Diethyl pyrocarbonate (DEPC), also called diethyl dicarbonate (

IUPAC The International Union of Pure and Applied Chemistry (IUPAC ) is an international federation of National Adhering Organizations working for the advancement of the chemical sciences, especially by developing nomenclature and terminology. It is ...

name), is used in the

laboratory A laboratory (; ; colloquially lab) is a facility that provides controlled conditions in which scientific or technological research, experiments, and measurement may be performed. Laboratories are found in a variety of settings such as schools ...

to inactivate

RNase Ribonuclease (commonly abbreviated RNase) is a type of nuclease that catalyzes the degradation of RNA into smaller components. Ribonucleases can be divided into endoribonucleases and exoribonucleases, and comprise several sub-classes within the ...

enzyme An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different mol ...

s in water and on laboratory utensils. It does so by the

covalent A covalent bond is a chemical bond that involves the sharing of electrons to form electron pairs between atoms. These electron pairs are known as shared pairs or bonding pairs. The stable balance of attractive and repulsive forces between atom ...

modification of

histidine Histidine (symbol His or H) is an essential amino acid that is used in the biosynthesis of proteins. It contains an Amine, α-amino group (which is in the protonated –NH3+ form under Physiological condition, biological conditions), a carboxylic ...

(most strongly),

lysine Lysine (symbol Lys or K) is an α-amino acid that is a precursor to many proteins. Lysine contains an α-amino group (which is in the protonated form when the lysine is dissolved in water at physiological pH), an α-carboxylic acid group ( ...

cysteine Cysteine (; symbol Cys or C) is a semiessential proteinogenic amino acid with the chemical formula, formula . The thiol side chain in cysteine enables the formation of Disulfide, disulfide bonds, and often participates in enzymatic reactions as ...

, and

tyrosine -Tyrosine or tyrosine (symbol Tyr or Y) or 4-hydroxyphenylalanine is one of the 20 standard amino acids that are used by cells to synthesize proteins. It is a conditionally essential amino acid with a polar side group. The word "tyrosine" is ...

residues. DEPC-treated (and therefore RNase-free) water is used in handling of

RNA Ribonucleic acid (RNA) is a polymeric molecule that is essential for most biological functions, either by performing the function itself (non-coding RNA) or by forming a template for the production of proteins (messenger RNA). RNA and deoxyrib ...

in the laboratory to reduce the risk of RNA being degraded by RNases. Water is usually treated with 0.1% v/v DEPC for at least 2 hours at 37 °C and then

autoclave An autoclave is a machine used to carry out industrial and scientific processes requiring elevated temperature and pressure in relation to ambient pressure and/or temperature. Autoclaves are used before surgical procedures to perform steriliza ...

d (at least 15 min) to inactivate traces of DEPC. Inactivation of DEPC in this manner yields CO₂ and

ethanol Ethanol (also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is an organic compound with the chemical formula . It is an Alcohol (chemistry), alcohol, with its formula also written as , or EtOH, where Et is the ps ...

. Higher concentrations of DEPC are capable of deactivating larger amounts of RNase, but remaining traces or byproducts may inhibit further biochemical reactions such as in vitro transcription. Furthermore, chemical modification of RNA such as carboxymethylation is possible when traces of DEPC or its byproducts are present, resulting in impaired recovery of intact RNA even after buffer exchange (after precipitation). Depc treated water

DEPC is unstable in water and susceptible to hydrolysis to carbon dioxide and ethanol, especially in the presence of a

nucleophile In chemistry, a nucleophile is a chemical species that forms bonds by donating an electron pair. All molecules and ions with a free pair of electrons or at least one pi bond can act as nucleophiles. Because nucleophiles donate electrons, they are ...

. For this reason, DEPC cannot be used with

Tris Tris, or tris(hydroxymethyl)aminomethane, or known during medical use as tromethamine or THAM, is an organic compound with the formula (HOCH2)3CNH2. It is extensively used in biochemistry and molecular biology as a component of buffer solutions ...

HEPES HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) is a zwitterionic sulfonic acid buffering agent. It is one of the twenty Good's buffers. HEPES is widely used in cell culture, largely because it is better at maintaining physiological pH ...

buffers. In contrast, it can be used with

phosphate-buffered saline Phosphate-buffered saline (PBS) is a buffer solution (pH ~ 7.4) commonly used in biological research. It is a water-based salt solution containing disodium hydrogen phosphate, sodium chloride and, in some formulations, potassium chloride and potass ...

MOPS MOPS (3-(''N''-morpholino)propanesulfonic acid) is a buffer solution, buffer introduced in the 1960s, one of the twenty Good's buffers. It is a structural analog to MES (buffer), MES, and like MES, its structure contains a morpholine ring. HEPES ...

. A handy rule is that enzymes or chemicals which have active -O:, -N: or -S: cannot be treated with DEPC to become RNase-free, as DEPC reacts with these species. Furthermore, DEPC degradation products can inhibit in vitro transcription. DEPC

derivatization Derivatization is a technique used in chemistry which converts a chemical compound into a product (the reaction's derivate) of similar chemical structure, called a derivative. Generally, a specific functional group of the compound participates ...

of histidines is also used to study the importance of histidyl residues in enzymes. Modification of

by DEPC results in carbethoxylated derivates at the N-omega-2 nitrogen of the

imidazole Imidazole (ImH) is an organic compound with the formula . It is a white or colourless solid that is soluble in water, producing a mildly alkaline solution. It can be classified as a heterocycle, specifically as a diazole. Many natural products, ...

ring. DEPC modification of histidines can be reversed by treatment with 0.5 M

hydroxylamine Hydroxylamine (also known as hydroxyammonia) is an inorganic compound with the chemical formula . The compound exists as hygroscopic colorless crystals.Greenwood and Earnshaw. ''Chemistry of the Elements.'' 2nd Edition. Reed Educational and Prof ...

at neutral pH. DEPC can also be used for probing the structure of double-stranded DNA.

References

{{Reflist

External links

DEPC at OpenWetWare life scientist wiki
Dicarbonates Ethyl esters

See also

References

External links