Processes for making amberketal from silylated hydroxyfarnesylacetonate

Abstract

It is provided a compound of formula (I) wherein R1, R2 and R3 are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond. The compound can release hydroxyfarnesylacetone, and can be used as starting material in the process of making Amberketal.

Description

[0001] ORGANIC COMPOUNDS

[0002] TECHNICAL FIELD

[0003] The present invention relates generally to compounds able to release hydroxyfarnesylacetone and their use, in particular as starting material for the preparation of Amberketal.

[0004] BACKGROUND

[0005] Amberketal provides a powerful and tenacious ambery and woody odour that is useful in fragrance compositions alone or in combination with other woody or ambery ingredients. Amberketal is traditionally prepared from Manool via a number of chemical transformations. Unfortunately, the supply of naturally derived Manool is limited. For this reason, in recent years’ new routes have been developed. For example, it is known from WO2021 / 209482 that (+)- Amberketal can be prepared from (5Z,9E)-6-(hydroxymethyl)-10,14-dimethylpentadeca- 5,9,13-trien-2-one (CAS 173198-97-5, (5Z,9E)-hydroxyfarnesylacetone, (5Z,9E)-HFA) by biocatalysis. Therefore, it is desirable to provide a new efficient and cost effective route to obtain HFA or its analogues.

[0006] SUMMARY

[0007] In accordance with a first aspect of the present invention there is provided a compound of formula (I) wherein Ri, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond.

[0008] In accordance with a second aspect of the present invention there is provided the use of a compound of formula (I)

[0009] wherein Ri, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond to release hydroxyfarnesylacetone. In accordance with a third aspect of the present invention there is provided the use of a compound of formula (I) wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond as starting material for the process of making Amberketal.

[0010] In accordance with a fourth aspect of the present invention there is provided a process of making (+)-Amberketal (a compound of formula (II)) from a compound of formula (I) by a SHC (squalene-hopene cyclase) mediated process. In accordance with a fifth aspect of the present invention there is provided a method of making the compound of formula (I) wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond.

[0011] In accordance with a sixth aspect of the present invention there is provided a process of making (+)-Amberketal (a compound of formula (II)) from a compound of formula (I) by a SHC (squalene-hopene cyclase) mediated process, that is obtained by the previously described method.

[0012] Certain embodiments of any aspect of the present invention may provide one or more of the following advantages:

[0013] • Access to hydroxyfarnesylacetone;

[0014] • control of the CC double bond configuration of hydroxyfarnesylacetone;

[0015] • improved purification and storage of hydroxyfarnesylacetone; and / or

[0016] • facile conversion to further products.

[0017] The details, examples and preferences provided in relation to any particular one or more of the stated aspects of the present invention will be further described herein and apply equally to all aspects of the present invention. Any combination of the embodiments, examples and preferences described herein in all possible variations thereof is encompassed by the present invention unless otherwise indicated herein, or otherwise clearly contradicted by context.

[0018] DETAILED DESCRIPTION

[0019] The present invention is based on the surprising finding that the compound of formula (I) can be used to release hydroxyfarnesylacetone (compound of formula (III)) in situ, which can be then further converted to other compounds. So in the first aspect of the invention, there is provided a compound of formula (I) wherein Ri, R2and R3 are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond.

[0020] C1-C4 alkyl includes methyl, ethyl, n-propyl, / -propyl, n-butyl, / -butyl and f-butyl.

[0021] For example, Si(RiR2R3) is trimethyl silyl (TMS), triethyl silyl (TES), triisopropyl silyl (TIPS), tertbutyl dimethyl silyl (TBDMS), or te / t-butyldiphenylsilyl (TBDPS).

[0022] For example, compound of formula (I) is selected from the group consisting of 10,14-dimethyl- 6-(((trimethylsilyl)oxy)methyl)pentadeca-5,9,13-trien-2-one (compound of formula (la)), 10,14- dimethyl-6-(((triethylsilyl)oxy)methyl)pentadeca-5,9,13-trien-2-one (compound of formula (lb)), 10,14-dimethyl-6-(((triisopropylsilyl)oxy)methyl)pentadeca-5,9,13-trien-2-one (compound of formula ( I c)) , 6-(((te / t-butyldimethylsilyl)oxy)methyl)-10, 14-dimethylpentadeca-5,9, 13-trien-2- one (compound of formula (Id)), and 6-(((tert-butyldiphenylsilyl)oxy)methyl)-10,14- dimethylpentadeca-5,9,13-trien-2-one (compound of formula (le)).

[0023] For example, the compound of formula (I) has the double bond between C-9 and C-10 in E- configuration and the double bond between C-5 and C-6 in Z-configuration.

[0024] In one embodiment, the compound of formula (I) is a starting material for the preparation of Amberketal.

[0025] For example, the compound of formula (I) with the double bond between C-9 and C-10 in E- configuration and the double bond between C-5 and C-6 in Z-configuration is a starting material for the preparation of (+)-Amberketal.

[0026] For example, the compound of formula (la) has a significantly lower boiling point than HFA, which allows for a better purification by distillation. The compound of formula (la) has a boiling point of about 152-158°C at 0.06 mbar in fractional distillation. In contrast to this, HFA, the compound of formula (III), has a boiling point of about 180°C at 0.04 mbar, when using a wiped film apparatus.

[0027] In a further aspect of the present invention, there is provided the use of the compound of formula (I) to release hydroxyfarnesylacetone (compound of formula (III)).

[0028] Hydroxyfarnesylacetone (HFA, compound of formula (III)) is an analogue of the compound of formula (I) with a free hydroxyl group.

[0029] In comparison to the use of HFA as such, the present invention shows several advantages: The compound of formula (I) has a higher thermal stability and therefore, it can be purified by conventional distillation, while HFA as such requires more elaborate purification process, for example wiped film distillation which is slow and higher in energy consumption and requires two distillation cycles to first remove impurities with a lower boiling point, followed by distillation of HFA as such; or column chromatography which requires high amounts of solvents. The compound of formula (I) is also more stable upon storage and does not degrade when exposed to air. So the use of the compound of formula (I) instead of HFA leads to a more economically viable production process. The in situ generated HFA is pure and can be used as is for further reactions.

[0030] For example, there is provided the use of the compound of formula (I) wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond as starting material for the process of making Amberketal (compound of formula (II)) wherein the compound of formula (I) releases hydroxyfarnesyl acetone, which is further converted to the compound of formula (II).

[0031] The in situ released hydroxyfarnesylacetone (HFA) can be used further for the process of making Amberketal (compound of formula (II)). The use of the compound of formula (I) is advantageous over the use of HFA (compound of formula (III)). The compound of formula (I) has a higher thermal stability (e.g. it does not undergo rearrangement to 1-(2-(6,10- dimethylundeca-1 ,5,9-trien-2-yl)cyclopropyl)ethan-1-one upon heating) , and it is more stable upon storage and does not degrade, so it is thereby not hindering the conversion in the biotransformation to Amberketal over time.

[0032] For example, there is provided herein the use of the compound of formula (I), wherein said compound has the double bond between C-9 and C-10 in E-configuration and the double bond between C-5 and C-6 in Z-configu ration as starting material for the process of making of (+)- Amberketal.

[0033] In a further aspect of the present invention, there is provided a process of making Amberketal (compound of formula (II))

[0034] from the compound of formula (I) wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond, by a SHC (squalene-hopene cyclase) mediated process.

[0035] The SHC (squalene-hopene cyclase) mediated process for the conversion of polyunsaturated alcohols like HFA to Amberketal and amberketal homologues is well described in international publication WO2021 / 209482.

[0036] In the present invention, for example, there is provided the process of making (+)-Amberketal, wherein the compound of formula (I) has the double bond between C-9 and C-10 in E- configuration and the double bond between C-5 and C-6 in Z-configuration.

[0037] In one specific embodiment there is provided the process of making Amberketal comprising the steps of: a) providing the compound of formula (I) wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, and wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond; b) releasing hydroxyfarnesylacetone; and c) contacting hydroxyfarnesylacetone with SHC.

[0038] For example, in step b), the compound of formula (I) is pretreated with acid to release HFA.

[0039] For example, the compound of formula (lb) is admixed with succinic acid / SDS, to release HFA. Thereafter, the pH is adjusted with NaOH to reach pH 5.6 (buffered mix) necessary for the biotransformation.

[0040] Alternatively, the compound of formula (la) readily releases HFA under the reaction conditions required for step c) at a pH of 5.0 - 6.0 (0.1 M succinic acid / sodium hydroxide buffer), and no pretreatment with acid is required. However, under acidic conditions, the release of HFA from the compound of formula (la) is faster. So if the compound of formula (la) is provided in the process of making Amberketal, the pretreatment with acid is optional.

[0041] The acid for the pretreatment of the compound of formula (I) to release HFA may be any acid, for example it can be selected from the group consisting of succinic acid, phosphoric acid, sulfuric acid, acetic acid, citric acid, maleic acid, hydrochloric acid, and others.

[0042] For example, there is provided the process of making (+)-Amberketal comprising the steps of: a) providing the compound of formula (I) wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, b) releasing hydroxyfarnesylacetone; and c) contacting hydroxyfarnesylacetone with SHC.

[0043] The compound of formula (I) can be prepared from HFA by direct etherification. Alternatively, the compound can be prepared in five steps from [3-farnesene (7,11-dimethyl-3- methylenedodeca-1 ,6,10-triene).

[0044] In a further aspect of the present invention, there is provided a method of making the compound of formula (I) wherein Ri, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond; comprising the steps of: a) providing [3-farnesene (compound of formula (V)); b) epoxidizing [3-farnesene to epoxide VI; c) reacting epoxide VI with R4CI to obtain the corresponding R4-ether chloride (compound of formula (VII)); d) forming the ketoester (compound of formula VIII); e) optionally isolating the ketoester; f) decarboxylating the ketoester to HFA (compound of formula III); g) subsequent re-protecting the liberated alcohol group with R5CI to obtain the compound of formula (I), h) optionally purifying the compound of formula (I), for example by fractional distillation.

[0045] In the method of making the compound of formula (I) as described above, either the isolation of the ketoester (step e)) or the purification of the compound of formula (I) (step h)) is carried out or both steps are carried out in the same sequence. The reaction scheme can be found in Figure 1.

[0046] In step a), [3-farnesene (compound of formula (V), 7,11-dimethyl-3-methylene-1 , 6,10- dodecatriene) is provided. Said compound is an isomer of farnesene. The compound of formula (V) may be (6E)-[3-farnesene, which may be referred to as (6E)-7,11-dimethyl-3-methylene- 1 ,6,10-dodecatriene. The compound of Formula (V) may be the compound with CAS number [18794-84-8] or the compound with CAS number [77129-48-7],

[0047] The epoxidation according to step b) of [3-farnesene to epoxide VI can be performed according to the procedure as described in international patent application WO 2022 / 268840. The epoxide of formula (VI, exo epoxide) is obtained in mixture with the corresponding 1 ,2-epoxide of [3-farnesene (7,11-dimethyl-3-methylene-1 ,6,10-dodecatriene). The reaction mixture can be used for next step(s), and the desired product can be isolated at later stage, respectively.

[0048] In step c), epoxide VI is reacted with R4CI to obtain the corresponding R4-ether chloride (compound of formula (VII)). Using R4CI to open the epoxide has the advantage that the epoxide VI reacts faster than the by-product (1 ,2-epoxide) and gives selectively the compound of formula (VII) in Z configuration.

[0049] For example, the R4CI in step c) is selected from the group consisting of TMSCI, TESCI and AcCI.

[0050] Step d) is the formation of the ketoester (compound of formula VIII) by reaction of R4-ether chloride (compound of formula (VII)) with methylacetoacetate.

[0051] Optionally, the obtained ketoester can be isolated at this stage from side products, like 1 ,2- epoxide and / or [3-farnesene (step e)). For example, the side products can be simply and efficiently removed by distillation or by extraction with an organic solvent, while ketoester VIII remains in the aqueous phase. From the side products, [3-farnesene can be recycled. Alternatively, the reaction mixture can be subjected to the next reaction steps, and the purification can be carried out at a later stage.

[0052] For example, if the ketoester bears a TMS-group (compound of formula (VIII) where R4= TMS), said group is readily removed when in contact with diluted aqueous NaOH. The so formed hydroxy-ketoester is more water-soluble and allows for the removal of impurities like farnesene, 1 ,2-epoxide and other impurities with an organic solvent, for example with isopropyl acetate, MTBE, heptane, cyclohexane, methyl-THF, or mixtures thereof, or others. In the case that the protecting group (R4) is not readily removed, methanol as co-solvent can be added to help solubilizing the ketoester in the water phase. This has the drawback to also solubilize impurities which are then more difficult to remove by extraction with an organic solvent.

[0053] In step f), the ketoester is decarboxylated under basic conditions, that simultaneously might cause the deprotection of the hydroxyl group, and hydroxyfarnesylacetone (compound of formula III) is obtained.

[0054] This crude product is subsequently protected with R5CI in step g) to yield crude compound of formula (I).

[0055] RsCI in step g) corresponds to the chloride of Si(RiR2Rs), wherein R1, R2 and R3 are independently selected from C1-C4 alkyl and phenyl. For example, R5CI is selected from the group consisting of trimethyl silyl chloride (TMSCI), triethyl silyl chloride (TESCI), triisopropyl silyl chloride (TIPSCI), tert-butyl dimethyl silyl chloride (TBDMSCI), or tert-butyldiphenylsilyl chloride (TBDPSCI). In a preferred embodiment, R5CI in step g) is TMSCI.

[0056] Optionally, the crude compound of formula (I) can be purified by simple and efficient techniques, for example by fractional distillation.

[0057] In one embodiment of the present invention, the R4CI in step c) is selected from the group consisting of TMSCI, TESCI and AcCI, and R5CI in step g) is TMSCI.

[0058] The described method offers access to HFA and provides a simple way of protecting it for storage. The method offers also the possibility to recover unreacted farnesene as well as the undesired epoxide by extraction.

[0059] In a further aspect of the present invention, there is provided a process of making Amberketal (compound of formula (II)) from the compound of formula (I) wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond, by a SHC (squalene-hopene cyclase) mediated process, wherein the compound of formula (I) is obtained by the method comprising the steps of: a) providing [3-farnesene (compound of formula (V)); b) epoxidizing [3-farnesene to epoxide VI; c) reacting epoxide VI with R4CI to obtain the corresponding Rzi-ether chloride (compound of formula (VII)); d) forming the ketoester (compound of formula VIII); e) optionally isolating the ketoester; f) decarboxylating the ketoester to HFA (compound of formula III); g) subsequent re-protecting the liberated alcohol group with R5CI to obtain the compound of formula (I), h) optionally purifying the compound of formula (I), for example by fractional distillation. The invention is now further described with reference to the following non-limiting examples. These examples are for the purpose of illustration only and it is understood that variations and modifications can be made by one skilled in the art.

[0060] EXAMPLES

[0061] General:

[0062] All reactions were performed under argon using solvents and reagents from commercial suppliers without further purification. Solvents for extraction and chromatography were technical grade and used without further purification. Flash chromatography was performed using commercially available prepacked silica gel cartridges. NMR spectra were recorded with Bruker AW 400 MHz or Avance III HD 400 MHz instruments. The chemical shifts for1H NMR spectra was reported in 5 (ppm) referenced to the residual proton signal of the deuterated solvent; coupling constants were expressed in Hertz (Hz).13C NMR spectra were referenced to the carbon signals of the deuterated solvent. The following abbreviations are used: s = singlet, d = doublet, t = triplet, q = quartet, quint. = quintuplet, m = multiplet, dd = double doublet, bs = broad singlet. GC / MS spectral data were obtained from an Agilent 6890 N and MSD 5975 using a column HP-5 MS, 30 m, 0.25 mm, 0.25 pm.

[0063] Example 1 : Preparation of (5Z,9E)-10,14-dimethyl-6-(((trimethylsilyl)oxy)methyl)pentadeca- 5,9,13-trien-2-one using method 1

[0064] Step 1 - (E)-2-(4,8-dimethylnona-3,7-dien-1-yl)-2-vinyloxirane:

[0065] The epoxidation of (E)-[3-farnesene was performed as described in WO2022268840 producing a mixture containing (E)-[3-farnesene and (E)-[3--farnesene epoxides (qNMR-purity: 29% (£)- P--farnesene, 34% (E)-2-(4,8-dimethylnona-3,7-dien-1-yl)-2-vinyloxirane and 26% (E)-2-(6,10- dimethylundeca-1 ,5,9-trien-2-yl)oxirane).

[0066] Step 2 - (((2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1-yl)oxy)trimethyl- silane:

[0067] A mixture of (E)-[3-farnesene and (E)-[3-farnesene epoxides containing (E)-2-(4,8- dimethylnona-3,7-dien-1-yl)-2-vinyloxirane (1614 g, 34%, 1 eq, 2.49 mol) was heated to 55°C, and chlorotrimethylsilane (273 g, 99%, 1 eq, 2.49 mol) was added dropwise over 10 minutes. The reaction temperature progressively rose to 78°C and at the end of the addition, the internal temperature was kept at 75°C until complete conversion of (E)-2-(4,8-dimethylnona-3,7-dien- 1-y|)-2-vinyloxirane was observed by GC (2h30min). The reaction mixture was cooled down, diluted with MtBE (2.0 I) and washed with saturated aqueous NaHCO3(2.0 I), with a 2 : 1 mixture of water and brine (2.0 I) and with brine (0.5 I). The organic phase was dried over MgSO4and concentrated under vacuum to give a light yellow transparent oil (1928 g) containing a mixture of (E)-[3-farnesene, (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane and (((2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1-yl)oxy)trimethylsilane (qNMR-purity: 25% (E)-[3-farnesene, 13% (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane and 30% (((2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1- yl)oxy)trimethylsilane (yield: 71%)). The crude mixture was used as is for the subsequent step.

[0068] Analytical data of (((2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1-yl)oxy)- trimethylsilane:

[0069] 1H NMR (CHLOROFORM-d, 25 °C) 5: 5.51 (t, J=8.0 Hz, 1 H), 5.06-5.17 (m, 2H), 4.19 (d, J=0.7 Hz, 2H), 4.17 (d, J=8.0 Hz, 2H), 1.93-2.28 (m, 8H), 1.68 (s, 3H), 1.60 (s, 6H), 0.14 (s, 9H) ppm.13C NMR (CHLOROFORM-d, 25 °C) 5: 142.0, 135.7, 131.3, 124.4, 124.4, 123.8, 60.5, 40.0, 39.9, 35.0, 26.7, 26.6, 25.7, 17.6, 16.0, -0.5 ppm.

[0070] GC-MS (El, 70 eV): 155 (31), 93 (24), 91 (15), 81 (21), 79 (14), 75 (17), 73 (72), 69 (100), 67 (17), 41 (52).

[0071] Step 3 - methyl (4Z,8E)-2-acetyl-9,13-dimethyl-5-(((trimethylsilyl)oxy)methyl)tetradeca-4,8,12- trienoate:

[0072] A suspension of potassium carbonate (736 g, 99%, 3 eq, 5.27 mol) in acetone (1.4 I) was heated to 50°C and methyl 3-oxobutanoate (619 g, 99%, 3 eq, 5.27 mol) was added dropwise over 15 minutes. The suspension was stirred 30 minutes before the addition of potassium iodide (9 g, 0.03 eq, 52.73 mmol) and of a solution of (((2Z,5E)-2-(2-chloroethylidene)-6,10- dimethylundeca-5,9-dien-1-yl)oxy)trimethylsilane (1928 g, 30 %, 1 eq, 1.76 mol) in acetone (0.6 I) in 25 minutes. The internal temperature was then raised to 63°C and the mixture was stirred until full conversion of (((2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1- yl)oxy)trimethylsilane was observed by GC (2h30min). The mixture was transferred to a round bottom flask and concentrated under vacuum. The residue was dissolved in a mixture of water (1 .5 I) and MtBE (1.51), the phases were separated and the aqueous phase was extracted once with MtBE (0.8 I). The organic phases were combined, washed twice with a 1 : 1 mixture of water and brine (1.5 I), once with brine (0.4 I), dried over MgSO4and concentrated under vacuum to give a yellow transparent oil (2284 g) containing a mixture of (E)-[3-farnesene, (E)- 2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane and methyl (4Z,8E)-2-acetyl-9,13-dimethyl-5- (((trimethylsilyl)oxy)methyl)tetradeca-4,8,12-trienoate (qNMR-purity: 20% (E)-p-farnesene, 11 % (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane and 27% methyl (4Z,8E)-2-acetyl- 9,13-dimethyl-5-(((trimethylsilyl)oxy)methyl)tetradeca-4,8,12-trienoate (yield: 86%)). The crude mixture was used as is for the subsequent step.

[0073] Analytical data of methyl (4Z,8E)-2-acetyl-9,13-dimethyl-5-(((trimethylsilyl)oxy)methyl)- tetradeca-4,8,12-trienoate:

[0074] 1H NMR (500 MHz, CHLOROFORM-d, 25°C): 5 = 5.15 (t, J = 7.5 Hz, 1 H), 5.06-5.12 (m, 2H), 4.15 (d, J = 11.9 Hz, 1 H), 4.11 (d, J = 11.9 Hz, 1 H), 3.73 (s, 3H), 3.48 (t, J = 7.5 Hz, 1 H), 2.62 (t, J = 7.5 Hz, 2H), 2.23 (s, 3H), 1.93-2.11 (m, 8H), 1.68 (d, J = 1.2 Hz, 3H), 1.60 (s, 3H), 1.59 (d, J = 0.9 Hz, 3H), 0.13 ppm (s, 9H) ppm.

[0075] 13C NMR (126 MHz, CHLOROFORM-d, 25°C): 5 = 202.7, 169.8, 141.5, 135.2, 131.3, 124.3, 123.9, 122.0, 59.7, 59.6, 52.4, 39.7, 34.8, 29.3, 26.7, 26.4, 25.7, 17.7, 16.0, -0.5 ppm.

[0076] GC-MS (El, 70 eV): 133 (43), 107 (25), 105 (23), 79 (27), 75 (25), 73 (72), 69 (100), 55 (24), 43 (72), 41 (70).

[0077] Step 4 - Hydroxyfarnesylacetone:

[0078] A 15 I Mini-Pilot reactor was charged with aqueous sodium hydroxide (4827 g, 5%, 4 eq, 6.03 mol) and the crude oil from step 3 containing methyl (4Z,8E)-2-acetyl-9,13-dimethyl-5- (((trimethylsilyl)oxy)methyl)tetradeca-4,8,12-trienoate (2283 g, 27%, 1 eq, 1.51 mol). The internal temperature reached 35°C during the addition. The mixture was stirred until full disappearance of methyl (4Z,8E)-2-acetyl-9,13-dimethyl-5-(((trimethylsilyl)oxy)methyl)- tetradeca-4,8,12-trienoate was observed by GO (1 h40min). Isopropyl acetate (2.5 I) was added to the reaction mixture which was stirred vigorously for 5 minutes. The phases were separated and the organic phase was concentrated under vacuum to give a light yellow oil (1159 g) containing a mixture of (E)-[3-farnesene and (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2- yl)oxirane (qNMR-purity: 36% (E)-[3-farnesene, 23% (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2- yl)oxirane). The aqueous phase was extracted once more with isopropyl acetate (1.3 I) and the organic phase was concentrated in vacuum to give a light yellow oil (122 g) containing a mixture of (E)-[3-farnesene and (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane (qNMR-purity: 30% (E)-[3-farnesene, 20% (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane). The resulting aqueous phase was basified by adding aqueous sodium hydroxide (284 g, 32%) to reach pH 14 and heated to 50°C. After 45 min, the pH dropped to 8 and additional aqueous sodium hydroxide (473 g, 32%) was added to reach pH 11. The mixture was stirred for 15 hours at 50°C before being cooled to 40°C at which temperature the phases were separated and the aqueous phase was extracted once with MtBE (2.5 I). The organic layers were combined and washed with a 2 : 1 mixture of water and brine (2.3 I), with water (1.5 I), with a 2 : 1 mixture of brine and water (1.5 I), with brine (0.5 I), dried over MgSO4and concentrated under vacuum to give a brown oil (508 g) containing hydroxyfarnesylacetone (qNMR-purity: 74%, yield: 89%). The crude product was used as is for the subsequent step.

[0079] Step 5 - (5Z,9E)-10,14-dimethyl-6-(((trimethylsilyl)oxy)methyl)pentadeca-5,9,13-trien-2-one:

[0080] A solution of crude hydroxyfarnesylacetone (458 g, 74%, 1 eq, 1 .22 mol) in tetra hydrofuran (4.0 I) was treated successively with triethylamine (160 g, 1.3 eq, 1.58 mol) and chlorotrimethylsilane (159 g, 1.2 eq, 1.46 mol) over 40 minutes. The internal temperature rose to 35°C during the addition. The resulting mixture was stirred at room temperature until full conversion of hydroxyfarnesylacetone was observed by GC (1 h). The mixture was quenched with aqueous saturated NaHCO3(2.0 I), the phases were separated and the organic phase was washed with NaHCO3(1.0 I), with a 1 : 1 mixture of water and brine (2.0 I), with brine (0.6 I), dried over MgSO4and concentrated under vacuum to give brown oil (557 g). The crude oil was purified by fractional distillation at 0.06 mbar and a boiling point range of 152-158°C to give (5Z,9E)-10, 14-dimethyl-6-(((trimethylsilyl)oxy)methyl)pentadeca-5,9, 13-trien-2-one (391 g, 0.94 mol, qNMR-purity: 84%, Yield: 77%).

[0081] Analytical data of (5Z,9E)-10,14-dimethyl-6-(((trimethylsilyl)oxy)methyl)pentadeca-5, 9,13- trien-2-one:

[0082] 1H NMR (CHLOROFORM-d, 25 °C) 5: 5.20 (t, J=7.3 Hz, 1 H), 5.05-5.14 (m, 2H), 4.14 (s, 2H), 2.44-2.51 (m, 2H), 2.33 (m, 2H), 2.13 (s, 3H), 2.01-2.11 (m, 6H), 1.94-2.01 (m, 2H), 1.68 (d, J=1.0 Hz, 3H), 1.57-1.62 (m, 6H), 0.13 (s, 9H) ppm.

[0083] 13C NMR (CHLOROFORM-d, 25 °C) 5: 208.0, 139.2, 134.9, 131.1 , 125.0, 124.2, 124.0, 59.6, 43.7, 39.6, 34.6, 29.8, 26.6, 26.64, 25.62, 21.8, 17.6, 15.9, -0.6 ppm.

[0084] GC-MS (El, 70 eV): 135 (17), 133 (49), 105 (17), 93 (19), 81 (33), 75 (30), 73 (60), 69 (95), 43 (100), 41 (68).

[0085] Example 2: Preparation of (5Z,9E)-10,14-dimethyl-6-(((trimethylsilyl)oxy)methyl)pentadeca-

[0086] 5,9,13-trien-2-one using method 2 Step 1 & 2: (2Z,5E)-2-(2-chloroethvlidene')-6.10-dimethvlundeca-5.9-dien-1-vl acetate

[0087] A solution of a mixture of (E)-[3-farnesene, (E)-2-(4,8-dimethylnona-3,7-dien-1-yl)-2- vinyloxirane and (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane, prepared as described in Step 1 of Example 1 , (100 g; (E)-[3-farnesene I (E)-2-(4,8-dimethylnona-3,7-dien-1-yl)-2- vinyloxirane I (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane) = 1 :0.7:0.5; 118 mmol (E)- 2-(4,8-dimethylnona-3,7-dien-1-yl)-2-vinyloxirane) in Me-THF (100 ml) at 5 °C was treated dropwise with acetyl chloride (10.2 ml, 142 mmol). The resulting mixture was stirred at 60 °C for 7h, at 50 °C for 16h and at 60 °C for 6h. After solvent evaporation under reduced pressure a crude mixture containing (2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1-yl acetate was isolated.

[0088] Analytical data of (2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1-yl acetate:1H NMR (400 MHz, CDCI3) 5 ppm 5.65 (br. t, J= 8.1 , 1 H), 5.15-5.05 (m, 2H), 4.64 (s, 2H), 4.16 (d, J = 8.1 , 2H), 2.06 (s, 3H), 2.25-1.95 (m, 8H), 1.68 (br. s, 3H), 1.61 (s, 6H).

[0089] 13C NMR (100 MHz, CDCI3) 5 ppm 170.74 , 139.54, 136.03, 131.36, 125.68, 124.21 , 122.94, 61.18, 39.63, 35.10, 26.65, 26.16, 25.66, 20.88, 17.66, 16.04.

[0090] GC-MS (El): 298 (1), 262 (1), 239 (1), 223 (1), 203 (2), 195 (3), 159 (2), 137 (6), 133 (10), 123 (7), 105 (13), 93 (17), 91 (18), 81 (28), 69 (100), 79 (13), 67 (16), 55 (10), 53 (10), 43 (41), 41 (47).

[0091] Step 3 - methyl (4Z,8E)-5-(acetoxymethyl)-2-acetyl-9,13-dimethyltetradeca-4,8,12-trienoate:

[0092] A suspension of potassium carbonate (627 g, 99%, 3 eq, 4.49 mol) in acetone (0.9 I) was heated to 60°C and methyl 3-oxobutanoate (527 g, 99%, 3 eq, 4.49 mol) was added dropwise over 10 min. The suspension was stirred for 40 min before potassium iodide (7 g, 0.03 eq, 44.91 mmol) and a solution of (2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1- yl acetate (1864 g, 24 %, 1 eq, 1.50 mol) in acetone (0.9 I) were added over 30 minutes. The internal temperature was raised to 63°C and the mixture was stirred until full conversion of (2Z,5E)-2-(2-chloroethylidene)-6,10-dimethylundeca-5,9-dien-1-yl acetate was observed by GO (2h30min). After concentration under vacuum, the residue was dissolved in a mixture of water (2.0 I) and MtBE (2.0 I). The phases were separated and the aqueous phase was extracted once with MtBE (1.0 I). The organic phases were combined, washed twice with a 2 : 1 mixture of water and brine (2.0 I), once with brine (0.6 I), dried over MgSO4and concentrated under vacuum to give a yellow transparent oil (2153 g) containing a mixture of (E)-[3-farnesene, (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane and methyl (4Z,8E)-5-(acetoxymethyl)-2- acetyl-9,13-dimethyltetradeca-4,8,12-trienoate (qNMR-purity: 24% (E)-[3- farnesene, 10% (E)- 2-(6,10-dimethylundeca-1 ,5,9-trien-2-yl)oxirane and 27±4% methyl (4Z,8E)-5-(acetoxymethyl)- 2-acetyl-9,13-dimethyltetradeca-4,8,12-trienoate (yield: quantitative)). The crude mixture was used as is for the subsequent step.

[0093] Analytical data of methyl (4Z,8E)-5-(acetoxymethyl)-2-acetyl-9,13-dimethyltetradeca-4,8,12- trienoate:

[0094] 1H NMR (400 MHz, CHLOROFORM-d, 25°C): 5 = 5.32 (t, J = 7.6 Hz, 1 H), 5.02-5.08 (m, 2H), 4.65 (d, J = 12.3 Hz, 1 H), 4.58 (d, J = 12.3 Hz, 1 H), 3.73 (s, 3H), 3.50 (t, J = 7.5 Hz, 1 H), 2.65 (t, J = 7.5 Hz, 2H), 2.24 (s, 3H), 2.06 (s, 3H), 1.93-2.11 (m, 8H), 1.68 (d, J = 1.0 Hz, 3H), 1.60 (s, 3H), 1.59 ppm (d, J = 1.0 Hz, 3H) ppm.

[0095] 13C NMR (101 MHz, CHLOROFORM-d, 25°C): 5 = 202.3, 170.9, 169.6, 136.8, 135.6, 131.3, 125.8, 124.2, 123.3, 61.6, 59.4, 52.4, 39.7, 35.3, 29.2, 26.7, 26.5, 26.4, 25.7, 20.9, 17.7, 16.0 ppm.

[0096] GC-MS (El, 70 eV): 139 (19), 133 (29), 107 (16), 81 (35), 79 (25), 69 (100), 67 (15), 55 (18), 43 (83), 41 (53).

[0097] Step 4 - Hydroxyfarnesylacetone

[0098] A 15 I Mini-Pilot reactor was charged with methyl (4Z,8E)-5-(acetoxymethyl)-2-acetyl-9,13- dimethyltetradeca-4,8,12-trienoate (2152 g, 27 %, 1 eq, 1.54 mol) which was treated with a solution of NaOH (691 g, 32%, 3.6 eq, 5.53 mol) in methanol (5.0 I) over 15 minutes at room temperature. The internal temperature reached 30°C during the addition. The mixture was stirred until full disappearance of methyl (4Z,8E)-5-(acetoxymethyl)-2-acetyl-9,13- dimethyltetradeca-4,8,12-trienoate was observed by GO (30 min). Heptane (3.0 I) was added to the reaction mixture which was stirred vigorously for 5 minutes. The phases were separated and the organic phase was concentrated under vacuum to give a light yellow oil (889 g) containing a mixture of (E)-[3-farnesene and (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2- yl)oxirane (qNMR-purity: 50% (E)-[3-farnesene, 20% (E)-2-(6,10-dimethylundeca-1 ,5,9-trien-2- yl)oxirane). The resulting aqueous methanolic phase was concentrated under vacuum (3.5 I of solvent was removed) and transferred back to the reactor. Water (2.5 I) and aqueous sodium hydroxide (180 ml, 32%) were added to the mixture (pH 14) which was heated to 54°C. After 1 h, the pH dropped to 12 and additional aqueous sodium hydroxide (40 ml, 32%) was added to reach pH 13. The mixture was stirred for 1 h at 53°C before being cooled to 40°C at which temperature the phases were separated and the aqueous phase was extracted once with MtBE (1.5 I) and once with MtBE (0.8 I). The organic layers were combined and washed twice with water (1.5 I), with brine (0.5 I), dried over MgSO4and concentrated under vacuum to give a brown oil (859 g) containing hydroxyfarnesylacetone (qNMR-purity: 45%, yield: 90%).

[0099] The crude mixture was used as is to prepare (5Z,9E)-10,14-dimethyl-6- (((trimethylsilyl)oxy)methyl)pentadeca-5,9,13-trien-2-one as described in Example 1 (Step 5).

[0100] Example 3: Preparation of (5Z,9E)-10,14-dimethyl-6-(((triethylsilyl)oxy)methyl)pentadeca- 5,9,13-trien-2-one:

[0101] A solution of crude hydroxyfarnesylacetone (14.4 g, 45%, 1 eq, 23.3 mmol) in tetrahydrofuran (150 mL) was treated successively with triethylamine (6.2 g, 99%, 2.6 eq, 60.6 mmol) and chlorotriethylsilane (8.5 g, 99%, 2.4 eq, 55 mmol) over 5 minutes at room temperature. The resulting mixture was stirred overnight at room temperature. The mixture was quenched with aqueous saturated NaHCO3(100 ml), the phases were separated and the organic phase was washed with a 2 : 1 mixture of water and brine (100 ml), dried over MgSO4and concentrated under vacuum to give a clear dark orange oil (20.9 g).

[0102] 5.0 g of the crude oil were purified by flash chromatography (Eluent: Heptane / MTBE) on SiO2yielding (5Z,9E)-10,14-dimethyl-6-(((triethylsilyl)oxy)methyl)pentadeca-5,9,13-trien-2-one (937 mg, GC-purity: 99%).

[0103] 15.7 g of the crude oil were purified by kugelrohr distillation at a pressure range of 0.05-0.017 mbar and a boiling point range of 200-240°C, yielding (5Z,9E)-10,14-dimethyl-6- (((triethylsilyl)oxy)methyl)pentadeca-5,9,13-trien-2-one (10.0 g, GC-purity: 87%).

[0104] Analytical data of (5Z,9E)- 10,14-dimethyl-6-(((triethylsilyl)oxy)methyl)pentadeca-5, 9,13-trien- 2-one:

[0105] 1H NMR (500 MHz, CHLOROFORM-d, 25°C): 5 = 5.18 (t, J = 7.3 Hz, 1 H), 5.05-5-14 (m, 2H), 4.16 (s, 2H), 2.44-2.51 (m, 2H), 2.32 (q, J = 7.3 Hz, 2H), 2.13 (s, 3H), 1.93-2.12 (m, 8H), 1.68 (d, J = 1.0 Hz, 3H), 1.60 (s, 3H), 1.59 (d, J = 1.2 Hz, 3H), 0.97 (t, J = 8.0 Hz, 9H), 0.62 ppm (q, J = 8.0 Hz, 6H) ppm.

[0106] 13C NMR (CHLOROFORM-d, 25° C) 5: 208.3, 139.7, 135.0, 131.2, 124.7, 124.3, 124.1 , 60.0, 43.9, 39.7, 34.7, 29.9, 26.8, 26.7, 25.7, 21.9, 17.6, 16.0, 6.8, 4.4 ppm.

[0107] GC-MS (El, 70 eV): 255 (23), 133 (52), 115 (36), 103 (23), 87 (31), 81 (22), 75 (35), 69 (100), 43 (60), 41 (61). Example 4: Preparation of (+)-Amberketal from TMS-HFA

[0108] (5Z,9E)-10, 14-dimethyl-6-(((trimethylsilyl)oxy)methyl)pentadeca-5,9, 13-trien-2-one (TMS E,Z- HFA) was cyclized to (+)-Amberketal with Bacillus megaterium squalene hopene cyclase variant BmeSHC#192. The reaction contained 135 g / l TMS E.Z-HFA, 182 g / l cells that had produced the SHC variant and 1.5 g / l SDS. The reactions (4 ml total volume) were run in 0.1 M succinic acid / NaOH buffer (pH 5.8) at 30°C under constant agitation. The reaction was started (t = 0) by adding the cells to the reaction mixture containing all other reaction components.

[0109] In Reaction 1 , the cells were added after stirring TMS E.Z-HFA for 24 h at 30°C in the presence of the reaction buffer (pH 5.8) and SDS. Reaction 2 was started without pre-treatment of TMS E,Z-HFA.

[0110] The results are shown in Figure 2. Under the conditions of Reaction 1 , TMS E.Z-HFA was first hydrolysed to hydroxyfarnesylacetone (E.Z-HFA), which was then cyclized to (+)-Amberketal by the SHC enzyme. Under the conditions of Reaction 2, TMS E.Z-HFA was hydrolyzed directly in the reaction mixture while produced E.Z-HFA was cyclized by the SHC enzyme to (+)- Amberketal. It was observed that (+)-Amberketal formation is faster when TMS E.Z-HFA is first hydrolysed to E.Z-HFA (Reaction 1) compared to the in situ deprotection / cyclization method without pre-treatment (Reaction 2).

[0111] Example 5: Preparation of (+)-Amberketal from different starting materials

[0112] The performance of TMS E,Z-HFA, (5Z,9E)-10,14-dimethyl-6-(((triethylsilyl)oxy)methyl)- pentadeca-5,9,13-trien-2-one (TES E,Z-HFA) and E,Z-HFA were compared in cyclization reactions run at constant molar concentration of E,Z-HFA. Bacillus megaterium squalene hopene cyclase variant BmeSHC#192 was used as catalyst. The reaction mixtures (4 ml total volume) contained 485 mM E,Z-HFA (135 g / l E,Z-HFA, 170 g / l TMS E,Z-HFA or 190 g / l TES E,Z-HFA) and 1.9 g / l SDS. They were run in 0.1 M succinic acid / NaOH buffer (pH 5.8) at 30°C under constant agitation. Reaction progress was followed by GC by measuring conversion of E,Z-HFA to (+)-Amberketal. (+)-Amberketal formation was slower using TMS E,Z-HFA as starting material because of the required hydrolysis to E,Z-HFA prior to cyclization by the SHC enzyme. However, total E,Z-HFA conversion to (+)-Amberketal was similar after 72 h of stirring. On the other hand, using TES E,Z-HFA as starting material, only very low levels of (+)- Amberketal were produced in the same time frame, which indicates that in situ hydrolysis of TES E,Z-HFA is rather slow or almost inexistent (Table 1).

[0113] Table 1. E.Z-HFA cyclization to (+)-Amberketal with three different E.Z-HFA feedstocks

[0114] Example 6: Preparation of (+)-Amberketal from TES-HFA after acidic pre-treatment

[0115] Reactions were run as outlined on Example 5 using TES E,Z-HFA as a starting material with the only exception that TES E,Z-HFA was first hydrolyzed prior to starting the cyclization reaction. TES E,Z-HFA was stirred at 30 °C in the presence of 1.5 g / l SDS in 0.1 M succinic acid (pH 2.6) for 5 min. The pH was set to 5.8 with 0.5 M NaOH and the SHC biocatalyst (cells that had produced SHC variant BmeSHC#192) was added to start the cyclization reaction. After 72 h, E,Z-HFA conversion to (+)-Amberketal was 82.6 %.

[0116] This result demonstrated that TES E,Z-HFA can be used as a feedstock for E,Z-H FA cyclization to (+)-Amberketal. However, compared to TMS E,Z-HFA, TES E,Z-HFA hydrolysis to E,Z-HFA needs to be done first under more acidic conditions (pH 2.6) than those available in the cyclization reaction itself (pH 5.8) where hydrolysis is only very poor as outlined in Example 5.

[0117] Cyclization of TES E,Z-HFA to (+)-Amberketal as described above at a concentration of 485 mM of E,Z-HFA showed 69.4% conversion after 72h. This result is very similar compared to the results described in Example 5 using E,Z-HFA as startig material.

Claims

Claims1 . A compound of formula (I)wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond.

2. The compound according to claim 1 , selected from the group consisting of 10,14- dimethyl-6-(((trimethylsilyl)oxy)methyl)pentadeca-5,9, 13-trien-2-one (compound of formula (la)), 10,14-dimethyl-6-(((triethylsilyl)oxy)methyl)pentadeca-5,9, 13-trien-2-one (compound of formula (lb)), 10,14-dimethyl-6-(((triisopropylsilyl)oxy)methyl)pentadeca- 5,9,13-trien-2-one (compound of formula (lc)), 6-(((tert-butyldimethylsilyl)oxy)methyl)- 10,14-dimethylpentadeca-5,9,13-trien-2-one (compound of formula (Id)), and 6-(((tert- butyldiphenylsilyl)oxy)methyl)-10,14-dimethylpentadeca-5,9,13-trien-2-one (compound of formula (le)).

3. The compound according to claim 1 or 2, wherein the compound has the double bond between C-9 and C-10 in E-configuration and the double bond between C-5 and C-6 in Z-configuration.

4. The compound according to any of claims 1 through 3 as starting material for the preparation of Amberketal.

5. Use of a compound of formula (I) according to claim 1wherein Ri, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond as a compound to release hydroxyfarnesylacetone.

6. The use of the compound of formula (I)wherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond, as starting material for the process of making Amberketal (compound of formula (II))wherein the compound of formula (I) releases hydroxyfarnesylacetone, which is further converted to the compound of formula (II).

7. The use according to claim 5 or 6, wherein the compound of formula (I) has the double bond between C-9 and C-10 in E-configuration and the double bond between C-5 and C-6 in Z-configuration.

8. A process of making Amberketal (compound of formula (II)) from the compound of formulawherein R-i, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond, by a SHC (squalene-hopene cyclase) mediated process.

9. The process according to claim 8, wherein the compound of formula (I) has the double bond between C-9 and C-10 in E-configuration and the double bond between C-5 and C-6 in Z-configuration.

10. The process according to claim 8 or claim 9, comprising the steps of: a) providing the compound of formula (I)wherein Ri, R2and R3 are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond; b) releasing hydroxyfarnesylacetone; and c) contacting hydroxyfarnesylacetone with SHC.11 . The process according to any of claims 8 to 10, wherein the compound of formula (I) is pretreated with acid.

12. A method of making the compound of formula (I)wherein R1, R2and R3are independently selected from C1-C4 alkyl and phenyl, wherein the wavy bonds are indicating an unspecified configuration at the adjacent carbon carbon double bond, comprising the steps of: a) providing [3-farnesene; b) epoxidizing [3-farnesene; c) reacting the epoxide of [3-farnesene with R4CI to obtain the corresponding R4-ether chloride, wherein R4CI is selected from the group consisting of TMSCI, TESCI and AcCI;d) forming the ketoester; e) optionally isolating the ketoester f) decarboxylating the ketoester g) subsequent re-protection the liberated alcohol group with R5CI to obtain the compound of formula (I), wherein R5CI corresponds to the chloride of Si(RiR2R3), wherein Ri, R2and R3are independently selected from C1-C4 alkyl and phenyl, h) optionally purifing the compound of formula (I), for example by fractional distillation.

13. The method according to claim 12, wherein the R4CI in step c) is selected from the group consisting of TMSCI, TESCI and AcCI.

14. The method according to claim 12 or 13, wherein the R5CI in step g) is TMSCI.

15. A process of making Amberketal according to any of claims 8 through 11 , wherein the compound of formula (I) is obtained by the method according to any of claims 12 through

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