Microsecond time-resolved mass spectrometry

a time-resolved mass spectrometry and microsecond technology, applied in material analysis, instruments, electric discharge tubes, etc., can solve the problems of chromophoric substrates or radioactive labeling, and achieve the effect of reducing the number of radioactive components

Inactive Publication Date: 2014-05-29
OHIO UNIV
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Problems solved by technology

The protein conformational changes or the formation of enzyme-substrate complex intermediates may occur within microseconds, making the d

Method used

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  • Microsecond time-resolved mass spectrometry
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  • Microsecond time-resolved mass spectrometry

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example 1

[0042]The rate of acid-induced unfolding of cytochrome c was investigated using a device similar to the exemplary embodiment of FIG. 1A. A rate of conformational change for cytochrome c may be generally expressed as:

FU

where F designates the folded protein conformation and U designates the unfolded protein conformation.

[0043]In that regard, a sample of cytochrome c in H2O was loaded into the first syringe 52 and a 4% acetic acid solution was loaded into the second syringe 54. Fast mixing occurred within the Tee-mixer 64. The liquid jet 50 was sampled for DESI-MS analysis at Ljet, distances of about 0.5 cm, 1.0 cm, 1.5 cm, 2.0 cm, 2.5 cm, and 3.0 cm, which are calculated to correspond to reaction times of 10.5 ms, 12.0 ms, 13.5 ms, 15.0 ms, 16.5 ms, and 18.0 ms, respectively.

[0044]FIG. 3A illustrates an exemplary spectrum acquired at Ljet=0.5 cm (10.5 ms) and demonstrates dominating peaks having m / z values ranging between +7 and +10, corresponding to the native protein ions, i.e., fol...

example 2

[0047]The rate of L-ascorbic acid (“L-AA”) induced reduction of 2,6-dichlorophenolindophenol (“DCIP”) was investigated using a device that is similar to the exemplary embodiment of FIG. 2. The reaction under investigation may be expressed as follows:

[0048]A solution of 40 mM LAA in 20 mM HCl and a solution of 0.2 mM DCIP in H2O were pumped via the HPLC pumps 84, 86 at a flow rate of 1.0 mL / min into the Tee-mixer 64 to produce a continuous jet 50 having a pH of 2.0. Desorption and ioniziation occurred with the DESI sprayer 10 having a spray solvent 26 of 20 mM aqueous NH4OAc buffer having a pH of 7.0 under nitrogen gas. An LCQ DECA mass spectrometer 30 (Thermo Finnigan, San Jose, Calif., USA) was used for mass analysis.

[0049]A vertical micrometer adjusted the height of Tee-mixer 64 relative to the DESI sprayer 10 and the mass spectrometer 30 for DESI-MS analysis. Ljet distances of 1.0 cm, 1.5 cm, 2.0 cm, 2.5 cm, 3.0 cm, 3.5 cm, and 4.0 cm were calculated to correspond to reaction tim...

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Abstract

A microsecond time-resolved mass spectrometry device and method of using desorption electrospray ionization (10) was described for the kinetic study of fast reactions. The device includes a liquid jet generator (64) that is configured to emit a continuous liquid jet (50) having a length. An ambient ionization source (10) is directed toward a selected variable location along the continuous liquid jet (50) to desorb at least a portion of the continuous liquid jet (50). A mass analyzer (30) analyzes a mass-to-charge ratio of an ionized sample that is within the desorbed portion of the continuous liquid jet (50). The acquired mass spectra reflect the reaction progress in different reaction times and, therefore, may be used to derive the reaction rate.

Description

TECHNICAL FIELD[0001]The invention is with regard to the apparatus and methodology of time-resolved mass spectrometry.BACKGROUND OF THE INVENTION[0002]Chemical kinetics is the study of rates at which a chemical reaction occurs. Such an investigation provides understanding of the reaction mechanisms along with the identification of possible reaction intermediates.[0003]Of particular interest are biochemical reactions, such as protein conformational change and enzymatic catalysis, which are responsible for many important biochemical and physiological processes. The protein conformational changes or the formation of enzyme-substrate complex intermediates may occur within microseconds, making the detection of intermediates technically challenging.[0004]Conventional techniques for kinetic study generally involve spectroscopic methods, such as UV and fluorescence spectroscopy. These methods have the benefit of high time resolution, for example, femtosecond or attosecond time scales. Howev...

Claims

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Application Information

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IPC IPC(8): G01N27/68G01N33/487
CPCG01N33/48707G01N27/68G01N1/00H01J49/0445H01J49/145H01J49/165H01J49/045G01N33/6848
Inventor CHEN, HAOMIAO, ZHIXIN
Owner OHIO UNIV
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