AMMONIUM SULFATE CAS N°: 7783-20-2

OECD SIDS
AMMONIUM SULFATE
5. TOXICITY ID: 7783-20-2
DATE: 18.04.2006
dihydrostreptomycin sulfate in 10-cm petri dishes; 48 hr
before the treatment 2x10e5 cells from the stock culture
were seeded in 10-cm petri dishes with 10 mL of complete
medium.
MUTAGEN TREATMENT: Before adding the mutagen the culture
medium was discarded and replaced by 10 mL of prewarmed MEM.
2 mL of ethyl methanesulfonate (EMS), dissolved in MEM were
added to each dish, to give a final concentration of 20 mM.
After 1 h of treatment at 37 deg C, the medium was removed
and the dishes were washed twice with 10 mL of prewarmed
phosphate-buffered saline (PBS).
POST-TREATMENT WITH AMMONIUM SULFATE or SODIUM CHLORIDE:
The hypertonic treatment was performed immediately after the
mutagen treatment. The washing solution was carefully
removed, then the cells were covered with 300 uL of PBS to
avoid drying. Then 300 uL of ammonium sulfate or sodium
chloride solution were added to give a final osmolality of
500, 750, 1000 or 1500 mOsm/kg H2O. Controls and EMS-treated
cultures received 600 uL of PBS (300 mOsm/kg H2O). The
osmolality of all solutions was measured with a
microosmometer. After 30 min at 37 deg C the treatment was
stopped by adding 5 mL of PBS. The supernatant of all dishes
was collected in centrifuge tubes so that no cells were
lost. Then the cells were trypsinized by adding 2 mL of
trypsin/EDTA. After 3 min the cells were collected with 2 mL
of medium and added to the supernatant. After centrifugation
the cells were counted and plated for the chromosomal
aberration and TGr mutation assay.
CHROMOSOMAL ABERRATIONS:
Immediately after the posttreatment with hypertonic salt
solutions 10e6 cells were transferred to 6-cm dishes and
cultivated for 13 or 16 hours, respectively, in complete
medium at 37 deg C, including a 2.5 h colcemid treatment.
The cells were fixed in methanol-acetic acid and stained
with 5% Giemsa. In some experiments cultures containing
BrdUrd were set up in parallel to ensure that only first
post-treatment metaphases were scored for chromosomal
aberrations.
TGr MUTATIONS:
5x10e5 cells were seeded in 10-cm dishes and cultivated for
7 days after the treatment. Then the medium was removed, the
cells were washed once with PBS and trypsinized. Then 10e5
cells were seeded in 6-cm dishes (10 per concentration) and
grown in 5 mL complete medium including 10.9 ug/mL of
6-thioguanine (TG). Cell survival tests were carried out in
parallel to the selection procedure. Colonies were fixed and
stained after 8 days of incubation.
Reliability: (3) invalid
Unsuitable test system (unphysiological, hypertonic test
conditions).
12-APR-2006 (145)
5.6 Genetic Toxicity 'in Vivo'
Type:
Micronucleus assay
Species: mouse Sex: male
Strain:
other: ddY
Route of admin.: i.p.
UNEP PUBLICATIONS 193