Patent Application: US-53938490-A

Abstract:
the modulation of prostaglandin levels can be realized through the dietary intake of specified ratios of activated omega 6 essentially fatty acids when combined with the appropriate amount of eicosapentaenoic acid , an omega 3 fatty acid . the modulation of prostaglandins levels can be determined by changes in physiological parameters which are related to prostaglandin levels in mammals . certain ratios of activated omega 6 essentially fatty acids and epa can have significant physiological benefits , whereas other ratios demonstrate detrimental physiological effects in mammals .

Description:
the invention consists of a defined combination of essential fatty acids containing gla and / or dgla with the appropriate weight amount of epa to modulate precursor pools for prostaglandin production in mammals . therefore , to fully describe the invention one must illustrate general methods of gla , dgla , and epa preparation . gla in the triglyceride form can be most easily extracted and refined from vegetable seed sources using standard technology common in the edible oil industry to create an oil suitable for human consumption as defined by international standards . 11 common sources of gla would include borage , black current , evening primrose seeds and oat bran . certain microorganisms can also be fermented to produce gla in the triglyceride form which can likewise be refined to meet international standards established for an edible oil . gla isolated in the triglyceride form can be chemically or biochemically transformed into free fatty acids , salts of free fatty acids , methyl or ethyl esters , or monoglycerides which can be further fractionated by standard techniques into fractions with higher gla content than found in the starting oils . finally , gla can be chemically synthesized by standard chemical techniques . a good natural source dgla does not exit , so that to make this essential fatty acid , one must either elongate the free fatty acid , methyl or ethyl esters of gla using standard techniques such as the malonic ester synthesis or to chemically synthesize the compound . like gla , epa can be easily extracted from natural sources such as plankton , krill , or marine animals . also like gla , epa can be fermented under controlled conditions . in both cases , the extracted oil should be refined to meet all international standards for edible oils . again like gla , the triglyceride form of epa can be altered either by chemical or biochemical means to produce free fatty acids , salts of free fatty acids , methyl or ethyl esters , or monoglycerides which can be further fractionated to give higher epa contents than the starting oil in the triglyceride form . epa can also be chemically synthesized . for the purpose of illustration only , the invention will be described in connection with the method of preparation in various pharmaceutical and food products and its use in the treatment in certain cardiovascular and immune disorders and hence by extension , its use in the prophylaxis of such cardiovascular and immune disorders . however , it is recognized that various changes and modifications to the illustrated examples can be made by those persons skilled in the art , all falling within the spirit and scope of the invention . fig1 describes the biochemical relationships of gla and epa that is important in the modulation of prostaglandins . it is the effect of epa as an inhibitior of the enzyme delta 5 desaturase that diverts the flow of gla into dihomo gamma linolenic acid ( dgla ) instead of its further metabolism into arachidonic acid ( aa ). as shown in the examples of the invention , the ratio of epa to gla is critically important in the successful modulation of prostaglandins . six hypertensive subjects with an average blood pressure of 150 / 92 were placed on a daily intake of 80 mg . of gla and 640 mg . of epa per day for 6 weeks . at the end of six weeks , their blood pressure was measured and was lowered to an average of 134 / 78 . their intake was then changed to 80 mg . gla and 320 mg . epa per day for another 2 weeks . at the end of this period , their blood pressure was measured and the average was found to be 140 / 83 . their intake was then changed to 80 mg . gla and 160 mg . epa per day for another 2 weeks . at the end of this 2 week period , their blood pressure was measured , and the average blood pressure was 146 / 88 . their intake was then changed to 80 mg . gla and 80 mg . epa for another 2 weeks . at the end of this period , their blood pressure was measured , and the average blood pressure reading was 156 / 95 . their intake was changed to 80 mg . gla and 40 mg . epa per day for another another 2 weeks . at the end of 2 weeks , their blood pressure was measured , and found to average 162 / 100 . at this point they were switched back to 80 mg . gla and 640 mg . epa for a final 2 weeks . the average blood pressure was reduced to 143 / 85 . the results concerning the cardiovascular effects of different gla and epa ratios are shown in table 1 . table 1______________________________________effect of combinations of activated omega 6 essential fattyacids and epa on blood pressure in hypertensive individualsweeks ratio of epa to gla average blood pressure______________________________________start none 150 / 926 8 : 1 134 / 782 4 : 1 140 / 832 2 : 1 146 / 882 1 : 1 156 / 952 0 . 5 : 1 162 / 1002 8 : 1 143 / 85______________________________________ these results show that some ratios of epa and gla are beneficial in the treatment of existing hypertension , whereas other ratios are detrimental to the existing disease condition . 36 healthy adults were split into 6 groups of 6 individuals . they were given the following amounts of dietary supplements for 6 weeks on a daily basis . at the end of 6 weeks , each individual was asked to evaluate their energy levels , digestive system responses , and skin condition based on the various levels of supplementation on a subjective scale comparing the initial starting point to the final point at the end of the study . the following questions were asked , and the individuals were asked to respond using a grading system ranging from + 2 ( significantly improved ), + 1 ( somewhat improved ), 0 ( no change ), - 1 ( somewhat worse ), - 2 ( significantly worse ). each of these questions was used to assess physiological function which is closely related to prostaglandin formation , and therefore serves as an indication as to how dietary supplementation with activated omega 6 essential fatty acids and epa could effect prostaglandin formation , and therefore utlimately physiological function . energy levels are related to fatigue . prostaglandins derived from aa ( such as thromboxane a 2 ) are powerful vasoconstrictors that restrict the size of the capillary bed , and ultimately reduce the transfer rate of oxygen to muscle tissue . this lack of oxygen transfer will increase the levels of lactic acid in the muscle which causes fatigue . on the other hand , prostaglandins derived from dgla ( such as pge 1 ) are powerful vasodilators which will increase the size of the capillary bed , thereby increasing oxygen transfer to muscle cells . however , if too much pge 1 is formed , the kidneys will undergo a corresponding increase in vasodilation resulting in increased urination , and subsequent electrolyte depletion . the electrolyte depletion will contribution to fatigue , thereby decreasing energy levels . therefore , at either extreme of the dgla to aa ratio in the cardiovascular system , fatigue is the end result . however , at the optimal balance of these two fatty acids , energy levels should increase . therefore , the levels of dgla to aa in the cardiovascular system will determine which prostaglandins are eventually produced , and thus ultimately determine the energy level of the individual . likewise , stool composition is a good indicator of prostaglandin formation in the gastrointensintal tract . vasoconstrictors such as thromboxane a 2 derived from aa will decrease the flow of water into the colon causing constipation . on the other hand , vasodilators such as pge 1 derived from dgla will increase the flow of water into the colon . if too much aa is formed by dietary supplementation with activated omega 6 essential fatty acids and an insufficient amount of epa , then an individual will develop constipation . on the other hand , if too much dgla is being formed , then an individual will develop diarrhea . again the appropriate balance of dgla to aa is reflected in the stool composition . thus the ratio of dgla to aa in the mucosa that lines the gastrointensinal tract will determine which prostaglandins are ultimately produced which are reflected in the stool composition . finally , the skin also responds to changes in the dgla to aa ratio which will manifest itself in the modulation of prostaglandins . prostaglandins such as pge 2 derived from aa are pro - inflammatory and will increase existing skin disorders such as eczema or produce dry skin . furthermore , another group of prostaglandins known as leukotrienes produced from aa which are a major factor in the promotion of inflammatory response and allergies . on the other hand , prostaglandins produced from dgla such as pge 1 are anti - inflammatory and tend to reduce skin disorders . moreover , leukotrienes cannot be formed from dgla , so that their levels will also be modulated by the ratio of dgla to aa in the skin . therefore , the ratio of dgla to aa in the skin will determine which prostaglandins are ultimately produced . in table 2 is shown the results of this study with the six groups of individuals using different ratios of activated omega 6 essential fatty acids and epa . table 2______________________________________effect of weight ratios of activated omega 6 essentialfatty acids and epa on physiological responsesrelated to prostaglandin formationgroup epa / gla ratio energy stool skin______________________________________1 40 : 1 - 1 . 5 ± 0 . 2 - 1 . 3 ± 0 . 2 + 1 . 5 ± 0 . 22 20 : 1 + 0 . 8 ± 0 . 3 - 0 . 2 ± 0 . 3 + 1 . 3 ± 0 . 23 10 : 1 + 1 . 8 ± 0 . 2 + 1 . 5 ± 0 . 2 + 0 . 5 ± 0 . 24 5 : 1 + 1 . 2 ± 0 . 2 + 0 . 7 ± 0 . 3 + 0 . 7 ± 0 . 25 1 : 1 - 1 . 2 ± 0 . 3 - 1 . 7 ± 0 . 3 - 0 . 8 ± 0 . 36 0 . 5 : 1 - 1 . 7 ± 0 . 2 - 1 . 7 ± 0 . 3 - 1 . 3 ± 0 . 2______________________________________ this data shows a number of results . at high ratios of epa to gla , energy levels and stool composition ( due to diarrhea ) worsened for the individuals at the end of the study compared to the start . both results can be explained by excessive vasodilation caused by too much dgla formation without a compensating production of aa to maintain an appropriate dgla to aa balance in the cardiovascular system and gastrointensinal tract . on the other hand , their skin condition improved . this is because the skin is not as senistive to high dgla levels as are the other systems . thus at high epa to gla weight ratios there are some benefits , although many negative effects are also observed . the ratio of 10 : 1 epa to gla in this study produced optimal results in the improvement of all three areas ( i . e . energy , stool composition and skin ). at the lowest ratios of epa to gla , a distinct decrease in the energy , stool composition ( due to constipation ) and the skin composition ( dryness and flaring up of eczema , if an existing condition prior to supplementation ) were observed . all of these effects can be explained by the increased levels of aa formation giving rise to a decreased dgla to aa ratio in these body tissues . correspondingly , the lowered dgla to aa ratio would lead to increased levels of vasoconstrictors and pro - inflammatory prostaglandins which adversely effect these physiological functions . these results indicate that there is a specific range of activated omega 6 essential fatty acids and epa weight ratios that modulate prostaglandin levels to the benefit of humans , whereas other ratios actually decrease the individual &# 39 ; s health . it should be noted that the ratios of activated omega 6 essential fatty acids and epa disclosed in the examples of prior art ( u . s . pat . no . 4 , 681 , 896 ) would have caused detrimental effects if given to patients with atopic disorders . 6 patients with clinical manifestations of rheumatoid arthritis were place on a daily regimen of 480 mg . epa and 120 mg . of gla for three months . at the end of the time period , all clinical signs of rheumatoid arthritis were significantly diminished by joint pain as assessed by their physician and self assessment . this reduction is joint pain can be related to formation of prostaglandins which are anti - inflammatory , and the simultaneous suppression of prostaglandins which are pro - inflammatory . 10 normal subjects were placed on a daily dose of 640 mg . epa and 80 mg . gla for 2 weeks . blood samples were taken , the platelets were isolated . measurements aggregation of platelets were taken before and after the supplementation program . in these platelet aggregation studies , the isolated platelets were stimulated with collagen ( 0 . 5 ug / ml ) and a thromboxane a 2 analog ( u46619 ) { 500 ng / ml }. the results are shown in table 3 . table 3______________________________________effects of epa and activated omega 6 essentialfatty acids on platelet aggregation . average before average afterparameter supplementation supplementation p______________________________________aggregation 117 ± 18 88 ± 27 & lt ; 0 . 01with u46619aggregation lag 66 ± 23 79 ± 36 & lt ; 0 . 05with collagen * ______________________________________ n . s . not statistically significant * lag time in seconds before aggregation these results can be summarized as follows : there was no change in the platelet count , but there was a statistically significant decrease in platelet aggregation times . this indicates that the decrease in platelet aggregation was due to modulation of prostaglandins in the platelets by the dietary supplementation with epa and gla . this modulation of prostaglandins decreased the tendency of these platelets to aggregate when stimulated by an external response . a 0 . 5 gram soft gelatin capsule containing 15 mg . gla and 60 mg . epa . the number of capsules taken on a daily basis to modulate prostaglandins on a short term basis ( up to 30 days ) would be 4 capsules per day . the amount ingested for a long term basis would be one capsule per day as less activated omega 6 essential fatty acids are required to maintain the tissue levels of dgla to aa once they are established . to illustrate the effect of the invention for the treatment of immune disorders , an aids patient with clinicial signs of karposi &# 39 ; s sarcoma was put on a dialy regime of 8 0 . 5 gram soft gelatin capsules containing 15 mg . gla and 60 mg . of epa for 60 days . after 60 days , the dosage was increased to 16 capsules per day for the next 6 months . three months after starting the program , the lesions associated with karposi &# 39 ; s sarcoma had disappeared . the lesions reappeared after eight months from the start of the program . this result shows that a relatively low amount of the invention can cause regression of cancerous lesions such those associated with karposi &# 39 ; s sarcoma . a 0 . 5 gram soft gelatin capsule containing 8 mg . dgla and 80 mg . epa . the number of capsules taken on a daily basis to modulate prostaglandins on a short term basis ( up to 30 days ) would be 4 capsules per day . the amount taken for a long term basis would be one capsule per day as less activated omega 6 essential fatty acids are required to maintain tissue levels of dgla to aa once they are established . another example of a pharmaceutical composition of the invention is a physiologically compatible intravenous emulsion suitable for injection . 1 . 0 grams of purified soybean lecithin containing 75 % phosphatidylcholine was dispersed in 100 ml . of distilled water buffered with 1 mm phosphate to ph 7 . 5 . to the dispersed lecithin was added 10 grams of oil containing 1 . 2 grams epa and 0 . 3 grams of gla and 2 . 25 g of glycerine . the material was emulsified with a branson w - 375 sonifier under a nitrogen atomsphere . the resulting dispersion consisted of an emulsion with an average particle size of 261 nm . to illustrate an example of a food product containing the invention , a dairy emulsion suitable for food use can be made by the dispersion of 1 . 0 grams of a purified soybean lecithin fraction consisting of 45 % phosphatidylcholine in 100 ml . of distilled water . to the dispersed lecithin is added 10 grams of oil containing 1 . 2 grams of epa and 0 . 3 g gla and 0 . 01 grams of artificial chocolate flavor . the mixture was homogenized and passed through a microfluidizing apparatus to produce a dispersion with an average particle size of 275 nm . to illustrate the potential of increasing the potency of the invention by fractionation of the active ingredients , in particular the gla component , the following example is given . 500 g . of refined borage oil was refluxed for 1 hour with a solution of 400 ml . of ethanol , 125 ml . of water , and 115 g . of koh . the mixture was cooled and 500 ml of crushed ice and 600 ml . of 0 . 4m h 2 so 4 was added . the layers were separated , and the upper layer was dried by the addition of 3 % by weight of mgso 4 . at this point , the triglycerides of the borage oil have been transformed into free fatty acids . the mgso 4 was filtered , and the free fatty acids are refluxed for 1 hour with 1000 ml . of methanol and 20 ml . of concentrated h 2 so 4 . after cooling , 1500 ml . of water was added and phases were separated . to the upper phase containing the methyl esters of gla was added 3 % by weight of mgso 4 . the mgso 4 was filtered and the solution was evaporated to dryness . at this point the concentration of the methyl esters of gla relative to other fatty acids was 22 . 1 % and the content of linoleic acid was 37 . 2 %, which was similar to that found in the starting borage oil . to further increase the potency of the methyl ester of gla the following procedures were used . 1400 g . of urea was dissolved in 5600 ml . of warm methanol / ethanol ( 2 : 1 v / v ). to the dissolved urea was added 500 g . of the methyl esters of gla derived from borage oil . the mixture was placed at 0 ° c . overnight . the mixture was filtered and washed with cold methanol . to eliminate any urea that may have stayed in solution , to the filtered solution was added 1000 ml . of 0 . 4m h 2 so 4 for every 2000 ml . of filtered solution . the upper phase was separated and dried with mgso 4 . the mgso 4 was filtered , and the solvent were evaporated under vacuum . from the starting 500 g . of unfractionated methyl esters of gla , the yield was 96 g . analysis of this fraction by gas liquid chromatography showed that the composition of this fraction was 92 . 4 % gla and 6 . 8 % linoleic acid . the total recovery of gla was 80 . 3 %. 1 . lands , w . e . m . &# 34 ; fish and human health &# 34 ; academic press , new york . 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