United States
Environmental Protection
Agency
Water Engineering
Research Laboratory
Cincinnati OH 45268
Research and Development
EPA/600/S2-87/002 May 1987
Project Summary
Factors Affecting the
Bioavailability of Cadmium
R. B. Corey, P. A. Helmke, D. R. Keeney, and G. C. Gerloff
Results are presented from two
studies designed to identify factors af-
fecting the bioavailability of Cd and
other trace metals in soils amended
with municipal sewage sludges and to
develop analytical methods for measur-
ing those factors. The research included
field, greenhouse, growth-chamber, and
laboratory studies. The field studies
were conducted on the agricultural re-
search stations at Arlington and
Hancock, Wisconsin. Growth-chamber
studies were done at the University
Biotron, and the greenhouse and lab-
oratory studies were conducted in
Department of Soil Science facilities at
the University of Wisconsin-Madison.
The first study was entitled "Heavy
Metal Bioavailability in Sludge-amended
Soils" and included (1) field studies
with two sludges containing about 200
mg Cd/kg to determine the effects of
application rate, sludge properties, soil
texture, and liming on uptake of Cd and
other trace metals by corn, (2) green-
house studies of the same factors but
with three sludges and ten soils, and (3)
laboratory studies to develop methods
for measuring parameters that have
been shown to affect bioavailability of
the trace metals. The second study was
entitled "Bioavailability of Cadmium"
and was concerned mainly with further
developing and applying methods for
predicting bioavailability of Cd and
other trace metals in soils amended
with sewage sludge.
This Project Summary was developed
by EPA's Water Engineering Research
Laboratory, Cincinnati, OH, to announce
key findings of the research project that
Is fully documented In a separate report
of the same title (see Project Report
ordering Information at back).
Introduction
Municipal sewage sludges have been
shown to be effective sources of plant
nutrients, particularly N and P, when
applied to agricultural land. However,
some sludges contain relatively high
levels of trace metals, which under some
circumstances can produce toxic re-
actions in plants and/or animals. The
trace metal Cd is of particular concern
because it can accumulate in sufficient
quantities in plant tissues to be toxic to
animals or humans ingesting those tis-
sues over an extended period. The pur-
pose of the investigations reported here
was to identify sludge and soil char-
acteristics that affect the bioavailability
of Cd and other trace metals and to
develop analytical methods for measuring
those factors. As our knowledge of the
sludge-soil-plant system increased during
these studies, it became apparent that a
particularly effective research approach
would be to devise methods that would
provide inputs to a computer model
describing solute transport and plant
uptake.
Transport of solutes in a soil system
can occur by mass flow (moving with the
labile soil water) or by diffusion (moving
in response to a concentration gradient).
Transport of metal downward in the soil
profile occurs dominantly by mass flow,
whereas transport to plant roots is mainly
a diffusion process (even for some con-
servative solutes such as nitrate), except
when the species being absorbed are at
very high concentrations. The soil and
plant factors that affect the rate at which
a plant absorbs a given metal from
solution in a diffusion-limited system are
shown in Equation 1. This equation is a
modification of an uptake equation that
describes the diffusive radial flux of solute
-------
(in our case a metal) from an isotropic
medium (soil) to a cylindrical sink (plant
root), in time, t assuming depletion of a
cylindrical volume of soil surrounding
each segment of root.
U = CMb(1
- exp
-27raAir0L»t
(D
D,0f
1.65 rn
Soil factors: C,, = initial concentration of
metal in soil solution
b = buffer power, the
change in concentra-
tion of total labile forms
(adsorbed + dissolved)
per unit of change in
concentration of dis-
solved metal
A| = fractional area of water
contacting root
0 - volumetric water con-
tent
D, = diffusion coefficient for
the metal in soil solution
f = conductivity factor re-
lated to length of dif-
fusion path
Plant factors: U = uptake per unit volume
of soil in time, t
a = root-absorbing power
(uptake flux density)/
(cone, at root surface)
r0 = root radius
rh = half-distance between
roots
LV = root density
We have developed analytical methods
that determine the proportions of metals
present as dissolved free ions, dissolved
complexes, and reactive adsorbed (labile
forms). These methods allow us to cal-
culate the dissolved metal concentration,
CN, and buffer power, b, in Equation 1.
We also developed methods for control-
ling metal-ion activities at realistic soil-
solution levels in solution cultures to
study the effects of speciation of dissolved
metals on the root-absorbing power, a,
and on competition among metal ions in
the uptake process.
Development of Analytical
Methods
Total Labile Metals
The accepted standard method for
determining labile elements is isotopic
exchange if suitable isotopes are avail-
able. Thus, 109Cd and 65Zn were equili-
brated with samples of sand and silt loam
soils with and without addition of 102
Mg sewage sludge/ha. Isotopically ex-
changeable Cd and Zn were determined
after 16 hr, and the results were com-
pared with amounts of Cd and Zn ex-
tracted by various diethylenetriamine-
pentaacetic acid (DTPA) and ethylenedia-
minetetracetic acid (EDTA) solutions.
Metal extraction by 0.005M EDTA in
0.01M CaCI2 at a 1:2.5 soihsolution ratio
gave the best agreement with isotopically
exchangeable values. DTPA extraction
consistently gave low results.
Metal Speciation In Soil
Suspensions
The method developed for determining
metal speciation in soil suspensions
involves equilibration of separate soil
samples with each of four solutions: (1)
0.01 M Ca(N03)2, (2) 0.01 M CaC12, (3)
0.01 M Ca(N03)2 + 10;45M EDTA (or hy-
droxyethylenediaminetriacetic acid
(HEDTA), and (4) 0.01 M Cad 2 + 0.005M
EDTA. If Cd is not detectable in the 0.01 M
Ca(N03)2 solution, Cd(N03)2 is added to
all solutions to give a final Cd concentra-
tion of 5 mg/L. Equilibration with Solution
1 gives total dissolved metals. The in-
crease in Cd concentration in Solution 2
over Solution 1 results from the formation
of cadmium chlorine complexes. Because
the stability constants for these complexes
and the CI" activity are known, the activity
of Cd2* can be calculated. The increases
in metal concentrations in Solution 3
over Solution 1 result from the formation
of metal-EDTA complexes. The activity of
any divalent metal ion, (M2t), can be
calculated from Equation 2 once the Cd2t
activity and relevant competitive stability
constant (K) are known:
tion 1. An overall competitive stability
constant (KN) for the natural complexes
can be calculated from Equation 3.
(M2+) = K(Cd2+)
[M-EDTA]
(2)
[Cd-EDTA]
If concentrations of any of the metals are
below detection limits in Solution 1 but
are detectable in Solution 2, activities
can be calculated, but information on
dissolved complexes cannot be obtained.
The Solution 4 extract is a measure of
labile metals. From the ratio of metal
activities in solution to labile metal con-
centrations, distribution coefficients can
be calculated for the partitioning between
solution and solid phases. Concentrations
of these metal ions can be determined
from the calculated activities and ionic
strength. Concentrations of dissolved
metal complexes (M-ch), are determined
by subtracting the calculated metal ionic
concentration from the respective total
metal concentration determined in Solu-
(M2lCd-ch]
(Cd+2HM-ch]
(3)
The method used for calculating free
metal ion activities assumes that the
addition of CaCI2 IN Solution 2 or EDTA in
Solution 3 does not alter the equilibrium
significantly; that is, the amount extracted
into solution is not a significant part of
the labile fraction. If this assumption does
not hold, the calculated activities will be
lower than those in a natural system.
Speciation of Dissolved Metals
Dissolved metals are partitioned be-
tween hydrated free-ions, ion pairs, and
organic and inorganic complexes. The
concentrations of total dissolved species
are usually within the detection limits of
sensitive analytical methods such as
flameless atomic absorption spectro-
photometry (FAAS), but present methods
that measure activities of free metal ions
are either not sensitive enough to mea-
sure the low activities present in most
soil solutions, or they are subject to in-
terference by components of the solution.
The method for determining metal specia-
tion in soil suspensions estimates free-
ion activities, but it requires contact with
a solid phase to maintain a constant
metal activity. In order to overcome these
limitations a sensitive method utilizing a
Donnan* equilibrium system was devel-
oped. In this method a cation-exchange
membrane separating a donor compart-
ment (through which the test solution is
pumped) from a much smaller compart-
ment containing an acceptor solution.
The latter is similar in composition to the
test solution with respect to major cations.
Free metal ions equilibrate across the
membrane in about 1 hr, but transfer of
complexed species is greatly retarded.
Thus, analysis of the acceptor solution
after 1 hr by FAAS gives the concentration
of the free ion, from which an activity
may be calculated. The detection limit is
determined by the detection limit of the
FAAS analysis.
Characterization of Soluble
Complexlng Agents
Chelating resins which provide a re-
servoir for metal ions and maintain
•Mention of trade names or commercial products
does not constitute endorsement or recommenda-
tion for use
-------
constant metal ionic activity are used to
control metal-ion activities so that
competitive selectivity coefficients can be
determined for dissolved ligands. The
chelating resin is placed in a dialysis bag
and equilibrated with the solution which
originally contains the dissolved ligand or
ligands but not the metal ion. The amount
of metal complex formed is the difference
between the total metal concentration
and the metal ion concentration supported
by the resin. Competitive stability con-
stant (K) is calculated from the following
equation where M and N are different
metals, L is the dissolved ligand and ML
and NL and the concentration of metal
ligands:
[ML] (N2+)
K = -
(4)
[NL]
A method was developed for estimating
the Cu-complexing capacity of dissolved
ligands. In this method, samples of solu-
tion containing the dissolved ligand re-
ceived varying amounts of added Cu2*.
The difference between the total Cu con-
centration determined by FAAS and Cu2+
concentration (determined either with the
Donnan-equilibri urn/cat ion-exchange
membrane system or with an ion-specific
electrode when Cu2+ activities are above
10~6M) gave the Cu-complexing capacity
at the measured Cu2+ activity.
Cadmium Adsorption Isotherms
A method was developed for adsorbing
Cd onto soil surfaces under conditions of
constant pH and Cd2+ activity. To produce
solutions with known and constant Cd2+
activity varying ratios of Cd-saturated
chelating resin to Ca-saturated chelating
resin were mixed and added to a dialysis
bag. A pH-adjusted, weak-acid resin was
also added for pH control. The resin mix-
ture in a dialysis bag was then equili-
brated with a soil sample in 0.01 M.
Ca(NO3)2, and the Cd adsorbed on the soil
was extracted with acid and determined.
The Cd adsorbed on the soil must be
small compared with that contained in
the chelating resin so that a nearly con-
stant, known Cd2+ activity is maintained.
Competition for adsorption sites on the
soil is limited to Cd2* and Ca2+ as the
resin adsorbs other metals that were
originally adsorbed on the soil. Competi-
tive adsorption at known metal activities
could be studied by including other metals
in the chelating resin.
Control of Solute Concentrations
In Nutrient Solutions
If nutrient solutions are to be used for
determining root-absorbing powers for
trace metals, concentrations of those
metals at the root surfaces must be
known. However, the nutrient solutions
do not buffer metal concentrations, and
even rapidly flowing solutions do not
maintain constant metal concentrations
at root surface when concentrations are
in normal soil solution ranges. For this
reason, a resin-buffered nutrient solution
system was developed. In this system, a
strong-acid resin controls the ratios of
Ca2+, Mg2+, K+, and Mn2+; a weak-acid
resin buffers pH and assists in controlling
ratios of- the previously mentioned
cations; a chelating resin controls the
ratios of trace metals, including Cd2t,
Zn2+, and Cu2+; and a strong-acid resin
partially saturated with (AI(OH)2+)n
polynuclear complexes controls P con-
centrations and assists in controlling
ratios of the major cations. Anions such
as NO3 and S04= must be maintained by
periodic additions. For most of the cations,
the ratios are controlled by the resins; but
the absolute concentrations are a function
of ionic strength. Thus the total solute
concentration as indicated by electrical
conductivity must be controlled within
rather narrow limits. This system has
been used successfully with the resins
enclosed in filter membrane bags, floating
free in the pot, or contained in a column
through which the nutrient solution is
recirculated.
Factors Affecting Cadmium
Bioavailability
Sludge Characteristics
Data from laboratory, greenhouse, and
field all show that sludge characteristics
are extremely important, if not dominant,
in controlling Cd (and Zn) availability to
plants. From 18% to 63% of the total Zn,
and from 12% to 63% of the total Cd in
the sewage sludges studied was isotopi-
cally exchangeable. The greatest per-
centage was for Chicago sludge, whereas
the lowest was for a sludge that received
soluble Fe during the treatment process.
When corrected for the effects of pH, the
fractions of isotopically exchangeable Zn
and Cd extracted by 0.01 M Ca(NO3)2 were
about the same for all sludges and soils
studied. This result suggests that the
mechanism by which solubility is con-
trolled is coprecipitation (solid solution)
of Zn and Cd with phosphates of Fe, Al, or
Ca, with hydrous oxides of Fe or Al, or
with CaC03 during the treatment process.
According to solid solution theory, the
solubility of a trace constituent should be
proportional to its relative concentration
on the surface of the precipitate. Thus for
equal concentrations of Cd in different
sludges, the solubility should decrease as
the amount of the precipitate's major
component (FePO4, AIP04, etc.) increases
with respect to the Cd concentration.
This effect is apparent when comparing
results for the Chicago sludge (low Fe, Al,
and P) with those for the Wisconsin
Rapids sludge (high Fe and P). Although
total concentrations of Cd were nearly
the same for both sludges, the isotopically
exchangeable Cd, solution Cd, and Cd in
barley leaves in the field were all much
higher for the Chicago sludge.
Similarly, in sludges with similar levels
of the major precipitant, the solubility of
Cd should increase with the total Cd
concentration. This result is borne out
(although not as clearly as would be
desired because of the low additions in-
volved) in the greenhouse study com-
paring plant uptake of Cd at equal Cd
additions from the Oshkosh (9 mg Cd/kg)
and Wisconsin Rapids (180 mg Cd/kg)
sludges.
If these observations on a few sludges
were found to hold as a general rule, Cd
solubility might be predicted on the basis
of sludge composition and verified by
analyzing for the capacity factor by iso-
topic dilution or ligand extraction and for
the intensity factor by equilibration with
Sr(N03)2 or Ca(NO3)2.
Sludge Application Rate
At high sludge application rates, the
properties of the sludge tend to determine
Cd and Zn availability, with the effects of
soil properties being limited primarily to
pH control. At low sludge rates, the
adsorptive properties of the soil appear to
exert some control.
Increasing additions of the high-Cd
Wisconsin Rapids sludge to soils in the
greenhouse resulted in proportionate
increases in DTPA-extractable Cd. Similar
increases in the levels of soluble DTPA-
extractable and isotopically exchangeable
Zn and Cd were observed in the field as
the rate of sludge application was in-
creased. However, these proportionate
increases in labile metal levels were not
consistently reflected in crop uptake. The
Cd concentration in corn tissue, par-
ticularly in the third greenhouse crop,
tended to plateau at about 80 Mg/ha
(18.3 kg Cd/ha) in the Plainfield sand
and 160 Mg/ha (36.6 kg Cd/ha) in the
Piano silt loam. Tissue Cd levels in the
field also tended to plateau at higher
sludge rates, but the effect was not as
apparent as in the greenhouse because
-------
of the lower rates used in the field. This
relationship was somewhat confounded
in the field by lower pH induced by the
high sludge rates.
In the field, the concentrations of Cd in
corn ear leaves tended to reach a plateau
between 1 and 1.5 mg/kg. Very little Cd
was found in the corn grain. In the
greenhouse, the Cd concentration in plant
tissue from the third corn crop reached a
plateau at about 7 mg/kg. This level is
much higher than in the field, even con-
sidering that the greenhouse tissue was
less mature. This result suggests that the
greater transpiration ratio of the green-
house crop may have resulted in greater
Cd uptake because of the increased
amount of Cd arriving at the root surface
by mass flow. Alternatively, Cd uptake in
the field may have been reduced by root
penetration into deeper soil regions not
affected by sludge addition.
Time Following Sludge
Application
In both the greenhouse and field
studies, DTPA-extractable Cd tended to
increase with time, but the effect ex-
tended over a longer period in the field,
probably because the sludge particles
were coarser and took longer to interact
completely with the soil. Mixing sewage
sludge with soil did not change the
isotopic exchangeability of either com-
ponent immediately (i.e., the concentra-
tion of isotopically exchangeable Zn in
the mixture remained the weighted aver-
age of the values for the soil and sludge).
However, mixing did increase the con-
centration of isotopically exchangeable
Cd by 30% to 40% in the mixture after 1
year.
The concentration of Cd in tissue tended
to increase from crop-to-crop, both in the
greenhouse and the field. This result is
contrary to those of most field studies
reported in the literature. However, in-
terpreting year-to-year differences in crop
Cd concentrations as due to changes in
lability of sludge Cd alone is questionable
because Cd uptake from a given soil
might also be affected by changes in
environmental conditions such as tem-
perature, moisture, pH, soluble salts, and
evapotranspiration.
SoilpH
The combination of liming and sludge
application resulted in a wide pH range
— greater than 2 pH units in the Piano
field plots. Isotopic exchange studies on
soil samples from these plots indicated
that the concentration of isotopically ex-
changeable Zn in the sludge-amended
soil was independent of pH. However,
isotopically exchangeable Cd at pH 6.2
after 1 year was only 80% of that mea-
sured at pH 5.4. Similarly, DTPA-extract-
able Cd on the limed plots in 1979 (after
3 years of equilibration) averaged 67% of
that of the unlimed plots. Graphs of the
negative logarithms of solution concen-
tration versus pH for control and sludge-
treated plots gave a slope of 1.0 for Zn
and 0.7 for Cd over a pH range of 4.6 to
6.8. An increase in 1 pH unit therefore
decreased the intensity factor of bioavail-
ability (concentration in solution) tenfold
for Zn but only fivefold for Cd. The values
of the slopes were independent of the
sludge application rate.
This marked decrease in soluble Cd
with increasing pH is not consistently
reflected in a corresponding decrease in
Cd uptake in the field, however. Although
the limed and unlimed plots on the Piano
soil differed by approximately 1 pH unit,
no significant effects of pH on Cd con-
centration in the leaves of corn or barley
were observed. Only in the corn leaf
tissue from Hancock at the high rates in
1978 and at all rates in 1979 was there a
markedly greater Cd concentration in corn
ear leaves from unlimed plots. Why the
increased solution Cd at lower pH did not
result in consistently greater Cd uptake
from the Piano soil is not apparent.
In contrast to the field studies, where
there was little effect of pH on Cd uptake,
addition of excess CaC03 to 10 soils in
the greenhouse caused marked decreases
in Cd uptake. This result is consistent
with others in the literature, indicating
relatively low Cd availability in calcareous
soils. The pH range in the greenhouse
study was higher than those used in the
field study, so the results are not directly
comparable.
Soluble Salts
Equilibration of Wisconsin Rapids
sludge with solutions of varying Ca con-
centrations resulted in solution Cd levels
that were directly related to Ca concen-
tration. This result is in accord with Cd-
Ca reactions on the chelating resin. Zinc,
Ni, and Fe also increased with increasing
Ca concentration, but only about half as
much as Cd. Similar results were obtained
with soils from the field plots when
equilibrated with varying concentrations
of Ca or Sr.
Tissue Cd levels in the second green-
house corn crop tended to be lower for
high-sludge treatments that had been
leached than for lower-sludge treatments
that had not been leached. This suggests
that Cd levels in the soil solution may
have been increased at higher salt levels.
Such a response implies that either the
salts in the sludge were present in the Ca
form or that the other cations induced
greater displacement of Ca from exchange
sites, which then competed with Cd for
more selective adsorption sites. This rela-
tionship is by no means clearcut, but it
does point out the need to control soluble
salt concentration 'following sludge ap-
plication, especially in greenhouse
studies.
Speciation of Dissolved Cd
Speciation studies indicated that most
of the Cd and Zn in the soil solution were
in the free-ion form, whereas Cu was
more than 90% complexed. In a Biotron
study using the resin-controlled nutrient
solution system, the Cd2+, Zn2+, and Cu2+
activities were controlled at levels char-
acteristic of a sludge-amended soil, but
total metal concentrations were changed
two to four orders of magnitude by
complexing with EDTA. Concentrations
of Cd in the plant shoots were not affected
by the presence of the soluble complex,
and concentrations of Zn and Cu were
increased less than 30% even though the
total Cu concentration in solution in-
creased 10,000 times. Metal uptake
therefore appears to be a function of the
free-ion concentration at the root surface.
Speciation results with Cd and Cu in
soil solutions from control and sludge-
amended soils by the Donnan membrane
equilibrium technique were compared
with those calculated by the GEOCHEM
program. The nine-ligand Mixture Model
of GEOCHEM generally tended to predict
higher proportions of free Cd2+ and Zn2*
than those experimentally determined.
Butter Power
The Cd buffer power is the change in
total labile Cd per unit change in dissolved
Cd. If curves relating adsorbed Cd to
solution Cd were linear, the distribution
coefficient, Cdadsorbed/Cdsolutlon, would
equal the buffer power. A survey of
literature values showed that distribution
coefficients decreased dramatically as
Cd/Ca ratios increased. The reason is
probably that sites with a very high affinity
for Cd are available at low Cd concentra-
tions; but at higher Cd concentrations,
these sites are filled, and Cd has to
compete with Ca for sites of lower Cd
specificity. Under these conditions, Cd
buffer powers determined at relatively
high Cd/Ca ratios drastically underesti-
-------
mate buffer powers at Cd/Ca ratios oc-
curring in normal soil solution. In fact,
the literature survey indicates that most
distribution coefficients have been deter-
mined at unrealistically high Cd values
and that buffer powers derived from those
values may be as much as three orders of
magnitude too low for normal soils. The
relationship of buffer power to uptake
shown in Equation 1 indicates that a
difference of this magnitude would have
a very great effect on predicted uptake.
Cadmium-Zinc Interactions
The addition of Zn(N03>2to unamended
Plainfield sand resulted in increased up-
take of Cd by plants grown in the green-
house, most likely as a result of increased
Cd in solution caused by displacement
from the relatively small number of ad-
sorption sites in this soil. Similar Zn
additions to Piano soil and to both soils
containing sludge resulted in little addi-
tional Cd uptake, probably because the
greater quantity of sorption sites furnished
by the soil and the sludge could accom-
modate the Zn with little Cd displacement.
These assumptions were supported by
the Cd concentrations determined in
0.01 M Ca(N03)2 extracts from the above
soil treatments. Zn addition to the Plain-
field soil greatly increased relative solu-
tion Cd levels, whereas the increases in
solution Cd levels in the Piano soil and
the sludge-amended soils were much
smaller.
Effects of Cd/Zn ratios in solution on
Cd and Zn uptake by tomatoes grown in
the resin-controlled nutrient system were
studied in a Biotron experiment. Ratios of
Cd/Zn were varied over four orders of
magnitude, with Cd varying from 10~93 to
1 (T 5M and Zn from 10'7 9 to 10'5 2M. Cu
was maintained at 10"113M. A linear re-
lationship (r2=0.94) was found between
log
-------
R. B. Corey, P. A. Helmke. D. R. Keeney. and G. C. Gerloffare with the University
of Wisconsin, Madison, Wl 53706.
J. A. Ryan is the EPA Project Officer (see below).
The complete report, entitled "Factors Affecting the Bioavailability of Cadmium,"
(Order No. PB 87-147 435/AS; Cost: $30.95, subject to change) will be
available only from:
National Technical Information Service
5285 Port Royal Road
Springfield, VA22161
Telephone: 703-487-4650
The EPA Project Officer can be contacted at:
Water Engineering Research Laboratory
U.S. Environmental Protection Agency
Cincinnati, OH 45268
United States
Environmental Protection
Agency
Center for Environmental Research
Information
Cincinnati OH 45268
BULK RATE
POSTAGE & FEES P/
EPA
PERMIT No. G-35
Official Business
Penalty for Private Use $300
EPA/600/S2-87/002
0000329 PS
CH?rfrnEAR80RN STREET
CHICAGO IL 606()4
------- |