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EFFECT OF SUSPENDED SEDIMENT ON THE
AVAILABILITY OF ADSORBED PHOSPHOROUS AND IRON
TO A GREEN ALGA

Thesis for the Degree of M. S.
MICHIGAN STATE UNIVERSITY
GEORGE CI KANDLER, JR.
1976

IX

r""*~

ABSTRACT

EFFECT OF SUSPENDED SEDIMENT ON THE
AVAILABILITY OF ADSORBED PHOSPHOROUS
AND IRON TO A GREEN ALGA

By

George C. Kandler, Jr.

This research concerns several important algal-nutrient-sediment
relationships. Scenedesmus guadricauda, a planktonic green alga, was
cultured with phosphorous being supplied in soluble and sediment-adsorbed
forms. Kaolinite clay was used to typify suspended sediment.
Phosphorous-kaolinite was prepared through a series of ion exchange
reactions. Growth chambers were specially designed for the experiments.

Scenedesmus guadricauda utilizes adsorbed phosphorus both
indirectly and directly from the clay particles. Algae grew extremely
well with P-clay as the only source of phosphorous. Besides making
phosphorous available to phytoplankton, suspended sediment also increases
the availability of iron when the pH of the medium is below 7.5, a con-

dition which normally causes iron to precipitate as ferric hydroxide.

EFFECT OF SUSPENDED SEDIMENT ON THE
AVAILABILITY OF ADSORBED PHOSPHOROUS
AND IRON TO A GREEN ALGA

By

George C. Kandler, Jr.

A THESIS

Submitted to
Michigan State University
in partial fulfillment of the requirements
for the degree of

MASTER OF SCIENCE

Department of Fisheries and Wildlife

1976

ACKNOWLEDGMENTS

I would like to thank the members of my graduate committee,
Dr. N. Kevern, Dr. E. Roelofs and Dr. B. Knezek, for their valuable
advice during the course of this research. I would also like to extend
my thanks to Dr. J. Gill for his help in the statistical analysis of
the data.

This research was supported by funds from the Michigan State

University Agricultural Experiment Station.

ii

TABLE OF CONTENTS

LIST OF TABLES AND FIGURES .

INTRODUCTION
METHODS AND PROCEDURES

Culture Conditions . .

Preparation of P- Kaolinite

Experimental Design .

Measurement of pH and Phosphorous Concentration
Measurement of Algal Growth.

RESULTS

Densities of Algal Cultures . .

Statistical Analysis of Growth Curves

Soluble Phosphorous Concentrations .
Relationship of pH to Nutrient Availability
Effect of Various Iron Sources on Algal Growth

DISCUSSION
LITERATURE CITED

Page

iv

LIST OF TABLES AND FIGURES

Table
l. Composition of Modified Rodhe VIII Culture Medium

Figure

l. Photographs<rfthe growth enclosure containing culture
flasks (A) and experimental chambers (B).

2. Photographs of the experimental chambers showing construction
(A,B), separation of algal cells and clay particles by the
partition (C) and the three experimental treatments (D)

3. Photographs of Scenedesmus guadricauda (450X) in culture
media free of clay (A) and containing clay (B)

4. Mean cell densities (: one standard error) of algal cultures
over the 16-day experimental period . . . . .

5. Mean concentrations and standard errors of soluble phos-
phorous in algal cultures over the l6-day experimental
period . . . . .

6. Ranges of pH values of algal cultures over the l6-day
experimental period . . . . . . .

7. Mean cell densities and ranges of algal cultures treated
with various iron sources over a 20-day period

8. Mean concentration and ranges of soluble phosphorous in
algal cultures treated with various iron sources over
a 20-day period .

9. Ranges of pH values of algal cultures treated with various
iron sources over a 20-day period .

iv

Page

15

18

22

24

27

29

32

INTRODUCTION

Sediment transported in runoff water is the most significant
source of suspended and deposited sediment in aquatic systems.
Suspended solid loads delivered to streams and lakes as sediment in
surface runoff are equivalent by weight to more than 700 times the
load from sewage (Johnson and Moldenhauer, l97D)._ The suspended
solid load carries with it a vast array of nutrients which are
ultimately deposited in lakes. Many investigators have shown that
surface drainage, especially from agricultural lands, contains high
amounts of phosphorous (Bernhardt gt_gl,, 1970; Engelbrecht and
Morgan, 1961; Gilchrist and Gillingham, l970; Keup, l968; Romkens
and Nelson, l974). Increased concentrations of inorganic phosphate
entering a natural water system are extremely important in that
phosphorous is a critical nutrient for algae and is most often the
element limiting algal growth. After the sediments are deposited,
they take on a very important role in regulating the phosphorous
cycle in natural water systems. The phosphates in the sediments
may be capable of supporting algal growth for some time even if all
inputs of phosphates to the system are eliminated. Sediments are
frequently capable of sorbing large amounts of added inorganic
phosphorous (MacPherson et;al,, 1958; Harter, l968; Shukla gt_al,,
197l; Williams gt_al,, l970). The phosphorous may be held

temporarily and subsequently released to growing macrophytes and

l

algae. PomFoy gt_al, (1965) have shown that the release of
exchangeable sediment ph05phate to the water is of sufficient
magnitude to sustain the growth of phytoplankton. These workers
also suggested that the development of algal blooms in many natural
waters depend upon phosphates from the sediments. Several investi-
gators have obtained excellent growth of Scenedesmus spp. in
cultures where mud was the only source of phOSphorous (Golterman
gt_§1,, 1969; Chiou and Boyd, 1974).

In well oxygenated lakes, there exists an oxidized microzone
at the sediment-water interface which acts as a barrier against
diffusion of phOSphates from the mud to the water (Mortimer, 1941).
Breakdown of the microzone is brought about by turbulence in the
waters which promotes particle suSpension. McKee et_al. (1970)
have found that wind induced currents, density currents and wave
action are the principal mechanisms for suspending and transporting
sediments within a lake.

Suspension of sediments increases their effect on the over-
lying water. Under completely mixed conditions, relatively rapid
release of phosphate occurs under both anaerobic and aerobic condi-
tions from a variety of lake sediments (Lee, 1970). Zicker gt_al,
(1965) found that agitation of the sediments by turbulence almost
doubles the rate of phosphate release from the sediments as com-
pared to undisturbed systems. These results point to the
importance of mixing processes in determining the relative flux

of phosphorous from the sediments. Large amounts of sediments are

mixed into the overlying waters under severe conditions of turbu-
lence and runoff.

The purpose of this research was to determine if algae
cultured in suspension with clay particles were able to directly
utilize sediment-adsorbed phosphorous. Growth comparisons were
made among cultures containing soluble phosphate and cultures
containing suspended clay particles as the only source of phosphate.
Although not an initial consideration, this research has also

shown that suspended sediments assist in the availability of iron.

METHODS AND PROCEDURES

Culture Conditions

 

All glassware used in the experiments was washed with
sodium-dichromate-sulfuric acid solution and rinsed thoroughly
with deionized water to assure the absence of phosphorous. To
prevent bacteria from contaminating the algal cultures, sterile
serological pipettes were used for all algal innoculations, trans-
fers and sampling. Nutrient media and culture flasks were auto-
claved for 30 minutes at 17 psi before use. Culture flasks and
experimental chambers were stoppered with polyurethane foam plugs
to allow for gaseous exchange while keeping out contaminating
particles.

An auxenic culture of the green alga, Scenedesmus

guadricauda (614), was obtained from the Indiana University

 

culture collection of algae. The alga was cultured in 300 ml
Erlenmeyer flasks in a specially built growth enclosure in which
temperature and photoperiod were regulated (Figure 1A). Culture
conditions in the flasks were: solution volume, 150 ml; temperature,
25 :_2°C; 14 hours illumination per day; light intensity of 125

ft-c provided by Westinghouse Agro-Lite fluorescent lights; con-
tinuous agitation of 100 oscillations per minute during the light

period; and initially equivalent nutrient concentrations.

Figure 1. Photographs of the growth enclosure containing
culture flasks (A) and experimental chambers (B).

 

A modified Rodhe VIII culture medium (Rodhe, 1948;
Golterman, 1960), with added micronutrient solution, was used for
stock cultures and growth experiments (Table l). One-milliliter
innoculates from stock cultures were transferred to new medium at
7- to 14-day intervals to ensure that cultures contained young,
rapidly dividing cells which would grow quickly when placed in
experimental medium.

Four days prior to an experimental run, a culture of algal
cells was centrifuged at 1300 rpm for 5 minutes, the supernatant
medium being discarded. The cells were innoculated into fresh
nutrient medium containing no phosphorous. During the four-day
period, the algae depleted their stored phosphorous and became
phosphorous-starved, a condition required for experiments involving

phOSphorous uptake. Scenedesmus quadricauda cells, when P-starved

 

for periods greater than 4 days, were observed to be in very poor
morphological condition.

Special chambers constructed of plexiglass were designed
for culturing algae under the experimental conditions (Figure 2).
The removable center partition of the chambers was designed to hold
a membrane filter 47 mm in diameter. A nucleopore membrane with
pore size of 3 microns was used because it allowed free movement
of nutrient ions across the membrane, while preventing the mixing
of algal cells and clay particles which were innoculated into oppo-
site sides of the chamber (Figure 2C). Conditions in the experi-
mental chambers were the same as in the culture flasks except for a

larger solution volume (450 ml).

TABLE l.--Composition of Modified Rodhe VIII Culture Medium

(Rodhe, 1948, Golterman, 1960).

 

 

 

Nutrients Concentration

Major mg/l
Ca(No3)2 - 4H20 10.0
MgSO4 - 7H20 20.0
KZHPO4 5.0
NH4N03 30.0
NaHCO3 80.0
Ferric Citrate 1.0
Citric Acid 1.0

Micronutrients
H3803 0.185
MnCl2 . 4H20 0.06
ZnCl2 0.01
CoCl2 ° 6H20 0.002
Na2M004 - 2H20 0.01
Na EDTA 0.1

2

 

Figure 2. Photographs of the experimental chambers showing
construction (A,B), separation of algal cells
and clay particles by the partition (C) and the
three experimental treatments (0).

IO

 

11

Preparation of P-Kaolinite

Kaolinite, an aluminum clay, was used to typify sediment
found in the aquatic system. Many investigators have shown the
phosphorous adsorbing capabilities of kaolinite (Stumm and Morgan,
1970; Smith, 1965; Muljadi et_al,, 1966; Upchurch et_al,, 1974).
The following procedure is slightly modified over the one used by
Helfrich (1972) for preparing P-kaolinite. Four liters of a 5%
clay suspension were homogenized in a blender. The suspension was
passed through a medium porosity 1R-120 ion exchange resin which
replaced Ca ions with H ions. A 1N solution of NaOH was added to
the eluant, maintaining the pH between 10 and 11 and replacing H
ions with Na ions. The Na-clay suspension was vigorously shaken
and allowed to settle for six hours, after which the clay in solu-
tion was siphoned off. The clay which had settled was discarded.
The clay suspension was once again passed through the ion exchange
column, replacing Na ions with H ions. The kaolinite becomes an
acidic aluminum clay which readily adsorbs phosphorous. One gram
of phosphate in the form of KH2P04 was dissolved in 1 liter 0.02M
HCl. The phosphate solution was added to the clay suspension
followed by continuous agitation for 24 hours to insure maximum
adsorption. The P-kaolinite suspension was then centrifuged at
8000 rpm for 5 minutes, discarding the supernatant. The P-clay
was air dried for 96 hours until a constant weight was achieved,
and then stored as a dry powder to retard bacterial growth.
Phosphorous analysis of the kaolinite after phosphorous-adsorption

revealed a concentration of 373 ug P/gram of clay.

12

Experimental Design

 

Twelve experimental chambers were used in a (4 X 3) experi-
ment designed to compare differences in growth attained using
different sources of phosphorous (Figure 18). In the first four
chambers, phosphorous was initially available in soluble form,
mixed freely with the algal cells. The next four chambers contained
phosphorous adsorbed to clay particles as the phosphate source.

The algal cells and clay particles were separated by a partition
allowing no physical contact between the two. The remaining four
chambers also contained P-clay as the only source of phosphate,

but the algae and clay were kept in mixed suspension with no parti-
tion present. Figure 20 illustrates each of the three chamber
types.

The initial phosphorous concentration in each chamber was
414 ug/liter, whether in soluble or adsorbed form. This concentra-
tion was achieved in the chambers containing clay by adding 0.5
gram of P-clay. In the chambers without clay, 1 m1 of a solution
containing 1.0468 grams KZHPO4 per liter of deionized water was
added. An appropriate volume of P-starved Scenedesmus quadricauda
culture was added to each of the chambers to obtain an initial
density of 150 cells per ml of medium. The solution in each chamber
was stirred thoroughly for one minute, three times daily, the first
being at the onset of the light period to aid the shaker in keeping
the solutions in each of the chambers in suspension during the light

period.

 

13

Measurement of pH and Phosphorous Concentration

Sampling was carried out every other day. At the end of the
dark period, the culture chambers were removed from the enclosure
and thoroughly stirred prior to sampling. Measurements of pH were
taken in each chamber with a Beckman pH meter and recorded to the
nearest tenth of a unit. In those chambers containing partitions,
pH readings were taken in both algal and clay sides. A 15-ml
sample was then pipetted from each chamber, but was taken from the
algal side only in the chambers with partitions. Three ml of each
sample were placed into small capped vials from which cell counts
were taken within two hours. The remaining 12 ml were expelled
into 15-m1 centrifuge tubes and centrifuged at 2500 rpm for 5 minutes.
Ten m1 of the clear supernatant were pipetted into clean, dry 125-ml
Erlenmeyer flasks for the ascorbic acid method of measuring soluble
phOSphorous concentrations (American Public Health Association,
1971).

In order to keep from decreasing culture volumes by too
large a factor over the experimental period, the recommended 20-m1
sample for phosphorous determination was halved. Reagents were
also added in one-half the usual amounts. Some preliminary experi-
ments using standard phosphate solutions of varying known concen-
trations showed that this decrease in volume of sample and reagents,
as long as it was kept proportional, did not alter the results of

the method.

14

Phorphorous concentrations were measured to determine at
what point in time phosphorous in solution reached concentrations

limiting to cell growth.

Measurement of Algal Growth

 

Algal densities were determined using Palmer-Maloney
counting chambers and a standard microscope equipped with a Whipple
square in the ocular. Counting chambers were loaded by pipette
from the 3-m1 samples taken at the beginning of the sampling day
and allowed to set 10 minutes before counting to facilitate the
settling of cells. Cell densities were recorded as algal cells

per ml. Figure 3 shows Scenedesmus quadricauda in culture medium

 

free of clay and containing clay.

15

Figure 3. Photographs of Scenedesmus guadricauda (450x)
in culture media free of clay (A) and containing
clay (B).

 

(A . “ \

 

Q' '
6* '3 o." "I I I ‘ I
Q 9 - .o" c ‘Q 1"“1 I

RESULTS

Phosphorous adsorbed to clay particles is available to

phosphate-limited Scenedesmus guadricauda cells. Some nutrients

 

in media containing suspended clay particles are more available to
algal cells than nutrients in media free of suspended clay. The
nutrients in some way become closely associated with or bound to

the sediment, whereas they are prevented from precipitating or from
forming chemical complexes with other nutrients. It is the author's
hypothesis that algae cultured in a sediment suspension not only
utilize dissolved nutrients but also remove the adsorbed nutrients
directly from the sediment particles. Suspended sediment tends to

enhance nutrient availability.

Densities of Algal Cultures

Mean densities of algal cells cultured under three condi-
tions of available phosphorous over a 16-day period are shown in
Figure 4. Algae cultured in a suspension of nutrient media and
phosphorous-clay reached a density of 137,000 :_5,000 cells per m1
as compared to 38,000 :_6,000 and 32,500 :_4,000 cells per ml in
cultures free of clay and partitioned from clay, respectively.
Algae cultured under the latter two conditions seemed to be limited
by something not limiting to algae cultured with suspended clay

particles.

17

18

Figure 4. Mean cell densities (:_one standard error)
of algal cultures over the 16-day experimental
period.

I00

0

Algal Cells per ML x lo3

 

19

 

 

 

.,l
0"!
: "/
-l ’0/
: /'
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.7
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- j /
_ / ,’
_ [I .ll’
- ,/
0/ IA
" ,/,’l
j/
,/
:‘ [Ill -— Algae w/0 CIOY
: I ----- Algae side of partition
: —'- Algae mixed w/ Clay
1 1 I l l l l ‘
O 2 4 6 8 IO I2 I4 l6

Day

20

Statistical Analysis of Growth Curves

Growth curve data were statistically handled in a split-
plot, repeat-measure design (Dr. John Gill, personal communication).
In such a design, a comparison can be made between treatments over
a period of time. The growth data of three successive experimental
runs were combined for each of the eight sampling days. An 8 X 8
matrix containing sums of products and sums of squares was made for
each of the three treatments. These three matrices were combined
into one 8 X 8 matrix containing covariances and variances. This
matrix was then inverted by computer. A multivariate parallel to
the "t-test" was used in determining which treatments were signifi-
cantly different. The values contained in the inverse matrix were
used in calculating test statistics, which were then compared with
appropriate critical values provided by Dr. Gill.

As shown in Figure 4, algae cultured in suspension with
clay particles attained significantly greater densities at a 99%
confidence level than algae cultured in media free of clay and
separated from clay. Statistical analysis also showed that algae
cultured without clay expressed significantly greater growth at a
99% confidence level than algae partitioned from the clay. This
difference is not apparent in Figure 4, probably due to the loga-

rithmic scale used in plotting data points.

Soluble Phosphorous Concentrations
Mean concentrations of soluble phosphorous in the algal

cultures over the 16-day experimental period are shown with their

 

21

standard errors in Figure 5. Soluble ph05phorous concentrations
lower than 20 ug/liter are limiting to growth of Scenedesmus

gpadricauda (Rodhe, 1948). Algae cultured in media free of clay

 

and separated from clay did not become phosphorous-limited over

the 16-day period, but did reach limiting concentrations on about ' a?
day 19 as compared to day 11 in clutures with suspended clay.

Phosphorous was being utilized at a much slower rate in cultures

not containing clay in mixed suspension. The cultures were being

limited by something other than phosphorous. g?

Relationship of pH to Nutrient Availability_

Figure 6 shows pH ranges over the l6-day period for each
culture condition. Initially, solutions containing clay were
slightly more acidic than solutions free of clay due to the acid
character of the P-kaolinite. Algal cultures free of clay and
separated from clay showed average pH values of 7.6 over the entire
l6-day period. Algae in suspension with clay particles exhibited
pH values increasing from 7.7 to 8.7, an indication of increased
growth and reduction of C02 concentrations during photosynthesis.

Effect of Various Iron Sources
on ATgal Growth

An experiment was designed to determine the effects of
various iron sources on the growth of algal cultures in an attempt
to establish if iron had limited growth in the previous experiment.
Twelve experimental chambers were established, each with an initial

iron concentration of 0.18 mg/liter as in the previous experiment.

22

Figure 5. Mean concentrations and standard errors of
soluble phosphorous in algal cultures over
the l6-day experimental period.

 

23

 

 

 

 

 

—--“'I\ , Algae "lo Clay
400‘) | --- Algae side of partition
\I\ —-- Algae mixed "/ Clay
I """ Extensions based on
fewer data points
350 ‘
’Z 250 ‘
22
I
O5
3
m 200 T
:1
o l
E l
9; \.\| t“
2 _ \ .|_. _ -|__ _|\ .\
Q. '50 . /|/‘./ \ x
/ / 1, x
/ \ \
/ ' \\ l
o A \l |\\ t‘
I 00 T / I, \\ ‘1‘
I \\ \
0,, \\ ‘1
I \\\‘1‘
so /, \| m
., \ k
I \l\ ‘ .\
I .\'_._._.-' ‘L - -\-:5
1 l l I l l l l l 1 4J
O0 2 4 6 8 l0 l2 I4 16 IB 20 22

Day

24

Figure 6. Ranges of pH values of algal cultures over
the 16-day experimental period.

9
B

7 -

9
8
7

:1:
ca.

9..
8..

25

Algae "lo Clay

\

1-———l~|——]————l-—l—l—-|

I I I I I I I I

Algal side of partition

I I I I I l I

Clay side of partition

 

7

84

I I I I I I I I

 

7

02468l0l2l4l6

J I

26

The way in which the iron was supplied varied among chambers. Four
of the chambers contained algae and P-clay in suspension as in the
previous experiment, with iron citrate as the iron source. The
remaining eight chambers contained an innoculate of algae in
nutrient media free of clay. In two of the chambers, initial iron
concentrations were supplied by ferric citrate as in the previous
experiment. The next two chambers were prepared as the preceding
two except that a concentration of iron, equal to the initial
concentration, was added every other day as ferric citrate solution.
The remaining four chambers were set up with iron being initially
supplied as Fe-EDTA in two of the chambers, and Fe-EDDHA in the
other two chambers.

Mean algal densities and ranges resulting from the experi-
ment are shown in Figure 7. Ferric citrate, added every other day
in concentration equal to the initial amount, produced algal
densities slightly greater than densities obtained in the clay
su5pensions. Fe-EDTA produced algal densities equal to those
densities in the clay suspensions. In the chambers where iron was
supplied as Fe-EDDHA, algal densities were 20,000 cells per ml less
than when clay was present. The lowest cell densities were achieved
in cultures provided with only an initial ferric citrate solution.

Mean soluble phosphorous concentrations and ranges of the
algal cultures receiving various iron sources are shown in Figure 8.
The concentrations of soluble phosphorous decreased in relation to
increases in algal densities. Algae cultured in chambers with ferric

citrate added every other day experienced phOSphorous-limitation on

 

eBI

27

Figure 7. Mean cell densities and ranges of algal
cultures treated with various iron sources
over a 20-day period.

I

O

Algal Cells per ML x lo3

 

28

   

 

.. _ — Fe-Citrate added initially only
-—--— Fe-Citrate added every other doll
------ Fe-EDTA

-— — Fe-EDDHA

Clay su5pension

 

8 l0 l2 l4 l6 l8 20
Day

29

Figure 8. Mean concentrations and ranges of soluble
phosphorous in algal cultures treated with
various iron sources over a 20-day period.

Phosphorous (ug /liter)

 

 

 

 

3O

 

 

 

 

 

 
 

 

 

 

 

 

- - — Fe -Citrate added initially only
400_‘\ --' — Fe-Citrate added every other day
\\\\k\ ------ Fe-EDTA
. \\ —-— -
\.\\\ Fe EDDHA .
\ ‘§‘I\ Clay suspensmn
o \\ \
\ ‘x I ,\
. x \
\
\ . Ix
300-. o \\ \ \
\ ‘l \ \‘
o ‘1 \ \
\I l‘ \ \\
\ “ ‘ l
. ‘. \ \
1‘ \ \
\O “\ \ \
200- r \
‘. \ \
o “ \ \
\ \ ‘. \ \
‘. \
, \
\. x \ \
\ .
o ‘1
l00-+ \ x \
.\ \\
\\I\° \
\\ __._\_A
02468l0|2l4l6182

Day

 

31

day 12, as compared to day 11 for algae cultured in clay suspension.
Algal cultures supplied with initial concentrations of Fe-EDTA,
Fe-EDDHA and Fe-citrate became phosphorous-limited on day 14, day

16 and day 20 respectively.

 

Ranges of pH values of the algal cultures in the iron P“?
source experiment are shown in Figure 9. Cultures with ferric I
citrate added only initially experienced low pH values over most
of the experimental period. In the other four culture conditions,
pH values increased to between 8.6 and 9.0. These higher values 943

are due to the increased algal growth and the subsequent reduction

in CO2 by photosynthetic activity.

32

Figure 9. Ranges of pH values of algal cultures treated
with various iron sources over a 20-day period.

33

 

 

 

 

 

 

 

 

 

 

9 Fe-Citrate added initially only
\1"—'—"_I-—I’l/
7 I I I I I I I I I I
9 - Fe-Citrate added every other day l/‘k _'
8 W /l/
\l/I
'7 I I I I I t l I I I
9 _ Fe-EDTA
/I
I _ /I——
Q. 8 .\l-——l——'——'/|
7 I I I I l I l I I I
9. Fe-EDDHA I I
/
/
8 -l\l . | '/"’I.——-—-I
7 1 1 l l l 1 I I l I

9 _ Clay suspension

7

8d'/

 

’
I

l-——l/

I I I I I I l I

o 2 4 6 8 l0 l2l416 l82
Day

 

21-? '5

DISCUSSION

The results of this research have shown that phosphate-

limited Scenedesmus gpadricauda can utilize this nutrient adsorbed

 

to clay particles. The only source of phosphate initially present
in the cultures was adsorbed to clay. The algal cells probably
obtain phosphate in two ways: (1) direct utilization off the clay
particles; and (2) utilization of dissolved orthophosphate desorbed
from the clay. During the first 4 to 6 days of the growth experi-
ments, many of the algal cells were seen to be tightly enveloped
by clay particles. This suggests that the P-limited algal cells
were somehow concentrating the P-clay particles resulting in
phosphate assimilation. The adsorbed phosphate is chemically
bonded to positively charged corners and edges of the clay (A13+)
(Stumm and Leckie, 1970; Stumm and Morgan, 1970). Concentrations
of the particles around algal cells would bring the phosphate ions

in contact with the cells. The long cellular extensions of the

Scenedesmus spp. may aid in this process. It may also be possible

 

that the algae are altering the chemical environment around them—
selves through metabolic activity, increasing the availability of
sediment-adsorbed phosphate, by increasing desorption of the
phOSphate ions.

The algae most likely obtain the bulk of their phosphate

supply from dissolved orthophosphate which was desorbed from the

34

 

35

clay particles. Adsorption-desorption of phosphorous is dependent
upon pH with maximum adsorption occurring over the slightly acid

pH range 4-7 (Murrmann and Peech, 1969). Desorption of phosphorous
increases rapidly above and below this range. Stumm and Morgan
(1970) found the maximum adsorption by kaolinite to be at a pH
near 3. Muljadi et_al, (1966) have shown that kaolinite sorption
is reversible over a pH range 5-9. The predominant dissolved

2', both

orthophosphate species of a pH range 5-9 are H2P04' and HPO4
of which are readily available to algal cells. Algae are able to
increase desorption of phosphate by increasing the pH of the medium
through photosynthesizing activity. The pH immediately adjacent
to a photosynthesizing algal cell should be higher than the pH in
the bulk of the medium. Since the sorption reaction is markedly
dependent upon pH, it is probable that the exchange takes place
at or near the surface of the algal cell rather than at a distance
from the cell. This is another point in defense of the idea that
algae obtained adsorbed phosphate directly from the sorption sites.
As shown in the results, algae cultured in medium free of
clay and partitioned from the clay suspension became iron-limited
in their growth, whereas algae cultured in suspension with clay
did not become iron-limited. Citrate can keep ferric iron in
solution only up to a pH value of about 7.5, above which the iron
precipitates as ferric hydroxide (Stumm and Morgan, 1970). Wetzel
(1975) has shown that ferric hydroxide has very low solubility
over a pH range 5-8. In the cultures free of clay suspension, the

iron is only minimally available due to agitation of the medium.

36

Suspended clay particles somehow prevent the iron in
ferric hydroxide from becoming unavailable, possibly by some
physical or chemical bonding to the clay. Ferric hydroxide
adsorbed onto clay particles increases the surface to volume
ratio of the particles, a condition which tends to keep them in rm;
suspension due to increased bouyancy. Wetzel (1975) has indicated i
that ferric hydroxide is adsorbed onto suspended particles from

which iron can be assimilated by phytoplankton. This would

 

explain why the algae cultured in suspension with clay particles
did not become iron-limited. In the chambers with partitions, the
clay held the iron in an available form in suspension, but the
algae were unable to utilize it because of the membrane partition.
Citrate was found to be a very poor iron chelater at pH
values greater than 7.5; however, extremely good growth was
evidenced in cultures where ferric citrate was added every other
day. The reasons for this increase in growth are twofold:
(1) iron was added in an initially available form every other day;
and (2) the instances of algae coming into contact with iron
increased due to the increasing iron concentrations. EDTA
(ethylene-diamine-tetraacetic acid) and EDDHA (ethylene-diaminedi-
o-hydroxyphenylacetic acid) are chelating agents which bind ferric
iron strongly over a pH range 4-9 (Cleton gt_gl,, 1963). Cultures
containing iron added as Fe-EDTA reached cell densities equal to
those reached by cultures containing a clay suspension. Growth in
cultures containing Fe-EDDHA was somewhat limited due to a light-

limiting effect caused by the red coloration of Fe-EDDHA.

37

It has been shown that suspended clay particles are just as
efficient in keeping iron available as some of the major chelaters
in use today. It has also been concluded that phosphate adsorbed
to sediments in natural water systems represents an available
nutrient source for phytoplankton. These results stress the
importance of sediment-adsorbed nutrients to algal growth when
sediments are agitated into the water column.

More research is needed on the role sediments play in

 

eutrophication of natural water systems. The key to understanding
sediment-nutrient-algae interactions lies in the complexity of the
microenvironment surrounding the interactions. A greater emphasis
should be placed on research involving the physical-chemical-

biological processes being carried out in these microenvironments.

LITERATURE CITED

38

5"“
ll

LITERATURE CITED

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“I

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Chiou, C. J. and C. E. Boyd. 1974. The utilization of phosphorous from
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Hydrobiologia. 45:345-355.

 

Cleton, F., A. Turnbull and C. A. Finch. 1963. Synthetic chelating
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Engelbrecht, R. S. and T. T. Morgan. 1961. Land drainage as a source
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39

4O

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41

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