Disaggregation of gold nanoparticles by Daphnia magna

Figures & data

Table 1. Size of Au-nanoparticles measured with three different methods, including dynamic light scattering (DLS), differential sedimentation centrifugation (DSC), and absorbance spectroscopy.

	Diameter (nm) by DLS		Diameter (nm) by DSC
Condition	Dh (nm)^a	Size distr^b (%)	Dh (nm)^a	Distribution (nm)^c	Absorbance spectroscopy λmax (nm)
AuNP	29 ± 0.3	12.8	25	7	529

^a D_h refers to the hydrodynamic diameter with ^bthe size distribution reported as a mean value (DLS) and ^cat 50% of the peak intensity (DSC).

Figure 1. Au nanoparticles were incubated under three conditions: TW (tap water), DTW (tap water containing Daphnia) and CTW (conditioned tap water, in which Daphnia had filtered the water for 24 h before removal). The media were then analysed with dynamic light scattering (A), disc centrifugation (B), absorbance spectroscopy (C) and trans emission electron microscopy (D). The particles and proteins in DTW and CTW were further analysed by SDS gel chromatrography (E) and mass spectrometry (F).

Figure 2. Absorbance spectra of Au nanoparticles when incubated in tap water (TW) at (A) 0 hour, i.e. the samples were taken right after the particles were added to the test tubes, (B) 1 hour, (C) 4 hour and (D) 24 hours (4 samples each), with (E) TEM image from particles in tap water (TW).

Figure 3. Absorbance spectra of Au nanoparticles when incubated with Daphnia (DTW) at (A) 0 h, i.e. a sample taken right after the particles were added to the test tubes, (C) 1 h, (E) 4 h and (G) 24 h (four samples each), with TEM images at time points (B) 0 h, (D) 1 h, (F) 4 h and (H) 24 h.

Table 2. Diameter of Au nanoparticles at different incubation times in TW, CTW and DTW, as determined by DLS, DSC and absorbance spectroscopy (N = 4 for each time point).

	Diameter (nm) by DLS			Diameter (nm) by DSC		Absorbance spectroscopy λmax (nm)
Condition	Peak 1	Peak % Pd	Peak 2	Peak 1	Peak 2	λmax(1)	λmax(2)
TW 0 h	28 ± 0.5	13	No	27	No	528	No
TW1 h	51 ± 0.9	16	No	33	No	528	636
TW 4 h	ND	ND	ND	44	No	528	671
TW 24 h	48 ± 1.8	16	Yes	97	No	527	700
CTW 0 h	37 ± 0.7	25	Yes	25	No	529	No
CTW 1 h	43 ± 1.0	16	Yes	25	No	529	No
CTW 4 h	53 ± 1.6	28	Yes	25	No	530	650^a
CTW 24 h	53 ± 1.7	32	Yes	28	63	529	650^a
DTW 0 h	35 ± 1.0	21	Yes	26	No	529	No
DTW 1 h	72 ± 2.9	35	Yes	27	No	529	627
DTW 4 h	88 ± 8.5	32	Yes	32	57	532	639
DTW 24 h	115 ± 7.5	21	Yes	18	45	532	646

Notes: Peak 1 and 2 refer to two separate size distributions of nanoparticles, as determined by DLS or DSC for which some samples only showed one size distribution. A “No” denotes that no second peak was recorded and “Yes” indicates a second peak bigger than the first peak. ND stands for not determined. For samples analyzed with absorbance spectroscopy, one or two λ_max values are observed, as indicated.

^aOnly half of the samples show a second maximum around 650 nm.

Figure 4. Absorbance spectra of Au nanoparticles when incubated in conditioned water (CTW) at (A) 0 h, i.e. samples were taken right after the particles were added to the test tubes, (B) 1 h, (C) 4 h and (D) 24 h (four samples each), with (E) TEM image from particles in conditioned water (CTW).

Figure 5. Images of (A) Daphnia (control) and (B) Daphnia exposed to Au nanoparticles after 4 h of incubation.

Table 3. Diameter of Au nanoparticles at different time points for BSA-coated particles (N = 3 for each time point).

	Diameter (nm) by DLS			Diameter (nm) by DSC		Absorbance spectroscopy λmax (nm)
Condition	Peak 1	Peak % Pd	Peak 2	Peak 1	Peak 2	λmax(1)	λmax(2)
TW 0 h	29 ± 0.5	8	No	20	No	523	No
TW 1h	60 ± 5.9	7	No	20	No	523	No
TW 4 h	27 ± 0.4	7	No	20	No	523	No
TW 24 h	23 ± 0.7	8	No	20	No	523	No
CTW 0 h	25 ± 0.5	9	Yes	20	No	523	No
CTW 1 h	23 ± 0.8	13	Yes	20	No	523	No
CTW 4 h	35 ± 0.4	9	Yes	18	No	–	–
CTW 24 h	36 ± 0.9	17	Yes	18	No	–	–
DTW 0 h	27 ± 0.5	11	Yes	18	No	525	No
DTW 1 h	53 ± 6.8	8	Yes	18	No	525	No
DTW 4 h	36 ± 0.6	10	Yes	18	No	525	No
DTW 24 h	39 ± 2.0	49	Yes	19	No	524	No

Notes: Peak 1 and 2 refer to two separate size distributions, as determined by DLS or DSC for which some samples only show one size distribution. A “No” denotes that no second peak was recorded and “Yes” indicates a second peak larger than the first peak. For samples analyzed with absorbance spectroscopy, one or two λ_max values are observed, as indicated.

Figure 6. Au nanoparticles were incubated under three conditions TW (tap water), DTW (tap water containing Daphnia) and CTW (conditioned tap water in which Daphnia had filtered the water during 24 h prior to being removed). The water was then analyzed with DLS, DSC, absorbance spectroscopy and TEM. In TW the particles aggregate. In DTW and CTW proteins and other biomolecules present bind to the particles affecting the size of the aggregates. As shown in the Venn diagram, the overlap in the number of identified proteins (n = 8) is very low compared to the overall number of proteins present in DTW (red, n = 90) vs. CTW (blue, n = 175).

Figure 7. BSA-coated Au nanoparticles were incubated under three conditions TW (tap water), DTW (tap water containing Daphnia) and CTW (conditioned tap water, pre-incubated with Daphnia for 24 h before removal). The media were then analyzed with DLS, DSC, UV–VIS and TEM showing that the particles do not aggregate but remain in their initial state.