FAO Nutrition Meetings
Report Series No. 40A,B,C
TOXICOLOGICAL EVALUATION OF SOME
ANTIMICROBIALS, ANTIOXIDANTS, EMULSIFIERS,
STABILIZERS, FLOUR-TREATMENT AGENTS, ACIDS AND BASES
The content of this document is the result of the deliberations of the
Joint FAO/WHO Expert Committee on Food Additives which met at Rome,
13-20 December, 19651 Geneva, 11-18 October, 19662
1 Ninth Report of the Joint FAO/WHO Expert Committee on Food
Additives, FAO Nutrition Meetings Report Series, 1966 No. 40;
Wld Hlth Org. techn. Rep. Ser., 1966, 339
2 Tenth Report of the Joint FAO/WHO Expert Committee on Food
Additives, FAO Nutrition Meetings Report Series, 1967, in press;
Food and Agriculture Organization of the United Nations
World Health Organization
SULFUR DIOXIDE AND RELATED SUBSTANCES
Chemical name Sulfur dioxide; Sulfurous acid anhydride
Chemical formula SO2
Molecular weight 64.07
Definition Sulfur dioxide contains not less than 95
per cent. SO2.
Description Sulfur dioxide is a colourless,
nonflammable gas, with a strong,
pungent, suffocating odour.
(See Sodium Sulfite)
Chemical names Potassium metabisulfite; Potassium
pyrosulfite; Potassium disulfite
Empirical formula K2S2O5
Molecular weight 222.33
Definition Potassium metabisulfite contains not
less than 90 per cent. of K2S2O5.
Description Potassium metabisulfite occurs as
colourless free-flowing crystals,
crystalline powder, or granules, usually
having an odour of sulfur dioxide
(See Sodium Sulfite)
SODIUM HYDROGEN SULFITE
Synonyms Sodium bisulfite; Sodium acid sulfite
Chemical name Sodium hydrogen sulfite
Chemical formula NaHSO3
Molecular weight 104.06
Definition Sodium hydrogen sulfite contains not
less than 95 per cent. of NaHSO3 and
not less than 60 per cent. SO2.
Description Sodium hydrogen sulfite is a white
crystalline or granular solid with an
odour of sulfur dioxide.
(See Sodium Sulfite)
Chemical name Sodium pyrosulfite
Chemical formula Na2S2O5
Molecular weight 190.11
Definition Sodium metabisulfite contains not less
than 95 per cent. of Na2S2O5 and not
less than 64 per cent. SO2.
Description Sodium metabisulfite is a white
crystalline solid with an odour of
(See Sodium Sulfite)
Chemical name Sodium sulfite
Chemical formula (a) Na2SO3 (anhydrous)
Definition (a) Sodium sulfite (anhydrous) contains
not less than 95 per cent. Na2SO3 and
not less than 48 per cent. SO2.
(b) Sodium sulfite heptahydrate contains
not less than 48 per cent. Na2SO3 and
not less than 24 per cent. SO2.
Description a) Sodium sulfite (anhydrous) is a white
powder with not more than a faint odour
of sulfur dioxide.
(b) Sodium sulfite heptahydrate is a
transparent or white crystalline solid
with not more than a faint odour of
The biological data as they were available at the end of 1964
have been summarized in the document entitled "Specifications for
Identity and Purity and Toxicological Evaluation of Some
Antimicrobials and Antioxidants" (FAO/WHO, 1965). Since its
publication some new experimental work has been carried out on this
substance. This and other work not included in the above document are
summarized in this monograph.
Four rats given oral doses of sodium metabisulfite as a 0.2 per
cent. solution eliminated 55 per cent. of the sulfur as sulfate in the
urine within the first 4 hours (Bhagat & Lockett, 1960). A rapid and
quantitative elimination of sulfites as sulfate was also observed in
man and dog (Rost, 1933).
Sulfite is a strong inhibitor of some dehydrogenases, e.g.
lactate dehydrogenase (heart) and malate dehydrogenase; 50 per cent.
inhibition by about 10-5 M sulfite (Pfleiderer et al., 1956).
Small amounts of sulfite are regularly formed in the intermediary
metabolism of the body in the catabolism of cystine by the
non-enzymatic decomposition of 8-sulfinyl pyruvic acid to pyruvic acid
and SO2. The stationary concentration of sulfite in the cells is too
small to be measured. However, 0.10-0.12 meq./100 ml was found in bull
seminal fluid (Larson & Salisbury, 1953).
Exposure of cells in tissue culture to various concentrations of
SO2 in the medium showed that strain L cells could tolerate 5 ppm
SO2 for periods of 8 hours provided a recovery period followed each
exposure. At higher concentrations (500-2000 ppm) of SO2 there was
inhibition of growth; at the 500 ppm level the growth was comparable
to control cultures. The addition of salts of SO2 caused stimulation
of growth at lower levels and complete inhibition at 2000 ppm NaHSO3
(Thompson & Pace, 1962).
Effect on thiamine
Treatment of foods with sulfites reduced their thiamine content
(Morgan et al., 1935; Williams et al,, 1935), It has been suggested
that the ingestion of SO2 in a beverage may effectively reduce the
level of thiamine in the rest of the diet (Hötzel, 1962).
Six rats ware given a diet providing 40 mg thiamine daily. At
weekly intervals an additional 160 mg thiamine was given and the
urinary excretion of thiamine measured on the following 2 days. When
the response, in terms of urinary output of thiamine, appeared to be
constant, 160 mg thiamine was given together with 120 mg potassium
metabisulfite. It was found that the addition of SO2 greatly reduced
the urinary output of thiamine, especially on the day when both were
given together (Causeret et al,, 1965).
In wine containing 400 ppm SO2, 50 per cent. of the thiamine was
destroyed in one week. However, no loss of thiamine was observed in 48
hours. The small amounts of SO2, resulting from the recommended
levels of usage in wine are therefore not likely to inactivate the
thiamine in the diet during the relatively short period of digestion
Interest in this aspect arises from the possibility that sulfate
formed metabolically from sulfite may serve to increase loss of
calcium in urine and faeces of man.
Levels of 0.5-0.7 per cent. calcium carbonate in the diet caused
increased faecal excretion and diminished urinary levels of Ca. Levels
up to 0.2 per cent. had no effect on the excretion of Ca. (Causeret &
Hugot, 1960). In a further experiment, diets containing 0.5 and 1 per
cent. calcium carbonate and 0.5 and 1 per cent. potassium
metabisulfite (2885 and 5770 ppm SO2 ) were administered to young
rats and the faecal and urinary excretion of Ca measured for 10 days.
At the lower level of dietary Ca (0.5 per cent.) both levels of the
metabisulfite caused a significant increase in the urinary excretion
of Ca but had no effect on the faecal excretion. At the higher dietary
Ca level (1 per cent.) the reverse was found. There was no difference
between the effects of the two levels of metabisulfite. This was
interpreted as being due to saturation of the body's capacity to
convert sulfite to sulfate (Hugot et al., 1965).
The levels of hepatic vitamin A were determined on both control
and test rats receiving 1.2 g/l potassium metabisulfite in the
drinking water (700 ppm as SO2). There was an insignificant decrease
in the vitamin A level in the liver of test animals after 10 days. In
another experiment, two groups of 40 rats each were kept for 4 months
on a diet containing only traces of vitamin A. The drinking water of
one group contained 1.2 g/l potassium metabisulfite. Hepatic vitamin A
levels were determined at the end or each month. A gradual reduction
in liver vitamin A levels was observed in both groups. The addition of
SO2 to the drinking water did not accentuate this reduction (Causeret
et al., 1965).
Rat. Groups of 20 Wistar rats (10 of each sex) were fed diets
containing 0.125, 0.25, 0.5, 1.0 and 2.0 per cent. of sodium hydrogen
sulfite (770-12 300 ppm as SO2) for 17 weeks. A group of 20 rats on
untreated diet served as controls. Immediately after preparation, all
diets ware stored at -18°C in closed glazed earthenware containers,
for not longer than 2 weeks. Measurements of loss of SO2 on keeping
each diet in air for 24 hours at room temperature revealed losses
amounting to 12.5, 10.0, 14.3, 8.2 and 2.5 per cent. of the sulfite
present in the diets as listed above, i.e. with Increasing SO2
content a decreasing proportion was lost.
After 124 days there was no effect on the growth of male rats. In
females, the 2.0 per cent. group grew as well as the controls; both
these female groups ware used for fertility studies, had given birth
to litters during the course of the test, and had raised their young.
The other female groups on lower levels of dietary sulfite were not
mated and showed significant depression of growth (as compared with
controls that had been mated). Haematological measurements at 7-8
weeks (all groups) and at 13 weeks (2 per cent. and controls) revealed
no effect of sulfite.
In the diet containing 2 per cent. sulfite, thiamine could not be
measured after 14 days at -18°C; at 1.0 per cent. and 0.25 per cent.
sulfite there was some loss of thiamine but this cannot be assessed
precisely since the initial values are not quoted. Measurements of
urinary thiamine excretion revealed substantial reduction at 1 week
and particularly at 13 weeks in all groups receiving more than 0.125
per cent, sulfite in the diet. Urine concentration tests were not
carried out on a sufficient number of animals to permit any firm
conclusion to be drawn.
Males and females of the control and 2 per cent. groups were
mated with rats drawn from the main colony. The only untoward
findings, with females of the 2 per cent. group, were lower weight of
the offspring at 7 and 21 days of life and 44.3 per cent. mortality as
compared with mortalities of 0, 2.8 and 3.8 per cent. in the other
groups of young rats. It is claimed that no changes were found in
relative organ weight (liver, heart, spleen, kidneys, adrenals,
testes) nor in microscopical appearance (above organs, stomach,
intestine, uterus, teeth and eyes). Since no measure of dispersion is
quoted, it is impossible to say weather the apparent severe reduction
in relative liver weight at the 0.125, 0.25, 0.5 and 1.0 per cent.
levels is significant (Centraal Instituut voor Voedingsonderzoek
Rat. A solution containing 1.2 g of potassium metabisulfite per
litre of water (700 ppm SO2) was administered to 80 weanling rats (40
of each sex) over a period of 20 months. A group of 80 rats given
distilled water served as controls. It was shown that the intake of
fluid by the test group was the same as that of the controls (but no
study appears to have been made of SO2 loss from the metabisulfite
solution). The intake of SO2 calculated from the consumption of water
was 30-60 mg/kg body-weight/day for males and 40-80 mg/kg
body-weight/day for females. The, following criteria provided no
evidence of toxic effect: growth rate, food intake, clinical
condition, haematological indices of blood and bone marrow (except
peripheral leucocyte count, which was increased in males), organ
weights (except spleen weight, which was heavier in females),
micropathological examination of a large number of tissues and
mortality rate. Fatty change in the liver was mostly slight or absent,
with a similar incidence and severity in test and control groups.
Reproduction studies over two generations revealed no effect except
for a slightly smaller number of young in each litter from test
animals and smaller proportion of males in each of these litters.
Growth of the offspring up to 3 months was almost identical in test
and control groups (Cluzan et al., 1965).
Four groups of 20 rats (10 of each sex on standard diet) were
given daily doses (30 ml/kg body-weight) of red wine containing 100
and 450 ppm SO2 an aqueous solution of potassium metabisulfite (450
ppm SO2) and pure water by oral intubation on 6 days each week for
four successive generations. The females ware treated for 4 and the
males for 6 months; the second generation was treated for 1 year. The
only effect seen was a slight reduction in hepatic cellular
respiration. All other parameters examined: weight gain, weight and
macroscopic or histological appearance of various organs, appearance
and behaviour, proportion of parturient females, litter size and
weight, biological value of a protein sample, showed no changes
attributable to SO2 (Lanteaume et al, 1965).
The long-term studies involving the administration of
metabisulfite in drinking water (700 ppm SO2) equivalent to 35 ml/kg
body-weight/day SO2 shows that this level apparently causes no toxic
effects in the rat.
From the long-term studies on metabisulfite in wine and water it
appears that the daily administration of fluid (30 ml/kg body-weight)
containing an equivalent of 450 ppm 50 SO2 (14 mg/kg body-weight/day)
produces no adverse effects in the rat.
Sulfite administered in food appears to be more toxic in animal
experiments than when administered in drinking water or wine. Further
studies are desirable [Note: text missing]