instrumental analysis chem 4811
DESCRIPTION
INSTRUMENTAL ANALYSIS CHEM 4811. CHAPTER 5. DR. AUGUSTINE OFORI AGYEMAN Assistant professor of chemistry Department of natural sciences Clayton state university. CHAPTER 4 ULTRAVIOLET AND VISIBLE MOLECULAR SPECTROSCOPY (UV-VIS). UV-VIS SPECTROSCOPY. - PowerPoint PPT PresentationTRANSCRIPT
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INSTRUMENTAL ANALYSIS CHEM 4811
CHAPTER 5
DR. AUGUSTINE OFORI AGYEMANAssistant professor of chemistryDepartment of natural sciences
Clayton state university
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CHAPTER 4
ULTRAVIOLET AND VISIBLE
MOLECULAR SPECTROSCOPY
(UV-VIS)
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UV-VIS SPECTROSCOPY
- Solutions allow a component of white light to pass through and absorb the complementary color of the component
- The component that passes through appears to the eye as the color of the solution
- This chapter deals with molecular spectroscopy (absorption or emission of UV-VIS radiation by molecules
or polyatomic ions)
- The spectrum is absorbance or transmittance or molar absorptivity versus wavelength
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Complementary Colors
λmax
380-420420-440440-470470-500500-520520-550550-580580-620620-680680-780
Color Observed
Green-yellow YellowOrange
RedPurple-red
VioletViolet-blue
BlueBlue-green
Green
Color Absorbed
VioletViolet-blue
BlueBlue-green
GreenYellow-green
YellowOrange
RedRed
UV-VIS SPECTROSCOPY
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Complementary Colors
UV-VIS SPECTROSCOPY
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Complementary Colors
Ru(bpy)32+
λmax = 450 nmColor observed with the eye: orange
Color absorbed: blue
Cr3+-EDTA complexλmax = 540 nm
Color observed with the eye: violetColor absorbed: yellow-green
UV-VIS SPECTROSCOPY
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UV RADIATION
- Wavelength range is 190 nm – 400 nm
- Involved with electronic excitations
- Radiation has sufficient energy to excite valence electrons in atoms and molecules
- Vacuum UV spectrometers are available that uses radiation between 100 Å – 200 nm (also electronic excitations)
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VISIBLE RADIATION
- Wavelength range is 400 nm – 800 nm
- Involved with electronic excitations
- Similar to UV
- Spectrometers therefore operate between 190 nm and 800 nm and are called UV-VIS spectrometers
- Can be used for qualitative identification of molecules
- Useful tool for quantitative determination
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ELECTRONIC EXCITATION
- Electrons in molecules move in molecular orbitals at discrete energy levels
- Energy levels are quantized
- Molecules are in the ground state when energy of electrons is at a minimum
- The molecules can absorb radiation and move to a higher energy state (excited state)
- An outer shell (valence) electron moves to a higher energy orbital
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ELECTRONIC EXCITATION
- Is the process of moving electrons to higher energy states
- Radiation must be within the visible or UV region in order to cause electronic excitation
- The frequency absorbed or emitted by a molecule is given as
ΔE = hνΔE = E1 – Eo
E1 = excited state energyEo = ground state energy
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ELECTRONIC EXCITATION
Three Distinct Types of Electrons Involved in Transition
Electrons in a Single Bond (Alkanes)- Single bonds are called sigma (σ) bonds
- Amount of energy required to excite electrons in δ bonds are higher than photons with wavelength greater than 200 nm
- Implies alkanes and compounds with only single bonds do not absorb UV radiation
- Used as transparent solvents for analytes
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ELECTRONIC EXCITATION
Three Distinct Types of Electrons Involved in Transition
Electrons in Double or Triple Bonds (Unsaturated)- Alkenes, alkynes, aromatic compounds
- These bonds are called pi (π) bonds
- π bond electrons are excited relatively easily
- These compounds absorb in the UV-VIS region
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ELECTRONIC EXCITATION
Three Distinct Types of Electrons Involved in Transition
Electrons Not Involved in Bonding Between Atoms- Called the n electrons (n = nonbonding)
- Organic compounds containing N, O, S, X usually contain nonbonding electrons
- n electrons are usually excited by UV-VIS radiation
- Such compounds absorb UV-VIS radiation
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En
ergy
s orbital
- Two s orbitals on adjacent atoms overlap to form a σ bond- Two molecular orbitals is the result
- Sigma bonding orbital (σ) is of lower energy than the atomic orbitals (filled with the two 1s electrons)
- Sigma antibonding orbital (σ*) is of higher energy than the atomic orbitals (empty)
ΔE = energy difference between σ and σ*
σ*
σ
ΔE
s orbital
ELECTRONIC EXCITATION
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ELECTRONIC EXCITATION
- p orbitals of atoms can also overlap along axis to form sigma bonds
- There are three p orbitals in a given subshell
- One of these p orbitals from adjacent atoms form sigma orbitals
- The other two p orbitals can overlap sideways to form π orbitals
- The result is pi bonding (π) and pi antibonding (π*) orbitals
- p orbital filled with 2 electrons has no tendency of forming bonds
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ELECTRONIC EXCITATION
σ
π
n
σ*
π*
Relative Energy Diagram of σ,π, and n electrons
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ELECTRONIC EXCITATION
- Energy required to excite electrons from σ to σ* is very high(higher than those available in the UV region)
- UV radiation is however sufficient to excite electrons in π to π* and n to π* or σ* antibonding
- Molecular groups that absorb UV or VIS light are called chromophores
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ABSORPTION BY MOLECULES
- Review quantum mechanics (beyond the scope of this text)
- Quantum mechanical selection rules indicate that some transitions are allowed and some are forbidden
- Electrons move from highest occupied molecular orbital (HOMO) to the lowest unoccupied molecular orbital (LUMO)
during excitation
- LUMO is usually an antibonding orbital
- π electrons are excited to antibonding π* orbitals
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ABSORPTION BY MOLECULES
- n electrons are excited to either σ* or π* orbitals
π → π* Transition- A molecule must possess a chromophore with an unsaturated bond
(C=O, C=C, C=N, etc)
n → π * or n → σ* Transition- A molecule must contain atoms with nonbonding electrons
(N, O, S, X)
- Lists or organic compounds and their λmax are available (table 5.3)
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TRANSITION METAL COMPOUNDS
- Solutions are colored
- Absorb light in the visible portion of the spectrum
- Absorption is due to the presence of unfilled d orbitals
λmax is due to - The number of d electrons
- Geometry of compound- Atoms coordinated to the transition metal
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ABSORPTIVITY (a)
- Defines how much radiation will be absorbed by a molecule at a given concentration and wavelength
- Is termed molar absorptivity (ε) if concentration is expressed in molarity (M, mol/L)
- Can be calculated using Beer’s Law (A = abc = εbc)
- If units of b is cm and c is M then ε is M-1cm-1 or Lmol-1cm-1
- Magnitude of ε is an indication of the probability of the electronic transition
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ABSORPTIVITY (a)
- High ε results in strong absorption of light
- Low ε results in weak absorption of light
- ε is constant for a given wavelength but different at different wavelengths
- εmax implies ε at λmax (see table 5.4 for some εmax values)
- ε is 104 – 105 for allowed transitions and 10 – 100 for forbidden transitions
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UV ABSORPTION CURVES
- Broad absorption band is seen over a wide range of wavelengths
- Broad because each electronic energy level has multiple vibrational and rotational energy levels associated with it
- Each separate transition is quantized
- Vibrational energy levels are very close in energy
- Rotational energy levels are even closer
- These cause electronic transitions to appear as a broad band
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SOLVENTS
- Many absorbing molecules are usually dissolved in a solvent
- Solvent must be transparent over the wavelength range of interest
- Solute must completely dissolve in solvent
- Undissolved particles may scatter light which will affect quantitative analysis
- Solvent must be colorless
- Examples: acetone, water, toluene, hexane, chloroform
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INSTRUMENTATION
- Radiation source
- Monochromator
- Sample holder
- Detector
- Computer
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- Constant intensity over all wavelengths
- Produce light over a continuum of wavelengths
- Tungsten lamp and deuterium discharge lamp are the most common
RADIATION SOURCE
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Tungsten Filament Lamp
- Just like an ordinary electric light bulb- Contains tungsten filament that is heated electrically
- Glows at a temperature near 3000 K- Produces radiation at wavelengths from 320 to 2500 nm
- Stable, robust, and easy to use
- Modern lamps are tungsten-halogen lamps (has quartz bulb)
Disadvantage- Low radiation intensity at shorter wavelengths (< 350 nm)
RADIATION SOURCE
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Dueterium (D2) Arc Lamp
- Made of deuterium gas (D2) in a quartz bulb
- D2 molecules are electrically excited and dissociated
- Produces continuum radiation at λs from 160 to 400 nm
- Stable, robust, and widely used
- Emission intensity is 3x that of hydrogen at short λs
RADIATION SOURCE
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Xenon Arc Lamps
- Electric discharge lamps
- Xenon gas produces intense radiation over 200 – 1000 nm upon passage of current
- Produce very high radiation intensity
- Widely used in visible region and long λ end of UV
RADIATION SOURCE
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- Disperse radiation according to selected wavelengths
- Allow selected wavelengths to interact with the sample
- Diffraction gratings are used to disperse light in modern instruments
Refer chapter 2
MONOCHROMATORS
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- Earlier detectors were human eye observation of color and intensity
- Modern instruments make use of photoelectric transducers (detection devices that convert photons into electric signal)
Examples- Barrier layer cells
- Photomultiplier tubes- Semiconductor detectors
DETECTOR
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Barrier Layer Cells
- Also called photovoltaic cells
- A semiconductor (selenium) is joined to a strong metal base (iron)
- Silver is coated on the semiconductor
- Current is generated at the metal-semiconductor interface(requires no external electrical power)
- Response range is 350 nm – 750 nm
DETECTOR
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Photomultiplier Tubes
- The most common
- Photoemissive cathode is sealed in an evacuated transparent envelope
- Also contains anode and other electrodes called dynodes
- Electrons from cathode hit dynodes which causes more electrons to be emitted
- Process repeats until electrons fall on anode (collector)
DETECTOR
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Semiconductor Detectors
- Silicon and germanium are the most widely used elements
- Others include InP, GaAs, CdTe
- Covalently bonded solids with λ range ~ 190 nm – 1100 nm
Photodiode Array- Consists of a number of semiconductors embedded in a single
crystal in a linear array
- Used as detector for HPLC and CE
DETECTOR
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- Called sample cells or cuvettes or cuvets
- Different types of sample holders are designed for solids, liquids, and gases
- Cells must be transparent to UV radiation
- Quartz and fused silica are commonly used as materials
- Glass or plastic cells can be used for only VIS region
- Material must be chemically inert to solvents
SAMPLE HOLDER
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- HF and very strong bases should not be put in cells
- Standard cell is the 1 cm pathlength rectangular cell
- Holds about 3.5 mL sample
- Flow through cells are available (for chromatographic systems)
- Larger pathlength or volume cells are used for gases
- Thin solid films can be analyzed using a sliding film holder
SAMPLE HOLDER
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Fiber Optic Probes
- Enables spectrometer to be brought to sample for analysis
- Enables collection of spectrum from microliter samples
- Can collect spectrum from inside almost every container
- Useful for hazardous samples
SAMPLE HOLDER
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Chromophore- A group of atoms that gives rise to electronic absorption
Auxochrome- A substituent that contains unshared electron pairs (OH, NH, X)
- An auxochrome attached to a chromophore with π electronsshifts the λmax to longer wavelengths
ABSORPTION DEFINITIONS
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Bathochromic- A shift to longer wavelengths or red shift
Hypsochromic - A shift to shorter wavelengths or blue shift
Hyperchromism- An increase in intensity of an absorption band (increase in εmax)
Hypochromism- A decrease in intensity of an absorption band (decrease in εmax)
ABSORPTION DEFINITIONS
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- Molecules with absorption due to π → π* transition exhibit red shift when dissolved in polar solvents as compared to
nonpolar solvents
- Used to confirm the presence of π → π* transitions in molecules
- Molecules with absorption due to n → π* transition exhibit blue shift when dissolved in solvents that are able to form
hydrogen bonds (same with n → σ* transition)
- Used to confirm the presence of n electrons in a molecule
- Blue shift of n → σ* puts molecules into the vacuum UV region
SOLVENT EFFECTS
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- A compound that contains both π and n electrons may exhibit two absorption maxima with change in solvent polarity
- π → π* transitions absorb ~ 10x more strongly than n → π* transition
- n → π* transition occur at longer wavelengths than π → π*
- Such a compound will exhibit two characteristic peaks in a nonpolar solvent such as hexane
- The two peaks will be shifted closer to each other in a polar and hydrogen bonding solvent such as ethanol
SOLVENT EFFECTS
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ANALYSIS OF A MIXTURE
- Occurs when there is more than one absorbing species
- All absorbing species will contribute to absorbance at most λs
- Absorbance at a given λ = sum of absorbances from all species
AT = ε1b1c1 + ε2b2c2 + ε3b3c3 + ….
For the same sample cellb1 = b2 = b3 = b
AT = b(ε1c1 + ε2c2 + ε3c3 + ….)
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APPLICATIONS
- Environmental monitoring
- Industrial quality control or process control
- Pharmaceutical quality control
- For measuring kinetics of a chemical reaction
- For measuring the endpoint of spectrophotometric titrations
- For spectroelectrochemistry in which redox reactions are studied by measuring the electrochemistry and spectroscopy simultaneously
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OTHER TECHNIQUES
- Methods for nontransparent particles suspended in a liquid(colloidal suspensions, precipitates)
- Used for analyzing the clarity of drinking water, liquid medications, beverages
Nephelometry- Measures the amount of radiation scattered by the particles
Turbidimetry- Measures the amount of radiation not scattered by the particles
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LUMINESCENCE
- Molecular emission
- Includes any emission of radiation
Emission Intensity (I)
I = kPoc
k is a proportionality constantPo is the incident radiant power
c is the concentration of emitting species
- Only holds for low concentrations
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Photoluminescence (PL)- Excitation by absorption and re-radiation (very short lifetime)
- Examples are fluorescence and phosphorescence
Chemiluminescence (CL)- Excitation and emission of light as a result of a chemical reaction
Electrochemiluminescence (ECL)- Emission as a result of electrochemically generated species
Bioluminescence- Production and emission of light by a living organism
LUMINESCENCE
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LUMINESCENCE
Fluorescence
- Instantaneous emission of light following excitation
- Excitation by photon absorption to a vibrationally excited singlet state followed by relaxation resulting in emission of a photon
- Emitted photon has lower energy (longer λ) than absorbed energy(due to the radiationless loss)
- Called the stokes fluorescence (excited state lifetime ~ 1-20 ns)
- A molecule that exhibits fluorescence is called fluorophore
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LUMINESCENCE
Phosphorescence
- Similar to fluorescence
- Excited state lifetime is up to 10 s
- Excitation by absorption of light to an excited singlet state, thenan intersystem crossing (ISC) to the triplet state, followed
by emission of a photon
- Photon associated with phosphorescence has lower energy than fluorescence
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MOLECULAR EMISSION SPECTROSCOPY
- Two electrons occupying a given orbital have opposite spins
- There are two possible electronic transitions
- The excited state is known as a singlet state if one of the electrons goes to the excited state without changing its spin
- The excited state is known as a triplet state if one of the electrons goes to the excited state and changes its spin for both
to have same spin
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MOLECULAR EMISSION SPECTROSCOPY
- Singlet state energy levels (S) are higher than triplet state energy levels (T)
- Ground state is a singlet state (So)
- Excited state singlet can undergo radiationless transition to excited state triplet (ISC)
Transition from ground state singlet to excited state triplet is forbidden
Relative energy of transitionAbsorption > Fluorescence > Phosphorescence
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MOLECULAR EMISSION SPECTROSCOPY
Excited Singlet State (S1)
Ground Singlet State (So)
Excited Triplet State (T1)
Rel
ativ
e E
nerg
y
Ab
sorp
tion
Flu
ores
cen
ce
Phosphore
scence
ISC
Radiationless transition to the lowest vibrational level in the excited state
Intersystem crossing (radiationless)
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Sourceλ selector
sample
monochromator
(λ selector)readout detector
InstrumentationComponents of the Fluorometer
MOLECULAR EMISSION SPECTROSCOPY
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MOLECULAR EMISSION SPECTROSCOPY
Applications- Analysis of clinical samples, pharmaceuticals,
environmental samples, steroids
Advantages- High sensitivity and specificity
- Large linear range
Disadvantage- Quenching by impurities and solvent
- Temperature, viscosity and pH must be controlled to minimize quenching