CN1620253A - Seal oil based lipid emulsions and uses thereof - Google Patents
Seal oil based lipid emulsions and uses thereof Download PDFInfo
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- A23D—EDIBLE OILS OR FATS, e.g. MARGARINES, SHORTENINGS, COOKING OILS
- A23D7/00—Edible oil or fat compositions containing an aqueous phase, e.g. margarines
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Abstract
The present invention provides seal oil based lipid emulsions that are rich in long chain .OMEGA.-3 fatty acids. The present invention further provides methods of preparing the emulsions and for the use of the emulsions in clinical settings for a variety of applications, including total parental nutrition.
Description
Technical field
The present invention relates to lipid emulsion system field, and relate to the sea dog oil base lipid emulsion that is used for parenteral administration specifically.
Background technology
The lipid emulsion is widely used as the basis of total parenteral absorption (TPN) infusion solution, and being used for can not provide heat and necessary fat acid with the patient that oral route is accepted nutrition to those.For example, in the sixties in 20th century, Intralipid is just developed in Europe
Intravenous lipid emulsion, and to 1975 the approval use in the U.S..The lipid emulsion that is used for parenteral use of present commercially available acquisition is made up of vegetable oil (such as soybean oil or safflower oil), emulsifying agent (such as the egg phosphatide class), G ﹠ W.The emulsion of these commercially available acquisitions comprises Liposyn
And LiposynII
(Abbott Laboratories); Travemusion
(Baxter Healthcare) and Soyacale
(Alpha Therapeutics).
The biochemical progress of lipid has confirmed that how unsaturated the biology of Omega-3 (ω-3) and Omega-6 (ω-6) aliphatic acid is essential.Aliphatic acid characterizes with the number of two keys and by the position of first pair key with respect to molecule methyl end by the C atom that they had.Thus, what name C18:2 ω-6 represented is the aliphatic acid with 18C atom and 2 two keys, and first double bond position is in the 6th C atom from the methyl end.Polyunsaturated fatty acid is important as the film of setting up Biochemical processes blocking-up and modulator.Biologically most important ω-6 aliphatic acid is arachidonic acid (AA; C20:4) and precursor, linoleic acid (C18:2).Alpha-linolenic acid is the parent material of long-chain omega-fatty acid eicosapentaenoic acid (EPA), clupanodonic acid (DPA) and DHA (DHA), and these long-chain omega-fatty acids are the essential fatty acids in the human body.As the precursor in high biological activity eiconaoids synthetic, AA and EPA affect inflammatory reaction (Burton JL.Lancet, 1989,433-5) and immunology resistance (Endrers S, Deng, Eur J Clin Invest, 1995,25:629-38) and angiocardiopathy (Burr ML, Deng, Lancet, 1989,2:757-61; Hu FB, etc., JAMA, 2002,287 (14): 1815-21), disorders of lipid metabolism, thrombosis process and neoplastic disease (Jeeski LI, etc., Pro Soc Exp Biol Med 1995,210:227-33; Tevar R, etc., J Parenter Enteral Nutr, 2002,26 (5): 285-9; BarberMD, etc., Nutr Cancer, 2001,40 (2): 118-24; Senzaki H, etc., WorldRev Nutr Diet, 2001,88:117-25).(for example skin lesion of some clinical symptoms, growth is ended, kidney is degenerated and metabolic rate increases) and the shortage of ω-6 aliphatic acid relation (Caldwell M.D. is arranged, Human essential fatty acids deficiency:A reviewin fat emulsion in parenteral nutrition.Meng H.C. and Wilmore D.W. (eds.) Chicago, IL:Amer.Med.Assoc.P.24,1978).
Epidemiological study (it shows the relation between food fat content and the angiocardiopathy generation) thinks that polyunsaturated fatty acid is that prevention is effective.In this content, the omega-fatty acid of finding in marine oil is according to proving useful especially.Recently, people recognize, keep the balance between the consumption of ω-3 and ω-6 aliphatic acid, are necessary to the protectiveness pharmacological effect of realizing these aliphatic acid.ω-3 is 1 with the ratio of ω-6 aliphatic acid: 2-1: the 4th, recommend (Helmut G. etc., JPEN, 1994,18:417-421).
As mentioned above, the lipid emulsion of most of commercially available acquisition is obtained from vegetable oil.It is dual being obtained from the relevant defective of vegetable oil with emulsion.At first, vegetable oil lacks omega-3 unsaturated fatty acid, and secondly, the vitamin K that contains significant quantity in the plant is (for example, at Intralipid
And LiposynII
10% emulsion in, the concentration that it is found that vitamin K1 is respectively 30.8 and 13.2 micrograms/dL).Vitamin K is one of key factor in the blood clotting process and may makes surgical procedure become complicated thus, particularly concerning the patient who accepts anticoagulant therapy.
Along with replenishing parenteral absorption with omega-fatty acid, present studies show that under some clinical condition, can draw various protectiveness effects
10,11, people develop fish oil base lipid emulsion, Omegaven
(Fresenius Kabi), and allowed on the European market, to use.Omegaven
Be 10% Fish Oil Emulsion, it contains the omega-fatty acid EPA and the DHA of high degree.Many biology beneficial effects according to the show all with use Omegaven
Be related, comprise LTC
5Producing increase, platelet aggregation reduction, immunologic function raising and oedema formation slows down.With Omegavene
With basic lipid emulsion such as Intralipid
Intravenous is bestowed the ratio of the ω-3 with ω-6 aliphatic acid that also provide best simultaneously.
Yet, contain very small amount of omega-fatty acid DPA in the fish oil.The same with polyunsaturated fatty acid EPA and DHA, DPA is that nutrition is important, and about 1/3rd of the long-chain omega-fatty acid that circulates in the blood of human body all belongs to DPA.Recent studies confirm that in fact EPA can change into DPA in vascular wall, and DPA is a potent agent under these environment originally.Though believing EPA usually is key factor in producing prostaglandin, wherein prostaglandin makes the arterial wall softness and does not contain atheromatous plaque, and this studies show that DPA can be that 10-20 doubly more has effect than EPA in this regard.DPA can play strong effectively anti-arteries and veins gruel type factor (the Schiefenneier M.﹠amp that activates thus; Yavin E., J Lipid Res, 2002,43 (1): 124-31; Hansen JB, etc., J Lipid Res, 2000,41 (9): 1376-83).
Another potential defective of using fish oil in the lipid emulsion is that the method for extracting fish oil depends on steam or heating at present, and it makes the omega-fatty acid oxidation.High temperature also has the acceleration omega-fatty acid is isomerizated into trans geometry from its natural cis geometry possibility.Oxidation and trans-fatty acid all are harmful to the health of human body.
Therefore, still need a kind of lipid emulsion, it contains omega-fatty acid EPA, DPA and the DHA of high degree, the oxidation and the trans-fatty acid of low content vitamin K and low ratio.
This background information is that the applicant believes that these information and the present invention have the relevance of possibility for the purpose of making Given information provides.There is no need to admit also should not be interpreted as aforesaid any information constitutes prior art of the present invention.
Summary of the invention
An object of the present invention is to provide Seal Oil-Based Lipid Emulsion And Its Use.One aspect of the present invention provides a kind of lipid emulsion, and this emulsion contains:
(a) the purifying seal oil of 5-40wt%;
(b) emulsifying agent of 0.5-5wt% and
(c) water,
The final pH of wherein said emulsion is 6.0-9.0.
Another aspect of the present invention provides a kind of alimentation composition, and said composition contains:
(a) contain the emulsifying agent of purifying seal oil, 0.5-5wt% of 5-40wt% and water the lipid emulsion and
(b) one or more nutrient compounds,
The final pH of wherein said composition is 6.0-9.0.
Another aspect of the present invention provides a kind of alimentation composition, and said composition contains: contain the purifying seal oil of 5-40wt%, the emulsifying agent of 0.5-5wt% and lipid emulsion and one or more therapeutic agents of water.
Another aspect of the present invention provides a kind of alimentation composition, and said composition contains: contain the purifying seal oil of 5-40wt%, the emulsifying agent of 0.5-5wt% and lipid emulsion and one or more diagnosticums of water.
Another aspect of the present invention provides the purifying seal oil of a kind of 5-40wt% of containing, the emulsifying agent of 0.5-5wt% and the purposes of lipid emulsion in total parenteral absorption of the patient of needs of water.
The purposes of the transmission carrier that the lipid emulsion that another aspect of the present invention provides the emulsifying agent of purifying seal oil, 0.5-5wt% of a kind of 5-40wt% of containing and water is used as therapeutic agent or diagnosticum.
Brief description
Fig. 1 has described the influence of liquid droplet size of the comparison 10% seal oil emulsion of lecithin and seal oil.
Fig. 2 has described the influence of pH to the zeta potential of 10% seal oil emulsion.
Fig. 3 has described the snperoxiaized influence of pH to 10% seal oil emulsion.
Fig. 4 has described the snperoxiaized influence to 10% seal oil emulsion of EDTA and/or vitamin E.
Fig. 5 has described the snperoxiaized influence of EDTA concentration to 10% seal oil emulsion.
Fig. 6 has described to contain the peroxide value of the oily emulsion of difference.Rhombus: Intralipid
, prepare by soybean oil; Square: Omegaven
, by Preparation fish; Triangle: 10% seal oil emulsion.
Fig. 7 has described the snperoxiaized influence of autoclaving time to 10% seal oil emulsion.
Fig. 8 has described the influence of autoclaving time to the pH of 10% seal oil emulsion.
Fig. 9 has described the influence of autoclaving time to the zeta potential of 10% seal oil emulsion.
Figure 10 has described the influence of autoclaving time to the grain graininess of 10% seal oil emulsion.
Detailed Description Of The Invention
The invention provides a kind of lipid emulsion, it contains seal oil as inner core lipid, is used for using use in treatment, is included in the application in total parenteral absorption.
Definition
Unless other explanation is arranged, all technology used herein and scientific terminology all have with the general technical staff of the technical field of the invention the identical implication usually understood.
Term " about " used herein is that relative nominal value has+/-5% variation. Should be understood that no matter whether this variation always is included in any given number range that this paper provides, particularly point out.
The component of lipid emulsion
According to the present invention, the emulsifying agent that the lipid emulsion contains seal oil and is dispersed in water. Emulsion can also contain annexing ingredient, such as antioxidant, and chelating agent, buffer, assistant for emulsifying agent, permeability (osmolality) improver, nertralizer etc., they are used for improving stability, uniformity and/or other characteristic of emulsion.
Compare present obtainable fish oil base emulsion, the more stable and resistance to oxidation more of sea dog oil base lipid emulsion of the present invention. In addition, contain essential fatty acid clupanodonic acid (DPA) in the seal oil base emulsion, its in the fish oil base emulsion only to exist on a small quantity.
1. seal oil
Lipid emulsion of the present invention contains seal oil as the inner core lipid. The sea dog of various kinds all is suitable as the source that described emulsion is used oil. In one embodiment of the invention, oil derives from harp seal (harp seal).
As mentioned above, at present contain the oil that derives from plant or contain fish oil as the inner core lipid in the obtainable lipid emulsion that is used for the treatment of purposes. Seal oil provides the many advantages that surpass fish oil or vegetable oil. For example, major part derives from the oil of plant all has the useful omega-fatty acid of low content and the vitamin K of relative high-load, and contains the omega-fatty acid of high-load and the vitamin K of low content in the seal oil. In addition, the omega-fatty acid content of seal oil is higher than the content that exists in fish oil, and for example, the omega-fatty acid content of harp seal oil is generally about 20-35%. In addition, the most of omega-fatty acid in the seal oil is in occupation of the 1st and the 3rd of substituted glycerol three ester molecules, and is identical with the position in the people, and the omega-fatty acid in the fish oil mainly occupies the 2-position. Therefore, this replacement mode of seal oil can allow the recipient absorb better lipid in the emulsion, because the lipid of seal oil more is similar to endogenous lipid.
With regard to the content of omega-fatty acid, seal oil contains the ratio of higher DHA (DHA)/eicosapentaenoic acid (EPA) than fish oil, and the more cholesterol of not expecting and the saturated fatty acid of low content. In addition, contain the essential fatty acid clupanodonic acid (DPA) of significant quantity in the seal oil, and the content that it exists is very low in fish oil. Be fit to very much in the seal oil of therapeutic purposes in preparation, above-mentioned characteristic gives seal oil superior liquid properties in addition, so that the semisolid that its unlikely formation can occluding vascular.
According to the present invention, the seal oil that is used for using as emulsion inner core lipid is highly purified and contains the polyunsaturated fatty acid, particularly omega-fatty acid of high concentration. It is high that ratio ω-6 aliphatic acid of the omega-fatty acid that contains in this oil is wanted.
The concentration of omega-fatty acid generally is about 20-35% of gross weight in the seal oil, and the concentration of ω-6 aliphatic acid generally is about 2-10% of gross weight. Yet those skilled in the art can understand, and it is to depend on the method for extraction and refining and different that the aliphatic acid of oil forms.
According to the present invention, the seal oil that in emulsion, uses contain gross weight greater than about 20% omega-fatty acid.In one embodiment, seal oil contain gross weight greater than about 25% omega-fatty acid.In another embodiment, seal oil contain gross weight greater than about 27% omega-fatty acid.
In seal oil, eicosapentaenoic acid (EPA) and DHA (DHA) occupy higher proportion in the omega-fatty acid content of oil.For example, in general, in harp seal oil, the concentration of eicosapentaenoic acid (EPA) and DHA (DHA) accounts for about 45-70% of omega-fatty acid gross weight, and leukotrienes accounts for about 0.5-1.5% of omega-fatty acid gross weight.
According to the present invention, in the seal oil that uses in emulsion of the present invention, DHA is about 1: 1 to about 3.5: 1 with the ratio of EPA.
The content of fatty acid of seal oil can also be the form of the ester of aliphatic acid, the ester of linoleic acid (18:2n6), arachidonic acid (20:4n6), leukotrienes (18:3n3), eicosapentaenoic acid or EPA (20:5n3) and DHA or DHA (22:6n3) for example, and the ester of other saturated fatty acid, as the ester of myristic acid (14:00), palmitic acid (16:00) and stearic acid (18:00).With regard to acid number (according to United States Pharmacopoeia 1993), generally contain the free fatty acids (about 0.2-0.9ml/g) of low content in the seal oil.Those skilled in the art can understand, and the amount of the fatty acid ester in the oil depends on the technology that is adopted in olein refining process.
According to the present invention, the free fatty acid content of used seal oil is less than about 0.5ml/g in the emulsion.In one embodiment, free fatty acid content is less than about 0.4ml/g.In another embodiment, free fatty acid content is less than about 0.3ml/g (all numerical value are all measured according to United States Pharmacopoeia 1993).
The quality of oil can be estimated by parameter known in the art, and for example, peroxide value, iodine number or aliphatic acid are formed.These parameters can be measured by many methods well known by persons skilled in the art, for example, parameter can be measured (also referring to the analytic process of describing in the United StatesPharmacopoeia XXI and XXII and British Pharmacopoeia 1993 and 1998) by gas chromatography.
According to the present invention, the peroxide value of the seal oil that in emulsion, uses for about 3mEq/kg to about 35mEq/kg and average iodine number be greater than 60mEq/kg.In one embodiment, the iodine number of seal oil is that about 85mEq/kg is to about 100mEq/kg.
The method of extraction and refined oil is well known in the art.For example, International Patent Application PCT/CA00/00028 has described a kind of method of using low-heat to come refining animal and plant oil.The use of low temperature can farthest reduce the harmful oxidation that exists in the purifying oil and the amount of trans-fatty acid.Additionally, seal oil is often than the more anti-autoxidation process of fish oil, and preliminary experiment proves, in seal oil, the external degree of oxidation of omega-3 polyunsaturated fatty acids is less than observed half in fish oil.According to the present invention, seal oil has the oxidation and the trans-fatty acid of low content.
In one embodiment of the invention, by low temperature method extraction and refining seal oil, so that farthest reduce oxidation.In another embodiment, processing temperature is remained on or be lower than 22 ℃.
Lipid emulsion of the present invention generally contains 5% seal oil to about 40% (w/v) of having an appointment.In one embodiment, emulsion contains 5% seal oil to about 30% (w/v) of having an appointment.In another embodiment, emulsion contains 5% seal oil to about 20% (w/v) of having an appointment.In another embodiment, emulsion contains 5% seal oil to about 15% (w/v) of having an appointment.In another embodiment, emulsion contains the seal oil of have an appointment 10% (w/v).
2. emulsifying agent
For preparing lipid emulsion of the present invention, one or more emulsifying agents are mixed with the seal oil component.The phospholipid in normally natural, the synthetic or semi-synthetic source of the emulsifying agent that is used for this purpose.Various examples of suitable emulsifiers are known in the art.The example of suitable emulsifying agent includes but not limited to Yolk lecithin, egg lecithin, L-α-DPPC (DPPC), DL-α-two palmityl phosphatidyl-ethanolamine (DPPE), dioleoyl phospholipid phatidylcholine (DOPC) and soybean lecithin.According to the present invention, the triglycerides in the emulsifying agent and the total concentration of free fatty should be low, so that farthest reduce the influence to the total oil concentration of emulsion.In one embodiment of the invention, the total concentration of triglycerides in the emulsifying agent and free fatty is less than about 3.5%.
In one embodiment of the invention, use lecithin as the emulsifying agent in the lipid emulsion.In another embodiment, use egg lecithin as emulsifying agent.In another embodiment, use contain the 80-85% phosphatid ylcholine and less than the egg lecithin of about 3.5% fat as emulsifying agent.Those skilled in the art can understand, and can have other component that can influence emulsifiability in egg lecithin sharply.For example, can contain in phosphatidyl-ethanolamine, lysophosphatidyl choline, lysophosphatidyl ethanolamine, sphingomyelins and other natural constituents one or more in the egg lecithin.
Generally contain 0.5% the emulsifying agent of having an appointment in the lipid emulsion of the present invention to about 5% (w/v).In one embodiment of the invention, emulsion contains 0.6% emulsifying agent to about 2% (w/v) of having an appointment.In another embodiment, emulsion contains 0.8% emulsifying agent to about 1.8% (w/v) of having an appointment.In another embodiment, emulsion contains 1.0% emulsifying agent to about 1.5% (w/v) of having an appointment.In another embodiment, emulsion contains the emulsifying agent of have an appointment 1.2% (w/v).
Aspect the granularity of the ratio of the lecithin in the emulsion and seal oil formed oil droplet in the decision emulsion is important.According to the present invention, the oil droplet granularity in the emulsion is lower than 1 μ m.Therefore, lecithin is about 1: 4 to about 1: 20 with the ratio of seal oil.In one embodiment of the invention, lecithin is about 1: 4 to about 1: 18 with the ratio of seal oil.In another embodiment, lecithin is about 1: 4 to about 1: 15 with the ratio of seal oil.In another embodiment, lecithin is about 1: 4 to about 1: 10 with the ratio of seal oil.
3. other additive
Can also contain annexing ingredient in the lipid emulsion of the present invention, such as antioxidant, chelating agent, permeability agent, buffer, nertralizer or the like, they are used for improving stability, uniformity and/or other characteristic of emulsion.
The present invention includes to the lipid emulsion and add one or more antioxidants, so that help to prevent to form the oxidation of fat acid of not expecting.The suitable antioxidant that can add in the lipid emulsion includes but not limited to alpha-tocopherol (vitamin E) and tocotrienols (tocotrienols).As known in the art, tocotrienols is the natural blend of tocotrienols and vitamin E extract, forms by concentrating in the rice bran oil distillate, and it has the antioxidation activity that is similar to alpha-tocopherol (vitamin E).Tocotrienols has the structure similar to vitamin E and contain three two keys on the carbon side chain of molecule.
When using, the concentration of the antioxidant that adds in emulsion is generally about 0.002 to about 1.0% (w/v).In one embodiment, the concentration of used antioxidant is about 0.02% to about 0.5% (w/v) in emulsion.
In one embodiment of the invention, tocotrienols is added in the emulsion as antioxidant.In another embodiment, in emulsion, add the tocotrienols of about 0.5% (w/v).In another embodiment, vitamin E is added in the emulsion as antioxidant.In another embodiment, in emulsion, add the vitamin E of about 0.02% (w/v).
Chelating agent be can also contain in the emulsion, the stability of emulsion and the formation of reduction oxidation of fat acid are used for improving.Suitable chelating agent is known in the art and is logical safe (GRAS) compound of thinking that the example includes but not limited to EDTA.In one embodiment of the invention, contain EDTA in the emulsion.In another embodiment, contain in the emulsion and have an appointment 1 * 10
-6M to 5 * 10
-5The EDTA of M concentration.
The permeability agent can also be mixed in the emulsion, be suitable for the numerical value that parenteral gives so that the permeability of emulsion is adjusted to.The amount and the type of the permeability agent of using in the parenteral emulsion are well known in the art.An example of suitable permeability agent is a glycerine.The concentration of permeability agent is generally about 2% to about 5% (w/v).The amount of the permeability agent of adding in emulsion in one embodiment of the invention, is about 2% to about 4%.The amount of the permeability agent of adding in emulsion in another embodiment, is about 2% to about 3%.In another embodiment, agent is added in the emulsion as permeability with the glycerine of about 2.25% (w/v).
The pH that it will be appreciated by those skilled in the art that emulsion can adjust by using buffer or nertralizer.According to the show, the pH value near physiology pH or on the tendency of the rarer fatty acid peroxidase of emulsion.Those skilled in the art can understand, can by use suitable alkali (in and on the aliphatic acid negative electrical charge), by using suitable buffer or its combination, regulate the pH of emulsion.Various types of alkali and buffer are applicable to emulsion of the present invention.Those skilled in the art can understand, and adding buffer to emulsion not only can influence final pH, but also influences the ionic strength of emulsion.High ionic strength can influence the zeta potential (being the surface charge of oil droplet) of emulsion negatively and therefore not expect.The general knowledge of selecting suitable buffer intensity to provide suitable pH and zeta potential (as defined herein) to be considered to belong to those skilled in the art.
In one embodiment of the invention, use NaOH to regulate the pH of emulsion.In another embodiment, regulate pH with buffer.In another embodiment, buffer is a PB.In another embodiment, NaOH and PB are added in the emulsion simultaneously.
The final pH of emulsion is generally about 6.0 to about 9.0.In one embodiment of the invention, the pH of emulsion is about 7.0 to about 8.5.In another embodiment, the pH of emulsion is about 7.0 to about 8.0.
Can also contain in the lipid emulsion and be useful on the component of adjusting stability of emulsion, for example amino acid or carbohydrate, for example fructose or glucose.The lipid emulsion can also be mixed with the nutrient that contains such as glucose, amino acid, vitamin or other parenteral absorption tonic.The lipid emulsion is mixed with the admixture therapeutic agent also is considered to belong to scope of the present invention." therapeutic agent " used herein is meant the material of physiology or pharmacological activity, it can produce part or whole body effect or multiple effect in animal, and is often referred to medicine, nutritional supplement, vitamin, mineral matter, enzyme, hormone, protein, polypeptide, antigen and other treatment or diagnoses useful compound.
The preparation of lipid emulsion
Lipid emulsion of the present invention can prepare by many routine techniques well known by persons skilled in the art.Generally speaking, at first the inner core lipid is mixed with emulsifying agent and antioxidant (if using it).Then, carry out constant agitation in the water simultaneously, prepare emulsion by this oil phase slowly is added into.If use the permeability agent, it is added in the water, mix with oil phase then.If necessary, can regulate pH, and can adjust final volume for water if necessary in this stage.
The oil droplet granularity of emulsion (being grain graininess) is an important parameters for the result of treatment of emulsion and quality.Because lipid granule can be removed from the body circulation in the mode that is similar to chylomicron, thus the granularity of the lipid granule in the emulsion need remain within the particle size range (being 0.4-1.0 μ m) of naturally occurring chylomicron or under.If grain graininess is greater than this scope, lipid granule can be deposited in liver, spleen and the lung, and (RahuiC.M. is etc., Am.J Hosp.Pharm.1992,49:2749-2755) to cause behind infusion producing significant fat load.Lipid with granule granularity is dispersed in the emulsion preferably, and often produces safety and more stable emulsion.Lipid emulsion of the present invention generally has the littler and homogeneous granules granularity more of lipid emulsion than commercially available acquisition.Suitable grain graininess is to have average diameter to be lower than 1 μ m, generally is that about 0.05 μ m is to about 0.5 μ m.In one embodiment of the invention, the average grain granularity is that about 100nm is to about 500nm.In another embodiment, the average grain granularity is that about 200nm is to about 400nm.In other embodiments, the average grain granularity for about 250nm to about 350nm and about 300nm about 350nm extremely.It will be appreciated by those skilled in the art that obtainable grain graininess depends on the composition and the preparation process of preparation.
Lipid emulsion with fair average grain graininess (being that diameter is less than 1 μ m) can prepare by many technology known in the art.For example, can handle carrying out homogeneous processing, sonication or Micro Fluid by the dispersion liquid that the interpolation oil phase makes in water as mentioned above.Can commercially available various suitable plant equipment, realize the preparation of emulsion.In one embodiment of the invention, make dispersion liquid pass through the homogenizer one or many.In another embodiment,, use 20 by using homogenizer, 000-27, the pressure of 000Psi produces emulsion.In another embodiment, by using homogenizer, use 180, the pressure of 000kPa produces emulsion.Those skilled in the art can understand, in this way the pressure of the grain graininess of Zhi Bei emulsion and oil content, emulsifier concentration and homogenizer and relevant by its number of times.The selection that is suitable for preparing the condition of emulsion of the present invention is considered to belong to those skilled in the art's ordinary skill category.
If necessary, in this stage, the emulsion of gained can be passed through for example 0.45 μ m membrane filtration of suitable filter.Perhaps, can handle, emulsion is sterilized by autoclaving.It is found that the emulsion that contains seal oil of the present invention is stable to handling maximum 60 minutes at 121 ℃ of following autoclavings.
Usually, remain on final emulsion in the aseptic condition and can be packaged in and carry out short-term or long-term preservation in the closed container.
The generation lipid oxidation also belongs to scope of the present invention to preparation lipid emulsion under inert atmosphere in the process so that farthest be reduced in.
In one embodiment of the invention, the lipid emulsion contains seal oil, the egg lecithin that contains 80-85% phosphatide, 0.002-0.5% tocotrienols or alpha-tocopherol of 0.5-5%, the 2-5% G ﹠ W of 5-20%.In another embodiment, the omega-fatty acid that contains 20-35% in the seal oil.In another embodiment, use temperature is remained on 22 ℃ or be lower than 22 ℃ technology, seal oil is carried out refining.In another embodiment, the average grain granularity of emulsion is that about 250nm is to about 350nm.In another embodiment, the final pH of emulsion is about 7 to about 8.
The stability of lipid emulsion
The oxidation that at utmost reduces lipid is a particular importance to the lipid emulsion, because the oxidation meeting causes developing into and becomes sour and produce the molecule with genotoxic potential.Just as known in the art, the existence of the character of concentration, the emulsifying agent of the granularity of oil droplet, oil and antioxidant is important decisive factor to the amount of the lipid oxidation that taken place in the emulsion.
The stability of lipid emulsion can include but not limited to creaming of emulsion and/or separation, accelerated ageing test, high speed centrifugation, measurement grain graininess and the ζ-Potential Distribution and the variation of vision-based detection emulsion by many known in the art and measure at this paper and described elsewhere test.Can estimate the state of oxidation of seal oil in advance by measuring peroxide value, and determine the quality of emulsion thus.The method of measuring the peroxide content of oil is (for example, referring to the analytic process of describing among the United States Pharmacopoeia XXI or the British Pharmacopoeia1993) known in the art.Seal oil generally has the low-peroxide value of about 4mEq/kg to about 32mEq/kg, depends on the method for manufacturing.Thus, in one embodiment of the invention, the peroxide value of seal oil is less than 35mEq/kg.In another embodiment, the peroxide value of seal oil is that about 5mEq/kg is to about 30mEq/kg.
The oxidation level of lipid emulsion can also be by measuring peroxide value, measuring by the consumption of monitoring oxygen or by detecting lipid peroxide.
Toxotest
Use standard method known in the art, can be external and the toxicity of body build-in test lipid emulsion (referring to, for example, Current Protocols in Toxicology, Maines, M. waits volume), J.Wiley ﹠amp; Sons, Inc., New York, NY).For example, can use the test of LimulusAmebocyte Lysate (LAL) blood clot (Marilyn J.Gould, NephrologyNews ﹠amp; Issues, November 1988, pp26), come the endotoxin in the testing in vitro emulsion to exist.Perhaps, or in addition, can use the standard acute toxicity test, wherein emulsion is administered to suitable experimental animal (in general being mouse or rat), come body build-in test emulsion.After using reasonable time, animal is killed, collect the tissue in the main organ and analyze.Result and control animals are compared, wherein use placebo or standardized product to control group.
Test lipid emulsion
Can adopt many clinical testings well known by persons skilled in the art, measure lipid emulsion of the present invention aspect total parenteral absorption or be used for the treatment of the effect of application.For example, study by standard biological is equivalent (referring to, for example, Current Protocols in Toxicology, Maines, M. waits (volume), J.Wiley ﹠amp; Sons, Inc., New York NY), can measure the ability that the lipid emulsion is absorbed by animal body.These research in, monitor suitable animal model infusion lipid emulsion of the present invention or the contrast emulsion (generally being the vegetable oil or the fish oil base emulsion of commercially available acquisition) afterwards, the disappearance situation of aliphatic acid from blood flow.Can also measure the fatty acid profile of liver lipid and measure distribution of fatty acids by standard procedure by the Histopathology of test example such as liver, kidney, the heart, lung, spleen or thymus gland.
The result of treatment of lipid emulsion of the present invention can obtain measuring in suitable animal disease model.For example, often use the mouse model of burning test the treatment helpfulness of lipid emulsion [referring to, Hayashi, Nutrition, 17:211 (2001)].In this case, the lipid emulsion is administered to mouse and monitors various physiologic parameters, for example, the regeneration of the nitrogen content of blood and urine or some protein such as blood albumin.
After short-term or long-term arrangement, also can carry out other biological utilisation Journal of Sex Research and toxicity test known in the art.
The purposes of lipid emulsion
Lipid emulsion of the present invention can be used as total parenteral absorption (TPN) product that is rich in cis-omega-fatty acid.For this purpose, if desired, lipid emulsion and for example plant oil based emulsion of the lipid emulsion that is rich in ω-6 can be merged, so that best ω-3: ω-6 aliphatic acid ratio is provided.In addition, can lipid emulsion of the present invention be mixed with the form of alimentation composition by standard technique, so that it contains one or more other nutrients, such as glucose, amino acid, vitamin and/or other parenteral absorption tonic.
As previously mentioned, known omega-fatty acid can influence inflammatory reaction and immunology resistance, and angiocardiopathy, fat metabolic disturbance, thrombosis process and neoplastic disease.Therefore, lipid emulsion of the present invention can also be used to be of value to patient (particularly through the patient with the anticoagulant treatment) after postoperative and the wound, experience the patient of early stage septicopyemia/general inflammatory responses syndrome (SIRS), be in the patient of high inflammatory process danger, the patient that its immunologic function need be supported, suffers from inflammatory bowel disease (Crohn ' s disease, ulcerative colitis) patient, suffers from inflammatory skin disease (psoriasis, atopy eczema) patient, the arthritic, cardiovascular patient, the cancer patient, consumptive's (as asthma) and diabetic.
Using the lipid emulsion to prepare admixture has one or more therapeutic or diagnostic compound compositions also to belong to category of the present invention.In this embodiment, the lipid emulsion plays the transmission carrier of therapeutic or diagnostic compound, and aforesaid beneficial effect is provided.Preparation lipid emulsion makes it contain one or more therapeutic or diagnostic compound, is that those skilled in the art use standard technique to realize easily.
TPN uses or the method for treatment beneficial effect is known in the art so that carry out to use the lipid emulsion to the patient.Generally, bestow emulsion by infusion in suitable time bar.The suitable dosage and the system of bestowing are that the technical staff of clinical field determines easily.
Kit
The present invention is also additional to provide the reagent that contains lipid emulsion box, is used for using to the patient.This kit can be provided at the proper dosage in the prescribed time-limit.
Kit of the present invention comprises one or more packings that contain the lipid emulsion or containers and cover teachings, and the guidance that wherein said guidance is normally written relates to the use and the dosage of lipid emulsion.Kit can also contain the additional container that contains one or more nutrients or therapeutic or diagnostic compound, and these nutrients or compound can be added in the emulsion before using.The packing that contains the lipid emulsion can be the form of UD or the form that is packing in enormous quantities (for example, multiple-unit container).The packaged form of dosage can be to make every dosage relevant with for example all interior dosage every day.Also kit and points for attention can be connected, wherein said points for attention are to be stipulated by the government organs that adjust to make, use or sell biological products, and this points for attention have reflected the permission of government organs to the manufacturing, use or the sale that are used for the human or animal and use.
For obtaining the present invention is better understood, can be with reference to following examples.Should be understood that these embodiment only enumerate for purposes of illustration.Therefore, their scopes that should not limit the present invention in any way.
Embodiment
Embodiment 1: preparation contains the lipid emulsion of harp seal oil
Phospholipid, seal oil, glycerine, tocotrienols or alpha-tocopherol and cholesterol are passed through stir about thoroughly mixing in 30 minutes at room temperature.Sterilized water is added in the mixture and with it stirred other 2-3 minute.Then, with whole mixture impouring homogenizer (EmulsiFelex
-C5, Avestin Inc., Ottawa, Canada) the chamber in and condition of high voltage (20,000-27,000psi) down for several times through this homogenizer.
In a kind of combination, seal oil 10%, egg lecithin 2.4%, tocotrienols 0.5%, glycerine 2.25% and cholesterol 5% make emulsion 20, and through behind the homogenizer 4 times, the particle mean size that fat drips is 452nm to 000Psi pressure down.27, after 000Psi increase pressure descended the process homogenizer once, grain graininess was reduced to 340nm.Yet when cholesterol was saved from combination, 20, the pressure of 000Psi can obtain the grain graininess of 280nm down.
Embodiment 2: the optimization that emulsion is formed
Because the lipid emulsion is thermodynamic unstable system, thus for preparation stable and effectively drug products is just essential considers many factors.These factors comprise character, each component concentrations and the compound method of oil and emulsifying agent.Yet it is unpractical that all factors under various content are tested, and therefore, often uses the design of factorial or part factorial.The preparation that factorial or part factorial are designed to screen various factors and optimize emulsion provides an effective way, referring to Phan-Tan-Luu, R. and Didier, M.Experimental design in emulsion andsuspension formulations:theoretical aspects.P.465-535; And Neilloud, F., Marti-Mestres, G., Maillols, H.Application ofexperimental methodology to emulsion and suspensions. P.535-557, Pharmaceutical emulsion and Suspensions.Neilloud, F. and Marti-Mestres compile, G., Marcel Dekker Inc., 2000).
What describe below is a sample of full factorial design, is used for estimating changing some factor of lipid emulsion to the sex result of 10% seal oil emulsion-stabilizing.
The excursion of three factors is shown in following table 1.
Table 1.
| Numbering | Factor | Content 1 (w/v) | Content 2 (w/v) |
| 1 | Seal oil | 10.00％ | 20.00％ |
| 2 | Egg lecithin | 1.20％ | 2.40％ |
| 3 | Cholesterol | 0.00％ | 0.50％ |
In this sample, the response of consideration is the grain graininess of emulsion.
Also estimate other parameter and the biological stability data of lipid oxidation by this method.
Full factorial design (2
3) and response be shown in following table 2.
Table 2.
| Numbering | Seal oil % | Egg lecithin % | Cholesterol % | Granularity nm |
| ?1 | ?10 | ?1.2 | ?0 | ?287±45 |
| ?2 | ?20 | ?1.2 | ?0 | ?324±48 |
| ?3 | ?10 | ?2.4 | ?0 | ?280±30 |
| ?4 | ?20 | ?2.4 | ?0 | ?290±46 |
| ?5 | ?10 | ?1.2 | ?0.5 | ?383±79 |
| ?6 | ?20 | ?1.2 | ?0.5 | ?373±57 |
| ?7 | ?10 | ?2.4 | ?0.5 | ?452±110 |
| ?8 | ?20 | ?2.4 | ?0.5 | ?363±62 |
| Matrix effect | -13 | ?4.5 | ?97.5 |
The calculating of matrix effect shows in this sample, and the existence of cholesterol and concentration are important factors for the grain graininess of decision this type emulsion.
Embodiment 3: the grain graininess of regulating the lipid emulsion
As mentioned above, the grain graininess of emulsion is relevant with many factors.Control grain graininess and this granularity remained on the stable tool to emulsion has a significant impact in certain scope, and determining the validity and the toxicity of emulsion.The emulsion of forming by following combination: seal oil 10%, egg lecithin 2.4%, tocotrienols 0.5%, glycerine 2.25% and cholesterol 5% make emulsion 20,000Psi.Through behind the homogenizer 4 times, it is 452nm that its average lipid drips granularity to pressure condition down.When pressure is increased to 27, during 000Psi, only through once, the granularity that this lipid drips drops to 340nm.Yet, if from then on economize de-cholesterol in the combination, 20, under the 000Psi pressure condition, just the grain graininess of emulsion can reach 280nm.
Embodiment 4: prepare 10% seal oil emulsion
With lecithin (12g, Lipoid E80
, Lipoid GmbH, Ludwigshafen, Germany), glycerine (22.5g, Sigma-Aldrich, Burlington, Ontario, Canada), (100g perhaps derives from Caboto Seafood Ltd. to seal oil, Baie Verte, Newfoundland and Labrador, Canada, perhaps derive from Seafreez Seafood Ltd, Catalina, Newfoundland and Labrador, Canada), vitamin E (0.2g, Sigma-Aldrich, Burlington, Ontario, Canada), 0.1M NaOH solution (2.0g, Sigma-Aldrich, Burlington, Ontario, Canada), K
2HPO
4(0.38g, Sigma-Aldrich, Burlington, Ontario, Canada), EDTA (0.75mg, Sigma-Aldrich, Burlington, Ontario, Canada) and water for injection, USP (Baxter Canada Ltd., Mississauga, Ontario, Canada) to 1000g, stir 30min.By making this mixture, prepare emulsion through high pressure homogenisers 4-8 time (EmulsiFlex-C5, Avestin, Ottawa, Ontario, Canada).Pressure is remained on 180,000kPa.With product by autoclave at 121 ℃ of following sterilization 20min and under aseptic condition, being packaged in bottle or the polybag.
The composition analysis of embodiment 5:10% seal oil emulsion
Use gas chromatography (GC) and infrared spectrophotometry, analyze the aliphatic acid of three kinds of different harp seal oil products and form, wherein said three kinds of different harp seal oil are available from different suppliers.In brief, use NaOH that seal oil is hydrolyzed into free fatty, carry out Methyl transportersization then.The fatty acid methyl ester of esterification is analyzed on the HP-5964 capillary gas chromatograph.Adopt identical process, the aliphatic acid standard sample esterification that will buy from Sigma.The aliphatic acid composition of quantitative analysis seal oil (referring to Fidler N, etc., J Lipid Res, 2000,41 (9): 1376-83).The results are summarized in following table 3.
Table 3: the composition of seal oil emulsion
| Aliphatic acid | Common name | ???? | ||
| Sample | ||||
| 1 | | | ||
| ?14:0 | Myristic acid | ????4.063 | ????4.367 | ??8.664 |
| ?14:1 | ????1.111 | ????1.034 | ??1.838 | |
| ?16:0 | Palmitic acid | ????6.406 | ????7.795 | ??9.478 |
| ?16:?1ω7 | ????17.581 | ????14.341 | ??16.632 | |
| ?18:?0 | Stearic acid | ????0.851 | ????1.249 | ??1.043 |
| ?18:1ω9 | ????22.029 | ????20.377 | ??16.88 | |
| ?18:1ω7 | ????4.555 | ????4.136 | ??3.035 | |
| ?18:2ω6 | Linoleic acid | ????1.938 | ????1.661 | ??1.277 |
| ?18:3ω6 | Gamma-Linolenic acid | ????0.226 | ????0.739 | ??n/a |
| ?18:3ω3 | Alpha-linolenic acid | ????0.722 | ????1.513 | ??0.569 |
| ?18:4ω3 | ????1.509 | ????？ | ??2.336 | |
| ?20:1ω9 | ????11.902 | ????12.403 | ??13.642 | |
| ?20:4ω6 | Arachidonic acid (AA) | ????0.544 | ????0.531 | ???n/a |
| ?20:5ω3 | Eicosapentaenoic acid (EPA) | ????8.062 | ????7.564 | ???6.985 |
| ?22:1ω11 | ????2.292 | ????3.563 | ???5.629 | |
| ?22:1ω9 | ????0.558 | ????0.699 | ???0.896 | |
| ?22:4ω6 | ????0.0903 | ????n/a | ????n/a | |
| ?22:5ω3 | Clupanodonic acid (DPA) | ????4.829 | ????5.452 | ???3.573 |
| ?22:6ω3 | DHA (DHA) | ????10.731 | ????12.575 | ???7.573 |
As can be seen from Table 3, the composition of the long-chain omega-3 polyunsaturated fatty acids in fish oil and the seal oil is very different.Though contain EPA and DHA in the fish oil, its DPA quantity is considerably less.The content of EPA is than DHA height in the fish oil.
The acid number of embodiment 6. seal oils
According to the US pharmacopeia measure acid number and with in and the mL quantity of the needed 0.1N alkali of free fatty in the 10.0g harp seal oil represent.
Oil from supplier #1: 2.356
Old harp seal oil from supplier #2: 2.262
Beater (old) oil from supplier #2: 1.695
Beater (immature) oil from supplier #2: 0.986
Bedlamer oil from supplier #2: 2.300
Beater: be in its life immature Greenland or the grey sea dog in 1 year, three to around age finished it and casted off a skin for the first time, form the soft spot grey crust that has.
Bedlamer: Greenland or grey sea dog childhood of about one to five years old, it has the crust of band spot.Harp seal is at the sexually matured distinctive saddle type (saddle) or harp (harp) the shape marking of growing gradually in early days.
Embodiment 7: content of beary metal and PCBs analyze
Carry out the ICP-mass spectral analysis of trace metal (USEPA method #3050B), mercury (USEPA method #245.5) and Polychlorinated biphenyls (PCBs) by PSC Analytical Services (Bedford, NS, Canada).
To by seal oil, fish oil (Omegaven
) and soybean oil (Intralipid
) preparation five kinds of oil samples and three kinds of lipid samples of latex test.But all samples is all less than the heavy metal and the PCBs that find detection level.
Embodiment 8: the various factors that influences seal oil emulsion quality
Granularity by oil droplet in the monitoring emulsion is estimated physical stability.Test is at acceleration environment, high speed centrifugation or carry out under hot conditions.Under 15 ℃ samples of latex (10mL) is carried out 41, the high speed centrifugation under the 600g is handled a week.Collect the aliquot (50 μ L) of emulsion every day, carry out the grain graininess analysis.In one group of experiment independently, samples of latex (10mL) remained in 37,45 and 60 ℃ the water-bath 5 days.Per 24 hours at interval, takes out the aliquot (50 μ L) of sample, carries out the grain graininess analysis.
Define chemical stability by peroxide value.Samples of latex remained in 37 ℃ the test tube.0th, collected sample and analysis peroxide value in 1,2,3 and 4 day.
With emulsion 0.2mL be added into 1.5mL isooctane/isopropyl alcohol (3: 2, v/v) in.With about 30 seconds of mixture vortex and under 2000xg centrifugal 2min.Collect top layer (20 μ L), and with 2.8mL methyl alcohol/1-butanols (2: 1, v/v) and 15 μ L rhodanate/Fe
2+Solution
*Mix.Then with the mixture vortex.At room temperature be incubated after the 20min, measure the absorbance under the 510nm.According to the calibration curve that the cumene hydroperoxide by a series of concentration known obtains, measure the concentration of peroxide.
* rhodanate/Fe
2+Solution passes through 3.94M thiocyanate salt solution and 0.072M Fe
2+Solution mixes with 1: 1 ratio and prepares.
8.1. the influence of the comparison oil droplet granularity of lecithin and seal oil
As previously mentioned, the oil droplet granularity is one of feature of emulsion.We find that the surfactant (lecithin) that uses is influential to the granularity of formed oil droplet.As shown in Figure 1, along with the ratio of lecithin with oil reduces, the oil droplet granularity increases.
8.2.pH influence to surface charge
Electric charge on the known oil droplets has influence to the stability of emulsion.High electric charge repels oil droplet each other.As a result, oil droplet can stop the long period in emulsion.The use zeta potential defines the electric charge on the oil droplets.Measure the seal oil emulsion under condition of different pH zeta potential and the results are shown in Fig. 2.This result confirms: along with the pH increase of emulsion, zeta potential increases.
8.3.pH to snperoxiaized influence
Contain polyunsaturated fatty acid in the seal oil, they are easy to peroxidating.Oxidation of fat acid is relevant with cancer, angiocardiopathy and inflammatory process.For the oil that contains unrighted acid, primary concern be the peroxidating problem of aliphatic acid.The degree of known peroxidization and/or speed are subjected to the influence of many factors, comprise pH.Estimate pH to the influence of the peroxide of seal oil emulsion and the results are shown in Fig. 3.The result shows: when the pH of emulsion was～7 or pH＜3, peroxidization was slow.More peroxidization takes place when pH is between 4-6.
Based on The above results, selecting PB to control emulsion pH is 7.Used K
2HPO
4Concentration be 2.5mM.Under this pH, find that zeta potential is in the 35-42mV scope.Though the PB of higher concentration can increase buffer capacity, it also can increase the ionic strength of solution, thereby can cause the quick reduction of zeta potential, and this does not expect.
8.4 antioxidant and chelating agent are to the influence of peroxide
Fig. 4 has shown when vitamin E (as antioxidant), EDTA and vitamin E/EDTA do not exist together or exist, the peroxidating result of seal oil emulsion.The result shows that EDTA and vitamin E all can reduce peroxidization significantly.The combination of EDTA and vitamin E is very effective.According to reports, the transition metal ions such as the Cu that have trace in the lecithin (from egg or soybean)
2+And Fe
3+Known these metal ions are superior catalyst to oxidation reaction.It is believed that the chelating agent such as EDTA can form complex compound with metal ion.As a result, the reaction of ion institute catalysis is suppressed.Yet, in the product of waiting to be infused in patient's blood plasma, exist a large amount of EDTA not expect.For finding out the MEC of EDTA in seal oil lipid emulsion, the EDTA of the different amounts of test.It is found that, when the concentration of EDTA is higher than 2 * 10
-6During M, peroxidization is eliminated (Fig. 5).Therefore, all seal oil latex products are selected this concentration.The concentration of vitamin E is 0.02%, itself and Fish Oil Emulsion (Omegaven
) numerical value be identical.
The sign of embodiment 9:10% seal oil emulsion
9.1. grain graininess and zeta potential
The homogeneous granules granularity is the ideal characterisitics of emulsion.Known zeta potential influences the stability of emulsion.Therefore, estimate the grain graininess and the zeta potential of the seal oil emulsion that adopts different component and under different condition, prepare.
(Beckman Coulter makes by grain analyser Delsa 440SX, Fullerton, California, USA), analysis finds that according to the grain graininess and the zeta potential of 10% seal oil emulsion of embodiment 4 preparations the result is respectively 320 ± 40nm and 44.7 ± 15.3mV.The pH of emulsion measures to find it is 7.5 ± 0.5 by pH meter.
9.2. aseptic test
Medical Microbiological Lab (St.John ' s, Newfoundland and Labrador, Canada) by Health Care Corporation checks aseptic.Aseptic evidence, product do not contain any germ contamination.Table 4 has gathered the seal oil of different batches and has used the aseptic check result of these seal oil products according to 10% seal oil emulsion of embodiment 4 preparations.
Table 4: aseptic test
| The description of seal oil | Specimen | The autoclaving time (minute) | Aseptic result's (+expression has bacterium) |
| Under 70 ℃, from the sea dog at unknown age, extract, do not have the chromatogram deodorization simultaneously, derive from supplier #1 | The seal oil that is obtained | ????0 | ????- |
| 10% seal oil emulsion | ????0 | ????- | |
| 10% seal oil emulsion | ????10 | ????- | |
| 10% seal oil emulsion | ????20 | ????- | |
| At room temperature extract from very old harp seal, the chromatogram deodorization derives from supplier #2 simultaneously | The seal oil that is obtained | ????0 | ????+ |
| 10% seal oil emulsion | ????0 | ????- | |
| 10% seal oil emulsion | ????10 | ????- | |
| 10% seal oil emulsion | ????20 | ????- | |
| At room temperature extract from one-year-old harp seal, the chromatogram deodorization derives from supplier #2 simultaneously | The seal oil that is obtained | ????0 | ????+ |
| 10% seal oil emulsion | ????0 | ????- | |
| 10% seal oil emulsion | ????10 | ????- | |
| 10% seal oil emulsion | ????20 | ????- | |
| At room temperature extract from Beaters, the chromatogram deodorization derives from supplier #2 simultaneously | The seal oil that is obtained | ????0 | ????+ |
| 10% seal oil emulsion | ????0 | ????- | |
| 10% seal oil emulsion | ????10 | ????- | |
| 10% seal oil emulsion | ????20 | ????- | |
| At room temperature extract from Bedlamers, the chromatogram deodorization derives from supplier #2 simultaneously | The seal oil that is obtained | ????0 | ????+ |
| 10% seal oil emulsion | ????0 | ????- | |
| 10% seal oil emulsion | ????10 | ????- | |
| 10% seal oil emulsion | ????20 | ????- |
9.3. endotoxin test: LALT
(Cape Cod, Massachusetts USA) buy endotoxin standard sample (0.2ng/mL) and LAL kit from Associates of Cape Cod Incorporated.Injectable sterile water USP (10mL/vial, Baxter Canada, Mississauga, Ontario, Canada) is contrasted as negative.Endotoxin standard sample (0.2ng/mL) is diluted to 0.1,0.05,0.025 and 0.0125ng/mL.The 10% seal oil emulsion of embodiment 4 also is diluted to 2,4,8,16 and 32 times.For testing, the negative contrast of 0.1mL, endotoxin standard sample (0.2,0.1,0.05,0.025 and 0.0125ng/mL) or seal oil emulsion and dilute sample thereof are added in the LAL kit of 0.1mL.Sample is shaken mixing fully.Sample is incubated 1 hour down and detection at 37 ± 1 ℃.Indicate endotoxic existence by the formation of gel.
External endotoxin test (LAL chemical examination) shows that the seal oil emulsion is non-pyrogenicity (＜0.006ng/mL or 10EU/ng).
Embodiment 10: the chemical stability of seal oil, vegetable oil and Fish Oil Emulsion relatively
According to the procedural test 10% seal oil emulsion (pressing embodiment 4 described preparations) of embodiment 8 descriptions, the vegetable oil emulsion (Intralipi of commercially available acquisition
) and the Fish Oil Emulsion (Omegaven of commercially available acquisition
) chemical stability (being defined as the peroxidating degree) and compare.The results are shown in Fig. 6, the result proves: the seal oil emulsion is all more stable than vegetable oil or Fish Oil Emulsion.Intralipid
And Omegaven
It all is the trade name of Fresenius-Kabi.
Embodiment 11: autoclaving is handled the physics of seal oil emulsion and the influence of chemical stability
Because the seal oil emulsion formulations desires to be used for intravenous infusion, therefore need carry out autoclaving and handle, so that deactivation bacterium and other pathogen.Because the polyunsaturated fatty acid of seal oil is to high temperature and pressure transient, thereby sample behind the autoclaving of emulsion carried out chemistry and reexamining of physical stability is necessary.Five kinds of seal oil latex products (20mL separately, seal oil derive from the sea dog of all ages and classes and by different technologies processing, and by embodiment 4 described preparation emulsions) were handled 0,10,20,30,60 minute under 121 ℃ autoclaving condition.The speed of the grain graininess of test sample, zeta potential and peroxidization (Fig. 7,8,9 and 10).Presentation of results, the parameter of measuring in all samples does not have marked change.According to the US pharmacopeia, the autoclaving volume is that the injectable goods of 500mL need 30 minutes.Used autoclaving condition has clearly proved the stability of seal oil emulsion formulations in these experiments.
Embodiment 12: the comparison of seal oil and Fish Oil Emulsion
Relatively press the 10% seal oil emulsion of embodiment 4 described preparations and the Fish Oil Emulsion (Omegaven of commercially available acquisition
) performance.
Table 5: preparation relatively
| Composition (per 100 grams) | Omegaven (10% Fish Oil Emulsion) | 10% seal oil emulsion |
| Total oil | 10g | ?10g |
| ?EPA(C20：5ω-3) | 1.25-2.82g | ?0.70-0.81g |
| ?DPA(C22：5ω-3) | N/A | ?0.34-0.55g |
| ?DHA(C22：6ω-3) | 1.44-3.09g | ?0.75-1.3g |
| Lecithin | 1.2g | ?1.2g |
| Glycerine | 2.5g | ?2.5g |
| Vitamin E | 0.015-0.0295g | ?0.02g |
| ?EDTA- | 0 | ?0.007g |
Table 6:Omegaven
Compare with the feature of 10% seal oil emulsion
| Feature | ?Omegaven (10% Fish Oil Emulsion) | 10% seal oil emulsion |
| Grain graininess (nm) | ?330±24 | ?320±40 |
| Zeta potential (mV) | ?48.3±17.2 | ?44.7±15.3 |
| pH | ?7.5-8.7 | ?7.0-8.0 |
| Endotoxin test (LAL) | ?- | ??- |
| Aseptic | ?- | ??- |
| Permeability (mosm/kg) | ?308-376 | 320 ± 20 (calculating) |
| Peroxide value | ?0.52 | ?0.14 |
| Shelf-life | Made back 1 year | Made back 2 years |
Embodiment 13: acute toxic test
Use the SD rat and rat is divided into two groups, every group of three animals.In 10 days time limit, give animal infusion seal oil or 1 hour every day of Fish Oil Emulsion by tail vein.The dosage of seal oil or Fish Oil Emulsion compares Omegaven
RD every day (0.1g/kg/ days) high 10 times (1g/kg body weight/day).Control group is handled with saline solution.After 10 days the infusion, with every animal kill and collect the tissue that comprises kidney, the heart and liver, (St.John ' s) carries out pathological examination with these fixation of tissue and the Pathology Lab by Health Care Corporation.(Omegaven contrasts, feeds in a organized way for kidney, liver, GI road and institute in the heart
With the seal oil emulsion) all similar, there is not the liver deposition of lipids.In rat model, do not observe acute toxicity.
Invention has been described, and obviously same problem can change by a lot of modes.These changes do not think to have broken away from the spirit and scope of the invention, and used these conspicuous to those skilled in the art improvement are included in the scope of following claim.
Claims (23)
1, a kind of lipid emulsion, this emulsion contains:
(d) the purifying seal oil of 5-40wt%;
(e) emulsifying agent of 0.5-5wt% and
(f) water,
The final pH of wherein said emulsion is 6.0-9.0.
2,, also contain the permeability agent of 2-5wt% according to the lipid emulsion of claim 1.
3, according to the lipid emulsion of claim 2, wherein said permeability agent is a glycerine.
4,, also contain the antioxidant of 0.002-1.0% according to claim 1,2 or 3 each lipid emulsions.
5, according to claim 1,2,3 or 4 each lipid emulsions, also contain 1 * 10
-6M to 5 * 10
-5The EDTA of M.
6, according to claim 1,2,3,4 or 5 each lipid emulsions, wherein said emulsifying agent is a lecithin.
7, a kind of alimentation composition, said composition contains:
(c) contain the emulsifying agent of purifying seal oil, 0.5-5wt% of 5-40wt% and water the lipid emulsion and
(d) one or more nutrient compounds,
The final pH of wherein said composition is 6.0-9.0.
8,, also contain the permeability agent of 2-5wt% according to the alimentation composition of claim 7.
9, alimentation composition according to Claim 8, wherein said permeability agent is a glycerine.
10, claim 7,8 or 9 each alimentation compositions also contain the antioxidant of 0.002-1.0%.
11, according to claim 7,8,9 or 10 each alimentation compositions, also contain 1 * 10
-6M to 5 * 10
-5The EDTA of M.
12, according to claim 7,8,9,10 or 11 alimentation composition, wherein said emulsifying agent is a lecithin.
13, a kind of contain claim 1,2,3,4,5 or 6 each the lipid emulsion and the composition of one or more therapeutic compounds.
14, a kind of claim 1,2,3,4,5 or 6 each lipid emulsion and one or more diagnostic compound compositions of containing.
15, claim 1,2,3,4,5 or 6 each lipid emulsions are used to prepare the purposes of alimentation composition.
16, claim 1,2,3,4,5 or 6 each lipid emulsions are used to prepare the purposes of therapeutic composition, and wherein said therapeutic composition is used for one or more therapeutic compounds are passed to the patient of needs.
17, claim 1,2,3,4,5 or 6 each lipid emulsions are used to prepare the purposes of diagnostic compositions, and wherein said diagnostic compositions is used for one or more diagnostic compounds are passed to the patient of needs.
18, claim 1,2,3,4,5 or 6 each the purposes of lipid emulsion in total parenteral absorption of the patient of needs.
19, according to the purposes of claim 18, wherein prepare described emulsion, with the second kind of emulsion combined administration that contains vegetable oil.
20, claim 7,8,9,10,11 or 12 each the purposes of alimentation composition in total parenteral absorption of the patient of needs.
21, according to the purposes of claim 20, wherein prepare described composition, with the second kind of emulsion combined administration that contains vegetable oil.
22, the composition of claim 13 is used for one or more therapeutic compound parenterals are passed to the patient's of needs purposes.
23, the composition of claim 14 is used for one or more diagnostic compound parenterals are passed to the patient's of needs purposes.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CA2366166 | 2001-12-27 | ||
| CA2,366,166 | 2001-12-27 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN1620253A true CN1620253A (en) | 2005-05-25 |
Family
ID=4170969
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNA028280695A Pending CN1620253A (en) | 2001-12-27 | 2002-12-23 | Seal oil based lipid emulsions and uses thereof |
Country Status (3)
| Country | Link |
|---|---|
| CN (1) | CN1620253A (en) |
| AU (1) | AU2002350353A1 (en) |
| WO (1) | WO2003055323A1 (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101822839B (en) * | 2010-03-31 | 2012-01-04 | 首都医科大学 | Preparation of nanometer structure lipid carrier by using seal oil as liquid-phase substrate and application |
| CN102579338A (en) * | 2012-03-02 | 2012-07-18 | 首都医科大学 | Preparation method and application of paclitaxel intravenous fat emulsion |
| CN105147609A (en) * | 2015-10-28 | 2015-12-16 | 辅必成（上海）医药科技有限公司 | Diethylstilbestrol fat emulsion injection |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| LU86601A1 (en) * | 1986-09-22 | 1988-04-05 | Oreal | PHOTOSTABLE COSMETIC COMPOSITION CONTAINING AN ETHYLRUTINE DERIVATIVE AS A PROTECTIVE AGENT AGAINST SALARY LIGHT AND ITS USE FOR PROTECTING THE SKIN AND HAIR |
| PL186618B1 (en) * | 1995-11-28 | 2004-02-27 | Braun Melsungen Ag | Lipidic compositions optimised in respect to hydrolysis and their application |
| EP1042001B1 (en) * | 1997-12-16 | 2002-04-03 | Chiron Corporation | Use of microparticles combined with submicron oil-in-water emulsions |
| CN1227731A (en) * | 1998-03-02 | 1999-09-08 | 孙淑磊 | Method for producing ham sausage containing omega-3 unsaturated fatty acid |
| CA2260397A1 (en) * | 1999-01-29 | 2000-07-29 | Atlantis Marine Inc. | Method of converting rendered triglyceride oil from marine sources into bland, stable food oil |
| PT102509A (en) * | 2000-08-16 | 2002-02-28 | Fundacao Essprit Icarus | PHARMACEUTICAL, COSMETIC AND DERMO-COSMETIC COMPOSITIONS, OF HYGIENE, FOOD AND FODDER, BASED ON FOCUS OIL, AND THEIR USE |
-
2002
- 2002-12-23 CN CNA028280695A patent/CN1620253A/en active Pending
- 2002-12-23 AU AU2002350353A patent/AU2002350353A1/en not_active Abandoned
- 2002-12-23 WO PCT/CA2002/001981 patent/WO2003055323A1/en not_active Application Discontinuation
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101822839B (en) * | 2010-03-31 | 2012-01-04 | 首都医科大学 | Preparation of nanometer structure lipid carrier by using seal oil as liquid-phase substrate and application |
| CN102579338A (en) * | 2012-03-02 | 2012-07-18 | 首都医科大学 | Preparation method and application of paclitaxel intravenous fat emulsion |
| CN105147609A (en) * | 2015-10-28 | 2015-12-16 | 辅必成（上海）医药科技有限公司 | Diethylstilbestrol fat emulsion injection |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2003055323A1 (en) | 2003-07-10 |
| AU2002350353A1 (en) | 2003-07-15 |
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| Date | Code | Title | Description |
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| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| EE01 | Entry into force of recordation of patent licensing contract |
Assignee: China vast Pharmaceutical Co Ltd Assignor: Norht Atlantic Biopharma lnc.,licensed from GENESIS|Group lnc.of Memorial University of Newfoundland Contract fulfillment period: From September 28, 2004 to December 23, 2020 Contract record no.: 051000030035 Denomination of invention: Seal oil based lipid emulsions and uses thereof License type: exclusive license Record date: 20050309 |
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| LIC | Patent licence contract for exploitation submitted for record |
Free format text: EXCLUSIVE LICENCE; TIME LIMIT OF IMPLEMENTING CONTACT: 2004.9.28 TO 2020.12.23 Name of requester: CHINA GUANGZENG MEDICINE CO., LTD. Effective date: 20050309 |
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| C12 | Rejection of a patent application after its publication | ||
| RJ01 | Rejection of invention patent application after publication |
Open date: 20050525 |