Quaternary ammonium compounds, dicoco alkyldimethyl, nitrites

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Study period:

From February 10, 2004 to March 13, 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Read across justification is presented from the structurally analogous quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides to the quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites.

In the stomach the gastic juice is acidic, made up of acids and enzymes. In such an evironment it is highly unlikely that the quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites substance (s) will remain ionically bound to each other and thus are prone to dissociation in which case the released cation(s) will associate with other anions and the released anion will associate with cations. Thererfore, it is suggested read-across that data from the corresponding quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides is considered approriate in that such substances are likely to dissociate in a similar manner.

Furthermore, in 1988, the US EPA, Office of Pesticides and Toxic Substances issued a Notice to producers, Formulators, Distributors and Registrants regarding quaternary ammonium compounds with regard to "Clustering" of such quaternary ammonium compounds.

Prior to this, EPA had required each quat compound to be individually coded and registered as a new chemical, even when the chemical structure of individual compounds differed only slightly in alkyl distribution and chain lengths. This procedure was continued with the new generations of quats having two, three, and four chains. As a result, EPA records showed that some 211 registered technical grade active ingredient products containing varying concentrations of Quats, each coded separately on the basis of alkyl chain length and percentage carbon distribution within the chain. At this time, there are approximately eight to ten thousands (8-10,000) registered end-use formulations.

However, questions were raised regarding whether the EPA could cluster or group the quats and pick one or more representative members of each cluster to be used in toxicity studies, instead of requiring separate studies on each quat. These same questions were raised when the EPA issued its March 4, 1987 Data Call-In Notice requiring all registrants of antimicrobial active ingredients to submit subchronic and chronic toxicological data to support the continued registration of their products.

In response to these questions, EPA·solicited information from industry, the public, academia, industry cooperative work groups, the state of California, and Canada. EPA then reviewed all of the assembled information along with the chemical structure of most of the quats. Based on the results of this review, EPA developed the following four groupings of currently registered quat compounds:

Group I. The alkyl or hydroxyalkyl (straight chain) substituted Quats
Group II. The non-halogenated benzyl substituted Quats (includes hydroxybenzyl, ethylbenzyl, hydroxyethybenzyl, napthylmethyl, dodecylbenzyl, and alkyl benzyl)
Group III. The di-and tri-chlorobenzyl substituted
Group IV. Quats with unusual substituents (charged heterocyclic ammonium compounds).

Fundamental to this discussion EPA determined that "X-" in all of these structures would be attributed to "any anionic species". Therefore, this would mean in terms of toxicological evaluation the coutner anion in such quaternary ammonium compounds could be regarded as; e.g halogen (Cl-, Br-, I-,), saccharinate or cyclohexylsulphamate. It is therefore suggested here that nitrite (NO2-) could also be regarded as a pertinent anion.

Since the US EPA deem that such a clustering of structures for toxicological evaluation is well founded then it would seem that to consider read-across data from quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides to the closely structurally analogous quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites to be equally justifiable.

Furthermore, in certain organic solvents it has been reported that the exchange constants between nitrite and chloride in quaternary ammonium salts (QAS) are approximately equal. [Zhurnal Analiticheskoi Khimii, 2010, Vol. 65, No. 6, pp. 579–584. (E.M. Rakhman’ko, M.S. Markovskaya, L.S. Stanishevskii, Yu.S. Zubenko, A.R. Tsyganov)]

Reason / purpose for cross-reference:

read-across: supporting information

Objective of study:

absorption

distribution

excretion

metabolism

Qualifier:

according to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Specific details on test material used for the study:

- Name of test material (as cited in study report): Arquad 2.10-40
- Physical state: Colourless to pale yellowish liquid
- Analytical purity: 40.5 % Didecyldimethylammonium chloride (CAS No 7173-51-5)
- Composition of test material, percentage of components: 40.2 % of DDAC, amine 0.43 %, Amine HCl 0.39 %,
- Purity test date: Oct. 20, 2004
- Lot/batch No.: WIR03048
- Storage condition of test material: At room temperature

Radiolabelled Material: [Methyl-14C]DDAC
- Physical state: Colourless liquid
- Radiochemical purity (if radiolabelling): 98 % (by HPLC), 99.2 % (by TLC)
- Specific activity (if radiolabelling): 1.11 GBq/mmol; 30.1 mCi/mmol (by gravimetric analysis): 1.22 GBq/mmol; 33 mCi/mmol
- Radioactive concentration: 148 MBq/mL, 4 mCi/mL
- Batch Number: CFQ13640
- Purity test date: Nov. 19, 2003
- Expiration date of radiochemical substance (if radiolabelling):
- Storage condition of test material: -20 °C in the absence of light and air is recommended

Radiolabelling:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories France, L’Arbresle, France. Caesarean Obtained, Barrier Sustained-Virus Antibody Free (COBS-VAF®).
- Age at study initiation: 7 wk old; for the bile collection group, animals were around 10 wk (males) 12 wk females).
- Weight at study initiation: 229 g males and 159 g females
- Fasting period before study: Overnight
- Housing: 2 or 3 animals/cage in oral groups and singly in dermal and bile collection group
- Diet (e.g. ad libitum): A04 C pelleted diet ad libitum
- Water (e.g. ad libitum): Filtered tap water ad libitum
- Acclimation period: At least 7 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22± 2 °C
- Humidity (%): 50±20%
- Air changes (per hr): 12 cycles/h of filtered, non-recycled air
- Photoperiod: 12 h light/ 12 h dark

Route of administration:

other: Oral gavage and dermal

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:

VEHICLE
- Amount of vehicle (if gavage): 10 mL/kg

TEST SITE
- Area of exposure: Interscapular/upper back region
- % coverage: 10 % (25 -30 cm2 for 250-300 g rat)

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Washed to remove all traces of test material by wiping with at least 3 cotton swabs dampened with water and diluted normal hand soap solution.
- Time after start of exposure: 6 h

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 1.5 mL of dosage form/kg ( Dose: 1.5 mg/kg)

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes

Duration and frequency of treatment / exposure:

6 h

Dose / conc.:

50 mg/kg bw/day (nominal)

Remarks:

Oral

Dose / conc.:

200 mg/kg bw/day

Remarks:

Oral

Dose / conc.:

1.5 mg/kg bw/day

Remarks:

Dermal

Dose / conc.:

15 mg/kg bw/day

Remarks:

Dermal

No. of animals per sex per dose / concentration:

9/sex/dose - Plasma/blood pharmacokinetics (Group 1-5)
5/sex/dose - Excretion balance (Group 6-8)
5/sex/dose bile collection (Group 9)

Control animals:

other: Yes: For the purposes of pre-dose sample analysis, plasma, blood and tissues will be collected from at least one untreated supplementary animal/sex using the above mentioned procedures.

Positive control reference chemical:

Not applicable

Details on study design:

- Rationale for animal assignment (if not random): Random

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled : Blood, plasma, serum, urine, feces, cage washes, bile, skin-application site
- Time and frequency of sampling: Oral groups: 0.5, 1, 2, 4, 8, 24, 48, 72 and 96 h post gavage on Day 1 (Groups 1, 2 and 5)
Dermal group: 3, 6, 7, 8, 10, 16, 24, 48 and 72 h post dermal application (Groups 3 and 4)
Excreta and tissue collection: 0-24, 24-48, 48-72, 96-120, 120-144 and 144-168 h after the radioactive gavage/dermal application (Group 6-8)
Bile collection: 0-3, 3-6, 6-12 and 12-24 h post injection/gavage (Group 9)

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled : urine, faeces, tissues, cage washes, bile
- Method type(s) for identification : Radio- HPLC for urine samples and LSC for biological samples

Statistics:

None

Type:

absorption

Results:

Low oral bioavailability (0.93 to 3.16 %); rapidly eliminated within a 48 h period

Type:

distribution

Results:

Radioactive levels generally decreased over time in all tissues

Type:

excretion

Results:

A mean 2.57±2.39 (males) and 1.14±0.69 % (females) of the radioactive dose was recovered over the 24 h period in the bile

Details on distribution in tissues:

Oral gavage:
50 mg/kg (single dose): The mean radioactivity levels were below quantifiable limits (Group 1) in all tissues except for intestines, kidneys and liver
200 mg/kg (single dose): The mean radioactivity levels were above quantifiable limits (Group 2) in half analysed tissues and organs at 24 h. Specifically high levels were present in adrenals, abdominal fat, eyes, heart, kidneys, liver, lungs, lymph nodes and pancreas.
50 mg/kg (repeated dose): The mean radioactivity levels were below quantifiable limits (Group 1) in all tissues except for intestines, kidneys, liver and mesenteric lymph nodes
Dermal application:
Single application (1.5 mg/kg): Radioactivity levels were below quantifiable limits in all tissues/organs at all time points except for the stripped skin from the application site and intestine (15 mg/kg)
Single application (15 mg/kg): Radioactivity levels were below quantifiable limits in all tissues/organs at all time points except for the stripped skin from the application site, adrenals, heart, kidneys, liver and lungs

Details on excretion:

Following single oral gavage at a nominal dose level of 50 mg/kg to rats, a mean 2.57±2.39 (males) and 1.14±0.69 % (females) of the radioactive dose was recovered over the 24 h period in the bile

Key result

Toxicokinetic parameters:

other: Oral: Mean plasma and blood radioactivity levels were all below quantifiable limits

Key result

Toxicokinetic parameters:

other: Dermal: Mean plasma and blood radioactivity levels were all below quantifiable limits

Metabolites identified:

no

Details on metabolites:

No metabolites nor parent drug were found in urine samples

Conclusion: Low dermal and oral absorption. The actual minimal fraction of the oral dose absorbed was 0.93 to 3.16%; this was eliminated rapidly, essentially within a 48-hour period.

Conclusions:

Interpretation of results (migrated information): no bioaccumulation potential based on study results.

Under the test conditions, the test material (containing 40.5 % DDAC) has low dermal and oral absorption. The actual minimal fraction of the oral dose absorbed was 0.93 to 3.16 %; this was eliminated rapidly, essentially within a 48 h period.

Executive summary:

The toxicokinetics of the test material was investigated in an OECD guideline 417conducted to GLP.

This study is assigned a reliability score of 1 in accordance with the criteria for assessing data quality set forth by Klimisch et al (1997).

The study was conducted to investigate the blood and plasma pharmacokinetics, tissue distribution and mass balance of total radioactivity of test material [14C-DDAC] following single dermal administration (at 1.5 and 15 mg/kg, as 6 h exposure over 10% of the body surface) and single (at 50 and 200 mg/kg) and repeated (at 50 mg/kg/d) oral gavage administrations to male and female Sprague-Dawley rats.   In addition, the elimination of radioactivity in bile after single oral administration at 50 mg/kg was investigated. Investigations included blood and plasma pharmacokinetics, tissue distribution and mass balance of total radioactivity.

The animals were divided into 9 groups; groups 1 to 5 (each of 9 animals/sex) for plasma/blood pharmacokinetics of radioactivity and tissue distribution, groups 6 to 8 (each of 5 animals/sex) principally for excretion balance and group 9 (4 animals/sex) for bile collection. Animals of groups 1 to 4 and 6 and 7 were treated once with the radiolabelled test item. Animals of groups 5 and 8 were treated once per day for 6 d with the unlabelled test item, followed by a single administration of the radiolabelled test material on Day 7.

Reference 2

Endpoint:

basic toxicokinetics in vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Study period:

1996

Reliability:

2 (reliable with restrictions)

Justification for type of information:

Since in the stomach the gastic juice is acidic, made up of acids and enzymes, it is suggested that in this evironment it is highly unlikely that the quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites substance (s) will remain ionically bound to each other and thus are prone to dissociation in which case the released cation(s) will associate with other anions and the released anion will associate with cations. Thererfore, it is suggested that read-across data from sodium nitrite is considered appropriate in order to arrive at a conclusion in terms of any health effects that may be incurred from exposure to "nitrite" whichever cation it may be associated with.

In addition, in 1988, the US EPA, Office of Pesticides and Toxic Substances issued a Notice to producers, Formulators, Distributors and Registrants regarding quaternary ammonium compounds with regard to "Clustering" of such quaternary ammonium compounds.

Prior to this, EPA had required each quat compound to be individually coded and registered as a new chemical, even when the chemical structure of individual compounds differed only slightly in alkyl distribution and chain lengths. This procedure was continued with the new generations of quats having two, three, and four chains. As a result, EPA records showed that some 211 registered technical grade active ingredient products containing varying concentrations of quats, each coded separately on the basis of alkyl chain length and percentage carbon distribution within the chain. At this time, there are approximately eight to ten thousands (8-10,000) registered end-use formulations.

However, questions were raised regarding whether the EPA could cluster or group the quats and pick one or more representative members of each cluster to be used in toxicity studies, instead of requiring separate studies on each quat. These same questions were raised when the EPA issued its March 4, 1987 Data Call-In Notice requiring all registrants of antimicrobial active ingredients to submit subchronic and chronic toxicological data to support the continued registration of their products.

In response to these questions, EPA solicited information from industry, the public, academia, industry cooperative work groups, the state of California, and Canada. EPA then reviewed all of the assembled information along with the chemical structure of most of the quats. Based on the results of this review, EPA developed the following four groupings of currently registered quat compounds:

Group I. The alkyl or hydroxyalkyl (straight chain) substituted Quats
Group II. The non-halogenated benzyl substituted Quats (includes hydroxybenzyl, ethylbenzyl, hydroxyethybenzyl, napthylmethyl, dodecylbenzyl, and alkyl benzyl)
Group III. The di-and tri-chlorobenzyl substituted
Group IV. Quats with unusual substituents (charged heterocyclic ammonium compounds).

Fundamental to this discussion EPA determined that "X-" in all of these structures would be attributed to "any anionic species". Therefore, this would mean in terms of toxicological evaluation the coutner anion in such quaternary ammonium compounds could be regarded as; e.g halogen (Cl-, Br-, I-,), saccharinate or cyclohexylsulphamate. It is therefore suggested here that nitrite (NO2-) could also be regarded as a pertinent anion.

Since the US EPA deem that such a clustering of structures for toxicological evaluation is well founded and that the counter anion could be regarded as "any anionic species" then it would seem that to consider available toxicological data on sodium nitrite, in order to evaluate any health effects that may be incurred from exposure to the nitrite anion (NO2-), is justifiable.

Reason / purpose for cross-reference:

read-across: supporting information

Objective of study:

absorption

distribution

excretion

metabolism

GLP compliance:

not specified

Radiolabelling:

not specified

Species:

other: Dogs, sheep and Ponies

Strain:

not specified

Sex:

not specified

Route of administration:

intravenous

Vehicle:

not specified

Control animals:

not specified

Nitrite in blood is highly reactive with haemoglobin and causes methaemoglobinaemia. The oxygen-carrying capacity of methaemoglobin is much less than that of haemoglobin.

Following intravenous administration of nitrite, the plasma half-life in the distribution stage in dogs, sheep, and ponies was found to be 48, 12, and 5 minutes, respectively. Oral administration of nitrite to experimental animals is not usually followed by detectable levels in tissues and bodily fluids, presumably due to the rapidity with which nitrite is oxidized to nitrate (Nitrate and nitrite are readily interconverted by oxidation-reduction reactions in biological systems [Reduction of nitrate (NO3-) to nitrite (NO2-) occurs by mammalian nitrate reductase and nitrate reductase activity of microorganisms in the oral cavity and upper gastrointestinal tract]. Thus the pharmacokinetics and metabolism of nitrate and nitrite cannot adequately be considered separately from one another; both are also intimately connected to the potential for in vivo formation of N-nitroso compounds.

Nitrite is not a normal constituent of urine in humans or animals; its presence is considered to indicate a bacterial infection of the urinary tract.

In blood, nitrite undergoes a coupled oxidation reaction with oxyhemoglobin to produce methemoglobin (Ferrous iron associated with haemoglobin is oxidized by nitrite to ferric iron, leading to the formation of methaemoglobin).

Unlike Hb, MetHb cannot exchange oxygen; hence, the presence of excess MetHb in the circulation proportionately reduces the ability of the blood to transfer oxygen. The rate of MetHb formation is species dependent, with the rate in humans being intermediate to the rates found in ruminants and pigs.

Nitrite, or the nitrosating species derived from it (N2O3 and N2O4), can combine in vitro or in vivo with secondary amines (such as ethylurea, DNA primidine and purine bases) or N-alkylamides to form, respectively, carcinogenic nitrosamines (R1NNOR2) and nitrosamides (R1NNO.COR2).

Humans are exposed to a wide range of amines through endogenous production or ingestion of numerous foods, beverages and medicines.

Conclusions:

Following intravenous administration of nitrite, the plasma half-life in the distribution stage in dogs, sheep, and ponies was found to be 48, 12, and 5 minutes, respectively. Oral administration of nitrite to experimental animals is not usually followed by detectable levels in tissues and bodily fluids, presumably due to the rapidity with which nitrite is oxidized to nitrate (Nitrate and nitrite are readily interconverted by oxidation-reduction reactions in biological systems [Reduction of nitrate (NO3-) to nitrite (NO2-) occurs by mammalian nitrate reductase and nitrate reductase activity of microorganisms in the oral cavity and upper gastrointestinal tract]. Thus the pharmacokinetics and metabolism of nitrate and nitrite cannot adequately be considered separately from one another; both are also intimately connected to the potential for in vivo formation of N-nitroso compounds.

Executive summary:

In a publication entitled "The metabolism of dietary nitrates and nitrites", (Biochem Soc Trans 1996;24(3):780-5, Walker R.) the metabolism of dietary nitrates and nitrites was investigated.

This study is assigned a reliability score of 2 in accordance with the criteria for assessing data quality set forth by Klimisch et al (1997).

In blood, nitrite undergoes a coupled oxidation reaction with oxyhemoglobin to produce methemoglobin (Ferrous iron associated with haemoglobin is oxidized by nitrite to ferric iron, leading to the formation of methaemoglobin).

Unlike Hb, MetHb cannot exchange oxygen; hence, the presence of excess MetHb in the circulation proportionately reduces the ability of the blood to transfer oxygen. The rate of MetHb formation is species dependent, with the rate in humans being intermediate to the rates found in ruminants and pigs.

Nitrite, or the nitrosating species derived from it (N2O3 and N2O4), can combine in vitro or in vivo with secondary amines (such as ethylurea, DNA primidine and purine bases) or N-alkylamides to form, respectively, carcinogenic nitrosamines (R1NNOR2) and nitrosamides (R1NNO.COR2).

Humans are exposed to a wide range of amines through endogenous production or ingestion of numerous foods, beverages and medicines.

Reference 3

Endpoint:

basic toxicokinetics in vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Study period:

1980

Reliability:

2 (reliable with restrictions)

Justification for type of information:

Since in the stomach the gastic juice is acidic, made up of acids and enzymes, it is suggested that in this evironment it is highly unlikely that the quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites substance (s) will remain ionically bound to each other and thus are prone to dissociation in which case the released cation(s) will associate with other anions and the released anion will associate with cations. Thererfore, it is suggested that read-across data from sodium nitrite is considered appropriate in order to arrive at a conclusion in terms of any health effects that may be incurred from exposure to "nitrite" whichever cation it may be associated with.

In addition, in 1988, the US EPA, Office of Pesticides and Toxic Substances issued a Notice to producers, Formulators, Distributors and Registrants regarding quaternary ammonium compounds with regard to "Clustering" of such quaternary ammonium compounds.

Prior to this, EPA had required each quat compound to be individually coded and registered as a new chemical, even when the chemical structure of individual compounds differed only slightly in alkyl distribution and chain lengths. This procedure was continued with the new generations of quats having two, three, and four chains. As a result, EPA records showed that some 211 registered technical grade active ingredient products containing varying concentrations of quats, each coded separately on the basis of alkyl chain length and percentage carbon distribution within the chain. At this time, there are approximately eight to ten thousands (8-10,000) registered end-use formulations.

However, questions were raised regarding whether the EPA could cluster or group the quats and pick one or more representative members of each cluster to be used in toxicity studies, instead of requiring separate studies on each quat. These same questions were raised when the EPA issued its March 4, 1987 Data Call-In Notice requiring all registrants of antimicrobial active ingredients to submit subchronic and chronic toxicological data to support the continued registration of their products.

In response to these questions, EPA solicited information from industry, the public, academia, industry cooperative work groups, the state of California, and Canada. EPA then reviewed all of the assembled information along with the chemical structure of most of the quats. Based on the results of this review, EPA developed the following four groupings of currently registered quat compounds:

Group I. The alkyl or hydroxyalkyl (straight chain) substituted Quats
Group II. The non-halogenated benzyl substituted Quats (includes hydroxybenzyl, ethylbenzyl, hydroxyethybenzyl, napthylmethyl, dodecylbenzyl, and alkyl benzyl)
Group III. The di-and tri-chlorobenzyl substituted
Group IV. Quats with unusual substituents (charged heterocyclic ammonium compounds).

Fundamental to this discussion EPA determined that "X-" in all of these structures would be attributed to "any anionic species". Therefore, this would mean in terms of toxicological evaluation the coutner anion in such quaternary ammonium compounds could be regarded as; e.g halogen (Cl-, Br-, I-,), saccharinate or cyclohexylsulphamate. It is therefore suggested here that nitrite (NO2-) could also be regarded as a pertinent anion.

Since the US EPA deem that such a clustering of structures for toxicological evaluation is well founded and that the counter anion could be regarded as "any anionic species" then it would seem that to consider available toxicological data on sodium nitrite, in order to evaluate any health effects that may be incurred from exposure to the nitrite anion (NO2-), is justifiable.

Reason / purpose for cross-reference:

read-across: supporting information

GLP compliance:

not specified

Radiolabelling:

not specified

Species:

rat

Strain:

Sprague-Dawley

Route of administration:

oral: gavage

Vehicle:

water

Control animals:

not specified

This publication reported that maximum levels of methaemoglobin (45 – 80%) were reached one hour after dosing Sprague-Dawley rats with 150 mg/kg bw sodium nitrite. The concentration

returned to normal after 24 hours if the animal survived.

Conclusions:

In in vivo Studies on Methemoglobin Formation by Sodium Nitrite the maximum levels of methaemoglobin (45 – 80%) were reached one hour after dosing Sprague-Dawley rats with 150 mg/kg bw sodium nitrite. The concentration returned to normal after 24 hours in surviving animals.

Executive summary:

Methemoglobin Formation by Sodium Nitrite was investigated in a publication entitled "In vivo Studies on Methemoglobin Formation by Sodium Nitrite" (International Archives of Occupational and Environmental Health 45, 97-104, 1980, Imaizumi K, Tyuma I, Imai K, Kosaka H, and Ueda Y)

This study is assigned a reliability score of 2 in accordance with the criteria for assessing data quality set forth by Klimisch et al (1997).

In in vivo Studies on Methemoglobin Formation by Sodium Nitrite the maximum levels of methaemoglobin (45 – 80%) were reached one hour after dosing Sprague-Dawley rats with 150 mg/kg bw sodium nitrite. The concentration returned to normal after 24 hours in surviving animals.

Description of key information

Read across data is presented on the "fragments" from which this substance is manufactured :-

Quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides, CAS Number 68391-05-9, EC Number 269-924-1 and from sodium nitrite, CAS Number 7632-00-0, EC Number 231-555-9.

The justification in taking this approach is as follows :-

In the stomach the gastic juice is acidic, made up of acids and enzymes. In such an evironment it is highly unlikely that the quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites substance (s) will remain ionically bound to each other and thus are prone to dissociation in which case the released cation(s) will associate with other anions and the released anion will associate with cations. Thererfore, it is suggested read-across data from the corresponding quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides is considered approriate in that such substances are likely to dissociate in a similar manner.

Furthermore, in 1988, the US EPA, Office of Pesticides and Toxic Substances issued a Notice to producers, Formulators, Distributors and Registrants regarding quaternary ammonium compounds with regard to "Clustering" of such quaternary ammonium compounds.

Prior to this, EPA had required each quat compound to be individually coded and registered as a new chemical, even when the chemical structure of individual compounds differed only slightly in alkyl distribution and chain lengths. This procedure was continued with the new generations of quats having two, three, and four chains. As a result, EPA records showed that some 211 registered technical grade active ingredient products containing varying concentrations of Quats, each coded separately on the basis of alkyl chain length and percentage carbon distribution within the chain. At this time, there are approximately eight to ten thousands (8-10,000) registered end-use formulations.

However, questions were raised regarding whether the EPA could cluster or group the quats and pick one or more representative members of each cluster to be used in toxicity studies, instead of requiring separate studies on each quat. These same questions were raised when the EPA issued its March 4, 1987 Data Call-In Notice requiring all registrants of antimicrobial active ingredients to submit subchronic and chronic toxicological data to support the continued registration of their products.

In response to these questions, EPA solicited information from industry, the public, academia, industry cooperative work groups, the state of California, and Canada. EPA then reviewed all of the assembled information along with the chemical structure of most of the quats. Based on the results of this review, EPA developed the following four groupings of currently registered quat compounds:

Group I. The alkyl or hydroxyalkyl (straight chain) substituted Quats

Group II. The non-halogenated benzyl substituted Quats (includes hydroxybenzyl, ethylbenzyl, hydroxyethybenzyl, napthylmethyl, dodecylbenzyl, and alkyl benzyl)

Group III. The di-and tri-chlorobenzyl substituted

Group IV. Quats with unusual substituents (charged heterocyclic ammonium compounds).

Fundamental to this discussion EPA determined that "X-" in all of these structures would be attributed to "any anionic species". Therefore, this would mean in terms of toxicological evaluation the coutner anion in such quaternary ammonium compounds could be regarded as; e.g  halogen (Cl-, Br-, I-,), saccharinate or cyclohexylsulphamate. It is therefore suggested here that nitrite (NO2-) could also be regarded as a pertinent anion.

Since the US EPA deem that such a clustering of structures for toxicological evaluation is well founded then it would seem that to consider read-across data from quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides to the closely structurally analogous quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites to be equally justifiable.

Similarly since the US EPA deem that the counter anion could be regarded as "any anionic species" then it would seem that to consider available toxicological data on sodium nitrite, in order to evaluate any health effects that may be incurred from exposure to the nitrite anion (NO2-), is equally justifiable.

Furthermore, in certain organic solvents it has been reported that the exchange constants between nitrite and chloride in quaternary ammonium salts (QAS) are approximately equal. [Zhurnal Analiticheskoi Khimii, 2010, Vol. 65, No. 6, pp. 579–584. (E.M. Rakhman’ko, M.S. Markovskaya, L.S. Stanishevskii, Yu.S. Zubenko, A.R. Tsyganov)]

Therefore, one study is reported on Quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides, CAS Number 68391-05-9, EC Number 269-924-1 and two publications are reported on sodium nitrite, CAS Number 7632-00-0, EC Number 231-555-9.

Quaternary ammonium compounds, di-C12-18-alkyldimethyl, chlorides, CAS Number 68391-05-9, EC Number 269-924-1

The toxicokinetics of the test material was investigated in an OECD guideline 417conducted to GLP.

The study was conducted to investigate the blood and plasma pharmacokinetics, tissue distribution and mass balance of total radioactivity of test material [14C-DDAC] following single dermal administration (at 1.5 and 15 mg/kg, as 6 h exposure over 10% of the body surface) and single (at 50 and 200 mg/kg) and repeated (at 50 mg/kg/d) oral gavage administrations to male and female Sprague-Dawley rats.   In addition, the elimination of radioactivity in bile after single oral administration at 50 mg/kg was investigated. Investigations included blood and plasma pharmacokinetics, tissue distribution and mass balance of total radioactivity.

The animals were divided into 9 groups; groups 1 to 5 (each of 9 animals/sex) for plasma/blood pharmacokinetics of radioactivity and tissue distribution, groups 6 to 8 (each of 5 animals/sex) principally for excretion balance and group 9 (4 animals/sex) for bile collection. Animals of groups 1 to 4 and 6 and 7 were treated once with the radiolabelled test item. Animals of groups 5 and 8 were treated once per day for 6 d with the unlabelled test item, followed by a single administration of the radiolabelled test material on Day 7.

Sodium nitrite, CAS Number 7632-00-0, EC Number 231-555-9 (1)

In a publication entitled "The metabolism of dietary nitrates and nitrites", (Biochem Soc Trans 1996;24(3):780-5, Walker R.) the metabolism of dietary nitrates and nitrites was investigated.

In blood, nitrite undergoes a coupled oxidation reaction with oxyhemoglobin to produce methemoglobin (Ferrous iron associated with haemoglobin is oxidized by nitrite to ferric iron, leading to the formation of methaemoglobin).

Unlike Hb, MetHb cannot exchange oxygen; hence, the presence of excess MetHb in the circulation proportionately reduces the ability of the blood to transfer oxygen. The rate of MetHb formation is species dependent, with the rate in humans being intermediate to the rates found in ruminants and pigs.

Nitrite, or the nitrosating species derived from it (N2O3 and N2O4), can combine in vitro or in vivo with secondary amines (such as ethylurea, DNA primidine and purine bases) or N-alkylamides to form, respectively, carcinogenic nitrosamines (R1NNOR2) and nitrosamides (R1NNO.COR2).

Humans are exposed to a wide range of amines through endogenous production or ingestion of numerous foods, beverages and medicines.

Sodium nitrite, CAS Number 7632-00-0, EC Number 231-555-9 (2)

Methemoglobin Formation by Sodium Nitrite was investigated in a publicatin entitled "In vivo Studies on Methemoglobin Formation by Sodium Nitrite" (International Archives of Occupational and Environmental Health 45, 97-104, 1980, Imaizumi K, Tyuma I, Imai K, Kosaka H, and Ueda Y)

In in vivo Studies on Methemoglobin Formation by Sodium Nitrite the maximum levels of methaemoglobin (45 – 80%) were reached one hour after dosing Sprague-Dawley rats with 150 mg/kg bw sodium nitrite. The concentration returned to normal after 24 hours in surviving animals.

Key value for chemical safety assessment

Bioaccumulation potential:

low bioaccumulation potential

Absorption rate - oral (%):

3.16

Additional information

Since , the "nitrite" functionality present in quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites accounts for only approximately 7% of the substance and the "quaternary ammonium compounds, di-C12 -18 -alkyldimethyl fragment" accounts for approximately 93% of the substance, it is therefore envisaged that the contribution to the toxicokinetic assessment should focus overwhelmingly on the "quaternary ammonium compounds, di-C12 -18 -alkyldimethyl fragment".

However, it should also be duly noted that Nitrite in blood is highly reactive with haemoglobin and causes methaemoglobinemia. Ferrous iron associated with haemoglobin is oxidized by nitrite to ferric iron, leading to the formation of methaemoglobin. Humans are considered to be more sensitive than rats in this respect.

With methemoglobinemia, the hemoglobin can carry oxygen, but is not able to release it effectively to body tissues.

In extreme cases the observed symptoms can be :-

• Bluish coloring of the skin
• Headache
• Fatigue
• Shortness of breath
• Lack of energy

Methaemoglobin concentrations in SD rats increased from 45% to 80% over 1 hour after an oral dose of sodium nitrite at 150 mg/kg bw and they returned to normal levels within 24 hours in surviving rats.

Given that the "nitrite" functionality present in quaternary ammonium compounds, di-C12-18-alkyldimethyl, nitrites accounts for only approximately 7%, it is anticipated that any elevated concentrations of Methaemoglobin formed is not going to be sufficient to result in the development of the symptoms listed above and within 24 / 48 hours the levels will return to normal.