2251
J. Phys. Chem. 1991,95,2251-2256
Solvatochromism and Time-Resolved Fluorescence of the Antitumor Agent Mitoxantrone and Its Analogues In Solution and in DNA Su Lin and Walter S . Struve*
Department of Chemistry and Ames Laboratory-USDOE, Iowa State University, Ames, Iowa 5001 1 (Received: September I I, 1990)
The electronic spectroscopy and fluorescence kinetics of 1,4-dihydroxy-5,8-[2-[2-[(2-hydroxyethyl)amino]ethy~]amino]9,IO-anthracenedione (mitoxantrone) and three closely related analogues have been studied in several solvents. The small solvatochromic blue shifts of their visible charge-transfer absorption bands in protic solvents are dominated by interactions with a solvent H-bonding donor, rather than by dipoledielectric solutesolvent electrostatics. These interactions are unrelated to the phenolic hydroxy groups or the distal N atoms on the side chains but must be localized to the carbonyl groups. The fluorescence decays of all four anthraquinones are controlled by subnanosecond nonradiative relaxation in all solvents studied. At least two decay mechanisms contribute to the observed fluorescence kinetics in solution: (a) subnanosecond internal conversion that is accelerated relative to that in 1,4-diaminoanthraquinoneby the presence of the flexible 1 ,rl-side chains in mitoxantrone and its analogues; (b) an additional decay mode that is accentuated in H-bonding solvents. A substantial normal isotope effect occurs in the fluorescence lifetimes of mitoxantrone in perdeuterated water and methanol but not in aprotic solvents. When bound to double-stranded calf thymus DNA, mitoxantrone displays a fluorescence lifetime similar to that in aprotic solvents, suggesting that H-bonding interactions with water are precluded by chromophore intercalation. DNA-bound ametantrone exhibits a lifetime longer than that in either H-bonding or aprotic solvents, indicating that immobilization of the side chains through binding of the distal N atoms to the DNA backbone may influence the decay kinetics. This technique therefore shows potential for elucidating the DNA binding modes for a large class of intercalative drugs.
Introduction The antitumor agent mitoxantrone (1,4-dihydroxy-5,8-bis[2[2-[(2-hydroxyethyl)amino]ethyl]amino]-9,lO-anthracenedione) is a substituted anthraquinone that has undergone phase I and phase I I clinical trials after proving effective in several tumor model systems and is now approved for clinical u ~ e . l - ~While the basis of its pharmacological activity is unclear, it is known to bind efficiently to DNA$,' probably through intercalation* of the flat anthraquinone chromophore between adjacent base pairs. It becomes preferentially attached to DNA and RNA in intact cells, unwinds covalently closed circular DNA, and effectively inhibits DNA repli~ation.~Formation of mitoxantrone-DNA adducts thus appears crucial to its clinical efficacy. The electronic spectroscopy of mitoxantrone and its analogues has been relatively little investigated, despite the need for bioanalytical assays of this drug. Its static electronic spectroscopy was recently studied by Lee and Dutta,Io who also obtained resonance Raman spectra for excitation wavelengths in the visible and near-UV regions. The origin band in the lowest energy absorption system (located at -682 nm in CHCIJ exhibits small solvatochromic blue shifts in polar solvents. This visible band system (which is absent in the unsubstituted anthraquinone but appears in I ,4-diaminoanthraquinone1I) has been assigned to an (1) Von Hoff, D. D.; Pollard, E.; Kuhn, J.; Murray, E.; Coltman, C. A. Cancer Res. 1980, 40, 1516. (2) Anderson, K. C.; Cohen, G. 1.; Garnick, M. B. Cancer Treat. Rep. 1982, 66,1929. 13) . , .la), Wallace. R. E.; Murdock. K. C.; Anaier. R. B.; Durham. E. Cancer Res. 1979, 39, 1570. (b) Murdock, K. C.; Crhild, R.;Fabio, P. F.; Angier, R. B.; Wallace, R. E.; Durr, F. E.; Citarella, R. V. J. Med. Chem. 1979, 22, 1024. (4) Von Hoff, D. D.; Coltman, C. A.; Forseth, B. Cancer Res. 1981.41, 1853. (5) Drewinko, B.; Yang, L. Y.; Barlogie, B.; Trujillo, J . M . Cancer Res. 1983, 43, 2648. (6) Lown, J. W.: Morgan, A. R.; Yen, S.-F.;Wang, Y.-H.; Wilson, W. D. Biochemisfry 1985, 24, 4028. (7) Rasenberg, L. S.;Carvlin, M. J.; Krugh, T. R. Biochemisrry 1986, 25, 1002. (8) Islam, S. A.; Neidle, S.;Gandecha, B. M.; Partridge, M.; Patterson, L. H.; Brown, J . R. J . Med. Chem. 1985, 28, 857. ( 9 ) Karpuscinski, J.; Darzynkiewicz, Z.; Traganos, F.; Melamed, M. R. Biochem. Pharmacol. 3981, 30, 23 I . (IO) Lee, B. S.; Dutta, P. K. J. Phys. Chem. 1989, 93, 5665.
CHART I
1 0
NHCH2CH2OH
Q
YHCH,CH,NHCH&H,OH
8
NHCH~CH~NHCH~CH~OH
OH
0
NHCH2CHaHCH,CH2OH
OH
0
NHCH2CHpNHCHsCHpOH
OH
0
NHCH2CH2N(CHS)a
OH
2
3
4
aminq-ring charge-transfer transition.1° Similar blue shifts appear in mitoxantrone's fluorescence band maximum. Since the analogous charge-transfer absorption band for 1,4-diaminoanthraquinone exhibits red shifts in polar solvents,'* the blue shifts in mitoxantrone have been ascribed to solvent H-bonding interactions with the phenolic hydroxy groups in the ground-state molecule.'0 A second peculiarity of mitoxantrone, reported in a recent chara~terization'~ of its fluorescence spectral shifts and depolarization in DNA environments, is its low fluorescence quantum yield. This has been attributed to mitoxantrone dimerization and/or aggregation:I0 the lower of the two excitonic ( 1 1) Inoue. H.: Hoshi. T.; Yoshino, J.; Tanizaki. Y. Bull. Chem. Soc. Jpn. 1972, 45, 1018. (12) Sinclair, R. S.;McAlpine, E. J. J. Soc. Dyers Colour. 1975, 91, 399. (13) Bell, D. H. Biochim. Biophys. Acra 1988, 949, 132.
0022-3654/91/2095-2251 $02.50/0 0 1991 American Chemical Society
2252 The Journal of Physical Chemistry, Vol. 95, No. 6, 1991
Lin and Struve TABLE I: Absorption a d Fluorescence Band Maxima (iuaoneters) for Mitoxantrone (3) and A n a l o w
1.0
w
comvd
0 2
a m a
0
8 a 0.0 500
600
700
WAVELENGTH (nm) Figure 1. Absorption spectra of 1 in (from left) methanol, acetone, and chloroform, solid curves; absorption spectra of ametantrone (2) in (from
left) water, methanol, acetone, and chloroform, dashed curves. states in an excited dimer frequently carries minimal oscillator strength to the ground-state dimer,I4 with the result that dimers and higher aggregates are essentially nonfluorescing. In this paper, we analyze solvent effects on the static absorption and time-resolved fluorescence spectroscopy of the four compounds 5,8-dihydroxy- 1,4-bis[ [2-hydroxyethyI]amino]-9,10anthracenedione (I), ametantrone (2), mitoxantrone (3), and 5,8-dihydroxy- 1,4-bis[[ 2-(dimethylamino)ethyl] amin0]-9,10anthracenedione (4, Chart I). This sequence of molecules isolates the effects of the phenolic hydroxy groups, the substituent chain length, and the presence of amino hydrogens and the distal nitrogen atoms. Emission profiles were obtained for each analogue in the same sequence of solvents by using time-correlated single-photon-counting spectroscopy. Finally, we describe time-resolved fluorescence studies for mitoxantrone bound to doublestranded DNA.
Experimental Section Compounds 1,2 (as the acetate salt), 3 (as the hydrochloride), and 4 were generously provided by K. C. Murdock of Lederle Laboratories, American Cyanamid Co. None of the solvents used exhibited background emission in steady-state or time-resolved studies: triply distilled deionized water, methanol (Fisher ACS certified >99.9%), cyclohexane (Aldrich), and acetone, ethyl acetate, DMSO, and CHCl3 (OmniSolv spectrophotometric grade). Double-stranded calf thymus DNA was purchased from Sigma Chemical Co. Absorption spectra were obtained on a Perkin-Elmer Lambda 3B UV-vis spectrophotometer; static fluorescence spectra were evaluated on a Spex Fluorolog 2 spectrofluorometer with 4,6-nm excitation and detection bandpass. The time-correlated single-photon-counting apparatus has been described previo~sly.'~The Canberra Series 30 multichannel analyzer (MCA) was superseded by an EG&G-Ortec ACE MCA plug-in board/lBM compatible computer combination. A synchronously pumped rhodamine 590 dye laser (Coherent CR599) was cavity-dumped to yield 4.8-MHz repetition rate, with output pulses exhibiting 12-ps fwhm. The photon-counting instrument function (obtained by detecting laser pulses scattered by nondairy creamer in water) was typically 80-ps fwhm. Solutions were housed between X/4 fused silica flats separated by a 130-pm Teflon spacer. Their concentrations were maintained at M (for compounds 2-4) and 0.99 (x2= 1.30). All of the photostable compounds show subnanosecond decay, with lifetimes ranging from 153 (for 2 in H 2 0 ) to 319 ps (for 1 in DMSO). These ultrafast decays account for the low emission yieldI3 of mitoxantrone. While dimerization further reduces the empirical quantum yield at elevated concentrations,’ 10 Table I1 shows that the monomer yield is intrinsically low due to ultrafast nonradiative decay. Some of the anthraquinone solutions (notably 1 in CHCl3 and mitoxantrone in DMSO) were photolabile; a photoproduct emission band centered at -660 nm was responsible for the observed nonexponentiality in these cases. The fluorescence lifetimes in Table 11 correlate with the absorption band positions in Table I, as is shown in Figure 5. They are considerably more sensitive to the solvent’s H-bond donating character than to its solvatochromic T* parameter: acetone, methanol, and chloroform all exhibit similar ** (0.71, 0.61, and 0.58 re~pectively~~), but the fluorescence lifetimes of 1-4 are markedly shorter in methanol than in the other two solvents. Water solutions of 2 and 3 (the other two anthraquinones are water-insoluble) exhibited by far the shortest lifetimes, 153 and 169 ps, respectively. Hence, an important nonradiative decay channel in these compounds is clearly modulated by H-bonding interactions with the solvent. This trend is not influenced by structural differences in the 1,4-diamino side chains, nor are they affected by the absence of the phenolic hydroxy group in ametantrone, 2. The site of the pertinent H-bonding interactions is thus localized a t the carbonyl groups. The subnanosecond fluorescence decays of 1,4-diaminoanthraquinone(compound 0 in Table 11) exhibit a similar pattern: this parent comound exhibits a shorter lifetime (574 ps) in methanol than in any of the aprotic solvents. This argues further against direct side-chain involvement in the H-bonding acceleration of the nonradiative decay in anthraquinones 1-4. The fluorescence lifetime of 3 exhibits a large deuterium isotope effect in H-bonding solvents: the decay times in D 2 0 and methanol-d4 are 625 and 933 ps, respectively. Given the lifetimes for 3 in the nondeuterated analogues in Table 11, these correspond to r l ( D ) / r ~ ( H )= 3.69 and 3.56. Much smaller isotope effects occur in the aprotic solvents, since the lifetimes of 3 in acetone-d6
(347 ps, vs 285 ps in acetone) and in DMSO-& (304 ps, vs 368 ps in DMSO) yield T ~ ( D ) / T ~ ( = H )1.21 and 1.13, respectively. In all cases, the charge-transfer absorption spectra of 3 in perdeuterated solvents are nearly identical (band shifts 5 3 nm) to those in the nondeuterated solvents. These results may be compared with the excited-state dynamics of 1,s-dihydroxyanthraquinone (DHAQ), 13-diaminoanthraquinone (1 ,5-DAAQ), and 2,6-diaminoanthraquinone (2,6DAAQ), which were elucidated by Flom and Barbarazs in 1985. Strong intramolecular H-bonding occurs between the phenolic hydroxy groups and carbonyl oxygens in DHAQ, and intermolecular excited-state proton transfer effects a large fluorescence Stokes shift (-6500 cm-I). The DHAQ fluorescence decay is little influenced by the solvent’s H-bonding capacity (-365 ps in n-hexane, -330 ps in ethanol). Intramolecular H-bonding is impossible in 2,6-DAAQ, which exhibits smaller fluorescence Stokes shifts (-3800 cm-I). The emission lifetime in 2,6-DAAQ is considerably shortened in H-bonding solvents, -425 ps in ethanol versus 1670 ps in acetonitrile. For 2,6-DAAQ, formation of an intermolecular 0.-H hydrogen bond between a solvent hydroxy group and the anthraquinone carbonyl creates a new, large-displacement vibrational accepting mode that accelerates SI So internal c o n v e r s i ~ n . *In ~ ~this ~ ~ context, the deuterium isotope effect in the fluorescence lifetime arises from the internal conversion rate’s sensitivity to the accepting mode frequency.26 1,5-DAAQ exhibits intermediate behavior in that it forms weak, intramolecular amino-carbonyl H bonds and slightly accelerated fluorescence decay in H-bonding solvents (-450-ps lifetime in ethanol, -510 ps in 2-methyltetrahydrofuran25). The small fluorescence Stokes shifts in compounds 1-4 (typically 500 cm-I in Table I) show no evidence of strong intramolecular H-bonding or excited-state proton transfer similar to that in DHAQ, despite the presence of the phenolic hydroxy groups in 1,3, and 4. The solvent H-bond donor strength exerts considerably more leverage on the fluorescence kinetics in 1-4 than in DHAQ (negligible) or 1,5-DAAQ (weak). The solvent dependence of the fluorescence dynamics in 1-4 is therefore intermediate between that of 2,6-DAAQ (where essentially all of the rapid internal conversion arises from formation of an intermolecular hydrogen bond) and that in 1,5-DAAQ (where both intra- and intermolecular H-bonding appear to contribute to the subnanosecond nonradiative relaxation. It is unlikely, however, that hydrogen bonding alone accounts for the observation of rapid internal conversion for 1-4 in aprotic as well as hydrogen-bond-donor solvents. Comparisons between the lifetimes of ametantrone (153-240 ps) and 1,4-diaminoanthraquinone (574-793 ps) in Table I1 are instructive here: The only structural difference between these two compounds (2 and 0) is the absence of alkylamino side chains on the 1,Camino groups. The low-frequency modes in the flexible side chains of 2 appear to provide a decay mechanism that is absent in 0, probably through accelerated SI So internal conversion. An antecedent for this is furnished by fluorescence quantum yield studies of the laser dye rhodamine Its fluorescence yield is strongly temperature-dependent (-0.4 at 298 K, -1.0 at sufficiently low temperatures) and has been linked to the temperature-dependent mobility of the chromophore’s diethylamino groups. These groups are immobilized by six-membered carbon rings in the closely related laser dye rhodamine 640, whose fluorescence yield is essentially unity irrespective of temperature. In this context, our measured fluorescence lifetime of 3 in the viscous solvent glycerol (323 ps, essentially single exponential) is interesting. The principal absorption band of 3 in glycerol peaks at -670 nm, the same wavelength as in methanol. According to the lifetimespectral shift correlations in Figure 2, the projected lifetime for 3 in glycerol should then be -260 ps. The discrepancy
-
-
-
B.27928
(26) Avouris, P.; Gelbart, W. M.; El-Sayed, M. A. Chem. Rev. 1977.77, 793.
(27) Huth, B. G.; Farmer, G . I.; Kagan, M. R. J . Appl. Phys. 1969, 40, 5 145.
(28) Drexhage, K. H. In Dye Lasers, Topics in Applied Physics, Springer-Verlag: New York, 1973; Vol. I, p 148.
The Journal of Physical Chemistry, Vol. 95, No. 6, 1991 2255
The Antitumor Agent Mitoxantrone Absorption Band Maxima (nanometers) for MitoxantroneDNA Solutionso
TABLE 111:
DNA, pg/mL mitoxantrone/base pair 75
50
IO 5 I 0
band 1
band 2
626 625 62 1 612 609 608
680 680 665 662 660 660
0.048 0.07I 0.357 0.7 I4 3.51 m
‘Mitoxantrone concentration 5
X
IOd M.
may stem from viscous slewing of the side chains’ mobility in mitoxantrone. (b) In summary, at least two nonradiative decay mechanisms appear to control the subnanosecond fluorescence kinetics in anthraquinones 1-4. The first is specific to H-bonding solvents and exhibits a normal deuterium isotope effect for internal conversion with a hydrogen-bond-accepting mode. The second is acceleration of internal conversion by the floppy side chain substituents in the 1,4-positions. Binding of Mitoxantrone to Double-Stranded DNA. The absorption spectra of 5 X IOd M mitoxantrone in a sequence of buffer solutions (0.2 M NaH2P04, pH 7.013) with increasing Figure 6. Minimum-energy configurations (after ref 8 ) for intercalation concentrations of double-stranded DNA were obtained, with the of 1,4-[ 2- [ 2- [ethylamino]ethyl]amino]-9,IO-anthracenedione between results shown in Table 111. The mitoxantrone charge-transfer C-G base pairs in DNA: (a) “straddling” mode with side chains in major absorption bands in water are Stokes-shifted by -20 nm upon and minor grooves; (b) “perpendicular” mode with both side chains in binding to DNA.’O The spectral changes are consistent with a major groove. Thick lines outline C-G base pairs, thin lines indicate mitoxantrone-DNA binding constant K 2 X lo5 M-I, as defined position of the I ,4-disubstituted anthraquinone. in the Scatchard equation modified by McGhee and von H i ~ p e l : ~ ~ binding mode in Figure 6b, although the former mode requires a higher activation energy through distortion of the DNA backbones). Second, DNA binding can immobilize the side chains (e.g., Here u is the number of adducted drug molecules per DNA base through H-bonding of the distal N atoms to the phosphate pair at equilibrium, L is the concentration of unbound drug backbone in addition to intercalation). Recent binding studies molecules in solution, and n is the number of base pairs (-3 for of similar intercalation agents have shown that the distal N atoms mitoxantrone6) occupied by a single drug molecule. These data are indeed pivotal to antitumor action.30 In particular, a key serve to establish a range of DNA concentrations (>75 pg/mL) determinant of antineoplastic activity in imidazoacridinones with for which essentially all of the visible light absorption is due to side chains analogous to those of mitoxantrone is the number of DNA-bound mitoxantrone. carbon atoms between the proximal and distal N atoms. The most A priori, the fluorescence lifetime of mitoxantrone can be potent activity is exhibited by compounds with two carbon spacers; influenced by its binding to DNA through at least two mechathe presence of additional carbons significantly reduces or destroys nisms. First, the chromophore’s H-bonding sites may be geothe antitumor action. metrically screened from ambient water by the adjacent base pairs. The lifetime of the principal 680-nm fluorescence component In computer simulations for intercalation of the morphologically of 3 in 170 pg/mL DNA is 259 ps (Figure 5 ) . This is far longer similar compound I ,4- [ 2- [2- [ethylamino]ethyl]amino]-9, IOthan the 169-ps lifetime in water and resembles the lifetimes of anthracenedione into DNA, Islam et a18 found a minimum-energy 3 in aprotic solvents. Hence, base-pair shielding of the mitoconformation in which the side chains are located in the major xantrone chromophore from the H-bonding solvent is suggested groove of the double helix. This geometry is shown in Figure 6, by our time-resolved fluorescence profiles as well as absorption along with an alternative, lower energy conformation in which spectra. More intriguing is the fluorescence lifetime for amethe chromophore straddles the base pairs and projects its side tantrone (2)in >300 pg/mL DNA (the larger DNA concentration chains into both major and minor grooves. In either case, here is mandated by the smaller DNA binding constant of amspace-filling models* indicate that the anthraquinone carbonyl etantroneZ9). This is 275 ps, which exceeds not only the I53 ps groups are occluded from the aqueous solvent by the adjacent decay time for 2 in water but also the lifetimes for 2 in any of DNA base pairs. Since H-bonding is highly directional (the donor, the aprotic solvents listed in Table 11. (This fact is highlighted hydrogen, and acceptor atoms are constrained to be nearly colin Figure 5, in which the data point for DNA-bound ametantrone linear), these intercalation geometries offer little opportunity for 2 lies considerably higher than those for 2 in any of the solvents). H-bonding between these chromophore binding sites and the DNA This implies the presence of an additional mechanism for lifetime base pairs. Such a picture is consistent with the fact that the broadening (other than the absence of chromophore H-bonding) absorption spectrum of 3 bound to DNA resembles that of 3 in and suggests that partial immobilization of the side chains through aprotic solvents (e.g., the principal band maxima of 3 in acetone binding of the distal N atoms to the DNA backbone may conand in DNA are at 676 and 680 nm, versus 660 nm for 3 in water, tribute to the observed decay. Controlled time-resolved Table I). The principal driving force for such chromophore influorescence experiments may thus have potential for separating tercalation is thus van der Waals attraction rather than H-bonding effects due to intercalation and distal N atom bonding. Questions between the near-planar anthraquinone and the base pairs and p e r ~ i s tas ~ ’to~the ~ ~relative contributions of these binding modes; is sensitive to the extent of spatial 0verlap2~between the respective *-electron systems. (In this sense, the “straddling” intercalation (30) (a) Cholody, W. M.;Martelli, S.;Paradziej-Lukowicz,J.; Konopa, geometry in Figure 6a is clearly more stable than the major groove J. J. Med. Chem. 1990, 33.49. (b) Atwell, G.J.; Rewcastle, G. W.; Baguley,
-
B. C.; Denny, W. A. J . Med. Chem. 1987,30,664. (29)Wakelin, L. P. G.; Chetcuti, P.;Denny, W. A. J . Med. Chem. 1990, 33,2039.
(31)Zee-Cheng, R. K.-Y.; Cheng, C. C. J . Med. Chem. 1978. 21, 291. (32)Feigon, J.; Denny, W. A.; Leupin, W.;Kearns, D. R.J. Med. Chem.
1984, 27, 450.
2256
J. Phys. Chem. 1991, 95, 2256-2260
they are currently researched by techniques such as effects of structural variations on antineoplastic activity?O nuclear magnetic resonance?' circular di~hroism,'~and computational simulat i o n ~ . * *Future ~ ~ work in our laboratory will examine fluorescence decays of other intercalative agents in poly[d(G-C)-d(G-C)], (33) Dinesen. J.; Jacobscn, J. P.; Hansen, F. P.; Pedersen, E. 9.; Eggert, H. J . Med. Chem. 1990,33, 93. (34) Wilson, D.; Jones, J . L. Intercalation Chemistry; Academic Press: New York, 1982. (35) Rao, S.N.; Remers, W. A. J . Med. Chem. 1990, 33, 1701.
poly[d(A-T)-d(A-T)], and other DNA types.
Acknowledgment. We are indebted to K. C. Murdock and V. Lee of Lederle Laboratories for the gift of anthraquinones 1-4. The Ames Laboratory is operated for the U.S. Department of Energy by Iowa State University under Contract No. W-7405Eng-82. This work was supported by the Office of Basic Energy Sciences. Registry No. 1 acetate, 131685-06-8; 2 acetate, 70711-40-9; 3, 65271-80-9; 4, 70476-63-0.
Optical Phase Conjugation in Nuclear Magnetic Resonance: Laser NMR Spectroscopy M. W.Evans Theory Center, Cornell University, Ithaca, New York 14853 (Received: June 5, 1990)
The role of optical phase conjugation in nuclear magnetic resonance (NMR) spectroscopy is discussed theoretically in terms of the extra quantized angular momentum J, produced by the conjugate product r of a powerful pump laser such as a Nd:YAG. Elementary first-order time-dependent perturbation theory is used to derive an expression for a novel molecular property Y,, tensor that mediates the induction by r of an orbital molecular magnetic moment m,, which couples in to the spin magnetic moment of the nucleus. The tensor Y,j is expressed in terms of the product of matrix elements of a transition molecular orbital magnetic moment multiplied by matrix elements of the dynamical molecular electronic orbital polarizability, a quantity that mediates the effect of r on the interaction Hamiltonian. It is shown that the conventional NMR detail splits according to Clebsch, Gordan, and LandB coupling between J, and I, the nuclear spin quantum number, a potentially useful phenomenon.
(1) Introduction This paper introduces a potentially useful analytical technique based on the effect of powerful pulses of pump laser radiation in conventional nuclear magnetic resonance spectrometers. It is shown theoretically that the conjugate product ( r )of the laser introduces a quantized orbital molecular angular momentum (J,) that couples to the nuclear spin angular momentum (I) through the Clebsch-Gordan theorem governing angular momenta in quantum mechanics. Being a quantized angular momentum, J, is described by two quantum numbers,' J, and M,,,both of which are capable of coupling to I and therefore of splitting the conventional N M R spectrum into detail dependent on new atomic or molecular property tensors. This provides experimental information on the tensors and can be used in the analytical laboratory to supplement conventional NMR spectroscopy by directing the pump laser in the same Z axis as the applied magnetic flux density B of the spectrometer. Part 2 of this paper defines the conjugate product of the pump laser and the interaction Hamiltonian with an atomic or molecular ensemble, thus defining the orbital electronic polarizability a,. Part 3 is a brief demonstration of conservation of Wigner rein the experversality ( T ) and parity inversion symmetry iment consisting of measuring nuclear resonance with r parallel or antiparallel to B of the N M R spectrometer. Part 4 introduces the orbital atomic or molecular magnetic dipole moment m, prqduced by r through a novel molecular property tensor &, which is defined with elementary first-order perturbation theory from the time-dependent Schrcidinger equation. Part 5 is a discussion, still in elementary terms, of the Clebsch-Gordan coupling between the quantized orbital angular momentum defined by J, and the quantized nuclear spin angular momentum I. This coupling de(P)273
( I ) Edmonds, A. R. Angular Momentum in Quanttyn Mechanics; Princeton University Press: Princeton, NJ. 1957. (2) Wigner, E. P. Z. Phys. 1927, 43, 624. (3) Wigner, E. P. Group Theory; Academic: New York, 1959.
0022-3654/91/2095-2256$02.50/0
fines new energy levels, transitions between which are picked up by the resonating probe of the N M R spectrometer in the usual way. The Clebsch, Gordan, and Land6 coupling greatly enriches the customary N M R spectrum, and a discussion closes the paper with estimates of the frequency splitting for a given pump laser electric field strength in volts per meter.
(2) Definition of the Conjugate Product r of the Pump Laser A description of the physical meaning of wave and optical conjugation has been given by Zel'Dovich et a1.4 Its effects are usually investigated experimentally4 by using a pump laser and a device such as a reflective holographic grating or by using four-wave mixing. It is increasingly used for a variety of purposes, including4 optical phase conjugation interferometry (using a phase conjugate mirror), phase conjugation resonance, light guides, pulse temporal spread, self-targeting, spatial/time modulation, bistable optical devices, tunable filters, nonlinear selection, nonlinear phase conjugate spectroscopy, and other effects. Approximately 150 publications per year are produced4 on optical phase conjugation, and the technology is readily available for the development of laser N M R spectrometers. Recently, it has been ~ h o w n ~ - " that J ~ -the ' ~ conjugate product r has the same P and T symmetries'* as static magnetic flux (4) Zel'Dovich, 9. Ya.; Pilipetsky, N. F.; Schumov, V. V. Principles of Phase Conjugation; Springer Verlag: New York, 1985. (5) Barron, L. D. tight Scattering and Optical Activify; Cambridge
University Press: Cambridge, 1982. (6) Barron, L. D. Chem. Soc. Reo. 1986, 15, 189. (7) Barron, L. D. In New Developments in Molecular Chirality; Mezey, P. G., Ed.;Reidel: Netherlands, 1990. (8) Evans, M . W. Phys. Reu. Left. 1990, 64, 2909. (9) Evans, M. W. Phys. Lett. A 1990. 146, 185. (10) Evans, M. W.; Heyes, D. M. Mol. Simul. 1990,4, 399. ( I 1) Evans, M. W.; Heyes, D. M. Phys. Scr. 1990, I ! ,304. (12) Barron, L. D. Nature 1975, 257, 372. (13) Evans, M. W. Opt. Lett. 1990, 15, 863. (14) Evans, M. W. J . Opf. Soc. Am., B, in press. ( 1 5 ) Evans, M. W. J . Chem. Phys. 1990, 93, 2329.
0 1991 American Chemical Society
