Screening Assessment - Part 1

Aromatic Azo and Benzidine-based Substance Grouping
Certain Benzidine-based Dyes and Related Substances

Environment Canada
Health Canada
November 2014

Table of Contents

List of Tables

1. Introduction

Pursuant to sections 68 or 74 of the Canadian Environmental Protection Act, 1999 (CEPA 1999) (Canada 1999), the Minister of the Environment and the Minister of Health conduct screening assessments of substances to determine whether these substances present or may present a risk to the environment or to human health.

The Substance Groupings Initiative is a key element of the Government of Canada’s Chemicals Management Plan (CMP). The Aromatic Azo and Benzidine-based Substance Grouping consists of 358 substances that were identified as priorities for assessment, as they met the categorization criteria under section 73 of CEPA 1999 and/or were considered as a priority based on human health concerns (Environment Canada and Health Canada 2007). Some substances within this Substance Grouping have been identified by other jurisdictions as a concern due to the potential cleavage of the azo bonds, which can lead to the release of aromatic amines that are known or likely to be carcinogenic.

While many of these substances have common structural features and similar functional uses as dyes or pigments in multiple sectors, significant diversity within the substance group has been taken into account through the establishment of subgroups. Subgrouping based on structural similarities, physical and chemical properties, and common functional uses and applications accounts for variability within this Substance Grouping and allows for subgroup-specific approaches in the conduct of screening assessments. This Screening Assessment considers substances that belong to the Benzidine-based Acid Dyes, Benzidine-based Direct Dyes, Benzidine-based Cationic Indicators, and Benzidine-based Precursors, and Benzidine Derivatives subgroups. Consideration of azo bond cleavage products (aromatic amines) is a key element of human health assessment in each subgroup. Some aromatic amines, commonly referred to as EU22 aromatic aminesFootnote[2], as well as associated azo dyes are restricted in other countries (EU 2006). Information on the subgrouping approach for the Aromatic Azo and Benzidine-based Substance Grouping under Canada’s Chemical Management Plan (CMP), as well as additional background information and regulatory context, is provided in a separate document prepared by the Government of Canada (Environment Canada and Health Canada 2013).

Ten Benzidine-based Acid Dyes and 25 Benzidine-based Direct Dyes originally constituted 2 subgroups of the Aromatic Azo and Benzidine-based Substance Grouping. One Benzidine-based Acid Dye (Acid Red 111) and one Benzidine-based Direct Dye (Direct Black 38) were previously assessed during the Challenge Initiative of the CMP. It was concluded that Acid Red 111 and Direct Black 38 did not meet the criteria under section 64 of CEPA 1999 (Environment Canada, Health Canada 2009, 2011). Similarly, two of the Benzidine-based Acid Dyes (NAAHDand Acid Red 99), one of the Benzidine-based Direct Dyes (Direct Violet 28), and one of the Benzidine-based Cationic Indicators (TDBD) were previously included as part of a screening assessment, in April 2008, of 145 persistent, bioaccumulative, and inherently toxic (PBiT) substances that were considered not to be in commerce. No significant new information has been identified for Acid Red 111 or Direct Black 38, and therefore these substances are not included in the current Screening Assessment. However, Acid Red 111 and Direct Black 38 are used in this report for read-across purposes due to their structural similarity to the other Benzidine-based Acid Dyes and Benzidine-based Direct Dyes in these subgroups.  In contrast, NAAHD, Acid Red 99, Direct Violet 28, and TDBD are included in the Benzidine-based Substances considered in this Screening Assessment because significant new information has been identified.

Information on Benzidine and 3,3′-dichlorobenzidine (3,3′-DCB) was used to inform this Screening Assessment. These two benzidine derivatives were previously assessed under the Priority Substances List (PSL) program and are listed on Schedule 1 (List of Toxic Substances) under CEPA 1999 (Canada 1993a, 1993b). Benzidine is regulated under the Prohibition of Certain Toxic Substances Regulation, 2012 (Canada 2012).

Screening assessments focus on information critical to determining whether substances meet the criteria as set out in section 64 of CEPA 1999, by examining scientific information to develop conclusions by incorporating a weight of evidence approach and precaution.Footnote[3]

This Screening Assessment includes consideration of information on chemical properties, environmental fate, hazards, uses and exposure, including additional information submitted by stakeholders. Relevant data were identified up to May 2014. Empirical data from key studies as well as some results from models were used to reach conclusions. When available and relevant, information presented in assessments from other jurisdictions was considered.

The Screening Assessment does not represent an exhaustive or critical review of all available data. Rather, it presents the most critical studies and lines of evidence pertinent to the conclusion.

The Screening Assessment was prepared by staff in the Existing Substances Programs at Health Canada and Environment Canada and incorporates input from other programs within these departments. The ecological and human health portions of this assessment have undergone external written peer review and consultation. Comments on the technical portions relevant to the environment were received from Dr. Harold Freeman (North Carolina State University, USA) and Dr. Gisela Umbuzeiro (University of Campinas, Brazil). Comments on the technical portions relevant to human health were received from Dr. Harold Freeman (North Carolina State University, USA), Dr. David Josephy (University of Guelph, Canada), Dr. Michael Bird (University of Ottawa, Canada) and Dr. Kannan Krishnan (University of Montreal, Canada). Additionally, the draft of this Screening Assessment was subject to a 60-day public comment period.  While external comments were taken into consideration, the final content and outcome of the Screening Assessment remain the responsibility of Health Canada and Environment Canada.

The critical information and considerations upon which the Screening Assessment is based are given below.

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2. Identity of Substances

This Screening Assessment focuses on 42 substances that belong to 5 subgroups that are part of the Aromatic Azo and Benzidine-based Substance Grouping. These subgroups are based on structural similarity and similar applications: Benzidine-based Acid Dyes (9 substances), Benzidine-based Direct Dyes (24 substances), Benzidine-based Cationic Indicators (2 substances), Benzidine-based Precursors (2 substances) and Benzidine Derivatives (5 substances) (Environment Canada and Health Canada. 2013). Two substances, Acid Red 111 and Direct Black 38, which were previously assessed in the Challenge Initiative, are included to inform the assessment.

For the purpose of this Screening Assessment, four subgroups (Benzidine-based Acid Dyes, Direct Dyes, Cationic Indicators and Precursors) are collectively referred to as “Benzidine-based Substances” while Benzidine-based Acid Dyes and Benzidine-based Direct Dyes are collectively referred to as “Benzidine-based Dyes.”

The identities of the individual substances in this Screening Assessment are presented in Tables 2-1 to 2-5. The Chemical Abstracts Service Registry Numbers (CAS RNs), Domestic Substances List (DSL) names, Colour Index (C.I.) generic names, and chemical acronyms of these substances are presented in Tables 2-1 to 2-5. Chemical acronyms are derived from the C.I. generic names when available; otherwise, they are based on the DSL names. A list of additional chemical names (e.g., trade names) is available from the National Chemical Inventories (NCI 2007).

Table 2-1. Identity of the Benzidine-based Acid Dyes
CAS RNDSL nameC.I. generic nameChemical acronym
3701-40-42,7-Naphthalenedisulfonic acid, 4-hydroxy-3-[[4′-[(2-hydroxy-1-naphthalenyl)azo]-2,2′-dimethyl[1,1′-biphenyl]-4-yl]azo]Acid Red 99N/A
6459-94-51,3-Naphthalenedisulfonic acid, 8-[[3,3′-dimethyl-4′-[[4-[[(4-methylphenyl)sulfonyl]oxy]phenyl]azo][1,1′-biphenyl]-4-yl]azo]-7-hydroxy-, disodium saltAcid Red 114N/A
6470-20-8[1,1′-Biphenyl]-2,2′-disulfonic acid, 4-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-4′-[(2-hydroxy-1-naphthalenyl)azo]-, disodium saltAcid Orange 56N/A
6548-30-71,3-Naphthalenedisulfonic acid, 8-[[3,3′-dimethoxy-4′-[[4-[[(4-methylphenyl)sulfonyl]oxy]phenyl]azo][1,1′-biphenyl]-4-yl]azo]-7-hydroxy-, disodium saltAcid Red 128N/A
10169-02-5[1,1′-Biphenyl]-2,2′-disulfonic acid, 4,4′-bis[(2-hydroxy-1-naphthalenyl)azo]-, disodium saltAcid Red 97N/A
68318-35-42,7-Naphthalenedisulfonic acid, 4-amino-3-[[4′-[(2,4-dihydroxyphenyl)azo]-3,3′-dimethyl[1,1′-biphenyl]-4-yl]azo]-5-hydroxy-6-[(4-sulfophenyl)azo]-, trisodium saltAcid Black 209N/A
68400-36-22,7-Naphthalenedisulfonic acid, 4-amino-5-hydroxy-6-[[4′-[(4-hydroxyphenyl)azo]-3,3′-dimethyl[1,1′-biphenyl]-4-yl]azo]-3-[(4-nitrophenyl)azo]-, disodium saltNANAAHD
83221-63-02,7-Naphthalenedisulfonic acid, 4-amino-3-[[4′-[(2,4-diaminophenyl)azo]-2,2′-disulfo[1,1′-biphenyl]-4-yl]azo]-5-hydroxy-6-(phenylazo)-, sodium saltNANAADD
89923-60-4Benzenesulfonic acid, 3,3′-[(2,2′-dimethyl[1,1′-biphenyl]-4,4′-diyl)bis[azo(4,5-dihydro-3-methyl-5-oxo-1H-pyrazole-4,1-NABADB

Abbreviations:
N/A, not applicable (chemical acronym);
NA, not available (C.I.generic name)

Table 2-2. Identity of the Benzidine-based Direct Dyes
CAS RNDSL nameC.I. generic nameChemical acronym
72-57-12,7-Naphthalenedisulfonic acid, 3,3′-[(3,3′-dimethyl[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[5-amino-4-hydroxy-, tetrasodium saltDirect Blue 14N/A
573-58-01-Naphthalenesulfonic acid, 3,3′-[[1,1′-biphenyl]-4,4′-diylbis(azo)]bis[4-amino-, disodium saltDirect Red 28N/A
992-59-61-Naphthalenesulfonic acid, 3,3′-[(3,3′-dimethyl[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[4-amino-, disodium saltDirect Red 2N/A
2150-54-12,7-Naphthalenedisulfonic acid, 3,3′-[(3,3′-dimethyl[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[4,5-dihydroxy-, tetrasodium saltDirect Blue 25N/A
2429-71-21-Naphthalenesulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[4-hydroxy-, disodium saltDirect Blue 8N/A
2429-74-52,7-Naphthalenedisulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[5-amino-4-hydroxy-, tetrasodium saltDirect Blue 15N/A
6420-06-01-Naphthalenesulfonic acid, 4-hydroxy-3-[[4′-[(1-hydroxy-5-sulfo-2-naphthalenyl)azo]-3,3′-dimethyl[1,1′-biphenyl]-4-yl]azo]-, disodium saltDirect Violet 28N/A
6420-22-02,7-Naphthalenedisulfonic acid, 5-amino-3-[[4′-[(6-amino-1-hydroxy-3-sulfo-2-naphthalenyl)azo]-3,3′-dimethyl[1,1′-biphenyl]-4-yl]azo]-4-hydroxy-, trisodium saltDirect Blue 295N/A
6449-35-01-Naphthalenesulfonic acid, 3-[[4′-[(6-amino-1-hydroxy-3-sulfo-2-naphthalenyl)azo]-3,3′-dimethoxy[1,1′-biphenyl]-4-yl]azo]-4-hydroxy-, disodium saltDirect Blue 151N/A
6548-29-42,7-Naphthalenedisulfonic acid, 4,4′-[(3,3′-dichloro[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[3-amino-, tetrasodium saltDirect Red 46N/A
6655-95-4Acetic acid, 2,2′-[[4,4′-bis[[1-hydroxy-6-[(4-methoxyphenyl)amino]-3-sulfo-2-naphthalenyl]azo][1,1′-biphenyl]-3,3′-diyl]bis(oxy)]bis-, tetrasodium saltDirect Blue 158N/A
16071-86-6Cuprate(2−), [5-[[4′-[[2,6-dihydroxy-3-[(2-hydroxy-5-sulfophenyl)azo]phenyl]azo][1,1′-biphenyl]-4-yl]azo]-2-hydroxybenzoato(4-)]-, disodiumDirect Brown 95N/A
67923-89-12,7-Naphthalenedisulfonic acid, 5-amino-4-hydroxy-3-[[4′-[(1-hydroxy-4-sulfo-2-naphthalenyl)azo]-3,3′-dimethoxy[1,1′-biphenyl]-4-yl]azo]-, trilithium saltNANAAH·3Li
70210-28-5Benzoic acid, 5-[[4′-[[6-amino-5-(1H-benzotriazol-5-ylazo)-1-hydroxy-3-sulfo-2-naphthalenyl]azo]-3,3′-dimethoxy[1,1′-biphenyl]-4-yl]azo]-2-hydroxy-4-methyl-, disodium saltNABABHS
71215-83-3Benzoic acid, 5-[[4′-[(2-amino-8-hydroxy-6-sulfo-1-naphthalenyl)azo]-2,2′-dichloro[1,1′-biphenyl]-4-yl]azo]-2-hydroxy-, disodium saltNABAHSD
71550-22-62,7-Naphthalenedisulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[5-amino-4-hydroxy-, tetralithium saltNANADB·4Li
72252-59-6[1,1′-Biphenyl]-3,3′-dicarboxylic acid, 4-[[5-[[5-(aminosulfonyl)-2-hydroxyphenyl]azo]-1-hydroxy-6-(phenylamino)-3-sulfo-2-naphthalenyl]azo]-4′-[[1-[[(3-carboxy-4-hydroxyphenyl)amino]carbonyl]-2-oxopropyl]azo]-, tetrasodium saltNABDAAH
75659-72-22,7-Naphthalenedisulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[5-amino-4-hydroxy-, monolithium trisodium saltNANADB·Li·3Na
75659-73-32,7-Naphthalenedisulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[5-amino-4-hydroxy-, dilithium disodium saltNANADB·2Li·2Na
75673-18-62,7-Naphthalenedisulfonic acid, 5-amino-4-hydroxy-3-[[4′-[(1-hydroxy-4-sulfo-2-naphthalenyl)azo]-3,3′-dimethoxy[1,1′-biphenyl]-4-yl]azo]-, monolithium disodium saltNANAAH·Li·2Na
75673-19-72,7-Naphthalenedisulfonic acid, 5-amino-4-hydroxy-3-[[4′-[(1-hydroxy-4-sulfo-2-naphthalenyl)azo]-3,3′-dimethoxy[1,1′-biphenyl]-4-yl]azo]-, dilithium monosodium saltNANAAH·2Li·Na
75673-34-61-Naphthalenesulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[4-hydroxy-, dilithium saltNANADB·2Li
75673-35-71-Naphthalenesulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[4-hydroxy-, monolithium monosodium saltNANADB·Li·Na
75752-17-92,7-Naphthalenedisulfonic acid, 3,3′-[(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis(azo)]bis[5-amino-4-hydroxy-, trilithium monosodium saltNANADB·3Li·Na

Abbreviations:
N/A, not applicable;
NA, not available (C.I.generic name)

Table 2-3. Identity of the Benzidine-based Cationic Indicators
CAS RNDSL nameC.I. generic nameChemical acronym
298-83-92H-Tetrazolium, 3,3′-(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis[2-(4-nitrophenyl)-5-phenyl-, dichlorideN/ATDBPD
1871-22-32H-Tetrazolium, 3,3′-(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis[2,5-diphenyl-, dichlorideN/ATDBD

Abbreviations:
N/A, not applicable;
NA, not available (C.I.generic name)

Table 2-4. Identity of the Benzidine-based Precursors
CAS RNDSL nameC.I. generic nameChemical acronym
91-92-9Naphthalenecarboxamide, N,N′-(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis[3-hydroxy-N/ANaphthol AS-BR
93940-21-71-Triazene-1-carbonitrile, 3,3′-(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)bis-N/ATCDB

Abbreviations:
N/A, not applicable;
NA, not available (C.I.generic name)

Table 2-5. Identity of the Benzidine Derivatives
CAS RNDSL nameC.I. generic nameChemical acronym
91-97-41,1′-Biphenyl, 4,4′-diisocyanato-3,3′-dimethyl-N/ATODI
119-90-41,1′-Biphenyl]-4,4′-diamine, 3,3′-dimethoxy-N/A3,3′-DMOB
119-93-71,1′-Biphenyl]-4,4′-diamine, 3,3′-dimethyl-N/A3,3′-DMB
366-29-01,1′-Biphenyl]-4,4′-diamine, N,N,N′,N′-tetramethyl-N/A4N-TMB
612-82-81,1′-Biphenyl]-4,4′-diamine, 3,3′-dimethyl-, dihydrochlorideN/A3,3′-DMB·2HCl

Abbreviations:
N/A, not applicable;
NA, not available (C.I.generic name)

Example chemical structures, molecular formulas, group descriptions and molecular weight ranges are presented in Table 2-6 for the subgroups of Benzidine-based Acid Dyes, Benzidine-based Direct Dyes and Benzidine-based Cationic Indicators. Due to overall chemical similarity, including common functional groups (as described in Table 2-6), sharing similar physical and chemical properties and lack of data for individual substances, data for these three application classes (for which groupings are based on the method and conditions of their use) will be used for ecological exposure estimation approaches (when warranted). This general approach is discussed further in the section below on selection of analogues for read-across. Despite their similarities, these substances may have different potential transformation products. This is discussed further in the Environmental Persistence section and in the Human Health Effects section. Individual chemical structures, molecular formulas and molecular weights are presented for all Benzidine-based Dyes and Benzidine-based Cationic Indicators in Appendix A, Tables A1 – A3.

Table 2-6. Example structures and descriptions for Benzidine-based Dyes and Benzidine-based Cationic Indicators
SubgroupExample structure for the subgroupGroup description with critical functional groupsMolecular weight
range (g/mol)
Benzidine-based Acid Dyes (n = 10)Chemical structure of Acid Red 111
C37H30N4O10S3Na2(Acid Red 111)
Benzidine fragment, 1–2 naphthalene groups, 2–3 azo groups, 2–4 sulfonic acid groups, 3–4 benzene rings699–920
Benzidine-based Direct Dyes (n = 25)Chemical structure of Direct Blue 14
C34H24N6O14S4Na4(Direct Blue 14)
Benzidine fragment, 0–2 naphthalene groups, 2–3 azo groups, 1–4 sulfonic acid groups, 2–5 benzene rings, 0–3 amino groups, 0–1 benzotriazol groups696–1134
Benzidine-based Cationic Indicators (= 2)Chemical structure of TDBD
C40H30N8O2 (TDBD)
Benzidine fragment, 2 tetrazoliums, 6 benzene rings655–818


Specific identities are presented for each of the Benzidine-based Precursors (Table 2-7) and Benzidine Derivatives (Table 2-8), given their more diverse chemical structure, physical and chemical properties.

Table 2-7. Structures and descriptions for specific Benzidine-based Precursors
Chemical nameChemical structureSubstance description with critical functional groupsMolecular weight (g/mol)
TCDBChemical structure of TCDB
C16H14N8O2
Benzidine fragment, 2 azo groups, 2 nitrile groups351
Naphthol AS-BRChemical structure of Naphthol AS-BR
C36H28N2O6
Benzidine fragment, 2 naphthol groups585
Table 2-8. Structures and descriptions for specific Benzidine Derivatives
Chemical nameChemical structureSubstance description with critical functional groupsMolecular weight (g/mol)
3,3′-DMBChemical structure of 3,3'-DMB
C14H16N2
Benzidine fragment212
3,3′-DMB·2HClChemical structure of 3,3'DMB 2HC1
C14H184CL2N2
Benzidine fragment, 2 chlorine ions285
3,3′-DMOBChemical structure of 3,3'-DMOB
C14H16N2O2
Benzidine fragment244
TODIChemical structure of TODI
C16H12N2O2
Biphenyl fragment, 2 isocyanate moieties264
4N-TMBChemical structure of 4N-TMB
C16H20N2
Benzidine fragment, 2 tertiary amines240


In general, Benzidine-based Direct Dyes are of higher molecular mass than the Benzidine-based Acid Dyes and Benzidine-based Cationic Indicators, while Benzidine Derivatives and the two Benzidine-based Precursors have much lower molecular masses, since most can be used to synthesize dyes or pigments. Acid and direct dyes may each contain one or more azo, naphthalene and sulfonic acid groups and have 2–5 benzene rings each. The Benzidine-based Direct Dyes in this report may also contain amino groups that the Benzidine-based Acid Dyes do not. The two Benzidine-based Cationic Indicators each contain two tetrazolium moieties.

2.1 Selection of Analogues and Use of (Q)SAR Models

Guidance on the use of read-across approaches has been prepared by various organizations, such as the Organisation for Economic Co-operation and Development (OECD). It has been applied in various regulatory programs, including the European Union’s (EU) Existing Substances Programme. The general method for analogue selection and the use of (quantitative) structure-activity relationship ((Q)SAR) models is provided in Environment Canada and Health Canada (2013). For characterization of human health effects, the basis for the use of analogues and/or (Q)SAR modelling data is documented in the Human Health Effects Assessment section of this report.

Analogues used to inform the ecological assessment were selected based on the availability of relevant empirical data pertaining to physical and chemical properties and aquatic ecotoxicity data for the Benzidine-based Dyes as well as the two Benzidine-based Cationic Indicators were identified from within each respective group of substances contained in this Screening Assessment. A few additional analogue data points from non-benzidine-based acid dyes (Acid Yellow 23, Acid Yellow 36, Acid Orange 7; Table 2-9) were also considered for the Benzidine-based Acid Dyes to contribute to the weight of evidence. Analogue data for the Benzidine-based Cationic Indicators were also identified from outside the Benzidine-based Cationic Indicators application class. Basic Brown 4, an azo-based cationic dye (Table 2-9), and a tetrazolium substance (2,3,5-triphenyltetrazolium chloride, or TTC; Table 2-9) provide read-across data for ecotoxicity. The US Environmental Protection Agency (US EPA), in its chemical categories document, notes that no relationship between structure and activity has been found for cationic dyes (US EPA 2010), which are similar to the Benzidine-based Cationic Indicators. That being said, some categories of basic dyes (e.g., triarylmethanes) have been found to share certain characteristics, such as high toxicity. Given a lack of available empirical information and an acceptable level of similarity based on physical and chemical properties as well as the environmental fate of soluble Benzidine-based Acid and Direct Dyes and Benzidine-based Cationic Indicators, bioaccumulation and soil toxicity data are shared across these three subgroups.

Analogue data for one of the Benzidine-based Precursors (Naphthol AS-BR) were also identified from a substance outside the application classes identified in this assessment (3-hydroxy-N-phenyl-2-naphthalenecarboxamide, or NHNP; Table 2-9).

Acceptable analogue candidates were identified within the Benzidine Derivatives group for four of the five substances. The substance 3,3′-DMB will be used as an analogue for 3,3′-DMB·2HCl and TODI for physical and chemical properties, persistence, bioaccumulation and ecotoxicity. 3,3′-DMB·2HCl is in a crystalline form that behaves as a salt and will release 3,3′-DMB when in solution. TODI rapidly hydrolyzes in water to its associated amine, 3,3′-DMB, also referred to as TODA. 3,3′-DMB will also be used as an analogue for 3,3′-DMOB to evaluate its bioaccumulation potential and ecotoxicity, since no empirical data are available. These substances are similar, with the exception of methoxy groups in the ortho position to the amines for 3,3′-DMOB instead of methyl groups for 3,3′-DMB. Therefore, it is expected that the substances will have similar bioavailability and mode of action.

Benzidine (CAS RN 92-87-5; Table 2-9) was selected as a suitable analogue for 4N-TMB to correct some physical and chemical property estimates. 4N-TMB is structurally similar to benzidine, with the difference that it is a tertiary amine, containing two methyl substituents on each nitrogen atom, thereby missing the amine component (–NH–) that could undergo N-hydroxylation.

Analogue data are very important to this Screening Assessment, since the physical and chemical properties of many dyes are not amenable to model prediction, as they are considered to be out of the model domain of applicability (e.g., structural and/or parametric domains).

Therefore, the applicability of (Q)SAR models to dyes is determined on a case-by-case basis. In some cases, empirical analogue data will be adjusted to account for structural differences between the substance being assessed and the analogue using the Experimental Value Adjustment (EVA) method in EPISuite version 4.1 (2012).

Table 2-9. Identity of analogues and parameters to be used to inform the physical and chemical properties, environmental fate and potential to cause ecological harm
Name (acronym)Chemical structure and formula (CAS RN)Molecular weight (g/mol)Parameters to be used in reportDescription
Acid Yellow 23Chemical structure of Acid Yellow 23
C16H8N4O9S2Na4(1934-21-0)
556.34Kow
Read-across for Benzidine-based Acid Dyes
AY23 is a monoazo acid dye. It differs in that it lacks the benzidine moiety and has a heterocyclic nitrogen moiety.
Acid Orange 7Chemical structure of Acid Orange 7
C16H11N2O4SNa (633-96-5)
350.33Kow
Read-across for Benzidine-based Acid Dyes
AO7 is a monoazo acid dye. It differs in that it has a lower molecular mass and lacks the benzidine moiety.
Acid Yellow 36Chemical structure of Acid Yellow 36
C18H14N3O3SNa (587-98-4)
375.38Kow
Read-across for Benzidine-based Acid Dyes
AY36 is a monoazo acid dye. It differs in that it has a lower molecular mass and lacks the benzidine moiety.
Basic Brown 4Chemical structure of Basic Brown 4
C21H26Cl2N8(5421-66-9)
461Ecotoxicity
Read-across for Benzidine-based Cationic Indicators
BB4 is a water-soluble cationic dye with two chloride ions. It differs in that it is a meta-phenylenediamine instead of benzidine based and has a lower molecular mass.
2,3,5-Triphenyltetrazolium chloride (TTC)Chemical structure of TTC
C19H15ClN4 (298-96-4)
335Ecotoxicity
Read-across for Benzidine-based Cationic Indicators
TTC contains the tetrazolium chloride moiety, which is almost exactly one-half of the structure of the two Benzidine-based Cationic Indicators.
2-Naphthalenecarboxamide, 3-hydroxy-N-phenyl- (NHNP)Chemical structure of NHNP
C36H28N2O6(92-77-3)
263Melting point
Read-across for Naphthol AS-BR (Benzidine-based Precursor)
This is about one half of the structure of Naphthol AS-BR.
BenzidineChemical structure of Benzidine
C12H12N2 (92-87-5)
184Henry’s Law constant, water solubility and Kow
Read-across to correct estimates for five Benzidine Derivatives
Benzidine is the critical fragment for all Benzidine Derivatives.

Abbreviations:
Kow, octanol–water partition coefficient

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3. Physical and Chemical Properties

Physical and chemical properties determine the overall characteristics of a substance and are used to determine the suitability of different substances for different applications. Such properties also play a critical role in determining the environmental fate of substances (including their potential for long-range transport), as well as their toxicity to humans and non-human organisms.

A subset of physical and chemical properties of Benzidine-based Acid Dyes, Benzidine-based Direct Dyes and Benzidine-based Cationic Indicators--namely, melting point, water solubility and log octanol–water partition coefficient (log Kow)--is important in terms of ecological and human health assessment. A summary of the experimental physical and chemical properties of substances in the Benzidine-based Dyes and Benzidine-based Cationic Indicators subgroups that are relevant to their environmental fate and ecotoxicity is presented in Table 3-1. Pivotal values, including either single mean data points (e.g., melting point and decomposition) or a range of values, have been chosen to represent the properties of each subgroup. Detailed substance-specific information on these substances can be found in Tables A4–A6 in Appendix A of this report.

All Benzidine-based Dyes and Benzidine-based Cationic Indicators in this assessment are relatively large (molecular weights greater than  600 g/mol), complex ionic molecules (acid and direct dyes are anionic molecules and cationic indicators are cationic molecules), tend to dissociate at environmentally relevant pH levels and are highly soluble in water (in excess of 1 g/L) due to the presence of solubilizing functional groups (Hunger 2003; Table 3-1). Benzidine-based Acid Dyes are sodium salts that contain solubilizing sulfonic acid groups, while Benzidine-based Cationic Indicators contain tetrazolium functional groups with solubilizing positive charges. Given their hydrophilicity and ionic character, as demonstrated by low pKa values (an indicator of acid dissociation), Benzidine-based Acid and Direct Dyes tend to have very low experimental log Kow values (Table 3-1).

While no quantitative data are available for the two Benzidine-based Cationic Indicators, they are basic substances that are known to dissociate in water and have low log Kowvalues (Øllgaard et al. 1998).

While experimental data are limited, all Benzidine-based Dyes and Benzidine-based Cationic Indicators are expected to have very low vapour pressures and very low Henry’s Law constant values (Øllgaard et al. 1998).

Table 3-1. A summary of experimental physical and chemical properties (at standard temperature of approximately 25°C) for the Benzidine-based Acid Dyes, Benzidine-based Direct Dyes and Benzidine-based Cationic Indicators
SubsetPropertyValue(s) or range (for more than 3 data points)Pivotal value(s) used in this  assessment (basis for selection)
Benzidine-based Acid DyesMelting point and decomposition (°C)170; 185; 190182 (mean)
Benzidine-based Acid DyesWater solubility (mg/L)greater than 500–65 000 (n = 4)greater than 500–65 000 (range is used)
Benzidine-based Acid DyesLog Kow (dimensionless)−0.017; 0.57; 0.70.42 (mean)
Benzidine-based Acid DyesEffective minimum cross-sectional diameter (Dmin, nm)0.96–1.15 (n = 10)Range used and discussed in text
Benzidine-based Acid DyesEffective maximum cross-sectional diameter (Dmax, nm)1.27–1.78 (n = 10)Range used and discussed in text
Benzidine-based Acid DyespKa (dimensionless)pKa1 = −0.67
pKa2 = −0.23
(sole pKa estimates)
Benzidine-based Direct DyesMelting point and decomposition (°C)109 – greater than 360 (n = 6)greater than 317 (mean)
Benzidine-based Direct DyesWater solubility (mg/L)1000–116 000 (n = 4)1000–116 000 (range is used)
Benzidine-based Direct DyesLog Kow (dimensionless)0.770.77 (sole value)
Benzidine-based Direct DyesEffective minimum cross-sectional diameter (Dmin, nm)0.95–1.27 (n = 25)Range used and discussed in text
Benzidine-based Direct DyesEffective maximum cross-sectional diameter (Dmax, nm)1.24–2.07 (n = 25)Range used and discussed in text
Benzidine-based Direct DyespKa (dimensionless)pKa1 = −1.5
pKa2 = −1.3
(sole pKa estimates)
Benzidine-based Cationic IndicatorsMelting point and decomposition (°C)189–255 (n = 4)209 (mean)
Benzidine-based Cationic IndicatorsWater solubility (mg/L)9000, 10 0009500 (mean)
Benzidine-based Cationic IndicatorsLog Kow (dimensionless)LowLow (no quantitative data)
Benzidine-based Cationic IndicatorsEffective minimum cross-sectional diameter (Dmin, nm)N/AN/A
Benzidine-based Cationic IndicatorsEffective maximum cross-sectional diameter (Dmax, nm)N/AN/A
Benzidine-based Cationic IndicatorspKa (dimensionless)pKa1 = not reliable
pKa2 = not reliable
NA

Abbreviations:
Ka, acid dissociation constant;
Kow, octanol–water partition coefficient;
N/A, not applicable;
NA, not available (C.I.generic name)

Data for each of the Benzidine-based Precursors and Benzidine Derivatives are shown individually in Tables 3-2 and 3-3, respectively, since there are important structural differences between these substances. More detailed data for each of these substances are also available in Appendix A, Tables A-7 and A-8. While physical and chemical properties also vary from one substance to another, generally, Benzidine Derivatives are water soluble, with low to moderate log Kow and log organic carbon–water partition coefficient (log Koc) values. The two Benzidine-based Precursors are less water soluble, with higher log Kow and log Koc values.

The only experimental physical and chemical property data available for the two Benzidine-based Precursors were for melting point; therefore, model estimates were used for the remaining properties for the neutral form of the substances (Table 3-2).

It is acceptable to use (Q)SAR models for Benzidine-based Precursors since they are relatively simple structures and fit in the model domains of applicability. Benzidine-based Precursors have high modelled log Kow and log Koc values and relatively low water solubility in comparison with the Benzidine Derivatives. However, there are differences between these two substances, as Naphthol AS-BR is practically water insoluble and does not ionize readily based on pKa, while TCDB is somewhat soluble and is predicted to ionize in water.

Table 3-2. Experimental and estimated physical and chemical properties (at standard temperature of approximately 25°C) for the Benzidine-based Precursors
ChemicalPropertyValue(s) or range (for more than 3 data points)Pivotal value used in this assessment (basis for selection)
Naphthol AS-BRMelting point (°C)246; 350290 (mean of estimated data)
Naphthol AS-BRBoiling point
(ºC)
927.49927.49 (sole estimated value)
Naphthol AS-BRVapour pressure
(Pa)
7.7 × 10−257.7 × 10−25 (sole estimated value)
Naphthol AS-BRHenry’s Law constant
(Pa·m3/mol)
1.96 × 10151.96 × 10−15 (sole estimated value)
Naphthol AS-BRLog Kow7.757.75 (sole estimated value)
Naphthol AS-BRLog Koc1.43 × 105; 8.21 × 1054.85 × 105 (mean of estimated values)
Naphthol AS-BRLog Koa25.8525.85 (sole estimated value)
Naphthol AS-BRWater solubility
(mg/L)
8.97 × 10−6; 1.44 × 10−51.15 × 10−6 (mean of estimated values)
Naphthol AS-BREffective minimum cross-sectional diameter (Dmin, nm)1.021.02 (sole estimated value)
Naphthol AS-BREffective maximum cross-sectional diameter (Dmax, nm)1.421.42 (sole estimated value)
Naphthol AS-BRpKa (dimensionless)pKa1 = 13.80
pKa2 = 13.10
pKa1 = 13.80
pKa2 =13.10
(sole estimated values)
TCDBMelting point (°C)250.21250.21 (sole estimated value)
TCDBBoiling point
(ºC)
580.51580.51 (sole estimated value)
TCDBVapour pressure
(Pa)
3.16 × 10−83.16 × 10−8 (sole estimated value)
TCDBHenry’s Law constant
(Pa·m3/mol)
5.81 × 10−95.81 × 10−9 (sole estimated value)
TCDBLog Kow5.135.13 (sole estimated value)
TCDBLog Koc2.2; 5.473.85 (mean of estimated values)
TCDBLog Koa16.7616.76 (sole estimated value)
TCDBWater solubility
(mg/L)
0.26; 32.8016.53 (mean of estimated values)
TCDBEffective minimum cross-sectional diameter (Dmin, nm)0.830.83 (sole estimated value)
TCDBEffective maximum cross-sectional diameter (Dmax, nm)1.051.05 (sole estimated value)
TCDBpKa (dimensionless)pKa1 = −3.9
pKa2 = −5.15
pKa1 = −3.9
pKa2 = −5.15
(sole estimated values)

Abbreviations:
Ka, acid dissociation constant;
Koa, octanol–air partition coefficient;
Koc, organic carbon–water partition coefficient;
Kow, octanol–water partition coefficient

The variance in the physical and chemical properties of the five Benzidine Derivatives is generally quite high due to small differences in their chemical structures (e.g., 3,3′-DMB·2HCl is a salt). Three substances, 3,3′-DMB, 3,3′-DMOB and 3,3′-DMB·2HCl, are primary aromatic amines that are not expected to become ionized under relevant physiological and environmental pH conditions, as indicated by their pKa values (Table 3.3). 3,3′-DMB is structurally identical to benzidine but contains two methyl (CH3−) substituents in the ortho position to the amino functional groups. 3,3′-DMB·2HCl is anticipated to behave as a salt and will produce 3,3′-DMB once released in solution. 3,3′-DMOB contains two methoxy functional groups (O–CH3–) in the ortho position to the two amino groups. TODI contains two isocyanate functional groups (–N=C=O) and two methyl groups in the ortho position to the isocyanate functional groups, which will hydrolyze readily when in contact with water (see Environmental Persistence section). 4N-TMB is a tertiary amine and lacks the N–H groups that undergo N-hydroxylation.

Any modelled values shown in Table 3-3 and Table A-8 in Appendix A are for the neutral forms of the five Benzidine Derivatives.

Using (Q)SAR models for these substances is acceptable, as the Benzidine Derivatives are simpler substances that fit in the model domains of applicability. It is noted that EPIWIN’s predictions for salts are uncertain; therefore, predictions generated for 3,3′-DMB·2HCl will not be considered in the assessment.

The modelled water solubilities, log Kow values and Henry’s Law constants for 3,3′-DMB, 3,3′-DMOB and 4N-TMB were determined using the EVA option in WATERNT, KOWWIN and HENRYWIN in EPISuite 4.1. The empirical value for the analogue is quantitatively adjusted based on structural fragment differences when the two chemicals are compared. In the present case, water solubility, log Kow value and Henry’s Law constant for the analogue substance benzidine were used to generate predictions for 4N-TMB or to correct predictions for 3,3′-DMB and 3,3′-DMOB.

Table 3-3. Experimental and estimated physical and chemical properties (at standard temperature of approximately 25°C) for the Benzidine Derivatives
ChemicalPropertyValue(s) or range (for more than 3 data points)Pivotal value used in this  assessment (basis for selection)
3,3′-DMBMelting point (°C)128–132 (n = 6)130 (mean of experimental data)
3,3′-DMBBoiling point (ºC)200–339 (n = 4)280 (mean of experimental data)
3,3′-DMBVapour pressure (Pa)9.226 × 10−5; 2.74 × 10−29.226 × 10−5 (sole experimental value)
3,3′-DMB
3,3′-DMB
Henry’s Law constant
(Pa·m3/mol)
6.373 × 10−7 – 2.59 × 10−2(n = 4)2.59 × 10−2 (sole EVA method value)
3,3′-DMBLog Kow2.3–3.0 (n = 4)2.4 (mean of experimental and EVA values)
3,3′-DMBLog Koc2.17–3.502.8 (mean of estimated values)
3,3′-DMBLog Koa10.9310.93 (sole estimated value)
3,3′-DMBWater solubility (mg/L)27–1300 (n = 5)51 (mean of MITI data and EVA values)
3,3′-DMBEffective minimum cross-sectional diameter (Dmin, nm)0.790.79 (sole estimated value)
3,3′-DMBEffective maximum cross-sectional diameter (Dmax, nm)0.860.86 (sole estimated value)
3,3′-DMBpKa (dimensionless)pKa1 = 4.6, 4.5, 3.3
pKa2 = 3.4–3.5
pKa1 = 4.13
pKa2 = 3.45
(mean of experimental data)
3,3′-DMB·2HClMelting point (°C)210; 340275 (mean of experimental data)
3,3′-DMB·2HClBoiling point (ºC)NANA
3,3′-DMB·2HClVapour pressure (Pa)NANA
3,3′-DMB·2HClHenry’s Law constant
(Pa·m3/mol)
NANA
3,3′-DMB·2HClLog KowNANA
3,3′-DMB·2HClLog KocNANA
3,3′-DMB·2HClLog KoaNANA
3,3′-DMB·2HClWater solubility10 000; 50 00030 000 (mean of experimental data)
3,3′-DMB·2HClEffective minimum cross-sectional diameter (Dmin, nm)0.770.77 (sole estimated value)
3,3′-DMB·2HClEffective maximum cross-sectional diameter (Dmax, nm)0.770.77 (sole estimated value)
3,3′-DMB·2HClpKa (dimensionless)NANA
3,3′-DMOBMelting point (ºC)136; 137137 (mean of experimental data)
3,3′-DMOBBoiling point (ºC)356; 417.2356 (sole experimental value)
3,3′-DMOBVapour pressure (Pa)1.66 × 10−5 – 9.45 × 10−41.66 × 10−5 (sole experimental value)
3,3′-DMOBHenry’s Law constant
(Pa·m3/mol)
1.83 × 10−8 – 7.45 × 10−57.45 × 10−5 (EVA method)
3,3′-DMOBLog Kow1.5; 2.081.7 (mean of experimental and EVA values)
3,3′-DMOBLog Koc1.99; 2.712.4 (mean of estimated values)
3,3′-DMOBLog Koa13.2113.2 (sole estimated value)
3,3′-DMOBWater solubility60; 146.8103.4 (mean of experimental and EVA values)
3,3′-DMOBEffective minimum cross-sectional diameter (Dmin, nm)0.790.79 (sole estimated value)
3,3′-DMOBEffective maximum cross-sectional diameter (Dmax, nm)0.860.86 (sole estimated value)
3,3′-DMOBpKa (dimensionless)pKa1 = 4.7; 4.24.7 (sole experimental value)
TODIMelting point (°C)70–11671 (mean of experimental data)
TODIBoiling point (ºC)314–373353 (mean of experimental data)
TODIVapour pressure (Pa)2.95 × 10−32.95 × 10−3 (sole estimated value)
TODIHenry’s Law constant
(Pa·m3/mol)
NANA
TODILog KowNANA
TODILog KocNANA
TODILog Koa10.4710.5 (sole estimated value)
TODIWater solubilityNANA
TODIEffective minimum cross-sectional diameter (Dmin, nm)0.770.77 (sole estimated value)
TODIEffective maximum cross-sectional diameter (Dmax, nm)0.780.78 (sole estimated value)
TODIpKa (dimensionless)NANA
4N-TMBMelting point (°C)108.5–195 (n = 5)194 (mean of experimental data)
4N-TMBBoiling point (ºC)353.7353.7 (sole estimated value)
4N-TMBVapour pressure (Pa)2.41 × 10−4; 2.17 × 10−32.17 × 10−3 (sole experimental value)
4N-TMBHenry’s Law constant
(Pa·m3/mol)
1.06 × 10−2; 4.94 × 10−14.94 × 10−1 (EVA method)
4N-TMBLog Kow3.53; 4.113.53 (EVA method)
4N-TMBLog Koc2.75; 3.07; 3.173 (mean of estimated values)
4N-TMBLog Koa9.489.48 (sole estimated value)
4N-TMBWater solubility8.23–33.8 (n = 5)22.6 (mean of 4N-TMB experimental and EVA values)
4N-TMBEffective minimum cross-sectional diameter (Dmin, nm)0.650.65 (sole estimated value)
4N-TMBEffective maximum cross-sectional diameter (Dmax, nm)0.650.65 (sole estimated value)
4N-TMBpKa (dimensionless)pKa1 = 6.14
pKa2 = 4.07
pKa1 = 6.14
pKa2 = 4.07
(sole estimated values)

Abbreviations:
EVA, Experimental Value Adjustment;
Ka, acid dissociation constant;
Koa, octanol–air partition coefficient;
Koc, organic carbon–water partition coefficient;
Kow, octanol–water partition coefficient;
MITI, Ministry of International Trade and Industry (Japan);
NA, not available

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4. Sources and Uses

4.1 Sources

All Benzidine-based Dyes, Benzidine-based Cationic Indicators, Benzidine-based Precursors and Benzidine Derivatives are anthropogenically produced and not expected to occur naturally in the environment.

In recent years (2005 to present), all substances included in this Screening Assessment have been included in surveys issued pursuant to section 71 of CEPA 1999. Nine substances were included in a survey for the 2005 calendar year (Canada 2006b), 2 of these substances were also included in surveys conducted pursuant to section 71 for the 2006 calendar year under the Challenge Initiative (Canada 2006a, 2006b, 2007, 2008, 2009), 5 substances were included in Phase One of the DSL Inventory Update survey (Canada 2009) and 33 substances were included in a survey conducted pursuant to section 71 of CEPA 1999 for the 2010 calendar year that focused on the Aromatic Azo and Benzidine-based Substance Group (Canada 2011). Three substances were included in both surveys conducted for the 2005 and 2010 calendar years.

No substances were reported as being manufactured or imported into Canada above the reporting threshold of the surveys. Direct Blue 14 was however reported as being imported into Canada, in quantities below or equal to the reporting threshold in the DSL Inventory Update survey. Acid Red 97 had declarations of stakeholder interest. This is consistent with the trend by manufacturers to phase out the use of benzidine-based dyes in the mid- to late 1970s and replace them with other dyes due to potential human health concerns (IARC 2010a). Additionally, restrictions on uses of dyes based on benzidine are in place in many jurisdictions, including Europe (Environment Canada and Health Canada 2013). Two European surveys, as well as a Japanese study, indicate that 3,3′-DMB and 3,3′-DMOB are still sometimes detected in some textiles and leather products, some of which were reported to be imported from other countries. Accordingly, these two Benzidine Derivatives could be present in imported products in Canada, as the Canadian textile market is predominantly composed of imported products. Testing of products on the Canadian market, however, did not identify these two derivatives in imported and domestic textile and leather products (Health Canada 2013).

4.1.1 Benzidine-based Acid Dyes

Based on the information submitted in response to industry surveys conducted for the years 2005 and 2006 under Canada Gazette notices issued pursuant to section 71 of CEPA 1999, no manufacture or import activity in Canada above the reporting threshold was reported for any of the 9 Benzidine-based Acid Dyes (Canada 2006a, 2006b, 2008, 2009, 2011b).

Acid Red 97 had a stakeholder interest expressed in the section 71 survey for the reporting year of 2010 (Environment Canada 2012), but manufacture or import of the substance above the reporting threshold was not reported. Acid Red 97 was reported to be used or sold in Canada in quantities less than the reporting threshold of 100 kg (2010 email from Ecological and Toxicological Association of Dyes and Organic Pigments Association to Program Development and Engagement Division, Environment Canada; unreferenced).

4.1.2 Benzidine-based Direct Dyes

Information on Canadian manufacture and import of Direct Blue 14 was collected in Phase One of the DSL Inventory Update in 2009 under Canada Gazette notices issued pursuant to section 71 of CEPA 1999 (Canada 2006b, 2008b, 2011b).

Fewer than four companies reported using Direct Blue 14 in quantities between 0 and 100 kg/year. No information was submitted on other Benzidine-based Direct Dyes in recent regulatory surveys issued under CEPA 1999.

Only one Benzidine-based Direct Dye was found on any European use lists: Direct Blue 295 was identified as an LPV chemical in the EU (ESIS ©1995–2012). No information was found on recent use of Direct Blue 14 by the EU.

4.1.3 Benzidine-based Cationic Indicators

No information was submitted on cationic indicators in recent regulatory surveys issued under CEPA 1999. No information is available from Substances in Preparations in Nordic Countries (SPIN), the US Toxic Substances Control Act (TSCA) or the European Chemical Substances Information System (ESIS).

4.1.4 Benzidine-based Precursors

No information was submitted on the two Benzidine-based Precursors (Naphthol AS-BR and TCDB) in recent regulatory surveys issued under CEPA 1999. No information is available from SPIN, TSCA or ESIS.

4.1.5 Benzidine Derivatives

No information was submitted on Benzidine Derivatives in recent regulatory surveys issued under CEPA 1999, with the exception of one stakeholder that identified itself as having an interest in one substance (3,3′-DMOB) (Canada 2006b).

TODI, 3,3′-DMB and 3,3′-DMOB are considered LPV chemicals by the EU (ESIS ©1995–2012). TODI is also registered in the European Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) initiative (ECHA 2012).

The national aggregate production volumes for TODI in the United States were 4 540–227 000 kg (i.e., 10 000–500 000 pounds) and 227 000–454 000 kg (i.e., 500 000 – 1 000 000 pounds) in the 2002 and 2006 reporting cycles under the US EPA’s Inventory Update Reporting program (US EPA 2010, 2012b). 3,3′-DMB·2HCl was last reported in 1990 at a quantity of 4540–227 000 kg (i.e., 10 000 – 500 000 pounds) (US EPA 2012b).

4.2 Uses

Most Benzidine-based Substances have common structural features and similar functional uses as colourants, which are used in various sectors, predominantly textile and leather, as well as research and development (Environment Canada and Health Canada 2013).

No manufacture or import activities were reported in response to the section 71 survey for the calendar year 2010 (Environment Canada 2012) or other recent surveys (Environment Canada 2006b, 2008, 2009) above the reporting threshold, however it was reported that Direct Blue 14 was imported for use as a laboratory substance for research and development below or equal to the reporting threshold of 100 kg (Canada 2009; Environment Canada 2009). General uses, globally, reported in publicly available sources for each subgroup are summarized in Table 4-1 (Merck Index 2001; Kirk-Othmer 2010; Sigma-Aldrich Canada 2010; Ullmann’s Encyclopedia 2010; US EPA 2010).

Table 4-1. General uses of Benzidine-based Substances and Benzidine Derivatives
Benzidine-based Acid DyesBenzidine-based Direct DyesBenzidine-based Cationic IndicatorsBenzidine-based PrecursorsBenzidine Derivatives
Dyes, pigments and/or colouring agents
Electro-optical devices
Food contact substances / food packaging
Laboratory agents (e.g., stains)
Laser and liquid crystal display
Leather
Paints and coatings
Paper
Pharmaceuticals
Plastic materials
Printing inks
Textiles
Chemical intermediates
Dyes, pigments and/or colouring agents
Electro-optical devices
Food contact substances / food packaging
Laboratory agents
Laser and liquid crystal display
Leather
Paints and coatings
Paper
Pharmaceuticals
Plastic materials
Printing inks
Textiles
Laboratory agentsCosmetic Regulations

Intermediates in the manufacturing of:

  • Dyes, pigments and/or colouring agent
  • Electro-optical devices
  • Food contact substances / food packaging
  • Laboratory agents
  • Laser and liquid crystal display
  • Paints and coatings
  • Paper
  • Pharmaceuticals

Intermediates in the manufacturing of:

  • Adhesives, elastomers, flexible and rigid foam plastics
  • Dyes, pigments and/or colouring agents
  • Printing inks


Benzidine-based Dyes may be regulated as colorants in Canada by various measures, depending on their uses and applications, as explained further below.

None of the substances in this Screening Assessment are on the List of Permitted Colouring Agents as permitted food colourants or identified for use in food packaging materials in Canada (2011 emails from the Food Directorate, Health Canada, to the Risk Management Bureau, Health Canada; unreferenced; Health Canada 2012).

None of the substances in this Screening Assessment are listed as permitted drug colourants, listed for use as non-medicinal ingredients in human drugs in the Therapeutic Products Directorate’s internal Non-Medicinal Ingredients Database (2011 email from the Therapeutic Products Directorate, Health Canada, to the Risk Management Bureau, Health Canada; unreferenced) nor have they been identified to be present in veterinary drugs (2011 email from the Veterinary Drugs Directorate, Health Canada, to the Risk Management Bureau, Health Canada; unreferenced) or in biologics in Canada (2011 email from the Biologics and Genetic Therapies Directorate, Health Canada, to the Risk Management Bureau, Health Canada; unreferenced). Direct Blue 14 was the only substance identified in the Drug Products Database as an active ingredient in an ophthalmic solution for humans for use in eye surgery (DPD 2012). None of the substances in this Screening Assessment are listed in the Natural Health Products Ingredients Database (NHPID 2008) as substances in natural health products nor identified in the Licensed Natural Health Products Database (LNHPD 2008) to be present in currently licensed natural health products.

None of the substances in this Screening Assessment are present as ingredients in cosmetic products notified to Health Canada under the Cosmetic Regulations, Food and Drugs Act (R.S.C., 1985, c. F-27) (2011 emails from the Consumer Product Safety Directorate, Health Canada, to the Existing Substances Risk Assessment Bureau, Health Canada; unreferenced). None of these substances are included on Health Canada’s Cosmetic Ingredient HotlistFootnote[4]

No uses of Benzidine-based Substances or Benzidine Derivatives were identified for pest control products registered in Canada (2011 email from the Pest Management Regulatory Agency, Health Canada, to the Risk Management Bureau, Health Canada; unreferenced).

In addition, no uses of Benzidine-based Substances were identified for military applications in Canada (2011 email from the Department of National Defence to the Risk Management Bureau, Health Canada; unreferenced). Two Benzidine Derivatives, 3,3′-DMOB and 3,3′-DMB, have been identified to be used in military applications in Canada for gas detector kits (2011 email from the Department of National Defence to the Risk Management Bureau, Health Canada; unreferenced).

4.2.1 Benzidine-based Acid Dyes

Acid dyes are used primarily in the textile industry for dyeing natural fibres and synthetics (e.g., wool, silk, nylon, polyesters, acrylic and rayon). The sulfonic acid groups react with the cationic amino groups in the fibres (ETAD 1995b). To a lesser extent, these dyes are used in other applications, such as leather, plastics, inks and paints (ETAD 1995b; Øllgaard et al. 1998; CII 2011).

The dominant use of Acid Red 97 was identified for dyeing leather and textiles (CII 2011).

4.2.2 Benzidine-based Direct Dyes

Direct dyes, because of their larger and planar structure, exhibit high affinity for cellulose fibres and are applied directly to cellulose-containing materials in a neutral dye bath (ETAD 1995b; Hunger 2003). They are used for dyeing cotton, rayon and, to a lesser extent, wool, silk and nylon (Environment Canada and Health Canada 2013). In addition to the general uses of Benzidine-based Direct Dyes, a specific therapeutic use of Direct Blue 14 as an active ingredient in an ophthalmic solution used in eye surgery was identified in Canada (DPD 2012), Direct Red 28 in testing hydrochloric acid in gastric contents and for testing amyloidosis (Merck Index 2001), and Direct Blue 15 as a tint, a type of photographic agent, for cinematographic films (HSDB 1983–).

4.2.3 Benzidine-based Cationic Indicators

TDBPD was identified as being used as a laboratory chemical and in scientific research and development (ChemIDplus 1993; Merck Index 2001; SPIN 2010). It is used as a laboratory agent and intermediate in the synthesis of a dark blue pigment (Merck Index 2001).

4.2.4 Benzidine-based Precursors

The substance TCDB is expected to be used not as a dye but as a stabilized diazo compound that can be used in print pastes for textile printing with azoic dyes (Kirk-Othmer 2010). It can also be used as a precursor or intermediate.

Naphthol AS-BR can be used as a precursor and coupling component to make Naphthol AS diazo pigments and for dyeing cloth and printing textiles with azoic dyes (Kirk-Othmer 2010).

4.2.5 Benzidine Derivatives

Benzidine Derivatives are primarily used as intermediates for the synthesis of colourants and chemicals that are later used in the preparation of other materials, such as textiles, foam, elastomers and plastics (HSDB 1983– ; Lide 2002; ChemicalLand21 2010; Kirk-Othmer 2010; US EPA 2010).

4N-TMB has been used as a chemical intermediate in the manufacture of triarylmethane dyes (Kirk-Othmer 2010).

TODI is used in the manufacture of urethane, plastics and plasticizers. More specifically, TODI is used as a polymer intermediate to produce urethane elastomers used to manufacture seals, automobile parts, specialty polyurethanes and polyurethane adhesives for food packaging (Aznar et al. 2011). TODI is also used as a reactant in the plastics pipe, pipe fitting and unlaminated profile shape manufacturing sector in the United States (US EPA 2010).

3,3′-DMB·2HCl was listed as an intermediate in pigment and dyes manufacturing (Merck Index 2001; Sigma-Aldrich Canada 2010) and in the synthesis of biphenyl and diphenyl ethers. It is also used as a reagent for gold and free chlorine in water (Merck Index 2001).

3,3′-DMOB and 3,3′-DMB are both used as chemical intermediates in the manufacture of azo dyes (Lewis 1997; ATSDR 2001; Merck Index 2001). 3,3′-DMOB may also be used in the production of dyes for dyeing leather, paper, plastic, rubber and textiles (IARC 1974a) and as a chemical intermediate in the production of o-dianisidine diisocyanate (IARC 1974a). 3,3′-DMB may be used as a reagent for gold and free chlorine in water (Merck Index 2001) and as a curing agent for urethane resins (Lewis 1997).

None of the Benzidine Derivatives were identified for use in food packaging materials in Canada (2011 emails from the Food Directorate, Health Canada, to the Risk Management Bureau, Health Canada; unreferenced). Although TODI was identified as a diisocyanate used in polyurethane adhesives in food packaging laminates in Europe (Aznar et al. 2011), there are no indications of this use in Canada (2012 email from the Food Directorate, Health Canada, to the Existing Substances Risk Assessment Bureau, Health Canada; unreferenced).

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5. Environmental Fate and Behaviour

The environmental fate of chemicals describes the processes by which chemicals move and are transformed in the environment. In this section, some general characteristics of the substances considered in this Screening Assessment will be discussed with respect to their environmental fate in different compartments in an effort to understand how organisms come into contact with the substances in a particular medium, the persistence of the substances in environmental compartments, and their degradation, distribution among media, migration in groundwater, removal from effluents by standard wastewater treatment methods and bioaccumulation in organisms.

As explained in Environment Canada and Health Canada (2013), the Equilibrium Criterion model or EQC (2003) is not applicable for strongly ionic acid and direct dyes and charged, basic substances such as cationic indicators, as they do not fall under the model domain. Therefore, the environmental fate and compartmentalization of these substances will be discussed qualitatively using information on physical and chemical properties. EQC will be used to help characterize and quantify the environmental behaviour of Benzidine Derivatives and Benzidine-based Precursors.

5.1 Releases to Water and Sediment

Dyes (including Benzidine-based Acid and Direct Dyes as well as Benzidine-based Cationic Indicators) have an inherently high affinity to substrates, which results in high fixation rates (ETAD 1995b; Environment Canada and Health Canada 2013). If released to natural waters or wastewater in an untransformed state, negatively charged Benzidine-based Acid and Direct Dyes are expected to primarily bind to suspended organic matter, specifically positively charged particulates, due to electrostatic interactions and eventually settle out to bed sediments or wastewater sludge (ETAD 1995b). Positively charged Benzidine-based Cationic Indicators are expected to have an affinity for ionic substrates such as dissolved organic material, which has a net negative charge due to the presence of humic and fulvic acids. Some ionic dyes can also bind to organic material via hydrogen bonds and van der Waals forces (Oster 1955).

Other factors, such as increasing molecular size, hardness of the water and salinity, as well as decreasing pH, are thought to favour some sorption of azo dyes to suspended solids (HSDB 1983– ; Øllgaard et al. 1998). It has been stated generally that, due to the recalcitrant nature of azo dyes in aerobic environments, they eventually end up in anaerobic sediments, shallow aquifers and groundwater (Razo-Flores et al. 1997). After partitioning to sediment or wastewater sludge, some azo dyes may bind reversibly and become resuspended, while others will bind irreversibly and remain buried.

A smaller proportion of ionic dyes may also reside in the water column due to their very high water solubility. Eventually, these dyes may form associations with organic material and settle out to sediments.

Certain dyes based on benzidine may also be biotransformed in sediment to benzidine derivatives. EQC analysis was performed for 3,3′-DMB, 3,3′-DMOB and 4N-TMB, which are relatively neutral substances except at lower pHs. Since 3,3′-DMB·2HCl is a salt of 3,3′-DMB and TODI hydrolyzes to 3,3′-DMB, these three substances are expected to behave similarly to the other two Benzidine Derivatives. If Benzidine Derivatives are completely released to water (at a level of 100%), EQC modelling indicates that more than 90% of the mass of Benzidine Derivatives will remain in water, with only a small percentage (0.2–9.3%) going to sediment. However, as indicated in Environment Canada and Health Canada (2013), sorption is an important fate process for aromatic amines in aquatic and sediment systems (Colon et al. 2002; Chen and Nyman 2009); therefore, it is anticipated that a greater percentage of Benzidine Derivatives will be found in sediment than is predicted by EQC.

When Benzidine-based Precursors are released to water only, EQC modelling indicates that 99.1% of TCDB will remain in water, with less than 1% going to sediment due to its moderate solubility. This contrasts with a prediction of 92.9% of Naphthol AS-BR going to sediment, with only 7.1% staying in water, due to its very low solubility.

5.2 Releases to Soil

There are two major routes for the release of azo dyes to soil: directly via the use or application of a dye in the environment and indirectly via the application of wastewater biosludge to agricultural land or deposition in landfills.

It is expected that ionic dyes will have high to moderate mobility in soil due to low soil–water adsorption coefficient (Kd) values (Øllgaard et al. 1998); this is tempered with the finding that they may also undergo ion exchange processes with clay in soil, which would retard leaching (HSDB 1983). Specifically, acid dyes have an inherently high affinity for substrates, with fixation levels ranging from 85% to 98% for acid dyes with more than one sulfonic acid group (ETAD 1995b).

Certain azo dyes may also biotransform in soil to Benzidine Derivatives. If Benzidine Derivatives are completely released to soil, EQC modelling for 3,3′-DMB, 3,3′-DMOB and 4N-TMB indicates that the majority (81–99%) of them are expected to remain in soil . Again, 3,3′-DMB·2HCl and TODI are expected to behave similarly.

Using EQC modelling, the two Benzidine-based Precursors, Naphthol AS-BR and TCDB, are both expected to remain in soil when released to that medium (99.9% and 96.6%, respectively), with only a small amount going to air in the case of TCDB (3.3%) and negligible amounts going to sediment ( less than  0.1% for both substances).

5.3 Releases to Air

Benzidine-based Dyes are not expected to be released to air and are not expected to partition to this compartment due to very low vapour pressures and Henry’s Law constants (HSDB 1983– ; Øllgaard et al. 1998). Water-soluble dyes such as Benzidine-based Acid and Direct Dyes as well as Benzidine-based Cationic Indicators are intended for use in water-based treatments, which also limits their release, as they are hydrophilic. While premixed dyes in their solid states may have some limited capacity for dispersal into the air as large particles, air is not considered to be a carrying medium for dyes, as these substances exhibit low or negligible volatility (ETAD 1995b; Øllgaard et al. 1998).

Benzidine Derivatives and Benzidine-based Precursors are also not expected to be released to air. However, if released entirely to air, the majority of Benzidine Derivatives and Benzidine-based Precursors are not expected to stay in this medium, but mostly are predicted by EQC modelling to be deposited to soil (78.6–97.5%).

Given low levels of volatility and physicochemical preference for partitioning to other media, it is also not expected that water-soluble or water-insoluble azo dyes will be subject to long-range atmospheric transport.

5.4 Environmental Persistence

In order to characterize the environmental persistence of Benzidine-based Dyes, Benzidine-based Cationic Indicators, Benzidine-based Precursors and Benzidine Derivatives, empirical and modelled data for these substances were considered under both aerobic and anaerobic conditions. In addition, the process of ecological biotransformation was explored, specifically with respect to the potential for Benzidine-based Dyes to degrade to benzidine derivatives.

5.4.1 Empirical Data for Persistence

Empirical biodegradation data related to the persistence of Benzidine-based Dyes and Benzidine-based Precursors are limited. Select empirical tests (Table 5-1) under aerobic conditions, using standard Organisation for Economic Co-operation and Development (OECD Test Guideline 301C) methodologies, showed no degradation after 42 and 28 days for Acid Red 114 and Direct Black 38, respectively (CHRIP ©2002-2012).

Some conflicting degradation data for azo and benzidine-based dyes have been found. For example, Idaka et al. (1985) reported relatively high colour loss for the benzidine dye Direct Red 28 at 85% and the azo dye Acid Orange 7 at 65% after 150 days. However, since sludge was acclimated to the dyes, longer test periods were used and the percent colour loss was provided instead of percent theoretical oxygen demand uptake, it makes the values difficult to compare with the results of standard OECD Test Guideline 301C tests. Also, it is unclear whether commercial formulations may have been used in some of these tests instead of high-purity dyes.

Table 5-1. Select empirical data for biodegradation of Benzidine-based Dyes under aerobic conditions
Subgroup (specific substance with data)MethodDegradation value (%)Footnote Table 5-1 [a]Degradation endpointTest duration (days)Reference
Benzidine-based Acid Dyes (Acid Red 114)OECD Test Guideline 301C0Biodegradation42CHRIP ©2002-2012
Benzidine-based Direct Dyes (Direct Black 38)OECD Test Guideline 301C0Biodegradation28CHRIP ©2002-2012
Footnote Table 5-1 a

Percent biodegradation at a given concentration of the test substance.

Return to footnote Table 5-1 a referrer

In addition to biodegradation tests under aerobic conditions, it is notable that data for Direct Blue 14 showed 50% degradation in anaerobic sediment after 16 days (Table 5-2).

Table 5-2. Select empirical data for biodegradation of Benzidine-based Dyes under anaerobic conditions
Subgroup
(specific substance with data)
MethodDegradation value (%)Footnote Table 5-2 [a]Degradation endpointTest duration (days)Reference
Benzidine-based Direct Dyes
(Direct Blue 14)
Not listed50Water/sediment biodegradation16Weber 1991
Footnote Table 5-2 a

Percent biodegradation at a given concentration of the test substance.

Return to footnote Table 5-2 a referrer

Significantly more empirical data were available on the Benzidine Derivatives. An empirical abiotic degradation study (hydrolysis) conducted according to OECD Test Guideline 111 (hydrolysis as a function of pH) and EU Method C.7 is available for TODI (ECHA 2012). Results of the study conducted at three pHs (pH 4, 7 and 9) and two temperatures (25°C and 50°C) show rapid hydrolysis rates for TODI, with half-lives of 16 hours (at 25°C and pH 7), 1.2 hours (at 50°C and pH 7) and equal to or less than 2 minutes under all other experimental conditions. The main hydrolysis product identified was 3,3′-DMB, but a second unidentified compound with a molecular weight of 240 g/mol was also observed (ECHA 2012).

Empirical biodegradation studies have been identified for 3,3′-DMB, 3,3′-DMOB and TODI (Table 5-3).

Results from these studies vary for 3,3′-DMB and 3,3′-DMOB. Indeed, 28-day ready biodegradability (Japanese) Ministry of International Trade and Industry (MITI) tests conducted with an initial substance concentration of 100 mg/L indicate that 3,3′-DMB and 3,3′-DMOB are not readily biodegradable (Kawasaki 1980; MITI 1992). Another study conducted by Baird et al. (1977) using conventional Warburg bioassay techniques with 3,3′-DMB and 3,3′-DMOB at a concentration of 20 mg/L at 25°C in activated sludge determined that both substances were completely degraded (100%) over 6 hours. It is noted that the activated sludge samples were chosen from two treatment facilities that combined domestic and industrial discharges; therefore, microorganisms may have been acclimated to aromatic amines. Also, the substances were degraded in spite of some inhibitory effects on the oxygen uptake of microorganisms, indicating that the effects may have been caused by the unidentified degradation products (Baird et al. 1977).

Biodegradation of 3,3′-DMOB was investigated using a 28-day OECD Test Guideline 301A test at six laboratories (Brown and Laboureur 1983). The tests were carried out using the substance at concentrations of 20 and 30 mg/L of sludge, predominantly from domestic discharges. The OECD Test Guideline 301A method was modified, and varying quantities of yeast extracts were added. Complete degradation (100%) was obtained at a high yeast concentration; however, a clear dependence of the extent of degradation on high levels of yeast was observed, indicating that the substance is “not readily biodegradable” (Brown and Laboureur 1983).

The aerobic and anaerobic biodegradabilities of 3,3′-DMOB were investigated by Brown and Hamburger (1987). 3,3′-DMOB was more than 75% degraded in a 28-day ready biodegradability screening test (OECD Test Guideline 301E) with 30 mg/L of innoculum sludge under aerobic conditions. 3,3′-DMOB was completely degraded after 42 days under anaerobic conditions using a protocol identical to the one used in Brown and Laboureur (1983).

Results from a 28-day ready biodegradability closed bottle test (OECD Test Guideline 301D) on TODI (ECHA 2012) showed 0% degradation, indicating that the substance is not readily biodegradable. Since TODI is known to rapidly hydrolyse to 3,3′-DMB in water, this result may also confirm the recalcitrant nature of 3,3′-DMB.

Table 5-3. Empirical data for degradation of Benzidine Derivatives
ChemicalMediumFate processDegradation valueDegradation endpoint (test duration)Reference
3,3′-DMBWaterBiodegradation6% (HPLC)
3% (BOD)
(100 mg/L)
% biodegradation (28 days)MITI 1992
3,3′-DMBWaste waterBiodegradation100%
(20 mg/L)
% biodegradation (6 h)Baird et al. 1977
3,3′-DMOBWaterBiodegradationResistant to biodegradation% biodegradation (28 days)Kawasaki 1980
MITI  1992
3,3′-DMOBWaste waterBiodegradation100%
(20 mg/L)
% biodegradation (6 h)Baird et al. 1977
3,3′-DMOBWaste waterBiodegradation0–100%% biodegradation (28 days)Brown and Laboureur 1983
3,3′-DMOBWaterBiodegradationgreater than 75%% biodegradation (28 days)Brown and Hamburger 1987
3,3′-DMOBWaterBiodegradation100%% biodegradation (anaerobic) (42 days)Brown and Hamburger 1987
TODIWaterHydrolysis100% in 29 h
t½ = 16 h
% hydrolysis (25°C)ECHA 2012
TODIWaterBiodegradation−4.8%% biodegradation (28 days)ECHA 2012

Abbreviations:
BOD, biological oxygen demand;
HPLC, high-performance liquid chromatography;
t½, half-life

5.4.2 Modelling of Persistence

In addition to the experimental data, a (Q)SAR-based weight of evidence approach (Environment Canada 2007) was applied using biodegradation models. These models are considered acceptable for use, as they are based on chemical structure, and the azo structure is represented in the training sets of all the BIOWIN models used, thereby increasing the reliability of the predictions. Given the ecological relevance of the water compartment, the fact that most of the available models apply to water and the fact that Benzidine-based Dyes are expected to be released to this compartment, aerobic biodegradation in water was primarily examined.

Tables A-9a–A-9e in Appendix A summarize the results of available (Q)SAR models for degradation in various environmental media. Aquatic degradation models used in this analysis were HYDROWIN (2010), BIOWIN Submodels 3–6 (BIOWIN 20010, DS TOPKAT (©2005–2009) and CATALOGIC (©2004–2011).

All of the model outputs for Benzidine-based Dyes and Benzidine-based Cationic Indicators as well as all but one model output for Benzidine-based Precursors (BIOWIN Submodel 4) consistently predicted that these substances would biodegrade slowly in water under aerobic conditions (Appendix A, Tables A9a–d). These results are consistent with information included in Environment Canada and Health Canada (2013), which outlines the general persistence of azo dyes in aerobic environments.

Table A-9e in Appendix A summarizes the results of available (Q)SAR models for degradation of Benzidine Derivatives in various environmental compartments. Biodegradation in water was primarily examined for substances in the Benzidine Derivatives subgroup, given the ecological importance of the water compartment and the fact that they are expected to be released to this compartment. Only TODI contains a functional group (isocyanate) expected to undergo hydrolysis. Atmospheric degradation predictions were also generated, since Benzidine Derivatives can be semi-volatile.

Gaseous amines have the potential to undergo atmospheric oxidation reactions with hydroxyl, nitrogen oxide or ozone radicals (Ge et al. 2011). Modelled and calculated half-lives of 0.052–0.25 day for the Benzidine Derivatives subgroup indicate that these substances rapidly photooxidize via reaction with hydroxyl radicals (Meylan and Howard 1993; AOPWIN 2010). Degradation of the Benzidine Derivatives via hydroxyl radicals will decrease if the substances are bound to particulate matter. Removal of amines by reactions with ozone is generally negligible (Ge et al. 2011), and no predictions could be made for the Benzidine Derivatives subgroup using AOPWIN (2010).

Overall, all model results generated for 3,3′-DMB, 3,3′-DMOB and 4N-TMB indicate that while some transformation of the parent structure may occur (primary degradation) for 3,3′-DMOB, all three substances are not expected to significantly biodegrade in the environment.

5.4.3 Aerobic Biodegradation of Benzidine-based Dyes and Benzidine-based Cationic Indicators

The available empirical and modelled persistence data for Benzidine-based Acid and Direct Dyes as well as Benzidine-based Cationic Indicators in aerobic environments are in agreement and show little to no biodegradation in the time scale of the studies. This is consistent with the understanding that dyes must be stable to end use applications in order to be effective and that most are generally considered non-degradable under environmentally relevant aerobic conditions (Pagga and Brown 1986; Øllgaard et al. 1998; ETAD 1995b).

5.4.4 Anaerobic Biodegradation of Benzidine-based Dyes and Benzidine-based Cationic Indicators

Under anaerobic or reducing conditions, biotic degradation of dyes may take place relatively rapidly (Yen et al. 1991; Baughman and Weber 1994; ETAD 1995b; Øllgaard et al. 1998; Isik and Sponza 2004). Dyes have a high tendency to cleave at the azo bond, with the formation of aromatic amines (Øllgaard et al. 1998; Hunger 2005). Direct Blue 14 has been documented to degrade in anaerobic sediment–water systems with a half-life of 16 days (Weber 1991). Total mineralization or further degradation of these metabolites could take place if they are transferred (e.g., by sediment resuspension) to aerobic environments (Øllgaard et al. 1998; Isik and Sponza 2004).

In order to account for potentially harmful transformation products, the scientific literature was consulted, and the (Q)SAR-based predictive tool CATALOGIC (©2004–2011) was used to determine what benzidine derivatives may be released from Benzidine-based Dyes and Benzidine-based Cationic Indicators in the environment. Simulations were generated using CATALOGIC’s metabolite simulator based on biological oxygen demand (BOD) data from MITI. Potential degradation products were prioritized based on the percent likelihood of formation, potential hazard and the known commercial activity of their parent compounds. Experimental evidence from Brown and Hamburger (1987) indicates that Direct Blue 14 and Acid Red 114 (a close analogue to Acid Red 111) have the potential to transform to 3,3′-DMB in sediment under anaerobic conditions. Based on the structural similarities between Acid Red 114 and Acid Red 111, it is anticipated that Acid Red 111 would also transform to 3,3′-DMB under similar conditions. Empirical degradation studies conducted with Direct Black 38 identified three different metabolites--namely, aniline (CAS RN 62-53-3) (Isik and Sponza 2004), benzidine and 4-biphenylamine (CAS RN 91-67-1) (Isik and Sponza 2004; Bafana et al. 2009a). However, none of these metabolites are included in the Benzidine Derivatives subgroup assessed in this report.

Modelled data from CATALOGIC are consistent with these empirical results. 3,3′-DMB was identified as a potential degradation product for Direct Blue 14, Acid Red 111 and Acid Red 114, but not for Direct Black 38. Therefore, both Direct Blue 14 and Acid Red 114 have the potential to transform to 3,3′-DMB, one of the Benzidine Derivatives being assessed in this report, under conditions that favour biodegradation of the compounds. The contribution of these transformation products will be discussed further in the Ecological Exposure Assessment section.

Anaerobic biodegradation of the Benzidine-based Cationic Indicators is not expected, as no azo bond is contained within their chemical structures.

5.4.5 Hydrolysis of Benzidine-based Dyes, Benzidine-based Cationic Indicators and Benzidine-based Precursors

The majority of the Benzidine-based Acid and Direct Dyes, Benzidine-based Cationic Indicators and Benzidine-based Precursors do not contain functional groups expected to undergo hydrolysis. This is consistent with published studies that note hydrolysis as being an insignificant factor in the cleavage of azo compounds (Baughman and Perenich 1988). However, four substances, Acid Orange 56 and BADB (Benzidine-based Acid Dyes), BDAAH (a Benzidine-based Direct Dye) and Naphthol AS-BR (a Benzidine-based Precursor), contain amide functional groups that were flagged by EPISuite as having the potential to undergo some degree of hydrolysis.

5.4.6 Summary of Persistence

Due to the persistence of Benzidine-based Dyes, Benzidine-based Cationic Indicators, Benzidine-based Precursors and Benzidine Derivatives subgroups in aerobic environments in combination with their moderate to high water solubility, it is expected that these substances will have relatively long residence times in water. As these substances are predicted to stay in the water for long periods of time, they may disperse widely from point sources of release. Eventually, due to electrostatic interactions with negatively charged particulate matter, they will be deposited to sediment, where they will persist under aerobic conditions and remain a source of exposure to organisms until buried due to sedimentation. Deeper layers of sediment are likely under anaerobic conditions, which will transform (reduce) the dyes via azo hydrolysis. Exposure of the benthos under anaerobic conditions is not expected to be significant. Short residence times in air are expected to result in low potential for long-range atmospheric transport. 

5.5 Potential for Bioaccumulation

In this assessment, a variety of lines of evidence have been used to determine the bioaccumulation potential of Benzidine-based Dyes, Benzidine-based Cationic Indicators, Benzidine-based Precursors and Benzidine Derivatives. Experimental data for traditional bioaccumulation metrics such as bioconcentration factor (BCF) are minimal and restricted to the water compartment for these substances. In addition, the use of (Q)SAR bioaccumulation modelling was not pursued for Benzidine-based Dyes and Benzidine-based Cationic Indicators, since these substances were outside the model domains of applicability.

5.5.1 Octanol–Water Partition Coefficient

As indicated in Table 4a, Benzidine-based Dyes and Benzidine-based Cationic Indicators have relatively high water solubilities ( greater than  500–116 000 mg/L), and a limited number of experimental data for dyes suggest relatively low log Kow values (below 0.77), which would also suggest a very low bioaccumulation potential according to equilibrium partitioning theory. This is consistent with the general view from other sources that note the very low bioaccumulation potential of ionic dyes (ETAD 1995b).

Benzidine-based Precursors have high modelled log Kowvalues. Naphthol AS-BR has a modelled log Kow of 7.75, which, at above 7, is just over the threshold of indicating reduced bioaccumulation potential (Arnot and Gobas 2003). TCDB has a modelled log Kow of 5.13, which may be indicative of a high potential for bioaccumulation.

Benzidine-based Derivatives are moderately to highly water soluble (8.23–50 000 mg/L), and most have relatively low log Kow values (below 3.0). Modelled log Kowvalues for 4N-TMB ranged from 3.53 to 4.11, which suggests a moderate bioaccumulation potential. Estimated log Koavalues ranging from 9.48 to 13.21 suggest that, given a terrestrial dietary exposure, substances in the Benzidine-based Derivatives subgroup would not have the potential to biomagnify in terrestrial food webs, as suggested by Gobas et al. (2003) and Kelly et al. (2007), assuming some degree of metabolism (at least a metabolic rate constant of greater than or equal to  0.03/day) and some degree of assimilation efficiency from the diet.

5.5.2 Bioconcentration Factor (BCF)

Estimated and experimental log Kow values were compared with experimental BCFs for fish for a number of dyes (Anliker et al. 1981; ETAD 1995b; Øllgaard et al. 1998). With respect to the data for six acid dyes and one direct dye, reported BCFs were less than 10, indicating that these very hydrophilic (ionic) dyes are not likely to bioconcentrate in aquatic organisms. Data available for Acid Red 114 (Table 5-4) illustrate low BCF values (42–84 L/kg) for carp exposed to two different concentrations.

Table 5-4. Empirical data for bioconcentration of Benzidine-based Dyes (Acid Red 114)
Test organismExperimental concentration (mg/L)Endpoint (BCF, L/kg)Reference
Common carp (Cyprinus carpio)0.242–76MITI 1992
Common carp (Cyprinus carpio)0.0252–84MITI 1992

No experimental BCF data were available for the Benzidine-based Precursors, but estimated data are presented in Table 5-5, as (Q)SAR models were deemed acceptable for these substances based on their simpler chemical structures.

Modelled data for Naphthol AS-BR show some potential for bioaccumulation, with values above 5000, without factoring in metabolism. However, when metabolism is taken into consideration, the BCF value lowers significantly. TCDB shows moderate to high BCF values (1120–6456) before metabolism is factored in, but values are of much lower concern when metabolism is considered. However, since the log Kow and all of the BCF data are modelled, there is a high degree of uncertainty regarding the bioaccumulation potential of Naphthol AS-BR and TCDB and reason for some concern.

Table 5-5. Modelled BCF data for Benzidine-based Precursors
SubstanceLog KowMetabolism constant (kM) (/day)Test organismModel and model basisValue wet weight
(L/kg)
Reference
Naphthol AS-BR7.75NAFishBCFBAF
Submodel 1 (linear regression)
5699BCFBAF 2010
Naphthol AS-BR7.750.2909 (10 g fish)FishBCFBAF
(with biotransformation rate; mid-trophic)
106.2BCFBAF 2010
Naphthol AS-BR7.750.304FishBCFmax12 302
(Log BCFmax: 4.09)
CPOPs 2008
Naphthol AS-BR7.750.304FishBCF corrected7.4
(Log BCF corrected: 0.871)
CPOPs 2008
TCDB5.13NAFishBCFBAF
Submodel 1 (linear regression)
1120BCFBAF 2010
TCDB5.130.192 (10 g fish)FishBCFBAF
(with biotransformation rate; mid-trophic)
1623BCFBAF 2010
TCDB5.130.00363FishBCFmax6456
(Log BCFmax: 3.81)
CPOPs 2008
TCDB5.130.00363FishBCF corrected645.7
(Log BCF corrected: 2.81)
CPOPs 2008


While experimental data were also limited, BCF values were available for one of the five Benzidine Derivatives (3,3′-DMB). Low BCFs ranging from 4.8 to 83 in fish over a period of 56 days indicate that 3,3′-DMB has a low bioaccumulation potential (Table 5-6).

Low BCFs (55–2617) have also been observed for benzidine in fish, Daphnia magna, mosquito larvae, snails and filamentous green algae (Lu et al. 1977; Freitag et al. 1985; Tsuda et al. 1997). Based on these values, it is anticipated that the substances from the Benzidine Derivatives subgroup have a low bioaccumulation potential.

Table 5-6. Empirical data for bioconcentration of Benzidine Derivatives (3,3′-DMB)
Test organismExperimental concentration (mg/L)Endpoint (BCF, L/kg)Reference
FishNot known47SRC 2011
Common carp(Cyprinus carpio)0.24.8–34MITI 1992
Common carp (Cyprinus carpio)0.0210–83MITI 1992


Modelled bioaccumulation factor (BAF)/BCF data for Benzidine Derivatives are in general agreement with experimental values, showing low bioaccumulation potential in the range of 5.2–1173.

5.5.3 Other Factors for Assessing Bioaccumulation Potential

As outlined in the Potential for Bioaccumulation section of Environment Canada and Health Canada (2013), due to the lack of empirical bioaccumulation data available for Benzidine-based Dyes and Benzidine-based Cationic Indicators, available data on water solubility, molecular weight and cross-sectional diameter are considered in order to determine bioaccumulation potential. Given their relatively high water solubility, ionic nature and high degree of dissociation under typical environmental conditions, the lipid partitioning tendency of these substances is expected to be limited. Also, bioaccumulation data resulting from exposures of organisms to these substances in soil and sediment are minimal and limited, in large part due to the high water solubility of these substances.

In general, Benzidine-based Dyes are relatively hydrophilic, large molecules with high molecular weight (696–1134 g/mol). The minimum and maximum cross-sectional diameters for Benzidine-based Dyes range from 0.95 nm (Dmin) to 2.07 nm (Dmax) (Table 4a). These characteristics suggest a low bioaccumulation potential for these substances.

Cross-sectional diameters for the Benzidine-based Cationic Indicators could not be determined by the models.

Effective diameters of the Benzidine Precursors ranged from 0.83 nm (Dmin for TCDB) to 1.42 nm (Dmax for Naphthol AS-BR) (Table 3-2). This indicates that Naphthol AS-BR may have somewhat less bioaccumulation potential than TCDB.

Benzidine Derivatives were found to have effective cross-sectional diameters that ranged from 0.65 nm (Dmin) to 0.86 nm (Dmax) (Table 3-3). Benzidine Derivatives evaluated in this assessment generally have low molecular weights (212–285 g/mol) and low effective cross-sectional diameters. Therefore, this would not be a factor in restricting the rate of uptake when crossing cell membranes.

5.5.4 Summary of Bioaccumulation Potential

Benzidine-based Dyes and Benzidine-based Cationic Indicators are expected to have a low bioaccumulation potential due to low observed bioconcentration in empirical tests. This is supported by and consistent with their physical and chemical properties (i.e., low log Kow, ionized at relevant environmental pH, high molecular weight, large cross-sectional diameters, high water solubility) and likely high degree of biotransformation by organisms. Benzidine-based Precursors are expected to be moderately bioaccumulative due to their high modelled BCF values without consideration of metabolism and physical and chemical properties (i.e., moderate to high log Kow, molecular weight and cross-sectional diameters that do not slow uptake rates). Low experimental and modelled BCF and BAF data for Benzidine Derivatives, physical and chemical property information (i.e., high water solubility, low log Kow) that suggests a low lipid partitioning tendency plus an inherent ability to be biotransformed in organisms indicate consistently that these substances have low bioaccumulation potential.

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6. Potential to Cause Ecological Harm

6.1 Ecological Effects Assessment

Only empirical data (from specific substances within the subgroups and analogues) were considered for assessment of the ecological effects potential of Benzidine-based Dyes given the high level of uncertainty associated with modelling the ecotoxicity of these substances. With respect to the Benzidine Derivatives, empirical and modelled data were considered.

6.1.1 Empirical Studies for the Aquatic Compartment

A variety of lines of evidence have been used to determine the ecotoxicological potential of the Benzidine-based Dyes, Benzidine-based Precursors and Benzidine Derivatives.

Limited empirical studies were available for Benzidine-based Dyes (Table 6-1). Most substances had very few or no empirical data, whereas other substances, such as Direct Blue 14, had multiple studies in the aquatic compartment.

One empirical toxicity study was available in which the acute toxicity of various dyes (46 in total) to fathead minnow (Pimephales promelas) was studied (Little and Lamb 1972). Bioassays were carried out according to published standard methods, and data sheets for each test were prepared, including pertinent information on the test organisms, dilution water and test conditions. The experiment was designed to estimate the threshold concentration at which 50% of the experimental animals survived after 96 hours (TL50). The TL50 for 29 dyes, including 2 dyes from the Benzidine-based Direct Dyes subgroup (Direct Brown 95 and Direct Black 38), was in excess of 180 mg/L (highest concentration tested).

Empirical toxicity studies for Benzidine-based Acid Dyes were available for Acid Red 114 (MITI 1992) and a formulated product, Lanasyn Scarlet F-3GL 130, which contains Acid Red 111 (Study Submission 2007). Since the Acid Red 111 study, which yielded a 48-hour median lethal concentration (LC50) of 11 mg/L for rainbow trout (Oncorhynchus mykiss), used a formulation with an unknown concentration of the Benzidine-based Acid Dye and additional study details were not reported, it was not considered to be used as a critical value but is considered for comparative purposes in the weight of evidence. The study on Acid Red 114 by MITI (1992) was done on common carp (Cyprinus carpio) and yielded the lowest value for Benzidine-based Acid Dyes, with an LC50 of 4 mg/L (Table 9a). These two studies suggest that Acid Red 111 and Acid Red 114 would have a moderate (bordering on high) potential for toxicity.

Toxicity studies on the freshwater shrimp (Desmocaris trispinosa) were available for two Benzidine-based Direct Dyes (Direct Blue 14 and Direct Red 2). The 96-hour LC50 for Direct Red 2 was reported as 4.96 mg/L, while that for Direct Blue 14 was 11.33 mg/L (Ogugbue and Oranusi 2006). The authors stated that the difference in the toxicities may be partially explained by the lower molecular weight of Direct Red 2 compared with Direct Blue 14 (i.e., lower molecular weight substances are transported more rapidly across the cell membrane). This pattern appeared more evident when a lower molecular mass substance (Mordant Black 17, CAS RN 2538-85-4) and two higher molecular weight substances (Reactive Red 4, CAS RN 17681-50-4and Reactive Yellow 2, CAS RN 50662-99-2) were included in the data set.

In general, when data from multiple taxa were available, fish species seemed to have a higher tolerance to Benzidine-based Dyes than other taxa (see Table 6-1). Invertebrate and algal species appeared to be more susceptible to Benzidine-based Dyes.

The above results are similar to what has been reported generally for acid and direct dyes in the literature (Øllgaard et al. 1998). The Danish Environmental Protection Agency (EPA) study summarized short-term test results for zebrafish, Daphnia magna, algae and bacteria from a study by the Ecological and Toxicological Association of Dyes and Organic Pigment Manufacturers (ETAD) covering 47 dyes of different chemical dye classes (although specific dyes included in the study were not reported). In 96-hour LC50 tests on zebrafish, toxicity was observed for two acid dyes between 1 and 10 mg/L, for three acid dyes between 10 and 100 mg/L and for six other acid dyes above 100 mg/L. For direct dyes, all seven dyes tested reported toxicity to zebrafish at levels above 100 mg/L. Effects were observed between 10 and 100 mg/L and above 100 mg/L in 48-hour median effective concentration (EC50) tests (endpoint not specified) on D. magna for nine acid dyes. Similar tests with direct dyes resulted in adverse effects on D. magna at levels above 100 mg/L. Algal toxicity (measured in 72-hour EC50tests) was observed in two cases for acid dyes below 1 mg/L, as well as above this level (for the remaining seven acid dyes), whereas toxicity results for the seven direct dyes tested were all above 1 mg/L. Algae appeared to be the most susceptible organisms for all dye classes tested; the effect was thought to be related to light inhibition at high dye concentrations (coloration of water can occur above 1 mg/L). Generally, bacteria were the least susceptible to most of the different classes of dyes tested, compared with the other organisms (median inhibitory concentrations [IC50s] greater than 100 mg/L).

Experience with over 200 acid dyes has led to the observation that the potential ecotoxicity of such substances may generally be predicted by the number of sulfonic acid groups present (US EPA 2002). Some monosulfonated and disulfonated dyes have shown high to moderate toxicity (i.e., acute values less than  1 mg/L and less than  100 mg/L, respectively) to fish and other aquatic organisms. Dyes with three or more acid groups showed low toxicity (i.e., acute values greater than  100 mg/L) towards fish and invertebrates. All acid dyes showed moderate toxicity to green algae, with further analysis suggesting that such effects may have been related to light shading. For these generalizations to be applicable, the acid dyes must have some water solubility, and molecular weights generally need to be near or below 1 000, which is the case for Acid Red 111.

Furthermore, Environment Canada has evaluated numerous acid dyes under the New Substances Notification Regulations and has generally found anionic dyes to be of low toxicity regardless of the number of sulfonic acid groups, but some exceptions have been found (e.g., when a reactive functional group is not hindered).

Few experimental toxicity tests are available for cationic indicators as well. Since cationic indicators are similar in physical and chemical properties to basic (cationic) dyes, some data available for these substances will be used as read-across. In general, cationic indicators are known to have high levels of toxicity that exceed those of acid and direct dyes (Øllgaard et al. 1998). While most available data originate from studies on triarylmethane basic dyes, these are quite different in chemical structure from azo- and benzidine-based dyes. Therefore, for comparative purposes, only benzidine-based basic dye and azo-based basic dye data (for BB4), as well as data for a tetrazolium substructure (TTC), are used in this report for read-across to cationic indicators, as these are structurally more similar substances.

Read-across toxicity data used for Benzidine-based Cationic Indicators range from 2.5 to 10 mg/L, with the most sensitive values being for algae. As noted for acid dyes, the low endpoint value for algae may be due to shading, so, while sensitive, this organism is not the most indicative of cationic indicator toxicity. Most data for Benzidine-based Cationic Indicators are quite old, with studies published in the 1950s, 1960s and 1970s. Fish toxicity data occupy a relatively tight range from 5 to 10 mg/L, with the most reliable, recent and among the most sensitive studies coming from Little and Lamb (1973, 1974), at 5.6 and greater than  5.6 mg/L for fathead minnow (Pimephales promelas) exposed to BB4 for 96 hours.

Table 6-1. Empirical data for aquatic toxicity from representative substances for the Benzidine-based Acid Dyes, Benzidine-based Direct Dyes and Benzidine-based Cationic Indicators subgroups
SubgroupTest organismType of test (duration)EndpointValue (mg/L)Reference
Benzidine-based Acid DyesFish (Cyprinus carpio)Acute (96 h)LC504Footnote Table 6-1[a](Acid Red 114)MITI 1992
Benzidine-based Acid DyesFish (Oncorhynchus mykiss)Acute (48 h)LC5011 (Acid Red 111)Study Submission 2007
Benzidine-based Direct DyesAlga (Scenedesmus subspicatus)Chronic (72 h)EC50 (biomass)4.7 (Direct Blue 15)Brown 1992
Benzidine-based Direct DyesFreshwater shrimp
(Desmocaris trispinosa)
Acute (96 h)LC504.96 (Direct Red 2)
11.3[a] (Direct Blue 14)
Ogugbue and Oranusi 2006
Benzidine-based Direct DyesFish (Oncorhynchus mykiss)Acute (96 h)LC50560 (Direct Blue 15)Douglas et al. 1986
Benzidine-based Direct DyesFish (Oryzias latipes)Acute (96 h)LC50greater than 1000 (Direct Blue 14)Tsuji et al. 1986
Benzidine-based Direct DyesFish (Pimephales promelas)Acute (96 h)LC50greater than 180 (Direct Black 38 and Direct Brown 95)Little and Lamb 1973
Benzidine-based Direct DyesFish (Danio rerio)Acute (96 h)LC50greater than 500 (Direct Blue 15)Brown 1992
Benzidine-based Cationic IndicatorsAlga
(Pseudokirchneriella subcapitata)
Chronic (14 days)Population growth10 (BB4)Ericson 1977
Benzidine-based Cationic IndicatorsAlga (Anacystis aeruginosa)Chronic (1 day)Population growth2.5 (TTC)Fitzgerald et al. 1952
Benzidine-based Cationic IndicatorsFish (Petromyzon marinus, Ptychocheilus oregonensis, Oncorhynchus tshawytscha, Oncorhynchus kisutch)Acute (24 h)LC505–10 (TTC)Applegate et al. 1957; MacPhee and Ruelle 1969
Benzidine-based Cationic IndicatorsFish (Pimephales promelas)Acute (96 h)LC505.6 – greater than  5.6
(BB4)
Little and Lamb 1973, 1974


Abbreviations:
EC50, the concentration of a substance that is estimated to cause some effect on 50% of the test organisms;
LC50, the concentration of a substance that is estimated to be lethal to 50% of the test organisms

Footnote Table 6-1 a

Robust study summaries were conducted to determine the quality of the studies and are available upon request.

Return to footnote Table 6-1 a referrer

A limited number of empirical ecotoxicity studies are available for 3,3′-DMB and TODI (see Table 6-2 and divided up by substance in Appendix A, Table A-10).

Results indicate that both substances are generally expected to be moderately toxic to aquatic organisms, as indicated by a 72-hour EC50 ( greater than or equal to 1.5–6.3 mg/L) in algae, a 48-hour EC50 ( greater than or equal to 1.5–4.5 mg/L) in Daphnia and a 96-hour LC50(0.25–13 mg/L) in fish. It is unclear whether the elevated aquatic toxicity value for fish (96-hour LC50 of 0.25 mg/L) for TODI (ECHA 2012) is driven by TODI or its corresponding amine 3,3′-DMB, since the substance hydrolyses rapidly when in contact with water. A chronic 21-day EC50 value of 0.64 mg/L for Daphnia (MITI 2000) for 3,3′-DMB is the most conservative empirical value available for the Benzidine Derivatives.

Table 6-2. Summary of empirical data for aquatic toxicity for Benzidine Derivatives
SubgroupTest organismType of test (duration)EndpointValue (mg/L)Reference
Benzidine Derivatives (3,3′-DMB and TODI)AlgaChronic (72 h)EC50greater than or equal to 1.5–6.3ECHA 2012; MITI 2000
Benzidine Derivatives (3,3′-DMB and TODI)DaphniaAcute (48 h)EC50greater than or equal to 1.5–4.5Kuhn 1989Footnote Table 6-2 [a]; ECHA 2012; MITI 2000
Benzidine Derivatives (3,3′-DMB and TODI)DaphniaChronic (21 days)EC500.64MITI 2000
Benzidine Derivatives (3,3′-DMB and TODI)FishAcute (96 h)LC500.25–13ECHA 2012; MITI 2000


Abbreviations:
EC50, the concentration of a substance that is estimated to cause some effect on 50% of the test organisms;
LC50, the concentration of a substance that is estimated to be lethal to 50% of the test organisms

Footnote Table 6-2 a

24-hour EC50 (behaviour) = 3.2 mg/L for 3,3′-DMB.

Return to footnote Table 6-2 a referrer

No experimental toxicity data are available for the Benzidine-based Precursors or read-across substances.

6.1.2 Empirical Studies for Other Environmental Compartments

Toxicological data were reported for a formulated product, trypan blue, which contains 80% Direct Blue 14 (Hulzebos et al. 1993). Seven-day and 14-day EC50 growth studies resulted in moderate (bordering on high) toxicity values of 263 (196–352) µg/g of soil and 290 (250–337) µg/g of soil, respectively, for lettuce (Lactuca sativa) (Table 6-3).

No empirical data were available on the toxicity of Benzidine-based Dyes in sediment.

Table 6-3. Empirical soil toxicity data from representative substances in the Benzidine-based Direct Dyes subgroup
SubgroupTest organismType of testEndpointValue (mg/kg of soil)Footnote Table 6-3 [a]Reference
Benzidine-based Direct Dyes (Direct Blue 14)Lettuce (Lactuca sativa)Chronic
(7 days)
EC50 (growth) (soil)196–352 (263)Hulzebos et al. 1993
Benzidine-based Direct Dyes (Direct Blue 14)Lettuce (Lactuca sativa)Chronic
(14 days)
EC50 (growth) (soil)250–337 (290)Hulzebos et al. 1993


Abbreviations:
EC50, the concentration of a substance that is estimated to cause some effect on 50% of the test organisms;

Footnote Table 6-3 a

Robust study summaries were conducted to determine the quality of the study and are available upon request.

Return to footnote Table 6-3 a referrer

No toxicity data were found on the toxicity of Benzidine Derivatives in soil or sediment. However, a study conducted by Chung et al. (1998) on the toxicity of benzidine congeners on 18 bacterial strains, including nitrogen-fixing Azotobacter vinelandii, determined that TODI, 3,3′-DMOB and 4N-TMB were not inhibitory to any of the bacterial species tested.

6.1.3 Modelled Results

Although empirical ecotoxicity data for the Benzidine-based Dyes and Benzidine-based Precursors were limited, predictions of ecotoxicity using (Q)SAR models are considered unreliable. Anionic dye classes are difficult to model because the properties of these dyes fall outside the domains of applicability of the available models and because of the error associated with estimation of log Kow values used as input to the models.

However, due to their less complex chemical structure and a lower tendency to ionize, aquatic toxicity predictions for three Benzidine Derivatives, 3,3′-DMB, 3,3′-DMOB and 4N-TMB, were modelled using ECOSAR (2011), as they are considered to be within the model domain of applicability (Table 6-4).

Predictions for 4N-TMB were determined using the EVA corrected log Kow value of 3.53. Model predictions are consistent with the empirical results available for 3,3′-DMB and TODI, indicating that these Benzidine Derivatives are expected to be moderately toxic to algae, Daphnia and fish.

Table 6-4. Summary of modelled data for aquatic toxicity for the Benzidine Derivatives
SubstancesTest organismType of testEndpointValue (mg/L)Reference
3,3′-DMB and 3,3′-DMOBFishAcute (96 h)LC509.06–31.91ECOSAR 2011
3,3′-DMB and 3,3′-DMOBFishChronic (14 days)LC501.81–21.19ECOSAR 2011
3,3′-DMB and 3,3′-DMOBFishChronicChV0.017–0.092ECOSAR 2011
3,3′-DMB and 3,3′-DMOBDaphniaAcute (48 h)LC501.26–2.22ECOSAR 2011
3,3′-DMB and 3,3′-DMOBDaphniaChronicChV0.014–0.033ECOSAR 2011
3,3′-DMB and 3,3′-DMOBAlgaAcute (96 h)EC507.99–11.28ECOSAR 2011
3,3′-DMB and 3,3′-DMOBAlgaChronicChV4.55–6.87ECOSAR 2011
4N-TMBFishAcute (96 h)LC508.35ECOSAR 2011
4N-TMBFishChronic (14 days)LC508.65ECOSAR 2011
4N-TMBFishChronicChV0.96ECOSAR 2011
4N-TMBDaphniaAcute (48 h)LC505.90ECOSAR 2011
4N-TMBDaphniaChronicChV0.87ECOSAR 2011
4N-TMBAlgaAcute (96 h)EC505.44ECOSAR 2011
4N-TMBAlgaChronicChV2.68ECOSAR 2011

Abbreviations:
ChV, chronic ecotoxicity end-point value;
EC50, the concentration of a substance that is estimated to cause some effect on 50% of the test organisms;
LC50, the concentration of a substance that is estimated to be lethal to 50% of the test organisms

Since no experimental data were available for Benzidine-based Precursors and they are at most only weakly ionized in the environment, modelled data for the neutral form are presented in Table 6-5 for Naphthol AS-BR and Table 6-6 for TCDB.

Note that warnings were received from ECOSAR (2011), CPOPs (2008) and ACD/I-Lab (2010-2011) for the majority of model output values because most toxicity values were in excess of the minimum of the range of estimated water solubilities for Naphthol AS-BR (8.97 × 10−6 mg/L) and TCDB (0.26 mg/L). The baseline toxicity model was chosen for ECOSAR over other structure–activity relationships (SARs) such as Amides, Phenylamines and Polyphenols, since it has the most chemicals in the training set and widest range of log Kow values. This is important for these two substances, since they both have high modelled log Kow values near the edge of the model domains. Given that no experimental data are available and that the toxicities are predicted using modelled log Kow data, this information should be considered with a high degree of uncertainty in the overall weight of evidence for toxicity. This is demonstrated by the 5 orders of magnitude toxicity range for Naphthol AS-BR.

Table 6-5. Summary of modelled data for aquatic toxicity of Naphthol AS-BR using ACD/I-Lab (2010-2011), CPOPs (2008) and the baseline toxicity model from ECOSAR (2011)
Test organismType of test (duration)EndpointValue (mg/L)Reference
FishChronicChV0.000 51Footnote Table 6-5 [a]ECOSAR 2011
FishAcute (96 h)LC500.006[a]ECOSAR 2011
FishAcuteLC507.3[a],Footnote Table 6-5[b]ACD/I-Lab 2010–2011
FishAcuteLC500.29CPOPs 2008
DaphnidChronicChV0.0018[a]CPOPs 2008
DaphnidAcute (96 h)LC500.007[a]ECOSAR 2011
DaphnidAcuteLC500.000 89[a],Footnote Table 6-5[c]ECOSAR 2011
DaphnidAcuteEC500.19[a]ACD/I-Lab 2010–2011
DaphnidAcuteLC500.16[a]CPOPs 2008
Green algaChronicChV0.033[a]ECOSAR 2011
Green algaAcute (96 h)EC500.032[a]ECOSAR 2011


Abbreviations:
ChV, chronic ecotoxicity end-point value;
EC50, the concentration of a substance that is estimated to cause some effect on 50% of the test organisms;
LC50, the concentration of a substance that is estimated to be lethal to 50% of the test organisms

Footnote Table 6-5 a

Data in excess of minimum from range of predicted water solubilities for Naphthol AS-BR (8.97 × 10−6 mg/L).

Return to footnote Table 6-5 a referrer

Footnote Table 6-5 b

Not reliable in ACD/I-Lab (2010–2011)

Return to footnote Table 6-5 b referrer

Footnote Table 6-5 c

Borderline reliability in ACD/I-Lab (2010–2011)

Return to footnote Table 6-5 c referrer

Table 6-6. Summary of modelled data for aquatic toxicity of TCDB using ACD/I-Lab (2010–2011), CPOPs (2008) and the baseline toxicity model from ECOSAR (2011)
Test organismType of test (duration)EndpointValue (mg/L)Reference
FishChronicChV0.054ECOSAR 2011
FishAcute (96 h)LC500.61Footnote Table 6-6 [a]ECOSAR 2011
FishAcuteLC502.9[a],Footnote Table 6-6[b]ACD/I-Lab 2010–2011
FishAcuteLC50less than or equal to 0.31[a]CPOPs 2008
DaphnidChronicChV0.083ECOSAR 2011
DaphnidAcute (96 h)LC500.52[a]ECOSAR 2011
DaphnidAcuteLC500.026Footnote Table 6-6 [c]ACD/I-Lab 2010–2011
DaphnidAcute (48 h)EC50less than or equal to 0.87[a]CPOPs 2008
DaphnidAcute (48 h)LC50less than or equal to 0.19CPOPs 2008
Green algaChronicChV0.51[a]ECOSAR 2011
Green algaAcute (96 h)EC500.80[a]ECOSAR 2011


Abbreviations:
ChV, chronic ecotoxicity end-point value;
EC50, the concentration of a substance that is estimated to cause some effect on 50% of the test organisms;
LC50, the concentration of a substance that is estimated to be lethal to 50% of the test organisms

Footnote Table 6-6 a

Data in excess of minimum from range of predicted water solubilities for TCDB (0.26 mg/L).

Return to footnote Table 6-6 a referrer

Footnote Table 6-6 b

Not reliable in ACD/I-Lab (2010–2011)

Return to footnote Table 6-6 b referrer

Footnote Table 6-6 c

Borderline reliability in ACD/I-Lab (2010–2011)

Return to footnote Table 6-6 c referrer

6.1.4 Carcinogenicity and Mode of Action

Limited information is available on the environmental carcinogenicity of benzidine-based dyes. As mentioned previously, cleavage of the azo bond under anaerobic or reducing conditions (e.g., deep layers of sediments) is known to result in aromatic amines, some of which are known to be potentially carcinogenic. Studies using Direct Black 38 have shown breakdown to benzidine and 4-aminobiphenyl, which have been found to be mutagenic and carcinogenic substances (Isik and Sponza 2004; Bafana et al. 2009a; IARC 2010e) and acutely toxic (LC50 less than  1 mg/L) to some crustaceans and juvenile fish (Øllgaard et al. 1998). However, because they are formed only in deep anoxic sediment, there is a lower likelihood that aquatic organisms will be exposed to these more harmful metabolites. Limited information was available for terrestrial organisms. The potential genotoxicity and carcinogenic effects of Benzidine Derivatives to sediment-dwelling biota remain a source of uncertainty.

Limited data are available on the deoxyribonucleic acid (DNA) and/or protein binding of Benzidine-based Substances and Benzidine Derivatives in the environment. All 44 Benzidine-based Substances and Benzidine Derivatives were profiled for structural alerts using the OECD QSAR Toolbox for DNA interactions and protein binding. These mechanistic-based profilers identify the potential of substances to cause genotoxicity or protein binding–related effects (e.g., chromosomal aberrations, acute inhalation toxicity and acute aquatic toxicity). Possible DNA binding was identified through nitrenium ion formation for 33 (18 Benzidine-based Direct Dyes, 9 Benzidine-based Acid Dyes, 2 Benzidine-based Cationic Indicators and 4 Benzidine Derivatives) of the 44 Benzidine-based Substances. Five substances (2 Benzidine-based Acid Dyes, Benzidine-based 1 Direct Dye, 1 Benzidine-based Precursor and 1 Benzidine Derivative) have structural alerts suggesting that they are capable of directly acylating proteins resulting in adduct formation, while the remaining 39 substances showed no protein binding potential.

These profiling results indicate that 33 of these substances may exert additional toxicity (mostly DNA binding and in some cases protein binding) to the standard narcotic response. These data suggest only a potential for these adverse effects to occur and do not confirm that they will occur. Consequently, the profiling results will be considered as a line of evidence in determining the overall potential for adverse effects that these substances may elicit. However, the profiling results are consistent with the moderate, bordering on high, acute aquatic toxicity results observed for some of the subgroups addressed in this assessment. This suggests that given significant bioavailability in the environment, the reactivity of these compounds can result in acute aquatic toxicity above the baseline of narcosis (as correlated with log Kow). The potential genotoxicity and carcinogenicity of these substances for human relevance are addressed in the Potential to Cause Harm to Human Health section.

6.1.5 Derivation of the PNEC and Rationalization of the Assessment Factor

Benzidine-based Acid Dyes

Due to the paucity of toxicity data, a grouping approach was used in the development of the predicted no-effect concentration (PNEC) for the Benzidine-based Acid Dyes subgroup. Toxicity data for Acid Red 114 were chosen as read-across for Benzidine-based Acid Dyes since Acid Red 114 shares chemical structural similarity with other members of the group as well as common physical and chemical properties as part of the same application class and because reliable toxicity information was available.

The 96-hour LC50 of 4 mg/L for common carp (MITI 1992) was selected as the critical toxicity value (CTV) because it was the most sensitive valid experimental value. A PNEC was derived by dividing the CTV by an assessment factor of 100 (to account for interspecies and intraspecies variability in sensitivity and to estimate a long-term no-effects concentration from a short-term LC50) to give a value of 0.04 mg/L.

Benzidine-based Direct Dyes

A grouping approach was also used in the development of the PNEC for the Benzidine-based Direct Dyes subgroup. Direct Blue 14 was chosen as the surrogate or structural analogue for read-across, given that it is the most representative for the grouping (i.e., 70–100% structural similarity with most substances according to the OECD QSAR Toolbox), shares similar physical and chemical properties as part of the same application class and has the most toxicity information available.

The CTV selected was the 96-hour LC50 of 11.3 mg/L for freshwater shrimp (Desmocaris trispinosa; Ogugbue and Oranusi 2006), as this was the most sensitive valid experimental value. The aquatic PNEC was then derived by dividing this value by an assessment factor of 100 (to account for differences in interspecies and intraspecies variability and to estimate a long-term no-effects concentration from a short-term LC50). Therefore, a PNEC of 0.11 mg/L was calculated for the Benzidine-based Direct Dyes.

The soil CTV was the 7-day EC50 (growth) for lettuce (Lactuca sativa) of 263 mg/kg soil (Hulzebos et al. 1993), as this was the most sensitive value from the sole experimental study. The soil PNEC was then derived by dividing this value by an assessment factor of 100 (to account for interspecies and intraspecies variability in sensitivity and the fact that this is the only study available, and because the 7-day test length may be considered intermediate between short and long term for vascular plants). The result of the adjustment is a calculated value of 2.63 mg/kg soil.

This soil PNEC will also be used in the weight of evidence for Benzidine-based Acid Dyes, since there is a paucity of soil toxicity data and because direct and acid dyes share similar physical and chemical properties.

Benzidine-based Cationic Indicators

The aquatic CTV selected was the 96-hour LC50 of 5.6 mg/L for fathead minnow (Pimephales promelas) using the structural analogue BB4 (Little and Lamb 1973), as this was the most valid sensitive experimental value. A PNEC was derived by dividing the CTV by an assessment factor of 100 (to account for interspecies and intraspecies variability in sensitivity and to estimate a long-term no-effects concentration from a short-term 96-hour LC50) to give a value of 0.056 mg/L. This fish toxicity endpoint value was chosen for Benzidine-based Cationic Indicators since few data are available and there are similarities between BB4 and the Benzidine-based Cationic Indicators grouping due to their sharing of common functional groups and certain physical and chemical properties.

Benzidine-based Precursors

A PNEC for Naphthol AS-BR was derived from the chronic toxicity value of 0.00051 mg/L using the baseline toxicity model as the most sensitive for fish, which is within 1 order of magnitude of the substance’s mean predicted water solubility. An assessment factor was not used, since the CTV is a very conservative modelled chronic value that is driven by a very high modelled log Kow, which is also conservative.

A separate PNEC for TCDB was derived from the chronic modelled toxicity value of 0.083 mg/L for Daphnia as the most sensitive value that is below the lowest predicted water solubility. An assessment factor of 10 was used to account for interspecies and intraspecies variability in sensitivity, to give a PNEC value of 0.0083.

Benzidine Derivatives

Based on available experimental and modelled data, Benzidine Derivatives share similar ecological toxicity, which may be due to common modes of action (due to the presence of benzenamine or aniline groups). Therefore, the most sensitive valid experimental value for 3,3′-DMB was selected as the CTV and used as read-across to define the Benzidine Derivatives subgroup.

The CTV selected was the chronic 21-day EC50immobilization value of 0.64 mg/L for Daphnia (MITI 2000) for 3,3′-DMB. A PNEC was derived for the Benzidine Derivatives by dividing this value by an assessment factor of 10 (to account for interspecies and intraspecies variability in sensitivity) to yield a value of 0.064 mg/L.

6.1.5 Ecological Effects Summary

Based on lines of evidence involving empirical and read-across aquatic ecotoxicity data, it may be concluded that Benzidine-based Acid and Direct Dyes as well as Benzidine-based Cationic Indicators may be expected to cause harm to aquatic organisms at low concentrations. Based on limited empirical soil ecotoxicity data, Benzidine-based Acid and Direct Dyes as well as Benzidine-based Cationic Indicators are not expected to cause harm to soil-dwelling organisms at low concentrations.

In addition, based on various lines of evidence involving empirical and modelled ecotoxicity data in various environmental compartments, it may be concluded that Benzidine Derivatives may be expected to cause harm to aquatic organisms at low concentrations. Data for soil- and sediment-dwelling organisms are not available.

Finally, based on various lines of evidence involving empirical ecotoxicity data in various environmental compartments, it may be concluded that Benzidine-based Precursors may cause harm to aquatic organisms at low concentrations. Data for soil- and sediment-dwelling organisms are not available.

6.2 Ecological Exposure Assessment

6.2.1 Releases to the Environment

No measured concentrations of substances in the Benzidine-based Acid and Direct Dyes, Benzidine-based Cationic Indicators, Benzidine-based Precursors or Benzidine Derivatives subgroups in the Canadian environment have been identified. Environmental concentrations have therefore been estimated from available information.

Anthropogenic releases of a substance to the environment depend on various losses that occur during the manufacture, industrial use, consumer/commercial use and disposal of the substance. In order to estimate releases to the environment occurring at different stages of the life cycle of the substances evaluated in this Screening Assessment, Environment Canada compiled information on the sectors and product lines relevant to the substances. In addition, an effort was made to compile emission factorsFootnote[5] for emissions to wastewater, land and air at different life cycle stages. Relevant factors were considered, uncertainties were recognized and assumptions were made, when necessary, during each stage, depending on information available.

The information compiled was to give an overview of the potential losses occurring at different stages of the life cycle and the receiving media involved, as well as to identify the life cycle stages that are likely the largest contributors to the overall environmental concentrations. Recycling activities and transfer to waste disposal sites (landfill, incineration) were also considered. However, unless specific information on the rate of or potential for release from landfills and incinerators was available, releases to the environment from disposal were not quantitatively accounted for.

This information was used to further develop exposure scenarios for the purpose of determining Predicted environmental concentrations (PECs).

6.2.2 Ecological Exposure

The general methodology used for characterizing ecological exposure to the Benzidine-based Dyes is to evaluate the highest release sector or use first and then determine if it is necessary to provide the same level of detail for other sectors or uses. If the highest release sector or use is determined to present low ecological concern, then all other sectors or uses would be judged to also be of low concern due to their lower releases and would therefore not be analyzed further. Otherwise, these other sectors or uses will be examined to determine the extent of the ecological concern.

Benzidine-based Dyes are used in various sectors, based on reported uses in the literature (e.g., Hunger 2003); however, the dominant sector for Benzidine-based Dyes is the textile and leather sector, as discussed in the Uses section of this report.

The textile wet processing sector was selected as the scenario resulting in highest exposure for the assessment of the ecological risks posed by Benzidine-based Dyes. The textile sector was chosen, as it is expected to be the major use sector for Benzidine-based Dyes, and the fraction of dyes lost to wastewater during textile dyeing operations is higher than that from all other use sectors. The laundering of textile products as well as textile wet processing are identified as major uses. During laundering of textile products, dyes can be lost to wastewater. The loss is estimated to be 2.5–5% by dividing the maximum 10% loss over the lifetime of textile products (Øllgaard et al. 1998) by a typical life expectancy of 2–4 years for textile products (Michigan Institute of Laundering and Dry Cleaning 2012). The average loss to wastewater from textile wet processing is reported to be 10% for acid dyes and 12% for direct dyes (OECD 2004), significantly higher than the 2.5–5% loss from the laundering of textile products. Higher releases are therefore expected from the textile wet processing sector.

Based on the reported uses from surveys and the literature, the textile wet processing sector was determined to be an important use sector for the Benzidine-based Acid and Direct Dyes. A quantitative exposure assessment based on the textile wet processing sector was chosen for the Benzidine-based Acid Dyes, because this was the only reported use from industry surveys conducted for the years 2005 and 2006 (Canada 2006b, 2008b). Although these surveys did not reveal the specific application of the Benzidine-based Direct Dyes within the textile sector, literature data indicate that they exhibit high affinity for cellulose fibres and are applied directly to cellulose-containing textiles (ETAD 1995b; Hunger 2003).

Exposure Scenario #1: Aquatic Exposure via Potential Releases from Textile Wet Processing Mills

The approach for estimating aquatic exposure for Benzidine-based Acid and Direct Dyes is to focus on textile wet processing as the major release sector and to evaluate their aquatic PECs from individual sites.

In total, 145 textile wet processing mills were identified in a survey conducted by Environment Canada in 1999 (Crechem 1998). Those mills included six different types (knit, woven, stock/yarn, wool, carpet and non-woven) and eight facilities whose types were unknown. The majority of these mills discharged their wastewaters to municipal wastewater treatment systems before reaching the aquatic environment.

An effort was made to obtain site-specific conditions needed for aquatic exposure calculations. For this purpose, the 145 mills were examined together with their local wastewater treatment systems and receiving water bodies. For 75 mills, information was available on wastewater treatment effluent flow and receiving river flow. For the remaining 70 mills, there was a lack of accurate information on local municipal wastewater treatment systems or their receiving water bodies. The 75 mills, however, cover all major mill types (knit, woven, non-woven, stock/yarn, wool and carpet) and are believed to provide sufficient representation of the textile dyeing operations conducted by the sector.

The aquatic PECs for the 75 mills were determined at 33 sites, based on an estimated amount of the Benzidine-based Acid or Direct Dyes used. This estimated amount is associated with any given single mill and determined from the highest quantity sold to one single textile mill from the CEPA 1999 section 71 survey data.

Other parameters considered in the calculation of aquatic PECs include the amount of the Benzidine-based Acid or Direct Dyes released from textile dyeing operations and removed by wastewater treatment systems, as well as the dilution by the receiving water. Since the use and release of the Benzidine-based Acid or Direct Dyes are intermittent, additional dilution by lagoons is also considered. The lagoon dilution is warranted by the fact that lagoons have much longer hydraulic retention times than the release durations for the Benzidine-based Acid or Direct Dyes. This additional dilution is not considered valid for primary or secondary wastewater treatment systems due to their relatively short hydraulic retention times.

The aquatic PEC results for the 75 mills are presented in Table 12 in the section on Characterization of Ecological Risk, and the calculations are explained in Appendix B. The results show that the aquatic PECs range from 0.05 to 24 µg/L for the Benzidine-based Acid Dyes and from 0.06 to 20 µg/L for the Benzidine-based Direct Dyes. These values are considered conservative because they are based on a maximum quantity of the Benzidine-based Acid or Direct Dyes used at each single mill and zero removal from on-site wastewater treatment.

Exposure Scenario #2: Soil Exposure via Biosolids Application to Land

Soil exposure to the Benzidine-based Acid or Direct Dyes in this Screening Assessment was estimated using a conservative scenario. In this scenario, biosolids produced from wastewater treatment systems at the 33 sites evaluated for the aquatic exposure were assumed to be applied to land at the maximum allowable rate over a substantial number of years. The scenario also assumed that the Benzidine-based Acid or Direct Dyes accumulated in soil do not incur any losses from degradation, volatilization, soil runoff or leaching. This conservative scenario yielded a soil PEC of 0.18 mg/kg for the Benzidine-based Acid Dyes and 0.24 mg/kg for the Benzidine-based Direct Dyes (see Appendix C for detailed calculations).

6.3 Characterization of Ecological Risk

The approach taken in this ecological screening assessment was to examine various supporting information and develop conclusions based on a weight of evidence approach and using precaution as required under CEPA 1999. Lines of evidence considered include information on physical and chemical properties, environmental fate, ecotoxicity and sources of the substances, as well as results from conservative risk quotient analyses, which are outlined below.

6.3.1 Benzidine-based Acid and Direct Dyes

Aquatic Risk Quotient Analysis

A risk quotient analysis, integrating conservative estimates of exposure with toxicity information, was performed for the aquatic medium to determine whether there is potential for ecological harm in Canada. The analysis was conducted for a set of 75 mills at 33 sites and is considered representative of the textile sector.

Sector-based site-specific industrial scenarios (considering the actual receiving water body) show PECs of 0.05–24 µg/L for Benzidine-based Acid Dyes and 0.06–20 µg/L for Benzidine-based Direct Dyes (Environment Canada 2012). PNECs of 0.04 and 0.11 mg/L were derived for Benzidine-based Acid and Direct Dyes, respectively (see the Ecological Effects Assessment section). The resulting risk quotients (PEC/PNEC) are 0.0013–0.6 for Benzidine-based Acid Dyes and 0.00055–0.16 for Benzidine-based Direct Dyes. These results show low concern in the aquatic compartment from the use of the two groups of dyes in textile dyeing. Therefore, harm to aquatic organisms is unlikely at these sites, due in part to the large dilution capacities of the locations.

Soil Risk Quotient Analysis

Risk quotients for soil were also determined by dividing the PECs by the PNEC (2.63 mg/kg). The soil risk quotient for Benzidine-based Acid Dyes was 0.068, while the soil risk quotient for Benzidine-based Direct Dyes was 0.091. These risk quotients are conservative estimates, since many conservative assumptions were used in their derivation. The major conservative assumptions used include a) maximum quantities of acid or direct dyes based on the upper limit of each individual dye reported by industry via regulatory surveys; b) no deduction of the quantities of acid or direct dyes that are released to lagoons and do not end up in land-applied biosolids; and c) no loss assumed for the dyes present in soil via degradation, soil runoff or leaching over 10 years of biosolids application.

The PECs for the Benzidine-based Acid and Direct Dyes in water and soil show that they are well below the PNECs. Since all other sectors are expected to have lower releases, they are not likely to present ecological concern and are therefore not given further exposure analysis.

Benzidine-based Cationic Indicators, Benzidine-based Precursors and Benzidine Derivatives

No recent commercial data have been submitted for Benzidine-based Cationic Indicators, Benzidine-based Precursors or Benzidine Derivatives, and stakeholder interest has been declared for only one Benzidine Derivative. This is consistent with an understanding that the commercial use of benzidine-based dyes are also decreasing due to restrictions in several jurisdictions. While these substances may be used in commerce in Canada below survey reporting thresholds of 100 kg/year, or, in the case of the Benzidine Derivatives, transformation products may be formed through azo bond cleavage, environmental concentrations are expected to be very low and dispersive. While the estimated PNECs for these groups of substances differ, given the lack of data on commercial activity and the assumption that releases are low, no significant ecological risk from exposure is expected.

6.3.2 Discussion of Weight of Evidence and Conclusion of Ecological Risk Characterization

The approach taken in this ecological screening assessment was to examine various supporting information and develop conclusions based on a weight of evidence approach and using precaution as required under CEPA 1999. Lines of evidence considered include results from a conservative risk quotient calculation, as well as information on persistence, bioaccumulation, ecological effects, sources, fate of the substance and presence and distribution in the environment. Various lines of evidence for each subgrouping are summarized below, along with relevant uncertainties leading to overall conclusions.

Benzidine-based Acid and Direct Dyes are anthropogenically produced and not expected to occur naturally in the environment. The majority of the Benzidine-based Acid Dyes and Benzidine-based Direct Dyes in this grouping were not found to be in commerce according to recent surveys. No data concerning concentrations of these substances in the Canadian environment have been identified. Benzidine-based Acid and Direct Dyes are complex anionic molecules that have relatively high water solubility (greater than 1 g/L) and are expected to dissociate at environmentally relevant pH levels. Since there is a relative paucity of data, Benzidine-based Acid Dyes were grouped together due to their similar physical and chemical properties as well as relatively similar chemical structures (e.g., sharing common functional groups, but varying in number). Also, log Kow data from azo-based acid dyes were used as read-across for the Benzidine-based Acid Dyes. Due to their high water solubility and affinity for oppositely charged organic particles, Benzidine-based Acid and Direct Dyes are expected to be found in water, sediment and soil. Given very low expected vapour pressures and Henry’s Law constants, they are unlikely to stay in air if released to this compartment. Therefore, long-range atmospheric transport is not anticipated to be of concern. Given their hydrophilicity and charged character, Benzidine-based Acid and Direct Dyes have low experimental log Kow values (0.42 and 0.77, respectively). Estimated and experimental log Kow values were compared with experimental BCFs for fish for a number of dyes (Anliker et al. 1981; Øllgaard et al. 1998; ETAD 1995b). The reported BCF values for Benzidine-Based Acid and Direct Dyes were low, ranging from 42 to 84, indicating that these dyes are not likely to bioconcentrate in aquatic organisms. Furthermore, Benzidine-based Acid and Direct Dyes are not expected to bioconcentrate due to their high molecular weights ( greater than 600 g/mol) and relatively large minimum and maximum effective cross-sectional diameters, which suggest slow uptake potential. Bioaccumulation resulting from exposures of organisms to these substances in soil and sediment is not well understood due to minimal and limited data, in large part due to the high water solubility of these substances. According to empirical and modelled data, Benzidine-based Acid and Direct Dyes are expected to biodegrade very slowly in aerobic environments and are therefore considered to be persistent in water, sediment and soil. However, Benzidine-based Acid and Direct Dyes may degrade and transform to certain benzidine derivatives if they reach anaerobic environments. A source of uncertainty is the potential carcinogenicity and mutagenicity of these benzidine derivatives and their potential effects on sediment-dwelling biota. Based on lines of evidence involving empirical specific and read-across aquatic and terrestrial ecotoxicity data, it may be concluded that Benzidine-based Acid and Direct Dyes may be expected to cause harm to aquatic organisms at moderately low concentrations (4 and 11 mg/L, respectively). Toxicity data are limited for the terrestrial environment and unavailable for sediment-dwelling organisms. A conservative exposure analysis of the textile sector was done because that sector was anticipated to present the highest potential ecological risk related to industrial releases to the environment for these substances. Risk quotients calculated from PECs for Benzidine-based Acid and Direct Dyes released from textile wet processing mills and PNECs in the aquatic environment as well as risk quotients calculated from PECs for the application of biosolids containing Benzidine-based Acid and Direct Dyes to soil and PNECs in the soil environment show that at conservatively predicted levels of release, Benzidine-based Acid and Direct Dyes are not likely to result in significant aquatic or soil exposure. Overall, the results of this assessment lead to the conclusion that Benzidine-based Acid and Direct Dyes have a low potential to cause ecological harm in Canada.

Benzidine-based Cationic Indicators are also anthropogenically produced and not expected to occur naturally in the environment. Neither of the two Benzidine-based Cationic Indicators was identified to be in commerce above the reporting thresholds according to recent surveys, nor were Canadian monitoring data found. Benzidine-based Cationic Indicators also have relatively high water solubility (approximately 9–10 mg/L) and are expected to dissociate at environmentally relevant pH levels. These two substances were found to have similar physical and chemical properties and chemical structures, so any data available for either was shared. Also, read-across data from an azo-based cationic dye analogue and the substance TTC, which contains the tetrazolium chloride moiety, were used for ecotoxicity. Since Benzidine-based Cationic Indicators have a high water solubility and affinity for negatively charged organic particles, they are expected to be found in water, sediment and soil, but not air. Since Benzidine-based Cationic Indicators are charged, they have low experimental log Kow values. While BCFs were not available for these substances, they are similar enough to Benzidine-based Acid and Direct Dyes in their physical and chemical properties to indicate that these dyes are not likely to bioconcentrate in aquatic organisms. Data on terrestrial bioaccumulation and in sediment are minimal and limited, in large part due to the high water solubility of these substances. Benzidine-based Cationic Indicators are expected to biodegrade very slowly in aerobic environments and are therefore considered to be persistent in water, sediment and soil. Based on lines of evidence involving empirical specific and read-across aquatic and terrestrial ecotoxicity data, it may be concluded that Benzidine-based Cationic Indicators may be expected to cause harm to aquatic organisms at moderately low concentrations (pivotal value of 5.6 mg/L). Toxicity data are unavailable for the terrestrial environment and for sediment. However, given that Benzidine-based Cationic Indicators are not known to be in commerce in Canada, it is not believed that they would have a harmful effect on the environment.

The Benzidine-based Precursors Naphthol AS-BR and TCDB are anthropogenically produced, not expected to occur naturally in the environment and not found to be in commerce above the reporting thresholds in recent surveys. Naphthol AS-BR is sparingly soluble and not expected to dissociate under environmentally relevant pH levels, while TCDB is moderately soluble and is expected to dissociate readily. These two substances were addressed separately, since their physical and chemical properties as well as chemical structures are very different. Models were used to predict most results, given that read-across data from other substances could not be used, as no close analogues with data could be found. Since Naphthol AS-BR has very limited solubility, it is expected to reside mostly in soil or sediment if released to the environment, while TCDB is expected to stay in the water column or bind with particles, given its acidic character. Both Benzidine-based Precursors have moderate to high estimated log Kowvalues, and experimental BCFs were not available for these substances. Modelled aquatic BCFs were moderate to high for TCDB and Naphthol AS-BR.  Benzidine-based Precursors are expected to biodegrade very slowly in aerobic environments and are therefore considered to be persistent in water, sediment and soil. Based on lines of evidence involving two sets of modelled aquatic toxicity data for acute and chronic endpoints, it may be concluded that both TCDB and Naphthol AS-BR may be expected to cause harm to aquatic organisms at low concentrations (pivotal values less than 1 mg/L). Toxicity data are unavailable for the terrestrial environment and for sediment. However, given that TCDB and Naphthol AS-BR are not known to be in commerce in Canada, it is not likely that either substance would have a harmful effect on the environment.

Benzidine Derivatives are low molecular weight (212–285 g/mol) substances that contain biphenyl. These anthropogenic substances are not expected to occur naturally in the environment, and none were found to be in commerce above the reporting thresholds in Canada according to recent surveys. Benzidine Derivatives have primarily been used as intermediates for the synthesis of colorants, but may also be used to a lesser degree to manufacture other chemicals (HSDB 1983– ). No data concerning concentrations of these substances in the Canadian environment have been identified. The Benzidine Derivatives are generally moderately soluble (greater than 10 mg/L), have low to moderate log Kow values (1.7–3.5) and will become ionic at low pH levels, as indicated by their pKa values (3.4–4.7). Based on these properties, as well as their high potential for binding to particulate matter and sediment (see Environment Canada and Health Canada 2013), Benzidine Derivatives are expected to be found in water, sediment and soil. Some Benzidine Derivatives may be considered semi-volatile; however, they are unlikely to stay in air if released to this compartment. Therefore, long-range transport is not anticipated to be of concern. This is further confirmed by calculated persistence data indicating that Benzidine Deratives are expected to degrade rapidly in air (Meylan and Howard 1993; AOPWIN 2010. However, other empirical and modelled data indicate that Benzidine Derivatives biodegrade slowly in aerobic environments and are considered to be persistent in water, sediment and soil (Kawasaki 1980; MITI 1992; ECHA 2012). Moderate to high water solubility, low to moderate log Kow values as well as low empirical BCFs (4.8–47) for 3,3′-DMB (MITI 1992) indicate that Benzidine Derivatives will not bioconcentrate in aquatic organisms. Empirical and modelled data indicate that Benzidine Derivatives are moderately toxic to fish and algae ( greater than 1 mg/L), but may be highly toxic to invertebrates ( less than 1 mg/L) (Kuhn 1989; ECHA 2012; MITI 2000; ECOSAR 2011). However, given that releases of Benzidine Derivatives to the aquatic compartment as well as to other media are expected to be minimal based on their non-commercial status, it is not believed that Benzidine Derivatives would have a harmful effect on the environment.

6.3.3 Uncertainties

In general, with the exception of certain Benzidine Derivatives, substances addressed in this report had limited data available. As a result, a read-across approach using data from selected analogues was the best alternative to estimating physical and chemical properties. In the case of Benzidine-based Precursors, only modelled data could be used.

This paucity of information necessitated the generation of model predictions for biodegradation and inference of bioaccumulative potential using available data on physical and chemical properties. Certain empirical physical, chemical and toxicological data were generated with formulated products. Therefore, when possible, available data from relevant analogues were also used to inform the read-across groupings. Long-term (chronic) toxicity data would be beneficial in evaluating these substances due to the fact that they are predicted to be persistent in the environment. The use of assessment factors in determining a PNEC is intended to address these uncertainties. While the soil and sediment exposure media were found to be important for Benzidine-based Acid and Direct Dyes, as well as Benzidine-based Cationic Indicators, effects data were not generally available.

The lack of measured environmental concentrations of these substances (e.g., monitoring data) in Canada resulted in the need to evaluate risk based on predicted concentrations in water near industrial point sources as well as sediment and soil. Conservative assumptions were made when using models to estimate concentrations in receiving water bodies, sediment and soil.

Given the use of some of these substances in other countries, it is possible that they may enter the Canadian market as components of manufactured items and/or consumer products. However, it is anticipated that the proportions of these substances released to the various environmental media would not be significantly different from those estimated here, given the conservative assumptions used in the textile sector exposure analysis.


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