3-hydroxy-2-methyl-4-pyrone

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

08 September 2014 - 05 January 2015

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Remarks:

Plasma Bioanalysis investigation to support Maltol in vivo genotoxicity studies

Objective of study:

bioaccessibility (or bioavailability)

Qualifier:

no guideline available

Principles of method if other than guideline:

The report indicated under Regulatory Test Guidelines 'Relevant sections of OECD Guideline 474, 1997 (OECD, 1997) and OECD Guideline 489, 2013 (OECD, 2014).'

In the study, the systemic exposure of rats following oral administration of Maltol was determined. Animals were dosed at 700 mg/kg/day (determined previously to be the maximum tolerated dose), using the same dosing regimen employed in Covance Study Number 8262049 (combined micronucleus and comet test). Whole blood was taken at 0.5, 1, 2 and 3 hours after dosing on Day 3. Plasma was isolated and stored frozen at <-50ºC until analysis. The method of analysis was gas chromatography.

GLP compliance:

yes

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Sigma Aldrich, Germany/STBB9440V
- Expiration date of the lot/batch: February 2016
- Purity test date: >99.9%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Stored at 15-25°C, protected from light

Radiolabelling:

no

Species:

rat

Strain:

other: Han Wistar Crl:WI(Han)

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River (UK) Ltd., Margate, UK
- Age at study initiation: Young adult 97/8 weeks)
- Weight at study initiation: 223-240g
- Housing: Animals were housed in wire topped, solid bottomed cages, with three animals per cage. Bedding was provided on a weekly basis to each cage by use of clean wood bedding (Aspen).
- Diet: ad libitum to SQC Rat and Mouse Maintenance Diet No 1, Expanded (Special Diets Services Ltd. Witham).
- Water: Mains water was provided ad libitum via water bottles
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 23°C
- Humidity (%): 44 to 59%
- Air changes (per hr): 15-20 air changes/hour.
- Photoperiod (hrs dark / hrs light): cycle of 12 hours light (0600 to 1800) and 12 hours dark

Route of administration:

oral: gavage

Vehicle:

other: 0.5% (w/v) aqueous methylcellulose

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Formulations were freshly prepared prior to each dosing occasion by formulating Maltol in 0.5% (w/v) aqueous methylcellulose (0.5% MC) at a concentration of 70 mg/mL for an administered dose of 700 mg/kg bw (dose volume of 10 mL/kg).

Formulations were prepared as follows: the test article was weighed and transferred to a mortar and pestle. A small volume of vehicle was added and mixed to form a smooth paste. The mixture was transferred to the formulation bottle and the mortar and pestle rinsed with the vehicle, which was subsequently added (together with any remaining vehicle) to the formulation bottle to achieve the final volume. Formulations were then mixed using a Silverson until visibly homogenous. To ensure homogeneity, dose formulations were stirred continuously (on a magnetic stirrer) immediately before and throughout dosing.

Duration and frequency of treatment / exposure:

0, 24 and 45 hours

Dose / conc.:

700 mg/kg bw/day

No. of animals per sex per dose / concentration:

6 males

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Covance Study Number 8262049 confirmed that Maltol administered orally at 700 mg/kg, once daily for three consecutive days was well tolerated with no adverse signs of toxicity and only minor body weight loss.

Details on dosing and sampling:

TOXICOKINETIC
- Tissues and body fluids sampled (delete / add / specify): blood, plasma.

- Time and frequency of sampling: Approximately 0.3 mL of whole blood was taken via the jugular vein at 0.5, 1, 2 and 3 hours after dosing on Day 3. Blood was collected at room temperature into lithium heparin tubes and mixed thoroughly using an automatic mixer for two minutes before placing in a cooled kryorack. Following centrifugation (2300 g, 4°C, 10 minutes) plasma was removed, split into two approximately equal aliquots in labelled polypropylene tubes and stored frozen at <-50°C prior to analysis.

-Other:
Bioanalysis Method: Preparation of Standard Solutions

Internal Standard Working Solution (100 μg/mL)
10 mg of Ethyl Maltol was dissolved in 100 mL of t-butyl methyl ether.

Standard Preparation (5mg/mL Maltol in Methanol)
50 mg of Maltol was dissolved in 10 mL of methanol.

Sample Preparation
Samples were prepared as follows:
1 Standards were prepared by spiking 50 uL aliquots of rat plasma with maltol
2 50 uL aliquots of test plasma were prepared
3 250 uL of water + 0.2% methanol was added to each aliquot
4 Aliquots were mixed for 2 min
5 250 uL of internal standard in t-butyl methyl ether (TBME) were added
6 Samples were mixed for 5 min and then centrifuged for 10 min at 3600 rpm
7 The organic layer was removed and submitted for analysis

Calibration Working Standards
A Standard (stock) Prepared to 5 mg/mL

GC parameters:
Column: INNOWax 10m x 0.1 mm x 0.1 μm film

Type:

other: Bioavailability

Results:

Detectable levels of Maltol were found in all animals in plasma samples isolated at 0.5, 1 and 2 hours after dosing. In animals 1-3M Maltol was also detected in plasma isolated 3 hours after dose administration.

Bioaccessibility (or Bioavailability) testing results:

Detectable levels of Maltol were found in all animals in plasma samples isolated at 0.5, 1 and 2 hours after dosing. In animals 1-3M (Table 1). Maltol was also detected in plasma isolated 3 hours after dose administration. Peak plasma levels were seen in the majority of animals 0.5 hours after dose administration. Animal 1M was the exception, with peak plasma levels occurring 1 hour after dose administration. At all time points, plasma values for animals 1M-3M (cage 1) were consistent with each other, as were plasma values for animals 4M-6M (cage 2), however, the levels of plasma between animals in cage 1 versus animals in cage 2 did differ. No technical reasons that could account for this difference were identified.

Table 1: Concentrations of Maltol detected in plasma

Cage Number	Animal	Maltol (μg/mL)/Sample Time (hrs)
	Number/Sex	0.5	1	2	3
1	1M	183.2	208.5	102.8	29
1	1M	134.2	105.3	25	17.6
1	1M	91.4	79/5	29	17.3
2	1M	45.6	25.1	16.5	ND
2	1M	38.2	20.6	16.7	ND
2	1M	42.4	30.8	23.9	ND

ND=not detected

Observations and body weights

Decreased activity was seen in all animals at 1 and 3 hours after dosing on Day 1, with piloerection also observed at 3 hours (Table 7.1). Prior to dosing on Day 2, all animals were observed to have decreased activity and piloerection, however, approximately 5.5 hours later (i.e. immediately after dosing) all animals were observed to be normal and no post dose observations were seen in any animal after dosing on Day 2 or Day 3.

No notable effect of treatment on body weights was observed (Table 7.2).

Conclusions:

In a plasma bioanalysis study, it was concluded that rats dosed with Maltol at 700 mg/kg/day were systemically exposed, with peak plasma levels occurring 0.5 to 1 hour after the final dose
administration.

Executive summary:

In a plasma bioanalysis study (8307783) to support the in vivo micronucleus/comet assay genotoxicity data (8262049), groups of 6 male rats were treated by oral gavage with Maltol (>99.9%) in 0.5% (w/v) aqueous methylcellulose at a dose of 700 mg/kg bw/day. The animals were dosed 3 times (0, 24, 45 hrs) and whole blood was taken via the jugular vein at 0.5, 1, 2 and 3 hours after dosing on Day 3. Plasma samples were prepared and analysed via gas chromatography for Maltol.

Detectable levels of Maltol were found in all animals in plasma samples isolated at 0.5, 1 and 2 hours after dosing. In animals of one cage, Maltol was also detected in plasma isolated 3 hours after dose administration. Peak plasma levels were seen in the majority of animals 0.5 hours after dose administration. Therefore, it was concluded that rats dosed with Maltol at 700 mg/kg/day were systemically exposed, with peak plasma levels occurring 0.5 to 1 hour after the final dose administration.

Reference 2

Endpoint:

basic toxicokinetics in vivo

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

Objective of study:

excretion

metabolism

Qualifier:

no guideline followed

Principles of method if other than guideline:

In a crossover design study, groups of two beagle dogs of each sex were given a single intravenous injection of 10 mg/kg bw maltol or ethyl maltol and urine samples were collected for 3, 6, 24, 48 and 72 hr. The second administration (crossover) was done 7 days after the first. The cumulative urinary excretion of ethyl maltol and maltol conjugates (sulfate and glucuronide) in percent of the administered dose was presented.

GLP compliance:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and of test material: Synthetic metabolites (sulphate, glucuronide) of Maltol were synthesised by the author according to methods in the literature and were considered pure compounds.

Radiolabelling:

no

Species:

dog

Strain:

Beagle

Sex:

male/female

Details on test animals or test system and environmental conditions:

The animals were maintained in metabolism cages and fed once daily; water was given ad libitum. This study was started with fresh (not prestressed) dogs.

Route of administration:

intravenous

Vehicle:

unchanged (no vehicle)

Duration and frequency of treatment / exposure:

IV study: Single dose once and then again 7 days later (crossover design)

Dose / conc.:

10 other: mg/kg bw

Remarks:

IV study

No. of animals per sex per dose / concentration:

IV study: 2 males and 2 females

Control animals:

no

Positive control reference chemical:

None

Details on dosing and sampling:

TOXICOKINETIC STUDY
Urine samples were collected for 3, 6, 24, 48 and 72 hr after each administration (7 days apart).

METABOLITE CHARACTERISATION STUDIES
The protocol also decribes ethyl maltol experiments.

Identification and Synthesis of Metabolites.
Urine excreted over a 24 hr period following intravenous administration of ethyl maltol (10 mg per kg) was examined by paper chromatography (system D, detector reagent a). No unchanged ethyl maltol (Rf 0.7) was found. However, treatment of the urine with mineral acid (refluxing equal volumes of urine and 6N sulfuric acid for 2 hr in oil bath) resulted in the formation of substantial quantities of ethyl maltol. This observation led to the conclusion that ethyl maltol is excreted as one or more conjugates, possibly as ethereal sulfate and/or β-glucuronide. Treatment of the urine with β-glucuronidase (0.2M pH 5 sodium acetate-acetic acid buffer, 1 mg β -glucuronidase per ml of urine, incubated 29 hr at 46C) also regenerated ethyl maltol, indicating the presence of a p-glucuronide. Sequential treatment first with 0.25N hydrochloric acid (15 min at 100C), known to hydrolyze sulfates but not β -glucuronides , then with β - glucuronidase or 6N sulfuric acid (conditions described above) resulted in incremental regeneration of ethyl maltol in both steps, and thus implicated both sulfate and glucuronide as conjugates. Final confirmation of the metabolic route was obtained by identification of the urine components of treated animals with synthetic metabolites.

Hydrolysis of the Metabolites and Colourimetric Determination of the 2-Alkyl 3-Hydroxy-1,4-pyrones Formed.
In order to be amenable for colourimetric determination as their ferric complexes, ethyl maltol and maltol were regenerated from the metabolites by graded acid hydrolysis. Heating the ethereal sulfates in 0.25N sulfuric acid for 15 min at 100" C caused their complete hydrolysis, whereas the glucuronides proved to be stable under these conditions. Hydrolysis of the glucuronides required refluxing in 3N sulfuric acid for 2 hr. Ethyl maltol and maltol were determined colourimetrically as their ferric complexes at 520 nm. Reagent: 1%ferric ammonium sulfate in 0.36M sulfuric acid. Concentrations were read from a standard curve linear in the range of 0.5 to 5 mg %

Isolation of Ethyl Maltol and Maltol from Urine.
The ethyl maltol and maltol formed by hydrolysis of the metabolites was extracted five times from the acid solution with chloroform. After replacement of the chloroform by 40% aqueous methanol the pyrones were determined as described above. Since chloroform extracts of normal urine heated with an equal volume of 6N sulfuric acid undergo a slight colour change in the presence of ferric ions, the intensity of this background colour was determined experimentally and applied as a correction. Mean value of 23 determinations. Absorbance at 520 nm 0.015 (range: 0.005 to 0.028). The colour inherent to excess reagent was found to be negligible (transmission vs water: 99%. Synthetic metabolites were added to normal dog urine, which was then put through the described two step hydrolysis, and extracted with chloroform after each step.

Recoveries: Metabolite Ethyl Maltol Maltol
Sulfate 83 83.5
Glucuronide 90.5 85

These recovery factors, as well as the background colour factor were included in the calculations of ethyl maltol and maltol excreted.


Urinary Excretion.
Determination of free ethyl maltol
Background and recovery. Four normal urine samples (20 ml each) were adjusted to pH 5.0, extracted with five times 20 ml of chloroform, the extract was evaporated in vacuo at 45" C, the residue dissolved in 5 ml of methanol, 1 ml of ferric ammonium sulfate reagent added, and diluted to 10 ml with water. The absorbance was measured at 520 nm against a reagent blank. The mean value for background was found to be 0.1. Recovery (normal urine fortified with ethyl maltol) amounted to 98%. Free ethyl maltol was determined by extraction of urine samples of dosed animals as described above.

Determination of total conjugates
2. 0 ml of urine of each dog of each of the two subsequent 24 hr collections were hydrolyzed by refluxing with equal volumes of 6N sulfuric acid for 2 hr. Extractions and assays were performed as described.

Separate determination of sulphate and glucuronide
An aliquot of the urine samples was mixed with the necessary amount of 2Nsulfuric acid to give a 0.25Nacid solution. The mixture was heated in a steam bath for 15 min, cooled, and extracted with five times 20 ml of chloroform. The combined chloroform extracts were processed and assayed as described. The aqueous phase from the chloroform extraction was mixed with an equal volume of 6N sulfuric acid and refluxed for 2 hr. The hydrolyzate was processed as described above.

Type:

metabolism

Results:

An average of 58.5% of the administered dose was excreted as a mixture of sulfate and glucuronic acid conjugates of maltol at 72hrs.

Type:

excretion

Results:

About 98% of the total urinary excretion of conjugates occurred within the first 24 h, males and females excreting an average of 42% and 73% of the administered dose, respectively.

Details on excretion:

About 98% of the total urinary excretion of conjugates occurred within the first 24 h, males and females excreting an average of 42% and 73% of the administered dose, respectively (Table 2).

Metabolites identified:

yes

Details on metabolites:

An average of 58.5% of the administered dose was excreted as a mixture of sulfate and glucuronic acid conjugates of maltol (Table 2).

Conclusions:

In the Beagle dog, Maltol are predominantly metabolized to sulfate and glucuronic acid conjugates, which are then eliminated in the urine.

Executive summary:

In a metabolism/excretion crossover design study (Rennhard, 1971), Maltol was administered to 2 Beagle dogs of each sex via IV injection at a dose level of 10 mg/kg bw. Urine samples were collected for 3, 6, 24, 48 and 72 hr. The second administration was done 7 days after the first administration. The cumulative urinary excretion of Maltol conjugates (sulfate and glucuronide) in percent of the administered dose was reported.

An average of 58.5% of the administered dose was excreted as a mixture of sulfate and glucuronic acid conjugates of Maltol. About 98% of the total urinary excretion of conjugates occurred within the first 24 h, males and females excreting an average of 42% and 73% of the administered dose, respectively.

Description of key information

A toxicokinetic assessment was conducted in accordance with REACH Annex VIII 8.8.1. The substance Maltol is a solid. It is an organic mono-constituent, with a purity of 80% to 100%, and a typical concentration of 99.9%.

A full ADME toxicokinetic study in the rat is not available. A metabolism/excretion crossover design study in the dog and a plasma bioanalysis study in the rat is available. The Joint FAO/WHO Expert Committee on Food Additives (JECFA) has indicated a metabolic pathway for Maltol [1]. The toxicokinetic analysis is based on this expert opinion, physicochemical and in vivo toxicological data. In vivo studies covering the oral route are available (acute, 90-day repeated dose in the rat and dog, read-across to Ethyl Maltol (CAS No. 4940-11-8) 90-day repeated dose in the rat, read-across to Ethyl Maltol one generation reproductive toxicity/combined chronic toxicity & carcinogenicity study in the rat). No studies via the inhalation route are available. For further details on study summaries, reference is made to the appropriate sections in the IUCLID 6 registration dossier.

Based on the available in vivo toxicological data, Maltol is readily absorbed via the oral route; absorption via the dermal and inhalational routes is expected to be low. Maltol will be distributed throughout the body. The JECFA expert opinion indicates that as the γ-pyrone ring contains a 3-hydroxy substituent, it is expected that maltol will be readily conjugated with glucuronic acid or sulfate and excreted in the urine.

The absorption rates of 50% (oral), 50% (dermal) and 100% (inhalation) are accepted for chemical risk assessment purposes.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Absorption rate - oral (%):

50

Absorption rate - dermal (%):

50

Absorption rate - inhalation (%):

100

Additional information

In accordance with the ECHA Guidance on Information Requirements and Chemical Safety Assessment, Chapter R.7C Section R.7.12 (Endpoint Specific Guidance), the physicochemical properties can provide an insight into the potential behaviour of Maltol in the body.

1.Physicochemical properties

Absorption - oral

The molecular weight of Maltol is 126.11 g/mol which is favourable for oral absorption (<500 g/mol). The log Kow (2.3 at 25°C) indicates it is lipophilic and the water solubility (5791.4 mg/L at 24°C) indicates it is soluble in water. These characteristics will facilitate transport of Maltol via passive diffusion and oral absorption is expected to occur.

Absorption – dermal

The log Kow and water solubility of Maltol are in the optimal range for dermal absorption though the molecular weight is above the favourable range (<100 g/mol). Some dermal absorption can be expected to occur.

Absorption – inhalation

The particle size distribution report for Maltol indicates an MMAD of 352.5 µm (d(0.1) 144.727 μm, d(0.5) 293.779 μm, d(0.9) 555.783 μm), so it is considered to have no dustiness. As none of the particles are available in the inhalable fractions of air (<100 μm), exposure via inhalation is expected to be negligible.

Distribution/Metabolism/Excretion

Based on the molecular weight, water solubility, log Kow and structure, Maltol is likely to be widely distributed, and is expected to be metabolised and excreted in the urine.

2. Other data in the literature

The JECFA indicate that Maltol and its synthetic derivative, Ethyl Maltol, are classified as γ-pyrones [1]. lt is a hydroxyl-substituted 4Hpyran-4-one and is expected be metabolized similarly to phenol, primarily undergoing phase II conjugation of the free hydroxy substituent. In a JECFA-reviewed metabolism/excretion crossover design study (no guideline), Maltol was administered to 2 Beagle dogs of each sex via IV injection at a dose level of 10 mg/kg bw. Urine samples were collected for 3, 6, 24, 48 and 72 hr. The second administration was done 7 days after the first administration. The cumulative urinary excretion of Maltol conjugates (sulfate and glucuronide) in percent of the administered dose was reported. An average of 58.5% of the administered dose was excreted as a mixture of sulfate and glucuronic acid conjugates of Maltol. About 98% of the total urinary excretion of conjugates occurred within the first 24 h, males and females excreting an average of 42% and 73% of the administered dose, respectively.

3. Information from other studies in the dossier

Absorption – oral

In a plasma bioanalysis study (no guideline), groups of 6 male rats were treated by oral gavage with Maltol in 0.5% (w/v) aqueous methylcellulose at a dose of 700 mg/kg bw/day.  Detectable levels of Maltol were found in all animals in plasma samples isolated at 0.5, 1 and 2 hours after dosing. Peak plasma levels were seen in the majority of animals 0.5 hours after dose administration.

In an acute oral study (equivalent or similar to OECD 401), the CD rat LD50 (male) was 1440 mg/kg bw.

In a sub-chronic repeated dose toxicity supporting study (Equivalent or similar to OECD 408), Maltol was administered to 1 group of Charles River male and female rats (10/sex/group) in the diet at dose levels of 0 and 1000 mg/kg bw/day daily for 90 days. There were 2 deaths (sex not specified). Decreased body-weight gain was reported in males and females after weeks 3 and 9, respectively, the male rats being more severely affected. A high incidence of albuminuria was observed in all treated rats. Microscopically, kidney lesions were noted in the rats receiving 1000 mg/kg bw/day. The NOEL (male/female) was <1000 mg/kg bw/day.

In a sub-chronic repeated dose toxicity supporting study (Equivalent or similar to OECD 409), Maltol was administered to 4 groups of Beagle male and female rats (4 per level, not necessarily distributed according to sex) in the diet at dose levels of 0, 125, 250 and 500 mg/kg bw/day daily for 90 days. Three of four animals (sex not specified) at 500 mg/kg bw/day died within 21-41 days, and the fourth was killed when it became moribund. Episcleritis, icteric mucous membranes, emesis, ataxia and prostration was noted before death. Severe weight loss (3.0-4.8 kg) was noted in animals that died before the end of the study. Pathological examination of the tissues revealed pulmonary oedema, hepatic and adrenal cortical and medullary necrosis, fatty degeneration of the myocardium and testicular degeneration. The NOEL (male/female) was 250 mg/kg bw/day.

In a subchronic repeated dose toxicity read-across key study (Equivalent or similar to OECD 408), Ethyl Maltol was administered to 4 groups of Charles River male and female rats (10/sex/group) in the diet at dose levels of 0, 250, 500, 1000 mg/kg bw/day daily for 90 days. Microscopically, kidney lesions were noted in the rats receiving 1000 mg/kg bw/day. The NOAEL (male/female) was 500 mg/kg bw/day. A NOAEL (male/female) of 500 mg/kg bw/day was also predicted for Maltol.

In a one generation reproductive toxicity/combined chronic toxicity & carcinogenicity read-across study (Similar to OECD415/453), Ethyl Maltol was administered to 4 groups of Charles River male and female rats (25/sex/group) in the diet at dose levels of 0, 50, 100 and 200 mg/kg bw/day daily for 2 years. Between 15 and 21 and 30 and 36 weeks, 10 males and 10 females per level were mated to produce two separate litters. All dose levels of ethyl maltol were well tolerated throughout the 2-year feeding period. The test and control animals grew and maintained the body weight in a comparable manner. All rats showed a tendency toward albuminuria; otherwise, urinalysis values were essentially normal. Pathologic changes consistent with aged rats were observed primarily in the pulmonary, endocrine, urinary, and cardiovascular systems. Neoplasia occurred in a random manner with no apparent relationship between number, location, or type of tumors and treatment. Ethyl maltol had no effect on gestation, parturition or lactation. Ethyl maltol had no effect on fetal development and no gross internal abnormalities were noted. The EFSA peer-reviewed NOAEL (parental/offspring) was ≥200 mg/kg bw/day. A NOAEL (parental/offspring) of ≥200 mg/kg bw/day was also predicted for Maltol.

Based on the physicochemical data and available in vivo toxicological data, there is systemic absorption after oral administration. For chemical safety assessment purposes, an oral absorption rate of 50% is accepted.

Absorption – dermal

Maltol is not a skin irritant or skin sensitiser. The ECHA guidance criteria (Chapter R.7C) state that 10% dermal absorption is used when the molecular weight of the substance is >500 and the log Pow is <-1 or >4, otherwise 100% dermal absorption is used. In general, dermal absorption will not be higher than oral absorption, so for chemical safety assessment purposes a dermal absorption rate of 50% is accepted.

Absorption – inhalation

There is no data on inhalation available. For chemical safety assessment purposes, an inhalation absorption rate of 100% is accepted, based on the most conservative approach.

Distribution/Metabolism/Excretion

Based on the expert opinion and available in vivo toxicological data, Maltol will be distributed throughout the body. The JECFA indicate that as the γ-pyrone ring contains a 3-hydroxy substituent, it is expected that maltol will be readily conjugated with glucuronic acid or sulfate and excreted in the urine.

[1] JECFA (2006). Safety evaluation of certain food additives. Who Food Additives Series:56. Prepared by the Sixty fifth meeting of the Joint FAO/WHO Expert Committee on Food Additives (JECFA). World Health Organization, Geneva..