Safety evaluation of certain food additives - ipcs inchem

ALKOXY-SUBSTITUTED ALLYLBENZENES 363
dependent (Drinkwater et al., 1976; Swanson et al., 1981; Miller et al., 1982, 1983;
Boberg et al., 1983; Gardner et al., 1995, 1996; Daimon et al., 1998). Sulfate
inhibition studies and in vivo–in vitro unscheduled DNA synthesis (UDS) assays of
myristicin, elemicin, estragole, methyl eugenol and the 1-hydroxy metabolites of
estragole and methyl eugenol (Boberg et al., 1983; Caldwell et al., 1992; Chan &
Caldwell, 1992; Hasheminejad & Caldwell, 1994) provide additional evidence that
the sulfate ester of the 1-hydroxy metabolite is the principal intoxication metabolite
in animals.
Additional biochemical studies have provided information on the influence of
dose and species on the formation of the 1-hydroxy metabolite, the cytochrome
P450 (CYP) isoenzymes that catalyse the 1-hydroxylation pathway and the
formation of protein and DNA adducts with the 1-hydroxy metabolite.
The metabolism in animals of methoxy- and methylenedioxy-substituted
allylbenzenes is determined by dose. At low doses in vivo, CYP-catalysed oxidation
and cleavage of the O-methylene of the methylenedioxy substituent of myristicin,
apiole or safrole are by far the predominant pathway, yielding polar
dihydroxyallylbenzene metabolites that are readily excreted either free or as sulfate
or glucuronic acid conjugates. At high doses in rodents, O-demethylenation
becomes saturated, and 1-hydroxylation and epoxidation of the allyl side-chain
compete. This change in the balance of metabolic pathways from O-dealkylation to
1-hydroxylation with dose has been demonstrated better for methoxy-substituted
allylbenzenes, notably estragole (Anthony et al., 1987), than for methylenedioxysubstituted
allylbenzenes (Smith et al., 2002). This shift may be due to the increased
oxidative cleavage of methylene from the methylenedioxy function. Since the 1hydroxylation
and the subsequent formation of the 1-sulfoxy metabolite have been
associated with the toxicity and carcinogenicity of these substances, the effect of
dose and, to a lesser extent, species significantly impacts the interpretation of the
results of toxicity and carcinogenicity studies.
In a three-part metabolism study (Beyer et al., 2006), groups of rats were
administered safrole, myristicin or elemicin, either individually or as constituents of
powdered nutmeg. Urinary metabolites were then compared with those of a human
consuming a relatively large dose of ground nutmeg. Male Wistar rats were
administered oral doses (100 mg/kg bw) of the nutmeg ingredients myristicin,
safrole or elemicin, and the urine was collected separately from faeces over the next
24 h. In a second phase of the study, rats were given an aqueous suspension of
ground nutmeg at 500 mg/kg bw from two different batches. In addition, urine
samples were collected from a nutmeg-abusing inpatient who reportedly consumed
large amounts of nutmeg powder. Urine samples of all dosed groups were treated
with glucuronidase and arylsulfatase, then acetylated and subjected to gas
chromatographic/mass spectrometric (GC/MS) analysis.
The 24-h urine of rats dosed with safrole or myristicin at 100 mg/kg bw
contained metabolites derived from O-demethylenation, epoxidation and hydrolysis
(as a 2,3-dihydroxy derivative) and 1-hydroxylation (see Figure 2), with the peak
area of the O-demethylenation metabolite being at least 10 times that of any other
metabolites. Rats treated with elemicin at 100 mg/kg bw primarily showed