Hexamethylenetetramine

Classification/MAK value: not yet established

Synonyms: hexamethyleneamine
hexamethylenetetraamine
hexamine
HMT
HMTA
methenamine
1,3,5,7-tetraazaadamantane
Trade names: Aminoform Formin
®
Ammoform Uritone
®
Cystamin Urotovet
®
Cystogen Urotropin
Chemical name (CAS): 1,3,5,7-tetraazatricyclo-
3,7
[3.3.1.1 ]decane
CAS number: 100-97-0
Structural formula: CH2
N N
C C
H2 N H2
H2 C CH2 CH2
N

Molecular formula: C6H12N4

Molecular weight: 140.19
Melting point: colourless, hygroscopic crystals
which do not melt but sublime at 260-
295 °C [1]
Solubility: in water (20 °C) 874 g/l
in g%:
water (25 °C) 66.7 [1]
water (12 °C) 81.3 [2]
chloroform (20 °C) 13.4 [2]
methanol (20 °C) 7.25 [2]
ethanol (20 °C) 2.89 [2]
acetone (20 °C) 0.65 [2]
ether (20 °C) 0.06 [2]
petroleum ether (20 °C) c0 [2]
Purity: technical grade: about 99%
356 Hexamethylenetetramine Volume 5

Stability: During storage of highly pure HMTA

(containing no volatile compounds) at
4 °C in the dark or at 20°C in daylight
in sealed glass containers for 6
months, no volatile organic com-
pounds are formed [3].
Thermal decomposition yields NOX,
CO, CO2 and formaldehyde (HCHO);
combustion yields CO2, N2, NOX and
traces of NH3 but no HCN [1]; hydro-
lysis yields HCHO and NH3.
Production: reaction of formaldehyde and am-
monia in the gas phase and/or in
aqueous solution [1, 2, 4]
OH
6HCHO + 4NH3 C6 H12N4 + 6H2O
H+

Uses: It is used in the production of phenolic

resins as a curing agent, in the pro-
duction of nitrilo-triacetic acid, as a
starting material in the production of
explosives (Hexogen (cyclonite), Oc-
togen), as a vulcanization accelerator
in the rubber industry, as a biocide in
metal-working fluids, etc. It was once
used as a preservative in the food-
stuff industry (to preserve fish) but
since 1.1.64 is no longer used in the
®
FRG. As methenamine or Urotropin ,
it is used to disinfect the urinary tract
[1,2,5].

Note
No dimethylnitrosamine was formed in in vitro nitrosation experiments with HMTA and
sodium nitrite in acidic solution [3].

1 Toxic Effects and Modes of Action

In man, hexamethylenetetramine is absorbed rapidly from the gastrointestinal tract.
Accumulation is not observed. Excretion of the orally administered substance takes place
largely in the urine (up to 80 % within 3 hours). Depending on the local pH, form-
aldehyde is released from hexamethylenetetramine more rapidly in the stomach than in
Volume 5 Hexamethylenetetramine 357

the urine. The side effects described after administration of relatively large doses of
hexamethylenetetramine (up to 6 g per day) in the therapy of acute and chronic infect-
ions of the urinary tract include gastrointestinal disorders, bladder irritation, haematuria
and skin eruptions.
Hexamethylenetetramine irritates human skin and mucous membranes and can cause
hypersensitivity reactions.
Oral administration of the substance to experimental animals produces only slight
acute toxicity. Long-term administration of relatively large doses does not have adverse
effects on body functions, body weight, organ weights or life expectancy.
Animal studies have not provided evidence of a teratogenic potential of hexa-
methylenetetramine .
In in vivo studies, weak mutagenic effects are seen only with sublethal doses. In vitro
mutagenicity studies also demonstrate that very high concentrations are required to
produce weak mutagenic effects which appear to result from hydrolytic release of
formaldehyde in aqueous media.
No carcinogenic effects are seen after oral administration of high doses of hexa-
methylenetetramine to rats and mice. However, local tumours have been described after
subcutaneous injection of the substance; they are probably primarily a reaction to
chronic local irritation at the injection site.

1.1 Pharmacokinetics
Hexamethylenetetramine is absorbed rapidly from the human gastrointestinal tract. The
hexamethylenetetramine concentration in plasma and serum reaches a maximum one to
two hours after oral administration (one daily dose of 1 g or two daily doses of 1 g at 12
hour intervals for 6 to 7 days). The half-life of the substance in plasma and serum was
determined as 4 hours. Accumulation was not observed. Excretion took place largely in
the urine (80 % of the dose of 1 g twice per day) [6, 7]. Hexamethylenetetramine is
distributed in body fluids such as saliva, bile, cerebrospinal and synovial fluids and
pleural exudate [8,9]. After oral administration of 1 g hexamethylenetetramine to preg-
nant women, it was demonstrated that the substance crosses the placental barrier and is
found in the amniotic fluid. After 4 hours, the level in the umbilical vein plasma was
similar to that in the maternal plasma; hexamethylenetetramine concentrations of the
same order of magnitude were also found in mother's milk [6].
After intravenous injection of a hexamethylenetetramine dose of 100 mg/kg body
weight into a dog whose kidneys had been removed surgically, the substance was found
largely in the aqueous phases of the blood. The hexamethylenetetramine concentration
increased slowly in the cerebrospinal fluid and more rapidly in the aqueous humour of
the eye. The substance entered the pericardial fluid most rapidly. There was very little
absorption to protein; there was no correlation between the protein level of the fluids and
their maximal hexamethylenetetramine concentrations [10]. The pH value of a 1 N solu-
tion of hexamethylenetetramine in water is 9.5; low pH values promote breakdown. In a
neutral solution at 38 °C, less than 1 % formaldehyde and ammonia are released from
hexamethylenetetramine within 6 hours. Under the conditions of the above study, no free
formaldehyde could be found in the blood of the treated dog [10].
358 Hexamethylenetetramine Volume 5

A study of the kinetics of decomposition of hexamethylenetetramine at 37.5 °C in

citrate-phosphate buffers at pH values between 2 and 7.4 revealed that formaldehyde
release is pH dependent. At pH 5.8 the half-life was 13.8 hours, at pH 2 it was 1.6 hours.
No decomposition was detected within 6 hours at pH 7.4. These results indicate, in
agreement with the observations of Langecker [10], that hexamethylenetetramine is
stable in blood but that at the usual urinary pH values it decomposes rapidly and releases
formaldehyde [11].
Whereas at pH 7 no formaldehyde is detectable, in urine at pH 5.6, peak concen-
trations are found about 2 hours after ingestion of hexamethylenetetramine. The
formaldehyde concentrations vary from 1 to 85 µg/ml, depending on the urinary pH, the
hexamethylenetetramine concentration and the time the urine is retained in the bladder.
At a constant urinary pH of 5.85, hexamethylenetetramine concentrations of 0.7, 1.0 and
1.7 mg/ml result in formaldehyde concentrations of 32, 40 and 60 µg/ml. If the excretion
rate increases from 31 ml urine per hour to 58 ml per hour, the formaldehyde
concentration sinks from 60 to 25 µg/ml [12, 13].
In the human stomach, 10–30% of an oral dose of hexamethylenetetramine is
converted to formaldehyde. Assuming a stomach volume of 0.5 1, a dose of 480 mg
hexamethylenetetramine can produce a local formaldehyde concentration of 123–396
µg/ml [14]. The excretion of hexamethylenetetramine and formaldehyde in the urine
within 48 hours of administration of various preparations (tablets, suspension, granulate,
tablets of hexamethylenetetramine salts with and without acid-resistant coatings) with an
average hexamethylenetetramine content of 480 mg was studied in 10 test persons. The
absorption and excretion rates were higher for the preparations without acid-resistant
coatings (within 3 hours without coating, 7–17 hours with coating). Independent of the
formulation, there were large differences in the amounts of the various preparations
excreted, the recoveries in urine varying between 16 and 83 % of the dose. However,
there were no significant differences between the preparations in the total amounts of
formaldehyde excreted (e.g., 12 hours after administration of hexamethylenetetramine,
the formaldehyde concentration in urine was in the range between 27 and 56 µg/ml) [15].

2 Effects in Man
There are no publications which describe the acute toxicity of ingested or inhaled
hexamethylenetetramine in man.
Hexamethylenetetramine has been used in relatively high doses (2–6 g/day) in the
therapy of acute and chronic urinary tract infections. In up to 7% of the patients who
ingested these doses for weeks at a time, reversible side effects such as gastrointestinal
symptoms (nausea, vomiting, abdominal cramp) and skin reactions (itching, urticaria,
erythematous eruptions) were observed. More rarely, reactions like inflammation of the
oral cavity, anorexia, headaches, respiratory distress and generalized oedema developed.
Very high doses (c 8 g/day) taken for several weeks had adverse effects on the urinary
tract (bladder irritation, albuminuria, haematuria) [3,13,14,16].
Exposure to solid hexamethylenetetramine as well as to the vapour and solutions can
cause irritation of skin and mucous membranes [16, 17] and dermatitis [1, 16].
Volume 5 Hexamethylenetetramine 359

3 Effects on Animals
3.1 Acute toxicity
There are no reports of the acute toxicity of inhaled hexamethylenetetramine.
Ingested hexamethylenetetramine had no toxic effects when fed to mice in doses of
up to 5 g/kg body weight and day for 10 days [18]. No deaths were observed after
administration of an 80 % aqueous solution of hexamethylenetetramine in doses of
10000 or 20000 mg/kg body weight by gavage to Wistar rats [19]. The published LD50
values are shown in Table 1.
The lethal hexamethylenetetramine doses (LDlo and LD50) for mice, rats, guinea pigs
and cats after administration of the substance by subcutaneous injection ranged from
200 to 450 mg/kg body weight [17, 22] (Table 2).
The LD50 after intravenous injection of hexamethylenetetramine into rats was given
as 9200 mg/kg body weight [10, 17, 23]. On the other hand, intravenous injection of a
dose of 10000 mg/kg body weight as an 80% aqueous solution led neither to deaths nor
to any signs of toxicity [19] (Table 2).

Table 1. Acute toxicity of orally administered hexamethylenetetramine

Species No. Administration Dose Duration of Observations Ref.

(strain) route and form mg/kg bw treatment

mouse n.s. oral 512 once LDlo* [20]

mouse n.s. oral 1853 once LD50 (for the [21]
(C3H) formulation
®
Urotovet )
mouse n.s. diet 5000 10 d no toxic effects [18]
mouse n.s. oral > 5000 once LD50* [16]
rat n.s. oral > 5000 once LD50* [16]
rat n.s. oral 9200 once LD50* [1]
rat 5 gavage 10000 once no deaths* [19]
(Wistar) 80% aqueous
solution
rat 5 gavage 20000 once no deaths* [19]
80% aqueous
solution

bw body weight
n.s. not specified
* symptoms of intoxication not described
360 Hexamethylenetetramine Volume 5

Table 2. Acute toxicity of single subcutaneous and intravenous doses of hexamethylenetetramine

Species Number Administration Dose Observations Ref.

(strain) sex route and form mg/kg bw

mouse n.s. subcutaneous 450 lethal dose* [17, 22]

(n.s.)

rat n.s. subcutaneous 200 lethal dose* [17, 22]

(n.s.)

guinea pig n.s. subcutaneous 300 lethal dose* [17, 22]

(n.s.)

cat n.s. subcutaneous 200 lethal dose* [17, 22]

(n.s.)

rat 6 intravenous 9200 LD50* [10, 17, 23]

(n.s.)

rat 14 ♂♀ intravenous 10000 no deaths; [19]

(Wistar) 80% aqueous no signs of
solution toxic effects

bw body weight
n.s. not specified
* symptoms of intoxication not described

On the intact skin of rabbits (groups of 3 male and 3 female Russian rabbits), 0.5 g
hexamethylenetetramine did not cause irritation (4 hour patch test). Similarly, no
irritation was seen after application of 0.1 g to the rabbit eye by instillation into the con-
junctival sac [24, 25].

3.2 Subchronic and chronic toxicity

Long-term inhalation studies with hexamethylenetetramine have not been carried out.
The reported effects of orally administered hexamethylenetetramine are shown in
Table 3.
To test its tolerability after oral intake, hexamethylenetetramine was administered to
BDII rats (2–2.5 months old) as a 40% aqueous solution by gavage at a dose of about 0.4
g/rat/day for 90 days (total dose 28.8 g). None of the treated animals died during the
study nor was their behaviour or development different from that of the controls. The
hair coat of the animals became discoloured yellow, and the discoloration increased with
the duration of treatment [26].
In a 333-day study, 6-week old BDII rats were given daily doses of 0.4 g hexa-
methylenetetramine in 1 ml water by gavage (total dose 94 g). There were no differences
between treated animals and controls in behaviour, development or weight gain during
the course of the study. Autopsy of animals which died or were killed at the end of the
study revealed neither substance-related organ changes nor tumours [26].
 Volume 5
Table 3. Subchronic and chronic toxicity of hexamethylenetetramine after oral or intramuscular administration

Species Number Administration Dose Duration of Observations Ref.

(strain) sex route mg/rat/d treatment

rat 10 gavage 400 90 days no deaths, no deviations from the norm in behaviour [26]
(BDII) or development; yellow discoloration of the fur
rat 15 ♂ gavage 400 333 days no deviations from the norm in behaviour, development [26]
(BDII) 15 ♀ or body weight gain; no effects on organs, no tumours

rat 16 ♂ diet 0.16% for life no deviations from the norm in motor activity, body [27]
(Wistar) 16 ♀ (c 100 mg/kg weight gain, survival, causes of death or relative organ
bw/day) weights; yellow discoloration of the fur
rat 5♂ intramuscular 200 90 days no deaths, no deviations from the norm in behaviour [26]
(BDII) 5♀ or development; yellow discoloration of the fur

bw body weight

Hexamethylenetetramine
361
362 Hexamethylenetetramine Volume 5

Two-month old Wistar rats were given 0.16% hexamethylenetetramine in the diet (c
100 mg/kg body weight per day) from the time of weaning for the rest of their lives.
Motor activity, body weight gain, survival, causes of death and relative organ weights
(liver, kidneys, adrenals and gonads) of the treated animals did not differ from those of
the controls. Autopsy revealed no evidence of treatment-related disease. Occasionally,
yellow discoloration of the hair coat was observed [27].
In a group of 2 to 2.5-month old BDII rats, administration of hexamethylenetetramine
doses of 0.2 g/rat by intramuscular injection, daily for 90 days (total dose 11.8 g) had
no conspicuous effects on behaviour or development. The treatment did not cause any
deaths. Here too, the hair coats of the animals became discoloured yellow [26] (Table 3).

4 Allergenic Effects
4.1 Man
Hexamethylenetetramine causes skin sensitization [1, 28]. Inhalation of hexamethylen-
etetramine can induce an asthma-like reaction in sensitized persons [29].
Exposure to a variety of chemicals led to asthma-like symptoms and finally to a
severe attack of asthma in one worker. Epicutaneous and provocation tests revealed
positive reactions to ethylene diamine and to hexamethylenetetramine [30].
Seven persons employed in the paint industry developed allergic reactions (asthmoid
symptoms, hay fever) after exposure to hexamethylenetetramine. Epicutaneous tests with
1 % solutions of hexamethylenetetramine or ethylene diamine produced toxic reactions
(4 + ) with immediate weal formation. The provocation test produced positive reactions
to both substances in all the patients [30].
In the available literature, however, only one case is described in which a positive
reaction can be ascribed exclusively to hexamethylenetetramine. The patch test (48
hours, occlusive) on a 54-year old foundry worker with allergic contact dermatitis
yielded positive results with hexamethylenetetramine (1 % in petrolatum) after both 48
and 72 hours but negative results with formaldehyde (2% aqueous solution) [31].

4.2 Animals
A sensitizing effect of hexamethylenetetramine was demonstrated in the guinea pig
maximization test (20 female Pirbright White; induction: intradermal 0.1 ml of a 30%
solution, epicutaneous 0.5 g for 48 hours; provocation: epicutaneous 0.2 ml of a 50%
solution) [32].

5 Reproductive and Developmental Toxicity

The results of reproductive toxicity studies carried out with hexamethylenetetramine are
shown in Table 4.
 Volume 5
Table 4. Reproduction and teratogenicity studies with hexamethylenetetramine

Species Number Admini- Concentration Duration of Results, Ref.

(strain) sex stration or dose treatment observations
route

rat P 16 ♂ diet 0.16% 3 months litter size not different from control values [27]
(Wistar) 16 ♀ (100 mg/kg/d) before mating
rat F1 16 ♂ diet 0.16% after weaning no deviation from control values in body weight gain, motor
(Wistar) 16 ♀ (100 mg/kg/d) for life activity, relative organ weights; yellow discoloration of hair coat
rat 9♀ gavage 1 g/kg/d d 7–17 adverse effects on maternal body weight gain; no effects on [33]
(Alpk:AP) post coitum litter size, survival or body weight gain of pups
rat P 1♂ drinking 1% 4 weeks before no deviation from control values in fertility, litter size or [19]
(Wistar) 2♀ water mating and until lactation
after weaning
rat F1 13 ♂ drinking 1% prenatal until 40 no deviation from control values in fertility, litter size,
(Wistar) 6♀ water weeks after birth lactation, body weight or growth
rat F2 15 ♂ drinking 1% prenatal until 40 no deviation from control values in fertility, litter size,
(Wistar) 11 ♀ water weeks after birth lactation, body weight or growth

Hexamethylenetetramine
rat F3 12 ♂ drinking 1% prenatal until 20 no deviation from control values in fertility, litter size,
(Wistar) 12 ♀ water weeks after birth lactation, body weight or growth
rat 6♂ drinking 1% 2 weeks before not teratogenic [34]
(Wistar) 12 ♀ water mating and
until weaning
dog 9♀ diet 600 ppm d 4–56 no adverse effects on number of pregnancies, body weight [35]
(beagle) (15 mg/kg/d) post coitum gain, duration of pregnancy or litter size; not teratogenic
dog 10 ♀ diet 1250 ppm d 4–56 no adverse effects on number of pregnancies, body weight
(beagle) (31 mg/kg/d) post coitum gain, duration of pregnancy or litter size; slight increase in the
number of stillborn pups, slight reduction in body weight gain
and survival of the the pups; not teratogenic

363
P parent generation
364 Hexamethylenetetramine Volume 5

One male and two female Wistar rats were given 1 % hexamethylenetetramine in the
drinking water for 4 weeks before mating. The treatment of the females was then
continued until the two litters had been weaned. The progeny were treated with 1 %
hexamethylenetetramine for 40 weeks and mated in week 26. The F2-generation was
treated similarly and mated at the age of 15 weeks. The F3-generation was also given 1
% hexamethylenetetramine. There were no differences between the treated animals and
the corresponding controls in fertility, litter size, lactation, body weight or growth [19].
Wistar rats (6 males and 12 females) were given 1 % hexamethylenetetramine in the
drinking water for 2 weeks before mating. The treatment of the females was then
continued during pregnancy and lactation. All females littered normally. There were no
malformations in the progeny [34].
Two-month old Wistar rats were given 0.16 % hexamethylenetetramine in the diet (c
100 mg/kg body weight and day) for 3 months before mating. There were no differences
in litter size between the control and treated animals. The progeny, which were treated
like the parents after weaning, did not differ from the controls in body weight gain or
motor activity. The relative organ weights (liver, kidneys, adrenals and gonads) were not
different in treated and untreated pups. Occasionally, the treatment caused yellow
discoloration of the hair coat [27].
Female beagle dogs (weighing 12 kg) were given a diet containing 600 or 1250 ppm
hexamethylenetetramine (c 15 and 31 mg/kg body weight and day) from day 4 to day 56
after mating. No treatment-related effects on the number of pregnancies, body weight
gain of the pregnant animals, duration of pregnancy or litter size were found. In the high
dose group, the number of stillborn pups was slightly increased (because of one litter in
which only 2 of 9 pups were born alive) and there were slight adverse effects on body
weight gain of the pups and on their survival until weaning. Organ or skeletal
malformations were not observed [35].
In a modified Chernoff-Kavlock assay for teratogenicity, hexamethylenetetramine
was classified as not teratogenic. In Alpk:AP rats (Wistar-derived), a hexa-
methylenetetramine dose of 1 g/kg body weight and day administered by gavage from
day 7 to day 17 of gestation had adverse effects on the body weight gain of the dams
(maternal toxicity); however, no effects on litter size, survival or body weight gain of the
pups were detected within the 5 days after birth [33].

6 Genotoxicity
6.1 In vitro studies
The results of the published in vitro genotoxicity studies with hexamethylenetetramine
are shown in Table 5.
In the Ames test in the Salmonella typhimurium strains TA1535, TA1537, TA1538,
TA98 and TA100 with and without a metabolic activation system (S9 mix from Aroclor
1254-induced rat liver), a hexamethylenetetramine concentration of 1000 µg/plate was
not mutagenic. However, positive results were obtained with the nitrosation product ob-
tained in the reaction of hexamethylenetetramine with nitrite in acetic acid solution [36].
Volume 5 Hexamethylenetetramine 365

Table 5. In vitro genotoxicity studies with hexamethylenetetramine

Test organism Concentration Results Ref.

µg/plate or M

S. typhimurium 1000 negative [36]

TA1535, TA1537,
TA1538, TA98, TA100
S. typhimurium 1000 with nitrosation product positive [36]
TA98 + NaNO2 in
acetic acid
S. typhimurium 200 negative [37]
TA98, TA100 1000 negative
5000 negative
S. typhimurium n.s. with nitrosation product positive [37]
TA98, TA100 + NaNO2 at
low pH
S. typhimurium n.s. negative, questionable [38]
TA1535, TA1537,
TA1538, TA98, TA100
S. typhimurium 10000 positive [38]
TA98, TA100 15000 positive
E. coli 5000 negative [39]
WP2uvrA
E. coli 500 negative [40]
P3478(pol A–-) 6000 positive
Bacillus subtilis 1000 positive [41]
H17 and M45
mouse lymphoma cells n.s. negative [42]
L5178Y TK+ / –
HeLa cells 1.0 × 10–3 M increase in chromosomal aberrations [43]
1.4 × 10–2 very cytotoxic,
to 7.0 × 10–2 M inhibition of growth and mitosis, nuclear
changes, chromosomal clumping

human leukocytes 1.8 × 10–5 M no growth inhibition, no chromatid or [43]

in culture chromosomal aberrations
1.4 × 10–2 very cytotoxic, inhibition of mitosis
to 7.0 × 10–2 M

n.s. not specified

Under similar test conditions (S. typhimurium strains TA98 and TA100 with and
without S9 mix from Aroclor 1254-induced rat liver), hexamethylenetetramine con-
centrations up to 5000 µg/plate were not mutagenic. Again, the product of the nitrosation
reaction with sodium nitrite at low pH was shown to have mutagenic activity [37].
366 Hexamethylenetetramine Volume 5

Negative results were also obtained in another study with the S. typhimurium strains
TA1535, TA1537, TA1538, TA98 and TA100 (concentrations not specified) with and
without S9 mix from Aroclor 1254-induced rat liver; in this case [38] it was considered
that poor solubility and inadequate penetration of the bacterial membrane could have
been responsible for the lack of effect.
In preincubation tests with the S. typhimurium strains TA1535, TA1537, TA1538,
TA98 and TA100 and Escherichia coli WP2uvrA with and without metabolic activation
(S9 mix from PCB-induced rat liver) a weak mutagenic effect was seen only in the
strains TA98 and TA100 in the concentration range above 10000 µg/plate and up to
15000 µg/plate [39].
In a DNA repair test in E. coli P3478 (pol A-) with a hexamethylenetetramine
concentration of 6 mg/plate, a positive result was obtained and ascribed to released
formaldehyde [40].
In the Rec assay with the Bacillus subtilis strains H17 and M45 both with and without
metabolic activation (S9 mix from Aroclor 1254-induced rat liver), 1 mg/ plate caused
DNA damage which was accounted for in terms of the electrophilic properties of
hexamethylenetetramine [41].
In a modified mouse lymphoma test (L5178Y TK+/–) in which the enzyme formalde-
hyde dehydrogenase and its cofactor NAD+ were added to the substrate during exposure
of the cells, no mutagenic effect was observed. With this modified test, it could be
demonstrated that hexamethylenetetramine is not mutagenic per se and that mutagenic
effects are only produced after hydrolytic release of formaldehyde [42].
High concentrations of hexamethylenetetramine (1.4 × 10–2 M to 7.0 × 10–2 M) are
very cytotoxic for cultured leukocytes and HeLa cells (inhibition of growth and mitosis,
nuclear changes, chromosome clumping). A concentration of 1.8 × 10–5 M caused
neither growth inhibition nor chromatid or chromosomal aberrations in cultured
leukocytes. In cultured HeLa cells, an increase in chromosomal aberrations above the
control values was seen at concentrations of 1×10–3 M or more [43].

6.2 In vivo studies

The results of the published in vivo genotoxicity studies with hexamethylenetetramine
are shown in Table 6.
After intraperitoneal injection of hexamethylenetetramine doses of 800–10000 mg/kg
body weight into male C3H or (C3H × 101)F1 mice and mating with females of the same
strain, the incidence of dominant lethal mutations was not different from that in the
controls. Oral doses of at least 25000 mg/kg were required to increase the incidence of
dead implants in C3H mice above control values. Evidence of a relatively weak
mutagenic effect on the germ cells of male mice was thus only obtained with sublethal
hexamethylenetetramine doses [43, 44].
In a study of the induction of chromosomal aberrations in bone marrow, male C3H
mice were given the preparation Urotovet® dissolved in water in oral doses of
Volume 5 Hexamethylenetetramine 367

Table 6. In vivo genotoxicity studies with hexamethylenetetramine

Test system Route Dose Duration of Results and Ref.

and organism mg/kg bw treatment Observations

dominant lethal test

mouse (♂) i.p. 8000–10000 once no induction [43, 44]
C3H of lethal mutations
(C3H×101)F1
mouse (♂) oral 25000 once increased number [44]
C3H of dead implants
test for chromosomal aberrations
mouse (♂) oral 1
, 91 , 271 once no clastogenic effects [21]
3
C3H
of the LD50*
mouse (♂) oral 1 1
, , 271 5d no clastogenic effects [21]
3 9
C3H
of the LD50*

bw body weight
* refers to an LD50 of 1853 mg/kg (Table 1 [21])

1
/3, 1/9 and 1/27 of the LD50 (LD50 given as 1853 mg/kg body weight) either once or, in
another series of experiments, on 5 consecutive days. Clastogenic effects were not
detectable in either experimental series [21].

6.3 Cell transformation

In a cell transformation test with BHK-21/cl.13 cells, hexamethylenetetramine caused a
dose-dependent increase in transformation frequency in the concentration range between
1 and 10000 µg/ml in the presence of a metabolic activation system (S9 mix from
Aroclor 1254-induced rat liver). The toxic and transforming effects of a hexamethylene-
tetramine concentration of 1000 µg/ml were comparable with those of a 20 µg/ml form-
aldehyde solution [45].

7 Carcinogenicity
The results of the carcinogenicity studies which have been carried out with hexa-
methylenetetramine are shown in Table 7. There are no long-term inhalation studies.
Local sarcomas were induced in 8 of 14 rats (57%) by repeated subcutaneous
injection of 35–40% hexamethylenetetramine solutions (total dose 25–30 g/animal) [46].
Ten-day old CTM mice and Wistar rats were given five hexamethylenetetramine
doses, each of 5 g/kg body weight as a 30 % aqueous solution (total dose 25 g/kg) by
subcutaneous injection on alternate days and were then observed for the rest of their
lives. Evidence for a carcinogenic effect was not obtained in either species [47].
 368
Table 7. Carcinogenicity studies with hexamethylenetetramine

Species Number Administration Concentration Duration of Results and Ref.

(strain) sex route or dose treatment/observation observations

Hexamethylenetetramine
rat 14 subcutaneous 35–40% solution (total repeatedly 8/14 rats (57%) with local [46]
(n.s.) dose 25–30 g/rat) sarcomas

mouse 39 ♂ subcutaneous 5 g/kg bw 5 d/life not carcinogenic [47]

(CTM) 44 ♀

rat 20 ♂ subcutaneous 5 g/kg bw 5 d/life not carcinogenic [47]

(Wistar) 20 ♀
rat 15 ♂ drinking water 0.1 % (total dose 5 g/rat) 50 weeks/life not carcinogenic [48]
(Sprague 15 ♀
Dawley)

rat 15 ♂ drinking water 0.1% + 0.2% NaNO2 50 weeks/life not carcinogenic [48]
(Sprague 15 ♀
Dawley)
rat (Wistar) 16 ♂ diet 0.16% (100 mg/kg/day) life not carcinogenic [27]
16 ♀
mouse 50 ♂ drinking water 0.5% (1.25 g/kg/day) 60 weeks/life not carcinogenic [47]
(CTM) 50 ♀

mouse 96 ♂ drinking water 1% (2.5 g/kg/day) 60 weeks/life not carcinogenic [47]

(CTM) 102 ♀

mouse 30 ♂ drinking water 1% (2.5 g/kg/day) 60 weeks/life not carcinogenic [47]

(C3Hf) 63 ♀

Volume 5
mouse 29 ♂ drinking water 1 % (2.5 g/kg/day) 60 weeks/life not carcinogenic [47]
(SWR) 27 ♀
 Volume 5
Table 7 (continued)

Species Number Administration Concentration Duration of Results and Ref.

(strain) sex route or dose treatment/observation observations

mouse 29 ♂ drinking water 5% (12.5 g/kg/day) 30 weeks/life slight reduction in growth and [47]
(CTM) 50 ♀ survival; not carcinogenic

rat 48 ♂ drinking water 1% (2.0–1.5 104 weeks/life not carcinogenic [47]

(Wistar) 48 ♀ or 2.5–2.0 g/kg/day)

rat 12 ♂ drinking water 5% 2 weeks/life 50% mortality; not carcinogenic [47]

(Wistar) 12 ♀

rat F1 24 ♂ drinking water 1% prenatal until 20 weeks no deviation from the norm in [34]
(Wistar) 24 ♀ p.p./>2 years organ weights, gross pathological
or microscopic changes; no
transplacental carcinogenesis

rat F1 13 ♂ drinking water 1% prenatal until 40 weeks no transplacental carcinogenesis [34]

(Wistar) 7♀ p.p./>2 years

Hexamethylenetetramine
rat F2 15 ♂ drinking water 1% prenatal until 40 weeks no transplacental carcinogenesis [34]
(Wistar) 11 ♀ p.p./>2 years

rat F3 12 ♂ drinking water 1% prenatal until 20 weeks no transplacental carcinogenesis [34]

(Wistar) 12 ♀ p.p./>2 years

rat F1 16 ♂ drinking water 2% prenatal until 50 weeks no transplacental carcinogenesis [34]

(Wistar) 16 ♀ p.p./>2 years

bw body weight
n.s. not specified

369
p.p. post partum
370 Hexamethylenetetramine Volume 5

Hexamethylenetetramine was administered for 50 weeks to Sprague-Dawley rats (8–

10 weeks old) at a concentration of 0.1 % in the drinking water, to one group on its own
(total dose 5 g/rat) and to a second group together with 0.2% sodium nitrite. The animals
were then observed for the rest of their lives. Tumours were not induced by
hexamethylenetetramine either alone or in combination with nitrite [48].
Likewise, when Wistar rats were given 0.16% hexamethylenetetramine in the diet (c
100 mg/kg body weight and day) for the whole of their lives (see Section 3.2 above), no
treatment-related tumours were found [27].
In long-term studies, groups of CTM mice were given hexamethylenetetramine in the
drinking water at concentrations of 0.5% and 1 %, C3Hf and SWR mice at 1 % for 60
weeks and CTM mice at 5 % for 30 weeks. All the mice were then observed for the rest
of their lives. In the mice treated with 5 % hexamethylenetetramine in the drinking
water, there were slight reductions in growth and survival. Slight delays in growth were
also seen in the SWR mice. Gross pathological and histological examination of animals
which died during the study or were killed at the end revealed no signs of treatment-
related pathological or tumorigenic changes [47].
In similar studies, Wistar rats were given hexamethylenetetramine in the drinking
water at a concentration of 1 % for 104 weeks or 5 % for 2 weeks. Half of the animals in
the 5 % group died during the course of the study; treatment-related pathological
changes were, however, not detected. The survivors recovered rapidly and developed no
symptoms of disease during the remainder of the study. Thus in the rats as well, there
were no gross pathological or microscopic changes or tumours which could be ascribed
to the treatment with hexamethylenetetramine [47].
Wistar rats were treated for two weeks before mating and the females also during
pregnancy and lactation with 1 % hexamethylenetetramine in the drinking water; the
progeny were also treated with 1 % hexamethylenetetramine for 20 weeks. No devi-
ations from the control values in organ weights were detected and no transplacental
carcinogenicity [34].
In other studies, Wistar rats were given 1 % hexamethylenetetramine in the drinking
water for 40 weeks in the F1 and F2 generations and for 20 weeks in the F3 generation.
The progeny from a group of parent animals treated with 2% hexamethylenetetramine
were also treated with 2% hexamethylenetetramine in the drinking water for 50 weeks.
All the animals were observed for the rest of their natural lives. No evidence for
transplacental carcinogenicity of hexamethylenetetramine was obtained in any of the
groups [34].

8 Manifesto (MAK value, classification)

Hexamethylenetetramine as the solid, in solution and as vapour irritates human skin and
mucous membranes. Animal studies and one human case suggest that it can cause
sensitization.
The data published to date provide no evidence of systemic effects of hexa-
methylenetetramine in man. The results of animal studies indicate that organ damage is
Volume 5 Hexamethylenetetramine 371

not to be expected in persons exposed to hexamethylenetetramine at work, especaily

because of the rapid elimination of the substance under the usual workplace conditions.
The animal studies also provide no evidence that teratogenic, mutagenic or car-
cinogenic effects may be expected in man.
Neither the known effects in man nor the data from animal studies are sufficient for
the establishment of a MAK value. Therefore hexamethylenetetramine can only be
included in Section IIb of the "List of MAK and BAT Values".

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completed 16.5.1991