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`APPLICATION NUMBER FEINor37 (C) GRP ART UNIT|FIL FEE REC'D TOT CLAIMS|IND CLAIMSATTY.DOCKET.NO
`
`
`
`
`
`60/920,483
`
`03/28/2007
`
`200
`
`NATNUT-14409
`
`23535
`MEDLEN & CARROLL, LLP
`101 HOWARD STREET
`SUITE 350
`SAN FRANCISCO, CA94105
`
`CONFIRMATION NO. 4952
`
`FILING RECEIPT
`
`Date Mailed: 04/20/2007
`
`It will not be examined for patentability and
`Receipt is acknowledged of this provisional Patent Application.
`will become abandoned not
`later than twelve months after its filing date. Be sure to provide the U.S.
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`incorporating the requested corrections (if appropriate).
`
`Applicant(s)
`
`Inge Bruheim, Volda, NORWAY;
`Asgier Saebo, Volda, NORWAY;
`Mikko Griinari, Espoo, FINLAND;
`
`Powerof Attorney:
`John Jones--44174
`
`If Required, Foreign Filing License Granted: 04/18/2007
`
`The country code and number of your priority application, to be usedfor filing abroad under the Paris Convention, is
`US60/920,483
`
`Projected Publication Date: None, application is not eligible for pre-grant publication
`
`Non-Publication Request: No
`
`Early Publication Request: No
`
`Title
`
`Bioeffective krill oil compositions
`
`PROTECTING YOUR INVENTION OUTSIDE THE UNITED STATES
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`Since the rights granted by a U.S. patent extend only throughoutthe territory of the United States and have
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`international application under the Patent Cooperation Treaty (PCT). An international (PCT) application
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0001
`
`page 0001
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`

`

`generally has the same effect as a regular national patent application in each PCT-member country. The
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`RIMFROST EXHIBIT 1005
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`RIMFROST EXHIBIT 1005 page 0002
`
`page 0002
`
`

`

`NOTGRANTED
`
`if the phrase "IF REQUIRED, FOREIGN
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`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0003
`
`page 0003
`
`

`

`'
`
`.
`
`U.S. PTO
`60/920483
`PTO/SB/16 (12-04)
`03/28/2007
`3
`Approvedfor use through 07/31/2006. OMB 0651-0032
`2. 23
`U.S. Patent and Trademark Office; U.S. DEPARTMENT OF COMMERCE
`== o
`o== 2 Under the Paperwork Reduction Act of 1995, no persons are required to respondto a collection of information unlessit displays a valid OMB control number.
`Ss
`PROVISIONAL APPLICATION FOR PATENT COVER SHEET
`CO ==
`This is a requestfor filing a PROVISIONAL APPLICATION FOR PATENTunder 37 CFR 1.53(c).
`C= ESpress Mail Label No. EV 851 964 955 US
`
`0
`Gwen Name(first and middle [if any])
`
`INVENTOR(S)
`Family Nameor Surname
`
`Residence
`City and either State or Foreign Count
`
`Inge
`Asgier
`Mikko
`
`Bruheim
`Saebo
`Griinari
`
`Norway
`Norway
`mg Fintand
`
`separately numbered sheets attached hereto
`Additional inventors are being named on the
`TITLE OF THE INVENTION (500 characters max):
`
`
`
`
`
`
`
` Directall correspondenceto:
`CORRESPONDENCE ADDRESS
` 23535
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`-
`. Aduplicative copy of this form is enclosed for fee processing.
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`
` The invention was made by an agencyof the United States Governmentor undera contract with an agencyofthe United States Government.
`No.
`C] Yes, the name of the U.S. Government.agency and the Government contract numberare:
`
`
`{A\
`
`
`Date March 28, 2007
`‘
`
`TYPED or PRINTED NAME
`REGISTRATION NO. 44,174
`(if appropriate)
`TELEPHONE 608/218-6900 Docket Number:_NATNUT-14409
`
`USE ONLY FORFILING A PROVISIONAL APPLICATION FOR PATENT
`This collection of information is required by 37 CFR 1.51. The information is required to obtain or retain a benefit by the public whichis to file (and by the USPTO
`to process) an application. Confidentiality is governed by 35 U.S.C. 122 and 37 CFR 1.11 and 1.14. This collection is estimated to take 8 hours to complete,
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`ADDRESS. SEND TO: Commissionerfor Patents, P.O. Box 1450, Alexandria, VA 22313-1450.
`if you need assistance in completing the form, call 1-800-PTO-9199 and select option 2.
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`METHOD OF PAYMENT OF FILING FEES AND APPLICATION SIZE FEE FOR THIS PROVISIONAL APPLICATION FOR PATENT
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`[_] Applicant claims small entity status. See 37 CFR 1.27.
`TOTAL FEE AMOUNT($)
`Accheck or moneyorderis enclosed to coverthefiling fee and application size fee (if applicable).
`200.00
`
`Bioeffective Krill Oil Compositions
`
`Paymentby credit card. Form PTO-2038is attached
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0004
`
`page 0004
`
`

`

`Express Mail Label No.: EV 851 964 955 US
`17667 U.S. PTO
`
`PATENT
`Attorney Docket No.: NATNUT-14409
`
`IN THE UNITED STATESPATENTAND TRADEMARKOFFICE
`mn |
`In re applicationof:
`Inge Bruheim,et al
`
`For:
`
`Anti-inflammatory Properties of Marine Lipid Compositions
`
`Mail Stop Provisional Patent Application
`Commissioner for Patents
`P.O. Box 1450
`Alexandria, Virginia 22313-1450
`
`CERTIFICATION UNDER37 C.F.R. § 1.10
`
`I herebycertify that this correspondence and the documents referred to as attached therein are
`being deposited with the United States Postal Service on March 28, 2007, in an envelope as
`"EXPRESS MAIL POST OFFICE TO ADDRESSEE"service under 37 C.F.R. § 1.10, Mailing
`
`pnt Application,
`
`TRANSMITTAL COVER SHEETFOR FILING PROVISIONAL APPLICATION
`(37 C.E.R. § 1.51(c)(1))
`
`This is a request for filing a PROVISIONAL APPLICATION FOR PATENTunder 37 C.F.R. § 1.53(c).
`
`1.
`
`2.
`
`The following comprises the information required by 37 C.F.R. § 1.51(c):
`
`The name(s) of the inventor(s) is/are (37 C.F.R. § 1.51(c)(1)(i)):
`
`Inge Bruheim
`Asgeir Saebo
`MikkoGriinari
`
`3.
`
`Address(es) of the inventor(s), as numbered above (37 C.F.R. § 1.51(c)(1)Gii)):
`
`Martavegen 6A, 6100 Volda, NO
`Bogardsvegen 12, 6100 Volda, NO
`Kultatinnantie 1B, 02660 Espoo, FI
`
`4.
`
`Thetitle of the invention is (37 C.F.R. § 1.51(c)(1)(iv)):
`
`Bioeffective Krill Oil Compositions
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0005
`
`page 0005
`
`

`

`Express Mail Label No.: EV 851 964 955 US
`
`PATENT
`Attorney Docket No.: NATNUT-14409
`
`5.
`
`The name,registration, and telephone numberofthe attorney (ifapplicable) is (37 C.F.R.
`§ 1.51(c)C1)C¥)):
`
`J. Mitchell Jones
`Reg. No.: 44,174
`Tel.:
`(608) 218-6900
`
`6.
`
`7.
`
`8.
`
`—_
`
`A Power of Attorney accompaniesthis cover sheet.
`
`The docket numberusedto identify this application is (37 C.F.R. § 1.51(c)(1)(vi)):
`
`Docket No.: NATNUT-14409
`
`The correspondence addressforthis application is (37 C.F.R. § 1.51(¢)CL)vii)):
`
`MEDLEN & CARROLL, LLP
`101 Howard Street, Suite 350
`San Francisco, California 94105
`
`Statement as to whether invention was madeby an agency of the U.S. Governmentor under
`contract with an agency of the U.S. Government. (37 C.F.R. § 1.51(c)(1)(viii)):
`
`This invention was madeby an agency of the United States Government, or under contract with an
`agency of the United States Government.
`
`x No.
`
`—
`
`Yes.
`
`9.
`
`Identification of documents accompanyingthis cover sheet:
`
`A.
`
`Documents required by 37 C.F.R. § 1.51(c)(2)-(3):
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`Specification:
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`No. of pages
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`39
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`Drawings:
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`No.of sheets
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`B.
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`Additional documents:
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`xX
`
`Claims:
`
`No. of claims
`
`56
`
`—
`
`—
`
`—_—
`
`_
`
`Powerof Attorney
`
`Small Entity Statement
`
`Assignment
`
`Other
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0006
`
`page 0006
`
`

`

`Express Mail Label No.: EV 851 964 955 US
`
`PATENT
`Attorney Docket No.: NATNUT-14409
`
`10.
`
`Fee
`
`The Filing Fee for this provisional application, as set forth in the Consolidated Appropriations Act
`for Fiscal Year 2005, is $200.00 for other than a small entity, and $100.00 for a small entity.
`
`The Application Size Fee for this provisional application, as set forth in the Consolidated
`Appropriations Act for Fiscal Year 2005, is $250.00 for other than a small entity, and $125.00 for
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`_
`
`Applicant is a small entity.
`
`11.
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`Small Entity Statement
`
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`
`Verified statement that this is a filing by a small entity under 37 C.F.R. § 1.27.
`
`12.
`
`Fee payment being madeat this time
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`Charge Deposit Account No. 08-1290 in the amount of $
`duplicate of this transmittal is enclosed for this purpose.
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`. An originally executed
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`Method of Fee Payment:
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`xX
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`Check in the amount of $200.00 enclosed.
`
`Charge Deposit Account No. 08-1290 in the amountof$.
`duplicate of this transmittal is enclosed for this purpose.
`
`
`
`. An originally executed
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`XxX
`
`The Commissioneris hereby authorized to charge any deficiency in the paymentofthe
`required fee(s), and/or credit any overpayment, to Deposit Account No.: 08-1290. An
`originally executed duplicate of this transmittal is enclosed for this purpose.
`
`28,2007 . Mitchell Jones
`
`Date:March
`
`Reg. No.: 44,174
`
`MEDLEN & CARROLL, LLP
`101 Howard Street, Suite 350
`San Francisco, California 94105
`(608) 218-6900
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0007
`
`page 0007
`
`

`

` -EVAS51964955y8
`
`Bioeffective krill oil compositions
`
`Inventors: Inge Bruheim (Volda, Norway), Asgeir Szebe (Eidsnes, Norway), and Mikko
`
`Griinari (Espoo, Finland).
`
`FIELD OF THE INVENTION
`
`This invention relates to methods for producing and refining phospholipids with desired
`
`omega-3 fatty acid levels, and in particular to phospholipid and fatty acid compositions
`
`obtained from Antarctic krill and methods for processing such phospholipids and fatty acids.
`
`BACKGROUNDOF THE INVENTION
`
`In the Southern Ocean, off the coast of Antarctica, Antarctic krill (Euphausia superba) can be
`
`found in large quantities, ranging from 300-500 million metric tons of biomass. It feeds on
`
`phytoplankton during the short Antarctic summer. During winter, however, its food supply ts
`
`limited to ice algae, bacteria, marine detritus as well as depleting body protein for energy [1].
`
`For this reason, the nutritional values of krill varies during the season and to some extent
`
`annually [2].
`
`In order to accommodate variations in food supply, krill has developed an
`
`efficient enzymatic digestive apparatus resulting in a rapid breakdown of the proteins into
`
`amino acids [3]. This autoproteolysis is highly efficient also post mortem, making it a
`
`challenge to catch and processthe krill in a way that preserves the nutritional quality of the
`
`krill.
`
`In orderto isolate the krill oil from the krill, solvent extraction methods have been used [7-8].
`
`Krill lipids have been extracted by placing the material in a ketone solvent (e.g. acetone) in
`
`order to extract the lipid soluble fraction. This method involves separating the liquid and solid
`
`contents and recovering a lipid rich fraction from the liquid fraction by evaporation. Further
`
`processing steps include extracting and recovering by evaporation the remaining soluble lipid
`
`fraction from the solid contents by using a solvent such as ethanol [8]. The compositions
`
`produced by these methods are characterized by containing at
`
`least 75 jg/g astaxanthin,
`
`preferably 90 g/g astaxanthin. Another krill lipid extract disclosed contained at least 250
`
`ug/g canastaxanthin [8], preferably 270 ug/g canastaxanthin.
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0008
`
`page 0008
`
`

`

`Krill oil compositions have been described as being effective for decreasing cholesterol,
`
`inhibiting platelet adhesion,
`
`inhibiting artery plaque formation, preventing hypertension,
`
`controlling arthritis symptoms, preventing skin cancer, enhancing transdermal
`
`transport,
`
`reducing the symptoms of premenstrual symptomsor controlling blood glucose levels in a
`
`patient.
`
`[9]
`
`In yet another application, a krill oil composition has been disclosed [4]
`
`comprising a phospholipid and/or a flavanoid. The phospholipid content in the krill
`
`lipid
`
`extract could be as high as 60% w/w and the EPA/DHAcontentas high as 35% (w/w).
`
`Furthermore, nutraceuticals, pharmaceuticals and cosmetics comprising the phospholipid
`extract were disclosed. Previously, it was also shownthat supercritical fluid extraction using
`neat CO. could be used to prevent the extraction of phospholipids in order to extract the
`
`lipid fraction from krill, which comprised of esterified and free astaxanthin [10].
`neutral
`Supercritical fluid extraction with solvent modifier has previously been used to extract marine
`phospholipids
`from salmon roe [11], but has not been previously used to extract
`
`phospholipids from krill meal.
`
`The methods described above rely on the processing of frozen krill that are transported from
`
`the Southern Ocean to the processing site. This transportation is both expensive and can
`
`result in degradation of the krill starting material. Data in the literature showing a rapid
`
`decomposition of the oil in krill explains why somekrill oil currently offered as an omega-3
`
`supplement
`
`in the marketplace contains very high amounts of partly decomposed
`
`phosphatidylcholine and also partly decomposed glycerides. Saether et al., Comp. Biochem
`
`Phys. B 83B(1): 51-55 (1986). The products offered also contain high levels of free fatty
`
`acids.
`
`Whatis needed in the art are methods for processing krill that do not require transport of
`
`frozen krill material over long distances and the products produced by those methods.
`
`SUMMARYOF THE INVENTION
`
`In a first aspect of the invention is a composition characterized by comprising at least 65%
`
`(w/w) phospholipids.
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0009
`
`page 0009
`
`

`

`In another aspect of the invention is a composition obtained from aquatic or marine sources,
`
`characterized by comprising 65% (w/w) phospholipids.
`
`In yet another aspect of the invention is a composition obtained from krill, characterized by
`
`comprising at least 65% (w/w) phospholipids.
`
`In another aspect of the invention is a composition obtained from krill, characterized by
`
`comprising at least 65% (w/w) phospholipids and at least 39% omega-3 fatty acids (w/w).
`
`In yet another aspect of the invention is a composition obtained from krill, characterized by
`
`comprising at least 65% (w/w) phospholipids, at least 39% omega-3 fatty acids (w/w) and at
`
`least 580 mg/kg astaxanthinesters.
`
`In another aspect of the invention is a composition obtained from krill, characterized by
`
`comprising at least 39% omega-3 fatty acids (w/w)and at least 580 mg/kg astaxanthin esters.
`
`In yet another aspect of the invention is a composition obtained from krill, characterized by
`
`comprising at least 65% (w/w) phospholipids and at least 580mg/kg astaxanthinesters.
`
`In yet another aspectis a krill oil effective for reducing insulin resistance, improving blood
`
`lipid profile, reducing inflammation or reducing oxidative stress.
`
`In some embodiments, the present invention provides a composition comprising at least 65%
`
`(w/w) of phospholipids, said phospholipids characterized in containing at least 35% omega-3
`
`fatty acid residues.
`
`In some preferred embodiments, the composition is derived from a marine
`
`or aquatic biomass.
`
`In some further preferred embodiments, the composition is derived from
`
`krill.
`
`In some embodiments, the composition comprises less than 2% free fatty acids.
`
`In
`
`some embodiments, composition comprises less than 10% triglycerides.
`
`In some preferred
`
`embodiments, the phospholipids comprise greater than 50% phosphatidylcholine.
`
`In some
`
`embodiments, the composition comprises at
`
`least 500 mg/kg astaxanthin esters.
`
`In some
`
`the composition comprises at least 500 mg/kg astaxanthin esters andatleast
`embodiments,
`36% (w/w) omega-3 fatty acids.
`In some embodiments, the composition comprises less than
`
`about 0.5g/100g total cholesterol. In some embodiments, the composition comprises less than
`
`about 0.45% arachidonic acid (w/w).
`
`RIMFROST EXHIBIT 1005
`
`RIMFROST EXHIBIT 1005 page 0010
`
`page 0010
`
`

`

`In some embodiments, the present invention providesa krill lipid extract comprising at least
`
`500 mg/kg astaxanthin esters and at
`
`least 36% (w/w) omega-3 fatty acids.
`
`In further
`
`embodiments,
`
`the present invention provides a krill
`
`lipid extract comprising at
`
`least 100
`
`mg/kg astaxanthin esters, at least 20% (w/w) omega-3 fatty acids, and less than about 0.45%
`
`arachidonic acid (w/w).
`
`In some embodiments, the present invention provides methods comprising administering the
`
`foregoing compositions to a subject in an amounteffective for reducing insulin resistance,
`
`reducing inflammation, improving blood lipid profile and reducing oxidativestress.
`
`In some embodiments, the present invention provides a krill lipid extract comprising greater
`
`than about 80% triglycerides and greater than about 90 mg/kg astaxanthin esters.
`
`In some
`
`embodiments, the krill lipid extract is characterized in containing from about 5% to about
`
`15% omega-3 fatty acid residues.
`
`In some embodiments,
`
`the krill
`
`lipid extract
`
`is
`
`characterized in containing less than about 5% phospholipids.
`
`In some embodiments,, the
`
`krill lipid extract is characterized in comprising from about 5% to about 10% cholesterol.
`
`In some embodiments, the present invention provides a krill meal composition comprising
`
`less than about 50g/kg total fat.
`
`In some embodiments, the krill meal composition comprises
`
`from about 5 to about 20 mg/kg astaxanthin esters.
`
`In some embodiments, the krill meal
`
`composition comprises greater than about 65% protein.
`
`In some embodiments, the krill meal
`
`composition of comprises greater than about 70% protein.
`
`In some further embodiments, the
`
`present invention provides an animal feed comprising the krill meal of Claim 21.
`
`In some embodiments, the present invention provides methods of increasing flesh coloration
`
`in an aquatic species comprising feeding said aquatic species a composition comprising the
`
`krill meal described above.
`
`In some embodiments, the present invention provides methods of
`
`increasing growth and overall survival rate of aquatic species by feeding the krill meal
`
`described above.
`
`In some embodiments,
`
`the present
`
`invention provides methods of producing krill oil
`
`comprising: a) providing krill meal; and b) extracting oil from said krill meal.
`
`In some
`
`embodiments, the krill meal! is produced by heat-treating krill.
`
`In some embodiments, the krill
`
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`meal
`
`is stored prior to said extraction step.
`
`In some embodiments,
`
`the extracting step
`
`comprises extraction by supercritical fluid extraction.
`
`In some embodiments, the supercritical
`
`fluid extraction is a two step process comprising an extraction with carbon dioxide and an
`
`extraction including ethanol as an entrainer.
`
`In some embodiments, the present invention
`
`provides an oil produced by the foregoing method.
`
`In some embodiments, the present
`
`invention provides methods of production of krill oil
`
`comprising: a) providing fresh krill; b) treating said fresh krill
`
`to denature lipases and
`
`phospholipases in said fresh krill to provide a denatured krill product; and c) extracting oil
`
`from said denatured krill product.
`
`In some embodiments, the denaturation step comprises
`
`heating of said fresh krill.
`
`In some embodiments, the denaturation step comprises heating
`
`said fresh krill after grinding.
`
`In some embodiments, the methods further comprise storing
`
`said denatured krill product at room temperature or below between the denaturation step and
`
`the extraction step.
`
`In some embodiments, the enzyme denaturation step is achieved by
`
`application of heat.
`
`In some embodiments, the extraction step comprises use of supercritical
`
`carbon dioxide, with or without use of a polar modifier.
`
`In some embodiments, the extraction
`
`step comprises use of ethanol.
`
`In some embodiments, the extraction step is comprises ethanol
`
`extraction followed by acetone to precipitation of phospholipids.
`
`In some embodiments,the
`
`denatured krill product is a meal.
`
`In some embodiments, the present invention provides anoil
`
`produced by the foregoing method.
`
`In some embodiments,
`
`the present
`
`invention provides a composition comprising an oil
`
`extracted from krill having a phosphatidylcholine content of greater then about 50% (w/w).
`
`In some embodiments, the oil has a phosphatidylcholine content of greater then about 70%
`
`(w/w).
`
`In some embodiments, the oil has a phosphatidylcholine content of greater then about
`
`80% (w/w).
`
`In some embodiments, the composition comprises less than 2% free fatty acids.
`
`In some embodiments, the composition comprises less than 10% triglycerides.
`
`In some
`
`embodiments, the composition comprises at least 500 mg/kg astaxanthin esters.
`
`In some
`
`embodiments, the composition comprises less than about 0.45% arachidonic acid (w/w).
`
`In some embodiments, the present invention provides composition comprising odorless krill
`
`oil.
`
`In some embodiments, the odorless krill oil comprises less than about 10 mg/kg (w/w)
`
`trimethylamine.
`
`In some further embodiments, the present invention provides an odorless
`
`krill oil produced by the method comprising: extracting a neutral krill oil from a krill oil
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`containing material by supercritical fluid extraction to provide a deodorized krill material,
`
`wherein said neutral krill oil contains odor causing compoundsandextracting a polar krill oil
`
`from said deodorized krill material by supercritical fluid extraction with a polar entrainer to
`
`provide an essentially odorlesskrill oil.
`
`In some embodiments,
`
`the present invention provides a composition comprising krill oil
`
`containing less than about 70 micrograms/kilogram (w/w) astaxanthin esters.
`
`In some
`
`embodiments,
`
`the compositions comprise less than about 50 micrograms/kilogram (w/w)
`
`astaxanthin esters.
`
`In some embodiments, the compositions comprise less than about 20
`
`micrograms/kilogram (w/w) astaxanthin esters.
`
`In some embodiments,
`
`the compositions
`
`comprise less than about 5 micrograms/kilogram (w/w)astaxanthin esters.
`
`In some embodiments, the present invention provides a krill oil produced by the process
`
`comprising: pumping fresh krill from a trawl onto a ship, heating the krill to provide a krill
`
`material, and extracting oil from the krill material.
`
`DEFINITIONS
`
`As used herein, "phospholipid" refers to an organic compound having the following general
`
`structure:
`
`O—C—R'
`
`Il
`
`——o——C——R?
`
`O——P——O——R?®
`
`wherein R1 is a fatty acid residue, R2 is a fatty acid residue or -OH, and R3 is a —-H or
`nitrogen
`containing
`compound
`choline
`(HOCH2CHN*(CH3);0H),
`ethanolamine
`
`(HOCH2CH2NH,), inositol or serine. Rl and R2 cannot simultaneously be OH. When R3is
`
`an —OH,the compoundis a diacylglycerophosphate, while when R3 is a nitrogen-containing
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`compound, the compoundis a phosphatide such as lecithin, cephalin, phosphatidyl!serine or
`
`plasmalogen.
`
`Asusedherein, the term omega-3 fatty acid refers to polyunsaturated fatty acids that have the
`
`final double bond in the hydrocarbon chain between the third and fourth carbon atoms from
`
`the methyl end of the molecule. Non-limiting examples of omega-3 fatty acids include,
`
`5,8,11,14,17-eicosapentaenoic acid (EPA), 4,7,10,13,16,19-docosahexanoic acid (DHA) and
`
`7,10,13,16,19-docosapentanoic acid (DPA).
`
`Asused herein, astaxanthin refers to the following chemical structure:
`
` O
`
`As used herein, astaxanthin esters refer to the fatty acids esterified to OH groupin the
`
`astaxanthin molecule.
`
`DETAILED DESCRIPTION OF THE INVENTION
`
`This invention discloses novel krill oil compositions characterized by containing high levels
`
`of astaxanthin, phospholipids and omega-3 fatty acids. The krill oils compositions are
`
`extracted from krill meal using supercritical fluid extraction (SFE) with a solvent modifier.
`
`The krill meal has been processed on board a ship in Antarctica using live krill as starting
`
`material in order to ensure the highest possible quality of the krill meal. The krill oils are
`
`extracted from the krill meal in two stages, in step 1 the neutral fraction is extracted using
`
`neat supercritical CO2. The neutral fraction consisted mostly of triglycerides and cholesterol.
`
`In stage 2,
`
`the polar lipids (phospholipids) are extracted by adding 20% ethanol
`
`to the
`
`supercritical COextraction medium. The polarkrill oil extracted where found to contain 94%
`
`phospholipids (w/w), 46% omega-3 fatty acids (w/w) and 580 mg/kgastaxanthinesters.
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`The present
`
`invention provides methods to avoid decomposition of glycerides and
`
`phospholipids in krill oil and compositions produced by those methods. The product obtained
`
`by these new methodsis virtually free of enzymatically decomposed oil constituents. The
`
`solution to the problem is to incorporate a protein denaturation step on fresh krill prior to use
`
`of any extraction technology. Denaturation can be achieved by thermal stress or by other
`
`means. After denaturation, the oil can be extracted by an optional selection of nonpolar and
`polar solvents including use of supercritical carbon dioxide. Krill is adapted to a very efficient
`nutrient digestion at very low temperatures. Therefore the enzymesare sensitive to heat and
`
`the step of applying thermal! denaturation of lipases and phospholipases does not imply use of
`
`very high temperatures. Surprisingly,
`
`it has been found that the use of mild denaturation
`
`conditions can greatly enhancethe quality of krill oil.
`
`Additionally, a major obstacle of several processes of extraction is the cost of removing
`
`water. This is particularly true for methods feasible for extraction of highly unsaturated lipids
`
`where freeze drying has been regarded as the method of choice to avoid oxidative breakdown
`
`of lipids. However, the lipids in krill are surprisingly stable against oxidative deterioration.
`
`Therefore, a process including moderate use of heat in the water removing processis feasible
`provided that the enzymes have been inactivated.
`
`A.
`
`Krill Processing
`
`The present invention provides methods for processing freshly caught krill at the site of
`
`capture and preferably on board a ship. After processing on board, the krill can be further
`
`subjected to extraction processes on board the ship or at a remote location away from theship.
`
`The processing steps described herein also allow for the storage of krill material, preferably a
`
`krill meal for from about 1,2, 3, 4, 5, 6, 8, 9, 10, 11, or 12 months to about 24 to 36 months
`
`prior to processing.
`
`In some preferred embodiments, freshly caug