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EC number: 200-073-0
CAS number: 50-97-5
Table 2. Calculation of Stimulation Indices per Dose Group
Test item concentration
Mean DPM peranimal (2 lymph nodes)a)
Vehicle Control Group (DMF)
Vehicle Control Group for the Positive Control Item (acetone/live oil (4+1, v/v))
Positive Control Group(25% α -HCA)
a) = Mean DPM/animal was determined by dividing the sum of the measured
values from lymph nodes of all animals within a group by the number of
animals in that group (5 animals)
* = Statistically significant (p<0.05) if compared to concurrent
Table 3. Calculation of the EC3 value
Test item concentration %
EC3 = (a-c) [(3-d)/(b-d)] + c = 18.2% (w/w)
a,b,c,d = Co-ordinates of the two pairs of data lying immediately above
and below the S.I. value of 3 on the LLNA dose response plot.
Table 4. Ear Thickness in the Main Experiment
Ear thickness gain
Difference day 1 to 3
Difference day 1 to 6
Mean per dose group
Mean per dose group (%)
OECD 429 (2018): Three groups each of five female mice were treated once
daily with the test item at concentrations of 10, 25, and 50% in DMF by
topical application to the dorsum of each ear for three consecutive
days. Three further groups each of five mice were treated with the
vehicle for the test item (DMF) or the vehicle for the positive control
item (acetone:olive oil (4+1 v/v)) only or with the positive control at
Five days after the first topical application the mice were injected
intravenously into a tail vein with radio-labelled thymidine (3H-methyl
thymidine). Approximately five hours after intravenous injection, the
mice were sacrificed, the draining auricular lymph nodes excised and
pooled per animal. Single cell suspensions of lymph node cells were
prepared from pooled lymph nodes, which were subsequently washed and
incubated with trichloroacetic acid overnight. The proliferative
capacity of pooled lymph node cells was determined by the incorporation
of 3H-methyl thymidine measured in a scintillation counter.
No mortality was reported and no signs of systemic toxicity were
observed. Some animal some animals showed an erythema of the ear skin
with statistically significant and biologically relevant increase in ear
Stimulation Indices of 1.8, 4.0, and 4.5 were determined with the test
item at concentrations of 10, 25, and 50% (w/w) in DMF. A dose response
was observed. The EC3 value calculated was 18.2 % (w/w).
The animals treated with the positive control item
alpha-Hexylcinnamaldehyde (25% (w/v)) showed a distinctly positive
response (mean S.I. of 8.3), corroborating the validity of the assay.
Based on the condition of the study, the test
item,α-phenyl-1H-benzimidazole-2-methanol was found to be a skin
sensitiser cat. 1B in accordance with the Globally Harmonized
Classification System and to the Regulation (EC) No. 1272/2008.
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