Reactions of Amine and Peroxynitrite: Evidence for Hydroxylation as

Jul 6, 2012 - ABSTRACT: Peroxynitrite is related to numerous diseases including cardiovascular diseases, inflammation, and cancer. In order to expand ...
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Reactions of Amine and Peroxynitrite: Evidence for Hydroxylation as Predominant Reaction and New Insight into the Modulation of CO2 Zhi Sun, Yong Dong Liu,* and Ru Gang Zhong College of Life Science & Bioengineering, Beijing University of Technology, Beijing 100124, P. R. China S Supporting Information *

ABSTRACT: Peroxynitrite is related to numerous diseases including cardiovascular diseases, inflammation, and cancer. In order to expand the understanding for the toxicology of peroxynitrite in biological system, the reactions of amine (morpholine as a probe) with peroxynitrite and the modulation of CO2 were investigated by using DFT methods. The results strongly indicate that the hydroxylation of amine by peroxynitrous acid ONOOH, which was previously overlooked by most studies, is predominant relative to the widely reported nitration and nitrosation in the absence of CO2. The product N-hydroxylamine is proposed to be mainly generated via nonradical pathway (two-electron oxidation). The modulation of CO2 exhibits two main functions: (1) inhibition of hydroxylation due to the promoted consumption of peroxynitrite via fast reaction of CO2 with ONOO¯ to form ONOOCO2¯; (2) dual effect (catalysis and inhibition) of CO2 toward nitration and nitrosation. As a new insight, amine does react with CO2 and produce inert amine carbamate R2NCOO¯. This reaction has the potential to compete with the reaction of CO2 and ONOO¯, which leads to inhibition of nitration and nitrosation. The concentration of CO2 could be a critical factor determining the final effect, catalysis or inhibition. As a new finding, HCO3¯ is probably an effective catalyst for the reaction of amine and CO2. Moreover, further studies on how the different types of the amine might affect the outcome of the reactions would be an interesting topic.

1. INTRODUCTION

suggested to be disease-related because of their high reactivity toward biological substrates.16

Reactive nitrogen oxide species (RNOS), which are known as dinitrogen trioxide (N2O3), peroxynitrite (ONOO¯), and other related species, are derived from the NO-related reactions (such as •NO/O2 or •NO/O2•¯) in biological systems.1,2 As a class of intermediates with strong bioactivity, RNOS were found to react with DNA, lipids, and amino acids, which could cause various modifications to biological tissues.3−5 A great deal of research indicates that the existence of RNOS is relevant to various diseases, such as neurodegenerative diseases, cardiovascular diseases, and cancer.6−10 Among the members of RNOS, peroxynitrite is especially important due to its rich chemistry in vivo. During some pathological processes, such as inflammation, the rapid reaction between •NO and superoxide O2•¯ (eq 1) is generally considered to be the main source of peroxynitrite ONOO¯ in biological systems.11 The fate of peroxynitrite mainly involves two reaction pathways: (a) protonation of ONOO¯ (pKa = 6.8) to generate peroxynitrous acid ONOOH (eq 2);12 (b) fast reaction with CO2 to form a short-lived intermediate nitrosoperoxycarbonate ONOOCO2¯ (eq 3).13,14 Both ONOOH and ONOOCO 2 ¯ undergo subsequent decomposition (eqs 4−5) to generate metastable radicals, such as •OH, •NO2, and CO3•¯.12,15 These radicals were © 2012 American Chemical Society

•

NO + O2•− → ONOO−

(1)

ONOO− + H+ → ONOOH −

ONOO + CO2 → ONOOCO2

(2) −

(3)

ONOOH → HO• + •NO2

(4)

ONOOCO2− → •NO2 + CO3•−

(5)

Reactions induced by peroxynitrite and its derived species have been widely investigated. Among them, most studies focused on the nitration of tyrosine (or the model compounds, phenol and its derivatives)17−19 because this reaction is viewed as an important biomarker for the formation of peroxynitrite in vivo.20 However, peroxynitrite was also found to induce nitrosative stress in subsequent research,18,21−23 and nitrosation is even preferred over nitration under alkaline conditions.23 In addition to phenol and its derivatives, amines are also capable of undergoing nitration and nitrosation by peroxynitrite.21−23 Received: May 3, 2012 Revised: July 5, 2012 Published: July 6, 2012 8058

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Scheme 1. Nitration and Nitrosation of Amine by Peroxynitrite in the Absence and Presence of CO2

Experimental studies22,23 have proposed a radical mechanism (Scheme 1) in which nitration and nitrosation of amine (morpholine, MorH) by peroxynitrite can be modulated by CO2. Recently, hydroxylation of amine by peroxynitrite has been found to be significant in the absence of CO2.24 All these reactions (nitration, nitrosation, and hydroxylation) are important in consisting of the rich chemistry of peroxynitrite in vivo, and understanding these reactions at the mechanistic level is of particular importance in developing therapeutic strategies against the peroxynitrite-related diseases. The mechanism of peroxynitrite-mediated nitration of tyrosine has been recently investigated by Gunaydin et al.19 using the CBS-QB3 method. Until now, however, few theoretical studies have directly focused on the reactions of amine with peroxynitrite. This is of significance because amine structures (e.g., amino acids) are ubiquitous in biological systems, and a considerable fraction of pharmaceuticals contains amine, amide, or other amine-like groups.25,26 Note that the different molecular structures between tyrosine (phenol as the basic structure) and amine (generally contains a sp3-hybridized N atom as the reaction site) may cause differences in mechanism and thermochemistry when reactions occur. Therefore, one aim of the present study is to gain more insight into the reaction mechanism of amine with peroxynitrite. With regard to the modulation of CO2, a low level of CO2 promotes nitration or nitrosation, but excess CO2 shows an inhibitory effect.22 Experimental studies proposed that the dual effect of CO2 results from the reaction of ONOO¯ with CO2 and the subsequent radical process,18,23 but few studies take the possible effect of the reaction between amine and CO2 into account, especially in the presence of excess CO2. It is known that biological carbamate formation occurs in CO2 transport by hemoglobin in vivo,27,28 and the reaction of amine with CO2 is utilized as an important technology to remove greenhouse gas CO2.29−31 Significantly, this reaction was also found to inhibit the N2O3-induced formation of nitrosamine in vivo.32 Based on the above statement, is it possible that, at least with high level of CO2, the reaction of amine and CO2 could affect the modulation of CO2 in the amine/peroxynitrite reaction system, which is overlooked before? Another aim of this study is to explore this possibility. In short, the reaction of amine with peroxynitrite is complicated at the mechanistic level. To better understand the chemistry of peroxynitrite in vivo, this paper focuses on the reaction of amine with peroxynitrite, including nitration, nitrosation, and hydroxylation. Morpholine is selected as a

probe because it gives few side reactions and produces wellcharacterized products in experimental studies.22,23 The modulation of CO2 on the reactions will be discussed, and new insights into the mechanism will be presented.

2. THEORETICAL METHODS Density functional theory (DFT) calculations were performed by using the B3LYP method (Becke’s three-parameter functional33 with the correlation functional of Lee, Yang, and Parr34), in conjunction with the 6-311+G(d,p) basis set.35 All the structures of the reactants, transition states, intermediates, and products were fully optimized. Vibrational frequencies were also calculated at the same level to characterize the nature of each stationary point. The intrinsic reaction coordinate (IRC)36 calculation was performed to confirm that every transition state connects with the corresponding reactant and product through the minimized-energy pathway. The solvent effects were considered by single-point calculations on the gas phase geometries with the conductor-like polarizable continuum model (CPCM)37 (water as the solvent), namely, CPCMB3LYP/6-311+G(d,p)//B3LYP/6-311+G(d,p). According to the results from Takano and Houk,38 UAKS (united atom topological model applied on radii optimized for the PBE1PBE/6-31G(d) level of theory) cavity was selected. All free energies reported are corrected by the solvation free energy calculated at the CPCM-B3LYP/6-311+G(d,p) level, and this strategy has recently been used to provide reliable estimations of energy.39−41 All calculations were performed with the Gaussian 03 program package.42 3. RESULTS AND DISCUSSION The reactions of morpholine with peroxynitrite will be discussed in the absence and presence of CO2, respectively. Furthermore, in the presence of CO2, the potential interaction between CO2 and morpholine will be explored to check whether and how the reactions are affected. 3.1. Reactions of Morpholine with ONOO¯/ONOOH. Under physiological conditions, ONOO¯ (pKa = 6.8) is partially protonated to ONOOH. The conformers of ONOO¯ and ONOOH have been discussed in detail elsewhere.43−46 Peroxynitrite exists in two conformers, cisONOO¯ and trans-ONOO¯, while peroxynitrous acid mainly exists in the form of cis, cis-ONOOH and trans, perp-ONOOH (Figure 1). The cis, cis-ONOOH was predicted to be slightly more stable than trans, perp-ONOOH, and the difference in free energy is only 1.7 kcal mol−1. All the four species were taken into consideration. 8059

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Table 1. Activation Free Energies (ΔG⧧sol, in kcal mol−1) and Reaction Free Energies (ΔGsol, in kcal mol−1) for the Nitration, Nitrosation, and Hydroxylation of Morpholine by ONOOH via Nonradical Pathways

Figure 1. B3LYP/6-311+G(d,p)-optimized structures of cis-ONOO¯, trans-ONOO¯, cis, cis-ONOOH, and trans, perp-ONOOH (distances in Å).

3.1.1. Reactions of Morpholine with ONOO¯/ONOOH via Nonradical Pathways. (a). Nitration of Morpholine. No feasible nonradical pathways were found for the nitration of morpholine (MorH) by ONOO¯. The transition states for cis, cis-ONOOH and trans, perp-ONOOH, TS1 and TS2 as shown in Figure 2, were located for the nitration of morpholine. Both TS1 and TS2 involve the transfer of an ONO moiety of ONO− OH to MorH to produce Mor−NO2 and the H-abstraction by an OH moiety to release a H2O molecule. As shown in Table 1, the nitration of morpholine by cis, cis-ONOOH (with the activation free energy ΔG⧧sol being 17.6 kcal mol−1) occurs easier than the nitration by trans, perp-ONOOH (ΔG⧧sol = 28.7 kcal mol−1), and both are highly exothermic with the ΔGsol being above −50 kcal mol−1. (b). Nitrosation of Morpholine. Similar to nitration, the nitrosation of morpholine by ONOO¯ hardly occurs via nonradical mechanism. Although the corresponding transition states were located, high activation free energies were predicted. Therefore, ONOO¯ per se is not an effective nitrosating agent in vivo. This result supports the assumption from Williams47 that the direct nitrosation by ONOO¯ is unlikely to occur because O22¯ is a very poor leaving group from its parent molecule ONOO¯ to generate nitrosating species ON+. Peroxynitrous acid ONOOH reacts with morpholine to form N-nitrosomorpholine (Mor−NO) via two four-membered cyclic transition states TS3 and TS4 for cis, cis-ONOOH and

reactions

ΔG⧧sol

ΔGsol

MorH + cis, cis-ONOOH → Mor−NO2 + H2O MorH + trans, perp-ONOOH → Mor−NO2 + H2O MorH + cis, cis-ONOOH → Mor-NO + H2O2 MorH + trans, perp-ONOOH → Mor−NO + H2O2 MorH + cis, cis-ONOOH → Mor−OH + H−ONO (concerted) MorH + cis, cis-ONOOH → MorHO + H−ONO (stepwise) MorH + trans, perp-ONOOH → MorHO + H−NO2 (stepwise) MorHO + H2O → Mor−OH + H2O (one H2O-assisted H-transfer) MorHO + 2H2O → Mor−OH + 2H2O (two H2Oassisted H-transfer)

17.6 28.7 24.5 16.9 14.7

−50.9 −54.5 −11.0 −21.5 −20.6

4.2

−18.3

6.4

−23.8

12.1

−7.0

8.8

−6.1

trans, perp-ONOOH, respectively, as shown in Figure 3. In contrast to nitration, trans, perp-ONOOH (ΔG⧧sol = 16.9 kcal mol−1 and ΔGsol = −21.5 kcal mol−1) shows a stronger nitrosating ability than cis, cis-ONOOH (ΔG⧧sol = 24.5 kcal mol−1 and ΔGsol = −11.0 kcal mol−1) via the nonradical pathway. As shown in Table 1, the nitrosation trans, perpONOOH could compete with the nitration kinetically, but the latter is more favored in thermodynamics. (c). Hydroxylation of Morpholine. With regard to hydroxylation, two different pathways (Scheme 2) were located to form N-hydroxymorpholine (Mor−OH): (a) concerted pathway, ONOOH-mediated hydroxylation via cyclic transition states TS5-1 and TS5-2; (b) stepwise pathway, oxygen transfer of ONOOH to MorH (namely, two-electron oxidation48−52) and subsequent H2O-assisted H-transfer reaction. For the concerted pathway in Scheme 2, the cis, cis-ONOOH reacts with morpholine to form Mor−OH via a six-membered cyclic transition state TS5−1 (Figure 4). Activation free energy for this hydroxylation was calculated to be 14.7 kcal mol−1, and the reaction is exothermic (ΔGsol = −20.6 kcal mol−1). Note that its activation free energy is slightly lower than the barriers of the nitration and nitrosation (see Table 1). It is unable to locate the hypothetic transition state TS5-2 (the dashed line in Scheme 2) for trans, perp-ONOOH. Because the isomerization from trans, perp-ONOOH to cis, cis-ONOOH occurs easily,53 we propose that trans, perp-ONOOH could first transform to

Figure 2. B3LYP/6-311+G(d,p)-optimized transition states for the nitration of morpholine by cis, cis-ONOOH and trans, perp-ONOOH via nonradical pathways (distances in Å). 8060

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Figure 3. B3LYP/6-311+G(d,p)-optimized transition states for the nitrosation of morpholine by cis, cis-ONOOH and trans, perp-ONOOH via nonradical pathways (distances in Å).

Scheme 2. Hydroxylation of Morpholine by ONOOH via Concerted and Stepwise Pathways

Compared with the first step (two-electron oxidation), the H-transfer appears to be the rate-limiting step for the stepwise pathway. However, even on this basis, compared with nitration and nitrosation (see Table 1), the hydroxylation is still the most favorable reaction among the nonradical pathways. Instead of the H2O-assisted H-transfer, another possibility is that the oxygen atom in IM may capture H+ in solution to generate Nhydroxymorpholine (see Scheme 2). 3.1.2. Reactions of Morpholine with ONOO¯/ONOOH via Radical Pathways. The radical pathways in the absence of CO2 were studied mainly on the basis of the experimentally proposed mechanisms shown in Scheme 1. Under physiological conditions, ONOOH is expected to undergo decomposition to give the “caged radical pair” [•NO2···HO•] in solution. Based on experimental results,54 about 70% of the caged radical pair [•NO2···HO•] form the stable NO3¯. Only about 30% escape as free radicals HO• and •NO2. These free radicals were suggested to oxidize MorH into Mor• radical, and the nitrated and nitrosated products are generated from the recombination of

cis, cis-ONOOH which then reacts with morpholine to give Mor−OH. For the stepwise pathway in Scheme 2, morpholine first reacts with ONOOH to form an intermediate IM which undergoes subsequent H-transfer to generate Mor−OH. In the first step, morpholine reacts with cis, cis-ONOOH and trans, perp-ONOOH via TS6-1 and TS6-2 (Figure 4), respectively. Both TS6-1 and TS6-2 involve the oxygen transfer from ONOOH to MorH and intramolecular H-abstraction of ONOOH with the release of cis-HONO and HNO 2 , respectively. Significantly, low activation free energies (ΔG⧧sol = 4.2 and 6.4 kcal mol−1, for cis, cis-ONOOH and trans, perpONOOH, respectively) were obtained and both are favorable in thermodynamics with ΔGsol being around −20 kcal mol−1. After the formation of IM, the H-transfer (second step) occurs easily with the assistant of H2O to form Mor−OH (one H2O molecule: ΔG⧧sol = 12.1 kcal mol−1 and ΔGsol = −7.0 kcal mol−1; two H2O molecules: ΔG⧧sol = 8.8 kcal mol−1 and ΔGsol = −6.1 kcal mol−1, see Table 1 and the corresponding transition structures TSwater-1 and TSwater-2 in Figure 4). 8061

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Figure 4. B3LYP/6-311+G(d,p)-optimized transition states for the hydroxylation of morpholine by ONOOH via concerted and stepwise pathways (distances in Å).

Figure 5. B3LYP/6-311+G(d,p)-optimized transition states for the one-electron oxidation of morpholine by HO•, •NO2, and CO3•¯ radicals (distances in Å).

activation energy was calculated to be 18−19 kcal mol−1 at the UCCSD/6-311++G(d,p) level by Zhao et al.53 This value is close to the reported value obtained by Bach et al.56,57 as well as the result from recent molecular dynamics simulation.58 (b). One-Electron Oxidation of Morpholine by HO• and • NO2 Radicals. As shown in Figure 5 (TS7), the oxidation of

these radicals. The detailed mechanisms of these reactions are discussed below. (a). Decomposition of ONOOH. The experimental activation free energy for the homolytic dissociation of ONOOH was reported to be about 16 kcal mol−1.55 The mechanism53,56,57 and dynamics58 have been recently studied in detail. The 8062

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different mechanisms. That is, the radical mechanisms are somewhat preferred over the nonradical mechanism for nitration and nitrosation, whereas the predominant hydroxylation prefers nonradical pathways, especially for the stepwise mechanisms. With regard to the predominant hydroxylation, indirect evidence for the nonradical pathways come from the experiments 48−52 in which the “two-electron oxidation” of methionine48 or selenomethionine50 by peroxynitrite occurs with a second-order kinetics, predicting a direct reaction between the substrate and peroxynitrite (ground-state ONOOH was suggested to be the two-electron oxidant). Although different substrates were used in these experiments,48,50 theoretical studies49,51,52 indicate that the twoelectron oxidation of NH3,49,52 H2S,49,52 Me2S,51 and Me2Se51 by ONOOH occur via very similar mechanisms. This implies that the two-electron oxidation of amine (such as morpholine) by peroxynitrite probably occur with a similar kinetics to those of Me2S and Me2Se, that is, a direct reaction of amine and ONOOH. On this basis, the hydroxylation of morpholine via the stepwise pathway proposed in Scheme 2 would be rational. 3.2. Reactions of Morpholine with ONOO¯/CO2. Different from the case in the absence of CO2, the modulation of the reactivity of peroxynitrite by CO2 is widely believed to be the result of the reaction of peroxynitrite with CO2 (eq 3).13,14,17,59 The reaction of ONOO¯ and CO2 is rapid (k = 3.0 × 104 M−1 s−1),13 and because the concentration of CO2 in vivo is high (about 1 mM), it makes the reaction of ONOO¯/CO2 a main reaction for ONOO¯ in biological system. Its product ONOOCO2¯ is expected to undergo fast decomposition to give the “caged radicals” [•NO2···CO3•¯] in solution (see Scheme 1). Similar to the case of ONOOH, about 70% of the caged radicals [•NO2···CO3•¯] form the stable NO3¯, and only about 30% escape as free radicals CO3•¯ and •NO2, which are responsible for the biological reactions of peroxynitrite.14,54 Note that, in the presence of CO2, the fast reaction of ONOO¯ with CO2 would continuously shift the equilibrium between ONOO¯ and ONOOH (eq 2) to the ONOO¯ side, and then the hydroxylation would be inhibited. This explains the experimental results that, only in the absence of CO2, hydroxylamine is the main product relative to nitramine and nitrosamine.24 Accordingly, only nitration and nitrosation are discussed below. 3.2.1. Nitration and Nitrosation of Morpholine by ONOO¯/ CO 2 via Radical Pathways. After the homolysis of ONOOCO2¯, the nascent one-electron oxidant CO3•¯ is expected to react with MorH to generate Mor• radical, which was then followed by the recombination of Mor• with •NO2 and •NO to complete nitration and nitrosation, respectively (see Scheme 1). As shown in Figure 5 (TS9), the oxidation of MorH by CO3•¯ mainly involves the H-abstraction from MorH by CO3•¯ to generate Mor• radical with a HCO3¯ anion released. The activation free energy was calculated to be 5.9 kcal mol−1 (Table 2), which is slightly lower than the activation energy 6.8 kcal mol−1 for the oxidation induced by •NO2 radical. Moreover, the thermodynamic data in Table 2 for the two reactions indicate that the oxidation by CO3•¯ is slightly more favored than that of •NO2. It infers that CO3•¯ [E°(CO3•¯/CO32¯) = 1.5 V] is a more effective oxidant than • NO2 [E°(•NO2/NO2¯) = 1.04 V].60 After the oxidation of morpholine, recombination of the radicals Mor•, •NO2, and •NO are expected to occur to

MorH by hydroxyl radical mainly involves the H-abstraction of MorH by HO• to generate Mor• radical with a H2O molecule released. The activation free energy was calculated to be 2.2 kcal mol−1 (see Table 2). Such low barrier height indicates that Table 2. Activation Free Energies (ΔG⧧sol, in kcal mol−1) and Reaction Free Energies (ΔGsol, in kcal mol−1) for the Nitration, Nitrosation, and Hydroxylation of Morpholine by ONOOH via Radical Pathways reactions •

•

MorH + HO → Mor + H2O MorH + •NO2 → Mor• + HONO MorH + CO3•¯ → Mor• + HCO3¯ Mor• + HO• → Mor−OH Mor• + •NO → Mor−NO Mor• + •NO2 → Mor−NO2

ΔG⧧sol

ΔGsol

2.2 6.8 5.9 NA NA NA

−16.7 8.3 3.7 −40.2 −32.0 −28.9

the hydroxyl radical as a potent oxidant could react very fast with morpholine to generate Mor• radical, and this is consistent with experiment.23 The oxidation of MorH by •NO2 mainly involves the Habstraction of MorH to generate Mor• radical. The transition state TS8 (Figure 5) was located for the reaction with the releasing of nitrous acid HONO. The activation free energy was calculated as 6.8 kcal mol−1, and the reaction was predicted to be endothermic. The different barrier heights indicate that the • NO2 is a kinetically and thermodynamically less potent oxidant than HO•. (c). Recombination of Radicals To Form Nitramine, Nitrosamine, and Hydroxylamine. After the oxidation of morpholine, recombinations of the nascent radicals Mor•, • NO2, •NO, and HO• are expected to occur to generate Mor−NO2, Mor−NO, and Mor−OH. An orbital analysis was shown in Scheme 3. The singly occupied orbital on N atom is Scheme 3. Radical Recombination To Form Mor−NO2, Mor−NO, and Mor−OH

almost orthogonal to the lone pair orbital, and the reaction is initiated by the attack of X• radical (•NO2, •NO, or HO•) on the singly occupied orbital. Once the radical X• is close enough to the N atom of Mor•, a covalent N−X bond forms to give Mor−NO2, Mor−NO, and Mor−OH. The energy data in Table 2 indicate that all radical combinations are highly exothermic (ΔGsol = −40.2, −32.0, and −28.9 kcal mol−1, for the formation of Mor−OH, Mor− NO, and Mor−NO2, respectively). However, it should be noted that these radical combinations should be diffusion-limited in biological milieu. Since the less reactive radicals (•NO2 and • NO) tend to accumulate over the course of the reaction, their transient levels will be much higher than HO•. Therefore, nitration and nitrosation will be favored over hydroxylation by radical recombination. 3.1.3. Further Discussion for the Morpholine/Peroxynitrite Reaction System. Comparison of the energy data in Tables 1 and 2 leads to an interesting conclusion that the nitration, nitrosation, and hydroxylation of amine probably occur via 8063

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Figure 6. B3LYP/6-311+G(d,p)-optimized transition states for the reactions of morpholine and CO2 (distances in Å).

MorCOOH → MorCOO− + H+

generate Mor−NO2 and Mor−NO, which is shown in Scheme 3. 3.2.2. Prediction of the Possible Effect of CO2 on Amine. The reaction of amine with CO2 to produce amine carbamate is a well-known technology to remove greenhouse gas CO229−31,61,62 (for morpholine, the rate constant k = ca. 2.2 × 104 M−1 s−1 at 25 °C30). Note that this value is close to the rate constant for the reaction of CO2 and ONOO¯ (k = 3.0 × 104 M−1 s−1).13 Furthermore, it has been found that the generation of morpholine carbamate (MorCOO¯) from the reaction of morpholine and CO2 does inhibit the nitrosation of morpholine by potent nitrosating agent N2O3.32 Therefore, it is worthy to explore the possibility whether a similar inhibition is present in the amine/peroxynitrite/CO2 reaction system. Four pathways were found for the reaction of morpholine and CO2 to generate morpholine carbamic acid (MorCOOH, eq 6) or carbamate (MorCOO¯, eq 7). As shown in Figure 6, they are the nonassisted pathway (TS10), H2O-assisted pathway (TS11 and TS12), MorH-assisted pathway (TS13), and HCO3¯-assisted pathway (TS14). MorH + CO2 → MorCOOH

(7)

Energy data in Table 3 indicate that the direct reaction of MorH and CO2 (nonassisted pathway) hardly occurs due to Table 3. Activation Free Energies (ΔG⧧sol, in kcal mol−1) and Reaction Free Energies (ΔGsol, in kcal mol−1) for the Reaction of Morpholine and CO2 reactions MorH MorH MorH MorH MorH

+ + + + +

CO2 CO2 CO2 CO2 CO2

→ MorCO2H + H2O → MorCO2H + H2O + 2H2O → MorCO2H + 2H2O + MorH → MorCO2H + MorH + HCO3¯ → MorCO2¯ + CO2 + H2O

ΔG⧧sol

ΔGsol

37.6 21.3 17.6 8.3 13.8

1.7 −2.2 3.2 −10.6 −4.3

the high activation free energy 37.6 kcal mol−1. The activation energy of the H2O-assisted pathway decreases to be 21.3 kcal mol−1 (one water molecule, TS11) and 17.6 kcal mol−1 (two water molecules, TS12). The MorH-assisted reaction occurs with a very low activation free energy 8.3 kcal mol−1 and the ΔGsol being −10.6 kcal mol−1. The results agree very well with the experimental assumption from Crooks and Donnellan29

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4. CONCLUSIONS Peroxynitrite behaves as an endogenous toxic agent that is related to various human diseases, such as inflammation, cardiovascular diseases, and cancer. To better understand the toxicology of peroxynitrite in biological systems, the reactions of amine (morpholine a probe) with peroxynitrite and the modulation of CO2 were investigated by using DFT methods. In the absence of CO2, the hydroxylation of amine by peroxynitrous acid ONOOH, which was previously overlooked by most studies, was found to be predominant relative to the widely reported nitration and nitrosation. For hydroxylation, the nonradical pathways are preferred, and the product hydroxylamine is mainly generated from the direct reaction of amine and ONOOH. In contrast to hydroxylation, the radical mechanisms are preferred over the nonradical mechanism for nitration and nitrosation. In the presence of CO2, the hydroxylation of amine is inhibited because of the promoted consumption of peroxynitrite by fast reaction of CO2 with ONOO¯ to form ONOOCO2¯. The homolysis of ONOOCO2¯ provides a relatively stable source of the oxidant CO3•¯, which facilitates the one-electron oxidation of amine, which exhibits catalysis. However, as a new insight into the modulation of CO2, amine does react with CO2 to form inert amine carbamate R2NCOO¯. It could have the potential to compete with the reaction of CO2 and ONOO¯ and then inhibit the nitration and nitrosation, at least in the presence of a high level of CO2. Therefore, the CO2 shows dual effect toward the nitration and nitrosation in the morpholine/peroxynitrite/CO2 reaction system, and the concentration of CO2 could be a critical factor determining the final effect, catalysis or inhibition. Moreover, as a new finding, HCO3¯ is probably an effective catalyst for the reaction between amine and CO2. Note that the model compound morpholine belongs to secondary amine; however, many biological amines are primary, tertiary, or even quaternary, or exist as amides. The corresponding reactions could occur via different mechanisms leading to different products. Therefore, further studies on how the nature of the amine might affect the outcome of the reactions would be an interesting topic.

that the reaction between amine and CO2 is single-step and termolecular, for both H2O and amine molecules being capable of assisting the reaction. Surprisingly, we even found that HCO3¯, the equilibrated species of CO2 in solution, is capable of catalyzing the reaction with quite low activation energy of 13.8 kcal mol−1. In this reaction, HCO3¯ actually plays a role of an effective base that helps to subtract the H atom from amine molecule and facilitate the attack of CO2 to the amine lone pair. Because HCO3¯ is an important physiological anion in vivo and is a widely used buffer in experiments, it is worthy to further investigate the newly found catalytic role of HCO3¯ for the reaction between amine and CO2. As a conclusion, the reactions between amine and CO2 in the morpholine/peroxynitrite/CO2 reaction system are possible, and the corresponding pathways assisted by a third molecule (H2O, MorH, or HCO3¯) are more preferred than the nonassisted pathway to produce amine carbamate. 3.2.3. Further Discussion for the Morpholine/Peroxynitrite/ CO2 Reaction System. After the formation of MorCOOH, deprotonation (eq 7) occurs to give MorCOO¯ which has been found to be stable toward strong nitrosating agent N2O3.32 In the present study, the possible reaction of MorCOO¯ with ONOOH was investigated. However, all attempts to find the relevant transition states failed. As shown in Figure 7, the

Figure 7. Highest occupied molecular orbital (HOMO) of morpholine carbamate MorCOO¯.

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distribution of HOMO on N and C atoms (the expected reaction sites) in MorCOO¯ are almost negligible in the direction perpendicular to the NCOO plane, which could explain the stability of MorCOO¯ toward ONOOH. This result provides another possible explanation for the experimental fact22 that the nitration and nitrosation of amine are inhibited by a high level of CO2 which could promote the formation of inert amine carbamate. Therefore, together with the reaction of ONOO¯ and CO2, the interaction between amine and CO2 could also have the potential to modulate the peroxynitritemediated nitration and nitrosation. Based on our results, the interesting experimental result that only low levels of CO2 show catalysis, whereas a high level of CO2 exhibits the inhibitory effect,22 could be explained as the following. For catalysis, the easy formation of ONOOCO2¯ (eq 3) and its fast decomposition (eq 5) provide a more stable source of one-electron oxidant CO3•¯ than the source of HO• (ONOOH, eq 4) in the case without CO2, which demonstrates the catalytic effect of CO2.22,23 For inhibition, with a high level of CO2, the reaction of amine and CO2 could become significant, which promotes the consumption of substrate amine and then inhibit the nitration and nitrosation.

ASSOCIATED CONTENT

S Supporting Information *

Cartesian coordinates for all the reactant complexes (RC), transition states (TS), and product complexes (PC) involved in the study. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*Fax +86-10-6739-2001; e-mail [email protected]. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS The authors thank the High Performance Computing (HPC) Center in Beijing University of Technology and Beijing Computing Center for providing the high-performance computing clusters. This research was supported by National Natural Science Foundation of China (No. 20903006), Beijing Natural Science Foundation (No. 2092008), and Beijing Nova Program (No. 2008B09). 8065

dx.doi.org/10.1021/jp304290r | J. Phys. Chem. A 2012, 116, 8058−8066

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(36) Gonzalez, C.; Schlegel, H. B. J. Chem. Phys. 1989, 90, 2154− 2161. (37) Tomasi, J.; Persico, M. Chem. Rev. 1994, 94, 2027−2094. (38) Takano, Y.; Houk, K. N. J. Chem. Theory Comput. 2004, 1, 70− 77. (39) Liu, P.; Sirois, L. E.; Cheong, P. H.-Y.; Yu, Z.-X.; Hartung, I. V.; Rieck, H.; Wender, P. A.; Houk, K. N. J. Am. Chem. Soc. 2010, 132, 10127−10135. (40) Krenske, E. H.; Petter, R. C.; Zhu, Z.; Houk, K. N. J. Org. Chem. 2011, 76, 5074−5081. (41) Liu, P.; Krische, M. J.; Houk, K. N. Chem.Eur. J. 2011, 17, 4021−4029. (42) 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.; et al. Gaussian 03, revision D.01; Gaussian Inc.: Wallingford, CT, 2004. (43) Liang, B.; Andrews, L. J. Am. Chem. Soc. 2001, 123, 9848−9854. (44) González Lebrero, M. C.; Perissinotti, L. L.; Estrin, D. A. J. Phys. Chem. A 2005, 109, 9598−9604. (45) Lo, W. J.; Lee, Y. P. Chem. Phys. Lett. 1994, 229, 357−361. (46) Bach, R. D.; Glukhovtsev, M. N.; Canepa, C. J. Am. Chem. Soc. 1998, 120, 775−783. (47) Williams, D. L. H. Nitric Oxide 1997, 1, 522−527. (48) Pryor, W. A.; Jin, X.; Squadrito, G. L. Proc. Natl. Acad. Sci. U.S.A. 1994, 91, 11173−11177. (49) Houk, K. N.; Condroski, K. R.; Pryor, W. A. J. Am. Chem. Soc. 1996, 118, 13002−13006. (50) Padmaja, S.; Squadrito, G. L.; Lemercier, J. N.; Cueto, R.; Pryor, W. A. Free Radical. Biol. Med. 1996, 21, 317−322. (51) Musaev, D. G.; Geletii, Y. V.; Hill, C. L. J. Phys. Chem. A 2003, 107, 5862−5873. (52) Olson, L. P.; Bartberger, M. D.; Houk, K. N. J. Am. Chem. Soc. 2003, 125, 3999−4006. (53) Zhao, Y. L.; Houk, K. N.; Olson, L. P. J. Phys. Chem. A 2004, 108, 5864−5871. (54) Goldstein, S.; Lind, J.; Merényi, G. Chem. Rev. 2005, 105, 2457− 2470. (55) Goldstein, S.; Czapski, G.; Lind, J.; Merenyi, G. Chem. Res. Toxicol. 2001, 14, 657−660. (56) Bach, R. D.; Dmitrenko, O.; Estevez, C. M. J. Am. Chem. Soc. 2003, 125, 16204−16205. (57) Bach, R. D.; Dmitrenko, O.; Estevez, C. M. J. Am. Chem. Soc. 2005, 127, 3140−3155. (58) Gunaydin, H.; Houk, K. N. J. Am. Chem. Soc. 2008, 130, 10036− 10037. (59) Uppu, R. M.; Squadrito, G. L.; Pryor, W. A. Arch. Biochem. Biophys. 1996, 327, 335−343. (60) Stanbury, D. M. Reduction Potentials Involving Inorganic Free Radicals in Aqueous Solution. In Advances in Inorganic Chemistry; Sykes, A. G., Ed.; Academic Press: New York, 1989; Vol. 33, pp 69− 138. (61) Vaidya, P. D.; Kenig, E. Y. Ind. Eng. Chem. Res. 2008, 47, 34−38. (62) Hartono, A.; da Silva, E. F.; Svendsen, H. F. Chem. Eng. Sci. 2009, 64, 3205−3213.

REFERENCES

(1) Wink, D. A.; Mitchell, J. B. Free Radical. Biol. Med. 1998, 25, 434−456. (2) Grisham, M. B.; Jourd’Heuil, D.; Wink, D. A. Am. J. Physiol. Gastrointest. Liver Physiol. 1999, 276, G315−G321. (3) Wink, D. A.; Kasprzak, K. S.; Maragos, C. M.; Elespuru, R. K.; Misra, M.; Dunams, T. M.; Cebula, T. A.; Koch, W. H.; Andrews, A. W.; Allen, J. S.; Keefer, L. K. Science 1991, 254, 1001−1003. (4) Davis, K. L.; Martin, E.; Turko, I. V.; Murad, F. Annu. Rev. Pharmacol. Toxicol. 2001, 41, 203−236. (5) Suzuki, T.; Mower, H. F.; Friesen, M. D.; Gilibert, I.; Sawa, T.; Ohshima, H. Free Radical. Biol. Med. 2004, 37, 671−681. (6) Rubbo, H.; DarleyUsmar, V.; Freeman, B. A. Chem. Res. Toxicol. 1996, 9, 809−820. (7) Boje, K. M. K. Front. Biosci. 2004, 9, 763−776. (8) Dedon, P. C.; Tannenbaum, S. R. Arch. Biochem. Biophys. 2004, 423, 12−22. (9) Sawa, T.; Ohshima, H. Nitric Oxide 2006, 14, 91−100. (10) Szabo, C.; Ischiropoulos, H.; Radi, R. Natl. Rev. Drug. Discov. 2007, 6, 662−680. (11) Huie, R. E.; Padmaja, S. Free Radical. Res. 1993, 18, 195−199. (12) Beckman, J. S.; Beckman, T. W.; Chen, J.; Marshall, P. A.; Freeman, B. A. Proc. Natl. Acad. Sci. U.S.A. 1990, 87, 1620−1624. (13) Lymar, S. V.; Hurst, J. K. J. Am. Chem. Soc. 1995, 117, 8867− 8868. (14) Squadrito, G. L.; Pryor, W. A. Chem. Res. Toxicol. 2002, 15, 885−895. (15) Meli, R.; Nauser, T.; Latal, P.; Koppenol, W. H. J. Biol. Inorg. Chem. 2002, 7, 31−36. (16) Squadrito, G. L.; Pryor, W. A. Free Radical. Biol. Med. 1998, 25, 392−403. (17) Lemercier, J. N.; Padmaja, S.; Cueto, R.; Squadrito, G. L.; Uppu, R. M.; Pryor, W. A. Arch. Biochem. Biophys. 1997, 345, 160−170. (18) Uppu, R. M.; Lemercier, J. N.; Squadrito, G. L.; Zhang, H. W.; Bolzan, R. M.; Pryor, W. A. Arch. Biochem. Biophys. 1998, 358, 1−16. (19) Gunaydin, H.; Houk, K. N. Chem. Res. Toxicol. 2009, 22, 894− 898. (20) Crow, J. P.; Ischiropoulos, H. Methods Enzymol. 1996, 269, 185−194. (21) Uppu, R. M.; Pryor, W. A. J. Am. Chem. Soc. 1999, 121, 9738− 9739. (22) Masuda, M.; Mower, H. F.; Pignatelli, B.; Celan, I.; Friesen, M. D.; Nishino, H.; Ohshima, H. Chem. Res. Toxicol. 2000, 13, 301−308. (23) Uppu, R. M.; Squadrito, G. L.; Bolzan, R. M.; Pryor, W. A. J. Am. Chem. Soc. 2000, 122, 6911−6916. (24) Kirsch, M.; Korth, H. G.; Wensing, A.; Lehnig, M.; Sustmann, R.; de Groot, H. Helv. Chim. Acta 2006, 89, 2399−2424. (25) Brambilla, G.; Martelli, A. Mutat. Res.Rev. Mut. Res. 2007, 635, 17−52. (26) Brambilla, G.; Martelli, A. Mutat. Res.Rev. Mut. Res. 2009, 681, 209−229. (27) Kilmarti, Jv; Rossiber, L. Physiol. Rev. 1973, 53, 836−890. (28) Vandegriff, K. D.; Benazzi, L.; Ripamonti, M.; Perrella, M.; Letellier, Y. C.; Zegna, A.; Winslow, R. M. J. Biol. Chem. 1991, 266, 2697−2700. (29) Crooks, J. E.; Donnellan, J. P. J. Chem. Soc., Perkin Trans. 2 1989, 331−333. (30) Al-Juaied, M.; Rochelle, G. T. Chem. Eng. Sci. 2006, 61, 3830− 3837. (31) da Silva, E. F.; Svendsen, H. F. Int. J. Greenhouse Gas Control 2007, 1, 151−157. (32) Kirsch, M.; Korth, H. G.; Sustmann, R.; de Groot, H. Chem. Res. Toxicol. 2000, 13, 451−461. (33) Becke, A. D. J. Chem. Phys. 1993, 98, 5648−5652. (34) Lee, C. T.; Yang, W. T.; Parr, R. G. Phys. Rev. B 1988, 37, 785− 789. (35) Frisch, M. J.; Pople, J. A.; Binkley, J. S. J. Chem. Phys. 1984, 80, 3265−3269. 8066

dx.doi.org/10.1021/jp304290r | J. Phys. Chem. A 2012, 116, 8058−8066