SUPPORTING INFORMATION Fractionating Nanosilver: Importance for Determining Toxicity to Aquatic Test Organisms Alan J. Kennedy†*; Matthew S. Hull‡; Anthony J. Bednar†; Jennifer D. Goss†; Jonas C. Gunter§; Jennifer L. BouldinΩ; Peter J. Vikesland‡; Jeffery A. Steevens† * U.S. Army Engineer Research and Development Center Environmental Laboratory, EP-R 3909 Halls Ferry Rd Vicksburg, MS, 39180 Voice: 601-634-3344 Fax: 601-634-2263
[email protected] † U.S. Army Engineer Research and Development Center, Vicksburg, MS, 39180 ‡Department of Civil and Environmental Engineering, Virginia Polytechnic Institute and State University, Blacksburg, VA § Luna Innovations, Blacksburg, VA Ω Arkansas State University, Jonesboro, AR
Number of pages in Supporting Information (including cover): 7 Number of tables in Supporting Information: 4 Number of figures in Supporting Information: 2
S1
Table S1. Quality control measures for the dynamic light scattering data presented in Table 1 for the tested Nano-silver (nAg) materials. The baseline index is provided as a measure of the quality of fit to the algorithm in the DLS software. PI = polydispersity index; BI = baseline index Hydrodynamic Diameter by intensity (nm) Material MQ water MHRW
ASAP-nAg
PI = 0.13; BI = 7.6
PI = 0.06; BI = 8.0
Citrate-nAg
PI = 0.29; BI = 8.4
PI = 0.28; BI = 8.8
EDTA-nAg
PI = 0.27; BI = 6.4
PI = 0.29; BI = 3.9
NanoComposix (10 nm)
PI = 0.39; BI = 0.0
PI = 0.22; BI = 8.8
NanoComposix (20 nm)
PI = 0.18; BI = 6.2
ND
NanoComposix (50 nm)
PI = 0.19; BI = 9.6
ND
NanoComposix (80 nm)
PI = 0.19; BI = 6.1
PI = 0.20; BI = 8.9
Polyvinyl pyrrolidone Ag
PI = 0.24; BI = 7.5
PI = 0.24; BI = 6.5
S2
Table S2. Electrophoretic mobility and zeta potential of nanosilver suspensions in milli-Q (MQ) water and moderately hard reconstituted water (MHRW) after 5 minutes and 24 hours of interaction with the media. In MQ Material
In MHRW (5 min.)
In MHRW (24h)
Electrophoretic Mobility (m2/v*s)
Zeta (mV)
Electrophoretic Mobility (m2/v*s)
Zeta (mV)
Electrophoretic Mobility (m2/v*s)
Zeta (mV)
Citrate-Ag
-3.6 ± 0.2
-46 ± 3
-2.1 ± 0.1
-27 ± 1
-1.3 ± 0.2
-17 ± 3
EDTA-Ag
-2.6 ± 0.2
-33 ± 2
-1.7 ± 0.2
-22 ± 2
-2.0 ± 0.1
25 ± 1
NC10-Ag
-1.7 ± 0.1
-21 ± 2
0.4 ± 0.3
5±4
-0.6 ± 0.2
-8 ± 3
NC20-Ag
-1.8 ± 0.1
-23 ± 1
-1.7 ±0.1
-21 ± 2
-0.8 ± 0.5
-11 ± 6
NC50-Ag
-2.7 ± 0.1
-34 ± 1
-0.9 ± 0.7
-12 ± 4
-1.6 ± 0.1
-21 ± 1
NC80-Ag
-1.5 ± 0.1
-20 ±1
-0.9 ± 0.1
-11 ± 1
-1.9 ± 0.1
-24 ± 2
PVP-Ag
-1.1 ± 0.1
-15 ± 1
-0.7 ± 0.1
-9 ±1
-0.5 ± 0.3
-6 ± 4
S3
Table S3. Silver (Ag) measured in the unmodified stock, a supernatant of the stock produced by ultracentrifugation at 100,000 x g for 60 minutes, percent dissolved in the stock measured by field flow fractionation (FFF) and percentage of nAg in the supernatant of the high Daphnia magna and Pimephales promelas exposure concentrations following ultracentrifugation.
Material
Stock concentration Supernatant in milli-Q concentration water (µg/L) (µg/L)
% Ag in stock supernatant
% dissolved by FFF
% Fractionated % Fractionated Silver in Silver in highest highest MHRW MHRW concentration concentration (P. promelas) (D. magna)
Calculated % fractionated at LC50 (P. promelas)
ASAP-Ag
10,300
NA
NA
NA
57%
57%
16%
Citrate-Ag
5,550
1100
20%
7%
6%
25%
21%
EDTA-Ag
15,800
520
4%
4%
7%
2%
3%
NC10
5,170
374
7%
5%
9%
10%
11%
NC20
10,147
610
6%
4%
7%
7%
7%
NC50
10,372
351
3%
4%
7%
7%
6%
NC80
1,560
36.8
2%
6-8%
ND
5%
4%
PVP-Ag
89,100
2750
3%
3%
3%
NA
NA
S4
Table S4. Change in total and fractionated (dissolved) silver concentrations of nanosilver particles after 48-h exposure to the fish Pimephales promelas. The percent increase or decrease in silver concentrations is provided for the no observable effect concentration (NOEC). Asterisks indicate a value below detection limits.
Material
Effect level (% survival)
NanoComposix 10
Total Silver (µg/L)
Fractionated Silver (µg/L) 0 48 Percent hours hours Change
0 hours
48 hours
Percent Change
NOEC (95 ± 6%)
22.4
18.3
-18%
2.6
3.1
+20%
NanoComposix 80
NOEC (100 ± 0%)
81.1
69.7
-14%
0.6
3.3
+432%
NanoComposix 80
NOEC (97 ± 6%)
93.1
82.8
-11%
2.0
3.4
+73%
EDTA-nAg
NOEC (98 ± 5%)
18.0
8.5
-53%
0.5 *
1.1
+120%
EDTA-nAg
NOEC (90 ± 8%)
31.5
20.0
-37%
1.3
2.7
+108%
S5
(a)
(b)
(c)
(d)
(f)
(e)
(g)
Figure S1. Field flow fractionation analysis of nano silver suspensions for (a) citrate-Ag (b) EDTA-Ag, (c) NC10, (d) NC20, (e) NC50, (f) NC80 and (g) PVP-Ag. The left peak corresponds to the dissolved concentration. S6
(a)
Percent Survival
100
AgNO3 (3 tests) ASAP-nAg
80
Citrate-nAg EDTA-nAg
60
NC10 NC20 NC50
40
NC80 PVP-nAg
20 0 0.1
1
10
100
Log Total Silver (ug/L) (b)
Percent Survival
100 80 60 40 20 0 1
10
100
1000
Log Total Silver (ug/L) Figure S2. Dose response for Daphnia magna (a) and Pimephales promelas (b) exposed to silver nitrate (AgNO3) and nanoparticle suspensions.
S7
