redox properties and prooxidant cytotoxicity of benzofuroxans-2009-chemija_vol_20_p109-115

7/31/2019 Redox Properties and Prooxidant Cytotoxicity of Benzofuroxans-2009-Chemija_VOL_20_p109-115

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chemija. 2009. vo. 20. No. 2. P. 109115 ltuos oks kd, 2009 ltuos oks kdos dyk, 2009

Redox properties and prooxidant cytotoxicity of

benzofuroxans: a comparison with nitrobenzenes

Jonas arlauskas1,

Val Miliukien1,

ilvinas Anuseviius1,

Lina Miseviien1,

Kastis Kriktopaitis

1

,Aura Nemeikaitnien2,

Irena Vitnien3,

Narimantas nas1, 3*

1 Institute of Biochemistry,Mokslinink 12,

LT-08662 Vilnius, Lithuania

2 Institute of Immunology

of Vilnius University,Molt pl. 29, LT-08409 Vilnius,

Lithuania

3 Vilnius Pedagogical University,

Student 39, LT-08106 Vilnius,

Lithuania

* corrspondng utor. e-: [email protected]

Te presence o =N(O)O- moiety in benzouroxans may coner them the electron-ac-

cepting properties similar to those o aromatic nitrocompounds or N-oxides which areused as bioreductively activated antitumour and / or cytotoxic agents. However, the redox

properties o benzouroxans have not been characterized so ar. We have studied the redox

properties o eight benzouroxans, determining the heats o ormation o their ree radicals

(H(e)), the peak-potentials o their electrochemical reduction in the aqueous medium

at pH 7.0 (Ep, 7

), and their reactivity towards a single-electron transerring avoenzyme,

erredoxin : NADP+ reductase (FNR). We ound that the reactivity o benzouroxans to-

wards FNR in general is similar to that o (poly)nitrobenzenes possessing analogous valueso H(e), and the benzouroxans are reduced in a single-electron way. However, the E

p, 7

o the irreversible electrochemical reduction o benzouroxans strongly deviated rom the

dependence o H(e) o nitrobenzenes on their Ep, 7

. In general, the cytotoxicity o ben-

zouroxans in bovine leukemia virus-transormed lamb kidney broblasts (line FLK) is

similar to that o nitroaromatic compounds possessing the analogous values o H(e ),

whereas their cytotoxicity in mice splenocytes considerably diers.Tis may be caused byother parallel cytotoxicity mechanisms, e. g. reactions o benzouroxans with SH groups.

In this work, or the rst time we identied benzouroxans as redox active compounds. Pre-

liminarily, one may conclude that the redox properties and cytotoxicity o benzouroxans

bear certain similarities (reactions with single-electron-transerring avoenzymes) and

dierences (electrochemical reduction, cytotoxicity) rom nitroaromatic compounds.

Key words: benzouroxans, nitrobenzenes, electrochemical reduction, electron-acceptingproperties, erredoxin : NADP+ reductase, ree radicals, oxidative stress, cytotoxicity

Abbreviations: Ar-NO2, aromatic nitrocompound; Ar>NO, aromatic N-oxide; Ar-NO

2.,

aromatic nitroanion-radical; H(e), heat o ormation o ree radical; FNR, erredoxin:

NADP+ reductase; kcat

/ Km

, bimolecular rate constant in enzymatic steady-state reactions;

E17, single-electron reduction potential at pH 7.0; E

p, 7, the reduction peak-potential in cyclic

voltammetry; O2, superoxide; NHE, normal hydrogen electrode; cL

50, compound concent-

ration or 50% cell death.

INTRODUCTION

One o the main mammalian cell cytotoxicity mechanisms onitroaromatic and nitrohetero-cyclic compounds (Ar-NO

2)

is their redox cycling, initiated by the avoenzyme-catalyzed

single-electron reduction. Te ormation o anion-radicals o

nitroaromatics (Ar-NO2.) is ollowed by their reoxidation by

molecular oxygen, which yields the parent Ar-NO2

and su-

peroxide (O2), the latter subsequently yielding other activated

oxygen species. Tis results in the oxidative stress-type cyto-

toxicity, i. e. peroxidation o lipids and the oxidative damage

to proteins and DNA [14]. It is assumed that i the oxidative

stress is the main actor o cytotoxicity o ArNO2, then their

cL50

(compound concentration or 50% cell death) shoulddecrease with an increase in their single-electron reduction

potential (E17) with the coecient log cL

50/ E1

7~ 10 V1

[14]. Te E17

values o nitroaromatic compounds deter-

mined by means o pulse-radiolysis, typically range rom

0.20 V to 0.50 V vs. normal hydrogen electrode (NHE)

[1, 5, 6]. AromaticN-oxides (Ar > NO) represent another


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Jonas arlauskas, Val Miliukien, ilvinas Anuseviius, Lina Miseviien, Kastis Kriktopaitis et al.110

important group o bioreductively activated compounds, in-

cluding the antitumour agent tirapazamine and its analogues

[79]. However, they are less ecient electron acceptors than

nitroaromatics, because their E17

values range rom 0.38 V

to 0.80 V.

Benzouroxans possess =N(

O)O- moiety (Fig. 1)which shares a similarity with both nitro- and N-oxidegroups. Tus, one may expect that benzouroxans may also

undergo a single-electron enzymatic reduction, and that

their electron-accepting potency may be similar to that o

nitroaromatic compounds or aromaticN-oxides. Tese datamay be o certain importance in medicinal chemistry and

toxicology because benzouroxans are used as activators o

guanylate cyclase and inhibitors o monoamino oxidases[10, 11]. Besides, benzouroxans comprise a new group o

explosive compounds [12]. Tis implies the possible ecotoxi-

cological problems due to the environmental contamination

by explosive residues. However, to our best knowledge, the

redox properties o benzouroxans have not been reported

so ar. In this paper, we present data on the electron-accept-

ing properties o several benzouroxans and their mamma-lian cell cytotoxicity, which are compared to the analogous

properties o nitrobenzenes.

EXPERIMENTAL

Te synthesis o benzouroxans was perormed according to

the established methods (Scheme). Mono-substituted deriva-

tives o benzouroxan (compounds 1, 2, 4 (Fig. 1)) were ob-

tained by a direct one-step oxidation o corresponding o-nit-roanilines by hypochlorite in basic medium (pH 8.09.5)[13]. Di-substituted benzouroxans (compounds 3, 5, 6, 7

(Fig. 1)) were synthesized rom the corresponding 1,2-dini-

trobenzene derivatives by a modied two-step method [14].

2-Nitrophenylazides were obtained by the nucleophilic sub-

stitution reaction and underwent cyclization during their

boiling with acetic acid [15]. Benzotriuroxan (compound 8

(Fig. 1)) was synthesized by the thermocyclization o 1,3,5-triazido-2,4,6-trinitrobenzene in boiled propionic acid as

previously described [16]. Te structure o the compounds

was identied by means o elemental analysis, NMR, and IR.

2,4,6-rinitrotoluene and 2,4,6-trinitrohenyl-N-methylnit-ramine (tetryl) were synthesized as described [4].

Voltammetric studies o benzouroxans and model

(poly)nitrobenzenes (0.51.0 mM) were perormed using

a Parstat 2273 potentiostate (Princeton Applied Research)under anaerobic conditions (purging the solution with ar-

gon or 20 min) at pH 7.0 (0.05 M K-phosphate and 0.1 M

KCl) and 25 C. A rod o glassy carbon (BAS, 2.0 mm diame-

ter) polished with a corundum abrasive served as a work-

ing electrode. A saturated Ag / AgCl electrode (+0.205 V vs.

NHE) was used as a reerence electrode. Te voltammetric

reduction peak-potentials o compounds studied in thiswork (E

p, 7) are presented with respect to this electrode. A

Pt electrode (apparent surace area 56 mm2) was used as an

auxiliary electrode.Te kinetic measurements were carried out spectro-

photometrically using a Hitachi-557 spectrophotometer in

0.1 M K-phosphate buer (pH 7.0) containing 1 mM EDA

at 25 C. Ferredoxin : NADP+ reductase (FNR, EC 1.18.1.2)

rom Anabaena was prepared as described [17], and was a

generous gif o Dr. Marta Martines-Julvez and Proessor Car-

los Gomez-Moreno (Zaragoza University, Spain). Te enzymeconcentration was determined spectrophotometrically using

Scheme.The synthesis o mono- and disubstituted-benzouroxans


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111Redox properties and prooxidant cytotoxicity o benzouroxans: a comparison with nitrobenzenes

459

= 9.4 mM1 cm1. Te rates o FNR-catalyzed oxidation o100 M NADPH by benzouroxans and nitroaromatic com-

pounds, corrected or the intrinsic NADPH-oxidase activity

o FNR, were determined according to 340

= 6.2 mM1 cm1.

Te catalytic constant (kcat

) and the bimolecular rate constant

(kcat

/ Km

) o the reduction o compounds were calculated

rom the LineweaverBurk plots. kcat

is the number o NA-DPH molecules oxidized by the single active center o FNR

per second. In separate experiments, cytochrome c (50 M)was added to the reaction mixture. Its reduction was moni-

tored according to 550

= 20 mM1 cm1.

Te heats o ormation o ree radicals o benzouroxans

and nitroaromatics (H(e)) were calculated using PC Spar-tan 04 Pro (Waveunction, Inc.) according to semiempirical

AM1 and PM3 methods, as described previously [18].

Te primary mice splenocytes were obtained rom

BALB / c mice as described previously [19]. Tese experi-

ments were approved by the Lithuanian Veterinary and Food

Service (License No. 0171, 2007). Te splenocytes (106/ml)were suspended in RPMI 1640 medium with 5% etal bovine

serum, penicillin (100 U/ml), and streptomycin (100 g/ml);their viability was determined afer 24 h o incubating the

splenocytes with the examined compounds according to the

rypan blue exclusion test. Te compounds were dissolved

in DMSO whose nal concentration in the medium (0.6%)

did not aect the splenocyte viability. Te culture o bovine

leukemia virus-transormed lamb kidney broblasts (lineFLK) was grown and maintained in Eagles medium supple-

mented with 10% etal bovine serum and antibiotics as de-

scribed previously [20]. In the cytotoxicity experiments, cells

(2.5 104/ml) were seeded on glass slides in 5 ml asks in thepresence or in the absence o compounds, and were grown

or 24 h. Further, the slides were rinsed 34 times with phos-

phate buer saline and stained with rypan blue. Te cells

on the slides were counted under a light microscope. Te sta-

tistical analysis was perormed using Statistica (version 4.3,StatSof, 1993).

All other reagents were obtained rom Sigma, unless

otherwise specied.

RESULTS AND DISCUSSION

Because o the similarity o =N(O)O- moiety o benzou-

roxans with the nitrogroup, their properties are urther dis-

cussed in the context o redox properties o nitroaromatic

compounds which are relatively well understood. According

to the numerous studies ([4] and reerences given therein),

the reactivity o nitroaromatics in avoenzyme-catalyzed

single-electron reduction reactions, which cause the oxida-tive stress-type cytotoxicity, increases with an increase in

their single-electron reduction potential (E17) with the coe-

cient log (kcat

/ Km

)/E17

= 8.010 V1, where kcat

/ Km

is the

bimolecular reaction rate constant. Tis is in accordance with

the outer-sphere mechanisms o single-electron transer [21].I the experimentally determined E1

7values o compounds are

absent, the heats o the ormation o ree radicals (H(e))

obtained by means o quantum mechanical calculations may

be used as an approximate substitute parameter describing

their reactivity [4]. Te H(e) values o benzouroxans are

given in able 1, together with the H(e) values o several

nitrobenzenes, obtained in our previous studies [4, 18]. Our

data show that the H(e) values o benzouroxans are simi-

lar to those o nitrobenzenes (1215) and are considerably

more negative than that o nitrobenzene with E17= 0.485 V

(able 1). It means that benzouroxans studied in this

work may be better electron acceptors than nitrobenzene.

Fig. 1. The ormulae o benzouroxans studied in this work


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According to quantum mechanical calculations, the Natom

o =N(O)O- moiety o benzouroxans is the most probable

electron-accepting site, because it possessed the highest posi-

tive Mulliken charge and the highest coecients o the lowest

unoccupied molecular orbital.

We have studied the reactions o benzouroxans with amodel single-electron-transerring avoenzyme erredox-in : NADP+ reductase (FNR). Its reactivity is not specic to-

wards the particular structures o nitroaromatic oxidants, and

it is characterized by a linear dependence on their log kcat

/ Km

on E17

or, to a lesser extent, on their H(e) ([4] and reer-

ences therein). It is evident that the reactivity o benzourox-ans towards FNR is similar to the reactivity o nitroaromatics

possessing analogous H(e) values (Fig. 2). Te reduction

o the benzouroxan derivative (5) (Fig. 1) by FNR is accom-

panied by a reduction o cytochrome cadded in the separateexperiments, at a rate twoold to the NADPH oxidation rate.

Te reduction o cytochrome c is inhibited by 6065% by100 U/ml superoxide dismutase. It shows that the reactionproceeds as a single-electron transer with the ormation o

a ree radical o benzouroxan, which urther is oxidized by

oxygen and yields superoxide.

Te electrochemical reduction o most nitroaromatic

compounds proceeds as an irreversible net our-electron

transer, with the ormation o hydroxylamines [22, 23]. Te

nitrogroups in polynitroaromatic compounds are reducedsubsequently as is evidenced by the appearance o three

reduction peaks o 2,4,6-trinitrotoluene (Fig. 3) and 2,4,6-

trinitrophenyl-N-methylnitramine (able). Te subsequentreactions o the reduction products sometimes result in the

appearance o electrochemically active condensation / po-lymerisation products which undergo electrochemical oxida-

Hf (e-) (kJ/mol)

-350-300-250-200

101

102

103

104

105

106

9

16

1011

1412

13

15

85

4

7

61

32

k/K

(M

s)

cat

m

-1

-1

Fig. 2.Relationship between the H(e) (AM1) values o nitroaromatic com-

pounds (circles) and benzouroxans (triangles) and their reactivity towards

erredoxin : NADP+ reductase (kcat

/ Km

). The numbers o compounds are taken

rom Table

Fig. 3. Cyclic voltammograms (v = 50 mV/s) o electrochemical reduction o

benzouroxan (1), 2,4,6-trinitrotoluene (2), and nitrobenzene (3) at 0.5 mM

compound concentration and pH 7.0. The starting potential is 0.5 V, the poten-

tials are given vs.Ag / AgCl. Because o the electrode contamination, only the

frst scans are shown

tion at positive potentials ([22, 23], Fig. 3). Our preliminary

data show that benzouroxans are also electrochemically ir-reversibly reduced at the range o the potentials o nitroben-

zene reduction (Fig. 3, able). Te reduction peak currents o

benzouroxans and nitrobenzenes studied are controlled by

diusion because they are directly proportional to the square

root o the potential scan rate, 101000 mV/s. In general, the

voltammetric reduction peak-potentials (Ep, 7

) o nitroaro-matic compounds increase with an increase in theirE1

7values

[24]. For this reason, a linear relationship has been observed

between the E17

values and the rst Ep, 7

o nitroaromatics

studied in this work (able), although it has been poorly

expressed(r2 = 0.7845, data not shown). On the other hand,there also exists a linear relationship between the values o

E17

and H(e) o nitroaromatics [4]. Tus, it results in the

appearance o linear although poorly expressed relation-

ships between the rst Ep, 7

values o nitroaromatics and their

H(e) (r2 = 0.7193 (AM1, Fig. 4), and r2 = 0.7337 (PM3,

data not shown). However, the Ep, 7

values o benzouroxans

strongly deviate rom this dependence (Fig. 4). For this rea-son, the mechanism(s) o the electrochemical reduction o

benzouroxans and their dependence on their structure are

the subject o our orthcoming studies.

We have also preliminarily investigated the cytotoxicity

o benzouroxans in two mammalian cell l ines, primary mice

splenocytes and bovine leukemia virus-transormed lamb

embryo kidney broblasts (line FLK). In both cell lines, theconcentrations o nitroaromatic compounds causing a 50%

cell death (cL50

) increase with a decrease in their E17

with

the coecient log cL50

/ E17

~ 10 V1 [4, 19]. Analogously,

cL50

increases with an increase in H(e) o nitroaromat-

ics [4]. aken together with the cytotoxicity decrease by the

Potential, V

-1.0 -0.8 -0.6 -0.4 -0.2 0.0 0.2 0.4

-50

-40

-30

-20

-10

0

10

1

2

3

Current,A


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Hf (e-) (kJ/mol)

-400 -350 -300 -250 -200 -150

-800

-600

-400

-200

16

9

8

10

11

12

14

13

15

43

7

5

62

1

E

(mV)

p,7

Fig. 4. Relationship between the H(e) (AM1) values o nitroaromatic com-

pounds (circles) and benzouroxans (triangles) and their electrochemical reduc-

tion peak-potentials (Ep, 7

). The numbers o compounds are taken rom Table, the

line corresponds to the frst-order regression drawn through the corresponding

parameters o n itroaromatic compounds

T a b l e . The heats o ormation o ree radicals o benzouroxans and nitroaromatic compounds (H(e)), their reactivities towards the single-electron

transerring favoenzyme erredoxin : NADP+ reductase (kcat

/ Km

), their single-electron reduction potentials determined by pulse-radiolysis (E17), their reduc-

tion peak-potentials in cyclic voltammetry (Ep, 7

, 50 mV/s), and their concentrations or 50% mammalian cell death (cL50

)

No. CompoundHf(e)(kJ/mol)

kcat

/ Km

(M1 s1)E1

7

(mV)E

p, 7

cL50

(M)

AM1 PM3 FNR vs. NHE

a

vs. Ag / AgCl Splenocytes FLKBenzofuroxans:

1. Benzofuroxan 231.0 225.8 3.7 0.30 102 452 60 8.0 250 30

2. 5-Methylbenzofuroxan 228.2 222.9 2.1 0.10 103 448, 0.800 94 8.0

3.5,6-Dimethoxy-benzofuroxan

228.9 221.9 4.7 0.20 103 408, 0.810 >300 20 4.0

4. 5-Chlorobenzofuroxan 256.1 245.2 1.6 0.10 103 397 47 5.0

5.1-Oxy-5H,8H-2-oxa-1,3,5,8-

tetraaza-cyclopenta[b]naphthalene-6,7-dione

265.2 282.7 6.7 0.70 103 634, 765 19 2.0 110 18

6.6,7-Dihydro-2,5,8-trioxa-1,3-diazacyclo-penta[b]

naphthalene-1-oxide240.6 229.5 1.0 0.10 103 457, 760 75 7.0

7.

[1,3] Dioxolo [4,5 : 4,5] ben-

zo [1,2-c]-[1,2,5]oxadiazole-1-oxide 243.4 231.8 3.1 0.10 103 698 22 3.0

8. Benzotrifuroxan 364.8 414.5 1.5 0.10 104 759 70 5.0 4.6 0.7

Nitrobenzenes:

9.2,4,6-Trinitrophenyl-N-

methylnitramine383.1 369.2 1.1 0.2 105b 156

305, 485,

6256.0 1.5c 1.5 0.3d

10. 2,4,6-Trinitrotoluene 310.7 316.4 1.0 0.1 104b 253470, 650,

81010 2.0c 25 5.0d

11. p-Dinitrobenzene 273.3 281.1 3.0 0.2 104b 257 337, 805 2.5 0.4c 8.0 2.0d

12. o-Dinitrobenzene 257.3 261.0 2.2 0.2 103b 287 452,807 60 15c 30 5.0d

13. p-Nitrobenzaldehyde 222.1 223.2 8.3 0.7 103b 315 634 40 6.0c 25 12d

14. m-Dinitrobenzene 254.6 262.1 2.5 0.2 103b 345 563,805 100 20c 90 20d

15. p-Nitroacetophenone 217.1 218.1 1.0 0.1 103b 355 532 60 8.0c 166 36d

16. Nitrobenzene 167.7 172.1 58 7.0

b

485 706 1000 200

c

4370 1370

d

a From Res. [5, 6]. b, dFrom Re. [4]. c From Re. [19].

antioxidants [4, 19], these data show that the main actor ocytotoxicity o nitroaromatics in these cell lines is the oxi-

dative stress caused by their avoenzyme-catalyzed redox

cycling. We have ound that the cytotoxicity o benzourox-

an in both cell lines is, at least partly, caused by the oxidative

stress. At the concentrations causing a 38 3.0% splenocyte

death (75 M benzouroxan) or 34 2.5% FLK cell death

(300 M benzouroxan), addition o 2.0 MN,N-diphenyl-p-phenylene diamine or 300 M deserrioxamine typically

increased the cell viability to 50% (data not shown). Ingeneral, the cL

50values o several benzouroxans in FLK cells

(able) are similar to those o nitroaromatic compounds

possessing the analogous H(e) values (Fig. 5A). How-

ever, the cL50

values o benzouroxans show a much larger

deviation rom the cytotoxicity o nitroaromatics in spleno-

cytes (Fig. 5B). Tese ndings are unexpected and need ur-

ther clarication. One o the possible explanations is that

H(e) values may not quite well reect the dierences

in the electron-accepting potency o nitroaromatic com-pounds and benzouroxans in an aqueous medium. Besides,

it is also possible that some other mechanism(s) o action o

benzouroxans, e. g.,their reactions with intracellular SH


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compounds [25] may also contribute to their cytotoxicity. A

more thorough investigation o the mechanism(s) o mam-malian cell cytotoxicity o benzouroxans is the subject o

our orthcoming studies.

CONCLUSIONS

In this work, or the rst time we have identied benzouro-

xans as the redox active compounds whose bioreductive

properties may be maniested at the physiological range o

redox potentials, 0.5 V vs. NHE. Although their E17

values

have not been determined, the quantum mechanical calcu-lations have revealed that the electron-accepting potency o

benzouroxans may be similar to that o a number o (poly)

nitroaromatic compounds, including the important explo-

sive 2,4,6-trinitrotoluene. Te benzouroxans are reducedin a single-electron way by a model avoenzyme, erredox-

in : NADP+ reductase, which implies the possibility o theirsingle-electron reduction by the other avoenzymes dehy-

drogenases-electrontranserases, e. g.,NADPH cytochromeP-450 reductase or NO-synthase, which may be important in

their mammalian cell cytotoxicity. Although the mammalian

cell cytotoxicity o benzouroxans may not entirely depend

on their electron-accepting properties due to their possible

parallel reactions, our data demonstrate the role o their re-dox cycling in their cytotoxicity. Tis may serve as a guideline

or the urther studies o the redox properties and cytotoxic-

ity o benzouroxans.

ACKNOWLEDGEMENTS

Tis work has been supported in part by the Agency or Inter-

national Science and echnology Development Programmesin Lithuania (COS Action CM0603). We thank Proes-

sor Carlos Gomez-Moreno and Dr. Marta Martinez-Julvez

(Zaragoza University, Spain) or their generous gif o erre-doxin : NADP+ reductase.

Rd 16 jnury 2009aptd 27 jnury 2009

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Hf (e-) (kJ/mol)

-400 -350 -300 -250 -200 -150

1

10

100

1000

Hf (e-) (kJ/mol)

-400 -350 -300 -250 -200 -150

1

10

100

1000

16

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(M)

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Jonas arlauskas, Val Miliukien, ilvinas Anuseviius,Lina Miseviien, Kastis Kriktopaitis, Aura Nemeikait-nien, Irena Vitnien, Narimantas nas

BENZOFUROKSAN REDOKS SAVYBS IR PROOK-SIDANTINIS CITOTOKSIKUMAS PALYGINTI SU

NITROBENZENAIS

S a n t r a u k a

Funkcins =N(O)O- grups gali suteikti benzouroksanams

elektronoakceptorines savybes, panaias nitroaromatini jungini

arba j N-oksid savybes. Pastarieji junginiai danai naudojamikaip bioredukciniu bdu aktyvuojami prienavikiniai ir / arba ci-

totoksiki agentai. aiau iki iol benzouroksan redoks savybs

nebuvo itirtos. Ityrme 8 benzouroksanus, nustatydami j lais-

vj radikal susidarymo ilumas (H(e)), j ciklins voltampe-

rometrijos redukcijos porencialus vandeninje terpje (Ep, 7

) ir j

vienelektronins redukcijos greiio konstantas avininiu ermentu

erredoksin: NADP+ reduktaze (FNR). Nustatme, kad FNR redu-

kuoja benzouroksanus ir (poli)nitrobenzenus vienelektroniniu

bdu ir panaiais greiiais, priklausaniais nuo j H(e) reikms.

aiau benzouroksan Ep, 7

ir H(e) nekoreliavo su nitrobenzen

H(e) ir Ep, 7

tarpusavio priklausomybe. Benzouroksan citotok-

sikumas galvij leukemijos virusu transormuotuose riuko inkst

broblastuose (FLK linija) buvo artimas nitroaromatini jungini

su atitinkamu H(e) citotoksikumui, taiau peli splenocituose

i jungini citotoksikumas smarkiai skyrsi. ai gali bti susieta

su kitais lygiagreiais benzouroksan citotoksikumo mechaniz-

mais, pvz., j reakcijomis su SH grupmis.

redox properties and prooxidant cytotoxicity of benzofuroxans-2009-chemija_vol_20_p109-115

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