Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

100.25 mg/m³

Most sensitive endpoint:

developmental toxicity / teratogenicity

Route of original study:

By inhalation

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

25

Dose descriptor starting point:

NOAEC

Value:

3 562.5 mg/m³

Modified dose descriptor starting point:

NOAEC

Value:

2 506.2 mg/m³

Explanation for the modification of the dose descriptor starting point:

The starting point NOAEC (neonatal systemic toxicity) obtained in the animal study according to OECD 443 was modified accounting for the differences between experimental and human exposure conditions. The conversion of an inhalatory rat NOAEC into a corrected inhalatory NOAEC to assess human inhalatory exposure was performed: to take into account the differences in exposure duration a factor of 0.75 was included (6 h for the rodent to 8 h for the worker), moreover a factor of 1.4 is included (the differences in exposure duration rat 7 d/wk and workers only 5 d/wk), additionally a factor of 0.67 is included (6.7 m³, as a respiratory volume for humans for an 8-hour shift divided by 10 m³, the respiratory volume for worker by light activity).

corrected NOAEC = 3562.5 mg/m³ * 0.75 * 1.4 * 0.67 = 2506.2 mg/m³

AF for dose response relationship:

1

Justification:

default

AF for differences in duration of exposure:

2

Justification:

default value for subchronic to chronic

AF for interspecies differences (allometric scaling):

1

Justification:

No allometric scaling is applied because a ventilation rate and food intake directly depends on the basal metabolic rate and has already been scaled according to the allometric principle (inhalation study and inhalation exposure in humans).

AF for other interspecies differences:

2.5

Justification:

default for remaining interspecies differences

AF for intraspecies differences:

5

Justification:

default value for workers

AF for the quality of the whole database:

1

Justification:

default value (GLP guideline study)

AF for remaining uncertainties:

1

Justification:

there are no further remaining uncertainties to be taken into account

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

196.3 mg/m³

Most sensitive endpoint:

acute toxicity

Route of original study:

By inhalation

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

37.5

Dose descriptor starting point:

LOAEC

Value:

10 990 mg/m³

Modified dose descriptor starting point:

LOAEC

Value:

7 363 mg/m³

Explanation for the modification of the dose descriptor starting point:

An acute inhalation LOAEC (6-hr) value in the rat of 3810 mg/m³ was used as the starting point for an acute inhalation systemic worker DNEL. A LOAEL has been assigned based on the presence of only minimal and reversible CNS effects immediately following dosing in rats. This LOAEC has been converted into an acute 15-minute value using Habers law = 10990 mg/m³ and was subsequently corrected by a factor of 0.67 accounting for differences in respiratory volume of workers (6.7 m³, as a respiratory volume for humans for an 8-hour shift divided by 10 m³, the respiratory volume for worker by light activity) = 7363 mg/m³

AF for dose response relationship:

3

Justification:

default factor to account for use of a LOAEC instead on a NOAEC

AF for interspecies differences (allometric scaling):

1

Justification:

No allometric scaling is applied because a ventilation rate and food intake directly depends on the basal metabolic rate and has already been scaled according to the allometric principle (inhalation study and inhalation exposure in humans).

AF for other interspecies differences:

2.5

Justification:

default for remaining interspecies differences

AF for intraspecies differences:

5

Justification:

default for workers

AF for the quality of the whole database:

1

Justification:

default

AF for remaining uncertainties:

1

Justification:

there are no further remaining uncertainties to be taken into account

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

14.2 mg/kg bw/day

Most sensitive endpoint:

developmental toxicity / teratogenicity

Route of original study:

By inhalation

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

100

Dose descriptor starting point:

NOAEC

Value:

3 562.5 mg/m³

Modified dose descriptor starting point:

NOAEL

Value:

1 421.4 mg/kg bw/day

Explanation for the modification of the dose descriptor starting point:

The starting point for a long-term systemic DNEL (dermal) value for workers was the NOAEC value of 3562.5 mg/m³ from the extended one generation reproductive toxicity study in rats via inhalation (Coder, P., 218; according to OECD 443). The starting point NOAEC obtained in the animal study was corrected accounting for the differences between experimental and human exposure conditions. The conversion of an inhalatory rat NOAEC into a corrected dermal NOAEL to assess human dermal exposure was performed: to take into account the differences in respiratory volume of rats versus humans a factor of 0.38 was included; to take into account the differences in exposure duration a factor of 0.75 was included (6 h for the rodent to 8 h for the worker), moreover a factor of 1.4 is included (the differences in exposure duration rat 7 d/wk and workers only 5 d/wk).

AF for dose response relationship:

1

Justification:

default (GLP-guideline study)

AF for differences in duration of exposure:

2

Justification:

default for subchronic to chronic

AF for interspecies differences (allometric scaling):

4

Justification:

default for rats

AF for other interspecies differences:

2.5

Justification:

default for remaining interspecies differences

AF for intraspecies differences:

5

Justification:

default for workers

AF for the quality of the whole database:

1

Justification:

default

AF for remaining uncertainties:

1

Justification:

there are no further remaining uncertainties to be taken into account

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

EXPOSURE DNEL DERIVATIONS FOR METHYLiso-AMYL KETONE

(CAS No. 110-12-3)

I. Introduction

Methyliso-amyl ketone (5-methylhexan-2-one) is currently listed as a harmful substance in the Annex I of the EU Dangerous Substance Directive 67/548/EEC and is rated Xn; R20 (Harmful by inhalation). Similarly, it is listed in Annex VI to EC Regulation 1272/2008 as Acute Toxicity, Category 4 (H332: Harmful if inhaled). It is not a Category 1 or 2 mutagen or carcinogen. Thus, DNEL/DMEL values for these endpoint were not derived. However, recently a reproductive hazard has been identified and the substance is classified and labelled as Repro Cat. 2. DNELvalues for these endpoints were derived (OECD 443, rats, whole body inhalation; NOAEC (maternal toxicity, systemic) = 1500 ppm = 7125 mg/m³; NOAEC (neonatal toxicity, systemic) = 75ß ppm = 3562.5 mg/m³. Summarized in Table 1 are the critical preliminary DNEL values derived for methyl isoamyl ketone.

Table 1: Critical Preliminary DNEL Values for Methyl Isoamyl Ketone

Exposure Pattern	DNEL
Exposure Pattern	Workers	General Population
Acute – inhalation, systemic	196.3 mg/m³(1)	146.5 mg/m³
Acute – inhalation, local	not calculated	not calculated
Acute – dermal, systemic	not calculated	not calculated
Acute – dermal, local	not calculated (2)	not calculated (2)
Acute – oral, systemic	not relevant (3)	not relevant (3)
Long-term – inhalation, systemic	100.25 mg/m³	17.8125 mg/m³
Long-term – inhalation, local	not relevant (4)	not relevant (4)
Long-term – dermal, systemic	14.2 mg/kg bwt/day	5.12 mg/kg bwt/day
Long-term – dermal, local	No data	No data
Long-term – oral, systemic	not relevant (3)	5.12 mg/kg bwt/day

1 Chapter 8 of the REACH TGD (Appendix R.8-8) indicates that acute exposure DNEL values are not normally required. Additionally, the lack of acute oral or dermal toxicity would indicate MiAK is non-hazardous under acute exposure scenarios. Given the assignment of R20 and Acute Toxicity (Category 4, H332), acute inhalation DNEL values (15-minute exposures) were derived for MiAK.

2 Assessment based on a lack of significant skin irritation.

3 It is assumed that there will be no acute or long-term oral exposures to MiAK in worker populations or acute oral exposures in consumer populations.

4 Eye and respiratory irritation thresholds are assumed to exceed thresholds for chronic systemic effects.

Summarized in Table 2 are the critical toxicological studies and values identified for MiAK.

Table 2: Dose descriptor for key studies of toxicological concern

Endpoint	Descriptor	Key Reference	Associated Effects/Comments
Acute Toxicity
Inhalation, rat 802, 1603, 3207 and 5878 ppm (measured)	LOAEL 802 mg/m³ (3810 mg/m³)	Katz, G.V., Basic Toxicity of Methyl Isoamyl Ketone, Eastman Kodak Company, Report No. 104791R, 1978	Reversible CNS effects; initial weight loss with weight gains in surviving animals. A LOAEL has been assigned based on the presence of only minimal and reversible CNS effects immediately following dosing.
Repeated Dose Toxicity
Inhalation, rat 212, 1025 and 2079 ppm (measured)	NOAEL = 212 ppm (1007 mg/m³)	Katz, G.V., Renner, C.J. and Terhaar, C.J. (1986). Subchronic inhalation toxicity of methyl isoamyl ketone in rats. Fundam. Appl. Toxicol. 6:498-505.	Increased absolute/relative liver/kidney weights; hyalin droplet degen. of kidney (males); minor/moderate tubular epithelium regeneration; hepatocyte hypertrophy
IOEL
NOAEL from study of Katz et al. (1986); assessment factor of 10	IOEL (8-hr TWA) = 20 ppm (95 mg/m³)	European Commission Directive 2000/39/CE	Refer to study of Katz et al. (1986)

Reproductive toxicity
NOAEL from study of Corder, R. (2018);	NOAEL (neonatal toxicity, systemic, rats, whole body inhalation, OECD 443) = 750 ppm corresponding to ca. 3562.5 mg/m³	Coder, R., OECD 443 with Methyl isoamyl ketone	Refer to study of Corder, P. (2018)

II. Descriptions and Rationale for DNEL Derivations

1. Acute toxicity

The acute oral toxicity of methyl isoamyl ketone (MiAK) is low with LD₅₀values reported in experimental animals of 3200 mg/kg bwt. In all studies reported, clinical signs indicating reversible effects on the central nervous system included one or more of the following: weakness, ataxia, rough hair coats, excitement, vasodilatation, slight tremors, restlessness, and prostration. All surviving animals gained weight normally.

The acute dermal toxicity of MiAK is low. In two studies, doses of 5, 10 or 20 mL/kg bwt (equivalent to 4, 8 and 16 g/kg bwt) MiAK were applied to the depilated abdomens of guinea pigs under occluded contact for 24-hours. There were no deaths observed in these studies. Except for weight loss at the two higher dose levels, there were no clinical signs indicating absorption or systemic toxicity. The dermal LD₅₀is considered to be > 20 mL/kg bwt (16 g/kg bwt).

In a key study, groups of 4 male rats were exposed via whole body inhalation to target concentrations of 800, 1600, 3200 or 6400 ppm MiAK (measured concentrations: 802, 1603, 3207 and 5878 ppm) for 6 hours. All rats at the highest exposure concentration died within 2.5 hours of the start of exposure. One of four rats at the 3207 ppm concentration died just prior to termination of exposure. At the highest concentration tested, eye irritation and narcosis were observed in all animals prior to death. Reversible central nervous system effects were also observed in survivors at all exposure concentrations. Although surviving rats in the 3207 ppm group lost weight over the first 3 days, subsequent weight gain was normal and all other rats gained weight normally over the 14-day observation period. A LOAEL of 802 ppm (3810 mg/m³) was set for this study based on the minimal appearance of reversible CNS effects immediately after exposures. These data are supported by four lesser studies in which groups of 3 rats were exposed via whole body inhalation to 3.88 mg/L (948 ppm), 21 mg/L (4000 ppm), 29 mg/L (5500 ppm), or 31.57 mg/L (7718 ppm) for 6 or 7 hours. At 7718 ppm, all animals exhibited signs of central nervous system depression and died shortly after hour 4 of exposure. All animals survived exposure at 5500 ppm for 7 hours but exhibited central nervous system effects including loss of coordination, prostration, and labored breathing. One of three animals exposed to 4000 ppm died at 5 hours into the 6-hr exposure period. Reversible central nervous system effects were observed in survivors. No deaths or clinical signs were observed in the 948 ppm exposure group. MiAK is considered to be harmful by the inhalation route.

In the case of acute worker and general population inhalation exposures, the LOAEL (6-hr) value of 3810 mg/m³, measured in rats, was selected as the appropriate dose descriptor for derivation of acute systemic inhalation DNEL values. No additional local or systemic acute DNEL values were derived. It should be noted that there is no IOEL STEL value assigned for MiAK that would be appropriate for the derivation of acute inhalation systemic DNEL values for workers.

2. Irritation/Sensitization

In experimental animals, MiAK caused only slight and transient dermal irritation when tested in guinea pigs following 24-hr occluded contact. Repeated dermal applications of the test material did not significantly exacerbate the irritative effects. Application of MiAK to the eyes of rabbits (either washed or unwashed) produced at most only slight irritation. In a primary dermal irritation study, slight irritation and some signs of desquamation were observed following 24-hr occluded contact. Repeat application of the test material did not significantly exacerbate the irritative effects. There were no clinical signs indicative of respiratory tract irritation in rats following acute inhalation exposure to up to 7700 ppm or subacute or subchronic exposure to up to 2000 ppm. The material was also not a skin sensitizer when tested in guinea pigs. Based on a weight-of-evidence assessment, MiAK would not be classified as a respiratory tract irritant.

3. Repeat dose toxicity

MiAK administered to rats via whole body inhalation at concentrations up to 2000 ppm for a total of 69 exposures and spanning 96 days produced treatment-related organ weight changes and microscopic effects in the livers and kidneys of both male and female rats at 1000 and 2000 ppm (Katz et al., 1986). The NOEC for systemic toxicity was determined to be 200 ppm (212 ppm measured) based on a lack of treatment-related organ weight changes and microscopic effects observed in the livers and kidneys at the higher exposure concentrations. Increased kidney weights were not accompanied by changes in clinical chemistry parameters. Hyaline droplet degeneration, observed in the proximal convoluted tubular epithelium of the kidneys of male rats, was not considered relevant to human risk assessment. Regeneration of tubular epithelium in both sexes was minor and neither kidney lesion impaired the health of the animals. Liver weight changes were accompanied by minor to moderate hepatocyte hypertrophy, but no changes in serum clinical chemistries were observed which would be suggestive of liver damage. No hepatocyte hyperplasia was observed by the inhalation route of exposure.

MiAK when administered by gavage to rats at a dose level of 2000 mg/kg bwt/day, five days/week over 90 days, produced statistically significant increases in two liver enzyme values, GOT and GPT, and microscopic changes in the stomach, liver and kidney. Changes in the stomach were thought to be due to irritation following prolonged contact with the test material and, as such, would not be a true indication of a target organ effect. The observed kidney effects are consistent with those seen with exacerbation of a commonly occurring effect observed in male laboratory rats following chemical exposure and which have not been observed in humans. Increases in liver enzymes, increases in absolute and relative liver weights, and histopathologic changes in the liver were indicative of the liver as a target organ for exposure to methyl isoamyl ketone by the oral route and consisted of effects described as pre-neoplastic, hepatocyte necrosis and bile-duct hyperplasia. Although liver effects were observed at extremely high oral doses and only minor liver effects were observed at comparable inhalation exposures. A NOEL or NOAEL could not be assigned to this oral gavage study.

The IOEL value (8-hr TWA) of 95 mg/m³had been selected in the past as the long-term inhalation systemic DNEL value for workers. Since the publication of this IOEL, there has been new scientific information on the toxicology of MiAK (OECD 443 study results, see below) that would invalidate the use of this for the setting of a worker DNEL value. Formerly, the NOAEC value of 212 ppm (1007 mg/m³) (Katz et al., 1986) had been selected as the starting dose descriptor for the derivation of a long-term inhalation systemic DNEL value for the general population. After correction for route-of-exposure, this same NOAEC value had served as the starting point for both the oral and dermal long-term systemic DNEL values for the general population and for the dermal long-term systemic DNEL for workers. These DNELs however, are now derived of the NOAEC value obtained from the recent OECD 443 study in rats.

4. Genotoxicity/carcinogenicity

MiAK was negative in anin vitro Ames mutagenicity assay conducted at plate incorporation levels of up to 5,000mg/plate. In a mouse lymphoma L5178Y mammalian cell gene mutation assay, MiAK was negative at concentrations up to 1200 µg/mL in the presence and absence of mammalian metabolic activation and showed no evidence of inducing chromosomal aberrations or polyploidy when tested in an in vitro cytogenetics test with Chinese hamster ovary cells (CHO cells) with and without metabolic activation up to a concentration of 1200 µg/mL. MiAK has not been tested for genotoxicity in vivo.

MiAK has not been tested for carcinogenicity.

5. Reproductive/developmental toxicity

In an OECD 421 reproductive/developmental toxicity screening study in rats, MiAK was administered by inhalation (whole body) at up to 5.0 mg/L for 6 hr/day, 7 days/week for a total of 51 (males) and 35-41 (females) consecutive days during premating, mating, pregnancy and early lactation. There were no gross or microscopic effects on the reproductive organs of either sex and there were no treatment-related effects on any reproductive parameters. Similarly, in 13-week exposure studies in rats by the oral and inhalation routes, there were no reported gross or microscopic changes in the reproductive organs of either sex at oral gavage doses up to 2000 mg/kg bwt/day or inhalation exposures up to 2000 ppm (6 hr/day, 5 days/week for a total of 69 exposures).

In and OECD 414 prenatal developmental toxicity test, pregnant female rats were exposed by whole-body inhalation to the test article at target concentrations(actual) of 380(379), 750(751) and 1500(1495) ppm. Animals were exposed from gestation day 0 to gestation day 19. There was a decreased in food consumption and body weight in the 1500 ppm dams, along with a reduced startle reflex at 1500 and 750 ppm. The NOAEL for maternal toxicity was 380 ppm and 750 ppm was the NOAEL for embryo/fetal toxicity based on reduced pup weights at 1500 ppm. There were no significant findings of any developmental defects in the pups at any dose.

In and OECD 443 extended one-generation reproductive toxicity test, rats were exposed by whole-body inhalation to the test article at target concentrations(actual) of 380, 750 and 1500 ppm. Overall mean analyzed exposure concentrations were 380, 750, and 1499 ppm for the F0 generation and 379, 750, and 1497 ppm for the F1 generation. Animals in the parental (F0) generation were exposed via whole-body inhalation for 6 hours daily for 70 consecutive days prior to mating and continuing through the day prior to euthanasia. Maternal exposure was suspended from Gestation Day 20 through Lactation Day 4 to prevent confounding effects on parturition and maternal nursing and nesting behavior. The offspring in the F1 generation were potentially exposed in utero during gestation. Direct offspring exposure during the pre-weaning period was demonstrated on a concurrent study, where quantifiable levels of MiAK were obtained in maternal milk and plasma obtained from dams exposed at the same target exposure concentrations (380 to 1500 ppm). Offspring selected to constitute the F1 generation were exposed beginning at weaning (Postnatal Day 28) and continuing through the day prior to euthanasia. Due to the lower body weight gains noted for the F1 pups during the neonatal and pre-weaning periods (see below), F1 animals assigned to Cohort 1B for follow-up reproductive assessments were bred to obtain an F2 generation. All procedures for F1 animals assigned to Cohort 1B were generally the same as those for the F0 generation/F1 litters, including suspension of exposure for the F1 generation maternal animals from Gestation Day 20 through Lactation Day 4. The offspring in the F2 generation were not directly exposed to test substance atmospheres but were potentially exposed in utero, and through maternal milk during the pre-weaning period.

The following parameters and end points were evaluated in this study: clinical signs, body weights, body weight gains, food consumption, estrous cycles, reproductive performance, parturition, litter viability and survival, anogenital distance, areolae/nipple anlagen, developmental landmarks, thyroid hormones, clinical pathology, gross necropsy findings, spermatogenic endpoints, organ weights, and histopathologic examinations.

The NOAEL for maternal toxicity was 1500 ppm and 750 ppm was the NOAEL for embryo/fetal toxicity based on reduced pup weights at 1500 ppm.

III. Mode of Action Considerations

For the critical studies identified in Table 2, a threshold mode of action was assumed.

IV. Modification of Relevant Dose Descriptors to the Correct Starting Point

Summarized in Table 3 are corrected dose descriptors derived from those in Table 2 (see Guidance Document, Chapter R.8, Appendix R.8-2).

Table 3: Corrected dose descriptors for key studies of toxicological concern

Endpoint	Most relevant dose descriptors		Corrected dose descriptors (Workers)
	Local	Systemic	Local	Systemic
Acute Toxicity
-oral	NA	NA	NA	NA
-dermal	NC	NC	NC	NC
-inhalation	NC	LOAEC (6-hr) = 3810 mg/m³	NC	7363 mg/m³
Repeated Dose Toxicity
-oral	NA	NA	NA	NA
-dermal	NC	NOAEC = 750 ppm = 3562.5 mg/m³	NC	1421.4 mg/kg bwt/day
-inhalation	NC	NOAEC = 750 ppm = 3562.5 mg/m³	NC	2506.2 mg/m³
Endpoint	Most relevant dose descriptors		Corrected dose descriptors (General Population)
	Local	Systemic	Local	Systemic
Acute Toxicity
-oral	NC	NC	NC	NC
-dermal	NC	NC	NC	NC
-inhalation	NC	LOAEC (6-hr) = 3810 mg/m³	NC	10,990 mg/m³
Repeated Dose Toxicity
-oral	NC	(use inhal. data)	NC	1024.2 mg/kg bwt/day
-dermal	NC	(use inhal. data)	NC	1024.2 mg/kg bwt/day
-inhalation	NC	NOAEC = 750 ppm = 3562.5 mg/m³	NC	890.625 mg/m³

NA - not applicable NC - not calculated

Workers

1. Acute

In the case of worker exposures, the oral route of exposure was not considered. Based on the low acute toxicity of MiAK by the dermal route, the dermal route of exposure was also not considered. An acute inhalation LOAEC (6-hr) value in the rat of 3810 mg/m³was used as the starting point for an acute inhalation systemic worker DNEL (see Guidance Document, Chapter R.8, Appendix R.8-8). Duration of exposure correction was applied based on an acute (15-minute) exposure and a correction for an increased respiratory volume for light work conditions (see Guidance Document, Chapter R.8.4.2). For workers, the 6-hour starting value (3810 mg/m³) was corrected to a 15-minute exposure duration using Haber’s Law (n=3) giving a modified dose descriptor as follows:

LOAEC_worker= 10,990 mg/m³(Haber’s Law corrected value)

= 10,990 mg/m³x (6.7 m³/8 hours / 10 m³/8 hours) (Light work corrected)

or 7363 mg/m³

2. Long-Term

dermal

The starting point for a long-term (repeated dose) dermal systemic DNEL value for workers was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (8 hours/d) x (7 d/week / 5 d/week)

or, NOAEC_corr= 3740.625 mg/m³

Conversion to a dermal equivalent dose is performed as discussed in the Guidance Document, Appendix R.8-2, Example B.4.

NOAEL_dermal,rat= NOAEC_corrx sRV_rat(0.38 m³/kg bwt for 8 hour exposure)

or, 1421.4 mg/kg bwt/day

inhalation

The starting point for a long-term (repeated dose) inhalation systemic DNEL value for workers was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure and for an increased respiratory volume for light work conditions (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (8 hours/d) x (6.7 m³/8 hours / 10 m³/8 -hours - light work corrected) * (7 d/week / 5 d/week)

or, NOAEC_corr= 2506.2 mg/m³

General Population

1. Acute

In the case of general population exposures, the oral and dermal routes of exposure were not considered based on the low acute toxicity of MiAK by either route. An acute inhalation LOAEC (6-hr) value in the rat of 3810 mg/m³was used as the starting point for an acute inhalation systemic general population DNEL (see Guidance Document, Chapter R.8, Appendix R.8-8). Duration of exposure correction was applied based on an acute (15-minute) exposure (see Guidance Document, Chapter R.8.4.2). For the general population, the 6-hour starting value (3810 mg/m³) was corrected to a 15-minute exposure duration using Haber’s Law (n=3) giving a modified dose descriptor as follows:

LOAEC_gen.pop.= 10,990 mg/m³(Haber’s Law corrected value)

2. Long-Term

oral and dermal

The starting point for a long-term (repeated dose) oral or dermal systemic DNEL value for the general population was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (24 hours/d)

or, NOAEC_corr= 890.625 mg/m³

Conversion to a dermal equivalent dose is performed as discussed in the Guidance Document, Appendix R.8-2, Example B.4.

NOAEL_dermal,rat= NOAEC_corrx sRV_rat(1.15 m³/kg bwt for 24 hour exposure)

or, 1024.2 mg/kg bwt/day

inhalation

The starting point for a long-term (repeated dose) inhalation systemic DNEL value for general population was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (24 hours/d)

or, 890.8125 mg/m³

V. Application of Assessment Factors to the Corrected Dose Descriptors

Summarized in Table 4 are endpoint-specific DNEL values for worker and general population exposures to MiAK derived from the corrected dose descriptors found in Table 3.

Table 4: Endpoint-specific DNEL values for MiAK

Endpoint	Corrected dose descriptor		Overall AF[1]	Endpoint-specific DNELs (Workers)
	Local	Systemic		Local	Systemic
Acute Toxicity
-oral	NA	NA	NA	NA	NA
-dermal	NC	NC	NC	NC	NC
-inhalation	NC	7363 mg/m³	15	NC	196.3 mg/m³
Repeated Dose Toxicity
-oral	NA	NA	NA	NA	NA
-dermal	NC	1421.4 mg/kg bwt/day	100	NC	14.2 mg/kg bwt/day
-inhalation	NC	2506.2mg/m³	25	NC	100.25 mg/m³
Endpoint	Corrected dose descriptor		Overall AF¹	Endpoint-specific DNELs (General Population)
Acute Toxicity
-oral	NC	NC	NC	NC	NC
-dermal	NC	NC	NC	NC	NC
-inhalation	NC	10,990 mg/m³	75	NC	146.5 mg/m³
Repeated Dose Toxicity
-oral	NC	1024.2 mg/kg bwt/day	200	NC	5.12 mg/kg bwt/day
-dermal	NC	1024.2 mg/kg bwt/day	200	NC	5.12 mg/kg bwt/day
-inhalation	NC	890.625 mg/m³	50	NC	17.8125 mg/m³

NA - not applicable NC - not calculated

Workers

1. Acute

In the case of acute inhalation worker exposure, the starting dose descriptor did not represent a clear NOAEL and thus an assessment factor (AF) of 3 was applied. An additional AF of 5 was applied (intraspecies, sensitive worker) to give an overall AF of 15 (Table 4).

2. Long-Term

dermal

An AF of 4 was applied to the corrected dose descriptor for dermal long-term systemic effects in workers (interspecies differences, allometric scaling). In addition, an AF of 2 (subchronic to chronic). an AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive worker) AF of 5 were applied. The overall AF obtained was 100 (Table 4).

inhalation

An AF of 2 was applied to the corrected dose descriptor for inhalation long-term systemic effects in workers (subchronic to chronic), an AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive worker) AF of 5 were applied. The overall AF obtained was 25 (Table 4).

General Population

1. Acute

In the case of acute inhalation exposure of the general population, the starting dose descriptor did not represent a clear NOAEL and thus an assessment factor of 3 was applied. An additional assessment factor (AF) of 2.5 for remaining interspecies differences and an AF of 10 was applied (intraspecies, sensitive individual) to give an overall AF of 75 (Table 4).

2. Long-Term

oral and dermal

An AF of 4 was applied to the corrected dose descriptor for oral long-term (repeated dose) systemic effects in the general population (interspecies differences, allometric scaling). In addition, an AF of 2 (subchronic to chronic), an AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive individual) AF of 10 were applied. The overall AF obtained was 200 (Table 4). These same AF values apply to the case of dermal exposure.

inhalation

No additional intraspecies (allometric scaling) factor was applied to the corrected dose descriptor for inhalation long-term systemic effects in the general population. An AF of 2 (subchronic to chronic), An AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive individual) AF of 10 were applied. The overall AF obtained was 50 (Table 4).

SUMMARY

The calculated DNEL values for MiAK are summarized in Table 1 at the beginning of this document.

[1]Overall assessment factors were assigned based on the ECHA guidelines.

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

17.812 mg/m³

Most sensitive endpoint:

developmental toxicity / teratogenicity

Route of original study:

By inhalation

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

50

Dose descriptor starting point:

NOAEC

Value:

3 562.5 mg/m³

Modified dose descriptor starting point:

NOAEC

Value:

890.625 mg/m³

Explanation for the modification of the dose descriptor starting point:

The starting point NOAEC (neonatal systemic toxicity) obtained in the animal study according to OECD 443 was modified accounting for the differences between experimental and human exposure conditions. The conversion of an inhalatory rat NOAEC into a corrected inhalatory NOAEC to assess human inhalatory exposure was performed: to take into account the differences in exposure duration a factor of 0.25 was included (6 h for the rodent to 24 h for the general population).

corrected NOAEC = 3562.5 mg/m³ * 0.25 * = 890.625 mg/m³

AF for dose response relationship:

1

Justification:

default

AF for differences in duration of exposure:

2

Justification:

default value for subchronic to chronic

AF for interspecies differences (allometric scaling):

1

Justification:

No allometric scaling is applied because a ventilation rate and food intake directly depends on the basal metabolic rate and has already been scaled according to the allometric principle (inhalation study and inhalation exposure in humans).

AF for other interspecies differences:

2.5

Justification:

default for remaining interspecies differences

AF for intraspecies differences:

10

Justification:

default value for general population

AF for the quality of the whole database:

1

Justification:

default value (GLP guideline study)

AF for remaining uncertainties:

1

Justification:

there are no further remaining uncertainties to be taken into account

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

146.5 mg/m³

Most sensitive endpoint:

acute toxicity

Route of original study:

By inhalation

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

75

Dose descriptor starting point:

LOAEC

Value:

10 990 mg/m³

Modified dose descriptor starting point:

NOAEC

Value:

10 990 mg/m³

Explanation for the modification of the dose descriptor starting point:

An acute inhalation LOAEC (6-hr) value in the rat of 3810 mg/m³was used as the starting point for an acute inhalation systemic general population DNEL. A LOAEL has been assigned based on the presence of only minimal and reversible CNS effects immediately following dosing in rats. This LOAEC has been converted into an acute 15-minute value using Habers law = 10990 mg/m³.

AF for dose response relationship:

3

Justification:

default factor to account for use of a LOAEC instead on a NOAEC

AF for interspecies differences (allometric scaling):

1

Justification:

No allometric scaling is applied because a ventilation rate and food intake directly depends on the basal metabolic rate and has already been scaled according to the allometric principle (inhalation study and inhalation exposure in humans).

AF for other interspecies differences:

2.5

Justification:

default for remaining interspecies differences

AF for intraspecies differences:

10

Justification:

default for the general population

AF for the quality of the whole database:

1

Justification:

default

AF for remaining uncertainties:

1

Justification:

there are no further remaining uncertainties to be taken into account

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

5.12 mg/kg bw/day

Most sensitive endpoint:

developmental toxicity / teratogenicity

Route of original study:

By inhalation

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

200

Dose descriptor starting point:

NOAEC

Value:

3 562.5 mg/m³

Modified dose descriptor starting point:

NOAEL

Value:

1 024.2 mg/kg bw/day

Explanation for the modification of the dose descriptor starting point:

The starting point NOAEC (neonatal systemic toxicity) obtained in the animal study according to OECD 443 was modified accounting for the differences between experimental and human exposure conditions. The conversion of an inhalatory rat NOAEC into a corrected dermal NOAEC to assess human dermal exposure was performed: to take into account the differences in exposure duration a factor of 0.25 was included (6 h for the rodent to 24 h for the general population), additionally a factor of 1.15 is included (1.15 m³, as a respiratory volume for humans for a 24h-day).

corrected NOAEC = 3562.5 mg/m³ * 0.25 * 1.15 = 1024.2 mg/m³

AF for dose response relationship:

1

Justification:

default (GLP-guideline study)

AF for differences in duration of exposure:

2

Justification:

default for subchronic to chronic

AF for interspecies differences (allometric scaling):

4

Justification:

default for rats

AF for other interspecies differences:

2.5

Justification:

default for remaining interspecies differences

AF for intraspecies differences:

10

Justification:

default for the general population

AF for the quality of the whole database:

1

Justification:

default

AF for remaining uncertainties:

1

Justification:

there are no further remaining uncertainties to be taken into account

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

DNEL (Derived No Effect Level)

Value:

5.12 mg/kg bw/day

Most sensitive endpoint:

developmental toxicity / teratogenicity

Route of original study:

By inhalation

DNEL derivation method:

ECHA REACH Guidance

Overall assessment factor (AF):

200

Dose descriptor starting point:

NOAEC

Value:

3 562.5 mg/kg bw/day

Modified dose descriptor starting point:

NOAEL

Value:

1 024.2 mg/kg bw/day

Explanation for the modification of the dose descriptor starting point:

The starting point NOAEC (neonatal systemic toxicity) obtained in the animal study according to OECD 443 was modified accounting for the differences between experimental and human exposure conditions. The conversion of an inhalatory rat NOAEC into a corrected oral NOAEC to assess human oral exposure was performed: to take into account the differences in exposure duration a factor of 0.25 was included (6 h for the rodent to 24 h for the general population), additionally a factor of 1.15 is included (1.15 m³, as a respiratory volume for humans for a 24h-day).

corrected NOAEC = 3562.5 mg/m³ * 0.25 * 1.15 = 1024.2 mg/m³

AF for dose response relationship:

1

Justification:

default (GLP-Guideline study)

AF for differences in duration of exposure:

2

Justification:

default for subchronic to chronic

AF for interspecies differences (allometric scaling):

4

Justification:

default for rats

AF for other interspecies differences:

2.5

Justification:

default for remaining interspecies differences

AF for intraspecies differences:

10

Justification:

default for the general population

AF for the quality of the whole database:

1

Justification:

default

AF for remaining uncertainties:

1

Justification:

there are no further remaining uncertainties to be taken into account

Hazard assessment conclusion:

no hazard identified

Hazard assessment conclusion:

no hazard identified

EXPOSURE DNEL DERIVATIONS FOR METHYLiso-AMYL KETONE

(CAS No. 110-12-3)

I. Introduction

Methyliso-amyl ketone (5-methylhexan-2-one) is currently listed as a harmful substance in the Annex I of the EU Dangerous Substance Directive 67/548/EEC and is rated Xn; R20 (Harmful by inhalation). Similarly, it is listed in Annex VI to EC Regulation 1272/2008 as Acute Toxicity, Category 4 (H332: Harmful if inhaled). It is not a Category 1 or 2 mutagen or carcinogen. Thus, DNEL/DMEL values for these endpoint were not derived. However, recently a reproductive hazard has been identified and the substance is classified and labelled as Repro Cat. 2. DNELvalues for these endpoints were derived (OECD 443, rats, whole body inhalation; NOAEC (maternal toxicity, systemic) = 1500 ppm = 7125 mg/m³; NOAECL (neonatal toxicity, systemic) = 75ß ppm = 3562.5 mg/m³. Summarized in Table 1 are the critical preliminary DNEL values derived for methyl isoamyl ketone.

Table 1: Critical Preliminary DNEL Values for Methyl Isoamyl Ketone

Exposure Pattern	DNEL
Exposure Pattern	Workers	General Population
Acute – inhalation, systemic	196.3 mg/m³(1)	146.5 mg/m³
Acute – inhalation, local	not calculated	not calculated
Acute – dermal, systemic	not calculated	not calculated
Acute – dermal, local	not calculated (2)	not calculated (2)
Acute – oral, systemic	not relevant (3)	not relevant (3)
Long-term – inhalation, systemic	100.25 mg/m³	17.8125 mg/m³
Long-term – inhalation, local	not relevant (4)	not relevant (4)
Long-term – dermal, systemic	14.2 mg/kg bwt/day	5.12 mg/kg bwt/day
Long-term – dermal, local	No data	No data
Long-term – oral, systemic	not relevant (3)	5.12 mg/kg bwt/day

1 Chapter 8 of the REACH TGD (Appendix R.8-8) indicates that acute exposure DNEL values are not normally required. Additionally, the lack of acute oral or dermal toxicity would indicate MiAK is non-hazardous under acute exposure scenarios. Given the assignment of R20 and Acute Toxicity (Category 4, H332), acute inhalation DNEL values (15-minute exposures) were derived for MiAK.

2 Assessment based on a lack of significant skin irritation.

3 It is assumed that there will be no acute or long-term oral exposures to MiAK in worker populations or acute oral exposures in consumer populations.

4 Eye and respiratory irritation thresholds are assumed to exceed thresholds for chronic systemic effects.

Summarized in Table 2 are the critical toxicological studies and values identified for MiAK.

Table 2: Dose descriptor for key studies of toxicological concern

Endpoint	Descriptor	Key Reference	Associated Effects/Comments
Acute Toxicity
Inhalation, rat 802, 1603, 3207 and 5878 ppm (measured)	LOAEL 802 mg/m³ (3810 mg/m³)	Katz, G.V., Basic Toxicity of Methyl Isoamyl Ketone, Eastman Kodak Company, Report No. 104791R, 1978	Reversible CNS effects; initial weight loss with weight gains in surviving animals. A LOAEL has been assigned based on the presence of only minimal and reversible CNS effects immediately following dosing.
Repeated Dose Toxicity
Inhalation, rat 212, 1025 and 2079 ppm (measured)	NOAEL = 212 ppm (1007 mg/m³)	Katz, G.V., Renner, C.J. and Terhaar, C.J. (1986). Subchronic inhalation toxicity of methyl isoamyl ketone in rats. Fundam. Appl. Toxicol. 6:498-505.	Increased absolute/relative liver/kidney weights; hyalin droplet degen. of kidney (males); minor/moderate tubular epithelium regeneration; hepatocyte hypertrophy
IOEL
NOAEL from study of Katz et al. (1986); assessment factor of 10	IOEL (8-hr TWA) = 20 ppm (95 mg/m³)	European Commission Directive 2000/39/CE	Refer to study of Katz et al. (1986)

Reproductive toxicity
NOAEL from study of Corder, R. (2018);	NOAEL (neonatal toxicity, systemic, rats, whole body inhalation, OECD 443) = 750 ppm corresponding to ca. 3562.5 mg/m³	Coder, R., OECD 443 with Methyl isoamyl ketone	Refer to study of Corder, P. (2018)

II. Descriptions and Rationale for DNEL Derivations

1. Acute toxicity

The acute oral toxicity of methyl isoamyl ketone (MiAK) is low with LD₅₀values reported in experimental animals of 3200 mg/kg bwt. In all studies reported, clinical signs indicating reversible effects on the central nervous system included one or more of the following: weakness, ataxia, rough hair coats, excitement, vasodilatation, slight tremors, restlessness, and prostration. All surviving animals gained weight normally.

The acute dermal toxicity of MiAK is low. In two studies, doses of 5, 10 or 20 mL/kg bwt (equivalent to 4, 8 and 16 g/kg bwt) MiAK were applied to the depilated abdomens of guinea pigs under occluded contact for 24-hours. There were no deaths observed in these studies. Except for weight loss at the two higher dose levels, there were no clinical signs indicating absorption or systemic toxicity. The dermal LD₅₀is considered to be > 20 mL/kg bwt (16 g/kg bwt).

In a key study, groups of 4 male rats were exposed via whole body inhalation to target concentrations of 800, 1600, 3200 or 6400 ppm MiAK (measured concentrations: 802, 1603, 3207 and 5878 ppm) for 6 hours. All rats at the highest exposure concentration died within 2.5 hours of the start of exposure. One of four rats at the 3207 ppm concentration died just prior to termination of exposure. At the highest concentration tested, eye irritation and narcosis were observed in all animals prior to death. Reversible central nervous system effects were also observed in survivors at all exposure concentrations. Although surviving rats in the 3207 ppm group lost weight over the first 3 days, subsequent weight gain was normal and all other rats gained weight normally over the 14-day observation period. A LOAEL of 802 ppm (3810 mg/m³) was set for this study based on the minimal appearance of reversible CNS effects immediately after exposures. These data are supported by four lesser studies in which groups of 3 rats were exposed via whole body inhalation to 3.88 mg/L (948 ppm), 21 mg/L (4000 ppm), 29 mg/L (5500 ppm), or 31.57 mg/L (7718 ppm) for 6 or 7 hours. At 7718 ppm, all animals exhibited signs of central nervous system depression and died shortly after hour 4 of exposure. All animals survived exposure at 5500 ppm for 7 hours but exhibited central nervous system effects including loss of coordination, prostration, and labored breathing. One of three animals exposed to 4000 ppm died at 5 hours into the 6-hr exposure period. Reversible central nervous system effects were observed in survivors. No deaths or clinical signs were observed in the 948 ppm exposure group. MiAK is considered to be harmful by the inhalation route.

In the case of acute worker and general population inhalation exposures, the LOAEL (6-hr) value of 3810 mg/m³, measured in rats, was selected as the appropriate dose descriptor for derivation of acute systemic inhalation DNEL values. No additional local or systemic acute DNEL values were derived. It should be noted that there is no IOEL STEL value assigned for MiAK that would be appropriate for the derivation of acute inhalation systemic DNEL values for workers.

2. Irritation/Sensitization

In experimental animals, MiAK caused only slight and transient dermal irritation when tested in guinea pigs following 24-hr occluded contact. Repeated dermal applications of the test material did not significantly exacerbate the irritative effects. Application of MiAK to the eyes of rabbits (either washed or unwashed) produced at most only slight irritation. In a primary dermal irritation study, slight irritation and some signs of desquamation were observed following 24-hr occluded contact. Repeat application of the test material did not significantly exacerbate the irritative effects. There were no clinical signs indicative of respiratory tract irritation in rats following acute inhalation exposure to up to 7700 ppm or subacute or subchronic exposure to up to 2000 ppm. The material was also not a skin sensitizer when tested in guinea pigs. Based on a weight-of-evidence assessment, MiAK would not be classified as a respiratory tract irritant.

3. Repeat dose toxicity

MiAK administered to rats via whole body inhalation at concentrations up to 2000 ppm for a total of 69 exposures and spanning 96 days produced treatment-related organ weight changes and microscopic effects in the livers and kidneys of both male and female rats at 1000 and 2000 ppm (Katz et al., 1986). The NOEC for systemic toxicity was determined to be 200 ppm (212 ppm measured) based on a lack of treatment-related organ weight changes and microscopic effects observed in the livers and kidneys at the higher exposure concentrations. Increased kidney weights were not accompanied by changes in clinical chemistry parameters. Hyaline droplet degeneration, observed in the proximal convoluted tubular epithelium of the kidneys of male rats, was not considered relevant to human risk assessment. Regeneration of tubular epithelium in both sexes was minor and neither kidney lesion impaired the health of the animals. Liver weight changes were accompanied by minor to moderate hepatocyte hypertrophy, but no changes in serum clinical chemistries were observed which would be suggestive of liver damage. No hepatocyte hyperplasia was observed by the inhalation route of exposure.

MiAK when administered by gavage to rats at a dose level of 2000 mg/kg bwt/day, five days/week over 90 days, produced statistically significant increases in two liver enzyme values, GOT and GPT, and microscopic changes in the stomach, liver and kidney. Changes in the stomach were thought to be due to irritation following prolonged contact with the test material and, as such, would not be a true indication of a target organ effect. The observed kidney effects are consistent with those seen with exacerbation of a commonly occurring effect observed in male laboratory rats following chemical exposure and which have not been observed in humans. Increases in liver enzymes, increases in absolute and relative liver weights, and histopathologic changes in the liver were indicative of the liver as a target organ for exposure to methyl isoamyl ketone by the oral route and consisted of effects described as pre-neoplastic, hepatocyte necrosis and bile-duct hyperplasia. Although liver effects were observed at extremely high oral doses and only minor liver effects were observed at comparable inhalation exposures. A NOEL or NOAEL could not be assigned to this oral gavage study.

The IOEL value (8-hr TWA) of 95 mg/m³had been selected in the past as the long-term inhalation systemic DNEL value for workers. Since the publication of this IOEL, there has been new scientific information on the toxicology of MiAK (OECD 443 study results, see below) that would invalidate the use of this for the setting of a worker DNEL value. Formerly, the NOAEC value of 212 ppm (1007 mg/m³) (Katz et al., 1986) had been selected as the starting dose descriptor for the derivation of a long-term inhalation systemic DNEL value for the general population. After correction for route-of-exposure, this same NOAEC value had served as the starting point for both the oral and dermal long-term systemic DNEL values for the general population and for the dermal long-term systemic DNEL for workers. These DNELs however, are now derived of the NOAEC value obtained from the recent OECD 443 study in rats.

4. Genotoxicity/carcinogenicity

MiAK was negative in an in vitro Ames mutagenicity assay conducted at plate incorporation levels of up to 5,000 mg/plate. In a mouse lymphoma L5178Y mammalian cell gene mutation assay, MiAK was negative at concentrations up to 1200 µg/mL in the presence and absence of mammalian metabolic activation and showed no evidence of inducing chromosomal aberrations or polyploidy when tested in an in vitro cytogenetics test with Chinese hamster ovary cells (CHO cells) with and without metabolic activation up to a concentration of 1200 µg/mL. MiAK has not been tested for genotoxicity in vivo.

MiAK has not been tested for carcinogenicity.

5. Reproductive/developmental toxicity

In an OECD 421 reproductive/developmental toxicity screening study in rats, MiAK was administered by inhalation (whole body) at up to 5.0 mg/L for 6 hr/day, 7 days/week for a total of 51 (males) and 35-41 (females) consecutive days during premating, mating, pregnancy and early lactation. There were no gross or microscopic effects on the reproductive organs of either sex and there were no treatment-related effects on any reproductive parameters. Similarly, in 13-week exposure studies in rats by the oral and inhalation routes, there were no reported gross or microscopic changes in the reproductive organs of either sex at oral gavage doses up to 2000 mg/kg bwt/day or inhalation exposures up to 2000 ppm (6 hr/day, 5 days/week for a total of 69 exposures).

In and OECD 414 prenatal developmental toxicity test, pregnant female rats were exposed by whole-body inhalation to the test article at target concentrations(actual) of 380(379), 750(751) and 1500(1495) ppm. Animals were exposed from gestation day 0 to gestation day 19. There was a decreased in food consumption and body weight in the 1500 ppm dams, along with a reduced startle reflex at 1500 and 750 ppm. The NOAEL for maternal toxicity was 380 ppm and 750 ppm was the NOAEL for embryo/fetal toxicity based on reduced pup weights at 1500 ppm. There were no significant findings of any developmental defects in the pups at any dose.

In and OECD 443 extended one-generation reproductive toxicity test, rats were exposed by whole-body inhalation to the test article at target concentrations(actual) of 380, 750 and 1500 ppm. Overall mean analyzed exposure concentrations were 380, 750, and 1499 ppm for the F0 generation and 379, 750, and 1497 ppm for the F1 generation. Animals in the parental (F0) generation were exposed via whole-body inhalation for 6 hours daily for 70 consecutive days prior to mating and continuing through the day prior to euthanasia. Maternal exposure was suspended from Gestation Day 20 through Lactation Day 4 to prevent confounding effects on parturition and maternal nursing and nesting behavior. The offspring in the F1 generation were potentially exposed in utero during gestation. Direct offspring exposure during the pre-weaning period was demonstrated on a concurrent study, where quantifiable levels of MiAK were obtained in maternal milk and plasma obtained from dams exposed at the same target exposure concentrations (380 to 1500 ppm). Offspring selected to constitute the F1 generation were exposed beginning at weaning (Postnatal Day 28) and continuing through the day prior to euthanasia. Due to the lower body weight gains noted for the F1 pups during the neonatal and pre-weaning periods (see below), F1 animals assigned to Cohort 1B for follow-up reproductive assessments were bred to obtain an F2 generation. All procedures for F1 animals assigned to Cohort 1B were generally the same as those for the F0 generation/F1 litters, including suspension of exposure for the F1 generation maternal animals from Gestation Day 20 through Lactation Day 4. The offspring in the F2 generation were not directly exposed to test substance atmospheres but were potentially exposed in utero, and through maternal milk during the pre-weaning period.

The following parameters and end points were evaluated in this study: clinical signs, body weights, body weight gains, food consumption, estrous cycles, reproductive performance, parturition, litter viability and survival, anogenital distance, areolae/nipple anlagen, developmental landmarks, thyroid hormones, clinical pathology, gross necropsy findings, spermatogenic endpoints, organ weights, and histopathologic examinations.

The NOAEL for maternal toxicity was 1500 ppm and 750 ppm was the NOAEL for embryo/fetal toxicity based on reduced pup weights at 1500 ppm.

III. Mode of Action Considerations

For the critical studies identified in Table 2, a threshold mode of action was assumed.

IV. Modification of Relevant Dose Descriptors to the Correct Starting Point

Summarized in Table 3 are corrected dose descriptors derived from those in Table 2 (see Guidance Document, Chapter R.8, Appendix R.8-2).

Table 3: Corrected dose descriptors for key studies of toxicological concern

Endpoint	Most relevant dose descriptors		Corrected dose descriptors (Workers)
	Local	Systemic	Local	Systemic
Acute Toxicity
-oral	NA	NA	NA	NA
-dermal	NC	NC	NC	NC
-inhalation	NC	LOAEC (6-hr) = 3810 mg/m³	NC	7363 mg/m³
Repeated Dose Toxicity
-oral	NA	NA	NA	NA
-dermal	NC	NOAEC = 750 ppm = 3562.5 mg/m³	NC	1421.4 mg/kg bwt/day
-inhalation	NC	NOAEC = 750 ppm = 3562.5 mg/m³	NC	2506.2 mg/m³
Endpoint	Most relevant dose descriptors		Corrected dose descriptors (General Population)
	Local	Systemic	Local	Systemic
Acute Toxicity
-oral	NC	NC	NC	NC
-dermal	NC	NC	NC	NC
-inhalation	NC	LOAEC (6-hr) = 3810 mg/m³	NC	10,990 mg/m³
Repeated Dose Toxicity
-oral	NC	(use inhal. data)	NC	1024.2 mg/kg bwt/day
-dermal	NC	(use inhal. data)	NC	1024.2 mg/kg bwt/day
-inhalation	NC	NOAEC = 750 ppm = 3562.5 mg/m³	NC	890.625 mg/m³

NA - not applicable NC - not calculated

Workers

1. Acute

In the case of worker exposures, the oral route of exposure was not considered. Based on the low acute toxicity of MiAK by the dermal route, the dermal route of exposure was also not considered. An acute inhalation LOAEC (6-hr) value in the rat of 3810 mg/m³was used as the starting point for an acute inhalation systemic worker DNEL (see Guidance Document, Chapter R.8, Appendix R.8-8). Duration of exposure correction was applied based on an acute (15-minute) exposure and a correction for an increased respiratory volume for light work conditions (see Guidance Document, Chapter R.8.4.2). For workers, the 6-hour starting value (3810 mg/m³) was corrected to a 15-minute exposure duration using Haber’s Law (n=3) giving a modified dose descriptor as follows:

LOAEC_worker= 10,990 mg/m³(Haber’s Law corrected value)

= 10,990 mg/m³x (6.7 m³/8 hours / 10 m³/8 hours) (Light work corrected)

or 7363 mg/m³

2. Long-Term

dermal

The starting point for a long-term (repeated dose) dermal systemic DNEL value for workers was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (8 hours/d) x (7 d/week / 5 d/week)

or, NOAEC_corr= 3740.625 mg/m³

Conversion to a dermal equivalent dose is performed as discussed in the Guidance Document, Appendix R.8-2, Example B.4.

NOAEL_dermal,rat= NOAEC_corrx sRV_rat(0.38 m³/kg bwt for 8 hour exposure)

or, 1421.4 mg/kg bwt/day

inhalation

The starting point for a long-term (repeated dose) inhalation systemic DNEL value for workers was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure and for an increased respiratory volume for light work conditions (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (8 hours/d) x (6.7 m³/8 hours / 10 m³/8 -hours - light work corrected) * (7 d/week / 5 d/week)

or, NOAEC_corr= 2506.2 mg/m³

General Population

1. Acute

In the case of general population exposures, the oral and dermal routes of exposure were not considered based on the low acute toxicity of MiAK by either route. An acute inhalation LOAEC (6-hr) value in the rat of 3810 mg/m³was used as the starting point for an acute inhalation systemic general population DNEL (see Guidance Document, Chapter R.8, Appendix R.8-8). Duration of exposure correction was applied based on an acute (15-minute) exposure (see Guidance Document, Chapter R.8.4.2). For the general population, the 6-hour starting value (3810 mg/m³) was corrected to a 15-minute exposure duration using Haber’s Law (n=3) giving a modified dose descriptor as follows:

LOAEC_gen.pop.= 10,990 mg/m³(Haber’s Law corrected value)

2. Long-Term

oral and dermal

The starting point for a long-term (repeated dose) orla or dermal systemic DNEL value for the general population was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (24 hours/d)

or, NOAEC_corr= 890.625 mg/m³

Conversion to a dermal equivalent dose is performed as discussed in the Guidance Document, Appendix R.8-2, Example B.4.

NOAEL_dermal,rat= NOAEC_corrx sRV_rat(1.15 m³/kg bwt for 24 hour exposure)

or, 1024.2 mg/kg bwt/day

inhalation

The starting point for a long-term (repeated dose) inhalation systemic DNEL value for general population was the NOAEC value of 3562.5 mg/m³from an extended one-generation reproductive rat inhalation toxicity study. This value was corrected for duration of exposure (see Guidance Document, Chapter R.8.4.2). A first-pass effect was discounted for the purposes of these calculations.

NOAEC_corr= NOAEC_ratx (6 hours/d) / (24 hours/d)

or, 890.8125 mg/m³

V. Application of Assessment Factors to the Corrected Dose Descriptors

Summarized in Table 4 are endpoint-specific DNEL values for worker and general population exposures to MiAK derived from the corrected dose descriptors found in Table 3.

Table 4: Endpoint-specific DNEL values for MiAK

Endpoint	Corrected dose descriptor		Overall AF[1]	Endpoint-specific DNELs (Workers)
	Local	Systemic		Local	Systemic
Acute Toxicity
-oral	NA	NA	NA	NA	NA
-dermal	NC	NC	NC	NC	NC
-inhalation	NC	7363 mg/m³	15	NC	196.3 mg/m³
Repeated Dose Toxicity
-oral	NA	NA	NA	NA	NA
-dermal	NC	1421.4 mg/kg bwt/day	100	NC	14.2 mg/kg bwt/day
-inhalation	NC	2506.2mg/m³	25	NC	100.25 mg/m³
Endpoint	Corrected dose descriptor		Overall AF¹	Endpoint-specific DNELs (General Population)
Acute Toxicity
-oral	NC	NC	NC	NC	NC
-dermal	NC	NC	NC	NC	NC
-inhalation	NC	10,990 mg/m³	75	NC	146.5 mg/m³
Repeated Dose Toxicity
-oral	NC	1024.2 mg/kg bwt/day	200	NC	5.12 mg/kg bwt/day
-dermal	NC	1024.2 mg/kg bwt/day	200	NC	5.12 mg/kg bwt/day
-inhalation	NC	890.625 mg/m³	50	NC	17.8125 mg/m³

NA - not applicable NC - not calculated

Workers

1. Acute

In the case of acute inhalation worker exposure, the starting dose descriptor did not represent a clear NOAEL and thus an assessment factor (AF) of 3 was applied. An additional AF of 5 was applied (intraspecies, sensitive worker) to give an overall AF of 15 (Table 4).

2. Long-Term

dermal

An AF of 4 was applied to the corrected dose descriptor for dermal long-term systemic effects in workers (interspecies differences, allometric scaling). In addition, an AF of 2 (subchronic to chronic). an AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive worker) AF of 5 were applied. The overall AF obtained was 100 (Table 4).

inhalation

An AF of 2 was applied to the corrected dose descriptor for inhalation long-term systemic effects in workers (subchronic to chronic), an AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive worker) AF of 5 were applied. The overall AF obtained was 25 (Table 4).

General Population

1. Acute

In the case of acute inhalation exposure of the general population, the starting dose descriptor did not represent a clear NOAEL and thus an assessment factor of 3 was applied. An additional assessment factor (AF) of 2.5 for remaining interspecies differences and an AF of 10 was applied (intraspecies, sensitive individual) to give an overall AF of 75 (Table 4).

2. Long-Term

oral and dermal

An AF of 4 was applied to the corrected dose descriptor for oral long-term (repeated dose) systemic effects in the general population (interspecies differences, allometric scaling). In addition, an AF of 2 (subchronic to chronic), an AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive individual) AF of 10 were applied. The overall AF obtained was 200 (Table 4). These same AF values apply to the case of dermal exposure.

inhalation

No additional intraspecies (allometric scaling) factor was applied to the corrected dose descriptor for inhalation long-term systemic effects in the general population. An AF of 2 (subchronic to chronic), An AF of 2.5 for remaining interspecies differences and an intraspecies (sensitive individual) AF of 10 were applied. The overall AF obtained was 50 (Table 4).

SUMMARY

The calculated DNEL values for MiAK are summarized in Table 1 at the beginning of this document.

[1]Overall assessment factors were assigned based on the ECHA guidelines.

Registration Dossier

Registration Dossier

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Diss Factsheets

5-methylhexan-2-one

Toxicological Summary

Administrative data

Workers - Hazard via inhalation route

Systemic effects

Long term exposure

DNEL related information

Acute/short term exposure

DNEL related information

Local effects

Long term exposure

Acute/short term exposure

DNEL related information

Workers - Hazard via dermal route

Systemic effects

Long term exposure

DNEL related information

Acute/short term exposure

DNEL related information

Local effects

Long term exposure

Acute/short term exposure

Workers - Hazard for the eyes

Local effects

Additional information - workers

I. Introduction

II. Descriptions and Rationale for DNEL Derivations

III. Mode of Action Considerations

IV. Modification of Relevant Dose Descriptors to the Correct Starting Point

Workers

Workers

General Population - Hazard via inhalation route

Systemic effects

Long term exposure

DNEL related information

Acute/short term exposure

DNEL related information

Local effects

Long term exposure

Acute/short term exposure

DNEL related information

General Population - Hazard via dermal route

Systemic effects

Long term exposure

DNEL related information

Acute/short term exposure

DNEL related information

Local effects

Long term exposure

Acute/short term exposure

General Population - Hazard via oral route

Systemic effects

Long term exposure

DNEL related information

Acute/short term exposure

DNEL related information

General Population - Hazard for the eyes

Local effects

Additional information - General Population

I. Introduction

II. Descriptions and Rationale for DNEL Derivations

III. Mode of Action Considerations

IV. Modification of Relevant Dose Descriptors to the Correct Starting Point

Workers

Workers