SYNTHESIS OF NEW BORON COMPOUNDS WITH AMOXICILLIN AND SOME OF ITS METAL COMPLEXES WITH USE THEM IN ANTIBACTERIAL, ASSESSMENT OF HEPATOPROTICTIVE AND KIDNEYACTIVITY, ANTICANCER AND ANTIOXIDANT APPLICATIONS

Objective: New ligand ({7-[2-amino-2-(4-hydroxyphenyl)-acetamido-3,3-dimethyl-6-oxo-2-thia-5-aza bicyclo[3,2,0]heptane-4-carboxylic boric anhydride}) with its Co (II), Ni (II) and Cu (II) complexes. And new mixed ligand copper complex was synthesized. Methods: The ligand was synthesized by the reaction of boric acid with amoxicillin (1:1) and the mixed ligand complex has been synthesized by the reaction of the ligand, 4-aminoantipyrine and Cu (II) ion (1:1:1). Results: All studied compounds were characterized by the spectral method: Fourier transform infrared, ultraviolet-visible, thermal analysis (TG and DTG), flame atomic absorption and nuclear magnetic resonance. Also CHNS, melting point, magnetic susceptibility and molar conductivity. Conclusion: According to the obtained data, all complexes were non electrolyte and the geometry was octahedral for all complexes. All synthesized compounds were tested as antibacterial agents against Escherichia Coli, Pseudomonas auroginosa as Gram-negative bacteria (G-) and Staphylococcus aureus epidermis as Gram-positive bacteria (G+). The results showed that copper complexes were more active in (10-2M) than the other compounds. The medicinal applications (hepatoprotective and kidney in serum of mice, histopathological of liver and kidney, anticancer and antioxidant in human cell were studied of the synthesized compounds and gave a good results in all tested.


INTRODUCTION
Boric acid and borates are exists in water, food and soil. They are used as least toxic pesticides to kill insects, mites, fungi, algae, fleas and wood decay fungi [1]. The mechanism of killing contains working to poison the stomach and absorbing the waxes which protect insects. Also boric acid used as antiseptic in talcum powder, mouth-washes, eyewashes and protective ointments. Another application includes reducing the flammability of cellulosic materials used in production of leather [2].
In the present work we are synthesis a new derivative of amoxicillin with boric acid, also the metal complexes of this derivative (ligand) with Co (II), Ni (II) and Cu (II) ions were synthesized. We are synthesis a mixed ligand complex from the reaction of amoxicillin and 4-aminoantipyrine with Cu (II) ion. The medincinal applications and the biological activity were studied for all synthesized compounds.

FT-IR Spectroscopy
The FT-IR spectrum of the ligandshowed bands at 3456, 3379 cm −1 which refer to stretching vibration of NH 2 [16]. The band of ν N-H (amide) was appeared at 3205 cm −1 [16]. The spectrum of exhibited absence of carboxylic OH band (3300-2621 cm −1 in amoxicillin) and appear new band at 1342 cm −1 , which due toν O-B [17,18]. The spectrum showed band at 3525 cm −1 which was assigned to phenolic OH group [16]. The band of ν C=O (carboxylic and β-lactam) was appeared at 1770 cm −1 [16,18]. While the band of ν C=O of amide appeared at 1683 cm −1 [16]. The data can be shown in Table 2. The spectra of complexes C 1 , C 3 , Mix.showed shifting in some band positions. The bands of ν NH 2 shifted to higher frequency of asymmetry bands and to lower frequency of symmetric bands (Table 2) and this refer to coordination with metal ions through the nitrogen atom of NH 2 [4]. The band of υ NH (amide) was shifted to higher frequency in all complexes and this is attributed to coordination with metal ions. The spectrum of mixed ligand showed a new band at 1560 cm −1 due to imine (υ C=N) [19]. All spectra of complexes exhibited new bands at lower frequency which refer toυ M-O, υ M-N and υ M-Cl [20]. The data can be shown in Table 2.

1
HNMR and 13 CNMR were used to characterized the ligand (L) and its metal complexes using d 6 -DMSO as solvent.

H-NMR Spectroscopy
The spectra of all compounds showed peak at δ 2.5 ppm which refer to chemical shift of DMSO as a solvent. The spectra of Lexhibited absence the proton peak of carboxylic OH (about δ 10 ppm in amoxicillin) [21] and appear new proton peak of B-OH at δ 8.74 pp [22] ( Table 3). The spectra of Co (II), Ni (II) and Cu (II) complexes (C 1 -C 3 ) were showed shifting to higher value in proton peak position of N-H amide and NH2 groups (Tables 4-6) compartion with the spectrum of ligand (L) [21]. The spectrum of (Mix.) exhibited further proton peaks which attributed to 4-aminoantipyrine [19]. The spectrum of (Mix.) also showed shifting in position of chemical shift of N-H amide and NH 2 compartion with the ligand (Table 7) because the complexation with metal ions through nitrogen atoms of N-H amide and NH 2 . The spectrum of (Mix.) also showed shifting to higher values in position peaks of N-CH and this is because formation of imin group in neighbring atom [21]. The spectra of the ligand and C 3 can be shown in Figs. 2-4.

CNMR Spectroscopy
The 13 CNMR data are listed in (Tables 8-12). The chemical shift of DMSO as a solvent appeared at δ 40 ppm. The 13 CNMR spectrum of the ligand (L) showed shifted to higher values of carboxylic C=O group

Al-Jebouri and Noorikhaleel
compartion with parent drug [23] and this is because the binding with B(OH) 2 . The spectra of metalcomplexes (C 1 -C 3 ) exhibited shifted to higher values in the chemical shift of C=O (β-lactam), C=O amide and CH-NH 2 and this is attributed to complexation with metal ions [23]. In (Mix.) spectrum, the carboxylic C=O band absent and appeared a new bands at δ 159.36, which assigned to δ C=N (imino group) [19].
As well as the C=O (β-lactam), C=O (amide) and CH-NH 2 bands were shifted to higher values and this is because coodination with metal ions.

Electronic spectra
The electronic spectra of synthesized compounds were recorded in methanol (10 −4 M) at room temperture. The spectral data were listed in Table 13. The electronic spectrum of ligand exhibited high intensity band in 356 nm (28089 cm −1 ), which due to π→π* transition [24]. In addition the spectrum of ligand showed low intensity bands at 373 nm (26809 cm −1 ), which assigned to n→π* transition [24]. The data were listed in Table 13. The spectrum of C 1 complex showed change in position of π→π* transition (Table 14).

Thermal analysis of the ligand and its metal complexes
The TG and DTG analysis were performed under nitrogen gas in the range heating 16-650°C and the heating rate (10°C/min). This technique was used to study the thermal stability of synthesized compounds as well as to characterize the suggested structures. The thermal decomposition data were listed in (Table 14) and the thermographs of the ligand (L) and C 1 complex were shown in (Figs. 2 and 8). The results showed that the stability of the ligand and its complexes was increase as the following order Mix. <C 1 <L 1 <C 2 <C 3 . The results of degradation exhibited good agreement in percentage of calculate and found mass loss and this confirm the suggested structures of synthesized compounds [29].

Biological activity
The antibacterial activity of the ligand (L) and its metal complexes have been evaluated against (P. auroginosa, E. coli (G-) and Staphylococcus aureus (G+)). The bacterial activity was test with two concentration (10 −2 and 10 −3 M) of the primary materials and all synthesized compounds. The C 3 and Mix. in 10 −2 M were the most effective against the studied microorganism. The ligand (L) exhibited small activity with S. aureus compartion with its complexes. All synthesized compounds in 10 −3 M were more active from the parint drug against S. aureus. The antibacterial data were listed in (Table 15). DMSO solvent was used as control [30].

Hepatoprotective and kidney evaluation
Hepatoprotective evaluations included assessment of liver function enzyme (GOT, GPT and ALP) and renal function test (Urea, Creatinine, TSP and Alb) in serum. The results were listed in (Tables 16 and 17).
The obtained results showed the positive effect of all studied compound on GOT, GPT, ALP, Urea, Creatinine, TSP and Alb.

Histopathological evaluation
The results of histopathological evaluation of liver and kidney tissue which treated with C 3 and Mix. showed the positive effect      of studied compounds and no negative effect were on tissue observed.

Anticancer activity (cytotoxicity assays)
Study of anticancer activity for amoxicillin, L, C 3 and Mix. was carried out on different cancer cell lines (AMJM, SOK-7 and CMF-3). The inhibition rate of cell growth (the percentage of cytotoxicity) was calculated as the following equation: A-B Inhibitionrate= ×100 A Where A and B are the optical density of control and the optical density of test.
The study of anticancer activity for the ligand exhibited high percentage of cytotoxicity comparison with amoxicillin, this is due to presence of boron element and this is agreement with literature [31]. The increase in percentage of cell inhibition due to presence of β-lactam ring in all studied compounds as well as presence of aromatic ring in Mix. (4-Aminoamtipyrine) and this is enhanced the anticancer activity [32]. Also found that the Schiff base ligand improve the anticancer activity [33]. The Schiff base linkage (-C=N) is an essential structural requirement for antitumor activities [33,34].Cytotoxicity assays of ligand and its complexes are shown in Tables 18-20 and Figs. 9-11 for L and C 3 .
DPPH radical scavenging activity (RSA)(antioxidant activity) 2,2-diphenyl-1-picryl-hydrazyl (DPPH) was used in RSA for the evaluation of antioxidant and this is the rapid technique for screening the RSA of specific compounds or extracts [35]. DPPH is a stable free radical that can accept an electron or hydrogen radical and get converted to a stable, diamagnetic molecule. The results of this study showed that the ligand exhibited a negligible DPPH activity. In