necessarily represent the decisions or the stated policy of the United Nations Environment Programme, the International Labour Organisation, or the World Health Organization.
Concise International Chemical Assessment Document 15
1,2-DIAMINOETHANE (ETHYLENEDIAMINE)
First draft prepared by
Mr R. Cary, Health and Safety Executive, Merseyside, United Kingdom,
Dr S. Dobson, Institute of Terrestrial Ecology, Cambridgeshire, United Kingdom, and Dr J. Delic, Health and Safety Executive, Merseyside, United Kingdom
Please note that the layout and pagination of this pdf file are not identical to those of the printed CICAD
Published under the joint sponsorship of the United Nations Environment Programme, the International Labour Organisation, and the World Health Organization, and produced within the framework of the Inter-Organization Programme for the Sound Management of Chemicals.
World Health Organization
Geneva, 1999
and the World Health Organization (WHO). The overall objectives of the IPCS are to establish the scientific basis for assessment of the risk to human health and the environment from exposure to chemicals, through international peer review processes, as a prerequisite for the promotion of chemical safety, and to provide technical assistance in strengthening national capacities for the sound management of chemicals.
The
Inter-Organization Programme for the Sound Management of Chemicals (IOMC) wasestablished in 1995 by UNEP, ILO, the Food and Agriculture Organization of the United Nations, WHO, the United Nations Industrial Development Organization, the United Nations Institute for Training and Research, and the Organisation for Economic Co-operation and Development (Participating Organizations), following recommendations made by the 1992 UN Conference on Environment and Development to strengthen cooperation and increase coordination in the field of chemical safety. The purpose of the IOMC is to promote coordination of the policies and activities pursued by the Participating Organizations, jointly or separately, to achieve the sound management of chemicals in relation to human health and the environment.
WHO Library Cataloguing-in-Publication Data 1,2-Diaminoethane (Ethylenediamine).
(Concise international chemical assessment document ; 15) 1.Ethylenediamines 2.Environmental exposure 3.Risk assessment I.International Programme on Chemical Safety II.Series
ISBN 92 4 153015 4 (NLM classification: QV 275) ISSN 1020-6167
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iii
FOREWORD . . . 1
1. EXECUTIVE SUMMARY . . . 4
2. IDENTITY AND PHYSICAL/CHEMICAL PROPERTIES . . . 5
3. ANALYTICAL METHODS . . . 5
4. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE . . . 6
5. ENVIRONMENTAL TRANSPORT, DISTRIBUTION, AND TRANSFORMATION . . . 6
6. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE . . . 7
6.1 Environmental levels . . . 7
6.2 Human exposure . . . 7
7. COMPARATIVE KINETICS AND METABOLISM IN LABORATORY ANIMALS AND HUMANS . . . 8
8. EFFECTS ON LABORATORY MAMMALS AND IN VITRO TEST SYSTEMS . . . 9
8.1 Single exposure . . . 9
8.2 Irritation and sensitization . . . 9
8.3 Short-term exposure . . . 9
8.4 Long-term exposure . . . 10
8.4.1 Subchronic exposure . . . 10
8.4.2 Chronic exposure and carcinogenicity . . . 10
8.5 Genotoxicity and related end-points . . . 10
8.6 Reproductive and developmental toxicity . . . 11
8.7 Immunological and neurological effects . . . 11
9. EFFECTS ON HUMANS . . . 11
10. EFFECTS ON OTHER ORGANISMS IN THE LABORATORY AND FIELD . . . 14
11. EFFECTS EVALUATION . . . 14
11.1 Evaluation of health effects . . . 14
11.1.1 Hazard identification and dose–response assessment . . . 14
11.1.2 Criteria for setting guidance values for EDA . . . 15
11.1.3 Sample risk characterization . . . 16
11.2 Evaluation of environmental effects . . . 16
12. PREVIOUS EVALUATIONS BY INTERNATIONAL BODIES . . . 18
13. HUMAN HEALTH PROTECTION AND EMERGENCY ACTION . . . 18
13.1 Human health hazards . . . 18
13.2 Advice to physicians . . . 18
13.3 Health surveillance advice . . . 18
13.4 Spillage . . . 18
iv
14. CURRENT REGULATIONS, GUIDELINES, AND STANDARDS . . . 18
INTERNATIONAL CHEMICAL SAFETY CARD . . . 19
REFERENCES . . . 21
APPENDIX 1 — SOURCE DOCUMENTS . . . 25
APPENDIX 2 — CICAD PEER REVIEW . . . 25
APPENDIX 3 — CICAD FINAL REVIEW BOARD . . . 26
RÉSUMÉ D’ORIENTATION . . . 27
RESUMEN DE ORIENTACIÓN . . . 29
FOREWORD
Concise International Chemical Assessment Documents (CICADs) are the latest in a family of publications from the International Programme on Chemical Safety (IPCS) — a cooperative programme of the World Health Organization (WHO), the International Labour Organisation (ILO), and the United Nations Environment Programme (UNEP). CICADs join the Environmental Health Criteria documents (EHCs) as authoritative documents on the risk assessment of chemicals.
CICADs are concise documents that provide summaries of the relevant scientific information concerning the potential effects of chemicals upon human health and/or the environment. They are based on selected national or regional evaluation documents or on existing EHCs. Before acceptance for publication as CICADs by IPCS, these documents undergo extensive peer review by internationally selected experts to ensure their completeness, accuracy in the way in which the original data are represented, and the validity of the conclusions drawn.
The primary objective of CICADs is characterization of hazard and dose–response from exposure to a chemical. CICADs are not a summary of all available data on a particular chemical; rather, they include only that information considered critical for characterization of the risk posed by the chemical. The critical studies are, however, presented in sufficient detail to support the conclusions drawn. For additional information, the reader should consult the identified source documents upon which the CICAD has been based.
Risks to human health and the environment will vary considerably depending upon the type and extent of exposure. Responsible authorities are strongly encouraged to characterize risk on the basis of locally measured or predicted exposure scenarios. To assist the reader, examples of exposure estimation and risk characterization are provided in CICADs, whenever possible. These examples cannot be considered as representing all possible exposure situations, but are provided as guidance only. The reader is referred to EHC 1701 for advice on the derivation of health-based guidance values.
While every effort is made to ensure that CICADs represent the current status of knowledge, new
information is being developed constantly. Unless otherwise stated, CICADs are based on a search of the scientific literature to the date shown in the executive summary. In the event that a reader becomes aware of new information that would change the conclusions drawn in a CICAD, the reader is requested to contact IPCS to inform it of the new information.
Procedures
The flow chart shows the procedures followed to produce a CICAD. These procedures are designed to take advantage of the expertise that exists around the world — expertise that is required to produce the high- quality evaluations of toxicological, exposure, and other data that are necessary for assessing risks to human health and/or the environment.
The first draft is based on an existing national, regional, or international review. Authors of the first draft are usually, but not necessarily, from the institution that developed the original review. A standard outline has been developed to encourage consistency in form.
The first draft undergoes primary review by IPCS to ensure that it meets the specified criteria for CICADs.
The second stage involves international peer review by scientists known for their particular expertise and by scientists selected from an international roster compiled by IPCS through recommendations from IPCS national Contact Points and from IPCS Participating Institutions. Adequate time is allowed for the selected experts to undertake a thorough review. Authors are required to take reviewers’ comments into account and revise their draft, if necessary. The resulting second draft is submitted to a Final Review Board together with the reviewers’ comments.
The CICAD Final Review Board has several important functions:
– to ensure that each CICAD has been subjected to an appropriate and thorough peer review;
– to verify that the peer reviewers’ comments have been addressed appropriately;
– to provide guidance to those responsible for the preparation of CICADs on how to resolve any remaining issues if, in the opinion of the Board, the author has not adequately addressed all comments of the reviewers; and
– to approve CICADs as international assessments.
Board members serve in their personal capacity, not as representatives of any organization, government, or industry. They are selected because of their expertise in human and environmental toxicology or because of their
1 International Programme on Chemical Safety (1994) Assessing human health risks of chemicals: derivation of guidance values for health-based exposure limits.
Geneva, World Health Organization (Environmental Health Criteria 170).
S E L E C T I O N O F H I G H Q U A L I T Y N A T I O N A L / R E G I O N A L A S S E S S M E N T D O C U M E N T ( S )
CICAD PREPARATION FLOW CHART
F I R S T D R A F T P R E P A R E D
REVIEW BY IPCS CONTACT POINTS/
SPECIALIZED EXPERTS
FINAL REVIEW BOARD 2
FINAL DRAFT 3
EDITING
APPROVAL BY DIRECTOR, IPCS
PUBLICATION SELECTION OF PRIORITY CHEMICAL
1 Taking into account the comments from reviewers.
2 The second draft of documents is submitted to the Final Review Board together with the reviewers’ comments.
3 Includes any revisions requested by the Final Review Board.
REVIEW OF COMMENTS (PRODUCER/RESPONSIBLE OFFICER), PREPARATION
OF SECOND DRAFT 1 P R I M A R Y R E V I E W B Y I P C S (REVISIONS AS NECESSARY)
experience in the regulation of chemicals. Boards are chosen according to the range of expertise required for a meeting and the need for balanced geographic
representation.
Board members, authors, reviewers, consultants, and advisers who participate in the preparation of a CICAD are required to declare any real or potential conflict of interest in relation to the subjects under discussion at any stage of the process. Representatives of nongovernmental organizations may be invited to observe the proceedings of the Final Review Board.
Observers may participate in Board discussions only at the invitation of the Chairperson, and they may not participate in the final decision-making process.
1. EXECUTIVE SUMMARY
This CICAD on 1,2-diaminoethane (ethylenedi- amine) was based on a review of human health concerns (primarily occupational, but also including an environ- mental assessment) prepared by the United Kingdom’s Health and Safety Executive (Brooke et al., 1997). Data identified up to the end of 1994 were covered in the origi- nal review. An additional literature search up to July 1997 was conducted to identify any new information that had been published since the review was completed.
Information on environmental fate and effects was based on the report of the German Chemical Society’s Adviso- ry Committee on Existing Chemicals of Environmental Relevance (BUA, 1997). The preparation and peer review of the source documents are described in Appendix 1.
Information on the peer review of this CICAD is presented in Appendix 2. This CICAD was approved as an international assessment at a meeting of the Final Review Board, held in Tokyo, Japan, on 30 June – 2 July 1998. Participants at the Final Review Board meeting are listed in Appendix 3. The International Chemical Safety Card (ICSC 0269) produced by the International Pro- gramme on Chemical Safety (IPCS, 1993) has also been reproduced in this document.
1,2-Diaminoethane (CAS No. 107-15-3), commonly known as ethylenediamine (EDA), is a synthetic colour- less to yellowish liquid at normal temperature and pressure. It is strongly alkaline and is miscible with water and alcohol. The main use for EDA is as an intermediate in the manufacture of tetraacetyl ethylenediamine, ethylenediaminetetraacetic acid (EDTA), organic flocculants, urea resins, and fatty bisamides. It is also used, to a much smaller extent, in the production of for- mulations for use in the printed circuit board and metal finishing industries, as an accelerator/curing agent in epoxy coatings/resins, and in the manufacture of phar- maceutical products. EDA is present as a contaminant (<0.5%) in commercially supplied fatty amines, which are used as wetting agents in bituminous emulsions. It is also used in the synthesis of carbamate fungicides, in surfactant and dye manufacture, and in photography development chemicals and cutting oils. EDA is a degra- dation product of ethylenebis(dithiocarbamate) fungi- cides.
No atmospheric effects are expected, as reaction of EDA with hydroxyl radicals is likely to be rapid (half-life 8.9 h), and washout of volatilized EDA is expected.
Volatilization to the atmosphere is likely from soil but not from water. Adsorption to soil particulates is strong through electrostatic binding; leaching through soil profiles to groundwater is not expected. Complex formation with metals and humic acids is expected. Bio- degradation is the most likely source of breakdown in the environment and should be quite rapid; adaptation
of microorganisms may improve degradation. Breakdown is less rapid in seawater than in fresh water. Bioaccumu- lation is unlikely.
EDA has moderate acute toxicity in animals. It is a primary irritant, being corrosive when undiluted, and is also a skin sensitizer. EDA has not been tested for muta- genicity to current regulatory standards, and there are no assays for clastogenic activity or for the potential to express activity in somatic cells in vivo. Thus, there is insufficient information to draw firm conclusions regard- ing the mutagenic potential of EDA. EDA was not carcinogenic in animals. Non-neoplastic effects on the liver (pleomorphic changes to hepatocytes) have been observed in rats following oral dosing for 2 years at 45 mg EDA/kg body weight per day and above, with no effects seen at 9 mg EDA/kg body weight per day.
Although the significance of these hepatic cell changes for human health is unclear, as well as whether or not they are a consequence of oral exposure (i.e., they might not occur via other routes, as they may be related to first-pass effects), they cannot be discounted, and the risk of their development should be characterized. In oral gavage dosing studies, effects on the rat eye (retinal atrophy and, at higher doses, cataract formation) were observed at doses of 100 mg EDA/kg body weight per day and above. Doses of 200 and 100 mg EDA/kg body weight per day and above were associated with renal damage in rats and mice, respectively. There was also some indication of effects in the spleen in mice and rats at doses of 400 mg EDA/kg body weight per day and above and in the thymus in rats at 800 mg/kg body weight per day. In inhalation studies, no effects were seen in rats at about 150 mg/m3 (60 ppm), and slight depilation was the only treatment-related effect observed at about 330 mg/m3 (132 ppm).
Because diluted EDA is a skin irritant and a skin sensitizer, there may be a risk of developing irritant and/or allergic dermatitis if suitable personal protective equipment is not used in the occupational environment where skin contact can occur. EDA is also capable of inducing a state of respiratory tract hypersensitivity and provoking asthma in the occupational environment, and this is considered to be the major health effect of con- cern.
The mechanism for the induction of the hypersen- sitive state is not proven, although the skin sensitizing potential of EDA and the limited evidence of immuno- logical involvement in workers with EDA-provoked asthma are suggestive of an immunological mechanism.
However, irrespective of the mechanism involved, the available data do not allow either elucidation of dose–
response relationships or identification of the thresholds for induction of the hypersensitive state or provocation of an asthmatic response. The sample risk characteriza- tion in this document has, in order to assess the risks of
other systemic effects, evaluated the risk of hepatic effects in occupationally exposed individuals. It con- cludes that when EDA is used in closed systems, the exposure, both measured and predicted from models, is substantially (by 100-fold or greater) less than the no- observed-effect level (NOEL) in rats; thus, adverse effects on the liver are unlikely.
Exposure of the general public to EDA could not be evaluated owing to the lack of available data.
Toxic thresholds for microorganisms may be as low as 0.1 mg EDA/litre. However, toxicity tests in cul- ture media should be treated with caution, as the EDA may complex with metal ions. Effects may therefore be indirect, resulting from the loss of bioavailability of essential elements. LC50s for invertebrates and fish range from 14 to >1000 mg/litre. A no-observed-effect concen- tration (NOEC) for Daphnia reproduction has been reported at 0.16 mg/litre.
Given the wide range of acute and chronic test results, a predicted no-effect concentration (PNEC) for aquatic organisms was taken as 16 :g/litre, based on application of an uncertainty factor of 10 to the lowest reported NOEC for Daphnia reproduction. Conservative assumptions for predicted environmental concentration (PEC) produce PEC/PNEC ratios indicating some concern from initial concentrations (i.e., at first release into the river or estuary). However, more refined exposure estimates indicate low risk to aquatic organisms.
2. IDENTITY AND PHYSICAL/CHEMICAL PROPERTIES
1,2-Diaminoethane (CAS No. 107-15-3) is more commonly known as ethylenediamine, with EDA used as a common abbreviation. Other common synonyms include dimethylenediamine, 1,2-ethanediamine, 1,2- ethylenediamine, beta-aminoethylamine, and ethane-1,2- diamine. EDA’s structural formula is shown below:
EDA is a colourless to yellowish hygroscopic liquid with an ammonia-like odour. Its molecular weight is 60.12. It is a strongly alkaline (pH of 25% EDA in water is 11.9), very volatile, pungent material, which fumes profusely in air. It has a melting point of about 8.5 °C, a boiling point of 116 °C (at 101.3 kPa), and a vapour pressure of 1.7 kPa at 25°C. EDA is miscible with water
and alcohol. The log octanol/water partition coefficient (log Kow) ranges from !1.2 to !1.52. pKa1 and pKa2 (calculated) are 10.71 and 7.56, respectively, indicating protonation at environmentally relevant pH. Additional physical/chemical properties are presented in the International Chemical Safety Card reproduced in this document.
Conversion factors for EDA at 20 °C and 101.3 kPa are as follows:
1 ppm = 2.50 mg/m3 1 mg/m3 = 0.40 ppm
3. ANALYTICAL METHODS
For monitoring concentrations of EDA in work- place air, NIOSH (1984–1989) uses a method that employs adsorption on silica gel and analysis by gas chromatography with flame ionization detection. A solvent-free sampling system is preferable because of more convenient handling, and it is a great advantage if derivatization can be achieved directly on the absorbent.
The Health and Safety Laboratories of the United King- dom’s Health and Safety Executive have evaluated a published method (Andersson et al., 1985; Levin et al., 1989; Patel & Rimmer, 1996). Air is sampled onto 1- naphthyl-isothiocyanate-impregnated filters, desorbed by acetonitrile, and analysed by high-performance liquid chromatography with ultraviolet detection. The method has a working range between 2.5 and 50 mg/m3 for a 5-litre air sample. The detection limit was found to be 0.08 mg/m3. The method generally meets the Comité Européen de Normalisation requirements on the overall uncertainty. Although the Comité Européen de Normali- sation requirements for desorption efficiency were not satisfied at 25 and 50 mg/m3, a smaller sample can be taken if necessary.
There are no reported methods for the biological monitoring of occupational exposure to EDA. However, analytical techniques based on solvent extraction of EDA and high-performance liquid chromatography have been reported and used in pharmacological studies (Cotgreave & Caldwell, 1983c), and these might form the basis for biological monitoring methods.
EDA can be measured in water using reverse- phase high-performance liquid chromatography with ultraviolet detection at 315 nm, following derivatization with acetylacetone. The limit of detection was reported to be 0.26 :g/litre (Nishikawa, 1987).
N C H H
C N H H H
H H
H
4. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE
EDA is not known to occur naturally. The main use for EDA is as an intermediate in the manufacture of tetraacetyl ethylenediamine, EDTA, organic flocculants, urea resins, and fatty bisamides. It is also used, to a much smaller extent, in the production of formulations for use in the printed circuit board and metal finishing industries, as an accelerator/curing agent in epoxy coat- ings/resins, and in the manufacture of pharmaceutical products. EDA is also present as a contaminant (<0.5%) in commercially supplied fatty amines, which are used as wetting agents in bituminous emulsions. It is also used in the synthesis of carbamate fungicides, in surfactant and dye manufacture, and in photography development chemicals and cutting oils. These are believed to be minor uses in the United Kingdom and were not inves- tigated in this review. EDA is a degradation product of ethylenebis(dithiocarbamate) fungicides.
Approximately 11 000 tonnes of EDA are imported into the United Kingdom each year, with very little being re-exported (Brooke et al., 1997). World production amounts to 100 000–500 000 tonnes annually.1 In 1992, annual production capacities were 18 000 tonnes for Germany, 54 000 tonnes for the Netherlands, 30 000 tonnes for Belgium, 25 000 tonnes for Sweden, about 159 000 tonnes for the United States, and 15 000 tonnes for Japan (BUA, 1997).
No measured concentrations of EDA in wastewater streams from manufacture and use are available.
However, estimates of EDA entering waste treatment from four European manufacturing plants were 200, 287, 5000–10 000, and 1000 kg/year. Use in photochemicals was estimated to lead to 1.1 tonnes being introduced into municipal sewage treatment plants in Germany. All figures are for 1992 or 1993 (BUA, 1997).
5. ENVIRONMENTAL TRANSPORT, DISTRIBUTION, AND TRANSFORMATION
Few experimental data are available on the distri- bution, transport, or fate of EDA in the environment.
However, qualitative, and some quantitative, estimates have been made on the basis of its physicochemical properties.
EDA has a moderately high vapour pressure and is expected to volatilize from soil (HSDB, 1997). In the atmosphere, it should react rapidly with photochemically produced hydroxyl radicals; no experimental rates are available for this proposed reaction, but a half-life of 8.9 h has been calculated.1 EDA may react with carbon dioxide to form an insoluble carbamate. The high water solubility of EDA means that volatilized chemical is also likely to be washed out by rain.2 The calculated dimen- sionless Henry’s law constant (air/water partition coeffi- cient) is extremely low (7.08 × 10–8); therefore, little evaporation would be expected from water. A half-life for volatilization of 45 years was estimated for a model river 1 m deep.2 An approximate Henry’s law constant is given in BUA (1997) as 1.77 × 10–4 PaAm3/mol.
Photodegradation is not expected, as the molecule contains no chromophores, which absorb radiation (HSDB, 1997).
Despite their miscibility with water, ethylene- amines can bind strongly to soil. There was a wide range of determined adsorption coefficients in experimental studies on six soil types (Table 1). Some reduction in variability occurred when results were normalized for organic carbon content, although this was less marked with EDA than with the other ethyleneamine studied.
Sorption to soil was rapid, with equilibrium occurring within a few hours. Electrostatic interaction between the positively charged ethyleneamine and negatively charged soils appeared to be the dominant factor in binding. Complex formation with metals and humic acids is expected. Sorption is greater to soils with high cation exchange capacity (Davis, 1993).
EDA at 200 mg/litre was incubated with adapted sewage sludge until there was no further decrease in chemical oxygen demand (COD); at that time (unspeci- fied), 97.5% of the chemical had been degraded. The rate of degradation was 9.8 mg COD/g per hour (Pitter, 1976).
EDA at 3, 7, and 10 mg/litre was incubated with sewage sludge (adapted and non-adapted), and percent biodegradation was determined 5, 10, 15, and 20 days later. Degradation rates were comparable for adapted and non-adapted sludge up to 15 days (at 56% and 55%, respectively); at 20 days, however, the values were 70%
and 47%, respectively. Based on this single point, it is not possible to conclude definitively that adaptation improves degradation. Nitrate and nitrite were measured throughout the incubation to correct for oxygen demand due to conversion of ammonia or organic nitrogen to these species. Such a correction was necessary for EDA
1 IUCLID (European Union database), 1st ed., 1996.
2 Syracuse Research Corporation modelling, summarized in HSDB (1997).
Table 1: Sorption of EDA to various soil types.a
Soil type pH
Cation exchange capacity (meq/100 g)
Fraction of organic carbon (Foc)b
Freundlich adsorption coefficient (Kd)b
Adsorption coefficient normalized for organic carbon (K = Kd/Foc)c
Sandy loam (Londo) 7.2 9.2 0.026 69 2700
Sandy clay loam 7.3 16.4 0.039 220 5600
Sandy loam (Cecil) 6.0 3.0 0.014 29 2100
Silty loam 6.0 15.6 0.034 238 7100
Clay 7.9 11.9 0.014 70 5000
Aquifer sand 9.6 6.9 0.0024 15 6200
a Data from Davis (1993).
b Values rounded.
c Mean 4800 ± 2000 (SD).
alone out of more than 50 compounds tested. Degrada- tion was also tested in a salt-water system using non- adapted sludge; EDA was degraded less effectively, with 16% of theoretical degradation after 20 days (Price et al., 1974). A comparable value at 16.6% was measured in seawater by Takemoto et al. (1981). EDA incubated with microorganisms isolated from river water and adapted to the compound over 28 days showed >80%
degradation relative to theoretical oxygen demand over 10 days (Mills & Stack, 1955).
Brief descriptions of the following degradation tests were also identified. EDA incubated with activated sludge at 100 mg/litre for 28 days showed 93–95% degra- dation relative to theoretical oxygen demand in a modified Ministry of International Trade and Industry (MITI) test (Japan Chemical Industry Ecology- Toxicology Information Center, 1992). Incubation with activated sludge at a concentration of 50 mg/litre led to degradation of 10%, 87.5%, and 94% after 5, 15, and 28 days, respectively.1
The high water solubility and low octanol/water partition coefficient indicate that bioaccumulation in organisms is unlikely.
6. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE
6.1 Environmental levels
There are no reports of monitoring of EDA levels in the aquatic environment or of measurements in effluents.
Residues of EDA in soil, 15 days post-treatment with the fungicide maneb, have been reported at
0.119 mg/kg for the top 1 cm (approximately) and at 0.044 mg/kg down to about 5 cm. Residues on tomatoes and beans were 0.053 and 0.239 mg/kg, respectively, immediately after spraying, falling to 0.047 and
0.094 mg/kg, respectively, after 14 days (Newsome et al., 1975).
6.2 Human exposure
The data available to the authors of this document are restricted mainly to the occupational environment.
The exposure assessments used in this report are based on either limited data or data modelled using the Esti- mation and Assessment of Substance Exposure (EASE) model. This is a general-purpose predictive model developed by the United Kingdom’s Health and Safety Executive for exposure assessment in the workplace. In its present form, the model is in widespread use across the European Union for the occupational exposure assessment of new and existing substances. Similarly, information on control measures has been derived from United Kingdom industry sources. Where data gaps exist, professional judgement has been used.
The number of employees exposed to EDA in the United Kingdom is not accurately known. For use as an intermediate in the manufacture of tetraacetyl ethylene- diamine, EDTA, organic flocculants, urea resins, and fatty bisamides, it is estimated that 140 employees will be potentially exposed. During the production of formu- lations for use in the printed circuit board and metal finishing industries and in the manufacture of epoxy coatings/resins and pharmaceutical products, it is esti- mated that 200 employees will be regularly exposed to EDA. The number of employees potentially exposed from use of EDA-based formulations in the printed circuit board and metal finishing industries is estimated to be about 100. EDA can also be released when indus- trial epoxy coatings/adhesives are applied, and this
1 Unpublished report from Akzo Research to Delamine, 1989 (cited in IUCLID).
activity has the potential to expose several thousand employees across a wide range of industries.
There are very few measured occupational expo- sure data available. EDA’s use as an intermediate in chemical synthesis takes place in closed systems. Meas- ured exposures for these manufacturing processes show that control is achieved to a level of less than 1.25 mg/m3 (0.5 ppm) 8-h time-weighted average (Hansen et al., 1984). Modelled data (EASE) are in good agreement, predicting comparable values of 0.53–1.3 mg/m3 (0.21–
0.52 ppm). Short-term peak exposures (sampling and hose uncoupling operations) were predicted to range between 16.8 and 33.3 mg/m3 (6.7 and 13.3 ppm), 15-min time-weighted average.
EDA’s use in the production of formulations usu- ally takes place in well-ventilated enclosed systems.
Measured exposure data are not available for these proc- esses. However, modelled exposure data indicate expo- sure levels of 5–20 mg/m3 (2–8 ppm) 8-h time-weighted average in the presence of local exhaust ventilation and 38–75 mg/m3 (15–30 ppm) 8-h time-weighted average in the absence of local exhaust ventilation. Corresponding short-term peak exposures during mixer charging oper- ations were estimated to be 5–25 mg/m3 (2–10 ppm) 15-min time-weighted average in the presence of local exhaust ventilation and 50–103 mg/m3 (20–41 ppm) 15-min time-weighted average in the absence of local exhaust ventilation.
The potential for exposure during the use of EDA formulations will be moderated by the low concentration of EDA present in the formulations. Very few exposure data are available, and there is scope for widely different use scenarios. There will be no appreciable occupational exposure if these products are used in enclosed venti- lated systems as indicated by measured exposure data (<2.5 mg/m3 [<1 ppm] 8-h time-weighted average) and modelled exposure data (0–0.25 mg/m3 [0–0.1 ppm] 8-h time-weighted average). Modelled exposure data for immersion processes, in the presence of local exhaust ventilation, predicted inhalation exposures of 0.5–2.5 mg/
m3 (0.2–1 ppm) 8-h time-weighted average. The potential for greatest inhalation exposure was predicted for situations where these formulations are brushed in open systems with only general dilution ventilation or sprayed in open systems in the presence of local exhaust ventilation. Under these conditions, modelled exposure data predict exposures in the range 2.5–5 mg/m3 (1–
2 ppm) 8-h time-weighted average. Short-term peak exposures during mixing and loading operations were estimated to be 5–10 mg/m3 (2–4 ppm) 15-min time- weighted average. Polyamines and alkanol polyamines, including EDA, have been reported to be released from hot bitumen during road paving (Levin et al., 1994). EDA concentrations generated during road paving were below 0.025 mg/m3 (0.01 ppm).
There will also be a potential for dermal exposure across the full range of industries handling EDA. Mod- elled data estimate dermal exposures in the range 0–0.15 mg/cm2 per day. However, the use of personal protective equipment is standard practice in all indus- tries using EDA. Therefore, in practice, dermal exposure will be considerably reduced by the use of personal protective equipment.
7. COMPARATIVE KINETICS AND METABOLISM IN LABORATORY ANIMALS
AND HUMANS
The toxicokinetics of EDA has received only limited study, and there are no studies following inhalation exposure. Studies in humans have been related to the clinical application of EDA and have demonstrated rapid absorption via the gastrointestinal tract, with at least 50% absorbed within the first 7 h;
absorbed EDA is rapidly removed from the plasma (Caldwell & Cotgreave, 1983; Cotgreave & Caldwell, 1983a,b, 1985). At least half the amount absorbed is excreted in the urine, largely as the acetylated metabolite N-acetylethylenediamine and, in smaller amounts, as the unchanged compound.
This toxicokinetic picture is supported and extend- ed by data from studies in experimental animals. Studies in rats and mice have demonstrated rapid and extensive uptake via the oral route and also via the respiratory tract following intratracheal instillation (about 70% or more of the applied dose was absorbed within 48 h) (McKelvey et al., 1982; Yang & Tallant, 1982; Yang et al., 1984b). Some (about 12% of the applied dose over 24 h) dermal absorption has also been observed in rats at non- irritant concentrations, with greater absorption at higher, skin-damaging concentrations (Yang et al., 1987). These animal studies have also demonstrated that EDA and/or its metabolites are widely distributed throughout the body and are rapidly eliminated, largely via the urine but also as carbon dioxide in the breath and a small amount via the faeces, providing evidence for some biliary excretion. It would seem reasonable to conclude that a similar situation with respect to distribution and excretion would pertain in humans. Examination of urinary metabolites in these animal studies demonstrated that EDA is also found in an acetylated conjugate form in the rat and mouse. There is evidence that this pathway may become saturated with increasing dose and that alternative metabolic pathways may be involved at higher doses in the mouse.
8. EFFECTS ON LABORATORY MAMMALS AND IN VITRO TEST SYSTEMS
A number of the available studies on both the toxicokinetics and toxicity of EDA have employed the base substance (EDA) and/or the hydrochloride salt (EDAA2HCl). The latter is used in pharmaceutical prepa- rations as a solubilizer to increase uptake of theophylline (this complex being known as aminophylline) and has been used as a preservative in skin creams (although it is unclear whether or not this still occurs). In general, the presence of the hydrochloride has little qualitative effect on the toxicokinetic or systemic toxicity properties of EDA, particularly following oral dosing, as it is likely that the hydrochloride salt would be formed anyway in the acidic environment of the stomach. However, the hydrochloride does seem to act in a neutralizing capacity to reduce the significant irritancy potential of EDA.
Studies using both forms of EDA are included in this review.
8.1 Single exposure
Studies in various animal species have shown EDA to be of moderate acute toxicity by the inhalation (rat 8-h LC50 estimated to be in the range of 4916–
9832 mg/m3 [1966–3933 ppm]), oral (rat LD50 values of 1160–3250 mg/kg body weight), and dermal (rabbit LD50 values of 550–2880 mg/kg body weight) routes of exposure (Smyth et al., 1941, 1951; Boyd & Seymour, 1946; Carpenter et al., 1948; NTP, 1982a,b; Yang et al., 1983; Dubinina et al., 1997). Few details exist of the toxic signs observed or of target organs.
8.2 Irritation and sensitization
There are a number of reports available on skin irritation in animals, but in general they all repeat the information from one original study (Smyth et al., 1951).
In that study, 0.01 ml undiluted EDA applied to the shaved backs of albino rabbits produced skin necrosis within 24 h. A recent report has also indicated EDA to be a skin irritant (Dubinina et al., 1997). Although no further information is available, such a response is consistent with EDA being strongly alkaline. Studies using EDAA2HCl have also resulted in skin irritation, although the neutralizing action of the hydrochloride may have influenced the severity of effects, particularly on dilution (Yang et al., 1983, 1987).
As with skin irritation, the reports that are available for eye irritation all largely reproduce data from one original study (Carpenter & Smyth, 1946). In this study, 0.005-ml solutions of 5% EDA or greater caused corneal injury, which again would be expected, given the alkaline properties of the substance. More recently, Dubinina et
al. (1997) stated that inflammatory responses in the rabbit eye were induced by “one drop” of EDA.
Overall, from the reports that are available, togeth- er with a consideration of its alkaline properties, it is reasonable to conclude that EDA is corrosive, with the capacity to produce severe chemical burns to the skin and eye.
EDA has been demonstrated to possess skin sensitizing potential in guinea-pig studies, generally using standard methodologies such as the Magnusson and Kligman maximization and Buehler tests
(Thorgeirsson, 1978; Erikson, 1979; Maurer et al., 1979;
Henck et al., 1980; Goodwin et al., 1981; Babiuk et al., 1987; Robinson et al., 1990; Dubinina et al., 1997; Leung
& Auletta, 1997). In four of these studies (Goodwin et al., 1981; Babiuk et al., 1987; Robinson et al., 1990; Leung &
Auletta, 1997), the investigators ensured that non-irritant challenge concentrations of EDA were used, providing clear evidence for a sensitization response. EDA also produced positive results in the local lymph node assay (Basketter & Scholes, 1992). In contrast to these positive results, EDA consistently produced negative results in the mouse ear swelling test (Gad et al., 1986; Cornacoff et al., 1988; Dunn et al., 1990). One study demonstrated the potential for EDA to cross-react with other alkylamines either as the inducing or as the challenge agent (Leung
& Auletta, 1997).
No studies are available on respiratory sensitiza- tion in animals.
8.3 Short-term exposure
In a 12-day study (NTP, 1982b), mice received gavage doses of between 50 and 600 mg EDA/kg body weight per day (administered as EDAA2HCl). Deaths were observed at 400 and 600 mg EDA/kg body weight per day. No effects were seen at 50 mg EDA/kg body weight per day. Renal effects (nephrosis and tubule
regeneration) were observed at 100 mg EDA/kg body weight per day and above. Lymphoid depletion in and necrosis of splenic follicles were observed at 400 mg EDA/kg body weight per day.
A 7 h/day, 30-day inhalation study in rats indicat- ed that the liver and kidney are potential target tissues, with local effects in the lungs also likely (Pozzanni &
Carpenter, 1954). No effects were observed in this study at an airborne exposure concentration of about 150 mg/
m3 (60 ppm). Slight depilatory effects were seen at 330 mg/m3 (132 ppm), becoming more marked at higher exposure concentrations. Treatment-related deaths were observed at 563 mg/m3 (225 ppm) and 1210 mg/m3 (484 ppm) (all animals died at 1210 mg/m3 [484 ppm]).
Cloudy swelling of cells in the liver and convoluted tubules of the kidneys were also observed at these expo-
sure concentrations. Degeneration of the convoluted tubules was seen in animals exposed to 1210 mg/m3 (484 ppm), as was congestion of the lungs and adrenals.
8.4 Long-term exposure 8.4.1 Subchronic exposure
Dietary studies in rats have also indicated that the liver is a target tissue, with changes in the size and shape of hepatocytes and their nuclei being noted at 1000 mg/kg body weight per day in a 90-day study (Yang et al., 1983).
Oral gavage studies using doses of between 100 and 1600 mg EDA/kg body weight per day (administered as EDAA2HCl) have been carried out in rats (NTP, 1982a).
Deaths were observed after 12 doses at 800 and 1600 mg EDA/kg body weight per day and after 800 mg EDA/kg body weight per day for 90 days. Renal tubular lesions (dilation of the lumen, necrosis, degeneration and regeneration of the epithelium) were seen at 200 mg EDA/kg body weight per day and above after 12 doses.
Similar renal lesions but of a less severe nature were seen only at 600 mg EDA/kg body weight per day and above after 90 days. This indicates recovery in the kid- ney, probably as a consequence of compensatory regen- eration. No effects were seen on the kidney at 100 mg EDA/kg body weight per day in either study. Ocular effects including cataract formation and retinal atrophy were observed in all dose groups. Minimal to moderate focal retinal atrophy was observed in 3 out of 10 females at 100 mg EDA/kg body weight per day; 2 males had mild to moderate retinal atrophy and 1 male had severe retinal atrophy at 200 mg EDA/kg body weight per day.
Lymphoid depletion and/or necrosis in spleen were observed at 800 mg EDA/kg body weight per day and in all decedents following 12 doses, and thymus weight was reduced at 800 mg EDA/kg body weight per day in the 90-day study. Uterine lesions (reduced uterine horn size and atrophy of the myometrium and endometrium) were seen after dosing for 90 days with 600 or 800 mg EDA/kg body weight per day, and reduced ovarian size was seen after 800 mg EDA/kg body weight per day for 90 days. Overall, a no-observed-adverse-effect level (NOAEL) was not identified from these studies, as effects on the eyes were seen at all dose levels. Only ocular effects were seen at the lowest-observed-adverse- effect level (LOAEL) of 100 mg EDA/kg body weight per day, and these were of a minimal to mild nature,
suggesting that this dose represented the lower end of the dose–response relationship for these effects.
In a 90-day study, mice received oral gavage doses of 25–400 mg EDA/kg body weight per day (NTP, 1982b). No effects were seen at 100 mg EDA/kg body weight per day. Renal lesions (cortical tubular
degeneration and/or necrosis) were observed at 200 and 400 mg EDA/kg body weight per day.
8.4.2 Chronic exposure and carcinogenicity There are two carcinogenicity studies in animals.
Both studies were performed to reasonably adequate standards, including extensive histopathology, and were negative for carcinogenic activity.
In the first study, groups of 99–225 F344 rats were orally dosed with 0, 20, 100, or 350 mg EDAA2HCl (equivalent to 0, 9, 45, or 158 mg EDA/kg body weight per day) for 2 years (Yang et al., 1984a). Non-neoplastic effects were similar to those described in studies of shorter duration (Yang et al., 1993), as indicated above (section 8.4.1). Effects were seen at 45 mg EDA/kg body weight per day, with a NOAEL of 9 mg EDA/kg body weight per day. Tracheitis was also observed, probably as a consequence of exposure to EDA in airborne dust derived from the diet.
In the second study, groups of 40–50 C3H/HeJ mice were dermally administered 0 or 0.25 mg aqueous EDA 3 times per week for a lifetime (DePass et al., 1984).
The dermal study included a positive control group that received 3-methylcholanthrene. Skin fibrosis and hyper- keratosis were observed in EDA-treated mice.
8.5 Genotoxicity and related end-points Only limited information is available on the geno- toxic potential of EDA. There is some evidence that EDA may be mutagenic in bacteria with and without metabolic activation (Hedenstedt, 1978; Hulla et al., 1981; Haworth et al., 1983; Leung, 1994). Although the most recent study (Leung, 1994) appears to be negative, there was a small response in Salmonella typhimurium TA100 and a positive, but not reproducible, response in TA1535.
Positive results have also been reported in these strains from the other studies, although only one of these (Haworth et al., 1983) was adequately reported. The only series of studies performed on mammalian cell systems in vitro (gene mutation and sister chromatid exchange in Chinese hamster ovary cells; unscheduled DNA synthesis in rat primary hepatocytes) were consistently negative (Slesinski et al., 1983), although there has been no assay for clastogenic activity. A sex-linked recessive lethal test in Drosophila melanogaster was negative following dosing by feeding or injection (Zimmering et al., 1985). There are no in vivo studies on somatic cells, but a dominant lethal study in rats up to doses inducing signs of toxicity (up to 500 mg EDAA2HCl/kg body weight per day in the diet) was negative (Slesinski et al., 1983).
Although there has been some evidence of muta- genicity in bacterial systems in a few limited studies, the
available evidence indicates that EDA is not genotoxic, with all results in mammalian cells in vitro and in vivo (dominant lethal assay) being negative. It should be noted that the overall database is limited, with no assays available for clastogenic activity or for genotoxic poten- tial in somatic cells in vivo.
8.6 Reproductive and developmental toxicity
The potential of EDA to affect fertility and development has been studied in rats in investigations conducted to modern regulatory standards. In a two- generation study in F344 rats, no effects on fertility or development in any of the generations were observed up to a dose level (225 mg EDA/kg body weight per day) that induced signs of parental toxicity (Yang et al., 1984b). Dose levels used in this study were 0, 50, 150, or 500 mg EDAA2HCl/kg body weight per day (equivalent to 0, 23, 68, and 225 mg EDA/kg body weight per day).
Effects on the uterus and ovaries have been seen following gavage dosing of rats with 600 and 800 mg EDA/kg body weight per day for 90 days (see section 8.4.1; NTP, 1982a). In a series of developmental toxicity studies in F344 rats, EDA was found to produce signs of fetotoxicity (increased resorptions) and delays in devel- opment at high dose levels (450 mg EDA/kg body weight per day) that induced clear signs of toxicity in the dams (DePass et al., 1987). Dose levels used in this study were 0, 50, 250, or 1000 mg EDAA2HCl/kg body weight per day (equivalent to 0, 23, 113, and 450 mg EDA/kg body weight per day). Some of the developmental effects appear to have been related, at least in part, to the reduced nutritional status of the animals. However, a clear NOAEL for developmental toxicity of 113 mg EDA/kg body weight per day was observed in these studies.
The results of a preliminary screening study in mice indicated no significant effects on development in the offspring of dams exposed to toxic doses of EDA (400 mg/kg body weight per day) by oral gavage (Hardin et al., 1987).
No effects on development were seen in the off- spring of New Zealand white rabbits dosed during preg- nancy with up to 178 mg EDAA2HCl/kg body weight per day (equivalent to 80 mg EDA/kg body weight per day), a dose that did not induce maternal toxicity (NTP, 1991;
Price et al., 1993). In a preliminary study, 2/20 pregnant rabbits receiving 100 mg EDA/kg body weight per day by gavage died, and decreased body weight was seen in survivors. At 400 mg/kg body weight per day, all the dams died.
8.7 Immunological and neurological effects
No studies are available that have specifically investigated the potential immunotoxicity of EDA.
Effects on lymphoid tissue in the spleen in mice and rats (see sections 8.3 and 8.4.1, respectively) and on the thymus in rats (see section 8.4.1) were observed in oral gavage dosing studies.
There are a few, mainly in vitro, studies on the effects of EDA on the release of (-aminobutyric acid from the retina, gut, and brain (Perkins & Stone, 1980;
Forster et al., 1981; Lloyd et al., 1982; Morgan & Stone, 1982; Sarthy, 1983; Kerr & Ong, 1984; Strain et al., 1984;
Hill, 1985; Erdo et al., 1986; Krantis et al., 1990; McKay &
Krantis, 1991). The general conclusion that can be drawn from these studies is that EDA can cause a calcium- independent release of (-aminobutyric acid that is insensitive to the presence of tetrodotoxin. EDA was also shown to have (-aminobutyric acid mimetic proper- ties (i.e., reduction of neuronal firing rate). This suggests that EDA could have a central nervous system depres- sant effect, but studies were not performed to address this possibility. It was reported that EDA elicited con- traction of the guinea-pig ileum that was mediated via neuronal release of (-aminobutyric acid. However, in the rat ileum, EDA acted directly on the mucosa, resulting in relaxation. Although these are interesting results, the toxicological significance of these findings is unclear;
they may, however, partly explain the central nervous system depressant and gastrointestinal effects seen in some of the animal studies at high doses.
9. EFFECTS ON HUMANS
No studies are available in which the effects, other than the respiratory effects summarized below, of repeat- ed exposure of humans to EDA are examined. No reports have been found in which genotoxicity, carcinogenicity, or reproductive toxicity following exposure to EDA in human populations has been studied.
A case report exists concerning a 36-year-old worker who died from cardiac collapse 55 h after being splashed by an accidental spillage of EDA (Niveau &
Painchaux, 1973). Exposure to an unquantified amount of EDA was in the order of a few minutes prior to the patient being washed. Four hours after the exposure, he presented with tachycardia (100 beats/min), anuria, and red/brown generalized erythema. The tachycardia increased (up to 140 beats/min), anuria persisted, and an expectorant cough, abdominal cramps, diarrhoea, and blackish vomiting appeared. The patient became hyper- kalaemic, and his red blood cell count decreased. Overall,
given the lack of information with respect to levels of exposure, few useful conclusions can be drawn from this case report.
The only information available on skin irritation in humans either is anecdotal or does not involve direct surface skin contact with EDA. In a brief report on the physicochemical properties of EDA, it is noted anecdot- ally that “the liquid, if not washed from the skin, causes blistering” (Boas-Traube et al., 1948). The other report available documents the results of intradermal skin tests with solutions (0.1–1%) of EDA on three individuals being tested for hypersensitivity following treatment with aminophylline (Kradjan & Lakshminarayan, 1981).
The skin response in two patients consisted of blistering rather than a weal and flare reaction, which normally typifies a sensitization response. Punch biopsies were obtained from one patient, and histopathological exami- nation of these indicated tissue necrosis and oedema of the epidermis and dermis. These responses suggested a direct corrosive effect of EDA.
No specific reports were found of the effects of EDA on the eyes of humans. In the original reports of the animal eye irritation study (see section 8.3), it is claimed that EDA is known to have produced loss of vision or slowly healing corneal burns in industrial use (Carpenter & Smyth, 1946). However, no further infor- mation or references were given.
EDA has been known for many years to be capable of inducing allergic skin reactions in humans. This has been observed both in the workplace and, most notably, in patients treated with aminophylline or with skin creams in which EDA was used as a stabilizer (Epstein &
Maibach, 1968; Petrozzi & Shore, 1976; Booth et al., 1979;
Wall, 1982; Hardy et al., 1983; Balato et al., 1984; Edman
& Moller, 1986; Nielsen & Jorgensen, 1987; Terzian &
Simon, 1992; Toal et al., 1992; Dias et al., 1995; Simon et al., 1995; Sasseville & Al-Khenaizan, 1997). The first reports of skin sensitizing effects in humans date back to the late 1950s, when cases were described of eczematous reactions in pharmacists who came into contact with EDA when using aminophylline (Baer et al., 1958; Tas &
Weissberg, 1958).
Subsequent to these early reports, numerous stud- ies and case reports have been published documenting the skin sensitizing properties of EDA both following clinical use and within the occupational setting, such that the substance has become incorporated into stan- dard series for patch testing (Fregert, 1981; Shehade et al., 1991). An example from the clinical setting is that of the report on a series of 13 patients who had used skin cream containing EDA for, paradoxically, dermatitic conditions (Provost & Jillson, 1967). Use of the cream in 11 of these patients had resulted in the sudden
appearance of a severe generalized patchy eczematous
eruption following, in all but one of the cases, an initial improvement upon using the cream. Patch testing was conducted using a 1% aqueous solution of EDA, producing skin reactions in all patients ranging from erythema and oedema to erythema vesiculation and oedema vesiculation, which extended beyond the patch test site. Other substances tested also induced responses, but not in a consistent manner, with at most only four individuals responding in any one test.
As well as the original reports in pharmacists working with EDA, cases of skin sensitization to EDA have been reported in the occupational environment in a number of different settings, including use of floor polish remover (English & Rycroft, 1989), use of coolant oils (Crow et al., 1978), and in wire-drawing (Matthieu et al., 1993; Sasseville & Al-Khenaizan, 1997). Positive responses to patch testing with EDA have also been observed in other occupational settings, such as the offshore oil industry (Ormerod et al., 1989), but positive responses to other substances, including other
polyamines, were also seen in such cases. Thus, it is unclear whether EDA had been responsible for inducing the sensitized state and/or cross-reacting following sensitization to another polyamine.
A large number of cases of occupational asthma reported to have been caused by exposure to EDA are available in the literature (Dernehl, 1951; Gelfand, 1963;
Popa et al., 1969; Valeyeva et al., 1975; Lam &
Chan-Yeung, 1980; Chan-Yeung, 1982; Hagmar et al., 1982; Matsui et al., 1986; Aldrich et al., 1987; Nakazawa
& Matsui, 1990; Lewinsohn & Ott, 1991; Ng et al., 1991, 1995). There are a few studies in which the potential for EDA to cause respiratory hypersensitivity has been examined using bronchial provocation testing and investigation of antibody formation. As EDA is corrosive, the vapour would be predicted to be a respi- ratory tract irritant, which is a complicating factor in interpreting the data available and in elucidating the underlying mechanism for any asthmatic responses seen.
Popa et al. (1969), in a well-conducted study, investigated 48 subjects with asthmatic symptoms caused by exposure to a number of low molecular weight chemicals, including EDA. None of the subjects had a history of respiratory disorder prior to occupational exposure, and the asthmatic response was associated only with occupational exposure in all cases. No informa- tion was given in the report on the workplace airborne concentrations of EDA to which these workers were exposed. A series of tests were performed in all subjects, including skin and inhalation tests with the test agent at sub-irritant concentrations; skin and inhalation tests to common allergens; skin tests (intradermal, scratch, and patch tests) using sub-irritant concentrations of the test substance; Prausnitz-Kustner transfer reaction (to test
for the presence of immunoglobulin E antibodies); and determination of precipitating antibodies to EDA. For the inhalation test, the sub-irritant concentration was determined in control asthmatic subjects, and a 2- to 10- fold dilution of this was used for the bronchial challenge.
No information was given on the airborne exposure concentrations generated under these test conditions.
Control inhalation tests with the diluent, physiological saline, were also conducted. It is not stated in the report whether or not the inhalation challenge tests were conducted in a blind manner.
Six subjects had an immediate, positive reaction to EDA in the workplace. Of these, four showed an immedi- ate, positive response following inhalation testing with sub-irritant concentrations of EDA. These subjects developed marked bronchoconstriction following inhala- tion exposure to EDA, with a reduction in forced expi- ratory volume in 1 s (FEV1) of 62% and an increase in respiratory resistance of 44%, compared with controls.
Although not stated in the report, these values are pre- sumed to be average changes. Intradermal skin tests with EDA were positive in these four subjects, whereas patch tests were negative. Inhalation challenges with common allergens were negative. The Prausnitz-Kustner test was positive in all subjects, and all had eosinophilia, deter- mined in the sputum, although not, except in one case, in the blood. No precipitating antibodies were found. In the two other subjects, the inhalation challenge test was negative. No precipitating antibodies were found, and the Prausnitz-Kustner test was negative in both subjects. Eosinophilia was absent. Inhalation challenges with other common allergens were also negative.
These data provide evidence that EDA may elicit an asthmatic response at sub-irritant concentrations and that the response is specific to EDA. Four out of six subjects responsive to EDA in the workplace also had a positive response to inhaled EDA at sub-irritant concen- trations. This demonstrates that the reaction is not a generalized response to an irritant. The positive Prausnitz-Kustner reaction may be indicative of an immunological component, but the test is not specific, and no firm conclusions can be drawn from it. It provides supporting evidence in this case. The evidence suggests that the subjects were hypersensitive to inhaled EDA and that a state of respiratory hypersensitivity had been induced by the substance.
Although a number of other studies are available, the information is of poor quality. Lam & Chan-Yeung (1980) and Chan-Yeung (1982) describe the case of one worker in a photographic laboratory who developed asthma after 2.5 years of exposure to a variety of chemi- cals, including EDA, but also other irritant substances.
The worker developed symptoms of sneezing, nasal discharge, productive cough and nocturnal cough, wheezing, and dyspnoea. The symptoms coincided with
the work shift and subsided at weekends. There was no previous history of asthma. No information was given in the report of the airborne concentrations of EDA (or other substances) to which the man was exposed at work. A series of controlled inhalation challenge expo- sures, designed to mimic work exposure conditions, were conducted with each of the chemicals to which the subject was exposed at work. The duration of exposure was determined by the patient’s tolerance, and exposure was terminated when eye irritation or cough was experi- enced. No information on the airborne exposure concen- trations of EDA generated under these test conditions was given in the report. A methacholine inhalation test for bronchial hyperreactivity was also performed. Pul- monary function tests were conducted pre- and post- challenge, and blood samples were taken before, during, and after each challenge. The subject showed marked bronchial reactivity to methacholine.
Exposure to an unknown concentration of vapour from a 1:25 solution of EDA was tolerated for 15 min.
This exposure produced a marked bronchoconstriction.
A late asthmatic response developed 4 h after the expo- sure, at which time FEV1 was reduced by 26% and continued to decrease over the next 3 h towards a 40%
reduction. A 26% reduction was still apparent after 24 h, despite treatment with bronchodilator drugs. This pat- tern of response to EDA was reproducible. The patient did not respond similarly to any of the other chemicals tested: formaldehyde, sulfur dioxide, and two colour developing agents that were stated to be irritants. Expo- sure to formaldehyde (vapour from a 1:4 solution) produced an immediate small (<20%), transient reduction in FEV1, whereas exposure to sulfur dioxide caused coughing and chest tightness and an immediate transient reduction of 25% in FEV1. There was no increase in plasma histamine concentration during the period of bronchoconstriction, although EDA was shown to cause in vitro histamine release from whole blood taken from the patient and from two control subjects. A skin test using 1:100 EDA and a precipitin test for antibodies to EDA were both negative. The patient subsequently had to give up work because of respiratory symptoms and became asymptomatic after 2 weeks. Subsequent testing with methacholine, 2.4 months after ceasing work, showed that the subject had a reduction in the previous bronchial hyperreactivity.
In conclusion, the subject showed an asthmatic response to EDA but not to formaldehyde or the colour developing agents. The pattern of response to sulfur dioxide was more immediate and suggestive of an irritation response. Overall, a clear pattern of asthmatic response that was specific to EDA was observed in this study. However, it is not possible to distinguish with certainty between an irritant response and a sensitization response, because it is possible that an irritant concen- tration was used for the bronchial challenge exposure,
although little immediate response was observed. In addition, although exposure to irritant concentrations of the other workplace chemicals did not elicit the same pattern, magnitude, or severity of response as that seen with EDA, since accurate exposure levels were not given, it is not possible to determine whether or not the EDA concentration used had the greatest irritant poten- tial. No evidence for any immunological involvement was found. In conclusion, this study provides only
circumstantial evidence that EDA caused a state of respiratory hypersensitivity in this subject.
A number of other case reports are available of individuals who exhibited asthmatic signs and symptoms associated with exposure to EDA in the workplace (Gelfand, 1963; Valeyeva et al., 1975; Matsui et al., 1986;
Nakazawa & Matsui, 1990; Ng et al., 1991). Although bronchial challenge testing with EDA produced
asthmatic responses in these subjects, they had personal and/or family histories of allergic disease and/or they had worked with and responded on challenge to other substances. Retrospective studies using the medical records of populations of workers using EDA have indicated that about 10% of such populations developed signs and symptoms of occupational asthma (Aldrich et al., 1987; Lewinsohn & Ott, 1991). No challenge tests were carried out with these surveys. Thus, these case reports and population-based studies provide only sup- porting circumstantial evidence for the involvement of EDA in producing occupational asthma.
Although it is clear from these reports that EDA can provoke an asthma attack, in many cases there is insufficient information to indicate whether or not the hypersensitive state was induced specifically by EDA.
However, from one well-conducted study, there is evidence that a hypersensitive state specific to EDA has been induced in workers and that an asthmatic response was provoked by sub-irritant concentrations of the substance. Overall, the results of this study, taken together with the supporting data from a substantial number of other reports of occupational asthma, indicate that EDA is capable of inducing a state of hypersensitiv- ity in the airways, such that subsequent exposure may trigger asthma. The mechanism by which the hypersensi- tive state is induced is not proven. Given the skin sen- sitizing potential of EDA and the limited evidence of immunological involvement in workers with EDA- provoked occupational asthma, an immunological mechanism would seem plausible. Irrespective of the mechanism involved, the data available (specifically the lack of information on airborne exposure concentrations under both work and challenge test conditions) do not allow elucidation of a dose–response relationship or the identification of levels of EDA that are not capable of inducing a hypersensitive state or of provoking an asthmatic response.
10. EFFECTS ON OTHER ORGANISMS IN THE LABORATORY AND FIELD
Results of acute ecotoxicity tests are given in Table 2.
A 21-day Daphnia magna reproduction test was conducted according to the German Federal Environment Agency (Umweltbundesamt) guidelines in a closed vessel. End-points measured included adult mortality, onset of production of young, and reproduction rate.
The most sensitive end-point was for reproductive rate, and a NOEC of 0.16 mg/litre was established (Kuhn et al., 1989). In a second study conducted according to Organ- isation for Economic Co-operation and Development (OECD) guidelines, a NOEC for reproduction was reported at 2 mg/litre (Mark & Hantink-de Rooy, 1992).
An early life stage test conducted under OECD guide- lines on three-spined stickleback (Gasterosteus acule- atus) showed no effects of EDA at 10 mg/litre (the highest concentration tested) over 28 days (Mark &
Arends, 1992).
Growth of lettuce (Lactuca sativa) plants over 7 days was studied in tests conducted under OECD Guideline 208; EC50 concentrations for EDA in soil (nominal) were >1000 mg/kg for 7-day and 692 mg/kg for 14-day growth periods (Hulzebos et al., 1993).
11. EFFECTS EVALUATION
11.1 Evaluation of health effects
11.1.1 Hazard identification and dose–response assessment
EDA is of moderate acute toxicity by all routes. In studies in animals, EDA is a primary skin and eye irritant, being corrosive when undiluted. It is also a skin
sensitizer. In repeated-dose toxicity studies by the oral and inhalation routes, effects on the liver and kidney have been observed, with pleomorphic changes to hepatocytes in rats being reported at the lowest oral doses used (45 mg/kg body weight per day and above for 2 years; NOAEL, 9 mg/kg body weight per day). In inhalation studies, there were no effects at 150 mg/m3 (60 ppm), although slight depilation was observed at the next highest concentration (330 mg/m3 [132 ppm]) and effects on the liver and kidney at higher concentrations still (approximately 500 mg/m3 [200 ppm] and above).
There has been some evidence of mutagenicity in bacterial systems in a few limited studies. However, much of the available evidence is negative, although the
