WO2007080514A2

WO2007080514A2 - A method for the extraction of lipid fractions from krill

Info

Publication number: WO2007080514A2
Application number: PCT/IB2007/000098
Authority: WO
Inventors: Peter Mose Larsen; Stephen John Fey; Jesper Breuning; Bent Ludvigsen
Original assignee: Krill A/S; Alfa Laval Copenhagen A/S
Priority date: 2006-01-13
Filing date: 2007-01-15
Publication date: 2007-07-19
Also published as: AR059659A1; WO2007080514A3

Abstract

According to the present disclosure there is provided a method for extracting lipid fractions from krill, wherein freshly captured krill is ground to produce a slurry, which is gently heated to a temperature below 90°C for less than 45 minutes, whereafter the liquid into an aqueous phase and a krill oil phase from which a krill oil extract is derived without the use of organic solvents. Moreover there is also provided a pharmaceutical composition for the treatment of thrombosis in a patient comprising an effective amount of a krill oil extract.

Description

A METHOD FOR THE EXTRACTION OF LIPID FRACTIONS FROM KRILL

FIELD OF THE INVENTION

This invention relates to a novel method and equipment for extracting lipid fractions from marine and aquatic animals such as krill, Calanus, fish and sea mammals, which method does not utilize high temperatures and/or organic solvent and thus preserves valuable components in the extracted animal material.

BACKGROUND OF THE INVENTION

Marine and aquatic animal oils and fractions thereof contain various therapeutic agents. For example, it is reported that various marine and aquatic animal oils have anti-inflammatory properties. Marine and aquatic animal oils are also reported as helpful in reducing the incidence of cardiovascular disease. Also, some marine and aquatic animal oils are reported as suppressing the development of certain forms of lupus and renal diseases. As a further example, krill may be used as a source of enzymes for debridement of ulcers and wounds or to facilitate food digestion. Also marine and aquatic oils contain various antioxidants, which may have potential therapeutic properties.

Krill is the common name for small, shrimp-like crustaceans that swarm in dense shoals, especially in Antarctic waters. It is one of the most important food sources (especially protein) for fish, some kind of birds and especially for baleen whales. Krill is also a good source of omega-3 fatty acid, which are well known for their beneficial effects on human health.

It is known in the art to use krill and/or marine enzymes for the treatment of a great variety of diseases in human and animals such as infections, inflammations, cancers, HIV/AIDS, pain, polyps, warts, haemorrhoids, plaque, wrinkles, thin hair, allergic itch, anti-adhesion, eye disease, acne, cystic fibrosis and immune disorders including autoimmune diseases and cancer.

It is also known in the art that krill and/or marine oils may be used for the treatment of autoimmune lupus and other autoimmune diseases and can also be used for treating cardiovascular diseases. Various methods for extracting marine and aquatic animal oils are known. For example, it is known to extract fish oil using organic solvents such as hexane and ethanol. It is also known to measure the fat content in fish muscle tissue using solvents such as acetone.

U.S. Pat. No. 4,331,695 describes a method using pressurized solvents which are gaseous at room temperature, such as propane, butane or hexane. The extraction is performed at preferred temperatures of 15 to 80° C on shredded vegetable or finely divided animal products. The extracted oils are then made to precipitate under high pressure and elevated temperatures of 50 to 200° C. However, hexane is a poor extraction solvent for marine animals such as krill. Furthermore, the high temperatures used in the precipitation step negatively alters the lipids.

Canadian Patent Application 2,115,571 describes a method for extracting oils from various brown and read algae species. The method provides, for example, extraction using nearly pure ethanol for 40 hours.

U.S. Pat. No. 5,006,281 describes a method for extracting oil from marine and aquatic animals such as fish. The marine and aquatic animal is first treated with an antioxidant compound, finely divided and centrifuged to separate the oil phase from the aqueous phase and solid phase. The oil phase is then further treated with antioxidant to remove undesirable odour or taste.

Canadian Patent 1 ,098,900 describes a method for extracting oils from krill. The method involves emulsifying fresh or defrosted krill in an aqueous medium. The oil fraction is recovered by centrifugation.

Folch published in J. biol. Chem. 226: 497-509 in 1957 an article entitled "A simple method for the isolation and purification of total lipids from animal tissues" proposes an extraction method using chloroform and methanol. This method is not commercially feasible because of the toxicity of the solvents involved.

However, most of the krill oil extract used for these treatments has only conserved its omega-3 fatty acids as active ingredients, which is a very small part of all the active ingredients of the krill itself. This fact dramatically reduces the potential of the krill and/or marine oil as a treatment for human diseases. There is an increasing demand for treatments using products derived from a natural source, therefore, it would be highly desirable to be provided with a krill and/or marine extract having an enhanced potential for prevention and/or treatment and/or management of disease.

US Patent 6,800,299 discloses a method for extracting lipid fractions from marine and aquatic animal material by acetone extraction. The resulting non-soluble and particulate fraction is preferably subjected to an additional solvent extraction with an alcohol, preferably ethanol, isopropanol or t-butanol or an ester of acetic acid, preferably ethyl acetate to achieve extraction of the remaining soluble lipid fraction from the marine and aquatic animal material. The remaining non-soluble particulate content is also recovered since it is enriched in proteins and contains a useful amount of active enzymes. Also provided herein is a krill extract. It is reported that these marine and aquatic animal oils have anti-inflammatory properties. Marine and aquatic animal oils are also reported as helpful in reducing the incidence of cardiovascular disease. As a further example the patent mentions that krill may be used as a source of enzymes for debridement of ulcers and wounds or to facilitate food digestion.

WO02102394A2 discloses a process for the preparation of a krill oil extract, which process includes the steps of placing krill and/or marine material in a ketone solvent to achieve extraction of the soluble lipid fraction from the krill; then separating the liquid and solid contents; then recovering a first lipid rich fraction from the liquid contents by evaporation of the solvent present in the liquid contents; then placing the solid contents in an organic solvent to achieve extraction of the remaining soluble lipid fraction from the krill material; then separating the liquid and solid contents; then recovering a second lipid rich fraction by evaporation of the solvent from the liquid contents; and finally recovering the solid contents. Diseases that can be treated and/or prevented by using the krill oil extract are inter alia cardiovascular diseases. In this respect it is mentioned that the Krill oil has been shown to decrease cholesterol in vivo, inhibit platelet adhesion and plaque formation and reduce vascular endothelial inflammation in a patient.

Canadian Patent 1 ,098,900 describes a method for extracting oils and producing proteins from krill comprising emulsification of lipids of krill in an aqueous medium, separation of the emulsion of lipids from the krill mass, alkaline extraction of proteins from the krill mass, separation of the protein extract produced from chitin integuments, and finally separation of protein from the protein extract. The document mentions that krill is a prospective source of food and other practically useful products such as chitin and lipids which find wide application in different branches, such as food industry, textile, and medicine.

WO03011873A2 discloses a phospholipid extract from inter alia krill, with therapeutic properties, such as those essential for the maintenance of a healthy cardiovascular system. The phospholipid extract comprises a variety of phospholipids, fatty acid, metals and a novel flavonoid. The method for the preparation of this extract is generally carried out by a method similar to the one described in US Patent 6,800,299 (see above; and utilises organic solvents), which procedure produces two successive lipid fractions and a dry residue enriched in protein, including active enzymes.

WO8401715A1 and WO09533471A1 disclose various aspects of so-called krill enzymes, which are water-soluble. It is mentioned that in krill a mixture of different enzymes exists, such as e.g. proteinases (with acidic and neutral-to-alkaline pH- optima), peptidases (exo- and endopeptidases), lipases, phospholipases, amylases and other carbohydrate degrading enzymes, phosphatases nucleases, nucleotidases and esterases. The proteolytic (trypsin-like) activity existing in a water extract from krill has been studied and described. WO09533471A1 disclose the use of one or more krill enzymes for the manufacture of an intravasal pharmaceutical composition for thrombolysis in a mammal host.

As appears from the above cited prior art current available technology for production of marine oils are usually based on a thermal process such as those used in the fish meal process, and in some particular cases on a solvent extraction method, such as hexane extraction or similar highly effective solvent. However the solvent method which is necessary to get the optimum oil quality, cannot be implemented on a fishing vessel,. In addition the most active solvents are not suitable for pharmaceutical processing.

Thus, the prior art disclose extraction methods involving the use of organic solvents or high temperatures or both - none of which are optimal from the point of view of the biological activity of the oil. SUMMARY OF THE INVENTION

In accordance with the present invention there is provided a novel marine lipid extract obtainable by a process with following characteristics:

• Processing temperature below 60 ⁰C, and more precisely with an optimum processing just under 27 ⁰C.

• Mechanical and physical disruption of the lipid cell membrane to facilitate low temperature extraction. • Processing takes place under inert gas to prevent oxidation or denaturation of fat and proteins

• Intermediate processing tanks kept at a minimum level to reduce residence time.

• The oil is frozen immediately after recovery to stabilize it.

In accordance with the present invention, there is provided a novel lipid extract obtainable by the process of the present invention. The novel lipid extract is derived from marine raw materials found in any marine environment around the world, for example, for krill, the Antarctic ocean (euphasia superba), the Pacific ocean (euphasia pacifica), the Atlantic ocean, the Indian ocean, in particular coastal regions of Mauritius Island and/or Reunion Island of Madagascar, Canadian West Coast, Japanese Coast, St-Lawrence Gulf and Fundy Bay, but other sources of marine raw materials obviously exist.

The present invention describes a method for extracting lipid fractions from marine raw materials, said method comprising the steps of:

• Feeding freshly captured raw material into a grinder to produce a slurry comprising particles of less than 5 mm prior to further processing • Ultra sound disintegration of cell membranes to expose the marine oil without heating the slurry appreciably

• Heating the slurry gently to a temperature below 60⁰C, preferably between 25°C and 27°C

• Separating the solid material from the liquid • Separating the liquid into an aqueous phase and a lipid phase wherein the extraction does not involve the use of organic solvents or temperatures above 60⁰C, and each process step takes place in an inert gas enviroment.

Additionally according to the present invention there is provided a apparatus for performing the process of the invention.

DETAILED DESCRIPTION OF THE INVENTION

Specifically, according to a first aspect of the present invention there is provided a method for extracting lipid fractions from marine raw materials, such as krill, said method comprising the steps of:

• supplying an inert gas (e.g. nitrogen) to reduce or exclude oxygen

• placing the raw material in a grinder to mechanically disrupt fat cell membranes

• ultra sound sonication of the grinded material to facilitate extraction of the soluble lipid fraction;

• separating the liquid and solid components;

• recovering a lipid rich fraction from the liquid component; wherein the extraction is performed at a temperature below 60 ⁰C, preferably below 30⁰C, and does not involve the use of organic solvents.

Specifically, according to a second aspect of the present invention there is provided an apparatus for carrying out the method of the present invention comprising • means for grinding and ultra sound sonication of the marine raw material,

• means for extracting the majority of the liquid fraction of the material,

• means for recovering the light phase of the liquid fraction,

• means for recovering a concentrated marine lipid,

• means for accumulating remaining solid components and subsequently pressing residue liquid therefrom,

• means for returning the additional extracted liquid to means for recovering the additional lipid extract Example 1

Preparation of the lipid extract of the present invention.

The invented process may be performed with an apparatus comprising one or more of components referred to in Figure 1 , and in particular:

53.01. Grinding of the product and addition of extraction water - Max. 5 mm holeplate grinder.

53.02. Feed pump for ultra sound sonicator, Contherm and decanter.

53.03. Heavy duty ultrasound sonication to disintegrate cell membrane. 53.04. Contherm scraped heat exchanger - Temperature condition by heating to maximum 30 ⁰C

53.05. Decanter centrifuge for extracting liquid.

53.06. Feed pump for separator

53.07. Concentrator centrifuge to recover emulsified fat phase 53.08. Clarifying centrifuge to recover the clean marine oil

53.09. Slurry pump for heavy and sludge phases from centrifuges

53.10. Solid phase pump

53.11. High pressure dewatering press - applying a pressure of minimum 15 bar to press residual liquid from product 53.12. Pump to return additional extracted liquid to a process step prior to the decanter

53.13. Press cake pump

53.14. Contherm pasturising the press cake before freezing.

A preferred embodiment comprises at least:

• means for grinding (53.01) and ultra sound sonication of (53.03) the marine raw material,

• means for extracting the majority of the liquid fraction of the material (53.05), • means for recovering the light phase of the liquid fraction (53.07),

• means for recovering a concentrated marine lipid (53.08), • means for accumulating (53.09, 53.10) remaining solid components and subsequently pressing residue liquid therefrom (53.11), and

• means for returning (53.12) the additional extracted liquid to means for recovering the additional lipid extract (53.07).

Preferably the entire process is carried out under an inert atmosphere, such as nitrogen or helium.

The inventors have also envisaged that the present invention may be carried out by applying supercritical CO2 extraction.

Claims

1. A method for extracting lipid fractions from marine raw materials, such as krill, said method comprising the steps of: • placing the raw material in a grinder to mechanically disrupt fat cell membranes;

• Ultra sound sonication of the ground material to facilitate extraction of the soluble lipid fraction;

• separating the liquid and solid components; • recovering a lipid rich fraction from the liquid component;

• recovering a protein rich cake; wherein the extraction is performed at a temperature below 60 ⁰C and does not involve the use of organic solvents.

2. A method as in claim 1 , wherein separating the liquid and solid components is effected by techniques selected from the group consisting of mechanical pressure, filtration, centrifugation and sedimentation.

3. A method as in claim 1 , wherein the extraction is performed at a temperature between 25⁰C and 27⁰C.

4. A method as in claim 1 , wherein the extraction is performed under an inert atmosphere.

5. A marine lipid extract obtainable by a method according to any one of claims 1 to 4.

6. A method according to any one of claims 1 to 4, wherein the marine raw material is krill.

7. A krill oil extract obtainable by a method according to claim 6.

8. An apparatus for carrying out the method of any one of the claims 1 to 4 comprising

• means for grinding (53.01) and ultra sound sonication of (53.03) the marine raw material, • means for extracting the majority of the liquid fraction of the material (53.05),

• means for recovering the light phase of the liquid fraction (53.07), • means for recovering a concentrated marine lipid (53.08)

• means for accumulating (53.09, 53.10) remaining solid components and subsequently pressing residue liquid therefrom (53.11), and

9. An apparatus for carrying out the method of any one of the claims 1 to 4 comprising:

• Feed pump for ultra sound sonicator, Contherm and decanter, • Heavy duty ultrasound sonication to disintegrate cell membrane,

• Contherm scraped heat exchanger - Temperature condition by heating to maximum 30 ⁰C,

• Decanter centrifuge for extracting liquid,

• Feed pump for separator, • Concentrator centrifuge to recover emulsified fat phase,

• Clarifying centrifuge to recover the clean marine oil,

• Slurry pump for heavy and sludge phases from centrifuges,

• Solid phase pump,

• High pressure dewatering press - applying a pressure of minimum 15 bar to press residual liquid from product,

• Pump to return additional extracted liquid to a process step prior to the decanter,

• Press cake pump, and

• Contherm pasturising the press cake before freezing.

10. An apparatus according to claim 8 or 9, wherein the method is carried out under an inert atmosphere.