ORIGINAL SUBMISSION
`
`RIMFROST EXHIBIT 1075 page 0001
`
`

`

`+
`.. '2 ,.j-
`.-
`
`I
`
`r
`
`.<
`
`Geheraify Recognized AS Safe
`_. 2,0482 Jacklight Lahe, bend, OR.,97702-3074
`- ..
`_.* < .
`541 638-y2
`. .
`.
`.
`.
`
`m c q u a t e e g r a s -a .. s go Q i at e s . co m . 1 ww w . g r a s - a s s o c i a t e s :c o m
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`
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`
`0 1 - 2 4 - 0 8 A l l : 3 5 O U T
`
`~
`
`January 18,2008
`
`Food and Drug Administration
`Center for Food Safety & Applied Nutrition
`Office of Food Additive Safety (HFS-200)
`51 00 Paint Branch Parkway
`College Park, MD 20740-3835
`
`Attention: Dr. Robert L. Martin
`
`Dear Dr. Martin:
`
`On behalf of Neptune Technologies & Bioressources of Lava1 (Quebec), Canada, we are submitting for FDA review a
`GRAS notification for High Phospholipid Krill Oil. The attached documentation contains the specific information that
`addresses the safe human food uses for the subject notified substance as discussed in the GRAS guidance document
`As part of the deliberations by our Expert Panel on the safety of the subject krill oil as a human food ingredient, two
`unpublished scientific studies were considered. They have been included as Appendix G in the submission. Even
`though each of the reports is identified as being confidential, we recognize that the content submitted cannot be held as
`confidential, and we hereby waive the designation of confidentiality.
`
`If additional information or clarification is needed as you and your colleagues proceed with the review, please feel free
`to contact me via telephone or email.
`
`We look forward to your feedback.
`
`Sincerely,
`
`Robert S, McQuate, Ph.D.
`CEO & Co-Founder
`GRAS Associates, LLC
`20482 Jacklight Lane
`Bend, OR 97702-3074
`541 -678-5522
`mcauate@was-assoiates.com
`www.gras-associates.com
`Enclosure: GRAS Notification - High Phospholipid Krill Oil (in triplicate)
`
`o o o o b ~
`
`Page I of 1
`
`RIMFROST EXHIBIT 1075 page 0002
`
`GRAS ASSOCIATES, LLC
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page I
`
`1. GRAS EXEMPTION CLAIM
`
`A. Claim of Exemption From the Requirement for Premarket Approval Pursuant to
`Proposed 21 CFR 170.36(~)(1)’
`
`High phospholipid krill oil, meeting the specifications described below, has been determined to
`be Generally Recognized As Safe (GRAS), in accordance with Section 201 (s) of the Federal
`Food, Drug, and Cosmetic Act. This determination was made by experts qualified by scientific
`training and experience; it is based on scientific procedures as described in the following
`sections; and the evaluation accurately reflects the conditions of the ingredient’s intended use
`in foods.
`
`01-24-08 A I l : 3 5 O U T
`
`Signed:
`
`Robert S. McQuate, Ph.D.
`GRAS Associates, LLC
`20482 Jacklight Lane
`Bend, OR 97702-3074
`
`Date
`
`B. Name and Address of Notifier
`
`Neptune Technologies and Bioressources
`2740 Pierre Peladeau Avenue, Suite H200
`Lava1 (Quebec)
`Canada H7T 3B3
`
`As the not if i e r , Ne p t u ne Techno I og i e s and B i o ress o u rces (‘I N e p t u ne”) accepts res p o n s i b i I it y
`for the GRAS determination that has been made for high phospholipid krill oil as described in
`the subject notification; consequently, high phospholipid krill oil meeting the conditions
`described herein is exempt from pre-market approval requirements for food ingredients.
`
`C. Common Name and Identity of the Notified Substance
`High phospholipid krill oil; also see Sections 1I.A and 1I.B.
`
`1 See 62 FR 18938 (1 7 April 1997).
`
`GRWS ~
`
`~
`
`LLC
`~
`
`~
`
`C
`
`~
`
`~
`
`~
`
`~
`
`$
`
`,
`
`
`
`RIMFROST EXHIBIT 1075 page 0003
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page 2
`
`D. Conditions of Intended Use in Food
`
`High phospholipid krill oil is intended to be added as a substitute or alternative to fish oils to
`the following food categories at per serving levels ranging from 150 - 500 mg: non-alcoholic
`beverages; breakfast cereals; cheeses; frozen dairy desserts; milk products; processed
`fruitdfruit juices; and medical foods. The maximum daily consumption of the subject krill oil
`will provide up to 2.2 g/person/day of EPA and DHA.
`
`E. Basis for the GRAS Determination
`
`Pursuant to 21 CFR § 170.30, high phospholipid krill oil has been determined to be GRAS on
`the basis of scientific procedures as discussed in the detailed description provided below.
`
`F. Availability of Information
`
`The data and information that serve as the basis for this GRAS Notification will be sent to the
`US Food and Drug Administration (FDA) upon request or will be available for review and
`copying at reasonable times at the offices of GRAS Associates, LLC, located at 20482
`Jacklight Lane, Bend, OR 97702-3074.
`
`RIMFROST EXHIBIT 1075 page 0004
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page 3
`
`II. CHEMISTRY AND MANUFACTURE OF NKOTM
`
`NKOTM is refined krill oil which is derived from the marine organism, Euphasia superba, which
`is harvested from Antarctic waters. Krill is recognized as a vital part of the marine food chain
`for baleen whales, mantas, whale sharks, crabeater seals and other seals, and a few seabird
`species.
`
`The following description of krill is given on Neptune’s website.*
`
`“Krill is a generic term used to designate all deepwater marine planktonic crustacean species. It looks
`like miniature shrimp, ranging between one and six centimeters in length. These 85 species represent
`the planet‘s most abundant animal biomass found in banks that may extend over several square
`kilometers. Krill fisheries can mainly be found in two ocean regions: the Antarctic Ocean and the North
`Pacific Ocean, along the Canadian and Japanese coasts. According to a World Health Organization
`estimate, the total quantity of krill in these two oceans exceeds 500 million tons. Approximately 1 10,000
`tons of krill are harvested every year, or about less than one-half of 0.1 % of the existing resources.”
`
`NKOTM is a whole lipid extract and is an opaque reddish oil with a seafood odor. The major
`components of NKOTM are triglycerides and phospholipids which are rich in eicosapentaenoic
`acid (EPA, C20:5 n-3 fatty acid) and docosahexaenoic acid (DHA, C22:6 n-3 fatty acid).
`NKOTM is an oil intended for use as a dietary ingredient as a source of omega-3 fatty acids,
`which as noted are found in NKOTM in their phospholipid form. No processing aids or
`additives are included in the final NKOTM product due to naturally occurring antioxidants that
`aid in NKOTM’s preservation. Additionally, no proprietary or coloring ingredients are added to
`the oil from Antarctic krill to produce NKOTM.
`
`A. Common or Usual Name of the Subject Material
`
`High phospholipid krill oil is the common or usual name of the material that is the subject of
`the GRAS evaluation. The specific substance evaluated is Neptune Krill OilTM, which is
`commonly referred to throughout this evaluation as NKOTM. As noted in the preceding
`paragraph, NKOTM is a mixture of triglycerides and phospholipids that contain numerous
`constituent fatty acids. The compositional character of NKOTM is described in detail in this
`Section.
`
`B. Manufacturing and Specifications for NKOTM
`
`1. Krill Source and Handling
`
`NKOTM is extracted from Euphasia superba. The current taxonomic placement of
`Euphasia superba is summarized below:
`
`2 See www.neptunebiotech.com/whatiskrill. html.
`
`GRA$ ~,~~~~~~~~~~ l.iC
`
`000005
`RIMFROST EXHIBIT 1075 page 0005
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page 4
`
`is produced from Antarctic krill, which are native to the Atlantic section of the
`NKO"
`Austral-Antarctic Circumpolar Ocean. In terms of their phylogeny, Antarctic krill are
`closely related to shrimp and are consumed as human food in a similar fashion. The
`Antarctic krill used in the production of NKO" are a non-genetically modified source,
`are fished from the wild, and are not the result of breeding. No toxic constituents are
`known to be present in NKO".
`
`The Antarctic krill used to extract NKOTM are fished in the Atlantic section of the Austral-
`Antarctic Circumpolar Ocean, originating specifically from Statistical Fishing Area 48, a
`fishing area designated by the Commission for the Conservation of Antarctic Marine
`Living Resources (CCAMLR). The Antarctic krill processed by Neptune are supplied by
`a Japanese firm that has been fishing Antarctic krill for more than two decades. The
`Antarctic krill are drained and placed in plastic molds for quick-freezing at about -30°C
`on board factory ships within a few hours of harvest. The resulting 12 kg blocks of
`Antarctic krill are individually wrapped in plastic film and are packaged two per carton
`prior to export to Canada; the packaged Antarctic krill are maintained at temperatures
`of -30 to -2OOC. Upon arrival in Canada, they are inspected and approved by the
`Canadian Food Inspection Agency (CFIA). Source manipulations on board the fishing
`vessels are made in compliance with regulations as they apply to the handling of krill
`intended for human consumption.
`
`2. NKO" Manufactwing Process
`
`The manufacturing of NKO" begins with the crushing of deep frozen Antarctic krill at a
`temperature of 4°C until the Antarctic krill particles are <5 mm in size. The lipids and
`proteins are separated and extracted from the crushed Antarctic krill via the addition of
`acetone under cold extractions conditions for a period of 2 hours. This step also takes
`place at a temperature of 4"C, and requires the addition of enough acetone to ensure a
`ratio of crushed Antarctic krill to acetone of 1 :6 (w/v). Filtration occurs through an
`organic solvent-resistant filter under reduced pressure to enable physical separation of
`the Antarctic krill lipids and proteins. The remaining crushed Antarctic krill is then
`subjected to a second round of extraction to ensure that all possible lipids and proteins
`are extracted. Following extraction, the Antarctic krill proteins and lipids are filtered and
`separated from each other with this process yielding an oil. Excess acetone is then
`evaporated and separated from any water remaining in the product. The oil undergoes
`filtration and purification to remove impurities and is then packaged in a modified NZ-
`containing atmosphere and stored as NKO". Figure 1 summarizing the overall NKO"
`manufacturing process.
`
`RIMFROST EXHIBIT 1075 page 0006
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page 5
`
`Figure 1. Manufacturing Process for the Production of r..tOTr
`
`~ Acetone
`
`I
`
`I Deep Frozen Raw Krill
`
`I
`
`Crushing of the Frozen Raw Krill
`
`Cold Extraction of Krill
`Lipids and Proteins
`
`Filtration of Krill Lipids
`and Proteins
`
`I
`
`I
`
`Oil
`
`Emporation of Acetone
`
`Krill Proteins
`
`Excess Acetone
`
`Separation of Oil and Water
`
`Filt ration/Purification I
`
`N KOTM
`
`3. Manufacturing Controls
`
`The process by which Neptune produces NKOTM occurs under Canadian Good
`Manufacturing Practices (GMP) requirements, as certified by the Natural Health
`Products Directorate (NHPD) of Canada. For GMP Certificate of Compliance
`(Certification Number 0001 349) and Site License (License Number 3001 91), see
`Appendix A.
`
`RIMFROST EXHIBIT 1075 page 0007
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page 6
`
`In addition to conforming to Canadian GMP standards, the Neptune production method
`is internally controlled under the Hazard Analysis Critical Control Points (HACCP)
`system. Under this system, Neptune has identified potential points for potential
`contamination or corruption of NKOTM. To ensure the quality of the final NKOTM
`product, production lots are consistently tested to ensure compliance with appropriate
`international standards addressing residual solvents, microorganisms, heavy metals,
`arsenic, and pesticides as discussed further.
`
`Additionally, Certificate of Registration for a Fish Processing Establishment
`(Certification Number 51 11) was granted to Neptune on February 17, 2005 and was
`renewed on February 17, 2006 by the CFlA following the implementation of a “Quality
`Management Program - Fish” (QMP) and is attached in Appendix A.
`
`4. Specifications for NKOTM
`
`The compositional specifications for NKOTM as established by Neptune are presented
`in Table 1.
`
`The compositional specifications established by Neptune address all relevant issues
`concerning marine oils. They address the quality of the oil by providing minimum
`requirements for fatty acids and phospholipids content, as well as markers of stability
`and purity. The specifications provide information concerning the fat, cholesterol, and
`protein contents of NKOTM, as well as common contaminants, such as pesticides,
`dioxins, and heavy metals, which may be present in marine oils. Finally, Neptune’s
`compositional specifications are similar to those for other marine-derived oils
`considering phospholipid or omega-3 fatty acid-rich oils and are consistent with the
`Codex Alimentarius standard for Edible Fats And Oils Not Covered By Individual
`Standards (CODEX STAN 19-1 981 (Rev. 2-1 999); see Appendix B).
`
`The analytical procedures employed in the analysis have been validated by a variety of
`sources, the most prominent of which are the American Oil Chemist’s Society (A.0.C.S)
`and the Association of Official Agricultural Chemists (A.0.A.C) International. Both of
`these organizations produce validated microbiological and chemical analytical methods.
`In addition, several methods were developed and validated by the Canadian
`government, and additional information on these methods appears in Appendix C.
`
`Results from the analyses for the detailed composition of three different batches of
`NKOTM are presented in Table 2. As can be seen, overall and detailed compositions
`are consistent among the three batches.
`
`RIMFROST EXHIBIT 1075 page 0008
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 7
`
`Requirement-
`
`Specification
`
`TABLE 1. SPECIFICATIONS AND COMPOSITIONAL PROFILE FOR NKO"
`I
`1
`Method( 8 )
`Description
`I Visual - Reddish opaque oil
`I Complies
`Olfactive - Light shrimp odor
`Brookfield 021, 1.5 rpm, 25°C
`I A.O. A. C. 984.20
`
`Appearance
`Odor
`Viscosity
`Humidity
`
`Complies
`1,500 CP
`
`1 CO.9%
`
`Peroxide value
`P-Anisidine Index
`Saponification value
`Iodine Value
`
`Total Protein
`Total Cholesterol
`Total Phospholipids
`Total Lipids as Fatty
`Acids
`Saturated fatty acids
`Monounsaturated fatty
`acids
`Polyunsaturated fatty
`acids
`Omega-3 Fatty Acids
`EPA
`DHA
`DPAa
`Omega-6FattyAcids
`Linoleic acid
`Omega-9 Fatty Acids
`Oleic acid
`Total Trans Fat
`Vitamin A
`Vitamin E
`Esterified Astaxanthin
`
`Identification
`A.O.A.C.965.33
`A.0.C.S Cd 18-90 (modified)
`A.O.A.C.920.160
`A.0.C.S Ja 14-91
`
`0.2 mEq peroxidekg
`1.2
`z 165.0 mg KOH/g
`>130.0 g 12/100 g
`
`BCA Colorimetric method
`A.O.A.C. 976.26 (modified)
`A.O.C.S. Ja7-86
`
`I
`
`A, 0.A.C.996. 06
`
`I A.O.A.C.996.06
`I A.O.A.C.996.06
`I A.O.A.C.996.06
`I A.O.A.C.991.39, 963.22, 996.06
`
`0.5 - 3.5 g/l Om
`0.8 - 1.2 gllOOg
`> 40.0 g/IOOg
`
`I
`
`30.0 g/lOO g
`
`I > 73.0 gI100 g
`I
`I > 12.0 g/100 g
`I > 32.0 g/100 g
`I > 30.0 g/lOOg
`
`AO.A.C.991.39, 963.22, 996.06
`A.O.A.C.991.39, 963.22, 996.06
`
`I A.O.A.C.991.39, 963.22, 996.06
`I A.O.A.C.991.39, 963.22,996.06
`
`A.O.A.C.991.39, 963.22, 996.06
`A.O.A.C.991.39, 963.22, 996.06
`A.O.A.C.991.39, 963.22, 996.06
`
`I A.O.A.C. 996.06
`I J. Sci. Agri. Vol 29 p697-702
`J. Sci. Agri. Vol29 p697-702
`HPLC
`
`> 15.0 g/10@
`> 9.0 gl100g
`> 0.4 gI100 g
`
`I
`I 1.5-2.5g/IOOg
`
`1.3 gI100 g
`> 6.0 g/lOOg
`> 5.0 g/100 g
`I co.1 g/lOOg
`I > 1OO.OlUlg
`> 0.5 lU/g
`> 150.0 mg/100 g
`
`RIMFROST EXHIBIT 1075 page 0009
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page 8
`
`TABLE 2. BATCH ANALYSES OF THREE LOTS OF NKO"
`Assessing the Compositional Guidelines for NKO"
`
`Requirement
`
`Specifidons
`
`Appearance
`Odor
`Vacosity
`Humidity
`
`I Peroxide value
`I Saponification value
`
`P-Anisidine
`
`Red opaque oil
`Light shrimp odor
`< 1,500 CP
`(0.9%
`
`I < 0.2 mEq peroxiddkg I
`I
`I 2 165.0 mg KOHlg
`
`< 1.2
`
`Iodine Value
`
`>130.0 gldl00 g
`
`060116
`
`Complies
`Complies
`564.0
`0.1%
`
`1 .o
`174.6
`
`137.9
`
`h ~ l C h o l e s t e r o l
`Total Phospholipids
`Total Lipids as Fatty
`Acids
`I Saturatedfaltyacids
`I acids
`I Polyunsaturatdw
`Monounsaturated
`
`acids
`
`I
`
`I
`I
`I
`
`0.8-1.2gllOOg
`2 40.0 g/lOOg
`
`2 73.0 g/lOO g
`
`<3O.OgllOOg
`
`2 12.0 g/lOO g
`
`232.0gllOOg
`
`I
`
`I
`I
`I
`
`1.1 gll00g
`47.3 g100 g
`88.5 glloo g
`
`29.8gll00g
`
`22.1 s(100 g
`
`36.7gll00g
`
`Lot Number
`06051 9
`
`I 060224
`
`Complies
`Complies
`476.5
`0.2%
`
`Complies
`Complies
`585.8
`0.1%
`
`I
`
`I
`
`I
`I
`I
`
`0.9
`
`170.8
`
`135.6
`
`I
`
`0.9
`
`177.6
`
`134.3
`
`l.Ogll00g
`44.8 gIlO0 g
`
`81.2 @I 00 g
`
`29.2gllOOg
`
`21.6 gll00 g
`
`34.2gllOOg
`
`I 0.9gllOOg
`43.8 dl00 g
`
`88.2 dl00 g
`I 28.9g400g
`I 21.9#lOOg
`I 37.4glOOg
`
`I
`
`I
`
`I
`1
`I
`
`Vimin A
`Vimin E
`Esterified Astaxanthin
`
`2 100.0 IU/g
`2 0.5 IU/g
`2 150.0 mg'100 g
`
`259.1 IU/g
`0.65 IU/g
`162.6 mgllOO g
`
`336.6 IU/g
`0.67 IU/g
`160.7 mg/lOO g
`
`269.9 IU/g
`0.70 lU/g
`154.0 mg/100 g
`
`000010
`RIMFROST EXHIBIT 1075 page 0010
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill OilTM
`Page 9
`
`5. Purity of NKOTM
`a. Chemical Characterization and Batch Analyses of NKOTM
`
`The potential impurities and incidental constituents present in NKOTM arise largely
`from environmental exposure of the Antarctic Krill. Neptune routinely analyzes
`production lots of NKOTM for the presence of 17 pesticides, as well as dioxins,
`PCBs, heavy metals, fluorine, and various microorganisms. The incidental
`constituents for which NKOTM are analyzed, as well as the methods of analysis and
`associated specifications are presented in Table 3. Results from the batch analyses
`of NKOTM for solvent residues and incidental metal and pesticide constituents are
`presented in Tables 4 and 5, respectively.
`
`C. Microbiological Characterization of NKOTM
`The Neptune production method is internally controlled under the Hazard Analysis Critical
`Control Points (HACCP) system. Under this system, Neptune has identified potential points
`for contamination or corruption of NKOTM. To ensure the quality of the final NKOTM product,
`production lots are consistently tested to ensure compliance with international standards
`addressing residual solvents, microorganisms, heavy metals, inorganic arsenic, and
`pesticides. The microorganisms for which NKOTM is analyzed, as well as the methods of
`analysis and associated specifications, are presented in Table 6. Results of the batch
`analyses of NKOTM for the presence of microorganisms are presented in Table 7.
`
`D. Stability of NKOTM
`
`Three tests have been conducted to ensure the stability of NKOTM over time. The peroxide
`values of NKOTM were measured, and the results indicate the extent of oxidation with oils, the
`Oil Stability Index (OSI), and the Oxygen Radical Absorption Capacity (ORAC).
`
`In the first test, the peroxide value of NKOTM was determined by the A.0.C.S Cd.8b.90 method
`to be 0.05 mEq peroxide/kg, indicating that it does not have a high rancidity potential.
`
`The second test conducted on NKOTM by Neptune examined the OS1 which is a measure of
`the resistance to oxidation and high temperatures over time. In this test, air heated to 97.8OC
`was passed through a sample of NKOTM for a period of 50 hours, during which the
`peroxide values of NKOTM were measured at regular intervals. The results indicate that
`NKOTM maintains a peroxide value of ~ 0 . 1 mEq peroxide/kg for over 50 hours at a
`temperature of 97.8OC, thereby confirming lack of appreciable rancidity potential.
`
`GRAS Associates, i.1.6
`
`0000Tz1
`RIMFROST EXHIBIT 1075 page 0011
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page IO
`
`TABLE 3. INCIDENTAL CONSTITUENTS OF NKO"
`I
`I GC
`
`Methods
`
`Specification
`
`c 10 mglkg
`
`Requirement
`
`Acetone
`
`Arsenic (inorganic)
`
`rcadmium
`Mercury
`Lead
`
`Hydride Generation Atomic
`Absorption
`1 A.O.A.C.986.15, 971.21, 999.1Ia
`A.O.A.C.986.15, 971.21, 999.11
`A.O.A.C.986.15. 971.21. 999.11
`
`c 0.1 mg/kg
`1 ~ 0 . 1 mglkg
`c 0.1 mglkg
`c 0.1 ma/kci
`
`I
`
`I A.O.A.C.970.52, US EPA 8081
`I Hexachlorobenzene
`I
`I
`I A.O.A.C.970.52, US EPA 8081
`I c 0.1 mg/kg
`IHCH
`I Beta-HCH
`I A.O.A.C.970.52, US EPA 8081
`I
`I c 0.05 mglkg
`I A.O.A.C.970.52, US EPA 8081
`I
`I Lindane
`I
`mglkg
`I
`I
`Other Organic or Inorganic Impurities or Toxins
`I Benzo(a)pyrene
`I US EPA610, 8310, and 8100
`I
`I c 2.0 ppb
`US EPA Method 1613 revision B
`Sum of PCDDIPCDF
`2.0 pS/g fat WHO-
`PCDDIF-TEQ/ g fat
`< 10.0 pg/g fat WHO-
`PCDDIF-TEQI g fat
`
`US EPA Method 1613 revision B
`
`Sum of PCDFIFlPCBs
`I
`Total aerobic bacteria
`Total coliforms and E. cdi
`Yeasts and Moulds
`Staphylococcus aureus
`Listeria monocytogenes
`Pseudomonas aeruginosa
`I MFHPB-20-MFO-11
`Salmonella spp.
`a A.O.A.C. Official Methods of Analysis, 18m Ed. Revision 1, 2006.
`Inductively coupled plasma mass spectrometry.
`A pseudomonas selective agar.
`
`Microbial Specifications
`MFHPB-33
`MFHPB-34
`M F HPB-32
`MFLP-21
`MFHPB-30
`CFC agar"
`
`I
`
`10 CFUIg
`10 CFUIg
`10 CFUIg
`10 CFUIg
`Not detected/25 g
`Not detected/25 g
`I Not detected/25 g
`
`RIMFROST EXHIBIT 1075 page 0012
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 11
`
`TABLE 4. BATCH ANALYSES OF 4 LOTS OF NKO" FOR THE PRESENCE
`OF SOLVENT RESIDUES
`
`Requirement
`
`Specification
`
`050201
`
`Lot Number
`I 050610
`I 050223
`
`I 050909
`
`Acetone
`
`< 10 mgkg
`
`7.0
`
`9.1
`
`1.7
`
`<I
`
`TABLE 5. BATCH ANALYSES OF 3 LOTS OF NKO" FOR THE PRESENCE
`OF INCIDENTAL METAL AND NON-METAL CONSTITUENTS,
`PESTICIDE RESIDUES AND ORGANIC OR NON-ORGANIC IMPURITIES
`
`000013
`RIMFROST EXHIBIT 1075 page 0013
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oil"
`Page 12
`
`TABLE 6. MICROBIOLOGICAL SPECIFICATIONS FOR NKO"
`
`Requirement
`Total aerobic bacteria
`I Total coliforms
`Escherichia coli
`Yeasts and Molds
`Staphylococcus aureus
`
`1 Listeria monocytogenes
`
`Methods
`MFHPB-33
`I MFHPB-34
`USP 27=
`MFHPB-32
`MFLP-21
`
`I MFHPB-30
`
`Pseudomonas aeruginosa
`Salmonella spp.
`
`Oxoid-CFC agar
`MFHPB-20-MFO-11
`
`I
`
`I
`
`Specification
`c 1000 CFU/g
`c 10 CFU/a
`NegativeR5 g
`c 100 CFU/g
`c 10 CFU/g
`Negativel25 g
`Negative/25 g
`Negativel25 g
`
`TABLE 7. BATCH ANALYSIS OF 3 LOTS OF NKO" FOR COMPLIANCE
`WITH MICROBIOLOGICAL SPECIFICATIONS
`
`Requirement
`
`Specification
`
`050827
`
`Lot Number
`050503
`
`I
`
`050729
`
`c 1000 CFU/g
`
`c 10 CFU/g
`
`e 10 CFUlg
`
`c 10 CFU/g
`
`c 10 CFU/g
`c 10 CFU/g
`Negative125 g Not Detected
`
`c 10 CFU/g
`Not Detected
`
`c 100 CFU/g
`
`e 10 CFU/g
`
`c 10 CFU/g
`
`e 10 CFU/g
`Not Detected
`10 CFU/g
`
`e 10 CFU/g
`
`c 10 CFU/g
`
`e 10 CFUlg
`
`c 10 CFUlg
`
`Total aerobic
`bacteria
`Total coliforms
`Escherichia coli
`Yeasts and
`Molds
`staphylocOCcus
`aureus
`Listeria
`monocytog enes
`
`Pseudomonas
`aeruginosa
`Salmonella spp.
`
`Negativel25 g Not Detected
`
`Not Detected
`
`1 Negative125 g I Not Detected I Not Detected
`I Negative/25 g I Not Detected I Not Detected
`
`I
`
`I
`
`Not Detected
`
`I Not Detected
`1 Not Detected
`
`I
`
`The ORAC value for an oil is an indicator of the oil's antioxidant capacity against the formation
`of peroxyl and hydroxyl radicals. The presence of peroxyl and hydroxyl radicals increases the
`likelihood of lipid oxidation and associated oxidative rancidity. ORAC analyses involve the
`addition of a fluorescent probe, which measures the scavenging capacity of the antioxidants
`contained in the oil. The results reported were compared to those of Trolox@, a water-soluble
`vitamin E analog which selves as a calibration standard. The ORAC of NKO" was reported
`to be 378 pmol Trolox@ equivalenug, an ORAC value that is more than 300-fold greater than
`that of commercially availably vitamin A and/or vitamin E preparations while also exhibiting
`more than 48-fold greater capacity than a commercial omega-3 18:12 fish oil that is available
`in North America. These results indicate that NKO"
`is a potent free radical scavenger
`indicating the antioxidant potential of the oil.
`
`GRAS Associates. 11.:
`
`000014
`RIMFROST EXHIBIT 1075 page 0014
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 13
`
`Additionally, long-term stability testing revealed that there is no significant change in the
`composition of NKO"
`following storage at 25OC over a period of 28 months, with an ambient
`relative humidity of 60%. The results of this stability test are presented in Table 8.
`
`TABLE 8. STABILITY TESTING CONDUCTED WITH NKO"
`
`Parameters
`
`Specification
`
`Results afbr 28 Months
`Bulk Oil 10303271
`
`E. Fatty Acid Composition of NKOm
`
`As an oil extracted directly from Antarctic krill, NKOm is composed primarily of triglycerides
`and phospholipids. These lipids contain a wide variety of naturally occurring fatty acids as
`their side chains, the most prominent of which are EPA and DHA. The fatty acid compositions
`of multiple production batches of NKO" were examined by Neptune and the mean values,
`with standard deviations, are presented in Table 9.
`
`F. Similarity of Krill Oil to Other Fish Oils
`
`Several forms of fish oil have GRAS status as acknowledged by FDA. Menhaden oil has been
`affirmed as GRAS.3 In addition, FDA has not objected to GRAS notifications submitted on
`tuna oil (FDA, 2002b), salmon oil (FDA, 2004a) and anchovy oil (FDA, 2004b).
`
`In FDA's review of tuna oil, the fatty acid content of tuna oil was compared to menhaden oil
`(FDA, 2002b). Table 10 captures the values used by FDA and compares them to those of krill
`oil.
`
`Additional information from the scientific literature on the distribution of fatty acids in several
`fish oils is shown in Table 11 and is compared to the Neptune data on krill oil. Krill oil contains
`a high level of the desirable n-3 unsaturated fatty acids that is comparable to that of salmon
`oil. Its ratio of n-3 to n-6 fatty acids is comparable to the other fish oils in this group.
`
`G. Natural Occurrence of Vaccenic Acid in Food Sources
`
`Vaccenic acid is one of the major fatty acids present in krill oil. The vaccenic acid in krill oil is
`in the cis- isomeric form as indicated by the low value of trans-fatty acids found in the oil (see
`Table 2). Vaccenic acid constitutes the majority of the monounsaturated fatty acids present in
`krill oil. Vaccenic acid is found in various fish oils in both the cis- and trans- forms at levels of
`
`RIMFROST EXHIBIT 1075 page 0015
`
`

`

`2-4% and can also be found in a number of other fats and oils. Table 12 lists the vaccenic
`acid concentrations in several widely used vegetable oils as found in two references, although
`it is noted that neither author indicated whether the vaccenic acid was in the cis- or trans-
`form. These oils are reported to be used in a wide variety of food applications.
`TABLE 9. FArpl ACID CONTENT OF NKOm
`I Mean Fatly Acid
`Content
`(Standard
`Deviation)
`0.20 (0.03)
`11.37 (0.57)
`0.22 (0.05)
`0.43 (0.06)
`22.17 (1.04)
`7.46 (1.40)
`1.27 (0.32)
`10.61 (1.18)
`8.54 (0.69)
`1.73 (0.22)
`0.18 (0.03)
`1.31 (0.60)
`3.57 (1.65)
`0.77 (0.12)
`0.08 (0.05)
`0.33 (0.10)
`
`Fatty Acid Content Of NKO" Lots
`I
`(%of total lipid)
`I050117 I 050201
`050223
`0.19
`0.18
`0.20
`11.29
`10.51
`12.07
`0.17
`0.19
`0.27
`0.51
`0.51
`0.46
`I 21.58
`23.4
`22.14
`7.37
`1.35
`11.14
`7.33
`1.94
`0.2
`1.43
`3.72
`0.75
`0.08
`0.37
`
`I
`
`I
`
`10.61
`9.07
`1.57
`0.21
`1.1
`
`I
`I 2.39
`
`11.12
`0.37
`0.45
`
`0.83
`7.34
`
`0.17
`2.47
`
`0.08
`0.43
`
`I
`I
`
`1
`I
`
`I
`I
`
`I
`I
`
`10.34
`I 8.14
`2.04
`0.17
`
`I 1.59
`I 4.27
`
`0.37
`
`I
`I 0.42
`I 0.47
`
`17.12
`
`16.03
`
`0.24
`
`1
`I 0.24
`I 0.41
`
`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 14
`
`*-
`
`FattyAcid
`
`212 :o (lauric)
`C14 :O (myristic)
`C14 :I (myristdei)
`C15 :O (pentadecanoic)
`C16 :O (mlrnitic)
`C16 :I (palrnitoleic)
`C18 :O (stearic)
`C18 :I n-7 (vaccenic)
`C18 :I n-9 (oleic)
`C18 :2 n-6 ( linoleic)
`C18 :3 n-3 (gamma-linoleic)
`C18 :3 n-6(alpha-lindei)
`C18 :4 n3(stearidonii)
`C20 :I (eicosenic)
`C20 :4 n-3 (eicosatetranoic)
`C20 :4 n-6 (arachidonic)
`C20 :5 n-3
`(eicosapentaenoic acid,
`E PA)
`C22 :I n-9 (erucic)
`C21 :5 n-3
`(heneicosapentaenoic)
`C22 :5 n-3
`(docosapentaenoic acid,
`DPA)
`C22 :6 n-3
`(docosahexaenoic acid,
`DHA)
`
`16.43
`
`17.35
`
`16.71 (0.65)
`
`0.54
`
`0.48
`
`0.46
`
`0.38
`
`0.53
`
`0.45
`
`1
`
`0.47 (0.10)
`
`0.47 (0.06)
`
`0.42 (0.08)
`
`0.46
`
`0.39
`
`10.53
`
`7.94
`
`9.70
`
`11.97
`
`9.50 (1 .54)
`
`OOC016
`RIMFROST EXHIBIT 1075 page 0016
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 15
`
`TABLE IO. FATTY ACID COMPOSITION OF KRILL OIL" COMPARED
`TO TUNA OIL AND MENHADEN OILb (g/lOO g)
`
`TABLE 11. COMPARISON OF FATTY ACID CONTENTS OF VARIOUS
`FISH OILS (PERCENTAGES)
`1 AnchoM I Menhaden' I Salmon' I Krill
`
`I Fatty
`
`I
`
`a From Lucas et al., 2005.
`
`RIMFROST EXHIBIT 1075 page 0017
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 16
`
`TABLE 12. VACCENIC ACID CONCENTRATION (% FATTY ACIDS)
`IN VARIOUS FATS AND OILS
`
`From Sauer et al., 1997
`
`Vaccenic acid and other isomers of 18:l fatty acids occur naturally in milk fats and have
`been found to be protective against cancers in animal and in vitro models, with some
`supporting epidemiological evidence in humans (Doyle et ai., 2005). The vaccenic acid
`present in dairy products is in the trans isomeric form. Cod et al. (2003) showed that the
`level of vaccenic acid and other trans1 8:l isomers in butter can be increased by varying
`the diets of the producing cows. The levels of trans-1 8: 1 and other C18 fatty acids in
`butter from cows on a conventional corn ration versus a corn ration supplemented with 2%
`sunflower oil and 1% fish oil is shown in Table 13.
`
`Table 13. DISTRIBUTION OF C18 FATTY ACIDS IN BUTTER'
`
`Acid
`
`18:O
`
`18: 1, trans-9
`18: 1 , trans-10
`
`18: 1. trans-1 2
`18: 1, Cis-9
`18:2, cis-9, cis-1 2
`1812 Cis-9, tmnS-11
`18:3 cis-9, cis-12, cis-1 5
`From Cor1 et al., 2003
`
`0.77
`25.35
`3.33
`0.51
`0.41
`
`Ration
`11.85
`
`Ration
`5.09
`
`2.68
`16.28
`2.48
`14.23
`2.93
`3.76
`0.46
`
`RIMFROST EXHIBIT 1075 page 0018
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 17
`
`111.
`
`INTENDED DIETARY USE
`
`A. Current Usage of NKO" in the Diet
`
`Krill oil is not presently added to foods and is not naturally present in foods consumed in the
`US. Neptune reported that krill has been used in human food since the 1970s, principally in
`Japan, Russia, Ukraine, and, more recently, France.
`
`Neptune submitted a notification to FDA in 2002 for use of krill oil in dietary supplements.
`FDA did not object to this use (FDA, 2002a). The supplement is sold in 300 and 500 mg
`capsules with a recommended dose of 1 - 2 capsules per day.
`
`Softgels containing the NKO" have been available as dietary supplements in North America
`for several years; in Canada and the US, NKO" has been commercially available since
`October of 2002, and in Japan, Korea, Singapore, and Hong Kong, NKO" has been available
`since December of 2004. A total of 120,000 kg of Neptune krill oil, which is equivalent to
`60,000,000 individual 500 mg softgel capsules, has been ingested by consumers. Throughout
`this time no serious adverse effects have been reported.
`
`B. Intended Uses
`
`Neptune intends to incorporate krill oil into the various food categories as listed in Table 14.
`The krill oil would function as a nutrient supplement as defined in 21 CFR 170.3(0)(20).
`Neptune research has indicated that use levels are self-limiting due to the strong taste which
`begins to be detected at levels of 300 - 500 mg per serving, depending on the food type. It
`should be noted that krill oil is intended to be used as a substitute or alternative to fish oil.
`
`TABLE 14. PROPOSED FOOD USES FOR NKO"
`
`I
`I
`
`Food Category
`
`Breakfast cereals
`
`Cheeses
`Fruit juices
`
`Milk products
`
`Medical foods
`
`I
`I
`
`Use Levels Per Serving
`
`300 mg
`
`300 mg
`150-250 mg
`
`300-500 mg
`
`300-500 mg
`
`C. Estimated Daily Intake
`
`The estimated daily intake of NKOm from proposed food uses above was calculated
`according to FDA guidelines (FDA, 1995) using serving size data and the mean (50th
`percentile) consumption of each food type of interest from the CSFll 1994-96 Survey (USDA,
`
`GRAS Associates L i C;
`
`00~019
`RIMFROST EXHIBIT 1075 page 0019
`
`

`

`GRAS Assessment for Neptune Technologies & Bioressources
`Neptune Krill Oilm
`Page 18
`
`2005). According to FDA