Luminol (C
8H
7N
3O
2) is a
chemical
A chemical substance is a form of matter having constant chemical composition and characteristic properties. Some references add that chemical substance cannot be separated into its constituent elements by physical separation methods, i.e., wi ...
that exhibits
chemiluminescence, with a blue glow, when mixed with an appropriate
oxidizing agent
An oxidizing agent (also known as an oxidant, oxidizer, electron recipient, or electron acceptor) is a substance in a redox chemical reaction that gains or "Electron acceptor, accepts"/"receives" an electron from a (called the , , or ). In ot ...
. Luminol is a white-to-pale-yellow crystalline solid that is soluble in most polar organic solvents, but insoluble in water.
Forensic
Forensic science, also known as criminalistics, is the application of science to Criminal law, criminal and Civil law (legal system), civil laws, mainly—on the criminal side—during criminal investigation, as governed by the legal standard ...
investigators use luminol to detect trace amounts of
blood
Blood is a body fluid in the circulatory system of humans and other vertebrates that delivers necessary substances such as nutrients and oxygen to the cells, and transports metabolic waste products away from those same cells. Blood in the c ...
at
crime scene
A crime scene is any location that may be associated with a committed crime. Crime scenes contain physical evidence that is pertinent to a criminal investigation. This evidence is collected by crime scene investigators (CSI) and law enforcement ...
s, as it reacts with the iron in
hemoglobin
Hemoglobin (haemoglobin BrE) (from the Greek word αἷμα, ''haîma'' 'blood' + Latin ''globus'' 'ball, sphere' + ''-in'') (), abbreviated Hb or Hgb, is the iron-containing oxygen-transport metalloprotein present in red blood cells (erythrocyte ...
. Biologists use it in cellular
assays
An assay is an investigative (analytic) procedure in laboratory medicine, mining, pharmacology, environmental biology and molecular biology for qualitatively assessing or quantitatively measuring the presence, amount, or functional activity of a ...
to detect
copper
Copper is a chemical element with the symbol Cu (from la, cuprum) and atomic number 29. It is a soft, malleable, and ductile metal with very high thermal and electrical conductivity. A freshly exposed surface of pure copper has a pinkis ...
,
iron
Iron () is a chemical element with symbol Fe (from la, ferrum) and atomic number 26. It is a metal that belongs to the first transition series and group 8 of the periodic table. It is, by mass, the most common element on Earth, right in f ...
,
cyanide
Cyanide is a naturally occurring, rapidly acting, toxic chemical that can exist in many different forms.
In chemistry, a cyanide () is a chemical compound that contains a functional group. This group, known as the cyano group, consists of ...
s, as well as specific
proteins
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, respo ...
via
western blot
The western blot (sometimes called the protein immunoblot), or western blotting, is a widely used analytical technique in molecular biology and immunogenetics to detect specific proteins in a sample of tissue homogenate or extract. Besides detect ...
ting.
When luminol is sprayed evenly across an area, trace amounts of an activating oxidant make the luminol emit a blue glow that can be seen in a darkened room. The glow only lasts about 30 seconds, but can be documented photographically. The glow is stronger in areas receiving more spray; the intensity of the glow does not indicate the amount of blood or other activator present.
Synthesis
Luminol is synthesized in a two-step process, beginning with 3-nitro
phthalic acid
Phthalic acid is an aromatic dicarboxylic acid, with formula C6H4(CO2H)2. Although phthalic acid is of modest commercial importance, the closely related derivative phthalic anhydride is a commodity chemical produced on a large scale. Phthalic aci ...
.
First,
hydrazine
Hydrazine is an inorganic compound with the chemical formula . It is a simple pnictogen hydride, and is a colourless flammable liquid with an ammonia-like odour. Hydrazine is highly toxic unless handled in solution as, for example, hydrazine ...
(N
2H
4) is heated with the 3-nitrophthalic acid in a high-boiling solvent such as
triethylene glycol
Triethylene glycol, TEG, or triglycol is a colorless odorless viscous liquid with molecular formula HOCH2CH2OCH2CH2OCH2CH2OH. It is used as a plasticizer for vinyl polymers. It is also used in air sanitizer products, such as "Oust" or "Clean and ...
and
glycerol
Glycerol (), also called glycerine in British English and glycerin in American English, is a simple triol compound. It is a colorless, odorless, viscous liquid that is sweet-tasting and non-toxic. The glycerol backbone is found in lipids known ...
. An
acyl substitution condensation reaction
In organic chemistry, a condensation reaction is a type of chemical reaction in which two molecules are combined to form a single molecule, usually with the loss of a small molecule such as water. If water is lost, the reaction is also known as a ...
occurs, with loss of water, forming 3-nitrophthalhydrazide. Reduction of the
nitro group
In organic chemistry, nitro compounds are organic compounds that contain one or more nitro functional groups (). The nitro group is one of the most common explosophores (functional group that makes a compound explosive) used globally. The nitr ...
to an
amino group
In chemistry, amines (, ) are compounds and functional groups that contain a basic nitrogen atom with a lone pair. Amines are formally derivatives of ammonia (), wherein one or more hydrogen atoms have been replaced by a substituent such ...
with
sodium dithionite
Sodium dithionite (also known as sodium hydrosulfite) is a white crystalline powder with a sulfurous odor. Although it is stable in dry air, it decomposes in hot water and in acid solutions.
Structure
The structure has been examined by Raman s ...
(Na
2S
2O
4), via a transient
hydroxylamine
Hydroxylamine is an inorganic compound with the formula . The material is a white crystalline, hygroscopic compound.Greenwood and Earnshaw. ''Chemistry of the Elements.'' 2nd Edition. Reed Educational and Professional Publishing Ltd. pp. 431–43 ...
intermediate, produces luminol.
The compound was first synthesized in Germany in 1902, but was not named "luminol" until 1934.
[
]
Chemiluminescence
To exhibit its luminescence, the luminol must be activated with an oxidant. Usually, a solution containing hydrogen peroxide
Hydrogen peroxide is a chemical compound with the formula . In its pure form, it is a very pale blue liquid that is slightly more viscous than water. It is used as an oxidizer, bleaching agent, and antiseptic, usually as a dilute solution (3%†...
(H2O2) and hydroxide
Hydroxide is a diatomic anion with chemical formula OH−. It consists of an oxygen and hydrogen atom held together by a single covalent bond, and carries a negative electric charge. It is an important but usually minor constituent of water. I ...
ions in water is the activator. In the presence of a catalyst
Catalysis () is the process of increasing the rate of a chemical reaction by adding a substance known as a catalyst (). Catalysts are not consumed in the reaction and remain unchanged after it. If the reaction is rapid and the catalyst recyc ...
such as an iron
Iron () is a chemical element with symbol Fe (from la, ferrum) and atomic number 26. It is a metal that belongs to the first transition series and group 8 of the periodic table. It is, by mass, the most common element on Earth, right in f ...
or periodate
Periodate is an anion composed of iodine and oxygen. It is one of a number of oxyanions of iodine and is the highest in the series, with iodine existing in oxidation state +7. Unlike other perhalogenates, such as perchlorate, it can exist in two ...
compound, the hydrogen peroxide decomposes to form oxygen and water:
:2 H2O2 → + 2 H2O
:H2O2 + KIO4 → KIO3 + O2 + H2O
Laboratory settings often use potassium ferricyanide
Potassium ferricyanide is the chemical compound with the formula K3 e(CN)6 This bright red salt contains the octahedrally coordinated 3−.html" ;"title="e(CN)6sup>3−">e(CN)6sup>3− ion. It is soluble in water and its solution shows some g ...
or potassium periodate
Potassium periodate is an inorganic salt with the molecular formula KIO4. It is composed of a potassium cation and a periodate anion and may also be regarded as the potassium Salt (chemistry), salt of periodic acid. Note that the pronunciation is ...
for the catalyst. In the forensic detection of blood, the catalyst is the iron present in haemoglobin
Hemoglobin (haemoglobin BrE) (from the Greek word αἷμα, ''haîma'' 'blood' + Latin ''globus'' 'ball, sphere' + ''-in'') (), abbreviated Hb or Hgb, is the iron-containing oxygen-transport metalloprotein present in red blood cells (erythrocyte ...
. Enzyme
Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products. A ...
s in a variety of biological systems may also catalyse the decomposition of hydrogen peroxide.
The exact mechanism of luminol chemiluminescence is a complex multi-step reaction, especially in aqueous conditions. A recent theoretical investigation has been able to elucidate the reaction cascade as shown below. Luminol is first deprotonated in basic conditions, then oxidized to the anionic radical. Which in turn has two paths available to give the key intermediate α-hydroxy- peroxide. After cyclization to the endoperoxide, the mono-anion will undergo decomposition without luminescence, if the pH is too low (< 8.2) for a second deprotonation. The endoperoxide dianion, however can give the retro-Diels-Alder product: 1,2-dioxane-3,6-dione dianion. And after chemiexcitation by two single-electron-transfers (SET) gives 3-aminophthalate dianion in its first singlet excited-state (S1). This highly instable molecule relaxes to the ground state, thereby emitting light of around 425 nm wavelength (purple-blue), the so-called chemiluminescence.
Use in criminal investigation
History
In 1928, German chemist H. O. Albrecht found that blood
Blood is a body fluid in the circulatory system of humans and other vertebrates that delivers necessary substances such as nutrients and oxygen to the cells, and transports metabolic waste products away from those same cells. Blood in the c ...
, among other substances, enhanced the luminescence of luminol in an alkaline solution of hydrogen peroxide. In 1936, Karl Gleu and Karl Pfannstiel confirmed this enhancement in the presence of haematin
Haematin (also known as hematin, ferriheme, hematosin, hydroxyhemin, oxyheme, phenodin, or oxyhemochromogen) is a dark bluish or brownish pigment containing iron in the ferric state, obtained by the oxidation of haem.
Haematin inhibits the synth ...
, a component of blood. In 1937, German forensic scientist Walter Specht made extensive studies of luminol's application to the detection of blood at crime scenes. In 1939, San Francisco pathologists Frederick Proescher and A. M. Moody made three important observations about luminol:
# although the test is presumptive, large areas of suspected material can be examined rapidly;
# dried and decomposed blood gave a stronger and more lasting reaction than fresh blood; and
# if the luminescence disappears, it may be reproduced by the application of a fresh luminol-hydrogen peroxide solution; dried bloodstains may thus be made luminescent repeatedly.
Theory
Crime scene investigator
Forensic science, also known as criminalistics, is the application of science to criminal and civil laws, mainly—on the criminal side—during criminal investigation, as governed by the legal standards of admissible evidence and criminal p ...
s use luminol to find traces of blood, even if someone has cleaned or removed it. The investigator sprays a solution of luminol and the oxidant. The iron in blood catalyses the luminescence. The amount of catalyst necessary to cause the reaction is very small relative to the amount of luminol, allowing detection of even trace amounts of blood. The blue glow lasts for about 30 seconds per application. Detecting the glow requires a fairly dark room. Any glow detected may be documented by a long-exposure photograph
A photograph (also known as a photo, image, or picture) is an image created by light falling on a photosensitive surface, usually photographic film or an electronic image sensor, such as a CCD or a CMOS chip. Most photographs are now create ...
.
Drawbacks
Luminol's use in a crime scene investigation is somewhat hampered by the fact that it reacts to iron- and copper
Copper is a chemical element with the symbol Cu (from la, cuprum) and atomic number 29. It is a soft, malleable, and ductile metal with very high thermal and electrical conductivity. A freshly exposed surface of pure copper has a pinkis ...
-containing compounds, bleaches, horseradish
Horseradish (''Armoracia rusticana'', syn. ''Cochlearia armoracia'') is a perennial plant of the family Brassicaceae (which also includes mustard, wasabi, broccoli, cabbage, and radish). It is a root vegetable, cultivated and used worldwide ...
, fecal matter
Feces ( or faeces), known colloquially and in slang as poo and poop, are the solid or semi-solid remains of food that was not digested in the small intestine, and has been broken down by bacteria in the large intestine. Feces contain a relat ...
, or cigarette smoke residue.
Application of luminol to a piece of evidence may prevent other tests from being performed on it; however DNA has been successfully extracted from samples exposed to luminol.Technical note about ''Hemaglow''
Related molecules
* Luminol: 5-amino-2,3-dihydro-1,4-phthalazinedione ; 3-amino-phthalhydrazide ; 1,4-phthalazinedione, 5-amino-2,3-dihydro ; CAS:
21-31-3br /> C
8H
7N
3O
2 – MW: 177.16
λ
abs (in 0.1 N NaOH) λ
max 1 : 347 nm & λ
max 2 : 300 nm; EC (at λ
max 1): 7650 L/mol × cm
λ
abs / λ
em (MeOH): 355/413 nm
* Luminol, sodium salt: sodium 3-amino-phthalhydrazide; CAS:
0666-12-0br /> C
8H
6N
3O
2Na – MW: 199.12
* Luminol hemihydrate: a
hydrate
In chemistry, a hydrate is a substance that contains water or its constituent elements. The chemical state of the water varies widely between different classes of hydrates, some of which were so labeled before their chemical structure was understo ...
of sodium 3-amino-phthalhydrazide; CAS:
06658-90-4br /> C
8H
6N
3O
2Na – MW: 217.16
* Luminol hydrochloride: 3-amino-phthalhydrazide hydrochloride; CAS:
4165-64-3br /> C
8H
7N
3O
2 · HCl MW: 213.62
* Isoluminol: 4-aminophthalhydrazide; CAS:
682-14-1br />C
8H
7N
3O
2 – MW: 117.16 (Xi)
* Isoluminol monohydrate: 4-aminophthalhydrazide monohydrate
C
8H
7N
3O
2 – MW: 195.15
* Isoluminol ABEI: 4-aminophthalhydrazide monohydrate; CAS:
6612-29-1
See also
*
Fluorescein
Fluorescein is an organic compound and dye based on the xanthene tricyclic structural motif, formally belonging to triarylmethine dyes family. It is available as a dark orange/red powder slightly soluble in water and alcohol. It is widely used ...
*
Diphenyl oxalate
Diphenyl oxalate (trademark name Cyalume) is a solid whose oxidation products are responsible for the chemiluminescence in a glowstick. This chemical is the double ester of phenol with oxalic acid. Upon reaction with hydrogen peroxide, 1,2-diox ...
References
External links
HowStuffWorks "How Luminol Works"Luminol demonstration in Turin
{{Authority control
Aromatic amines
Chemical tests
Forensic chemicals
Hydrazides
Phthalazines
Lactams
Chemiluminescence