Synthetic Cationic Cholesteric Liquid Crystal Polymers

Abstract

We report the synthesis of six multifunctional cationic cholesteric liquid crystals polyesters functionalized with choline, amine, and amide groups to obtain new chemical formulations involving macromolecular features with new properties added to those of precursor chiral cholesteric polyesters. They are designed as PTOBDME-choline [(C34H36O8)n─C5H13N]; PTOBEE-choline [(C26H20O8)n─C5H13N]; PTOBDME-ammonium [(C34H36O8)n─C5H13N]; PTOBEE-ammonium [(C26H20O8)n─C5H13N]; PTOBUME-amide (C33H33O9N)n; and PTOBEE-amide (C26H19O9N)n. Structural characterization is performed by NMR. Thermal behavior is studied by thermogravimetry (TG) and differential scanning calorimetry (DSC), showing all the polymers endothermic transition from crystal phase to liquid crystal mesophase. Chirality is determined by optical rotatory dispersion (ORD). The cationic cholesteric liquid crystal polymers described here have proved to act as nonviral vectors in gene therapy, transfecting DNA to the nucleus cell.

1. Introduction

Cholesteric liquid crystal polyesters have received much attention in the last few years for their interesting chemical, optical, mechanical, and biological properties. Due to their anisotropic formulation and amphiphilic nature, their molecules are able to self-associate and/or aggregate in blocks to form species with supramolecular ordered structure, which presents desirable material properties.

Two cholesteric liquid crystal polyesters, named PTOBDME and PTOBEE in Figure 1, were obtained by polycondensation reaction. Although only racemic materials were used in their synthesis, a cholesteric, chiral morphology, theoretically unexpected, was found. Evidence of this was obtained when a white solid, recrystallized, as the second fraction, from toluene mother liquor after the filtration of the polymer, was identified as ─PTOBDME, with [α]²⁵₅₈₉ = −1.43 (1.538g/100 ml, toluene) [1, 2] and ─PTOBEE, with a value of [α]²⁵₅₈₉ = −2.33 (0.0056 mol/l, toluene) [3], respectively. The synthetic method [4], based on the previously reported by Bilibin [5], leads to obtain two or more fractions of different kinetic rates, with different enantiomeric excess. Not always, the enantiomer in excess is the same.

Figure 1.

Monomeric unit of cholesteric liquid-crystalline PTOBEE (m = 1) and PTOBDME (m = 9). The three different zones of the monomer: mesogen, spacer, and flexible side chain are indicated. The asterisk indicates the chiral center. Torsion angle ϕ, along the bond containing the asymmetric carbon atom, is indicated. Aromatic-end acid and aliphatic-end alcoholic groups are also specified.

We are interested in the molecular design and chemical modifications of these multifunctional cholesteric liquid crystals to obtain new chemical formulations involving macromolecular features with new properties added. Our main interest being to introduce cationic charge, hence favoring the creation of hydrogen bonds, through intra and intermolecular interactions, giving secondary structures with long-range supramolecular order, and enabling to interact with molecules of interest, such as biological molecules (lipids, DNA, and oligonucleotides) and metal surfaces. The functional groups selected to be introduced at the end of the main chains were Choline [─CH₂─CH₂─N─(CH₃)₃] and ammonium [─CH₂─CH₂─CH₂─NH─(CH₃)₂] and amide groups (─CONH₂) at the end of the lateral hydrophobic chains.

The new synthetized cationic polymers reported here have proved to be able to interact with negatively charged DNA, forming polyplexes, which are able to condense and successfully transfect the new DNA into the nucleus cell, protecting it from damage during the transfection process, acting as nonviral vectors in Gene Therapy [6, 7]. Besides, they are sensitive to pH changes, acting as polycationic efficient transfection agents possessing substantial buffering capacity below physiological pH. These vectors have shown to deliver genes as well as oligonucleotides, both in vitro and in vivo, by protecting DNA from inactivation by blood components. Their efficiency relies on extensive endosome swelling and rupture that provides an escape mechanism for the polycation/DNA complexes [8].

2. Materials

The new cholesteric liquid crystal polymers so designed have been synthesized as follows: PTOBDME-choline [(C₃₄H₃₆O₈)_n─C₅H₁₃N]; PTOBEE-choline [(C₂₆H₂₀O₈)_n─C₅H₁₃N]; PTOBDME-ammonium [(C₃₄H₃₆O₈)_n─C₅H₁₃N]; PTOBEE-ammonium [(C₂₆H₂₀O₈)_n─C₅H₁₃N]; PTOBUME-amide [(C₃₃H₃₃O₉N)_n; and PTOBEE-amide (C₂₆H₁₉O₉N)_n.

2.1. Synthesis of cholesteric PTOBDME-choline [(C₃₄H₃₆O₈)_n─C₅H₁₃N]

Poly[oxy(1,2-dodecane)oxycarbonyl-1,4-phenylene-oxy-1,4-terephthaloyl-oxy-1,4-phenylene-carbonyl]-oxy-N, N, N-trimethylethan-1-ammonium (Choline) chloride, II in Figure 2, was obtained through polycondensation reaction between: 4,4′-(terephthaloyldioxydibenzoic chloride) TOBC, I in Figure 2, the racemic mixture of DL-1,2-dodecanediol, and choline chloride. Notation similar to precursor cholesteric liquid crystal PTOBDME [1, 2] is used.

Figure 2.

Synthetic process of cholesteric liquid-crystalline PTOBDME-choline (m = 9) (II) and PTOBEE-choline (m = 1) (III). Monomeric units are indicated, together with aliphatic end groups and choline aromatic end groups. The asterisks indicate the chiral centers (¹²C*) and (⁴C*), respectively. Torsion angles ϕ, along (¹²C─¹¹C) and (⁴C─³C) bonds, respectively, are shown.

2.2. Synthesis of cholesteric PTOBEE-choline [(C₂₆H₂₀O₈)_n─C₅H₁₃N]

The structure of Poly[oxy(1,2-butane)oxycarbonyl-1,4-phenylene-oxy-1,4-terephthaloyl-oxy-1,4-phenylene-carbonyl]-oxy-N,N,N-trimethylethan-1-aminium (choline) chloride is shown in III of Figure 2. The polycondensation included DL-1,2-butanediol. Notation similar to precursor cholesteric liquid crystal PTOBEE [3, 4] is used.

2.2.1. Preparation of PTOBDME-choline and PTOBEE-choline

The dichloride, TOBC, was obtained by reaction between thionyl chloride and 4,4′-(terephthaloyldioxydibenzoic) acid (TOBA), previously synthesized from terephthaloyl chloride and 4-hydroxybenzoic acid [5].

The polycondensation reaction between TOBC and the racemic mixture, the corresponding glycol, takes place in presence of 1/7 equimolecular choline chloride. The preparation of these compounds was performed on melting due to the insolubility of choline chloride in the solvents used in the synthesis of PTOBDME or PTOBEE precursors, diphenyl oxide, or chloronaphthalene.

A mixture of 0.0054 mol of the glycol, either DL-1,2-dodecanediol or DL-1,2-butanediol, from Flucka Chemie GmBH (Buchs, Switzerland) and 0.000775 mol of choline chloride from Sigma-Aldrich Chemie GmBH (Steinheim, Germany) were placed into a flask of 50 ml contained in a bath with a high-temperature transfer agent, while a current of dry nitrogen from Praxair (Madrid, Spain) was used to purge the system at room temperature and then maintained in the rest of the reaction. The mixture was stirred and heated to 110°C to whole dissolution of the choline chloride into diol. The bath was cooled to 80°C, and 0.0062 mol of TOBC was added; this temperature was maintained for 15 minutes. The bath was heated up to 190°C, the mixture was melted, and emission of HCl was observed. After 60 minutes, 15 ml of chloronaphthalene from Sigma-Aldrich Chemie GmBH (Steinheim, Germany) was added. The reaction mix was maintained into the solvent stirring at 190°C for 150 minutes. Then, it was poured into 150 ml of toluene from Merck KGaA (Darmstadt, Germany), decanting PTOBDME-choline or PTOBEE-choline, respectively, which was filtered, washed with ethanol, and vacuum dried.

2.3. Synthesis of cholesteric PTOBDME-ammonium [(C₃₄H₃₆O₈)_n─C₅H₁₃N]

The structure of Poly[oxy (1,2-dodecane)-oxy-carbonyl-1,4-phenylene-oxy-1,4-terephthaloyl-oxy-1,4-phenylene-carbonyl]-oxy-3-dimethyl amine-1-propyl choride is shown in II of Figure 3.

Figure 3.

Synthetic process of cholesteric liquid-crystalline PTOBDME-ammonium (II), and PTOBEE-ammonium (III). Monomeric units are indicated, together with aliphatic end groups and ammonium aromatic end groups. The asterisks indicate the chiral centers (⁴C*) and (¹²C*), respectively. Torsion angles ϕ, along (⁴C─³C) and (¹²C─¹¹C) bonds, are shown.

2.4. Synthesis of Cholesteric PTOBEE-ammonium [(C₂₆H₂₀O₈)_n─C₅H₁₃N]

The structure of Poly[oxy(1,2-butane)oxycarbonyl-1,4-phenylene-oxy-1,4-terephthaloyl -oxy-1,4-phenylene-carbonyl]-oxy-3-dimethylamine-1-propyl choride is shown in III of Figure 3.

2.4.1. Preparation of PTOBMDE-ammonium and PTOBEE-ammonium

PTOBDME-ammonium chloride, II in Figure 3, and PTOBEE-ammonium chloride, III in Figure 3, were obtained through polycondensation reaction between 4 and 4′-(terephthaloyldioxydibenzoic chloride) TOBC, I in Figure 3, and the racemic mixture of DL-1,2-dodecanediol and DL-1,2-butanediol, respectively, and then reaction with 3-Dimethylamino-1-propanol. Notation of cholesteric liquid crystal PTOBDME and PTOBEE precursors is used. Next, a typical preparation of PTOBDME-ammonium chloride is shown.

Into a flask of 50 ml, TOBC (0.0079 mol) and 1,2-dodecanediol (0.0079 mol) from Flucka Chemie GmBH (Buchs, Switzerland) and diphenyl oxide (19.7 ml) of from Sigma-Aldrich Chemie GmBH (Steinheim, Germany) were mixed, while the system was purged with stream of dry nitrogen from Praxair (Madrid, Spain), for 30 min at room temperature. Then, while maintaining the gas current, the flask was transferred to a bath at 200°C for 2 hours; since the liberation of HCl is still observed, the temperature of the bath was descended to 160°C, the polycondensation was stopped and was not observed HCl formation. 3-Dimethylamino-1-propanol (0.2 ml, 0.00156 mol) was added to the reaction mix, and the liberation of HCl returned again. After 2 hours, the reaction finished. The result of the polycondensation reaction was poured into 200 ml of toluene from Merck KGaA (Darmstadt, Germany), decanting PTOBDME, which was filtered, washed with ethanol, and vacuum dried.

2.5. Synthesis of cholesteric PTOBUME-amide [(C₃₃H₃₃O₉N)_n

Poly[oxy(1,2-undecan-11-amidyl)-oxycarbonyl-1,4-phenylene-oxy-1,4-terephthaloyl-oxy-1,4-phenylene-carbonyl], VII in Figure 4, was obtained through polycondensation reaction between 4 and 4′-(terephthaloyldioxydibenzoic chloride) TOBC and the racemic mixture of DL-10,11-dihydroxyundecanemide (V in Figure 4) [11, 12, 13, 14, 15]. Similar notation has been used than with precursor cholesteric liquid crystal PTOBDME, Figure 1.

Figure 4.

Synthetic method of cholesteric liquid-crystalline PTOBUME-amide. The asterisk indicates the chiral center (¹¹C*). Torsion angle ϕ, along ¹⁰C─¹¹C bond.

2.5.1. Preparation of undec-10-enoyl chloride (II in Figure 4)

To a stirred solution of 0.118 mol of undec-10-enoic acid in 100 ml of toluene, at 25°C, 0.078 mol of oxalyl chloride was added during 30 minutes. The solution was stirred for 30 minutes after emission of HCl gas had completed. The mixture reaction was concentrated to about half the initial volume by using a vacuum pump equipped with a sodium hydroxide trap. This solution was used directly to prepare undec-10-enamide (III in Figure 4).

2.5.2. Preparation of undec-10-enamide

A NH₃ gas stream was used to purge the stirred solution of undec-10-enoyl chloride, cooled in a bath of dry ice/acetone. The NH₃ stream was produced by boiling to reflux ammonia solution generated by reaction between 100 g of ClHN₄ solved into 300 ml of H₂O and 76 g of NaOH solved in 50 ml of water at 10°C. The reflux condenser and a NaOH trap were connected between the ammonia solution and the mixture reaction to prevent moisture. After 30 minutes of reaction, when a white solid had precipitated and HCl gas emission was not observed, the reaction flask was allowed to warm to room temperature. The mixture reaction was concentrated to a residue on a rotatory evaporator. The solid was partitioned between 10% aqueous sodium hydroxide and dichloromethane, and the aqueous phase was washed three times with additional dichloromethane. The combined dichloromethane extracts were washed with brine, dried with anhydrous sodium sulfate, and concentrated to a solid on a rotatory evaporator [12, 13, 14, 15]. The solid was recrystallized in a mix chloroform/hexane (1:1) to give pure undec-10-enamide—yield (80%) and melting point 87°C (III in Figure 4).

¹H NMR (CDCl₃ 300 MHz, δ; (ppm)): δ 7.19, (dd, 2H) (7.03), δ 5.80, (m, 1H) (5.82), δ 5.32-5.22, (bs, 1H,) (5.13), δ 4.96, (dd, 1H J = 7.2 Hz) (4.88), δ 2.21, (t, 2H, J = 7.6 Hz) (2.34); δ 2.02 (2.13), (m, 2H), δ 1.62 (t, 2H, J = 7.4 Hz) (1.53), δ 1.33-1.28, overlapped (10H) (1.33, 1.30, 1.30, 1.30, 1.29). In tilted numbers are the calculated shifts.

¹³C NMR (CDCl₃ 100 MHz, δ; (ppm)): 175.2 (173.6), 139.6 (139.1), 114.5 (115.7), 36.3 (38.7), 34.2 (33.9), 29.4 (29.7, 29.7, 29.6, 28.9, 28.6) and 25.4 (25.3). HRMS m/z calc. For C₁₁H₂₁NONa + [M + Na] + 206.2; found 206.2.

2.5.3. Preparation of 10-11 epoxy undecanamide

To a stirred solution of undec-10-enamide (7 g;) in 108.4 ml of acetone, NaHCO₃ (26.4 g) was added, and then, 5.2 ml of water was added carefully. The resultant thick mixture was strongly stirred, while a solution of 40.6 g of oxone in 158 ml of water was added dropwise during 45 min. The reaction was monitored by thin layer chromatography (TLC) using a mix of ethyl acetate/hexane 2:1. After the reaction was complete, the acetone was removed by evaporation. The remaining solution was acidified with HCl 10% to pH 2 at 10°C and followed rapid extraction with 250 ml of dichloromethane. The aqueous phase was washed three times with additional dichloromethane. The combined organic phase was washed with brine, dried with anhydrous sodium sulfate, and concentrated to a white solid on a rotatory evaporator (IV in Figure 4).

¹H NMR (CDCl₃ 300 MHz, δ; (ppm)): δ 5.44, (bs, 2H), δ 2.90, (m, 1H), δ 2.74, (dd, 1H J = 4.6 Hz), δ 2.46, (dd, 1H J = 5.0 Hz), δ 2.21, (t, 2H, J = 7.6 Hz); δ 1.62 (t, 2H, J = 7.4 Hz), δ 1.51, (m, 2H), δ 1.44 (m, 2H), δ 1.33-1.28, (bs, 8H); ¹³C NMR (CDCl₃ 100 MHz, δ; (ppm)): 173.8 (1C) (CONH₂), 137.6 (1C) (═CH─C), 115.7 (1C) (H₂C═), 38.7 (1C) (─H₂C─CONH₂), 29.7 (3C), 28.7 (2C), 25.3 (1C); HRMS m/z calc. For C₁₁H₂₁NO₂Na + [M + Na]+; found.

2.5.4. Preparation of 10-11 of dihydroxyundecanamide

The previously obtained 10-11 epoxy undecanamide was stirred during 8 hours at 60°C in aqueous HCl 10%. The reaction was monitored by TLC using a mix of ethyl acetate/hexane 2:1. An oil, not miscible with water, was obtained. The mixture reaction was extracted with dichloromethane, and the aqueous phase was washed three times with additionally dichloromethane. The combined organic phase was washed with brine, dried with anhydrous sodium sulfate, and concentrated to a yellow oil on a rotatory evaporator. Yield (75%) (V in Figure 4).

¹H NMR (CDCl₃ 300 MHz, δ; (ppm)): δ 5.44, (bs, 2H), δ 3.78, (m, 1H), δ 3.60, (dd, 1H J = 11.0 Hz), δ 3.48, (dd, 1H J = 2.21, (t, 2H, J = 7.6 Hz); δ 1.62 (t, 2H, J = 7.4 Hz);), δ 1.51, (m, 2H), δ 1.44 (m, 2H), δ 1.33-1.28, (bs, 8H);

¹³C NMR (CDCl₃ 100 MHz, δ; (ppm)): HRMS m/z calc. For C₁₁H₂₃NO₃H₃O+ [M + H₃O] + 236.2; found 236.2.

2.5.5. Preparation of TOBC

In the course of 20 minutes, 20 g TOBA were added to 350 ml thionyl chloride from Sigma-Aldrich Chemie GmBH (Steinheim, Germany), while stirring rapidly at room temperature (VI in Figure 4).

The solution was boiled with the reflux condenser. When the emission of HCl had finished and most of the sediment had dissolved, the hot solution was filtered and cooled down to 0°C for a day. The obtained product that separated out was filtered, vacuum dried, and recrystallized in chloroform, from SDS Votre Partenaire Chimie (Peypin, France).

Yield: 14 g (60%).

2.5.6. Preparation of PTOBUME-amide.

A mixture of TOBC (5.5 g; 0.012 mol), 10-11 of dihydroxyundecanamide (2.7 g; 0.012 mol) in 3 ml of diphenyl oxide from Sigma-Aldrich Chemie GmBH (Steinheim, Germany) was purged with dry nitrogen from Praxair (Madrid, Spain) for 25min at room temperature. Then, while maintaining the gas stream, the flask was transferred to a bath containing a high-temperature heat-transfer agent. The polycondensation was carried out for 360 minutes at 200°C. The reaction gets completed when emission of HCl had finished. The reaction mixture was poured into 300 ml of toluene from Merck KGaA (Darmstadt, Germany), decanting PTOBUME-amide. After 12 hours, it was filtered, washed with ethanol, and vacuum dried. After 3 weeks, a second fraction of polymer was precipitated of the toluene mother liquors, which was filtered, washed with ethanol, and vacuum dried.

Yield first fraction 3.0 g (38.5%); yield first and second fraction 0.1 g (40.0%).

2.6. Synthesis of Cholesteric PTOBEE-amide (C₂₆H₁₉O₉N)_n

Poly[oxy(1,2-butan-4-amidyl)-oxycarbonyl-1,4-phenylene-oxy-1,4-terephthaloyl-oxy-1,4-phenylene-carbonyl], VI in Figure 5, was obtained through poly-condensation reaction between 4 and 4′-(terephthaloyldioxydibenzoic chloride) TOBC and the racemic mixture of DL-3,4-dihydroxybutanamide (IV in Figure 5). The same notation has been used with precursor cholesteric liquid crystal PTOBEE, Figure 1.

Figure 5.

Synthetic method of cholesteric liquid-crystalline PTOBEE-amide.

2.6.1. Preparation of 3,4-dihydroxybutanoic acid

To a stirred mixture of 10 g of but-3-enoic acid solved in 130 ml acetone, 34 g. NaHCO₃ in 65 ml mili-Q water was added carefully. The resultant mixture was strongly stirred, while a solution of 51.1 g oxone in 200 ml of water was added dropwise during 120 min. The reaction was monitored by thin layer chromatography (TLC) using a mix of ethyl acetate/diethyl ether 1:1. After the reaction was complete, the acetone was removed by evaporation. The remaining solution was acidified with HCl 10% to pH 2 at 10°C and followed of rapid extraction with 250 ml of ethyl acetate. The aqueous phase was washed three times with additional ethyl acetate. The combined organic phase was washed with brine, dried with anhydrous sodium sulfate, and concentrated to a white solid on a rotatory evaporator (III in Figure 5).

2.6.2. Preparation of 4-hydroxydihydrofuran-2(3H)-one

To 3,4-dihydroxybutanoic acid into a flask equipped with a Dean Stark adapter filled with a toluene column finally connected to a refrigerant, 0.5 ml trifluoroacetic acid was added in 100 ml toluene heating to 110°C, mixing for 3 hours. The reaction product was removed with ethanol, washed in ethyl acetate, and dried. The reaction was monitored by thin layer chromatography (TLC) using a mix of ethyl acetate/diethyl ether 1:1 (IV in Figure 5).

2.6.3. Preparation of 3,4-dihydroxybutanamide

To the 4-hydroxydihydrofuran-2(3H)-one, 100 ml NH₃ 33% was added stirring at 70°C with reflux for 12 h. The reaction product was removed with ethanol, filtered, and washed with water several times (V in Figure 5).

2.6.4. Preparation of PTOBEE-amide

In a three-neck round-bottom flask, 0.2 g 3,4-dihydroxybutanamide was added dropwise to 1 g TOBC solved in 100 ml 1,1,2,2-Tetrachloroethane. The reaction was stirred at 90°C for 20 hours. The reaction product was filtered, washed in 50 ml ethanol, 100 ml water, 200 ml NaHCO₃ (10%), 200 ml HCl (5%), 300 ml water, and 200 ml ethanol, and dried.

3. Characterization techniques

3.1. Conventional NMR techniques

The obtained polymers are characterized by ¹H-NMR, ¹³C-NMR, COSY (Homonuclear Correlation Spectroscopy), TOCSY (Total Correlation Spectroscopy), NOESY (Nuclear Overhauser Effect Spectroscopy, through-space correlation method), HSQC (Heteronuclear Single-Quantum Correlation spectroscopy), and HMBC (Heteronuclear Multiple Bond Correlation) for correlations between carbons and protons that are separated by two, three, and sometimes four bonds, in conjugated systems. Direct one-bond correlations being suppressed.

The experiments were performed in a Bruker 300 MHz NMR spectrometer and VARIAN 400 and 500 MHz spectrometers. The solvents used were DMSO-d₆ and CDCl₃, from Merck KGaA (Darmstadt, Germany), at 25°C. ¹H chemical shifts were referenced to the residual solvent signal at δ = 2.50 ppm (DMSO-d₆) relative to tetramethylsilane (TMS). All the spectra were processed and analyzed with MestReNova v.11.0.4 software [9]. Predicted ¹H and ¹³C-NMR chemical shifts were calculated from the formula with ChemDraw Professional, v.15.1.0.144 [10].

3.2. Thermal behavior

Thermal stability was studied by Thermogravimetry on a Mettler TA4000-TG50 at heating rate of 10°C/min with nitrogen purge between 30 and 600°C. Thermal behavior was determined by differential scanning calorimetry (DSC) in a Mettler TA4000/DSC30/TC11 calorimeter, with series of heating/cooling cycles in a temperature range between 0 and 230°C.

3.3. The optical activity

The optical activity of the polymers was measured as optical rotatory dispersion (ORD) at 25°C in DMSO from Scharlau Chemie, in a Perkin Elmer 241 MC polarimeter with wavelengths: λ_Na = 589 nm, slit = 5 mm, integration time = 50 s; λ_Hg = 574 nm, slit = 14 mm, integration time = 50s; λ_Hg = 546 nm, slit = 30 mm, integration time = 50 s; λ_Hg = 435 nm, slit = 5 mm, integration time = 50s; λ_Hg = 365 nm, slit = 2.5 mm, integration time = 50 s.

4. Structural characterization by NMR

4.1. Structural characterization of PTOBDME-choline

The designation of the ¹H and ¹³C-NMR chemical shifts, in DMSO-d₆, of the monomeric unit and the end groups of Polyester PTOBDME-choline, is given in Table 1. All the spectra have been analyzed and interpreted the help of MestReNova [9]. The predicted theoretical values, also in Table 1, have been calculated by ChemDraw [10]. Similar notations as those assigned with precursor cholesteric liquid crystal polyesters PTOBDME [1, 2] have been used.

Set of signal of system ( ‘ ) and ( “ )				Set of signal of system without apostrophe ( )				Calculated chemical shift
Atom	^1H(ppm)	Atom	^13C(ppm)	Atom	^1H(ppm)	Atom	^13C(ppm)	^1H(ppm)	^13C(ppm)
^20’H	8.36	^20’C	130.4	^20H	8.36	^20C	130.4	8.04	130.2
		^19’C	133.3			^19C	133.3		135.4
		^18’C	163.5			^18C	163.4		165.2
		^17’C	153.9			^17C	154.4		155.6
^16’H	7.50	^16’C	122.1	^16H	7.55	^16C	122.2	7.26	121.5
^15’H	8.08	^15’C	130.8	^15H	8.15	^15C	131.0	8.13, 8.11	130.3
		^14’C	128.8			^14C	127.6		126.9
		^13’C	166.7			^13C	164.7		165.9
Hc’	5.45	^12’C	72.3	Hc	5.26	^12C	73.7	4.55	70.3
Hc”	4.39	^12”C	60.0					3.81	70.8
Ha’Hb’	4.63, 4.52	^11’C	65.7	Ha, Hb	3.95, 3.89	^11C	46.4	4.80, 4.55	67.5
Ha”Hb”	4.52	^11”C	67.8					4.53, 4.28	70.8
Hd’	1.83	^10’C	30.0	Hd	1.77	^10C	31.2	1.71	30.7
Hd”,He”	1.92 1.78	^10”C						1.44	34.0
Hf’, Hg’	1.45	^9’C	24.6	Hf, Hg	1.35	^9C	24.4	1.29	23.3
Hf”, Hg”	1.53, 1.44	^9”C	25.6					1.29	23.1
^8’H	1.24	^8’C	28.9 m*	^8H	1.24	^8C	28.9 m*	1.29	29.6
^7’H	1.24	^7’C	28.9 m*	^7H	1.24	^7C	28.9 m*	1.29	29.6
^6’H	1.24	^6’C	28.9 m*	^6H	1.24	^6C	28.9 m*	1.26	29.6
^5’H	1.24	^5’C	28.9 m*	^5H	1.24	^5C	28.9 m*	1.26	29.6
^4’H	1.24	^4’C	28.6 m*	^4H	1.24	^4C	28.6 m*	1.26	29.3
^3’H	1.24	^3’C	31.3	^3H	1.24	^3C	31.3	1.26	31.8
^2’H	1.24	^2’C	22.1	^2H	1.24	^2C	22.1	1.26	22.7
^1’H	0.85	^1’C	14.0	^1H	0.85	^1C	14.0	0.86	14.1

Table 1.

¹H and ¹³C-NMR chemical shifts (ppm) observed and calculated for the repeating unit and the aliphatic end group polyester PTOBDME-choline.

^*

Signal of ⁴C to ⁸C at 28.8 ppm is a multiplet from 28.9 to 28.6.

The symbol ( ‘ ) and without it ( ) distinguish the two independent system of the repeating unit, the symbol ( “ ) is used to mark signals due to the aliphatic end group.

Considering the monomer structure, three zones can be differentiated in the ¹H-NMR spectrum, corresponding to the mesogen, including aromatic protons between 11.0–7.00 ppm, the spacer where methylene and methine protons directly attached to oxygen atoms are observed, with signals between 6 and 3 ppm, and the flexible side chain formed by aliphatic protons between 2 and 0.8 ppm. The main feature of the proton spectrum is the presence of higher number of peaks than those expected for the monomeric unit. Hydrogen atoms H_a and H_b are bonded to 11C atom, allocated in α position with respect to the asymmetric carbon atom ¹²C*. For that reason, they are diastereotopic and their 1H-NMR signals, usually indistinguishable, split in two easily differentiated. The same effect is observed for H_d and H_e, bonded to ¹⁰C, and for H_f and H_g, both bonded to ⁹C.

The presence of two independent ¹H-NMR sets of signals are observed in the spectrum, one marked with ( ‘ ) and the other without it ( ). They are attributed to two conformers gg and gt of the spacer within the repeating unit respectively. The same effect has been reported for PTOBDME and PTOBEE, and accordingly, similar nomenclature is used to identify the signals. A third set of signals, marked with ( “ ), is assigned to the aliphatic end group.

In the aromatic zone singlet at 8.36 ppm belongs to ²⁰H and doublets at 7.50 and 8.08 ppm are assigned to ^16’H and ^15’H, respectively, and doublets at 7.55 and 8.15 ppm to ¹⁶H and ¹⁵H; similar assignation was previously carried out in precursor PTOBDME [1]. In the spacer zone, multiplet at 5.45 ppm is interpreted due to H_c’, and the double doublets at 4.63 and 4.52 ppm correspond to H_a’ and H_b’. These peaks presented correlation signals in COSY and were related with other aliphatic signals H_d’H_e’ (1.83 ppm) and H_f’H_g’ (1.45 ppm) by TOCSY experiment. Multiplet at 5.26 ppm was assigned to H_c and double doublets at 3.95 and 3.89 ppm to H_a and H_b, and they showed COSY correlations and were related with signals at 1.77 ppm (H_d) and 1.35 ppm (H_f H_g) respectively by TOCSY experiment. The peaks assigned to H_a”, H_b”, and H_c” due to aliphatic end group are overlapped with H_b’ (4.52 ppm) and with ²¹H (4.54 ppm) (in Table 2), and they were assigned through TOCSY correlations observed for signal at 4.52 ppm (not observed for 4.63 ppm, H_a’), with the multiplet at 4.39 ppm (H_c”) and confirmed by HSQC. By this method, carbon C¹¹’ (65.7 ppm) was correlated with signals at 4.63 (H_a’) and 4.52 (H_b’) and carbon ¹¹”C (67.8 ppm) with 4.52 ppm (H_a”) and (H_b”). Signals at 1.83 and 1.45 ppm assigned to (H_d’) and (H_f’, H_g’) and correlated by COSY with H_c’(5.45 ppm). Peaks at 1.77 ppm (H_d H_e) and 1.35 ppm (H_f H_g) are correlated by COSY with H_c (5.26 ppm). Signals at 1.92, 1.78 ppm are related with H_c” (4.39 ppm) by COSY experiments, and they were assigned to H_d” and H_e”. They are also related with Hf” (1.53 ppm) and Hg”(1.44 ppm) by the same experiment.

PTOBDME-choline							PTOBEE-choline
Observed chemical shifts			Calc. chemical shifts				Observed chemical shifts		Calc. chemical shifts
Atom ^1H(ppm)			Atom ^13C(ppm)				Atom ^1H(ppm)		Atom ^13C(ppm)
	DMSO		DMSO	^1H calc	^13C calc		DMSO		DMSO	^1H calc	^13C calc
^21H, ^21’H	4.54, 4.76	^21C ^21’C	62.6 58.8	4.69	58.1	^13H ^13’H	4.56 4.75	^13C ^13’C	58.8 58.5	4.69	58.1
^22H, ^22’H	3.33 3.85	^22C ^22’C	55.6 64.0	3.70	66.5	^14H ^14’H	3.32 3.84	^14C ^14’C	55.0	3.70	66.5
^23H, ^23’H	2.74 3.21	^23C ^23’C	43.2 53.0	3.30	54.4	^15H ^15’H	2.74 3.22	^15C ^15’C	42.5 52.7	3.30	54.4

Table 2.

Observed ¹H and ¹³C-NMR chemical shifts (ppm), in DMSO-d6, for the -N, N, N-trimethylethan- 1-ammonium (Choline) oxy benzoate hydrochloride end group, in Polyester PTOBDME-choline and Polyester PTOBEE-choline, and theoretical calculated values.

Choline end group showed in Table 2, two set of signals probably due to conformational equilibrium: Multiplets assigned to ²¹H (4.54 ppm), ²²H (3.33 ppm), and ²³H (2.74 ppm), correlated in COSY, and another set was multiplets ²¹’H (4.76 ppm), ²²’H (3.85 ppm) and a singlet ²³’H (3.21 ppm).

The HSQC experiment allowed the direct allocation of carbon atoms linked to hydrogens, confirming the assignation of the proton signals overlapped in the ¹HNMR experiment. The correlation of carbon atom ¹¹’C (65.7 ppm) with H_a’ (4.63 ppm) and H_b’ (4.52 ppm); correlation between ¹¹”C (67.8 ppm) and H_a” and H_b”; and correlation between carbon ²¹C at (62.6 ppm) and ²¹H at (4.54 ppm), are observed in Table 2.

4.2. Structural characterization of PTOBEE-choline

The assignment of the ¹H and ¹³C-NMR chemical shifts, in CDCl₃ and DMSO-d₆, of the monomeric unit and the end groups of PTOBEE-choline are given in Table 3, with the predicted values calculated by ChemDraw Professional [10]. Similar notations as those designated for precursor cholesteric liquid crystal PTOBEE [4] have been used.

Set of signal of system (’) and (“)						Set of signal of system without apostrophe ( )						Calculated chemical shifts
Atom	^1H(ppm)		Atom	^13C(ppm)		Atom	^1H(ppm)		Atom	^13C(ppm)		Atom	Atom
	CDCl3	DMSO		CDCl3	DMSO		CDCl3	DMSO	DMSO	CDCl3	DMSO	^1H	^13C
^12’H	8.34	8.35	^12’C	130.4	130.1	^12H	8.34	8.35	^12C	130.4	130.0	8.04	130.2
			^11’C	133.8	133.2				^11C	132.2	131.6		135.4
			^10’C	163.6	163.3				^10C	163.6	163.3		165.2
			^9’C	154.5	154.2				^9C	154.5	153.8		155.6
^8’H	7.36	7.48, 7.50	^8’C	121.7	121.7	^8H	7.34	7.53, 7.51	^8C	121.7	122.0	7.26	121.5
^7’H	8.18	8.08, 8.06	^7’C	131.5	130.6	^7H	8.16	8.11, 8.09	^7C	131.5	130.6	8.13, 8.11	130.3
			^6’C	128.3	128.7				^6C	127.9	127.3		126.9
			^5’C	165.4	166.4				^5C	165.4	164.7		165.9
Hc’	5.46	5.38	^4’C	73.8	73.1	Hc	5.25	5.20	^4C	75.1	74.4	4.55	72.5
Hc”	4.15	4.36	^4”C		61.6^*							3.81	73.0
Ha’, Hb’	4.60, 4.53	4.63, 4.52	^3’C	65.6	64.8	Ha, Hb	3.76, 3.74	3.94, 3.91	^3C	45.23	45.6	4.80, 4.55	67.2
Ha”, Hb”	4.53	4.52 ^*	^3”C	67.8	67.1							4.53, 4.28	70.5
Hd’	1.90	1.86	^2’C	24.4	23.0	Hd	1.86	1.81	^2C	24.4	24.2	1.75	23.5
Hd”, He”	1.88, 1.13	1.96, 1.80	^2”C		^*							1.48	26.8
^1’H	1.09	1.02	^1’C	9.8	9.2	^1H	1.03	0.95	^1C	9.6	9.8	0.96	7.8
^1”H	1.13	1.04^*	^1”C		8.73^*							0.96	7.6

Table 3.

Observed and calculated ¹H and ¹³C-NMR chemical shifts (ppm) for polyester PTOBEE-choline in DMSO-d₆ and CDCl₃. Repeating unit and the aliphatic end group.

*Overlapped signal

In the ¹H-NMR experiment in CDCl3, observed chemical shifts are ¹²H singlet at (8.34 ppm), ⁸H doublet at (7.34 ppm), ⁷H doublet at (8.16 ppm), ⁸’H doublet at (7.36 ppm) and ⁷’H doublet at (8.18 ppm). Multiplets at 5.46 and at 5.25ppm are interpreted as H_c’ and H_c, respectively. The double doublet at 4.60 ppm is assigned to H_a’ and correlates in COSY with H_c‘ signal. An overlapped signal at 4.53 ppm is identified as H_b’, with COSY and TOCSY cross signal with H_a’. At 4.15, a weak multiplet is assigned to H_c”, it presented COSY correlation with signal 4.53 ppm, indicating the presence of H_a” and H_b”. The two double doublets at 3.76 and 3.74 ppm were identified as H_a and H_b and presented the expected COSY correlation with H_c (5.25 ppm). The overlapped signal at 1.90 ppm is identified as H_d‘, with cross signal with H_c’. A very weak COSY cross signal between 4.15 and H_d”(1.88 ppm) is observed. Triplet at 1.09 ppm is due to ¹’H, with TOCSY correlation with H_c’, while triplet at (1.03 ppm) is ¹H, with TOCSY correlation with H_c. The weak triplet at 1.13 ppm corresponded with ¹”H. As in PTOBDME-choline, the choline end group shows, in Table 2, two set of signals due to conformational equilibrium. Multiplets ¹³H, ¹⁴H are observed at 4.56 and 3.32 ppm, respectively, and ¹⁵H at 2.74 ppm, correlated in COSY experiments, and another set was ¹³’H and ¹⁴’H multiplets at 4.75 and 3.84 ppm, respectively, singlet ¹⁵’H at 3.22 ppm.

HSQC experiment was performed to determine the chemical shift of carbons bonded to the assigned hydrogen. The complex signal at 4.53 in proton presented several correlations with carbons. H_a’(4.60 ppm) and H_b’ (4.53 ppm) showed correlation with carbon ³’C (65.6 ppm). Another correlation with overlapped signal at 4.53 ppm H_a”H_b” was observed with carbon ³”C (67.8 ppm). Signals corresponding to ¹C, ²C, and ⁴C of aliphatic end group were not observed due to the low concentration. ¹³C-NMR experiment allowed the assignation of the carbons not attached to hydrogens matching the calculated model.

4.3. Structural characterization of PTOBDME-ammonium

The structure of PTOBDME-ammonium, as depicted II in Figure 3, is confirmed by ¹H and ¹³C-NMR, with the chemical shifts given in Table 4. In the aromatic zone of the ¹H-NMR spectrum of PTOBDME-ammonium, in DMSO-d₆, a singlet at 8.36 ppm belongs to ²⁰H and doublets at 7.50 and 8.07 ppm are assigned to ^16’H and ^15’H, respectively, and doublets at 7.57 and 8.13 ppm to ¹⁶H and ¹⁵H. In the spacer zone where methylene and methines attached to oxygen are observed, a multiplet at 5.45 ppm, and the double doublets at 4.64 ppm, 4.48 ppm correspond to H_c’, H_a’, and H_b’, respectively; these signals present correlation signals of COSY and are related with other aliphatic signals: H_d’, H_e’ (1.81 ppm) and H_f’, H_g’ (1.42 ppm) by TOCSY. Multiplet at 5.26 ppm is assigned to H_c and double doublets at 3.94 and 3.88 ppm, to H_a and H_b, they show COSY correlations and are related with signals H_d H_e (1.77 ppm) and H_f H_g (1.33 ppm), in the TOCSY experiment. In the set of signals due to aliphatic end group, H_a”, H_b”, and H_c” are overlapped with H_b’ (4.48 ppm) and with ²¹H (4.38 ppm), according to TOCSY correlations observed for signal H_b’ (4.48 ppm) and not observed for H_a’ (4.63 ppm) and confirmed by HSQC. Aliphatic signals at 1.81 ppm (H_d’, H_e’) and 1.42 ppm (H_f’, H_g’) have TOCSY correlation with 5.45 ppm (H_c’), also COSY correlation. Signals at 1.77 ppm (H_d H_e) and 1.33 ppm (H_f H_g) show TOCSY correlation with H_c (5.26 ppm). The signal at 1.33 ppm cannot be observed in the ¹H sprectrum due to the overlapping with CH₂, but it was clearly observed in TOCSY 2D. Signals at 1.91 and 1.78 ppm, related with H_c”(4.38 ppm) by COSY and TOCSY, are assigned to H_d” and H_e” and are also related with H_f”(1.51 ppm) and H_g”(1.42 ppm) by the same experiment.

Set of signal of system (‘) and (“)				Set of signal of system without apostrophe ( )				Calculated chemical shift
Atom	^1H (ppm	Atom	^13C(ppm)	Atom	^1H(ppm)	Atom	^13C(ppm)	^1H(ppm)	^13C(ppm)
^20’H	8.36	^20’C	130.4	^20H	8.36	^20C	130.4	8.04	130.2
		^19’C	133.3			^19C	133.3		135.5
		^18’C	163.5			^18C	163.4		165.2
		^17’C	153.8			^17C	154.3		155.7
^16’H	7.50	^16’C	122.1	^16H	7.57	^16C	122.4	7.26	121.5
^15’H	8.07	^15’C	131.0	^15H	8.13	^15C	131.0	8.13;8.11	130.3
		^14’C	128.9			^14C	127.6		127.0
		^13’C	166.6			^13C	164.8		166.0
Hc’	5.45	^12’C	72.6	Hc	5.26	^12C	73.4	4.55	70.4
Hc”	4.38	^12”C	60.4					3.81	70.9
Ha’, Hb’	4.64, 4.48	^11’C	66.0	Ha, Hb	3.94, 3.88	^11C	46.4	4.80, 4.55	67.6
Ha”, Hb”	4.48	^11”C	67.8					4.53, 4.28	70.9
Hd’, He’	1.81	^10’C	30.3	Hd, He	1.77	^10C	31.3	1.71	30.8
Hd”, He”	1.91 1.78	^10’C	33.6					1.44	34.1
Hf’, Hg’	1.42	^9’C	24.6	Hf, Hg	1.33	^9C	24.3	1.29	23.4
Hf”, Hg”	1.51;1.42	^9”C	25.3					1.29	23.2
^8’H	1.22	^8’C	28.6 m*	^8H	1.22	^8C	28.6 m*	1.29	29.7
^7’H	1.22	^7’C	28.6 m*	^7H	1.22	^7C	28.6 m*	1.29	29.7
^6’H	1.22	^6’C	28.6 m*	^6H	1.22	^6C	28.6 m*	1.26	29.7
^5’H	1.22	^5’C	28.5 m*	^5H	1.22	^5C	28.5 m*	1.26	29.7
^4’H	1.22	^4’C	28.1 m*	^4H	1.22	^4C	28.1 m*	1.26	29.4
^3’H	1.22	^3’C	31.3	^3H	1.22	^3C	31.3	1.26	31.9
^2’H	1.22	^2’C	22.1	^2H	1.22	^2C	22.1	1.26	22.8
^1’H	0.84	^1’C	14.0	^1H	0.84	^1C	14.0	0.86	14.1

Table 4.

¹H and ¹³C-NMR chemical shifts (ppm) observed and calculated for the repeating unit of polyester PTOBDME-ammonium chloride and the aliphatic end group.

^*

Signal of ¹³C at 28.8 ppm is a multiplet from 28.9 to 28.7.

The symbol (‘) and with no apostrophe ( ) distinguish the two independent systems of the repeating unit, the symbol (“) is used to mark the signals of the aliphatic end group.

Ammonium end group shows ²³H, ²²H, ²⁴H multiplets at 3.28 ppm, 2.17 ppm, and 2.79 ppm, respectively . ²³ H at 3.28 ppm was overlapped with signal of H₂O of the deuterated solvent, and it was assigned due to the COSY and TOCSY correlations with signals at 4.38 ppm (²¹H) and at 2.17 ppm (²²H) and HSQC correlation with ²³C at 54.0 ppm. ²¹H signal was overlapped at 4.38 ppm, and it was identified by COSY correlations with 2.17 ppm (²²H) and TOCSY correlations with 3.28 ppm (²¹H). The polymer holds positive charge due to ammonium proton ²⁵H observed at 10.33 ppm.

The HSQC experiment confirmed the direct assignation of carbon atom ¹¹’C (66.0 ppm) linked to protons H_a’ (4.63 ppm) and H_b’ (4.48 ppm). Signal H_b’ exhibits correlation with ¹¹”C (67.8 ppm), linked to H_a” and H_b”. Two cross signal are observed for H_c” (4.38 ppm), one with carbon atom at ¹²”C (60.4 ppm) and another with ²¹H linked to carbon atom ²¹C (62.3 ppm). The correlations of carbon atom ¹⁰”C (33.6 ppm) with H_d”(1.91 ppm) and H_e”(1.78 ppm), and carbon atom ⁹”C (25.6 ppm) with H_f”(1.51 ppm) and H_g”(1.42 ppm) confirmed the previous assignation.

4.4. Structural characterization of PTOBEE-ammonium

Table 6 shows the assignation of ¹H and ¹³C-NMR chemical shifts (ppm) observed of polyester PTOBEE-ammonium chloride and calculated for the repeating unit and the aliphatic end group. In the ¹H-NMR experiment, in CDC_l3, peaks observed at 8.34, 7.36, 7.34, 8.18, and 8.16 ppm are assigned to ¹²H singlet, ^8’H doublet, ⁸H doublet and ^7’H doublet, and ⁷H doublet, respectively. Peak at 5.46 ppm is H_c‘ and 5.25 ppm is H_c. The double doublet at 4.60 ppm is interpreted as H_a’ because of its shape and the COSY correlation with H_c‘. An overlapped signal at 4.53 ppm is attributed to H_b’, by COSY and TOCSY cross signal with H_a‘. A weak multiplet at 4.15 ppm is assigned to H_c”, and this signal presented COSY correlation with H_b’, indicating the presence of H_a” and H_b”. The two double doublets at 3.76 and 3.74 ppm are identified as H_a and H_b and presented the expected COSY correlation with H_c. The overlapped signal at 1.90 ppm is H_d‘ correlated with H_c’. A very weak COSY cross signal is observed between H_c” and 1.88 ppm (H_d”). Triplet signal at 1.09 ppm with TOCSY correlation with H_c’ is interpreted as H_e’, while triplet at 1.03 ppm with TOCSY correlation with H_c was assigned to H_e. The weak triplet at 1.13 ppm corresponds to H_e”.

Signals of proton ammonium end group in DMSO-d₆ are observed at (Table 5): ¹⁶H singlet (2.81 ppm), ¹⁵H multiplet (3.25 ppm), ¹⁴H multiplet (2.18 ppm), and ¹³H multiplet overlapped at (4.39 ppm) but presented COSY correlations with 2.18 ppm and TOCSY correlations with 3.25 ppm. The compound is positively charged, with the ammonium proton ¹⁷H observed at 10.3 ppm in DMSO-d₆ and 13.2 ppm in CDCl₃.

PTOBDME-Ammonium						PTOBEE-Ammonium
Observed chemical shifts				Calc. chemical shifts		Observed chemical shifts						Calc. chemical shifts
Atom	^1H(ppm)	Atom	^13C(ppm)	^1H(ppm)	^13C(ppm)	Atom	^13C(ppm)	^1H(ppm)	Atom	^1H(ppm)	^13C(ppm)	^1H(ppm)	^13C(ppm)
	DMSO		DMSO				DMSO	CDCl3		DMSO	CDCl3
^21H	4.38	^21C	62.3	4.25	63.0	^13H	4.39	4.49	^13C	61.3	61.9	4.25	63.0
^22H	2.17	^22C	23.8	2.19	22.5	^14H	2.18	2.42	^14C	23.1	24.5	2.19	22.5
^23H	3.28	^23C	54.0	3.24	55.2	^15H	3.25	3.14	^15C	53.8	55.5	3.24	55.2
^24H	2.79	^24C	42.5	2.90	45.0	^16H	2.81	2.82	^16C	41.9	43.1	2.90	45.0
^25H	10.33			7.0		^17H	10.33	13.2				7.0

Table 5.

¹H and ¹³C-NMR chemical shifts (ppm) observed for the 3-dimethylamine-1-propyl benzoate hydrochloride end group in polyester PTOBDME-ammonium and polyester PTOBEE-ammonium, and calculated values.

Set of signal of system (’) and (“)						Set of signal of system without apostrophe ( )						Calculated chemical shift
Atom	^1H(ppm)		Atom	^13C(ppm)		Atom	^1H(ppm)		Atom	^13C(ppm)		Atom	Atom
	CDCl3	DMSO		CDCl3	DMSO		CDCl3		DMSO	CDCl3	DMSO	^1H	^13C
^12’H	8.34	8.36	^12’C	130.4	129.9	^12H	8.34	8.36	^12C	130.4	129.9	8.04	130.2
			^11’C	133.8					^11C	132.2			135.4
			^10’C	163.6					^10C	163.6			165.2
			^9’C	154.5					^9 C	154.5			155.6
^8’H	7.36	7.53	^8’C	121.7	121.7	^8H	7.34	7.53	^8C	121.9	121.7	7.26	121.5
^7’H	8.18	8.13	^7’C	131.5	130.4	^7H	8.16	8.11	^7C	131.5	130.4	8.13, 8.11	130.3
			^6’C	128.3					^6C	127.9			126.9
			^5’C	165.4					^5C	165.4			165.9
Hc’	5.46	5.39	^4’C	73.8	73.02	Hc	5.25	5.20	^4C	75.1	74.4	4.55	72.5
Hc”	4.15	4.40	^4”C		*							3.81	73.0
Ha’, Hb’	4.60, 4.53	4.64 4.53	^3’C	65.6	64.6	Ha, Hb	3.76, 3.74	3.96, 3.92	^3C	45.23	45.6	4.80, 4.55	67.2
Ha”, Hb”	4.53	4.48*	^3”C	67.8	*							4.53, 4.28	70.5
Hd’	1.90	1.86	^2’C	24.4	23.06	Hd	1.90	1.82	^2C	24.4	24.1	1.75	23.5
Hd”, He”	1.88, 1.13	1.90, 1.86*	^2”C		*							1.48	26.8
^1’H	1.09	1.04	^1’C	9.8	8.73	He	1.03	0.96	^1C	9.6	8.38	0.96	7.8
^1”H	1.13	1.06*	^1”C		8.73*							0.96	7.6

Table 6.

¹H and ¹³C-NMR chemical shifts (ppm) observed for polyester PTOBEE-ammonium chloride, both the repeating unit and the aliphatic end group, in DMSO-d6 and in CDCl₃, and calculated values.

*

Overlapped signal.

HSQC experiment exhibits several correlations of the complex proton signal at 4.53 ppm, carbon atoms. Double doublet H_a’(4.60 ppm) H_b’ (4.53) correlates with ³’C at (65.6 ppm). Another correlation is observed between proton H_a” and ³”C (67.8 ppm). Correlation between the overlapped signal of proton ¹³H (4.39 ppm), within the ammonium end group, and ¹³C (61.9 ppm) in CDCl₃ is observed.

4.5. Structural characterization of PTOBUME-amide

The structures of undec-10-enamide, 10-11-epoxy-undecanamide, and 10,11-dihydroxyundecanamide (III, IV and V in Figure 4) were confirmed by ¹H-NMR, ¹³C-NMR, registered in DMSO-d₆ at 25°C in a Bruker 300 MHz NMR spectrometer. Chemical shifts and Mass spectrometry results are given in Section 2.3.

The structure of PTOBUME-amide, VII in Figure 4, has also been confirmed by ¹H-NMR, ¹³C-NMR, COSY and HSQC, obtained in VARIAN 400 and 500 MHz spectrometers, also at room temperature. The solvent used were DMSO-d₆ and CDCl₃ from Merck KGaA (Darmstadt, Germany). The spectra were processed and analyzed with the help of MestReNova 11.0.4 [9]. The chemical shifts are given in Table 7. Theoretical values predicted by ChemDraw Professional, v. 15.1.0.144. Tilted values are chemical shifts registered in CDCl₃.

System (‘)				System without apostrophe ( )				Theoretical chemical shifts
Atom	^1H	Atom	^13C	Atom	^1H	Atom	^13C	Atom	^1H	Atom	^13C
^19’H	8.36	^19’C		^19H	8.36	^19C	130.4	^19H	8.04	^19C	130.2
		^18’C				^18C	133.7			^18C	135.5
		^17’C				^17C	163.7			^17C	165.2
		^16’C				^16C	154.7			^16C	153.7
^15’H	8.07	^15’C		^15H	8.07	^15C	121.6	^15H	7.52	^15C	121.5
^14’H	7.50	^14’C		^14H	7.50	^14C	131.3	^14H	8.18	^14C	130.3
		^13’C				^13C	128.2			^13C	126.9
		^12’C				^12C	165.7			^12C	165.9
Hc’	5.76	^11’C		Hc	5.45	^11C		Hc	5.16	^11C	70.3
Ha’, Hb’	4.95, 4.30	^10’C		Ha, Hb	4.23, 4.18	^10C		Ha,Hb	4.78, 4.53	^10C	66.0
Hd’, He’	1.74	^9’C		Hd, He	1.55	^9C		Hd, He	1.67	^9C	30.7
Hf’, Hg’	1.37	^8’C		Hf, Hg	1.22	^8C		Hf, Hg	1.25	^8C	23.3
^7’H	1.22	^7’C		^7H	1.22	^7C		^7H	1.25	^7C	29.6
^6’H	1.22	^6’C		^6H	1.22	^6C		^6H	1.26	^6C	29.6
^5’H	1.22	^5’C		^5H	1.22	^5C		^5H	1.30	^5C	28.9
^4’H	1.22	^4’C		^4H	1.22	^4C		^4H	1.30	^4C	28.6
^3’H	1.52	^3’C		^3H	1.52	^3C		^3H	1.53	^3C	25.3
^2’H	2.28	^2’C		^2H	2.28	^2C		^2H	2.34	^2C	38.7
NH2’	7.0	^1’C		NH2	7.0	^1C		NH2	7.03	^1C	173.6
Experimental signals “end group”								Theoretical chemical shifts end group
Hc”	4.18			^11”C				Hc”	4.57	^11”C	73.6
Ha”, Hb”				^10”C				Ha”, Hb”	3.86, 3.80	^10”C	64.3
Hd”, He”	1.62			^9”C				Hd”, He”	1.67	^9”C	30.5
Hf”, Hg”	1.22			^8”C				Hf”, Hg”	1.25	^8”C	25.6
^7”H	1.22			^7”C				^7”H	1.25	^7”C	29.6
^6”H	1.22			^6”C				^6”H	1.26	^6”C	29.6
^5”H	1.22			^5”C				^5”H	1.30	^5”C	28.9
^4”H	1.22			^4”C				^4”H	1.30	^4”C	28.6
^3”H	1.52			^3”C				^3”H	1.53	^3”C	25.3
^2”H	2.28			^2”C				^2”H	2.34	^2”C	38.7
NH2 “	10.7			^1”C				NH2 “	7.03	^1”C	173.6
CH2	4.36			CH2		61.2		CH2	4.29	CH2	60.9
CH3	1.35			CH3		14.3		CH3	1.30	CH3	14.1

Table 7.

¹H and ¹³C-NMR chemical shifts (ppm) observed and calculated for chiral Polyesteramide PTOBUME-amide.

4.6. Structural characterization of PTOBEE-amide

The structure of PTOBEE-amide, VI in Figure 5, has also been confirmed by ¹H-NMR, ¹³C-NMR, COSY and HSQC, obtained in VARIAN 400 and 500 MHz spectrometers, at room temperature. The solvent used was DMSO-d₆ from Merck KGaA (Darmstadt, Germany). The experimental chemical shifts analyzed from the spectra are given in Table 8. Theoretical values predicted by ChemDraw Professional, v. 15.1.0.144.

System (‘)				System without apostrophe ( )				Theoretical chemical shifts
Atom	^1H	Atom	^13C	Atom	^1H	Atom	^13C	Atom	^1H	Atom	^13C
^12’H	8.32	^12’C	131.36	^12H	8.32	^12C	131.36	^12H	8.04	^12C	130.2
		^11’C				^11C				^11C	135.4
		^10’C				^10C				^10C	165.2
		^9’C				^9C				^9C	155.7
^8’H	8.08	^8’C	132.35	^8H	8.08	^8C	132.35	^8H	8.18	^8C	121.5
^7’H	7.45	^7’C	123.06	^7H	7.45	^7C	123.06	^7H	7.52	^7C	130.3
		^6’C				^6C				^6C	126.9
		^5’C				^5C				^5C	165.9
Hc’	5.67	^4’C		Hc	5.43	^4C		Hc	4.56	^4C	70.2
Ha’, Hb’	4.63, 4.49	^3’C		Ha,Hb	4.32, 4.22	^3C		Ha,Hb	4.78, 4.53	^3C	66.5
Hd’, He’	3.22, 3.10	^2’C		Hd, He	1.05	^2C		Hd, He	2.46	^2C	38.9
NH2’	7.61	^1’C	17.47	NH2	7.61	^1C		NH2	7.03	^1C	173.6

Table 8.

¹H and ¹³C-NMR chemical shifts (ppm) observed and calculated for chiral Polyesteramide PTOBEE-amide.

5. Thermal stability and differential scanning calorimetry (DSC)

The presence of choline group at the end of polymer chains causes in PTOBDME-choline a decrease in the thermal stability range compared to precursor PTOBDME. A 5% weight loss is observed for PTOBDME-choline at 230°C, while PTOBDME loses 5% weight at about 280°C. The thermal stability of PTOBEE-Choline is similar to that of polyester PTOBEE. PTOBEE-choline has 5% weight loss at 281°C, and PTOBEE at 280°C (see Figures 6 and 7). In the thermal stability curve of PTOBDME-choline, the first degradation step observed at 230°C is followed by two other weight loss step at 280 and 448°C. Two decomposition steps are observed at 280 and 466°C in PTOBEE-choline.

Figure 6.

(a) Thermogravimetry of PTOBDME-choline and PTOBEE-choline; (b) DSC analysis of PTOBDME-choline and (c) DSC of PTOBEE-choline, both at 10°C/min.

Figure 7.

(I) Thermogravimetry of precursor PTOBDME; (II) DSC analysis of PTOBDME; (III) DSC of PTOBEE: (a) first heating process of the original sample, (b) subsequent cooling down, (c) second heating process; and (d) DSC of (─)PTOBEE. First heating run of the original sample. All at 10°C/min.

In the DSC experiment of PTOBDME-choline, performed at 10°C/min, Figure 6(b), a glass transition can be observed at 58.2°C, in the first heating run, and a weak endothermic peak at 99.5°C is interpreted as due to the first order transition from crystal phase to liquid crystal state. An exothermic peak at 171.2°C is also observed which is not explained, but the beginning of a second endothermic peak at 200°C can be attributed to fusion to the isotropic. In the cooling process, two exothermic peaks at 155°C and at 175 are observed, probably associated to crystal formation. In the second heating, a very broad endothermic peak at 100.2°C is observed again associated to the transition to liquid crystal mesophase.

In the DSC experiment of PTOBEE-choline at (10°C/min), Figure 6(c), a glass transition can be observed at 60°C, and an endothermic peak at 130.2°C is attributed to the transition crystal to liquid crystal. A decreasing of baseline from 183.7°C to the end of heating was also observed in the first heating run due to a nonconcluded endothermic process or to the beginning of degradation to the polymer. A broad exothermic peak observed the cooling around 145°C would correspond to a crystallization from the mesophase state. In the second heating, only two glass transitions can be observed at 65 and 85°C.

The presence of ammonium chloride group at the end in the polymer chains, in Figure 8-I, produces a decrease of the thermal stability range compared to precursor polyesters. At 278°C, PTOBEE-ammonium chloride loses 10% weight and PTOBDME-ammonium chloride at 260° C, while precursor PTOBDME and PTOBEE at 310°C. In the thermal stability curve of the ammonium-polymers, the first degradation step observed at 228 and 230°C, respectively, was not observed in PTOBEE and PTOBDME. The two next inflexion points at 310 and 311°C and 466 and 471°C were equivalents to the observed in the precursor polyesters, which would indicate the same type of decomposition to principal core of the chain.

Figure 8.

(I) Thermogravimetric curve of PTOBDME-ammonium and PTOBEE-ammonium; (II) DSC analysis of PTOBDME-ammonium; (III) and PTOBEE-ammonium chloride. All at 10°C/min of.

In the DSC experiment of PTOBDME-ammonium chloride, at 10°C/min, Figure 8-II, a very broad exothermic peak centered at 96.8°C, is observed in the first heating, associated to low enthalpy value, which can be attributed to crystal to crystal transitions, involving molecular reordering between crystalline phases. An endothermic peak at 146.9°C is interpreted due to the transition to liquid crystal mesophase; finally, an exothermic peak at 186.8°C is observed. In the cooling run, very weak exothermic peaks at 154.4 and at 104.1°C were observed due crystallization process. In the second heating, a broad exothermic peak centered at 75.2°C, an endothermic peak at 149.1°C, and finally, an exothermic peak at 179.8°C were observed again.

The DSC experiment of PTOBEE ammonium choride, at 10°C/min, Figure 8-III, shows in the first heating run a broad exothermic peak centered at 69.1°C, and a very strong endothermic peak at 146.2°C due to the fusion transition from crystalline phase to liquid crystal mesophase, and finally, a weak endothermic peak at 173.3°C, perhaps due to a partial fusion to isotropic. During the cooling, an exothermic peak appeared at 166°C would correspond to a crystallization from the mesophase state, and in the second heating, the broad exothermic peak observed in the first heating was observed to higher temperature centered at 114.8°C; the two endothermic peaks were again observed at 147.6 and 170.1°C.

The thermogravimetric curve and the DSC analysis of PTOBUME-amide are given in Figure 9. At 265°C, it loses 5% weight. At 340°C, a first decomposition step begins, followed by another three at 400, 450, and 510°C. In the first heating of the DSC, an endothermic peak is observed at 160°C interpreted as the transition to the mesophase state. In the cooling run, several week exothermic peaks could be associated to crystal formation processes.

Figure 9.

(a) Thermogravimetric curve of PTOBUME-amide; (b) DSC analysis of PTOBUME-amide.

6. Optical characterization

6.1. Optical activity of PTOBDME-ammonium and PTOBEE-ammonium

As in the polyester precursors PTOBEE-ammonium chloride and PTOBDME-ammonium chloride presented an unexpected optical activity and chiral morphology, although they were synthesized starting from equimolar quantities of TOBC and the racemic mixture of the corresponding glycol. The obtained chirality has been evaluated by optical rotatory dispersion, in Figure 10, the values of optical activity are given as [α]^25°C, at different wavelengths. Table 9 shows the measured values.

Figure 10.

Optical activity of PTOBDME-ammonium chloride and PTOBEE-ammonium choride. Expressed as [α]^25°C in DMSO-d₆ at different wavelengths.

Polymers (0.2 g/100 ml in DMSO)	Hg (365 nm)	Hg (435 nm)	Hg (546 nm)	Hg (578 nm)	Na (589 nm)
PTOBEE-ammonium chloride	+12.44°	+5.75°	+2.85°	+3.44°	+5.75°
PTOBDME-ammonium chloride	+9.00	+5.06	−0.55	−1.15	−4.04

Table 9.

Optical activity of PTOBDME-ammonium and PTOBEE-ammonium, expressed by optical rotatory dispersion.

In the optical characterization of precursor cholesteric liquid crystal polyesters [1, 3], even an increase of chirality was observed for a second fraction of the polymer, obtained by precipitation, after days of reaction of the liquors mother with respect to the initial first fraction of the polymer. The optical activity of PTOBDME-choline, PTOBEE-choline, PTOBUME-amide and PTOBEE-amide, has not been studied at the end of the present article but will be reported in the future.

7. Conclusions

The synthetic methods of six new multifunctional cationic cholesteric liquid crystal polymers designed as PTOBDME-choline [(C₃₄H₃₆O₈)_n─C₅H₁₃N]; PTOBEE-choline [(C₂₆H₂₀O₈)_n─C₅H₁₃N]; PTOBDME-ammonium [(C₃₄H₃₆O₈)_n─C₅H₁₃N]; PTOBEE-ammonium [(C₂₆H₂₀O₈)_n─C₅H₁₃N]; PTOBUME-amide (C₃₃H₃₃O₉N)_n and PTOBEE-amide (C₂₆H₁₉O₉N)_n are given and their characterization by ¹H, ¹³C-NMR, COSY, and HSQC is reported.

The NMR analysis let us to conclude that the enantiomeric polymer chains present stereo regular head-tail, isotactic structure, explained in terms of the higher reactivity of the primary hydroxyl group in the glycol, with respect to the secondary one, through the polycondensation reaction.

According to our previous experience, each enantiomer, with two independent sets of signals observed by ¹H and ¹³C-NMR, differentiated with apostrophe (‘) and without it ( ), could be attributed to two diastereomeric conformers: gg and gt, related with two possible staggered conformations, of the torsion along the chemical bond containing the asymmetric carbon atom in the spacer, along the copolymer backbone, with two possible helical screw sense of the polymer chain and in all the studied polymers. Chirality in racemic PTOBDME was proposed to be due to the kinetic resolution of a preferable helical diastereomer, such as Sgt, with respect to the possible four forms, while the R/S ratio of asymmetric carbon atoms remained 50:50.

The presence of choline group or ammonium chloride groups at the end of polymer chains causes in precursor polyesters a decrease in their thermal stability range. PTOBDME-choline losses 5% weight at 230°C (PTOBDME at 280°C). The thermal stability of PTOBEE-choline is similar to that precursor PTOBEE, with 5% weight loss at 281°C.

At 260°C, PTOBDME-ammonium loses 10% weight and PTOBEE-ammonium at 278°C (precursor PTOBDME and PTOBEE at 310°C).

All the synthetized cationic liquid crystal polymers show in DSC an endothermic peak assigned to the first order transition from crystalline phase to liquid crystal mesophase: PTOBDME-choline at 99.5°C; PTOBEE-choline at 130.2°C; PTOBDME-ammonium at 146.9°C; and PTOBEE-ammonium at 146.2°C.

At 265°C, PTOBUME-amide loses 5% weight. At 340°C, it has a first decomposition step, followed by another three at 400, 450, and 510°C. In the DSC first heating, it shows the endothermic peak due to the mesophase transition at 160°C.

Optical ORD values are provided for the second fractions of PTOBDME-ammonium and PTOBEE-ammonium.

Acknowledgments

The author thanks Dr. Javier Sanguino Otero for his valuable help during the development of this Project. She also thanks the financial support obtained in the Project “Nuevos vectores no virales basados en polímero cristal-líquido colestérico (PCLC) y su uso para transfección génica”. PTR1995-0760-OP.