ABSTRACT

With regard to repeat dose toxicity, 1 dam in the 40 mg/kg group died during delivery on day 22 of gestation and 1 dam in the 160 mg/kg group died on day 22 of gestation. Increased liver weights and decreased ovary weights were observed in females of the 160 mg/kg group. In the histopathological findings, a decrease in the degree and incidence of cytoplasmic lipid droplets in the liver was observed in males in the groups treated with 40 mg/kg or more and slight increase in glycogen deposits in the liver were observed in females in the groups treated with 40 mg/kg or more. The NOEL for repeat dose toxicity is considered to be 10 mg/kg/day for both sexes.

With regard to the reproductive/developmental toxicity, the viability index in the 160 mg/kg group tended to be low and the body weights of the pups showed a tendency toward low values on days 0 and 4 of lactation in this group. The NOELs for reproductive and developmental toxicity are considered to be 160 mg/kg/day for males and 40 mg/kg/day for females and their offspring.

2-Hydroxybenzaldehyde was not mutagenic to Salmonella typhimurium TA100, TA98, TA1535, TA1537 and Escherichia coli WP2 uvrA, with or without an exogeneous metabolic activation system.

2-Hydroxybenzaldehyde induced structural chromosomal aberrations and polyploidy in CHL/IU cells in the absence and presence of an exogeneous metabolic activation system.

SUMMARIZED DATA FROM THE STUDY

1. Repeat Dose and Reproductive/Developmental Toxicity １）

Purity	:	99.3%
Test Species/strain	:	Rat/Crj:CD (SD)
Test method	:	OECD Combined Repeat Dose and Reproductive/Developmental Toxicity Screening Test
Route	:	Oral (gavage)
Doses	:	0 (Vehicle), 2.5, 10, 40 and 160 mg/kg/day
Number of animals/group	:	Males, 12; females, 12
Vehicle	:	Olive oil
Administration period	:	Males, 49 days Females, from 14 days before mating to day 3 of lactation
Terminal kill	:	Males, day 50 of administration Females, day 4 of lactation
GLP	:	Yes

　Test results:

<Repeat dose toxicity>

No effects of the administration of the test article were observed in terms of clinical signs, body or organ weights, food consumption, hematological, blood chemical and gross pathological findings. One dam in the 40 mg/kg group died during delivery on day 22 of gestation and 1 dam in the 160 mg/kg group died on day 22 of gestation. In females, increased liver weights and decreased ovary weights were observed in the 160 mg/kg group. Histopathological examination showed a decrease in the degree and incidence of cytoplasmic lipid droplets in the liver in the groups treated with 40 mg/kg or more in males and a slight increase of glycogen deposits in the livers in females of the groups treated with 40 mg/kg or more. The NOEL for repeat dose toxicity is considered to be 10 mg/kg/day for both sexes.

<Reproductive and developmental toxicity>

No effects of the administration of the test article were observed on the estrous cycle, mating and conception, gestation length or delivery, the number of females delivering live pups, the gestation index, the numbers of corpora lutea and implantation sites or the implantation index. In the 160 mg/kg group, all pups of 2 dams died by day 4 of lactation. However, no effects of the test article were observed on nursing in the groups treated with 40 mg/kg or less. Regarding the pups, the viability index in the 160 mg/kg group tended to be low and the body weights of the pups in this group showed a tendency toward low values on days 0 and 4 of lactation. However, there were no effects on the numbers of live and dead pups, the sex ratio, delivery index or live birth index. The NOELs for reproductive and developmental toxicity are considered to be 160 mg/kg/day for males and 40 mg/kg/day for females and their offspring.

2. Genetic Toxicity

2-1. Bacterial test ２）

Purity	:	above 95 %
Test species/strains	:	Salmonella typhimurium TA100, TA1535, TA98, TA1537, Escherichia coli WP2 uvrA
Test methods	:	Guideline for Screening Mutagenicity Testing of Chemicals (Japan) and OECD (471 and 472)
Procedures	:	Plate incorporation method
Solvent	:	DMSO
Positive controls	:	-S9 mix, 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (TA100, WP2, TA98), Sodium azide (TA1535) and 9-Aminoacridine (TA1537) +S9 mix, 2-Aminoanthracene (five strains)
Dosage	:	-S9 mix, 0, 15.6, 31.3, 62.5, 125, 250 and 500 μg/plate (TA100), 0, 31.3 - 1000 (TA1535, TA98 and TA1537) and 0, 125 - 4000 μg/plate (WP2) +S9 mix, 0, 15.6 - 500 (TA100), 0, 62.5 - 2000 μg/plate (TA1535, TA98 and TA1537) and 0, 125 - 4000 μg/plate (WP2).
S9	:	Rat liver, induced with phenobarbital and 5,6-benzoflavone
Plates/test	:	3
Number of replicates	:	2
GLP	:	Yes

　Test results:

The chemical did not induce gene mutations in the S. typhimurium and E. coli strains. Toxicity was observed at 500 μg/plate (TA100 and TA1537), 1000 μg/plate (TA1535 and TA98) and 4000 μg/plate (WP2) without S9 mix and at 500 μg/plate (TA100), 1000 μg/plate (TA1535) and 1500 μg/plate (WP2, TA98 and TA1537) with S9 mix.

Genetic effects:
Salmonella typhimurium TA100, TA1535, TA98, TA1537

	+	?	-
Without metabolic activation:	[ ]	[ ]	[*]
With metabolic activation:	[ ]	[ ]	[*]

Escherichia coli WP2 uvrA

	+	?	-
Without metabolic activation:	[ ]	[ ]	[*]
With metabolic activation:	[ ]	[ ]	[*]

2-2. Non-bacterial in vitro test (chromosomal aberration test) ２）

Purity	:	More than 95%
Type of cell used	:	Chinese hamster lung (CHL/IU) cells
Test method	:	Guidelines for Screening Mutagenicity Testing of Chemicals (Japan)
Solvent	:	Dimethylsulfoxide
Positive controls	:	-S9 mix, Mitomycin C +S9 mix, Cyclophosphamide
Doses	:	-S9 mix (continuous treatment): 0, 0.0050, 0.010,0.020 mg/ml -S9 mix (short-term treatment): 0, 0.050, 0.10, 0.20 mg/ml +S9 mix (short-term treatment): 0, 0.050, 0.10, 0.20 mg/ml
S-9	:	Rat liver, induced with phenobarbital and 5,6-benzoflavone
Plates/test	:	2
GLP	:	Yes

　Test results:

This chemical induce structural chromosomal aberrations and polyploidy in the absence and presence of an exogeneous metabolic activation system.

Lowest concentration producing cytogenetic effects in vitro:
without metabolic activation (continuous treatment ):	0.020 mg/ml (clastogenicity) 0. 020 mg/ml (polyploidy)
Without metabolic activation (short-term treatment):	0.10 mg/ml (clastogenicity)
With metabolic activation (short-term treatment):	0.10 mg/ml (clastogenicity) 0. 050 mg/ml (polyploidy)

Genotoxic effects:

	clastogenicity			polyploidy
	+	?	-	+	?	-
Without metabolic activation:	[*]	[ ]	[ ]	[*]	[ ]	[ ]
With metabolic activation:	[*]	[ ]	[ ]	[*]	[ ]	[ ]

1)	The tests were performed by Bozo Research Center Inc, 1284, Kamado, Gotemba-shi, Shizuoka, 412, Japan. Tel +81-550-82-2000 Fax +81-550-82-2379
2)	The tests were performed by the Hatano Research Institute, Food and Drug Safety Center, 729-5 Ochiai, Hadano-shi, Kanagawa, 257, Japan. Tel +81-463-82-4751 Fax +81-463-82-9627