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The microwave spectrum of a vitamin: Nicotinamide
JOURNAL
OF MOLECULAR
SPECTROSCOPY
145, 1-t 1 ( 199 1)
The Microwave Spectrum of a Vitamin: Nicotinamide BEAT VOGELSANGER,RONALD D. BROWN, PETER D. GODFREY, AND ANTHONY P. PIERLOT Centre,for High-Resolution Spectroscopy and Opto-Electronic Technology, Monash Universit?: Clayton, Victoria 3168, .4ustralia We have observed the microwave spectrum of nicotinamide and three deuterated isotopomers in the range 57-78 GHz, identifying four species that we conclude are the ground and vibrationally excited E-conformer and a split ground state of the Z-conformer. Some hyperfme splitting patterns were observed and dipole moments of the two conformers were determined. A detailed ab initio molecular-orbital investigation was also undertaken, involving complete geometry optimization at the 6-3 1 G level of basis functions. The extent of torsional twist of the amide groupin the two conformers was in good agreement with the theoretical calculations and similar satisfactory agreement was found between observed quadrupole hyperfme patterns and patterns computed from theoretically derived coupling constants. The observed relative energies of E- and Z -nicotinamide are qualitatively in agreement with predictions of the molecular-orbital calculations but the separation is numerically greater. 0 1991 Academic Press. Inc
INTRODUCTION
The shape of the nicotinamide molecule (Fig. 1) has been the subject of much interest, particularly with respect to the conformation of the amide group relative to the pyridine ring. The X-ray structure of the crystal was reported over 30 years ago ( I), the conclusion being that the oxygen of the amide was approximately entgegen to the pyridine nitrogen but twisted out of the plane of the pyridine ring by rotation around the C-C bond by about 156” from the zusammen position. The conformation of nicotinamide was further studied via dipole moment measurements in benzene solution by Kuthan et al. (2), following earlier measurements of Purcell and coworker ( 3, 4). They obtained a moment intermediate between values estimated, by several lines of calculation, for the Z- and E-conformations, concluding that two different rotamers were present but finding a puzzling temperature dependence of the dipole moment. Theoretical studies by Bijhm and Kuthan (5)) in which the torsional angle around the C-C bond was optimized using an ab initio STO-3 G basis molecular orbital calculation, the other geometrical parameters being constrained to CNDO/2 values, indicated that both Z- and E-conformers are nonplanar. A more complete treatment by Iaconis et al. (6) in which all parameters apart from the pyridine ring were optimized, the pyridine ring parameters being held at the values derived from a separate geometry optimization for pyridine itself, all done using the STO-3 G basis, predicted twist angles of 24 and 158”, respectively, around the C-C bond, the Econformer being more stable by about 1 kJ mole-’ . Other SCF-CI calculations by El1
0022-2852/91 $3.00 Copyright 0 All n&s
1991 by Academic Press, Inc.
of reproduction m any form resewed.
VOGELSANGER
2
ET AL.
Z-nicotinamide
E-nicotinamide FIG. 1. Structure
of the two conformers
of nicotinamide
with labeling of the nuclei.
Shahawy et a/. (7) predicted the Z-conformer to be the more stable. Thus there is some disagreement between different theoretical approximations, while experimental evidence is complicated by the unknown effects of solvent or the crystal lattice. TABLE I Observed
Transitions
(MHz)
E-nlcotinamik vibr. ground state TRANSlTlON 880-771 881.770 981-872 982-871 990.88, 991.880 IO 7 3. IO 7 4. IO 9 I IO 9 2. IO IO 0 IO 10 I II 74-10 II 7 S-10 II 8 3-10 I1 84-1073 II 9 2-10 II 9 3-10 I1 IO 1 II 10 2 13 6 7 13 6 8 14 6 8 14 6 9 IS 5 IO 16 4 I2 16 5 II 16 7 9 I6 7 10 II 5 13 17 7 IO 17 7 II 18 5 14 20 5 16 27 3 24 28 3 2S 28 4 25 29 2 27 29 3 26 31 031 13 II 2 14 II 3 IS II 4 16 II 5 17 II 6 23 II 12 -
9 9 9 9 9 9
10 10 I2 I2 13 13 14 IS 15 I5 IS 16 16 16 17 19 26 27 27 28 28 30 13 14 15 16 17 23
OBS. FREQ.
6 6 8 8 9 9 6 6 7
4 3 2 I L 0 5 4 4
8 8 9 9 5 5 5 5 4 3 4 6 6 4 6 6 4 4 4 4 3 3 4 i 10 IO IO 10 IO IO
3 2 2 I 8 7 9 8 II I3 12 IO 9 I2 I, IO 13 I5 23 24 24 26 25 30 3 4 5 6 7 I3
DC
of the Four Different E-nrotinamide wbr. excited safe OBS. FREQ.
W
Species of Nicotinamide 2.n,mtinamlde state I
Z-N~tiMllllde state I,
OBS. FREQ.
W
0.004 0.004 0.004 -0.004 -0.012 -0012 -0.060 a038 0.03 I 0.03 I 0.063 0.063 -0.05 I 0.03s 0.002 0.003 -0.033 -0.033 -0.070 -0.070 0.068 -0.01 I 0.045 0.061 0.077
-0.002 0.070 -0.101
60663.387 71195.079 71181.289
a.c40 0041 -0.1 is
0.074 -0.038
73332.154 73302.354
0.055 -0.041
59392.030
-0.003
58613.758 58592.622 58567.036
0.018 -0.0 I 3 0 035
59424.675 59424.675 61613.359 61613.359 67201.796 67201.796 58208.690 58208.690 69390.569 69390X9 76978 R42 7*971 Ro? 6039md3 60390.843 65987.61 I 65987.61 I 71578.696 71578.696 77167.626 71167.626 59104.767 59073.329 61256.483 61186.576 58245.412
o.w3 0.003 o.cm7 0.007 -0.w3 -0.003 -0.044 -0.019 0.016 0.016 0.011 0.01 I -0.047 0.053 0.019 0.020 OSKW 0.009 -0.533 -0.003 0.059 -0.041 0.007 -0.023 0.W
59415.983 59415.983 61603.990 61603.990 67191.977 6,191 477 S8199.247 58199.247 69380.081 69380.081 74967 950 74967.950 60380.948 60380.948 63976.961 65976.961 71567.479 71567.479 77155.921 77155.921 59096.308 59068.400 61247.954 61185.815
-0.023 -0.023 0.018 0.018
60766S95 71207.029 71191.001 59056.320 73342.695 73307.957
-0.070 0.063 -0.049 -0.029 0.060 -0.031
60672.238 71198.960 71184.981 73335.797 73305.648
57254.013 59265.631 59783.689 59453.105 61232.913 59500.625
57299.218 59322.836
-0.035 0.037
58679.374
58623.184 58600.988
-0.037 -0.019
58638.674 58621.476 58W.234
-0.028 0.028 -0.004
58541.617
0.008
58543.616
-0.004
58595.827 58235.692
-0.045 0.029
W.-
594 10.468 594 10.468 61598.452 61598.452 67185.71 I 67185.71, 58194SlS 58194.515 69373.815 69373.815 749643.945 7496Q.945 60376.326 60376.326 65971.483 6597 I.483 71561.223 71561.223 77148.911 77148.911 59092.970 5906S.351 61244.781 61183.357 58181 288
59461.932 59461.932 61650.160 61650.160 67244.076 67244.076 58239.458 58239.458 69432.376 69432.376 75026.129 75026.129 60420.95 1 60420.95 1 66023.375 66023.375 71620.017 71620.017 77214.445 77214.445 59127.040 59093.794 61277.978 61203.975 58286.403 60855.183 60325.551 71231.893 71214.747 58964.166 73366.106 13328.944 mw82:;;;
” , ,
OBS. FREQ.
:g;: -0.018 0.004 0.04 I 0.041
__.._._
t:::: -0.053 0.034 “-012
MICROWAVE
SPECTRUM
OF NICOTINAMIDE
3
In view of the biological significance of nicotinamide it was felt desirable to attempt to establish unambiguously the conformers of this molecule when not affected by environmental influences. Accordingly, we have investigated its microwave spectrum in the gas phase and, because the theoretical studies quoted above do not seem satisfactory by contemporary standards, we have made further ab initio molecular-orbital calculations that have included a more thoroughgoing geometrical optimization and have employed a larger basis set. EXPERIMENTAL
DETAILS
A millimeter-wave spectrometer, based on observed absorption in a supersonic expanding plume of nicotinamide vapor entrained in argon and described previously (8 ), was used for the measurements. The source was a phase-locked millimeter-wave klystron or a K-band klystron with tripler. Stark modulation at voltages up to 6000 V were employed at a plate separation of 3.5 cm. Accurate line frequencies were obtained by repetitive scans using a data-collection system based on a VAX 1 1 / 780 computer. Measurements were made in the frequency range 57-78 GHz. Commercially available nicotinamide was used in a volatilization tube, heated to 160- 17O”C, and a stream of argon at pressures of about 300 hPa was bubbled through the melted sample before being expanded through a nozzle 500 pm in diameter. The liquid evaporated at rates typically of about 0.3 g hr-’ . We were unable to detect any decomposition of the nicotinamide under these conditions. Nicotinamide-N-d1 and nicotinamide- N, N-d2 were generated by dissolving nicotinamide in D20/H20 1: 1. After 24 hr at 40°C. the water was removed in a “Ro-
60360
60390
60410
60400
Frequency
60420
/MHz
FIG. 2. Typical strong rotational transitions [ 11 7,4- 106,5/ 117,5- 106,4] for the four different species of nicotinamide. The transition lines correspond to (from left to right) state II of the Z- , state I of the Z- , vibr. excited state of the E- , and vibr. ground state of the E-conformer. The integration time to record this spectrum was 100 min. (6 scans averaged).
VOGELSANGER
ET AL.
TABLE II
Calculated and Observed Spectroscopic Parameters. Geometric Parameters. and Dipole Moment Components” of Nicotinamide E-nicotinamide vibr. ground state
E-niwtinamide vibr. excaed state
3954.5
3891.2262(211
3888.7292(29
3953.0
3888.l8OOf39)
3887.8089(44)
1242.6
1236.2442(16)
1234.7691(18
1238.0
1231.413(25)
1230.893(27)
953.5
944.7920(141
946.9075(2
957.3
949.855(29)
950.462(42)
0.0361(12)
0.0262(
0.0458~31)
0.0822(66:
0.219~22)
0.334(14)
0.329(20)
0.320(29)
E-lWXiMlll1& talc. 6.3lG
-0.00.547(73) -0.00375(30)
-4.497
(reference ‘Statntically
I
Z-nlcomwnide stale
13:
-0.065~131
0”
-0.S I6(5 1,
0”
24
32
33
26
-3.768
-5.535
1
-8.174
Z-mcotmarmde
state II
0.0224(221 0.111(10) 0.336(30) 0”
0’
0’
39
46
23
24
-8.325
-8.851
194
I84
I85
197
190
191
194
I86
I89
I89
177
180
46
74
63
61
88
81
336
335
334
338
338
338
I05
105
106
100
95
96
I8
28
26
25
38
35
0.75
0.315(2V
I.49
l.l5(2)Y
I.15
WFrom Stark-effect
Z-oicotinamide talc. 6.3lG
0.316(2P
0.67
0.209(2)Y
0.200(4)Y
5.98
CO.2
0.96
I .88
l.l58(7lY
l.O5(4)Y
6.06
4.5(4)2
4.5(4)7
2.02
1.20(l)
1.09(5)
6.09
4.5(4)
4.5(4)
measurements.
Tlx field was cahbrated wth the 413-404 tnnsltion
of S@ usmg the dipole moment oh = 1.63305(4)D
IO). not signliicant pammeler
YFrom rnea~uremenf~ of Individual ‘From measuremen&
(F-test.
99%
Stark lobes.
confidence mterval: see reference I I ). Numbers m parenlhesls represent the standard daubon
of unresolved clumps of Stark lobes for two different transmons.
of the fit only.
Numbers m parsthea
are an esbmate of the error.
tavap,” providing a mixture of nicotinamidedo/d1 /d:. Both possible monosubstitutions on the amide nitrogen were obtained in sufficient quantities for the spectra to be observed and analyzed. A sample containing more than 90% nicotinamideN, Nd2 was obtained by repeating the above procedure twice with DzO instead of DzO/ H20. RESULTS
The observed spectrum in the vicinity of 60 GHz is ph-type, dominated by R-branch series: .!7,(5-6) (J 1 )6.~J-7~/J7,~J-7~- (J - 1 )6,c.-6) with J = 10, 11. 12, - * - and &(5_7) (J 1)7.~J-~~/J8,~J-8~ - (.I - 1 )7.(~-7j with J = 8, 9, - - - . This pattern of lines, the spacings between two transitions in each of these series being about 2.2 GHz, was recognizably similar to a theoretical spectrum generated by using geometries of nicotinamide that were predicted by fully optimized ab initio molecular-orbital calculations using a 6-3 1 G basis and the Gaussian 88 code ( 9). All other transitions in this range are much weaker. It was then straightforward to arrive at a complete assignment of lines which revealed that four complete spectra of different species were
MICROWAVE
SPECTRUM OF NICOTINAMIDE TABLE III
Spectroscopic ConstantsX for N-deuterated Nicotinamide?
mcotmamide-N-d]
E-n1wunam1de
E-lW3UKVlll&
wbr.ground
wbr.excited
state
Z-mcounamlde state
state
I
Id,lo,]
A iMHr
3776.41638(58)
3776.15471I31
377s 98144(87)
B/MHz
1224.3Y.51 I I)
3774.65384
1223.202 (18)
1921
1218.332127~
1~18.0271191
c/MHz
932 8719t961
934.61 I (151
937.917t241
Y38.2bS( 16)
RMS emx ikHz
24
41
rn
No. of lines
IS
I4
13
-4.839
-6.31 I
-9 815
Al&
mcobnamide-N-d,
36 I? -10.125
Ld,ll,]
A /MHz
3857 6056X591
3854.82557021
3858.lYl541811
1857.7980( I01
B/MHz
1202.9S3O(71~
1201.6228(64)
1197.651111l
1197.1981141
C/MH7
9?3.5582(58)
925.5742031
RMSelPXkH2 No. of lmes AI&
Y28.3804f901
9X929(
24
21
34
44
I4
I4
13
I3
-1916
-5.667
x597
~9.092
121
nicotmam~deN&d2
/
A/MHZ
3747.68462(48)
3745.7.5356(S?)
3750.7SO4( 1 I
BMHZ
I lYl.7387(40)
I lYO.7016~44)
I185.4680193~
I185 1?34(65)
C/&ma
9124381127)
914.0466(.30,
917.08011671
417.4668i471
RMS error kHz
22
24
53
No. of lmes
I4
I4
16
-5042
-6.456
~9.979
A /PAZ
‘Table wth all observed
uansmon
frequencies
of the N-druterated
3750.57S?5(76,)
species IS avdllable
37 I6 -10.34I
from the authors
upon request
present (frequencies of lines are listed in Table I). A small section of the spectrum, showing a transition from each of the four vibrational species of nicotinamide, is presented in Fig. 2. Spectroscopic constants (Table II), derived by least-squares fitting to a semirigid rotor Hamiltonian, were close to those predicted for the Z- and Econformers of nicotinamide. Line widths were typically 220-250 kHz for the species subsequently identified as E-nicotinamide. The strongest transitions come from the ground state of this species, those of the species we have identified (see below) as the excited E-species being 42( 4)% the intensity of those attributed to the ground state. Lines assigned to the Z-species were about 45 ( 5 ) % the intensity of the corresponding ground state E-lines and were noticeably broader. For the dominant series, the lines of the two E-species always appeared as doublets with spacings of about 5 MHz for
6
VOCELSANGER
ET AL.
TABLE IV 6-3 1 G Optimized Parameters” for Nicotinamide
148.84
148.75
122.98
122.88
135.26
135.45
98.85
98.86
99.23
99.24
123.90
117.98
120.68
121.03
118.14
117.75
123.82
123.56
117.69
117.75
164.45
21.10
163.86
21.76
172.95
-170.72
-1.96
2.10
-414.304724
-4 14.302880 4.84
0
I -3.050 0.05
0.141 ~3.098 2.957
2.999
1.823
1.833
1.702
I.418
-3.524
-3.251
0.75
0.67
I.49
5.98
1.15
0.96
2.02
XFully optimized 6-3 I G ban
molecular
geometry
set of the Gaussnn
lpyndme
88 package 191
rhe amide groups are gwen m this tile
nng kept planar).
calculated
Only the geomemcal
with the
parameters of
The full set of parameters IS avalable
from the
authors upon request ~lnlemuclear
distances are m pm,
bond angles are m degree
nicotinamide (see Fig. 2) and nicotinamide-N-d1 [ a!(I I )I, while the spacings were about 2.5 MHz for nicotinamide-N-d1 [ dclo,] and nicotinamide-N,N-&. It is not feasible to distinguish between the two conformers of nicotinamide via their respective rotational constants because they are almost identical. This is the situation for the deuterated species also (Table III 1. Quadrupole coupling constants computed via molecular-orbital calculations (Table IV) proved to be very simlar for the two conformers, indicating that it would not be possible to distinguish them from analyzes of hyperfine multiplets. We must therefore rely on measured dipole moment components because molecular-orbital calculations predict that the Z- and E-species differ substantially in their values of l*b, and hence ptotal, the Z-species being the more polar (Table IV). We also predicted significantly higher p-values for Z- as compared with E- from our simple bond-moment calculations, and similarly contrasting moments had been reported from previous semiempirical and bond-moment calculations (2) and earlier ab initio molecular-orbital computations (6).
MICROWAVE
SPECTRUM
OF
I
NICOTINAMIDE
In the frequency range covered, all of the strong transitions of nicotinamide for J > 7 exhibit both linear and quadratic Stark effects, the former dependent mainly on pa, and the latter predominantly on p*hand pLc.Values of pa were determined from measurements on individual Stark lobes, as was also the case for determinations of ph/pC and (IL: + CL;)for the E-species (see Fig. 3a). In the case of the Z-species, (CL; + ) was derived from measurements on a “bunch” of Stark lobes (Fig. 3b). Although
of
a
59460
59462
Frequency
/ 62594
62595
62596
Frequency
59466
59464
I MHz
62597
62596
I
62599
I MHz
FIG. 3. Stark effect of two different transitions of E-nicotinamide. (a) Stark effect dominated by Ii near term, lobes are fully resolved. The Stark lobes within this spectrum can be attributed to 1Ml = 1-5 of the transition 8s,0-7,,0 (shifted to lower frequencies) and 8s.,-77,, (shifted to higher frequencies). (b) The Alcomponents of the transitions 127.6-1 l& 12,,5-1 16,5bunch together, giving rise to a single unresolved Star-h lobe. The integration time was 140 min for (a) (52 scans), and 25 min for(b) ( 12 scans).
8
VOGELSANGER
ET AL.
we were unable to obtain statistically significant values for all components, moderately reliable values were nevertheless obtained for the total moment for all four molecular species, the two species identified as ground and excited E-states having p N 1.2 D, the two Z-states having p = 4.5( 4)D, these values being the basis of our identification of species. The determined values of the dipole moment components are included in Table II. If we use these two values for the dipole moments of the conformers in the formula utilized by Kuthan et al. (2) to derive the proportions of the two conformers in dioxane or benzene solutions from the value of p, 3.4 D, that they measured in these solutions, we obtain 46% E- and 54% Z-conformer, i.e., the two conformers appear to occur in very nearly equal concentrations in these solutions. This is noticeably different from the relative concentrations in the gas phase (see below), perhaps indicating that solvent stabilizing effects are significantly different for the two conformers. It is also possible that the solution dipole moment measurements could be influenced by sources of systematic error such as intermolecular association. Some additional degree of confirmation of our identification of conformers comes from the fact that the inertial defects of the four observed species, which differ by approximately a factor of two between the Z- and E-species, are close to the predicted values derived from molecular-orbital calculations (see Table II). The observed Avalues correspond to a torsional angle of 166 (2) ’ for the E-species and of 22 (2) ’ for the Z-conformer, respectively, these values being derived from the 6-3 1 G optimized structures by varying the torsional angle to achieve the observed inertial defect. These fitted torsional angles are close to those in the fully optimized structure obtained by molecular-orbital calculations (Table IV). Substitution coordinates may be derived for the amide hydrogens by using the rotational constants of the deuterated species, with either the do (shown in Table II) or the d2 species as parent species. The coordinates for E- and Z- are very similar,
TABLE V Differences
(pm) in Substitution
Coordinates for Amide Hydrogens - 1x 1(Z-nicotinamide)
1x) (E-nicotinamide)
irl =
bl rq10,t
6 -7
-3
-3
-7
-7
-4
IVJ1410,l
9
6
5
6
4
6
5
5
-14
-7
-15
-38
-4
-8
-15
-36
lol l411,l
-3
-3
-2
-1
-3
-3
-2
-I
lbl Iqll,l
109
5
4
7
6
4
-7
-16
-1
3
-7
M ldw,l
M l~llll
z hrst mlumn: Secondcolumn:
-10
-7
lr Wbxinamide) Ix1(E-nimtinamide)
- Irl (Z-nicohnamide W (Z-nicotinamide
state I) state II)
-2
3 -II
MICROWAVE
SPECTRUM
OF NICOTINAMIDE
9
because of the accidental near coincidence of the inertial axis with the amide twist axis, but differences between E- and Z-coordinates tend to support our vibrational assignment (see Table V). None of the strong transitions exhibit nuclear quadrupole splitting, splitting being predicted only for a few of the weakest lines of J > 15. We were able to resolve some hyperfine multiplets for the E-species but lines of the Z-species were mostly too broad to resolve. For the ground state of E-nicotinamide all observed multiplet patterns had spacings that deviated by less than 20% from values calculated from the coupling constants listed in Table V. The corresponding transitions of the excited state showed the same shapes as those of the ground state but the splitting appeared to be about 10% smaller. An example of such a transition of both E-species is given in Fig. 4. We were able to make approximate estimates of the relative abundances of the four species of nicotinamide from the observed relative line intensities together with the measured dipole moments. These indicated t?at the excited state of E-nicotinamide had a beam concentration of about 50% of that of the ground state which, depending on the degree of vibrational cooling in the beam, corresponds to an excitation energy of 120-220 cm-’ (depending on whether we assume a vibrational temperature of 250 or 450 K, the latter corresponding to no vibrational cooling). The two other species, i.e., the two states of Z-nicotinamide, were similarly estimated to be of equal relative concentration in the beam, about 4% of the ground-state E-species, corresponding to an excitation energy of 550- 1050 cm-‘. The same ordering of energies of states, i.e., a significant energy difference between ground and excited E-states, the two states of the Z-conformer being higher again, was found when some transitions were reobserved with the nozzle heated to 3 15”C, intensities being carefully compared with those observed at 180°C. The Z- : E-energy separation seems to be higher than predicted by ab initio molecular-orbital calculations based on 3-2 1, 4-3 1, or 6-3 1 G functions. All three basis sets calculate the separation to be about 400 cm-‘. Qualitatively, however, the observed relative energies agree with molecular-orbital predictions. We suggest that the two states of the Z-nicotinamide arise from the splitting of the ground state from tunneling through the barrier of the planar conformation of this conformer. Presumably the barriers to rotation to the E-conformation and the planar E-barrier are too high for us to observe other splittings. Although all lines studied for most of the isotopomers of nicotinamide were assigned to the four states of the two conformers as just discussed, in the case of the dideuterocompound several additional relatively weak lines were observed in the immediate vicinity of the strongest transitions. These presumably are to be attributed to one or more additional low-lying vibrational states for this isotopomer. In summary, we conclude that we have observed four species of nicotinamide in the vapor phase, a ground and excited state of the E-conformer and a split ground state of the Z-conformer. The vibrational/conformational assignment is based on observed dipole moments and relative line intensities, conformational assignment being assisted by observed linewidths and some hyperfine splitting patterns. The good agreement of observed substitution coordinates, inertial defects, and relative energies with those derived from ab initio molecular orbital calculations further supports the assignments. All features present in the observed spectrum are satisfactorily accounted for in terms of our assignments.
10
VOGELSANGER
58963
ET AL.
58964
Frequency
58965 I
58966
MHz
b
59056
Frequency
59057
59058
f MHz
FE. 4. Observed and calculated nuclear hyperfine pattern ofthe I75,~3I 64.12transition of both vibrational ground (a) and excited (b) states of E-nicotinamide. The calculated patterns are based on the ab initio values of the coupling constant given in Table IV. The integration time was 140 min ( 100 scans) for (a) and 280 min (200 scans) for (b).
ACKNOWLEDGMENTS B.V. acknowledges the grant of a Nachwuchsstipendium ofthe Schweizerischer Nationalfonds zur FCirderung der wissenschaftlichen Forschung. The research was supported by a grant from the Australian Research Council. RECEIVED:
July 2, 1990
MICROWAVE
SPECTRUM
OF NICOTINAMIDE
11
REFERENCES W. B. WRIGHT AND G. S. D. KING. .&~a Crystallogr. 7, 283-288 ( 1954). J. KUTHAN. L. MUSIL, AND V. JEHLICKA,Collect. Czech. Chem. Commwi. 42, 283-291 ( 1977). W. P. PURCELL,J. Phys. Chem. 68, 2666-2670 ( 1964). W. P. P~JRCELL AND J. A. SINGER,J. Phq’s. Chem. 69,4097-4101 (1965). S. BOHM AND J. KUTHAN. C’ollecr. C‘-_eChC’henl. Common. 48, 1842-1853 ( 1983). E. IACONIS. N. RCJSSO.AND M. TOS~ANO, J. Chem Res. 1984, 416-417 ( 1984). A, S. EL-SHAHAWY,M. M. GIRGIS,AND M. T. ISMAIL,Spectrochim. .&~a Purl A43, 1371-1375 ( 1987). R. D. BROWN, J. G. CROFTS, P. D. GODFREY, D. MCNAUGHTON. AND A. P. PIERLOT,J. Mol. Slrucr. 190, 185-193 (1988). Y. GAUSSIAN88. M. J. FRISCH,M. HEAD-GORDON.H. B. SCHLEGEL,K. RAGHAVACHARI,J. S. BINKL~Y. C. GONZALEZ, D. J. DEFREES.D. J. Fox, R. A. WHITESIDE,R. SEEGER,C. F. MELIUS.J. BAKER, R. L. MARTIN, L. R. KAHN. J. J. P. STEWART.E. M. FLUDER.S. TOPIOL,ANDJ. A. POPLE.Gaussian. Inc.. Pittsburgh PA. 10 F. J. LOVAS. J, Phys. Chem. Re/. Datu 14, 395-488 ( 1985). I1 R. L. COOK. R. T. WALDEN, AND G. E. JONES.J. zMol. Speclrosc. 53, 370-378 ( 1974 ). I. .?. 3. 4. 5. 6. 7. X.
OF MOLECULAR
SPECTROSCOPY
145, 1-t 1 ( 199 1)
The Microwave Spectrum of a Vitamin: Nicotinamide BEAT VOGELSANGER,RONALD D. BROWN, PETER D. GODFREY, AND ANTHONY P. PIERLOT Centre,for High-Resolution Spectroscopy and Opto-Electronic Technology, Monash Universit?: Clayton, Victoria 3168, .4ustralia We have observed the microwave spectrum of nicotinamide and three deuterated isotopomers in the range 57-78 GHz, identifying four species that we conclude are the ground and vibrationally excited E-conformer and a split ground state of the Z-conformer. Some hyperfme splitting patterns were observed and dipole moments of the two conformers were determined. A detailed ab initio molecular-orbital investigation was also undertaken, involving complete geometry optimization at the 6-3 1 G level of basis functions. The extent of torsional twist of the amide groupin the two conformers was in good agreement with the theoretical calculations and similar satisfactory agreement was found between observed quadrupole hyperfme patterns and patterns computed from theoretically derived coupling constants. The observed relative energies of E- and Z -nicotinamide are qualitatively in agreement with predictions of the molecular-orbital calculations but the separation is numerically greater. 0 1991 Academic Press. Inc
INTRODUCTION
The shape of the nicotinamide molecule (Fig. 1) has been the subject of much interest, particularly with respect to the conformation of the amide group relative to the pyridine ring. The X-ray structure of the crystal was reported over 30 years ago ( I), the conclusion being that the oxygen of the amide was approximately entgegen to the pyridine nitrogen but twisted out of the plane of the pyridine ring by rotation around the C-C bond by about 156” from the zusammen position. The conformation of nicotinamide was further studied via dipole moment measurements in benzene solution by Kuthan et al. (2), following earlier measurements of Purcell and coworker ( 3, 4). They obtained a moment intermediate between values estimated, by several lines of calculation, for the Z- and E-conformations, concluding that two different rotamers were present but finding a puzzling temperature dependence of the dipole moment. Theoretical studies by Bijhm and Kuthan (5)) in which the torsional angle around the C-C bond was optimized using an ab initio STO-3 G basis molecular orbital calculation, the other geometrical parameters being constrained to CNDO/2 values, indicated that both Z- and E-conformers are nonplanar. A more complete treatment by Iaconis et al. (6) in which all parameters apart from the pyridine ring were optimized, the pyridine ring parameters being held at the values derived from a separate geometry optimization for pyridine itself, all done using the STO-3 G basis, predicted twist angles of 24 and 158”, respectively, around the C-C bond, the Econformer being more stable by about 1 kJ mole-’ . Other SCF-CI calculations by El1
0022-2852/91 $3.00 Copyright 0 All n&s
1991 by Academic Press, Inc.
of reproduction m any form resewed.
VOGELSANGER
2
ET AL.
Z-nicotinamide
E-nicotinamide FIG. 1. Structure
of the two conformers
of nicotinamide
with labeling of the nuclei.
Shahawy et a/. (7) predicted the Z-conformer to be the more stable. Thus there is some disagreement between different theoretical approximations, while experimental evidence is complicated by the unknown effects of solvent or the crystal lattice. TABLE I Observed
Transitions
(MHz)
E-nlcotinamik vibr. ground state TRANSlTlON 880-771 881.770 981-872 982-871 990.88, 991.880 IO 7 3. IO 7 4. IO 9 I IO 9 2. IO IO 0 IO 10 I II 74-10 II 7 S-10 II 8 3-10 I1 84-1073 II 9 2-10 II 9 3-10 I1 IO 1 II 10 2 13 6 7 13 6 8 14 6 8 14 6 9 IS 5 IO 16 4 I2 16 5 II 16 7 9 I6 7 10 II 5 13 17 7 IO 17 7 II 18 5 14 20 5 16 27 3 24 28 3 2S 28 4 25 29 2 27 29 3 26 31 031 13 II 2 14 II 3 IS II 4 16 II 5 17 II 6 23 II 12 -
9 9 9 9 9 9
10 10 I2 I2 13 13 14 IS 15 I5 IS 16 16 16 17 19 26 27 27 28 28 30 13 14 15 16 17 23
OBS. FREQ.
6 6 8 8 9 9 6 6 7
4 3 2 I L 0 5 4 4
8 8 9 9 5 5 5 5 4 3 4 6 6 4 6 6 4 4 4 4 3 3 4 i 10 IO IO 10 IO IO
3 2 2 I 8 7 9 8 II I3 12 IO 9 I2 I, IO 13 I5 23 24 24 26 25 30 3 4 5 6 7 I3
DC
of the Four Different E-nrotinamide wbr. excited safe OBS. FREQ.
W
Species of Nicotinamide 2.n,mtinamlde state I
Z-N~tiMllllde state I,
OBS. FREQ.
W
0.004 0.004 0.004 -0.004 -0.012 -0012 -0.060 a038 0.03 I 0.03 I 0.063 0.063 -0.05 I 0.03s 0.002 0.003 -0.033 -0.033 -0.070 -0.070 0.068 -0.01 I 0.045 0.061 0.077
-0.002 0.070 -0.101
60663.387 71195.079 71181.289
a.c40 0041 -0.1 is
0.074 -0.038
73332.154 73302.354
0.055 -0.041
59392.030
-0.003
58613.758 58592.622 58567.036
0.018 -0.0 I 3 0 035
59424.675 59424.675 61613.359 61613.359 67201.796 67201.796 58208.690 58208.690 69390.569 69390X9 76978 R42 7*971 Ro? 6039md3 60390.843 65987.61 I 65987.61 I 71578.696 71578.696 77167.626 71167.626 59104.767 59073.329 61256.483 61186.576 58245.412
o.w3 0.003 o.cm7 0.007 -0.w3 -0.003 -0.044 -0.019 0.016 0.016 0.011 0.01 I -0.047 0.053 0.019 0.020 OSKW 0.009 -0.533 -0.003 0.059 -0.041 0.007 -0.023 0.W
59415.983 59415.983 61603.990 61603.990 67191.977 6,191 477 S8199.247 58199.247 69380.081 69380.081 74967 950 74967.950 60380.948 60380.948 63976.961 65976.961 71567.479 71567.479 77155.921 77155.921 59096.308 59068.400 61247.954 61185.815
-0.023 -0.023 0.018 0.018
60766S95 71207.029 71191.001 59056.320 73342.695 73307.957
-0.070 0.063 -0.049 -0.029 0.060 -0.031
60672.238 71198.960 71184.981 73335.797 73305.648
57254.013 59265.631 59783.689 59453.105 61232.913 59500.625
57299.218 59322.836
-0.035 0.037
58679.374
58623.184 58600.988
-0.037 -0.019
58638.674 58621.476 58W.234
-0.028 0.028 -0.004
58541.617
0.008
58543.616
-0.004
58595.827 58235.692
-0.045 0.029
W.-
594 10.468 594 10.468 61598.452 61598.452 67185.71 I 67185.71, 58194SlS 58194.515 69373.815 69373.815 749643.945 7496Q.945 60376.326 60376.326 65971.483 6597 I.483 71561.223 71561.223 77148.911 77148.911 59092.970 5906S.351 61244.781 61183.357 58181 288
59461.932 59461.932 61650.160 61650.160 67244.076 67244.076 58239.458 58239.458 69432.376 69432.376 75026.129 75026.129 60420.95 1 60420.95 1 66023.375 66023.375 71620.017 71620.017 77214.445 77214.445 59127.040 59093.794 61277.978 61203.975 58286.403 60855.183 60325.551 71231.893 71214.747 58964.166 73366.106 13328.944 mw82:;;;
” , ,
OBS. FREQ.
:g;: -0.018 0.004 0.04 I 0.041
__.._._
t:::: -0.053 0.034 “-012
MICROWAVE
SPECTRUM
OF NICOTINAMIDE
3
In view of the biological significance of nicotinamide it was felt desirable to attempt to establish unambiguously the conformers of this molecule when not affected by environmental influences. Accordingly, we have investigated its microwave spectrum in the gas phase and, because the theoretical studies quoted above do not seem satisfactory by contemporary standards, we have made further ab initio molecular-orbital calculations that have included a more thoroughgoing geometrical optimization and have employed a larger basis set. EXPERIMENTAL
DETAILS
A millimeter-wave spectrometer, based on observed absorption in a supersonic expanding plume of nicotinamide vapor entrained in argon and described previously (8 ), was used for the measurements. The source was a phase-locked millimeter-wave klystron or a K-band klystron with tripler. Stark modulation at voltages up to 6000 V were employed at a plate separation of 3.5 cm. Accurate line frequencies were obtained by repetitive scans using a data-collection system based on a VAX 1 1 / 780 computer. Measurements were made in the frequency range 57-78 GHz. Commercially available nicotinamide was used in a volatilization tube, heated to 160- 17O”C, and a stream of argon at pressures of about 300 hPa was bubbled through the melted sample before being expanded through a nozzle 500 pm in diameter. The liquid evaporated at rates typically of about 0.3 g hr-’ . We were unable to detect any decomposition of the nicotinamide under these conditions. Nicotinamide-N-d1 and nicotinamide- N, N-d2 were generated by dissolving nicotinamide in D20/H20 1: 1. After 24 hr at 40°C. the water was removed in a “Ro-
60360
60390
60410
60400
Frequency
60420
/MHz
FIG. 2. Typical strong rotational transitions [ 11 7,4- 106,5/ 117,5- 106,4] for the four different species of nicotinamide. The transition lines correspond to (from left to right) state II of the Z- , state I of the Z- , vibr. excited state of the E- , and vibr. ground state of the E-conformer. The integration time to record this spectrum was 100 min. (6 scans averaged).
VOGELSANGER
ET AL.
TABLE II
Calculated and Observed Spectroscopic Parameters. Geometric Parameters. and Dipole Moment Components” of Nicotinamide E-nicotinamide vibr. ground state
E-niwtinamide vibr. excaed state
3954.5
3891.2262(211
3888.7292(29
3953.0
3888.l8OOf39)
3887.8089(44)
1242.6
1236.2442(16)
1234.7691(18
1238.0
1231.413(25)
1230.893(27)
953.5
944.7920(141
946.9075(2
957.3
949.855(29)
950.462(42)
0.0361(12)
0.0262(
0.0458~31)
0.0822(66:
0.219~22)
0.334(14)
0.329(20)
0.320(29)
E-lWXiMlll1& talc. 6.3lG
-0.00.547(73) -0.00375(30)
-4.497
(reference ‘Statntically
I
Z-nlcomwnide stale
13:
-0.065~131
0”
-0.S I6(5 1,
0”
24
32
33
26
-3.768
-5.535
1
-8.174
Z-mcotmarmde
state II
0.0224(221 0.111(10) 0.336(30) 0”
0’
0’
39
46
23
24
-8.325
-8.851
194
I84
I85
197
190
191
194
I86
I89
I89
177
180
46
74
63
61
88
81
336
335
334
338
338
338
I05
105
106
100
95
96
I8
28
26
25
38
35
0.75
0.315(2V
I.49
l.l5(2)Y
I.15
WFrom Stark-effect
Z-oicotinamide talc. 6.3lG
0.316(2P
0.67
0.209(2)Y
0.200(4)Y
5.98
CO.2
0.96
I .88
l.l58(7lY
l.O5(4)Y
6.06
4.5(4)2
4.5(4)7
2.02
1.20(l)
1.09(5)
6.09
4.5(4)
4.5(4)
measurements.
Tlx field was cahbrated wth the 413-404 tnnsltion
of S@ usmg the dipole moment oh = 1.63305(4)D
IO). not signliicant pammeler
YFrom rnea~uremenf~ of Individual ‘From measuremen&
(F-test.
99%
Stark lobes.
confidence mterval: see reference I I ). Numbers m parenlhesls represent the standard daubon
of unresolved clumps of Stark lobes for two different transmons.
of the fit only.
Numbers m parsthea
are an esbmate of the error.
tavap,” providing a mixture of nicotinamidedo/d1 /d:. Both possible monosubstitutions on the amide nitrogen were obtained in sufficient quantities for the spectra to be observed and analyzed. A sample containing more than 90% nicotinamideN, Nd2 was obtained by repeating the above procedure twice with DzO instead of DzO/ H20. RESULTS
The observed spectrum in the vicinity of 60 GHz is ph-type, dominated by R-branch series: .!7,(5-6) (J 1 )6.~J-7~/J7,~J-7~- (J - 1 )6,c.-6) with J = 10, 11. 12, - * - and &(5_7) (J 1)7.~J-~~/J8,~J-8~ - (.I - 1 )7.(~-7j with J = 8, 9, - - - . This pattern of lines, the spacings between two transitions in each of these series being about 2.2 GHz, was recognizably similar to a theoretical spectrum generated by using geometries of nicotinamide that were predicted by fully optimized ab initio molecular-orbital calculations using a 6-3 1 G basis and the Gaussian 88 code ( 9). All other transitions in this range are much weaker. It was then straightforward to arrive at a complete assignment of lines which revealed that four complete spectra of different species were
MICROWAVE
SPECTRUM OF NICOTINAMIDE TABLE III
Spectroscopic ConstantsX for N-deuterated Nicotinamide?
mcotmamide-N-d]
E-n1wunam1de
E-lW3UKVlll&
wbr.ground
wbr.excited
state
Z-mcounamlde state
state
I
Id,lo,]
A iMHr
3776.41638(58)
3776.15471I31
377s 98144(87)
B/MHz
1224.3Y.51 I I)
3774.65384
1223.202 (18)
1921
1218.332127~
1~18.0271191
c/MHz
932 8719t961
934.61 I (151
937.917t241
Y38.2bS( 16)
RMS emx ikHz
24
41
rn
No. of lines
IS
I4
13
-4.839
-6.31 I
-9 815
Al&
mcobnamide-N-d,
36 I? -10.125
Ld,ll,]
A /MHz
3857 6056X591
3854.82557021
3858.lYl541811
1857.7980( I01
B/MHz
1202.9S3O(71~
1201.6228(64)
1197.651111l
1197.1981141
C/MH7
9?3.5582(58)
925.5742031
RMSelPXkH2 No. of lmes AI&
Y28.3804f901
9X929(
24
21
34
44
I4
I4
13
I3
-1916
-5.667
x597
~9.092
121
nicotmam~deN&d2
/
A/MHZ
3747.68462(48)
3745.7.5356(S?)
3750.7SO4( 1 I
BMHZ
I lYl.7387(40)
I lYO.7016~44)
I185.4680193~
I185 1?34(65)
C/&ma
9124381127)
914.0466(.30,
917.08011671
417.4668i471
RMS error kHz
22
24
53
No. of lmes
I4
I4
16
-5042
-6.456
~9.979
A /PAZ
‘Table wth all observed
uansmon
frequencies
of the N-druterated
3750.57S?5(76,)
species IS avdllable
37 I6 -10.34I
from the authors
upon request
present (frequencies of lines are listed in Table I). A small section of the spectrum, showing a transition from each of the four vibrational species of nicotinamide, is presented in Fig. 2. Spectroscopic constants (Table II), derived by least-squares fitting to a semirigid rotor Hamiltonian, were close to those predicted for the Z- and Econformers of nicotinamide. Line widths were typically 220-250 kHz for the species subsequently identified as E-nicotinamide. The strongest transitions come from the ground state of this species, those of the species we have identified (see below) as the excited E-species being 42( 4)% the intensity of those attributed to the ground state. Lines assigned to the Z-species were about 45 ( 5 ) % the intensity of the corresponding ground state E-lines and were noticeably broader. For the dominant series, the lines of the two E-species always appeared as doublets with spacings of about 5 MHz for
6
VOCELSANGER
ET AL.
TABLE IV 6-3 1 G Optimized Parameters” for Nicotinamide
148.84
148.75
122.98
122.88
135.26
135.45
98.85
98.86
99.23
99.24
123.90
117.98
120.68
121.03
118.14
117.75
123.82
123.56
117.69
117.75
164.45
21.10
163.86
21.76
172.95
-170.72
-1.96
2.10
-414.304724
-4 14.302880 4.84
0
I -3.050 0.05
0.141 ~3.098 2.957
2.999
1.823
1.833
1.702
I.418
-3.524
-3.251
0.75
0.67
I.49
5.98
1.15
0.96
2.02
XFully optimized 6-3 I G ban
molecular
geometry
set of the Gaussnn
lpyndme
88 package 191
rhe amide groups are gwen m this tile
nng kept planar).
calculated
Only the geomemcal
with the
parameters of
The full set of parameters IS avalable
from the
authors upon request ~lnlemuclear
distances are m pm,
bond angles are m degree
nicotinamide (see Fig. 2) and nicotinamide-N-d1 [ a!(I I )I, while the spacings were about 2.5 MHz for nicotinamide-N-d1 [ dclo,] and nicotinamide-N,N-&. It is not feasible to distinguish between the two conformers of nicotinamide via their respective rotational constants because they are almost identical. This is the situation for the deuterated species also (Table III 1. Quadrupole coupling constants computed via molecular-orbital calculations (Table IV) proved to be very simlar for the two conformers, indicating that it would not be possible to distinguish them from analyzes of hyperfine multiplets. We must therefore rely on measured dipole moment components because molecular-orbital calculations predict that the Z- and E-species differ substantially in their values of l*b, and hence ptotal, the Z-species being the more polar (Table IV). We also predicted significantly higher p-values for Z- as compared with E- from our simple bond-moment calculations, and similarly contrasting moments had been reported from previous semiempirical and bond-moment calculations (2) and earlier ab initio molecular-orbital computations (6).
MICROWAVE
SPECTRUM
OF
I
NICOTINAMIDE
In the frequency range covered, all of the strong transitions of nicotinamide for J > 7 exhibit both linear and quadratic Stark effects, the former dependent mainly on pa, and the latter predominantly on p*hand pLc.Values of pa were determined from measurements on individual Stark lobes, as was also the case for determinations of ph/pC and (IL: + CL;)for the E-species (see Fig. 3a). In the case of the Z-species, (CL; + ) was derived from measurements on a “bunch” of Stark lobes (Fig. 3b). Although
of
a
59460
59462
Frequency
/ 62594
62595
62596
Frequency
59466
59464
I MHz
62597
62596
I
62599
I MHz
FIG. 3. Stark effect of two different transitions of E-nicotinamide. (a) Stark effect dominated by Ii near term, lobes are fully resolved. The Stark lobes within this spectrum can be attributed to 1Ml = 1-5 of the transition 8s,0-7,,0 (shifted to lower frequencies) and 8s.,-77,, (shifted to higher frequencies). (b) The Alcomponents of the transitions 127.6-1 l& 12,,5-1 16,5bunch together, giving rise to a single unresolved Star-h lobe. The integration time was 140 min for (a) (52 scans), and 25 min for(b) ( 12 scans).
8
VOGELSANGER
ET AL.
we were unable to obtain statistically significant values for all components, moderately reliable values were nevertheless obtained for the total moment for all four molecular species, the two species identified as ground and excited E-states having p N 1.2 D, the two Z-states having p = 4.5( 4)D, these values being the basis of our identification of species. The determined values of the dipole moment components are included in Table II. If we use these two values for the dipole moments of the conformers in the formula utilized by Kuthan et al. (2) to derive the proportions of the two conformers in dioxane or benzene solutions from the value of p, 3.4 D, that they measured in these solutions, we obtain 46% E- and 54% Z-conformer, i.e., the two conformers appear to occur in very nearly equal concentrations in these solutions. This is noticeably different from the relative concentrations in the gas phase (see below), perhaps indicating that solvent stabilizing effects are significantly different for the two conformers. It is also possible that the solution dipole moment measurements could be influenced by sources of systematic error such as intermolecular association. Some additional degree of confirmation of our identification of conformers comes from the fact that the inertial defects of the four observed species, which differ by approximately a factor of two between the Z- and E-species, are close to the predicted values derived from molecular-orbital calculations (see Table II). The observed Avalues correspond to a torsional angle of 166 (2) ’ for the E-species and of 22 (2) ’ for the Z-conformer, respectively, these values being derived from the 6-3 1 G optimized structures by varying the torsional angle to achieve the observed inertial defect. These fitted torsional angles are close to those in the fully optimized structure obtained by molecular-orbital calculations (Table IV). Substitution coordinates may be derived for the amide hydrogens by using the rotational constants of the deuterated species, with either the do (shown in Table II) or the d2 species as parent species. The coordinates for E- and Z- are very similar,
TABLE V Differences
(pm) in Substitution
Coordinates for Amide Hydrogens - 1x 1(Z-nicotinamide)
1x) (E-nicotinamide)
irl =
bl rq10,t
6 -7
-3
-3
-7
-7
-4
IVJ1410,l
9
6
5
6
4
6
5
5
-14
-7
-15
-38
-4
-8
-15
-36
lol l411,l
-3
-3
-2
-1
-3
-3
-2
-I
lbl Iqll,l
109
5
4
7
6
4
-7
-16
-1
3
-7
M ldw,l
M l~llll
z hrst mlumn: Secondcolumn:
-10
-7
lr Wbxinamide) Ix1(E-nimtinamide)
- Irl (Z-nicohnamide W (Z-nicotinamide
state I) state II)
-2
3 -II
MICROWAVE
SPECTRUM
OF NICOTINAMIDE
9
because of the accidental near coincidence of the inertial axis with the amide twist axis, but differences between E- and Z-coordinates tend to support our vibrational assignment (see Table V). None of the strong transitions exhibit nuclear quadrupole splitting, splitting being predicted only for a few of the weakest lines of J > 15. We were able to resolve some hyperfine multiplets for the E-species but lines of the Z-species were mostly too broad to resolve. For the ground state of E-nicotinamide all observed multiplet patterns had spacings that deviated by less than 20% from values calculated from the coupling constants listed in Table V. The corresponding transitions of the excited state showed the same shapes as those of the ground state but the splitting appeared to be about 10% smaller. An example of such a transition of both E-species is given in Fig. 4. We were able to make approximate estimates of the relative abundances of the four species of nicotinamide from the observed relative line intensities together with the measured dipole moments. These indicated t?at the excited state of E-nicotinamide had a beam concentration of about 50% of that of the ground state which, depending on the degree of vibrational cooling in the beam, corresponds to an excitation energy of 120-220 cm-’ (depending on whether we assume a vibrational temperature of 250 or 450 K, the latter corresponding to no vibrational cooling). The two other species, i.e., the two states of Z-nicotinamide, were similarly estimated to be of equal relative concentration in the beam, about 4% of the ground-state E-species, corresponding to an excitation energy of 550- 1050 cm-‘. The same ordering of energies of states, i.e., a significant energy difference between ground and excited E-states, the two states of the Z-conformer being higher again, was found when some transitions were reobserved with the nozzle heated to 3 15”C, intensities being carefully compared with those observed at 180°C. The Z- : E-energy separation seems to be higher than predicted by ab initio molecular-orbital calculations based on 3-2 1, 4-3 1, or 6-3 1 G functions. All three basis sets calculate the separation to be about 400 cm-‘. Qualitatively, however, the observed relative energies agree with molecular-orbital predictions. We suggest that the two states of the Z-nicotinamide arise from the splitting of the ground state from tunneling through the barrier of the planar conformation of this conformer. Presumably the barriers to rotation to the E-conformation and the planar E-barrier are too high for us to observe other splittings. Although all lines studied for most of the isotopomers of nicotinamide were assigned to the four states of the two conformers as just discussed, in the case of the dideuterocompound several additional relatively weak lines were observed in the immediate vicinity of the strongest transitions. These presumably are to be attributed to one or more additional low-lying vibrational states for this isotopomer. In summary, we conclude that we have observed four species of nicotinamide in the vapor phase, a ground and excited state of the E-conformer and a split ground state of the Z-conformer. The vibrational/conformational assignment is based on observed dipole moments and relative line intensities, conformational assignment being assisted by observed linewidths and some hyperfine splitting patterns. The good agreement of observed substitution coordinates, inertial defects, and relative energies with those derived from ab initio molecular orbital calculations further supports the assignments. All features present in the observed spectrum are satisfactorily accounted for in terms of our assignments.
10
VOGELSANGER
58963
ET AL.
58964
Frequency
58965 I
58966
MHz
b
59056
Frequency
59057
59058
f MHz
FE. 4. Observed and calculated nuclear hyperfine pattern ofthe I75,~3I 64.12transition of both vibrational ground (a) and excited (b) states of E-nicotinamide. The calculated patterns are based on the ab initio values of the coupling constant given in Table IV. The integration time was 140 min ( 100 scans) for (a) and 280 min (200 scans) for (b).
ACKNOWLEDGMENTS B.V. acknowledges the grant of a Nachwuchsstipendium ofthe Schweizerischer Nationalfonds zur FCirderung der wissenschaftlichen Forschung. The research was supported by a grant from the Australian Research Council. RECEIVED:
July 2, 1990
MICROWAVE
SPECTRUM
OF NICOTINAMIDE
11
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