Tailoring Tripodal Ligands for Zinc Sensing

MARM 200805/20/2008

Zhaohua DaiDepartment of Chemistry & Physical Sciences, NY

zdai@pace.edu

Zinc in Brain• The second most abundant transition metal in human body• Stimulates ~200 enzymes and proteins• More Zn2+ in brain than in any other organ• Zn2+ and Cu2+ are implicated in Alzheimer’s, Parkinson’s, and

Amyotrophic Lateral Sclerosis (ALS)• Complicated roles• Tools are needed to image Zn2+ distribution and kinetics

N

NHO2S

R1

OR2

R3

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), 191C. J. Fahrni and T. V. O’Halloran, J. Am. Chem. Soc., 1999, 121, 11448

Koh et al. Science 1996, 272, 1013–1016

Tripodal Ligands for Zinc Sensing

HOOC

OO

R

OH

R

NNN

N N

N

ZP1: R = ClZP2: R = HZP3: R = F

ZP4: R1 = H, R2 = H, R3 = ClZP5: R1 = F, R2 = H, R3 = ClZP6: R1 = Cl, R2 = H, R3 = ClZP7: R1 = OMe, R2 = H, R3 = ClZP8: R1 = H, R2 = F, R3 = F

HOOC

O O

N

N

N

HO

R3R2

NH

R1

HOOC

O

N

N

N

HO

NH O

ZNP1

NN

N

HN

O OHO

COOH

ZnAF-2

NN

N

NH

O OHO

COOH

ZnAF-1

NN

N

NH

O OHO

COOHF F

ZnAF-1F

NN

N

HN

O OHO

COOHF F

ZnAF-2F

NN

N

HN

O OHO

COOH

ZnAF-2M

NN

N

HN

O OHO

COOH

ZnAF-2MM

NHN

O OHO

COOH

ZnAF-3

N

N

HNHN

O OHO

COOH

ZnAF-4

N

NHN

O OHO

COOH

ZnAF-5

N

N

NN

N

HN

OMe

O

ZnAF-R2 NO

COOHHOOC

OO

R

OH

R

NN

N N S

ZS1: R = ClZS2: R = F

S

Our First Generation Tripods

N

NN

2CH3

1

Castagnetto, J.M.; Canary, J.W. Chem. Commun., 1998, 203

N

NNN

logZn2+

Cu2+

11.0016.15

Zn2+/Cu2+

selectivity 10-5

2

Anderegg, G. et al. Helv. Chim. Acta 1977, 60, 123-140

Sensitivity: 8-Hydroxyquinoline Based Tripods

N

N

NN

N

N

NN

OH HO

log = 12.07

N

N

NN

OH HO

O2S

N

SO2

N

N

N

NN

OH

O2S

N

log = 14.34

log = 13.07

log = 11

N

O

N

OZn

1:2 Metal:Ligand Complexlog = 16.76log = 8.66log = 8.1

Ratiometric Time-Resolved Fluorescence

N

N

N

HO

O2S

SO2

N

NN

N

N

N

O

O2S

SO2

N

NN Zn

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 Zn2+

Treated with zinc pyrithione

Chiral Fluorescent Probes for Zn2+

1. Higher Zn2+/Cu2+ Selectivity

Stereochemical Control

2. Better contrast

Fertile Optical Information: Differential Circularly Polarized Fluorescence Excitation (CPE)

Strategy to Higher Zn/Cu Selectivity: Preorganized Chiral Ligands

PiperidinesPodands

N

N N

N

N

N N

N

H HH

H H

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

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

CuII d9

ZnII d10

N

NNN

H H

N

NNN

H H

CuX

py Npy

py

py Cupy

py

X

N

STRONG GEOMETRICAL REQUIREMENTS FOR COORDINATION TO METAL

GEOMETRY DICTATED ONLY BY STERICS

4

3

Xu, X.

Binding Constants

Zn2+/Cu2+

selectivity 10-4 10-1

logCu2+

Zn2+

14.810.1

12.011.2

N

NNN

H H

N

NNN

H H

Obtained by Xu, X. through potentiometric titration

TPA: 10-5

320 360 400 440 480 5200

10

20

30

40

50

60

70

80

90

Zn2+

I f

(nm)

Fluorescent Sensors

Dai, Z.; Xu, X.; Canary, J.W. Chem. Commun., 2002, 1414-5.

NN N

N

H

H

BITE

LIG

HT

16

Zn(II)/Cu(II) Selectivity

N

NNN

H H

N

NNN

H H

Zn2+ 7.44 7.08

Cu2+ 7.64 7.06

Zn/Cu 0.6 1.0

log

16 17

More Highly Preorganized Chiral Ligangds

N

N N

N

N

N N

N

H HH

H H

N NHN

H

HN

Piperidines Quinuclidines

Representative Synthesis

HC CH2 C Cl

O CH3NHOCH3

HC CH2 C N

O CH3

OCH3

LiN

Br

HC CH2 C OCH3

O i-PrM

gCl

CH 3NHOCH 3

N

N N

H

H H

N

MeOH

B(OH)2

N

N N

H

H

Br Br

H

N

Br

NH3

HC CH2 CHN

Br

OH

HC CH2 CN

BrO

HC CH2 CHN

Br

OMs

MsCl

3

33

Pd0

58%68%

323

24

2526

282919

3

3

27

(+)DIP-Cl

B

Cl

(+)DIP-Cl =

X-Ray Structure of Cu(II) Complexes

Zn2+ 11.0 7.1 8.95

Cu2+ 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

Zn2+/Cu2+

Selectivity:

log

NN N

N

N NHN

H

H

N

N

N N

N

H

H

Increase Contrast: Reducing Background

• Lower sensor background

• Diminish background from non-analyte

Chiral Fluorescent Sensors: Fertile Optical Information: Differential Circularly Polarized Fluorescence Excitation (CPE)

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

RLRL

RL

FF

Fk

FF

FFkS

10ln10

)101(2A

A

cd

S

Conversion

Differential Circularly Polarized Fluorescence Excitation (CPE)

Changes in F will be very large when changes in BOTH fluorescence AND circular dichroism are large.

A

IKF

10*

0

a

b

A

A

b

a

b

a

b

a

F

F

10

10

CPE utilized only F part of FDCD raw data for analysis.

: CD ellipticity; : Fluorescence quantum yield.

200 220 240 260 280 300 320 340

-5

-4

-3

-2

-1

0

1

2

200 220 240 260 280 300 320 340

-5

0

5

10

15

320 360 400 440 480 5200

200

400

600

800

1000

CPE Reduces Background from Free Ligand

/nm

Rel

ati

ve

Inte

nsi

ty I

f

Zn2+

/nm

Ell

ipti

city

/

md

eg

Zn2+

/nm

CP

E

F

Zn2+

Free ligand

[Zn(L)]2+

NN N

N

H H

(S,S)-17

Dai, Z.; Proni, G.; Mancheno, D.; Karimi, S.; Berova, N.; Canary, J.W. J. Am. Chem. Soc., 2004, 126, 11760

200 220 240 260 280 300 320 340-30

-25

-20

-15

-10

-5

0

5

300 330 360 390 420 450 480 510 5400

200

400

600

800

1000

1200

CPE SELECTS AGAINST PROTEIN-BASED BACKGROUND FLUORESCENCE

/nm

Rel

ati

ve

Inte

nsi

ty I

f

Lysozyme

Zn2+

CP

E F

/nm

Zn2+

Lysozyme

Lysozyme+

[Zn(L)]2+

Dai, Z.; Proni, G.; Mancheno, D.; Karimi, S.; Berova, N.; Canary, J.W. J. Am. Chem. Soc., 2004, 126, 11760

200 220 240 260 280 300 320 340-70

-60

-50

-40

-30

-20

-10

0

10

260 280 300 320 340-4

-2

0

2

4

6

Ell

ipti

city

/

md

eg

Zn2+

/nm

NN N

N

H H

(R,R)-17

Tailoring Tripodal Ligands for Zinc Sensing

Zhaohua Dai and James W. Canary,  New J. Chem., 2007, 31, 1708-1718.

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

Chiral Fluorescent Sensor for Hg2+

N

SO

HO

N NO

O

HO

O

O

OH

HOOC

N

SO

HO

N N

O

OH

O

COOH

1

2O

OH

O

COOH

COOH

We intend to use these ligands to further develop CPE.

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)