Analytical spectroscopy and structure of biomolecules using an ab

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Anal. Chem. 1902, 64, 2726-2734

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Analytical Spectroscopy and Structure of Biomolecules Using an ab Initio Computational Method Yeong Choi and David M. Lubman' Department of Chemistry, The University of Michigan, Ann Arbor, Michigan 48109

An ab lnltlo computational method has been applied to the study of blologlcally and pharmaceutlcally Important phenethylamlne and Its derlvatlves (tyramlne, dopamlne, and tyroslne) In order to predlct the structure of m e of thelr stable conformers and consequently thelr spectrorcoplc propertles. The structures were optlmlzed at the HFi3-21G level and thelr stabllltks were determlned by performlng vlbratlonal frequency calculationson the optlmkedstructures. I n the case of phenethylamlne and tyramlne, three stable conformers were found, two of which have an ethylamlnefolded structure and the other has an ethylamlne-extended structure wlth the former two lylng about 0.01 eV hlgher In energy than the latter. The stable conformers of phenethylamlne found In the present study are compared wHh the resultsobtained by experhnentsand other theoretkal methods From the resultlng structural calculatlons, I R and Raman vlbratlonal frequencles of phenethylamlneand tyramlne were predlcted whlch are partlcularly useful for the analyses of the experlmental spectra when avallable. I n partlcular, the capablllty to vlwallze the vlbratlonal motlon responslbk for a partlcular peak enhances lts usefulness In the assignment of the features In the IR spectra. Ioniratlon potentlak,electron affInltles, and the charge dendty dlstrlbutlon of phenethylamlne and Its derlvatlves were predlcted, and thelr utlllty In the Interpretatlon of the experimental spectroscoplc data Is demonstrated.

INTRODUCTION There has recently been a great deal of interest in the study of biological molecules by a variety of spectroscopic techniques. These have ranged from studies of the trace detection of biological species1+ to studies of their structure, function, and reactivity.'-12 The former are important for analytical (1)Lubman, D. M.; Li, L. In Lasers and Mass Spectrometry; Lubman D. M., Ed.; Oxford University Press: Oxford, U.K., 1990; pp 353-381. (2)Li, L.; Lubman, D. M. Anal. Chem. 1988,60,2591-2598. (3)Wehry, E. L.; Gore, R. R.; Dickinson, R. B. In Lasers in Chemical Analysis; Hieftje, G. M., et al., Eds.; The Humana Press: Clifton, NJ, 1981; Chapter io. (4)Strojny, N.; desilva, J. A. In Lasers in Chemical Analysis; Hieftje, G. M.. et al.. Eds.: The Humana Press: Clifton. NJ. 1981.Chauter 11. ( 5 )'See: Analytical Laser Spectroscopy;Omenettb, N. Ed.; Chemical Analysis Series, Vol. 50; John Wiley & Sons, Inc.: New York, 1979. (6)See: Detectors for Liquid Chromatography; Yeung, E. S., Ed.; John Wiley & Sons, Inc.: New York, 1986. (7)Pullman, B.; Coubelis, J.-L.; Courriere, Ph.; Gervois, J.-P. J. Med. Chem. 1972,15,17-23. (8)Pauling, P. In ConformationofBiological molecules and polymers; Berg", E. D., et al., Eds.; The Jerusalem symposia on Quantum Chemistry and Biochemistry V; Academic Press Inc.: New York, 1973; pp 505-516. (9)Pullman, B.; Courrier, P. In Conformation of Biological molecules and polymers; Bergman, E. D., et al., Eds.; The Jerusalem symposia on Quantum Chemistry and Biochemistry V; Academic Press Inc.: New York, 1973;pp 547-570. (10)Pullman, B.; Berthod, H.; Courriere, Ph. Int. J. Quantum Chem., Quantum Biol. Symp. 1974,1,93-108. (11)Martin, M.; Carbo, R.; Petrongolo, C.; Tomasi, J. J. Am. Chem. SOC.1975,97,1338-1347.

purposes while the latter have been particularly important in understanding the properties of molecules for drug design and pharmacological activity. The various studies used to probe these molecules have involved different spectroscopic techniques in many different environments ranging from gasphase jet studies12-18 to in vitro studies in the liquid phase a t physiological pH.lS2l However, any particular spectroscopic method can only provide a limited amount of information on any system and the experimental spectra of these molecules may not be easily interpretable by themselves. In particular, many of these biomolecules are thermally labile and spectroscopic measurements of these molecules are often plagued by artifacts due to thermal decomposition or other background impurities that may not be easily distinguishable in these experiments. Thus, there has been a motivation toward developing computational methods that can be used to predict a priori a broad range of spectroscopic prope r t i e ~ . ~Such ~ - ~ calculations ~ can be used to study the properties of various biological substances which are difficult to measure accurately by present experimental methods. Such computational methods can be extremely valuable for developing an accurate analytical data base for spectral properties of molecules in combination with experimental methods. In addition, computational methods have found an incremingly important role in analyticalchemistry more generally.22-n There are several classesof molecules that are being studied rather extensively because of their potent biological activity and pharmaceutical importance. These include various derivatives of catechol, indole and phenyl compounds. In particular, phenethylamine is a precursor for molecules such as tyramine (adrenergic),tyrosine (amino acid), and dopamine (neurotransmitter) and is structurally related to amphetamine (stimulant) and various other important hallucinogens. The indole group of compounds includes derivativesof tryptamine, such as serotonin (neurotransmitter) and NJV-dimethyl(12)Mazurek, A. P.; Weinstein, H.; Osman, R.; Topiol, S.; Ebersole, B. J. Int. J. Quantum Chem., Quantum Biol. Symp. 1984,11,183-194. 1988, (13)Cable, J. R.; Tubergen, M. J.;Levy, D. H. J. Am. Chem. SOC.

110,7349-7355. (14)Cable, J. R.; Tubergen, M. J.; Levy, D. H. J.Am. Chem. SOC.1989, 11 1, 9032-9039. (15)Breen, P. J.; Warren, J. A.; Bernstein, E. R.; Seeman, J. I. J. Am. Chem. SOC.1987,109, 3453-3455. (16)Seeman, J. I.; Secor,H. V.;Breen, P. J.;Grassian,V. H.;Bernstein, E.R. J. Am. Chem. SOC.1989,111, 3140-3150. (17)Hager, J.; Ivanco, M.; Smith, M. A.; Wallace, S. C. Chem. Phys. 1986,105,397-416. (18)Bickle, G. A.; Leach, G. W.; Demmer, D. R.; Hager, J. W.; Wallace, S. C. J. Chem. Phys. 1988,88,1-8. (19)Spiro, T.G.In Chemical andBiochemical Applications of Lasers; Moore, C. B., Ed.; Academic Press: New York, 1974;Chapter 2. (20)See: Spectroscopy in Biochemistry; Bell, S. E., Ed.; CRC Press, Inc.: Boca Raton, FL, 1981;Vol. 1. (21)See: Carey, P. R. Biochemical Applications of Raman and Resonance Raman Spectroscopies; Academic Press: New York, 1982. (22)Stanton, D. T.; Jura, P. C. Anal. Chem. 1990,62,2323-2329. (23)Stuper,A.J.;BrOgger,W.E., Jurs,P.C. ComputerAssistedStudies of Chemical Structure and BiologicalFunction;Wiley-Interscience: New York, 1979. (24)Barber, A. S.;Small, G. W. Anal. Chem. 1989,61,2658-2664. (25)Rogers, L. B. Anal. Chem. 1990,62,703A-711A. (26)Small, G.W.; Jura, P. C. Anal. Chem. 1984,56,1214-1323. (27)Morrow, J. C.; Baer, T. J. Phys. Chem. 1988,92,6567-6571.

0003-2700/92/0364-2726$03.00/0 0 1992 Amerlcan Chemical Soclety

ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1992

tryptamines (hallucinogens). Thus, it is important to develop an analytical data base for spectroscopicdetection and study of the basic structure of these moleculea. Some of these species have been studied in gas-phase ultraviolet absorption spectroscopy2.B and in the condensed phase in the ultraviolet, visible, and infrared regions of the spectrum.24-32 However, the information availableon many of these important species is clearly rather limited or nonexistent. Here, computational methods can, in principle, be used to predict many of the properties and spectra of these molecules. Traditionally, theoretical predictions on the structure and electronicproperties of these classes of compounds have been performed by using semiempirical methods such as CNDO (completeneglect of differentialoverlap),INDO (intermediate neglect of differential overlap), PCILO (perturbative configuration interaction using localized orbitals), and EHT (extended Htickel theory).7-12 However, the accuracy of the predictions by these semiempiricalmethods is often limited, in part, due to crude approximations inherent in these methods. Ab initio computational methods can be expected to yield much more accurate informationthanthe semiempirical methods. However, general application of ab initio methods has been limited to considerably smaller molecules and biomolecules have been considered as too large to be studied by these methods. For molecules of this size, most of which are of low symmetry, such computations have been prohibitively expensive and time-consuming,and it is only recently that the application of ab initio calculations to these biomolecules has become reasonably feasible. This is due to (a) improved algorithms in computational methodology and (b) easier access to supercomputers, such as CRAY Y/MP, as a result of rapid advances in computer technology. Thus, ab initio methods have the potential for wider use beyond the tool of theoretical and computational chemists. In principle, such calculations can be used by a broad range of analytical and structural chemists for its accurate predictive power on structural, electronic,and spectroscopic properties of various atoms, radicals, ions, solids, and now larger polyatomics. In the present study, phenethylamine,tyramine, dopamine, and tyrosine, shown in Figure 1,are examined using the ab initio method. Even with the capability of the present supercomputer, calculations on biologically and pharmaceutically interestingmoleculea that are sip;nificantlylarger would still be difficult. However, it is expected that as computer software and hardware improve significantly over the next decade, these methods will be extended to even larger systems. Nonetheless, accurately determined parameters of these biomolecules provide an excellent building block for semiempirical calculations on larger biological systems. More significantly,ab initio methods can provide a broad range of information on biologically interesting systems. In this work, ab initio computational methods are used to obtain several stable conformers by optimizing the structure of biomolecules and calculating the corresponding vibrational frequencies. Vibrational frequency calculations are shown to provide relatively accurate predictionsof the IR and Raman spectra of these molecules where experimental data are available, and visualization of particular vibrational modes (28) Li, L.; Lubman, D. M. Anal. Chem. 1987,59, 2538-2541. (29) Johnson, C.R.;Ludwig, M.; Asher, S. A. J.Am. Chem. SOC.1986, 108,905-912. (30) Ludwig, M.;Asher, S. A. J.Am. Chem. SOC.1988,110,1006-1011. (31) Lee, N.-S.; Hsieh, Y.-Z.; Paisley, R. F.; Morris, M. D. Anal. Chem. 1988,60,442-446. (32) McGlashen, M. L.; Davis, K. L.; Morris, M. D. Anal. Chem. 1990, 62,846849. (33) Smith, A. Molecular Editor, software package for Apple Macintosh; Intellimation: Santa Barbara, CA. (34) Allan, D. S.;Seeger, D. M.; Korzeniewski, C. Appl. Spectrosc. 1990,44, 1579-1581.

0 C H 2 C H 2 N H 2

2727

PHENETHYLAMINE

TY RAMlNE

DOPAMINE

HO

0 CH,

0

-pH-OCII,H

TY R O S l N E

NH,

Figure 1. Phenethylamine and substituted phenethylamines.

using a software program, Molecular Editor,33J4 helps the assignment of the features observed in the IR spectra. In addition, the vibrational frequency calculations are also used to differentiate stable conformers (local minima in the potential energy surface) from transition states. Ab initio calculations can also provide a variety of information on electronic properties of these molecules. Within the context of Koopmans' theorem, ionization potentials (IP's) and electron affinities (EA'S) are predicted which will be very useful for studies of biomolecules by laser-induced fluorescence13J4 or multiphoton ionization spectroscopic techniques.2A13-18

COMPUTATIONALDETAILS The calculations for the present study have been performed available on CRAY Y/MP at using the Gaussian 90program the San Diego Supercomputer Center. A 3-21G split-valence basis set was used for all the calculations. Although it is the smallest basis set requiredfor freqency calculations, the predictive power of the calculations using this basis set is well documented3' and using a 6-31G basis set does not seem to change the results significantly. Calculations on phenethylamineand its derivatives using a basis set larger than 3-21G would be much too expensive and time-consuming even for the CRAY Y/MP. For the readers who are not familiar with ab initio methods, there are many excellent books and review articles a ~ a i l a b l e . ~ ~ . ~ ~ ~ (36) Frisch, M.J.; Head-Gordon, M.; Trucks, G. W.; Foreeman, J. B.; Schlegel, H. B.; Raghavachari,K.; Robb, M.; Binkley, J. S.;Gonzalez, C.; Defrees, D. J.; Fox, D:J.; Whiteeide, R. A.; Seeger, R.; Melius, C. F.; Baker, J.; Martin, R. L.; Kahn, L. R.; Stewart, J. J. P.; Topiol, S.;Pople, J. A. Gaussian 90,revieion F Gaussian, Inc.: Pittsburgh, PA, 1990. (36) Frisch, M. Gaussian90 User's GuideandProgrammer's Reference, revision I version; Gaussian, Inc.: Pittsburgh, PA, 1991. (37) Hehre, W. J.; Radom, L.; Schleyer, P.v.R.; Pople, J. A. Ab Initio Molecular Orbital Theory;John Wiley&Sona, Inc.: New York, 1986 (see also references therein). (38) For example, see: Pulay, P.; Lee, J.-G.; Boggs, J. E. J. Chem. Phys. 1983, 79, 3382-3391. (39) Simons, J. J. Phys. Chem. 1991,95, 1017-1029. (40) Szabo, A,; Ostlund, N. S. Modern Quantum Chemistry, revised 1st ed.; McGraw-Hill Co.: New York, 1989.

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ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1992

Table I. Relative Computational Time vs Basis Set for Phenethylamine Calculation no. basis re1 basis set functions computationaltime STO-3G 3-21G 6-31G* 6-31+G*

56 103 157 204

1 11 62 176

(A) Geometry Optimization, Most initial parameters of phenyethylamine were obtained from the experimental geometries of benzene and ethylamine. The two dihedral angles of phenethylamine which are responsible for the stability of the molecules were semiempirically determined values which were reported in the literature.7-9 For the optimization of the subsequent molecules (tyramine, dopamine, and tyrosine), the 3-21G optimized parameters of phenethylamine were adopted as the starting geometries and the additional parameters were approximated by using a reasonable guess obtained from other related molecules. All the optimization calculations were carried out using a Murtaugh-Sargent optimization algorithm instead of the default Berny a l g ~ r i t h m . The ~ ~ Murtaugh-Sargent method usually converges more slowly than the Berny method and thus takes a longer computational time, but occasionally, it performs more reliably, especially in the cases where the dihedral angles to be Due to computer time constraint, optimized are close to neither an exhaustive search for all the stable conformersnor the global search of the conformational surface for the most stable conformer have been attempted in the present study. (B) Vibrational Frequency. Calculated normal-mode vibrational frequencies provide two important pieces of information;37 first, they may be used to characterize stationary points on the potential energy surface, i.e., to distinguish local minima which have all real frequencies from saddle points which have a single imaginary frequency, and second, they may be used as an aid in assigning fundamental vibrational modes observed in the IR and Raman spectra. Vibrational frequency calculations were performed at the HF/3-21G level using the 3-21G optimized parameters. Atotal of 3n-6 normal-modevibrational frequencies are predicted which are typically about 8-15% larger than the experimental values.37 This offset is due to the effects of anharmonicity of the potential energy surface near the local minimum and the deficiencies of the HF procedure. The latter deficienciescan be removed by including polarization functions to the basis set and by including electron correlation.% Therefore, when comparing the calculated and experimental frequencies, it is necessaryto "scale down" the calculated values by about 10 % .37 A checkpoint file saved from the optimization procedure was used as a starting point for the frequency calculation in order to save some computational time. (C) Computational Time. When the electronic SchrMinger equation is actually solved, evaluation of two-electron integrals is the most time-consuming step. The computational time required for a single cycle of HF energy calculation is approximately proportional to n4, where n is the number of basis functions used in the calculation.a*37 The computational time for this step can be significantly reduced if the chemcial species under consideration is highly symmetrical since the number of two-electron integrals to be evaluated is reduced. Table I shows the number of basis functions and the approximate relative computational time for single-point H F calculations of phenethylamine in which several basis seta were used. It is estimated that the computational time for single-point H F calculations on phenethylamine using 3-21G, 6-31G*, and 6-31+G* basis sets is 11,62,176timeslonger,respectively,thanthatusing theminimum STO-3G basis set. A single-point H F calculation on phenethylamine using the 3-21G basis set takes slightly less than 10 min of CRAY Y/MP CPU time, thus showing that the calculations, particularly optimization calculations, using a basis set larger than 3-21G quickly become excessively expensive and time(41)Schaefer,H. F. Quantum Chemistry: the development ofab initio methods in molecular electronic structure theory; Oxford University Press: New York, 1984.

Table 11. Stable Conformers, Their Dihedral Angles, and the Total Energies compd dl d2 ET (Hartree) AE(eV) Phenethylamine 90.0 folded -63.3 -361.7716 90.0 folded 63.3 -361.7716 90.0 -361.7713 0.01 extended 180.0 180.0 180.0 0.11 TS -361.7685 Tyramine 90.0 -62.7 -436.2117 folded 90.0 62.7 -463.2 117 folded extended 90.0 179.9 -436.2113 0.01 0.0 TS 61.3 -436.2073 0.20 Dopamine folded(a)a 90.0 -63.3 -510.6559 90.0 folded(a)a 63.3 -510.6559 90.0 extended(a)a 179.8 -510.6555 0.01 -510.6556 co.01 folded(b)" -510.6444 0.31 folded(c)a Tyrosine 90.0 -69.3 folded -622.7777 (a),(b),and (c) refer to theorientationsofthe two hydroxygroups in dopamine. See Figure 5a-c, respectively, for the corresponding configurations.

kH

H

H

H/

'H

Figure 2. 3-210 optlmized phenethylamine.

consuming. A 3-21G vibrational frequency calculation on phenethylamine using the Gaussian 90 program on CRAY Y/MP required approximately 30 CPU min.

RESULTS AND DISCUSSION (A) Structures of Stable Conformers. Table I1 summarizes the results of the optimization calculations performed to identify some of the stable conformers and the transition states of phenethylamine, tyramine, dopamine, and tyrosine. Three stable conformers each were found for phenethylamine and tyramine whereas five stable conformers were found for dopamine. Only one stable conformer was located for tyrosine via these computational methods. As mentioned in the previous section, no attempts have been made to locate the global minimum for all the molecules studied here. The two relevant dihedral angles which are mainly responsible for the stability of the molecule and the total energy of each of the conformers are also presented in Table 11,as well as the energy difference with respect to the most stable conformer identified in the present study. In the followingdiscussion, the structure and atomic positions in each molecule are described by referring to the phenethylamine structure shown in Figure 2 which forms the basic structure for all the molecules considered in the present study. (i) Phenethylamine. Three stable conformers have been found for phenethylamine, two of which are equivalent and mirror image to each other. Figure 2 shows the 3-21G optimized structure of one of the two more stable pheneth-

ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1992 0'

C8H2NH2

c3

hc5 H

A-? H (a) d l

-

HH*

-

-90.0'

(bl) d2 83.3'

quency calculations whereas Carbo et al. performed singlepoint ab initio calculations on the possible conformers as determined by semiempirical methods, and (3) the NMR experimental results of amphetamine in D2O cannot be used to support the results of ab initio calculations because phenethylamine is present in different chemical environments. More specifically, the ab initio calculations were performed on free phenethylamine whereas the NMR experiment was performed on amphetamine in D2O where solvation effects may be important. In the crystal form of phenethylamine, only the ethylamine-extended form seems to dominate,possibly because it leads to more efficient crystalpacking forces.9~~~ However, in solution,both the ethylamineextended and -folded forms coexist.9 The results of the present study are in agreement with those of the previous study using PCILO in which three stable conformers were f0und.7.8~11The present study determines the d2 dihedral angle more precisely and indicates that there is a small energy difference (about 0.01 eV) between the ethylamine-folded and ethylamine-extended conformers. Whether this represents the real situation or the consequence ofusingarelativelysmall3-21Gbasissetcannotbedetermined by the present study. To clarify this point, more extensive calculations with basis sets more flexible than 3-21G and perhaps the inclusion of a correlation effect will be required. In addition to the three stable conformers described above, a transition state was located for phenethylamine which yields one imaginary frequency in the vibrational frequency calculation. In this structure, d l = d2 = 180.0', which is equivalent to having all the carbon and nitrogen atoms on the same plane as the benzene ring. This transition state lies 0.11 eV (about 2.8 kcal/mol) in energy above the two most stable ethylamine-folded conformers. (ii) Tyramine, Dopamine, and Tyrosine. Tyramine is a substituted phenethylamine where a hydroxy group on the benzene ring is in the para position (Cl) to the ethylamine chain. As in the case of phenethylamine, three stable conformershave been identified for tyramine. Figure 4 shows a snapshot of the most stable conformer of tyramine found in the present study with the ethylamine chain folded. All 3-21G optimized parameters of tyramine do not differ significantly (less than 0.2 A and 1.0' for bond distance and angle, respectively) from those of phenethylamine. Using the same definition of dihedral angles as in phenethylamine, the two equivalent mirror-image conformers have d l = 90.0' in common and d2 = +62.7 and -62.7'. These conformers are slightly more stable by 0.01 eV than the conformer with d l = 90.0° and d2 = 179.9'. The first two conformers correspond to the ethylamine-folded form while the third conformer corresponds to the ethylamine-extendedform. The semiempirical calculations on tyramine using the PCILO method found two ethylamine-folded conformers at the two dihedral angles, d l = 90' and d2 = +60 and =60°, approximately in agreement with the result of the present study? But for the ethylamine-extended conformer, the PCILO method found d l and d2 to be 120 and 180°, respectively, as compared to 90 and 179.9' observed in the present study. Other optimized parameters specific to tyramine are OC1= 1.38 A, HO = 0.96 A, OClC2 = 117.3', and HOC1 = 112.8'. A transition state has been located at d l = ' 0 and d2 = 61.3', which is equivalent to rotating the ethylamine chain of a stable conformer along the C 7 4 4 axis by 90' counterclockwise. The energy difference between the most stable conformers found and this transition state is 0.20 eV (about 4.6 kcal/mol). Dopamine is a substituted phenethylamine where two hydroxy groups are in the para (Cl) and meta (C6) positions

'eNH2 0

0

'

-

H

(b2) 62

-63.3'

d

e

H

NH2 (b3) d2

-

180.0'

Flguro 3. Newman prolections of phenethylamine conformers along the (a) C7-C4 and (b) CGC7 axes.

ylamine conformers found. Carbon atoms are arbitrarily numbered 1through 8 and the nitrogen and hydrogen atoms are represented by N and H s , respectively. The bond angles and distances are represented in degrees and angstroms, respectively. The parameters of the benzene ring do not seem to change significantly from one conformer to another and the stability of the structure depends strongly on how the ethylamine chain is arranged. This can.be more clearly elucidated by referring to Figure 3 where the two relevant dihedral angles, d l and d2, are shown in Newman projections along (a) C7-C4 and (bl-3) C&C7 axes, respectively. The dihedral angles d l and d2 correspond to the angles C8C7C4C3 and NC8C7C4, respectively. Parta b l and b2 of Figure 3 correspond to the two most stable conformers found with d2 = +63.3 and -63.3', respectively. The sign of the dihedral angle depends on whether the front axis (N-CS) is rotated clockwise (+) or counterclockwise(-) in order tosuperimpose it to the back-axis (C7-C4), as indicated in the figure by the arrows. Figure 3b3 represents another stable conformer with d2 = 180.0°,which is slightly less stable (about 0.01 eV) with respect to the other two conformers. The ethylamine chain in the first two more stable conformers is folded whereas the one in the third conformer is extended. Figure 4 shows a snapshot of the more stable conformer of phenethylamine with the ethylamine chain folded. Previous studies on phenethylamine show contradicting results on the structure of its most stable conformer.ll Whereas the calculations using the semiempirical methods ( 0 0 ,INDO, and PCILO) identified the ethylamine-folded form as the most stable conformer for phenethylamine, the EHT calculations showed the ethylamine-extended form as the most stable conformer, which is in agreement with the NMR experimental results where amphetamine in D2O was shown to be in the ethylamine-extended form.I1 Carbo et al.11 performed ab initio calculations on phenethylamine at the Hartree-Fock level using the minimum STO-3G basis set and, on the basis of their results, concluded that the most stable conformer was the ethylamine-extended form, in agreement with the results of the EHT calculations. This is in constrast with the results obtained in the present study where the ethylamine-foldedform is shown to be slightly more stable than the ethylamine-extended form. There are several reasons to believe that our results are more reliable than Carbo's: (1)the calculations in the present study used the split-valence 3-21G basis set which describes the nonsphericalanisotropic aspects of molecular charge distribution more adequately than the minimum STO-3G basis set which Carbo et al. used in their calculations,(2) for all the optimized structures considered in the present study, the stability of the conformer was verified by performing vibrational fre-

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(42) Tsoucaris, P.G. Acta Crystallogr. 1961, 14, 909-914.

(a) PHENETHYLAMINE

(b) TYRAMINE

(c) DOPAMINE

(d) TYROSINE

H-J

Flguro 4. Snapshots of the stable conformer of (a) phenethylamine, (b) tyramine, (c) dopamine, and (d) tyrosine.

to the ethylamine chain on the benzene ring. A total of five stable conformers were identified for dopamine. As shown in Figure 5, there are three possible configurations for the two hydroxy groups in dopamine. Optimization of the two dihedral angles have been performed on the configuration of Figure Sa which has the lowest energy of the three possible configurations. In accord with the cases of phenethylamine and tyramine, the dopamine with the configuration of Figure 5a has two stable conformers which are isoenergetic and mirror image to each other. These two conformers correspond to the ethylamine-foldedstructure with d l = 90.0° in common and d2 = +63.3 and -63.3". Other optimized parameters specific to this stable conformer are OC1= 1.39 A, H 0 1 = 0.96 A, OC6 = 1.37 A, H 0 2 = 0.97 A, OClC6 = 114.3*, OC6C5 = 120.6O, HOlCl = 113.5O, and H02C6 = 110.1O. As in the cases of phenethylamine and tyramine, all 3-21G optimized parameters common to both tyramine and dopamine do not change significantly (less than 0.1 A and 1.0" for bond distance and angle, respectively). Another stable conformer of dopamine with Figure 5a configuration was found with d l = 90.0° and d2 = 179.8", corresponding to the ethylamine-extended structure. This conformer is slightly higher in energy (about 0.01 eV) than the other two conformers. Using the optimized ethylamine-folded structure as a starting geometry, optimization calculationswere performed on dopamine with the configurations of Figure 5b,c. There is almost no energy difference between the optimized Figure 5a,b (less than 0.01 eV) whereas optimized Figure 5c is about 0.31 eV (7.1 kcal/mol) higher in energy than the optimized Figure 5a. From the symmetry consideration, it is reasonable

(a)

(b)

Y Y

\

o

Y

(c) H\O

H

\H

Flguro 5. Possible configurations for hydroxy groups in dopamine.

ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1992

Table IV. Predicted Vibrational Frequencies, IR Intensities, and Raman Activities of Tyramine freq IR Raman freq IR YN (cm-1) inten act. YN (cm-1) inten

H

Flgure 6. 3-210 optimized tyroslne.

Table 111. Predicted Vibrational Frequencies, IR Intensities, and Raman Activities of Phenethylamine freq IR Raman freq IR Raman (cm-1) inten act. a (cm-1) inten act. 1 2 3 4

5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27

3397 3309 3044 3032 3025 3015 3010 2941 2903 2885 2851 1659 1597 1577 1512 1504 1484 1464 1397 1369 1342 1318 1244 1199 1192 1189 1145

1 0 7 15 8 0 2 10 27 9 29 16 2 1 1 7 3 6 6 1 3 2 1 0 1 0 0

29 52 100 3 32 39 8 20 48 37 32 4 10 4 7 1

5 0 1 1 6 6 1 3 4 1 2

28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54

1117 1076 1067 1041 1039 1029 1004 995 973 894 875 842 790 746 733 711 642 584 495 424 417 340 266 256 148 98 45

1 3 0 6 3 1 1 0 0 0 0 0 16 4 22 100 0 6 4 0 4 0 15 34 3 1 2

1 0 0 3 2 0 1 14 1 2 2 3 1 6 0 2 2 1 2 0 1 0 6 2 1 1 3

to speculate that there are more stable conformers for dopamine with the hydroxy groups in either Figure 5b or 5c configuration. Figure 4 shows a snapshot of the most stable conformer of dopamine found with the ethylamine chain folded. Tyrosine is a substituted tyramine where the -COOH group is attached to the ethylamine chain at the C8 position. Structural optimization of tyrosine was a nontrivial matter due to the increased number of parameters compared with the other molecules in the present study. The 3-21G optimized stable conformer is shown in Figure 6 and a snapshot of this conformer in Figure 4. Using the same definition of dihedral angles as in phenethylamine, d l = C8C7C4C3 = 90.0° AND d2 = NC8C7C4 = 69.3O, which correspond to the ethylamine-folded structure. Other dihedral angles specific to tyrosine are M 9 C 8 C 7 = -114.7O and OC9C8C7 = 63.9'. Bysymmetry, there should beanother stable conformer with d2 = -63.9O, as well as other stable conformers,which have not been explored due to the computer time constraint. (B)Vibrational Frequencies. Tables I11 and IV show the predicted vibrational frequencies, their relative IR intensities, and Raman activities for phenethylamine and tyramine, respectively. The IR intensities and Raman activities are normalized to u43 at 711 cm-1 and u3 at 3044 cm-l, respectively, for the vibrational frequencies of phen-

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29

3523 3395 3308 3045 3038 3020 3020 2938 2901 2885 2851 1658 1615 1579 1523 1511 1485 1438 1397 1360 1343 1319 1261 1238 1193 1189 1177 1145 1103

28 1 0 2 6 5 7 11 32 13 31 17 18 7 36 2 3 12 8 6 4 4 2 29 1 3 72 3 11

87 46 81 100 43 36 26 32 83 56 50 6 19 5 1 10 10 0 1 1 10 10 2 2 4 1 9 3 0

30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57

1101 1045 1041 1024 1020 1004 901 897 870 833 812 770 718 694 661 574 493 438 424 409 362 291 271 257 212 128 82 41

2731

Raman

act. 1 5 2 1 0 2

66 2

5 7 1

5

0 3 4 4 18 0 3

31 1 8 12 9 1 100 4 1 11 2 1 4 8 3 6 97 45 2 3 1 3

1 3 0 1 0 3 1 2 1 3 3 0 0 1 4

I

100,

I/

Ill I I

u x D 3 6 0 0 3 o o o ~ 2 o o o 1 5 0 0 1 o o o 6 0 0

0

WAVENUMBER (an-1)

Flgure 7. Predlcted I R spectrum of phenethylamlne. Dotted llnes lndlcate the locatkins of the experlmental peaks.

ethylamine in Table 111,and to ~ 4 at 2 718 cm-l and u4 at 3045 cm-l, respectively, for the vibrational frequenciesof tyramine in Table IV. Although frequency calculations predict only the fundamentals (3n - 6 in the case of nonlinear molecules with n atoms), it is quite useful for the assignments of the experimental spectra which include not only fundamentals but also overtone and combination bands. It is well-known that simple vibrational frequency calculations such as the ones used in the present study predict the in-plane vibrational modes more accurately than the out-of-plane vibrational modes which are extremely sensitive to the basis set used in the calculation.43 Inclusion of electron correlationeffects may improve the prediction significantly,particularly for the outof-plane vibrational mode calculations. Figure 7 shows a simulated IR spectrum of phenethylamine which includes only the fundamental vibrational modes. In this figure, the locations of the experimental vibrational frequencies are indicated with the dotted lines in the middle of the spectrum (43) Barstis, T.; Lubman, D. M. Unpublished resulta, University of Michigan, 1992.

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ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1992

I

0 h

Table V. Calculated Molecular Orbital Energies (eV) of Phenethylamines I

compd

SHOMO

HOMO

LUMO

SLUMO

phenethylamine tyramine dopamine tyrosine

-9.16 (9.35)" -9.39 (9.35)" -9.09 (8.90)" -9.30

-8.84 (8.99)" -8.32 (8.41)" -8.18 (8.18)" -8.33

4.21 3.95 4.08 4.01

4.15 4.48 4.50 4.37

a

The experimental IP's are in the parentheses. From refs 46 and

47.

0 I

I

Flgure8. Visualization of a vibrationalmode of phenethybmineobtained by using the Molecular Edltor program.33

for comparison purposes. (The relative intensities are not included). Although not exactly identical, mostly due to the presence of the overtone and combination peaks, the approximate regions where major fundamental peaks appear are quite well predicted. The experimental IR spectrum of phenethylamine in the vapor phase contains nine major peaks at v = 3037 (m), 2943 (s),2860 (m), 1613 (m), 1500 (m), 1454 (m), 1075 (m), 744 (s), and 699 (s) cm-1,M as indicated in the simulated IR spectrum by the dotted lines in Figure 7. By comparing the experimental and theoretical frequencies and intensities of the peaks, it is possible to speculate on the tentative assignment of the peaks in the experimental IR spectrum. For example, the most intense peak in the experimental IR spectrum of phenethylamine at v = 743 cm-1 is most likely the fundamental v43 which is predicted at v = 711 cm-l as the most intense peak. Also, the three closely spaced peaks at v = 3037,2943, and 2860 cm-l observed in the 3050-2850-cm-l region of the experimental IR spectrum probably correspond to the three peaks predicted at v3 = 3032 cm-l, vg = 2903 cm-l, and v11 = 2851 cm-1, respectively, as can be deduced from the spacings and the intensities of these peaks. In order to make a more definitive assignment of the features in these spectra, it is often quite useful to be able to actually visualize a specific vibrational mode that is responsible for a particular peak. Theoretical vibrational modes can be visualized by using the Molecular Editor software program333which graphicallyrepresents a mode by oscillating between the two extremes of vibrational motion. The two extremes of a particular vibrational mode are calculated by adding and subtracting the nuclear displacements from the equilibrium nuclear positions, all of which is available from the Gaussian output. This capability is especially useful when there are ambiguities in assigning the peaks that are closely spaced. Figure 8, for example, shows the vibrational mode v43 predicted to be the most intense peak a t 711 cm-l. As can be seen from Figure 8a,b, which represent the two extremes of vibrational motion, the peak at v u = 711 cm-l is mainly due to the rocking motion of the -NH2 group. On a computer, (44)Pouched, C. J. The Aldrich Library of FT-IR Spectra Vapor Phase, ed. I; Aldrich Chemical Co., Inc.: Milwaukee, WI, 1989;Vol. 3, p 1164.

the molecule can be rotated in three dimensions which enhances the visualizationof the vibrational mode responsible for a particular peak in the spectrum. Due to experimental difficulties, the Raman spectra of phenethylamine and substituted phenethylamines in the gas phase are not found in the literature. When the experimental Raman spectra of these compounds become available in the future, the predicted vibrational frequencies and Raman activities presented in Tables I11 and IV will be particularly useful for the analyses of the experimental data. For phenethylamine and tyramine, the most intense peaks are predicted to be at v = 3044 and 3045 cm-l, respectively, when scaled down by 10% from the calculated values. (C) Ionization Potentials and Electron Affinities. Ionization potentials (IP's) and electron affinities (EA'S) are two of the most fundamental properties of chemical species which are the key to understanding oxidation and reduction processes as well as for estimating their electronegativities. The ionization potential is particularly important for experiments in photoionization or multiphoton ionization spectroscopies.192 The simplest way to estimate these properties is by invoking Koopmans' theorem45in which the IP and EA are associated with the negatives of the energies of the orbitals the electron is ejected from and captured into, respectively. Since the orbital energies are obtained from a HartreeFock calculation on the neutral species, the Koopmans' theorem approximation includes neither the relaxation of the rest of the electrons upon gain or loss of an electron nor the difference in correlation energies between the neutral and ionic species. IP's are predicted much better than EA'S by invoking Koopmans' theorem in the HF calculations. The fact that IP's are accurately predicted, often within a few tenths of an electronvolt, is actually quite fortuitous due to the relative magnitudes and sign of relaxation and correlation effects in neutrals and the corresponding cations. Table V shows the energies of the SHOMO (second highest occupied molecular orbital), HOMO (highest occupied molecular orbital), LUMO (lowest unoccupied molecular orbital), and SLUMO (second lowest unoccupied molecular orbital) of the most stable conformers of phenethylamine, tyramine, dopamine, and tyrosine. These energies,expressed in electronvolts (eV),were calculated at the HF/3-21G level and obtained by invoking the Koopmans' theorem. The negatives of the energies of the SHOMO, HOMO, LUMO, and SLUMO correspond to the second IP (ionizationpotential), first IP, first EA (electron affinity), and the second EA of the molecules, respectively. The experimental ionization potentials of the compounds obtained by photoelectron spectro~cop?~*~~ are also included in Table V in parentheses. Considering the fact that the experimental deviations in the photoelectron spectra are typically 0.12-0.20 eV,46947the ionization potentials of phenethylamine, tyramine, and dopamine are quite accurately predicted within the margin of the experimental deviation at (45) Koopmans, T. Physica 1934,1,104-113.

(46)Domelsmith, L.N.;Munchausen,L. L.; Houk, K. N. J. Am. Chem. SOC.1977,99,4311-4321. (47) Domelamith, L. N.; Houk, K. N. Int. J. Quantum Chem., Quantum Biol. Symp. 1978,5,257-268.

ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1902

this level of calculation. It appears from Table V that the first ionization potentials are somewhat more accurately predicted by the calculation than the second ionization potentials. Although not available in the literature, the first and second ionization potentials of tyrosine are predicted to be 8.33 and 9.30 eV, with its first ionization potential quite close to that of tyramine. In contrast to the accurate prediction of the ionization potentials, the prediction of electron affinities of these molecules is not expected to be quantitatively accurate due to the additiveness of relaxation and correlation effects in neutrals and correspondinganions. However,it is well-known that among the related molecules, relaxation and correlation effects often remain nearly constant so that the HF calculations invoking Koopmans' theorem approximation can predict the relative eledron affinitiesquite well when adjusted with a constant scaling fador.@ Nevertheless, one has to be cautious when applying Koopmans' theorem to predict the electron affiiities of the molecules since there are complications associated with the fact that the unfilled orbitals corresponding to electron affinities are usually buried in the continuum region. Therefore, as more diffuse basis seta are used for calculations, the low-lying unoccupied orbitals may actually correspond to discretized continuum states, not the anion s t a t e ~ . ~ + ~Since 1 extensive discussion on this subject is beyond the scope of this paper, interested readers are referred to the references for detail.4'62 Although no experimental data are availableat the present time for phenethylamine and ita derivatives considered in the present study, it can be predicted that the magnitude of the fiit electron affiiities will be in the order phenethylamine > dopamine > tyrosine > tyramine, as shown in Table V. Once the electron affinity of one of these molecules is determined experimentally, for example by using electron transmission spectroscopy,52then on the basis of the scaling factor between the calculated and experimental values, the electron affiiities of the rest of the related molecules may be predicted quantitatively. (D) Total Atomic Charges. Parta a and b of Figure 9 show the total atomic chargesof phenethylamineand tyrosine, respectively, which were obtained from the Mulliken population analyses of the HF/3-21G ~alculations.3*3~Figure 9a also includes the atomic charges obtained from the HF/STO3G calculations in parentheses.ll When relative atomic charges are compared, there is little difference between the two calculations except for the carbon atom, C4, which is predicted to be negatively and positively charged according to the 3-21G and STO-3G calculations,respectively.11 Other carbon atoms on the benzene ring (C144, C5, and C6) are all negatively charged with similar magnitudes. A carbon atom on the ethylamine group, C7, is negatively charged and the other carbon atom, C8, is less negatively charged due to a larger electronegativity of the neighboring nitrogen atom which is the most negatively charged atom of the compound. In tyrosine, shown in Figure 9b, C1 is positively charged due to the neighboring oxygen atom and the carbon atom C9 is also positively charged due to the two neighboring oxygen atoms in the carbonyl group. Sincethe nitrogen atom is more negatively charged than the three oxygen atoms present in the compound, other conditions being considered equal, the nitrogen is the most likely atom predicted to be protonated. (48)For example, see: Balaji, V.; Ng, L.; Jordan, K. D.; Paddon-Row, M. N.; Patney, H. K. J. Am. Chem. SOC.1987,109,6957-6969. (49) Chao, J. S.-Y.; Falcetta, M. F.; Jordan, K. D. J.Chem. Phys. 1990, 93, 1125-1135. (50) Falcetta, M. F.;Jordan, K. D. J. Phys. Chem. 1990,94,5666-5669. (51) Falcetta, M. F.; Jordan, K. D.J. Am. Chem. SOC.1991,113,29032909. (52) Jordan, K. D.; Burrow, P. D. Acc. Chem. Res. 1978,11,341-348.

ma

0.30

(0.W

0.24 H

0.21 (0.02)

(4.08)

/

H 0.23 (0.07)

(a) TOTAL ATOMIC CHARGES OF PHENETHYLAMINE 0.32 ti

\

\ 0.27

0.25

0 26

(b) TOTAL ATOMIC CHARGES OF TYROSINE

Figure 9. Total atomic charges of (a) phenethylamineand (b) tyrosine.

It is very important to obtain information concerning the protonated form of a molecule in order to understand the structureactivity relationship. In addition to being the most abundant form at the physiological value of pH = 7.4, the protonated form is the main form of a molecule responsible for its pharmacologicalresponses by means of ionic interaction between the cationic head and some anionic receptor site." In order to further investigate these problems, studies on protonated forms of phenethylamine and substituted phenethylamines are in progress.

CONCLUSIONS The present study demonstrates the utility of the ab initio computational method to study the structural and spectroscopicproperties of biomoleculeswhich are frequentlydifficult to determine experimentally. Specifically, phenethylamine and its derivatives are pharmaceutically and biologically interesting molecules which have been studied by various semiempirical methods. Using the ab initio method, structural and spectroscopic properties of these biomolecules have been predicted and compared with those obtained by semiempirical methods. Several stable conformers were identified for phenethylamine, tyramine, and dopamine, and their relative energies were predicted. In the case of phenethylamine and tyramine, transition states were also identified. In particular, two ethylamine-folded (dl = 90.0° and d2 Y 60.0°) and one ethylamine-extended (dl = 90.0° and d2 = 180.0°) stable conformerswere found for both phenethylamine and tyramine, with the first two about 0.01 eV lower in energy than the last one. While there is general agreement between the results of ab initio calculations presented in this study and those by several semiempirical methods, the ab initio calculations yield more precise values for the two dihedral angles that are responsiblefor the stability of the conformers.

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ANALYTICAL CHEMISTRY, VOL. 64, NO. 22, NOVEMBER 15, 1992

IR vibrational frequencies were reasonably well predicted when the calculated frequencies were scaled down by 10% to compensatefor the deficiencies of the computational method used in the present study. The capability to visualize a vibrational mode responsible for a particular peak enhances ita usefulness for the assignment of the features in the IR spectra. Invoking Koopmans’ theorem, the experimental ionization potentials were quite accuratelypredicted whereas calculated electron affinities need to be scaled in order to predict the experimental values. It is important to remember that, when comparing theoretical predictions with the experimental data, one has to take into consideration the environment of the molecule. For example, whereas the calculations are performed on molecules in their free forms, approximately equivalent to their being in the gas phase, molecules studied experimentally are often present in highly perturbed condensed-phase environments in which solvation effects (NMR)or crystal-packingefficiency

(X-ray crystallography) may be important. In these cases, direct comparisons between the experimental data and the theoretical predictions are not possible. In order to further investigate some of these problems, studies on protonated phenethylamine and substituted phenethylamines are in progress.

ACKNOWLEDGMENT We would like to thank C. Korzeniewski for helpful suggestions regarding the Molecular Editor program. We acknowledge financial support of this work under NSF Grant No. CHE 9022610. Most of the computations in the present study were performed on CRAY YIMP at the San Diego Supercomputer Center.

RECEIVED for review March 17, 1992. Accepted August 6, 1992.