Supporting Information
Environmental Science & Technology
Copper redox transformation and complexation by reduced and
oxidized soil humic acid: 2. Potentiometric titrations and dialysis cell
experiments
Felix Maurer,1 Iso Christl,1* Beate Fulda,1 Andreas Voegelin,2 and Ruben Kretzschmar1
1
Institute of Biogeochemistry and Pollutant Dynamics, ETH Zurich, CHN, CH-8092 Zurich,
Switzerland.
2
Eawag, Swiss Federal Institute of Aquatic Science and Technology, CH-8600 Dübendorf,
Switzerland.
* Corresponding author, e-mail address: [email protected]
Number of pages: 11
Number of figures: 6
Number of tables: 3
Contents
S1: Cu(I) sensitivity of the copper ion selective electrode (Fig. S1) .............................. S2
S2: Total Cu(I) determination by complexation with bathocuproine (Fig. S2 & S3, and
Tab. S1).................................................................................................................. S3
S3: Calculation of inorganic copper speciation (Tab. S2 & S3)..................................... S6
S4: Cu(I) formation during titration experiments plotted as fraction of total added copper
(Fig. S4).................................................................................................................. S8
S5: Conditional stability constants for Cu-HA binding (Fig. S5) .................................... S9
S6: Speciation for solid phase samples (Fig. S6) ....................................................... S10
S1
60
40
20
Cu-ISE potential (mV)
80
100
S1: Cu(I) sensitivity of the copper ion selective electrode
0
Cu(II) only (1, 3, 10, 30, 100 μM)
In presence of 30 μM Cu(I)
10.0
9.5
9.0
8.5
2+
−log{Cu }
8.0
7.5
7.0
Figure S1. Measured Cu-ISE electrode potential as a function of Cu2+ activity in solution for
calibration experiments in the presence and absence of Cu+. Titration experiments were
performed in 200 µM histamine (Fluka) for stabilization of both Cu+ and Cu2+, 0.1 M NaCl, and
20 mM phosphate buffer to adjust the pH value to 7.0. For Cu+ and Cu2+ additions, a 2 mM Cu(I)
stock solution prepared in 0.1 M HCl and 1 M NaCl and a 2 mM Cu(II) stock solution prepared
from CuCl2 were used, respectively. Best linear fit for the Cu(II) data is shown by the solid line
with slope of 27.6 per logarithmic increase in calculated Cu2+ activities. The arrow illustrates the
increase in Cu-ISE electrode potential due to the Cu+ spike to the solution containing 1 µM of
Cu2+.
S2
S2: Total Cu(I) determination by complexation with bathocuproine
Experimental. The absorbance of the Cu(I)-bathocuproine complex and its interactions with
untreated humic acid (HA) solutions were tested by measuring the UV–vis absorbance spectra of
mixtures of HA (concentrations: 0, 46, 93, and 186 mg L–1), bathocuproine (Alfa Aesar,
concentrations: 0 and 10 mM), Cu(I) (from 1 mM Cu+ stock solution prepared from CuCl
(Merck, p.a.) in 0.1 M HCl (Sigma-Aldrich, p.a.) and 1 M NaCl (VWR, p.a.), concentrations: 0,
15, 30, and 100 µM) in 100 mM phosphate buffer at pH 7 (prepared from NaH2PO4·H2O and
Na2HPO4·2 H2O, both Merck, p.a.). The amount of acid added by the addition of Cu(I) stock
solution was balanced by the same volume of 0.1 M NaOH (Sigma-Aldrich, p.a.) to avoid effects
on pH. Each combination of humic acid concentration, bathocuproine, and Cu(I) concentration
was prepared in duplicate, leading to a total number of 64 samples.
Data analysis. The dataset was analyzed by fitting the following model with interactions to the
data by linear regression:
y = β0 + β1·x1 + β2·x2 + β3·x3 + β12·x1·x2 + β13·x1·x3 + β23·x2·x3 + β123·x1·x2·x3 + ε
(E1)
where y is the absorbance of the mixture at 484 nm, and x1, x2, and x3 are the concentrations of
HA, Cu(I), and bathocuproine in the sample, respectively. ε represents the residual error.
Results. Figure S2 shows that the UV–vis absorbance of the HA and the Cu(I)-bathocuproine
complex are additive, that is, the sum of the UV–vis absorbance of the complex of Cu(I) with
bathocuproine alone and the absorbance of the HA alone equals the absorbance of the mixture of
HA, Cu(I), and bathocuproine.
Table S1 lists the estimated parameters and test results for the linear fit to equation (E1).
Graphical analysis of residuals showed that model assumptions (constant, zero variance and
normal distribution of residuals) are fulfilled sufficiently (Figure S3).
The only parameters that significantly influenced the measured UV–vis absorbance (i.e., p-value
< 0.01) were the cuvette absorbance (intercept β0), the HA concentration (β1), and the interaction
of Cu(I) and bathocuproine concentrations (β23). Molar absorbance of HA was found to be
0.0031 per (mg L–1) at 484 nm, which is the same as determined by Maurer et al.1 (0.00308 per
(mg L–1)). The interaction of Cu(I) and bathocuproine concentrations (β23) corresponds to the
molar absorbance coefficient of the Cu(I)- bathocuproine complex (11156 ±10 M–1 cm–1) that
can be used for quantification of Cu(I) (compare with 12700 M–1 cm–1 found by Moffett et al.2).
The presence of Cu(I) or bathocuproine had no significant influence on the absorbance of HA (p
> 0.01 for the corresponding interaction parameters β12 and β13). Further, also the HA
concentration was found to have no significant influence on the absorbance of the Cu(I)bathocuproine complex (p > 0.01 for the corresponding three-fold interaction parameter β123).
Therefore, the method measured always total Cu(I) present in the sample including Cu(I) bound
to HA.
From the standard error of the residuals (0.0095), a detection limit (±3 standard errors) of 0.029
in absorbance units or about 3 µM Cu(I) can be deduced. The residual standard error was found
to depend strongly on the number of solutions that were added to a cuvette before measurement
of the absorbance (number of pipette actions), however. For this reason, the residual standard
error for the main experiments (see main article) could be lowered considerably by using only
two solutions (sample and reagent solution) for preparation of the sample.
S3
0.7
0.4
0.3
0.0
0.1
0.2
Absorbance
0.5
0.6
buffer
HA
BC
CuI
BC.CuI
BC.CuI.HA
sum
400
450
500
550
600
650
700
Wavelength (nm)
Figure S2. UV–vis absorbance spectra for 15 µM Cu(I), 93 mg L–1 HA, and 10 mM
bathocuproine (BC) illustrating the additivity of the absorbance of HA and the Cu(I)bathocuproine complex. The dashed line shows the calculated sum of the spectra recorded
from the HA and Cu(I)-bathocuproine (BC.CuI) solutions only.
S4
Table S1. Parameter estimates, estimated standard error, and p-value of t-tests testing
for significance of the parameter for the linear model given in equation (E1).
1.0
3
2
1
-2
-1
0
1
2
Fitted values
Theoretical Quantiles
Scale-Location
Residuals vs Leverage
0.5
3
1
0 1 2
0.5
1.0
Fitted values
1.5
0.5
Cook's distance
-4
0.5
48
12
-2
35
1.0
1.5
Standardized residuals
93
34
34
93
1.5
0.0
Standardized residuals
0.5
35
-1 0
-0.01
93
-3
35
0.01
34
Normal Q-Q
Standardized residuals
Residuals vs Fitted
-0.03
Residuals
0.03
Parameter
Estimate
Std. Error
p
–2
–3
<10–16
β0
7.44·10
3.59·10
<10–16
β1
3.11·10–3
3.37·10–5
0.899
–8.64·10–6
6.80·10–5
β2
–3
–3
0.257
β3
5.81·10
5.07·10
0.588
β12
3.48·10–7
6.39·10–7
–7
–5
0.994
β13
3.88·10
4.77·10
–2
–2
<10–16
β23
1.12·10
9.62·10
–7
–7
0.649
4.14·10
9.04·10
β123
0.00951 (56 degrees of freedom)
ε
R2
0.9995
0.0
0.1
0.2
93
0.3
1
0.4
Leverage
Figure S3. Graphical residual analysis for the linear model fit of equation (E1) shown in Table
S1.
S5
S3: Calculation of inorganic copper speciation
Inorganic copper speciation was determined using a fixed ratio of complexed to free species
concentrations calculated using laws of conservation of mass and mass action.
For Cu+, the relationship between total ([Cu+]tot) and free ([Cu+]) Cu+ was formulated as:
[Cu+]tot = [Cu+] + [CuCl] + [CuCl2–] + [CuCl32–] = {Cu+}/γ1 + {CuCl} + {CuCl2–}/γ1 +
{CuCl32–}/γ2 = {Cu+} · (1/γ1 + KCuCl · {Cl–} + KCuCl2 · {Cl–}2/γ1 + KCuCl3 · {Cl–}3/γ2) = [Cu+] ·
γ1 · (1/γ1 + KCuCl · {Cl–} + KCuCl2 · {Cl–}2/γ1 + KCuCl3 · {Cl–}3/γ2) = αI* · [Cu+],
using [] and {} to denote concentrations and activities, respectively, and γ1 and γ2 to denote
activity coefficients for species with charge 1 or 2, respectively. Ki refer to stability constants
listed in Table S3.
For a NaCl concentration of 0.1 M, the ratio of total ([Cu+]tot) and free ([Cu+]) Cu+ concentration
(αI*) becomes
αI* = γ1 · (1/γ1 + KCuCl · {Cl–} + KCuCl2 · {Cl–}2/γ1 + KCuCl3 · {Cl–}3/γ2) = 0.78 · (1/0.78 + 103.1 ·
0.078 + 105.68 · 0.0782/0.78 + 105.02 · 0.0783/0.36) = 3064
For Cu2+, a similar calculation leads to:
[Cu2+]tot = [Cu2+] + [CuOH+] + [Cu(OH)2] = {Cu2+}/γ2 + {CuOH+}/γ1 + {Cu(OH)2}
= {Cu2+} · (1/γ2 + KCu(OH)+ · {OH–}/γ1 + KCu(OH)2 · {OH–}2) = γ2 · [Cu2+] · (1/γ2 + KCu(OH)+ ·
{OH–}/γ1 + KCu(OH)2 · {OH–}2) = αII* · [Cu2+]
For a NaCl concentration of 0.1 M, the ratio of total ([Cu2+]tot) and free ([Cu2+]) Cu2+
concentration (αII*) becomes
αII* = γ2 · (1/γ2 + KCu(OH)+ · {OH–}/γ1 + KCu(OH)2 · {OH–}2) = 0.36 · (1/0.36 + 10(6.5–7)/0.78 +
10(11.8–14)) = 1.32
Table S2 lists values for αI* and αII* calculated for concentrations of NaCl used here and in the
first part of this study3.
Table S2. Total dissolved to free ratios for Cu2+ (αII*) and Cu+ (αI*) used for calculation
of inorganic speciation in solutions with selected NaCl concentrations.
–
{Cl } (M)
γ1
γ2
αI *
αII*
0.1
0.078
0.78
0.36
3064
1.32
NaCl concentration (M)
0.073
0.023
0.058
0.020
0.80
0.86
0.40
0.56
1716
212.8
1.16
1.21
S6
Table S3. Stability constants used for calculation of inorganic copper speciation.
no.
1
2
3
4
5
6
7
8
9
10
reactions
Cu+ + Cl– = CuCl (aq)
Cu+ + 2 Cl– = CuCl2–
Cu+ + 3 Cl– = CuCl32–
Cu2+ + OH– = Cu(OH)+
Cu2+ + 2 OH– = Cu(OH)2
Cu2+ + e– = Cu+
Cu(0) = Cu2+ + 2 e–
CuCl(s) = Cu+ + Cl–
Cu2O(s) = 2 Cu+ + H2O – 2 H+
CuO(s) = Cu2+ + H2O – 2 H+
Eh0 (V)
0.153
0.34
log K at 25 °C, I=0 M
3.10
5.68
5.02
6.5
11.8
2.59
–11.49
–6.73
–2.17
7.66
reference
Martell et al.4
Wang et al.5
Wang et al.5
Martell et al.4
Morel and Hering6
Moffett and Zika7
Lindsay8
Martell et al.4
Lindsay8
Lindsay8
S7
S4: Cu(I) formation during titration experiments plotted as fraction of
total added copper
1.4
1.3
1.2
reduced HA (N2)
reoxidized HA (N2)
reoxidized HA (O2/N2)
Total Cu(I) to total Cu ratio
1.1
1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0.0
1 3 6 23 1 3 6 23 1 3 6 23 1 3 6 23
+0
+ 24
+ 48
+ 72
Reaction time (h)
Figure S4. Ratios of measured total Cu(I) to total added copper in the titrations solutions 1, 3, 6,
and 23 h after Cu2+ addition for each of the four copper spikes (indicated by dotted vertical
lines). The nominal copper loadings resulting from copper spikes were 9.5 mmol kg–1, 46 mmol
kg–1, 180 mmol kg–1, and 600 mmol kg–1 during the first, second, third, and fourth day,
respectively. The same data are shown in Figure 1A (main article) as absolute concentrations of
total Cu(I). Symbol types correspond to HA treatments and experimental conditions during
titration. Error bars represent a 95% confidence interval (±2 standard errors).
S8
100
12
S5: Conditional stability constants for Cu-HA binding
10
1
0.01
Cu(I)-Cl species
+
5
10 20
50 100 200
−1
Cu(I) tot (mmol kg )
500
Cu
1
100
2
2
5
(d)
10 20
50 100 200
−1
Cu(I) tot (mmol kg )
500
Cu(II)-HA
7
0.01
0.1
Fraction of total Cu(II) (%)
10
9
8
c
1
10
11
(c)
−1
(L kg )
[HA-Cu(II)]
[HAtot][Cu(II)]
Cu(I)-HA
0.001
7
6
12
1
log K =
(b)
0.1
Fraction of total Cu(I) (%)
10
9
8
[HA-Cu(I)]
c
log K =
[HAtot][Cu(I)]
−1
(L kg )
11
(a)
2+
6
0.001
Cu
1
2
5
10 20
50 100 200
−1
Cu(II) tot (mmol kg )
500
1
2
5
10 20
50 100 200
−1
Cu(II) tot (mmol kg )
500
Figure S5. Conditional stability constants for binding of (a) Cu+ and (c) Cu2+ to humic acid as
calculated from dialysis cell experiment data for Cu+ (Figure 2b) and titration experiment data
corresponding to 23 h equilibration for Cu2+ (Figure 2a). Speciation of (b) Cu(I) and (d) Cu(II) as
a function of copper loading in solutions containing 1 g L–1 HA and 0.1 M NaCl. The species are
plotted as fractions of total copper.
S9
S6: Speciation for solid phase samples
(a) 0.5
Cu
Calculated Eh (V)
0.4
2+
CuO(s)
0.3
0.2
Cu2O(s)
-
CuCl2
0.1
0.0
Cu(0)
-0.1
-0.2
-11
-10
(b)
-9
-8
-7
-6
-5
-4
log total inorganic copper (M)
-3
Calculated Eh (V)
0.3
0.2
Cu(II)-HA
0.1
0.0
Cu(I)-HA
Cu(0)
-0.1
-11
-10
-9
-8
-7
-6
-5
log total copper (M)
untrox redox untranox redanox
L1
L2
-4
-3
untrox redox untranox redanox
H1
H2
Figure S6. Calculated redox potentials as a function of (a) calculated total inorganic copper and
(b) total added copper in HA solutions used for preparation of solid phase samples described in
the first part of this study.3 Fractions of redox species based on LCF of Cu K-edge XANES
spectra were used as input data (see Table S2 in Supporting Information of Fulda et al.3).
Redox potentials were calculated using the Nernst equation and free Cu+ and Cu2+ activities
derived from isotherms corresponding to Cu+ binding (Figure 2b) and Cu2+ binding after 23 h
equilibration (Figure 2a) without correction for ionic strength deviations. Areas divided by lines
depict in plot (a) the most abundant inorganic copper redox species or oversaturation of a solid
phase as predicted by thermodynamics for the given experimental conditions (L: 0.023 M NaCl
(dashed lines) or H: 0.078 M NaCl (solid lines), 25 °C). For plot (b), the speciation calculation
additionally included HA copper species. Adjacent lines reflect the range of HA concentrations
used in the titration experiments. Symbol type corresponds to redox treatment (untrox: untreated
HA under oxic conditions; redox: reduced HA under oxic conditions; untranox: untreated HA under
anoxic conditions; redanox: reduced HA under anoxic conditions) and type (1: added as Cu(I); 2:
added as Cu(II)) and amount (L: 7.5 mmol kg–1; H: 150 mmol kg–1) of copper added (see first
part of this study3). Error bars represent a 95% confidence interval for the derived redox
potentials and were calculated using Monte Carlo estimation procedures with 1000 calculations
per point and assuming standard errors for the Cu+ isotherm parameters as shown in Figure 2b
(main article), and an absolute standard error of 5% for species fractions estimated by XANES
LCF.
S10
References
(1)
(2)
(3)
(4)
(5)
(6)
(7)
(8)
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S11