LUMINESENCEThe 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

TYPES OF LUMINESENCE Bioluminescence Chemiluminescence

Electrochemiluminescence Crystalloluminescence Electroluminescence Mechanoluminescence

Triboluminescence Fractoluminescence Piezoluminescence

Radioluminescence Sonoluminescence Thermoluminescence

PHOTOLUMINESENCE “Photoluminescence (PL) is the spontaneous

emission of light from a material under optical excitation.”

It is futher subdivided into two typesa. Flourescenceb. Phosphorescence

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.

• 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.

The deactivation processes can be broadly categorized into two groups given below.

• Nonradiative deactivation Vibrational Relaxation Internal Conversion Intersystem Crossing

• Radiative deactivation Fluorescence Phosphorescence

JABLONSKI DIAGRAM

DIFFERENCE BETWEEN PHOSPHORESCENCEAND FLUORESCENCE

The emission could proceed either from a singlet or triplet state.

short-live electrons (<10-5 s) in the excited state of fluorescence

The emission could proceed only from a singlet state.

longer lifetime of the excited state (second to minutes)

PhosphorescenceFluorescence

FLUORESCENCE AND PHOSPHORESCENCE

Fluorescence Phosphorescence

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.

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

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 temperature2) A Rotating-shutter device commonly called a phosphoroscope and

.

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.

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

PHOSPHOROSCOPE

INSTRUMENTATION Excitation Source Filters/Monochromators for excitation

radiations Phosphorscope(sample Cell) Filters and Monochromators for emission Detectors

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

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)

PHOSPHOROSCOPE1.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 .

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

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

FIG:SCHEMATIC DRAWING OF GATED PHOSPHORIMETER SETUP.

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.

APPLICATIONS OF PHOSPHORESCENCE SPECTROSCOPY

Pharmaceutical Applications

Clinical Applications

Environmental Applications

Forensic Applications

Entertainment Applications

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.

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.

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.

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 .

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.

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).

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

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THANKS