METHOD 14:1

Printed with effect from 1st March 2014

CALCIUM - VOLUMETRIC METHOD

1: Scope and Field of Application

This method is for the determination of the total calcium content of feeding stuffs.

2: Principle
The sample is ashed, the ash treated with hydrochloric acid and the calcium precipitated
as calcium oxalate. The precipitate after filtering and washing is dissolved in acid and the
liberated oxalic acid is titrated with standard potassium permanganate solution.

3: Reagents
3:1 Ammonia (d=0.88g/ml).
3:2 Hydrochloric acid solution, 50% (V/V): dilute an appropriate volume of concentrated
hydrochloric acid (d=1.18g/ml) with an equal volume of water.
3:3 Nitric acid (d=1.42g/ml).
3:4 Ammonium chloride solution: 5g ammonium chloride per 100ml.
3:5 Ammonium oxalate, cold saturated solution.
3:6 Citric acid monohydrate solution: 30g citric acid monohydrate per 100ml.
3:7 Sulphuric acid solution, 20% (V/V): 20ml sulphuric acid (d=1.84g/ml) per 100ml.
3:8 Potassium permanganate solution, 0.1N.
3:9 Bromocresol green indicator solution: dissolve 0.04g of bromocresol green in 20ml
ethanol and dilute to 100ml with water.

4: Apparatus
4:1 Muffle furnace capable of being maintained at 550oC.
4:2 Platinum, silica or porcelain crucibles for ashing.
4:3 Glass filter crucibles, No.4 porosity.

5: Procedure
5:1 Dissolution of sample
Weigh to the nearest 0.001g, approximately 5g of the prepared sample into the crucible
(4:2) and incinerate at a temperature not exceeding 550oC until all the organic matter has
been destroyed. Allow to cool, moisten the ash with water and cautiously add 10ml of
hydrochloric acid (3:2), avoiding loss by use of a cover glass. Wash the cover glass with
water adding the washings to the crucible and evaporate to dryness. Continue the heating
for at least one hour in order to dehydrate any silica that might be present. Cool, add
20ml water, 40ml of hydrochloric acid (3:2), bring to the boil and then filter into a 250ml
graduated flask. Wash the crucible and filter with hot water collecting the washings in
the flask. Cool, make up to the mark and mix.

5:2 Determination
Transfer an aliquot of the solution from 5:1 containing 10 to 40mg of calcium into a
250ml beaker, add 1ml of citric acid solution (3:6) and 5ml of ammonium chloride
solution (3:4). Make the volume up to approximately 100ml with water, bring to the boil,

14:1/1
 add 8 to 10 drops of bromocresol green indicator solution (3:9) and 30ml of a warm
solution of ammonium oxalate (3:5). If any precipitate forms dissolve it by adding a few
drops of hydrochloric acid (3:2). Neutralise very slowly with ammonia (3:1), stirring
continuously until a pH 4.4-4.6 is obtained (i.e. when the indicator changes colour). Place
the beaker on a steam bath and keep it there for 30 minutes to allow the precipitate, which
has formed, to settle. Remove the beaker from the steam bath and allow it to stand for
one hour. Transfer the precipitate to the glass filter crucible (4:3) with water and wash
the beaker and the precipitate with water until the excess ammonium oxalate is removed.
(The absence of chloride in the washing water indicates that they have been sufficiently
washed). Rinse the outside of the glass filter crucible with water and discard the rinsings.
Place the crucible containing the precipitate in the original 250ml beaker, add 50ml of
sulphuric acid (3:7) and water to give a total volume of about 100ml and heat the contents
to 70-80oC in order to dissolve the precipitate. Titrate the hot solution with potassium
permanganate (3:8) until a pink colour persists for one minute.

6: Expression of the Results

6:1 Calculate the Calcium (Ca) content using the formula

Ca = [ (S x 2.004) / 1000 ] x 100

W

Where;

S = Sample titration (ml), where 1ml potassium permanganate = 2.004mg Calcium

1000 = factor for conversion of mg to g
W = Mass of sample used (g)

6:2 Take the result as the arithmetic mean of 2 determinations, if the requirement for
repeatability (7:) is satisfied. If it is not, then the determinations should be repeated.

6:3 Report the result as a percentage to one decimal place.

7: Repeatability
The difference between results of two parallel determinations on the same sample must
be less than 10% in relative value to the highest value obtained.

8: NOTE: where the magnesium content of the sample exceeds that of the calcium, the
calcium oxalate should be re-dissolved and re-precipitated before titration with potassium
permanganate.

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