CYHEXATIN
EXPLANATION
Cyhexatin was reviewed by the JMPR in 1970, 1973, 1974, 1975,
1977, 1978, 1979, 1982, 1983 and 1985 (Annex 1, FAO/WHO 1971a,
1974a, 1975a, 1976a, 1978a, 1979a, 1980a, 1983a, 1984, 1986a). An
ADI of 0.008 mg/kg bw was allocated in 1980. A teratology study in
the New Zealand White rabbits at dose levels of 0.75 and 3 mg/kg
bw/day and a multi-generation study in rats (top dose level 100 ppm)
considered in 1970 did not demonstrate biological effects. Since
the evaluation in 1985, cyhexatin has been withdrawn from use as an
acaricide in a number of countries as a result of new teratology
studies. The 1988 meeting was requested to evaluate these studies
but insufficient data were available at that time. These data and
other teratology studies have now been submitted and reviewed.
Short-term studies
Rabbits
Groups of 4 virgin female hybrid Hy/Cr White New Zealand
rabbits were given doses of 96% pure cyhexatin (technical) of 0,
0.5, 1.0 or 3.0 mg/kg bw/day by gavage for 13 days. Dosing volume
was 5 ml/kg bw of a suspension of the test material in a 0.5 aqueous
solution of carboxymethyl cellulose. Suspensions were prepared
weekly. The rabbits were housed individually and monitored daily for
condition and behaviour; body weight and food consumption were
recorded weekly. On day 13 blood samples were taken two hours
post-dosing and fecal samples were obtained. On Day 14 the animals
were sacrificed and necropsied and samples of liver, kidney and
ovary saved for analysis.
There were no effects noted on appearance, behaviour or gross
pathology. Body weights, liver, kidney and ovary weights and food
consumption were similar in all groups. Tissues analyses results
indicated dose-related increases in blood, liver, kidney and fecal
tin concentrations. No effect was observed in ovary (Monnot et al.
1989a).
Groups of 4 virgin female rabbits of the same strain as in the
oral study were given daily dermal doses of 0, 0.5, 12.0 or 3.0 mg
cyhexatin (96%)/kg bw for 13 days. Dose volume was 0.25 ml/kg bw of
suspensions in 0.5 aqueous carboxymethyl cellulose solutions. The
suspensions were applied to the shaved back of each rabbit at sites
which were not treated more than twice without an interval of at
least seven days. At application the site was given a slight
massage for about one minute. The rabbits were monitored for
clinical conditions, effects at application site, body weights and
food consumption. Blood, fecal and tissue samples were taken
similarly to the oral study.
There were no effects on appearance, behaviour or gross
pathology. There was a dose-related increase in the incidence of
erythema at all dose levels. Desquamation and atonia were observed
at all treated sites with no dose-relationship but with a slight
increase with time. "Cracking" of the skin was observed in one
rabbit at 1 mg/kg bw and 3 at 3 mg/kg bw. At 3 mg/kg bw no increase
in body weight was observed in contrast to weight gains in controls
and the rabbits in the other two groups with the exception of one
rabbit in the 0.5 mg/kg bw group which lost weight; however, this
rabbit was stated to have been delivered on Day 9 of the treatment
and, thus, was not comparable to the rest of the animals. Food
consumption was slightly lower in the second week in 2 rabbits at
the 3 mg/kg bw level. No effects were noted on liver, kidney or
ovary weights. Tissue tin levels tended to be variable between
animals. Mean levels of tin were increased with dose in liver,
feces and, very slightly, in blood. Fecal levels were higher in
this study than in the preceding oral study (Monnot et al. 1989b).
Special studies on teratology
Oral studies
Rat
Groups of 30 female Sprague-Dawley derived Charles River COBS
CD rats (Portage, Michigan, USA) were mated (1:1 basis) at 84 days
of age. The day a copulatory plug was observed was designated
Gestation Day 0. On Days 6-15 of gestation the females were given
doses of cyhexatin (95.6% pure) of 0, 0.5, 1.0 or 5.0 mg/kg bw/day
by gavage. The test material was suspended in corn oil for
administration with the dose level adjusted to correct for purity.
The dosing suspensions were prepared once for the study and agitated
with a magnetic stirrer during dose administrations. Dose volume
was 2.0 ml/kg. The animals were monitored during the study for
signs of toxicity, body weight, food and water consumption. On Day
20 of gestation the females were sacrificed and the uterus and ovary
examined. Other organs were examined for gross pathology. Fetuses
were weighed, sexed and examined for external malformations. Half
the fetuses were examined for visceral malformations and the other
half for skeletal malformations.
There were no clinical signs of toxicity and no treatment-
related gross pathology. During dosing the 5 mg/kg bw group had
slightly lower body weight gains and food consumption but overall
weight gain did not differ between groups and the differences were
not considered to be biologically significant. There were no
treatment-related effects on water consumption, number of fetuses
and viable fetuses per dam, number of resorptions (all early), or
fetal body weight or sex ratio. Absolute and relative liver weights
and mean gravid uterus weight were higher in the 5 mg/kg bw group.
The total incidence of malformations was 4(3), 3(2), 2(2) and 6(6)
fetuses (litters) at 0, 0.5, 1.0 and 5.0 mg/kg bw, respectively.
Microphthalmia was observed only in treated animals in 1, 2, and
2(2) fetuses at 0.5, 1.0 and 5.0 mg/kg bw, respectively. This
malformation was not listed in the submitted historical control data
for 35 studies. Tail anomalies (short; short and thread-like; or
short and bent) were observed in 1, 0, 1 and 3(3) fetuses (litters)
at 0, 0.5, 1.0 and 5.0 mg/kg bw, respectively. The other malformed
fetus at 5 mg/kg bw had fused skull bones. The mid- and low dose
fetuses with microphthalmia both had (multiple defects including
cleft palate (both), cleft lip (0.5 mg/kg bw), micrognathia
(1.0 mg/kg bw) and malformed skull bones (1.0 mg/kg bw). These
other anomalies were not observed at 5.0 mg/kg bw. Visceral
anomalies were observed only in control and low dose fetuses.
Development variations were observed in some fetuses in each group;
however, the occurrence of 25 or 27 pre sacral vertebrae was
observed only at 5.0 mg/kg bw in 6(6) fetuses (litters). Although
the incidence of malformations was slightly higher in the high dose
group, it cannot be considered to be indicative of a teratogenic
effect. However, the vertebral variations of 5.0 mg/kg bw suggest a
fetototoxic effect. The NOAEL for this study is considered to be
1.0 mg/kg bw (Aldridge et al. 1986).
Rabbit
In a teratology probe study, groups of 7 female New Zealand
White rabbits (Hazleton-Dutchland) was artificially inseminated and
then dosed with 0, 5, 10 or 20 mg cyhexatin/kg bw/day on Days 6-18
of gestation (Day of insemination = Day 0). The test material was
suspended in corn oil at concentrations to provide the appropriate
doses in 1 ml/kg bw and administered with a 4-inch 12 gauge dosing
needle. One analysis of each solution is stated to indicate
concentrations of 99-103% of target. There were 2, 7 and 7 deaths
at 5, 10 and 29 mg/kg bw, respectively, during the dosing period.
Autopsy revealed lesions in the trachea and lungs of all animals
that died suggesting that improper dosing technique contributed to
the deaths. Two animals each at 10 and 20 mg/kg and all 5 surviving
animals at 5 mg/kg bw had stomach erosions or ulcers. Four of the 5
surviving animals at 5 mg/kg bw had only resorptions (Berdasco
et al. 1986).
A second probe was carried out in the same strain of rabbits
prepared for treatment as above (7 rabbits/dose level) but dosed
with 0, 1, 5 or 10 mg cyhexatin/kg bw/day on Days 7-19 of gestation.
In this study the test material was suspended in 0.5% aqueous
Methocel at concentrations to provide the appropriate doses in
1 ml/kg bw and administered with a # 10 juvenile Foley catheter. One
analysis of each solution indicated concentrations of 80-101% of
target. Again, deaths were observed only in treated groups with 2,
1 and 1 deaths at 1, 5 and 10 mg/kg bw. Tracheal and lung lesions
were not seen in this study. Stomach erosion/ulcers were observed
in one animal at 5 mg/kg bw and 4 at 10 mg/kg bw. Peritoneal blood
was observed in 4 females at 10 mg/kg bw and was considered to be
evidence of recent abortion. Body weight gains were reduced at both
5 and 10 mg/kg bw/day. At 10 mg/kg bw, 4 females totally resorbed
and/or aborted their fetuses. Both of the other surviving females
had resorptions and the number of viable fetuses was significantly
reduced compared to controls. At 5 mg/kg bw, there was an increased
incidence of resorptions, with one litter totally resorbed, and a
decreased number of fetuses. The 1 mg/kg bw group was similar to
controls (Berdasco et al. 1986).
As a result of the above probe studies, the teratology study
was conducted in groups of 20 females of the same strain of rabbit
(5 months old at insemination) with dose levels of 0, 0.5, 1.0 and
3.0 mg cyhexatin (95.5%)/kg bw/day given Days 7-19 of gestation (Day
of artificial insemination = Day 0 of gestation) as a suspension in
0.5% aqueous Methocel (1 ml/kg bw) and administered with 5 cc
plastic syringes and 16 inch long flexible feeding tubes. The
suspensions of test material were prepared once (concentration
adjusted for purity of test material) and resuspended daily with a
Brinkman polytron homogenizer and maintained in suspension during
treatment with a magnetic stirrer. Analyses of the suspensions at
intervals during the study indicated sample concentrations of
70-104%, 91-125% and 55-137% of target at the 0.5, 1.0 and 3.0 mg/kg
bw dose levels, suggesting that homogeneity was difficult to attain
especially at the highest concentration. The homogeneity problems
may have related to the particle size of the cyhexatin used in the
suspension but no data were available for this parameter. The
rabbits were housed individually in suspended wire cages and
maintained on Purina Certified Rabbit Chow # 5322 and tap water
available ad libitum. The rabbits were monitored throughout the
study for clinical condition and body weight. Females which died or
which aborted or delivered prematurely were autopsied. On Day 29,
all surviving females were sacrificed and the uterus and ovary of
each examined. Fetuses were weighed, sexed and examined for
external, visceral and skeletal malformations and variations.
There were no treatment-related effects on mortality (1-4
females/group died, all but one of which (low dose) animals showed
red fluid in the thoracic cavity which may indicate injury during
dosing), or general signs of toxicity. Five rabbits aborted: one at
0.5 mg/kg bw and 4 at 3.0 mg/kg bw; two rabbits delivered on Day 28:
one each at 1.9 and 3.0 mg/kg bw; and two rabbits totally resorbed
their litters: one each at 0.5 and 3.0 mg/kg bw. Body weight
changes were extremely variable in all groups and did not appear to
be related to treatment. Food intakes were not measured. Liver to
body weight ratios were not dose-related and there were no gross
pathological lesions indicative of a treatment-related effect. The
number of viable fetuses per dam was reduced in the 3.0 mg/kg bw
group. The number of dams with resorptions did not differ between
groups but the number of dams with two or more resorptions was
increased at 3.0 mg/kg bw. Total percent post-implanatation loss
was increased in a dose-related manner and was 2x and 3x the control
value of 1.0 and 3.0 mg/kg bw, respectively. Fetal body weights and
sex ratio were not affected by treatment. A few fetuses in each
group had malformations but the only observation which appeared to
be dose-related was hydrocephalus in 8(4) fetuses (litters) at
3.0 mg/kg bw. In this group, one aborted fetus had a dome-shaped
head but was not examined viscerally. The variability of results
and lack of dose relationships may have been related either to the
questionable homogeneity of the test or to maternal infection.
There was no consistent evidence of maternal toxicity in this study.
The NOAEL for this study was 0.5 mg/based on increased
post-implantation loss at the nominal 1.0 mg/kg bw level (Schardein
et al. 1986).
To confirm the results of the above study, another study was
conducted in a different laboratory with groups of 27 artificially
inseminated New Zealand White (Hazleton-Dutchland) female rabbits.
The test material, technical cyhexatin of 94.8-95.5% purity, was
suspended in 0.5 aqueous Methocel at concentrations to provide doses
of 0.75 or 3.0 mg/kg bw in a volume of 1 ml/kg bw. Again, no data
were available on particle size of the cyhexatin. No analyses of
administered suspensions were reported. Controls received 0.5%
aqueous Methocel at the same volume. The suspensions were prepared
once and were maintained with continuous magnetic stirring
throughout the study (Hanley, 1989). Analyses pre- and post-test
indicate concentrations were 89-106% of target. The suspensions of
cyhexatin were given by oral gavage Days 7 through 19 of gestation
(Day of insemination = Day 0 of gestation). The rabbits were housed
singly and maintained on Purina Rabbit Chow (8 oz/day) and tap water
(ad libitum). The type of cages and the room conditions are not
described. Body weights and clinical condition of the rabbits were
monitored throughout the study. On Day 28 of gestation, Cesarian
sections were performed on all rabbits and the uterus of each was
weighted and examined. Fetuses were weighed, sexed and examined for
viability and external and visceral malformations.
There were 2, 7 and 4 deaths at 0, 0.75 and 3.0 mg/kg bw,
respectively, of which 2, 5 and 2 rabbits had lung or thoracic
cavity lesions suggesting possible problems during dosing.
Abortions occurred in three rabbits at 0.75 mg/kg bw (one dam died
after aborting one fetus) and in 12 rabbits at 3.0 mg/kg bw. Among
the animals that aborted at 3.0 mg/kg bw, 5 had hairballs in the
stomach and one had slight abdominal ascites. No significant
pathology was observed in the other rabbits that aborted. The
rabbits with litters were not examined for gross pathology.
Individual body weight changes were quite variable but in the
3.0 mg/kg bw most of the rabbits lost weight Days 7-10 (20/23 cf
8/23 and 12/25 at 0 and 0.75 mg/kg bw, respectively). During the
remainder of the dosing period the number of females losing weight
was slightly higher than in the other groups and the mean body
weight change was lower than in controls. At Day 20, mean body
weight was statistically significantly lower in the 3.0 mg/kg bw
group than in the controls. This was considered to be a treatment-
related effect. Body weight changes in the 0.75 mg/kg bw group were
not considered to be different than the controls. Absolute and
relative liver weights did not differ between the groups but there
were only seven animals remaining at termination in the 3.0 mg/kg bw
group. There was an increased incidence of resorption and a
decreased number of live fetuses in the 3.0 mg/kg bw group.
However, with only seven dams in this group, the significance of
these observations is not certain. There were no apparent effects on
fetal weights or sex ratio. The fetuses were examined only for
external and visceral malformations and the total incidences of any
malformation were 3(3), 10(7) and 11(5) fetuses (litters) at 0, 0.75
and 3.9 mg/kg bw, respectively. The most common malformation was
hydrocephalus with incidences of 2(2), 7(5) and 9(4) fetuses
(litters), respectively. Dilated cerebral ventricles were observed
in one fetus at 0.75 and 3.0 mg/kg bw in litters with no
hydrocephalic fetuses. Incidences of malformations in control
groups from 48 studies (mainly oral but some inhalation indicate
that hydrocephalus was observed in only one of these studies (#43)
in one fetus (of 91) (overall incidence 1/2936 fetuses and 1/839
litters). Dilated cerebral ventricles were observed in a total of 3
fetuses from 2 litters (2 fetuses in study #11 and one in Study
#16). The incidence of this malformation is higher in the
concurrent controls and is increased further in the treated groups.
At 3.0 mg/kg bw there was evidence of a maternal effect with reduced
body weight gain during the dosing period and a high number of
abortions. There was no clear indication of maternal toxicity at
0.75 mg/kg bw which may be considered to be the NOEL for maternal
toxicity. A NOAEL for the induction of hydrocephalus was not
demonstrated in this study (Kirk et al. 1989a).
Two studies using cyhexatin from another source were carried
out in a different laboratory. Hybrid HY/Cr White New Zealand type
rabbits (Charles River, France, 16-18 weeks old) were mated by the
supplier and delivered to the laboratory on Gestation Day 1 or 2
(Day of mating was considered Day 0 of gestation). Groups of 24
mated females were given doses of 0, 3.0 (technical) or 3.0 (pure)
mg/kg bw/day in the first study and 0, 0.5, 0.75 or 1.0 mg
(technical)/kg bw/day in the second study on Days 6-18 of gestation.
Purity of the technical test material was 96% and no adjustment was
made for purity when the dosing suspensions were prepared weekly in
0.5% aqueous carboxymethyl cellulose at concentrations to provide
the appropriate doses in a volume of 5 ml/kg bw. The suspensions
were stated to have been "under constant agitation before and during
daily dosing". No analyses for homogeneity were performed.
Particle size of the cyhexatin was 20-50 microns for technical
material and 20-60 microns for the pure material prior to
suspension. The animals were dosed by intragastric intubation
(details of method not given). The rabbits were housed individually
in plastic cages and were given access to 200 g of pelleted rabbit
diet (UAR-112-Usine d'Alimentation Rationnelle)/day. Tap water was
available ad libitum. The animals were monitored for clinical
signs, body weight and food consumption throughout the study.
Animals which died or aborted were autopsied. All surviving females
were sacrificed on Day 29 and the ovary and uterine contents
examined. The number, sex, and body weights of fetuses were
recorded and the fetuses were examined for external visceral and
skeletal malformations. The heads of half the fetuses were removed
for sectioning.
There were 1-3 deaths in each group of rabbits (except the
group given pure cyhexatin in which there were no deaths) but the
incidence was not dose-or treatment-related. In the first study
three rabbits aborted in the group given technical material
(3.0 mg/kg bw) but none in the controls or those given pure
cyhexatin. In the second study abortions were observed only in the
controls (3) and 0.5 mg/kg bw (2) groups. None of the rabbits
delivered prematurely and none resorbed their entire litter. Body
weights and food consumptions were somewhat variable but differences
did not appear to be related to treatment. There were no apparent
treatment-related effects on numbers of viable fetuses or in the
numbers of resorptions. The group given 3.0 mg/kg bw of pure
cyhexatin had slightly fewer viable fetuses than the other groups as
a result of fewer corpora lutea and implantations. Mean fetal
weight and sex ratio were not affected by treatment. In each of the
two studies, one fetus was observed with a dome-shaped head
associated with dilated ventricles of the brain; in the first study
the fetus was in the 3.0 mg/kg bw technical cyhexatin group, in the
second the fetus was in the 0.5 mg/kg bw group. One fetus given
pure cyhexatin at 3.0 mg/kg bw also had dilated ventricles of the
brain but without a dome-shaped head. The total incidence of major
malformations was 1, 2 and 2(2) fetuses (litters) at 0, 3.0
(technical) and 3.0 (pure) mg/kg bw/day in the first study and 1(1),
3(3), 4(4) and 3(3) fetuses (litters) at 0, 0.5, 0.75 and 1.0 mg/kg
bw/day in the second study. Retinal detachment was observed in 2, 3
and 1 fetus at 0.5, 0.75 and 1.0 mg/kg bw but was not observed in
controls or the 3.0 mg/kg bw groups.
In these studies there were no apparent treatment-related
maternal toxicity, embryo/fetotoxicity, or teratogenic effects. The
NOAEL was the highest dose tested: 3.0 mg/kg bw (Monnot, 1989a & b).
Dermal studies
Rabbits
Groups of 16 artificially inseminated New Zealand White rabbits
(Hazleton-Dutchland, USA) were given doses of 0, 0.5, 1.0 or 3.0 mg
cyhexatin (94.8-95.5% pure)/kg bw/day dermally on Days 7-19 of
gestation (Day of insemination = Day 0 of gestation). The test
material was suspended in 0.5% aqueous Methocel at concentrations to
provide the appropriate dose in a volume of 1 ml/kg bw. The
suspensions were prepared once and kept with constant agitation
throughout the dosing period. The test substance was applied
uniformly to a clipped ear (about 10 x 15 cm) on the back of each
animal and covered with an occulusive bandage and a lycra/spandex
jacket for a 6-hour period. The rabbits were housed individually
and given 8 oz of Ralston Purina rabbit chow daily. Tap water was
available ad libitum. The animals were monitored throughout the
study for clinical signs of toxicity, local effects of treatment,
body weights and food consumption. Cesarian section was performed
on Day 28 of gestation and the ovary and uterine contents were
examined. Maternal and liver and uterine weights were recorded and
liver and skin samples retained for histopathological examination.
Fetuses were examined for viability, body weight, sex and external
and visceral malformations. The fetuses were not examined for
skeletal effects.
In this study, dosing suspensions had mean concentrations of
75.6 to 114.2% of the target. The low mean was at the highest
concentration in the post-dosing analyses. There was a dose-related
increase in lesions at the site of application (erythema/eschar,
edema and fissuring/scaling) which increased with dosing. Fissuring
was observed in 0, 6, 9 and 16 rabbits at 0, 0.5, 1.0, and
3.0 mg/kg bw, respectively, with maximum mean scores (24 hours after
application) of 0, 1.7, 2.3 and 2.9, respectively (scale 0-4).
There were no clinical signs of general toxicity noted. One rabbit
at 0.5 mg/kg bw and two at 3.9 mg/kg bw delivered their litters
prior to Cesarean section (Days 27, and Days 26 and 27,
respectively). None of the rabbits aborted. There were no apparent
treatment-related effects on maternal body weight gains or on liver
weights. Histopathological examination of the treated skin of
rabbits given 3.0 mg/kg bw showed slight to moderate acanthosis and
generally slight hyperkeratosis with very slight to slight
inflammation. There were no treatment-related effects on the number
of fetuses or resorptions, fetal sex ratio or fetal body weights.
One dam in each of the control and 3.0 mg/kg bw groups had a
hairball in the stomach, lost considerable weight during gestation
and had fetuses with low body weights (litter means of 13.6 g and
21.5 g, respectively). Both of these females had runts and fetuses
with malformations. The total incidence of any malformation was
8(4), 1(1), 1(1) and 8(5) in the 0, 0.5, 1.0 and 3.0 mg/kg bw
groups, respectively. Hydrocephalus was observed in 4(3) fetuses
(litters) in the 3.0 mg/kg bw groups but not in any other group
although dilated cerebral ventricles were seen in 3 fetuses from one
control litter and one fetus in the 3.0 mg/kg bw group. In
contrast, dilation of the renal pelvis was observed only in controls
in 4(3) fetuses (litters). Multiple head and limb malformations
(anonychia, aprosopia, brachydactyly) were observed in one control
fetus, retroesophageal subclavian in one 0.5 mg/kg bw fetus and one
3.0 mg/kg fetus, multiple facial anomalies (including anophthalmia)
in one fetus at 1.0 mg/kg bw, and cleft palate and persistent
truncus arteriosus with ventricular septal defect each in one fetus
at 3.0 mg/kg bw. The cleft palate occurred in a fetus with
hydrocephalus. Since hydrocephalus was also observed in fetuses of
rabbits given cyhexatin orally the incidence of this lesion is
considered to be treatment-related. The NOAEL for teratogenicity in
this study is 1.0 mg/kg bw. Dermal lesions at the dosing site were
observed at all dose levels but no systemic toxicity or
embrytoxicity was noted at any of the dose levels tested (Kirk
et al. 1987b).
In a similar study performed in a different laboratory with
cyhexatin from another source, groups of 24 mated hybrid Hy/Cr New
Zealand White female rabbits (Charles River, France) were given
daily dermal doses of 0, 0.5, 1.0 or 3.0 mg cyhexatin (96% pure)/kg
bw/day Days 6-18 of gestation (day of mating = Gestation Day 0).
The females were mated by the supplier and delivered Day 1 or 2 of
gestation. The test suspensions were prepared weekly in 0.5%
aqueous carboxymethyl cellulose at concentrations (2, 4 and 12
mg/ml) to provide the appropriate dose in a volume of 0.25 ml/kg bw.
The suspensions were applied to shaved areas on the backs of the
rabbits so that test material was not applied "more than twice on
the same site without an interval of at least seven days". The test
site was given a slight massage for about one minute after
application of the test material. It is not indicated if the site
was occluded or if any other precautions were taken to prevent oral
ingestion of the test material from the application site. The
animals were housed individually in plastic cages. Each rabbit was
given 200 g pelleted rabbit diet (UAR-112-Usine d'Alimentation
Rationnelle) each day and tap water was available ad libitum. The
animals were monitored for clinical condition, body weight and food
consumption throughout the study. Local effects were evaluated 24
hours after the last application. On Gestation Day 29 the females
were sacrificed and the ovaries and uterine contents examined. The
numbers of implantations, numbers of live and dead fetuses, fetal
weights and sex were recorded. The fetuses were examined for
external, visceral and skeletal malformations.
The test suspensions were not analysed for achieved
concentration or homogeneity. There were no deaths and no gross
signs of system toxicity in any of the groups. Erythema, atonia and
desquamation were observed at the treatment site of all three dose
levels. Cracking of the skin was observed in 6/24 rabbits at 1.0
mg/kg bw and 11/24 rabbits at 3.0 mg/kg bw. Mean scores for
cracking were 0, 0, 0.3 and 0.6 on a scale of 0-3. Edema was not
observed in any of the rabbits. There were no apparent treatment-
related effects on maternal body weights or food consumption. Two
females aborted: one at 0.5 mg/kg bw and one at 3.0 mg/kg bw. No
treatment-related effects were noted on numbers of implantations,
resorptions or live fetuses or on fetal weights or sex. The total
numbers of fetuses (litters) with malformations were 3(3), 3(3),
5(4) and 3(3) at 0, 0.5, 1.0 and 3.0 mg/kg bw, respectively.
Dilated ventricles of brain were observed in two fetuses: one each
at 0.5 and 1.0 mg/kg bw. At 1.0 mg/kg bw the fetus also had
anasarca and cleft palate. No fetus at 3.0 mg/kg bw had these
malformations. One fetus in each group including controls had
unilateral or bilateral retinal detachment. Skeletal malformation
(vertebral with or without rib malformations or pelvic girdle
malformations) were observed in 2(2), 1(1), 2(2) and 2(2) fetuses
(litters) at 0, 0.5, 2.0 and 3.0 mg/kg bw, respectively. The
remaining malformed fetus at 1.0 mg/kg bw had multiple abdominal
(visceral) and vertebral malformations. Arthrogyposis, considered
to be a minor defect, was observed in one fetus at 1.0 mg/kg bw and
2 fetuses from one litter at 3.0 mg/kg bw. Other skeletal anomalies
and variations were observed in some fetuses in all of the groups
and did not appear to be treatment-related.
The only treatment-related effect observed in this study was
local irritation at the site of application which was observed at
all three dose levels. The apparent NOAEL for teratogenicity in
this study was 3.0 mg/kg bw (Monnot, 1989c).
COMMENTS
The oral study in Sprague-Dawley rats, utilizing dose levels of
0.5-5.0 mg cyhexatin/kg bw/day (95.6% purity) administered on days
6-15 of gestation did not result in any maternal toxicity or signs
of teratogenicity. An increased incidence of vertebral variations
at 5 mg cyhexatin/kg bw/day was interpreted as evidence of
fetotoxicity.
The results of the rabbit oral teratogenicity studies available
to the Meeting were discrepant (see Table 1). In two cases, the
results were negative with respect to all parameters measured, with
NOAELs of 3 and 1 mg/kg bw/day (top doses). In a third study, the
NOAEL was 0.5 mg/kg bw/day based on increased post-implantation
losses. However, this study is of uncertain validity. The
concentrations of test material administered varied from 70-104%,
91-125%, and 55-137% of the nominal at 0.5, 1 and 3 mg/kg bw/day,
respectively, suggesting non-homogeneity in the test material.
Hydrocephalus was observed in 8 pups from 4 litters at 3 mg/kg
bw/day. The distribution of these malformations raised the
possibility that they may have been induced by infection. The
second positive study failed to show a NOAEL at the lowest dose
tested (0.75 mg/kg bw/day). This dose level caused maternal
toxicity and hydrocephalus. The abortion rate was high in both test
groups (0.75 and 3 mg/kg bw/day) as was maternal mortality in all
groups. Again the possibility of infection cannot be ruled out.
The existence of an additional rabbit teratology study using
different manufacturing batches from widely separated sources was
brought to the attention of the Meeting. This study was reported to
be negative by the testing laboratory but was not available for
review by the Meeting. In view of the conflicting results in the
positive studies, and the problems with dose levels in one of the
studies, together with the knowledge of 3 negative studies as well
as a completed but unreported study, the Meeting determined that
cyhexatin should be reviewed again in 1991 when data, known to be in
process of development, should be available. Meanwhile, the present
ADI remains unchanged.
TOXICOLOGICAL EVALUATION
Level causing no toxicological effect
Mouse: 3 mg/kg bw/day
Rat: l mg/kg bw/day
Dog: 0.75 mg/kg bw/day.
Estimate of acceptable daily intake
0-0.008 mg/kg bw.
Studies which will provide information valuable in the continued
evaluation of the compound
1. Complete specifications (including nature and content of
impurities) and physical properties of test materials used in
all studies.
2. Data on absorption and tissue distribution in the pregnant
female rabbit.
3. Submission of a rabbit teratology study known to exist.
4. Submission of a two-generation reproduction study in rats known
to be in progress.
TABLE 1. TOXICOLOGICAL EVALUATION RESULTS OF RABBIT ORAL TERATOGENICITY STUDIES
EMBRYO/FETAL TOXICITY MATERNAL TOXICITY TERATOGENICITY
NOAEL LOAEL EFFECT NOAEL LOAEL EFFECT NOAEL LOAEL EFFECT
STUDY I1 0.5 1.0 Increased post 3.0 - No effects 1.0 3.0 Hydrocephalus
implantation
loss
STUDY II2 - 0.75 Abortion 0.75 3.0 Reduced - 0.75 Hydrocephalus
maternal
body weight
STUDY III3 3 - No effect 3 - No effect 3 - No effect
STUDY IV3 1 - No effect 1 - No effect 1 - No effect
1 IRDC Laboratories
2 Dow, USA Laboratories
3 Hazleton, France
TABLE 2. ORAL (GAVAGE) TERATOLOGY STUDIES WITH CYHEXATIN
Laboratory IDRC9 Dow USA Hazleton, France Hazleton, France
Test material
purity 95.6% 94.8-95.5% 96% 96%
source Dow Dow Oxon Oxon
form powder powder powder powder
Dose suspension
vehicle 0.5% Methocel 0.5% Methocel 0.5% CMC 0.5% CMC
volume 1 ml/kg bw 1 ml/kg bw 5 ml/kg bw 5 ml/kg bw
preparation once once weekly weekly
agitation during dosing continuous during dosing during dosing
doses (mg/kg bw) 0, 0-5, 1.0, 3.0 0, 0.75, 3.0 0, 3.0, 3.0 (pure) 0. 0.5, 0.75, 1.0
Experimental animals
strain NZWb, SPFc NZW Hy/Cr NZ Hy/Cr NZ
source Hazleton, Pa, USA Hazleton, Pa, USA Charles River, France Charles River, France
age about 5 months not given 16-18 weeks 16-18 weeks
weight (kg) 3.62 - 4.32 3.5 - 4.5 3.10 - 4.35 3.05 - 4.25
insemination artificial artificial matedd matedd
acclimation 14 days 3 weeks none none
diet Purina #5322 Purina #5322 UARe UAR-112
feeding ad libitum 8 oz/day 200 g/day 200 g/day
cage wire not given plastic plastic
housing individual individual individual
a CMC = carboxymethyl cellulose
b NZW = New Zealand white
c SPF = Specific pathogen free
d mated by supplier, delivered Day 1-2 of gestation
e UAR = Usine d'Alimentation Rationnelle
f rabbits received 4oz food/day prior to start of study (insemination)
g IRDC = International Research and Development Corporation, Michigan, USA.
REFERENCES
Aldridge, D., Schwartz C.A., Keller, K.A. & Schardein, J.L. (1986)
Cyhexatin -Oral teratology study in Sprague-Dawley rats (with
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Research and Development Corporation (IDRC). Submitted to WHO by
Dow Chemical Co., Midland, Michigan, USA.
Berdasco, N.M., Johnson, K.A., Wolfe, E.L. & Hanley, T.R. Jr (1986)
Cyhexatin: oral teratology probe study in New Zealand White rabbits
Unpublished Report from Mammalian and Environmental Toxicology
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Co., Midland, Michigan, USA.
Kirk, H.D., Johnson, K.A. & Hanley, T.R., Jr (1987b) Dermal
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Monnot, G. (1989a) Cyhexatin - oral (gavage) teratology study in
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