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WHO FOOD ADDITIVES SERIES: 48

SAFETY EVALUATION OF CERTAIN
FOOD ADDITIVES AND CONTAMINANTS

ALIPHATIC ACYCLIC DIOLS, TRIOLS,
AND RELATED SUBSTANCES

First draft prepared by Dr P.J. Abbott, Professor A.G. Renwick2 and Professor I.G. Sipes3
1
Australia New Zealand Food Authority, Canberra, Australia
2Clinical Pharmacology Group, University of Southampton, Southampton, United Kingdom
3 Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona, USA

Evaluation

Introduction

Estimated daily intake

Metabolic considerations

Application of the Procedure for the Safety Evaluation of Flavouring Agents

Consideration of combined intakes

Conclusions

Relevant background information

Explanation

Additional considerations on intake

Biological data

Biochemical data

Absorption and transformation

Metabolism

Toxicological studies

Acute toxicity

Short-term studies of toxicity

Long-term studies of toxicity and carcinogenicity

Genotoxicity

Reproductive toxicity

References

1. EVALUATION

1.1 Introduction

The Committee evaluated a group of 31 flavouring agents1 that included aliphatic acyclic diols, triols, and related substances (see Table 1) using the Procedure for the Safety Evaluation of Flavouring Agents (see Figure 1, Introduction). All members of this group are aliphatic acyclic primary alcohols, aldehydes, acids, or related esters with one or more additional oxygenated functional groups. The group contains four subgroups: glycerol (No. 909) and 15 related glycerol esters and acetals (Nos 910, 911, 912, 913, 914, 915, 916, 917, 918, 919, 920, 921, 922, 923, and 924); propylene glycol (No. 925) and four related esters, acetals, and ketals (Nos 926, 927, 928, and 929); lactic acid (No. 930) and four lactate esters (Nos 931, 932, 934, and 935); and pyruvic acid (No. 936), its corresponding aldehyde (No. 937), two pyruvate esters (Nos 938 and 939) and one acetal of pyruvic acid (No. 933).

Table 1. Summary of results of safety evaluations of aliphatic acyclic diols, triols, and related substances

Flavouring agent

No.

CAS No. and structure

Step A3b Does intake exceed the threshold for human intake?

Step A4 Is the flavouring agent or are its metabolites endogenous?

Conclusion based on current intake

Structural class I

Glycerolc,d

909

56-81-5

Yes
Europe: 17 000
USA: 220 000

Yes Glycerol is endogenous

Evaluation not finalized

1,2,3-Tris[(1´-ethoxy)]-propane

913

67715-82-6

No
Europe: 0
USA: 140

NR

No safety concern

Glyceryl monostearatec

918

123-94-4

No
Europe: 0
USA: 230

NR

Evaluation not finalized

Glyceryl monooleatec

919

111-03-5

No
Europe: ND
USA: 860

NR

Evaluation not finalized

Triacetinc

920

102-76-1

Yes
Europe: ND
USA: 83 000

Yes Expected to be hydrolysed to glycerol, which is endogenous

Evaluation not finalized

Glyceryl tripropanoatec

921

139-45-7

No
Europe: 0.1
USA: 280

NR

Evaluation not finalized

Tributyrinc

922

60-01-5

No
Europe: 31
USA: 2

NR

Evaluation not finalized

Glycerol 5-hydroxy-decanoatec

923

26446-31-1

No
Europe: 4
USA: 0

NR

Evaluation not finalized

Glycerol 5-hydroxydo-decanoatec

924

26446-32-2

No
Europe: 4
USA: 0

NR

Evaluation not finalized

Propylene glycolc,e

925

57-55-6

Yes
Europe: ND
USA: 2 400 000

Yes Expected to be oxidized to lactic acid, which is endogenous

Evaluation not finalized

Propylene glycol stearatec

926

142-75-6

Yes
Europe: ND
USA: 66 000

Yes Expected to be oxidized to propylene glycol and subsequently to lactic acid

Evaluation not finalized

1,2-Di[(1-ethoxy)-ethoxy)] propanec

927

67715-79-1

No
Europe: 7
USA: 150

NR

No safety concern

Lactic acid

930

598-82-3

Yes
Europe: ND
USA: 47 000

Yes Lactic acid is endogenous

No safety concern

Ethyl lactatef

931

97-64-3

Yes
Europe: 1900
USA: 760

Yes Expected to be hydrolysed to lactic acid, which is endogenous

No safety concern

Butyl lactate

932

138-22-7

No
Europe: 380
USA: 24

NR

No safety concern

Potassium 2-(1´-ethoxy) ethoxy-propanate

933

No
Europe: ND
USA: 1400

NR

No safety concern

cis-3-Hexenyl lactate

934

61931-81-5

No
Europe: 38

USA: 5NR

No safety concern

Butyl butyryl lactate

935

7492-70-8

No
Europe: 280
USA: 1400

NR

No safety concern

Pyruvic acid

936

127-17-3

No
Europe: 35
USA: 69

NR

No safety concern

Pyruvaldehyde

937

79-98-8

No
Europe: 120
USA: 3

NR

No safety concern

Ethyl pyruvate

938

617-35-6

No
Europe: 1
USA: 20

NR

No safety concern

Isoamyl pyruvate

939

7779--72-8

No
Europe: 0
USA: 0

NR

No safety concern

Structural class III

3-Oxohexanoic acid glyceride

910

91052-72-1

Yes
Europe: 0
USA: 270

Yes Expected to be hydrolysed to glycerol, which is endogenous

No safety concern

3-Oxooctanoic acid glyceride

911

91052-68-5

No
Europe: 34
USA: 0

NR

No safety concern

Heptanal glyceryl acetal (mixed 1,2 and1,3 acetals)

912

1708-35-6

No
Europe: 4
USA: 0

NR

No safety concern

3-Oxodecanoic acid glyceridec

914

91052-69-6

Yes
Europe: 0
USA: 270

Yes Expected to be hydrolysed to glycerol, which is endogenous

Evaluation not finalized

3-Oxododecanoic acid glyceridec

915

91052-70-9

No
Europe: 73
USA: 0

NR

Evaluation not finalized

3-Oxotetradecanoic acid glyceridec

916

91052-73-2

Yes
Europe: 0
USA: 270

Yes Expected to be hydrolysed to glycerol, which is endogenous

Evaluation not finalized

3-Oxohexadecanoic acid glyceridec

917

91052-71-0

No
Europe: 43
USA: 0

NR

Evaluation not finalized

4-Methyl-2-pentyl-1,3- dioxolane

928

26563-74-6

No
Europe: 0
v

NR

No safety concern

2,2,4-Trimethyl-1,3-oxycyclopentane

929

1193-11-9

No
Europe: 0.3
USA: 0.2

NR

No safety concern

CAS: Chemical Abstracts Service; ND: no data available; NR: not required for evaluation because consumption of the substance was determined to be of no safety concern at Step A3 of the Procedure.

a

Step 2: All of the flavouring agents in this group are expected to be metabolized to innocuous products.

b

The thresholds for human intake are 1800 µg/day for structural class I and 90 µg/day for structural class III. All intake values are expressed in µg/day.

c

Further information is required to determine whether this substance is currently used as a flavouring agent.

d

An ADI ‘not specified’ was established for glycerol by the Committee at its twentieth meeting (Annex 1, reference 41), which was maintained at the present meeting.

e

An ADI of 0–25 mg/kg bw was established for propylene glycol by the Committee at its seventeenth meeting (Annex 1, reference 32), which was maintained at the present meeting.

f

Ethyl lactate was included in the group ADI ‘not specified’ for lactic acid and its salts that was established by the Committee at its twenty-sixth meeting (Annex 1, reference 59), which was maintained at the present meeting.

The Committee previously evaluated three members of the group. Glycerol (No. 909) was considered at the twentieth meeting, when an ADI "not specified" was established (Annex 1, reference 41). Propylene glycol (No. 925) was considered at the seventh meeting, when an ADI of 0–20 mg/kg bw was established (Annex 1, reference 7); it was further considered at the seventeenth meeting, when the ADI was increased to 0–25 mg/kg bw (Annex 1, reference 32). Ethyl lactate (No. 931) was considered at the eleventh (Annex 1, reference 14), twenty-third (Annex 1, reference 50), twenty-fourth (Annex 1, reference 53), and twenty-sixth meetings (Annex 1, reference 59). At its twenty-sixth meeting, the Committee included ethyl lactate in the group ADI ‘not specified’2 with lactic acid.

Nine of the 31 substances (Nos 909, 929, 930, 931, 932, 934, 936, 937, and 938) have been detected as natural components of foods in cocoa, milk, cider, cognac, asparagus, tomato, and mushrooms (Maarse et al., 1999).

1.2 Estimated per capita intake

The total annual production of the 31 substances in this group of flavouring agents for use in food was reported to be 140 000 kg in Europe (International Organization of the Flavor Industry, 1995) and 21 000 000 kg in the USA (Lucas et al., 1999). These values are equivalent to a total daily per capita intake of 20 000 µg in Europe and 2 800 000 µg in the USA. The large difference in the annual volume of production between Europe and the USA is due to the inclusion in the USA of figures on the use of glycerol, triacetin, and propylene glycol as solvents in the preparation of compound flavour mixtures.

In Europe, three flavouring agents, namely, glycerol (17 000 µg/day), ethyl lactate (1900 µg/day), and butyl lactate (380 µg/day), accounted for approximately 97% of the total per capita intake. In the USA, three substances, namely glycerol (220 000 µg/day), triacetin (83 000 µg/day), and propylene glycol (2 400 000 µg/day), accounted for 96% of the total annual daily per capita intake. The per capita intakes of individual substances are shown in Table 2.

Table 2. Annual volumes of use of aliphatic acyclic diols, triols and related substances used as flavouring agents in Europe and the USA

Substance (No.)

Most recent annual volume (kg)

Intakea

 

Annual volume in naturally occurring foods (kg)b

Consumption ratioc

 

 

µg/day

µg/kg bw per day

 

 

Glycerol (909)

Europe

120 000

17 000

280

24 000 000

200

USA

1 700 000

220 000

3700

 

14

1,2,3-Tris[(1’-ethoxy)ethoxy]propane (913)

Europe

0

0

0

 

USA

1000

140

2.3

 

 

3-Oxohexadecanoic acid glyceride (917)

Europe

300

43

0.7

 

USA

0

0

0

 

 

Glyceryl monostearate (918)

Europe

0

0

0

 

USA

1800

230

4

 

 

Glyceryl monooleate (919)

Europe

NR

NA

NA

 

USA

6500

860

14

 

 

Triacetin (920)

Europe

NR

NA

NA

 

USA

630 000

83 000

1400

 

 

Glyceryl tripropanoate (921)

Europe

0.5

0.1

0.002

 

USA

2100

280

5

 

 

Tributyrin (922)

Europe

220

31

0.5

 

USA

15

2

0.03

 

 

Glycerol 5-hydroxydecanoate (923)

Europe

25

4

0.007

 

USA

0

0

0

 

 

Glycerol 5-hydroxydodecanoate (924)

Europe

25

4

0.07

 

USA

0

0

0

 

 

Propylene glycol (925)

Europe

NR

NA

NA

 

USA

18 000 000

2 400 000

40 000

 

 

Propylene glycol stearate (926)

Europe

NR

NA

NA

 

USA

500 000

66 000

1100

 

 

1,2-Di[(1-ethyoxy)ethyoxy]propane (927)

Europe

49

7

0.1

 

USA

1200

150

2.5

 

 

Lactic acid (930)

Europe

NR

NA

NA

11 000 000

 

USA

360 000

47 000

780

 

30

Ethyl lactate (931)

Europe

13 000

1900

32

250 000

19

USA

5800

760

13

 

43

Butyl lactate (932)

Europe

2600

380

6

+

 

USA

190

24

0.4

 

 

Potassium 2-(1´-ethoxy)ethoxypropanate (933)

Europe

NR

NA

NA

 

USA

10 000

1400

23

 

 

cis-3-Hexenyl lactate (934)

Europe

270

38

0.6

+

 

USA

36

5

0.1

 

 

Butyl butyryllactate (935)

Europe

1900

280

5

 

USA

11 000

1400

24

 

 

Pyruvic acid (936)

Europe

250

35

0.6

3 900 000

16 000

USA

520

69

1.1

 

7 500

Pyruvaldehyde (937)

Europe

810

120

2

3100

38

USA

22

3

0.05

 

140

Ethyl pyruvate (938)

Europe

5

1

0.02

210

42

USA

150

20

0.3

 

1.4

Isoamyl pyruvate (939)

Europe

0

0

0

 

USA

0

0

0

 

 

3-Oxohexanoic acid glyceride (910)

Europe

0

0

0

 

USA

2000

270

4.5

 

 

3-Oxooctanoic acid glyceride (911)

Europe

240

34

0.6

 

USA

0

0

0

 

 

Heptanal glyceryl acetal (mixed 1,2 and 1,3 acetals) (912)

Europe

27

4

0.07

 

USA

0.05

0

0

 

 

3-Oxodecanoic acid glyceride (914)

Europe

0

0

0

 

USA

2000

270

4.5

 

 

3-Oxododecanoic acid glyceride (915)

Europe

510

73

1.2

 

USA

0

0

0

 

 

3-Oxotetradecanoic acid glyceride (916)

Europe

0

0

0

 

USA

2000

270

5

 

 

4-Methyl-2-pentyl-1,3-dioxolane (928)

Europe

0.1

0

0

 

USA

1.4

0.2

0.003

 

 

2,2,4-Trimethyl-1,3-oxyacylopentane (929)

Europe

2

0.3

0.005

2300

1200

USA

1.8

0.2

0.003

 

1300

NA, not applicable; NR, not reported; +, reported to occur naturally in foods (Maarse et al., 1999), but quantitative data were not available; –, not reported to occur naturally in foods

a

Intake expressed as µg/person per day calculated as follows: [(annual volume, kg) x (1 x 109 µg/kg)/ (population x survey correction factor x 365 days)], where population (10%, ‘eaters only’) = 32 x 106 for Europe and 26 x 106 for the USA. The correction factor = 0.6 for Europe and 0.8 for the USA, representing the assumption that only 60% and 80% of the annual volume of the flavour, respectively, was reported in the poundage surveys (International Organization of the Flavor Industry, 1995; Lucas et al., 1999). Intake expressed as µg/kg bw per day calculated as follows: [(µg/person per day)/body weight], where body weight = 60 kg. Slight variations may occur from rounding.

b

Quantitative data from Stofberg & Grundschober (1987)

c

Calculated as follows: (annual consumption in food, kg)/(most recently reported volume as a flavouring agent, kg)

1.3 Metabolic considerations

The aliphatic esters of propylene glycol, lactic acid and pyruvic acid and their parent compounds would all be expected to be readily absorbed from the gastrointestinal tract. Hydrolysis of the aliphatic esters is catalysed largely by hepatic esterases to give the component alcohol and carboxylic acid or aldehyde. After hydrolysis of glycerol esters in the intestine, glycerol is also readily absorbed. Glycerol, pyruvic acid, and lactic acid are endogenous in humans. Glycerol and pyruvic acid are metabolized completely and are not excreted. Lactic acid is also mainly metabolized, although urinary excretion may occur if the blood concentration is high. Propylene glycol can be metabolized, but high doses are likely to be excreted largely unchanged in the urine.

Glycerol is metabolized via the glycolytic pathway after it has been converted in the liver to glycerol-3-phosphate. Glycerol-3-phosphate is then oxidized to yield dihydroxyacetone phosphate, which is isomerized to glyceraldehyde-3-phosphate, eventually yielding pyruvic acid.

Pyruvic acid follows two primary routes of metabolism. Under aerobic conditions, it is converted to acetyl coenzyme A and enters the citric acid cycle. Under anaerobic conditions, primarily in muscles as a result of strenuous physical activity, pyruvic acid is reduced by lactic dehydrogenase to lactic acid.

Lactic acid diffuses through muscle tissue and is transported to the liver in the bloodstream. In the liver, it is converted to glucose by gluconeogenesis. Lactic acid can also be further catabolized in the lactic acid cycle (also known as the Cori cycle).

Propylene gycol can be oxidized to lactic acid via two biochemical pathways. If propylene glycol is phosphorylated, it can be converted to acetol phosphate, lactaldehyde phosphate, lactyl phosphate, and then lactic acid. If it is not phosphorylated, propylene glycol is successively oxidized to lactaldehyde, methylgloyoxal, and lactic acid.

1.4 Application of the Procedure for the Safety Evaluation of Flavouring Agents

Step 1 Twenty-eight of the 31 flavouring agents in this group are linear, simple branched aliphatic compounds. Twenty-two of these are in structural class I (Cramer et al., 1978) because they contain fewer than three different types of functional group (Nos 909, 913, 918–927, and 930–939). Six of these 28 substances are in structural class III because they contain three or more different types of functional group (Nos 910, 911, 914–917). The three remaining substances in the group are in structural class III because they are cyclic acetals and ketals (Nos 912, 928, and 929).

Step 2 The data on the metabolism of individual members of the group were sufficient to allow conclusions about their probable metabolic fate. The aliphatic esters of propylene glycol (Nos 926, 927, 928, and 929), lactic acid (Nos 931, 932, 934, and 935), and pyruvic acid (Nos 938 and 939) are expected to be hydrolysed to their component alcohol and carboxylic acid. Glycerol esters (Nos 910, 911, 912, 913, 914, 915, 916, 917, 918, 919, 920, 921, 922, 923, and 924) are expected to be hydrolysed to glycerol and carboxylic acids. Esters of propylene glycol are expected to be hydrolysed to propylene glycol and component acid. Esters of lactic acid and pyruvic acid are expected to be hydrolysed to lactic acid and pyruvic acid, respectively, and the corresponding alcohol. Acetals (Nos 927 and 933) are expected to be hydrolysed to their component alcohols and aldehydes, while ketals (Nos 928 and 929) are expected to be hydrolysed to their component ketones and alcohols. Glycerol (No. 909), lactic acid (No. 930), and pyruvic acid (No. 936) are endogeneous and are metabolized through the glycolytic and citric acid pathways. Propylene glycol (No. 925) is oxidized to lactic acid. For all substances in this group, therefore, the evaluation proceeded via the A side of the scheme.

Step A3 The daily per capita intakes of 22 of the substances in this group are below the threshold of concern for their respective structural classes (class I, 1800 µg; class III, 90 µg). These substances would not be expected to be of safety concern. The daily per capita intakes of the remaining nine substances (Nos 909, 910, 914, 916, 920, 925, 926, 930, and 931) exceed the threshold of concern for their respective structural classes. Evaluation of these substances therefore proceeds to step A4.

Step A4 Glycerol (No. 909), lactic acid (No. 930), and ethyl lactate (No. 931) are endogenous in humans and are therefore not expected to be a safety concern. Triacetin (No. 920), 3-oxohexanoic acid glyceride (No. 910), 3-oxodecanoic acid glyceride (No. 914), and 3-oxotetradecanoic acid glyceride (No. 916) are glycerol esters and are hydrolysed to glycerol. Propylene glycol (No. 925) and propylene glycol stearate (No. 926) are not endogenous in humans; however, the ester is expected to be hydrolysed to propylene glycol and stearic acid. Propylene glycol is known to be oxidized to lactic acid in mammals. These substances would therefore not be expected to be a safety concern.

1.5 Consideration of combined intakes

In the unlikely event that all 23 substances in structural class I were to be consumed concurrently on a daily basis, the estimated per capita consumption in Europe and the USA would exceed the human intake threshold for class I. The estimated per capita consumption in Europe and the USA for combined intake of the eight flavouring agents in structural class III would also exceed the human intake threshold for class III. Given that the substances are expected to be efficiently metabolized by known metabolic pathways, the Committee considered that the combined intake would not give rise to concerns about safety.

1.6 Conclusions

On the basis of the predicted metabolism, the Committee concluded that the 31 aliphatic acyclic diols, triols, and related substances in this group would not raise safety concerns at the current levels of intake when used as flavouring agents. In applying the Procedure, the Committee noted that all of the available data on toxicity are consistent with the results of the safety evaluation.

2. RELEVANT BACKGROUND INFORMATION

2.1 Explanation

This monograph summarizes the key data relevant to the evaluation of the 31 flavouring agents in this group. All members of this group are aliphatic acyclic primary alcohols, aldehydes, acids, or related esters with one or more additional oxygenated functional groups. The group consists of four subgroups: glycerol (No. 909) and 15 related glycerol esters and acetals (Nos 910, 911, 912, 913, 914, 915, 916, 917, 918, 919, 920, 921, 922, 923, and 924); propylene glycol (No. 925) and four related esters, acetals, and ketals (Nos 926, 927, 928, and 929); lactic acid (No. 930) and four lactate esters (Nos 931, 932, 934, and 935); and pyruvic acid (No. 936), its corresponding aldehyde (No. 937), two pyruvate esters (Nos 938 and 939) and one acetal of pyruvic acid (No. 933).

2.2 Additional considerations on intake

Quantitative data on natural occurrence and consumption ratios have been reported for seven flavouring agents in the group, which indicate that they are consumed predominantly from traditional foods (i.e., consumption ratio > 1) (Stofberg & Kirschman, 1985; Stofberg & Grundschober, 1987) (Table 2).

The daily per capita intake of this group of flavouring substances is 20 000 µg/day in Europe (330 µg/kg bw per day) and 2 500 000 µg/day in the USA (14 000 µg/kg bw per day). Glycerol accounted for approximately 85% (17 000 µg/day) of the total per capita intake in Europe; in the USA, propylene glycol accounted for 96% (2 400 000 µg/day) of the total per capita intake.

2.3 Biological data

2.3.1 Biochemical data

(a) Absorption and transformation

Little specific information was available on the absorption and transformation of individual members of this group of flavouring substances. The esters, acetals, and ketals of glycerol, lactic acid, and pyruvic acid would be expected to be readily absorbed, as would the parent compounds. After hydrolysis of glycerol esters in the intestine, glycerol is readily absorbed. Glycerol and pyruvic acid are metabolized completely and are not excreted. Lactic acid is also largely metabolized, although urinary excretion may occur if the blood concentration is high. Propylene glycol can be metabolized, but at high concentrations is likely to be largely excreted unchanged in the urine.

Propylene glycol given orally to three persons at a dose of 1038 mg (0.017 g/kg bw) was rapidly absorbed and eliminated in the urine and saliva (Hanzlik et al., 1939). In a study of the pharmacokinetics of propylene glycol in humans, multiple oral doses were rapidly absorbed, and its rate of clearance from blood was dose-dependent (Yu et al., 1985).

In studies of the minor pathways of metabolism of propylene glycol, administration to rats in drinking-water resulted in excretion unchanged in the urine (Van Winkle, 1941). Propylene glycol given orally to rabbits was conjugated with glucuronic acid and excreted in the urine (Miura, 1911; Fellows et al., 1947).

(b) Metabolism

(i) Hydrolysis of linear and branched-chain esters

In general, aliphatic esters of propylene glycol, lactic acid, and pyruvic acid are expected to be hydrolysed to their component alcohol and carboxylic acids. The hydrolysis is catalysed by classes of enzymes recognized as carboxylesterases or esterases (Heymann, 1980), the most important of which are the B-esterases, which, in mammals, predominate in hepatocytes (Heymann, 1980; Anders, 1989). The rates of hydrolysis follow first-order kinetics, with hydrolysis of the straight-chain esters occurring approximately 100 times more rapidly than that of branched-chain esters (Butterworth et al., 1975; Longland et al., 1977; Grundschober, 1977; Leegwater & van Straten, 1979).

Glycerol esters are hydrolysed to glycerol and the corresponding carboxylic acids (see Figure 1). The hydrolysis is catalysed by intestinal lipase (Tietz, 1986), which attacks the ester bonds at carbons 1 and 3. The ester bond at carbon 2 is more resistant to hydrolysis, possibly because of its stereochemistry and steric hindrance. The beta-monoglyceride can, however, spontaneously isomerise to the alpha-form (3-acylglycerol), permitting further hydrolysis to yield glycerol.

FIGURE 1

Figure 1. Hydrolysis of glycerol esters in humans

The rate of hydrolysis of glycerol esters depends on the surface area of the lipid–water interfaces, which increases greatly with the churning peristaltic movements of the intestine and the emulsifying action of bile acids. Lipase is rapidly denatured at these interfaces; however, colipase, a pancreatic protein that forms a 1:1 complex with lipase, inhibits the surface denaturation of lipase and anchors it to the lipid–water interface (Voet & Voet, 1990).

Studies of the hydrolysis of the glycerol fatty acid esters (tributyrin (No. 922) (Pilz, 1959; Pilz & Johann, 1967), glycerol 5-hydroxydecanoate (No. 923) (Als, 1975), and glycerol 5-hydroxydodecanoate (No. 924) (Als, 1975) showed complete hydrolysis to glycerol and the corresponding fatty acids, butyric acid, 5-hydroxy-decanoic acid, and 5-hydroxydodecanoic acid, respectively.

Hydroxylated and keto acids formed by hydrolysis of glycerol esters such as 5-hydroxydecanoic acid and 5-hydroxydodecanoic acid may form lactones by acid-catalysed intramolecular cyclization to yield five-member rings (see Figure 2). In aqueous media, equilibrium is established between the open-chain hydroxy-carboxylic acid and the lactone. At basic pH, the equilibrium favours the open-chain hydroxycarboxylate anion, but the lactone predominates at acidic pH. 5-Hydroxy-decanoic acid and 5-hydroxydodecanoic acid may form the delta-lactones delta-decalactone and delta-dodecalactone, respectively. Their metabolic fate can be predicted on the basis of an analogy with the known biotransformation of structurally related aliphatic lactones previously considered by the Committee (Annex 1, reference 132). Linear saturated 5-hydroxycarboxylic acids, which are formed from delta-lactones, are converted, via acetyl coenzyme A (CoA) to hydroxythioesters, which then undergo beta-oxidation and cleavage to yield an acetyl CoA fragment and a new beta-hydroxy-thioester reduced by two carbons. Even-numbered carbon acids continue to be oxidized and cleaved to yield acetyl CoA, while odd-numbered carbon acids yield acetyl CoA and propinyl CoA. Acetyl CoA enters the citric acid cycle directly, while propionyl CoA is transformed into succinyl CoA, which then enters the citric acid cycle.

FIGURE 2

Figure 2. Equilibrium between delta-lactones and their corresponding hydroxycarboxylic acids

Esters of propylene glycol are hydrolysed to propylene glycol and their component acid. In the presence of pancreatic lipase, propylene glycol stearate (No. 926) was hydrolysed to propylene glycol and stearic acid (Balls & Matlock, 1938).

Esters of lactic acid are hydrolysed to lactic acid and the corresponding alcohol. In rat plasma, ethyl lactate (No. 931) was hydrolysed to ethyl alcohol and lactic acid (Falke et al., 1981).

Esters of pyruvic acid are expected to be hydrolysed to pyruvic acid and the corresponding alcohol.

(ii) Hydrolysis of acetals and ketals

In general, acetals are hydrolysed to their component alcohols and aldehydes. Studies on the hydrolysis of 1,2,3-tris[(1´-ethoxy)ethoxy] propane (No. 913), which is readily hydrolysed to yield acetaldehyde and glycerol (DeSimone, 1976), support this conclusion.

Acetals of propylene glycol have also been shown to be hydrolysed to their component alcohol and aldehyde. In vitro, 1,2-di[(1´-ethoxy)ethoxy]propane (No. 927) was completely hydrolysed to acetaldehyde and propylene glycol (DeSimone, 1976). Potassium 2-(1´-ethoxy) ethoxypropanoate (No. 933), an acetal of lactic acid, was completely hydrolysed to lactic acid, acetaldehyde, and ethanol in simulated stomach fluids (Moreno et al., 1984). Aldehydes are oxidized to their corresponding carboxylic acids, which are subsequently metabolized through known biochemical pathways (Voet & Voet, 1990). For example, pyruvaldehyde (No. 937), the aldehyde of pyruvic acid, was metabolized to pyruvic acid when incubated with rat liver homogenate (Bonsignore et al., 1968).

Ketals are hydrolysed to their component ketones and alcohols. The related compound, benzaldehyde propylene glycol acetal, was hydrolysed in simulated gastric fluid and, to a lesser extent, in intestinal fluid (Morgareidge, 1962). Similarly, 2,2,4-trimethyl-1,3-oxacyclopentane (No. 929) would be expected to be hydrolysed in humans to yield acetone and propylene glycol.

(iii) Metabolism of glycerol (No. 909)

Glycerol is endogenous in the human body. It enters the glycolytic pathway after its conversion in the liver to glycerol-3-phosphate by glycerol kinase. Glycerol-3-phosphate is then oxidized by glycerol-3-phosphate dehydrogenase to yield dihydroxyacetone phosphate (see Figure 3), which is then isomerized to glyceral-dehyde-3-phosphate, eventually yielding pyruvic acid.

FIGURE 3

Figure 3. Metabolism of glycerol in humans

(iv) Metabolism of pyruvic acid (No. 936) and lactic acid (930)

Pyruvic acid is endogenous in the human body. It is a critical metabolic intermediate, and its fate depends on the oxidation state of the cell (see Figure 4). Under aerobic conditions, pyruvic acid is converted to acetyl CoA and enters the citric acid cycle, where it is completely metabolized. Under anaerobic conditions, lactate dehydrogenase catalyses the reduction of pyruvic acid to lactic acid and the oxidation of NADH to NAD+, primarily in muscles.

FIGURE 4

CoA, coenzyme A

Figure 4. Metabolism of pyruvic acid in humans

Lactic acid diffuses from the muscles and is transported through the bloodstream to oxygen-rich tissues such as the heart and liver, where it is catabolized further through the lactic acid cycle (also known as the Cori cycle) (see Figure 5), or converted to glucose via gluconeogenesis. Even in fully oxygenated muscle tissue, as much as 50% of the metabolized glucose is converted to lactic acid by way of pyruvic acid (Voet & Voet, 1990).

FIGURE 5

Figure 5. Lactic acid cycle (Cori cycle)

In resting women who received intravenous injections of [2-14C]pyruvate, analysis of blood glucose 1 h later showed 96% conversion of pyruvic acid to glucose (Hostetler et al., 1969). When [2-14C]pyruvate was incubated with liver slices from fasted normal rats, 86% had been used after 90 min of incubation. Of the radiolabel associated with metabolized pyruvic acid, 23% was associated with glycogen and glucose, 16% with CO2, and 16% with lactic acid. In the presence of glycerol, the use of pyruvic acid was increased to 95%, accompanied by a decrease in conversion to glycogen (16%) and CO2 (8.6%) and an increase in the production of lactic acid (Teng et al., 1953).

(v) Metabolism of propylene glycol (No. 925)

Propylene glycol can be oxidized to lactic acid via one of two pathways, depending on whether the glycol is phosphorylated (Rudney, 1954; Miller & Bazzano, 1965). In studies in vitro with rat liver, the free glycol was successively oxidized to lactaldehyde, methylglyoxal (pyuvaldehyde), and lactic acid (see pathway 1, Figure 6) (Ting et al., 1964; Miller & Bazzano, 1965), while the phosphorylated glycol followed the pathway of acetyl phosphate, lactaldehyde phosphate, lactyl phosphate, and lactic acid (Ruddick, 1972; see pathway 2, Figure 6). Lactate is subsequently converted to pyruvate, which enters the citric acid cycle and/or the gluconeogenesis pathway (Ruddick, 1972; Wittman & Bawin, 1974).

FIGURE 6

Figure 6. Metabolism of propylene glycol in mammals

2.3.2 Toxicological studies

(a) Acute toxicity

LD50 values after oral administration were available for 12 of the 31 substances in this group. In rats, the values ranged from 2000 to 31 000 mg/kg bw (Nos 909, 920, 922, 925, 930–933, 912, and 929), indicating little acute toxicity of this group by the oral route (Smyth et al., 1941; Fassett & Roudabush, 1952; Dominguez-Gil & Cadorniga, 1971; deGroot et al., 1974; Bailey, 1976; Bartsch et al., 1976; Moreno, 1976, 1977, 1978; Moreno et al., 1984; Clary et al., 1998). The available values for mice ranged from 1100 to 5000 mg/kg bw (Nos 922, 935, and 928) (Gast, 1963; Moran et al., 1980; Moreno, 1980).

(b) Short-term studies of toxicity

The results of short-term and long-term studies of toxicity conducted with substances in this group are summarized in Table 3.

Table 3. Results of short-term studies of toxicity with aliphatic acyclic diols, triols and related substances

Flavouring agent (No.)

Species, sex

No. of test groupsa/ no. per groupb

Route

Length (days)

NOEL (mg/kg bw per day)

Reference

Glycerol (909)

Rat, M,F

10/10

Diet

140

5000

Guerrant et al. (1947)

 

Mouse, M,F

1/81

Oral

365

No tumours at 5000 mg/kg bw

Witschi et al. (1989)

 

Human, M,F

1/14

Oral

50

No adverse effects reported at 24 000 mg/kg bw

Johnson et al. (1933)

 

Rat, M,F

3/18

Diet

350

20 000

Annex 1, reference 41

 

Rat, M,F

3/48

Diet

730

10 000

Annex 1, reference 41

 

Rat

3/22

Diet

730

10 000

Annex 1, reference 41

3-Oxooctanoic acid glyceride (911)

Rat, M,F

5/10

Diet

14

10

Gill & van Miller (1987)

3-Oxotetradecanoic acid glyceride (916)

Rat, M,F

5/10

Diet

14

10

Gill & van Miller (1987)

Tributyrin (922)

Rat,

NR1/66

Diet

245

Marked hyperplasia and papillomatous growth in fore stomach at 7500 mg/kg bw

Salmon & Copeland (1949)

Glycerol 5-hydroxydecanoatec (923)

Rat, M,F

2/8-17

Diet

343

150

Wilson (1961

Glycerol 5-hydroxydodecanoatec (924)

Rat, M,F

2/8-17

Diet

343

300

Wilson (1961)

Propylene glycol (925)

Rat, M,F

2/10

Diet

730

1800

Morris et al. (1942)

 

Rat, M,F

4/60

Diet

730

1300

Gaunt et al. (1972)

 

Dog, M,F

2/10

Diet

730

2000

Weil et al. (1971)

2,2,4-Trimethyl-1,3-oxacyclopentane (929)

Rat, M,F

2/5

Gavage

14

38

de Groot et al. (1974)

Lactic acid (930)

Rat, M,F

2/?

Diet

730

No increase in tumour incidence at 5000 mg/kg bw

Maekawa et al. (1991)

Pyruvaldehyde (937)

Rat, M

3/30
2/10

Oral

224

No increase in tumour incidence at 2500 mg/kg bw

Takahashi et al. (1989)

M, male; F, female; NR, not reported

a

Does not include control animals

b

Both male and female animals

c

delta-Decalactone and delta-dodecalactone are metabolites of glycerol 5-hydroxydecanoate and glycerol 5-hydroxydodecanoate, respectively.

Glycerol (No. 909)

Groups of five young rats of each sex were fed a diet containing glycerol at a concentration of 0, 1, 3, 6, 10, 15, 20, 30, 40, 50, or 60% (equivalent to 0, 1000, 3000, 6000, 10 000, 15 000, 20 000, 30 000, 40 000, 50 000, or 60 000 mg/kg bw per day) for 20 weeks. There was no significant difference in the body-weight gain at concentrations of glycerol ­ 30%, but reduced body-weight gain was observed at > 40%. Histological examination revealed no treatment-related changes at < 10%. The pathological changes observed at concentrations > 10% were marked hydropic and fatty degeneration of liver parenchymal cells. The NOEL was 5% glycerol in the diet, equivalent to 5000 mg/kg bw per day (Guerrant et al., 1947).

Ten men and four women were given glycerol orally at a dose calculated to result in an average daily intake of 24 000 mg/kg bw per day, for 50 days. No toxic effects were reported. The only effect was a slight tendency towards an increase in body weight (Johnson et al., 1933).

3-Oxooctanoic acid glyceride (No. 911) and 3-oxotetradecanoic acid glyceride (No. 916)

Groups of five male and five female Fischer 344 rats were given diets containing either 3-oxooctanoic acid or 3-oxotetradecanoic acid as esters of hydrogenated palm oil at a concentration calculated to provide a dose of 10 mg/kg bw per day, for 14 days. Detailed clinical examinations were conducted daily, and food consumption was measured on days 7 and 14. No physical signs of toxicity, abnormal body-weight gain, abnormal food consumption, or treatment-related effects were observed at necropsy. The absolute and relative weights of the liver and kidney were increased by 10% in female rats, but this effect was not considered to be biologically significant as no histological changes were found in these tissues and there were no other observed toxic effects (Gill & van Miller, 1987).

Glycerol 5-hydoxydecanoic acid (No. 923) and glycerol 5-hydroxydodecanoic acid (No. 924)

Studies were available on the lactones, delta-decalactone and delta-dodecalactone, which are formed from the hydrolysis of their respective glycerol esters glycerol 5-hydroxydecanoic acid and glycerol 5-hydroxydodecanoic acid. Groups of rats were fed a mixture of 30% delta-decalactone, 60% delta-dodecalactone, and 10% butyric acid at a concentration of 0.01% or 1% delta-decalactone or delta-dodecalactone in the diet for 49 weeks. These concentrations were calculated to result in average daily intakes of 1.5 or 150 mg/kg bw delta-decalactone and 3 or 300 mg/kg bw delta-dodecalactone. Histological examination revealed no adverse effects in any group. Haematology, blood chemistry, and urinary analysis showed no significant difference between test and control groups (Wilson, 1961).

2,2,4-Trimethyl-1,3-oxacyclopentane (No. 929)

In rats given 2,2,4-trimethyl-1,3-oxacyclopentane at a dose of 3.8 or 38 mg/kg bw per day for 14 days, there were no signs of toxicity at either dose (deGroot et al., 1974).

(c) Long-term studies of toxicity and carcinogenicity

Glycerol (No. 909)

In a study of the tumour promoting potential of glycerol, groups of male and female C3H mice, 6–8 weeks old, were given various carcinogens followed by 0, 0.5, or 1% (v/v) glycerol solution until they were 1 year old. Animals in the control group received either 5% (v/v) glycerol (equivalent to 5000 mg/kg bw per day) or water. The animals were killed, and the incidences of liver and lung tumours were recorded. Among males, the incidence of liver tumours was 23% in those given glycerol and 39% in those given water. The tumour incidence in the lung was 21% with glycerol abd 41% with water. Similar results were obtained for female mice. Thus, lower incidences of liver and lung tumours were seen after glycerol treatment. No treatment-related adverse effects were reported (Witschi et al., 1989).

A study in which Sprague-Dawley rats were given glycerol in the diet at a concentration of 0, 5, 10, or 20% (equivalent to 0, 5000, 10 000 or 20 000 mg/kg bw per day) for 50 weeks was evaluated previously by the Committee (Annex 1, reference 41). No significant treatment-related effects were found on growth rate or gross or histological appearance. The NOEL was 20 000 mg/kg bw per day (Atlas Chemical Co., 1969).

A study in which Sprague-Dawley rats were given glycerol in the diet at a concentration of 0, 5, 10, or 20% (equivalent to 0, 2500, 5000, or 10 000 mg/kg bw per day) for 2 years was evaluated previously by the Committee (Annex 1, reference 41). No significant treatment-related effects were found on growth rate or gross or histological appearance. Changes observed in relative kidney weights were not accompanied by histopathological changes. The NOEL was 10 000 mg/kg bw per day (Atlas Chemical Co., 1969).

A study in which Long-Evan rats were given glycerol in the diet at a concentration of 0, 5, 10, or 20% glycerol (equivalent to 0, 2500, 5000, or 10 000 mg/kg bw per day) for 2 years was evaluated previously by the Committee (Annex 1, reference 41). There were no significant treatment-related effects. The NOEL was 10 000 mg/kg bw per day (Hine et al., 1953).

Tributyrin (No. 922)

Groups of rats were fed a diet containing tributyrin, butyric acid, or ethyl butyrate to examine the occurrence of gastric lesions. Tributyrin was given at a concentration of 15 or 25% (equivalent to 7500 and 12 500 mg/kg bw per day) for 3–35 weeks. The animals has severely reduced body-weight gain, which represented approximately one-third of that of the control group. The 66 rats receiving tributyrin that were necropsied showed greatly enlarged stomachs with numerous irregular protuberances on the external surface. Microscopic examination revealed hyperplasia, hyperkeratosis, and occasional ulceration. The mucosa of the forestomach was covered in papillomas, resulting in a significant thickening of the forestomach wall (Salmon & Copeland, 1949).

Propylene glycol (No. 925)

A study in which rats were given propylene glycol in the diet at a concentration of 2.45% or 4.9% (equivalent to 900 and 1800 mg/kg bw per day) for 2 years was evaluated previously by the Committee (Annex 1, reference 33). No treatment-related adverse effects were found on growth, and histological examination revealed no treatment-related effects (Morris et al., 1942).

A study in which rats received propylene glycol in the diet at a concentration of 0, 310, 630, 1300, or 2500 mg/kg bw per day for 2 years was evaluated previously by the Committee (Annex 1, reference 33). No treatment-related adverse effects on body-weight gain, haematological, urinary, or clinical chemical end-points, or organ weights were found. The NOEL was 1300 mg/kg bw per day (Gaunt et al., 1972).

A study in which dogs received propylene glycol in the diet at a concentration of 0, 2000, or 5000 mg/kg bw per day for 2 years was evaluated previously by the Committee (Annex 1, reference 33). Increased erythrocyte destruction was found at the higher dose. No significant treatment-related effects on haematological, clinical chemical, or urinary end-points, or on gross or histological appearance were found (Weil et al., 1971).

Lactic acid (No. 930) and pyruvic acid (No. 936)

As pyruvic acid is reduced to lactic acid in vivo, data on lactic acid can be used to evaluate the safety of pyruvic acid.

Groups of male and female Fischer 344 rats were fed diets containing the calcium salt of lactic acid at a concentration of 0, 2.5, or 5% for 2 years, calculated to provide a dose of 0, 2500, or 5000 mg/kg bw per day, respectively. No adverse effects were observed, and no evidence was found of a significant dose-related increase in the incidence of tumours in any organ or tissue of treated animals. No specific dose-related changes were observed in any of the haematological and biochemical parameters measured (Maekawa et al., 1991).

Pyruvaldehyde (No. 937)

In a study to examine the tumour promoting potential of pyruvaldehyde in a two-stage model of stomach carcinogenesis, groups of male Wistar rats were given drinking-water containing 0.25% pyruvaldehyde for 32 weeks, alone or after 8 weeks’ treatment with a known tumour initiator, N-methyl-N-nitro-N-nitrosoguanidine (MNNG). Pyruvaldehyde alone caused no increase in the incidence of stomach hyperplasia or tumours. In rats pretreated with MNNG, pyruvaldehyde did not enhance the development of adenocarcinomas in the pylorus of the glandular stomach, but it significantly increased the incidence of hyperplasia (Takahashi et al., 1989).

(d) Genotoxicity

The results of studies of genotoxicity with these substances are shown in Table 4.

Table 4. Results of studies of the genotoxicity of aliphatic acyclic diols, triols and related substances

No.

Flavouring agent

End-point

Test system

Concentration

Results

Reference

In vitro

909

Glycerol

Reverse mutation

S. typhimurium

11 780 ppm

Negative

Cortruvo et at. (1977)

 

 

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537, TA1538

1000 µg/plate

Negativeb

Doolittle et al. (1988)

 

 

Reverse mutation

S. typhimurium TA97, TA102

0–10 000 µg/plate

Negativec

Fujita et al. (1994)

 

 

Reverse mutation

S. typhimurium TA98, TA100

0.05–1000 µg/plate

Negatived

Haresaku et al. (1985)

 

 

Reverse mutation

S. typhimurium TA98, TA100, TA1537, TA1538

10 000 µg/plate

Negativeb

Haworth et al. (1983)

 

 

Reverse mutation

S. typhimurium TA92, TA94, TA98, TA100, TA1535, TA1537

50 000 µg/plate

Negativeb

Ishidate et al. (1984)

 

 

Reverse mutation

S. typhimurium TA98, TA100. TA1535, TA1537, TA1538

1000 µg/plate

Negativeb

Lee et al. (1988)

 

 

Reverse mutation

S. typhimurium

NR

Negative

McCann & Ames (1976)

 

 

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537, TA1538

5-5000 µg/plate

Negativeb

Shimizu et al. (1985)

 

 

Reverse mutation

S. typhimurium TA100

1000 µmol/plate

Negative

Stolzenberg & Hine (1979)

 

 

Reverse mutation

S. typhimurium TA100

500 µg/ml

Negativeb

Yamaguchi (1982)

 

 

Reverse mutation

E. coli WP2uvrA

5-5000 µg/plate

Negativeb

Shimizu et al. (1985)

 

 

Reverse mutation

Saccharomyces cerevisiae D3

12 000 ppm

Negative

Cortruvo et at. (1977)

 

 

Modified reverse mutation

E. coli Sd-4-73

12–31 µg/plate

Negative

Szybalski (1958)

 

 

Gene mutation

Chinese hamster ovary cells K1-BH4, hprt locus

0–1000 µg/ml

Positiveb

Doolittle et al. (1988)

 

 

Sister chromatid exchange

Chinese hamster ovary cell line

100-1000 µg/ml

Negativeb

Doolittle et al. (1988)

 

 

Sister chromatid exchange

Chinese hamster ovary cells

1000 µg/ml

Negativeb

Lee et al. (1988)

 

 

Chromosomal aberration

Chinese hamster ovary cell line

1000 µg/ml

Negative

Doolittle et al. (1988)

 

 

Chromosomal aberration

Chinese hamster lung fibroblasts

1000 µg/ml

Negative

Ishidate et al. (1984)

 

 

Chromosomal aberration

Chinese hamster ovary cells

1000 µg/ml

Negativeb

Lee et al. (1988)

 

 

Mutation

Chinese hamster ovary cells

1000 µg/ml

Negativeb

Lee et al. (1988)

 

 

Unscheduled DNA synthesis

Rat hepatocytes

1000 µg/ml

Negative

Doolittle et al. (1988)

 

 

Unscheduled DNA synthesis

Rat hepatocytes

10–100 000 µg/ml

Negative

Fautz et al. (1991)

 

 

Unscheduled DNA synthesis

Rat hepatocytes

1000 µg/ml

Negative

Lee et al. (1988)

918

Glycerol mono-stearate

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537, TA1538

50 µg/plate

Negativeb

Blevins & Taylor (1982)

 

 

Reverse mutation

S. typhimurium TA97, TA98, TA100

0–400 µg/plate

Positiveb

Kuroda et al. (1985)

925

Propylene glycol

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537, TA1538

1–10 000 µg/plate

Negative

Clark et al. (1979)

 

 

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537

230 µg/plate

Negativeb

Florin et al. (1980)

 

 

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537

100–10 000 µg/plate

Negativeb

Haworth et al. (1983)

 

 

Reverse mutation

S. typhimurium TA92, TA94, TA98, TA100, TA1535, TA1537

10 000 µg/plate

Negativeb

Ishidate et al. (1984)

 

 

Reverse mutation

S. typhimurium TA98, TA100

NR

Negativeb

Kawachi et al. (1981)

 

 

Reverse mutation

S. typhimurium

NR

Negative

McCann & Ames (1976)

 

 

Reverse mutation

S. typhimurium TA100

1000 µmol/plate

Negative

Stolzenberg & Hine (1979)

 

 

Host-mediated mutation

S. typhimurium TA1530 and G46

0.01–0.25 ml

Negative

Weir (1974)

 

 

Host-mediated mutation

Saccharomyces cerevisiae

0.01–0.25 ml

Positive

Weir (1974)

 

 

Mutation

Bacillus subtilis rec

NR

Negatived

Kawachi et al. (1981)

 

 

Chromosomal aberration

Human embryonic lung cells

0.001-0.1 µg/ml

Negative

Weir (1974)

 

 

Chromosomal aberration

Hamster lung fibroblasts

32 000 µg/ml

Positived

Ishidate et al. (1984)

 

 

Chromosomal aberration

Hamster lung fibroblasts

NR

Positived

Kawachi et al. (1981)

 

 

Micronucleus formation

Chinese hamster ovary Don-6 cell line

3800–23 000 µg/ml

Negative

Sasaki et al. (1980)

 

 

Micronucleus formation

Human fibroblastic cell line HE2144

3800–23 000 µg/ml

Negative

Sasaki et al. (1980)

 

 

Sister chromatid exchange

Hamster lung fibroblasts

NR

Negatived

Kawachi et al. (1981)

 

 

Sister chromatid exchange

Hamster lung fibroblasts

NR

Negative

Kawachi et al. (1981)

 

 

Sister chromatid exchange

Chinese hamster ovary Don-6 cell line

3800–23 000 µg/ml

Positive

Sasaki et al. (1980)

 

 

Sister chromatid exchange

Human fibroblastic cell line HE2144

7600 µg/ml

Negative

Sasaki et al. (1980

 

 

Mutation

Mice

2700 mg/kg bw

Negative

Solt & Neale (1980)

 

 

Chromosomal aberration

Mice

0.6–24 mg/kg bw

Negative

Vargova et al. (1980)

 

 

Chromosomal aberration

Rat

30–5000 mg/kg bw

Negative

Weir (1974)

 

 

Micronucleus formation

Mice

0–20 000 mg/kg bw

Negative

Hayashi et al. (1988)

930

Lactic acid

Reverse mutation

S. typhimurium TA97, TA98, TA100, TA104

0.5–2.0 µl/plate

Negativeb

Al-Ani & Al-Lami (1988)

 

 

Chromosomal aberration

Chinese hamster ovary K1 cells

900–1400 µg/ml

Positiveb

Morita et al. (1990)

931

Ethyl lactate

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537, TA1538

NR

Negativeb

Clary et al. (1998)

935

Butyl butyryl lactate

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537, TA1538

0–3600 µg/plate

Negativeb

Wild et al. (1983)

 

 

Micronucleus formation

Mice

1500 mg/kg bw

Negative

Wild et al. (1983)

 

 

Sex-linked recessive lethal mutation

Drosophila melanogaster

5400 µg/ml

Negative

Wild et al. (1983)

936

Pyruvic acid

Reverse mutation

S. typhimurium TA100

200 µg/plate

Negativeb

Yamaguchi (1982)

 

 

Reverse mutation

S. typhimurium TA98, TA100

10–10 000 µg/plate

Negativeb

Bjeldanes & Chew (1979)

937

Pyruvaldehyde

Reverse mutation

S. typhimurium TA98

30 µg/plate

Positiveb

Yamaguchi (1982)

 

 

Reverse mutation

S. typhimurium TA98, TA100, TA104

NR

Positived

Kato et al. (1989)

 

 

Reverse mutation

S. typhimurium

76 µg/plate

Positive

Kim et al. (1987)

937

Pyruvaldehyde

Reverse mutation

S. typhimurium TA100, TA102, TA104

5–500 µg/plate

Positiveb

Shane et al. (1988)

 

 

Reverse mutation

S. typhimurium TA102, TA104

0–72 µg/plate

Positiveb

Migliore et al. (1990)

 

 

Reverse mutation

S. typhimurium TA98, TA100, TA102

0.29-29 µg/plate

Positiveb

Aeschbacher et al. (1989)

 

 

Reverse mutation

S. typhimurium TA100

1000 µg/plate

Positived

Nagao et al. (1986)

 

 

Reverse mutation

S. typhimurium TA100

220-500 µg/plate

Positived

Dorado et al. (1992)

 

 

Reverse mutation

E. coli WP2uvrA/pKM101

NR

Positived

Kato et al. (1989)

 

 

Chromosomal aberration

Chinese hamster ovary cells

10–50 µg/ml

Positive

Nishi et al. (1989)

 

 

Chromosomal aberration

Human lymphocytes

320 µg/ml

Positiveb

Migliore et al. (1990)

 

 

Sister chromatid exchange

Chinese hamster ovary cells

7.2 µg/ml

Negative

Tucker et al. (1989)

 

 

Sister chromatid exchange

Chinese hamster ovary cells

14–36 µg/ml

Positive

Tucker et al. (1989)

 

 

Sister chromatid exchange

Chinese hamster ovary cells

7.2–54 µg/ml

Positive

Faggin et al. (1985)

 

 

Sister chromatid exchange

Human lymphocytes

320 µg/ml

Positiveb

Migliore et al. (1990)

 

 

Micronucleus formation

Human lymphocytes

320 µg/ml

Positiveb

Migliore et al. (1990)

 

 

DNA strand breaks

Calf thymus DNA

NR

Positive

Rahman et al. (1990)

 

 

Micronucleus formation

Rat

400 mg/kg bw

Negative

Martelli et al. (1989)

 

 

Micronucleus formation

Rat

800 mg/kg bw

Weakly positive

Martelli et al. (1989)

 

 

Sister chromatid exchange

Mice duodenal cells

400 mg/kg bw

Negative

Migliore et al. (1990)

 

 

Sister chromatid exchange

Mice duodenal cells

600 mg/kg bw

Weakly positive

Migliore et al. (1990)

 

 

Unscheduled DNA synthesis

Rat pyloric mucosa cells

50 mg/kg bw

Negative

Furihata et al. (1985)

 

 

Unscheduled DNA synthesis

Rat pyloric mucosa cells

200–600 mg/kg bw

Positive

Furihata et al. (1985)

 

 

Chromosomal aberration

Mice duodenal cells

400–600 mg/kg bw

Negative

Migliore et al. (1990)

938

Ethyl pyruvate

Reverse mutation

S. typhimurium TA98, TA100, TA1535, TA1537

32–20 000 µg/plate

Negativeb

Anderson & Jensen (1984)

a With and without ozonation

b With and without metabolic activation

c With metabolic activation

d Without metabolic activation

e Administered in drinking-water

f Administered by gavage

The only consistently positive results both in vitro and in vivo were found with pyruvaldehyde (No. 937). This substance caused reverse mutation in bacteria and chromosomal aberrations in Chinese hamster ovary and human cells, sister chromatid exchange in Chinese hamster ovary cells, micronuclei in human lymphocytes and in rat cells, and unscheduled DNA synthesis in rat cells. Pyruvaldehyde is a natural component of some foods and is readily oxidized to the endogenous substance, pyruvate, in vivo. The estimated intake from its use as a flavouring agent is well below the estimated intake from natural sources.

(e) Reproductive toxicity

Glycerol (No. 909)

A seven-generation study of reproductive toxicity in rats given glycerol at a concentration of 0 or 30% (equivalent to 15 000 mg/kg bw per day) was evaluated previously by the Committee (Annex 1, reference 41). On average, the pups of treated dams weighed 20% less than those of the control group (Guerrant et al., 1947).

Propylene glycol (No. 925)

In a study to examine the potential of di(2-ethylhexyl) phthalate and its metabolites to cause testicular damage in rats after oral administration, a control group of six male Sprague Dawley rats were given propylene glycol orally at a dose of 2000 mg/kg bw per day for 5 days. On day 6, the animals were killed, and the testis, the ventral lobes of the prostate, and the liver were removed and studied. The testes of animals given propylene glycol were reported to contain occasional degenerated cells most of which were in early meiotic prophase or undergoing meiotic division (Sjoberg et al., 1986).

The effects of 15 chemicals, including propylene glycol, on differential ovarian follicle counts and reproductive performance were compared. Propylene glycol had no effect on reproductive function (Bolon et al., 1997).

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ENDNOTES

1 When evaluating these flavouring agents, the Committee questioned whether some substances in this group (see footnote to Table 1) were used as flavouring agents and therefore appropriate to be evaluated using this Procedure. Information to address this question will be sought from relevant manufacturers.

2 ADI ‘not specified’ is used to refer to a food substance of very low toxicity, which, on the basis of the available data (chemical, biochemical, toxicological, and other) and the total dietary intake of the substance arising from its use at the levels necessary to achieve the desired effect and from its acceptable background levels in food, does not, in the opinion of the Committee, represent a hazard to health. For that reason, and for reasons stated in individual evaluations, the establishment of an ADI expressed in numerical form is not deemed necessary. An additive meeting this criterion must be used within the bounds of good manufacturing practice, i.e., it should be technologically efficacious and should be used at the lowest level necessary to achieve this effect, it should not conceal food of inferior quality or adulterated food, and it should not create a nutritional imbalance.



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       Toxicological Abbreviations