Cis−Trans Isomerizations of β-Carotene and Lycopene: A Theoretical

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Cis-Trans Isomerizations of β-Carotene and Lycopene: A Theoretical Study Wen-Hsin Guo, Cheng-Yi Tu, and Ching-Han Hu* Department of Chemistry, National Changhua UniVersity of Education, Changhua 50058, Taiwan ReceiVed: March 5, 2008; ReVised Manuscript ReceiVed: June 24, 2008

The all-trans to mono-cis isomerizations of polyenes and two C40H56 carotenes, β-carotene and lycopene, at the ground singlet (S0) and triplet (T1) states are studied by means of quantum chemistry computations. At the S0 state of polyenes containing n acetylene units (Pn), we find that the energy barrier of the central CdC rotation decreases with n. In contrast, however, at the T1 state, the rotational barrier increases with n. For the C40H56 carotenes, the rotational barriers of lycopene are lower than those of their β-carotene counterparts. This difference renders the rotational rates of lycopene to be 1-2 orders of magnitude higher than those of β-carotene at room temperature. For both these carotenes, the barrier is lowest for the rotation toward the 13-cis isomer. The relative abundances are in the following order: all-trans > 9-cis > 13-cis > 15-cis. Although the 5-cis isomer of lycopene has the lowest energy among the cis isomers, its formation from the all-trans form is restricted, owing to a very large rotational barrier. The possible physiological implications of this study are discussed. 1. Introduction Carotenes play an important role in biological systems. They act as antennas in the light-harvesting units in chlorophyll and bacteriochlorophyll, and they also protect biological tissues from the deleterious effects of singlet oxygen and triplet bacteriochlorophyll.1,2 Dietary carotenes are known to exhibit antioxidation properties.3,4 For example, Di Mascio et al. have shown that lycopene (Scheme 1) is most effective among carotenes in quenching singlet oxygen.5 Some carotenes, such as β-carotene (Scheme 1), are converted to retinol (vitamin A); therefore, they thus are vital for vision, skin and membrane protection, and anticancer activity.6 The protective effects of carotenoids have been well-documented.3,4,7-11 It has been recognized by many studies that these effects are dependent upon their stereochemistry. For example, the provitamin A activity of carotene is correlated with its stereogeometry. In living organisms, the conversion of β-carotene to retinol is performed via regiospecific, central cleavage at its (E)-configured 15,15′-position.12 The isomerization of β-carotene would probably decrease the provitamin A activity. Although the all-trans isomers of carotenes are apparently the most stable, cis isomers were found to be present in minor amounts. In human serum and tissues, it has been discovered that more than 50% of lycopene and ∼5% of β-carotene exist in cis conformations.13 The cis isomers of lycopene were found to be more bioavailable than the all-trans isomer.14 Food processing, heating, and illumination would definitely facilitate the cis-trans isomerizations of carotenoids and thus would alter the bioactivity of these compounds.15,16 In an extract of the thylakoid membranes of a thermophilic blue-green alga and of spinach, Ashikawa et al. found approximately 80% all-trans isomer and 20% cis isomers including 9-cis, 9,13-di-cis, 9,9′-di-cis, 13-cis, and 15-cis isomers (in decreasing order of quantity).17 Different types of carotenes were observed in the cytochrome b6f (Cyt b6f) complexes obtained from various species. For one example, 9-cis-β-carotene was found to be the predominant component in Cyt b6f complex of * Corresponding author.

spinach.18 Further, 9-cis-R-carotene is bound to the Cyt b6f complex of green alga Bryopsis corticulans.19 Interestingly, it was observed that in the reaction center of light-harvesting system of photosynthetic bacteria, the configuration of the carotenoid is 15-cis; in contrast, carotenoid in the antenna complexes exhibit all-trans configuration.1 An important question has been raised by Ashikawa et al. with regard to whether or why different isomers in the photosystem have optimal functions, i.e., photoprotection in the reaction center and lightharvesting in the antenna complex.1 Because of the relevance to their functions, the isomerization of carotenoids has attracted considerable attention from chemists. Spectroscopic identifications of various cis and trans isomers of β-carotene have been reported by Koyama’s group; in their study, isomerizations at the S0 and T1 states were investigated under thermal and triplet-sensitized conditions.20 Doering and co-workers reported cis-trans thermal isomerizations among β-carotene and its cis isomers.21 At and above physiological temperature they found that 13-cis and 15-cis isomers are relatively abundant, and this leads to the inference that the 13cis and 15-cis isomers of β-carotene are true anticarcinogenic agents, whereas all-trans-β-carotene is relegated to the reservoir of the cis isomers.21 Doering’s research brought to attention 9-cis, 13-cis, and 15-cis isomers in addition to all-trans-βcarotene. After photoexcitation to its optically allowed singlet state, carotene decays to the lower singlet (S1) and triplet (T1) states within 1 ps. The lifetimes of S1 and T1 states are in the picosecond and nanosecond ranges in light-harvesting systems, respectively.20 In solution, the triplet lifetime of a carotene is even longer, being at the microsecond range.22 In the presence of light, therefore, it is thus essential to consider the isomerization process that occurs via the T1 state. In the present investigation, the cis and trans isomers of β-carotene and lycopene, and the transitions between them at the S0 and T1 states, are studied using theoretical methods. All-trans polyenes have been used as model compounds for understanding the cis-trans isomerizations of carotenes, through both experimental and theoretical means.21,23-25 Doering and

10.1021/jp8019705 CCC: $40.75  2008 American Chemical Society Published on Web 08/28/2008

Cis-Trans Isomerization of β-Carotene and Lycopene

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SCHEME 1: Structure and Numbering Schemes for all-trans-β-Carotene and all-trans-Lycopene

coauthors measured the enthalpies of activation of semirigid polyenes; they incorporated all the double bonds, but the central one, into fused six-membered rings to preclude the formation of conformers other than central-cis isomers. Polyenes with up to nine acetylene units (P9) have been investigated, and the enthalpies of activation were reported by Doering’s group.24 In this study, we include the central-cis isomerizations of all-trans polyenes up to P15. 2. Computational Approach The computations were performed using the density functional theory (DFT) and CASMP2 approaches implemented in the Gaussian03 series of programs.26 In the CASMP227 approach, electron correlation corrections are added to the CASSCF theory,28 and it is used in our model polyene compounds. The B3LYP density functional was applied to all of the compounds in this study; it is a hybrid method that includes Becke’s threeparameter mixing of the nonlocal exchange potential29 and the nonlocal correlation functional proposed by Lee, Yang, and Parr.30 In the study by Bernardi et al., it has been demonstrated the B3LYP/6-31G(d) theory is able to obtain rotational barriers of small polyenes (up to all-trans-octatetraene) that are within 1 kcal/mol from the CASPT2 predictions.31,32 Harmonic vibrational frequencies were computed via analytic energy second derivatives at the B3LYP/6-31G(d) level. These frequencies were used to verify genuine minimum, transition states, and were used to compute zero-point vibrational energy corrections. These frequencies were also used to compute enthalpy and free energy corrections. For small polyenes (P1-P4), we applied the CASMP2/6-311G(d)//CASSCF/6-31G(d) theory. The B3LYP results of these polyenes were then compared with those of CASMP2 and CASPT2. The singlet transition states of polyenes or carotenes have a strong diradical, open-shell singlet character and hence should be defined at least by a two-determinant wave function. Within unrestricted DFT, this issue is resolved by spin correction, which was suggested by Yamaguchi et al.33 The corrections turn out to be very small (within 1 kcal/mol) in all cases. 3. Results and Discussion 3.1. Polyenes. In the case of small polyenes, the energy barriers computed using B3LYP/6-31G(d) for rotating the central double bonds are compared with those computed using highlevel ab initio theories (see Table 1). It should be noted that the rotational barriers predicted for P1-P4 using DFT are in reasonably good agreement with those predicted using the

TABLE 1: Barriers of Activation Energy (∆Eq), Barriers of Activation Enthalpy (∆Hq), Vertical and Adiabatic Singlet-Triplet Energy Separations (∆E′S-T and ∆ES-T) of All-Trans Polyenes (in kcal/mol) Predicted by DFT and ab Initio Theoriesa ∆Eq

∆Hq ∆E′S-T ∆ES-T

P1 B3LYP/6-31G(d) 57.4 (58.4)b 57.7 104.1 CASMP2/6-311G(d)//CAS(2,2)/ 64.4 65.4 110.3 6-31G(d) CASPT2/6-31G(d)//B3LYP/ 58.3 59.6 104.5 6-31G(d) P2 B3LYP/6-31G(d) 48.7 (49.3)b 48.8 CASMP2/6-311G(d)//CAS(4,4)/ 55.7 55.8 6-31G(d) CASPT2/6-31G(d)//B3LYP/ 50.3 50.0 6-31G(d) P3 B3LYP/6-31G(d) 40.5 (41.0)b 40.4 CASMP2/6-311G(d)//CAS(6,6)/ 45.8 45.6 6-31G(d) CASPT2/6-31G(d)//B3LYP/ 40.7 40.4 6-31G(d) P4 B3LYP/6-31G(d) 36.4 (36.7)b 36.2 CASMP2/6-311G(d)//CAS(8,8)/ 40.6 40.8 6-31G(d) CASPT2/6-31G(d)//B3LYP/ 35.8 6-31G(d)

75.4 81.4

60.4 64.1

54.5 59.9

74.5

58.6 61.9

41.3 44.3

58.6

48.6 53.6

32.9 35.3

a CASPT2/6-31G(d)//B3LYP/6-31G(d) predictions are adopted from previous theoretical studies (refs 32 and 34). b B3LYP/6-31G(d) predictions adopted from Bernardi et al. (ref 32).

CASMP2 and CASPT2 theories. It is observed that ∆Eq (difference of electronic energy plus zero-point vibrational energy) and ∆Hq (difference of enthalpy at 298 K) predicted using B3LYP are very close to those predicted using the CASPT2 theory. In contrast, the CASMP2 predictions overestimate the barriers by several kilocalories per mole. In all the cases, the differences between the values obtained using B3LYP and CASPT2 are less than 1 kcal/mol. Our B3LYP/6-31G(d) predicted ∆Eq and ∆Hq to be slightly smaller than those predicted by Bernardi et al.;32 this difference is attributed to the inclusion of spin correction in the singlet transition states. In addition, vertical (singlet and triplet energies computed at the S0 geometry) and adiabatic (energies computed at the S0 and T1 optimized geometries, respectively) singlet-triplet energy separations (∆E′S-T and ∆ES-T) predicted using B3LYP

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TABLE 2: Barriers of Activation Energy (∆Eq), Enthalpy (∆Hq), and Gibbs Free Energy (∆Gq), the Reaction Energy (∆E), Enthalpy (∆H), and Gibbs Free Energy (∆G), and the Vertical and Adiabatic Singlet-Triplet Energy Separations (∆E′S-T and ∆ES-T) of All-Trans Polyenes (in kcal/mol) Predicted using B3LYP/6-31G(d)a ∆Eq

∆Hq

∆Gq

57.7 (58.1)b 48.8 40.4 (38.9)b 36.2 32.2 (32.1)b 29.9 27.6 (27.5)b 26.0 24.5 (24.5)b 23.4 22.2 (22.4)b 21.4 20.5 (21.1)b 19.8 19.1 (20.1)b

S0 58.0 (65)c 48.6 41.0 (42.2)d 36.5 32.5 30.1 27.9 26.3 24.8 23.7 22.6 21.7 20.9 20.2 19.6

P1 P2 P3 P4 P5 P6 P7 P8 P9 P10 P11 P12 P13 P14 P15

57.4 48.7 40.5 36.4 32.4 30.0 27.8 26.2 24.7 23.6 22.4 21.6 20.7 20.0 19.3

P1 P2 P3 P4 P5 P6 P7 P8 P9 P10 P11 P12 P13 P14 P15

17.8 17.6 0.0 -0.5 1.0 0.6 4.4 4.0 5.9 5.5 7.6 7.2 8.4 8.0 9.2 8.9 9.7 9.4 10.2 9.9 10.5 10.2 10.8 10.5 11.0 10.7 11.2 10.9 11.3 11.1

∆E ∆H ∆G ∆E′S-T ∆ES-T 0.0 0.0 2.0 2.1 2.1 2.2 2.1 2.2 2.1 2.2 2.1 2.2 2.1 2.2 2.2

0.0 0.0 2.0 2.0 2.0 2.1 2.1 2.2 2.2 2.2 2.2 2.2 1.6 1.6 2.2

0.0 0.0 1.9 2.0 1.8 2.1 1.7 2.1 1.4 1.9 0.8 1.4 2.9 3.1 0.8

0.0 0.0 0.3 2.5 2.9 2.9 3.0 3.0 3.0 3.0 3.0 3.0 3.0 3.0 3.0

0.0 0.0 0.3 2.1 2.9 3.0 3.1 3.1 3.1 3.1 3.2 2.5 3.2 2.6 3.2

0.0 0.0 0.1 2.9 3.0 2.3 1.7 2.5 2.1 2.3 2.0 3.7 1.7 3.7 1.6

104.1 75.4 58.6 48.6 42.0 37.3 33.9 31.2 29.1 27.5 26.1 25.0 24.1 23.3 22.7

60.4 54.5 41.3 33.0 27.2 23.1 19.9 17.4 15.4 13.7 12.3 11.0 10.0 9.0 8.2

T1 17.5 0.6 1.9 4.8 6.3 7.9 8.7 9.5 10.0 10.4 10.7 11.0 11.2 11.4 11.5

a Available experimental results and predictions from previous predictions are included in parentheses. b Experimental measurement for the semirigid counterparts of Pn from Doering and Sarma (ref 24). c Experimental rotational barriers (∆Gq) (ref 44). d Experimental rotational barriers ∆Gq) (ref 45).

are in good agreement with those obtained from the CASPT2 theory but are smaller than those obtained using the CASMP2 theory. The comparison between B3LYP and ab initio theories encourages the application of B3LYP to the computation of the potential energy surfaces for the isomerizations at the ground state (S0) and first triplet state (T1) of polyenes. All-trans polyenes (H-(CHdCH)n-H, Pn) up to P15 are examined using the B3LYP/6-31G(d) method, and the results are summarized in Table 2. The geometries of singlet and triplet Pn are all planar, the only exception being triplet ethene, which adopts a D2d geometry. For simplicity, we illustrated only the geometrical parameters of P9, P11, P13, and their central-cis isomers in Figure 1. The geometrical parameters of all the polyenes are illustrated in the Supporting Information. Among the singlet polyenes, we observed the bond-length alternations and found that the extent of alternation becomes less apparent for larger polyenes. In contrast, the bond lengths of triplet all-trans Pn reveal switching between the double and single bonds, particularly at the central region. The longest bond in the T1 state is the central double bond.

The transition states involved in the all-trans to cis rotations at both S0 and T1 surfaces of P9, P11, and P13 are illustrated in Figure 2 (results of all polyenes are illustrated in the Supporting Information). As can be observed in Figure 2, the geometry of the transition states at the S0 and T1 surfaces are very similar; the most noticeable difference is observed in the case of the torsional angle at the central double bond. The geometries of the transition states reveal that they exhibit strong diradical nature; therefore, the electronic nature of the transition states at the S0 and T1 states mainly differ by the spin couplings of the diradical electrons. With regards to the energy, we found that for polyenes with more than four acetylene units, the transition states of T1 are higher in energy by no more than 1 kcal/mol than those of S0. The predicted singlet-triplet energy separations in Table 2 reveal the electronic nature of the polyenes. As a result of the extended π-conjugation system, the relative energy of the T1 state decreases when n increases. We observed that both ∆E′S-T (singlet and triplet energies computed at the S0 geometry) and ∆ES-T (singlet and triplet energies computed at the S0 and T1 geometries) decrease with an increase in n. On the basis of the studies on polyenes using the Hubbard and Pariser-Parr-Pople models, Tavan and Schulten demonstrated fairly linear relations between the singlet excitation energies and 1/(2n + 1).23 Interestingly, similar correlations were observed for our predicted values of ∆E′S-T and ∆ES-T (Figure 3). For the S0 state, the rotational barriers are smaller in extended polyenes. The above-mentioned trend of ∆Eq versus n in Pn is attributed to the stabilization of the open-shell singlet diradical by the polyacetylene units: the larger the unit, the smaller the barrier. The imaginary vibrational frequencies also decrease with n (see the Supporting Information). The rotational barriers of Pn predicted in this work are in good agreement with the experimental results of Doering et al., which were derived from the kinetic data of semirigid polyenes.24 The rotational barriers for the S0 and T1 states of polyenes are plotted against n in Figure 4a. At the S0 state, ∆Eq decreases with an increase in n. This trend is attributed to the stabilization of the transition state in larger polyenes. The geometries of the transition states of polyenes contain CdC double bonds rotated to ∼90° (refer to the Supporting Information); therefore, the more extended π-conjugation systems are capable of stabilizing the transition states. In contrast to ∆Eq of polyenes in the S0 state, that of polyenes at the T1 state increases with n. This seemingly contradictory result is a consequence of the fact that, at larger n, the extended π-conjugation system stabilizes the T1 minima to a larger extent than it stabilizes the transition states. This is demonstrated by the dependency of ∆ES-T on n, as shown in Figure 4a. Since the energies of the transition states for S0 and T1 are nearly degenerate because they are a singlet and triplet pair of openshell diradicals, respectively, the rotational barrier in the T1 state is proportional to n. This correlation explains how the potential energy surfaces of the Pn change with the length of polyenes. With respect to the S0 minima, as n of Pn increases, the relative energies of T1 decrease more drastically than those of the transition states (refer to Figure s4 of the Supporting Information). Linear correlations are observed when we plot ∆Eq of S0 and T1 against ∆ES-T. As illustrated in Figure 4b, the rotational barriers of singlet Pn are directly proportional to ∆ES-T, whereas those of the triplet Pn are inversely proportional to ∆ES-T. An exception to the aforementioned dependency is triplet P1, for which the rotational barrier is the largest among the triplet polyenes studied. Unlike the other triplet Pn compounds, the

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Figure 1. Geometrical parameters (in angstroms) of singlet (S0) and triplet (T1) polyenes (P9, P11, P13), and their corresponding central-cis isomers optimized at B3LYP/6-31G(d) level. Geometrical parameters of S0 and T1 states are shown in the upper and lower entries, respectively.

minimum of P1 adopts a nonplanar, D2d symmetry. Its transition state, however, has a planar geometry. The electronic structure of P1 is, therefore, not comparable to that of the other polyenes. The cis isomers at the S0 and T1 states are higher in energy than their all-trans counterparts. 3.2. β-Carotene and Lycopene. The potential energy surfaces of the isomerizations of all-trans-β-carotene and lycopene were investigated using the B3LYP approach. For simplicity, the structures of all the isomers and transition states are summarized in the Supporting Information. We include the optimized structures of the two carotenes and their 13-cis and

15-cis isomers in Figure 5. The structures and magnitudes of the imaginary frequencies for the transition states corresponding to the all-trans to 13-cis and 15-cis isomerizations are shown in Figure 6. The optimized structures of β-carotene and lycopene reveal the following trends similar to those observed in polyenes; bondlength alternation at the S0 state and the bond-length inversion at the T1 state. Bond-length alternation is apparent at the central part of the compound and is less apparent at the peripheral region of the conjugated chains. In contrast to the marked difference between the geometries of S0 and T1, their transition

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Figure 2. Geometrical parameters (in angstroms) and magnitude of imaginary vibrational frequencies (νimag, in i cm-1) of the isomerization transition states (TSs) of singlet (S0) and triplet (T1) polyenes (P9, P11, P13) located at B3LYP/6-31G(d) level. The results of S0 and T1 states are shown in the upper and lower entries, respectively.

Figure 3. Singlet-triplet energy separation (∆ES-T, b, R2 ) 0.9940), and vertical singlet-triplet energy separation (∆E′S-T, 9, R2 ) 0.9993,), vs 1/(2n + 1) for polyenes P2-P15. The energies are in kcal/mol.

states (Figure 6) are very similar. For each isomerization, the transition states at S0 and T1 surfaces are nearly degenerate (vide infra). The imaginary vibrational frequencies of the S0 state are higher than their counterparts at the T1 state. The rotational barriers and energies of isomerization reactions involved in the isomerizations of all-trans-β-carotene and lycopene are summarized in Tables 3 and 4. Relative energies (including zero-point vibrational energy corrections) of all-transβ-carotene and lycopene, their mono-cis isomers, and the transition states of isomerizations at the S0 and T1 states are schematically illustrated in Figures 7 and 8. For both β-carotene and lycopene, at their S0 and T1 states, the lowest rotational barrier occurs at the 13-14 CdC bond, rather than the 15-15′ CdC bond. This observation is contradictory, for the experiments performed by Doering’s group

Figure 4. (a) Rotational barrier (∆Eq) of S0 ([) and T1 (2) and the singlet-triplet energy separation (∆ES-T, 9) vs n; (b) ∆Eq of S0 (0, R2 ) 0.9971) and T1 (O, R2 ) 0.9793) states vs ∆ES-T, for P2-P15. The energies are in kcal/mol.

revealed that the rotational rates for the conversion of the alltrans isomer to the 13-cis isomer of β-carotene are higher than those for the conversion of its all-trans isomer to the 15-cis isomer.21 We attribute this result to the stabilization of the transition state by methyl substitution at the 13th carbon. The stabilization effect of methyl group on the rotational barrier of

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Figure 5. Geometrical parameters (in angstroms) of all-trans, 13-cis, and 15-cis isomers of singlet (S0) and triplet (T1) carotenes optimized at B3LYP/6-31G(d) level. Geometrical parameters of S0 and T1 states are shown in the upper and lower entries, respectively.

an alkene has been studied by Jarowski et al.34 To verify this proposal, we examined the rotational barriers of trans-2-butene and 2-methyl-trans-butene at the B3LYP/6-31G(d) level. Indeed, the energy of the rotational barrier of 2-methyl-trans-butene is lower than that of trans-2-butene by 1.3 kcal/mol.

At S0, the rotational barrier for the conversion of the alltrans to mono-cis isomers for both β-carotene and lycopene are in the following order: 13-cis < 15-cis < 11-cis < 9-cis < 7-cis (< 5-cis, lycopene). Nevertheless, we see that for all isomers except for 7-cis rotations, the energies of the rotational barriers

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Figure 6. Geometrical parameters (in angstroms) and magnitude of imaginary vibrational frequencies (νimag, in i cm-1) of the isomerization transition states (TSs) of singlet (S0) and triplet (T1) carotenes located at B3LYP/6-31G(d) level. The results of S0 and T1 states are shown in the upper and lower entries, respectively.

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TABLE 3: Barriers of Activation Energy (∆Eq), Enthalpy (∆Hq), and Gibbs Free Energy (∆Gq), the Reaction Energy (∆E), Enthalpy (∆H), and Gibbs Free Energy (∆G) of all-trans-β-Carotene into its Cis Isomers, and the Adiabatic Singlet-Triplet Energy Separations (∆ES-T) of Isomers (in kcal/mol) Predicted using B3LYP/6-31G(d) ∆Eq S0 13-cis 15-cis 11-cis 9-cis 7-cis T1 13-cis 15-cis 11-cis 9-cis 7-cis

∆Hq ∆Gq

22.6 (27.7)c 23.2 (28.9)c 24.2 26.9 27.4

22.5 23.2 24.2 26.9 27.3

9.1 9.7 10.9 13.4 14.0

9.5 8.6 2.6 10.0 9.0 3.2 11.1 11.2 5.3 13.7 12.8 1.3 14.2 13.5 4.6

22.7 23.1 23.6 26.8 27.3

∆H ∆G ∆Erq a ∆ES-T

∆E 1.1 (1.3)c 2.6 (1.7)c 5.3 0.9 5.2

1.0 2.6 5.1 0.8 5.1

1.6 2.5 5.9 1.2 5.6

21.5 20.6 18.9 26.0 22.2

3.3 4.0 5.9 1.9 5.3

1.6 1.7 4.2 0.3 3.0

6.5 6.5 5.6 12.1 9.4

13.8b 15.3 14.4 13.8 14.2 13.3

a Rotational barrier for the mono-cis to all-trans reverse reaction. ∆ES-T of all-trans-β-carotene. c Arrhenius parameters obtained by Doering et al. (ref 21).

b

TABLE 4: Barriers of Activation Energy (∆Eq), Enthalpy (∆Hq), and Gibbs Free Energy (∆Gq), the Reaction Energy (∆E), Enthalpy (∆H), and Gibbs Free Energy (∆G) of all-trans-Lycopene into its Cis Isomers, and the Adiabatic Singlet-Triplet Energy Separations (∆ES-T) of Isomers (in kcal/mol) Predicted using B3LYP/6-31G(d) ∆Eq S0 13-cis 15-cis 11-cis 9-cis 7-cis 5-cis T1 13-cis 15-cis 11-cis 9-cis 7-cis 5-cis b

∆Hq

∆Gq

∆E

∆H

∆G

∆Erq a

∆ES-T b

21.1 21.9 22.2 24.1 27.6 35.4

20.9 21.6 21.8 23.9 27.4 35.1

20.5 21.1 22.2 23.9 27.7 35.5

1.2 2.8 5.4 1.0 5.5 0.2

1.1 2.7 5.2 0.9 5.2 0.0

1.4 2.4 5.8 0.9 6.1 1.1

19.9 19.1 16.8 23.1 22.1 35.2

9.3 10.0 10.4 12.4 15.9 23.7

9.1 9.7 10.0 12.1 15.5 23.4

9.2 9.7 10.9 12.7 16.3 24.5

2.6 3.4 5.2 1.6 5.2 0.0

2.8 3.5 5.1 1.5 5.0 -0.1

1.6 2.4 5.3 1.8 5.4 0.4

6.7 6.6 5.2 10.8 10.7 23.7

11.9 13.4 12.6 11.8 12.5 11.6 11.7

a Rotational barrier for the mono-cis to all-trans reverse reaction. ∆ES-T of all-trans-lycopene.

of lycopene are higher than those of their counterparts in β-carotene. Therefore, at room temperature, the isomerization rates of lycopene are higher by 1-2 orders of magnitude than those of β-carotene. Despite the fact that both these carotenes possess 11 conjugated π-bonds, the presence of the β-ionone ring in β-carotene reduces electron delocalization. ∆ES-T of all-transβ-carotene is 13.8 kcal/mol; in contrast, ∆ES-T of all-translycopene is lower by ∼2 kcal/mol. Similarly, ∆ES-T values of the lycopene isomers are lower than those of their β-carotene counterparts (see Tables 3 and 4). The low-lying triplet of lycopene could be responsible for its significant antioxidative effects among carotenes. On the basis of experimental measurements, Doering and Sarma extrapolated their results and predicted a rotational barrier (∆Hq) of 22.4 kcal/mol for semirigid P11.24 Our DFT computation for the rotational barrier of P11 (22.2 kcal/mol, see Table 2) is in close agreement with Doering and Sarma’s prediction. Considering β-carotene and lycopene that involve a branched P11 conjugated system, the predicted energies of the 15-cis rotational barriers in

Figure 7. Potential energy diagrams of all-trans-β-carotene, their mono-cis isomers, and the transition states of isomerization at the S0 (a) and T1 (b) states. The energies (in kcal/mol) are presented with reference to the all-trans isomer of S0.

β-carotene and lycopene are 23.2 and 21.6 kcal/mol, respectively. The energy of the central (15-cis) rotational barrier of β-carotene is higher than that of P11, whereas the energy of the rotational barrier of lycopene is lower than that of P11. Despite the fact that our computed order of reactivities and relative stabilities for alltrans, 13-cis, and 15-cis isomers of β-carotene are in the same order with those predicted by Doering et al.,21 our values are ∼5 kcal/ mol higher in energy than those predicted by Doering (Table 3). At present, we cannot offer an explanation to justify this noticeable deviation. For both the carotenes, the all-trans isomers are the most stable at the S0 and T1 states. The order of the relative energies of the mono-cis isomers is also similar for both the electronic states as follows: (5-cis, lycopene) < 9-cis < 13-cis < 15-cis < 7-, 11-cis. Thus, thermodynamically, it is easier to rotate a methylated double bond (5-cis, 9-cis, and 13-cis) than an unmethylated double bond, i.e., 15-cis, 7-cis, and 11-cis. Among the methylated cis isomers, it is easier to rotate the peripheral ones (5-cis and 9-cis). In contrast, in the case of unmethylated double bonds, the central 15-cis isomer is more susceptible to rotation. It has to be noted that, although 5-cis-lycopene is a thermodynamically stable product, it has an extraordinarily high rotational barrier. The relative stabilities of the isomers, as shown in Figures 7 and 8 and in Tables 3 and 4, are generally in good agreement with the experimental values. In chloroplasts and solvents, the 9-cis and 13-cis isomers of β-carotene and lycopene were found to be the predominant cis isomers.15 In human serum and tissues, the 9-cis, 13-cis, and 15-cis isomers of β-carotene and lycopene are present.13 From the experiments on both pure β-carotene and carrot juices, 13-cis isomer was found to be the dominant cis isomer, whereas under lighted storage, the accumulation of 9-cis isomer was favored.35,36 In green vegetables and in fruits, 9-cis-β-carotene was found to be the major cis isomer.37 In Figure 7, we can observe

12166 J. Phys. Chem. B, Vol. 112, No. 38, 2008 that the relative energies of the 9-cis and 13-cis isomers are comparable, whereas the formation of 9-cis involves a larger rotational barrier. Aman et al. revealed that in heated chloroplast isolates, pure 13-cis-β-carotene and 13-cis-lutein are the predominant isomers, whereas 9-cis is the predominant cis isomer in heated chloroplast.15 This observation is justified as the formation of 13cis-β-carotene is kinetically favored, whereas the formation of 9-cisβ-carotene is thermodynamically favored. The 11-cis isomer of β-carotene has received less attention as its spectrum is very similar to that of the all-trans isomer, making its identification more difficult than other isomers.38 In addition, it has been pointed out by Hu et al. that the 11-cis isomer would experience significant steric repulsions, thereby rendering it be unstable to coexist with other isomers at room temperature.38 Our results are in accordance with these predictions, i.e., the 11-cis isomer is the least stable species on both the S0 and T1 potential energy surfaces. Kinetically, 11-cis-β-carotene has the smallest barrier toward all-trans-β-carotene and thus can most easily elude experimental identification. In addition, 7-cis is also an unstable species. At the ground state, 7-cis-β-carotene is only 0.1 kcal/mol lower in energy than 11-cis-β-carotene. The rotational barrier of 7-cis-β-carotene, however, is larger than that of 11-cis-β-carotene. 7-cis-β-Carotene was observed by Tsukida and Saiki, whereas 11cis-β-carotene was not.39 The thermal isomerization reaction of mono-cis isomers with all-trans-β-carotene reveals that the experimental isomerization rate is in the following order: 7-cis < 9-cis < 13-cis < 15-cis < 11-cis.20,38 The rate of isomerization according to our prediction is in the order of 9-cis < 7-cis < 13-cis < 15-cis < 11-cis. The energy barrier of thermal isomerization of 9-cis-βcarotene to all-trans-β-carotene was 26.0 kcal/mol, which is significantly higher than those of other cis isomers. The di-cis isomers of 9-cis-β-carotene were identified by Kuki et al.20 Overall, our results are in reasonable agreement with the experimental results, in which the composition of isomers in natural or heated β-carotene was investigated. The predicted rotational barriers at S0 suggest that, within hours, the all-trans isomer is in equilibrium with the first three or four lowest energy isomers. Experimentally, when the thermal isomerization reaction is carried out for 30 min with the alltrans isomer as the starting material, the amounts of 7-cis and 11-cis isomers are negligible.20 Although 9-cis-β-carotene has the next higher population after all-trans-β-carotene, the very large rotational barrier forbids its early observation. The relative populations of the isomers obtained from our thermodynamic data are consistent with the experimental observations, wherein 13-cis, 9-cis, and 15-cis isomers are the products formed by the isomerization of all-trans-β-carotene at 80 °C.20 As revealed by Koyama’s group, the 13-cis and 15-cis isomers of β-carotene are efficiently isomerized toward the all-trans configuration upon triplet excitation.1,20 Our predictions (Table 3) show that at the T1 state, the 11-cis, 13-cis, and 15-cis isomers are most susceptible to isomerization. The theoretical predictions are in good agreement with the observations (11-cis was not included in the triplet sensitization experiment). However, our prediction for the isomerization efficiency of the triplet 9-cis isomer remains questionable when compared with the experimental values. The triplet-sensitized photoisomerization of the all-trans and mono-cis isomers of β-carotene shows that the decrease of the isomers at the initial stage of irradiation is in the following order: all-trans < 7-cis < 9-cis < 13-cis < 15-cis.40 According to the computed rotational barriers, the tendency for isomerization is 9-cis < all-trans < 7-cis < 13-cis < 15-cis < 11-cis at T1. Again, we found that the predicted activities agree well with

Guo et al.

Figure 8. Potential energy diagrams of all-trans-lycopene, their monocis isomers, and the transition states of isomerization at the S0 (a) and T1 (b) states. The energies (in kcal/mol) are presented with reference to the all-trans isomer of S0.

the experimental results, with the exception for the 9-cis isomer. According to our prediction, the 9-cis isomer is most stable with respect to isomerization. In addition, the relative compositions of the isomers after 1000 or 3000 pulses of irradiation for tripletsensitized photoisomerization are in the order all-trans . 9-cis > 13-cis > 15-cis.40 We can summarize the rule of thumb for evaluating the relative stability of carotene isomers on the basis of our computational data: (1) the all-trans isomers are the lowest energy isomers; (2) the methylated cis isomers are lower in energy than the unmethylated cis isomers, and among them the peripheral cis isomers are lower in energy; (3) among the unmethylated cis isomers, central-cis (15-cis) isomers are lower in energy. These general features are valid for both S0 and T1 potential energy surfaces. Although the information for the isomerizations of pure carotenes may not be applied to chloroplast-bound carotenes, the isomerization processes of the two important carotene species examined in this study should provide valuable insight for future studies. The relative stability of the 9-cis isomers of β-carotene and lycopene may have important physiological implications. As mentioned in the Introduction, cis isomers including 9-cis, 9,13di-cis, and 9,9′-di-cis isomers are the predominant cis isomers in blue-green algae and spinach.17 9-cis-β-Carotene was found to be the predominant component in Cyt b6f of spinach,18 whereas 9-cis-R-carotene is bound to the Cyt b6f complex of green algae.19 Zhang et al. have reported the existence of stoichiometrically bound (9-cis or 15-cis) β-carotene in the Cyt b6f extracts obtained from a thermophilic bacterium, spinach, and a green alga.41 It was suggested that the function of the protein-bound β-carotene is to provide protection of the Cyt b6f complex against the toxicity of photosynthetically produced

Cis-Trans Isomerization of β-Carotene and Lycopene O2.41 In vitro experiments of Levin and Mokady suggest that 9-cis-β-carotene has a higher antioxidant potency than that of the all-trans isomer.42 Liu and Osawa showed that the cis isomers, especially 9-cis-astaxanthin, exhibit higher antioxidant activities in vitro than the all-trans isomer.43 Being more stable and more thermodynamically favored than other cis isomers, the presence of these 9-cis isomers, particularly in Cyt b6f complexes, may play important roles in antioxidant functions. 4. Conclusion We have studied the mono-cis rotations of all-trans polyenes, β-carotene, and lycopene. The B3LYP method has been demonstrated to be able to provide reliable predictions for the rotational barriers of the species in this study. In the case of polyenes, we found that ∆Eq for Pn isomerization at S0 decreases with an increase in n; in contrast, ∆Eq increases with n at the T1 state. The difference is attributed to the fact that, with respect to S0, the relative energy of T1 decreases at a faster rate than the energy of the transition state with an increase in n. A linear correlation was observed between ∆ES-T and 1/(2n + 1) for Pn, which was in agreement with the prediction of Tavan and Schulten. In addition, we found that ∆Eq of Pn is linearly proportional to ∆ES-T at S0 and inversely proportional to ∆ES-T at T1. Therefore, the singlet-triplet energy separation is a very good indicator for the stability of a polyene cis-trans isomerization. For carotenes at the S0 state, lycopene has lower rotational barriers and higher rotational rates than its β-carotene counterparts. Lycopene has a more low-lying triplet state than β-carotene, which may be responsible for its enhanced singletoxygen quenching ability. On the basis of our computed data, it can be suggested that, at room temperature, the low-lying isomers of carotenes are in equilibrium with the all-trans isomer in the following order: all-trans > 9-cis > 13-cis > 15-cis. The rule of thumb for evaluating the relative stability of carotene isomers derived from this study is as follows: (1) all-trans isomers are the most stable; (2) the methylated cis isomers are lower in energy than the unmethylated cis ones, and the peripheral methylated cis isomers are lowest in energy; (3) among the unmethylated cis isomers, central-cis is the most stable. Among all the cis isomers, the 9-cis isomers are extraordinarily stable. The presence of 9-cis isomers in photosynthetic systems may be attributed to their stability and antioxidative ability. Acknowledgment. The authors acknowledge that the National Science Council of Taiwan, Republic of China, supported this work. We also thank the National Center for High-Performance Computing in Taiwan for computer time and facilities. Supporting Information Available: Geometrical parameters of compounds and transition states optimized at the B3LYP/631G(d) level. This material is available free of charge via the Internet at http://pubs.acs.org. References and Notes (1) Koyama, Y.; Fujii, R. The Photochemistry of Carotenoids; Kluwer Academic: Dordrecht, The Netherlands, 1999. (2) Telfer, A. Philos. Trans. R. Soc. London, Ser. B 2002, 357, 1431– 1440. (3) Edge, R.; McGarvey, D. J.; Truscott, T. G. J. Photochem. Photobiol., B 1997, 41, 189–200. (4) Sies, H.; Stahl, W. Am. J. Clin. Nutr. 1995, 62, 1315S–1321S. (5) Di Mascio, P.; Murphy, M. E.; Sies, H. Arch. Biochem. Biophys. 1989, 274, 532–538. (6) Rock, C. L. Carotenoids in Health and Disease; Mercer Dekker: New York, 2004. (7) Ames, B.; Shigenaga, M.; Hagen, T. Proc. Natl. Acad. Sci. U.S.A. 1993, 90, 7915–7922.

J. Phys. Chem. B, Vol. 112, No. 38, 2008 12167 (8) Di Mascio, P.; Murphy, M. E.; Sies, H. Am. J. Clin. Nutr. 1991, 53, 194S–200S. (9) Burton, G. W.; Ingold, K. U. Science 1984, 224, 569–573. (10) Truscott, T. G. J. Photochem. Photobiol., B 1996, 35, 233–235. (11) Polidori, M. C.; Stahl, W.; Eichler, O.; Niestroj, I.; Sies, H. Free Radical Biol. Med. 2001, 30, 456–462. (12) Woggon, W.-D.; Kundu, M. K. Carotenoids in Health and Disease; Mercer Dekker: New York, 2004. (13) Stahl, W.; Schwarz, W.; Sundquist, A. R.; Sies, H. Arch. Biochem. Biophys. 1992, 294, 173–177. (14) Boileau, A. C.; Merchen, N. R.; Wasson, K.; Atkinson, C. A.; Erdman, J. W. J. Nutr. 1999, 129, 1176–1181. (15) Aman, R.; Schieber, A.; Carle, R. J. Agric. Food Chem. 2005, 53, 9512–9518. (16) Shi, J.; Le Maguer, M. Crit. ReV. Food Sci. Nutr. 2000, 40, 1–42. (17) Ashikawa, I.; Miyata, A.; Koike, H.; Inoue, Y.; Koyama, Y. Biochemistry 1986, 25, 6154–6160. (18) Yan, J.; Liu, Y.; Mao, D.; Li, L.; Kuang, T. Biochim. Biophys. Acta 2001, 1506, 182–188. (19) Zuo, P.; Li, B.-X.; Zhao, X.-H.; Wu, Y.-S.; Ai, X.-C.; Zhang, J.P.; Li, L.-B.; Kuang, T.-Y. Biophys. J. 2006, 90, 4145–4154. (20) Kuki, M.; Koyama, Y.; Nagae, H. J. Phys. Chem. 1991, 95, 7171– 7180. (21) Doering, W. v. E.; Sotiriou-Leventis, C.; Roth, W. R. J. Am. Chem. Soc. 1995, 117, 2747–2757. (22) Burke, M.; Land, E. J.; McGarvey, D. J.; Truscott, T. G. J. Photochem. Photobiol., B 2000, 59, 132–138. (23) Tavan, P.; Schulten, K. Phys. ReV. B 1987, 36, 4337–4358. (24) Doering, W. v. E.; Sarma, K. J. Am. Chem. Soc. 1992, 114, 6037– 6043. (25) Nagae, H.; Kakitani, Y.; Katoh, T.; Mimuro, M. J. Chem. Phys. 1993, 98, 8012–8023. (26) Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.; Montgomery, J. A., Jr.; Vreven, T.; Kudin, K. N.; Burant, J. C.; Millam, J. M.; Lyengar, S. S.; Tomasi, J.; Barone, V.; Mennucci, B.; Cossi, M.; Scalmani, G.; Rega, N.; Petersson, G. A.; Nakatsuji, H.; Hada, M.; Ehara, M.; Toyota, K.; Fukuda, R.; Hasegawa, J.; Ishida, M.; Nakajima, T.; Honda, Y.; Kitao, O.; Nakai, H.; Klene, M.; Li, X.; Knox, J. E.; Hratchian, H. P.; Cross, J. B.; Adamo, C.; Jaramillo, J.; Gomperts, R.; Stratmann, R. E.; Yazyev, O.; Austin, A. J.; Cammi, R.; Pomelli, C.; Ochterski, J.; Ayala, P. Y.; Morokuma, K.; Voth, G. A.; Salvador, P.; Dannenberg, J. J.; Zakrzewski, V. G.; Dapprich, S.; Daniels, A. D.; Strain, M. C.; Farkas, O.; Malick, D. K.; Rabuck, A. D.; Raghavachari, K.; Foresman, J. B.; Ortiz, J. V.; Cui, Q.; Baboul, A. G.; Clifford, S.; Cioslowski, J.; Stefanov, B. B.; Liu, G.; Liashenko, A.; Piskorz, P.; Komaromi, I.; Martin, R. L.; Fox, D. J.; Keith, T.; Al-Laham, M. A.; Peng, C. Y.; Nanayakkara, A.; Challacombe, M.; Gill, P. M. W.; Johnson, B.; Chen, W.; Wong, M. W.; Gonzalez, C.; Pople, J. A. Gaussian 03, revision B.05; Gaussian, Inc.: Pittsburgh, PA, 2003. (27) McDouall, J. J.; Peasley, K.; Robb, M. A. Chem. Phys. Lett. 1988, 148, 183. (28) Hegarty, D.; Robb, M. A. Mol. Phys. 1979, 38, 1795. (29) Becke, A. D. J. Chem. Phys. 1993, 98, 5648–5652. (30) Lee, M.-T.; Hu, C.-H. Organometallics 2004, 23, 976–983. (31) Andersson, K.; Malmqvist, P.-A.; Ross, B. O. J. Chem. Phys. 1992, 1218–1226. (32) Bernardi, F.; Garavelli, M.; Olivucci, M.; Robb, M. A. Mol. Phys. 1997, 92, 359–364. (33) Yamaguchi, K.; Jensen, F.; Dorigo, A.; Houk, K. N. Chem. Phys. Lett. 1988, 149 (5-6), 537–542. (34) Jarowski, P. D.; Diederich, F.; Houk, K. N. J. Phys. Chem. A 2006, 110, 7237–7246. (35) Chen, B. H.; Peng, H. Y.; Chen, H. E. J. Agric. Food Chem. 1995, 43, 1912–1918. (36) Pesek, C. A.; Warthesen, J. J. J. Agric. Food Chem. 1990, 38, 1313– 1315. (37) Watanabe, K.; Kon, M.; Takahashi, B.; Hirota, S. Food Sci. Technol. Res. 1999, 5, 308–310. (38) Hu, Y.; Hashimoto, H.; Moine, G.; Hengartner, U.; Koyama, Y. J. Chem. Soc., Perkin Trans. 2 1997, 2699–2710. (39) Tsukida, K.; Saiki, K. J. Nutr. Sci. Vitaminol. 1982, 28, 311–313. (40) Hashimoto, H.; Koyama, Y. J. Phys. Chem. 1988, 92, 2101–2108. (41) Zhang, H.; Huang, D.; Cramer, W. A. J. Biol. Chem. 1999, 274, 1581–1587. (42) Levin, G.; Mokady, S. Free Radical Biol. Med. 1994, 17, 77–82. (43) Liu, X.; Osawa, T. Biochem. Biophys. Res. Commun. 2007, 357, 187–193. (44) Douglas, J. E.; Rabinovitch, B. S.; Looney, F. S. J. Chem. Phys. 1955, 23, 315–323. (45) Cundall, R. B.; Palmer, T. F. Trans. Faraday Soc. 1961, 57, 1936–1941.

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