Ultraviolet
and visible absorption spectrophotometry is the measurement of the absorption
of monochromatic radiation by solution of chemical substances, in the range of
185 nm to 380 nm, and 380 nm to 780 nm of the spectrum, respectively.
The
magnitude of the absorption of a solution is expressed in terms of the
absorbance, A, defined as the
logarithm to base 10 of the reciprocal of transmittance (T) for monochromatic radiation:
logarithm to base 10 of the reciprocal of transmittance (T) for monochromatic radiation:
For
convenience of reference and for ease in calculations, the specific absorbance
of a 1 percent w/v solution is adopted in this pharmacopoeia for several
substances unless otherwise indicated, and it refers to the absorbance of a 1
per cent w/v solution in a 1 cm cell and measured at a defined wavelength.
It is
evaluated by the expression
A (1%, 1
cm) = A/cl,
Where c is
the concentration of the absorbing substance expressed as percentage w/v and 1
is the thickness of the absorbing substance.
Unless otherwise
stated, measure the absorbance at the prescribed wavelength using a path length
of 1 cm and at 24° to 26°. Unless otherwise stated, the measurements are
carried out with reference to the same solvent or the same mixture of solvents.
Apparatus
A spectrophotometer,
suitable for measuring in the ultraviolet and visible ranges of the spectrum
consists of an optical system capable of producing monochromatic light in the
range of 200 nm to 800 nm and a device suitable for measuring the absorbance.
The two empty
cells used for the solutions under examination and the reference liquid must
have the same spectral characteristics. Where double-beam-recording instruments
are used, the solvent cell is placed in the reference beam.
Control
Of wavelengths
Verify the
wavelength scale using the absorption maxima of holmium perchlorate solution,
the line of hydrogen of deuterium discharge lamp or the lines of a mercury
vapour are shown below. The permitted tolerance is ± 1 nm for the range 200 nm
to 400 nm and ± 3 nm for the range 400 nm to 600 nm
Control
of absorbance
Check the
absorbance using suitable filters or a solution of potassium dichromate UV at
the wavelengths indicated in Table 1, which gives for each wavelength the exact
values and permitted limits of the specific absorbance. The tolerance for the
absorbance is ± 0.01.
Use
solutions of potassium dichromate UV which has been previously dried to
constant weight at 130°. For the control of absorbance at 235 nm, 257 nm, 313
nm and 350 nm, dissolve 57.0-63.0 mg of potassium dichromate UV in 0.005 M
sulphuric acid and dilute to 1000.0 ml with the same acid. For the control of
absorbance at 430 nm, dissolve 57.0-63.0 mg of potassium dichromate UV in 0.005
M sulphuric acid and dilute to 100.0 ml with the same acid.
Limit Of
stray light
Stray light
may be detected at a given wavelength with suitable filters or solutions; for
example, absorbance of a 1.2 per cent w/v solution of potassium chloride in a 1
cm cell should be greater than 2.0 at about 200 nm when compared with water as
reference liquid.
Resolution
power
When stated
in a monograph, record the spectrum of 0.02 per cent v/v solution of toluene in
hexane.
No comments:
Post a Comment