1. Tailoring Tripodal Ligands for Zinc Sensing MARM 2008 05/20/2008 Zhaohua Dai Department of Chemistry & Physical Sciences, NY zdai@pace.edu

2. Zinc in Brain The second most abundant transition metal in human body Stimulates ~200 enzymes and proteins More Zn 2+ in brain than in any other organ Zn 2+ and Cu 2+ are implicated in Alzheimer’s, Parkinson’s, and Amyotrophic Lateral Sclerosis (ALS) Complicated roles Tools are needed to image Zn 2+ distribution and kinetics TSQ, Zinquin High sensitivy Poor Zn(II)/Cu(II) selectivity M. P. Cuajungco, G. J. Lees, R. R. Kydd, R. E. Tanzi, A. I. Bush, Nutr. Neurosci., 1999, 2(4), 191 C. J. Fahrni and T. V. O’Halloran, J. Am. Chem. Soc., 1999, 121, 11448 Koh et al. Science 1996, 272, 1013–1016

3. Tripodal Ligands for Zinc Sensing

4. Our First Generation Tripods 1 Castagnetto, J.M.; Canary, J.W. Chem. Commun., 1998, 203 log  Zn 2+ Cu 2+ 11.00 16.15 Zn 2+ /Cu 2+ selectivity 10 -5 2 Anderegg, G. et al. Helv. Chim. Acta 1977, 60, 123-140

5. Sensitivity: 8-Hydroxyquinoline Based Tripods

6. Ratiometric Time-Resolved Fluorescence Royzen, M.; Durandin, A.; Young, Jr., V.G.; Geacintov, N.E.; Canary, J.W. J. Am. Chem. Soc., 2006, 128, 3854-5.  f (1) = 0.64 ns  f (2) = 24.9 ns A549 Cells No added Zn 2+ Treated with zinc pyrithione

7. Chiral Fluorescent Probes for Zn 2+ 1. Higher Zn 2+ /Cu 2+ Selectivity Stereochemical Control 2. Better contrast Fertile Optical Information: Differential Circularly Polarized Fluorescence Excitation (CPE)

8. Strategy to Higher Zn/Cu Selectivity: Preorganized Chiral Ligands PiperidinesPodands The design of a metal ion selective ligand must involve a high degree of preorganization for a specific metal ion and also a high degree of ‘disorganization’ or ‘mismatch’ for other metal ions. ------------- Peter Comba Comba, P. Coord. Chem. Rev. 1999, 185-186, 81

9. Inner Sphere Requirements E. A. Ambundo, M.-V. Deydier, A. J. Grall, N. Aguera-Vega, L. T. Dressel, T. H. Cooper, M. J. Heeg, L. A. Ochrymowycz and D. B. Rorabacher, Inorg. Chem., 1999, 38, 4233 Cu II d 9 Zn II d 10 STRONG GEOMETRICAL REQUIREMENTS FOR COORDINATION TO METAL GEOMETRY DICTATED ONLY BY STERICS 4 3 Xu, X.

10. Binding Constants Zn 2+ /Cu 2+ selectivity 10 -4 10 -1 log  Cu 2+ Zn 2+ 14.8 10.1 12.0 11.2 Obtained by Xu, X. through potentiometric titration TPA: 10 -5

11. Fluorescent Sensors Dai, Z.; Xu, X.; Canary, J.W. Chem. Commun., 2002, 1414-5. BITE LIGHT 16

12. Zn(II)/Cu(II) Selectivity Zn 2+ 7.44 7.08 Cu 2+ 7.64 7.06 Zn/Cu 0.6 1.0 log  16 17

13. More Highly Preorganized Chiral Ligangds Piperidines Quinuclidines

14. Representative Synthesis

15. X-Ray Structure of Cu(II) Complexes

16. Zn 2+ 11.0 7.1 8.95 Cu 2+ 16.15 7.1 7.0 10 -5 1 90* Stereochemical Approach to Improved Zn(II)/Cu(II) Selectivity 15% acetonitrile/aqueous buffer pH 7.19 * Z. Dai, et al. unpublished Zn 2+ /Cu 2+ Selectivity: log 

17. Increase Contrast: Reducing Background Lower sensor background Diminish background from non-analyte Chiral Fluorescent Sensors: Fertile Optical Information: Differential Circularly Polarized Fluorescence Excitation (CPE)

18. Fluorescence-detected Circular Dichroism (FDCD) J-8100 Circular Dichroism System with FDCD Attachment Nehira; Berova; Nakanishi; et al. J. Am. Chem. Soc. 1999, 121, 8681  F = Two channels of data Conversion

19. Differential Circularly Polarized Fluorescence Excitation (CPE) Changes in  F will be very large when changes in BOTH fluorescence AND circular dichroism are large. CPE utilized only  F part of FDCD raw data for analysis.  : CD ellipticity;  : Fluorescence quantum yield.

20. CPE Reduces Background from Free Ligand /nm Relative Intensity I f Zn 2+ /nm Ellipticity  /mdeg Zn 2+ /nm CPE  F Zn 2+ Free ligand [Zn(L)] 2+ Dai, Z.; Proni, G.; Mancheno, D.; Karimi, S.; Berova, N.; Canary, J.W. J. Am. Chem. Soc., 2004, 126, 11760

21. CPE SELECTS AGAINST PROTEIN-BASED BACKGROUND FLUORESCENCE /nm Relative Intensity I f Lysozyme Zn 2+ CPE  F /nm Zn 2+ Lysozyme + [Zn(L)] 2+ Dai, Z.; Proni, G.; Mancheno, D.; Karimi, S.; Berova, N.; Canary, J.W. J. Am. Chem. Soc., 2004, 126, 11760 Ellipticity  /mdeg Zn 2+ /nm

22. Tailoring Tripodal Ligands for Zinc Sensing Zhaohua Dai and James W. Canary, New J. Chem., 2007, 31, 1708-1718.

23. Conclusion Achieved solid Zn(II)/Cu(II) selectivity through a stereochemical approach Developed a new approach for analysis: CPE CPE may be used to improve contrast in detecting metal ions by fluorescent, chiral ligands with low background CPE may be used to diminish interference from fluorescent non-analytes CPE needs further development

24. Chiral Fluorescent Sensor for Hg 2+ We intend to use these ligands to further develop CPE.

25. Acknowledgement Prof. James W. Canary (NYU) Prof. Nina Berova Mike Isaacman Cho Tan Amanda Mickley Patrick Carney Nikhil Khosla NSF (JWC) Pace University (Startup Fund, Scholarly Research Fund, Kenan Award)