Dietary formulation comprising arachidonic acid and methods of use

A novel formulation which contains about 2-60% by calories of a C.sub.20 or longer .omega.3 fatty acid moiety and about 2-60% by calories of an arachidonic acid moiety has been developed. This formulation is particularly useful for restoring arachidonic acid levels, increasing immunity or minimizing risk of infection in critically ill patients, and treating functional essential fatty acid deficiency in patients suffering from end stage liver disease. A structured lipid which has a C.sub.20 or longer .omega.3 fatty acid moiety and an arachidonic acid moiety has also been developed.

BACKGROUND OF THE INVENTION 
Protein calorie malnutrition is a common complicating condition in patients 
with alcoholic chronic liver disease (Mendenhall et al. Am J Med 
1984;76:211-222; Mendenhall et al. Am J Clin Nutr 1986;43:213-218) and 
non-alcoholic chronic liver disease (O'Keefe et al. Lancet 1980;2:615-617; 
Morgan et al. Gut 1976;17:113-118). Factors contributing to the high 
prevalence include poor dietary intake, elevated resting energy 
expenditure, and nutrient malabsorption (McCullough A J and Tavill A S. 
Seminars in Liver Disease 1991;11:265-277). Patients with end stage liver 
disease complicated by portal hypertension are particularly likely to be 
malnourished and, when hospitalized, frequently require active nutritional 
therapy. While the effects of malnutrition in chronic liver disease on 
fatty acid nutrition have not been extensively studied, because of an 
increased resting energy expenditure, fat malabsorption and abnormal fat 
catabolism, these patients may have significant abnormalities in fatty 
acid metabolism (Cabre et al. Am J Gastroent 1988;83:712-717; Palombo et 
al. Gastroenterology 1987;93: 1170-1177). One potential mechanism for such 
a disturbance would be an inadequate intake of essential fatty acids as 
part of the global protein calorie malnutrition. 
Dietary fatty acids are classified according to their chain length. Long 
chain fatty acids contain 14 carbons or greater and can be further 
characterized by the number of double bonds contained in their structure 
into saturated, monounsaturated and polyunsaturated subgroups. The two 
fatty acids essential in human nutrition are linoleic acid and 
alpha-linolenic acid from which polyunsaturated fatty acids of the omega 6 
series and omega 3 series are formed through enzymatic desaturation and 
elongation by the liver. The body cannot convert omega 3 fatty acids to 
omega 6 fatty acids or vice versa. Patients with advanced liver disease 
may have an impaired ability to form polyunsaturated fatty acids, 
including arachidonic and eicosapentaenoic acid, from their essential 
fatty acid precursors, potentially altering membrane composition and 
eicosanoid production. 
It has now been demonstrated that there is a defect in elongation and 
desaturation in end stage liver disease, resulting in dramatically reduced 
levels of the polyunsaturated fatty acids such as arachidonic, 
eicosapentaenoic, and docosahexaenoic acid. Although there may be some 
degree of concurrent linoleic acid deficiency, the ratio of 
linoleic/arachidonate confirms that elongation and desaturation is the 
limiting factor. Arachidonic acid levels in tissue phospholipids are 
usually very tightly regulated, such that arachidonic acid levels are 
stable at varying levels of linoleic acid content in the diet. Only with 
essential (linoleic) fatty acid deficiency or with the consumption of 
omega 3 fatty acids in place of omega 6 fatty acids do arachidonic acid 
levels fall. Given the importance of arachidonic acid in second messenger 
metabolism, it is reasonable to suspect that some of the untoward side 
effects of end stage liver disease, including decreased immunocompetence 
and increased risk of infection and infectious mortality, may be a 
consequence of functional essential fatty acid deficiency. In the normal 
diet arachidonic acid is very low, because it is only found in any 
quantity in the membranes of animal flesh. To add arachidonic acid alone 
to a diet formulation is problematic, since arachidonic acid 
supplementation in normal individuals is pro-inflammatory and 
immunosuppressive. In the alternative, to give eicosapentaenoic acid (EPA) 
and docosahexaenoic acid (DHA), which provide benefits regarding 
immunosuppression, would likely lead to a worsening of arachidonic acid 
levels, and potentially essential fatty acid deficiency. 
Thus a need still exists to develop a novel dietary formulation which can 
normalize membrane composition to accomplish two goals: (1) restore 
arachidonic acid levels to normal and reverse signs related to essential 
fatty acid deficiency, and (2) provide added immune-enhancing benefits of 
EPA and DHA. 
Accordingly, an object of the invention is to provide a novel formulation 
which can restore arachidonic acid levels and which can increase immunity 
of a subject. 
Another object of the invention is to provide a method of restoring 
arachidonic acid levels in a subject by administering the novel 
formulation of the invention. 
Another object of the invention is to provide a method of increasing 
immunity or minimizing a risk of infection in a subject by administering 
the novel formulation of the invention. 
Another object of the invention is to provide a method of treating 
essential fatty acid deficiency in a subject by administering the novel 
formulation of the invention. 
A still further object of the invention is to provide a structured lipid 
which provides a high energy fat source and fatty acids which assist in 
fighting infection and treating essential fatty acid deficiency. 
These and other objects and features of the invention will be apparent from 
the following description and from the claims. 
SUMMARY OF THE INVENTION 
The present invention features a novel formulation which includes about 
2-60% of the caloric requirements of a patient with a C.sub.20 or longer 
.omega.3 fatty acid moiety and about 2-60% of the caloric requirements 
which provides an arachidonic acid moiety. Preferably, the formulation 
includes about 10-20% by calories of an arachidonic acid moiety highly 
enriched in arachidonic acid (i.e., 50%) and about 20% by calories of a 
formula containing substantial amounts of a C.sub.20 or longer .omega.3 
fatty acid moiety (i.e., certain fish oils). The .omega.3 fatty acid 
moiety is selected from the group consisting of an eicosapentaenoic (EPA) 
acid moiety, a docosahexaenoic (DHA) acid moiety, and mixtures thereof. 
The formulation should include a source of carbohydrate. The source of 
carbohydrates can be any simple or complex carbohydrate, e.g., 
monosaccharides, disaccharides, or oligosaccharides. Examples of preferred 
carboxydrates include but are not limited to corn starch, dextrose and 
glucose. 
The formulation further should include a source of protein. The source of 
protein can be any protein hydrolysate or peptide mixtures of high 
biologic values, e.g., meat or soy proteins. The protein hydrolysate are 
preferably partially hydrolyzed in nature and include a substantial 
fraction of variable chain length peptides, e.g., medium or short chain 
peptides, e.g., di- and tri-peptides, but has less than about 10% free 
amino acids, more preferably less than about 5% free amino acids. In a 
preferred embodiment, only the highest biological value proteins are 
hydrolyzed, e.g., whey, lactalbumin, casein, egg white, egg solids, soy, 
or delactosed milk solids. In other preferred embodiments, the protein 
source is lactose-free, and free amino acids are preferably avoided in the 
formulation of the invention. 
The invention also features formulations which include, in addition to the 
components described above, vitamins and minerals in accordance with, or 
approximately, the Recommended Dietary Allowance (RDA), now called the 
Reference Daily Intake (RDI). The formulation of the invention can also 
contain nutrients not recommended by the RDA, e.g., nucleotides, 
beta-carotene, carnitine, and taurine. 
The formulation may also include, in addition to the components described 
above, inactive ingredients such as emulsifiers, artificial sweeteners 
and/or flavoring. Any formulation preferably includes essential amino 
acids (albeit not in free form), essential fatty acids, essential and 
non-essential vitamins and minerals. The formulations of the present 
invention may be in the form of a dietary supplement or used as a total 
enteral or parenteral feeding regimen. If the latter, these essential 
nutrients are required while even in a supplement, the addition insures 
that the patient is obtaining these nutrients. When the formulation is in 
the form of a dietary supplement, the formulation should provide about 
5-60% of total energy expenditure in terms of calories. 
The invention also features a method of restoring arachidonic acid levels 
in a subject. The method includes administering to the subject the 
formulation of the invention which includes about 2-60% by calories of a 
C.sub.20 or longer .omega.3 fatty acid moiety and about 2-60% by calories 
of an arachidonic acid moiety. This method is particularly useful in 
treating subjects suffering from chronic liver disease. 
In another aspect, the invention features a method of increasing immunity 
or minimizing a risk of infection in a subject. The method includes 
administering to the subject the formulation of the invention. The method 
of the invention is particularly useful for patients who are critically 
ill for a variety of reasons including surgery, burns, trauma, cancer, 
AIDS, multistem organ failure, sepsis or inflammatory process which can 
also impair fatty acid elongation and desaturation. It is also useful for 
individuals who may have an infection at the time of the administration of 
the diet or may be at high risk of infection due to some immunocompromise. 
Individuals at risk of infection include those suffering with secondary 
immunosuppression due to chemotherapy or diabetes mellitus, 
protein-malnourished patients, or patients undergoing surgery, e.g., 
abdominal or thoracic surgery. 
In still another aspect, the invention features a method of treating 
essential fatty acid deficiency in a subject. The method includes 
administering to the subject the described formulation. Such treatment is 
beneficial, for example, for improvement in growth, nitrogen balance, 
vision (where DHA is critical), higher cerebral function, red and white 
blood cell and platelet function and sodium and fluid balance. 
The invention also features a structured lipid which provides a high energy 
fat source and fatty acids which assist in fighting infection and treating 
essential fatty acid deficiency. The structured lipid has a glycerol 
backbone with three fatty acids linked thereto. Preferably, the structured 
lipid includes at least .omega.3 C.sub.20 or longer .omega.3 fatty acid 
moiety and at least one arachidonic acid moiety. In preferred embodiments, 
the fatty acid moiety or the arachidonic acid moiety is at position 
R.sub.2. The structured lipid further includes a C.sub.8 -C.sub.12 fatty 
acid moiety, preferably a C.sub.12 fatty acid moiety. In one preferred 
embodiment, the C.sub.8 -C.sub.12 fatty acid moiety is at position R.sub.1 
or R.sub.3 and the arachidonic acid C.sub.12 at R.sub.2 while in others 
the C.sub.8 -C.sub.12 fatty acid moiety is at position R.sub.2. 
The following description and non-limiting examples further elucidate the 
invention. 
DETAILED DESCRIPTION OF THE INVENTION 
Patients with chronic liver disease have significant abnormalities in fatty 
acid metabolism which can lead to essential fatty acid deficiency. 
Hospitalized patients with chronic liver disease have significantly lower 
omega 3 and omega 6 essential fatty acids and their metabolites compared 
to control patients. There is evidence of both an acute deficiency as 
measured by the fatty acid composition of plasma triglyceride and chronic 
deficiency as reflected by the phospholipid fatty acids. Administering the 
formulation of the invention to these patients would be beneficial both in 
restoring arachidonic acid levels to normal and reversing signs related to 
essential fatty acid deficiency, and providing added immune-enhancing 
benefits of EPA and DHA. As the result of the administration of the 
formulation of the invention, essential fatty acid deficiency in tissue is 
reduced and serum and tissue lipid levels increase. 
The formulation of the invention is made by blending the fat fraction, 
containing at least the C.sub.20 or longer .omega.3 fatty acid moiety and 
the arachidonic acid moiety, proteins, carbohydrates, and any additional 
additives, and homogenizing the mixture into a stable emulsion. 
Sources for the .omega.3 fatty acids are plant oils, marine plankton, 
fungal oils, and fish oils, preferably menhaden, salmon, anchovy or 
herring oils. Arachidonic acid is commercially available from Martek, Inc. 
as a fungal derivative which contains 50% arachidonic acid. 
The preferred protein is a protein hydrolysate. The protein hydrolysate may 
be any suitable partially hydrolyzed protein or protein hydrolysate 
utilized in a nutritional formula such as soy protein hydrolysate, casein 
hydrolysate, whey protein hydrolysate, animal and vegetable protein 
hydrolysates, partially hydrolyzed whey, casein or soy proteins, and 
mixtures thereof. Soy or casein protein hydrolysates comprising a 
substantial proportion of variable chain length peptides, e.g., medium 
chain and short chain peptides, e.g., di- and tri-peptides, but having 
less than about 10% free amino acids, preferably less than about 5% free 
amino acids, are preferred. For greatest use, the protein source should be 
lactose-free so it can be used for lactose intolerant patients. 
When choosing a protein source, the biological value of the protein should 
be considered first, with the highest biological values being found in 
casein, whey, lactalbumin, egg albumin, and whole egg proteins. Next, the 
cost should be considered, the lowest cost with the best biological value 
being the best combination. 
The source of carbohydrate may be any simple monosaccharides, 
disaccharides, oligosaccharides, or complex carbohydrates. Examples 
include fructose, dextrose, glucose, maltodextrin, corn syrup and corn 
starch. Carbohydrate sources which may be utilized in the formulation of 
the invention include hydrolyzed or nonhydrolyzed starches. 
Emulsifiers may be added for stability purposes to the formulation of the 
invention, e.g., emulsifiers such as soybean phospholipids. This may be 
required for parenteral products. 
Flavoring may also be added to the emulsion to make it more palatable for 
enteral use. Flavoring can be in a form of flavored extracts, volatile 
oils, chocolate flavoring, peanut butter flavoring, cookie crumbs, vanilla 
or any commercially available flavoring. 
The formulation of invention may also contain a stabilizer such as 
.lambda.-carrageenan. .lambda.-carrageenan increases the viscosity of the 
formula without forming a gel structure, thus retarding the precipitation 
of insoluble calcium and phosphorus salts if included in the formula. 
Xanthan gum or other standard stabilizers may also be used as a stabilizer 
in the same fashion as .lambda.-carrageenan. 
While the formulation of the invention is preferably provided in a 
ready-to-feed form, it may also be concentrated by increasing the percent 
total solids of the formula or made in powder form, both procedures being 
well known to those skilled in the art. The concentrate or powder are then 
reconstituted for feeding by adding water (tap or deionized-sterilized 
water). 
The structured lipid of the invention may be manufactured by any 
conventional means such as transesterification but the use of blocking 
groups which allow positioning of the residues at specific locations is 
preferred. Those skilled in the art are familiar with the variety of 
techniques useful for directing the residues to particular locations and 
they need not be set forth here in detail. It appears that the use of the 
C.sub.20 or longer .omega.3 fatty acid moiety or more preferably, the 
arachidonic acid moiety in the R.sub.2 position with C.sub.8 -C.sub.12 
fatty acid moiety, preferably a C.sub.12 fatty acid moiety, in the R.sub.1 
or R.sub.3 positions leads to a most preferred triglyceride. The preferred 
triglyceride improves absorption into the body as a whole and allows for 
ease of incorporation of the long chain fatty acid found at the R.sub.2 
position. In another preferred embodiment, it appears that the use of a 
C.sub.8 -C.sub.12 fatty acid moiety, preferably a C.sub.12 fatty acid 
moiety, at R.sub.2 position results in endogenous formation of the 
structured lipid in intestinal cells. 
Certain terms used herein are described below for clarity. 
As used herein, the term "C.sub.20 or longer .omega.3 fatty acid moiety" 
refers to eicosapentaenoic (EPA) acid moiety, docosahexaenoic (DHA) acid 
moiety, or mixtures thereof. 
As used herein, the term "critically ill patients" refers to patients who 
are suffering from a total or partial dysfunction of the gastro-intestinal 
tract due to disease or stress of injury such as surgery, cancer, acute 
diabetes, AIDS, malnutrition, trauma or sepsis. The term "critically ill 
patients", as used herein, is also intended to include hypercatabolic 
patients. These critically ill individuals are often hospitalized and must 
receive most or all of their daily nutritional requirements parenterally 
and/or enterally in order to sustain protein synthesis and to minimize the 
likelihood of becoming malnourished, to maintain nutritional status, or to 
improve nutritional status. 
As used herein, the term "patients suffering from chronic liver disease" 
refers to patients who are suffering from alcoholic liver cirrhosis, liver 
cirrhosis caused by chronic infection after acute inflammation of the 
liver or immunologic liver diseases characterized by chronic inflammation 
without known reason. Metabolic derangements associate with chronic liver 
disease include, but are not limited to: increased plasma glucagon; 
hyperinsulinemia; increased plasma epinephrine and cortisol; decreased 
liver and muscle carbohydrate stores; accelerated gluconeogenesis; 
hypoglycemia; hyperammonemia; increased plasma aromatic amino acid; 
increased plasma methionine, glutamine, asparigine and histidine; and 
decreased plasma branched chain amino acids.