Feeding high-oleic peanuts to layer hens enhances egg yolk color and oleic fatty acid content in shell eggs

Feeding high-oleic peanuts to layer hens enhances egg yolk color and oleic fatty acid content in shell eggs Ondulla T. Toomer,∗,1 Amanda M. Hulse-Kemp,† Lisa L. Dean,∗ Deborah L. Boykin,‡ Ramon Malheiros,§ and Kenneth E. Anderson§ U.S. Department of Agriculture, Agricultural Research Service, Market Quality & Handling Research Unit, Raleigh, NC 27695, USA; † U.S. Department of Agriculture, Agricultural Research Service, Genomics and Bioinformatics Research Unit, Raleigh, NC 27695, USA; ‡ U.S. Department of Agriculture, Agricultural Research Service, Office of the Area Director, Stoneville, MS 38776, USA; and § Department of Poultry Science, North Carolina State University, Raleigh, NC 27695, USA from layer hens fed the HO peanut + corn diet had reduced egg weights relative to the controls (P = 0.0001). Eggs produced from layer hens fed the HO peanut diet had greater yolk color scores (P < 0.0001), HO fatty acid (P < 0.0001), and β -carotene (P < 0.0001) levels in comparison to the controls. Eggs produced from hens fed the control diet had greater palmitic and stearic saturated fatty acids (P < 0.0001), and trans fat (P < 0.0001) content compared to eggs produced from hens fed the HO peanut diet. All egg protein extracts from all treatments at each time point were non-reactive with rabbit anti-peanut agglutinin antibodies. This study identifies HO peanuts as an abundant commodity that could be used to support local agricultural markets of peanuts and poultry within the southeastern United States and be of economic advantage to producers while providing a potential health benefit to the consumer with improved egg nutrition.

ABSTRACT Previous studies have identified normaloleic peanuts as a suitable and economical broiler feed ingredient. However, no studies to date have examined the use of high-oleic (HO) peanut cultivars as a feed ingredient for laying hens and determined the impact of feeding HO peanuts on performance and egg nutritive qualities. This project aimed to examine the use of HO peanuts as a feed ingredient for layer hens to determine the effect on performance, egg lipid chemistry, and quality of the eggs produced. Forty-eight 40-wkold layer hens were fed a conventional soybean meal + corn control diet or a HO peanut + corn diet for 10 wk in conventional battery cages. Body and feed weights were collected weekly. Pooled egg samples were analyzed for quality, lipid analysis, and peanut protein allergenicity. There were no significant differences in hen performance or egg quality as measured by USDA grade quality, egg albumen height, or egg Haugh unit between the treatment groups. However, eggs produced

Key words: high-oleic peanuts, feed ingredients, layer hens, shell eggs, β -carotene 2018 Poultry Science 0:1–18 http://dx.doi.org/10.3382/ps/pey531

INTRODUCTION

tities of these grains and high protein oilseeds are imported from South America (Economic Research Service, USDA, 2017) and routinely from Iowa and Illinois, the top producing soybean and corn states (Agweek, 2009) in the U.S. Mid-West for animal feed production. This study aims to examine the use of high-oleic peanuts, a valuable oilseed crop abundantly grown also within the U.S. southeast, with Georgia producing approximately 46% of U.S. peanuts annually (National Peanut Board, 2018). In various parts of the world, peanut meal from normal-oleic peanuts (groundnuts) is utilized as a common protein source for feeding livestock (Cilly et al., 1977; Olomu and Offiong, 1980, 1985; Aletor and Olonimoyo, 1992; Venkataraman et al., 1994; Donkoh et al., 1999; Naulia and Singh, 2002). Yet, in the United States 80% of peanut production is used for peanut butter, oil production, and snacks for human consumption

For decades, the poultry industries (broilers, eggs, turkeys) have been of great economic importance and represented a significant portion of the agricultural products sold in the southeastern (Georgia, Arkansas, Alabama, North Carolina, South Carolina, and Mississippi) United States (APHIS, USDA, 2015). Within this region, the need for poultry feed components such as corn and soybean meal far exceeds the ability to produce these ingredients locally. Hence, large quan-

Published by Oxford University Press on behalf of Poultry Science Association 2018. This work is written by (a) US Government employee(s) and is in the public domain in the US. Received April 23, 2018. Accepted October 30, 2018. 1 Corresponding author: [email protected]

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∗

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MATERIALS AND METHODS Experimental Design, Animal Husbandry and Dietary Treatments To test this hypothesis, we compared 2 isonitrogenous and isocaloric dietary treatments prepared 1 wk prior to the onset of the study and maintained in feed storage bins in North Carolina State University (NCSU) Feed Mill in a cool dry location. Treatment 1 was a high-oleic peanut + corn diet (18% crude protein, 11% fat, and 1,400 metabolizable energy) prepared using aflatoxinfree whole non-roasted high-oleic peanuts with the testa (skin) intact crushed using a Roller Mill. Prior to inclusion within the diet, whole raw high-oleic peanuts were crushed using a Roller Mill. The high-oleic peanut + corn diet (HO PN) was composed predominately of yellow corn, corn gluten, whole high-oleic peanuts (nonroasted, testa intact), and wheat bran, supplemented with amino acids L-lysine, L-methionine, L-tryptophan, L-threonine along with NCSU vitamin and mineral premix (Figure 1). Treatment 2 was a soybean meal + corn diet (SBM), a control conventional mash diet (18% crude protein, 7% fat, and 1,400 metabolizable energy), composed predominately of yellow corn, corn gluten, soybean meal, wheat bran, and poultry fat, with NCSU vitamin and mineral premix (Figure 1). Experimental diets were analyzed and determined to be free of aflatoxin and microbiological contaminants by the North Carolina Department of Agriculture and Consumer Services, Food and Drug Protection Division Laboratory (Raleigh, NC). Forty-eight 40-wk-old White Leghorn layer hens (NCSU University maintained Poultry Flock) were randomly assigned to 24 animals per treatment. Animals were housed in 1 room with 2 sets of conventional battery cages housing 24 hens each on each side of the room designated as left and right (1 hen per cage; 12 cages upper row + 12 cages lower row) with 12 animals randomly assigned to 2 replicate pens within each treatment group. To account for treatment variability due to environmental factors such as ventilation and/or temperature in the room, 12 animals per treatment were housed on separate sides of the room to ascertain the effect of environmental differences between each side of the room. Animals were provided feed and water ad libitum and housed individually with 14 h of light daily for 10 wk. Body and feed weights were recorded weekly. Daily shell eggs were collected, and weights were recorded from each hen. Weekly, shell eggs were analyzed for quality (yolk color DSM, albumen height, Haugh unit [HU]) and USDA grading by the Layer Hen & Small Flock Management Lab, Department Poultry Science, NCSU. The Institutional Animal Care and Use Committee at NCSU approved all experimental animal protocols prior to the onset of this study.

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(American Peanut Council, 2017). Although earlier research studies demonstrated that peanut meal prepared from normal-oleic peanuts is a suitable and economical poultry feed ingredient (Costa et al., 2001; Pesti et al., 2003), few studies have examined the use of modern high-oleic peanut cultivars (80% oleic fatty acids and 2% linoleic fatty acids) as a feed ingredient for laying hens. In addition, early poultry feeding studies using normal-oleic peanuts determined lysine as the first limiting dietary amino acid, followed by methionine and tryptophan (Douglas and Harms, 1959; Waldroup and Harms, 1963). However, relatively recent introduction of affordable purified threonine and other amino acids has made the inclusion of purified amino acids in poultry rations economically reasonable. Studies by Moreira et al. (2014, 2016) demonstrated the human health benefits of moderate consumption of high-oleic peanuts; however, few studies have examined how feeding peanuts high in monounsaturated fatty acids to egg-producing hens and poultry may alter the fatty acid and lipid profile of poultry meat and eggs produced. Studies by Shapira et al. (2008) demonstrated that an extruded linseed-supplemented diet fed to layer hens increased the omega-3 polyunsaturated fatty acid (PUFA) content in table eggs compared to control eggs, with fortified eggs yielding approximately a 3.8-fold increase in omega 3 PUFA, 6.4-fold increase in alpha-linolenic acid (18:3), and 2.4-fold increase in docohexaenoic acid (DHA). Moreover, laying hens fed diets with varying levels of fish meal (0% control diet to 20%) had increased DHA in egg yolk relative to increased dietary inclusion of fish meal compared to the controls with the exception of 15 and 10% fish meal, which led to the same increase of DHA levels in egg yolk (Howe et al., 2002). However, the inclusion of greater than 1 to 2% of marine oils (primary source PUFA) in poultry diets compromises the oxidative stability of the meat and/or eggs (Lopez-Ferrer et al., 2001; Baeza et al., 2013), while producing off-flavors (Hargis et al., 1991). Other reports have demonstrated that linseed inclusion in the diets of hens also produces off-flavors in the meat and eggs produced similar to those of hens fed diets with marine oils (Woods and Fearon, 2009). Therefore, in this study we aimed to 1) examine the usefulness of whole high-oleic peanuts as a cost-effective and suitable protein and energy-rich feed ingredient for 40-wk-old layer hens, and 2) determine the effects of feeding a high-oleic peanut diet to egg-producing layer hens on the nutritive value, lipid, and fatty acid composition of the shell eggs produced. We conjecture that the eggs produced from layer hens fed a diet with higholeic peanuts will have improved lipid and fatty acid profiles with enhanced healthy monounsaturated fatty acids compared to poultry fed a conventional layer hen diet.

EGGS FROM LAYER HENS FED HIGH-OLEIC PEANUTS

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Layer Hen Feed Analysis Total nitrogen was determined in homogenized samples by combustion using an Elementar N cube analyzer (Elementar Americas, Mt. Laurel, PA) according to AOAC 990.03 (2007). A Kjeldahl conversion factor of 6.25 (mixed food) was used to calculate total protein in the samples. Total fat as triglycerides was determined in the samples gravimetrically after Soxhlet extraction. Samples were extracted for 6 h using continuous extraction with hexane (AOAC 920.39, 1990).

Egg Quality and Grading Albumen height and HU (Haugh, 1937) were recorded with the TSS QCD system (Technical Services and Supplies, Dunnington, York, UK). Egg albumen height and HU were calculated to determine egg albumen quality. Yolk color was determined by using TSS QCD System yolk color scan which is calibrated to the DSM Color Fan, which consists of a series of 15 colored plastic tabs arranged as a fan corresponding to the range of yolk colors found in yellow from light yellow to

orange-red (a color index 1 to 15 to distinguish the yolk color density from lightest to darkest color intensity), earlier defined by Vuilleumier (1969). Eggs were visually inspected for exterior and interior grading in accordance with the USDA Standards (USDA 2010). The exterior of eggs was examined to ensure that a clean, smooth, defect-free, oval shape surface was present, lacking thin spots or textured appearance. Interior grading was conducted by the candling method to examine the air cell, the albumen, and yolk and to examine for meat and blood spots and cracks. Eggs were sized weekly by treatment (USDA, 2010). Egg sizing was classified by a minimum net weight per egg: peewee (<42.6 g), small (42.6 < 56.8 g), medium (49.7 < 56.8 g), large (56.8 < 63.9 g), extra-large (63.9 < 70.9 g), and jumbo (>70.9 g).

Egg Samples—Protein Extraction Shell eggs were collected, labeled, and weighed daily from each laying hen. All eggs produced by an individual layer hen were pooled in a sterile Whirlpak bag (Thermo Fisher Scientific, Rockford, IL) and homogenously mixed using a Stomacher Lab Blender.

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Figure 1. Layer hen dietary treatments and composition. (A) Treatment 1 was a high-oleic peanut + corn diet (HOPN: 18% crude protein, 11% fat, and 1,400 metabolizable energy) composed predominately of yellow corn, corn gluten, whole high-oleic peanuts with the skins intact (nonroasted) and wheat bran, supplemented with amino acids L-lysine, L-methionine, L-tryptophan, L-threonine along with vitamin and mineral premix. Prior to inclusion within the diet, whole raw high-oleic peanuts were crushed using a Roller Mill. (B) Treatment 2 was a control, conventional soybean meal + corn layer hen mash diet (SBM:18% crude protein, 7% fat, and 1,400 metabolizable energy (kCal/lb.)) composed predominately of yellow corn, corn gluten, soybean meal, wheat bran and poultry fat, with vitamin and mineral premix. (C) Composition table of experimental diets in percent by weight.

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SDS-PAGE Analysis and Immunoblotting Protein extracts from pooled shell egg samples (75 μg per lane) were loaded on a 10% polyacrylamide Trisglycine gel (Criterion TGX, Bio-Rad, Hercules, CA) and electrophoretically separated under constant voltage with Tris/Glycine/SDS buffer according to the manufacturer’s instructions (Bio-Rad). Proteins were visualized by Coomassie Brilliant Blue staining and digitally imaged. SDS-PAGE resolved proteins that were transferred to nitrocellulose membranes electrophoretically for immunoblotting procedures using Midi TransBlot Turbo Transfer Pack (Bio-Rad) The membranes were washed 3 times in Tris-buffered saline with 0.1% Tween (TBST) and subsequently blocked in 5% nonfat dry milk (NFDM) in TBST for 1 h at room temperature with shaking. To examine the potential peanut allergenicity of protein extracts from shell egg samples, membranes were incubated at 4◦ C overnight with shaking in 5% NFDM in TBST containing rabbit IgG antipeanut agglutinin primary antibody 1:1000 dilution (LifeSpan Biosciences, Seattle, WA). Membranes were washed 3 times in TBST and incubated in 5% NFDM in TBST containing secondary antibody donkey antirabbit IgG-HRP (Santa Cruz Biotechnology, Dallas, TX) dilution 1:1,000 at room temperature with shaking for 1 h. Bio-detection was determined utilizing chromogenic peroxidase substrate CN/DAB (chloronaphthol and diaminobenzidine, Thermo Fisher Scientific) detection of HRP (horseradish peroxidase) activity.

Lipid and Fatty Acid Analysis Lipid and fatty acid analysis (total fat, total oleic fatty acid, and total linoleic fatty acid) of homogenous pooled total egg samples (experimental weeks 2, 3, 4, 6, 7, 8, and 9) and feed samples from both treatment groups were analyzed using direct methylation modified methods as described by Wang et al. (2000) in the Market Quality & Handling Research Unit. Pooled total egg samples were thawed at room temperature and vortexed to obtain a homogenous mixture. One milliliter of internal standard (tridecanoic acid, 1 mg/mL, Fluka Chemical Corp., Milwaukee, WI) in hexane was added to a screw-capped glass tube. The

hexane (Thermo Fisher Scientific, Waltham, NJ) was evaporated in a stream of nitrogen. One hundred milligrams of pooled total egg sample was added to the tube, and the weight was recorded. One milliliter of methanol (Thermo Fisher) was added, and the contents of the tube were vortexed. Three milliliters of methanolic hydrochloric acid (Sigma Chemical Corp., St. Louis, MO) was subsequently added. The tube was capped, vortexed, and placed in a water bath for 1 h at 95◦ C. After removing from the water bath and cooling to room temperature, 8 mL of 0.88% sodium chloride (Sigma) in water was added and the tube was vortexed. Subsequently, 3 mL of hexane was added. The tube was vortexed for 30 s and centrifuged for 15 min at 1,000 × g using the IEC Model K centrifuge. The top organic layer was transferred to a clean glass tube containing approximately 50 mg of sodium sulfate (Sigma) to dry the solvent. The layer was then transferred to a crimp top autosampler vial. The samples were run on the PEAS XL Autosampler GC (Perkin Elmer, Shelton, CT) with Kel Fir Fame 5 and Kel Fir Fame 6 Standards (Matreya LLC, State College, PA). The column used was an SGE BPX-70 (70% Cyanopropyl Polysilphenylenesiloxane, SGE Analytical Science, Austin, TX, 30 m length, 0.25 mm i.d., 0.25μ df). The temperature program was 60◦ C for 2-min hold, 10◦ C per minute to 180◦ , 0-min hold, and then 4◦ C per minute to 235◦ C with 0-min hold for a total of 27.75 min. The injector was split at 40 mL/min at 220◦ C. The detector was FID at 250◦ C. The carrier gas was helium at 1.85 mL/min. Fatty acids were identified by retention time matches using the Kel Fir Fame 6 Standard. The results were calculated as total fat (g/100 g) as triglycerides using response factors calculated from the Kel Fir Fame 5 Standard (Ngeh-Ngwainbi et al., 1997). Homogeneous egg samples from each treatment group were chemically analyzed for crude fat, cholesterol, β -carotene, and 36 differing fatty acids (butyric, caproic, caprylic, undecanoic, lauric, tridecanoic, myristic, myristoleic, pentadecylic, pentadecenoic, palmitic, palmitoleic, margaric, margaroleic, stearic, oleic, n9 t elaidic, linoleic, linolenic, n6 gamma-linolenic, arachidic, gadoleic, eicosadienoic, homo-gamma-linolenic, eicosatrienoic, arachiconic, n3 timnodonic, heneicosanic, behenic, erucic, brassic, docosahexaenoic, lignoceric, nervonic, omega 3, and omega 6) content at weeks 1, 5, and 10 experimentally (Tables 5, 6, 8, and 9). Analysis of 36 differing fatty acids on all 775 egg samples was cost-prohibitive at all experimental time points (week 1 to 10). Therefore, only eggs produced at weeks 1, 5, and 10 were analyzed for the full panel of 36 fatty acids by an external vendor (ATC Scientific). For β -carotene and comprehensive lipid and fatty acid analysis, ATC Scientific (Little Rock, AR) analyzed pooled total egg samples for experimental weeks 1, 5, and 10 of experimental samples only due to cost restrictions. Pooled total egg samples were analyzed for the following lipids and fatty acids: crude fat,

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Total proteins were extracted from pooled shell egg samples (experimental weeks 1, 3, 5, 7, and 10) in 20 mM Tris-HCL pH 8.1 extraction buffer supplemented with 1% HALT protease inhibitor (Thermo Fisher Scientific) using 1 mL of homogenous pooled shell egg sample to 10 mL of extraction buffer. Samples were vortexed and centrifuged for 10 min at 4◦ C, and the supernatants (protein extracts) were used for analysis. Protein concentration of protein extracts was determined using a BCA Protein Assay Kit (Thermo Fisher Scientific). Protein extracts were aliquoted and stored at –20◦ C.

EGGS FROM LAYER HENS FED HIGH-OLEIC PEANUTS

Statistical Analysis-Fatty Dietary Treatments Biochemical components of the 2 diets were measured in triplicate and then compared using a t-test in SAS software version 9.4 (PROC TTEST). The diets were investigated for equal variance: if variances were detected to be significantly different, the Satherwaithe t-test P-value was reported and if variances were not statistically significant, the pooled t-test P-value was reported for all biochemical components.

Statistical Analysis-Hen Performance Performance of the laying hens was measured in number of total eggs produced, feed intake, feed conversion ratio, and body weight for each animal over 10 wk. Analysis of variance was performed on each phenotype using a general linear mixed model mixed model (PROC GLIMMIX in SAS software version 9.4). In the first analysis room, treatment, week, and their interactions were included as fixed effects. Animals were treated as replications for diet treatments, measurements taken on individual eggs were treated as subsamples for each animal, and weekly measurements were repeated measures for each animal. Random effects included animals within treatment and room and week by animal within treatment and room with residual error representing subsampling error or variability between eggs for each animal. In the second analysis, week was treated as a trend with other factors treated same as initial analysis. For all models, residuals were plotted to verify that residuals were independent and normally distributed. Means were separated at P = 0.05 using least squares means with Tukey-Kramer adjustment for multiple comparisons.

Statistical Analysis-Egg Quality One egg per animal per week for 10 wk was used for egg quality measurements including number of meat spots and number of blood spots, egg weight (grams), albumen height, HU, and yolk color measured by Roche. Confidence intervals were evaluated between

treatment groups for number of meat spots and number of blood spots.

Statistical Analysis-Egg Chemistry, Lipid and Fatty Acid Analysis β -Carotene levels and full lipid and fatty acid profiles were run on a single egg from each animal measured on weeks 1, 5, and 10. A separate egg was utilized for the β carotene measurement and the full lipid and fatty acid profiles. These measurements were similarly treated as hen performance above with the initial analysis utilizing the week as classification effect. Since there were just 3 weekly measurements, a trend was not used to model the week effect. Total fats, oleic acid, linoleic acid, and the oleic-to-linoleic acid (O/L) ratio were measured on weeks 2, 3, 4, 6, 7, 8, and 9 for eggs from all animals. These data were initially treated with week as a class, and then analyzed for a trend if a significant interaction was observed.

RESULTS AND DISCUSSION Feed Analysis Although both experimental dietary treatments were formulated to be isocaloric and isonitrogenous, the high-oleic peanut + corn diet had significantly greater amounts of lipid compared to the conventional soybean meal diet, with 9.8 and 6.9% lipid, respectively (P = 0.006, Table 1). Therefore, the corn within the soybean meal + corn diet provided the predominant source of energy, whereas the lipids from the higholeic peanuts in combination with the corn within the high-oleic + corn diet provided dietary energy for egg-producing layer hens. Nevertheless, the percentages of nitrogen and protein were similar between both experimental dietary treatment groups (Table 1). The soybean meal + corn experimental diet had a significantly greater (P-value < 0.0001) content of saturated fatty acids (26.4%) relative to the saturated fatty acid content in the high-oleic peanut + corn diet (14.3%), with the following saturated fatty acids content being significantly greater (P < 0.0001) in the conventional soybean meal + corn diet (Table 1): myristic acid (14:0), palmitic acid (16:0), and stearic acid (18:0). At small quantities, arachiadic acid (20:0), behenic (22:0), lignoceric acid (24:0), and cerotic acid (26:0) were the predominant saturated fatty acids found within the high-oleic peanut + corn experimental diet (Table 1). The high-oleic peanut + corn diet had significantly greater (P < 0.0001) amounts of monounsaturated fatty acids in comparison to the soybean meal + corn diet, with a oleic acid (18:1) content of 72.18 ± 0.18 and 38.04 ± 0.19, respectively (Table 1). Although gondoic acid (20:1) and erucic acid (22:1)

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butyric acid, caproic acid, caprylic acid, capric acid, lauric acid, tridecanoic acid, myristic acid, myristoleic acid, pentadecylic acid, palmitic acid, palmitoleic acid, margaric acid, stearic acid, oleic acid, n9 t elaidic acid, linoleic acid, linolenic acid, arachidic acid, behenic acid, brassic acid, lignoceric acid, and nervonic acid. Methods used to determine β -carotene content in eggs were determined using AOAC 958.05 (1990) color of egg yolk. Egg cholesterol was determined using methods described by Zhang et al. (1999). Egg fat hydrolysis methods were determined using AOAC method 954.02 (1990), and fatty acid profile methods were determined using AOCS Ce 2–66/Ce 1e 91 methods.

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TOOMER ET AL. Table 1. Chemical lipid and fatty acid analysis of dietary treatments.1

% Lipid % Nitrogen % Protein

High-oleic peanut + corn diet (HO-PN)

P-value2

6.88 ± 0.02 2.78 ± 0.27 17.4 ± 1.7

9.79 ± 0.39 2.76 ± 0.04 17.2 ± 0.3

0.006 0.90 0.90

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

< 0.0001 0.003 < 0.0001 < 0.0001 0.42 < 0.0001 < 0.0001 < 0.0001 0.0002 < 0.0001 < 0.0001 < 0.0001 0.18 < 0.0001 0.14 < 0.0001 < 0.0001 < 0.0001

0.446 0.126 21.4 4.43 0.072 4.83 38.0 28.3 1.24 0.165 0.314 0 0 0 0 1.34 26.4 1.07

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

0.010 0.010 0.1 0.17 0.120 0.05 0.2 0.2 0.02 0.040 0.030 0.00 0.00 0.00 0.00 0.01 0.1 0.01

0 0 7.32 0.085 0 2.54 72.2 11.3 0.34 1.07 1.38 2.05 0.85 1.25 0.40 6.40 14.3 0.79

0.00 0.00 0.08 0.00 0.00 0.02 0.2 0.2 0.00 0.01 0.04 0.03 0.07 0.02 0.29 0.10 0.07 0.01

1 Three replicate samples were collected from each dietary treatment and were analyzed for lipid, nitrogen, protein, lipid, and fatty acid content. Each value represents the mean ± the standard error for each sample. Fatty acid content (g/100 g sample). 2 Measurements were tested for equal variance. When variances were detected to be statistically significant Satherwaithe P-values were reported, else pooled P-values were reported.

monounsaturated fatty acids were found in greater content in the high-oleic peanut + corn diet in comparison to the soybean meal + corn diet, they were in small quantities (Table 1). Monounsaturated fatty acids myristoleic acid (14:0) and palmitoleic acid (16:1) were found to be in greater quantities in the soybean meal + corn diet (0.126 ± 0.01, 4.43 ± 0.17) relative to the high-oleic peanut + corn diet (0.0, 0.085 ± 0.07), respectively (Table 1). The soybean meal + corn diet had significantly greater amounts of (P < 0.0001, Table 1) the PUFA, linoleic acid (28.29 ± 0.160) in comparison to the higholeic peanut + corn diet (11.28 ±0.200). Additionally the soybean meal + corn diet had significantly greater contents of omega 3 fatty acid (linolenic acid) in comparison to the high-oleic peanut + corn diet (P = 0.0002, Table 1). Normal-oleic peanuts have an O/L of approximately 1.5 to 1.0 (Chamberlin et al., 2014), whereas high-oleic peanuts have an O/L ≥ 9.0 (Davis et al., 2016, 2017). The calculated O/L ratio in the high-oleic peanut + corn diet and soybean meal + corn diets was 6.40 ± 0.10 and 1.344 ± 0.01, respectively (Table 1). Commonly, 95 out of 100 peanut kernels must pass the minimal fatty acid chemistry profile threshold of ≥74% oleic acid and ≤8% linoleic acid to be classified as a high-oleic peanut lot (Sweigart et al., 2011). Research has demonstrated that high-oleic peanut lots may not be meeting this threshold due to either contamination with conventional normal-oleic peanuts (Andersen et al., 1998; Sweigart et al., 2011) and/or the presence of immature peanut kernels within the lot that has yet to fully express the high-oleic oil chemistry of a ma-

ture kernel (Pattee et al., 1974; Singkham et al., 2010). Consequently, with kernel-to-kernel sampling within a lot of high-oleic peanuts there will be a natural distribution of O/L oil content (Davis et al., 2017), which may explain oleic acid values of 72.18 ± 0.180, linoleic acid values of 11.28 ± 0.20, and an O/L value of 6.40 ± 0.10 for oil chemistry of the high-oleic peanut + corn diet analyzed.

Layer Hen Performance There were no significant differences in the body weights of layer hens between the experimental treatment groups at any of the time points measured (week 1 to 10, Table 2). However, there was a significant week-to-week (wk) and treatment × week (trmt × wk) effect on body weights of the layer hens from both treatment groups, P = 0.029 and P = 0.0005, respectively (Table 2). This indicates a trend associated with the HO peanut diet for a body weight loss, whereas the control group had a weight gain over the same period. Feed conversion ratio was calculated as the ratio of egg mass (grams) to feed weight (grams) to determine the amount of feed utilized for egg production. Thus, layer hens with greater feed conversion ratios were more efficient with the utilization of feed for shell egg production. Although there were no overall effect on feed conversion (P = 0.077), feed conversion was affected at biweekly experimental time points (4, 6, 8, and 10 wk) for hens fed the control soybean meal diet in comparison to hens fed the high-oleic peanut diet (Table 2). Although the feed conversion at these time points

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Myristic acid (14:0) Myristoleic acid (14:1cis) Palmitic acid (16:0) Palmitoleic acid (16:1cis) Margaric acid (17:0) Stearic acid (18:0) Oleic acid (18:1) Linoleic acid (18:2) α -Linolenic acid (18:3cis) Arachidic acid (20:0) Gondoic acid (20:1cis) Behenic acid (22:0) Erucic acid (22:1cis) Lignoceric acid (24:0) Cerotic acid (26:0) Oleic/linoleic ratio (O/L) Saturated fat (%) P/S

Soybean + corn diet (SBM)

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Table 2. Performance of layer hens fed a diet with high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1 A.

Body weight (g) SBM 1 2 3 4 5 6 7 8 9 10

1,464 1,462 1,460 1,477 1,464 1,461 1,469 1,485 1,495 1,505

± ± ± ± ± ± ± ± ± ±

37 37 37 37 37 37 37 37 37 37

1469 1451 1445 1429 1436 1409 1429 1437 1421 1435

± ± ± ± ± ± ± ± ± ±

37 37 37 37 37 37 37 37 37 37

B.

Treatment (Trmt) Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm × Wk Trend

Feed intake(g feed/bird/day) P-value 0.93 0.83 0.78 0.37 0.59 0.33 0.45 0.37 0.17 0.19

SBM 154 112 87 96 91 107 102 113 110 107

± ± ± ± ± ± ± ± ± ±

3 3 3 3 3 3 3 3 3 3

HO-PN 155 104 78 96 84 113 105 113 109 113

± ± ± ± ± ± ± ± ± ±

3 3 3 3 3 3 3 3 3 3

Body weight

Feed conversion ratio (g egg/g feed)

P-value

SBM

HOPN

P-value

0.69 0.08 0.05 0.90 0.12 0.16 0.65 1.00 0.86 0.21

na na 0.568 ± 0.018 0.600 ± 0.018 0.630 ± 0.018 0.530 ± 0.018 0.491 ± 0.018 0.554 ± 0.019 0.522 ± 0.018 0.533 ± 0.018

na na 0.589 ± 0.018 0.536 ± 0.018 0.600 ± 0.018 0.452 ± 0.018 0.468 ± 0.018 0.487 ± 0.018 0.475 ± 0.018 0.466 ± 0.018

na na 0.42 0.01 0.24 0.003 0.36 0.01 0.06 0.01

Feed intake

Feed conversion ratio

F-test

P-value

F-test

P-value

F-test

P-value

0.63 0.64 < 0.0001 2.03 3.24 1.13 0.61 26.93

0.43 0.43 0.98 0.03 0.0005 0.34 0.81 < 0.0001

< 0.0001 1.30 0.04 128.97 3.24 1.98 0.76 3.31

0.97 0.26 0.84 < 0.0001 0.0005 0.03 0.67 0.07

5.65 0.71 1.33 31.2 2.64 1.99 1.29 3.14

0.02 0.40 0.25 < 0.0001 0.01 0.05 0.25 0.08

1 Layers (24 per dietary treatment) were fed a HO-PN or SBM diet for 10 wk. (A) Body weight, feed intake, and feed conversion ratio. Reported mean ± standard error; na = (not applicable). Feed conversion could not be calculated because ample time was needed for hens to be in full egg production. Trend test was used to compare the slopes (variable vs. time) for hens fed HOPN or SBM diets. (B) Treatment and interaction statistics.

was lower in the HO PM, the values were close in numerical value (Table 2). Thus, in the model, there were significant treatment effects (P = 0.022), week effects (P < 0.0001), treatment × week effects (trt × wk) P = 0.008). However, there were no significant differences in feed conversion between the treatment groups at week 3, 5, 7, and 9 (Table 2).

Egg Grading, Quality and Production All eggs produced at all time points (week 1 to 10) between both treatment groups were graded as USDA Grade AA of superior quality, with thick, firm egg whites and defect-free egg yolks. Additionally, the shells were clean and without defects. There were minimal number of blood spots or number of meat spots, and there was no statistical difference at the 95% confidence interval between eggs produced from both feeding treatment groups (data not shown). The most widely used measurement of albumen (egg white) quality is the Haugh unit, by Raymond Haugh in 1937 (Stadelmann et al., 1995). This method consists of measuring the height and thickness of the albumen. Hence, fresher, higher quality eggs have thicker egg whites and thus higher HU values. There were no significant differences in the egg HU (Table 3), number of eggs produced (Table 3), or the egg albumen height (Table 4) between the treatment groups at any of the time points measured (week 1 to 10), whereas there were significant week effects on these 3 variables (number of eggs P < 0.0001, HU P < 0.0001, albumen height P < 0.0001). Moreover, there was a significant trmt × rm effect on egg albumen height (P = 0.001, Table 4).

The yolk color Roche score was examined weekly to determine the yolk color intensity of eggs produced from both dietary treatments at all time points measured (Table 4). The yolk color Roche value was significantly greater at all time points (P < 0.0001) measured in eggs produced from layer hens fed the high-oleic peanut + corn diet in comparison to eggs produced from layer hens fed the soybean meal + corn diet (Table 4), indicating a more intense yellow-orange colored yolk in these eggs, with main effects of treatment (P < 0.0001). Additionally, there were significant week (P < 0.0001) and trmt × wk (P < 0.0001) effects on the yolk color Roche score, in which the Roche score diminished weekly particularly in eggs produced from layer hens fed the conventional soybean meal + corn diet. Moreover, there were significant rm × wk (P < 0.0001) effects on yolk color Roche, particularly only in eggs produced from layer hens fed the high-oleic peanut + corn diet. In contrast, Pesti et al. (2003) reported that yolk color in eggs produced from layer hens fed peanut meal to be slight and not noticeable upon casual observation. However, in this study there were clearly observable differences in the yolk color in eggs produced from layer hens fed the high-oleic peanut diet in comparison to eggs produced from layer hens fed the soybean meal diet. Eggs produced from layer hens fed the soybean meal + corn diet had egg weights that were significantly greater in size than eggs from layer hens fed the higholeic peanut + corn diet at all time points measured (week 1 to 10), with significant treatment (P = 0.0001) and week (P = 0.003) effects on egg weights (Table 3). Interestingly, the total number of eggs produced

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Week Week Week Week Week Week Week Week Week Week

HOPN

8

TOOMER ET AL.

Table 3. Egg production parameters and quality of eggs produced by layer hens fed a diet with high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1 A.

Number of eggs produced SBM 1 2 3 4 5 6 7 8 9 10

± ± ± ± ± ± ± ± ± ±

5.58 7.46 5.67 6.42 6.50 6.38 6.00 6.79 6.38 6.38

0.24 0.24 0.24 0.24 0.24 0.24 0.24 0.24 0.24 0.24

HO-PN

P-value

± ± ± ± ± ± ± ± ± ±

0.72 0.55 0.81 0.72 0.47 0.06 0.90 0.63 0.63 0.81

5.71 7.25 5.75 6.29 6.25 6.21 6.04 6.63 6.21 6.29

B.

0.24 0.24 0.24 0.24 0.24 0.24 0.24 0.24 0.24 0.24

SBM 58.5 60.3 59.0 58.7 58.5 59.8 59.5 59.3 59.5 59.5

± ± ± ± ± ± ± ± ± ±

0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7

HO-PN

P-value

± ± ± ± ± ± ± ± ± ±

0.01 < 0.0001 0.003 < 0.0001 0.009 0.001 0.004 0.003 0.02 0.02

55.8 56.5 55.9 54.8 55.8 56.3 56.5 56.2 57.1 57.1

# Eggs produced (#) F-test

Treatment (Trmt) Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm × Wk

0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7

SBM 92.6 95.2 92.8 94.6 94.4 94.1 92.2 92.7 88.1 88.1

± ± ± ± ± ± ± ± ± ±

0.8 0.8 0.8 0.9 0.8 0.8 0.8 0.9 0.8 0.8

F-test

0.77 0.70 0.50 < 0.0001 0.97 0.08 0.95

P-value

± ± ± ± ± ± ± ± ± ±

0.67 0.45 0.60 0.92 0.98 0.21 0.88 0.65 0.86 0.86

0.8 0.8 0.8 0.9 0.8 0.8 0.8 0.9 0.8 0.8

Haugh unit (HU)

P-value

14.30 1.00 0.09 2.89 0.65 0.43 0.69

HO-PN 93.1 94.3 93.4 94.8 94.3 95.6 92.0 92.2 87.9 87.9

Egg weight (g)

P-value

0.09 0.15 0.47 24.10 0.35 1.70 0.40

Haugh unit (HU)

F-test

0.0001 0.32 0.76 0.003 0.75 0.92 0.72

P-value

0.01 1.19 11.37 40.97 0.66 1.08 1.22

0.94 0.28 0.002 < 0.0001 0.74 0.38 0.28

1 Twenty-four animals per dietary treatment were fed either an HO-PN or SBM diet for 10 wk. (A) Egg data. Egg Haugh unit (HU) was calculated to determine egg albumen quality. Reported mean ± standard error. (B) Treatment and interaction statistics.

Table 4. Egg quality and β -carotene levels of eggs produced by layer hens fed a diet of high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1 Albumen height2 (mm)

A. SBM Week Week Week Week Week Week Week Week Week Week

1 2 3 4 5 6 7 8 9 10

8.60 9.18 8.62 9.01 8.93 8.92 8.55 8.64 7.82 7.82

± ± ± ± ± ± ± ± ± ±

0.16 0.16 0.16 0.16 0.16 0.16 0.16 0.16 0.16 0.16

HO-PN

P-value

± ± ± ± ± ± ± ± ± ±

0.78 0.10 0.92 0.43 0.53 0.43 0.38 0.28 0.41 0.41

8.54 8.81 8.60 8.82 8.79 9.09 8.35 8.39 7.64 7.64

0.16 0.15 0.16 0.16 0.16 0.16 0.16 0.16 0.15 0.15

Treatment (Trmt) Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm × Wk Trend

SBM 5.09 5.74 4.67 4.18 4.48 4.11 3.76 3.72 2.87 2.87

± ± ± ± ± ± ± ± ± ±

0.15 0.15 0.15 0.15 0.15 0.15 0.15 0.16 0.15 0.15

HOPPN

P-value

SBM

HO-PN

P-value

± ± ± ± ± ± ± ± ± ±

< 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001

11.44 ± 0.35 na na na 7.74 ± 0.35 na na na na 8.27 ± 0.26

12.21 ± 0.35 na na na 12.75 ± 0.39 na na na na 12.40 ± 0.36

0.12 na na na < .0001 na na na na < .0001

6.63 6.97 6.64 6.14 6.48 6.49 5.62 6.10 5.96 5.96

Albumen height2

B.

β -Carotene4 (ppm)

Yolk color3 (Roche score 1 to 15)

0.15 0.15 0.15 0.16 0.15 0.15 0.15 0.16 0.15 0.15

β -Carotene4

Yolk color3

F-test

P-value

F-test

P-value

F-test

P-value

0.71 0.60 11.60 38.20 0.89 1.05 1.04 na

0.40 0.44 0.001 < 0.0001 0.53 0.40 0.40 na

301.00 1.22 4.12 49.60 11.16 5.43 1.47 60.40

< 0.0001 0.28 0.05 < 0.0001 < 0.0001 < 0.0001 0.16 < 0.0001

128.06 2.69 1.58 45.90 55.09 6.80 9.42 128.06

< 0.0001 0.11 0.22 < 0.0001 < 0.0001 0.002 0.0002 na

1 Twenty-four animals per dietary treatment were fed either an HO-PN or SBM diet for 10 wk. (A) Egg data. Each value represents the mean ± the standard error. Trend test compared the slopes (variable vs. time) for the 2 diets. 2 Egg albumen height (mm) was calculated to determine egg albumen quality. 3 Yolk color was determined using the Roche Color Fan color index 1 to 15 to distinguish the yolk color from lightest to darkest color intensity. 4 Eggs collected weeks 1, 5, and 10 were analyzed for β -carotene content; testing at other time points was cost prohibitive and reported as na = not applicable. (B) Treatment and interaction statistics.

from layer hens fed the high-oleic peanut + corn diet categorized by weight (United States Department of Agriculture egg classification system) was 60% medium size eggs, 35% large size eggs, 3% extra-large size eggs, and 2% small size eggs. The total number of eggs produced from layer hens fed the soybean meal + corn diet

categorized by weight (United States Department of Agriculture egg classification system) was 66% large size eggs, 24% medium size eggs, 10% extra-large size eggs, and 0.3% small size eggs. These results are similar to results reported by Van Elswyk et al. (1994) demonstrating that layer hens fed unsaturated fatty acids in

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Week Week Week Week Week Week Week Week Week Week

Egg weight (g)

9

Total palmitoleic acid3 F-test P-value 643.26 < 0.0001 0.25 0.62 0.09 0.77 0.32 0.73 61.32 < 0.0001 2.00 0.14 0.16 0.860 Total stearic acid3 F-test P-value 46.60 < 0.0001 0.03 0.86 1.01 0.32 116.20 < 0.0001 1.54 0.22 1.33 0.27 2.29 0.110 Total palmitic Acid3 F-test P-value 74.8 < 0.0001 0.01 0.90 1.05 0.31 28.88 < 0.0001 10.68 < 0.0001 1.12 0.33 0.01 0.99 Total choleserol2 F-test P-value 10.50 < 0.0001 0.34 0.06 0.08 0.08 7.38 < 0.0001 2.38 0.10 4.79 0.01 0.19 0.82 Trmt Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm × Wk

B.

1 Twenty-four animals per dietary treatment were fed either an HO-PN or SBM diet for 10 wk. (A) Egg data. Eggs collected at 1, 5, and10 wk were analyzed by ATC Scientific for lipid and fatty acid content. Each value represents the mean ± standard error. 2 Units of mg/100 g sample. 3 Units of g/100 g sample. (B) Treatment and interaction statistics.

Total oleic acid3 F-test P-value 653.36 < 0.0001 0.37 0.55 0.34 0.56 90.25 < 0.0001 43.04 < 0.0001 1.69 0.19 0.69 0.51

< 0.0001 < 0.0001 < 0.0001 48.2 ± 0.5 53.1 ± 0.5 56.0 ± 0.5 39.5 ± 0.5 38.2 ± 0.5 41.6 ± 0.5 < 0.0001 < 0.0001 < 0.0001 1.98 ± 0.06 1.52 ± 0.06 1.52 ± 0.06 2.88 ± 0.06 3.29 ± 0.06 3.37 ± 0.06 < 0.0001 < 0.0001 < 0.0001 7.37 ± 0.13 5.91 ± 0.14 6.11 ± 0.13 8.80 ± 0.13 7.67 ± 0.13 8.05 ± 0.13 < 0.0001 < 0.0001 < 0.0001 23.1 ± 0.2 21.4 ± 0.2 21.0 ± 0.2 25.9 ± 0.2 26.9 ± 0.2 26.9 ± 0.2 0.35 0.01 < 0.0001 385 ± 6 373 ± 7 392 ± 6 377 ± 6 349 ± 6 360 ± 6 Week 1 Week 5 Week 10

Total oleic acid3 (18:1) HO-PN P SBM Total palmitoleic acid3 (16:1) SBM HO-PN P Total stearic acid3 (18:0) SBM HO-PN P Total palmitic acid3 (16:0) SBM HO-PN P P Total choleserol2 HO-PN SBM

Similar to the egg yolk color, β -carotene levels were significantly greater in eggs produced from layer hens fed the high-oleic peanut + corn diet in comparison to eggs produced from layer hens fed the soybean meal + corn diet at week 5 (P < 0.0001, Table 4) and week 10 (P < 0.0001, Table 4). Hence, implying the β -carotene content in the eggs produced was influenced by the lipid/fatty acid composition and/or polyphenolic components of the high-oleic peanuts. β -Carotene is a carotenoid found in plants that gives plants rich yellow and deep orange hues and is a precursor of vitamin A (retinol). Today, there is a growing body of scientific literature, which indicates the health benefits of β -carotene and other carotenoids in the prevention of chronic diseases in humans (Woodside et al., 2015; Zaheer, 2017). Carotenoids represent a group of lipid soluble antioxidant-rich bioactive compounds present in plants. Conventional commercially produced eggs yolks are rich in carotenoids, lutein, and zeaxanthin (Zaheer, 2017). However, lutein and zeaxanthin content within the egg is highly subject to

A.

Egg β-Carotene, Lipid and Fatty Acid Content

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the diet produced reduced size eggs in response to the menhaden (fish) oil feeding in comparison to the controls. This attribute could be of potential benefit within the commercial egg production industry by reducing the number of oversized eggs produced in older laying hens. Layer hens typically start to lay eggs around 5 mo (20 to 21 wk) of age and continue to lay for 12 mo to 52 wk (FAO, 2003). Birds typically begin producing eggs in their 20th or 21st week and continue for slightly over a year (52 wk), with eggs tending to increase in size until the end of the egg production cycle (Silversides and Scott, 2001; Padhi et al., 2013). Egg size has been shown to be influenced by genetic factors (breed, breeding system, body weight, and age), nutritional factors (energy and feed intake, protein, lipids, feeding program), and pullet management (lighting program, stress, and disease management). For example, egg size is directly influenced by body weight; in that for every 45 g of body weight increase from 18 wk of age, there is a 0.5-g increase in egg size (Duraisamy, 2011). Therefore, as the layer hen ages in the production cycle and increases in body weight there is a proportionate increase in egg size (Duraisamy, 2011). Thus, a feeding program, which aims to reduce egg size, is of significant interest in the commercial egg production industry as a mechanism to manage egg size with increasing production age of layer hens. In conclusion, the only reduction in performance due to feeding layer hens’ high-oleic peanut + corn diet was smaller egg weights over the 10-wk trial, while all other production parameters were similar. In future studies, we aim to examine the effects of a high-oleic peanut diet on egg size in late-stage egg production.

Table 5. Lipid and fatty acid content of eggs produced by layer hens fed a diet with high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1

EGGS FROM LAYER HENS FED HIGH-OLEIC PEANUTS

P-value < 0.0001 0.13 0.80 < 0.0001 < 0.0001 0.22 0.16 F-test 101.12 2.39 0.07 107.16 6.72 1.55 1.87 P-value < 0.0001 0.07 0.54 < 0.0001 0.09 0.01 0.41 F-test 26.26 3.33 0.39 38.10 2.54 4.58 0.91 P-value < 0.0001 0.69 0.27 < 0.0001 < 0.0001 0.01 0.34 F-test 39.59 0.16 1.24 50.58 8.33 4.51 1.10 Trmt Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm × Wk

1 Twenty-four animals per dietary treatment were fed either an HO-PN or SBM diet for 10 wk. (A) Egg data (units g/100 g sample). Eggs collected at 1, 5, and 10 wk were analyzed by ATC Scientific for lipid and fatty acid content. Each value represents the mean ± standard error. (B) Treatment and interaction statistics.

P-value < 0.0001 0.29 0.77 < 0.0001 < 0.0001 0.40 0.25 F-test 150.23 1.13 0.09 202.07 13.90 0.94 1.41

1.11 ± 0.02 < 0.0001 0.84 ± 0.03 < 0.0001 0.84 ± 0.03 < 0.0001 Total linoleic acid (18:2) 1.40 ± 0.03 1.24 ± 0.03 1.05 ± 0.03 < 0.0001 < 0.0001 < 0.0001 acids

1.26 ± 0.03 0.93 ± 0.03 1.00 ± 0.03 Omega 6 fatty 1.59 ± 0.03 1.31 ± 0.03 1.19 ± 0.03 0.013 ± 0.001 < 0.0001 0.004 ± 0.002 0.09 0.007 ± 0.002 < 0.0001 Omega 3 fatty acids 0.023 ± 0.002 0.007 ± 0.002 0.014 ± 0.002 4.14 ± 0.31 0.71 6.29 ± 0.36 < 0.0001 4.59 ± 0.33 < 0.0001 n9 elaidic acid (18:1) 4.30 ± 0.32 8.78 ± 0.32 6.99 ± 0.32 Week 1 Week 5 Week 10 B.

P-value HO-PN SBM P-value HO-PN

Omega 6 fatty acids

SBM P-value HO-PN

Omega 3 fatty acids

SBM P-value HO-PN

n9 elaidic acid (18:1)

SBM A.

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oxidation during, egg processing, storage, and transport and/or cooking (Zaheer, 2017). Studies by Olson et al. (2008) demonstrated that carotenoid pigments found naturally within plant-based feedstuff of layer hen diets are transferred to the egg yolk of the eggs produced. Egg yolk color has been shown to also be affected by several different components within the diet, such as lipid profile (Suksombat et al., 2006) and type and concentration of carotenoids (Leeson and Caston, 2004; Karadas et al., 2006). Interestingly, studies by Chung et al. (2004) revealed that the carotenoid lutein was more bioavailable from enriched eggs in comparison to lutein found in spinach and dietary supplements. This research also demonstrated improved intestinal absorption of lutein when consumed with dietary lipids, suggesting that eggs may be superior delivery system for some carotenoids (Chung et al., 2004). Karadas et al. (2006) demonstrated that supplementing the diets of female quail with natural carotenoids (alfafa concentrated, tomato powder, and/or marigold extract) enriched egg yolks with carotenoids lutein, zeaxanthin, lycopene, and β -carotene. This work parallels earlier research by Jiang et al. (1994), which demonstrated that dietary supplementation of vitamin E and β -carotene in the diets of layer hens enriched egg yolk proportionately with vitamin E and β -carotene levels in comparison to the controls. However, these feeding regimens are not commercially viable due to the associated cost of the inclusion of specialty feed ingredients (dietary carotenoid supplements, alfalfa, tomato powder, marigold) in the diets of layer hens to enrich the eggs (Jiang et al., 1994). In general, this study implies that feeding layer hens’ high-oleic peanuts may be an economical and effective means to enrich egg nutrition and yolk color and may be of value to peanut and egg producers globally. Today, consumer demand for food products of superior quality and nutrition has led to increased interest and research in livestock feeding studies with aims to enrich and/or modify the lipid and nutrient profile of the eggs and meat produced. Although current feeding studies have demonstrated the health benefits of the moderate consumption of high-oleic peanuts in the diet (Moreira Alves et al., 2014, Alves et al., 2014; Moreira et al., 2016; Barbour et al., 2017), there are no previous studies which have examined the use, economic, and health advantages of utilizing high-oleic peanuts as a feed ingredient to enrich eggs and meat produced with heart healthy monounsaturated fatty acids. Earlier feeding trials revealed that the fatty acid composition of poultry meat (Marion and Woodroof, 1963; Hulan et al., 1988; Yau et al., 1991) and egg yolk (Cruickshank, 1934; Sell et al., 1968; Ohtake and Hoshino, 1976; El-Hussainy et al., 1983) is readily altered by dietary manipulation of fatty acids. Therefore, in this study we aimed to determine the effects of feeding layer hens’ high-oleic peanut diet on the fatty acid and

Total linoleic acid (18:2)

TOOMER ET AL.

Table 6. Fatty acid content of eggs produced by layer hens fed a diet with high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1

10

11

EGGS FROM LAYER HENS FED HIGH-OLEIC PEANUTS

Table 7. Total linoleic and oleic fatty acid content of eggs produced by layer hens fed a diet with high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1 A.

2 3 4 6 7 8 9

Total oleic acid (18:1) (%)

SBM

HO-PN

P-value

± ± ± ± ± ± ±

9.5± 0.4 10.3 ± 0.4 10.4 ± 0.4 10.4 ± 00.4 10.0 ± 0.4 11.1 ± 0.40 10.0 ± 0.37

< 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001

13.5 12.6 14.0 13.1 13.4 13.3 12.7

B.

Treatment (Trmt) Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm × Wk Trend

0.4 0.4 0.4 0.4 0.4 0.4 0.4

SBM 36.9 36.6 28.2 29.7 29.5 31.5 31.5

± ± ± ± ± ± ±

0.6 0.6 0.5 0.5 0.6 0.6 0.6

Total linoleic acid (18:2)

Oleic: linoleic ratio

HO-PN

P-value

± ± ± ± ± ± ±

< 0.0001 0.004 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001

49.6 38.9 37.1 37.2 38.9 40.7 40.8

0.5 0.6 0.5 0.5 0.5 0.5 0.5

Total oleic acid (18:1)

SBM 2.76 3.03 2.03 2.28 2.21 2.38 2.49

± ± ± ± ± ± ±

0.09 0.09 0.09 0.09 0.09 0.09 0.09

HO-PN

P-value

± ± ± ± ± ± ±

< 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001

5.28 3.80 3.59 3.60 3.89 4.21 4.22

0.09 0.09 0.09 0.09 0.09 0.09 0.09

Oleic: linoleic ratio

F-test

P-value

F-test

P-value

F-test

P-value

165.10 0.29 0.45 1.52 1.65 1.34 1.8 na

< 0.0001 0.59 0.51 0.17 0.14 0.24 0.10 na

541.07 0.94 1.36 96.89 18.40 3.59 0.27 0.45

< 0.0001 0.34 0.25 < 0.0001 < 0.0001 0.002 0.95 0.50

672.75 0.03 0.14 44.46 19.04 1.58 1.68 0.04

< 0.0001 0.86 0.71 < 0.0001 < 0.0001 0.16 0.13 0.84

1 Twenty-four animals per dietary treatment were fed either an HO-PN or SBM diet for 10 wk. (A) Egg data. Eggs collected at weeks 2, 3, 4, 6, 7, 8, and 9 were analyzed for lipid and fatty acid content at the Market Quality & Handling Research Unit at. Each value represents the mean ± standard error. Trend test compared the slopes (variable vs. time) of the 2 diets. (B) Treatment and interaction statistics.

lipid profile of shell eggs produced from layer hens for 10 wk. In this study, total cholesterol from eggs produced from layer hens fed the high-oleic peanut + corn diet was significantly greater than the total cholesterol from eggs produced from layer hens fed the conventional soybean meal + corn diet at week 5 (P = 0.01) and week 10 (P < 0.0001) experimentally (Table 5), with significant treatment effects (P < 0.0001), wk effects (P < 0.0001), and rm × wk effects (P = 0.010). Total saturated fatty acids, palmitic acid and stearic acid, were significantly higher in the eggs produced from layer hens fed the conventional soybean meal + corn diet in comparison to eggs produced from layer hens fed the high-oleic peanut + corn diets at week 1 (P < 0.0001), week 5 (P < 0.0001), and week 10 (P < 0.0001) of the experiment (Table 5), with significant treatment (P < 0.0001), wk (P < 0.0001), and trmt × wk effects (P < 0.0001 palmitic acid only). Additionally, trans fat n9 elaidic acid was significantly higher in eggs produced from layer hens fed the conventional soybean meal + corn diet at week 5 (P < 0.0001) and week 10 (P < 0.0001) experimentally (Table 6), with trmt (P < 0.0001), wk (P < 0.0001), trmt × wk (P < 0.0001), and rm × wk (P = 0.010) effects. Eggs produced from layer hens fed the soybean meal + corn diet had significantly higher content of monounsaturated fatty acid, palmitoleic acid (Table 5), and polyunsaturated fatty acid, linoleic acid (Table 6) at week 1 (P < 0.0001), week 5 (P < 0.0001), and week 10 (P < 0.0001) experimentally, with trmt (P < 0.0001) and trmt × wk (P < 0.0001) effects. Conversely, the content of monounsaturated fatty acid,

oleic acid content was significantly higher in eggs produced from layer hens fed the high-oleic peanut + corn diet at week 1 (P < 0.0001), week 5 (P < 0.0001), and week 10 (P < 0.0001) experimentally (Table 5), with trmt (P < 0.0001), wk (P < 0.0001), and trmt × wk (P < 0.0001) effects. Also, eggs produced at week 2, 3, 4, 6, 7, 8, and 9 by hens fed the high-oleic peanut + corn diet had significantly greater total oleic acid content in comparison to eggs produced by hens fed the soybean meal conventional diet, with significant treatment (P < 0.0001), wk (P < 0.0001), trmt × wk (P < 0.0001), and rm × wk (P = 0.002) effects (Table 7). Moreover, eggs produced from layer hens fed the higholeic peanut + corn diet had higher O/L ratios than the other treatment group at all the time points measured (P < 0.0001, Table 7), with significant treatment (P < 0.0001), wk (P < 0.0001), and trmt × wk (P < 0.0001) effects. In parallel, eggs produced from layer hens fed the soybean meal + corn diet had significantly greater levels of total linoleic acid in comparison to eggs produced from layer hens fed the high-oleic peanut + corn diet at weeks 2, 3, 4, 6, 7, 8, and 9 (P < 0.0001, Table 7), with significant treatment effects (P < 0.0001). Lastly, chemical analysis was conducted to determine the total contents of omega 3 and omega 6 family of PUFA. Although the levels of omega 3 and omega 6 PUFA were very low in eggs from both treatment groups, eggs produced from layer hens fed the soybean meal + corn diet had significantly greater levels of omega 3 and omega 6 PUFA in comparison to eggs produced from layer hens fed the high-oleic peanut + corn diets at week 1 (omega 3 P < 0.0001, omega 6 P < 0.0001), week 5 (omega 3 P = 0.09, omega 6

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Week Week Week Week Week Week Week

Total linoleic acid (18:2) (%)

0.84 0.32 < 0.0001 0.04 0.09 0.37

1 Twenty-four animals per dietary treatment were fed either an HO-PN or SBM diet for 10 wk. (A) Egg data. Eggs collected at weeks 1, 5, and 10 were analyzed by ATC Scientific for lipid and fatty acid content (g/100 g sample). Each value represents the mean ± standard error. (B) Treatment and interaction statistics.

P-value 0.13 0.05 0.23 < 0.0001 < 0.0001 0.24 0.35 2.27 3.93 1.48 45.27 8.17 1.44 1.07

F-test

P-value < 0.0001 0.14 0.41 < 0.0001 0.02 0.01 0.04 F-test 34.00 2.24 0.70 37.91 3.98 4.90 1.01 P-value < 0.0001 0.58 0.31 0.20 0.16 0.20 0.31 F-test 24.24 0.31 1.05 1.66 1.85 1.64 1.20 P-value < 0.0001 0.03 < 0.0001 < 0.0001 0.04 0.18 0.41 19.16 5.04 9.72 7.56 3.32 1.75 0.90 F-test P-value

< 0.0001

F-test 23.24 0.04 1.00 25.26 3.44 2.51 1.01 Trmt Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm× Wk

Total n6 c,c,c gamma-linolenic acid (18:3) Total linolenic acid (18:3) Total margaric acid (17:0) Total pentadecylic acid (15:0) Total tridecanoic acid (13:0) B.

− 2.3E-18 ± 0.006 0.010 ± 0.006 0.25 0.178 ± 0.006 0.193 ± 0.006 0.07 0.248 ± 0.015 0.124 ± 0.015 < 0.0001 0.121 ± 0.009 0.065 ± 0.009 < 0.0001 0.007 ± 0.006 0.046 ± 0.007 < 0.0001 0.164 ± 0.006 0.196 ± 0.007 < 0.0001 0.084 ± 0.015 0.041 ± 0.017 0.06 2.8 E-17 ± 0.009 0.006 ± 0.011 0.69 1.1E-17 ± 0.006 0.025 ± 0.006 < 0.0001 0.158 ± 0.006 0.193 ± 0.006 < 0.0001 0.136 ± 0.015 0.066 ± 0.015 < 0.0001 0.061 ± 0.009 0.075 ± 0.010 0.29 0.017 ± 0.002 0.013 ± 0.002 0.16 0.026 ± 0.002 0.015 ± 0.002 < 0.0001 0.031 ± 0.002 0.021 ± 0.002 < 0.0001 Week 1 Week 5 Week 10

HO-PN SBM SBM

HO-PN

P

SBM

HO-PN

P

SBM

HOPN

P

SBM

HO-PN

P

Total n6 c,c,c gamma-linolenic acid (18:3) Total linolenic acid (18:3) Total margaric acid (17:0) Total pentadecylic acid (15:0) Total tridecanoic acid (13:0) A.

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P < 0.0001), and week 10 (omega 3 P < 0.0001, omega 6 P < 0.0001) experimentally (Table 6). Moreover, there were significant treatment (P < 0.0001) and wk (P < 0.0001) effects on omega 3 and omega 6 PUFA contents in eggs in both treatment groups (Table 6). Although the content of total tridecanoic in eggs produced from both feeding treatments was ≤ 0.031, total tridecanoic acid content (Table 8) was significantly greater in eggs produced from layer hens fed the soybean meal + corn diet at week 5 (P < 0.0001) and week 10 (P = 0.000), with significant treatment (P < 0.0001), week (P < 0.0001), and trmt × wk (P = 0.040) effects. Similarly, levels of total pentadecyclic acid and total margaric acid in eggs of were very low. However, eggs produced from layer hens fed the high-oleic peanut + corn diet had significantly greater contents of total pentadecyclic acid and total margaric acid in comparison to eggs produced from a traditional soybean meal diet at weeks 5 and 10 (Table 8). Moreover, there were significant treatment (P < 0.0001), room (P = 0.030), trmt × rm (P < 0.0001), wk (P < 0.0001), and trmt × wk (P = 0.040) effects on total pentadecyclic levels between treatment groups (Table 8). There were only significant treatment effects (P < 0.0001) on total margaric levels between the treatment groups (Table 8). Total linolenic acid levels were significantly greater in eggs produced from layer hens fed the soybean meal + corn diet at week 1 (P < 0.0001) and week 10 (P < 0.0001) in comparison to eggs of the other treatment group (Table 8), with significant treatment (P = 0.0001), wk (P < 0.0001), trmt × wk (0.020), and rm × wk (P = 0.010) effects. Nevertheless, total n6, c, c, c, gamma-linolenic acid content was only significantly greater in eggs produced from layer hens fed the soybean meal + corn diet at week 1 (P < 0.0001) in comparison to eggs produced from layer hens fed the high-oleic peanut + corn diet (Table 8), with significant wk (P < 0.0001) and trmt × wk (P < 0.0001) effects. Total homo-gamma-linolenic acid levels were significantly greater only at week 10 (P = 0.010) in eggs produced from layer hens fed the high-oleic peanut + corn diet in comparison to the other treatment group (Table 9), with significant room (P < 0.0001), wk (P < 0.0001), rm × wk (P < 0.0001), and trmt × rm × wk (P = 0.010) effects. Total arachidonic acid levels were significantly greater only at week 1 in eggs produced from layer hens fed the soybean meal + corn diet in comparison to the other treatment group, with significant rm (P < 0.0001), trmt × rm (P = 0.020), wk (P < 0.0001), trmt × wk (P = 0.040), and rm × wk (P = 0.020) effects (Table 9). The remaining fatty acids analyzed included caproic acid, caprylic acid, capric acid, undecanoic acid, myristoleic acid, gadoleic acid, and brassic acid. The contents of these fatty acids were ≤ 0.04 g/100 g of feed or were undetectable (data not shown).

P

TOOMER ET AL.

Table 8. Fatty acid content of eggs produced by layer hens fed a diet with high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1

12

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EGGS FROM LAYER HENS FED HIGH-OLEIC PEANUTS

Table 9. Fatty acid content of eggs produced by layer hens fed a diet with high-oleic peanuts (HO-PN) or a conventional diet with soybean meal (SBM).1 A.

Week 1 Week 5 Week 10

Treatment (Trmt) Room (Rm) Trmt × Rm Week (Wk) Trmt × Wk Rm × Wk Trmt × Rm × Wk

Total arachidonic acid (20:4)

SBM

HO-PN

P-value

SBM

HO-PN

P-value

0.016 ± 0.005 0.020 ± 0.005 0.091 ± 0.005

0.010 ± 0.005 0.021 ± 0.006 0.113 ± 0.006

0.47 0.90 0.01

0.152 ± 0.007 0.046 ± 0.007 1.3E-17 ± 0.007

0.132 ± 0.007 0.061 ± 0.008 2.7E-17 ± 0.007

0.03 0.15 1.00

Total homo-gamma-linolenic acid

Total arachidonic acid

F-test

P-value

F-test

P-value

1.68 23.02 2.00 165.27 3.48 10.72 4.75

0.20 < 0.0001 0.16 < 0.0001 0.03 < 0.0001 0.01

0.11 11.87 5.57 233.53 3.25 3.97 1.44

0.47 < 0.0001 0.020 < 0.0001 0.04 0.02 0.24

1 Twenty-four animals per dietary treatment were fed either an HO-PN or SBM diet for 10 wk. (A) Egg data. Eggs collected at weeks 1, 5, and 10 were analyzed by ATC Scientific for lipid and fatty acid content (g/100 g sample). Each value represents the mean ± standard error. (B) Treatment and interaction statistics.

Figure 2. Immunoreactivity of protein extracts from pooled total egg samples from eggs produced by layer hens fed a diet with high-oleic peanuts (HOPN) vs. a conventional diet with soybean meal (SBM) at week 1. Protein extracts from pooled egg samples (75 μ g per lane) were run electrophoretically on a 10% polyacrylamide gel. To determine immunereactivity, resolved proteins were transferred to a nitrocellulose membrane and immunoblotted with rabbit IgG anti-peanut agglutinin antibody (1:1000). Biodetection was determined with chromogenic peroxidase substrate-based detection of HRP activity. (A) Immunoblotting results of total egg proteins from hen(s) #1 to 12 (HOPN) and hen(s) 13 to 24 (SBM). Extracted proteins (75 μ g) from peanut flour were utilized as a positive control. (B) Immunoblotting results of total egg proteins from hen(s) #25 to 36 (SBM) and hen(s) 37 to 48 (HOPN). Extracted proteins (75 μ g) from peanut flour were utilized as a positive control.

Temporal Changes of Egg Fatty Acids and Lipids Two components enter into the influence of diet and the rate at which the dietary change will affect the performance criteria. First is hen age and second is the nutrient composition of the diet. The age of the hen

has been shown (Al-Batshan et al., 1994) to negatively influence the utilization of some of the nutrients in the diet. Therefore, as diets are changed during a phase feeding program in the layer industry understanding interaction of age and the speed in which the diet change will have an impact on the production criteria is important. This study also examined the rate (week to week)

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B.

Total homo-gamma-linolenic acid (20:3)

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TOOMER ET AL.

at which the feeding of high-oleic peanuts had on hen performance and the quality and nutrient composition of the eggs they produced. This study demonstrates that egg fatty acid and lipid composition can be greatly influenced rapidly (approximately 1 to 2 wk) with dietary modification of dietary fat fed to egg-producing hens. Eggs from the high-oleic peanut feeding group had a reduction in the content of saturated fatty acids (total palmitic and total stearic) and an increase in oleic fatty acid content that was very apparent after 1 to 2 wk of feeding the high-oleic peanut diet in comparison to the controls (Table 5). Additionally, there was a significant trmt × wk interaction shown in Table 2 indicating that the introduction of a new diet to a laying hen can have an impact on body weight and feed intake within 7 d of the change. However, the influence of a dietary change on egg production characteristics does not come into play until after approximately 4 wk as shown by the drop in feed conversion of 0.536 g egg/g feed for the hens on the high-oleic peanut diet, which accounts for the changes in production parameters of feed consumption and egg weight. Interestingly, there was no influence of diet on egg quality parameters, with the principal factor on quality

being the hen age. In the high-oleic peanut diet, there was an impact on yolk color and β -carotene content in eggs from the high-oleic peanut diet having darker (P < 0.0001) yellow yolks in comparison to eggs from the soybean meal diet (Table 4). After 1 wk of feeding a high-oleic peanut diet, the yolk color DSM was significantly greater than the controls. This effect was more pronounced over the experimental time period, with egg yolk color score decreasing in intensity weekly of control eggs, such that by week 9 the yolk color score of the control eggs were approximately 2-fold less than eggs from the high-oleic peanut feeding group. In addition, there was a significant temporal reduction in β -carotene content in eggs produced from hens fed the soybean meal control diet from week 1 to 10 in comparison to eggs produced from hens fed the high-oleic peanut diet over time. In summary, the predominant saturated fatty acids found within experimental egg and feed samples were palmitic fatty acid (16:0) and stearic fatty acid (18:0), with concentrations of these saturated fatty acids higher in feed samples and eggs produced from hens fed the conventional soybean meal +corn diet. Additionally, the unsaturated trans fatty acid that has been implicated in heart disease (Tardy et al., 2011),

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Figure 3. Immunoreactivity of protein extracts from pooled total egg samples from eggs produced by layer hens fed a diet with high-oleic peanuts (HOPN) vs. a conventional diet with soybean meal (SBM) at week 5. Protein extracts from pooled egg samples (75 μ g per lane) were electrophoretically ran on a 10% polyacrylamide gel. To determine immunereactivity, resolved proteins were transferred to a nitrocellulose membrane and immunoblotted with rabbit IgG anti-peanut agglutinin antibody (1:1000). Biodetection was determined with chromogenic peroxidase substrate-based detection of HRP activity. (A) Immunoblotting results of total egg proteins from hen(s) #1 to 12 (HOPN) and hen(s) 13 to 24 (SBM). Extracted proteins (75 μ g) from peanut flour were utilized as a positive control. (B) Immunoblotting results of total egg proteins from hen(s) #25 to 36 (SBM) and hen(s) 37 to 48 (HOPN). Extracted proteins (75 μ g) from peanut flour were utilized as a positive control.

EGGS FROM LAYER HENS FED HIGH-OLEIC PEANUTS

15

n9 t elaidic, was significantly elevated in eggs produced from layer hens fed the conventional soybean meal corn diet. Overall, the predominant unsaturated fatty acids found within the experimental egg and feed samples were palmitoleic acid (omega 7 monounsaturated fatty acid) and oleic acid (omega 9 monounsaturated fatty acid), with contents of palmitoleic acid in higher concentrations in the feed and eggs produced from hens fed the conventional soybean meal + corn diet. Conversely, oleic acid content was greatest in the feed and egg samples from layer hens fed the high-oleic peanut + corn diet.

Potential Transfer of Peanut Allergens in Eggs produced in Layer Hens fed the High-Oleic Peanut Diet To determine the potential transfer of allergenic peanut proteins to eggs produced from layer hens fed a high-oleic peanuts + corn diet, protein extracts from all eggs were analyzed by western immunoblotting methods at weeks 1 (Figure 2), 5 (Figure 3), and 10 (Figure 4) experimentally. All egg protein extracts from both treatments at each time point were non-reactive

with rabbit anti-peanut agglutinin antibodies, whereas only protein samples from the positive control of peanut flour were reactive. In summary, all egg samples were non-allergenic for peanut antigens. In contrast, earlier studies by Armentia et al. (2006) and Faeste et al. (2014) reported hypersensitivity reactions in sensitized patients to allergens found in the meat consumed of animals fed diets containing the allergenic proteins. Nevertheless, these studies investigated the transfer of parasitic fish larva (Anisakis simplex, parasitic nematode found in fish that causes allergic reaction in some human consumers) allergenic peptides from the diet to the meat produced, in contrast to examination of the transfer of proteins/peptides found in feedstock rations. Therefore, this study aimed to address the safety assessment of eggs produced from layer hens fed a high-oleic peanut diet by utilizing immunoblotting techniques with peanut-specific antibodies. In summary, these studies demonstrate that eggs produced from layer hens fed high-oleic peanuts are non-reactive with peanut antibodies and therefore should not elicit an allergic response in peanutsensitized individuals. Furthermore, earlier research has demonstrated that a great proportion of dietary proteins and/or amino acids consumed in the diet

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Figure 4. Immunoreactivity of protein extracts from pooled total egg samples from eggs produced by layer hens fed a diet with high-oleic peanuts (HOPN) vs. a conventional diet with soybean meal (SBM) at week 10. Protein extracts from pooled egg samples (75 μ g per lane) were run electrophoretically on a 10% polyacrylamide gel. To determine immunereactivity, resolved proteins were transferred to a nitrocellulose membrane and immunoblotted with rabbit IgG anti-peanut agglutinin antibody (1:1000). Biodetection was determined with chromogenic peroxidase substrate-based detection of HRP activity. (A) Immunoblotting results of total egg proteins from hen(s) #1 to 12 (HOPN) and hen(s) 13 to 24 (SBM). Extracted proteins (75 μ g) from peanut flour were utilized as a positive control. (B) Immunoblotting results of total egg proteins from hen(s) #25 to 36 (SBM) and hen(s) 37 to 48 (HOPN). Extracted proteins (75 μ g) from peanut flour were utilized as a positive control.

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TOOMER ET AL.

of livestock are either catabolized during first-pass metabolism of the intestine, liver, intestinal cells, or tissues (Stoll et al., 1998), and therefore are not found directly in the eggs or meat produced. In conclusion, this study helps validate the use of high-oleic peanuts as a valuable feed ingredient for poultry and means to enrich the eggs produced with heart healthy unsaturated fatty acids of health benefit to the health conscious consumer.

Supplementary data are available at Poultry Science online.

ACKNOWLEDGMENTS The authors would gratefully like to acknowledge the following: Prestage Department of Poultry ScienceNCSU, NCSU Feed Mill, Birdsong Peanuts, Dr. Adam Fahrenholz, and Market Quality & Handling Research Unit for their contributions to this study and to Sabrina Whitley-Ferrell, Thien C. Vu and Rasi Fitria for all of their administrative and technical assistance.

REFERENCES Agweek. 2009. Top 10 corn and soybean producing states. Accessed Oct. 2018. http://www.agweek.com/news/3787583top-10-corn-and-soybean-producing-states Al-Batshan, H. A., S. E. Scheideler, B. L. Black, J. D. Garlich, and K. E. Anderson. 1994. Duodenal calcium uptake, femur ash, and eggshell quality decline with age and increase following molt. Poult. Sci. 73:1590–1596. Aletor, V. A., and F. I. Olonimoyo. 1992. Feeding differently processed soya bean Part 1. Effect on performance, protein utilization, relative organ weights, carcass traits and economics of producing broiler-chickens. Nahrung 36:357–363. Alves, R. D., A. P. Moreira, V. S. Macedo, A. R. J. de C´ assia Gon¸calves, R. Mattes, and N. M. Costa. 2014. Regular intake of high-oleic peanuts improves fat oxidation and body composition in overweight/obese men pursuing a energy-restricted diet. Obesity 22:1422–1429. American Peanut Council. 2017. The Peanut Industry-U.S. Peanut Supply. Accessed Oct. 2018. https://www.peanutsusa.com/ about-peanuts/the-peanut-industry3/18-u-s-peanut-supply.html Andersen, P. C., K. Hill, D. W. Gorbet, and B. V. Brodbeck. 1998. Fatty acid and amino acid profiles of selected peanut cultivars and breeding lines. J. Food Compos. Anal. 11:100–111. AOAC 920.39. 1990. Fat (crude) or ether extract in animal feed. Pages 79 in Official Methods of Analysis of the Association of Official Analytical Chemists. K. Helrich, ed. 15th edn. AOAC Inc, Arlington, VA. AOAC 954.02 1990. Fat (crude) or ether extract in pet food. Pages 79–80 in Official Methods of Analysis of the Association of Official Analytical Chemists. 15th edn. AOAC Inc, Arlington, VA. AOAC 958.05 1990. Chemical analysis methods for egg and egg products. Pages 853 in Official Methods of Analysis of the Association of Official Analytical Chemists. 15th edn. AOAC Inc, Arlington, VA. AOAC 990.03. 2006. Protein (crude) in animal feed, combustion method. Pages 30–31 in Official Methods of Analysis of the Association of Official Analytical Chemists. W. H. Horwitz, and G. W. Latimer, Jr. Assistant ed. 18th edn. AOAC International, Gaithersburg, MD. APHIS-Animal and Plant Health Inspection Service-USDA. 2015. Overview of U.S. Livestock, Poultry, and Aquaculture Produc-

Downloaded from https://academic.oup.com/ps/advance-article-abstract/doi/10.3382/ps/pey531/5238158 by guest on 11 January 2019

SUPPLEMENTARY DATA

tion. Accessed Oct. 2018. https://www.aphis.usda.gov/animal health/nahms/downloads/Demographics2015.pdf Armentia, A., F. J. Martin-Gil, C. Pascual, M. Mart´ın-Esteban, A. Callejo, and C. Mart´ınez. 2006. Anisakis simplex allergy after eating chicken meat. J. Investig. Allergol. Clin. Immunol. 16:258– 263. Baeza, E., P. Chartrin, V. Gigaud, S. Tauty, K. Meteau, M. Lessire, and C. Berri. 2013. Effects of dietary enrichment with n-3 fatty acids on the quality of raw and processed breast meat of high and low growth rate chickens. Br. Poult. Sci. 54:190–198. Barbour, J. A., P. R. C. Howe, J. D. Buckley, J. Bryan, and A. M. Coates. 2017. Cerebrovascular and cognitive benefits of high-oleic peanut consumption in healthy overweight middle-aged adults. Nutr. Neurosci. 20:555–562. Chamberlin, K. D., N. A. Barkley, B. L. Tillman, J. W. Dillwith, R. Madden, M. E. Payton, and R. S. Bennet. 2014. A comparison of methods used to determine the oleic/linoleic acid ratio in cultivated peanut. Agric. Sci. 5:227–237. Chung, H. Y., H. M. Rasmussen, and E. J. Johnson. 2004. Lutein bioavailability is higher from lutein-enriched eggs than from supplements and spinach in men. J. Nutr. 134:1887–1893. Cruickshank, E. M. 1934. Studies in fat metabolism in the fowl. Biochem. J. 28:965–977. Costa, E. F., B. R. Miller, G. M. Pesti, R. I. Bakalli, and H. P. Ewing. 2001. Studies on feeding peanut meal as a protein source for broiler chickens. Poult. Sci. 80:306–313. Cilly, V. K., G. N. Lodhi, and J. S. Ichhponani. 1977. Mustard cake a substitute for groundnut cake in egg-type and meat-type chick diets. J. Agric. Sci. 89:759–765. Davis, J. P., J. M. Leek, D. S. Sweigart, P. Dang, C. L. Butts, R. B. Sorensen, C. Y. Chen, and M. C. Lamb. 2017. Measurements of oleic acid among individual kernels harvested from test plots of purified runner and Spanish high oleic seed. Peanut Sci. 44:134– 142. Davis, J. P., K. M. Price, L. L. Dean, D. S. Sweigart, J. M. Cottonaro, and T. H. Sanders. 2016. Peanut oil stability and physical properties across a range of industrially relevant oleic acid/linoleic acid ratios. Peanut Sci. 43:1–11. Donkoh, A., C. C. Atuahene, D. M. Anang, and S. K. Ofori. 1999. Chemical composition of solar-dried blood meal and its effect on performance of broiler chickens. Anim. Feed Sci. Technol. 81:299– 307. Douglas, C. R., and R. H. Harms. 1959. Peanut oil meal as a source of protein in broiler diets. Poult. Sci. 38:786–790. Duraisamy, R. 2011. Egg Size Management-Enhancing Early Egg Size by Proper Management. National Institute of Animal Nutrition and Physiology. Accessed Mar. 2018. https://en.engormix.com/poultry-industry/articles/egg-sizemanagement-t35097.htm. Economic Research Service (ERS)-USDA. 2017. Soybeans and Oil Crops: Trade-Major Foreign Soybean Exporters and Importers. Accessed Oct. 2018. https://www.ers.usda.gov/ topics/crops/soybeans-oil-crops/trade/#foreign El-Hussainy, O., N. E. Asker, A. Ghazalah, and A. Hamid. 1983. Effect of dietary lipid on egg production and fatty acid contents in liver and egg yolk. Ann. Agric. Soc. Moshtohor. 20: 289–310. Fæste, C. K., K. R. Jonscher, M. M. Dooper, W. Egge-Jacobsen, A. Moen, A. Daschner, E. Egaas, and U. Christians. 2014. Characterisation of potential novel allergens in the fish parasite Anisakis simplex. EuPA Open Proteomics 4:140–155. Food and Agriculture Organization of the United Nations (FAO), 2003. Egg Marketing. A guide for the production and sale of eggs. Accessed Mar. 30, 2018. http://teca.fao.org/ sites/default/files/technology files/Egg%20Marketing.pdf Hargis, P. S., M. E. Van Elswyk, and B. M. Hargis. 1991. Dietary modification of yolk lipid with menhaden oil. Poult. Sci. 70:874– 883. Howe, P. R., J. A. Downing, B. F. Grenyer, E. M. Grigonis-Deane, and W. L. Bryden. 2002. Tuna fishmeal as a source of DHA for n-3 PUFA enrichment of pork, chicken, and eggs. Lipids 37:1067– 1076. Hulan, H. W., R. G. Ackman, W. M. N. Ratnayake, and F. G. Proudfoot. 1988. Omega-3 fatty acid levels and performance of broiler

EGGS FROM LAYER HENS FED HIGH-OLEIC PEANUTS

Sell, J. L., S. H. Choo, and P. A. Kondra. 1968. Fatty acid composition of egg yolk and adipose tissue as influenced by dietary fat and strain of hen. Poult. Sci. 47:1296–1302. Shapira, N., P. Weill, and R. Loewenbach. 2008. Egg fortification with n-3 polyunsaturated fatty acids (PUFA): nutritional benefits versus high n-6 PUFA western diets, and consumer acceptance. Isr. Med Assoc. J. 10:262–265. Silversides, F. G., and T. A. Scott. 2001. Effect of storage and layer age on quality of eggs from two lines of hens. Poult. Sci. 80:1240– 1245. Singkham, N., S. Jogloy, T. Kesmala, P. Swatsitang, P. Jaisil, and N. Puppala. 2010. Genotypic variability and genotype by environment interactions in oil and fatty acids in high, intermediate, and low oleic acid peanut genotypes. J. Agric. Food Chem. 58:6257– 6263. Stadelmann, W. J. 1995. Quality identification of shell eggs. Pages 51–64 in Egg Science and Technology. W. J. Stadelmann, and O. J. Cotterill, eds. 4th edn. Food Products Press, Binghamton, NY. Stoll, B., J. Henry, P. J. Reeds, H. Yu, F. Jahoor, and D. G. Burrin. 1998. Catabolism dominates the first-pass intestinal metabolism of dietary essential amino acids in milk protein-fed piglets. J. Nutr. 128:606–614. Suksombat, W., S. Samitayotin, and P. Lounglawan. 2006. Effects of conjugated linoleic acid supplementation in layer diet on fatty acid compositions of egg yolk and layer performances. Poult. Sci. 85:1603–1609. Sweigart, D. S., C. A. Homich, and D. A. Stuart. 2011. Rapid single kernel refractive index test that differentiates regular from high oleic peanuts. Proc. Am. Peanut Res. Educ. Soc. 43:27–28. Tardy, A. L., B. Morio, J. M. Chardigny, and C. Malpuech-Brug`ere. 2011. Ruminant and industrial sources of trans-fat and cardiovascular and diabetic diseases. Nutr. Res. Rev. 24:111–117. United States Department of Agriculture (USDA)Agricultural Marketing Service. 2010. Voluntary Shell Egg Grading Regulations. 7 CFR part 56. Fed. Regis. 7 Part 56:41–64. Van Elswyk, M. E., B. M. Hargis, J. D. Williams, and P. S. Hargis. 1994. Dietary menhaden oil contributes to hepatic lipidosis in laying hens. Poult. Sci. 73:653–662. Venkataraman, L. V., T. Somasekaran, and E. W. Becker. 1994. Replacement value of blue-green-alga (Spirulina platensis) for fishmeal and a vitamin-mineral premix for broiler chicks. Br. Poult. Sci. 35:373–381. Vuilleumier, J. P. 1969. The ‘Roche Yolk Colour Fan’—An Instrument for Measuring Yolk Colour. Poult. Sci. 48:767–779. Waldroup, P. W., and R. H. Harms. 1963. Amino acid supplementation of peanut meal diets for broiler chicks. Poult. Sci. 42:652–657. Wang, Y., H. Sunwoo, G. Cherian, and J. S. Sim. 2000. Fatty acid determination in chicken egg yolk: a comparison of different methods. Poult. Sci. 79:1168–1171. Woods, V. B., and A. M. Fearon. 2009. Dietary sources of unsaturated fatty acids for animals and their transfer into meat, milk and eggs: a review. Livestock Sci. 126:1–20. Woodside, J. V., A. J. McGrath, N. Lyner, and M. C. McKinley. 2015. Carotenoids and health in older people. Maturitas 80:63– 68. Yau, J. C., J. H. Denton, C. A. Bailey, and A. R. Sams. 1991. Customizing the fatty acid content of broiler tissues. Poult. Sci. 70:167–172. Zaheer, K., 2017. Hen egg carotenoids (lutein and zeaxanthin) and nutritional impacts on human health: a review. J. Food. 15:474– 484. Zhang, R. Z., L. Li, S. T. Liu, R. M. Chen, and P. F. Rao. 1999. An improved method of cholesterol determination in egg yolk by HPLC. J. Food Biochem. 23:351–361.

Downloaded from https://academic.oup.com/ps/advance-article-abstract/doi/10.3382/ps/pey531/5238158 by guest on 11 January 2019

chickens fed redfish meal or redfish oil. Can. J. Anim. Sci. 68:533– 547. Haugh, R. R. 1937. The Haugh unit for measuring egg quality. US Egg Poult. Mag. 43:552–555; 572–573. Jiang, Y. H., R. B. McGeachin, and C. A. Bailey. 1994. Alphatocopherol, beta-carotene, and retinol enrichment of chicken eggs. Poult. Sci. 73:1137–1143. Karadas, F., E. Grammenidis, P. F. Surai, T. Acamovic, and N. H. Sparks. 2006. Effects of carotenoids from lucerne, marigold and tomato on egg yolk pigmentation and carotenoid composition. Br. Poult. Sci. 47:561–566. Leeson, S., and L. Caston. 2004. Enrichment of eggs with lutein. Poult. Sci. 83:1709–1712. L´opez-Ferrer, S., M. D. Baucells, A. C. Barroeta, J. Galobart, and M. A. Grashorn. 2001. N-3 enrichment of chicken meat: Use of precursors of long-chain polyunsaturated fatty acids: linseed oil. Poult. Sci. 80:753–761. Marion, J. E., and J. G. Woodroof. 1963. The fatty acid composition of breast, thigh, and skin tissues of chicken broilers as influenced by dietary fats. Poult. Sci. 42:1202–1207. Moreira, A. P., T. F. Teixeira, R. D. Alves, M. C. Peluzio, N. M. Costa, J. Bressan, R. Mattes, and R. C. Alfenas. 2016. Effect of a high-fat meal containing conventional or high-oleic peanuts on post-prandial lipopolysaccharide concentrations in overweight/obese men. J. Hum. Nutr. Diet. 29:95–104. Moreira Alves, R. D., A. P. Boroni Moreira, V. S. Macedo, J. Bressan, A. R. de C´ assia Gon¸calves, R. Mattes, and N. M. Brunoro Costa. 2014. High-oleic peanuts: new perspective to attenuate glucose homeostasis disruption and inflammation related obesity. Obesity 22:1981–1988. National Peanut Board. 2018. Peanut Country, USA. Accessed Oct. 2018. http://www.nationalpeanutboard.org/peanut-info/ peanut-country-usa.htm. Naulia, U., and K. S. Singh. 2002. Effect of substitution of groundnut with soybean meal at varying fish meal and protein levels on performance and egg quality of layer chickens. Asian Australas. J. Anim. Sci. 15:1617–1621. Ngeh-Ngwainbi, J., J. Lin, and A. Chandler. 1997. Determination of total, saturated, unsaturated, and monounsaturated fats in cereal products by acid hydrolysis and capillary gas chromatography: A collaborative study. J. Assoc. Official Anal. Sci. Int. 80:359– 372. Ohtake, Y., and Y. Hoshino. 1976. Influences of dietary fat and oil on the fatty acid distribution in egg yolk lipids of laying hens. Jpn. J. Zootech. Sci. 47:430–440. Olomu, J. M., and S. A. Offiong. 1980. Effect of groundnut meal as the sole supplementary dietary-protein on the performance of broiler-chickens. Trop. Agric. 57:173–181. Olomu, J. M., and S. A. Offiong. 1985. Performance of brown eggtype pullets fed diets based on groundnut meal, with and without supplementation with fish-meal or blood meal. Trop. Agric. 62:289–293. Olson, J. B., N. E. Ward, and E. A. Koutsos. 2008. Lycopene incorporation into egg yolk and effects on laying hen immune function. Poult. Sci. 87:2573–2580. Padhi, M. K., R. N. Chatterjee, S. Haunshi, and U. Rajkumar. 2013. Effect of age on egg quality in chicken. Indian J. Poult. Sci. 48:122–125. Pesti, G. M., R. I. Bakalli, J. P. Driver, K. G. Sterling, L. E. Hall, and E. M. Bell. 2003. Comparison of peanut meal and soybean meal as protein supplements for laying hens. Poult. Sci. 82:1274–1280. Pattee, H. E., E. B. Johns, J. A. Singleton, and T. H. Sanders. 1974. Composition changes of peanut fruit parts during maturation. Peanut Sci. 1:57–62.

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