Fluorescence

Topics Definition Theory’s Factor’s Instrumentation Applications

Luminescence Emission of previously absorbed radiation

Types : Luminescence is two types 1. Florescence 2.Phosphorescence

Fluorescence When a beam of light is incident on certain substances, they emmit visible light or radiations. ( The substance is fluorescent substance. Measurement is called as fluorimetry. )

Phosphorescence When a beam of light is incident on certain substances, they emit radiations continuously even after the incident light is cut off . (The substance is phosphorescent. Measurement of radiations is Phosphorimetry. )

Types of fluorescence 1. Based upon the wave length of emitted radiation when compared to absorbed radiation - Stokes's fluorescence - Anti- Stokes's fluorescence - Resonance fluorescence

2. Based upon the phenomenon - Sensitized fluorescence - Direct line fluorescence - Stepwise fluorescence - Thermally assisted fluorescence

Theory 1.Singlet and triplet states 2.Excited-state state process in molecules 3.Relation between fluorescence intensity and concentration 4.Factors affecting fluorescence

Singlet and triplet states A molecular electronic state in which all of the electron spins are paired is called a singlet state. (spine state) If ‘no’ un pair electrons are present (n=0), there is only n+1 or 0+1 spin state. Such state is called a singlet state. Similarly, systems having 1,2,3,4…… unpaired electrons refer to doublet, triplet, quartet etc. state respectively

Excited –state processes in molecules 1.Collisional deactivation 2.Fluorescence 3.phosphorescence

processes in molecules Absorption Collisional deactivation Fluorescence Phosphorescence Intersystem crossing Vibrational relaxation v0 v3 v5 v7 v0 v3 v5 v7 S1 T v0 v3 v5 v4 v2 S0

Relation between intensity of fluorescence and concentration We know that “less number of molecules absorb lesser radiation and so emit lesser radiation”. “similarly more number of molecules absorb more radiation”.

Fluorescence intensity=Q x Ia Ia -intensity of absorbed light Q – fluorescence efficiency Q= fluorescence quanta emitted EMR quanta absorbed

Since Emission is proportional to Absorption. Ia = Io- It I0 = Intensity of incident light It = Intensity of transmitted light It = I0 e –act Ia = I0 – I0 e –act Ia = I0 – (1-e –act) Ia = I0(1-(1-act)) because e –act =1-act

Ia = Io (1-1+act) Ia = I0 x act F.I = Q x Ia F.I = QI0act Q = constant for a particular substance I0 = constant for an instrument a = molecular extinction coefficient T = path length C = concentration

Fluorescence intensity is proportional to the concentration of the substance. If the concentration is high it does not fallow due to quenching effect. concentration % intensity Low con High con

Quenching Quenching is the decrease in fluorescence intensity Types : Self or concentration quenching Chemical quenching(ph,o2,halides,heavy metals) Static quenching Collisional quenching

Factors Nature of molecules Nature of substitutent’s Effect of concentration Light Methods of illumination Oxygen Photodecomposition Temperature and viscosity Adsorption

Nature of molecules Compounds which absorb U.V and VIS radiations shows the fluorescence. Molecules having conjugated double bond are suitable. Aliphatic and Saturated cyclic organic compounds are not suitable.

Nature of substitutents Electron donating groups like –NH2, -OH enhance the fluorescence. some of the groups like -SO3H,-NH4 and Alkyl groups not effect Phosphorescence and Fluorescence. Electron withdrawing groups like -COOH, -NO2,-N=N- and Halides decrease or even destroy the Fluorescence

Effect of concentration Fluorescence α Concentration (only for small values ) Concentration should low to get good results weak strong

Light: Monochromatic light is essential. Methods of illumination :Right angle method is only used ,another method is frontal method. Oxygen : It shows two effects - Oxidation of fluorescent substance in to non-fluorescence substance. - Quenching the fluorescence. Photodecomposition : Radiation passing through the solutions is weak adequate for the measuring.

Photodecomposition : Radiation passing through the solution is weak adequate for the measuring Temperature and viscosity : variations in temperature and viscosity will cause variations in the frequency Adsorption: For the fluorimetry we need 10 – 100 weaker solution than the spectrophotometry

Instruments Source : mercury vapor lamp, xenon arc lamp, tungsten lamp. Filters and monochromators primary filter secondary filter Sample cell Detectors

Instruments Source :Mercury vapor lamp, Xenon arc lamp, Tungsten lamp. Filters and monochromators primary filter secondary filter Sample cell Detector’s

Single beam fluorimeter cuvette Low pass filter High pass filter Photomultiplier tube Tungsten source

Double beam fluorimeter Sample Solution Secondary Filter Primary Filter Photo Multiplier Tube Lamp Primary Filter Secondary Filter Reference or blank

Applications Inorganic chemistry Organic chemistry Biological Food products Pharmaceutical Clinical Natural products

Notes Fluorescence- relaxation through the emission of a photon Generally occurs on the ms-ns timescale. Phosphorescence is the emission of a photon from a excited triplet state Phosphorescence is an extremely rare property

Notes Absorption occurs on the 10-15 seconds Relaxation from the ground state occurs through the fastest available process Most molecules do not fluoresce because the excited vibrational states of S0 overlap with the S1 state and relaxation can take place rapidly by vibrational relaxation Generally on the picoseconds timescale

Thank you