Article pubs.acs.org/JAFC
Lipid-Encapsulated Echium Oil (Echium plantagineum) Increases the Content of Stearidonic Acid in Plasma Lipid Fractions and Milk Fat of Dairy Cows Melissa L. Bainbridge,† Adam L. Lock,§ and Jana Kraft*,† †
Department of Animal Science, University of Vermont, 570 Main Street, Burlington, Vermont 05405, United States Department of Animal Science, Michigan State University, 474 South Shaw Lane, East Lansing, Michigan 48824, United States
§
S Supporting Information *
ABSTRACT: The objective of this study was to evaluate the impact of feeding lipid-encapsulated echium oil (EEO) on animal performance and milk fatty acid profile. Twelve Holstein dairy cows were used in a 3 × 3 Latin Square design with 14 day periods. Treatments were a control diet (no supplemental fat), 1.5% dry matter (DM) as EEO and 3.0% DM as EEO. Treatments had no negative effect on animal performance (dry matter intake, milk yield, and fat yield). The milk fat content of total n-3 fatty acids and stearidonic acid (SDA) increased with EEO supplementation (P < 0.001). The proportion of SDA increased in all plasma lipid fractions with EEO supplementation (P < 0.001). Transfer of SDA from EEO into milk fat was 3.4 and 3.2% for the 1.5 and 3% EEO treatments, respectively. In conclusion, EEO increases the content of n-3 fatty acids in milk fat; however, the apparent transfer efficiency was low. KEYWORDS: blood metabolites, γ-linolenic acid, bioactive fatty acids, plasma lipid fractions, transfer efficiency
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mammary gland for production of milk fat,13 resulting in a low transfer efficiency of EPA and DHA into milk.14 Recent research demonstrates the two shorter carbon chain n-3 FA, αlinolenic acid (ALA; 18:3 9c,12c,15c) and stearidonic acid (SDA; 18:4 6c,9c,12c,15c), show similar health benefits to EPA and DHA.15,16 SDA bypasses the initial rate-limiting step of δ6-desaturase in the biosynthesis of EPA and DHA and, thus, is converted 4−5-fold more efficiently to EPA than ALA.17 Moreover, recent research suggests that uptake of FA by the mammary gland is based on the fatty acid composition of plasma triacylglyceride (TAG), as an intravenous infusion of a TAG emulsion of ALA exhibited a higher transfer efficiency (28%) to milk fat when compared to DHA (11%).18 Echium oil (EO) from the seed of Echium plantagineum, a member of the Boraginaceae family, is a plant-based source of SDA (13% of total FA), ALA (31% of total FA), and γ-linolenic acid (GLA; 18:3 6c,9c,12c; 9% of total FA). GLA has been shown to possess anticarcinogenic and anti-inflammatory properties,19,20 and several studies have shown the health benefits of echium oil, as well as its promise as an alternative to fish oils.21,22 Kitessa and Young23 demonstrated an increase in the n-3 FA content of milk with inclusion of a formaldehydetreated EO on a pasture-based diet using five dairy cows. Lipid protection may provide more resilience against rumen biohydrogenation and thus may result in higher incorporation into milk fat.11 There can also be considerable variation in response to supplementation depending upon basal diet and inclusion level;24 thus, an evaluation of EO fed at incremental
INTRODUCTION The increased understanding of the relationship between nutrition and health has led to a growing interest in promoting functional foods enriched with bioactive compounds. One class of bioactive compounds is n-3 fatty acids (n-3 FA), which are known for their protective effects against inflammation,1 neurological disorders,2 and cardiovascular diseases,3,4 as well as their importance to growth and development.5 Current dietary sources do not meet the requirements for n-3 FA in the human diet,6 driving the pursuit to develop functional foods enriched with n-3 FA. The fatty acid profile of milk is predominantly influenced by the diet of dairy cows,7 and research has demonstrated that FA derived from supplemental oils can be incorporated into milk fat.8,9 There are several challenges, however, when supplementing dairy cows with oils rich in polyunsaturated FA (PUFA). One is the extensive biohydrogenation of dietary unsaturated FA by rumen bacteria, which often exceeds 85%.10 This produces trans-fatty acid isomers that decrease lipogenesis in the mammary gland, leading to milk fat depression, and reduce the amount of unsaturated FA available to the mammary gland for milk fat synthesis.10 Various protection methods have been developed to try to minimize ruminal biohydrogenation of feed-derived unsaturated FA, such as chemical treatment, calcium salt, protein aldehyde matrix, and encapsulation with saturated lipid. Hence, protecting n-3 FA from bacterial degradation in the rumen is essential to increase the amount of n-3 FA in milk while maintaining animal performance.11 Docosahexaenoic acid (DHA; 22:6 4c,7c,10c,13c,16c,19c) and eicosapentaenoic acid (EPA; 20:5 5c,8c,11c,14c,17c) are longchain, highly unsaturated n-3 FA that are preferentially incorporated into plasma cholesterol esters and phospholipids.12 Both plasma lipid fractions are unavailable to the © 2015 American Chemical Society
Received: Revised: Accepted: Published: 4827
February April 22, April 22, April 22,
15, 2015 2015 2015 2015 DOI: 10.1021/acs.jafc.5b00857 J. Agric. Food Chem. 2015, 63, 4827−4835
Article
Journal of Agricultural and Food Chemistry
diet. EO was purchased from Technology Crops International (Winston-Salem, NC, USA) and lipid-encapsulated with hydrogenated vegetable oil by Jefo (Saint-Hyacinthe, Canada) using the spray cooling method with a prilling automizer. Lipid encapsulation technology is based on the inherent property of saturated lipids being resistant to enzymes in the rumen, yet susceptible to enzymatic digestion in the small intestine.11 EO contained 35% ALA, 15% 18:1 9c, 15% SDA, 10% 18:2 9c,12c, 10% GLA, and 3% 18:0. The lipidencapsulated supplement contained 25% EO, and the fatty acid profile of the supplement is presented in Table 2. A 10 day period before the
levels on conserved forages is needed to determine the optimal inclusion rate for commercial feasibility. It is necessary to determine which plasma lipid fractions EO-derived FA are preferentially incorporated into and hence to what extent they can be incorporated into milk fat. We hypothesized that the inclusion of lipid-encapsulated echium oil (EEO) in the diet of dairy cows fed a total mixed ration (TMR) would increase the ALA, SDA, EPA, DHA, and GLA contents of milk fat. Our objectives were to (1) evaluate the effects of feeding an EEO supplement on milk production parameters, (2) determine the transfer efficiency of EO-derived FA to milk fat by supplementing incremental levels of EEO, and (3) assess the incorporation of EO-derived FA into plasma lipid fractions.
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Table 2. Fatty Acid Composition (Grams per Kilogram) of Lipid-Encapsulated Echium Oil (EEO)
MATERIALS AND METHODS
Experimental Design. All experimental procedures were approved by the Michigan State Institutional Animal Care and Use Committee. Twelve multiparous Holstein dairy cows in late lactation (229 ± 62 DIM) were used in a 3 × 3 Latin Square design with 14 day experimental periods. Cows averaged body weights of 749 ± 69 kg and body condition scores of 3.56 ± 0.39 (1−5 in 0.25 increments25) at the start of the study. Diets were formulated to meet NRC (2001) nutrient requirements (Table 1) and fed as TMRs. Treatments were diets containing no supplemental fat (control; CON), diets supplemented with EEO at 1.5% of DM (low EEO; LEO), and EEO at 3% of DM (high EEO; HEO). EEO replaced soy hulls in the
Table 1. Ingredient and Nutrient Composition of the Treatment Diets
fatty acid
EEO
12:0 14:0 16:0 18:0 cis-9 18:1 cis-11 18:1 cis-9,cis-12 18:2 20:0 cis-6,cis-9,cis-12 18:3 cis-9,cis-12,cis-15 18:3 cis-6,cis-9,cis-12,cis-15 18:4 22:0 cis-11,cis-14,cis-17 20:3 24:0 cis-15 24:1
0.12 0.81 41.5 33.4 4.33 0.14 3.96 0.39 2.65 9.03 3.40 0.06 0.12 0.06 0.04
treatment a
ingredient, % of DM corn silage alfalfa silage dry ground corn soybean meal soyhulls vitamin−mineral mixd echium oil supplement nutrient composition DM, % NDF,e % of DM CP,f % of DM starch, % of DM total fatty acids, % of DM cis-9,cis-12 18:2 cis-6,cis-9,cis-12 18:3 cis-9,cis-12,cis-15 18:3 cis-6,cis-9,cis-12,cis-15 18:4 total SFA total MUFA total PUFA total n-3 total n-6 n-6/n-3 ratio
CON
LEOb
HEOc
33.9 17.0 23.8 12.7 9.2 3.5 0.0
33.9 17.0 23.8 12.7 7.7 3.5 1.5
33.9 17.0 23.8 12.7 6.2 3.5 3.0
48.8 27.2 17.0 28.5 2.72 1.33 0.00 0.20 0.0 0.58 0.59 1.54 0.20 1.34 6.61
47.7 26.2 15.9 28.4 3.71 1.25 0.03 0.31 0.04 1.49 0.59 1.63 0.35 1.28 3.63
47.4 25.3 16.3 28.5 4.94 1.31 0.06 0.43 0.08 2.39 0.65 1.89 0.51 1.37 2.69
start of the trial served as a preliminary period. Milk samples were collected during the last 3 days of this period, and cows were subsequently blocked by fat-corrected milk (FCM) yield and assigned to treatments. Cows were housed in individual tie stalls, blocked from feed at 8:00 a.m., fed once daily at 10:00 a.m., and milked twice daily at 4:00 a.m. and 3:00 p.m. Corn silage and alfalfa silage DM were determined twice weekly, and TMR was adjusted accordingly. Diets were fed ad libitum (12−17% orts) throughout the experiment. Fresh drinking water was available continuously. Data and Sample Collection. Milk weights and samples were taken at each milking during the last 4 days of each period; an aliquot was preserved with 2-bromo-2-nitropropane-1,3-diol and analyzed for fat, protein, lactose, and milk urea nitrogen (MUN) by mid-infrared spectroscopy by the Michigan Herd Improvement Association (Universal Lab Services, Lansing, MI, USA). Another aliquot was stored at −20 °C without preservative for fatty acid analysis. Feed offered and refusals were weighed and recorded daily. Feed ingredients and TMR were sampled during the last 4 days of each period and stored at −20 °C until further analysis. Blood was collected from coccygeal vessels into evacuated tubes containing K2EDTA (Becton Dickenson, Franklin Lakes, NJ, USA) at 9:00 a.m. on the last day of each period. Blood samples were placed immediately on ice, and plasma was obtained within 2 h of blood collection by centrifugation at 900g for 15 min at 4 °C and stored at −20 °C. Feed and Orts Samples. Samples for each period were composited and dried in a forced-air oven at 65 °C for 48 h. Dried samples were weighed to determine dry matter and ground with a Wiley mill using a 1 mm screen (Arthur H. Thomas, Philadelphia, PA, USA). Individual feed ingredients and orts were analyzed for neutral detergent fiber (NDF) with heat-stable α-amylase and sodium sulfite,26 crude protein (CP),27 and starch composition28 (Cumberland Valley Analytical Services Inc., Hagerstown, MD, USA). Reported NDF values are inclusive of ash. The fatty acid composition of feed samples was determined according to a modified method of Sukhija and Palmquist.29 Glyceryl tridecanoate (Nu-Check Prep, Elysian, MN, USA) was used as an internal standard (1 mg/mL in acetone). Two milliliters of toluene and 2 mL of 5% methanolic sulfuric acid (Sigma-
a
CON, control (0% of DM as lipid-encapsulated echium oil). bLEO, 1.5% of DM as lipid-encapsulated echium oil. cHEO, 3% of DM as lipid-encapsulated echium oil. dVitamin−mineral mix contained (DM basis) 30.1% limestone, 25.3% sodium bicarbonate, 10.1% salt, 7.07% urea, 6.00% potassium chloride, 5.98% dicalcium phosphate, 5.68% magnesium sulfate, 5.68% animal fat, 3.94% trace mineral premix and vitamins, and 0.21% selenium yeast 600 (600 mg of Se/kg). eNDF, neutral detergent fiber. fCP, crude protein. 4828
DOI: 10.1021/acs.jafc.5b00857 J. Agric. Food Chem. 2015, 63, 4827−4835
Article
Journal of Agricultural and Food Chemistry Table 3. Fatty Acid Intake (Grams per Day) of Treatments treatment CONa
LEOb
HEOc
SE
P value
total fatty acids 16:0 18:0 cis-9 18:1 cis-9,cis-12 18:2 cis-9,cis-12,cis-15 18:3 cis-6,cis-9,cis-12 18:3 cis-6,cis-9,cis-12,cis-15 18:4 ∑ otherd
726a 114a 21.5a 145a 358a 50.4a 0.30a 0.04a 35.7a
1000b 250b 125b 146a 338b 84.5b 8.50b 10.6b 34.8a
1329c 382c 228c 162b 356a 116c 17.1c 21.7c 44.5b
27.0 7.54 4.48 3.94 8.24 2.34 0.32 0.40 1.02
