CHEM2403 Results sheet for Experiment 1

Name: SID:
Lab day:
Experiment partners:
Contents
Title:............................................................................................................................................................2
Author:........................................................................................................................................................3
Aim:.............................................................................................................................................................3
Calibration:..................................................................................................................................................3
Molar Absorption:.......................................................................................................................................5
Iron %:.........................................................................................................................................................5
Occupation%:..............................................................................................................................................6
Table or figure:............................................................................................................................................8
STUDY 01.................................................................................................................................................9
STUDY NO 02...........................................................................................................................................9
Release%:..................................................................................................................................................10
Title:
1. Iron content in Ferritin
2. Iron storage in Protein

Author:

Aim:
1. To represent the utilization of spectrophotometry in follow examination of iron.
2. To see how iron is put away in living cells.

Why it is important?
A spectrophotometer estimates the measure of light sent through a substance and is an important
instrument in science. Without it, deciding the compound make-up of materials would be truly
challenging and not extremely exact

Calibration:
Measured absorbance:
Fe(II) concentration
Solution Absorbance (at 561 nm)
(M)
Blank 0.000 0.000
-4
Standard 1 1.00 x 10 0.115

Standard 2 1.00 x 10-4 0.230

Standard 3 1.00 x 10-4 0.601

Standard 4 1.00 x 10-4 1.201

 Figure 2

A cross-segment of the protein is displayed to one side. The breaks in the twofold lines address the protein channels. The
mathematical example inside the protein addresses the microcrystalline mineral design of [FeO (OH)]8[FeO(H2PO4)] that
structures when Fe (II) enters the protein circle, is oxidized by oxygen to Fe(III), and responds with water and phosphate.

Figure 3

Ferrozine2-, an organic ligand that binds Fe (II) to form a purple complex

Equation 1
Molar Absorption:
The path length b as shown below in graph is equal to 1cm therefore according to it the molar
absorption € is equal to 2.79 * 10-4 M-1cm-1 for the ferrozine iron complex at maximum wavelength.

€ = 2.79 * 10-4 M-1cm-1

Iron %:
1. Total iron content of ferritin

Concentration of Fe(II) in ferritin solution A: 384.9 M

Amount of Fe(II) in 25.0 mL solution A 9622.5 moles

537416.025g

As mentioned earlier in the Introduction, the mineral core of ferritin is made up of

[FeO(OH)]8[FeO(H2PO4)], which has a formula weight of 879.7 g/mole. Since there are 9 atoms
of iron per molecule of core, the core weight is divided by 9, giving 97.70 grams of core for
every mole of iron present. So the weight of the iron core in milligrams can be calculated using
 Amount of ferritin in 1.00 mL of stock 21111.765g

(2) Total iron content of ferritin 21111.0 % (w/w)

Occupation%:
From the total iron content of the ferritin sample obtained above (x), and given:
Atomic weight Fe = 55.85 g / mol
Formula weight Fe core in ferritin = 97.74 g / mol Fe
Number of Fe-binding sites in ferritin = 4500
Molecular weight apoferritin = 4.74 x 105 g / mol
Use the following steps to calculate the % of the possible Fe-binding sites occupied in the ferritin
molecules in your sample.
100 g of ferritin contains:
(1) How many grams of Fe? 5.37*105 g

(2) How many moles of Fe? 9622.5 moles

(3) How many g of Fe core? 940118.25 g

(4) How many grams of apoferritin? 919006.485 g

(total mass – Fe core)

(5) How many moles of apoferritin? 1.94 moles

From your answers to (2) and (5) above, calculate the number of moles of Fe per mole of
apoferritin:
 9960

This value is the number of Fe atoms per molecule of ferritin.

(3) Calculate the percentage of the 4500 possible Fe-binding sites occupied:
Solution Absorbance (at 561 nm)
110%
Blank 0.000

Treated ferritin solution A 0.1074

Table or figure:
Total iron determination

Labelled concentration of Fe(II) standard: 1.00 x 10-4 M

Concentration of stock dihydroxyfumarate 5.0 x 10-3 M

Study 1 (1.00 mL DHF reagent)

Time (min) after ferritin Absorbance Concentration of released Fe(II) /

addition (at 561 nm) M
(to be completed during analysis)
 1 0.0758 271.68
2 0.0933 334.40
3 0.1098 393.54
4 0.1248 447.3
5 0.1384 483.1
6 0.151 541.2
7 0.1631 584.5
8 0.174 623.6
9 0.1851 663.4
10 0.1954 700.3
11 0.2065 740.1
12 0.2153 771.6
Study 2 (2.00 mL DHF reagent)

Time (min) after ferritin Absorbance Concentation of released Fe(II) / M

addition (at 561 nm) (to be completed during analysis)
1 0.0696 249.4
2 0.112 401.4
3 0.1222 437.9
4 0.1354 485.3
5 0.1501 537.9
6 0.1593 570.9
7 0.1716 615.0
8 0.1827 654.8
9 0.1931 692.1
10 0.203 727.5
11 0.2126 762.0
12 0.2234 800.7
 STUDY 01
Study no 01
900
800
700
600
concentration

500
400
300
200
100
0
1 2 3 4 5 6 7 8 9 10 11 12
Time

STUDY NO 02

Study No 02
900
800
700
600
concentration

500
400
300
200
100
0
1 2 3 4 5 6 7 8 9 10 11 12
Time

Release%:
By the help pf this equation:

Initial rates of iron release

2*10-4
Study 1 (1.00 mL DHF reagent)
 4*10-4
Study 2 (2.00 mL DHF reagent)

Proportion of total iron released in time studies

Amount of Fe(II) in 25.0 mL of treated ferritin solution B

Amount of Fe(II) in 25.0 mL of treated ferritin solution B

Study 1 13650 mol Fe

7.62*105g Fe

Study 2 14425 mol Fe

8.0*105g Fe

Finally, calculate the % of the total iron released as Fe(II) during the time course of each study.

DHF
reagent(ml)
(4) % of total iron released as Fe (II)

Study 1 14.18%

Study 2 14.99% approx. 15%