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AVeline et al.
the apolipoproteins of human low density lipoprotein (LDL)26
and bovine serum albumin (BSA).27 This compound is impor-
tant considering the lack of convenient sources for the study of
•OH reactions in biologically relevant systems28,29 and the
general effort put into the development of new, selective
•
photochemical OH generators.30-41
We report here the results of a study of the photochemistry
of the bishydroperoxy naphthaldiimide, NP-III, and related
molecules. In order to elucidate the potential bioreactivity of
NP-III, it was necessary to investigate the structural effects
(nature of the substituent on the nitrogen atom, mono- or
bifunctionality of the molecule) on the primary and subsequent
photochemistry of these compounds. In this manner, we have
demonstrated a variety of possible mechanisms by which
photoactivation of NP-III could result in biological damage and
shown that the exact mechanism in operation will depend on
the microenvironment of the excited molecule.
Figure 1. Chemical structures of N-(2-hydroperoxy-2-methoxyethyl)-
1,8-naphthalimide (NP-II) and N,N′-bis(2-hydroperoxy-2-methoxy-
ethyl)-1,4,5,8-naphthaldiimide (NP-III).
potential application in light-induced tissue-repair and welding
of meniscal cartilage, articular cartilage and cornea has been
achieved in Vitro.13,14 The combination of high photoactivity
and DNA-intercalation has also prompted the use of some
naphthalenic derivatives as sequence-specific DNA-photo-
nucleases.15 Depending on the chemical structure of the
compound used, various mechanisms have been proposed,
involving free radical formation,16 photogeneration of carbo-
cation,17 electron-transfer from oxidizable guanine residues
(G),18 or hydrogen abstraction from thymine,19 but a lack of
experimental evidence precludes the confirmation of these
proposals.
Experimental Section
Chemicals. All reagents used were supplied by Sigma Chemical
Co. (St. Louis, MO), Fisher Chemical (Pittsburgh, PA) or Aldrich
Chemical Co. (Milwaukee, WI). Organic solvents were of spectro-
scopic grade from Fisher, deuterated acetonitrile (CD3CN: 99.6 atom
% D) was from Aldrich and Phosphate Buffered Saline Salt Solution
(PBS, pH ) 7.2) was purchased from Gibco BRL (Grand Island, NY).
Figures 1 and 2 show the chemical structures of the naphthalimide
and naphthaldiimide derivatives studied. 1,8-Naphthalimide (1) was
provided by Aldrich and used as supplied. NP-II, NP-III, and
compounds 2 and 3 were synthesized and purified as previously
described.20,25,42 All the naphthalenic derivatives were stored in solid
form at -20 °C. Solutions were prepared immediately before use and
were protected from light at all times. In case of low hydrophilicity,
a concentrated solution of naphthalenic derivative in acetonitrile was
diluted with PBS (the pH of the resulting solution was verified to be
7.2, and the percentage by volume of acetonitrile in the final sample
was <1%).
The most intriguing molecules of this family are the hydro-
peroxy derivatives NP-II and NP-III (presented in Figure 1),
which have been designed with the goal of generating the
hydroxyl radical (•OH) upon near-UV irradiation and named
“photo-Fenton reagents”.20,21 Photoexcitation of these com-
pounds has been shown to cause DNA-strand breaks20,21 and
in the case of the naphthaldiimide (NP-III), a sequence-specific
cleavage at the 5′G of 5′-GG-3′ sites has been observed.21,22
NP-III has also been used to investigate oxidative modifications
General Techniques. Ground-state absorption properties were
studied using a Cary 2300 UV-visible spectrophotometer or a Hewlett-
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•
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