Molecular Luminescence
Emission of a photon as an excited state molecule
returns to a lower state
• Chemoluminescence
• Bioluminescence
• Crystalloluminescence
• Electroluminescence
• Photoluminescence
• Radioluminescence
• Sonoluminescence
• Thermoluminescence
• Triboluminescence
http://www.shef.ac.uk/content/1/c6/01/89/68/luminescence.jpg
Jablonski Diagram
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Absorption
Selection Rules:
DJ = 0, 1
Dv = 1, 2, 3, …
DS = 0 (i.e. S  S, T  T)
Very Fast  10-14 – 10-15 sec.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Vibrational Relaxation
• Excited molecule rapidly
transfers excess vibrational
energy to the solvent /
medium through collisions.
• Molecule quickly relaxes
into the ground vibrational
level in the excited
electronic level.
• Non-radiative process
• 10-11 – 10-10 sec.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Internal Conversion
• Transfers into a lower
energy electronic state of
the same multiplicity
without emission of a
photon.
• Favored when there is an
overlap of the electronic
states’ potential energy
curves.
• Non-radiative process
(minimal energy change)
• ~10-12 s between excited
electronic states.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Fluorescence
• Radiative transition
between electronic states
with the same multiplicity.
• Almost always a
progression from the
ground vibrational level of
the 1st excited electronic
state.
• 10-10 – 10-6 sec.
• Occurs at a lower energy
than excitation.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Stokes Shift
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Franck-Condon Factor
Relationship
between the shape
of the excitation and
fluorescence bands.
shift
P.R. Callis et. al., Chem. Phys. Lett, 244 (1995), 53-58.
Ingle and Crouch,
Spectrochemical Analysis
External Conversion
• Non-radiative transition
between electronic states
involving transfer of energy
to other species (solvent,
solutes).
• Also referred to as
quenching.
• Modifying conditions to
reduce collisions reduces
the rate of external
conversion.
• Occurs on a comparable
time scale as fluorescence.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Intersystem Crossing
• Similar to internal
conversion except that it
occurs between electronic
states with different
multiplicities.
• Slower than internal
conversion.
• More likely in molecules
containing heavy nuclei.
• More likely in the
presence of paramagnetic
compounds.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Luminol Chemoluminescence
www.wikipedia.org
Phosphorescence
• Radiative transition between electronic states of
different multiplicities.
• Much slower than fluorescence (10-4 – 104 s).
• Even lower energy than fluorescence.
www.wikipedia.org
Stokes Shift
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Dissociation
Ingle and Crouch, Spectrochemical Analysis
Predissociation
• Occurs if the molecule
enters a vibrational level
above the dissociation limit
during an internal
conversion.
• Dissociation and
predissociation are more
likely in molecules that
absorb at low l.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Quantum Yield
Fraction of absorbed photons that are
converted to luminescence, fluorescence
or phosphorescence photons.
fL =
F L,p
F A,p
May approach unity in favorable cases.
Fluorescence Quantum Yield
All activation and deactivation processes discussed
so far can be described using first order rate
constants.
dnS1
dt
= k A nS0 - (k F + k nr )nS1
nS1, nS0 = population densities of S1 and S0.
kA = rate of absorption
kF = rate of fluorescence
knr = rate of non-radiative deactivation processes.
A continuously illuminated sample volume (V) will
reach steady-state.
dnS1
dt
= k A nS0 - (k F + k nr )nS1 = 0
nS1 =
nS 0 k A
k F + k nr
FA,p = kAnS0V
FF,p = kFnS1V
unitless but
Fluorescence
FF,p
k F describes
=
Quantum Efficiency fF =
F A,p k F + k nr photons/molecule
of a Molecule:
fF =
typically ~ 106 – 109 s-1
k F + k ec
kF
+ k ic + k isc + k pd + k d
kec = external conversion (S1  S0)
kic = internal conversion (S1  S0) typically 105-107 s-1
kisc = intersystem crossing (S1  T1) typically 106-109 s-1
kpd = predissociation
kd = dissociation
Time Scales of Processes
http://micro.magnet.fsu.edu/primer/techniques/fluorescence/fluorescenceintro.html
Can put in terms of nS0:
FF,p = nS0kAfFV
Proportional to the number of fluorophores, the
rate of absorption (i.e. e), the quantum yield
and the volume of the sample measured.
Are you getting the concept?
For a given fluorophore under steady state conditions,
excitation of a 1 cm3 sample volume yields the following
first-order rate constants: kf = 5 x 107 s-1, knr = 9 x 105 s-1,
and ka = 1 x 1014 s-1 and an overall rate of fluorescence
photon emission of 9.8 x 1019 photons/second. What is
the molecule number density in the ground electronic
state?
Phosphorescence Quantum Yield
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Phosphorescence Quantum Yield
Product of two factors:
- fraction of absorbed photons that undergo
intersystem crossing.
- fraction of molecules in T1 that phosphoresce.
æ k isc
fP = çç
è k F + k nr
öæ k P
÷÷çç
øè k P + k'nr
ö
÷÷
ø
knr = non-radiative deactivation of S1.
k’nr = non-radiative deactivation of T1.
Is phosphorescence possible if kP < kF?
Conditions for Phosphorescence
kisc > kF + kec + kic + kpd + kd
kP > k’nr
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Luminescence Lifetimes
Emitted Luminescence will decay with time according to:
F L (t )  F0L e

t
L
Φ L (t) luminescence radiant power at time t
Φ
0
L
τL
luminescence radiant power at time 0
luminescence lifetime
 F  (k F  knr )
1
 P  (k P  k 'nr ) 1
~10-5 – 10-8 s
~10-4 – 10 s
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Quenching
Static Quenching
Lumophore in ground state and quencher form dark
complex. Luminescence is only observed from
unbound lumophore. Luminescence lifetime not
affected by static quenching.
Dynamic Quenching/Collisional Quenching
Requires contact between quencher and excited
lumophore during collision (temperature and viscosity
dependent). Luminescence lifetime drops with
increasing quencher concentration.
Long-Range Quenching/Förster Quenching
Result of dipole-dipole coupling between donor
(lumophore) and acceptor (quencher). Rate of energy
transfer drops with R-6. Used to assess distances in
proteins (good for 2-10 nm).
Fluorescence Resonance Energy Transfer (FRET)
http://www.olympusfluoview.com/applications/fretintro.html
Are you getting the concept?
Determine the type of quenching being demonstrated
in the figures below if the fluorescence lifetime of
receptor 1 is unchanged with increasing addition of 3.
S. Amemiya et al., Chem. Commun.,1997, 1027.
Fluorescence or Phosphorescence?
p – p* transitions are most favorable for fluorescence.
 e is high (100 – 1000 times greater than n – p*)
 kF is also high (absorption and spontaneous
emission are related).
 Fluorescence lifetime is short (10-7 – 10-9 s for
p – p* vs. 10-5 – 10-7 s for n – p*).
Nonaromatic Unsaturated Hydrocarbons
Luminescence is rare in nonaromatic hydrocarbons.
Possible if highly
conjugated due to
p – p* transitions.
Seyhan Ege, Organic Chemistry, D.C. Heath
and Company, Lexington, MA, 1989.
Aromatic Hydrocarbons
Fluorescent
Low lying p – p* singlet state
Phosphorescence is weak
because there are no n electrons
Ingle and Crouch, Spectrochemical Analysis
Heterocyclic Aromatics
Aromatics containing carbonyl or
heteroatoms are more likely to phosphoresce
n – p* promotes intersystem crossing.
Fluorescence is often weaker.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Aromatic Substituents
• Electron donating groups usually increase fF.
• Electron withdrawing groups usually decrease fF.
Ingle and Crouch, Spectrochemical Analysis
Halogen Substituents
Internal Heavy Atom Effect
Promotes intersystem crossing.
fF decreases as MW increases.
fP increases as MW increases.
P decreases as MW increases.
Ingle and Crouch, Spectrochemical Analysis
Increased Conjugation
fF increases as conjugation increases.
fP decreases as conjugation increases.
Hypsochromic effect and bathochromic shift.
Ingle and Crouch, Spectrochemical Analysis
Rigid Planar Structure
fF = 1.0
fF = 0.2
fF = 0.8
not fluorescent
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Ingle and Crouch,
Spectrochemical Analysis
Metals
Metals other than certain lanthanides and actinides
(with f-f transitions) are usually not themselves
fluorescent.
A number of organometallic complexes are fluorescent.
Skoog, Hollar, Nieman, Principles of Instrumental Analysis,
Saunders College Publishing, Philadelphia, 1998.
Fluorescence and Phosphorescence
Which effect is used more regularly?
SciFinder Scholar Citations
Fluorescence
Phosphorescence
… Labels/Tags
… Dyes
1729
25
10612
56
www.wikipedia.org
Fluorescence or Phosphorescence?
Commercially Available Phosphorescence Labels
Erythrosin derivative
Eosin derivative
http://www.invitrogen.com/
Fluorescence or Phosphorescence?
Publications in Analytical Chemistry
Fluorescence …
Phosphorescence…
1380
106
• Phosphorescence is rarer than fluorescence => Higher selectivity.
• Phosphorescence: Analysis of aromatic compounds in
environmental samples.