A 13-week subchronic toxicity study of dietary administered morin in F344 rats

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Abstract

A subchronic toxicity study of a flavonoid morin was performed in both sexes of F344 rats with dietary administration at concentrations of 0%, 0.625%, 1.25%, 2.5% and 5% (w/w) for 13 weeks. No mortality or abnormal clinical signs were observed throughout the experimental period in any group. Although a slight tendency for increase in food intake was noted in both sexes of the 2.5% and 5.0% groups, slight non-significant body weight decrease was observed in 5.0% males. Significant increases in alanine transaminase (ALT; over 2.5%), alkali phosphatase (ALP; 1.25% and 5.0%) and relative liver weights (1.25% and 2.5%) in males and in γ-glutamyl transpeptidase (γ-GT), aspartate transaminase (AST), ALT, relative liver weights in the 2.5% and 5.0% females and ALP in 5.0% females were noted. Increased urea nitrogen and relative kidney weights at dose of 1.25% and above and creatinine at 5.0% were observed also in females. On histopathological observation, hepatocyte hypertrophy was detected in 3 of 10 5.0% females. Based on the above findings, the no-observed-adverse-effect level (NOAEL) for both sexes was estimated to be 0.625% (299 and 356 mg/kg b.w./day for males and females, respectively).

Introduction

Morin (2′,3,4′,5,7-pentahydroxyflavone), a constituent of many fruits and herbs, e.g. mulberries, figs and other Moraceae, is a flavonoid which is used as a food additive because of its antioxidant activity (Hanasaki et al., 1994, Ramanathan et al., 1994). In addition, it possesses anti-inflammatory potential through modulatory effects on lipoxygenase and cycloxygenase activities in the arachidonic cascade (Baumann et al., 1980, Galvez et al., 2001, Nakadate et al., 1984). Morin is also reported to be active against phenobarbital promotion of rat liver tumor development after initiation with N-nitrosodiethylamine (DEN) (Denda et al., 1989) and to inhibit 4-nitroquinoline 1-oxide (4-NQO)-induced rat tongue carcinogenesis either in the initiation or post-initiation stage, and azoxymethane (AOM)-induced rat colorectal carcinogenesis in the post-initiation stage. These effects are considered to be due to induction of phase II enzymes and/or suppression of cell proliferation activity by morin (Kawabata et al., 1999, Tanaka et al., 1999).

However, quercetin, a positional isomer of morin and a naturally occurring flavonol in plants, induces gene mutations in Salmonella with and without exogenous metabolic activation, mutations in Drosophila, and chromosomal aberrations and sister chromatid exchange in mammalian cell cultures (Brown, 1980, Carver et al., 1983, IARC, 1983, Meltz and MacGregor, 1981, Nagao et al., 1981, National Toxicology Program, 1992). Renal toxic lesions, with increased severity of chronic nephropathy and tumors of the tubular epithelium, were also found in an oral carcinogenicity study in male rats (Dunnick and Hailey, 1992), although other long-term rat studies indicated no carcinogenic effects (Hirono et al., 1981, Ito et al., 1989). Toxicological effects of morin are therefore conceivable, but no data in vivo are available except for the documented pro-oxidant activity in isolated rat liver nuclei (Sahu and Gray, 1997) and oxidative effects on hemoglobin in vitro (Kitagawa et al., 2004). While morin induces gene mutations in Salmonella, the activity is weak (Nagao et al., 1981). The present study was therefore conducted to determine toxic effects and to establish a no-observed-adverse-effect level (NOAEL) with oral administration to F344 rats.

Section snippets

Test chemical

The morin sample (CAS no. 480-16-0, Fig. 1) obtained from San-Ei Gen F.F.I. (Osaka, Japan) was a yellowish brown powder with a purity of >95% (w/w), which was extracted from branch, trunk and root parts of Broussonetia xanthoxylum MARTIUS using ethanol extraction and purification. The compound is insoluble in water or oil but soluble in ethanol or alkali solutions, and stable at room temperature in a dark place (internal data of San-Ei Gen F.F.I.). For the present study the compound was mixed

Results

Neither mortality nor deterioration in general conditions were observed in any of the groups. A slight reduction of body weight gain was noted in 5.0% males without statistical significance (Table 1). As shown in Table 1, data for food consumption and total chemical intake showed a tendency for increase in both sexes of the 2.5% and 5% groups, resulting in greater than expected doses of the test substance. Hematological data are shown in Table 2, Table 3. WBCs for females of the 2.5% and 5.0%

Discussion

In the present study of morin administration in the diet at concentrations of 0%, 0.625%, 1.25%, 2.5% or 5.0% to male and female F344 rats for 13 weeks, a number of test substance-related changes were observed in organ weights and parameters in hematology, serum biochemistry and histopathology. For example, the 2.5 and 5.0% females exhibited significantly increased WBCs and the 5.0% females significantly decreased segmented neutrophils. Although adverse nutritional effects should be avoided

Acknowledgement

This study was supported by a Grant-in-Aid from the Ministry of Health, Labour and Welfare of Japan.

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