Metal ion burst: Examining metal ion diffusion using ultrafast fluorescence spectroscopy in reverse...
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- 1. METAL ION BURST: EXAMINING METAL ION DIFFUSION USING ULTRAFAST FLUORESCENCE SPECTROSCOPY IN REVERSE MICELLES CHELSEY CROSSE ORALS PRESENTATION APRIL 29, 2014
2. to develop methods to measure the impact of metal ions on a wide range of timescales. 1 RESEARCH FOCUS: Example: Proposed work with Dr. Crans observing interaction of Cu2+ and -amyloid. All current work is directed in this capacity. Illustration courtesy of Alzheimers Disease Research, a program of the American Health Assistant Foundation 3. A STEP BACK: TIME RESOLVED SPECTROSCOPY 2 Initialization t0 t Changes Equilibrium 0 1 2 3 0 5 10 15 20 Absorbance (~620nm) Time Example Kinetic Trace at Single Wavelength 0 0.2 0.4 0.6 0.8 1 400 500 600 700 Absorbance (t~8) Wavelength (nm) Example Spectrum at Single Time 4. Rate of observation Rate of initialization 3 LIMITS OF TIME RESOLVED SPECTROSCOPY t0 1s Pouring Human Eye Photolysis Release Optical Gating Highest Possible Time Resolution 1ps (approximate temporal orders) Micro Channel Plate (electronic limit) 5. pH jump Photoacids excited by femtosecond pulses Releases H+ Regenerates Well characterized release dynamics Caged-metal organic complexes Cages photolysed by femtosecond pulses Releases metal ion Does not regenerate Release dynamics not characterized on fs scale to our knowledge 4 PHOTOLYSIS RELEASE TECHNIQUES HA hn A- +H+ time HA AMB hn A+M +B M2+ M2+ hn H+ hn H+ Donten, M. L., Hamm, P., & VandeVondele, J. (2011). A Consistent Picture of the Proton Release Mechanism of oNBA in Water by Ultrafast Spectroscopy and Ab Initio Molecular Dynamics. The Journal of Physical Chemistry B , 115, 1075-1083. First Step 6. 5 METAL ION DETECTION BY FLUORESCENT PROBE 1. Photolysis 2. Diffusion 3. Interaction 1. Photolysis 2. Diffusion 3. Interaction 1. Photolysis 2. Diffusion 3. Interaction 1. Photolysis 2. Diffusion 3. Interaction (1) hnM2+ (3) Interaction M2+ Probe (2) DiffusionM2+ M2+ 7. 6 Requirements: Ultrafast probe technique Well characterized probe Limited diffusion distance (1) hnM2+ M2+ Probe M2+ M2+ METAL ION DETECTION BY FLUORESCENT PROBE 8. Energy of electronic states depends on: Molecule geometry Interactions with environment Changes can be observed as: Changes in fluorescence intensity Shifts in fluorescence spectra 7 TIME RESOLVED FLUORESCENCE ON MOLECULAR TIMESCALE System Coordinate t t FluorescenceIntensity 9. Detection of fluorescence signal using non-linear optical gating. 8 FLUORESCENCE UPCONVERSION wgated =wfl +wgate Non-Linear Crystal Gated Fluorescence Signal (gated) 10. Portion of fluorescence signal sampled can be changed by: Changing gate arrival time temporal region Changing crystal angle spectral region 9 FLUORESCENCE UPCONVERSION: TEMPORAL RESOLUTION Non-Linear Crystal Gated Fluorescence SignalIntensity Crystal Out Crystal In 11. Each frequency requires realignment of crystal angle Kinetic traces can be used to reconstruct temporal fluorescence spectra 10 FLUORESCENCE UPCONVERSION: EXAMPLE DATA Zhang, X. -X.; Wrth, C.; Zhao, L.; Resch-Genger, U.; Ernsting, N. P.; Sajadi, M. Femtosecond broadband fluorescence upconversion spectroscopy: Improved setup and protometric correction. Review of Scientific Instruments 2011, 82, 063108. t FluorescenceIntensity 12. 11 FLUORESCENCE UPCONVERSION: TEMPORAL RESOLUTION BBO Sample Nd:Vanadate Pump Laser Mode-locked Ti:Sapphire Laser Optical Delay Stage BBO Elliptical Mirror Monochromator PMT LabVIEW VI Used successfully by Levinger group to examine water environments I have rebuilt most of this system, repairs & replacements have prevented completion Gate Excitation Gated Signal 13. I have experience aligning necessary optical systems: Assembled & successfully mode-locked Ti:Sapphire oscillator system Used & adjusted FROG system Designed & built other non-linear system 12 FLUORESCENCE UPCONVERSION: PERSONAL EXPERIENCE 14. 13 RELATIVE TIMESCALES Fluorescence lifetime: ~10-9 s Diffusion of metal ion: 10-13-10-6s (1) hnM2+ M2+ Probe M2+ M2+ 15. 14 Requirements: Ultrafast probe technique Well characterized probe Limited diffusion distance (1) hnM2+ (3) Interaction M2+ Probe (2) DiffusionM2+ M2+ METAL ION DETECTION BY FLUORESCENT PROBE 16. Ideal fluorophore: 1. Inert 2. Selective & sensitive 3. Detectable by our system, currently: Absorption: 400-450nm Emission: 450-500nm Detectable intensity 15 FLUOROPHORE SELECTION 17. Initial characterization: Steady State Fluorescence Fluorescence Lifetime (TCSPC) Have been able to observe quenching by Cu2+. 16 COUMARIN 343 (C343) Previously used successfully by Levinger group in fluorescence upconversion studies. 18. 0 500 1000 1500 2000 2500 3000 3500 4000 4500 5000 450 460 470 480 490 500 510 520 530 540 550 FluorescenceIntensity[counts] Wavelength (nm) Steady State Fluorescence Spectra for Solutions of C343 excited by 372nm NanoLED with Varying Concentrations of CuSO4 0.0E+00 5.5E-06 1.1E-05 2.2E-05 4.4E-05 1.1E-04 2.2E-04 Concentration CuSO4 (M) 17 C343: STEADY STATE FLUORESCENCE 0 1 2 3 4 5 0.0E+00 1.0E-04 2.0E-04 Thousands Concentration CuSO4 [M] Peak Fluorescence Intensity 19. 18 C343: FLUORESCENCE LIFETIME Fluorescence Lifetime Measurements of 2.2E-5 [M] C343 with Varying Concentration of CuSO4 20. Used to determine complex stoichiometry. 1. Measure absorbance of mixture solution at different mole fractions 2. Subtract the absorbance of the pure compounds in solution at the concentration measured 19 JOB PLOT Harris Quantitative Chemical Analysis Fig. 19-8 A(PX, x)= A(solution, x)- A(P,[P])- A(X,[X]) 21. C343: BEERS LAW 20 In cooperation with Angela Warner. A =ebc y = 0.253x R = 0.9636 0 0.1 0.2 0.3 0.4 0.5 0.6 0 0.5 1 1.5 2 2.5 Absorbance Concentration of C343 (10-5M) Beer's Law Plot C343 at 409 nm y = 0.3417x R = 0.9997 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0 0.5 1 1.5 2 2.5 PeakAbsorbance Concentration (10-5M) Beer's Law Plot of Tryptophan at 212 nm C343: R = 0.96364 22. JOB PLOT: TRYPTOPHAN 21 0 0.005 0.01 0.015 0.02 0.025 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 CorrectedAbsorbance Mole Fraction of Cu2+ Job Plot of Tryptophan with CuSO4 Corrected Abs @204nm Corrected Abs @212nm Corrected Abs @219nm Corrected Abs @230nm Corrected Abs @246nm Corrected Abs @279nm Corrected Abs @300nm In cooperation with Angela Warner. 0 0.2 0.4 0.6 0.8 200 250 300 350 Absorbance Wavelength (nm) Absorbance Spectrum of Tryptophan (2.2 10-5 M) 23. 22 Requirements: Ultrafast probe technique Well characterized probe Limited diffusion distance (1) hnM2+ (3) Interaction M2+ Probe (2) DiffusionM2+ M2+ METAL ION DETECTION BY FLUORESCENT PROBE 24. Open solution: Sensor-ion interaction determined by Brownian diffusion Large probability distribution Limited measurement resolution Confined environment: Sensor-ion interaction probability enhanced by proximity Increased reaction times have been shown previously Choose reverse micelles (RMs) Levinger group has expertise in characterization & preparation. 23 CHARACTERIZATION OF METAL ION RELEASE M2+ Probe M2+ Probe 25. 24 Ultrafast probe technique Fluorescence upconversion Currently being reassembled Well characterized probe Analytical characterization techniques have been validated Still in the process of testing candidates Limited diffusion distance Confinement in reverse micelles Relying on expertise of Levinger group METAL ION DETECTION BY FLUORESCENT PROBE 26. Immediate: Identify fluorophore Build upconversion experiment Characterize full fluorophore system using fluorescence upconversion Future: Extend upconversion to metal ion burst Measure metal ion dynamics in interesting systems 25 GOALS M Probe 27. Dr. Nancy Levinger Dr. Debbie Crans Angela Warner Ben Wiebenga-Sanford All the Levinger & Crans group folks Barisas (use of the IBH) CIF Jenee Cyran & Laura Tvedke 26 ACKNOWLEDGEMENTS 28. 27 ANATOMY OF A REVERSE MICELLE Organic solvent Aqueous suspension Surfactant molecules 29. 28 REVERSE MICELLE: TERNARY PHASE DIAGRAM Size Described in number of water molecules (w0) in each RM Easily adjusted by changing proportions of solution Well documented characteristics Rate enhancements observed in confined reaction systems Magalhaes Phase diagram of a lyotropic mixtrue sodium bis (2 ethylexyl) sulfosuccinate-codecanol-water:(1998) Confinement papers from Levinger (78-80 PRF) 30. Photolytic release of metal ions into solution Complex dissociation has been observed to be faster than 10 s Variety of compounds with different optical absorbances Diffusion time of metal ion has yet to be characterized 29 CAGED METAL ORGANIC COMPLEXES H. M. D. Bandara, D. P. Kennedy, E. Akin, C. D. Incarvito, and S. C. Burdette, "Photoinduced Release of Zn2+ with ZinCleav-1: a Nitrobenzyl-Based Caged Complex", Inorganic Chemistry 48 (17), 8445-8455 (2009). H. M. D. Bandara, T. P. Walsh, and S. C. Burdette, "A Second-Generation Photocage for Zn2+ Inspired by TPEN: Characterization and Insight into the Uncaging Quantum Yields of ZinCleav Chelators", Chemistry-a European Journal 17 (14), 3932-3941 (2011). K. L. Ciesienski and K. J. Franz, "Keys for Unlocking Photolabile Metal-Containing Cages", Angewandte Chemie-International Edition 50 (4), 814-824 (2011). K. L. Ciesienski, K. L. Haas, and K. J. Franz, "Development of next-generation photolabile copper cages with improved copper binding properties", Dalton Transactions 39 (40), 9538-9546 (2010). 31. Because we are observing quenching effects, we will be working in lower intensity regions. It may be necessary to increase instrument sensitivity. OPAGE would gate signal in an OPA crystal to account for this It has been tried before, but have been some challenges 30 OPAGE 32. 31 C343 CHARACTERIZATION: QUALITATIVE FLUORESCENCE 0 200000 400000 600000 800000 1000000 1200000 1400000 440 460 480 500 520 540 Intensity Emission [nm] 0 drops 1 drop 0.01 M 2 drops 0.01 M 1 drop 0.1M 2 dr