UREA
First draft prepared by
Dr P. Olsen
Institute of Toxicology, National Food Agency of Denmark
Ministry of Health, Soborg, Denmark
1. EXPLANATION
Urea is a white crystalline powder with a cooling saline taste
(Merck, 1968). Urea occurs naturally in mammals and is an excretory
end-product of amino acid metabolism. Urea is formed in the liver.
Urea has not been evaluated previously by the Joint FAO/WHO Expert
Committee on Food Additives.
Urea is used in sugar-free chewing gum to adjust the texture.
A heavy user of chewing gum may consume approximately. Chewing gum
may contain up to 3% urea, and intake from this source could be up
to 300 mg urea/day. The Committee considered urea only for
evaluation in relation to its use in chewing gum.
2. BIOLOGICAL DATA
2.1 Biochemical aspects
2.1.1 Absorption, distribution, and excretion
Urea has little or no nutritional value to monogastric mammals
(Briggs, 1967). Ruminants are able to utilize urea as a source for
food protein (Blood & Henderson, 1963). Urea present in the blood
of ruminants appears to be actively transported across the rumen
wall into the lumen and used as a nitrogen source (Schmidt-Nielsen,
1958).
A study in pregnant rats which were injected subcutaneously
with urea dissolved in 0.9% NaCl solution showed that urea diffused
readily through the placenta. The concentrations of urea in the
maternal liver, thigh muscle and in the whole fetus were equal two
hours after injection (Luck & Engle, 1929).
In dogs injected intraperitoneally with 3% urea solution, urea
diffused throughout the body and was present in tissue fluid at
concentrations equal to, or greater than, that present in the
extracellular fluid (Grollman & Grollman, 1959).
The distribution of urea was determined in 4 young pigs given
15N-labelled urea in the diet. Fifty-two per cent of the
administered 15N-labelled urea was excreted in the urine after 48
hours and 1.9% during the subsequent 48 hours. Faecal 15N
excretion over 96 hours accounted for only 1.3% of the amount
administered. Less than 1% of the 15N was found in the liver,
muscle and blood cells, study concluded that this indicates
incorporation of 15N in body proteins. Only 60% of the
administered 15N was recovered (Grimson et al., 1971).
In another study, the distribution of 14C-labelled urea after
intraperitoneal injection was determined by radioactivity analysis
and autoradiography techniques in the brain and spinal fluid of
fasted cats. In the brain and cerebrospinal fluid the highest urea
concentration was reached 6 hours following injection.
Autoradiography showed dense areas in the cerebral and cerebellar
cortex. White matter showed the least radioactivity (Schoolar
et al., 1960).
Renal excretion of urea is rapid and chiefly by glomerular
filtration (Sollmann, 1957). Renal tubular secretion (Sollmann,
1957) and reabsorption also occur (Mountcastle, 1974).
2.1.2 Biotransformation
Urea is an excretory end-product of amino acid metabolism in
mammals. The formation of urea takes place in the liver. This is a
cyclic process in which the initial step is the reaction between
carbon dioxide and ammonia to yield carbamyl phosphate. Carbamyl
phosphate reacts with ornithine to form citrulline which combines
with aspartate to form argininosuccinate. This product is cleaved
to fumarate and arginine. The terminal step is the hydrolysis of
arginine, yielding urea and regenerating ornithine. This cycle of
reactions involves several enzymes including carbamyl phosphate
synthetase, ornithine carbamylase, argininosuccinate synthetase and
arginine-lyase. The fetal liver was capable of synthesizing urea 28
days (in pigs), and 19 days (in rats) after gestation (Kennan &
Cohen, 1959).
2.1.3 Effects on enzymes and other biochemical parameters
No information available.
2.2 Toxicological studies
2.2.1 Acute toxicity studies
The results of acute toxicity studies with urea are summarized
in Table 1.
The clinical symptoms observed in cattle included ataxia,
weakness, abdominal pain, dyspnoea, excessive salivation, frothing,
violent struggling and bellowing. Acute urea toxicity in cattle may
be due to ammonia formed by the rapid breakdown of urea by rumen
microorganisms (Blood & Henderson, 1963).
The clinical signs of acute urea toxicity in ponies were
typical of severe central nervous system derangement:
incoordination, dilated pupils, sluggish pupillary response to
light, depressed palpebral and corneal reflexes, slow respiratory
rate, rapid and weak peripheral pulse, cold and clammy skin, and
pressing of the head against fixed objects until falling at death
(Hintz et al., 1970).
Table 1. Results of acute toxicity studies with urea.
Species Sex Rte Dosage, LD/MLD Reference
mg/kg bw
Dog ? sc 3 000-9 000 LD Abderhalden, 1935
Dog ? iv 3 000 LD Abderhalden, 1935
Rabbit ? sc 1 000-2 000 LD Abderhalden, 1935
Hamster ? iv 4 000-8 000 LD Abderhalden, 1935
Sheep(1) ? po 160 LD Satapathy & Panda, 1963
Cattle M po 511 MLD Dinning et al., 1948
Cattle(1) F po 600 MLD Stiles et al., 1970
Cattle(2) M po 1080 MLD Stiles et al., 1970
Ponies ? po 3461 LD Hintz et al., 1970
(1): Not adapted to urea
(2): Adapted to urea
2.2.2 Short-term toxicity studies
2.2.2.1 Dogs
Twelve unilaterally nephrectomized dogs were injected
subcutaneously with 10% urea solution (3 000-4 000 mg/kg bw) every 8
hours over a period of 45 days. Serum urea levels ranged from
600-700 mg/100 ml ´ hour after injection. Except for a mild
drowsiness and increased diuresis urea did not induce any severe
toxic symptoms (Balestri et al., 1971).
2.2.2.2 Ruminants
A gradual increase in the amount of urea in rations up to
1762 mg/kg bw/dy to steers over a period of 70 days did not cause
distress (Dinning et al., 1948). However, without adaptation to
urea, doses of 166 mg/kg bw/dy and 232 mg/kg bw/dy urea caused
sudden death in sheep and cattle, respectively (Satapathy & Panda,
1963). Tolerance to urea was reduced in starving ruminants and in
ruminants on a low protein diet (Blood & Henderson, 1963).
2.2.3 Long-term toxicity/carcinogenicity studies
2.2.3.1 Mice
Three groups of 50 C57B1/6 mice of each sex were administered
either 0.45% (approx.674 mg/kg bw/day) 0.90% (approx.1350 mg/kg
bw/day), or 4.5% (approx.6750 mg/kg bw/day) urea (no information on
purity reported) in the diet for 1 year. The control group
comprised 100 mice of each sex. The identity of urea was confirmed
by melting point comparison. Biochemical and haematological
parameters were not included in the study. No body weight
depression was noted at terminal necropsy for mice of either sex at
any dose levels. Survival of all treated groups were unaffected.
Among treated female mice there was a significant increased
occurrence of malignant lymphomas in the middle dose-group. The
incidence of malignant lymphomas was 10/92 in controls and 7/43,
10/38 (p=0.008) and 9/50 in the low-, middle-, and high-dose groups,
respectively. The increased incidence of malignant lymphomas among
middle-dose female mice was of questionable biological significance
since the occurrence was not dose-related. Urea was non-
carcinogenic in this study (Fleischman et al., 1980).
2.2.3.2 Rats
Groups of 50 Fischer 344 rats of each sex were administered
either 0, 0.45% (approx.225 mg/kg bw/day), 0.90% (approx.450 mg/kg
bw/day) or 4.5% (approx.2 250 mg/kg bw/day) urea (no information on
purity reported, identity of urea was confirmed by melting point
comparison) in the diet for 1 year. Biochemical and haematological
parameters were not included in the study. No body weight
depression was noted at terminal necropsy for rats of either sex at
any dose levels. The middle-dose male rats showed decreased
survival (89%) relative to controls (95%)(statistics not reported).
The survival of the other dose groups remained unaffected.
Among treated male rats, there was a significant increased
linear trend (p=0.008), and a higher proportion of interstitial cell
adenomas of testis in the high-dose group (p=0.004). The incidence
of interstitial cell adenomas was 21/50 in the controls, and was
27/48, 25/48, and 35/50 in the low- middle- and high-dose groups,
respectively. The statistically significant increased incidence of
interstitial cell adenomas in male rats was of questionable
biological significance since this tumour may occur in 100% of
controls. Urea was non-carcinogenic in this study (Fleischman
et al., 1980).
2.2.3.3 Ruminants
A calf received 4.3% urea (approx. 1290 mg/kg bw) in feed over
a period of 12 months caused. Increased diuresis was observed
throughout the experiment. Histologically, renal hyaline
degeneration, tubular casts and several areas of liver necrosis were
found (Hart et al., 1939).
2.2.3 Long-term toxicity/carcinogenicity studies
No information available.
2.2.4 Reproduction studies
No information available.
2.2.5 Special studies on genotoxicity
The results of genotoxicity studies with urea are summarized in
Table 1.
2.3 Observations in humans
2.3.1 Blood values, distribution, metabolism, excretion and effects
on other parameters.
The absorption of urea was studied in 8 healthy fasting male
volunteers by means of a colon perfusion technique. Only 5% of the
urea perfused through the colon was absorbed. The authors concluded
that the colon was relatively impermeable to urea (Wolpert et al.,
1971).
The mean concentrations of blood urea in healthy human subjects
were 28.9 mg/100 ml (range 16-54 mg/100 ml) in 298 men and 21.7
mg/100 ml (range 12-47 mg/100 ml) in 278 woman. Urea levels tended
to increase with age (Keating et al., 1969).
Correlation between blood urea and the content of urea in
parotid fluid has been found (Shannon & Prigmore, 1961).
The normal value of urea in saliva (unstimulated) was reported
to be 3.3 mM/l (200 mg/l) with a range of 2.4-12.5 mM/l. Daily
production of saliva varied from 500-1 500 ml (Geigy, 1981a).
Average daily urinary excretion of urea in adults was estimated
to be 20.6 g. The urinary excretion of urea was proportional to
protein intake and was increased on a high protein diet. Urea
excretion was decreased during growth and pregnancy or due to action
of insulin, growth hormone and testosterone (Geigy, 1981b).
Urea excretion was also diminished in cases of reduced urea
formation due to liver diseases (Geigy, 1981b) and nephropathies
(Mountcastle, 1974).
The enzyme system necessary for urea synthesis in human fetuses
was functional when mesonephric glomeruli were present (Kennan
et al., 1959).
Urea has been shown to have a neutralizing effect on acidified
plaque layers produced in the oral cavity after consumption of
fermentable carbohydrates (Imfield, 1984 & 1985).
2.3.2 Toxicity
Four healthy male human subjects received an oral dose of
15 grams urea (approx.250 mg/kg bw), blood urea rose from 30 mg/100
ml (mean level prior to treatment) to a mean level of 42 mg/100 ml
(range: 40-46) within 15 to 60 minutes. The increased blood urea
levels returned to normal after 3 hours. Fifteen patients with
renal impairment, after similar oral treatment with 15 g urea,
showed a rise in blood urea from 50 mg/100 ml (mean level prior to
treatment; range: 26-220) to a mean level of 75 mg/100 ml (range:
38-299). The increased blood urea levels returned to the levels
observed prior to treatment after more than 4 hours (Archer & Robb,
1925).
Table 1. Results of genotoxicity tests for urea
Test System Test Object Concent. Result Reference
of urea
In vitro bacterial S.typhimurium TA98, TA100 ? Neg. Ishidate, et al., 1981
mutagenicity assay TA1537
Mammalian cell Mouse lymphoma TK locus 329-628 µM/l Pos. (2) Garberg, et al., 1988
mutation assay (1) assay
Chromosomal aberration Chinese hamster fibroblast 16 mg/ml Pos. (2) Ishidate & Odashima, 1977
assay (1) cell
Chromosomal aberration Chinese hamster fibroblast 13 mg/ml Pos. (2) Ishidate et al., 1981
assay (1) cell
Chromosomal aberration Human leucocytes 50 µM (4) Pos. (3) Oppenheim & Fishbein, 1965
assay
In vivo Chromosomal Bone marrow cell 25 g/kg Pos. Chaurasia & Sinha, 1987
aberration assay bw (5)
(1) With and without metabolic activation.
(2) Only positive without metabolic activation; negative with metabolic activation.
(3) The authors considered the positive result as a non-specific effect of high-molarity urea solution on cell division.
(4) Concentration, probably per l.
(5) The applied oral dose appears unrealistically high, it exceeds lethal dose by several times.
Six healthy subjects were given oral treatment of 2 000 to
3 000 mg/kg bw urea hourly for a period of 24 hours to induce
azotaemia. Serum urea-nitrogen values ranged from 60-120 mg/100 ml
(approx. blood urea of 128-257 mg/100 ml; [conversion factor for
"blood urea" {serum urea} to "blood urea-nitrogen" = 2.14]) (Eknoyan
et al., 1969).
No toxic effects were found in humans if the blood
urea-nitrogen was below 45 mg/100 ml (approx. blood urea of
96 mg/100 ml). Loss of appetite, nausea and vomiting developed at
about 70 mg/100 ml (approx. blood urea of 150 mg/100 ml) (Crawford &
McIntosh 1925).
Signs of malaise, vomiting, weakness, lethargy, and bleeding
were noted in patients with renal failure who were loaded with urea
in the blood at levels of 300-600 mg/100 ml for 60 to 90 days.
Blood urea concentrations below 300 mg/100 ml were well tolerated by
the patients (Johnson et al., 1972).
80 patients were hospitalized after ingestion of urea
fertilizer mistaken for table salt. The symptoms observed were
nausea, persistent violent vomiting, excitement, and severe general
convulsions. Complete recovery of all patients was observed within
a few days (Steyn, 1961).
Six healthy human subjects were maintained at serum
urea-nitrogen concentrations at 60 to 120 mg/100 ml (approx. blood
urea of 128-257 mg/100 ml) over a period of 24 hours. Prolonged
bleeding time and a drastic reduction of the blood platelet
adhesiveness was observed in 5/6 subjects (Eknoyan et al., 1969).
Oxygen uptake in human blood platelets in vitro was reduced
7%, 14%, and 19% at urea levels of 100, 300, and 500 mg/100 ml,
respectively (Schneider et al., 1967).
The relationship between plasma urea concentration and low
birth weight in infants of non-toxaemic mothers was investigated.
16 infants with low birth weight had a statistically significantly
higher mean plasma urea concentration of 23.2 mg/100 ml in
comparison with a mean value of 18.6 mg/100 ml in 90 infants with
normal birth weight (p<0.02) (McKay & Kilpatrick, 1964).
Ingestion of 60 grams of urea per day (approx. 1 000 mg/kg
bw/day), in divided doses, over a period of 3 1/4 days, resulted in
prolonged clearance time of glucose in adults (Perkoff et al.,
1958).
The irritant potential of urea dissolved in water was
determined on human scarified skin. On the third day following
daily application, a solution of 7.5% urea showed slight skin
irritation, and a solution of 30% urea showed marked skin
irritation. A solution of 30% urea did not affect normal skin
(Frosch & Kligman, 1977).
Intra-amniotic injection of up to 300 ml 30% urea solution has
been used to induce therapeutic abortion (Anteby et al., 1973).
2.3.3 Drug interactions
Treatment of 40 men suffering from sulfonamide-resistant
gonorrhoea with urea (500 mg/kg bw/dy) for a period of 3 days
enhanced the effect of sulfonamide in 52% of the patients. A
combination of urea and sulfathiazole inhibited the growth of
gonococci in vitro, although neither alone was effective
(Schnitker & Lenhoff, 1944).
The inhibitory effect of sulfadiazine on the growth of E.coli
in vitro was enhanced in combination with urea (Tsuchiya et al.,
1942).
2.3.4 Use in human medicine
Urea has been used in human medicine as diuretic at doses of 15
to 60 grams/day. The mechanism of the diuretic effect originates
from increased glomerular filtration due to osmotic action of urea
(Sollmann, 1957).
In the oral therapy of sickle-cell anaemia, urea at doses of
667-2 000 mg/kg bw/day, in divided doses, was given for periods of
3 weeks to 9 months. Side effects included increased diuresis,
thirst, gastrointestinal discomfort, nausea and vomiting (Bensinger
et al., 1972).
3. COMMENTS
The Committee reviewed biochemical studies, short-term toxicity
studies in dogs and ruminants, carcinogenicity studies in rats and
mice, mutagenicity studies, and studies on effects in human
volunteers. It noted that most of the available data were either
inadequate or of little relevance for the evaluation of urea as a
food additive. As urea is a naturally-occurring constituent of the
body, the Committee carried out its evaluation in accordance with
the principles relating to materials of this type outlined in
Annex 1, reference 76.
4. EVALUATION
Since urea is a natural end-product of amino acid metabolism in
humans, and that approximately 20 grams/day is excreted in the urine
in adults (proportionately less in children) the Committee concluded
that the use of urea at levels of up to 3% in chewing-gum was of no
toxicological concern.
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