Complexation Equilibria of Vitamin B9, Glycine Oligopeptides with Di- and Trivalent Metal Ions

It is well known that group vitamins B complex form wide organic-compounds that cannot be synthesized by humans and necessary for the tropism of human beings and need to be part of our daily intake. It occurs in living cells as essential substances for growth. Folic acid, one of important known vitamin B family known as vitamin B9 is structurally composed of pteroic acid and glutamic acid connected via an amide linkage. It is essential for normal human cell division, cell growth, formation of red blood cells and energy production. It also gives a definite effect in inhibiting the growth of tumors. The use of folic acid in dietary supplements has been dramatically increasing in recent years because of folates established role in reducing the prevalence of neural tube defects and its role in reducing the risk of cardiovascular disease and anemia. Chemically, the solubility of folic acid is crucial for drug delivery, absorption, transferring in the human body and crystallization in the manufacturing process. Glycine oligopeptides such as glycylglycine, glycyl-L-phenylalanine and glyclglycylglycine are biological molecules that are made up of simplest amino acid glycine playing many important roles in various biochemical processes. Considering the biological importance of folic acid (their possible application in medicine by combining the therapeutic properties of folic ligand and the other ligand in one compound and in chemical modeling of the transport and storage of some metal ions in living systems), the considerable attention has been paid in recent years on the study of the complexation Complexation Equilibria of Vitamin B9, Glycine Oligopeptides with Diand Trivalent Metal Ions


INTRODUCTION
It is well known that group vitamins B complex form wide organic-compounds that cannot be synthesized by humans and necessary for the tropism of human beings and need to be part of our daily intake. It occurs in living cells as essential substances for growth 1 . Folic acid, one of important known vitamin B family known as vitamin B9 is structurally composed of pteroic acid and glutamic acid connected via an amide linkage 2 . It is essential for normal human cell division, cell growth, formation of red blood cells and energy production [3][4][5] . It also gives a definite effect in inhibiting the growth of tumors 6 . The use of folic acid in dietary supplements has been dramatically increasing in recent years because of folates established role in reducing the prevalence of neural tube defects and its role in reducing the risk of cardiovascular disease and anemia 7,8 . Chemically, the solubility of folic acid is crucial for drug delivery, absorption, transferring in the human body and crystallization in the manufacturing process 9 . Glycine oligopeptides such as glycylglycine, glycyl-L-phenylalanine and glyclglycylglycine are biological molecules that are made up of simplest amino acid glycine playing many important roles in various biochemical processes [10][11][12][13][14][15] .
Considering the biological importance of folic acid (their possible application in medicine by combining the therapeutic properties of folic ligand and the other ligand in one compound and in chemical modeling of the transport and storage of some metal ions in living systems), the considerable attention has been paid in recent years on the study of the complexation Complexation Equilibria of Vitamin B9, Glycine Oligopeptides with Di-and Trivalent Metal Ions equilibria of hydroxamates, phenolates, amino acids, nonprotein amino acids and vitamins with metal ions [16][17][18][19][20][21][22][23][24][25][26] . The fact that, the ternary systems are somewhat better models for complicated biological systems, as the importance of ternary complexes in biochemical systems is beyond question. So, it is worthwhile to assemble information on their formation, stability and structure and on the mutual influence of two ligands bound to the same metal ion. In this study, a UV-visible spectrophotometric investigation of metal ion-vitamin B9glycine peptide unit was performed to elucidate the complexation equilibria of these systems.
UV-Visible spectrophotometric measurements: Di-and trivalent metal ion complex species were spectrophotometrically studied in solutions at about 298 K using a compact, double-beam Jasco V-530 UV/Visible Scanning Spectrophotometer with standard 1 cm quartz cells working on its current supply (220V) device. UV/visible bandwidth, scan speed and data interval used are 0.1 nm, 100 nm/min and 1 nm, respectively. The solutions of different binary metal-ligand complex species stability determination were done at various concentration ratios [TL : TM = 1 : 3, L (vitamin B9) = FA (folic acid), M (metal ion) = Cu(II), Co(II), Ni(II), Fe(III), Cr(III) and Al(III) metal ions, P (peptide = G, GG, GP and GGG glycine oligopeptides], while measurements for various mixed ligand complex species were done at concentration ratios TL: TM : TP =1:1:1. Order of mixing of solutions of different reagents was maintained strictly throughout the work. The vitamin B9 solution was first added to the metal ion solution, then glycine oligopeptide was added to this binary solution and again kept for a few minutes to reach complete equilibrium. The pH adjustment was done using Techne pH meter (model 3540) with glass and calomel electrode assembly and checked frequently with buffer solutions of Buffer, Hydrion ® Tablets (pH values of 4.00 ± 0.02, 7.00 ± 0.02 and 9.00 ± 0.02). The pH of desired solution was adjusted using hydrochloric acid (Panreac, Spain) and sodium hydroxide (Across Organics, USA) solution of suitable concentration. Each measurement was repeated at least four times.

RESULTS AND DISCUSSION
Ultraviolet-visible absorption spectroscopic measurements of vitamin B9 (folic acid, FA) in different pH medium (pH ≈ 3, 4, 5, 6, 7, 8 and 9) were shown below in Fig. 1. Based on the above UV-visible spectroscopic results, it is concluded that the protonation equilibria of folic acid was affected by the pH changes of the aqueous medium and due this equilibria could be explained based on the molecular structure of folic acid (Scheme-I).
In the present study, we followed the UV-visible absorption spectra of folic acid different pH media, in which the measurements indicates two different maximum absorption bands at two different wavelengths (λ = 313 and 376 nm) and the absorbance increases with the increment of the basic media of the folic acid solution. These transitions involves energies  energy of 33220 cm -1 for AlGP; 306 nm with energy of 33111 cm -1 for CrG; 306 nm with energy of 33111 cm -1 for CrGG; 306 nm with energy of 33111 cm -1 for CrGGG; 307 nm with energy of 33220 cm -1 for CrGP complex species. As seen above, each iron(III), aluminum(III) and chromium(III) metal binary complex species involving glycine oligopeptides give an absorption band in which Fe(III), Al(III) and Cr(III) have d 5 , P 6 and d 3 electronic configuration with spectroscopic ground term symbol of 6 S, 1 S and 4 F, respectively. These orbitals are non-degenerate states and cannot split by either an octahedral or tetrahedral fields 25,27 , hence d-d transition should be expected for all of the above complexes. However, the single absorption band observed is assigned to π → π * transition of the chromophoric groups in the complexes.
• The ternary systems involving copper(II)-folic acidglycine oligopeptides complex species was found to display two absorption maximum wavelengths (λmax) at 300, 341 nm with energies of 32463, 37372 cm -1 , respectively, for glycine; 300, 345 nm with energies of 32463, 37914 cm -1 , respectively, for glycylglycine; 304, 350 nm with energies of 32895, 37914 cm -1 , respectively, for glycylglycylglycine; and 298, 360 nm with energies of 32895, 38997 cm -1 , respectively, for glycyl-L-phenylalanine complex species. Copper(II) has a d 9 configuration with one unpaired electron. All the copper(II) complexes are either blue or green. In this case, copper(II) has a d 9 electronic configuration, which implies a spectroscopic ground state term symbol of 2 D. The 2 D orbitals splitted in a tetrahedral field into two sub-energy levels, namely 2 T2 and 2 E. Hence d-d transition is expected from 2 T2 → 2 E transitions. The first UV-visible absorption band in each metal complex is assigned to metal ligand charge transfer, while the second band is assigned to 2 T2 → 2 E transitions due to a tetrahedral geometry.