week
Experiment
Page
no.
1
Extraction of Nickel as dimethyl glycoxime
1
2
a) Determination of Iron by chloride extraction
3
b) Separation of Iodine
5
3
Separation of mixture of K2Cr2O7 and KMnO4 by column chromatography
8
4
Paper chromatography
5
6
a) Separation of amino acids
10
b)Separation of metal ions from three groups
13
Paper chromatography
a) Separation of halides
15
b) Separation of sugars
17
Thin layer chromatography
a) preparation of Thin layer chromatography
20
b) Separation of chlorophyll
21
7
Separation of nitro phenol isomers by thin layer chromatography
23
8
a) Determination of an-ion-exchange resin
25
b) preparation of column
25
9
Determination of the total cation concentration in water
28
10
Gas – Liquid chromatography
30
Separation of a hydrocarbon mixture
11
Visit
12
Final Exam
Experiment (1)
Extraction of Nickel as the dimethy glyoxime complex
Theory:
Nickel (200-400 µg) forms the red dimethyglyoxime complex in a
slightly alkaline medium; it is only slightly soluble in chloroform
(35-50 µg Ni L-1). The optimum pH range for extraction of the
nickel complex is 7-12 in the presence of citrate. The nickel
complex absorbs at 366 nm and also at 465-470 nm.
Chemicals:
Amm. Nickel sulphate (0.135 g), citric acid (A.R.) 5 g, ammonia,
dimethyglyoxime, chloroform and aluminium salt.
Procedure:
A homogenous solution of nickel (200-400 µg) and aluminium
(500 µg) or iron (500 µg) is prepared. Transfer 10.0 ml of this
solution (Ni content about 200 µg) to a beaker containing 90 ml of
water, add 5.0 g of A.R. citric acid, and then dilute ammonia
solution until the pH is 7.5. Cool and transfer to a separatory funnel,
add 20 ml of dimethylglyoxime solution (1) and, after standing for a
minute or two, 12 ml of chloroform. Shake for 1 minute, allow the
phases to settle out, separate the red chloroform layer, and determine
the absorbance at 366 nm in a l.0 cm absorbance cell against a blank.
Extract with a further 12 ml of chloroform and measure the
absorbance of the extract at 366 nm; very little nickel will be found.
Test for the iron or aluminium in the aqueous layer.
-1-
Repeat the experiment in the presence of 500 µg of iron (III) and
500 µg of aluminium ion; on interference will be detected.
Note:
The dimethyglyoxime reagent is prepared by dissolving 0.50 g of
A.R. dimethyglyoxime in 250 ml of ammonia solution and diluting
to 300 ml with water.
-2-
Experiment (2)
a) Determination of Iron by chloride extraction
Theory:
The extraction of iron(III) chloride from hydrochloric acid with
diethyl ether (probably as the solvated complex H[FeCI 4] has long
been known, but the amount of metal extracted depends upon the
concentration of the acid and passes through a maximum at about
6M-hydrochloric acid.
Elements that extract well as chloride complexes include Sb(V),
As(III), Ga(III), Ge(IV), Tl(III), Hg(II), Mo(VI), Pt(II), and Au(III).
Elements which are partially extracted include Sb(III), As(V), V(V),
Co(II), Sn(II), and Sn(IV). Many solvents with donor oxygen atoms,
including di-isopropyl ether, ββ′-dichlorodi ethyl ether, ethyl acetate,
butyl acetate, and pentyl acetate, have been employed. In most cases
the optimum extraction depends upon the acid concentration.
The extraction of large amounts of iron is conveniently made with
iso-butyl acetate: this solvent has the merit of low volatility and of
almost negligible temperature rise during the extraction (unlike
diethyl ether).
.
To gain experience in the procedure, experimental details are
given for the extraction of iron (III) in hydrochloric acid solution
with diethyl ether.
-3-
Procedure:
Weigh out 16.486 g of A.R. hydrated ammonium iron (III)
sulphate and dissolve it in 250 cm3 of 6M-hydrochloric acid in a
graduated flask. Extract 5ml of the iron (III) solution (which
contains 200mg of Fe) with three 5ml portions of pure diethyl ether
(1): shake gently for 3 minutes during each extraction. Combine the
three ether extracts and strip the iron from the ether by shaking with
25ml of water: approximately 99.9per cent of the iron is removed by
this method. Boil off any ether remaining in the aqueous extract on a
water- bath (caution!), and determine the iron by titration with
standard 0.1N-potassium dichromate after previous reduction to the
iron (II) state. The iron recovered should not be less than 99.6 per
cent (2).
Notes: The factors of importance in the diethyl ether extraction of
iron are:
a) The iron must be in the iron (III) state, since iron (III) chloride is
not extracted.
b) The hydrochloride acid concentration must be close to 6M.
c) The extraction should be carried out in subdued light, since ether
photochemically reduces iron (III).
d) The ether should be free from ethanol and peroxides because these
reduce iron (III) chloride.
e) The concentration of anions other than chloride should be kept low.
f)Heat is generated by the mixing of the ether and the hydrochloride
acid iron (III) chloride solution so that cooling of the mixture under
the tap or in ice is essential.
-4-
b) Separation of Iodine
You will be given a homogenous mixture of iodine and sodium
chloride in distilled water.
.
Theory:
The iodine is separated by an extraction process. The aqueous
iodine-salt solution, is shaken together with an approximately equal
volume of carbon tetrachloride (CCL4). Water and carbon
tetrachloride do not mix (they are insoluble in one another) hence
two liquid phases will coexist here. Iodine vastly prefers to dissolve
in CCL4, thus it migrates from the aqueous (H20) phase into the nonaqueous (CCI4) phase - we say that CC14 “extracts” the I2 from the
aqueous phase. Cl has no such tendency (its solubility in CCL 4 is
nil)
and
hence
it
remains
behind
in
the
water
phase.
Procedure:
Pour all of filtrate into a clean 125 ml separatory funnel
(suspended in an iron ring on a ring stand) whose stopcock is closed.
Pour in 20 ml of carbon tetrachloride *, watch the added liquid to see
whether it dissolves, floats as an upper layer, or settles as a lower
layer. (Which liquid has the greated density CCI4 or H20?). Insert the
separatory funnel stopper, and shake the closed funnel for about 10
sec. (Fig. 1). Shake vigorously enough so as to mix the aqueous and
non-aqueous phases intimately. Keep your hand on the stopper, if
internal pressure builds up, the stopper may pop out. To vent the
inside pressure, hold the stoppered separatory funnel upside down,
allow the liquids to drain away from the stopcock, and which the tip
still pointing up, open the stopcock momentarily. Close the
-5-
stopcock, turn the separatory funnel upright, remove the top stopper,
and allow the layers to separate as completely as possible.
Carefully drain the lower layer through the stopcocker closing it
just before the last drop of lower layer goes through. What will the
lower layer be? Add another 20 ml portion of fresh CC1 4 to the
remaining upper layer and repeat once more, drawing the second
batch of CCL4 into a separate container. (Why must remove the top
stopper from the separatory funnel each time before attempting to
drain out the lower layer?). Repeat a third time.
Separately save the second and third portions of CCl 4, note and
compare their color with the first portion. Show these carbon
tetrachloride solutions, to your laboratory instructor. Save the
aqueous layer.
-6-
Iodine test*:
In the presence of iodine, the white color of the starch paper
changes to deep blue. Dip your stirring rod into the solution and then
touch it to a piece of the starch paper. Be sure your stirring rod is
thoroughly rinsed before and after each use.
Chloride test:
To a test tube containing the sample (the aqueous layer), add 2
drops of nitric acid (HN03, 6M HN03) and then 1 drop of silver
nitrate solution (0.05 M AgNO3). You will observe a precipitate.
Exercise:
Suggest a volumetric method for the determination of iodine and
chloride after the separation. Discuss the theory of the indicator used
in each method.
*
Iodine and its vapours are very corrosive and toxic. Do not allow it to touch
your skin or clothing, avoid inhalation.
-7-
Experiment (3)
Separation of mixture of potassium permanganate and
potassium dichromate by column chromatography
Theory:
The separation of mixture containing both the potassium
dichromate and potassium permanganate pass from the separation
column containing material alumina acid, solvent using distilled
water as the loyal and adsorption dichromate on the surface of
alumina acid more adsorption permanganate therefore concludes
permanganate first. If checked consider dichromate layer orange in
the separation column you will notice a yellow minutes of chromate
potassium accompanying the potassium dichromate and who is with
him in equilibrium as shown in the following equation:
Cr2O 72  + H2O ↔ 2HCrO 4
Chemicals:
1- 16-18 gm of alumina acid.
2- 1% of potassium dichromate solution.
3- 1% of potassium permanganate solution.
4- Commingle sizes equal from both potassium dichromate and
potassium permanganate.
5- 1mol/L sulfuric acid.
6- 1mol/L nitric acid.
-8-
Procedure:
1- Prepare the separation column from alumina acid the length of
column 12-15 cm.
2- Open the tap separation column and waited until the water level
slightly above the surface of alumina.
3- Convey very carefully 5ml of the mixture to be separated by
pipette in one go to the highest separation column.
4- Put 100ml of distilled water in the separating funnel installed over
the brink separation column and leave the water is equal to a fixed
rate of 5ml per minute.
5- Process repeated washing with water several times until the violet
permanganate layer separating from layer dichromate yellow.
6- Extract the isolated permanganate on the column by washing the
column by 50ml nitric acid solution, which concentration 1 mol /
liter by the rate of about 2 ml per minute and collect the effluent in
conical flask.
7- Extract the isolated potassium dichromate on the column by
washing the column by 50ml sulfuric acid solution, which
concentration 1 mol / liter and collect the effluent in conical flask.
8- For quantitative analysis both permanganate and dichromate used
spectral methods or by volumetric analysis.
-9-
Experiment (4)
Separation of amino acid by paper chromatography
Theory:
Paper chromatography is a techniques used to separate complex
mixtures of drugs, metal ions, amino acids and dyes. One of the
classic applications of paper chromatography is the separation of
amino acids. The sample is spotted on the paper, eluted and
subsequently visualised by reaction with ninhydrin to form a purple
or brown colour. Qualitative identification is made by calculation of
Rf values for each spot.
Rf = distance solute moved / distance solvent moved.
Quantitative determination can be made by comparison of the
intensity of the colour with standards.
Chemicals:
1- Reagents:L-leucine, DL-aspartic acid, L-lysine and mixture of
the three amino acids. Use 1% aqueous solution.
2- Solvent: n-butanol: acetic acid gloicol: water
60 :
15
:
25 by volume.
3- Sheets: Whatman chromatographic paper.
4- Equipments: Chromatography tank, capillaries, hair dryer,
spray bottle.
5- Locating reagent: Ninhydrin (0:2 gm in 100 ml acetone).
- 10 -
Procedure:
1- Place sufficient solvent into the bottom of the tank cover the lid
and
allow
the
tank
to
be
saturated
with
the
solvent.
2- Take a sheet of Whatman chromatography paper (about 10-30
cm) and place it on a piece of clean paper on a bench.
3- Draw a fine line with a pencil along the width of the paper and
about 1.5 cm from the lower edge.
4- Along this line place four equally spaced (about 2 cm apart) small
circles with a lead pencil.
5- Label the paper at the top with the name of each of the three
amino
acids
and
label
the
fourth
unknown.
6- Use a fine capillary or tooth pick to place drops of the solutions to
be chromatographed and dry each drop before applying the next
drop.
7- After spotting, allow the paper to air dry for 5 minutes then use a
hot-air hair dryer for one minute.
8- Place the spotted paper in the chromatographic tank and make the
development using the ascending technique.
9- Close the tank with lid, allow solvent to flow for about 1.5 hou.
10- Remove the paper and immediately mark the position of the
solvent front with a lead pencil.
11- After the chromatogram has dried: spray the paper with 0.2
ninhydrin solution.
12- Put the sprayed paper in an oven at 105° C for two minutes.
The amino acids will form purple or blue spots. The colour is
stable for some weeks if kept in the dark away from acid vapours.
- 11 -
13- For permanent record, circle the position of each spot with a
pencil.
14- Compute the Rf values and record them in a table for both for
single amino acids and for the components of the mixture (compare).
- 12 -
b) Chromatography, ascending or descendingseparation of metal ions from three groups (Co2+, Ni2+,
Cu2+, Fe3+)
Theory:
The separation and identification of metal cations from different
groups by paper chromatography.
Chemicals:
1- Reagents: A solution containing 20 mg/mI of chlorides of
each of: (Co2+, Ni2+, Cu2+, Fe3+).
2- Solvent: Acetone: conc. HCL: water
86 :
6
:
8 by volume.
3- Sheets: Whatman chromatographic paper.
4- Equipments: Chromatography tank, capillaries, hair dryer,
spray bottle.
5- Locating reagent: Ammonia.
Procedure:
1) Place 50 ml of solvent in the bottom of the tank. cover the lid.
2) Prepare the paper, and place three spots of the mixture, evenly
spaced, along the base line.
.
3) Form the paper into a cylinder and secure with the tongued clips,
Part G. Place the cylinder, with the spotted end down, in the tank
taking care not to let the paper touch the glass walls.
4) Close the tank with the lid.
- 13 -
5) If the chromatogram is observed closely, a yellow band due to Cu
may be seen moving behind the solvent front. The locating reagents
are prepared as above.
6) After the solvent front has travelled about 1/2 of the length of the
paper, remove the chromatogram from the tank, mark the solvent
front with a pencil, open out and dry.
7) After the chromatogram has dried: spray the paper with ammonia
solution.
This shows Nickel is blue spot, Cobalt is yellow brown spot, Copper
is olive green spot, and Iron is reddish brown.
8) Compute the Rf values.
- 14 -
Experiment (5)
a) Chromatography, ascending the separation of anions
the halides CI, Br, I
Theory:
Separation of anions (the halides), and their locating on paper
chromatograms.
Chemicals:
1- Reagents: (a) Potassium chloride/water - 10 mg/ml.
(b) Potassium bromide/water - 10 mg/ml.
(c) Potassium iodide/water - 10 mg/ml.
(d) A solution of the above salts in the same concentrations.
(e) A solution of any two to of above salts as an unknown.
2- Solvent: iso-propanol: acetone: 0.880 ammonia solution.
40
:
30 :
30 by volume.
3- Sheets: Whatman chromatographic paper.
4- Equipments: Chromatography tank, capillaries, hair dryer,
spray bottle.
5- Locating reagent: 0.1 m/L Silver nitrate.
Procedure:
1) Place 50 ml solvent in bottom of the tank. Replace the lid.
2) Use a fine capillary or tooth pick to place drops of the solutions to
be chromatographed and dry each drop before applying the next
drop.
3) After spotting, allow the paper to air dry for 5 minutes then use a
hot-air hair dryer for one minute.
- 15 -
4) Place the spotted paper in the chromatographic tank and make the
development using the ascending technique.
5) Close the tank with lid.
6) Remove the paper and immediately mark the position of the
solvent front with a lead pencil.
7) After the chromatogram has dried: spray the paper with Silver
nitrate solution.
This shows Chloride is gray spots, Bromide is yellowish gray spots,
and Iodide is yellow spots.
8) For permanent record, circle the position of each spot with a
pencil.
9) Compute and compare the Rf values of each anion, when run
individually and when run in a mixture with the others.
- 16 -
b) Chromatography, ascending or descending the
separation of sugars
Theory:
a) The separation and identification of individual sugars and
mixtures of naturally occurring sugars by paper chromatography.
b) Determination of Rg values.
Chemicals:
1- Reagents: D (+) – glucose.
D (+) – xylose.
Lactose.
Fruit juices.
2- Solvent: ethyl acetate: pyridine: water.
55 :
25 :
20 by volume.
3- Locating reagent: m- phenylene diamine - 0.5 g.
Stannous chloride - 1.2g.
acetic acid - 20 ml.
Ethanol - 80 ml.
Procedure:
1) Place 50 ml of solvent in the tank, replace the lid.
2) Prepare 25 x 25 cm paper with eight origins and number one
through eight.
3) Onto origins 1, 2 and 8, spot one drop of lactose solution.
4) Allow the spot to dry spontaneously, or by blowing over them a
current of air from a hair dryer.
5) Spot a dry of glucose solution on to origins 1, 3 and 8.
6) Dry again.
- 17 -
7) Spot a drop of xylose solution on to origins 1, 4 and 8.
8) On to origins 5, 6 and 7, spot a drop of different fruit juices.
9) From the paper into a cylinder and secure with the tongued clips,
part G. Place the cylinder, with the origins end down, into the tank
taking care not to let the paper touch the glass walls. Close the tank
with the lid.
10) No observation can be made while the chromatogram is running
because the sugars are colorless.
11) Allow the chromatogram to run until the solvent front reaches
nearly to the top of the paper or, preferably, overnight.
12) Remove the chromatogram from, the tank, mark the solvent
front
with
a
pencil,
open
out
and
dry
(see
Note).
13) Pour the locating reagent into the drip tray, and dip the
chromatogram.
14) Heat the dipped paper for up to 5 minutes in an oven at 100-105
C. The sugars from dark yellow to brown spots.
15) Compare and compute the Rg values of each sugar when run
individually and when run in a mixture with the others.
Conclusions:
(i) Colourless sugars can be separated and identified by paper
chromatography through the use of a locating reagent which
transforms
them
into
coloured
spots
on
the
paper.
(ii) The Rg values of sugars are the same whether run individually or
in
mixtures
with
each
other.
.
(iii) Chromatography is a useful technique for the analysis of sugar
mixtures.
.
- 18 -
Note:
If the chromatogram is allowed to run overnight, the solvent front
reaches the to evaporate top of the paper and continues to evaporate
off the edge. In this case, no true front is available for the
computation of Rf values. In such chromatograms, it is the practice
to use an Rg value, based on the distance traveled by the glucose as
the preference point, and computed by the following formula:
Rg=
distance the substance has run from the origin x 100
distance the glucose has run from the origin
Thus, the Rg for glucose itself is 100.
- 19 -
Experiment (6)
Thin layer chromatography (TLC)
When a separation of components in a mixture is attempted, TLC
has several advantages over paper chromatography:
1. The time taken to achieve separations is far less than that
for paper chromatography.
2. The resolution of components is usually superior.
3. It is possible to apply to coated plates a variety of
corrosive location reagents that would destroy paper
chromatogram.
4. The non-fluorescing inorganic adsorbents used in TLC
provide a greater contrast to fluorescent spot than does
chromatographic paper.
5. The wide range of adsorbents available enhances the
flexibility of the method.
a) Preparation of the plates:
1- Plates themselves should be cleaned with detergent solution, then
rinsed
thoroughly
with
water
and
allow
to
dry.
2- Silica gel G is slurried with disi1led water. Generally, two parts
of water are mixed thoroughly with one part of adsorbent.
3- The slurry is then applied to the plates in a thickness of 0.25 mm.
- 20 -
4- The plates are left in the air for 2 hours and then dried in an oven
at 105 - 110°C for one hour and stored in a disicator.
b) Separation of chlorophyll and related compounds by
TLC
Theory:
The depend method of separating the components of green plants
on the adsorption difference of green material on the surface of silica
gel existing in the form of thin layer. This method is similar to the
method largely separation chromatography by column.
Chemicals:
Fresh leaves, sand, chloroform, mortar and TLC silica gel plate.
Solvent: isopropyl alcohol: petroleum ether: acetone.
10
:
50
:
50 by volume.
Procedure:
1) Place 50 ml of solvent in the tank, replace the lid.
2) Prepare the thin layer plate (Silica gel are mixed with distilled
water).
3) Obtain a small quantity of green plant material and grind it in a
morter with some sand.
4) Extract the mashed plant material with 5ml of acetone or
chloroform then filter the liquid, which should be coloured deep
green.
5) The solutions of chlorophyll are spotted on the same plate about 2
cm apart using a capillary tube. Spotting is carried out following the
same technique used in paper chromatography.
- 21 -
6) The plates are placed in tightly covered tank containing the
mobile phase. When the solvent moves about 12 cm the plates are
removed and air-dried.
7) Describe the colour separations which you observed (The green
substance is chlorophyll. The orange band is due to a closely related
group of compounds called carotenes. The yellow band is produced
by xanthophylls).
8) Calculate the Rf values.
- 22 -
Experiment (7)
Separation of nitro phenol isomers by thin layer
chromatography
Theory:
Chromatography is thin layer of an important way to separate
images isomers of organic compounds from each other. The
experience separation of a mixture composed of ortho and meta and
para nitrophenol (o-nitrophenol, m-nitrophenol and p- nitrophenol)
good example of the use of thin layer Chromatography.
Chemicals:
1- Prepare the 0.1% from an aqueous solution of each of the three
images nitrophenol isomers.
2- A mixture of unknown contains two from phenols the previously
mentioned.
3- Solvent: methanol: benzene.
5
:
95 by volume.
4- Locating reagent: 1% alc. KOH.
Procedure:
1) Prepare the thin layer plate is used the silica gel as constant
medium.
2) Place of solvent (stationary phase) in the tank.
3) Cover the lid and allow the tank to be saturated with the solvent.
4) Draw a fine line with a pencil along the width of the thin layer
plate and about 2 cm from the lower edge.
- 23 -
5) Use a fine capillary to place drops of the solutions to be
chromatographed and dry each drop before applying the next drop.
6) Place the spotted plate in the chromatographic tank and make the
development using the ascending technique.
7) Close the tank with lid, allow solvent to flow. Remove the paper
and immediately mark the position of the solvent front with a lead
pencil.
8) After the chromatogram has dried: spray the plate with 1%
alcohol KOH.
9) Describe the colour separations which you observed (the
yellowish orange spot is o-nitrophenol. The yellow spots are
produced by m-nitrophenol and p-nitrophenol).
10) Calculate the Rf values.
- 24 -
.
Experiment (8)
Determination of the capacity of an ion-exchange resin
Theory:
The total ion-exchange capacity of a resin is dependent upon the
total number of ion-active groups per unit weight of material, and
the greater the number of ions the greater will be the capacity. The
total ion-exchange capacity is usually expressed in milli equivalent /
gm of exchanger. The exchange capacity of a cation-exchange resin
may be measured in the laboratory by determining the number of
milligram equivalents of sodium ion which are absorbed by 1 gm of
the dry resin in the hydrogen form. Similarly, the exchange capacity
of a strongly basic anion exchanger is evaluated by measuring the
amount of chloride ion taken by 1 gm of the dry resin in the
hydroxide form.
Determination of the capacity of anion-exchange resin:
Chemicals:
1- Air dried strongly basic anion exchanger (Dowex 20-50 chloride
form).
2- Sodium nitrate solution Ca. 0.25 M.
Procedure: 1- Dry off purified resin by placing it in an evaporating
dish (over with a watch glass supported on two glass rods to provide
protection from dust while giving access to the air.
2- Leave in warm place (25-35°C) until the resin is completely freerunning (2-3 days).
- 25 -
3- Partly fill a small burette, provided with a glass wool plug at the
lower end, with distilled water taking care to displace any trapped
air from beneath the glass wool plug.
4- Weigh out accurately about 0.5 gm of the air-dried resin in a
weighing bottle.
5- Transfer the resin with the aid of a small camel hair brush through
a dry funnel into the column.
6- Add sufficient distilled water to cover the resin.
7- Adjust the level of the out let tube so that the liquid in the column
will drain to a level about 1 cm above the resin beads.
8- Fill a 250 ml separating funnel with Ca 0.25 M NaNO 3 and allow
this solution to drop into the column at the rate of about 2 ml /
minute. Collect the effluent in a 500 ml conical flask, and titrate
with standard 0.1 N AgNO3 using potassium chromate as indicator
(Mohr method).
The reaction which occurs may be written as:

R+C1- + NaNO3
R+NO 3 + NaC1
Calculation:
The capacity of the resin expressed as meq / gm is given by:
N x V / W (where: N is the normality of AgNO3, V is the volume
of AgNO3 and W is the weight of the resin.
- 26 -
Standardisation of AgNO3 solution:
1- Pipette 25 ml of standard 0.1 N NaC1 into a 250 ml conical flask.
2- Add 1 ml of indicator solution K2CrO4.
3- Add AgNO3 solution from a burette, swirling the liquid
constantly, until the red colour formed by the addition of each drop
begins to disappear more slowly (match the colour against a white
filter paper), this is an indication that most of the chloride has been
precipitated.
4- Continue the addition drop by drop until a faint but distinct
change in colour occurs. The faint reddish- brown colour should
persist after brisk shaking.
5- Calculate the normality of AgNO3 solution from the relation:
N x V = N′ x V′
- 27 -
Experiment (9)
Determination of the total cation concentration in water
Theory:
The following procedure is a rapid one for the determination of the
total cations present in water particularly that used for industrial ion
exchange plant but may be used for all samples of water, including
tap water. When water containing dissolved ionised solids is passed
through a cation exchanger in the hydrogen form, all cations are
removed and replaced by hydrogen ions. By this means, any
alkalinity present in the water is destroyed and the neutral salts
present in solution are converted into the corresponding mineral
acids. The effluent is titrated with 0.02M sodium hydroxide using
screened methyl orange as indicator.
Chemicals:
Amberlite resin, 0.1M NaOH, methyl orange indicator and tap
water.
Procedure:
1) Prepare column of Amberlite resin (H-form). The level of the
water should never be permitted to drop below the upper surface of
the resin in the column.
2) Pass 150 ml of the sample of water under test through the column
at a rate of 3-4 ml per minute, and discard the effluent.
3) Now pass two 120 ml portions through the column at the same
rate.
- 28 -
4) Collect the effluents separately, and titrate each with standard
0.1M NaOH using screened methyl orange as indicator.
From the results of the titration, calculate the milli-equivalents of
calcium present in the water. It may be expressed, if desired, as the
equivalent (E.M.A.) in terms of mg CaCO3 per L of water (i.e., parts
per million of CaCO3). In general, if the titre is A ml of NaOH of
molarity B for an aliquot volume of V ml, the E.M.A. is given by
(AB × 50 × 1000) / V.
Commercial samples of water are frequently alkaline due to the
presence of hydrogen carbonates, carbonates, or hydroxides. The
alkalinity is determined by titrating a 100 ml sample with 0.02 M
hydrochloric acid using screened methyl orange as indicator (or to a
pH of 3.8). To obtain the total cation content in terms of CaCO3, the
total methyl orange alkalinity is added to the E.M.A.
Results:
(NV)NaOH = (NV)H+ = (NV)CaCO3 = (wt/eq.wt x 1000) CaCO3
Total hardaness ppm CaCO3 = wt x 106 / 150
- 29 -
Experiment (10)
Gas - Liquid Chromatography
Separation of a hydrocarbon mixture
Gas chromatography is probably the most valuable separation
technique developed to date. A small amount of the liquid sample is
injected into a hot injection chamber where it is changed into a gas.
The gas is partitioned between the inert, carrier gas (usually helium)
and a nonvolatile, high molecular weight, organic stationary phase
that is coated on a solid support. Because of their differences in
solubility in the stationary phase, the sample components are
separated.
Apparatus:
Gas chromatography with a nonpolar column, in this experiment,
the effects of column temperature on the separation of series of
hydrocarbons will be investigated. Also the determination of each
hydrocarbon in the sample will be carried out.
Procedure:
1- Prepare a mixture of propanol, heptane and octane in different
proportion. Using an appropriate column set the following
parameters on the gas chromatograph:
a- Column temperature 60°C.
b- Injection chamber temperature at least 135°C.
c- Detector temperature at least 150°C.
d- Gas flow rate, 30 cm3/min.
e- Filament current to 150 m A (in case of TCI) detection.
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f- Record the length of the column.
2- After the instrument has come to equilibrium inject 2 µl of the
mixed hydrocarbon sample and allow the three components to elute.
3- Carry out the experiment with column temperature at 110, 100,
75, 65 and 60°C while keeping the flow rate constant at 30 cm3/min.
4- Determine which temperature is optimum, and adjust the column
to that temperature.
5- Successively inject and record the chromatogram of 3 µl portions
of the sample until the height of the first peak is identical for three
injections.
6- Successively inject and record the chromatograms of 0.5, 1.0, 1.5,
2.0 and 2.5 µl portions of pure propanol, heptane and octane.
Calculations:
1- Use the chromatograms of the sample to calculate the adjusted
retention time (tR) for each of the three sample component peaks.
Qualitatively determine the sample component that is responsible for
each of the three sample peaks by comparing the retention times of
the sample peaks with those of the pure hydrocarbons.
2- Calculate the resolution of the column. Rs =
t R 2  t R1
1 / 2( w1  w2 )
The resolution is complete if Rs > 1 .5.
3- Calculate the number of theoretical plates N = 16 (t R /w)
2
and the height equivalent of a theoretical plate H.E.T.P = L / N
for each of the three sample components in one of the sample
chromatogram.
4- Prepare a working curve for each of the pure hydrocarbon by
plotting peak height against the injected volume of the hydrocarbon.
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5- Use e three working curves to determine the volume of each
hydrocarbon in the sample.
6- Use the volumes of each component in the sample to determine
percentage of each (v/v %) in the sample.
Percentage =
Volume of component
Volume of sample injected
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X 100
‫‪References:‬‬
‫‪1- J. Bassett et.al., Vogel’s textbook of quantitative inorganic‬‬
‫‪analysis, fourth edition, , Longman, (1978).‬‬
‫‪ -2‬الكيمياء التحليلية تجارب عملية في طرق التحليل اآللي‪ ،‬تأليف‪ :‬أ‪.‬د‪ .‬عبد الغني حمزة‬
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