Conformational analysis: 3JHCOC and 3JHCCC Karplus relationships for methylene 1H nuclei

Article abstract of DOI:10.1002/mrc.4264

NAMFIS (NMR Analysis of Molecular Flexibility In Solution) was applied to 1-[2-(benzyloxy)phenyl]ethanone using quantitative ¹H-¹H NOE distances and ³J proton-carbon coupling constant (CC) restraints for averaged methylene

Full text of DOI:10.1002/mrc.4264

Research article
Received: 31 January 2015
Revised: 26 April 2015
Accepted: 28 April 2015
Published online in Wiley Online Library: 27 May 2015
(wileyonlinelibrary.com) DOI 10.1002/mrc.4264
3
3
Conformational analysis: JHCOC and JHCCC
1
Karplus relationships for methylene H nuclei
Craig D. Grimmer* and Cathryn A. Slabber
NAMFIS (NMR Analysis of Molecular Flexibility In Solution) was applied to 1-[2-(benzyloxy)phenyl]ethanone using quantitative
3
3
1H-1H NOE distances and J proton-carbon coupling constant (CC) restraints for averaged methylene proton 3JHCOC and JHCCC
pathways H2-3J-X imposed by density functional theory-generated Karplus relationships. Comparison of the NOE-only versus
the NOE + CC conformational selections illustrates that the experimentally measured average 3J coupling constants of methylene
protons can be used for solution conformational analysis, potentially valuable in the study of small-molecule drugs and natural
products which lack the typically studied H1-3J-X Karplus relationships. Copyright © 2015 John Wiley & Sons, Ltd.
Keywords: NMR; 1H; 13C; conformational analysis; DFT; Karplus
carbon atom. In a molecular system where free rotation is impeded,
and where an average J CC of methylene protons can be mea-
sured, it should be possible to extract environmental information
in a manner similar to that applied to a methine proton. The diffi-
Introduction
3
The ‘Karplus equation’,[1,2] has become one of the most important
contributions to NMR spectroscopy[3] and conformational analysis,
3
describing the relationship between the vicinal J coupling con-
stant (CC) and the dihedral angle, Θ, between two spin-coupled
nuclei.[4,5] Originally applied to three-bond homonuclear coupling,
the relationship has been extended to heteronuclear coupling and
numerous examples of both homonuclear and heteronuclear rela-
tionships can be found in the literature.[6–10] Knowledge of the rela-
tionship between the CC and the dihedral angle for I ≠ 0 nuclei is a
key factor in determining molecular shape, fundamental to the un-
derstanding of the interactions between small molecules and pro-
teins and the design of new, more selective, more effective, drugs.
In conformational analysis, the application of a Karplus equation
is to translate a 3J CC measurement into a dihedral angle relation-
ship between two nuclei in an environment where ‘the dihedral
angle adopts a prevailing value over time’.[11] For protons bound to
carbon, there are three cases to consider – CH, CH2 and CH3. The
free rotation of methyl groups means that 3J coupling information
between methyl hydrogen atoms and another nucleus provides lit-
tle conformational information. Methine (CH) protons are the typi-
cal subjects,[12] offering valuable insight into molecular structure.
There is, however, the intermediate case of methylene protons,
CH2; while non-equivalent methylene protons with distinctly differ-
ent chemical shifts and coupling patterns can essentially be treated
in the same way as methine protons, provided a definitive assign-
ment can be made, it is the case of seemingly equivalent methylene
protons, exhibiting what appears as a singlet signal in the 1H NMR
spectrum, which presents an interesting problem because of the in-
ability to distinguish between the ‘magnetically equivalent’ protons
and their relationship with a third, coupled nucleus (H or X). The
magnetic equivalence is, however, only applicable to the 12C
isotopomer, in which the coupling to all potential coupling partners
(or absence thereof) is the same for both protons. In a 13C
isotopomer, the three-bond coupling between each of the two
methylene partners and a third nucleus is different, because the di-
hedral angles between those methylene protons and the coupled
partner are different, by the Karplus equation, arising from the
roughly 108° separation of the protons on the sp3-hybridised
culty in extracting the structural information lies in that while in
any individual conformer of a particular geometry each of the two
methylene protons sits in a unique isotopomeric coupling environ-
ment, the spectroscopic measurement can only observe a confor-
mational average.
Computational chemistry software packages like Gaussian[13,14]
possess the ability to calculate CCs given a fixed geometry, usually
the result of another calculation. Typically, the calculation of NMR
parameters follows a geometry optimisation. This calculation repre-
sents only one conformation and while it is hoped that this is the
dominant and energetically most favourable structure (a global
minimum), experimental evidence in an NMR spectrum often indi-
cates that this is not the case and that there may be a number of
conformations present.[15]
To investigate the possibility of using average 3J CCs of methylene
protons in conformational analysis, we have combined a computa-
3
tionally derived assessment of the individual J coupling relation-
3
ships with experimentally determined average J values, obtained
from an EXSIDE[16] experiment, and quantitative NOE-derived intra-
molecular 1H-1H distances and used these parameters to select
those conformations likely to represent the solution speciation.
Methods
Synthesis (SI)
1-[2-(Benzyloxy)phenyl]ethanone was prepared by condensation of
2-hydroxyacetophenone with benzyl bromide in refluxing acetone
in the presence of anhydrous potassium carbonate.[17–19]
*
Correspondence to: Craig D. Grimmer, School of Chemistry and Physics,
University of KwaZulu-Natal, Pietermaritzburg, South Africa. E-mail: craig.
grimmer@gmail.com
School of Chemistry and Physics, University of KwaZulu-Natal, Pietermaritzburg,
South Africa
Magn. Reson. Chem. 2015, 53, 590–595
Copyright © 2015 John Wiley & Sons, Ltd.

3JHCOC and 3JHCCC relationships for methylene 1H nuclei
NMR data (SI)
Experiments were performed on a Bruker Avance-III 500 spec-
trometer at 30°C. The sample for the quantitative NOE experi-
ments (10mg in 600μl) was degassed and sealed to remove
dissolved oxygen and prevent moisture absorption during the
experiments. Quantitative NOESY experiments were performed
with mixing times in the 50–300ms range, and internuclear
distances were calculated from cross-peak volumes at a mixing
time consistent with linear behaviour,[20–28] using the isolated-
spin-pair-approximation.[29] A customised hard-X-pulse EXSIDE-
3
type experiment,[16,30] was used to measure J CCs with selective
excitation at the required 1H frequency using a shaped pulse
(180° Gaussian Cascade Q3). Shaped pulses were tested for selec-
tive excitation with a customised 1-D DPFGSE experiment.[31]
Conformers (SI)
A conformational search was performed using Schrodinger’s
MacroModel[32,33] software producing 1185 structures. To this was
added one conformation determined by single crystal X-ray
diffraction.[34]
Figure 1. Relevant atom numbers from the Gaussian and MacroModel
atom number maps, arrowed NOE contacts between 1H nuclei, and
internuclear distances (Å).
DFT calculations (SI)
Calculation of CCs was performed using the Gaussian-09 W
(C.01)[13,14] suite of programs at the MPW1PW91[35,36] level with
the 6–31 + g(d,p) basis set using the ‘mixed’ method[37,38] for the
relevant nuclei in restricted geometries. A 3J versus dihedral angle
(Θ) relationship was established using the optimum Karplus-type
relationship, the parameters for which were determined using the
Origin software package.[39]
NAMFIS (SI)
NMR Analysis of Molecular Flexibility In Solution (NAMFIS)[15,40] is a
means by which the individual contributing conformations of a
small flexible molecule can be extracted from an average NMR
spectrum. A set of possible conformations is generated, and these
are compared with the experimentally determined proton-proton
distances, from quantitative Nuclear Overhauser Enhancement
measurements (NOE), and/or dihedral angles, from 3J CC measure-
ments. The result is a best-fit set of conformers, characterised by a
sum-of-squared-differences (SSD) value, each associated with a
mole fraction of the total population.
Figure 2. HSQCETGPLRJC spectrum of A2OB in DMSO-d6.
•
The aryl-O-CH2-aryl fragment occurs in 126 small molecule
structures found in the Protein Data Bank,[41] more than 1700
structures in the Cambridge Structural Database,[42] and
forms a component of a novel range of COX-2 inhibitors;[43]
The aceto-substituent impedes internal movement of the
structure and creates the possibility that the dihedral angle
adopts a prevailing value over time.[11]
Results and discussion
While ostensibly simple, 1-[2-(benzyloxy)phenyl]ethanone, abbr
A2OB, (Figs 1 and 2) presents a number of attractive features:
•
1
•
•
The methylene signal in the H NMR spectrum is a singlet,
clearly separated from all other signals, facilitating clean selec-
tive irradiation for the EXSIDE[16] experiment;
From the NOESY spectrum of A2OB, there are five structurally sig-
nificant NOE contacts (Fig. 1), together with the reference distance
of 2.5Å.
The methylene moiety is part of the bridge between two aro-
matic rings, with 3 J coupling pathways to both rings, poten-
tially offering structural insight into the spacial relationship
between the bridge and both rings;
There are four structurally significant 3J coupling relationships for
the bridge between the aromatic rings, shown for the relevant
isotopomers in Table 1.
All 1H signals can be assigned without ambiguity, leading to
unambiguous NOE correlations;
Using the HSQCETGPLRJC[16,30] experiment, the average long-
range CCs of the methylene protons have been determined. The
spectrum is shown in Fig. 2, providing the average 3J CCs between
the methylene protons involved in the three-bond-to-carbon con-
•
•
The single methylene fragment presents only four 3 J coupling
pathways, computationally manageable at a reasonable level
of theory;
2
tacts in the aromatic rings, as well as the average J CC between
Magn. Reson. Chem. 2015, 53, 590–595
Copyright © 2015 John Wiley & Sons, Ltd.
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C. D. Grimmer and C. A. Slabber
Table 1. Isotopomeric 3 J methylene coupling pathways in A2OB
Table 2. Values of Karplus equation parameters (A, B, C and D) for
Equation (1)
Pathway/
constant (25-11-10-2) (26-11-10-2) (25-11-12-13)a (26-11-12-13)a
3JHCOC
3JHCOC
3JHCCC
3JHCCC
A
B
C
D
8.4 0.1
8.4 0.1
4.0 0.2
ꢀ0.9 0.1
1.9 0.1
3.7 0.1
ꢀ0.8 0.05
2.3 0.05
ꢀ1.7 0.1
ꢀ1.6 0.1
ꢀ0.1 0.07 ꢀ0.1 0.06
Atom pathways
Atom pathways
ꢀ3.0 0.3
2.8 0.3
14.6 0.7
ꢀ12.0 0.9
(numbers from Fig. 1):
(numbers from Fig. 1):
aSimilar values obtained for pathways 25-11-12-14, 26-11-12-14;
25-11-10-2 (3JHCOC
26-11-10-2 (3JHCOC
)
)
25-11-12-13/14 (3JHCCC
)
isotopomer with a 13C atom located at atom position 14 (SI).
26-11-12-13/14 (3JHCCC
)
+108° (in terms of 25-11-12-13) for average pathway 25/26-11-12-
13. Using the NOE and average 3 J CC constraints, with the density
functional theory (DFT)-derived Karplus relationships, the NAMFIS
scripts make a best fit selection of from a set of conformational can-
didates (SI). The outcome of the process for the NOE-based and
NOE + CC-based selections is shown in Table 3; the selected con-
formers and their relative populations (SI). Common conformers
are highlighted in bold text (220, 711).
The SSD for each selection is zero and, ironically, disappointing
because this provides no immediate guide as to the comparison
of the sets. From a relatively large number of conformers, it is easy
for NAMFIS to provide an excellent match. Even with this wide selec-
tion (1185 + 1), in which we could potentially find a completely dif-
ferent conformational mixture for each of the two sets, it is pleasing
to note that there is a significant degree of cross-over (33–40%), in
the selection of conformers 220 and 711. The X-ray conformation is
not selected despite reports of other NAMFIS studies indicating the
presence of the solid-state structure amongst the solution
conformations.[49–52]
the methylene protons and the quaternary carbon of the terminal
phenyl ring. The spectral region encompassed by the singlet at
5.24 ppm in the 1H spectrum corresponds to a number of methy-
lene signals, that for the all-12C isotopomer, and, masked by this,
the methylene proton signals of the various 13C isotopomers –
these are the species that provide all of the 13C information. By
irradiating the region of the 12C isotopomer, one simultaneously ir-
radiates the signals of the 13C isotopomers, even though those
signals are masked (excluded from this are the 1J 1H-13C isotopomer
signals, visible in the 1H spectrum as 13C satellites).[44]
From the splitting of the signals in the indirect dimension, the ab-
3
3
solute values of the J CCs can be determined as JHCOC = 2.65 Hz
and 3JHCCC = 4.11 Hz.
Using the results of the CC calculations from Gaussian,[13,14] for
23 restricted, optimised geometries, the 3 J versus dihedral angle re-
lationships between the methylene protons (atoms 25 and 26 in
Fig. 1) and the appropriate carbon atoms for the 13C isotopomers
(atoms 2 and 13 in Fig. 1), have been determined (Figs 3 and 3a-
SI). The isotopomer with a 13C atom located at atom position 14 is
the same as that with a 13C atom at position 13.
The dihedral angles for the chosen conformers of the NOE-based
selection (Fig. 4) are shown in Table 4, with the calculated average
3 J CCs according to the DFT-Karplus equations.
Different forms of the Karplus equation can be found in the
literature.[5,45,46] The curves presented in Figs 3 and 3a-SI have been
plotted according to the relationship shown in Eqn (1),[47] for a
phase-shifted form[48] of the equation, and the values for the con-
stants are presented in Table 2.
Keeping in mind that the selection in the table is based solely
upon six NOE-distances, there is a pattern in the conformers that
3
have been chosen by NAMFIS – conformers that present JHCOC
values of around 6 Hz, 85% of the population, despite a variety of
25-11-10-2 angles. Those conformers with 25-11-10-2 angles in
the range ꢀ161° 12° represent 60% of the population. The obvi-
ous ‘odd man out’ is 711 (14%), with a 25-11-10-2 angle of ꢀ68°
and a 3JHCOC value of 1 Hz, the closest (albeit poor) match to the ex-
perimentally measured average 3JHCOC CC (2.6Hz).
3J ¼ A: cos2ðΘ þDÞ þ B: cosðΘ þDÞ þ C
(1)
The plots illustrate that dihedral angles delivering appropriate
average 3 J CCs are ꢀ92°and ꢀ24° (in terms of angle 25-11-10-2)
for the average pathway 25/26-10-11-2, and ꢀ164°, ꢀ77°, +9° and
There is little common ground amongst the 25-11-12-13 dihedral
angles although there is a strong presence of conformers presenting
3JHCCC values around 4 Hz (220, 1020, 763, 746 and 760), some 60% of
3
the mixture, with selections presenting JHCCC values around 5 Hz
(711, 1121, 748, 1137 and 1146) making up the remainder of the mix-
ture. Conformer 763 (13%) represents the closest individual match
to the experimentally measured average 3JHCCC CC (4.3 vs 4.1 Hz).
While the focus of the analysis is the link between the two rings,
the relative position of the acetyl group cannot be ignored.
Governed by three distance constraints (Fig. 1), most of the selected
conformers have the methyl hydrogen atoms turned inward (70%),
towards the ether moiety (Fig. 4a-SI). The obvious misfits, based on
a graphical inspection, are conformers 711, 746, 760 and 1146
(30%) with the acetyl group turned carbonyl-inwards or side-on
to the benzyl ether (Fig. 4b-SI).
3
The dihedral angles for the chosen conformers of the NOE + CC-
based selection (Fig. 5) are shown in Table 5, with the calculated
average 3 J CCs according to the DFT-Karplus equations.
Figure 3. JHCOC for pathways 25/26-10-11-2 and average 3JHCOC, plotted as
a function of dihedral angle 25-11-10-2. The offset between 25 and 26 in
relation to 2 is 116i° 5° over 23 geometries.
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3JHCOC and 3JHCCC relationships for methylene 1H nuclei
Table 3. NMR Analysis of Molecular Flexibility in Solution conformational selections
A2OB (NOE)
A2OB (NOE + CC) [13C iso-13]a
A2OB (NOE + CC) [13C iso-14]a
Number of candidate conformers
Number of NOE, J constraints
Number of selected conformers
Conformer, (%)
1185 + 1 (XRD)
6
1185 + 1 (XRD)
6, 2
1185 + 1 (XRD)
6, 2
10
8
8
220 (18.6)
1020 (16.1)
711 (14.5)
1121 (13.6)
763 (13.5)
746 (9.6)
748 (4.7)
1137 (3.7)
1146 (3.4)
760 (2.0)
0
711 (31.9)
499 (20.2)
1156 (18.0)
347 (11.7)
220 (8.0)
1159 (7.3)
1117 (1.5)
1120 (1.4)
—
711 (33.9)
499 (19.7)
1156 (19.2)
1159 (9.0)
347 (8.6)
220 (7.0)
1117 (1.5)
1120 (1.3)
—
—
—
SSD
0
0
CC, coupling constant; XRD, X-ray diffraction; SSD, sum of squared differences; NAMFIS, NMR Analysis of Molecular Flexibility in Solution.
aNAMFIS analysis of the isopomerically identical 13C-at-position-13 and 13C-at-position-14 delivers the same selection of conformations, in the same order,
up to the 70% level; thereafter, the conformers selected are the same with small differences in the population levels.
Figure 4. NMR Analysis of Molecular Flexibility in Solution conformational
Figure 5. NMR Analysis of Molecular Flexibility in Solution conformational
selection (NOE-based), ten conformers.
selection (NOE + coupling constant-based), eight conformers.
Table 4. NMR Analysis of Molecular Flexibility in Solution conformational selection, NOE-based (SI)
Conformer (%)
Angle 25–2 (°)
Angle 26–2 (°)
3JHCOC (Hz)
Angle 25–13 (°)
Angle 26–13 (°)
3JHCCC (Hz)
220 (18.6)
1020 (16.1)
711 (14.5)
1121 (13.6)
763 (13.5)
746 (9.6)
ꢀ166.1
ꢀ151.9
ꢀ68.1
ꢀ51.3
ꢀ36.3
+52.0
6.1
6.4
1.0
6.4
6.3
6.3
5.7
6.3
6.3
6.3
ꢀ72.0
ꢀ86.6
ꢀ13.8
+176.9
+109.8
ꢀ156.9
+128.6
ꢀ56.8
ꢀ52.0
+13.7
+170.6
+157.8
ꢀ133.2
+60.7
4.4
3.7
5.2
5.0
4.3
3.7
5.2
5.1
5.2
3.7
+40.2
+156.4
ꢀ36.5
+158.9
ꢀ59.5
ꢀ45.4
ꢀ46.3
+154.6
ꢀ150.9
+43.7
ꢀ5.4
+87.9
748 (4.7)
ꢀ173.1
ꢀ161.7
ꢀ163.4
+39.6
+6.8
1137 (3.7)
1146 (3.4)
760 (2.0)
ꢀ173.0
ꢀ168.7
ꢀ101.5
Applying the NOE-distance and the CC constraints, NAMFIS se-
lects a somewhat different set of conformations. Conformers 711,
347 and 1159 have 25-11-10-2 angles in the region ꢀ61° 7° and
3JHCOC values of around 1 Hz (50% of the population). Excluding
the conformers at < 2% level, the remainder of the selection is
made up of three conformers, 499, 1156 and 220. These three
conformers have completely different 25-11-10-2 angles but
conformers 1156 and 220 (26% of the population) present 3JHCOC
Magn. Reson. Chem. 2015, 53, 590–595
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C. D. Grimmer and C. A. Slabber
Table 5. NMR Analysis of Molecular Flexibility in Solution conformational selection, NOE + coupling constant-based
Conformer (%)
Angle 25–2 (°)
Angle 26–2 (°)
3JHCOC (Hz)
Angle 25–13 (°)
Angle 26–13 (°)
3JHCCC (Hz)
3 J constraint
711 (31.9)
499 (20.2)
1156 (18.0)
347 (11.7)
220 (8.0)
—
—
2.6
1.0
2.3
6.3
1.0
6.1
1.0
2.0
1.4
—
—
4.1
5.2
3.8
3.0
2.5
4.4
5.6
2.9
3.2
ꢀ68.1
ꢀ89.4
+37.2
ꢀ54.6
ꢀ166.1
ꢀ58.9
ꢀ82.5
ꢀ75.6
+52.0
+28.4
+152.3
+63.2
ꢀ51.3
+59.6
+32.7
+43.1
ꢀ13.8
ꢀ85.2
ꢀ139.1
+73.3
ꢀ133.2
+158.7
+105.4
ꢀ42.9
+170.6
+34.2
ꢀ72.0
+151.3
ꢀ109.7
ꢀ98.9
1159 (7.3)
1117 (1.5)
1120 (1.4)
+135.0
+144.8
Table 6. Dihedral angles and density functional theory-calculated average 3J coupling constants for conformers X-ray diffraction, 499, 711 and 821
Conformer
Angle 25–2 (°)
Angle 26–2 (°)
3JHCOC (Hz)
Angle 25–13 (°)
Angle 26–13 (°)
3JHCCC (Hz)
3J constraints
—
—
2.6
0.9
2.3
1.0
0.9
—
—
4.1
5.6
3.8
5.2
5.6
XRD
499
711
821
ꢀ58.4
ꢀ89.4
ꢀ68.1
ꢀ59.5
+61.2
+28.4
+52.0
+59.5
ꢀ33.3
ꢀ85.2
ꢀ13.8
+149.0
ꢀ152.9
+158.7
ꢀ133.2
+31.8
XRD, X-ray diffraction.
Consideration has been given to the HCOC and HCCC bond
angles for pathways 25/26-11-10-2 and 25/26-11-12-13, and the re-
sultant average 3J CCs that arise from the conformation of A2OB as
determined by X-ray diffraction,[34] based on the proposal that this
conformation may be present in solution,[49–52] although this is not
always the case.[47] There is an excellent match for the crystal struc-
ture in the conformer pool, conformer 821 (Fig. 5c-SI), but the crys-
tal structure (XRD) has been added anyway.
The values for the angles of the pathways and the calculated
3J CCs for conformer XRD are presented in Table 6 – dihedral
angles for pathways 25/26-11-10-2 of ꢀ60° and +60°, respec-
3
tively, with a JHCOC value of 0.9Hz, dihedral angles for path-
ways 25/26-11-10-13 of ꢀ30° and ꢀ150°, respectively, and a
3JHCCC value of 5.6Hz. Amongst the common conformers, con-
former 711 provides the closest match within the NOE-based
(14%) and the NOE + CC-based (32%) selections.
The best individual match for the CC constraints is conformer
Figure 6. NMR Analysis of Molecular Flexibility in Solution conformers 499
(light), 711 (medium) and X-ray diffraction (dark).
3
3
499, with JHCOC of 2.3Hz and JHCCC of 3.8 Hz and angles 25-11-
10-2 of ꢀ89° and 25-11-12-13 of ꢀ85° (Table 6). While conformer
XRD is not chosen by NAMFIS, the root mean square deviation on
an ‘all atoms’ basis between this conformation and conformer 499
is 0.9, and for conformer 711, 1.0[53] (Fig. 6).
Despite the similar root mean square deviation values, it is clear
from the graphical comparison (Fig. 6a-SI) that conformer 499, from
the application of CC constraints, is a better match for the crystal
structure.
values of around 6 Hz, consistent with the majority of the NOE-
based selection. Conformer 499 (20%), with a 25-11-10-2 angle of
3
ꢀ89°, presents the closest JHCOC value (2.3Hz) to the constraint
(2.6 Hz).
There is a less commonality amongst the NOE +CC conformers in
terms of the 25-11-12-13 angle and the 3JHCCC values. Notably, the
‘odd man out’ from the NOE selection, conformer 711, is the top
selection, fitting neatly into the NOE+ CC-based selection, with
3JHCOC of 1.0 Hz and 3JHCCC of 5.2 Hz, at a substantial level of 32%.
3
Conformer 499 (20%) presents the closest match to the JHCCC
Conclusion
constraint, followed closely by conformer 220 (8%), common to
both selections.
With regard to the acetyl substituent, conformers with the
methyl hydrogen atoms turned inward (Fig. 5a-SI) dominate the se-
lection (98%). The obvious misfit (Fig. 5b-SI), with the acetyl group
turned carbonyl-inwards, is conformer 1117 (2%).
NAMFIS has been applied to a key conformational element of 1-[2-
(benzyloxy)phenyl]ethanone using 1H-1H distances and 1H-1H dis-
tances combined with DFT-calculated average coupling constants
for H2-3 J-X pathways. Good agreement between the distance-only
and the distance-CC conformational selections indicates that the
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Copyright © 2015 John Wiley & Sons, Ltd.
Magn. Reson. Chem. 2015, 53, 590–595

3JHCOC and 3JHCCC relationships for methylene 1H nuclei
theoretical treatment of average 3 J versus Θ relationships for meth-
ylene protons, combined with measured average 3 J coupling con-
stants, is a potentially useful means of providing structural insight in
cases of restricted movement of methylene groups.
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