Preparation and thermal stability of optically active 1,2,4-triazolium-based ionic liquids

The synthesis of optically active ionic liquids, in a four-step reaction sequence, is described. In the first step an oxirane ring of cyclohexene oxide was opened with 1,2,4-triazole, yielding a racemic mixture of (1R,2R)and (1S,2S)-2-(1H-1,2,4-triazol-1-yl)cyclohexanol. Kinetic resolution of the racemate by a lipase catalyzed transesterification with vinyl acetate followed by alkylation (quaternization) of the triazole ring resulted in the appropriate optically active salts formation. After the anion metathesis, thermally stable novel chiral ionic liquids were obtained.


Introduction
The growing interest in recent years in chiral ionic liquids inclined us 1 to prepare and investigate some properties of a few 1,2,4-triazolium-based salts.Ionic liquids (ILs) are salts consisting of big organic cations and inorganic or organic anions with melting points lower than 100 °C (373.15K). 2 The existence of an enormous range of cation-anion combinations, 3,4 gives lot of possibilities for modification of IL's structure resulting in diverse chemical and physical properties.By changing the cation or anion in an ionic liquid molecule, such property as density, melting point, viscosity, or solubility in water or other solvents -can be changed and fine-tuned.That is why ILs are often called 'designer solvents' or considered as 'task-specific' compounds, which fulfill and influence the outcome of technological demands in various applications.At present, ionic liquids are widely used as solvents in chemical synthesis 5 as well as in electrochemistry 6 and in reactions carried out with enzymes or microorganisms. 7Obviously, beside achiral ILs, their chiral, optically active analogues (CILs), became a subject of intensive study in recent years.As it was established, chiral ionic liquids can act as catalysts for asymmetric induction 8 or as supplements influencing reaction stereoselectivity. 9They can also be used as a chiral solvents in stereoselective polymerization, 10 as a chiral phase for gas chromatography 11 or as chiral shift reagents in NMR. 12 They were also applied in production of chiral liquid crystals. 13Beside the applications mentioned above, some ILs posses special and unique antibacterial or antifungal activities, which is promising for future development of new disinfectants, sanitizers, preservatives or highly toxic biocides. 14For example Pernak et al. 15 obtained ionic liquids based on acesulfamate and choline, and tested them as insect feeding deterrents, fixatives for soft tissues in histopathological diagnosis and preservatives for blood.
It was found that quaternized nitrogen heterocycles in IL molecules give usually lower melting compounds than those containing aliphatic ammonium ions, and for this reason, mostly imidazolium or pyridinium cations were used in ionic liquids preparation.A few achiral ILs containing a quaternized triazole ring in the molecule are also described.Because the geometry and coordinating properties of 1,2,4-triazoles are similar to those of imidazoles, one can also expect they can provide interesting and valuable ionic liquids.Moreover, as it is known that the presence of a 1,2,4-triazole ring in a molecule often creates interesting pharmacological activities, many 1,2,4-triazole derivatives exhibit antibacterial, antifungal, anticancer, antitubercular, analgesic or anti-inflammatory properties. 16Unique physical properties of ILs in combination with their possible biological activity seemed to be very interesting, and inclined us 1 to prepare a few new 1,2,4-triazolium based ionic liquids.A similar investigation was undertaken by Spanish researchers 17 and some of the results are convergent, however the employed procedures and final products were different.

Results and Discussion
Herein, we describe a simple and efficient synthetic procedure for the preparation of optically active triazolium quaternary salts as potential ionic liquids.The salts were prepared from enantiomerically enriched (1S,2S)-2-(1H-triazol-1-yl)cyclohexanol and (1R,2R)-2-(1H-triazol-1yl)cyclohexyl acetate obtained through lipase mediated kinetic resolution of the racemate.The starting racemic trans-(±)-2-(1H-1,2,4-triazol-1-yl)cyclohexanol (±)-3, was synthesized according to the method described by Yus and co-workers 18 by stirring equimolar amounts of 1,2,4-triazole (2) and cyclohexene oxide (1) without catalyst and in the absence of any solvent.We found that the solvent-free ring opening of cyclohexene oxide by 1,2,4-triazole is regioselective and the reaction yield can be improved by a slight increase of the temperature as well as the extension of the reaction time (Table 1).a Isolated yield of pure product after recrystallization from i-PrOH.
Kinetic resolution of the racemic alcohol enantiomers (±)-3 was performed by enantioselective acetylation catalyzed by commercially available lipases using a 3-fold molar excess of vinyl acetate as the acyl donor (Scheme 1).In preliminary studies of enzyme catalyzed acetylation of (±)-3, we have checked various solvents to establish the most convenient one for the reaction.The media employed were chloroform, MTBE, THF, 1,4-dioxane and 2-methyl-2butanol (tert-amyl alcohol).The reactions carried out in chloroform, MTBE and THF were sluggish, due to the low solubility of the substrate, and much higher reaction rates were observed in dioxane and 2-methyl-2-butanol.These two solvents were tested in order to find the proper lipase for the reaction.For that purpose, two of the most frequently used immobilized enzymes were examined, the lipase from: Pseudomonas cepacia (Amano PS-C) and Candida antarctica (Novozym SP 435).
In most cases, obtained enantioselectivities (Table 2) were fairly good but not fully satisfying, except for the entry rac-2.Nevertheless, the initial screening experiments reveal useful information: the ractions carried out in 2-methyl-2-butanol run faster than in dioxane, and Amano PS-C is a more effective catalyst than Novozym 435 (Table 2: entry rac-1 vs rac-3, rac-2 vs rac-4).Furthermore, we found that the reactions catalyzed by Amano PS-C gave a better product enantioselectivity.Thus, the most effective kinetic resolution of (±)-3 was achieved in 2-methyl-2-butanol, at room temperature in the presence of Amano PS-C lipase.On the basis of these findings, the reaction was studied on a bigger scale and the process was slightly modified by increasing the temperature to 30 °C and terminating the experiment when 50% of conversion was achieved.Both the substrate and the product were isolated with high enantiomeric excesses (98%) (enantioselectivity of the reaction E >200) and in satisfying yields (Table 2: entry rac-5).The best result in terms of optical purity of the residual alcohol (>99%) was obtained when the conversion slightly exceeded 60% (Table 2: entry rac-6).
The stereochemical preference of the PS-C lipase in the acetylation reaction of (±)-3 towards one of the enantiomers was determined by assignement of the absolute configuration of the unreacted alcohol (±)-3 by the method based on a double derivatization described by Mosher. 20his was achieved by transformation of the unreacted alcohol enantiomer into two diastereomeric esters by reacting it separately with the chiral auxiliaries [(R)-and (S)-αmethoxy-α-phenylacetic acid (MPA)] and comparison of the chemical shifts in 1 H NMR spectra of the resulting two derivatives (Figure 1).On the basis of the finding 21 that in the α-methoxy-α-phenylacetic acid esters of secondary alcohols the most representative and stable conformer (by 0.6-1.0kcal/mol) is the sp conformer [in which the methoxy group, the Cα carbon, the carbonyl group of the MPA fragment, and the H(7') hydrogen of the alcohol fragment are in the same plane (see Figure 2)], and evaluation of differences in chemical shifts of the appropriate protons in the esters, the absolute configuration of the investigated enantiomer has been assigned as (1S,2S).
According to Riguero et al. 21the structure can be also calculated on the basis of changes in proton chemical shifts of the asymmetric carbon substituents in the substrate (L1 -triazole protons and L2 -protons of -CH2 group in cyclohexane ring) in both diastereisomers.These differences in the chemical shifts are represented by Δδ and it is the sign of this parameter (+ or -) that provides information about the configuration.For a particular substituent (e.g., L1), Δδ is defined as the difference in chemical shifts of a given signal of the substituent (δL1) in the two considered spectra (diastereoisomers).
Δδ RS L1 = δL1(R) -δL1(S) = 7.98 -7.68 = 0.30 > 0 Δδ RS L2 = δL2(R) -δL2(S) = 2.04 -2.23 = -0.19< 0 The calculated result and examination of the NMR spectra of the (R)-and (S)-MPA esters are consistent with the three-dimensional structure proposed (Figure 2).It is obvious that if the absolute configuration of the slower reacting enantiomer of (±)-3 is (S) on C(7') carbon, both protons in the triazole ring H(3') and H(5') of the (S)-MPA ester are shielded by the phenyl ring (due to the space-orientated anisotropic effect) while the same protons in the (R)-MPA ester remain unaffected.The opposite effect is observed for aliphatic protons H(8') which are shielded in (R)-MPA derivative while in (S)-MPA remain unaffected.The suggested assignment is also in agreement with Kazlauskas rule, 22 according to which the preferably accepted enantiomer in lipase catalyzed transesterification reactions posses (R) configuration on the alcoholic center.
In Figure 3 the molecular structure of (+)-3 is shown.The crystal structure was chiral but not polar and was composed of antiparallel polar hydrogen-bonded helical chains passing in the b crystallographic direction with the intermolecular hydrogen bonds O( 12)-H( 12 3: entry 2).The second notice concerns the reaction of (+)-3 with ethyl bromide, which can be performed successfully only in pressurized reactor (Table 3: entry 3).The synthesized triazole salts with halogen anions (5a-b; 6a-h) are solid.The exchange of bromide anion to the trifluoroacetate with use of (CF3COO)2Pb led to the corresponding liquid at room temperature salts 7a-d, obtained with excellent isolated yields (94-99%) (Table 4).DSC measurements exhibited that these novel CILs have a glass transition temperature (Tg) ranging from -17 to -9 °C.Triazolium salts containing trifluoroacetate as a counteranion can be considered to be chiral RTILs since DSC plots are unambiguously characteristic for amorphous materials for all the cases studied.DSC determinations revealed that all CILs had good thermal stabilities up to at least 150 °C.The corresponding DSC traces are shown in Figure 4.

Conclusions
New types of optically active, liquid at room temperature triazolium salts have been prepared.Lipase-catalyzed transesterification was established as a simple, efficient and straightforward technique for the kinetic resolution of trans-(±)-2-(1H-1,2,4-triazol-1-yl)cyclohexanol.Due to the method based on the double derivatization, the absolute configuration of the chiral intermediate product of CILs was deduced as (1S,2S)-2-(1H-1,2,4-triazol-1-yl)cyclohexanol.Furthermore, an X-ray diffraction analysis of a single crystal of alcohol (+)-3 unambiguously confirmed the proposed chemical structure as the (1S,2S)-configuration.Until now, very few racemic and enantiomerically pure triazolium quaternary salts with melting points below 100 °C are known.We here report that (1S,2S)-2-(1H-1,2,4-triazol-1-yl)cyclohexanol can be selectively quaternized at the N-4 atom in the triazole ring by using a threefold molar excess of alkyl iodides or bromides yielding solid salts, which after anion exchange become liquids at room temperature.The metathesis was achieved with lead trifluoroacetate, and has proven to be a novel versatile procedure for anion exchange in ILs.Four new triazolium salts which are claimed to be ionic liquids show low glass transition temperatures and possess a high degree of thermal stability.Antibacterial and antifungal activities of the prepared compounds are being evaluated.

Experimental Section
General.All commercially available reagents (Aldrich, Fluka and POCH) were used without further purification.Novozym SP 435 (lipase from Candida antarctica immobilized on a macroporous acrylic resin), and Amano PS-C (lipase from Pseudomonas cepacia immobilized on ceramic particles) were purchased from Novo Nordisk Co. and Amano Pharmaceutical Co. respectively and were used without any treatment.Melting points were obtained with an MPA100 Optimelt SRS apparatus.Thin-layer chromatography was carried on TLC aluminum plates with silica gel Kieselgel 60 F254 (Merck) (0.2 mm thickness film) and the compounds were visualized in iodine vapors.Preparative plate chromatography was performed with DC-Fertigplatten Kieselgel 60 F254 (5 x 20 cm with 0.25 mm thickness layer).The chromatographic analyses (GLC) were performed with an HP Series II 5890 instrument equipped with a flame ionization detector (FID) and fitted with HP-50+ (30 m) semipolar column.Helium (2 mL/min) was used as carrier gas; Tinjector 280 °C, Tcolumn 100 °C (3 min) and 100-280 °C (10 °C/min); retention times (tR) are given in minutes under these conditions.Column chromatography was performed using Silica gel 60 (Merck) of 40-63 μm.Mixture of 95:5 v/v chloroform/methanol was used as eluent.The enantiomeric excesses of resulting esters and alcohols were determined by HPLC analysis which were performed on a Shimadzu CTO-10ASV equipped with UV detector STD-20A and chiral column Chiralcel OD-H (Diacel), using mixtures of nhexane/isopropyl alcohol as mobile phase in appropriate ratio given in experimental section; flow (f) is given in mL/min; racemic alcohols and esters were used as standards.Optical rotations were measured on a P20 polarimeter (Belligham & Stanley Ltd., line D spectrum of sodium) in 2 dm of length cuvette.Absorption of electromagnetic radiation waves from UV/VIS extent was made on spectrophotometer Cary 3. The X-ray data were measured using Xcalibur R Oxford Diffraction apparatus with a ccd camera-detector applying CuKα monochromatic radiation. 1H NMR and 13 C NMR spectra were measured on a Varian Mercury 400BB spectrometer, operating at 400 MHz for 1 H nuclei and 100 MHz for 13 C nuclei, if not indicated otherwise; chemical shifts (δ) are given in parts per million (ppm) related to tetramethylsilane (TMS) as internal standard; multiplicity (s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet), coupling constant (J) in hertz (Hz) assignment.Mass spectra were recorded on a Micro-mass ESI Q-TOF spectrometer at the Mass Spectrometry Laboratory, Institute of Biochemistry and Biophysics (IBB), PAN.IR spectra were measured with SPECORD M80 spectrometer.Samples were prepared in paraffinic oil.Elemental analyses were performed on a Elementar Analysensysteme GmbH -VARIO EL III (Element Analyzer: CHNS).Glass transition temperatures (Tg) were recorded on a Thermal Analysis DSC Q200 differential scanning calorimeter with heating rate at 5 °C/min after initially cooling samples from -70 to 150 °C under nitrogen in Tzero hermetic aluminium pans.

General procedure for anion exchange (7a-7d)
To the stirred solution of the appropriate bromide salt (0.1 g) in distilled water (3 ml), (CF3COO)2Pb was added drop by drop at 0 °C till all bromide anions became precipitated.Then the reaction suspension was filtrated by using Pasteur pipette tipped with wool.To a sustain permeate toluene (15 ml) was added in order to remove the last trace of water azeotropicly.This procedure was repeated 5-times and then the solid residue was rinsed with diethyl ether (3 x 3 ml).The collected washings were evaporated under reduced pressure yielding brown semitransparent oil.The oil was diluted with acetone (2 ml) and filtered through active carbon.The product was evaporated to dryness to give transparent colorless oil.

Figure 1 .
Figure 1.Newman projection of model for configurational correlation of MPA esters.Red marked protons are shielded by the phenyl ring of chiral auxiliary (MPA).

Figure 4 .
Figure 4. DSC plots for a family of enantiopure chiral triazolium salts.

Table 4 .
Properties of CILs after anion metathesis in triazolium salts a Glass transition temperature; the data were determined by DSC.b c solution in chloroform.