What is the procedure for UV-spectrophotometer calibration?
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Answer Posted / rakesh
1.0 CONTROL OF ABSORBANCES
PROCEDURE: REPARATION OF POTASSIUM DICROMATE SOLUTION: Dry
K2Cr2O7 at 130oC for 2 hours. Weigh accurately about the
quantity of sample equal to 60mg into 1000ml volumetric
flask. Add 0.005m Sulphuric acid to dissolve and dilute
with the same solvent.
Find out the absorbances of (A 1% 1cm) above said solution
at wavelengths of a 235nm, 257nm, 313nm and 350nm.
Calculate the (1% 1cm) value using the following expression.
Absorbance X 1 X 1000 / Weight of K2Cr2O7 in g X 100
Check the value for each wavelength with the tolerances
given four wavelengths.
PREPARATION OF REAGENT:
(i) Dilute 3ml of concentrated sulphuric acid and make up
100ml with distilled water.
(ii) Dilute 20ml to 2000ml with distilled water.
2. LIMIT OF STRAYLIGHT:
The absorbance of a 1.2-% w/v solution of potassium
chloride at a path-length of 1cm should be greater than 2.0
at about 200nm when compared with water as reference liquid.
1.2% POTASSIUM CHLORIDE SOLUTION:
Take 1.2 g of Potassium chloride previously dried at 105oC
for two hours into the 100ml volumetric flask. Add water to
dissolve and make up to volume with water.
3. RESOLUTION POWER:
Record the spectrum of a 0.02% v/v solution of Toluene in
Hexane UV. The ratio of the absorbance at the maximum at
about 269nm to that at the minimum at about 266nm is not
less than 1.5 unless other wise specified in the monograph.
0.02% v/v TOLUENE:
Take 0.02ml of Toluene into the 100ml volumetric flask. Add
Hexane to volume.
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