Toxicological evaluation of some food
additives including anticaking agents,
antimicrobials, antioxidants, emulsifiers
and thickening agents
WHO FOOD ADDITIVES SERIES NO. 5
The evaluations contained in this publication
were prepared by the Joint FAO/WHO Expert
Committee on Food Additives which met in Geneva,
25 June - 4 July 19731
World Health Organization
Geneva
1974
1 Seventeenth Report of the Joint FAO/WHO Expert Committee on
Food Additives, Wld Hlth Org. techn. Rep. Ser., 1974, No. 539;
FAO Nutrition Meetings Report Series, 1974, No. 53.
PROPIONIC ACID AND ITS CALCIUM, POTASSIUM AND SODIUM SALTS
Explanation
These compounds have been evaluated for acceptable daily intake
by the Joint FAO/WHO Expert Committee on Food Additives (see Annex I,
Refs No. 6 and No. 13) in 1961 and 1965.
Since the previous evaluation, additional data have become
available and are summarized and discussed in the following monograph.
The previously published monograph has been expanded and is reproduced
in its entirety below.
BIOLOGICAL DATA
BIOCHEMICAL ASPECTS
At concentrations up to 0.004 mol/l the acid is fungistatic (Peck
& Rosenfeld, 1938). Sodium and calcium propionate inhibit moulds and
fungi at specific concentrations varying from 0.0125% to 1.25% at pH
5.5 and sodium propionate at 1.6% to 6% inhibits various bacteria
(Keeney & Boyles, 1943). In tests with a number of microorganisms it
has been shown that the bacteriostatic and fungistatic activity of
sodium propionate was greater in acid than in neutral or slightly
alkaline solution, which suggests that the antimicrobial action is due
to the undissociated acid (Heseltine, 1952b and Preservatives Report
1959).
Propionic acid is not a component of the edible fats and oils,
but arises in the intermediary metabolism of the body as the terminal
three-carbon fragment in form of propionyl coenzyme A in the oxidation
of odd-number carbon fatty acids. Oxidation of the side-chain of
cholesterol by rat liver mitochondria yields propionate as the
immediate product of cleavage (Mitropoulos & Myant, 1965). Propionates
are metabolized and utilized in the same way as normal fatty acids and
even after large doses no significant amounts of propionic are
excreted in the urine (Bässler, 1957). In vitro propionic acid is
completely oxidized by liver preparations to CO2 and water
(Huennekens et al., 1951).
The metabolic fate of propionates varies in microorganisms. Some
have enzyme systems converting succinate to propionyl-coenzyme A and
by various further steps to propionate, CO2 or propionyl phosphate.
Others convert propionic acid to B-alanine or directly to CO2. The
inhibiting effect for microbials is probably related to competition
with acetate in the acetokinase systems, to blockage of pyruvate
conversion to acetyl-coenzyme A and to interference with B-alanine in
pantothenic acid syntheses (Bässler, 1959). In mammals observations
have shown easy absorption from the gastrointestinal tract (Dawson et
al., 1964) and absence of any excretion in the urine whatever the mode
of administration. Decomposition by bacteria in the gut also occurs
(Hermann et al., 1938). In the mouse 14C-tagged propionate acts as
a precursor of body fat (Kleiber et al., 1953) and is incorporated
into the odd-numbered fatty acids of milk fat (James et al., 1956).
1-14C-labelled propionate fed to fasted rats produced 50% of the
activity as expired CO2 within two hours, the remainder being
incorporated into glucose, glycogen, succinate, malate, fumarate and
proteins (Buchanan et al., 1943; Lorber et al., 1950; Pritchard &
Tove, 1960). Rabbits oxidize propionate without keto-intermediates
(Mackay et al., 1940a; Mackay et al., 1940b). Dairy cows, when
injected i.v. with propionic acid labelled with 1-C14, produce active
lactose, casein and butter fat and convert 2-C14 acid to asparaginic
acid and serine (Assoc. Food & Drug Officials). Propionate increases
glucose output in phlorhidzinised animals (Rittenberg et al., 1937),
and has a glycogenic effect in fasting rats without being involved
in cholesterol synthesis (Buchanan et al., 1943). The in vitro
antiketogenic action of propionate was not confirmed in vivo.
Nine rabbits, made diabetic with alloxan, were given orally 1000 and
3000 mg/kg bw sodium propionate daily without significant useful
effect on their acetonuria and 30% of the animals died at the higher
dosage level. 740 mg/kg bw oral sodium propionate did not alter the
quantitative urinary excretion of volatile carboxylic acids but a
higher proportion of acetic acid appeared in normal rabbits' urine and
a proportionate rise in diabetic animals. Sugar excretion beyond that
of the diabetic level was noted (Maurer & Lang, 1956). Studies on
liver and kidney mitochondrial extracts or intact mitochondria and
homogenates of cardiac or skeletal muscle tissue showed conversion of
propionate to CO2 and water. The pathways involve combination with
coenzyme A in the same way as acetic acid, becoming carboxylated to
methylmalonyl-coenzyme A and undergoing a quantitative conversion to
succinate which then enters the Krebs citric acid cycle (Beck et al.,
1957; Flavin & Ochoa, 1957). Intact rat liver mitochondria fix
14C-labelled bicarbonate to propionate with conversion to succinate
in the presence of a cosubstrate while guinea-pig or beef liver
mitochondria and pig heart enzymes act without cosubstrate (Friedberg
et al., 1956). In vitro 1 g liver (rat) oxidize 3 mg/hom (Friedberg
et al., 1956), 1 g heart muscle (dog) 0.23 mg/hom (Cavert & Johnson,
1956). Skeletal muscle (dog) metabolizes 1-14C-labelled propionate,
which contributes 9% to 13% of the CO2 produced (Lifson, 1957).
Rabbit skeletal muscle extract markedly stimulates by its
fluorophosphate-forming enzyme system carboxylative activity of pig
heart enzymes (Flavin et al., 1957). Other pathways use direct
oxidation to pyruvate or direct condensation with acetate to produce
fatty acids with odd numbers of C-atoms in mouse adipose tissue and
ruminants (Bässler, 1959). Oxidation of 1-14C-labelled propionate to
CO2 proceeded in the same way and at the same rate in mouse adipose
tissue as in liver, while 2-14C propionate was incorporated
preferentially into newly synthesized fatty acids at much increased
rate in mouse adipose tissue compared with liver. The minor direct
oxidation of 2-14C propionate proceeded at equal rates in adipose and
liver tissue. Direct condensation of propionate and CO2 to succinate
or 4-carbon intermediates did not occur (Feller & Feist, 1957, Wakil,
1962). Conversion of propionic acid B-alanine was shown (Rendina &
Coon, 1957). High propionate concentrations inhibit acetate and other
metabolism in tissue cultures, homogenates or mitochondrial
preparations by competition for coenzymes A, e.g. formation of
ketobodies from pyruvate and acetate, activation and oxidation of
fatty acids, acetate metabolism and acetoacetate formation are
effected (Lang & Bässler, 1953). The production of 14CO2 from
labelled acetate by rat liver homogenates was almost abolished by
propionate in only one-tenth the concentration of acetate (Pennington
& Appleton, 1958). Inhibition of catalase activity has been reported
(Lück, 1957). In man and in animals these enzyme-inhibiting activities
are of little significance in vivo, as the fast rate of metabolism
prevents accumulation of the high concentrations that would be
necessary. In human plasma propionic acid represents 0% to 4% of the
total fatty acid and is a by-product of normal intermediate
metabolism. Absorbed propionate is removed by the liver, kidneys,
heart, muscle and adipose tissue. The liver can deal with 4.5 g free
acid or 5.8 g sodium propionate per hour, the isolated dog's heart
metabolizes 0.24 mol/hour per 100 g tissue and isolated dog's
gastrocnemius metabolizes 1-14C-labelled propionate such that 9% to
13% of CO2 produced is being contributed by the labelled substrate.
The normal breakdown of various amino acids yields propionate of
propionyl-coenzyme A and fatty acids with odd numbers of C-atoms yield
also propionate (Bässler, 1959). Propionate occurs in human sweat
through glycogenolysis in the sweat glands and from excess in tissue
fluids after the needs of fatty acid metabolism have been met
(Heseltine, 1952a).
TOXICOLOGICAL STUDIES
Acute toxicity
No LD50 estimations are published in the literature because of
the very low acute toxicity of propionates (Heseltine, 1952a;
Sollmann, 1957). The CNS effect is a direct action of the propionate
on nervous tissue (Samson et al., 1956).
Other
ED50* LD50 dosages
Animal Route mg/kg bw Effects References
Frog i.m. - - 100 Twitching, Fodera, 1894
paresis for
24 hours
Rat i.p. 2 156 - - Unconscious Samson et al.,
(sod,salt) in 18-20 1956
minutes
i.v. 1 330 - - Unconscious Samson et al.,
(sod.salt) in 5 seconds 1956
s.c. - - 2 800 Tiredness Samson et al.,
(sod.salt) 1956
oral - - 5 600 Nil Samson et al.,
(sod.salt) 1956
(free acid) - 2 600 - - U.S. Food & Drug
Cat s.c. - - 1 000 Sleep Mayer, 1886
(sod.salt)
Rabbit i.v. - - 1 320 Death Hermann, 1930a
(free acid)
(sod.salt) - - 2 200 Dyspnoea, Mayer, 1886
narcotic
paresis &
urination
(24 hours)
Dog s.c. - - 900 Nil Knoop & Jost,
(free acid) 1923
i.v. - - 500 Dyspnoea, Mayer, 1886
(free acid) narcotic
weakness
- - 370 Temporary Hermann, 1930a
BP fall
(sod.salt) - - 570 Vomiting, Knoop & Jost,
weakness 1923
* ED50 - dose producing temporary unconsciousness in 50% of animals.
Short-term studies
Rat
Four groups of one control and two weanling test rats were pair-
fed for four to five weeks on diets containing 1% sodium or calcium
propionate and 3% sodium or calcium propionate. No effect on growth
was observed (Harshbarger, 1942). In another experiment 14 groups of
15 male animals were each fed on diets containing 0.075% or 3.75%
sodium propionate in combination with varying proportions of other
additives at 50 times their commercial level of usage. Transient
growth depression was noted with 3.75% sodium propionate which later
became normal. Final weights of the 3.75% groups were significantly
depressed. Food consumption was reduced in all diets compared with
controls and feed efficiency was very poor with 3.75% propionate.
Haematological findings were normal at 16 weeks. Histopathology of
cerebellum, cerebrum, heart, trachea, oesophagus, thyroid, salivary
gland, thymus, adrenals, pancreas, bladder, testes, kidneys, lymph-
glands, lung, stomach, liver, spleen, small gut showed kidney
abnormalities in diets containing chlorine dioxide-treated flour, but
no consistent change related to propionate. Mortality was not
adversely affected in any group and weights of liver, left kidney,
heart and spleen were comparable with controls (Graham & Grice, 1955).
Over a period of 39 days 5% sodium propionate in a rat free from
biotin, relic acid or vitamin B12 reduced growth rate and food
intake. The addition of biotin, relic acid or especially vitamin B12
inhibited this effect. Diets containing 3.0 and 6.0% sodium propionate
also produced a growth retardation which was overcome by the addition
of vitamin B12 (25 µg/kg diet), more effectively with the 3.0%
propionate diet (Hogue & Elliot, 1964).
Rabbit
Normal and alloxan-diabetic animals were fed daily on 1000 mg/kg
bw of sodium propionate. Normal animals showed no adverse effects but
a small amount of acetic acid appeared in their urine. Diabetic
animals excreted unchanged amounts of ketone bodies but urinary
concentrations of volatile fatty acids and glucose increased. No
propionate appeared in tho urine of either normal or diabetic animals
(Maurer & Lang, 1956).
Long-term studies
Rat
Groups of 13 male and 13 female weanling rats were fed on diets
containing 0.075% and 3.75% sodium propionate in a baked bread in
combination with various proportions of other additives at 50 times
their usual commercial level for one year. No effect on growth,
mortality rate or body weight was seen although food consumption was
slightly depressed on all diets compared with controls. No other
toxicological effects were found as judged by histopathology of
bladder, small gut, spleen, stomach, pancreas, adrenal, kidney, liver,
heart, lung, thyroid, brain or gonads or weights of heart, liver,
spleen and kidneys. The kidneys of female rats on diets containing
high levels of chlorine-dioxide-treated flour showed tubular
degeneration and minor glomerulonephritic changes (Graham et al.,
1954).
OBSERVATIONS IN MAN
In an adult male daily oral doses of 6000 mg of sodium propionate
rendered the urine faintly alkaline but had no other effect (Bässler,
1959).
Solutions of propionate applied to the eye in concentrations up
to 15% in man and up to 20% in rabbits had no irritating effect
(Theodore, 1950). Propionic acid is a moderate irritant of skin
causing stinging pain and subsequent hyperpigmentation (Oettel, 1936).
No sensitization from topical use has been reported, nor has it any
anticoagulant effect (Heseltine, 1952a).
Two male and two female volunteers were treated locally with 0.05
or 0.1% histamine phosphate and inhibition of the reaction by 7.5% or
15% sodium propionate was measured. A moderate potency of about 1/7.5
of that diphenhydramine was found (Heseltine, 1952a).
Comments;
There are no toxicological studies of longer duration than one
year. However, propionate is a normal intermediary metabolite, and a
normal constituent of foods.
An evaluation may be made based on the metabolic information.
EVALUATION
Estimate of acceptable daily intake for man
Not limited*
* See relevant paragraph in the seventeenth report (pages 10-11).
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See Also:
Toxicological Abbreviations