Cancer and Saccharin (Sweet N' Low, Sweet Twin and Necta Sweet)

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SACCHARIN

Explanation

Saccharin was evaluated by the Joint Expert Committee on Food
Additives in 1967, 1974, 1978 and 1980 (see Annex I, Refs. 14, 34,
48 and 54). In 1978, the Committee changed the ADI from 5 mg/kg to a
temporary ADI of 2.5 mg/kg and withdrew the conditional ADI of
15 mg/kg for dietetic purposes only. The decision to reduce the ADI
and to restrict the use of saccharin was based primarily on the
results of animal studies which indicated that excessive and long-term
ingestion of saccharin was potentially a carcinogenic hazard for
humans. At the 1980 meeting the temporary ADI of 2.5 mg/kg was
extended pending the completion of current investigations. Further
studies have been received and evaluated and the previous monograph
has been revised.

Saccharin may be produced in various ways, starting from toluene
(Remsen & Fahlberg, 1879), phthalic anhydride or phthalic acid
(Maumee, 1951) and o-chlorotoluene (Bungard, 1967). Accordingly, a
series of different chemical impurities could find their way to the
final product. The most widely used methods for the manufacture of
saccharin are the Remsen-Fahlberg and the Maumee processes (Munro et
al., 1974).

Although a universally applicable method for identification of
impurities in saccharin regardless of its production method is not
available, a recently published procedure (Stavric et al., 1976) for
isolation and identification of organic solvent soluble impurities
gives very satisfactory results for saccharin produced by the Remsen-
Fahlberg method. The major impurity in saccharins produced by this
procedure was identified as o-toluene-sulfonamide, one of the
intermediate reaction products in the preparation of saccharin
(Stavric et al., 1974b). Methods for isolation, identification and
quantitation of water-soluble impurities have also been reported
(Stavric et al., 1974a; Nelson, 1976).

BIOLOGICAL DATA

BIOCHEMICAL ASPECTS

Saccharin and saccharin salts (sodium, ammonium, calcium) have
been in use since the late nineteenth century, salt forms being more
soluble but of the same sweetening power as the acid form.

Absorption

The absorption of ingested saccharin in animals and man occurs
rapidly. With a pKa of 2.2, saccharin exists in acidic media
predominantly in the unionized form, which is the more readily
absorbed form in a number of animal species. Saccharin is more
completely absorbed from the guinea-pig (pH 1.4) and rabbit (pH 1.9)
stomach, than from the rat's stomach (gastric pH 4.2) (Ball, 1973;
Minegishi et al., 1972). In vitro perfusion of rat stomach and small
intestine with a solution of saccharin demonstrated considerable
absorption from the stomach at pH 1.0 and slow absorption from the
small intestine, i.e. less than 9% in 2 hours (Kojima et al., 1966).
The gastrointestinal absorption of saccharin appears to be somewhat
greater in monkeys than in rats (Pitkin et al., 1971a). In monkeys,
and also most probably in man, both gastric acidity and degree of
absorption are intermediate between those of the rabbit and guinea-pig
on one side, and the rat on the other (Ball, 1973). This also means
that the degree of absorption of saccharin could be dependent on food
intake which affects the acidity of the gastric contents.

Distribution and excretion

Lethco & Wallace (1975) studied the distribution of radioactivity
in organs and tissues of rats at various time intervals (1, 2, 4, 8,
24, 48 and 72 hours) following a single oral administration of
(3-14C)saccharin (50 mg/kg). Traces of radioactivity were found in
almost all organs 1 hour after dosing. Fat, brain and spleen contained
only minute quantities of 14C. Kidney, urinary bladder and liver
contained the highest amount of 14C, which peaked at 4 and 8 hours.
In subsequent experiments, rinsing the bladders of the treated rats
with 8, 0.5 ml portions of a 0.9% saline solution, they found that a
significant portion of the 14C activity was retained by or bound to
the bladder tissue (Lethco & Wallace, 1975).

In similar experiments, Matthews et al. (1973) found that a
single oral dose (1 mg/kg bw) of 14C-saccharin (labelled in the
carbonyl group) was rapidly absorbed from the gastrointestinal tract,
with peak tissue radioactivity occurring within 15 minutes of dosing.
With repeated dosing, within a single day or over a period of several
days, there was an accumulation of saccharin in some tissues,
particularly in the urinary bladder. The bladders of rats which
received daily doses of saccharin contained 19 times as much saccharin
as did the bladders of animals which received only a single dose.
After the removal of saccharin from the diet, almost complete tissue
clearance resulted in 3 days (Matthews et al., 1973).

Saccharin crosses the placenta of the rat (West, 1979; Ball et
al., 1977) and monkeys (Pitkin et al., 1971a,b) to a limited extent.
However, clearance from foetal tissues may be slower than from
maternal tissues, particularly the urinary bladder (Ball et al., 1977;
Pitkin et al., 1971a,b). Additionally, saccharin has also been found
to be excreted in the milk of goats (Carlson et al., 1923) and rats
(West, 1979).

Recently, Sweatman & Renwick (1980) studied the tissue
distribution and pharmacokinetics of saccharin in a series of
experiments. When saccharin was included in the diet of rats at a
level of 5%, there was a marked diurnal variation in the concentration
of saccharin in their plasma. Feeding male rats diets containing 1-10%
saccharin for 22 days resulted in tissue concentrations of saccharin
in the kidney and bladder which were greater than the concentration of
saccharin in the plasma, while the concentrations of saccharin in the
liver, lungs, spleen, adrenal, fat and muscle were less than in the
plasma. When female rats were fed a diet containing 5% saccharin, the
concentration of saccharin in the plasma and organs was greater than
for the male rats fed the 5% diet, particularly the concentration in
the kidney and urinary bladder. Additionally, the concentrations of
saccharin in the plasma and tissues of the male rats fed the diets
containing 7.5% or 10% saccharin had a higher than predicted
concentration, based on a linear extrapolation from the lower dose
levels. The authors attributed this to the animals' reduced ability to
eliminate saccharin.

A single i.v. dose of [3H] saccharin was given to male and
female anaesthetized rats at levels from 1 to 1000 mg/kg. The plasma
clearance of the [3H] saccharin was found to be dose dependent with
the high dosages resulting in a 60% decrease in clearance. When rats
were pretreated with probenecid prior to the i.v. administration of
[3H] saccharin at levels of 50-200 mg/kg, a 70% reduction in plasma
clearance was observed (Sweatman & Renwick, 1980). These findings were
similar to those of Bourgoignie et al. (1980) who used unanaesthetized
male and female rats to examine the mechanism by which saccharin was
excreted via the kidneys, using a range of [3H] saccharin
concentrations from 1 to 80 mg/dl in plasma. The authors concluded
that saccharin was excreted by a combination of filtration and tubular
excretion without resorption. The latter appeared to be carrier
mediated, since competitive inhibition was observed between saccharin
and para-aminohippurate.

Side-effects

During its long history of use, saccharin has been accused of
being responsible for a number of adverse effects both in human
beings and laboratory animals. Saccharin has been implicated in the
development of photosensitive skin eruptions in humans (Taub, 1972),
and as a hypoglycaemic agent in animals (Thompson & Mayer, 1959). Low

dietary concentrations of saccharin have been shown to alter the
concentration of serum lipid components in rats fed chemically-defined
diets (Purdom et al., 1973).

No effect was noted on human nitrogen balance or protein
utilization in man by doses up to 4 g of saccharin/day. Daily doses of
5 g of saccharin reduced albumin absorption and utilization by 0.94%
(Neumann, 1926a,b). No abnormal effects on total urinary nitrogen
excretion or uric acid output were noted by other investigators after
giving saccharin orally (Folin & Herter, 1912). No deleterious effects
on blood sugar, kidney function, vitamin utilization, blood
coagulation or enzyme activity were detected in man (NAS-NRC, 1955).

Metabolism

It has long been assumed that saccharin undergoes very little
metabolic conversion under normal dietary usage in animals and man
(NRC, 1974).

Rat

Four male and 2 female Sprague-Dawley rats were dosed with
14C-saccharin (40 mg/kg) uniformly labelled in the benzene ring. More
than 90% of the radioactivity was recovered in the urine collected for
96 hours after dosing. Only 1 radioactive spot was detected on the TLC
of the whole urine, which was identified as saccharin (Byard &
Golberg, 1973). Three to 4% of the dose was excreted in the 48-hour
collection of faeces as unmetabolized saccharin. During the first
4-8-hour collection period, no more than 0.3% of an oral dose of
14C-saccharin was excreted in the bile as unchanged saccharin.
Induction of mixed function oxidase activity in rat liver by treatment
with phenobarbitone had no influence on saccharin metabolism (Byard &
Golberg, 1973).

Kennedy et al. (1972), using albino rats treated with 10 or 20 µC
of sodium saccharin labelled with 14C in the carbonyl group, obtained
rapid excretion with total recovery from 90.21% to 100.57%. In 3 of
the 4 rats the amount recovered in the urine ranged from 82.65% to
96.95%, while in 1 animal it was only 67.91%. However, in this animal
a total of 31.34% of the radioactive material appeared in the faeces.
Most of the radioactivity excreted was within 24 hours of ingestion.
Expired air and tissues contributed very little to the total 14C
recovery. The same authors demonstrated the presence of trace amounts
of 2 hydrolytic products of saccharin (o-sulfamoylbenzoic acid and
ammonium carboxybenzene sulfonate) in urinary extracts. Minegishi et
al. (1972) failed to demonstrate the presence of any hydrolytic
products in the urine of rats given 35S-saccharin. Urine contained
70% of the administered saccharin and the remainder was in the faeces.
Similar results were found for rats given repeated daily doses of
saccharin for 4-5 weeks prior to 35S-saccharin administration,

indicating the lack of an induction mechanism for saccharin metabolism
(Minegishi et al., 1972). Lethco & Wallace (1975) obtained 56-87% of
the administered 14C-carbonyl labelled saccharin in urine and 10-40%
in the faeces. Ninety-nine per cent. of the urinary 14C was unchanged
saccharin, while the remainder was identified as the metabolite
o-sulfamoylbenzoic acid. Matthews et al. (1973) also failed to
demonstrate the presence of any saccharin metabolites in the urine of
rats given multiple low doses of 14C-saccharin (1 mg/kg bw).

Mouse, hamster, rabbit, guinea-pig and dog

Metabolism and rates of saccharin elimination appear similar in
these species. The fate of radioactive saccharin in the rabbit was
studied by Ball (1973) and by Lethco & Wallace (1975). Lethco &
Wallace (1975) made comparative studies of the metabolism of saccharin
in dog, rabbit, guinea-pig and hamster. Similar comparative studies by
Byard (1972) were done with mice, golden hamsters, guinea-pigs and
dogs, while Minegishi et al. (1972) studied saccharin metabolism in
guinea-pigs. These authors concluded that these species do not
metabolize saccharin.

Ball et al. (1974) failed to detect any metabolites in the urine
of female Wistar albino rabbits treated orally with a single dose of
20 mg/kg of (3-14C)saccharin. Seventy-two per cent. of the 14C was
excreted in the urine in 24 hours and 15% in the faeces. In 48 hours,
96% of the radioactivity was recovered. Similarly no metabolites were
found in urine of rats treated in the same way, which were kept up to
12 months on diets containing 1% or 5% sodium saccharin.

Monkey

Following a single oral dose of uniformly ring-labelled
14C-saccharin (1 mg/kg and 10 mg/kg) to young female rhesus monkeys,
98% of the radioactivity was excreted in the urine within 96 hours.
A very small amount of 2 hydrolytic products of saccharin
(o-sulfamoylbenzoic acid and ammonium carboxybenzene sulfonate) were
detected in urinary extracts (prepared by ethyl acetate extraction of
acidified urines) (Pitkin et al., 1971a). Byard & Golberg (1973) found
that 14C-saccharin (40 mg/kg) was not metabolized in male rhesus
monkeys. They also demonstrated that "artifactual metabolites" of
saccharin were produced if the urine was extracted under acidic
conditions. No induction of saccharin metabolism was found in groups
of rhesus monkeys receiving sodium saccharin (0, 20, 100 and
500 mg/kg/day) for 79 months. Metabolic studies conducted using
radioactive saccharin with these monkeys on several occasions during
the test indicated rapid, almost exclusively urinary excretion of
unmetabolized saccharin (McChesney et al., 1977).

Man

Orally administered saccharin appeared in the urine of man within
a half hour of dosing and was completely eliminated unchanged in 16-18
hours (Staub & Staehelin, 1936), some 90% being excreted in the urine
within 24 hours (Folin & Herter, 1912). Intravenous sodium saccharin
in doses of 2.5 g has been used without adverse effects in sick and
healthy people to determine circulation time (Fishberg et al., 1933).
Studies conducted with human subjects (McChesney & Golberg, 1973)
suggested that some degree of saccharin metabolism may occur in man,
since oral doses of saccharin could not be recovered quantitatively as
saccharin in the urine. Subsequent, more definitive studies, using
14C-labelled saccharin (Byard et al., 1974) demonstrated that the
reduced recovery was an artifact thought to be due to the binding of a
portion of the urinary saccharin to unidentified urinary constituents.

Four men received 500 mg of 14C-saccharin (uniformly labelled in
the benzene ring) and excreta was collected at intervals up to 96
hours post-dosing. More than 98% of the 14C was recovered within 48
hours (92.3% in urine, 5.8% in faeces). An additional 0.3% was
excreted in the 48-72-hour collection interval. None of the detected
saccharin was found to have been metabolized. Consequently, it was
concluded that man, like other species, does not metabolize saccharin
(Byard et al., 1974).

TOXICOLOGICAL STUDIES

Special studies on carcinogenicity: other than dietary exposure

Saccharin and croton oil together were tested for dermal
co-carcinogenicity in mice. Although the treated group showed a
greater incidence of skin papillomas compared with the control, the
difference was not statistically significant (Salaman & Roe, 1956).

Paraffin wax pellets containing saccharin when implanted in the
mouse bladder induce a significant incidence of bladder tumours and
this was interpreted as demonstrating a co-carcinogenic effect (Allen
et al., 1957).

Hicks et al. (1973, 1975) reported a rat model system for testing
bladder carcinogens wherein each rat was subjected to a pre-test
treatment consisting of a single instillation into the bladder of up
to 2 mg of methyl-N-nitrosourea (MNU). The test treatment consisted of
adding saccharin either to the diet or drinking-water at dosage levels
up to 4 g/kg/day. This treatment produced a significantly higher
incidence of bladder tumours with a dramatically reduced latent period
in the MNU plus saccharin treated rats compared to the untreated
control or animals receiving a single instillation of MNU or animals
receiving saccharin in their diet or drinking-water without any MNU
pretreatment. Although there does not appear to be any correlation

between the incidence of bladder tumours or bladder calculi (Hicks et
al., 1975), considerably more research is needed to evaluate the
biological significance of this model. Additionally, other
laboratories have been unable to reproduce these results (Mohr et al.,
1978; Green & Rippel, 1979; Green et al., 1980; Hooson et al., 1980).
Some of the suggested experimental variables which may have affected
the results include the quantity of MNU required to produce a
subcarcinogenic or initiator dose (Hooson et al., 1980); the length
and/or storage conditions for MNU; purity of MNU; or time lapse
between preparation of the MNU solutions and the treatment of the
animals (Mohr et al., 1978).

Subsequently, Cohen et al. (1978, 1979) developed a similar
2-stage cancer model wherein N[4-(5-nitro-2-furyl)-2-thiazolyl]
formamide (FANFT) served as the initiator, being administered in the
diet to rats at a level of 0.2%. In their initial studies, FANFT was
included in the diet for 6 weeks prior to feeding a saccharin (5%)
containing diet or a control diet for 2 weeks prior to the saccharin
(5%) diet. Both regimes resulted in a significant increase in the
incidence of bladder tumours. In subsequent studies on the rat, the
use of FANFT was decreased to 4 (Fukushima et al., 1981) and then 2
weeks (Cohen et al., 1982). The latter study employed various
treatment regimes that included FANFT- and saccharin-containing diets,
freeze ulceration, and the i.p. injection of cyclophosphamide. The
authors found that when bladder mucosal proliferation was increased,
following freeze ulceration and cyclophosphamide injection, the
bladder appears to be more sensitive to the promoting effects of
saccharin.

Using an experimental model similar to that of Cohen et al.
(1979), Nakanishi et al. (1980) administered N-butyl-N-
(4-hydroxybutyl) nitrosamine (BBN) in the drinking-water at a level of
0.001% for 4 weeks, either concurrently with or prior to feeding a
diet containing 5% saccharin. In the co-administration study,
saccharin enhanced the induction of urinary bladder hyperplasia
(simple, papillary or nodular) and papillomas, while the sequential
administration only enhanced the induction of papillary or nodular
hyperplasia.

In vitro promotional assay systems using saccharin have also
been reported. Mondal et al. (1978) reported that an initiating but
non-transforming dose of 3-methylcholanthrene (3MC) was needed in a
C3H/10T1/2 oncogenic transformation system before saccharin produced a
significant number of transformed cells. Sivak & Tu (1980) initiated
BALB/c3T3 cells with 3MC prior to the addition of saccharin, which was
not found to be active as a direct transforming or promoting agent of
Type III transformed foci.

Special studies on carcinogenicity: feeding studies

Mouse

Groups of 50 female Swiss mice (9-14 weeks of age) were fed one
of the following diets for 18 months: control (2 groups); 10% sucrose;
or 5% saccharin. Seven days prior to the start of the study, half of
the mice received a single intragastric instillation of 0.2 ml
polyethylene glycol while a similar group of mice received an equal
amount of polyethylene glycol which contained 50 µg benzo(a) pyrene
(BP), to ascertain whether the dietary treatments might be
co-carcinogenic with BP. Neither body weight nor survival in any of
the treatment groups was different from controls. The administration
of BP clearly increased the incidence of neoplasms arising in the
forestomach epithelium, but the other dietary treatments had no effect
upon this type of neoplasm. Although no neoplasms of the urinary
bladder were seen on careful macroscopic examination of all animals,
the bladders were not examined microscopically (Roe et al., 1970).

Kroes et al. (1977) fed diets containing 0, 0.2 and 0.5% sodium
saccharin during a 6-generation study in which 50 male and 50 female
mice from the F0, F3b and F6a generations remained on the test diets
for 20 months. There were no significant treatment-related tumorigenic
effects.

In a study performed in duplicate at the Bio-Research Consultants
Inc., groups of 25 male and 25 female mice (8 weeks of age) were fed
diets containing 0, 10 000 or 50 000 ppm (0, 1 or 5%) saccharin in the
diet for 24 months. The incidence of bladder tumours in the various
groups was as follows: male controls, 1 of 19; 1% saccharin (males),
none of 14 in the first replicate and none of 15 in the second; 5%
saccharin (males), 1 of 15 in the first replicate and 2 of 19 in the
second. Similar groups of female mice were also tested and none of the
female mice developed bladder tumours (Homburger, 1978).

Miyaji (1974) fed 50 male and 50 female mice (30 days of age)
diets containing 0, 0.2, 1.0 or 5.0% saccharin for 21 months. No
adverse treatment-related effects were reported.

Hamster

Groups of 30 male and 30 female hamsters, 8 weeks of age,
received the following levels of saccharin in their drinking-water for
the balance of their lifetime: 0, 0.156, 0.312, 0.625 or 1.25%.
Average survival time was 50-60 weeks. The overall incidence of
tumours in the control animals was 10.1% in 168 controls, while in the
saccharin group consisting of 299 animals the incidence was 14.7%. The
tumour types were similar in both groups and no urinary tract
neoplasms were found in either group (Althoff et al., 1975).

Rat (1-generation feeding studies)

Fitzhugh et al. (1951) performed the first 2-year saccharin
feeding study, where rats received a control diet (7 male and 9
female), or diets containing 1.0% (10 male and 10 female) or 5.0%
saccharin (9 male and 9 female). Saccharin had no apparent effect upon
mortality, haematology or organ weight (liver, kidney and spleen), but
did result in a slight growth depression in the 5% group. The only
significant pathological changes observed were in the 5% group where 7
animals (sex unspecified) had lymphosarcomas as well. The urinary
bladder was not histologically examined.

In another study, groups of 20 male and 20 female rats were fed
diets containing 0, 0.005, 0.05, 0.5 or 5.0% saccharin for 2 years. A
similar group was also given 1 ml of a 1% aqueous solution of trypan
blue s.c. every 2 weeks for 1 year as a positive control. In the 5%
group and the trypan blue group, mortality was higher than in the
controls. Mortality was lower than for the controls in the 0.005%
group. Retardation of growth was observed in the males and females of
the 5% group despite greater feed consumption.

One female and 4 male rats in the 5% saccharin group had bladder
calculi and 1 male had kidney calculi. The female rat with calculi had
an extensive transitional cell papilloma of the bladder, while another
5% female without calculi had hyperplasia and papillomatosis. No
evidence of nematodes was found in any of these animals (Lessel,
1967).

Schmahl (1973) reported spontaneous tumours in rats which were
fed diets containing either 0, 100 or 250 mg/kg of sodium saccharin
(52 males and 52 females per group). The incidence of Trichosomoides
crassicauda was 16% and was associated with a mild cystitis
condition, but no bladder tumours were observed in the control or
saccharin-treated groups.

Miyaji (1974) fed groups of 54 male rats, which were 40 days of
age, diets containing 0, 0.2, 1.0 or 5% saccharin for 28 months. No
significant tumorigenic effects were observed.

In a study conducted by Litton Bionetics, which was performed in
duplicate, 26 male and 26 female rats were fed diets containing 0, 1
or 5% saccharin for 24 months (NRC, 1974). The following was reported:
"The incidence of all tumors at 24 months in the untreated control
groups was 45% and 60% for the males, 80% and 55% for the females in
the two replicate experiments. It is important to note the extent of
the variation, indicative of the great variability of the assay as
well as the high tumor incidence in the controls."

However, "a single papilloma of the urinary bladder was found in
a female of the high dose saccharin group in the second replicate; no
other bladder tumor was found in the study".

Homburger (1978) fed groups of 25 male rats (8 weeks of age)
diets containing 0, 10 000 or 50 000 ppm (0, 1 or 5%) saccharin for 24
months. The study was performed in duplicate. The incidence of bladder
tumours in the various groups was as follows: controls, 1 of 16; 1%
saccharin, 1 of 13 in the first replicate and 1 of 15 in the second;
5% saccharin, 1 of 12 in the first replicate and none of 14 in the
second.

Groups of 54-56 male rats were fed either a control diet or a
diet containing 2.5 g of sodium saccharin/kg/day for 28 months. Ten to
16 rats in each group were killed at 12 months. Haematological-blood-
biochemistry and pathological examinations were performed on the rats
when sacrificed. The internal organs of all rats dying during the
study were examined macroscopically. A significant growth depression
was observed in the treated group, but no differences in mortality
were found. No bladder abnormalities were reported for either control
or treated animals (Furuya et al., 1975).

Groups of 60 male and 60 female weanling rats were fed diets
containing sodium saccharin produced by the Remsen-Fahlberg procedure
to provide daily doses of 0, 90, 270, 810 or 2430 mg of saccharin/kg/
day. The study ran for 26 months. Food consumption, body weight and
clinical examinations were conducted weekly on all rats. The animals
were free of the bladder parasite Trichosomoides crassicauda. One
bladder tumour was found in a male control, 1 each in a male and
female of the 90 mg/kg group and 2 male animals in the 810 mg/kg
group. The tumours were all transitional cell papillomas. Three
bladder calculi were observed grossly and 67 animals were found to
have bladder calculi small enough to pass through the urethra. The
presence of bladder calculi was not associated with the saccharin
treatment or with the presence of bladder tumours. Saccharin
administration was not accompanied by an increase in tumour incidence,
although high doses were associated with reduced body weight in both
sexes. No decrease in food consumption was observed. There was a
decreased longevity in male rats. Diarrhoea was observed in the
highest dose group, but was not accompanied by pathological evidence
of enteritis. The haematological parameters measured were not affected
by saccharin and saccharin had no effect upon the chemical composition
of the urine (Munro et al., 1975).

Rat (2-generation studies)

Tisdel et al. (1974) fed 20 male and 20 female rats one of the
following saccharin-containing diets for approximately 3 months prior
to mating: 0, 0.05, 0.5 or 5.0%. The saccharin used in this study was
produced by the Remsen-Fahlberg procedure. Following weaning, the F0
generation was killed and the F1 generation was weaned on to their
parents' diet, which they consumed until the study was terminated 100
weeks later. Thus, the F1 generation animals were exposed to
saccharin and its impurities during gestation, throughout lactation

via the dam's milk and for the remainder of their lifetime via the
diet. Transitional cell carcinomas of the urinary bladder were found
exclusively in the males of the 5.0% group with an incidence rate of 7
animals. One male in the 0.5% group had an epithelial hyperplasia of
the urinary bladder which the authors considered to be of a
precancerous type. Although no bladder tumours were observed in the
female rats, squamous cell carcinomas of the uterus were found in 1
animal in the 0.05% group and 2 animals in the 0.5% and 5.0% groups.

In a combination 3-generation chronic feeding study reported by
Taylor & Friedman (1974) and Taylor et al. (1980), 10 males and 20
females were fed one of the following diets containing sodium
saccharin produced by the Remsen-Fahlberg procedure: 0, 0.01, 0.1,
1.0, 5.0 or 7.5%. These animals were mated after 3 months on test on a
1 male to 2 female basis. The pups were weaned on to their parents'
diet and the F0 generation was killed. The study was terminated after
28 months. The reproductive data for this study are presented in
another section. The average body weights of treated groups was
generally less than that of the control animals; the lowest body
weights were observed in the 7.5% group in which the body weight was
depressed by 15% compared to the control, but survival was not
affected by dietary treatment. The most significant pathological
finding was the incidence of bladder tumours, with 1 tumour being
found in a male control, 1 in a male on 5% sodium saccharin and a
total of 9 fed the 7.5% sodium saccharin diet (7 males and 2 females).
All the tumours were diagnosed as transitional cell carcinomas except
for 1 tumour in a male in the 7.5% saccharin group which was diagnosed
as a transitional cell papilloma.

Since the Tisdel et al. (1974) study and the study by Taylor &
Friedman (1974) have used saccharin which contained levels of o-TS up
to 4660 ppm (0.466%) (Stavric et al., 1973), it was suggested (NRC,
1974) that o-TS might be the chemical responsible for the bladder
tumours observed in these 2-generation studies.

Arnold et al. (1977, 1980) fed groups of 50 male and 50 female
rats one of the following diets for 141 weeks: control, 2.5 mg
o-TS/kg/day, 25 mg o-TS/kg/day, 250 mg o-TS/kg/day with 1% NH4Cl in
the drinking-water or 5% sodium saccharin. The saccharin used in these
experiments was produced by the Maumee procedure containing no
detectable amounts of o-TS. After the F0 animals had been on test for
3 months, they were mated on a 1:1 basis. The pups were weaned on to
their parents' diet and both generations remained on their respective
diets for their lifetime (30-32 months). The reproductive data are
presented in another section. The animals were free of the bladder
parasite Trichosomoides crassicauda. The only treatment-related
effects, other than on lactation index and litter size, were a
decreased growth rate in the 2 250 mg/kg o-TS groups and the saccharin
group, and the incidence of bladder tumours.

This is the first study where F0 male animals receiving sodium
saccharin in their diet had a statistically higher incidence of
bladder tumours than did the control groups (P <0.05). The authors
suggest this observation may be attributable to the length of time on
test plus the fact that the animals were only 30 days old when the
experiment was initiated.

Special epidemiological studies

Available reports of epidemiological studies do not provide clear
evidence to support or refute an association between bladder cancer in
males and the use of saccharin.

Evaluation of these studies on saccharin is difficult for many
reasons. In some cases, only a very limited number of patients was
studied. In others, a distinction between consumption of saccharin and
cyclamates was not made, or smoking had a confounding effect.

In 2 recently published epidemiological studies in humans, the
authors concluded that no association between saccharin consumption
and the incidence of bladder cancer was present (Morrison & Buring,
1980; Wynder & Stillman, 1980).

Special studies on mutagenicity

A review on this subject was recently published (Kramers, 1975).
A predominant feature emerging from a review of the literature on
saccharin's mutagenic effects is a lack of consistency in the results.
As an explanation, it was suggested that the detected effects might be
due to a contaminant(s), present in varying amounts in different
samples of saccharin (Kramers, 1975).

This suggestion is supported by the findings of Stoltz et al.
(1977) who reported mutagenic activity with the Ames Salmonella assay
for some, but not all, samples of organic solvent extracts of various
saccharin samples produced by the Maumee and Remsen-Fahlberg methods.

The US Technology Assessment panel (OTA, 1977) commissioned a
battery of 12 short-term tests, of which 10 had been completed prior
to the publication of the panel's report. The sister chromatid
exchange studies conducted with human lymphocytes, the mouse lymphoma
forward mutation test and the Chinese hamster ovary cell test for
chromosome aberration all gave positive results, while negative tests
included the Ames Salmonella test, mitotic recombination in yeast,
unscheduled DNA synthesis, the Pol A test, Drosophila sex-linked
recessive lethal test, in vitro transformation test and the
induction of plasminogen activator. Subsequently, Mondal et al. (1978)
reported that saccharin did not produce oncogenic transformation of
mouse embryo fibroblasts (C3H/10T1/2) unless an initiating dose of
3-methylcholanthrene was used.

Batzinger et al. (1977) orally administered saccharins of
differing degrees of purity to mice and then collected the urine to
test for mutagenic activity in the Ames Salmonella test. By modifying
the growth and selection medium, they were able to detect mutagenic
activity for all but the most purified samples of saccharin.

When yeast was grown in a medium containing saccharin of
different purities, Moore & Schmick (1979) found an inverse
relationship between the incidence of mitotic crossing-over and the
purity of the saccharin sample. The results of Saxholm et al. (1979)
with the mouse embryo fibroblast test (C3H/10T1/2) and sister
chromatid exchange, as well as those by Abe & Sasaki (1977) and Wolfe
& Rodin (1978), are somewhat divergent, possibly due to use of
saccharins with different purities or the use of different endpoints.
Simmon et al. (1982) reported on the results of the two independent
laboratories which attempted to confirm previous short-term positive
studies when saccharin was introduced into the medium of the following
tests: mouse lymphoma, human lymphocyte SCE, the CHO cytogenetic
analyses as well as the standard and modified Salmonella test. As the
findings from both laboratories were not always similar, the authors
concluded that the differences observed were attributable in part to
intrinsic test system variability generated at high concentrations of
a relatively non-toxic material.

Various impurities found in Remsen-Fahlberg-produced saccharin
have been tested in the following systems: ortho- and para-
toluenesulfonamide were negative with Drosophila (Kramers, 1977) and
in a modified Salmonella assay (Poncelet et al., 1979, 1980) but
weakly mutagenic in the Ames Salmonella test and the Basc test in
Drosophila melanogaster (Eckhardt et al., 1980), while ortho- and
para-sulfamoylbenzoic acid were ineffective mutagens in the Ames
Salmonella assay, the Basc test in Drosophila melanogaster and
micronucleus test in mice (Eckhardt et al., 1980). Such impurities as
o-sulfobenzoic acid, ammonium o-sulfobenzate, p-sulfobenzoic acid and
p-sulfamoylbenzoic acid were also negative in a modified Salmonella
assay (Poncelet et al., 1979, 1980).

Special studies on pharmacokinetics

Following the oral administration of 14C-saccharin to pregnant
rats on the twenty-first day of gestation, Ball et al. (1977) found
that 0.6% of the administered dose was detectable in the foetuses and
radioactivity in their urinary bladder was 10 times higher than in
other tissues 26 hours post-dosing.

The pharmacokinetic properties of saccharin in man have recently
been reported. Colburn et al. (1981) had a man (who had not previously
consumed saccharin on a routine basis) and a woman (who was a regular
user of saccharin) ingest a single 100 mg dose of saccharin, following
which blood and urine samples were obtained during the next 24 hours.

The authors calculated the half-life and apparent volume of
distribution to be 1.2 and 6.6 hours, and 41.6 and 13.1 litres,
respectively. While no measurable saccharin was detected in the plasma
of either subject prior to the test, the female subject was found to
be excreting 17.4 µg/hour during the 12 hours prior to the start of
the test. The female subject had chronically used saccharin up to 10
days prior to the test period. The authors proposed that a deep
peripheral second compartment existed in the female subject due to her
chronic use of saccharin. Sweatman et al. (1981) administered
saccharin to 3 adult male subjects and periodically determined the
concentration of saccharin during the next 400 minutes after oral
(2 g) and i.v. (10 mg/kg) doses. The authors found that their i.v.
data fitted a two-compartment open model with the half-life of the
second phase being 70 minutes. Following oral ingestion, the decrease
in plasma saccharin levels was more complex; however, if the same dose
of saccharin was given after a meal, the peak plasma levels were lower
and were delayed by 100-120 minutes versus the i.v. dosing situation.
After 96 hours, 90% of the latter dose had been recovered in the urine
and 8% in the faeces.

Special studies on reproduction

Mouse

Groups of 21 pregnant mice received 40-168 mg/kg bw per day of
saccharin through the production of 3 successive litters without
deleterious effect on growth, litter number and pups per litter when
compared with controls fed sugar. No histological studies were
performed (Lehmann, 1929).

A 6-generation study was carried out by Kroes et al. (1977) in
which mice were fed diets containing 0, 0.2 or 0.5% sodium saccharin
during the entire study. The criteria of reproductive performance
were: fertility index; viability index (percentage survival at 5
days); lactation index (percentage survival at 21 days); body weight
at 21 days; and number of infertile males. No consistent changes in
any of these parameters over the 6 generations were observed. Although
body weight at 21 days was suppressed in some groups of some
generations, this response was not consistently observed in later
generations at the same treatment level. Additionally, there was no
evidence of any teratogenic effects.

Rat

One aspect of a long-term feeding study with saccharin performed
at the Wisconsin Alumni Research Foundation (WARF) reported by Tisdel
et al., 1974 involved the mating of rats from the same treatment
groups, on a 1:1 basis, after they had received one of the following
diets for 14 weeks: 0, 0.05, 0.5 or 5% saccharin, produced by the
Remsen-Fahlberg procedure. The mothers continued to receive the test
diets during mating, gestation and lactation.

"Pregnant animals were isolated about 5 days before parturition
and allowed to deliver and care for their young until the litters were
weaned 21 days after birth. Survival of progeny was recorded through
day 28."

"Data recorded for each litter included: identification of
parents, number of pups born (total, alive and dead); number of
survivors 4 and 21 days after birth; and weight of survivors through
28 days of age. The results showed no effect of saccharin on mating
efficiency, survival of pups born alive, or weight gain of surviving
offspring. All groups fed saccharin had smaller average litter sizes
and reduced average percent live births, when compared to controls,
but neither response appeared to be related to dose level."

"The differences were not statistically analyzed, but examination
of data on individual animals shows that the lower average values are
attributable to extremely poor performance of one or two animals in
each group, not to a general reduction. The apparent effect is
therefore probably within the customary experimental variability."

"Surviving and dead fetuses were examined for gross
abnormalities; no evidence of teratogenicity was found" (NAS, 1974).

A 3-generation reproductive study (Taylor & Friedman, 1974;
Taylor et al., 1980) was carried out in which rats were continuously
fed diets containing 0, 0.01, 0.1, 1.0, 5.0 and 7.5% sodium saccharin
produced by the Remsen-Fahlberg procedure. In the F1a animals, the
male offspring of dams receiving 5.0% or 7.5% saccharin weighed 12-20%
less than controls and the females weighed 17-29% less than control
animals, but both sexes were able to overcome some of these initial
weight differences during a chronic feeding portion of the study. In
the F2a litters, the fertility and viability indices were unaffected
by treatment, but average litter size was slightly decreased for dams
receiving 5.0% or 7.5% sodium saccharin; the survival index, weaning
index and body weights of these pups were below controls. In the F2b
litters, only body weight at weaning was lower than controls for these
same two groups.

During a 2-generation lifetime feeding study by Arnold et al.
(1977, 1980), 50 male and 50 female rats were fed one of the following
diets (tap water ad libitum except where noted): control; 2.5 mg/kg
ortho-toluene-sulfonamide (o-TS); 25 mg/kg o-TS; 250 mg/kg o-TS plus
drinking-water containing 1% NH4Cl; or 5% sodium saccharin produced
by the Maumee procedure, which contained no detectable o-TS. o-TS is a
major impurity in Remsen-Fahlberg-produced saccharin, and o-TS is a
known inhibitor of the enzyme carbonic anhydrase (Miller et al.,
1956). The animals were mated on a 1:1 basis after 3 months on test.
The only treatment-related effect on reproduction was a significant
depression in the lactation index in the 2.5 mg/kg o-TS group and a
significant decrease in the litter size of the 250 mg/kg o-TS group.

There were no treatment-related effects upon fertility, gestation or
viability indices or any effect upon the body weight of pups or the
number of males in the litter.

Special studies on teratology

Teratogenic studies with mice (Tanaka, 1964; Lorke, 1969; Kroes
et al., 1977), rats (Bough et al., 1967; Fritz & Hess, 1968; Lessel,
1970; Taylor & Friedman, 1974) and rabbits (Bough et al., 1967;
Klotzsche, 1969; Lessel, 1970; Tanaka et al., 1973) have been negative
to date.

Lederer & Pottier-Arnould (1973) fed pregnant female rats diets
containing 0.3% saccharin throughout gestation. The pups from
saccharin-treated dams had a 37.9% incidence of lens anomalies versus
12.4% incidence for the control animals.

Acute toxicity

LD50
Animal Route (mg/kg bw) Reference

Mouse Oral 17 500 Taylor et al., 1968
i.p. 6 300 Taylor et al., 1968
17 500 Tanaka, 1964

Hamster Oral (F) 8 700
(8-day LD50 value) (M) 7 400 Althoff et al., 1975

Rat Oral 14 200-17 000 Taylor et al., 1968
i.p. 7 100 Taylor et al., 1968

Rabbit Oral 5 000-8 000 (LD) Folin & Herter, 1912

Dog i.v. 2 500 (LD) Becht, 1920

Short-term studies

Rat

Kennedy et al. (1976) fed groups of 10 male and female
weanling rats one of the following diets for 13 weeks: (1) control;
(2) 20 000 ppm (2%) sodium saccharin; (3) 20 000 ppm (2%)
o-sulfamoylbenzoic acid (o-SABA); (4) 20 000 ppm (2%) ammonium
o-carboxybenzene sulfonate (A-o-CBS); (5) 100 ppm (0.01%) sodium
saccharin plus 450 ppm (0.045%) o-SABA and 450 ppm (0.045%) A-o-CBS;
(6) 500 ppm (0.05%) sodium saccharin plus 2250 ppm (0.225%) o-SABA and

2250 ppm (0.225%) A-o-CBS; (7) 2000 ppm (0.2%) sodium saccharin plus
9000 ppm (0.9%) of o-SABA and 9000 ppm (0.9%) A-o-CBS. (A-o-CBS and
o-SABA are hydrolytic products of saccharin.) Weight gain and feed
consumption were determined on a weekly basis and behavioural changes
were looked for. Haematological studies (RBC, total and differential
WBC, Hgb and haematocrit), clinical blood chemistry (glucose, BUN,
serum alkaline phosphatase and SGPT) and urinalysis (albumin, glucose,
microscopic elements, pH and specific gravity) were conducted prior to
starting the study, at the midpoint and upon termination. Every animal
was autopsied and examined histologically. The liver, kidney, spleen
and gonad weights were determined and organ/body weight ratios
calculated. No consistent or significant changes in any parameter were
observed.

Groups of 14 male and 14 female rats (75-100 g) received either a
control diet or one containing 0.5% sodium saccharin for 38 days. The
inclusion of saccharin in the diet depressed body weight gains and
feed intake, but efficiency of feed utilization was similar to
controls. Although diarrhoea was a common observation, no overt
behavioural changes were apparent. Gross and microscopic inflammatory
and hydropic changes in the liver and kidneys of the saccharin-treated
group were reported, but the extent of these changes was not described
(Taylor et al., 1968).

Groups of 25 rats (5 males and 20 females) were fed diets
containing 0, 1.0 or 10% saccharin for 36 weeks. Similar groups of
rats were fed diets containing 0, 0.1 or 1.0% saccharin for a
lifetime. One female from each group in the second experiment was
mated and 4 progeny from each litter were fed a diet containing the
same amount of saccharin as their parents for a lifetime. Growth was
retarded at the 10% level, but no adverse effects were seen at the
lower levels upon histological examination of the major organs (Fantus
& Hektoen, 1923).

In a 4-week feeding study, Anderson (1979) fed rats diets
containing 0, 1, 3, 5 or 7.5% sodium saccharin and observed: (1)
transient diarrhoea in the 5% and 7.5% group; (2) a dose-dependent
decrease in urinary ammonia; and (3) a decreased faecal odour. These
observations led the author to hypothesize that diets with high levels
of sodium saccharin may lead to changes in the intestinal microflora.
To test the hypothesis, Anderson & Kirkland (1980) fed rats diets
containing 0 or 7.5% saccharin for 10 days. The saccharin diet
produced an increased weight of the caecal tissues and contents; did
not decrease the total number of anaerobic microbes, but did result in
the inability to detect a specific anaerobic microbe; increased the
number of aerobic microbes; and reduced the amount of ammonia produced
from urea by Proteus vulgaris.

Dog

In addition to the rat study cited above, Kennedy et al. (1976)
also fed the same 7 diets to groups of 3 male and 3 female beagle dogs
for 16 weeks. In addition to the parameters indicated for the rats,
the authors also determined SGOT and protein-bound iodine in this
study. No significant treatment-related effects were observed.

One male and 1 female dog received 150 mg/day of saccharin in
their food for 18 months without any adverse effects on weight,
fertility or other bodily functions. Their pups developed normally
(Bonjean, 1922).

When given doses of 175-350 mg/day for 100 days, dogs developed
hyperaemia of the lungs, liver, myocardium and kidney, as well as
cloudy swelling of renal glomeruli and convoluted tubules (de Nito,
1936).

Groups of 4 dogs (sex unspecified) received 6 daily doses of
sodium saccharin (0.065 g/kg dissolved in water) for 11 months via
intragastric intubation. The dogs were observed daily for survival,
general physical condition, behaviour, food and water consumption, and
character and frequency of excreta. The animals were weighed weekly.
Laboratory tests included renal and hepatic function tests which were
performed at 0, 1, 2, 4, 9 and 11 months on test. During the second
half of the study, 1 dog receiving saccharin became anorexic and
eventually died, but no abnormal organ pathology was observed. After
10 months of test, all saccharin-treated dogs developed soft, poorly-
formed stools without frank diarrhoea. Body weight, haemoglobin, total
leucocyte count, non-protein nitrogen and phenosulfonephthalein
retention were within normal limits and total erythrocyte count,
bilirubin and urinary analyses were also unaffected. Gross and
histological appearance of viscera were normal in the test group
(Taylor et al., 1968).

Monkey

McChesney et al. (1977) reported no pathological or any other
significant changes in growth, haematology or clinical chemistry in
groups of 3 rhesus monkeys of each sex receiving 0 (control) or 500 mg
of saccharin/kg/day and in groups of 2 male and 2 female monkeys
receiving 20 or 100 mg of saccharin/kg/day 6 days per week for 79
months. One animal in each experimental group and 2 control animals
died before termination of the experiment. None of the deaths were
attributable to the treatment. Metabolic studies conducted on several
occasions during the test indicated a rapid, almost exclusively
urinary excretion of unmetabolized saccharin.

OBSERVATIONS IN MAN

Doses of 1.5-3.0 g of saccharin/day in man caused a persistent
sweet metallic taste (Carlson et al., 1923). Single doses of 5-10 g
have been tolerated and even 100 g orally is said to have caused no
harm. A few non-fatal cases of acute poisoning and allergic response
have been reported (NAS-NRC, 1955).

During high-intake balance studies, 3 male volunteers received
0.3 g of sodium saccharin/day for a maximum of 4 months and 1-1.5 g
of sodium saccharin/day for a maximum of 2 months. All of the
administered saccharin was fully accounted for. Seven volunteers
received 0.15-0.3 g of saccharin/day for 1.3 months without adverse
effects except for an increased urine output (Folin & Herter, 1912).
Doses of 90-180 mg of saccharin/day were well tolerated by children
aged 10-12 years for 13 months (Jessen, 1890). Diabetic patients have
received as much as 4.8 g daily for 5 months without adverse effect
(Neumann, 1926a,b) and 0.4-0.5 g/day for 15-24 years without any
adverse effects (National Academy of Sciences - National Research
Council, 1955).

The principal adverse effects that have been reported from
saccharin are as follows:

(1) Mild digestive disturbances were noted by Herter & Folin (1911) in
volunteers ingesting doses of 1-1.5 g of saccharin/day. Loose stools
were observed in clinical studies when subjects consumed cyclamates
plus saccharin in doses of about 7 g/day (Berryman et al., 1968). At
these dosages, the saccharin intake was about 0.7 g/day. Evidence from
other studies indicates that cyclamate alone at intakes of 5-7 g/day
may cause loose stools.

(2) Allergic responses, principally skin reactions of a phototoxic or
photosensitivity type occur but appear to be of low incidence and, in
some cases, may have been due to cyclamate being ingested at the same
time (Fujita et al., 1965; Stritzler & Samuels, 1956; Kingsley, 1966;
Boros, 1965; Meisel, 1952; Gordon, 1972; Taub, 1972). Some authors
have suggested that there may be a cross-sensitivity to sulfonylureas
and similar drugs known to cause phototoxic skin reactions. Contact
dermatitis and photosensitivity or phototoxic reactions have not been
noted in persons occupationally exposed to saccharin (NAS, 1974).

Comments

The present Committee reviewed further findings from
epidemiological studies that did not reveal any evidence for a
saccharin-associated increase in bladder tumours. Final assessment of
the epidemiological aspect of saccharin consumption will be made
following the review of a large-scale study, now in progress but

nearing completion. The Committee decided to extend the temporary ADI
of 0-2.5 mg/kg for saccharin to 1984 and required the submission of
the results of a long-term feeding study in rats and epidemiological
studies.

EVALUATION

Estimate of temporary acceptable daily intake for man

0-2.5 mg/kg bw.

FURTHER WORK OR INFORMATION

Required by 1984

(1) Submission of the results of a long-term feeding study, currently
in progress.

(2) Submission of the results of the epidemiological study, currently
in progress.

REFERENCES

Abe, S. & Sasaki (1977) Chromosome aberrations and sister chromatid
exchanges in Chinese hamster cells exposed to various chemicals,
J. natl. Cancer Inst., 58, 1645

Allen, M. J. et al. (1957) Cancer of the urinary bladder induced in
mice with metabolites of aromatic amines and tryptophan,
Br. J. Cancer, 11, 212

Althoff, J. et al. (1975) A chronic study of artificial sweeteners in
Syrian golden hamsters, Cancer Lett., 1 21

Anderson, R. L. (1979) Response of male rats to sodium saccharin
ingestion: Urine composition and mineral balance, Fd. Cosmet.
Toxicol., 17, 195

Anderson, R. L. & Kirkland, J. J. (1980) The effect of sodium
saccharin in the diet on caecal microflora, Fd. Cosmet.
Toxicol., 18, 353

Arnold, D. L. et al. (1977) Long term toxicity study with
orthotoluene-sulfonamide and saccharin, Toxicol. Appl.
Pharmacol., 41, 164, abstract No. 78

Arnold, D. L. et al. (1980) Long-term toxicity of orthotoluene-
sulfonamide and sodium saccharin in the rat, Toxicol. Appl.
Pharmacol., 52, 113

Ball, L. M. (1973) The metabolism of saccharin and related compounds.
Report 4, from the Department of Biochemistry, St Mary's Hospital
Medical School, London, UK. Unpublished report submitted to WHO

Ball, L. M., Renwick, A. G. & Williams, R. T. (1974) The fate of 14C
saccharin in rats chronically fed on saccharin, Biochem. Soc.
Trans., 2, 1084

Ball, L. M., Renwick, A. G. & Williams, R. T. (1977) The fate of
14C saccharin in man, rat and rabbit and of 2-sulphamoyl 14C
benzoic acid in the rat, Xenobiotica, 7, 189

Batzinger, R. P., Ou, S. Y. L. & Bueding, E. (1977) Saccharin and
other sweeteners: Mutagenic properties, Science, 198, 944

Becht, F. C. (1920) Influence of saccharin on the catalases of the
blood, J. Pharmacol. Exp. Therap., 16, 155

Berryman, G. H. et al. (1968) A case for safety of cyclamate and
cyclamate-saccharin combinations, Am. J. Clin. Nutr., 21, 673

Bonjean, E. (1922) Rev. Hyg., 44, 50, cited in Chemisch.
Weekbl., 40, 26-32

Boros, E. (1965) An experience with artificial sweeteners,
J. Am. Med. Assoc., 194, 571

Bough, R. G. et al. (1967) Unpublished report submitted by Boots Pure
Drug Co. to WHO

Bourgoignie, J. J., Hwang, K. H. & Bricker, N. S. (1980) Renal
excretion of 2,3-dihydro-3-oxobenzisosulfonazole (saccharin),
Am. J. Physiol., 238(1), F10-5

Bungard, G. (1967) Die SusStoffe Der Deut Apotheker, 19, 150

Byard, J. L. (1972) Observations on the metabolism of saccharin,
Toxicol. Appl. Pharmacol., 22, 291, abstract No. 46

Byard, J. L. & Golberg, L. (1973) The metabolism of saccharin in
laboratory animals, Fd. Cosmet. Toxicol., 11, 391

Byard, J. L. et al. (1974) Excretion and metabolism of saccharin in
man. II. Studies with 14C-labelled and unlabelled saccharin,
Fd. Cosmet. Toxicol., 12, 170

Carlson, A. J. et al. (1923) Studies on the physiological action of
saccharin, J. Metabl. Res., 3, 451

Cohen, S. M. et al. (1979) Promoting effect of saccharin and
DL-tryptophan in urinary bladder carcinogenesis, Cancer Res.,
39, 1207

Cohen, S. M. et al. (1978) Co-carcinogenicity testing of saccharin and
DL-tryptophan following oral initiation with N-[4-(5-nitro-2-
furyl)-2-thiazolyl] formamide. In: Guggenheim, B., ed., Health
and sugar substitutes, Proceedings of the ERGOB Conference on
Sugar Substitutes, Geneva, 30 October - 1 November 1978, Basel,
S. Karger, 70 pp.

Cohen, S. M. et al. (1982) Effect of regenerative hyperplasia on the
urinary bladder: Carcinogenicity of sodium saccharin and
N-[4-(5-nitro-2-furyl)-2-thiazolyl] formamide, Cancer Res.,
42, 65

Colburn, W. A., Bekersky, I. & Blumenthal, H. P. (1981) A preliminary
report on the pharmacokinetics of saccharin in man: single oral
dose administration, J. Clin. Pharmacol., 21, 147

de Nito, G. (1936) Sull'azione tossica della saccarina. Ricerche
isto-patologiche, Boll. Soc. Ital. Biol. Sper., 11, 934

Eckhardt, K. et al. (1980) Mutagenicity study of Remsen-Fahlberg
saccharin and contaminants, Toxicol. Lett., 7, 51

Fantus, B. & Hektoen, L. (1923) Saccharin feeding of rats, J. Amer.
Pharm. Ass. Sci. Ed., 12, 318

Fishberg, A. M., Hitzig, W. M. & King, F. H. (1933) Measurement of
the circulation time with saccharin, Proc. Soc. Exptl. Biol.
Med., 30, 651

Fitzhugh, O. G., Nelson, A. A. & Frawley, J. P. (1951) A comparison of
the chronic toxicities of synthetic sweetening agents, J. Am.
Pharm. Assoc., 40, 583

Folin, O. & Herter, C. (1912) US Dept. Agric. Rep. No. 94

Fritz, H. & Hess, R. (1968) Prenatal development in the rat following
administration of cyclamate, saccharin and sucrose,
Experientia, 24, 1140

Fujita, H. et al. (1965) Five cases which showed diffuse erythema and
edematous papules possibly caused by saccharin, Acta Derm.
(Kyoto), 60, 303

Fukushima, S. et al. (1981) Effect of L-tryptophan and sodium
saccharin on urinary tract carcinogenesis initiated by
N-4-(nitro-2-furyl)-2-thiazolyl formamide, Cancer Res., 41,
3100

Furuya, T. et al. (1975) Long term toxicity study of sodium cyclamate
and saccharin sodium in rats, Japanese J. of Pharmacol., 25
(Suppl.), 55 (abstract No. 60)

Gordon, H. H. (1972) Allergic reaction to saccharin, Am. J. Obstet.
Gynecol., 113, 1145

Green, U. & Rippel, W. (1979) Bladder calculi in rats treated with
nitrosomethylurea and fed artificial sweeteners, Exp. Path.,
17, 561

Green, U. et al. (1980) Syncarcinogenic action of saccharin or sodium
cyclamate in the induction of bladder tumors in MNU-pretreated
rats, Fd. Cosmet. Toxicol., 18, 575

Herter, C. & Folin, O. (1911) Influence of saccharin on the nutrition
and health of man. Report No. 94, US Department of Agriculture,
Government Printing Office, Washington, D.C.

Hicks, R. M., Wakefield, J. St J. & Chowaniec, J. (1973)
Co-carcinogenic action of saccharin in the chemical induction of
bladder cancer, Nature, 243, 347

Hicks, R. M., Wakefield, J. St J. & Chowaniec, J. (1975) Evaluation of
a new model to detect bladder carcinogens or co-carcinogens;
results obtained with saccharin, cyclamate and cyclophosphamide,
Chem. Biol. Interactions, 11, 225

Homburger, F. (1978) Negative lifetime carcinogen studies in rats and
mice fed 50 000 ppm saccharin. In: Galli, C. L., Paoletti, R. O.
& Vettorazzi, G., eds, Chemical toxicology of food, Amsterdam,
Elsevier/North Holland Biomedical Press

Hooson, J. et al. (1980) Orthotoluenesulphonamide and saccharin in the
promotion of bladder cancer in the rat, Brit. J. Cancer, 42,
129

Jessen, F. (1890) Zur Wirkung des Saccharin, Arch. F. Hyg., 10, 64

Kennedy, G. L., Jr, Faucher, O. E. & Calandra, J. C. (1972) Metabolic
fate of saccharin in the albino rat, Fd. Cosmet. Toxicol.,
10, 143

Kennedy, G. L., Jr, Faucher, O. E. & Calandra, J. C. (1976) Subacute
toxicity studies with sodium saccharin and two hydrolytic
derivatives, Toxicology, 6, 133

Kingsley, H. J. (1966) Sensitivity to saccharin, Cent. Afr. J. Med.,
12, 243

Klotzsche, C. (1969) Zur Frage der teratogenen und embryotoxischen
Wirkung von Cyclamat, Saccharin und Saccharose,
Arzneimittel-Forsch., 19, 925

Kojima, S. & Ichibagase, H. (1966) Synthetic sweetening agents. X.
Thin-layer chromatography of sodium cyclamate, saccharin sodium
and dulcin, Yakuzaigaku, 26, 115

Kramers, P. G. N. (1975) The mutagenicity of saccharin, Mutation
Research, 32, 81

Kramers, P. G. N. (1977) Mutagenicity of saccharin in Drosophila:
The possible role of contaminants, Mutation Research, 56, 163

Kroes, R. et al. (1977) Long term toxicity and reproduction study
(including a teratogenicity study) with cyclamate, saccharin, and
cyclohexylamine, Toxicology, 8, 285

Lederer, J. & Pottier-Arnould, A. M. (1973) Influence of saccharin on
embryonal development in the pregnant rat, Le Diabète, 21, 13

Lehmann, K. B. (1929) Fütterungsversuche mit und Ohne Saccharin an
Mäusepaaren, Zugleich ein Beitrag zum Studium der Frage Minimaler
Giftwirkung, Arch. F. Hyg., 101, 39

Lessel, B. (1967) Unpublished report submitted by Boots Pure Drug Co.
to WHO

Lessel, B. (1970) Carcinogenic and teratogenic aspects of saccharin.
SOS/70 Proceedings, 764 pp. In: Third International Congress,
Food Science and Technology, Washington, D.C.

Lethco, E. J. & Wallace, W. C. (1975) The metabolism of saccharin in
animals, Toxicology, 3, 287

Lorke, D. (1969) Untersuchungen von Cyclamat und Saccharin auf
embryotoxische und teratogene Wirkung an der Maus,
Arzneimittel-Forsch., 19, 920

Matthews, H. B., Fields, M. & Fishbein, L. (1973) Saccharin:
distribution and excretion of a limited dose in the rat,
J. Agr. Food Chem., 21, 916

Maumee (1951) Chem. Eng. News (1963), 41, 76 (December 9)

McChesney, E. W. & Golberg, L. (1973) The excretion and metabolism of
saccharin in man. I. Methods of investigation and preliminary
results, Fd. Cosmet. Toxicol., 11, 403

McChesney, E. W., Coulston, F. & Benitz, K. F. (1977) Six-year study
of saccharin in rhesus monkeys, Toxicol. Appl. Pharmacol.,
41, 164 (abstract No. 79)

Meisel, E. H. (1952) A letter to the editor: Headache from saccharin,
J. Amer. Med. Assoc., 148, 163

Miller, W. H., Dessert, A. M. & Roblin, R. O., Jr (1956) Heterocyclic
sulfonamides as carbonic anhydrase inhibitors, J. Amer. Chem.
Soc., 72, 4893

Minegishi, K. I., Asahina, M. & Yamaha, T. (1972) The metabolism of
saccharin and the related compounds in rats and guinea pigs,
Chem. Pharm. Bull., 20, 1351

Miyaji, T. (1974) Discussion about food additives: Saccharin, Nippon
Riusho, 32, 839

Mohr, U. et al. (1978) Syncarcinogenic action of saccharin and sodium
cyclamate in the induction of bladder tumours in MNU-pretreated
rats. In: Guggenheim, B., ed., Health and sugar substitutes,
Proceedings of the ERGOB Conference on Sugar Substitutes, Geneva,
30 October - 1 November 1978, Basel, S. Karger, 64 pp.

Mondal, S., Brankow, D. W. & Heidelberger, C. O. (1978) Enhancement of
oncogenesis in C3H/10T1/2 mouse embryo cell cultures by
saccharin, Science, 201, 1141

Moore, C. W. & Schmick, A. (1979) Genetic effects of impure and pure
saccharin in yeast, Science, 205, 1007

Morrison, A. S. & Buring, J. E. (1980) Artificial sweeteners and
cancer of the lower urinary tract, New Engl. J. Med., 302,
537

Munro, I. C., Stavric, B. & Lacombe, R. (1974) The current status of
saccharin. In: Wimek, C. L. ed, Toxicology Annual 1974, New
York, Dekker, p. 71

Munro, I. C. et al. (1975) A carcinogenicity study of commerical
saccharin in the rat, Toxicol. Appl. Pharmacol., 32, 513

Nakanishi, K. et al. (1980) Effect of sodium saccharin and caffeine on
the urinary bladder of rats treated with N-butyl-N-(4-hydroxy-
butylnitrosamines), GANN, 71, 490

National Academy of Sciences - National Research Council (1955) The
safety of artificial sweeteners for use in food, Publication No.
386

National Academy of Sciences - National Research Council (1974)
Safety of saccharin in the human diet. Committee of Food
Protection, NAS 1974, Washington, D.C.

National Research Council (1974) Safety of saccharin and sodium
saccharin in the human diet. Prepared for the Food and Drug
Administration. PB-238 137

Nelson, J. J. (1976) Quantitation of o- and p-sulfamoylbenzoic acids
in commercial saccharin by high-performance liquid
chromatography, JAOAC, 59, 243

Neumann, R. O. (1926a) Wird die Ausnutzung des Nahrungseiweisses durch
Saccharin beeinflusst? Archiv fur Hygiene Bakteriologie, 96,
265

Neumann, R. O. (1926b) Bemerkungen zu der vorstehenden Erwiderung von
W.A. Uglow auf meine Arbeit "Wird die Ausnutzung des
Nahrungseiweisses durch Saccharin beeinflusst"? Archiv fur
Hygiene Bakteriologie, 97, 275

Office of Technology Assessment (1977) Cancer testing technology and
saccharin, Washington, DC, US Govt. Print. Off., 149 pp.

Pitkin, R. M. et al. (1971a) Saccharin metabolism in Macaca mulatta,
Proc. Soc. Exp. Biol. Med., 137, 803

Pitkin, R. M. et al. (1971b) Placental transmission and fetal
distribution of saccharin, Am. J. Obstet. Gynecol., 111, 280

Poncelet, F., Mercier, M. & Lederer, J. (1980) Saccharin: Para forms
of some impurities are not mutagenic in Salmonella typhimurium,
Fd. Cosmet. Toxicol., 18, 453

Poncelet, F. et al. (1979) Absence of mutagenic activity in
Salmonella typhimurium of some impurities found in saccharin,
Fd. Cosmet. Toxicol., 17, 229

Purdom, M. E., Hyder, K. & Pybas, M. D. (1973) Effects of saccharin on
rats fed chemically defined diets, J. Amer. Diet. Assoc., 63,
635

Remsen, I. & Fahlberg, C. (1879-1880) On the oxidation of substitution
products aromatic hydrocarbons. IV. On the oxidation of
orthotoluenesulphamide, Amer. Chem. J., I, 426

Roe, J. C., Levy, L. S. & Carter, R. L. (1970) Feeding studies on
sodium cyclamate, saccharin and sucrose for carcinogenic and
tumor-promoting activity, Fd. Cosmet. Toxicol., 8, 135

Salaman, M. H. & Roe, F. J. C. (1956) Further tests for tumor
initiating activity: N,N-di-(2-chloroethyl)-p-aminophenyl-butyric
acid (CB 1348) as an initiator of skin tumor formation in the
mouse, Brit. J. Cancer, 10, 363

Saxholm, H. J. K. et al. (1979) Carcinogenesis testing of saccharin.
No transformation or increased sister chromatid exchange observed
in two mammalian cell systems, Europ. J. Cancer, 15, 509

Schmahl, D. (1973) Fehlen einer kanzerogener. Wirkung von Cyclamat,
Cyclohexylamin und Saccharin bei ratten, Arzneimittel-Forsch.,
23, 1466

Simmon, V. F. et al. (1982) A comparative investigation of the
mutagenic and clastogenic effects of saccharin, ascorbic acid,
pantothenic acid and ergocalciferol in several in vitro
mammalian and microbial assay. Special report prepared by SRI
International, Melo Park, CA, and Litton Bionetics Inc.,
Kensington, MD, and submitted to JEFCA

Sivak, A. & Tu, A. S. (1980) Cell culture tumor promotion experiments
with saccharin, phorbol myristate acetate and several common food
materials, Cancer Letters, 10, 27

Staub, H. & Staehelin, R. (1936) Saccharin. Med. Press Circ., 193,
419

Stavric, B. et al. (1973) Unpublished report submitted to WHO

Stavric, B., Lacombe, R., Munro, I. C., By, A. W., Klassen, R. &
Ethier, J. (1974a) Studies on water soluble impurities in
commercial saccharins. 167th Amer. Chem. Soc. National Meeting,
Los Angeles, 31 March - 5 April 1974 (Abstract ANAL, 171)

Stavric, B., Lacombe, R., Watson, J. R. & Munro, I. C. (1974b)
Isolation, identification and quantitation of o-
toluenesulphonamide, a major impurity in commercial saccharins,
J. Assoc. Off. Anal. Chem., 57, 678-681

Stavric, B., Klassen, R. & By, A. W. (1976) Impurities in commercial
saccharin. I. Impurities soluble in organic solvents, J. Assoc.
Off. Anal. Chem., 59, 1051

Stoltz, D. R. et al. (1977) The mutagenicity of saccharin impurities.
I. Detection of mutagenic activity, J. Environ. Pathol.
Toxicol., 1, 139

Stritzler, C. & Samuels, L. (1956) Urticaria caused by sensitivity to
saccharin, Arch. Derm., 74, 433

Sweatman, T. W. & Renwick, A. G. (1980) The tissue distribution and
pharmacokinetics of saccharin in the rat, Toxicol. Appl.
Pharmacol., 55, 18

Sweatman, T. W., Renwick, A. G. & Burgers, C. D. (1981) The
pharmacokinetics of saccharin in man, Xenobiotica, 11, 531

Tanaka, R. (1964) LD50 of saccharin or cyclamate for mice embryo in
the 7th day of pregnancy (fetal median lethal dose: FLD50),
J. Iwate Med. Assoc., 16, 330-337

Tanaka, S. et al. (1973) Studies on the teratogenicity of food
additives. Effects of sodium saccharin on the development of rats
and mice, J. Food Hyg. Soc. Japan, 14, 371-379

Taub, S. J. (1972) Untoward reactions of saccharin, Eye, Ear, Nose
and Throat Mon., 51, 405

Taylor, J. D. et al. (1968) Toxicological studies with sodium
cyclamate and saccharin, Fd. Cosmet. Toxicol., 6, 313

Taylor, J. M. & Friedman, L. (1974) Combined chronic feeding and
three-generation reproduction study of sodium saccharin in the
rat, Toxicol. Appl. Pharmacol., 29, 154 (abstract No. 200)

Taylor, J. M., Weinberger, M. A. & Friedman, L. (1980) Chronic
toxicity and carcinogenicity to the urinary bladder of sodium
saccharin in the in utero-exposed rat, Toxicol. Appl.
Pharmacol., 54, 57

Thompson, M. M. & Mayer, J. (1959) Hypoglycemic effects of saccharin
in experimental animals, Am. J. Clin. Nutr., 7, 80

Tisdel, M. O. et al. (1974) Long-term feeding of saccharin in rats.
In: Inglett, G. E., ed., Sweetness, symposium, Westport, Conn.,
AVI Publishing Co., 145 pp.

West, R. W. (1979) The exposure of fetal and suckling rats to
saccharin from dosed maternal animals, Toxicol. Lett., 7, 409

Wolfe, S. & Rodin, B. (1978) Saccharin-induced sister chromatid
exchanges in Chinese hamster and human cells, Science, 200,
543

Wynder, E. L. & Stillman, S. D. (1980) Artificial sweetener use and
bladder cancer: a case-control study, Science, 207, 1214

See Also:
Toxicological Abbreviations
Saccharin (FAO Nutrition Meetings Report Series 44a)
Saccharin (FAO Nutrition Meetings Report Series 48a)
SACCHARIN (JECFA Evaluation)
Saccharin (IARC Summary & Evaluation, Supplement7, 1987)
Saccharin (IARC Summary & Evaluation, Volume 22, 1980)

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