biochemistry Biochemistry or biological chemistry is the study of chemical processes within and relating to living organisms. A sub-discipline of both chemistry and biology, biochemistry may be divided into three fields: structural biology, enzymology ...

, ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)) is a chemical compound used to observe the

reaction kinetics Chemical kinetics, also known as reaction kinetics, is the branch of physical chemistry that is concerned with understanding the rates of chemical reactions. It is to be contrasted with chemical thermodynamics, which deals with the direction in wh ...

of specific

enzyme Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different molecule ...

s. A common use for it is in the

enzyme-linked immunosorbent assay The enzyme-linked immunosorbent assay (ELISA) (, ) is a commonly used analytical biochemistry assay, first described by Eva Engvall and Peter Perlmann in 1971. The assay uses a solid-phase type of enzyme immunoassay (EIA) to detect the presence ...

(

ELISA The enzyme-linked immunosorbent assay (ELISA) (, ) is a commonly used analytical biochemistry assay, first described by Eva Engvall and Peter Perlmann in 1971. The assay uses a solid-phase type of enzyme immunoassay (EIA) to detect the presence ...

) to detect the binding of molecules to each other. It is commonly used as a

substrate Substrate may refer to: Physical layers *Substrate (biology), the natural environment in which an organism lives, or the surface or medium on which an organism grows or is attached ** Substrate (locomotion), the surface over which an organism lo ...

with

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% ...

for a

peroxidase Peroxidases or peroxide reductases ( EC numberbr>1.11.1.x are a large group of enzymes which play a role in various biological processes. They are named after the fact that they commonly break up peroxides. Functionality Peroxidases typically ca ...

enzyme (such as

horseradish peroxidase The enzyme horseradish peroxidase (HRP), found in the roots of horseradish, is used extensively in biochemistry applications. It is a metalloenzyme with many isoforms, of which the most studied type is C. It catalyzes the oxidation of various or ...

) or alone with blue multicopper oxidase enzymes (such as

laccase Laccases () are multicopper oxidases found in plants, fungi, and bacteria. Laccases oxidize a variety of phenolic substrates, performing one-electron oxidations, leading to crosslinking. For example, laccases play a role in the formation of l ...

bilirubin oxidase In enzymology, a bilirubin oxidase, BOD or BOx, () is an enzyme encoded by a gene in various organisms that catalyzes the chemical reaction :2 bilirubin + O2 \rightleftharpoons 2 biliverdin + 2 H2O This enzyme belongs to the family of oxidoreduct ...

). Its use allows the reaction kinetics of

peroxidases Peroxidases or peroxide reductases ( EC numberbr>1.11.1.x are a large group of enzymes which play a role in various biological processes. They are named after the fact that they commonly break up peroxides. Functionality Peroxidases typically ca ...

themselves to be followed. In this way it also can be used to indirectly follow the reaction kinetics of any

-producing enzyme, or to simply quantify the amount of hydrogen peroxide in a sample. The formal reduction potentials for ABTS are high enough for it to act as an electron donor for the reduction of oxo species such as molecular oxygen and hydrogen peroxide, particularly at the less-extreme pH values encountered in biological catalysis. Under these conditions, the sulfonate groups are fully deprotonated and the mediator exists as a dianion. :ABTS^–· + e^– → ABTS^2– E° = 0.67 V vs

SHE She most commonly refers to: *She (pronoun), the third person singular, feminine, nominative case pronoun in modern English. She or S.H.E. may also refer to: Literature and films *'' She: A History of Adventure'', an 1887 novel by H. Rider Hagga ...

:ABTS + e^– → ABTS^–· E° = 1.08 V vs

This compound is chosen because the enzyme facilitates the reaction with hydrogen peroxide, turning it into a green and

soluble In chemistry, solubility is the ability of a substance, the solute, to form a solution with another substance, the solvent. Insolubility is the opposite property, the inability of the solute to form such a solution. The extent of the solub ...

end-product. Its new

absorbance Absorbance is defined as "the logarithm of the ratio of incident to transmitted radiant power through a sample (excluding the effects on cell walls)". Alternatively, for samples which scatter light, absorbance may be defined as "the negative lo ...

maximum of 420 nm

light Light or visible light is electromagnetic radiation that can be perceived by the human eye. Visible light is usually defined as having wavelengths in the range of 400–700 nanometres (nm), corresponding to frequencies of 750–420 te ...

(ε = 3.6 × 10⁴ M^–1 cm^–1) can easily be followed with a spectrophotometer, a common laboratory instrument. It is sometimes used as part of a

glucose Glucose is a simple sugar with the molecular formula . Glucose is overall the most abundant monosaccharide, a subcategory of carbohydrates. Glucose is mainly made by plants and most algae during photosynthesis from water and carbon dioxide, usi ...

estimating reagent when finding glucose concentrations of solutions such as

blood serum Serum () is the fluid and solute component of blood which does not play a role in clotting. It may be defined as blood plasma without the clotting factors, or as blood with all cells and clotting factors removed. Serum includes all proteins not ...

. ABTS is also frequently used by the food industry and agricultural researchers to measure the

antioxidant Antioxidants are compounds that inhibit oxidation, a chemical reaction that can produce free radicals. This can lead to polymerization and other chain reactions. They are frequently added to industrial products, such as fuels and lubricants ...

capacities of foods. In this assay, ABTS is converted to its radical cation by addition of

sodium persulfate Sodium persulfate is the inorganic compound with the formula Na2 S2 O8. It is the sodium salt of peroxydisulfuric acid, H2S2O8, an oxidizing agent. It is a white solid that dissolves in water. It is almost non-hygroscopic and has good shelf-lif ...

. This radical cation is blue in color and absorbs light at 415, 645, 734 and 815 nm. The ABTS radical cation is reactive towards most antioxidants including phenolics,

thiols In organic chemistry, a thiol (; ), or thiol derivative, is any organosulfur compound of the form , where R represents an alkyl or other organic substituent. The functional group itself is referred to as either a thiol group or a sulfhydryl grou ...

and

Vitamin C Vitamin C (also known as ascorbic acid and ascorbate) is a water-soluble vitamin found in citrus and other fruits and vegetables, also sold as a dietary supplement and as a topical 'serum' ingredient to treat melasma (dark pigment spots) a ...

. During this reaction, the blue ABTS radical cation is converted back to its colorless neutral form. The reaction may be monitored spectrophotometrically. This assay is often referred to as the

Trolox equivalent antioxidant capacity The Trolox equivalent antioxidant capacity (TEAC) assay measures the antioxidant Antioxidants are compounds that inhibit oxidation, a chemical reaction that can produce free radicals. This can lead to polymerization and other chain reactions ...

(TEAC) assay. The reactivity of the various antioxidants tested are compared to that of

Trolox Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) is a water-soluble analog of vitamin E sold by Hoffman-LaRoche. It is an antioxidant like vitamin E and it is used in biological or biochemical applications to reduce oxidative stres ...

, which is a water-soluble analog of

vitamin E Vitamin E is a group of eight fat soluble compounds that include four tocopherols and four tocotrienols. Vitamin E deficiency, which is rare and usually due to an underlying problem with digesting dietary fat rather than from a diet low in vitami ...

Applications for functional food analysis

Based on the special chemical properties of formed free radicals, ABTS assay has been used to determine the antioxidant capacity of food products. For example, polyphenol compounds, which widely exist in fruit, can quench free radicals inside human body, thus prevent oxidative damage by free radicals. The antioxidant potency of plant extract or food product has been measured by ABTS assay. One example with detailed method is the antioxidant activity analysis of Hibiscus products.

References

{{DEFAULTSORT:Abts Biochemistry methods Enzyme kinetics Sulfonic acids Benzothiazolines