Laser spectroscopic study of estrogen and its hydrated clusters in a supersonic jet

○Fumiya Morishima, Yoshiya Inokuchi, Takayuki EbataGraduate School of Science, Hiroshima Univ.

IntroductionIntroduction

A

D

D

Fukuzawa et al. Journal of Computational Chemistry

• In estrogen receptor• In estrogen receptor

Cegelski et al. Drug Development Research

• In membrane• In membrane

estriol(E3)

A

D

estrone(E1)

A

D

β-estradiol(E2)

A

D

estrogen estrogen

B

C

B

C

B

C12

3 4 56

7

8

910

1112

1314

15

16

17

18

3

16

17

3

16

17

IntroductionIntroduction

Investigate the structure of bare estrogen and itshydrated clusters to clarify their hydrogen bonding ability.

estriol(E3)

A

D

estrone(E1)

A

D

β-estradiol(E2)

A

D

estrogen estrogen

B

C

B

C

B

C

3

16

17

3

16

17

3

16

17

Experimental and ComputationalExperimental and Computational

vacuum chamber

He gas

IR laser

UV laser

~4 atm

Supersonic jet

Vacuum chamber

heater temperatureheater temperature

estrone : ~150℃

estradiol : ~130℃

estriol : ~190℃

Electronic spectra LIF

To differentiate bands UV-UV hole-burning(HB)

Vibrational spectra IR-UV double resonance

experimentalexperimental

Quantum chemical calculation M05-2X / 6-311++G**

S1-S0 Electronic Spectra of Bare EstrogenS1-S0 Electronic Spectra of Bare Estrogen

3520035150351003505035000

band 1

band 3

band 2

band aband bband cband d

band 1band 2

a b cd

1 2

1 23 4

56

7 8 9

UV wavenumber / cm-1

LIF and UV-UV HB spectra of bare estrogenLIF and UV-UV HB spectra of bare estrogen

Number ofconformer

= 4

= 6 or more?

= 2

Each conformer of estrogen originates from orientation of OH group(s)

estrone

estradiol

estriol

3800370036003500

IR Spectra of Bare EstroneIR Spectra of Bare Estrone

UV wavenumber / cm-1

1 2

1

23657

3658

IR wavenumber / cm-1

IR spectra in the regionof OH stretching vibrationIR spectra in the regionof OH stretching vibration

cis-estrone

trans-estrone

estrone(E1)

cis-estrone

trans-estrone

3520035150351003505035000

A

DNumber ofconformer = 2

LIF

3520035150351003505035000

cis-estrone

1 2

UV wavenumber / cm-1

Assignment for cis- and trans- Conformation of EstroneAssignment for cis- and trans- Conformation of Estrone

trans-estrone

Comparison of LIF spectra of estrone and results of TD-DFT calculationComparison of LIF spectra of estrone and results of TD-DFT calculation

cis-estrone trans-estrone

3520035150351003505035000

band 1

band 3

band 2

band aband bband cband d

band 1band 2

a b cd

1 2

1 23 4

56

7 8 9

UV wavenumber / cm-1

estrone

estradiol

estriol

Assignment for cis- and trans- Conformation of EstrogenAssignment for cis- and trans- Conformation of Estrogen

cis- trans-

37503700365036003550

IR Spectra of Bare EstradiolIR Spectra of Bare Estradiol

UV wavenumber / cm-1

ab c

d

a

b

c

d

IR wavenumber / cm-1

β-estradiol(E2)

3656

3655

3657

3658

3642

3641

A-ring OHD-ring OH

A-ring OH stretching vibrations… A-ring OH stretching vibrations…

a, c show two bands b, d show only one band.

A

D

estrone(E1)

AD

3520035150351003505035000

IR spectra in the regionof OH stretching vibrationIR spectra in the regionof OH stretching vibration

Number ofconformer = 4

cis- trans-

LIF

16

17

Conformations of A-ring & D-ringConformations of A-ring & D-ring

anti: ~14cm-1 gauche(+): 0cm-1

H1 6 C 1 3 C

H

M e

O1 7 C H

1 6 C 1 3 C

H

M e

O1 7 C

gauche(-): ~73cm-1

H

1 6 C 1 3 C

H

M e

O1 7 C

• cis• trans

• anti• gauche(+)

anti gauche(+)

H1 6 C 1 3 C

H

M e

O1 7 C H

1 6 C 1 3 C

H

M e

O1 7 C

Four conformers

cistrans

antigauche(+)×

Conformations of A-ring & D-ringConformations of A-ring & D-ring

16

17

3520035150351003505035000

Assignment of D-ring OH ConformationsAssignment of D-ring OH Conformations

a b

c d

3800370036003500 3800370036003500

D-ring OH A-ring OH

IR wavenumber / cm-1

UV wavenumber / cm-1

a : antib : gauche(+) c : anti

d : gauche(+)

3656 3655

3657 3658

3642

3641

cis-anti

trans-gauche(+)

cis-gauche(+)

trans-anti

d: trans-gauche(+)b: cis-gauche(+)

a: cis-anti c: trans-anti

35050 35100 35150 35200

UV wavenum ber / cm -1

ab c

d

cis-anti

trans-gauche(+)

cis-gauche(+) trans-anti

Assignment of bare estradiolAssignment of bare estradiol

With water vapor

Without water vapor

Hydrated Cluster of Estradiol on A-ringHydrated Cluster of Estradiol on A-ring

UV wavenumber / cm-1

AB

CD

ab c

d

LIF and UV-UV HB spectraof monohydrated cluster of estradiolLIF and UV-UV HB spectraof monohydrated cluster of estradiol

β-estradiol(E2)

35200351003500034900348003470034600

band B

band C

band D

Red shift = 350 cm-1

~340 cm-1

38003700360035003400IR wavenumber / cm-1

38003700360035003400IR wavenumber / cm-1

IR Spectra of Hydrated Clusters of Estradiol on A-ringIR Spectra of Hydrated Clusters of Estradiol on A-ring

trans-anti-(H2O)

3641band C

trans-anti-(H2O)

trans-anti

ν3(H2O)

36573641

3527

3746

ν1(H2O)trans-gauche(+)

-(H2O)

band D

trans-gauche(+)

ν3(H2O)

3657

3658

35273746

ν1(H2O)

trans-gauche(+)-(H2O)

UV wavenumber / cm-134850348003475034700

C

DLIF

A

B

Each conformer forms hydrogen bondon A-ring OH as proton donor.

ν(D-ring)ν(A-ring) ν(D-ring)ν(A-ring)

** *

3520035150351003505035000

Another Isomer of Hydrated Cluster of EstradiolAnother Isomer of Hydrated Cluster of Estradiol

ab c

d

H1 6 C 1 3 C

H

M e

O1 7 C

OH

H

1 6 C 1 3 C

H

M e

O1 7 C

H

OH

Hgauche(+) typeproton donor

anti typeproton acceptor

B

C

D

Different patterns of hydrogen bond are conceivable for D-ring OH

3520035150351003505035000

37503700365036003550 37503700365036003550

IR Spectra of Bare EstriolIR Spectra of Bare Estriol

UV wavenumber / cm-1

IR wavenumber / cm-1

IR spectra in the regionof OH stretching vibration

IR spectra in the regionof OH stretching vibration

1

2

3

4

5

6

7

8

9

12

34

5

6

7 8 9

estriol(E3)

3723

37213538

3535

3722

3657

3656

3658

3663

3645

3644

3536

3655

3656

3657

3658 3664

3658

3658

3644

3644

A-ring OH

cis region

trans region

IR wavenumber / cm-1

A-ring OH

Band 1 - 6 Bare estriol

Band 7 - 9 Hydrated clusteror its vibronic band

A

Dcis- trans-

LIF

Predicted Conformers of Bare cis-EstriolPredicted Conformers of Bare cis-Estriol

Conformational nameRelative energy / cm-1

17

16

3c_16a_17a810

3c_16g-_17a208

3c_16g+_17a740

3c_16a_17g+796

3c_16g-_17g+0.00

3c_16g+_17g+466

3c_16a_17g-243

3c_16g+_17g-46.3

Intramolecularhydrogen bondIntramolecularhydrogen bond

3c_16a_17a810

3c_16g-_17a208

3c_16g+_17a740

3c_16a_17g+796

3c_16g-_17g+0.00

3c_16g+_17g+466

3c_16a_17g-243

3c_16g+_17g-46.3

Conformational nameRelative energy / cm-1

1 6 C 1 7 C

1 3 CO

1 8 C

H

H H

H1 6 C 1 7 C

1 3 C

O

1 8 C

H

HHH

1 6 C 1 7 C

1 3 C

O

1 8 C

H

H H

H

H

1 6 C 1 7 C

1 3 C

O

1 8 C

H

HH 1 6 C 1 7 C

1 3 CO

1 8 C

H

H

H

H1 6 C 1 7 C

1 3 C

O

1 8 C

H

HH

H

H

1 6 C 1 7 C

1 3 CO

1 8 C

H

H H

H

1 6 C 1 7 C

1 3 C

O

1 8 C

H

H

H

Predicted Conformers of Bare cis-EstriolPredicted Conformers of Bare cis-Estriol

380037503700365036003550

IR Spectra of Bare cis-EstriolIR Spectra of Bare cis-Estriol

3

2

1

IR wavenumber / cm-1

3520035150351003505035000UV wavenumber / cm-1

12

3estriol(E3)

3c_16g-_17g+

3c_16g-_17a

3c_16g+_17g-

3

17

16

3658

3662

3656

3657

3643

3645

Comparison of IR spectra of cis-estriol and results of frequency calculationComparison of IR spectra of cis-estriol and results of frequency calculation

3c_16g-_17g+0.00

1 6 C 1 7 C

1 3 CO

1 8 C

H

H H

H

3c_16g+_17g-46.3

1 6 C 1 7 C

1 3 C

O

1 8 C

H

HHH

3c_16g-_17a208

1 6 C 1 7 C

1 3 C

O

1 8 C

H

H H

H

Bare estriol

Form the intramolecular

hydrogen bond

between16OH and 17OH

LIFcis-

SummarySummary

estrone(E1) β-estradiol(E2) estriol(E3)

monomer

Assign the conformation of bare estrogen.

Estriol forms intramolecular hydrogen bond

1:1 hydrated cluster of estradiol

On A-ringEach conformer forms hydrogen bondon A-ring OH.

On D-ringDifferent pattern of hydrogen bond are conceivable.

Future work Hydrated clusters of estrone / estriol Methoxy estrogen

H1 6 C 1 3 C

H

M e

O1 7 C

OH

H

1 6 C 1 3 C

H

M e

O1 7 C

H

OH

H

Thank you for kindly listening to my presentationand my poor English

Relative energy of bare estroneRelative energy of bare estrone

/ cm-1

anti gauche(-)gauche(+)

Relative energy of bare estradiolRelative energy of bare estradiol

/ cm-1

H1 6 C 1 3 C

H

M e

O1 7 C H

1 6 C 1 3 C

H

M e

O1 7 C

H

1 6 C 1 3 C

H

M e

O1 7 C

35200351503510035050

* * *

3480034700

UV wavenumber / cm-1

≈

estradiol(A)-H2O

estradiol(monomer)

estradiol(D)-H2O

Relative energy of clusters

/ cm-1

3c_16g-_17g+ 0.003c_16g+_17g- 46.33c_16g-_17a 2083c_16a_17g- 243

3c_16g+_17g+ 4663c_16g+_17a 7403c_16a_17g+ 7963c_16a_17a 810

3t_16g-_17g+ 22.43t_16g+_17g- 46.33t_16g-_17a 2073t_16a_17g- 249

3t_16g+_17g+ 4643t_16g+_17a 7303t_16a_17a 796

3t_16a_17g+ 805

1 6 C 1 7 C

1 3 CO

1 8 C

H

H

H

H1 6 C 1 7 C

1 3 C

O

1 8 C

H

HH

H

H

1 6 C 1 7 C

1 3 CO

1 8 C

H

H H

H

1 6 C 1 7 C

1 3 C

O

1 8 C

H

H

H

1 6 C 1 7 C

1 3 CO

1 8 C

H

H H

H1 6 C 1 7 C

1 3 C

O

1 8 C

HH

HH

1 6 C 1 7 C

1 3 C

O

1 8 C

H

H H

H

H

1 6 C 1 7 C

1 3 C

O

1 8 C

H

HH

Relative energy of bare estriolRelative energy of bare estriol

Intramolecular hydrogen bond

16g-_17g+ 16g+_17g- 16g-_17a 16a_17g-

16g+_17g+ 16g+_17a 16a_17a 16a_17g+

/ cm-1

IR Spectra of Bare EstrogenIR Spectra of Bare Estrogen

estradiol

estrone

estriol

71238

9456

1

2

a

b

c

d

IR wavenumber / cm-1

IR spectra in the region of OH stretching vibrationIR spectra in the region of OH stretching vibration

37503700365036003550

cis-anti

trans-anti trans-gauche(+)

cis-gauche(+)

Predicted ConformersPredicted Conformers

M05-2X / 6-311++G**

~410 cm-1

~260 cm-1

Barrier between each D-ring conformation of estradiolBarrier between each D-ring conformation of estradiol

~820 cm-1

Without ZPE correction

H1 6 C 1 3 C

H

M e

O1 7 C

H

1 6 C 1 3 C

H

M e

O1 7 C

H

1 6 C 1 3 C

H

M e

O1 7 C

810 cm-1

cistrans

Rotation of A-ring OHRotation of A-ring OH

Without ZPE correction

π(HOMO)

π(HOMO)

π*(LUMO)

π*(LUMO)

cis-anti

trans-anti

HOMO and LUMO of EstradiolHOMO and LUMO of Estradiol

35200351503510035050UV wavenumber / cm -1

cis-antitrans-gauche(+)

cis-gauche(+)

trans-anti

a

bc

d

cis-anti

trans-gauche(+)

cis-gauche(+)trans-anti

Assignment of A-ring OH conformationsAssignment of A-ring OH conformations

estradiol(estradiol(DD)-H)-H22OO

Estradiol(D)-H2O

cis-antitrans-gauche(+)cis-gauche(+)

trans-anti

* * *

35200351503510035050U V wa venum be r / c m

-1

cis-anti

trans-gauche(+)cis-gauche(+)trans-anti

H1 6 C 1 3 C

H

M e

O1 7 C

OH

H

1 6 C 1 3 C

H

M e

O1 7 C

H

OH

Hgauche(+) typeproton donor

anti typeproton acceptor

red shift

blue shift

3240032000316003120030800 31800314003100030600

1-cis-naphthol

1-trans-naphthol

2-trans-naphthol

2-cis-naphthol

M05-2X / 6-311++G**

TD-DFT calculationTD-DFT calculation

352503520035150351003505035000

TD-DFT Calculation for conformers of bare estriolTD-DFT Calculation for conformers of bare estriol

UV wavenumber / cm-1

trans- 16g+_17g- 16g+_17a 16g+_17g+ 16a_17g- 16a_17a 16g-_17a 16a_17g+ 16g_17g+

Position / cm-1 35140 35141 35144 35146 35149 35150 35153 35155

Relative energy / cm-1 46 730 464 249 796 207 805 22

21

3

5

46

cis- 16g+_17g- 16g+_17a 16g+_17g+ 16a_17g- 16a_17a 16g-_17a 16a_17g+ 16g_17g+

Position / cm-1 35051 35054 35057 35058 35061 35061 35065 35067

Relative energy / cm-1 46 740 466 243 810 208 0 796

cis- trans-

Lifetime

Time / ns

Time / ns95908580757065605550

80757065605550

Monomer

Cluster

τ = 5 ns

τ = 15 ns

Predicted Conformers of Bare trans-EstriolPredicted Conformers of Bare trans-Estriol

Conformational nameRelative energy / cm-1

3

17

16

3t_16a_17a796

3t_16g-_17a207

3t_16g+_17a730

3t_16a_17g+805

3t_16g-_17g+22.4

3t_16g+_17g+464

3t_16a_17g-249

3t_16g+_17g-46.3

Predicted Conformers of Bare trans-EstriolPredicted Conformers of Bare trans-Estriol

3t_16a_17a796

3t_16g-_17a207

3t_16g+_17a730

3t_16a_17g+805

3t_16g-_17g+22.4

3t_16g+_17g+464

3t_16a_17g-249

3t_16g+_17g-46.3

Conformational nameRelative energy / cm-1

1 6 C 1 7 C

1 3 CO

1 8 C

H

H H

H1 6 C 1 7 C

1 3 C

O

1 8 C

H

HHH

1 6 C 1 7 C

1 3 C

O

1 8 C

H

H H

H

H

1 6 C 1 7 C

1 3 C

O

1 8 C

H

HH 1 6 C 1 7 C

1 3 CO

1 8 C

H

H

H

H1 6 C 1 7 C

1 3 C

O

1 8 C

H

HH

H

H

1 6 C 1 7 C

1 3 CO

1 8 C

H

H H

H

1 6 C 1 7 C

1 3 C

O

1 8 C

H

H

H

375036503550IR wavenumber / cm-1

7

1238

9456

IR spectra of Bare EstriolIR spectra of Bare Estriol

16g-_17g+

16g+_17g-

16g-_17a

16a_17g-

16g+_17g+

16g+_17a

16a_17a16a_17g+

1 6 C 1 7 C

1 3 CO

1 8 C

1 6 C 1 7 C

1 3 C

O

1 8 C

1 6 C 1 7 C

1 3 CO

1 8 C

1 6 C 1 7 C

1 3 CO

1 8 C

1 6 C 1 7 C

1 3 C

O

1 8 C

1 6 C 1 7 C

1 3 C

O

1 8 C

1 6 C 1 7 C

1 3 C

O

1 8 C

1 6 C 1 7 C

1 3 C

O

1 8 C

16g-_17g+

16g+_17g-

16g-_17a

16a_17g-

16g+_17g+

16g+_17a

16a_17a

16a_17g+

cis region

trans region

Intramolecula

rhydrogen bond

Relative Energy of Monohydrated Estriol IsomersRelative Energy of Monohydrated Estriol Isomers

3t_16g+_17g-_(A)-H2O 3t_16g+_17g-_(D)-H2O

0.00 cm-1~1,000 cm-1

More stable

Relative Energy of Monohydrated Estradiol IsomersRelative Energy of Monohydrated Estradiol Isomers

cis-gauche(+)-(A)-H2O

cis-gauche(+)-(D)-H2O

cis-anti-(D)-H2O

0.00 cm-1

~330 cm-1

~460 cm-1

More stable

cis-anti-(A)-H2O62.9cm-1