UV-Visible and Infrared Spectroscopic Methods in Environmental Analysis - Fundamentals of Environmental Sampling and Analysis - page 205

For example, a blank sample without a light-absorbing chemical should be 100%
transparent, that is, T ¼ 100% or A ¼ 0.
When a calibration curve, that is, a plot of A vs. known concentrations ( C )of
standard solutions ( A ¼ a C þ b) is acquired, a linear range is always sought for
the measurement of analyte concentration in samples (Chapter 2). Care should
be excised on the limitations of the Beer's law. The linearity of
the Beer's law is sometimes deviated by chemical and instrumental factors,
and the analyst should be aware of the solutions. The nonlinearity may be
caused by: (a) electrostatic interactions between molecules in close proximity,
(b) scattering of light caused by particulates in the sample, (c) fluorescence or
phosphorescence of the sample, (d) changes in refractive index at high analyte
concentration, (e) shifts in chemical equilibria as a function of concentration,
(f) nonmonochromatic radiation (this deviation can be minimized by using a
relatively flat part of the absorption spectrum such as the maximum of an
absorption band), (g) stray light that reaches the detector without passing
through the sample.
With IR, instrumental deviations from the Beer's law are more common than
with UV and visible wavelengths. This is because infrared absorption bands are
relatively narrow. For dispersive IR (Section 8.3.2), the low intensity of sources and
low sensitivities of detectors in this region require the use of relatively wide
monochromator slit widths, leading to nonlinear relationship between absorbance
and concentration.

EXAMPLE 8.1. A spectrometric method was developed to determine nitrite (NO 2 ,
MW¼46 g/mol) in human saliva and rain water (Helaleh and Korenaga, J. AOAC Int.,
84:53-58, 2001). The method was reported to have an absorption maximum of 546 nm
and molar absorptivity of 4:610 4 L/(mol cm). What range of concentrations can be
measured, so the absorbance remains within the range 0.05-0.80, using a 1.0 cm cell?

SOLUTION: For A ¼ 0:05,

A
e l ¼

0:05
ð4:6110 4 L=cm=molÞ1cm ¼ 1:0810 6 mol=L ¼ 0:050 mg=mL

C ¼

For A¼0.80,

A
e l ¼

0:80
ð4:6110 4 L=cm=molÞ1cm ¼ 1

7410 5 mol

C ¼ : = L ¼ 0 : 798 m g = mL

Note that a conversion factor of 461000 is used to convert mol/L into mg/mL. In the
absorbance range 0.05-0.80, the above calculated concentration range compares favorably
with the linear range (0.054-0.816 mg/mL) reported in this paper.

EXAMPLE 8.2. Beer's law for multiple components. Permanganate (KMnO 4 ; l¼525nm)
and chromate (K 2 Cr 2 O 7 ; l ¼ 440 nm) can be determined in the same sample by visible
light spectroscopy. The absorbance readings of two standard solutions (0.01 M KMnO 4
and 0.02 M K 2 Cr 2 O 7 ) and a sample containing two analytes obtained from a UV-VIS