1. A Working Protocol for Assignment of AbsoluteA Working Protocol for Assignment of Absolute
Configuration from UVConfiguration from UV--Visible CircularVisible Circular DichroismDichroism
Spectra and Theoretical CalculationsSpectra and Theoretical Calculations
Andrew Pudzianowski
Computer-Aided Drug Design
Bristol-Myers Squibb PRI
June 5, 2006
2. The Problem: What is “Isomer A?”The Problem: What is “Isomer A?”
• Many racemic compounds made and tested
• Interesting compounds often separated by chiral LC:
Isomers “A” and “B”
• One enantiomer may be more biologically active than the
other (if just binary diastereomers).
• Let’s say “isomer A” is more active: Is it R or S? That is,
what is its absolute configuration?
Can’t say without further experimental work….
3. Absolute Configuration: ExperimentalAbsolute Configuration: Experimental
MethodsMethods
• Stereospecific synthesis: Definitive, but can be time-
consuming and expensive.
• Crystallography: Phase ambiguity in small-molecule
diffraction…absolute configuration not a routine result.
• Chiroptical spectroscopy: Polarized light source. CD
(absorption) or ORD (dispersion). Easy experiment, not
destructive. Spectral bands have opposite sign (+ or -)
for enantiomers. Shapes, intensities related to electronic
structure, require interpretation….
• VCD: Chiral IR or Raman spectroscopy.
• NMR: New methods.
4. Experimental Methods: ComplementaryExperimental Methods: Complementary
ApproachesApproaches
• Physical methods can be complementary: One method
can strengthen another’s results, so….
• Multiple methodologies are good!
• Role of UV-vis CD: Many attractive properties – easy to
run; strong spectral features; direct link to molecular
electronic structure – can be important part of structural
tool kit. But…
Need general methodology to interpret spectra – expect great
diversity of compound types, so no “special rules.”
Methodology must be dependable, else one could just as well
flip a coin…heads it’s R, tails it’s S!
5. CD SpectroscopyCD Spectroscopy
• What’s measured: Differential absorption of right
and left circularly polarized light passing through
a medium. Observed as…
• Circular dichroism: ∆ε = εL – εR …. Difference in
absorption coefficients. Zero in optically inactive
media. Units: L/(mol-cm). Often expressed as…
• Molar ellipticity: [Θ] ≈ 3300 ∆ε …. Related to an
angle, where units should resemble °/mol (but
don’t quote me).
• Spectrum: Record ∆ε as wavelength varies.
6. Example CD SpectrumExample CD Spectrum
Note direct relationship
between CD and regular UV
absorption: Same electronic
states and same excitations
involved. (Same transitions,
slightly different experiments.)
Simplicity of CD spectrum is
attractive.
N. Harada et al., J. Am. Chem. Soc. 109, 1661 (1987)
7. Optical Activity: It’s Helical In HereOptical Activity: It’s Helical In Here
A simplified but not simplistic view:
Chiral molecules have the symmetry properties of helices.
Classic asymmetric carbon is
equivalent to conical helix
(point group C1).
Atropisomeric molecules are
related to cylindrical helix (point
group C2)…
Helical displacement of electron
density in a transition gives rise to
differential absorption of polarized
light!
8. Hook to Molecular Structure?Hook to Molecular Structure?
Since CD spectrum reflects electronic
structure…
Can electronic structure calculationsCan electronic structure calculations
“predict” CD?“predict” CD?
Answer: Given certain limits, a big YESYES.
9. Quantum Mechanics: TransitionsQuantum Mechanics: Transitions
System in state i, one of many possible states. Mathematical
representation: Ψi (state vector, wave function)
Observable property A. Mathematical representation: operator Â.
Calculate value of A: expectation value
ii AˆA ΨΨ=
Mathematical operation – Integration for wave functions, matrix
multiplication for state vectors.
For system transition from state i to excited state j, value of A…
jiij ΨΨ= AˆA
10. QM ofQM of ChiropticalChiroptical SpectroscopySpectroscopy
Energy: Absorption of light kicks molecule from state i (often
ground so i = 0) to excited state j. Excitation occurs at energy
given below, where H is Hamiltonian for the molecule. (Usually
converted to wavelength λij and expressed in nm.)
ji
ˆE ΨΨ= Hij
ji
ˆ ΨΨ= µµij
v
Intensity: Strength of ordinary absorption is determined by electric
dipole moment µ for transition (transition dipole)…
Strength of chiroptical absorption (rotatory strength) is determined by
interaction of electric dipole with magnetic dipole moment m…
ijji
2
ˆˆˆ
2
)Im( Ψ∇×Ψ•Ψ∇Ψ=•= r
mcE
he
mR
ij
jiijij
vv
µ
11. Quantum Chemical CalculationsQuantum Chemical Calculations
Wavefunctions: States expressed as finite combinations of MO’s
with specified occupancies by electrons (configurations). These
states are used to evaluate the energies and intensities of bands
in the spectral range of interest.
Programs:
•Gaussian 03: ab initio / DFT with calculation of chiroptical properties
for UV-visible (electronic) range. Uses TD-DFT for electronic
transitions…same machinery for states and transitions as CIS.
•Jaguar: Faster ab initio / DFT for geometries and LMP2 energies.
•AMPAC: Very fast geometry optimizations with semiempirical SAM1
methodology. Good filter between conf search and above calcs.
12. UV/CD ProtocolUV/CD Protocol
DFT geometry
optimization
SAM1 geometry
optimization
Conformations
Macromodel Monte Carlo,
Merck Molecular Force Field
Jaguar 6.5 program,
B3LYP/6-31+G(d,p)
AMPAC 8.16 program:
Semiempirical SCF/MO
Conformer
energy cutoff:
≤ 3.0 kcal/mol
TD-DFT electronic
transitions
Local MP2 energy
Fixed geometry
Gaussian 03 program,
TD-B3LYP/6-31+G(d,p)
λ and R values
Jaguar 6.5 program,
LMP2/6-31+G(d,p)
Compute UV/CD bands
and plot spectrum
MULTSPEC program,
GNUPlot
13. How It’s Done: Some detailsHow It’s Done: Some details
Brief details of the UV-CD protocol….
Conformations: MacroModel MCMM with MMFF.
Refine geometries: First AMPAC/SAM1, then Jaguar at
B3LYP/6-31+G(d,p) level of theory.
Refine energies: Jaguar LMP2/6-31+G(d,p) on optimized
geometries from previous step.
Excitations: Gaussian 03 TD-B3LYP/6-31+G(d,p) with 25
excited states per conformer.
Plots: Per conformer, feed wavelengths and rotatory
strengths, along with conformer energy, to MULTSPEC
program…Boltzmann-weighted sum of Gaussian bands
for calculated spectrum.
Assign (R) or (S): Compare with experimental CD bands.
16. AA spirospiro acetalacetal troponoidtroponoid compoundcompound
TD-DFT B3LYP/6-31+G(d,p) spectrum
Note that strengths and
positions of longer wavelength
bands are off here, but pattern
is readily matched to
experimental spectrum.
N. Harada et al., J. Am. Chem. Soc. 109, 1661 (1987)
17. A closer lookA closer look……
Same with last two bands shiftedTD-DFT B3LYP/6-31+G(d,p) spectrum
If two longest-wavelength bands are shifted left by 60 nm and spectrum is
replotted (right), get a much closer match to experimental spectrum. Thus, long
wavelengths may sometimes be too long but are not qualitatively wrong.
Incidentally, a “bigger” TD-DFT calculation with the 6-311+G(d,p) basis set
doesn’t improve the results. Perhaps DFT functionals other than B3-LYP?....
18. BMS: Application to Eg5 InhibitorsBMS: Application to Eg5 Inhibitors
N
H3C
CH3
N
O
OH
CH3
CH3
*
TD-DFT CD for (R) configuration
Cpd 32: C. M. Tarby et al., Bioorg. Med. Chem.
Lett. 16, 2095-2100 (2006)…racemic
Below right: UV-CD spectrum (methanol) for
active enantiomer…
Not hard to assign (R) configuration here.
19. Another BMS Example:Another BMS Example:
Calculated CD Spectrum for (R)Calculated CD Spectrum for (R) EnantiomerEnantiomer
*
Fused ring
system
Aromatic
ring
TD-DFT B3LYP/6-31+G(d,p)
Four rotatable bonds
Below right: UV-CD spectrum (methanol)…
Not hard to assign (R) configuration here.
20. Conformer Contributions: EnergiesConformer Contributions: Energies
Note what happens to conformer energies as theory gets better…
Note: Basis set for B3LYP and MP2 entries is 6-31+G(d,p).
7.446.155.61.47
0.280.330.500.01.26
----identical to
conf 4
1.05
4.393.531.90.74
2.021.821.90.53
3.983.051.90.22
0.000.000.000.00.01
(Full MP2)LMP2 Erel
(kcal/mol)
B3LYP Erel
(kcal/mol)
SAM1 ∆Hrel
(kcal/mol)
MMFF Erel
(kcal/mol)
MCMM
conf #
21. How important are conformers?How important are conformers?
Look at individual (R) conformer spectra…Look at individual (R) conformer spectra…
Calculated CD
spectra for
isolated
conformers and
relative LMP2
energies
(Boltzmann
weights in
parentheses).
Conf 3: 2.02 kcal/mol (0.033)Conf 1: 0.0 kcal/mol (1.00)
Conf 4: 3.98 kcal/mol (0.0012)Conf 2: 0.33 kcal/mol (0.57)
22. How important are conformers? (cont.)...How important are conformers? (cont.)...
Conf 5: 4.39 kcal/mol (0.00061)
TD-DFT composite CD spectrum
While isolated confs can have drastically
different spectra, composite CD spectrum for
all 5 confs (left) looks amazingly like weighted
sum of confs 1 and 2. Boltzmann factor drops
10-fold per 1.36 kcal/mol increment above
lowest energy… confs above ~3 kcal/mol
relative energy contribute very little to overall
spectrum.
23. Summary and ConclusionsSummary and Conclusions
CD spectrum is correlated with molecular structure by established theory.
Computing resources and algorithms now provide enough power to express
theory at rather high level, so….
B3LYP and LMP2 with 6-31+G(d,p) basis set are reliable for providing
structures and conformer energies (for Boltzmann weights), and TD-DFT can
reliably calculate UV-vis CD spectra, all on a useful time scale.
Absolute configuration can be assigned from CD spectra, within some
limits….
No more than 4, perhaps 5 rotatable bonds
Correlation with experimental spectrum may not be easy if calculated
positions and strengths of bands are off…look at pattern.
CD-based assignments can’t always be made, so technique provides
additional method without superseding others.
VCD and UV-vis CD can cross-check each other, as well as other methods.
They can also point up ambiguities in experimental data and suggest
another look when stereo assignment is a priority.
24. AcknowledgmentsAcknowledgments
Bill Pitts, Al Dyckman (Discovery Chemistry)
Adrienne Tymiak (DAS), Steve Gozo (AR&D) (initial
discussions about UV-vis and VCD at BMS)
Atsu Apedo (DAS) (Experimental CD spectra)
Jack Gougoutas (AR&D), Wes Cosand (MMS) (Gaussian
03 acquisition)
Malcolm Davis, Brian Claus, Jano Jusuf (CADD) and Shibu
Nair, Brian Wong (DTCS) (SGI and Linux support)
