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4500-Norg NITROGEN (ORGANIC)*

* Approved by Standard Methods Committee, 1997.

 

4500-Norg B. Macro-Kjeldahl Method

 

1. General Discussion

 

    a. Principle: In the presence of H2SO4, potassium sulfate (K2SO4), and cupric sulfate (CuSO4) catalyst, amino nitrogen of many organic materials is converted to ammonium. Free ammonia also is converted to ammonium. After addition of base, the ammonia is distilled from an alkaline medium and absorbed in boric or sulfuric acid. The ammonia may be determined colorimetrically, by ammonia-selective electrode, or by titration with a standard mineral acid.

    b. Selection of ammonia measurement method: The sensitivity of colorimetric methods makes them particularly useful for determining organic nitrogen levels below 5 mg/L. The titrimetric and selective electrode methods of measuring ammonia in the distillate are suitable for determining a wide range of organic nitrogen concentrations. Selective electrode methods and automated colorimetric methods may be used for measurement of ammonia in digestate without distillation. Follow equipment manufacturer's instructions.

 

2. Apparatus

 

    a. Digestion apparatus: Kjeldahl flasks with a total capacity of 800 mL yield the best results. Digest over a heating device adjusted so that 250 mL water at an initial temperature of 25C can be heated to a rolling boil in approximately 5 min. For testing, preheat heaters for 10 min if gas-operated or 30 min if electric. A heating device meeting this specification should provide the temperature range of 375 to 385C for effective digestion.

    b. Distillation apparatus: See Section 4500-NH3.B.2a.

    c. Apparatus for ammonia determination: See Section 4500-NH3.C.2, D.2, F.2, or G.2.

 

3. Reagents

 

    Prepare all reagents and dilutions in ammonia-free water.

   All of the reagents listed for the determination of Nitrogen (Ammonia), Section 4500-NH3.C.3, D.3, F.3, or G.3, are required, plus the following:

    a. Digestion reagent: Dissolve 134 g K2SO4 and 7.3 g CuSO4 in about 800 mL water. Carefully add 134 mL conc H2SO4. When it has cooled to room temperature, dilute the solution to 1 L with water. Mix well. Keep at a temperature close to 20C to prevent crystallization.

    b. Sodium hydroxide-sodium thiosulfate reagent: Dissolve 500 g NaOH and 25 g Na2S2O3 5H2O in water and dilute to 1 L.

    c. Borate buffer solution: See Section 4500-NH3.B.3b.

    d. Sodium hydroxide, NaOH, 6N.

 

4. Procedure

 

    a. Selection of sample volume and sample preparation: Place a measured volume of sample in an 800-mL kjeldahl flask. Select sample size from the following tabulation:

                  

  Organic Nitrogen  
  in Sample Sample Size
  mg/L mL
   0-1 500
    1-10 250
  10-20 100
  20-50     50.0
  50-100     25.0

 

       If necessary, dilute sample to 300 mL, neutralize to pH 7, and dechlorinate as described in Section 4500-NH3.B.4b.

    b. Ammonia removal: Add 25 mL borate buffer and then 6N NaOH until pH 9.5 is reached. Add a few glass beads or boiling chips such as Hengar Granules #12 and boil off 300 mL. If desired, distill this fraction and determine ammonia nitrogen. Alternately, if ammonia has been determined by the distillation method, use residue in distilling flask for organic nitrogen determination.

    For sludge and sediment samples, weigh wet sample in a crucible or weighing bottle, transfer contents to a kjeldahl flask, and determine kjeldahl nitrogen. Follow a similar procedure for ammonia nitrogen and organic nitrogen determined by difference. Determinations of organic and kjeldahl nitrogen on dried sludge and sediment samples are not accurate because drying results in loss of ammonium salts. Measure dry weight of sample on a separate portion.

    c. Digestion: Cool and add carefully 50 mL digestion reagent (or substitute 6.7 mL conc H2SO4, 6.7 g K2SO4, and 0.365 g CuSO4) to distillation flask. Add a few glass beads and, after mixing, heat under a hood or with suitable ejection equipment to remove acid fumes. Boil briskly until the volume is greatly reduced (to about 25 to 50 mL) and copious white fumes are observed (fumes may be dark for samples high in organic matter). Then continue to digest for an additional 30 min. As digestion continues, colored or turbid samples will become transparent and pale green. After digestion, let cool, dilute to 300 mL with water, and mix. Tilt flask away from personnel and carefully add 50 mL sodium hydroxide-thiosulfate reagent to form an alkaline layer at flask bottom. Connect flask to a steamed-out distillation apparatus and swirl flask to insure complete mixing. The pH of the solution should exceed 11.0.

    d. Distillation: Distill and collect 200 mL distillate. Use 50 mL indicating boric acid as absorbent solution when ammonia is to be determined by titration. Use 50 mL 0.04N H2SO4 solution as absorbent for manual phenate or electrode methods. Extend tip of condenser well below level of absorbent solution and do not let temperature in condenser rise above 29C. Lower collected distillate free of contact with condenser tip and continue distillation during last 1 or 2 min to cleanse condenser.

    e. Final ammonia measurement: Use the titration, ammonia-selective electrode, manual phenate, or automated phenate method, Sections 4500-NH3.C, D, F, and G, respectively.

    f. Standards: Carry a reagent blank and standards through all steps of the procedure.

 

5. Calculation

 

    See Section 4500-NH3.C.5, D.5, F.5, or G.5.

 

Table 4500-Norg:I. Precision Data for Kjeldahl Nitrogen Method

Based on Mean of Triplicate Analyses of

Nicotinic Acid

 

  Relative
Nicotinic Recovery of  Standard Standard
Lab/ Acid N Deviation Deviation
Analyst mg N/L % mg/L  %
1/1 5     93.3 0.16    3.46
1/2 5 101  0.16    3.17
1/1 10     87.7  0.16    1.84
1/2 10     91.5 0.28    3.06
1/1 20     95.7 0.16    0.84
1/2 20     95.7 0.58    3.03
2/1 0.5     97.4  0.005    1.04
2/2 0.5     95.3  0.027    5.46
3/1 0.5     87.3  0.130 29.9
4/1 0.5 113  0.235 41.7
2/1 1.0 103  0.012    1.15
2/2 1.0 101  0.046    4.63
3/1 1.0     84.3  0.081    9.66
4/1 1.0     99.3  0.396 39.9
2/1 2.0 104 0       0  
2/2 2.0     99.2  0.029    1.44
3/1 2.0     89.2  0.071    3.98
4/1 2.0 112  0.139    6.18

                                                                    

6. Precision and Bias

 

    Two analysts in one laboratory prepared reagent water solutions of nicotinic acid and digested them by the macro-kjeldahl method. Ammonia in the distillate was determined by titration. Results are summarized in Table 4500-Norg:I.

 

 

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7. Bibliography

 

Kjeldahl, J. 1883. A new method for the determination of nitrogen in organic matter. Z. Anal. Chem. 22:366.

 

Phelps, E.B. 1905. The determination of organic nitrogen in sewage by the Kjeldahl process. J. Infect. Dis. (Suppl) 1:225.

 

McKenzie, H.A. & H.S. Wallace. 1954. The Kjeldahl determination of nitrogen: A critical study of digestion conditions. Aust. J. Chem. 7:55.

 

Morgan, G.B., J.B. Lackey & F.W. Gilcreas. 1957. Quantitative determination of organic nitrogen in water, sewage, and industrial wastes. Anal. Chem. 29:833.

 

Boltz, D.F., ed. 1978. Colorimetric Determination of Nonmetals. Interscience Publishers, New York, N.Y.

 

Jones, M. & D. Bradshaw. 1989. Copper: An alternative to mercury; more effective than zirconium in kjeldahl digestion of ecological materials. Commun. Soil Sci. Plant Anal. 20:1513.

 

 

Standard Methods for the Examination of Water and Wastewater. 20th Ed. American Public Health Association, American Water Works Association, Water Environment Federation.