Molecular Fluorescence Principles and Applications
, by Valeur, Bernard; Berberan-Santos, Má; rio Nuno
- ISBN: 9783527328468 | 3527328467
- Cover: Paperback
- Copyright: 3/25/2013
This second edition of the well-established bestseller is completely updated and revised with approximately 30% additional material, including two new chapters on applications, which has seen the most significant developments. The comprehensive overview written at an introductory level covers fundamental aspects, principles of instrumentation and practical applications, while providing many valuable tips. For complex, physical, food and photochemists, molecular physicists, lecturers and students of chemistry and physics, and molecular biologists.
Bernard Valeur received his engineering diploma from the ?cole Sup?rieure de Physique et de Chimie Industrielles de Paris (E.S.P.C.I.) and his PhD degree from the Universit? Pierre-et-Marie-Curie (Paris, France), followed by postdoctoral research at the University of Illinois at Urbana-Champaign (USA). After being an associate professor at E.S.P.C.I, he became full professor of physical chemistry at the Conservatoire National des Arts et M?tiers (Paris) in 1979, where he is emeritus professor since 2008. Professor Valeur is a member of the laboratory Photophysique et Photochimie Supramol?culaires et Macromol?culaires at the ?cole Normale Sup?rieure de Cachan since 1996. From 1995 to 2000 he served as an elected member of the French Comit? National de la Recherche Scientifique. He is the winner of the 2012 Gregorio Weber Award for Excellence in Fluorescence Theory and Applications and author of over 170 articles or book chapters, five books, and the editor of one book. In addition, he is a member of several editorial boards.
M?rio Nuno Berberan-Santos graduated in chemical engineering from Instituto Superior T?cnico (IST, Technical University of Lisbon, Portugal). After a brief stay at the National Research Council of Canada (Ottawa), he received his PhD in chemistry from IST in 1989. He was a post-doctoral fellow with Bernard Valeur at Conservatoire National des Arts et M?tiers (Paris, France), and at Laboratoire pour l'Utilisation du Rayonnement Electromagn?tique (Univ. Paris-Sud, Orsay, France). He is full professor of Physical Chemistry at IST, and was invited full professor at the ?cole Normale Sup?rieure de Cachan (France). He is a member of several editorial advisory boards and is president of the Portuguese Chemical Society (2010-2012). He has authored over 180 publications, including 150 papers in scientific journals, several book chapters, and was the editor of one book.
M?rio Nuno Berberan-Santos graduated in chemical engineering from Instituto Superior T?cnico (IST, Technical University of Lisbon, Portugal). After a brief stay at the National Research Council of Canada (Ottawa), he received his PhD in chemistry from IST in 1989. He was a post-doctoral fellow with Bernard Valeur at Conservatoire National des Arts et M?tiers (Paris, France), and at Laboratoire pour l'Utilisation du Rayonnement Electromagn?tique (Univ. Paris-Sud, Orsay, France). He is full professor of Physical Chemistry at IST, and was invited full professor at the ?cole Normale Sup?rieure de Cachan (France). He is a member of several editorial advisory boards and is president of the Portuguese Chemical Society (2010-2012). He has authored over 180 publications, including 150 papers in scientific journals, several book chapters, and was the editor of one book.
Preface to the First Edition XV
Preface to the Second Edition XVII
Acknowledgments XIX
Prologue XXI
1 Introduction 1
1.1 What Is Luminescence? 1
1.2 A Brief History of Fluorescence and Phosphorescence 2
1.2.1 Early Observations 3
1.2.2 On the Distinction between Fluorescence and Phosphorescence: Decay Time Measurements 10
1.2.3 The Perrin–Jablonski Diagram 12
1.2.4 Fluorescence Polarization 14
1.2.5 Resonance Energy Transfer 16
1.2.6 Early Applications of Fluorescence 17
1.3 Photoluminescence of Organic and Inorganic Species: Fluorescence or Phosphorescence? 19
1.4 Various De-Excitation Processes of Excited Molecules 20
1.5 Fluorescent Probes, Indicators, Labels, and Tracers 21
1.6 Ultimate Temporal and Spatial Resolution: Femtoseconds, Femtoliters, Femtomoles, and Single-Molecule Detection 23
General Bibliography: Monographs and Books 25
Part I Principles 31
2 Absorption of Ultraviolet, Visible, and Near-Infrared Radiation 33
2.1 Electronic Transitions 33
2.2 Transition Probabilities: The Beer–Lambert Law, Oscillator Strength 39
2.3 Selection Rules 46
2.4 The Franck–Condon Principle 47
2.5 Multiphoton Absorption and Harmonic Generation 49
Bibliography 51
3 Characteristics of Fluorescence Emission 53
3.1 Radiative and Nonradiative Transitions between Electronic States 53
3.1.1 Internal Conversion 56
3.1.2 Fluorescence 56
3.1.3 Intersystem Crossing and Subsequent Processes 57
3.1.3.1 Intersystem Crossing 58
3.1.3.2 Phosphorescence versus Nonradiative De-Excitation 60
3.1.3.3 Delayed Fluorescence 60
3.1.3.4 Triplet–Triplet Transitions 61
3.2 Lifetimes and Quantum Yields 61
3.2.1 Excited-State Lifetimes 61
3.2.2 Quantum Yields 64
3.2.3 Effect of Temperature 66
3.3 Emission and Excitation Spectra 67
3.3.1 Steady-State Fluorescence Intensity 67
3.3.2 Emission Spectra 68
3.3.3 Excitation Spectra 71
3.3.4 Stokes Shift 72
Bibliography 74
4 Structural Effects on Fluorescence Emission 75
4.1 Effects of the Molecular Structure of Organic Molecules on Their Fluorescence 75
4.1.1 Extent of the π-Electron System: Nature of the Lowest-Lying Transition 75
4.1.2 Substituted Aromatic Hydrocarbons 77
4.1.2.1 Internal Heavy Atom Effect 77
4.1.2.2 Electron-Donating Substituents: –OH, –OR, –NH2, –NHR, –NR2 78
4.1.2.3 Electron-Withdrawing Substituents: Carbonyl and Nitro Compounds 78
4.1.2.4 Sulfonates 79
4.1.3 Heterocyclic Compounds 80
4.1.3.1 Compounds with Heteronitrogen Atoms 80
4.1.3.2 Coumarins 81
4.1.3.3 Xanthenic Dyes 82
4.1.3.4 Oxazines 84
4.1.3.5 Cyanines 85
4.1.3.6 BODIPY Fluorophores 86
4.1.4 Compounds Undergoing Photoinduced ICT and Internal Rotation 87
4.2 Fluorescence of Conjugated Polymers (CPs) 92
4.3 Luminescence of Carbon Nanostructures: Fullerenes, Nanotubes, and Carbon Dots 93
4.4 Luminescence of Metal Compounds, Metal Complexes, and Metal Clusters 96
4.5 Luminescence of Semiconductor Nanocrystals (Quantum Dots and Quantum Rods) 103
Bibliography 105
5 Environmental Effects on Fluorescence Emission 109
5.1 Homogeneous and Inhomogeneous Band Broadening – Red-Edge Effects 109
5.2 General Considerations on Solvent Effects 110
5.3 Solvent Relaxation Subsequent to Photoinduced Charge Transfer (PCT) 112
5.4 Theory of Solvatochromic Shifts 117
5.5 Effects of Specifi c Interactions 119
5.5.1 Effects of Hydrogen Bonding on Absorption and Fluorescence Spectra 119
5.5.2 Examples of Effects of Specifi c Interactions 120
5.5.3 Polarity-Induced Inversion of n−π* and π−π* States 123
5.6 Empirical Scales of Solvent Polarity 124
5.6.1 Scales Based on Solvatochromic Shifts 124
5.6.1.1 Single-Parameter Approach 124
5.6.1.2 Multiparameter Approach 126
5.6.2 Scale Based on Polarity-Induced Changes in Vibronic Bands (Py Scale) 129
5.7 Viscosity Effects 129
5.7.1 What is Viscosity? Significance at a Microscopic Level 129
5.7.2 Viscosity Effect on the Fluorescence of Molecules Undergoing Internal Rotations 132
5.8 Fluorescence in Solid Matrices at Low Temperature 135
5.8.1 Shpol’skii Spectroscopy 136
5.8.2 Matrix Isolation Spectroscopy 137
5.8.3 Site-Selection Spectroscopy 137
5.9 Fluorescence in Gas Phase: Supersonic Jets 137
Bibliography 138
6 Effects of Intermolecular Photophysical Processes on Fluorescence Emission 141
6.1 Introduction 141
6.2 Overview of the Intermolecular De-Excitation Processes of Excited Molecules Leading to Fluorescence Quenching 143
6.2.1 Phenomenological Approach 143
6.2.2 Dynamic Quenching 146
6.2.2.1 Stern–Volmer Kinetics 146
6.2.2.2 Transient Effects 148
6.2.3 Static Quenching 152
6.2.3.1 Sphere of Effective Quenching 152
6.2.3.2 Formation of a Ground-State Nonfluorescent Complex 153
6.2.4 Simultaneous Dynamic and Static Quenching 154
6.2.5 Quenching of Heterogeneously Emitting Systems 158
6.3 Photoinduced Electron Transfer 159
6.4 Formation of Excimers and Exciplexes 162
6.4.1 Excimers 163
6.4.2 Exciplexes 167
6.5 Photoinduced Proton Transfer 168
6.5.1 General Equations for Deprotonation in the Excited State 170
6.5.2 Determination of the Excited-State pK* 172
6.5.2.1 Prediction by Means of the Förster Cycle 172
6.5.2.2 Steady-State Measurements 173
6.5.2.3 Time-Resolved Experiments 174
6.5.3 pH Dependence of Absorption and Emission Spectra 174
6.5.4 Equations for Bases Undergoing Protonation in the Excited State 178
Bibliography 179
7 Fluorescence Polarization: Emission Anisotropy 181
7.1 Polarized Light and Photoselection of Absorbing Molecules 181
7.2 Characterization of the Polarization State of Fluorescence (Polarization Ratio and Emission Anisotropy) 184
7.2.1 Excitation by Polarized Light 184
7.2.1.1 Vertically Polarized Excitation 184
7.2.1.2 Horizontally Polarized Excitation 186
7.2.2 Excitation by Natural Light 187
7.3 Instantaneous and Steady-State Anisotropy 187
7.3.1 Instantaneous Anisotropy 187
7.3.2 Steady-State Anisotropy 188
7.4 Additivity Law of Anisotropy 188
7.5 Relation between Emission Anisotropy and Angular Distribution of the Emission Transition Moments 190
7.6 Case of Motionless Molecules with Random Orientation 191
7.6.1 Parallel Absorption and Emission Transition Moments 191
7.6.2 Nonparallel Absorption and Emission Transition Moments 192
7.6.3 Multiphoton Excitation 196
7.7 Effect of Rotational Motion 199
7.7.1 Free Rotations 200
7.7.1.1 General Equations 200
7.7.1.2 Isotropic Rotations 201
7.7.1.3 Anisotropic Rotations 203
7.7.2 Hindered Rotations 206
7.8 Applications 207
Bibliography 210
8 Excitation Energy Transfer 213
8.1 Introduction 213
8.2 Distinction between Radiative and Nonradiative Transfer 218
8.3 Radiative Energy Transfer 219
8.4 Nonradiative Energy Transfer 221
8.4.1 Interactions Involved in Nonradiative Energy Transfer 221
8.4.2 The Three Main Classes of Coupling 224
8.4.3 Förster’s Formulation of Long-Range Dipole–Dipole Transfer (Very Weak Coupling) 226
8.4.4 Dexter’s Formulation of Exchange Energy Transfer (Very Weak Coupling) 233
8.4.5 Selection Rules 233
8.5 Determination of Distances at a Supramolecular Level Using FRET 235
8.5.1 Single Distance between the Donor and the Acceptor 235
8.5.2 Distributions of Distances in Donor–Acceptor Pairs 239
8.5.3 Single Molecule Studies 242
8.5.4 On the Validity of Förster’s Theory for the Estimation of Distances 242
8.6 FRET in Ensembles of Donors and Acceptors 243
8.6.1 FRET in Three Dimensions: Effect of Viscosity 243
8.6.2 Effects of Dimensionality on FRET 247
8.6.3 Effects of Restricted Geometries on FRET 250
8.7 FRET between Like Molecules: Excitation Energy Migration in Assemblies of Chromophores 250
8.7.1 FRET within a Pair of Like Chromophores 251
8.7.2 FRET in Assemblies of Like Chromophores 251
8.7.3 Lack of Energy Transfer upon Excitation at the Red Edge of the Absorption Spectrum (Weber’s Red-Edge Effect) 252
8.8 Overview of Qualitative and Quantitative Applications of FRET 252
Bibliography 258
Part II Techniques 263
9 Steady-State Spectrofl uorometry 265
9.1 Operating Principles of a Spectrofl uorometer 265
9.2 Correction of Excitation Spectra 268
9.3 Correction of Emission Spectra 268
9.4 Measurement of Fluorescence Quantum Yields 269
9.5 Possible Artifacts in Spectrofl uorometry 271
9.5.1 Inner Filter Effects 271
9.5.1.1 Excitation Inner Filter Effect 271
9.5.1.2 Emission Inner Filter Effect (Self-Absorption) 272
9.5.1.3 Inner Filter Effects due to the Presence of Other Substances 274
9.5.2 Autofl uorescence 274
9.5.3 Polarization Effects 275
9.5.4 Effect of Oxygen 275
9.5.5 Photobleaching Effect 276
9.6 Measurement of Steady-State Emission Anisotropy: Polarization Spectra 277
9.6.1 Principles of Measurement 277
9.6.2 Possible Artifacts 279
9.6.3 Tests Prior to Fluorescence Polarization Measurements 279
Appendix 9.A Elimination of Polarization Effects in the Measurement of Fluorescence Intensity 281
Bibliography 283
10 Time-Resolved Fluorescence Techniques 285
10.1 Basic Equations of Pulse and Phase-Modulation Fluorimetries 286
10.1.1 Pulse Fluorimetry 286
10.1.2 Phase-Modulation Fluorimetry 286
10.1.3 Relationship between Harmonic Response and δ-Pulse Response 287
10.1.4 General Relations for Single Exponential and MultiExponential Decays 290
10.2 Pulse Fluorimetry 292
10.2.1 Light Sources 292
10.2.2 Single-Photon Timing Technique (10 ps–500 μs) 292
10.2.3 Streak Camera (1 ps–10 ns) 294
10.2.4 Fluorescence Upconversion (0.1–500 ps) 295
10.2.5 Optical Kerr-Gating (0.1–500 ps) 297
10.3 Phase-Modulation Fluorimetry 298
10.3.1 Introduction 298
10.3.2 Phase Fluorimeters Using a Continuous Light Source and an Electro-Optic Modulator 300
10.3.3 Phase Fluorimeters Using the Harmonic Content of a Pulsed Laser 302
10.4 Artifacts in Time-Resolved Fluorimetry 302
10.4.1 Inner Filter Effects 302
10.4.2 Dependence of the Instrument Response on Wavelength – Color Effect 304
10.4.3 Polarization Effects 304
10.4.4 Effects of Light Scattering 304
10.5 Data Analysis 305
10.5.1 Pulse Fluorimetry 305
10.5.2 Phase-Modulation Fluorimetry 306
10.5.3 Judging the Quality of the Fit 306
10.5.4 Global Analysis 307
10.5.5 Fluorescence Decays with Underlying Distributions of Decay Times 308
10.6 Lifetime Standards 312
10.7 Time-Resolved Polarization Measurements 314
10.7.1 General Equations for Time-Dependent Anisotropy and Polarized Components 314
10.7.2 Pulse Fluorimetry 315
10.7.3 Phase-Modulation Fluorimetry 317
10.7.4 Reference Compounds for Time-Resolved Fluorescence Anisotropy Measurements 318
10.8 Time-Resolved Fluorescence Spectra 318
10.9 Lifetime-Based Decomposition of Spectra 318
10.10 Comparison between Single-Photon Timing Fluorimetry and Phase-Modulation Fluorimetry 322
Bibliography 323
11 Fluorescence Microscopy 327
11.1 Wide-Field (Conventional), Confocal, and Two-Photon Fluorescence Microscopies 328
11.1.1 Wide-Field (Conventional) Fluorescence Microscopy 328
11.1.2 Confocal Fluorescence Microscopy 329
11.1.3 Two-Photon Excitation Fluorescence Microscopy 331
11.1.4 Fluorescence Polarization Measurements in Microscopy 333
11.2 Super-Resolution (Subdiffraction) Techniques 333
11.2.1 Scanning Near-Field Optical Microscopy (SNOM) 333
11.2.2 Far-Field Techniques 337
11.3 Fluorescence Lifetime Imaging Microscopy (FLIM) 340
11.3.1 Time-Domain FLIM 341
11.3.2 Frequency-Domain FLIM 342
11.4 Applications 342
Bibliography 346
12 Fluorescence Correlation Spectroscopy and Single-Molecule Fluorescence Spectroscopy 349
12.1 Fluorescence Correlation Spectroscopy (FCS) 349
12.1.1 Conceptual Basis and Instrumentation 350
12.1.2 Determination of Translational Diffusion Coefficients 355
12.1.3 Chemical Kinetic Studies 356
12.1.4 Determination of Rotational Diffusion Coefficients 359
12.1.5 Cross-Correlation Methods 360
12.2 Single-Molecule Fluorescence Spectroscopy 360
12.2.1 General Remarks 360
12.2.2 Single-Molecule Detection in Flowing Solutions 361
12.2.3 Single-Molecule Detection Using Fluorescence Microscopy Techniques 363
12.2.4 Single-Molecule and Single-Particle Photophysics 367
12.2.5 Applications and Usefulness of Single-Molecule Fluorescence 371
Bibliography 372
Part III Applications 377
13 Evaluation of Local Physical Parameters by Means of Fluorescent Probes 379
13.1 Fluorescent Probes for Polarity 379
13.1.1 Examples of Photoinduced Charge Transfer (PCT) Probes for Polarity 380
13.1.2 Pyrene and Its Derivatives 384
13.2 Estimation of “Microviscosity,” Fluidity, and Molecular Mobility 384
13.2.1 Various Methods 385
13.2.2 Use of Molecular Rotors 386
13.2.3 Methods Based on Intermolecular Quenching or Intermolecular Excimer Formation 389
13.2.4 Methods Based on Intramolecular Excimer Formation 390
13.2.5 Fluorescence Polarization Method 393
13.2.5.1 Choice of Probes 393
13.2.5.2 Homogeneous Isotropic Media 393
13.2.5.3 Ordered Systems 395
13.2.5.4 Practical Aspects 395
13.2.6 Concluding Remarks 397
13.3 Temperature 398
13.4 Pressure 402
Bibliography 404
14 Chemical Sensing via Fluorescence 409
14.1 Introduction 409
14.2 Various Approaches of Fluorescence Sensing 410
14.3 Fluorescent pH Indicators 412
14.3.1 Principles 412
14.3.2 The Main Fluorescent pH Indicators 417
14.3.2.1 Coumarins 417
14.3.2.2 Pyranine 417
14.3.2.3 Fluorescein and Its Derivatives 419
14.3.2.4 SNARF and SNAFL 419
14.3.2.5 pH Indicators Based on Photoinduced Electron Transfer (PET) 420
14.4 Design Principles of Fluorescent Molecular Sensors Based on Ion or Molecule Recognition 420
14.4.1 General Aspects 420
14.4.2 Recognition Units and Topology 422
14.4.3 Photophysical Signal Transduction 424
14.4.3.1 Photoinduced Electron Transfer (PET) 424
14.4.3.2 Photoinduced Charge Transfer (PCT) 425
14.4.3.3 Excimer Formation or Disappearance 427
14.4.3.4 Förster Resonance Energy Transfer (FRET) 427
14.5 Fluorescent Molecular Sensors of Metal Ions 427
14.5.1 General Aspects 427
14.5.2 Fluorescent PET Cation Sensors 430
14.5.3 Fluorescent PCT Cation Sensors 430
14.5.4 Excimer-Based Cation Sensors 430
14.5.5 Cation Sensors Based on FRET 430
14.5.6 Hydroxyquinoline-Based Cation Sensors 432
14.5.7 Concluding Remarks on Cation Sensors 435
14.6 Fluorescent Molecular Sensors of Anions 436
14.6.1 Anion Sensors Based on Collisional Quenching 437
14.6.2 Anion Sensors Based on Fluorescence Changes upon Anion Binding 437
14.6.2.1 Urea and Thiourea Groups 438
14.6.2.2 Pyrrole Groups 439
14.6.2.3 Polyazaalkanes 440
14.6.2.4 Imidazolium Groups 443
14.6.2.5 Anion Binding by Metal Ion Complexes 443
14.6.3 Anion Sensors Based on the Displacement of a Competitive Fluorescent Anionic Molecule 444
14.7 Fluorescent Molecular Sensors of Neutral Molecules 445
14.7.1 Cyclodextrin-Based Fluorescent Sensors 446
14.7.2 Boronic Acid-Based Fluorescent Sensors 449
14.7.3 Porphyrin-Based Fluorescent Sensors 452
14.8 Fluorescence Sensing of Gases 453
14.8.1 Oxygen 453
14.8.2 Carbon Dioxide 456
14.8.3 Nitric Oxide 456
14.8.4 Explosives 456
14.9 Sensing Devices 458
14.10 Remote Sensing by Fluorescence LIDAR 460
14.10.1 Vegetation Monitoring 461
14.10.2 Marine Monitoring 462
14.10.3 Historic Monuments 462
Appendix 14.A. Spectrophotometric and Spectrofluorometric pH Titrations 462
Single-Wavelength Measurements 462
Dual-Wavelength Measurements 463
Appendix 14.B. Determination of the Stoichiometry and Stability Constant of Metal Complexes from Spectrophotometric or Spectrofluorometric Titrations 465
Definition of the Equilibrium Constants 465
Preliminary Remarks on Titrations by Spectrophotometry and Spectrofluorometry 467
Formation of a 1 : 1 Complex (Single-Wavelength Measurements) 467
Formation of a 1 : 1 Complex (Dual-Wavelength Measurements) 469
Formation of Successive Complexes ML and M2L 470
Cooperativity 471
Determination of the Stoichiometry of a Complex by the Method of Continuous Variations (Job’s Method) 471
Bibliography 473
15 Autofluorescence and Fluorescence Labeling in Biology and Medicine 479
15.1 Introduction 479
15.2 Natural (Intrinsic) Chromophores and Fluorophores 480
15.2.1 Amino Acids and Derivatives 481
15.2.2 Coenzymes 488
15.2.3 Chlorophylls 490
15.3 Fluorescent Proteins (FPs) 491
15.4 Fluorescent Small Molecules 493
15.5 Quantum Dots and Other Luminescent Nanoparticles 497
15.6 Conclusion 501
Bibliography 502
16 Miscellaneous Applications 507
16.1 Fluorescent Whitening Agents 507
16.2 Fluorescent Nondestructive Testing 508
16.3 Food Science 511
16.4 Forensics 513
16.5 Counterfeit Detection 514
16.6 Fluorescence in Art 515
Bibliography 518
Appendix: Characteristics of Fluorescent Organic Compounds 521
Epilogue 551
Index 553
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