Radiolytic reduction of merocyanine 540 in methanol solution

Journal of Radioanalytical and Nuclear Chemistry, Articles, Vol. 190, No. 1 (1995) 97-102

R A D I O L Y T I C R E D U C T I O N OF MEROCYANINE 540 IN M E T H A N O L SOLUTION

J. MAYER, R. KRASIUKIANIS

Institute of Applied Radiation Chemistry, Technical University, Wr6blewskiego 15, 93-590.L6d~ (Poland)

(Received November 1, 1994)

Radiolytic reduction of merocyanine 540 (MC) in acidic (0.02 tool. dm -3 in H2SO4) and neutral methanol solution was studied by pulse radiolysis. The spectra centered around 400 and 700 nm of the MC reduced transients recorded in acidic methanol and in neutral solution were found to be quite similar but they disappeared with different rates suggesting that different radicals (MCH" and MC'-) were responsible for these spectra. The rate constant of "CH2OH reaction with MC was found to be 7.108 mo1-1 �9 3- s -1.

Merocyanine 540 (MC) is an anionic lipophilic polymethine dye which was used as a probe of changes in membrane potential in various systems (e.g.). 1,2 MC is also of therapeutic interest in connection with its selective antineoplasmic and antiviral activities (e.g.), 3 Light-activated MC has been shown to be effective against certain types of tumour cells and enveloped viruses, including human immunodeficiency virus (HIV-1)J -7 The mechanism for its in vivo reactivity was suggested to involve electron transfer reactions 8 hence the study of such processes using pulse radiolysis seems to be reasonable.

Radiation induced oxidation and reduction of MC was investigated in neutral aqueous 2-propanol in the time scale from mseconds up to minutes but the rate parameters of the reactions responsible for reduction were not found, s The reduction of MC by solvated electron in aqueous methanol 9 and methanoP ~ solution was

observed previously and the rate constant for this reaction was found to be in the range (6.0 § 6.6) �9 109 mo1-1 �9 dm 3. s-1. 9,1~ Under anaerobic conditions MC was shown to undergo radiolytic reduction leading to formation of the corresponding radical anion, MC--. In all cases the spectra recorded after pulse irradiation of At-saturated neutral solutions and assigned to MC-- were very similar and had a broad absorption band centered around 400 nm. The radical anion should exist in equilibrium with its protonated form, a neutral radical MCH-:

MC.-+ H§ �9 ~MCH. (1)

J. MAYER, R. KRASIUK1ANIS: RADIOLYTIC REDUCTION

In acidic solution this equilibrium should be shifted to the right and theneutral radical s!lould be the stable form. In this paper we report the radiolytic reduction of MC by 'CH2OH radicals in acidic and neutral methanol solution.

Experimental

MC (Aldrich, -97%) and methanol (Merck, for spectroscopy) were used as received. The solutions prepared Were kept in the dark in order to avoid photodecomposition of MC. The samples contained in a quartz cell of 1.4 cm path length were bubbled with argon for 40 minutes prior to irradiation. Pulse radiolysis experiments were made with a linear accelerator (ELU-6) delivering 17 ns or 3 kts electron pulses (6-8 MeV). Transient differential absorption spectra were recorded using appropriate interference filters (Zeiss) t0 minimize photodecomposition of MC. The pulse radiolysis system used was described in details elsewhere. 1H3 Electron pulses were calibrated by thiocyanate dosimeter. Computer simulation of the kinetic curves was made using ACUCHEM program) 4

Results and discussion

The structure of the MC molecule is shown below:

O • / CH2~CHs N

5" CH CH--CH CH~ / S

N \

O (CH2)3CH 3 (iH2)3

- +

SOsNa MC

In acidic methanol solvated electrons (e 7) are rapidly converted into H atoms:

e 7 + CH3OH~=-->H- + CHaOH (2)

98

J. MAYER, R. KRASIUKIANIS: RADIOLYTIC REDUCTION

The latter and other radical products of radiolysis (e.g. CH~ and CH30") react with

the solvent molecules to form hydroxymethyl radicals .CHzOH: 15,16

CHaOH + H.--).CH2OH + H 2 (3)

The absorption changes recorded after 17 ns pulse irradiation of acidic methanol solution (0.02 mol. dm -3 I-I2SO 4) containing MC in low concentration (8.10-6 mol. �9 dm -3) were similar to the ones recorded in neutral solution, l~ However, the precise comparison of the spectra was impossible due to low intensity of. the signals recorded in acidic solution in this case. The low efficiency of reduction of MC in acidic methanol may be due to the slow rate of reaction of MC with hydroxymethyl radicals (see discussion later in the text) and efficient recombination of the latter radicals (2.8 - 109 mo1-1 �9 dm 3, s-l). 16

The absorption changes recorded after pulse in'adiation of more concentrated solution of MC (5- 10 -4 mol, dm -a) in acidic (0.02 mol- dm -3 I-LzSO4) (Fig. la) and neutral methanol solutions (Fig. lb) were indeed very similar. In the range 425-600 nm the measurement could not be done due to strong absorption of the parent compound. The spectrum recorded 1.6 Vts after the pulse in acidic methanol and assigned to the MC neutral radical. MCH'. is weaker but similar to the one obtained in neutral solution, In both spectra the absorption bands are centered around 400 and 700 nm, Acidic and basic forms of radicals often differ in absorption spectra and in most cases the acidic form absorbs at shorter wavelengths than the basic one. 17 Our result may suggest that the speclrum of the MC neutral radical, MCH-, is very similar to the spectrum of the corresponding radical anion, MC--. An alternative explanation that the same acid-base

form of the MC reduced transient is stable both in neulral and acidic (0.02 mol. dm -3 H2SO 4) methanol does not seem to be probable. At longer times the decay of the MC reduced transient was observed [inset (2) in Fig. lb] leading to formation of more stable product [inset (1) Fig. tb]. This process was observed in both systems, neutral and acidic one but the decay rates were different. In neutral solution during 80 ItS ca. 27% of MC.- disappeared comparing with more than 50% of the radicals (MCH.) in acidic system. The data strongly support the hypothesis that the spectra observed in neutral and acidic solutions representdifferent transients. The spectra recorded 80 kts after the pulse are shown in Fig. 1. Similar absorption changes were found by SARNA et al., after pulse irradiation of MC in neutral aqueous methanol. 9

The computer analysis of bleaching kinetic curves recorded in solutions containing MC in low concentration (Fig. 2a, b, d) was done using ACUCHEM program 14 and the rate constant of MC reduction by CH2OH radicals was found to be equal to 7 �9 108 mol -~ �9 3- s -1. In acidic methanol the simple scheme of competition between decay of hydroxymethyl radicals in reaction with MC and in recombination process was

99


,x 0.08 <1

006

Q04

0.02

< 0.20A <:1

a)

o o

I h

I

r /oo.

61

b)

I Jm 50O 6O0 7OO 8O0

Wavelength, nm

Fig. 1. The differential absorption spectra recorded 1.6 (m) and 80 ps (o), respectively, after pulse irradiation 3 3 of MC (5- 10 -4 tool- din- ) in Ar-saturated acidic (0.02 tool. dm- H2SO4) (a) and neutral (b)

methanol solution; pulse 17 ns, dose 40 Gy. Insets: oscilloscope traces recorded for .g= 330 nm, 10 ItS per division, 7.8% per division (1), and 2=400 nm, 10 ItS per division, 7.3% per division (2)

considered (Fig. 2a, b). In these calculations the rate constant of .CH2OH recombination was assumd to be 2 .8 .10 9 mo1-1 - d m 3 �9 s - l . 16 In neutral solution the redaction of MC by solvated electrons was additionally included with the rate constant equal to 6.6.109 mo1-1 �9 d m 3 . s -1 (F ig . 2(1). 1~ The decay of e~ by recombination [k(e~ + e~) =

= 3 �9 10 9 m o l -s � 9 3- s - l ] , 15 which was also taken into consideration, was found to have little effect on the rate constant of CH2OH raction with MC in neutral solution.

For higher concentration of MC (5- 10 4 rnol. dm -3) the initial rapid growth of absorption attributed to reduced forms of MC was found both in acidic (0~ tool � 9 -3

H2SO4) (Fig. 2c) and neutral methanol (Fig. 2e). Comparison of the latter results with the kinetic profile corresponding to the decay of e~ in neutral solution (Fig. 2f) showed

100


i i i (a)

..... ! ............ ! .................. ~ ..... ~ ............... -

i-~!i~i~i;-ii!~il;i112! i;-!i~i-~iiill ~-!ii~;i ~ ~ : _ ~ ! : -

(c)

........... ~ ~ . ~ ~ ~ ,

~@~ ........... ii .... iii iii~i iiiii

.......... ?-I ............. ........... : ......... ........ ............. .............. ........... I

............ .................... !i! I

: :{d) ....................... 2. ........... ~ ........................ .. ..........................................................

....................... i ...................... : .................... ~ ............... i ...................

J

.................................... ,y~-- ' - -o , / -~,~. . , , , ~ . ~ , ~ .~ . -- .- - - - - - - - ....

Fig. 2. Oscilloscope traces recorded after pulse irradiation fordeareated solutions of MC in acidic (0.02 mol.

�9 dm -3 H2SO4) (a, b, c) and neutral methanol (d, 10 e, f): a - 8 . 10-6 mol- dm -3 MC, pulse 17 ns, dose 42 Gy, A= 575 rim, 100 Its per division, 3.7% per division; b - 8 �9 10 -6 mol. dm -3 MC, pulse 3 Its, dose 982 Gy, A= 500 nm, 100 Its per division, 4.2% per division; c - 5 . 1 0 -4 mol �9 dm -3 MC, pulse 17 as, dose 40 Gy, 2 = 379 nrn, 500 ns per division, 6.2% per division; d - 1 - 10 -5 tool- dm -3 MC, pulse 17 ns, dose 39 Gy, 2,= 575 rim, 10 Its per division, 8.3% per division; e - 5- 10 -4 tool- dm -3 MC, pulse 17 ns, dose 40 Gy, ~,= 399 ran, 200 ns per division, 7.0% per division; f - 5 �9 10 -4 mol . �9 dm -3 MC, pulse 17 ns, dose 40 Gy, ,~= 621 rim, 200 ns per division, 3.8% per division

101


that this growth cannot be due to the reaction of MC with e~. In acidic solution e~ reacts rapidly with CH3OH ~ [k(e~+ CH3OH~)= 5.2.101~ mo1-1, dm 3. s-i] 16 and disappear during the 17 ns pulse. In both solutions, CHsO./.CH2OH radicals are formed at such short time scale i.e., during the 17 ns pulseJ 9 The rate constant for the reaction of MC with these radicals in ns time scale has to be at least one order of magnitude higher than the above calculated value (7. 10s mo1-1, dm 3. s-l). According to GETOFF et al. 19 CH30. radicals are the primary radiolytic transients of methnol in addition to H', e~ etc., but not 'CH2OH species. The radical CH30. may react with MC during the pulse whereas -CH2OH radicals are responsible for after the pulse reduction.

Maybe this "fast" process corresponds to the reaction of MC with dry electron. Dry electrons were assumed to be unreactive towards protonated forms of alcohol 18 hence they could survive in acidic solution. The concentration of MC in our experiments was too low in order to be competitive with the solvation process of dry electrons hence the dry electron mechanism seems to be much less probable than theCH30, mechanism.

References

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L. J. JENSKd, Biochim. Biophys. Acta, 1146 (1993) 136. 3. L NORTH, H. HEYNDORFF,. (3. LEVY, J. Photochem, Photobiol. B: Biol., 17 (1993) 99. 4, W. H. H. GO'NTHER, R: SEARLE, F. SIEBER, Phosph0rt~ , Sulfur and Silicon, 67 (1992) 417. 5. W. H. H. GONTHER, R. SEARLE, F, SIEBER, Seminars in Hematology, 29 (1992) 88.: 6. S. PERVAIZ, M. BATI'AGLINO, J, L. MATTHEWS, K. S. GULLIYA, Cancer Chemother. Pharmacol.,

31 (1993) 467. 7. M. HOEBEKE, A. SERET, J. PIE~I'E, A. VAN DE VORST, Biochemistry, 32 0993) 2730. 8, A. HARRIMAN, L. C. T. SHOUTE, P. NETA, J. Phys. Chem., 95.(1991) 2415. 9. T. SARNA, B. PlLAS, C. LAMBERT, E. J. LAND, T. G. TRUSCOTT, J. Photochem. PhotobiO1. A, 58(3)

(1991) 339. i0. R. KRASIUKIANIS, J. MAYER, Radiat. Phys. chem., (i995) 45 (2) (1995) 309, 11. s. KAROLCZAK, K. HODYR, R. JLUBIS, J. KROH, J. Radioanal. NucL Chem:, 10t (1986) 177. 12. J. MAYER, R. KRASIUKIANIS,J. Chem. ~ . Faraday Trans., 87 (1991) 2943. I3. S. KAROLCZAK, K. HODYR, M. POEOWII~KI,. Radiat. Phys. Chem., 39 (1992)1. 14. W.BRAUN, J..T. HERRON, D. K. KAHANER, Int. L Chem. Kin., 20 (1988) 5i. 15. D. W. JOHNSON, G; A~ SALMON, Radiat. Phys. Chem., I0 (1977) 294. 16. H~:J. JUNG, N. GETOFF, Z. Natufforsch., 46a (199!) 625. 17. A. J. SWALLOW, in: The Study of Fast Processes and Transient Species by Electron Pulse Radiolysis,

J. H. BAXENDALE, F. BUS!, (Eds), D. Reidel Publishing Company, 1982, p. 289. 18. W. J. HUNT, in: Advances~in Radiation Chemlstry~ M. BURTON, J. L. MAGEE (Eds), Wiloy-lnterscience,

New York, 1976. 19. N. GETOFF, A. RI'ITER, F. SCHWORER, P. BAYER, Radiat. Phys. Chem., 41 (!993) 797~

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Radiolytic reduction of merocyanine 540 in methanol solution