ÖZET Bu çalışmada, nitrik asit perklorik asit ortamında, seryum(IV) mono ve dinitrato komplekslerinin oluşum denge sabitleri (K, ve K2) ile seryum(4), seryum mono- ve dinitrato komplekslerinin 400 nm dalga boyu için molar absorptivitesi s0, e-ve e2 hesaplanmış ve ilgili türlerin absorbanslarının aditivitesinden (toplamsallığından) yararlanarak teorik absorbans değerleri ile deneysel veriler karşılaştırılmıştır. İyonik şiddet ve hidrojen iyonu derişimi sabit tutularak (1,0 M) nitrik asit - perklorik asit derişimlerinin değişimi ile seryum(IV) türlerinin, ve ayrıca amino asit türlerinin hidrojen iyonu derişimine göre dağılımları incelenmiştir. Çalışma ortamında seryum(IV) ve amino asit türlerinin dağılımları belirlendikten sonra yükseltgenme kinetiği spektrofotometrik yöntemle araştırılmış ve L-fenil alanın, L-aspartik asit ve L-serin'in seryum(IV) ile reaksiyonlarının yalancı birinci mertebe hız bağıntısına uyduğu, L-metionin'in yükseltgenme reaksiyonunun ise 2. mertebeye uyduğu bulunmuştur. Seryum(IV), amino asit, nitrat iyonu ve hidrojen iyonu derişimleri, sıcaklık gibi parametreler değiştirilerek hız sabitleri saptanmış ve bu bilgilerin ışığında reaksiyon mekanizması önerilmiştir. Reaksiyonun radikal mekanizması üzerinden yürüdüğünü kanıtlamak için akrilamid poiimerizasyonu gerçekleştirilmiş ve L-fenil alanin ile L-aspartik asit uç gruplu poliakrilamid'in en yüksek verimle elde edilmesi için polimerizasyon şartiarı değiştirilmiştir. Ayrıca poiimerlerin metallerle kompleks yapma özelliğinden yararlanarak L-aspartik asit uç gruplu poliakrilamid'e uranii ve bakır iyonu bağlanması spektrofotometrik yöntemle izlenmiştir. Son zamanlarda üçlü poiimer-metal-protein komplekslerinin farmakolojik ve immünolojik önemi nedeniyle polimer-bakır kompleksi ile Bovin serum albuminin üçlü kompleksi elde edilmiş ve bu kompleks spektrofotometrik ve viskozimetrik yöntemle incelenmiştir. VI - SUMMARY KINETICS AND REACTION MECHANISM OF THE OXIDATION OF SOME AMINO ACIDS WITH CERIUM(IV) Oxidative decarboxylation of amino acids is of interest both from a pure chemical standpoint and from the point of view of its bearing a light on the mechanism of amino acid metabolism. Therefore, investigation of amino acids in the context of non enzymatic chemical oxidation processes constitutes a potential area for intensive experimentation. In this work, cerium(IV), being a strong oxidant, has been utilized for investigating the oxidation kinetics of some amino acids. Knowing the distribution of cerium(IV) species is beneficial for better understanding this type of kinetics. Distribution of cerium(IV) species in aqueous media was studied by measuring the total optical absorbance of Ce(IV) species in differing nitric acid-perchloric acid mixture solutions. The formation of the nitrate complexes is expressed by the equilibria K 3-*- Ce(4) + HN03 ^=Ce(N03) +H* Ce(NQ3 ) + HN03 = Ce(N03 );+ + H^ where Kj and K2 are the equilibrium constants for the formation of mono- and dinitrato- complexes of cerium(IV) and Ce(4) represents all other forms of the metal including the free ion and the hydroxo species. [Ce(4)] = [Ce**] + [Ce(OH)3`] + [Ce(OH),;'] The total molar absorption coefficient for cerium (et) is defined by the equation et = (£0 + e, K, n + e, K, K, n: ) / (1 + K, n + K, K: n2 ) where n = [HNOJ / [H~] and e0, e, and s2 are the molar absorption coefficients of the Ce(4), mono- and dinitrato- species respectively. The - v i i -solutions of the above equation for a set of data yielded the following results with regression coefficients ranging between 0.9992 and 0.9997: K, =0.520 s0 =140 K2 = 0.650 e, =817 e2 =247 Using the above results, the relative distribution of Ce(IV) species in HN03 - HCI04 mixture solutions of different concentration was investigated while the ionic strength and hydrogen-ion concentration was maintained at 1.0, i.e. 1=1.0 and [H+]=1.0. Taking advantage of the additivity of the individual absorbances of the related species, the expected (theoretical) absorbances of Cerium(IV) at 400 nm wavelength were computed, and these came out to be in accordance with experimental absorbances. For the hydrogen ion concentrations less than or equal to 1.0 M, relative distribution computations performed on the amino acid species (phenyl alanine, aspartic acid, serine and methionine) showed that the protonated (cationic) forms of amino acids were prevalent in > 99% percentages. In the chosen medium (HN03 - HCI04 mixture containing 1.0 M H* ions), the oxidation kinetics of some amino acids were followed spectrophotometrically. At the wavelength chosen for eliminating spectral interference, i.e. X - 395 nm, neither the amino acid nor the oxidation product absorbed light and the only species responsible for light absorption was Ce(fV) species. The rate of the oxidation reaction of L-phenyi alanine with Ce(IV) was followed by measuring the absorbance at 395 nm due to Ce(IV) species in the excess of amino acid. Under the experimental conditions employed, the reactions obeyed pseudo-first order kinetics laws, and the rate constants (k'obs) were calculated from the slope of the lines drawn with In^ - A^ ) vs. time, where At and Aw are the instantaneous and final absorbances. it was established that at constant phenyl alanine concentrations and ionic strengths, the rate constants did not vary with the initial Ce(IV) concentrations. In solutions containing constant Ce(IV) and H* ion concentrations and ionic strength (1=1.0 M) but different nitrate concentrations, the variation of pseudo-first order rate constants with L-phenyl alanine concentrations was followed in the form of (1/kobs) vs. (1/[HL]) plots. vm -The values of K,, K2, Kg, Kg Kg and k: computed from non linear regression analysis of the curves using the slopes and intercepts, where Kg, Kg and Kg are the complexation equilibrium constants of the 3+ Ce(4), Ce(N03 ) and Ce(N03 );` species respectively with the amino acid and k: is the decomposition rate constant of the activated complex, i.e. the Ce(IV;%pecies-amino acid complex, are as follows: K, = 0.526 K` =1.890 k= = 7.30x1 0`4 `.o Jo K2 = 1.730 ^ =5.220 Kg, = 0.274 l2 The value found for K, was in accordance with the previously reported value, i.e., the equilibrium constant for the formation of cerium(IV) mono nitrate. The temperature effect was studied in HN03 - HCI04 solutions, and it was found that at constant [H+ ], the rate constants increased with increasing temperature and decreasing nitrate ion concentration. When the total nitrate ion concentration and ionic strength were kept constant in HN03 -NaN03 solutions of different H'ion concentration, the rate constants were found to increase with both acidity, i.e., [H~ ], and temperature. The calculated activation energies (Ea) increased with increasing nitrate concentration. A high concentration of nitrate ion probably slowed down the electron transfer from the organic substrate to cerium by occupying the coordination sites around the metai ion, and this effect was exhibited in an elevated activation energy. Since the Ce(IV)-Ce(lll) redox couple assumes the highest standard reduction potential in HCI04 medium where complexation of Ce(IV) is minimal, the oxidizing power of tetravalent cerium is decreased in nitrate media. When the effect of inert electrolyte cations on the rate constants was investigated, it was established that the rate constants were inversely proportional to the atomic radii of the cations. Silver(l) nitrate had a catalytic effect on the reaction, and the rate constants increased with temperature. The overall scheme of reaction mechanism was postulated as follows: - IX -(i) Nitric acid complexaiion: Ce(4) +HX ^ [CeX,](4-°-+H- [CeX]3* (X=NOj andHX=HN03) (ii) Amino acid protonation: R R H2 N-CH-COOH + H^HJ* -CH-COOH (HL) (H2 L)+ (iii) Zwitter ion formation: R R H2 N-CH-COOH 5= H3 N^ -CH-COO` (iv) Activated complex formation between Ce(IV) species and amino acid: [CeX, ](4ih + HL =^ C, (activated complex) (v) Decomposition of activated complex: R I C, ? H,N-CH-COO` + [CeX, ](3-°*+H* slow (vi) Fast reaction steps of radical conversion (decarboxylation), double bound formation and hydration: R R I I fast H, M-CH-COO' > H, N-C-H + CO, R R 1 i fast H2 N-C-H + [CeXi ]MH)+ > HN = C-H + [CeX, f`'' + H` fast HN = CHR + H:0 > RCHO + NH, - x -where i=0, 1 or 2; K- = equilibrium constant for Cj complex formation, and k^ is the rate constant for the decomposition of the activated complex. The oxidation kinetics of L-aspartic acid with Ce(IV) was studied using a known excess of the amino acid, and it was revealed that pseudo-first order kinetics with respect to Ce(IV) was followed. When the aspartic acid concentration and ionic strength were kept constant, the initial Ce(IV) concentration did not influence the rate constant. In HN03 - HCI04 mixture solutions of constant Ce(IV) concentration, the dependence of the rate constant on L-aspartic acid concentration ([HL]) and on the nitrate ion concentration were studied; the rate constant increased with [HL] and decreased with [NO;]. Non-linear regression analysis of the (1/k'obs) vs. (1/[HL]) curves yielded the following findings by using the slope and intercepts values: K, = 0.589 K^ = 0.454 kJo = 8.90x10^ K2 = 0.601 K^ = 2.820 1^=1.110 Thus these K-, and K2 values matched with the previously calculated ones from Ce(IV)-HN03 - HCI04 equilibria. in nitric acid-perchloric acid media, the temperature effect was studied and activation parameters calculated. The corresponding activation energies (Ea ) were found to increase with [NO; ]. As for the catalytic effect of AgN03 on the Ce(IV)-L-aspartic acid reaction, the rate constants increased linearly with silver(l) ion concentration. The rate constants were found to increase with [HL] and decrease with [NO;]. The rate constants again increased with temperature in catalyzed medium, and the activation energies were reduced when compared to those of uncataiyzed media. In the pseudo-first order oxidation of L-serine with Ce(IV), results similar to those of L-phenyl alanine and L-aspartic acid were obtained, probably indicating that an identical mechanism of Ce(IV) oxidation was applicable to all the amino acids studied. The concentrations of L-serine were varied for different nitrate media, and regression analysis of (1/k'obs) vs. (1/[HL]) using the siope and intercept values yielded the following values: XT-4 Ki =0.660 K^ = 7.81 kJo = 4.94x10 K2 = 0.561 Kjx =16.10 K^ = 16.60 Again these K1 and K2 values are in accordance with previously reported ones. The complexation equilibrium constant of Ce(4)-HL activated complex was smaller than those obtained with Ce(N03 ) -HL and Ce(N03 )22~-HL for all the amino acids (HL) studied. When the spectrochemical series of ligands in octaedral field are considered: (NO3 < halogenides < OH` < H2 O < NH3 < CN` ) water is on the far right of nitrate, indicating that aqua-complexes are stronger than the corresponding nitrato-complexes. Thus it should be harder for an amino acid molecule (HL) to displace a water molecule from the coordination sphere of the Ce(IV) ion than a nitrate ion. This may account for the fact that hL or l^ is greater than K^. The Ce(IV) oxidation of L-methionine was a second order reaction. The methionine oxidation was much faster than those of L-phenyl alanine, L- aspartic acid and L-serine. Stochiometric investigations revealed that the latter three amino acids consumed 2 moles of Ce(IV) while L-methionine consumed 4 moles of Ce(IV) per mole of amino acid. The reaction products in each case were identified by preparative oxidation. Carbon dioxide was detected by Ba(OH)2 solution, NH3 by Nessler reagent, and the corresponding aldehydes by derivatizing with 2,4 - dinitrophenylhydrazine in the form of hydrazone-precipitates. The final oxidation products were identified as phenyl acetaldehyde from L-phenyl alanine, formyl acetic acid from L-aspartic acid, hydroxy acetaldehyde from L-serine and acrolein from methionine. Polymerization of acryiamide was undertaken for the purpose of proving that Ce(IV) oxidation of the a-amino acids proceeds according to the radical mechanism. a-Amino acid or Ce(iV) alone did not initiate polymerization under experimental conditions, but the free radicals generated from the a-amino acid-Ce(IV) reaction were capable of initiating the polymerization of acryiamide. The optimal polymerization conditions were investigated for maximizing the conversion efficiency of L-phenyl alanine and L-aspartic acid. XllSpectrophotometry investigation of the polymer-metai complexes was carried out by the uranyl(VI) and copper(ll) complexes of polyacrylamide containing L-aspartic acid terminal groups. By adding different amounts of protein solutions to polymer-copper complexes, the variation of absorbance at 400nm (A^) with time was followed. The absorbances after 24 hours were plotted against (n^/hp) rations, i.e., the mole ratio of bovine serum albumin to that of the polymer. The formation water-insoluble complexes increased up to a (nBSA/np ) ratio of 1.0, beyond which (n^/rip > 1) BSA addition causes the system to be water-soluble. Recently in the field of biopolymers, investigations on the protein- metal-polymer ternary complexes have been intensified. Research has been initiated on the possible utilization of the synthesized ternary complexes of this work as radioprotectors due to the fact that immunological activity of such compounds has been somewhat established in the literature. - xm 108