Presentation on theme: "20040300 XMUGXQ PFS0401 Principles of Fluorescence Spectroscopy Chemistry Department XMU."— Presentation transcript:
20040300 XMUGXQ PFS0401 Principles of Fluorescence Spectroscopy Chemistry Department XMU
20040300 XMUGXQ PFS0401 Chapter Four Factors Influencing Fluorescent Emission
20040300 XMUGXQ PFS0401 Factors Influencing Fluorescent Emission 4.1 Effect of SolventEffect of Solvent 4.2 Effect of TemperatureEffect of Temperature 4.3 Effect of pHEffect of pH 4.4 Effect of Hydrogen bondEffect of Hydrogen bond 4.5 Effect of Heavy atomEffect of Heavy atom 4.6 Effect of SurfactantEffect of Surfactant
20040300 XMUGXQ PFS0401 4.1 Effect of Solvent The phenomena of solvent effect Shifting Emission Wavelengths Changing quantum yield Changing anisotropy Changing fluorescence lifetime
20040300 XMUGXQ PFS0401 General and Specific Solvent effects General solvent effect Specific solvent effect dielectric constant n refractive index Specific chemical interaction Hydrogen bonding Complexation Charge transfer Acid-base reaction Solvent properties Solute properties as well as solvent properties Reflect the freedom of motion of the electrons in the solvent molecules, and the dipole moment of these molecules.
20040300 XMUGXQ PFS0401 General solvent effect Franck-Condon principle Solvent relaxation a b 0-0 hv A c d 0-0 hv F Relaxation **
20040300 XMUGXQ PFS0401 The Lipper equation Stoke’s shift
20040300 XMUGXQ PFS0401 The Lipper equation a radius of cavity in which the fluorophore reside *, dipole moment of ground state and excited states, reapectively
20040300 XMUGXQ PFS0401 The Lipper equation Orientation polarizability * > Gas phaseIn solution f (n) f ( ) Solvent relaxation ff How about = n 2 ?
20040300 XMUGXQ PFS0401 Example 1 n ff em hexane1.8741.3720.001135350.4 methanol33.11.3260.30989740531.1 Assume * - = 20 Debye A unit-charge separation of 4.2 Å Example 2 Assume * - = 20 Debye ex = 350 nm In nonpolar solution, observed Why?
20040300 XMUGXQ PFS0401 Derivation of Lipper Equation
20040300 XMUGXQ PFS0401 Application of Lipper Equation For a given fluorophore
20040300 XMUGXQ PFS0401 Determination of dipole moment of excited state For a given solvent, measure the dipole moment
20040300 XMUGXQ PFS0401 Specific solvent effect General solvent effect: the effect of the properties of solvent on the emitting behavior of fluorophore. Specific solvent effect: changing to an new species that fluoresces differently, duo to the reaction between fluorophore and the solvent molecule. Discrimination Electronic polarizability, molecular polarizability S mall change of solvent constituent could cause large shift of emission wavelength. Spectrum shape, not only emission wavelength, change Not follow the lipper equation
20040300 XMUGXQ PFS0401 Effect on the structural detail of spectrum
20040300 XMUGXQ PFS0401 Effect of pH Acid-base reaction of ground state fluorophore Difference in fluorescent characteristics between conjugate acid and base em, HA em, A F F pH em, HA Non-fluorescent F pH em, A Non-fluorescent F pH
20040300 XMUGXQ PFS0401 Effect on the composition of fluorophore Changing pH may change the composition of metal- ligand compound
20040300 XMUGXQ PFS0401 Acid-base reaction of excited state fluorophore pK a * = pK a pKapKa * * HA + A + hv em,A Non- fluorescent A pH pKapKa * F pKapKa pKapKa * Radiating takes place prior to acid- base reaction
20040300 XMUGXQ PFS0401 Acid-base reaction of excited state fluorophore pK a * < pK a pKapKa * * HA + A + hv em,A Non- fluorescent A pH pKapKa * pKapKa Acid-base reaction finished before radiating pKapKa * F pH
20040300 XMUGXQ PFS0401 Acid-base reaction of excited state fluorophore pK a * > pK a pKapKa * * HA + A + hv em,A Non- fluorescent A pH pKapKa * pKapKa Acid-base reaction finished before radiating F pH pKapKa *
20040300 XMUGXQ PFS0401 Excited-state intra-molecule proton transfer 水杨酸酯
20040300 XMUGXQ PFS0401 Effect of hydrogen bond Ground state: changing absorption as well as emission spectrum Excited state: changing emission spectrum Effect on n → * transition n ** Hydrogen bond Blue shift absorption
20040300 XMUGXQ PFS0401 Effect on *→n transition n ** Hydrogen bond Solvent relaxation Intensify solvent effect Red shift emission Blue shift
20040300 XMUGXQ PFS0401 Changing the type of low-energy transition Blue shift Changing the transition type Intensify emission
20040300 XMUGXQ PFS0401 Effect on quantum yield Generally, intensity IC, decrease quantum yield When transition-type changing occurs, intensity emission Intra-molecular hydrogen bond Intensity IC, F is 100 times lower than that of 5-hydroxyl- quinoline Almost same absorption Why?
20040300 XMUGXQ PFS0401 Effect of heavy atom Intra-molecule In the solvent
20040300 XMUGXQ PFS0401 4.6 effect of surfactant Surfactant Aggregation Micelle Critical micelle concentration CMC Surfactants used in fluorimetry cation C 16 H 33 N + (CH 3 ) 3 Br - 溴化十六烷基三甲铵 CTAB
20040300 XMUGXQ PFS0401 Surfactants used in fluorimetry anion C 12 H 25 SO 4 - Na + 十二烷基硫酸钠 C 12 H 25 SO 3 - Na + 十二烷基磺酸钠
20040300 XMUGXQ PFS0401 Surfactants used in fluorimetry Amphoteric 两性型 Sulfobetaine, SB-12 N- 十二烷基 -N,N- 二甲基铵 -3- 丙烷 -1- 磺酸 C 12 H 25 N + (CH 3 ) 2 (CH 2 ) 3 SO 3 -
20040300 XMUGXQ PFS0401 Surfactants used in fluorimetry Neutral 非离子型 Triton X-100
20040300 XMUGXQ PFS0401 Application of surfactant Sensitize fluorescence intensity Decrease the quenching of oxygen Decrease the quenching due to collision Increase the solubility of fluorophore in water Decrease the interference from the other species Micelle-sensitized fluorimetry Micelle-stabilized room temperature phosphorimetry Simulating membranous micro-environment