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Determination of Particulate Organic Carbon (POC) and Nitrogen (PON)
in Seawater
John H. Martin
- Scope and Field of Application
This procedure describes a method for the determination of particulate
organic carbon (POC) and nitrogen (PON) in seawater. The assay is appropriate
for measuring open ocean and coastal concentrations of 1.0 to 800 µg
POC/l and 0.1 to 160 µg PON/l, provided that a large enough sample
is provided.
- Definition
- The particulate organic carbon content is defined as µg
POC /l seawater.
- The particulate organic nitrogen content is defined as µg
PON /l seawater.
- Principle
A dried sample of particulate matter (on a filter or freeze dried) is
combusted at 900 to 1,000° C. The organic carbon is converted into
CO
and the organic nitrogen
into N
gas; these are separated on a column and measured by thermal conductivity.
- Apparatus
Any analyzer with high temperature combustion and subsequent detection
of CO
and N
by TCD gas chromatography can be used. None of the commercially available
analyzers is particularly recommended.
- Reagents
- Hydrochloric acid (HCL: reagent grade)
- Acetanilide, EDTA (reagent grade) or other non hydrophile
substances with a C:N ratio close to those of the samples are used
as standards.
- Sampling
- General Precautions
During the entire procedure of sampling, filtration, storage,
fuming and analysis, care must be taken to avoid contamination by
both organic and inoranic forms of carbon and nitrogen. Acid cleaning
and rinsing with deionized water of water samplers, filtration bottles,
filtration device and storage containers is recommended. During
storage, particularly when deep frozen, samples must not be exposed
to other sources of volatile carbon and nitrogen.
- Sample Acquisition
A representative aliquot of the water from the sampler or the
contents of the entire sampler is taken. In the former case homogeneity
of the entire sample must be maintained; e.g., by gentle agitation
prior to the taking of an aliquot. In the latter case all water
must be taken from the bottle including the dregs. Sample size is
chosen in accordance to the sensitivity of the analyzer and
in situ concentrations of the respective environment under investigation
(0.5 to 2 l in the euphotic zone, 5 to 10 l in deep water). For
deep water sampling it is preferred to take duplicates or triplicates
to achieve good numbers rather than daily sampling of one sample
of discrete depths.
Prescreening of water samples may exclude mucilagious or chain-forming
algae and large faecal pellets and strings. If prescreening is necessary
due to zooplankton and/or infrequent large particles a 200 micron
mesh should be used and this must be reported in the protocols.
Removal of such particles from the filters is also recommended.
From sediment trap samples a liquid or freeze dried aliquot can
be used.
- Filtration
Glass fiber filters (GF/F) or quartz fiber filters are used. The
latter have the advantage of not melting in the C/N analyzer furnace
tube. Filters are precombusted in a muffle furnace for 4 hours at
500°C. With increasing sample size, retention of smaller particles
on fiber filters is increased. Therefore, the sample size should
be kept as constant as possible to maintain a consistent filter
loading. A pressure differential of less than 0.5 atm should be
used in pressue or vacuum filtration. To avoid cell lysis filters
must not run dry. Generally salt residues do not cause a problem,
therefore filters should not be rinsed with deionized water. Rinsing
with filtered seawater is useful when mucilagious particles are
sticking to the filtration device.
- Storage of Samples
Samples can be either kept deep frozen at -20°C or in a desiccator
after drying at 60°C for 5 hours. When drying filters in an
oven care must be taken to avoid contamination from volatile organics.
Samples from sediment traps can be stored as dried powder. Storage
time is nearly unlimited. In the case of deep-freezing samples are
desiccated before analysis.
- Procedures
- Acidification of Filters
In certain regions and seasons filter loads may contain carbonate
minerals amounting to a considerable portion of total particulate
carbon, e.g. coccolithophorid blooms, foraminifera, coral reef environments.
Removal of inorganic carbon can be done by fuming the dried filters
with concentrated hydrochloric acid for 24 hrs. Thereafter filters
are dried again. When rinsing the filters with acid, losses in particulate
organic carbon and nitrogen of up to 40 % have been observed. In
the case of acid rinsing of filters the rinse should be collected
and analyzed for dissolved organic carbon and nitrogen. When low
amounts of carbonate minerals in the samples are expected, acid
can be added directly to the sample capsule of the analyzer. Whether
samples were acidified or not and what mode of acidification was
used must be reported in the protocols.
- Standards
The standard should be not hydrophilic and should have a C/N ratio
close to that of the samples. Pure substances such as acetanilide
and EDTA, are easily analyzed and may not fully account for the
instruments performance for samples with more refractory components.
A reference standard containing more refractory components should
be prepared and used by the JGOFS community. Standards should be
run every 20 to 40 samples to check the drift of the analyzer.
- Blanks
The machine blank is usually included by the analyzer calibration.
A blank for filters, filtration, handling and storage is best achieved
by refiltering the same volume of filtered seawater as for the samples
through a precombusted filter and thereafter follow exactly the
same procedure as for the samples. This should also account for
filtration artefacts such as adherence of dissolved organic matter
to the fibers and formation of colloids during filtration. One blank
should be run for every 10 samples.
- Calculation and Expression of Results
The calculation of the results is dependent on the computing software
linked to the analyzer. Carbon and nitrogen content of samples and blanks
are computed according to the results of the standard measurements. Thereafter
the blank is substracted from the sample. This number is divided by the
volume filtered. The results are given in µg/l and also in molar
units in accordance with the JGOFS standard formats.
- Quality Control/Quality Assessment
- Accuracy
The exact nature of organic particles on the filters is unknown.
Therefore the accuracy must be regarded as 100 % provided that the
analyzer is adequately calibrated and an appropriate standard is
used.
- Precision
Precision is defined as the replicability of a homogeneous standard
or sample material. Precision for the standard has to be within
±5 %. The limit of detection varies somewhat depending on the
instrument used. Samples should contain at least 10 micrograms of
nitrogen and 15 micrograms of carbon.
- Quality control
Between 10 and 20 % of the samples should be run as triplicates.
The coefficient of variation of replicate samples should be not
higher than ±10--15 %. Operation of the machine should mainly
follow the manual of the manufacturer. If changes are introduced
these should be discussed with the manufacturer and harmonized with
the colleagues using the same model.
- Notes
Notes about general precautions and possible modifications have been made
in the sections 5 to 9. Backward compatibility should be checked following
intercomparisons with other laboratories.
- Intercomparison
Intercomparison with one or more labs should be carried out whenever feasible.
Care must be taken that samples for such intercomparison come from the
same water bottle or cast and are handled identically prior to the analysis.
Bulk sediment trap samples, whenever available should also be used, to
achieve intercomparison for the more refractory part of POC/PON.
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