Phospholipids and fish and shellfish feed containing same

Abstract

Provided is a phospholipid to which DHA and / or EPA is bound, wherein: the DHA and / or the EPA is contained at a total amount of 2-50 wt% relative to the entirety of constituent fatty acids in the phospholipid; and (the total amount of DHA and / or EPA bound to position 2 of the phospholipid) / (the total amount of DHA and EPA bound to position 1 and position 2 of the phospholipid)(weight ratio) is 0.2-0.8.

Description

[Technical field]

[0001] The present invention relates to a phospholipid or a method for producing the same, and a fish and shellfish feed containing the phospholipid. [Background technology]

[0002] In the future, as the global population increases, it will become increasingly necessary to farm fish and shellfish, so it is extremely important to develop inexpensive, nutritious and functional feed to increase yields. In addition to trace elements such as proteins, carbohydrates and vitamins, lipids are also important components of fish feed.

[0003] Highly unsaturated fatty acids such as docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) contained in fish oils are particularly important as nutrients for seafood, and although they can be ingested in the form of triglycerides such as refined fish oil, it is more effective to ingest them bound to phospholipids. In particular, for young fish that do not have strong digestive abilities, it is said that feeding DHA and / or EPA bound to phospholipids to seafood as a compound feed or as a fortifier for Artemia or rotifers will provide greater nutritional benefits.

[0004] Phospholipids bound with DHA and / or EPA are mainly contained in oils and fats extracted from fish eggs such as salmon roe and squid, but there are problems with these being in limited supply and expensive.

[0005] As a method for producing phospholipids bound with DHA and / or EPA, Patent Document 1 discloses a method in which lysolecithin is bound to DHA and / or EPA using phospholipase A2. Although the supply of raw materials is stable, this method uses expensive enzymes, takes a long time to manufacture, and is poor in productivity. In addition, most of the binding sites of DHA and / or EPA are at the second position of phospholipids.

[0006] Patent Document 2 discloses an artificial formula feed for juvenile tuna fish containing a phospholipid fraction bound with DHA and / or EPA, which is prepared from crude fish oil extracted from fish heads as the main raw material. However, since the raw material is fish heads, the supply is unstable, and most of the binding sites of DHA and / or EPA are at the second position of phospholipids. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] JP 2016-28612 A [Patent Document 2] JP 2013-59293 A Summary of the Invention [Problem to be solved by the invention]

[0008] The object of the present invention is to provide a DHA- and / or EPA-bound phospholipid that improves the growth performance and stress resistance of cultured fish, and fish and shellfish feed containing the phospholipid, and to provide a method for producing the phospholipid at low cost, with high productivity, and capable of supplying the phospholipid stably. [Means for solving the problem]

[0009] As a result of extensive research into solving the above-mentioned problems, the inventors have discovered that the above-mentioned problems can be solved by adjusting the proportion of DHA and EPA in the total constituent fatty acids of phospholipids within a specific range, and by setting the ratio (weight ratio) of the total amount of DHA and EPA bound to the 2nd position of phospholipid to the total amount of DHA and EPA bound to the 1st and 2nd positions of phospholipids within a specific range, thereby completing the present invention.

[0010] That is, the first aspect of the present invention relates to a DHA- and / or EPA-bound phospholipid, which contains 2 to 50% by weight of DHA and / or EPA in total in the total constituent fatty acids of the phospholipid, and has a weight ratio of (total amount of DHA and EPA bound to the 2-position of the phospholipid) / (total amount of DHA and EPA bound to the 1-position and 2-position of the phospholipid) of 0.2 to 0.8. Preferably, the raw material of the phospholipid is derived from a plant. A second aspect of the present invention relates to a feed composition for fish and shellfish, which contains the above-mentioned DHA and / or EPA-bound phospholipid. The third aspect of the present invention relates to a fish and shellfish feed containing 0.4 to 1.5% by weight of phospholipids in total, the phospholipids containing 2 to 50% by weight of DHA and / or EPA in total of all the constituent fatty acids of the phospholipids, and the weight ratio of (the total amount of DHA and EPA bound to the 2-position of the phospholipids) / (the total amount of DHA and EPA bound to the 1-position and 2-position of the phospholipids) being 0.2 to 0.8. Preferably, the raw material of the phospholipids is of plant origin. Preferably, the fish and shellfish is red sea bream fry. The fourth aspect of the present invention relates to a method for producing a DHA- and / or EPA-bound phospholipid, comprising adding an alkali metal catalyst in an amount of 2 to 10 times (by weight) equivalent to the acid value of the mixture, the mixture having a mixing ratio of 15:85 to 60:40 (weight ratio) of phospholipid to oils and / or fatty acid esters containing DHA and / or EPA as constituent fatty acids, the mixture containing 5 to 60% by weight in total of DHA and EPA in the total constituent fatty acids of the mixture, and reacting the mixture at 30 to 100°C for 5 to 120 minutes. Effect of the Invention

[0011] According to the present invention, there are provided DHA and / or EPA-bound phospholipids that improve the growth performance and stress resistance of cultured fish, and fish and shellfish feed containing the phospholipids, and further, a production method that can supply the phospholipids inexpensively, productively, and stably. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] The present invention will now be described in further detail. The DHA and / or EPA-bound phospholipid of the present invention refers to a phospholipid in which a phosphate group and a fatty acid containing DHA and / or EPA are bound to a glycerin skeleton. In the DHA and / or EPA-bound phospholipid, a phosphate group is bound to the phospholipid, and constituent fatty acids containing DHA and EPA are bound to the 1st and 2nd positions of the phospholipid. The constituent fatty acids of the DHA and / or EPA-bound phospholipid may contain only one of DHA and EPA, or may contain both. In addition, the DHA and / or EPA-bound phospholipid may contain phospholipid molecules to which DHA and / or EPA are not bound, in addition to phospholipid molecules to which DHA and / or EPA are bound.

[0013] The DHA (docosahexaenoic acid) is an ω-3 unsaturated fatty acid, a 22-carbon carboxylic acid (22:6) containing six double bonds. Similarly, the EPA (eicosapentaenoic acid) is an ω-3 unsaturated fatty acid, a 20-carbon carboxylic acid (20:5) containing five cis-type double bonds.

[0014] The DHA and / or EPA-bound phospholipid of the present invention preferably contains DHA and EPA in total of 2-50% by weight, more preferably 3-30% by weight, even more preferably 5-25% by weight, and particularly preferably 7-20% by weight, based on the total fatty acids constituting the phospholipid. If it is less than 2% by weight, it may be expensive or the balance with other nutrients may be lost, resulting in insufficient effect as a feed. If it is more than 50% by weight, it may be expensive or the production efficiency may be poor. The type of the constituent fatty acids other than DHA and EPA in the DHA and / or EPA-bound phospholipid of the present invention is not particularly limited.

[0015] In the DHA and / or EPA-bound phospholipid of the present invention, the weight ratio of (total amount of DHA and EPA bound to the 2-position of the phospholipid) / (total amount of DHA and EPA bound to the 1-position and 2-position of the phospholipid) is preferably 0.2 to 0.8, more preferably 0.3 to 0.7, and even more preferably 0.4 to 0.6. If it is less than 0.2, processing such as hydrolysis of only the fatty acid at the 2-position after the transesterification described below may be required, which may result in high costs. If it exceeds 0.8, the effect as a feed may be inferior.

[0016] As the source of DHA and / or EPA used in producing the DHA and / or EPA-bound phospholipids of the present invention, it is preferable to use oils and fats derived from fish oil, marine animal oil, and algae, which are rich in DHA and / or EPA as constituent fatty acids, oils and fats in which DHA and / or EPA have been concentrated by fractionation or hydrolysis of these oils and fats, or fatty acid esters containing DHA and / or EPA as constituent fatty acids.

[0017] Examples of the fish oil include oils and fats obtained from fish and shellfish such as liver oil, herring, mackerel, salmon, sardine, and Antarctic krill, and tuna orbital oil. Examples of the marine animal oil include oils and fats obtained from seals, fur seals, sea lions, etc. Examples of the algae-derived oils and fats include oils and fats obtained from Euglena algae, Aurantiochytrium, Schizochytrium, etc.

[0018] The phospholipid source used in producing the DHA and / or EPA-bound phospholipid of the present invention can be exemplified by plant-derived lecithin such as soybean lecithin, rapeseed lecithin, sunflower lecithin, etc., which are rich in phospholipids, animal-derived lecithin such as egg yolk lecithin, lecithin obtained by degreasing these with acetone to increase the phospholipid concentration, or lecithin derived from seafood.Among these, plant-derived lecithin is preferred from the viewpoint of cost and supply.

[0019] The DHA and / or EPA-bound phospholipid of the present invention can be mixed directly into a compound feed to produce a fish and shellfish feed. Alternatively, a fish and shellfish feed composition containing the DHA and / or EPA-bound phospholipid may be prepared and mixed with other feed ingredients to produce a fish and shellfish feed. The fish and shellfish feed composition may contain fats and oils in addition to the DHA and / or EPA-bound phospholipid.

[0020] The other feed ingredients are not particularly limited, but examples thereof include protein sources such as fish meal, soybean meal, corn meal, gluten, and casein, oil sources such as amino acids, fish oil, and vegetable oil, lecithin sources such as soybean lecithin and egg yolk lecithin, carbohydrates such as dextrin and starch, and nutritional components such as vitamins, minerals, and taurine. The DHA and / or EPA-bound phospholipid of the present invention can also be used as a raw material for compound feed for poultry such as chickens, pigs, and cows.

[0021] The content of the DHA and / or EPA-bound phospholipid of the present invention in the fish and shellfish feed composition is preferably 5% by weight or more, more preferably 5-50% by weight, even more preferably 10-50% by weight, particularly preferably 20-50% by weight, and most preferably 30-40% by weight. If it is less than 5% by weight, lipid components other than phospholipids in the fish and shellfish feed composition will be relatively high, resulting in a poor lipid balance in the feed and poor effect as a feed. If it is more than 50% by weight, the DHA and / or EPA-bound phospholipid components will be excessive, and the effect may be reduced.

[0022] The content of the DHA and / or EPA-bound phospholipid of the present invention in the fish and shellfish feed is preferably 0.4 to 1.5% by weight, more preferably 0.4 to 1.2% by weight, and even more preferably 0.4 to 1.0% by weight, based on the total amount of the fish and shellfish feed. If the content is less than 0.4% by weight, sufficient growth effect may not be obtained. If the content is more than 1.2% by weight, the DHA and / or EPA-bound phospholipid component may be excessive, resulting in reduced effect.

[0023] The fish and shellfish feed composition containing the DHA and / or EPA-bound phospholipid of the present invention can be produced, for example, as follows. A mixture is prepared by mixing the raw phospholipid with fats and / or oils containing DHA and / or EPA as constituent fatty acids. In the mixture, the mixing ratio of the raw phospholipid to the fats and / or oils and / or fatty acid ester is preferably 15:85 to 60:40 (weight ratio), more preferably 20:80 to 60:40, and even more preferably 30:70 to 50:50. If the ratio of the raw phospholipid is less than 15, the phospholipid content in the product is reduced, which may result in poor production efficiency. If the ratio of the raw phospholipid exceeds 60, the fluidity of the reaction liquid in the transesterification described below may be poor, reducing reactivity, or making post-reaction treatment difficult.

[0024] The fats and / or oils and / or fatty acid esters contain DHA and / or EPA as constituent fatty acids. The higher the total content of DHA and EPA in the total constituent fatty acids of the mixture, the higher the total content of DHA and EPA in the phospholipid after the reaction, which is preferable, but from the viewpoint of cost, the total content of DHA and EPA in the total constituent fatty acids of the mixture is preferably 5 to 60% by weight, more preferably 10 to 50% by weight, and even more preferably 15 to 40% by weight.

[0025] It is preferable to use phospholipids that have been degreased with acetone in advance to remove unnecessary triglycerides, as this increases the total content of DHA and EPA in the phospholipids after the reaction.

[0026] In order not to inactivate the alkali metal catalyst, it is preferable to remove the water content in the mixture of the raw material phospholipid and the fats and / or fatty acid esters containing DHA and / or EPA as constituent fatty acids to 0.1% by weight or less before adding the catalyst. When removing the water content, for example, the mixture may be heated to 30 to 100°C and stirred under reduced pressure of 1 kPa or less to remove the water content. In this case, it is preferable to add silicon because it can suppress foaming during reduced pressure.

[0027] After removing the moisture, an alkali metal catalyst capable of catalyzing the transesterification of fats and oils is then added in an amount of 2 to 10 times (weight) equivalent relative to the acid value of the mixture. The amount added is more preferably 3 to 9 times equivalent, even more preferably 4 to 8 times equivalent, and particularly preferably 4 to 7 times equivalent. If the amount added is less than 2 times equivalent, the reaction may not proceed, and if it exceeds 10 times equivalent, the phospholipid may deteriorate or decomposition may proceed. The alkali metal catalyst may be of any type as long as it can catalyze the transesterification of fats and oils, but sodium methylate is preferred because it is widely used.

[0028] After adding the alkali metal catalyst, the composition for fish and shellfish feed containing the DHA and / or EPA-bound phospholipid of the present invention can be obtained by reacting for 5 to 120 minutes at preferably 30 to 100° C. The reaction temperature is more preferably 40 to 90° C., further preferably 50 to 80° C., and particularly preferably 50 to 70° C. If the reaction temperature is less than 30° C., the reaction may not proceed easily, and if it exceeds 100° C., the phospholipid may deteriorate or decompose.

[0029] As described above, the reaction time is preferably 5 to 120 minutes, more preferably 10 to 100 minutes, further preferably 15 to 80 minutes, and particularly preferably 20 to 60 minutes. If the reaction time is less than 5 minutes, the reaction may not be completed, and if it exceeds 120 minutes, the phospholipid may deteriorate or decompose.

[0030] After the reaction is completed, it is preferable to neutralize the mixture with an acid as necessary and remove excess acid used for neutralization by vacuum treatment. After neutralization and vacuum treatment, the phospholipid may be concentrated with acetone or may be used as it is. The DHA and / or EPA-bound phospholipid of the present invention can be obtained by separating the phospholipid components from the obtained fish and shellfish feed composition by a conventional method.

[0031] The fish and shellfish feed containing DHA and / or EPA-bound phospholipid of the present invention can be widely used for fish and shellfish in general and poultry.Specifically, the fish and shellfish include red sea bream, yellowtail, shrimp, amberjack, tuna, salmon, trout, etc., and it is particularly preferable to use it for red sea bream juveniles.Furthermore, the poultry include chicken, pig, cow, etc. EXAMPLES

[0032] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples. In the examples, "parts" and "%" are by weight.

[0033] <Method for measuring the content of DHA and / or EPA-bound phospholipids in a composition or salmon roe oil> The composition obtained in the examples or the salmon roe oil was precipitated by adding ice-cold acetone, and the weight of the dried product was measured and used as the weight of DHA and / or EPA-bound phospholipids.

[0034] <Method for measuring the content of DHA and EPA in the total fatty acids constituting the phospholipids contained in the composition or sujiko oil> After adding ice-cold acetone to the composition or sujiko oil obtained in the examples to precipitate the phospholipids, 10 mg of the dried phospholipids were dissolved in 2 ml of isooctane, and 1 ml of a 0.2 M methanol solution of sodium methylate was added and heated at 60°C for 10 minutes while stirring. The mixture was neutralized with acetic acid, water was added, and the upper isooctane layer was collected and analyzed by gas chromatography. The gas chromatograph used was an Agilent 7890. The column used was an Agilent DB-23 (length 30 m, inner diameter 0.25 mm, film thickness 0.25 μm), and the inlet temperature was 260°C, the detection temperature was 260°C, and the oven temperature was constant at 200°C, and the DHA and EPA contents in the total fatty acids constituting the phospholipids were obtained as B% by weight.

[0035] On the other hand, 200 mg of the composition obtained in the Examples or the phospholipid obtained from the salmon roe oil was added with 2 ml of 0.2 M Tris-HCl pH 8.0 buffer and stirred thoroughly, and 50 μl of a porcine pancreatic phospholipase A2 solution (1000 U / ml) was added, and 10% by weight CaCl 2 100μl of aqueous solution was added and reacted at 60℃ for 3 hours to selectively hydrolyze only fatty acids bound to the 2-position of phospholipids. Phospholipids and hydrolyzed fatty acids were extracted from the reaction solution with a chloroform-methanol=2:1 solution, the phospholipid fraction was removed by TLC, and only the free fatty acids generated by hydrolysis of the 2-position of phospholipids were separated and the composition of the free fatty acids was measured using boron trifluoride as per the standard method. The composition of the free fatty acids measured in this way was considered to be the composition of the constituent fatty acids at the 2-position of phospholipids, and the DHA and EPA content in the constituent fatty acids at the 2-position of phospholipids was A% by weight, and the DHA and EPA content in the total constituent fatty acids of phospholipids was B% by weight. The ratio of the total amount of DHA and EPA bound to the 2-position of phospholipids to the total amount of DHA and EPA bound to the 1-position and 2-position of phospholipids was calculated by (A / 2B).

[0036] <Method for measuring the content of DHA and EPA in the total phospholipids contained in the composition or sujiko oil> When measuring fatty acids as shown in the method for measuring the DHA and EPA contents in the compositions obtained in the Examples or in the salmon roe oil, tricaprin was mixed as a standard and the DHA and EPA contents in the total phospholipids were calculated.

[0037] <Method of measuring weight gain rate> The average weight of the red sea bream fry at the start of the feeding test was taken as 100, and the average weight at the end of the test was calculated as a percentage to obtain the rate of increase or decrease (%). The rate of increase or decrease obtained in Comparative Example 5 or 7 was set as 1, and the ratio of the rate of increase or decrease obtained in other Examples or Comparative Examples was calculated. The obtained rate of increase or decrease ratio was evaluated based on the following criteria. 5 points: The ratio of increase / decrease rate to Comparative Example 5 or 7 is 1.2 or more 4 points: The ratio of the increase / decrease rate to Comparative Example 5 or 7 is 1.1 or more and less than 1.2 3 points: The ratio of the increase / decrease rate to Comparative Example 5 or 7 is 0.95 or more and less than 1.1 2 points: The ratio of the increase / decrease rate to Comparative Example 5 or 7 is 0.9 or more and less than 0.95 1 point: The ratio of increase / decrease rate to Comparative Example 5 or 7 is less than 0.9

[0038] <Method of measuring survival rate> The number of surviving red sea bream fry at the end of the feeding test was calculated as a percentage, with the number of fry at the start of the test taken as 100, to obtain a survival rate (%). The survival rate obtained in Comparative Example 5 or 7 was set to 1, and the survival rate ratios obtained in other Examples or Comparative Examples were calculated. The survival rate ratios obtained were evaluated based on the following criteria. 5 points: The survival rate ratio to Comparative Example 5 or 7 is 1.0 or more. 4 points: The survival rate ratio to Comparative Example 5 or 7 is 0.975 or more and less than 1.0 3 points: The survival rate ratio to Comparative Example 5 or 7 is 0.95 or more and less than 0.975 2 points: The survival rate ratio to Comparative Example 5 or 7 is 0.92 or more and less than 0.95. 1 point: The survival rate ratio to Comparative Example 5 or 7 is less than 0.92

[0039] <How to measure blood cortisol levels> After the feeding test, an equal amount of diethyl ether was added to the plasma sample collected from the red sea bream fry, and the mixture was stirred in a vortex mixer. The separated upper layer (ether layer) was collected, and the ether was removed under a nitrogen gas stream to obtain a dried sample. A commercially available kit (Oxford Biochemical Research's "Cortisol EIA Kit") was used to analyze the cortisol concentration. The blood cortisol value obtained in Comparative Example 5 or 7 was set to 1, and the ratio of the blood cortisol value obtained in other Examples or Comparative Examples was calculated. The obtained blood cortisol value ratio was evaluated based on the following criteria. 5 points: The ratio of blood cortisol level to Comparative Example 5 or 7 is less than 0.8 4 points: The ratio of blood cortisol value to Comparative Example 5 or 7 is 0.8 or more and less than 0.9 3 points: The ratio of blood cortisol value to Comparative Example 5 or 7 is 0.9 or more and less than 1.1 2 points: The ratio of blood cortisol level to Comparative Example 5 or 7 is 1.1 or more and less than 1.2 1 point: The ratio of blood cortisol level to Comparative Example 5 or 7 is 1.2 or more

[0040] (comprehensive evaluation) A comprehensive evaluation was performed based on the results of weight gain rate, survival rate, and blood cortisol level. The evaluation criteria were as follows: A: The total score of the weight gain rate, survival rate, and blood cortisol level is 14 or 15 points. B: No score below 3 on the evaluation scores for weight gain rate, survival rate, and blood cortisol level, and the total score is 12 or 13 points C: At least one score of 3 or less for the weight gain rate, survival rate, and blood cortisol level, no score of 1 or 2, and a total score of 12 or 13. D: At least one score below 3 for the weight gain rate, survival rate, and blood cortisol level, and no score below 1, for a total of 11 points E: The total score of the evaluation points for weight gain rate, survival rate, and blood cortisol level is 10 points or less, or there is one or more points of 1.

[0041] (Reference Example 1) Sujiko Oil Commercially available sujiko oil (Marine Tech Co., Ltd.'s "Sujiko Emulsified Oil") is a composition containing DHA and / or EPA-bound phospholipids. As shown in Table 1, the content of DHA and / or EPA-bound phospholipids in sujiko oil was 21.9% by weight. In addition, ice-cold acetone was added to sujiko oil to obtain precipitated phospholipids, and the total amount of DHA and EPA in the total constituent fatty acids of the phospholipids was 41.3% by weight. Furthermore, the weight ratio of (total amount of DHA and EPA bound to the 2nd position of the phospholipids) / (total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipids) in the total phospholipids was 0.85.

[0042] (Reference Example 2) Synthesis of DHA and / or EPA-bound phospholipids using phospholipase A2 (based on Production Example 1 and Example 1 of Patent Document 1) In a 1L glass reaction vessel, 200g of glycerin (manufactured by Sakamoto Pharmaceutical Co., Ltd.) was added with 15g of lysophosphatidylcholine (manufactured by Tsuji Oil Mills, "SLP-White Lyso"), 6g of phospholipase A2 (manufactured by Sanyo Fine, "Lisonase", activity: 50,000 U / g), 6g of fatty acid hydrolyzed from ethyl esters containing 78.5% by weight of DHA and EPA in the total constituent fatty acids, 6g of glycine, 6g of alanine, and 2ml of 2M calcium chloride aqueous solution, and the mixture was stirred and reacted at 50°C for 48 hours under a reduced pressure of 300 Pa to obtain 13g of DHA and / or EPA-bound phospholipid. The DHA and EPA content in the total constituent fatty acids of this phospholipid was 36.5% by weight, and further, the weight ratio of (the total amount of DHA and EPA bound to the 2nd position of the phospholipid) / (the total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipid) in the total phospholipid was 0.95.

[0043] (Reference Example 3) Analysis of DHA- and EPA-bound phospholipids contained in fish meal Lipids were extracted from fish meal ("Anchovy Meal" manufactured by Higashimaru Co., Ltd.) using a solvent of chloroform:methanol 2:1, and the extracted lipids were precipitated with acetone as per the usual method to recover phospholipids. The DHA and EPA content in the total fatty acids constituting the phospholipids was 39.8% by weight, and the weight ratio of (total amount of DHA and EPA bound to the 2nd position of the phospholipid) / (total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipid) in the phospholipids was 0.86.

[0044] [Table 1]

[0045] Example 1: Preparation of a fish and shellfish feed composition containing DHA and / or EPA-bound phospholipids 100g of tuna neutral oil (manufactured by Yokoseki Oil Industries Co., Ltd., "Refined Tuna Oil L") containing 25.3% by weight of DHA and 6.5% by weight of EPA in the total constituent fatty acids of the oil and fat was mixed with 50g of defatted soybean lecithin (manufactured by Tsuji Oil Mills Co., Ltd., "SLP-White"), 15mg of silicon (manufactured by Shin-Etsu Chemical Co., Ltd., "KS-69") was added, and the mixture was stirred at 60°C for 30 minutes while reducing the pressure to 500 Pa with a vacuum pump to remove moisture, reducing the moisture from 0.3% to 0.02%. The acid value of this mixture was 8.7mgKOH / g. The total content of DHA and EPA in the total constituent fatty acids of this mixture was 24.5% by weight.

[0046] The temperature was maintained at 60°C, and the pressure was returned to normal, and 7 g (5.6 times the equivalent amount of the acid value of the mixture) of sodium methylate (Kanto Chemical Co., Ltd.) was added, and the pressure was reduced again and the mixture was stirred for 30 minutes at 60°C. After the reaction was completed, the mixture was neutralized with 7 ml of acetic acid, and the pressure was reduced to 500 Pa for 1 hour, and then the mixture was filtered to remove solid matter, to obtain a fish and shellfish feed composition containing DHA and / or EPA-bound phospholipid. The reaction conditions in this example are shown in Table 1.

[0047] The content of DHA and / or EPA-bound phospholipids in the fish and shellfish feed composition obtained above was 25.1% by weight. In addition, ice-cold acetone was added to the fish and shellfish feed composition to obtain precipitated phospholipids, and the total amount of DHA and EPA in the total fatty acids constituting the phospholipids was 14.9% by weight. In the total phospholipids, the weight ratio of (the total amount of DHA and EPA bound to the 2nd position of the phospholipid) / (the total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipid) was 0.48.

[0048] Example 2: Preparation of fish and shellfish feed A fish and shellfish feed was prepared according to the composition in Table 2. Specifically, the feed contained 50 parts by weight of fish meal (Higashimaru Co., Ltd.'s "Anchovy Meal"), 20 parts by weight of soybean oil cake (J-Oil Mills Co., Ltd.'s "Soy Pro"), 5 parts by weight of fish oil (Kanematsu Shintoa Foods Co., Ltd.'s "Fish Farming Feed Oil"), 1 part by weight of soybean lecithin (Fujifilm Wako Pure Chemical Industries, Ltd.'s "Lecithin, soybean derived"), 2 parts by weight of the fish and shellfish feed composition prepared in Example 1, dextrin (Nacalai Tesque, Inc.'s "Dextrin"), and 1 part by weight of the fish and shellfish feed composition prepared in Example 2. A fish and shellfish feed was prepared by mixing 4 parts by weight of α-starch ("Okanol" manufactured by Asahi Chemical Industry Co., Ltd.), 4 parts by weight of a vitamin mixture ("Kadai Vitamin" manufactured by Kokin Chemical Co., Ltd.), 3 parts by weight of a mineral mixture ("Kadai Mineral" manufactured by Kokin Chemical Co., Ltd.), 3 parts by weight of activated gluten ("A-Glu SS" manufactured by Glico Nutrition Foods Co., Ltd.), and 4 parts by weight of cellulose ("KC Flock" manufactured by Nippon Paper Industries Co., Ltd.).

[0049] The contents of DHA and EPA derived from the DHA and EPA bound to phospholipids contained in the seafood feed composition blended into the obtained seafood feed, and the content (wt%) of DHA and / or EPA-bound phospholipids contained in the feed, in which the (total amount of DHA and EPA bound to position 2 of phospholipids) / (total amount of DHA and EPA bound to positions 1 and 2 of phospholipids) (weight ratio) was 0.2 to 0.8, are shown in Table 2.

[0050] [Table 2]

[0051] Example 3: Preparation of fish and shellfish feed A seafood feed was prepared in the same manner as in Example 2, except that soybean lecithin was not added and the amount of the seafood feed composition added was changed from 2 parts by weight to 3 parts by weight according to the formulation in Table 2. Table 2 shows the contents of DHA and EPA derived from the DHA and EPA bound to phospholipids contained in the seafood feed composition blended in the obtained seafood feed, and the contents (wt%) of DHA and / or EPA-bound phospholipids contained in the feed, in which the (total amount of DHA and EPA bound to the 2-position of the phospholipid) / (total amount of DHA and EPA bound to the 1-position and 2-position of the phospholipid) (weight ratio) is 0.2 to 0.8.

[0052] (Comparative Example 1) Preparation of fish and shellfish feed A seafood feed was prepared in the same manner as in Example 2, except that the amount of soybean lecithin added was changed from 1 part by weight to 3 parts by weight and no composition for fishery feed was added according to the formulation in Table 2. Table 2 shows the contents of DHA and EPA derived from the DHA and EPA bound to phospholipids contained in the composition for fishery feed blended in the obtained seafood feed, and the content (wt%) of DHA and / or EPA-bound phospholipids contained in the feed, in which the (total amount of DHA and EPA bound to the 2-position of the phospholipid) / (total amount of DHA and EPA bound to the 1-position and 2-position of the phospholipid) (weight ratio) was 0.2 to 0.8.

[0053] (Comparative Example 2) Preparation of fish and shellfish feed Fish and shellfish feed was prepared in the same manner as in Example 2, except that the amount of soybean lecithin added was changed from 1 part by weight to 2 parts by weight, and the amount of the fish and shellfish feed composition added was changed from 2 parts by weight to 1 part by weight, according to the formulation in Table 2. Table 2 shows the contents of DHA and EPA derived from DHA and EPA bound to phospholipids contained in the fish and shellfish feed composition blended in the obtained fish and shellfish feed, and the contents (wt%) of DHA and / or EPA-bound phospholipids contained in the feed, in which the (total amount of DHA and EPA bound to the 2-position of phospholipids) / (total amount of DHA and EPA bound to the 1-position and 2-position of phospholipids) (weight ratio) is 0.2 to 0.8.

[0054] (Comparative Example 3) Preparation of fish and shellfish feed A seafood feed was prepared in the same manner as in Example 2, except that, according to the formulation in Table 2, fish oil was not added, the amount of soybean oil meal added was changed from 20 parts by weight to 18 parts by weight, and the amount of the composition for fish and shellfish feed added was changed from 2 parts by weight to 9 parts by weight. Table 2 shows the contents of DHA and EPA derived from DHA and EPA bound to phospholipids contained in the composition for fish and shellfish feed blended in the obtained seafood feed, and the contents (weight %) of DHA and / or EPA bound phospholipids contained in the feed, in which the (total amount of DHA and EPA bound to the 2-position of phospholipids) / (total amount of DHA and EPA bound to the 1-position and 2-position of phospholipids) (weight ratio) is 0.2 to 0.8.

[0055] (Comparative Example 4) Preparation of fish and shellfish feed A seafood feed was prepared in the same manner as in Example 2, except that the composition for seafood feed was not added, the amount of fish oil added was changed from 5 parts by weight to 5.96 parts by weight, the amount of soybean lecithin added was changed from 1 part by weight to 1.6 parts by weight, and 0.44 parts by weight of salmon roe oil was added, according to the formulation in Table 2. Table 2 shows the contents of DHA and EPA derived from the DHA and EPA bound to phospholipids contained in salmon roe oil added to the obtained seafood feed, and the contents (wt%) of DHA and / or EPA-bound phospholipids contained in the feed, in which the (total amount of DHA and EPA bound to the 2-position of the phospholipid) / (total amount of DHA and EPA bound to the 1-position and 2-position of the phospholipid) (weight ratio) was 0.2 to 0.8.

[0056] (Example 4) Feeding test of red sea bream juveniles A feeding test of red sea bream fry was carried out using the fish and shellfish feed prepared in Example 2. In the feeding test, red sea bream fry obtained from Ogata Suisan Co., Ltd. were raised in a 500L tank by feeding them with a commercially available feed for marine fry until they reached an average weight of 3.5g. The fish were housed in a 100L tank with running water at a density of 15 fish / tank, and a feeding test was carried out using the feed of Example 2 at a water temperature of 19.9 to 23.2°C, with 3 tanks per test area, feeding frequency 2 times / day, and feeding days 40 days. The initial body weight, final body weight, weight gain rate, survival rate, and blood cortisol level measurements, as well as the evaluation results, are shown in Table 3.

[0057] [Table 3]

[0058] (Example 5, Comparative Examples 5 and 6) Feeding test of red sea bream juveniles A red sea bream juvenile feeding test was carried out in the same manner as in Example 4, except that the fishery feed of Example 2 was changed to the fishery feed of Example 3 (Example 5), the fishery feed of Comparative Example 1 (Comparative Example 5), or the fishery feed of Comparative Example 2 (Comparative Example 6) according to Table 3. The initial body weight, final body weight, weight gain rate, survival rate, and blood cortisol level measurements, as well as the evaluation results, are shown in Table 3.

[0059] As is clear from the results in Table 3, compared to red sea bream fry (Comparative Example 5) fed a seafood feed composition containing DHA and / or EPA-bound phospholipids or seafood feed that did not contain salmon roe oil, red sea bream fry (Examples 4 and 5) fed a seafood feed containing 2 to 50% by weight of DHA and / or EPA in total based on the total fatty acids constituting the phospholipids and 0.4 to 1.5% by weight of phospholipids in which the (total amount of DHA and EPA bound to the 2nd position of the phospholipid) / (total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipid) (weight ratio) was 0.2 to 0.8, showed good results in both growth performance (weight increase rate, survival rate) and stress resistance (blood cortisol level).

[0060] On the other hand, red sea bream fry (Comparative Example 6) fed a seafood feed containing 2-50% by weight of DHA and / or EPA in total among all the constituent fatty acids of the phospholipids, and having a phospholipid content of (total amount of DHA and EPA bound to position 2 of the phospholipid) / (total amount of DHA and EPA bound to positions 1 and 2 of the phospholipid) (weight ratio) of 0.2-0.8, was low at 0.251% by weight among all the seafood feed, and had a lower survival rate than Comparative Example 5, and a poor overall evaluation.

[0061] (Example 6) Feeding test of red sea bream juveniles A feeding test of red sea bream juveniles was carried out in the same manner as in Example 4, except that the test was carried out on different days according to Table 4. The initial body weight, final body weight, weight gain rate, survival rate, and blood cortisol level measurements, as well as the evaluation results, are shown in Table 4.

[0062] [Table 4]

[0063] (Comparative Examples 7 to 9) Feeding test of red sea bream juveniles A red sea bream juvenile feeding test was carried out in the same manner as in Example 6, except that the fish and shellfish feed of Example 2 was changed to the fish and shellfish feed of Comparative Example 1 (Comparative Example 7), the fish and shellfish feed of Comparative Example 3 (Comparative Example 8), or the fish and shellfish feed of Comparative Example 4 (Comparative Example 9) according to Table 4. The initial body weight, final body weight, weight gain rate, survival rate, and blood cortisol level measurements, as well as the evaluation results, are shown in Table 4.

[0064] As is clear from the results in Table 4, compared with red sea bream fry (Comparative Example 7) fed a seafood feed composition containing DHA and / or EPA-bound phospholipids or seafood feed that did not contain salmon roe oil, red sea bream fry (Example 6) fed a seafood feed containing 2 to 50% by weight of DHA and / or EPA in total based on the total fatty acids constituting the phospholipids and 0.4 to 1.5% by weight of phospholipids in which the (total amount of DHA and EPA bound to the 2nd position of the phospholipid) / (total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipid) (weight ratio) was 0.2 to 0.8, showed good results in both growth performance (weight increase rate, survival rate) and stress resistance (blood cortisol level).

[0065] On the other hand, red sea bream fry (Comparative Example 8) fed a seafood feed containing a total of 2-50% by weight of DHA and / or EPA in the total fatty acids constituting the phospholipids in the entire seafood feed, and having a high phospholipid content of 2.259% by weight, with a weight ratio of (the total amount of DHA and EPA bound to the 2nd position of the phospholipids) / (the total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipids) of 0.2-0.8, had a lower weight gain rate than Comparative Example 7 and a poor overall evaluation. Furthermore, red sea bream fry (Comparative Example 9) fed seafood feed containing a total of 2 to 50% by weight of DHA and / or EPA in the total fatty acids constituting the phospholipids in the entire seafood feed and no phospholipids in which the (total amount of DHA and EPA bound to the 2nd position of the phospholipids) / (total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipids) (weight ratio) was 0.2 to 0.8 had better stress resistance than Comparative Example 7, but their growth performance (weight increase rate, survival rate) was the same and their overall evaluation was poor.

Claims

1. A DHA- and / or EPA-bound phospholipid, which contains 2 to 50% by weight of DHA and / or EPA in total of all the constituent fatty acids of the phospholipid, and has a weight ratio of (the total amount of DHA and EPA bound to the 2-position of the phospholipid) / (the total amount of DHA and EPA bound to the 1-position and 2-position of the phospholipid) of 0.2 to 0.

6.

2. The DHA and / or EPA-bound phospholipid according to claim 1, wherein the source of the phospholipid is derived from a plant.

3. A fish and shellfish feed composition comprising the DHA and / or EPA-bound phospholipid according to claim 1 or 2.

4. The fish and shellfish feed contains 0.4 to 1.5% by weight of phospholipids in total, the phospholipids containing 2 to 50% by weight of DHA and / or EPA in total of all the constituent fatty acids of the phospholipid, and the weight ratio of (the total amount of DHA and EPA bound to the 2nd position of the phospholipid) / (the total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipid) is 0.2 to 0.

6.

5. The fish and shellfish feed according to claim 4, wherein the raw material of the phospholipid is derived from a plant.

6. The fish and shellfish feed according to claim 4 or 5, wherein the fish and shellfish are red sea bream fry.

7. A method for producing a DHA- and / or EPA-bound phospholipid, which contains 2 to 50% by weight of DHA and / or EPA in total of all constituent fatty acids of the phospholipid, and has a weight ratio of (total amount of DHA and EPA bound to the 2nd position of the phospholipid) / (total amount of DHA and EPA bound to the 1st and 2nd positions of the phospholipid) of 0.2 to 0.8, comprising: A method for producing DHA and / or EPA-bound phospholipids, comprising adding an alkali metal catalyst in an amount of 2 to 10 times (by weight) equivalent to the acid value of a mixture of phospholipids and fats and / or fatty acid esters containing DHA and / or EPA as constituent fatty acids at a mixing ratio of 15:85 to 60:40 (weight ratio), the mixture containing 5 to 60% by weight of DHA and EPA in total of the constituent fatty acids of the mixture, and reacting the mixture at 30 to 100°C for 5 to 120 minutes.

8. The method for producing DHA and / or EPA-bound phospholipids according to claim 7, wherein the raw material for the phospholipids is derived from plants.

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