Chapter 20

Spectrophotometers
 Overview

20-1 Lamps and Lasers: Sources of Light

20-2 Monochromators
20-3 Detectors
20-4 Optical Sensors
20-5 Fourier Transform Infrared Spectroscopy
20-6 Dealing with Noise
 Spectrophotometers
In a single-beam spectrophotometer:
• Light from a source is separated into a narrow
band of wavelength by a monochromator, passed
through the sample in a sample holder, and
measured by the detector.
• Using the reference cell containing the solvent or
reagent blank, we measure the irradiance P0
striking the detector.
• We replace the blank with the sample and if it
absorbs some radiation, we measure the
irradiance P striking the detector.
 Spectrophotometers
• P/P0, which is a number between 0 and 1, is the transmittance,
T. Instruments convert this to absorbance, which is
proportional to concentration.

Disadvantages of single-beam instruments:

• The sample and the reference must be placed alternately in the

beam.

• For measurements at multiple wavelengths, the reference must

be run at each wavelength.

• Poor for measuring absorbance versus time for kinetics studies

because both the source and the detector response slowly drift.
 Spectrophotometers
A double-beam spectrophotometer:
• The beam is alternately sent through the reference and
sample cells.
• It is chopped several times a second.
• When light passes through the sample, P is measured;
through the reference, P0 is measured.
• P is compared to P0, T is obtained and converted to A.
 Spectrophotometers
In this chapter, we will consider the components of
spectrophotometers (Figures 20-1 and 20-2):

1. Light sources – lamps and lasers

2. Wavelength selectors – gratings and monochromators
3. Detectors – photomultiplier tubes, photodiodes, and
charged coupled devices for UV-vis and thermocouples,
ferroelectric materials, and photoconductive detectors for
IR.

We will also consider Fourier transform infrared

spectroscopy and the use of interferometers to improve the
speed of IR spectroscopy.
Spectrophotometers
 20-1: Lamps and Lasers: Sources of Light

• A tungsten lamp is an excellent

source of continuous visible and
near-IR radiation, giving useful
radiation in the range 320 to 2500
nm.
• UV usually employs a deuterium
arc lamp in which an electric
discharge (spark) causes D2 to
dissociate and emit UV radiation
from 110 to 400 nm.
• In a UV-vis instrument, a change is
made between deuterium and
tungsten lamps when passing
through 360 nm so that the lamp
of highest intensity is always used.
 20-1: Lamps and Lasers: Sources of Light

• Infrared radiation (4 000 to 200 cm-1) is obtained

using a silicon carbide glowbar, which emits
approximately the same spectrum as a blackbody at
1 000 K.

• Lasers give isolated lines of a single wavelength.

• Laser light is:
1. Monochromatic
2. Extremely bright
3. Collimated
4. Polarized
5. Coherent
 20-1: Lamps and Lasers: Sources of Light

• A necessary condition for lasing is population

inversion, in which a higher energy state has
a greater population, n, than a lower energy
state in the lasing medium.
• In the figure, this occurs when the population
of E2 exceeds that of E1.

• Molecules in the ground state, E0, of the

lasing medium are pumped to excited state
E3 by broadband radiation from a powerful
lamp or an electric discharge.
 20-1: Lamps and Lasers: Sources of Light

• Molecules in E3 rapidly relax to E2, which has a relatively long

lifetime.
• After a molecule in E2 decays to E1, it rapidly relaxes to the
ground state, E0, thereby keeping the population of E2 greater
than the population of E1.
 20-1: Lamps and Lasers: Sources of Light

• A photon with an energy that exactly spans two states can

be absorbed to raise a molecule to an excited state.

• Alternatively, the same photon can stimulate the excited

molecule to emit a photon and return to the ground state.
This is called stimulated emission.

• When a photon emitted by a molecule falling from E2 to E1

strikes another molecule in E2, a second photon can be
emitted with the same phase and polarization as the first
photon.

• If there is population inversion n2 > n1, one photon stimulates

the emission of many photons as it travels through the laser.
 20-1: Lamps and Lasers: Sources of Light

• Pump energy is directed through the side of the lasing

medium to create the population inversion.
• One end of the laser cavity is a mirror that reflects all
light (0% transmittance). The other end is a partially
transparent mirror that reflects most of the light (1%
transmittance).

• Photons with energy E2 – E1 that bounce back and forth

between the mirrors stimulate an avalanche of new
photons.

• The small amount of light passing through the mirror on

the right is the useful output of the laser.
 20-2: Monochromators

• A monochromator disperses light into its component

l’s and selects a narrow band to pass on to the
sample or detector.
• A grating has closely ruled lines, each behaving as a
separate source of radiation.
• Different wavelengths of light are diffracted at different
angles from the grating.
• Rotation of the grating allows different wavelengths to
reach the sample.
• The reflection grating shown in Figure 20-6 is ruled
with a series of closely spaced, parallel grooves with
repeat distance, d.
 20-2: Monochromators

• When adjacent light waves are in phase, they

reinforce each other. When they are not in
phase, they partially or completely cancel each
other.
 20-2: Monochromators

• Fully constructive interference occurs when the

difference in the length of the two paths is an integral
multiple of the wavelength of light.
• In Figure 20-6, the difference is a - b and constructive
interference occurs when:
nλ  a  b
n is the diffraction order and a = dsinθ and b = -dsinФ

• Substituting this in for constructive interference, the

grating equation is:
nλ  d (sinθ  sin  )
 20-2: Monochromators

• Resolution measures the ability to separate two

closely spaced peaks.

• The greater the resolution, the smaller the difference

(Δ  ) between two wavelengths that can be
distinguished from each other.

The resolution of a grating is given by the equation:

• The greater the 

number of grooves Resolution   nN
(smaller the 
spacing), the higher
the resolution.
 20-2: Monochromators

• To select a narrower band of l’s from the

monochromator, we decrease the exit slit width.
• A narrower bandwidth gives better performance.
• Decreasing the exit slit width decreases the
selected bandwidth (wavelengths) and decreases
the energy to the detector.
• There is a trade-off between resolution and signal.

• The narrower the exit slit, the greater the ability to

resolve closely spaced peaks and the noisier the
spectrum. En (grating)
Relative efficiency 
E (grating)
 20-2: Monochromators

• Stray light – l’s outside the bandwidth expected

from the monochromator reaches the detector
from:

1.Unwanted diffraction orders

2.Outside of the instrument when the sample
compartment is not perfectly sealed.

• Error from stray light is most serious when sample

absorbance is high.

• Filters are used to permit only certain bands of l’s

from passing through.
 20-3: Detectors

• A detector produces an electric signal when it is

struck by photons.
• Detector response is a function of l of incident
light.

• A Photomultiplier Tube contains a photoemissive

surface (photocathode) as well as several
additional surfaces (dynodes) that emit a cascade
of electrons when struck by an electron from a
photosensitive area.
 20-3: Detectors

• Photon strikes the

photosensitive surface and
electrons are ejected.
• They are accelerated and
strike the dynode with
more than their original
kinetic energy.
• Each dynode is more
positive than the previous.
• Each electron causes
emission of several
additional electrons.
• 106-107 electrons reach the
anode for each incident
photon.
 20-3: Detectors

• Instruments with photomultiplier tube detectors scan

through a spectrum one l at a time.
• A photodiode array is essential for rapid
spectroscopy.
• Rows of p-type silicon on a substrate of n-type
silicon create a series of pn junction diodes.
• A reverse bias is applied to each diode, drawing
electrons and holes away from the junction. There
is a depletion region at each junction, in which there
are few electrons and holes.
• The junction acts as a capacitor with charge stored
on either side of the depletion region.
 20-3: Detectors

• At the beginning of a measurement, each diode is fully

charged.
• Photons absorbed in the semiconductor create mobile
electron-hole pairs.
• The more radiation that strikes each diode, the less charge
remains at the end of the measurement.
• The state of each diode is determined at the end of the
cycle by measuring the current needed from the instrument
to recharge the diode.
 20-3: Detectors

• A photodiode array spectrophotometer records the

entire spectrum at once in a fraction of a second.
• White light is passed through the sample, then a
polychromator disperses light into its component
wavelengths and directs the light at the photodiode
array.
• Each diode receives a different wavelength.
 20-3: Detectors

Advantages:
• Allow faster spectral acquisition (<1 s) than dispersive
instruments, which require several minutes.
• Have almost no moving parts.

Disadvantages:
• The resolution of ~0.1nm attainable with a dispersive
instrument and the wavelength accuracy are better than
those of a photodiode array (~0.5-1.5 nm).
• Stray light is less in a dispersive instrument than a diode
array instrument.
 20-3: Detectors

A charge coupled device is an extremely sensitive detector

that stores photo-generated charge in a two-dimensional
array.
• Constructed of p-doped Si on an n-doped substrate.
• Capped with an insulating layer of SiO2, on top of which
is a pattern of conducting Si electrodes.
• Light absorbed in the p-doped region introduces an
electron into the conduction band and a hole is left in the
valence band.
• The electron is attracted to and stored at the positive
electrode.
• The hole migrates to the n-doped substrate.
 20-3: Detectors

• Electrodes store 105 electrons before they spill out into

adjacent elements.
• Electrons stored in each pixel of the top row are moved into
the serial register at the top and charge is read.
• The next row moves up and is read until the entire array has
been read.
• Charge transfer is efficient – loss of only five of every 106
electrons.
 20-3: Detectors

Infrared detectors: Infrared photons are not energetic

enough to eject electrons from a photosensitive surface
or promote electrons from the valence band of Si to the
conduction band.

• Thermocouples are junctions of specific alloys that

have a predictable and reproducible relationship
between temperature and voltage.
• If a thermal couple is blackened to absorb radiation,
its temperature and therefore voltage becomes
sensitive to radiation.
• These convert a temperature gradient into electricity.
 20-3: Detectors

• In a ferroelectric material, such as deuterated

triglycine sulfate (DTGS), the dipole moments of
molecules remain aligned in the absence of an
external field.
• This gives the material a permanent electric
polarization.
• When IR radiation is absorbed, the polarization
changes and a voltage develops across the
material.
• DTGS is a common detector in Fourier transform
spectrometers.
 20-3: Detectors

• Photoconductive detectors
are semiconductors have a
small band gap that
corresponds to IR
energies.
• When they absorb a
photon, an electron is
excited into the conduction
band and collected at an
electrode.
• IR – mercury/cadmium
telluride (MCT) and NIR –
lead sulfide (PbS) and
indium antimonide (InSb).
 20-4: Optical Sensors

• When light passes from one medium into another,

its path is refracted (bent) and is described by
Snell’s law: n sinθ  n sinθ
1 1 2 2
• Optical fibers carry light by total internal reflection.
• The fiber has a high-refractive-index transparent
core (n1) enclosed in a lower-refractive index
transparent cladding (n2):
 20-4: Optical Sensors

• When n1 (core) > n2 (cladding), Snell’s law:

n1
n1 sinθ i  n 2 sinθ r  sin θ r  sin θ i
n2

• If n1/n2 >1, there is a range of i in which

essentially all of the light is reflected at the walls of
the core and none enters the cladding.
 20-4: Optical Sensors

• Optical sensors for specific analytes are made by

placing a chemically sensitive layer at the end of a
fiber; called optodes.
• A flat material in which light is totally reflected
because n1 (material) > n2 (surroundings) is called
a waveguide.
• Sensors can be made by putting a chemically
sensitive layer onto a waveguide.
• The light is totally reflected if i exceeds critical =
n2/n1.
• The electric field penetrates the cladding to a
small degree.
 20-4: Optical Sensors

• Evanescent wave – the part

of the light that penetrates
the wall.
• Figure 20-24a shows a
sensor based on attenuated
total reflectance.
• The evanescent wave
extends into the sorbent
beads.
 20-4: Optical Sensors

• When caffeine is present • Each peak in the

l’s absorbed by caffeine spectrum arises from a
are attenuated. vibrational mode of
caffeine.
 20-4: Optical Sensors

• A surface plasma
wave (surface
plasmon) propagates
along the boundary
between a metal and
an insulator
(dielectric).
• Figure 20-25 shows
the essentials of a
surface plasmon
resonance
measurement.
 20-5: Fourier Transform Infrared Spectroscopy

• To observe all wavelengths at once and detect

them simultaneously, an interferometer is used to
produce an interference pattern that contains all of
the IR data.

An interferometer has the following parts:

1. Collimated light source
2. Stationary mirror
3. Movable mirror
4. Beam splitter
5. Detector
 20-5: Fourier Transform Infrared Spectroscopy

• Source radiation strikes the beamsplitter at point O.

• 50% is reflected to a stationary mirror at distance OS.
• 50% is transmitted to a movable mirror at distance
OM.
 20-5: Fourier Transform Infrared Spectroscopy

• The beams travel back to the beamsplitter and

half of each is transmitted and half reflected.
• One recombined wave travels toward the detector
and one travels toward the source.
• Paths OS and OM are not equal.
• The movable mirror is moved to the left at a
constant velocity.
• The difference in paths between the light beams
2(OM-OS) is called retardation, d.
• Constructive interference occurs whenever d is an
integer multiple of the wavelength.
 20-5: Fourier Transform Infrared Spectroscopy

• A minimum appears
(destructive interference)
when d is a half-integer
multiple of the wavelength.
• Light reaching the detector
goes through a series of
maxima and minima.
• A graph of output light
intensity versus retardation,
d, is called an interferogram.
• Fourier analysis of the
interferogram gives the
spectrum.
 20-6: Dealing with Noise

• Signal averaging can

improve the quality of
data.
• The signal is measured
from the middle of the
baseline noise to the
middle of the noisy
peak.
• By this definition, the
bottom trace in the
figure has a signal-to-
noise ratio of 14/9 =
1.6.
 20-6: Dealing with Noise

• A more common measurement of noise, which

requires a digitized signal and a computer, is the
root-mean-square (rms) noise:
 i
(A  A ) 2

rms noise  i
n
• The rms noise is ~5 times less than the peak-to-
peak noise.

• Using this, the noise in the bottom scan is 9/5 =

1.8, and the signal-to-noise ratio is 14/1.8 = 7.8.
 20-6: Dealing with Noise

• When the spectrum is recorded twice and the results

added, the signal is the same in both spectra and adds
to give twice the value of each spectrum.
• Collecting n spectra increases the signal by n times.
• Noise is random, so it may be positive or negative at any
point. If n spectra are recorded, the noise increases in
proportion to the square root of n.
• The signal-to-noise ratio therefore increases by:

n
 n
n
• So, in order to increase the signal-to-noise ratio by a
factor of 10, we must average 100 spectra.
 20-6: Dealing with Noise

Types of noise
1. White noise – normal, random,
Gaussian noise arises from
causes such as random motions
of electrons within a circuit.
2. 1/f noise – also called drift,
greatest at zero frequency and
decreases in proportion to
1/frequency.
3. Line noise – also called
interference or whistle noise, at
discrete frequencies such as the
60-Hz transmission-line
frequency. 44