Day 15 - May 26
Weather Report
Time: 12: 35 PM Pacific Daylight Time
Air Temperature: 6.6 Degrees Celsius
Sea Temperature: 6.7 Degrees Celsius
Wind Speed: 22 Knots
Ship's speed: 1.6 Knots
Latitude: N 50.01
Longitude: W 145.02
Filtration of the sea water has
been mentioned as a daily process that occurs after samples are
collected by the CTD. Below is an explanation of the chlorophyll
filtration protocol, written by Natalie, a graduate student from the
University of Western Ontario.
Chlorophyll filtrations begin after
samples of sea water at differing depths have been collected from the
CTD. Usually samples are collected in dark bottles from depths of
100 meters, 80 m, 60 m, 40 m, 25 m, 15 m, 10 m, 5 m, and 0 m below the
surface of the water. Samples of water are also collected at the
maximum chlorophyll depth and the depth at which light intensity is 1%
of the intensity at the surface of the water, both of which vary from
sampling station to sampling station.
Once the samples are collected, they
are run through the filtration rig. The rig is hooked up to a
vacuum pump such that when water is poured on top of the filter, the
vacuum sucks the water through the filter, leaving behind the cells
that will be broken apart for quantification of chlorophyll content.
Prior to running each sample through
the rig, the bottle containing the sample is gently inverted four to six times
to ensure that the contents within the sample are well-mixed, after
which a portion of the sample is measured using a graduated cylinder
pre-washed in distilled water; aliquots of 100mL are measured for
depths below 40 m while aliquots of 50mL are measured for depths at 40m
and above. Each sample is run through the rig twice to ensure
confidence in the chlorophyll measurements attained.
After all of the water from a given
aliquot has been sucked through by the vacuum, the filter is collected
off of the rig and placed in a small culture tube. Once all of
the samples have been run and the filters collected, 8 mL of acetone
are added to each tube. The acetone extracts the chlorophyll from
the filter and places it in suspension. Each tube is then capped,
wrapped in tin foil, and placed in a freezer for 24 hours to allow
sufficient chlorophyll extraction to occur.
After the 24 hours have passed, the
samples are removed from the freezer and placed in a dark room that we
call the "chlorophyll bubble" for one hour to enable the samples to
warm to room temperature. The room is kept dark to ensure that
the chlorophyll content within each tube does not degrade from light
exposure. In this room is a machine called a fluorometer; the
fluorometer gives a measurement of chlorophyll content within each
tube. Each tube is gently inverted three times to mix the acetone
and the extracted chlorophyll contents, wiped clean with a Kimwipe, and
then placed in the machine's cuvette holder. The machine then
produces a value of chlorophyll content within each tube on the digital
monitor.
However, we are only looking for
chlorophyll a, whereas each tube contains a mixture of chlorophyll a,
chlorophyll b, and other non-target components. To figure out
chlorophyll content, three drops of a 10% solution of hydrochloric
acid (HCl) are added to each tube after the first reading. The
HCl will deactivate chlorophyll a contents in each sample by breaking
apart its porphyrin ring, and as a result the fluorometer no longer
includes chlorophyll a in its readings. After the acid addition,
the tube is gently inverted three times, wiped clean with a Kimwipe,
and placed in the cuvette holder. A second reading for the sample
is recorded, and the difference between the reading before the acid was
added and after acid addition gives a measure of chlorophyll a content
in each sample.
This procedure repeats each time a CTD sampling occurs.
Chlorophyll Filtration Protocol in the Classroom
Photosynthesis WebQuest
Another Photosynthesis WebQuest