Environmental Consequences of Nanotechnologies: Nanoparticle Dispersion in Aqueous Media: SOP-T-1

Abstract

Homogenous dispersions of nanoparticles (NPs) for use in bioassay media for exposures/characterizations are a crucial need in ecotoxicology assays. The present protocol provides guidance and step-by-step methods for: (1) creating a working stock from nanoparticle powder and nanoparticle aqueous suspensions, (2) spiking working stock suspensions into aqueous bioassay media. The protocol also provides guidance on optimization of test media and organism health. Peer-reviewed publications and standards, with adaptations for aquatic bioassays are included. Users are directed to follow the protocol section relevant to their need and parent material (e.g. aqueous NPs or powder NPs). This protocol was developed on procedures created specifically for metal nanoparticles, but may have broader application. In an effort to validate procedural consistency, values associated with reference materials (RM) are included. The standard references include powderbased and aqueous suspensions from the National Institute of Standards and Technology (NIST) and NanoComposix (NC); including aqueous suspension of NIST Reference Material citrate coated 30 nm Au and NC citrate coated 30 nm Ag, and powdered NIST standard reference material 1898 TiO2 nanomaterial. The Ag and Au reference materials were selected to represent moderately soluble and relatively insoluble materials, respectively. If followed correctly, near-homogenous particle dispersions of the reference materials and a best case scenario for dispersion of test materials in aqueous media should be generated based on best procedural guidance available.

Assessing nanomaterial exposures in aquatic ecotoxicological testing: Framework and case studies based on dispersion and dissolution

“…Case study 1: non-dispersible material A dispersion pretest, performed according to Coleman et al (2015), determined that the AgNP powder was not dispersible in ultrapure water by bath or probe sonication. Since AgNPs may release toxic ions into test media (Kennedy et al, 2015b), two comparative bioassays were conducted; one that included the settled particles and another that adapted elutriate methods Example durations are based on typical daily water renewals used in bioassay test methods.…”

“…"Dispersible" is functionally and conservatively defined as !1% of the original concentration. "Stable" is functionally defined as ±20% of the initial concentration (Petersen et al, 2015;OECD, 2012;Coleman et al, 2015Coleman et al, , 2017aColeman et al, , 2017b.…”

“…Bioassays were conducted to compare two methods; one where an attempt was made to perform a serial dilution with particles present; one where only the top 200 ml of the supernatant after 1 h particle settling was used. The AgNP powder for both assays was probe sonicated in MHRW (20 mg/l) according to Coleman et al (2015). Serial dilutions were created from the stock for the particle testing in nominal concentrations of 10-0.08 mg/l while bath sonicating until adding organisms.…”

“…First, it was determined if the ENM was dispersible in ultrapure water; water column exposures may be of lower importance for ENMs that will not disperse in ultrapure water by available methods (Coleman et al, 2015), with sediment testing potentially more relevant. These issues are common to "difficult to test substances" (e.g.…”

“…The powder materials (TiO 2 , AgNP-powder) were dispersed in ultrapure water (Milli-Q Plus, 18.2 mX/cm, Billerica, MA) using available methods (Coleman et al, 2015;Taurozzi et al, 2011Taurozzi et al, , 2013 to determine dispersibility ( Figure 1). Stocks of the AgNP and TiO 2 powders were prepared in ultrapure water first by bath sonication (Fisher Scientific model FS-60, 130 W, Pittsburgh, PA), Table 1.…”