The effect of nitro substitution on the photochemistry of benzyl benozhydroxamate: Photoinduced release of benzohydroxamic acid

Article abstract of DOI:10.1016/j.jphotochem.2011.10.005

The redox congener of the important signaling agent nitric oxide (NO), nitroxyl or nitrosyl hydride (HNO) has also been demonstrated to induce distinct physiological effects. The aim of this study was to determine if benzohydroxamic acid, which was selected as a stable model compound of HNO donors, could be released by the o-nitrobenzyl photolabile protecting group (PPG) in a wavelength-dependent manner. It was expected that selective irradiation of the o-nitrobenzyl chromophore would favor the release of benzohydroxamic acid over undesired products associated with N-O bond cleavage. Quantum yields for the release of benzohydroxamic acid protected by the o-nitrobenzyl PPG increased at longer wavelengths, with a concomitant decrease in the yield of minor products. Through the use of triplet photosensitizers, triplet quenchers, computational methods, and the position of the nitro substituent, insights into the nature of the mechanism were suggested.

Full text of DOI:10.1016/j.jphotochem.2011.10.005

Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
Contents lists available at SciVerse ScienceDirect
Journal of Photochemistry and Photobiology A:
Chemistry
journal homepage: www.elsevier.com/locate/jphotochem
The effect of nitro substitution on the photochemistry of benzyl
benozhydroxamate: Photoinduced release of benzohydroxamic acid
Whitney R. Grither, James Korang, Jacob P. Sauer, Matthew P. Sherman, Pamela L. Vanegas,
Miao Zhang, Ryan D. McCulla∗
Saint Louis University, Department of Chemistry, Monsanto Hall, 3501 Laclede Ave., St. Louis, MO 63103, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
The redox congener of the important signaling agent nitric oxide (NO), nitroxyl or nitrosyl hydride (HNO)
has also been demonstrated to induce distinct physiological effects. The aim of this study was to determine
if benzohydroxamic acid, which was selected as a stable model compound of HNO donors, could be
released by the o-nitrobenzyl photolabile protecting group (PPG) in a wavelength-dependent manner.
It was expected that selective irradiation of the o-nitrobenzyl chromophore would favor the release of
benzohydroxamic acid over undesired products associated with N–O bond cleavage. Quantum yields for
the release of benzohydroxamic acid protected by the o-nitrobenzyl PPG increased at longer wavelengths,
with a concomitant decrease in the yield of minor products. Through the use of triplet photosensitizers,
triplet quenchers, computational methods, and the position of the nitro substituent, insights into the
nature of the mechanism were suggested.
Received 10 May 2011
Received in revised form
28 September 2011
Accepted 3 October 2011
Available online 18 October 2011
Keywords:
Hydroxamate
Nitroxyl
Nitrosyl hydride
Photolabile protecting group
Nitrobenzyl
© 2011 Elsevier B.V. All rights reserved.
Wavelength dependent
1. Introduction
spatial and temporal control over the generation of HNO, several
investigators have examined methods of photochemically gener-
The redox congener of the important signaling agent nitric
oxide (NO), nitroxyl or nitrosyl hydride (HNO), has also been
demonstrated to induce distinct physiological effects [1–5]. The
therapeutic utilization of HNO has been used to inhibit alde-
hyde dehydrogenases (ALDH) in the treatment of alcoholism [6,7].
Recent efforts have focused on application of HNO in cardiovas-
cular conditions, such as the treatment of congestive heart failure,
atherosclerosis, and vascular thrombosis [8–13]. Additionally, HNO
donors afforded neuroprotection from excitotoxic assault by down-
regulating N-methyl-d-aspartate (NMDA) receptors [14]. Biological
targets of HNO have been identified as thiols, transition metals,
iron–sulfur clusters, and DNA [15–18].
The rapid dehydrative dimerization of HNO to nitrous oxide
and water necessitates in situ generation to investigate the chem-
istry and underlying biochemistry of HNO [19,20]. As shown in
Scheme 1, the exogenous generation of HNO can be achieved
through the decomposition of HNO donors such as Angeli’s salt
(NaN2O3) and Piloty’s acid (PhSO2NOH) [21]. However, release of
HNO from these donors depends upon the spontaneous degrada-
tion of the donors, which is not a controlled event. To provide better
ating HNO in solution. Acyl nitroso compounds undergo hydrolysis
to release HNO, and adducts between 9,10-dimethylanthracene
and acyl nitroso derivatives have been photoinduced to undergo
a retro-Diels–Alder reaction that releases the acyl nitroso HNO
donor [22]. The photo-induced fragmentation of 3,5-diphenyl-
1,2,4-oxadiazole-4-oxide produces an acyl nitroso intermediate
whose decomposition also releases HNO [23]. However, these
methods either require aqueous insoluble reagents or incident
wavelengths that are absorbed by biomolecules, which often leads
to photodamage and phototoxicity. Photolabile protecting groups
(PPG) afford researchers exquisite spatial, temporal, and concentra-
tion control of the release of bioactive agents at wavelengths that
induce little photodamage [24,25]. A seemingly straightforward
strategy to generate HNO would be to protect a HNO donor with a
PPG. Upon photolysis, the caged HNO donor would be liberated. In
addition, this could lead to more rapid generation of HNO due to
residual vibrational energy carried over from the initial excitation.
A
potential complicating factor facing the release of an
organic based HNO donor caged by PPG is that they
a
contain bonds that are potentially photosensitive. For
example, N,O-diacylhydroxylamines, hydroxamic acids, and
N-alkylbenzohydroxamates have all been shown to undergo N–O
bond homolysis upon irradiation [26–31]. Alkyl benzohydroxa-
mates undergo triplet state Norrish Type II photoelimination to
∗
Corresponding author. Tel.: +1 314 977 2850.
E-mail address: rmccull2@slu.edu (R.D. McCulla).
1010-6030/$ – see front matter © 2011 Elsevier B.V. All rights reserved.
doi:10.1016/j.jphotochem.2011.10.005

2
W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
energy conformation was determined from a number of rationally
selected conformers that were optimized with HF/6-31G(d). These
geometries were then further refined at the B3LYP/6-31G(d) level of
theory, which were confirmed as minima by vibrational frequency
calculations. Vertical excitation energies and oscillator strengths
for singlet excited states were calculated using time-dependent
DFT methods [39]. Structures and the difference between the
ground and excited state density matrices were visualized using
ChemCraft 1.6 [40].
Scheme 1.
2.2. Materials
produce benzamide and the corresponding carbonyl compound
[31,32]. The photolysis of arenesulfonylhydroxamates leads to a
number of products that are consistent with S–N bond homolysis
[33]. However, most of these processes required long irradiation
times. Therefore, a PPG that is released efficiently and whose
excitation does not lead to N–O bond homolysis should be capable
of releasing HNO donors.
The aim of this study was to determine if mimics of HNO
donors could be released by the o-nitrobenzyl (2NBn) PPG [24].
Benzohydroxamic acid (1) was selected to serve as model of
sulfonylhydroxamate HNO donors because 1 was expected to
be stable after release by 2NBn. Since all HNO donors degrade
to release HNO, a stable model (i.e. 1) was used instead of
attempting to directly release HNO donors, thus simplifying quan-
tification of the products for this initial study. The 2NBn PPG was
selected since 2NBn has been thoroughly investigated [34]. Also,
the 2NBn chromophore was expected to absorb light at higher
wavelengths, which was expected to limit N–O bond homolysis
reactions [35–37]. To examine this hypothesis, the photochem-
istry of nitro-substituted derivatives of benzyl benzohydroxamate
(Bn-1) shown in Scheme 2 (i.e. 4-nitrobenzyl benzohydroxamate
(4NBn-1), 2-nitrobenzyl benzohydroxamate (2NBn-1), and benzyl
4-nitrobenzohydroxamate (Bn-4N-1)) was examined.
Commercial materials were obtained from Sigma–Aldrich or
Fisher Scientific and used as provided unless specified otherwise.
HPLC grade acetonitrile was dried by distilling over calcium hydride
under inert atmosphere following procedure found in Armarego
and Perrin [41]. Water was purified by a Milli-Q system. Benzyl
benzohydroxamate (Bn-1) and 4-nitrobenzyl benzohydroxamate
(4NBn-1) were prepared by the literature procedures [42,43].
2.2.1. Benzyl 4-nitrobenzohydroxamate (Bn-4N-1)
4-Nitrobenzoyl chloride (0.68 g, 4.10 mmol) and O-
benzylhydroxylamine hydrochloride (0.65 g, 4.07 mmol) were
dissolved in 60 mL of dry CH2Cl2 under nitrogen atmosphere.
The reaction was allowed to stir for 15 min, and then 0.70 mL
(8.65 mmol) of pyridine was added at 0 ◦C. The reaction mixture
was allowed to proceed for 2 h at room temperature before being
diluted with CH2Cl2 and washed three times with water. The
organic layer was dried with anhydrous Na2SO4 and concentrated
under reduced pressure. The resulting solid was recrystallized
in toluene to give a colorless solid (0.49 g, 44.2%) with the same
spectral profile as reported in the literature for the desired product
[44]. Lit. m.p. = 166–167 ◦C, obs. m.p. = 167–168 ◦C.
2.2.2. 2-Nitrobenzyl benzohydroxamate (2NBn-1)
2. Materials and methods
The desired compound was prepared following a modified
procedure [45]. Benzohydroxamic acid (1.4 g, 10.0 mmol) was dis-
solved in 20 mL EtOH followed by the addition of 0.84 g (15.0 mmol)
KOH dissolved in the minimum amount of water. In 50 mL of
EtOH, 2-nitrobenzyl bromide (1.8 g, 8.3 mmol) was added drop-
wise. The solution was heated and allowed to reflux for 4.5 h and
all of the 2-nitrobenzyl bromide had reacted. The mixture was
then concentrated under reduced pressure and reconstituted with
80 mL of water. The product was extracted with chloroform (3×
60 mL), and the combined organic phases were washed with water
(60 mL) and dried over anhydrous magnesium sulfate. The solvent
was removed under reduced pressure, and the solid residue was
then purified by column chromatography Rf = 0.39 (CH2Cl2/EtOAc
9:1) to yield a pale yellow solid. This solid was recrystallized in
EtOH to yield 0.65 g (28.7%) of a colorless solid. Lit. m.p. = 121 [46]
obs. m.p. = 119–120 ◦C; 1H NMR (CDCl3) ı = 8.76 (s br, 1H), 8.09 (d,
J = 8.2 Hz, 1H), 7.95 (d, J = 7.6 Hz, 1H), 7.71 (m, 3H), 7.53 (m, 2H), 7.46
(m, 2H), 5.47 (s, 2H); IR (ATR) ꢀ˜ (cm−1) = 3190, 3050, 2950, 1636,
1521, 1336.
2.1. Computational methods
All calculations were performed with the Gaussian 09 suite
of programs for electronic structure calculations [38]. The lowest
2.3. Irradiations
Quantum yield measurements were carried out with a 75 W
Xe arc lamp focused on a monochromator for wavelength selec-
tion. Slit widths allowed 6 nm of linear dispersion from the set
wavelength. Samples (4.8 mL) in a 1 cm quartz cell were placed
in a permanently mounted cell holder such that all of the exiting
light hit the sample without further focusing. For all direct quan-
tum yield measurements, the concentration of the starting material
was confirmed as sufficient to obtain a minimum absorbance of
2 at the selected wavelength, and the samples were irradiated
Scheme 2.

W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
3
until approximately 15% conversion of the starting material was
reached. Sensitized photoreactions were carried out at sufficient
excess concentration of the photosensitizers to ensure at least 99%
of the incident irradiation was absorbed by the sensitizer. Quench-
ing experiments used 20 to 100-fold molar excess of the triplet
quenchers. Prior to photolysis, most samples were sparged with
argon for at least 15 min to remove oxygen. Other samples were car-
ried out under freeze–pump–thaw conditions to rigorously exclude
oxygen. Photolysis of azoxybenzene was used as an actinometer
[47].
Due to the slow rates of conversion, percent yields of the
observed products were determined from photolysis reactions car-
ried out at 254 nm with a Luzchem LZC-4 photoreactor using
low-pressure Hg lamps. Samples were held in standard fused-silica
test tubes, and the starting concentrations (0.20 mM) were low-
ered to ease quantification of the loss of starting material. To ease
quantification of the products, photoreactions were carried out to
30% conversion of the starting material. Thus, all percent yields
are reported in respect to the theoretical maximum that could be
obtained after 30% consumption of the starting material.
electron density accumulation in the excited state, and light gray
corresponds to depletion from the ground state. For Bn-1, the char-
acter of the first excited state corresponds to a n, ␲* state centered
on the hydroxamate. Inspection of the difference density plot of
2NBn-1 and 4NBn-1 shows that the S1 state is characterized by a (n,
NO2) → (␲*, NO2) transition. The difference density of the S1 state of
Bn-4N-1 reveals that electron density is transferred from the ben-
zyl moiety to the nitro functional group, which suggests the S1 of
Bn-4N-1 resembles a donor-bridge-acceptor intramolecular charge
transfer (CT) state [53,54]. Visualization of the S3 state of 4NBn-1
revealed transfer of electron density from the benzohydroxamate
to the nitro group, which was similar to the donor-bridge-acceptor
CT state observed for Bn-4N-1.
Potentially, release of 1 from 2NBn-1 would be decreased if
pathways from higher energy excited states led to N–O homoly-
sis. While internal conversion from higher singlet excited states
(i.e. S2–S5) to S1 is generally faster than other processes, poor over-
lap between the electronic or vibronic wave functions of two states
results in slow internal conversion [55]. For 2NBn-1, the S2, S3, and
S5 states were characterized by depletion of electron density from
the benzohydroxamate moiety and an increase on the ␲* orbital of
the nitro group. Since the orbitals of the benzohydroxamate and the
nitro groups were not conjugated, excitation into any of these states
had the potential to allow processes not associated with S1, which
was expected to increase the possibility N–O bond homolysis.
Evidence indicating that irradiation of 2NBn-1 at longer wave-
lengths would lead to excitation of the 2NBn chromophore was
obtained from the UV/Vis spectra of 1, Bn-1, 2NBn-1, 4NBn-1,
and Bn-4N-1 and 2-nitrotoluene. As shown in Fig. 2a, the molar
extinction coefficient (ε) for 1 and 2-nitrotoluene dropped below
100 M−1 cm−1 at 300 nm and 368 nm, respectively. This indicated
that 2-nitrotoluene had a lower energy S1 state than 1, and there-
fore, the S1 state of 2NBn-1 was expected to be predominantly
associated with the 2NBn moiety. The absorption spectra for 2NBn-
1 resembled a composite of the spectra obtained for 2NBn and
1, which suggested that the two observed absorption bands arose
from localized chromophores.
2.4. General methods
The concentrations of the starting materials and products at sev-
eral time points during the photoreaction were quantified by HPLC
analysis carried out with an Agilent 1100 Series instrument fitted
with a diode array detector using a 150 mm/4.6 ␮m Eclipse XDB-
C18 column. Products were identified by comparison to authentic
samples and GC/MS analysis using a Shimadzu GCMS QP2010S
using a DB-5 column for separations. The UV/Vis absorption spec-
tra were obtained using a Shimadzu PharmaSpec UV-1700 system,
and infrared spectra were obtained using a Perkin-Elmer Paragon
FT-IR. NMR spectra were obtained using a Bruker DRX-400.
3. Results
A number of previous studies have shown the 2NBn PPG to
undergo an intramolecular hydrogen abstraction from the benzyl
position generating an aci-nitro intermediate whose subsequent
thermal chemistry releases the “caged” molecule [34,48–52]. For-
mation of the aci-nitro intermediate has been observed to form
in less than 10 ps by picosecond pump–probe spectroscopy. For
the photolysis of 2NBn-1, the release of 1 was expected to domi-
nate if the first excited singlet state (S1) of 2NBn-1 favors hydrogen
abstraction by the nitro group over N–O bond homolysis. For the
parent Bn-1, photolysis led to predominantly N–O bond cleavage
through a triplet Norrish Type II mechanism [31]. To better under-
stand the excited states involved, the nature of the excited states of
Bn-1, 2NBn-1, 4NBn-1, and Bn-4N-1 were investigated by absorp-
tion spectroscopy and time-dependent density functional theory
(TD-DFT) calculations.
For Bn-4N-1 and 4NBn-1, TD-DFT calculations found transitions
with charge-transfer character had the largest oscillator strengths.
To determine if the broad absorption bands centered near 265 nm
for Bn-4N-1 and 4NBn-1 arose from excitation into these CT states,
the absorption spectra of Bn-4N-1 and 4NBn-1 were obtained in
chloroform, isopropanol, tetrahydrofuran, and acetonitrile. If the
polarity of the ground state and excited state for the dominant
absorption band were different, a positive solvatochromic shift
would be expected. However, only a small bathochromic shift was
observed for both compounds (Bn-4N-1; ꢁmax CHCl3 (262.9 nm)
ACN (264.1 nm), 4NBn-1; ꢁmax CHCl3 (266.0 nm) ACN (267.0 nm)).
This small positive solvatochromic shift indicated that for both Bn-
4N-1 and 4NBn-1 the polarity of the ground state and excited state
were similar.
The singlet ground (S0) and triplet state (T1) geometries of Bn-1,
2NBn-1, 4NBn-1, and Bn-4N-1 were optimized at the B3LYP/6-
31G(d) level of theory. Zero point energies were included when
determining the energy of T1 relative to S0 at their equilibrium
geometries. Vertical excitation energies and oscillator strengths for
singlet excited states were calculated using time-dependent DFT
methods [39], TD-B3LYP/6-31G(d). The vertical excitation energies
(singlets), oscillator strengths, and T1 energies are listed in Table 1.
The nature of the excited states was characterized through
inspection of the difference of the single electron density matrix
of the excited (Pex) and ground states (P0). The difference between
the electronic density of S1 and S0 is depicted in Fig. 1 for Bn-1
(a), Bn-4N-1 (b), 2NBn-1 (c), and 4NBn-1 (e). Additionally, the dif-
ference densities of the S2 state of 2NBn-1 (d) and S3 of 4NBn-1
(f) are also illustrated. In Fig. 1, dark gray contours correspond to
Since all HNO donors spontaneously decompose in certain con-
ditions, benzohydroxamic acid (1) was selected as a stable model
of a HNO donor. By investigating the photochemistry of benzo-
hydroxamates and the photoactivated release of 1 from 2NBn-1,
it was expected that insights valuable to the use of this PPG to
release HNO donors would be gained. The parent benzyl benzo-
hydroxamate (Bn-1) has been investigated previously, and it was
concluded that Bn-1 undergoes a triplet Type II photoelimination
to yield benzamide (2) and benzaldehyde (3) [31]. In similar con-
ditions and at lower concentrations, we found that photolysis of
Bn-1 yields mainly 2 and 3 as the primary photoproducts, and
these results are summarized in Fig. 3. In the original work, it was
stated that the carbonyl products were generally not observed, and
an unknown by-product was often present. In our photolysis of
Bn-1, benzyl alcohol and benzoic acid were confirmed as products

4
W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
Table 1
Computationally predicted electronic configurations, excitation energies, and oscillator strength of selected excited states.
State
Electron configurationa
Vert. ex. Eb
fc
E (T1 − S0)d
Bn-1
S1
S2
n(CO), ␲*(CO)
n(CO), ␲*(CO)
102.3 (279.3 nm)
114.2 (250.2 nm)
0.0019
0.0282
T1
n(CO), ␲*(CO, aryl)
74.1
2NBn-1
S1
S2
S3
S4
S5
T1
n(NO2), ␲*(NO2)
n(CO), ␲*(NO2)
n(CO), ␲*(NO2)
n(NO2), ␲*(NO2)
Tr(aryl), ␲*(NO2)
n(CO), ␲*(NO2, aryl)
86.9 (329.1 nm)
88.6 (322.58 nm)
97.3 (293.81 nm)
98.9 (289.00 nm)
102.1 (279.96 nm)
0.0116
0.0018
0.0009
0.0008
0.0254
52.7
4NBn-1
S1
S2
S3
T1
n(NO2), ␲*(NO2)
n(CO), ␲*(NO2)
CT (aryl, NO2)
86.8 (329.6 nm)
91.5 (312.4 nm)
95.4 (299.81 nm)
0.0000
0.0001
0.0138
n(CO), ␲*(CO, aryl)
77.0
63.5
Bn-4N-1
S1
S2
T1
CT (aryl, NO2)
CT (aryl, NO2)
␲(aryl), ␲*(NO2, aryl)
82.2 (347.8 nm)
85.2 (335.7 nm)
0.0056
0.0008
a
Singlet characterized by the difference between the excited and ground state electron density matrix. Triplet characterized by examining singly occupied molecular
orbitals.
b
Vertical excitation energy (kcal mol−1).
Oscillator strength.
c
d
kcal mol−1, includes ZPE correction.
by chromatographic comparison to authentic samples. Typically,
benzyl alcohol and benzoic acid were minor products; however,
considerable variation in their product yields was observed. It was
also confirmed that photosensitization by irradiating Bn-1 in an
excess of benzophenone at 350 nm led to the same products. Thus,
the previously postulated Type II photoelimination from the triplet
state was verified.
While photolysis of 2NBn-1 was expected to release 1, it was
unclear if the addition of the nitro functional group would activate
other side reactions, and thus, the effect of nitro substitution on
the photochemistry of Bn-1 was examined. Unlike Bn-1, the lowest
excited singlet of Bn-4N-1 was predicted by TD-DFT to be a charge
transfer state (essentially a zwitterionic diradical) [54]. The photol-
ysis of Bn-4N-1 in wet acetonitrile led to 3, 4-nitrobenzamide (4)
and 4-nitrobenzoic acid (5) with a mass balance near 75%. A similar
result was obtained when the solvent was dried; however, the mass
balance dropped to 40%. The products and quantum yields for these
products are shown in Table 2. To ensure the observed products
did not arise from a thermal reaction, control solutions were kept
at 35 ◦C for 4 days in the dark. No decomposition of Bn-4N-1 or the
appearance of products was detected in these control experiments.
Regardless of the conditions, the quantum yields for these reac-
tions were very low (less than 0.005). The ratio of 4:5 was 1:4,
respectively, and their combined yield roughly correlated with
the yield of 3. However, when the photolysis was run in dry ace-
tonitrile, the ratio of 4:5 decreased to 1:1. It was also observed
that performing the photolysis under ambient atmosphere had
no effect on the observed quantum or product yields. Addition-
ally, rigorous exclusion of oxygen by five freeze–pump–thaw cycles
had no effect on the product ratio obtained in a 2% water ace-
tonitrile mixture upon photolysis at 294 nm. Thus, it was believed
that a significant portion of 5 arose from the hydrolysis of excited
Bn-4N-1 and not oxidation by a trace amount of advantageous
oxygen.
The TD-DFT predicted electronic configuration of S1 for both
2NBn-1 and 4NBn-1 was n(NO2), ␲*(NO2); however, only for
2NBn-1 was the location of nitro group amenable to abstrac-
tion of the benzyl hydrogen leading to the release of 1. Thus,
investigation of the photochemistry of 4NBn-1 was expected to
provide insight into side reactions with the potential to com-
pete with the release of the 2NBn. Photolysis of 4NBn-1 in wet

W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
5
Fig. 1. TD-B3LYP/6-31G(d) excited-state difference density plots (excited-state less the ground-state density). A dark gray shading indicates an area where electron density
is accumulated in the excited-state, and light gray shading represents an area from where the electron density is depleted from S0. (a) S1 Bn-1, (b) S1 Bn-4N-1, (c) S1 2NBn-1,
(d) S2 2NBn-1, (e) S1 4NBn-1, and (f) S3 4NBn-1. The surfaces were determined with a 0.023 a.u. isocountour value, and hydrogen atoms were not shown to improve figure
clarity.
Table 2
Product and quantum yields of the photolysis of benzyl 4-nitrobenzohydroxamate (Bn-4N-1).
Entry
1
Solvent
ꢁ (nm)
3
4
5
MeCN/2%H2O
254
0.0030 7a
(84 15)b
0.00063 29a
(67 2)b
0.00057 14a
(17 3)b
0.0023 12a
(61 13)b
2
MeCN/2%H2O
294
254
0.00023 3a
(14 1)b
0.00089 42a
(66 17)b
3
MeCN
(38 9)b
(20 11)b
(24 2)b
a
˚
95% confidence interval of last digits.
b
% Yield relative to loss of Bn-4N-1 95% confidence interval of last digit(s). % Yields determined at 30% conversion of Bn-4N-1.
acetonitrile produced a complex product mixture, and the dom-
inant products in this mixture were 4-nitrobenzaldehyde (6),
benzamide (7), benzoic acid (8), and 4-nitrobenzyl alcohol (9).
Product percent yields of these products are listed in Table 3.
Similar to Bn-4N-1, the quantum yields for the formation of these
products were less than 0.006 at 294 nm. Quantum yields at
254 nm could not be accurately measured due to the slow rate of
photoreaction.

6
W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
Table 3
Product and quantum yields of the photolysis of 4-nitrobenzyl benzohydroxamate (4NBn-1).
Entry
ꢁ (nm)
Solvent
Additive/technique
6
7
8
9
1
2
3
4
5
6
7
8
254
254
254
294
294
294
294
360
MeCN/2%H2O
MeCN
MeCN/2%H2O
MeCN/2%H2O
MeCN
MeCN/2%H2O
MeCN/2%H2O
MeCN/2%H2O
(52 24)a
(50 15)a
(74 28)a
0.0041 16c
0.0057 8c
0.0011 4c
(54 10)a
(83 17)a
(40 15)a
0.0012 5c
0.0012 2c
0.0012 3c
0.0018 5c
(13 16)a
(6.0 4.4)a
(6.7 0.4)a
(15 3)a
(3.0 1.8)a
(13 2)a
b
O2
(3.0 3.9)a
0.00038 14c
0.00059 18c
0.00017 8c
n.d.e
0.0022 14c
0.0013 6c
n.d.e
FPTd
Dienef
Ph2COh
–
0.00033 14c
(75 9)a
g
g
–
(18 6)a
a
% Yield relative to loss of 4NBn-1 95% confidence interval of last digit(s). % Yields determined at 30% conversion of 4NBn-1.
b
c
d
e
f
Sparged with O2 gas for 20 min before photolysis.
˚
95% confidence interval of last digit(s).
Freeze–pump–thaw cycles.
Not detected.
The addition of 2,4-dimethyl-1,3-pentadiene (80 mM).
Could not be determined.
Benzophenone (80 mM) sensitized photoreaction.
g
h
Table 4
Product and quantum yields of the photolysis of 2-nitrobenzyl benzohydroxamate (2NBn-1).
Entry
Solvent
ꢁ (nm)
1
7
10
1
2
3
MeCN/2%H2O
MeCN/2%H2O
MeCN/2%H2O
254
294
340
0.064 12a (45 7)b
0.14 1a
0.033 7a (37 4)b
0.042 9a (40 4)b
0.059 25a
0.062 4a
0.18 4a
0.053 11a
0.029 8a
a
˚
95% confidence interval of last digit(s).
b
% Yield relative to loss of 2NBn-1 95% confidence interval of last digit(s). % Yields determined at 30% conversion of 2NBn-1.
The stated aim of this work is to use the 2NBn PPG to photo-
chemically release 1 as a way to gain insight into the release of
HNO donors. One concern to the yield of 1 was the photosensitive
N–O bond of benzohydroxamates. The UV/Vis spectrum of 2NBn-1
closely resembled a superposition of 1 and 2-nitrotoluene, which
indicated that transitions associated with the two absorption bands
were localized on either the 1 or the 2NBn chromophores. Since the
ꢁmax of 1 was 225 nm, the first absorption band centered at 225 nm
for 2NBn-1 was assigned to the benzohydroxamate chromophore,
and shoulder beginning
at 260 nm and extending out to 370 nm to the 2NBn chro-
mophore. It was thought possible that since absorption appeared
localized that wavelength-dependent photochemistry might be
observed. Thus, the irradiation was carried out at three different
wavelengths: 254, 294, and 340 nm. These wavelengths were
selected since they covered the range of the UV absorption by
2NBn-1 and corresponded with the reported quantum yields for the
azoxybenzene actinometer. As seen in Table 4, the major product
at all wavelengths was 1, and quantum yields for the formation of
For Bn-4N-1, the formation of the benzoic acid product appeared
to arise from hydrolysis of an excited state. However, unlike Bn-
4N-1, no change in the product or quantum yields of benzoic acid
was observed when the photolysis of 4NBn-1 was carried out in
dry acetonitrile. Therefore, the effect of oxygen, triplet sensitiz-
ers, and triplet quenchers was examined. Rigorous exclusion of O2
by five freeze–pump–thaw cycles prevented the formation of ben-
zoic acid (8) and had little effect on the quantum yields of 6, 7,
and 9. Conversely, sparging the solution with O2 prior to irradi-
ation resulted in a 3-fold increase in the yield of 8. The addition
of a 20-fold excess of the triplet quencher 2,4-dimethylpentadiene
(80 mM) also resulted in a 10-fold reduction in the quantum yield of
benzoic acid while having a minimal effect on the quantum yields
of 7, and 9. The quantum yield of 6 could not be measured due
to a co-eluting peak in the chromatography. Irradiation of 4NBn-1
at 360 nm in excess concentrations (80 mM) of the photosensitizer
benzophenone resulted in benzoic acid being the major product.
Again, the yield of 6 could not be measured due to a co-eluting
peak.

W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
7
1 increased at longer wavelengths. The minor products 7 and
2-nitrobenzaldehyde (10) were observed at all wavelengths, and
the quantum yields for the formation of 7 and 10 were constant,
within experimental error, at the three wavelengths examined. The
release of the 2NBn PPG typically yields a 2-nitrosobenzaldehyde
remnant, which is known to undergo thermal or photoinduced
decomposition and was not detected in this study [34].
phenyl 4-nitrobenzoate, where irradiation in ethanol led to trans-
esterification rather than the expected photo-Fries rearrangement
[57].
Unlike Bn-4N-1, the lowest energy singlet excited state for
4NBn-1 was n(NO2), ␲*(NO2) rather than a CT state. Since
the S1 electronic configuration of 2NBn-1 was also n(NO2),
␲*(NO2), the photochemistry of 4NBn-1 was expected to resemble
While the above result clearly indicates that release of 1 by the
2NBn PPG was the dominant process, it was unclear how 7 and
10 were being generated. To determine if these minor products
were the result of a Type II photoelimination as observed for Bn-1,
the effects of common triplet quenchers and photosensitizers were
examined. As seen in Table 5, the addition of 1,3-cyclohexadiene
and p-terphenyl led to slightly decreased quantum yields of 1.
Additionally, oxygen quenched the formation of 1. The quan-
tum yields of 7 and 10 were largely unaffected or even slightly
increased upon the addition of the triplet quenchers. The T1 energy
predicted by B3LYP/6-31G(d) suggested energy transfer for pho-
tosensitizers with triplet energies above 53 kcal mol−1 should be
thermodynamically favorable. However, no reaction was observed
2NBn-1 more than Bn-4N-1 if internal conversion to S1 was effi-
cient. Additionally, since the position of the nitro group is critical
for the photoinduced release of 2NBn, the photoreactions of 4NBn-
1 were expected to provide insights into processes that could be
competitive with the release of 1 from 2NBn-1. Therefore, the low
quantum yields for the photoreactions of 4NBn-1 in Table 3 were
encouraging for the potential of 2NBn-1 to release 1.
It was considered that benzoic acid could potentially arise
from the hydrolysis of the S3 CT state of 4NBn-1, as with Bn-
4N-1. However, when 4NBn-1 was photolyzed in dry acetonitrile,
no change in the amount of benzoic acid (8) was observed as
shown in Table 3 (entries 1 and 2). While water content had no
effect, exposure to O2 led to higher product yields of benzoic
acid (entry 3). Likewise, rigorous exclusion of oxygen prevented
the formation of benzoic acid (entries 4 and 6). The effect of
O2 can also be used to explain the discrepancy between the
reported product yields at 254 nm and the quantum yields at
294 nm (entries 1 and 4). For the photoreactions used to deter-
mine product percent yields, a greater photon flux generated by
low pressure mercury bulbs was used, and thus, the photoreactions
were complete in less than hour. Due to the comparatively lower
photon flux generated by the Xe arc lamp, quantum yield mea-
surements were preformed over the course of 12 h, and multiple
time-points were analyzed from the same sample. Thus, the quan-
tum yield measurements were at higher risk for contamination
by O2.
with photosensitizers with triplet energies under 60 kcal mol−1
.
Only benzophenone, which has a triplet energy of 69 kcal mol−1
[56], was able to photosensitize the formation of 1, 7 and 10.
4. Discussion
In order to develop photoactivated HNO precursors, the overall
goal of these studies was to examine the effect of nitro substitu-
tion on the photochemistry of Bn-1. Homolysis of the N–O bond
was expected to be the main challenge to the release of the HNO
donor mimic 1, and thus, the effect of nitro substitution at dif-
ferent positions on the parent Bn-1 on N–O bond homolysis was
examined.
The nature of the S1 state was found to have an effect on the
observed photochemistry. Results from TD-DFT calculations indi-
cated that irradiation of Bn-4N-1 would generate a CT state. Since
the S1 state of Bn-1 was n, ␲*, it was expected that the photo-
chemistry of Bn-4N-1 would be different than the parent. Unlike
Bn-1, which only produced benzoic acid in trace amounts, pho-
tolysis of Bn-4N-1 in wet acetonitrile led to a significant amount
of 4-nitrobenzoic acid (5) as seen in entries 1 and 2 of Table 2.
Additionally, removal of water (Table 2, entry 3) led to a decrease
in the amount of 5 compared to the other nitro-containing prod-
uct (4), and thus, the formation of 5 appeared to arise from the
hydrolysis of photoexcited Bn-4N-1. The enhancement of solvol-
ysis by nitro substitution had also been previously reported for
Since triplet sensitization by benzophenone dramatically
increased the yield of benzoic acid (Table 3, entry 8), it was con-
cluded that benzoic acid arose primarily from a T1 reaction with
advantageous oxygen in solution. This hypothesis was supported
by the 4-fold decrease in the quantum yield of benzoic acid when
a diene triplet quencher was added to the solution as shown in
Table 3 entry 7. The percent yields of the other major products 6
and 7 were not affected by water, O2, or dienes, and thus, the forma-
tion of 6 and 7 was proposed to arise from the S1 state. It is possible
that 6 and 7 were the result of an N–O bond homolysis followed by
disproportionation as shown in Scheme 3. The minor alcohol prod-
uct 9 was likely the result of the nascent alkoxy radical, arising from

8
W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
Table 5
Quantum yields for 2-nitrobenzyl benzohydroxamate (2NBn-1) with various triplet photosensitizers and quenchers.
a
Entry
Additives
T1
ꢁ (nm)
1
7
10
1
2
3
4
5
6
7
Oxygen
1,3-Cyclohexadiene
p-Terphenyl
23
53
58
45
50
59
69
340
340
340
340
360
360
360
0.047 4b
0.10
1
0.062
0.015
0.062 18
n.d.
n.d.
n.d.
3
4
0.11
0.13
n.d.c
n.d.
3
2
0.041 10
0.099 32
n.d.
n.d.
n.d.
Acridine
9-Fluorenone
2-Acetonaphthalene
Benzophenone
n.d.
0.063 33
0.058 30
0.024 17
a
Triplet energies taken from Ref. [56].
b
c
˚
95% confidence interval of last digit(s).
Not detected.
N–O bond homolysis, abstracting a hydrogen atom from a solvent
molecule.
that irradiation at longer wavelengths would favor the release of
the 2NBn PPG by the mechanism shown in Scheme 4. Indeed, an
increase in the quantum yield for 1 was observed as the wavelength
of irradiation was increased as shown in Table 4. This indicated that
localized absorption by the 2NBn chromophore favored abstraction
of the benzyl hydrogen leading to the aci-nitro intermediate (I) and
the subsequent steps leading to the release of 1 [34].
Since the absorption band centered at 225 nm of 2NBn-1 was
believed to be localized on the benzohydroxamate chromophore,
it was a concern that irradiation at shorter wavelengths would
increase the percent yields of products associated with N–O bond
cleavage. The 2NBn chromophore was expected to absorb light at
higher wavelengths than the benzohydroxamate moiety of 2NBn-
1. The use of monochromatic irradiation at longer wavelengths
has been used to selectively activate one photoreactive center over
another center on the same molecule [35,37]. Thus, it was expected
The B3LYP/6-31G(d) T1 energy of 2NBn-1 was 52.7 kcal mol−1
,
however, only irradiation of benzophenone (T1 = 69 kcal mol−1
[56]) and not 2-acetonaphthone (T1 = 59 kcal mol−1 [56]) was able
to induce the release of 1 as shown in Table 5 entries 4–7. This sug-
gested that the actual triplet energy of 2NBn-1 was between 60
and 70 kcal mol−1 and the release of 1 can occur through the triplet
state. Supporting this assertion was the observed quenching of the
formation of 1 by the addition the triplet quenchers of p-terphenyl
and 1,3-cyclohexadiene (entries 2 and 3). Picosecond-pump probe
experiments have been used to suggest that the triplet state of
2NBn can abstract the benzylic hydrogen leading to a triplet birad-
ical that converts to I [49]. However, the quenching was modest,
which indicates that most of the release of 1 upon direct photolysis
of 2NBn-1 occurs through S1.
As shown in Scheme 5, three different processes were consid-
ered as possible pathways to the minor products 7 and 10. The
first was abstraction of the benzylic hydrogen by the carbonyl ini-
tiating a Type II photoelimination (Scheme 5a). The second was
N–O bond homolysis followed by disproportionation (Scheme 5b).
Finally, it was considered possible that the minor products 7 and
10 were generated from an alternative degradation of the aci-nitro
intermediate (I) (Scheme 5c).
Since both pathways (a) and (b) shown in Scheme 5 were
most likely to arise from excitation of the benzohydroxamate
chromophore, the quantum yields of 7 and 10 were expected to
decrease at longer wavelengths as the absorption by the 2NBn
moiety increased. However, irradiation at 340 nm still generated
7 and 10 in significant amounts (Table 4, entry 3). A few different
possibilities were considered to explain this observation at longer
wavelengths. Since benzophenone triplet sensitization yielded sig-
nificant amounts of 7 and 10 compared to 1, it was considered that
the formation of 7 and 10 arose from triplet state after intersys-
tem crossing. However, triplet quenchers had no effect or even
increased the observed quantum yields for 7 and 10 (Table 5, entries
1–3), and thus, it was concluded that the formation of 7 and 10 pre-
dominantly proceeds through a singlet excited state under direct
photolysis conditions. Only a very small difference in energy of S1
and S2 was predicted by TD-DFT calculation, and thus, S2 should be
accessible at longer wavelengths. The S2 state of 2NBn-1 was char-
acterized by TD-DFT as n(CO), ␲*(NO2), which could potentially
undergo a Type II mechanism (Scheme 5a). However, for 7 and 10
to arise from S2, slow relaxation from S2 to S1would be required,
which was considered unlikely. The much lower quantum yields
of 6 and 7 for 4NBn-1 suggest the minor products 7 and 10 were
due to a photochemical processes specific to 2NBn-1, which argues
Fig. 2. UV/Vis spectra in acetonitrile. (a) Solid line (2-nitrotoluene), short dash line
(1). (b) Short dash line (2NBn-1), solid line (4NBn-1), long dash line (Bn-4N-1).

W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
9
Scheme 3.
Scheme 4.
against the N–O bond homolysis mechanism (Scheme 5b). Thus, it
was thought most likely that the minor products 7 and 10 arose
from an alternative degradation of I (Scheme 5c).
(Table 4, entry 1) and benzophenone photosensitization at 360 nm
(Table 5, entry 7).
For the benzophenone-sensitized photoreaction, analysis of the
B3LYP/6-31G(d) molecular orbitals of the triplet state (T1) of 2NBn-
1 found that the two SOMO’s were centralized on the carbonyl
oxygen and the nitro substituent. While abstraction of the ben-
zylic hydrogen by the nitro substituent would eventually lead to
I and eventually 1, abstraction by the carbonyl would lead to a
Type II elimination (Scheme 5a) and the minor products 7 and
If 1, 7, and 10 arose solely from the decomposition of I, it
would be expected that the ratio of the products would remain
constant over the wavelengths examined. However, the amount
of 1 compared to 7 and 10 decreases at shorter wavelengths. This
indicates that another process leading to 7 and 10 becomes opera-
tive at shorter wavelengths. It is possible that excess vibrational
energy generated with the higher energy photons leads to N–O
bond homolysis (Scheme 5b). Alternatively, excitation into a ␲, ␲*,
such as S5 which has a high oscillator strength, eases intersystem
crossing into n, ␲* triplet states [58]. This assertion was consistent
with the similar product ratio observed for 254 nm directirradiation
Fig. 3. Yields of products upon photolysis of benzyl benzohydroxamate (Bn-1).
aPercent yields relative to loss of Bn-1 at 30% conversion, buncertainty reported
as 95% confidence interval.
Scheme 5.

10
W.R. Grither et al. / Journal of Photochemistry and Photobiology A: Chemistry 227 (2012) 1–10
10. Thus, product yields for the benzophenone sensitization may
reflect competition for the benzylic hydrogen between the carbonyl
(Scheme 5a) and the nitro (Scheme 5c) functional groups.
[15] B. Lopez, D. Wink, J. Fukuto, Arch. Biochem. Biophys. 465 (2007) 430–436.
[16] M. Doyle, S. Mahapatro, R. Broene, J. Guy, J. Am. Chem. Soc. 110 (1988) 593–599.
[17] S. Maraj, S. Khan, X. Cui, R. Cammack, C. Joannou, M. Hughes, Analyst 120 (1995)
699–703.
[18] H. Ohshima, I. Gilibert, F. Bianchini, Free Radic. Biol. Med. 26 (1999) 1305–1313.
[19] F. Kohout, F. Lampe, J. Am. Chem. Soc. 87 (1965) 5795–5796.
[20] V. Shafirovich, S.V. Lymar, Proc. Natl. Acad. Sci. U. S. A. 99 (2002) 7340–7345.
[21] K.M. Miranda, H.T. Nagasawa, J. Toscano, Curr. Top. Med. Chem. 5 (2005)
649–664.
[22] Y. Adachi, H. Nakagawa, K. Matsuo, T. Suzuki, N. Miyata, Chem. Commun. (2008)
5149–5151.
[23] A.D. Cohen, B.B. Zeng, S.B. King, J.P. Toscano, J. Am. Chem. Soc. 125 (2003)
1444–1445.
[24] G. Mayer, A. Heckel, Angew. Chem. Int. Ed. 45 (2006) 4900–4921.
[25] R.S. Givens, P.G. Conrad, A.L. Yousef, J. Leel, CRC Handbook of Organic Photo-
chemistry and Photobiology, second ed., CRC Press, Boca Raton, 2004 (Chapter
69).
[26] C. Walling, A.N. Naglieri, J. Am. Chem. Soc. 82 (1960) 1820–1825.
[27] T. Sakurai, S. Yamada, H. Inoue, Chem. Lett. (1983) 4975–4978.
[28] T. Sakurai, H. Yamamoto, S. Yamada, H. Inoue, Bull. Chem. Soc. Jpn. 58 (1985)
1174–1181.
[29] B. Hosangadi, P. Chhaya, M. Nimbalkar, N. Patel, Tetrahedron 43 (1987)
5375–5380.
[30] E. Lipczynska-Kochany, Chem. Rev. 91 (1991) 477–491.
[31] J.E. Johnson, M. Arfan, R. Hodzi, L.R. Caswell, S. Rasmussen, Photochem. Photo-
biol. 51 (1990) 139–144.
5. Conclusions
The stated aim of this work was to determine if selective exci-
tation of the 2NBn chromophore within 2NBn-1 would result in
increased release of 1. Quantum yields for the formation of 1
increased at longer wavelengths and the percent yields of the minor
products decreased, which supported the proposed hypothesis. The
totality of these results also suggested that all of the products
formed during direct photolysis of 2NBn-1 can be formed from
either the S1 or the T1 state. Given the wavelength-dependent quan-
tum yields for 1, the relatively constant quantum yields for the
minor products 7 and 10 over several wavelengths were taken as
an indication that two different processes were responsible for the
formation of 7 and 10. At longer wavelengths, it was suspected
that an alternative decomposition of I gave rise to these minor
products. Irradiation at higher energies was thought to allow easier
access to the triplet manifold through a ␲, ␲* singlet. Photosensiti-
zation indicated that equal amounts of 1 and 7 were formed from T1.
The position of the nitro substituent also influenced the observed
photochemistry. When the lowest energy excited state had charge
transfer character, hydrolysis became possible. Positioning of the
nitro substituent to disfavor the hydrogen abstraction dramatically
decreased the quantum yields for the loss of the starting materials,
and thus, the addition of a nitro substituent was not expected to
increase the amount of N–O bond homolysis.
[32] R. White, K. Oppliger, J. Johnson, J. Photochem. Photobiol. A 101 (1996) 197–200.
[33] M.M. Aly, A.M. Fahmy, F.F. Abdellatif, M.Z.A. Badr, Bull. Pol. Acad. Sci., Chem. 35
(1987) 47–52.
[34] Y. Il‘ichev, M. Schworer, J. Wirz, J. Am. Chem. Soc. 126 (2004) 4581–4595.
[35] A. Blanc, C.G. Bochet, J. Org. Chem. 67 (2002) 5567–5577.
[36] C. Bochet, Angew. Chem. Int. Ed. 40 (2001) 2071–2073.
[37] A. Blanc, C.G. Bochet, Org. Lett. 9 (2007) 2649–2651.
[38] M.J. Frisch, G.W. Trucks, H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Cheeseman,
G. Scalmani, V. Barone, B. Mennucci, G.A. Petersson, H. Nakatsuji, M. Caricato,
X. Li, H.P. Hratchian, A.F. Izmaylov, J. Bloino, G. Zheng, J.L. Sonnenberg, M. Hada,
M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda,
O. Kitao, H. Nakai, T. Vreven, J.A. Montgomery Jr., J.E. Peralta, F. Ogliaro, M.
Bearpark, J.J. Heyd, E. Brothers, K.N. Kudin, V.N. Staroverov, R. Kobayashi, J.
Normand, K. Raghavachari, A. Rendell, J.C. Burant, S.S. Iyengar, J. Tomasi, M.
Cossi, N. Rega, J.M. Millam, M. Klene, J.E. Knox, J.B. Cross, V. Bakken, C. Adamo,
J. Jaramillo, R. Gomperts, R.E. Stratmann, O. Yazyev, A.J. Austin, R. Cammi, C.
Pomelli, J.W. Ochterski, R.L. Martin, K. Morokuma, V.G. Zakrzewski, G.A. Voth,
P. Salvador, J.J. Dannenberg, S. Dapprich, A.D. Daniels, Ö. Farkas, J.B. Foresman,
J.V. Ortiz, J. Cioslowski, D.J. Fox, Gaussian 09, Gaussian, Inc., Wallingford, CT,
2009.
[39] R. Stratmann, G. Scuseria, M. Frisch, J. Chem. Phys. 109 (1998) 8218–8224.
[40] G.A. Zhurko, D.A. Zhurko, ChemCraft, Pilmus Inc., Milpatas, CA, 2007.
[41] W.L.P. Armarego, D.D. Perrin, Purification of Laboratory Chemicals, Pergamon
Press Inc., Elmsford, NY, 1998.
[42] O. Miyata, T. Koizumi, H. Asai, R. Iba, T. Naito, Tetrahedron 60 (2004) 3893–3914.
[43] A.S. Singha, B.N. Misra, Indian J. Chem. 21B (1982) 361–363.
[44] A. Gissot, A. Volonterio, M. Zanda, J. Org. Chem. 70 (2005) 6925–6928.
[45] O. Brady, F. Peakin, J. Chem. Soc. (1930) 226–229.
References
[1] W. Flores-Santana, C. Switzer, L.A. Ridnour, D. Basudhar, D. Mancardi, S.
Donzelli, D.D. Thomas, K.M. Miranda, J.M. Fukuto, D.A. Wink, Arch. Pharm. Res.
32 (2009) 1139–1153.
[2] N. Paolocci, M. Jackson, B. Lopez, K. Miranda, C. Tocchetti, D. Wink, A. Hobbs, J.
Fukuto, Pharmacol. Therapeut. 113 (2007) 442–458.
[3] J.M. Fukuto, M.D. Bartberger, A.S. Dutton, N. Paolocci, D.A. Wink, K.N. Houk,
Chem. Res. Toxicol. 18 (2005) 790–801.
[4] D.A. Wink, K.M. Miranda, T. Katori, D. Mancardi, D.D. Thomas, L. Ridnour, M.G.
Espey, M. Feelisch, C.A. Colton, J.M. Fukuto, P. Pagliaro, D.A. Kass, N. Paolocci,
Am. J. Physiol. Heart Circ. Physiol. 285 (2003) H2264–H2276.
[5] K.M. Miranda, N. Paolocci, T. Katori, D.D. Thomas, E. Ford, M.D. Bartberger, M.G.
Espey, D.A. Kass, M. Feelisch, J.M. Fukuto, D.A. Wink, Proc. Natl. Acad. Sci. U. S.
A. 100 (2003) 9196–9201.
[6] E.G. DeMaster, B. Redfern, H.T. Nagasawa, Biochem. Pharmacol. 55 (1998)
2007–2015.
[7] H.T. Nagasawa, E.G. DeMaster, B. Redfern, F.N. Shirota, D.J. Goon, J. Med. Chem.
33 (1990) 3120–3122.
[46] P. Mamalis, J. Green, D. Mchale, J. Chem. Soc. (1960) 229–238.
[47] N.J. Bunce, J. LaMarre, S.P. Vaish, Photochem. Photobiol. 39 (1984) 531–533.
[48] M. Schworer, J. Wirz, Helv. Chim. Acta 84 (2001) 1441–1458.
[49] R. Yip, Y. Wen, D. Gravel, R. Giasson, D. Sharma, J. Phys. Chem. 95 (1991)
6078–6081.
[8] N. Paolocci, T. Katori, H. Champion, M. St John, K. Miranda, J. Fukuto, D. Wink,
D. Kass, Proc. Natl. Acad. Sci. U. S. A. 100 (2003) 5537–5542.
[9] P. Pagliaro, D. Mancardi, R. Rastaldo, C. Penna, D. Gattullo, K.M. Miranda, M.
Feelisch, D.A. Wink, D.A. Kass, N. Paolocci, Free Radic. Biol. Med. 34 (2003)
33–43.
[10] C.G. Tocchetti, W. Wang, J.P. Froehlich, S. Huke, M.A. Aon, G.M. Wilson, G. Di
Benedetto, B. O’Rourke, W.D. Gao, D.A. Wink, J.P. Toscano, M. Zaccolo, D.M. Bers,
H.H. Valdivia, H. Cheng, D.A. Kass, N. Paolocci, Circ. Res. 100 (2007) 96–104.
[11] J.M. Fukuto, K. Chiang, R. Hszieh, P. Wong, G. Chaudhuri, J. Pharmacol. Exp. Ther.
263 (1992) 546–551.
[12] E. Bermejo, D.A. Sáenz, F. Alberto, R.E. Rosenstein, S.E. Bari, M.A. Lazzari,
Thromb. Haemost. 94 (2005) 578–584.
[13] A. Ellis, C.G. Li, M.J. Rand, Br. J. Pharmacol. 129 (2000) 315–322.
[14] W. Kim, Y. Choi, P. Rayudu, P. Das, W. Asaad, D. Arnelle, J. Stamler, S. Lipton,
Neuron 24 (1999) 461–469.
[50] D. Gravel, R. Giasson, D. Blanchet, R. Yip, D. Sharma, Can. J. Chem. 69 (1991)
1193–1200.
[51] R. Yip, D. Sharma, R. Giasson, D. Gravel, J. Phys. Chem. 89 (1985) 5328–5330.
[52] R. Yip, D. Sharma, R. Giasson, D. Gravel, J. Phys. Chem. 88 (1984) 5770–5772.
[53] L. Hviid, J.W. Verhoeven, A.M. Brouwer, M.N. Paddon-Row, J. Yang, Photochem.
Photobiol. Sci. 3 (2004) 246–251.
[54] D.E. Falvey, C. Sundararajan, Photochem. Photobiol. Sci. 3 (2004) 831–838.
[55] N. Turro, V. Ramamurthy, W. Cherry, W. Farneth, Chem. Rev. 78 (1978)
125–145.
[56] S.L. Murov, I. Carmichael, G.L. Hug, Handbook of Photochemistry, second ed.,
Marcel Dekker Inc., New York, NY, 1993.
[57] R. Finnegan, D. Knutson, J. Am. Chem. Soc. 90 (1968) 1670–1671.
[58] N. Turro, Modern Molecular Photochemistry, University Science Books, Sausal-
ito, CA, 1991 (Chapter 6).