2007

Instrumental Analysis:Spectrophotometric Methods

•Understand interaction between light and matter

(absorbance, excitation, emission, luminescence,fluorescence, phosphorescence)

•Describe the main components of a spectrophotometer,

(sources, monochromators, detectors, interferometer, grating, ATR, ICP, )

•Make calculations using Beer’s Law(analyse mixture absorption)

•Understand the mechanism and application of UV-Vis, FTIR, Luminescence, atomic spectroscopy

By the end of this part of the course, you should be able to:

Background knowledge:

What you are expected to know before the course:

Error analysis in quantitative analysisSolve linear equationsComplementary colourExponential and logarithm

What you are recommended to know before the course:

Least square fittingBasic quantum chemistryMolecular symmetry

If you have difficulty to understand above topics, find extra reading materials!

Or discuss with me after the lecture.

If you are trying to learn above topics, please let me know.

Today’s lecture: (Instruments based on light interaction with matter)

• Properties of light• Molecular electronic structures• Interaction of photons with molecules• Spectrophotometer components

• Light sources• Single and double beam instruments• Monochrometers• Detectors

• Fluorescence spectroscopy

Next week’s lecture:

• Fourier transformed infrared spectroscopy• Interferometer

• Atomic spectroscopy• Quantitative analysis

• Beer’s law• Method validation• Dilution and spike

Light travelling speed:

in other media: c/n (n = refractive index, generally >1)

in a vacuum: c=2.998 x 108 m s-1 (n=1 exactly, in air n=1.0002926)

c/n=

Therefore:

Energy is inversely proportional to wavelength

but proportional to wavenumber

And of course, the relationship between energy and frequency:

E = h = hc/ = hc h = Planck’s constant (6.626 x 10-34 J s)

= wavenumber (most common units = cm-1)

~

~

Light is energy in the form of electromagenetic field

Review on properties of light:photon

Wavelength (): Crest-to-crest distance between waves Frequency (): Number of complete oscillations that the wave makes each second

units: number of oscillations/sec or s-1 or Hertz |(Hz)

Frequency Scanning Techniques: a few definitions

Emission method: source of light is sample

Absorption method: intensities of a source with and without the sample in place are compared

Spectrum: a plot of intensity vs. frequency/wavelength

In quantitative analysis:

common to work at 1 wavelength

running a spectrum is an important initial step (to select best conditions)

Fig. 18-2

Regions of Electromagnetic Spectrum-the “colour” of light

Electronic structures of simple molecule

S0

S1

T1

Bond length

D

Ground state

Excited stateSinglet

Excited stateTriplet

En

erg

y

Dissociated states

Vib

rati

on

sta

tes

Interaction between photon and molecule

S0

S1

T1

D

S0

S1

T1

S0 S1 transition

A F

IR

UV

-vis

P

Electronic structures

Singlet and triplet

Bond length for ground and excited states

Vibrational structures-infrared absorption/transmission (FTIR)

Internal conversion

Intersystem crossing

Photon adsorption excitation (Beer’s law, UV-vis)

Frank Condon condition and The Stokes' shift

Radionless relaxation and vibration relaxation

Luminescence-fluorescence/phosphorescence

Key concept from energy diagram

Type of optical spectroscopy

UV-vis absorption spectroscopy (UV-Vis)FT-IR absorption/transmission spectroscopy (FTIR)Atomic absorption spectroscopy (AAS)Atomic fluorescence spectroscopy (AFS)X-ray fluorescence spectroscopy (XFS)

What you will learn:

The excitation mechanism

Monochromator design

Instrument principle

Quantitative methods

Optical spectrophotometer components

Monochromators

Filters

Grating+slit

prism

Excitation sources

Deuterium Lamp

Tungsten Lamp

Laser

X-ray tube

Mercury lamp

Xenon lamp

Silicon carbide globar

Flame

Furnaces

Plasmas

Hollow-cathode lamp

Detectors

PMT

CCD/CID

Photodiode

Thermocouple

MCT

Pyroelectric detector

UV

UV-vis

X-ray, UV, vis, IR

X-ray

UV-vis

UV-vis

IR

What is the advantage and disadvantage?

Design of optical spectrophotometersSingle Beam vs. Double Beam

Fig. 13-12, pg. 315 "Instrument designs for photometers and spectrophotometers”

(a) single-beam design

(b) dual channel design with beams separated in space but simultaneous in time

(c) double-beam design in which beams alternate between two channels."

(a)

(b)

(c)

Q: what’s the advantage of double beam spectrophotometer?

Light sources

Black-body radiation for vis and IR but not UV

- a tungsten lamp is an excellent source of black-body radiation

- operates at 3000 K

- produces from 320 to 2500 nm

For UV:

- a common lamp is a deuterium arc lamp

- electric discharge causes D2 to dissociate and emit UV radiation (160 – 325 nm)

- other good sources are:

Xe (250 – 1000 nm)

Hg (280 – 1400 nm)

( How much in cm-1, J, Hz and eV?)

Lasers:

- high power

- very good for studying reactions

- narrow line width

- coherence

- can fine-tune the desired wavelength (but choice of wavelength is limited)

- £££ expensive £££

What is the important properties of a source?

BrightnessLine widthBackgroundStabilityLifetime

Sample a source containers:for UV: quartz (won’t block out the light)

for vis: glass [ 800nm (red) to 400 nm (violet)]

for IR: NaCl (to or 15384 nm or 650 cm-1)

KBr (to 22222 nm or 450 cm-1)

CsI (to 50000 nm or 200 cm-1)

Optical transmission coefficient Best material: diamond, why?

High transmissionChemically inert

Mechanically strong

Criteria

Monochromators

Early spectrophotometers used prisms

- quartz for UV

- glass for vis and IR

These are now superseded by:

Diffraction gratings:

- made by drawing lines on a glass with a diamond stylus

ca. 20 grooves mm-1 for far IR

ca. 6000 mm-1 for UV/vis

- can use plastic replicas in less expensive instruments

Think of diffraction on a CD

http://www.mrfiber.com/images/cddiffract.jpg

http://www.ii.com/images/prism.jpg

http://www.veeco.com/library/nanotheater_detail.php?type=application&id=331&app_id=34

10mx10m

Why?

Monochromators: cont’d

Polychromatic radiation enters

Second concave mirror focuses each wavelength at different point of focal plane

Orientation of the reflection grating directs only one narrow band of wavelengths to exit slit

The light is collimated the first concave mirror

Reflection grating diffracts different wavelengths at different angles

http://oco.jpl.nasa.gov/images/grating_spec-br.jpg

What is the purpose of concave mirrors?

Interference in diffraction

d

d sin()+d sin()=n

n=1, 2, 3 In-phase

n=1/2, 3/2, 5/2 out-phase

>0<0

Bragg condition

Phase relationship

Monochromators: reflection grating

Monochromators: reflection grating

Each wavelength is diffracted off the grating at a different angle

Angle of deviation of diffracted beam is wavelength dependent diffraction grating separates the incident beam into its constituent wavelengths components

Groove dimensions and spacings are on the order of the wavelength in question

In order for the emerging light to be of any use, the emerging light beams must be in phase with each other

Resolution of grating:

=nN

Angular resolution:As: d sin()+d sin()=n

So: n =d cos()

Therefore: =n/[d cos()]

What does this mean?

n: diffraction orderN: number of illuminated groves

Monochromators: slit

Bottom line:

- it is usually possible to arrange slits and mirrors so that the first order (n = 1) reflection is separated

- a waveband of ca. 0.2 nm is obtainable

However, the slit width determines the resolution and signal to noise ratio

Large slit width: more energy reaching the detector higher signal:noise

Small slit width: less energy reaching the detector BUT better resolution!

Detectors

Choice of detector depends upon what wavelength you are studying

Want the best response for the wavelength (or wavelength range) that you are studying

In a single-beam spectrophotometer, the 100% transmittance control must be adjusted each time the wavelength is changed

In a double-beam spectrophotometer, this is done for you!

: Radiation-----charger converter

Photomultiplier-single channel, but very high sensitivity

- Light falls on a photosensitive alloy (Cs3Sb, K2CsSb, Na2KSb)

- Electrons from surface are accelerated towards secondary electrodes called dynodes and gain enough energy to remove further electrons (typically 4-12, to 50 with GaP).

- For 9 stages giving 4 electrons for 1, the amplification is 49 or 2.6 x 105)

- The output is fed to an amplifier which generates a signal

- To minimise noise it is necessary to operate at the lowest possible voltage

What decide the sensitive wavelength?

Photodiode Array-multiplex, but low sensitivity

Good for quick (fraction of a second) scanning of a full spectrum

Uses semiconductor material:

Remember: n-type silicon has a conduction electron – P or As doped

p-type silicon has a ‘hole’ or electron vacancy – Al or B doped

A diode is a pn junction:

under forward bias, current flows from n-Si to p-Si

under reverse bias, no current flows

boundary is called a depletion layer or region

Photodiode Array

- Electrons excited by light partially discharge the condenser

- Current which is necessary to restore the charge can be detected

- The more radiation that strikes, the less charge remains

- Less sensitive than photomultipliers several placed on placed on single crystal

- Different wavelengths can be directed to different diodes

- Good for 500 to 1100 nm

- For some crystals (i.e. HgCdTe) the response time is about 50 ns

Could you compare photodiode with CCD detector?

Photodiode Array Spectrophotometer

- For photodiode array spectrophotometers, a white light passes through sample

- The grating polychromator disperses the light into the component wavelengths

- All wavelengths are measured simultaneously

- Resolution depends upon the distance between the diodes and amount of dispersion

No moving parts! Simple mechanical and optical design, very compact.

Photodiode Array Spectrophotometers vs Dispersive Spectrophotometers

Dispersive Spectrophotometer:

- only a narrow band of wavelengths reaches the detector at a time

- slow spectral acquisition (ca. 1 min)

- several moving parts (gratings, filters, mirrors, etc.)

- resolution: ca. 0.1 nm

- produces less stray light greater dynamic range for measuring high absorbance

- sensitive to stray light from outside sources i.e. room light

Photodiode Array Spectrophotometer:

- no moving parts rugged

- faster spectral acquisition (ca. 1 sec)

- not dramatically affect by room light

What are the components 1 to 10?From: http://www.oceanoptics.com/

Property of luminescence spectrum

Fluorescence vs phosphorescence

1. Phosphorescence is always at longer wavelength compared with fluorescence2. Phosphorescence is narrower compared with fluorescence3. Phosphorescence is weaker compared with fluorescence

Absorption vs emission

1. absorption is mirrored relative to emission2. Absorption is always on the shorter wavelength compared to emission3. Absorption vibrational progression reflects vibrational level in the electronic excited

states, while the emission vibrational progression reflects vibrational level in the electronic ground states

4. 0 transition of absorption is not overlap with the 0 of emission

Why?

Why?

Fluorescence spectroscopy

Fluorescence spectroscopy

Emission spectrum: hold the excitation wavelength steady and measure the emission at various wavelengths

Excitation spectrum: vary the excitation wavelength and vary the wavelength measured for the emitted light

Light source

Excitationmonochromator

Referencediode

8% of li

ght

Beamsplitter

sample

EmissionMonochromator

Amplifier ComputerPMT

Q: why the emission is measured at 90 relative to the excitation?

Fluorescence spectroscopy: well defined molecules

• Describe the main components of a spectrophotometer and distinguish between single double beam instruments

• Describe suitable sources for ultraviolet (UV)/visible (vis), infra red (IR) and atomic absorption (AA) instruments

• Describe and assess advantages and disadvantages of various monochromators e.g. Prism, diffraction gratings

• Explain how to asses the quality of grating

• Explain how photomultipliers and diode detectors work• Explain the advantage of multiplex detecting• Describe the luminescence spectroscopy and energy transfer process• Compare the emission and absorption spectrum

Summary of spectrophotometric techniques