Chemiluminescence

Not to be confused with Fluorescence.

A chemoluminescent reaction in an Erlenmeyer flask

Chemiluminescence (also chemoluminescence) is the emission of light (luminescence) as the result of a chemical reaction. There may also be limited emission of heat. Given reactants A and B, with an excited intermediate ◊,

[A] + [B] → [◊] → [Products] + light

For example, if [A] is luminol and [B] is hydrogen peroxide in the presence of a suitable catalyst we have:

${\ce {{\underset {luminol}{C8H7N3O2}}+{\underset {hydrogen\ peroxide}{H2O2}}->3-APA[\lozenge ]->{3-APA}+light}}$

where:

• 3-APA is 3-aminophthalate
• 3-APA[◊] is the vibronic excited state fluorescing as it decays to a lower energy level.

General description

The decay of this excited state[◊] to a lower energy level causes light emission. In theory, one photon of light should be given off for each molecule of reactant. This is equivalent to Avogadro's number of photons per mole of reactant. In actual practice, non-enzymatic reactions seldom exceed 1% Q_C, quantum efficiency.

In a chemical reaction, reactants collide to form a transition state, the enthalpic maximum in a reaction coordinate diagram, which proceeds to the product. Normally, reactants form products of lesser chemical energy. The difference in energy between reactants and products, represented as $\Delta H_{rxn}$, is turned into heat, physically realized as excitations in the vibrational state of the normal modes of the product. Since vibrational energy is generally much greater than the thermal agitation, it rapidly disperses in the solvent through molecular rotation. This is how exothermic reactions make their solutions hotter. In a chemiluminescent reaction, the direct product of the reaction is an excited electronic state. This state then decays into an electronic ground state and emits light through either an allowed transition (analogous to fluorescence) or a forbidden transition (analogous to phosphorescence), depending partly on the spin state of the electronic excited state formed.

Chemiluminescence differs from fluorescence or phosphorescence in that the electronic excited state is the product of a chemical reaction rather than of the absorption of a photon. It is the antithesis of a photochemical reaction, in which light is used to drive an endothermic chemical reaction. Here, light is generated from a chemically exothermic reaction. The chemiluminescence might be also induced by an electrochemical stimulus, in this case is called electrochemiluminescence.

Bioluminescence in nature: A male firefly mating with a female of the species Lampyris noctiluca.

The first chemiluminescent compound to be discovered was 2,4,5-triphenylimidazole (lophine), which was reported, in 1877, to emit light when mixed with potassium hydroxide in aqueous ethanol in the presence of air. A standard example of chemiluminescence in the laboratory setting is the luminol test. Here, blood is indicated by luminescence upon contact with iron in hemoglobin. When chemiluminescence takes place in living organisms, the phenomenon is called bioluminescence. A light stick emits light by chemiluminescence.

Liquid-phase reactions

Chemiluminescence in aqueous system is mainly caused by redox reactions.

Chemiluminescence after a reaction of hydrogen peroxide and luminol

• Luminol in an alkaline solution with hydrogen peroxide in the presence of iron or copper, or an auxiliary oxidant, produces chemiluminescence. The luminol reaction is

${\ce {{\underset {luminol}{C8H7N3O2}}+{\underset {hydrogen\ peroxide}{H2O2}}->3-APA[\lozenge ]->{3-APA}+light}}$

Gas-phase reactions

Green and blue glow sticks

• One of the oldest known chemiluminescent reactions is that of elemental white phosphorus oxidizing in moist air, producing a green glow. This is a gas-phase reaction of phosphorus vapor, above the solid, with oxygen producing the excited states (PO)₂ and HPO.
• Another gas phase reaction is the basis of nitric oxide detection in commercial analytic instruments applied to environmental air-quality testing. Ozone is combined with nitric oxide to form nitrogen dioxide in an activated state.

NO+O₃ → NO₂[◊]+ O₂

The activated NO₂[◊] luminesces broadband visible to infrared light as it reverts to a lower energy state. A photomultiplier and associated electronics counts the photons that are proportional to the amount of NO present. To determine the amount of nitrogen dioxide, NO₂, in a sample (containing no NO) it must first be converted to nitric oxide, NO, by passing the sample through a converter before the above ozone activation reaction is applied. The ozone reaction produces a photon count proportional to NO that is proportional to NO₂ before it was converted to NO. In the case of a mixed sample that contains both NO and NO₂, the above reaction yields the amount of NO and NO₂ combined in the air sample, assuming that the sample is passed through the converter. If the mixed sample is not passed through the converter, the ozone reaction produces activated NO₂[◊] only in proportion to the NO in the sample. The NO₂ in the sample is not activated by the ozone reaction. Though unactivated NO₂ is present with the activated NO₂[◊], photons are emitted only by the activated species that is proportional to original NO. Final step: Subtract NO from (NO + NO₂) to yield NO₂

Infrared chemiluminescence

In chemical kinetics, infrared chemiluminiscence (IRCL) refers to the emission of infrared photons from vibrationally excited product molecules immediately after their formation. The intensities of infrared emission lines from vibrationally excited molecules are used to measure the populations of vibrational states of product molecules.

The observation of IRCL was developed as a kinetic technique by John Polanyi, who used it to study the attractive or repulsive nature of the potential energy surface for gas-phase reactions. In general the IRCL is much more intense for reactions with an attractive surface, indicating that this type of surface leads to energy deposition in vibrational excitation. In contrast reactions with a repulsive potential energy surface lead to little IRCL, indicating that the energy is primarily deposited as translational energy.

Enhanced chemiluminescence

Enhanced chemiluminescence is a common technique for a variety of detection assays in biology. A horseradish peroxidase enzyme (HRP) is tethered to an antibody that specifically recognizes the molecule of interest. This enzyme complex then catalyzes the conversion of the enhanced chemiluminescent substrate into a sensitized reagent in the vicinity of the molecule of interest, which on further oxidation by hydrogen peroxide, produces a triplet (excited) carbonyl, which emits light when it decays to the singlet carbonyl. Enhanced chemiluminescence allows detection of minute quantities of a biomolecule. Proteins can be detected down to femtomole quantities, well below the detection limit for most assay systems.

Applications

• Gas analysis: for determining small amounts of impurities or poisons in air. Other compounds can also be determined by this method (ozone, N-oxides, S-compounds). A typical example is NO determination with detection limits down to 1 ppb. Highly specialised chemiluminescence detectors have been used recently to determine concentrations as well as fluxes of NOx with detection limits as low as 5 ppt.
• Analysis of inorganic species in liquid phase
• Analysis of organic species: useful with enzymes, where the substrate is not directly involved in the chemiluminescence reaction, but the product is
• Detection and assay of biomolecules in systems such as ELISA and Western blots
• DNA sequencing using pyrosequencing
• Lighting objects. Chemiluminescence kites, emergency lighting, glow sticks (party decorations).
• Combustion analysis: Certain radical species (such as CH* and OH*) give off radiation at specific wavelengths. The heat release rate is calculated by measuring the amount of light radiated from a flame at those wavelengths.
• Children's toys.
• Glow sticks.

Biological applications

Chemiluminescence has been applied by forensic scientists to solve crimes. In this case, they use luminol and hydrogen peroxide. The iron from the blood acts as a catalyst and reacts with the luminol and hydrogen peroxide to produce blue light for about 30 seconds. Because only a small amount of iron is required for chemiluminescence, trace amounts of blood are sufficient.

In biomedical research, the protein that gives fireflies their glow and its co-factor, luciferin, are used to produce red light through the consumption of ATP. This reaction is used in many applications, including the effectiveness of cancer drugs that choke off a tumor's blood supply. This form of bioluminescence imaging allows scientists to test drugs in the pre-clinical stages cheaply. Another protein, aequorin, found in certain jellyfish, produces blue light in the presence of calcium. It can be used in molecular biology to assess calcium levels in cells. What these biological reactions have in common is their use of adenosine triphosphate (ATP) as an energy source. Though the structure of the molecules that produce luminescence is different for each species, they are given the generic name of luciferin. Firefly luciferin can be oxidized to produce an excited complex. Once it falls back down to a ground state a photon is released. It is very similar to the reaction with luminol.

${\ce {Luciferin{}+O2{}+ATP->[{\text{Luciferase}}]Oxyluciferin{}+CO2{}+AMP{}+PPi{}+light}}$

Many organisms have evolved to produce light in a range of colors. At the molecular level, the difference in color arises from the degree of conjugation of the molecule, when an electron drops down from the excited state to the ground state. Deep sea organisms have evolved to produce light to lure and catch prey, as camouflage, or to attract others. Some bacteria even use bioluminescence to communicate. The common colors for the light emitted by these animals are blue and green because they have shorter wavelength than red and can transmit more easily in water.

In April 2020, researchers reported to have genetically engineered plants to glow much brighter than previously possible by inserting genes of the bioluminescent mushroom Neonothopanus nambi. The glow is self-sustained, works by converting plants' caffeic acid into luciferin and, unlike for bacterial bioluminescence genes used earlier, has a relatively high light output that is visible to the naked eye.

Chemiluminescence is different from fluorescence. Hence the application of fluorescent proteins such as Green fluorescent protein is not a biological application of chemiluminescence.

See also

• Eclox
• Electrochemiluminescence
• List of light sources
• Lyoluminescence
• Will-o'-the-wisp