Faults in the Radiocarbon Method and Problems in its Calibration
67
part of the smallest cells of picocyanobacteria may pass through the membrane
or glass fiber filter with pores of 0.6-0.8 mm, such as are most often used for
primary production experiments. Finally, part of the labeled photosynthesis
assimilates can be used by algae as material for respiration and respired as
14C02. These latter losses are unimportant in a short-term 2- to 4-h incubation,
but might become significant after 12-h exposures (Bidwell 1977).
5. Possible losses of 14C assimilated by algae not held back on filters
during filtration of experimental samples, are evaluated by numerous authors
as between 0 and 60% of the total 14C carbon assimilated during incubation
(Fogg 1958, 1966; Fogg et al. 1965; Thomas 1971; Ryther et al. 1971; Berman
and Holm Hansen 1974; Jensen 1983; Lignell1990; Pujo-Pay et al. 1997). The
great variability in results reflects, in part, the insufficiency of the methods for
measuring radioactivity of 14C organic matter in filtrates. The faults were in
impurities contained in the working solution of 14C-carbonate, in inadequacy
in the technique of 14C02 extraction from the filtrates, in the use of too small
volumes of filtrates (3-6ml) for direct measurements of the radioactivity or
14C-labeled organic matter in filtrates (Thomas 1971; Williams and Yentsch
1976; Sharp 1977). Nevertheless, a selection of data has been obtained by a
more substantial method (see Table 2.2). Thus, if the researcher does not have
the possibility of assessing these losses personally, I advise using this table. It
will be a better approximation to the correct numbers than ignoring filtration
losses, which are considerable.
During incubation experiments to estimate filtration losses of assimilated
14C, a significant part (to 30%) of labeled organic matter excreted by algal cells
can be respired by bacteria even during short-term incubations (Jensen 1985).
In this case, injection of antibiotics into the samples (20mgl- 1 streptomycin or
1 mg I-I metoxithin + vancomycin) is recommended half an hour before charging them with 14C-bicarbonate (Jensen and Sondergaard 1985). Before hand,
the researcher must, however, obtain experimental evidence that the antibiotic does not inhibit the photosynthesis of phytoplankton in the samples of
water from the basin under investigation. Another matter concerns the selection of optimal filters for local working conditions. The filters should be
selected by the researcher himself experimentally. First of all it is necessary to
establish the minimal pore size of the filters to give a maximal gain in radioactivity (with dark controls subtracted). Selection from a series of filters with a
decreasing scale of pore size of the kind of filter which combines maximum
gain in radioactivity with an acceptable rate of filtration: some 5 min for a
sample of 250ml with a filter of diameter 20mm.
For an approximate evaluation of losses of 14C due to filtration of samples,
a rather simple procedure can also be recommended (Bullion 1975). After
incubation of water samples with an enlarged charge of 14C-bicarbonate, they
are normally filtered through filters of the kind otherwise used for primary
production estimations. The filtrates in portions of 20-30 ml are then refiltered
through an extradense membrane filter with 0.05-mm pores. After filtration,
the filters are placed into scintillation vials, treated with 0.2 ml 0.1 N HCI, and
67
part of the smallest cells of picocyanobacteria may pass through the membrane
or glass fiber filter with pores of 0.6-0.8 mm, such as are most often used for
primary production experiments. Finally, part of the labeled photosynthesis
assimilates can be used by algae as material for respiration and respired as
14C02. These latter losses are unimportant in a short-term 2- to 4-h incubation,
but might become significant after 12-h exposures (Bidwell 1977).
5. Possible losses of 14C assimilated by algae not held back on filters
during filtration of experimental samples, are evaluated by numerous authors
as between 0 and 60% of the total 14C carbon assimilated during incubation
(Fogg 1958, 1966; Fogg et al. 1965; Thomas 1971; Ryther et al. 1971; Berman
and Holm Hansen 1974; Jensen 1983; Lignell1990; Pujo-Pay et al. 1997). The
great variability in results reflects, in part, the insufficiency of the methods for
measuring radioactivity of 14C organic matter in filtrates. The faults were in
impurities contained in the working solution of 14C-carbonate, in inadequacy
in the technique of 14C02 extraction from the filtrates, in the use of too small
volumes of filtrates (3-6ml) for direct measurements of the radioactivity or
14C-labeled organic matter in filtrates (Thomas 1971; Williams and Yentsch
1976; Sharp 1977). Nevertheless, a selection of data has been obtained by a
more substantial method (see Table 2.2). Thus, if the researcher does not have
the possibility of assessing these losses personally, I advise using this table. It
will be a better approximation to the correct numbers than ignoring filtration
losses, which are considerable.
During incubation experiments to estimate filtration losses of assimilated
14C, a significant part (to 30%) of labeled organic matter excreted by algal cells
can be respired by bacteria even during short-term incubations (Jensen 1985).
In this case, injection of antibiotics into the samples (20mgl- 1 streptomycin or
1 mg I-I metoxithin + vancomycin) is recommended half an hour before charging them with 14C-bicarbonate (Jensen and Sondergaard 1985). Before hand,
the researcher must, however, obtain experimental evidence that the antibiotic does not inhibit the photosynthesis of phytoplankton in the samples of
water from the basin under investigation. Another matter concerns the selection of optimal filters for local working conditions. The filters should be
selected by the researcher himself experimentally. First of all it is necessary to
establish the minimal pore size of the filters to give a maximal gain in radioactivity (with dark controls subtracted). Selection from a series of filters with a
decreasing scale of pore size of the kind of filter which combines maximum
gain in radioactivity with an acceptable rate of filtration: some 5 min for a
sample of 250ml with a filter of diameter 20mm.
For an approximate evaluation of losses of 14C due to filtration of samples,
a rather simple procedure can also be recommended (Bullion 1975). After
incubation of water samples with an enlarged charge of 14C-bicarbonate, they
are normally filtered through filters of the kind otherwise used for primary
production estimations. The filtrates in portions of 20-30 ml are then refiltered
through an extradense membrane filter with 0.05-mm pores. After filtration,
the filters are placed into scintillation vials, treated with 0.2 ml 0.1 N HCI, and