Chemiluminescence method

Molecular luminescence spectrometry

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Chinese name: Chemiluminescence method
Discipline: analytical chemistry
English translation: ChemiLuminescence method

Abbreviation: CL
Nature: A Class of Molecular Luminescence Spectroscopy
Industrial application: Trace metal ions, various inorganic compounds, etc

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Fundamentals

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Chemiluminescence (CL for short) is molecular luminescence Spectral analysis It is mainly based on the principle that the concentration of the substance to be measured in the chemical detection system has a linear quantitative relationship with the chemiluminescence intensity of the system under certain conditions. It is a trace analysis method that uses the instrument to detect the chemiluminescence intensity of the system and determine the content of the substance to be measured. The essential difference between chemiluminescence and other luminescence analysis is that the energy sources absorbed by the system to generate luminescence (light radiation) are different. To generate chemiluminescence, the system must have a light radiation reaction that generates detectable signals and a chemical reaction that can provide a single reaction step with sufficient energy to cause the luminescence phenomenon at one time^[1] 。

Luminescence classification

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According to the characteristics of energy supply reaction Chemiluminescence analysis Divided into:

1) Ordinary chemiluminescence analysis (energy supply reaction is ordinary chemical reaction);

2) Biochemiluminescence analysis (energy supply reaction is biochemical reaction (BCL for short);

3) Electrochemiluminescence analysis (energy supply reaction is electrochemical reaction, abbreviated as ECL), etc.

According to the determination method, the method can be divided into:

1) Direct determination of CL analytical method;

2) Coupling reaction CL analysis method (determine a component in the system by coupling reaction);

3) Time resolved CL analysis (that is, multi-component determination is realized by using the time difference of multi-component effects on the same chemiluminescence reaction);

4) CL analysis of solid phase, gas phase and liquid phase;

5) Enzyme-linked immunosorbent assay^[2] 。

Luminescent agent and principle

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Chemiluminescence is the light radiation produced by the absorption of chemical energy by certain material molecules. Any chemiluminescence reaction includes two key steps, namely chemical excitation and luminescence. Therefore, two conditions must be met for a chemical reaction to become a luminescence reaction: first, the reaction must provide enough energy (170~300KJ/mol); second, these chemical energies must be able to be absorbed by some material molecule to produce an electronic excited state, and there must be enough Fluorescence quantum yield 。 Most of the chemiluminescence reactions studied are Redox reaction And most of them are liquid phase chemiluminescence reactions.

The luminous efficiency of chemiluminescence reaction refers to the ratio of the number of luminous components in the reaction to the number of molecules participating in the reaction. For general chemiluminescence reaction, the value is about 10 -6 Typical luminous agents, such as Luminol, have a luminous efficiency of 0.01, and the luminous reactions with luminous efficiency greater than 0.01 are rare. Several commonly used light-emitting agents with high luminous efficiency and their luminous mechanisms are summarized as follows.

Luminol and its derivatives

The derivatives of luminol mainly include isoluminol, 4-aminohexyl-N-ethyl isoluminol, AHEI, ABEI, etc. Luminol can be oxidized by some oxidants under alkaline conditions to generate chemiluminescence reaction, and emit chemiluminescence with the maximum emission wavelength of 425nm.

Under normal circumstances, Lumino and hydrogen peroxide Chemiluminescence reaction is quite slow, but the reaction is very fast when there are some catalysts. The most commonly used catalyst is metal ions. In a large concentration range, the concentration of metal ions is proportional to the luminous intensity, which can be used for chemiluminescence analysis of some metal ions. This reaction can be used to analyze organic compounds containing metal ions, achieving high sensitivity. Secondly, the inhibition of organic compounds on luminol chemiluminescence reaction was used to determine the organic compounds with quenching effect on chemiluminescence reaction. The third is the indirect determination of inorganic or organic compounds through coupling reaction. The fourth is to label derivatives of luminol, such as isoluminol (ABEI), onto carboxylic acids and ammonia compounds, and then react with hydrogen peroxide potassium ferricyanide under alkaline conditions for chemiluminescence detection after separation by high performance liquid chromatography or liquid chromatography. Other separation methods can also be used, such as labeling the newly synthesized chemiluminescence reagent isoluminol isothiocyanate to yeast RNA, centrifugation and dialysis, and then chemiluminescence detection. In addition, N two （B two Carboxypropionyl) isoluminol, and its properties were studied.

Lustre essence

Glossy essence exists in the form of nitrate. In alkaline medium, hydrogen peroxide Oxidize it into four membered ring peroxide The intermediate is then cracked to generate an excited pyridinone and emit light. It can be used to determine some important reducing substances in clinical medicine, such as ascorbic acid , creatinine glutathione 、 Glucuronic acid , lactose, glucose.

Lofarine

Luo powder is the earliest chemiluminescence reagent recorded in the literature, but it has not been applied until Marino et al. applied it to the determination of Co in 1979. This reagent has been used for the analysis and determination of many elements.

Oxalates peroxide

Oxalate chemiluminescence reactions mostly generate peroxyoxalyl intermediates, so these reactions are also called peroxyoxalyl chemiluminescence reactions. The application of peroxyoxalate chemiluminescence analysis also depends on the development of new fluorescent derivative reagents.

acridinium ester

McCapr et al. synthesized a series of acridine ester compounds. Research on the chemiluminescence mechanism of this kind of reagent shows that the luminous efficiency is closely related to the pKa of the dissociatable acid group in the reagent, and pKa should generally be less than 11. Acridine ester compounds are a kind of promising non radioactive nucleic acid probe markers. They are used as DNA light-emitting probes with high luminous quantum yield and good stability. The markers have no effect on the kinetics of hybridization reaction and the stability of hybrids, and can directly conduct chemiluminescence reaction in alkaline medium.

The above five Chemiluminescent agent Chemiluminescence has high quantum yield, stable aqueous solution, and can be directly oxidized by a variety of oxidants to give off light. It can also be given off by many metals above the catalytic luminescence reaction. Many inorganic, organic and biochemical components can also enhance or inhibit their luminescence, so it is widely used. It is reported that o-phenanthroline, base salicylic acid, rhodamine B gallic acid , coumarin, cortin, alizarin violet, hematoxylin essence, Peihuaqing, triphenylmethane dyes, acetone, ethanol, hydroxylamine, etc. These reagents are highly commercialized, inexpensive, and convenient to use, but their chemiluminescence quantum yields are low. Therefore, it is very critical to study sensitizers to improve their chemiluminescence quantum yields.