Claims:I CLAIM:
1. A process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain comprises of following steps;
a. adding a solution comprising of predetermined concentrations of aromatic dicarboxylic acid, diphenyl chlorophosphate, and LiCl in pyridine in an RB flask and continuously stirred at room temperature for predetermined time to form a mixture;
b. increasing the temperature of the said mixture of step(a) to 115°C and adding predetermined concentrations of 4,4'-diaminodiphenyl and CHBB followed by continuously stirring for predetermined time to form a suspension and;
c. cooling the said suspension of step(b) and pouring into methanol to form a precipitate followed by filtration, washing the said precipitate with methanol and drying in vacuum to form poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety.

2. A process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain comprises of following steps;
a. adding a solution comprising of 5mmol of aromatic dicarboxylic acid, 12 mmol of diphenyl chlorophosphate, and 10 mmol of LiCl in pyridine in an 100mL RB flask and continuously stirred at room temperature for 30 minutes to form a mixture;
b. increasing the temperature of the said mixture of step(a) to 115°C and adding 2.5 mmol of 4,4'-diaminodiphenyl and 2.5 mmol of 2,6-bis(4-hydroxybenzylidene)cyclohexanone (CHBB) followed by continuously stirring for 3 hours to form a suspension and;
c. cooling the said suspension of step(b) and pouring into methanol to form a precipitate followed by filtration, washing the said precipitate with methanol and drying in vacuum to form poly(ester-amides) possessing 2,6-bis(4-hydroxy benzylidene)cyclohexanone moiety.

3. The process as claimed in claim 1 and 2 wherein the said CHBB is synthesized by a process comprising of following steps:
a. adding p-hydroxy benzaldehyde (0.2 mol) in 100 mL of dry methanol in a container followed by drop wise addition of cyclohexanone (0.1 mol) under constant shaking to form a mixture;
b. drop wise addition of fuming sulphuric acid (5 mL) to the said mixture of step (a) under ice cold condition wherein colour changes to bright yellow initially, then dark green and finally pink and forms a precipitate;
c. filtering the said precipitate of step(b) followed by washing with aqueous methanol and recrystallising from aqueous methanol to form purified CHBB.

4. The process as claimed in claim 1 and 2 wherein the said aromatic dicarboxylic acid comprises of wherein
5. An antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain comprises of wherein prepared by a processes as claimed in claim 1 and 2
6. An antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain comprises of wherein

Dated this 24th day of February 2020


For KAVITHA ERRA KALAPPA
By her Patent Agent

Dr.B.Deepa
, Description:Form 2

THE PATENT ACT, 1970
(39 of 1970)
&
THE PATENT RULES, 2003
COMPLETE SPECIFICATION
(See section 10 and rule 13)

“SYNTHESIS, CHARACTERIZATION AND STUDY OF CYTOTOXICITY OF THERMOTROPIC LIQUID CRYSTALLINE POLY(ESTER-AMIDES) CONTAINING 2,6-BIS(3-METHOXYBENZYLIDENE)CYCLOHEXANONE MOIETY IN THE MAIN CHAIN”

in the name of KAVITHA ERRA KALAPPA an Indian National having address at Post-Graduate and Research Department of Chemistry, Presidency College (Autonomous), Chennai -600 005, Tamil Nadu, India.

The following specification particularly describes the invention and the manner in which it is to be performed
FIELD OF THE INVENTION:
The present invention relates to synthesis of antibacterial and anticancer activity exhibiting novel compounds. More particularly the present invention relates to a process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) and products thereof.
BACKGROUND OF THE INVENTION:
The thermotropic liquid crystalline poly(ester-amides) (TLCPs) incorporating flexible methylene groups, aromatic groups, and arylidene-keto moiety were reported.1-3 The bisbenzylidene cyclohexanone has both mesogenic and photo-active properties. They were found to be a potential mesogen that imparted thermotropic liquid crystalline property to the polymeric materials.4 Phase transitions of liquid crystals were reported by Marin et al. 5 by using Hot-stage Optical Polarized Microscopy (HOPM), Differential Scanning Calorimetry (DSC), and X-ray Diffraction (XRD) studies. The poly(ester-amides) containing 2,5-bis(benzylidene) cyclopentanone moiety were reported by Kannappan et al.6 Suhas Thatte and co workers,7 discussed the new and improved drug delivery methodology in the several polymer-drug systems. The first drug delivery application is reported using hydrogel in 1960.8 They found that these compounds were cytotoxic to a number of human tumours in vitro, particularly towards colon cancer and leukemic cells. Thus an attempt has been made to synthesize novel thermotropic liquid crystalline poly(ester-amides) compounds which exhibits antibacterial and anticancer activity.
References
1. Ling A-L, Hsu K-Y, Li C-H and Chang T-C 1992 Studies on thermotropic liquid crystalline polymers: Synthesis and properties of poly(ether ester amide)s Polymer 33 2217
2. Aharoni S H 1988 Hydrogen-bonded highly regular strictly alternating aliphatic-aromatic liquid crystalline poly(ester amides) Macromolecules 21 1941
3. Chen W-H, Chang T-C and Higashi F 1988 Liquid-crystalline copoly(ester amide)s prepared from hydroxybenzoic and 4-aminobenzoic acids J. Polym. Sci. Part A: Polym. Chem. 26 3269
4. Murali M and Samui A B 2010 Bisbenzylidene cycloalkanone: a versatile molecule as a polymer building block J. Mater. Chem. 20 2714
5. Marin L, Damaceanu M D and Timpu D 2009 New thermotropic liquid crystalline polyazomethines containing luminescent mesogens Soft Materials 7 1
6. Kannapan V and Reuben Jonathan D 2013 A study on the synthesis and bactericidal efficacy of certain poly(ester-amides) containing 2,5-bis(benzylidene)cyclopentanone moiety in the main chain J. Chem. Pharm. Res. 5 382
7. Thatte S, Datar K and Ottenbrite RM 2005 Perspectives on: polymeric drugs and drug delivery systems J. Bioactive Compatible Polymers 20 585
8. Wichterle O. and Lim D 1960 Hydrophilic gels for biological use Nature 185, 117
OBJECT OF THE INVENTION:
The main object of the present invention is to devise a novel process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain.
Another object of the present invention relates to antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain.
Yet another object of the present invention is to synthesis the novel thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain by direct polycondensation method.
Further object of the present invention is to utilize the synthesized novel thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain for antibacterial and anticancer studies.
BRIEF DESCRIPTION OF DRAWINGS:
Figure 1(a,b) depicts the FT-IR spectrum of random poly(ester-amides) (a) PBDT and (b) PBDA.
Figure 2(a,b) depicts the 1H NMR spectrum of random poly(ester-amides) (a) PBDT and (b) PBDA.
Figure 3(a,b) depicts the 1C NMR spectrum of random poly(ester-amides) (a) PBDT and (b) PBDA.
Figure 4 depicts the DSC thermograms of random poly(ester-amides)
Figure 5(a-c) depicts the Hot-stage optical polarized micrographs for (a) PBDT at 25ºC, crystalline phase of the mesogens (b) PBDT at 181ºC, crystal to crystal transition, and (c) PBDT at 186ºC, crystalline to nematic transition.
Figure 6(a-c) depicts Hot-stage optical polarized micrographs for (a) PBDI at 24ºC, crystalline phase of the mesogens (b) PBDI at 170ºC, crystal to crystal transition, and (c) PBDI at 182ºC, crystalline to nematic transition.
Figure 7(a-c) depicts Hot-stage optical polarized micrographs for (a) PBDP at 25ºC, crystalline phase of the mesogens (b) PBDP at 189ºC, crystal to crystal transition, and (c) PBDP at 230ºC, crystalline to nematic transition.
Figure 8(a-c) depicts Hot-stage optical polarized micrographs for (a) PBDA at 22ºC, crystalline phase of the mesogens (b) PBDA at 170ºC, crystal to crystal transition, and (c) PBDA at 200ºC, crystalline to nematic transition.
Figure 9(a-c) depicts Hot-stage optical polarized micrographs for (a) PBDAz at 22ºC, crystalline phase of the mesogens (b) PBDAz at 175ºC, crystal to crystal transition, and (c) PBDAz at 190ºC, crystalline to nematic transition.
Figure 10(a,b) depicts X-ray diffraction pattern of random poly(ester-amides) (a) PBDT and (b) PBDA.
Figure 11(a-e) depicts Scanning electron micrographs of random poly(ester-amides) (a) PBDT, (b) PBDI, (c) PBDP, (d) PBDA, and (e) PBDAz.
Figure 12 depicts A graphical representation of the polymer effect on cancer cells by % cell viability is shown
Figure 13 depicts affected MCF-7 cell line at different concentration (a) PBDT and (b) PBDAz
Figure 14(a-e) depicts inhibition effects of random poly(ester-amides) (a) PBDT (b) PBDI (c) PBDP (d) PBDA and (e) PBDAz on the growth of pathogenic bacteria. (Fig Label: a = 2 µg/µl, b = 4 µg/µl, c = 6 µg/µl, d = 8 µg/µl, e = 10 µg/µl, f = 12 µg/µl, and N = Standard (positive control) norflaxin) 10 µg/µl
SUMMARY OF THE INVENTION:
The present invention discloses a process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain and products thereof. Five thermotropic liquid crystalline poly(ester-amides) were synthesized by polycondensation method. The poly(ester-amides) were synthesized from varying dicarboxylic acids with a common diamine namely 4,4'-diamino benzene and a common diol namely 2,6-bis(4-hydroxybenzylidene))cyclohexanone. For qualitative characterization, viscosity measurements and solubility data were used for these synthesized poly(ester-amides). The spectroscopic techniques such as FT-IR, 1H NMR, 13C NMR were performed to investigate the microstructural features of these synthesized poly(ester-amides). The thermal phase transition behavior of these poly(ester-amides) were studied by Differential Scanning Calorimetry (DSC) and Hot-stage Optical Polarized Microscopy (HOPM). The degree of crystallinity was assessed by X-ray diffraction (XRD) patterns. Scanning Electron Microscopic (SEM) technique was used to illustrate the morphology of these poly(ester-amides). The synthesized copolymer was subjected into in vitro anti-cancer activity studies against human breast cancer (MCF-7) cell line and also these copolyesters displayed potential bactericidal activity against pathogenic bacteria.
DETAILED DESCRIPTION OF THE INVENTION:
The present invention discloses a process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) and products thereof.
A series of five thermotropic liquid crystalline poly(ester-amides) containing bisbenzylidenecyclohexanone moiety in the polymer backbone was synthesized. In the current work, poly(ester-amides) are a category of polymeric materials which contains both ester and amide linkages and are synthesized by the copolymerization of a diacid with that of a diamine and a diol.

Experimental
Chemicals
Aldrich samples of diphenyl chlorophosphate, terephthalic acid, isophthalic acid, phthalic acid, adipic acid, azelaic acid, and 4,4'-diamino diphenyl were used as received. Lithium chloride
(SD Fine), 4-hydroxy benzaldehyde, and cyclohexanone (Fluka) were used as received. Merck sample of pyridine, Merck LR sample of methanol, SD-Fine AR samples of N,N-dimethyl acetamide (DMAc ) and N,N-dimethyl formamide (DMF), and Aldrich spectral grade DMSO-d6 were used.
Preparation of monomer
Synthesis of arylidene-keto diol: The arylidene-keto diol namely 2,6-bis(4-hydroxybenzylidene)cyclohexanone (CHBB) was synthesized by the method reported in literature.7
Preparation of 2,6-bis(4-hydroxybenzylidene)cyclohexanone (CHBB): In a 250 mL conical flask, a solution containing p-hydroxy benzaldehyde (0.2 mol) in 100 mL of dry methanol was taken. To this solution, cyclohexanone (0.1 mol) was added drop wise and the mixture was shaken well. Then with external cooling in an ice bath, fuming sulphuric acid (5 mL) was added drop wise so that an exothermic reaction took place. The reaction mixture turned bright yellow initially, then dark green and finally pink. The precipitated diol was filtered and washed with aqueous methanol. It was filtered and recrystallised from aqueous methanol. Yield: 95%
(m.p.: 195 oC). It is represented in Scheme 1.

Scheme 1. Synthesis of 2,6-bis(4-hydroxybenzylidene))cyclohexanone (CHBB).
Synthesis of poly(ester-amides): Two methods were employed to synthesize the poly(ester-amides) which depends upon the type of dicarboxylic acid monomer (aromatic or aliphatic).
The procedure for the synthesis of a typical poly(ester amide) derived from aromatic dicarboxylic acid is represented here. Three poly(ester-amides) were prepared by the direct polycondensation method using diphenyl chlorophosphate (DPCP) as the condensation agent in the mole ratio 1:1:2 of a diamine, a diol and a diacid in pyridine solution. This method avoids the tedious preparation of acid chloride and hence, yield of the polymer is high.
In a 100 mL round-bottomed flask, a solution containing aromatic dicarboxylic acid
(5 mmol), DPCP (12 mmol), and LiCl (10 mmol) in pyridine was continuously stirred at room temperature for 30 minutes. Then the temperature was raised to 115°C and 4,4'-diamino diphenyl (2.5 mmol) and the diol (2.5 mmol) were added and the stirring was continued for 3 hours. The reaction mixture was cooled and poured into methanol. The poly(ester-amide) was precipitated, filtered and washed with methanol and dried in vacuum. It is represented in Scheme 2.

Scheme 2. Synthesis of poly(ester-amides) PBDT (I), PBDI (II) and PBDP (III) derived from aromatic dicarboxylic acid
In Table 1, the various poly(ester-amides) prepared were listed along with the polymer code, percentage of yield and inherent viscosities.

Table 1. List of monomers used, polymer code of poly(ester-amides) with percentage of yield and inherent viscosities (?inh)
S. No. Common diamine: diphenyl diamine Polymer Code Yield (%) ?inh (dL/g)
Diol Diacid / Diacid chloride
1. CHBB Terephthalic acid PBDT - I 77 1.20
2. CHBB Isophthalic acid PBDI - II 75 1.17
3. CHBB Phthalic acid PBDP - III 72 1.08
4. CHBB Adipoyl dichoride PBDA - IV 58 0.56
5. CHBB Azelaoyl dichloride PBDAz - V 60 0.58
CHBB: 2,6-(bis(4-hydroxybenzylidene))cyclohexanone
Characterization methods
Solubility: In a small stoppered test tube, about 50 mg of the polymer containing 1 mL of the solvent was taken and kept for 24 hours with occasional shaking. In various solvents, the solubility was tested qualitatively.
Viscosity: The inherent viscosity (?inh) was determined in DMAc solution by using Ubbelohde viscometer in which the pure solvent had a flow rate of 470 seconds at 30°C. In each case, dry poly(ester-amide) sample (25 mg) was dissolved in 25 mL of DMAc and kept aside for 12 hours with occasional shaking. From the flow time measurement at 30°C, the ?inh was calculated.
Fourier Transform Infrared (FT-IR) spectroscopy: Shimadzu FT-IR instrument was used to record the FT-IR spectra. The sample was taken in the form of potassium bromide pellet.
1H and 13C NMR Spectra: Bruker AVANCE III 500 MHz instrument was used to record the 1H and 13C NMR spectra. DMSO-d6 was used as solvent with TMS as internal reference.
Differential Scanning Calorimetry (DSC): NETZSCH DSC 200F3 differential scanning calorimeter using 5mg samples under nitrogen atmosphere was used for thermal analysis of these poly(ester-amides) at a heating rate of 10°C/min.
Hot-stage Optical Polarized Microscopy (HOPM): The polarized optical microscopic technique was used to identify the mesophases using Olympus BX51P model with a source of 100W halogen lamp housing by using Linksys 32 software for Hot Stage temperature controller.
X-Ray Diffraction (XRD): GE-Inspection Technology Diffractometer System XRD 3003 TT model made in Germany with a source of copper target operating voltage 40 Kv 300 mAo current rate was used to record X-ray diffraction measurements. They were taken to assess the degree of crystallinity of these powdered poly(ester-amides).
Scanning Electron Microscopy (SEM): Hitachi S-3000 Hz scanning electron microscopy was used to investigate the morphology of these poly(ester-amides).
Cytotoxic anticancer evaluation of synthesized polymer.
MTT assay (MTT = 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl--tetrazolium bromide = 5mg/ml in PBS). The cancer activity of samples on MCF7 cell was determined by the MTT assay. Cells (1 × 105/well) were plated in 0.2 ml of medium/well in 96-well plates. Incubate at 5% CO2 incubator for 72 hours. Then, added various concentrations of the samples in 0.1% DMSO for 48hrs at 5% CO2 incubator. View the images under Inverted microscope 40X and take the photos. After removal of the sample solution and 20µl/well MTT reagent was added. Viable cells were determined by the absorbance at 540nm. 50% inhibition of cell viability (IC50 value) was determined graphically. The effect of the samples on the proliferation of MCF7 cells was expressed as the % cell viability, using the following formula:

Calculation

Antibacterial Activity (Agar Disc Diffusion Method)

Preparation of inoculums
Stock cultures were maintained at 4°c on slant of nutrient agar. Active cultures for experiments were prepared by transferring a loop full of cells from the stock cultures to test tubes of nutrient broth for bacteria that were incubated at 24hrs at 37ºc. The Assay was performed by agar disc diffusion method.

Determination of antibacterial activity
Antibacterial activity of sample was determined by disc diffusion method on Muller Hinton agar (MHA) medium. The Muller Hinton Agar medium was weighed as 3.8gms and dissolved in 100ml of distilled water and add 1gm of agar. Then the medium is kept for sterilization. After sterilization the media was poured in to sterile petriplates and were allowed to solidify for 1hr. After the medium was solidified, the inoculums were spread on the solid plates with sterile swab moistened with the bacterial suspension. Discs were prepared with 20 µl sample of respective concentrations (5, 10, 15, 20, 25 and 30 µg) and positive control 10 µl (10 µg) norflaxin and placed on MHA plates. These plates were incubated for 24 hrs at 37ºc. Then the microbial growth was determined by measuring the diameter of zone of inhibition.

Results and Discussion
Solubility
These poly(ester-amides) were found to be soluble in highly polar solvents such as DMAc, DMSO, and TFA, partially soluble in moderately polar solvents and insoluble in non-polar solvents such as benzene and hexane. This is because of the inter-molecular interactions of polar solvents with ether linkages of the polymer backbone. In Table 2, the results of the solubility of these poly(ester-amides) are represented.
Table 2. Solubility of the poly(ester-amides) in common organic solvents
S.No. Polymers Hexane Benzene CHCl3 EtOAc THF Acetone DMF DMAc CH3CN TFA DMSO
1 I -- -- -- -- -- +- +- ++ +- ++ ++
2 II -- -- -- -- +- ++ ++ ++ ++ ++ ++
3 III -- -- -- -- +- ++ ++ ++ ++ ++ ++
4 IV -- -- +- +- ++ ++ ++ ++ ++ ++ ++
5 V -- -- +- +- ++ ++ ++ ++ ++ ++ ++
++ =Freely Soluble; -- = Insoluble; +- = Partially soluble Viscosity
In Table 1, the ?inh values of all these five poly(ester-amides) are represented and were found to be in the range of 0.56–1.20 dL/g. It is noted that the poly(ester-amides) synthesized from aromatic dicarboxylic acids have higher ?inh values than those prepared from aliphatic diacid chloride. Since the data shows higher viscosity values, they are reasonably of higher molecular weight.
Spectral studies
By FT-IR spectra, the ester and amide functional groups present in the poly(ester-amide) chain were identified. It showed characteristic absorption at = 1630–1750 cm-1 due to ester and amide C=O stretching frequency and an absorption at = 3240–3380 cm-1 due to the amide N-H stretching frequency. They are represented in Figures 1(a) and 1(b) for PBDT and PBDA, respectively.
By 1H and 13C NMR spectra, the structural units present in the poly(ester-amide) chain were identified. A singlet was appeared in the range ??9.5–10.15 ppm is due to the presence of secondary amide proton. Due to the presence of aromatic protons, peaks were observed in the range of ??6.5–8.54 ppm. They are represented in Figures 2(a) and 2(b) for PBBT and PBBA, respectively.
In the 13C NMR spectra of the poly(ester-amides), the signal in the range of ? 188–200 ppm is due to the carbonyl carbon of the ???-unsaturated ketone. Due to the carbonyl carbon of the amide and ester groups, the signals in the range of ? 171–185 ppm and ? 160 –170 ppm were appeared which indicates the formation of poly(ester-amide). They are represented in Figures 3(a) and 3(b) for PBDT and PBDA, respectively.
Thermal analysis and phase behavior
By using Differential Scanning Calorimetry (DSC) and Hot-stage Optical Polarized Microscopy (HOPM), the thermal and phase behaviors of the five poly(ester-amides) were investigated.
Differential Scanning Calorimetry (DSC): In Table 3, the glass transition temperature (Tg), crystal to crystal transition (TK1?K2), melting temperature (Tm), isotropization temperature (Ti), and liquid crystalline range (?T) obtained from DSC are tabulated.
Table 3. Differential Scanning Calorimetry (DSC) data of poly(ester-amides)
S.No. Polymers Differential Scanning Calorimetry (DSC)
Tg (ºC) TK1?K2 (ºC) Tm (ºC) Ti (ºC) ?T
1 I 22.5 180.0 330.0 460.0 130.0
2 II 33.5 178.5 336.0 438.5 102.5
3 III 23.3 163.3 325.8 393.3 67.5
4 IV 32.9 175.4 312.9 477.9 165.0
5 V 25.4 180.4 300.4 390.4 90.0

In Figure 4, the DSC thermograms of all poly(ester-amides) are shown. By analyzing the DSC data, we could interpret that the poly(ester-amides) I and III synthesized from aromatic dicarboxylic acids have lower glass transition temperature (Tg) values than those synthesized from aliphatic diacid chloride monomers. In these polymers, an exothermic peak appears before melting transition, which seems to be related to crystal to crystal transition resulted from different crystalline polymorphs. The liquid crystalline range (????of the poly(ester-amide) derived from adipic acid monomer is broader than compared to the poly(ester-amide) derived from other monomer azelaic acid. This may be due to effective molecular packing by coplanar geometry. Also, the liquid crystalline range (?T) of the poly(ester-amide) derived from isophthalic acid aromatic acid monomer is broader than compared to the poly(ester-amide) derived from other monomers which may be due to the rigidity of m-phenylene moiety while comparing with o-phenylene or p-phenylene moiety in the polymer chain.
Hot-stage Optical Polarized Microscopy (HOPM): Mesophase identification of poly(ester-amides) have been achieved by Hot-stage Polarized Optical Microscopy (HOPM). From Figures 5 to 9, the different textures of poly(ester-amides) have been identified at different temperatures. The poly(ester-amides) exhibited high birefringence when it analyzed with a HOPM..In Figures 5(a) to 9(a), crystalline phases of the mesogens has been shown. By increasing the temperature, crystal to crystal transition was observed. They are shown in Figures 5(b) to 9(b). This on further heating, a crystalline to nematic transition of the mesogens takes place. They are shown in Figures 5(c) to 9(c). On further heating of these poly(ester-amides), the nematic phase lost its birefringence and transformed to an isotropic phase where there is no texture identification i.e. it appears as black domain.
X-Ray Diffraction studies
In Figures 10(a) and 10(b), the X-Ray diffraction pattern of poly(ester-amides) PBDT and PBDA has been shown that indicates the semi-crystallinity of these polymers with an amorphous background which may be due to the presence of carbonyl and C=C groups. In the region 2? = 10 – 50°, these poly(ester-amides) showed few reflection peaks that confirms the presence of semi-crystallinity.
Morphological study by SEM
In Figure 11, the SEM images of the poly(ester-amides) have been shown that illustrates the morphology of these poly(ester-amides).
Cytotoxic Anticancer Evaluation of Synthesized Polymer
Viable cells were determined by the absorbance. Measurements were performed and the concentration required for a 50% inhibition of viability (IC50) was determined graphically. The anti cancer activity and anti bacterial activity have been studied by Chitra and Roop Singh. The absorbance was measured with a UV-Spectrophotometer using wells without sample containing cells as blanks. The effect of the polymer on the proliferation of MCF-7 was expressed as the % cell viability. In Figure 12, a graphical representation of the polymer effect on cancer cells by % cell viability is shown.The affected MCF-7 cell line at different concentration was shown in Figure 13. IC of the polymer was determined and was shown in Table 4. The study on anti-cancer activity containing arylidene ketone moiety was studied by Rajam and Roop Singh.
Table 4. MCF7 Cell line
S. No.
Concentration
(µg/mL) Absorbance
(540 nm) % Cell viability
PBDT PBDAz
1 100 0.02 1.7 7.1
2 50 0.05 4.2 13.2
3 25 0.11 9.4 25.5
4 12.5 0.27 23.0 47.9
5 6.25 0.56 47.8 64.2
6 3.12 0.83 70.9 87.7
7 Control cells 1.17 100 100
Bactericidal Study
The antibacterial activity of all the five poly(ester-amides) were assayed against Escherichia coli, Salmonella typhi, Staphylococcus aureus, Proteus vulgaris, Proteus mirabilis, and Klebsiella pneumonia by disc diffusion method. The zone of inhibition for each concentration against all the test bacteria is depicted in table 6. The zone of inhibition for the standard positive control norflaxin disc 10 µg/µl is shown in table 6. They are represented in Figures 14(a) to 14(e). Analysis of the data in table 5suggests that the poly(ester-amide) derived from aliphatic diacid choride exhibited substantial antibacterial activity towards all the test bacteria E. coli, S. typhi, S. aureus, P. vulgaris, P. mirabilis, and K. pneumonia in all the concentrations. S. Kothai and coworkers reported on the antibacterial activity of certain copolyesters derived from aliphatic diacid chloride. Similar observation was reported by V. Kannapan and D. Reuben Jonathan in a series of copolyesteramides derived from arylidene diols.
Comparing the poly(ester-amides) derived from aromatic dicarboxylic acids, the poly(ester-amide) derived from phthalic acid exhibited greater antibacterial activity. Comparing the poly(ester-amides) derived from aliphatic dicarboxylic acids,the poly(ester-amide) derived from azelaic acid exhibited greater antibacterial activity. The bacteria Klebsiella pneumonia injected in the plate PBDAz has no strain effect.
Table 6. Inhibition effects of the five poly(ester-amides) on the growth of Escherichia coli, Salmonella typhi, Staphylococcus aureus, Proteus vulgaris, Proteus mirabilis, and Klebsiella pneumonia
Plate No. Polymer code / Microorganisms Zone of inhibition (mm)
2 µg/µl 4 µg/µl 6 µg/µl 8 µg/µl 10 µg/µl 12 µg/µl N
10 µg/µl
PBDT
1 E. coli 0 0 0 0 0 6.7 6.1
2 S. typhi 0 0 0 0 0 6.3 6.2
3 S. aureus 0 0 0 0 0 6.1 6.1
4 P. vulgaris 0 0 0 0 0 0 6.1
5 P. mirabilis 0 0 0 0 6.1 6.3 6.1
6 K. pneumonia 0 0 0 0 0 6.5 6.2
PBDI
1 E. coli 0 0 0 0 6.5 7 7
2 S. typhi 0 0 0 0 0 6.1 7
3 S. aureus 0 0 0 0 0 6.1 7.2
4 P. vulgaris 0 0 6.1 6.3 6.5 6.7 6.2
5 P. mirabilis 0 0 0 0 0 0 6.1
6 K. pneumonia 0 0 0 0 0 0 6.1
PBDP
1 E. coli 6.2 6.7 6.8 6.9 7.1 7.3 18
2 S. typhi 0 0 6.3 6.5 6.9 7.5 14
3 S. aureus 0 0 0 0 0 6.4 15
4 P. vulgaris 6.4 6.9 7.1 7.4 7.6 8.1 15
5 P. mirabilis 6.2 6.5 6.8 8.1 8.7 9.6 14
6 K. pneumonia 0 6.4 6.7 6.9 7.2 7.6 17
PBDA
1 E. coli 6.3 6.5 6.9 7.3 7.6 7.9 21
2 S. typhi 6.4 6.6 6.8 7.0 7.4 7.6 14
3 S. aureus 6.2 6.4 6.9 7.1 7.5 8.2 20
4 P. vulgaris 7.2 7.6 8.1 8.9 9.7 10.5 13
5 P. mirabilis 7.3 7.5 7.9 8.2 9.1 9.8 13
6 K. pneumonia 6.1 6.3 6.4 6.8 6.9 7.2 15
PBDAz
1 E. coli 6.2 6.4 6.6 6.8 7.3 7.7 16
2 S. typhi 7.3 7.6 7.9 8.4 9.2 9.7 24
3 S. aureus 7.8 8.1 8.9 9.8 10.3 10.8 26
4 P. vulgaris 6.8 7.6 7.9 8.4 9.3 9.7 24
5 P. mirabilis 7.4 7.8 8.1 8.5 9.1 9.6 14
6 K. pneumonia 0 0 0 0 0 0 13

To conclude a series of five poly(ester-amides) have been synthesized successfully by direct polycondensation method. Solubility studies reveal that these poly(ester-amides) are highly soluble in polar organic solvents and the viscosity studies reveal that these are of high molecular weight. These are characterized by FT-IR, 1H-NMR, and 13C-NMR spectral studies that supports the structural assignments. From the thermal studies such as DSC and HOPM, the phase transition and texture behavior of these poly(ester-amides) were identified. The degree of crystallinity were confirmed by X-ray diffractograms. The morphology of these five poly(ester-amides) are illustrated by SEM. Because of their high thermal stability as thermotropic liquid crystals, they might be utilized for flame retardant applications and optical device technology.
In one of the preferred embodiment the present invention shall disclose a process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain. The process of synthesis comprises of following steps;
a. adding a solution comprising of predetermined concentrations of aromatic dicarboxylic acid, diphenyl chlorophosphate, and LiCl in pyridine in an RB flask and continuously stirred at room temperature for predetermined time to form a mixture;
b. increasing the temperature of the mixture of step(a) to 115°C and adding predetermined concentrations of 4,4'-diaminodiphenyl and CHBB followed by continuously stirring for predetermined time to form a suspension and;
c. cooling the suspension of step(b) and pouring into methanol to form a precipitate followed by filtration, washing the precipitate with methanol and drying in vacuum to form poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety.

In another preferred embodiment the present invention shall disclose a process of synthesis of antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain. The process of synthesis comprises of following steps;
a. adding a solution comprising of 5mmol of aromatic dicarboxylic acid, 12 mmol of diphenyl chlorophosphate, and 10 mmol of LiCl in pyridine in an 100mL RB flask and continuously stirred at room temperature for 30 minutes to form a mixture;
b. increasing the temperature of the mixture of step(a) to 115°C and adding 2.5 mmol of 4,4'-diaminodiphenyl and 2.5 mmol of 2,6-bis(4-hydroxybenzylidene)cyclohexanone (CHBB) followed by continuously stirring for 3 hours to form a suspension and;
c. cooling the suspension of step(b) and pouring into methanol to form a precipitate followed by filtration, washing the said precipitate with methanol and drying in vacuum to form poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety.

According to the invention in the process described above, the CHBB is synthesized by a process comprising of following steps:
a. adding p-hydroxy benzaldehyde (0.2 mol) in 100 mL of dry methanol in a container followed by drop wise addition of cyclohexanone (0.1 mol) under constant shaking to form a mixture;
b. drop wise addition of fuming sulphuric acid (5 mL) to the mixture of step (a) under ice cold condition in which colour changes to bright yellow initially, then dark green and finally pink and forms a precipitate;
c. filtering the precipitate of step(b) followed by washing with aqueous methanol and recrystallising from aqueous methanol to form purified CHBB.

In accordance with the invention, in the process described above, the aromatic dicarboxylic acid comprises of in which
In yet another preferred embodiment the present invention shall disclose an antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain comprises of in which prepared by a processes described

In further preferred embodiment the present invention shall disclose an antibacterial and anticancer activity exhibiting thermotropic liquid crystalline poly(ester-amides) possessing 2,6-bis(4-hydroxybenzylidene)cyclohexanone moiety in main chain comprises of in which

Certain modifications and improvements will occur to those skilled in the art upon a reading of the foregoing description. The above-mentioned details are provided to serve the purpose of clarifying aspects of the invention and it will be apparent to one skilled in the art that they do not serve to limit the scope of the invention. All modifications and improvements have been deleted herein for the sake of conciseness and readability but are properly within the scope of the present invention. It is understood that the foregoing detailed description is given merely by way of illustration and that modification and variations may be made therein without departing from the spirit and scope of the invention