329 - Investigation on the electrochemical properties of catecholmines and melanins

Bioelectrochemistry a n d Bioenergetic$ 7 (x98o) 74x--748 f . ElectroanaL Chem. 116 (x98o) 74t--748 Elsevier Sequoia S.A., L a u s a n n e - P r i n t e d in I t a l y Short

Communieatim~

329Investigation on the Electrochemical of Cateeholamines and Melanins *

Properties

b y J. ~[ATYSIK a n d M. PRZEGALI~SKI I n s t i t u t e o f C h e m i s t r y ° x~Iaria S k l o d o w s k a U n i v e r s i t y PL ~f. S k l o d o w s k i e j 3, L u b l i n , P o l a u d R e v i s e d m a n u s c r i p t r e c e i v e d S e p t e m b e r x x t h 198o

Summat'¥

T h e m e l a n i z a t i o n process was i n v e s t i g a t e d using e l e c t r o c h e m i c a l methods. O n t h e a.c. a n d m u l t i s w e e p c u r v e s t h e p a r t i c u l a r stages o c c u r as s e p a r a t e d , c o n s e c u t i v e l y rising a n d d e c r e a s i n g peaks, c o r r e s p o n d i n g t o g r a d u a l association o f p r e c u r s o r molecules. A n i n t e r e s t i n g effect is t h e rise of d.c. w a v e s a t p o t e n t i a l s c h a r a c teristic for HzO z ( a p p e a r i n g u s u a l l y if Oz is p r e s e n t in t h e solution) in d e o x y g e n a t e d solutions of melanins. T h e differences i n t h e c u r v e s of c a t e c h o l - m e l a n i n a n d i n d o l e - m e l a n i n p e r m i t to distinguish t h e s e c o m p o u n d s using p o l a r o g r a p h i c m e t h o d s . T h e r e s u l t of p r a c t i c a l i m p o r t a n c e is t h e possibility of a fast det e r m i n a t i o n of s m a l l a m o u n t s of a d r e n a l i n e a n d n o r a d r e n a l i n e b y use o f multisweep methods.

Introducdon A n essential s t r u c t u r a l e l e m e n t of c a t e c h o l a m i n e s a n d m e l a n i n s is t h e s y s t e m o f h y d r o x y l (or quinone) g r o u p s in t h e orgho position a t t h e a r o m a t i c r i n g [I]_ S u c h a s t r u c t u r e ensures t h e i r s t r o n g c h e l a t i n g a n d reversible r e d - o x p r o p e r t i e s [z, 3]- I t is j u s t t h e s e p r o p e r t i e s t h a t a r e m a i n l y responsible for t h e f u n c t i o n s of c a t e c h o l a m i n e s a n d m e l a n i n s in living o r g a n i s m s a n d t h e y also m a k e t h e s e c o m p o u n d s c o n v e n i e n t o b j e c t s for e l e c t r o c h e m i c a l s t u d i e s [4, 5]T h e m e t h o d s of t h e i n v e s t i g a t i o n s i n c l u d e d d . c . , a_c., s.w. (square wave) p o l a r o g r a p h y , m u l f i s w e e p a n d e l e c t r o c a p i l l a r y m e a s u r e m e n t s since * Presented at the 5th Internationa/ Symposium on Bioelectrochemistry, 3 - 8 S e p t e m b e r x979, W e i m a r (D.D_R.).

o3o2--4598/8o/o74x-o748

~

x98o E l s e v i e r S e q u o i a S . A .

742

Matysik a n d ]Przeg-.alifigki

these techniques p e r m i t a v e r y quick or even continuous recording of t h e r e d - o x p r o c e s s e s a n d a d s o r p t i o n effects. The present investigations dealt with catechol-melanin and indolemelanin as well as with adrenaline and noradrenaline as soluble comp o u n d s o c c u r r i n g i n li~-ing o r g a n i s m s , c o n t a i n i n g t h e e s s e n t i a l b L t a c t u r a l e l e m e n t s i m i l a r t o t h a t o f mela~ing_

Kslmrimental Melanins were o b t a i n e d w i t h o u t use of enzymes, b y t h e air oxidation o f c a t e c h o l a n d D O P A i n s o l u t i o n o,x zvlr K O H . T h e p r o d u c t w a s p r e cipitated w i t h hydrochloric acid at p H = z a n d purified b y dialysis. T h e d . c . s t u d i e s w e r e c a r r i e d o u t u s i n g t h e p o l a r o g r a p h L P - 6 o , (LABOR_~TOR,~t PfizS-rROJ~), a.c.--using p o l a r o g r a p h P L P - A C , I C h F , P . A . N . , s . w . - u s i n g p o l a x o g r a p h O H - I o 4, RADELKIS, t h e m u l t i s w e e p m e a s u r e m e n t s w e r e m a d e o n a D i g i s c o p e p o l a r o g r a p h O P - 3 , TELPOD a t s w e e p r a t e s f r o m z t o z o V s -1. T h e o s c i H o p o l a x o g r a p h i c o b s e r v a t i o n s w e r e d o n e o n t h e P o l a x o s c o p e K ~ I ~ I K 1>--576, t o o b t a i n t h e r e l a t i o n s h i p d U / d t = f ( U ) . T h e electrocapillary curves of melanins were o b t a i n e d from avera g e d d r o p - t i m e o f 5o d r o p s o f H g a t c o r r e s p o n d i n g p o t e n t i a l s . The e l e c t r o l y t e u s e d w a s a I z~Ir w a t e r s o l u t i o n o f K O H o r x z~r N H a N O a N H a O H buffer, p H = 8,5T h e c h a x a c t e r i s t i c s o f t h e D . M . E , w e r e 2 . x m g s -x. a n d ~ . 4 s d r o p t i m e . :am t h e r e f e r e n c e e l e c t r o d e sex~'ed a S . C . E . T h e m e a s u r e m e n t s w e r e m a d e i n t h r e e (digiscope, s.w., a . c . p o l a r o g r a p h y ) o r t w o e l e c t r o d e s y s t e m , a t 293 K . If necessary, the oxygen was removed from the solutions using nitrogen.

Results and d/~ .... ;on

I t t u r n e d o u t t h a t m e l a n i z a t i o n o f c a t e c h o l a m i n e s is a c o n v e n i e n t object for the study. Simultaneously with the solution turning brown, t h e multisweep polaxography curves registered t h e stages of the process a s c o n s e c u t i v e r i s i n g a n d f a l l i n g o f r e v e r s i b l e s i g n a l s (peaks), c o r r e s p o n d i n g m o s t p r o b a b l y t o t h e d e g r e e s o f p o l y m e r i z a t i o n (Fig. 1). C o n s i d e r i n g t h a t on t h e a.c. polaxograms a t t h e p o t e n t i a l of t h e m e n t i o n e d signals t h e well p r o n o u n c e d peak occur b u t no polarographic waves were observed o n t h e d . c . c u r v e s w e c o n c l u d e t h a t t h e a . c . a n d m u l t i s w e e p pea-ks a r e capacitive a n d n o t faxadaic. The a.c. a n d electrocapillaxy curves show t h a t t h e s i g n a l axe c o n n e c t e d t o e l e c t r o d e p r o c e s s e s t a k i n g p l a c e i n t h e i n t e r v a l o f t h e a d s o r p t i o n o f t h e c o m p o u n d (Fig. 2, 3)- T h e first c h a n g e s in t h e curves b e c o m e visible a l r e a d y after seconds a n d t h e period of quick changes lasts for a b o u t I h. I t t u r n e d o u t t h a t t h e r a t e of t h e m e l a n i z a t i o n p r o c e s s is q u i c k e n e d b y t h e p r e s e n c e o f h e a v y m e t a l i o n s .

l~-lectrochemlcal

Properties

of Catecholamines

and

~[elanins

743

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.20

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f - 0.5

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--30 ]j 2 ] •-40 F i g . I. ~ [ u l t i s , v e e p c u r , ' e s o f a d r e n a l i n e L 5 X I o -a :lir i n N H ~ N O s - i - N H a O H , pH ~8,5, in the p r e s e n c e o f a i r . S w e e p r a t e ~ _~o V s-Z. L t h e s u p p o r t i n g e l e c t r o l y t e ; z. t h e s u p p o r t i n g e l e c t r o l y t e - I - a d r e n a l i n e . a f t e r o r a i n ; 3- t h e s u p p o r t i n g e l e c t r o l y t e q - a d r e n a l i n e , a f t e r x o r a i n ; 4- t h e s u p p o r t i n g e l e c t r o l y t e + a d r e n a l i n e , a f t e r r h .

A strange effect was the occurrence of the d.c. polarographic waves o f i n d o l e - m e l a n i n e i n d e o x i d i z e d i ~ / K O H s o l u t i o n a t U~_ ~ - o , 9 5 V a n d U¢_. ~----1,38 V w h i c h a r e c h a r a c t e r i s t i c f o r t h e p r e s e n c e [6] o f H O ± a n d HoO~ (Fig. 4)- T h e f o r m a t i o n o f H z O . - in t h e r e a c t i o n b e t w e e n q u i n o n e s a n d O H - i o n s w a s d e s c r i b e d b y LAZAROV a n d c o w o r k e r s [7]

Q + oH-

~ Q - : - + OH"

Matysik

744

and

Przegali6~ki

IP

.1:3 Fig. z. polarograms. I. I ~f KOH, deoxid i z e d s o l u t i o n ; 2. a s I q - c a t e c h o l - m e l a n i n 2 0 o lzg c m -a. a m p l i t u d e ~ i o i n V , f r e q u e n c y ~ 7 6 I-Iz. a.c_

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F i g . 3E l e c t r o c a p i l l a r y c u r v e s ( d r o p t i m e v s . U)_ i . I ~'~1FK O I - I , d e o . ~ d i z e d s o l u t i o n ; ~. a.s I -k ¢~techoI-mel~-i-; 3-as i q-indolc melanin.

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Electrochemical

Properties

of

Catecholamines

a~d

xXlelanins

745

(where Q q u i n o n e fo~m) a n d t h e c o u r s e o f t h i s r e a c t i o n s h o u l d b e forced by following irreversible dimerization : z

O H " --> H202

however, this explanation makes objections due to energetic reasons. -1;9

•

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.

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L z ~1[ K O H .

d e o x i d i z e d s o l u t i o n ; 2. a s t -t- i n d o l e m e l a n i n

x o o ~ g c m -a .

T h e similarity between t h e polarographic b e h a v i o u r of the deoxid i z e d m e l a n i n e s o l u t i o n s a n d H202 s o l u t i o n s i n I zl~/ K O H o c c u r s also i n t h e o t h e r m e t h o d s . S.w. p e a k s c o r r e s p o n d i n g t o H202 a n d t o m e l a n i n e solutions are located at the same potential values, they have similar a s y m e t r i c s h a p e a n d i d e n t i c a l h a l f - w a w e w i d t h (240 m V ) . I n t h e a.c. polaxograms of b o t h c o m p o u n d s no peaks (corresponding to t h e H , 0 2 e l e c t r o r e d u c t i o n ) o c c u r e d . O n t h e o s c i l l o p o l a r o g r a m s , a t p o t e n t i a l s corr e s p o n d i n g t o s.w. p o l a r o g r a p h i c w a v e s , i n b o t h c a s e s t h e r e d i d n o t appear any oscillopolarographic indentation. B y e l e c t r o r e d u c t i o n o f t h e Tl+z a n d P b +± i n d e o x i d i z e d m e l a n i n e solutions effects of h i d d e n limiting currents occurred characteristic for t h e r e d u c t i o n o f t h e s e i o n s i n t h e p r e s e n c e o f 0 3 o r H202. S i n c e i n t h e d.c. c u r v e s o f c a t e c h o l e - - m e l a n i n o n l y a w a v e c o r r e s p o n d i n g t o H±O 2

746

xNfatysik and

Przegali6ski

r e d u c t i o n o c c u r s , it offers a p o s s i b i l i t y f o r d i f f e r e n t i a t i o n b e t w e e n t h e t w o m e l a n i n s (Fig. 5)Another practical result of the oscillopolarographic and multisweep investigations may be seen in the possibility of a quick determination of low concentration of hydrolyzing salts of heavy metals in the presence of catecholamines as well as a possibility of quantitative determination o f c a t e c h o l a m i n e s (Fig. 6) ( a d r e n a l i n e , n o r a d r e n a l i n e D O P A ) a f t e r i n t r o d u c t i o n o f s o m e i n o r g a n i c c a t i o n s t o t h e s o l u t i o n [8].

1.9 ~

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x. ,~t K O H .

d e o x i d i z e d s o l u t i o n ; 2. a s x -1- ~ t e c h o l

m e l a n i n 2 0 o izg c m -=.

In order to verify the results as an analytical method, we examined pha~luaceutical solutions of adrenaline for injections, o,27 cm s of the c o n t e n t o f t h e i n j e c t i o n w a s d i l u t e d b y 9,73 c m a a m m o n i u m b u f f e r a n d a f t e r a d d i t i o n o f C u ffi+ t o c o n c e n t r a t i o n x-5 × xo - s 1~/, t h e s o l u t i o n w a s examined using the multisweep polarograph. The results of these examinations were very promising, however, the method brought no positive r e s u l t s i n u n d e r p r o t e i n i z e d o r g a n i c fluids. T h e m e c h a n i a m o f f o r m i n g of oscillographic signals in the Me~+-catecholamine system already at l o w s u b s t r a t e c o n c e n t r a t i o n is b a s e d o n t h e c o u r s e o f r e v e r s i b l e r e d - o x processes of catecholamine complexed cations under continuous complex adsorption, enhancing the concentration at the electrode surface_

Electrochemical

Properties

of Catecholamines

aRd

Melanins

747

'40 ;> .30 -20

-10 -1.6

f

-1.

U{V)

0

:.j

.-10

- -2.0

-30

-40 F i g . 6. M u l t i ~ v e e p c u r v e s , r . i . 5 × xo - s .~f C u ÷-* i n N H , N O s - I - N I - [ a O H . p K = 8. 5 i n t h e p r e s e n c e o f asr- ; 2. a s r i n t h e p r e s e n c e o f a d r e n a l i n e 3- x r o - s _1I s w e e p r a t e = 2 0 V s - t .

T h i s is i m p l i e d b y t h e increase o f signals d u r i n g t h e d r o p t i m e o f t h e D . M . E . a n d b y t h e f a c t t h a t t h e p o t e n t i a l s of t h e i n d e n t a t i o n s o r pea-ks axe c h a r a c t e r i s t i c n o t for c a t e c h o l a m i n e s b u t f o r t h e m e t a l used. I t s e e m s t h a t in t h e a d s o r b e d l a y e r m e l a n i n e r e t a i n s b o t h t h e c h e l a t i n g a n d r e d - o x s y s t e m s , which, t o g e t h e r w i t h t h e flat o r i e n t a t i o n of t h e a r o m a t i c r i n g a t t h e D . M . E . ( a n d t h e ~ e l e c t r o n s interactions), m a k e s possible ion a d s o r p t i o n a n d e l e c t r o n t r a n s m i s s i o n b e t w e e n t h e e l e c t r o d e s u r f a c e a n d m e t a l ions.

Referenees G . A . Sw.~,,~, Progress in the'Chemistry of Organic zVatural Products. S p r i n g e r V e r l a g . "~Vien, N e w Y o r k (x974) [ 2 ] T . S A R N A a n d S_ ~_.UKIEWICZ. Folia Histochem. Cytochem_ 1 0 (Z972) 265 [ 3 ] I-][.G. C A S S Z D Y a n d A . A _ K u . ~ , Oxydation-Reduction Polyn~rs, I n t e r s c i e n c e Publ., New York (I965) [I]

748 [4"[- ~'[5] M. [6] J . [7] S. [8] ~[.

Matysik and Przegali6ski I-IORAK a n d J_R. G~v_T_~I-rEo J~olo P h a r m a c o l . T ( I 9 7 I) 429 PRzEc;_.~..x~sw.I. P h . D . Thesis. UMCS. : L u b l i n (t976) CHODKOIVSKI, Rob'n_ Chb'~n. 2~ (x953) 3o9 LAZAROV. A_ TROFINOV a n d T, PoPOV, Z. P h y s . C h e m . I I (x967) z45 PRZEGALIIqSKI a n d J . ~IATYSIK, T a l a n t a 27 (z98o) 9 z o