Glycerides, C16-18 mono-, di- and tri-, hydrogenated, citrates, potassium salts

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Acceptable, well-documented publication meeting basic scientific principles. Study was part of subchronic oral toxicity study. Test compound was a mixture of short- and long-chain acyl triglyceride. Information is used in a read.across approach. For justification of read-across and for further details please refer to the read-across report.

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Genetic Toxicology Studies of SALATRIM Structured Triacylglycerols. Lack of Genetic Damage in in Vitro Mammalian Cell Assays and the in Vivo Micronucleus Assay

Author:

Hayes, J.R. et al.

Year:

1994

Bibliographic source:

J. Agrlc. Food Chem. 42, 521-527

Reference 2

Reference Type:

publication

Title:

Subchronic Toxicity Studies of SALATRIM Structured Triacylglycerols in Rats. 3. Triacylglycerols Composed of Stearate, Acetate, Propionate, and Butyrate

Author:

Hayes, J.R. et al.

Year:

1994

Bibliographic source:

J. Agric. Food Chem. 42, 552-562

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

although test was performed within a oral subchronic toxicity study

Deviations:

yes

Remarks:

, no positive controls

Principles of method if other than guideline:

The SALATRIM family of triacylglycerols differs from other fats in the ratio of short-chain fatty acids (SCFA) to long-chain fatty acids (LCFA) and in that stearic acid is the major LCFA. These fats have caloric availability values (4.5-6 kcal/g) lower than that of corn oil (9 kcal/g). SALATRIM 234CA lot A019 and SALATRIM 234CS lot A018 were tested in the in vivo bone marrow micronucleus assay. Rats received these SALATRIM fats or corn oil at 10% (w/w) in the diet for 13 weeks.
The in-life and bone marrow smear preparation portions of the study were part of a 13-week subchronic toxicity study conducted at Hazleton Wisconsin, Inc., Madison, WI, and the micronucleus assay portion of the study was conducted at Hazleton Washington, Vienna, VA. Detailed information on the experimental design and methods for the subchronic study can be found in the paper for the subchronic toxicology portion of that study (Hayes et al., 1994).

GLP compliance:

not specified

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

short- and long-chain triacylglycerols (SALATRIM 234CA)

IUPAC Name:

short- and long-chain triacylglycerols (SALATRIM 234CA)

Constituent 2

Reference substance name:

short- and long-chain triacylglycerols (SALATRIM 234CS)

IUPAC Name:

short- and long-chain triacylglycerols (SALATRIM 234CS)

Constituent 3

Reference substance name:

SALATRIM 234CA

IUPAC Name:

SALATRIM 234CA

Constituent 4

Reference substance name:

SALATRIM 234CS

IUPAC Name:

SALATRIM 234CS

Details on test material:

- Name of test material (as cited in study report): SALATRIM 234CA lot A019 and SALATRIM 234CS lot A018

SALATRIM 234CA lot A019 and SALATRIM 234CS lot A018 are typical SALATRIM fats. SALATRIM 234CA lot A019 was produced from triacetin, tripropionin, tributyrin, and hydrogenated canola oil. This results in a relatively random distribution of acetic, propionic, and butyric acids among the SCFA esterified to glycerol. SALATRIM 234CS lot A018 was similar except the precursor fat was hydrogenated cottonseed oil. This resulted in a fat similar to SALATRIM 234CA lot A019 with stearic acid still the predominant LCFA but with a slightly different distribution of other LCFA.

Members of the SALATRIM family of structured triacylglycerols are typical triacylglycerols composed of fatty acids esterified to a glycerol backbone. The unique characteristic of this family of fats is the triacylglycerols contain a preponderance of stearic, acetic, propionic, and/
or butyric acids. Triacylglycerols that contain one stearic acid and two short-chain fatty acids (SCFA) predominate among the mixed triacylglycerols that comprise these fats. Because of the limited absorption of stearic acid compared to certain other fatty acids and the fewer carbons available for energy production from the SCFA, these fats have lower caloric availabilities (4.5-6 kcal/g) than fats such corn oil (9 kcal/g). SALATRIM fats are produced by interesterification among high-stearate fats, such as hydrogenated canola oil and hydrogenated soybean oil, and triacetin, tripropionin, and tributyrin. Because the natural precursor fats contain mixed triacylglycerols, SALATRIM fats contain small quantities of other fatty acids such as oleic and palmitic, among others. SALATRIM fats can have different physical and functional characteristics because the ratios of the SCFA can be varied.

Test animals

Species:

rat

Strain:

other: Crl:CD BR VAF/Plus

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI, USA
- Diet: ad libitum

Administration / exposure

Route of administration:

oral: feed

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
SALATRIM fats and corn oil were mixed separately with NIH-07 Rat and Mouse Ration 5018 (Purina Mills, Inc., St. Louis, MO) and fed ad libitum.

Duration of treatment / exposure:

13 weeks
Groups of 20 male and 20 female Crl:CD BR VAF/Plus rats from Charles River Laboratories, Portage, MI, were exposed to either of the two SALATRIM fats or corn oil at 10% (w/w) of the diet for at least 13 weeks.

Frequency of treatment:

mixed in diet

Post exposure period:

none

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
10 % (w/w)
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
approx. 7000 mg/kg bw/d
Basis:
other: actual ingested as calculated by measured food consumption

No. of animals per sex per dose:

20

Control animals:

other: 10% corn oil in diet as negative control group

Positive control(s):

Because these data were collected from a 13-week subchronic toxicity study, a positive control for micronuclei formation was not included.

Examinations

Tissues and cell types examined:

bone marrow

Details of tissue and slide preparation:

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
At necropsy, duplicate bone marrow slides for each rat were prepared for clinical pathology. One set of unstained bone marrow slides was shipped to Hazelton Washington for the micronucleus assay.

DETAILS OF SLIDE PREPARATION/METHOD OF ANALYSIS:
Upon arrival, the slides were fixed in methanol and stained with acridine orange. A bone marrow slide from each of the 20 rata per group (with the exception of the SALATRIM 234CS lot A018 group, where only 18 slides were available) was analyzed using fluorescent microscopy. One thousand polychromatic erythrocytes per rat were scored for micronuclei. Identification of micronuclei was based upon the criteria of Schmid (1976). The scoring unit was the micronucleated cell, not the number of micronuclei. Therefore, a cell that contained more than one micronucleus was scored as a single micronucleated cell. The frequency of micronucleated cells was expressed as percent micronucleated cells based upon the total number of polychromatic erythrocytes scored. The normal frequency of micronucleated erythrocytes for this strain of rat is 0.0 - 0.4 % .
Analyses were conducted separately for each SALATRIM-treated group and each sex combination.

Evaluation criteria:

The criteria for a positive response was a significant increase in micronucleated polychromatic erythrocytes compared with the corn oil control group. If no increase was found, the test fats were considered negative in this assay.

Statistics:

Statistical analysis of the data was by analysis of variance on the square root arcsine transformation.
Tukey’s Studentized range test (Sokal and Rohlf, 1981) was used to determine statistical significance (p < 0.05) from the corn oil control group.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

not examined

Additional information on results:

Comparison of the data for the two SALATRIM fats with that from the corn oil group indicated that neither SALATRIM fat increased the incidence of micronucleated polychromatic erythrocytes. Therefore, the SALATRIM fats were considered to be negative in the assay.

Any other information on results incl. tables

Table 3:Percent Micronucleated Polychromatic Erythrocytes in Bone Marrow of Rats Fed Diets Containing 10% SALATRIM 234CA Lot A019, SALATRIM 234CS Lot

A018, and Corn Oil for 13 Weeks*

 

treatment	males (% MN/1000 PCE)	females (% MN/1000 PCE)	combined male and female (% MN/1000 PCE)
corn oil	0.18 ± 0.11	0.17 ± 0.13	0.17 ± 0.12
SALATRIM 234CA lot A019	0.25 ± 0.17	0.17 ± 0.11	0.21 ± 0.14
SALATRIM 234CS lot A018	0.21 ± 0.17	0.12 ± 0.12	0.16 ± 0.14

MN = micronucleated cells

PCE = polychromatic erythrocytes

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
The test items were negative in this assay. The data confirm the prediction that SALATRIM fats lack genotoxic potential.

Executive summary:

In a Crl:CD BR VAF/Plus rats bone marrow micronucleus assay, 20 animals/sex/dose were fed with SALATRIM 234CA and 234CS at a dose of 7000 mg/kg/d daily for 13 weeks. Bone marrow cells were harvested post-treatment.  

There were no signs of toxicity during the study. A positive control was not included because the data were collected from a 13-week subchronic toxicity study. There was no significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow after any treatment time.

This study is classified as acceptable.  This study satisfies the requirement for Test Guideline OECD 474 for in vivo cytogenetic mutagenicity data.