Anti-cancer and Antioxidant Activities of Some New Synthesized Mannich Bases Containing an Imidazo (2, 1-B) Thiazole Moiety

Synthesis and preliminary biological evaluation of imidazo (2, 1-b) Thiazole derivatives is reported. Under Mannich conditions, a series of new imidazo (2, 1-b) Thiazole derivatives were synthesized. Starting from the reaction of 2- amino thiazole with 4- bromo phenyl bromide to produce 5-(4-bromo phenyl) imidazo (2, 1-b) thiazoles, following by introduce the substituted aminomethyl at position 6-by reacting with different aromatic amines under Mannich conditions to afford 6-secondary amine-5-(4-bromo phenyl) imidazo (2,1-b) thiazole in high yields. FT-IR, 1H NMR, and 13C NMR techniques were used to characterize the synthesized derivatives. In addition, all compounds were tested for their antioxidant activity, and three of them were examined for cytotoxic activity against kidney cancer using an MTT assay. The product (A9) has shown an excellent anti-cancer activity with 69.69 value.


Synthesis 6-(4-bromophenyl) imidazo [2, 1-b] thiazole (compound 1A)
A mixture of 2-aminothiazole (0.01 mol) and 4-bromophenacyl bromide (0.01 mol) was heated in ethyl alcohol (80 mL) under reflux for 18-20 hours. The reaction was monitored by TLC (petroleum ether/ethyl acetate 1:2) until there were no more starting materials. After filtering the reaction mixture, a solution of NaOH (5%) was added to bring the pH of the mixture to around (10)(11). The mixture was then allowed for 4 hours to acquire the most quantity of precipitate possible during the digestion process. The precipitate was then filtered, rinsed in distilled water, dried, and re-crystallized with EtOH, yielding yellowish-orange crystals with an Excellent yield (90 %). [21,22], and the physical properties of this compound 1A are shown in Table 1.

Synthesis of Mannich bases compounds (2A-13A)
Drops of conc. HCl solution were added to a combination of compound 1A (1 mmol) and formaldehyde (37 %) (1 mmol) until the pH of the reaction medium reached 5-4, then agitated for 30 minutes. Then one of the different amines (primary or secondary) was added (1 mmol) and the mixture was heated for 4-18 hours. The solid mass was filtered, washed, dried, and purified with EtOH once the reflux was halted and the reaction mixture had cooled to room temperature [23]. A synthetic route of various Mannich bases derivatives is shown in the scheme 1 and the physical properties of these compounds 2-13 A are shown in Table 2. and FT-IR spectra data (cm -1 ) of Mannich bases derivatives 2-13 A are shown in Table 3, 1 HNMR spectra data (δ ppm) of these derivatives 2-13 A are shown in Table 4, 13 CNMR spectral data (ppm) was listed in Table 5. The technique for synthesizing several Mannich bases  R=  ,  ,  ,

Anticancer Activity
Using 3-(dimethyltiazole-2-yl) 2, 5-diphenyltetrazolium bromide, the cytotoxic effect of the novel imidazo [2, 1-b] thiazole derivatives on the kidney cancer cell line (HEK293) was assessed (MTT). After treating HEK293 cells separately with compounds 4A, 8A, and 9A, the MTT test was used to compute cell viability and inhibition rate on the tumor cell line using varied quantities of compounds, and the parameters of assessment were determined. The viability percentage of treated cells was calculated in contrast to the normal cell line WRL.

Scheme 3: mechanism of formation compound 1A
The formation of 1A as given approval by FT-IR spectrum data which includes disappearance of NH 2 band at 3200-3300 cm -1 and (C=O) band at 1700 cm -1 new bands are forming of (C=N) imidazo at (1679-1662) cm -1 due to formation of imidazole ring. The FT-IR spectra data (cm-1) of compound (1A) were shown in Table (2a) The 1 HNMR spectrum data of compound [1A] included the appearance of multi signals of aromatic rings protons at (8.14-7.36). The 13 C-MR data of compound [1A] included the appearance of (N-CH=C imidazole) at (108.7) ppm and (C=N) at (146.5) ppm.

Synthesis of Mannich derivatives 2-13 A
The mechanism for the formation of new Mannich derivatives as described in Scheme4:

Scheme 4: mechanism of Mannich bases derivatives
The FT-IR of Mannich bases derivatives includes the appearance of (N-H) stretching bands at (3436-3371) cm -1 . Also, FT-IR data have shown that the (CH) aliphatic bands at (2995-2927) cm -1 . FT-IR characteristic data are reported in Table 3. 1 H-NMR spectrum data Mannich derivative included appearance of singlet signal of (-CH2) protons at δ= (3.36) ppm, and singlet signal of (-NH) at δ= (9.85) ppm, 1 H-NMR spectrum data of Mannich bases derivative as listed in the Table 4. 13 CNMR data appearance δ at 36.7 of (CH 2 ) aliphatic. 13 CNMR data of Mannich bases derivative as listed in the Table 5.

Antioxidant activity DPPH Radical Scavenging Activity
Antioxidant activity and a significant scavenging percentage against the DPPH free radical were found in some of the newly synthesized compounds. As a result, the compounds that were tested demonstrated antioxidant capabilities and were chosen for future investigation. As a consequence, inhibitory concentrations (IC50) were calculated and recorded in Table 7. In this study, we employed Phongpaichit's antioxidant activity classification system, which is based on IC50 range values. For more information on these ranges, see Table 6. 1-The highest antioxidant activity and the lowest IC 50 values were found for compound (3A), (12A) and (4A) respectively. Table 10, the antioxidant activities and IC50 values of the other compounds varied, with a range of high, middle, and low activities.

Anticancer Activity
The cytotoxicity of three of the novel Manach bases was examined in vitro using the MTT assay against kidney cancer (HEK293 cancer cell line). To determine the cytotoxic effect of 6-(4-bromophenyl) imidazo [2, 1-b] thiazol-5-yl methyl -naphthalene-1-amine 4A on a kidney cancer cell line, the MTT method was applied (HEK293). The MTT Assay was used to evaluate cell viability and inhibition rate on a tumor cell line using different dosages of chemical 4A. In comparison to the normal cell line WRL-68, the proportion of treated cells that survived was calculated. On HEK293 cells, the cytotoxic effect of compound 4A in concentrations ranging from 12.5 to 400 mg.ml -1 resulted in a dose-dependent decrease in cell viability (Table 8). By raising the concentration of 4A chemicals, cell viability was lowered. The lowest HEK293 cell viability (percentage) was seen at 400 mg.ml -1 and the maximum HEK293 cell viability was observed at 12.5 mg.ml -1 . With an IC50 value of 101.9 g/ml, compound 4A showed strong cytotoxic action. The effect of compound 4A on WRI-68 cell line normalization yielded an IC50 of 122.6 mg.ml -1 (Figure 1).

Conclusion
New Mannich bases were synthesized using a well-known technique in this work. Because of their strong aromaticity, some of these novel bases showed good potential as an antioxidant activity and a high scavenging percentage against the DPPH free radical. Moreover, the applied procedure gets many of advantages such as high amount of yield, reactants and less time reaction. Other bases were also evaluated as cytotoxic agents and found to have a high cytotoxic effect with an IC50 value of 69.69 gm/ml. The antioxidant and anticancer effects of imidazo (2,1-b) thiazole derivatives were found to be in good agreement with other findings in the literature.