Microtox

Kamlet, M. J., Doherty, R. M., Veith, G. D., Taft, R. W., Abraham, M. H. (1986) Solubility properties in polymers and biological media. 7. An analysis toxicant properties that influence inhibition of bioluminescence in Photobacterium phosphoreum (the Microtox test). Environ. Sci. Technol. 20, 690-695. 

Toxicity in estuarine sediments—use of Mutatox and Microtox to evalu- 173 ate the acute toxicity and genotoxicity of organic sediments Toxicity tests for the analysis of pore water sediment a comparison of 4 174... 

Toxicity of sediments comparison between Photobacterium phosphoreum 177 and Microtox system... 

Parrott, J.L. and J.B. Sprague. 1993. Patterns in toxicity of sublethal mixtures of metals and organic chemicals determined by Microtox and by DNA, RNA, and protein content of fathead minnows (Pimephales promelas). Canad. Jour. Fish. Aquat. Sci. 50 2245-2253. 

Argese, E., C. Bettiol, A.V. Ghirardini, M. Fasolo, G. Giurin, and P.F. Ghetti. 1998. Comparison of in vitro submitochondrial particle and Microtox assays for determining the toxicity of organotin compounds. Environ. Toxicol. Chem. 17 1005-1012. 

Middaugh, D.P., S.M. Resnick, S.E. Lantz, C.S. Heard, and J.G. Mueller. 1993. Toxicological assessment of biodegraded pentachlorophenol Microtox and fish embryos. Arch. Environ. Contam. Toxicol. 24 165-172. 

But the most-used toxicity tests are based on bioluminescence inhibition, the responses of which are sometimes difficult to interpret particularly for waste-water quality monitoring. A comparison between a bioluminescence test kit (Microtox) and a respirometry approach for the toxicity study of seven organic and five inorganic toxic compounds was performed [54]. The bioluminescent response proved to have a higher sensitivity to toxicants but was less representative of the effects on activated sludge compared to respirometry, due to the nature of the microorganisms involved in each procedure. 

Recent studies, including the use of Microtox and ToxAlert test kits [55,56], were carried out for the determination of the toxicity of some non-ionic surfactants and other compounds (aromatic hydrocarbons, endocrine disruptors) before implementation on raw and treated wastewater, followed by the identification and quantification of polar organic cytotoxic substances for samples with more than 20% inhibition. Furthermore, the study of their contribution to the total toxicity was obtained using sequential solid-phase extraction (SSPE) before liquid chromatography-mass spectrometry (LC-MS) detection. This combined procedure allows one to focus only on samples containing toxic substances. 

The toxicity of chlorinated compounds is connected with the chlorine content. The decrease of toxicity of chlorophenols with decreasing number of chloro substituents, expressed as EC50, using the Microtox method [291], is shown in Table 21a. [290], cf. also Refs. [292,293],... 

When using purified triolein, most samples are amenable to bioassay after di-alytic enrichment. For example, Microtox bioassay of dialysates of SPMDs shows that the SPMDs made with the purified triolein have lower acute toxicities than dialysates from SPMDs made from unpurified triolein (Johnson, 2001). Finally, examination of the dialysates using the yeast estrogen screen (YES) assay (Routledge and Sumpter, 1996) demonstrated that the purification procedure removes all background estrogenic activity (Lebo et ah, 2004). Use of triolein purified by this process expands the potential applicability of SPMD sample extracts to include numerous bioassay procedures (see Chapter 6) and GC-MS as a standard analysis technique. 

Table 6.1 Toxicological Evaluation of Polycyclic Aromatic Hydrocarbons (PAHs) with Microtox Basic Test and Mutatox. Reprinted with permission from the American Petroleum Institute (Huckins et al., 2002)...

Sample type Microtox toxicity ECso Mutatox genotoxicity... 

Freshly prepared blank SPMD, carried through Microtox and Mutatox test. 

Microtox values are 5-minute EC50s with 95% confidence intervals (in parentheses). 

Microtox responds well to a wide array of hydrophobic chemicals (see the work of Johnson [1998] in which a variety of pesticides, industrial chemicals and petroleum products were tested). Apparently, Microtox is highly responsive to compounds with a narcosis mode of toxicity (Johnson, 1998 Sabaliunas et al., 1998). As shown by Johnson (1998) chemicals that elicit narcosis include multiple chemical classes. 

Sabaliunas et al. (2000) showed that a Daphnia pulex immobilization test (Daphtoxkit F) was far more sensitive than Microtox to a mixture of insecticides seqnestered in SPMDs. This is not surprising because the OCPs and pyrethroid pesticides present in the enriched SPMD extracts are neurotoxins, and the effect thresholds can be much lower than narcosis-type toxicants. Thus, if insecticides are the contaminants of concern, the Daphtoxkit F approach may have some advantage over Microtox. 

Using a preemptive approach, Lebo et al. (2004) have shown that oleic acid and methyl oleate can be removed from triolein prior to use of the triolein in SPMDs (see Chapters 4 and 5). Dialysates from SPMDs prepared using triolein purified by the Lebo et al. (2004) method exhibited lower acute toxicity (Microtox assay) than SPMDs prepared with unpurified triolein. Also, the YES assay demonstrated that the purification method had removed all background estrogenic activity from SPMD extracts. Eor these reasons, the use of triolein purified by the method of Lebo et al. (2004) is standard for all SPMD studies conducted at CERC, USGS. Also, SPMDs with triolein purified by the Lebo et al. (2004) method are available from the commercial vendor upon request. 

Cleveland, L. Little, E.E. Petty, J.D. Johnson, B.T. Lebo, J.A. Orazio, C.E. Dionne. J. Crockett, A. 1997, Toxicological and chemical screening of Antarctica sediments Use of whole sediment toxicity tests, Microtox, Mutatox, and semipermeable membrane devices (SPMDs). Mar. Pollut. Bull 34 194-202. 

Johnson, B.T. 1998, Microtox Toxicity Test System—New Developments and Applications. In Microscale Testing in Aquatic Toxicology Wells, P.G., Lee, K., Blaise, C., Eds CRC Press Washington, D.C. pp. 201-218. 

Microbics Corporation 1992, Condensed Protocol for Basic Test, Using Organic Solvent Sample Solubilization. Microtox Manual, Vol. Ill Condensed Protocols, Carlsbad, CA 13, 226-232. 

Gustavson, K.E., Svenson, A., and Harkin, J.M. Comparison of toxicities and mechanism of action of / -alkanols in the submitochondrial particle and the Vibrio fischeri bioluminescence (Microtox ) bioassay. Environ. Toxicol. Chem., 17(10) 1917-1921, 1998. 

Somasundaram, L.. Coats, J.R., Racke. K.D., and Stahr, H.M. Application of the Microtox system to assess the toxicity of pesticides and their hydrolysis metabolites. Bull Environ. Contam. Toxicol, 44(2) 254-259,1990. 

The first steps in bypassing of the biological, technological, and financial burden of live stock culturing or maintenance were made more than 20 years ago through the development of a bacterial luminescence inhibition test [34,35] this bioassay is presently known and used worldwide as the Microtox test. The revolutionary principle of this test is that it uses a lyophihzed strain of a (marine) bacterium Photobacterium phosphoreum). This makes the bioassay apphcable anytime, anywhere, without the need for continuous culturing of the test species. 

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