WO2018189199A1 - Pyridinium salts and uses thereof - Google Patents

Abstract

A 1,3,5-trisubstituted pyridinium salt of general formula I (I) wherein Xn- is a counterion in which n is an integer from 1 to 4; wherein R2 and R3 are each independently a substituent group comprising Ar, wherein Ar is an aryl or heteroaryl group, which is substituted or unsubstituted; wherein R1 is a substituent group linked to the quarternary nitrogen of the pyridinium ring by an aliphatic carbon; and wherein, when R1 is benzyl, R2 and R3 are not both phenyl, 3-MeO phenyl, 4-Br phenyl, 3-CF3 phenyl, 2-Me phenyl or 4-(4-tert BuPh) phenyl; when R1 is 3-MeO phenethyl, R2 and R3 are not both 3-MeO phenyl; and when R1 is dimethoxyphenethyl, R2 and R3 are not both phenyl.

Description

Pyridinium salts and uses thereof

The present invention relates to pyridinium salts, uses of pyridinium salts as antimicrobials and medicaments and processes for their production.

Background of the Invention

Polysubstituted pyridine and pyridinium salts such as the coenzymes NAD⁺ and NADP⁺ [ 1 ] are involved in many essential biochemical processes. They are also part of other biologically active natural products such as juliprosine [2], the antibacterial alkaloid ficuseptine [3-5], and the neurotoxic metabolite 1 -methyl-4-phenylpyridinium (MPP⁺) [6,7] (Scheme A). In addition, synthetic pyridiniums have been developed to facilitate gene delivery [8], or act as platelet activation antagonists [9], and benzyl idenehydrazinyl pyridiniums have been investigated as antimicrobial agents [ 10]. The synthesis of polysubstituted pyridiniums however typically involves multistep procedures, or harsh reaction conditions, and the development of rapid and mild methodologies for the facile preparation of new polysubstituted pyridinium analogues is sought after.

The Chichibabin reaction for the synthesis of polysubstituted pyridines was first reported nearly 100 years ago [ 1 1]. The reaction involves the condensation of aldehydes with an amine to yield pyridiniums in a single synthetic step. Initially, the reaction required elevated temperatures and pressure, or acid catalysis [ 12-17], but the intrinsic instability of aldehydes under these forcing conditions often resulted in product yields that were at best mediocre.

Typically, three products can be isolated from the acid-mediated condensation of amines with acetaldehydes in the Chichibabin reaction (Scheme 1); a 1,2,3,5-tetrasubstituted pyridinium 1 (often the major product), which is formed via the auto-oxidation of the product 1,2,3,5- dihydropyridinium 2, and 1 ,3,5-trisubstituted pyridinium salts 3 (a minor product often referred to as the 'abnormal' Chichibabin product).

Scheme 1. Three products 1-3 isolated from the acid-mediated Chichibabin reaction.

Triflate salts of lanthanides have been used in room temperature reactions to promote the Chichibabin condensation reaction between amines and acetaldehydes, giving 1, 2, or 3 in varying amounts [18]. More recently, the condensation of benzylamine and aryl acetaldehydes in a 50 mol% solution of ytterbium triflate in water provided the first report of a selective synthesis of pyridinium salts 3 [19-21]. However, the use of such rare-earth Lewis acid catalysts raises sustainability and cost issues. Glacial acetic acid [21] was also reported to promote the condensation of phenethylamine and phenylacetaldehyde to the pyridinium 3 but such harsh reaction conditions are unsuitable for intrinsically unstable substrates such as arylacetaldehydes.

An aim of the present invention is to provide an improved process for the synthesis of polysubstituted pyridinium salts and novel pyridinium salts and uses thereof.

Summary of the Invention

Accordingly, in a first aspect, the present invention provides a 1 ,3,5-trisubstituted pyridinium salt of general formula I

whe ein X^n~ is a co nterion in which n is an integer from 1 to 4;

wherein R² and R³ are each ndependently a substituent group comprising Ar, wherein Ar is an aryl or heteroaryl group, which is substituted or unsubstituted;

wherein R¹ is a substituent group linked to the quarterna y nitrogen of the pyridinium ring by an aliphatic carbon; and

wherein, when R¹ s benzyl, R² and R³ are not both phenyl, 3-MeO phenyl, 4- ϊ r phenyl, 3-CF₃ phenyl, 2-Me phenyl o 4-(4-tert BuPh) phenyl;

when R¹ is 3-MeO phenethyl, R² and R³ are not both 3-MeO phenyl; and

when R¹ is dimethoxyphenethyl, R² and R³ are not both phenyl.

In a second aspect, the present invention provides se of a 1,3,5-trisubstituted pyridin^"u salt of general formula I as an antimicrobial,

wherein X^n~ is a counterion in which n is an integer from 1 to 4;

whe ein R² and R³ are each independently a substituent group comprising Ar, wherein Ar is an aryl or heteroaryl group, which is substituted or unsubstituted;

wherein R¹ is a substituent group which comprises (i) Ar¹, wherein Ar¹ is an aryl or heteroaryl group which is substituted or unsubstituted; or (ii) an aliphatic group compris^'ng at least 6 carbon atoms, wherein the substituent group is linked to the quarternary nitrogen of the pyridinium ring by an aliphatic carbon; and wherein R¹ is not 4-hydroxyphenethyl or 4-carboxyphenethyl when R² and R³ are both phenyl; and

where^'n R¹ is not 3-hydroxyphenethy or 3-hydroxyphenyl 1 -hydroxy prop-2-yl when R² and R³ are both 3,4-dimethoxyphenyl.

In a third aspect, the present invention provides a process for the product^'on of a 1,3,5- trisubstituted pyridinium salt, which comprises reacting an aryl or heteroaryl acetaldehyde with an amine in the presence of a phos - hate catalyst and water, so as to form the 1,3,5-trisubstituted pyridinium salt.

It has surp isingly been found that "abnormal" Chichibabin pyridin^'um salts tri-substituted at positions 1,3 and 5 may be produced in h^'gh yield by reacting the ary or heteroaryl acetaldehyde with the amine in the presence of a phosphate catalyst n aqueous conditions. The reaction is facile, may be performed in one step and tolerates ox datively sensitive aromatics. Th^'s enables rapid access to a wide ange of pyridinium salts hitherto unknown. The use of such reactants and relatively mild reaction conditions enables the pyridinium salts to be prepared inexpensively. The eactants used under these reaction conditions also provide a process which is environmentally friendly.

Pyridinium salts obtainab e by the process of the invention have been found to have antimicrobial and, in particular, antibacterial properties which may make them useful in antibioitic, antiseptic and antifouling applications.

According to the first aspect, a 1,3,5 tri-substituted pyridinium salt of general formula I is provided in which R¹ is the substituent at pos^'tion 1, R² is the substituent at position 3 and R³ is the substituent at position 5 of the pyridin^'um ring. R¹ may be any substituent group provided that it is linked to the quarternary nitrogen of the pyridinium ring by an aliphatic carbon, t has been observed that there is a lack of reactivity when aromatic amines s ch as aniline 6m or 2- aminobenzimidazole 6n have been used in an attempt to make pyridinium salts according to the i vention. The presence of the aliphatic carbon adjacent the nucleophilic amine group appears advantageous in the production of the pyridinium salts of the invention. In one arrangement, R¹ comprises Ar¹, wherein Ar¹ is an aryl or heteroaryl group which is substituted or unsubstituted. According to this arrangement, Ar¹ may be linked to the quarternary nitrogen of the pyridinium ring by a linker compris^'ng one or more aliphatic ca bons, typically a C1-C3 linker such as a one or two carbon linker.

Ar¹ may be aryl or heteroaryl. Thiophene was found to be useful as a heteroaryl substituent and the pyridinium possessed good antibacterial activity. Aryl substituents, namely those not containing heteroatoms ^"n their rings, were generally found to possess antibacterial activity. It is preferred that Ar¹ contains no more than one or two aromatic rings. Larger structures may be stearically unfavourable.

Typical y, A ^{1 '}s unsubstituted, mono-substituted o di-substituted. For antibacte ial activity t is preferred that Ar¹ does not bear a 4-hydroxy subst^'tuent, 4-carboxy substitutent or a 3- hydroxy substituent at least when Ar¹ is mono-subst^'tuted. Where R² and R³ are both phenyl, R¹ is not 4-hydroxyphenethyl or 4-carboxyphenethyl because this compound does not possess antibacterial activity. Similarly, when R² and R³ are both 3,4-dimethoxyphenyl, R¹ is not 3- hydroxyphenethyl or 3-hydroxy -hydroxy prop-2-yl.

Generally, Ar¹ may bear one or two substituents selected from nitro, methoxy, hydroxyl and halogen. Halogen substituents, alone or in combination with another substituent such as hydroxyl, were found to have good antimicrobial act^'vity. Advantageously, Ar¹ bears a chloro or bromo substituent.

Typically, Ar¹ is phenyl or thiophene-2-yl.

In a further arrangement, R¹ is an aliphatic group which comprises at east six carbon atoms. It ^'s found that smaller aliphatic substituents such as C1-C5 substituents did not possess antibacterial activity. Where Ri is an aliphatic substituent, Ri will necessarily be linked to the quarternary n^'trogen of the pyridinium ring by an aliphatic carbon from the substituent. Ri may comprise hexyl or cyclohexyl. Hexyl is typically a straight chain hexyl optionally bearing a substituent such as hydroxyl. Where R¹ comprises cyclohexyl, this may be linked to the quarternary nitrogen directly or by a linker which may be a C1-C3 linker, such as a one or two carbon linker. The cyclohexyl may be substituted or unsubstituted. In one arrangement, the 1,3,5-tri-substituted pyridinium may be derivatised for pharmaceutical use, for example by derivatisation at R¹. a further arrangement, the 1,3,5-tri-substituted pyridinium may be adsorbed to, attached to or form part of a surface, for example at R¹. The surface may be any surface where it is desirable to confer antimicrobial properties. For example, the surface may be from a medical device such as a surgical device or implant. The surface may be a hospital surface where sterility or low microbial activity is required. The surface may be for food use, such as a food preparation surface or utensil. The surface may comprise sanitary ware. The surface may be a marine surface where antifoul^'ng properties are required.

R² and R³ are each independently a substituent g oup compris ng Ar, wherein Ar ^'s an ary or heteroaryl group, which is substituted or unsubstituted. Thus, the substituent gro p consists of Ar or inc udes Ar with other moieties. Although R² and R³ need not be the same, they may most straightforwardly arise from a common reactant such as an aldehyde as discussed herein. It is ikely that large Ar groups may be stearically unfavourable and so it is p eferred that Ar has no more than one or two aro atic rings. General y, the substituent group consists of Ar with no more than one aromatic ring. Advantageously, Ar is phenyl, preferably unsubsituted phenyl, 4-hydroxy phenyl or 4-methoxyphenyl. These substituents have been fo nd to exhibit useful antimicrobial behaviour. Where Ar is heteroaryl, th^'s may be thiophene-2-yl, which is another substituent which has exhibited antimicrobial behaviour. R² and R³ are preferably not 3,4-dimethoxyphenyl because such a subst^'tuent is less useful when used as an antibacterial. R² and R³ are preferably not indole substituents. Mono-substitued Ar groups are generally preferred.

The counterion X_n- ay be any suitable counterion, including CI-, Br-, CF₃C0₂-, CF₃CO₂-, SO4^2-, PO4^3-, and CO3^2-. The trifluoroacetate counterion is used in a number of examples described herein.

The 1,3,5-tri-substituted pyridinium salts of the present invention may be used as an antimicrobial agent. Such use includes inhibition or prevention of microbial cell growth. Such uses may be therapeutic or non-therapeutic or prevention. Typically, a therapeutic use comprises treatment or prevention of a condition in a subject in need of such treatment or prevention. The subject may be a h man or non-human animal subject. The treatment may be by any suitable ode of administration. An example of s ch treatment is topical treatment for an infection.

Pharmaceutical compositions suitable for med^'cal use may be provided which comprise a 1,3,5-tri-substituted pyridinium salt according to the invention together with a suitable carrier or excipient. In this aspect, the ,3,5-tri-substituted pyridinium salts of the nvention are provided for use as a medicament.

In a further aspect, the 1,3,5-tri-substituted pyridinium salts of the present i vention may be sed top cally on subjects as a disinfectant, for example to prevent carriage of microorganisms from one subject to another. Such use inc udes use as a microbial decontaminant. Such use may be regarded as non-therapeutic.

Further non-therapeutic uses include t eatment of inanimate surfaces for example to disinfect medical devices, surfaces for food use or hospital su faces.

The antimicrobial activity of the 1,3,5-tri-s bstituted pyridinium salts of the present invention is advantageously antibacterial activity, includ^'ng antibacterial activity against gram positive bacteria such as Staphylococcus, Bacillus, and Micrococcus, as well as Mycobacterium. Particular bacteria include Staphylococcus aureus, Staphylococc s carnosus, Bacillus subtilis, Mycobacterium aurum and Micrococcus luteus. The 1,3,5-tri-substituted pyridinium salts may also have antifungal activity, for example against Candida.

Particu arly preferred 1,3,5-tri-substituted pyridinium salts of the present invention are set out below:

According to the process of the present invention, an aryl or heteroaryl acetaldehyde is reacted with an amine in the presence of a phosphate catalyst and water so as to for a ,3,5-tr - substituted pyr dinium salt. It is thought that one molecule of the amine reacts w^'th three molecu es of the aryl o eteroaryl acetaldehyde in a condensation reaction so as to form the pyridinium ring. It is preferred that a mo ar excess of aryl or heteroaryl acetaldehyde over amine is present, preferably with a mo ar ratio of aryl or heteroaryl acetaldehyde to amine of 3 or more, or 4 or more such as in the range 4 to 6, for example around 5. Such a molar rat^'o produces the desired pyr^'din^'um salt in good yield.

Phosphate catalyst is present in a catalytically effective amount which is typically at least 0.05, advantageously at least 0.1M, preferably at least 0.2 , typically up to 0.5M. A preferred range is 0.2 to 0.3M, such as around 0.25M. The phosphate catalyst may be an inorganic phosphate or pyrophosphate, a sugar phosphate such as a disaccharide phosphate, for example glucose- 1- phosphate, or a nucleobase monophosphate such as uridine 5 '-phosphate.

The reaction may be performed at a temperature of at least 35°C, advantageously at at least 60°C and preferably at a temperature of 60°C to 100°C, such as around 100°C. It -s found that higher temperatures result in increased reaction rates without significant unwanted side reactions. The reaction may be performed for at least 2 hours to provide desired products, usual y for at least 10 hours, such as in the range 10 to 15 hours. A typ^'cal reaction time is 2 hours.

Typically, the reaction is performed at a pH in the range 5 to 7 and preferably around 6. At a pH below 6 slower reaction rates are observed. At a pH above 6 reaction conditions favour polymerisation of the acetaldehyde.

The reaction may be carried out in an aqueous medium which is typically buffered by the phosphate component. A cosolvent may be employed to aid the solubility of the amine and aldehyde reactants and methanol may be selected as a suitable cosolvent, typically in a volume ratio of : 1. A water osolvent ratio above is poss^'ble where less cosolvent is used to solubilise the reaction components. Other suitable cosolvents include water-miscible cosolvents such as acetonitrile, ethanol or isopropanol. Su^'table amines and aldehydes are chosen so as to provide the 1,3,5-tri-substituted pyridinium salts as described here^'n. Thus, the amine may have the formula R¹-NH₂ where^'n R¹ s a substituent group linked to the nitrogen by an al^'phatic carbon. It is thought that the nitrogen of an amine acts as a nucleophile in the reaction. Accordingly, R¹ is preferably selected so as not to reduce the nucleophilicity of the nitrogen. R¹ may be defined as above.

The aryl or heteroaryl acetaldehyde typically has the formula R²-CH₂-CHO. Alternatively, the aryl or heteroaryl acetaldehyde may comprise a mixture of compounds of the formulae R²- CH₂-CHO and R³-CH₂-CHO. R² and R³ are each independently a substituent group compr^'sing Ar, wherein Ar is an aryl or heteroaryl group, which is substituted or unsubstituted. R² and R³ may be defined as above.

The invention will now be described in further detail, by way of example only, with reference to the following specific Examples.

Detailed Description of the Invention

Examples

1. Phosphate-mediated synthesis of 1,3,5-pyridiniums

We have recently reported a phosphate mediated biomimetic Pictet-Spengler synthesis of THIAs [22]. The mild reaction conditions allowed even the least stable arylacetaldehydes to generate THIAs in good yields [22-24]. This biomimetic Pictet-Spengler condensation reaction is selective towards the amine component, requiring a phenethylamine substrate that is meta- substituted with a strong electron-donating group (e.g. a hydroxyl group) such as ^"n 3- hydroxyphenethylamine. The condensation of this phenethylamine for example with an aldehyde such as phenylacetaldehyde 4a in potassium phosphate (KPi) buffer gave the THIA 5 (Scheme 2). Chichibabin and Pictet-Spengler condensations both involve amine and aldehyde components, imine formation and activation, and therefore by analogy with the Pictet-Spengler reaction we postulated that the Chichibabin reaction could be promoted by phosphates. This hypothesis was investigated by reacting phenylacetaldehyde 4a with tyramine (4- hydroxyphenethy amine) 6a (unreactive under biomimetic Pictet-Spengler conditions) in a 1.2: 1 ratio (as reported for the synthesis of THIAs) in KPi buffer (pH 6), at 60 °C for 12 h. No THIA was fo ed, but seve al compounds were detected in trace quantities, with the main product of the reaction (5% yield) being the 'abnormal' Chich^'babin pyr^'dini m salt 3a (Scheme 2).

In order to optimise reaction conditions to favour formation of the pyridinium product 3a, the ratio of 4a to 6a was inc eased from 1.2: 1 to 5 : 1 to account for the stoichiometry of the reaction, and a range of buffers were screened at 0.1 M (pH 6) as media for the reaction (Table 1).

Table 1. Influence of bu fer conditions on the formation of 3a (Scheme 2(ii)).¹

The four phosphate-based buffers tested (Table 1, entries 1-4) catalysed the production of pyridinium salt 3a, whereas other buffers or water alone (Table 1, entries 5-8) did not significantly promote the reaction. These results suggested that phosphates promote not only the synthesis of THIAs, but also that of 1,3,5-trisubstituted pyridinium salts 3. All four phosphates, glucose- -phosphate (Glc-l-P), inorganic pyrophosphate (PPi), uridine 5'- phosphate (UMP) and inorganic phosphate (Pi) are naturally abundant and essential to cell survival. They are involved in buffering cellular pH, for storing genetic information (UMP and other nucleotide building blocks for DNA and RNA) and transferring b^'ochemical i formation (PPi is a by-product of the hydrolysis of ATP mediated by kinases). Hence, the synthesis of

,3,5-trisubstituted pyridiniums, as we as THIAs, is likely to proceed at low levels under mild conditions in vivo. This could explain the occurrence of plant natural products such as the haouamines [19]. The in vivo condensation of aldehydes such as allysine with itself or with lysine has also been reported: the resulting polyfunct^'onal pyridinium salt cross-links n elastin s believed to be an age-related intermolecular cross-linking process [25,26].

The reaction conditions were f rther improved by increasing the KPi buffer concentration to 0.25 M and elevating the temperature to 00 °C. The reaction pH was however kept at 6 since more acidic conditions resulted in slowe eaction rates and more alkaline conditions favoured the polymerisat on of phenylacetaldehyde. Final y, the solubility of both amine and aldehyde substrates was enhanced by using a 1 : 1 mixture of KPi buffer with methanol. The combinat^'on of these new conditions was rewarded with the production of the 3a in 70% isolated yield. Aldehyde 4a was then reacted with a range of (hetero)arylethylamines 6b-6g, a benzylamine 6h, aliphat c amines 6i-61, and aromatic and heteroaromatic amines 6m, 6n to form the corresponding 1,3,5-t isubstituted pyridinium salts 6b-61 n 3-72% isolated yields (Table 2). Functionalities including hydroxyls, ha ogens and carboxylates were well tolerated. The lack of reactivity observed with aromatic amines such as aniline 6m or 2-aminobenzimidazole 6n may originate from their poor nucleoph^'licity as well as steric hindrance.

Table 2. One-step hosphate mediated synthesis of 1,3,5-pyridiniums 3.¹

In contrast to the amines, the reaction was highly selective towards aldehyde substrates (Table 3). While tyramine 6a reacted with arylacetaldehydes and (hetero)arylacetaldehydes, no reaction was observed with aliphatic aldehydes with the exception of 3-methylbutyraldehyde 4r: reaction of 6a with 4r produced the pyridinium 3r in a low yield of 5% and also the pyridinium lr (10% yield). The formation of lr suggested that the phosphate-mediated reaction was analogous to a Chichibabin type condensation where the aldehyde directed the reaction towards either the Chichibabin prod ct 1 via an oxidation reaction or 1,3,5-trisubstituted pyridiniums 3 via an elimination step (Scheme 1). Table 3. Versatility of the phosphate-mediated pyridinium synthesis towards aldehydes 4.¹

In this study, we have demonstrated that the synthesis of 3 can also be promoted by phosphates in good isolated yields. Phosphate acid/base catalysis can p omote keto-enol equilibria, proton transfers, and eliminations, which typically occur during the Chichibabin pyridine synthesis. Here, the excel ent nucleoph Tc and leaving group abilit^'es of phosphates may further facilitate the reaction. Here, in the final elimination step producing 3, loss of proton 3-H can be assisted by phosphate, and the substituent at C-2 then leaves. If the substituent at C-2 is a good leaving group (to uene in the case of arylacetaldehydes), 1,3,5-trisubstituted pyridiniums 3 will predominantly be formed (Scheme 1). Otherwise, the 1,2,3,5-tetrasubstituted intermediate will slowly oxidise to generate the corresponding 1,2,3,5-tetrasubstituted pyridinium Chichibabin salt 1.

2. Antimicrobial activity of 1,3,5-pyridiniums toward 5. aureus

There is a growing need for new antibiotics and over the last 30 years the number of new antibiotics entering the clinic has decreased significantly. During that time there has been a constant rise in the resistance of pathogenic bacteria to the antibiotics in use. One of the major pathogens in hospitals and increasingly in the community is Staphylococcus aureus. This bacterium has acquired resistance over the last 25 years to methici lin and in the last 15 years to vancomycin. In the light of th^'s need for new antibiotics to treat S. aureus infections, selected 1,3,5-pyr dinium compounds were tested for their ability to inhibit the growth of S. aureus. Agar plate diffusion assays and liquid min^'mal inhibitory concentration (MIC) tests were carried out to assess the potency of the compounds.

Some examples of the agar plate diffusion assays against a kanamycin control are shown in Figure 2 and the MIC data in Table 4. The data demonstrated the potency of some of the compounds compared to the antibiotic kanamycin.

Figure 2. Examples of the agar plate diffusion assays against S. a reus with selected pyridiniu

400 μg/mL) and the kanamycin control (at 100 μg/mL).

From the compounds tested it was notable that the two most potent anti-bacterials against S. aureus were compounds 3e and 3g (Figure 3). Both had an aromatic ring at C-3 and C-5 of the pyridinium ring, indeed for 3r where an isopropyl group was present, no growth inhibitory properties were noted. In addition, 3g had a 5-bromo-3-hyd oxypheny group attached via an ethyl spacer to N- , whereas in 3e a 3-bromophenyl group was present. Overall this data suggests that aromatic substituents at positions C-3 and C-5 are preferred, and that substitution of the phenyl rings is tolerated, as demonstrated by 3o and 3q. Also, that substitution of the N- 1 phenethyl group by a halogen leads to good inhibitory growth properties. However, it is not essential for an aromat^'c group to be present at N-l, for example as with 3j, for inhibitory growth effects to be conferred.

Figure 3. Structures o 3e a d 3g

These results point to compounds that can now be made to explore the structure activity relationship of the various substituents that can be placed on the C-l, C-3, and C-5 positions of the pyridine ring.

3. Experimental Section

3.1. General Information and Methods 3.1.1. Chemistry.

All reagents were obtained f om commercial sources and used as received unless otherwise stated. TLC was performed on Kieselgel 60 F₂₅₄ precoated plastic plates and compounds visual^'sed by exposure to UV light, potassium permanganate, phosphomolybdic acid (PMA) or ninhydr^'n. Flash column chromatography was carried out using silica gel (particule size 40- 63 μm). Preparative HPLC were performed on a Varian Prostar instrument equipped with an autosampler, a UV-vis^'ble detector and a DiscoveryBIO wide Pore C 8-10 Supelco co umn (25 x 2.12 cm). Elutions were monitored at 280 nm and carried out according to Gradient 1: 5 to 90% of aceton^'trile against water (0.1% trifluoroacetic acid). NMR: ¹H and ¹³C NMR spectra were recorded at 298 K at the field indicated using Bruker spectrometers Avance 500, and Bruker Avance ΙΠ 600. Coupling constants (/) are measured in Hertz (Hz) and multiplicities for ¹H NMR couplings are shown as s (singlet), d (doublet), t (triplet), hept (heptet) and m (multiplet). Chemical shifts (in ppm) are given relative to tetramethylsilane and referenced to res dual protonated solvent. Infrared spectra were recorded on a Perkin Elmer Spectrum 100 FTIR spectrometer. Mass spectrometry analyses were performed at the UCL Chemistry Mass Spectrometry Fac^' ity using a Finnigan MAT 900 XP and Micro Mass Quattro LC mass spectrometer: TFA refers to the CF₃C0₂ salt and NMR signals are not recorded for TFA n the ¹³C NMR data.

3.1.2. Biological Assays

Plate zone asssays: Staphylococcus aureus was grown overnight in LB broth (Oxoid) and 100 μL was spread onto the surface of LB agar plates using a ste ile spreader. A sterile glass pipette with a diameter of 6 mm was used to punch out wells and the agar re oved from the wells. 50 μL of compo nd, at a concentration of 400 μg/mL in RO water, or a control solution (positive control kanamycin at 100 μg/mL; negative control 1: 12.5 DMSO:water) was pipetted into each well. The LB Agar plates were then left to incubate at 37 °C for 18-24 h. The zone diameters were measured at 8 h.

Minimimal Inhibitory Concentration (MIC): 50 mL of LB broth was inocu ated with a mL overnight culture of Staphylococcus aureus, then 450 μ·L of the inoculated broth was pipetted into each wel of a 96 deep square well plate. 50 μ·L of compound was pipetted into each well; the compound was taken from the 256 μg/mL to 2 μg/mL dilution series. A 5 mg/mL stock concentration of the test compounds was diluted with Reverse Osmosis (RO) water in clean, sterile .5 mL Eppendorf Micro-centrifuge tubes to a starting concentration of 256 μg/mL. A 2-fold dilution series was made using RO water down to a concentration 2 μg/mL. There were 8 different dilutions in each series ranging from 256 μg/mL to 2 μg/mL. After growth of the plate at 37 °C for 18-24 h with shaking, the wells with no growth were recorded as the MIC.

3.2 Procedure A for synthesis of the 1 ,3, 5 -substituted pyridinium salts

Amine ( equiv.) and aldehyde (5 equiv.) were dissolved in 10 mL of a : 1 mixture of methanol/phosphate buffer (0.25 M solution at pH 6). The resulting solution was stirred at 100 °C for 12 h. The crude mixture was cooled to rt and pur^'fied by preparative HPLC (Gradient 1). Fractions containing the desired product were combined, concentrated under vacuum and co-evaporated with methanol (3 x 20 mL). l-(4-Hydroxyphenethyl)-3,5-diphenyIpyridinium.TFA (3a.TFA). Compound 3a was prepared accord^'ng to procedure A from tyramine.HCl (87 mg, 0.50 mmo ) and phenylacetaldehyde (300 mg, 2.50 mmol). The crude product was pur^'fied by preparat^'ve PLC (Gradient 1, retention t^' e 5.5 min) to give 3a.TFA as a pale yellow o^'l (162 mg, 70%). V_max(neat)/cm^-1 3064, 671, 16 3, 1596, 517; ¹H NMR (500 MHz; CD₃OD) δ 3.29- 3.32 (2 , m, CH₂CH₂N⁺), 4.95 (2H, t, J = 6.7 Hz, CH₂N⁺), 6.74 (2Η, d, J = 8.5 Hz, 3"-H, 5"- H), 6.98 (2H, d, J = 8.5 Hz, 2"-H, 6"-H), 7.56-7.60 (6H, m, Ph), 7.72-7.76 (4H, m, Ph), 8.91 (2H, d, J = .7 Hz, 2-H, 6-H), 8.95 ( H, t, J = .7 Hz, 4-H); ¹³C NMR ( 25 MHz; CD₃OD) δ 37.5, 64.6, 16.7, 127.2, 128.5, 130.6, 13 .1, 131.3, 34.5, 141.4, 141.7, 42.5, 58.1; m/z [HRMS ES+] found [M-TFA]⁺ 352.1700. C₂₅H₂₂NO⁺ requ res 352.1701. l-Phenethyl-3,5-diphenylpyridinium.TFA (3b.TFA). Compound 3b was prepared according to procedure A from phenethylamine (6 mg, 0.50 mmol) and phenylacetaldehyde (300 mg, 2.50 mmol). The crude product was purified by preparative HPLC (Gradient 1, retent^'on time 16.5 min) to give 3b.TFA as a pale ye low oil (121 mg, 54%). v_max(neat)/cm^_1 3064, 683, 597, 482¹H; NM ^. (500 MHz; C ·₃OD) δ 3.50 (2H, t, J = 6.9 Hz, CH₂CH₂N⁺), 5. 0 (2H, t, J = 6.9 Hz, CH₂N⁺), 7.28 (2H, d, J = 6.8 Hz, 2"-H, 6"-H), 7.33-7.41 (3H, m, Ph), 7.64-7.66 (6H, m, Ph), 7.80-7.83 (4H, m, Ph), 9.05 (1H, t, J = 1.6 Hz, 4-H), 9.07 (2H, d, J = 1.6 Hz, 2-H, 6-H); ¹³C NMR (125 MHz; CD₃OD) δ 38.2, 64.2, 17.9, 128.5, 130.0, 130.5, 131.4, 134.5, 136.9, 141.6, 41.7, 42.6; m/z [HRMS ES+] found [M-TFA]⁺ 336.1750. C₂₅H₂₂N⁺ requ res 336.1752. 3,5-Diphenyl-l-[2-(pyridin-2-yl) ethyl]pyridinium.TFA (3c.TFA). Compound 3c was prepared according to procedu e A from 2-(2-aminoethyl)pyridine (35 mg, 0.29 mmol) and phenylacetaldehyde (170 mg, 1.4 mmol). The crude product was pur f ed by preparative HPLC (Gradient 1, retention time 13.5 min) to give 3c.TFA as a pale yellow oil (68 mg, 52%). v_max(neat)/cm^-1 3077, 1635, 1598, 483; ¹H MR (500 MHz; CD₃OD) 5 3.85 (2H, t, J = 7.4 Hz, CH₂CH₂N⁺), 5.21 (2H, t, J = 7.4 Hz, CH₂N⁺), 7.57-7.64 (6Η, m, Ph), 7.73 (1Η, app. t, J = 6.4 Hz, 5"-H), 7.83 (1H, d, J = 7.6 Hz, 3"-H), 7.87-7.89 (4H, m, Ph), 8.27 ( H, t, J = 7.6 Hz, 4"-H), 8.68 (1H, d, J = 4.9 Hz, 6"-H), 9.04 (1H, t, J = 1.5 Hz, 4-H), 9.27 (2H, d, J = 1.5 Hz, 2-H, 6-H); ¹³C NMR (125 MHz; CD3OD) 5 37.3, 61.6, 126.0, 127.8, 128.8, 29.5, 130.8, 131.6, 134.7, 142.2, 142.3, 143.2, 44.4, 146.6, 154.5; m/z [HRMS ES+] found [M-TFA]⁺ 337.1708. C₂₄H₂₁N₂ ⁺ requires 337.1705.

3,5-Diphenyl-l-[2-(thiophen-2-yl)ethyl]pyridinium.TFA (3d.TFA). Compound 3d was prepared according to procedure A from thiophene-2-ethylamine (36 mg, 0.28 mmol) and phenylacetaldehyde ( 70 mg, .4 mmol). The crude prod ct was purified by preparative HPLC (Gradient 1, retention time 16.2 min) to give 3d.TFA as a pale yel ow oil (48 mg, 38%). V_max(neaty) cm^-1 3065, 2940, 680, 597, 483; ¹H NMR (500 MHz; CD₃OD) δ 3.67 (2H, t, J = 6.6 Hz, CH₂CH₂N⁺), 5.02 (2H, t, J = 6.6 Hz, CH₂N⁺), 6.87 ( Η, m, 3"-H), 6.96 ( H, dd, J = 5. and 3.5 Hz, 4"-H), 7.33 ( H, dd, J = 5. and 1.1 Hz, 5"-H), 7.56-7.62 (6H, m, Ph), 7.77- 7.79 (4H, m, Ph), 8.98 (1H, t, J = 1.7 Hz, 4-H), 9.04 (2H, d, J = 1.7 Hz, 2-H, 6-H); ¹³C NMR (125 MHz; CD3OD) δ 32.2, 64.4, 26.7, 128.5, 128.7, 128.8, 30.8, 131.6, 134.7, 138.5, 141.9, 142.0, 142.9; m/z [ RMS ES+] found [M-TFA]⁺ 342.1302. C₂₃H₂₀NS⁺ requires 342. 316. l-(2-[3-Bromophenethyl])-3,5-diphenyIpyridinium.TFA (3e.TFA). Compo nd 2e was prepared according to procedure A from 3-bromophenethylamine (100 mg, 0.50 mmol) and phenylacetaldehyde (300 mg, 2.5 mmol). The crude product was purified by preparative HPLC (Gradient 1, retenfon time 17.2 min) to give 3e.TFA as a pale yellow oil (131 mg, 50%). V_max(neaty) cm^-1 3086, 2979, 1685, 1601, 1488; ¾ NMR (500 MHz; CD3OD) δ 3.42 (2H, t, J = 7.2 Hz, CH₂CH₂N⁺), 5.00 (2H, t, J = 7.2 Hz, CH₂N⁺), 7.21-7.27 (2Η, m, 2"-H, 5"-H), 7.42- 7.47 (2H, m, 4"-H, 6"-H), 7.57-7.61 (6H, m, Ph), 7.79 (4H, d, J = 6.3 Hz, 2 x 2'-H, 6'-H), 8.97 (1H, t, J = .7 Hz, 4-H), 9.09 (2H, d, J = 1.7 Hz, 2-H, 6-H); ^,3C NMR (125 MHz; CD₃OD) 6 37.8, 63.9, 124.0, 128.8, 129.1, 130.8, 131.6, 131.8, 131.9, 133.3, 134.7, 139.8, 141.9, 142.0, 143.0; m/z [HRMS ES+] found [M-TFA]⁺ 414.0840. C₂₅H₂₁ ⁷⁹BrN⁺ requires 4 4.0857. l-[2-(3-Nitrophenethyl)]-3,5-diphenyIpyridinium.TFA (3f.TFA). Compound 3f was prepared according to procedu e A from 3-nitrophenethylamine (17 mg, 0.10 mmol) and phenylacetaldehyde (60 mg, 0.5 mmol). The crude product was purified by preparative HPLC (Gradient 1, retention time 16.0 min) to give 3f.TFA as a pale yellow oil (22 mg, 45%). v_max(neaty) cm^"1 3070, 1655, 1598, 561 ; ¹H NMR (500 MHz; CD₃OD) δ 3.59 (2H, t, J = 7.4 z, CH₂CH₂N⁺), 5.04 (2 , t, J = 7.4 Hz, CH₂N⁺), 7.59-7.64 (7Η, m, 5"-H and Ph), 7.71 (1H, d, J = 7.6 Hz, 6' '-Η), 7.83 (4H, dd, 7 = 7.8 and .7 Hz, 2 x 2'-H, 6'-H), 8.16 (1H, d, J = 8.2 Hz, 4"-H), 8.20 (1H, s, 2"-H), 9.04 (1H, t, 7 = .6 Hz, 4-H), 9.21 (2H, d, J = 1.6 Hz, 2-H, 6- H); ¹³C NMR (125 MHz; CD₃OD δ) 37.7, 63.6, 123.6, 25.1, 28.8, 130.8, 31.4, 31.6, 134.7, 136.6, 139.5, 142.0, 142. , 143. , 49.5; m/z [HRMS ES+] found [M-TFA]⁺ 381.1604. C₂5H₂₁N₂02⁺ requi es 38 . 603.

3-(2-Aminoethyl)-4-bromophenol (6g). The reaction was carried out under anhydrous cond ions. To a solution of 2-(2-bromo-5-methoxyphenyl)ethylamine [28] (120 mg, 0.52 mmol) n dichloromethane (20 mL) at -78 °C, boron tribromide (1.3 mL, .3 mmol; 1 M solution in hexane) was added. The mixture was warmed to rt and stirred for 20 h, then cooled to -78 °C and water (50 mL) added dropwise. The aqueous layer was extracted with dich oromethane (3 x 50 mL), filtered and concentrated under reduced pressure to give 6g [29] as a colourless oi (110 mg, 97%). ¹H NMR (600 MHz; D2O) δ 3.09 (2 , t, J = 7.5 Hz, CH₂CH₂N), 3.29 (2H, t, J = 7.5 Hz, CH₂N), 6.77 ( Η, dd, 7 = 8.7 and 3.0 Hz, 6-H), 6.90 (1H, d, 7 = 3.0 Hz, 2-H), 7.52 (1H, d, J = 8.7 Hz, 5-H); ¹³C NMR (150 MHz; D2O) δ 33.9, 39.7, 1 4.4, 117.1, 118.7, 134.6, 37.7, 56.1; m/z [HRMS ES+] found [MH]⁺ 2 6.0030. C₈H₁₁ ⁷⁹BrNO requires 216.0024. l-(2-Bromo-5-hydroxyphenethyl)-3,5-diphenylpyridinium.TFA (3g.TFA). Compound 6g (50 mg, 0.23 mmol) and phenylacetaldehyde (33 mg, 0.28 mmol) in 10 mL of a 1 : 1 mixture of acetonitri e/phosphate buffer (0.1 M solution at pH 6) were stirred at 60 °C for 12 h. The crude product was purified by preparative HPLC (Gradient 1, retention time 6.5 min) and fractions containing the desired product were combined, concentrated and co-evaporated with methanol (3 x 20 mL) to give 3g.TFA (16 mg, 13%) as a pale yellow oil ¹H NMR (600 MHz; CD₃OD) δ 3.51 (2H, t, J = 6.6 Hz, CH₂CH₂N⁺), 5.05 (2H, t, J = 6.6 Hz, CH₂N⁺), 6.66-6.70 (2Η, m, 4"- H, 6"-H), 7.37 (1H, d, J = 8.3 z, 3"-H), 7.57-7.62 (6H, m, Ph), 7.74-7.77 (4H, m, Ph), 8.95 (2H, d, J = .6 Hz, 2-H, 6-H), 9.02 (1H, t, J = 1.6 Hz, 4-H); ¹³C NMR (125 MHz; CD₃OD) δ 38.0, 63.3 114.2, 117.9, 1 9.3, 128.7, 130.8, 13 .6, 134.7, 135.0, 137.4, 142.0, 142.2, 42.9, 159. ; m/z [HRMS EI+] found [M-TFA]⁺ 430.0815. C₂₅H₂₁ ⁷⁹BrNO⁺ requires 430.0807.

l-(3-Hydroxybenzyl)-3,5-diphenylpyridinium.TFA (3h.TFA). Compound 3h was prepared according to proced re A, from 3-aminomethylphenol (62 mg, 0.50 mmol) and phenylacetaldehyde (300 mg, 2.50 mmol). The crude product was purif^'ed by preparat^'ve HPLC (Gradient 1, retention time 14.4 min) to g^'ve 3h.TFA as a pale yellow oil (147 mg, 65%). V_max(neaty) cm^-1 3070, 2927, 1661, 1586, 1482; 1H NMR (500 MHz; CD3OD) δ 5.88 (2H, s, CH₂N⁺ , 6.88 (1Η, d, J = 7.9 Hz, 6"-H , 6.97 (1H, s, 2^*'-H), 7.02 (1H, d, J = 7.9 Hz, 4"-H), 7.27 (1H, t, J = 7.9 Hz, 5"-H), 7.57-7.63 (6H, m, Ph), 7.84-7.88 (4H, m, Ph), 9.04 (1H, s, 4- H), 9.30 (2H, s, 2-H, 6-H); ¹³C NMR (125 MHz; CD₃0

) δ 66.0, 116.4, 17.9, 120.5, 28.8, 30.8, 31.6, 13 .9, 134.7, 136.1, 141.9, 142.3, 143.3, 159.7; m/z [HRMS ES+] found [M- TFA]⁺ 338.1513. C₂₄H₂₀NO⁺ equires 338. 539. l-(2,3-DihydroxypropyI)-3,5-diphenylpyridinium.TFA (3LTFA). Compound 3i was prepared accord^'ng to procedure A, from 3-aminopropane- ,2-diol (45 mg, 0.49 mmol) and phenylacetaldehyde (300 mg, 2.50 mmol). The crude product was purified by preparative HPLC (Gradient 1, retention time 3.0 min) to give 3i.TFA as a pale yellow oil (147 mg, 72%). v_max(neat)/cm ¹ 3292, 3070, 1667, 1597, 1485; *H NMR (500 MHz; CD₃OD) δ 3.60 (1H, dd, J = 11.4 and 6.0 Hz, CHHOH), 3.76 ( H, dd, J = 11.4 and 4.7 Hz, CHHOH), 4. 5-4. 8 ( H, m, CHOH), 4.74 (1H, dd, J = 13.2 and 8.5 Hz, CHHN⁺), 4.97 (1H, dd, J = 13.2 and 3.0 Hz, CHHN⁺), 7.57-7.64 (6Η, m, Ph), 7.87-7.9 (4Η, m, Ph), 9.03 (1Η, t, J= .7 Hz, 4-H), 9.21 (2H, d, J = 1.7 Hz, 2-H, 6-H); ¹³C NMR (125 MHz; CD3OD) δ 64.3, 65.7, 7 .9, 128.8, 130.8, 31.5, 134.9, 141.8, 142.5, 42.9; m/z [HRMS EI] found [M-TFA]⁺ 306.1488. C₂₀H₂₀NO₂ ⁺ requires 306. 489. l-(6-Hydroxyhexyl)-3,5-diphenylpyridinium.TFA (3j.TFA). Compound 3j was prepared according to p ocedure A, from 6-hydroxyhexy amine (59 mg, 0.50 mmol) and phenylacetaldehyde (300 mg, 2.50 mmol). The crude product was purif^'ed by preparative HPLC (Gradient 1, retention time 15.3 min) to give 3j.TFA as a pale yellow oil ( 53 mg, 69%) V. _max(neaty) cm^-1 3070, 2932, 598, 1484; ¹H NMR (500 MHz; CD₃OD) 5 1.49-1.85 (6H, m, 3 x CH₂), 2. 4-2.19 (2Η, m, CH₂), 3.67 (2Η, t, J = 6.3 Hz, CH₂OH), 4.75 (2H, t, J = 7.8 Hz, CH₂N⁺), 7.58-7.65 (6 , m, Ph), 7.88-7.92 (4Η, m, Ph), 9.03 (1Η, t, J = 1.6 Hz, 4-H), 9.29 (2H, d, J = 1.6 Hz, 2-H, 6-H); ¹³C NMR (125 MHz; CD₃OD) δ 26.2, 26.8, 28.9, 32.5, 62.6, 63.4, 28.8, 130.8, 131.6, 134.9, 141.8, 142.0, 143.2; m/z [HRMS ES+] found [M-TFA]⁺ 332.2018. C₂₃H₂6NO⁺ requ^'res 332.2014.

1- (4-Carboxybutyl)-3,5-diphenylpyridinium.TFA (3k.TFA). Compound 3k was prepared according to procedure A, from 4-aminobutyric acid (51 mg, 0.49 mmol) and phenylacetaldehyde (300 mg, 2.50 mmol). The crude product was purified by preparat^'ve HPLC (Gradient 1, retent on time 14.2 min) to give 3k.TFA as a pale yel ow oil ( 34 mg, 63%). v_max(neat)/c ^-1 3200, 2943, 668, 598, 483; ¹H NMR (500 MHz; CD₃OD) δ 2.43 (2H, m, CH₂CH₂C0₂H), 2.56 (2H, t, J = 6.8 Hz, CH₂C0₂H), 4.82 (2H, t, J = 7.3 Hz, CH₂N⁺), 7.56- 7.64 (6Η, m, Ph), 7.88-7.92 (4Η, m, Ph), 9.00 (1Η, t, J = .6 Hz, 4-H), 9.28 (2H, d, J = 1.6 Hz,

2- H, 6-H); ¹³C NMR (125 MHz; CD3OD) δ 27.6, 31.3, 62.7, 128.8, 130.8, 131.5, 134.9, 14 .9, 142.2, 143. , 175.7; m/z [HRMS ES+] found [M-TFA]⁺ 318.1485. C₂₁H₂₀NO₂ ⁺ requ^'res 3 8. 494. l-Cyclohexyl-3,5-diphenylpyridinium.TFA (31.TFA). Compound 31 was prepared according to procedure A from cyclohexylamine (50 mg, 0.50 mmol) and phenylacetaldehyde (300 mg, 2.50 mmol). The crude product was purified by preparative HPLC (Gradient 1, retention time 16.2 min) to g^'ve 31.TFA as a pale yellow oil (1 13 mg, 53%). ¹H NMR (500 MHz; CD3OD) δ .46 (1H, app. qt, J = 3.2 and 3.6 Hz, 4"-Hax), 1.63 (2H, app. q, J = 10.2 Hz, 2 x 2"-Hax), 1.82 ( H, br d, J = 13.2 Hz, 4"-H_eq), 2.05 (2H, br d, J = 3.8 Hz, 2 x 2"-H_eq), 2.18 (2H, qd, J = 2.3 and 3.6 Hz, 2 x 3"-Hax), 2.32 (2H, br d, J = 10.6 Hz, 2 x 3"-H_eq), 4.82 (1H, m, CHN⁺), 7.56-7.64 (6Η, m, Ph), 7.89-7.92 (4Η, m, Ph), 9.00 ( Η, d, J = 1.6 Hz, 4-H), 9.25 (2H, d, J = 1.6 Hz, 2-H, 6-H); ¹³C NMR (125 MHz; CD3OD) δ 25.5, 26.6, 34.3, 74.3, 129.0, 30.8, 131.5, 135.0, 140.5, 142.1, 43.3; m/z (ES+) 314 (M⁺, 100%), 232 (12); m/z [HRMS ES+] found [M- TFA]⁺ 314.1906. C₂₃H₂₄N⁺ requires 314. 909.

3,5-Bis-(4-hydroxyphenyl)-l-(4-hydroxyphenethyI)pyridinium.TFA (3o.TFA).

Compound 3o was prepared according to procedure A, from tyramine.HCl (5.0 mg, 0.028 mmol) and 4-hydroxyphenylacetaldehyde [22] (17 mg, 0.13 mmol). The crude product was purified by preparat^'ve HPLC (Gradient 1, retention time 13.5 min) to give 3o.TFA as a pale ye low oil (9.2 mg, 66%). 1H NMR (600 MHz; CD₃OD) δ 3.28 (2H, t, J = 6.7 Hz, CH₂CH₂N⁺),

4.87 (2H, t, J = 6.7 Hz, CH₂N⁺), 6.73 (2Η, d, J = 8.2 Hz, 3"-H, 5"-H), 6.94-6.98 (6H, m, 2 x (3'-H, 5'-H), 2"-H, 6"-H), 7.57 (d, J = 8.2 Hz, 2 x 2Ή, 6'-H), 8.71 (2H, d, J = 1.6 Hz, 2-H, 6-H), 8.77 ( H, t, J = .6 Hz, 4-H); ¹³C NMR (150 MHz; CD₃OD) δ 37.7, 64.6, 116.9, 1 7.5, 125.6, 127.5, 130.1 , 131.3, 139.2, 139.9, 142.5, 158.3, 161.2; m/z [HRMS ES+] fo nd [M- TFA]⁺ 384.1602. C₂₅H₂₂N0₃ ⁺ requires 384.1600. l-(4-Hydroxyphenethyl)-3,5-di(thiophen-2-yI)pyridinium.TFA (3p.TFA). Compound 2p was prepared according to procedure A, from tyramine.HCl (5.0 mg, 0.028 mmo ) and thiophen-2-yl-acetaldehyde [22] (17 mg, 0.14 mmol). The crude product was purified by preparative HPLC (Gradient 1, retention time 13.5 min) to give 3p.TFA as a pale ye low oil (5.6 mg, 42%). ¹H NMR (600 MHz; CD₃OD) δ 3.29 (2H, t, J = 6.8 Hz, CH₂CH₂N⁺), 4.87 (2H, t, J = 6.8 Hz, CH₂N⁺), 6.73 (2Η, d, J = 8.5 Hz, 3"-H, 5"-H), 6.99 (2H, d, J = 8.5 Hz, 2"-H, 6"-H), 7.27 (2H, dd, J = 4.9 and 3.8 Hz, 4'-H), 7.75-7.78 (4H, m, 2 x 3"-H, 5"-H), 8.79 ( H, t, J = 1.7 Hz, 4-H), 8.83 (2H, d, J = .7 Hz, 2-H, 6-H); ¹³C NMR (150 MHz; CD₃OD) δ 37.5, 64.8, 6.9, 127.3, 129.7, 130.3, 131.2, 131.3, 36.4, 136.7, 136.9, 39.7, 58.3; m/z [HRMS ES+] found [M-TFA]⁺ 364.0820. C₂₁Hi₈NOS₂ ⁺ requires 364.0830. l-(4-Hydroxyphenethyl)-3,5-bis-(4-methoxyphenyl)pyridinium.TFA (3q.TFA).

Compound 3q was prepared according to procedure A, from tyramine.HCl (10 mg, 0.056 mmol) and 4-methoxyphenylacetaldehyde [24] (38 mg, 0.25 mmol). The crude product was purified by preparative HPLC (Gradient 1, retention time 6.2 min) to give 3q.TFA as a pale yellow oil ( 7 mg, 58%). ¹H NMR (600 MHz; CD3OD) δ 3.29 (2H, t, J = 6.7 Hz, CH₂CH₂N⁺),

3.88 (6H, s, 2 x O e), 4.90 (2H, t, J = 6.7 Hz, CH₂N⁺), 6.74 (2Η, d, J = 8.5 Hz, 3"-H, 5"-H), 6.97 (2H, d, J = 8.5 Hz, 2"-H, 6"-H), 7.1 (4H, d, J = 8.7 Hz, 3'-H, 5'-H), 7.68 (4H, d, J = 8.7 Hz, 2'-H, 6'-H), 8.78 (2H, d, J= 1.6 Hz, 2-H, 6-H), 8.83 (1H, t, J = 1.6 Hz, 4-H); ¹³C NMR ( 50 MHz; CD₃OD) δ 37.7, 56.0, 64.7, 116.1, 116.9, 26.9, 27.5, 30.3, 131.3, 39.8, 140.4, 142.3, 158.3, 163.1; m/z [HRMS ES+] found [M-TFA]⁺ 412.1909. C₂7H₂6N0₃ ⁺ requ res 412. 913.

1- (4-Hydroxyphenethyl)-3,5-di(isopropyl)pyridinium.TFA and l-(4-Hydroxyphenethyl)-

2- isobutyl-3,5-di(isopropyl)pyridinium.TFA (3r.TFA and lr.TFA). Compounds 3r and lr were prepared according to procedure A, from tyramine. CI (52 mg, 0.30 mmol) and isovaleraldehyde (120 mg, 1.4 mmol). The c ude products were purified by preparative HPLC (Gradient 1, retention times 3r 15.3 min and lr 16.2 min) to give 3r.TFA (6 mg, 5%) and lr.TFA (13 mg, 0%) as pale yellow oi s. Compo nd 3r: ¹H NMR (500 MHz; CD₃OD) δ 1.28 (12H, d, J = 6.7 Hz, 2 x C (CH₃)₂), 3.05-3.17 (4Η, m, CH(CH₃)₂, CH₂CH₂N⁺), 4.77 (2H, t, J = 6.6 Hz, CH₂N⁺), 6.65 (2H, d, J = 8.4 Hz, 3'-H, 5'-H), 6.79 (2H, d, J = 8.4 z, 2'-H, 6'-H), 8.29 (2H, s, 2-H, 6-H), 8.34 (1H, s, 4-H); ¹³C NMR (125 M z; CD3OD) 6 23.2, 33.2, 37.6,

64.1, 1 6.8, 27.2, 130.9, 14 .7, 43.2, 150.7, 158.2; m/z [HRMS ES+] found [M-TFA]⁺ 284.20 9. Ci₉H₂₆NO⁺ requires 284.2014. Compo nd lr: ¹H NMR (500 MHz; CD₃OD) δ . 2 (6H, d, J = 6.7 Hz, CH(CH₃)₂), .20 (6Η, d, J = 6.9 Hz, CH(CH₃)₂), 1.3 (6Η, d, J = 6.8 Hz, C (CH₃)₂), 1.97-2.03 (1Η, m, CH(CH₃)₂), 2.92 (2H, d, J = 7.5 Hz, 2 x CH₂CH(CH₃)₂), 2.98 (1H, hept, J = 6.9 Hz, CH(CH₃)₂), 3. 5 (2H, t, J = 6.4 Hz, CH₂CH₂N⁺), 3.32-3.36 ( H, m, CH(CH₃)₂), 4.86 (2H, t, J = 6.4 Hz, CH₂N⁺), 6.65 (2Η, d, J = 8.5 Hz, 3'-H, 5'-H), 6.74 (2H, d, / = 8.5 Hz, 2'-H, 6'-H), 8.14 ( H, s, 6-H), 8.30 ( H, s, 4-H); ¹³C NMR ( 25 MHz; CD₃OD) δ

22.2, 23.0, 23.5, 31.0, 31.5, 32.8, 36.8, 37.0, 61.7, 1 6.8, 127.1, 31. , 42.7, 43.3, 147.7, 50.6, 53.7, 158.3; m/z [HRMS ES+] found [M-TFA]⁴ 340.2650. C₂₃H₃₄NO⁺ requires

340.2640.

4. Synthesis of further 1,3,5-trisubstituted pyridinium salts Synthesis of amines

The generation of a new library of pyridinium salts was carried out from a series of substituted pheny ethylamines as well as from aminoalcohols of vary^'ng chain length using the same protoco as that above but a d fferent purification method. Seven of the required amines were commercially available (Scheme 3).

Scheme 3 Commercially available amines

Due to restrictions on the purchase of some phenylethylamine derivatives these were prepared by borane reduction of the corresponding phenylacetonitri e (Table 5). Twelve amine hydrochlorides were prepared in this way. Table 5 Synthesis of substituted phenylethylamines

The commercially available 4-(2-aminoethyl)benzoic acid hydrochloride was used to prepare the corresponding methyl ester 9 (Scheme 4).

Scheme 4 Esterification of 8

Methy 3-(2-aminoethyl)benzoate hydrochloride 12 was prepared via the Henry reaction of methyl 3-formylbenzoate 10 and nitromethane fol owed by reduction of the obtained nitroalkene 11 (Scheme 5). Unexpected y, borane red ction of this nitroalkene 11 also resulted in complete reduct^'on of the ester to the alcohol 13.

Scheme 5 Henry reaction followed by reduction of nitroalkene Synthesis of pyridinium salts

A l^'brary of N-substituted 3,5-d^'pheny pyridinium salts were prepared by the Chichibab^'n- type reaction of phenylacetaldehyde with the amines and amine hydrochlor^'des detailed above.

The final synthesis and pur^'ficat^'on route to the pyridinium salts is outlined in Scheme 6. The pyridinium salts synthes^'sed by this method are outlined in Table 6.

Scheme 6 Synthesis and purification of further pyridinium salts. The pyridinium salts were purified by HPLC. Table 6 Pyrid^'nium salt yields

Testing of pyridinium salts against bacteria

Activity was observed for some of the py dinium chlorides against five bacteria (B. subtilis, S. carnosus, S. aureus, M. luteus and M. aurum). Results are shown in Table 7. Pyridinium salts bearing hydroxyalkyl substituents n general do not display activity (entries A-D) with the exception being substituted hydroxyhexyl pyridinium salt (entry E). This except on may be the result of the increased lipophilicity due to the hexyl chain, which will potentially display the same hydrophobic interactions as those observed with the phenylethylamine derived pyridinium salts. The cyclohexylethy derived pyridini m salt shows activity across all the above bacteria, with particularly good activity for S. aureus, M. luteus and M. a rum (entry F). This suggests that unsaturated ring systems may be tolerated. Bromo- and chloro-substituted phenylethylamine (o, m, or p) derived pyridinium salts all show good activity against the bacteria with the exception of the o-chloro substit ted phenylethylamine pyridinium salt against B. subtilis (entry G-L). o-Methoxy phenylethylamine pyridinium salt had act^'vity across all the above bacteria (entry M). Whereas m- and p-methoxy phenylethylamine pyridinium salts did inhibit the growth of the majority of the bacteria (entries N-O), o- Nitrophenylethylamine pyridinium salt was not suitably purified so was not evaluated against the bacteria, m- and p-nitrophenylethylamine tyridinium salts showed moderate activity against the majority of the bacter^'a (entries Q-R). p-carboxyphenylethylam^'ne pyridinium salt d^'d not show any activity against the above bacteria, presumably due to unfavourable hydrophilic interactions (entry U). The m-methyl ester phenylethylamine derived pyridiniu salts generally had good activity against the bacter^'a, with the exception of B. subtilis (entry W). Conversely the p-methyl ester derived pyridinium salt displayed poor antibacterial activity, only inhibit^'ng the growth of S. aureus and M. ciurunt (entry X). The 3,5 dihydroxyphenylpropyl pyr^'d^'n^'um salt showed limited activity as inhibition was only observed against M. aurum (Table 7, entry Y).

Table 8

Summary of antimicrobial 1,3,5-trisubsituted pyridinium cations

References

1. Pollack, N.; Dolle, C; Ziegler, M. The power to reduce: pyridine nucleotides - small molecules with a multitude of functions. Biochem. J. 2007, 402, 205-2 8.

2. von Datwyler, P.; Ott-Longon^', R.; Schopp, E.; Hesse, M. Ju iprosine, a further alkalo^'d isolated from Prosopis juliflora. Helv. Chim. Acta 1981, 64, 959-1963.

3. Baumgartner, B.; Erde meier, C.A. J.; Wright, A.D.; Rali, T.; Sticher, O. An antimicrob^'al alkaloid from Ficus septica. Phytochemistry, 1990, 2 ·, 3327-3330.

4. Bracher, F.; Daab, J. Total synthesis of the indolizidinium alkaloid fic septine. E ir. J.

Org. Chem. 2002, 2288-2291.

5. Snider, B.B.; Neubert, BJ. Synthesis of ficuseptine, juliprosine, and juliprosopine by b^'omimetic intramolecular chichibabin py idine syntheses. Org. Lett. 2005, 7, 27 5-2718.

6. Arora, P.K.; Riachi, N.J.; Fiedler, G.C.; Singh, M.P.; Abdallah, F.; Harik, S.I.; Sayre, L.M.

Struct re-neurotoxicity trends of analogues of -methyl-4-phenylpyridinium (MPP⁺) the cytotoxic metabolite of the dopaminergic neurotoxin MPTP. Life Sei. 1990, 46, 379-390.

7. Wimalasena, D.S.; Perera, R.P.; Heyen, B.J.; Balasooriya, I.S.; Wimalasena, K. Vesicula Monoamine Transpo ter Substrate/Inhibitor Activity of MPTP/MPP+ Derivatives: A structure-activity study. J. Med. Chem. 2008, 5 , 760-768.

8. Pijper, D.; Bulten, E.; Smisterova, J.; Wagenaar, A.; Hoekstra, D.; Engberts, J.B.F.N.;

Hulst, R. Novel b odegragable pyridinium amphiphiles for gene delivery. Eur. J. Org. Chem. 2003, 4406-4412.

9. Trova, M.P.; Wissner, A.; Carroll, M.L.; Kerwar, S.S.; Pickett, W.C.; Schaub, R.E.;

Tor ey, L.W.; Kohler, C.A. Analogues of platelet activating factor. 8. Antagonists of PAF containing an aromatic ring I nked to a pyr^'dinium r^'ng. J. Med. Chem. 1993, 36, 580-590.

10. Alptuziin, V.; Parlar, S.; Tash, H.; Erciyas, E. Synthesis and antimicrobial activity of some pyridinium salts. Molecules 2009, 14, 5203-5215.

11. Ch^'ch babin, A.E. Uber condensationen der aldehyde mit ammonok zu pyridinbasen. J.

Prakt. Chem. 1924, 107, 22-128.

12. Frank, R.L.; Seven, R.P. Pyridines. IV. A study of the Chich^'babin synthesis. /. Am. Chem.

Soc. 1949, 71, 2629-2635.

13. Eliel, E.L.; McBride, R.T.; Kaufmann, S. Abnormal Chichibabin reactions. The condensation of phenylacetaldehyde and homoveratric aldehyde with ammonia. J. Am. Chem. Soc. 1953, 75, 4291-4296.

4. Farley, CP.; Eliel, E.L. Chich^'babin reactions with phenylacetaldehyde. J. Am. Chem. Soc.

1956, 78, 3477-3484.

15. Charman, H.B.; Rowe, J.M. Condensation of aldehydes w^'th ammoni m salts to give substituted pyridines. Chem. Commun. 1971, 476—477.

6. Suyama, K.; Adachi, S. Reaction of alkanals and amino acids or primary amines. Synthesis of 1,2,3,5- and 1,3,4,5-substituted quaternary pyridinium salts. J. Org. Chem. 1979, 44, 417-1420.

17. Ma, T.; Zhang, S.; Li, Y.; Yang, F.; Gong, C; Zhao, J. Synthesis and characterisation of soluble polyimides based on a new fluorinated diamine: 4-Phenyl-2,6-bis[3-(4'amino- 2'trifluoromethyl-phenoxy)phenyl] pyridine. J. Fluorine Chem. 2010, 131, 724-730. 18. Yu, L.-B.; Chen, D.; L^', J.; Ramirez, J.; Wang, P. G. Lanthanide-promoted reactions of aldehydes and amine hydrochlorides in aqueous solution. Synthesis of 2,3- dihydropyridinium and pyridini m derivatives. J. Org. Chem. 1997, 62, 208-211.

9. Burns, N.Z.; Baran, P.S. On the origin of the haouamine alkaloids. Ange . Chem. Int. Ed.

2008, 47, 205-208.

20. Burns, N.Z.; Jessing, M.; Baran, P.S. Total synthesis of hao amine A: the indeno- tetrahydropyr^'dine core. Tetrahedron 2009, 65, 6600-6610.

21. Dagorn, F.; Yan, L.-H.; Gravel, E.; Leblanc, K.; Mac uk, A.; Poupon, E. Particular behavior of 'C₆C₂ un^'ts' in the Chichibabin pyrid^'ne synthesis and biosynthetic implications. Tetrahedron Lett. 2011, 52, 3523-3526.

22. Pesnot, T.; Gershater, M.C.; Ward, J. .; Hailes, H.C. Phosphate ediated b^'omimetic synthes^'s of tetrahydroisoquinolines. Chem. Commun. 2011, 47, 3242-3244.

23. L ehman, B.R.; Lamming, E.D.; Pesnot, T.; Smith, J.M.; Hailes, H.C; Ward, J.M. One- pot triangular chemoenzymatic cascades for the syntheses of chiral alkaloids from dopamine. Green Chem. 2015, 17, 852-855.

24. Pesnot, T.; Gershater, M.C.; Ward, J.M.; Hailes, H.C. The Catalytic Potential of Coptis japonica NCS2 Revealed - Development and Uti isation of a Fluorescamine-Based Assay. Adv. Syn. Cat. 2012, 354, 2997-3008.

25. Davril, M.; Han, K.-K. Isolation and characterization of a highly cross- inked peptide from elastin of porcine aorta. FEBS Lett. 1974, 43, 33 -336.

26. Umeda, H.; Takeuchi, M.; Suyama, K. Two new elastin cross-I nks hav^'ng pyridine skeleton. J. Biol. Chem. 2001, 276, 12579- 2587.

27. Pereira, A.M.; Abreu, A.C.; Simoes, M. Action of kanamycin against s ngle and dual species biof ^'lms of Escherichia coli and Staphylococcus a reus. J. Microbiol. Res. 2012, 2, 84-88.

28. Liang, J.T. ; Liu, J. ; Sh^'reman, B.T. ; Tran, V. ; Deng, X. ; Mani, N.S. A practical synthesis of regioisomeric 6- and 7-methoxytetrahydro-3-benzazepines. Org. Process Res. De . 2010, 14, 380-385.

29. Chao, H.J.; Tuerd^", R.T.; Nerpin, T.; Roberge, J.Y.; Liu, Y.; Lawrence, R.M.; Rehfuss, R.P.; Clark, C.G.; Q^'ao, J.X.; Gungor, T.; Lam, P.Y.S.; Wang, T.C.; Ruel, R.; L'Heureux, A.L.; Thibeault, C; : outhillier, G.; Schnur, D.M. PCT Int. Appl. 2005, WO 200511351 .

Claims

Claims:

1. A 1,3,5-trisubstituted pyridinium salt of general formula I

wherein X"- is a counterion in which n is an integer from 1 to 4;

wherein R² and R³ are each independently a substituent group comprising Ar, wherein Ar is an aryl or heteroaryi group, which is substituted or unsubstituted;

wherein R¹ is a substituent group linked to the quartemary nitrogen of the pyridinium ring by an aliphatic carbon; and

wherein, when R¹ is benzyl, R² and R³ are not both phenyl, 3-MeO phenyl, 4-Br phenyl, 3-CF₃ phenyl, 2-Me phenyl or 4-(4-tert BuPh) phenyl;

when R¹ is 3-MeO phenethyl, R² and R³ are not both 3-MeO phenyl; and

when R¹ is dimethoxyphenethyl , R² and R³ are not both phenyl.

2. A 1,3,5-trisubstituted pyridinium salt according to claim 1, wherein R¹ comprises (i) Ar¹, wherein Ar¹ is an aryl or heteroaryi group which is substituted or unsubstituted; or (ii) an aliphatic group comprising at least 6 carbon atoms.

3. A 1,3,5-trisubstituted pyridinium salt according to claim 2, wherein R^! comprises Ar¹ linked to the quartemary nitrogen of the pyridinium ring by an aliphatic carbon.

4. A 1 ,3,5-trisubstituted pyridinium salt according to claim 3, wherein the aliphatic carbon is from a one or two carbon linker between Ar¹ and the quartemary nitrogen of the pyridinium ring.

A 1 ,3,

5-trisubstituted pyridinium salt according to any one of claims 2 to 4, wherein monosubstituted or disubstituted.

6. A 1,3,5-trisubstituted pyridinium salt according to claim 5, wherein Ar¹ bears one or two substituents selected from nitro, methoxy, hydroxyl and halogen.

7. A 1,3,5-trisubstituted pyridinium salt according to claim 6, wherein Ar¹ bears a chloro or bromo substituent.

8. A 1 ,3,5-trisubstituted pyridinium salt according to any one of claims 2 to 7 wherein Ar³ is phenyl.

9. A 1,3,5- tri substituted pyridinium salt according to any one of claims 2 to 4, wherein Ar ¹ is thiophen-2-yl.

10. A 1 ,3,5- trisubstituted pyridinium salt according to claim 2, wherein R¹ comprises hexyl or cyclohexyl.

11. A 1 ,3,5-trisubstituted pyridinium salt according to claim 10, wherein R¹ comprises cyclohexyl linked to the quaternary nitrogen of the pyridinium ring by a one or two carbon linker.

12. A 1,3,5-trisubstituted pyridinium salt according to any preceding claim, wherein R¹ is adsorbed to, attached to or forms part of a surface.

13. A 1,3,5-trisubstituted pyridinium salt according to claim 12, wherein the surface is a medical device surface, a surface for food use, a hospital surface, a sanitary ware surface or a marine surface.

14. A 1 ,3,5-trisubstituted pyridinium salt according to any preceding claim, wherein R² and R³ are each independently a substituent group which consists of Ar with no more than one aromatic ring.

15. A 1,3,5-trisubstituted pyridinium salt according to any preceding claim, wherein R² and R³ are the same.

16. A 1 ,3,5-trisubstituted pyridinium salt according to any preceding claim, wherein Ar is phenyl.

17. A 1 ,3,5-trisubstituted pyridinium salt according to claim 16, wherein Ar is unsubstituted phenyl, 4-hydroxyphenyl or 4-methoxy phenyl.

18. A 1,3,5- trisubstituted pyridinium salt according to any one of claims 1 to 15, wherein Ar is thiophen-2-yl.

19. A 1 ,3,5-trisubstituted pyridinium salt according to any preceding claim, wherein the counterion is selected from CI-, Br, CF3CO2^', CF3CO2 , SO4^2-, P0₄ ^3-, and CO3^2-.

20. A 1,3,5- trisubstituted pyridinium salt according to claim 1, in which the pyridinium cation is selected from:

21. Use of a 1,3,5-trisubstituted pyridinium salt of general formula I as an antimicrobial,

wherein X"^~ is a counterion in which n is an integer from 1 to 4;

wherein R² and R³ are each independently a substituent group comprising Ar, wherein Ar is an aryl or heteroaryl group, which is substituted or unsubstituted;

wherein R¹ is a substituent group which comprises (i) Ar¹, wherein Ar¹ is an aryl or heteroaryl group which is substituted or unsubstituted; or (ii) an aliphatic group comprising at least 6 carbon atoms, wherein the substituent group is linked to the quartemary nitrogen of the pyridinium ring by an aliphatic carbon; and

wherein R¹ is not 4-hydroxyphenethyl or 4-carboxyphenethyl when R² and R³ are both phenyl; and

wherein R¹ is not 3-hydroxyphenethyl or 3-hydroxyphenyl 1 -hydroxy prop-2-yl when R² and R³ are both 3,4-dimethoxyphenyl.

22. Use according to claim 20, wherein R¹ comprises (i) Ar¹, wherein Ar¹ is an aryl or heteroaryl group which is substituted or unsubstituted; or (ii) an aliphatic group comprising at least 6 carbon atoms.

23. Use according to claim 22, wherein R¹ comprises Ar¹ linked to the quartemary nitrogen of the pyridinium ring by an aliphatic carbon.

24. Use according to claim 23, wherein the aliphatic carbon is from a one or two carbon linker between Ar¹ and the quartemary nitrogen of the pyridinium ring.

25. Use according to any one of claims 22 to 24, wherein Ar¹ is monosubstituted or disubstituted.

26. Use according to claim 25, wherein Ar¹ bears one or two substituents selected from nitro, methoxy, hydroxyl and halogen.

27. Use according to claim 26, wherein Ar¹ bears a chloro or bromo substituent.

28. Use according to any one of claims 22 to 27 wherein Ar¹ is phenyl.

29. Use according to any one of claims 22 to 24, wherein Ar ¹ is thiophen-2-yl.

30. Use according to claim 22, wherein R¹ comprises hexyl or cyclohexyl.

31. Use according to claim 30, wherein R^l comprises cyclohexyl linked to the quaternary nitrogen of the pyridinium ring by a one or two carbon linker.

32. Use according to any one of claims 21 to 31, wherein R¹ is attached to or forms part of a surface.

33. Use according to claim 32, wherein the surface is a medical device surface, a surface for food use, a hospital surface, a sanitary ware surface, or a marine surface.

34. Use according to any one of claims 21 to 33, wherein R² and R³ are each independently a substituent group which consists of Ar with no more than one aromatic ring.

35. Use according to any one of claims 31 to 34, wherein R² and R³ are the same.

36. Use according to any one of claims 21 to 35, wherein Ar is phenyl.

37. Use according to claim 36, wherein Ar is unsubstituted phenyl, 4-hydroxyphenyl or 4- methoxy phenyl.

38. Use according to any one of claims 21 to 35, wherein Ar is thiophen-2-yl.

39. Use according to any one of claims 21 to 38, wherein the counterion is selected from Cl\ Br, CF3CO2 , CF3CO2-, S0₄ ²-, PO4³-, and CO3² .

40. Use according to claim 21, in which the pyridinium cation is as defined in claim 20.

41. Use according to any one of claims 21 to 40, wherein the antimicrobial is an antibacterial.

42. Use according to claim 41, wherein the antibacterial is effective against Staphylococcus, Bacillus, Mycobacterium and/or Micrococcus.

43. A 1,3,5-trisubstituted pyridinium salt as defined in any one of claims 21 to 40, for use as a medicament.

44. A process for the production of a 1,3,5-trisubstituted pyridinium salt, which comprises reacting an aryl or heteroaryl acetaldehyde with an amine in the presence of a phosphate catalyst and water, so as to form the 1,3,5-trisubstituted pyridinium salt.

45. A process according to claim 44, wherein the molar ratio of aryl or heteroaryl acetaldehyde to amine is in the range 4 to 6.

46. A process according to claim 44 or claim 45, wherein the concentration of phosphate is preferably in the range 0.2M to 0.3M.

47. A process according to anyone of claims 44 to 46, wherein the temperature is at least 60°C.

48. A process according to claim 47, wherein the temperature is around 100°C.

49. A process according to any one of claims 44 to 48, wherein the pH is around 6.

50. A process according to any one of claims 44 to 49, wherein the reaction time is at least 10 hr.

51. A process according to any one of claims 44 to 50, which is carried out in the presence of water and a water-miscible cosolvent.

52. A process according to claim 51 , wherein the volume ratio of water to cosolvent is 1.

53. A process according to any one of claims 44 to 52, wherein the phosphate catalyst is inorganic phosphate or pyrophosphate, a sugar phosphate or a nucleobase monophosphate.

54. A process according to anyone of claims 44 to 53, wherein the amine has the formula R¹-NH₂, wherein R¹ is a substituent group linked to the nitrogen by an aliphatic carbon.

55. A process according to claim 54, wherein R¹ is as defined in any one of claims 21 to 34.

56. A process according to any one of claims 44 to 55, wherein the aryl or heteroaryl acetaldehyde has the formula R²-CH₂-CHO or is a mixture of compounds of the formulae R²- CH2-CHO and R³-CH₂-CHO, wherein R² and R³ are each independently a substituent group comprising Ar, wherein Ar is an aryl or heteroaryl group, which is substituted or unsubstituted.

57. A process according to claim 56, wherein R² and R³ are as defined in any one of claims 35 to 38.