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`of polyunsaturated fatty acids and a person of ordinary skill in the art reading
`
`Table 4 would understand that they are included in the polyunsaturated fatty acids.
`
`59. The specification states that “Table 5 … details the lipids and other
`
`compounds (non-metal) of the extract.” ’351 Patent at col.17 l.49-col.18 l.7.
`
`Table 5 lists lipid classes as “≥ X g/100g extract.” The way Table 5 is reported
`
`indicates that the lipids are measured as g/100g of the extract, meaning that this is
`
`a quantitative measure of the lipid classes in the extract.
`
`60. Although not clear what type of analysis was conducted in the
`
`specification, the fatty acid composition (also known in the art as fatty acid profile)
`
`of total lipids is typically measured by conducting a fatty acid methyl ester
`
`(“FAME”) analysis. To do so lipids are dissolved in a suitable solvent and an acid
`
`or based catalyzed reaction or set of reactions is performed to remove all fatty acyl
`
`groups from their respective esterified positions in triacylglycerols, phospholipids,
`
`cholesterol esters and all other lipid classes, and together with native free fatty
`
`acids, they are esterified to methyl groups to yield free fatty acids. FAME
`
`mixtures are then subjected to gas liquid chromatography which produces a series
`
`of mostly separated FAME peaks corresponding to the original fatty acids in the
`
`lipid, which must then be calibrated for differential response with an external
`
`standard of containing representative FAME. Once calibrated the relative areas
`
`under the FAME peaks are proportional to their abundance in the original lipid.
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`of polyunsaturated fatty acids and a person of ordinary skill in the art reading
`
`Table 4 would understand that they are included in the polyunsaturated fatty acids.
`
`59. The specification states that “Table 5 … details the lipids and other
`
`compounds (non-metal) of the extract.” ’351 Patent at col.17 l.49-col.18 l.7.
`
`Table 5 lists lipid classes as “≥ X g/100g extract.” The way Table 5 is reported
`
`indicates that the lipids are measured as g/100g of the extract, meaning that this is
`
`a quantitative measure of the lipid classes in the extract.
`
`60. Although not clear what type of analysis was conducted in the
`
`specification, the fatty acid composition (also known in the art as fatty acid profile)
`
`of total lipids is typically measured by conducting a fatty acid methyl ester
`
`(“FAME”) analysis. To do so lipids are dissolved in a suitable solvent and an acid
`
`or based catalyzed reaction or set of reactions is performed to remove all fatty acyl
`
`groups from their respective esterified positions in triacylglycerols, phospholipids,
`
`cholesterol esters and all other lipid classes, and together with native free fatty
`
`acids, they are esterified to methyl groups to yield free fatty acids. FAME
`
`mixtures are then subjected to gas liquid chromatography which produces a series
`
`of mostly separated FAME peaks corresponding to the original fatty acids in the
`
`lipid, which must then be calibrated for differential response with an external
`
`standard of containing representative FAME. Once calibrated the relative areas
`
`under the FAME peaks are proportional to their abundance in the original lipid.
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`Areas are integrated and normalized to (divided by) the sum of all areas to yield
`
`proportions of each fatty acid in the lipid; the sum of all FAME in a sample added
`
`to 100%, and this is considered a weight-for-weight (w/w) of total fatty acids
`
`percentage. For instance, DHA might be 10% w/w of fatty acids in the mixture.
`
`Quantitative analysis is often performed as an additional measure. Normally an
`
`internal standard will be added quantitatively (on the basis of milligrams per
`
`milliliter) to the FAME mixture prior to analysis. Once responses are calibrated,
`
`the internal standard can be used to calculate the concentration of each particular
`
`fatty acid per unit weight of a sample. For instance, 100 mg of a lipid might
`
`contain 8 mg of DHA, and the concentration of DHA would be said to be 80 mg
`
`per gram lipid. These procedures both presume proper identification of the identity
`
`of fatty acids which requires separate criteria.
`
`61. Example 1 of the ‘351 Patent purports to be an analysis that
`
`“illustrates the isolation and molecular characterization of the phospholipids from
`
`the extract.” This analysis does not demonstrate a particular phospholipid
`
`molecular species exists in the sample tested. Any chemically definable class of
`
`lipids within a lipid sample can be analyzed for their fatty acid composition
`
`(profile). For instance, free fatty acids can be purified and then their fatty acid
`
`profile can be determined. When a fatty acid profile is performed on a
`
`phospholipid such as phosphatidylcholine the profile does not distinguish between
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`fatty acids on the sn-1 and sn-2 positions; this can be accomplished using
`
`additional analysis steps, such as phospholipase A2 to selectively cleave the sn-2
`
`position with subsequent separation and profiling of the free fatty acids and 1-
`
`monoglycerols. In special cases the results of a fatty acid analysis of
`
`phosphatidylcholine can be used to unambiguously drawn chemical conclusions
`
`about fatty acids bound to the sn-1 and sn-2 positions. For instance, if DHA was
`
`found to be between 51 and 100% of the fatty acids on a phosphatidylcholine, then
`
`at least some phosphatidylcholine molecules must have DHA on both positions.
`
`62.
`
`It is not clear from the specification whether the same samples were
`
`tested in Tables 3, 4, and 5, or in any other Tables in the ‘351 Patent. Nor is there
`
`any information about how these samples were prepared.
`
`63. The specification identifies an extraction method that is “similar to the
`
`one described in commonly owned PCT publication number WO 00/23546”
`
`(“Beaudoin I”; Ex. 1002), which is incorporated by reference. (’351 Patent
`
`col.18.) No variation of this extraction procedure is mentioned or described. The
`
`table below compares the extraction process in Beaudoin I and the ‘351 Patent.
`
`The methods are virtually identical.
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`Beaudoin I
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`‘351 Patent (’351 Patent col.18
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`Beaudoin ’299 patent
`
`l.32-col.19 l.9)
`
`The starting material consisting
`
`Preferably, freshly harvested and
`
`of freshly harvested and preferably
`
`finely divided marine and aquatic
`
`finely divided marine and aquatic
`
`animal material is subjected to acetone
`
`animal material is subjected to acetone
`
`extraction, for at least about two hours
`
`extraction, for at about two hours and
`
`and preferably overnight. Col. 18, l. 53-
`
`preferably overnight. p. 5, l. 21-25.
`
`55.
`
`However extraction time is not
`
`However, extraction time is not
`
`critical to the yield of lipid extraction.
`
`critical to the yield of lipid extracted.
`
`To facilitate extraction, it is preferable
`
`Particle sizes of comminuted crustacean
`
`to use particles of less than 5mm in
`
`less than 5 mm are preferred. The
`
`diameter. Extraction is preferably
`
`extraction is preferably conducted under
`
`conducted under inert atmosphere and at
`
`an inert atmosphere and at a temperature
`
`a temperature in the order of about 5°C
`
`of about 5 degrees Celsius or less. Col.
`
`or less. p. 5, l. 25-29.
`
`18, l. 55-60.
`
`Preferably, the beginning of the
`
`The mixture may be agitated
`
`
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`Beaudoin I
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`‘351 Patent (’351 Patent col.18
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`Beaudoin ’299 patent
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`l.32-col.19 l.9)
`
`extraction will be conducted under
`
`during extraction and a volume ratio of
`
`agitation for about 10 to 40 minutes,
`
`about 6:1 of acetone to biomass is
`
`preferably 20 minutes. Although
`
`generally most preferred. Col. 18, l. 60-
`
`extraction time is not critical, it was
`
`62.
`
`found that a 2 hour extraction with 6:1
`
`volume ratio of acetone to marine and
`
`aquatic animal material is best. p. 6, l.
`
`30 – p. 6, l. 2.
`
`The solubilized lipid fractions are
`
`The solubilized lipid fraction is
`
`separated from the solid material by
`
`separated from the solid starting
`
`standard techniques including, for
`
`material by known techniques, for
`
`example, filtration, centrifugation or
`
`example, by filtration, centrifugation or
`
`sedimentation. Filtration is preferably
`
`sedimentation. Filtration is preferred.
`
`used. p. 6, l. 6-7.
`
`Col. 18, l. 63-66.
`
`After separation by filtration on
`
`The residue is optionally washed
`
`
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`Beaudoin I
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`‘351 Patent (’351 Patent col.18
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`Beaudoin ’299 patent
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`l.32-col.19 l.9)
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`an organic solvent resistant filter (metal,
`
`with acetone to recover more lipid and
`
`glass or paper) the residue is optionally
`
`the acetone removed by flash
`
`washed with pure acetone, preferably
`
`evaporation or spray drying. Water
`
`two volumes 10 (original volume of
`
`residue is allowed to separate from the
`
`material) to recover yet more lipids. The
`
`lipid extract at low temperature. Col. 18,
`
`combined filtrates are evaporated under
`
`l. 66 – Col. 19., l. 2.
`
`reduced pressure. Optionally, flash
`
`evaporation or spray drying may be
`
`used. The water residue obtained after
`
`evaporation is allowed to separate from
`
`the oil phase (fraction I) at low
`
`temperature. p. 6, l. 8-14.
`
`The solid residue collected on the
`
`The solid residue left on the filter
`
`filter is suspended and extracted with
`
`from the initial extraction is suspended
`
`alcohol, such as ethanol, isopropanol, t-
`
`and extracted with 95/5 ethyl
`
`butanol or alternatively with ethyl
`
`acetate/ethanol, preferably two volumes
`
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`Beaudoin I
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`Beaudoin ’299 patent
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`l.32-col.19 l.9)
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`acetate, preferably two volumes
`
`(original volume of material). Col. 19, l.
`
`(original volume of material). p. 6, l. 15-
`
`3-5.
`
`17.
`
`The filtrate is evaporated leaving
`
`The filtrate is evaporated yielding
`
`a second fraction of lipids (identified as
`
`a second fraction of lipids. Extraction
`
`fraction II). Although the extraction
`
`period is not critical although it is
`
`period is not critical, it was found that
`
`preferred to extract for about 30 minutes
`
`an extraction time of about 30 minutes
`
`at a temperature below about 5 degrees
`
`is sufficient at temperatures below about
`
`Celsius. Col. 19, l. 5-9.
`
`5°C. p. 6, l. 17-20.
`
`
`
`64. Example 2 of the ‘351 Patent is a study with the objective “[t]o
`
`evaluate the photoprotective potential of krill extract against UVB-induced skin
`
`cancer.” Mice were treated with “krill extract” or placebo soy extract, either
`
`orally, topically, or both. (‘351 Patent at col.24, l.48 - col.5 l.67.) The mice were
`
`then exposed to UVB radiation and monitored for malignant tumors. (Col. 25, ll.
`
`
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`41-51.) The mice receiving krill extract had a lower frequency of malignant
`
`tumors than those receiving soy. (Col. 25 at Table 11.) The ‘351 Patent does not
`
`say anything about the composition of the krill extract used or how it was made.
`
`65.
`
`In Example 3, a 7 year old girl was reportedly treated with 2 grams
`
`per day of “the present krill extract.” (Col. 26, ll. 3-8.). The ‘351 Patent report
`
`that after 1.5 months, the girl showed increased learning ability, improved motor
`
`function. (Col. 26, ll. 7-13.). The ‘351 Patent do not say anything about the
`
`composition of the krill extract used or how it was made.
`
`2.
`
`DETAILED DESCRIPTION OF PRIOR ART REFERENCES
`
`2.1 Beaudoin I (Ex. 1002)
`
`2.1.1 Beaudoin I (the reference)
`
`66. WO 00/23546 (“Beaudoin I”; Ex. 1002), “Method of Extracting
`
`Lipids from Marine and Aquatic Animal Tissues,” is directed to krill extracts, a
`
`process for producing them, and the beneficial uses of the extracts. It lists Adrien
`
`Beaudoin and Genevieve Martin as inventors.
`
`67. Beaudoin I published on April 27, 2000. Therefore I understand that
`
`it is prior art under at least 35 U.S.C. § 102(b) even under Patentee’ earliest alleged
`
`priority date. This reference was cited by the Examiner during prosecution of the
`
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`‘351 Patent and the parent ‘348 Patent. It is also cited in the specification of the
`
`‘351 Patent. (’351 Patent at 1:43-48)
`
`68. Beaudoin I discloses that lipid fractions from marine and aquatic
`
`animals such as krill have a variety of applications in the medical, nutraceutical,
`
`and cosmetics fields, and are useful as dietary supplements. (Ex. 1002 at pp.1-2.)
`
`Beaudoin I cites marine oils for their beneficial anti-inflammatory properties, their
`
`utility in cardiovascular disease, lupus, and renal diseases, and their uses as dietary
`
`supplements due to the beneficial effects of omega-3 fatty acids in krill oil. (Id.)
`
`Beaudoin I notes that krill contain high concentrations of EPA and DHA. (Id. at
`
`p.2.)
`
`69. Beaudoin I discusses that various methods of extracting oils from
`
`marine and aquatic animals were known. For example, he cites the classical
`
`technique of Folch et al., published in 1957 in J. Biol. Chem. 226:497-509, “A
`
`simple method for the isolation and purification of total lipids from animal
`
`tissues.” (Id. at p.3.) He notes that the method is not commercially feasible
`
`because of the toxicity of the solvents (chloroform and methanol). (Id.) He also
`
`notes that another prior art extraction method, described in U.S. Patent 4,331,695
`
`to Zosel, is not preferred because hexane is a poor extraction solvent for krill and
`
`because the process used high temperatures which negatively altered the lipids that
`
`were precipitated. (Id. at p. 2.)
`
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`70. Beaudoin I teaches an alternate method for extracting lipid fractions
`
`
`
`from marine and aquatic animal material by using a ketone such as acetone,
`
`followed by subjecting the non-soluble and particulate fraction to additional
`
`solvent extraction with an alcohol such as ethanol, isopropanol, t-butanol, or an
`
`ester of acetic acid, such as ethyl acetate. (Id., at Abstract.) More specifically,
`
`Beaudoin I presents the following process:
`
`•
`
`Finely divided marine and aquatic animal material is subjected to
`
`acetone extraction, preferably at low temperature: “Extraction is preferably
`
`conducted under inert atmosphere and at a temperature on the order of about 5°C
`
`or less.” (Id. at p. 5.) The extraction is preferably carried out for about two hours
`
`with a 6:1 volume ratio of acetone to marine and aquatic material. (Id. at pp.5-6.)
`
`•
`
`The solubilized lipid fractions are separated from the solid material by
`
`standard techniques, such as filtration. (Id. at p.6.) The residue is optionally
`
`washed with pure acetone, preferably two volumes, to recover yet more lipids.
`
`(Id.) The combined filtrates are evaporated under reduced pressure, and the water
`
`residue obtained after evaporation is allowed to separate from the oil phase
`
`(identified as fraction I) at low temperature. (Id.)
`
`•
`
`The solid residue collected on the filter is suspended and extracted
`
`with alcohol (such as ethanol, isopropanol, t-butanol or alternatively with ethyl
`
`
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`acetate, preferably two volumes. (Id.) The filtrate is evaporated, leaving a second
`
`fraction of lipids (identified as fraction II). (Id.) Beaudoin I again notes that
`
`extraction is carried out at low temperature: “Although the extraction period is not
`
`critical, it was found that an extraction time of about 30 minutes is sufficient at
`
`temperatures below about 5°C.” (Id.)
`
`71. Beaudoin I states: “Temperature of the organic solvents, except t-
`
`butanol, and the temperature of the sample are not critical parameters, but it is
`
`preferable to be as cold as possible. However, in the case of t-butanol which is
`
`solid at room temperature, it is important to warm it before using it and to perform
`
`the extraction at 25°C immediately.” (Id.)
`
`72. Beaudoin I used this extraction process with krill, and analyzed the
`
`resulting extracts: “Results on krill oils obtained in accordance with the method of
`
`the present invention (fraction I extracted with acetone and fraction II extracted
`
`with ethyl acetate) are provided in Tables 13, 14, 15, 16, 17 and 18.” (Id. at p.
`
`10.) Table 14 details the lipid class composition of krill oil from E. pacifica. (Id.
`
`at p. 23.) It shows the area percentages of free fatty acids in fractions I and II (23.7
`
`and 20.3, respectively), as well as the amounts of phospholipids or other polar
`
`materials in these fractions (54.1 and 8.5, respectively). (Id.) In these samples,
`
`Fraction I was made by a sample-acetone ratio of 1:6 (w/v), which was incubated 2
`
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`hours at 4°C. Fraction II was made with a sample-ethyl acetate ratio of 1:2 (w/v),
`
`which was incubated 30 minutes at 4°C, following a first extraction with acetone.
`
`(Id.)
`
`73. Tables 15 and 16 show the fatty acid composition of krill oil (E.
`
`pacifica). Both Fractions I and II show the presence of EPA (20:5 n-3) and DHA
`
`(22:6 n-3). (Id. at pp. 23-26.) In Table 15, EPA is present at 17.4% and 8.6%
`
`(wt/wt%) in fractions I and II, respectively, and DHA is present at 13.2% (fraction
`
`I) and 6.6% (fraction II). (Id. at pp. 23-25.) In Table 16, EPA is present at 26.4%
`
`(fraction I) and 17.4% (fraction II), and DHA is present at 20.5% (fraction I) and
`
`14.4% (fraction II). (Id. at pp. 25-26.)
`
`74. Table 18 shows the amount of astaxanthin in E. pacifica fractions.
`
`(Id. at p. 27.) Fraction I had 93.1 µg/g oil, and Fraction II had 121.7 µg/g. (Id.)
`
`75. Beaudoin I discloses that “Table 19 shows the best mode of the
`
`method in accordance with the present invention for lipid extraction of aquatic
`
`animal tissues.” (Id. at p. 11.) Table 19 is reproduced below:
`
`
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`(Id. at p. 28.)
`
`
`
`76. Beaudoin I teaches that krill oil extracts made according to the
`
`disclosed process contain high proportions of the omega-3 fatty acids EPA (20:5)
`
`and DHA (22:6): “Figures 1 to 6 show chromatograms of fatty acid composition
`
`of E. pacifica lipids. On each of them, high proportions of 20:5 and 22:6 fatty
`
`acids (characteristic of marine and aquatic oils) are noticeable and represented by
`
`two distinct peaks. Data are shown in Table 12.” (Id. at p. 11.)
`
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`77. Beaudoin I notes: “Table 13 shows also that fraction I is comprised of
`
`
`
`10,0% of volatile matter and humidity after evaporation of the solvent. For the
`
`same test, the fraction II gives a value of 6,8%. To get rid of traces of solvents, it
`
`is important to briefly heat (to about 125°C, for about 15 min) the oil under
`
`nitrogen.” (Id. at p. 10.) (“To get rid of traces of organic solvents, lipid fractions I
`
`and II are warmed to about 125°C for about 15 minutes under inert
`
`atmosphere.”).)2
`
`78. Beaudoin I explains that extracts have various uses including at least
`
`medical application, nutraceutical, and cosmetics. Beaudoin I also notes that the
`
`extracts were consumed by a human: “[o]ne of the inventors, Dr. Adrien
`
`Beaudoin, has ingested the different lipid fractions of krill. No side effect profile
`
`was observed.” (Id. at p. 12.)
`
`79. Claim 1 is directed to seven-step extraction method, as follows:
`
` 1. A method for extracting lipid fractions from marine and
`
`aquatic animal material, said method comprising the steps of:
`
`(a)
`
`placing marine and aquatic material in a ketone solvent,
`
`preferably acetone to achieve extraction of the soluble lipid fraction
`
`from said marine and aquatic animal material;
`
`2
`As will be discussed herein, Patentee have asserted that these heating steps are part of the Beaudoin
`extraction process. I disagree. (See infra Section 11.2.1.1 and Exhibit A.1.)
`
`
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`(b)
`
`separating the liquid and solid contents;
`
`
`
`(c)
`
`recovering a first lipid rich fraction from the liquid
`
`contents by evaporation of the solvent present in the liquid contents;
`
`(d)
`
`placing said solid contents in an organic solvent selected
`
`from the group of solvents consisting of alcohol, preferably ethanol,
`
`isopropanol or t-butanol and esters of acetic acid, preferably ethyl
`
`acetate to achieve extraction of the remaining soluble liquid fraction
`
`from said marine and aquatic animal material;
`
`(e)
`
`separating the liquid and solid contents;
`
`(f)
`
`recovering a second lipid rich fraction by evaporation of
`
`the solvent from the liquid contents;
`
`(g)
`
`recovering the solid contents.
`
`(Id. at p. 29.)
`
`80. Beaudoin I notes several advantages of his extraction process and the
`
`resulting krill extracts. First, his process results in translucent oils that are more
`
`attractive than those obtained by the Folch procedure. (Id. at p. 7.) His process
`
`also uses less toxic solvents, and achieves higher levels of lipids extracted from dry
`
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`krill, as compared to the Folch procedure. (Id. at p. 8.) He also notes that his
`
`extraction process removes bacteria from the extracts. (Id.)
`
`2.1.2 Beaudoin (testing)
`
`81.
`
`I have reviewed the Expert Witness Report of Suzanne Budge, Ph.D
`
`(“Budge Declaration,” Ex. 1041). As described in her report, Dr. Budge extracted
`
`samples of both Euphausia pacifica and Euphausia superba krill according to the
`
`extraction method disclosed in Beaudoin I and II. I have reviewed Beaudoin I and
`
`II and Dr. Budge’s report, and in my opinion Dr. Budge properly performed the
`
`extractions in accordance with Beaudoin I and II.
`
`82. Dr. Richard van Breemen analyzed the samples prepared by Dr.
`
`Budge for the presence of phosphatidylcholine (“PC”) containing two EPA
`
`molecules; PC containing two DHA molecules; and PC containing an EPA
`
`molecule and a DHA molecule. I have also reviewed the Declaration of Richard
`
`van Breemen, Ph.D (Ex. 1040). I believe that Dr. van Breemen conducted these
`
`tests properly and that he did, in fact, confirm the presence of each of these
`
`phospholipids in each of the samples tested.
`
`83. Chemir analyzed the samples for the presence of fatty acids, metals,
`
`and astaxanthin. I spoke with Chemir prior to the testing to confirm that they
`
`would be conducting the appropriate analyses. I also reviewed the Expert Witness
`
`
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`Report of Albert C. Lee, Ph.D. and confirmed that they conducted the analyses in
`
`accordance with the methods described in Dr. Lee’s report. (“Lee Declaration,”
`
`Ex. 1045)
`
`84. Avanti Polar Lipids (“Avanti”) analyzed the samples for total lipids3
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`and phospholipids. I spoke with Avanti prior to the testing to confirm that they
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`would be conducting the appropriate analyses. I also reviewed the Expert Witness
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`Report of Jeff D. Moore, Ph.D. and confirmed that they conducted the analyses in
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`accordance with the methods described in Dr. Moore’s report.4 (“Moore
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`Declaration,” Ex. 1044).
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`85. The Beaudoin I and Beaudoin II (Ex. 1003) extractions have also been
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`performed no fewer than four additional times, and each time the resulting extract
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`was tested for the presence of phosphatidylcholine (“PC”) containing two EPA
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`molecules; PC containing two DHA molecules; and PC containing an EPA
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`molecule and a DHA molecule. At Neptune’s request, Dr. Earl L. White analyzed
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`two different Beaudoin extracts. In addition, Bjorn Ole Haugsgjerd has prepared
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`two different Beaudoin extracts (Ex. 1047 and 1048), which were tested by Drs.
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`3 I note that one set of E. superba samples were not tested for total lipids
`4 Having reviewed the test result provided by Avanti and Chemir with respect to the three sets of Beaudoin samples
`(one E. Pacifica and two E. superba, each at three temperatures), I observed that an entire set of the E. superba
`samples (BEA-S0, BEA-S1, BEA-S2) are anomalous. Lipids were found in the BEA-S0, BEA-S1, BEA-S2 E.
`superba samples at each temperature, but phospholipids were only detected at less than 0.5%. These outlier results
`are not considered within the realm of possible lipid extraction results using freshly frozen E. superba and therefore
`excluded from further analysis.
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`Thomas Gundersen (Ex. 1049 and 1050) and Richard B. Van Breemen (Ex. 1066).
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`In each of these samples, the Claimed Phospholipids were found.
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`2.2 Beaudoin II (Ex. 1003)
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`2.2.1 Beaudoin II (reference)
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`86. A second Beaudoin reference is also directed to krill extracts, a
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`process for producing them, and the beneficial uses of the extracts. CA 2,251,265
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`(“Beaudoin II,”), entitled “Process for Lipid Extraction of Aquatic Animal Tissues
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`Producing a Dehydrated Residue,” lists Adrien Beaudoin and Genevieve Martin as
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`inventors.
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`87.
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`I understand that Beaudoin II is a Canadian patent application that
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`was published on April 21, 2000, but which was filed in Canada on October 1998.
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`Therefore I understand that Beaudoin II is prior art under at least 35 U.S.C.
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`§ 102(b) even under Patentee’ earliest alleged priority date. Beaudoin II is cited on
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`the face of the ’351 Patent.
`
`88. Beaudoin II describes a method for extracting lipid fractions from
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`krill. (Beaudoin II at p. 2.) First, fresh or frozen krill is suspended in cold acetone
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`at low temperature (5°C or lower). (Id. at p. 2.) A ratio of krill-acetone of 1:6
`
`(w/v) and an incubation time of 2 hours is preferred, and it is preferable to grind
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`the material to ~5 mm before contacting it with acetone. (Id.)
`
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`89. After filtration, the residue is washed with two volumes of pure
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`
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`acetone. (Id.) The combined filtrates are evaporated under reduced pressure. (Id.)
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`The water residue obtained after evaporation is allowed to separate from the oil
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`phase (identified as fraction I) at low temperature. (Id.) The solid residue
`
`collected on the filter is suspended and extracted with two volumes of 100%
`
`ethanol. (Id.) The ethanol filtrate is evaporated leaving a second fraction of lipids
`
`(identified as fraction II). (Id.) Beaudoin II notes that the temperature of the
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`organic solvents and sample are not critical parameters, but that they are preferably
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`as cold as possible. (Id.)
`
`90.
`
` Beaudoin II teaches that oil and other fractions from aquatic animal
`
`biomass (including krill biomass) have a variety of uses. He recognizes the high
`
`levels of the beneficial fatty acids EPA and DHA in krill, which are useful as
`
`dietary supplements for humans. (Id. at p.4.) Beaudoin II also notes that these
`
`marine products are rich in vitamins A, D, E, and K and carotenoids (which
`
`include astaxanthin), and that these compounds are extracted with lipids. (Id. at p.
`
`5.)
`
`91. Beaudoin II discusses the levels of lipids extracted by his method as
`
`compared to the classical procedure of Folch. (Id. at p. 6.) He also states that his
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`method preserves the krill sample such that there is no bacterial growth. (Id.) He
`
`also discloses the fatty acid composition of E. pacifica lipids, including high
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`proportions of DHA and EPA, which he notes are characteristic of marine oils.
`
`(Id. at p. 7 and Figs. 1-4.)
`
`92. Beaudoin II also discloses that his krill extracts were consumed by a
`
`human: “One of the inventors, Mr. Adrien Beaudoin, has tasted the different lipid
`
`fractions. No side effect was observed. The fraction I has the taste of the cod liver
`
`oil and the insoluble material tastes like salty shrimps.” (Id. at p. 8.)
`
`93. Diagram 1 shows a krill lipid extraction process of Beaudoin II, which
`
`starts with 1,000 kg fresh krill, and results in 40 kg of krill oil:
`
`
`
`(Id. at p. 9.)
`
`
`
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`94. Table 11 provides the “Optimal conditions for lipid extraction of
`
`
`
`aquatic animal tissues (suggested procedure)”:
`
`
`
`
`
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`(Id. at p. 20.)
`
`
`
`2.2.2 Beaudoin II (testing)
`
`95. Dr. Budge prepared extracts of E. superba and E. pacifica according
`
`to the methods in Beaudoin I and Beaudoin II and these extracts where then tested
`
`by Dr. van Breemen, Chemir, and Avanti. I explain the results of this testing in
`
`detail below.
`
`2.3 Neptune Prospectus (Ex. 1011)
`
`96.
`
`I also reviewed a Neptune document, entitled “Final Prospectus” (the
`
`“Neptune Prospectus,” Ex. 1011), dated May 11, 2001. The Neptune Prospectus
`
`seems to have issued in connection with an initial public offering. I understand
`
`that based on a priority date of July 29, 2002 or later, this reference is prior art
`
`under at least 35 U.S.C. § 102(b).5 This reference was not of record during the
`
`prosecutions of the ‘348, or ’351 Patents.
`
`97.
`
`In the Neptune Prospectus, Neptune described its mission as
`
`“developing, marketing and exploiting
`
`innovative high-yield
`
`technological
`
`processes in the production of value-added natural product, such as Omega-3 type
`
`oils.” (Neptune Prospectus at p. 10.) Neptune targeted krill as a source for omega-
`
`5
`I understand that even if Neptune were accorded the benefit of the July 27, 2001 filing date of the
`Provisional Application, the Neptune Prospectus would still be prior art, under 35 U.S.C. § 102(a). I understand that
`Neptune would then have an opportunity to eliminate this reference as prior art by proving an invention date earlier
`than the date of the reference.
`
`
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`3 rich oil because its lipid content is a major source of polyunsaturated fatty acids,
`
`mainly docosahexanoic acid (DHA) and eicosapentanoic acid (EPA), both of
`
`which are types of omega-3 fatty acids. (Id. at p. 11) Krill also contains powerful
`
`antioxidants such as astaxanthin. (Id.) The prospectus notes that krill is a good
`
`target because it is “abundant, accessible, and there is a potential for long-term
`
`exploitation.” (Id.)
`
`98. The Neptune Prospectus points out that polyunsaturated omega-3 fatty
`
`acids are beneficial for patients with cardiovascular diseases, arthritis, asthma,
`
`inflammatory conditions associated with immune system diseases, and neoplastic
`
`diseases. (Id.)
`
`99. The Neptune Prospectus describes its krill oil as containing “powerful
`
`natural carotenoids (vitamin A, beta-carotene, trans-retinols, astaxanthin and
`
`canthaxantin) and a liposoluble vitamin (vitamin E), which have numerous known
`
`antioxidant properties. . . . The Company’s krill oil concentrate is of extremely
`
`high quality as it does not oxidize during the Neptune OceanExtract™ process . . .
`
`which increases its biological effects and absorption by the human body.” (Id. at
`
`p. 12.)
`
`100. The Neptune Prospectus further states that its extraction process,
`
`when performed on krill, uses no heat at the raw material processing stage. (Id. at
`
`p. 13.) It defines the process as a “cold extraction process that enables the
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`
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`extraction of Omega-3 polyunsaturated oil, protein concentrates and amino acid
`
`concentrates from marine biomasses such as krill, calanus and other crustaceans.
`
`Krill oil is extracted from the raw material, i.e. the krill, by successive immersions
`
`in organic solutions and through filtrations.” (Id.) The Neptune Prospectus refers
`
`to this as the OceanExtract™ process, and asserts that it “destroys bacteria,
`
`resulting in products that are safe and healthy for human consumption. . . .” (Id.)
`
`101. According to the Neptune Prospectus, the extraction process was
`
`developed in 1998 and 1999 in the Biology Department of the Faculty of Science
`
`at Université de Sherbrooke. (Id.) The Neptune Prospectus notes that the
`
`Université filed two patent applications relating to lipid extraction, CA 2,251,265
`
`and PCT/CA/99/00987.6 (Id. at p. 15) According to the Neptune Prospectus, the
`
`PCT application
`
`(i.e., Beaudoin
`
`I, Ex. 1002) “includes
`
`the Neptune
`
`OceanExtract™ extraction process” described in the Neptune Prospectus. (Id.)
`
`Dr. Adrien Beaudoin (one of the inventors on Beaudoin I and II), was on the
`
`Neptune research advisory board. (Id. at p. 15.)
`
`2.4 Neptune Press Release (Ex. 1012)
`
`102. On June 14, 2001, Neptune published a press release titled “Neptune
`
`Technologies & Bioressources Soon to Obtain a Major Patent in Over 30
`
`
`6
`These are Beaudoin II and Beaudoin I, respectively.
`
`
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`
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`Countries” (“Neptune Press Release,” Ex. 1012). The patent application that was
`
`the subject of
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