Fenitrothion

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

30 June 1992 - 2 October 1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EPA OPP 83-1 (Chronic Toxicity)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EPA OPP 82-7

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Fenitrothion

EC Number:

204-524-2

EC Name:

Fenitrothion

Cas Number:

122-14-5

Molecular formula:

C9H12NO5PS

IUPAC Name:

O,O-dimethyl O-3-methyl-4-nitrophenyl phosphorothioate

Test material form:

liquid

Specific details on test material used for the study:

Fenitrothion
Batch No.: 90617
Purity: 94.3 %

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on species / strain selection:

Standar strain and species

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Canada
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 50-53 days
- Weight at study initiation: 237-289 g (males), 168-215 g (females )
- Fasting period before study: None
- Housing: Individual
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 15 days

DETAILS OF FOOD AND WATER QUALITY:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-24
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: 30 June 1992 - 2 October 1992

Administration / exposure

Route of administration:

oral: feed

Details on route of administration:

Dietary incorporation

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

Dietary concentrations were 0, 6, 20, 60 and 200 ppm; corresponding to 0, 0.40, 1.32, 3.99 and 13.8 mg/kg bw/d in males; 0, 0.46, 1.56, 4.85 and 17.6 mg/kg bw/d in females.

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

Daily (continuous)

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 ppm

Remarks:

Control (basal diet)

Doses / concentrations 2

Dose / conc.:

6 ppm

Remarks:

0.40 and 0.46 mg/kg bw/d in males and females, respectively

Doses / concentrations 3

Dose / conc.:

20 ppm

Remarks:

1.32 and 1.56 mg/kg bw/d in males and females, respectively

Doses / concentrations 4

Dose / conc.:

60 ppm

Remarks:

3.99 and 4.85 mg/kg bw/d in males and females, respectively

Doses / concentrations 5

Dose / conc.:

200 ppm

Remarks:

13.8 and 17.6 mg/kg bw/d in males and females, respectively

No. of animals per sex per dose:

12/sex (main groups)
15/sex (cholinesterase activity assessment)

Control animals:

yes, concurrent no treatment

Details on study design:

Groups of 12 male and 12 female Sprague-Dawley rats for the main study and groups of 15 male and 15 female Sprague-Dawley rats for cholinesterase assessment received fenitrothion, in powdered Certified PMI Rodent Chow at concentrations of 0, 6, 20, 60 and 200 ppm (corresponding to 0, 0.40, 1.32, 3.99 and 13.8 mg/kg bw/d in males; 0, 0.46, 1.56, 4.85 and 17.6 mg/kg bw/day in females) for 13 weeks.

Positive control:

Not required

Examinations

Observations and examinations performed and frequency:

All animals were observed twice a day for their toxic signs and mortality. Body weight was recorded weekly and days of behavioural testing. Food consumption was recorded weekly for all animals.

Main study animals: Functional observational battery (FOB) and motor activity were tested at prestudy and 4th, 8th and 13th weeks of treatment.

Cholinesterase test animals: Erythrocyte, plasma and brain cholinesterase activities were determined on 5 rats/sex/group at 4, 8 and 13 weeks.

Sacrifice and pathology:

At the completion of the study, 6 rats/sex/group were randomly selected for perfusion and those in the control and high dose groups underwent a neuropathological evaluation. The remaining animals in each group (6/sex) had their brains removed and then were subjected to necropsy. The brain of each of non-perfused rats in the control and high dose groups was subsequently analyzed for glial fibrillary acidic protein (GFAP).

Statistics:

Study parameters were assessd and compared to controls using appropriate statistical analyses.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Four females in the 200 ppm group showed tremors on one or two days during the second week of treatment. The 200 ppm group females also showed a higher incidence of muzzle staining and brown staining along the tail during the study compared to the control group.

Mortality:

no mortality observed

Description (incidence):

No animals died and none were sacrificed prior to study completion

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Both males and females in the 200 ppm group and females in the 60 ppm group had significantly reduced body weights compared to the control group on Day 7. The 200 ppm group's body weight remained significantly lower on Day 14 and 21 for males and from Days 7 to 42 for females

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Food consumption by the 200 ppm group males and females were significantly lower than the control group from Days 0 to 7. Food consumption by the 200 ppm group females remained significantly reduced from Days 7 to 14 but then increased significantly from Days 28 to 56 and from 84 to 91.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Reductions in blood and brain cholinesterase levels were noted for males in the 60 and 200 ppm groups and females in the 20, 60 and 200 ppm groups.

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, non-treatment-related

Description (incidence and severity):

No significant differences considered to be related to treatment were seen for the qualitative components of the FOB. Significantly lower values for the 200 ppm group females in forelimb grip strength during the 4th week and in hindlimb grip strength during the 8th week occurred. These differences were considered not to be indicative of neurotoxicity taking lower body weights for the 200 ppm group females. Body temperature values as well as motor activity levels were not significantly different between control and fenitrothion-treated groups.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Neuropathological findings:

no effects observed

Description (incidence and severity):

No gross pathological findings or neuropathological changes (including brain measurements) seen were attributed to treatment with fenitrothion. No significant increases in glial fibrillary acidic protein were detected between the control and high dose groups for either male or females.

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Effect levels

Key result

Dose descriptor:

NOAEL

Effect level:

20 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical biochemistry

Target system / organ toxicity

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

60 ppm

System:

nervous system

Organ:

neurons

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Any other information on results incl. tables

Summary of bodyweights (g)

Dose level	0 ppm	6 ppm	20 ppm	60 ppm	200 ppm
Males
Day  0	264.2	262.7	265.3	266.0	263.6
Day  7	318.4	314.7	320.6	319.6	298.4**
Day 14	359.4	355.1	364.3	362.5	336.8**
Day 21	388.6	384.7	395.2	394.9	372.3*
Day 26	413.1	410.9	421.6	417.7	393.1
Day 27	409.3	406.0	419.0	415.4	392.5
Day 28	414.0	409.8	423.8	421.1	401.7
Day 35	439.1	428.7	453.3	447.8	429.5
Day 42	465.2	454.5	475.0	473.5	456.8
Day 49	488.2	472.8	496.0	494.2	479.1
Day 54	503.2	495.3	511.1	509.7	484.4
Day 55	496.6	492.9	508.4	505.3	482.3
Day 56	502.2	488.3	512.1	506.4	497.4
Day 63	518.8	503.1	254.2	524.5	509.1
Day 70	536.7	520.4	539.1	541.2	525.5
Day 77	553.6	538.6	553.6	558.7	539.7
Day 84	563.4	549.3	564.8	570.6	552.3
Day 89	569.5	570.8	581.6	584.5	554.7
Day 90	563.2	564.4	573.0	579.3	551.2
Day 91	566.8	556.0	569.7	577.7	558.8
Females
Day  0	189.2	188.9	186.8	185.6	186.5
Day  7	213.8	211.3	209.0	204.9*	186.0**
Day 14	226.8	226.4	225.4	219.9	190.2**
Day 21	244.1	242.7	238.1	234.7	210.6**
Day 26	249.7	251.4	246.6	243.4	216.2**
Day 27	251.3	250.7	247.8	242.5	215.4**
Day 28	255.9	253.2	248.1	246.5	226.4**
Day 35	269.3	264.8	261.0	257.4	246.4**
Day 42	280.8	276.5	270.5	268.6	216.8*
Day 49	290.8	280.4	277.0	279.6	277.1
Day 54	288.6	283.3	279.3	284.3	271.2
Day 55	287.1	280.4	277.9	282.7	268.5
Day 56	295.8	286.1	279.1	284.9	282.2
Day 63	300.7	292.5	288.6	296.5	284.7
Day 70	307.0	300.6	291.4	304.2	291.6
Day 77	314.0	308.0	301.5	313.0	299.8
Day 84	319.0	313.6	306.3	315.8	305.3
Day 89	319.1	313.8	306.2	317.3	305.5
Day 90	316.5	311.0	302.9	312.3	302.8
Day 91	318.4	312.2	304.7	315.6	307.2

*significantly different to controls (p<0.05); **p<0.01

Summary of food consumption (g/day)

Dose level	0 ppm	6 ppm	20 ppm	60 ppm	200 ppm
Males
Day  0 - 7	196.5	194.7	201.6	200.9	182.2**
Day  7 - 14	203.2	197.2	208.9	207.7	194.8
Day  14 - 21	206.3	202.9	212.0	211.5	209.5
Day  21 - 28	202.2	199.0	204.5	204.8	211.2
Day  28 - 35	204.2	199.0	210.9	204.8	211.8
Day  35 - 42	213.2	199.4	204.1	209.8	214.8
Day  42 - 49	217.0	197.9**	208.5	211.9	213.3
Day  49 - 56	208.8	197.0	205.3	206.0	213.3
Day  56 - 63	211.1	200.5	207.7	208.1	213.6
Day  63 - 70	215.3	203.1	205.5	207.0	210.1
Day  70 - 77	215.6	205.4	206.2	211.9	210.6
Day  77 - 84	208.9	204.3	207.5	214.9	215.3
Day  84 - 91	199.6	194.7	199.3	204.0	206.7
Females
Day  0 - 7	139.3	137.1	137.5	135.7	119.3**
Day  7 - 14	143.3	138.8	142.3	140.4	100.9**
Day  14 - 21	151.1	149.9	145.7	148.1	145.2
Day  21 - 28	145.7	141.9	140.8	143.9	154.4
Day  28 - 35	149.6	141.0	141.9	151.2	167.4**
Day  35 - 42	152.9	145.4	141.4	150.4	173.9**
Day  42 - 49	151.6	143.1	146.8	153.9	180.7**
Day  49 - 56	148.2	141.6	139.1	147.5	164.8**
Day  56 - 63	150.4	147.3	145.1	159.1	165.8
Day  63 - 70	151.8	145.0	143.8	153.6	160.3
Day  70 - 77	152.0	145.9	146.2	153.0	163.2
Day  77 - 84	149.6	146.8	145.9	150.3	161.2
Day  84 - 91	142.9	139.8	135.6	145.8	159.9*

*significantly different to controls (p<0.05); **p<0.01)

Summary of FOB

Dose level	0 ppm	6 ppm	20 ppm	60 ppm	200 ppm
Females
Forelimb grip strength (g)
Week 3	1120.4	1095.0	1100.0	1084.2	998.8**
Week 7	1091.3	1111.3	1066.7	1102.5	1036.3
Week 12	1182.5	1157.1	1051.3	1036.7	971.7
Hindlimb grip strength (g)
Week 3	773.3	797.1	760.0	765.8	737.5
Week 7	766.3	792.5	755.8	726.3	658.8**
Week 12	908.3	982.1	915.0	884.6	808.8

*significantly different to controls (p<0.05); **p<0.01

Summary of cholinesterase activity

Dose level	0 ppm	6 ppm	20 ppm	60 ppm	200 ppm
Males
Plasma cholinesterase (U/L)
Week 4	349.0	350.4	292.2	220.0**	181.4**
Week 8	379.0	299.0	295.4	196.0*	155.8**
Week 13	359.5	299.8	313.0	231.2*	216.6**
Erythrocyte cholinesterase (U/L)
Week 4	2255.0	2117.0	2131.4	1711.0*	1206.4**
Week 8	2106.0	2054.8	1839.8	1519.4**	1205.8**
Week 13	2186.5	1978.0	1978.8	1815.0	1299.2**
Brain cholinesterase (U/L)
Week 4	10.06	10.08	9.62	8.00**	3.52**
Week 8	8.30	7.60	8.66	7.19	3.49**
Week 13	9.53	9.26	9.51	8.18**	4.00**
Females
Plasma cholinesterase (U/L)
Week 4	1533.0	1194.6	720.4	513.4*	314.0***
Week 8	2159.4	1914.4	938.8**	505.6**	324.8**
Week 13	2888.4	1396.0	976.2	661.8**	376.8***
Erythrocyte cholinesterase (U/L)
Week 4	2331.6	2218.6	1880.4*	1499.6**	1162.2**
Week 8	1601.0	1589.8	1332.6	1175.8	822.5
Week 13	1650.0	1889.8	1668.2	1217.6	1021.8
Brain cholinesterase (U/L)
Week 4	10.23	9.74	8.72*	4.58**	2.12**
Week 8	9.13	8.74	8.08	3.89**	2.01**
Week 13	9.40	9.62	8.92	3.99**	2.18**

*significantly different to controls (p<0.05); **p<0.01; ***p<0.001

Applicant's summary and conclusion

Conclusions:

A NOAEL of 20 ppm (equivalent to 1.32 and 1.56 mg/kg bw/d in males and females respectively) can be determined for this study based on the inhibition of cholinesterase activity at higher dietary concentrations.

Executive summary:

The neurotoxicity of fenitrothion was investigated in a 90 -day rat study. Groups of 12 male and 12 female Sprague-Dawley rats (main study) and groups of 15 male and 15 female rats (cholinesterase assessment) received fenitrothion in the diet at concentrations of 0, 6, 20, 60 and 200 ppm. Dietary concentrations were calculated to be equal to mean intakes of 0, 0.40, 1.32, 3.99 and 13.8 mg/kg bw/d in males; 0, 0.46, 1.56, 4.85 and 17.6 mg/kg bw/d in females) for 13 weeks. All animals were observed twice a day for their toxic signs and mortality. Body weight was recorded weekly and days of behavioral testing. Food consumption was recorded weekly for all animals. Functional observational battery (FOB) and motor activity assessments were performed pre-test and during Weeks 4, 8 and 13.  At termination, 6 rats/sex/group were randomly selected for perfusion and those in the control and high dose groups underwent a neuropathological evaluation. The remaining animals in each group (6/sex) had their brains removed and then were subjected to necropsy. The brain of each of non-perfused rats in the control and high dose groups was subsequently analyzed for glial fibrillary acidic protein (GFAP). Cholinesterase activities (erythrocyte, plasma, brain) were measured at 4, 8 and 13 weeks (5/sex). No animal died and none were sacrificed prior to scheduled termination. No treatment-related toxic signs were noted in male rats. Four females in the 200 ppm group showed tremors on one or two days during the second week of treatment. The 200 ppm group females also showed a higher incidence of muzzle staining and brown staining along the tail during the study compared to the control group. Both males and females in the 200 ppm group and females in the 60 ppm group had significantly reduced body weights compared to the control group on Day 7. The 200 ppm group's mean body weight remained significantly lower on Day 14 and 21 for males and from Days 7 to 42 for females. Food consumption for the 200 ppm group males and females were significantly lower than the control group from Days 0 to 7. Food consumption for the 200 ppm group females remained significantly reduced from Days 7 to 14 but then increased significantly from Days 28 to 56 and from 84 to 91. No significant differences considered to be related to treatment were seen for the qualitative components of the FOB. Significantly lower values for the 200 ppm group females in forelimb grip strength during the 4th week and in hindlimb grip strength during the 8th week occurred. These differences were considered not to be indicative of neurotoxicity taking lower body weights for the 200 ppm group females. Body temperature values as well as motor activity levels were not significantly different between control and fenitrothion-treated gropus. Reductions in blood and brain cholinesterase levels were noted for males in the 60 and 200 ppm groups and females in the 20, 60 and 200 ppm groups. No gross pathological findings or neuropathological changes (including brain measurements) seen were attributed to treatment with fenitrothion. No significant increases in glial fibrillary acidic protein were detected between the control and high dose groups for either male or females. A NOAEL of 20 ppm (equivalent to 1.32 and 1.56 mg/kg bw/d in males and females respectively) can be determined for this study based on the inhibition of cholinesterase activity at higher dietary concentrations.