Regioselectivity in the adiabatic photocleavage of DNA-based oxetanes

Article abstract of DOI:10.1039/d0ob01974g

Direct absorption of UVB light by DNA may induce formation of cyclobutane pyrimidine dimers and pyrimidine-pyrimidone (6-4) photoproducts. The latter arise from the rearrangement of unstable oxetane intermediates, which have also been proposed to be the electron acceptor species in the photoenzymatic repair of this type of DNA damage. In the present work, direct photolysis of oxetanes composed of substituted uracil (Ura) or thymine (Thy) derivatives and benzophenone (BP) have been investigated by means of transient absorption spectroscopy from the femtosecond to the microsecond time-scales. The results showed that photoinduced oxetane cleavage takes place through an adiabatic process leading to the triplet excited BP and the ground state nucleobase. This process was markedly affected by the oxetane regiochemistry (head-to-head, HH, vs. head-to-tail, HT) and by the nucleobase substitution; it was nearly quantitative for all investigated HH-oxetanes while it became strongly influenced by the substitution at positions 1 and 5 for the HT-isomers. The obtained results clearly confirm the generality of the adiabatic photoinduced cleavage of BP/Ura or Thy oxetanes, as well as its dependence on the regiochemistry, supporting the involvement of triplet exciplexes. As a matter of fact, when formation of this species was favored by keeping together the Thy and BP units after splitting by means of a linear linker, a transient absorption at ～400 nm, ascribed to the exciplex, was detected. This journal is

Full text of DOI:10.1039/d0ob01974g

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Regioselectivity in the adiabatic photocleavage of
DNA-based oxetanes†
Cite this: Org. Biomol. Chem., 2020,
18, 9117
Alejandro Blasco-Brusola, Ignacio Vayá * and Miguel A. Miranda
*
Direct absorption of UVB light by DNA may induce formation of cyclobutane pyrimidine dimers and pyri-
midine–pyrimidone (6-4) photoproducts. The latter arise from the rearrangement of unstable oxetane
intermediates, which have also been proposed to be the electron acceptor species in the photoenzymatic
repair of this type of DNA damage. In the present work, direct photolysis of oxetanes composed of substi-
tuted uracil (Ura) or thymine (Thy) derivatives and benzophenone (BP) have been investigated by means of
transient absorption spectroscopy from the femtosecond to the microsecond time-scales. The results
showed that photoinduced oxetane cleavage takes place through an adiabatic process leading to the
triplet excited BP and the ground state nucleobase. This process was markedly affected by the oxetane
regiochemistry (head-to-head, HH, vs. head-to-tail, HT) and by the nucleobase substitution; it was nearly
quantitative for all investigated HH-oxetanes while it became strongly influenced by the substitution at
positions 1 and 5 for the HT-isomers. The obtained results clearly confirm the generality of the adiabatic
photoinduced cleavage of BP/Ura or Thy oxetanes, as well as its dependence on the regiochemistry, sup-
porting the involvement of triplet exciplexes. As a matter of fact, when formation of this species was
favored by keeping together the Thy and BP units after splitting by means of a linear linker, a transient
absorption at ∼400 nm, ascribed to the exciplex, was detected.
Received 26th September 2020,
Accepted 29th October 2020
DOI: 10.1039/d0ob01974g
rsc.li/obc
Benzophenone (BP) is a well-known photosensitizing moiety
that is present in a variety of drugs.14 It can induce damage to
Introduction
The origin of human diseases such as skin cancer can be associ- DNA, and its main target is the thymine (Thy) nucleobase.15–17
ated in many cases to photoinduced DNA damage.1–3 Thus, In this regard, excitation of BP with UVA light generates its
when DNA absorbs UV light, a cascade of reactions may lead to triplet excited state (3BP*), which can then induce formation of
the formation of cyclobutane pyrimidine dimers (CPDs) and to a CPDs through a formal [2 + 2] cycloaddition between two adja-
lesser extent pyrimidine–pyrimidone (6-4) photoproducts ((6-4) cent thymine units, or alternatively it may lead to oxetane
PPs), which may result in the appearance of mutations and cell derivatives through a triplet exciplex 3[BP⋯Thy]*.18–20
death.4–7 The CPD photolesions can also arise from photosensiti-
The photoreactivity of monomeric pyrimidine bases such as
zation, where an endogenous or exogenous photosensitizer (PS) thymine or uracil (Ura) interacting with BP is highly influenced
can absorb UVA light to generate long-lived reactive species such by the substituents at position C5 of the nucleobase, mainly
as excited triplet states that will be the initiators of the damaging due to steric hindrance. In this context, CPDs formation in
reactions.8,9 Nature has developed efficient mechanisms to repair solutions containing BP and Ura derivatives is completely
these lesions. In this context, the nucleotide excision repair oper- blocked by the presence of a bulky group at the C5 position of
ates in humans, while in other organisms such as bacteria or the nucleobase.21 In connection with oxetanes, the Paternò-
plants the involvement of a photolyase reverses the damage Büchi photoreaction between Thy and BP derivatives has been
through a photoinduced electron transfer process.10–13
reported in the course of model studies related to the photoen-
zymatic repair of DNA (6-4)PPs.22–27 Interestingly, direct photo-
lysis of some of these oxetanes results in a rare adiabatic clea-
vage to form 3BP* and Thy in its ground state.23,26 This
process has recently been investigated in detail for two head-
to-head (HH) and head-to-tail (HT) oxetane regioisomers
obtained from 1,3-dimethylthymine (DMT) and BP (see
Scheme 1).18 Irradiation of any of the two isomers induces
instantaneous formation of its first excited singlet state, which
rapidly evolves to a singlet biradical through C–C bond scis-
Departamento de Química/Instituto de Tecnología Química UPV-CSIC,
Universitat Politècnica de València, Camino de Vera s/n, 46022 València, Spain.
E-mail: mmiranda@qim.upv.es, igvapre@qim.upv.es
†Electronic supplementary information (ESI) available: Synthesis and character-
ization of the uracil and thymine derivatives and the corresponding intermedi-
ates by means of 1H and 13C NMR and HRMS. Synthetic schemes, femtosecond
transient absorption and laser flash photolysis spectra and kinetic traces. See
DOI: 10.1039/d0ob01974g
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Scheme 1 Schematic representation of the photoinduced cyclorever-
sion process of the HH and HT oxetanes composed of BP and DMT.18
For clarification, the HH regioisomer follows the blue arrows, while the
HT oxetane follows the red ones.
Fig. 1 Chemical structure of the investigated systems. (A) Oxetanes
arising from the intermolecular photoreaction between BP and Ura
(from HH-1 to HT-4) or Thy (HH-5 and HT-5) derivatives. (B) Oxetanes
arising from the intramolecular photoreaction between BP and Thy units
covalently linked by spacers of different nature.
sion. For the HH-isomer, step (III) to reach the excited triplet
exciplex 3[BP⋯DMT]* has been found to be much more
efficient than for the HT-isomer, which may follow step (IV) to
generate a triplet biradical that evolves to regenerate either the
starting oxetane or the separated Thy and BP chromophores in
the ground state. Alternatively, the triplet exciplexes are also
formed from the HT oxetane, albeit to a lesser extent; these
gated systems, direct steady-state photolysis with UVC light
gave rise to the starting materials BP and the Ura or Thy
derivatives as the only photoproducts. The synthesis of HH-1
has been described earlier;28 however, we have slightly modi-
fied the synthetic procedure to get the different HH- and HT-
regioisomers (see Scheme 2). Briefly, solutions containing BP
(0.2 M) and the nucleobase derivative (0.1 M) in acetonitrile
were placed into Pyrex tubes and irradiated ca. 6 h under nitro-
gen in a Luzchem photoreactor emitting at λmax = 350 nm (12
× 8 W lamps). The crudes were purified by column chromato-
graphy to get the final HH- and/or HT-oxetanes as white
solids. The synthesis of the precursor nucleobase derivatives is
explained in detail in the ESI.†
Due to the chemical instability of the uracil-derived HT-1
and HT-2, which could not be isolated in pure form, only the
photophysical properties of their HH-regioisomers were inves-
tigated. Thus, nanosecond laser flash photolysis (LFP)
measurements were performed upon excitation at 266 nm in
deaerated acetonitrile, using BP as a reference. Under these
conditions, photolysis of either HH-1 or HH-2 resulted in the
instantaneous formation of a transient absorption peaking at
530 nm, where 3BP* displays its maximum.29 This result
agrees well with the previous observations for the HH BP-DMT
oxetane.18 Thus, after the laser pulse, a nearly quantitative
adiabatic population of 3BP* was achieved for both oxetanes
3
species finally dissociate to give BP* and DMT in the ground
state. Consequently, the adiabatic cycloreversion of the two
investigated oxetanes seems to be markedly influenced by the
HH- vs. the HT-regiochemistry.
With this background, the aim of the present work is to
establish the generality and scope of the adiabatic cyclorever-
sion process and to gain further insight into its mechanistic
features, using a variety of oxetanes with different regiochemis-
tries and substitution patterns at positions 1 and 5 of the
nucleobase. With this goal, different oxetanes have been
obtained by the intermolecular photoreaction between BP and
Ura or Thy derivatives. In addition, oxetanes arising from the
analogous intramolecular process between Thy and BP units
covalently connected by spacers of different nature have also
been studied in order to evaluate the possible influence of the
linker on the photolytic splitting. Thus, the photobehavior of a
variety of HH- and HT-oxetanes (see Fig. 1) has been investi-
gated by means of nanosecond laser flash photolysis (LFP)
and, in some cases, femtosecond transient absorption spec-
troscopy. As a matter of fact, the obtained results confirm the
generality of the adiabatic photocleavage in Thy/BP oxetanes,
as well as its dependence on the HH- vs. HT-regiochemistry.
Moreover, they indicate that the ring opening process is highly
influenced by the nature of the substituents at positions 1 and
5 of the nucleobase.
Results and discussion
The photobehavior of oxetanes resulting from the inter-
molecular photoreaction between BP and Ura or Thy deriva-
Scheme 2 Schematic representation of the photoinduced cyclorever-
tives (see Fig. 1A) was studied first. In general, for all investi- sion process of oxetanes composed of BP and DMT.
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group at position 1) as well as the corresponding Thy ana-
logues HH-5 and HT-5. As stated above for the related com-
pounds HH-3 and HT-3, the ns LFP decay traces at 530 nm for
HH-4 and HT-4 (Fig. S11A in ESI†) evidenced a remarkable
regioselectivity (end-of-pulse 3BP* ratio ca. 4 : 1) in the adia-
batic cycloreversion of the investigated oxetanes. Thus, replace-
ment of the methyl group with a methyl acetate substituent at
position 1 for the tert-butyl derivatives mainly affected the
adiabaticity of the HT-isomer, which decreased from 35% in
HT-3 to 20% in HT-4. This could be associated with a possible
stabilization of the triplet biradical (see Scheme 1) by the influ-
ence of the methyl acetate moiety, which could hinder for-
mation of the triplet exciplex (step III) thus favoring step VII to
generate the starting oxetane HT-4 or the separated BP and
Thy chromophores in the ground state. For the thymine deriva-
tives HH-5 and HT-5 (Fig. S11B in ESI†), a regiodifferentiation
was again observed in the adiabatic ring-opening (end-of-pulse
3BP* ratio ca. 7 : 3), which was comparable to the results
recently reported for BP-DMT oxetanes.18
In view of the photoreactivity observed for the nucleobase-
derived oxetanes as a function of the substitution at positions
1 and 5, the photobehavior of polycyclic oxetanes (see Fig. 1B)
whose splitting could lead to BP and Thy chromophores co-
valently linked through a spacer of different nature was also
investigated. Their synthesis was achieved following the
reported procedures,26,30 by means of the Paternò-Büchi
photoreaction between the BP and Thy units of the precursor
dyads 1 and 2 (see Fig. 4). Thus, photolysis of the dyad 1 con-
taining the BP and Thy chromophores linked through a sugar
moiety resulted in the formation of HH-6, HT-6, HH-7 and
HT-7,26 whereas HH-8 was obtained from irradiation of dyad 2,
where BP and Thy are connected through a linear chain of ten
linking atoms.30 Steady state photolysis of oxetanes HH-6,
HT-6, HH-7 and HT-7 gave rise in all cases to dyad 1 as the
only photoproduct, whereas, under the same conditions,
oxetane HH-8 led cleanly to dyad 2.
Fig. 2 LFP decay traces at 530 nm for (A) HH-1 (blue) and BP (black),
and (B) HH-2 (blue) and BP (black) after excitation at 266 nm in deaer-
ated acetonitrile.
(see Fig. 2). The decay traces were properly fitted by a one-
order exponential law with lifetimes of about 5 μs for BP and
HH-1 and shorter for HH-2 (ca. 2 μs). The difference in the
triplet lifetimes could be associated with a faster BP triplet
quenching by HH-2 than by HH-1.
Since the influence of the regiochemistry on the photobeha-
vior of the HT-uracil derivatives could not be evaluated due to
their chemical instability, two additional and more stable sub-
stituted uracil oxetanes, namely HH-3 and HT-3, were syn-
thesized with a bulky tert-butyl group at position 5. In this
context, LFP measurements evidenced clear differences
between the two regioisomers (see Fig. 3). Analysis of the
kinetic traces at 530 nm showed again an almost quantitative
3
end-of-pulse adiabatic population of BP* for HH-3, while this
process was much less efficient (ca. 35%) for HT-3. Therefore,
a strong regioselectivity was observed in the photoinduced
ring-opening reaction, which is in line with the results recently
reported for the HH and HT BP-DMT oxetanes;18 noteworthy,
in the case of 3 the difference in the degree of adiabaticity
found between the HH- and the HT-regioisomers was mark-
edly enhanced. Thus, the presence of a tert-butyl substituent at
position 5 had little if any effect on the photoreactivity of the
HH derivative, but it clearly affected the photobehavior of the
HT-isomer.
In order to complete the picture on the influence of the
substituents at positions 1 or 5 on the adiabatic cycloreversion
of nucleobase-derived oxetanes, two additional uracil-derived
compounds were synthesized, namely HH-4 and HT-4 (with a
bulky tert-butyl substituent at position 5 and a methyl acetate
Since the lifetime of 3BP* in dyad 1 was very short (∼20
ns),26 reliable transient absorption experiments could only be
performed in the femtosecond time-scale. Under the employed
experimental conditions, the degree of photodegradation was
kept below 5%. As it is shown in Fig. 5, excitation of the
different regio- and stereoisomeric oxetanes HH-6, HT-6, HH-7
and HT-7 at 280 nm in acetonitrile led to the formation of the
typical triplet–triplet absorption band of BP (see Fig. S12 in
ESI†);31,32 with maximum at ca. 530 nm with a time constant
of about 9.5 ps. Accordingly, the photoinduced cycloreversion
Fig. 3 LFP decay traces at 530 nm for BP (black), HH-3 (blue) and HT-3
(red) after excitation at 266 nm in deaerated acetonitrile.
Fig. 4 Chemical structures of dyad 1 and dyad 2.
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Fig. 5 Femtosecond transient absorption spectra from 0.5 to 40 ps for
(A) HH-6, (B) HT-6, (C) HH-7 and (D) HT-7. The insets show the kinetic
traces at 530 nm. All measurements were performed at λexc = 280 nm in
acetonitrile.
Fig. 6 Femtosecond transient absorption measurements after exci-
tation at 280 nm in acetonitrile: (A) spectra from 0.5 to 40 ps and (B)
kinetic traces at 340 nm (gray) and 530 nm (blue) for HH-8; (C) spectra
recorded at 10 ps for the BP-derived chromophore (black) and HH-8
(blue); (D) kinetic traces at 400 nm for the BP-derived chromophore
(black) and HH-8 (green).
for the intramolecular oxetanes HH-6, HT-6, HH-7 and HT-7
also operates as an adiabatic process.
Table 1 Relationship between the degree of adiabaticity and the substi-
tution at positions N1 and C5 of the nucleobase for the photoreversion
of the indicated oxetanes
Finally, the photophysical properties of HH-8 were investi-
gated and compared to those of dyad 2.30 Kinetic analysis of
the transient peaking at 530 nm in the ns-μs time-scale upon
LFP at 266 nm revealed that the decay traces in both systems
could be fitted by a first order exponential law, with lifetimes
of ca. 55 ns. More interestingly, HH-8 evidenced once more a
complete adiabatic photoreversion process (see Fig. S13 in
ESI†). However, the spectra and the decay traces were noisy,
due to the short triplet lifetime compared to the duration of
the pulse. Hence, oxetane HH-8 was also submitted to femto-
second transient absorption measurements (λexc = 280 nm),
which fully confirmed formation of the triplet excited state of
the BP-derived chromophore with maximum at ca. 530 nm
(ref. 30) (see Fig. 6A and B). As previously observed for the
BP-DMT regioisomers,18 an absorption at about 400 nm with a
Substituent at N1
Substituent at C5
Adiabaticity (%)
HH-1
HH-2
HH-3
HT-3
HH-4
HT-4
HH-5
HT-5
CH3
CH2CO2CH3
CH3
H
H
tert-Bu
tert-Bu
tert-Bu
tert-Bu
CH3
100
100
100
35
100
20
CH3
CH2CO2CH3
CH2CO2CH3
CH2CO2CH3
CH2CO2CH3
75
38
CH3
Conclusions
flat profile, which is absent for the BP-derived chromophore, The UV-induced photocleavage of a variety of oxetanes arising
was formed in ca. 3.5 ps (see Fig. 6C and D); this can be associ- from the inter- or intramolecular Paternò-Büchi photoreaction
ated with the formation of an intermediate excited triplet exci- between BP and Ura or Thy chromophores has been investi-
plex between the BP and Thy chromophores (see Scheme 1).
gated by means of transient absorption spectroscopy from the
In summary, photolysis of the investigated oxetanes femtosecond to the microsecond time-scales. In all cases, an
resulted in an adiabatic population of 3BP* after ring opening. adiabatic process is observed, which leads to the BP unit in its
In this context, a strong regioselectivity was detected for the triplet excited state. The degree of adiabaticity depends on the
oxetanes 1–5, being the degree of adiabaticity much higher for oxetane regiochemistry and on the substitution pattern at posi-
the HH than for the HT isomers (see Table 1). In addition, the tions 1 and 5 of the nucleobase. In general, a nearly quantitat-
process was significantly affected by the substitution at posi- ive adiabatic cycloreversion is observed for the HH-regio-
tion N1 and C5 of the nucleobase for the HT derivatives, isomers; by contrast, for the HT-oxetanes the process is not
whereas a less marked effect was noticed for the HH isomers.
fully adiabatic and becomes strongly influenced by the nature
Likewise, the photoinduced cleavage of the polycyclic oxe- of the substituents at positions 1 and 5. The highest regio-
tanes was found to occur as an adiabatic process. Interestingly, differentiation (4 : 1 ratio) is detected between HH-4 and HT-4.
for the tricyclic oxetane HH-8, a transient absorption at These results are consistent with the intermediacy of a triplet
∼400 nm is detected, which is ascribed to formation of the exciplex, whose formation is more favorable in the HH-regio-
purported triplet exciplex between the linked benzophenone isomers. In summary, the adiabatic cycloreversion of oxetanes
and thymine chromophores.
derived from aromatic ketones and pyrimidine bases appears
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to be a rather general process, which occurs in all the investi- quartz cuvettes at room temperature in deaerated conditions
gated compounds. Regioselectivity (HH vs. HT) and substi- (25 min N2 bubbling), using 10 mL of fresh solution in order
tution-dependence are interesting features of this process.
to avoid data acquisition from photodegraded products.
Control experiments indicated that the degree of decompo-
sition of all oxetanes after photolysis was lower than 5%.
Femtosecond transient absorption experiments were per-
formed using a pump–probe system. The femtosecond pulses
were generated with a mode-locked Ti-Sapphire laser of a
compact Libra HE (4 W power at 4 kHz) regenerative amplifier
delivering 100 fs pulses at 800 nm (1 mJ per pulse). The output
of the laser was split into two parts to generate the pump and
the probe beams. Thus, tunable femtosecond pump pulses
were obtained by directing the 800 nm light into an optical
parametric amplifier. In the present case, the pump was set at
280 nm and passed through a chopper prior to focus onto a
rotating cell (1 mm optical path) containing the samples in
organic solution. The white light used as probe was produced
after part of the 800 nm light from the amplifier travelled
through a computer controlled 8 ns variable optical delay line
and impinge on a CaF2 rotating crystal. This white light was in
Experimental section
Chemicals and reagents
1,3-Dimethyluracil, benzophenone (BP), dimethyl sulphate,
lithium diisopropylamide (LDA), ethyl formate 97%, thiourea,
chloroacetic acid, thymine-1-acetic acid, ethyl bromoacetate,
ethyl 4-bromobutyrate, were purchased from Sigma-Aldrich.
Ethyl-8-bromooctanoate was purchased from Fluorochem.
Methyl 3,3-dimethylbutanoate was purchased from Alfa-aesar.
Sodium hydroxide (NaOH), potassium hydroxide (KOH), mag-
nesium sulphate (MgSO4), hydrochloric acid 37% (HCl) and
acetonitrile spectroscopic grade were purchased from
Scharlab.
Spectroscopic techniques
1
The H NMR and 13C NMR spectra were recorded at 400 and turn split in two identical portions to generate reference and
100 MHz, respectively, using a Bruker AVANCE III instrument; probe beams that then were focused on the rotating cell con-
chemical shifts are reported in ppm.
taining the sample. The pump and the probe beams were
High-resolution mass spectrometry (HRMS) was performed made to coincide to interrogate the sample. The power of the
in an Ultra Performance Liquid Chromatography (UPLC) pump beam was set to 180 μW. Under these conditions, the
ACQUITY system (Waters Corp.) with a conditioned autosam- degree of photodegradation of all oxetanes was lower than 5%.
pler at 4 °C. The separation was accomplished on an ACQUITY A computer-controlled imaging spectrometer was placed after
UPLC BEH C18 column (50 mm × 2.1 mm i.d., 1.7 μm), which this path to measure the probe and the reference pulses to
was maintained at 40 °C. The analysis was performed using obtain the transient absorption decays/spectra. The experi-
acetonitrile and water (60 : 40 v/v containing 0.01% formic mental data were treated and compensated by the chirp using
acid) as the mobile phase with a flow rate of 0.5 mL min−1
,
the ExciPro program.
and injection volume was 5 μL. The Waters ACQUITY™
XevoQToF Spectrometer (Waters Corp.) was connected to the
Steady-state photolysis
UPLC system via an electrospray ionization (ESI) interface. Irradiations were performed in a Luzchem multilamp photo-
This source was operated in positive ionization mode with the reactor emitting at λmax = 350 nm (12 × 8 W lamps). Solutions
capillary voltage at 1.5 kV at 100 °C and the temperature of the containing the uracil or thymine derivative (0.1 M) and BP (0.2
desolvation was 300 °C. The cone and desolvation gas flows M) were irradiated through Pyrex for ca. 6 h; the crude pro-
were 40 L h−1 and 800 L h−1, respectively. The collision gas ducts containing the HH- and HT-oxetanes were purified by
flow and collision energy applied were 0.2 mL min−1 and 12 V, silica gel chromatography (hexane : ethyl acetate 80 : 20 v/v).
respectively. All data collected in Centroid mode were acquired
Oxetane HH-1
using Masslynx™ software (Waters Corp.). Leucine-enkephalin
was used at a concentration of 500 pg μL−1 as the lock mass 1H NMR (400 MHz, CDCl3) δ 7.44–7.29 (m, 10H), 5.20 (d, J =
generating an [M + H]+ ion (m/z 556.2771) and fragment at m/z 8.8 Hz, 1H), 4.94 (d, J = 8.8 Hz, 1H), 3.04 (s, 3H), 3.01 (s, 3H);
120.0813 and flow rate of 50 μL min−1 to ensure accuracy 13C NMR (100 MHz, CDCl3) δ 167.0, 151.7, 143.3, 138.7, 128.7,
during the MS analysis.
128.4, 128.3, 128.2, 125.9, 125.1, 96.0, 70.6, 60.7, 35.4, 27.3;
Steady-state absorption spectra were recorded in a JASCO HRMS (ESI): m/z calcd for C19H19N2O3 [M + H]+: 323.1396,
V-760 spectrophotometer. Laser Flash Photolysis (LFP) found: 323.1391
measurements were performed using a pulsed Nd:YAG L52137
V LOTIS TII at the excitation wavelength of 266 nm. The single
Oxetane HH-2
pulses were ca. 10 ns duration, and the energy was ∼12 mJ per 1H NMR (400 MHz, CDCl3) δ 7.44–7.26 (m, 10H), 5.24 (d, J =
pulse. The laser flash photolysis system consisted of the 8.8 Hz, 1H), 5.02 (d, J = 8.8 Hz, 1H), 4.47 (d, J = 17.6 Hz, 1H),
pulsed laser, a 77 250 Oriel monochromator and an oscillo- 3.77 (s, 3H), 3.37 (d, J = 17.6 Hz, 1H), 3.06 (s, 3H); 13C NMR
scope DP04054 Tektronix. The output signal from the oscillo- (100 MHz, CDCl3) δ 168.9, 166.6, 151.9, 142.5, 138.4, 128.7,
scope was transferred to a personal computer. Absorbances of 128.5, 128.4, 126.1, 125.4, 95.3, 70.2, 59.4, 52.7, 48.0, 27.5;
all solutions were adjusted at ∼0.20 at 266 nm in acetonitrile. HRMS (ESI): m/z calcd for C21H21N2O5 [M + H]+: 381.1450;
All UV and LFP measurements were done using 10 × 10 mm2 found: 381.1446.
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Oxetane HH-3
d’Educació Cultura
i
Esport (PROMETEO/2017/075 and
GRISOLIAP/2017/005) is gratefully acknowledged.
1H NMR (400 MHz, CD3CN) δ 7.41–7.23 (m, 10H), 4.94 (s, 1H),
3.21 (s, 3H), 2.70 (s, 3H), 0.98 (s, 9H); 13C NMR (100 MHz,
CDCl3) δ 169.7, 152.0, 144.2, 139.0, 128.3, 127.8, 127.6, 125.2,
124.4, 91.1, 83.5, 61.5, 35.8, 29.7, 27.4, 24.6; HRMS (ESI): m/z References
calcd for C23H27N2O3 [M + H]+: 379.2022; found: 379.2027.
1 K. H. Kraemer, Proc. Natl. Acad. Sci. U. S. A., 1997, 94, 11–
14.
Oxetane HT-3
2 J.-S. Taylor, Acc. Chem. Res., 1994, 27, 76–82.
3 F. Urbach, J. Photochem. Photobiol., B, 1997, 40, 3–7.
4 H. Mukhtar and C. A. Elmets, Photochem. Photobiol., 1996,
63, 356–357.
5 B. M. Sutherland, Photochem. Photobiol., 1996, 63, 375–
377.
1H NMR (400 MHz, CD3CN) δ 7.84–7.20 (m, 10H), 5.84 (s, 1H),
3.17 (s, 3H), 2.66 (s, 3H), 0.96 (s, 9H); 13C NMR (100 MHz,
CDCl3) δ 169.1, 153.0, 144.5, 128.2, 127.7, 127.4, 127.2, 125.9,
89.5, 84.3, 66.0, 35.9, 34.2, 29.7, 27.5; HRMS (ESI): m/z calcd
for C23H27N2O3 [M + H]+: 379.2022; found: 379.2026.
6 S. Tommasi, M. F. Denissenko and G. P. Pfeifer, Cancer
Res., 1997, 57, 4727–4730.
7 J. Cadet, A. Grand and T. Douki, Top. Curr. Chem., 2015,
356, 249–275.
8 J. Cadet, S. Mouret, J. L. Ravanat and T. Douki, Photochem.
Photobiol., 2012, 88, 1048–1065.
9 B. Epe, M. Pflaum and S. Boiteux, Mutat. Res., 1993, 299,
135–145.
Oxetane HH-4
1H NMR (400 MHz, CDCl3) δ 7.37–7.23 (m, 10H), 4.97 (s, 1H),
4.84 (d, J = 17.6 Hz, 1H), 3.82 (s, 3H), 3.73 (d, J = 17.6 Hz, 1H),
2.84 (s, 3H), 1.10 (s, 9H); 13C NMR (100 MHz, CDCl3) δ 169.8,
169.1, 152.1, 143.7, 139.1, 128.7, 128.4, 127.8, 125.4, 124.6,
90.6, 83.6, 59.7, 52.6, 48.3, 36.2, 27.5, 24.4; HRMS (ESI): m/z
calcd for C25H29N2O5 [M + H]+: 437.2076; found: 437.2071.
10 J. Li, Z. Liu, C. Tan, X. Guo, L. Wang, A. Sancar and
D. Zhong, Nature, 2010, 466, 887–890.
Oxetane HT-4
1H NMR (400 MHz, CDCl3) δ 7.79–7.19 (m, 10H), 5.81 (s, 1H),
4.80 (d, J = 17.6 Hz, 1H), 4.07 (d, J = 17.6 Hz, 1H), 3.80 (s, 3H),
2.79 (s, 3H), 1.04 (s, 9H); 13C NMR (100 MHz, CDCl3) δ 169.2,
168.9, 152.0, 142.6, 141.4, 128.1, 127.8, 127.5, 127.3, 127.1,
126.0, 89.8, 83.3, 66.0, 52.5, 47.1, 36.1, 27.8, 27.3; HRMS (ESI):
m/z calcd for C25H29N2O5 [M + H]+: 437.2076; found: 437.2073.
11 T. Todo, H. Ryo, K. Yamamoto, H. Toh, T. Inui, H. Ayaki,
T. Nomura and M. lkenaga, Science, 1996, 272, 109–112.
12 T. Todo, H. Takemori, H. Ryo, M. Ihara, T. Matsunaga,
O. Nikaido, K. Sato and T. Nomura, Nature, 1993, 361, 371–
374.
13 T. Todo, H. Tsuji, E. Otoshi, K. Hitomi, S.-T. Kim and
M. Ikenaga, Mutat. Res., 1997, 384, 195–204.
14 K. Surana, B. Chaudhary, M. Diwaker and S. Sharma,
MedChemComm, 2018, 9, 1803–1817.
15 F. Bosca and M. A. Miranda, J. Photochem. Photobiol., B,
1998, 43, 1–26.
16 T. Delatour, T. Douki, C. D’Ham and J. Cadet, J. Photochem.
Photobiol., B, 1998, 44, 191–198.
Oxetane HH-5
1H NMR (400 MHz, CDCl3) δ 7.38–7.25 (m, 10H), 4.78 (d, J =
17.6 Hz, 1H), 4.64 (s, 1H), 3.79 (s, 3H), 3.58 (d, J = 17.6 Hz,
1H), 2.85 (s, 3H), 1.76 (s, 3H); 13C NMR (100 MHz, CDCl3) δ
169.0, 168.8, 151.9, 143.8, 138.6, 128.7, 128.4, 128.1, 125.6,
125.1, 91.3, 65.3, 52.7, 48.3, 27.5, 23.7; HRMS (ESI): m/z calcd
for C22H23N2O5 [M + H]+: 395.1607; found: 395.1607.
17 J. E. Rogers and L. A. Kelly, J. Am. Chem. Soc., 1999, 121,
3854–3861.
Oxetane HT-5
18 A. Blasco-Brusola, M. Navarrete-Miguel, A. Giussani,
D. Roca-Sanjuán, I. Vayá and M. A. Miranda, Phys. Chem.
Chem. Phys., 2020, 22, 20037–20042.
19 M. C. Cuquerella, V. Lhiaubet-Vallet, F. Bosca and
M. A. Miranda, Chem. Sci., 2011, 2, 1219–1232.
20 P. Miró, M. Gómez-Mendoza, G. Sastre, M. C. Cuquerella,
M. A. Miranda and M. L. Marín, Chem. – Eur. J., 2019, 25,
7004–7011.
1H NMR (400 MHz, CDCl3) δ 7.55–7.18 (m, 10H), 5.29 (s, 1H),
4.64 (d, J = 23.6 Hz, 1H), 4.04 (d, J = 23.6 Hz, 1H), 3.78 (s, 3H),
2.78 (s, 3H), 1.52 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 169.1,
151.7, 141.8, 141.1, 128.3, 128.0, 127.7, 127.6, 125.3, 125.1,
90.5, 87.1, 52.5, 46.1, 27.8, 20.1; HRMS (ESI): m/z calcd for
C22H23N2O5 [M + H]+: 395.1607, found: 395.1606.
21 V. Vendrell-Criado, V. Lhiaubet-Vallet, M. Yamaji,
M. C. Cuquerella and M. A. Miranda, Org. Biomol. Chem.,
2016, 14, 4110–4115.
Conflicts of interest
There are no conflicts to declare.
22 S. Encinas, N. Belmadoui, M. J. Climent, S. Gil and
M. A. Miranda, Chem. Res. Toxicol., 2004, 17, 857–862.
23 A. Joseph and D. E. Falvey, J. Am. Chem. Soc., 2001, 123,
3145–3146.
Acknowledgements
Financial support from the Spanish Government (RYC-2015- 24 A. Joseph, G. Prakash and D. E. Falvey, J. Am. Chem. Soc.,
17737 and CTQ2017-89416-R) and from the Consellería
2000, 122, 11219–11225.
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25 G. Prakash and D. E. Falvey, J. Am. Chem. Soc., 1995, 117, 29 J. A. Bell and H. Linschitz, J. Am. Chem. Soc., 1963, 85, 528–
11375–11376. 532.
26 N. Belmadoui, S. Encinas, M. J. Climent, S. Gil and 30 A. Blasco-Brusola, I. Vayá and M. A. Miranda, J. Org. Chem.,
M. A. Miranda, Chem. – Eur. J., 2006, 12, 553–561. 2020, DOI: 10.1021/acs.joc.0c02088, accepted.
27 S.-T. Kim, K. Malhotra, C. A. Smith, J.-S. Taylor and 31 N. Tamai, T. Asahi and H. Masuhara, Chem. Phys. Lett.,
A. Sancar, J. Biol. Chem., 1994, 269, 8535–8540. 1992, 198, 413–418.
28 A. Joseph and D. E. Falvey, Photochem. Photobiol. Sci., 2002, 32 W. M. Kwok, X. Guan, L. M. Chu, W. Tang and
1, 632–635.
D. L. Phillips, J. Phys. Chem. B, 2008, 112, 11794–11797.
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