Garlmer River Data



AUTHORS/CONTACTS: Joan E. Sheldon and Dr. William J. Wiebe

Revision Date: June 7, 1997

Initially we tried the standard Koroleff (1976, 1983) method which uses no magnesium reagent and instead uses citrate to complex any natural magnesium and calcium and thus avoid a precipitate. Humics remained in solution and interfered with recovery of ammonium. A color blank using nitroprusside solution without phenol (Flebbe, 1982) could not completely correct for the interference: "corrected" recoveries of ammonium added to humic- colored water were low.

Sample Volume:
15 mL
Sample Collection:
Filter through ashed GF/F filter in glass or plastic apparatus, ashed or used only for similar samples.
Sample Storage:
In polyethylene bottle at -20 oC or lower

Magnesium reagent:
Dissolve 45 g NaCl and 20 g MgSO4.7H2O in about 200 mL water. Add 1 N NaOH dropwise until a slight precipitate forms. Add a few boiling chips and boil (to drive off ammonia) until < 200 mL. Cool and dilute to 200 mL. A slight precipitate will remain. Use an automatic dispenser.
Dissolve 0.2 g Na2Fe(CN)6NO.2H2O and 19 g C6H5OH in 500 mL water. Use an automatic dispenser. Store in a dark bottle in refrigerator.
Dilute 750 mg available Cl- as Clorox® bleach (see protocol below) to 500 mL with 0.5 N NaOH. Use an automatic dispenser. Store in a dark bottle in refrigerator.
Dissolve 7.9 g Na2S2O3 or 12.4 g Na2S2O3.5H2O in 500 mL water. Store in refrigerator.
Potassium Iodide:
Dissolve 0.5 g KI in 50 mL 1 N H2SO4. Make fresh each time.

10 mM Primary:
Dry (NH4)2SO4 overnight at 50 oC. Dissolve 0.6607 g and make up to 1 L with water. Store in refrigerator.
1 mM Secondary:
1 mL Primary Standard + 9 mL water, or similar convenient amount. Make fresh each day.
Make fresh each day.
5 uM: 4.975 mL water + 25 uL Secondary Std.
10 uM: 4.950 mL water + 50 uL Secondary Std.
20 uM: 4.900 mL water + 100 uL Secondary Std.

Pyrex®, test tubes, 20 x 150 mm, with Teflon®-lined screw caps, kept only for this procedure. Before first use, wash and dry tubes and caps and ash tubes as usual, then run reagent blanks in all tubes to "burn out" any remaining ammonia. After each use, leave any remaining sample in tube, cap, and keep until next use.

Basically the Koroleff (1983) method but using slightly more than half the suggested phenol and hypochlorite (Koroleff, 1976; D. Bronk, pers. comm.) to reduce phenol vapors in the lab. The Mg hydroxide precipitation procedure is used to clear humic color from the water. The sum of NH4++ NH3 is determined.
Available Cl-:
Make up potassium iodide solution and add 1 mL Clorox®. Titrate with thiosulfate solution until color is gone. 1 mL thiosulfate reacts with 3.54 mg available Cl-, so (mL Clorox® needed to give 750 mg available Cl-) = (750 ÷ 3.54) ÷ (mL thiosulfate used). Discard Clorox® if < 12 mL thiosulfate is used (Parsons, et al. 1984).
Discard old sample in test tubes (collect as hazardous waste), rinse tubes 4 times with 18 megohm-cm water, and shake out as much water as possible. Small drops will remain, but these are fairly consistent among all tubes and thus will not affect the analysis.

Make up triplicate blanks and working standards in tubes, 5 mL each. Pipette 5 mL of each sample into 3 tubes, noting the salinity of each sample. Add 100 uL of magnesium reagent to each blank, standard, and sample with salinity <= 5 ppt and vortex. To each tube add 250 uL nitroprusside/phenol, vortex, immediately add 250 uL hypochlorite, vortex, and cap. Keep tubes in a dark place until read at least 6 hours but not more than 30 hours later.

Handle carefully to avoid resuspending the precipitate. Read the absorbance in a 1 cm cell at 630 nm (against the absorbance of water), using a sample sipper or Pasteur pipette to draw off the supernatant. Settings for the sample sipper for the Shimadzu Model UV160U spectrophotometer are pump speed medium, sip time 4 sec, dwell time 1 sec, purge time 4 sec, 1 rinse. If using a conventional cell and Pasteur pipette, rinse once with sample.

Corrections for salinity are obtained by running standards in waters of various salinities. For example, salt factors were obtained for 0, 5, 10, ... 34 ppt water made by mixing aged Gulf Stream water with 18 megohm-cm water, and factors for other salinities were interpolated. Salt factor = 1 + (salinity in ppt) × 0.0062

Correct standards for dilution by magnesium reagent because not all samples will receive it (e.g. 20 uM standard is really 19.61 uM after addition of magnesium reagent).

Plot the corrected standard concentration vs. absorbance. The slope of this line is the calibration factor F. F should be approximately 55-60 for a 1 cm cell.

Correct sample calculations for dilution by magnesium reagent where necessary: Dilution factor = 1.02 if sample salinity was <= 5 ppt (i.e. magnesium reagent was added) or Dilution factor = 1 if sample salinity was > 5 ppt (i.e. no magnesium reagent).

Sample concentration (uM) = F × Salt factor × (sample abs - average blank abs) × Dilution factor

Limit of Detection:
2 std. dev. above blank approx. = 0.1 uM

approx. 40 uM directly or up to 150 uM if diluted after color development (Koroleff, 1983).
95% confidence limits for prediction of a concentration near the mean for a typical standard curve (8.75 uM) approx. = ± 0.48 uM for a triplicate determination. (Confidence limits for prediction near the ends of a standard curve are, of course, somewhat larger.) For some trials with ammonium-spiked river water, recoveries were still 10-20% lower than expected but these results showed no pattern with humic fluorescence or salinity, so perhaps other factors such as heavy metals interfere with the analysis in some rivers (Koroleff, 1983).

Flebbe, P. 1982. Biogeochemistry of carbon, nitrogen, and phosphorus in the
aquatic subsystem of selected Okefenokee Swamp sites. Okefenokee Ecosystem Investigations no. 8.

Koroleff, F. 1976. Determination of ammonia. Pages 126-133 in Grasshoff, K.,
editor. Methods of seawater analysis. Verlag Chemie, Weinheim.

Koroleff, F. 1983. Determination of ammonia. Pages 150-157 in Grasshoff, K., M.
Ehrhardt, and K. Kremling, editors. Methods of seawater analysis: second, revised and extended edition. Verlag Chemie, Weinheim.

Parsons, T.R., Y. Maita, and C.M. Lalli. 1984. A manual of chemical and biological
methods for seawater analysis. Pergamon Press, New York.