Catalysis of singlet oxygen production in the reaction of hydrogen peroxide and hypochlorous acid by 1,4-diazabicyclo[2.2.2]octane (DABCO)

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

Vol. 134, No. 2, 1986

Pages 777-782

January 29, 1986

CATALYSIS OF SINGLET OXYGEN PRODUCTION IN THE REACTION OF HYDROGEN PEROXIDE AND HYPOCHLOROUS ACID BY I~4-DIAZABICYCLO[2.2.2]OCTANE (DABCO) Jeffrey

R. Kanofsky

Medical S e r v i c e , Edward Hines, J r . Veterans A d m i n i s t r a t i o n H o s p i t a l , Hines, IL 60141 and Department of M e d i c i n e , Loyola U n i v e r s i t y S t r i t c h School of Medicine, Maywood, IL 60153 Received

October

24,

1985

SUMMARY: The k i n e t i c s of the s i n g l e t oxygen p r o d u c t i o n in the hydrogen p e r o x i d e plus hypochlorous acid r e a c t i o n were s t u d i e d by measuring the time course of the s i n g l e t oxygen emission at 1268 nm. The a d d i t i o n of 1 , 4 - d i a z a b i c y c l o [ 2 . 2 . 2 ] o c t a n e (DABCO) increased the peak i n t e n s i t y of the chemiluminescence, but decreased i t s d u r a t i o n . The increased r a t e of s i n g l e t oxygen p r o d u c t i o n l i k e l y accounts f o r the enhancement of s i n g l e t oxygen dimol emission r e p o r t e d in 1976 by Deneke and K r i n s k y (J. Am. Chem. Soc. 98, 3041-3042). This phenomenon was not seen when s i n g l e t ox-ygen was generated w i t h the r e a c t i o n of hypobromous acid and hydrogen p e r o x i d e . Thus, the enhancement of red chemiluminescence by DABCO should not be regarded as a general t e s t f o r the p r o d u c t i o n of s i n g l e t oxygen in complex biochemical systems. ©1986AcademicPress, Inc. In 1976, Deneke and K r i n s k y the L02 dimol This r e s u l t ,

emission

in the r e a c t i o n

which has s u b s e q u e n t l y

been d i f f i c u l t DABCO reduces

reported

to r a t i o n a l i z e z02 dimol

with

emission

DABCO quenches the chemical

that

DABCO enhanced

of H202 with

been confirmed

HOCI ( 1 ) . (2,3),

other observations

in the gas phase (4)

reactivity

that and t h a t

of 102 in s o l u t i o n

(5-10).

The study of Deneke and K r i n s k y

tigators

to use the enhancement of red chemiluminescence

DABCO as a s p e c i f i c systems (11-12)

test

for

has

has caused some i n v e s by

z02 in complex biochemical

even though DABCO has been shown to enhance ~02

A b b r e v i a t i o n s used: 1,4-diazabicyclo[2.2.2]octane, DABCO; hydrogen p e r o x i d e , H202; hypobromous a c i d , HOBr; hypochlorous a c i d , HOCl; n e g a t i v e l o g a r i t h m of the deuterium ion c o n c e n t r a t i o n , p2H; deuterium o x i d e , 2H20; sodium bromide, NaBr.

777

0006-291X/86 $1.50 Copyright © 1986 by Academic Press, Inc. All rights of reproduction in any form reserved.

Vol. 134, No. 2, 1986

dimol

emission

report

that

reaction sible

only

in

the

reaction

DABCO i n c r e a s e s

of

for

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

H202 w i t h

the

rate

HOCI and t h a t

t h e enhanced

z02 dimol

by Deneke and K r i n s k y

of

H202 w i t h

of

HOCI.

I now

102 p r o d u c t i o n

this

catalysis

is

chemiluminescence

in

the

respon-

reported

(1,2).

MATERIALS AND METHODS S t u d i e s o f z02 monomol c h e m i l u m i n e s c e n c e in t h e H20e + HOCI r e a c t i o n and t h e H202 + HOBr r e a c t i o n were c a r r i e d o u t with a highly sensitive chemiluminescence spectrometer (13-16). For t h e s e e x p e r i m e n t s , t h e a n a l o g o u t p u t o f t h e s p e c t r o m e t e r was c o u p l e d to an IBM PC computer w i t h a DASCON-1 a n a l o g to digital interface board ( M e t r o b y t e C o r p . , S t o u g h t o n , MA). The emission intensity was sampled at 4 Hz and t h e IBM computer was used to c a l c u l a t e the integral of the emission intensity o v e r t h e r e a c t i o n p e r i o d and t h e i n t e g r a l of the square o f t h e emission intensity over the reaction period. The s o u r c e s , methods o f p u r i f i c a t i o n and methods o f assay f o r H202, HOCI and HOBr were r e p o r t e d p r e v i o u s l y (13-16). A l a n i n e , 2H20, 99.8%, and t a u r i n e were o b t a i n e d from Sigma Chemical Co. DABCO was o b t a i n e d from A l d r i c h Chemical Co. T r i m e t h y l a m i n e was a p r o d u c t o f Eastman Kodak Co. Other i n o r g a n i c c h e m i c a l s were r e a g e n t g r a d e . The sodium p h o s p h a t e b u f f e r used was made w i t h 2H20 and a d j u s t e d to an a p p a r e n t pH of 7.4 with a glass electrode. T h i s c o r r e s p o n d s to a p2H o f 7.0 ( 1 7 ) . For s t u d i e s o f t h e H202 + HOCI r e a c t i o n , a stock s o l u t i o n o f DABCO was p r e p a r e d in b u f f e r and t h e p2H a d j u s t e d to 7.0 w i t h p h o s p h o r i c a c i d . A l l e x p e r i m e n t s were done in t r i p l i c a t e and are r e p o r t e d as the mean ± t h e s t a n d a r d e r r o r . Hypohalous a c i d in 1.5 ml of b u f f e r was p l a c e d in t h e s p e c t r o m e t e r . The r e a c t i o n was then i n i t i a t e d by t h e r a p i d i n j e c t i o n o f an a d d i t i o n a l 1.5 ml of b u f f e r c o n t a i n i n g H202. For most e x p e r i m e n t s , an amine u s u a l l y DABCO was added to t h e H=O= s o l u t i o n p r i o r to i t s injection.

RESULTS AND DISCUSSION The k i n e t i c s

of

=02 p r o d u c t i o n

H202 + HOBr r e a c t i o n s

were s t u d i e d

course

z02 e m i s s i o n

of

t h e monomol

intensity tration

at of

1268 nm is

to

concentrations order

to

the

square

of

(150 ~M, each)

slow down t h e

rate

t h e Hz02 + HOCI and

by m o n i t o r i n g at

1268 nm.

proportional

z02 as opposed to t h e dimol

proportional

in

directly

for

the of of 778

the

time

The e m i s s i o n to

emission

the

concen-

which

zO 2 c o n c e n t r a t i o n

is

(18).

H202 and HOCI were used the

reaction

sufficiently

Low

Vol. 134, No. 2, 1986

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

~8 6 v

4

2 E

-= 0 ' 6

0

' ~o'do

Time(s)

o

'3'o'

Fig. I. E f f e c t o f DABCO on the time course o f 1268 nm chemiluminescence in the H202 + HOCl system. A. p2H 7, 100 mM sodium p h o s p h a t e , 150 uM H202, 150 ~M HOCl, 2H20 s o l v e n t . B. c o n d i t i o n s as in A w i t h 300 ~M DABCO added.

to

study

they

its

kinetics.

Buffers

enhance the

102 e m i s s i o n

As shown

Fig.

and s h o r t e n s reaction control

of

in the

of

H202 + HOCl.

not

by a f a c t o r

DABCO i n c r e a s e s

duration

experiments

HOCl ( d a t a

I,

made w i t h

the

of the

peak

No 1268 nm e m i s s i o n

Table

I

added t o

used,

since

30 ( 1 3 ) .

1268 nm e m i s s i o n

w h e r e DABCO i s

shown).

2H20 w e r e

is

intensity in

detected

either

demonstrates

the

that

in

H202 o r the

total

TABLE I . E f f e c t o f DABCO C o n c e n t r a t i o n on the 1268 nm Chemiluminescence o f the H202 + HOCl and H202 + HOBr R e a c t i o n s DABCO (mM)

H20z + HOCIa

H202 + HOBrb

/Idt /12dt )c /Idt )c fl2dt (arb. units) c (arb. units (arb. units (arb. units) c 0.0

1.00 ± 0.03

1.0 ± 0.09

1 O0 ± 0.04

1.0

0.03

0.80 ± 0.04

2.3 ± 0.1

0 96 ± 0.07

0.87 +- 0.11

0.i

0.63 ± 0.03

4.8 ± 0.2

0 98 -+ 0.05

0.86 + 0.14

0.3

0.64 ± 0.06

8

± 0.5

0 97 + 0.04

1.14 ± 0.06

1.0

0.56 ± 0.04

10

± 2

0 94 _+ 0.04

0.93 -+ 0.09

3.0

0.62 ± 0.01

13

± I

0 97 ± 0.07

0.98 ± 0.17

± 0.11

a c o n d i t i o n s : p2H 7 . 0 , 100 mM sodium p h o s p h a t e , 150 vM HOCI 150 vM H202, 2H20 s o l v e n t . b C o n d i t i o n s : pZH 7 . 0 , i 0 0 mM sodium p h o s p h a t e , 10 mM NaBr, 150 vM HOBr, 150 ~M H202, 2H20 s o l v e n t . CArbitrary units, 1.0 w i t h o u t DABCO.

each column n o r m a l i z e d to g i v e a v a l u e o f

779

Vol. 134, No. 2, 1986

yield

of

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

z02,

emission

which

intensity,

emission,

however,

square

of

the

by t h e

addition

z02 dimol

not

proportional

is

decreased

is

of

proportional

DABCO.

to

the

to

the

and t h i s

to

reported

postulate

integral

rate the

direct

a change

in

of

added to

their

the

concentrations

high

The d u r a t i o n determined the

rate

of

the

by t h e m i x i n g

of

the

of

the

of

but

duration

(5-10)

the

faster

If than

emission

the

reaction

Primary

form

amines,

taurine

to

(300 ~M),

In

is

DABCO

to

detect

when DABCO was the

result

(0.4

of

M) used.

was more l i k e l y

their

system

and not

H202 + HOCl system may

by two mechanisms. decrease of is

the

of

the

both

contrast,

the

by t h e

then of

the

amines

rate

both

(300 ~M),

are g r e a t l y

tertiary

amine,

effect 780

peak will

readily

is

HOCl

HOCI is

the

of

t h e H202

with

emission

When e i t h e r

inten-

The of

react

react

peak i n t e n s i t y

has at most a m i n i m a l

peak e m i s s i o n

chemically

alanine

the

quenching

chemiluminescence.

(19-21).

(300 ~M) o r

Direct

added amine w i t h

and s e c o n d a r y

chloramines

the

determined

1268 nm c h e m i l u m i n e s c e n c e

shown).

the

and t h e d u r a t i o n

H202 + HOCl s y s t e m , the

of

t h e H202 + HOCI r e a c t i o n ,

HOCI to

failed

study

It

of

M) and NaOCl

their

Amines may a l s o

intensity

decrease.

in

will

emission

+ HOCI r e a c t i o n . (19-21).

amines

not t h e d u r a t i o n of

102 f o r m a t i o n

was p r o b a b l y

properties

102 e m i s s i o n

z02 by amines

sity,

increased

H202 + HOCl r e a c t i o n .

The a d d i t i o n decrease

this

H202 ( 1 . 2

emission

of

the

enhancement

chemiluminescence but

of

is

effect

Deneke and K r i n s k y

(1),

of

by Deneke and K r i n s k y .

on 102 p h o s p h o r e s c e n c e .

system

the

The z02 dimol

quantity

explain

a novel,

the duration

of

integral

The i n c r e a s e d

sufficient

emission

necessary

to

by DABCO.

1268 nm e m i s s i o n ,

caused by DABCO is of

is

with

the

primary

added to

and t h e decreased

the

duration (data

not

trimethylamine

on t h e

kinetics

of

~02

of

Vol. 134, No. 2, 1 9 8 6

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

A

12 ~ I0 :>,

B

,

8 <

6 4

~2 i

i

i

:50 0 Time (s)

0

,

30

Fig. 2 . E f f e c t of DABCO on the time course of 1268 nm chemiluminescence in the H202 + HOBr system. A. p2H 7, 100 mM sodium phosphate, i0 mM NaBr, 150 uM Hz02, 150 ~M HOBr, 2HzO solvent. B. c o n d i t i o n s as in A with 300 ~M DABCO added.

production

(data

not

shown).

that

the

perturbations

demonstrate

caused by DABCO, a l a n i n e , a direct

reaction

catalyze

the

Addition minute

which

of

intermediate stable.

then

H202 to

earlier

produces formed

As shown in

Fig.

of

the

changed

of

reacts

rapidly of

no l i g h t .

This

reaction

of

any i n t e r m e d i a t e

HOBr a l s o

2,

the

the

reaction

and t h e

the

integral

square of

amine.

with

kinetics of the

part

an u n s t a b l e

H202 to

suggests

form

formed

are not

the

I02 than

greatly

~02. i

any not

be

(22). the altered

intensity

intensity

DABCO to to

cannot

produces

emission

may be r e l a t e d

that

DABCO and HOCI i s

the e m i s s i o n

of

influence

relatively

DABCO when compared to H202. 781

due to

DABCO may

i s much f a s t e r

The i n a b i l i t y

HOBr w i t h

large

DABCO and HOCl p r e p a r e d

H202 w i t h

H202 + HOBr r e a c t i o n of

in

of

by DABCO.

reactivity

are

data.

Neither

integral

and t a u r i n e

available

H202 + HOCI r e a c t i o n by DABCO.

H202 + HOCI system

H202 + HOCI by f o r m i n g

by the

from t h e

The r e a t i o n

the

a solution

The n a t u r e

determined

of

emission durations

of

between HOCl and the

reaction

intermediate

The s h o r t e n e d

are the low

nor

Vol. 134, No. 2, 1986

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

The data presented

in t h i s

study demonstrate

enhancement of red chemiluminescence used as a general

test

for

that

the

by DABCO should not be

z02 in complex biochemical

systems.

ACKNOWLEDGEMENTS I wish to thank W i l l i a m in performing

experiments.

GM-32974 from the National Veterans A d m i n i s t r a t i o n

Wardman f o r This

technical

work is supported

Institutes

of Health

assistance by g r a n t

and the

Research S e r v i c e . REFERENCES

1 .

2. 3. 4. 5. 6. 7. 8. 9. I0. II. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22.

Deneke, C. F . , and K r i n s k y , N. I . (1976). J. Am. Chem. Soc. 98, 3041-3042. Deneke, C. F . , and K r i n s k y , N. I . (1977). Photochem. P h o t o b i o l . 25, 299-304. L e n g f e l d e r , E., Cadenas, E., and S i e s , H. (1983). FEBS L e t t . 164, 366-370. O g r y z l o , E. A. and Tang, C. W. (1970). J. Am. Chem. Soc. 92, 5034-5036. Quannes, C., and Wilson, T. (1968). J. Am. Chem. Soc. 90, 6527-6528. Foote, C. S., Denny, R. W., Weaver, L . , Chang, Y . , and P e t e r s , J. (1970). Ann. N. Y. Acad. Sci. 171, 139-145. Smith, W. F . , J r . (1972). J. Am. Chem. Soc. 9__44, 186-190. Foote, C. S. (1979). in S i n g l e t Oxygen, Wasserman, H. H., and Murray, R. W., Eds., Academic Press, New York, pp. 139-171. Monroe, B. M. (1985). in S i n g l e t 02, Vol. I , P h y s i c a l Chemical Aspects, F r i m e r , A. A . , Ed., CRC Press, Boca Raton, F l o r i d a , pp. 177-224. Monroe, B. M. (1977). J. Phys. Chem. 81, 1861-1864. B o v e r i s , A . , Cadenas, E., and Chance, B. (1980). Photobiochem. Photobiophys. i , 175-182. Cadenas, E., B o v e r i s , A . , and Chance, B. (1980). Biochem. J. 187, 131-140. Kanofsky, J. R., (1983). J. B i o l . Chem. 258, 5991-5993. Kanofsky, J. R., (1984). J. Photochem. 25, 105-113. Kanofsky, J. R., !1984). J. B i o l . Chem. 259, 5596-5600. Kanofsky, J. R., W r i g h t . , J . , M i l e s - R i c h a r d s o n , G. E., Tauber, A. I . (1984). J. C l i n . I n v e s t . 74, 1489-1495. Salomaa, P., Schaleger, L. L . , and Long, F. A. (1964). J. Am. Chem. Soc. 86, 1-7. Bader, L.W., and O g r y z l o , E. A. (1964). Discuss. Faraday Soc. 377, 46-56. H o r r i s , J. C. (1967). in P r i n c i p l e s and A p p l i c a t i o n s of Water C h e m i s t r y , Faust, S. D., and Hunter, J. V., Eds., John Wiley & Sons, New York, pp. 23-53. Thomas, E. L . , Grisham, M. B . , and J e f f e r s o n , M. M. (1983). J. C l i n . I n v e s t . 72, 441-454. Weiss, S. J . , K l e i n , R., S l i v k a , A . , and Wei, M. (1982). J. C l i n . I n v e s t . 70. 598-607. McKeown, E., and Waters, W. A. (1966). J. Chem. Soc. ( B ) , 1040-1046. 782