ITMI20121681A1 - METHOD OF PRODUCTION OF ETHYLIC ESTERS OF ACID FAT POLYINSATURES AND THEIR USE IN FORMULATIONS FOR ORAL USE IN MAN - Google Patents

Description

"METHOD OF PRODUCTION OF ETHYL ESTERS OF POLYUNSATURATED FATTY ACIDS AND THEIR USE IN FORMULATIONS FOR ORAL USE IN MAN"

The present invention relates to a new chemical method for preparing ethyl esters of polyunsaturated fatty acids, derived from semi-finished precursors, their purification and their use in the preparation of formulations for oral use.

State of the art

Polyunsaturated fatty acids, in particular linear ones, bearing carbon-carbon double bonds in a "cis" (or "Z") configuration and separated from each other by a saturated carbon atom, having eighteen or more total carbon atoms , have long been known as important or essential nutritional factors in humans, entering the constitution of membrane phospholipids and the biosynthesis of prostaglandins.

More specifically, those called "omega-3", characterized by the fact of having, in the IUPAC name, the last double bond between the third and fourth carbon atom starting from the terminal methyl of the chain, have been recognized as important in the prevention and treatment of dyslipidemias and the damage resulting from them on the cardiovascular system.

Some semi-finished products deriving from oil extracted from marine fish, especially of the genus Sardinia and Gadus (cod) are particularly rich in "omega-3" fatty acids. This oil consists of triglycerides in which the main fatty acid residues are the acid (a // - Z) -5,8, ll, 14,17-eicosapentaenoic, or EPA, and the acid (a // - Z) -4,7,10,13,16,19-docosahexaenoic, or DHA, of formula (I) and (II), respectively.

FORMULA (I) FORMULA (II)

(all-Z) -4,7,10,13,16,19-docosahexaenoic acid (DHA) (all-Z) -5,8,11,14,17-eicosapentaenoic acid

Molecular Weight = 328.50

Molecular Weight = 302.46 Molecular Formula = C22H3202 Molecular Formula = C20H3002

EPA is important in the biosynthesis of series 3 prostaglandins and thromboxanes, while DHA is the major constituent of the sensitive membranes of the retina and brain.

Fish oil is obtained from fish, or their liver, by mechanical or extractive processes.

EPA and DHA, together with other omega-3 acids and other minor components are isolated from fish oil as acids, salts, esters or other functional derivatives, by chemical or enzymatic processes. In particular, commercial "EE fish oil" means a semi-finished product deriving from oil, extracted from fish of various kinds, through transesterification processes and consisting mainly of the ethyl esters of the original fatty acids. Various specialties containing mixtures of EPA and DHA and their derivatives in various capacities are on the market as food supplements.

Especially in Italy it is particularly sought after, because it enters the composition of pharmaceutical specialties, a mixture of ethyl esters of EPA and DHA at a very high level: EPA plus DHA greater than 85% of the total, with EPA not less than 40% and DHA not less than 34%; Total Omega-3s greater than 95%; EPA / D HA ratio between 0.9 and 1.5.

A mixture of this quality has been obtained, up to now, from natural fish oil, by preliminary refining (filtration, bleaching, deodorization, neutralization, washing, drying), transesterification to obtain the mixture of raw ethyl esters, treatment with urea to separate the derivatives of saturated and monounsaturated fatty acids, distillation to obtain the desired product. Distillation must be performed with the technique, and therefore the equipment, of "molecular" or short-path distillations (US201 1091947A1 - AK Biotech). The overall yield is around 25% by weight of fish oil and the molecular distillation equipment is very expensive.

There is, however, a chemical method of preparation of the ethyl esters of Omega-3 fatty acids, reported in WO20 12/088620 A2 with the title "Omega-3 concentrated". This method claims a process which, starting from oils of various natural origin, animal or vegetable, by means of alkaline hydrolysis followed by esterification with acid catalysis, obtains a mixture of ethyl esters with a total omega-3 title greater than 80%. However, a purification by molecular distillation of the crude ethyl esters is required. The oil used is of low quality, with total Omega-3 from 30 to 50%, the hydrolysis is done at high temperature, the precipitated sodium salts are discarded, the waters containing glycerin are sent as they are to esterification. large quantities of acids and intermediate fatty acids are not isolated which, in this way, are neither purified nor washed. The final yield is low: about 20-25% by weight of the oil and Omega-3 recovery of 50-60% of the original content.

Description of the invention

It has now been found that a mixture of ethyl esters of EPA and DHA of the "pharmaceutical" quality required in Italy and described above can be obtained by means of a completely chemical process, simple and executable with normal reactors and equipment in an industrial chemical plant.

In short, it involves subjecting the EE fish oil of the market, of the best quality available, in a suitable solvent, to alkaline hydrolysis by means of a defined quantity of alkali, at a controlled temperature and in the presence of antioxidants. A precipitate is obtained consisting of the alkaline salts of the fatty acids constituting the esters of the oil itself. This precipitate can be separated from the mother liquors by filtration or kept together with them in the subsequent phases; the choice is a consequence of the quality of the oil used and the EPA / DHA ratio in it, because the precipitate is richer in EPA than the oil itself, while the mother liquors are enriched in DHA. From the alkaline salts, by acidification at controlled pH and temperature, in an aqueous environment, the corresponding acids are obtained which, after extraction and washing, are transformed into the desired esters by acid catalysis in the corresponding alcohol. The mixture of esters, suitably concentrated, extracted and washed, is decoloured with activated carbon and, if necessary, purified with urea. The yield is between 52 and 82% by weight of the starting oil, depending on the need for purification.

All processing takes place in an inert atmosphere and in the presence of antioxidants and radical inhibitors.

The differences, and the consequent improvements, with respect to the previous technique, reported in WO 2012/088620 A2, are the following:

- The starting oil is commercial "EE fish oil", as already defined, of high quality, type "45:35", ie having an EPA content higher than 45% and a higher DHA content 35%. The alkaline hydrolysis procedure, preferably carried out in isopropanol, leads to obtaining the sodium salts of Omega-3 fatty acids as solid phase, separable from mother liquors with excellent yields and free from unwanted impurities and by-products.

- The separation of the sodium salts of the Omega-3 fatty acids allows to correct any imbalances in the EPA / DHA ratio which must be between 0.9 and 1.5; this is because it has been found that the precipitate is naturally enriched in EPA, while the filtrate is found enriched in DHA.

- The presence of antioxidants from the early stages of processing greatly slows down the degradation of Omega-3 derivatives.

- The isolation, by solvent extraction, of the free Omega-3 fatty acids and their purification by appropriate washing, is a further advantage for the quality of the finished product.

- The esterification of Omega-3 fatty acids takes place with acid catalysis, in absolute ethanol and in very mild conditions; the acids used are truly in a catalytic quantity and we have preferably chosen the one which, being reported in the analytical procedure of the European Pharmacopoeia, is probably the most suitable for giving quantitative results without damaging the product.

- The mixture of Omega-3 ethyl esters obtained is neutralized under conditions such as to avoid new hydrolysis, for example with sodium ethoxide, extracted with a solvent and washed with water.

- It is further purified by treatment with decolouring charcoal selected from those used for food or pharmaceutical use.

- When the purification treatment through the formation of adducts with urea is useful, this is used in mild conditions and in an approximately equal quantity, by weight, with respect to the esters to be treated, thus avoiding high volumes and the formation of unwanted by-products.

- No distillation of the finished product is ever necessary, much less "molecular".

- The final yield ranges from 50 to 80% by weight of the starting oil.

- 60 to 95% of the total Omega-3 content of the starting oil is recovered.

The mixture of Omega-3 ethyl esters, which is liquid, can be packaged in soft gelatin capsules, like various specialties already on the market; adsorbed on inert support, such as, for example, precipitated silica, diatomaceous earth or the like, with suitable solid excipients, binders, disaggregating and lubricating agents, as well as antioxidants, it can also be packaged in tablets or sealed capsules.

Detailed description of the invention

The process of the invention is carried out in an inert atmosphere: a quantity of commercial EE fish oil, of minimum quality 45:35, preferably distilled, is dissolved from three to four volumes of 96% ethanol or absolute ethanol or isopropanol, preferably isopropanol. 1% by weight of antioxidant selected from tocopherol, sodium or potassium metabisulphite, BHA, BHT, ascorbic acid or others admitted for pharmaceutical use, preferably BHT or BHA, is added. At a temperature between 0 ° and 60 ° C, preferably from 15 ° to 20 ° C, a saturated solution of sodium or potassium hydroxide is added, in an amount from 1 to 2 equivalents / equivalent, preferably 1.1 eq / eq., In one of the alcohols mentioned above, preferably absolute ethanol. It is then stirred at the same temperature for a time ranging from 1 to 12 hours. The precipitated solid can be filtered by washing it with absolute ethanol or isopropanol, or the entire reaction mixture can be brought to residue under vacuum; in the first case, the mother liquors and the washes are discarded. The filtered solid or the residue of the concentration are dissolved in four or five volumes of water; the solution is acidified with a non-oxidizing mineral acid, such as hydrochloric or sulfuric acid at various concentrations, and repeatedly extracted with a water-immiscible solvent, such as toluene, petroleum ether or liquid aliphatic or cycloaliphatic hydrocarbons, ethyl acetate, ethyl ether or isopropyl, dichloromethane or chloroform. The combined extracts are washed with water and brought to residue under vacuum, obtaining a mixture of the acids corresponding to the acyl radicals of the triglycerides used. Three to four volumes of absolute ethanol are loaded onto said mixture. An acid catalyst is added, in an amount from 0.01 to 0.2 mol / mol, preferably 0.1 mol / mol, selected from non-oxidizing and anhydrous mineral acids, such as hydrochloric acid gas, sulfuric acid, phosphoric acid, or acid 4-toluenesulfonic, benzenesulfonic, methanesulfonic, or Lewis acids such as aluminum trichloride, iron trichloride, tin tetrachloride, titanium tetrachloride, boron trifluoride, also as a complex with ethyl ether, boron trichloride, zinc dichloride or others; preferably boron trichloride. The mixture is stirred at a temperature from 0 ° to 60 ° C for a time from 1 to 12 hours. The reaction environment is neutralized with mineral or organic alkali, preferably sodium ethoxide, and the residue is brought to a vacuum. It is taken up with a solvent selected from those, immiscible with water, already listed in the previous step and washed repeatedly with water. The organic phase is treated with decolorizing carbon, chosen from among those for pharmaceutical or food use, in quantities from 1/10 to 1/20 on the weight of the oil, filtered, washing the filter with the same solvent, and brought to residue under vacuum. If necessary, the residue is washed again with deionized water, at a temperature from 20 ° to 60 ° C, two or three times; it then dries well under vacuum.

If analytically necessary, the residue is again taken up with methanol, from three to four volumes, and urea, from 0.9 to 1.1 times by weight, homogenized under heat and left to crystallize by slowly cooling down to a temperature between 0 ° and 15 ° C. The crystallized solid is filtered, washing it with a little methanol, and the filtrate is brought to residue under vacuum. The residue is repeatedly washed with deionized water, at a temperature from 20 ° to 60 ° C and dried carefully under vacuum. The ester yield ranges from about 50 to 80% by weight of the oil used.

The mixture of Omega-3 ethyl esters prepared with the process of the invention can be diluted in a volatile solvent, such as petroleum ether, dichloromethane or ethanol and the solution is mixed with precipitated silica or diatomaceous earth in such quantities as to obtain a paste. consistent. This pasta is loaded onto the arelle of an oven that can be heated and vacuum-packed. The whole is dried, with a vacuum below 50 mbar and a temperature not exceeding 60 ° C, until the residual solvent in the solid mass is below the permitted limits. The residual solid mass is ground in a special granulator, added with excipients that allow the formation of tablets, such as lactose, microcrystalline cellulose, polyvinylpyrrolidone or PVP, talc and magnesium stearate and compressed with suitable punches. Or, with the addition of dry lubricants only, such as talc and magnesium stearate, the mass is loaded into a special capsule filling machine to fill hard gelatin capsules.

Here are some examples of preparation, both of Omega-3 ethyl esters, and of tablets and capsules object of the invention.

EXAMPLE 1

Fish oil EE “45/35” (minimum 45% EPA and 35% DHA) 100 g, dissolved in absolute ethanol 300 ml. Under stirring, 1.0 g BHT and a 98% 12.8 g sodium hydroxide solution in 150 ml absolute ethanol are added; the mixture is then stirred at room temperature for 12 hours. It is cooled to 0 ° C and the precipitated solid is filtered, washing it with a little absolute ethanol cooled to 0 ° C. The solid obtained is dissolved in 500 ml of water, 200 ml dichloromethane is added and the mixture is brought to pH ~ 2 with 37% hydrochloric acid, stirring well. It is stirred for a few minutes and the phases are separated; the aqueous phase is extracted again with 100 ml dichloromethane. The combined extracts are washed with 100 ml deionized water and brought to residue under vacuum; an oily residue of free Omega-3 fatty acids is obtained, 74.5 g. These fatty acids are dissolved in absolute ethanol 400 ml, a 1M solution in heptane of boron trichloride, 23.7 ml, is added, stirred briefly and left at room temperature, in the dark, for 12 hours. It is then brought to reflux for 1 hour and cooled to 0 ° C. The pH is brought to neutral with a 21% solution of sodium ethoxide in ethanol and all the solvent is distilled under vacuum to a residue in a 60 ° C bath. The residue is taken up with petroleum ether, fraction 30 ° - 50 ° C, 400 ml and 100 ml deionized water. It is stirred well and the phases are separated, discarding the lower aqueous phase. The organic phase is washed again with 100 ml deionized water two more times. The organic phase washed with "Eno Anticromo" decolouring carbon 8 g, for 40 - 45 'is treated and the carbon is filtered, washing the filter with petroleum ether 30 - 50. The filtrate is brought to residue under vacuum; this residue is washed twice with deionized water heated to 55 ° C, 200 ml each time and dried well under vacuum at a maximum temperature of 70 ° C. The oily residue, consisting of the ethyl esters of Omega-3 fatty acids, weighs 73 g. The EPA content is 48.2%; that in DHA, 34.3%; that in total Omega-3, 90%.

EXAMPLE 2

EE “45/35 distilled” fish oil (minimum 45% EPA and 35% DHA), 100 g, dissolved in 96% ethanol 300 ml. Under stirring, 1.0 g BHT and a 98% 12.8 g sodium hydroxide solution in 96% ethanol 150 ml are added; the mixture is then stirred at room temperature for 12 hours. All the solvent is distilled under vacuum. The residue is dissolved in 500 ml deionized water, obtaining a clear solution, 200 ml dichloromethane is added and the mixture is brought to pH ~ 2 with 37% hydrochloric acid, stirring well. It is stirred for a few minutes and the phases are separated; the aqueous phase is extracted again with 100 ml dichloromethane. The combined extracts are washed with 100 ml deionized water and brought to residue under vacuum; an oily residue of free Omega-3 fatty acids is obtained, 91.0 g. These fatty acids are dissolved in absolute ethanol 400 ml, a 1M solution in heptane of boron trichloride, 23.7 ml, is added, stirred briefly and left at room temperature, in the dark, for 12 hours. It is then brought to reflux for 1 hour and cooled to 0 ° C. The pH is brought to neutral with a 21% solution of sodium ethoxide in ethanol and all the solvent is distilled under vacuum to a residue in a 60 ° C bath. The residue is taken up with petroleum ether, fraction 30 ° - 50 ° C, 400 ml and 100 ml deionized water. It is stirred well and the phases are separated, discarding the lower aqueous phase. The organic phase is washed again with 100 ml deionized water two more times. The organic phase is treated, washed with "Eno Anticromo" bleaching carbon 9 g, for 40 - 45 'and the carbon is filtered, washing the filter with petroleum ether 30 - 50. The filtrate is brought to residue under vacuum; this residue is washed twice with deionized water heated to 55 ° C, 200 ml each time and dried well under vacuum at a maximum temperature of 70 ° C. The oily residue, consisting of the ethyl esters of Omega-3 fatty acids, weighs 82 g. The EPA content is 47.3%; that in DHA, 45.5%; that in total Omega-3, 96%.

EXAMPLE 3

Fish oil EE “45/35” (minimum 45% EPA and 35% DHA) 100 g, dissolved in isopropanol 300 ml. Under stirring, 1.0 g BHA and a 98% 12.8 g sodium hydroxide solution in 150 ml absolute ethanol are added; the mixture is then stirred at room temperature for 12 hours. It is cooled to 0 ° C and the precipitated solid is filtered, washing it with isopropanol. The solid obtained is dissolved in 500 ml water, 200 ml ethyl acetate is added and the mixture is brought to pH ~ 2 with 37% hydrochloric acid, stirring well. It is stirred for a few minutes and the phases are separated; the aqueous phase is extracted again with 100 ml ethyl acetate. The combined extracts are washed with 100 ml deionized water and brought to residue under vacuum; an oily residue of free Omega-3 fatty acids is obtained, 80.5 g. These fatty acids are dissolved in absolute ethanol 400 ml, a 1M solution in heptane of boron trichloride, 23.7 ml, is added, stirred briefly and left at room temperature, in the dark, for 12 hours. It is then brought to reflux for 1 hour and cooled to 0 ° C. The pH is brought to neutral with a 21% solution of sodium ethoxide in ethanol and all the solvent is distilled under vacuum to a residue in a 60 ° C bath. The residue is taken up with petroleum ether, fraction 30 ° - 50 ° C, 400 ml and 100 ml deionized water. It is stirred well and the phases are separated, discarding the lower aqueous phase. The organic phase is washed again with deionized water 100 ml for two more times. The organic phase is treated, washed with "Eno Anticromo" bleaching carbon 8 g, for 40 - 45 'and the carbon is filtered, washing the filter with petroleum ether 30 - 50. The filtrate is brought to residue under vacuum; this residue is washed twice with deionized water heated to 55 ° C, 200 ml each time and dried well under vacuum at a maximum temperature of 70 ° C. The oily residue, consisting of the ethyl esters of Omega-3 fatty acids, weighs 75 g. The EPA content is 47.2%; that in DHA, 39.3%; that in total Omega-3, 91%.

EXAMPLE 4

Fish oil EE "45/35 distilled" (minimum 45% EPA and 35% DHA) 100 g, is dissolved in isopropanol ml 300. Under stirring, 1.0 g BHA and a 98% sodium hydroxide solution 12 are added, 8 g in absolute ethanol 150 ml; the mixture is then stirred at room temperature for 12 hours. It is cooled to 0 ° C and the precipitated solid is filtered, washing it with a little isopropanol cooled to 0 ° C. The solid obtained is dissolved in 500 ml of water, 200 ml dichloromethane is added and the mixture is brought to pH ~ 2 with 37% hydrochloric acid, stirring well. It is stirred for a few minutes and the phases are separated; the aqueous phase is extracted again with 100 ml dichloromethane. The combined extracts are washed with 100 ml deionized water and brought to residue under vacuum; an oily residue of free Omega-3 fatty acids is obtained, 82 g. These fatty acids are dissolved in absolute ethanol 400 ml, a 1M solution in heptane of boron trichloride, 23.7 ml, is added, stirred briefly and left at room temperature, in the dark, for 12 hours. It is then brought to reflux for 1 hour and cooled to 0 ° C. The pH is brought to neutral with a 21% solution of sodium ethoxide in ethanol and all the solvent is distilled under vacuum to a residue in a 60 ° C bath. The residue is taken up with petroleum ether, fraction 30 ° - 50 ° C, 400 ml and 100 ml deionized water. It is stirred well and the phases are separated, discarding the lower aqueous phase. The organic phase is washed again with 100 ml deionized water two more times. The organic phase washed with "Eno Anticromo" decolouring carbon 8 g, for 40 - 45 'is treated and the carbon is filtered, washing the filter with petroleum ether 30 - 50. The filtrate is brought to residue under vacuum; this residue is washed twice with deionized water heated to 55 ° C, 200 ml each time and dried well under vacuum at a maximum temperature of 70 ° C. The oily residue, consisting of the ethyl esters of Omega-3 fatty acids, weighs 77.5 g. The EPA content is 47.4%; that in DHA, 48.5%; that in total Omega-3, 97.2%.

EXAMPLE 5

Fish oil EE “45/35” (minimum 45% EPA and 35% DHA), 100 g, is dissolved in 96% ethanol 300 ml. Under stirring, 1.0 g BHT and an 85% 20.7 g potassium hydroxide solution in 150 ml 96% ethanol are added; the mixture is then stirred at room temperature for 12 hours. All the solvent is distilled under vacuum. The residue is dissolved in 500 ml deionized water, obtaining a clear solution, 200 ml dichloromethane is added and the mixture is brought to pH ~ 2 with 37% hydrochloric acid, stirring well. It is stirred for a few minutes and the phases are separated; the aqueous phase is extracted again with 100 ml dichloromethane. The combined extracts are washed with 100 ml deionized water and brought to residue under vacuum; an oily residue of free Omega-3 fatty acids is obtained, 91.5 g. These fatty acids are dissolved in absolute ethanol 400 ml, a 1M solution in heptane of boron trichloride, 23.7 ml, is added, stirred briefly and left at room temperature, in the dark, for 12 hours. It is then brought to reflux for 1 hour and cooled to 0 ° C. The pH is brought to neutral with a ~ 10% solution of sodium hydroxide in 96% ethanol and all the solvent is distilled under vacuum, up to a residue, on a bath at 60 ° C. The residue is taken up with 400 ml ethyl acetate and 100 ml deionized water. It is stirred well and the phases are separated, discarding the lower aqueous phase. The organic phase is washed again with 100 ml deionized water two more times. The organic phase is treated, washed with "Eno Anticromo" bleaching carbon 9 g, for 40 - 45 'and the carbon is filtered, washing the filter with ethyl acetate. The filtrate is brought to residue under vacuum; this residue is washed twice with deionized water heated to 55 ° C, 200 ml each time and dried well under vacuum at a maximum temperature of 70 ° C. The oily residue, consisting of the ethyl esters of Omega-3 fatty acids, weighs 80.1 g. The EPA content is 43.6%; that in DHA, 38.5%; that in total Omega-3, 86.7%.

EXAMPLE 6

Fish oil EE “45/35” (minimum 45% EPA and 35% DHA) 100 g, dissolved in isopropanol 300 ml. Under stirring, 1.0 g BHA and a 98% 12.8 g sodium hydroxide solution in 150 ml absolute ethanol are added; the mixture is then stirred at room temperature for 12 hours. The precipitated solid is filtered, washing it with a little isopropanol at room temperature. The solid obtained is dissolved in 500 ml water, 200 ml dichloromethane is added and the mixture is brought to pH ~ 2 with 37% hydrochloric acid, stirring well, 200 ml dichloromethane is added and the mixture is brought to pH ~ 2 with hydrochloric acid 37%, shaking well. It is stirred for a few minutes and the phases are separated; the aqueous phase is extracted again with 100 ml dichloromethane. The combined extracts are washed with 100 ml deionized water and brought to residue under vacuum; an oily residue of free Omega-3 fatty acids is obtained, 85.4 g. These fatty acids are dissolved in absolute ethanol 400 ml, a 1M solution in heptane of boron trichloride, 23.7 ml, is added, stirred briefly and left at room temperature, in the dark, for 12 hours. It is then brought to reflux for 1 hour and cooled to 0 ° C. The pH is brought to neutral with a ~ 10% solution of sodium hydroxide in 96% ethanol and all the solvent is distilled under vacuum, up to a residue, on a bath at 60 ° C. The residue is taken up with 400 ml ethyl acetate and 100 ml deionized water. It is stirred well and the phases are separated, discarding the lower aqueous phase. The organic phase is washed again with 100 ml deionized water two more times. The organic phase is treated, washed with "Eno Anticromo" decolouring carbon 8 g, for 40 - 45 'and the carbon is filtered, washing the filter with ethyl acetate. The filtrate is brought to residue under vacuum; this residue is washed twice with deionized water heated to 55 ° C, 200 ml each time and dried well under vacuum at a maximum temperature of 70 ° C. The oily residue, consisting of the ethyl esters of Omega-3 fatty acids, weighs 76.3 g. The EPA content is 45.7%; that in DHA 36.3%; that in total Omega-3, 88.7%

EXAMPLE 7

The Omega-3 ethyl ester residue of Example 5), 80.1 g, is added with urea, 88.0 g and methanol, 255 ml. The mixture is homogenized by heating to 55 ° - 60 ° C. It is cooled slowly, in 4 - 6 hours, to 10 - 12 ° C. It is held about 1 hour and the precipitated crystalline solid is filtered, washing it with methanol, 79 ml. The filtrate is brought to residue, under vacuum, in a bath at 60 ° C. The residue is taken up with deionized water 220 ml at 55 ° C, stirred well and decanted, discarding the lower aqueous phase.

It is repeated in the same way with other 220 ml deionized water, at 55 ° C, on the upper oily phase.

The washed oily phase is dried under vacuum at 70 ° C. Purified Omega-3 ethyl esters are obtained, 67.5 g. The EPA content is 44.7%; that in DHA, 43.3%; that in total Omega-3, 94.7%.

EXAMPLE 8

The Omega-3 ethyl ester residue of example 6), 76.3 g, is added with urea, 69.0 g and methanol, 200 ml. The mixture is homogenized by heating to 55 ° - 60 ° C. It is cooled slowly, in 4 - 6 hours, to 10 - 12 ° C. It is held about 1 hour and the precipitated crystalline solid is filtered, washing it with methanol, 62 ml. The filtrate is brought to residue, under vacuum, in a bath at 60 ° C. The residue is taken up with deionized water 173 ml at 55 ° C, stirred well and decanted, discarding the lower aqueous phase.

It is repeated in the same way with other 173 ml deionized water at 55 ° C on the upper oily phase.

The washed oily phase is dried under vacuum at 70 ° C. Purified Omega-3 ethyl esters are obtained, 66.3 g. The EPA content is 47.8%; that in DHA, 42.3%; that in total Omega-3, 96.8%.

EXAMPLE 9

100 g of Omega-3 ethyl esters deriving from one of the preparation methods described in the previous examples, are dissolved in 300 ml of ethanol for food use. This solution is poured, under stirring, on 100 g of precipitated, amorphous silica. The mixture obtained, stirred until completely homogenized, is dried in a vacuum oven at 40 ° C to a residual ethanol lower than 5000 ppm and passed in an oscillating screen equipped with a mesh of the desired fineness or in a suitable mill. The fine granulate obtained is combined, in variable proportions, with excipients suitable for the packaging of tablets and subjected to punching to obtain tablets of the desired dosage. For example: above granulate (Omega-3 ethyl esters / 1/1 precipitated silica) 1000 mg; microcrystalline cellulose 160 mg; croscarmellose sodium 100 mg; PVP (polyvinylpyrrolidone) 60 mg; talc 20 mg; magnesium stearate 60 mg.

Capsule format punch 21 x 10.

EXAMPLE 10

The fine granulate (Omega-3 ethyl esters / 1/1 precipitated silica) obtained as in the previous example, is dry mixed with lubricating powders, such as talc and magnesium stearate; typically, for 1000 mg of granules, corresponding to 500 mg of Omega-3 ethylesters, 20 mg of talc and 60 mg of magnesium stearate are added, mix thoroughly and fill, flush or with a piston filling machine, into sealed capsules of hard jelly, size 0 or 00

Claims (14)

CLAIMS 1. Process for obtaining ethyl esters of a // - Z) -5,8, ll, 14,17-eicosapentaenoic acid (EPA) and of acid (a // - Z) -4,7,10,13,16 , 19-docosahexaenoic DHA from an EE fish oil, having an EPA content greater than 45% and a DHA content greater than 35%, comprising: a) Hydrolysis in a suitable solvent using alkaline hydroxides of the starting oil in the presence of antioxidants; b) Possible separation of the solid sodium salts of the fatty acids obtained from the hydrolysis of step a) precipitated in the reaction environment; c) Dissolution in water of the alkaline hydrolysis product of step a), both as a solid sodium salt separated in step b), and as a total residue of the hydrolysis reaction; d) Precipitation, by acidification of the aqueous solution of step c), of the fatty acids corresponding to the alkaline salts obtained from hydrolysis a), their extraction with a suitable solvent, washing with water and concentration; e) Esterification with acid catalysis in ethanol or in ethanol / water mixtures of the fatty acids obtained in step d), to obtain their ethyl esters; f) Isolation of ethyl esters by neutralization of the reaction environment, concentration, extraction by a suitable solvent of the ethyl esters themselves, washing with water, treatment with bleaching carbon, concentration, final washing with water and drying. g) Possible purification by treatment with urea in methanol; steps a) -g) being carried out in an inert atmosphere. 2. Process according to claim 1, wherein the solvents used in step a) are 96% ethanol, absolute ethanol (greater than 99%) and isopropanol and the antioxidants used are selected from tocopherol, sodium or potassium metabisulfite, BHA, BHT, ascorbic acid, preferably lipophilic antioxidants, more preferably BHT or BHA. 3. Process according to claim 1 or 2 in which the hydrolysis is carried out with sodium or potassium hydroxide in molar excess preferably of 1.1 moles per mol, at a temperature between 15 ° and 20 ° C, for a time of 12 ± 1 hours. 4. Process according to one or more of claims 1 to 3 in which the sodium salts in step b) are separated from the mother liquors at room temperature or between 0 ° and 5 ° C and are washed with absolute ethanol or isopropanol. 5. Process according to one or more of claims 1 to 4 wherein the dissolution of step c) is carried out at room temperature. 6. Process according to one or more of claims 1 to 5 wherein the precipitation of the fatty acids of step d) occurs by adding a slight stoichiometric excess of mineral acid, preferably aqueous hydrochloric acid or dilute sulfuric acid, at a temperature between 0 ° and 20 ° C, up to pH ~ 2; the precipitated acids are extracted with water immiscible solvents, preferably petroleum ether or dichloromethane; the concentration takes place by vacuum distillation of all the solvent, until an oily residue of prevalent Omega-3 fatty acids is obtained. 7. Process according to one or more of claims 1 to 6 wherein the esterification of step e) takes place in ethanol, preferably absolute (greater than 99%), in the presence of acid catalysts, in quantities from 0.01 to 0, 2, preferably 0.1 moles per mol. 8. Process according to claim 7 in which the acid catalyst is boron trichloride, as it is or in a solution of heptane or dichloromethane, and the reaction is carried out for about 12 hours at room temperature and for about an hour at reflux of the solvent. 9. Process according to one or more of claims 1 to 8 in which the isolation of ethyl esters occurs after neutralization with dilute solutions of sodium or potassium hydroxide or with sodium ethoxide in ethanol solution. 10. Process according to one or more of claims 1 to 9 in which the eventual purification of step g) is carried out by adding an amount of urea from 0.9 to 1.1 times by weight. 11. Process according to one of claims 1 to 10 in which the hydrolysis a) is carried out in the presence of potassium hydroxide and step b) is absent. 12. Ethyl esters of a // - Z) -5,8, 11, 14,17-eicosapentaenoic acid (EPA) and of acid (a // - Z) -4,7,10,13,16,19- docosahexaenoic (DHA) obtained from the process of claims 1-10 adsorbed on solid inert supports, preferably precipitated amorphous silica. 13. Pharmaceutical or nutraceutical compositions comprising as active ingredient the ethyl esters of claim 12. 14. Compositions according to claim 13 in the form of sealed hard gelatin capsules or tablets. Milan, 8 October 2012