1.1 Range of molar absorptivity

Electronic excitation of outer valence (i.e. bonding) electron

How probable for this electronic excitation? (allowed transition, or forbidden)

judged by the range of = 104 -105 L mol-1 cm-1, strong absorption < 103 L mol-1 cm-1, low intensity

*MhM excitation

1.2 Which electron get excited?1.2.1 Organic molecules

, (bonding) and n (non-bonding) orbitals*, * (anti-bonding) orbitals

* E large ( < 150 nm, out of range) = 10 -10,000 Lmol-1cm-1

n * E smaller ( = 150 - 250 nm) = 200-2000 Lmol-1cm-1

* n * E smallest ( = 200 - 700 nm) = 10-10,000 Lmol-1cm-1

Ideal for UV-Vis spectrometry of organic chromophore

1.2.2 Inorganic moleculesMost transition metal ions are colored (absorption in Vis) due to d d electronic transition

Fig. 14-4 (p.371)Fig. 14-3 (p.370)

1.2.3 Charge-transfer absorption

A: electron donor, metal ionsD: electron acceptor, ligand > 10,000

DAhAD excitation

Fig. 14-5 (p.371)

2.1Approximation of T and A

A = -log T = log (P0/P) = ··b·c: molar absorptivity at one particular

wavelength (L·mol-1cm-1)b: path length of absorption (cm)c: molar concentration (mol·L-1)

Fig. 6-25 (p.158)

Fig. 13-1 (p.337)

Light loss due to reflection (17.3%), scattering, …

PP

PPA

PP

PPT

solution

solvent

solvent

solution

0

0

loglog

2.2 Application of Beer’s law to mixtures

Absorbance is additive

Atotal = A1 + A2 + … = 1bc1 + 2bc2

For a 2-component mixture, we measure the absorption at two different wavelength, respectively

A1 = 1,1·b·c1 + 2,1·b·c2

A2 = 1,2·b·c1 + 2,2·b·c2

2.3 Limitations of Beer’s law

2.3.1 Real deviations

At low concentration

A = -log T = log (P0/P) = ··b·c

At c > 0.01 M solute-solute interaction, hydrogen-bond, …can alter the electronic absorption at a given wavelength

dilute the solution

2.3.2 Chemical effects analyte associates, dissociates or reacts with a solvent to

give molecule with different

Example: acid-base equilibrium of an indicator

430 570 HIn 6.30 x102 7.12x103 (measured in HCl solution)In- 2.06 x 1049.61 x102 (measured in NaOH solution)

What’s the absorbance of unbuffered solution at c = 2 x 10-5M?

073.0][][

236.0][][

1088.0][

1012.1][

570,570,570

430,430,430

5

5

HInbInbA

HInbInbA

MHIn

MIn

HInIn

HInIn

5 a 102.1K InHHIn

][][

][][

1042.1][

]][[ 5

IncHIn

InHHInInHKa

Fig. 13-3 (p.340)=[HIn] + [In-]

InHHIn

2.3.3 Instrumental deviations due to polychromatic radiation

Beer’s law applies for monochromatic absorption only.

If a band of radiation consisting of two wavelength ’, and ”Assuming Beer’s law applies to each wavelength

bc"bcm

bc"

bc

PPPP

PPPP

A

PP"

PP

bcPP

A

"0

''0

"0

'0

"0

'0

"0

''0

'0

1010log

"'log

absorbance Measured

10"h wavelengtsecond For the

10'

''

log'

length first waveFor

Fig. 7-11 (p.176)

2.3.3 Instrumental deviations due to polychromatic radiation

bc"bcm PPPP

PPPPA "

0'

0

"0

'0

"0

'0

1010log

"'log

absorbance Measured

Fig. 13-4 (p.341)

′″

′″

′″ Non-linear calibration curve

How to avoid :

Select a wavelength band near its maximum absorption where the absorptivity changes little with wavelength

Fig. 13-5 (p.341)

2.3.4 Other physical effects

stray light – the scattering, reflection radiation from the instrument, outside the nominal wavelength band chosen for measurement

mismatched cell for the sample and the blank

3.1 Standard deviation of c

TTs

css

sTcbTT

c

Tb

c

Tc

TT

cc

Tc

log434.0

)(

434.0

log1

2

2

222

3.2 Sources of instrumental noise Case I Limited readout resolution (31/2-digit displays 0.1% uncertainty from 0%T -100% T) Thermal noise in thermal detector, etc (particularly for IR and neat IR spectrophotometer)

Case IIShot noise in photon detector (random emission of photon from the light source orrandom emission of electrons from the cathode in a detector)

Case III Flicker noise,

Fail to position sample and blank cells reproducibly in replicate measurements (as a result, different sections of cell window are exposed to radiation, and reflection and scattering losses change)

1ksT

TTksT 22

TksT 3

3.2 Sources of instrumental noise

Fig. 13-3 (p.344)

4.1 Designsa. Single beam

b. Double-beam-in-space

c. Double-beam-in-time

Advantage of double beam configuration

• Compensate for fluctuation in the radiant output, drift in transducer, etc.

• Continuous recording of spectra

Fig. 13-13 (p.352)

Shimadzu UV-2450 Spectrophotometer

Wavelength Range

190 to 900nm (performance guaranteed range). Extendable to 1,100nm through the use of the optional photomultiplier. (The measurable range maybe restricted in the shorter wavelength side depending on the type of photomultipler used.)

Monochromator System

UV-2450: Single monochromator with a high-performance blazed holographic grating in the aberration corrected Czerny-Turner mounting.

Resolution 0.1nmSpectral Bandwidth 0.1, 0.2, 0.5, 1, 2 and 5nm

Wavelength Accuracy ±0.3nm

Wavelength Repeatability ±0.01nm

Wavelength Scanning Speed FAST, MEDIUM, SLOW, and SUPER SLOW

Light Source 50W halogen lamp (2,000 hours of life) and D2lamp (500 hours of life)Light Source lamp switching Selectable between 282nm and 393nm

Stray Light UV-2450: Less than 0.015% at 220nm and 340nmDetector Photomultiplier R-928Photometric System Double beam, direct ratio system with dynode feedback

Photometric Mode Absorbance (Abs.), transmittance (%), reflectance (%) and energy (E).

Photometric Range

Abosrbance: -4~5 Abs. (0.001 Abs. increments)Transmittance: 0~999.9% (0.01 increments)Reflectance: 0~999.9% (0.01% increments)

Photometric Accuracy

±0.002Abs(0~0.5Abs), ±0.004Abs(0.5~1Abs),±0.3T (0~100%T) (all determined with NIST 930D standard filter)

Photometric Repeatability 0.001Abs (0~0.5Abs), ±0.1%T

Baseline Correction Selectable with storage in firmware

Baseline Flatness Within ±0.001Abs. (excluding noise, 2nm slit width and SLOW wavelength scanning speed)

Drift Less than 0.0004 Abs. per hour (after 2 hours warm-up)Dimensions 570 (W) x 660 (D) x 275 (H) (mm)Weight 36kgPower Supply AC 100V/120V/220V/240V, 50/60Hz 250VA (swithc-selectable)

Shimadzu UV-2450 Spectrophotometer

d. Effects of monchromator exit slit width on spectraNarrow exit slit width improves the spectrum resolutionbut it also significantly reduce the radiant power

Trade-off between resolution and S/N ratio

e. Multichannel spectrometerNo monochromator, but disperses transmitted light and measures “all wavelength at once”

No Scanning-simple and fastMore expensiveLimited resolution

Fig. 13-15 (p.353)

5.1Quanlitative spectra

Solvent effects on the UV-Vis spectra

Polar solvents “blur” vibrational features

Polar solvents shift absorption maximan * blue shift * red shift -

UV-Vis not reliable for qualitative but excellent for quantitative analysis

5.2 Quantitative analysis- Determining the relationship between A and c

External StandardsStandard-Addition

Fig. 14-14 (p.382)

5.3 Spectrophotometric kinetics

Fig. 14-16 (p.384)