CONFORMATIONAL SPECIFIC SPECTROSCOPY OF JET COOLED 3-(4-HYDROXYPHENYL)-N-BENZYL-

PROPIONAMIDE (HNBPA)

ESTEBAN E. BAQUERO, V. ALVIN SHUBERT, AND TIMOTHY ZWIER Department of Chemistry, Purdue University

West Lafayette, IN 47907

N

HO

OH

N

H

O

OH

IntroductionIntroduction

Motivation

- Learn about Electronic Energy Transfer (EET)

- Test Theories of Fluorescence Resonant Energy Transfer (FRET) measurements - Conformational specific spectroscopy allows us to test chromophoric distance and orientation dependence of EET. - Dispersed fluorescence measurements can be used to measure conformational efficiency of EET.

HNBPA

Flexible

Rigid

DeMember JR, Filipescu N. J. Am.Chem. Soc. 90, 6425 (1968)

Experimental methodsResonant 2 photon ionization (R2PI): Records spectra in mass selective fashion

Biomolecule* (S1)

Biomolecule (S0)

Biomolecule+ + e-

Hol

e-bu

rn

Pro

be

Conformer A Conformer B

Hol

e-bu

rn

Pro

be

R2PI: Electronic SpectrumUV-UV Hole-burning: Conformation

specific electronic spectrum

4

3

2

1

36000359003580035700356003550035400

P-Cresol35338 cm-1

R2PI Spectrum of HNBPAR2PI Spectrum of HNBPA

UV-UV Hole-burning Spectra of HNBPAUV-UV Hole-burning Spectra of HNBPA -Two major conformers and one minor were found

14

12

10

8

6

4

2

0

Ion

Inte

nsity

(arb

itrar

y un

its)

3740037200370003680036600364003620036000358003560035400Photo Energy (cm

-1)

R2PI HNBPA

HB at 3567 cm-1

(also origin)

HB at 35517 cm-1

(origin at 35368 cm-1

)

P-Cresol 35338 cm-1 Toluene 37477.4 cm-1

B

A

HB at 35670 cm-1 (also origin)

HB at 35517 cm-1 (origin at 35368)

Minor Conformer

Laser Induced Fluorescence Spectrum of p-cresol

I Appel, K. Kleinermanns Ber Busenges. Phys. Chem. 91, 140-152 (1987)

CH3

OH

N

H

O

OH

UV-UV Hole-burning Spectra of HNBPAUV-UV Hole-burning Spectra of HNBPA

12

10

8

6

4

2

0

Ion

Inte

nsity

(arb

itrar

y un

its)

3740037200370003680036600364003620036000358003560035400Photo Energy (cm

-1)

HB at 35670 cm-1

(also origin)

HB at 35517 cm-1

(origin at 35368 cm-1

)

Origin 400 cm-1

800 cm-1

1238 cm-1

1600 cm-1

Origin440 cm

-1

800 cm-1

1270 cm-1

6a10 121

0 6a20 122

0

6a10 121

0 6a20

S2 0 00

Toluene-like origin

Photon Energy (cm-1)

A

B

UV-UV Hole-burning Origin Region

-4

-2

0

2

4

ion

inte

nsity

(ar

bitr

ary

units

)

36000359003580035700356003550035400photon energy (cm

-1)

0 28

4571 87

97

111149

202

016

38

52/54

70/74

127

137

164 1

74 18

8

31/3

4

A

B

R2PI Showing Progressions of Conformer A

0.15

0.10

0.05

0.00

-0.05

ion

inte

nsity

(ar

bitr

ary

units

)

35520355003548035460354403542035400353803536035340Photon energy (cm

-1)

0

28

45

55

71 87

97

111

128137

45 42 41

28 27

26 26 26

26

124.

5

10.54 cm10.54 cm-1-1

20.10 cm20.10 cm-1-1

31.16 cm31.16 cm-1-1

38.80 cm38.80 cm-1-1

48.74 cm48.74 cm-1-1

66.29 cm66.29 cm-1-1

113.36 cm113.36 cm-1-1

9.69 cm9.69 cm-1-1

21.38 cm21.38 cm-1-1

31.35 cm31.35 cm-1-1

37.43 cm37.43 cm-1-1

49.56 cm49.56 cm-1-1

65.74 cm65.74 cm-1-1

100.25 cm100.25 cm-1-110.60 cm10.60 cm-1-1

18.07 cm18.07 cm-1-1

23.36 cm23.36 cm-1-1

31.29 cm31.29 cm-1-1

38.02 cm38.02 cm-1-1

57.62 cm57.62 cm-1-1

70.80 cm70.80 cm-1-1 9.62 cm9.62 cm-1-1

16.32 cm16.32 cm-1-1

22.67 cm22.67 cm-1-1

31.55 cm31.55 cm-1-1

40.43 cm40.43 cm-1-1

60.05 cm60.05 cm-1-1

74.77 cm74.77 cm-1-1

Fundamentals

Conformer A

28cm-1 45 cm-1 111cm-1

Conformer B

16cm-1 34cm-1 38cm-1

So Resonant Ion-dip Infrared Spectroscopy (RIDIRS)

Biomolecule *(S1)

Biomolecule+ + e-

Biomolecule (A) NH or OH stretch

(S0, v=1)

UV Source fixed: Provides selectivity IR Source tuned

Laser Timing

50-200nsec

IRHole-burn

UVprobe

Active Baseline Subtraction

3000 3200 3400 3600 3800Wavenumbers (cm-1)

SubtractedSignal

UV only

UV +IR

Difference

IR Spectra of HNBPAIR Spectra of HNBPA

0.4

0.3

0.2

0.1

0.0

-0.1

-0.2Inte

nsi

ty (

Fra

ctio

nal

Dep

leti

on

)

370036803660364036203600Wavenumbers (cm

-1)

Conformer B

Conformer A

1.0

0.5

0.0

-0.5

Inte

nsi

ty (

Fra

ctio

nal

Dep

leti

on

)

350034903480347034603450Wavenumbers (cm

-1)

Conformer A

Conformer B

N-H Stretch Region O-H Stretch Region

Calculated Structures for HNBPACalculated Structures for HNBPA-Conformational search is done using AMBER force field in MacroModel package.

- DFT calculations are then made using Gaussian03 at the B3LYP/6-31+G* level.

0.393 kcal/mol

0.313 kcal/mol0.000 kcal/mol

0.143 kcal/mol 0.884 kcal/mol

0.882 kcal/mol0.315 kcal/mol

0.400 kcal/mol

Folded Perpendicular (FP) Extended Facing (EF) Extended Perpendicular (EP) Folded Facing (FF)

Calculated IR Hydride Stretch RegionCalculated IR Hydride Stretch Region

0.000 kcal/mol

0.313 kcal/mol

0.882 kcal/mol

300

250

200

150

100

50

0

Infra

red

Inte

nsity

(KM

/mol

e)

376037403720370036803660364036203600Frequency (cm

-1)

300

250

200

150

100

50

0

Infra

red

Inte

nsity

(KM

/mol

e)

376037403720370036803660364036203600Frequency (cm

-1)

0.315 kcal/mol

UV-UV Hole-burning Spectra of HNBPAUV-UV Hole-burning Spectra of HNBPA

12

10

8

6

4

2

0

Ion

Inte

nsity

(arb

itrar

y un

its)

3740037200370003680036600364003620036000358003560035400Photo Energy (cm

-1)

HB at 35670 cm-1

(also origin)

HB at 35517 cm-1

(origin at 35368 cm-1

)

Origin 400 cm-1

800 cm-1

1238 cm-1

1600 cm-1

Origin440 cm

-1

800 cm-1

1270 cm-1

6a10 121

0 6a20 122

0

6a10 121

0 6a20

S2 0 00

Toluene-like origin

Photon Energy (cm-1)

A

B

Summary

EvidenceEvidence Conformer AConformer A Conformer BConformer B

UV SpectrumUV Spectrum FPFP EP>EF≈FFEP>EF≈FF

IR SpectrumIR Spectrum FPFP FF>EPFF>EP

SS22 0 00000 FFFF EP>EF≈FFEP>EF≈FF

Tentative Tentative AssignmentAssignment FPFP EPEP

Folded Perpendicular (FP) Extended Facing (EF) Extended Perpendicular (EP) Folded Facing (FF)

Future WorkFuture Work

-Conformational assignments by double resonant IR-microwave spectroscopy (collaboration with Brian Dian and Brooks Pate at the University of Virginia). TA06, TA07, TA08, TI09, TI11, RI06, FC03, FC04.

- Dispersed fluorescence spectra will be taken in order to probe (EET) and its conformational dependence.

- New systems will be studied such as:

N-Z-L Serine benzyl ester

ConclusionsConclusions

- Electronic energy transfer could be tested in bichromophore systems such as HNBPA were the system’s flexibility allows the sampling of many distances and orientations between chromophores.

- HNBPA may already show evidence for conformational dependence (EET), which needs further study.

N

HO

OH

N

H

O

OH

HNBPA

Flexible

AcknowledgementsAcknowledgements

PeoplePeople Prof. Timothy S. ZwierProf. Timothy S. Zwier

The Zwier GroupThe Zwier Group Alvin Shubert Alvin Shubert Jasper ClarksonJasper Clarkson

Tracy LeGreveTracy LeGreveWilliam “Bill” JamesWilliam “Bill” JamesJaime StearnsJaime StearnsNathan PillsburyNathan PillsburyJosh NewbyJosh NewbyTalitha SelbyTalitha Selby

FundingFundingNational Science FoundationNational Science Foundation