Hypochlorous acid
Encyclopedia
Hypochlorous acid is a weak acid
with the chemical formula
HClO. It forms when chlorine
dissolves in water. It cannot be isolated in pure form due to rapid equilibration with its precursor. HClO is an oxidizer, and as its sodium salt sodium hypochlorite, (NaClO), or its calcium salt calcium hypochlorite
, (Ca(CIO)2) is used as a bleach, a deodorant
, and a disinfectant.
, HCIO converts alkene
s to chlorohydrins.
In biology
, hypochlorous acid is generated in activated neutrophils by myeloperoxidase-mediated peroxidation of chloride ions, and contributes to the destruction of bacteria
.
In water treatment, hypochlorous acid is the active sanitizer in hypochlorite-based products (e.g. used in swimming pools).
In food service and water distribution, specialized equipment to generate weak solutions of HOCl from water and salt is sometimes used to generate adequate quantities of safe (unstable) disinfectant to treat food preparation surfaces and water supplies.
to water
gives both hydrochloric acid (HCl) and hypochlorous acid:
When acids are added to aqueous salts of hypochlorous acid (such as sodium hypochlorite in commercial bleach solution), the resultant reaction is driven to the left, and chlorine gas is evolved. Thus, the formation of stable hypochlorite bleaches is facilitated by dissolving chlorine gas into basic water solutions, such as sodium hydroxide.
The acid can also be prepared by dissolving dichlorine monoxide
in water; under standard aqueous conditions, anhydrous hypochlorous acid is impossible to prepare due to the readily reversible equilibrium between it and its anhydride:
The presence of light or transition metal oxides of copper, nickel, or cobalt accelerates the exothermic decomposition into hydrochloric acid
and oxygen
:
solution, hypochlorous acid partially dissociates into the anion hypochlorite OCl−:
Salts of hypochlorous acid are called hypochlorite
s. One of the best-known hypochlorites is NaClO, the active ingredient in bleach.
HClO is a stronger oxidant than chlorine under standard conditions.
HClO reacts with HCl to form chlorine gas:
inhibitor that, in sufficient quantity, could completely inactivate proteins containing sulfhydryl groups
. This is because HClO oxidises sulfhydryl groups, leading to the formation of disulfide bond
s that can result in crosslinking of protein
s. The HClO mechanism of sulfhydryl oxidation is similar to that of chloramine
, and may only be bacteriostatic, because, once the residual chlorine is dissipated, some sulfhydryl function can be restored. One sulfhydryl-containing amino acid can scavenge up to four molecules of HOCl. Consistent with this, it has been proposed that sulfhydryl groups of sulfur-containing amino acid
s can be oxidized a total of three times by three HClO molecules, with the fourth reacting with the α-amino group. The first reaction yields sulfenic acid
(R-SOH) then sulfinic acid (R-SO2H) and finally R-SO3H. Each of those intermediates can also condense with another sulfhydryl group, causing cross-linking and aggregation of proteins. Sulfinic acid and R-SO3H derivatives are produced only at high molar excesses of HClO, and disulfides are formed primarily at bacteriocidal levels. Disulfide bonds can also be oxidized by HClO to sulfinic acid. Because the oxidation of sulfhydryls and disulfides
evolves hydrochloric acid
, this process results in the depletion HClO.
side-chains, with the chlorine from HClO displacing a hydrogen, resulting in an organic chloramine. Chlorinated amino acid
s rapidly decompose, but protein
chloramines are longer-lived and retain some oxidative capacity. Thomas et al. concluded from their results that most organic chloramines decayed by internal rearrangement and that fewer available NH2
groups promoted attack on the peptide bond
, resulting in cleavage of the protein
. McKenna and Davies found that 10 mM or greater HClO is necessary to fragment proteins in vivo. Consistent with these results, it was later proposed that the chloramine undergoes a molecular rearrangement, releasing HCl
and ammonia
to form an amide
. The amide group can further react with another amino group
to form a Schiff base
, causing cross-linking and aggregation of proteins.
is the most reactive because HClO reacts with both the heterocyclic NH group and the amino group. In similar manner, TMP
with only a heterocyclic NH group that is reactive with HClO is the second-most reactive. AMP
and CMP
, which have only a slowly reactive amino group, are less reactive with HClO. UMP
has been reported to be reactive only at a very slow rate. The heterocyclic NH groups are more reactive than amino groups, and their secondary chloramines are able to donate the chlorine. These reactions likely interfere with DNA base pairing, and, consistent with this, Prütz has reported a decrease in viscosity of DNA exposed to HClO similar to that seen with heat denaturation. The sugar moieties are nonreactive and the DNA backbone is not broken. NADH can react with chlorinated TMP and UMP as well as HClO. This reaction can regenerate UMP and TMP and results in the 5-hydroxy derivative of NADH. The reaction with TMP or UMP is slowly reversible to regenerate HClO. A second slower reaction that results in cleavage of the pyridine ring occurs when excess HClO is present. NAD+ is inert to HClO.
s in lipid
s, but not saturated bond
s, and the OCl−
ion does not participate in this reaction. This reaction occurs by hydrolysis
with addition of chlorine
to one of the carbons and a hydroxyl
to the other. The resulting compound is a chlorhydrin. The polar chlorine disrupts lipid bilayer
s and could increase permeability. When chlorhydrin formation occurs in lipid bilayers of red blood cells, increased permeability occurs. Disruption could occur if enough chlorhydrin is formed. The addition of preformed chlorhydrins to red blood cells can affect permeability as well. Cholesterol
chlorhydrins have also been observed, but do not greatly affect permeability, and it is believed that Cl2
is responsible for this reaction.
exposed to hypochlorous acid lose viability
in less than 100 ms due to inactivation of many vital systems. Hypochlorous acid has a reported of 0.0104–0.156 ppm and 2.6 ppm caused 100% growth inhibition in 5 minutes. However it should be noted that the concentration required for bactericidal activity is also highly dependent on bacterial concentration.
oxidation is a major factor in the bacteriocidal nature of chlorine solutions. He proposed that the active agent or agents diffuse across the cytoplasmic membrane to inactivate key sulfhydryl
-containing enzyme
s in the glycolytic pathway
. This group was also the first to note that chlorine solutions (HOCl) inhibit sulfhydryl
enzyme
s. Later studies have shown that, at bacteriocidal levels, the cytosol
components do not react with HOCl. In agreement with this, McFeters and Camper found that aldolase
, an enzyme
that Knox et al. proposes would be inactivated, was unaffected by HOCl in vivo
. It has been further shown that loss of sulfhydryl
s does not correlate with inactivation. That leaves the question concerning what causes inhibition of glucose
oxidation. The discovery that HOCl blocks induction of β-galactosidase by added lactose
led to a possible answer to this question. The uptake of radiolabeled substrates by both ATP hydrolysis and proton co-transport
may be blocked by exposure to HOCl preceding loss of viability. From this observation, it proposed that HOCl blocks uptake of nutrients by inactivating transport proteins. The question of loss of glucose oxidation has been further explored in terms of loss of respiration. Venkobachar et al. found that succinic dehydrogenase was inhibited in vitro by HOCl, which led to the investigation of the possibility that disruption of electron transport could be the cause of bacterial inactivation. Albrich et al. subsequently found that HOCl destroys cytochrome
s and iron-sulfur cluster
s and observed that oxygen uptake is abolished by HOCl and adenine nucleotides are lost. It was also observed that irreversible oxidation of cytochrome
s paralleled the loss of respiratory activity. One way of addressing the loss of oxygen uptake was by studying the effects of HOCl on succinate-dependent electron transport. Rosen et al. found that levels of reductable cytochrome
s in HOCl-treated cells were normal, and these cells were unable to reduce them. Succinate dehydrogenase was also inhibited by HOCl, stopping the flow of electrons to oxygen. Later studies revealed that Ubiquinol oxidase activity ceases first, and the still-active cytochrome
s reduce the remaining quinone. The cytochrome
s then pass the electron
s to oxygen
, which explains why the cytochrome
s cannot be reoxidized, as observed by Rosen et al. However, this line of inquiry was ended when Albrich et al. found that cellular inactivation precedes loss of respiration by using a flow mixing system that allowed evaluation of viability on much smaller time scales. This group found that cells capable of respiring could not divide after exposure to HOCl.
replication. When bacteria are exposed to HOCl, there is a precipitous decline in DNA synthesis
that precedes inhibition of protein
synthesis, and closely parallels loss of viability. During bacterial genome replication, the origin of replication
(oriC in E. coli) binds to proteins that are associated with the cell membrane, and it was observed that HOCl treatment decreases the affinity of extracted membranes for oriC, and this decreased affinity also parallels loss of viability. A study by Rosen et al. compared the rate of HOCl inhibition of DNA replication of plasmids with different replication origins and found that certain plasmids exhibited a delay in the inhibition of replication when compared to plasmids containing oriC. Rosen’s group proposed that inactivation of membrane proteins involved in DNA replication are the mechanism of action of HOCl.
and methionine
oxidation. A recent examination of HOCl's bactericidal role revealed it to be a potent inducer of protein aggregation. Hsp33, a chaperone known to be activated by oxidative heat stress, protects bacteria from the effects of HOCl by acting as a holdase, effectively preventing protein aggregation. Strains of E. coli and Vibrio cholerae lacking Hsp33 were rendered especially sensitive to HOCl. Hsp33 protected many essential proteins from aggregation and inactivation due to HOCl, which is a probable mediator of HOCl's bactericidal effects.
and sodium hypochlorite.
of chlorine gas in alkaline solutions.
Acid
An acid is a substance which reacts with a base. Commonly, acids can be identified as tasting sour, reacting with metals such as calcium, and bases like sodium carbonate. Aqueous acids have a pH of less than 7, where an acid of lower pH is typically stronger, and turn blue litmus paper red...
with the chemical formula
Chemical formula
A chemical formula or molecular formula is a way of expressing information about the atoms that constitute a particular chemical compound....
HClO. It forms when chlorine
Chlorine
Chlorine is the chemical element with atomic number 17 and symbol Cl. It is the second lightest halogen, found in the periodic table in group 17. The element forms diatomic molecules under standard conditions, called dichlorine...
dissolves in water. It cannot be isolated in pure form due to rapid equilibration with its precursor. HClO is an oxidizer, and as its sodium salt sodium hypochlorite, (NaClO), or its calcium salt calcium hypochlorite
Calcium hypochlorite
Calcium hypochlorite is a chemical compound with formula 2. It is widely used for water treatment and as a bleaching agent...
, (Ca(CIO)2) is used as a bleach, a deodorant
Deodorant
Deodorants are substances applied to the body to affect body odor caused by bacterial growth and the smell associated with bacterial breakdown of perspiration in armpits, feet and other areas of the body. A subgroup of deodorants, antiperspirants, affect odor as well as prevent sweating by...
, and a disinfectant.
Uses
In organic synthesisOrganic synthesis
Organic synthesis is a special branch of chemical synthesis and is concerned with the construction of organic compounds via organic reactions. Organic molecules can often contain a higher level of complexity compared to purely inorganic compounds, so the synthesis of organic compounds has...
, HCIO converts alkene
Alkene
In organic chemistry, an alkene, olefin, or olefine is an unsaturated chemical compound containing at least one carbon-to-carbon double bond...
s to chlorohydrins.
In biology
Biology
Biology is a natural science concerned with the study of life and living organisms, including their structure, function, growth, origin, evolution, distribution, and taxonomy. Biology is a vast subject containing many subdivisions, topics, and disciplines...
, hypochlorous acid is generated in activated neutrophils by myeloperoxidase-mediated peroxidation of chloride ions, and contributes to the destruction of bacteria
Bacteria
Bacteria are a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria have a wide range of shapes, ranging from spheres to rods and spirals...
.
In water treatment, hypochlorous acid is the active sanitizer in hypochlorite-based products (e.g. used in swimming pools).
In food service and water distribution, specialized equipment to generate weak solutions of HOCl from water and salt is sometimes used to generate adequate quantities of safe (unstable) disinfectant to treat food preparation surfaces and water supplies.
Formation, stability and reactions
Addition of chlorineChlorine
Chlorine is the chemical element with atomic number 17 and symbol Cl. It is the second lightest halogen, found in the periodic table in group 17. The element forms diatomic molecules under standard conditions, called dichlorine...
to water
Water
Water is a chemical substance with the chemical formula H2O. A water molecule contains one oxygen and two hydrogen atoms connected by covalent bonds. Water is a liquid at ambient conditions, but it often co-exists on Earth with its solid state, ice, and gaseous state . Water also exists in a...
gives both hydrochloric acid (HCl) and hypochlorous acid:
- Cl2 + H2O HClO + HCl
When acids are added to aqueous salts of hypochlorous acid (such as sodium hypochlorite in commercial bleach solution), the resultant reaction is driven to the left, and chlorine gas is evolved. Thus, the formation of stable hypochlorite bleaches is facilitated by dissolving chlorine gas into basic water solutions, such as sodium hydroxide.
The acid can also be prepared by dissolving dichlorine monoxide
Dichlorine monoxide
Dichlorine monoxide, Cl2O, also known as, oxygen dichloride, dichlorine oxide or chlorine oxide is a chlorine oxide. It is a brownish yellow gas at room temperature which can explode in high concentrations when heated or sparked....
in water; under standard aqueous conditions, anhydrous hypochlorous acid is impossible to prepare due to the readily reversible equilibrium between it and its anhydride:
- 2 HOCl Cl2O + H2O K(0°C) = 3.55 dm3mol−1
The presence of light or transition metal oxides of copper, nickel, or cobalt accelerates the exothermic decomposition into hydrochloric acid
Hydrochloric acid
Hydrochloric acid is a solution of hydrogen chloride in water, that is a highly corrosive, strong mineral acid with many industrial uses. It is found naturally in gastric acid....
and oxygen
Oxygen
Oxygen is the element with atomic number 8 and represented by the symbol O. Its name derives from the Greek roots ὀξύς and -γενής , because at the time of naming, it was mistakenly thought that all acids required oxygen in their composition...
:
- 2 Cl2 + 2 H2O → 4 HCl + O2
Chemical reactions
In aqueousWater
Water is a chemical substance with the chemical formula H2O. A water molecule contains one oxygen and two hydrogen atoms connected by covalent bonds. Water is a liquid at ambient conditions, but it often co-exists on Earth with its solid state, ice, and gaseous state . Water also exists in a...
solution, hypochlorous acid partially dissociates into the anion hypochlorite OCl−:
- HClO OCl− + H+
Salts of hypochlorous acid are called hypochlorite
Hypochlorite
The hypochlorite ion, also known as chlorate anion is ClO−. A hypochlorite compound is a chemical compound containing this group, with chlorine in oxidation state +1.Hypochlorites are the salts of hypochlorous acid...
s. One of the best-known hypochlorites is NaClO, the active ingredient in bleach.
HClO is a stronger oxidant than chlorine under standard conditions.
- 2HClO(aq) + 2 + 2 Cl2(g) + 2 E = +1.63 V
HClO reacts with HCl to form chlorine gas:
- HClO + HCl → H2O + Cl2
Reactivity of HClO with biomolecules
Hypochlorous acid reacts with a wide variety of biomolecules, including DNA, RNA, fatty acid groups, cholesterol and proteins.Reaction with protein sulfhydryl groups
Knox et al. first noted that HClO is a sulfhydrylThiol
In organic chemistry, a thiol is an organosulfur compound that contains a carbon-bonded sulfhydryl group...
inhibitor that, in sufficient quantity, could completely inactivate proteins containing sulfhydryl groups
Thiol
In organic chemistry, a thiol is an organosulfur compound that contains a carbon-bonded sulfhydryl group...
. This is because HClO oxidises sulfhydryl groups, leading to the formation of disulfide bond
Disulfide bond
In chemistry, a disulfide bond is a covalent bond, usually derived by the coupling of two thiol groups. The linkage is also called an SS-bond or disulfide bridge. The overall connectivity is therefore R-S-S-R. The terminology is widely used in biochemistry...
s that can result in crosslinking of protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
s. The HClO mechanism of sulfhydryl oxidation is similar to that of chloramine
Chloramine
Chloramines are derivatives of ammonia by substitution of one, two or three hydrogen atoms with chlorine atoms. Monochloramine is an inorganic compound with the formula NH2Cl. It is an unstable colourless liquid at its melting point of -66° temperature, but it is usually handled as a dilute...
, and may only be bacteriostatic, because, once the residual chlorine is dissipated, some sulfhydryl function can be restored. One sulfhydryl-containing amino acid can scavenge up to four molecules of HOCl. Consistent with this, it has been proposed that sulfhydryl groups of sulfur-containing amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
s can be oxidized a total of three times by three HClO molecules, with the fourth reacting with the α-amino group. The first reaction yields sulfenic acid
Sulfenic acid
A sulfenic acid is an organosulfur compound and oxoacid with the general formula RSOH, where R ≠ H. Simple sulfenic acids, such as methanesulfenic acid, CH3SOH, are highly reactive and cannot be isolated in solution. In the gas phase the lifetime of methanesulfenic acid is about one minute...
(R-SOH) then sulfinic acid (R-SO2H) and finally R-SO3H. Each of those intermediates can also condense with another sulfhydryl group, causing cross-linking and aggregation of proteins. Sulfinic acid and R-SO3H derivatives are produced only at high molar excesses of HClO, and disulfides are formed primarily at bacteriocidal levels. Disulfide bonds can also be oxidized by HClO to sulfinic acid. Because the oxidation of sulfhydryls and disulfides
Disulfide bond
In chemistry, a disulfide bond is a covalent bond, usually derived by the coupling of two thiol groups. The linkage is also called an SS-bond or disulfide bridge. The overall connectivity is therefore R-S-S-R. The terminology is widely used in biochemistry...
evolves hydrochloric acid
Hydrochloric acid
Hydrochloric acid is a solution of hydrogen chloride in water, that is a highly corrosive, strong mineral acid with many industrial uses. It is found naturally in gastric acid....
, this process results in the depletion HClO.
Reaction with protein amino groups
Hypochlorous acid reacts readily with amino acids that have amino groupAmine
Amines are organic compounds and functional groups that contain a basic nitrogen atom with a lone pair. Amines are derivatives of ammonia, wherein one or more hydrogen atoms have been replaced by a substituent such as an alkyl or aryl group. Important amines include amino acids, biogenic amines,...
side-chains, with the chlorine from HClO displacing a hydrogen, resulting in an organic chloramine. Chlorinated amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
s rapidly decompose, but protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
chloramines are longer-lived and retain some oxidative capacity. Thomas et al. concluded from their results that most organic chloramines decayed by internal rearrangement and that fewer available NH2
Amine
Amines are organic compounds and functional groups that contain a basic nitrogen atom with a lone pair. Amines are derivatives of ammonia, wherein one or more hydrogen atoms have been replaced by a substituent such as an alkyl or aryl group. Important amines include amino acids, biogenic amines,...
groups promoted attack on the peptide bond
Peptide bond
This article is about the peptide link found within biological molecules, such as proteins. A similar article for synthetic molecules is being created...
, resulting in cleavage of the protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
. McKenna and Davies found that 10 mM or greater HClO is necessary to fragment proteins in vivo. Consistent with these results, it was later proposed that the chloramine undergoes a molecular rearrangement, releasing HCl
HCL
HCL or HCl can stand for:* Hairy cell leukemia, an uncommon and slowly progressing B cell leukemia* Hardware compatibility list...
and ammonia
Ammonia
Ammonia is a compound of nitrogen and hydrogen with the formula . It is a colourless gas with a characteristic pungent odour. Ammonia contributes significantly to the nutritional needs of terrestrial organisms by serving as a precursor to food and fertilizers. Ammonia, either directly or...
to form an amide
Amide
In chemistry, an amide is an organic compound that contains the functional group consisting of a carbonyl group linked to a nitrogen atom . The term refers both to a class of compounds and a functional group within those compounds. The term amide also refers to deprotonated form of ammonia or an...
. The amide group can further react with another amino group
Amine
Amines are organic compounds and functional groups that contain a basic nitrogen atom with a lone pair. Amines are derivatives of ammonia, wherein one or more hydrogen atoms have been replaced by a substituent such as an alkyl or aryl group. Important amines include amino acids, biogenic amines,...
to form a Schiff base
Schiff base
A Schiff base, named after Hugo Schiff, is a compound with a functional group that contains a carbon-nitrogen double bond with the nitrogen atom connected to an aryl or alkyl group, not hydrogen....
, causing cross-linking and aggregation of proteins.
Reaction with DNA and nucleotides
Hypochlourous acid reacts slowly with DNA and RNA as well as all nucleotides in vitro. GMPGuanosine monophosphate
Guanosine monophosphate, also known as 5'-guanidylic acid or guanylic acid and abbreviated GMP, is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside guanosine. GMP consists of the phosphate group, the pentose sugar ribose, and the nucleobase...
is the most reactive because HClO reacts with both the heterocyclic NH group and the amino group. In similar manner, TMP
Thymidine monophosphate
Thymidine monophosphate, also known as 5'-thymidylate, thymidylate, or TMP, is a nucleotide that is used as a monomer in DNA. It is an ester of phosphoric acid with the nucleoside thymidine. TMP consists of a phosphate group, the pentose sugar deoxyribose, and the nucleobase thymine...
with only a heterocyclic NH group that is reactive with HClO is the second-most reactive. AMP
Adenosine monophosphate
Adenosine monophosphate , also known as 5'-adenylic acid, is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid and the nucleoside adenosine. AMP consists of a phosphate group, the sugar ribose, and the nucleobase adenine...
and CMP
Cytidine monophosphate
Cytidine monophosphate, also known as 5'-cytidylic acid or simply cytidylate, and abbreviated CMP, is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside cytidine...
, which have only a slowly reactive amino group, are less reactive with HClO. UMP
Uridine monophosphate
Uridine monophosphate, also known as 5'-uridylic acid and abbreviated UMP, is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside uridine...
has been reported to be reactive only at a very slow rate. The heterocyclic NH groups are more reactive than amino groups, and their secondary chloramines are able to donate the chlorine. These reactions likely interfere with DNA base pairing, and, consistent with this, Prütz has reported a decrease in viscosity of DNA exposed to HClO similar to that seen with heat denaturation. The sugar moieties are nonreactive and the DNA backbone is not broken. NADH can react with chlorinated TMP and UMP as well as HClO. This reaction can regenerate UMP and TMP and results in the 5-hydroxy derivative of NADH. The reaction with TMP or UMP is slowly reversible to regenerate HClO. A second slower reaction that results in cleavage of the pyridine ring occurs when excess HClO is present. NAD+ is inert to HClO.
Reaction with lipids
Hypochlorous acid reacts with unsaturated bondSaturation (chemistry)
In chemistry, saturation has six different meanings, all based on reaching a maximum capacity...
s in lipid
Lipid
Lipids constitute a broad group of naturally occurring molecules that include fats, waxes, sterols, fat-soluble vitamins , monoglycerides, diglycerides, triglycerides, phospholipids, and others...
s, but not saturated bond
Saturation (chemistry)
In chemistry, saturation has six different meanings, all based on reaching a maximum capacity...
s, and the OCl−
Hypochlorite
The hypochlorite ion, also known as chlorate anion is ClO−. A hypochlorite compound is a chemical compound containing this group, with chlorine in oxidation state +1.Hypochlorites are the salts of hypochlorous acid...
ion does not participate in this reaction. This reaction occurs by hydrolysis
Hydrolysis
Hydrolysis is a chemical reaction during which molecules of water are split into hydrogen cations and hydroxide anions in the process of a chemical mechanism. It is the type of reaction that is used to break down certain polymers, especially those made by condensation polymerization...
with addition of chlorine
Chlorine
Chlorine is the chemical element with atomic number 17 and symbol Cl. It is the second lightest halogen, found in the periodic table in group 17. The element forms diatomic molecules under standard conditions, called dichlorine...
to one of the carbons and a hydroxyl
Hydroxyl
A hydroxyl is a chemical group containing an oxygen atom covalently bonded with a hydrogen atom. In inorganic chemistry, the hydroxyl group is known as the hydroxide ion, and scientists and reference works generally use these different terms though they refer to the same chemical structure in...
to the other. The resulting compound is a chlorhydrin. The polar chlorine disrupts lipid bilayer
Lipid bilayer
The lipid bilayer is a thin membrane made of two layers of lipid molecules. These membranes are flat sheets that form a continuous barrier around cells. The cell membrane of almost all living organisms and many viruses are made of a lipid bilayer, as are the membranes surrounding the cell nucleus...
s and could increase permeability. When chlorhydrin formation occurs in lipid bilayers of red blood cells, increased permeability occurs. Disruption could occur if enough chlorhydrin is formed. The addition of preformed chlorhydrins to red blood cells can affect permeability as well. Cholesterol
Cholesterol
Cholesterol is a complex isoprenoid. Specifically, it is a waxy steroid of fat that is produced in the liver or intestines. It is used to produce hormones and cell membranes and is transported in the blood plasma of all mammals. It is an essential structural component of mammalian cell membranes...
chlorhydrins have also been observed, but do not greatly affect permeability, and it is believed that Cl2
Chlorine
Chlorine is the chemical element with atomic number 17 and symbol Cl. It is the second lightest halogen, found in the periodic table in group 17. The element forms diatomic molecules under standard conditions, called dichlorine...
is responsible for this reaction.
Mode of disinfectant action
Escherichia coliEscherichia coli
Escherichia coli is a Gram-negative, rod-shaped bacterium that is commonly found in the lower intestine of warm-blooded organisms . Most E. coli strains are harmless, but some serotypes can cause serious food poisoning in humans, and are occasionally responsible for product recalls...
exposed to hypochlorous acid lose viability
Viability
Viable or viability is the ability of a thing to maintain itself or recover its potentialities.Viable or viability may also refer to:...
in less than 100 ms due to inactivation of many vital systems. Hypochlorous acid has a reported of 0.0104–0.156 ppm and 2.6 ppm caused 100% growth inhibition in 5 minutes. However it should be noted that the concentration required for bactericidal activity is also highly dependent on bacterial concentration.
Inhibition of glucose oxidation
In 1948, Knox et al. proposed the idea that inhibition of glucoseGlucose
Glucose is a simple sugar and an important carbohydrate in biology. Cells use it as the primary source of energy and a metabolic intermediate...
oxidation is a major factor in the bacteriocidal nature of chlorine solutions. He proposed that the active agent or agents diffuse across the cytoplasmic membrane to inactivate key sulfhydryl
Thiol
In organic chemistry, a thiol is an organosulfur compound that contains a carbon-bonded sulfhydryl group...
-containing enzyme
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
s in the glycolytic pathway
Glycolysis
Glycolysis is the metabolic pathway that converts glucose C6H12O6, into pyruvate, CH3COCOO− + H+...
. This group was also the first to note that chlorine solutions (HOCl) inhibit sulfhydryl
Thiol
In organic chemistry, a thiol is an organosulfur compound that contains a carbon-bonded sulfhydryl group...
enzyme
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
s. Later studies have shown that, at bacteriocidal levels, the cytosol
Cytosol
The cytosol or intracellular fluid is the liquid found inside cells, that is separated into compartments by membranes. For example, the mitochondrial matrix separates the mitochondrion into compartments....
components do not react with HOCl. In agreement with this, McFeters and Camper found that aldolase
Aldolase
Aldolase A is an enzyme that catalyses a reverse aldol reaction: The substrate, fructose 1,6-bisphosphate is broken down into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate . This reaction is a part of glycolysis. Three aldolase isozymes , encoded by three different genes, are...
, an enzyme
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
that Knox et al. proposes would be inactivated, was unaffected by HOCl in vivo
In vivo
In vivo is experimentation using a whole, living organism as opposed to a partial or dead organism, or an in vitro controlled environment. Animal testing and clinical trials are two forms of in vivo research...
. It has been further shown that loss of sulfhydryl
Thiol
In organic chemistry, a thiol is an organosulfur compound that contains a carbon-bonded sulfhydryl group...
s does not correlate with inactivation. That leaves the question concerning what causes inhibition of glucose
Glucose
Glucose is a simple sugar and an important carbohydrate in biology. Cells use it as the primary source of energy and a metabolic intermediate...
oxidation. The discovery that HOCl blocks induction of β-galactosidase by added lactose
Lactose
Lactose is a disaccharide sugar that is found most notably in milk and is formed from galactose and glucose. Lactose makes up around 2~8% of milk , although the amount varies among species and individuals. It is extracted from sweet or sour whey. The name comes from or , the Latin word for milk,...
led to a possible answer to this question. The uptake of radiolabeled substrates by both ATP hydrolysis and proton co-transport
Co-transport
Co-transport, also known as coupled transport or secondary active transport, refers to the simultaneous or sequential passive transfer of molecules or ions across biological membranes in a fixed ratio...
may be blocked by exposure to HOCl preceding loss of viability. From this observation, it proposed that HOCl blocks uptake of nutrients by inactivating transport proteins. The question of loss of glucose oxidation has been further explored in terms of loss of respiration. Venkobachar et al. found that succinic dehydrogenase was inhibited in vitro by HOCl, which led to the investigation of the possibility that disruption of electron transport could be the cause of bacterial inactivation. Albrich et al. subsequently found that HOCl destroys cytochrome
Cytochrome
Cytochromes are, in general, membrane-bound hemoproteins that contain heme groups and carry out electron transport.They are found either as monomeric proteins or as subunits of bigger enzymatic complexes that catalyze redox reactions....
s and iron-sulfur cluster
Iron-sulfur cluster
For biological Fe-S clusters, see iron-sulfur proteins.Iron-sulfur clusters are ensembles of iron and sulfide centres. Fe-S clusters are most often discussed in the context of the biological role for iron-sulfur proteins. Many Fe-S clusters are known in the area of organometallic chemistry and as...
s and observed that oxygen uptake is abolished by HOCl and adenine nucleotides are lost. It was also observed that irreversible oxidation of cytochrome
Cytochrome
Cytochromes are, in general, membrane-bound hemoproteins that contain heme groups and carry out electron transport.They are found either as monomeric proteins or as subunits of bigger enzymatic complexes that catalyze redox reactions....
s paralleled the loss of respiratory activity. One way of addressing the loss of oxygen uptake was by studying the effects of HOCl on succinate-dependent electron transport. Rosen et al. found that levels of reductable cytochrome
Cytochrome
Cytochromes are, in general, membrane-bound hemoproteins that contain heme groups and carry out electron transport.They are found either as monomeric proteins or as subunits of bigger enzymatic complexes that catalyze redox reactions....
s in HOCl-treated cells were normal, and these cells were unable to reduce them. Succinate dehydrogenase was also inhibited by HOCl, stopping the flow of electrons to oxygen. Later studies revealed that Ubiquinol oxidase activity ceases first, and the still-active cytochrome
Cytochrome
Cytochromes are, in general, membrane-bound hemoproteins that contain heme groups and carry out electron transport.They are found either as monomeric proteins or as subunits of bigger enzymatic complexes that catalyze redox reactions....
s reduce the remaining quinone. The cytochrome
Cytochrome
Cytochromes are, in general, membrane-bound hemoproteins that contain heme groups and carry out electron transport.They are found either as monomeric proteins or as subunits of bigger enzymatic complexes that catalyze redox reactions....
s then pass the electron
Electron
The electron is a subatomic particle with a negative elementary electric charge. It has no known components or substructure; in other words, it is generally thought to be an elementary particle. An electron has a mass that is approximately 1/1836 that of the proton...
s to oxygen
Oxygen
Oxygen is the element with atomic number 8 and represented by the symbol O. Its name derives from the Greek roots ὀξύς and -γενής , because at the time of naming, it was mistakenly thought that all acids required oxygen in their composition...
, which explains why the cytochrome
Cytochrome
Cytochromes are, in general, membrane-bound hemoproteins that contain heme groups and carry out electron transport.They are found either as monomeric proteins or as subunits of bigger enzymatic complexes that catalyze redox reactions....
s cannot be reoxidized, as observed by Rosen et al. However, this line of inquiry was ended when Albrich et al. found that cellular inactivation precedes loss of respiration by using a flow mixing system that allowed evaluation of viability on much smaller time scales. This group found that cells capable of respiring could not divide after exposure to HOCl.
Depletion of adenine nucleotides
Having eliminated loss of respiration Albrich et al. proposes that the cause of death may be due to metabolic dysfunction caused by depletion of adenine nucleotides. Barrette et al. studied the loss of adenine nucleotides by studying the energy charge of HOCl-exposed cells and found that cells exposed to HOCl were unable to step up their energy charge after addition of nutrients. The conclusion was that exposed cells have lost the ability to regulate their adenylate pool, based on the fact that metabolite uptake was only 45% deficient after exposure to HOCl and the observation that HOCl causes intracellular ATP hydrolysis. It was also confirmed that, at bacteriocidal levels of HOCl, cytosolic components are unaffected. So it was proposed that modification of some membrane-bound protein results in extensive ATP hydrolysis, and this, coupled with the cells inability to remove AMP from the cytosol, depresses metabolic function. One protein involved in loss of ability to regenerate ATP has been found to be ATP synthetase. Much of this research on respiration reconfirms the observation that relevant bacteriocidal reactions take place at the cell membrane.Inhibition of DNA replication
Recently it has been proposed that bacterial inactivation by HOCl is the result of inhibition of DNADNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
replication. When bacteria are exposed to HOCl, there is a precipitous decline in DNA synthesis
DNA synthesis
DNA synthesis commonly refers to:*DNA replication - DNA biosynthesis *Polymerase chain reaction - enzymatic DNA synthesis *Oligonucleotide synthesis - chemical synthesis of nucleic acids...
that precedes inhibition of protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
synthesis, and closely parallels loss of viability. During bacterial genome replication, the origin of replication
Origin of replication
The origin of replication is a particular sequence in a genome at which replication is initiated. This can either be DNA replication in living organisms such as prokaryotes and eukaryotes, or RNA replication in RNA viruses, such as double-stranded RNA viruses...
(oriC in E. coli) binds to proteins that are associated with the cell membrane, and it was observed that HOCl treatment decreases the affinity of extracted membranes for oriC, and this decreased affinity also parallels loss of viability. A study by Rosen et al. compared the rate of HOCl inhibition of DNA replication of plasmids with different replication origins and found that certain plasmids exhibited a delay in the inhibition of replication when compared to plasmids containing oriC. Rosen’s group proposed that inactivation of membrane proteins involved in DNA replication are the mechanism of action of HOCl.
Protein unfolding and aggregation
HOCl is known to cause post-translational modifications to proteins, the notable ones being cysteineCysteine
Cysteine is an α-amino acid with the chemical formula HO2CCHCH2SH. It is a non-essential amino acid, which means that it is biosynthesized in humans. Its codons are UGU and UGC. The side chain on cysteine is thiol, which is polar and thus cysteine is usually classified as a hydrophilic amino acid...
and methionine
Methionine
Methionine is an α-amino acid with the chemical formula HO2CCHCH2CH2SCH3. This essential amino acid is classified as nonpolar. This amino-acid is coded by the codon AUG, also known as the initiation codon, since it indicates mRNA's coding region where translation into protein...
oxidation. A recent examination of HOCl's bactericidal role revealed it to be a potent inducer of protein aggregation. Hsp33, a chaperone known to be activated by oxidative heat stress, protects bacteria from the effects of HOCl by acting as a holdase, effectively preventing protein aggregation. Strains of E. coli and Vibrio cholerae lacking Hsp33 were rendered especially sensitive to HOCl. Hsp33 protected many essential proteins from aggregation and inactivation due to HOCl, which is a probable mediator of HOCl's bactericidal effects.
Hypochlorites
Hypochlorites are the salts of hypochlorous acid; commercially important hypochlorites are calcium hypochloriteCalcium hypochlorite
Calcium hypochlorite is a chemical compound with formula 2. It is widely used for water treatment and as a bleaching agent...
and sodium hypochlorite.
Production of hypochlorites using electrolysis
Solutions of hypochlorites can be produced by electrolysis of an aqueous chloride solution. Chlorine gas is produced at the anode, while hydrogen forms at the cathode. Some of the chlorine gas produced will dissolve forming hypochlorite ions. Hypochlorites are also produced by the disproportionationDisproportionation
Disproportionation, also known as dismutation is used to describe a specific type of redox reaction in which a species is simultaneously reduced and oxidized so as to form two different products....
of chlorine gas in alkaline solutions.