Glycerol trioctanoate

Carcinogenicity

Description of key information

Oral (2-year oral gavage study): NOAEL male rat: 2390 mg/kg bw/day

Key value for chemical safety assessment

Carcinogenicity: via oral route

Link to relevant study records

Reference

Endpoint:

carcinogenicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Remarks:

Acceptable, well documented technical report which meets basic scientific principles.

Reason / purpose for cross-reference:

reference to same study

Principles of method if other than guideline:

Male rats were exposed to corn oil, safflower oil and tricaprylin via gavage. After 15 months (interim evaluation) and 2 years, animals were sacrificed for gross and histopathological examinations to evaluate the treatment-related effects on neoplasm incidence.

GLP compliance:

yes

Species:

rat

Strain:

other: Fischer 344/N

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Simonsen Laboratories (Gilroy, CA)
- Age at study initiation: 7 weeks
- Weight at study initiation: 126 - 159 g (range)
- Housing: groups of 5 animals in polycarbonate cages with hardwood chips
- Diet: NIH-07 open-stock mash diet (Zeigler Bros., Inc., Gardners, PA), ad libitum
- Water: Worcester public water, ad libitum
- Acclimation period: 14 to 22 days; prior to study start, five rats from each study were randomly selected and killed for parasite evaluation and gross observation of disease.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-22
- Humidity (%): 47.1 ± 4.9
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Tricaprylin was dispensed into vials for gavage dosing on a weekly basis. After dispensing, the test item was stored at 4°C for no more than 3 weeks.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Accelerated bulk chemical stability studies were performed using gas chromatography. The test substance was found to be stable as a bulk chemical for 2 weeks when stored protected from light at temperatures up to 60°C. The stability of tricaprylin was monitored periodically by ultraviolet spectroscopy and gas chromatography. In addition, the peroxide concentration was determined prior to use. The acceptable peroxide concentration was set at 2 mEq/L. A bottle was discareded if it exceeded this specification. No significant degradation of the bulk chemical was observed throughout the study.

Duration of treatment / exposure:

24 months (50 males per group)
15 months (10 males per group, interim group)

Frequency of treatment:

daily; 5 days/week

Post exposure period:

None

Dose / conc.:

2 390 mg/kg bw/day (actual dose received)

Remarks:

2.5 mL/kg bw/day, dose calculated based on a density of 0.9540 g/cm³

Dose / conc.:

4 770 mg/kg bw/day (actual dose received)

Remarks:

5 mL/kg bw/day, dose calculated based on a density of 0.9540 g/cm³

Dose / conc.:

9 540 mg/kg bw/day (actual dose received)

Remarks:

10 mL/kg bw/day, dose calculated based on a density of 0.9540 g/cm³

No. of animals per sex per dose:

60 males/group; 10 from each group were sacrificed after 15 months

Control animals:

yes, concurrent no treatment

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily, findings were recorded at least monthly

BODY WEIGHT: Yes
- Time schedule for examinations: at study initiation, than weekly for 13 weeks, and monthly thereafter

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at 15 months. Blood was collected from the posterior vena cava.
- How many animals: 10
- Parameters checked: haematocrit, haemoglobin, erythrocyte count, mean erythrocyte haemoglobin, mean erythrocyte haemoglobin concentration, mean erythrocyte haemoglobin volume, reticulocyte count, total and differential leukocyte count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at 15 months from the posterior vena cava
- How many animals: 10
- Parameters checked: potassium, total protein, albumin, cholesterol, alanine aminotransferase, creatine kinase, sorbitol dehydrogenase, bile acids

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, on all animals. Animals found in a moribund state, selected for the 15-month interim evaluations, or surviving to the end of the 2-year study were killed by the use of "Biotol", an ultra fast-acting barbiturate. All organs and tissues were examined for gross lesions.
HISTOPATHOLOGY: Yes, on all animals. All of the following tissues were preserved in 10% neutral puffered formalin, embedded in paraffin, sectioned, and stained with hematoxylin and eosin for microscopic examination: Adrenal gland, bone and marrow, brain, esophagus, gross lesions, heart, kidney, large intestine (cecum, colon, rectum), liver, lung, mammary gland, lymph nodes (mandibular and mesenteric), pancreas, parathyroid gland, pituitary gland, preputial gland, salivary gland, skin, small intestine (duodenum, jejunum, ileum), spleen, stomach, testis with epididymis and seminal vesicle, thymus, thyroid gland, tissue masses, trachea, and urinary bladder.

Other examinations:

Brain, kidney (right) and liver of each animal were weighed at the 15 month interim evaluation.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

9540 mg/kg bw/day: Clinical findings including dyspnoea, ataxia and lethargy were observed in 50/60 animals in the high-dose group. However, most of the animals generally recovered prior to the next daily dosing and the incidence of clinical findings declined during the second half of the study.

Mortality:

mortality observed, treatment-related

Description (incidence):

9540 mg/kg bw/day: 30 animals died, other group: comparable survival rats
The two-year survival rate was lower in test animals compared with the untreated controls with statistical significance for the high-dose group only (31/50, 30/50, 31/50, 23/53 for control, 2390, 4770, 9540 mg/kg bw/day, respectively (Table 1)). Twenty-three rats of the high-dose group died or were killed between weeks 33 and 85. Ten of these animals were found dead and 13 were killed moribund. Although the cause of death or moribund condition could not-be-determined in 20 of these animals, clinical findings were noted in all of the moribund animals. Of these, eight rats died or were killed between weeks 45 and 49, when the incidence of clinical findings was highest. The average body weight of these eight animals was 316 g, which was significantly less than the mean group body we
ight of approximately 360 g for high-dose group at 11 months. Only one of the moribund animals had a pulmonary mass that might have explained the dyspnoea. Up to four animals in the high-dose group died or were killed in each subsequent month.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

9540 mg/kg bw/day: Mean body weights were decreased in test animals compared to controls throughout the study. However, the difference was less than 5% after week 61. No effects were observed in the low- and mid-dose group.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

A dose-dependent decrease in food consumption was determined in test animals (table 3) which resulted in a decreased protein consumption per day (protein consumption (g/rat/day): 3.88, 3.45, 3.09 and 2.70, for control, low-, mid- and high-dose animals, respectively). This effect is most probably due to the fact that rats exposed to tricaprylin received more than 10% of their caloric intake from the test item (10.4%, 19.8% and 35.4% for the low-, mid- and high-dose animals, respectively). Thus, the effect on food consumption is considered as non-adverse.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

Significant increases in the haematocrit, haemoglobin and erythocyte counts were determined for the high-dose group. No effects were observed in the low- and mid-dose group.

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

Test animals of the mid-dose group revealed a statistically significant lower absolute kidney weight. As the relative organ weight is not different from the control and the effect is not observed in the low- and high-dose group, the effect is considered as incidental and not treatment-related.

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No adverse effects observed.
A significant decrease in the incidence of nephropathy was determined in the high-dose group (control: 46/50, 2390 mg/kg bw/day: 42/50, 4770 mg/kg bw/day: 45/50, 9540 mg/kg bw/day: 27/49) with a dose-related decrease in severity (control: 2.0, 2390 mg/kg bw/day: 1.5, 4770 mg/kg bw/day: 1.7, 9540 mg/kg bw/day: 0.9). As the severity of nephropathy is normally related to the amount of protein in the diet, the decrease in severity induced by tricaprylin might be due to the lower food consumption in test animals. No further findings have been determined in control vs test animals.

Histopathological findings: neoplastic:

effects observed, treatment-related

Description (incidence and severity):

dose-related increase in the incidences of pancreatic exocrine hyperplasia and adenoma; proliferative lesions of the forestomach.
Tricaprylin induced dose-related increases in the tumor incidences of pancreatic exocrine hyperplasia reaching statistically significance for the mid-and high-dose group (control: 8/49, 2390 mg/kg bw/day: 9/49, 4770 mg/kg bw/day: 18/49, 9540 mg/kg bw/day: 28/50), adenoma (control: 2/49, 2390 mg/kg bw/day: 6/49, 4770 mg/kg bw/day: 13/49, 9540 mg/kg bw/day: 18/50) and proliferative lesions in the forestomach (basal cell hyperplasia: control: 4/50, 2390 mg/kg bw/day: 7/50, 4770 mg/kg bw/day: 12/49, 9540 mg/kg bw/day: 21/52; squamous cell papilloma: control: 0/50, 2390 mg/kg bw/day: 0/50, 4770 mg/kg bw/day: 3/50, 9540 mg/kg bw/day: 10/53) reaching statistically significance for squamous cell papilloma in the high-dose group. In contrast, a decrease in the incidence of mononuclear cell leukemia was observed in high-dose animals (control: 23/50, 2390 mg/kg bw/day: 28/50, 4770 mg/kg bw/day: 22/50, 9540 mg/kg bw/day: 9/53). The incidence of pancreatic islet hyperplasia and adenoma or carcinoma decreased in a dose-dependent, but not significant manner (hyperplasia: control: 6/49, 2390 mg/kg bw/day: 5/48, 4770 mg/kg bw/day: 3/49, 9540 mg/kg bw/day: 1/49; adenoma/carcinoma: control: 5/49, 2390 mg/kg bw/day: 2/48, 4770 mg/kg bw/day: 3/49, 9540 mg/kg bw/day: 1/49)

Key result

Dose descriptor:

NOAEL

Remarks:

carcinogenicity

Effect level:

2 390 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Dose descriptor:

LOAEL

Remarks:

carcinogenicity

Effect level:

4 770 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

histopathology: neoplastic

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

4 770 mg/kg bw/day (actual dose received)

System:

gastrointestinal tract

Organ:

pancreas

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

4 770 mg/kg bw/day (actual dose received)

System:

gastrointestinal tract

Organ:

other: forestomach

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

no

Table 1: Mortality data after tricaprylin exposure

	control	2390 mg/kg bw/day	4770 mg/kg bw/day	9540 mg/kg bw/day
number of animals included in the study	60	60	60	60
sacrificed for 15-month interim evaluation^a	10	10	10	7
natural deaths	4	7	4	13
moribund kills	15	13	15	17
animals surviving until study termination	31^d	30	31^d	23^d
mean survival (days)^b	651	639	642	553
survival analysis^c	P = 0.004	P = 0.944	P = 0.969	P = 0.014

a: censored from survival analyses

b: mean of all death including uncensored, censored and terminal sacrifice

c: The result of the life table trend test (Tarone, 1975) is shown in the control column whereas the results of life table pairwise comparisons (Cox, 1972) with the controls are listed in the dosage columns.

d: Includes 2 rats for the control and 1 rat for the mid- and high-dose groups that died during the last week of the study.

 

Table 2: Effects on body weight after tricaprylin exposure

	body weight (g)
weeks on study	control	2390 mg/kg bw/day	4770 mg/kg bw/day	9540 mg/kg bw/day
1	146	144	145	145
2	172	171	169	168
3	199	198	193	190
4	219	215	215	207
5	243	241	239	231
7	263	259	255	247
8	266	265	257	254
9	283	282	277	266
10	300	295	285	278
12	306	301	294	283
13	306	301	296	284
14	317	311	303	289
17	332	329	319	303
21	351	350	339	319
25	363	367	354	330
29	376	372	357	335
33	381	379	371	345
37	386	387	376	353
41	388	389	377	347
45	389	389	382	358
49	388	395	388	368
53	401	408	396	380
57	404	411	394	380
61	403	416	400	390
65	408	419	399	392
69^a	406	421	400	392
73	408	422	403	395
77	414	423	406	403
81	413	419	405	404
85	405	412	396	399
89	402	412	390	395
93	400	413	393	397
97	393	406	386	391
101	387	404	379	379
104	386	391	371	366
mean (week 1–13)	246	243	239	232
mean (week 14–52)	367	367	357	335
mean (week 53–104)	402	413	394	390

a: interim evaluation occurred during week 67

 

Table 3: Effects on food consumption after tricaprylin exposure

	food consumption per animal per day (g)
weeks on study	control	2390 mg/kg bw/day	4770 mg/kg bw/day	9540 mg/kg bw/day
2	14.2	13.7	12.5	11.4
5	16.9	15.9	16.0	14.4
9	17.1	16.5	16.4	13.6
13	13.0	12.1	9.9	8.9
17	20.2	16.9	16.0	13.5
21	15.8	14.9	12.5	10.0
25	18.3	18.3	16.5	13.5
29	17.8	16.2	14.0	12.2
33	17.8	16.3	16.0	13.7
37	18.3	15.6	14.2	12.1
41	15.2	12.8	11.5	8.8
45	16.7	14.9	12.5	10.8
49	20.3	17.7	15.8	13.3
53	18.2	16.2	14.2	12.1
57	20.4	17.5	14.1	12.7
61	17.9	16.5	14.2	12.5
65	16.8	13.7	11.9	10.6
69	18.6	17.3	16.5	15.6
73	16.8	14.1	12.9	10.2
77	17.7	14.3	12.9	11.2
81	17.7	14.5	13.2	10.7
85	15.9	14.3	12.2	10.3
89	13.9	12.6	10.6	9.5
93	16.6	14.4	15.9	15.1
97	16.5	12.3	12.3	12.9
101	13.7	12.3	9.4	8.7
104	13.5	10.6	9.2	7.6
mean	17.0	15.1	13.6	11.8

 

Table 4: Haematology data after tricaprylin exposure at the 15-month interim evaluation

	control	2390 mg/kg bw/day	4770 mg/kg bw/day	9540 mg/kg bw/day
number of animals	9	10	8	7
haematocrit (%)	45.6 ± 1.1	50.7 ± 2.7	49.2 ± 2.3	51.7 ± 2.0*
haemoglobin (g/dL)	15.6 ± 0.4	17.3 ± 0.8	16.7 ± 0.6	17.5 ± 0.6*
erythrocytes (10^6/ µL)	8.77 ± 0.31	9.72 ± 0.41	9.30 ± 0.34	9.91 ± 0.32*

*: significantly different from control by Dunn´s or Shirley´s test (P ≤ 0.05)

 

Table 5: Organ weights of the kidney (right) after tricaprylin exposure at the 15-month interim evaluation

	control	2390 mg/kg bw/day	4770 mg/kg bw/day	9540 mg/kg bw/day
number of animals	10	10	10	7
body weight at necropsy	430 ± 15	419 ± 11	402 ± 15	406 ± 12
absolute	1.349 ± 0.04	1.23 ± 0.036	1.194 ± 0.041*	1.328 ± 0.053
relative	3.15 ± 0.07	2.95 ± 0.07	2.98 ± 0.07	3.27 ± 0.08

*: significantly different from the control by William´s or Dunnett´s test (P ≤ 0.05)

Organ weights are given in grams; organ-weight-to body-weight ratios are given as mg organ/g bw

 

Table 6: Histopathological findings after tricaprylin exposure

		Untreated control	2390 mg/kg bw/day	4770 mg/kg bw/ day	9540 mg/kg bw/day
nephropathy incidence		46/50 (92%)	42/50 (84%)	45/50 (90%)	27/49 (55%)*
nephropathy severity grade		2.0 ± 0.12	1.5 ± 0.13**	1.7 ± 0.11*	0.9 ± 0.13**
forestomach: basal cell hyperplasia		4/50	7/50	12/49	21/52
mononuclear cell leukemia (trend test)		23/50 (46%) p=0.029	28/50 (56%) p=0.205	22/50 (44%) p=0.525	9/53 (17%) p=0.071
pancreatic islet hyperplasia		6/49	5/48	3/49	1/49
pancreatic islet adenoma/carcinoma		5/49	2/48	3/49	1/49

* Significantly different (p≤0.05) from the control group by the logistic regression test (overall rate) or the Mann-Whitney U test (severity)

**p≤0.01

Average severity grade is given as mean f standard error. 0 = none, 1 = minimal, 2 = mild, 3 = moderate, and 4 = marked

Table 7: Incidences of Proliferative Lesions of the Exocrine Pancreas

	Untreated Control	2390 mg/kg bw/day	4770 mg/kg bw/day	9540 mg/kg bw/day
Hyperplasia -overall rate	8/49 (16%)	9/49 (18%)	18/49 (37%)*	28/50 (56%)**
Adenoma – overall rate	2/49 (4%)	6/49 (12%)	13/49 (27%)	18/50 (36%)
Adenoma – terminal rate	2/30 (7%)	6/30 (20%)	11/31 (35%)	16/23 (70%)
First incidence (days)	729 (terminal sacrifice)	729 (terminal sacrifice)	518	485
Logistic regression test	p<0.001	p=0.129	p=0.002	p<0.001

*Significantly different (p≤0.05) from the control group by the logistic regression test

**p≤0.01

Beneath the control incidence are the p values associated with the trend test. Beneath the dosed group incidences are the p values corresponding to pairwise comparisons between the controls and that dosed group. The logistic regression test regards neoplasms as nonfatal.

Table 8 Incidences of Squamous Cell Papilloma (Neoplasms of the Forestomach)

	Untreated Control	2390 mg/kg bw/day	4770 mg/kg bw/day	9540 mg/kg bw/day
Overall rate	0/50 (0%)	0/50 (0%)	3/50 (6%)	10/53 (19%)
First incidence (days)	-	-	623	623
Logistic regression test	p<0.001	-	p=0.118	p<0.001

Beneath the control incidence is the p value associated with the trend test. Beneath the dosed group incidence are the p values corresponding to pairwise comparisons between the controls and that dosed group. The logistic regression test regards neoplasms as nonfatal.

Conclusions:

A statistically significant dose-related increase in pancreatic acinar cell hyperplasia and adenoma was found, but there were no acinar carcinomas. The incidence of mononuclear leukemia was less, and nephropathy findings were reduced. No level of evidence of carcinogenicity was assigned by the study authors.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

2 390 mg/kg bw/day

Study duration:

chronic

Species:

rat

Quality of whole database:

The available information comprise an adequate and reliable (Klimisch score 2) study with the target substance.

System:

gastrointestinal tract

Organ:

pancreas

Carcinogenicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Carcinogenicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Mode of Action Analysis / Human Relevance Framework

Not applicable.

Justification for classification or non-classification

The available data on carcinogenicity are inconclusive according to Regulation (EC) 1272/2008.

Additional information

Repeated dose toxicity, 2-year oral gavage study

CAS 538-23-8

The chronic oral toxicity of glycerol trioctanoate (CAS 538-23-8), also named tricaprylin, was investigated in male F344/N rats in comparison to corn oil and safflower oil (key, 1994). The National Toxicology Program (NTP) designed studies to evaluate the role of several oils in altering cancer rates in male rats. Control male rats receiving corn oil (often used as vehicle) have previously been shown to have a higher incidence of pancreatic proliferative lesions and a lower incidence of mononuclear cell leukaemia than untreated control males. To evaluate 3 gavage vehicles for potential systemic toxicity and carcinogenicity, corn oil, safflower oil, and tricaprylin (CAS 538-23-8) were administered by gavage to male F344/N rats for 2 years. Sixty animals per dose were administered tricaprylin at 2.5, 5 or 10 mL/kg bw once daily for 5 days per week by oral gavage. The doses were equivalent to 2390, 4770 and 9540 mg/kg bw/day as calculated using a density of 0.954 g/cm³. Sixty control animals remained untreated.

All non-neoplastic study results (including clinical signs and mortality) are summarized under the endpoint repeated dose toxicity. In high-dose animals a decrease in the incidence of mononuclear cell leukaemia was observed; this is not considered to have toxicological significance. Proliferative lesions of the forestomach (basal cell hyperplasia, squamous cell papilloma) were seen in the mid- and high-dose groups. These are considered to be treatment-related, but are most likely not relevant for humans as humans do not have a forestomach. A dose-dependent increase in the incidence of proliferative lesions of the exocrine pancreas (hyperplasia and adenoma) was seen; the overall rates were statistically significant in the mid- and high-dose group. The mechanism by which dietary fats or triglycerides stimulate proliferation of the exocrine pancreas in the rat is unclear, but the effect has been reported in public literature. No pancreatic carcinoma was observed in rats given glycerol trioctanoate in this study. Based on the high doses administered to rats over 2 years via oral gavage, the absence of pancreatic carcinoma in this study and the establishment of a NOAEL, no clear evidence of genotoxic carcinogenicity is shown for glycerol trioctanoate (CAS 538-23-8). The NOAEL for non-genotoxic carcinogenicity was found to be 2390 mg/kg bw/day based on the study results.