WO1998016233A1

WO1998016233A1 - Formulations containing phosphatidylethanolamine

Info

Publication number: WO1998016233A1
Application number: PCT/GB1997/002756
Authority: WO
Inventors: David Frederick Horrobin; Bengt Herslof
Original assignee: Scotia Holdings Plc
Priority date: 1996-10-11
Filing date: 1997-10-08
Publication date: 1998-04-23
Also published as: GB9621293D0; AU4567997A; ZA979054B

Abstract

Phosphatidylethanolamine in the form of a preparation derived from egg yolk and containing more than 20 % by weight of the phosphatidylethanolamine, preferably more than 40 %, very preferably more than 60 %, and most preferably more than 70 %.

Description

FORMULATIONS CONTAINING PHOSPHATIDYLETHANOLAMINE

Field of Invention

The invention relates to formulations containing unsaturated fatty acids.

Background

In previous patent applications and publications, we and others have claimed the use of preparations containing arachidonic acid (AA) and/or docosahexaenoic acid (DHA) for various therapeutic and nutritional purposes. These highly unsaturated essential fatty acids may be used for a wide variety of conditions, but particularly for psychiatric and other central nervous system and eye disorders; for cardiovascular and metabolic disorders: and for inflammatory disorders.

The psychiatric, eye and central nervous system disorders include schizophrenia, depression, multiple sclerosis. Huntington's disease, dementias including Alzheimer's disease, and eye diseases such as retinitis pigmentosa and senile macular degeneration. The cardiovascular and metabolic, disorders include peripheral, cerebral and coronary arterial disease, hypertension, elevated blood cholesterol and triglycerol levels, low HDL cholesterol levels, diabetes and "syndrome X" a syndrome of insulin resistance often associated with hypertension and obesity. The inflammatory and auto-immune diseases include rheumatoid arthritis, osteoarthritis. asthma, inflammatory bowel disease and a range of other conditions. Under appropriate circumstances other diseases may also be treated. AA and DHA may also be used in foods or as nutrients, food additives or nutritional supplements since AA and DHA are essential constituents of all the cell membranes in the human and animal bodv. The Invention

We have sought improved ways of administering AA and DHA and broadly we have found that using phosphatidylethanolamine CPE) isolated from egg yolks is a particularly convenient way of doing so. Among other aspects therefore, as claimed herein, the invention lies in pharmaceutical and nutritional uses of a product of egg yolk enriched so that it contains more than 20% by weight of PE. preferably more than 40%. very preferably more than 60%. and most preferably more than 70%.

Egg yolk is a good source of AA and DHA but these fatty acids are found in only relatively low concentrations in the whole egg yolk. We have found that much of the AA and DHA is in fact found in the PE fraction of the egg yolk phospholipids. that this PE fraction can be isolated on a large scale and that it can be formulated and used either in pharmaceutical dosage forms or in foods or nutritional supplements to raise the concentration of AA and DHA at appropriate sites.

PE prepared in pure form from egg yolks by the techniques described later or by any other suitable technique is a phospholipid with an ethanolamine head group in the Sn3 position and two fatty acids in the Sn2 and Snl positions. Especially in the Sn2 position these fatty acids in PE are high in AA and DHA. Depending on the precise source of eggs, between 5 and 30% of the total fatty acids may be AA and between 5 and 30% may be DHA. Usually these values are in the 10-20% range.

We have found that this PE is a powder which can be formulated as such or as a tablet or hard gelatin capsules, and which can also be dissolved in appropriate vegetable oils to give a PE solution in triglycerides. Such liquids can be encapsulated, incorporated into products for skin care on the treatment of skin diseases, used to form emulsions of various types or incorporated into any appropriate dosage form known to those skilled in the an. Test Work

We have administered pure egg yolk PE formulated as a 20% solution in corn oil to normal, chow fed rats and have demonstrated the efficiency of the preparation in raising the plasma, red cell membrane, liver, heart, and other tissue concentrations of AA and DHA. The material is therefore readily assimilated into the body and provides an excellent deliven' vehicle for AA and DHA. Moreover, the PE has also been used in tests to see whether it can restore certain aspects of normal brain function in old mice. These tests have shown that the PE is effective in improving brain function, so demonstrating that it is an effective system for producing pharmacological effects in the brain. For example, in 4 month old mice, membrane AA concentration in the of the brain was 20.7 nmol/mg ±1.5. whereas in 22 month old mice the concentration had fallen significantly to 16.6 nmol/mg. On feeding the PE preparation to the old mice at a dose which provided lOmg of AA/day/mouse. the hippocampal AA concentration rose to 22.9 x 12.1 nmol/mg. In addition this biochemical change was associated with a functional change. One of the most important neurotransmitters in the brain is glutamate which can be released from nerve cells by exposing them to calcium. Isolated hippocampal nerve endings from 4 month old mice show a surge of glutamate release when exposed to calcium. In contrast, the AA depleted membranes from 22 month old mice show- no such effect. However, the nerve endings from 22 month old mice which had been treated with PE showed normal glutamate release in response to calcium.

Isolation of Lipids

PE can be isolated from egg yolk, preferably egg yolk from hens' eggs. Spray-dried egg yolk powder can be purchased on the open market. This spray-dried egg yolk powder is extracted with organic solvents, using any appropriate method but preferably a mixture of hexane and isopropanol. The polar lipids. mainly PE and phosphatidylcholine CPC). will dissolve in the solvent and the non-soluble matter can be removed by filtration. The extract solution is then applied to a chromatographic column with an appropriate stationary phase known to those skilled in the art. such as silica gel. The polar lipids will be absorbed by the stationary phase. The column is then washed with solvent mixture of the same proportion as the extraction mixture which will remove all of the non-absorbed lipids. The absorbed lipids can then be eluted from the column by a step-wise gradient procedure based on mixtures of hexane. isopropanol and water. This separates the PC and the PE into separate fractions. The eluted fraction containing PE is collected and the solvent mixture is removed from the PE by evaporation and freeze-drying. There are many possible variations of this isolation procedure but the value of the product is not affected by the precise nature of the procedure used.

In a typical example 150 kg of egg yolk powder is mixed with 300 litres of hexane/isopropanol (85/15 vol/vol). The slurry is filtered and the filter cake is washed with the solvent mixture to a total volume of 450 litres extract. This volume is transferred onto a column containing 85 kg of silica gel to absorb the polar lipids. 600 litres of hexane/isopropanol/water (60/36/4 vol/vol/vol) are then pumped through the column and the fraction containing PE is collected (340 litres). The fraction is evaporated and freeze-dried to yield 2.5 kg of > 95 % PE by weight (as determined by HPLC).

The PE may be used in a pure form or in a form which contains a mixture of egg yolk phospholipids containing not less than 20% of PE. This PE or PE-enriched product may then be employed to make pharmaceutical or nutritional or skin care preparations which may be administered by oral, enteral. parenteral or topical routes. The PE doses used may range from lmg to 200g/day, preferably lOmg and 20g more preferably 50mg to 5g/day.

Examples

Examples of preparations suitable for administration in the above amounts for purposes described herein are:- 1. Hard gelatin capsules, each containing 20- 1 OOOmg of PE in the form of one of the following: a) The above freeze dried product containing from 20% to 99.9% PE, b) The freeze dried product carried by an appropriate vegetable oil carrier such as corn oil, soya oil, sunflower oil, safflower oil or evening primrose oil or a marine oil such as sardine or tuna oil.

2. Oils carrying PE as in example lb but formulated as an emulsion for oral administration. The emulsion may contain 5 to 50%> of the oil emulsified with emulsifying agents known to those skilled in the art including natural, synthetic and semi-synthetic agents including galactolipids.

3. Emulsions as in 2 but sterilised and formulated for intraveneous administration.

4. Oils carrying PE as in example lb, sterilised and formulated for intrasmuscular or sub-cutaneous injection.

5. Oils carrying PE as in example lb formulated for skin care or topical adminsitration using patch or other technologies.

Claims

1. Phosphatidylethanolamine in the form of a preparation derived from egg yolk and containing more than 20%> by weight of the phosphatidylethanolamine, preferably more than 40%), very preferably more than 60% and most preferably more than 70%.

2. A pharmaceutical or skin care composition comprising as a source of AA and DHA phosphatidylethanolamine in the form claimed in claim 1.

3. A food or nutritional supplement comprising as a source of AA and DHA phosphatidylethanolamine in the form claimed in claim 1.

4. When for use in therapy, phosphatidylethanolamine in the form claimed in claim 1.

5. The use of phosphatidylethanolamine in the form claimed in claim 1 , for the preparation of a medicament for treatment of eye, central nervous system or psychiatric disorders; cardiovascular and cardiovascular metabolic disorders; inflammatory and autoimmune disorders.

6. Treatment of the disorders of claim 5, by administration of phosphatidylethanolamine in the form claimed in claim 1.

7. The subject of claim 2, 3, 4, 5 or 6, for administration of the phosphatidylethanolamine in daily amounts of lmg to 200g preferably lOmg to 20g more preferably 50mg to 5g.