Toxicity, Uptake, and Accumulation of Nano and Bulk Cerium Oxide Particles in Artemia Salina

Toxicity, uptake, and accumulation of nano and bulk cerium oxide particles in Artemia salina

Einstein Mariya David Sugantharaj David, Madhav Madurantakam Royam, Suresh Kumar Rajamani Sekar, Bhuvaneshwari Manivannan, Swathy Jalaja Soman, Amitava Mukherjee, Natarajan Chandrasekaran*

Centre for Nanobiotechnology, VIT University, Vellore 632 014, India

Supplementary Information

*Corresponding author

Dr. N. Chandrasekaran

Senior Professor & Director

Centre for Nanobiotechnology, VIT University

Vellore-632014

Email: ,

Phone No: +91 4162202624; fax: +91 416 2243092.

1. Material and methods

1.1 Microscopic studies

Morphological changes in the control, nano and bulk CeO2 treated A. salina was imagined using phase contrast microscopy (Zeiss Axiostar Optical Microscope, USA). After 48 h of interaction, the control and treated A. salina nauplii were fixed on a glass slide using 2% of glutaraldehyde and viewed under a microscope at 40X magnification. The changes in morphology in degradation of organelles, loss of appendages, damages in gut were noted by the acquisition of images using the inbuilt camera in the microscope.

2. Results:

2. 1 Microscopic studies

The uptake nano and bulk cerium oxide particles into A. salina nauplii were examined by optical microscopy (Fig. S4). Nauplii in the control group showed no visible morphological changes or damage to the appendages and antennae. No uptake of Ce particles in the gut of nauplii present in the control group was either seen (Fig S4 A). The A. salina nauplii exposed to different concentrations of nano (Fig S4 B–G), and bulk CeO2 (Fig S4 H–M) where represented. Nauplii treated with 0.3 mg/L of nano CeO2 did not show any uptake, whereas, lesser uptake was observed in gut lining of A. salina treated with 3 mg/L. Further, the ingestion of nanoparticles increased for higher exposure concentrations. The uptake of nano CeO2 (35 mg/L) from the mouth to the entire gut lining was clearly seen. For the bulk particle treatment the lower exposure concentrations (0.7, 7 and 14 mg/L), did not show any uptake. Ingested particles from labrum to whole gut were seen in 52 and 100 mg/L exposure to A. salina. Further, this was confirmed using quantification of uptake of Ce into A. salina form Table 1.

Figures:

Scanning electron microscopy

Fig S1: SEM image of nano (A) and bulk (B) CeO2

XRD pattern of nano and bulk CeO2 particles

Fig S2: XRD pattern of nano (A) and bulk (B) CeO2

Sedimentation profile of nano and bulk CeO2 particles in seawater

Fig S3A: Sedimentation profile of the different concentrations of nano cerium oxide particles in seawater medium at different time intervals (0, 1, 2, 4, 8, 12, 24, and 48 h). The data are presented as mean ± SD, n-3.

Fig S3B: Sedimentation profile of the different concentrations of bulk cerium oxide particles in seawater medium at different time intervals (0, 1, 2, 4, 8, 12, 24, and 48 h). The data are presented as mean ± SD, n-3.

The in vivo reactive oxygen species generation using fluroscance micrscopy.

Fig S4: The in vivo reactive oxygen species generation was confirmed using fluorescence micrscopy. The control Artemia salina devoid of nano and bulk CeO2 treatment is shown in A. Treatment of nano CeO2 (25 and 32 mg/L) are represented in B and C. Bulk CeO2 (52 and 100 mg/L) are represented in D and E.

Microscopic studies

Fig S 5: The morphological changes in the control (A), nano CeO2 of 0.3, 3, 9, 18, 25and 32 mg/L (B–G) and bulk CeO2 of 0.7, 7, 14, 28, 56 and 100 mg/L (H-M) particles treated Artemia salina.