WO2019092137A1

WO2019092137A1 - Enzymatic method for forming estolides

Info

Publication number: WO2019092137A1
Application number: PCT/EP2018/080671
Authority: WO
Inventors: Audrey ROBIC; Pascal AUFFRAY
Original assignee: Produits Chimiques Auxiliaires Et De Synthese
Priority date: 2017-11-08
Filing date: 2018-11-08
Publication date: 2019-05-16
Also published as: FR3073231A1; FR3073231B1; EP3707268A1

Abstract

The invention relates to a method for forming estolides from hydroxylated fatty acids characterised in that it is carried out in the presence of a lipase, in particular of Streptomyces avermitilis, preferably without a solvent, in a rapid and effective manner and with a higher number of EN estolides, preferably higher than 1.1. Optionally, the terminal acid function of the estolide can be protected by adding alcohol to the reaction mixture, optionally followed by the addition of an organic acid to the reaction mixture to protect the terminal hydroxyl function of the estolide, using a one-pot method.

Description

ENZYMATIC PROCESS FOR THE FORMATION OF ESTOLIDES

FIELD OF THE INVENTION The present invention relates to the preparation of estolides from hydroxylated fatty acids and more particularly to an enzymatic method using a lipase.

DESCRIPTION OF THE PRIOR ART

The estolides are polyesters derived from fatty acids. These are biodegradable compounds that are of industrial importance as lubricants, plasticizers, emulsifiers or moisturizers. They find their application in the automotive industry, cosmetics and food. This wide range of applications is related to their thermo-oxidative stability, their viscosity and their low melting point. The estolides also have biological and texturing properties and an adjustable hydrophilic / lipophilic balance. The estolides bring a moisturizing power to the products, they have a role of thermal barrier, they improve the elasticity of the fibers, and they give a shine effect. The estolides can represent up to 20% of the composition of a cosmetic product.

The preparation of estolides is mainly by chemical processes from unsaturated fatty acids in the presence of catalyst. The estolides formed by these processes are generally of low purity and require long and expensive purification steps.

For example, US Pat. No. 5,380,894 describes a chemical process for the formation of estolides from unsaturated fatty acids, in particular, where the unsaturations are localized in position Δ ⁵ and Δ ⁶ . No estolide is characterized in this patent. Also, WO2013 / 009471 discloses a chemical process for forming estolides from unsaturated fatty acids. Although the description is intended to be very generic, the examples are limited to the use of oleic acid. It should be noted that these chemical processes do not allow the regio-specific targeting of the bond between the fatty acids of these estolides but lead to estolides containing mixtures of regio-isomers. Of Moreover, these chemical processes require elaborate purification processes for products intended for food, cosmetic or pharmaceutical applications.

An alternative to lead to a better regio-specific homogeneity consists of hydroxylated fatty acids. For example, WO2008 / 040864 discloses a chemical process for forming estolides from hydroxylated fatty acids using essentially 12-hydroxystearic acid. The terminal acid function of the resulting estolide is then esterified in the presence of an alcohol. Also, the publication WO2011 / 037778 describes a process for the formation of estolides from hydroxylated fatty acids essentially using 12-hydroxystearic acid. The terminal acid function of the resulting estolide is then esterified in the presence of an alcohol and then the terminal alcohol function is protected with an aliphatic acid. Also, US Pat. No. 2,785,978 describes a process for the formation of estolides from hydroxylated fatty acids such as ricinoleic or 12-hydroxystearic acid. The terminal acid function of the resulting estolide is then esterified in the presence of polyglycerol. However, these processes are carried out at high temperatures of up to 210 ° C.

Esterification of hydroxylated fatty acids can also be carried out enzymatically. For example, US Pat. No. 7,125,694 describes the enzymatic esterification of 9,10-dihydroxystearic acid in the presence of Lypozyme ™ lipase and an alcohol. However, the presence of oligomers is not reported. Similarly, the publication US2013 / 0102041 (A1) describes the preparation of a double ester of ricinoleic acid ((9Z, 12R) -12-hydroxyoctadec-9-enoic acid) enzymatically from a methyl ester in the presence of Novozym ^® 435 lipase (lipase from Candida antarctica and immobilized) and a saturated fatty acid. Here again, it is not oligomers but only a hydroxylated fatty acid whose alcohol function and acid function are protected in ester form, because the product formed contains only one ricinoleate chain.

The preparation of estolides enzymatically from hydroxylated fatty acids has also been described. For example, Hayes describes the preparation of estolides from lesquerolic acid (14-hydroxy-eicosen-1-oic acid) (DG Hayes et al., JAOCS, vol 72, No. 11, 1309-1316 ( 1995)) in an aqueous medium. Only lipases from Pseudomonas sp., Candida rugosa and R. arrhizus allowed the preparation of oligomers. The yields of estolides with high degree of polymerization are low, the formation of monoestolide (with only one lesquerolic unit) is predominant, mainly due to the presence of octadecenoic acid mixed with lesquerolic acid. The reaction times are also very long, of the order of 65 hours. Esterification of the terminal acid function of these estolides is in a subsequent step. On the other hand, Horchani and Bodalo report the enzymatic preparation of estolides from ricinoleic acid in the presence of immobilized Staphylococcus xylosus lipase, respectively (H. Horchani et al., J. Mol., Cat.B Enz., Vol. 75, No. 11, 35-42 (2012)) or Candida rugosa lipase (Bodalo, A. et al, Biochem, Eng J, vol 44, 214-219 (2009)). Horchani reports a conversion rate of 65% in 48 hours in the presence of desiccant at 55 ° C; the non-immobilized enzyme is not thermostable (beyond 55 ° C). Bodalo reports the formation of estolides with, at best, an acid number of 57.5 mg KOH / g in 48 h; the enzyme is not thermostable (above 50 ° C). More recently, Greco-Duarte reports the enzymatic preparation of mono or diesters from polyols and ricinoleic acid resulting from the hydrolysis of triglycerides of castor oil (J. Greco-Duarte et al, Fuel Vol 202, 196-205 (2017)) in the presence of lipases such as Candida rugosa (Lipomod ™ 34 MDP), Candida antarctica (Novozym ^® 435) or Rhizopus miehei (Lipozyme RM IM ™). Greco-Duarte also reports the formation of estolides, mainly dimers in the presence of polyols and oligomers in the absence of polyols, only with the enzyme Lipomod ™ 34 MDP. Preparing estolides enzymatically from the 10S-hydroxy-8E-octadecenoic acid is also described in the presence of a lipase such as Novozym ^® 435 (from Candida antarctica and immobilized), Lipozyme RM IM ™ (from Rhizopus meihei and immobilized) or Lipozyme ™ TM IM (from Thermomyces lanuginosus and immobilized) (I. Martin-Arjol, J. Am., Chem Soc., vol 92, No. 8, 1125-1141 ( 2015)). The reactions are carried out however over long periods of 160 hours. The preparation of polyester-type estolides enzymatically from hydroxylated fatty acids, for example ricinoleic acid, methyl ricinoleate or a mixture of 12-hydroxystearic acid and 12-hydroxy-dodecanoic acid, is also described in the presence of a lipase such as Novozym ^® 435 (from Candida antarctica and immobilized) in the presence of a desiccating agent is also described (US2010 / 0204435; US8,729,176B2). The preparation of polyglycerol polyricinoleate estolides is described in two stages first comprising the esterification of ricinoleic acid catalyzed by a free or immobilized 1,3-nonspecific lipase, preferably derived from Candida rugosa, followed by esterification of the polyricinoleic acid with lipase catalyzed polyglycerol of Rhizopus oryzae or Rhizopus arrhizus, free or immobilized (WO2008 / 031908). The pot preparation of estolide of polyglycerol polyricinoleate has more recently been described by Ortega-Requena (2014) with the lipase Novozym ^® 435, lipase B from Candida antarctica immobilized (S. Ortega-Requena et al, Biochem. Eng. J 84, 91-97 (2014)).

By modifying the reaction conditions, it is also possible to prepare enzymatically mixtures of macrolide lactones and estolides from 12-hydroxystearic acid or ricinoleic acid or else 16-hydroxy-acid. palmitic in the presence of a lipase such as that from Pseudomonas fluorescens or Pseudomonas stutzeri in toluene at 40 ° C. The formation of oligomer mixtures containing up to octamers has thus been observed (A. Todes, Pure Appl. Chem., 87 (1), 51-58 (2015)).

None of these documents implements a lipase of Streptomyces avermitilis for the enzymatic synthesis of estolides, whether esters or oligomers. Such a lipase was studied by B. Wang in 2010, but only its hydrolytic activity has been demonstrated. It is suggested that this lipase may also be useful in the food field or for the synthesis of pharmaceuticals (B. Wang, Acta Microbiol, Sinica, 50 (2), 236-243 (2010)).

There is a need to develop rapid formation processes of estolides under mild, solvent-free or desiccant and high yielding conditions. There is a need to develop estolide forming processes using a high temperature stable lipase without having to immobilize the solid support lipase. There is also a need to develop processes capable of producing estolides with a high degree of purity in a simple and inexpensive manner. There is a need to develop rapid and effective biological processes for the formation of estolides, especially for their use in the cosmetic, pharmaceutical and food industry. There is also a need to develop processes capable of producing estolides having a high number of estolides greater than 1.1, an acid number of less than 60 mgKOH / g, a viscosity at 40 and 100 ° C varies depending on the application but with the lowest possible viscosity index, a pour point of less than -20 ° C and a good resistance to oxidation. There is also a need to develop monotopic processes capable of producing estolides whose hydroxyl and terminal acid functions are protected. SUMMARY OF THE INVENTION

It is therefore an object of the present invention to produce estolides from hydroxylated fatty acids in the presence of a lipase of Streptomyces avermitilis. The process can be conducted without a solvent, at a temperature chosen between 20 ° C and 95 ° C depending on the starting hydroxylated fatty acid, without the addition of desiccant. The temperature is selected so that the starting hydroxylated fatty acid is in the liquid phase. Advantageously, the lipase of Streptomyces avermitilis is thermostable and can therefore be used in the process at a high temperature (between 40 ° C. and 95 ° C.) without immobilization on a solid support. The process allows the formation of estolides with a high (or even greater than or equal to 1.1) solids number in less than 48 hours, even more preferably in 24 hours or less. Advantageously, the process allows the formation of estolides with a number of estolides greater than or equal to 2.0, or greater than or equal to 3.0. Advantageously, the estolide formed has a number of estolides having a value of 1.5 to 2.0, or 2.0 to 2.5, or 2.5 to 3.0, or 3.0 to 3.5 , or 3.5 to 4.0, or 4.0 to 4.5, or 4.5 to 5.0. Optionally, the process may be conducted in the presence of an aliphatic or aromatic hydrocarbon or ether type apolar solvent.

Optionally, an alcohol is added to the reaction medium to esterify the terminal acid function of the estolide once the acid number is below 60. In general, the alcohol is added at a rate of 15 to 35% equivalent of the initial hydroxylated fatty acid. Preferably, the alcohol is added at a level of 15 to 20%, or 20 to 25%, or 25 to 30%, or 30 to 35%, or about 25% equivalent. of the initial hydroxylated fatty acid.

Optionally also, an organic acid is added to the reaction medium to protect the free hydroxyl function of the estolide once the acid number of the estolide whose terminal acid function is esterified is below 60, or once that the index The acid acid of the estolide whose terminal acid function is not esterified is between 10 and 40. In general, the acid is added at a level of 15 to 35% equivalent of the initial hydroxylated fatty acid. Preferably, the acid is added at a level of 15 to 20%, or 20 to 25%, or 25 to 30%, or 30 to 35%, or about 25% equivalent of the initial hydroxylated fatty acid.

DESCRIPTION OF THE FIGURES

FIG. 1 is a diagram of the number of estolides (EN) as a function of the lipases evaluated in the presence of ricinoleic acid as detailed in Example 2.

FIG. 2 is a diagram of the number of estolides (EN) as a function of the lipases evaluated in the presence of 12-hydroxystearic acid as detailed in Example 7.

FIG. 3 is a diagram of the number of estolides (EN) for two lipases as a function of time in the presence of ricinoleic acid at 40 ° C. as detailed in Example 3.

FIG. 4 is a diagram of the number of estolides (EN) for two lipases as a function of time in the presence of ricinoleic acid at 50 ° C. as detailed in Example 3.

FIG. 5 is a diagram of the number of estolides (EN) for two lipases as a function of the temperature in the presence of ricinoleic acid at 24 hours as detailed in Example 3.

SETTINGS

The acid number (IA) is calculated as the mass of KOH (mg) required to neutralize 1 g of acid or reaction sample.

The EN value represents the number of hydroxylated fatty acids added to a starting hydroxylated fatty acid such as a dimer having an EN of 1, a trimer having an EN of 2, and an equimolar mixture of dimer and trimer having an EN of 1.5 . The number of estolides (EN) is therefore the average number of hydroxylated fatty acids added to the starting hydroxylated fatty acid (AGH). EN is calculated according to the formula:

MW KOH x 1000

EN =

IA x MW AGH Where IA is the acid number, MW KOH is the molar mass of potash, and MW AGH is the molar mass of hydroxylated fatty acid.

The viscosity is measured according to ASTM standard D445 which consists in measuring the flow time of a volume of liquid contained in a calibrated glass capillary tube at a given temperature (40 ° C. and 100 ° C. for lubricants).

The pour point is measured according to ASTM D97 which consists of measuring the lowest temperature at which a movement of a liquid sample is observed.

DETAILED DESCRIPTION OF THE INVENTION It has been advantageously demonstrated by the efforts of the present inventors that a lipase of Streptomyces avermitilis may also be used in esterification, in particular on hydroxylated fatty acids, rapidly and efficiently leading to the formation of estolides. with high conversion rates and estolide numbers (EN). This lipase demonstrated a rather unexpected high catalytic activity with fast conversion rates. This lipase thus appears more advantageous than those known from the literature reported above. In addition, this lipase has high stability at high temperatures, even without being immobilized on a solid support, making it possible to prepare hydroxylated fatty acid estolides with a high melting temperature, such as up to 95 ° C.

It is therefore an object of the present invention to produce estolides from hydroxylated fatty acids in the presence of a lipase of Streptomyces avermitilis. The process can be carried out without a solvent, especially at a temperature chosen between 20 ° C. and 95 ° C. depending on the starting hydroxylated fatty acid, without the addition of desiccant. The temperature is selected so that the starting hydroxylated fatty acid is in the liquid phase. Thus, the reaction is conveniently initiated at a temperature above the melting point of the hydroxylated fatty acid. The process allows the formation of estolides with a rapidly increasing number of estolides (EN) in less than 48 hours, even more preferably in 24 hours or less.

Optionally, the process may be conducted in the presence of an aliphatic or aromatic hydrocarbon or ether type apolar solvent. For example, the solvent may be toluene, methyl tert-butyl ether (MBTE), hexane, isooctane, or diisopropyl ether. In particular, the use of a solvent is advantageous when the starting hydroxylated fatty acid has a high melting point, such as above 95 ° C.

Optionally, an alcohol is added to the reaction medium to esterify the terminal acid function of the estolide once the acid number is below 60. The amount of alcohol to be added is evaluated as a function of the d residual acid. The lower the acid number, the smaller the amount of alcohol to add. In general, the alcohol is added at a level of 15 to 35% equivalent of the initial hydroxylated fatty acid. Preferably, the alcohol is added at a level of 15 to 20%, or 20 to 25%, or 25 to 30%, or 30 to 35%), or about 25% equivalent of the initial hydroxylated fatty acid.

Optionally also, an organic acid is added to the reaction medium to protect the free hydroxyl function of the estolide once the acid number of the estolide whose terminal acid function is esterified is between 10 and 40 inclusive, or once the acid number of the estolide whose terminal acid function is not esterified is below 60. The amount of acid to be added is evaluated as a function of the residual acid number. The lower the acid number, the smaller the amount of acid to add. In general, the acid is added at a level of 15 to 35% equivalent of the initial hydroxylated fatty acid. Preferably, the acid is added at a level of 15 to 20%, or 20 to 25%, or 25 to 30%, or 30 to 35%, or about 25% equivalent of the hydroxylated initial fatty acid. It is therefore an object of the present invention to provide a method for producing estolides in the presence of a lipase of Streptomyces avermitilis, these estolides advantageously having a number of estolides (EN or n-1) greater than or equal to 1, 1. In particular, the enzymatic process allows the preparation of estolides (formula II) from hydroxylated fatty acids (formula I) according to the following scheme (1):

(1) n HO-CHRi-Q-COOH + Lipase → H- (O-CHRi-Q-CO) _n -OH

I II

Where R 1 is selected from saturated or mono-unsaturated hydrocarbon groups, linear or branched from one to twelve carbon atoms (Cl to Cl 2) optionally substituted with a hydroxyl group, Q is a divalent non-cyclic aliphatic group saturated or monounsaturated from six to twelve carbon atoms (C6 to Cl2) optionally substituted by a hydroxyl group, and n is a value greater than or equal to 2.1, preferably greater than or equal to 2.5.

It is also an object of the present invention to provide a monotopic method for producing ester estolides, whose terminal acid function is protected in the presence of a lipase of Streptomyces avermitilis, these estolides advantageously having a number of estolides (EN or n-1) greater than or equal to 1.1. In particular, the enzymatic process allows the preparation of estolides (formula V) from hydroxylated fatty acids (formula I) in the presence of an alcohol (formula IV) according to scheme (2):

(2) n HO-CHRi-Q-COOH + HOR ₃ + Lipase → H- (O-CHRi-Q-CO) _n- OR ₃

I IV V

Where R 1 is selected from saturated or mono unsaturated hydrocarbon groups, linear or branched from one to twelve carbon atoms (Cl to Cl 2) optionally substituted by a hydroxyl group, R ₃ is selected from (saturated) alkyl groups, linear or branched from one to twenty-two carbon atoms (C1 to C22), optionally substituted with one or more, especially 1 to 7, hydroxyl groups, or radicals of polyether alcohols (such as polyglycerol or polyalkylene glycol), radicals wherein polyether alcohols are selected such that the HOR ₃ molecule is a polyether alcohol, Q is a divalent saturated or monounsaturated non-cyclic aliphatic group of six to twelve carbon atoms (C6 to C12), optionally substituted by a hydroxyl group, and n is a value greater than or equal to 2.1, preferably greater than or equal to 2.5.

It is also an object of the present invention to provide a monotopic process for producing ester estolides, whose terminal acid function and terminal hydroxyl function are protected in the presence of a lipase of Streptomyces avermitilis, these estolides advantageously having a number of (EN or n-1) greater than or equal to 1.1. In particular, the enzymatic process allows the preparation of estolides (formula VI) from hydroxylated fatty acids (formula I) in the presence of an acid. organic (formula III) and an alcohol (formula IV) according to scheme (3): (3) n HO-CHRi-Q-COOH + R ₂ -COOH + HOR ₃ + Lipase → R ₂ -CO- (O-CHRi-Q-CO) _n -OR ₃ I III IV VI

Where R 1 is selected from saturated or mono-unsaturated hydrocarbon groups, linear or branched from one to twelve carbon atoms (Cl to Cl 2) optionally substituted by a hydroxyl group, R ₂ is selected from saturated or unsaturated hydrocarbon groups, linear or branched from one to twenty-two carbon atoms (Cl to C22), R ₃ is selected from linear or branched (saturated) alkyl groups of one to twenty-two carbon atoms (C1 to C22), optionally substituted by one or more, especially 1 to 7, hydroxyl groups, or radicals of polyether alcohols, the radicals of polyether alcohols being chosen such that the molecule HOR ₃ represents a polyether alcohol, Q is a divalent non-cyclic aliphatic saturated or mono unsaturated group from six to twelve carbon atoms (C6 to C12), optionally substituted by a hydroxyl group, and n is a value greater than or equal to 2.1, preferably greater than 2.5 or less. In Schemes (1), (2) and (3) above, for the compounds of formulas I to VI, the variables R 1, R ₂ , R ₃ , Q, and n can be more particularly defined as follows.

For example, R 1 can be selected from the following groups: -CH ₃ , -C ₂ H ₅ , -C ₃ H ₇ , -C ₄ H ₉ , -C ₅ H 11, -C ₆ H ₃ , -C ₇ H ₁₅ , -C ₈ H ₁₇ , -C ₉ H 19, -CioH ₂ i, -C nH ₂₃ , -Ci ₂ H ₂₅ , -CH = CH ₂ , -CH = CH-CH ₃ , -CH = CH-C ₂ H ₅ , -CH = CH-C ₃ H ₇ , -CH = CH-C ₄ H ₉ , -CH = CH-C ₅ H n, -CH = CH-C ₆ H 3,

-CH = CH-CioH ₂ i, -CH ₂ -CH = CH-CH ₃ , -CH ₂ -CH = CH-C ₂ H ₅ , -CH ₂ -CH = CH-C ₃ H ₇ , -CH ₂ - CH = CH-C ₄ H ₉ , -CH ₂ -CH = CH-C ₅ H n, -CH ₂ -CH = CH-C ₆ H ₃ , -CH ₂ -CH = CH-C ₇ H ₅ ,

-C ₂ H ₄ -CH = CH-CH ₃ , -C ₂ H ₄ -CH = CH-C ₂ H ₅ , -C ₂ H ₄ -CH = CH-C ₃ H ₇ , -C ₂ H ₄ -CH = CH-C ₄ H ₉ , -C ₂ H ₄ -CH = CH-C ₅ H n, -C ₂ H ₄ -CH = CH-C ₆ H ₃ , -C ₂ H ₄ -CH = CH-C ₇ H ₅ ,

-CH ₂ -CH (OH) -CH ₃ , -CH ₂ -CH (OH) -C ₂ H ₅ , -CH ₂ -CH (OH) -C ₃ H ₇ , -CH ₂ -CH (OH) -C ₄ H ₉ , -CH ₂ -CH (OH) -C ₅ Hn, -CH ₂ -CH (OH) -C ₆ H ₃ , -CH ₂ -CH (OH) -C ₇ H ₅ , -CH ₂ -CH (OH) -C ₈ H ₇ , -CH ₂ -CH (OH) -C ₉ H ₉ , -CH ₂ -CH (OH) -C ₁₀ OH-C ₂ H ₄ -CH (OH) -CH ₃ , -C ₂ H ₄ -CH (OH) -C ₂ H ₅ -C ₂ H ₄ -CH (OH) -C ₃ H ₇ -C ₂ H ₄ -CH (OH) -C ₄ H ₉ -C ₂ H ₄ -CH (OH) -C ₅ Hn, -C ₂ H ₄ -CH (OH) -C ₆ H ₃ , -C ₂ H ₄ -CH (OH) -C ₇ H ₅ , -C ₂ H ₄ -CH (OH) -C ₈ H ₇ and -C ₂ H ₄ -CH (OH) -C ₉ H ₉ . For example, R ₂ may be selected from the following groups: linear alkyl Cl to C22 (saturated) such as -CH _3, -C ₂ H _5, -C3H7, -C4H9, -C5H11, -C ₆ Hi ₃ Hi ₇ -C ₅ -C ₈ Hi _7, - C9H19, -CioH ₂ i, -CnH _23, -C ₂ H _25, -C ₂₇ H _3, -C ₂ H ₄ 9, -C ₅ H ₃ i, -C ₆ H _33, -C ₇ H _35, -CisH ₃₇ - Ci9H _{March 9,} -C ₂₀ H _41, -C ₂₁ H ₄ 3, -C ₂₂ H _45; alkyl branched C2 to C22 (saturated) of the formula - C _r H _2r CH (CsH _2s + i) Cth ₂ t + i where r is an integer from 0 to 19, s and t are integers from 1 to 20 and the sum r + s + t varies from 2 to 21; and C2-C22 alkenyl (unsaturated) of the formula -C _u H _2u -CH = CH ₂ or -C _m H _2m- (CH = CH-CH ₂ ) p-CqH _2q + i where u is an integer of 0 at 20, m and q are integers from 0 to 19, and p is an integer of 1 to 3.

For example, R ₃ is selected from the following groups: a linear alkyl (saturated), such as -CH _3, -C ₂ H _5, -C ₃ H _7, -C4H9, -C5H11, -C ₆ Hi _3, -C ₇ Hi ₅ -C ₈ Hn, -C9H19, -G0H21, -CnH _23, -C ₂ H _25, -C ₂₇ H _3, -C ₂ H ₄ 9, -C ₅ H i _3, -C ₆ H ₃₃ , -C ₇ H _35, -CisH _37, -Ci9H _{September 3,} -C ₂ oH ₄ i, -C ₂ iH ₄₃ - C ₂₂ H _45; a branched (saturated) alkyl of the formula -C _r H _2r -CH (CsH _2s + 1) CtH ₂ t + i where r is an integer from 0 to 9, s and t are integers of 1 to 20 and the sum r + s + t varies from 2 to 22; said linear or branched alkyl being optionally substituted with 1 to 7 hydroxyls; a polyether alcohol radical (such that R ₃ OH corresponds to the polyether alcohol) formed from one or more, in particular 1 or 2, of said linear or branched alkyls substituted with 2 to 7 hydroxyls as monomers. Preferentially, R ₃ is selected from the following groups: methyl, ethyl, n-propyl, iso-propyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, 2 methylpentyl, 3-methylpentyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl, heptyl, octyl, 2-ethylhexyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl , nonadecyl, eicosyl, henicosyl, docosyl, isooctadecyl, 2-butyloctyl, 2-octyldodecyl, 2-hexyldecyl, 2-octyldecyl, 2-hydroxyethyl, 2-hydroxypropyl, 2,3-bis (hydroxy) propyl, 3-hydroxy (2,2-bis (hydroxymethyl) propyl, 2,3,4-ter (hydroxy) butyl, 2,3,4,5-tetra (hydroxy) pentyl, 2,3,4,5,6-penta (hydroxy hexyl, a polyetheralcohol radical formed from: ethylene glycol, 1,2-propyleneglycol, 1,3-propyleneglycol, 1,4-butanediol, pentamethyleneglycol, hexamethyleneglyco 1,1,1,1-tris (hydroxymethyl) methane, 1, 1, 1- tris (h ytroxymethyl) ethane, 1,1,1-tris (hydroxymethyl) propane, glycerol, thetitol, tetritol, erythritol, mesoerythritol, xylitol, ribitol, arabinitol, arabitol, pentitol, pentaerythritol, mannitol, allitol, altritol, hexitol, glucitol, sorbitol, dulcitol, volemitol, or a mixture thereof as the monomer (s). Preferably, the polyether alcohol radical is a polyglycerol radical (so that R 3 OH is polyglycerol), polyerythritol (so that R 3 OH is polyerythritol), polypentaerythritol (so that R 3 OH is polypentaerythritol), polymannitol (so that that R 3 OH is polymannitol), a polyalkylene glycol (such that R 3 OH is polyalkylene glycol) or a copolymer of alkylene glycols (so that R 3 OH is the copolymer of alkylene glycols). Preferably, the polyether alcohol radical is a polyglycerol radical, a polypentaerythritol radical, and a polyalkylene glycol radical (for example poly ((Ci-C8) alkylene glycol), in particular poly ((Ci-C6) alkylene glycol). preferably poly ((C 1 -C 3) alkylene glycol) such as polyethylene glycol PEG or polypropylene glycol PPG) or a copolymer of alkylene glycols, especially of two alkylene glycols (for example from (C ₁ -C ₈₎ alkylene glycols, especially (C ₁ -C ₆ ) alkylene glycols, preferably (C 1 -C 3) alkylene glycols - it may be for example a copolymer of ethylene glycol and propylene glycol).

Q may be selected from the following divalent groups: -C ₆ Hi2-, -C ₇ H ₁₄ -, -C ₈ Hi ₆ -, -C9H18-, -C10H20-, -C11H22-, -C12H24-, -CH = CH -C ₄ H ₈ -, -CH = CH-C ₅ H 10-, -CH = CH-C ₆ Hi 2 -, -CH = CH-C ₇ Hi 4,

-CH = CH-CioH ₂₀ -, -CH ₂ -CH = CH-C ₃ H ₆ -, -CH ₂ -CH = CH-C ₄ H ₈ -, -CH ₂ -CH = CH-C ₅ H 10-, -CH ₂ -CH = CH-C ₆ Hi ₂ -, - CH2-CH = CH-C ₇ Hi4-,

-C ₂ H ₄ -CH = CH- C2H4-, -C2H ₄ -CH = CH-C ₃ H ₆ -, -C ₂ H ₄ -CH = CH-C ₄ H ₈ -, -C2H ₄ -CH = CH -C ₅ H 10-, -C ₂ H 4 -CH = CH-C ₆ H 2 -, -C ₂ H ₄ -CH = CH-C ₇ Hi 4,

-CH ₂ -CH (OH) -C ₄ H ₈ -, - CH ₂ -CH (OH) -C ₅ H 10-, -CH ₂ -CH (OH) -C ₆ H ₂ -, -CH ₂ -CH ( OH) -C ₇ H ₄ -, -CH ₂ -CH (OH) - CsHie-, -CH ₂ -CH (OH) -C ₉ H ₈ -, -CH ₂ -CH (OH) -CioH ₂₀ -, -C 2H ₄ -CH (OH) -C ₃ H ₆ -, -C ₂ H ₄ -CH (OH) -C ₄ H ₈ -, -C ₂ H ₄ -CH (OH) -C ₅ H 10-, -C ₂ H ₄ -CH (OH) Hi2- -C ₆ alkyl, -C ₂ H ₄ -CH (OH) -C ₇ Hi ₄ -, -C ₂ H ₄ -CH (OH) -C ₈ Hi6- and -C ₂ H ₄ -CH (OH) - C ₉ Hi ₈ -. n is a value selected from 2.1 to 8.0 by incrementing by 0.1 (ie 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2 , 8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0 , 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5 , 3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5 , 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7 , 8, 7.9, or 8.0). Advantageously, n is a value of 2.1 to 2.5, or 2.5 to 3.0, or 3.0 to 3.5, or 3.5 to 4.0, or 4.0 to 4.5 , or 4.5 to 5.0, or 5.0 to 5.5, or 5.5 to 6.0, or 6.0 to 6.5, or 6.5 to 7.0, or 7.0 at 7.5, or 7.5 to 8.0. Advantageously, n is 3.0 to 6.0 or 3.5 to 5.5. Preferably, the lipase is the lipase LpsA2 isolated from the strain Streptomyces avermitilis having in particular the accession number WP O 10987960 of the National Center for Biotechnology Information, of the US National Library of Medicine with the amino acid sequence SEQ ID NO. 2 next:

MLP WK VLRPLT ALLLT V AV AL AAT VP AH AD SAP S S G WND YS CK

PSAAHPRPVVLVHGTLGNSVDNWLGLAPYLEHRGYCVFSLDYGQ

LSGVPFFHGLGPIDKSAEQLQVFVDKVLTATGATKADLVGHSQGG

MMPRYYLKFLGGAGKVNALVGIAPNNHGTTLSGLTNLLPYFPGAE

DLLSTATPGLADQVVGSAFMAKLNAGGDTVAGVHYTVIATQYDE

VVTPYRTAFLSGSDVHNVLLQDLCPLDLSEHVAIGLIDRIAFHEVTN

ALDPAHATRTTCASVFS.

As an alternative, the lipase used in the process according to the invention may also be a lipase with a sequence having at least 85%, in particular at least 86%, at least 87%, at least 88%, at least 89% at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%), at least 99% %, at least 99.3%, or at least 99.6% identity with the sequence SEQ ID NO. 2.

The percentages of identity to which reference is made in the context of the disclosure of the present invention are determined on the basis of an overall alignment of the sequences to be compared, that is to say on an alignment of the sequences taken in their entirety over the entire length using any algorithm well known to those skilled in the art such as the Needleman and Wunsch-1970 algorithm. This sequence comparison can be performed using any software well known to those skilled in the art: for example needle software using the parameter "Gap open" equal to 10.0, the "Gap extend" parameter equal to 0.5 and a "Blosum 62" matrix. The needle software is for example available on the ebi.ac.uk world wide website under the name "Align".

When the lipase according to the invention has an amino acid sequence which is not 100% identical to the reference sequence SEQ ID NO: 2 but which has at least 85%, in particular at least 86%, at least 87%, at least 88%, at least 89%, at least 90%), at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least less than 96%, at least 97%, at least 98%>, at least 99%, at least 99,3%>, or at least 99,6%) of identity with such a reference sequence, it may have insertions, deletions or substitutions with respect to the reference sequence. When it comes to substitutions, the substitution is preferably carried out by an "equivalent" amino acid, that is to say any amino acid whose structure is close to that of the original amino acid and is therefore unlikely to alter the biological activities of lipase. Examples of such substitutions are presented in the following table:

Alternatively or in combination with one or more of the features described above for lipases that have an amino acid sequence that is not 100% identical to the SEQ ID NO: 2 reference sequence, the sequence of the lipase used in the process according to the invention has at least one of the following characteristics, and preferably all of the following characteristics, after optimal global alignment with SEQ ID NO: 2:

the amino acids of the lipase sequence aligned with positions 43 and 80 of SEQ ID NO: 2 are identical to the amino acids of SEQ ID NO: 2 at positions 43 (C) and 80 (C) respectively, these amino acids for forming a first disulfide bridge;

the amino acids of the lipase sequence aligned with the positions 128-132 of SEQ ID NO: 2 are identical to the amino acids of SEQ ID NO: 2 at positions 128-132, so as to form a conserved pentapeptide structure of sequence GHSQG (SEQ ID NO: 3);

the amino acids of the lipase sequence aligned with the positions 130, 221 and 253 of SEQ ID NO: 2 are identical to the amino acids of SEQ ID NO: 2 at positions 130 (S), 221 (D) and 253 ( H) respectively, these amino acids corresponding to the 3 amino acids of the active site of the lipase;

the amino acids of the lipase sequence aligned with the positions 246 and 281 of SEQ ID NO: 2 are identical to the amino acids of SEQ ID NO: 2 at positions 246 (C) and 281 (C) respectively, these amino acids to form a second disulfide bridge.

Alternatively or in combination with one or more of the features described above relating to lipases that have an amino acid sequence that is not 100% identical to the reference sequence SEQ ID NO: 2, the lipase comprises a sequence of 256 amino acids having at least 90%>, especially at least 91%>, at least 92%>, at least 93%>, at least 94%>, at least 95%>, at least 96%>, at least minus 97%>, at least 98%>, at least 99%>, at least 99.2%>, at least 99.6%>, or even 100% identity with positions 31-286 of SEQ ID NO 2. In fact, amino acids 1-30 of the sequence SEQ ID NO: 2 correspond to the signal peptide of the LpsA2 lipase and are therefore not important for the activity of the lipase. A lipase comprising a sequence of 256 amino acids 100% identical to positions 31-286 of SEQ ID NO: 2 should therefore have the same activity as SEQ ID NO: 2 and therefore corresponds to a preferred embodiment of the invention. Such a lipase may or may not include an N-terminal signal peptide, which may be different from that corresponding to amino acids 1- Of SEQ ID NO: 2. Those skilled in the art may select such a signal peptide from those known in the art.

According to a preferred embodiment of the invention, the lipase sequence used in the process according to the invention will have the following characteristics after optimal global alignment with SEQ ID NO: 2:

- at least 85%, including at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% %, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.3%, or at least 99.6% identity with the sequence SEQ ID NO . 2;

the presence of a 256-amino acid sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.2%, at least 99.6%, or even 100% identity with positions 31-286 of SEQ ID NO: 2;

the amino acids of the lipase sequence aligned with the positions 43, 80, 128-132, 221, 246, 253 and 281 of SEQ ID NO: 2 are identical to the amino acids of SEQ ID NO: 2 at positions 43 ( C), 80 (C), 128-132 (GHSQG pentapeptide, SEQ ID NO: 3), 221 (D), 246 (C), 253 (H) and 281 (C) respectively; and

optionally, one or more substitutions (advantageously all the substitutions) with respect to SEQ ID NO: 2 is (are) preferably carried out by an "equivalent" amino acid, as defined above .

According to a particularly preferred embodiment of the invention, the lipase sequence used in the process according to the invention will have the following characteristics after optimal global alignment with SEQ ID NO: 2:

- at least 85%, including at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% %, at least 95%, at least 96%, at least 97%, at least 98%, at least minus 99%, at least 99.3%, or at least 99.6% identity with the sequence SEQ ID NO. 2; and

the presence of a 256 amino acid sequence having 100% identity with positions 31-286 of SEQ ID NO: 2 (this ensures that the amino acids of the lipase sequence aligned with positions 43, 80, 128-132, 221, 246, 253 and 281 of SEQ ID NO: 2 are identical to the amino acids of SEQ ID NO: 2 at positions 43 (C), 80 (C), 128-132 (GHSQG pentapeptide, SEQ ID NO : 3), 221 (D), 246 (C), 253 (H) and 281 (C) respectively).

In general, the lipase can be produced in recombinant form in expression systems such as bacteria, yeasts or fungi. Recombinant protein expression systems are known to those skilled in the art and include, for example, Bacillus strains (eg, B. subtilis), Escherichia strains (eg, E. coli), Pseudomonas strains (eg, P. aeruginosas). , Saccharomyces (eg, S. cerevisiae), Yarrowia (eg, Y. lipolytica), Pichia (eg, P. pastoris), Trichoderma (eg, T. reesei), Aspergillus. The lipase is preferably produced in recombinant form in a host cell, such as Escherichia coli, by methods well known to those skilled in the art, and preferably it is produced as described in Example 1. The lipase may also be advantageously be produced in recombinant form in a Yarrowia or Pichia host cell for applications in the food industry, such as Yarrowia lipolytica or Pichia pastoris. The lipase can also be immobilized on a support so that it can be recycled. For example, the lipase may be deposited on a resin or a mineral support. For example, resins can be selected from polystyrene as Lewatit ^® VP OC 1064 MD-PH from Lanxess, a copolymer of styrene and divinylbenzene such as Diaion ^® HP20 home Resindion Sri (Binasco, Italy), polymethacrylate methyl such as Diaion ^® HP2MG home Resindion Sri (Binasco, Italy), phenol-formaldehyde such as AMBERLITE ^® XAD761 from Rohm and Haas, polyvinyl chloride, polyethylene polyamides, poly silicon (hydroxyethyl) methacrylate, the lignin, or polysaccharides of the starch type, cellulose, or ethyl cellulose. Among the mineral supports, calcium carbonate, activated carbon, among others, can be used as immobilization supports. The supports and techniques for immobilizing enzymes are well known to those skilled in the art, such as those described in publication WO2016 / 151115. The methods according to the present invention can use the lipase at a level of 5 to 10% by weight, such as 5, or 6, or 7, or 8, or 9, or 10% by weight, expressed in terms of ratio of the mass of dry cells containing the lipase used on the mass of the hydroxylated starting acid. For example, the methods according to the present invention can use a wide range of hydroxylated fatty acids of formula (I) such as 5-, 6-, or 7-hydroxy-eicosanoic acid; 8-, 9-, 10-, or 11-hydroxy-octadecanoic acid; 7-, 8-, or 9-hydroxy-hexadecanoic acid; 11-, 12-, or 13-hydroxy docosanoic acid; 12-, or 13-hydroxy-5-docosenoic acid (all described in US Patent 5,380,894); 12,13,17-trihydroxy-9 (Z) -octadecenoic acid (described in US Pat. No. 5,852,196); 10-hydroxystearic acid (described in US Pat. No. 4,582,804); 12-hydroxystearic acid (described in Patent FR2906530); 7,10-dihydroxy-8-octadecenoic acid (described in US Patent 5,900,496); ricinoleic acid (with the hydroxyl at position 12, described in patent FR2942628); 8-, 9-, 10, 11-, or 12-hydroxy-lauric acid, 8-, 9-, 10, 11-, or 12-hydroxy-tridecyl acid, 8-, 9-, , 10, 11-, 12-, or 13-hydroxy-myristic acid, 8-, 9-, 10,11-, 12-, or 13-hydroxy-pentadecyl acid, 8-, 9-, 10-, ,

11-, 12-, or 13-hydroxy-palmitic acid, 8-, 9-, 10,11-, 12-, or 13-hydroxy-margaric acid, 8-, 9-, 10-, 11-, , 12-, or 13-hydroxy-stearic acid, 8-, 9-, 10, 11-,

12-, or 13-hydroxy-nonadecyl, 8-, 9-, 10,11-, 12-, or 13-hydroxy-arachidic acid, 8-, 9-, 10, 11-, 12-, , or 13-hydroxy-heneicosanoic acid, 8-, 9-,

10, 11-, 12-, or 13-hydroxy-behenic acid, 8-, 9-, 10,11-, 12-, or 13-hydroxytricosanoic acid, 8-, 9-, 10-, 11-, 12-, or 13-hydroxy-lignoceric acid, 8-, 9-, 10,11-, 12-, or 13-hydroxy-pentacosanoic acid, 8-, 9-, 10, 11- , 12-, or 13-hydroxy-cerotic acid, 8-, 11-, 12-, or 13-hydroxy-myristoleic acid, 8-, 1 1-, 12-, or 13-hydroxy-palmitoleic acid, 8-, 9-, 10,11-, 12-, or 13-hydroxy-sapienic acid, 8-, 11-, 12-, or 13-hydroxy-oleic acid, 10-hydroxy- 12-octadecenoic acid, 13-hydroxy-9-octadecenoic acid, 8-, 11-, 12- or 13-hydroxy-elaidic acid, 8- or 11-hydroxy-linoleic acid, 11 hydroxy-linolelaic acid, 8- or 11-hydroxy-alpha-linolenic acid, 8- or 11-hydroxy-gammalinolenic acid, 8-, 9-, 10, 11-, or 12-hydroxy-acid. erucic, 8-, 9-, 10,11-, 12-, or 13-hydroxy-nervonic acid, 10,13-dihydroxy-myristic acid, 10,13-dihydroxy-pentadecyl acid, 10,13-dihydroxy-palmi acid 10,13-dihydroxy-margaric acid, 10,13-dihydroxy- stearic acid, 10,13-dihydroxy-nonadecyl acid, 10,13-dihydroxy-arachidic acid, 10,13-dihydroxy-heneicosanoic acid, 10,13-dihydroxy-behenic acid, 10 , 13-dihydroxy-tricosanoic acid, 10,13-dihydroxy-lignoceric acid, 10,13-dihydroxy-pentacosanoic acid, or 10,13-dihydroxy-cerotic acid, individually or a mixture of them. Preferably, the processes according to the present invention can use the hydroxylated fatty acids of formula (I) such as 10-hydroxystearic acid, 12-hydroxystearic acid, 13-hydroxystearic acid, ricinoleic acid, 10-hydroxy acid 12-octadecenoic, 13-hydroxy-9-octadecenoic acid, or 10,13-dihydroxystearic acid, individually or a mixture of them.

For example, the processes according to the present invention can use a wide range of organic acids of formula (II) such as acetic, propionic, butyric, valeric, caproic, enanthic, caprylic, pelargonic, capric, undecylic, lauric, tridecyl, myristic, pentadecyl, palmitic, margaric, stearic, nonadecyl, arachidic, henicosanoic, or behenic or a mixture thereof. Preferably, the processes according to the present invention may use the acids of formula (II) such as palmitic, stearic or arachidic acid.

For example, the processes according to the present invention can use a wide range of alcohols of formula (III) such as methanol, ethanol, propanol, isopropanol, butanol, 2-butanol, pentanol, 2-pentanol, 3-pentanol, n hexanol, 2-methyl-pentanol, 3-methylpentanol, 2,2-dimethylbutanol, 2,3-dimethylbutanol, tert-butanol, heptanol, octanol, 2-ethylhexanol, nonanol, decanol, undecanol, dodecanol, tridecanol, tetradecanol , pentadecanol, hexadecanol, heptadecanol, octadecanol, nonadecanol, eicosanol, henicosanol, docosanol (behenyl alcohol), isooctadecanol, 2-butyloctanol, 2-octyldodecanol, 2-hexyldecanol, iso-stearyl alcohol such as 2- (4,4- diméthylpentan-2-yl) -5,7,7-triméthyloctan-l-ol (Fineoxocol ^® 180) or 2- (3-methylhexyl) -7-decan-l-ol (Fineoxocol ^® 180N), iso-palmityl alcohol ( Fineoxocol ^® 1600), 2-octyldecanol, glycerol, polyglycerol, glycol, polyethylene glycol (adjuvant E1521, PEG8, PEG400, PEG600, PEG1000, PEG2000, PEG4000, PEG8000), 1,2-propyleneglycol, polypropylene glycol, 1,3-propylene glycol, 1,4-butanediol, thetitol, tetritol, erythritol, xylitol, ribitol, arabinitol, arabitol, pentitol, pentaerythritol, mannitol, allitol, altritol, hexitol, glucitol sorbitol, dulcitol, volemitol, polyerythritol, polypentaerythritol, polymannitol, or a mixture thereof. Preferably, the processes according to the present invention can use an alcohol of formula (III) such as octanol, polyglycerol, or 2-ethylhexanol, preferably 2-ethylhexanol. It is another object of the present invention to produce estolides having a number of EN estolides selected from 1.1 to 7.0 by incrementing by 0.1 (ie 1.1, 1.2, 1.3, 1, 4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2, 6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5, 1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0,). Advantageously, EN is a value of 2.0 to 2.5, or 2.5 to 3.0, or 3.0 to 3.5, or 3.5 to 4.0, or 4.0 to 4.5 , or 4.5 to 5.0, or 5.0 to 5.5, or 5.5 to 6.0, or 6.0 to 6.5, or 6.5 to 7.0. Advantageously, EN is a value of 2.0 to 5.0 or 2.5 to 4.5.

It is another object of the present invention to produce estolides having at least one or at least two of the following characteristics: an acid number of less than 60, preferably between 10 and 55;

- A melting point between -40 ° C and + 10 ° C;

- A viscosity between 50 and 650 cSt at 40 ° C measured according to ASTM D445;

A viscosity of between 10 and 60 cSt at 100 ° C. measured according to ASTM D445;

A pour point of -25 ° C. to -30 ° C. or -40 ° C. measured according to ASTM D97;

- A pale yellow to amber color.

EXAMPLES General Conditions Ricinoleic acid (purity over 80%) is obtained from TCI Europe (Zwijndrecht, Belgium). 12-hydroxystearic acid (85% purity) is obtained from Alfa Aesar (Karlsruhe, Germany). The 10-hydroxystearic acid is isolated from the reaction of a oleic acid on oleic acid (> 99% purity) obtained from TCI Europe (Zwijndrecht, Belgium). The lipase of the present invention (E1) is prepared by the recombinant expression of the lipase gene of Streptomyces avermitilis in the host cell Escherichia coli. Novozym®435 (E2) lipase from Candida antarctica is immobilized and obtained from Novozymes (Bagsvaerd, Denmark). The IMMCRL-T2-150 lipase derived from Candida rugosa and immobilized (E3) is obtained from Chiral Vision (Leiden, Holland). LE-11 lipase from non-immobilized Candida rugosa (E4) is obtained from ASA Spezialenzyme GmBH (Wolfenbuttel, Germany). The lipase DSM40783 from Streptomyces violaceoruber (E5) is obtained from the Leibniz Institute DSMZ, German Collection of Microorganisms and Cell Cultures (Braunschweig, Germany). The acid number (IA) and the number of estolides (EN) are calculated as previously described.

Example 1 Preparation of Lipase from Streptomyces avermitilis

The lipase gene of Streptomyces avermitilis was cloned into Novagen's pET26-b (+) expression vector (Merck Biosciences Subdivision of Merck KGaA (Darmstadt, Germany)) and this vector was integrated into Escherichia coli host cells. (BL21 (DE3)) from Novagen.

The cells were cultured in the autoinducer medium (Overnight Express Instant TB Medium Express obtained from Merck Chemicals Ltd. Subdivision of Merck KGaA (Darmstadt, Germany)) in the presence of kanamycin (60 μg / ml). The medium was inoculated from a glycerol stock. The culture was incubated for 2 hours at 37 ° C and then for 23 hours at 25 ° C (up to DOeoonm = 11). After centrifugation, the cell pellets were recovered and then lyophilized. In the following examples, the amounts of lipase (El) used are expressed in milligrams of lyophilized cells.

The synthetic gene is obtained from IDT Technologies (Leuven, Belgium) according to the nucleic sequence SEQ ID NO. 1 next:

ATGCTGCCGTGGAAGCGCGTCCTTCGTCCGTTGACGGCATTGTTG CTTACCGTAGCAGTCGCACTTGTTCCCGCCGCCACTGCCCATGCA GACTCTGCGCCCTCTAGTGGGTGGAACGACTATAGCTGCAAACCT TCTGCGGCTCACCCACGTCCCGTAGTCTTGGTGCACGGCACATTA GGTAACTCTGTAGATAATTGGCTTGGGCTGGCTCCTTATCTTGAA CACCGTGGGTATTGTGTTTTCTCATTAGACTACGGGCAACTGTCT GGAGTACCCTTCTTTCATGGTTTAGGCCCAATCGATAAATCCGCA GAGCAGCTTCAAGTGTTTGTGGACAAAGTGTTGACAGCCACGGG TGCTACTAAGGCGGATCTGGTCGGTCATTCCCAGGGTGGGATGAT GCCGCGTTATTACCTGAAATTCCTTGGGGGTGCAGGCAAGGTCA ATGCCCTGGTGGGAATCGCTCCTAATAATCATGGCACCACTTTGT CCGGGCTGACTAACCTGCTTCCTTATTTTCCCGGGGCAGAGGACT TGCTTTCCACTGCAACGCCTGGTCTGGCGGACCAAGTCGTAGGTA GCGCCTTTATGGCGAAGTTAAACGCGGGAGGGGATACTGTAGCA GGGGTTCATTATACAGTCATTGCCACACAGTACGACGAGGTCGTT ACCCCGTATCGTACCGCATTTTTGTCTGGCTCGGATGTACATAAT GTTTTGTTACAAGATTTGTGTCCACTGGACCTTAGCGAGCACGTC GCTATTGGGTTGATCGACCGTATCGCTTTTCATGAGGTCACAAAT GCGTTAGATCCGGCTCACGCGACGCGCACAACTTGTGCCTCCGTG TTCTCCTAA

The protein sequence SEQ ID NO. 2 of lipase from Streptomyces avermitilis is as follows:

MLP WK VLRPLT alllt V AV G VP AAT AH AD SAP SG WND S YS CKP SAAHPRPVVLVHGTLGNSVDNWLGLAPYLEHRGYCVFSLDYGQLS GVPFFHGLGPIDKSAEQLQVFVDKVLTATGATKADLVGHSQGGMM PRY YLKFLGG AGKVN AL VGI APNNHGTTL S GLTNLLP YFPG AEDLL STATPGLADQVVGSAFMAKLNAGGDTVAGVHYTVIATQYDEVVTP YRTAFLSGSDVHNVLLQDLCPLDLSEHVAIGLIDRIAFHEVTNALDP AH ATRTTC AS VF S

Example 2 Comparative Study of Lipases on Ricinoleic Acid

The formation of estolides from ricinoleic acid was carried out in parallel with our lipase derived from Streptomyces avermitilis (E1) prepared according to Example 1, and three commercial enzymes: lipase Novozym®435 (E2), lipase immobilized C. rugosa (E3) and non-immobilized (E4). The reactions were conducted without solvent by mixing 1 g of ricinoleic acid with 90 mg of enzyme or cells containing the unmodified pET26 vector or nothing for the two negative controls, with magnetic stirring at 40 ° C. The reactions were conducted in duplicate. The results presented in Table 1 and FIG 1 are the arithmetic means. Table 1

The reaction with Streptomyces avermitilis lipase produced estolides with a number of estolides two-and-a-half times higher than the reaction conducted with non-immobilized C. rugosa lipase, and nine times higher than the reaction conducted with Novozym®435 lipase which is the most commonly studied commercial enzyme for the formation of estolides. The latter does not seem to have favored the formation of estolides below that obtained by the two negative controls.

Example 3: Study of the Effect of Temperature and Duration on the Formation of Estolides of Ricinoleic Acid

The formation of estolides from ricinoleic acid was conducted in parallel with our Streptomyces avertimilis lipase (E1) and a homologue of Streptomyces violaceoruber (E5). The reactions were conducted without solvent by mixing 1 g of ricinoleic acid with 90 mg of enzyme or cells containing the unmodified pET26 vector or nothing for the two negative controls, with magnetic stirring at 40 ° C, 50 ° C or 75 ° C for 24h, 48h or 96h. The reactions were conducted in duplicate, the results shown in Table 2 and FIGS. 3-5 are the arithmetic means. Table 2

Number

Lipase or white Temperature Duration Index

Estolide (EN) of Acid (IA)

24 h 159 0.2

Without enzyme 40 ° C 48h 160 0,2

96h 159 0.2

24 h 159 0.2

Without enzyme 50 ° C 48h 157 0,2

96h 159 0.2

Without enzyme 75 ° C 24h 157 0.2

24 h 150 0.2 pET26 no

40 ° C 48h 156 0,2 changed

96h 151 0.2

24 h 153 0.2 pET26 no

50 ° C 48h 150 0,2 modified

96h 151 0.2 pET26 no

75 ° C 24h 157 0.2 changed

24 h 47 3.0

El 40 ° C 48h 47 3.0

96h 45 3.2

24 h 47 3.0

El 50 ° C

48h 39 3.8

El 50 ° C 96h * 37 4.0

El 75 ° C 24h 36 4.2

E5 40 ° C 24 hrs 129 0.5 Number

Lipase or white Temperature Duration Index

Estolide (EN) of Acid (IA)

48h 114 0.7

96h 87 1.2

24 h 116 0.6

E5 50 ° C 48h 100 0.9

96h 87 1.2

* without duplicate.

The reaction with Streptomyces avermitilis lipase (E1) produced estolides with a high number of estolides as early as 24 hours of incubation from 40 ° C (see FIGS. 3 to 5). El lipase has demonstrated high efficiency and high stability at high temperatures, especially up to 75 ° C, with an EN above 4 after 24 hours of incubation (see FIG. 5).

Example 4: Ascending Scale for the Formation of Estolides of Ricinoleic Acid

Formation of estolides from ricinoleic acid was conducted with our lipase from Streptomyces avertimilis (E1). The reaction was conducted without solvent by mixing 42 mL of ricinoleic acid with 3.5 g of enzyme, with magnetic stirring at 55 ° C for 24 hours. The titration of a sample of the reaction medium indicated an IA of 51 and therefore an EN of 2.7. The estolide formed is a thick yellow oil containing suspended biomass. Even at -20 ° C, the oil remains fluid. The estolide was taken up in 150 mL of ethyl acetate and then filtered through Clarcel ^® ceca DIC B. The filter was rinsed with 90 mL of ethyl acetate. The solvent was then evaporated under vacuum at 40 ° C. 33.86 g of a viscous amber oil were recovered. The titration of a sample indicated an AI of 58 and therefore an EN of 2.2. The physicochemical properties of viscosity are shown in Table 3 below. Table 3

Example 5 Study of the Esterification of Estolides of Ricinoleic Acid with 2-Ethylhexanol

Esterification of the acid function of ricinoleic acid estolides with 2-ethylhexanol was conducted with our lipase from Streptomyces avermitilis (E1). The reaction was conducted without solvent by mixing 60 mL of ricinoleic acid with 5.74 g of enzyme, with magnetic stirring at 55 ° C, for 24 hours. The titration of a sample of the reaction medium indicated an IA of 55 and therefore an EN of 2.4. 8 ml of 2-ethylhexanol were added to the reaction medium under the same reaction conditions. After 24 hours, the titration of a sample of the reaction medium indicated an AI of 21.

The estolide was taken up in 150 mL of ethyl acetate and then filtered through Clarcel ^® ceca DIC B. The filter is rinsed with 90 mL of ethyl acetate. The solvent and excess 2-ethylhexanol were then evaporated under vacuum at 40 ° C to constant mass. A viscous oil was recovered. The titration of a sample indicated an AI of 21. The rheological properties are shown in Table 4 below.

Table 4

Protection of the acid function of ricinoleic acid estolides with 2-ethylhexanol induces a decrease in viscosity at 40 ° C associated with a slight decrease in viscosity at 100 ° C and a sharp increase in the viscosity index. Example 6: Study of Estolide Formation from 12-Hydroxystearic Acid

Formation of estolides from 12-hydroxystearic acid was conducted with our lipase of S. avertimilis (E1). The reactions were conducted without solvent by mixing 1 g of 12-hydroxystearic acid with 90 mg of enzyme or cells containing the unmodified pET26 vector or nothing for the two negative controls, with magnetic stirring at 75 ° C for 24 hours. The reactions were conducted in duplicate, the results presented in Table 5 are the arithmetic means.

Table 5

The reaction with Streptomyces avermitilis lipase (E1) produced estolides with a high number of estolides with 12-hydroxystearic acid as early as 24 hours.

Example 7 Comparative Study of Lipases on 12-Hydroxystearic Acid

The formation of estolides from 12-hydroxystearic acid was conducted in parallel with our S. avermitilis lipase and three commercial enzymes: Novozym®435 (E2), immobilized C. rugosa lipase (E3) and not immobilized (E4). The reactions were conducted without solvent by mixing 1 g of 12-hydroxystearic acid with 90 mg of enzyme or cells containing the unmodified pET26 vector or nothing for the two negative controls, with magnetic stirring at 76 ° C for 24 hours. The reactions were conducted in duplicate, the results presented in Table 6 are the arithmetic means. Table 6

The reaction with Streptomyces avermitilis lipase produced estolides with an estolide number eight to ten times higher than the reactions with immobilized and non-immobilized C. rugosa lipases and Novozym®435 which is the the most commonly studied commercial enzyme for the formation of estolides. These lipases do not seem to have favored the formation of estolides since their number of estolides remained of the same order of magnitude as those obtained with the two negative controls.

Example 8: Study of Estolide Formation from 12-Hydroxystearic Acid, Variation in Temperature

Formation of estolides from 12-hydroxystearic acid was conducted with our lipase from Streptomyces avermitilis (E1). The reactions were carried out without solvent by mixing 1 g of 12-hydroxystearic acid with 90 mg of enzyme with magnetic stirring at 81 ° C. for 5 h and then at 75 ° C. for 48 h. The reactions were conducted in duplicate, the results presented in Table 7 are the arithmetic means.

Table 7

* in duplicate The reaction with Streptomyces avermitilis lipase (E1) produced estolides with a high number of estolides as early as 24 hours of incubation. This number of estolides reached 3.4 after 48 hours of incubation. El lipase has demonstrated high stability at high temperatures, especially up to 81 ° C for at least 5 hours. Example 9 Immobilization of Streptomyces Avermitilis Lipase on Solid Support

27 g of Diaion ^® HP20 resin (available from Resindion Sri (Binasco, Italy)) was suspended in 100 mL of deionized water. The water is removed and the resin is resuspended in 100 mL of a cell-breaking supernatant containing Streptomyces avermitilis lipase. The resin is kept in suspension with stirring for 96 hours at 25 ° C. The supernatant is then evacuated. The resin is then dried under vacuum at 50 ° C.

BIBLIOGRAPHIC REFERENCES

US2,785,978

US4,582,804

US5,380,894

US5,852,196

US5,900,496

US7,125,694

US8,729,176

US2010 / 0204435

US2013 / 0102041

WO2008 / 031908

WO2008 / 040864

WO2011 / 037778

WO2013 / 009471

WO2016 / 151115

FR2906530

FR2942628

A. Bodalo et al., Biochem. Eng. J, vol. 44, 214-219 (2009)

J. Greco-Duarte et al, Fuel Vol 202, 196-205 (2017) DG Hayes et al. JAOCS, vol. 72, No. 11, 1309-1316 (1995)

H. Horchani et al. J. Mol. Cat. B Enz., Vol. 75, No. 1 1, 35-42 (2012)

I. Martin-Arjol, J. Am. OR Chem. Soc., Vol. 92, no. 8, 1125-1141 (2015) A. Todes, Pure Appl. Chem., 87 (1), 51-58 (2015)

B. Wang, Acta Microbiol. Sinica, 50 (2), 236-243 (2010)

S. Ortega-Requena et al., Biochem. Eng. J, vol. 84, 91-97 (2014) Needleman and Wunsch, J. Mol. Biol. 48,443-453 (1970)

Claims

A process for the formation of estolides from a hydroxylated fatty acid characterized in that it is conducted in the presence of a lipase having the amino acid sequence SEQ ID NO. 2 next:

MLP WKRVLRPLT ALLLT V AV AL VP AAT AH AD S APS S G WND YS CKP S AAHP

RPVVLVHGTLGNSVDNWLGLAPYLEHRGYCVFSLDYGQLSGVPFFHGLGP

IDKSAEQLQVFVDKVLTATGATKADLVGHSQGGMMPRYYLKFLGGAGKV

NALVGIAPNNHGTTLSGLTNLLPYFPGAEDLLSTATPGLADQVVGSAFMAK

LNAGGDTVAGVHYTVIATQYDEVVTPYRTAFLSGSDVHNVLLQDLCPLDL

SEHVAIGLIDRIAFHEVTNALDPAHATRTTCASVFS,

or a sequence having at least 85% identity with the sequence SEQ ID NO. 2.

2. Method according to claim 1, characterized in that the lipase has the amino acid sequence SEQ ID NO. 2 or a sequence having at least 90%, in particular at least 95% identity with the sequence SEQ ID NO. 2.

3. Method according to claim 1 or 2, characterized in that, after optimal global alignment with SEQ ID NO: 2, the amino acids of the lipase sequence aligned with positions 43, 80, 128-132, 221, 246 , 253, and 281 of SEQ ID NO: 2 are identical to the amino acids of SEQ ID NO: 2 at positions 43 (C), 80 (C), 128-132 (GHSQG pentapeptide, SEQ ID NO: 3), 221 ( D), 246 (C), 253 (H) and 281 (C) respectively.

4. Method according to any one of claims 1 to 3, characterized in that the lipase comprises a sequence of 256 amino acids having at least 90%, especially at least 95%>, and preferably 100% identity, with the positions 31-286 of SEQ ID NO: 2.

5. Method according to any one of claims 1 to 4, characterized in that the product estolide has a number of estolides greater than or equal to 1.1.

6. Method according to claim 5, characterized in that the product estolide has a number of estolide greater than or equal to 2.0.

7. Method according to claim 6, characterized in that the product estolide has a number of estolide greater than or equal to 3.0.

8. Method according to any one of claims 1 to 7, characterized in that it is conducted without solvent.

9. Process according to any one of claims 1 to 8, characterized in that the hydroxylated fatty acid is selected from the group consisting of 10-hydroxystearic acid, 12-hydroxystearic acid and 13-hydroxystearic acid. ricinoleic acid, 10-hydroxy-12-octadecenoic acid, 13-hydroxy-9-octadecenoic acid, 10,13-dihydroxystearic acid, and mixtures thereof.

10. Process according to any one of claims 1 to 9, characterized in that the reaction is initiated at a temperature above the melting point of the hydroxylated fatty acid.

11. Process according to any one of claims 1 to 10, characterized in that an alcohol is added to the reaction medium once the acid number of the reaction medium is below 60.

12. Process according to claim 11, characterized in that the alcohol is chosen from the group consisting of methanol, ethanol, propanol, isopropanol, butanol, 2-butanol, pentanol, 2-pentanol, 3-pentanol and n-hexanol. , 2-methyl-pentanol, 3-methyl-pentanol, 2,2-dimethylbutanol, 2,3-dimethylbutanol, tert-butanol, heptanol, octanol, 2-ethylhexanol, nonanol, decanol, undecanol, dodecanol, tridecanol, tetradecanol, pentadecanol , hexadecanol, heptadecanol, octadecanol, nonadecanol, eicosanol, henicosanol, docosanol, iso-octadecanol, 2-butyloctanol, 2-octyldodecanol, 2-hexyldecanol, iso-stearyl alcohol such as 2- (4,4-dimethylpentan-2-yl) 5,7,7-trimethyloctan-1-ol or 2- (3-methylhexyl) -7-decan-1-ol, iso-palmitic alcohol, 2-octyldecanol, glycerol, polyglycerol, glycol, polyethylene glycol, 1,2- propylene glycol, polypropylene glycol, 1,3-propylene glycol, 1,4-butanediol, threitol, tetritol, erythritol, xylitol, ribitol, arabinitol, arabitol, pen titol, pentaerythritol, mannitol, allitol, altritol, hexitol, glucitol, sorbitol, dulcitol, volemitol, polyerythritol, polypentaerythritol, polymannitol, and mixtures thereof.

13. Process according to claim 11, characterized in that the alcohol is 2-ethylhexanol.

14. Process according to any one of claims 1 to 10, characterized in that an organic acid is added to the reaction medium once the acid number of the reaction medium is below 60.

15. Process according to any one of claims 1 1 to 13, characterized in that an acid is added to the reaction medium once the acid number of the reaction medium is between 10 and 40.

16. The method of claim 14 or 15, characterized in that the acid is selected from the group consisting of acetic acid, propionic, butyric, valeric, caproic, enanthic, caprylic, pelargonic, capric, undecylic, lauric, tridecylic , myristic, pentadecyl, palmitic, margaric, stearic, nonadecyl, arachidic, henicosanoic, behenic and mixtures thereof.

17. Process according to any one of claims 1 to 16, characterized in that the estolide formed has a number of estolides EN at a value of 1.5 to 2.0, or 2.0 to 2.5, or 2.5 to 3.0, or 3.0 to 3.5, or 3.5 to 4.0, or 4.0 to 4.5, or 4.5 to 5.0.

18. Process according to any one of Claims 1 to 4, characterized in that the estolide is formed according to the following scheme (1)

HO-CHRi-Q-COOH + Lipase → H- (O-CHRi-Q-CO) _n -OH (1)

I II

Where R 1 is selected from saturated or mono-unsaturated, linear or branched hydrocarbon groups of one to twelve carbon atoms (C 1 -C 12) optionally substituted by a hydroxyl group, Q is a divalent non-cyclic aliphatic saturated or mono unsaturated group from six to twelve carbon atoms (C6 to Cl2) optionally substituted by a hydroxyl group, and n is a value greater than or equal to 2.1, preferably greater than or equal to 2.5.

19. Process according to any one of Claims 1 to 4, characterized in that the estolide is formed according to the following scheme (2)

HO-CHRi-Q-COOH + HOR ₃ + Lipase → H- (O-CHRi-Q-CO) _n- OR ₃ (2)

I IV V

Where R 1 is selected from saturated or mono-unsaturated hydrocarbon groups, linear or branched from one to twelve carbon atoms (C 1 -C 12) optionally substituted by a hydroxyl group, R ₃ is selected from linear or branched alkyl groups of one to twenty-two carbon atoms (C1 to C22) optionally substituted with one or more hydroxyl groups, and the radicals of polyether alcohols, the radicals of polyether alcohols being chosen so that the molecule HOR3 represents a polyether alcohol, Q is a divalent saturated or monounsaturated non-cyclic aliphatic group of six to twelve carbon atoms (C6 to Cl 2), optionally substituted by a hydroxyl group, and n is a value greater than or equal to 2.1, preferably greater than or equal to 2.5.

20. Process according to any one of claims 1 to 4, characterized in that the estolide is formed according to the following scheme (3):

n HO-CHR-Q-COOH + R ₂ -COOH + HOR ₃ Lipase + → R ₂ -CO- (CHR-0-Q-CO) _n -OR ₃ (3)

Wherein R 1 is selected from saturated or mono-unsaturated hydrocarbon groups, linear or branched from one to twelve carbon atoms (Cl to Cl 2) optionally substituted with a hydroxyl group, R ₂ is selected from saturated hydrocarbon groups or unsaturated, linear or branched from one to twenty-two carbon atoms (Cl to C22), R3 is selected from linear or branched alkyl groups of one to twenty-two carbon atoms (C1 to C22) optionally substituted with one or more hydroxyl groups, and the radicals of polyether alcohols, the radicals of polyether alcohols being chosen so that the molecule HOR3 represents a polyether alcohol, Q is a divalent non-cyclic aliphatic saturated or monounsaturated group of six to twelve carbon atoms (C6 to C1 2), optionally substituted by a hydroxyl group, and n is a value greater than or equal to 2.1, preferably greater than or equal to 2.5.

21. Process according to claim 20, characterized in that R ₂ is selected from - CH 3, -C ₂ H ₅ , -C ₃ H ₇ , -C ₄ H 9, -C ₅ H 11, -C ₆ H 3, -C ₇ H 15, -C ₈ H 17, -C ₉ H 19, -CioH ₂ , -CnH ₂ 3, -C12H25, -C13H27, -C14H29, -C15H31, -C16H33, -C17H35, -C18H37, -C19H39, -C20H41, - C21H43, -C22H45, -C _r H _2r CH (CsH _{2s +} i) Cth ₂ t ₊ i, -C _u H _2u -CH = CH ₂ and -C _m H _2m - (CH = CH-

CH ₂ ) pC _q H _{2q +} i where r is an integer from 0 to 19, s and t are integers from 1 to 20, the sum r + s + t varies from 2 to 21, m and q are integers of 0 at 19, and p is an integer of 1 to 3.

22. A method according to any one of claims 19 to 21, characterized in that R3 is selected from a linear alkyl of the formula -CH 3, -C ₂ H _5, -C3H7, -C4H9, - C5H11, -C ₆ HI3, -C7H15, -C8H17, -C9H19, -CioH ₂ i, ₂ -CnH 3, -C ₂ H ₂₅ ₂ -Ci3H 7 -Ci4H ₂ 9 -

C 15 H 31, -C 16 H 33, -C ₁₇ H 35, -C 18 H 37, -C ₁₉ H 39, -C ₂₀ H 41, -C ₂ H 43, or -C ₂₂ H 45; branched alkyl of formula -C _r H _2r -CH (CsH _2s + i) Cth ₂ t + i where r is an integer from 0 to 19, s and t are integers from 1 to 20 and the sum r + s + t varies from 2 to 22; said linear or branched alkyl being optionally substituted with 1 to 7 hydroxyls; and a polyglycerol radical, polyerythritol, polypentaerythritol, polymannitol, a polyalkylene glycol or a copolymer of alkylene glycols.

23. Process according to any one of claims 18 to 22, characterized in that R 1 is selected from -CH ₃ , -C ₂ H ₅ , -C ₃ H ₇ , -C ₄ H ₉ , -C ₅ H 11, -C ₆ H ₃ , -C ₇ H ₅ , -C ₈ H ₁₇ , -C ₉ H ₁₉ , -C ₁₀ H ₂₁ , -C ₁₁ H ₂₃ , -C ₁₂ H 25, -CH = CH ₂ , -CH = CH-CH ₃ , -CH = CH-C ₂ H ₅ , -CH = CH- C ₃ H ₇ ,

-

-CH = CH-CioH ₂ 1, -CH ₂ -CH = CH-CH ₃ , -CH ₂ -CH = CH-C ₂ H ₅ , -CH ₂ -CH = CH-C ₃ H ₇ , -CH ₂ - CH = CH-C ₄ H ₉ , -CH ₂ -CH = CH-C ₅ Hn, -

CH ₂ -CH = CH-C ₆ H ₃ , -CH ₂ -CH = CH-C ₇ Hi ₅ ,

-CH ₂ -CH = CH- C9H19, -C ₂ H ₄ -CH = CH-CH _3, -C2H ₄ -CH = CH-C ₂ H _5, -C ₂ H ₄ -CH = CH-C ₃ H ₇ , -C2H4- CH = CH-C ₄ H _9, -C2H ₄ -CH = CH-C ₅ H n, -C ₂ H ₄ -CH = CH-C ₆ Hi _3, -C ₂ H ₄ -CH = CH- C ₇ Hi ₅ ,

-CH ₂ -CH (OH) -CH ₃ , -CH ₂ -CH (OH) -C ₂ H ₅ , -CH ₂ -CH (OH) -C ₃ H ₇ , -CH ₂ -CH (OH) -C ₄ H ₉ , -CH ₂ -CH (OH) -C ₅ Hn, -CH ₂ -CH (OH) -C ₆ H ₃ , -CH ₂ -

CH (OH) -C ₇ Hi _5, -CH ₂ -CH (OH) -C ₈ Hi _7, -CH ₂ -CH (OH) -C ₉ Hi9, -CH ₂ -CH (OH) - C10H21, -C ₂ H ₄ -CH (OH) -CH ₃ , -C ₂ H ₄ -CH (OH) -C ₂ H ₅ , -C ₂ H ₄ -CH (OH) -C ₃ H ₇ , -C ₂ H ₄ -CH (OH) - C ₄ H _9, -C2H ₄ -CH (OH) -C ₅ Hn, -C ₂ H ₄ -CH (OH) -C ₆ Hi _3, -C ₂ H ₄ -CH (OH) - C ₇ Hi _5, -C ₂ H ₄ -CH (OH) -C ₈ Hi ₇ and -C2H ₄ -CH (OH) -C ₉ Hi9.

24. Process according to any one of claims 18 to 23, characterized in that Q is selected from -C ₆ Hi 2 -, -C ₇ H ₁₄ -, -C ₈ H ₆ -, -CH ₁₈ -, -C ₁₀ H ₂₀ - , -C ₁₁ H ₂₂ -, -C ₁₂ H 24 -, -CH = CH-C ₄ H ₈ -, -CH = CH-C ₅ H 10-, -CH = CH-C ₆ H ₂ -, -CH = CH-C ₇ Hi ₄ -, -CH = CH- C ₈ Hie-, -CH = CH-C ₉ Hi ₈ -, -CH = CH-CioH ₂₀ -, -CH ₂ -CH = CH-C ₃ H ₆ -, -CH ₂ - CH = CH- -C ₄ H ₈ -, -CH ₂ -CH = CH-C ₅ Hio-, -CH ₂ -CH = CH-C ₆ Hi ₂ -, -CH ₂ -CH = CH-C ₇ Hi ₄ - , -CH ₂ -

-CH ₂ -CH = CH-C ₉ Hi ₈ -, -C2H4-CH = CH-C ₂ H ₄ -, -C ₂ H ₄ -CH = CH-

C ₃ H ₆ -, -C ₂ H ₄ -CH = CH-C ₄ H ₈ -, -C2H ₄ -CH = CH-C ₅ Hio-, -C ₂ H 4 CH = CH-C ₆ Hi2- - C ₂ H ₄ -CH = CH-C ₇ Hi ₄ -, -C ₂ H ₄ -CH = CH-C ₈ Hi ₆ -, -CH ₂ -CH (OH) -C ₄ H ₈ -, -CH ₂ -CH ( OH) -C ₅ H 10-, -CH ₂ -CH (OH) -C ₆ Hi _{2 -} , -CH ₂ -CH (OH) -C ₇ H ₄ -, -CH ₂ -CH (OH) -C ₆ H-, - CH ₂ -CH (OH) -C ₉ H ₈ -, -CH ₂ -CH (OH) -CioH ₂ o-, -C ₂ H ₄ -CH (OH) -C ₃ H ₆ -, - C ₂ H ₄ -CH (OH) -C ₄ H ₈ -, -C ₂ H 4 CH (OH) -C Hio- _5, -C ₂ H 4 CH (OH) -C ₆ Hi2-, -C2H4-

CH (OH) -C Hi4- _7, -C ₂ H ₄ -CH (OH) -C ₈ Hi6- and -C2H ₄ -CH (OH) -C ₉ Hi ₈ -. Process according to any one of claims 18 to 24, characterized in that a selected value of 2.5 to 3.0, or 3.0 to 3.5, or 3.5 to 4.0, or 4, 0 to 4.5, or 4, at 5.0, or 5.0 to 5.5, or 5.5 to 6.0.