Silver

Acute Toxicity: other routes

Administrative data

Endpoint:

acute toxicity: other routes

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

other: not rated (non-standard test system)

Data source

Reference

Reference Type:

publication

Title:

Acute toxic effects and gender-related biokinetics of silver nanoparticles following an intravenous injection in mice

Author:

Xue, Y. et al.

Year:

2012

Bibliographic source:

J. Appl. Toxicol. 32, 890 - 899

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Silver nanoparticles were intravenously injected into mice. Mice were exposed to different dosages of silver nanoparticles (7.5, 30 or 120 mg/kg). Toxic effects were assessed via general behaviour, serum biochemical parameters and histopathological observation of the mice.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Silver

EC Number:

231-131-3

EC Name:

Silver

Cas Number:

7440-22-4

Molecular formula:

Ag

IUPAC Name:

Silver

Test material form:

solid: nanoform

Details on test material:

- Name of test material (as cited in study report): silver nanoparticles (Shanghai Huzheng Nano Technology Co. Ltd, Shanghai, China)(manufacturer's stated average particle size of 15 nm (12 - 20 nm)
- Physical state: powder

Test animals

Species:

mouse

Strain:

ICR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Qingiongsham Experimental Animal Center, Nanjing, China)
- Age at study initiation: 5 weeks
- Housing: five mice of the same sex were housed in a stainless steel cage containing sterile paddy husk as bedding .
- Diet (ad libitum): standard pellet diet
- Water (ad libitum)
- Acclimation period: a week

All experimental procedures conformed to the guide for the care and use of laboratory animals (National Research Council, 1996)*.

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 2°C
- Humidity: 60 ± 10%
- Photoperiod (hrs dark / hrs light): 12/12

*Reference
- National Research Council. 1996. guide for the CAre and Use of Laboratory Animals. National Academy Press: Washington, DC.

Administration / exposure

Route of administration:

intravenous

Vehicle:

physiological saline

Details on exposure:

The silver nanoparticles were characterized using a transmission electron microscope (JEM-2100, Jeol, Japan) and a scanning electron microscope (S4800, Hitachi, Japan). Upon dispersing the silver nanoparticles in saline and prior to intravenous injection into animals, the particle size distribution of silver nanoparticles in the suspension was measured with a laser diffraction particle size analyzer (Zetasizer nano-ZS90, Malvern, UK).
Transmission electron microscope was used to measure the primary particle size of more than 400 random particles and compare the average with manufacturer's specifications. The silver particles were uniform in size and shape. Particles with diameter ranging from 10 to 30 nm accounted for 96% of the total particle count, with an average diameter of 21.8 nm. The scanning electron microscope imageshowed the silver aprticles to have good dispersion. However, as measured by laser diffraction particle analyzer at the time of injection, silver particles suspended in saline were 3.1 - 117 nm in diameter with a peak maximum at 90.5 nm accounting for 91% of the toal particle count. The results of particle characterization show that mice were exposed to agglomerates of silver nanoparticles that were larger than the primary nanoparticles.

Prior to dosing, silver nanaoparticles suspensions were prepared in saline. Mice were injected via a tail vein with saline or the test substance at a volume of 10 µL/g bw (doses of 7.5, 30 and 120 mg/kg). The experimental doses were selected based on the results of a preliminary study where a lethal dose of >200 mg/kg bw was observed.

Doses:

7.5, 30 and 120 mg/kg

No. of animals per sex per dose:

6 males/6 females

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 7 days or 14 days
- Frequency of observations and weighing: immediately after dosing, the vital signs and toxic effects of mice in each group were recorded. The body weight of each animal was measured at the initiation of treatment, then once a week during the treatment period thereafter.
- Necropsy of survivors performed: yes
Half of the mice in each group were killed on day 7 and the remaining mice were killed on day 14 following treatment. The brain, heart, lungs, liver, spleen, kidneys, testicle or ovary were carefully removed by incision. The organs were microscpically examined. Also, weights of organs (heart, lungs, liver, spleen and kidneys) were taken and relative organ weights were calculated based on body weight.
- Other examinations performed: blood samples were collected for haematological examination. Before necropsy about 1 mL of blood was drawn by removing the right eyeball and allowed to clot for biochemical analyses of serum. Serum biochemical parameters, including albumin, alkaline phosphatase, blood urea nitrogen, creatinine, glutamic oxalacetic transaminase, glutamic pyruvic transaminasae, lactate dehydrogenase, total cholesterol and total protein were analysed.

Statistics:

The quantitative data were expressed as means ± standard deviation. A multiple variance analysis and Duncan's multiple range tests were performed to compare the relative organ weights, and results of the blood biochemistry between the three experimental groups and the control mice. The software used for the data analysis was SPSS 13.0 for Windows (SPSS Inc., Chicago, IL, USA).

Results and discussion

Effect levels

Remarks on result:

other: no mortality was observed after single intravenous application of nanoAg at doses as high as 120 mg/kg

Mortality:

No mortality was reported by the authors.

Clinical signs:

No abnormal behaviour or clinical symptoms were observed in the experimental animals.

Body weight:

No significant changes in body weights of the animals were observed throughout the study period.

Gross pathology:

At necropsy, discoloured lungs, darkened liver and slight oedema were visually observed in 2/6 mice on day 7 and in 3/6 mice on day 14 for the high-dose group. The other organs looked normal in all experimental animals.

Other findings:

- Organ weights: no significant changes in relative organ weights of the animals were observed.

- Histopathology: lungs from high-dose silver nanoparticles exposure demonstrated thickened alveolar walls and infiltration of focal inflammatory cells on day 7, but these changes diminished by day 14. Slight interstitial oedema and inflitration of inflammatory cells were observed in lungs in the mid-dose group, compared with slight and negligible changes in the control group. Oedema and loose cytoplasm on liver cells were found in the high-dose silver nanoparticles treatment mice. No inflitration of inflammatory cells was present on liver cells in all mice. There were no remarkable histopathological changes in brain, heart, spleen, kidneys, testicles or ovaries in all of the experimental animals.

- Serum biochemistry:
- 7 days after administration: a statistically significant increase in the lactate dehydrogenase level was observed in the low-dose group when compared with the control group. Furthermore, a statistically significant decrease in the total protein and albumin levels was noted in the low-dose and the mid-dose groups when compared with the control group (table 1 in the field "Any other information on results incl. tables" below).
- 14 days after administration: on day 14, the mid-dose group exhibited significantly higher lactate dehydrogenase and blood urea nitrogen levels than those in the control group. The low dose group displayed a glutamic oxalacetic transaminase level that was significantly lower than that of the control group (table 1 in the field "Any other information on results incl. tables" below).
- The other serum biochemical parameters such as glutamic pyruvic transaminase, alkaline phosphatase, creatinine and total cholesterol did not show significant differences between the treatment and the control groups. Neither were there any significant dose-related changes in serum biochemistry parameters. Some parameters showed relatively large standard deviations, which could be due to inter-animal variation.

Any other information on results incl. tables

Table 1

		Experimental group (mg/kg)
	Biochemical parameters	7.5 mg/kg	30 mg/kg	120 mg/kg	Control (saline)
7 days	ALB (g/L	13.83 ± 1.47*	13.33 ± 1.03	15.92 ± 0.92	16.17 ± 1.83
	LDH (IU/L)	789.00 ± 341.03*	575.33 ± 65.70	640.20 ± 53.30	429.33 ± 44.00
	TP (g/L)	50.17 ± 4.71*	48.67 ± 3.61*	55.33 ± 2.66	55.33 ± 3.78
14 days	BUN (mmol/L)	7.28 ± 1.25	8.93 ± 1.03*	7.67 ± 1.74	6.55 ± 1.67
	GOT (IU/L)	100.00 ± 17.19*	139.00 ± 34.88	129.33 ± 20.89	131.00 ± 17.65
	LDH (IU/L)	555.67 ± 122.11	667.00 ± 217.24*	536.00 ± 77.24	464.17 ± 123.79

* Significant difference from the control value P < 0.05

ALB: albumin

LDH: lactate dehydrogenase

TP: total protein

BUN: blood urea nitrogen

GOT: glutamic oxalacetic transaminase

Applicant's summary and conclusion

Conclusions:

Silver nanoparticles (i.v) exerted no obvious acute toxicity in the mice with the 14-day observation period. However, inflammatory reactions in lung and liver cells were induced in mice treated at the highest dosel level (120 mg/kg).