VisualChem The interactive chemistry textbook
中文

Protein by Kjeldahl nitrogen

Digest, distil and titrate: measure the nitrogen in milk, flour, peanuts and gelatin, convert it to protein with 6.25 and with Jones’s special factors, and see how much a “protein content” depends on the factor.

Difficulty ●●○ ≈ 25 min

Aim Determine the nitrogen in foods by the Kjeldahl method, subtract the blank correctly, and compare the protein contents given by different conversion factors.
Digest → distil into boric acid → titrate with 0.1000 M HCl · blank 0.10 mL

Kjeldahl nitrogen: how much protein is in milk

NXorganic→ΔHX2SOX4,CuNHX4X+\ce{N_{organic} ->[H2SO4, Cu][\Delta] NH4+} NHX3+B(OH)X3+HX2O→NHX4X++B(OH)X4X−\ce{NH3 + B(OH)3 + H2O -> NH4+ + B(OH)4-} B(OH)X4X−+HX+→B(OH)X3+HX2O\ce{B(OH)4- + H+ -> B(OH)3 + H2O}
Kjeldahl flask 05101520253035404550STIR50100150200
Burette reading 0.00mL
Receiver pH4.86

Kjeldahl flask: sample and conc. sulfuric acid

Digest, then distil, then titrate the borate in the receiver.

Each sample’s nitrogen is back-calculated from the protein listed as N × 6.25 in Table 6 of Jones (USDA Circular 183, 1931), itself from Atwater and Bryant’s food-composition tables; the conversion factors are from Table 5 of the same circular. Receiver: 50 mL of 4 % boric acid, about 100 mL after distillation; its pH uses the pKa of boric acid, 9.237, through the buffer equation, and is never more acidic near the end point than boric acid alone; the mixed-indicator colour is computed from the absorption spectra of methyl red and bromocresol green. The model assumes complete digestion and distillation, with one H⁺ consumed per NH₃.

Procedure

  1. Choose whole milk (5.000 g), digest it, then distil the NH₃ into boric acid.
  2. Titrate the receiver with 0.1000 mol/L HCl: 1 mL at a time at first, then drop by drop near the end point; record the reading when the indicator turns from green to grey-violet.
  3. Repeat for flour, peanuts and gelatin, recording %N and the protein from both factors.

What to notice

  • Milk reaches the end point near 18.95 mL: 0.528 % nitrogen, 3.30 % protein (× 6.25) or 3.37 % (× 6.38).
  • The receiver starts near pH 8.0 (a borate buffer), reaches about 4.9 at the end point, and drops below 4.4 two drops later.
  • Gelatin comes out near 91 % protein with 6.25 but only about 81 % with 5.55; peanuts fall from 25.8 % to 22.5 %, while milk rises slightly.

In a real lab

  • Digest in a fume cupboard: boiling concentrated sulfuric acid gives off SO₂ and white SO₃ fumes; keep heating for a while after the solution clears so that all the nitrogen is converted.
  • Cool and dilute the digest before adding NaOH, or it will boil violently; keep the end of the condenser below the surface of the boric acid so no ammonia escapes.
  • Run a blank with every batch, and check the recovery with a standard of known nitrogen content such as ammonium sulfate or acetanilide.

Related lessons

All experiments