Primary Productivity of Phytoplankton
213
pH measurements. A void direct sunlight and store in an ice chest. Measurements
of alkalinity and pH should be performed as soon as possible (see Exercise 8).
Particularly in nutrient-poor, soft waters, direct analysis of DIC will be necessary
(see pp. 111 and 117).
4. After samples have been collected from all depths, open each sample bottle in the
darkened box. Inoculate each bottle with 1.0 ml of NaH 14 C0 3 [2 to 5 microcuries
(IlCi)/ml] with a spring-operated syringe by the following procedure. Score the
neck of the 14C-ampoule (or use prescored ampoules) and snap-break the ampoule
open. Fill the syringe to remove all air bubbles. The syringe should be equipped
with a 10- to 15-cm hypodermic needle or cannula. Transfer 1.0ml of the tracer
solution slowly into the bottom of the sample bottles. Immediately stopper the
bottles as soon as the tracer has been injected to avoid double inoculation or
omission. Time zero of the incubation is taken to the nearest minute at midpoint
of inoculation of the series of bottles.
Caution: Although the amount of radioactivity employed is very small, the half-life
of 14C is very long (5760 yr), and the isotope must not be introduced into the
body. All handling of the isotope must be done with extreme care. Disposable
gloves are recommended, and all excess solutions and the glassware must be
disposed of properly and cleaned per the directions of the instructor.
The amount of 14C required varies with growth rates, population density, light,
temperature, length of incubation, amount of sample filtered, and other factors.
Under many conditions of moderate productivity, 1 to 31lCi per 130-ml sample,
a 4-h incubation, and filtration of a 50-ml subs ample yields sufficient radioactivity
in the algae for good statistical radio assay without excessive layering of the
phytoplankton on the filter.
5. Make certain that each bottle is sealed tightly. Mix each gently but thoroughly
by repeated inversion and cap the dark bottles with aluminum foil to assure that
no light leaks through the stopper joint. In a shaded area, rapidly attach the bottles
to bottle spreaders with the dark bottle first in the center and then light bottles
on the ends. Lower the bottles to the depth from which they were collected.
Incubate for 3 to 4 h (not exceeding 6 h) between 0900 and 1500 h of the day.
Make certain that the bouy does not shade the upper bottles.
6. Perform analyses of pH, alkalinity, and/or DIC as soon as possible. Water
temperatures should be taken at each sampling depth.
7. At the end of the incubation, retrieve the bottles and place them into a light-proof
box as rapidly as possible. The time of the end of the incubation is the midpoint
of the retrieval operation.
8. Aliquots from each bottle, starting with the surface samples (usually most active),
are transferred by pi petting (no mouth pipetting) as rapidly as possible to the
filtration apparatus. Use membrane filters with a pore size of0.45llm (e.g., Millipore
HA or equivalent) and vacuum not exceeding 0.5 atm (380 mm Hg) to reduce the
possibility of rupturing more fragile cells as they aggregate on the filter. Filtration
should be performed in a semi darkened area to minimize further photosynthesis.
The vacuum should be released immediately after the water has passed the filter
to avoid rapid air desiccation.
To estimate extracellular release of dissolved organic carbon by the phytoplankton, obtain samples ofthe filtrate at this point (see discussion below, p. 219).
The amount of water to be filtered is governed by the observed density of the
phytoplankton. In oligotrophic waters, 100-ml subsamples should be filtered; in
mesotrophic waters filtration of 50-ml usually is adequate. Because the f1-radiation
213
pH measurements. A void direct sunlight and store in an ice chest. Measurements
of alkalinity and pH should be performed as soon as possible (see Exercise 8).
Particularly in nutrient-poor, soft waters, direct analysis of DIC will be necessary
(see pp. 111 and 117).
4. After samples have been collected from all depths, open each sample bottle in the
darkened box. Inoculate each bottle with 1.0 ml of NaH 14 C0 3 [2 to 5 microcuries
(IlCi)/ml] with a spring-operated syringe by the following procedure. Score the
neck of the 14C-ampoule (or use prescored ampoules) and snap-break the ampoule
open. Fill the syringe to remove all air bubbles. The syringe should be equipped
with a 10- to 15-cm hypodermic needle or cannula. Transfer 1.0ml of the tracer
solution slowly into the bottom of the sample bottles. Immediately stopper the
bottles as soon as the tracer has been injected to avoid double inoculation or
omission. Time zero of the incubation is taken to the nearest minute at midpoint
of inoculation of the series of bottles.
Caution: Although the amount of radioactivity employed is very small, the half-life
of 14C is very long (5760 yr), and the isotope must not be introduced into the
body. All handling of the isotope must be done with extreme care. Disposable
gloves are recommended, and all excess solutions and the glassware must be
disposed of properly and cleaned per the directions of the instructor.
The amount of 14C required varies with growth rates, population density, light,
temperature, length of incubation, amount of sample filtered, and other factors.
Under many conditions of moderate productivity, 1 to 31lCi per 130-ml sample,
a 4-h incubation, and filtration of a 50-ml subs ample yields sufficient radioactivity
in the algae for good statistical radio assay without excessive layering of the
phytoplankton on the filter.
5. Make certain that each bottle is sealed tightly. Mix each gently but thoroughly
by repeated inversion and cap the dark bottles with aluminum foil to assure that
no light leaks through the stopper joint. In a shaded area, rapidly attach the bottles
to bottle spreaders with the dark bottle first in the center and then light bottles
on the ends. Lower the bottles to the depth from which they were collected.
Incubate for 3 to 4 h (not exceeding 6 h) between 0900 and 1500 h of the day.
Make certain that the bouy does not shade the upper bottles.
6. Perform analyses of pH, alkalinity, and/or DIC as soon as possible. Water
temperatures should be taken at each sampling depth.
7. At the end of the incubation, retrieve the bottles and place them into a light-proof
box as rapidly as possible. The time of the end of the incubation is the midpoint
of the retrieval operation.
8. Aliquots from each bottle, starting with the surface samples (usually most active),
are transferred by pi petting (no mouth pipetting) as rapidly as possible to the
filtration apparatus. Use membrane filters with a pore size of0.45llm (e.g., Millipore
HA or equivalent) and vacuum not exceeding 0.5 atm (380 mm Hg) to reduce the
possibility of rupturing more fragile cells as they aggregate on the filter. Filtration
should be performed in a semi darkened area to minimize further photosynthesis.
The vacuum should be released immediately after the water has passed the filter
to avoid rapid air desiccation.
To estimate extracellular release of dissolved organic carbon by the phytoplankton, obtain samples ofthe filtrate at this point (see discussion below, p. 219).
The amount of water to be filtered is governed by the observed density of the
phytoplankton. In oligotrophic waters, 100-ml subsamples should be filtered; in
mesotrophic waters filtration of 50-ml usually is adequate. Because the f1-radiation