Chemiluminescence is the phenomenon of energy released as light when the excited state of molecules by oxidation returns to the ground state. Organic substances which exhibit chemiluminescene are Cypridina luciferin, firefly luciferin, oxalate, luminol and lucigenin etc.
Cypridina luciferin analogs achieve chemiluminescence through reaction with superoxide (O2–) or singlet oxygen (1O2). Using this characteristic, CLA and MCLA have been used in the research of the functions of leukocytes. The maximum emission wavelengths of CLA and MCLA are 380nm and 465nm, respectively. FCLA developed by Goto et al. is characterized by emitting light at a longer wavelength (532nm). Furthermore, Red-CLA developed by Teranishi emits at an even longer wavelength. Red-CLA exhibits high emission intensity by reaction with superoxide, and can be used for efficient analysis of superoxide at the longest wavelength (610nm).
Firefly luciferin reacts with ATP in the presence of luciferase and magnesium ion to provide oxyluciferin via luciferyl-adenylic acid. The light with a wavelength of 562nm is emitted when activated molecules return to the ground state by decomposition of oxyluciferin. Using this characteristic, firefly luciferin is used in the trace detection of ATP and the activity measurement of nucleotide phosphatase etc.
Luminol reacts with hydrogen peroxide in the presence of metals such as iron or its complexes to emit strong blue light with a wavelength of 460nm. This reaction is called the luminol test and is applied to the identification of blood stains in forensic science. Luminol is also used for trace detection of hydrogen peroxide and metals which catalyze this reaction.
On the other hand, oxalates are oxidized by hydrogen peroxide etc. to produce 1,2-dioxetanediones. When these substances are decomposed, they transfer energy to coexisting fluorescent substances which are elevated to the excited state. These excited fluorescent substances emit light during relaxation to the ground state. Thus, the emission wavelength can be changed by choosing coexisting fluorescent substances. This technique is applied to HPLC detection systems.