3B Scientific Polarimeter with 4 LEDs (115 V, 50__60 Hz) - Manual (Page 7)

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• Measure the sample thickness and place the
sample cell in the measurement chamber.
Note:
100 ml of liquid in the sample cell corresponds to a
sample thickness of 1.9 dm, 75 ml to 1.43 dm, 50 ml
to 0.96 dm, and 25 ml to 0.44 dm.
• Measure the angle of rotation for each of the
different LEDs.
• In the next step, keep the concentration the
same but reduce the sample thickness to 1.43
dm (75 ml) and repeat the measurement.
• Make further measurements with sample
thicknesses of 0.96 dm (50 ml) and 0.44 dm (25
ml).
• Finally, prepare sugar solutions of higher
concentrations (20 g, 30 g and 40 g in 100 ml)
and measure the angles of rotation in the same
way as in the first series.
• Set out the results in a table and plot graphs of
the angle of rotation as a function of
concentration and sample thickness for each
light colour.
6.2 Determine the specific rotation of saccharose
The specific rotation
[ ]
α
is a constant for any given
substance, and for a given light wavelength
λ and
temperature T it is defined by the equation:
[ ]
l c
T
⋅
α
= α
λ
(1)
α = observed angle of rotation
c = concentration of the dissolved substance
l = thickness of the sample solution.
Values given in the literature are usually those for
the yellow D line of sodium (
λ = 589 nm) at a
temperature of 20 °C.
• Prepare the sugar solution (50 g in 100 ml), by
weighing out 50 g of sugar, dissolving it in about
60 cm
3 of distilled water, and making the volume
up to 100 cm 3 in the cylindrical sample cell.
• Measure the sample thickness and place the
sample cell in the measurement chamber.
• Measure the angle of rotation with yellow light.
• Calculate the specific rotation using Equation 1
and compare it with the quoted value.
Quoted values
[ ]
20
D α
for specific rotation:
Saccharose +66.5°
D-glucose +52.7°
D-fructose -92.4°
(Values from Aebi, Einführung in die praktische
Biochemie [Introduction to Practical Biochemistry],
Karger 1982.)
6.3 Inversion of saccharose
Acids cause saccharose to split into D-glucose and D-
fructose, releasing the two components in equal
quantities. During this process the dextro-rotation is
steadily reduced until finally the angle of rotation
becomes negative (anti-clockwise). This phenomenon
is called inversion. The resulting glucose/fructose
mixture is therefore called invert sugar, and is a
constituent of some food products, such as synthetic
honey.
• Start to prepare a sugar solution (30 g in 100 ml),
by first weighing out 30 g of sugar and dissolving
it in about 60 cm
3 of distilled water (50° C).
• Carefully (wearing safety goggles) add 15 ml of
25% hydrochloric acid.
• Make up the volume to 100 cm 3 in the sample
cell and place it in the measurement chamber.
• Immediately start a stop-watch and measure the
angle of rotation.
• Repeat the measurement of the angle of rotation
at intervals of 5 minutes and compile all the
results in a table.
• After 30 minutes, bring your series of
measurements to an end and plot the inversion
curve.
6.4 Measure the concentration of a substance of
known specific rotation - example: cane sugar
in cola
• Fill the sample cell with 100 ml of cola.
• Using the yellow LED, determine the angle of
rotation and its direction.
• Calculate the sugar content using the following
equation obtained by rearrangement of
Equation 1:
[ ]
l
c
⋅ α
α
=
⎥⎦
⎤
⎢⎣
⎡
3
cm
g