Hawaii Ocean Time-series (HOT)
in the School of Ocean and Earth Science and Technology at the University of Hawai'i


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SUMMARY: Seawater samples are collected at discrete depths in 12-liter Niskin bottles. The water samples are prefiltered (202 µm) and transferred to specially designed, precalibrated filtration bottles, pressure filtered through combusted GF/F filters and stored frozen for subsequent analysis. In the laboratory, the filters are dried and analyzed for C and N using a Perkin-Elmer model 2400 analyzer.

1. Principle

Particulate carbon (PC) and particulate nitrogen (PN), including both inorganic and organic forms, are present in seawater primarily as by-products of biological activity. Living organisms also contribute a variable amount (<1% in deep waters to >50% in the euphotic zone) to the total particulate carbon and nitrogen content of seawater.

Both PC and PN can be measured using commercially-available instruments which detect the gaseous by-products of high-temperature combustion. The Perkin-Elmer model 2400 CHN analyzer used in the HOT-GOFS project, combines the classical Pregal and Dumas methods for the determination of carbon and nitrogen, respectively. The samples are combusted in a pure oxygen environment, the gases mixed and separated by frontal chromatography. The separated gases are determined stepwise by a thermal conductivity detector.

2. Precautions

Care must be taken to ensure that contamination with PC and PN is avoided. Samples should, therefore, be kept away from paper, wood, food or other carbon-containing compounds. The filtration system that is set up to collect these samples is constructed of plastic. Plastic contains carbon and, therefore, is a potential source of contamination. The carboys and tubing used in this system must be kept clean and free of abrading surfaces.

3. Sampling, Filtration and Storage

3.1. Seawater samples are collected in 12-liter Niskin bottles and transferred directly to acid-cleaned filtration bottles. The samples are transferred via Tygon tubing which incorporates an in-line 202 µm Nitex screen prefilter to remove zooplankton or any other rare particles which might otherwise affect the precision of the estimate. The filtration bottles are 4- and 12-liter polyethylene aspirator bottles fitted with a valve assembly and tubing connection.

Once the rosette sampler is on deck, the vent valve of each Niskin bottle is opened and one end of the drawing tube is attached to the outflow spigot of the sampling bottle and the other end to the tubing connector on the cap of the filtration bottle. Particular attention is paid to the orientation of the in-line screened drawing tube (the shorter, larger bore section is attached to the Niskin bottle). The filtration bottle valve is opened and 100-200 ml is run through the transfer tube and valve assembly to rinse the sampling bottle. The bottle and cap are rinsed 3 times in this fashion. During the rinsing and filling operation, the filtration bottle cap valve is used to control and direct the sample flow. After rinsing,the cap is placed on the bottle mouth (without tightening), the valve is opened and the polyethylene bottled is filled to the calibration mark.

3.2. After filling, the filtration bottles are inverted and placed into the filtration rack. The contents are then pressure filtered (4-7 psi nitrogen gas) through combusted in-line 25 mm GF/F filters.
3.3. Following filtration, clean forceps are used to transfer each filter to a clean plastic petri dish containing a 3 cm square piece of combusted foil. The sample is labeled and stored frozen (-20°C). Usually only a few ml or less of water remains in the carboy after filtration; this residual water is ignored in assessing the volume filtered unless it appears to be significant compared to the volume of the carboy. All appropriate data are entered on the data sheet.

4. Analysis

Standard procedures for instrument warm-up are followed. Primary PC/PN standards are prepared using acetanilide (C8H9NO; mol. wt. = 135.16). Standards are made to encompass the range 20-500 ug/sample. Blanks are prepared by analyzing combusted GF/F "field filters" and the mean value subtracted from the sample value. Typical ranges in analytical blanks, expressed as percentages of the total signal produced by a sample in the course of a given analysis, are 5-17% for both PC and PN.

5. Data Reduction and Calculations

External standard data are used to prepare a standard curve of C (or N) versus corrected signal counts, and linear regression statistics are calculated. An EXCEL spreadsheet is used to calculate PC and PN (µg l-1) for each sample based on standard curve, corrected signal counts and volume of seawater filtered.

6. Precision and Accuracy

Replicate samples are routinely analyzed to estimate the precision of our PC and PN analyses. The average coefficients of variation for such replicate analyses are 12% and 10%, respectively. Accuracy is estimated from determinations of the C and N contents of reference standards analyzed along with samples during each analytical run. This accuracy, expressed as the mean percent difference between the analytical determination and the known C and N contents of the standards, is found to be 6% for PC and 13% for PN. Note that the estimates of accuracy apply only to standards containing >10 µg N and >70 µg C. Below these levels, we have found the performance of the PE-2400 to be suspect. Therefore sample volumes large enough to keep above this lower limit are used.

7. Equipment/Supplies

  • PE-2400 (or equivalent) Carbon/Nitrogen analyzer with integrator

  • Cahn (or equivalent) electronic balance

  • combusted 2.5 cm GF/F filters (450°C, 4 hours)

  • ethanol-cleaned forceps for handling filters

  • combusted foil (450°C, 4 hours)

  • aluminum foil for covering work area

  • clean petri dishes

  • low pressure filtration apparatus (4-7 psi)

  • valved polyethylene aspirator bottles

8. Reagents

  • acetanilide standard

9. References

  • Sharp, J. H. 1974. Improved analysis for particulate organic carbon and nitrogen from seawater. Limnology and Oceanography, 19, 984-989.