Introduction/Theory:

Essential to the proper formation of teeth and bones calcium (Ca) is a mineral that can be found in
cereals fruits and vegetables. Predominantly important during childhood ones calcium intake is a
key determinant of bone mass in adults, and it also influences the rate. Dairy products are an
excellent source of bio-available calcium, and with increase intake can reduce osteoporosis, a
disease that affects millions where bones become fragile (bone thinning) that over time leads to
bone fracture.
In this experiment the percentage of calcium present in milk will be determined using back titration
with EDTA. EDTA is a hexaprotic ion (chelating agent): meaning that each of the acid oxygen’s
and each of the amine nitrogen’s can donate one electron pair, and it works by binding to metal ions
(i.e. forms a complex with calcium ions.) Ca2+ + EDTA→ Ca[EDTA]4-. The chemical equation
shows that the EDTA will react with the Ca in a 1:1 molar ratio. In this titration, a solution
containing the free metal ion (i.e. calcium) with a solution of chelating agent (EDTA) is titrated
against the standard Ca2+ ion solution. The endpoint is usually measured with an indicator ligand
that forms a colored complex with the free metal ion, which in this case a blue → violet colour
change is seen.

Figure 1: EDTA(ethylenediaminetetraacetic Figure 2: A structure of how EDTA bonds with

acid) molecule the calcium ion

Image taken from: http://www.chm.bris.ac.uk/motm/edta/EDTA.gif Image taken from: http://openlearn.open.ac.uk/file.php/2986/S_

[Accessed 11/04/10] 1_002i.jpg[Accessed 11/04/10]

Experimental: Carried out as stated in lab manual (pg 10 - 11)

Results:

Table 1: Showing the mass of powdered milk used.

Sample # 1 2 3
Mass of empty conical flask /g 114.79 89.78 133.94
Mass of powder and conical flask /g 115.78 90.77 134.93
Mass of milk powder used /g 0.99 0.99 0.99

Table 2: Showing the masses of EDTA and CaCO3 used.

EDTA CaCO3
Mass of empty beaker /g 32.0848 32.0680
Mass of beaker and salt /g 35.7874 32.6830
Mass of salt used /g 3.7026 0.6150

Calculation for Mass of EDTA needed to make std solution

Mr EDTA: 372.24 gmol-1
No. of moles = 0.04 moles
Therefore mass of salt required for 25mL = (372.24 gmol-1×0.04 moles)/4
Mass of EDTA = 3.7224g

Table 3: Titre volumes of CaCO3 standard solution vs 25mL 0.04M EDTA and Milk solutions

Sample # 1 2 3
Initial Burette Reading /cm3 1.0 0.0 0.0
Final Burette Reading /cm3 38.6 37.5 37.0
Volume of CaCO3 /cm3 37.6 37.5 37.0

Table 4: Titre volumes of CaCO3 standard solution vs 25mL 0.04M EDTA.

Sample Blank # 1 2 3
Initial Burette Reading /cm3 0.0 0.0 0.0
Final Burette Reading /cm3 40.4 40.6 40.5
Volume of CaCO3 /cm3 40.4 40.6 40.5
Treatment of Results:
Determination of the concentration of EDTA
Mass of EDTA used = 3.7026g
Mr EDTA: 372.24 gmol-1
No. of moles EDTA = 0.04 moles
No. of moles of EDTA present in 250mL
Moles = m/ Mr = 3.7026g / 372.24 gmol-1 = 9.947 x10-3 moles
Therefore there is 9.947 x10-3 moles present in 250mL of solution
Concentration = (9.947 x10-3 moles / 250mL) x 1000 = 0.0398 M

Determination of the concentration of CaCO3

Mass of CaCO3 used = 0.6150g
Mr CaCO3: 100.09 gmol-1
No. of moles of CaCO3 present in 250mL
Moles = m/ Mr = 0.6150g / 100.09 gmol-1= 6.144 x10-3 moles
Therefore there is 6.144 x10-3 moles present in 250mL of solution
Concentration = (6.144 x10-3 moles / 250mL) x 1000 = 0.0246 M

Determination of the number of moles of Ca that reacted with 25mL of EDTA

Sample blank 1 used 40.4mL of CaCO3
Therefore in 40.4mL there is (0.0246M /1000) x 40.4mL = 9.938x10-4 moles used
So 9.938x10-4 moles of Ca reacted with 25mL of EDTA

Sample 1 used 37.6mL of CaCO3

Therefore in 37.6mL there is (0.0246M /1000) x 37.6mL = 9.925x10-4 moles used
So 9.925x10-4 moles of Ca reacted with 25mL of EDTA

So the number of moles of calcium in sample 1

= Total calcium to react with 25 mL of EDTA – moles of calcium that react with residual moles of
calcium
= 9.938x10-4 moles - 9.925x10-4 moles = 1.3 x10-6 moles

Determination of percentage of Ca in milk sample 1

Mr Ca = 40.078 gmol-1
Mass = moles x Mr = 1.3 x10-6 moles x 40.078 gmol-1 = 5.21 x10-5 g in 0.9900 g of milk
So the amount of Ca in 100g of powder milk = 521mg

Average calcium concentration

= (521mg + 573mg+524)/3 = 539mg
Standard deviation = 29.19
Therefore calcium concentration in milk samples = 539 ±29.19 mg
Conclusion/Discussion : This lab has shown how the amount of calcium present in milk can be
found. The theoretical mass of calcium in powder milk is 900-1190 mg per 100g. For this
experiment the mass attained 539 ±29.19 mg. The size of the std. dev shows a large degree of error
and since the type of milk was not stated the accuracy cannot be determined. Colour changes for
calcium-EDTA titration was seen in the blanks titres where Solochrome Black was used as the
indicator. In the beginning the solution of the flask was royal blue colour well before endpoint
(excess Ca2+ ions present to complex with indicator). Centre this then turned a purple colour at
endpoint (all Ca2+ ions complexed by EDTA, indicator completely uncomplexed). In the case of
the samples colour changes were also noticed as they each started off as cloudy (opaque) bluish
sample solutions well before the end point, followed by a trace of bluish-purple colour just before
endpoint an finally a purple colour at endpoint.

Answers to Questions:
1. Explain the mechanism by which EDTA complexes with Ca++
Ethylene Diamine Tetra Acetic acid (EDTA) is one of the most common chelating agents used in
chemistry.

EDTA

EDTA has this chelating effect because it forms octahedral complexes with calcium as shown
below. This complex is more favored than the monodentate ligands of water and other bidentate
ligands. So these ligands are replaced by EDTA*4. They also react in a 1:1 ratio Ca2+ + EDTA4-→
[CaEDTA]2-.

Diagram 1 showing CaEDTA]2-

 2. An Amperometic titration method can be used to determine the calcium concentration in the
presence of barium. This method of titration measures the concentration of analytes in
solution by means of an electric current. It takes advantage of the fact that the calcium is
smaller than the barium ion so it will move faster if a voltage is applied. Voltage is applied
across the indicator electrode (Pt) and reference electrode (Hg/calomel) is held constant, the
current passing through the cell is measured as a function of the titrated volume added. So
an electrical cell has to be made and titre volumes are added by a microburet. Then a graph
current vs volume of reagent added is plotted. To straight lines with different gradients one
will be a negative gradient and the other positive will be obtained after which it will be
extrapolated to where they intercept. This point is the volume of reagent added.

References:
1. CHEM 2460 Principles of Chemical Analysis Laboratory Manual.pg 17-21
2. James N Miller & Jane C Miller, “Statistics and Chemometrics for Analytical” Chemistry,
5th Ed(2005) Publ. Pearson Education Limited pg 114
3. http://www.csudh.edu/oliver/che230/labmanual/calcium.htm [Accessed 10/04/10]
4. WO SAY DISCUSSION HELP http://www.ajcn.org/cgi/reprint/83/2/310.pdf [Accessed
10/04/10]
5. http://www.cerlabs.com/experiments/10875404367.pdf[Accessed 10/04/10]