Spectrophotometry

Key Concepts
Lamberts Law of Absorption
Beers Law
Beer-Lambert Law
Absorption Cross-Sections
Photometric quantities
Spectrophotometer
The Cary 50 Spectrophotometer

Lamberts Law of Absorption

Lambert described how intensity changes with distance
in an absorbing medium.
The intensity I0 if a beam of light decreases exponentially as it passes though a
uniform absorbing medium with the linear decay constant .
Restatement: In a uniform absorbing medium, the intensity of a beam of light
decreases by the same proportion for equal path lengths traveled.
The linear decay constant is a characteristic of the medium. It has units of
reciprocal length. is the path length over which the intensity is attenuated to 1/e.

I0

I I 0 e l
I

I ( x ) I 0 e x

I(x)

The distance traveled through the medium is called

the path length.

d I I d x
I I 0e

Johann Heinrich Lambert

1728-1777

dI
I
dx
I
x
e
I0

Photo: http://www-history.mcs.st-andrews.ac.uk/history/PictDisplay/Lambert.html

Lamberts Law of Absorption (base 10)

Typically base 10 is used in photometry.

I I 0e

I 010

kx

k ln 10

I
x
kx
e 10
I0
k is the path length over which the intensity is attenuated to 1/10.

I
10 k x
I0

Lamberts Law Example

If one slab of absorbing material of thickness l reduces the intensity of a beam of light to half.

I0

I
1
10 k l
I0
2

Then two slabs of the same absorbing material will then reduce the intensity of a beam
of light to one quarter.

I0

I
1
10 k 2 l
I0
2

1
4

And three slabs will reduce the intensity of a beam of light to one eight.

l
I0

I
1
k 3l
10

I0
2

Beers Law
Beer found that Lamberts linear decay constant k for a solution of an
absorbing substance is linearly related to its concentration c by a constant,
the absorptivity , a characteristic of the absorbing substance.
Restatement: The linear decay constant k is linear in concentration c with a
constant of proportionality .
(August Beer, 1825-1863)

k c
Typical units are: k cm1; c M (moles/liter); M1cm1
A colored absorber has an absorptivity that is dependent on wavelength of the light ().
The absorptivity is the fundamental property of a substance. This is the property that
contains the observable spectroscopic information that can be linked to quantum
mechanics (also see absorption cross section.)

Photometric Quantities
In photometry we measure the intensity of light and characterize its
change by and object or substance. This change is typically expresses as
percent transmittance or absorbance.

Transmittance (T)
Frequently when your primary
interest is the light beam

I
T
I0

usually given in percent

Absorbance (A) (AKA optical density, O.D.)

Used almost exclusively when your interest
concerns the properties of the material

I
log T
A log
I0

by convention, base 10 logs are used

Beer-Lambert Law
Lamberts and Beers Laws are combined to describe the attenuation of light by a
solution. It is easy to see how the two standard photometric quantities can be written in
terms of this law.

I I 010

c x

Transmittance

Absorbance

I
T
I0

I
log T
A log
I0
A cx

T 10 c x

Cross-Sections and Absorptivity

the connection to single particles and molecules
The absorption of light by particles (and single molecules) is characterized
by an absorption cross section C. In this model the particle is replaced by
a perfectly absorbing sphere with a cross sectional area C. This cross
section is a property of the particle and is not related to its geometric cross
sectional area. The concentration of particles per unit volume is N.

NC
k NC ln 10

typical units are: C cm2; N cm3

N A C ln 10 10 3

liter
3
cm

The cross section can be directly related to the molar

absorptivity. NA is Avagadros number.
units are: C cm2; N cm3; NA mole1; M1cm1

Efficiency
The absorption efficiency Q of a particle is the ratio of its absorption cross
section C to its geometric cross section Cgeo.
Absorption efficiency is dimensionless.

C
Q
C geo

Extension to Scattering and Extinction

Attenuation of light by absorption and scattering both obey Lamberts
Law. Thus we can extend our treatment of absorption to scattering and
extinction. (Recall that extinction is the effect of absorption + scattering.)

Cext Cabs Csca

Qext Qabs Qsca

The scattering efficiency can be much larger than unity.

Extinction paradox: Qext = 2 (Qabs = 1; Qsca = 1)
for an perfectly absorbing particle very large compared to the
wavelength of light.

ext abs sca

A ext cx abs sca cx

Note:
All of these quantities are in general wavelength dependent.
Our discussion has not included the mechanism (cause) of absorption and scattering.
There are many different mechanisms that cause of absorption and scattering.

Instrumentation
Spectrometer: measures I vs .
Simply measures the spectrum of the light (e.g. emission spectroscopy).

Spectrophotometer: measures I/I0 vs .

Measures how the sample changes the spectrum of the light (e.g. transmission,
reflection, scattering, fluorescence).

All spectrophotometers contain a spectrometer.

-meter: the detector is electronic
-graph: light intensity recorded on film
photometer: measures I/I0 without selection.

The Spectrophotometer
Measures absorbance as a function of wavelength
Components: light source, monochromator, sample cell, detector, optical
system.

sample cell
slit

diffraction grating

monochromator

light source

detector

Cary 50 UV-Vis Spectrophotometer

Computer controlled acquisition
of absorption spectra

monochromator
balance the forces:

detector

sample

light
source

Can you find the diffraction

grating and the slit?

www.varianinc.com

Making a Measurement with the Cary 50

First, measure the baseline using a blank sample. This is raw I0. The
blank sample is the cuvette with deionized water (everything but your
nanoparticles). This corrects for any absorption due to the cuvette,
water, and variations of the light intensity of the light source,
monochromator, etc.
Second, measure the zero by inserting the beam block. This corrects
the instrument for the detector background.
Third, measure your sample. This is the raw I. The Cary 50
automatically calculates the corrected intensities (I and I0) by
subtracting the zero from each of the raw intensities.
Subsequent measurements do not require re-measuring the blank and
zero, simply repeat step 3.

I
raw I zero
T
I 0 raw I 0 zero
A log T

Applications of
Spectrophotometry
Spectroscopy
Chemical Analysis:

trace analysis, pH, forensic, in situ monitoring,

remote monitoring, geology, astronomy, ....
Particle size
Thin film characterization
Color matching
Optics