Abstract
This project developed methodology for in vitro toxicity assessment of contaminated sites using the Promega® MTS
[3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxy-methoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay performed on
human cells (HepG2 and Skin fibroblasts). The project included the development of a method for extracting
contaminants from soil based on leaching and centrifugation. A number of solvents and surfactants were assessed
for their suitability as extracting agents. The Zwitterionic surfactant CHAPS ({3[(3-Cholamidopropyl)
dimethylammonio] propanesulphonic acid}), which is an irritant in vivo, was found suitable for in vitro toxicity
assessment applications. CHAPS was found to be the least toxic surfactant in vitro when tested on skin fibroblasts
(NOEC: 1800±577 ppm, IC50: 4000±577 ppm) and HepG2 cells (NOEC: 833±289 ppm, IC50: 5300±287 ppm). The
chosen surfactant was used in three different methods for extraction of Toluene and Xylene spiked in 2 g and 10g
soil. The combination comprising of 0.1% (s/w) CHAPS and cosolvent 1% (w/w) Isopropanol, at their respective
NOEC (No Observed Effective Concentration) toxicity values, showed good recovery of the nonpolar organic
compounds in comparison to the recovery by 0.1% CHAPS and 0.5% CHAPS. The study found additive interactions
to be the most common form of toxicity for 16 concentration combinations of Formaldehyde (polar), Toluene and
Xylene (nonpolar) when compared to predicted toxicity (R2=0.943, P<0.0001). When assessing the in vitro toxicity of
unknown (blind) contaminated soil samples, the Hazard Index (HI) predicted from the chemical analyses results
showed a relatively good correlation (R2>0.7062, n=26) when compared to the experimental toxicity results on
HepG2 cells. Furthermore, the comparison of Australian Health Investigation Levels (HIL) with in vitro toxicity testing
gave similar correlation (R2>0.6882, n=26) on HepG2 cells. The overall project suggests the potential application of
the zwitterionic surfactant (CHAPS) in sampling contaminants from soils in an in vitro toxicity assessment. This study
demonstrates the application of in vitro toxicity assessment using human cells for the prediction of toxic risk as a
sentinel to human toxicity from a contaminated site.