Chem 222

#15 Ch18 & 21Mar 1, 2005

Announcement

• Before the spring break, you have two weekends to write up a report.

• Reports for 4 exps are due right after the spring break.

• After the spring break, you have only one weekend, and so you must be busy.

Finish writing reports for KH 3-14 & Exp 19 & 20 early.

Homework

• Review Ch 16-1, 16-4• Problems 16-1, 16-3, 16-5, 16-14Ch 18 Problems:18-10, 18-12, 18-16, 18-22

21-B, 21-1, 21-10, 21-12, 21-18• Read Ch 10

Questions• 18-4 Which molecular process

corresponds to the energies of microwave, infrared, visible, and ultraviolet of photons

• 18-8 Why does a compound whose visible absorption is at 480 nm (blue-green) appear to be red?

Combined electronic vibrational, rotational transitions

What happens to absorbed energy

Florescence Phosphorescence

Life time 10-4 to 10-2 s 10-8 to 10-4 s

Why is fluorescence spectroscopy more sensitive than absorbance ?

<

(Emission)Absorbance(Excitation)

λEX > λEM or λEX < λEM ?

18-6 Luminescence

• Fluorescence and phosphorescence are examples of luminescence, which is emission of light from any excited state of molecule.

Essentials of a luminescence experiment

λex

λem

Memorize this diagram

Comparison between absorbance and fluorescence

• P0 Sample P

If concentration is low, let’s say P0 ~ 0.99 P We have to detect 1 % differencefrom high background

P0 Sample

0.01P

λex

λem

Low background because λex ≠ λem

I = kcP0

Fluorescence Imaging

GFP (Green Fluorescence Protein)isolated from jelly fish exhibits fluorescence

http://deepgreen.stanford.edu/Immuno-fluorescence microscopy

Molecular Beacon

Detection limit 1 nM

Besides calculations• Q1. Draw a diagram of an

luminescence spectrophotometer

• Q2. What is the difference Between absorption and fluorescence spectrophotometer

• Spelling ? Monochromatorcuvet

A typical questions for quiz

Excitation and Emission Spectra

Ene

rgy

Abs

Ene

rgy

Em

Why emission spectrum is the mirror image of absorption spectrum?

Excitation Excitation

Immunoassays (Ch 19-5)

Ch 21 Atomic Spectroscopy

• 21-1 Overview2000-3000 K

Why do we need a laser ?

Memorize the diagrams

λ

Atomic Emission

Fe + hot plasma Fe* Fe + hν

Atomic Emission Spectrum

Multi-element Analysis

Excited State

Extremely Narrow Line0.001 nm Resolution

Comparison of tooth by Atomic spectroscopy (ICP-mass; see 21-6)

Applications Material ScienceTrace Metal in BiologyWaste Control (Metals)

Analysis of Minerals in Environment

Modern tooth from Poland

Tooth from Norway AD 1800

Measurements of Lead (Pb) in Sugar by AA

Pb (ppb)

http://www.shimadzu.com/apps/appnotes/c122-e047.pdf

Concentration (ppb)

Abs

orba

nce

Q. How much is ppb? Pb/solution = 10-[Q1]

Blood lead levels as low as 0.1 µg per mLcan impair mental and physical development

21-2 Atomization: Flames, Furnaces, and Plasmas

• Most flame spectrometer uses a premix burner.

Fuel, Oxidant, Sample are mixed before introduction to the flame

Break droplets intosmaller particles

2400 – 2700 K ( acetylene + air)

Flame

2-1 Furnaces Usable instead of flame

• An electrically heated graphite furnace offers greater sensitivity than that afforded by flames and requires less sample.

Will be heated up to 2100 ºC

•90 ºC (Dry sample)• 125 ºC 20 s (Remove Solvent)•1400 ºC 60 s (Charring:

Remove Organic)• 2100 ºC 10s (Fe atomized)

Measurement of Fe in protein

21-3 How temperature affects atomic spectroscopyBolzmann distributionN*/N0 = (g*/g) exp(-∆E/kT)

g: number of states∆E = E* - EN and N* population of energy states E and

E*

E* __ ___ __ g* =3 E* __ __ g* =2N* N*

E ___ ___ g =2 __ __ __ g=3N N

∆E = 0.5kTIf ∆E = kT N*/N = (3/2)exp(-1)If ∆E = 2kT (3/2)exp(-[Q1])

The effect of temperature on excited-

state population

The lowest excited state of a sodium atom lies 3.371 ×10-19 J/atom above the ground state.

g* =2 and g =1.

Estimate the fraction of sodium atoms in the excited state at 2600 K.

(N*/N) =[Q1]×exp[-[Q2]/(2600×1.381×10-23)]

= 1.67 × 10-4 only 0.02 %

What happens if you increase Tto 6000 K?

At 2610 K N*/N = 1.74 × 10-4

The effects of temperature on absorption and emission

∆E = hc/λ Higher Fraction

Inductivity coupled plasma

6,000-10,000 K

Argon gas In

27 MHz RF Accelerate e- in Ar

2-4 Background correction

Background0.3