GHS: Health hazards Classification and labelling including basic toxicology

GHS: Health hazards Classification and labelling including basic toxicology Lennart Dock Swedish Chemicals Agency lennart.dock@kemi.se ITP299EU 2014 15 September - 7 October 2014

Alle Ding sind Gift und nichts ohn Gift; alein die Dosis macht, das ein Ding kein Gift ist Response 100% Dose Paracelsus 1493-1541 (Theophrastus Bombastus Philippus Aureolus Paracelsus von Hohenheim) All is poison, nothing is without poison; only the dose determines what is not a poison

Methyl mercury toxicity 100 80 Fetal toxicity Paresthesia Ataxia Prevalence, % 60 40 Dysarthria Deafness 20? 0 1 10 100 1000 Methyl mercury in hair, mg/kg Death WHO (1990), EHC 101

Basic concepts Some chemicals produce local effects Most chemicals produce systemic effects effects do not occur at the site of first contact Require absorption and distribution to a distant site where the effects are produced Most chemicals are metabolised before excretion Adverse effects are usually observed in one or two organs (target organs)

ADME Chemicals Skin Lung Gastrointestinal tract Absorption Distribution Metabolism Other tissues Blood Liver Excretion Kidney Hair Breast milk Urine Faeces

Biotransformation Lipohilic Hydrophilic

Biotransformation of paracetamol Lethal dose 10-15 g - or even less Within hours: nausea, vomiting, pallor, sweating Between 24 and 72 hours: signs of increasing liver damage, possibly kidney failure At 3 to 5 days: massive hepatic necrosis, kidney failure, multiple organ failure, and death.

Hazard Class Acute Toxicity Oral Dermal Inhalation Skin Corrosion/Irritation Serious Eye Damage/Irritation Sensitization Respiratory Skin Germ Cell Mutagenicity Carcinogenicity Reproductive Toxicity STOT - Single Exposure STOT - Repeated Exposure Aspiration hazard Health hazards Hazard Category 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 Corrosive 1A 1B 1C 1A 1A Decrease in severity 1A 1B 2 1A 1B 2 1A 1B 2 1 2 1 2 Irritant 2 3 1 2A 2B 1B 1B 1 2 3 Lactation

Acute toxicity Single exposure High dose Clinical symptoms Treatment Recovery Chronic toxicity Repeated exposure Low dose No overt clinical symptoms Treatment not always possible Sustained/irreversible damage

Weight of evidence Human epidemiological studies Human case reports Toxicity testing, in vivo studies Toxicity testing, in vitro mammalian cell assays Non-testing methods

Acute oral and dermal toxicity: LD50: Dose causing death in 50% of the treated animals following single exposure via oral or dermal route. Dose causing evident toxicity Preferred species rat and rat/rabbit. Acute toxiciy 100% Lethality 50% LD50 Acute inhalation toxicity: LC50: Concentration causing death in 50% of the treated animals following 4 hours of exposure by inhalation. Preferred species rat. GHS distinguishes between exposure to dust, mist, gas and vapour Dose Note: Present OECD TG do not have lethality (LD50, LC50) as an endpoint

Inhalation Classification criteria: Acute toxicity Exposure Route Oral (mg/kg bw) Dermal (mg/kg/bw) Gases (ppmv) Vapours (mg/l) Dust/mists (mg/l) Category 1 Category 2 Category 3 Category 4 Category 5 ATE 5 5 < ATE 50 50 < ATE 300 300 < ATE 2000 2000 < ATE 5000 ATE 50 50 < ATE 200 200 < ATE 1000 1000 < ATE 2000 2000 < ATE 5000 ATE 100 100 < ATE 500 500 < ATE 2500 2500 < ATE 20000 ATE 0.5 0.5 < ATE 2.0 2.0 < ATE 10 10 < ATE 20 ATE 0.05 0.05 < ATE 0.5 0.5 < ATE 1.0 1.0 < ATE 5 Equivalent to oral or dermal route Equivalent to oral or dermal route Equivalent to oral or dermal route ATE = acute toxicity estimate

Acute Toxic Class Method (OECD TG 423) Recommended starting dose GHS Cat.1 >0-5 GHS Cat.2 >5-50 GHS Cat.3 >50-300 GHS Cat.4 >300-2000 GHS Cat.5 >2000-5000 Category 5 or Unclassified

Classification of a mixture: acute oral toxicity Concentration ATE Classification Substance 1 1% 225 mg/kg Substance 2 3% 100 mg/kg Substance 3 10% acute tox, cat 3 Water 86% Note: The relevant ingredients to be considered when classifying a mixture for acute toxicity are those present at 1%

Additivity formula for classification of a mixture based on the acute toxicity of the ingredients 1) Data available for all ingredients 100 ATE mixture n Ci ATEi C i i n ATE i = concentration of ingredient i (% w/w or % v/v) = the individual ingredient from 1 to n = the number of ingredients = Acute Toxicity Estimate of ingredient i

Inhalation Conversion from range value/category to point estimate Exposure Route Oral (mg/kg bw) Category or experimental estimate 0 < category 1 5 5 < category 2 50 50 < category 3 300 300 < category 4 2000 2000 < category 5 5000 Dermal (mg/kg/bw) 0 < category 1 50 50 < category 2 200 200 < category 3 1000 1000 < category 4 2000 2000 < category 5 5000 Gases (ppmv) Vapours (mg/l) 0 < category 1 100 100 < category 2 500 500 < category 3 2500 2500 < category 4 20000 Category 5 equivalent 0 < category 1 0,.5 0,5 < category 2 2,.0 2,0 < category 3 10 10 < category 4 20 Category 5 equivalent Dust/mists (mg/l) 0 < category 1 0,05 0.05 < category 2 0.5 0.5 < category 3 1.0 1.0 < category 4 5 Category 5 equivalent Converted point estimate 0.5 5 100 500 2500 5 50 300 1100 2500 10 100 700 4500 0.05 0.5 3 11 0.005 0.05 0.5 1.5

Classification of a mixture: acute oral toxicity Concentration ATE Classification Substance 1 1% 225 mg/kg Substance 2 3% 100 mg/kg Substance 3 10% 50 300 mg/kg Water 86% 100 ATE mixture = 1 100 ATE 225 + mixture 3 100 + n 10 100 Ci ATEi acute tox, cat 3 ATE mixture = 743 Point estimate: 100 classification: acute tox, cat 4

Additivity formula for classification of a mixture based on the acute toxicity of the ingredients 2) Data not available for all ingredients 100 C unknownif 10% ATE mixture n C i ATE i C i i n ATE i = concentration of ingredient i (% w/w eller % v/v) = the individual ingredient from 1 to n = the number of ingredients = Acute Toxicity Estimate for ingredient i

Label elements Category 1 Category 2 Category 3 Category 4 Category 5 Oral H300: Fatal if swallowed H300: Fatal if swallowed H301: Toxic if swallowed H302: Harmful if swallowed H303: May be harmful if swallowed Dermal H310: Fatal in contact with skin H310: Fatal in contact with skin H311: Toxic in contact with skin H312: Harmful in contact with skin H313: May be harmful in contact with skin Inhalation H330: Fatal if inhaled H330: Fatal if inhaled H331: Toxic if inhaled H332: Harmful if inhaled H333: May be harmful if inhaled Danger Danger Danger Warning Warning

Skin corrosion/irritation Skin corrosion: Irreversible damage dermal exposure Skin irritation: Damage showing reversibility within 14 days after treatment

Skin corrosion/irritation category 1: Corrosive Corrosive* in 1 of 3 animals Sub-category Exposure Observation 1A 3 minutes 1 hour 1B > 3 min to 1 hour 14 days 1C > 1 hour to 4 hours 14 days *visible necrosis through the epidermis and into the dermis

Skin corrosion/irritation Category 2: Irritant Category 3: Mild irritant Criteria a) Mean value of 2.3-4.0 for erythema/eschar or for oedema in at least 2 of 3 tested animals from gradings at 24, 48 or 72 hours after patch removal or, if reactions are delayed, from grades on 3 consecutive days after the onset of skin reactions; or b) Inflammation that persists to the end of the observation period (normally 14 days) in at least 2 animals, particularly taking into account alopecia (limited area), hyperkeratosis, hyperplasia, and scaling; or c) In some cases where there is pronounced variability of response among animals, with very definite positive effects related to chemical exposure in a single animal but less than the criteria above. Mean value of 1.5-2.3 for erythema/eschar or for oedema in at least 2 of 3 tested animals from gradings at 24, 48 or 72 hours or, if reactions are delayed, from grades Not on in 3 consecutive CLP days after the onset of skin reactions (when not included in the irritant category above)

Grading of skin reactions (OECD TG 404) Erythema and Eschar Formation No erythema 0 Very slight erythema (barely perceptible) 1 Well defined erythema 2 Moderate to severe erythema 3 Severe erythema (beef redness) to eschar formation preventing grading of erythema 4 Oedema Formation Maximum possible: 4 No oedema 0 Very slight oedema (barely perceptible) 1 Slight oedema (edges of area well defined by definite raising) 2 Moderate oedema (raised approximately 1 mm) 3 Severe oedema (raised >1 mm and extending beyond area of exposure) 4 Maximum possible: 4

Test results (OECD TG 404) Animal no. Degree of erythema after Degree of oedema after Mean 24/48/72h 2.3? 1h 24h 48h 72h 7d 1h 24h 48h 72h 7d Erythema Oedema 1 3 3 3 2 0 1 2 2 2 0 Mean score = 2.7 Mean score = 2.0 Yes No 2 3 3 3 3 0 1 2 2 1 0 Mean score = 3 Mean score = 1.7 Yes No 3 1 1 1 0 0 1 1 1 1 0 Mean score = 0.66 Mean score = 1.0 No No 2/3 positive responders (exceeding mean score 2.3 for erythema) Skin Irritant Category 2

Classification of a mixture: skin corrosion/irritation Sum of ingredients classified as: Concentration triggering classification of a mixture as: Skin Corrosive Category 1* Skin Irritant Category 2 Skin Irritant Category 3 Skin Category 1 5% 1% but < 5% Skin Category 2 10% 1% but < 10% Skin Category 3 10% (10 x Skin Category 1) + Skin Category 2 (10 x Skin Category 1) + Skin Category 2 + Skin Category 3 10% 1% but < 10% 10% *If sub-categories are used: The sum of all ingredients of a mixture classified as Skin Corrosive Category 1A, 1B or 1C, respectively, should each be 5% in order to classify the mixture as either Skin Corrosive Category 1A, 1B or 1C. If the sum of the Skin Corrosive Category 1A ingredients is < 5% but the sum of Category 1A+1B ingredients is 5%, the mixture should be classified as Skin Corrosive Category 1B. If the sum of the Skin Corrosive Category 1A+1B ingredients is < 5% but the sum of of Category 1A+1B+1C ingredients is 5% the mixture should be classified as Skin Corrosive Category 1C.

Classification of a mixture when the additivity approach does not apply Ingredient Concentration Mixture classified as: Skin Acid with ph 2 1% Category 1 Base with ph 11.5% 1% Category 1 Other corrosive (Category 1) ingredient 1% Category 1 Other irritant (Category 2/3) ingredient, including acids and bases. 3% Category 2

Label elements Category 1 Category 2 Category 3 Category 1A Category 1B Category 1C H314: Causes severe skin burns and eye damage H315: Causes skin irritation H316: Causes mild skin irritation Danger Warning Warning

Serious eye damage/eye irritation Serious eye damage: Not fully reversible damage (decay of vision) following single topical exposure Eye irritation: full reversibility within 21 days after treatment

Serious eye damage Category 1: Irreversible effects on the eye Criteria If, when applied to the eye of an animal, a substance produces: At least in one animal effects on the cornea, iris or conjunctiva that are not expected to reverse or have not fully reversed within an observation period of normally 21 days; and/or At least in 2 of 3 tested animals, a positve response of: Corneal opacity 3 and/or iritis > 1.5 calculated as the mean score following grading at 24, 48 and 72 hours after instillation of the test material

Reversible effects on the eye Category 2A: Irritating to eyes Category 2B: Mildly irritating to eyes Criteria If, when applied to the eye of an animal, a substance produces: At least in 2 of 3 tested animals, a positive response of: Corneal opacity 1; and/or iritis > 1; and/or conjunctival redness 2; and/or conjunctival oedema (chemosis) 2 calculated as the mean score following grading at 24, 48 and 72 hours after instillation of the test material, and which fully reverses within an observation period of normally 21 days. When the effects listed above are fully reversible within 7 days of observation. Not in CLP

Classification of a mixture: serious eye damage/eye irritation Sum of ingredients classified as: Concentration triggering classification of a mixture as: Serious eye damage Eye irritation Category 1 Category 2/2A Skin Category 1 + Eye Category 1 a 3% 1% but < 3% Eye Category 2 10 x (skin Category 1 + eye category 1) a + eye Category 2 10% b 10% a If an ingredient is classified as both skin Category 1 and eye Category 1 its concentration is considered only once in the calculation b A mixture may be classified as eye Category 2B when all relevant ingredients are classified as eye Category 2B

Classification of a mixture when the additivity approach does not apply Ingredient Concentration Mixture classified as: Eye Acid with ph 2 1% Category 1 Base with ph 11.5% 1% Category 1 Other corrosive (eye Category 1) ingredient Other eye irritant (eye Category 2) ingredient 1% Category 1 3% Category 2

Label elements Category 1 Category 2A Category 2B H318: Causes serious eye damage H319: Causes serious eye irritation H320: Causes eye irritation Danger Warning Warning

Aspiration toxicity Entry of a liquid or solid chemical product directly through the oral or nasal cavity, or from vomiting, into the trachea and lower respiratory system. May result in chemical pneumonia and varying degrees of pulmonary injury which may be fatal Physicochemical properties Human experience No test method available

Aspiration toxicity Category Category 1 Category 2 Criteria Chemicals known to cause human aspiration toxicity hazards or to be regarded as if they cause human aspiration toxicity hazard. A substance is classified in Category 1: a) Based on reliable and good quality human evidence, or b) If it is a hydrocarbon and has a kinematic viscosity of 20.5 mm 2 /s, measured at 40 C Chemicals which cause concern owing to the presumption that they cause human aspiration toxicity hazard. Not in CLP On the basis of existing animal studies and expert judgement that takes into account surface tension, water solubility, boiling point, and volatility, substances, other than those classified in Category 1, which have a kinematic viscosity 14 mm 2 /s, measured at 40 C

Classification of a mixture:aspiration toxicity Category 1 A mixture which contains a total of 10% of a substance or substances classified in Category 1, and has a kinematic viscosity of 20.5 mm 2 /s measured at vid 40 C A mixture which separates into two or more distinct layers, one of which contains 10% of a substance classified in Category 1 and has a kinematic viscosity of 20,5 mm 2 /s measured at 40 C Category 2* A mixture which contains a total of 10% of a substance or substances classified in Category 2, and has a kinematic viscosity of 14 mm 2 /s measured at vid 40 C A mixture which separates into two or more distinct layers, one of which contains 10% of a substance classified in Category 1 and has a kinematic viscosity of 14 mm 2 /s measured at 40 C *The use of expert judgement that considers surface tension, water solubility, boiling point, volatility is critical, especially when Category 2 substances are mixed with water

Label elements Category 1 Category 2 H304: May be fatal if swallowed and enters airways H305: May be harmful if swallowed and enters airways Danger Warning

Allergy An adverse immune reaction to a protein (or allergen) that develops after repeated exposure to the causative agent. Sensitisation (induction of specialised immunological memory) takes place on initial exposure to an allergen during which time no adverse reaction appears. Elicitation is the reaction provoked by repeated exposure to the allergen

Sensitisation Skin sensitisation: Evidence for an immunological reaction (e.g. erythema, oedema, contact urticaria) in humans or test animals. Respiratory sensitisation: Asthma in exposed humans; consider size of exposure (number of people and concentration). Currently no test system available. Other: Rhinitis conjunctivitis alveolitis

Sensitisation Adaptive immune responses are systemic in nature; skin sensitisation does not result only from dermal exposure respiratory sensitisation requires protection of both skin and respiratory tract ingestion of allergen can induce respiratory effects

Immune response Type 1 Type 4 Immediate Antibody-mediated Allergic rhinitis Bronchial asthma Urticaria (hives) Delayed Cell-mediated Inflammatory response

Respiratory sensitisation Category 1 Sub-category 1A Subcategory 1B Criteria A substance is classified as a respiratory sensitizer: a) If there is evidence in humans that the substance can lead to specific respiratory hypersensitivity, and/or b) If there are positive results from an appropriate animal test* Substances showing a high frequency of occurrence in humans; or a probability of occurrence of a high sensitization rate in humans based on animal or other tests. Severity of reaction may also be considered Substances showing a low to moderate frequency of occurrence in humans; or a probability of occurrence of a low to moderate sensitization rate in humans based on animal or other tests. Severity of reaction may also be considered *presently there are no recognized and validated animal models available

Classification of a mixture: respiratory sensitisation Ingredient classified as: Respiratory sensitizer Category 1 Respiratory sensitizer Sub-category 1A Respiratory sensitizer Sub-category 1B Concentration triggering classification of a mixture as: Solid/Liquid Respiratory Sensitizer Category 1 Gas 0.1%* 0.1%* 1.0% 0.2% 0.1% 0.1% 1.0% 0.2% *Some competent authorities may require SDS and/or supplemental labelling for mixtures containing a sensitizing ingredient above this concentration

Skin sensitisation Category 1 Sub-category 1A Subcategory 1B Criteria A substance is classified as a skin sensitizer: a) If there is evidence in humans that the substance can lead sensitization by skin contact in a substantial number of persons, or b) If there are positive results from an appropriate animal test Substances showing a high frequency of occurrence in humans and/or a high potency in animals can be presumed to have the potential to produce significant sensitization in humans. Severity of reaction may also be considered Substances showing a low to moderate frequency of occurrence in humans and/or a low to moderate potency in animals can be presumed to have the potential to produce sensitization in humans. Severity of reaction may also be considered

Classification of a mixture: skin sensitisation Ingredient classified as: Skin sensitizer Category 1 Skin sensitizer Sub-category 1A Skin sensitizer Sub-category 1B Concentration triggering classification of a mixture as: Skin sensitizer Category 1 0.1%* 1.0% 0.1% 1.0% *Some competent authorities may require SDS and/or supplemental labelling for mixtures containing a sensitizing ingredient above this concentration

Label elements Respiratory sensitization Category 1/1A/1B H334: May cause allergy or asthma symptoms or breathing difficulties if inhaled Skin sensitization Category 1/1A/1B H317: May cause an allergic skin reaction Danger Warning

Specific target organ toxicity (STOT)

STOT single exposure Category Category 1 Category 2 Criteria Substances that have produced significant toxicity in humans or that, on the basis of evidence from studies in experimental animals, can be presumed to have the potential to produce significant toxicity in humans following single exposure. Substances are classified in Category 1on the basis of: a) Reliable and good quality evidence from human cases or epidemiological studies; or b) Observations from appropriate studies in experimental animals in which significant and/or severe toxic effects, of relevance to human health, were produced at generally low exposure concentrations. Substances that, on the basis of evidence from studies in experimental animals can be presumed to have the potential to be harmful to human health following single exposure. Substances are classified in Category 2 on the basis of observations from appropriate studies in experimental animals in which significant toxic effects, of relevance to human health, were produced at generally moderate exposure concentrations. In exceptional cases, human evidence can also be used to place a substance in Category 2.

STOT single exposure Supporting classification Not supporting classification Substance-related deaths Major functional changes in the central or peripheral nervous systems (sight, hearing, sense of smell) or other organ systems (for example the lung) Severe disturbances (eg haematological disturbances suggesting that they are due to decreased bone marrow production of blood cells) Severe organ damage, especially heart, nervous system, stem cell populations Only change in body weight gain, food consumption or water intake Small changes (clin chem) which are of doubtful or minimal toxicological importance Changes in organ weights with no evidence of organ dysfuntion Adaptive responses (enzyme induction, liver hypotrophy, macrophage migration in the lung) Species-specific mechanism

STOT single exposure: Guidance values Route of exposure Guidance value ranges for:* Unit Category 1 Category 2 Category 3 Oral (rat) mg/kg bw C 300 300 < C 2000 Dermal (rat or rabbit) Inhalation (rat) gas Inhalation (rat) vapour Inhalation (rat) dust/mist/fume mg/kg bw C 1000 1000 < C 2000 ppmv/4h C 2500 2500 < C 20000 mg/l/4h C 10 10 < C 20 mg/l/4h C 1.0 1.0 < C 5.0 Guidance values do not apply (based on human data) *The guidance values are intended for guidance purposes to be used as part of the weight-of-evidence approach and to assist in decision about classification. They are not intended as strict demarcation values.

STOT single exposure Category Category 3 Transient target organ effects Criteria This category only includes narcotic effects and respiratory tract irritation. These are target organ effects for which a substance does not meet the criteria to be classified in Categories 1 or 2 indicated above. These are effects which adversly alter human function for a short duration after exposure and from which humans may recover in a reasonable period without leaving significant alteration of structure or function.

Classification of a mixture: STOT SE Ingredient classified as: Generic concentration limits triggering classification of the mixture as: Category 1 Category 2 Category 1 STOT 1.0% a 1.0 C < 10% c 10% b Category 2 STOT 1.0% a - 10% b a between 1.0 and 10% an SDS would be required. A label may be required in some jurisdictions b at a concentration of 10% both an SDS and a label would generally be expected c if a Category 1 STOT ingredient is present between 1.0 and 10%, some authorities classify the mixture as STOT 2

Label elements Category 1 Category 2 Category 3 H370: Causes damage to organs H371: May cause damage to organs H335: May cause respiratory irritation or H336: may cause drowsiness or dizziness Danger Warning Warning

STOT repeated exposure Category Category 1 Category 2 Criteria Substances that have produced significant toxicity in humans or that, on the basis of evidence from studies in experimental animals, can be presumed to have the potential to produce significant toxicity in humans following repeated exposure. Substances are classified in Category 1 on the basis of: a) Reliable and good quality evidence from human cases or epidemiological studies; or b) Observations from appropriate studies in experimental animals in which significant and/or severe toxic effects, of relevance to human health, were produced at generally low exposure concentrations.. Substances that, on the basis of evidence from studies in experimental animals can be presumed to have the potential to be harmful to human health following repeated exposure. Substances are classified in Category 2 on the basis of observations from appropriate studies in experimental animals in which significant toxic effects, of relevance to human health, were produced at generally moderate exposure concentrations. In exceptional cases, human evidence can also be used to place a substance in Category 2

STOT repeated exposure: Guidance values Guidance value ranges for:* Route of exposure Unit Category 1 Category 2 Oral (rat) mg/kg bw/day C 10 10 < C 100 Dermal (rat or rabbit) Inhalation (rat) gas Inhalation (rat) vapour Inhalation (rat) dust/mist/fume mg/kg bw/day C 20 20 < C 200 ppmv/6h/day C 50 50 < C 250 mg/l/6h/day C 0.2 0.2 < C 1.0 mg/l/6h/day C 0.02 0.02 < C 0.2 *The guidance values are intended for guidance purposes to be used as part of the weight-of-evidence approach and to assist in decision about classification. They are not intended as strict demarcation values.

Classification of a mixture STOT RE Ingredient classified as: Generic concentration limits triggering classification of the mixture as: Category 1 Category 2 Category 1 STOT 1.0% a 1.0 C < 10% c 10% b Category 2 STOT 1.0% a - 10% b a between 1.0 and 10% an SDS would be required. A label may be required in some jurisdictions b at a concentration of 10% both an SDS and a label would generally be expected c if a Category 1 STOT ingredient is present between 1.0 and 10%, some authorities classify the mixture as STOT 2

Label elements Category 1 Category 2 H372: Causes damage to organs through prolonged or repeated exposure H373: May cause damage to organs through prolonged or repeated exposure Danger Warning

Genotoxicity

Definitions Mutation: a permanent change in the amount or structure of the genetic material in a cell In germ cells genetic damage can be transferred to the next generation (via sperms, eggs) In somatic cells genetic damage can be transferred to daughter cells (e.g. in the liver, skin) Mutagen: an agent that gives rise to an increased occurence of mutations in populations of cells and/or organisms Genotoxicity: applies to agents or processes which alter the structure, information content or segregation of DNA, including replication processes

Reverse mutation

Chromosome abberrations (in vitro; mammalian cells): Large deletions and rearrangements Unequal partition of chromosomes between daughter cells (nondisjunction) Sister chromatid exchange

Category 1 Category 1A Germ cell mutagenicity Criteria Substances known to induce heritable mutations or to be regarded as if they induce heritable mutations in the germ cells of humans. Substances known to induce heritable mutations in germ cells of humans Classification is based on positive evidence from human epidemiological studies. Category 1B Substances to be regarded as if they induce heritable mutations in the germ cells of humans. Classification in Category 1B is based on: Positive result(s) from in vivo heritable germ cell mutagenicity tests in mammals; or Positive result(s) from in vivo somatic cell mutagenicity test in mammals, in combination with some evidence that the substance has potential to cause mutations to germ cells. It is possible to derive this supporting evidence from mutagenicity/genotoxicity tests in germ cells in vivo, or by demonstrating the ability of the substance or its metabolite(s) to interact with the genetic material of germ cells; or Positive results from tests showing mutagenic effects in the germ cells of humans, without demonstration of transmission to progeny; for example, an increase in the frequency of aneploidy in sperm cells of exposed people.

Germ cell mutagenicity Category 2 Criteria Substances which cause concern for humans owing to the possibility that they may induce heritable mutations in the germ cells of humans Classification in Category 2 is based on: Positive evidence obtained from experiments in mammals and/or in some cases from in vitro experiments, obtained from: Somatic cell mutagenicity tests in vivo, in mammals; or Other in vivo somatic cell genotoxicity tests which are supported by positive results from in vitro mutagenicity assays. Substances which are positive in in vitro mammalian mutagenicity assays, and which also show chemical structure activity relationship to known germ cell mutagens, should be considered for classfication as Category 2 mutagens.

Classification of a mixture: Germ cell mutagenicity Ingredient classified as: Concentration limits triggering classification of a mixture as: Category 1A mutagen Category 1B mutagen Category 1A mutagen 0.1% - Category 1B mutagen - 0.1% Category 2 mutagen Category 2 mutagen - - 1.0% -

Label elements Category 1A Category 1B Category 2 H340: May cause genetic defects H341: Suspected of causing genetic defects Danger Warning

Carcinogenicity Evidence that exposure increases the incidence of tumours Non-threshold Genotoxic carcinogens Threshold Non-genotoxic carcinogens

Normal cell DNA damage Multistage carcinogenesis Apoptosis (programmed cell death) Mutagen Repair Initiation Initiated Mutations cell Promotion Neoplastic Angiogenesis cell Progression Tumour

Important factors to consider Confounding effects of excessive toxicity localized effects Effects only at very high dose levels exceeding Maximal Tolerated Dose (MTD) (e.g. >10% retardation in weight gain) Specific tumour types e.g. forestomach tumours following administration by gavage of an irritating or corrosive, non-mutagenic chemical Unusual tumour types or tumours occuring with reduced latency may add to the weight of evidence (even if not statistically significant) Unique mechanism e.g. lack of human relevance of kidney tumours in male rats associated with compounds causing kidney damage (α-2-microglobulin nephropathy)

Carcinogenicity testing Duration 18-30 months (mice or rats) Male and female animals 1-2 Control groups 3 dose groups 50 animals/group 400-500 animals total

Category Category 1 Category 1A Category 1B Carcinogenicity Criteria Known or presumed human carcinogens A substance is classified in Category 1 for carcinogenicity on the basis of epidemiological and/or animal data. A substance may be further distinguished as: Known to have carcinogenic potential for humans, classification is largely based on human evidence Presumed to have carcinogenic potential for humans, classification is largely based on animal evidence The classification in Category 1A or 1B is based on strength of evidence together with additional considerations. Such evidence may be derived from: Human studies that establish a casual relationship between human exposure to a substance and the development of cancer (known human carcinogen); or Animal experiments for which there is sufficient evidence to demonstrate animal carcinogenicity (presumed human carcinogen) In addition, on a case-by-case basis, scientific judgement may warrant a decision of presumed human carcinogenicity derived from studies showing limited evidence of carcinogenicity in humans together with limited evidence of carcinogenicity in experimental animals.

Carcinogenicity Category Category 2 Suspected human carcinogen Criteria The placing of a substance in Category 2 is done on the basis of evidence obtained from human and/or animal studies, but which is not sufficiently convincing to place the substance in Category 1, based on strength of evidence together with additional considerations. Such evidence may be derived either from limited evidence of carcinogenicity in human studies or from limited evidence of carcinogenicity in animal studies.

Classification of a mixture: Carcinogenicity Ingredient classified as: Concentration limits triggering classification of a mixture as: Category 1 carcinogen Category 2 carcinogen Category 1A Category 1B Category 1A carcinogen 0.1% - Category 1B carcinogen - 0.1% Category 2 carcinogen - - - 0.1%* 1.0%** *If a Category 2 carcinogen is present in the mixture at a concentration between 0.1 and 1.0% an SDS would be required. A label may be required in some jurisdictions **If a Category 2 carcinogen is present in the mixture at a concentration of 1.0% both an SDS and a label would generally be expected

Label elements Category 1A Category 1B Category 2 H350: May cause cancer H351: Suspected of causing cancer Danger Warning

www.kemi.se Reproductive toxicity

Reproductive toxicity Effects on fertility Genetic changes in germ cells Infertility Alterations in sex hormone levels Ovarian dysfunction Sexual behaviour Effects on development Growth Organ and tissue development and function Acquisition of normal organ function between conception and puberty May cause Death (in utero or postnatally) Structural defects Growth retardation Functional defects E.g. neurodevelopment Childhood cancer Confounder: maternal toxicity

WHO/UNEP (2013): State of the Science of Endocrine Disrupting Chemicals 2012

Reproductive toxicity Category Criteria Category 1 Known or presumed human reproductive toxicant Category 1A Category 1B Substances are classified in Category 1 for reprodutive toxicity when they are known to have produced an adverse effect on sexual function and fertility, or on development, in humans, or when there is evidence from animal studies, possibly supplemented with other information, to provide a strong presumption that the substance has the capacity to interfere with reproduction in humans. The classification of a substance is further distinguished on the basis of whether the evidence for classification is primarily from human data or from animal data. Known human reproductive toxicant Largely based on evidence from humans Presumed human reproductive toxicant Largely based on data from animal studies. Data should provide clear evidence of an adverse effect on sexual function and fertility or on development in the absence of other toxic effects, or if occuring together with other toxic effects the adverse effect on reproduction is considered not to be a secondary non-specific consequence of other toxic effects. However, when there is mechanistic information that raises doubt about the relevance of the effect for humans, classification in Category 2 may be more appropriate.

Reproductive toxicity Category Category 2 Criteria Suspected human reproductive toxicant Substances are classified in Category 2 for reproductive toxicity when there is some evidence from human or experimental animals, possibly supplemented with other information, of an adverse effect on sexual function and fertility, or on development, and where the evidence is not sufficiently convincing to place the substance in Category 1 The effects should have been observed in the absence of other toxic effects, or if occuring together with other toxic effects the adverse effect on reproduction is considered not to be a secondary non-specific consequence of other toxic effects. If deficiencies in the study make the quality of evidence less convincing, Category 2 could be the more appropriate classification.

Additional category Effects on or via lactation Effects on or via lactation are allocated to a separate single category. It is recognised that for many substances there is no information on the potential to cause adverse effetcs on the offspring via lactation. However, substances which are absorbed by women and have been shown to interfere with lactation, or which may be present (including metablites) in breast milk in amounts sufficient to cause concern for the health of a breastfed child, should be classified and labelled to indicate this property hazardous to breastfed babies. This classifiaction can be assigned on the basis of: a) Human evidence indicating a hazard to babies during the lactation period; and/or b) Results of one or two generation studies in animals which provide clear evidence of adverse effect in the offspring due to transfer in the milk or adverse effects on the quality of the milk; and/or c) Absorption, metabolism, distribution and excretion studies that indicate the likelihood that the substance is present in potentially toxic levels in breast milk.

Classification of a mixture: reproduction toxicity Ingredient classified as: Category 1A reproductive toxicant Concentration limits triggering classification of a mixture as: Category 1 reproductive toxicant Category 1A Category 1B Category 2 reproductive toxicant Effects on or via lactation 0.1% a - - - 0.3% b Category 1B reproductive toxicant - 0.1% a - 0.3% b Category 2 reproductive toxicant - - 0.1% c - 3.0% d Effects on or via lactation - - - a between 0.1 and 0.3% an SDS would be required. A label may be required in some jurisdictions b at a concentration of 0.3% both an SDS and a label would generally be expected c between 0.1 and 3.0% an SDS would be required. A label may be required in some jurisdictions d at a concentration of 3.0% both an SDS and a label would generally be expected 0.1% a 0.3% b

Label elements Category 1A Category 1B Category 2 Additional category H360: May damage fertility or the unborn child H361: Suspected of damaging fertility or the unborn child H362: May cause harm to breast-fed children Danger Warning

Highly Hazardous Pesticides Working definition by FAO/WHO Joint Meeting* WHO: Classes 1a or 1b GHS: Mutagenicity Cat. 1A or 1B GHS: Carcinogenicity Cat. 1A or 1B GHS: Reproductive toxicity Cat. 1A or 1B Stockholm Convention: Annex A or B or Annex D, para 1 Montreal protocol High incidence of severe or irreversible adverse effects *http://www.fao.org/fileadmin/templates/agphome/documents/pests_pesticides/code/report.pdf

Some hot topics in toxicology Endocrine disruptors Combined exposures ( cocktail effect ) Non-monotonic dose-response Nanomaterials Non-animal testing

Some useful links http://www.oecd-ilibrary.org/environment/oecd-guidelines-for-the-testing-ofchemicals-section-4-health-effects_20745788 OECD: Test guidelines: Health effects http://echa.europa.eu/web/guest/information-on-chemicals/cl-inventory-database ECHA: Classification and Labelling Inventory database http://www.echemportal.org/echemportal/participant/page.action?pageid=0 echemportal: The OECD Global Portal to Information on Chemical Substances

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