Technique of Measuring Phytoplankton Primary Production
47
be used in the absorber. In this case, it is discharged directly into the scintillation vial and radioassayed after addition of the scintillation cocktail (Sorokin
1987).
The alkaline - acid persulfate combustion described above can be
replaced by an alternative UV-combustion method (Armstrong et a1. 1966;
'lbomas 1971; Williams and Yentsch 1976; Williams et a1. 1976). The combustion of labeled organic matter to 14C02 in this case proceeds under the prolonged action of strong ultraviolet radiation in the presence of oxidizers such
as hydrogen peroxide or persulfate. For UV-combustion, the subsamples of
filtrate 50 or 100mi after extraction of labeled CO2 from them are neutralized
to pH 9.5 and placed in test tubes of pure fused quartz. Then two to three
drops of 30% H 20 2 or 0.5 g dry Na-persulfate are added to them. They are
closed with glass stoppers or glass balls and illuminated under a 1200-W round
mercury lamp (Hanovia 189, with fan) for 3-4h. After cooling, the 14C02 thus
formed is extracted from the samples as described above in the apparatus, or
using a simple device described by Thomas (1971), which resembles that used
for cleaning 14C-carbonate working solution (see Fig. 2.1). The combusted
sample is transferred into the 0.5-0.7-1 filtration flask, at the bottom of which
the bar of the magnetic stirrer is placed. The flask is closed with a rubber
stopper supplied with a glass tube with a small funnel at its top end. Under
the stopper to this tube a beaker is attached containing 3-5 ml of 5 % hyamine
solution in methanol, which serves as CO2 absorber after closing. The flask is
slightly emptied, and 5 ml 20% H 2S04 is added via this tube to the combusted
sample. The labeled CO2 evoluting from the acidified sample is consumed by
the hyamine solution. To accelerate this procedure, the contents of flask are
stirred with a magnetic stirrer. This device is kept for 1-2 days. Then the contents of the beaker is transferred into the scintillation vial and the radioactivity of the 14C02 consumed is assayed after adding the scintillation cocktail. To
control the wet combustion and to make the necessary correction, two or three
portions of labeled protein hydrolyzate with a known radioactivity (Ro) are
also subjected to the same wet combustion procedures, being dissolved in
50 ml of water. Their radioactivity (Rs) is measured after combustion. The
ratio RslRo = Kn is then used to correct the results of exudate combustion
by the appropriate techniques, including the alkaline-persulfate combustion
described above.
The loss in 14C-assimilates due to filtration (Kex) can then be calculated as
follows: Kex = (RI - R d) VIKn Ra 50, if RI and Rd are average radioactivity of
BaC03 on filters after CO2 distillation of 50-ml combusted samples of filtrates,
after filtration of light and dark incubations, cpm; V is the total volume of
filtrate, measured before subsampling; Kn is the correction coefficient for combustion completeness; Ra the joint radioactivity of phytoplankton on filters
from light incubation corrected for dark 14C02 uptake, cpm; and 50 the volume
of subsamples combusted, ill1. Then the Kc correction coefficients for the loss
in 14C-assimilates during filtration will be equal to Kc = 1 + Kex. Examples of
Kc-coefficients are given in Table 2.2.
47
be used in the absorber. In this case, it is discharged directly into the scintillation vial and radioassayed after addition of the scintillation cocktail (Sorokin
1987).
The alkaline - acid persulfate combustion described above can be
replaced by an alternative UV-combustion method (Armstrong et a1. 1966;
'lbomas 1971; Williams and Yentsch 1976; Williams et a1. 1976). The combustion of labeled organic matter to 14C02 in this case proceeds under the prolonged action of strong ultraviolet radiation in the presence of oxidizers such
as hydrogen peroxide or persulfate. For UV-combustion, the subsamples of
filtrate 50 or 100mi after extraction of labeled CO2 from them are neutralized
to pH 9.5 and placed in test tubes of pure fused quartz. Then two to three
drops of 30% H 20 2 or 0.5 g dry Na-persulfate are added to them. They are
closed with glass stoppers or glass balls and illuminated under a 1200-W round
mercury lamp (Hanovia 189, with fan) for 3-4h. After cooling, the 14C02 thus
formed is extracted from the samples as described above in the apparatus, or
using a simple device described by Thomas (1971), which resembles that used
for cleaning 14C-carbonate working solution (see Fig. 2.1). The combusted
sample is transferred into the 0.5-0.7-1 filtration flask, at the bottom of which
the bar of the magnetic stirrer is placed. The flask is closed with a rubber
stopper supplied with a glass tube with a small funnel at its top end. Under
the stopper to this tube a beaker is attached containing 3-5 ml of 5 % hyamine
solution in methanol, which serves as CO2 absorber after closing. The flask is
slightly emptied, and 5 ml 20% H 2S04 is added via this tube to the combusted
sample. The labeled CO2 evoluting from the acidified sample is consumed by
the hyamine solution. To accelerate this procedure, the contents of flask are
stirred with a magnetic stirrer. This device is kept for 1-2 days. Then the contents of the beaker is transferred into the scintillation vial and the radioactivity of the 14C02 consumed is assayed after adding the scintillation cocktail. To
control the wet combustion and to make the necessary correction, two or three
portions of labeled protein hydrolyzate with a known radioactivity (Ro) are
also subjected to the same wet combustion procedures, being dissolved in
50 ml of water. Their radioactivity (Rs) is measured after combustion. The
ratio RslRo = Kn is then used to correct the results of exudate combustion
by the appropriate techniques, including the alkaline-persulfate combustion
described above.
The loss in 14C-assimilates due to filtration (Kex) can then be calculated as
follows: Kex = (RI - R d) VIKn Ra 50, if RI and Rd are average radioactivity of
BaC03 on filters after CO2 distillation of 50-ml combusted samples of filtrates,
after filtration of light and dark incubations, cpm; V is the total volume of
filtrate, measured before subsampling; Kn is the correction coefficient for combustion completeness; Ra the joint radioactivity of phytoplankton on filters
from light incubation corrected for dark 14C02 uptake, cpm; and 50 the volume
of subsamples combusted, ill1. Then the Kc correction coefficients for the loss
in 14C-assimilates during filtration will be equal to Kc = 1 + Kex. Examples of
Kc-coefficients are given in Table 2.2.