A New Insight into the Mechanism of NADH Model Oxidation by Metal

Subscriber access provided by Kaohsiung Medical University

B: Biophysics; Physical Chemistry of Biological Systems and Biomolecules

A New Insight into the Mechanism of NADH Model Oxidation by Metal Ions in Non-Alkaline Media Jin-Dong Yang, Bao-Long Chen, and Xiao-Qing Zhu J. Phys. Chem. B, Just Accepted Manuscript • DOI: 10.1021/acs.jpcb.8b03453 • Publication Date (Web): 11 Jun 2018 Downloaded from http://pubs.acs.org on June 11, 2018

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

The Journal of Physical Chemistry

A New Insight into the Mechanism of NADH Model Oxidation by Metal Ions in Non-Alkaline Media Jin-Dong Yang,†,§ Bao-Long Chen,† and Xiao-Qing Zhu*,†,‡ †

The State Key Laboratory of Elemento-Organic Chemistry, ‡ Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), College of Chemistry, Nankai University, Tianjin 300071, P. R. China §

Center of Basic Molecular Science, Department of Chemistry, Tsinghua University, Beijing 100084, China

E-mail: [email protected] ___________________________________________________________________________________________ ABSTRACT: For a long time, it has been controversial that the three-step (e-H+-e) or two-step (e-H•) mechanism was used for the oxidations of NADH and its models by metal ions in non-alkaline media. The latter mechanism has been accepted by the majority of researchers. In this work, 1-benzyl-1,4-dihydronicotinamide (BNAH) and 1-phenyl-l,4-dihydronicotinamide (PNAH) are used as NADH models, and ferrocenium (Fc+) metal ion as an electron acceptor. The kinetics for oxidations of the NADH models by Fc+ in pure acetonitrile were monitored by using UV-Vis absorption and quadratic relationship between of kobs and the concentrations of NADH models were found for the first time. The rate expression of the reactions developed according to the three-step mechanism is quite consistent with the quadratic curves. The rate constants, thermodynamic driving forces and KIEs of each elementary step for the reactions were estimated. All the results supported the three-step mechanism. The intrinsic kinetic barriers of the proton transfer from BNAH+• to BNAH and the hydrogen atom transfer from BNAH+• to BNAH+• were estimated by using Zhu equation, the results showed that the former is 11.8 kcal/mol, and the latter is larger than 24.3 kcal/mol. It is the large intrinsic kinetic barrier of the hydrogen atom transfer that makes the reactions choose the three-step rather than two-step mechanism. Further investigation of the factors affecting the intrinsic kinetic barrier of chemical reactions indicated that the large intrinsic kinetic barrier of the hydrogen atom transfer originated from the repulsion of positive charges between BNAH+• and BNAH+•. The greatest contribution of this work is the discovery of the quadratic dependence of kobs on the concentrations of the NADH models, which is inconsistent with the conventional viewpoint of the “two-step mechanism” on the oxidations of NADH and its models by metal ions in the non-alkaline media.

________________________________________________________________________ ▉

INTRODUCTION

It is well-known that nicotinamide adenine dinucleotide coenzyme (NADH) is one type of the most important two-electron sources in living body (eq 1) and participates in many important substrate metabolisms, electron transports and energy storage processes.1-3

For example, in the respiratory

system, NADH can transmit two electrons to iron ion in Fe-S cluster through FMN, and realizes the storage of chemical energies in the ATP.4-7

In chemical laboratories and industries, the oxidations

of NADH models as two-electron sources by metal ions have also attracted the widespread attention and interest of many chemists,8-11 since these reactions have been widely applied in energy 1

ACS Paragon Plus Environment

The Journal of Physical Chemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

chemistry,12-14

Page 2 of 26

green chemistry,15-18 materials chemistry,19-21 and many others.

However, it has

been controversial that the mechanism of NADH and its model oxidations by metal ions (M+) (eq 2) is the three-step (e-H+-e) or two-step (e-H•).

By summarizing the previous investigations, a

prevailing view seems that the mechanism is dependent on the nature of reaction media.22-25, 26-27

(1)

NADH + 2M+ → NAD+ + 2M + H+

(2)

When the reactions (eq 2) are carried out in water or basic media, the reactions are generally considered to occur by the three-step mechanism (Scheme 1):22-25

The first step reaction is

electron transfer from NADH to M+ to produce NADH+• and M; the second is proton transfer from the formed NADH+• to water or a base (B:) to give NAD• and H3O+ or BH+; the third is electron transfer from the formed NAD• to another M+ to yield NAD+ and M. mole of NADH consumes two moles of M+.

The net reaction is that one

However, when the reactions are carried out in

non-alkaline media, the reactions are generally considered to occur by the two-step mechanism (Scheme 2):26-27

The first step reaction is electron transfer from NADH to M+ to produce NADH+•

and M and the second is hydrogen atom transfer from one formed NADH+• to another formed NADH+• to yield NAD+ and NADH2+ (i.e., the disproportionation of NADH+•). The net reaction is that one mole of NADH consumes one moles of M+.

The main experimental evidence for the

three-step mechanism is that the reaction stoichiometry is 1/2 for NADH/M+ and bases can greatly promote the rate of the reactions. The main experimental evidence for the two-step mechanism is that the reaction stoichiometry is 1/1 for NADH/M+, and the thermodynamic driving force of the hydrogen atom transfer from NADH+• to NADH+• is quite large. Scheme 1.

Three-step Mechanism of Reactions (Eq 2) in Water or Basic Media

2


Page 3 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Scheme 2.

Two-step Mechanism of Reactions (Eq 2) in Non-alkaline media

However, since the core structure of NADH and its models is dihydropyridine, they should act as not only two-electron donors, but also organic bases,28 which means that even in non-alkaline media, the three-step mechanism for the oxidations of NADH and its model by metal ions is also possible (Scheme 3). And in this mechanism, the reaction stoichiometry is also 1/1 for NADH/M+. Specially, from the kinetic point of view, if the reactions take place by the two-step mechanism (Scheme 2), the second step hydrogen atom transfer reaction should be very difficult. The reason is that both the reactants (NADH+• and NADH+•) carry positive charge, which is not conducive to the contact between them due to the electrostatic repulsion.

We therefore reasoned that regardless

of the basicity of reaction media, the reactions (eq 2) should use the three-step (e-H+-e) mechanism. Scheme 3.

Three-step Mechanism of Reactions (Eq 2) in Non-alkaline media

To prove the three-step mechanism of the reactions in non-alkaline media (Scheme 3), in this work, 1-benzyl-1,4-dihydronicotinamide (BNAH), 1-phenyl-l,4-dihydronicotinamide (PNAH) and 3



Page 4 of 26

their corresponding 4,4’-dideuterated compounds (BNAD and PNAD) were used as NADH models (Scheme 4), ferrocenium (Fc+) was used as an electron acceptor of metal ion, and pure acetonitrile was selected as a non-alkaline medium.

The detailed mechanism of the NADH model oxidations

by Fc+ in dry acetonitrile was investigated. Scheme 4.

▉

Structures and Name Abbreviations of NADH Models Examined in This Work

RESULTS AND DISCUSSION

The redox potentials of BNAH and PNAH as well as BNA+ and PNA+ in acetonitrile were measured by using two electrochemical methods CV and OSWV (Figure 1); the detailed experimental results are summarized in Table 1.

-1.420 V

0.220 V -0.2

-0.1

0.0

0.1

0.2

0.3

0.4

0.5

-1.8

0.6

-1.7

-1.6

-1.5

-1.4

-1.3

-1.2

E (V vs Fc)

E (V vs Fc)

Figure 1. Cyclic voltammogram (black line) and Osteryong square wave voltammogram (red line) of BNAH (left) and BNA+ (right) in anhydrous deaerated acetonitrile with 0.1M (n-Bu)4NPF6 as supporting electrolyte with a scan rate of 0.1 V/s.

Table 1. Oxidation Potentials of NADH Models and Their Radicals (NAD• Models) together with Free Energy Changes of Electron Transfer from NADH Models and NAD• Models to Fc+ in Acetonitrile NADH models BNAH PNAH

Eox(NADH)a CV OSWV 0.248 0.220 0.376 0.360

Eox(NAD•)b CV OSWV -1.445 -1.420 -1.152 -1.130

4


Go1(ET)c 5.1 8.3

Go3(ET) d -32.7 -26.1

Page 5 of 26

Oxidation potentials of NADH and NAD • models in acetonitrile derived from the peak potentials in CV and OSWV at 0.1 V/s at 298 K (V vs Fc). The reproducibility is 5 mV or better. c Free energy changes of the electron transfer from NADH models and d from NAD• to Fc+ in acetonitrile, which were derived from the OSWV data29 of Eox(NADH) and Eox(NAD•) multiplied by 23.06, respectively. The unit is kcal/mol. a,b

BNAH was treated with Fc+ in pure acetonitrile to yield Fc and the corresponding pyridinium cation derivative (BNA+).

The stoichiometry of the reactions can be

determined by using spectrophotometric titrations (see Figure 2). the stoichiometry of the reaction is 1/1 for BNAH/Fc+.

The results showed that

But when the reaction occurs in the

presence of pyridine, the stoichiometry of the reaction is 1/2 for BNAH/Fc+.

b

2.0

+

[BNAH] : [Fc ]

1.5

0:5 1:5 2:5 3:5 4:5 5:5

1.5

1.0

Abs

a

Abs

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


1.0

0.5 0.5

0.0

0.0 500

600

700

0.0

800

0.2

(nm)

0.4

0.6

0.8

1.0

+

[BNAH] : [Fc ]

Figure 2. (a) Spectral titrations for the reactions of various volume of 5.02 mM BNAH with 1.5 mL Fc+ (5.01 mM) in the pure acetonitrile to determine the reaction stoichiometry. (b) Titration plots in the absence (black line) and presence (red line) of pyridine.

Kinetics for the oxidations of NADH models (BNAH, BNAD, PNAH and PNAD) by Fc+ in acetonitrile at 298 K were monitored through the disappearance of Fc+ UV absorbance at 617 nm by using the stopped-flow technique.

When the concentration of

NADH models is 10 times greater than that of Fc+, the decay of Fc+ UV absorption at 617 nm well obeyed the pseudo-first-order kinetic rate law (Figure 3).

Since the

pseudo-first-order rate of the reactions can be influenced by the accumulated Fc, kinetic runs were carried out under the condition that the concentration of Fc keeps a near constant (about 2.5 mM) during the reactions.

The dependence of the observed pseudo-first-order

rate constants kobs of the reactions in pure acetonitrile at 298 K on the initial concentrations of NADH models are summarized in Table 2.

When the observed rate constants kobs of the

reaction were correlated to the concentrations of NADH models (Figure 4), it is surprising to find that the relation of kobs with the initial concentrations of NADH models is not a straight 5



line but a parabolic (i.e., quadratic) curve, which means that the oxidation of the NADH models by Fc+ in pure acetonitrile is not the first but second order with respect to NADH models (BNAH, BNAD, PNAH and PNAD).

a

b

4

0.10

0.10

-lnAbs

Abs

Abs

-lnAbs

4

3

3

0.05

0.05 0

1

t (s) 0

5

t (s)

0.00

0.00 0.0

2.5

0

5.0

5

10

15

t (s)

t (s)

20

4.5

c

d

4.0

0.09

4

0.09

-lnAbs

-lnAbs

3.5

3

3.0

Abs

0.06

Abs

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 6 of 26

2.5

0

20

40

60

80

0.06 2 0

100

40

80

120

160

t (s)

t (s)

0.03

0.03

0.00

0.00 0

50

100

150

0

200

100

t (s)

200

300

400

500

t (s)

Figure 3. Time profile of the UV absorbance at 617 nm due to Fc+ for the reactions of Fc+ (0.25 mM) with NADH models in deaerated acetonitrile at 298 K: (a) for BNAH, (b) for BNAD, (c) for PNAH, (d) for PNAD (top to bottom: 2.50, 3.75, 5.00, 6.25, 10.00 mM). (Inset) The corresponding pseudo-first-order plots.

Table 2. The Observed Rate Constants kobs of the Reactions of NADH Models with Fc+ (0.25 mM) at Various Concentrations of NADH Models in Pure Acetonitrile at 298 K NADH models BNAHa BNADb PNAHc PNADd

2.5 (mM) 0.614 0.161 0.012 0.0042

kobs (s-1) 5 (mM) 6.25 (mM) 2.63 3.97 0.71 1.17 0.047 0.088 0.017 0.028

3.75 (mM) 1.43 0.39 0.031 0.0093

a

Standard deviation   0.25 × 10-1. b Standard deviation   0.26 × 10-1. 0.29 × 10-2. d Standard deviation   0.27 × 10-3.

6


10 (mM) 9.76 2.98 0.19 0.069 c

Standard deviation  

Page 7 of 26

0.20 +

+

BNAD + Fc + BNAH + Fc -1

)

-1

PNAD + Fc + PNAH + Fc

0.15

kobs (s )

9

kobs (s

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


6

3

0.10

0.05

0 0.00

0.003

0.006

0.009

0.003

0.012

[BNAH] (M)

0.006

0.009

[PNAH] (M)

(a)

(b)

Figure 4. Dependence of kobs on [NADH model] in pure acetonitrile at 298 K (● for common isotope and ■ for the heavy isotope).

According to the final products and the stoichiometry of the reactions of BNAH with Fc+ in pure acetonitrile, two possible mechanisms can be proposed as shown in Scheme 2 and Scheme 3, respectively.

From the two reaction mechanisms, it is clear that the first

step in the two mechanisms is the same, i.e., electron transfer from BNAH to Fc+ to yield BNAH+• and Fc.

The difference of the two mechanisms is the following-up step (i.e., the

second step): one is proton transfer from BNAH+• to BNAH to form BNA• and BNAH2+ (i.e. deprotonation of BNAH+•), the other is hydrogen atom transfer from BNAH+• to BNAH+• to form BNA+ and BNAH2+ (i.e. disproportionation of BNAH+•).

Considering that the

NADH model oxidations by Fc+ in dry acetonitrile is the second order rather than the first order with respect to BNAH (Figure 4), the two-step mechanism (Scheme 2) can be ruled out, because in the two-step mechanism, only one elementary step needs BNAH as the reactant, and BNAH is the first order.

But, in the three-step mechanism (Scheme 3), there

are two elementary steps with BNAH as the reactant, which can make the concentration of BNAH have a power of two in the rate law of the reactions. In order to test the three-step mechanism of the reactions, the rate expression of the reactions was derived from the three-step mechanism.

Since the free energy change of the

third step electron transfer reaction in the three-step mechanism is a quite large negative value (-32.7 kcal/mol in Table 1), the rate of the third step electron transfer reaction should be diffusion-controlled, which means that the overall rate of the BNAH oxidation by Fc+ in the pure acetonitrile should be determined by the rates of the first two steps. 7


Because the


Page 8 of 26

free energy change of the first step electron transfer reaction is a positive value (5.1 kcal/mol), it is clear that the first step electron transfer reaction is reversible.

Therefore,

BNAH+• would be an unstable intermediate, and the concentration of BNAH+• can be estimated by using the steady-state approach. Since the first step electron transfer reaction is reversible, the rate of Fc+ disappearance would be equal to the sum of the rates of the second step proton transfer reaction and the third step electron transfer reaction: The total rate for the disappearance of Fc+ is -d[Fc+]/dt = k2[BNAH+•][BNAH] + k3[BNA•][Fc+]

(3)

Since the rate of the electron transfer from BNA• to Fc+ (the third step reaction) is diffusion-controlled and the reactant BNA• is the product of the second step reaction, the rate of Fc+ disappearance in the third step reaction would be equal to the rate of BNA• formation in the second step reaction (eq 4). k3[BNA•][Fc+] = k2[BNAH+•][BNAH]

(4)

When eq 4 is introduced into eq 3, then -d[Fc+]/dt = 2k2[BNAH+•][BNAH]

(5)

If the steady-state approximation is used, [BNAH+•] = k1[BNAH][Fc+]/(k-1[Fc] + k2[BNAH])

(6)

Eq 7 can be obtained from eq 5, -d[Fc+]/dt = 2k1k2[BNAH]2[Fc+]/(k-1[Fc] + k2[BNAH]) = kobs[Fc+]

(7)

In eq 7,

2k1k2 [BNAH]2  k1[Fc]  k2 [BNAH]

(8)

k1[Fc] [BNAH] 1   kobs 2k1k2 [BNAH] 2k1

(9)

kobs

kobs  2k1[BNAH]

(10)

If eq 8 is divided by the concentration of BNAH on both sides, and then inverted, eq 9 can be formed. From eq 9, it follows that if the three-step mechanism (Scheme 3) for the NADH model oxidations by Fc+ in dry acetonitrile is correct, the plots of [BNAH]/kobs vs 8


Page 9 of 26

1/[BNAH] should be straight lines and the line slope is k-1[Fc]/2k1k2, the intercept is 1/2k1. The experimental results show that the plots of [NADH]/kobs vs 1/[NADH] are all good straight lines for BNAH, PNAH and their 4.4’-dideuterated compounds as the reducing agents (see Figure 5), meaning that the three-step mechanism for the NADH model oxidations by Fc+ in dry acetonitrile (Scheme 3) conforms to the experimental results. Since the slope (S) of the lines is k-1[Fc]/2k1k2 and k1/k-1 = K1 = exp(-Go1/RT), k1, k-1 and k2 can be derived from the line slopes (S) and G1o, the results are summarized in Table 3.

0.015

+

0.6

PNAD + Fc + PNAH + Fc

+

[PNAH]/kobs

BNAD + Fc + BNAH + Fc

[BNAH]/kobs

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


0.010

0.005

0.000

0.4

0.2

0.0

0

100

200

300

400

0

100

1/[BNAH]

200

300

400

1/[PNAH]

(a)

(b)

Figure 5. Dependence of [NADH]/kobs on 1/[NADH] (● for common isotopes and ■ for the heavy isotopes).

Table 3. The Forward and Reverse Rate Constants for the First Step Reactions, the Rate Constants of the Second Step Reactions together with the KIEs of the First Step and the Second Step Reactions NADH models BNAH BNAD PNAH PNAD

Sa (×105) 0.98 3.55 49.4 147.0

k1 ×10-3 b,c 3.19 2.94 0.17 0.16

k-1 ×10-7 b,d 1.61 1.49 18.4 18.2

k2 ×10-5 b 6.30 1.79 28.0 9.39

KIE1 d

KIE2 e

1.08

3.52

1.13

2.98

a

S is the line slopes for the plots of [NADH]/kobs vs 1/[NADH]. b The rate constants, the unit is M-1 s-1. k1 obtained from the kinetic runs in the present of pyridines. d Kinetic isotope effect for the initial electron transfer. e Kinetic isotope effect for the second step proton transfer. c

From Table 3, it is clear that the rate constants for the three elementary reactions distribute in the order of k-1 (1.61 × 107) > k2 (6.30 × 105) > k1 (3.19 × 103) for BNAH as hydride donor and the kinetic isotope effects (KIE) in the initial electron transfer and the 9



followed proton transfer are 1.08 and 3.52, respectively.

Page 10 of 26

Since the observed KIE for

BNAH oxidation by Fc+ in dry acetonitrile (3.8, 3.6, 3.5, 3.4 and 3.2 when the initial concentration of BNAH is 2.5, 3.75, 5.0, and 10.0 mM, respectively) is quite close to the KIE of the proton transfer reaction, the reaction of proton transfer from BNAH+• to BNAH would be rate-determined, even though the rate constant of the proton transfer from BNAH +• to BNAH is about 200-fold larger than that of the initial electron transfer.

The reason is

that BNAH+• is a high energy reaction intermediate and the concentration of BNAH+• in the reaction system should be very small. 6.30

The value is 4.95 × 10-5 mM estimated by using eq

Since the concentration of BNAH+• (4.95 × 10-5 mM) is about 5000-fold smaller than

that of Fc+ (0.25 mM), it is clear that the rate of the proton transfer from BNAH+• to BNAH becomes 25-fold lower than that of the initial electron transfer. In order to further prove the three-step mechanism of the NADH model oxidations by Fc+ in the non-alkaline media, two following predictions can be made according to the three-step mechanism (Scheme 3) and the rate expression (eq 8):

(1) When an organic base

with stronger basicity than BNAH, such as pyridine (Py),28 is added to the reaction system, the stoichiometry of BNAH and Fc+ would change from the former 1:1 into 1:2, the plot of kobs vs [BNAH] would become a straight line, and the observed KIE at the C-H/D position of BNAH would be secondary.

The reason for the last prediction is that pyridine can greatly

accelerate the proton transfer reaction and make the initial electron transfer become the only rate-limiting step.

As a matter of fact, all the predictions are well verified by the

experimental tests (see Figures 2, 6 and 7).

(2) From eq 8, it is clear that if k2[BNAH]

increases and become much larger than k-1[Fc], eq 8 can be simplified into eq 10, i.e., a linear plot of kobs vs [BNAH] with the slope of 2k1 can be obtained.

The experimental

result is that when extra [Fc] is not added and excessive [BNAH] increases beyond 10 mM, the plot of kobs vs [BNAH] becomes a good straight line (Figure 8).

Evidently, these

additional experimental results further prove the three-step mechanism of NADH model oxidations by Fc+ in non-alkaline media.

10


Page 11 of 26

75

y = 6514x + 7.1 r = 0.996

-1

kobs (s )

60

45

30

15 2

4

6

8

10

[BNAH] (mM) Figure 6. Plot of kobs vs [BNAH] in the presence of pyridine in acetonitrile at 298 K.

a

b

+

PNAH + Fc + Py -1 kobs = 0.82 (s )

4.5

0.09

4.0

-lnAbs

+

PNAD + Fc + Py

0.06

Abs

+

PNAD + Fc + Py -1 kobs = 0.81 (s )

3.5

+

PNAH + Fc + Py

0.03

3.0 2.5 0.0

0.00 0

2

4

6

8

10

KIEobs = 1.01 0.5

1.0

1.5

2.0

t (s)

t (s)

Figure 7. (a) Time profiles of the absorbance at 617 nm due to the reaction of 0.25 mM Fc+ with 2.5 mM PNAH (red line) and PNAD (black line) in the pyridine-buffered deaerated acetonitrile at 298 K; (b) Fitted first-order plots.

120

-1

kobs (s )

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


90

y = 5849x - 19.1 r = 0.999

60

30 10

15

20

25

[BNAH] (mM) Figure 8. Dependence of kobs vs [BNAH] (beyond 10 mM) in dry acetonitrile at 298 K, when extra [Fc] is not added.

11



Page 12 of 26

Based on the above examinations, it is clear that the mechanism of the NADH model oxidations by Fc+ in non-alkaline media is three steps rather than two steps.

Why the

mechanism is the same (the three-step mechanism) whenever the reactions take place in non-alkaline or basic media?

In order to find out the reason why the NADH model

oxidations by Fc+ in dry acetonitrile choose the three-step mechanism (Scheme 3) rather than the two-step mechanism (Scheme 2), thermodynamic driving forces, concentrations of reactants and intrinsic kinetic barriers for the proton transfer from BNAH+• to BNAH and the hydrogen atom transfer from BNAH+• to BNAH+• were examined respectively. According to the oxidation potentials of BNAH (0.220 V vs Fc) and BNA• (-1.420 V vs Fc) and pKa values of BNAH+• (4.7)28 and BNAH2+ (8)28 in acetonitrile, the thermodynamic driving forces of three elementary reactions in the three-step mechanism can be obtained, 5.1,31 -4.5,32 and -32.7 (kcal/mol) for the first step electron transfer, the second step proton transfer, and the third step electron transfer, respectively (Scheme 3).

The thermodynamic

driving forces of two elementary reactions in the two-step mechanism can be also obtained, 5.1 and -42.3 (kcal/mol)32 for the first step electron transfer and the second step hydrogen atom transfer, respectively (Scheme 2).

Comparing their thermodynamic driving forces, it

is found that the thermodynamic driving forces of the proton transfer (-4.5 kcal/mol) is much smaller than that of the hydrogen atom transfer in acetonitrile (-42.3 kcal/mol),33 which indicates that the second step reaction of the BNAH oxidation by Fc+ in dry acetonitrile is not thermodynamically controlled. BNAH+• + BNAH → BNA• + BNAH2+

(11)

-d[BNAH+•]/dt = k2P[BNAH+•]×[BNAH]

(12)

BNAH+• + BNAH+• → BNA+ + BNAH2+

(13)

-d[BNAH+•]/dt = k2H[BNAH+•]2

(14)

From the proton transfer reaction (eq 11) in the three-step mechanism and the hydrogen atom transfer reaction (eq 13) in the two-step mechanism, it is found that the reactants of the two reactions are not the same.

For the proton transfer reaction, the

reactants are BNAH+• and BNAH, but for hydrogen atom transfer reaction, the two reactants are both BNAH+•.

Since the concentration of BNAH+• in the reaction system is quite small

(4.95 × 10-5 mM), it is clear that the large difference of the concentration between BNAH 12


Page 13 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(2.5 mM) and BNAH+• (4.95 × 10-5 mM) in the reaction system should be a factor that cannot be ignored to make the second step reaction of the reaction more favourable to choose the proton transfer than the hydrogen atom transfer.

However, from the rate

expression of the two elementary reactions (eqs 12 and 14), it is clear that the reaction rate not only depends on the concentration of reactants, but also depends on the activation free energy of the reactions.

Because the concentration of the reactants is an algebraic

relationship with the rate of the reactions, and the activation free energy is an exponential relationship with the rate of the reactions, it is conceived that the difference of the activation free energy between the proton transfer reaction and the hydrogen atom transfer reaction may be a main factor to make the second step reaction of the reaction choose proton transfer rather than hydrogen atom transfer.

Since activation free energy of a chemical reaction is

determined by thermodynamic driving force and intrinsic kinetic barrier of the reaction, and the second step reaction of the BNAH oxidation by Fc+ is not thermodynamically controlled, the intrinsic kinetic barrier difference between the proton transfer reaction and the hydrogen atom transfer reaction may be a decisive factor to make the second step reaction of the BNAH oxidation by Fc+ choose proton transfer rather than hydrogen atom transfer. In our previous paper,34 we developed a new chemical kinetic model (Figure 9) to estimate activation free energies or intrinsic kinetic barriers of various chemical reactions (eq 15).

Evidently,

from the new chemical kinetic model (Figure 9), eq 16 can be derived. In eq 16, G≠(XL/Y) is activation free energy of the reaction (XL + Y → X + YL), G≠Y(XL) is the free energy change of XL going from the initial state to the transition state, and G≠XL(Y) is the free energy change of Y going from the initial state to the transition state.

Since the sum of G≠Y(XL) and G≠XL(Y) is

approximately equal to the sum of G≠o(XL) and G≠o(Y) (see Figure 10),35 eq 17 can be obtained from eq 16. In eq 17, G≠o(XL) is the free energy change of XL going from the initial state to the transition state in the self-exchange reaction (XL + X → X + XL) and call as the thermo-kinetic parameter of XL.36,37,38 G≠o(Y) is the free energy change of Y going from the initial state to the transition state in the self-exchange reaction (YL + Y → Y + YL) and call as the thermo-kinetic parameter of Y.36,37,38 Since G≠o(XL) is equal to ½[G≠(XL/X) + Go(XL)] (eq 18)37 and G≠o(Y) is equal to ½[G≠(YL/Y) - Go(YL)] (eq 19),37 eq 20 can be obtained from eq 17 and was named as Zhu equation in the previous papers.36,37,38 In eq 20, Go(XL/Y) is thermodynamic driving force of 13



Page 14 of 26

the reaction (XL + Y → X + YL), G(XL/X) is the activation free energy of the self-exchange reaction (XL + X → X + XL), and G(YL/Y) is the activation free energy of the self-exchange reaction (YL + Y → Y + YL). The validity of eq 20 has been verified by predicting the activation free energies of 59904 hydride transfer reactions, 34,36 the activation free energies of 5886 hydrogen atom transfer reactions37 and the KIEs of 4556 hydride transfer reactions in acetonitrile at 298 K.38 From eq 20, it is clear that if Go(XL/Y) is zero, the average value of G≠(XL/X) and G≠(YL/Y) is equal to the activation free energy of the reaction (XL + Y → X + YL), G≠(XL/Y), which means that the average value of G≠(XL/X) and G≠(YL/Y) is the intrinsic kinetic barrier of the reaction (XL + Y → X + YL) and is symbolized by G≠o(XL/Y) (eq 21). If eq 21 is introduced into eq 20, eq 22 or eq 23 can be obtained.39 XL + Y → X + YL

(15)

(L = e, e2, H+, H•, H-, Cl-, Br•, CN-, NO, NO+.......) Mechanism:

XL + Y → [X...L...Y]≠ → X + YL

Figure 9. Kinetic model for the reaction (XL + Y → X + YL) described by two reverse Morse-type free energy curves: the left one (red) refers to the chemical process of XL to release L, the right one (black) refers to the chemical process of Y to capture L, the intersecting point refers to the transition state (TS), r is the distant between the two reactants in TS.

14


Page 15 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60




Figure 10 Relations of activation free energies between the reaction (XL + Y → X + YL) and its two self-exchange reactions (XL + X → X + XL and YL + Y → Y + YL). Morse-type free energy curves of the four partial reactions: XL → X + L (bold red line), Y + L → YL (bold black line), YL → Y + L (dashed black line), and X + L → XL (dashed red line).

G≠(XL/Y) = G≠Y(XL) + G≠XL(Y)

(16)

G≠(XL/Y) = G≠o(XL) + G≠o(Y)

(17)

Since G≠o(XL) = ½[G≠(XL/X) + Go(XL)]

(18)

G≠o(Y) = ½[G≠(YL/Y) - Go(YL)]

(19)

G≠(XL/Y) = ½[G≠(XL/X) + G≠(YL/Y)] + ½Go(XL/Y)

(20)

½[G≠(XL/X) + G≠(YL/Y)] ≡ G≠o(XL/Y)

(21)

G≠(XL/Y) = G≠o(XL/Y) + ½Go(XL/Y)

(22)

G≠o(XL/Y) = G≠(XL/Y) - ½Go(XL/Y)

(23)

or

From eq 23, it is clear that intrinsic kinetic barrier of a chemical reaction, G≠o(XL/Y), is equal to the activation free energy of the reaction, G≠(XL/Y), reduced by the half of the thermodynamic driving force of the reaction, Go(XL/Y). For the proton transfer reaction (XL = BNAH+•, Y = BNAH, L= H+) (eq 11), since G≠(XL/Y) is 9.5 kcal/mol,40 and Go(XL/Y) is -4.5 kcal/mol, G≠o(XL/Y) value of the reaction 11.8 kcal/mol can be obtained by using eq 23. For the hydrogen atom transfer reaction (XL = BNAH+•, Y = BNAH+•, L= H•) (eq 13), because the second step reaction of BNAH oxidation by Fc+ in dry acetonitrile is the proton transfer (eq 11) rather than hydrogen atom transfer (eq 13), it is clear that the rate of the hydrogen atom transfer reaction in the two-step mechanism should be slower than the rate of 15



the proton transfer reaction in the three-step mechanism.

Page 16 of 26

If the rate of the hydrogen atom

transfer reaction is assumed to be equal to the rate of the proton transfer reaction (k2H[BNAH+•]2 = k2P[BNAH+•]×[BNAH] according to eqs 12 and 14), the activation free energy of the hydrogen atom transfer reaction can be estimated according to the concentrations of BNAH (2.5 mM) and BNAH+• (4.95 × 10-5 mM) and the rate constant of the proton transfer reaction (k2P = 6.30× 105 M-1s-1 in Table 3) by using Eyring equation. The result is that the activation free energy of the hydrogen atom transfer reaction, G≠(XL/Y), is 3.12 kcal/mol.

Thus, the intrinsic kinetic barrier of the hydrogen atom

transfer reaction (24.3 kcal/mol) can be obtained from G≠(XL/Y) (3.12 kcal/mol) and Go(XL/Y) (-42.3 kcal/mol) by using eq 23.

In fact, for the hydrogen atom transfer

reaction, the real value of the intrinsic kinetic barrier could be much greater than 24.3 kcal/mol. When the intrinsic kinetic barrier of the hydrogen atom transfer reaction (˃ 24.3 kcal/mol) and that of the proton transfer reaction (11.8 kcal/mol) is compared, it is found that the former is much greater than the latter, which should be the decisive factor to make the NADH model oxidations by Fc+ in acetonitrile choose the proton transfer (eq 11) rather than the hydrogen atom transfer (eq 13) as its second step reaction.

If the intrinsic kinetic

barrier of the hydrogen atom transfer reaction were equal to the intrinsic kinetic barrier of the proton transfer reaction (11.8 kcal/mol), k2H should be 4.4 ×1019 (M-1s-1) and the rate of the hydrogen atom transfer reaction (9.31 × 104 M/s) should be larger than that of the proton transfer reaction (7.25 × 10-5 M/s) by about 109 times, even though the concentration of BNAH+• (4.95 × 10-5 mM) is much less than that of BNAH (2.5 mM) in the reaction system. Evidently, the much lower BNAH+• concentration than that of BNAH in the reaction system is not a decisive factor to determine the mechanism of the NADH model oxidations by Fc+ in non-alkaline media. A new question will be produced naturally why the intrinsic kinetic barrier of the hydrogen atom transfer is much greater than that of the proton transfer?

In order to reply this question, the

factors that affect the intrinsic kinetic barrier of chemical reactions need to be clarified first.

Since

the intrinsic kinetic barrier of a chemical reaction is equal to the average value of activation free energies of two corresponding self-exchange reactions, G≠(XL/X) and G≠(YL/Y) (eq 21), it is 16


Page 17 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


clear that the factor that affects the activation free energy of self-exchange reactions is the factor that affects the intrinsic kinetic barrier of the reaction. From the new kinetic model (Figure 9), it is clear that for a self-exchange reaction (XL + X → X + XL), the activation free energy of the reaction, G(XL/X), is mainly determined by two following factors: Go(XL) and the distance between XL and X (r).20

From

Figure 11, it is clear that when Go(XL) increases, the thermo-kinetic parameter of XL, Go(XL), increases, and at the same time, the thermo-kinetic parameter of X, Go(X), increases as well, which naturally makes the activation free energy of the self-change reaction, G(XL/X), increases little. G(XL/X) is quite small.

Thus, it is clear that the effect of Go(XL) on

From Figure 12, it is clear that when r increase, the

thermo-kinetic parameter of XL, Go(XL), increase, but at the same time, the thermo-kinetic parameter of X, Go(X), decrease, which naturally makes the activation free energy of the self-change reaction, G(XL/X), increase greatly.

Thus, the effect of r on

G(XL/X) is quite large and the larger r is, the greater G(XL/X) is.

Since r is mainly

determined by the composition, structure and charge of the reactants, it is clear that the composition, structure and charge of reactants should be the three most important factors that affect the activation free energy of the self-change reaction, G(XL/X), i.e., the intrinsic kinetic barrier of the chemical reactions.

Figure 11. Effect of Go(XL) of XL on the activation free energies of the self-exchange reaction, G(XL/X).

17



Page 18 of 26

Figure 12. Effect of distance between the two reactants in TS (r) on the activation free energy of the self-exchange reaction, G(XL/X).

From the proton transfer reaction (eq 11) and the hydrogen atom transfer reaction (eq 13), it is clear that the composition and structure of the corresponding reactants are the same, and the only difference is the charge.

In detail, for the proton transfer reaction, one

reactant (BNAH+•) carries one positive charge and the other reactant (BNAH) is uncharged, but for the hydrogen atom transfer reaction, the two reactants (BNAH+• and BNAH+•) all carry a positive charge.

Due to electrostatic repulsion between the same charges, it is

plausible that the distant between the two reactants in TS for the hydrogen atom transfer reaction should be much longer than that for the proton transfer reaction, which directly makes the hydrogen transfer reaction have much greater intrinsic kinetic barrier (Figure 12 and Scheme 5).

Obviously, it is the positive charge of BNAH+• that makes the NADH

model oxidations by Fc+ in acetonitrile go through the three-step rather than two-step mechanism. Scheme 5. Effect of Charge of Reactants on the Distance (r) between the Two Reactants in Transition State (TS)

18


Page 19 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

▉


CONCLUSIONS

In this work, the detailed mechanism of NADH model oxidations by Fc+ in dry acetonitrile were examined, the results suggest that the NADH model oxidations by metal ion (Fc+) in non-alkaline media is the three-step rather than two-step mechanism. that the radical cations of NADH models carry the positive charge.

The main reason is

It is the positive charge

of the radical cations that prevents them from approaching in the reaction.

Based on the

work in this paper, we can make an important conclusion: regardless of the basicity of reaction media the oxidations of NADH and its models by metal ions always take place by the three-step mechanism.

The difference lies in that if the basicity of the reaction media is greater than the

basicity of NADH or its models, the proton-acceptor in the reaction is the media (shown in Scheme 1); otherwise, the proton-acceptor is NADH or its models (shown in Scheme 3).

▉

EXPERIMENTAL SECTION Materials.

Solvents and reagents were obtained from commercial sources and used as

received unless otherwise noted.

Reagent grade acetonitrile was refluxed over KMnO4 and K2CO3

for at least eight hours and doubly distilled over P2O5 under argon and stored in the glove-box before use.

The BNAH and PNAH were synthesized according to literature methods.41

The

dideuterated compounds BNAD and PNAD were prepared from the corresponding monodeuterated compounds by three cycles of oxidations with p-chloranil in dimethylformamide and reduction with dithionite in deuterium oxide.42,43

The ferrocenium hexafluorophosphates (Fc+PF6-) was

synthesized according to published procedures.23,44 Stoichiometry Measurements.

The stoichiometry in acetonitrile was measured through

reaction at various molar ratios of BNAH/Fc+ by monitoring the changes of the absorption decay at 617 nm due to Fc+ on a HITACHI UV-3000 spectrometer.

A typical process was described. 5.02

mM BNAH and 5.01 mM Fc+ solutions were prepared and mixed in an Ar-filled glovebox.

The

constant Fc+ (1.5 mL) was added to a three-neck UV cell which can be sealed from the air and vapor. Then aliquots (0.3, 0.6, 0.9, 1.2 mL et al) of NADH solution with appropriate volume of acetonitrile to make sure a 3 mL total volume, were gradually syringed into it until the 617 nm absorption died away.

The measurements in pyridine-buffered solution were the same as

described above, except that the solvent was 1:10 pyridine/acetonitrile in volume. 19



Page 20 of 26

Kinetic Measurements. (1) General Process:

The kinetic runs were performed on an Applied Photophysics

SX.18MV-R stopped-flow spectrophotometer at 298 K in presence and absence of pyridine in acetonitrile.

The SX.18MV-R, connected to a superthermostat circulating bath to regulate the

temperature of cell compartments, has a dead time of about 1 ms with very high sensitivity. solutions used for kinetics were prepared in an Ar-filled glovebox.

All

These measurements of the

reactions were under pseudo-first-order conditions with the concentrations of excess NADH models at least 10 times larger than Fc+.

The progress of the reaction was monitored by following the

decrease of the ferrocenium absorption at 617 nm in the visible region.

All runs were repeated

more than five times to ensure the reliability of the data. (2) In Base Solution: The kinetic runs in base solution were performed by adding enough pyridine (pyridine: acetonitrile = 1:10 in volume) to act as the base in the deprotonation step. typical process is as follows: 0.2 oC with a water bath.

A

Prior to mixing, the reactant solutions were thermostated to 25.0 ±

2.77 mM BNAH in pyridine-acetonitrile solution was mixed with equal

volume of 0.25 mM Fc+ in the Stopped-flow syringe.

For the data collection, the kinetic traces

were recorded on an Acorn computer and analyzed by Pro-K Global analysis/simulation software or translated to PC for further analysis by Origin Software through a single-exponential kinetic equation to calculate the pseudo-first-order rate constants and then converted to second-order rate constants by linear correlation of the pseudo-first-order rate constants against the concentrations of the excessive reactants.

The reactions were all followed for three or more half-lives and plots of

ln(A0 - A∞)/(At - A∞) vs t gave r > 0.999. (3) In Neutral Solution: The measurements in neutral solution were performed under the condition of [NADH] >> [Fc+] < [Fc] in deaerated acetonitrile at 298 K including a constant amount of the product Fc (10 times more than Fc+, 2.5 mM) to ensure the steady-state approximation.

In each run, the NADH models varies in five different concentrations, typically

from 2.5 mM to 10 mM.

The decay profile of absorption at 617 nm due to Fc+ and the

corresponding fitted first-order plots were shown in Figure 3. The reaction timescales were depended on the activity of compounds.

Various concentration

of NADH models were used to derive the dependence of the pseudo-first-order rate constants on the concentrations of excess NADH models. The pseudo-first-order rate constants were calculated by 20


Page 21 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


a single-exponential kinetic equation and then converted to obtain the slopes and intercepts of [NADH]/kobs vs 1/[NADH] plots.

AUTHOR INFORMATION Corresponding Author *E-mail: [email protected] ORCID Xiao-Qing Zhu: 0000-0003-2785-640X Notes The authors declare no competing financial interest.

ACKNOWLEDGEMENTS Financial support from the National Natural Science Foundation of China (Grant Nos. 21672111, 21602116, 21472099, 21390400 and 21102074) and the 111 Project (B06005) is gratefully acknowledged.

In addition, we would especially like to thank Prof. Lu Yun of Southern Illinois

University at Edwardsville for thoroughly modifying the language of the article and giving a high evaluation of the scientific value of the article!

▉ (1)

REFERENCES Walker, J. E. The NADH: Ubiquinone Oxidoreductase (complex I) of Respiratory Chains. Quarterly reviews of biophysics, 1992, 25, 253-324.

(2)

Ying, W. NAD+/NADH and NADP+/NADPH in Cellular Functions and Cell Death: Regulation and Biological Consequences. Antioxidants & redox signaling, 2008, 10, 179-206.

(3)

Weiss, H.; Friedrich, T.; Hofhaus, G.; Preis, D. EJB Reviews 1991. Springer, Berlin, Heidelberg, 1991: 55-68.

(4)

Sazanov, L. A.; Hinchliffe, P. Structure of the Hydrophilic Domain of Respiratory Complex I from Thermus Thermophiles. Science, 2006, 311, 1430-1436.

(5)

Yagi, T.; Matsuno-Yagi, A. The Proton-translocating NADH−quinone Oxidoreductase in the Respiratory Chain: the Secret Unlocked. Biochemistry 2003, 42, 2266-2274. 21



(6)

Page 22 of 26

Ohnishi, T. Iron–sulfur Clusters/semiquinones in Complex I. Biochimica Biophysica Acta (BBA)-Bioenergetics, 1998, 1364, 186-206.

(7)

Wang, S.; Cazelles, R.; Liao, W.-C.; Vázquez-González, M.; Zoabi, A.; Abu-Reziq, R.; Willner, I. Mimicking Horseradish Peroxidase and NADH Peroxidase by Heterogeneous Cu2+-Modified Graphene Oxide Nanoparticles Nano Lett. 2017, 17, 2043−2048.

(8)

McSkimming, A.; Colbran, S. B. The Coordination Chemistry of Organo-hydride Donors: New Prospects for Efficient Multi-electron Reduction. Chem. Soc. Rev., 2013, 42, 5439-5488.

(9)

Binstead, R. A.; McGuire, M. E.; Dovletoglou, A.; Seok, W. K.; Roecker, L. E.; Meyer, T. J. Oxidation of Hydroquinones by Ruthenium Complexes [(bpy)2(py)RuIV(O)]2+ and [(bpy)2(py)RuIII(OH)]2+. Proton-coupled Electron Transfer. J. Am. Chem. Soc. 1992, 114, 173-186.

(10)

Fukuzumi, S.; Fujioka, N.; Kotani, H.; Ohkubo K.; Lee, Y.-M.; Nam, W. Mechanistic Insights into Hydride-Transfer and Electron-Transfer Reactions by a Manganese (IV)−Oxo Porphyrin Complex. J. Am. Chem. Soc. 2009, 131, 17127-17134.

(11)

Oppelt, K. T.; Gasiorowski, J.; Egbe, D. A. M.; Kollender, J. P.; Himmelsbach, M.; Hassel, A.

W.;

Sariciftci,

N.

S.;

Knör,

G.

Rhodium-Coordinated

Poly(arylene-ethynylene)-alt-Poly(arylene-vinylene) Copolymer Acting as Photocatalyst for Visible-Light-Powered NAD+/NADH Reduction. J. Am. Chem. Soc. 2014, 136, 12721−12729. (12)

Zhang, D.; Wu, L. Z.; Zhou, L.; Yang, Q. Z.; Zhang, L. P.; Tung, C. H. Photocatalytic Hydrogen Production from Hantzsch 1,4-dihydropyridines by Platinum(II) Terpyridyl Complexes in Homogeneous Solution. J. Am. Chem. Soc. 2004, 126, 3440-3441.

(13)

Schwarz, D. E.; Cameron, T. M.; Hay, P. J.; Scott, B. L.; Tumas, W.; Thorn, D. L. Hydrogen Evolution from Organic “Hydrides”. Chem. Commun. 2005, 47, 5919-5921.

(14)

Morkovnik, A. S.; Ivakhnenko, E. P.; Bogachev, Y. G.; Tertov, B. A.; Berberova, N. T.; Okhlobystin, O. Y. Hydrogen Evolution at Interaction of Certain Hydroheteroaromatics with Dehydrogenating agents. Khim. Geterotsikl. Soedin. 1988, 2, 203-208.

(15)

Chenault, H. K.; Whitesides, G. M. Regeneration of Nicotinamide Cofactors for Use in Organic Synthesis. Appl. Biochem. Biotechnol. 1987, 14, 147-197. 22


Page 23 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

(16)


Chen, Q.-A.; Chen, M.-W.; Yu, C.-B.; Shi, L.; Wang, D.-S.; Yang, Y.; Zhou, Y.-G. Biomimetic Asymmetric Hydrogenation: in situ Regenerable Hantzsch Esters for Asymmetric Hydrogenation of Benzoxazinones. J. Am. Chem. Soc. 2011, 133, 16432-16435.

(17)

Lo, H. C.; Fish, R. H. Biomimetic NAD+ Models for Tandem Cofactor Regeneration, Horse Liver Alcohol Dehydrogenase Recognition of 1, 4-NADH Derivatives, and Chiral Synthesis. Angew. Chem. Int. Ed, 2002, 41, 478-481.

(18)

Htet, Y.; Tennyson, A. G. NAD+ as a Hydride Donor and Reductant. J. Am. Chem. Soc. 2016, 138, 15833−15836.

(19)

Padhi, S. K.; Fukuda, R.; Ehara, M.; Tanaka, K. Comparative Study of C^N and N^C Type Cyclometalated Ruthenium Complexes with a NAD+/NADH Function. Inorg. Chem. 2012, 51, 8091-8102.

(20)

Nakamura. K.; Fuji, M.; Ohno, A.; Oka, S. NAD(P)+-NAD(P)H Model. 52. Reduction of Olefins by Hantzsch Ester on Silica Gel. Tetrahedron Lett. 1984, 25, 3983-3986.

(21)

Taniuchi, H.; Fujibayashi, Y.; Okazawa, H.; Yonekura, Y.; Konishi, J.; Yokoyama, A. Cu-Pyruvaldehyde-bis(N4-methylthiosemicarbazone) (Cu-PTSM), a Metal Complex with Selective NADH-Dependent Reduction by Complex I in Brain Mitochondria : A Potential Radiopharmaceutical for Mitochondria-Functional Imaging with Positron Emission Tomography (PET). Biol. Pharm. Bull. 1995, 18, 1126-1129.

(22)

Carlson, B. W.; Miller, L. L. Oxidation of NADH by Ferrocenium Salts. Rate-limiting One-electron Transfer. J. Am. Chem. Soc. 1983, 105, 7453-7454.

(23)

Carlson, B. W.; Miller, L. L.; Neta, P.; Grodkowski, J. Oxidation of NADH Involving Rate-limiting One-electron Transfer. J. Am. Chem. Soc. 1984, 106, 7233-7239.

(24)

Miller, L. L.; Valentine, J. R. On the Electron-proton-electron Mechanism for 1-benzyl-1, 4-dihydronicotinamide Oxidations. J. Am. Chem. Soc. 1988, 110, 3982-3989.

(25)

Powell, M. F.; Wu, J. C.; Bruice, T. C. Ferricyanide Oxidation of Dihydropyridines and Analogs. J. Am. Chem. Soc. 1984, 106, 3850-3856.

(26)

Shunichi, F.; Yuji, K.; Toshio, T. Electron Transfer Reactions of an NADH Model with Iron(III)complexes. A two-step Electron Transfer Mechanism. Chem. Lett. 1982, 11, 1591-1594. 23



(27)

Page 24 of 26

Fukuzumi, S.; Tokuda, Y.; Kitano, T.; Okamoto, T.; Otera, J. Electron-transfer Oxidation of 9-substituted 10-methyl-9, 10-dihydroacridines. Cleavage of the Carbon-hydrogen vs. Carbon-carbon Bond of the Radical Cations. J. Am. Chem. Soc. 1993, 115, 8960-8968.

(28)

Anne, A.; Hapiot, P.; Moiroux, J.; Neta, P.; Saveant, J.-M. Dynamics of Proton Transfer from Cation Radicals. Kinetic and Thermodynamic Acidities of Cation Radicals of NADH Analogs. J. Am. Chem. Soc. 1992, 114, 4694-4701.

(29)

For evaluating the standard one-electron redox potentials of analyte with irreversible electrochemical processes, OSWV has been verified to be a more exact electrochemical method than CV (see Zhu, X.-Q.; Zhang, M.-T.; Yu, A.; Wang, C.-H. Cheng, J.-P. J. Am. Chem. Soc. 2008, 130, 2501-2516).

(30)

The concentration of BNAH+• in the reaction system (4.95 × 10-5 mM) was estimated by using eq (6) according to the initial concentrations of BNAH (2.5 mM), Fc (2.5 mM) and Fc+ (0.25 mM) and the related data in Table 3.

(31)

5.1 kcal/mol is derived from the oxidation potential of BNAH (0.22 V vs Fc+) by using G = 23.06 × Eox(BNAH).

(32)

-4.5 kcal/mol is derived from the pKa value of BNAH+• (4.7) reduced by the pKa value of BNAH2+ (8) and then multiplied by -1.364.

(33) The thermodynamic driving force of the second step hydrogen atom transfer (-42.3 kcal/mol) in the two-step mechanism (Scheme 2) is derived from the free energy change of the net reaction of NADH models with Fc+ (-32.1 kcal/mol) in Scheme 3 and 2 fold of the molar free energy change of the first step electron transfer reaction (10.2 kcal/mol) according to Hess law. (34)

Zhu, X.-Q.; Deng, F.-H.; Yang, J.-D.; Li, X.-T.; Chen, Q.; Lei, N.-P.; Meng, F.-K.; Zhao, X.-P.; Han, S.-H.; Hao, E.-J. et al. A Classical but New Kinetic Equation for Hydride Transfer Reactions. Org. Biomol. Chem. 2013, 11, 6071-6089.

(35)

According to the curvature of the Morse curves, the sum of Go(XL) and Go(Y) should be slightly less than the sum of GY(XL) and GXL(Y), but the difference has been extensively verified to be less than the uncertainties of the experimental measurements of G(XL/Y).34,36,38 24


Page 25 of 26 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

(36)


Li, Y.; Zhu, X.-Q. Theoretical Prediction of Activation Free Energies of Various Hydride Self-Exchange Reactions in Acetonitrile at 298 K. ACS Omega, 2018, 3, 872-885.

(37)

Fu, Y.-H.; Shen, G.-B.; Li, Y.; Yuan, L.; Li, J.-L.; Li, L.; Fu, A.-K.; Chen, J.-T.; Chen, B.-L.; Zhu, L. et al. Realization of Quantitative Estimation for Reaction Rate Constants Using only One Physical Parameter for Each Reactant. ChemistrySelect 2017, 2, 904-925.

(38)

Shen, G.-B.; Xia, K.; Li, X.-T.; Li, J.-L.; Fu, Y.-H.; Yuan, L.; Zhu, X.-Q. Prediction of Kinetic Isotope Effects for Various Hydride Transfer Reactions Using a New Kinetic Model. J. Phys. Chem. A 2016, 120, 1779-1799.

(39)

Eq 22 (also known as Zhu equation36,37,38) is different from Marcus equation (G = Go + ½Go + Go2/16Go).

The main difference is that the G≠o(XL/Y) in eq 22 is a variable

when Go(XL/Y) (chemical reaction) changes, but the G≠o in Marcus equation is a constant when Go (chemical reaction) changes. From Marcus’ two initial papers that yield Marcus equation (Marcus, R. A. J. Chem. Phys. 1956, 24, 966-978 and 979-989), it is clear that the prerequisite of Marcus equation is that Go is a constant when Go (chemical reaction) changes. obtained. region.

If  were not a constant, Marcus equation could not be

In addition, if  were not a constant, there would not be the Marcus inverted However, so far, some chemists, such as H. Mayr et al. still mistakenly use the

Go in the Marcus equation as a variable to estimate the intrinsic kinetic barriers of chemical reactions (see H. Mayr, et al. J. Am. Chem. Soc. 2017, 139, 1499-1511; Pure Appl. Chem. 2017, 89(6), 729-744; Acc. Chem. Res. 2016, 49, 952-965). The main reason is that they lack a correct understanding of Marcus theory and neglect the prerequisite of Marcus equation (see reference 38). (40)

Derived from the rate constant (k1 = 3.19 × 103 M-1s-1) of the first step electron transfer reaction by using Eyring equation.

(41)

Zhu, X.-Q.; Tan, Y.; Cao, C.-T. Thermodynamic Diagnosis of the Properties and Mechanism of Dihydropyridine-type Compounds as Hydride Source in Acetonitrile with “Molecule ID Card”. J. Phys. Chem. B. 2010, 114, 2058-2075.

(42)

Fukuzumi, S.; Nishizawa, N.; Tanaka, T. Mechanism of Hydride Transfer from an NADH Model Compound to p-benzoquinone Derivatives. J. Org. Chem. 1984, 49, 3571-3578. 25



(43)

Page 26 of 26

Caughey, W. S.; Schellenberg, K. A. Characterization of an Intermediate in the Dithionite Reduction of a Diphosphopyridine Nucleotide Model as a 1, 4-Addition Product by Nuclear Magnetic Resonance Spectroscopy. J. Org. Chem. 1966, 31, 1978-1982.

(44)

Nesmeyanov, A. N.; Materikova, R. B.; Lyatifov, I. R.; Kurbanov, T. K.; Kochetkova, N. S. Sym-polymethylferricinium Hexafluorophosphates. J. Organomet. Chem. 1978, 145, 241-243.

TOC Graphic：

26


A New Insight into the Mechanism of NADH Model Oxidation by Metal

Recommend Documents