Wen-Sheng Chung (鍾文聖 )

Department of Applied Chemistry, National Chiao Tung University

Hsinchu, Taiwan

Ditopic Fluorescent Sensing of Heavy Metal Ions

Based on Functionalized Calix[4]arenes

方教授泰山榮退教育學術回顧與感恩研討會 December 24, 2011

方教授泰山榮退教育學術回顧與感恩研討會 December 24, 2011

OHOH

OHHO

p-tert-Butylcalix[4]arene

Advantages of using calix[4]arene as a scaffold for molecular sensing studies include: (1) flexible in modification, (2) conformational flexibility, and (3) multiple reaction sites.

Upper rim

Lower rim

OHOH HO

OH

OH OHOH OH

• Gutsche, C. D. Calix[4]arenes Revisited; The Royal Society of Chemistry: Cambridge, U.K., 1998.

(c)

OR

OR

OR OR OROR

OROR

RO

OR OR

OR

HOOHOHOH

cone partial cone 1,2-alternate 1,3-alternatene

R X Y Z

C C

+

YX

C C

ZR YXC C

ZR+ - +

-

Ex. 1 R1 C N O +

R2

R3 R5

R4 NC OR1

R3

R2 R4R52-isoxazolines

Ex. 2 R1 C N O +

NC OR1

R2 R3

R2 R3

isoxazoles

Ex. 3 R1 N N N +

NN NR1

R2 R3

R2 R3

triazoles

• Click Chemistry by Sharpless, K. B.; Fokin, V. V. et al. J. Am. Soc. Chem. 2005, 127, 210.

What is our approach?Using the powerful and versatile 1,3-dipolar cycloaddition and its subsequent ring-opening reactions to create arrays of bifunctional compounds with appended chromophores and/or fluorophores for molecular sensing studies.

Ring-Opening Products of Isoxazolines or Isoxazoles

• For a leading reference please see: Kozikowski, A. Acc. Chem. Res. 1984, 17, 410.• Tetrahedron Lett. 2006, 47, 7179; 9077.

Using Mo(CO)6 in Calix[4]arenes

R

N O

RR

C

R

R

O OHO

N OH

NH2 OH

NOH

-amino alcohol

-hydroxy ketone-unsaturated ketone1,3-diene

-hydroxy nitrile

-unsaturated oxime-unsaturated oxime

isoxazoline

R

NH2 O

-amino ,-unsaturatedketone

H2, Raney Ni

Mo(CO)6

• J. Chinese Chem. Soc. 2000, 47, 173.

Calix[4]arene with a 'lid'

Fluorogenic Chemosensor

Ref : Fabbrizzi, L.; Poggi, A. Chem. Soc. Rev. 1995, 197.

Signalingmoiety

Recognitionmoiety

Signals: Fluorescence quenching or enhancement and/ or color changes

Chemosensor

Analyte

Figure 1. Fundamentals and Functions of a Metal Ion Chemosensor.

• High sensitivity• Low background• Fast response time• Small amount of ligands• Low cost

• Allosteric Effects on the Triazole-modified Calix[4]crown toward K+ and Pb2+ Ions.

• 1,3-Alternate Calix[4]arene as a Homoditopic Fluorescent Chemosensor for Ag+ Ions.

• The concept of allosteric proteins was developed in the early 1960s by Monod and Koshland.

• Allosteric is derived from the Greek root allo, means “the other”.

• Kramer, R. Chem. Rev. 2004, 104, 3161.

Scheme 1. Synthetic pathways for fluoroionophores 4 and 5

N3

3

CuI, H2O/THF, 50 oC 24 h

82%

OHOH OOO O

O O

O

O

O

tetraethyleneglycol ditosylate

Cs2CO3, MeCN, reflux 24 h

35%

2

O OCuI, H2O/THF, rt 24 h

3

85%

N NN

5

O O

O O

O

O

O

N NN N

N N

4

1

-100

-75

-50

-25

0

25

4 5

Cu2+Hg2+Cr3+Pb2+

Li+ Na+ K+ Mg2+Ca2+Ba2+ Cd2+Ag+Ni2+Mn2+Zn2+

(I -

I o)/I o *

10

0%

Figure 1. Fluorescence intensity changes ((I – Io)/Io 100%) of fluoroionophores 4 and 5 (each of 10 M) in MeCN/CHCl3 (1000:4, v/v) at 298K upon addition of various metal perchlorates (10 equiv).

O

N NN

5

These results suggest that Hg2+, Cu2+ and Cr3+ ions can be recognized by the mono-triazole group of sensor 5 alone; however, the complexation of Pb2+ requiresthe coordination of the two triazole groups of 4.O O

O O

O

O

O

N NN N

N N

4

Off

4 Pb2+

O O

O O

O

O

O

N NN N

N N

Pb2+

O

N NN

5

Pb2+

400 450 500 550 600-1

0

1

2

3

4

5

6

7

8

[K+]/[Pb2+] ([Pb2+] = 100 uM)10080604020105.02.51.00.50.20

Free (10 uM)

Free + 10 eq K+

Flu

ore

sce

nce

Inte

nsi

ty (

a.u

.)

Wavelength (nm)

Figure 2. Fluorescence emission change for the 4Pb2+ complex in CH3CN/CHCl3

(700:3, v/v) upon addition of K+. Excitation wavelength was at 367 nm.

Allosteric Effect of 4•Pb2+ by K+ Ion

O O

O O

O

O

O

N NN N

N N

K+

Pb2+

Off On

K+

O O

O O

O

O

O

N NN N

N N

Pb2+

4 Pb2+ 4 K+

400 450 500 550 600-1

0

1

2

3

4

5

6

7

8

Free + 10 eq Pb2+

Free (10 uM)

[Pb2+]/[K+] ([K+] = 100 uM)00.20.51.02.55.01020406080

Flu

ore

sce

nce

Inte

nsi

ty (

a.u

.)

Wavelength (nm)

Figure 3. Fluorescence emission change for the 4K+ complex in CH3CN/CHCl3 (700:3,

v/v) upon addition of Pb2+. Excitation wavelength was at 367 nm.

Allosteric Effect of 4•K+ by Pb2+ Ion

O O

O O

O

O

O

N NN N

N N

K+

Pb2+

Off On

K+

O O

O O

O

O

O

N N

N N

N N

Pb2+

4 Pb2+ 4 K+

O O

O O

O

O

O

N NN N

N N

O O

O O

O

O

O

N NN N

N N

Pb2+

O O

O O

O

O

O

N NN N

N NPb2+

Pb2+

On

Off Strong emission

K+

K+

K+

Allosteric Binding

• Chang, K.-C. et al Org. Lett. 2007, 9, 3363.

• Watkinson and Todd in Chem. Soc. Rev., 2011, 40, 2848–2866, “Chemical sensors that incorporate click-derived triazoles” mentioned that: Chung and co-workers were the first to utilize click chemistry on calixarene frameworks to construct sensors.

OO

1,3-alternate calix[4]arene

Cation

Cation

OO

OO

Cation

O O

Chromogenic calix[4]arene sensor, with bistriazoles as the metal ion binding sites and azo groups as the signaling units, showed selective coloration toward Ca2+ and Pb2+ ions.

OHOH OO

N N

N N

N N

N NN

OCH3

6

OHOH OO

N N

N N

N N

N NN

OCH3

M2+

M2+ = Ca2+ or Pb2+

M2+

N

OCH3

N

OCH3

6 M2+

6 6Ca2+ 6Pb2+

• Chang, K.-C.; Chung, W.-S. et al. Tetrahedron Lett. 2007, 48, 7274.• I-Ting Ho et al. Chem. Asian. J. 2011, 6, 2738. Special issue on

the 10th anniversary of Click chemistry.

2 + Cu2+

+ Cu2+/CH3COO-

+ Cu2+/F-

2·Cu+

OO HO O

O O

NH2

NH2

OHO OH O

N N

OO

2

Cu2+

H

H H

H

A- = CH3COO- or F-

A-

2·Cu+·A-

Cu+

OO HO O

O

NNHH

-A

Cu+O

H H

a

bc

d

• Senthilvelan, A.; Ho, I-T.; Chang, K.-C.; Lee, G.-H.; Liu, Y.-H.; Chung, W.-S. Chem. Eur. J. 2009, 15, 6152

Fluorescence heteroditopic sensing of Cu(II) and anions

OH OOHOO

HN

O

NH

O

OH2N

O

NH2

O

Cu2+

O OHOO

O O

NH2

NH2

O O

HNHNO O

Cu+

Cu+

Fluorescence off Fluorescence on

2 x

L2 L2 2Cu+

Recognition site

Selective Fluorescence Turn-on Chemosensor for Two Cu(II) Ions

Fluorescence(signaling moiety)

• I-Ting Ho, J.-H. Chu, W.-S. Chung Eur. J. Org. Chem. 2011

• Allosteric Effects on the Triazole-modified Calix[4]crown toward K+ and Pb2+ Ions.

1,3-Alternate Calix[4]arene as a Homoditopic Fluorescent Chemosensor for Ag+ Ions.

OO

OO

N

NN NN

N

NH

H

O O

HNH

L3

Synthesis, UV/vis, and Fluorescence screening of L3

O O

O

OO

O

N

N N

NN

N

NN

HH

H

H

HH

2.29 Å

2.69 Å

(a) (b)

300 400 5000.0

0.2

0.4

0.6

0.8

1.0

347

Abs

orba

nce

Wavelength (nm)

332 nm

367

387

0 [Ag+]/[L3]

5.0

(a)

400 450 500 550 6000123456789 10

4 3 2

1

0 [Ag+]/[L3]

FL

Inte

nsity

(a.

u.)

Wavelength (nm)

416 nm(b)

[L3] = 20 MMeOH-CHCl3 (49:1, v/v) ex = 372 nm

UV-Vis titration spectrum Fluorescence titration spectrum

The non-linear least-squares analysis for 1:2 ligand-to-metal complexation of L3

0.0 0.2 0.4 0.6 0.80

2

4

6

8

10

FL In

tensi

ty a

t 416 n

m

[Ag+], mM

R2 = 0.9932

= K1*K

2 = 4.10E8 M-2

K1 = 4.46E3 M-1

K2 = 9.2E4 M-1

y = (Io + [L]*b*K

1*x+I

max**x 2̂)/(1+K

1*x+*x 2̂)

Figure 4. The non-linear least-squares fitting of L3 with AgClO4.

• Jiwan, J.-L. H.; Branger, C.; Soumillion, J.-Ph.; Valeur, B. J. Photoch. Photobio. A 1998, 116, 127-133.

The higher K2 revealed that a positive allosteric effect participated in the complexation of L3 with the second equiv of Ag+.

K1 = 4.46 x103 M-1

K2 = 9.2 x104 M-1

1H NMR Titration of L3 with 2 equivalents of AgClO4

456789 ppm

2.0

1.0

0.5

0 ☆

Figure 5. The 1H NMR titration of L3 (3 mM) in the presence of different amount of AgClO4 in CD3OD-CDCl3 (9:1, v/v). (a) 0, (b) 0.5, (c) 1.0 and (d) 2.0. #: CHD2OD, *: CD3OH, ※ : H2O, ☆ : external CHCl3, ◎ : internal CHCl3.

◎

g’h’

☆

☆

☆

b’

※

a’*

*

*

*

f’-CH2-bridge

#

gh

b af

-CH2-bridge

L3

OO

OO

N

NN NN

N

NH

H

O O

HNH

a

dc

e

b

g

h

f

L3 (Ag+)2

OO

OO

N

NN NN

N

NH2

O O

NH2

Ag+

Ag+

UV-Vis and Fluorescence Titration of 4 with AgClO4 in MeOH-CHCl3 (49:1, v/v)

400 450 500 550 6000

2

4

6

8

10 0 0.2 0.4 0.8 1.0 1.2 1.4 2.0 2.5 3.0 5.0 7.0 9.0

FL In

tensi

ty (

a. u

.)

Wavelength(nm)

416 nm [Ag+]/[4]

300 350 400 450 5000.0

0.1

0.2

0.3

0.4

Abso

rbance

Wavelength (nm)

0

9

[Ag+]/[4]

(a)

0.0 0.2 0.4 0.6 0.8 1.00.0

0.5

1.0

1.5

2.0

2.5

3.0

(Io-I

)*X

at 4

16 n

m

X, [4]/([4]+[Ag+])

(b)

0 4 8 12 16 200.0

0.5

1.0

1.5

2.0I o/

I at 4

16 n

m

[Ag+], M

y = a +b*xa = 1.01b = 4.09E4

R2 = 0.989

Ka = 4.09E4 M-1

OO

OO

NO O

N

N

NN NN

N

4

Stern-Volmer PlotIo/I = 1 + Ksv[M]

1H NMR Titration of 4 with AgClO4

Figure 6. The 1H NMR titration of 4 (3 mM) in the presence of different amount of AgClO4 in CD3OD-CDCl3 (3:1, v/v). (a) 0, (b) 0.5, (c) 1.0 and (d) 1.5. #: CHD2OD, *: CD3OH, ※ : H2O, ☆ : external CHCl3, ◎ : internal CHCl3.

456789 ppm

1.5

1.0

0.5

0 ☆

◎g’ h’

☆

☆

☆ b’

※

a’

*

*

*

*

f’-CH2-bridge

#

gh

b af

-CH2-bridge

OO

OO

NO O

N

NNN NN

N

a

bc

de

f

g

h

OO

OO

NO O

N

NNN NN

NAg+

4 Ag+

ESI-MS spectra

L3·(Ag+)2 formula: [C84H68N8O6Ag2]2+

4·Ag+

formula: [C84H64N8O6Ag]+

OO

OO

N

NN NN

N

NH

H

O O

HNH

Ag+

Ag+

Ag+

OO

OO

NO O

N

N

NN NN

N

• L3 exhibited as a homoditopic fluorescent chemosensor for Ag+ ions.

• A strong fluorescence enhancement was due to complexation-induced rigidity of its structure.

• Compound 4 without the ring-opening enaminone moieties did not have the ability to recognize two Ag+ ions simultaneously.

• The complexation of 4 with Ag+ led to a severe fluorescence quenching due to an inverse PET from anthracenes to the Ag+ bound nitrogen atoms of the triazole rings.

Fe, NH4Cl

4

L3

OO

OO

N

NN NN

N

NH

H

O O

HNH

OO

OO

NO O

N

N

NN NN

N

OO

OO

NO O

N

N

NN NN

N

Ag+

Ag+

Fluorescece Quenched

Ag+2 x

L3 (Ag+)2

OO

OO

N

NN NN

N

NH2

O O

NH2

Ag+

Ag+

Fluorescece Enhanced

4 Ag+

• I.-T. Ho; K.-C. Haung, and W.-S. Chung Chem. Asian J. 2011, 6, 2738.

Fluorescence TitrationFluorescence Titration

The absorbance changes of the triazole linked anthracene at 387 nm were almost saturated by adding 1 equiv of Ag+, whereas the absorbance of the enaminone at 332 nm was still increasing by the addition of more than 1 equiv of Ag+. The results indicated that the first equiv of Ag+ prefer to be bound with the bis-triazole sites, and the reoriented structure helped to bind the second equiv of Ag+ by the bis-enaminone sites.

The absorbance changes of the triazole linked anthracene at 387 nm were almost saturated by adding 1 equiv of Ag+, whereas the absorbance of the enaminone at 332 nm was still increasing by the addition of more than 1 equiv of Ag+. The results indicated that the first equiv of Ag+ prefer to be bound with the bis-triazole sites, and the reoriented structure helped to bind the second equiv of Ag+ by the bis-enaminone sites.

300 400 5000.0

0.2

0.4

0.6

0.8

1.0

347

Ab

sorb

an

ce

Wavelength (nm)

332 nm

367

387

0 [Ag+]/[L]

5.0

(a)

0 1 2 3 4 5 6 7 8 9 10

0.00

0.05

0.10

0.15

0.20

0.25

A at 332 nm of L A at 387 nm of L A at 387 nm of 4

A

Equiv of Ag+

(b)

The Binding Sequence of the L3 The Binding Sequence of the L3

Summary

• We have designed and synthesized various hetero- and homoditopic fluorescent chemosensors for metal ions based on calix[4]arenes functionalized by 1,3-dipolar cycloaddition reactions followed by ring opening reactions.

• These hosts are found to be selective sensors for different metal ions and/or anions depending on the ligands appended. Allosteric effects were observed in the upper-rim calix[4]crown and lower-rim triazolyl anthracenes.

Acknowledgements

Dr. I-Ting Ho

Financial Support $$$National Science Council, Taiwan MOE ATU projectNational Chiao Tung University

Dr. A. Senthilvelan Dr. Kai-Chi Chang Dr. Jean-Ho Chu Mr. Kuan-Chang Haung

Lifetime measurementsProf. Eric W.-G. Diau

X-ray analysisProf. Shie-Ming PengDr. Gene-Hsiang LeeMr. Yi-Hung Liu (National Taiwan University)

Mr. Wen-Hsing WuMr. Fu-Ming YangDr. Jun Luo

Science II Building of NCTU (科學二館 )

Thanks for Your Kind Attentions