Acknowledgements
This work was supported in part by the Undergraduate Research
Participation Program of the National Science Foundation. Grant
number GY-5878.
My thanks to the Faculty and Staff of the Hopkins Marine Station
for their generous assistance.
Abstract.
cpor uptake by populations of mixed phytoplankton are in-
vestigated by centrifugation and liquid scintillation techniques
Plankton in culture densities of 0.03 mg/ml to 0.41 mg/ml in
pDT solutions of 0.3 to 100 ppb show DDT concentrations (gm
DDT gm dry weight plankton) of 210ppb to 170 ppm. Concentration
factors comparing final DDT concentrations in cells with initial
solution concentrations are found between 700 and 13,000. Phyto-
plankton take up between 40 and 70% of the DDT in solution in
less than four hours and generally less than 15 minutes. Formalin
killed cells take up the same amount of DOT as live cells. Lugol
preservative killed cells take up less DOT than live cells.
Cells incubated in dark B.O.D. bottles take up the same amount of
DDT as cells incubated in the light. Less dense cultures con-
centrate DDT more than dense cultures. Plankton appear to con-
centrate DDT moee in dilute DDT solutions than in concentrated
solutions. Phyto plankton containing DDT removed to dilute DDT
solution retained 704 to 90% of their DDT after two hours.
Introduction
DDT (1,1,1 trichloro-2,2 bis (p-chlorophenyl) ethane) is
found to be a wold wide polutant and Rudl (1964) provides a
very excellent general backtround to the problem of pesticides in
ecosystems. This chlorinated hydrocarbon is found in birds of
the Antarctic (Woodwell, 1967) and is found to cause reproductive
failure in sea birds (Hickey and Anderson, 1968) as well as
behavior abnormalities in such animals as the Fiddler crabs
(Odum, Woodsell et. al..,1969). The pesticide is found to be
highly mobile (Risebrough, 11967) and is found to concentrate in
the food web (Woodwell, 1967).
Since marine phytoplankton is the base of this food web,
the effects of DDT on these important plants is important for the
world ecosystem. Butler (-) reports that DDT and other hydrocarbons
such as heptachlor, chlordane, aldrin, and toxaphene decreased
productivity of phytoplankton by 50-90% when incubated in 1 ppm
pesticide for four hours. Ukeles (1963) assesses growth of pure
cultures of marine phytoplankton and finds that some concentrations
in the range of 0.2 - 0.6 ppm retard growth of some species.
Wurster (1968) demonstrated that DDT inhibits photosynthesis and
reduces primary productivity by 50-90% in ranges of 150-100 ppb.
Some natural populations show DDT concentrations of 0.04 ppm
(Woodwell, 1967).
To better understand the effects of DOT on phytoplankton
growth and photosynthesis as well as on the food chain, the
processes involved in DT uptake by plankton need to be studied
and this is the purpose of this research project. Södergren (1968)
-2-
has conducted studies of C labelled DDT uptake by chlorella
but no studiesehaveubeen made on natural populations of phytoplankton.
The major question to be investigated is whether uptake is
active or passive, as this is very important especially in
consideration of the effects on the food chain. Accumulation
of substances by living phytoplankton is essentially a process of
absorption by metabolic processes, and adsorbtion by physical
and ion exchange phenomena. Death may increase or decrease uptake
as death may open more sights for uptake and does not necessarily
measure only adsorbtion (Cushing, 1968). Uptake of some radionuclides
shows light and temperature dependence (Gutknect, 1958). Mac
Isacc and Dugsdale, (1967) studied the kinetics of nitrate and
ammonia uptake by natural populations of marine phytoplankton
and suggest that it follows a Michaelis-Menton expression for
enzyme reactions. To test whether uptake is dependent on active
transport, metabolic poisons can be useditoestop metabolically linked
transport while diffusion uptake can continue. (Giese, 1968) Methods
are suggested by Wedemeyer (1965) to distinguish between adsorption
and absortion, and kinetics for diffusion and facilitated
diffusion uptake.
The tests conducted for this project were comparisons of
uptake by live cells and cells killed with Lugol fixative solution,
by live cells and cells killed with formalin, and by cells
exposed to light and cells kept in the dark. These tests are
only preliminary to the investigation of uptake and many experiments
need to be carried out for a fuller understanding of the problem.
There are many problems in investigations of DDT uptake
by plankton. Codistillation of DDT and H20 (Bowman, 1964)
adsorption of DDT to glass and other materials (Cole, 1961), and culture
and collection techniques are a few of these problems.
Materials and Methods
Phytoplankton for experiments were collected between 8:00 am
and 11:00 am in 10" phytoplankton nets 425 towed behind a
row boat at a depth of 15-20 meters. The collections were made
about 102 meters from the shore of Mussel Point of the Hopkins
Marine Station in Pacific Grove, California. Tows were made for
times from 10 minutes to one hour and placed in one pint mason
jars and kept in the dark in an ice filled styrofoam ice bucket during
subsequent tows and until use. These tows were diluted for use
or cultured as described in appendix. Each day samples of the
tows were taken and identified. Dry weight determinations were
made of cultures used in each experiment (Parson and Strickland,
1968).
C1 labelled DDT in ethanol (100 ppm, specific activity
19 mcmM) was micro-pipetted into a 50-100 ml millipore filtered
(0.45) sea water and aliquots from this stock were innoculated
into the cultures. The solution was always drawn up into the
pipette and flushed back into the bottle to assure that the
first aliquot did not lose more DDT through adsorption to glass
than subsequent aliquots..
Cells were killed for Experiments 1 and 2 and 4 with several
drops of Lugol's fixative.
Cells for Experiments 3 and 4 were killed with 2-3 ml of
56 formalin.
The dark tests in Experiments 5 and 6 were carried out in B.O.D.
bottles masked with two layers of plastic electrician's tape
and then wrapped in aluminum foil.
-2-
Incubations were made in either incubators with two 30
florescent lights 8-10 cm from the bottles or in a water bath
in a green house where natural light was available.
At appropriate intervals duplicate 10 ml aliquots of
culture were removed by volumetric pipette from cultures and
placed in glass 12 ml centrifuge tubes and rotated 75 cranks by
hand. Two ml samples of supernate were taken and the rest
discarded. 0.2 ml DMF was then added to the 0.3-0.5 ml loosely
packed pellet and poured into a scintillation vial. The tube
was then rinsed with 0.5 ml distilled water which was poured out
the same side of the tube as the pellet to prevent as much loss
of radioactivity as possible. Bray's scintillation fluid was used
for counting (Table X). Separate blanks were made for
supernate and pellet samphes as these count with different
efficiencies. For these blanks, the same procedure as above is
used on plankton from the culture which has not been exposed
to radioactivity. Counts were corrected for background and
pellet counts were corrected by subtracting the count of supernate
of equivalent volume. Concentration of DDT in plankton is
described as grams DDT per gram dry weight phytoplankton. Con-
centration of DOT in water is described as grams DDT per gram
water. Weights of DDT are determined by multiplying disintegrations
per minute by a factor of the specific activity. The factor
used for the experiments (all except Experiment 7) is
8.36 x 10-12 gms DDT/dpm. In Experiment 7 the factor is
62.5 x 10-12 gms DDT/dpm as the radioactivite DDT was diluted
by a factor of 7.5 with cold DDT, also in ethanol.
Tests for retention of DDT were accomplished by taking
10 ml samples of culture, centrifuging, removing 9.5 ml of
-3.
supernate and replacing it with filtered sea water. These
tubes were then allowed to rest in the water bath for the appropriate
interval and collected in the usual manner.
Calculations of initial solution concentrations were made
either by reading 2 ml of the water blank (Experiments 1 and 3)
or by taking total counts in a 10 ml aliquot (Experiements 5 and 6).
Five times the supernate count is added to the pellet count and
and this total is divided by 10 to get the counts perml which gives
the amount of DDT present. In the case of Experiment 7, the
amount is calculated by the amount added. O.75 ml 50 ppm DDT
in ethanol and 50 100 ppm C14DDT were added to 30 ml H20.
10 ml was added to 100 ml of culture to give 100 ppb and 5 was
added to give 50 ppb.
Results
Tahle I gives data for comparison of c' por uptake by Lugol
presergative killed and untreated cells. The initial solution con-
centration is calculated from the dpm of 2 ml of the water con-
trol blank whith is 0.45u millipore filtered sea water of an equal
volume as the test cultures.
Column Felapsed time" indicates the elapsed time from the in-
troduction of C DDT. Under each heading, the dpm of 0;5ml contain-
ing the pellet is listed in pairs and the next column gives the
average of the two. Further information is listed below the table.
The amount of DOT is proportional to the dpm and final uptake
figures are calculated. A concentration factor is determined com-
paring the initial solution concentration to the final phyto-
plankton concentration.
This data is also presented in fugure 1.
The results of this experiment suggested that live cells take
up between six and seven times as much DDT as Lugol preservative
killed cells. However, the fact that the highest value was only
three times the background count suggested that the results were
not definitive. The loss of 504 of the DDT through codistillation
did not reduce the counts of the live cells which suggested that
once DDT is taken up, it remained bound. As some counts were more
than double that of their duplicate, the figures can not be taken
to be anv more accurate than a factor of two.
-2-
Table II from experiment 2 gives data for comparison of
ppr uptake by Lugol preservative killed cells and untreated
cells incubated in a solution of 1.7 ppb DDT. The initial sol-
ution concentration is calculated from the dpm of 2 ml of the water
control blank which is O.45u millipore filtered sea water of an
equal volume as the test cultures.
Column under "elapsed time" indicates the time elapsed
since the introduction of CDDT. Under each heading, the dpm
of 0.5 ml containing the pellet is listed im pairs. The column
"Cor" gives the corrected value of the pellet. The counts of a
volume of supernate aqual to the volume of the pellet is sub-
tracted from the pellet reading. The column "g" is the percent
of the solution (in ten ml - 5 x supernate plus pellet count) that
is found in the pellet.
The amount of DDT is proportional to the dpm and final uptake
figures are calculated. A concentration factor is determined com-
paring initial solutin concentration and final phytoplankton con-
centration.
This data is presented in figure 1.
This experiment gave the same conclusion as experiment 1,
namely that live cells take up more DDT than Lugol preservative
killed cells, by a factor of 20. The results here were much more
reliable as the counts were considerably above background. Even
so, these figues are probably good within a factor of 1.5 - 2.
both experiments showed concentration by live cells of DDT on the
order of 1,000 fold on a dry weight basis.
-3-
These experiments suggested that uptake may be involved with
the life processes of the cells.
Experiment 3, Table III compares the uptake of C DDT by form-
alin kelled and untreated cells. The first column indicates the
slapsed time from the introduction of C DDT. The second column is
the dpm in 2 ml of water control which is the same volume of fil-
tered sea water as volume of the cultures being tested. The in-
itial concentration and codistillation data are determined from
this control.
The other data under four colums are as follows. "Pel" is
the dom of the pellet in a volume of supernate denoted below. The
colum labelled "Sup" is the dpm of 2 ml of supernate. The colum
under "Cor" is the corrected value of the pellet. The counts of
a volume of supenate equal to that of the pellet is subtracted
from the pellet counts to give a corrected pellet count. The column
under "ø" is the percent of DDT in solution taken up by the plank-
ton. The total counts are determined by multiplying the supernate
counts by 5 and adding the corrected pellet count.
The dpm is proportional to the amount of DDT in the cells and
conversions have been made below the table for final concentra¬
tions in the cells. These are compared tothe initial solution con-
centration by the concentration factor.
This data is presented in figure 2
The counts of this experiment were well above background.
They showed that live and formalin killed cells took up the DDT
approximately in the same amounts. A concentration factor of 1.3 x
10 was found. This suggested that the life processes were not in-
volved. Examination of the appendix shows that this is the only
-4.
phytoplankton used which was cultured. Perhaps these cells were
unhealthy in the cultures. Table IV, experiment 4, however, com-
paring alive Lugol preservative killed, and formalin killed cells
of a fresh tow showed that the Lugol's again took up considerably
less DDT than the other two which were approximately the same.
The values of table IV are given as dpm in 0.5 ml pellet. From
this it was assumed that experiment 3 was valid.
Experiment 5, Table V, and experiment 6, Table VI, compare
uptake of CDDT by cells incubated in the light and in the dark,
the former in 0.3 ppb and the latter in 1.2 ppb DDT solution.
The data in the four columns under the headings is arranged
as follows: "Sup" is the dpm of 2 ml supernate. "Pel"is the dpm
of the pellet in a volume of supernate denoted below it. "Cor"
is the corrected value of the pellet. The counts of a volume of
supernate volume equal to that of the pellet is subtracted to
give counts in the pellet. The "ø" is the percentage of DDT taken
from solution by the plankton. It is determined from the ratio of
the corrected pellet count to the total count (5 x supernate count
plus the corrected pellet count).
The dpm are proportional to the amount of DDT in the cells and
conversions have been made for final concentrations in the cells.
These are compared to the initial solution concentrations by the con-
centration factor.
This data is presented in figure 3.
Experiments 5 and 6 indicate that uptake was not related to
the photosynthetic process which it inhibits. The initial concen-
trations for these experiments were determined by taking a reading
of the stock solution and calculated from this according to di-
lution. The actual values may have been below thecalculated values
because of adsorbtion to glass. Experiment 5 had counts which were
close to background and were therefore less accurate than those of
experiment 6 which shows high counts. These cells show concentration
factors of 10 and 10 on a dry weight basis.
Experiment 7, Table VII, compares the uptake of plankton in-
cubated in C DDT at concentrations of 50 ppb and 100ppb. These are
the concentrations wheih Wurster found to inhibit photosynthesis.
The data in the four columns under the headings is arranged
as denoted below: "Pel" is the dpm of the pellet."Sup" is the dpm
of 2 ml of supernate. "Cor" is the corrected walue of the pellet.
is the percentage of DDT taken from the solution by the cells.
The plankton concentrated theDDT on the order of 2,000 times on a
dry weight basis giving concentratinns in the cells of between
100ppm and 200 ppn.
Experiment 8, Table VIII, investigates the retention of DDI
by cells placed in a CHDT reduced environment.
The second column of Table VIII shows the percentage of orig.
inal counts retained in the cells after the elapsed time im the
clean environment.
This experiment showed that 70% to 90% of the DDT taken up
remained in the cells when placed in a radioactive environment con-
taining 1/20th of the concentration (9.5 out of 10 ml were changed).
The retention may actuallty have been a little higher as probably
some cells were lost in the centrifuge process.
Table IX shows the data for ølive cells, delineates the
experiment number from which they were taken, cell densities,accord-
ing to dry weight, initial solution concentrations, and final con-
centrations in the fells. The concentration factor compares the last
two numbers. "Percent of supernate" shows the amount of DDT taken ur
y the phytoplankton from the solution.
Plankton showed uptake of between 306 and 704 of the DDT in
solution in less than four hours and mostly in less than 15 minutes.
Experiment 6 which was run for 13 hours showedthe greatest percentage
of uptake. Figure 3 showedthe small increase in the amount taken
up after four hours. Comparison of experiments 2,3, and 6 suggested
that concentration factors depended on cell densities with less
dense cultures concentrating DDT more. Experiment 7 suggested that,
very generally, more concentration occurs in less concentrated (DDT)
solutions. The 50 ppb cultures showed a slightly higher concentration
factor than 100 ppb where culture densities were the same. This
dependence on DDT solution concentration is further suggested by
experiment 3 in comparison which experiment 7. Exact relationships
can not be established form this data.
Discussion
The results are confusing with regard to the difference
between Lugol killed and formalin killed cells. It does not
clarify whether uptake is active or passive. Uptake is thought
to be dependent on lipid concentration which is 6% dry weight in
mixed plankton (Parsons and Strickland, 1968). Perhaps the Lugol
solution attacks and alters the lipid in the cell. This is
suggested by the fact that Lodine, which is a constituent of
Bone tCstto es,
Lugol's oxidizes the unsaturated double bonds and this may alter
the fat content, whereas formalin probably does not affect the
lipid.
Södergren, in his study of C1pDr uptake by Chlorella,
found that uptake in batch cultures by live cells and by cells
killed with mercuric chloride is the same. This corresponds to
the experiments with formalin killed cells.
Wuster found that photosynthetic inhibition in a given
solution concentration depends on cell density. These results
are consistent with findings here that less dense cultures
concentrate DDT more than dense cultures.
The dry weight concentrations demonstrated by these
phytoplankton suggest that this uptake would be of ecological
significance especially as concentration factors are higher in
low density low concentration situations which are present in the
oceans. On a wet weight basis (assuming 1% of wet weight is
dry weight) these figures suggest that a 10 fold concentration
of DDT is possible in dilute conditions. Concentrations of this
magnitude would be of ecological significance for the food web as
vast quantities of phytoplankton are consumed by herbevores
-2-
which could further concentrate the substance.
Certainly many more of the factors determining the uptake
are yet to be determined. Such tests would include examination
of adsorption to silicious shells of diatoms without soft
parts, and tests with metabolic poisons to determine if uptake
is dependent on metabolic activity. Sodergren found that con-
tinuous culture methods (Meyers and Clark, 1944) are much more
reliable than batch cultures to measure uptake as batch cultures
are rarely consistent in many of the parameters which determine
algal activity.
Table I
Duplicate determination and average dpm from filtered sea water
control, aliye, and Lugol preservative killed plankton incubated
in 0.3 ppb DDT solution. Experiment 1.
DPM
Elapsed
Lugol
Alive
Water
time
Pellet
Pellet
control
Hours
0.5 ml. avg.
0.5 ml
avg.
2 ml avg.
10
64
0.25
44
16
54
22
71
84
66
13
39
0.50
64
44
0.75
2
80
41
48
2.0
78
18
19
23
104
4.5
103
15
22
37
2
102
Background - 36
Initial solution concentration 0.3 ppb. Percent remaining after co-
distillation - 528.
Dry weight. O.41 mg/ml.
Final alive concentration - 210 ppb. Concentration factor - 700.
Final Lugol concentration - 31 ppb. Concentration factor - 100.
Table II
Experiment 2. Duplicate determination of dpm from filtered sea
water control, alive,and Lugol preservative killed plankton in-
cubated in 7.7 ppb C DDT solution. Corrected pellet dpm and per-
cent of DDT taken up from solution.
DPM
Elapsed
Water
Alive
Lugol
time
Pel. Sup. Cor. %
hours
control Pel. Sup. Cor.
.5ml 2ml
.5ml 2m1
2 ml
411
521
0.1
541
86
411
401
761
26
70
46
729
881
165
341
168
801
2.0
16
619. 42
721
Background 29. Above gounts have had background subtracted.
Dry weight 0.41 x 10-2 gm/ml.
Initial solution concentration - 1.7 ppb. Amount remaining after
codistillation - 89%.
Final Alive concentration - 1.42 ppm. Concentrarion factor - 10
Final Lugol concentration - less than control.
Table III
Experiment 3. Duplicate determination of dpm from filtered sea
water control, alive and formalin killed plankton incubated in
1.7 ppb C14DDT solution. Corrected pellet dpm and percent of DDr
taken up from solution.
DPM
Elapsed
Formalin
Water
Alive
time
Sup
4
Pel
Cor
Sup
Pel
Cor
Hours
control
.2 ml 2 ml
.2 ml 2 ml
2 ml
46
680
46
252
70
650
182
632
.08
282
108
916
940
922
933
176
38
620
600
206
159
719
73.
252
.41
672
690
625
183
650
225
646
65.
66.
192
222
63
232
.74
614
630
1670* 202 1650
165
545
524
34
212
190
586
230
605
1.16
556
580
240
214
550
529
170
623
42
643
640
226
765
2.08
218
705
708
206
725
725
170
Background - 27
Dry weight. .03 mg/ml.
Initial solution concentration - 1.2 ppb. Concentration Tactor
1.) K 10
Final alive concentration - 16 ppm. Concentration factor - 1.3 x 10".
Final Formalin concentration - 16 ppm. Concentration factor -
1.3x 10
Percent remaining after codistillation - 78%.
* Point rejected for extreme deviation from duplicate.
Table IV
Experiment 4. Dpm from 0.5 ml pellet of alive, Lugol preservative
killed, and formalin killed plankton after incubation for 1 hour.
Elapsed time
Hours
Alive
Formalin
Lugol
Pellet
Pellet
Pellet
0.5 ml
0.5 ml
0.5 ml
807
Background - 28.
Table V
Experiment 5. Duplicate determination of dpm from Phytoplankton
incubated in light and darkness in O.3 ppb CDDT solution. Cor-
rected pellet and percent of DDT taken up from solution.
DPM
Elapsed
Dark
Light
time
Sup
Pel
Cor
Sup
Cor
Pel
Hours
.3 ml 2 ml
.3 ml 2 ml
109 50
50
102
134
117
0.08
128
48
113
156 50
154
165
162
0.75
55
28
166
165
128
144 46
29
19
142
135
1.00
38
170
158
34
22
62
24
137
142
154
155
2.00
14
164
354
152
14
154
142
166
168
27
3.00
143
20
170
Background - 32
Stock solution count - 1,780. Diluted 1/25. Initial concentration-
3 ppb.
Cell density -.06 x 10.gms/ml.
Final congentration for both - 2 ppm. Concentration factor.
7 x 10.
Table VI.
Experiment 6. Duplicate determination of dpm from phytoplankton
incubated in light and darkness in 1.2 ppb C+ DDT solution. Cor-
rected pellet and percent of DDT taken up from solution.
DPM
Elapsed
Dark
Light
time
Sup
4
Pel
Cor
Pel
Sup
Cor
Hours
.3ml 2ml
.3 ml 2m1
37
634
660
41
534
191
565
202
0.08
690
174
780
136
173
872
56
893
53
1140
976
0.5
176
124
85:
57
924
153
894
132
1025
72
98.
1.05
900
114
1120
112
55
1120
186
169
870
953
957
54
1.5
835
830
117
127
850 61
98.
940
104
765
2.5
845
1
620
129
80 1011 71
960
1230
133 1267
13.0
108
1340
Background - 29. The above counts have had the background subtracted.
Dry weight -.2 mg/ml.
Initial concentration - 1.24 ppb.
Final concentration: Light - 5.3 ppm. Concentration factor - 3x 10"
Dark - 4.2 ppm.
Tahle VII.
Experiment 7. Duplicate determination of dpm from phtoplankton
incubated for eight hours in 50 ppb and 100 ppb C DDT solutions.
Corrected pellet and percent of DDT taken up from solution.
DPM
Elapsed
100 ppb
50 ppb
time
Sup Cor %
%
Sup
Pel
Pel
Cor
Hours
144 1011
805
102
101
79
8.0
262 1010
1036
180
36
530
512
Background - 30
Dry Weight. O.037 mg/ml.
Initial solution concentration-50 ppb.
Final concentration-109 ppm. Concentration factors - 2 x 102.
Initial solution concentration - 100 ppb.
Final concentration-170 ppm. Concentration factor - 1.7 x 10°.
Counts must be multiplied by 7.5 to correct for dilution of
radioactive DDT with non-radioactive DDT.
Table VIII.
14
Experiment 8. Percent of C DDT retained when phytoplankton
are removed to DDT solutions 1/20th the concentration of the
solution in which the DDT was taken-up.
DPM
Elapsed
8 of
Trial 2
time
Trial
Sup
6
Pel
Sup
%
Pel
orig.
Cor
Cor
Hours
3ml
2m1
count
.3ml 2m1
(1) (2)
960
80
1230
66
1011
72
133 1207
1087
1340
42
807
907 78
1080
987
54
97 76
1140
906
1210 88
1070
1.00
109 96 1220
1098 92
32
1360
980
40
83
1020
915
961
2.00
916
78
90 76
23
915
925
Background - 27. The above counts are corrected for background.
Dry weight -.2 x 10-2 gm/ml.
Retention - 70%- 90% of counts.
Table IX.
Cumulative Data for Alive Cells
Conc.
Final
Initial
Exp'ment Dr
Factor
Conc.
Conc.
Weight
ppb
ppm
gms/m
700
0.41 x 10-3
.210
0.3
825
1.42
0.41 x 10-3
1.7
13000
16
1.2
0.03 x 10-3
7000
2.0
0.06 x 10"
0.3
4000
0.2 x 10-3
5.0
1.2
2100
0.04 x 10-
109
1700
100
170
% of
Supernate
42-46
42
50
70
Table (
Counting Procedure
Contents of Bray's Scintillation fluid.
900 ml Dioxane
100 ml Methanol
20 ml Ethylene Glycol
g Naphthalene
60
PPO
g POPOP
.2
Machine
Nuclear Chicago Unilux II Two Channel Scintillation Counter.
Machine settings
ChannelE
Channel. A
9.9
2.2
Jpper discriminator
0.5
Lower discriminator
E 200
D 900
Attenuator
C
Figure 1
Comparison of C DDT Uptake
by Lugol killed and Untreated Cells
Figure 1 shows data from Tables IV and V. It shows radioactive
disintegration per minute emitted from samples of untreated (open
point) and Lugol preservative killed cells (solid point). One
set of points corresponds to cells incubated at 0.3 ppb for
4.5 hours, and the other to cells incubated at 1.7 ppb for
2 hours. DDT contained in the cells is proportional to the dpm.
The final point for untreated cells at 1.7 ppb corresponds to
1.5 ppm on a dry weight basis. The final point for the 0.3 ppb
incubation corresponds to 20 ppm on a dry weight basis.
O0(
DPM
800
60C
40
200
65
—
—

ELAPSED TIME
HOURS
8
Figure 2
Comparison of C1 Uptake by
Formalin killed and Untreated Cells
Figure 2 shows data from Table VI. It shows radioactive
disintegration per minute emitted from samples of untreated and
formalin killed phytoplankton incubated in 1.2 ppb DDT for two
hours. Two samples of untreated cells (open point) and two of
killed cells (solid point) are shown. Both appear to accumulate
the same amounts of DDT. The amount of DDT in the plankton is
proportional to the dpm. The average final concentration is
16 ppm.
00
DPM
80
60
400
200
10
05
ELAPSED TIME

HOURS
20
Figure 3.
Comparison of C14DDT Uptake by
Cells Incubated in Light and Dark.
Figure 3 shows data from Tables VIII and IX. Points correspond
to dpm of cells incubated in 1.2 ppb for 13 hours and 0.3 ppb for
3 hours. The amount of DDT in the cells is proportional to the
dpm. The final concentrations of DDT in the cells on a dry
weight basis is 2 ppm for 0.3 ppb and 4 ppm for 1.2 ppb.
0
O

I
S
L
0
O
O

4
A
Appendix
Culture and Incubation Data
This table providss information concerning collection and use of
of phytoplankton in the experiments. Column one gives experiment number
and a short indication of the nature of the experiment. This column also
provides the date of collection. Column two gives the concentration of
CDDT used in the experiment. Column three indicates mode of collection
The collction labelled "cultured" indicates that a tow was placed in
culture media (follows) and used subsequently. The "Depth" colum indi-
cates the depth at which the collection was made. The "Location"
column indicates location at which collections were made. The buoy re-
ferred to is the bell buoy approximately 1,000 meters off the shore
of Mussell Point of Hopkins Marine Station in Pacific Grove, California.
The "Age" column gives approximate age of the collection when th experi-
mentwas started. "Treatment" explains how the tow was diluted. The filter-
ed water referred to is filtered with 0.45 millipore filter. "Contain-
er" indicates th vessels used for experimental incubations. "Incubation"
describes the temperature and light conditions of incubation. Foot candle
measuremnts were made with a light meter. The incubator had two
flourescent light bulbs 8 or 10 cm. from the cultures. "Constitution"
indicates theimajor plankton types present in the collection ad identi-
fied by Dr. Isabella Abbott of the Hopkins Marine Station.
ond a
Culture and Incubation Data
Depth
Collection
Initial
Experiment
Date
Conc.
meters
ppb
Description
power skiff
15
0.3
May 22
Alive/Lugol
power skiff* 15
1.7
May 22
Alive/Lugol
cultured
1.2
May 31
Alive/Formalin
row boat
May
Alive/Lugol/Form.
row boat
0.3
May 26
Light/Dark
1.24
row boat
May 28
Light/Dark
50-100
row boat
May 28
High concentration
row boat
May 28
Retention
* Not collected by author.
15
15
10-15
10-15
Location
100 yds of
bell bouy
100 yds of
bell bouy
150 yds of
bell bouy
150 yds of
bell bouy
150 yds of
bell bouy
150 yds of
bell bouy
150 yds of
bell bouy
(con't
Age
hours
Treatment
tow diluted
250 ml tow
850 ml filtered
H20
tow diluted
250 ml tow
850 ml filtered
H20
culture
Chot diluted
tow diluted
400 ml tow
350 ml filtered
H20
tow diluted
250 ml tow
750 ml filtered
H20
tow diluted
250 ml tow
750 ml filtered
H,0
tow diluted
250 ml tow
750 ml filtered
120
Container
250 ml
Erlenmeyer flask
aluminum foil
cover
250 ml
Erlenmeyer flask
aluminum foil
cover
125 ml B.O.D.
50 ml in
125 ml
Erlenmeyer flask
500 ml in
100 ml B.O.D.
500 ml in
100 ml B.O.D.
125 ml B.O.D.
500 ml in
100 ml B.O.D.
Incubation
130 C
water bath
130 C
water bath
14° C
incubator
13° C
water bath
3000 foot¬
candles
130 C
water bath
1100 foot -
candles
140
incubator
14°
incubator
13
water bath
Constitution
Rhizosolenia alta (2 spp)
Chaetoceros (2 spp)
Nitzchia pacifica
Rhizosolenia alta (2 spp)
Chaetoceros (2 spp)
Nitzchia pacifica
Rhizosolenia alta
Chaetoceros
Nitzchia pacifica
Skeletonemia
Chaetochos
Rhizosolenia alta
R. hebetata
Chaetoceros (2 spp)
Nitzchia pacifica
Dinoflagellates
Rhizosolenia alta
Chaetoceros
Nitzchia pacifica
Skeletonemia
Rhizosolenia alta
Chaetoceros
Nitzchia pacifica
Skeletonemia
Rhizosolenia alta
Chaetoceros
Nitzchia pacifica
Skeletonemia
Algal Culturing Media
IR Food Chain Resarch Group
Sea Water - filtered 1-3
Nitrate 500 ug-at N/1
Phosphate 50 ug-at P/1
Silicate 250 ug-at Si/l.
Acid to neutralize silicate to ph 7.
Minor trace metals
ecl3.6H,0 5 X 102 gm/1
Mgla,N0,-2 H20 2.5 x 10-5gm/1
Sodium ethylenediaminetetracetate 5 x 10-gm/1.
Vitamins
1077 gm/1
3iotin 10 gm/1
Thiamine hydrochloride 10 gm/1
Tris Buffer 7.9-8.1 1 m1/1
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