CA2445738A1 - Method for enriching nervonic acid in expressed milk of ruminants - Google Patents
Method for enriching nervonic acid in expressed milk of ruminants Download PDFInfo
- Publication number
- CA2445738A1 CA2445738A1 CA002445738A CA2445738A CA2445738A1 CA 2445738 A1 CA2445738 A1 CA 2445738A1 CA 002445738 A CA002445738 A CA 002445738A CA 2445738 A CA2445738 A CA 2445738A CA 2445738 A1 CA2445738 A1 CA 2445738A1
- Authority
- CA
- Canada
- Prior art keywords
- milk
- nervonic acid
- ruminants
- feed additive
- enriched
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
Links
- GWHCXVQVJPWHRF-KTKRTIGZSA-N (15Z)-tetracosenoic acid Chemical compound CCCCCCCC\C=C/CCCCCCCCCCCCCC(O)=O GWHCXVQVJPWHRF-KTKRTIGZSA-N 0.000 title claims abstract description 163
- GWHCXVQVJPWHRF-UHFFFAOYSA-N cis-tetracosenoic acid Natural products CCCCCCCCC=CCCCCCCCCCCCCCC(O)=O GWHCXVQVJPWHRF-UHFFFAOYSA-N 0.000 title claims abstract description 163
- XJXROGWVRIJYMO-SJDLZYGOSA-N Nervonic acid Natural products O=C(O)[C@@H](/C=C/CCCCCCCC)CCCCCCCCCCCC XJXROGWVRIJYMO-SJDLZYGOSA-N 0.000 title claims abstract description 162
- 239000008267 milk Substances 0.000 title claims abstract description 93
- 210000004080 milk Anatomy 0.000 title claims abstract description 93
- 235000013336 milk Nutrition 0.000 title claims abstract description 91
- 241000282849 Ruminantia Species 0.000 title claims abstract description 32
- 238000000034 method Methods 0.000 title claims abstract description 25
- 239000003674 animal food additive Substances 0.000 claims abstract description 50
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- 235000012054 meals Nutrition 0.000 claims abstract description 36
- 210000003746 feather Anatomy 0.000 claims abstract description 34
- 239000003112 inhibitor Substances 0.000 claims abstract description 23
- 230000015556 catabolic process Effects 0.000 claims abstract description 15
- 238000006731 degradation reaction Methods 0.000 claims abstract description 15
- 230000000813 microbial effect Effects 0.000 claims abstract description 15
- 210000004767 rumen Anatomy 0.000 claims abstract description 12
- 108010046377 Whey Proteins Proteins 0.000 claims description 33
- 102000007544 Whey Proteins Human genes 0.000 claims description 25
- 235000021119 whey protein Nutrition 0.000 claims description 24
- 239000000047 product Substances 0.000 claims description 15
- 238000004519 manufacturing process Methods 0.000 claims description 14
- 150000004669 very long chain fatty acids Chemical class 0.000 claims description 13
- 102000014171 Milk Proteins Human genes 0.000 claims description 12
- 108010011756 Milk Proteins Proteins 0.000 claims description 12
- 235000018102 proteins Nutrition 0.000 claims description 11
- 102000004169 proteins and genes Human genes 0.000 claims description 11
- 108090000623 proteins and genes Proteins 0.000 claims description 11
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- 239000002253 acid Substances 0.000 claims description 4
- 239000005018 casein Substances 0.000 claims description 4
- BECPQYXYKAMYBN-UHFFFAOYSA-N casein, tech. Chemical compound NCCCCC(C(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(CC(C)C)N=C(O)C(CCC(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(C(C)O)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(COP(O)(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(N)CC1=CC=CC=C1 BECPQYXYKAMYBN-UHFFFAOYSA-N 0.000 claims description 4
- 235000021240 caseins Nutrition 0.000 claims description 4
- 241001656403 Lunaria Species 0.000 claims description 3
- 229940116540 protein supplement Drugs 0.000 claims description 3
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- 239000000470 constituent Substances 0.000 claims 2
- 241000283690 Bos taurus Species 0.000 abstract description 66
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- 239000000194 fatty acid Substances 0.000 description 18
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- 150000004665 fatty acids Chemical class 0.000 description 18
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 5
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 4
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- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 235000014698 Brassica juncea var multisecta Nutrition 0.000 description 3
- 235000006008 Brassica napus var napus Nutrition 0.000 description 3
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- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 3
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- 201000011452 Adrenoleukodystrophy Diseases 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
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- 102000009030 Member 1 Subfamily D ATP Binding Cassette Transporter Human genes 0.000 description 2
- 108010049137 Member 1 Subfamily D ATP Binding Cassette Transporter Proteins 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
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- 239000001103 potassium chloride Substances 0.000 description 2
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- 241000283707 Capra Species 0.000 description 1
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- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 1
- 229920001353 Dextrin Polymers 0.000 description 1
- 239000004375 Dextrin Substances 0.000 description 1
- 102000002322 Egg Proteins Human genes 0.000 description 1
- 108010000912 Egg Proteins Proteins 0.000 description 1
- 241001105021 Fragilariopsis cylindrus Species 0.000 description 1
- 241000287828 Gallus gallus Species 0.000 description 1
- 206010017993 Gastrointestinal neoplasms Diseases 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 108010068370 Glutens Proteins 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 235000019738 Limestone Nutrition 0.000 description 1
- 241000928579 Malania oleifera Species 0.000 description 1
- 229920002774 Maltodextrin Polymers 0.000 description 1
- 239000005913 Maltodextrin Substances 0.000 description 1
- 240000004658 Medicago sativa Species 0.000 description 1
- 235000017587 Medicago sativa ssp. sativa Nutrition 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- SXZWBNWTCVLZJN-NMIJJABPSA-N N-tricosanoylsphing-4-enine-1-phosphocholine Chemical compound CCCCCCCCCCCCCCCCCCCCCCC(=O)N[C@@H](COP([O-])(=O)OCC[N+](C)(C)C)[C@H](O)\C=C\CCCCCCCCCCCCC SXZWBNWTCVLZJN-NMIJJABPSA-N 0.000 description 1
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- 241000589515 Pseudoalteromonas atlantica Species 0.000 description 1
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 1
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 description 1
- 235000019764 Soybean Meal Nutrition 0.000 description 1
- 241001051088 Sphaerotheca humuli Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 241000577145 Thlaspi perfoliatum Species 0.000 description 1
- 241001331104 Tropaeolum speciosum Species 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- 230000009102 absorption Effects 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
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- 210000004185 liver Anatomy 0.000 description 1
- 235000020121 low-fat milk Nutrition 0.000 description 1
- 239000000395 magnesium oxide Substances 0.000 description 1
- CPLXHLVBOLITMK-UHFFFAOYSA-N magnesium oxide Inorganic materials [Mg]=O CPLXHLVBOLITMK-UHFFFAOYSA-N 0.000 description 1
- 235000012245 magnesium oxide Nutrition 0.000 description 1
- AXZKOIWUVFPNLO-UHFFFAOYSA-N magnesium;oxygen(2-) Chemical compound [O-2].[Mg+2] AXZKOIWUVFPNLO-UHFFFAOYSA-N 0.000 description 1
- 229940035034 maltodextrin Drugs 0.000 description 1
- WPBNNNQJVZRUHP-UHFFFAOYSA-L manganese(2+);methyl n-[[2-(methoxycarbonylcarbamothioylamino)phenyl]carbamothioyl]carbamate;n-[2-(sulfidocarbothioylamino)ethyl]carbamodithioate Chemical compound [Mn+2].[S-]C(=S)NCCNC([S-])=S.COC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OC WPBNNNQJVZRUHP-UHFFFAOYSA-L 0.000 description 1
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- 230000004060 metabolic process Effects 0.000 description 1
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- 229940029985 mineral supplement Drugs 0.000 description 1
- 210000000944 nerve tissue Anatomy 0.000 description 1
- 230000003955 neuronal function Effects 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 239000002417 nutraceutical Substances 0.000 description 1
- 235000021436 nutraceutical agent Nutrition 0.000 description 1
- 235000020660 omega-3 fatty acid Nutrition 0.000 description 1
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- 210000000578 peripheral nerve Anatomy 0.000 description 1
- WTJKGGKOPKCXLL-RRHRGVEJSA-N phosphatidylcholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCC=CCCCCCCCC WTJKGGKOPKCXLL-RRHRGVEJSA-N 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
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- 239000011573 trace mineral Substances 0.000 description 1
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- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 1
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Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23D—EDIBLE OILS OR FATS, e.g. MARGARINES, SHORTENINGS, COOKING OILS
- A23D9/00—Other edible oils or fats, e.g. shortenings, cooking oils
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/20—Animal feeding-stuffs from material of animal origin
- A23K10/26—Animal feeding-stuffs from material of animal origin from waste material, e.g. feathers, bones or skin
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/158—Fatty acids; Fats; Products containing oils or fats
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/10—Feeding-stuffs specially adapted for particular animals for ruminants
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C2230/00—Aspects relating to animal feed or genotype
- A23C2230/10—Animal milk with modified composition due to a specific feed
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Polymers & Plastics (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Animal Husbandry (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- Physiology (AREA)
- Molecular Biology (AREA)
- Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Birds (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Fodder In General (AREA)
- Feed For Specific Animals (AREA)
Abstract
The present invention is directed to a new method and feed additive and a method for enriching nervonic acid (NA) in expressed milk of ruminants, for example, dairy cattle. The feed additive comprises a source of NA and an effective amount of an inhibitor of microbial degradation of NA in the rumen of ruminants consisting of feather meal.
Description
Title: METHOD FOR ENRICHING NERVONIC ACID IN
EXPRESSED MILK OF RUMINANTS
Field of the Invention:
This invention relates to fortification of ruminant milk, in particular it concerns methods for enriching very long chain fatty acids in dairy milk in particular a feed additive and method for enriching nervonic acid in expressed milk of dairy cattle.
Background Of The Invention:
Nervonic acid (NA) or cis-15-tetracosenoic acid (24:1n-9) is a very long chain fatty acid (VLCFA - greater than or equal to 22 carbons). It has the common name nervonic acid because it is abundant in nerve tissue, particularly in the white matter of brain and in myelinated, peripheral nerves. Except for a few rare plants, nervonic acid is not found in plants in quantities higher than 0.01 % of total fatty acids. Nervonic acid is found from 10 to 100 fold higher concentration in animal tissues. In plants, if present, nervonic acid is found in triglyceride form;
in animals, nervonic acid is present as a major fatty acid of the phospholipid sphingomyelin.
Nervonic acid is a normal component of the diet of omnivorous humans.
Concentrated dietary sources of nervonic acid include human milk . Poor sources of nervonic acid are milk from ruminants, meat and plant foods. Nervonic acid is considered to be a conditionally essential nutrient, particularly in premature infants, formula fed babies and in patients with certain degenerative brain diseases like multiple sclerosis and adrenoleukodystrophy (Medical Hypotheses (1994) 42: 237-242). The main dietary form of nervonic acid is as sphingomyelin. Nervonic acid-containing sphingomyelin may have special physiological functions.
Sphingomyelin is being investigated as a nutraceutical for reducing the risk of gastrointestinal cancers and cardiovascular disease (Eur J Cancer Prev (2002) 11: 193-197; J
Nutr (1997) 127:1055-1060).
Dietary nervonic acid readily raises the nervonic acid content in the sphingomyelin of some tissues. Bettger et al. (Lipids (1997) 32: 51-55; Nutr Res (1996) 16: 1761-1765) have shown, using a rat bioassay, that dietary nervonic acid significantly elevates the nervonic acid content of sphingomyelin in liver, heart, skeletal muscle and adipose tissue in developing rats. Barre and Holub (Lipids (1992) 27: 315-320) have shown that human adults fed a dietary supplement rich in nervonic acid raises the nervonic acid content of platelet sphingomyelin.
Nervonic acid is a major fatty acid in the sphingomyelin fraction of milk from monogastric animals, including humans. However, the content of nervonic acid in the sphingomyelin in the milk of ruminants is roughly 10-fold lower than other VLCFA. It is not clear why the nervonic acid content is so much lower than other VLCFA in the milk of ruminants. The nervonic acid content in the milk of omnivorous, monogastric animals depends, in part, on the dietary intake of nervonic acid. Researchers have shown that feeding nervonic acid rich diets to rats or mice (model monogastric animals) will increase the nervonic acid of the sphingomyelin fraction of the expressed milk (J Nutr Biochem (2003) 14:160-165; Lipids (1998) 33:
993-1000).
There is commercial interest in the use of nervonic acid-rich lipids as dietary supplements. US 5,194,448 (owned by Croda International, in the United Kingdom), teaches the production and use of nervonic as triglyceride isolated from the Lunaria plant, and US 5,994,404 (also in the name of Croda International) teaches various nervonic acid compositions from plants, animals and/or microbial products, including, the seed oils of Cardamine gracea, Heliphila longifola, Thlaspi perfoliatum, Tropaeolum speciosum, Lunaria biennis, Lunaria annua and Malania oleifera; the moulds Neocallismastix frontalis, Erysiphe graminis and Sphaerotheca humuli; the bacterium Pseudomonas atlantica; the yeast Saccharomyces cerevisiae and the marine diatom Nitzschia cylindrus.
'There have been a number of reports on efforts to enrich bovine milk with various fatty acids. A recent review outlines a number of these (Ashes, J. R. et al.
(1997)). For example, methods have been developed to increase the level of omega-3 fatty acids in the flesh of beef cattle (US 5,290,573), sows (US 5,106,639; DE 3808885;
Taugbol, O.
et al., Zentralbl. Veterinarmed. A., (1993) 40(6): 437-443), poultry (US
5,012,761; JP
04271754; US 5,133,963; US 5,069,903), and eggs (KR 9311396; US 5,069,903).
DHA
has also been added as a dietary supplement to infant formula as discussed above, and milk. Sources of DHA for supplementing milk or infant formula include fish ' products, fatty acid containing microbial oils (US 5,374,657; US 5,397,591;
US
5,4"07,957), or fatty acids extracted from a mixture of egg yolk and coconut oil (US
4,670,285).
Indeed, researchers have been able to increase DHA content in the expressed milk of humans (Harris, W.S. et al., Am. J. Clin. Nutr. 40(4): 780-785,1984;
Henderson, R.A., Lipids, 27(11): 863-869,1992; US 5,069,903), sows (Taugbol, O. et al., Zentralbl, Veterinarmed. A. 40(6): 437-443,1993), and rats (Yonekubo, A., et al. J. Nutr.
123(10):
1703-1708,1993). However, researchers have had difficulty obtaining significant levels of DHA in cow's milk. (Hebeisen, D.F., et al. Int. J. Vitam. Nutr.
Res., 63(3):
229-233,1993).
A method, a feed additive and a feed to increase DHA content in expressed milk of dairy cattle has been disclosed by one of the present inventors in US
5,932,257 and in Wright, T.; McBride, B. ; and Holub, B, World Rez~. Nutr. Diet 83:160-165,1998.
However, there are no reported studies of an attempt to enrich bovine milk or low-fat milk products with nervonic acid. Yet in light of the importance of nervonic acid in the diet it is clear that it is desirable to have a method of enriching milk products with nervonic acid.
SUMMARY OF THE INVENTION
Surprisingly, the present inventors have found the use of a feed additive comprising feather meal and a source of NA such as oil extracted from the seeds of the plant Lunaria annua is able to dramatically increase the content of VLCFA
in the expressed milk of a ruminant.
The NA is in the whey protein fraction of expressed milk of dairy cattle fed a novel feed additive containing NA, and inhibitors of microbial degradation of NA in the rumen of the cattle. The feed additive does not affect the ability of the cattle to digest the feed by normal symbiotic digestion. The feed additive is also palatable to the cattle, and therefore food consumption is not decreased. As a result, the health of the cattle is maintained and their productivity is not reduced. In addition, because NA is present in the whey protein fraction, the expressed milk is suitable for the production of low-fat dairy products and milk protein fractions.
The present inventors found that when dairy cattle are fed the feed additive throughout lactation, the levels of NA in the whey protein fraction of expressed milk are between 26% and 34% of the fatty acids in sphingomyelin (see table 3).
Broadly stated, the present invention relates to a feed additive for dairy cattle which comprises a source of NA and inhibitors of microbial degradation of NA
in the rumen of dairy cattle. The source of NA and the inhibitors of microbial degradation of NA are present in the feed in an amount sufficient to enhance the concentration of NA in the milk of ruminants, preferably dairy cows, fed with feed containing the additive. In an embodiment of the invention, the source of NA
is the seed oil of Lunaria annua and Lunaria biennis (the Honesty plant). Various methods of extracting seed oils from oil bearing seeds are well known to those skilled in the art (see "Baileys Industrial Oil and Fat Products" ed. D. Swern, Vol. 2, pages 175 et.
seq. 4th Edition, Pub 1982, John Wiley & Sons Inc.).
The inhibitors of microbial degradation comprise feather meal. In one embodiment the inhibitors of microbial degradation consists of feather meal.
Preferably the feed additive comprises an amount of feather meal sufficient to increase the concentration of NA in milk from cattle consuming the feed additive.
The invention also contemplates a feed containing the feed additive.
The invention also relates to a method of producing milk in dairy cattle which is enriched for NA comprising feeding dairy cattle a diet containing a feed additive of the invention for a period of time longer than one day and preferably for at least about 7 days and milking the dairy cattle to obtain milk enriched for NA.
The invention further relates to expressed milk from dairy cattle enriched with NA which is produced by feeding cattle a diet containing a feed additive of the invention for a period of at least two days, preferably at least about 7 days, and milking the dairy cattle to obtain milk enriched for NA. The expressed whole milk of the invention preferably containing about 16% of NA in the sphingomyelin fraction (see Table 2).
The invention further relates to a NA-enriched dairy product produced using the expressed milk of the invention. The NA-enriched dairy product is preferably selected from the group consisting of cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant milk and butter. In another preferred embodiment the dairy product is a low-fat dairy product. In yet another preferred embodiment the dairy product is a milk protein product. The milk protein products is preferably selected from the group consisting of dried whey protein concentrate, dried whey, dried skim milk, dried milk protein, dried buttermilk, casein, acid casein, a protein drink, a protein bar or a protein supplement.
In one embodiment, the invention relates to the use of feather meal as an inhibitor of microbial degradation of NA in the rumen of dairy cattle in an amount sufficient to increase the concentration of NA in milk. In another embodiment the feather meal is added to the feed for dairy cattle.
Other features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples while indicating preferred embodiments of the invention are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
Table 1. Effect of a Nervonic Acid Supplement (Canstar) on Feed Intake and Milk Production of Dairy Cows' Parameter MeasuredDay Day Day Day 7 Olive CanstarOlive Canstar Dry Matter Intake (k /da ) 16.5 16.8 16.7 16.4 Milk Production (k /da ) 28.0 27.0 25.0 22.0 Milk Protein Content (%) 3.52 3.46 3.60 3.51 Milk Fat Content (%) 2.85 2.60 3.10 2.90 * Values represent the means of n=6 cows. Values within a parameter are not significantly altered by the nervonic acid supplement (Canstar) or by the number of days on the experiment (Student's t-test, P > 0.05).
Table 2. Effect of a Nervonic Acid Supplement (Canstar) on the Fatty Acid Composition of Sphingomyelin from Whole Milk of Dairy Cows*
Fatty Acid Day Day Day Day (mol%) 1 1 7 7 Olive Canstar Olive Canstar 16:0 34.5(3.2) --28.4(3.3) 25.2(11.8) 29.4(8.4) 18:0 9.7 (1.0) 8.9 (1.0) 7.8 (3.2) 11.5(3.6) 18:1 3.7 (0.4) 2.7 (1.2) 4.7 (2.2) 8.7 (3.6) 22:0 18.8(2.4) 21.6 (3.3) 24.9(6.0) 10.7(2.3) 23:0 15.8(3.6) 20.4 (3.8) 15.6(5.3) 8.8 (3.2) 24:0 14.4(2.8) 14.7 (2.8) 16.4(9.4) 7.4 (2.6) 24:1 1.3 (0.2) 1.0 (0.8) 1.9 (0.5) 1 (7.1)a 6.0 * Values are means (standard deviations) of n = 6 cows. Value for nervonic acid (24:1) on day 7 of the trial (Canstar Day 7) is significantly higher than that of the Olive Day 7 and the Day 1 controls (Student's t-test, P< 0.001).
a The range of NA (24:1) values for Canstar Day 7 are 11.5 to 25.0%
Table 3. Effect of a Nervonic Acid Supplement (Canstar) on the Composition of Very Long Chain Fatty Acids in Sphingomyelin of Isolated Whey Protein from Milk*
Very Long Chain Fatty Olive #1 Olive #2 Canstar #1 Canstar *2 Acid (mol%) 22:0 41.9 69.2 30.8 41.9 23:0 30.4 16.6 24.0 12.8 24:0 25.7 14.0 18.4 11.2 24:1 2.0 0.1 26.7 34.0 *Values for the nervonic acid (24:1) content of the sphingomyelin in the whey protein fraction of cows milk, day 7 of the experiment. Nervonic acid content is increased over 10-fold by the Canstar supplement.
As hereinbefore mentioned, the present invention relates to a feed additive for dairy cattle which comprises a source of NA, and inhibitors of microbial degradation of NA in the rumen of dairy cattle. Preferably, these components are present in an amount sufficient to increase the concentration of NA in the whey protein fraction of the expressed milk of dairy cows fed a diet containing the additive. "An amount sufficient" as used herein would be understood by a person skilled in the art to be the amount which results in an increased concentration of NA in milk.
The NA in the feed additive may be a NA concentrate containing 4 to 98% NA, or it may be a component of an extract from a source known to contain NA; for example, it may be a component of an extract derived from fish, such as fish meal, or from myelinated nervous tissue from other animals. NA concentrates may be obtained from commercial sources for example, CROSSENTIAL N25. Indeed any source of NA which is palatable to cattle and which does not compromise the taste qualities of milk produced may be used.
According to an embodiment of the invention, the feed additive comprises feather meal and a Lunaria oil based product containing NA. The NA may be made from the seeds ( or any other part ) of the plant Lunaria annua, and contains an amount of NA which will provide about 0.1 to 0.8% NA by weight of the feed supplement as described herein. In another embodiment there is 0.2 to 0.6%NA
by weight of the feed supplement, and in yet another embodiment the nervonic acid content of the supplement is 0.375%.
The feather meal may be made from the feathers of broiler chickens, although the feathers of any foul including turkeys and geese, may be used. Feather meal may be selected which has 93.9% protein, and a total fat content of 2.6% (dry measurement basis). It will be appreciated that the fish meal and feather meal may be obtained from commercial sources for example, Ralston Purina, Shurgain, Masterfields, ADM & Purina Mills.
In a preferred embodiment, the feed additive contains 5 to 60% feather meal, preferably 10 to 20% by weight of the total feed additive; and 5 to 20 % of Canstar oil preferably 8 to 15% by weight of the total feed additive.
In a more preferred embodiment, the feed additive contains 15% feather meal by weight of the total feed additive and 10% of Canstar oil by weight of the total feed additive.
The feed additive may contain a carbohydrate fraction such as soft white wheat or corn. Preferably the feed additive contains soft white wheat.
The feed additive may be added to a basal feed which may contain a carbohydrate fraction, a protein fraction, a lipid fraction and/or a vitamin/mineral fraction. Examples of components in carbohydrate fractions include corn silage, alfalfa hay, Timothy hay, wheat straw, barley grain, canola meal, oat grain, mixed straw, and corn. Typical components in the vitamin/mineral fraction include magnesium oxide, limestone, potassium chloride, sodium chloride, and a trace mineral supplement, containing zinc, copper, manganese, selenium, vitamins A, D
and E. Commercial sources of these components are Ralston Purina, Shurgain, Masterfields, ADM & Purina Mills. Typical protein feeds include soybean meal, corn gluten meal and distillers dried grains. Typical lipid feeds including vegetable fat sources (Church & Dwight Co.).
The feed containing the feed additive may be pelleted for feeding to dairy cattle, or the feed additive and basal feed may be fed to the cattle in a total mixed ration or as separate ingredients.
In an embodiment of the invention a feed is provided comprising (a) a basal feed containing 2 to 10%, preferably 5-7%, mixed straw; 40 to 55%, preferably 45 to 47%
corn silage; 35 to 50%, preferably 42 to 45% high moisture corn, and 2-4% of a vitamin/mineral fraction, each percentage being a percentage of the total weight of the basal feed; and (b) a feed additive comprising 1 to 7% feather meal, 0.5 to 3.5%
Canstar oil, each percentage being a percentage of the total weight of the feed (dry weight basis). In a preferred embodiment of the invention the feed additive comprises about 2.0%, feather meal and about 1.5% Canstar oil , each percentage being a percentage of the total weight of the feed (dry weight basis). In a more preferred embodiment, the feed additive comprises about 1.0% feather meal and about 1.0% Canstar oil, each percentage being a percentage of the total weight of the feed (dry weight basis) As illustrated in the examples herein, the use of a feed additive comprising feather meal and a source of NA such as oil extracted from the seeds of the plant Lunaria annua provides milk with an increased concentration of NA.
Accordingly, the invention also relates to the use of feather meal as an inhibitor of microbial degradation of NA in the rumen of dairy cattle. The use of feather meal is characterized in that the feather meal is added to the feed for dairy cattle.
According to a preferred embodiment, the use is characterized in that an amount sufficient to enrich milk from dairy cattle with NA is added to feed additive, which itself may be added to cattle feed.
The invention also relates to a method of producing milk in dairy cattle which is enriched for NA comprising feeding the dairy cattle a diet containing the feed additive of the invention for a period of at least 2 days, and preferable about 7 days, and milking the dairy cattle to obtain milk enriched for NA. The cattle may be fed a basal feed containing the feed additive, or the feed additive and basal feed may be fed to the cattle in a total mixed ration or as separate ingredients.
The cattle are preferably fed throughout lactation and for at least two days, preferably for at least 7 days, and more preferably 12 days, in order to obtain expressed milk with a NA content which is greater than or equal to 16% of the total fatty acids of sphingomyelin in whole milk. Whey protein isolated from such milk will have greater than 26% of the VLCFA in sphingomylein as NA Typically the cattle are fed the feed additive (which may be part of a basal feed) twice daily. The amount of feed additive given to the cattle ranges from 600 grams to 7.5 kg per animal per day.
The method of the invention for producing expressed milk enriched for NA may be applied to any breed of dairy cattle, for example, Ayshire, Guernsey, Holstein, Jersey, Brown Swiss, Dutch Belted, Canadienne and Milking Shorthorn. It will be appreciated that the method may also be applied to other ruminant species such as sheep and goats to produce expressed milk enriched for NA.
The expressed milk from dairy cattle enriched for NA produced by a method of the invention contains levels of NA typically in the range of 11.5 to 25% (see Table 2) of the fatty acids in sphingomyelin in the milk. These levels are as high as, or higher than NA levels found in human expressed milk which are typically in the range of 15.5 to 20% (Am J Clin Nutr (1984) 40: 1103-1119). The taste of the milk enriched for NA produced by the method of the invention is not altered and it is therefore suitable for human consumption. Further, the method achieves these concentrations of NA in milk using only feather meal as the inhibitor of NA
degradation in the rumen.
It will be appreciated that other NA-enriched dairy products can be produced by using the method described herein. For example, cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant formula, and butter enriched for NA may be produced using the method of the invention. In addition, low-fat dairy products and milk protein products can also be produced by using the method described herein. For example, low-fat , cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant formula, and butter enriched for NA may be produced using the method of the invention. Examples of milk protein products enriched for NA
include whey protein supplements for athletes and whey protein-fortified beverages and high protein bars.
The milk dairy products and milk protein products of the present invention enriched for NA are nutritionally superior products to conventional milk products.
The milk and dairy products may be of particular benefit with respect to the various factors for preventing degenerative brain diseases like multiple sclerosis and adrenoleukodystrophy. The benefits of the invention also extend beyond the production of NA-enriched food products for human consumption. For example, dairy cattle that are fed with a feed of the invention can be expected to exhibit improved health effects associated with NA , since NA is an essential nutrient for growth, development, and neuronal functioning in animals.
The use of feathermeal as a feed additive to 'mask' the impact of the ruminant GI tract on dietary fatty acid digestion, metabolism and absorption and/or to override the program of directed synthesis of certain molecular species of sphingomyelin in milk is just one manifestation of this NA enrichment technology.
The feathermeal/NA may also be administered within microcapsules to be ingested orally or in the coating of microcapsules. Suitable materials for the microcapsules include gelatin, gum arabic, food starch, malto dextrin, lactose, dextrin, corn syrup solids, the materials disclosed in U.S. Pat. No. 3,455,838 (which is hereby incorporated by reference), and mixtures thereof. The feathermeal/ NA may also be entrapped in an edible solid vehicle composed of a film forming material or a plasticizing agent. Alternatively the feathermeal/NA could be gavaged or freely ingested in a liquefied form. Other vehicles may also be used for the delivery of feathermeal/NA. In addition, the feathermeal and the NA may be administered in separate capsules or vehicles.
to Another aspect of the current invention includes inhibitors other than feathermeal, which can facilitate the enrichment of nervonic acid in milk of ruminants.
These inhibitors or facilitating agents of NA enrichment may also be administered either separately or in combination with NA as described above. The physical form of the inhibitors or facilitating agents for NA can be a solid, liquid or gas. They can also be a microbe (living organism) or a mechanical device and they can be administered by oral, transdermal, or inhalation routes.
The following non-limiting examples are illustrative of the present invention:
EXAMPLES
Example 1 NA Enrichment of Milk by Feeding a Lipid and Protein Rich Supplement containing NA and Feather Meal Experimental Design Eighteen Holstein cows housed at the Elora Research Station, University of Guelph, were used in this experiment and divided equally into three groups;
control, B, and C.
Throughout the study, cows were fed and milked twice daily. The diet of the control group was the total mixed ration (TMR) routinely fed at the research facility.
On a percent as-fed basis, it consisted of about: 2.9% hay, 16.7% haylage, 57.8% corn silage, 12.1% high-moisture corn, 10.6% commercial supplement pellet, as well as a mineral premix (see appendix H). The basal diet of groups B and C on a percent as-fed basis was: 4% straw, 60.6% corn silage, 33.6% high-moisture corn. A
mineral premix and 2kg of non-pellet supplement were top-dressed onto the basal diet of these two groups.
During days 1 to 14 of the trial, groups B and C received the same supplement (supplement A). Supplement A contained the following ingredients on a percent as-fed basis: 25% wheat, 15% feather meal, and 60% canola meal. On days 15 to 21, supplements B and C were fed to groups B and C respectively. Supplement B
contained (percent as-fed basis): 25% wheat, 15% feather meal, 50% canola meal, and 10% olive oil. Supplement C was identical to supplement B, only the 10% olive oil was replaced with 10% canstar oil. The canstar oil was composed of 85% canola oil and 15% crossential N25, an oil containing 25% nervonic acid purchased from Croda Oleochemicals, UK. During days 22 to 28 of the trial, groups B and C received supplement A. On the final 7 days of the trial (day 29 to 35), all eighteen cows were fed the routine TMR.
Milk samples were collected from the morning and evening milkings every Monday, Wednesday, and Friday on days 1 to 14 and 22 to 35, and daily during days 15 to 21. The samples obtained on these days were frozen at B20°C for fatty acid analysis. After consuming the oil-containing supplement for 7 consecutive days (day 22), two randomly selected cows from groups B and C were segregated during the morning milking. Their milk was collected and stored separately for protein fraction analysis.
Sphingomyelin Fatty Acid Analysis of Whole Milk For each sample, 0.12 g of freeze-dried whole milk was placed into a pre-rinsed test tube. 4.0 ml of 2:1 chloroform:methanol and 25 ~1 of 5% BHT in 2:1 chloroform:methanol were added to each test tube. NZ was added and the samples were vortexed for 1 minute. Test tubes were stored at 4 ~C for 2 hours and centrifuged at 2500 RPM for 20 minutes. After pipetting the supernatant from each test tube into other pre-rinsed test tubes, 0.8 ml of 88% KCl in distilled water were added to the samples. N2 was added and the samples were vortexed for 30 seconds followed by centrifugation at 2500 RPM for 20 minutes and placed in the freezer overnight. Once the lower phase was transferred from each sample into pre-rinsed test tubes, the upper phase was discarded.
Each sample was dried completely under Nz and 201 of 2:1 chloroform:methanol was added. Test tubes were capped immediately to avoid evaporation. In the case of evaporation, an additional 201 of 2:1 chloroform:methanol was added. Samples were vortexed for 15 seconds and the thin layer chromatography (TLC) plate was spotted using a 101 Hamilton syringe.
~2 After the last spot dried, the TLC plate was placed in the TLC tank containing chloroform:methanol:acetic acid:H20 (50:37.5:3.5:5:2). The plate ran until it was approximately one inch from the top of the plate and was removed from the tank and dried under a fume hood. The plate was then sprayed with 0.1% 8-anilinonapthalene-1-sulphonic acid (ANS) and viewed under UV light to identify the phospholipid bands. The sphingomyelin and phosphatidylcholine bands were marked, and scraped horizontally with a blade into disposable. 3.0 ml of 6%
HzS04 in methanol and 5.0 ~1 of 17:0 standard were added to these test tubes and they were filled with Nz and vortexed for 3 seconds. Samples were placed in an oven at 80°C for 16 hours to transmethylate.
After removing the samples from the oven, they were cooled to room temperature and 2.5 ml of hexane was added. After filling with Nz and vortexing for 15 seconds, 1.0 ml of deionized water was added and each test tube was filled with Nz again and vortexed for an additional 5 seconds.
The test tubes were then centrifuged at 1000 RPM for 1 minute. The top layer of each sample was transferred removed into minivials, filled with Nz, and stored in the freezer.
Each sample was dried completely under Nz. 3 ~l of CSz were added before injecting 1 ~l into the gas chromatograph (Hewlett-Packard 5890A). The amount of each fatty acid was identified by the area under each peak.
The results of this experiment are shown in Tables 1 and 2 , below.
Table 1. Effect of a Nervonic Acid Supplement (Canstar) on Feed Intake and Milk Production of Dairy Cows*
Parameter MeasuredDay Day 1 Day Day OliveCanstar Olive Canstar Dry Matter Intake (k /da ) 16.5 16.8 16.7 16.4 Milk Production (k /da ) 28.0 27.0 25.0 22.0 Milk Protein Content (%) 3.52 3.46 3.60 3.51 Milk Fat Content (%) 2.85 2.60 3.10 2.90 * Values represent the means of n=6 cows. Values within a parameter are not significantly altered by the nervonic acid supplement (Canstar) or by the number of days on the experiment (Student's t-test, P > 0.05).
Table 2. Effect of a Nervonic Acid Supplement (Canstar) on the Fatty Acid Composition of Sphingomyelin from Whole Milk of Dairy Cows*
Fatty Acid Day 1 Day 1 Day 7 Day 7 (mol%) Olive Canstar Olive Canstar 16:0 34.5 (3.2) 28.4 (3.3) 25.2 (11.8) 29.4 (8.4) 18:0 9.7 (1.0) 8.9 (1.0) 7.8 (3.2) 11.5 (3.6) 18:1 3.7 (0.4) 2.7 (1.2) 4.7 (2.2) 8.7 (3.6) 22:0 18.8 (2.4) 21.6 (3.3) 24.9 (6.0) 10.7 (2.3) 23:0 15.8 (3.6 20.4 (3.8 15.6 (5.3) 8.8 (3.2 24:0 14.4 (2.8 14.7 (2.8) 16.4 (9.4 7.4 (2.6 24:1 1.3 0.2) I .0 (0.8 1.9 (0.5 16.0 (7.1 )a * Values are means (standard deviations) of n = 6 cows. Value for nervonic acid (24:1) on day 7 of the trial (Canstar Day 7) is significantly higher than that of the Olive Day 7 and the Day 1 controls (Student's t-test, P< 0.001).
a The range of NA (24:1) values for Canstar Day 7 are 11.5 to 25.0%
The data clearly indicate that the supplemented cows had higher levels of nervonic acid than standard bovine milk. The control samples in this trial contained levels similar to the levels of nervonic acid found in commercially available, pasteurized, homogenized milk. The milk obtained from the cows fed the nutritional supplement had nervonic acid levels that were more like human milk than standard cow's milk.
No ill effects were observed in animals consuming the feed supplement. The NA content (16% of the fatty acids in the milk sphingomyelin) increased in expressed milk within 3 days after the cows first started consuming the feed supplement containing NA. Milk production and feed consumption were similar to production/consumption observed when the cows were fed conventional feeds.
Example 2 NA Enrichment of Whey Protein from Milk by Feeding a Lipid and Protein Rich Supplement containing NA and Feather Meal Experimental Design Same as for Example 1.
Skimming and Pasteurization Milk Samples Aliquots of each milk sample collected on day 22 were skimmed by centrifugation at 13,000 rpm for 30 minutes at 4°C. The remaining milk was skimmed by allowing the cream to rise to the surface of the milk, and removing the cream with a beaker. This step was repeated four times to ensure most of the fat was removed.
All samples were pasteurized according to Ontario government standards by heating to 72°C for not less than 16 seconds.
Preparation of Whey Whey was separated from the skim milk by first warming the sample to 31~C
using a water bath, adding a rennet-water solution, and allowing the milk to curd for 50 minutes. Once a curd was formed, it was gently agitated and gradually heated to 39°C. After holding this temperature for 60 minutes, the whey was removed.
Concentrating the Whey Proteins The whey proteins were concentrated via ultrafiltration (UF) at 4 °C.
Membranes with a pore size suitable to retain molecules with a molecular weight greater than 10,000 were used. The nitrogen pressure required for the procedure was maintained at 40 psi.
Aliquots of whey from each sample were injected into the UF. When 3m1 of whey remained in the UF, water was injected to further concentrate the whey proteins (diafiltration). The whey remaining in the OF (retentate) is highly concentrated with whey proteins. Samples were placed in the Dura-Dry MP freeze dryer for 42 hours.
Sphingomyelin Fatty Acid Analysis of Isolated Whey Protein Same procedure , as for whole milk (example 1) except started with 0.15g of freeze-dried whey protein instead of whole milk.
Results The results of this experiment are shown in Table 3, below.
Table 3. Effect of a Nervonic Acid Supplement (Canstar) on the Composition of Very Long Chain Fatty Acids in Sphingomyelin of Isolated Whey Protein from Milk*
Very Long Chain Fatty Olive #1 Olive #2 Canstar #1 Canstar *2 Acid (mol%) 22:0 41.9 69.2 30.8 41.9 23:0 30.4 16.6 24.0 12.8 24:0 25.7 14.0 18.4 11.2 24:1 2.0 0.1 26.7 34.0 *Values for the nervonic acid (24:1) content of the sphingomyelin in the whey protein fraction of cows milk, day 7 of the experiment. Nervonic acid content is increased over 10-fold by the Canstar supplement..
These results clearly demonstrate that isolated whey protein from cows fed the NA supplement (Canstar) is enriched in nervonyl-sphingomyelin relative to whey obtained from the milk of unsupplemented (Olive) cows.
Having illustrated and described the principles of the invention in a preferred embodiment, it should be appreciated to those skilled in the art that the invention can be modified in arrangement and detail without departure from such principles.
We claim all modifications coming within the scope of the following claims.
All publications, patents and patent applications are herein incorporated by reference in their entirety to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated by reference in its entirety.
EXPRESSED MILK OF RUMINANTS
Field of the Invention:
This invention relates to fortification of ruminant milk, in particular it concerns methods for enriching very long chain fatty acids in dairy milk in particular a feed additive and method for enriching nervonic acid in expressed milk of dairy cattle.
Background Of The Invention:
Nervonic acid (NA) or cis-15-tetracosenoic acid (24:1n-9) is a very long chain fatty acid (VLCFA - greater than or equal to 22 carbons). It has the common name nervonic acid because it is abundant in nerve tissue, particularly in the white matter of brain and in myelinated, peripheral nerves. Except for a few rare plants, nervonic acid is not found in plants in quantities higher than 0.01 % of total fatty acids. Nervonic acid is found from 10 to 100 fold higher concentration in animal tissues. In plants, if present, nervonic acid is found in triglyceride form;
in animals, nervonic acid is present as a major fatty acid of the phospholipid sphingomyelin.
Nervonic acid is a normal component of the diet of omnivorous humans.
Concentrated dietary sources of nervonic acid include human milk . Poor sources of nervonic acid are milk from ruminants, meat and plant foods. Nervonic acid is considered to be a conditionally essential nutrient, particularly in premature infants, formula fed babies and in patients with certain degenerative brain diseases like multiple sclerosis and adrenoleukodystrophy (Medical Hypotheses (1994) 42: 237-242). The main dietary form of nervonic acid is as sphingomyelin. Nervonic acid-containing sphingomyelin may have special physiological functions.
Sphingomyelin is being investigated as a nutraceutical for reducing the risk of gastrointestinal cancers and cardiovascular disease (Eur J Cancer Prev (2002) 11: 193-197; J
Nutr (1997) 127:1055-1060).
Dietary nervonic acid readily raises the nervonic acid content in the sphingomyelin of some tissues. Bettger et al. (Lipids (1997) 32: 51-55; Nutr Res (1996) 16: 1761-1765) have shown, using a rat bioassay, that dietary nervonic acid significantly elevates the nervonic acid content of sphingomyelin in liver, heart, skeletal muscle and adipose tissue in developing rats. Barre and Holub (Lipids (1992) 27: 315-320) have shown that human adults fed a dietary supplement rich in nervonic acid raises the nervonic acid content of platelet sphingomyelin.
Nervonic acid is a major fatty acid in the sphingomyelin fraction of milk from monogastric animals, including humans. However, the content of nervonic acid in the sphingomyelin in the milk of ruminants is roughly 10-fold lower than other VLCFA. It is not clear why the nervonic acid content is so much lower than other VLCFA in the milk of ruminants. The nervonic acid content in the milk of omnivorous, monogastric animals depends, in part, on the dietary intake of nervonic acid. Researchers have shown that feeding nervonic acid rich diets to rats or mice (model monogastric animals) will increase the nervonic acid of the sphingomyelin fraction of the expressed milk (J Nutr Biochem (2003) 14:160-165; Lipids (1998) 33:
993-1000).
There is commercial interest in the use of nervonic acid-rich lipids as dietary supplements. US 5,194,448 (owned by Croda International, in the United Kingdom), teaches the production and use of nervonic as triglyceride isolated from the Lunaria plant, and US 5,994,404 (also in the name of Croda International) teaches various nervonic acid compositions from plants, animals and/or microbial products, including, the seed oils of Cardamine gracea, Heliphila longifola, Thlaspi perfoliatum, Tropaeolum speciosum, Lunaria biennis, Lunaria annua and Malania oleifera; the moulds Neocallismastix frontalis, Erysiphe graminis and Sphaerotheca humuli; the bacterium Pseudomonas atlantica; the yeast Saccharomyces cerevisiae and the marine diatom Nitzschia cylindrus.
'There have been a number of reports on efforts to enrich bovine milk with various fatty acids. A recent review outlines a number of these (Ashes, J. R. et al.
(1997)). For example, methods have been developed to increase the level of omega-3 fatty acids in the flesh of beef cattle (US 5,290,573), sows (US 5,106,639; DE 3808885;
Taugbol, O.
et al., Zentralbl. Veterinarmed. A., (1993) 40(6): 437-443), poultry (US
5,012,761; JP
04271754; US 5,133,963; US 5,069,903), and eggs (KR 9311396; US 5,069,903).
DHA
has also been added as a dietary supplement to infant formula as discussed above, and milk. Sources of DHA for supplementing milk or infant formula include fish ' products, fatty acid containing microbial oils (US 5,374,657; US 5,397,591;
US
5,4"07,957), or fatty acids extracted from a mixture of egg yolk and coconut oil (US
4,670,285).
Indeed, researchers have been able to increase DHA content in the expressed milk of humans (Harris, W.S. et al., Am. J. Clin. Nutr. 40(4): 780-785,1984;
Henderson, R.A., Lipids, 27(11): 863-869,1992; US 5,069,903), sows (Taugbol, O. et al., Zentralbl, Veterinarmed. A. 40(6): 437-443,1993), and rats (Yonekubo, A., et al. J. Nutr.
123(10):
1703-1708,1993). However, researchers have had difficulty obtaining significant levels of DHA in cow's milk. (Hebeisen, D.F., et al. Int. J. Vitam. Nutr.
Res., 63(3):
229-233,1993).
A method, a feed additive and a feed to increase DHA content in expressed milk of dairy cattle has been disclosed by one of the present inventors in US
5,932,257 and in Wright, T.; McBride, B. ; and Holub, B, World Rez~. Nutr. Diet 83:160-165,1998.
However, there are no reported studies of an attempt to enrich bovine milk or low-fat milk products with nervonic acid. Yet in light of the importance of nervonic acid in the diet it is clear that it is desirable to have a method of enriching milk products with nervonic acid.
SUMMARY OF THE INVENTION
Surprisingly, the present inventors have found the use of a feed additive comprising feather meal and a source of NA such as oil extracted from the seeds of the plant Lunaria annua is able to dramatically increase the content of VLCFA
in the expressed milk of a ruminant.
The NA is in the whey protein fraction of expressed milk of dairy cattle fed a novel feed additive containing NA, and inhibitors of microbial degradation of NA in the rumen of the cattle. The feed additive does not affect the ability of the cattle to digest the feed by normal symbiotic digestion. The feed additive is also palatable to the cattle, and therefore food consumption is not decreased. As a result, the health of the cattle is maintained and their productivity is not reduced. In addition, because NA is present in the whey protein fraction, the expressed milk is suitable for the production of low-fat dairy products and milk protein fractions.
The present inventors found that when dairy cattle are fed the feed additive throughout lactation, the levels of NA in the whey protein fraction of expressed milk are between 26% and 34% of the fatty acids in sphingomyelin (see table 3).
Broadly stated, the present invention relates to a feed additive for dairy cattle which comprises a source of NA and inhibitors of microbial degradation of NA
in the rumen of dairy cattle. The source of NA and the inhibitors of microbial degradation of NA are present in the feed in an amount sufficient to enhance the concentration of NA in the milk of ruminants, preferably dairy cows, fed with feed containing the additive. In an embodiment of the invention, the source of NA
is the seed oil of Lunaria annua and Lunaria biennis (the Honesty plant). Various methods of extracting seed oils from oil bearing seeds are well known to those skilled in the art (see "Baileys Industrial Oil and Fat Products" ed. D. Swern, Vol. 2, pages 175 et.
seq. 4th Edition, Pub 1982, John Wiley & Sons Inc.).
The inhibitors of microbial degradation comprise feather meal. In one embodiment the inhibitors of microbial degradation consists of feather meal.
Preferably the feed additive comprises an amount of feather meal sufficient to increase the concentration of NA in milk from cattle consuming the feed additive.
The invention also contemplates a feed containing the feed additive.
The invention also relates to a method of producing milk in dairy cattle which is enriched for NA comprising feeding dairy cattle a diet containing a feed additive of the invention for a period of time longer than one day and preferably for at least about 7 days and milking the dairy cattle to obtain milk enriched for NA.
The invention further relates to expressed milk from dairy cattle enriched with NA which is produced by feeding cattle a diet containing a feed additive of the invention for a period of at least two days, preferably at least about 7 days, and milking the dairy cattle to obtain milk enriched for NA. The expressed whole milk of the invention preferably containing about 16% of NA in the sphingomyelin fraction (see Table 2).
The invention further relates to a NA-enriched dairy product produced using the expressed milk of the invention. The NA-enriched dairy product is preferably selected from the group consisting of cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant milk and butter. In another preferred embodiment the dairy product is a low-fat dairy product. In yet another preferred embodiment the dairy product is a milk protein product. The milk protein products is preferably selected from the group consisting of dried whey protein concentrate, dried whey, dried skim milk, dried milk protein, dried buttermilk, casein, acid casein, a protein drink, a protein bar or a protein supplement.
In one embodiment, the invention relates to the use of feather meal as an inhibitor of microbial degradation of NA in the rumen of dairy cattle in an amount sufficient to increase the concentration of NA in milk. In another embodiment the feather meal is added to the feed for dairy cattle.
Other features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples while indicating preferred embodiments of the invention are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
Table 1. Effect of a Nervonic Acid Supplement (Canstar) on Feed Intake and Milk Production of Dairy Cows' Parameter MeasuredDay Day Day Day 7 Olive CanstarOlive Canstar Dry Matter Intake (k /da ) 16.5 16.8 16.7 16.4 Milk Production (k /da ) 28.0 27.0 25.0 22.0 Milk Protein Content (%) 3.52 3.46 3.60 3.51 Milk Fat Content (%) 2.85 2.60 3.10 2.90 * Values represent the means of n=6 cows. Values within a parameter are not significantly altered by the nervonic acid supplement (Canstar) or by the number of days on the experiment (Student's t-test, P > 0.05).
Table 2. Effect of a Nervonic Acid Supplement (Canstar) on the Fatty Acid Composition of Sphingomyelin from Whole Milk of Dairy Cows*
Fatty Acid Day Day Day Day (mol%) 1 1 7 7 Olive Canstar Olive Canstar 16:0 34.5(3.2) --28.4(3.3) 25.2(11.8) 29.4(8.4) 18:0 9.7 (1.0) 8.9 (1.0) 7.8 (3.2) 11.5(3.6) 18:1 3.7 (0.4) 2.7 (1.2) 4.7 (2.2) 8.7 (3.6) 22:0 18.8(2.4) 21.6 (3.3) 24.9(6.0) 10.7(2.3) 23:0 15.8(3.6) 20.4 (3.8) 15.6(5.3) 8.8 (3.2) 24:0 14.4(2.8) 14.7 (2.8) 16.4(9.4) 7.4 (2.6) 24:1 1.3 (0.2) 1.0 (0.8) 1.9 (0.5) 1 (7.1)a 6.0 * Values are means (standard deviations) of n = 6 cows. Value for nervonic acid (24:1) on day 7 of the trial (Canstar Day 7) is significantly higher than that of the Olive Day 7 and the Day 1 controls (Student's t-test, P< 0.001).
a The range of NA (24:1) values for Canstar Day 7 are 11.5 to 25.0%
Table 3. Effect of a Nervonic Acid Supplement (Canstar) on the Composition of Very Long Chain Fatty Acids in Sphingomyelin of Isolated Whey Protein from Milk*
Very Long Chain Fatty Olive #1 Olive #2 Canstar #1 Canstar *2 Acid (mol%) 22:0 41.9 69.2 30.8 41.9 23:0 30.4 16.6 24.0 12.8 24:0 25.7 14.0 18.4 11.2 24:1 2.0 0.1 26.7 34.0 *Values for the nervonic acid (24:1) content of the sphingomyelin in the whey protein fraction of cows milk, day 7 of the experiment. Nervonic acid content is increased over 10-fold by the Canstar supplement.
As hereinbefore mentioned, the present invention relates to a feed additive for dairy cattle which comprises a source of NA, and inhibitors of microbial degradation of NA in the rumen of dairy cattle. Preferably, these components are present in an amount sufficient to increase the concentration of NA in the whey protein fraction of the expressed milk of dairy cows fed a diet containing the additive. "An amount sufficient" as used herein would be understood by a person skilled in the art to be the amount which results in an increased concentration of NA in milk.
The NA in the feed additive may be a NA concentrate containing 4 to 98% NA, or it may be a component of an extract from a source known to contain NA; for example, it may be a component of an extract derived from fish, such as fish meal, or from myelinated nervous tissue from other animals. NA concentrates may be obtained from commercial sources for example, CROSSENTIAL N25. Indeed any source of NA which is palatable to cattle and which does not compromise the taste qualities of milk produced may be used.
According to an embodiment of the invention, the feed additive comprises feather meal and a Lunaria oil based product containing NA. The NA may be made from the seeds ( or any other part ) of the plant Lunaria annua, and contains an amount of NA which will provide about 0.1 to 0.8% NA by weight of the feed supplement as described herein. In another embodiment there is 0.2 to 0.6%NA
by weight of the feed supplement, and in yet another embodiment the nervonic acid content of the supplement is 0.375%.
The feather meal may be made from the feathers of broiler chickens, although the feathers of any foul including turkeys and geese, may be used. Feather meal may be selected which has 93.9% protein, and a total fat content of 2.6% (dry measurement basis). It will be appreciated that the fish meal and feather meal may be obtained from commercial sources for example, Ralston Purina, Shurgain, Masterfields, ADM & Purina Mills.
In a preferred embodiment, the feed additive contains 5 to 60% feather meal, preferably 10 to 20% by weight of the total feed additive; and 5 to 20 % of Canstar oil preferably 8 to 15% by weight of the total feed additive.
In a more preferred embodiment, the feed additive contains 15% feather meal by weight of the total feed additive and 10% of Canstar oil by weight of the total feed additive.
The feed additive may contain a carbohydrate fraction such as soft white wheat or corn. Preferably the feed additive contains soft white wheat.
The feed additive may be added to a basal feed which may contain a carbohydrate fraction, a protein fraction, a lipid fraction and/or a vitamin/mineral fraction. Examples of components in carbohydrate fractions include corn silage, alfalfa hay, Timothy hay, wheat straw, barley grain, canola meal, oat grain, mixed straw, and corn. Typical components in the vitamin/mineral fraction include magnesium oxide, limestone, potassium chloride, sodium chloride, and a trace mineral supplement, containing zinc, copper, manganese, selenium, vitamins A, D
and E. Commercial sources of these components are Ralston Purina, Shurgain, Masterfields, ADM & Purina Mills. Typical protein feeds include soybean meal, corn gluten meal and distillers dried grains. Typical lipid feeds including vegetable fat sources (Church & Dwight Co.).
The feed containing the feed additive may be pelleted for feeding to dairy cattle, or the feed additive and basal feed may be fed to the cattle in a total mixed ration or as separate ingredients.
In an embodiment of the invention a feed is provided comprising (a) a basal feed containing 2 to 10%, preferably 5-7%, mixed straw; 40 to 55%, preferably 45 to 47%
corn silage; 35 to 50%, preferably 42 to 45% high moisture corn, and 2-4% of a vitamin/mineral fraction, each percentage being a percentage of the total weight of the basal feed; and (b) a feed additive comprising 1 to 7% feather meal, 0.5 to 3.5%
Canstar oil, each percentage being a percentage of the total weight of the feed (dry weight basis). In a preferred embodiment of the invention the feed additive comprises about 2.0%, feather meal and about 1.5% Canstar oil , each percentage being a percentage of the total weight of the feed (dry weight basis). In a more preferred embodiment, the feed additive comprises about 1.0% feather meal and about 1.0% Canstar oil, each percentage being a percentage of the total weight of the feed (dry weight basis) As illustrated in the examples herein, the use of a feed additive comprising feather meal and a source of NA such as oil extracted from the seeds of the plant Lunaria annua provides milk with an increased concentration of NA.
Accordingly, the invention also relates to the use of feather meal as an inhibitor of microbial degradation of NA in the rumen of dairy cattle. The use of feather meal is characterized in that the feather meal is added to the feed for dairy cattle.
According to a preferred embodiment, the use is characterized in that an amount sufficient to enrich milk from dairy cattle with NA is added to feed additive, which itself may be added to cattle feed.
The invention also relates to a method of producing milk in dairy cattle which is enriched for NA comprising feeding the dairy cattle a diet containing the feed additive of the invention for a period of at least 2 days, and preferable about 7 days, and milking the dairy cattle to obtain milk enriched for NA. The cattle may be fed a basal feed containing the feed additive, or the feed additive and basal feed may be fed to the cattle in a total mixed ration or as separate ingredients.
The cattle are preferably fed throughout lactation and for at least two days, preferably for at least 7 days, and more preferably 12 days, in order to obtain expressed milk with a NA content which is greater than or equal to 16% of the total fatty acids of sphingomyelin in whole milk. Whey protein isolated from such milk will have greater than 26% of the VLCFA in sphingomylein as NA Typically the cattle are fed the feed additive (which may be part of a basal feed) twice daily. The amount of feed additive given to the cattle ranges from 600 grams to 7.5 kg per animal per day.
The method of the invention for producing expressed milk enriched for NA may be applied to any breed of dairy cattle, for example, Ayshire, Guernsey, Holstein, Jersey, Brown Swiss, Dutch Belted, Canadienne and Milking Shorthorn. It will be appreciated that the method may also be applied to other ruminant species such as sheep and goats to produce expressed milk enriched for NA.
The expressed milk from dairy cattle enriched for NA produced by a method of the invention contains levels of NA typically in the range of 11.5 to 25% (see Table 2) of the fatty acids in sphingomyelin in the milk. These levels are as high as, or higher than NA levels found in human expressed milk which are typically in the range of 15.5 to 20% (Am J Clin Nutr (1984) 40: 1103-1119). The taste of the milk enriched for NA produced by the method of the invention is not altered and it is therefore suitable for human consumption. Further, the method achieves these concentrations of NA in milk using only feather meal as the inhibitor of NA
degradation in the rumen.
It will be appreciated that other NA-enriched dairy products can be produced by using the method described herein. For example, cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant formula, and butter enriched for NA may be produced using the method of the invention. In addition, low-fat dairy products and milk protein products can also be produced by using the method described herein. For example, low-fat , cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant formula, and butter enriched for NA may be produced using the method of the invention. Examples of milk protein products enriched for NA
include whey protein supplements for athletes and whey protein-fortified beverages and high protein bars.
The milk dairy products and milk protein products of the present invention enriched for NA are nutritionally superior products to conventional milk products.
The milk and dairy products may be of particular benefit with respect to the various factors for preventing degenerative brain diseases like multiple sclerosis and adrenoleukodystrophy. The benefits of the invention also extend beyond the production of NA-enriched food products for human consumption. For example, dairy cattle that are fed with a feed of the invention can be expected to exhibit improved health effects associated with NA , since NA is an essential nutrient for growth, development, and neuronal functioning in animals.
The use of feathermeal as a feed additive to 'mask' the impact of the ruminant GI tract on dietary fatty acid digestion, metabolism and absorption and/or to override the program of directed synthesis of certain molecular species of sphingomyelin in milk is just one manifestation of this NA enrichment technology.
The feathermeal/NA may also be administered within microcapsules to be ingested orally or in the coating of microcapsules. Suitable materials for the microcapsules include gelatin, gum arabic, food starch, malto dextrin, lactose, dextrin, corn syrup solids, the materials disclosed in U.S. Pat. No. 3,455,838 (which is hereby incorporated by reference), and mixtures thereof. The feathermeal/ NA may also be entrapped in an edible solid vehicle composed of a film forming material or a plasticizing agent. Alternatively the feathermeal/NA could be gavaged or freely ingested in a liquefied form. Other vehicles may also be used for the delivery of feathermeal/NA. In addition, the feathermeal and the NA may be administered in separate capsules or vehicles.
to Another aspect of the current invention includes inhibitors other than feathermeal, which can facilitate the enrichment of nervonic acid in milk of ruminants.
These inhibitors or facilitating agents of NA enrichment may also be administered either separately or in combination with NA as described above. The physical form of the inhibitors or facilitating agents for NA can be a solid, liquid or gas. They can also be a microbe (living organism) or a mechanical device and they can be administered by oral, transdermal, or inhalation routes.
The following non-limiting examples are illustrative of the present invention:
EXAMPLES
Example 1 NA Enrichment of Milk by Feeding a Lipid and Protein Rich Supplement containing NA and Feather Meal Experimental Design Eighteen Holstein cows housed at the Elora Research Station, University of Guelph, were used in this experiment and divided equally into three groups;
control, B, and C.
Throughout the study, cows were fed and milked twice daily. The diet of the control group was the total mixed ration (TMR) routinely fed at the research facility.
On a percent as-fed basis, it consisted of about: 2.9% hay, 16.7% haylage, 57.8% corn silage, 12.1% high-moisture corn, 10.6% commercial supplement pellet, as well as a mineral premix (see appendix H). The basal diet of groups B and C on a percent as-fed basis was: 4% straw, 60.6% corn silage, 33.6% high-moisture corn. A
mineral premix and 2kg of non-pellet supplement were top-dressed onto the basal diet of these two groups.
During days 1 to 14 of the trial, groups B and C received the same supplement (supplement A). Supplement A contained the following ingredients on a percent as-fed basis: 25% wheat, 15% feather meal, and 60% canola meal. On days 15 to 21, supplements B and C were fed to groups B and C respectively. Supplement B
contained (percent as-fed basis): 25% wheat, 15% feather meal, 50% canola meal, and 10% olive oil. Supplement C was identical to supplement B, only the 10% olive oil was replaced with 10% canstar oil. The canstar oil was composed of 85% canola oil and 15% crossential N25, an oil containing 25% nervonic acid purchased from Croda Oleochemicals, UK. During days 22 to 28 of the trial, groups B and C received supplement A. On the final 7 days of the trial (day 29 to 35), all eighteen cows were fed the routine TMR.
Milk samples were collected from the morning and evening milkings every Monday, Wednesday, and Friday on days 1 to 14 and 22 to 35, and daily during days 15 to 21. The samples obtained on these days were frozen at B20°C for fatty acid analysis. After consuming the oil-containing supplement for 7 consecutive days (day 22), two randomly selected cows from groups B and C were segregated during the morning milking. Their milk was collected and stored separately for protein fraction analysis.
Sphingomyelin Fatty Acid Analysis of Whole Milk For each sample, 0.12 g of freeze-dried whole milk was placed into a pre-rinsed test tube. 4.0 ml of 2:1 chloroform:methanol and 25 ~1 of 5% BHT in 2:1 chloroform:methanol were added to each test tube. NZ was added and the samples were vortexed for 1 minute. Test tubes were stored at 4 ~C for 2 hours and centrifuged at 2500 RPM for 20 minutes. After pipetting the supernatant from each test tube into other pre-rinsed test tubes, 0.8 ml of 88% KCl in distilled water were added to the samples. N2 was added and the samples were vortexed for 30 seconds followed by centrifugation at 2500 RPM for 20 minutes and placed in the freezer overnight. Once the lower phase was transferred from each sample into pre-rinsed test tubes, the upper phase was discarded.
Each sample was dried completely under Nz and 201 of 2:1 chloroform:methanol was added. Test tubes were capped immediately to avoid evaporation. In the case of evaporation, an additional 201 of 2:1 chloroform:methanol was added. Samples were vortexed for 15 seconds and the thin layer chromatography (TLC) plate was spotted using a 101 Hamilton syringe.
~2 After the last spot dried, the TLC plate was placed in the TLC tank containing chloroform:methanol:acetic acid:H20 (50:37.5:3.5:5:2). The plate ran until it was approximately one inch from the top of the plate and was removed from the tank and dried under a fume hood. The plate was then sprayed with 0.1% 8-anilinonapthalene-1-sulphonic acid (ANS) and viewed under UV light to identify the phospholipid bands. The sphingomyelin and phosphatidylcholine bands were marked, and scraped horizontally with a blade into disposable. 3.0 ml of 6%
HzS04 in methanol and 5.0 ~1 of 17:0 standard were added to these test tubes and they were filled with Nz and vortexed for 3 seconds. Samples were placed in an oven at 80°C for 16 hours to transmethylate.
After removing the samples from the oven, they were cooled to room temperature and 2.5 ml of hexane was added. After filling with Nz and vortexing for 15 seconds, 1.0 ml of deionized water was added and each test tube was filled with Nz again and vortexed for an additional 5 seconds.
The test tubes were then centrifuged at 1000 RPM for 1 minute. The top layer of each sample was transferred removed into minivials, filled with Nz, and stored in the freezer.
Each sample was dried completely under Nz. 3 ~l of CSz were added before injecting 1 ~l into the gas chromatograph (Hewlett-Packard 5890A). The amount of each fatty acid was identified by the area under each peak.
The results of this experiment are shown in Tables 1 and 2 , below.
Table 1. Effect of a Nervonic Acid Supplement (Canstar) on Feed Intake and Milk Production of Dairy Cows*
Parameter MeasuredDay Day 1 Day Day OliveCanstar Olive Canstar Dry Matter Intake (k /da ) 16.5 16.8 16.7 16.4 Milk Production (k /da ) 28.0 27.0 25.0 22.0 Milk Protein Content (%) 3.52 3.46 3.60 3.51 Milk Fat Content (%) 2.85 2.60 3.10 2.90 * Values represent the means of n=6 cows. Values within a parameter are not significantly altered by the nervonic acid supplement (Canstar) or by the number of days on the experiment (Student's t-test, P > 0.05).
Table 2. Effect of a Nervonic Acid Supplement (Canstar) on the Fatty Acid Composition of Sphingomyelin from Whole Milk of Dairy Cows*
Fatty Acid Day 1 Day 1 Day 7 Day 7 (mol%) Olive Canstar Olive Canstar 16:0 34.5 (3.2) 28.4 (3.3) 25.2 (11.8) 29.4 (8.4) 18:0 9.7 (1.0) 8.9 (1.0) 7.8 (3.2) 11.5 (3.6) 18:1 3.7 (0.4) 2.7 (1.2) 4.7 (2.2) 8.7 (3.6) 22:0 18.8 (2.4) 21.6 (3.3) 24.9 (6.0) 10.7 (2.3) 23:0 15.8 (3.6 20.4 (3.8 15.6 (5.3) 8.8 (3.2 24:0 14.4 (2.8 14.7 (2.8) 16.4 (9.4 7.4 (2.6 24:1 1.3 0.2) I .0 (0.8 1.9 (0.5 16.0 (7.1 )a * Values are means (standard deviations) of n = 6 cows. Value for nervonic acid (24:1) on day 7 of the trial (Canstar Day 7) is significantly higher than that of the Olive Day 7 and the Day 1 controls (Student's t-test, P< 0.001).
a The range of NA (24:1) values for Canstar Day 7 are 11.5 to 25.0%
The data clearly indicate that the supplemented cows had higher levels of nervonic acid than standard bovine milk. The control samples in this trial contained levels similar to the levels of nervonic acid found in commercially available, pasteurized, homogenized milk. The milk obtained from the cows fed the nutritional supplement had nervonic acid levels that were more like human milk than standard cow's milk.
No ill effects were observed in animals consuming the feed supplement. The NA content (16% of the fatty acids in the milk sphingomyelin) increased in expressed milk within 3 days after the cows first started consuming the feed supplement containing NA. Milk production and feed consumption were similar to production/consumption observed when the cows were fed conventional feeds.
Example 2 NA Enrichment of Whey Protein from Milk by Feeding a Lipid and Protein Rich Supplement containing NA and Feather Meal Experimental Design Same as for Example 1.
Skimming and Pasteurization Milk Samples Aliquots of each milk sample collected on day 22 were skimmed by centrifugation at 13,000 rpm for 30 minutes at 4°C. The remaining milk was skimmed by allowing the cream to rise to the surface of the milk, and removing the cream with a beaker. This step was repeated four times to ensure most of the fat was removed.
All samples were pasteurized according to Ontario government standards by heating to 72°C for not less than 16 seconds.
Preparation of Whey Whey was separated from the skim milk by first warming the sample to 31~C
using a water bath, adding a rennet-water solution, and allowing the milk to curd for 50 minutes. Once a curd was formed, it was gently agitated and gradually heated to 39°C. After holding this temperature for 60 minutes, the whey was removed.
Concentrating the Whey Proteins The whey proteins were concentrated via ultrafiltration (UF) at 4 °C.
Membranes with a pore size suitable to retain molecules with a molecular weight greater than 10,000 were used. The nitrogen pressure required for the procedure was maintained at 40 psi.
Aliquots of whey from each sample were injected into the UF. When 3m1 of whey remained in the UF, water was injected to further concentrate the whey proteins (diafiltration). The whey remaining in the OF (retentate) is highly concentrated with whey proteins. Samples were placed in the Dura-Dry MP freeze dryer for 42 hours.
Sphingomyelin Fatty Acid Analysis of Isolated Whey Protein Same procedure , as for whole milk (example 1) except started with 0.15g of freeze-dried whey protein instead of whole milk.
Results The results of this experiment are shown in Table 3, below.
Table 3. Effect of a Nervonic Acid Supplement (Canstar) on the Composition of Very Long Chain Fatty Acids in Sphingomyelin of Isolated Whey Protein from Milk*
Very Long Chain Fatty Olive #1 Olive #2 Canstar #1 Canstar *2 Acid (mol%) 22:0 41.9 69.2 30.8 41.9 23:0 30.4 16.6 24.0 12.8 24:0 25.7 14.0 18.4 11.2 24:1 2.0 0.1 26.7 34.0 *Values for the nervonic acid (24:1) content of the sphingomyelin in the whey protein fraction of cows milk, day 7 of the experiment. Nervonic acid content is increased over 10-fold by the Canstar supplement..
These results clearly demonstrate that isolated whey protein from cows fed the NA supplement (Canstar) is enriched in nervonyl-sphingomyelin relative to whey obtained from the milk of unsupplemented (Olive) cows.
Having illustrated and described the principles of the invention in a preferred embodiment, it should be appreciated to those skilled in the art that the invention can be modified in arrangement and detail without departure from such principles.
We claim all modifications coming within the scope of the following claims.
All publications, patents and patent applications are herein incorporated by reference in their entirety to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated by reference in its entirety.
Claims (21)
1. A feed additive for ruminants comprising the components:
(a) a source of nervonic acid; and (b) a source of inhibitors of microbial degradation of nervonic acid in the rumen said inhibitors being present in amounts to enhance the production of nervonic acid in the milk of ruminants.
(a) a source of nervonic acid; and (b) a source of inhibitors of microbial degradation of nervonic acid in the rumen said inhibitors being present in amounts to enhance the production of nervonic acid in the milk of ruminants.
2. A feed additive for ruminants comprising the components:
(a) a source of nervonic acid; and (b) a source of inhibitors of microbial degradation of nervonic acid in the rumen comprising feather meal or its constituents, said inhibitors being present in amounts to enhance the production of nervonic acid in the milk of ruminants.
(a) a source of nervonic acid; and (b) a source of inhibitors of microbial degradation of nervonic acid in the rumen comprising feather meal or its constituents, said inhibitors being present in amounts to enhance the production of nervonic acid in the milk of ruminants.
3. The feed additive of claims 1 or 2 wherein said inhibitors are present in amounts to enhance the production of nervonic acid in the milk of ruminants to levels greater than 16% of the very long chain fatty acids in the sphingomyelin of the milk.
4. A feed additive for ruminants comprising the components:
(a) a source of nervonic acid; and (b) a source of inhibitors of microbial degradation of nervonic acid in the rumen comprising feather meal or its constituents, said inhibitors being present in amounts to enhance the production of nervonic acid in the whey protein fraction of the milk of ruminants.
(a) a source of nervonic acid; and (b) a source of inhibitors of microbial degradation of nervonic acid in the rumen comprising feather meal or its constituents, said inhibitors being present in amounts to enhance the production of nervonic acid in the whey protein fraction of the milk of ruminants.
5. The feed additive of claim 4 wherein the inhibitors are present in amounts to enhance the production of nervonic acid in the whey protein fraction of milk of ruminants to levels of 26% to 34% of the very long chain fatty acids in the sphingomyelin in whey protein fraction of the milk.
6. The feed additive of claims 1, 2, 3, 4 or 5, wherein said source of nervonic acid is the Lunaria plant.
7. The feed additive of claim 6, wherein said source of inhibitors of microbial degradation of nervonic acid in the rumen of the ruminants is feathermeal.
8. A feed containing the feed additive as claimed in any preceding claim 1 to 7.
9. A method of producing milk in ruminants comprising feeding the ruminants a feed containing a feed additive according to any of claims 1 to 7 for a period of at least 2 days and milking the ruminants to obtain milk enriched for nervonic acid.
10. Expressed milk from ruminants which milk is produced using a method as claimed in claim 9.
11. The expressed milk as claim in claim 10, which contains greater than 16% nervonic acid as percent of the very long chain fatty acids in the milk.
12. A nervonic acid-enriched dairy product produced using expressed milk from ruminants enriched for nervonic acid as claimed in claim 11.
13. A nervonic acid-enriched dairy product as claimed in claim 12 which is cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant formula, or butter.
14. The expressed milk as claimed in claim 10, which contains 26% to 34% nervonic acid as a percent of the very long chain fatty acids in sphingomyelin in the whey protein fraction of the milk.
15. A nervonic acid-enriched low-fat dairy product produced using expressed milk from ruminants enriched for nervonic acid as claimed in claim 14.
16. A nervonic acid-enriched low-fat dairy product as claimed in claim 15 which is cheese, yogurt, cream, ice-creams, powdered milk, evaporated milk, infant formula, or butter.
17. A nervonic acid-enriched milk protein product produced using expressed milk from ruminants enriched for nervonic acid as claimed in claim 14.
18. A nervonic acid-enriched milk protein product as claimed in claim 17 which is a dried whey protein concentrate, dried whey, dried skim milk, dried milk protein, dried buttermilk, casein, acid casein, a protein drink, a protein bar or a protein supplement.
19. Use of feather meal as an inhibitor of microbial degradation of NA
in the rumen of ruminants.
in the rumen of ruminants.
20. Use of feather meal as claimed in claim 19 characterized in that the feather meal is added to the feed for ruminants.
21. Use of feather meal as claimed in claim 19 characterized in that the feather meal is added to feed additive in an amount sufficient to enrich milk from the ruminants with NA.
Priority Applications (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CA002445738A CA2445738A1 (en) | 2003-10-21 | 2003-10-21 | Method for enriching nervonic acid in expressed milk of ruminants |
PCT/CA2004/001856 WO2005036981A1 (en) | 2003-10-21 | 2004-10-21 | Method for enriching nervonic acid in expressed milk of ruminants |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CA002445738A CA2445738A1 (en) | 2003-10-21 | 2003-10-21 | Method for enriching nervonic acid in expressed milk of ruminants |
Publications (1)
Publication Number | Publication Date |
---|---|
CA2445738A1 true CA2445738A1 (en) | 2005-04-21 |
Family
ID=34427715
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CA002445738A Abandoned CA2445738A1 (en) | 2003-10-21 | 2003-10-21 | Method for enriching nervonic acid in expressed milk of ruminants |
Country Status (1)
Country | Link |
---|---|
CA (1) | CA2445738A1 (en) |
-
2003
- 2003-10-21 CA CA002445738A patent/CA2445738A1/en not_active Abandoned
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