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Penentuan kadar protein

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					Determination of Protein Content




   SMK Negeri 13 Bandung
        Protein in Human Body
 Protein has many function in human body:
  - Energy source
  - Tissue and cell reparation and development
  - As antibody and hormone and enzym sinthesys
  - As leveling substance for acid and base balance
    in cell
 In human body, protein through a cycle in which
  protein decomposed to smaller component (amino
  acid/peptide). Amino acid and peptide form new
  protein molecules to replaced the broken one.
  (There is no life long protein used)
Protein in Food
        According to the source, 2
         kinds of protein known:
         - Animal Protein
           (fish, meat, egg, milk)
         - Plant Protein
           (Beans, rice, wheat and
            several fruits)
        Protein content in food
         stuff are various
            Protein

One of big biomolecules besides
 polysacharide, lipid and polynucleotid
 which are the main former of live
 creature
Protein is a polymer of about 20
 amino acid linked by the peptide
 bonding
          Amino Acid
Amino acid is the main molecule of protein
 former/compiler
Amino acid is an organic substance with
 carboxyl (COOH) and amine (NH2) group.
 Amino acid structure in general is a C
 atom linked to 4 groups/atom: amine
 (NH2), gugus carboxcyl (COOH), hydrogen
 atom (H), and one rest group (R, residue).
 R group distinguish an amino acid with
 another.
AMINO ACID STRUCTURE




                       C
        PEPTIDE BONDING
 Peptide bonding is amide bonding which is link 2
  amino acids. One peptide has end-N amino acid with
  free NH3+ group and end-C amino acid with free COO-
  group.
ANALYSIS OF PROTEIN CONTENT

 The determination of N developed by
  Johan Kjeldahl (Danish chemist), that’s
  why it called Kjeldahl’s method
 The Kjeldahl method of nitrogen analysis
  is the worldwide standard for calculating
  the protein content in a wide variety of
  materials ranging from human and
  animal food, fertilizer, waste water and
  fossil fuels.
Kjeldahl’s Method Step
Diagram of Kjeldahl’s Method
Kjeldahl’s Apparatus




         http://www.brooklyn.cuny.edu/
                  Kjeldahl’s Steps

Decomposition   Distillation   Trapping   Titration
                Calculation

Nitrogen calculation       Protein calculation
                            It is possible to calculate
                             the amount of crude
                             protein in the sample.
                             Although there are
                             differences between
                             different samples, the
                             amount of "crude protein"
                             (CP) can be found by
                             multiplying the percent
                             Nitrogen by a factor
                             (usually 6.25).
                            CP = %N x 6.25

                       http://www.brooklyn.cuny.edu/
                  Digestion
 Weighing out approximately 1 gm of the sample
  containing protein, and placing the sample into a
  digestion flask, along with 12-15 ml of concentrated
  sulfuric acid (H2SO4).
 Adding seven grams of potassium sulfate and a
  catalyst, usually copper.
 Bringing the digestion tube/flask and mixture to a
  "rolling boil" (about 370oC to 400oC) using a heating
  a block.
 Heating the mixture in the tube/flask until white
  fumes can be seen, and then continuing the heating
  for about 60-90 mins, the solution should be clear.
 Cooling the tube/flask and cautiously adding 250 mls
  of water.
                 Distillation
 Raising the pH of the mixture using sodium
  hydroxide (45% NaOH solution). This has the effect
  of changing the ammonium (NH4+) ions (which are
  dissolved in the liquid) to ammonia (NH3), which is a
  gas.
 Separating the nitrogen away from the digestion
  mixture by distilling the ammonia (converting it to a
  volatile gas, by raising the temperature to boiling
  point) and then trapping the distilled vapors in a
  special trapping solution of about 15 ml HCl
  (hydrochloric acid) in 70 ml of water.
 removing the trapping flask and rinsing the
  condenser with water so as to make sure that all the
  ammonia has been dissolved.
                Titration
 Adding an indicator dye to the
  acid/ammonia trapping solution.
 Putting a standard solution of NaOH
  (sodium hydroxide) into the buret slowly
  adding small amounts of the sodium
  hydroxide solution to the acid solution with
  the dye.
 Watching for "endpoint" has been reached
  and that now all the acid has been
  neutralized by the base.
 Performing a calculation to find the amount
  of ammonia, and thus nitrogen.
             Calculation




 Moles ammonia represent moles nitrogen
 gms nitrogen =
  moles nitrogen x atomic mass
  (gN = molesN x 14.0067)
%nitrogen =
 (gms nitrogen / gms sample) x 100
 %N = (gN / gS) x 100

				
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posted:1/2/2012
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