The contents of
one flask were analyzed for each incubation period.
The flasks were swirled gently to effect extraction of
the toxin. The contents of each flask were transferred
to a 250-ml separatory funnel, rinsing the flask with
chloroform from a wash bottle. The separatory funnel
was inverted gently six to eight times, and the phases
were allowed to separate. Vigorous shaking was
avoided because it results in the formation of an
emulsion. The lower chloroform layer was drawn off
through ca. 40 g of anhydrous sodium sulfate in a
Butt extractor tube. The chloroform extract was
collected in a 300-ml round-bottom flask. The extraction
was repeated twice more with 50-ml portions
of chloroform. After the last extraction the sodium
sulfate was rinsed with approximately 10 ml of
chloroform. The combined chloroform extracts were
evaporated almost to dryness with a rotary evaporator,
and then were quantitatively transferred to a 25- or