(S)-p-mentha-1,8-diene

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because a pre-natal developmental toxicity study is available

other:

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Study period:

1977

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: Read-across from an analogue substance for which there is available information (Klimish=4)

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
See cross-reference to justification of read-across.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Treatment with the highest dose level (1000 mg/kg) of d-limonene resulted in death of dams with less than 40% mortality. The significant decrease of body- weight gain and food consumption were temporarily observed in dams given 500 and 1000 mg/kg of d-limonene, but no anomalies were observed in the general behaviour of dams given 250 and 500 mg/kg of d-limonene during the gestation.

Key result

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

body weight and weight gain

mortality

Remarks on result:

other: Based on the read-across from an analogue substance.

Key result

Abnormalities:

no effects observed

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
- External examination of fetuses showed no anormalies.
- Visceral and skeletal examinations revealed some anormalies such as incomplete lobulation of the lungs, enlargement of the foramen ovale and retarded ossification of the middle phalanx of fore limbs in addition to the 5th sternebrae. These did not appear to be dose-dependent and restored to normal during the postnatal development.
- Other non specific anormalies involved the lumber ribs in fetuses and offsprings, formation of the accessory ossification center of the 5th sternebrae in offsprings and the atrial septal defect detected in only 2 fetuses of a litter from dams treated with 250 mg/kg bw/day of d-limonene.

Dose descriptor:

NOAEL

Effect level:

> 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: fetotoxicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Table 1: Effect of d-limonene on prenatal development of rabbit fetuses

Dose (mg/kg bw)	Control	250	500	1000
No. of pregnant animals	10	10	10	18
No. of dead clams	0	0	0	6
(%)				33
No. of examined clams	10	10	10	10
No. of implantations	96	94	85	91
(mean ± S.E.)	9.50 ± 0.25	9.40 ± 0.21	8.50 ± 0.33	9.10 ± 0.25
No. of resorbed fetuses	5	4	4	8
No. of dead fetuses	3	5	0	3
No. of live fetuses	88	85	81	80
Sex ratio (Male/Female)	0.73 (37/51)	1.13 (45/50)	0.62 (31/50)	1.11 (38/42)
Fetus body weight (g)
Male (mean ± S.E.)	44.39 ± 1.33	48.09 ± 1.07 *	44.76 ± 1.51	43.22 ± 0.96
Female (mean ± S.E.)	45.64 ± 1.00	47.45 ± 1.08	46.14 ± 1.21	45.13 ± 1.10
Placental weight (g)
Male (mean ± S.E.)	5.76 ± 0.17	5.84 ± 0.17	5.95 ± 0.29	5.77 ± 0.19
Female (mean ± S.E.)	5.87 ± 0.19	5.70 ± 0.15	6.16 ± 0.18	5.87 ± 0.23

* Significantly different from the control at 5% level

Table 2: Prenatal examinations of rabbit fetuses

Dose (mg/kg bw)	Control	250	500	1000
External examination
No. of examined fetuses	91	90	81	83
No. of malforrned fetuses	0	0	0	0
Visceral examination
No. of examined fetuses	88	85	81	80
No. of malformed fetuses
Atrial septal defect (%)	0	2 (2.4)	0	0
No. of minor abnormality
Incomplete lobulation of lungs (%)	11 (12.5)	16 (18.8)	19 (23.5)	19 (23.8)
Enlargement of foramen ovale (%)	2 (2.3)	2 (2.4)	5 (6.2)	4 (5.0)
Skeletal examination
No. of examined fetuses	86	87	81	80
No. of malformed fetuses	0	0	0	0
No. of variation
Left lumbar rib (%)	18 (20.9)	26 (29.9)	14 (17.3)	25 (31.3)
Right lumbar rib (%)	16 (18.6)	22 (25.3)	14 (17.3)	22( 27.5)
Ossification pattern
Retarded ossification of 5th sternebrae (%)	11 (12.8)	14 (16.1)	8 (9.9)	18 (22.5)
Retarded ossification of middle phalanx of fore limbs (%)	2 (2.3)	3 (3.4)	0	6 (7.4)

Table 3: Absolute organ weights of rabbit offsprings

	Male				Female
	Control	250	500	1000	Control	250	500	1000
No. of offsprings	13	12	8	13	10	13	16	9
Final body weight (g)	893.0 ± 45.3	1021.2 ± 45.4 **	931.9 ± 55.5	957.3 ± 52.4	1005.5 ± 53.7	1093.1 ± 46.6	860.0 ± 31.6 *	1071.7 ± 58.1
Liver (g)	36.49 ± 2.52	45.08 ± 1.52 *	38.67 ± 3.10	34.91 ± 2.76	41.79 ± 3.39	13.95 ± 3.18	34.68 ± 1.90	48.30 ± 6.14
Lungs (g)	5.53 ± 0.35	6.25 ± 0.26	5.98 ± 0.36	5.46 ± 0.18	5.91 ± 0.25	5.94 ± 0.28	5.72 ± 0.18	5.93 ± 0.45
Heart (g)	2.63 ± 0.17	3.50 ± 0.16 **	2.86 ± 0.17	3.04 ± 0.19	3.19 ± 0.17	3.38 ± 0.21	2.72 ± 0.12 *	3.34 ± 0.20
Spleen (g)	0.71 ± 0.05	0.77 ± 0.04	0.71 ± 0.08	0.81 ± 0.04	0.64 ± 0.06	0.74 ± 0.05	0.74 ± 0.04	0.78 ± 0.07
Thymus (g)	2.31 ± 0.20	2.51 ± 0.23	2.11 ± 0.38	1.96 ± 0.11	2.40 ± 0.30	2.77 ± 0.19	1.671.14 *	2.36 ± 0.31
Kidneys (g)	7.67 ± 0.42	9.80 ± 0.46 **	8.29 ± 0.29	8.58 ± 0.52	8.82 ± 0.46	8.40 ± 0.30	7.86 ± 0.37	9.56 ± 0.55
Thyroids (mg)	75.89 ± 8.35	102.68 ± 4.18*	86.74 ± 10.97	80.93 ± 7.41	82.78 ± 8.00	89.90 ± 4.11	75.75 ± 5.43	96.30 ± 9.67
Adrenals (mg)	69.65 ± 6.02	9.1.93 ± 1.06 **	78.79 ± 5.89	71.36 ± 6.00	82.23 ± 4.37	94.24 ± 5.07	87.64 ± 4.18	105.39 ± 15.11
Testes or Ovaries (mg)	180.42 ± 17.15	272.86 ± 16.46 **	185.62 ± 23.78	162.84 ± 20.59	46.31 ± 7.90	46.10 ± 2.80	43.53 ± 2.69	47.77 ± 3.59

* Significantly different from the control at 5% level

** Significantly different from the control at 1% level

Table 4: Relative organ weights per 100 g of rabbit offsprings

	Male				Female
	Control	250	500	1000	Control	250	500	1000
No. of offsprings	13	12	8	13	10	13	16	9
Final body weight (g)	893.0 ± 45.3	1021.2 ± 45.4 **	931.9 ± 55.5	957.3 ± 52.4	1005.5 ± 53.7	1093.1 ± 46.6	860.0 ± 31.6 *	1071.7 ± 58.1
Liver (g/100 g)	4.08 ± 0.25	3.99 ± 0.22 *	4.16 ± 0.26	3.52 ± 0.10	4.25 ± 0.17	4.03 ± 0.21	4.02 ± 0.16	4.04 ± 0.32
Lungs (g/100 g)	0.61 ± 0.03	0.55 ± 0.03	0.65 ± 0.04	0.58 ± 0.03	0.62 ± 0.02	0.55 ± 0.02 *	0.67 ± 0.03	0.51 ± 0.02 **
Heart (g/100 g)	0.29 ± 0.01	0.31 ± 0.01	0.31 ± 0.01	0.31 ± 0.02	0.33 ± 0.01	0.31 ± 0.01	0.32 ± 0.01	0.29 ± 0.01 *
Spleen (g/100 g)	0.08 ± 0.01	0.07 ± 0.01	0.08 ± 0.01	0.08 ± 0	0.07 ± 0.01	0.07 ± 0.01	0.09 ± 0	0.07 ± 0.01
Thymus (g/100 g)	0.25 ± 0.02	0.22 ± 0.02	0.25 ± 0.03	0.20 ± 0.01 *	0.24 ± 0.03	0.25 ± 0.01	0.195 ± 0.01	0.20 ± 0.02
Kidneys (g/100 g)	0.85 ± 0.04	0.86 ± 0.04	0.90 ± 0.03	0.88 ± 0.02	0.91 ± 0.02	0.80 ± 0.01	0.91 ± 0.01	0.83 ± 0.03 *
Thyroids (mg/100 g)	8.21 ± 0.69	9.10 ± 0.51	9.21 ± 0.87	8.18 ± 0.42	8.40 ± 0.56	8.41 ± 0.45	8.81 ± 0.57	8.13 ± 0.41
Adrenals (mg/100 g)	7.82 ± 0.60	8.37 ± 0.40	8.58 ± 0.77	7.18 ± 0.28	8.61 ± 0.51	8.79 ± 0.57	9.15 ± 0.42	9.04 ± 0.14
Testes or Ovaries (mg/100 g)	23.68 ± 1.31	19.68 ± 0.94 *	19.48 ± 1.69	16.35 ± 1.56	5.15 ± 1.19	4.33 ± 0.32	5.19 ± 0.43	3.84 ± 0.32

* Significantly different from the control at 5% level

** Significantly different from the control at 1% level

Table 5: Effects of d-limonene on gross differentiations of rabbit offsprings

	Control	250	500	1000
No. of examined offsprings	23	25	24	22
Days of gross differentiation after birth Opening of the ear-shell
6th day (%)	0	0	1 (4.2)	0
7th day (%)	23 (100)	25 (100)	23 (95.8)	22 (100)
Coating with the hair
2nd day (%)	7 (30.4)	0	0	0
3rd day (%)	16 (69.6)	25 (100)	24 (100)	22 (100)
Odontiasis
At birth (%)	23 (100)	25 (100)	24 (100)	22 (100)
Opening of the eyelids
9th day (%)	0	0	0	3 (13.6)
10th day (%)	11 (47.8)	4 (16.0)	13 (54.2)	5 (22.7)
11th day (%)	4 (17.4)	15 (60.0)	10 (41.7)	12 (54.5)
12th day (%)	3 (13.0)	5 (20.0)	1 (4.2)	2 (9.1)
13th day (%)	5 (21.7)	1 (4.0)	0	0

Table 6: Effects of d-limonene on postnatal development of rabbit offsprings

	Control	250	500	1000
No of dams	3	3	3	3
No. of still-birth (Male/Female)	1 (1/0)	0	0	1 (0/1)
No. of offsprings (Male/Female) At birth	28 (15/13)	27 (14/13)	26 (8/18)	27 (15/12)
1st week	28 (15/13)	27 (14/13)	25 (8/17)	26 (14/12)
2nd week	26 (14/12)	27 (14/13)	25 (8/17)	26 (14/12)
3rd week	24 (14/10)	27 (14/13)	25 (8/17)	25 (14/11)
4th week	23 (13/10)	26 (13/13)	25 (8/17)	22 (13/ 9)
5th week	23 (13/10)	25 (12/13)	25 (8/17)	22 (13/ 9)
6th week	23 (13/10)	25 (12/13)	25 (8/17)	22 (13/ 9)
7th week	23 (13/10)	25 (12/13)	24 (8/16)	22 (13/ 9)
Weanling rate (%)	79.3 (81.2/76.9)	92.6 (85.7/100)	92.3 (100/88.9)	78.6 (86.7/69.2)

Table 7: Postnatal examinations of rabbit offsprings

	Control	250	500	1000
No. of dams	3	3	3	3
No. of examined offsprings	23	25	24	22
Sensory function	Normal	Normal	Normal	Normal
External examination No. of malformed offsprings	0	0	0	0
Visceral examination No. of malformed offsprings	0	0	0	0
No. of minor abnormality Incomplete lobulation of lungs (%)	2 (8.7)	1 (4.0)	0	0
Accessory spleen (%)	2 (8.7)	0	0	0
Protrusion of gall bladder (%)	1 (4.3)	1 (4.0)	0	0
Skeletal examination No. of malformed offsprings	0	0	0	0
No. of variation Left lumbar rib (%)	4 (17.4)	4 (16.0)	4 (16.7)	4 (18.2)
Right lumbar rib (%)	2 (8.7)	6 (24.0)	6 (25.0)	4 (18.2)
Translocation of caudal vertebrae (%)	1 (4.3)	0	1 (4.2)	0
Ossification pattern Retarded ossification of 5th sternebrae (%)	0	2 (8.0)	0	1 (4.5)
Accessory ossification center of 5th sternebrae (%)	1 (4.3)	2 (8.0)	0	3 (13.6)

Conclusions:

Based on the read-across from the analogue substance d-limonene, l-limonene is not considered to have teratogenic potential in rabbit, the NOAEL for fetal toxicity was estimated to be greater than 1000 mg/kg bw/day. The NOAEL for maternal toxicity was estimated to be 250 mg/kg bw/day based on the decreased bodyweight gain.

Executive summary:

Based on the read-across from the analogue substance d-limonene, l-limonene is not considered to have teratogenic potential in rabbit, the NOAEL for fetal toxicity is estimated  to be greater than 1000 mg/kg bw/day. The NOAEL for maternal toxicity is estimated to be 250 mg/kg bw/day based on the decreased bodyweight gain.

Cross-reference 2

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Study period:

1977

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: Read-across from an analogue substance for which there is available information (Klimish=4)

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
See cross-reference to justification of read-across.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Key result

Dose descriptor:

NOAEL

Effect level:

591 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

body weight and weight gain

Remarks on result:

other: Based on the read-across from an analogue substance.

Key result

Abnormalities:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

591 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Increased incidences of abnormal skeletal formation in fetuses and decreased bodyweight gain in male offsprings born to dams administered with 2363 mg/kg-bw/day

Remarks on result:

other: Based on the read-across from the analogue substance d-limonene.

Key result

Abnormalities:

effects observed, treatment-related

Localisation:

skeletal: rib

Description (incidence and severity):

Lumber rib and fused rib in the fetuses

Key result

Developmental effects observed:

yes

Lowest effective dose / conc.:

2 363 mg/kg bw/day

Treatment related:

yes

Relation to maternal toxicity:

not specified

Dose response relationship:

no

Relevant for humans:

not specified

Conclusions:

Based on the read-across from the analogue substance d-limonene, the NOAEL for maternal and fetal toxicity of l-limonene was estimated to be 591 mg/kg bw/day based on the decreased bodyweight gain in dams and increased incidences of abnormal bone formation in fetuses.

Executive summary:

Based on the read-across from the analogue substance d-limonene, the NOAEL for maternal and fetal toxicity of l-limonene was estimated to be 591 mg/kg bw/day based on the decreased bodyweight gain in dams and increased incidences of abnormal bone formation in fetuses.

Cross-reference 3

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: Read-across from an analogue substance for which there is available information (Klimish=4)

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
See cross-reference to justification of read-across.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Key result

Dose descriptor:

NOAEL

Effect level:

591 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

body weight and weight gain

mortality

Remarks on result:

other: Based on the read-across from an analogue substance.

Key result

Abnormalities:

no effects observed

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Prolongation of the ossification of metacarpals and proximal phalanges in fetuses, decreased bodyweight gain (male offsprings) and organ weights at 2869 mg/kg bw/day

Key result

Dose descriptor:

NOAEL

Effect level:

591 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: prolongation of the ossification of metacarpals and proximal phalanges in fetuses, decreased bodyweight gain (male offsprings) and organ weights at 2869 mg/kg bw/day

Remarks on result:

other: Based on the read.across from an analogue substance.

Key result

Abnormalities:

effects observed, treatment-related

Localisation:

other: prolongation of the ossification of metacarpals and proximal phalanges in fetuses, decreased bodyweight gain (male offsprings) and organ weights at 2869 mg/kg bw/day

Key result

Developmental effects observed:

yes

Lowest effective dose / conc.:

2 869 mg/kg bw/day

Treatment related:

yes

Relation to maternal toxicity:

not specified

Dose response relationship:

no

Relevant for humans:

not specified

Conclusions:

Based on the read-across from the analogue substance d-limonene, the NOAEL for maternal toxicity of l-limonene was estimated to be 591 mg/kg bw/day based on the deaths and decreased bodyweight gain and the NOAEL for fetal toxicity was estimated to be 591 mg/kg bw/day based on the delayed skeletal formation and decreased bodyweight gain.

Executive summary:

Based on the read-across from the analogue substance d-limonene, the NOAEL for maternal toxicity of l-limonene was estimated to be 591 mg/kg bw/day based on the deaths and decreased bodyweight gain and the NOAEL for fetal toxicity was estimated to be 591 mg/kg bw/day based on the delayed skeletal formation and decreased bodyweight gain.

Cross-reference 4

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Read-across from an analogue substance for which there is available data (Klimish =2).

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
See cross-reference to justification of read-across.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not specified

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

not examined

Other effects:

not specified

Key result

Dose descriptor:

NOAEL

Based on:

test mat.

Sex:

male

Basis for effect level:

other: male-rat specific nephrotoxicity at all dose levels (considered as not relevant for humans)

Remarks on result:

not determinable

Remarks:

no NOAEL identified

Key result

Dose descriptor:

NOAEL

Effect level:

600 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: decrease of bodyweight gains at 1200 and 2400 mg/kg bw/day;

Key result

Dose descriptor:

LOAEL

Effect level:

1 200 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

body weight and weight gain

Key result

Dose descriptor:

NOAEL

Effect level:

600 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: mortality at 2400 mg/kg bw/day; occurrence of clinical signs of toxicity (rough hair coats, lethargy and excessive lacrimation) at 1200 and 2400 mg/kg bw/day

Key result

Dose descriptor:

LOAEL

Effect level:

1 200 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

clinical signs

Key result

Critical effects observed:

not specified

Conclusions:

Based on the read-across from the analogue substance d-limonene, the NOAEL of l-limonene for male and female rats was estimated to be 600 mg/kg bw/day. The LOAEL for female and male rats were considered to be 1200 and 150 mg/kg bw/day, based on observation of clinical signs and nephropathy, respectively. As nephrotoxicity and accumulation of hyaline droplets were observed in male rats at all dose levels, no NOAEL for male rats could be identified in this study.

Executive summary:

A 13-week subchronic toxicity study was performed similarly to OECD Guideline 408 and in compliance with GLP with the analogue substance d-limonene which was administered through gavage to groups of 10 F344/N rats/sex/dose mixed in corn oil at dose levels of 0, 150, 300, 600, 1200 and 2400 mg/kg bw/day for 13 weeks (5 days/week).  

Five of 10 males and 9/10 female rats that received 2400 mg/kg bw/day died during week 1. Final mean body weights of male rats in 600, 1200 or 2400 mg/kg bw/day groups were 6, 12 or 23% lower than that of the vehicle controls. Final body weight of the female rat that received 2400 mg/kg bw/day and lived to end of the study was 11% lower than the mean of the vehicle controls. Rough hair coats, lethargy and excessive lacrimation were observed at 1200 or 2400 mg/kg bw/day. No treatment-related histopathologic lesions were observed in female rats. Nephropathy was identified in all groups of male rats, and there was a dose-related increased severity of the lesion in dosed groups. Nephropathy was characterized by degeneration of epithelium in the convoluted tubules, granular casts within tubular lumens, primarily in the outer stripe of the outer medulla, and regeneration of the tubular epithelium. Hyaline droplets (protein reabsorption droplets) were observed in the epithelium of proximal convoluted tubules in all groups of male rats, including vehicle controls. This mechanism of nephrocarcinogenicity has been proven as being male-rat specific and not relevant for humans.

Based on the read-across from the analogue substance d-limonene, the NOAEL of l-limonene for female rats was considered to be 600 mg/kg bw/day. When considering the non relevance of the nephrotoxic effects for humans, the NOAEL for male rats would be 600 mg/kg bw/day, based on decrease of bodyweight gains at 1200 and 2400 mg/kg bw/day. The LOAEL for female and male rats were considered to be 1200 and 150 mg/kg bw/day, based on observation of clinical signs and nephropathy, respectively. As nephrotoxicity and accumulation of hyaline droplets were observed in male rats at all dose levels, no NOAEL for male rats was identified.

Cross-reference 5

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Read-across from an analogue substance for which there is available data (Klimish =2).

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
See cross-reference to justification of read-across.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not specified

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

not examined

Other effects:

not specified

Key result

Dose descriptor:

NOAEL

Effect level:

500 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: mortality at 2000 mg/kg bw/day; decreased bodyweight gain and occurence of clinical signs of toxicity (rough hair coats and decreased activity) at 1000 and 2000 mg/kg bw/day

Key result

Dose descriptor:

LOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

Key result

Critical effects observed:

not specified

Conclusions:

Based on the read-across from the analogue substance d-limonene, the NOAEL of l-limonene was estimated to be 500 mg/kg bw/day. The LOAEL was estimated to be 1000 mg/kg bw/day for both female and male mice, based on observation of clinical signs in both sexes and decreased bodyweights in males.

Executive summary:

A 13-week subchronic toxicity study was performed similarly to OECD Guideline 408 and in compliance with GLP with the analogue substance d-limonene which was administered through gavage to groups of 10 B6C3F1 mice/sex/dose mixed in corn oil at dose levels of 0, 125, 250, 500, 1000 or 2000 mg/kg bw/day for 13 weeks (5 days/week). Animals were observed twice daily for clinical signs of toxicity and bodyweights were recorded weekly. Necropsy performed on all animals and microscopic examination of specified tissues was performed for all control and high dose animals scheduled to be killed at the end of the treatment period.

One of 10 males and 2/10 females that received 2000 mg/kg bw/day and 1/10 females that received 500 mg/kg bw/day died before the end of the studies. Several animals in other groups died as a result of gavage error. Clinical signs of rough hair coats and decreased activity were observed at the two highest doses. Final mean bodyweights of mice that received 1000 or 2000 mg/kg bw/day were 10% lower than that of the vehicle controls for males and 2% lower for females. An alveolar cell adenoma was observed in the lung of 1/10 females that received 2000 mg/kg bw/day.

 

Based on the read-across from the analogue substance d-limonene, the NOAEL of l-limonene was estimated to be 500 mg/kg bw/day. The LOAEL was estimated to be 1000 mg/kg bw/day for both female and male mice, based on observation of clinical signs in both sexes and decreased bodyweights in males.

Results and discussion

Overall reproductive toxicity

Reproductive effects observed:

not specified