Screening and categorising azo-based substances
In Stage 2 of the Inventory Multi-Tiered Assessment and Prioritisation (IMAP) Framework, we continue to carry out pre-prioritisation work. This includes screening and categorising about 3,000 unassessed azo-based substances listed in the Australian Inventory of Chemical Substances (the Inventory).
Our aim was to develop a step-wise, semi-automated screening methodology to identify azo-based substances that, with up to date and currently available knowledge, would be considered as low concern for human health.
Azo-based substances are a group of aromatic and aliphatic substances with the general chemical structure in the form of ‘R1-N=N-R2’. They are aliphatic when R1 and R2 are alkyl and aromatic if one or more of R1 or R2 are aryl. Azo-based substances are generally large molecules that are poorly absorbed across biological membranes.
Our approach to identifying azo-based substances of low concern to human health is primarily based on the potential to release aromatic amines that may have human health concerns.
The azo linkage is usually the most easily changed (labile) portion of an azo-based substance. It may easily undergo enzymatic reduction to release aromatic amines. The released aromatic amines are much more readily absorbed across biological membranes compared with the azo-based substance. These aromatic amines may potentially cause:
- human health effects related to skin sensitisation and mutagenicity as well as
- systematic toxicity including carcinogenicity
Azo-based substances are typically used as colourants and dyes. Public exposure may occur from the use of azo dyes in cosmetics and textiles. Worker exposure may occur from the dyeing process of textiles.
The risk of workers from the occupational use in dyeing is generally of low concern except in the case of mutagenicity. While the potentially high exposure is of concern for sensitisation and systemic toxicity, controls typically in place at sites where this occurs would normally reduce this.
The risk of the public is predominantly on the use of azo dyes in textiles and cosmetics.
We identified low concern azo-based substances based on the approaches in the step-wise screening methodology below. We determine substances that meet features based on the approaches to meet the Tier I criteria for human health.
Should further information become available; we would review these assessment outcomes.
Step-wise screening methodology
The step-wise screening methodology involves the following steps:
- pre-screening for cosmetic use, hazard flags, bioavailability and metabolite information
- applying the threshold of toxicological concern (TTC) approach
- assessing aromatic amines with skin sensitisation potential
- investigating aromatic amines with genotoxicity and carcinogenicity potential
We applied semi-automated pre-screening to aromatic azo-based substances using available simplified molecular-input line-entry system (SMILES) structural presentation. In collaboration with the Laboratory of Mathematical Chemistry (LMC), we built a computational workflow using the OASIS Pipeline Profiler software. The software accesses the Organisation for Economic Co-operation and Development (OECD) quantitative structure–activity relationship (QSAR) Toolbox database and the OASIS Tissue Metabolic Simulator (TIMES) software.
We simulated the metabolism (skin sensitisation and genotoxicity) of the substances using OASIS TIMES included in the OASIS Pipeline Profiler software.
We evaluated the bioavailability for azo-based substances to break down to aromatic amines. We considered the following main factors reducing bioavailability:
- high molecular weight
- high degree of sulfonation (increased water solubility)
- azo bond protection via complexation to a transition metal complex
We directed substances that are metabolised to amines of concern and are considered bioavailable for further evaluation.
We will consider substances with unknown components or no verified SMILES representation for further evaluation.
We pre-screened substances for existing cosmetic use(s) and human health hazard flags (existing hazard classification). If a substance had cosmetic use(s) we considered it to present greater exposure and, therefore, potential for human health risks. We will consider substances with existing hazard classification(s) for a further evaluation. For azo-based substances that had known cosmetic use(s) (and salts of these azo-substances), we conducted further, more detailed evaluations.
We considered a more detailed screening approach for azo-based substances in combination with other components, such as the cations associated with anionic dyes.
For azo-based substances without existing human health hazard concerns or known cosmetic use(s) we evaluated their potential to release aromatic amines of concern (skin sensitisation, genotoxicity, or carcinogenicity).
Threshold of toxicological concern (TTC)
Considering the significant use of these chemicals as textile colourants, we considered the main source of consumer exposure to be from direct contact of the skin with the textiles coloured with azo-based substances. We will separately evaluate the exposure and potential risk from azo-based substances with reported use in cosmetics. It was not part of this evaluation (see above Pre-screening).
We evaluated the applicability of the classical TTC approach for these azo-based substances. Using TTC approaches (classical and based on genotoxicity alerts), we evaluated the risk of systemic toxicity following consumer exposure via the skin contact with everyday textile products coloured with azo-based substances.
We based the exposure estimate on the default model that the German Federal Institute for Risk Assessment’s “Textiles and Leather” Working Group developed.
Based on the TTC approaches, there is no unreasonable risk for systemic toxicity (other than carcinogenicity) from exposure via the skin contact with everyday textile products coloured with these azo-based substances.
Aromatic amines with skin sensitisation potential
We used the dermal sensitisation threshold (DST) and dermal sensitisation quantitative risk assessment (QRA) approaches to determine the risk for skin sensitisation induction arising from the aromatic amine metabolites of azo dyes used in textiles.
We based the exposure estimate for the aromatic amines with skin sensitisation concern using:
- the default model developed by the “Textiles and Leather” Working Group established at the German Federal Institute for Risk Assessment and
- the median and 95th -percentile consumer exposure level estimated based on the reported levels of sensitising aromatic amines in over 200 textile products (national and international)
There is negligible risk of sensitisation induction resulting from exposure to these sensitising aromatic amines through textiles dyed with azo dyes. The azo-based substances that have no other health hazard indications, other than producing aromatic amine metabolites with sensitisation concern, are of low concern for human health and low priority for further evaluation. However, we acknowledge that the elicitation of sensitisation may still occur in already sensitised individuals from this route of exposure.
Aromatic amines with genotoxicity and carcinogenicity potential
We based the genotoxicity and carcinogenicity concern of the aromatic amines that are:
- on the European Union (EU) 22 carcinogenic amines regulated in textile (EU22 amines)
- uncommon benzidine derivatives
- potentially genotoxic/carcinogenic identified from other sources
EU22 aromatic amines
We separately evaluated azo-based substances that metabolised into EU22 aromatic amines (NICNAS Tier II assessments):
- Selected benzidine-congener-based dyes
- Selected benzidine-congener-based pigments
- Dyes that could release selected carcinogenic amines (listed on the Inventory)
- Dyes that could release selected carcinogenic amines (not listed on the Inventory)
Uncommon benzidine derivatives
We separately evaluated azo-based substances that metabolised into uncommon benzidine derivatives:
Other aromatic amines of potential concern
Based on the TTC approach above, the estimated exposure may pose the risk for the azo-based substances with potential to be metabolised into genotoxic aromatic amines. Other aromatic amines of potential concern were identified from:
- carcinogenicity classifications from the International Agency for Research on Cancer
- substances classified for genotoxicity and/or carcinogenicity in the Hazardous Chemical Information System
- Bruschweiler and co-workers (Bruschweiler et al, 2014; Bruschweiler & Merlot, 2017)
- IMAP assessment report outcomes
We separately evaluated the potential concerns associated with azo-based substances that metabolised into these aromatic amines. We based our evaluation on:
- weight of evidence analysis from available data
- presenting mechanisms action for genotoxicity and carcinogenicity
- identifying functional groups (and their structural relationship) contributing to or cancelling out the concerns
- structure-activity relationship (SAR) profiling to support read-across and fill in data gaps
We identified SAR profiles linked with mechanisms of genotoxicity and/or carcinogenicity. Based in these, azo-based substances that potentially metabolise to aromatic amines with corresponding genotoxic and/or carcinogenic SAR profiles will be prioritised for further evaluation. Azo-based substances that potentially metabolise to aromatic amines with negative genotoxicity and/or carcinogenicity activity are of low concern for human health and low priority for further evaluation.
Last update 12 December 2019