The document discusses luminescence and phosphorescence spectroscopy. It defines luminescence as light emission from a substance when an electron returns to the ground state from an excited state. Phosphorescence is luminescence from a triplet excited state with a longer lifetime than fluorescence which occurs from a singlet state. The document describes various types of luminescence and provides details on instrumentation, sample preparation, and applications of phosphorescence spectroscopy in different fields such as pharmaceutical, clinical, environmental, and forensic analyses.

1. LUMINESENCE
The term 'luminescence' was introduced in 1888 by
Eilhard Wiedemann .
“Luminescence is the emission of light by a
substance. It occurs when an electron returns
to the electronic ground state from an excited
state and loses it's excess energy as a photon.”
Luminescence spectroscopy is a collective name
given to three related spectroscopic techniques.
They are;
 Molecular fluorescence spectroscopy
 Molecular phosphorescence spectroscopy
 Chemiluminescence spectroscopy

2. TYPES OF LUMINESENCE
 Bioluminescence
 Chemiluminescence
 Electrochemiluminescence
 Crystalloluminescence
 Electroluminescence
 Mechanoluminescence
 Triboluminescence
 Fractoluminescence
 Piezoluminescence
 Radioluminescence
 Sonoluminescence
 Thermoluminescence

3. PHOTOLUMINESENCE
“Photoluminescence (PL) is the spontaneous
emission of light from a material under optical
excitation.”
It is futher subdivided into two types
a. Flourescence
b. Phosphorescence

4. ACTIVATION AND DEACTIVATION IN
PHOSPHORESENCE
 The absorption of a photon of suitable energy
causes the molecule to get excited from the ground
state to one of the excited states. This process is
called as excitation or activation and is governed
by Franck-Condon principle.
 According to this principle, the electronic transition
takes place so fast (~10-15 s) that the molecule
does not get an opportunity to execute a vibration,
 i.e., when the electrons are excited the internuclear
distance does not change.
• The basis for the principle is that the nuclei are
very massive as compared to the electrons and
therefore move very slowly.

5. • At the ground state , the molecular orbitals are occupied
by two electrons. the spins of the two electrons in the
same orbital must be antiparallel. This implies that the
total spin, S, of the molecule in the ground state is
zero [½ + ( ½)].
• This energy state is called “singlet state” and is labeled
as S0.
• The electron spins in the excited state achieved by
absorption of radiation may either be parallel or
antiparallel. Accordingly, this may be a triplet (parallel)
or a singlet (antiparallel) state.

6.  The deactivation processes can be broadly categorized
into two groups given below.
• Nonradiative deactivation
 Vibrational Relaxation
 Internal Conversion
 Intersystem Crossing
• Radiative deactivation
 Fluorescence
 Phosphorescence

7. JABLONSKI DIAGRAM

8. DIFFERENCE BETWEEN PHOSPHORESCENCE
AND FLUORESCENCE
Phosphorescence Fluorescence
 The emission could  The emission could
proceed either from a proceed only from a
singlet or triplet state. singlet state.
 short-live electrons  longer lifetime of the
(<10-5 s) in the excited excited state (second
state of fluorescence to minutes)

9. FLUORESCENCE AND
PHOSPHORESCENCE
 Fluorescence  Phosphorescence

10. PHOSPHORESCENCE SPECTROSCOPY
Phosphorescence Spectroscopy is the
spectroscopic study of the radiation
emitted by the lifetime of
phosphorescence.
Phosphorescence has been observed
from a wide variety of compounds and is
differentiated from fluorescence by the
long-lived emission of light after
extinction of the excitation source. The
first analytical uses of phosphorescence
were published in 1957 by Kiers et. al.

11. FIG:SRAL2O4 ,PIGMENTS IN THE DARK AND THAN PIGMENTS IN
THE DARK AFTER 4 MIN

12. PHOSPHORIMETRY
 Spectrophosphorimeter is similar to a
Spectrofluorimeter except that the former
instrument must be fitted with
1) a sample system which is maintained at liquid
nitrogen temperature
2) A Rotating-shutter device commonly called a
phosphoroscope and
.

13. SAMPLE PREPARATION
 The majority of phosphorescence measurements are carried out in
rigid media at the temperature of liquid nitrogen
 The criteria for solvent selection are:
 good solubility of the analyte
 formation of a clear rigid glass at 77 K
 low phosphorescence background (high purity)
 For polar compounds, ethanol is an excellent solvent and small
 For non-polar compounds, the most popular solvent is a mixture of
diethyl ether, isopentane and ethanol in the ratio of 5:5:2 respectively
 The samples are placed in a narrow quartz tube(internal diameter
varying from 1 to 3 mm).
 The dimensions are a compromise between too small a diameter
and too large a diameter.
 The sample tubes are placed in liquid nitrogen held in a quartz
Dewar flask, and the latter placed in the sample holder known as a
phosphoroscope.

14. Phosphorescence can also be observed from
solid samples at room temperature, and the
compounds can be divided into two types.
 The first includes inorganic salts and oxides such
as the rare earths, europium and uranium
 The second type of compounds are those which
exhibit phosphorescence when absorbed onto
certain substrates such as
paper, cellulose, silica, etc.
 Polar organic molecules, when absorbed onto
filter paper from solutions containing 1 N
NaOH and thoroughly dried, exhibit
phosphorescence.
 Studies have shown that the phosphorescence
could be enhanced by the addition of heavy
atoms, for example iodine, silver, lead

15. PHOSPHOROSCOPE

16. INSTRUMENTATION
 Excitation Source
 Filters/Monochromators for excitation radiations
 Phosphorscope(sample Cell)
 Filters and Monochromators for emission
 Detectors

17. EXCITATION SOURCE
 High intensity source of UV light are used
 Lasers – A laser makes it possible to have narrow wavelength intervals
that offer very high energy irradiation. This is useful when a large amount
of energy is needed to produce the Phosphorescence in the sample.
 Photodiodes – Photodiodes are specialized diodes that can be
configured in a manner that allows electrons to flow towards the sample
so that the excess energy excites the phosphorescent particles.
 Xenon Arcs – Arcs of Xenon can produce the right amount of radiation for
Phosphorescent materials.
 Mercury Vapor – Since mercury vapor can create ultraviolet radiation
when electrical current is passed through it, it is good for use with
materials that shows Phosphorescence under the ultraviolet radiation.
Care should be taken as Intense UV light is hazardous for health

18.  Filters and monochromators used in a
phosphorimeter device.
 Filters
 Absorption
 Interfernce
 Monochromators allow wavelength adjustment.
Monochromators make it possible to do so with a
diffraction grating.
 The primary filters that excite the sample provide the appropriate wavelength
 while the secondary filters monochromate the emitted light when sent to the
detector.
 Phosphoresence spectroscopy detectors may have
a
 single channel (single wavelength from sample)
 or multiple channels (multiple wavelengths detection)

19. PHOSPHOROSCOPE
1.The Becquerel or rotating disc phosphoroscope
 A rotating disk excitation optical chopper, with three open
and three larger opaque areas, is used to alternately
excite the sample and allow phosphorescence to be
measured.
 By measuring the phosphorescence intensity at several
time intervals along the emission decay curve, a recorder
trace of the decay with respect to time can be produced.
 The analytical precision and accuracy for quantitative
measurements is improved by rotating the sample tube.
This minimizes variation in the signal due to sample
inhomogeniety resulting from imperfect glass formation at
lowtemperatures .

20. 2.The Rotating-Can Phosphoroscope:
 It consists of hollow cylinder having one or more slit which are
equally spaced in the circumference.
 This is rotated by a variable-speed motor(>1000rpm)
 when the rotating-can is rotated by a motor the sample is excited
show fluorescence and phosphorescence
 For Fluorescence The emission monochromator is blocked by the
can so that fluorescence and scattered radiation cannot be detected.
 As the can rotates, the excitation beam is blocked and the
fluorescence and scattered radiation decay to a negligible amount.
 Further rotation of the can will bring the window into alignment with
the emission monochromator entrance slit and phosphorescence
radiation will pass into the emission monochromator and onto the
sample photomultiplier

22. PULSED SOURCE-TIME
RESOLVED PHOSPHORIMETRY
 was first proposed by Fisher and Winefordner in 1972.
 A pulsed source produces higher peak intensities than a
continuously operated xenon lamp resulting in a greater peak
phosphorescence emission intensity.
 Pulsed source phosphorimetry has the advantage of time
resolution compared with a mechanically modulated system
permitting the analysis of organic phosphor with short lifetimes
(0.1 to 50 microsec)
 The xenon lamp produces a burst of energy with a width a half
peak intensity of less than 10 μsec.
 The signals from the sample photomultiplier are gated and
both the
 delay of the start of the gate after the start the flash
 Timing is performed by a crystal clock and is highly accurate.
 a quantum-corrected reference photomultiplier is used to
monitor the flash intensity and the signals from th sample and
reference

23. FIG:SCHEMATIC DRAWING OF GATED
PHOSPHORIMETER SETUP.

24. THE PERKIN ELMER LS 55 LUMINESCENCE
SPECTROMETER
 It is a very versatile
instrument that allows
measurement of
fluorescence,
phosphorescence ,
chemiluminescence and
bioluminescence of a liquid,
solid, powder, or thin film
sample
 Fluorescence data are
collected at the instant of the
flash while
phosphorescence data are
collected in the dark period
between each flash.

25. APPLICATIONS OF PHOSPHORESCENCE
SPECTROSCOPY
 Pharmaceutical Applications
 Clinical Applications
 Environmental Applications
 Forensic Applications
 Entertainment Applications

26. PHARMACEUTICAL APPLICATIONS
 The majority of phosphorescence applications have
been applied in the drug and pharmaceutical field
and in the analysis of pesticides
 A number of the sulphonamide class of drugs
exhibit phosphorescence as do
phenobarbital, cocaine, procaine, chlorpromazin
and salicylic acid.

27. CLINICAL APPLICATIONS
 The phosphorescence intensity of the rare earths
increases tremendously when they are covalently
bound to certain molecules and this feature has
been used in the analysis of transferin in blood
 dual-wavelength phosphorimeter used to measure
microvascular PO2 (µPO2) in different depths in
tissue and demonstrates its use in rat kidney.

29. Entertainment Applications
 use of phosphor coated stamps and envelopes has
appeared which gives a fascinating insight into the
practical use of phosphorescence
 The rare earths and uranyl elements phosphoresce and a
number of them, particularly europium and terbium, are
used as phosphors in lamps and TV tubes.

30. ENVIRONMENTAL APPLICATIONS
 Phosphorescence has been used in the detection
of air and water-borne pollutants
 for the analysis of impurities in polycyclic aromatic
hydrocarbons and in petroleum products .

31. CONCLUSION
Although the applications of phosphorescence have
been somewhat limited in the past, the introduction
of new instrumentation and advances made in room
temperature phosphorescence have lead to an
increase in its use, particularly in clinical chemistry ,
the forensic, environmental and pharmaceutical
fields.

32. REFERENCES
1.Kiers, R.J., Britt, R.D., Wentworth, W.E., Anal. Chem., 29, 202
(1957).
2. Fluorescence Detection in Liquid Chromatography., Rhys
Williams, A.T., PerkinElmer Limited (1980).
3. Schulman, E. M., Walling, C., J. Phys. Chem., 77, 902 (1973).
4. Hollifield, H. C., Winefordner, J. D., Anal. Chem., 40, 1759
(1968).
5. Zweidinger, R., Winefordner, J. D., Anal. Chem., 42, 639 (1970).
6. Fisher, R. P., Winefordner, J. D., Anal. Chem., 44, 948 (1972).
7. Harbaugh, K.F., O’Donnell, C. M., Winefordner, J. D., Anal.
Chem., 45, 381 (1973.)
8. Sawicki, E., Pjaff, J. D., Anal. Chim. Acta., 32, 521 (1965).
9. Giffard, L. A., Miller, J. N., Burns, D. T., Bridges, J. W., J.
Chromatog., 103, 15(1975).
10. de Lima, C. G., Nichola, E. M., Anal.Chem., 50, 1658 (1978).
11. O’Donnell, C. M., Winefordner, J. D., Clin. Chem., 21, 285
(1975).

33. 12. Aaron, J. J., Winefordner, J. D., Anal.Chem., 44, 2127
(1972).
13. Vo. Dinh, T. Winefordner, J. D., Appl. Spec. Rev., 13, 261
(1977).
14. Saunders, L. B., Winefordner, J. D., Talanta, 16 407 (1969).
15. Aaron, J. J. Winefordner, J. D., Anal. Chem., 19, 21 (1972)
16. Hollifield, H. C., Winefordner, J. D., Talanta, 12, 860 (1965).
17. Winefordner, J. D., Latz, H. W., Anal. Chem., 35, 1517
(1963).
18. Perry, A. W., Winefordner, J. D., Anal. Chem., 43, 781
(1971).
19. Aaron, J. J., Kaleel, E. M., Winefordner, J. D., J. Agric. Food
Chem., 27, 1233 (1979).

34. THANKS