCN in seedlings of Lathyrus was

accompanied likewise by an increase in fermentation. Also in
this object the pasteur-reaction is present. It is, however, remark-
able that the pasteur-reaction is inhibited by HCN in the expe-
riments of GéNévois. At certain HCN concentrations the fermen-
tation was already increased, whilst the respiration was not yet
affected.

Experiments. The cyanic acid was supplied by adding KCN to
the twice destilled water in which the root tips were suspended.
Table 23 shows the effect of the KCN on the respiration:

From table 23 it is clear that the respiration is decreased by
KCN and the fermentation increased. The inhibition of the res-
piration by KCN being of the same order of magnitude as that
caused by glucose (see table 22), the R.Q. is remarkably higher
in KCN than in glucose. This points to a stronger increase of
the aerobic fermentation in KCN at an equal inhibition of the
respiration. One may explain this by the assumption that HCN
inhibits the pasteur-reaction in Vicia as GéNévois found in
Lathyrus.

For the determination of the effect of HCN on the auxin
content the root tips were ground after the end of the experiment
and extracted with ether. The liquid from the Warburg vessek
was washed with ether. Table 24 gives the result of the experi-
ments.

For each experiment is indicated in the table how long the root
tips have been in the respiration vessels, that means how long
they have been exposed to the action of the KCN. The results

of the experiments are rather irregular; they all agree in one
point: the auxin content was never decreased by KCN. In the
two experiments at the top of the table 24 there was no influence
of KCN on the auxin content; the small differences being within
the experimental error. In the other experiments, however, the
auxin content of the tips in KCN was distinctly higher than that
of normal tips. From this we may conclude that the auxin content
of root tips can be increased under the influence of KCN.

§ 4. The influence of increased oxygen concentrations on the
respiration and the auxin content of root tips.

The results of the experiments of the preceding paragraph
point to an increase of the auxin content when the oxygen con-
sumption is reduced. It was now to be investigated, whether an
increase in respiration was also accompanied by a decrease in
auxin content.

The increase in respiration was obtained by replacing the air
in the Warburg vessels by pure oxygen. In small organisms the
respiration seems to be independent from the oxygen tension
within wide limits, (e.g. Kubowitz und Warburg 1929; Shoup,

1930). In higher plants the relation between oxygen tension and
respiration has also been investigated, as a rule however the
respiration was estimated, only as carbon dioxide production.
In this case this method is not applicable, since the carbon dioxide
is produced partly by respiration, partly by aerobic fermentation.
Since the latter increases if respiration decreases, the production
of carbon dioxide is an inadequate measure. This method often
yields complications.

Mack (1930) for instance found in seedlings of wheat a mini-
miim in the production of carbon dioxide between 9,8% and 20%
of oxygen, dependent upon the temperature. Also Thomas and
Fidler (1933) found in young apples a minimum in carbon
dioxide production between 3% and 5% of oxygen. At the same
time they determined the alcohol formed in these apples. At low
oxygen tensions the alcohol formation was rather high; it
decreased when the oxygen tension increased. In young apples
the alcohol formation in 3% of oxygen equalled that in air. In
mature apples fermentation was higher and even in plain oxygen
it did not quite disappear.

Especially these experiments clearly prove that the carbon
dioxide production depends upon the intensities of respiration
and fermentation both; this production is therefore inapplicable
for the estimation of the respiration if the latter is subjected to

Since normal root tips do contain only small amounts of auxm,
it would become hard to estimate the latter, if the auxin content
really decreased. For that reason tips were used, which were
suspended in a 7% glucose solution; these tips have a higher-
auxin content than those in water. Table 21 shows that the
oxygen consumption of root tips in 7% of glucose is about 30%
lower than that of tips in water.

It was not a matter of course that the respiration of root tips
in 7% of glucose could be increased by increasing the oxygen
tension. Fortunately, however, this proved to be the case.

The increased oxygen tension was obtained by conducting for
several minutes practically pure oxygen (99,5%) from a bomb
through the vessels. Immediately after filling them with oxygen
the manometers were placed in the thermostat and the estimation
of the respiration started. Table 25 gives the results.

Firstly table 25 shows that respiration in pure oxygen has
almost an intensity 2]/^ times as great as in air. From table 22
we may conclude that the oxygen consumption in water is about
143% of that in 7% of glucose. In pure oxygen an increase is

TABLE 25. Effect of increased oxygen tension on respiration and auxin content of root tips in 7% glucose solution.

Expt. 1.

(ll-8-'36;

Expt. 2.

(13-8-'36)

l]/i hours
XYj hours

hours
hours
2J/2 hours

2}^ hours
2}/2, hours

30
30

40
40
40

30
30

air
oxygen

air
oxygen
oxygen

air

oxygen

6,5° ± 0,4
3,4° ± 0,4

1,2° ± 0,9
9,0° ± 0,9
2,4° ± 0,4

12,9° ± 0,7
6,8° ± 0,5

TABLE 26. Effect of low oxygen tension on respiration and auxin content of root tips.

Expt. 1.
(30-ll-'36)

Expt. 2.
(17-8-'36)

Expt. 3.

(24-8-'36)

Expt. 4.

(25-8-'36)

134 hours
IK hours

l}/2 hours
l]/2 hours

40 minutes
40 minutes

45 minutes
45 minutes
2 hours
2 hours

IM hours
V/2 hours

hours
l]/2 hours

30
30

40
40
40
40

20
20

28
28

air
nitrogen

194,5
21,9

122,7
5,8

170,4
4,2

3,0quot;
3,9'

3,5'
2,6=

5,0°
5,3°
6,0°
5,1°

3,9°
2,5°

5,6°
2,0°

±0,5
±0,6

±0,6
±0,4

±0,6
±0,9
±1,0
±0,8

±0,6
±0,5

Expt. 5.

(28-8-'36)

Expt. 6.

(27-8-'36)

89,4
6,1

124,5
10,1

found up to 240%; that means that not only the inhibition of
the respiration by glucose is balanced but even that the respira-
tion is much higher than in normal conditions.

The last column of table 25 shows that the increased respiration
is accompanied by a lower auxin content. Also this result indicates
that the intensity of respiration affects the auxin content of the
root tips.

§ 5. The influence of low oxygen concentrations on the respira-
tion and the auxin content of root tips.

From the preceding paragraphs we know that the auxin
content of the root tips decreases, when oxygen consumption
increases and reversely, that the auxin content increases, when
the oxygen consumption decreases. The decrease of the respira-
tion was obtained by adding glucose or KCN. The inhibition
obtained was at most ± 50%. The question rises, whether the
auxin content will still increase if the respiration is still more
inhibited. The substances used to inhibit the respiration, glucose
and KCN, cannot be applied in higher concentrations, since these
probably would interfere with other processes. Hoogerheide (1935)
showed that HCN inhibits at high concentrations as well the
aerobic as the anaerobic fermentation. For that reason it was
tried to obtain a greater decrease of the respiration by reducing
the oxygen tension in the vessels, by replacing the air by
nitrogen.

The nitrogen applied did not contain more than 0,1% of
oxygen, according to the analysis by the factory. Since strictly
anaerobic conditions were not strived at, these traces of oxygen
were not absorbed; also the liquid in which the root tips were
suspended was not freed from oxygen, so that the root tips still
showed traces of respiration. Table 26 shows the results of these
experiments.

Since the oxygen consumption is extremely low the „apparent
respiratory quotientquot; is very high. The figures of the last column
represent the auxin contents of the roots tips. These prove at
these extremely low oxygen tensions to be about equal to those
in air. In one case (exp. 6) the auxin content is even lower in
low oxygen tension than in air. It seems, therefore, that the
increase in auxin content does not occur if the respiration is
reduced too much.
§ 6. Discussion of the results.

On p. 308 we discussed the drawback of the large diameter
of the root tips. The question rises, how much this factor has

The oxygen diffuses to the oxygen consuming cells through
the surrounding tissue of the root. The track of the diffusion
to the greater part of the cells proves to be so long, that the
diffusion caimot keep pace with the want of oxygen of these
cells. This follows from the increase of the respiration at increa-
sing oxygen tensions. Only the cells in the most external layers
of the root tip will receive more oxygen than wanted. One
may therefore distinguish between two zones in the root tip:
an internal one, where the oxygen tension acts as limiting factor
on the respiration, and an external one, where this process is
limited by other factors. These two zones may behave differently,
when the respiration is changed.

An increase of the oxygen tension will only have an influence
in the internal area, where the oxygen is limiting factor. Here
the respiration of the cells will increase.

KCN will exert its action mostly and soonestly upon the cells
of the periphery. By Warburg's investigations (1928 a) has been
shown, that the inhibition of the respiration by HCN exists in
a blocking up of the haemin ferment, which activates the oxygen.
At increasing HCN concentrations a continually increasing portion
of haemin ferment will be blocked. Consequently at a certain
HCN concentration the haemin ferment will become limiting
factor for the respiration. This will happen firstly in those celli^
where the intensity of the respiration is highest, that means
in cells of the periphery. A much higher HCN concentration will
be wanted to reduce the respiration in the central area, where
it is limited by oxygen deficiency.

When the oxygen consumption of the root tip has been
depressed by means of KCN to 50%, still oxygen and not the
quantity of haemin ferment is limiting factor in the central
tissue. This view is endorsed by the following experiment. Root
tips were put into a 51 X 10-= mol KCN solution. In this solution
the respiration sank to 50% of that of root tips in water. The
air in the vessels was replaced by pure oxygen and consequently
the respiration increased with 44% again.

Thus we may conclude that the increase of the respiration at
increasing oxygen tensions and the decrease after addition of
KCN are partly located in different areas of the root tips. Al-
though perhaps in these cases the reaction is located in cells
of different tissues, the type of the responses indicates that the
reactions have identical physiological characters: increase in

respiration results into a lower R.Q., decrease into a higher R.Q.
On p. 312 it was already discussed that these changes of the R.Q.

By the investigations by Kluyver and Hoogerheide (1934, 1936)
and Hoogerheide (1935) a relation between metabolism and redo.-c
potential of the cell has been made probable. They found e.g.
in Saccharomyces cerevisiae during the anaerobic fermentation
always an Eh of about—43 millivolts. With increasing respiration
also the redox potential rises in a definite way, so that a definite

Endorsed by these experiments we may assume that the altera-
tions in respiration stated in the cells of the root tip are accom-
panied by changes in the redox potential. We learned that the
auxin content decreased with increasing respiration and increased
with decreasing respiration. This may be also formulated: the
auxin content increases with sinking redox potential, it decreases
with rising redox potential.

The suggestion, that the auxin content of a tissue would
depend upon the redox potential has already been made by
van Overbeek (1935). This investigator found that in the coleoptiles
of the corn variety nana more auxinj was inactivated than in
coleoptiles of normal corn. Moreover the nana-coleoptiles further
proved to be richer in catalase. If the catalase content of normal
coleoptiles was increased artificially, the auxin inactivation in-
creased also in these coleoptiles. This made van Overbeek con-
clude that the difference in auxin inactivation in normal and
nana coleoptiles is to be due to a difference in „oxidation level
(rH?)quot;.

A photochemical oxidation of auxin was stated by Skoog (1935).
He found that auxin solutions were inactivated by X-rays.
Ordinary light proved to have the same effect, if a small quantity
of eosin was added to the auxin solution. Since the presence of
oxygen was wanted for the inactivation of auxin, it should be
a photo-oxidation.

In our case we thus can explain the decrease of the auxin
content at higher oxygen tensions by a rise of the redox potential
in a number of cells, by which the oxidation of auxin is accele-
rated. The increase of the auxin content in the presence of KCN
should, reversely, be ascribed to a sinking redox potential

ï) It is remarkable that Bonner and Thimann (1935) in Avena. coleoptiles
did not find any influence of HCN on the auxin content of the cells.

The mechanism of the changes in auxin content leaves room
for different possibilities:

1.nbsp;The oxidation of auxin can be thought to be a reversible
process. In that case the oxidation product should be reduced to
auxin at a sinking redox potential. No data are available in the
literature on the reversibility of the auxin oxidation. Therefore
this hypothesis is lacking any ground.

2.nbsp;The production of auxin is greater at a low redox potential.
This possibility is disproved by the resutls of the experiments

aerobic fermentation
fore also the redox potential at low oxygen pressure must have
been much lower than in the experiments with KCN. It was,
however, found that the auxin content did not markedly shift
in comparison with that of root tips in air. That means a decline
in comparison with the auxin content of tips in KCN, which
cannot be ascribed to oxidation of the auxin. It seems probable
that the process, by which auxin is produced, is checked if the
redox potential becomes too low.

3.nbsp;Finally the most probable and simpliest way to explain
the increase in auxin content with decreasing respiration is the
assumption that the oxidation of auxin is slackened at a reduced
redox potential.

The aim of the experiments reported in this chapter was to
elucidate the glucose effect. The result can be resumed as fol-
lows: a high glucose concentration reduced the respiration of the
root tips. In the mean time the R.Q. increases. It seems justified
to exegete this as a decrease of the redox potential. This reduces
the oxidative inactivation of auxin, so that the auxin content
of the root tip is increased.

The influence of phosphate buffer solutions on the respiration
and the production of auxin in the root tips.

Auxin is an acid. According to Dolk and Thimann (1931) salts
of auxin do not cause curvatures in the Averti test. Therefore
the pH of the agar blocks applied in this test never should
exceed pH 7. Bonner (1934) showed that the auxin in the Avena
coleoptile partly occurs as inactive salt. By soaking cylinders of
coleoptiles in acid buffer solutions the auxin could be liberated

These facts are important in connection with the experiments
described in the preceding chapter. It was stated that the auxin
content of root tips increased, if the respiration decreased in
favour of the fermentation. It is possible that in this shifting of
the metabolism organic acids accumulate. In the applied method
of extraction these acids perhaps might be extracted too and
finally reach the agar blocks. In that case these agar blocks could
obtain an extremely low pH by which the auxin could be
liberated from its salt unilaterally in the coleoptile. The test
plants then would show much stronger curvatures than would
agree with the auxin content of the agar blocks applied.

It was investigated whether this possible source of error
actually comes into the play. To this purpose root tips, which
had stood on glucose agar, were extracted with ether. After
evaporation of the ether the résidu was not solved in 0,2 cm»
of McIlvaine's buffer solution, but in the same quantity of
destilled water. The pH of this solution was determined colori-
metrically. This experiment was repeated for a number of times;
the pH of the extract always proved to be higher than pH 4.

This shows, that if any organic acids may come into the agar
block, their quantity must be extremely small. Moreover, the
extract was not solved in water but in a buffer solution pH
± 5,4. There is, therefore, no reason to be afraid for an
influence of organic acids on the magnitude of the curvatures
of the test plants.

§ 2. The influence of phosphate buffer solutions on the produc-
tion of auxin and on the respiration of the root tips.

Originally I intended to suspend the root tips in all experiments
on respiration in buffer solutions. This proved, however, not to
be applicable, since the phosphate buffer solutions used proved
to affect the production of auxin in the root tips. This influence
will be described below.

The method in measuring the respiration and the auxin was
completely the same as in the experiments described above. The
buffer solutions were prepared after Kolthoff (1923) from solu-
tions of mol Na2HP04 and KH2PO4. In experiments with

glucose the latter was solved in the buffer solution. The pH was
determined either colorimetrically with a Hellige comparator
Î) In the case that the concentration of the acid would be so high, that
it would overbalance the buffer capacity of the McIlvaine solution (see
p. 287).

(table 27), or potentiometrically with the hydrogen electrode
(tables 28 and 29).

In table 27 the influence of a buffer solution of pH 5,5 on the
respiration of root tips is represented. The respiration in water
equals that in buffer solution. Glucose affects the respiration in
the same way in buffer solution and in water; the consumption
of oxygen is decreased by it.

Also the next table 28 shows that the influence of glucose on
the respiration of root tips is not shifted by the presence of
phosphate.

When considering the figures in the fifth column of table 28,
however, one states that the auxin content of root tips in glucose
buffer solution does not differ from that of tips in buffer
solutions alone. Here no effect of glucose on the auxin content
can be stated!

In fact the auxin content of the root tips in table 28 is much
higher than it uses to be of root tips in water. This indicates
that the phosphate solution causes an increase of the auxin
content of the same order of magnitude as glucose does.

For that reason the auxin contents of root tips in water and
in phosphate solutions were compared; the results are given in
table 29.

TABLE 29 Respiration and airxin content of root tips in water, phosphate
buffer and 7% glucose.

The auxin content of tips in phosphate solution proves indeed
to be much higher than that of tips in water. The difference is
of the same order of magnitude as that caused by glucose.

The described influence of a phosphate solution on the auxin
content of the root tip is still completely obscure. It is possible
that the buffer solution changes the pH in the root tips and that
this change affects the auxin content. It is possible as well, that
the influence of the phosphate solution has nothing to do with
its buffering capacity. Perhaps these facts correlate with the
influence of salts, reported earlier (van Raalte, 1936). It was
stated that the amount of auxin, delivered by root tips to agar
was considerably larger, if the agar contained KCl or BaClg.
According to Thimann (1934) I used the term „osmotic actionquot;.
This author namely supposed that the effect of glucose on the
auxin production consisted in an „osmotic actionquot;: the auxin
would be extracted from the tip into the agar by osmotic force.
In a verbal discussion Prof. C. E. B. Bremekamp critisized this

explanation of the salt effect. He remarked that such an osmotic
attraction was hard to visualize. I believe indeed, that for this
phenomenon the term „osmoticquot; should preferably not be used,
until more experimental evidence has been given to such an
effect.

1.nbsp;The influence of different concentrations of glucose on the
delivery of auxin by root tips has been determined (Chapter II,
§ 1 p. 288).

2.nbsp;Glucose has no increasing effect on the delivery of auxin
by the tip of the Avena coleoptile (Chapter II, § 2 p. 289).

3.nbsp;Glucose has no increasing effect on the delivery of auxin
by the cotyledons of Raphanus (Chapter II, § 3 p. 293).

4.nbsp;The presence of auxin inactivating substances in ground
root tips was proved (Chapter II, § 5 p. 295).

5.nbsp;The effect of glucose on the delivery of auxin by the root
tip is not due to reactions in the agar or in the boundary
between agar and root tip (Chapter II, § 5 p. 296).

6.nbsp;The effect of glucose on the delivery of auxin consists in
an increase of the total amount of auxin, present in the root tip.
In the root tip, therefore, auxin can be synthetisized from other
substances (Chapter III, p. 303).

7.nbsp;The oxygen consumption of root tips is smaller in 7% or
10% solutions of glucose than in water; the respiratory quotient
and the auxin content is higher (Chapter IV, § 2 p. 309).

8.nbsp;The decrease of the oxygen consumption of root tips by
means of KCN is associated by an increase of the respiratory-
quotient and of the auxin content (Chapter IV, § 3 p. 313).

9.nbsp;The oxygen consumption of root tips is increased, the auxin
content decreased, by a high oxygen tension of the milieu
(Chapter IV, § 4 p. 315).

10.nbsp;If the oxygen tension is reduced very strongly (to 3—10%
of that of normal air) the respiratory quotient strongly increases,
the auxin content, however, remains about on the same level
as in normal air (Chapter IV, § 5 p. 318).

11.nbsp;In 1/15 mol phosphate solutions the oxygen consumption
of root tips is the same as in water; the auxin content, however,
is higher (Chapter V, § 2 p. 322).

1.nbsp;The proof, that auxin can be synthetisized in the root tip
of Vicia from other substances. The importance of this fact for
our knowledge of the growth of the root and the literature on
this subject have already been discussed in Chapter III (p. 299)
to which may be referred here.

2.nbsp;The proof of a correlation between the auxin content of
the cells and their respiratory metabolism. What is the impor-
tance of the latter correlation for our knowledge of the growth

In Chapter IV the higher auxin content, resulting from a
reduced redox potential was explained as a consequence of a
reduced oxidation of auxin. For this phenomenon, however, also
a more indirect explanation can be found. One can suppose that
the auxin content of the cells is determined by the rate of auxin
production and by the rate of auxin consumption during the
growth at the other hand (the inactivation of auxin is left out
of consideration). When the rate of growth is reduced by some
external factor and the production of auxin is not affected, or
at least to a much smaller degree, the auxin content of the cells
consequently will increase. In our experiments the rate of growth
possibly may be reduced as a consequence of a reduced oxygen
consumption or of a high salt (phosphate) concentration of the
milieu. In these cases also the auxin content of the root tips
increases. An exception must be made for the experiments in
nitrogen. This can be explained by assuming that a too strongly
reduced redox potential stops not only the auxin consumption,
but also its production, so that the auxin of the cells does not
shift any more. The reduction of the auxin content of root tips
in pure oxygen can, according to this explanation, be ascribed
to an increased rate of growth, by which the auxin consumption
would increase.

All results, reported in Chapter IV and V, can be explained
in this way. The whole explanation, however, is based on the
hypothesis, that auxin is actually consumed in the growth process
of the root cells. Our knowledge of the function of auxin in the
root is still too scanty to assume such an hypothesis. Only during
the last year data have been published, which point into the
direction of an auxin consumption by the root. It may perhaps
be useful to survey briefly the literature on this point.

of roots of Avena was inhibited by pure auxin. Nielsen (1930)
Boyen Jensen (1933) and others found an inhibiting effect of
hetero-auxm on the growth of roots. According to these results,
the ceHs of roots and cells of stems react in an opposite way oii
these hormones.

CzAJA (1935) tried to explain this opposite reaction. He
thought that his experiments allowed the conclusion that the
growth of a cell is inhibited by a bipolar supply of auxin, accele-
rated by an unipolar supply. He tried to prove that in the root
two opposite flows of auxin occur: one from the aerial parts
traveling downwards and another from the root tip traveling
upwards. These flows would meet in the growing region and
mhibit the growth rate in this zone. In a decapitated root the
latter flow would be suppressed, the cells would receive auxin
only from one side and so the growth rate would be increased.
As a proof for this conception Czaja placed decapitated roots
horizontal: they curved positively geotropically.

This theory of Czaja, however, met with serious criticism This
criticism partly attacked his experiments in which inhibition of
growth by opposite flows of auxin had to be proven (Jost and
Reisz, 1936; Lane, 1936). At the other hand the results of Czaja's
experiments with decapitated roots could not be reproduced and
proved to be inconclusive (Faber, 1936; Thimann, 1936).

Much more evidence than Czaja's theory has the conception,
that the effect of auxin is dependent on its concentration. As
early as 1925 Jost had ah-eady suggested this possibility to
explain the opposite behaviour of stem and root. In the literature
issued during the last year a number of data has been reported
indeed, which indicate that low concentrations of (hetero-) auxin

Uy^b) derive from their experiments that hetero-auxin supplied
to roots, which contain only little auxin, accelerates the growth
while the same external concentration would inhibit the growth
of roots containing much auxin of their own. Also the experi-
ments by Jost and Reisz (1936) weakly point to a growth
accelerating effect in roots of low hetero-auxin concentrations
Amlong (1936) supplied hetero-auxin unilaterally to roots of
Vicia. A concentration of 10-« mol caused a negative curvature
higher concentrations positive ones. The concentration of 10-'
mol therefore would accelerate the growth.

Geiger—Huber and Burlet (1936) cultivated under sterile
conditions isolated excised roots and studied the effect of hetero-
auxm on the growth. They stated that concentrations of 2 86 lO-i^

_2.86.10—quot;' accelerate the growth, with the maximum accelera-
tion at a concentration of 2.86.10-quot;. Hetero-auxin concentrations
of 2.86.10—'—2.86.10-5 inhibit the growth. The authors conclude
from their experiments, that auxin is indispensable for the growth
of the root. They further stated that the hetero-auxin is inactiv-
ated by the root, also in concentrations, which inhibit the growth.

From the cited literature we perhaps may conclude that auxin
is indispensable for the growth of the root. The fact, that the
concentration has to be extremely low for an acceleration of
the growth rate, indicates that the quantity of auxin consumed
by the growth process is very small. If this holds true, no con-
siderable increase in the auxin content can be expected if the
growth is checked. For this reason it seems more probable that
the increase of the auxin content, reported in Chapter IV and V,
should be ascribed to a decreased inactivation rather than to
a decreased consumption, due to an inhibition of the growth.
To discriminate between these two possibilities, however, still
more should be revealed on the function of auxin in the growth
process of the root.

This work has been carried out in the Botanical Institute of
the State-University, Utrecht; it was started under the direction
of the late Prof. Dr. F. A. F. C. Went and continued under the
direction of Prof. Dr. V. J. Koningsberger.

Amlong, H. U. 1936. Zur Frage der Wuchsstoffwirkung auf das Wurzel-
wachstum. Jahrb. f. wiss. Botanik, 83, p. 773.

Avery Jr., G. S. 1935. Differential distribution of a phytohormone in the
developing leaf of Nicotiana, and its relation to polarized growth. Bull.
Torrey Botan. Club, 62, p. 313.

Barker, J. 1933. Analytic studies in Plant Respiration IV. The Relation
of the Respiration of Potatoes to the Concentration of Sugars and
the Accumulation of a Depressant at Low Temperatures. Proc. Roy.
Soc. London, B. 112 pp. 316 and 336.

Bonner, James. 1934. The Relation of Hydrogen Ions to the Grovrth Rate
of the Avena Coleoptile. Protoplasma 21, p. 406.

Bonner, James. 1936. The Growth and Respiration of the Avena Coleoptile.
Joum. gen. Phys. 20, p. 1.

Bonner, James and Thimann, Kenneth. V. 1935. Studies on the Growth
Hormone of Plants. VII. The Fate of the Growth Substance in the
Plant and the Nature of the Growth Process. Joum. gen. Phys. 18,
p. 649.

Boysen Jensen, P. 1911. La transmission de I'irritation phototropique dans
I'Avena. Oversigt over det Kgl. Danske Vidensk. Selsk. 1911, p. 1.

Boysen Jensen, P. 1933b. Die Bedeutung des Wuchsstoffes für das Wachstum
20 p.nbsp;Krümmung der Wurzeto von Viciaquot;

Boysen Jensen P. 1936. U^er die Verteilung des Wuchsstoffes in den
Keimstengeln und Wurzeln während der phototropischen iS geo^o-
pischen Krummung. Det. Kgl. Danske Videnskabemes Selskab 13 dI

hormonale Wirkung quot;der Organspitze bei der
geotropischen Krummung. Berichte deutsch, bot. Ges. 42, p 356

Emerson R 1927. The Effect' of certain respiratory Inhibitors on th.
Respiration of Chlorella. Journ. Gen. Phys. 10, p 469 ''''''^^^

Fiedler, Herbert. 1936 Entwicklungs- und reizphysiologische Untersuchun
gen an Kulturen isolierter Wurzelspitzen. Ztschr f BotLik SO n

Geiger-Hüber Max. 1935. Ueber den Einflusz der kLSiSL des At
Twins, Tl! f, Atmungsgeschwindigkeit .oT^lrJ^^^.

Hawker, Lilian. E. 1932. Experiments on the Perception of Gravity by
Roots. New Phytologist, 31, p. 321.

Heyn, A. N. J. 1935. The chemical Nature of some Growth Hormones as
determined by the Diffusion Method. Proc. Kon. Akad. Wetensch.
Amsterdam 38, p. 1074.

Hoogerheide, J. C. 1935. Bijdrage tot de Kennis van de Reactie van Pasteur.
Thesis, Delft.

Hopkins, E. F. 1924. Relation of low Temperatures to Respiration and
Carbohydrate Changes in Potatoe Tubers. Hot. Gazette, 78, p. 311.

Hulssen, C. j. van 1936. Adendialing, Gisting en Groei. Een Onderzoek
over de Werking van Auxinen en van Biotine. Thesis, Utrecht.

iNMAN O. L. 1921. Comparative Studies on Respiration. XVI. Effects of
hypotonic and hyigt;ertonic Solutions upon Respiration. Joum. Gen.
Phys., 3. p. 533.

Jost, L. und Reisz, Elisabeth. 1936. Zur Physiologie der Wuchsstoffe.
Zeitschr. f. Botanik, 30, p. 335.

Kluyver, A. J. und Hoogerheide, J. C. 1934. Ueber die Beziehungen zwischen
den ' Stoffwechselvorgängen der Mikroorganismen und dem Oxydo-
reduktionspotential des Mediums. Biochem. Zeitschr., 272, p. 197.

Kluyver, A. J. und Hoogerheide, J. C. 1936. Beziehimgen zwischen den
Stoffwechselvorgänge von Hefen und Milchsäurebakterien und dem
Redox-Potential im Medium. III Neue Versuche mit Hefearten. Enzy-
mologia 1, p. 1.

Kögl, F., Haagen Sivirr, A. J. und Erxleben, H. 1933. Ueber ein Phytohormon
der Zellstreckung. Reindarstellung des Auxins aus menschlichem Harn.
4 Mitteilung über pflanzliche Wachstumsstoffe. Ztschr. f. physiol.
Chemie. 214, p. 241.

Kögl, f., Haagen Smit, A. J. und Erxleben, H. 1934. Ueber den Einflusz
der Àuxine auf das Wurzelwachstum und über die chemische Natur
des Auxins der Graskoleoptilen. 12. Mitteilung über pflanzliche Wuch-
stoffe. Ztschr. f. physiol. Chemie, 228, p. 104.

Kosinski,' J. 1902. Die Atmung bei Hungerzuständen und unter Einwirkung
von mechanischen und chemischen Reizmitteln bei Aspergillus niger.
Jahrb. f. wiss. Botanik, 37, p. 137.

Kübowitz, f. und Warburg, O. 1929. Atmung bei sehr kleinen Sauerstoff-
drucken. Biochem. Ztschr., 214, p. 5.

Laan P. A. van der. 1934. Der Einflusz von Aethylen auf die Wuchsstoff-
bildung bei Avena und Vicia. Recueil Trav. bot. néerl. 31, p. 691.

Lane, Roy. H. 1936. The Inhibition of Roots by Growth Hormon. Amer.
Journ. of Bot., 23, p. 532.

Larsen Poul. 1936. Ueber einen wuchsstoffinaktivierenden Stoff aus Phaseo-
lus' Keimpflanzen. Planta, 25, p. 311.

Mack, W. B. 1930. The Relation of Temperature and the partial Pressure
of Oxygen to Respiration and Growth in germinating Wheat. Plant
Physiology, 5, p. 1.nbsp;• , , .

Maige, a. et Nicolas, G. 1910. Recherches sur l'influence des solutions
sucrées de divers degrés de concentration sur la respiration, la tur-
gescence et la croissance de la cellule. Ann. des Sciences nat. Botanique,
9- ème série, 12, p. 315.

Meyerhop, o. 1925. Ueber den Einflusz des Sauerstoffes auf die alkoholische
Gärung der Hefe. Biochem. Ztschr. 162, p. 43.

Nagao, Masayüki 1936. Studies on the Growth Hormones of Plants 1 The
Production of Growth Substance in Root Tips. Science ReS tLSu
Imp. University, serie 4, Biology, 10, p. 721.nbsp;xoKonu

lierenden Stoff: Rhizopin. Jahrb. f. wiss. Botanik, 73,
OVERBEEK, J. VAN, 1933. Wuchsstoff, Lichtwachstum^reaktion Und Phoio

tropismus bei Raphanus. Recueil Trav. bot. néerl. 30, p 537
ovemeek J. van, 1935. The Growth Hormone and the Wf Tvne or

ovemeek J. van. 1936^ Growth Substance Curvatures of A:vena in Light
and Dark. Journ. Gen. Phys., 20, p 283nbsp;^

Ts''fe;men?-r^^^^^^ ^ la connaissance de la théorie
desjermentations proprements dits. C. R. de l'Acad. des Sciences Tb!

Auxm and Auxin Precursor. Journ. Gen. Phys. 20, p 311
Soding Hans. 1929. Weitere Untersuchungen über die Wuchshormonp rf».

Waeburg, O. 1926. Ueber die Wirkung von Blausäureaethylester (Aethyl-
carbamin) auf die Pasteursche Reaktion. Biochem. Ztschr. 172, p. 432.
Wabburg, O. 1928a. Ueber die katalytischen Wirkungen der lebendigen
Substanz. Berlin.

traduits par E. Aubel et L. Génévois. Paris.
Went, F. W. 1928. Wuchsstoff und Wachstum. Recueil Trav. bot. néerl., 25,
p. 1.

Went, F. W. 1932. Eine botanische Polaritätstheorie. Jahrb. f. vyiss. Botanik,
76, p. 528.

Proc. Kon. Akad. Wetensch. Amsterdam, 34, p. 875.
Wey, H. G. van der, 1932. Der Mechanismus des Wuchsstofftransportes.
Recueil Trav. bot. néerl. 29, p. 381.

Bij de hogere plant kan iedere cel, onder geschikte uitwen-
dige omstandigheden, tot auxinevorming overgaan.

De methode, waarmee Amlong meent een invloed van auxine
op de rekbaarheid van de celwand bij wortels te kunnen aan-
tonen, is hiervoor niet geschikt.

De hoge affiniteit voor zuurstof van foetale haemoglobine in
vergelijking met die van het volwassen zoogdier, staat niet in
verband met de ontwikkeling in de uterus.

Bozler bewijst niet, dat de kontraktiele elementen in de gladde
spier viskeuze eigenschappen hebben.

Het verdient aanbeveling, naast de Gymnospermae en
Angiospermae een derde hoofdafdeling „Chlamydospermaequot; te
onderscheiden, welke de geslachten Ephedra, Gnetum, en Wel-
witschia omvat.

Uit de frequentie van het pollen van Carpinus in pollen-
diagrammen, kunnen geen kwantitatieve konklusies worden ge-
trokken omtrent het aandeel van deze boom in de samenstelling
van de bossen gedurende het postglaciaal.

De verminderde virulentie van tabaksmozaiek virus na toe-
voegen van trypsine, berust op een inwerking van deze stof op
het virus zelf; niet op een inwerking op de plant.

Ten onrechte konkludeert Beale, dat de door haar in mozaiek-
zieke cellen waargenomen kristallen zouden kunnen overgaan
in het kristallijne tabaksvirus proteine.

De opvatting van Waksman over de oorzaak van de ontgin-
ningsziekte is niet juist.

	1 hour	3 hours	5 hours
agar -f 5% glucose	1,6° ± 0,2	1,5° ± 0,5	2,1° ± 0,5
plain agar	0,5° ± 0,3	3,4° ± 0.9	3,4° ± 0,6

plain agar	10,0° ± 2,0	agar 5% glucose	7,2°	±1,2
plain agar	4,2° ±1,5	agar 5% fructose	5,6°	±0,8

	XYi hours	1 hour
plain agar	5,2° ± 0,6	7,2° ± 0,7
agar 5% glucose	7,2° ± 1,0	—
agar 10% glucose	3,0° ± 0,8	—
agar 5% fructose	3,3° ± 0,3	4,7° ± 1,2
agar 10% fructose	4,1° ± 0,9	—

plain agar	3,3°	±0,9	agar 5% glucose	4,4°	±0,8
plain agar	2,7°	±0,7	agar 5% glucose	2,8°	±0,6
plain agar	2,1°	±0,7	agar 5% glucose	3,6°	±0,7

			check agar blocks
			without auxin
plain agar	5,6°	±1,4
agar 5% glucose	6,1°	±1,1
agar 5% glucose -1- 1% asparagine	4,2°	±0,5	2,5° ± 0,9
agar 5% glucose 1% peptone Poulenc	14,1°	±0,9	0,7° ± 0,7

		check agar block
		without auxin
plain agar	4,9° ±1,1
plain agar	5,9° ± 0,5
agar 5% glucose	3,2° ± 1,0
agar 5% glucose	6,3° ± 1,3
agar 5% glucose 1% peptone	7,9° ± 0,6
agar 5% glucose 1% peptone	4,8° ± 0,8	1,0° ± 0,6

		agar -f- 5%			agar 10%
plain agar	2,7° ± 0,8	ordinary glucose	11,7°	±0,8	purified glucose	11,8°	±0,9
plain agar		agar -1- 10%			agar 10%
plain agar	j 3,3° ± 0,5	Îordinary glucose	11,7°	±1,7	purified glucose	13,5°	±1,1

Glucose concentration in the agar	Amount of auxin given off
0	2,9°	±1,0
1%	2,1°	±0,4
3%	3,5°	±0,6
5%	3,4°	±0,5
10%	4,9°	±0,6
15%	1,6°	±0,4

plain agar	10% glucose agar	plain agar
no glucose		glucose previously applied
1,7° ± 0,5	5,9° ± 1,1	8,2° ± 0,9

Experiment	tips from	number of tips extracted	curvature of test plants	ratio
1. (21-ll-'35) glucose aga-		10	5,7° ± 0,8	4,1
	plain agar	50	6,9° ± 0,8	4,1
2. (10-12-'3.5)	glucose agar	15	6,4° :t 0,6
2. (10-12-'3.5)	plain agar	25	3,5° 0,6	3,0
3. (9-12-'35)	! glucose agar	30	10,8° ± 0,7
3. (9-12-'35)	plain agar	50	8,0° ± 0,7	2,3
4. (2-12-'35)	glucose agar	30	4,0° It 0,4
4. (2-12-'35)	plain agar	75	2,7° 0,4	3,7
5. (3-12-'35)	glucose agar	15	3,2° ±0,6
5. (3-12-'35)	plain agar	75	8,1° ± 0,9	2,0
6. (19-12-'35)	glucose agar	15	2,7° ± 0,5
6. (19-12-'35)	plain agar	50	1,9° ± 0,4	4,5

	set 1	set 2
delivery from 10 h. 10—11 h. 10	6,1 ± 1,2	4,0 ± 0,5
„ 11 h. 10—12 h. 10	9,7 ± 1,2	7,6 ± 1,3
„ 12 h. 10—13 h. 10	10,6 ± 1,1	13,7 ± 0,9
„ 13 h. 10-15 h. 40	10,2 ± 0,8	10,0 ± 0,8
„ 15 h. 40—17 h. 10	5,2 ± 0,8	9,1 ± 1,5

lot no.:	1	2	3	4	5
mm' oxygen during first 30 minutes	85,2	83,3	84,6	88.3	84,9
„ „ „ second 30 min.	82,5	84,2	81,8	89,6	84,2
„ „ „ third 30 minutes	74,7	74,0	75,9	80,7	76,4
total respiration in ly^ hours	242,	241,5	242,3	258,6	245,4
largest difference: 258,6 — 241,5	= 17,1 mm'		= ± 7%.
TABLE 18. Oxygen consumption of 5	lots of 20 tips. Liquid medium
2 cc. phosphate buffer, concentration			p.H. 6,8.
lot no.:	1	2	3	4	5
mm' oxygen during first 30 minutes	53,5	52,6	57,9	58,0	52,3
„ „ „ second 30 min.	55,7	55.7	56,0	65,0	57,0
„ „ „ third 30 minutes	54,1	53,2	55,7	56,0	50,5
„ „ „ fourth 30 min.	48,0	48.5	50.9	59,5	52,8
total respiration in 2 hours	211,3	210,0	220,5	238,5	212,6

	Expt. 1.	(17-4-'36)	Expt. 2.	(20-4-'36)	Expt. 3.	(22-4-'36)
mm» oxygen		5%		10%		1 7%
hours	no glucose	glucose	no glucose	glucose	no glucose	glucose
of 30 tips:	161,7	150,9	134,9	94,6	131,1	90,7
	(100%)	(93,3%)	(100%)	(70,1%)	(100%)	(72,2%)

			oxygen
Experiment	tips in	number of	consumption	amount of
Experiment	tips in	of tips	mm» hours	auxin
1. (ll-5-'36)	water 10% of glucose
		60	310,0 (100%)	7,1° (100%)
		60	192,5 (62%)	12,3° (173%)
2. a3-5-'36)
	water 10% j of glucose	! 60	343,2 (100%)	7,5° (100%)
3. (i4-5-'36)	water 10% j of glucose	j 60	191,8 (73%)	28,8°* (384%)
3. (i4-5-'36)	1 water 10% of glucose
		60	292,5 (100%)	3,4° (100%)
		60	195,9 (67%)	±7,4° * (512%)
4. (15-5-'36)	water ! 10% of glucose i
		60	311,6 (100%)	5,1° (100%)
		60	222,6 (71%)	10,2° (200%)

	mm'Os	mm'COa	P.Q (lt;a) 1,09 1,87
Expt. 1. (16-ll-'36) water 7% of glucose	hours 168,8 (100%) 131,4 (77,8%)	hours 183,3 246,2	P.Q (lt;a) 1,09 1,87
Expt. 2. (17-ll-'36) water 7% of glucose	166,0 (100%) 115,3 (69,5%)	150,3 144,7	0,91 1,26
Expt. 3. (23-ll-'36) water 10% of glucose	169,5 (100%) 85,6 (50,6%)	173,1 167,0	1,02 1,97
Expt. 4. (24-ll-'36) water 10% of glucose	185,7 (100%) 98,4 (53,0%)	217,1 184,7	1,17 1,88


■ • V- Vf / '

	concentration	number		® quot; ' CO,
	of KCN	of tips	hours	hours	R.Q.
Expl. 1.
(20-7-'36)	0	20	102,8 (100%)	—	—
	77 X 10-6 niol	20	89,2 (86,7%)	—	—
	31x10-5 mol	20	64,4 (62,1%)	—	—
	154 X 10-5 mol	20	40,6 (39,4%)	—	—
Expl. 2.
C5-10-'36gt;	0	30	158,6 (100%)	261,2	1,65
C5-10-'36gt;	51 X 10-5 niol	30	91,6 (57,8%)	326,4	3,56
Expt. 3.
(6-10-'36)	0	20	187,9 (100%)	206,6	1,10
(6-10-'36)	51 X 10-5 mol	1 20	134,3 (65,1%)	354,8	2,64

	concentration of KCN		number of tips	time m respiration vessel	m m ' Q hours	i auxin content
Expt. 1. (21-7-'36)	0 20 X 10-5	mol	60 60	2 hours 2 „	249,8 (100%) 181,7 (72,8%)	7,0° i 0,7 7,9° ± 0,6
Expt. 2. (23-7-'36)	0 30 X 10-5	mol	60 60	2 hours 2 „	300,4 (100%) 182,2 (60,0%)	4,8° ± 0,4 5,5° ± 0,2
Expt. 3. (24-7-'36)	0 25 X 10-5	mo]	60 60	2gt;i hours 2m „	315,1 (100%) 224,1 (71,0%)	6,5 ' ± 0,4 9,9° ± 0,6
Expt. 4. (30-7-'36)	0 25 X 10-5 0 51 X 10-5	mol mol	30 30 30 30	y^ hours ^ „ „ 2^4 „	168,6 (100%) 92,0 (55,0%) 153,8 (91,2%) 89,2 (53,0%)	1,1° ± 0,4 3,0° ± 0,7 1,4° ± 0,6 6,6° ± 0,9
Expt. 5. (3-8-'36)	0 25 X 10-5 51 X 10-5 102 X 10- 5 205 X 10-5	mol mol mol mol	30 30 30 30 30	254 hours 2% „ 2j4 „ 2^ „ 2j4 „	133,0 (100%) 95,7 (72.0%) 83,2 (62,5%) 83,6 (62,8%) 66,4 (50,0%)	1,1° ± 0,3 2,4° ± 0,5 4,0° ± 0,6 4,6° ± 0,5 3,3° ± 0,7
Expt. 6. (4-8-'36)	0 51 X 10-5 51 X 10-5	mol mol	60 60 30	234' hours „ 2m „	291,6 (100%) 154,4 (53,0%) 76,4 (52,4%)	3.3° ±0,3 8,1° ± 0,8 3,3° ± 0,5

liquid medium	m m ' ^ ;-
liquid medium	hours
water	85.0
water	104,6
buffer pH 5,5	101,6
buffer pH 5,5	103,9
buffer pH 5,5 7% glucose	72,6