1,2,3-trichloropropane

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: - scientifically sound study, according to OECD 408 - journal publication presenting only limited numerical results

Data source

Reference

Reference Type:

publication

Title:

Cardiopathic effect of 1,2,3-trichloropropane after subacute and subchronic exposure in rats.

Author:

Merrick BA, Robinson M, Condie LW.

Year:

1991

Bibliographic source:

J Appl Toxicol. 1991 Jun;11(3):179-87. PMID: 1918791

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Principles of method if other than guideline:

in addition the results from a 10 d dose range finder study are included

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

1,2,3-trichloropropane

EC Number:

202-486-1

EC Name:

1,2,3-trichloropropane

Cas Number:

96-18-4

Molecular formula:

C3H5Cl3

IUPAC Name:

1,2,3-trichloropropane

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles Rivers Laboratories, Inc. Portage, MI, U.S.A.
- Age at study initiation: 70 d at arrival, 80 at start of administration
- Weight at study initiation: not reported
- Fasting period before study: no
- Housing: housed in polycarbonate cages containing hardwood chip bedding (Absorb-Dri, Maywood, NY),no. of animals per cage not reported
- Diet (e.g. ad libitum): ad libitum with Purina Rodent Chow (No. 5001, Ralston Purina Co., St. Louis, MO)
- Water (e.g. ad libitum): distilled drinking water that had been carbon-filtered and UV-irradiated
- Acclimation period: 10 d


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-22
- Humidity (%): 40-60
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: 1,2,3-trichloropropane was dissolved in corn oil at 1 mL/kg.

VEHICLE
- Justification for use and choice of vehicle (if other than water): corn oil is a standard vehicle for organic solvents
- Concentration in vehicle: 90 d study: 0.01, 0.05, 0.1 and 0.4 mol/L (= 1.47, 7.37, 14.74, 58.97 g/L)
10 d study: 0.01, 0.05, 0.2 and 0.8 mol/L (= 1.47, 7.37, 29.49, 117.94 g/L)
- Amount of vehicle (if gavage): 1 mL/Kg
- Provider: Sigma Chemical Company (St. Louis, MO, U:S.A.)

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Duration of treatment / exposure:

10d and 90d,

Frequency of treatment:

single daily exposure via gavage

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
0.01, 0.05, 0.1 and 0.4 mmol/Kg bw (= 1.47, 7.37, 14.74, 58.97 mg/Kg bw)
Basis:
other: nominal via gavage in the 90 d study

Doses / concentrations 2

Remarks:

Doses / Concentrations:
0.01, 0.05, 0.2 and 0.8 mmol/Kg bw (= 1.47, 7.37, 29.49, 117.94 mg/Kg bw)
Basis:
other: nominal via gavage in the 10 d study

No. of animals per sex per dose:

not reported

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: 10 d range finding study: high dose selected as one-quarter of the reported acute oral dose of 440 mg/Kg/d (= 3.0 mmol/Kg bw); 90 d study: based on the results of the 10 d study 0.4 was selected as the high dose
- Rationale for animal assignment (if not random): random

Positive control:

no

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No data


DETAILED CLINICAL OBSERVATIONS: No data


BODY WEIGHT: Yes
- Time schedule for examinations: 90 d study: weekly; 10 d study: prior to first dosing and at sacrifice


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No


FOOD EFFICIENCY: No


WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: twice weekly


OPHTHALMOSCOPIC EXAMINATION: No data


HAEMATOLOGY: No data

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at terminal sacrifice
- Anaesthetic used for blood collection: Yes (60 mg/Kg Nembutal (Abbott Las, Chicago, IL, U.S.A.)
- Animals fasted: No
- How many animals: all
- Parameters examined: not reported in detail, at least ALT, AST, BUN and creatinine


URINALYSIS: No


NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, see below
HISTOPATHOLOGY: Yes, see below
organ weights + lights microscopy: liver, kidneys, lung, spleen, heart, adrenal glands, thymus and brain
histopathology: analysis of high dose and controls; low and mid dose only for tissues where adverse effects were seen in the high dose group
liver, kidneys, lung, spleen, heart, adrenal glands, thymus, brain, alimentary canal (tongue, esophagus, stomach, duodenum,
jejunum, ileum, caecum, colon and rectum), femur bone, mandibular and mesenteric lymph nodes, mammary glands, thigh muscle, nasal cavity, sciatic nerve, ovaries, pancreas, parathyroids, pituitary, preputial or clitoral glands, prostate, salivary glands, seminal vesicles, skin, testis and epididymis, thyroid gland, urinary bladder, uterus, Zymbal's glands and all gross lesions.
- femurs and nasal structures decalcified prior to trimming and fixation
- grading from 1 - 4 of the severity of inflammatory, degenerative and necrotic lesions in tissuesd.
Histopathological evaluation was performed by a board-certified veterinary pathologist (Pathology Associates Inc., Ijamsville, MD).

Statistics:

- analysis of variance (ANOVA), with differences between control and 1,2,3-TCP-treated groups evaluated by Duncan's multiple comparison
tes
- not normally distributed data: Wilcoxin Rank Sum test for treatment differences and the Kruskal-Wallis Rank Sums test for differences among means.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

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):

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

effects observed, treatment-related

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

effects observed, treatment-related

Details on results:

CLINICAL SIGNS AND MORTALITY
- no mortalities or adverse clinical signs reported

BODY WEIGHT AND WEIGHT GAIN
- reduced body weight gain in the males and females of the high dose groups both in the 10 d and the 90 d study
- see table 1

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
not examined

FOOD EFFICIENCY
not examined

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
not effects detected

OPHTHALMOSCOPIC EXAMINATION
not examined

HAEMATOLOGY
- slightly reduced white blood cell count in males after the 10-day exposure but no such effect in females or any of the dose groups in the 90 d study

CLINICAL CHEMISTRY
- 10 d study: significantly increased ALT and AST values at the high dose
- 90 d study: significantly raised ALT and AST levels at the high dose in female rats only, raised values in males were compromised by very high ALT and AST levels in male controls, mirrored by histopathological findings in this control group

URINALYSIS
not examined

NEUROBEHAVIOUR
not examined

ORGAN WEIGHTS
- absolute organ weights:
heart, spleen, thymus and lung weight reduction in the high doses of both sexes of both studies
- relative organ weights:
generally no differences in relative weights of heart, lung and spleen (and ovaries in females)
relative thymus weight decreased in the high dose groups of both sexes in the 10 d study (but not in the 90 d study)
generally raised relative liver and kidney weights in the two highest doses groups in both sexes in both studies.
effects in the relative brain weights were explained by the reduced body weights in the high dose groups


GROSS PATHOLOGY
no macroscopically visual changes reported

HISTOPATHOLOGY: NON-NEOPLASTIC
- heart:
diffuse inflammation-associated necrosis of cardiac myocardiumin both sexes in all treatment groups in the 90 d study but only in the high dose group in the 10 d study. The prevalence for males was higher then for females. There is no clear dose response to this effect. (see table 2). The severity of cardiac lesions is reported to increase dose dependently (no detailed values given).
This cardiac lesions were reported by the authors to occur also in Long Evans rats and F344 rats if treated for 10 d at 0.8 mmol/Kg bw/d (high dose of the 10 d study).
- thymus:
atrophy in the high doses groups of both sexes in the 10 d study, no effects in the 90 d study
- liver:
10 d study: mild centrilubolar hepatic necrosis in one female and two males of the high dose
90 d study:
females: minimal liver necrosis in one animal in the 0.05 mmol/Kg dose group and in two animals of the high dose group
males: mild to moderate liver necrosis with increasing dose related incidence (1 - 4) in the respective dose groups. The findings were confounded by mild hepatic lesions in four of the male control animals


HISTOPATHOLOGY: NEOPLASTIC (if applicable)
- increased prevalence of bile duct hyperplasia in both sexes in the high dose group in the 90 d study
- plasma cell hyperplasia in the mandibular lymph nodes at variable extent in control males and females, but increased incidence in males exposed to high dose after 10 days; in the 90 d study a dose dependently increased prevalence of mandibular lymph node hyperplasia was found for both sexes
- No lesions in mesenteric, subcutis, pancreatic, bronchiolar or mediastinal lymph nodes
subchronic exposure conditions.
- Several other proliferative and neoplastic lesions in individual animals at high dose treatment: broncho-alveolar adenoma in a male, an adenocarcinoma of a female mammary gland, a forestomach squamous cell papilloma in a male and forestomach squamous cell hyperplasia in a male.

Effect levels

Dose descriptor:

NOAEL

Sex:

male/female

Basis for effect level:

other: at the lowest dose in the 90 d study inflammation and necrosis in the heart was seen. Nevertheless no clear dose response could be seen for this effect neither in males nor females.

Remarks on result:

not determinable

Remarks:

no NOAEL identified

Target system / organ toxicity

Critical effects observed:

yes

Lowest effective dose / conc.:

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

System:

cardiovascular

Organ:

heart

Treatment related:

yes

Dose response relationship:

no

Relevant for humans:

not specified

Any other information on results incl. tables

- Table 1: Final body weights of rats after subacute and subchronic exposure to 1,2,3 -trichloropropane

	Dose mmol/kg/day)	Body weight (g) after 10-day exposure	Dose (mmol/kg/day)	Body weight (g) after 90-day exposure
Male
	0.00	451 ± 8	0.00	529 ± 13
	0.01	410 ± 13	0.01	528 ± 16
	0.05	436 ± 7	0.05	509 .±. 11
	0.20	417 ± 8	0.10	533 ± 16
	0.80	337 ± 12*	0.40	426 ± 13*
Female
	0.00	253 ± 4	0.00	301 ± 5
	0.01	256 ± 3	0.01	301 ± 9
	0.05	262 ± 4	0.05	307 ± 10
	0.20	260 ± 3	0.10	301 ± 6
	0.80	195 ± 6*	0.40	258 ± 7*

Body weights were measured on the day of sacrifice. Values represent means ± SEM for 10 rats per group. An asterisk indicates means that are significantly different from the control value at P < 0.05.

- Table 2: Incidence of organ histopathology of rats after subacute and subchronic exposure to 1,2,3 -trichloropropane

Target organ	Male					Female
mmol/kg/day x 10 days
	0.00	0.01	0.05	0.20	0.80	0.00	0.01	0.05	0.20	0.80
Heart
Inflammation	1	0	0	0	10	0	0	0	0	10
Degeneration	0	0	0	0	10	0	0	0	0	10
Necrosis	0	0	0	0	10	0	0	0	0	10
Mandibular lymph	4	-	-	-	7	1	-	-	-	0
node
Thymus Diffuse atrophy	0	0	0	0	6/8	0	0	0	0	6/6
mmol/kg/day x 90 days
	0.00	0.01	0.05	0.10	0.40	0.00	0.01	0.05	0.10	0.40
Heart
Inflammation	0	3	2	3	8	0	1	2	1	6
Degeneration	0	0	0	1	5	0	0	1	1
Necrosis	0	2	1	2	6	0	0	1	0	7
Liver
Bile duct	1	0	0	0	4	0	0	0	0	8
hyperplasia .,...
Mandibular lymph node	2	0	1	0	9	1	1	2	3	5

Tissues were prepared for histological evaluation by staining with hematoxylin and eosin. Tissues of 10 animals were evaluated for each dose group unless otherwise indicated. A dash means that the tissues were not evaluated.

Applicant's summary and conclusion

Conclusions:

The toxicity of 1,2,3-trichloropropane was tested in Sprague Dawley rats in a 90 d repeated dose study with oral treatment via gavage (5 d/wk). Results for a 10 d dose range finding experiment are included.
A NOAEL cannot be defined as a raised prevalence of inflammation-associated diffuse necrosis of the cardiac myocardium was seen in all dose groups of the 90 d experiment but with an uncommon dose response pattern. In addition slight renal and liver weight effects and corresponding clinical chemical and histological findings in the liver were determined in the two highest dose groups of the 90 d study. As neoplastic lesions a significant increase in bile duct hyperplasia above the age related background was reported and a higher incidence of adenoma papilloma and carcinoma scattered over different tissues.

Executive summary:

In the present study (Merrick 1991) the toxicity of 1,2,3-trichloropropane was tested in Sprague Dawley rats in a 90 d repeated dose study with oral treatment via gavage (5 d/wk) according to OECD TG 408. Results for a 10 d dose range finding experiment are included.

A NOAEL could not be derived from the reported results as a raised prevalence of inflammation-associated diffuse necrosis of the cardiac myocardium was seen in all dose groups of the 90 d experiment but with an uncommon dose response pattern.

Animals aged 80 d at study initiation were treated with 1,2,3 -trichloropropane in corn oil, weighted weekly and their water consumption was determined twice weekly. At terminal sacrifice blood samples were taken for clinical chemical analysis and hematology. These animals were subjected to gross necroscopy and full histological examination.

No mortalities and adverse clinical signs were reported. Reduced body weight gain was found in the males and females of the high dose groups both in the 10 d and the 90 d study. No treatment related effects were seen regarding water consumption of treated animals as compared to controls. Hematologic examination revealed a slightly reduced white blood cell count in males after the 10-day exposure but no such effect in females or any of the dose groups in the 90 d study, therefore this finding is regarded as incidental. For the 10 d study: significantly increased ALT and AST values at the high dose are reported. Accordingly also in the 90 d study significantly raised ALT and AST levels at the high dose in female rats only were determined. Raised values in males of the 90 d study were compromised by very high ALT and AST levels in male controls, mirrored by histopathological findings in this control group. Several absolute organ weights are reduced (heart, spleen, thymus and lung) in the high doses of both sexes of both studies but as the body weights are reduced in these groups the more relevant relative organ weights of heart, lung and spleen (and ovaries in females) showed generally no differences. Nevertheless the relative thymus weight was decreased in the high dose groups of both sexes in the 10 d study (but not in the 90 d study) and generally raised relative liver and kidney weights in the two highest doses groups in both sexes in both studies were reported. Effects in the relative brain weights were explained by the reduced body weights in the high dose groups.

Several non-neoplastic lesions were revealed by the histologic examination, the most prominent being found in the heart as diffuse inflammation-associated necrosis of cardiac myocardiumin both sexes in all treatment groups in the 90 d study but only in the high dose group in the 10 d study. The prevalence for males was higher then for females and there is no clear dose response to this effect. The severity of cardiac lesions is reported to increase dose dependently (no detailed values given). This cardiac lesions were reported by the authors to occur also in Long Evans rats and F344 rats if treated for 10 d at 0.8 mmol/Kg bw/d (high dose of the 10 d study).

In thymus atrophy in the high doses groups of both sexes in the 10 d study is reported, but no effects in the 90 d study, therefore this effect is not regarded as adverse. For the liver mild centrilubolar hepatic necrosis in one female and two males of the high dose was detected in the 10 d study. In the 90 d study in females minimal liver necrosis in one animal in the 0.05 mmol/Kg dose group and in two animals of the high dose group was found while in males mild to moderate liver necrosis with increasing dose related incidence (1 - 4) in the respective dose groups was detected. The latter findings were confounded by mild hepatic lesions in four of the male control animals.

Several indications for preneoplastic lesions were also noted as there are an increased prevalence of bile duct hyperplasia in both sexes in the high dose group in the 90 d study, plasma cell hyperplasia in the mandibular lymph nodes at variable extent in control males and females, but increased incidence in males exposed to high dose after 10 days plus in the 90 d study a dose dependently increased prevalence of mandibular lymph node hyperplasia.

Other proliferative and neoplastic lesions in individual animals at high dose treatment comprised broncho-alveolar adenoma in a male, an adenocarcinoma of a female mammary gland, a forestomach squamous cell papilloma in a male and forestomach squamous cell hyperplasia in a male.

No lesions in mesenteric, subcutis, pancreatic, bronchiolar or mediastinal lymph nodes subchronic exposure conditions.

A derivation of a NOAEL or a LOAEL is not possible for the 90 d study. All adverse effects occur at the highest dose with two exceptions. Firstly relative weight effects in the liver and liver enzyme effects are seen also at the second highest dose (0.1 mmol/Kg/d = 14.74 mg/Kg/d) but these effects are confounded by a uncommon mild liver necrosis in 4 of the control animals. Secondly a significant prevalence of inflammation-related degeneration and necrosis of cardiac myocardium was seen in males of the high dose and to a lesser extent in the females of this dose level. In both sexes there is no clear dose response: 3/10 males showing inflammation at 0.01 mmol/Kg/d, 2/10 at 0.05, 3/10 at 0.1 and 8/10 at 0.4. The respective prevalences for females are 1/10, 2/10, 1/10 and 8/10.

Prevalences of degeneration and necrosis show a comparable variability.

Using the concept of LOAEL and NOAEL strictly would imply a to set the LOAEL for this study to the lowest dose (0.01 mmol/Kg/d = 1.47 mg/Kg bw/d). Nevertheless the complete absence of cardiac effects in a 120 d gavage study and even a 2 year chronic gavage study at comparable dose levels (see chapter 7.5.1: Hazleton (1983) / key (120d study on F344 rats) (8 - 250 mg/Kg bw/d) and chapter 7.7: National Toxicology Program (1993) / Rat and Mouse (3 - 30 and 6 - 60 mg/Kg bw/d, respectively)) renders the correlation between the cardiopathic effects and the 1,2,3 -trichloropropane exposition questionable.