Toxicity of Anemone Extracts — Martin 
303 
two 1-min periods. After being centrifuged for 
30 min at 1000 g the supernatant was decanted 
and centrifuged again. The resulting superna- 
tant extract was turbid but did not contain cells 
or debris. It was either decanted or, when it 
was found covered with a lipid layer, removed 
with syringe and needle; then its pH was deter- 
mined. The extract was dialyzed for 4 hr through 
commercial cellophane tubing against ten times 
its volume of 0.001 M sodium phosphate (pH 
6.2) in 0.8% sodium chloride with 1% acti- 
vated charcoal (Weinberger, 1936). The dial- 
ysis was repeated once with fresh solution. All 
operations were performed at room temperature. 
Extracts of the other lower metazoa were pre- 
pared in the same manner. All extracts were as- 
sayed immediately after their preparation. The 
potency of extracts of each species was com- 
pared with that of an extract of A. elegantissima 
prepared on the same day. 
ASSAY: The extract was given intraperi- 
toneally to DAL-Swiss albino mice weighing 
17 to 23 g. The mice were placed in cages in 
lots of four to eight (Russell et al., I960), and 
observed for 24 hr; their survival time was 
recorded. The abdominal cavities of the mice 
were inspected post mortem and mice with 
intraperitoneal hemorrhage were discarded. This 
condition was found in animals that showed 
severe pain reaction immediately after the in- 
jection and died within 4 min. The discarded 
animals were replaced by supplementary ones 
to complete the series. Each extract was as- 
sayed on a series of at least four mice. 
EXPERIMENTS AND RESULTS 
Preliminary experiments explored the effect 
of varying doses of extracts of A. elegantissima 
on the survival time of the injected mice. Only 
the area of LD<)<* was considered. A dose effect 
curve showed that with decreasing doses the 
survival time increased. At the dose of 20 cc 
of extract per kilo of mouse, both the range 
and the mean of the survival time were within 
practical limits; therefore, this dose was se- 
lected as a convenient reference. 
The pH of all the extracts varied from 6.0 
to 6.6. 
At a dose of 20 cc of extract of A . elegantis- 
sima per kilo of mouse, the mean survival time 
of the injected mice varied from 8 to 36 min 
among the ten series tested. Between some of 
these series the difference in survival time was 
significant at the 2% level by the Chi-square 
test. The cause of these differences is unknown. 
The differences showed no correlation with vari- 
ations in the size, sites of origin, or dates of 
collection of the specimens, or with the pH 
of the extracts. They were not relevant for the 
present study. 
Table 1 shows that the potency of extracts 
may vary from one species of anemone to 
another. The highest potency, as estimated by 
the survival time of mice after intraperitoneal 
injection, was found for the extracts of A. ele- 
gantissima and A. xanthogrammica. Extracts of 
other anemones were far less potent even when 
their dose was tripled. With extracts of T. lofo- 
TABLE 1 
Comparison of the Survival Time of Mice after Intraperitoneal Injection 
of Extracts of Anemones 
SPECIES 
NO. OF 
BATCHES 
NO. OF 
MICE 
DOSES CC OF 
extract/kg 
OF MOUSE 
SURVIVj 
M 
Mean 
\L TIME IN 
INUTES 
Range 
PROPORTION 
OF MICE 
SURVIVING 
24 HR 
Anthopleura elegantissima 
10 
48 
20 
15.7 
5-47 
1/48 
A. xanthogrammica 
2 
10 
20 
9.2 
6-15 
0/10 
Metridium senile 
1 
6 
20 
174.1 
80-240 
0/6 
Corynactis calif or nica 
1 
5 
60 
138.5 
118-180 
0/5 
Tealia crassicornis 
1 
6 
60 
342.8 
252-480 
0/6 
T. lofotensis 
1 
4 
20 
336 
3/4 
T. lofotensis 
1 
4 
60 
460-540 
2/4 
T. coriacea 
1 
4 
20 
264-640 
2/4 
T. coriacea 
1 
4 
60 
480-900 
2/4 
