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, Volume: 19( 9)

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Analysis and Speciation of Ternary Complexes of Copper Metal Ion with Pharmaceutical Ligand β-Diketone Ligand and Amino Acids

*Correspondence:

Sayujjta R. Vaidya, Jaishree J. Chamargore Department of Chemistry, Vivekananda Arts Sardar Dalipsing, Commerce and Science College, India, E-mail: srvaidyachem007@gmail.com, jaishreechamargore@gmail.com.

Abstract

The equilibrium studies of the mixed ligand complexes of copper(II) ion with drug β-diketone as primary ligand and the amino acids viz. Glycine, L-Leucine, Tryptophan, DL-Serine, DL-Valine, DL- Alanine, β-phenyl alanine, DL-methionine as secondary ligand were determined pH metrically at 30^oC and ionic strength of 0.1 M NaClO₄ in 50% (v/v) THF-water medium. The calculations have been made using the stability constant of generalized species computer programme

Keywords

Equillibrium constant; Amino acids; Mixed ligand complexes

Introduction

β-Dike tones and their metal complexes are associated with various pharmacological and biological properties such as anti-inflammatory [1], heptoprotective [2], antitumor [3], antiviral [4], anticancer activity [3]. They are also used in gastrointestinal and respiratory disorder [5]. Their biological activities as evidenced from their anticancer [6], antitumour, antioxidant [7], anti-inflammatory [8], antiviral [9] and immunodulatory activities [10]. In biological front, many of the β-dike tones were reported to be physiologically active and find applications in the treatment of many diseases. Their biological activity enhances on the formation of metal complexes. Literature survey reveals that there is an enormous growth of the study of metal complexes of β-dike tones in last few decades. A lot of work has been published on the study of β-dike tone complexes in solid as well as in solution. The complete formation of a complex may be predicted on the basis of its stability constants in solution. Glycine is the neutral, aliphatic, optically inactive non-essential, glycogenic amino acid [11-16]. It can be synthesized from CO₂ and NH₃ by glycine synthase or transamination of glyoxylate and in metabolism of serine and choline. It plays an important role in haeme synthesis. Heame is a tetra pyrol ring system with transition metal iron. The nitrogen from each pyrol is denied from glycine. It can form serine, creatine and purine.

Leucine [17] is neutral essential ketogenic amino acid and forms an acetoacetate and acetate. It is branched chain amino acid and taken up by brain and muscle. In leucine metabolism, transamination gives α-keto isocaproic acid, which is converted in to corresponding CoA, this is similar to oxidative decarboxylation of alfaketo glutarate and pyruvate. The enzyme complex is very important in the body of living organism.

Tryptophan is important amino acid. A metabolite of tryptophan 5-hydroxytryptophan (5-HTP), has been suggested as a treatment for epilepsy [18] and depression, Since 5-HTP readily crosses the blood brain barrier and in addition is rapidly decarboxylated to serotonin (5-hydroxytryptamine or 5-HT) [19]. Clinical trials, however, are regarded as inconclusive and lacking [20]. Serotonin has a relatively short half-life since it is rapidly metabolized by monoamine oxidase. Due to conversion of 5-HTP into serotonin by the liver, there may be a significant risk of heart valve disease from serotonin’s effect on the heart [21,22].

Serine is one of the naturally occurring proteinogenic amino acids. Only the L-stereoisomer appears naturally in proteins. D-serine was only thought to exist in bacteria until relatively recently, it was the second D amino acid discovered to naturally exist in humans, present as a signalling molecule in the brain, soon after the discovery of D-aspartate. Had D amino acids been discovered in humans sooner, the glycine site on the NMDA receptor might instead be named the D-serine site [23]. D-Serine is being studied in rodents as a potential treatment for schizophrenia and L-serine is in FDA approved human clinical trials as a possible treatment for ALS [24,25]. A 2011 meta-analysis found adjunctive sarcosine to have a medium effect size for negative and total symptoms [26].

Valine is essential amino acid [27] widely distributed but rarely occurs in amount exceeding 10%. It is branched chain amino acid and can be derived from alanine by the introduction of two methyl group present on α-carbon atom. This is glycogenic. On deamination, it forms methyl malonyl-CoA in place of two H atoms of the methyl group. Alanine is a nonessential amino acid, meaning it can be manufactured by the human body, and does not need to be obtained directly through the diet. Alanine is found in a wide variety of foods, but is particularly concentrated in meats. Good sources of alanine include:

Animal sources

Meat, Seafood, Caseinate, Dairy products, Eggs, Fish, Gelatin, Lactalbumin

Vegetarian sources

Beans, nuts, seeds, soy, whey, brewer's yeast, brown rice, bran, corn, legumes, whole grains. Alanine plays a key role in glucose–alanine cycle between tissues and liver. In muscle and other tissues that degrade amino acids for fuel, amino groups are collected in the form of glutamate by transamination. Glutamate can then transfer its amino group through the action of alanine aminotransferase to pyruvate, a product of muscle glycolysis, forming alanine and α-ketoglutarate. The alanine formed is passed into the blood and transported to the liver. A reverse of the alanine aminotransferase reaction takes place in liver. Pyruvate regenerated forms glucose through gluconeogenesis, which returns to muscle through the circulation system. Glutamate in the liver enters mitochondria and degrades into ammonium ion through the action of glutamate dehydrogenase, which in turn participate in the urea cycle to form urea [28].

Phenylalanine is aromatic essential glucogenic and ketogenic amino acid. In metabolism phenylalanine is converted in to tyrosine. In metabolism homogenstic acid is formed which undergoes cleavage and form fumarate and acetoacetate. The hormones such as adrenaline, noradrenaline, tyrosine and melanin pigment formed from tyroxine. Several abnormalities observed in phenyl nine metabolisms such as phenylketonuria and alkaptonaria.

Methionine [29] is essential glycogenic amino acid. It is the only common amino acid possessing an ether linkage. Cereals have sufficient quantity of mehionine whereas pulses lack in it. It is methylation product of homocysteine. Apart from its role as a protein constituent and as an essential amino acid, methionine is particularly important as a donor of active methyl groups.

Copper is a transition metal ion and is used by various enzymes in the body in different biochemical reactions. These reactions may be creating, decreasing the body’s inflammatory blood clotting [30] etc. Copper is absorbed by the body at two main sites such as small intestine and stomach. Copper does not float through the blood stream as copper ion but is carried by proteins. Two main carrier proteins especially for copper are ceruloplasmin [31] and albumin; these can carry many things including copper. Copper is stored in proteins called metallothione [32-34].

Survey of literature reveals that no work has been reported on complex tendencies of drug 1-[3,4,5-triethoxyphenyl-3(2-Hydroxyphenyl) propane 1, 3 dione with transition metal ion copper (II) in THF water solution. Therefore in order to understand the complex formation tendencies of 1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione it was though worthwhile to determine the formation constant 1:1:1 ternary complexes of 1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione with copper (II) in the presence of amino acids in 50%(v/v) THF-water medium at 30^°C at a fixed ionic strength 0.1 M NaClO₄.

Materials and Methods

Experimental

Micro analysis of the ligand is performed at the Central Drug Research Institute (CDRI). The H1 NMR spectra of ligand were recorded on EM-360 spectrophotometer at RSIC, Punjab University, Chandigarh (India) [35]. IR spectra of ligand were recorded in KBr pellet on a FTIR-4100 Jasco in the region 4000 cm-1-400 cm-1.

Reagents and chemicals

The glass distilled water was collected in a Stoppard bottle and always used fresh. Its pH was about 6.60 to 6.80. THF: HPLC grade THF was freshly distilled and used. Commercial THF was purified by standard method described by Vogel [34]. All other chemicals like perchloric acid, sodium perchlorate and sodium hydroxide were of AR grade, obtained either from B.D.H. (London) or E. Merck, Reidal (Germany). The solutions of above reagents were prepared in CO₂ free glass distilled water by taking precautions to avoid errors in their concentrations. Exact normalities were obtained by standard methods.

Instruments

An Elico model LI-120 digital pH meters in conjunction with an Elico combined electrode consisting of glass and reference electrodes in a single entity of the type CL-51 was used for the pH measurements. Synthesis and characterization of 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 dione [35].

Series of amino acids were of Anal R quality, obtained from Fluka (Germany) and were used as secondary ligands. These were recrystallized and their purity was checked by their M.P. Fresh solution of β-dike tone in freshly distilled THF and acid solutions in glass distilled water were prepared before performing the titrations. The potentiometric titration technique for the study of the mixed ligand complexes includes the titration of free HClO4 (A). Free HClO₄+Ligand 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 dione Drug (A+L). Free HClO₄+Ligand 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 dione+Metal ion (A+L+M) Free HClO₄+Ligand Amino acids (A+R). Free HClO₄+Ligand Amino acids+Metal ion (A+R+M). Free HClO₄+Ligand Amino acids+Ligand 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 dione+Metal ion (A+R+L+M).

Against standard solution of sodium hydroxide, were drug 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 dione and amino acid are two ligands. The ionic strength of solution was maintained constant i.e. 0.1 M by the addition of appropriate amount of sodium perchlorate solution. All the titrations are carried out at 30°C in an inert atmosphere by bubbling oxygen free nitrogen gas through the assembly containing the electrodes to keep out CO₂. The formation constant of ternary complexes were determined by computational programme SCOGS [36] to minimize the standard derivation.

Result and Discussion

Binary metal complexes

The proton ligand constant and metal ligand stability constant of 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 dione (L) and amino acids with Copper (II) determined in 50% (v/v) THF-Water mixture at 30°C and ionic strength μ=0.1 M NaClO4 are given in Table 1.

Ternary metal complexes

In the ternary systems, the mixed ligand titration curve coincides with acid+drug complex curve up to pH 2.8 and deviates afterwards. Theoretical composite curve remains towards left of the experimental mixed ligand curve, which is the indicative of mixed ligand complex formation. Since the mixed ligand curve do not coincides with either of the individual metal ligand binary complex curves, the formations of 1:1:1 complex by simultaneous equilibria was inferred.

The primary ligand 1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione (L1) form 1:1 and secondary ligand amino acids such as Glycine, Tryptophan, Serine, Valine, Alanine, Phenylalanine and Methionine form 1:1 and 1:2 complexes with Cu (II). It is evident from the figure of the percentage concentration species Cu (II)-1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione (L)-amino acids system, that the percentage distribution curve of free metal decreases sharply with increasing pH.

Species distribution studies

To visualize the nature of the equilibria and to evaluate the calculated stability constant of ternary complexes Cu (II)-1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione (L1)-glycine, species distribution curves have been plotted as a function of pH at temperature 30°C and ionic strength μ=0.1 M NaClO4 using SCOG program. From the SCOG distribution curve it is concluded that the formation of ternary complex started only after the metal-primary complex has attained its maximum concentration. This indicates that the metal-primary ligand complex Cu (II)-1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione (L1) is formed first and the secondary ligand Cu (II)-glycine coordinated to it, resulting the formation of ternary complex.

According to this method in this system ternary complex of 1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione (L1) with glycine, leucine, tryptophan, serine, valine, alanine, phenylalanine, and methionine show the following types of the concentration species distribution.

Where M=Metal; L=β-diketone; R=Amino acid

The stability constant of ternary complexes. The relative stabilities of the binary and ternary complexes are quantitatively expressed in terms of β111, β20, β02, KL, KR, Kr and log K values which are presented in Table 1. The values of β111, β20 and β02, reveals the preferential formation of ternary complexes over both binary complexes of 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 Dione and Glycine ligands. The considerably high positive values of KL and KR indicate that the ternary complex is highly stable as compared to the metal complex of primary and secondary ligands. In addition to this, the positive values of Kr support the higher stability of ternary complexes than the binary complexes. The observed slight negative values of log K is in accordance with HSAB principle [Greaser R and Singh H, inorg. Chem. 9(1970) 1238] which states that the log K value is less negative when the secondary ligand coordinates through an oxygen and a nitrogen atom viz. amino acids. In order to demonstrate the mechanism of the formation of ternary complex, various equilibria involved in the process are taken in to consideration. In addition to equilibrium reaction 5, the following reaction equilibria are also possible for the formation of ternary complex.

TABLE 1. The proton ligand constant and metal ligand stability constant of 1-[3,4,5-triethoxy phenyl Propane 1,3 dione and amino acids with copper (II) determined in 50% (v/v) THF-water mixture at 30^°C and ionic strength µ=0.1 M NaClO₄.

ML+R → MLR (6)

MR+L → MLR (7)

The other way of characterizing the ternary complexes is by disproportionate reaction.

ML2+MR2 → 2MLR (8)

This reaction is possible only if both the ligands form 1:1 and 1:2 complexes

ML₂+MR → MLR+ML (4)

MR₂+ML → MLR+MR (5)

ML+MR → MLR+M (6)

The reactions (4) and (5) corresponds to the systems containing one ligand which forms only 1:1 complex and the other forms both 1:1 and 1:2 complexes. The equilibrium reaction (6) represents the system containing the ligands which forms 1:1 complexes with the metal ion. This reaction is possible only if the sufficient concentration of ML and MR are available. The stability of ternary complexes in terms of secondary ligands is also examined in the present study. The observed order of stability of Cu (II) complexes is as follows.

For Cu 1-[3,4,5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1,3 dione Amino acid Leucine R2?Glycine R1?Valine R5?Tryptophan R3?Methionine R8?Serine R4?Alanine R6?Phenylalanine R7 Among these, Kr is a statistical relationship and gives the relative stability of mixed ligand chelate with the overall stabilities of binary chelates. It can be seen from Table 2 that the Kr values are positive for all the ternary chelates investigated in the present work which shows that β111>β20>β02. The magnitudes of Kr are almost same in the range of 0.99 to 1.4 for all the systems, suggesting that the ternary chelate in every case is more stable than binary ones. In addition to Kr, two more constants KR and KL are calculated and used to discuss the relative stability of mixed ligand and binary complexes.

TABLE 2. Parameters based on some relationship between the formation of ternary complexes of Copper (II) metal ion with 1-[3, 4, 5-triethoxyphenyl-3(2-hydroxyphenyl) propane 1, 3 dione (L) in the presence of amino acids (1:1:1) system. [Temp=30^°C; Ionic strength=0.1 M NaClO4; Medium=50% (V/V) THF-Water].

Conclusion

In all the systems studies, the value of KL is greater than K20. Same trend is seen from all the systems. Similarly the value of KR for all the system with some exceptions is greater than K02. These higher values of KL and KR indicate that the formation of ternary complexes is favorable than the formation of 1:2 binary complexes of both the ligands. The positive values of Kr in all the systems also confirm that the ternary complexes are more stable than their corresponding binary complexes. A simpler way of this comparison is to obtain Δ log K values. In the present work Δ log K values are calculated for all the systems by using relationship. Δ log K=log βMLR-(log KML+log KMR). It is observed from the Table that Δ log K values of all the ternary systems are negative values which indicate the extra stabilization of ternary complexes.

Acknowledgment

The Authors Declared that the research Work was carried out on their Own.

Conflict of Interests

The author declares that there is no conflict of interests regarding the publication of this paper.

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