UltravioletVisible Spectroscopy

UV-vis spectroscopy
- involves the spectroscopy of photons; uses light in the visible, UV and near infrared (NIR) ranges, where molecules undergo electronic transitions. - used in the quantitative determination of solutions of transition metal ions and highly conjugated organic compounds
2

Spectral Distribution of Radiant Energy

Wave Number (cycles/cm)

X-Ray 200nm

UV 400nm

Visible 800nm

IR

Microwave

WAVELENGTH(nm)

Molecules have quantized energy levels: ex. electronic energy levels. hv energy energy

} = hv

Q: Where do these quantized energy levels come from? A: The electronic configurations associated with bonding.
Each electronic energy level (configuration) has associated with it the many vibrational energy levels.

s* p*

2p
p s

2p

s* 2s

2s s

Ethane

C C

s*
hv

s* s

s
H C C H H H H H

s*

max = 135 nm (a high energy transition)

Absorptions having max < 200 nm are difficult to observe because everything (including quartz glass and air) absorbs in this spectral region.

C C

s* p*
hv

s* p*

= hv =hc/
p s

p s

p*

Example: ethylene absorbs at longer wavelengths: max = 165 nm = 10,000

C O

s* p*
n
hv

s* p*
n

p s
n

p s

p*

The n to pi* transition is at even lower wavelengths but is not as strong as pi to pi* transitions. It is said to be forbidden. Example: Acetone: ns* max = 188 nm ; = 1860 np* max = 279 nm ; = 15

Instrumentation

Instrument Parts
Source of visible light quartz halogen lamp (same to automobile) Source of ultraviolet light deuterium arc lamp (200 400 nm) Grating 1 selects a narrow band of wavelengths to enter monochromator Monochromator selects an even narrower bandto pass through the sample Photomultiplier tube detector; creates electric current proportional to P

Light Sources
UV Spectrophotometer
1. 2. 3. Hydrogen Gas Lamp Mercury Lamp Deuterium (200-400 nm)

Visible Spectrophotometer

1.
2. 1.

Tungsten Lamp
Xenon Arc lamps (200-1000 nm) Carborundum (SiC)

InfraRed (IR) Spectrophotometer

TUNGSTEN LAMP
Excellent source of continuous visible and near IR radiation Operates at 3000 K Useful range: 320 2500 nm

Deuterium arc lamp

Source of UV radiation Electric dicharge (spark) causes D2 to dissociate and emit ultraviolet radiation Useful range: 200 400 nm

Mercury and Xenon lamps

Electric discharge (spark) lamps Source of UV and visible radiation

Light emitting diodes (LED)

Provide narrow bands of visible and near IR radiation

Carborundum / Globar
Source of IR radiation Useful range: 4000 200 cmMade up of silicon carbide Heated to 1500 K by electric current

LASER (as a source of radiation)

Assignment! Discuss the principles, advantages and disadvantages (if any)

Fixed Wavelength Instrument

LED serve as source Pseudo-monochromatic light source No monochromator necessary/ wavelength selection occurs by turning on the appropriate LED 4 LEDs to choose from sample beam of light LEDs photodiode

Scanning Instrument
Scanning Instrument monochromator

Tungsten Filament (vis)

slit

slit

Photomultiplier tube cuvette

Deuterium lamp Filament (UV)

Diode array Instrument

mirror Diode array detector 328 individual detectors Tungsten Filament (vis) slit

slit

cuvette Deuterium lamp Filament (UV)

monochromator

Advantages/disadvantages
Scanning instrument
High spectral resolution Long data acquisition time (several minutes) Low throughput (amount of work performed by a computer within a given time)

Diode array
Fast acquisition time (a couple of seconds), compatible with on-line separations High throughput (no slits) Low resolution (2 nm)

Definition of Resolution

Spectral resolution is a measure of the ability of an instrument to differentiate between two adjacent wavelengths

UV / visible Spectroscopy

UV / visible Spectroscopy

ALWAYS
use in conjunction with NMR and infrared spectra.

HPLC-UV
HPLC Pump

Mobile phase

Sample loop

6-port valve

HPLC column UV detector

syringe Solvent waste

UV / visible Spectroscopy
1. Bathochromic shift (red shift)
lower energy, longer wavelength

CONJUGATION.

2. Hypsochromic shift (blue shift)

higher energy, shorter wavelength.

3. Hyperchromic effect
increase in intensity

4. Hypochromic effect
decrease in intensity

 Every instrument has a useful range for a particular analyte. Often, you must determine that range experimentally. This is done by making a dilution series of the known solution. These dilutions are used to make a working curve.

Make a dilution series of a known quantity of analyte and measure the Absorbance. Plot concentrations v. Absorbance.

What concentration do you think the unknown sample is?

In this graph, values above A=1.0 are not linear. If we use readings above A=1.0, graph isnt accurate.

The best range of this spectrophotometer is A=0.1 to A=1.0, because of lower errors. A=0.4 is best.

Relating Absorbance and Transmittance

Absorbance rises linearly with concentration. Absorbance is measured in units. Transmittance decreases in a nonlinear fashion. Transmittance is measured as a %. Absorbance = log10
(100/% transmittance)

Precision and Accuracy

Precision Accuracy

Precision + Accuracy

Precision Accuracy +

Precision + Accuracy +

MONOCHROMATOR
Disperses light into its component wavelengths and selects a narrow band of wavelengths to pass on to the sample or detector Consists of entrance and exit slits, mirrors and grating to disperse light

GRATING
A reflective or transmissive optical component with a series of closely ruled lines Different wavelengths of light are reflected or transmitted at different angles from the grating

Czerny-Turner Grating Monochromator

DIFFRACTION
Bending of light rays by a grating

REFRACTION
Bending of light rays by a prism or lens

GRATING
Made up of parallel grooves with distance d Aluminum coating reflectivity SiO2 coating protection * When adjacent rays are IN PHASE, they reinforce one another and provide a source of radiation

Resolution of Grating
Resolution ability to separate two closely spaced peaks / = nN = wavelength n = diffraction order N = number of grooves that are illuminated * The greater the resolution, the smaller the difference between 2 s that can be distinguished from each other.

Dispersion
Measures the ability to separate wavelengths differing by through the difference in angle f (radians)

Dispersion Devices
Non-linear dispersion Temperature sensitive

Linear Dispersion Different orders

Dispersion of polychromatic light with a prism

Infrared

monochromatic Ray
Red Orange Yellow Green Blue Violet

SLIT

Polychromatic Ray

PRISM

Ultraviolet

Polychromatic Ray

Monochromatic Ray

Prism - spray out the spectrum and choose the certain wavelength () that you want by moving the slit.

Stray Light
Wavelengths outside the bandwidth expected from the monochromator Caused by diffraction into unwanted orders and angles; unintended scattering; sample compartment not perfectly sealed Results to an increased measured absorbance

Stray Light Sample Problem

If the true absorbance of a sample is 2.00 and theres 1.0% stray light, find the apparent absorbance. Tapp = P + S / Po + S = 0.010 + 0.010 / 1 + 0.010

Filters
Remove/block wide bands of radiation from a signal to give the desired wavelength Placed in the beam Simplest types:
Colored glass Interference filters
Pass radiation in reference region; reflect other s

Wide Band-pass filter

Narrow Band-pass filter

DETECTORS

Detector
Produces an electrical signals when struck by photons Ex: phototube emits electrons from a photosensitive, negatively charged surface when struck by visible light/UV rays Detector response is dependent on the wavelength of incident photons

Photomultiplier Tube Detector

Electrons emitted from photosensitive surface strike another surface (dynode) High sensitivity at low light levels Cathode material determines spectral sensitivity Good signal/noise Shock sensitive

Anode

The Photodiode Detector

Wide dynamic range Very good signal/noise at high light levels Solid-state device

Schematic Diagram of a Photodiode Array

Same characteristics as photodiodes Solid-state device Fast read-out cycles

Conventional Spectrophotometer

Schematic of a conventional single-beam spectrophotometer

Conventional Spectrophotometer

Optical system of a double-beam spectrophotometer

Conventional Spectrophotometer

Optical system of a split-beam spectrophotometer