NOTES
AND
tube prevents particles from entering the
tube before sampling.
The collecting bottle is a 300-ml polythene bottle with two tubes fitted into the
screw cap. The wider tube is connected by
rubber tubing to one arm of the U-shaped
sampling tube. The other tube liberates
air slowly during sampling. The latter tube
is of flexible polyvinyl chloride ( PVC) ; in
use it is sealed by being folded into a small
clip made from tubing. Until this seal is
broken, no water can enter the bottle. The
seal is broken, as the tide rises, by means of
a float attached to the tube. The rate at
which water enters the bottle is controlled
by the bore of a capillary tube contained
within the PVC tubing. For the present
sampler, this was chosen to give a rate of
flow approximately equal to that produced
by a filtering animal in the same position
( approximately 2-3 liters/hr ) . When used
in less than 1 m, the capillary is omitted,
since otherwise the sampler may not
operate.
FLOTATION
COMMENT
601
In use, the bottle is attached to the stake.
A rod pushed into the substratum supports
the float. The sealed fine tube is fixed by
a length of thread to the float, the seal
being broken when the thread is pulled
taut by the float. The stage of the tide at
which the sampler will operate is controlled
by adjusting the length of the thread.
If desired, a little formalin
may be
placed in the sample bottle so that the
sample is preserved as it is collected. Small
filters of plankton netting can be incorporated in the wide tube through which the
sample is taken to separate the larger
planktonic organisms at sampling.
We are indebted to the Central Electricity Generating Board for a grant and
facilities.
J. C. BAYES
A. D. ANSELL
Department of Zoology and
Planktoti Laboratory,
University of Southampton,
England.
TECHNIQUE FOR SEPARATING FECAL PELLETS AND SMALL
MARINE ORGANISMS FROM SANDY
A technique has been developed for
flotation separation of low-density particles,
such as fecal pellets or organisms, from
water-wet sediments. The method employs
a flotation
medium that is not watermiscible (carbon tetrachloride)
after treating the wet sediment with detergent. It
is frequently necessary to keep sediments
wet in order to prevent aggregation. However, inexpensive, water-miscible
flotation
media of the proper density ordinarily have
viscosities too high for other than centrifugation methods. Usable solutions are
generally expensive, dangerous, or both.
Carbon tetrachloride has the high density
(1.58) and low viscosity (0.97 centipoise at
2OC) of an ideal flotation medium. Unfortunately, water is not miscible with CC14,
1 Contribution
No. 92 from the Narragansett
Alarinc Laboratory.
This work is supported
by
U.S. Public Health Service Grant WP 00023.
and water wets glass preferentially.
Adding
CC14 to water-wet particles results in the
formation of water globules that enclose
groups of particles that stick to the walls
of a glass vessel. A detergent, being a mixture of wetting and emulsifying
agents,
allows the water to break up into tiny
droplets, freeing the grains for flotation
separation. The present method was developed to separate fecal pellets from sand.
Since pellets are denser (density about
1.19 5 0.03) than organisms (the density
of most invertebrates
is less than 1.12,
Anderson 1959), the technique could also
be used to separate small animals from
sand.
Methods of flotation separation of organisms from marine sediments have been
suggested by Teal (1960), Anderson (1959),
and Birkett ( 1957). Birkett used carbon
tetrachloride for flotation of larger forms,
including mollusks, from wet sediment and
602
NOTES
AND
stated that it produced neither osmotic
effects on the flesh nor chemical corrosion
of the shell. However, since Birkett did not
use detergent, it is evident that his method
could not be used with small, water-wet
organisms. If a separation of very small
particles (0.5-200 p) is required, the centrifugation
methods of Lammers (1962,
1963) are applicable.
METHOD
A small sample of fecal pellets and sand
that has never been dried is placed in a polyethylene or polypropylene beaker and most
of the water decanted. Concentrated detergent solution is added (about 25 g of
Alconox per liter of water) and the mixture
agitated. As much of the liquid as possible
is carefully poured off. Carbon tetrachloride, several times the volume of the
particles, is added, and the mixture is
stirred thoroughly.
The pellets rise to the
surface while the sand remains on the
bottom. The water that remains with the
grains forms tiny droplets. The liquid and
particles are transferred to a polyethylene
funnel that has a piece of rubber tubing
slipped over the stem with a spring clamp
closing the tubing. The fractions then may
be isolated as in a separatory funnel.
This arrangement obviates the separatory
funnel’s stopcock, which might become
clogged. Plastic vessels are used instead
of glass since they are not wet by water
and therefore the water-wet grains do not
adhere to their walls. A wash bottle containing CC& is useful when transferring
particles from beaker to funnel.
The Ccl, is removed from the particles
by filtration, using a high-speed filter paper
( Reeve Angel No. 711). The particles are
cleaned of Ccl, by pouring warm dilute
detergent solution (30-40C)
over them,
then rinsing with warm water while they
remain on the filter paper. A stream of
COMMENT
water from a wash bottle easily washes
pellets or sand off the paper.
Sand-pellet fractions were produced by
water elutriation.
In this manner, pellets
could be removed from sediments that were
primarily silt and clay by first removing the
fine material at low flow rates, then the
sand-pellet mixtures at higher flow rates.
Pellets dealt with vary from about 0.06 to
about 0.8 mm in length and are associated
with hydraulically
equivalent sand and silt
of about 0.04 to 0.35 mm median diameter.
Care should be taken not to ingest or
breathe the vapor of carbon tetrachloride,
and skin contact should be avoided. Ingestion of 5 cc is considered lethal, and the
maximum allowable concentration
in air
is 25 ppm (Merck and Co. 1960). Exposure
to carbon tetrachloride is especially dangerous after consumption of alcohol.
This technique makes quantitative
estimates of total amount of pellets in a sediment possible and can help to avoid much
tedious work in separating small organisms
from sediment.
WILLIAM
P. DILLON
Graduate School of Oceanography,
Unitiersity of Rhode Island,
Kingston.
REFERENCES
ANDERSON, R. 0.
1959. A modified
flotation
technique for sorting bottom fauna samples.
Limnol. Oceanog., 4: 223-225.
BIRKETT, L. 1957. Flotation techniques for sorting grab samples.
J. Conseil, Conseil Perm.
Intern. Exploration
Mer, 22: 289-292.
LAMMERS, W. T. 1962. Density gradient separation of plankton and clay from river water.
Limnol. Oceanog., 7 : 224-229.
-.
1963.
Density
gradient
separation
of
organic and inorganic particles by centrifugation.
Science, 139: 1298-1299.
MERCK AND Co., INC. 1960. The Merck index.
7th ed. Rahway, N.J. 1642 p.
TEAL, J. M.
1960. A technique for separating
nematodes and small arthropods from marine
muds.
Limnol. Oceanog., 5: 341-343.