Analytical process for testing mixtures for toxic constituents

Details Analytical process for testing mixtures for toxic constituents Analytical process for testing mixtures for toxic constituents

1993-09-02

2001-05-29

Bhat, Nina (Department: 1761)

Chemistry: analytical and immunological testing

Including chromatography

Utilizing paper or thin layer plate

C436S172000, C436S178000, C422S070000, C435S252100, C210S198300, C210S615000, C210S658000

Reexamination Certificate

active

06238928

ABSTRACT:

For many years, luminescent bacteria have been used in the toxicity testing of chemical compounds (see, for example, A. A. Bulich; G. Bitton, B. J. Dutkay, “Toxicity Testing Using Microorganisms”, CRC Press, Boca-Raton, Fla., USA, pages 57 to 74). The principle of the luminescent bacteria test is based on substances toxic to bacteria leading to a reduction in the bioluminescence produced by the bacteria. This reduction is evaluated by measurement. In the case of a mixture of various chemical compounds, it has hitherto only been possible to evaluate the (integral) toxicity of the mixture as a whole. No information could be obtained as to the toxicity of the individual components involved.
Now, the problem addressed by the present invention was to provide a process for testing mixtures for toxic constituents based on the luminescent bacteria test which would enable the measured toxicities to be assigned to the individual components of the mixture. At the same time, the important boundary condition that the analytical process itself would not have any harmful effect on the luminescent bacteria would be satisfied.
According to the invention, the solution to this problem is characterized in that the mixture to be tested is first separated into fractions by chromatography, the separated fractions are contacted with luminescent microorganisms in the chromatographic system itself and the toxicity of the fractions is determined by measurement of the bioluminescence.
The fractions are advantageously separated by thin layer chromatography or column liquid chromatography. In the case of thin layer chromatography, the TLC plate is wetted with a suspension of luminescent microorganisms and the local bioluminescence assigned to the individual fractions is measured.
To this end, the TLC plate is advantageously contacted with a photographic film and the bioluminescence assigned to the individual fractions is evaluated on the exposed film either visually or by densitometry.
Where separation is carried out by column liquid chromatography, the suspension of luminescent microorganisms is continuously mixed with the eluate from the chromatographic column and the bioluminescence of the mixture is measured by means of a throughflow photometer.
The invention affords the following advantages:
1. Whereas conventional detection processes based on chromatography provide information on the identity, quantity and/or chemical structure of a mixture component, the process according to the invention provides direct information on the biological effect of a certain component of the mixture, i.e. the toxicity of a mixture can be directly assigned to discrete individual components of that mixture. The quality of the information obtainable by chromatographic separations is thus improved.
2. The selective detection of toxicity enables the effort and costs involved in the structural elucidation of unknown toxic compounds in mixtures to be considerably reduced because the work involved in structural elucidation (for example by isolation and subsequent spectroscopic examination) can be concentrated on those components of the mixture to which a toxic effect can be assigned on the basis of the toxicity detection results. Potential applications of this analytical process include the structural elucidation of unknown toxic components in wastewaters and the detection of biocides in various products.
The invention is illustrated by the following Examples in conjunction with the accompanying drawing.


REFERENCES:
patent: 3370175 (1968-02-01), Jordon et al.
patent: 4357420 (1982-11-01), Bostick et al.
patent: 4581335 (1986-04-01), Baldwin
patent: 4879249 (1989-11-01), Baldwin et al.
patent: 0293775 (1988-12-01), None
patent: 85/00890 (1985-02-01), None
patent: 90/04037 (1990-04-01), None
“Detection . . . Plates”; Bjurseth et al, Science, vol. 215 Jan. 1, 1982.

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