TER VERKRIJGING VAN DEN GRAAD VAN
DOCTOR IN DE WIS- EN NATUURKUNDE
AAN DE RIJKSUNIVERSITEIT TE UTRECHT,
OP GEZAG VAN DEN RECTOR-MAGNIFICUS.
Dr. L. S. ORNSTEIN, HOOGLEERAAR IN DE
FACULTEIT DER WIS- EN NATUURKUNDE.
VOLGENS BESLUIT VAN DEN SENAAT DER
UNIVERSITEIT TE VERDEDIGEN TEGEN DE
BEDENKINGEN VAN DE FACULTEIT DER
WIS- EN NATUURKUNDE OP
MAANDAG 30 MEI 1932.
DES NAMIDDAGS TE 4 UUR

Nu het einde van mijn studietijd is aangebroken, is het
mij een behoefte dank te 2;eggen aan allen, die aan mijn
wetenschappelijke vorming hebben meegewerkt.

Hooggeleerde Van Romburgh, Nierstrasz, Kruyt
en Moll, zeer erkentelijk ben ik U voor de leiding, welke
Gij mij gegeven hebt in mijn eerste studiejaren.

Hooggeleerde Jordan, het onder Uw leiding in aan-
raking komen met de dierlijke physiologie is voor mij van
blijvende waarde geweest. Uw colleges, vooral het college
over de theoretische grondslagen van de biologie, heb ik
met groote belangstelling gevolgd.

Hooggeleerde Wes ter dijk, het verheugt mij, dat een
diepere beschouwing van het ziekteprobleem niet aan mijn
opleiding ontbroken heeft. Voor het vele dat ik op Uw
laboratorium en Uw excursies geleerd heb, dank ik U zeer.

Hooggeleerde Pulle, U wekte mijn belangstelling voor
de plantengeographie en de systematiek. Van Uw colleges
en de excursies onder Uw leiding, heb ik een zeer aan-
gename herinnering behouden.

Hooggeleerde Went, Hooggeachte Promotor, in groote
mate gaan mijn gevoelens van dank naar U uit. Gij zijt
het in de eerste plaats, die mij geleerd hebt zelfstandig te
werken. Uw diep geworteld geloof in de beteekenis van
de natuurwetenschap was mij steeds tot steun in tijden
van tegenslag. Dat Gij mij toestond dit proefschrift buiten
Uw laboratorium te bewerken, heb ik opgevat als een
groot blijk van vertrouwen. Ik ben U zeer erkentelijk voor
de warme belangstelling, die U voor mijn werk bleef
koesteren; na een bespreking met U was ik steeds met
nieuwen moed bezield.

Hooggeleerde Van Iterson, zonder Uw medewerking
zou de bewerking van dit proefschrift niet mogelijk ge-
weest zijn. Ik ben U zeer dankbaar voor de onbekrompen
wijze, waarop U mij in de gelegenheid hebt gesteld mijn

werk in Uw laboratorium uit te voeren. Voor Uw belang-
stelling en het vele, dat ik als Uw assistent van U heb
geleerd, betuig ik U mijn hartelijken dank.

Waarde Van den Honert, ik ben niet vergeten, dat Gij,
hoewel Uw tijd zeer beperkt was, nog gelegenheid wist te
vinden om mij met de moeilijkheden van Uw apparaat
bekend te maken. Hoewel ik in enkele opzichten tot een
andere opvatting ben gekomen, vermindert dit niet de
beteekenis, welke Uw werk voor mij heeft gehad.

Den Heer Kiers zeg ik dank voor de moeite aan de
vertaling van dit proefschrift gegeven.

Het personeel van het Laboratorium voor Technische
Botanie te Delft wensch ik te bedanken voor de hulp mij
bij mijn werk verleend, in het bijzonder ben ik U, Waarde
Van Kampen, veel verphcht voor Uw kundige technische
hulp. Ook het personeel van het Botanisch Laboratorium
te Utrecht wensch ik mijn dank te betuigen voor de hulp,
mij vroeger verleend.

C.nbsp;The Influence of Temperature on the
CO2 Concentration-Assimilation Curve
when Phenylurethane has been Added 540

C.nbsp;Experiments on the Influence of Light
Intensity on the Assimilation of Hormi-
dium Cultivated in Weak Artificial Light 544

D.nbsp;The Influence of Light Intensity on Assi-
milation at Different Temperatures.... 546

B.nbsp;Experiments with Low Concentrations of
KCN, Showing a Stimulating Influence 565

2.nbsp;Experiments on the Induction of Pho-
tosynthesis at Different Temperatures 595

A.nbsp;The Assimilation Reacts to the Addition
of Uncommon Substances in the Same
Way under Different Conditions____... 601

C.nbsp;The Explanation of the Parallelism of
Respiration and Photosynthesis ........ 607

E.nbsp;An Attempt to Explain the too Large
Deviations of Some Determinations on
the Influence of Temperature on Photo-
synthesis from the Average, and a Few
Remarks on Optimum Curves......... 612

In this chapter a full discussion of the literature will
not be aimed at. Very extensive surveys are a.o. to be
found in the monographs of Stiles and Spoehr; I also
refer to the short review of literature of van den Honert.
In this place only that which pertains to the problem of
this paper will be discussed.

In the year 1905 Blackman stated in his well-known
paper: „When a process is conditioned as to its rapidity
by a number of separate factors, the rate of the process
is limited by the pace of the ,,slowestquot; factor.quot; In
Blackman's opinion the relation between an external
factor and the rate of photosynthesis can be represented
by a straight curve with a sharp break.

This statement is founded on experiments of Miss
Matthaei (1904), and is further supported by experiments
of Blackman and Matthaei (1905), and of Blackman
and Smith (1911).

The accuracy of these experiments was of such a nature
that a possibiHty of other interpretation remained (criticism
of Brown and Heise (1917, 18). Experimental attacks on
the theory of limiting factors came from the part of many
investigators. Especially the paper of Harder (1921),
which was altogether devoted to this question, is of impor-
tance. He stated that in case of low „concentrationquot; of
one factor, an increase in the concentration of the other
(non-limiting) factor, will cause an acceleration of the
process. According to Harder the relationship between
photosynthesis and the external factors can be expressed
by a smooth curve.

So, the problem seemed to be settled against Blackman's
formulation, were it not that van den Honert (1928, 30)
seemed to have confirmed this theory experimentally, in
very accurate experiments on the influence of the COg
factor, made with films of the filamentous alga Hormidium
flaccidum. The relation between COg concentration and
assimilation, represented in a graph, shows a straight curve
with a sharp break, resembling the scheme proposed by
Blackman.

It must also be mentioned that Boysen Jensen
and Müller (1929) discovered a Blackman curve for the
action of light on shade plants of Marchantia polymorpha.
The number of points, determined on this curve is,
however, rather small.

The light intensity-assimilation curve of van den
Honert, on the other hand, shows a logarithmic type (as
Harder's curve), but van den Honert is of opinion
that a better agreement with Blackman's scheme should
be possible, if the light intensity on the light- and shade-
side of the assimilating cells could be equalized.

A reliable curve of the Blackman-type obtained experi-
mentally is an argument of more importance than ten
curves showing a gradual course, and constitutes therefore
a great support for the adherents of Blackman's formula.
A smooth curve may also occur (even if Blackman's
postulate were true), if the experimental technique were
inexact and not all other factors than the one studied were
kept constant (cf. also Boysen Jensen and Müller).

The fact that van den Honert did not find a
Blackman-curve when the influence of light was studied,
therefore, does not preclude the possibility that under
improved experimental conditions a Blackman-curve would
indeed appear. It is true that van den Honert worked
with a film one-cell in thickness, but objections arise
against the manner of illumination used. A solution of the

question how Homidium behaves under improved experi-
mental conditions has been sought in Chapter VI.

Van den Honert explains his COg concentration-assi-
milation curve by assuming that in case of low CO2
concentrations the diffusion of CO2 will limit the assimi-
lation, and that the direct proportionality arises from the
quick absorption of CO2 in the organism.

Van den Honert succeeded in making it quite plau-
sible that the diffusion process limits indeed, by showing
that the process is not at all sensitive to temperature, which
is in conformity with the properties of the diffusion of
CO2 in water (cf. also Bohr 1897 and 99).

Though van den Honert has shown that a curve with
a proper resemblance to the Blackman-scheme is ob-
tainable, this may be a coincidence and an exception. The
same opinion is held by van den Honert in the theore-
tical discussion of his results (v. d. H. 1930, p. 245).
The fact itself does not speak for the general validity of
Blackman's theory.

In the experiments of Warburg (1919, 20), in which
the diffusion plays no part, as he could prove by change
in temperature, a logarithmic curve was obtained.

In other experiments Warburg ascertained that narcotics
retard the assimilation. I, therefore, have asked myself how
the CO2 concentration-assimilation curve of narcotized
Hormidium would proceed, and whether it would be possible
in this way to cause the Blackman-type to disappear. It
may be possible that the limiting action of diffusion of
CO2 will be excluded by this interference. This question
will be treated in Chapter V.

B. Do the External Factors Act Directly on the Photo-
synthetic Process, or Do Internal Protoplasmic Processes
Interact? «

(1919, 20, 22, 24) have very thoroughly studied the assi-
milation of carbon dioxide, and they have arrived experi-
mentally at a theory of photosynthesis. I am not going
to consider the contents of these theories here, in which
case the opinion of other investigators should have to be
mentioned too, but only a tendency which these theories
have in common.

Photosynthesis is considered here as a reaction in an
„unalterablequot; system (test-tube), while the possibility that
this system itself (the organism) changes under the
influence of the varying factors, is ignored and not even
suggested

With high light intensity and high concentration of COg,
temperature is the limiting factor of photosythesis. It is
supposed that under these conditions the rate of the whole
process is controlled by the speed of a pure chemical part
of it (Blackman reaction). In the theories of Willstatter
and Stoll, Warburg, a.o., this chemical reaction or
reaction complex is, as a matter of fact, considered as a
stage of the photosynthetic process itself. To be sure, this
possibility may not be denied, but that it should be obvious
is not at all evident. In fact it is possible, that internal
processes determine the intensity of the assimilation, and
that the accelerated action after an increase of temperature
is merely based on the acceleration of these internal
processes (alterations of the system in which the reaction
occurs). In this case some „branch-chainquot; of the process
will be studied, and not a part of the reaction: 6 COg
6 HgO Cfi HiaOfi -I- 6 Og.

Quite another conception than the first mentioned,
which we may call „chemicalquot; (on account of the internal
regulatory processes being neglected) we find in the work

1) I do not mean the „adaptationquot; to the factors of the milieu
after a long time, which is certainly acknowledged by these inves-
tigators, but an immediate reaction of the plasma during the experiment.

of Spoehr and Mac Gee (1923), Kostytschew and his
collaborators (1926—31), Harder (1930), and Arnold
(1931). The first mentioned investigators assume a depen-
dence of the assimilation on the respiration. They observed
that after a long period of darkness, the assimilatory intensity
ran parallel to that of the respiration and to the increase
in sugar-content. From this they concluded that active
products of carbohydrate-metabolism influenced assimi-
lation. The experimental foundation of the theory is weak,
but still the assimilation is looked upon as a physiological
process being dependent on internal processes.

Kostytschew and collaborators published a series of
investigations by which it was shown that assimilation is
only slightly influenced by the prevailing external factors.
Fluctuations occurred, without there being a noticeable
external cause, so that they came to the theory (Kostyt-
schew 1931), that assimilation is dependent on internal
factors and on the after-effect of external factors („höchst
komplizierte Reizkettenquot;). The irregular assimilation they
found appears partly to be under the influence of move-
ments of the stomata, but can also appear when the
stomata are quite open.

It was even shown that under very favourable circum-
stances an evolution of CO2 may take place. Boonstra
(1930), however, could not succeed in confirming in Holland
this phenomenon, which has been observed in four different
climates in Russia.

Kostytschew was also criticized by Boysen Jensen
and Müller (1929), who observed a more regular course
of the assimilation. According to them Kostytschew's
results are probably a consequence of experimental errors.
We wonder indeed how it is possible that rather regular
curves, as obtained by Matthaei (1904), Willstätter
and Stoll (1918), Lundegârdh (1921), would have been
possible if the assimilation would always be subject to

such large fluctuations. As a result of his opinion Ko-
stytschew denies any value of experiments of short
duration, in which external factors are varied.

Harder (1930) found that the intensity of assimilation
in aquatics changes with time. He found no fluctuations
as Kostytschew a.o. did, but a gradual increase of the
assimilation in light, followed by a decrease. He explains
his results by distinguishing a number of reaction swhich
must all be of internal nature, called by him „Akti-
ierungquot;, „Gegenreaktionquot;, „Ermüdungquot; and „Anpassungquot;.

Arnold (1931), who almost at the same time arrived
at similar results with Elodea, agrees with this opinion.

Other investigators, who did not find a constant course
of the assimilation-intensity, are Ewart (1897, 98),
Pantanelli (1904), Lubimenko (1905), Willstätter
and Stoll (1918), Johansson (1923, 26), Plantefol
(1927), Montfort and Neydel (1928).

Willstätter and Stoll ascribed this fact to an accu-
mulation of products of assimilation; the explanation is
altogether „chemicalquot;.

Johansson explains the decrease in intensity by the
closing of the stomata, so his explanation does not call
real internal factors into play either.

Of great importance is, therefore, the work of Montfort
and Neydel (1928), who observed the same phenomena
with leaves of Hydrophyllacea, in which stomata are
wanting. They could show that neither accumulation of
products of photosynthesis (a decrease began after a short
illumination), nor desiccation (the leaves were in water),
nor alteration of the position of the chloroplasts (after
change in position more light gave less assimilation) caused
the decrease, so that it is obvious to account for these
facts by means of internal reactions, the more so as the
preliminary treatment also proved to have influenced the
reaction. With strong light these investigators, too, did

find evolution of COg, however, not alternating with
periods of strong assimilation, as Kostytschew found.

Very large fluctuations within the lapse of an extremely
short time were found by Maximow and Krassnos-
selsky—Maximow (1928). They do not know, whether the
cause is internal or a result of the movement of the stomata.

Beljakoff (1929), however, did not find these large
„jumpsquot;, and ascribes them to experimental errors.
Moreover he found that the stomata do not react rapidly
on light, temperature and moisture, in contrast to the
above-mentioned investigators.

Unusual assimilation-curves, which also point to a very
complicated composition of the assimilatory-process were
found by Lundegârdh and his collaborators Stocker
(1927), Walther (1927), Yoshii (1928). Their curves are
characterised by several optima and minima.

Van den Honert (1928, p. 9; 1930, p. 158), however,
severely criticizes the apparatus of Lundegârdh and
remarks that inasmuch the measurements are made imme-
diately after the illumination, the absorption of CO2 cannot
be constant in the first minutes. To this I would add that
for another reason it is undesirable to measure in the first
minutes. The opinion that the assimilation is connected
with very complicated processes, demands that the organism
is allowed some time for the creation of a state of equih-
brium. Though the truth of this assertion may be doubted,
yet the possibility must be taken into account.

In connection with this last point the work of Tsi-
Tung Li (1929) should also be mentioned, who found
an immediate considerable increase of assimilation after
altering the intensity of the hght or its composition (red-
white). Tsi-Tung Li explains this by considering the
available surface to be a limiting factor. When taken out
of the darkness into light there will be a large surface
available in the beginning, which will diminish later on.

So he explains the phenomenon by an internal limiting
factor, though not an active internal one. The case may
be more compUcated, but it clearly demonstrates the
danger of premature measurements.

In the preceding pages we have mentioned a number
of modern investigations, where there was more or less
reason to consider the assimilation connected with other
vital functions. The study of this correlation has become
the principal object of the present paper.

In this the conception of Spoehr and Mc Gee have
served as a working-hypothesis. I have always asked myself
how both assimilation and respiration behave under different
conditions. I was led to their hypothesis, not because a
direct relation between photosynthesis and respiration
seemed likely a priori, but because of the fact that, if photo-
synthesis should be connected with protoplasmic activity,
the oxygen consumption of the protoplasm could be taken
as a fair measure of this activity.

For this reason the influence of temperature on photo-
synthesis is compared with the influence of this factor
on respiration (Chapter VI). In itself the determination
of the influence of temperature on assimilation is also of
importance, because the literature shows, in addition to the
none too rehable determinations with leaves of higher
plants, only the data of Emerson (1929), who worked with
Chlorella pyrenoidea. All other determinations are more
or less fragmentary.

The influence of four different substances on respiration
and assimilation is investigated in the Chapters VIII—XL
Since photosynthesis is considered to be a chain process
by many investigators (a.o. Briggs, Warburg, van den
Honert) and since different reactions possibly control
the rate of photosynthesis under different conditions, the
influence of these chemicals is determinined both in light
of low and of high intensity.

Finally, in Chapter XII, I have tried to find an answer
to the question whether any processes precede the assi-
milation. If the assimilation is really dependent on internal
processes we may expect that the equilibrium will not be
immediately established. We know already the so-called
„photochemical inductionquot; of assimilation since the inquiry
of Warburg (1920), but it is doubtful whether we have
really to do here with a photochemical phenomenon or
with physiological reactions which must precede the
assimilation. It has been tried to determine the nature of
this „precedingquot; reaction.

In the course of the investigations the necessity became
apparent to study respiration as well. Its magnitude during
photosynthesis should be established. Fluctuations in the
intensity of respiration might be the cause of fluctuations
in our analytical results. The study of respiration becomes
a necessity when a causal relation between this form of
metabolism and assimilation is claimed (Spoehr and
Mc Gee). This problem will be treated in Chapter III.

With the exception of a few, which are mentioned in
Chapter V, and which were made with the apparatus of
van den Honert (1928, 30), the experiments were made
m a new constructed, very simple, but at the same time
very accurate apparatus, according to the manometrical
method.

The advantage of van den Honert's method, the expe-
rimentation with a film of Hormidium of one cell in
thickness was maintained in it. The assimilation of this
film is, according to van den Honert very constant during
a long time. Kostytschew's objection against the
determination of the influence of external factors, which
would be impossible on account of the inconstant course,
does not hold here.

In order to check my results with those of van den
Honert (analyses of COg concentration) the assimilatory
quotient was determined (Chapter IV). This was all the
more necessary, because van den Honert had found
deviations of this quotient from unity in a few determi-
nations.

A strain of the filamentous alga Hormidium flaccidum
served as experimental object. The same strain was used
by van den Honert in his investigations. He determined
the strain as H. flaccidum, which species is subdivided in
in different sub-species in Pascher's Flora. This strain
forms, when cultivated in a suitable way, a film of only a
single cell layer on the nutrient solution and is, in this
state, very suitable for the investigation of photosythesis.
I obtained another strain, the sub-species Hormidium
nitens from the collection of Dr. E. G. Pringsheim at
Prague. This latter strain was free from bacteria, which
is not the case with the former. With the latter strain it
appeared more difficult to obtain „filmsquot;. Moreover the
film is less close, which causes loss of material when
removed to other liquid culture media (which was necessary
in the experiments of Ch. VIII, IX, XI). For these reasons
I mostly used van den Honert's strain; the quantity of
bacteria is, moreover, small in a well growing culture. For
the sake of comparison other organisms (Stichococcus
bacillaris, Oocystis spec.) were tested occasionally.

The cultivation succeeds very well in Erlenmeyer
flasks (contents 150 cm^), which, closed by a cotton-wool

stopper, are placed in a cool room before a north-window;
the conditions of cultivation were therefore not constant!
In summer the light must be dimmed a litde with white
paper. Very good cultures were obtained at a temperature
of 15°—18° C. A lower temperature is not directly injurious
but the growth becomes very slow. At higher temperatures
Pringsheim's strain soon forms less fine films; van den
Honert's strain may keep well, provided large quantities
are constantly transferred. If this is not done, the alga
gets overgrown with bacteria. At least once a week the
cultures must be transferred; in winter this may be done
less frequently. In the summer of 1930 I grew the algae
in a Hear son's cool biological incubator, in which the
temperature was kept at 15°—16°. The flasks are filled
with 20—30 cm3 nutrient solution of the following compo-
sition, given by van den Honert; water distilled from
glass into glass, containing:

In this medium also other algae, such as Stichococcus
bacillaris, S. minor and Oocystis spec, appeared to grow
well.

Besides on the windowsill, I also cultivated the algae
with a constant source of light. For this purpose Erlen-
meyer flasks (contents 1 litre) were used, shut at the
top by a rubber stopper containing two passages. With the
aid of an aquarium-pump indoorair continually bubbles
through the liquid. The flasks were placed round a 75 Watt
milkglass Philips lamp burning day and night and sur-
rounded by a watercooler. After three or four weeks the
liquid becomes darkgreen. Hormidium is in this case
usually unicellular and forms no film.

A few experiments (those mentioned in Chapter V) were
made with the apparatus of van den Honert. According
to his method a current pf air containing CO2 is led at a
great speed over a film of Hormidium. The used air is
not directly removed, but is again led over the alga by
means of a small pump. The supply of air containing CO^
and the removal of air samples for the analysis takes place
very slowly in comparison to the speed of the circulating
current of air. After some time a state of equilibrium is
attained and the tension of CO« of the circulating air
becomes rather constant. With the aid of a gasanalysis
apparatus of Krogh the tension of CO2 is determined
accurately within 0.001 %. The same had been done before
with the supplied air. From the quantity of air removed
per unit of time and the difference in COo contents be-
tween supplied and removed air the assimilation of carbon
dioxide may be calculated.

We refrain here from an extensive description. The reader
can find it in the publications ofvandenHonert (1928,30).

The method is very accurate. An objection, however, is
the long duration of the experiment; a disturbance in the
intricate apparatus may easily set in. The greatest diffi-
culty is the handling of the Krogh apparatus. This apparatus
suffers greatly from leakages, which often occur, especially
(as in my case), when it has already been used before, and
the stopcocks become older and less true. Moreover the
making of a great number of analyses is very trying work.
Another drawback is that the apparatus soon gets dirty; the
regular cleaning takes up much time. A last objection of
the method is, in my opinion, that it may be called accurate,
but it is not supple. The course of the process is difficult
to follow. When the air is removed, one never knows if
the equilibrium is already attained.

For all these reasons I have given up working with this
apparatus, and constructed a new one, according to the
manometrical method, which fitted better to the purpose
of the investigation.

For the present a few more remarks about the apparatus
of van den Honert. Some minor improvements have
been introduced. The glass tube, numbered 23 in fig. 8
of van den Honert (1928, 130) has been replaced by
an unbreakable brass one. The valves, shown in fig. 9 are
constructed more simply. Instead of string and a melted
wax and resin mixture, a piece of rubber tube is used. This
is put onto tube 1, after which tube 5 is passed round it,
so that a watertight closure is effected.

L Description of the apparatus. This apparatus (cf. fig. 1)
is an application of the microrespirometer of Krogh for
the assimilation of carbon dioxide. Two vessels (a and h)
are connected by means of a manometer (c), one of these
vessels (b) serves as control, and contains only air and
water. It served only to enable us to compare the varying
pressure of the air with the constant pressure in the other
vessel, and to preclude the effect of slight fluctuations in
temperature.

The principle of the method is to allow the alga to
assimilate or respire in a space of constant COo tension,
so that only the oxygen tension changes. This change is
indicated by the rise or fall of the liquid in the manometer.
By adding mercury out of a calibrated capillary tube (d),
the original pressure can be restored and the quantity of
Og evolved or absorbed, which is equal to the volume of
mercury displaced can be read. The constant tension of
CO2 is reached with a so-called buffer-solution of Nag CO3
and Na HCO3. So the method, contrary to that of van den
Honert, is based on the measurement of alterations in the

tension of oxygen and not on the determination of the
carbon dioxide concentration.

The assimilation- and the control-vessel are very shallow
and made of nickel plated brass (cf. fig. 1 A). The inner
diameter is 106 millimeter, and the outer 124, the height
only 4 mm. They are covered with a glass plate (ƒ) 3 mm
thick (a cover of a glass box). Between vessel and plate
is a greased rubber ring (e). The plate is clasped on the

vessel with the aid of a brass ring (/i) and five clamping-
screws (;). In order to prevent cracking of the glass there
is also a rubber ring (g) between ring and glass.

On the bottom of the vessels we put 20 cubic centimetres
of the buffer mixture. There is an opening in it leading
through a twice bent glass tube to the manometer. To
prevent the liquid from entering this opening a short tube
4 mm long (k) has been soldered on it. Besides the assimil-
ation* vessel contains an opening (/) communicating with
the calibrated capillary tube (d). This connection is esta-

blished by the twice bent tube (m), in which there is a
small bulge (n), which serves as a mercury reservoir. The
capillary tube (d) is a pipette of one cubic centimetre.

cm® and can be read with
an accuracy of 0.001 cm.^
The displacement of the
mercury is effected by rai-
sing and lowering a level-
glass with mercury (o), which
glass communicates with the
capillary tube.

The bent tube, which con-
nects vessel and calibrated
tube, contains also a three-
way stopcock (p) connected
with a side-tube (q). This
tube (q) serves to drive gas
through it, but was only
used in the experiments for the determination of the
assimilatory-quotient and is further unnecessary.

The vessels are put on tripods (r), so that the water of
the thermostat basin, in which the apparatus is put may
freely circulate around them.

The manometer (c) is a U-shaped capillary tube, 0.5 mm
wide, open at the top, and has a capillary connection (5)
with the vessels. The opening at the top can be closed
with rubber tubing and a clamping-screw (f).

As a manometer liquid paraffin oil coloured red with
Sudan III is used. Behind the tube is a scale division on
millimeter-paper.

algal film is fixed on the inside of the glass cover (ƒ) in the
following way: A plate altogether free from grease is laid
in a somewhat larger glass basin. This basin is filled with
nutrient solution, so that the plate is a few millimeters
below the surface of the liquid. On the water floats a
paraffined paper ring (diam. ± 8 cm), within which a qua
ntity of algae is transferred. The basin is put on the
window-sill under a glass cover. After two or three days a
round homogeneous film has formed itself, which is suitable
for the experiment. The liquid is sucked away with a pipette,
the film is pushed towards the middle of the plate, where
it stays as a humid layer. The last liquid is removed by
means of filter-paper and the paper ring taken away. The
plate is placed upside down on the assimilation-vessel,
after which this is shut. The film should not be too
humid, as drops would form making contact with the
buffer-mixture, neither may it be too dry lest the film
should dry up. After some practice this is easily perfor-
med. The vessel must also be placed stricdy horizontal
with the aid of a water-level in order to prevent the
liquid from sinking to one side.

When unicellular algae serve as object, the cell-suspension
is first centrifuged for some time; the clear liquid is poured
off and the remaining thick suspension is removed to a
clean glass plate with a brush.

3. The regulation of temperature. The contents of the
waterbasin, in which the apparatus stands, must be kept
at a very constant temperature, as the method is highly
sensitive to fluctuations in temperature, on account of the
great volume of gas of the vessels. A toluol thermostate is
practically not sufficient. I preferred to regulate the tempe-
rature by hand, as a reading had to take place every five
minutes, and I had an opportunity to control the tempe-
rature at that moment. Under the water-basin there is a

micro-burner (u) for heating purposes, while, in the basin,
bent glass tubes (y), through which tapwater or ice-water
streams, effect the cooling. By regulating the flame and
the speed of the stream of cooling-water the temperature
of the basin can be controlled very accurately. Slight fluc-
tuations were compensated by keeping at hand little basins
of warm and cold water or pieces of ice. The temperature
is read from a thermometer with a scale division to 0.1°.
The capacity of the basin is 70 litres. This regulation of
temperature, which may seem to be primitive, allows an
accuracy up to about 0.01° without any trouble, as within
a short time a great skill is obtained. In the basin is a stirring-
apparatus (w) driven by an electro-motor.

4.nbsp;The regulation of CO^-concentration. In most cases
a solution of the following composition was used as a
COa-buffer:

Such a solution is at 16° balanced with air containing
0.7 % CO2, at 23° with 1.1 %, which was determined by
means of the COg generator of van den Honert and the
gas analysis apparatus of Krogh. With this concentration
CO2 is no more limiting; alteration of the COg concentration
from 0.2% to 1.0% did not give noticeable alteration of
the CO2 assimilation under the conditions of my experi-
ments, as appeared from preliminary experiments.

5.nbsp;Illumination. The light source was an ordinary 150
Watt Philips metal filament lamp (x), when experiments
were made with the hght-factor in maximum. At about 20°
an alteration in the distance of the glowing filament of the
lamp from 15 to 12 centimeters has no result at all. As a
rule the distance was 13 or 14 cm. The lamp hangs halfway
in the water; so between lamp and algae there is nothing
else than a layer of water from 8 to 9 cm thickness.

When the influence of the light-factor was examined,
a different arrangement was used. As a lightsource a
Leybold projector lantern was used, in which, instead
of carbon-points a Philips 1000 W. cinema-lamp was
placed. The cone of light is vertically thrown on the assi-
milation-vessel with the aid of a 45° slanting mirror, while
care should be taken that the axis of the cone falls on the
centre of the vessel or close to it, so that the whole algal
film is equally illuminated. In order to secure a uniform
dimming of all wave-lengths the light of the lantern is
weakened by means of so-called neutral light-filters
(Plotnikov). These filters are made of fine mesh brass
wire supplied to us by the Newark Wire Cloth Company,
Newark, U.S.A. Other screens sold by the firm of
F. Köhler, Leipzic, proved to be useless for my purposes.

The screens were coppered. Then they were hung on a copper
wire in a 5 % solution of NaOH, to which 1 % potassiumpersulphate
powder was added. After about 5 minutes the surface was dull black
from the CuO formed. The screens were washed in water, dried
with a soft piece of cloth and clasped between metal rings. The
diameter of the screens is 10 cm. The following gauges were used:

The quantity of light which these screens allow to pass
was determined by means of a Moll-thermo-element
connected with a Moll-galvanometer and illuminating the
element alternatively with or without a filter put before
it. The relation of the galvanometer deflections gave the

percentage of light transmitted (with the filters used it
varied between 32 and 61 %).

. By combining two filters a still lower intensity of light
can be obtained. Here great attention should be paid that
the screens are parallel to one another, and that their
wires do not run parallel, but form a certain angle
(Plotnikov). If this precaution is not taken, moire-
phenomena may be obtained and consequendy a very
irregular illumination. It appeared for this reason that the
screens could be placed in one place only, viz. between
the two sets of lenses of the Ley bold projector lantern,
in close proximity of the front lens. Images or shadows
of the screens appeared when the screen was mounted
in any other place.

A plane parallel cuvet filled with water is placed between
the two lenses, 10 cm in diameter measuring.

The vessels are submerged in the water basin to a depth
of 6 cm below the surface (in the experiments with the
Leybold lantern).

The distance of the top of the light cone, which falls
between the two lenses, to the alga is ± 95 cm.

The voltage fluctuated during my experiments between
218 and 225 V.; very often it was much more constant.
The inconstant voltage is without any importance, when
the work is done with a strong intensity of light; in case
of weak intensity a correction must be made (cf. the experi-
ments on the influence of light intensity, Ch. VI).

When measuring the respiration all light was shut off
by means of an opaque round plate covering the glass plate.

After closing the vessels, filling the water-basin, turning
on the screw clamp above the manometer, the experiment
can begin. We must wait some time till the equilibrium
has been obtained. Any air-bubbles which there might

be under the vessels are removed with a bent pipette.
It takes rather a long time before equilibrium has become
established, at least 30 minutes. This is probably caused
by the changes of temperature and by the changes of
CO2- and water-vapour tension. The absorption of light
heats the vessel a litde. The heating effect is about Ve^
calculated from the expansion of the gas, when the lamp
is at 13 cm distance.

Now the manometer is put at 0 and the level of the
mercury in the calibrated tube is read. As a rule both mani-
pulations are repeated every five minutes. We wait till a
row of from 5 to 7 practically equal values have been read;
the evolution of O2 is apparently constant now, and the
assimilation per hour can be calculated. In case of low
intensity of assimilation or when measuring respiration,
the experiment lasts longer in order to decrease the chance
of a faulty reading.

The quantity measured does not precisely correspond
to the volume of gas evolved, as part of the oxygen dissolves
in the buffer-liquid, and nitrogen escapes from it. Moreover,
the newly formed oxygen mixes with escaping carbon
dioxide and water vapour. The increase of the oxygen
tension is not quite proportional to the formation of oxygen,
because the total volume of gas becomes larger. This may
practically be neglected, since the oxygen tension increases
only by a few percents during the experiment. The dissolved
quantity may therefore be taken as proportional to the
volume formed. Since the absolute value of assimilatory
CO2 is of secondary importance to us, a correction is not
necessary. The same may be said of the carbon dioxide
and the water vapour. Consequently the measured amount
of assimilation is somewhat too small, the measured amount
of respiration too large.

The above remarks about the corrections do not pertain,
when the investigations are made at various temperatures.

The vessel, in which the first experiments were made (table 6),
contained 37 cm® of gas and 104 cm» of liquid. When 370 mm»
of gas had been formed, the tension of the oxygen had increased
by 1 %, that of the nitrogen decreased by 1 %. Now oxygen dissolves
in the buffer-mixture, and a little nitrogen escapes. (We consider the
solubility of the gases in buffer-mixture to be equal to the solubility
of gases in pure water). The solubility of oxygen is greater, conse-
quently more gas dissolves than is given off. The higher the tempe-
rature the smaller this difference will be. At 10° the solubility of the
two gases is 0.03802 and 0.01857, the difference 0.01945; at 20°t
0.03102 and 0.01542, the difference 0.01560. The second difference
is 0.00385 smaller than the first, i.e. when the volume of gas increases
by 1 %, per cubic centimetre of liquid 0.0385 mm® less gas dissolves
at 20° than at 10°. The total liquid contains, consequently,104 x 0.0385

tively, the assimilation at 20° has been determined too high, and
this amount must be deduced.

CO2 and water vapour mix with this newly formed volume of
oxygen. The tension of both gases is greater at the highest tempe-
rature. At 16°C the tension of CO2 appeared to be 0.7 %, at 23°:
1.1 %. The difference in tension between 10° and 20° is 0.5 %;
roughly estimated. The tension of water vapour is 9.1 mm and
17.4 mm at both temperatures; a difference of 8.3 mm or in per cent
8 3

deduced from the values found at 20°, so in total 1.1 0.5 1.1 ==
2.7 %. The Qio of photosynthesis found (2.30, cf table 7) will be
corrected 2.30 — 0.06 = 2.24.

In case of the respiration this correction has to applied in opposite
direction. The same amount must be added to the value found. But
these determinations (table 9; and those of assimilation in May 1931,
table 8) were made in smaller vessels, containing 20 cm® of liquid
and 32 cm® of gas. The correction for the solubility of the gases is,
therefore, about five times smaller, or 0.2 %. The correction for the
CO2 tension was 0.5 %, roughly estimated. In some experiments
this tension was zero. Therefore, we put this correction at 0.3 %
on an average. Then the total correction becomes 0.2 0.3 1.1 =

1.6 %. The Qio of respiration found (2.06, cf table 10) will be corrected
2.06 0.03- 2.09.

Another correction to be made, when the influence of
temperature is studied, is the one for the expansion of the
gas in case of a rise of temperature. The volume is reduced
to the lowest temperature of the experimental series, by-

According to van den Honert a correction must be
made for the increase of the material, a „growth-correctionquot;,
which should be about 1 % per hour at 20°. I found a small
increase indeed, which however, as van den Honert
himself already stated was rather subject to variations.
It appeared e.g. to be a little stronger in weak light, and
so it seems that it is not exclusively founded on growth.
Moreover I was faced by the difficulty, how to execute
the correction at other temperatures. Since the duration
of my experiments was on the whole shorter as those of
van den Honert, it seemed warranted to drop the whole
correction, which is moreover of little importance.

Operating with the apparatus as described above is very
simple and not at all tiring for the investigator. An experi-
ment hardly ever fails on account of technical difficulties.
The accuracy of the method is, moreover, very great, the
determination possible with an accuracy within a few
cubic millimeters. The course of the process can always
be followed, so that the course of an experiment may be
changed if it seems advisable. Moreover, incidental pheno-
mena can readily be recognized as such. Besides the
advantage of the method of van den Honert, the feature
of the thin film of algae, has been preserved.

Changes in the solution of the cell-layer may be easily
effected in the following simple manner: the glass plate
is again placed in a basin, a paraffined paper ring is laid
round the film and liquid is added (containing the factor
studied e.g. a narcotic). The films are so tight that this
manipulation can take place without loss of material.

A. Introduction, On good grounds it has always been
supposed that respiration goes on during assimilation. It
appears that different processes as e.g. protoplasmic current
keep going on. These processes cannot be imagined without
respiration.

Whenever the amount of the assimilation is determined,
the amount of the respiration must be added to the value
measured, in order to evaluate the exact amount of photo-
synthesis. For want of a better standard the respiration
is here determined in the dark before or after the deter-
mination of assimilation, under the supposition that the
difference of respiratory activity in the light and in the
dark would be slight.

Yet there are other possibihties, viz. that the respiration
would have been changed by the supply of products of
photosynthesis or by direct action of light on the protoplasm,
or by an increase in temperature. The first possibilities
were mentioned by Meijer and Deleano (1911, 1913)
and called the „ergastogeneousquot; action, and the „plasmo-
geneousquot; action of the light. The third may especially
be of interest in experiments made with leaves. Blackman
and Miss Matthaei (1905) found in sunlight temperature
increases from 4° to 12°, which large differences, however,
were later contradicted by Willstatter and Stoll (1918),
Lundegârdh (1924) and Johansson (1926), who could

find but small differences of a few degrees at most, when
the light passed water, and the transpiration of the leaves
was not hindered.

That an „ergastogeneousquot; influence of light exists,
is fairly certain. Borodin ascertained (1876, 81) that
the respiration is stronger after an exposure to light than
before. Aereboe (1893) confirmed this. These investigators
put this down to an increase of products of photosynthesis.
An increase after an exposure, following a long period
of darkness, was further observed by Matthaei (1904),
Meijer and Deleano (1911, 13), Kniep (1914), Pan-
tanelli (1915), Harder (1915), Plaetzer (1917), War-
burg and Negelein (1922), Spoehr and Mac Gee
(1923). The last-mentioned authors were able to show
that the increase of sugar content runs parallel to this.
Warburg proved that an addition of glucose stimulates
the respiration, which fact could be confirmed by Emerson
(1927), Genevois (1927) and by myself (Chapter X).

Meyer and Deleano accentuated that these experiments
do not prove that at the same time light could not exercise
a plasmogeneous influence. Experiments made in air free
from CO«, which had to solve this point, gave doubtful
results. On the other hand these investigators did succeed
in finding a plasmogeneous action of light on the respiration
of another nature. By means of alternate exposures (in the
day time light, at night dark) a rhythmic respiration was
obtained. The rhythm continued when the leaf remained
in the dark afterwards.

Miss Plaetzer noticed that these rhythmic phenomena
did not appear in Cladophora. With Spirogyra on the
other hand, she found a rise at night, probably in connection
with nuclear division.

The experiments on the influence of illumination on the
respiration of non-chlorophyllaceous parts of plants give little
reason to believe in an important „plasmogeneousquot; influence

of light in case of green plants. The numerous investigations,
yielded few results, sometimes an increase, but mostly a
slight decrease or no change was found. A slight increase
may, moreover, have been caused by a rise of temperature
(Lowschin, 1908). A discussion of the literature on this-
subject is to be found with de Boer (1928).

A better insight into this problem is the more desirable
if one considers that sometimes the assimilation is relatively
slight and only litde larger than the respiration. Here the
ignorance about the exact amount of respiration might
be a dangerous source of errors.

B. Experiments. In the first place the intensity of
respiration was determined before and after a period of
illumination in which assimilation took place. The result

		\ \
		\
.5 1 -tt	Light Assimilallon	\ \ \ \ \

-'-		1 1 1 ''quot;»»

graphically repre-
sented in fig. 2,
shows clearly that
the respiration has
increased markedly
(from 13.5 mm® to
27 per hour), to
return to the old
value after some
Fig. 2. Rate of respiration determined before time. Several ex-
and after a period of assimilation. The full periments yielded

More examples of this phenomenon are the experiments
made in June 1931; see table 22 in Chapter XIL The
respiration is very high after a long period of illumination
by daylight; it keeps decreasing during the determinations,
of respiration in darkness.

The result of the experiment, mentioned above, indicates
that the respiration during the exposure to light had been
at least twice as great as in the period before it. It stands
to reason that this experiment does not show, whether
this increase is of an ergastogeneous or plasmo geneous

In order to esta-
blish this the res-
piration was exa-
mined in a COg-
free vessel over a
solution of 1 %
Fig. 3. Increase of respiration in a COa-free KOH, instead of
vessel after exposure to light. Full lines respi- over a buffer-mix-

lines respiration in the light diminished with qq ^hg followine
theassimilationofrespiratoryC02(exp.values);
the broken line represents the probable course of experiment made
respiration before, during, and after exposure, with Oocystis spec..
In spite of the lack of CO2 (no assimilation!) graphically repre-
there is a remarkable increase of respiration sented in fig 3
during and after the exposure.nbsp;gj^^^ ^^ impression

of the influence of exposure (lamp 150 W. at 13 cm). An
increase of respiration may no doubt be concluded.

This experiment shows furthermore the remarkable
feature that the respiration could also be determined during
the exposure. The amount that is determined now is of
course not the pure respiration, for part of the carbon
dioxide formed by the respiration (x) will be assimilated
once more, after which O2 evolves. The amount deter-
mined is, therefore, the respiration — x, the real respiration
in this experiment amounted consequently to 51 -f x per
hour. This is already higher than the rate of respiration

before in the dark (39). For the rest this experiment shows the
phenomenon much less beautifully than I always found it
with Hormidium. I chose this experiment as an example, as
the increase after the exposure is shown here very markedly.

A very beautiful example is given by e.g. experiment 5,
made-with a Hormidium strain of Pringsheim grown in
constant light (fig. 4).

During the exposure the respiration rises from 39.5 to
87 X, so the increase is at least more than 100 % here.
For the sake of control, the alteration in volume was also
measured, after the equilibrium of temperature being
re-established in the dark. This determination expresses
the respiration during the time of exposure followed by
10 minutes of darkness; this value, too, is very high
(80.5 y). The rise after exposure is small, but visible.
Again the experiment was repeated, first in light of lower
intensity, then in the same. In the light of low intensity
the rise is much smaller, in strong light it was almost as
large as before.

The rise also takes place with the algae, cultivated on the
windowsill; both with Pringsheim's and van den
Honert's strain. A positive result was also obtained with
a culture in constant light of Stichococcus bacillaris;
increase from 40 to 48 -f x.

These experiments distinctly show that the respiration
during the illumination (assimilation) may be much greater
than one would expect from a determination after or before
the measuring of the assimilation. When we can make it
clear, that the increase is not due to a rise of temperature,
then the plasmogeneous action of light has been proved.
In the first place, it be once more mentioned here, it can
be ascertained from the expansion of the gas, that the
assimilation-vessel becomes ± warmer. The general
heating can consequently be neglected. Also the film,
attached to a thin glass plate, can hardly be warmer.

There may exist an unequal temperature in the different
parts of the cells, but the probability of this supposition
is not very great. The increase of respiration was sometimes
twice or three times as much, or more. As we shall see in
Chapter VII the Qjo of respiration at higher temperatures
is a litde lower than 2. In order to explain such an increase

we should have to accept an average rise of temperature
of 10° to 20° at the least. Now I noticed that owing to a
short stay of 10 minutes in the dark at 41° the assimilation
had afterwards considerably decreased at lower temperature.
In the face of this, an almost constant assimilation at 34°,
as we found in Chapter VII, would hardly be possible, if
the alga had a temperature of from 44° to 54° on the average.

A second argument against a strong rise of temperature
we meet in the hght intensity-assimilation curve (Chapter VI)
In strong hght this curve runs parallel to the abscissa.
Yet the assimilation in strong light is very sensitive to
temperature (Chapter VII). It may be objected that the
light intensity-assimilation curve in itself is possibly an
optimum curve and that the fall may accidentally have
been counterbalanced by the action of the temperature.
This does not seem to be very likely, and so this parallel
course proves (when this subtle counterargument will be
relinquished), that no heating (worth mentioning) takes
place in case of an increase of light intensity.

In the third place the influence of red light on the
respiration has been examined in experiment 9, where a
material increase of respiration from 12 to 33 x was
met with. The Wratten filter 71 A^ was put on the vessel;
this filter allowed about 64 % of the energy and only
radiation gt; 600 m [jl to pass (determined with thermopile).
Yet the respiration was only 15.5 x', and, therefore,
as x' will be about equal to x (because COg concentration
is the limiting factor), much less than in white Hght. This
seems to disprove the influence of temperature.

In the last place the after-effect of hght on respiration,
which is distinctly ascertainable even one hour after the
exposure (fig. 3 and 4), seems to disprove a rise of tempe-
rature.

By these considerations the plasmogeneous influence of
light on the examined algae is made very plausible.

The objection may be made that the respiration is
measured in air, free from COg, and the rise is wholly or
partly due to the lack of the COg factor For this reason
two experiments mentioned in Chapter VI (exp. 29 and 30)
are of great importance. In a vessel containing COg there
appeared to be no assimilation in strong light, probably
because the assimilation of the cells (cultivated in weak
light) was hindered by very strong light. During the exposure
the respiration proved to be 2—4 times as great as in the dark.

I have also tried to ascertain whether light may cause
an increase of respiration in white plants, because an influence
of temperature will be smaller here, though the literature
gave me little hope. Yet in experiment 10, a suspension
of Saccharomyces Vordermannii appeared to increase the
respiration from 152 to 183. White leaves of Opiismenus
from the hothouse, stuck on the glass with strips of paper,
gave the following results:

It is not certain that these leaves do not contain any
chlorophyll. In that case the result can at most be unfa-

Noack (1925) stated an increase of O2 absorption from leaves
illuminated in absence of CO2 after 84 hours, caused by photo-
oxidative processes. Simultaneously the leaves grew yellow. These
processes probably play no part in my experiments, as the cells
proved to be quite normal after the experiment, and were able to
assimilate normally.

When we closely consider the figures of the increase
of respiration in glucose and fructose (Chapter X, table 20),
it strikes us that this increase is only rather slight,
usually no more than 50 %, often much less. So it is no
exaggeration to say that the plasmogeneous action of light
exceeds the influence of the ergastogeneous action by far.

The knowledge that there is a plasmogeneous action of
light is, besides being important in itself, also significant
for us in connection with the inquiry about the assimilation
of CO2. In what follows we shall many times have to
deal with the question, whether light (or other external
factors) controls the intensity of assimilation directly as a
factor in the photosynthetic process itself, or whether the
action is indirect and is caused by the intermediary of the
protoplasm. It goes without saying that the latter conception
becomes more likely, now that we have ascertained a very
strong influence of light on protoplasm.

The fact that the respiration has increased during the
assimilation of COg is of practical importance, especially
for the method of determining the assimilation. A minimum
value has only been ascertained; an exact quantitative
determination will very likely be among the impossibilities.
When the rate of assimilation is very high the respiration
may be neglected, without a serious objection. Matters
change, however, when the rate of assimilation is low.
Especially in strong light the error will be very large, and
may give rise to altogether wrong conclusions, so that
this knowledge warns us at least to be cautious when we
interpret the results. In the literature it is easy to find
examples which are, for this reason, of doubtful value.
One example I will mention.

Lundegârdh (1924) has found very interesting opti-
mum curves. He found e.g. an optimum curve when

studying the influence of light intensity in case of low
CO2 concentration. In these experiments the assimilation
was rather small. The decrease of assimilation in strong
light may now well be explained by a large increase of
respiration. I do not mean to say that Lundegârdh
cannot be right, but I will only point out how cautious
we must be in the explanation of such a case.

Notwithstanding the fact that there is an increase of
respiration in light, yet in my experiments I have stuck
to the oldfashioned way of correcting the respiration. We
had perhaps better multiply the values found by 1.5 or 2.
But how does the respiration behave during an exposure
with other intensities of light? We saw that, in strong
light, there is a large increase, but how is the quantitative
relation? I always determined the respiration after an
exposure, except in the experiments in Chapter V, when
I did not yet know this influence.

One remark should be made. The results recorded in
the tables 15—23 of van den Honert and in table 2 of
myself (experiments made with the van den Honert
apparatus) seem at first to be litde in conformity with
those described above. A lower concentration of COo is
continually found in light than in darkness. The difference
between the method of van den Honert and my own
method can account for this discrepancy. In the apparatus
of van den Honert a molecule CO2 formed by respir-
ation, and afterwards evolved, is repeatedly offered to
the cells and will have a great chance of being assimilated
again; in other words: x will be very great. In my apparatus,
as used in the above experiments, it will directly be bound
by the alkali. The unknown x in my results will possibly
be not very great. Moreover the light intensity in the
experiments of van den Honert is considerably lower,
and consequently the increase of respiration rather slight
(cf. fig. 4).

By assimilatory quotient I mean, with Stiles (1925),
the quotient of the volume Og evolved and the absorbed
CO2, in contrast with Willstatter and Stoll (1918) who
define the notion reversely. Stiles is quite right, when
he observes that the reverse of a quotient, which is historic-
ally always used in this form, makes no sense.

The knowledge of this quotient was for me of importance
for two reasons. In the first place because van den Honert
(1928, 30) contrary to Maquenne and Demoussy
(1913), Willstatter and Stoll (1918), and Johansson
(1926), who had always found that the quotient approxi-
mates to unity, had noticed a deviating quotient of 1.1
with Hormidium. The second reason is that I would
investigate to what extent my method, in which the assi-
milation of carbon dioxide is stated by measuring of the
O2 evolved, is in conformity with the method of van den
Honert, based on COg determinations.

In order to determine the quotient the assimilation
vessel is thoroughly cleaned from CO2 buffer. In the vessel
there is only air containing carbon dioxide and the algal
film. The CO2 concentration is increased by expired
air blown, after deep breathing, through the tube q (Fig. 1)
which has been inserted only for this purpose in the
apparatus. Then follows an exposure to light. If the volumes
of gas evolved and absorbed should be equal, the
manometer liquid will have to remain at rest. If, on the
other hand CO2 absorption or O2 evolution predominates,
a pressure difference will arise. When this difference
has been determined, the vessel is opened and CO«
buffer is added, after which the assimilation can be mea-
sured, i.e. the development of Og. We now add to this

value the difference stated before and find the COg absorp-
tion. In this way the quotient has been determined four
times, cf. table 1. The respiratory quotient, which is
perhaps very deviating from unity is not taken into account,
which is admissible as the respiration is slight in propor-
tion to the assimilation.

The quotient found deviates only little from unity.
Van den Honert found 1.09 and 1.13, amounts that
are not very unlikely. Yet there seems to me to be no
reason for the opinion of van den Honert, that there
should be a connection between the high assimilatory quotient
and the synthesis of oil as first product of assimilation.
Moreover, I never succeeded in demonstrating fat in the
Hormidium cell either with osmic acid or Sudan III.
Pascher (1914) who describes the formation of reserve
material in detail, tells nothing about such an oil
synthesis.

The second conclusion that may be derived from these
experiments, is that it is immaterial, whether the assimi-
lation of COg is determined by measuring the COg-absorp-
tion or the Oo-evolution.

In order to determine the quotient the assimilation
vessel is thoroughly cleaned from COg buffer. In the vessel
there is only air containing carbon dioxide and the algal
film. The CO2 concentration is increased by expired
air blown, after deep breathing, through the tube q (Fig. 1)
which has been inserted only for this purpose in the
apparatus. Then follows an exposure to light. If the volumes
of gas evolved and absorbed should be equal, the
manometer liquid will have to remain at rest. If, on the
other hand CO2 absorption or Oo evolution predominates,
a pressure difference will arise. When this difference
has been determined, the vessel is opened and COo
buffer is added, after which the assimilation can be mea-
sured, i.e. the development of Oo. We now add to this

value the difference stated before and find the CO2 absorp-
tion. In this way the quotient has been determined four
times, cf. table 1. The respiratory quotient, which is
perhaps very deviating from unity is not taken into account,
which is admissible as the respiration is slight in propor-
tion to the assimilation.

The quotient found deviates only litde from unity.
Van den Honert found 1.09 and 1.13, amounts that
are not very unlikely. Yet there seems to me to be no
reason for the opinion of van den Honert, that there
should be a connection between the high assimilatory quotient
and the synthesis of oil as first product of assimilation.
Moreover, I never succeeded in demonstrating fat in the
Hormidium cell either with osmic acid or Sudan III.
Pascher (1914) who describes the formation of reserve
material in detail, tells nothing about such an oil
synthesis.

The second conclusion that may be derived from these
experiments, is that it is immaterial, whether the assimi-
lation of CO2 is determined by measuring the COo-absorp-
tion or the 0lt;,-evolution.

Van den Honert (1928, 30) investigated the influence
of CO2 concentration on assimilation. His results deviated
remarkably from those of other investigators. When the
CO2 concentration was low the assimilation rose directly
proportionally to the tension of CO2, when it was high it
was altogether independent of it. Between the two regions

there was a small transition. The curve of van den Honert
therefore approximates the Blackman-scheme.

I have repeated this experiment with the pure Hormidium
strain of E. G. Pringsheim. All corrections were carried

The experiments of this chapter were all made with the
van den Honert apparatus.

out as by van den Honert. The constant rate of assi-
milation at high CO2 concentration has been put at 100.
The illumination was effected by 6 lamps as used by
van den Honert in a row, with a reflector of white
paper behind. The latter was absent in van den Honert's
experiments; the intensity of light was therefore somewhat
higher.

Figure 5 gives the graphical representation of the
experimental result. The curve perfectly agrees with the
one found by van den Honert.

Van den Honert has been able to make it probable
that the direct proportionality in low CO2 concentration
arises here, because a diffusion process controls the whole
photosynthetic process, and that in case of high COg
concentration a chemical process is limiting. In the latter
case a rise of temperature gives a considerable acceleration,
in the former it remains without any influence. This is,
as van den Honert explained, in conformity with the
properties of the (purely physical) CO, diffusion, taking
into account the solubility of the gas at different tempe-
ratures.

According to van den Honert the sharp break in the
curve is caused by a rapid fixation of the COo by the
organism.

The curve found with Hormidium deviates altogether
from those, determined by other investigators, e.g. from
Warburg's curve, representing the relation between COo
concentration and assimilation in Chlorella. This may be
caused by the fact that, in the experiments of Warburg,
the diffusion of CO« was no limiting factor. At least
he found a great influence of temperature in low CO2
concentrations.

Finally it may be observed here, that it also appeared
from the preliminary experiments with the manometric
method that in case of high COo concentration (0.1—1.0 %)

the assimilation is quite independent of the concentration.
No optimum curve has been found, as has been observed
by Lundegârdh (1921, 24) for higher plants.

B. The Influence of Carbon Dioxide Concentration when
the Assimilation has been Retarded by Narcotics,

It is a well-known fact that the assimilation may be
depressed by the addition of narcotics. We may ask whether,
by means of a suchlike retardation, we cannot eliminate
the interference of the diffusion-process, and in this way
measure the influence of COg concentration on the assi-
milatory process itself.

For this purpose I chose a concentration of phenylure-
thane, which according to Warburg retards the assimi-
lation: 2.0—2.4 X lOquot;'^ mol. Moreover I made some
experiments with antipyrin, examined by Ewart (1896)
and Jacobi (1899) for its retarding action; I experimented
with a weak concentration, viz. ± 0.001 mol.

The influence of COg concentration on photosynthesis after
the addition of a narcotic.

Exp. 21, 24-5-30. Pringsheim's strain. Antipyrin conc.
0.001 mol.
-33.5
188
149
122
110
200
-40
85

Table 2 shows the results of these experiments. It strikes
us that the second observation in experiment 19 has
yielded a rather low value. This is in accordance with the
further experiments made with phenylurethane (Chapter IX),
where it has been shown that the retardation is stronger
at the outset, and diminishes in course of time. The rate
of assimilation becomes rather constant some hours after
the addition of the narcotic. All other observations fall
in this constant region. The rate of assimilation in high
concentration of COg is again arbitrarily put as 100. Here
we must, however, not forget that this 100 cannot be
compared with the 100 of normal assimilation, as in higher
CO2 concentration the assimilation is retarded as well as
in low concentration.

When we represent the result of exp. 19 graphically (fig. 6),
a curve is obtained which deviates very considerably from the
CO2 concentration-assimilation curve found before. The
„Blackman-typequot; has disappeared, the transition is very
gradual. The rate of assimilation increases in case of high
CO2 concentrations where normally no increase takes place.

The curve very strongly calls to mind the curve of
Warburg, represented in fig. 7. In order to make com-

parison easier the concentration of COo is also given in
0.001 %. It seems not unlikely that the same part of the
process has been studied in both cases, possibly the
chemical binding of the COg.

Van den Honert is of opinion that a curve as I obtained in
fig. 6 (represented in his publications in fig. 17), represents the
quantity of the COj-assimilatory agent (probably COa-chlorophyll)
at different COg concentrations, and at the same time the rate of
the limiting process. It is true that he speaks only of the photochemical
part of the process, but from what follows it appears that this is
presumably a slight error, and that the limiting process is meant,
which may be chemical as well as photochemical. He thinks that the
determination of this curve will enable us to find the sequence of
the different parts of the chain process. If, he argues, a lowering
of the CO2 cortcentration should cause a retardation, when light
were the limiting factor, then CO2 would be a 'factor reacting in
the photochemical process, whereas, should this lowering be of in-
fluence, when temperature were limiting, CO^ would be a factor
in the chemical process.

It will be seen below that the reaction is very sensitive to tempe-
rature. A lowering of the CO« tension acts, therefore, retarding,
when a chemical process is limiting. When we apply van den
Honert's argument to my result, CO2 would be a factor in the
chemical process and this consequently would precede the photo-
chemical one. Now, this would refute the theory of Willstatter
and Stoll and agree with a theory as formulated by Warburg.

In my opinion this conclusion goes too far. It is certain that the
chemical reaction is retarded by a lowering of the CO2 concentration.
But from this it does not follow that CO2 itself takes part in this
reaction. Let us imagine that a photochemical conversion of CO2
precedes. This reaction is not limiting, (in the case considered here)
goes „fasterquot; than the other processes, and the product of this
reaction will be dependent on the quantity of the CO2 only (it being
indifferent for my considerations if this CO2 is free or bound to
another substance, for instance to chlorophyll). The concentration
of the latter is dependent on the C02-suppIy, so that the same will,
therefore, hold good for the quantity of substance that has been
converted photochemically. In this way the chemical reaction could
consequently be dependent on the CO2 concentration, even if COj
itself does not react in this chemical reaction.

For that reason the problem of the sequence of the processes
must as yet in my opinion remain unsolved.

I will also point out that it is not necessary that the chemical
reaction, which comes to the foreground here, is the same as the
chemical reaction that is limiting in a normal organism when tempe-
rature is the limiting factor. Different parts of the chemical reaction
complex may be unequally sensitive to these chemicals.

C. The Influence of Temperature on the CO2 Concentration-Assimil-
ation Curve when Phenylurethane has been Added.

In order to answer the question whether the reaction which controls
the process after the addition of narcotics is sensitive to temperature,
I have determined this CO2 concentration-assimilation curve at two
temperatures. Now that we know the general trend of the curve
the fixing of a few points suffices.

The results of both experiments have been recorded in table 3.
In exp. 23 the temperatures 19.2° and 22.8° have been compared
and in exp. 24 the temperatures 19.2° and 22.3°.

The few figures point to a strong increase at higher temperature.
In later experiments (Chapter IX) it has been shown that the retar-
dation, strong at the beginning, becomes less in course of time. Owing
to this the values at the lower temperature have been found a little
too low. Yet the third determination in exp. 23 falls already Sy,
hours after the beginning of the narcosis, the much higher 7th value
must, therefore, for the greater part be explained by the rise in
temperature, as the retardation is fairly well constant after 5 hours
(cf. exp. 71 and 72).

The influence of the COg concentration after the addition of phenyl-
urethane at different temperatures.
Exp. 23, 14-4-1930. Van den Honert's strain. Phenylurethane-
concentration 2.2 x lOquot;^ mol.

Exp. 24, 14-5-1930. Van den Honert's strain. Phenylurethane-
concentration 2.2 X 10quot;^ mol.

Already many investigators have examined the influence
of light intensity on COo assimilation. This relation, repre-
sented graphically, usually yielded a curve of logarithmic
type. Higher land plants and aquatics are unsuitable for
this work as the light intensity will rapidly diminish in
the leaf. Such experiments do not prove that, if the
illumination had been ideal, no curve agreeing with the
Blackman-scheme, would have appeared. Boy sen
Jensen and Müller (1929) lately drew the attention
once more to this fact. This objection also perhaps pertains

to Harder's experiments (1921) with Fontinalis. The
same criticism was made by van den Honert (1928, 30)
against the experiments of Warburg (1919), in which
thin cell-suspensions were employed, weakening the light
only 10—20 %, but in which some cells would, however,
be much less illuminated. The objection is perhaps not a
serious one as the average diminution of the light is only
0 to about 15 %. Moreover the cells that receive less light,
will be relatively small in number and exercise litde
influence on the total result. Van den Honert has
avoided this objection by using an algal film of one cell
thickness where only the unequal illumination of light-
and shade-side remains. Essentially the curve of van den
Honert does not deviate from that of other investigators.
However I thought it desirable to reinvestigate the problem,
as an objection may be raised against the way in which
van den Honert illuminated the algae.

He placed 1—6 incandescent lamps in a row. Using the same
arrangement I determined the distribution of light intensity in the
assimilation vessel of the apparatus of van den Honert (with
the aid of a thermopile). Taking 10 for the light intensity in the
middle of the vessel, and placing at first one lamp before the middle
of that vessel, the intensity at 8 cm distance from the middle
appears to be only 5. This is somewhat more unfavourable than in
the experiments of van den Honert, in which, only a central
part of ± 11 cm in length is used. The distribution of light in the
vessel shows to be unequal. Now when we put 6 lamps in a row
the intensity in the middle of the vessel is about 40, and at 8 cm
distance from the centre only 30.

Equal distribution of the light intensity is attained by
throwing the centre of the light cone of a Leybold
projector lantern on the algal film. The variation of light
intensity is obtained by placing wire cloth screens in the

bundle of light. The light intensity of the undimmed
projection lamp is arbitrarily put as 1000. As a very strong
light I used an ordinary 150 W. Philipslamp; the intensity
of it at a distance of 14 cm agrees with about 2300 (1000 is
± 3000 Lux, roughly determined with a photometer).
The slight fluctuations in the voltage were measured with
a voltmeter. According to Hoist (1920) a correction of
3.5 % for the light intensity must be made for each per
cent variation of voltage.

B. Experiments on the Influence of Light Intensity on the
Assimilation of Hormidium Cultivated in Daylight.

We find the result of exp. 27 graphically represented
in fig. 8. Evidendy a curve of a logarithmic type has been
obtained, notwithstanding the improved arrangement of

The influence of light intensity on photosynthesis.
Exp. 25, 10-3-31. Van den Honert's strain. Temp. 22.00°.

the experiment. The shape of the curve corresponds to
those of Warburg (1919), Harder (1921), Bose (1924)
a.o. An approximation of the Blackman-scheme is appa-
rently out of the question.

C. Experiments on the Influence of Light Intensity on the
Assimilation of Hormidium Cultivated in Weak Artificial
Light.

The way of cultivation is of great influence on the shape
of the curve. Experiment 28 has been made with a cell-

suspension, cultivated near a lamp of low intensity of light,
burning continually. The distinguishing feature of this
culture appeared to be that the assimilation optimum was
reached at a much lower light intensity (611), which appears
very distinctly from fig. 9. The alga has adapted itself to
weak hght, it has become a „shade plantquot;. The curve shows
a weak „optimum typequot;. It is however not impossible
that this is caused by an increase of respiration in the
light of high intensity (cf. Chapter III).

Two remarkable results, which also demonstrate the
influence of the preliminary treatment, were obtained with
an algal culture, cultivated in the same way as mentioned
above, a young cell-suspension, 17 days old. In light of
the 150 W. lamp at 15 cm distance not the slightest assi-
milation appeared to take place; on the contrary the On
absorption was greatly enhanced. This experiment was
repeated with the same result on the same day.

On the next day the experiment was repeated with
another cell-suspension and with a similar result. I verified
the apparatus with an algal film from the windowsill, which
proved to assimilate in a normal way. What was the cause?

I supposed that the high intensity of Hght had been in-
jurious to the cells, and an assimilation optimum would
exist at very low light intensity. In order to settle this point
exp. 30 was continued and a weaker light applied (lamp
at 40 cm distance). It was remarkable that the great Og
absorption (—47) had totally disappeared and had passed
into a very slight evolution of Og (-}- 8). So the supposition
proved to be correct.

This experiment is moreover a remarkable example of
an increase of respiration in hght, demonstrated in
Chapter III. The absorption of oxygen had become at least
from 2 to 4 times as great in the light.

I intended to determine the entire hght-curve of this
most interesting phenomenon. When, however, I wanted
to repeat the experiment a month later, I could not get
another result than that of figure 9, in spite of several
attempts with old and young cultures. Without any doubt
this diminishes the value of the discovery, and an expla-
nation must be given with some restriction, though a
mistake in the determinations is to be considered as out of
the question.

The phenomenon calls to our mind the evolution of
COg so often found by Kostytschew and other Russian
investigators, with favourable illumination, granting that
the evolution or absorption of Oo is determined in my
experiments.

The above instance remained exceptional. In all other
experiments I found a strong assimilation which continued
to exist for hours at a stretch without any variation worth
mentioning.

Harder (1921) investigated the behaviour of the assi-
milation in case of a simultaneous change of the factors

light and carbon dioxide. He found that an increase of
CO2 also has got of influence when the light intensity
is low and vice versa. A similar result was obtained by
Lundegârdh (1921, 22).

We make a similar experiment for the factors light and
temperature, when we deter mine the light intensity-
curve at two different temperatures. The result, which
we find in the graphical representation in fig. 10, is that
in case of low light intensity, the influence of temperature
is only slight. Another experiment gave a similar result.

It is certain that the curve never makes a much larger
angle with the abscissa, when temperature is increased, as
Harder had observed by varying the factors light and COg.
In this respect there is a greater agreement with the
opinion of Blackman.

From both experiments it does not appear convincingly,
whether there is still a small increase, or that it is totally
absent. The question

-1

E /

5 / .

..... 1 1

Uffhtmienstfy
-,-1-1--1

pear from other ex-
periments (Chapter
VIII and Chapter X),
that a stimulation of
the assimilation in low
light intensity is very
well possible. Possibly
the rate of assimilation
is not in the first place

internal factors may also control the rate of photosynthesis,
also when light is the external limiting factor. Seeing that

temperature will influence these internal protoplasmic pro-
cesses, it is of importance to look for a possiWe influence
of temperature in case of weak light.

Van den Honert has already made statements about
this question. He found, however, a somewhat lower
assimilation at higher temperatures.

Van den Honert thought he could prove by this, that the Qk,
of the photochemical part of photosynthesis is ± 1 (Warburg is
of the same opinion), in my opinion, however on insufficient grounds.
If the velocity of the process should really be determined by the velocity
of a photochemical reaction (let us suppose this to be the case), yet
the sensitivy to temperature could not exercise any influence. At
both temperatures an equal quantity of light energy is given, and
in case all the absorbed light is photochemically at work, the coeffi-
cient of temperature found cannot be anything else but the
quotient of the quantities of light supplied in both cases. In our
instance this quotient will be 1, because the light intensities are
equal.

Photochemistry is repeatedly faced by the same difficulty. Q,o
must always be ascertained when the factor light is in maximum.
It is then often an accompanying chemical process of which the
Qio is determined, but this difficulty cannot be overcome and the
problem remains unsolved as yet.

As we stated before, it will appear in the following chapters that
even when light intensity is the limiting factor, the intensity of
assimilation may be varied through changes in the internal factors.

The supposition that the photochemical stage of photosynthesis
controls the entire process exclusively, is therefore unwarranted.

Table 5 shows the results of a series of experiments, in
which the problem is treated more accurately.

The voltage fluctuated a litde. Therefore, the assimi-
lation velocity is reduced to equal light intensity. The
rate of assimilation is supposed to be proportional to the
light intensity.

Experiment 33 seems to show a distinct increase at higher
temperature. However, results of the experiments 34 and
35 are not so evident. But for a difference between the
first and the second observation the assimilation is distinctly

constant. It rather seems that a kind of adaptation to the
weak light takes place, than that temperature is of influence.

A similar little jump is also to be seen in experiment 36,
where I experimented under constant temperature.
Experiment 33 is distinguished by a repetition of this
jump. Adaptation and temperature are working here in
the same direction.

At all events the influence of temperature at low light
intensity is small. Yet the fact that adaptation and tempe-
rature together cause a rather considerable effect, proves
that the assimilation is not exclusively dependent on the
external factor light, but that the extent to which the light
energy is used, is also under the influence of internal
factors.

The influence of the factor temperature on the assimi-
lation, when neither light nor COg are limiting, has also
been repeatedly investigated. The result is often expressed
in the coefficient Q^,. Some American investigators as
Crozier, Navez, Emerson (their works are especially
to be found in The Journal of General Physiology)
violently oppose the use of this coefficient, which they
wish to replace by the temperature-characteristic yc, the
energy of activation. It is true that Q^q is not a constant;
^og Qio is inversely proportionate to the product of the
absolute temperatures. This seems no objection to me,
as in biology only a rather limited range of temperature is
studied, and a constant Qio, therefore, fairly well cor-
responds with a constant pi. The conclusion they try to get
from the study of these coefficients, viz. whether one or
more processes control the rate of the total process
studied, is therefore equally possible whether the results
are expressed in Qjo or in [ji. So there is no objection to

use Qio, which remains a convenient symbol. A conversion
in [X is moreover readily performed.

A danger of working with Q^ is that the temperature
interval of 10° is' rather great, and it does not state
deviations, e. g. optima and minima. For these reason
Lundegârdh (1924) uses the coefficient Qj. Only an
investigator who experiments with an utterly refined
method, has a right to measure by this standard. Otherwise
he runs the risk of determining apparent variations
occasioned by small experimental errors. For instance,
the Qi 1.08 changes with an error of 5 % already in
1.13 or 1.03, which seems to be a very great difference.
This criticism applies to Lundegârdh himself; with
detached leaves it is impossible to experiment accurately.
Besides, L. measures already after a preliminary heating
of 5 minutes. The question remains, whether the assimilation
has already become constant.

The values Lundegârdh finds for Qjo with cucum-
ber, potato and tomato leaves are from 5—15° : 5.0,
5.6 and 6.4; 20—30° : 1.6, 2.1 and 1.6; so Q^ falls con-
siderably.

Blackman and Matthaei (1905), and Blackman
and Smith found temperature coefficients between 2.0
and 2.5.

Miss van Amstel (1916) found a lower Q^ with
Elodea, in which case presumably the diffusion of carbon
dioxide acted disturbingly, as she supposed herself.

A lower Qi„ of 1.5 was also found by Willstätter and
Stoll (1918) with green leaves, while yellow leaves gave
a still lower Qm.

Warburg (1919) found with Chlorella a high coefficient
between 5° and 10° of 4.5—5. It fell regularly at higher
temperatures.

The determinations of Yabusoe (1924) suffer from
too great an interval of temperature. Emerson (1929)
could prove that the direct proportionality between tem-
perature and assimilation, as claimed by Yabusoe, is a
result of this large interval and is therefore only imaginary.
In reality the curve has a sigmoid shape.

The experiments of Emerson in this matter are the
most important, as he determined complete temperature
curves with an accurate method (of Warburg) with a
lower plant (Chlorella), in which case the other factors
may easily be brought in maximum. Emerson's result
is in principle in accordance with that of Warburg, the
increase of the assimilation diminishes with the rise of
temperature, in other words [x,■ in which Emerson
expresses his result, is not constant and falls gradually.

Finally van den Honert (1928, 30) investigated the
influence of temperature with Hormidium flaccidum. He
found from 12° to 20° a Qio of 1.87.

Ecological investigations about the influence of tempe-
rature under natural circumstances cannot of course be
put on the same level with investigations where all factors
are in maximum and will not be discussed here.

It has been my intention to study this temperature
influence on a larger range with Hormidium.

Further I felt interested in the influence of temperature
on respiration. It will appear later on (Ch. VIII—XI)
that there are reasons to assume a connection between
assimilation of carbon dioxide and respiration.

On the one hand the possibility exists that in case
temperature is „limiting factorquot;, in reality a part of the
assimilatory chain process is investigated, viz. a dark
chemical part of it (an opinion held by many investigators).

On the other hand the possibility must not be excluded,
that, while the influence of temperature on assimilation

is studied, yet nothing else is done than to determine the
influence on the total life-intensity of the organism as a
whole, namely when the vitality of the organism would
primarily determine the intensity of assimilation. The
intensity of respiration would in that case be the direct
criterion of this influence of temperature on the organism,
the intensity of assimilation on the other hand only an
indirect result. Temperature would in this case act as an
indirect factor only, viz. by means of accelerating proto-
plasmic processes, other than photosynthesis sensu stricto.
This acceleration of the protoplasmic processes might
then be the cause of the acceleration of photosynthesis.

It is quite imaginable that protoplasm produces an
essential substance for the photosynthetic process, for
instance an enzyme, and that it is the rate of this process
that controls the total rate of assimilation, but there are
other possibilities as well (cf. Spoehr and Mac Gee).

The conception given above would at once be refuted
if the influence of temperature on assimilation and respi-
ration should prove to be quite different, but it would be
supported, though not proved, when both processes were
influenced consistently. For these reasons an inquiry
about the influence of temperature on respiration became
necessary, in addition to that on assimilation.

A simultaneous investigation of the influence of tempe-
rature on assimilation and respiration has been carried
out by Lundegârdh (1924). In his results nothing can
be discovered of a parallelism between the two processes.
The determination of the temperature coefficient of respi-
ration is, however, rather difficult in case of detached
leaves. Respiration decreases in the dark, whereas, according
to Meyer and Deleano (1911, 13), cutting causes a
temporary rising, and a sudden increase of temperature
would temporarily give too high a respiration as well.

of temperature desirable. With each measurement of the
assimilation a correction for the respiration must be made.
Virtually a determination of respiration should be made
after each measurement of assimilation. If the Q^o of the
respiration is known one determination is sufficient.
Moreover an unavoidable error is made, because it has
been shown in Chapter III that respiration strongly increases
in light. This correction would, however, be quite super-
fluous if the Qio of both processes should prove to be
equal. Anticipating my result; I may say already now
that this is, approximately, the case. This conveys, therefore,
a great simplification. It is sufficient to determine the
influence of temperature on the apparent assimilation,
which will be of the same order of magnitude as the
influence on the real assimilation.

My method deviates from those of some other investi-
gators. While Warburg, Lundegârdh, Emerson a.o.
start their measurement, as soon as the temperature
equilibrium has set in, I waited till the assimilation
appeared to be constant, in other words, until the establish-
ment of a physiological equilibrium.

According to Blackman the determination should have
to be made in an infinitely short time. If the assimilation
could be measured in this way, the temperature curve
at injurious temperature, too, should be quite normal,
according to Blackman's conception, and would not
show any optimum-type. The injurious action of the high
temperatures could be eliminated in the results in this
way. The supposition underlying this would be right,
if indeed we studied a purely chemical reaction. If, how-
ever, Harder (1930) and Kostytschew (1931) are right
and „höchst komplizierte Reizkettenquot; are connected with
the assimilation, we may not at all expect that the final
rate of assimilation would immediately set in. The
alterations in the protoplasm, which would determine the

final rate of assimilation, would precede the establishment
of the equilibrium. In order not to neglect this possibility,
it is better always to check the constancy of the reaction.

The range of temperature investigated lies between
4° and 34°. Above 30° it is already difficult to observe a
constant assimilation. It may be that secondary factors,
as desiccation of the algal film are also responsible here.

The 150 W. lamp at 13 cm distance was always used
for illumination. My results are to be found in table 6.

In a few series of experiments the variations are rather
great, much greater at any rate than might be expected
with this accurate method. A possible explanation of these
variations will follow later (Chapter XIII § E).

The influence of temperature on COg assimilation (in November).
Van den Honert's strain.

I have represented this experimental series graphically
and drawn a probable curve between the determined
points. In this way the assimilation could be read off at 20°.
In order to compare all experimental series with one other,
the assimilation at 20° was put as 100. The rate of assi-
milation at 20° is a physiological measure for the quantity
of material used. This measure was also taken by van
den Honert. All values must consequently be multiplied
by a conversion-factor (4th column of the table). The
values calculated have been recorded in fig. 11. It appears
that in spite of the individual variations a smooth curve
can freely be drawn between the points. The increase of
the rate of assimilation with the temperature is, consequently,
very regular. Pronounced optima, as found by Lundegârdh,
are altogether wanting.

Qio can be calculated from the curve. These values
have been combined in table 7. In the last column
the determined Qio has been corrected in the way as
discussed in the second chapter. At low temperatures Qy,
is largest. Apart from this it is much more uniform than
Emerson found with Chlorella. Above 10° it is fairly
well constant. Since log Q^ is inversely proportional to
the product of the absolute temperatures (log Qjq

of activation), the characteristic (jl is also nearly constant.
\i calculated from this formula, usually put in the form:

Qio being constant shows that the total chemical reaction
complex is possibly determined by one part of the process.

These experiments were all made in the month of
November 1930. A few experiments performed in May
1931 gave similar results, cf. table 8.

The influence of temperature on COg assimilation in May.
Van den Honert's strain,
Exp. 44, 16-5-31.

It is of importance to know this, because it appears,
that, in spite of the different properties which the way of
cultivation can create (Chapters VI, VIII), the tempe-
rature coefficient of the reaction remains unchanged.

The experiments mentioned here have all been made
with the Hormidium strain of van den Honert. A
similar result was obtained with Pringsheim's strain.

It is remarkable that my results give a higher Qio than
van den Honert found with the same alga, though the
difference is not very large. He found from 12—20° 1.87,
whereas I find 2.15. Deviating values were sometimes
stated, so that this want of uniformity is apparently
inherent in the material used.

The measuring of Qio of the respiration presents some
difficulties. A falling respiration will be found when algae
are taken from the window and placed in darkness. To
prevent this the culture was placed in the dark on the
preceding evening.

Good results were also obtained by the experiments
33—35 in Chapter VI, in which periods of darkness
alternated with weak illumination.

In table 9 the results of the experiments are combined.
Only with one experiment (50) the results deviate and a
high Qio was^found at a rather high temperature. In the
other experiments the determined value was reduced

again to a respiration of 100 at 20°. I thought it better to
exclude from this the deviating result of exp. 50, which
was possibly caused by too high heating. The values
calculated are graphically represented in fig. 12.

The individual deviations are also very large here. The
respiration is rather small, and the experimental error for
that reason larger. Yet the curve can be pretty accurately
determined. The great resemblance with the temperature

curve of the assimilation is very striking. Qk, has been
determined from the curve, the values are to be found in
table 10.

Qio is but a litde lower than the coefficient of the assimi-
lation. This agreement is of course no proof that the
assimilation proceeds under the control of other processes,
connected with the respiration, but seeing that we shall

observe other points of agreement between the two processes
later on, it is certainly important that the temperature
coefficient does not speak against the idea of an indirect
action of temperature on the assimilation.

It is interesting that with Hormidium the respiratory Qio
is even a little more constant than the Qio of assimilation.
This may point to the fact that the whole respiratory
reaction complex at all temperatures is possibly controlled
by one part of the process.

Temperature-coefficient of respiration, compared with
the coefficient of assimilation.

Warburg investigated the influence of prussic acid
on assimilation. He stated that ptissic acid has, in a weak
concentration, a retarding influence on assimilation, when
temperature is the limiting external factor, that this
retardation, however, does not appear when light is the
limiting factor. From these'facts Warburg concluded
that photosynthesis is at least composed of two stages,
one of which, the Blackman reaction (the chemical
part of the assimilation), would be sensitive to H C N

only, and that this process would be brought about by
a heavy metal catalysis. Another fact found by Warburg
is that assimilation, in case of increasing concentration of
the cyanide, cannot be retarded beyond a certain limit.
This limit is the compensation point of carbon dioxide
assimilation and respiration. Warburg is of opinion that
a high prussic acid concentration completely retards the
spHtting off of the Og from the COg, but not of the Og
from the respiratory products. The photochemical mechanism
remains, therefore, unimpaired, but only the dark reaction
in which an active derivate of COg is produced, is inhibited.

Warburg could not find a similar action on respiration.
On the contrary, respiration appeared stimulated by con-
centrations where the assimilation became greatiy retarded.
Retardation does not set in until in a concentration of
Vio mol. (Warburg 1921).

Another investigator who made assimilation experiments
with prussic acid was Emerson (1929). He found that
Chlorella, grown in a nutrient solution deficient in Fe,
are poor in chlorophyll, and more sensitive to H C N than
normal cells. Therefore Emerson is of opinion that
chlorophyll not only takes part in the photochemical
reaction, but also in the chemical one. I would, however,
observe that the chlorophyll content of cells grown in a
solution deficient in Fe, is certainly the only visible
point of difference, but that it is . also to be expected that
it is the Fe containing enzyme that will have been
diminished by this treatment.

I studied the influence of potassium cyanide on respiration
and assimilation with different light intensities again for
Hormidium (van den Honert's strain).

B. Experiments with Low Concentrations of K CN, Showing
a Stimulating Influence,

54. This concentration will probably soon diminish by
evaporation. Table 26 gives the result, which appears even
more clearly from fig. 13. It shows that the assimilation
is increased by the addition of K C N. It is important that
this does not occur only, when temperature is the limiting
factor, but that the whole light intensity-assimilation
curve rises to a higher level. In strong light the increase
is 27 %, in weak hght 18 %.

different light intensities.
Exp. 54, 26-3-31. Van den Honert's strain. Temp. 18.00°.

In the following experiments, recorded in table 12, a
similar stimulation, both of assimilation and respiration
is found.

Experiments 55 and 60 were carried out in the same
way as exp. 54. The evaporation of prussic acid was prevented

in the other experiments by adding K C N in an equal
concentration to the buffer mixture. In some experiments
the increase is rather considerable. Both in experiment

Fig. 13. Stimulating influence of the addition of a low concentration
of KCN on photosynthesis in Hormidium. The assimilation is in-
creased in low and in high intensity of light; increase in respiration 33%.

The influence of low KCN concentrations, not added to the buffersolution.
Exp. 55, 30-3-31. Temp. 19.00°. KCN mol.

55 and in 57 the assimilation rises with the same amount
in strong and weak light. In exp. 57 an initial strong increase
seems gradually to become a little smaller. The fact,
found in experiment 54, viz. that the respiration behaves
like the assimilation, is generally confirmed. That the
respiration cannot be determined with great precision
must not be overlooked, however.

It appears in experiment 58, contrary to what was found
in exp. 54, that the concentration Vioooo mol« already retards
the assimilation. After the lapse of a few hours, however,
a recovery sets in and the retardation passes into an
acceleration. Later on we shall repeatedly meet with this
phenomenon. Also the respiration recovers a little.

corrected

Assimil- ation	Increase in per cent
1132
-43
1167	3
-57	33
1140	1
-54	26
mol., not

mol. KCN

Time	trans-
	mitted
12.05	2300
13.00	0
15.05	2300
16.05	0
17.05	2300
18.10	0

voltage

Exp. 60, 15-5-31. Temp. 19.00°. KCN

C. Experiments with Higher Concentrations of KCN,
Showing a Retarding Influence,

In these experiments KCN was added, besides to the
alga, also to the buffer-mixture.

In the first place I call the attention to exp. 61, which
shows the same type as 58, described above, and even
more distinctly. The addition of Viooo mol. KCN causes
an initial fall of 13 %, which, after some hours has changed
into an increase of 21 %.

Retarding influence of KCN, changing in course of time
into an increasing one. Van den Honert's strain.

Most remarkable is the result of the experiments 62
and 63, which were made two days later, in quite the same
way as exp. 61 (cf. table 14).

The retardation is extremely strong in exp. 62, some-
what less in 63. In both the assimilation does not show
any inclination to recover; it has on the contrary, diminished

again after three hours in exp. 63. Respiration is little
affected, in exp. 62 a slight decrease, in exp. 63 even an
increase.

TABLE 14.

Strong retardation of photosynthesis after the addition

of mol. KCN.
5000

Exp. 62, 7-5-31. Temp. 19.00°. KCN

The momentary disposition of the alga is apparently
of great influence on the rate of retardation. In those days
warmer weather had set in. This remarkable behaviour

Retardation of assimilation and respiration by the addition
of higher KCN concentrations.

is all the more striking, when we compare it with exp. 64
and 65 of an earlier date. The concentration of K C N
was much stronger, Viooo and V400 ^oU the retardation
not stronger than in exp. 62.

Results of experiment 64 are given in table 15. Contrary
to Warburg's result with Chlorella, we find that the
assimilation is retarded in weak as well as in strong light,
although to a somewhat smaller measure in weak light
(fig. 14). Exp. 64 is the only example where a corresponding
strong retardation of the respiration is found. In very
strong light the assimilation seems to decrease. It is not
setded yet, if this really points to the existence of a light-

assimilation optimum; perhaps the retardation had become
a litde stronger after some time.

It has been investigated whether the retardation is
reversible. The respiration recovered immediately indeed,
the assimilation increased, though a considerable retar-
dation remained. The next morning, however,it appeared
that also the assimilation had regained its old value.

In exp. 65 (table 15) the retardation is stronger, and,
besides, increases a little in course of time. In weak Hght
the retardation is distinct, though again somewhat smaller
than in strong light. There is again a flat assimilation-
optimum. We may perhaps explain this by an increase of
respiration in the stronger Hght.

D. Experiments, Showing a Complete Inhibition of Assimi-
lation after the Addition of KCN,
Finally, in two experiments, a complete stopping of the
carbon dioxide assimilation was observed (table 16).

The concentrations V2000 and Viooo molv which were used,
show how this phenomenon of retardation is independent
of the concentration used, but seems to depend on the
individual disposition of the alga. In the first experiment
the respiration had not been changed; in the second it
had been retarded, though not altogether wanting. In
exp. 66 there is still at first some assimilation, which
disappears gradually; in exp. 67 the assimilation is
direcdy stopped altogether.

Both experiments are contradictory to the results of
Warburg, as according to him the retardation would
stop at the compensation point of assimilation and
respiration. An explanation of this deviating result of
Warburg may perhaps be attempted, when we observe
that the assimilation under the influence of K C N was
not directly brought to a standstill. When the assimilation
is determined directly after the adding of K C N, as

Complete inhibition of photosynthesis after the addition of KCN.
Exp. 66, 13-5-31. Temp. 19.00°. KCN ^

Warburg did, we may cotitinually find a slight assimilation.
As for the rest it is not excluded that Chlorella behaves
otherwise than Hormidium.

1) Apparent assimilation (-25.5) respiration in the dark (21)
= — 4.5. The respiration in light is apparently stronger than in
darkness.

Let us combine the results of the above experiments
in a few words. It appeared that Warburg's results
could not be confirmed. The addition of potassium cyanide
caused a retardation not only when temperature, but also
when light was the limiting factor. So on this ground it is
not possible to distinguish two processes, one of which
would be sensitive to K C N („Blackman reactionquot;).

It is, moreover, important that this retardation may
disappear in course of time, and pass into an increase.
This speaks strongly against Warburg's conception. It
seems to me that the action of the KCN is much more
complicated.

The fact that the same concentration of KCN on
different days, can influence Hormidium (cultivated under
rather constant conditions before a north-window) very
differentiy, seems unaccountable in a simple way. It
cannot be assumed that the quantity of heavy metal,
probably Fe, which is, no doubt, present in unhmiting
amount, would be so highly subjected to fluctuations
in the cells.

This is also a reason to doubt Emerson's results.
Even small differences in the way of cultivation cause a
much changed sensitivity. This may also be the cause of
the different behaviour of normal-Chlorella and Chlorella
cultivated in a sugar solution deficient in Fe.

Another fact on which Warburg has built up his theory,
the impossibihty to retard the assimilation below the
compensation-point of assimilation and respiration, could
not be confirmed with Hormidium.

It stands to reason that it is not excluded that Chlorella
and Hormidium behave quite differently in this respect.
At any rate it appears, however, that no general validity
may be placed on these facts and the theory built up on
them.

Moreover it is an important fact that the assimilation
can be stimulated by a small dose of KCN and that to
the same extent, no matter whether either temperature
or light is the external factor which limits the process.
This fact, added to the observation that a uniform retar-
dation also enters under both conditions, speaks strongly
against the conception that in both cases different pro-
cesses govern the assimilation. On the contrary, the
uniformity of the controlling process is made plausible
by these facts.__

The stimulation of the assimilation in weak light is also
important for another reason, because it appears from it
that it is not the available light energy as such that limits
the rate of assimilation. If this were the case, a stimulation
would be impossible. That this stimulation is possible
shows us that the assimilation, also when light is limiting,
is not exclusively dependent on the quantity of light energy
added, but at the same time on internal processes. (It may
be possible that they themselves are connected again with
the hght intensity). Consequently it is not true either
that the photochemical part of the process exclusively
controls the entire reaction-chain under these circumstances,
as is now generally accepted.

Other investigators who found a stimulation of the
assimilation, are Bose (1924), who obtained stimulation
with much more diluted solutions, and Schmucker
(1928), who discovered an increase of the assimilation of
Cabomba aquatica in diluted ether of 0.125 % and alcohol
0.3—1 percent. Sabalitschka and Weidling (1926)
found a 100 % increase in acetaldehyd; Schmucker
could not confirm this statement (cf. further Ch. X).

We have seen that assimilation can be uniformly
retarded and uniformly stimulated under different circum-
stances. In case of retardation, we might think of an injury
of the assimilatory apparatus, in other words of direct

action of the poison on the assimilatory process. The fact
that also stimulation has been found, and even a retardation
may pass into a stimulation, points to the action being
indirect, and that the assimilation is affected „from the
interiorquot;. Therefore it is obvious that we must inquire
into the other metabolic processes of the organism. The
extent of the respiration can give us a hint on this metabolic
state. We find indeed that a stimulation of the assimilation
goes parallel with a stimulation of the respiration. When
the assimilation was slightly retarded, the respiration too
appeared to have decreased. These facts speak for the
idea that the increase or decrease of assimilation is a
result of the behaviour of other vital processes.

A strong retardation of the respiration accompanied a
corresponding decrease of the assimilation only in one
case (exp. 64). In the other experiments the contrary
proved to be the case. This fact which is in accordance with
the results of older investigations on the action of K C N
and narcotics makes the above idea, which lays a connection
between assimilation and other vital processes uncertain
again. It does not imply a refutation, however. In order
to escape this difficulty we need only take that in case
of highly injurious stimuli a strong retardation arises
through the direct action of the poison on the assimilatory
apparatus. Another possibility is that a stronger poisoning
calls into existence an increase in respiration, in the same
way as e.g. wounding may act as a respiratory stimulus.

It was Claude Bernard who discovered the retarding
action of narcotics on the assimilation of carbon dioxide
(1878). After him others, among whom Bonnier and

Mangin (1885), Ewart (1896, 97, 98), Jacobi (1899),
Irving (1911), Thoday (1913), also dealt with this
subject. As a result of these investigations it appeared a.o.,
that photosynthesis is retarded by concentrations that have
no influence on respiration.

The question has been thoroughly reinvestigated by
Warburg (1919, 20), who stated that the assimilation is
retarded in about the same degree, whether light, COg or
temperature is the limiting factor of the process. From
this Warburg concluded that photosynthesis is a surface-
phenomenon, just like respiration. The action increases
in a homologous series with the adsorbabihty. Warburg
confirmed that higher concentrations are necessary to
retard the respiratory process.

In Chapter V I discussed already some experiments
in which the assimilation of COo was investigated when
phenylurethane or antipyrin had been added. Both sub-
stances appeared to retard the assimilation.

Though Warburg already showed that the assimi-
lation altogether recovers after removal of the narcotic,
I performed a few experiments on the matter.

The assimilation was measured before, during and after
the narcosis. The narcotic was removed by transferring
the alga on its old nutrient medium, part of which had been
reserved for this purpose. This is not an absolute removal,
but it is certainly a very strong dilution. The experiments
showed that the recovery is perfect. There may even be
a slight increase after the removal. This manifests itself
distinctly in experiment 69. The increase amounts to
25 %, notwithstanding a shght loss of material during the
transfer. This is much more than could be caused by
„growthquot;. According to van den Honert the correction
for growth should amount to 5.7 per cent in this experiment.
We see the phenomenon is more complicated than a simple
physical removal of the narcotic from the surfaces.

I thought it interesting to determine the action of phenyl-
urethane on the whole light intensity-assimilation curve,
just as was done when studying the influence of potassium
cyanide. An example of such an experiment we find in
table 17, graphically represented in fig. 15. In this figure
the experimental points are numbered, the numbers indi-
cadng the time at which the determinadons were made.
It directly appears, in this and all following experiments,
that the assimilation recovers from the first decrease, as

will be clear from a comparison of the points 5 and 9. A
curve has been drawn through the points 7, 8 and 9. The
decrease appears to be uniform over the whole region,
which is in accordance with Warburg's results with
urethane, hut also with my own results with potassium cyanide.
Respiration has also diminished by 13 per cent.

Experiment 72 (table 18) gives the same results. Recovery-
is even more apparent. A comparison between the numbers 4,
7 and 9 shows this, but at the same time it is clear that
the difference between 7 and 9 is not great, so that after
some time a constant rate of assimilation seems to be
reached. Retardation of respiration is as large as of assimi-
lation. This points again to the possibility of a connection
between the two processes, of which examples were also
found in the preceding Chapters (VII and VIII).

One may ask whether also respiration shows an initial
strong, retardation that decreases later on. An answer to
this question is sought in experiments 73—76 recorded
in table 19. In these experiments a recovery of respiration
always occurred, though the results are not altogether
convincing. The finest example is supplied by exp. 76.
Retardation decreases uniformly, and the depression of the
respiration is in an ideal way in accordance with that of the

assimilation. Whatever may be the cause of this recovery,
it might be based on the urethane being oxidized, at any
rate it is of importance, as the uniform retardation of
assimilation and respiration is here illustrated in an unam-
biguous way.

This supposition of the oxidizing of the urethane is,
however, unlikely on other grounds. In the first place it

appears that the retardation has not quite disappeared even
after several hours, but has only diminished. Also the
experiments in Chapter V seem to contradict it. How
could I have found the deviating result, represented in
fig. 6, if the urethane had lost its efficacy? And finally
there is the. analogy with the action of KCN and Ba CI
(Chapter VIII and XI), where oxidation is out of the
question.

The influence of phenylurethane on assimilation and
respiration in course of time.
Exp. 73, 22-4-31. Temp. 19.00°. Phenylurethane 2.9 X 10-^ mol.

I consider this analogy of great importance. The assimi-
lation reacts in all cases in a similar way, it does not matter
which poison is the cause. The conclusion is evident that
the initial large retardation is caused by a temporary lowering
of the vital intensity of the protoplasm, which slowly
recovers from the „shockquot;. The decrease, and recovery

afterwards, cf the respiration also points in this direction.
So it is not a mere matter of course that the urethane
exclusively acts by diminishing the available surface of the
assimilatory apparatus. On the contrary, it appears that
something may be said in favour of an indirect influence
of the injurious substance. Also the above-mentioned fact
that the assimilation has been accelerated after the removal
of the narcotic, and not simply recovers its former
rapidity, is an argument in favour of this conception.

I made another attempt to prove, in a more direct way,
that the urethane is not oxidized by the cells (exp. 77).
I applied to a recovering algal film the same dose of
urethane. In case the assimilation did not fall again, I
should have proved that the urethane was not oxidized.
However, the reverse appeared to be true. The retardation
of 26 % had fallen to 7 %, a new dose enhanced it to
30 %, after some time it was 15 %. Though the result is in
accordance with the supposition of oxidation of urethane,
it has not conclusively proved it. It is not at all unlikely,
that, owing to the absorption of the alga, the concentration
of urethane in the liquid of the film had quickly diminished,
and that, therefore, the addition of fresh urethane had
caused the new decrease of assimilation.

Experiments 78 and 79 give us an insight into the influence
of other concentrations.

The very slight concentration in exp. 78 (2.4 X 10quot;^ mol.)
causes a small decrease (ass. 5—14 %, resp. 10 %), after
which no noticeable recovery sets in. Stimulation does
not occur here.

Exp. 79 is made with a stronger solution (1.2 X 10quot;^ mol.)
The retardation is considerable, but diminishes gradually
(from 77 % to 50 %). Respiration has been retarded less
strongly, but recovers in the same way (from 15 % to
5 %). This, too, is in accordance with the results with
potassium cyanide; assimilation is more sensitive to high
concentrations of the injurious substance than respiration.

The preceding three chapters more or less all point
to a connection between photosynthesis and respiration.
It was a matter of course that an investigation of the assimil-
ation under circumstances under which respiration increases
was now to be taken in hand.

A great many investigators, of whom Borodin (1876)
was the first (further: Aereboe (1893) Meyer and
Deleano (1911, 13), Kniep (1914), Pantanelli (1914),
Harder (1915), Plaetzer (1917) have shown the increase
of respiration after illumination. This increase was ascribed
to the production of oxidizable materials. Warburg (1922),
Emerson (1927) and Genevois (1927) directly showed
an increase of respiration by adding sugars to the algal
suspension. In a preliminary experiment I could confirm
this with Hormidium (a pure culture; increase 33 % in
0.7% fructose). If I should also succeed in stimulating the
assimilation under these conditions, then the relation
between assimilation and respiration, or, at least, between
assimilation and other protoplasmic processes connected
with respiration, would be very convincingly obvious,
because from a chemical point of view a retardation of
assimilation should be expected rather than an acceleration,
as sugars are indeed the final products of assimilation.

Spoehr and Mc Gee (1923) have observed that the
sugars disappear out of the leaves in darkness, and that
after a long stay in the dark the rate of assimilation has
diminished. After continued exposure to light, both the
rates of assimilation (cf. also Osterhout and Haas,
1918; Harder, 1930; Arnold 1931) and of respiration
increase with time as the sugar content increases. They

consider this increase of assimilation as a result of the increase
of sugar-content and respiration.

So the connection between assimilation and respiration,
according to Spoehr and Mac Gee, consists in a depen-
dence of the former on the latter. It would appear, however,
even more convincingly, when the supply of sugar took
place from the exterior, because in that case it would be
proved that the increase of sugar content was really the
cause of this phenomenon.

Treboux (1903) examined the action of sugars, and
found that they lower assimilation in Elodea. The concen-
trations studied were high, and the lowering was probably
caused by osmotic action, as Treboux thought.

I added the sugar by dropping a number of drops of
a sugar solution on the algal film. Beforehand I removed
the nutrient solution for the greater part by means of
filter-paper, so that the final solution, was only a little
more diluted than the solution added.

The sugars investigated were glucose and fructose. The
action was investigated both in strong and in weak light.
The Hormidium strains of Pringsheim and van den
Honert were both used. The results of the experiments
are to be found in table 20.

From these experiments an increase of assimilation
appears, when a sugar solution of a concentration of
0.7—1.0 % was added, both with the pure culture of
Pringsheim and van den Honert's strain. In a 2 %
solution Pringsheim's strain gave a slight lowering of
CO2 assimilation. In strong light the increase is, to be
sure, not exceedingly large, the values found are 5, 12,
9, 12, 5 and 6%; but an increase of 655—730 in exp. 82
and another of 1097—1199 and 1233 in exp. 83 far exceed
any experimental error. I consider the increase in weak
light of great importance. This result is a confirmation of
the results of experiments communicated in Chapter VIII,

in which it has been proved that in low light intensity-
assimilation is stimulated by the addition of a small dose
of KCN.

The influence of glucose and fructose on assimilation and
respiration.

Exp. 80, 8-5-3L Pringsheim's strain. Temp. 19.00°. Glucose 1 %.

It is peculiar that the respiration shows a decrease in
two experiments. It is more or less clear that respiration
decreases when the alga is taken away from daylight
(Chapter III), but it is strange that an addition of the
sugar does not counterbalance this effect.

Exp. 83, where I met with this phenomenon, has been
continued. The sugar was'removed and assimilation and
respiration were again determined. The removal appeared
to result in a new small increase. The assimilation rose
from 1233 to 1277, and was now 16 % higher that in the
beginning. Further the increase of respiration was note-
worthy, which recovered from 63 to 78, and was now

even 5 % higher than at the outset. This may point to the
action of the sugar being complicated, too. It may be that
the stimulating action is checked by an injurious, possibly
an osmotic, action, and that this accounts for the divergence
of the results.

One more experiment was made with a culture (van den
Honert's strain) which had stood in the dark continually
for 62 hours (exp. 87). The concentration added was 0.7 %.
Contrary to the expectation, that now the influence would
be great, it appeared not to be so. Respiration really increased
by 90 %, but assimilation proved to diminish by 24 per cent.

These few deviating results may, however, not turn
our attention from the important fact, that it has become
evident in this chapter, that an addition of sugars, which
in most cases brings about an increase of respiration, causes,
at the same time, an increase of assimilation. This fact
points to the assimilation being affected by the protoplasm.
According to its being provided with more sugar the
assimilation increases, altogether in accordance with the
conception of Spoehr and Mac Gee.

The increase also takes place in weak light and this
speaks in favour of the unity of the assimilatory process
which in its entirety is controlled by internal processes, and
not exclusively by external factors.

Now that it appears that the external factor light (when
it is limiting factor) alone does not determine the intensity
of the assimilation, but that the effect is also dependent
on internal factors, we have to consider the possibiUty,
that the increase of assimilation in stronger light is not
exclusively brought about by more energy available for
the photosynthetic process, but that the protoplasm, too, is
affected by the light intensity, and that the effect of stronger
light on the protoplasm promotes the assimilation again.
The protoplasm therefore may ultimately determine the
rate of the assimilation; the action of the external factor

This opinion is supported by the fact that light exerts
a strong influence on the protoplasm (Chapter III). It
appears that the organic system itself does not remain
unaltered, that among the factors in the photosynthetic
process, light is probably not the only variable.

In connection with the described increase of the assimi-
lation by the addition of respiratory material, it should
be noted here that Sabalitschka and Weidling
(1926) found a stimulation of the assimilation of as much
as 100 % by the addition of acetaldehyde, which can also
serve as respiratory material. The concentration was small
(0.016—0.032 %) and the increase is, according to them,
effected by a stimulating action of the substance. This
result could not be confirmed by Schmucker (1928).

Finally it may be mentioned that attempts have been made
to stimulate respiration and assimilation by putting the
algal film in water distilled from glass into glass. After
remaining there for some time, this water was again replaced
by fresh. For Shibata (1929) has found with Chlorella,
that washing with distilled water enhances respiration.
In my experiment neither respiration nor assimilation was
affected by this treatment.

While in the preceding chapter the question was treated,
how carbon dioxide assimilation behaves under circumstances
in which respiration is promoted, we shall now put the
question how assimilation behaves if respiration has under-
gone a strong decrease. The reply to the first question
appeared to be that under these conditions assimilation

was promoted as well. Will this parallelism also repeat itself
in case of retardation?

In the preceding chapters we have found instances of
a strong retardation of photosynthesis, but a great decrease
of respiration is still wanting (except exp. 64' with KCN,
where both processes appeared to be greatly retarded).
We find the best example in the chapter on the influence
of urethane, where both respiration and assimilation were
slightly retarded.

The question was now to find a substance that has a
retarding influence on the respiration. I found this
substance in the paper of Shibata on the antagonistic
action of electrolytes on respiration (1929). According to
this research the respiration of Chlorella in a solution of
one mol. Ba Clg is diminished by 80 per cent. This substance
seemed suitable to me for that reason, and also, because
the depressed respiration, according to the statement of
Shibata, remains rather constant, contrary to the result
with (likewise strongly retarding) Ca-salts. The concen-
tration I used in my experiments amounts to 0.5 mol.

In Chapter III the decrease of respiration in the dark
has been stated. In order to prevent that a decrease of
respiration could partly be caused by the influence of
darkness (this influence being high in the month of June,
as a result of the high light intensity), the algae were placed
in the dark for some hours before the experiment.

The nutrient solution was removed by once washing
the algal film with distilled water before the addition of
Ba Clo solution. In table 21 the results of the experiments
have been given.

Exp. 89 and 90 have been made in strong light, exp. 91 in
weak light. In the former experiments, both respiration and
assimilation appear to be strongly diminished, the former
(40 and 56 %) even a httle more than the latter (25 and
44 %), but the agreement is nevertheless very sufficient.

The influence of 0.5 mol. Ba Clj on assimilation and respiration.
Van den Honert's strain.
Exp. 89, 13-6-31. Temp. 21.00°.

Exp. 89 gives retardation as a function of time. Altogether
in accordance with the results of the experiments with
potassium cyanide and urethane, we see the recovery of
both assimilation and respiration. So it seems that here
we have a very general reaction before us, of respiration
as well as of assimilation. In the uniform retardation of
the two processes and the simultaneous recovery, I see an
indication of a connection between both vital processes.

It is important that Shibata, too, regularly finds a
recovery after an initially strong decrease, and that after
the addition of very different electrolytes. From Shibata's
figures this is to be seen at a glance. Ca only excites a con-
tinual decrease in high concentrations. The activity of this
substance is analogous to the action of higher KCN
concentrations on the assimilation.

BaCla has been replaced again by distilled water in
exp. 90. Respiration recovers altogether, assimilation only
partly. We have already an example of this phenomenon
in exp. 64, where the removal of KCN had a similar
effect. We must admit that this certainly does not point
to a direct connecdon between the processes of respiration
and assimilation.

Exp. 91 has been carried out in weak light. The retar-
dation of the assimilation seems to be a litde smaller
(analogy with action of KCN); against a retardation of
respiration of 48 %, there is a decrease of 22 % only.

In the last experiment (92) the influence of 1 mol.
Ba CI2 was investigated. Here it appeared that the assimi-
lation was totally, or almost totally inhibited (decrease 574
to 2). On the other hand the respiradon still amounted
to 23, against 48 before. To strong stimuli, therefore,
assimilation is more sensitive than respiration. This also
appeared to us when studying the influence of higher con-
centrations of KCN and of phenylurethane.

1. Introduction. In the preceding chapters we have
more than once been led to the conclusion that the
influence of the external factors is not direct, but that the
rate of the assimilation is determined by the inter-

mediary of the protoplasm. Even when light is limiting
and the increase direcdy proportional to its intensity, it
appeared that the assimilation could be stimulated by
factors that may cause a similar increase in respiration.
When this conception of the indirect action of external
factors, is right, then it is to be expected that the assimi-
lation will not reach its full rate directly after the beginning
of the exposure to light, but that some time will pass before
the protoplasm has adapted itself to the new conditions
and the assimilation has come to a constant intensity.

Warburg (1920) already established that in Chlorella
the assimilation is indeed smaller in the first minutes, and
within a short time rises from nothing to the final value.

Quite in accordance with his physico-chemical con-
ception of the assimilatory process Warburg is of opinion
that this increase in assimilatory activity is due to the
existence of a photochemical induction-time in photo-
synthesis.

He came to this discovery, because he began intentionally
to investigate, whether photosynthesis is characterized by
a photochemical induction period, as has been proved
to be the case in other photochemical processes. Therefore,
we can readily understand that, when he stated that
photosynthesis, too, showed an induction period, he did not
think of any other way of explanation of the induction
phenomenon, though he remarked that the induction of
photosynthesis is not quite identical with the induction
of the photochemical reaction of hydrogen and chlorine.

Possibly the influence of light on assimilation is more
or less indirect; a change of the protoplasmic structure
might be caused by illumination, and of this protoplasmic
change the start of the assimilatory process might partly,
or totally,^ be the result.

Certainty is altogether wanting, and calling the pheno-
menon quot;photochemical inductionquot; is consequentiy hypo-

thetical. An explanation as proposed here, by which the
induction is explained by means of preceding protoplasmic
processes that are induced by light, seems equally probable.

It seemed interesting to investigate whether this induction
time could also be observed with Hormidium, in order to
settle which of the two hypotheses was true. This could be
done by investigating the influence of temperature on the
phenomenon. If the process were of a physiological nature
then temperature had to influence it, just as temperature
affects other physiological processes, where light plays a
part (e.g. phototropical phenomena). On the other hand
no distinct influence of temperature is to be expected
from a phenomenon that is merely photochemical.

2. Experiments on the Induction of Photosynthesis at
Different Temperatures. The presence of the phenomenon
was demonstrated at 20°. When this appeared to be entirely
in accordance with Warburg's result, the experiment
was repeated at 14° and 26°, in order to answer the second
question. The experiment was made as follows. The algal
film was repeatedly illuminated for 0.5, 1, 1.5, 2 or
3 minutes. Between the exposures the culture was in the
dark for a much longer period. After due correction for
respiration the assimilation during a period of exposure
could be determined. The respiration was not constant
but continually falling, and had to be determined several
times during the experiment. The short exposures had no
observable influence on respiration (determined in the
dark), so that an error was possibly not made in the
respiration-correction (see Ch. Ill; yet an increase of
respiration during the short exposure, without an after-
effect on the respiration in darkness, is equally possible).

The illumination was effected with the projector lantarn
at full capacity. A new lamp with a littie more candle
power had been placed in it. In spite of this fact the

light was no longer quite in maximum at 26° and the
assimilation about 25 % lower than it could have been.
Yet I gave the preference to this illumination, as in this
case the light passed a layer of water of 16 cm thickness,
by which desiccation of the algal film which can easily
occur in a prolonged experiment at this temperature,
could be prevented.

The assimilation in the first minutes after the beginning of the
illumination. Van den Honert's strain.

Exp. 93, 5-6-31. Temp. 20.00°. Assim. at 10.35 :795; per minute 13.25,
reduced 10. Respir. at 11.20:73, at 12.50:54, at 15.47:50, at

Exp. 94, 8-^-31. Temp. 14.00°. Assim. at 11.25 : 549; per minute 9.15,
reduced 6. Respir. at 12.55 : 46, at 15.50 ; 38, at 18.35 : 32.

Exp. 95, 9-6-31. Temp. 26.00°. Assim. at 10.50 :972; per minute 16.2,
reduced 13. Respir. at 11.40 : 72, at 12.31:50, at 14.50:47, at

Before the periodical exposure the assimilation in constant
light was determined. This is the optimum velocity which
the assimilation can reach at alternative exposure.

In order to facihtate a comparison the optimum rate at
20° was put at 10 per minute. The optimum rate at 14°
and 26° reduced to this value, can be calculated from
fig. 11 in chapter VII. We find 5.75 and 14.8. Now that
the Hght is not quite in maximum, the increase will be
a litde slighter, especially at high temperature. It seems
to me better to round the numbers to 6, 10 and 13. The
results of the three experiments are to be found in table 22.

The great accordance with the results of Warburg is
evident from these data. This is distinctly shown by fig. 16.
It appears that the final optimum rate is fairly well reached
after two minutes. What strikes the eye most is the great
difference between the three curves. At 26° already 1 mm=^
has been assimilated after 12 seconds, at 14° this takes
55 seconds. This is certainly not a right measure for the
induction process. If the possibility for assimilation is
given as a result of the running down of the induction
period, a larger quantity will be assimilated at higher
temperatures, because the assimilation does proceed more
rapidly at higher temperature. But this needs not point to
a faster rate of the processes contributing the induction-chain.

In order to setde this point we have to see whether the
optimum speed of the process is sooner reached at higher
temperature than at lower. In practice it is difficult to
state where the transition-point between curved and straight
line is situated. It is much easier to fix another point; viz.
the point at which the speed is one half of the final
speed. A cross in- dicates this point in the figure.

This point is reached at 14^ after 64, at 20° after 54,
and at 26° after 24 seconds. A considerable difference is
apparent. In spite of the fact that the optimum rate at
26° is much greater than at 14°, this higher rate ofassi-

milation is reached much quicker than the lower at 14°.
Therefore, the in- duction is greatiy influenced by tem-
perature, so that a chemical protoplasmic process plays a

We can also get
an impression to
what extent tem-
peratur einfluences
the induction, when
we state the tem-
peraturecoefficient
from the values
found at 14° and
26°. Qjo appears to
be 2.2 and is stri-
kingly in accor-
dance with the
temperature-coeffi-
cients of respiration
and photosynthe-
sis. This affords a
new argument for
the conception that
induction is a process of a protoplasmic nature.

It has been established, therefore, that, before assimilation
reaches its full intensity, a reaction takes place which is
sensitive to temperature and caused by light. This points
to a new indirect action*of both factors in the photo-
synthetic process. A chemical reaction must occur in the
protoplasm under the influence of both factors, before the
proper photosynthetic activity may begin.

The increase of assimilation, described above, must be
distinguished from that which Osterhout and Haas
(1918), Spoehr and Mac Gee (1923), Harder (1930)
and Arnold (1931) observed during the course of the
experiment.

Harder could demonstrate that the assimilation of
Cladophora and Fontinalis somedmes kept increasing the
whole day. This rise was not interrupted by periods of
darkness, on the contrary this caused an increase of even
longer duration. Finally an opdmum was reached, and a
decrease set in. Thus Harder showed that the assimi-
lation under constant conditions is subject to alteration.

Osterhout and Haas found a similar phenomenon
with Ulva thallus. They stated, however, that the assimi-
lation was already constant after ly^ hours.

Finally, Spoehr and Mac Gee observed this increase
with leaves which had been in the dark for a long space
of dme. As said before, this increase paralleled an increase
in sugar content and respiration, and they pointed to the
latter factors as the cause of the increase in assimilation.

It appeared already to van den Honert (1928, 30)
that an increase during the whole day, as was found by
Harder, does not occur in Hormidium, and that the assimir
ladon remains fairly constant. Only a sUght increase of
about 1 % an hour at 20° was stated. This result was
regularly confirmed in my experiments. Van den Honert
thought that he could explain this by an increase of the
living material.

However, when I look through my protocols, it is
evident that there exists an initial increase, after a long
stay in the dark and also in the dark wintermonths. A few
examples will illustrate this:

exposure. Ass. at 20°, strong light:' 22 in 5 min., 54 in 10,
29 in 5, 31, 32, 35, 34, 35, 37, 35, 34. The last 6 values
are fairly constant; so the assimilation increased during 45
minutes.

Exp. 19th Nov. '30. Ass. per 5 min.: 28, 39, 44, 45, 43,
41, 42, 42. First determination after 10 minutes' exposure.
After 20 min. the assimilation is constant.

Exp. 22nd Dec. '30. Exposure from 1.26 to 2.35;
1.40—1.50 : 23; then 28 in 10 min., 17 in 5, 82.5 in 30.
Constant after 34 min. exposure.

Exp. 24th April '31. After 16 hours' darkness. First
determination after 20 min. exposure. Ass. 56 in 5 min.,
62 in 5, 62 in 5, 132 in 10, 127 in 10, 127 in 10; constant
after i 25 minutes.

The above examples, to which I could add a few more,
give an impression of the assimilation during the first
hour. There is, apparently also in Hormidium, an increase
of assimilation, as Osterhout and Haas found in Ulva.
The only difference is that the constant rate is reached earlier.

In the preceding chapters we have seen that, all other
conditions being constant the assimilation can be stimulated
by the addition of certain substances such as KCN,
narcotics and sugars; so it is not at all impossible that
substances formed during the assimilation, stimulate the
assimilation, as Spoehr and Mac Gee think. I do not
mean by this to give an explanation of the increase as found
by Harder. It would seem to me very unlikely that this
increase of long duration would entirely be a result of
improved nutritional conditions. I only want to point
here to one of the possible causes of this phenomenon.

It also appeared that the respiration increased during
the exposure by purely plasmogeneous action of the light
(Ch. III). If it is true that the assimilation can be influenced ,
by other vital processes, the increase of assimilation may also
be explained by this plasmogeneous influence of light.

It was mentioned before that some of Warburg's
results are contradictory to mine. Of one of them we
could say how Warburg might have arrived at his
results, but not with any amount of certainty. It seems
much more probable that the differences are due to the
experimental material, and that Hormidium and Chlorella
behave differently. Only a thorough, comparative study
of the assimilation of these and other organisms, which I
consider of the greatest importance, can throw light upon
this subject.

Should my opinion be right, it seems better to base
my considerations on my own results. It is of more
importance that we understand the assimilation in one
organism than that we attach too much importance to
differences, possibly based on individual peculiarities.
Especially if actually assimilation is controlled by other
vital functions, there is no reason to be astonished at a
different behaviour. There would be more reason for such
astonishment if we had always studied the quot;realquot; photo-
synthesis, and the plasmatic factor had showed itself to
be negligible.

We need not, therefore, to worry about the apparant
discrepancy of our results and those of Warburg. It
seems rather a support for the idea that photosynthesis is
controlled by the organism which has an individuality of
its own.

A. The Assimilation Reacts to the Addition of Uncommon
Substances in a Similar Way under Different Conditions,

added, is as much retarded in strong as in weak hght
(temperature or hght intensity as hmiting factors). We
meet this in case of retardation owing to three akogether
different causes, e.g. addition of a. phenylurethane, h.
KCN, c. Ba Clg. The recovery from this injury acts
similarly, both in high and low hght intensity.

II. The assimilation may be stimulated, and again
the increase is of the same extent under both conditions,
whether this stimulation is caused by a. diluted KCN,
or b. glucose and fructose.

Therefore we may conclude that the assimilatory process
behaves as a unity; it does not matter whether either hght
or temperature is the limiting factor.

In relation to this conclusion the work of Briggs may
be mentioned here. Briggs (1922) found that the photo-
synthetic activity of plants grown in culture solutions
devoid of one of the elements necessary for normal growth
was less than that of normal plants. This depression
resulted whether light or temperature was the limiting
factor. However, Briggs' results were contradicted by
Gregory and Richards (1929). These investigators
stated a shght depression of photosynthesis in Hght of low
intensity, whereas the depression proved to be very
marked in high light intensity. These very interesting
results, however, may not be compared with the results
of my experiments mentioned above, in which the imme-
diate reaction of photosynthesis to the addition of unusual
substances has been studied.

A first argument for a dependence of the assimilatory
process on internal factors is the inconstancy of the
assimilation while the external factors are kept constant.

Osterhout and Haas, Spoehr and Mac Gee, but
especially Harder and Arnold have described this

phenomenon with other organisms. In Harder's experi-
ments the assimilation appeared to increase during the
whole day. The decrease, which followed subsequently,
and also the decrease that Ewart, Pantanelli, Montfort
and Neydel, and Arnold could demonstrate, shows a
similar dependence on internal factors. The results of
Ewart and Pantanelli are partly based on injury and
accumulation of chloroplasts. An increase as described
above also proved to exist in my experiments with
Hormidium. The assimilation was inconstant after a long
period of darkness. However, a constant rate of assimilation
was reached after a rather short time (± ^ hour).

Another reason for the dependence of the assimilation
on internal factors is the parallelism with the respiration
(with Hormidium), which occurs in many cases.

The first striking parellelism between the two processes
is that both assimilation and respiration may be stimulated
with the same amount of greatly diluted KCN. Moreover,
the assimilation is stimulated by the addition of sugars
which in the first place may be looked upon as respiratory
material, and usually stimulate the respiration. This seems
to be important because it refutes a naive application of
the mass-law to the process, according to which law a
reaction product (sugar) should retard the reaction (photo-
synthesis) instead of stimulating it.

This parallelism is the more important as an increase in
the rate of assimilation results, no matter whether the external
conditions are such that light or temperature is limiting.

Other examples of a parallelism between the two processes
are instances of a slight retardation of photo- synthesis
(in weak and strong light!), which are always accompanied
by a small decrease of respiradon. This was found in case
of retardation by KCN, phenylurethane and Ba CL. The
recovery of a slight retardation, which always occurs in course
of time, proceeds in a similar way in both processes.

The assimilation is more considerable in strong than in
weak light. Nevertheless we have seen that it can increase
in weak light under changed internal conditions (addition
of sugars). So we supposed that the increased assimilation
in strong light, besides being a result of the greater quantity
of light that can do photosynthetic work (direct action), is
also a result of internal factors, changed under the influence
of light (indirect action). There is all the more reason for
this opinion, as we were able to show that the protoplasm
is strongly affected by Hght. Respiration increased consi-
derably, which is, to be sure, caused by the formation of
sugars (ergastogeneous acdon; at the same time a cause
for greater assimilation), but for the greater part Hght
affects the protoplasm directly (plasmogeneous action). The
actively assimilating organism also respires actively.

Temperature affects both processes in a similar way.
The idea of a possible connection between them would
be rather untenable, if this had not been the case. Heating
causes a very regular acceleration of the two processes,
so that the types of the resulting curves are altogether the
same. The coefficients of temperature differ only very
little; from 15°—25° Qio is 1.96 for the assimilation, 1.91
for the respiration. The resemblance is so great that it
could be accounted for by a full dependence of assimilation
on respiration (Spoehr and Mac Gee).

The optimum rate of photosynthesis does not set in
immediately after the beginning of the exposure to Hght.
Temperature is of great influence on the duration of this
induction-period, by which it is proved that the phenomenon
is not founded on photochemical induction (Warburg).
So an internal chemical reaction (or a chain of reactions)
must precede assimilation. The temperature coefficient of
this induction time resembles the coefficient of respiration,
though a very accurate determination seems impossible.
From 14°—26° Q^ amounts to about 2.2.

In addition to our statements of a correlation between
assimilation and respiration in Hormidium, it may be
mentioned here that Pies ter (1912), Boysen Jensen
(1918), Henrici (1918), Spoehr and Mac Gee (1923),
Gregory and Richards (1929), found a correlation be-
tween the intensity of assimilation and the intensity of
respiration in other organisms.

These were the arguments in favour of a relation between
assimilation and respiration, now a few will follow that seem
to point in another direction. KCN, phenylurethane or
BaClg in high concentrations, causing a strong decrease
of assimilation, affect the respiration in quite another
manner. An entire cessation of the first process may be
accompanied by a rather strong or slight retardation, or
even by a slight rise of the respiration. This result is in
accordance with the results of other investigators (Jacobi,
Irving, Thoday, Warburg).

Another discrepancy arose after removal of KCN or
Ba Clg in my experiments. The respiration recovered in
this case at once tç the initial value, the assimilation
recovered incompletely. In case of K C N the assimilation
also managed to recover after a longer period, but at all
events there was a difference in the time in which a com-
plete recovery set in.

The parallelism of the two processes was to us a ground
for the dependence of the assimilation on internal factors,
and the few cases in which this parallelism did not
hold, certainly cannot invalidate the occurrence of this
phenomenon. It is plausible that, when an organism is
injured in the first place the normal vital functions as
metabolism, growth, propagation are stopped, without
death following inevitably. On the contrary; the organism
will defend itself against the injurious influences and

consequently may manifest an increased oxygen con-
sumption. It is a well-known fact that e.g. wounding
stimulates the consumption of oxygen.

In our case a high concentration of injurious material
retards the normal function of the organism, the assimilatory
apparatus (or perhaps also an internal regulatory process),
suffers direct injury. But the organism is not yet dead,
it can defy much larger doses of poison before respiration
will at last come to a standstill.

If this opinion is correct the quantitative determination
of respiration in these stronger concentrations has lost its
importance for the study of photosynthesis.

In exp. 63 I even stated an increase of respiration of 14 %
in high concentration of KCN. Irving (1911) and
Warburg (1921) found a strong stimulation in high
concentrations, in which assimilation is greatly retarded.
They found, however, a gradual decrease of respiration
after exposure to very high concentrations.

This may show the cause for the difference in
Warburg's and my own results. The „pathologicalquot;
rise of respiration may have started a litde earlier in
Chlorella, and may also be a litde stronger than in
Hormidium, so that a parallehsm might have become
totally hidden.

So the non-appearance of parallelism in case of strong
retardation need not be contradictory to the parallelism
observed in case of moderate retardation.

The other discrepancies are no more of great importance.
The assimilation, after the poison being removed, did
not recover directly, though the respiration had recovered.
But ultimately we may also take this as an argument in
favour of a connection with internal processes (recovery
from injury, which takes time).

The discrepancy mentioned in the chapter on the
influence of sugars — a retardation of respiration was

found twice, though the assimilation had increased — is
neither very convincing. After the removal of the sugar
the respiration appeared to increase, so that the presence
of sugar, though beneficial to some processes, appeared
to be injurious to others (perhaps osmodc acdon). So the
influence is probably more complicated than one would
superficially think (see also Genevois, 1927) and the devia-
tion found is, therefore, no more an important argument.

We now come to discuss the quesdon in which way
we must explain the parallelism of assimiladon and
respiration.

1.nbsp;Spoehr and Mac Gee think that the assimilation
is direcdy dependent on the respiradon, and that very
active splitting products of carbohydrates, which arise by
respiration, play a part in the assimilatory process. We
shall see if there is any reason to accept such a direct
connection.

2.nbsp;One may also have the idea that assimilation does
not depend direcdy on the respiratory processes them-
selves, but on some protoplasmic functions in general, and
that, consequendy, there is a certain parallelism with the
consumption of oxygen, which is so to say a resultant
of all protoplasmic functions.

3.nbsp;Finally we may suppose that this parallelism is not
based on a causal dependence of the assimilation on other
processes, but that it is based on a joint cause, which acts
on all processes in a similar way. We might for instance
imagine that the colloidal structure of the plasmatic proteins
changes, and consequently the processes which depend on
such a structure.

Let us first consider the third case. We can easily see
that a retardation of both processes may be caused by a

structural change. The surfaces where the vital processes
occur may be destroyed, or taken up by strange substances
by which essential substances are expelled.

In this connection it may be mentioned here that Briggs
(1922) holds the opinion that the weaker assimilation of
plants grown in insufficient nutrient solution, would be
caused by a retarded development of the reactive surface.

The above theory could not, however, explain the recovery
of a function. If this recovery had been found only once,
another explanation would certainly have been possible.
The phenylurethane might have been oxidized, the KCN
dissociated otherwise under the influence of a changed pH
(a result of paralysis of funcdon), etc. The very fact that
the recovery is the same under all influences (probably
also after extreme temperature (cf. p. 612—613), addition of
chloroform (Irving), or electrolytes (Shibata) — points,
however, to an active interference of the organism.

That the alteration of the assimilation under uncommon
influences would be simply caused by a passive structural
change seems, therefore, very improbable. The organism
must always assist with an action of its own, either passively
(e.g. more sugars, more oxidation), or actively, which will
be the case in a recovery.

So the first mentioned two possibilities remain. Does
the respiration supply the energy necessary for the assimi-
latory process direcdy (1), or is the assimilation controlled
by definite vital functions, which in their activity are more
or less dependent on the total vitality, consequently also
on the oxygen consumption, which we may take as a
criterion of the vitality (2).

The temperature coefficients of assimilation and respir-
ation do not confhct with the first opinion, as they are

almost equal. The equality offers even a somewhat greater
support for this opinion. Dependence on other vital
functions might be in accordance with different Qiq. It is
inessential and improbable that all vital functions of the
same organism are affected by temperature in the same way.

Neither does the dissimilar retardation in high concen-
trations of injurious substances speak against a direct relation,
if we assume that the superficial functions are injured,
and the increase of respiration has a quite different cause;
it might be the result of the active reaction against the
injurious influence.

Other facts, however, seem to contradict such a depen-
dence. We found that light causes respiration to increase.
However, in the experiments in Chapter XII the agreement
is less satisfactory. In those experiments we started to
determine both assimilation and respiration. The latter
proved to be very high, which seems plausible, since in the
month of June the alga got daylight already for a great
many hours before the experiment began and the nights
were short. It appeared now that respiration fell considerably
in the course of the day, in spite of repeated illuminations
of short duration. The assimilation, however, was able to
rise nearly to the first stated level within a few minutes,
without the respiration being interrupted appreciably in
its fall. The possibility remains, however, that the respir-
ation did indeed increase during the short period of illumi-
nation, but that this short exposure had no after-effect.
But even if this is not the case, the hypothesis need not be
abandoned yet, though it cannot be maintained unaltered.
One may claim that a certain amount of respiration is
necessary, but that, as soon as it has been reached, a further
increase can only be of little influence. Respiration is
here considered to be a factor that can be limiting, or not.

The experiments on the influence of sugars, however,
do not agree very well with this amended hypothesis.

Parallelism after a treatment with sugars was not clearly
expressed. It is true that respiradon increased in general,
but this increase did not agree much with that of assimi-
lation. However, it seems to me too early to reject the
interesting hypothesis of Spoehr and Mc Gee in the
face of my experiments.

Personally I am of the opinion that the remaining second
case explains the facts in the most satisfactory way. There
is no direct causality between assimilation and respiration,
but a relation between assimilation and some protoplasmic
functions, which are, however, (like all vital functions)
closely connected with respiration. The connection between
the two processes of assimilation and respiration may be
less close now. We need not be surprised when, after
KCN or Ba Clg being removed, respiration recovers more
quickly than assimilation, while the somewhat irregular
behaviour in sugar-solutions is no more disquieting.

Whatever the explanation of the parallelism of respiration
and assimilation may be, we must claim a connection
between photosynthesis and other protoplasmic processes.

b.nbsp;the parallel stimulation or slight retardation after the
addition of chemicals;

d.nbsp;a chemical process preceding the assimilation of which
Qio is almost equal to the coefficient of respiration;
are all grounds for the assumption of a close connection
between assimilation and other protoplasmic functions.

We have mentioned repeatedly the reaction to abnormal
influences. It may be well to discuss this again at the
hand of the diagram represented in fig. 17. The hnes 1
and 2 show the cessation of the assimilation after more

or less time. This happened in exp. 66 and 69 after the
addi- don of KCN in exp. 92 after the addition of Ba CU.
A quick, and a successive slow, decrease (line 3) was also
found with KCN. Type 4 was very frequently found, a
rather strong retardation, followed by recovery. It mani-
fested itself in experiments with the three substances in-
vestigated. Type 5, retardation, passing into an increase,
resulted in experiments with KCN. Type 6 and type 7

give the experimental results with
diluted KCN and sugars. The
results obtained all show the
gradual transitions between very
strong- and almost immediate-
retardation and stimulation.

The types 4—7 were also
determined for the respiration.
In case of a stronger injury of
the assimilation, respiration be-
haves differently.

Another interesting fact was
that in the experiments with KCN
the effect did not depend on the
concentration of this substance.
A moderate retardation was ob-
tained with a strong concen-
tration, and a strong retardation another day with a weaker
one. Consequently, the incidental disposition of the alga
determines whether the effect will be strong or weak.

I had already drawn this diagram, when I was struck
by the great resemblance with a figure in Miss Irving's
work (1911; fig. 7, p. 1083). This diagram bears on the
influence of chloroform on respiration of cherry-laurel
leaves and young barley shoots. My types of curves are
to be found back there. The concentrations used are rather
high, and retarded the assimilation completely in her

experiments. It is, however, of great interest that we find
a similar mode of reaction for the respiration of these
plants and for the assimilation of Hormidium, while this
parallelism pardy applies to the respiration of this alga.

Shibata (1929) too, gives curves that are fundamentally
the same, and in which all my types can be found back.
He investigated the antagonistic action of electrolytes on
the respiration of Chlorella.

The deviations of some points on my temperature-
assimilation curve will also be very plausibly explained
by assuming the occurring of reaction-type 4, as a result
of the action of high or low temperature (cf. the following
paragraph).

So it seems that the diagram indicates a very general
reaction of the organism on a factor, uncommon to its
ordinary conditions of life.

E. An Attempt to Explain the too Large Deviations of
Some Determinations on the Influence of Temperature on
Photosynthesis from the Average, and a Few Remarks
on Optimum Curves,
We saw in the chapter on the influence of temperature
on assimilation (Chapter VII) that the deviations of the
individual determinations from the average curve were in
many cases larger than might be expected in connection
with the accuracy of the method. When discussing this
fact it was already pointed out that attempts would
be made later on to account for these variations with the
aid of other results. These results were obtained in the
Chapters VIII, IX and XI. It has been discussed in the
former paragraph that assimilation reacts in principle in
the same way to injurious influences, however different
they may be (addition of KCN, phenylurethane or Ba CL):
after the initial strong decrease of assimilation a recovery
of the function may start.

Now in the first place it strikes us while studying the
results of experiments on the influence of temperature,
when no high temperatures (30° and upwards) were investi-
gated in the experimental series, deviations did not appear.
This shows very distinctly from the experiments carried
out in May 1931 (table 8), in which high temperatures
were avoided, but also from exp. 38 and 43 (table 6).

An example of great variations is to be found in exp. 42.
At 19.5° the assimilation is 317, at 23.5° it is 513, at 31.5°
678, at 23.5 again 449, as we see much lower, in spite of
the difference in time of 2 h. 35 min. The last determination
in this experiment, also at 19.5° gives 312, in other words
the assimilation is again nearly as high as 5 h. 45 min.
ago; the assimilation that had been found much too low at
23.5°, has almost completely recovered at 19.5. This bears
a strong resemblance to the results obtained with poisons.

So I think I am able to explain this variation in the
determinations from the normal reaction to the injurious
influence which in this case had been the high temperature.

In exp. 41 we observe something similar, though a little
less pronounced. After a rise of temperature to 30° the
next determination at 22° is not higher than the one at
22°, booked 2 h. 45 min. earlier, while the following values
at 18° and 14° were higher than before. (The first deter-
mination at 18° is rather a little too low).

In experiments 39 and 40 the influence of a lowering in
temperature has been investigated. The first values of
these experiments are represented by the evidently lowest
points at 30° and 29° in fig. 11. This is easy to understand.
From the very cold windowsill the algae were suddenly
exposed to these high temperatures, so it is a matter of
course that the lowering action makes itself strongly felt now.

In exp. 40 the low temperature at 4° has also been
investigated. The next reading at 14° turned out 4 %
lower than the preceding one, in spite of a 5 hours' difference

in time. So a low temperature has apparently the same
influence as higher ones. This is a new base for the
generality of this reaction. There are, however, no obser-
vations as to the appearance of recovery after low tempera-
tures. So we see that the deviations from the average curve
can be explained from the physiological reaction to an
injurious influence.

Not only the variations of individual determinations,
but also the less steep course of the curve at higher
temperature (fall of Q^) can be explained by the physio-
logical reaction to injurious stimulants.

It may be mentioned here that, if my explanation
is true, this might be to a certain extent a support for
Blackman's classical theory about the optimum.

Blackman (1905) is of opinion that a slighter increase
of the rate of reaction at high temperature is a result of
the injurious action of this temperature, and that in this
way the optimum-curves, so often met with in physiology,
might be explained. Even before the optimum were reached,
a less strong rise of the curve might be caused by the
influence of this injurious action.

Therefore, the lower intensity of assimilation when
experimenting again at lower temperature might support
Blackman's opinion. But the difference between this
conception and my own results is, that in my experiments
the injury is followed by a recovery.

Neydel (1930), working with Cladophora, noted an imme-
diate recovery of the assimilation after the exposure to high
temperature. In my own experiments the recovery takes a
rather long time, but I believe this difference is not
essential.

A transition between Blackman's theory and the con-
ception of the real physiological optimum is apparent in
my results. The reversibility of the optimum curve seems
to be determined by the time wanted for the recovery.

I was not able to state a light-intensity optimum of
photosynthesis in intensities up to 8000 Lux with algae
cultivated in daylight. This is entirely in accordance with
the results of van den Honert. However, algae culti-
vated in weak artificial light, showed a slight decrease of
assimilation in high light intensity (fig. 9). In two other
cases the assimilation was even brought to a standstill, and
the absorption of oxygen began to predominate.

Finally, carbon dioxide did not show any influence in
high concentrations in van den Honert's experiments.
I can confirm this altogether. Up to a concentration of
1 % the curve is quite parallel with the abscissa and does
not show any tendency to fall.

In the fifth chapter, in which the relation between CO.,
concentration and assimilation has been established, the
peculiar Blackman curve seemed to disappear altogether,
when the reaction was retarded by the addition of narcotics,
and to show a logarithmic type, established by other
investigators (e.g. Harder). One may think here of a
slower chemical combination of the COg (or a product
derived from it) with other substances. Complications
might also be assumed. The question was put whether
the respiration with different COg concentrations had
perhaps become deviating, but this proved not to be the
case in a testing experiment. Whatever the explanation of
the COo concentration-assimilation curve after the addition of
urethane or antipyrin must be, the remarkable fact remains
that the Blackman curve has altogether disappeared.

The light intensity-assimilation curve of van den
Honert and the criticism on his method of illumination
led to the opinion that a better approach to the Blackman's

scheme might be possible. It is therefore of interest that
this did not prove to be so and the curve kept a same
logarithmic type with an improved experimental technique,
and did not show any tendency to approach a straight curve
with a sharp break.

The preceding considerations, in which the probability of
an indirect action of external factors was claimed on more
than one ground, makes it very unhkely that Blackman-
curves will often be met with. These curves sometimes
appear in incidental circumstances, such as the COo
concentration-assimilation curve of van den Honert.

A very simple, but at the same time very accurate
manometrical apparatus is described. Films of only a
single cell layer of the filamentous alga Hormidium
flaccidum or the sub-species H. nitens, served as experi-
mental objects.

A parallelism between assimilation and respiration was
repeatedly observed. Both processes can be stimulated
by the addition of highly diluted KCN or of sugar to
about the same degree. Not too strong a retardation of
photosynthesis, caused by higher concentrated KCN, by
Ba Clg or phenylurethane is accompanied by a decrease
of respiration. This parallelism disappears in case of a
strong retardation of assimiladon. When the retardation
is not too strong a recovery will follow. This occurs
simultaneously and in about the same measure for both
processes.

Temperature affects them both in a similar way; Q^
is at moderate temperatures rather constant: it is from
15° to 25° for the assimiladon 1.96, for the respiration 1.91.

The influence of various substances on assimiladon is
of equal strength, whether light or temperature is the
limiting factor, which for K C N is contrary to the results

Another fact found by Warburg in the case of
Chlorella, viz. the impossibility of retarding the assimilation
below the compensation-point of assimilation and respiration
by means of KCN, could not be confirmed with
Hormidium.

It will last a few minutes before the assimilation after
a period of darkness reaches a constant final speed. This
phenomenon is highly sensitive to temperature, and con-
sequently is not caused by photochemical induction
(Warburg). A chemical process must precede the beginning
of the assimilation. The Qio of this process is, determined
roughly, 2.2, and of about the same magnitude as that of
respiration and assimilation.

After a long period of darkness an initial increase of
photosynthesis, was found, but after half an hour it proved
to be fairly constant.

The facts mentioned led to the conclusion that the
assimilation is dependent on an internal factor. If this
internal factor is the extent of the available, active surface
— a supposition made by Briggs in order to explain
subnormal assimilation of young and starving plants —
then this factor is nevertheless directly dependent on other
active internal factors (as was a.o. proved by the recovery
of the function after the addition of toxic chemicals).

Respiration is increased during the exposure to hght.
It could be made plausible that this increase cannot be
due to a rise in temperature. The increase is partially
caused by the production of carbohydrates, but for the
most important part it is a result of the action of light on
the protoplasm.

Light, therefore, causes other protoplasmic reactions in
addition to photosynthetic. Consequently light may affect
the assimilatory process indirecdy.

The light intensity-assimilation curve showed, under
improved experimental conditions, the normal image of
a logarithmic curve. It does not at all approach the scheme
proposed by Blackman.

The Blackman-curve, which was found by van den
Honert for the influence of COg, disappears when the
assimilation is retarded by phenylurethane or antipyrin.

I started this investigation at the Botanical Laboratory
of Utrecht University and I should like to express my
grateful thanks to Professor F. A. F. C. Went for the
undiminished interest he took in my work up till the end.

By far the greater part of the investigation was carried
out at the Laboratory of Technical Botany at Delft and
I am very much obliged to Professor G. van Iterson
for his valuable help, wherefore I am thanking him
particularly.

F. Aereboe, Forsch, a. d. Gebiete d. Agrikult. phys. 16, 429, 1893.
J. E. van Amstel, Ree. trav. bot. néerl. 13, 1, 1916.
A. Arnold, Planta 13, 529, 1931.

u. Chem. N, F. 68, 500, 1899.
A. E. H. R. Boonstra, Planta 10, 108, 1930.
J. Borodin, Just's Bot. Jahresber. 4, Abt. II, 919, 1876; Mém.

acad. imp. sei. St. Pétersbourg, Vlle Sér. 28, No. 4, 1, 1881.
J. C. Bose, The Physiology of Photosynthesis. London 1924.
P. Boysen Jensen, Bot. Tidskr. 36, 219, 1918.

Phil. Journ. Sei. C. Bot. 12, 85, 1917.
R. Emerson, Journ. Gen. Physiol. 10, 469, 1927; Journ. Gen.

Physiol. 12, 609, 1929; Journ. Gen. Physiol. 12, 623, 1929.
A. J. Ewart, Journ. Linn. Soc. Bot. 31, 364, 1896; Journ. Linn.

Soc. Bot. 31, 554, 1897; Ann. of Bot. 12, 415, 1898.
L. Genevois, Biochem. Zeitschr. 186, 461, 1927.

F.nbsp;G. Gregory and F. J. Richards, Ann. of Bot, 43, 119, 1929.
R. Harder, Jahrb. f. wiss. Bot. 56, 254, 1915; Jahrb. f. wiss. Bot.

60, 531, 1921; Planta 11, 263, 1930.
M. Henrici, Verhandl. nat.-forsch. Ges. Basel, 30, 43, 1919.

T. H. van den Honert, Diss. Utrecht, 1928; Ree. trav. bot.
néerl. 27, 149, 1930.

N. Johansson, Svensk Bot. Tidskr. 17, 215, 1923; Svensk Bot.
Tidskr. 20, 107, 1926.

H.nbsp;Kniep, Internat. Rev. Hydrobiol. u. Hydrogr. 7, 1, 1914.
S. Kostytschew, Planta 13, 778, 1931.

1916; Biochem. Journ. 14, 267, 1920.
Tsi-Tung Li, Ann. of Bot. 43, 587, 1929.
A. Löwschin, Beih. Bot. Centralbl. 23, Abt. I, 54, 1908.
W. Lubimenko, Comp. rend. acad. sei. 143, 609, 1906.
H. Lundegiirdh, Svensk Bot. Tidskr. 15, 46, 1921; Der Kreislauf
der Kohlensäure in der Natur. Jena 1924; Biochem. Zeitschr.
154, 195, 1924; Flora 121, 273, 1927.
L. Maquenne and E. Demoussy, Comp. rend. acad. sei. 156,
506, 1913.

deuts. bot. Ges. 46, 383, 1928.
A. Meyer and N. T. Deleano, Zeitschr. f. Bot. 3, 657, 1911;
Zeitschr. f. Bot. 5, 225, 1913.

C.nbsp;Montfort and K. Neydel, Jahrb. f. wiss. Bot. 68, 801, 1928.
K. Neydel, Biochem. Zeitschr. 228, 451, 1930.

1, 1, 1918; Journ. Gen. Physiol. 1, 295, 1918.
E. Pantanelli, Jahrb. f. wiss. Bot. 39, 167, 1904; Ber. deuts. bot.

Ges. 32, 547, 1914.
A. Pascher, Die Süsswasserflora Deutschlands, Oesterreichs und
der Schweiz. H. 6: Chlorophyceae III. Jena, 1914.

H.nbsp;Plaetzer, Verhandl. phys. med. Ges. Würzburg, 45, 31, 1917.
L. Plantefol, Ann. sei. nat. Bot. lOe Ser. 9, 1, 1927.

Th. Schmucker, Biochem. Zeitschr. 195, 149, 1928.
M. Shibata, Sei. Reports Tohoku Imp. Univ. 4e Ser. 4, 431, 1929.
H. A. Spoehr, Photosynthesis, New York 1926.

Washington 1923.
W. Stiles, Photosynthesis, London 1925.
O. Stocker, Flora 121, 334, 1927.

O. Warburg, Biochem. Zeitschr. 100, 230, 1919; Biochem. Zeitschr.
103, 188, 1920; Biochem. Zeitschr. 119, 134, 1921; Naturwissen-
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R. Willstätter and A. Stoll, Untersuchungen über die Assimi-
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Bij de bestudeering van levensprocessen moet
steeds de mogelijke actieve invloed van het
organisme op het bestudeerde proces in het oog
worden gevat.

Het onderzoek van Smith, Dustman en Shull
houdt geen weerlegging in van de meening van
Schmucker, dat aan actieve (vitale) processen bij
de transpiratie van het blad een belangrijke rol
moet worden toegeschreven.

F. Smith, R. B. Dustman en Ch. A. Shull, Bot Gazette 91, 395,
1931. Th, Schmuckcr, /ahrb. ƒ. wiss. Bot. 68, 771, 1928,

De waarneming van Navez, dat de tempera-
tuur-coëfflciënten van ademhaling en geotropischen
reactie- en presentatietijd overeenstemmen, wijst
op een indirecten invloed van de temperatuur op
het geotropisch proces.

Wanneer de invloed van een factor op de
ontwikkehng van microörganismen onderzocht
wordt, mag niet volstaan worden met de bepaHng
van de „eindopbrengstquot;, maar moet het geheele
verloop van de ontwikkeling worden nagegaan.

Het is waarschijnhjk dat er zich tusschen den
primairen en den secundairen wand van het katoen-
haar een net van cellulose bevindt, waarvan de
draden in de richting van de as van het haar en
loodrecht op deze richting verloopen.

De veranderingen in de samenstelling van het
woud in den postglacialen tijd zijn in de eerste
plaats door wijzigingen van het klimaat veroorzaakt.

De vaststelling van den phylogenetischen samen-
hang van de soorten is wel een hulpmiddel, maar
niet het einddoel van de systematiek.

Aan het zoogenaamde compensatiepunt van
ademhaling en koolzuurassimilatie mag als oeco-
logische factor geen groote waarde worden
toegekend.

Het is niet mogelijk soorten van het geslacht
Fusarium te identificeeren naar hun gedrag tegen-
over groeivertragende stoffen (als malachiet-groen)
in den voedingsbodem, zooals door COONS en
Strong is voorgesteld.

G. h. Goons en M. G. Strong, Agcic. Exp. Stat. Mich. St. Coll.,
Techn. Ball. No. 115, 1931,

Wegens het zwak-parasitaire karakter van
Cytospora zijn cuhuurmaatregelen ter bestrijding
van Cytospora-ziekten van meer belang dan de
toepassing van directe bestrijdingsmiddelen.

De geleiding van een impuls door een zenuw
gaat met vermeerderd zuurstofverbruik gepaard.

De meening van Merker, dat het aan de opper-
vlakte verschijnen van aardwormen na een regenbui
door de afneming van het zuurstofgehalte van den
bodem veroorzaakt zou zijn, is door Focke
afdoende weerlegd.

mesh	diam. of wire (inches)	size of opening
24 x 24	0.0100	0.0317
30 x 30	0.0100	0.0233
35 x 35	0.0080	0.0206
40 x 40	0.0090	0.0160
45 x 45	0.0090	0.0132
50 x 50	0.0090	0.0110
55 x 55	0.0090	0.0092
60 x 60	0.0090	0.0077

Number of experiment	Date	[ Strain i 1	Tempera- ture	Oj evolution per hour	Difference between Oj evolution and CO-j ab- sorption per hour	Assimilatory Quotient O2/CO2
13	11-9-30	Pringsheim's	22.00	192	— 18	0.92
14	16-9-30	»	22.00	1068	1- 66	1,07
15	19-9-30	»	22.00	957	— 6	0.99
16	10-7-31	V. d. Honert's	21.50	610	28	1,05

Number of experiment	Date	Strain	Tempera- ture	O2 evolution per hour	Difference between Oj evolution and COj ab- sorption per hour	Assimilatory Quotient Oo/COj
13	11-9-30	Pringsheim's	22.00	192	— 18	0.92
14	16-9-30	gt;gt;	22.00	1068	66	1.07
15	19-9-30	tt	22.00	957	— 6	0.99
16	10-7-31	V. d. Honert's	21.50	610	28	1.05

Time	Light 1	Assimilation in mm^ per hour	Corrected CO2 conc. in 0.001 %	Corrected assimilation
11.50	darkness	-29
14.25	strong light	95	135 !	92
16.00	tt	105	173 j	100
17.10	If	76.5	50.5 !	72
19.00 !	»	64	37 1	59
20.45 1	gt;gt;	75	40.5 Î	68
22.00	tr	114	168	102

	Light intensity			1
Time	Transmitted	Voltage	Corrected	Assimilation
11.30	1000	2191/2	952	171
12.45	611	2221/4	609	136
14.15	415	22214	413	102
15.15	254	2221/2	254	66
16.20	2300	—	2300	211
17.05	1000	222V4	1004	171
18.20	0	—	—	-35
Exp. 26, 13-3-31. Pringsheim's strain. Temp. 20.00°.
10.25	1000	2211/4	988	493
11.05	611	222	611	351
11.50	415	222	415	271
12.55	254	2221/2	256	181
14.00	174	222	174	134
15.30	0	—	0	-35

Time	Light	Assimilation in mm® per hour	i Corrected CO2 conc. ; in 0.001 %	Corrected assimilation
~ 10.45	darkness	-20.5	—	_
13.45	strong light	108	100	—
15,10		146.5	136	90
16.15		121	55	74
17,20	»	97.5	19	59
19,15	It	65	10.5	39
20.50		167.5	367	100
22.00	quot;	152	132	90

Time	Temperature 1	Light	CO2 concentration	Assimilation
11.05	19.2°	darkness	_	-43
12.20	gt;f	strong light	425	124
15.10	tt	tgt;	90	87
16.50	22.8°	darkness	—	-62
18.00	Ji	strong light	397	193
20.10	tt	»	154	159
21.15	tt	ft	71	137

Time	Light intensity 1			Assimilation
Time	Transmitted	Voltage	Corrected	Assimilation
11.00	611	2201/4	613	1 350
11.40	335	220	335	213
12.35	174	2201/2	175	120
13.40 1	1000	221%	1024	465
14.25 :	2300	—	2300	539
15.30 ,	0	— i	0	-26

Time	Temperature	Voltage	Apparent assimilation respiration	Assimilation reduced to equal light intensity
11.35	10.00	220	93 17	110
13.55	16.00	22VU	95 30	122
16.15	22.00	2201/2	97 1- 43	139
18.25	28.00	2231/2	95 -r 59	146

10.25	14.00	2I8V4	132 t- 24	159
12.30	20.00	221	134 1 40	171
14.40	26.00	j 2I8V4	112 1 56	172
16.30	20.00	2181/4	128 1 40 1)	173
17.30	14.00 1	2221/4 1	' 166 i- 19 1	179
	Exp. 36,	2-6-31. Van den Honert's strain.
		Light intensity „140quot;.
11.25	20.00	2201/2	94 i 28	121
13.25	20.00	221	106 ) 28	132
15.45	20.00	221	106 28 1)	132

Time	1 j Temperature	Assimilation	Assimilation converted
15,55	20,00	420	100
16,45	24,00	542	129
17.25	28,00	736	175
19.50	31,00	928	221
20,40	34,00	1025 1	244
Exp.	38, 11-11-30. Conversion factor -		100 465
10.30	17.00	390	84
11.30	21.00	483	104
14,00	■25,00	646	139
14,55	29,00	818	176
16,35	21,00	499	107

Temperature	Qio determined	Qio corrected
5—15	2.84	2.77
10—20	2,30	2,24
15—25	2.02	1.96
20—30	1.93	1.87

Time	Temp,	Assim.	Q,o determ.
10.10	13,40	292
11.20	18,40	404	gt;) 2.18
13.15	23,40	636	)gt; 2,10
14.20	28,40	848	)
	Exp. 45,	19-5-31.
10.55	10,00	356	)
13,45	15.00	568	) 2,25
14,50	20,00	811	)[) 2.07
16,05	25,00	1177	)j 1.85
17.20	30,00	1504

Time	Tempe-	Respir-	Respir.
Time	rature	ation	converted
11.25	8.00	57	41
13.30	13.00	96	i 70
15.50	18.00	117	i 85
Exp. 49, 18-2-31. Conversion factor			io6 52
10.50	9.00	24	46
12.10	14.00	37	71
13.30	19.00	40.5	78
14.50	24.00	67	129
16.05	29.00	92	177
17.10	34.00	125	240
18.30	19.00	40	77
	Exp. 50, 5-3-31.
11.10 !	33.50	92	—
12.40 '	28.50	43	_
14.20	23.50	28.5	_
15.30	18.50	18	_
16.50	28.50	57	—

Temperature	Qi„ respiration determined	Qio respiration corrected	Qio assimilation corrected
10—20	2.06	2.09	2.24
15—25	1.87	1.91	1.96
20—30	1.80	1.85	1.87


	■quot; ! ■:.







	■gt;i • V

: -, -fm






■
	• l. ' ■ li ■ ■

Time	Temperature	Assimilation	Assimilation converted
10.40	16.00	360	79
11.55	12.00	233	51
13.50	12.00	226	50
15.05	8.00	154	34
16.00	4.50	±84	± 18
16.55	12.50	260	57
17.55	17.50	384	84
20.00	22.50	531	117

Solution	Time	Li trans- mitted	!ght intens voltage	ity corrected	Assimil- ation	Increase in per cent
normal	10.35	2300	_	(2300)	413
gt;f	11.15	611	220	611	295
ft	12.05	205	2211/2	210	130
tt mol. KCN	13.00	0	—	0	-29
tt mol. KCN	14.25	1000	—	(1040)	488	30
tt	15.15	205	223	215	158	18
tt	16.05	455	223	477	316	25
tt	16.50	1000	223	1048	478	27
tt	17.25	2300	—	(2400)	526	27
tt	18.30	0	—	0	-38.5	33

Solution		Light intensity			Assimil-	Increase
Solution	Time	trans- mitted	voltage	corrected	ation	in per cent
normal gt;t » —— mol. KCN 50000	10.15 11.00 11.45 12.40	2300 611 181 0	2221/2 223	2300 606 181 0	504 382 154 -42
normal gt;t » —— mol. KCN 50000	14.00	2300	—	2300	541	7
ft it tt	14.45 15.30 16.25	611 181 0	224 2241/2	621 185 0	408 165 -45.5	3 6 8

15.20	2300	—	—	766	1
16.10	0	—	—'	-63.5
17.30	2300	—	—	659	— 14 1
18.30	0	—	_	-57	— 10
19.30	2300	—	—	777	1
20.20	0	— i	—	-61	— 4

Solution	Time	Light intensity	Assimil- ation per hour	Increase or decrease in per cent
normal	11.35	2300	941
ti	12.30	0	— 51
^ n^ol. KCN 5000	13.40	2300	±512
gt;t	13.50	gt;gt;	±344
»	14.00	ft	±230
gt;t	14.10	tt	± 170
gt;gt;	14.20	tt	± 152	— 84
»	14.55	0	— 44	— 14

normal			with phenylurethane
Time	Assimil-	Respir-	Time	Assimil-	Respir-	Decrease in
	ation	ation		ation	ation	per cent
10.25	519		13.10		48	— 11
11.30		54	14.15	412		— 21
			14.45		52	— 4
			16.35	497		— 4
			17.40		51	— 6

13.50	318		— 49
14.40		22	— 17
15.55	453		— 21
16.30		21	— 21
17.25	524		— 15
18.25		24.5	— 8

normal	17.35	2300	—	—	1015
tt	18.25	0	—	—	-66
0.5 mol. Ba CI2	20.00	2300	—	i -	± 569	— 44
rr	20.45	0	—		-29	— 56
aqua dest.	22.15	2300	—	! —	640	— 37
»	23.00	0	—	1	-70	f 6
	Exp.	91, 25-6-31. Temp. 21.00°.
normal	1 17.15!	139	2211/4	139	158
i gt;»	18.25 !	0	—	0	-66
0.5 mol. Ba Clu	20,05	139	2231/4	143	127	— 22
ft	21.10	0	—	0	-29.5	— 48

Time	Duration in minutes	Exposure in minutes	Assimil- ation	Assim. in X minutes	Assim. reduced
13.55	66	6x 1	24	4.0	3.0
14.55	60	5x2	81	16.2	12.2
16.35	56	4x3	115.5	28.9	21.8
17.35	56	8x 0.5	11	1.4	1.1
19.55	66	6x 1.5	59.5	9.9	7.5

14.00	60	8x 1	16	2.0	1.4
15.00 I	54	5x2	46	9.2	6.0
16.45 i	54	10 X 0.5	4.5	0.5	0.3
17.35 1	48	4X3	67	16.8	11.0
19.40 1	48	5x 1.5	24	4.8	3.1

13.25	43	4x2	85.5	21.4	17.2
14.05	46	6x 1	57.5	9.6	7.8
15.40	' 48	8x 0.5	29.5	3.7	i 3.0
16.30	50	4x3	174	43.5	34.9
17.20	48	5x 1.5	93	18.6	14.9