]RC/CAU/ 1st March 1968
CCt44DMRY REZATING TO FE RIVORT NO. .37
TnLE: - THE CILLNWATIC EFFECT OFSKCKE ON TETRAHWWA
Lrf.VMjUUVr.LON
The reason-given for this work was that the coefficient of variation
(c.v.) for results obtained with Paramecium was too high. Although they
were able to reduce the c.v. from aa, 20% to 12-15% with Paramecium by
changing techniques, they reckoned to get down to a c.v. of 5% using
Tetrahymena. 7he reason is partly due to the fact that Tetrahymena can
be grown in sterile culture.
7he Initials CAMT are used throughout for the standard test. 7he
results are given either as seconds to kill or as see-Ix 1CP, deflned
as ciliastatic effectiveness W.
Pp 9-16 are devoted to a description of the test apparatus etc. A 50 ml
puff of whole smoke or vapour phase - more usually the latter - is taken
by syringe and Is passed over a 5 gl%drop of culture. '1he time to kill
Is recorded. Descriptions of culture solutions for T-vorax and T-pyriformiE
are given - the culture solution for the latter is particularly complex
having 42 constituents.
Pp 16-17 Cover the details of carrying out the test. .7he pH of the
culture solution used (5-5 - 5.8) is low (relative to both the human system
and to the R. & D.E. system where it in close to 7).
Pp 17-19 Cover the d6taile of reproducibility (for vapour phase) which is
clearly very good (Table 1 and 2, pp 18 and 19).
Pp 20 Day to day variation La shown to be sllght-(Table 3, p.20).
Pp 2D-38 Cover Results
(1) The effect of drop size is shown (Table 4).
(2) The effect of temperature is shown to be slight over the rantre
5-300C (Table 5). r110
BATCo document for Province of BritiSh Columbia 19 April 1999

-2-
(3) Photosensitivity was not observed.
(4) Increasing culture age 1.9 connected with a lower pH value and
to high values of c.
(5) The effect of increasing pH Is to reduce c (Table 6). 7his
result Is in accord with R. & D.E. reported results for
Paramecium and whole smoke.
(6) Rather different results were obtained with T.vorax and T. pyriform I
(Table 7). This may be due to differences in the culture solution.
(7) '1he Influence of tobacco moisture an (vapour phase) toxicity la
given In Table 9. This results again is In accord with R. & D.E.
reported results for Paramecium.
(8) Cigarette pressure drop variation appears to have no effect
(Table 10). -
(9) Paper porosity appears to have some effect on the toxicity of
vapour phase, Interacting with puff number (Table 11).
(10) Acetate and paper filters have a slight effect, as they frequently
do In R. & D.E. Paramecium tests (Table 12).
(11) Perforated paper was tested against unperforated paper. Perforations
reduced the toxicity (Table 13).
(12) Tbbacao filters had a slight effect (Tables 14 and 15); Gauloise
and Player's tobaccos wore compared as filters.
(13) The Influence of puff duration (at constant volume) was tested,
and was found to be very slight. Mere ls-an optimum duration
(Table 16).
(14) The Influence of various puff volumes (all of 12 seconds duration)
was tested. A slight but definite effect was found (Table 17).
(15) Me Influence of puff volume at constant speed was tested; there
was no difference between puff volumes of 20-50 ml (Table IS).
(16) There was only a very slight effect of varying the puff-flow speed r%j
In the smoke cell (Table 19).
BATCo document for Province of British Columbia 19 April 1999

-3-
(17) The effect of varying the concentration of gas phase on the
c value is-given in Table 2o and Figure 11. (There is scope
for some interesting arithmetic to explain the relationship
between puffs and time to kill in cillastasis work.)
The results suggests two other points.
(1) Since the relationship between concentration and time
to kill Is exponential, it becomes inconvenient to measure the time
to kill at low levels of toxicity - below any 20% of the normal
concentration of the vapour phase for example. Measurement of
puffs to kill Is more convenient in this respect.
(II) It would be more meaningful if Hamburg, using their
dose-response relationship, were to give their results on the basis
of % change In delivery of toxic material. For example, they could
calculate the filtration efficiencies of the filters tested.
(18) There was a -slight increase (Table 21) in toxicity towards the
butt end of the Player's cigarette, and a marked increase with
the Gauloise. This result fits with results reported for some
air-cured tobaccos In R. & D.E.
(19) Table 22 gives the results obtained with several different
cigarettes.
The most important points in the Discussion (PP 39-40) are:
(1) They recognize that you cannot directly test the effect of
smoke an cilia in Protozoa; the culture solution always
intervenes. One should therefore try to Minimize the effect
of the culture solution.
(2) They point out the difficulties of working with whole smoke
you cannot see the organisms through it. They say one 2 second
puff is not enough for cillastasis.
BATCo document for Province of British Columbia 19 April 1999

-4-
(3) They do not like the Australian and R. & D.E. system of using
intermediate fresh air puffs - because of selectivity of some
smoke constituents. This argument Is scarcely valid since the
smoker breathes fresh air between puffs of smoke.
(4) They do not like the IDLOO methods (like B. & D.E. solution
method) because the smoke is not fresh. This is true, but
unstable compounds such as acrolein do Influence the results.
so presumably only less stable compounds than acrolein are
lost. They also criticise the method because, they say, there is
selective solution. This Is in fact probably less true of the
solution method than of drop metho& such as theirs and ours.
They say the simplest system Is to use the vapour phase alone.
This has never been acceptable In the work in Australia,
Battelle (Frankfurt) or B. & D.E., for the obvious reason
that the smoker uses whole smoke.
(5) They say, that the examination of single puffs does not permit
conclusions about whole cigarettes. This is accepted for some
-cured tobacco cigarettes, where they and we have reported
air
a large Increase In activity towards the butt end. It Is
less important for flue-cured tobacco cigarettes. There Is a
difference in the Hamburg and R. & D.E. findings in that they
find an increase In toxicity per puff (of vapour phase) towards
the butt and, but we h&ve never found differences In toxicity
of lat and 2nd halves of flue-cured tobaoco cigarettes (for
whole smoke). This does not seem to be due to the lower
sensitivity of our system.
They say that by comparing smoke analysis figures with CART
CM1
values no substance can be Indicated.
'They say that G. L. C. examination of the drop after ciliantanis 0%
BATCo document for Province of British Columbia 19 April 1999

-5-
points to HCX as being active. 'This argument seems q4te
KLCL."061J,,
unsound since compounds such as acrolein will not.remain
unaltered after having exerted their toxic effect.
V@
B. Comber.
CZ1
C=O
BATCo, document for Province of BritiSh Columbia 19 April 1999