Olefinic quaternary ammonium compounds

Abstract

The invention relates to olefinic quaternary ammonium (OQA) compounds of Formula (I), methods of their synthesis and materials comprising them.

Description

[0001] OLEFINIC QUATERNARY AMMONIUM COMPOUNDS

[0002] 1. FIELD OF THE INVENTION

[0003] The invention relates generally to olefinic quaternary ammonium compounds, methods of their synthesis and materials comprising them.

[0004] 2. BACKGROUND TO THE INVENTION

[0005] Microbial infection is a huge challenge to global human health. Pathogenic microorganisms are a particular problem in hospitals and other healthcare settings, adversely affecting the functioning of medical devices, drugs, bone and dental grafts and surgical equipment. In addition, the widespread misuse of antibiotics and disinfectants has contributed to the emergence of antibiotic-resistant bacteria that pose a serious threat to human health. The World Health Organisation has appealed to researchers worldwide to prioritize efforts in combating the spread of such bacteria.

[0006] Quaternary ammonium compounds (QACs) are some of the most commonly used antimicrobial compounds. QACs comprise a positively charged nitrogen atom bound to four other groups, balanced by a negatively charged anion. Most commonly, the four other groups are hydrocarbon groups, often where at least one hydrocarbon group is longer than the others.

[0007] QACs such as benzalkonium chloride (BAC) and dimethyldidecylammonium chloride (DDAC) are biocides regularly found in disinfectant and antiseptic formulations in homes, healthcare facilities, animal welfare facilities and industry.

[0008] Without being bound by theory, it is believed that the major driving force for QAC activity originates from the interaction of the cationic group with the lipophilic bacterial cell walls / components. Subsequent insertion of hydrophobic segments of the QAC into the lipids of bacterial cytoplasmic membrane leads to the leakage of vital intracellular components and concomitant cell death.

[0009] Quaternary ammonium groups have also been incorporated into larger structures to confer antibiotic properties on those materials. Synthetic polymers with quaternary ammonium functionality can be produced by quaternization of reaction polymers (postpolymerisation) or by polymerization of monomers containing quaternary ammonium moieties. For example, quaternary ammonium groups have been added to polymers such as chitosan and peptide antibiotics, to enhance properties such as solubility and antimicrobial activity. Rusli et. al. describe the synthesis of a series of homo and copolymers containing quaternary ammonium groups, which were found to be active against strains of P. aeruginosa, S. aureus and C. albicans. (Rusli, Periayah, Loh, Lakshminar, & Parthiban, 2023).

[0010] Quaternary ammonium groups have also been bound to surfaces as antimicrobial treatments, such as to textiles, to medical instruments and to high-touch surfaces in public spaces.

[0011] Although QACs and quaternary ammonium-functionalised materials provide useful biocidal tools, there are still hurdles and disadvantages. For example, despite QACs being ubiquitous in society today, their chemical synthesis has remained largely the same since first described in the mid-19th century; namely quaternisation of amines by alkylation. QACs are primarily synthesised by monoalkylation of tertiary amines, (the Menshutkin reaction) or via exhaustive alkylation of ammonia, primary or secondary amines. These reactions are often conducted at elevated temperatures (60-100°C), which can increase the susceptibility of QAC to degradation. Additionally, alkylating agents used such as alkyl halides and alkyl sulfates are acutely toxic, potentially carcinogenic, and a concern with respect to environmental contamination.

[0012] A further hurdle is that although QACs are considered "biodegradable" they are often cytotoxic and may persist in the environment. As well as presenting potential health and environmental risks, incomplete degradation may lead to microenvironments containing sub-inhibitory concentrations of QACs, which can generate antimicrobial resistance.

[0013] It is therefore an object of the invention to go at least partway to alleviating some of the problems associated with QACs and the synthesis of quaternary ammonium- functionalised antimicrobial polymers and biomaterials and / or to at least provide the public with a useful choice.

[0014] In this specification where reference has been made to patent specifications, other external documents, or other sources of information, this is generally for the purpose of providing a context for discussing the features of the invention. Unless specifically stated otherwise, reference to such external documents is not to be construed as an admission that such documents, or such sources of information, in any jurisdiction, are prior art, or form part of the common general knowledge in the art. 3. SUMMARY OF THE INVENTION

[0015] In one aspect the invention provides a compound of Formula I as depicted below. Compounds of Formula I are olefinic quaternary ammonium (OQA) compounds which may have similar properties to QACs used in medicine and industry.

[0016] The invention also provides a compound of Formula IA and processes for preparing compounds of Formula I and IA.

[0017] In one aspect the invention provides an antimicrobial formulation comprising a compound of Formula I or IA and excipients.

[0018] In one aspect the invention provides a compound of Formula I or IA which is bound to the surface of a material via a trioxysilane.

[0019] In one aspect the invention provides a biomaterial comprising one or more compounds of Formula I or IA.

[0020] 4. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] The invention will now be described by way of example only and with reference to the drawings in which:

[0022] Figure 1 shows the preparation of quaternary ammonium modified allenamide-LYRAK 15 by synthesis of LYRAK using standard Fmoc-SPPS protocol, installation of the allenamide handle on-resin using protocol by Cameron et al. (Cameron, Harris, & Brimble, 2020) and aza-Michael quaternisation step either a) on-resin, or b) in solutionphase on the liberated peptide.

[0023] Figure 2 shows Scheme 1 outlining one embodiment of Process B where an allenamide intermediate compound reacts with triethylamine (TEA) to form the OQA compounds of the invention 2a and 2a*.

[0024] 5. DETAILED DESCRIPTION OF THE INVENTION

[0025] 5.1 Definitions and abbreviations

[0026] As used herein the term "comprising" means "consisting at least in part of". When interpreting each statement in this specification that includes the term "comprising", features other than that or those prefaced by the term may also be present. Related terms such as "comprise" and "comprises" are to be interpreted in the same manner.

[0027] The term "about" as used herein means a reasonable amount of deviation of the modified term such that the end result is not significantly changed. For example, when applied to a value, the term should be construed as including a deviation of+ / - 5% of the value. As used herein the term "and / or" means "and" or "or", or both. As used herein "(s)" following a noun means the plural and / or singular forms of the noun.

[0028] Asymmetric centers may exist in the compounds described herein. The asymmetric centers may be designated as (R) or (S), depending on the configuration of substituents in three-dimensional space at the chiral carbon atom. All stereochemical isomeric forms of the compounds, including diastereomeric, enantiomeric, and epimeric forms, as well as d-isomers and l-isomers, and mixtures thereof, including enantiomerically enriched and diastereomerically enriched mixtures of stereochemical isomers, are within the scope of the invention.

[0029] Individual enantiomers can be prepared synthetically from commercially available enantiopure starting materials or by preparing enantiomeric mixtures and resolving the mixture into individual enantiomers. Resolution methods include conversion of the enantiomeric mixture into a mixture of diastereomers and separation of the diastereomers by, for example, recrystallization or chromatography, and any other appropriate methods known in the art. Starting materials of defined stereochemistry may be commercially available or made and, if necessary, resolved by techniques well known in the art.

[0030] The compounds described herein may also exist as conformational or geometric isomers, including cis, trans, syn, anti, entgegen (5), and zusammen (Z) isomers. All such isomers and any mixtures thereof are within the scope of the invention.

[0031] Also within the scope of the invention are any tautomeric isomers or mixtures thereof of the compounds described. As would be appreciated by those skilled in the art, a wide variety of functional groups and other structures may exhibit tautomerism. Examples include, but are not limited to, keto / enol, imine / enamine, and thioketone / enethiol tautomerism.

[0032] The compounds described herein may also exist as isotopologues and isotopomers, wherein one or more atoms in the compounds are replaced with different isotopes. Suitable isotopes include, for example,1H,2H (D),3H (T),12C,13C,14C,16O, and18O. Procedures for incorporating such isotopes into the compounds described herein will be apparent to those skilled in the art. Isotopologues and isotopomers of the compounds described herein are also within the scope of the invention.

[0033] Also within the scope of the invention are salts of the compounds described herein, including pharmaceutically acceptable salts. Such salts include acid addition salts and base addition salts. Acid addition salts can be prepared by reacting compounds, in free base form, with inorganic or organic acids. Examples of inorganic acids include, but are not limited to, hydrochloric, hydrobromic, nitric, sulfuric, and phosphoric acid. Examples of organic acids include, but are not limited to, acetic, trifluoroacetic, propionic, succinic, glycolic, lactic, malic, tartaric, citric, ascorbic, maleic, fumaric, pyruvic, aspartic, glutamic, stearic, salicylic, methanesulfonic, benzenesulfonic, isethionic, sulfanilic, adipic, butyric, and pivalic. Base addition salts can be prepared by reacting compounds, in free acid form, with inorganic or organic bases. Examples of inorganic base addition salts include alkali metal salts, alkaline earth metal salts, and other physiologically acceptable metal salts, for example, aluminium, calcium, lithium, magnesium, potassium, sodium, or zinc salts. Examples of organic base addition salts include amine salts, for example, salts of trimethylamine, diethylamine, ethanolamine, diethanolamine, and ethylenediamine.

[0034] The compounds described herein may form or exist as solvates with various solvents. If the solvent is water, the solvate may be referred to as a hydrate, for example, a monohydrate, a di- hydrate, or a tri-hydrate. All solvated forms and unsolvated forms of the compounds described herein are within the scope of the invention.

[0035] The general chemical terms used herein have their usual meanings.

[0036] The term "alkyl" as used herein refers to a straight-chain or branched saturated acyclic hydrocarbon group having up to 30 carbon atoms and includes any C1-C26, C1-C22, Ci- Cis, C1-C10, or C1-C7 alkyl group.

[0037] The term "alkenyl" refers to a straight-chain or branched acyclic hydrocarbon group having at least one double bond, and having up to 30 carbon atoms, and includes any C2-C26, C2-C22, C2-C18, C2-C10, or C2-C7 alkenyl group.

[0038] The term "alkynyl" refers to a straight-chain or branched acyclic hydrocarbon group having at least one triple bond, and having up to 30 carbon atoms, and includes any C2- C26, C2-C22, C2-C18, C2-C10, or C2-C7 alkynyl group.

[0039] The prefix "Cx-Cy", wherein x and y are each an integer, when used in combination a carbon-containing substituent, refers to the number of carbon atoms in the substituent.

[0040] The term "halo" refers to F, Cl, Br and I. The term "thiol" refers to -SH. The term "thio" when used to prefix a functional group, means that a sulfur atom is present or that the functional group is the sulfur equivalent of an oxygen-containing functional group.

[0041] The term "carbocycle" as used herein refers to a saturated or unsaturated non-aromatic mono-, bi- or tricyclic alkyl group. In some embodiments, carbocycle groups have from 3 to 12, from 3 to 10, from 3 to 8, from 3 to 6, from 3 to 5 carbon atoms in the ring(s). In some embodiments, carbocycle groups have 5 or 6 ring carbon atoms. Examples of monocyclic carbocycle groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

[0042] The term "aryl" as used herein refers to a cyclic aromatic hydrocarbon group that does not contain any ring heteroatoms. Aryl groups include monocyclic, bicyclic and tricyclic ring systems. Examples of aryl groups include, but are not limited to, phenyl, azulenyl, heptalenyl, biphenyl, fluorenyl, phenanthrenyl, anthracenyl, indenyl, indanyl, pentalenyl, and naphthyl. In some embodiments, aryl groups have from 6 to 20, 6 to 14, 6 to 12, or 6 to 10 carbon atoms in the ring(s). In some embodiments, the aryl groups are phenyl or naphthyl. Aryl groups include aromatic-carbocycle fused ring systems such as indanyl and tetra hydronaphthyl.

[0043] The term "heterocycle" as used herein refers to a saturated or unsaturated non-aromatic ring system containing 3 or more ring atoms, of which one or more is a heteroatom. In some embodiments, the heteroatom is nitrogen, oxygen, or sulfur. In some embodiments, the heterocycle group contains one, two, three, or four heteroatoms. Heterocycle groups include partially unsaturated and saturated ring systems, for example, imidazolinyl and imidazolidinyl. Heterocycle groups include fused and bridged ring systems containing a heteroatom, for example, quinuclidyl.

[0044] The term "heteroaryl" as used herein refers to an aromatic ring system containing 5 or more ring atoms, of which, one or more is a heteroatom. In some embodiments, the heteroatom is nitrogen, oxygen, or sulfur. Heteroaryl groups include, but are not limited to, pyrrolyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, thiophenyl, benzothiophenyl, furanyl, benzofuranyl, indolyl, azaindolyl (pyrrolopyridinyl), indazolyl, benzimidazolyl, pyrazolopyridinyl, triazolopyridinyl, benzotriazolyl, benzoxazolyl, benzothiazolyl, imidazopyridinyl, isoxazolopyridinylxanthinyl, guaninyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, quinoxalinyl, and quinazolinyl. Heteroaryl groups include fused ring systems in which all of the rings are aromatic, for example, indolyl, and fused ring systems in which only one of the rings is aromatic, for example, 2,3-dihydroindolyl.

[0045] The term "heteroatom" is intended to include oxygen, nitrogen, sulfur, selenium, or phosphorus. In some embodiments, the heteroatom is selected from the group consisting of oxygen, nitrogen, and sulfur.

[0046] In some embodiments, the functional groups described above are substituted with other groups. As used herein, the term "substituted" is intended to mean that one or more hydrogen atoms in the group indicated is replaced with one or more independently selected suitable substituents, provided that the normal valency of each atom to which the substituent / s are attached is not exceeded, and that the substitution results in a stable compound. The term "stable" as used herein, unless indicated otherwise, refers to compounds which possess stability sufficient to allow manufacture and which maintain their integrity for a period of time sufficient to be useful for the purposes described herein.

[0047] The term "amino acid" as used herein, refers to a compound comprising both amino and carboxylic acid functional groups, and includes a, 0, y and 6 amino acids. The term "amino acid derivative" as used herein, refers to amino esters (amino acid esters) and amino amides (amino acid amides).

[0048] The term "peptide" as used herein, refers to a compound comprising two or more amino acids joined by peptide bonds. The C-terminus of a peptide may comprise an amino acid derivative, for example an amino ester or amino amide (also known as a carboxamide when present at the C-terminus of a peptide, eg -CONH2 or CONHR). Peptides may be defined using either the standard 3 letter or one letter code for amino acids.

[0049] It is intended that reference to a range of numbers disclosed herein (for example, 1 to 10) also incorporates reference to all rational numbers within that range (for example, 1, 1.1, 2, 3, 3.9, 4, 5, 6, 6.5, 7, 8, 9 and 10) and also any range of rational numbers within that range (for example, 2 to 8, 1.5 to 5.5 and 3.1 to 4.7) and, therefore, all sub-ranges of all ranges expressly disclosed herein are hereby expressly disclosed. These are only examples of what is specifically intended and all possible combinations of numerical values between the lowest value and the highest value enumerated are to be considered to be expressly stated in this application in a similar manner.

[0050] Whenever a range is given in the specification, for example, a temperature range, a time range, or a composition range, all intermediate ranges and subranges, as well as all individual values included in the ranges given are intended to be included in the disclosure. In the disclosure and the claims, "and / or" means additionally or alternatively. Moreover, any use of a term in the singular also encompasses plural forms.

[0051] 5.2 The olefinic ammonium compounds of the invention

[0052] The invention provides novel olefinic quaternary ammonium (OQA) compounds with a range of applications.

[0053] In one aspect the invention provides a compound of Formula I

[0054] wherein R and R’ are independently selected from H, -(C1-12)-alkyl, -(C2-12)-alkenyl, -(C2-12)-alkynyl (which latter three groups are optionally substituted with one or more substituents selected from the group consisting of halo, nitro, -N3, =O, -CN, -OR10a, - SR10b, -COR10c, -COOR10d, -NR10eR10f, -CONR10gR10h, carbocyclea1, aryla1, heterocyclea1, heteroaryla1, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), carbocycleb1, arylb1, heterocycleb1, heteroarylb1(which four groups are optionally substituted with one or more substituents selected from halo, nitro, -N3, -CN, -OR11a, - SR11b, -COR11c, -COOR11d, -NR11eR11f, -CONR11gR11h, carbocyclea2, aryla2, heterocyclea2, heteroaryla2, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), or wherein R and R’, together with the N atom to which they are attached, represent a 3-10-membered heterocyclic or heteroaryl ring which may additionally contain one or more heteroatoms selected from O, S and N (which heterocyclic or heteroaryl ring is optionally substituted by one or more substituents selected from halo, nitro, -N3, -CN, - OR12a,-SR12b, -COR12c, -COOR12d, -NR12eR12f, -CONR12gR12h, carbocyclea3, aryla3, heterocyclea3, heteroaryla3, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), or wherein R is H and -NHR’ represents an amino acid or amino acid derivative or a peptide, or wherein -NRR’ represents an amine-functionalised polymer or material; wherein R1, R2and R3are independently selected from -(C1-14)-alkyl or-(C1-14)carbocycle or R1and R2together with the N atom to which they are attached, form a 3-14- membered heterocyclic or heteroaryl ring which may additionally contain one or more heteroatoms selected from O, S and N (which heterocyclic or heteroaryl ring is optionally substituted with one to three methyl groups); and wherein -(C1-14)-alkyl may be optionally substituted with aryl10; wherein R4, R5and R6are independently selected from H, halo, nitro, -N3, -CN, -OR13a, - SR13b, -COR13c, -COOR13d, -NR13eR13f, -CONR13gR13h, -(C1-6)-alkyl, -(C2-6)-alkenyl, -(C2-6)- alkynyl (which latter three groups are optionally substituted with or more substituents selected from the group consisting of halo, nitro, =O, -N3, CN, -OR14a,-SR14b, -COR14c, - COOR14d, -NR14eR14f, -CONR14gR14h, carbocyclea4, aryla4, heterocyclea4, heteroaryla4, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), carbocycleb2, arylb2,heterocycleb2and heteroarylb2(which four groups are optionally substituted with one or more substituents selected from halo, nitro, -N3, -CN, -OR15a,- SR15b, -COR15c, -COOR15d, -NR15eR15f, -CONR15gR15h, carbocyclea5, aryla5, heterocyclea5, heteroaryla5-(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl); wherein each carbocyclea, aryla, heterocycleaand heteroarylais independently selected from a 4-14-membered ring as earlier defined, optionally substituted with one or more substituents selected from halo, nitro, =O, -N3,-CN, -OR16a,-SR16b, -COR16c, -COOR16d, - NR16eR16f, -CONR16gR16h; wherein each carbocycleb, arylb, heterocycleband heteroarylbis independently selected from a 4 to 14-membered ring as earlier defined; wherein each R10-16ato R10-16his independently selected from H, -(C1-10)-alkyl, -(C2-10)- alkenyl, -(C2-10)-alkynyl (which latter three groups are optionally substituted with one or more substituents selected from the group consisting of halo, nitro, -N3, -CN, =O, -OR17a, -SR17b, -COR17c, -COOR17d, -NR17eR17f, -CONR17gR17h, carbocycleb3, arylb3, heterocycleb3, heteroarylb3, -(C1-6)-alkyl, -(C2-6)-alkenyl, -(C2-6)-alkynyl and -(C3-6)-cycloalkyl); or the pairs R10-16ewith R10-16fand / or R10-16gwith R10-16hmay form, together with the N atom to which they are attached, a 3-14 membered heterocyclic or heteroaryl ring which may contain one or more heteroatoms selected from O, S and N, which heterocyclic ring is optionally substituted by one or more substituents selected from halo, nitro, -N3, CN, - (C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl; wherein each R17ato R17his independently selected from H, -(C1-10)-alkyl, -(C2-10)-alkenyl and -(C2-10)-alkynyl; wherein aryl10is a 6-14-membered aryl group; wherein indicates that a double bond is present at one of the two possible positions and X is selected from the group consisting of Cl, Br, I, CF3COO, CH3COO, CCl3COO, tetrafluoroborate (BF4), hexafluorophosphate (PF6), bis- trifluoromethanesulfonimide (NTf2), trifluoromethanesulfonate (OTf), dicyanamide (N(CN)2), hydrogen sulphate (HSO4), ethyl sulphate (EtOSO3) and 1-butyl-3- methylimidazolium tetrachloroferrate. In one embodiment R4, R5and R6are all H. In one aspect the invention provides a compound of Formula IA wherein R, R’, R1, R2, R3, X and are as defined for Formula I. The following preferred embodiments relate to compounds of Formula I and Formula IA: Where the compounds relate to Formula I, it is preferred that R4, R5and R6are all H. In one embodiment R is H and R’ is -(C1-12)-alkyl optionally substituted with one or more substituents selected from the group consisting of halo, nitro, -N3, =O, -CN, -OR10a,- SR10b, -COR10c, -COOR10d, -NR10eR10f, -CONR10gR10h, carbocyclea1, aryla1, heterocyclea1, heteroaryla1, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl). In one embodiment R’ is -(C1-12)-alkyl substituted with -(C6-14)-aryl, preferably phenyl. In one embodiment R’ is -(C1-12)-alkyl substituted with -(C6-14)-aryl and -COO(C1- C10alkyl), preferably -COOtBu. In one embodiment R’ is -CH(CH2Ph)COOtBu. In one embodiment R’ is -(C1-12)-alkyl substituted with -SR10awherein R10ais methyl. In one embodiment R’ is -(C1-12)-alkyl substituted with -CONR10gR10hwherein R10gis H and R10his -(C1-C10alkyl). In one embodiment R10his -(C1-C10alkyl) substituted with - CONR17gR17hwherein R17gand R17hare both H. In one embodiment R10his -(C1-C10alkyl) is substituted with -(C6-14)-aryl wherein aryl is phenyl. In one embodiment R’ is -(C1-C10alkyl) substituted with a heterocycle. In one embodiment the heterocycle is indole. In one embodiment R is H and R’ is selected from n-butyl, n-pentyl, n-hexyl, n-heptyl, n- octyl, n-nonyl, n-decanyl, n-undecanyl, and n-dodecanyl or R’ is benzyl. In one embodiment R is H and -NHR’ represents an amino acid, amino acid ester, amino acid amide or a peptide. In one embodiment the amino acid ester is phenylalanine tert-butyl ester. In one embodiment R is H and -NHR’ represents a dipeptide. In one embodiment the dipeptide comprises an amino acid selected from the group comprising methionine, serine, tryptophan, tyrosine, glycine, glutamine and asparagine. In one embodiment the dipeptide comprises phenylalanine amide. In one embodiment -NRR’ represents an amine-functionalised polymer or material. In one embodiment the material is selected from glass, silicon and titanium. In one embodiment the peptide is a 1-10 amino acid oligopeptide. In one embodiment R1, R2and R3are -(C1-14)-alkyl, preferably methyl or ethyl. In one embodiment R1and R2are methyl and R3is -(C6-16)-alkyl, preferably -(C6-14)- alkyl. In one embodiment R1and R2are methyl and R3is cyclohexyl. In one embodiment R1and R2are methyl and R3is -(C1-6)-alkyl substituted with phenyl. In one embodiment X is Cl. In one embodiment the compound is of Formula IA wherein R is H, R’ is selected from n- butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decanyl, n-undecanyl, and n- dodecanyl; R1and R2are methyl and R3is n-decanyl. Preferably, R’ is selected from n- octyl, n-nonyl, n-decanyl, n-undecanyl, and n-dodecanyl. In one embodiment the compound is of Formula IA wherein R is H and R’ is n-dodecanyl; R1and R2are methyl and R3is selected from n-hexyl, n-heptyl, n-octyl, n-nonyl, n- decanyl, n-undecanyl, n-dodecanyl, n-tridecanyl and n-tetradecanyl. Preferably, R3is selected from n-octyl, n-nonyl, n-decanyl, n-undecanyl, n-dodecanyl, n-tridecanyl and n- tetradecanyl. In one embodiment the compound is of Formula IA wherein R is H and R’ is Bn; R1and R2are methyl and R3is selected from n-decanyl, n-undecanyl, n-dodecanyl, n-tridecanyl, n-tetradecanyl, n-pentadecyl and n-hexadecyl. Preferably R3is selected from n- dodecanyl, n-tridecanyl, n-tetradecanyl, n-pentadecyl and n-hexadecyl. In one embodiment the compound is of Formula IA wherein R is H and R’ is - CH(CH2PH)COOtBu: R1and R2are methyl and R3is selected from n-hexyl, n-heptyl, n- octyl, n-nonyl, n-decanyl, n-undecanyl, n-dodecanyl, n-tridecanyl, n-tetradecanyl, n- pentadecyl and n-hexadecyl. The compounds of Formula I and IA exist as isomers as illustrated below with respect to Formula IA. The left hand isomer (α,β-unsaturated) is thermodynamically preferred while the right hand isomer (β,γ-unsaturated) is the kinetic product of the reaction of the invention. Higher reaction temperatures favour the α,β-unsaturated isomer. 5.3 The processes of the invention The inventors have surprisingly discovered that OQA compounds can be made by aza- Michael addition of tertiary amines to allenamides. Accordingly, the invention provides processes for making OQA compounds taking advantage of this knowledge. The allenamide starting material can be functionalised in many ways. Therefore, the processes of the invention provide simple means of obtaining a wide variety of OQA compounds. 5.3.1 Process A In one aspect the invention provides a process of making a compound of Formula I comprising reacting an allenamide compound of Formula II with a tertiary amine of Formula III,

[0055] wherein R, R’, R1-R5, X and are as defined for Formula I. Process A of the invention utilises the unique reactivity, availability and stability of the allenamide group (II). Process A could be considered to be a “click transformation”. Click transformations are easy to perform, give rise to their intended products in very high yields with little or no byproducts, work well under many conditions, and are relatively unaffected by the nature of the groups being connected to each other. The allenamide compound of Formula II can be prepared by any method known in the art. Examples include methods described in (Xie & Ma, 2002) (Trifonov & Oharovats, 1987) (Trifonov & Orahovats, 1986) (Audin, Drut-Grevoz, & Paris, 1993) (Tomida, Nagaki, & Yoshida, 2011) (Mbofana & Miller, 2014). We note that where R4-R6of Formula I are all H, the allenamide (II) can be made by reacting NHRR’ with 3-butynoic acid, as per Process B as described below. In one embodiment the reaction takes place in a solvent that is a cyclic carbonate solvent. In one embodiment the reaction takes place in a solvent selected from propylene carbonate (PC), ethylene carbonate (EC) and acetonitrile (MeCN). In one embodiment a proton source is added to reaction mixture, preferably HCl. In one embodiment about 0.25 to about 0.5 mol proton source is added for each mol of tertiary amine (III). In one embodiment about 1 to about 10 mol tertiary amine (III) is added for each mol of allenamide compound (II). Where the allenamide compound of Formula II comprises an allenamide-functionalised peptide, the reaction can be carried out in solution or attached to a solid phase polymeric support. In some cases the potentially reactive side-chains of the peptide will need to be protected prior to aza-Michael quaternization. A person skilled in the art would readily understand the protection strategies needed (Isidro-Llobet, Alvarez, & Albericio, 2009). The preparation of allenamide-modified peptides is described in Cameron et al. (Cameron, Harris, & Brimble, 2020). Once attached to a solid phase polymeric support, the allenamide (II) is reacted with the tertiary amine (III) to provide a OQA compound of Formula I, once cleaved from the support. Example 7 describes the synthesis of the peptide LYRAK using both solution and solid phase peptide synthesis (SPPS). LYRAK is an OQA compound of Formula I wherein R is H and NHR’ represents a peptide. 5.3.2 Process B While Process A provides a convenient synthesis of a wide range of OQA compounds, it is possible to modify the reaction to provide a one pot synthesis using simpler starting materials in which an allenamide is made and reacted in situ (Process B). Accordingly, in another aspect the invention provides a process of making a compound of Formula IA comprising reacting an amine (IV) with 3-butynoic acid (V) in the presence of a peptide coupling reagent and a tertiary amine of Formula III wherein R, R’, R1-R3, X and are as defined for Formula I. Without being bound by theory, it is believed that Process B proceeds via aza-Michael addition of the tertiary amine (III) to an allenamide intermediate (Formula II above where R4-R6are H). The allenamide intermediate need not be isolated for the reaction to proceed to Formula IA. The compound of Formula IV is a primary or secondary amine, preferably a primary amine. In one embodiment the amine (IV) is an amino acid, amino ester, amino amide or peptide. In one embodiment the amino acid is an ^^-amino acid. In one embodiment the amino acid is an L-amino acid. In one embodiment the peptide is a linear or cyclic oligopeptide, preferably a 1-30, 25, 20, 15 or 10 amino acid oligopeptide, preferably a 1- 10 amino acid oligopeptide. The amine (IV) is reacted with 3-butynoic acid (V) using a peptide coupling agent. A peptide coupling agent is an agent that catalyses the coupling of one amino acid's carboxyl group to the amino group of a second amino acid. Peptide coupling agents are well known and a person skilled in the art would be able to select a suitable agent for the particular reaction to be carried out.

[0056] In one embodiment the peptide coupling agent is Mukaiyama reagent (2-Chloro-l- methylpyridinium iodide (CMPI)). In one embodiment the peptide coupling agent is EEDQ (N-ethoxycarbonyl-2-ethoxy-l,2-dihydroquinoline).

[0057] While both CMPI and EEDQ are excellent, commonly used peptide coupling agents, one agent may be preferred over the other for the particular reaction being carried out. For example, the use of CMPI sometimes generates an undesirable byproduct, lowering the yield and purity of the products. For these reactions, EEDQ is preferred.

[0058] However, EEDQ does not react well in a highly basic environment so a high concentration of the tertiary amine (III) may hinder formation of the allenamide. In these circumstances the use of sym-collidine may enhance the reaction, as discussed below.

[0059] Process B can be carried out in one or two stages. In the one-stage reaction, all reagents are mixed and the reaction provides compounds of Formula IA, via the allenamide intermediate.

[0060] In the two-stage reaction, the allenamide intermediate is generated before the azaMichael addition of tertiary amine (III) to give compounds of Formula IA. The allenamide intermediate may be isolated or purified, or may merely be reacted in situ in the second stage. Formation of the allenamide intermediate may be monitored by HCLC.

[0061] Although the one-stage reaction is simpler, carrying out the reaction in two stages may be advantageous in some circumstances because the reagents can be optimised for each of the two stages, or it may facilitate simpler purification of the final OQA.

[0062] For example, in one embodiment, in the first stage, the 3-butynoic acid (V) is reacted with EEDQ and sym-collidine. Once the allenamide intermediate is formed (monitored by HPLC), further tertiary amine (III) is added, along with a proton donor to facilitate the aza-Michael addition of the tertiary amine to the allenamide.

[0063] Where EEDQ is the peptide coupling agent, the two-stage process may be preferred. Where CMPI is the peptide coupling agent, the one-stage process may be preferred. The relative amounts of the reagents depend on whether the reaction is proceeding in one or two stages, and whether it is carried out in solution phase or attached to a solid phase polymeric support.

[0064] In one embodiment, the tertiary amine (III) is added to the 3-butynoic acid (V) and amine (IV) and the reaction proceeds in one stage.

[0065] In one embodiment, the tertiary amine (III) is not added to the 3-butynoic acid (V) until formation of the allenamide intermediate is substantially complete.

[0066] In one embodiment 2, 3, 5, 10, 15 or 20 molar equivalents of tertiary amine (III) are added relative to the amount of amine (IV). In one embodiment the tertiary amine (III) is present in excess.

[0067] In one embodiment the molar ratio of 3-butynoic acid (V) to amine (IV) in the reaction is between about 1 to 2.

[0068] In one embodiment a proton source is added to reaction mixture, preferably HCI. In one embodiment about 0.25 to about 0.5 mol proton source is added for each mol of tertiary amine (III). In the one-stage reaction, the proton source is preferably added after about 30 to about 60 min of reaction. In the two-stage reaction the proton source is added with the tertiary amine (III).

[0069] In one embodiment the reaction takes place in a solvent that is a cyclic carbonate solvent. In one embodiment the reaction takes place in a solvent selected from PC, EC and MeCN. Cyclic carbonates are considered to be quite "green" as their production involves the consumption of carbon dioxide which is 100% atom economic. They are largely biodegradable, non-toxic and safe to use.

[0070] Process B is exemplified in Scheme 1 below where 1 is the allenamide intermediate compound that, upon reaction with triethylamine (TEA) forms the OQA compounds of the invention 2a and 2a*.

[0071] Scheme 1

[0072] Scheme 1 is shown in more detail in Figure 2. As in Process A, where the amine (IV) comprises a peptide, Process B can be carried out in solution or attached to a solid phase polymeric support. Example 10 outlines the synthesis of three OQA polymyxin analogues using the two-stage Process B carried out on a solid phase polymeric support. Once the peptide was made using standard SPPS techniques, the allenamide handle was introduced by coupling 3-butynoic acid using EEDQ in accordance with Cameron et al. (Cameron, Harris, & Brimble, 2020). The OQA moiety was then introduced by aza- Michael addition of a tertiary amine to the allenamide. As would be understood by a person skilled in the art, where Process B is carried out during on-resin (during SPPS), the ratios of reagents may differ from those used for the solution phase reaction. For example, in one embodiment the ratio of 3-butynoic acid (V) to resin-bound amine (IV) is about 10. Excess reagents may be used due to ease of purification. As would also be understood by a person skilled in the art, this is typical in on-resin chemistry, so as to enhance efficiency due to the ease of removal of excess reagents simply by filtration and washing of the solid phase resin-supported compound. In both Process A and Process B, the counterion X is determined by the reagents used in the process. The proton donor, peptide coupling agent and resin cleavage reagent all may contribute a counterion for the compounds of Formulae I and IA. However, the final purification step will ultimately dictate the nature of the counterion X. For example, the solvents used to purify the compounds of the invention by HPLC typically include a charge modifying agent (usually TFA), but the anion may be exchanged for Cl by freeze drying 2 or 3 times in dilute HCl. The nature of X does not affect the functioning of the compounds of Formulae I and IA. 5.3.3 Preferred embodiments of the processes of the invention The following preferred embodiments relate to both Process A and Process B: In one embodiment R is H and R’ is -(C1-12)-alkyl optionally substituted with one or more substituents selected from the group consisting of halo, nitro, -N3, =O, -CN, -OR10a,- SR10b, -COR10c, -COOR10d, -NR10eR10f, -CONR10gR10h, carbocyclea1, aryla1, heterocyclea1, heteroaryla1, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl). In one embodiment R’ is -(C1-12)-alkyl substituted with -(C6-14)-aryl, preferably phenyl. In one embodiment R’ is -(C1-12)-alkyl substituted with -(C6-14)-aryl and -COO(C1- C10alkyl), preferably -COOtBu. In one embodiment R’ is -(C1-12)-alkyl substituted with -SR10awherein R10ais methyl. In one embodiment R’ is -(C1-12)-alkyl substituted with -CONR10gR10hwherein R10gis H and R10his -(C1-C10alkyl). In one embodiment R10his -(C1-C10alkyl) substituted with - CONR17gR17hwherein R17gand R17hare both H. In one embodiment R10his -(C1-C10alkyl) is substituted with -(C6-14)-aryl wherein aryl is phenyl. In one embodiment R’ is -(C1-C10alkyl) substituted with heterocycle. In one embodiment the heterocycle is indole. In one embodiment R is H and -NHR’ represents an amino acid, amino acid ester, amino acid amide or a peptide. In one embodiment the amino acid ester is phenyl alanine tert- butyl ester. In one embodiment R is H and -NHR’ represents a dipeptide. In one embodiment the dipeptide comprises an amino acid selected from the group comprising methionine, serine, tryptophan, tyrosine, glycine, glutamine and asparagine. In one embodiment the dipeptide comprises phenylalanine amide (phenylalanine carboxamide). In one embodiment the peptide is a 1-10 amino acid oligopeptide. In one embodiment -NRR’ represents an amine-functionalised polymer or material. In one embodiment the material is selected from glass, silicon and titanium. In one embodiment R1, R2and R3are -(C1-14)-alkyl, preferably methyl. In one embodiment R1and R2are methyl and R3is -(C6-14)-alkyl. In one embodiment R1and R2are methyl and R3is cyclohexyl. In one embodiment R1and R2are methyl and R3is -(C1-6)-alkyl substituted with phenyl. In one embodiment X is Cl. For Process A, in one embodiment R4, R5and R6are independently selected from -(C1-12)-alkyl. In one embodiment R4, R5and R6are all H. In one aspect the invention provides a compound of Formula IA or a process of making a compound of Formula IA selected from , , , ,

[0073]

[0074]

[0075] 5.4 Properties of the OQA compounds of the invention

[0076] The OQA compounds of the invention share the antimicrobial properties demonstrated by QACs but are generally expected to be more environmentally friendly. The peptide bond of the OQA, when cleaved, gives rise to a zwitterionic OQA. Loss of the net positive charge is expected to render the molecule unable to bind to the membrane of microbes. In addition, the bond cleavage that occurs naturally in biodegradation results in separation of the hydrophobic ends of the molecule. These are needed to insert into the bacterial membrane to effect disruption, so the OQA is expected to forfeit its bioactivity.

[0077] Table 1: Comparison of existing QACs with OQAs of the invention

[0078] The processes of the invention have been used to prepare novel OQA compounds that mimic common commercially available QACs. As shown in Example 9, the OQA compounds of the invention have comparable or improved antimicrobial activity compared to their commercially available QAC counterparts.

[0079] In Example 11, OQA analogues of polymyxin B were tested for antimicrobial activity against their QAC analogues. Polymyxin B is an antimicrobial peptide which disrupts the membrane structures of Gram-negative bacteria. QAC-polymyxins were developed in an attempt to broaden the activity of polymyxin (Ongwae, et al., 2020). Many have been found to be more potent than polymyxin B itself towards Gram-positive bacteria but can retain potent activity against Gram-negative bacteria.

[0080] The results in Example 11 show that the OQA analogues of polymyxin B were even more potent agents against E. coli and S. aureus strains than were their QAC analogues. Example 12 shows that the OQA compounds of the invention had shorter half-lives than DDAC and so will not linger in the marine environment for as long as QACs.

[0081] Accordingly, the OQA compounds and processes of the invention have great potential in the battle against infectious diseases. OQA compounds combine the efficacy seen in related QACs with superior biodegradability properties. OQA compounds are readily cleaved into inactive components. This reduces the time in which these compounds persist in the environment at sub-lethal concentrations, where they could contribute to antimicrobial resistance.

[0082] The processes of the invention also provide ways of functionalising complex molecules such as polymyxin with a quaternary ammonium moiety, without the use of toxic alkylating agents.

[0083] In one aspect the invention provides an antimicrobial formulation comprising a compound of Formula I or IA and excipients.

[0084] 5.5 Biomaterials comprising the OQA compounds of the invention

[0085] As well as having been attached to antibiotic peptides such as polymyxin B, quaternary ammonium (QA) moieties have been incorporated onto a range of materials to confer antimicrobial properties (Jiao, et al., 2017). As would be appreciated by a person in the art, the same chemistry can be used to incorporate OQA compounds of the invention onto materials which need to be resistant to microorganisms and biofilm formation. OQA compounds may be incorporated via a polymer grafted onto the surface of the material or by direct reaction with a surface-bound initiator.

[0086] For example, De Zoysa and Sorojini (De Zoysa & Sarojini, 2016) describe functionalising glass, silicon and titanium surfaces with (3-aminopropyl)triethoxysilane (APTES). This reaction provides amine groups that may be converted to an allenamide and then reacted with a tertiary amine to provide OQA-functionalised surfaces as shown below.

[0087]

[0088] Similarly, other positions in the OQA compounds of the invention can be reacted with activated surfaces. For example, alkyne substituents on a OQA compound could react with azide-derivativised surfaces, generated using (3-azidopropyl)trimethoxysilane (AzPTMS). Similarly, azide substituents on a OQA compound could react with alkyne- derivatised surfaces.

[0089] Consequently, the compounds of Formula I and IA include within their scope, compounds in which -NRR' represents an amine-functionalised polymer or material.

[0090] In one aspect the invention provides a compound of Formula I or Formula IA which is bound to the surface of a material via a trialkyloxysilane. In one aspect the invention provides a biomaterial comprising one or more compounds of Formula I or Formula IA.

[0091] In one embodiment the material is selected from glass, filter paper, stainless stell, titanium, gold, silicon wafer, silicon rubber, PVDF membrane, polymer microsphere, polyolefin and polypropylene. In one embodiment the material comprises or coats a medical device, for example a surgical tool, catheter, heart valve, pacemaker, stent, prosthetic joint or contact lens.

[0092] 6. EXAMPLES

[0093] General Experimental Methods

[0094] Reagents

[0095] All reagents were used as supplied. Solvents for RP-HPLC were purchased as HPLC grade and used without further purification. O-(6-chlorobenzotriazol-l-yl)- / V, / V, / V', / V'- tetramethyluronium hexafluorophosphate (HCTU), and Fmoc-amino acids were purchased from CS Bio (Shanghai, China). Fmoc amino acids were supplied with the following side-chain protection: Fmoc-Arg(Pbf)-OH (Pbf = 2, 2, 4,6,7- pentamethyldihydrobenzofuran-5-sulfonyl), Fmoc-Asn(Trt)-OH (Trt = trityl), Fmoc- Lys(Boc)-OH (Boc = tert-butyloxycabonyl) Fmoc-Ser(tBu)-OH (tBu = tert-butyl) and Fmoc-Tyr(tBu)-OH. 3-Butynoic acid, 2-chloro-l-methylpyridinium iodide (CMPI or Mukaiyama reagent), / V-ethoxycarbonyl-2-ethoxy-l,2-dihydroquinoline (EEDQ), L- phenylalanine tert-butyl ester hydrochloride (HzN-Phe-OtBu.HCI) and 4-[(R,S)-o-[l-(9H- fluoren-9-yl)]methoxycarbonylamino]-2,4-dimethoxy]phenoxyacetic acid (Fmoc-Rink amide linker) were purchased from AK Scientific (Union City, CA, USA). Boc-Lys-OMe.HCI and 2-chlorotrityl chloride (2-CTC) polystyrene resin were purchased from ChemPep (Wellington, Florida, USA). Di-tert-butyl dicarbonate (BoczO) and H-Gly-OBzI.HCI were purchased from GL Biochem (Shanghai, China). Fmoc-Sar-OH was purchased from Chem-Impex Int'l. Inc. (Wood Dale, IL, USA). / V, / V-di methyl butylamine (DMBA) was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Deuterochloroform (CDCI3) and dimethyl sulfoxide-de (DMSO-de) were purchased from Cambridge Isotope Laboratories (Tewksbury, MA, USA). Chloroform (CHCI3), L-cysteine, Boc-L-Cys-OMe, / V, / V-diethylethanamine (triethylamine, TEA), / V, / V-diethylmethylamine (DEMA), N,N- diisopropylethylamine (DIPEA), / V, / V-dimethylcyclohexylamine (DMCHA), N,N- dimethyldecylamine (DMDA), / V, / V-di methylethylamine (DMEA), / V, / V-dimethylhexylamine (DMHA), / V, / V-di methyloctylamine (DMOA), / V, / V-di methyl phenethylamine (DMPEA), N,N- dimethyltetradecylamine (DMTDA), / V-methylmorpholine (NMM), piperidine, 4-methyl- l,3-dioxolan-2-one (propylene carbonate, PC), triisopropylsilane (TIPS), trimethylamine (TMA), and 2,4,6-trimethylpyridine (sym-collidine), were purchased from Sigma Aldrich (St. Louis, MO, USA). l,3-Dioxolan-2-one (ethylene carbonate, EC) was purchased from Janssen Chemica (Beerse, Belgium). / V, / V-dimethylformamide (DMF) (synthesis grade), and acetonitrile (MeCN) (HPLC grade) were purchased from Thermo Fisher Scientific (Hampshire, NH, USA). Acetic acid (AcOH), dichloromethane (CH2CI2), and hydrochloric acid (HCI) were purchased from ECP Limited (Auckland, New Zealand). Trifluoroacetic acid (TFA) was purchased from Oakwood Chemicals (Estill, SC, USA). Diethyl ether (Et2O) was purchased from Avantor Performance Materials (Center Valley, PA, USA). Milli-Q high purity deionised water (MQ H2O) was available from a Sartorius Arium® Pro Ultrapure Water System from Sartorius Stedim Biotech (Gottingen, Germany).

[0096] Aminomethyl polystyrene (AM PS) resin (0.85 mmol / g) was made in-house by Yann Hermant according to literature (Harris, Yang, & Brimble, 2011).

[0097] General procedures for purification and analysis

[0098] Analytical reverse phase high-performance liquid chromatography (RP-HPLC) was performed on a Waters (Waltham, MA, USA) Alliance equipped with a Phenomenex (Torrance, CA, USA) Luna C18 column (100 A, 5 pm, 4.6 mm x 250 mm) using a linear gradient as specified, where solvent A was 0.1% TFA in MQ H2O and solvent B was 0.1% TFA in MeCN. All analytical RP-HPLC runs were performed on a 30 minute gradient of 5- 95% solvent B, ca. 3% B / min. Chromatograms were recorded at 214 nm and 254 nm. Semi-preparative RP-HPLC was performed on a Waters 1525 Binary HPLC pump equipped with a Waters 2489 UV / visible detector using a Waters Xterra Prep MS C18 OBD (125 Å, 10 μm, 19 mm × 300 mm), with chromatograms recorded at 214 nm and 254 nm. Gradient systems for semi-preparative RP-HPLC were adjusted according to elution times and peak profiles obtained from analytical RP-HPLC chromatograms. High-resolution mass spectrometry (HRMS) was performed on a on a Bruker (Billerica, MA, USA) micrOTOF-Q Mass Spectrometer in electrospray ionisation (ESI) positive mode. Low-resolution mass spectrometry was performed on a Waters Quattro micro API Mass Spectrometer in ESI positive mode. Nuclear magnetic resonance (NMR) spectra were recorded at room temperature (rt) in CDCl3 on either a Bruker AVANCE 400 or a Bruker AVANCE 500 spectrometer operating at 400 / 500 MHz for1H nuclei and 100 / 125 MHz for13C nuclei, respectively. Chemical shifts are reported in parts per million (ppm) on the δ scale from tetramethylsilane (TMS) at 0.00 ppm. Shifts were referenced to the residual solvent peaks CDCl3at δC= 77.16 ppm for13C spectra and TMS at δH= 0.00 ppm for1H spectra. The1H NMR shift values are reported as chemical shift (δ), multiplicity (s = single, d = doublet, t = triplet, q = quartet, m = multiplet, dd = doublet of doublets, ddd = doublet of doublets of doublets, td = triplet of doublets, qd = quartet of doublets), coupling constant (J in Hz), relative integral and assignments.13C NMR values were reported as the chemical shift (δ), the degree of hybridisation and assignment. Structural assignments were achieved with the aid of DEPT135, COSY and HSQC experiments where required. Infrared (IR) spectra were recorded on a Perkin Elmer Spectrum 100 FT-IR spectrometer using a diamond ATR sampling accessory. Example 1: Synthesis of compounds of Formula I and Formula IA Allenamide 1 was prepared and isolated according to the protocol by Cameron et al. (Cameron, Harris, & Brimble, 2020). To allenamide 1 (2 mg, 0.007 mmol) dissolved in CH2Cl2(200 μL) was added TEA (30 equiv.) and HCl (0.035 mmol, 5 equiv., 5 M, aq.). Aliquots (2 μL) of the reaction mixture were diluted in H2O / MeCN (1:1, v / v, 100 μL) at select time intervals for reaction monitoring by injection to RP-HPLC (214 nm). This reaction was repeated without the addition of HCl and similarly monitored by RP-HPLC. RP-HPLC showed the reaction proceeded via the allenamide intermediate. Mass spectrometry suggested that the two products corresponded to olefinic ammonium compounds as a result of an aza-Michael addition of TEA to the allenamide moiety 1. This was unexpected since TEA has been reported to catalyse thia-Michael reactions exclusively via base catalysis instead of the nucleophilic initiation pathway (Desmet, et al., 2017), (Frayne & Northrop, 2018). Each species was isolated and analysed by NMR which revealed that 2a and 2a* were indeed OQAs, with isomer 2a being the kinetic olefinic-containing product and isomer 2a*, the thermodynamic α,β-unsaturated product. Additionally, OQA 2a was observed to slowly isomerise to 2a* during the reaction, likely catalysed by the presence of excess base. Characterisation IR νmax(neat): 3258, 2989 (alkene), 1730 (ester), 1670 (amide, olefinic alkene), 1151 (ester), 704 (phenyl) cm-1;1H NMR (400 MHz, CDCl3): δH= 8.44 (d, J = 7.8 Hz, 1H, HN), 7.27–7.23 (m, 4H, Ar-H), 7.19 (t, J = 6.32 Hz, 1H, Ar-H), 5.73 (d, J = 3.9 Hz, 1H, Ha-11), 5.52 (d, J = 4.0 Hz, 1H, Hb-11), 4.58 (ddd, J = 9.7, 7.9, 5.5 Hz, 1H, H-2), 3.53 (d, J = 16.2 Hz, 1H, Ha-9), 3.45 (q, J = 7.2 Hz, 6H, H-12), 3.38 (d, J = 16.2 Hz, 1H, Hb- 9), 3.16 (dd, J = 14.0, 5.5 Hz, 1H, Ha-3), 2.95 (dd, J = 14.0, 9.8 Hz, 1H, Hb-3), 1.38 (s, 9H, H-15), 1.14 (t, J = 7.1 Hz, 9H, H-13);13C NMR (100 MHz, CDCl3): δC= 170.6 (C, C=O), 167.0 (C, C=O), 141.6 (C, C-4), 137.3 (C, C-10), 129.4 (CH, C-6), 128.5 (CH, C- 5), 126.8 (CH, C-7), 119.5 (CH2, C-11), 81.9 (C, C-14), 55.3 (CH, C-2), 52.8 (CH2, C- 12), 37.4 (CH2, C-3), 36.5 (CH2, C-9), 27.9 (CH3, C-15), 7.8 (CH3, C-13); HRMS (ESI / Q- TOF) m / z: [M]+calcd for C23H37N2O3+, 389.2799; found, 389.2790 IR νmax (neat): 3260, 2983 (alkene), 1729 (ester), 1674 (amide, conjugated alkene), 1156 (ester), 720 (phenyl) cm-1;1H NMR (500 MHz, CDCl3): δH = 7.88 (d, J = 7.7 Hz, 1H, HN), 7.19–7.28 (m, 5H, Ar-H), 6.61 (s, 1H, H-9), 4.75 (ddd, J = 7.7, 7.3, 7.3 Hz, 1H, H-2), 3.47 (q, J = 7.2 Hz, 6H, H-12), 3.17 (dd, J = 14.0, 6.3 Hz, 1H, Ha-3), 3.01 (dd, J = 14.0, 8.3 Hz, 1H, Hb-3), 2.24 (s, 3H, H-11), 1.40 (s, 9H, H-15), 1.27 (t, J = 7.1 Hz, 9H, H-13);13C NMR (125 MHz, CDCl3): δC = 170.3 (C, C=O), 163.1 (C, C=O), 145.7 (C, C-4), 137.3 (C, C-10), 129.7 (CH, C-6), 128.4 (CH, C-5), 126.7 (CH, C-7), 125.8 (CH, C-9), 81.9 (C, C-14), 55.4 (CH, C-2), 53.1 (CH2, C-12), 37.9 (CH2, C-3), 28.0 (CH3, C-15), 12.8 (CH3. C-11), 8.3 (CH3, C-13); HRMS (ESI / Q-TOF) m / z: [M]+calcd for C23H37N2O3+, 389.2799; found, 389.2796. Example 2: Investigation of coupling reagents (Mukaiyama reagent and EEDQ) H2N-Phe-OtBu.HCl (5 mg, 0.019 mmol, 1 equiv.) was dissolved in the specified solvent (200 μL) to which was added TEA (2.7 μL, 0.019 mmol, 1 equiv.) to neutralise the HCl salt. 3-Butynoic acid (3.3 mg, 0.038 mmol, 2 equiv.) and TEA (5.4 μL, 0.038 mmol, 2 equiv.) were dissolved and incubated in the same solvent (200 μL) for 10 min at rt. These two mixtures were added in rapid succession to the coupling reagent (0.037 mmol, 1.9 equiv.), and the mixture shaken for up to 1h. Somewhat surprisingly, the reaction with EEDQ showed no consumption of starting materials, whereas the reaction with Mukaiyama reagent gave rapid conversion to allenamide 1 and OQA 2a, with 68% and 21% conversions at only 10 min, respectively. Proceeding with Mukaiyama induced coupling for 10 min, the transformation to OQA 2a was promoted with the addition of HCl (5 equiv.) and excess TEA (10 equiv.), affording product 2a at 85% conversion after 24 h. Subsequent testing of different stoichiometric amounts of HCl while maintaining 10 equiv. of TEA revealed that half the molar equivalence of acid to base was optimal. Example 3: Optimisation of solvent and Brønsted acid Step 1: H2N-Phe-OtBu.HCl (5 mg, 0.019 mmol, 1 equiv.) was dissolved in the specified solvent (200 μL) to which was added TEA (2.7 μL, 0.019 mmol, 1 equiv.) to neutralise the HCl salt. 3-Butynoic acid (3.3 mg, 0.038 mmol, 2 equiv.) and TEA (5.4 μL, 0.038 mmol, 2 equiv.) were dissolved and incubated in the same solvent (200 μL) for 10 min at rt. These two mixtures were added in rapid succession to Mukaiyama reagent (9.4 mg, 0.037 mmol, 1.9 equiv.), and was shaken for 10 min. Step 2: To the reaction mixture was added excess TEA (27.0 μL, 0.19 mmol, 10 equiv.) and a Brønsted-Lowry acid (0, 2.5, 5, or 7.5 equiv.). The mixture was further shaken until maximum conversion to quaternary ammonium compound (QAC) 2a was observed on analytical RP-HPLC (214 nm). Aliquots (2 μL) of the reaction mixture were diluted in H2O / MeCN (1:1, v / v, 100 μL) at select time intervals for reaction monitoring by injection to RP-HPLC. Both steps of the reactions were performed at the same temperature, either at rt, 40 or 60°C as specified. The solvents tested were ethylene carbonate (EC), propylene carbonate (PC), acetonitrile (MeCN), N,N-dimethylformamide (DMF) and dichloromethane (CH2Cl2). The cyclic carbonates performed the best, with EC the best performing solvent, providing 88% conversion of OQA 2a within 1 h (albeit aided slightly by the requisite increased reaction temperature (40°C, given mp: 36.4°C for EC)). The reaction rate in PC and MeCN were comparable with each other, offering ∼90% conversions after 5 h. DMF was undoubtedly the poorest performing solvent tested with only 33% conversion of 2a after 23 h. Although EC was the superior solvent in terms of reaction kinetics, the requisite elevated temperature increased base-induced isomerisation and fails to adhere to the green chemistry principle of reactions being conducted under ambient conditions. Therefore, propylene carbonate was the preferred solvent. It was used in the remainder of the experiments aimed at optimising a green chemistry protocol to prepare OQA compounds by aza-Michael addition to allenamides. Investigation of alternate proton sources to HCl, trifluoroacetic acid (TFA) and acetic acid (AcOH), further demonstrated the importance of proton transfer as the rate of conversion to 2a increased with acid strength (HCl>TFA>AcOH). Note that HCl is in aqueous solution, where water can also function as a proton donor for the rate- determining proton transfer. Nevertheless, HCl was found to be the proton source of choice. Example 4: Synthesis of compounds of Formula I and IA using a range of tertiary amine nucleophiles A range of compounds of the invention were prepared using different tertiary amines including with those containing alkyl substituents of varying lengths, a cycloalkane substituent, an aromatic substituent and a cyclic tertiary amine. Step 1: H2N-Phe-OtBu.HCl (5 mg, 0.019 mmol, 1 equiv.) was dissolved in propylene carbonate (200 μL) to which was added the tertiary amine (0.019 mmol, 1 equiv.) to neutralise the HCl salt. 3-Butynoic acid (3.3 mg, 0.038 mmol, 2 equiv.) and tertiary amine (0.038 mmol, 2 equiv.) were incubated in propylene carbonate (200 μL) for 10 min at rt. These two mixtures were rapidly added to Mukaiyama reagent (9.4 mg, 0.037 mmol, 1.9 equiv.), and was shaken for 10 min at rt. Step 2: To the reaction mixture was added excess of the same tertiary amine used in Step 1 (0.19 mmol, 10 equiv.) and aq. HCl (19.4 µL, 5 M, 0.095 mmol, 5 equiv.). The mixture was further shaken at rt until maximum conversion to QAC 2 was observed on analytical RP-HPLC (214 nm). Aliquots (2 μL) of the reaction mixture were diluted in H2O / MeCN (1:1, v / v, 100 μL) at select time intervals for reaction monitoring by injection to RP-HPLC. Pleasingly, all eleven tertiary amines tested were amenable to the aza-Michael addition, with excellent product selectivity and superb conversions to the respective olefinic products (85-90% conversions) in most cases. The compounds produced are shown below. Note that only one isomer is depicted but that both isomers are produced in the reaction.

[0099] Unsurprisingly, steric hindrance had a major influence on the rate of reaction for the remaining tertiary amines examined. Substitution of a single ethyl group of TEA for a methyl group, N,N-diethylmethylamine (DEMA), resulted in an approximate five-fold improvement in the rate of conversion, requiring only 1 h (2d). Furthermore, the series of dimethylalkylamines with alkyl chains lengths from C4to C10provided similar rapid conversions (2e-2h), while only C14 (2i) required the reaction to be extended beyond 1 h. The least hindered tertiary amines, trimethylamine (TMA) and N,N- dimethylethylamine (DMEA), however only gave moderate conversions to QACs 2b (76% at 2 h) and 2c (82% at 75 min), respectively, due to their volatility. Intriguingly, N,N- dimethylphenethylamine (DMPEA) provided the best conversion (90% at 45 min) to its corresponding QAC, 2j, despite the potential bulk imparted by the phenyl group. Without being bound by theory, it is believed that the intervening two-carbon spacer between the phenyl group and the nucleophilic nitrogen may be rendering the steric effects of the phenyl group to be negligible in the immediate area surrounding the nitrogen. This may be substantiated by the result where N,N-dimethylcyclohexylamine (DMCHA), lacking a spacer between the bulky cyclohexane group and the nitrogen atom, gave a notably poorer conversion rate to QAC 2k (80% at 60 min). N-methylmorpholine (NMM), as expected from its reduced basicity and steric hindrance, proved to be the least reactive, but still offered 89% conversion to QAC 21 at 24 h.

[0100] Example 5: Synthesis of compounds of Formula I and IA using a range of primary amine substrates

[0101] To assess the reaction with respect to the primary amine substrate, a series of dipeptides with unprotected side-chain functionalities was prepared by standard Fmoc- SPPS on Rink-amide aminomethyl polystyrene (PS) resin. The C-terminal residue was kept constant as phenylalanine to facilitate RP-HPLC reaction monitoring, while the N- terminal residue presented various unprotected side-chain functionalities, namely arginine (guanidino), histidine (imidazole), serine (hydroxyl), asparagine (side-chain carboxamide), methionine (thioether), tyrosine (tyrosyl) and tryptophan (indole). Upon liberation of the peptides from the resin with TFA and purification, the TFA salt was exchanged for HCI to avoid trifluoroacetylation during the reactions. Taking N,N- dimethylhexylamine (DMHA) as the model tertiary amine nucleophile, the optimised procedure was carried out on these dipeptides in accordance with Scheme X.

[0102] Scheme X

[0103] Step 1: The specified substrate (0.019 mmol, 1 equiv.) was dissolved in propylene carbonate (200 pL) to which was added DMHA (3.37 pL, 0.019 mmol, 1 equiv.) to neutralise the HCI salt. 3-Butynoic acid (3.3 mg, 0.038 mmol, 2 equiv.) and DMHA (6.74 pL, 0.038 mmol, 2 equiv.) were incubated in propylene carbonate (200 pL) for 10 min at rt. These two mixtures were added in rapid succession to Mukaiyama reagent (9.4 mg, 0.037 mmol, 1.9 equiv.), and was shaken for 10 min at rt.

[0104] Step 2: To the reaction mixture was added excess DMHA (33.7 pL, 0.19 mmol, 10 equiv.) and aq. HCI (19.4 pL, 5 M, 0.095 mmol, 5 equiv.). The mixture was further shaken at rt until maximum conversion to OQA 2 was observed on analytical RP-HPLC (214 nm). Aliquots (2 pL) of the reaction mixture were diluted in H2O / MeCN (1: 1, v / v, 100 pL) at select time intervals for reaction monitoring by injection to RP-HPLC. The compounds produced are shown below along with their yields.

[0105] Pleasingly, the reaction was robust in the presence of thioether, hydroxyl, side-chain carboxamide and indole functional groups, with the dipeptides achieving excellent conversions (87-96% conversions) to their respective QACs (3f-6f) within 30-45 min. The reaction in the presence of the phenol group of tyrosine yielded 70% conversion to QAC 7f after 10 min. However, further conversion could not be obtained as the routine protocol of providing excess TEA and HCI after 10 min resulted in formation of an unknown by-product, speculated to be a Mukaiyama adduct at the phenol group. Unfortunately, conversion to neither allenamide or desired olefinic ammonium compound could be effected in the presence of guanidine (Arg) or indole (His) functional groups. A sarcosine-phenylalanine dipeptide was also prepared to assess if the reaction could be applied to a secondary amine, which surprisingly posed little challenge as conversion to the corresponding OQA 8f was comparable to primary amines. Example 6: Synthesis of compounds of Formula I and IA that mimic common QACs - BAC and DDAC

[0106] Mimics of BAC and DDAC were made using the protocol set out below.

[0107] Step 1: To the specified substrate (0.019 mmol, 1 equiv.) was added 3-butynoic acid (3.3 mg, 0.038 mmol, 2 equiv.), EEDQ (9.1 mg, 0.037 mmol, 1.9 equiv.) and sym- collidine (0.038 mmol, 2 equiv.) in propylene carbonate (200 pL) for 1 h at rt.

[0108] Step 2: To the reaction mixture was first added the tertiary amine (0.29 mmol, 15 equiv.), followed by aq. HCI (19.4 pL, 5 M, 0.095 mmol, 5 equiv.). The mixture was further shaken at rt until maximum conversion to QAC 2 was observed on analytical RP- HPLC (214 nm). Aliquots (2 pL) of the reaction mixture were diluted in H2O / MeCN (1 : 1, v / v, 100 pL) at select time intervals for reaction monitoring by injection to RP-HPLC.

[0109] Reaction with DMHA transformed the allenamide intermediate to OQA mimic 9f in 81% conversion after 2 h. Using this EEDQ one-pot methodology, OQA mimics of DMDA with similar backbone lengths, lOf and llh, were also prepared and afforded each at 24% and 43% isolated yields, respectively. Mimics lOf and llh differ in that llh closely replicates the length of alkyl substituents bonded to the nitrogen atom, whereas lOf solely mimics the backbone length regardless the position of the nitrogen. Characterisation OQA 9f was isolated from the reaction mixture in PC using IEX SPE and further purified by semi-preparative RP-HPLC, affording OQA 9f as an amorphous white solid (1.9 mg, 23%); RP-HPLC: tR = 17.0 min; IR vmax (neat): 2927, 2859, 1669, 1550, 1455, 1200, 800, 718, 700 cm-1; HRMS (ESI / Q-TOF) m / z: [M]+calculated for C19H31N2O+: 303.2431, observed: 303.2430;13C NMR (125 MHz, CDCl3): δC= 167.4 (C=O, C-6), 145.4 (C, C- 8), 138.5 (C, C-4), 128.6 (2 × CH, C-2), 128.2 (2 × CH, C-1), 127.3 (CH, C-3), 115.6 (CH2, C-9), 65.2 (CH2, C-10), 52.5 (2 × CH3, C-16), 43.9 (-NH-CH2, C-5), 36.6 (CH2, C- 7), 31.5 (CH2), 29.6 (CH2), 25.7 (CH2), 23.7 (CH2, C-11), 22.5 (CH2), 14.0 (CH3, C-15);1H NMR (500 MHz, CDCl3): δH= 9.42 (br s, 1H, NH amide), 7.37-7.11 (m, 5, Bz-H), 5.61 (d, J = 3.2 Hz, 1H, Ha-9), 5.43 (d, J = 3.5 Hz, 1H, Hb-9), 4.34 (d, J = 4.2 Hz, 2H, H-5), 3.90 (t, J = 7.5 Hz, 2H, H-10), 3.78 (s, 2H, H-7), 3.37 (s, 6H, H-16), 1.42 (br s, 2H, H-11), 1.52-1.45 (m, 4H, H-6 and H-13), 1.24-1.14 (m, 6H, H-12 to H-14), 0.80 (t, J = 6.2 Hz, 3H, H-15) QOC 10f was isolated from the reaction mixture in PC using IEX SPE and further purified by semi-preparative RP-HPLC, affording QOC 10f as an amorphous white solid (2.0 mg, 24%); RP-HPLC: tR= 24.8 min; IR vmax(neat): 2923, 2854, 1668, 1554, 1463, 1201, 800, 719 cm-1; HRMS (ESI / Q-TOF) m / z: [M]+calculated for C22H45N2O+: 353.3526, observed: 353.3525;13C NMR (125 MHz, CDCl3): δC = 167.1 (C=O, C-11), 145.6 (C, C- 13), 115.3 (CH2, C-14), 65.1 (CH2, C-15), 52.5 (2 × CH3, C-21), 40.4 (-NH-CH2, C-10), 36.4 (CH2, C-12), 32.0 (CH2), 31.5 (CH2), 29.7 (CH2), 29.5 (CH2, N-CH2-CH2-), 29.4 (CH2), 27.3 (CH2), 25.8 (CH2), 23.8 (CH2, N-CH2-CH2-), 22.8 (CH2-CH3), 22.5 (CH2-CH3), 14.3 (CH3), 14.0 (CH3);1H NMR (500 MHz, CDCl3): δH = 8.94 (t, J = 4.9 Hz, 1H, NH amide), 5.64 (d, J = 3.6 Hz, 1H, Ha-14), 5.43 (d, J = 3.7 Hz, 1H, Hb-14), 3.98 (t, J = 8.4 Hz, 2H, H-15), 3.76 (s, 2H, H-12), 3.46 (s, 6H, H-21), 3.15 (dt, J = 7.3, 6.0 Hz, 2H, H- 10), 1.52-1.42 (m, 4H, H-9 and H-16), 1.26-1.17 (m, 20H, H-2 to H-8, H-17 and H-18), 0.81 (t, J = 7.0 Hz, 3H, H-1), 0.81 (t, J = 7.0 Hz, 3H, H-20) OQA 11h was isolated from the reaction mixture in PC using IEX SPE and further purified by semi-preparative RP-HPLC, affording OQA 11h as an amorphous white solid (3.7 mg, 43%); RP-HPLC: tR = 25.8 min; IR vmax (neat): 2924, 2855, 1668, 1552, 1466, 1260, 1093, 1020, 802, 719 cm-1; HRMS (ESI / Q-TOF) m / z: [M]+calculated for C23H47N2O+: 367.3683, observed: 367.3683;13C NMR (125 MHz, CDCl3): δC= 167.1 (C=O, C-8), 145.7 (C, C-10), 115.3 (CH2, C-11), 65.6 (CH2, C-12), 52.8 (2 × CH3, C-22), 40.4 (-NH- CH2, C-7), 36.6 (CH2, C-9), 32.0 (CH2), 31.9 (CH2), 29.6 (CH2), 29.5 (CH2), 29.4 (CH2), 29.4 (CH2), 29.1 (CH2), 27.2 (CH2), 26.2 (CH2), 23.9 (CH2), 22.8 (CH2-CH3), 22.8 (CH2- CH3), 14.2 (CH3), 14.2 (CH3);1H NMR (500 MHz, CDCl3): δH= 8.94 (t, 1H, NH amide), 5.64 (d, J = 3.6 Hz, 1H, Ha-11), 5.43 (d, J = 3.7 Hz, 1H, Hb-11), 3.89 (t, J = 8.3 Hz, 2H, H-12), 3.80 (s, 2H, H-9), 3.41 (s, 6H, H-22), 3.15 (dt, J = 7.2, 7.1 Hz, 2H, H-7), 1.52- 1.45 (m, 4H, H-6 and H-13), 1.30-1.12 (m, 22H, H-2 to H-5 and H-14 to H-20), 0.81 (t, J = 6.8 Hz, 3H, H-1 or H-20), 0.79 (t, J = 6.9 Hz, 3H, H-1 or H-20) Example 7: Synthesis of quaternary ammonium-modified allenamide LYRAK 15 LYRAK was selected as a model peptide on which to test the processes of the invention. The synthesis is set out in Figure 1. The allenamide was prepared on-resin by acylation with 3-butynoic acid in CH2Cl2 to functionalize the N-terminus of 11, prior to aza-Michael addition of DMHA as discrete steps. With resin-bound allenamidyl LYRAK 12 in hand, the on-resin aza-Michael quaternization was effected with a solution of DMHA (10 equiv.) and HCl (5 equiv.) in PC (Scheme a) of Figure 1). Gratifyingly, significant conversion to the desired olefinic ammonium-modified peptide 15 was observed and the product was also stable to TFA mediated resin cleavage. Cleaving small portions of the resin beads at 1, 4 and 20 h timepoints revealed the greatest conversion (76%) to QAC-LYRAK 15 was achieved within 4 h, while at extended time points, further base-induced isomerisation to the thermodynamic isomer begans to impede overall conversion to QAC-LYRAK 15. Consequently, cleavage of peptidyl-resin after the 20 h incubation yielded QAC-LYRAK 15 in ca. 73% purity, which was purified by RP-HPLC to yield the pure peptide in 9% isolated yield.

[0110] Aza-Michael quaternisation in solution-phase on fully unprotected allenamidyl-LYRAK, was attempted, despite the presence of the potentially reactive primary amino side-chain of lysine. Taking resin-bound LYRAK 12, TFA-mediated resin cleavage was effected to afford fully unprotected allenmidyl-peptide 14. With the unprotected allenamidyl-LYRAK 14 in hand, TFA salts of basic side-chains were first neutralised with DMHA (2 equiv.), followed by addition of DMHA (10 equiv.) and HCI (5 equiv.) in PC. A chemoselective conversion (83%) towards QAC-LYRAK 15 was achieved within 18 h, with no evidence of self-cyclisation or polymerisation of allenamidyl-LYRAK 14 through the side-chain amine and allenamide handle. The reaction readily proceeded in the presence of tyrosine and arginine side-chains, functionalities that were challenging for our one-pot solution-phase protocol.

[0111] Furthermore, this aza-Michael quaternization was tolerant of the C-terminal carboxylic acid, whereas the one-pot procedure would likely be incompatible, specifically with regards to the coupling step.

[0112] Since this reaction no longer involved a one-pot coupling and aza-Michael addition, we speculated that the use of a protic solvent would significantly enhance the rate of reaction. Accordingly, the reaction was also carried out in 1: 1 HzO / MeCN (v / v) without addition of HCI. Remarkably, this drastically enhanced the reaction rate with 89% conversion towards QAC-LYRAK 15 attained within 1 h. Resin cleavage according to the general protocol and purification by RP-HPLC yielded peptide 15 as a white powder (1.1 mg, 9% yield, purity). RP-HPLC tR = 18.2 min, MS (ESI+): m / z [M]+observed, [] requires.

[0113] These two approaches to quaternisation on solid-phase and solution-phase were then attempted on allenamidyl LYRAK using TEA, a more hindered tertiary amine. As expected, the reaction on solid-phase was notably slower with TEA, giving reasonable conversion (63%) to TEA-LYRAK 16 after 36 h. However, when the solution-phase transformation was attempted in 1: 1 HzO / MeCN (v / v), a competing by-product was observed. Mass of this new by-product suggested that it may be a hydration product of allenamidyl LYRAK in the form of a 0-ketoamide (Supporting Info). The absence of this by-product with DMHA is likely owing to aza-Michael reaction possessing a significantly higher reaction rate due to reduced steric effects and thus outcompeting the hydration reaction. Therefore, the aza-Michael addition of hindered tertiary amines may be incompatible with reactions performed in water as the solvent.

[0114] Both curious and encouraged by the apparent selectivity of the addition towards tertiary amines over their primary amine counterparts, we wished to further explore this unprecedented reactivity. Allenamidyl-LYRAK 14 was initially incubated with Boc-Lys- OMe.HCI for 18 h in 1: 1 HzO / MeCN (v / v), but again no reactivity was observed between the primary amines and allenamide. To further validate this, a competitive reaction was established with the addition of 3 equiv. of DMHA to this mixture and incubated for a further 1 h. After only 10 min, the transformation to DMHA-LYRAK 15 was already evident (34% conversion), while after 1 h, DMHA-LYRAK 15 was exclusively afforded at 89% conversion. Therefore, this confirms the chemoselective nature of the aza-Michael addition of tertiary amines to the allenamide, thereby enabling late-stage diversification of peptides with quaternary ammonium moieties.

[0115] Example 8: Synthesis of further compounds of Formula IA that mimic common QACs

[0116] Ala: N-(4-(butylamino)-4-oxobut-l-en-2-yl)-N,N-dimethyldecan-l-aminium chloride & Alb: (E)-N-(4-(butylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyldecan-l-aminium chloride

[0117] A1a

[0118] To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (200 pL), sym-collidine (107 pL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (200 pL), followed by butylamine (40.0 pL, 0.405 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. A sample (2 pL) was taken for reaction monitoring via analytical RP-HPLC using a 5-95% B 3% B / min gradient where solvent A was H2O with 0.1% TFA and solvent B was MeCN with 0.1% TFA. N,N-dimethyldecylamine (1446 pL, 6.07 mmol), then HCI (5 M, 400 pL, 2.0 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 1 h. A sample (2 pL) was taken for reaction monitoring and the reaction mixture was agitated for another 2 h. Reaction completion was confirmed using analytical RP-HPLC. H2O (ca. 2.00 mL) was added to the reaction mixture then using semi-preparative RP-HPLC with a 5% B isometric flow, the excess reagents were separated from the crude product. Following return to baseline, the crude product was collected using a 95% B isometric flow. The collected fractions were diluted with water (ca. 10.0 mL), then lyophilised. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), and then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound A1a as a colourless gum (8.86 mg, 6.1% yield, >95% purity) and A1b as a colourless gum (5.30 mg, 3.7% yield, >95% purity). A1a (8.86 mg, yield, 6.1%) H1NMR (CDCl3, 400 MHz) δ 8.83 (1H, t, J = 5.4 Hz), 5.71 (1H, d, J = 3. 9 Hz), 5.53 (1H, d, J = 3.9 Hz), 4.07-3.98 (2H, m), 3.76 (2H, s), 3.54 (6H, s), 3.26-3.19 (2H, m), 1.59-1.47 (4H, m), 1.41-1.20 (16H, m), 0.90 (3H, t, J = 7.4 Hz), 0.87 (3H, t, J = 7.2 Hz); C13NMR (CDCl3, 101 MHz) δ 167.2, 145.4, 115.3, 64.8, 52.3, 39.9, 36.4, 31.8, 31.3, 29.5, 29.46, 29.4, 29.3, 26.1, 23.7, 22.7, 20.3, 14.1, 13.8; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C20H41N2O]+: 325.3213; found: 325.3211. A2a: N-(4-(octylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyldecan-1-aminium chloride A2a was prepared following the method described for preparation of compound A1a substituting octylamine for butylamine. To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (200 µL), followed by octylamine (67 µL, 0.405 mmol) was added. The reaction mixture was then agitated at r.t. for 3 h. The reaction was diluted with ethyl acetate (20 mL), washed sequentially with HCl (1M, 20 mL), saturated sodium bicarbonate (20 mL) and brine (20 mL) and then concentrated in vacuo. The residue was dissolved in MeCN (200 µL). N,N-dimethyldecylamine (953 µL, 4.0 mmol), then HCl (5 M, 400 µL, 2.0 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 3 h. The mixture was diluted with water. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm). The fractions were combined, diluted with a solution of HCl (0.01 M), and then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound A2a as a white powder (15.52 mg, Yield, 9.3%, >95% purity). H1NMR (CDCl3, 400 MHz) δ 8.88 (1H, t, J = 5.6 Hz), 5.70 (1H, d, J = 3. 6 Hz), 5.50 (1H, d, J = 3. 6 Hz), 4.16-4.02(2H, m), 3.77 (2H, s), 3.56 (6H, s), 3.21 (2H, q, J = 14.5, 6.0 Hz), 1.60-1.43 (4H, m), 1.34-1.20 (24H, m), 0.90-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 167.2, 145.6, 115.3, 64.8, 52.4, 40.4, 36.4, 32.0, 29.7, 29.6, 29.6, 29.5, 29.4, 27.3, 26.3, 23.9, 22.9, 14.3; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C24H49N2O]+: 381.3839; found: 381.3839. A3a: N-(4-(dodecylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyldecan-1-aminium chloride To a solution of 3-butynoic acid (68.1 mg, 0.80 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.80 mmol), then EEDQ (190 mg, 0.77 mmol) in MeCN (200 µL), followed by dodecylamine (74.0 mg, 0.40 mmol) was added. The reaction mixture was then agitated at r.t. for 5 h. The reaction was diluted with ethyl acetate (20 mL), washed sequentially with HCl (1M, 20 mL), saturated sodium bicarbonate (20 mL) and brine (20 mL) and then concentrated in vacuo. The residue was dissolved in MeCN (200 µL). N,N- dimethyldecylamine (953 µL, 4.0 mmol), then HCl (5 M, 400 µL, 2.0 mmol) was added to the mixture. The reaction mixture was then agitated at r.t. for 5 h. The mixture was diluted with water. Purification of the crude product was performed using semi- preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound A3a as a colourless gum (22 mg, 11.6% yield, >95% purity). H1NMR (CDCl3, 400 MHz) δ 8.10 (1H, brs), 5.67 (1H, d, J = 3. 9 Hz), 5.56 (1H, d, J = 3.9 Hz), 3.64-3.53 (4H, m), 3.34 (6H, s), 3.26-3.19 (2H, m), 1.59-1.47 (4H, m), 1.31-1.20 (32H, m), 0.87 (6H, t, J = 7.4 Hz); C13NMR (CDCl3, 101 MHz) δ 166.9, 145.5, 115.8, 66.2, 52.7, 40.3, 36.7, 32.0, 31.9, 29.71, 29.69, 29.6, 29.4, 29.37, 29.34, 29.26, 29.15, 29.12, 27.0, 26.0, 23.5, 22.8, 22.7, 14.2, 14.1; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C28H57N2O]+: 437.4465; found: 437.4463. B1a: N-(4-(decylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyloctan-1-aminium chloride & B1b: (E)-N-(4-(decylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyloctan-1-aminium chloride Compound B1a and B1b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (200 µL), followed by decylamine (80 µL, 0.405 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-Dimethyloctylamine (1249 µL, 6.07 mmol), then HCl (5 M, 400 µL, 2.02 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M) and then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound B1a as a colourless gum (23.9 mg, 14.4% yield, >95% purity) and B1b as a colourless gum (13.4 mg, 8.1% yield, >95% purity). Yield, 14.4%; H1NMR (CDCl3, 400 MHz) δ 8.77 (1H, brs), 5.67 (1H, s), 5.52 (1H, s), 4.08-3.93 (2H, m), 3.71 (2H, s), 3.34 (6H, s), 3.26-3.19 (2H, m), 1.59-1.47 (4H, m), 1.38-1.14 (24H, m), 0.88-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 167.2, 145.3, 115.3, 64.7, 52.4, 40.2, 36.3, 31.9, 31.7, 29.65, 29.63, 29.4, 29.1, 27.2, 26.1, 23.7,22.7, 22.6, 14.13, 14.06; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C24H49N2O]+: 381.3839; found: 381.3837. Yield, 8.1%; H1NMR (CDCl3, 400 MHz) δ 8.74 (1H, brs), 7.54 (1H, s), 3.62-3.47 (8H, m), 3.32-3.21 (2H, m), 2.47 (3H, s), 1.66-1.52 (4H, m), 1.69-1.54 (24H, m), 0.90-0.84 (6H, m); C13NMR (CDCl3, 101 MHz) δ 163.3, 147.9, 122.9, 65.3, 53.1, 39.9, 31.9, 31.6, 29.65, 29.63, 29.57, 29.37, 29.26, 29.1, 27.2, 26.3, 23.6, 22.7, 22.6, 14.14, 14.07, 12.28, ; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C24H49N2O]+: 381.3839; found: 381.3839. B2a: N-(4-(decylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyltetradecan-1- aminium chloride Compound B2a was prepared using the method described for the preparation of compound B1a, replacing N,N-Dimethyloctylamine with N,N-Dimethyltetradecylamine. To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (200 µL), followed by decylamine (80 µL, 0.405 mmol) was added. The reaction mixture was then agitated at r.t. for 3 h. The reaction was diluted with ethyl acetate (20 mL), washed sequentially with HCl (1M, 20 mL), saturated sodium bicarbonate (20 mL) and brine (20 mL) and then concentrated in vacuo. The residue was dissolved in MeCN (200 µL). N,N- Dimethyltetradecylamine (1819 µL, 6.0 mmol), then HCl (5 M, 400 µL, 2.0 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. The mixture was diluted with water. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm). The fractions were combined, diluted with a solution of HCl (0.01 M), and then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound B2a as a white powder (25.4 mg, 12.7% yield, >95% purity). Yield, 12.7%; H1NMR (CDCl3, 400 MHz) δ 8.90 (1H, brs), 5.70 (1H, brs), 5.50 (1H, brs), 4.12-3.93 (2H, m), 3.80 (2H, s), 3.53 (6H, s), 3.26-3.14 (2H, m), 1.61-1.47 (4H, m), 1.38-1.14 (36H, m), 0.88-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 167.1, 145.5, 115.2, 65.0, 52.6, 40.3, 36.4, 32.0, 29.8, 29.74, 29.70, 29.6, 29.5, 29.45, 29.42, 29.38, 27.2, 26.2, 23.8, 22.8, 14.2; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C30H61N2O]+: 465.4778; found: 465.4778. B3a: N-(4-(decylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyldecan-1-aminium chloride and B3b: (E)-N-(4-(decylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyldecan-1-aminium chloride Compound B3a and B3b was prepared using a method similar to that described for the preparation of compound A1a and A1b; To a solution of 3-butynoic acid (33.6 mg, 0.4 mmol) in MeCN (200 µL), sym-collidine (53 µL, 0.4 mmol), then EEDQ (94 mg, 0.38 mmol) in MeCN (200 µL), followed by decylamine (40.0 µL, 0.2 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-dimethyldecylamine (715 µL, 3.0 mmol), then HCl (5 M, 200 µL, 1.0 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M) and lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound B3a as a white powder (27.4 mg, 15.4% yield, >95% purity), and B3b as a white power (6.6 mg, 3.7% yield, >95% purity). Yield, 15.4%; H1NMR (CDCl3, 400 MHz) δ 8.90 (1H, brs), 5.67 (1H, d, J = 3. 1 Hz), 5.54 (1H, d, J = 3.9 Hz), 4.04-3.89 (2H, m), 3.71 (2H, s), 3.53 (6H, s), 3.19 (2H, t, J = 7.4 Hz), 1.57- 1.43 (4H, m), 1.38-1.14 (28H, m), 0.88-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 167.2, 145.2, 115.4, 64.7, 52.4, 40.3, 36.3, 31.93,31.87, 29.7, 29.629.53, 29.48, 29.39, 29.36, 29.3, 27.2, 26.1, 23.7, 22.70, 22.68, 14.13, 14.11; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C26H53N2O]+: 409.4152; found: 409.4154. Yield, 3.7%; H1NMR (CDCl3, 400 MHz) δ 8.65 (1H, brs), 7.41 (1H, s), 3.65-3.47 (8H, m), 3.30-3.18 (2H, m), 2.46 (3H, s), 1.65-1.43 (4H, m), 1.38-1.14 (28H, m), 0.88-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 163.3, 147.5, 123.4, 65.5, 53.1, 40.0, 32.0, 31.9, 29.8, 29.7, 29.6, 29.5, 29.42, 29.39, 29.2, 19.1, 27.2, 26.3, 23.6, 22.75. 22.70,14.17, 14.15; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C26H53N2O]+: 409.4152; found: 409.4152. B4a: N-(4-(decylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyldodecan-1-aminium chloride and B4b: (E)-N-(4-(decylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyldodecan-1-aminium chloride Compound B4a and B4b was prepared using a method similar to that described for the preparation of compound A3a; To a solution of 3-butynoic acid (68.1 mg, 0.80 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.80 mmol), then EEDQ (190 mg, 0.77 mmol) in MeCN (200 µL), followed by decylamine (62.8 mg, 0.40 mmol) was added. The reaction mixture was then agitated at r.t. for 5 h. The reaction was diluted with ethyl acetate (20 mL), washed sequentially with HCl (1M, 20 mL), saturated sodium bicarbonate (20 mL) and brine (20 mL) and then concentrated in vacuo. The residue was dissolved in MeCN (200 µL). N,N-dimethyldodecylamine (852 mg, 4.0 mmol), then HCl (5 M, 400 µL, 2.0 mmol) was added to the mixture. The reaction mixture was then agitated at r.t. for 5 h. The mixture was diluted with water. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound B4a as a colourless gum (6 mg, 3.2% yield, >95% purity) and B4b as a colourless gum (17.5 mg, 9.3% yield, >95% purity). Yield, 3.2; H1NMR (CDCl3, 400 MHz) δ 8.86 (1H, brs), 5.70 (1H, s), 5.51 (1H, s), 4.10-4.98 (2H, m), 3.77 (2H, s), 3.54 (6H, s), 3.21 (2H, m), 1.65-1.43 (4H, m), 1.38-1.14 (32H, m), 0.88-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 167.1, 145.4, 115.3, 64.9, 52.5, 40.3, 36.4, 32.0, 29.7, 29.6, 29.5, 29.45, 29.40, 29.36, 27.2, 26.2, 23.8, 22.8, 14.2; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C28H57N2O]+: 437.4465; found: 437.4462. Yield, 9.3%; H1NMR (CDCl3, 400 MHz) δ 8.59 (1H, brs), 7.28 (1H, brs), 3.65-3.46 (8H, m), 3.24 (2H, brs), 2.45 (3H, s), 1.64-1.47 (4H, m), 1.38-1.14 (32H, m), 0.88-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 163.3, 147.9, 122.9, 65.3, 53.1, 40.0, 31.9, 29.74, 29.66. 29.6, 29.5, 29.4, 29.37, 29.3, 29.2, 27.2, 26.3, 23.6, 22.7, 14.1, 12.3; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C28H57N2O]+: 437.4465; found: 437.4465. B5a: N-(4-(butylamino)-4-oxobut-1-en-2-yl)-N,N-dimethylhexan-1-aminium chloride and B5b: (E)-N-(4-(butylamino)-4-oxobut-2-en-2-yl)-N,N- dimethylhexan-1-aminium chloride Compound B5a and B5b was prepared using a method similar to that described for the preparation of compound A1a; To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (200 µL), followed by butylamine (40.0 µL, 0.405 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-dimethylhexylamine (784 µL, 6.07 mmol), then HCl (5 M, 400 µL, 2.02 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound B5a as a colourless gum (7.78 mg, 6.4% yield, >95% purity) and B5b as a colourless gum (2.31 mg, 1.9% yield, >95% purity). Yield, 6.4%; H1NMR (CDCl3, 400 MHz) δ 8.83 (1H, t, J = 5.2 Hz), 5.70 (1H, d, J = 3.4 Hz), 5.51 (1H, d, J = 3.4 Hz), 4.10-4.98 (2H, m), 3.74 (2H, s), 3.53 (6H, s), 3.26-3.27 (2H, m), 1.58- 1.43 (4H, m), 1.38-1.14 (8H, m), 0.92-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 167.1, 145.4, 115.3, 64.8, 52.3, 39.9, 36.3, 31.4, 31.3, 25.7, 23.6, 22.4, 20.3, 13.9, 13.8; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C16H33N2O]+: 269.2587; found: 269.2586. Yield, 1.9%; H1NMR (CDCl3, 400 MHz) δ 8.65 (1H, brs), 7.37 (1H, s), 3.65-3.46 (8H, m), 3.28 (2H, t, J = 6.1 Hz), 2.46 (3H, s), 1.64-1.47 (4H, m), 1.38-1.14 (8H, m), 0.88-0.81 (6H, m); C13NMR (CDCl3, 101 MHz) δ 163.4, 147.6, 123.2, 65.5, 53.2, 39.6, 31.2, 31.18, 25.9, 23.5, 22.4, 20.3, 13.9, 13.8, 12.3; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C16H33N2O]+: 269.2587; found: 269.2586. C1a: N-(4-(benzylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyltetradecan-1- aminium chloride and C1b: (E)-N-(4-(benzylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyltetradecan-1-aminium chloride Compound C1a and C1b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (61.6 mg, 0.73 mmol) in MeCN (200 µL), sym-collidine (97 µL, 0.73 mmol), then EEDQ (172.1 mg, 0.69 mmol) in MeCN (200 µL), followed by benzylamine (40.0 µL, 0.37 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-dimethyltetradecylamine (1668 µL, 5.5 mmol), then HCl (5 M, 360 µL, 1.8 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound C1a as a colourless gum (15.0 mg, 8.3% yield, >95% purity) and C1b as a colourless gum (2.39 mg, 1.3% yield, >95% purity). Yield, 8.3%; H1NMR (CDCl3, 400 MHz) δ 9.41 (1H, s), 7.36-7.30 (2H, m), 7.25-7.15 (3H, m), 5.66 (1H, d, J = 2.5 Hz), 5.50 (1H, d, J = 2.5 Hz), 4.38 (2H, d, J = 5.3 Hz), 3.99-3.90 (2H, m), 3.77 (2H, s), 3.43 (6H, s), 1.51-1.38 (2H, m), 1.30-1.17 (22H, m), 0.86 (3H, t, J = 6.9 Hz); C13NMR (CDCl3, 101 MHz) δ 167.3, 145.2, 138.5, 128.5, 128.1, 127.2, 115.5, 64.8, 52.3, 43.8, 36.4, 32.0, 29.74, 29.70, 29.65, 29.58, 29.43, 29.40, 26.1, 23.7, 22.7, 14.2; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C27H47N2O]+: 415.3683; found: 415.3682. Yield, 1.3%; H1NMR (CDCl3, 400 MHz) δ 9.28 (1H, s), 7.58 (1H, s), 7.40-7.34 (2H, m), 7.32-7.26 (2H, m), 7.24-7.18 (1H, m), 4.47 (2H, d, J = 5.5 Hz), 3.58-3.44 (8H, m), 2.45 (3H, s), 1.66-1.52 (2H, m), 1.31-1.17 (22H, m), 0.88 (3H, t, J = 6.8 Hz); C13NMR (CDCl3, 101 MHz) δ 163.7, 148.0, 138.7, 129.0, 128.5, 127.6, 124.0, 66.2, 53.6, 44.0, 32.4, 30.2, 30.14, 30.12, 30.0, 29.9, 29.8, 29.5, 26.7, 24.0 23.2, 14.6, 12.8; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for C27H47N2O: 415.3683; found: 415.3681. C2a: N-(4-(benzylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyloctan-1-aminium chloride and C2b: (E)-N-(4-(benzylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyloctan-1-aminium Compound C2a and C2b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (61.6 mg, 0.73 mmol) in MeCN (200 µL), sym-collidine (96.8 µL, 0.73 mmol), then EEDQ (172 mg, 0.69 mmol) in MeCN (200 µL), followed by benzylamine (40.0 µL, 0.36 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-dimethyloctylamine (1129 µL, 5.5 mmol), then HCl (5 M, 360 µL, 1.8 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound C2a as a colourless gum (8.41 mg, 5.7% yield, >95% purity) and C2b as a colourless gum (5.96 mg, 4.1% yield, >95% purity). Yield, 5.7%; H1NMR (CDCl3, 400 MHz) δ 9.32 (1H, t, J = 5.5 Hz), 7.36-7.30 (2H, m), 7.25-7.15 (3H, m), 5.66 (1H, d, J = 2.5 Hz), 5.50 (1H, d, J = 2.5 Hz), 4.39 (2H, d, J = 5.3 Hz), 3.96- 3.90 (2H, m), 3.78 (2H, s), 3.41 (6H, s), 1.53-1.38 (2H, m), 1.30-1.17 (10H, m), 0.86 (3H, t, J = 6.9 Hz); C13NMR (CDCl3, 101 MHz) δ 167.3, 145.2, 138.4, 128.5, 128.1, 127.2, 115.6, 65.1, 52.4, 43.8, 36.5, 31.7, 29.3, 29.0, 26.0, 23.7, 22.6, 14.1; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C21H35N2O]+: 331.2744; found: 331.2742. Yield, 4.1%; H1NMR (CDCl3, 400 MHz) δ 9.26 (1H, s), 7.54 (1H, s), 7.40-7.33 (2H, m), 7.32-7.26 (2H, m), 7.24-7.18 (1H, m), 4.47 (2H, s), 3.68-3.36 (8H, m), 2.45 (3H, s), 1.62-1.50 (2H, m), 1.33-1.18 (10H, m), 0.87 (3H, t, J = 6.7 Hz); C13NMR (CDCl3, 101 MHz) δ 163.4, 147.9, 138.4, 128.7, 128.2, 127.4, 123.6, 66.0, 53.2, 32.1, 29.9, 29.8, 29.7, 29.6, 29.6, 29.5 26.4, 23.7, 22.9, 14.3, 12.5; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C21H35N2O]+: 331.2744; found: 331.2742. C3a: N-(4-(benzylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyldecan-1-aminium chloride and C3b: (E)-N-(4-(benzylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyldecan-1-aminium chloride Compound C3a and C3b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (200 µL), followed by benzylamine (44.0 µL, 0.405 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-dimethyldecylamine (1446 µL, 6.07 mmol), then HCl (5 M, 400 µL, 2.02 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound C3a as a colourless gum (8.86 mg, 5.6% yield, >95% purity) and C3b as a colourless gum (2.2 mg, 1.4% yield, >95% purity). Yield, 5.6%; H1NMR (CDCl3, 400 MHz) δ 9.32 (1H, t, J = 5.6 Hz), 7.36-7.25 (4H, m), 7.23-7.15 (1H, m), 5.63 (1H, d, J = 3.5 Hz), 5.50 (1H, d, J = 3.5 Hz), 4.39 (2H, d, J = 5.9 Hz), 3.65 (2H, s), 3.64-3.57 (2H, m), 3.27 (6H, s), 1.57-1.44 (2H, m), 1.30-1.17 (14H, m), 0.87 (3H, t, J = 6.9 Hz); C13NMR (CDCl3, 101 MHz) δ 167.6, 145.0, 138.4, 128.5, 128.0, 127.2, 115.8, 65.1, 52.6, 43.7, 36.5, 31.9, 29.5, 29.45, 29.36, 29.3, 26.1, 23.6, 22.7, 14.1; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C23H39N2O]+: 359.3057; found: 359.3056. Yield, 1.4%; H1NMR (CDCl3, 400 MHz) δ 9.25 (1H, t, J = 5.8 Hz), 7.36-7.19 (5H, m), 7.10 (1H, s), 4.45 (2H, d, J = 6.1 Hz), 3.51-3.41 (2H, m), 3.37 (6H, s), 2.44 (3H, s), 1.61-1.47 (2H, m), 1.30-1.17 (14H, m), 0.88 (3H, t, J = 6.9 Hz); C13NMR (CDCl3, 101 MHz) δ 163.2, 148.4, 138.3, 128.6, 128.0, 127.2, 122.8, 65.6, 52.5, 43.6, 31.9, 29.40, 29.38, 29.2, 29.0, 26.2, 23.4, 22.7, 14.2, 12.4; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C23H39N2O]+: 359.3057; found: C4a: N-(4-(benzylamino)-4-oxobut-1-en-2-yl)-N,N-dimethyldodecan-1- aminium chloride and C4b:(E)-N-(4-(benzylamino)-4-oxobut-2-en-2-yl)-N,N- dimethyldodecan-1-aminium chloride Compound C4a and C4b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (61.6 mg, 0.73 mmol) in MeCN (200 µL), sym-collidine (96.8 µL, 0.73 mmol), then EEDQ (172 mg, 0.69 mmol) in MeCN (200 µL), followed by benzylamine (40.0 µL, 0.36 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-dimethyldodecylamine (1129 µL, 5.5 mmol), then HCl (5 M, 360 µL, 1.8 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound C4a as a colourless gum (16.99 mg, 10.1% yield, >95% purity) and C4b as a colourless gum (2.93 mg, 1.7% yield, >95% purity). Yield, 10.1%; H1NMR (CDCl3, 400 MHz) δ 9.38 (1H, s), 7.36-7.30 (2H, m), 7.25-7.15 (3H, m), 5.66 (1H, d, J = 2.5 Hz), 5.50 (1H, d, J = 2.5 Hz), 4.39 (2H, d, J = 5.3 Hz), 3.96-3.90 (2H, m), 3.77 (2H, s), 3.44 (6H, s), 1.52-1.39 (2H, m), 1.30-1.17 (18H, m), 0.86 (3H, t, J = 6.9 Hz); C13NMR (CDCl3, 101 MHz) δ 167.3, 145.2, 138.4, 128.5, 128.0, 127.2, 115.6, 64.9, 52.3, 43.8, 36.4, 31.9, 29.64, 29.57, 29.42, 29.38, 26.1, 23.7, 22.7, 14.2; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C24H43N2O]+: 387.3370; found: 387.3369. Yield, 1.7%; H1NMR (CDCl3, 400 MHz) δ 9.21 (1H, s), 7.47 (1H, s), 7.36-7.30 (2H, m), 7.25-7.15 (3H, m), 4.46 (2H, d, J = 3.9 Hz), 3.61-3.38 (8H, m), 2.44 (3H, s), 1.61-1.49 (2H, m), 1.36-1.17 (18H, m), 0.88 (3H, t, J = 6.9 Hz); C13NMR (CDCl3, 101 MHz) δ 167.4, 148.0, 138.2, 128.6, 128.1, 127.2, 123.2, 65.6, 53.2, 43.5, 32.0, 29.64, 29.61, 29.5, 29.4, 29.38, 29.1, 26.3, 23.6, 22.8, 14.2, 12.4; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C24H43N2O]+: 387.3370; found: C5a: N-(4-(benzylamino)-4-oxobut-1-en-2-yl)-N,N-dimethylhexadecan-1- aminium chloride and C5b: (E)-N-(4-(benzylamino)-4-oxobut-2-en-2-yl)-N,N- dimethylhexadecan-1-aminium chloride Compound C5a and C5b was prepared using the method described for the preparation of compound C4a and C4b, replacing N,N-dimethyldodecylamine with N,N- Dimethylhexadecylamine. To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (200 µL), sym-collidine (107 µL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (200 µL), followed by benzylamine (44.0 µL, 0.405 mmol) was added. The reaction mixture was then agitated at r.t. for 3 h. The reaction was diluted with ethyl acetate (20 mL), washed sequentially with HCl (1M, 20 mL), saturated sodium bicarbonate (20 mL) and brine (20 mL) and then concentrated in vacuo. The residue was dissolved in MeCN (200 µL). N,N-Dimethylhexadecylamine (2019 µL, 6.0 mmol), then HCl (5 M, 400 µL, 2.0 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 72 h. The mixture was diluted with water. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm). The fractions were combined, diluted with a solution of HCl (0.01 M), and then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound C5a as a colourless gum (17.32 mg, 9.1% yield, >95% purity) and C5b as a colourless gum (9.99 mg, 5.2% yield, >95% purity). Yield, 9.1%; H1NMR (CDCl3, 400 MHz) δ 9.55 (1H, t, J = 5.92 Hz), 7.40-7.34 (2H, m), 7.29-7.26 (1H, m) 7.26-7.23 (1H, m), 7.22-7.16 (1H, m), 5.67 (1H, d, J = 3.7 Hz), 5.49 (1H, d, J = 3.7 Hz), 4.41 (2H, d, J = 6.0 Hz), 3.94-3.14 (4H, m), 3.39 (6H, s), 1.55-1.45 (2H, m), 1.31-1.17 (26H, m), 0.88 (3H, t, J = 6.8 Hz); C13NMR (CDCl3, 101 MHz) δ 167.3, 145.6, 138.7, 128.6, 128.3, 127.3, 115.6, 65.6, 52.6, 44.0, 36.7, 32.1, 29.9, 29.9, 29.8, 29.7, 29.6, 29.5, 29.5, 26.1, 23.9, 22.9, 14.3; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C29H51N2O]+: 443.3996; found: 443.3994. Yield, 5.2%; H1NMR (CDCl3, 400 MHz) δ 9.23 (1H, s), 7.48 (1H, s), 7.38-7.32 (2H, m), 7.32-7.26 (2H, m), 7.24-7.18 (1H, m), 4.47 (2H, s), 3.67-3.33 (8H, m), 2.45 (3H, s), 1.67-1.49 (2H, m), 1.27-1.21 (26H, m), 0.88 (3H, t, J = 6.8 Hz); C13NMR (CDCl3, 101 MHz) δ 163.4, 147.9, 138.4, 128.7, 128.2, 127.4, 123.6, 66.0, 53.2, 32.1, 29.9, 29.8, 29.7, 29.6, 29.6, 29.5 26.4, 23.7, 22.9, 14.3, 12.5; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C29H51N2O]+: 443.3996; found: 443.3994. D1a: (S)-N-(4-((1-(tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut- 1-en-2-yl)-N,N-dimethylhexan-1-aminium chloride and D1b:(S,E)-N-(4-((1- (tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut-2-en-2-yl)-N,N- dimethylhexan-1-aminium chloride Compound D1a and D1b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (52.2 mg, 0.62 mmol) in MeCN (200 µL), sym-collidine (82 µL, 0.62 mmol), then EEDQ (146 mg, 0.59 mmol) in MeCN (200 µL), followed by L-phenylalanine tert-butyl ester (80 mg, 0.31 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-Dimethylhexylamine (1055 µL, 6.07 mmol), then HCl (5 M, 156 µL, 0.78 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound D1a as a colourless gum (10.23 mg, 5.7% yield, >95% purity) and D1b as a colourless gum (2.4 mg, 1.3% yield, >95% purity). Yield, 5.7%; [α]D20+ 6.2 (c 0.24, CH2Cl2); H1NMR (CDCl3, 400 MHz) δ 9.07 (1H, s), 7.35-7.30 (1H, m), 7.27-7.15 (4H, m), 5.53 (1H, d, J = 3.6 Hz), 5.47 (1H, d, J = 3.6 Hz), 4.59-4.51 (1H, m), 3.94 (1H, d, J = 14.9 Hz), 3.92-3.79 (2H, m), 3.48 (1H, d, J = 14.9 Hz), 3.44 (3H, s), 3.36 (3H, s), 3.19-3.02 (2H, m), 1.58-1.46 (2H, m), 1.35 (9H, s), 1.32-1.21 (6H, m), 0.85 (3H, t, J = 7.1 Hz); C13NMR (CDCl3, 101 MHz) δ 170.7, 167.6, 145.3, 137.3, 129.6, 128.4, 126.7, 115.7, 81.7, 65.5, 55.5, 53.1, 52.1, 37.4, 36.7, 31.4, 28.0, 25.7, 23.6, 22.4, 13.9; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C25H41N2O3]+: 417.3112; found:. Yield, 1.3%; H1NMR (CDCl3, 400 MHz) δ 8.88 (1H, d, J = 6.1), 7.33-7.28 (3H, m), 7.27-7.25 (2H, m), 7.21-7.15 (1H, m), 4.67-4.56 (1H, m), 3.64-3.48 (8H, m), 3.16-3.06 (2H, m), 2.36 (3H, s), 1.63-1.50 (2H, m), 1.34 (9H, s), 1.32-1.21 (6H, m), 0.88 (3H, t, J = 7.1 Hz); C13NMR (CDCl3, 101 MHz) δ 170.4, 163.3, 148.3, 137.0, 129.6, 128.3, 126.7, 122.6, 81.7, 65.6, 55.3, 53.0, 38.0, 31.2, 28.0, 25.9, 23.5, 22.5, 13.9, 12.6; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C25H41N2O3]+: 417.3112; found: 417.3110. D2a: (S)-N-(4-((1-(tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut- 1-en-2-yl)-N,N-dimethyldecan-1-aminium chloride and D2b:(S,E)-N-(4-((1- (tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut-2-en-2-yl)-N,N- dimethyldecan-1-aminium chloride Compound D2a and D2b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (52.2 mg, 0.62 mmol) in MeCN (200 µL), sym-collidine (82 µL, 0.62 mmol), then EEDQ (146 mg, 0.59 mmol) in MeCN (200 µL), followed by L-phenylalanine tert-butyl ester (80 mg, 0.31 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N-dimethyldecylamine (1446 µL, 6.07 mmol), then HCl (5 M, 156 µL, 0.78 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound D2a as a colourless gum (7.5 mg, 4.8% yield, >95% purity) and D2b as a colourless gum (21.1 mg, 13.4% yield, >95% purity). Yield, 4.8%; [α]D20-21.3 (c 0.18, CH2Cl2); H1NMR (CDCl3, 400 MHz) δ 9.36 (1H, d, J = 7.2 Hz), 7.41- 7.33 (2H, m), 7.30-7.22 (2H, m), 7.20-7.14 (1H, m), 5.54 (1H, d, J = 3.6 Hz), 5.44 (1H, d, J = 3.6 Hz), 4.58-4.47 (1H, m), 4.08 (1H, d, J = 15.9 Hz), 4.03-3.89 (2H, m), 3.48 (1H, d, J = 15.9 Hz), 3.50 (3H, s), 3.39 (3H, s), 3.19-3.02 (2H, m), 1.58-1.46 (2H, m), 1.36 (9H, s), 1.32-1.21 (14H, m), 0.87 (3H, t, J = 7.1 Hz); C13NMR (CDCl3, 101 MHz) δ 170.7, 167.7, 145.4, 137.5, 129.7, 128.4, 126.6, 115.5, 81.6, 65.4, 55.7, 53.3, 52.0, 37.4, 36.7, 31.9, 29.6, 29.5, 29.4, 29.3, 28.0, 26.1, 23.9, 22.7, 14.2; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C29H49N2O3]+, 473.3738; found: 473.3738. Yield, 13.4%; [α]D20-2.3 (c 0.10, CH2Cl2); H1NMR (CDCl3, 400 MHz) δ 8.68 (1H, d, J = 7.1), 7.27-7.15 (5H, m), 7.05 (1H, s), 4.67-4.56 (1H, m), 3.61-3.44 (8H, m), 3.15-2.99 (2H, m), 2.32 (3H, s), 1.63-1.50 (2H, m), 1.33 (9H, s), 1.32-1.21 (14H, m), 0.86 (3H, t, J = 7.1 Hz); C13NMR (CDCl3, 101 MHz) δ 170.7, 163.3, 148.8, 136.9, 129.5, 128.3, 126.7, 121.8, 81.9, 65.3, 55.1, 52.8, 52.6, 37.9, 31.9, 29.4, 29.2, 29.1, 27.9, 26.2, 23.5, 22.7, 14.1, 12.5; HRMS (ESI / Q-TOF) m / z [M]+calc. for [C29H49N2O3]+, 473.3738; found: 473.3738. D3a: (S)-N-(4-((1-(tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut- 1-en-2-yl)-N,N-dimethyltetradecan-1-aminium chloride and D3b: (S,E)-N-(4- ((1-(tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut-2-en-2-yl)-N,N- dimethyltetradecan-1-aminium chloride Compound D3a and D3b was prepared using a method similar to that described for the preparation of compound A1; To a solution of 3-butynoic acid (52.2 mg, 0.62 mmol) in MeCN (200 µL), sym-collidine (82 µL, 0.62 mmol), then EEDQ (146 mg, 0.59 mmol) in MeCN (200 µL), followed by L-phenylalanine tert-butyl ester (80 mg, 0.31 mmol) was added. The reaction mixture was then agitated at r.t. for 1 h. N,N- Dimethyltetradecylamine (1841 µL, 6.07 mmol), then HCl (5 M, 156 µL, 0.78 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 5 h. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm) with a linear 30-95% B 1% B / min gradient. The fractions were combined, diluted with a solution of HCl (0.01 M), then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound D3a as a colourless gum (26.87 mg, 11.6% yield, >95% purity) and D3b as a colourless gum (4.2 mg, 1.8% yield, , >95% purity). Yield, 11.6%; [α]D20-13.1 (c 0.08, CH2Cl2); H1NMR (CDCl3, 400 MHz) δ 9.12 (1H, d, J = 6.88 Hz), 7.37-7.32 (2H, m), 7.28-7.22 (2H, m), 7.20-7.14 (1H, m), 5.54 (1H, d, J = 3.6 Hz), 5.45 (1H, d, J = 3.6 Hz), 4.59-4.51 (1H, m), 3.98 (1H, d, J = 15.8 Hz), 3.92-3.71 (2H, m), 3.52 (1H, d, J = 15.8 Hz), 3.41 (3H, s), 3.30 (3H, s), 3.22-3.03 (2H, m), 1.59-1.47 (2H, m), 1.37 (9H, s), 1.35-1.20 (22H, m), 0.88 (3H, t, J = 6.8 Hz); C13NMR (CDCl3, 101 MHz) δ 170.9, 167.9, 145.9, 137.8, 128.8, 127.0, 116.0, 82.0, 66.4, 55.9, 53.6, 52.7, 37.8, 37.2, 32.4, 30.2, 30.12, 30.06, 30.0, 29.84, 29.82, 29.77, 28.4, 26.4, 24.2, 23.2, 14.6; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C33H57N2O3]+: 529.4364; found: 529.4361. Yield, 1.8%; H1NMR (CDCl3, 400 MHz) δ 8.86 (1H, d, J = 7.4), 7.30-7.26 (2H, m), 7.25-7.21 (1H, m), 7.21-7.09 (2H, m), 4.66-4.57 (1H, m), 3.64-3.40 (9H, m), 3.16-3.00 (2H, m), 2.34 (3H, s), 1.62-1.45 (2H, m), 1.34 (9H, s), 1.32-1.21 (22H, m), 0.87 (3H, t, J = 6.8 Hz); C13NMR (CDCl3, 101 MHz) δ 170.9, 163.7, 149.0, 137.5, 129.9, 128.7, 127.1, 122.7, 82.2, 65.9, 55.6, 53.1, 53.0, 38.3, 32.4, 30.12, 30.10, 30.01, 29.89, 29.84, 29.82, 29.5, 28.3, 26.6, 23.9, 23.2, 14.6, 12.9; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C33H57N2O3]+: 529.4364; found: 529.4363. D4a: (S)-N-(4-((1-(tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut- 1-en-2-yl)-N,N-dimethylhexadecan-1-aminium chloride and D2b: (S,E)-N-(4- ((1-(tert-butoxy)-1-oxo-3-phenylpropan-2-yl)amino)-4-oxobut-2-en-2-yl)-N,N- dimethylhexadecan-1-aminium chloride Compound D4a was prepared using the method described for the preparation of compound D3a and D3b, replacing N,N-Dimethyltetradecylamine with N,N- Dimethylhexadecylamine. To a solution of 3-butynoic acid (68.1 mg, 0.809 mmol) in MeCN (100 µL), sym-collidine (107 µL, 0.809 mmol), then EEDQ (190 mg, 0.769 mmol) in MeCN (100 µL), followed by L-phenylalanine tert-butyl ester (103 mg, 0.4 mmol) in 1:1 MeCN:H2O (400 µL) was added. The reaction mixture was then agitated at r.t. for 3 h. The reaction was diluted with ethyl acetate (20 mL), washed sequentially with HCl (1M, 20 mL), saturated sodium bicarbonate (20 mL) and brine (20 mL) and then concentrated in vacuo. The residue was dissolved in MeCN (200 µL). N,N- Dimethylhexadecylamine (2019 µL, 6.0 mmol), then HCl (5 M, 400 µL, 2.0 mmol) was added to the reaction mixture. The reaction mixture was then agitated at r.t. for 24 h. The mixture was diluted with water. Purification of the crude product was performed using semi-preparative RP-HPLC using a Phenomenex Luna C18(2) column (100 Å; 5 µm; 10 × 250 mm). The fractions were combined, diluted with a solution of HCl (0.01 M), and then lyophilised. The dry samples were dissolved in a solution of HCl (0.01 M, ca. 3.00 mL) and lyophilised two more times to afford compound D4a as a colourless gum (0.95 mg, >95% purity) and D4b as a colourless gum (0.42 mg, >95% purity). D4a (0.95 mg) H1NMR (CDCl3, 400 MHz) δ 9.21 (1H, d, J = 7.56 Hz), 7.34-7.30 (2H, m), 7.25-7.22 (2H, m), 7.20-7.14 (1H, m), 5.54 (1H, d, J = 3.9 Hz), 5.44 (1H, d, J = 3.9 Hz), 4.59- 4.52 (1H, m), 3.92 (1H, d, J = 16.16 Hz), 3.81-3.63 (2H, m), 3.53 (1H, d, J = 16.16 Hz), 3.35 (3H, s), 3.26 (3H, s), 3.21-3.00 (2H, m), 1.61-1.43 (2H, m), 1.38 (9H, s), 1.29-1.19 (26H, m), 0.88 (3H, t, J = 6.8 Hz); C13NMR (CDCl3, 101 MHz) δ 170.9, 167.9, 146.0, 137.9, 128.7, 127.0, 116.0, 82.0, 66.5, 55.9, 53.5, 52.7, 37.8, 37.2, 32.4, 30.18, 30.16, 30.12, 30.06, 29.95, 29.85, 29.81, 29.70 28.4, 26.4, 24.1, 23.2, 14.6; HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C35H61N2O3]+: 557.4677; found: 557.4674. D4b (0.42 mg) HRMS (ESI / Orbitrap MS) m / z [M]+calc. for [C35H61N2O3]+: 557.4677; found: 557.4675. Example 9: Comparison of anti-microbial activity of compounds of the invention against common QACs A range of compounds of the invention were tested for anti-microbial activity against BAC, DDAC and BDMTDAC, shown below. Bacterial strains were grown in cation adjusted Mueller Hinton (MH) broth at 37 °C with shaking (200 rpm). MIC assays were performed in accordance with the CLSI recommended protocol (Performance standards for antimicrobial susceptibility testing; sixteenth informational supplement, 2023). Briefly, a two-fold dilution series of the test compounds (from 64 μM to 0.125 μM, final) was prepared in triplicate in polypropylene 96-well plates, using cation adjusted MH media. Cultures of bacteria were diluted accordingly in fresh media before adding 50 μL of inoculum to each well of the MIC plate, to achieve a final volume of 100 μL with a uniform CFU / ml of ~5 x105in each well. A growth control (untreated) and sterility control (non-inoculated) well was included for each test compound replicate. Plates were incubated at 37°C with shaking (with the exception of Enterococcus which remained static) for 18 h before determining the MIC. MICs were determined as the lowest concentration at which growth did not occur. The results are set out in Table 2. Table 2: Minimum inhibitory concentrations (MIC) values of OQA and QAC compounds towards E. Coli and S. aureus A second set of tests was carried out under the same conditions as set out above. In addition to the first set of antibacterial assays, an antifungal assay was performed. The antifungal activity of the compounds towards C. albicans was evaluated in accordance with the CLSI recommended protocols (National committee for clinical laboratory standard, 2002, reference method for broth dilution antifungal susceptibility testing of yeasts. approved standard-second edition m27-a2, vol. 22 no. 15. National committee for clinical laboratory standards, Wayne, PA. 2002). Briefly, RPMI 1640 media (with glutamine and phenol red, without bicarbonate) was used to prepare a triplicate two-fold dilution series of the test compounds (from 64 mM to 0.125 mM, final) in polypropylene 96-well plates. Five colonies (diameter of around 1 mm) of C. albicans were selected from 24h-old-culture and resuspended in 0.85% saline for preparing the inoculum. After vortex mixing, the suspension was diluted to provide a solution with absorbance equivalent to that of a 0.5 McFarland standard, approx. 0.1 OD at 530 nm (using 1 cm path length cuvette). This suspension was diluted twice further, once at a ratio of 1:50 and at a ratio of 1:20 using RPMI 1640 media, to achieve a final inoculum of 0.5 x 103to 2.5 x 103CFU / mL. The antifungal assay was performed by adding 50 μL of diluted C. albicans to 50 μL of test compound solution. The MIC was determined as the lowest test concentration at which no turbidity was seen in any of the three duplicates after the plates were incubated at 35 °C for 48 hours. The assay was carried out three times independently, and the lowest concentration at which agreement was observed across all three biological replicates was the established minimum inhibitory concentration (MIC). The results are set out in Table 3. Table 3: Minimum inhibitory concentration (MIC) values of OQA and QAC compounds against S. aureus ATCC 29213, E. coli ATCC 25922 and C. albicans

[0119] Example 10: Preparation of OQA analogues of polymyxin

[0120] OQA analogues of polymyxin were prepared using the following methodology:

[0121] Allyl (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-aminobutanoate (Fmoc-Dab-O-allyl)1

[0122] To a solution of Fmoc-Dab(Boc)-OH (1 g, 2.25 mmol) dispersed in water (5 mL) containing sodium bicarbonate (0.19 g, 2.25 mmol, 1 equiv.) was added dropwise a solution of Aliquat 336 (0.8 g, 1.9 mmol, 0.85 equiv.) and allyl bromide (1.02 mL, 11.9 mmol, 5.3 equiv.) dissolved in CH2CI2 (4 mL) with stirring. The reaction was kept overnight stirring at rt before dilution with water (15 mL). The aqueous phase was extracted with CH2CI2 (3 x 25 mL). The combined organic layers were washed with saturated brine (2 x 25 mL), filtered and concentrated in vacuo. The crude product was dissolved in a solution of TFA / CH2CI2 (20% v / v, 20 mL) and stirred for 2 h at rt. The reaction mixture was concentrated under a gentle stream of N2. The crude product was redissolved in FW / MeCN (1 : 1, v / v, 20 mL), lyophilised and used without further purification. (3 / ?)-2-(((allyloxy)carbonyl)amino)-3-(tert-butoxy)butanoic acid (Alloc- Thr(tBu)-OH)

[0123] 1. allyl chloroformate, aq. NaHCO3, THF / water (3:1 , v / v) 0°C, 5 min

[0124] 2. rt, 20 h

[0125] To a solution of H-Thr(tBu)-OH (1 g, 5.71mmol) dispersed in THF / water (3: 1, v / v, 50 mL) containing sodium bicarbonate (0.96 g, 11.4 mmol, 2 equiv.) at 0°C was added allyl bromide (0.73 mL, 6.3 mmol, 1.1 equiv.) with stirring for 5 min. The reaction mixture warmed to rt and was kept stirring overnight. Excess sodium bicarbonate was added, and the reaction mixture was stirred for a further 30 min. The reaction mixture was concentrated in vacuo and water (50 mL) was added. The aqueous solution was acidified to pH 2 using hydrochloric acid (1 M, aq.) and was extracted with CH2CI2 (3 x 50 mL). The combined organic layers were washed with saturated brine (2 x 50 mL), filtered and concentrated in vacuo. The crude product was redissolved in H2O / MeCN (1 : 1, v / v, 20 mL), lyophilised and used without further purification.

[0126] General method 1: Coupling of the first amino acid residue to 2-CTC resin

[0127] To 2-CTC resin (350 mg, 0.13 mmol, 0.38 mmol / g loading) pre-swollen in CH2CI2 (3 mL) for 5 min, was added Fmoc-Dab-O-allyl (255 mg, 0.532 mmol, 4 equiv.), and DIPEA (185.3 pL, 1.1 mmol, 8 equiv.) in CH2CI2 (3 mL). The reaction mixture was agitated for 1 h at rt. The resin was filtered and washed with DMF (5 x 3 mL).

[0128] General method 2: Standard removal of Fmoc protecting groups

[0129] / Va-Fmoc of the ensuing nascent peptide was removed by treatment of the resin with 20% piperidine in DMF (v / v, 3 mL) for 2 x 5 min at rt. The resin was filtered and washed with DMF (5 x 3 mL).

[0130] General method 3: Standard coupling of Fmoc-amino acids

[0131] Following the removal of / Va-Fmoc protection, a mixture of the desired Fmoc-AA-OH (4 equiv.), HCTU (3.8 equiv.), and DIPEA (8 equiv.) dissolved in DMF (3 mL) was added to the resin. The reaction mixture was shaken for 1 h at rt. The resin was filtered and washed with DMF (5 x 3 mL).

[0132] General method 4: Removal of Ny-Dde-Dab protecting group

[0133] The peptidyl resin (0.13 mmol) was treated with a solution of hydroxylamine hydrochloride (1.66 g, 23.9 mmol, 180 equiv.) and imidazole (1.18 g, 17.3 mmol, 130 equiv.) in / V-methyl-2-pyrrolidinone (3 mL) and agitated for 3 h at rt. The resin was filtered and washed with DMF (5 x 3 mL).

[0134] General method 5: Standard removal of Allyl and Alloc protecting groups

[0135] The peptidyl resin (0.13 mmol) was treated with tetrakis(triphenylphosphine)-palladium(0) [Pd(PPh3)4] (153.7 mg, 0.13 mmol, 1 equiv.), phenylsilane (164.1 pL, 1.3 mmol, 10 equiv.) in CH2CI2 (3 mL) and agitated for 3 h at rt. The resin was filtered and washed with CH2CI2 (2 x 3 mL), sodium diethyldithiocarbamate (3 x 3 mL, 0.5 w / w in DMF), and DMF (3 x 3 mL).

[0136] General method 6: On-resin cyclisation of linear peptides

[0137] To the peptidyl resin (0.13 mmol) was added a solution of PyAOP (208.0 mg, 0.39 mmol, 3 equiv.) and DIPEA (139.0 pL, 0.78 mmol, 6 equiv.) in DMF (3 mL) and agitated for 12 h at rt. The resin was filtered and washed with DMF (5 x 3 mL).

[0138] General method 7: Coupling of 3-butynoic acid to peptides on-resin

[0139] Introduction of the allenamide moiety to peptides was performed according to the protocol by Cameron et al. To the / Va-amino deprotected peptidyl resin (0.133 mmol) was added a solution of 3-butynoic acid (111.8 mg, 1.33 mmol, 10 equiv.), EEDQ (312.5 mg, 1.26 mmol, 9.5 equiv.), sym-collidine (158.2 pL, 1.20 mmol, 9 equiv.) in CH2CI2 (3 mL). The reaction mixture was agitated for 2 h at rt. The resin was filtered and washed with CH2CI2 (5 x 3 mL).

[0140] General method 8: Global side chain deprotection and cleavage of peptide from resin

[0141] Following removal of the final / Va-Fmoc protecting group for the corresponding peptide, the peptidyl resin was treated with a mixture of TFA / H2O / TIPS (95:2.5:2.5, v / v / v, 1 mL / 0.01 mmol of peptidyl resin) for 2 h at rt. The filtrate was concentrated under a stream of N2, and the peptide was precipitated using cold diethyl ether (ca. 50 mL). The precipitate was isolated by centrifugation (4000 rpm, 5 min) and the supernatant was discarded. The pellet was resuspended in fresh, cold diethyl ether, and the trituration was repeated. The precipitate was dissolved in FW / MeCN (1: 1, v / v, 20 mL) and lyophilised.

[0142] General method 9: Elimination and exchange of TFA counterion to chloride counterion

[0143] Peptides were redissolved in 10 mM aq. HCI (100 pL) in FbO / MeCN (1: 1, v / v, 5 mL) and lyophilised. This procedure was repeated twice more. Synthesis of quaternary ammonium-modified polymyxin B nonapeptide (QA- PMBN) analogues – General method Using 2-chlorotrityl resin (0.133 mmol), initial coupling of Fmoc-Dab-γ-NH2-(OAllyl) was performed according to General method 1. The peptide was elongated with iterative cycles of Fmoc removal and amino acid couplings in accordance with General methods 2 and 3, respectively, up to the terminal Fmoc-Thr(tBu)-OH residue. Removal of the orthogonal Dde protecting group was performed according to General method 4. Alloc- Thr(tBu)-OH was coupled to the peptidyl resin using PyAOP (545.9 mg, 1.05 mmol, 7.9 equiv.), and DIPEA (360.7 µL, 2.13 mmol, 16 equiv.) dissolved in DMF (3 mL) and was shaken for 1 h at rt. Removal of the orthogonal allyl and Alloc protecting group was performed according to General method 5. On-resin cyclisation was performed using General method 6. After the final Fmoc was removed (General method 2), the allenamide handle was introduced by coupling 3-butynoic acid using EEDQ for 2 h according to General method 7. The peptidyl resin was split into six 0.022 mmol portions. Preparation of QA-PMBN1 The quaternary ammonium moiety was introduced by addition of DMHA (38.5 µL, 0.22 mmol, 10 equiv.) and aq. HCl (22 µL, 0.11 mmol, 5 equiv., 5 M, aq.) to the peptidyl resin in propylene carbonate for 22 h. Resin cleavage according to General method 8, purification by RP-HPLC, and TFA to HCl salt exchange according to General method 9 yielded peptide QA-PMBN1 as an amorphous white powder (2.1 mg, 8%); tR= 25.1 min; MS (ESI+) m / z [M]+observed 1159.1, [C55H96N15O12]+requires 1158.7. Preparation of QA-PMBN2 The quaternary ammonium moiety was introduced by addition of DMDA (38.5 µL, 0.22 mmol, 10 equiv.) and aq. HCl (22 µL, 0.11 mmol, 5 equiv., 5 M, aq.) to the peptidyl resin in propylene carbonate for 22 h. Resin cleavage according to General method 8, purification by RP-HPLC, and TFA to HCl salt exchange according to General method 9 yielded peptide QA-PMBN2 as an amorphous white powder (1.2 mg, 4% yield); tR = 33.6 min; MS (ESI+) m / z [M]+observed 1215.1, [C59H104N15O12]+requires 1214.8. Preparation of QA-PMBN3 The quaternary ammonium moiety was introduced by addition of DMTDA (38.5 µL, 0.22 mmol, 10 equiv.) and aq. HCl (22 µL, 0.11 mmol, 5 equiv., 5 M, aq.) to the peptidyl resin in propylene carbonate for 22 h. Resin cleavage according to General method 8, purification by RP-HPLC, and TFA to HCl salt exchange according to General method 9 yielded peptide QA-PMBN3 as an amorphous white powder (1.4 mg, 5% yield); tR = 42.3 min; MS (ESI+) m / z [M]+observed 1271.3, [C63H112N15O12]+requires 1270.9. Example 11: Comparison of antimicrobial activity of polymyxin analogues of the invention The antimicrobial activity of PMB analogues 1-3 was tested using the protocol outlined in Example 9. Tested in cation adjusted MHB, activity in non-cation adjusted media is approx. 2-fold greater (MIC = ½). The results are shown in Table 4. Table 4: Antimicrobial activity of PMB analogues Example 12: Stability of OQA compounds in seawater The stability in seawater of several OQAs of the invention was assessed according to the OECD Guidelines for Testing of Chemicals Biodegradability in Seawater. (OECD Guideline for Testing of Chemicals (306): Biodegradability in Seawater Paris, 1992) The closed bottle method was employed as it enables lower test concentrations to be employed. Experiments were performed using enriched seawater (Aerobic plate count at 25°C was 6.9×106cfu / 100 mL, APHA 9215B). ThOD is the Theoretical oxygen demand of the compound. Results for NaOAc confirm that the seawater used for testing is biologically active. Compounds of the invention B3a, B3b and C5a were prepared as described in Example 8. The percentage biodegradation of the QACs and DDAC was measured at 5, 15 and 28 days. The results are shown in Table 5. The compounds of the invention had a significantly shorter half-life than DDAC.

[0144] Table 5: % Biodegradation of QAC / OQA compounds in seawater

[0145] 7. REFERENCES

[0146] Audin, Drut-Grevoz, & Paris. (1993). Synthetic Communications, 23 #8, 1139-1145.

[0147] Cameron, A. J., Harris, P. W., & Brimble, M. A. (2020). On-Resin Preparation of Allenamidyl Peptides: A Versatile Chemoselective Conjugation and Intramolecular Cyclisation Tool. Angewandte Chemie International Edition, 59 (41), 18054- 18061.

[0148] De Zoysa, G. H., & Sarojini, V. (2016). Feasibility Study Exploring the Potential of Novel Battacin Lipopeptides as Antimicrobial Coatings. ACS Appin. Mater. Interfaces, 1373-1383.

[0149] Desmet, G. B., Sabbe, M. K., D'hooge, D. R., Espeel, P., Celasun, S., Marin, G. B., . . . Reyniers, M.-R. (2017). Thiol-Michael Addition in Polar Aprotic Solvents: Nucleophilic Initiation or Base Catalysis? Polym. Chem., 8(8), 1341-1352.

[0150] Frayne, S. H., & Northrop, B. H. (2018). Evaluating Nucleophile Byproduct Formation during Phosphine- and Amine-Promoted Thiol-Methyl Acrylate Reactions. J. Org. Chem., 83 (17), 10370-10382.

[0151] Harris, P. W., Yang, S. H., & Brimble, M. A. (2011). Harris, P. W. R.; Yang, S. H.; Brimble, M. A. An Improved Procedure for the Preparation of Aminomethyl Polystyrene Resin and Its Use in Solid Phase (Peptide) Synthesis. Tetrahedron Lett. 2011, 52 (45), 6024-6026. https: / / doi.Org / 10.1016 / j.tetlet.2011.09.0. Tetrahedron Lett., 52 (45), 6024-6026.

[0152] Isidro-Llobet, A., Alvarez, M., 8<. Albericio, F. (2009). Amino Acid-Protecting Groups. Chem. Rev., 109(6), 2455-2504. doi: https: / / doi.org / 10.1021 / cr800323s

[0153] Jiao, Y., Niua, L.-n., Maa, S., Li, J., Tayd, F. R., & Chena, J.-h. (2017). Quaternary ammonium-based biomedical materials: State-of-the-art, toxicological aspects and antimicrobial resistance. Progress in Polymer Science, 71, 53-90.

[0154] Mbofana, C. T., & Miller, S. J. (2014). Journal of the American Chemical Society, 136 #8, 3285-3292.

[0155] OECD Guideline for Testing of Chemicals (306): Biodegradability in Seawater Paris. (1992). doi: https: / / doi.org / 10.1787 / 9789264070486-en

[0156] Ongwae, G. M., Morrison, K. R., Allen, R. A., Kim, S., Im, W., Wuest, W. M., & Pires, M. M. (2020). Broadening Activity of Polymyxin by Quaternary Ammonium Grafting. ACS Infect Dis., 6(6), 1427-1435.

[0157] Performance standards for antimicrobial susceptibility testing; sixteenth informational supplement. (2023). Wayne, PA.

[0158] Rusli, W., Periayah, M. H., Loh, X. J., Lakshminar, R., & Parthiban, A. (2023). Synthesis, characterization and cell selectivity of poly(quaternary ammonium chlorides): Effect of degree of quaternization and copolymer composition. Mater. Adv. doi: 10.1039 / D3MA00661A Tomida, Y., Nagaki, A., & Yoshida, J.-I. (2011). Journal of the American Chemical Society, 133 #11, 3744-3747. Trifonov, L. S., & Oharovats, A. S. (1987). Helvetica Chimica Acta, 69, 262-270. Trifonov, L. S., & Orahovats, A. S. (1986). Helvetica Chimica Acta, 69, 1585-1587. Xie, H., & Ma, S. (2002). Journal of Organic Chemistry, 67 #18, 6575-6578.

Claims

WHAT WE CLAIM:

1. A compound of Formula Iwherein R and R’ are independently selected from H, -(C1-12)-alkyl, -(C2-12)-alkenyl, - (C2-12)-alkynyl (which latter three groups are optionally substituted with one or more substituents selected from the group consisting of halo, nitro, -N3, =O, -CN, -OR10a,- SR10b, -COR10c, -COOR10d, -NR10eR10f, -CONR10gR10h, carbocyclea1, aryla1, heterocyclea1, heteroaryla1, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), carbocycleb1, arylb1, heterocycleb1, heteroarylb1, (which four groups are optionally substituted with one or more substituents selected from halo, nitro, -N3, -CN, -OR11a,- SR11b, -COR11c, -COOR11d, -NR11eR11f, -CONR11gR11h, carbocyclea2, aryla2, heterocyclea2, heteroaryla2, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), or R and R’, together with the N atom to which they are attached, represent a 3-10- membered heterocyclic or heteroaryl ring which may additionally contain one or more heteroatoms selected from O, S and N (which heterocyclic or heteroaryl ring is optionally substituted by one or more substituents selected from halo, nitro, -N3, -CN, -OR12a, -SR12b, -COR12c, -COOR12d, -NR12eR12f, -CONR12gR12h, carbocyclea3, aryla3, heterocyclea3, heteroaryla3, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), or R is H and -NHR’ represents an amino acid or amino acid derivative or a peptide, or -NRR’ represents an amine-functionalised polymer or material; wherein R1, R2and R3are independently selected from -(C1-14)-alkyl or-(C1- 14)carbocycle or R1and R2together with the N atom to which they are attached, form a 3-14-membered heterocyclic or heteroaryl ring which may additionally contain one or more heteroatoms selected from O, S and N (which heterocyclic or heteroaryl ring is optionally substituted with one to three methyl groups); and wherein -(C1-14)-alkyl may be optionally substituted with aryl10;wherein R4, R5and R6are independently selected from H, halo, nitro, -N3, -CN, -OR13a, -SR13b, -COR13c, -COOR13d, -NR13dR13f, -CONR13gR13h, -(C1-6)-alkyl, -(C2-6)-alkenyl, -(C2-6)-alkynyl (which latter three groups are optionally substituted with or more substituents selected from the group consisting of halo, nitro, =O, -N3, CN, -OR14a,- SR14b, -COR14c, -COOR14d, -NR14dR14f, -CONR14gR14h, carbocyclea5, aryla5, heterocyclea5, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl), carbocycleb2, arylb2,heterocycleb2and heteroarylb2(which four groups are optionally substituted with one or more substituents selected from halo, nitro, -N3, -CN, -OR15a,- SR15b, -COR15c, -COOR15d, -NR15eR15f, -CONR15gR15h, carbocyclea6, aryla6, heterocyclea6, heteroaryla6, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl); wherein each carbocyclea, aryla, heterocycleaand heteroarylais independently selected from a 4-14-membered ring as earlier defined, optionally substituted with one or more substituents selected from halo, nitro, =O, -N3,-CN, -OR16a,-SR16b, - COR16c, -COOR16d, -NR16dR16f, -CONR16gR16h; wherein each carbocycleb, arylb, heterocycleband heteroarylbis independently selected from a 4 to 14-membered ring as earlier defined; wherein each R10-16ato R10-16his independently selected from H, -(C1-10)-alkyl, -(C2-10)- alkenyl, -(C2-10)-alkynyl (which latter three groups are optionally substituted with or more substituents selected from the group consisting of halo, nitro, -N3, -CN, =O, - OR17a,-SR17b-COR17c, -COOR17d, -NR17eR17f, -CONR17gR17h, carbocycleb3, arylb3, heterocycleb3, heteroarylb3, -(C1-6)-alkyl, -(C2-6)-alkenyl, -(C2-6)-alkynyl and -(C3-6)- cycloalkyl); or the pairs R10-16eand R10-16fand / or R10-16gand R10-16hmay form, together with the N atom to which they are attached, a 3-14 membered heterocyclic or heteroaryl ring which may contain one or more heteroatoms selected from O, S and N, which heterocyclic ring is optionally substituted by one or more substituents selected from halo, nitro, -N3, CN, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl; wherein each R17ato R17his independently selected from H, -(C1-10)-alkyl, -(C2-10)- alkenyl and -(C2-10)-alkynyl; wherein aryl10is a 6-14-membered aryl group; wherein indicates that a double bond is present at one of the two possible positions and X is selected from the group consisting of Cl, Br, I, CF3COO, CH3COO, CCl3COO, tetrafluoroborate (BF4), hexafluorophosphate (PF6), bis- trifluoromethanesulfonimide (NTf2), trifluoromethanesulfonate (OTf), dicyanamide(N(CN)2), hydrogen sulphate (HSO4), ethyl sulphate (EtOSO3) and 1-butyl-3- methylimidazolium tetrachloroferrate.

2. A compound of claim 1 wherein R is H and R’ is -(C1-12)-alkyl optionally substituted with one or more substituents selected from the group consisting of halo, nitro, -N3, =O, -CN, -OR10a, -SR10b, -COR10c, -COOR10d, -NR10eR10f, -CONR10gR10h, carbocyclea1, aryla1, heterocyclea1, heteroaryla1, -(C1-6)-alkyl, -(C2-6)-alkenyl and -(C2-6)-alkynyl.

3. A compound of claim 2 wherein R’ is -(C1-12)-alkyl substituted with -(C6-14)-aryl, preferably phenyl.

4. A compound of claim 2 or claim 3 wherein R’ is -(C1-12)-alkyl substituted with - COO(C1-C10alkyl), preferably -COOtBu.

5. A compound of any preceding claim wherein R’ is -SR10awherein R10ais methyl.

6. A compound of any preceding claim wherein R’ is -(C1-12)-alkyl substituted with - CONR10gR10hwherein R10gis H and R10his -(C1-C10alkyl).

7. A compound of claim 6 wherein R10his -(C1-C10alkyl) substituted with -CONR17gR17hwherein R17gand R17hare both H.

8. A compound of claim 7 wherein R10his -(C1-C10alkyl) substituted with -(C6-14)-aryl wherein -(C6-14)-aryl is phenyl.

9. A compound of any preceding claim wherein R’ is -(C1-C10alkyl) substituted with heterocycle, preferably indole.

10. A compound of claim 1 wherein R is H and -NHR’ represents an amino acid or amino acid derivative or a peptide.

11. A compound of claim 10 wherein the peptide is a 1-10 amino acid oligopeptide.

12. A compound of claim 1 wherein -NRR’ represents an amine-functionalised polymer or material, preferably glass, silicon or titanium.

13. A compound of any preceding claim wherein R1, R2and R3are -(C1-14)-alkyl, preferably methyl.

14. A compound of claim 13 wherein R1and R2are methyl and R3is -(C6-14)-alkyl optionally substituted with phenyl.

15. A compound of claim 13 wherein R1and R2are methyl and R3is cyclohexyl.

16. A compound of claim 13 wherein R1and R2are methyl and R3is -(C1-6)-alkyl substituted with –(C6-C14)aryl, preferably phenyl.

17. A compound of any preceding claim wherein X is Cl.

18. A process of making a compound of Formula I comprising reacting an allenamide compound of Formula II with a tertiary amine of Formula III,wherein R, R’, R1-R3, X andare as defined in claim 1.

19. A process of claim 18 wherein R1-R3are all H, and the allenamide (II) is made by reacting NHRR’ with 3-butynoic acid.

20. A process of making a compound of Formula IA comprising reacting an amine of Formula IV with 3-butynoic acid (V) in the presence of a peptide coupling reagent and a tertiary amine of Formula III., , , efined in claim 1.

21. A process of any one of claims 18 to 20 wherein the reaction takes place in a cyclic carbonate solvent, preferably selected from propylene carbonate (PC), ethylene carbonate (EC) and acetonitrile (MeCN).

22. A process of any one of claims 18 to 21 wherein R is H and -NHR’ is a peptide and the reaction is carried out on a solid phase polymeric support.

23. A process of any one of claims 18 to 21 wherein -NRR’ is an amine-functionalised polymer or material.

24. A process of any one of claims 17 to 23 wherein the peptide coupling reaction is Mukaiyama reagent or EEDQ.

25. A process of any one of claims 18 to 24 wherein 2, 3, 5, 10, 15 or 20 molar equivalents of tertiary amine (III) are present.

26. A process of any one of claims 18 to 25 wherein a proton source (preferably HCl) is added to the reaction.

27. A compound of any one of claims 1 to 17 which is bound to the surface of a material via a trialkyloxysilane.

28. A material comprising one or more compounds of any one of claims 1 to 17.

29. A compound or material of claim 27 or 28 wherein the material is selected from glass, filter paper, stainless stell, titanium, gold, silicon wafer, silicon rubber, PVDF membrane, polymer microsphere, polyolefin and polypropylene.

30. A compound or material of any one of claims 27 to 29 wherein the material comprises or coats a medical device, for example a surgical tool, catheter, heart valve, pacemaker, stent, prosthetic joint or contact lens.

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