dna , with its building block dna bases 
 that store the genetic information , is indispensable in protein biosynthesis . 

investigations on long - distance charge transfer through dna are of 
 ongoing interest . moreover , the interaction of excited triplet states of aromatic 
 carbonyl compounds with dna bases is also of interest for biophysical 
 and biochemical studies .
these studies may serve as a useful strategy for understanding structure 
 and function of the bases in dna . in the present study
, we want to 
 extend the investigations on the interactions between an aromatic 
 ketone derivative , 3,3,4,4-benzophenone tetracarboxylic 
 acid , with selected dna bases .
3,3,4,4-benzophenone 
 tetracarboxylic acid along with 4-benzophenone monocarboxylic acid 
 ( 4-bc ) are derivatives of benzophenone , and their structures are similar . 

to get an idea about the kinetics of triplet bptc , the previous work 
 on 4-bc should be considered ; however , studies on the photochemical reaction 
 of 4-bc are challenging because of an existing overlap in the spectra 
 of triplet , ketyl radical , and ketyl radical anion between 380 and 
 720 nm .
although the overlapped spectra of these radicals with similar 
 molar extinction coefficients are comparable , the formation of these 
 radicals also depends on the type of quenchers and radical quantum 
 yields .
therefore , the various absorptions of the radicals need to 
 be considered carefully when measuring reaction rate constants . in 
 our experiments , we have employed different approaches to analyze 
 these kinetic behaviors . in this paper
, we present the kinetics 
 of the photoreaction of triplet bptc with various dna bases in aqueous 
 solution .
the ph - dependence of the quenching reaction rate constants 
 is investigated thoroughly for all quenchers .
in addition , from cyclic 
 voltammetric measurements , we propose a proton - coupled electron transfer 
 ( pcet ) by means of a stepwise mechanism from thymine to triplet bptc .
before discussing the results , 
 it is helpful to describe first all relevant calculations . as mentioned 
 earlier , two kinetic problems arise when analyzing the data obtained 
 from time - resolved laser flash photolysis : ( i ) the overlap in the 
 decay of the excited triplet states and the pseudo - first - order growth 
 of the corresponding radicals and ( ii ) 
 the overlap in the decay of the excited triplet states and the pseudo - first - order 
 growth of the corresponding radicals and their simultaneous second - order 
 decay . to deal with these kinetics , we 
 applied two alternative approaches . according to the low concentrations 
 of bptc ( 1  10 mol l ) used , triplet  triplet annihilation is negligible .
the interaction 
 of triplet bptc with quencher is a pseudo - first - order reaction .
thereupon , 
 the observed quenching rate constant , kqobs , is obtained 
 by monitoring the triplet decay of bptc at fixed wavelengths , applying 
 the stern 
volmer relation , eq 1a or eq 1b:1a1bhere , kd is the 
 decay rate constant of triplet bptc ; kqobs is the observed 
 quenching rate constant .
0 = ( 1/kd ) is the lifetime of triplet bptc in the absence 
 of quencher ;  = ( 1/kobs ) is actual lifetime of triplet bptc in the presence of quencher . 

reaction scheme 1 illustrates the two kinetic problems , where k1 is the first - order growth of radicals,2and k2 is the 
 second - order rate of the disappearance of radicals : ( i ) if the 
 system shows a combination of parallel reactions , a decay process 
 and the first - order growth of radicals ( pathway i , scheme 1 , denoted here as model i ) , the following equation 
 applies:3where a , a , and a are the absorbances 
 at time 0 , t , and infinity , respectively .
using kobs obtained from eq 3 and employing eq 1b with various concentrations 
 of the quencher gives values of kqobs .
( ii ) if the system shows a combined decay process , 
 a first - order growth of radicals and an overlap by their simultaneous 
 second - order decay ( pathway ii , scheme 1 , denoted 
 here as model ii ) , the time - resolved absorbance for the system is 
 expressed by eq 44and56thus , taking k1 obtained from the simulation , eqs 46 , and applying eq 2 with various 
 concentrations of the quencher gives the value of kqobs .
the observed quenching rate constant is expressed according to the 
 suggestion of yurkovskaya et al .
, with 
 the assumption that a polyprotic acid symbolized by hna , initial concentration c0 can undergo n - proton dissociation in water and 
 form the corresponding base conjugates .
each step has its own dissociation 
 constant , kaj , j = 1 , .. , n. the fraction of each species 
 is given by eq 7,7with 0  j  k ; 0  k  n ; ka0 = 1 . for example , for triplet 

bptc , pka1 * = 2.1 and pka2 * = 4.7 ( see the text ) , whereas for thymine , pka = 9.9 .
the molar fractions , 
 depicted in figure 1 , of all species in the 
 solution are , hence , calculated by eq 7 .
the 
 ph - dependence of kqobs for the reaction of triplet bptc with thymine 
 is then divided into four regions on the basis of their pka values and molar fractions : each pair of reactants is characterized by 
 the so - called intrinsic quenching rate constant kqi ( i = 1 ... 4 ) , and 
 the kqobs can be treated as a summation of kqi multiplied by the molar fraction of the 
 corresponding species according to eq 8 .
further 
 details of this equation can be found in the supporting 
 information.8finally , a multiple regression employed on 
 eq 8 with known parameters ( kqobs , kaj , kaj * , [ h ] ) estimates the intrinsic values , kqi .
similar expressions of the ph - dependence of the 
 observed quenching rate constants were applied for photochemical reactions 
 of bptc with the other quenchers .
absorption spectra of bptc recorded 
 in the absence and in the presence of thymine are shown in figure 2 .
it is likely that there is no association between 
 bptc and thymine in the ground state , since the spectrum of the mixture 
 of both shows an addition of their separate spectra .
similar behaviors 
 are observed with the other quenchers at different ph 's . by 
 changing the ph of the solution , it is possible to control which main 
 reactant species are present in the solution .
vis 
 spectroscopic measurements , we determined that bptc has two pka values : pka1 = 
 3.20  0.10 and pka2 = 5.12  
 0.10 .
the fact that only two pka values are observed implies that there are 
 two deprotonation steps of bptc characterized by two - proton dissociation 
 for each .
base equilibria:9a9bon the basis of the pka values and contribution of species ( figure 1 , dashed lines ) , it is unambiguous that eq 9a predominates at ph > 3.2 , whereas eq 9b dominates at ph > 5.1 .
absorption spectra of 
 bptc ( 4  10 mol l ) without 
 and with thymine ( 1.5  10 mol l ) in water at ph = 2.0 .
deprotonated forms of bptc show absorption spectra 
 at longer wavelengths compared with its neutral state .
bptch4 is found to have an absorption maximum at 261 nm ( at ph = 2.1 ) , 
 whereas the spectrum of bptch2 is shifted 
 to 269.5 nm ( ph = 4.1 ) .
laser flash photolysis 
 of bptc ( 1  10 mol l ) in aqueous solutions at ph = 2.0 and 12.0 gives spectra as figure 3 .
triplet bptc 
 shows a strong absorption peak at max = 590 nm in 
 water at ph = 2.0 , shifting to max = 550 nm at ph 
 = 12.0 .
because of these results , we have chosen 590 and 550 nm as 
 observation wavelengths for kinetic traces of all quenching experiments 
 in the acidic media and in the basic media , respectively .
the decay 
 of triplet bptc , observed at both wavelengths max = 590 and 550 nm in water follows a first - order kinetics with kd = ( 57 )  10 s , in agreement with reported values ( 7.1  10 and 6.8  10 s ) .
transient absorption spectra of bptc ( 1  10 mol l ) in water , monitored by transient absorbances 
 between 360 and 690 nm obtained right after laser flash .
because two pka values are observed in the ground state , it is expected 
 that bptc also behaves in the same way in the excited triplet state . 

the ph - dependence 
 of the absorbance ratio at 590 and 550 nm obtained immediately after 
 laser flash is shown in figure 4 .
the pka * values for triplet bptc are determined from the turning points of 
 this t1  tn absorbance titration curve as pka1 * = 2.1  
 0.2 and pka2 * = 4.7  0.2 .
this indicates that triplet 
 bptc is a stronger acid in comparison with its ground - state one .
triplet 
 absorbance titration curve of 1  10 mol 
 l bptc in water , monitored by the ratio of transient 
 absorbances at 590 and 550 nm , obtained right after laser flash .
it is realized in figure 1 that bptch4 exits mainly in the solution 
 of ph < 2.1 , whereas its dianionic bptch2 dominates within 2.1 < ph < 4.7 .
note 
 here that at ph = 3.2 , for instance , the light - absorbing species could 
 be both neutral bptch4 and dianionic bptch2 , which then generate their corresponding triplet 
 states . however , 308 nm irradiation of bptc solutions at ph = 3.2 
 results in the main species bptch2 because of the rapid acid  base equilibrium eq 10.10otherwise , at ph > 4.7 
 the species bptc is mainly present 
 because of the rapid equilibrium eq 11 although 
 bptch2 could be initially excited ( e.g. , 
 at ph = 5.1).11 as already pointed 
 out , all experimental traces for triplet bptc decay were monitored 
 at obs = 590 nm in acid solutions and at obs = 550 nm in alkaline solutions , respectively .
all experimental 
 data for the measured quenching rate constants were fitted with either 
 model i or model ii .
triplet bptc and thymine can exist 
 in either neutral or deprotonated forms , depending on the ph of the 
 solution ( see table 1 ) . in the presence of 
 thymine
, the lifetime of triplet bptc is decreased , and the long - lived 
 bptc ketyl radical anion ( bptc ) appears , 
 max = 630 nm ( see below ) .
the formation of the ketyl 
 radical anion implies that the photochemical primary step is an electron 
 transfer . in the range of ph
= 2.010.0 , we found that the 
 transient kinetics for this system correspond to model ii .
data analysis 
 for this system by model ii in the solution of ph = 6.4 is presented 
 in figure 5 .
it can be seen that the 
 total radical absorbance is high ( aradical/atriplet0 = 
 0.29 ) with [ thy ] = 6  10 mol l ( figure 5 ) as a result of the high radical 
 quantum yield of bptc at ph = 6.4 .
plotting k1 vs [ thy ] at ph = 6.4 ( figure 5 , inset ) gives a good linearity and zero intercept , showing 
 that the model accurately describes the kinetic behavior .
the spectrum 
 of bptc over the range of 610660 
 nm was obtained by laser flash irradiation of the solution of bptc 
 ( 1.0  10 mol l ) and 
 thymine ( 1.5  10 mol l ) ( figure 6 ) .
bptc absorbs at max = 630 nm , and the decay monitored 
 at obs = 630 nm obeys a second - order kinetics ( k2/bptc = 3  10 cm s ) .
in addition , 
 inbar et al . reported that the pinacol 
 yield of the 4-bc + 4-bc reaction is minor .
these lead to the postulation that after an initial 
 electron transfer followed by proton transfer , ketyl radicals ( bptch ) 
 are formed , which dissociate into free radical ions and thereafter 
 randomly encounter to undergo the pinacol reaction ( bptch + 
 bptch ) .
decays ( obs = 590 nm ) of triplet bptc 
 ( 1  10 mol l ) + thy 
 ( concentration increases from top to bottom : 01.5 mm ) in water 
 at ph = 6.4 .
bptc ketyl radical anion spectrum in the presence of thymine 
 ( 1.5  10 mol l ) at ph 
 = 2.0 .
inset : decay profile ( obs = 630 nm ) of bptc , which forms pinacol after the fast protonation 
 of bptc to bptch. on the other hand , the formation of bptc could not be observed at ph = 12.0 , even at high concentrations 
 of thymine ( [ thy ] = 1.5  10 mol l ) .
enol tautomerism of thy ( scheme 2 ) , which significantly 
 changes the chemical properties of thy .
in addition , 
 it is possible that the proton transfer from the oxidized thymine 
 is strongly hindered at ph = 12.0 and the electron reverts back to 
 the thymine molecule instead of remaining on the bptc .
therefore , 
 no quenching reaction of triplet bptc by thy occurs 
 ( i.e. , in eq 8 , kq4 = 0 ) .
the ph - dependence of the observed quenching 
 rate constants is presented in figure 7 .
although 
 the quenching reaction follows a complex reaction system that shows 
 multiple ( de)protonation equilibria in the ground and excited states , 
 the ph - dependent kqobs is adequately established .
figure 7 depicts a continuous downhill and flat behavior 
 of kqobs . the decrease in kqobs with ph
is characterized by 
 two changes ( at ph  2.1 and 4.7 ) which coincide with the two - deprotonation 
 stages of triplet bptc .
moreover , kqobs reaches plateaus in the regions , that are attributed to the equilibria of the 
 corresponding reactant pairs .
note that in the range of ph < pka , thymine exists in the neutral form ( figure 1 , inset ) and triplet bptc shows two steps of deprotonation 
 ( pka1 * = 2.1 and pka2 * = 4.7 ) .
therefore , it allows the prediction 
 of the chemical reactivity between triplet bptc and thymine in the 
 orderperhaps , for a more negative net charge , the 
 reduction is more difficult and the quenching rate constant becomes 
 lower .
kq5 = 
 0 implies that the deprotonation of the deoxyribose group of dthy 
 do not much influence the overall quenching rate constant .
ph - dependence of the observed quenching rate constant for the reaction 
 of triplet bptc with thymine .
the solid line is the simulation according 
 to eq 8 with kq4 = 0 .
the decay of triplet bptc in the presence of dthy obeys model i. the 
 appearance of the long - lived transient ( bptc ) at obs = 630 nm is observed , which then decays 
 according to a second - order kinetics on the milliseconds time scale . 

this is interpreted by an electron transfer mechanism from dthy to 
 triplet bptc , forming the radical pair followed by fast escape of 
 these radical ions from the charge - transfer successor complex .
thereafter , 
 depending on ph of the solution , the pinacol reactions , bptch 
 + bptch , bptch + bptc , or bptc+bptc occur . note 
 that unlike thymine , a keto 
enol tautomerism is no longer achieved 
 due to the deoxyribose group of thymidine . the ph - dependence 
 of kqobs
can be expressed by eq 12 and is 
 presented in figure 8 ( see tables 1 and 2 for parameters).12the main reactions at each ph region and their 
 corresponding quenching rate constants , kqi , obtained by the best fit ( solid line , figure 8) , are summarized in table 1 .
note also that kq5 = 0 implies that 
 the deprotonation of the deoxyribose group ( pka = 12.9 ) of dthy does not much influence the overall quenching 
 rate constant .
therefore , a similar explanation , such as the quenching 
 reaction by thymine for ph < 10 , is given for the variation of kqobs with ph .
a further decrease in kqobs at ph > 10 is caused by 
 the shift of the dthyh2 equilibrium toward its anionic form .
an effect 
 of coulombic repulsion may hinder the encounter of bptc and result in dthyh ; hence , the 
 quenching rate constants get smaller . the reactive species existing 
 in solution at different ph 's are shown in table 1 .
for the whole range of ph = 2.012.0 , we found that 
 the decay of triplet bptc in the presence of ade follows model i with 
 the formation of bptc .
the appearance of 
 transient bptc leads to the conclusion 
 that an electron transfer reaction from ade to triplet bptc is followed 
 by fast diffusion of the reactants .
the ph - dependence of kqobs is described by eq 13 and is presented in 
 figure 9 ( see tables 1 and 2 for parameters).13it is seen that kq1 = 0 , which implies no quenching reaction of triplet bptc by adeh2 under strong acidic conditions ( ph < 2 ) .
the increase in kqobs up to ph  2.8 can be explained 
 by a shift of the triplet bptc equilibrium toward its dianionic form , eq 10 .
the main reactant 
 pair within 2.1 < ph < 4.2 is bptch2 + adeh2 .
thus , the maximum 
 quenching rate constant here can be attributed to the molecular net 
 charge of the reactants ( coulombic interaction ) .
the further decrease 
 in kqobs at ph  2.85 is caused by a drop in the 
 adeh2 concentration following the equilibrium 
 as eq 14:14at ph > 5 , an explanation similar to that 
 of the quenching reaction by thymidine for the change of kqobs with ph 
 is used .
ph - dependence of the observed quenching rate constant for the reaction 
 of triplet bptc with dthy .
ph - dependence of the observed quenching rate constant 
 for the reaction of triplet bptc with dade .
the decay profile and kinetic 
 analysis for the quenching of triplet bptc by dade at ph = 4.0 is 
 shown in figure 10 .
it can be seen that the 
 total radical absorbance increases slightly with the concentration 
 of dade .
kinetic treatment by model i within the range of ph = 2.012.0 
 is applied .
together with the observation of bptc at obs = 630 nm , it leads to the conclusion that 
 electron transfer from dade to triplet bptc occurs , followed by a 
 fast diffusion step .
the ph - dependence of kqobs is expressed 
 by eq 13 ( see tables 1 and 2 for parameters ) .
values obtained from 
 the best fit ( solid line , figure 11 ) are summarized 
 in table 1 .
once again , an explanation similar 
 to that of the quenching reaction by adenine for the variation of kqobs with ph is used .
decay profiles ( obs = 590 nm ) for the 
 bptc + dade system in water at ph = 4.0 .
one question remains 
 that is related to the quenching reactions of triplet bptc by thymine 
 at ph = 2.010 : is electron transfer followed by proton transfer ? 
 to check the possibility , we have investigated the ph - dependence of 
 the oxidation potential of thymine by cyclic voltammetry .
whatever 
 the reaction mechanism may be , a global nernst equation can be applied 
 for the thermodynamics of the redox reaction:15here , e is the electrode 
 potential ; eap0,ox / red is the apparent standard 
 oxidation potential of the corresponding redox couple ; and [ox ] 
 and [red ] are the total concentrations of the oxidized and 
 reduced species in solution , respectively .
therefore , the ph - dependence 
 of eap0 for the oxidation of thymine can be described by eq 16 ( see the supporting information ) , as shown in figure 12.16here , pka , thyh = 3.2 is the dissociation constant 
 of thymine radical cation .
the apparent 
 oxidation potential vs ag / agcl of thymine ( 1 mm ) as a function of 
 ph . solid line : linear fitting with a slope of 55.2 mv / ph . between ph
= 28 , eap0 shows a linear 
 variation with ph , together with the characteristic slope of approximately 
 59 mv per ph unit .
this indicates a one - proton - coupled one - electron 
 transfer during the oxidation of thymine .
this observation and the above kinetic analysis of the quenching 
 reaction by thymine suggest that the possible pathway of the quenching 
 reaction of triplet bptc by thymine at ph around 28 is initially 
 an electron transfer , followed by proton transfer ( scheme 3 ) . at ph around 810 ,
the quenching of the triplet state of 3,3,4,4-benzophenone 
 tetracarboxylic acid by dna bases in aqueous solution was investigated 
 using time - resolved laser flash photolysis .
the observation of the 
 bptc ketyl radical anion ( max = 630 nm ) confirms 
 that the primary photochemical step is electron transfer .
although 
 the forms of the triplet bptc and quenchers depend on the ph of the 
 solution , we have been able to establish a relationship of the quenching 
 rate constant of each pair of reactants with the overall quenching 
 rate constant .
particularly , the ph - dependence of the apparent 
 standard potential of thymine in aqueous solution was investigated 
 by cyclic voltammetry . from the kinetic analysis of the quenching 
 reactions ,
the ph - dependent oxidation potential of thymine indicates 
 that proton - coupled electron transfer by means of a stepwise mechanism 
 from thymine to triplet bptc in the solutions of ph  28 
 proceeds . in the strong basic solution ( ph = 12.0 ) , no quenching reaction 
 between triplet bptc and thymine takes place , whereas it is observed 
 for thymidine .
the abbreviations used in this article can 
 be found in the section results and discussion .
y. lin , chemistry department , chinese academy 
 of science , beijing and was used without further purification .
dna 
 bases adenine ( 99% ) , adenosine ( 99% ) , thymine ( 97% ) , and thymidine 
 ( 99% ) , purchased from alfa - aesar , were also used as received .
all experiments were carried out at room temperature in buffered 
 aqueous solutions . the buffer solutions ( 0.01 mol l ) employed , covering the ph range from 3.0 to 11.0 with ( a ) hcl 
kh2po4 , ph = 3.05.0 ; ( b ) kh2po4na2hpo4 , ph = 5.09.0 ; 
 ( c ) na2hpo4naoh , ph = 9.011.0 , 
 ph = 2.0 and ph = 12.0 , were adjusted with hcl and naoh , respectively . 

transient absorption spectra 
 were obtained by time - resolved laser spectroscopy using a lambda physik 
 lpx-120 xecl excimer laser ( 308 nm , pulse energy up to 100 mj , pulse 
 width 10 ns ) .
the monitoring system included a 150 w xenon lamp ; a 
 hamamatsu r928 photomultiplier tube ; a obb / pti monochromator , model 
 101/102 ; and a digital storage oscilloscope , 9410a lecroy .
the irradiation 
 was carried out in a 1  1 cm rectangular quartz 
 cell .
all solutions were treated identically and deoxygenated by bubbling 
 with high - purity argon for 15 min .
cyclic voltammetric measurements 
 were performed with an autolab - pges aut 73227 potentiostat ( metrohm ) . 

a three - electrode cell configuration was used : a pt counter electrode , 
 a ag / agcl reference electrode , and a glassy carbon working electrode 
 ( 0.03 cm area ) .
the working electrode was polished with 
 diamond paste in water after each single scan to remove possible follow - up 
 products of the oxidative process on the electrode surface .
the background 
 current was always subtracted from the current response for further 
 calculation . for the triplet state .
 the kinetics of triplet state quenching of 3,3,4,4-benzophenone 
 tetracarboxylic acid ( bptc ) by dna bases adenine , adenosine , thymine , 
 and thymidine has been investigated in aqueous solution using time - resolved 
 laser flash photolysis . 
 the observation of the bptc ketyl radical 
 anion at max = 630 nm indicates that one electron 
 transfer is involved in the quenching reactions . 
 the ph - dependence 
 of the quenching rate constants is measured in detail . as a result 
 , 
 the chemical reactivity of the reactants is assigned . 
 the bimolecular 
 rate constants of the quenching reactions between triplet bptc and 
 adenine , adenosine , thymine , and thymidine are kq = 2.3  109 ( 4.7 < ph < 9.9 ) , kq = 4.0  109 ( 3.5 < ph < 
 4.7 ) , kq = 1.0  109 ( 4.7 
 < ph < 9.9 ) , and kq = 4.0  
 108 m1 s1 ( 4.7 < 
 
 ph < 9.8 ) , respectively 
 . moreover , it reveals that in strong basic 
 medium ( ph = 12.0 ) a keto  
 enol tautomerism of thymine inhibits 
 its reaction with triplet bptc . 
 such a behavior is not possible for 
 thymidine because of its deoxyribose group . 
 in addition , the ph - dependence 
 of the apparent electrochemical standard potential of thymine in aqueous 
 solution was investigated by cyclic voltammetry . 
 the e/ph  59 mv / ph result is characteristic 
 of proton - coupled electron transfer . 
 this behavior , together with 
 the kinetic analysis , leads to the conclusion that the quenching reactions 
 between triplet bptc and thymine involve one proton - coupled electron 
 transfer . 
Introduction
Results and Discussion
Conclusion
Experimental Section