Chemistry: analytical and immunological testing – Nuclear magnetic resonance – electron spin resonance or other...
Patent
1986-09-30
1989-01-31
Castel, Benoit
Chemistry: analytical and immunological testing
Nuclear magnetic resonance, electron spin resonance or other...
422 68, 436149, G01J 342, G01N 2162
Patent
active
048015520
DESCRIPTION:
BRIEF SUMMARY
The invention relates to a method for measuring absorbance difference spectra of radical reactions using a modulated perturbation of the concentration of reactants in, for example, radical complexes induced by radiation, the yield of substances and/or molecular states produced by recombination and by escaping radicals being influenced. The invention also relates to a spectrometer for carrying out the above-named method, which spectrometer is provided with means for radiation of the material to be investigated, means for receiving the said material, cooling and/or heating means, and detection means for the detection of the resulting absorbance spectrum of the said material.
The above method and spectrometer are known from the article by H. J. den Blanken and A. J. Hoff in Biochem. Biophys. Acta 681 (1982), pages 365-374. In this known method the magnetic resonance of intermediate triplet states in photosynthetic material is detected by means of triplet-minus-singlet absorbance difference (T-S) spectra. In this process a modulated perturbation of the concentration of intermediate triplet states is produced by means of irradiated microwaves which resonate between two of the triplet sublevels. The resulting change in the absorbance is then detected and recorded as a function of wavelength. In this manner an absorbance difference spectrum is obtained and recorded which reflects the bleaching of tripletcarrying pigments, and bandshifts and the appearance of new bands due to triplet-induced changes in the electrostatic interaction between pigments which adjoin the pigment carrying the triplet state.
Although good results have been obtained with the above method in the investigation of reactions in photosynthetic pigment materials such as plants, alga or bacteria, this method can be performed only in a limited range of temperature at very low temperatures near the absolute zero point to avoid spin-lattice relaxation between the triplet sublevels. As a result of this, temperature-induced changes in the triplet-minus-singlet absorbance difference spectrum, which reflect changes in pigment configuration or the presence of other excited states such as charge transfer states cannot be detected. In addition, this method is limited only to investigation of reactions in which intermediate triplet states are present.
The object of the invention is to eliminate the above-named problems and to provide a method and a spectrometer with which absorbance difference spectra of radical complexes can be measured over a wide temperature range with great accuracy so that all the substances taking part in a radical reaction can be investigated.
According to the invention this is achieved in a method of the type mentioned in the introduction in the manner that a modulated magnetic field is exerted on the material and in that the changes in the absorbance of the material generated as a result thereof are phase-sensitively detected over a wide frequency range, it being possible to vary the temperature of the material over a large adjustment range which may include room temperature.
According to the invention the spectrometer mentioned in the introduction is further characterized by a device for exerting a modulated magnetic field on the material, the detection means being provided with a phase-sensitive detector which converts the changes in absorbance generated into a dc absorbance, the cooling and/or heating means being adjusted to a temperature in a wide temperature range.
With the method and spectrometer according to the invention radical reactions can be investigated with advantage to determine, for example, the difference in concentration of reactants. The present so-called magneto-optical difference spectroscopy (MODS) measurements can be performed with great accuracy; thus, a signal
oise ratio is obtained which may be 1,000 times better than in the prior art. This method can also be used in any frequency range where absorbance of radiation occurs.
Thus, with the present method absorbance difference spectra have been measured of
REFERENCES:
patent: 3442592 (1969-05-01), Grosjean
Hoff et al.; Magneto-Optical Absorbance Difference Spectroscopy; Chem. Phys. Lett. vol. 114, No. 1, 15 Feb. 1985, pp. 39-43.
Ke et al.; Versatile Spectrophotometer for Photosynthesis and Other Biophysical Measurements; Rev. Sci. Instrum. 56(1) Jan. 1985, pp. 26-31.
Den Blanken et al.; High-Resolution Optical Absorption-Difference Spectra of the Triplet State of the Primary Donor in Isolated Reaction Centers of the Photosynthetic Bactria Rhodopseudomonas Sphaeroides R-26 and Rhodopseudomonas Viridis Measured with Optically Detected Magnetic Resonance at 1.2K. Biochim. Biophysica. Acta. 681 (1982) pp. 365-374.
Kolesnikova et al.; Study of Charge Separation at the Centers of Photosystem II of Higher Plants by Magnetic Modulation, Abs. 103807 1982.
Bartiromo et al.; An Apparatus for Magneto-Optical Spectroscopy by Magnetic Modulation, J. of Phys. E. 10, 1977.
Alexander Lyle Alfandary
Castel Benoit
Ryksuniversiteit Leiden
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