Sodium 6-amino-5-[[2-[(cyclohexylmethylamino)sulphonyl]phenyl]azo]-4-hydroxynaphthalene-2-sulphonate

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics

Type of information:

other: Expert statement

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The absorption, distribution, metabolism and excretion of FAT 20003/K have been predicted in the absence of toxicokinetic studies. There are no guidelines on how to conduct this type of modelling but the methods described are well accepted scientifically.

Data source

Reference

Reference Type:

other: Expert statement

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Principles of method if other than guideline:

The absorption, distribution, metabolism and excretion of FAT 20003/K have been predicted in the absence of toxicokinetic studies. There are no guidelines on how to conduct this type of modelling but the methods described are well accepted scientifically.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

Sodium 6-amino-5-[[2-[(cyclohexylmethylamino)sulphonyl]phenyl]azo]-4-hydroxynaphthalene-2-sulphonate

EC Number:

251-257-2

EC Name:

Sodium 6-amino-5-[[2-[(cyclohexylmethylamino)sulphonyl]phenyl]azo]-4-hydroxynaphthalene-2-sulphonate

Cas Number:

32846-21-2

Molecular formula:

C23H26N4O6S2.Na

IUPAC Name:

sodium 6-amino-5-({2-[cyclohexyl(methyl)sulfamoyl]phenyl}diazenyl)-4-hydroxynaphthalene-2-sulfonate

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Administration / exposure

Statistics:

None

Results and discussion

Toxicokinetic / pharmacokinetic studies

Details on absorption:

Absorption

Oral route

At the low pH in the stomach (approximately pH 2), FAT 20003 K is likely to be converted from the salt to the free acid, which is expected to have a lower water solubility than the salt (as predicted by EPIWIN QSAR). However, the pH in the GI tract moves from acidic to alkaline, so there is opportunity for complete dissolution to occur whilst inside the small intestine, which would facilitate absorption The moderate log Pow indicates that FAT 20003 K is likely to be absorbed by passive diffusion, although the molecular weight of greater than 500 is likely to reduce this absorption to some degree.

A combined repeated dose oral toxicity study with the reproduction/ developmental toxicity screening test was performed in Wistar rats. No mortality occurred in the control or any of the dose groups during the study period and there were no adverse effects of toxicological relevance on any of the parameters investigated (clinical signs, FOB, body weight, food consumption, haematology, blood coagulation, clinical biochemistry, urine, organ weights, pathology, histopathology, reproduction and development of the pups). FAT 20003 K is a red dye and might be expected to cause discolouration of the internal organs, although it is uncertain if the colour would be discernible from the tissue colour. However, the colour would be obvious if present in urine, assuming that Phase 1 biotransformation did not remove the colour by oxidation of the conjugated electron system in the molecule. The study reports that the urine colour was generally unaffected indicating that no (visually-detectable) FAT 20003 K is present in the urine. However, a lack of red colour in the urine does not prove that absorption of the parent molecule did or did not occur, as the primary excretion could be via bile based on a molecular weight greater than 300.

The lack of oral toxicity seen in the repeated dose study is supported by the acute oral toxicity study. No mortalities occurred in the control or dose groups during the study period and there were no observed substance related gross organ changes.

FAT 20003 K therefore has the potential to undergo absorption from the GI tract, although toxicological studies show that there were no adverse toxicological effects following oral exposure. There is no information to deduce the extent of any oral absorption, although the red-stained faeces could be indicative that a significant amount passed through the GI tract unabsorbed, although some could have been absorbed then excreted back into the intestine via the biliary duct.

The OECD QSAR application toolbox was used to apply Lipinski's Rule of Five. FAT 20003 K was predicted to be not orally bioavailable based on these rules. For clarification, bioavailability is not the same as absorption in this context. A molecule can be absorbed in the GI tract but then fully metabolised in the liver or excreted via biliary duct and therefore not bioavailable to the systemic circulation. The toolbox is also designed to predict possible metabolites that can be produced by phase 1(e.g. oxidation/reduction) and phase 2 (e.g. conjugation) biotransformations in the liver, based on the structure of the parent molecule. This biotransformation can occur during the first pass effect, but can also occur if the unmetabolised molecule passes into the systemic circulation and returns through the liver.

Based on the available evidence, it can be concluded that FAT 20003K has the potential for absorption to a limited extent, via the oral route.
Inhalation route

There is no information available regarding the absorption or toxicity of FAT 20003 K via inhalation.

The vapour pressure of FAT 20003 K is predicted to be very low indicating that inhalation exposure from volatilisation is unlikely to be a potential route of exposure.

The particle size (L50D) of the test material is 4.14 µm. REACH endpoint specific guidance (R.7c) states that respirable particles are < 15 µm. However, according to other REACH guidance (chapter R.14) respirable particles are regarded as being < 10 µm. Therefore, assessment of potential respiration will consider both these limits. There are more than 95% of particles which are < 10 µm, and 60% which are less than 5 µm and therefore, there is a very high potential for deposition in the alveoli .The substance is then sufficiently soluble to dissolve in the alveolar fluid and be fully absorbed. Larger particles would mainly be cleared from the lungs by the mucociliary escalator and then swallowed making them available for absorption via the GI tract.


Dermal route

There is no information available regarding the absorption or toxicity of FAT 20003 K following dermal exposure. Dermal absorption is influenced inter alia by water solubility, log Pow and molecular weight. REACH endpoint specific guidance (R.7c) indicates that the water solubility (100-10000 mg/L) and log Pow ( >1, < 4) of FAT 20003 K favours dermal absorption, but the molecular weight above 500 reduces the likelihood. However, the REACH guidance on dermal absorption only allows a reduction from 100% absorption to 10% if molecular weight is > 500 and log P is < 1 or > 4. In this case, only one of these criteria is met. In conclusion, it is considered that the rate of dermal absorption would be low due to the molecular weight of FAT 20003 K.

Details on distribution in tissues:

Distribution

There is minimal information available relating to the distribution of FAT 20003/K.

The OECD toolbox predicts that FAT 20003 K will not be bioavailable. If this is the case, then it will not distribute to the systemic circulation. The substance is unlikely to bioaccumulate in fat, based on the physico-chemical properties (log Pow 1.07, moderate water solubility, molecular weight > 500 and structure).

The repeated dose oral toxicity study does not indicate any significant discolouration of the internal organs although the red colour might be indiscernible from the tissue colour.

Overall, the indications are that distribution of FAT 20003 K would be minimal.

Details on excretion:

Excretion

The repeated dose toxicity study reports that the faeces were stained a red colour. This indicates several possibilities; there is also the possibility that the red colour could be due to both unabsorbed parent and excreted parent/metabolite(s). Several of the metabolites predicted by the OECD toolbox have intact conjugation and hence could therefore retain the red-coloured of the parent. The molecular weights of the parent and coloured metabolites are much greater than 300 and this increases the likelihood of excretion via the biliary duct rather than via urine, which could also explain the lack of red colour in the urine.

However, the OECD toolbox also predicts certain metabolites that would not be expected to be coloured, have molecular weights less than 300 and which are also quite polar due to the phase I and II transformations in the liver. Therefore, excretion of these metabolites in the urine cannot be excluded.

In summary, excretion of FAT 20003K is expected via bile and GI tract, but not via urine. Excretion via bile is expected for metabolites with molecular weights greater than 300.. Excretion of smaller non-coloured metabolites via urine is possible.

Metabolite characterisation studies

Details on metabolites:

Metabolism
Although the test material was stable to hydrolysis at environmental pHs (pH 4-9), at the pH in the stomach FAT 20003 K may undergo hydrolysis leading to smaller molecules that have an increased likelihood of being absorbed. In addition to this, there is a possibility that the azo group could be reduced to an amino by the action of intestinal microflora, with subsequent absorption of the amine compound.

In addition to this, potential metabolites of FAT 20003 K in the liver and skin have been predicted using OECD Toolbox (Table 1). Twenty-seven metabolites were predicted for liver metabolism (15 are bioavailable and 12 not bioavailable) and 28 metabolites for skin metabolism (13 are bioavailable and 15 not bioavailable). Six metabolites were common in both liver and skin (M1, M5, M6, M11, M13 and M14).
The main predicted biotransformations in liver and skin include breakage of the azo bond, hydroxylation reactions, demethylation and oxidation, which are considered as the normal types of phase I and phase II transformations. It is clear from the number of predicted metabolites that FAT 20003 K can potentially undergo a significant degree of metabolism.

With the exception of formaldehyde, the SMILES codes for all the above predicted metabolites were entered into both PubChem and ChemSpider websites, but no identification was found.

Toxicologically relevant metabolites
Formaldehyde:
The first obvious metabolite that is known to be potentially toxic is formaldehyde. This is regarded as carcinogenic to humans (group 1) according to the Hazardous Substance DataBase (HSDB).
However, it should be stressed that the underlying data for this is based on the inhalation route, and confined to site-specific respiratory neoplasms, specifically nasal area The data is also related to inhalation of formaldehyde, whereas in this case any inhalation will be for the FAT 20003 K. It is also expected that any formation of formaldehyde in vivo will be rapidly followed by phase I transformation (oxidation) to formic acid and this would be rapidly excreted probably as a formate salt. Therefore, if this possible metabolite forms, it is not considered to present any concern.

Aromatic C-nitroso metabolites:
Due to the presence of the nitroso group, this predicted metabolite warranted further attention. However, unlike the N-nitroso compounds, a generic search (google) for aromatic C-nitroso compounds did not reveal any information of particular concern. An analogous compound containing the nitrosonaphthyl moiety (2-nitroso-1-napthol) was found on the HSDB, but there was very little animal toxicity data (LD50 mice (i.p.) 42 mg/kg). The CCRIS database indicated a negative Ames test on 2-nitroso-1-naphthol, on the TA98 Salmonella strain. However, potential carcinogenic effects cannot be excluded based on the presence of the nitroso- functional group.

Metabolites with aromatic –OH and –NH2 present:
Where there are no structural alerts and the metabolites are similar to the parent, it is expected that the toxicity will be similar to the parent. However, for those metabolites where an aromatic amine is part of the structure, enhanced toxicity compared to the parent, cannot be excluded. It is known that certain aromatic amines are associated with human carcinogenicity and carcinogenicity in rat. This does not mean that these metabolites are carcinogens, but the potential is considered to be there.

In addition, all the metabolites which have phenolic groups present should also be considered potentially more toxic than the parent, due to the oxidation of phenols which is known to produce quinones/ semi-quinones which can become involved in binding to the –SH or –NH2 groups in proteins leading to inactivation of the protein. They are also associated with superoxide anion production ultimately leading to formation of hydroxyl radicals which can cause cell damage.

Bioaccessibility (or Bioavailability)

Bioaccessibility (or Bioavailability) testing results:

No data

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
There is insufficient information available to be able to determine the rate and extent of any absorption that may occur and the true toxicokinetic pathway of FAT 20003/K.

Executive summary:

The absorption, distribution, metabolism and excretion of FAT 20003/K have been predicted in the absence of toxicokinetic studies. The test substance is expected to have limited potential for oral absorption and the uptake via the inhalation route is expected to be potentially high, due to particle size. FAT 20003/K is expected to have low dermal absorption. Indications are that distribution of FAT 20003/K in the body would be minimal. A significant number of potential metabolites are predicted using the OECD toolbox (table 1). Some of these metabolites could be toxicologically relevant with potentially enhanced toxicity relative to the parent. Excretion of FAT 20003/K is expected via bile and GI tract, but not via urine. Excretion via bile is expected for metabolites with molecular weights greater than 300. Excretion of smaller non-coloured metabolites via urine is possible. Although there is a lack of information available regarding the toxicokinetic pathway of FAT 20003/K, there was no evidence of toxicity seen in the repeated dose toxicity study.