A comparative study on the effects of hemp seed oil versus four different UFA-rich seed oils in the diets of laying hens on egg production, egg quality and yolk fatty acids

Abstract

This study compared the effects of hemp seed oil versus four different UFA-rich seed oils in the diet of laying hens on egg production, quality, and fatty acid profile of the yolk. Soybean, sunflower, corn, canola, and hemp seed oils were included in the hens' diets in equal proportions. A total of one hundred and twenty 42-week old White Leghorn hens were placed in five diet groups with 8 replicates. The trial lasted 84 days and data were collected on egg production, quality, and fatty acid profile of the yolk. The data showed that none of the incorporated seed oils affected egg production or quality. However, hemp seed oil altered yolk colour values similarly to canola oil by increasing the L* value of the yolk while decreasing the a* value (P < 0.05). Hemp oil increased the PUFA content in the yolk, similar to soybean, corn, and sunflower oil, but unlike the latter, it also enriched the n-3 fatty acids in the yolk (P < 0.05). In conclusion, hemp seed oil can be safely used in the diet of chickens without having negative effects on egg production and quality like other seed oils. Furthermore, hemp seed oil can improve the desirable fatty acid content in the yolk and has the potential to produce n-3 design eggs.

Introduction

Lipids are concentrated sources of energy and are used in poultry nutrition to improve energy stability, palatability, and physical quality of the feed. During the peak phase of laying hens, the intensity of lipid metabolism in the body increases, and the oils in the feed enter the body of the eggs via the ovaries (Gao et al. 2021). The vegetable oils commonly used in feed are rich in PUFAs (polyunsaturated fatty acids) and the lipids in the egg yolk come mainly from the oils in the feed. Therefore, oils play a crucial role in the production performance and egg quality of laying hens (Gao, Zhang, Li, Zheng and Xu 2021). In addition, fats and oils provide varying amounts of linoleic acid, which is an essential nutrient for chickens, and it has been reported that diets should contain at least 1% linoleic acid (Leeson and Summers 2009). It has been reported that high levels of linoleic acid in feed can improve egg production performance (Bohnsack et al. 2002). Another purpose of using lipid sources in egg production is to modulate the fatty acid profile (FA) of the yolk or design functional eggs. Therefore, vegetable lipids, including oils from seeds rich in UFA, n-3, and n-6 (omega 3 and omega 6, respectively) FA are preferred in the diet of hens (Simopoulos 2001). Changes in the fatty acid content of egg yolk can improve the health image of eggs and increase their value as a food supplement (Reda et al. 2020). In particular, eggs enriched with n-3 FA are becoming increasingly popular following the discovery that n-3 is an important micronutrient for normal growth and has been linked to the treatment and prevention of various diseases, including cancer (Leeson and Summers 2009, Reda, El-Kholy, Abd El-Hack, Taha, Othman, Allam and Alagawany 2020, Yang et al. 2018, Yang et al. 2006). Clinical studies have shown that not only n-3, but also the total amount of PUFA, when replacing SFA in the human diet, have beneficial effects on the cardiovascular system (Jakobsen et al. 2009) and the egg has become a common source to increase the dietary intake of PUFA in humans. Soybean oil (SBO) is widely used in commercial egg production due to its high content of polyunsaturated fatty acids (PUFA) and linolenic acid (Dei 2011) and has become almost a standard feed item in poultry diets. Corn oil (CO) and sunflower oil (SFO) were commonly used in chicken diets due to their high content of n-6 fatty acids, especially linoleic acid (Reda, El-Kholy, Abd El-Hack, Taha, Othman, Allam and Alagawany 2020). SO, CO and SFO appear to be suitable lipid sources for producing eggs with enriched PUFAs, but due to economic concerns and competition in human diets, the search for alternative lipids that could replace them in poultry diets continues. Canola oil (CAO) has become a popular lipid source for diets due to its high content of oleic acid, known as n-9 (omega-9), and studies on the effects of CAO in chicken diets on egg yolk fatty acids have increased (Gül et al. 2012). Hemp seed oil (HSO), a by-product of hemp plant (Cannabis sativa) seeds, has the potential to be used in poultry diets due to its excellent micronutrient composition, such as high levels of PUFA and n-3 FAs, and its antioxidant capacity (Oomah et al. 2002). However, due to the legal status of hemp in many countries (Silversides and Lefrancois 2005), HSO cannot be evaluated as a feed additive and further studies are needed to use hemp seed oil as a lipid source in chicken feed. The nutritional properties of the lipid sources used in this study have been well identified and a wide body of knowledge is available in the scientific literature. Plant-based lipids, including some seed oils, have been widely studied in poultry diets and their advantages over animal fat sources have been elaborated. However, comparative studies on lipids of the same origin are needed to determine the crucial usability of newer lipids such as HSO in poultry nutrition. The idea of this study was to distinguish the merits of five different lipid sources of the same origin for chicken nutrition and to present their implications and reveal the usability of hempseed oil for commercial egg production. In this context, the current study investigated five seed-based lipid sources with high UFA content - soybean, corn, sunflower, canola, and hemp seed oils and their effects on egg-laying performance, egg quality, egg yolk colour values, and egg yolk fatty acid composition.

Materials And Methods

The animal experiment was conducted in the Poultry Unit of Prof. Dr. Orhan Düzgüneş Application and Research Farm of the Faculty of Agriculture, Selcuk University.

Animal management procedures

A total of one hundred and twenty 42-week-old White Leghorn chickens were randomly allocated to five treatment groups with dietary oils. Each treatment group contained 24 birds, with 8 replicates and 3 chickens per compartment. The experimental diets were formulated to be isocaloric and isonitrogenic and to meet the nutrient requirements of the National Research Council (NRC 1994) for laying hens. The ingredients and nutrient composition of the experimental feeds are shown in Table 1 and the fatty acid composition of the oil sources in Table 2.

Crude oils from soybean, sunflower, corn, and canola oils were obtained from a commercial local feed factory (Çöğenler Feed Factory, Konya-Turkey). Crude hemp seed oil was extracted by cold pressing (at about 45–50°C) from hemp seeds (Cannabis sativa) using an expeller (Karaerler Machine, model NF 100, Ankara). The metabolic energy value of the seed-based oil sources was ≌ 8800 Kcal/Kg. The experiment lasted 84 days and a 16-h light and the 8-h dark program was applied, feed and water were supplied ad-libitum. The cage dimensions were 40×50×40 cm and corresponded to a total floor area of 2,000 cm2.

Production performance and egg quality

At the beginning and the end of the experiment the hens were weighed, and the weight gain (WG) was determined from the differences. Feed intake (FI) and egg weight (EW) were recorded fortnightly. Egg production (EP) was recorded daily, and egg mass was calculated using the formula egg production × egg weight/period (days). Feed conversion ratio (FCR) was calculated using the ratio of feed intake to egg mass. Egg quality analyses were performed on all eggs collected on the last 2 consecutive days of the three 28-day periods (32 eggs from each group, 160 for each day of analysis). The breaking strength of eggshells was measured with a cantilever system using a force reader instrument (Orka Food Technology Ltd, Ramat Hasharon, Israel). The broken eggshells were rinsed under running water and dried in an oven at 60°C for 12 hours. The eggshells were weighed with a 0.001 g precision balance. The thickness of the eggshells (with membrane) was measured at three points on the eggs (one point on the air cell and two random points on the equator) using a digital micrometre (Mitutoyo Inc., Japan). The colour values of the yolk were determined with a portable colour metre (Minolta Chroma Metre CR 400; Minolta Co., Osaka, Japan) and the parameters L*, a* and b* corresponded to brightness (− 100 / + 100, dark/white), redness (− 100 / + 100, green/red) and yellowing (− 100 / + 100, blue/yellow), respectively.

Determination of fatty acids: On the last three consecutive days of the experiment, a total of 80 eggs were collected from each replicate, 16 from each treatment, and the yolk separated. The yolk oil was extracted using the ethanol/chloroform (1:1) solution method (Kovalcuks and Duma 2014). The fatty acid methyl esters of the yolk and seed oils were prepared according to the method recommended in EU Regulation 2568/91. The oils were weighed into screw-capped glass tubes (0.10 g) and dissolved in 10.0 mL of hexane. Then 100 µL of 2N potassium hydroxide solution in methanol was added to the tubes and shaken vigorously for 30 s. The tubes were then shaken for 5 minutes. The tubes were centrifuged at 2500 × g for 5 min and the upper layer was withdrawn into a small vial and stored at 0°C until the time of analysis (Ayyildiz et al. 2015). The fatty acid composition of the methyl esters was detected by gas chromatography (GC; Shimadzu GC-2010 Plus, 86 Japan) using an FID detector and an HP -88 column (100m x 250 µm x 0.20 µm id). The temperature at the injection block was 250°C and the heating programme of the column oven was set to 2 min at 50°C, 4 min between 50°C − 250°C, and 10 min at 250°C. The carrier gas used was helium with a flow rate of 1.3 mL/min. Fatty acids were detected based on the retention time (min) and area (%) of the identified peaks and classified with fatty acid standards (Supelco® 37 Component FAME Mix, Merck- Germany) and expressed as a percentage.

Statistical analysis

The experiment was designed as a complete randomized model and data were analysed using the one-way ANOVA procedure with Minitab software. Duncan's multiple range test was used to determine the differences among treatments that were found to be significantly different (P < 0.05).

Results

The results of adding seed oils to the diet on egg production, egg quality, and yolk colour values are summarised in Table 3.

Discussion

Previous studies have confirmed that dietary lipid types do not affect egg production and egg weight (Abbasi et al. 2019, Ceylan et al. 2011). It was reported that egg yield, egg weight, and feed intake did not change with the inclusion of different lipid sources (cotton, corn, flax, soybean, olive, sunflower, fish, tallow, and rendering) in the diet (Balevi and Coskun 2000). Omidi et al. (2015) used five different lipids rich in UFAs (fish, olive, grapeseed, canola, and soybean oils) in the diet and found that performance parameters during the laying period did not differ significantly between treatments. It has long been known that vegetable oils in the diet of chickens can increase egg weight, while fats from animal sources have less effect on it (Jensen et al. 1958, Shutze et al. 1958, Treat et al. 1960). To test this hypothesis, Whitehead et al. (1993) reported that corn oil in the diet of hens increases egg weight compared to fish oil, coconut oil, and tallow. Thus, egg size may increase when hens are fed diets high in oil and rich in unsaturated fatty acids (Galea 2011) which affects shell quality. It has been reported that feeding canola oil, which is high in linoleic acid, results in higher egg weight than feeding poultry fat (Bohnsack, Harms, Merkel and Russell 2002). The results of some studies comparing the effects of different vegetable oils in the diet of chickens showed that egg quality parameters were not influenced by the oil sources. For example, Batkowska et al. (2020) showed that the inclusion of 2.5% linseed or soybean oil in the diet for laying hens had no negative effect on egg quality. Similarly, Küçükersan and Küçükersan (2010) found no differences in egg quality when high UFA oils such as sunflower, fish, soybean, and hazelnut oil were included in the feed at a concentration of 3%. A recent study proved that linseed and soybean oil have no effect on quality traits, but increase the weight of chicken eggs (Batkowska et al. 2021). However, some studies reported that olive and sunflower oil in the diet increased the weight of eggs compared to sesame, cottonseed, hazelnut, maize, soybean, and fish oil, but the thickness of the eggshell was not changed by the lipid sources in the diet (Güçlü et al. 2008). Depending on the amount of added oil in the diet, the amounts of other feeds may change so that the fatty acid profile and nutrient composition of the diet may vary, and these changes may affect egg quality parameters. In our study, the experimental diets were formulated with the same amount of feed ingredients (Table 1.) and with the same content of seed-based oils (3.7% of the diet), so that the FA changes in the experimental diets were caused by the added oils and the FAs of the experimental diets in this study clearly had no effect on the egg quality parameters.

The colour of the egg yolk is another effective factor that can influence consumer choice as much as the size of the eggs and can be easily influenced by feed ingredients. For this reason, feed premixes containing carotenoids, which provide the preferred dark yellow yolk colour, are often used in the diets of laying hens. It is reported that unrefined seed oils can contain a detectable amount of carotenoids (Franke et al. 2010), but this amount depends on the carotenoid content of the seed. In addition, the literature mentions that different vegetable oils, depending on their content or mixed oil sources in the diet, can alter the pigmentation of the egg yolk (Faitarone et al. 2016). In this study, a* value, indicating redness, decreased significantly with the intake of HSO and decreased slightly with the intake of CAO, compared to CO, SBO, and SFO. The L* value, indicating the brightness of the yolk, increased in parallel with the decrease in a* value in the CAO and HSO groups. The results of the current study have shown that HSO and CAO can cause a pale-yellow yolk colour, which is not desired by customers, and this fact should be considered when selecting seed-based oils for laying hen diets.

The importance of the FA profile of egg yolk has increased with evidence linking the FA profile of the diet to cardiovascular disease in humans. Since it has been understood that the issue of dietary oil is important in designing a PUFA and Ʃn-3 enriched egg, studies have been presented that used lipids high in UFA or PUFA such as vegetable, nut, and fish oils in the diet of chickens. (Alagawany et al. 2019). The results of our study showed that dietary seed oils altered the DHA, ƩPUFA, and Ʃn-3 content of egg yolk (P < 0.05), and this change was directly related to the FA composition of the oil in the diet. Previous studies have also shown that SO, CO and SFO improved the n-6 content of egg yolk (Güçlü, Uyanık and İşcan 2008, Kang et al. 2006, Oliveira et al. 2010). CAO, which has the highest oleic acid content, increased the ƩMUFA content but decreased the ƩPUFA content of egg yolk. Similarly, CAO was reported to enrich the oleic acid content of the yolk instead of SO in the hens' feed (Gül, Yörük, Aksu, Kaya and Kaynar 2012). In addition, feeding laying hens with diets high in MUFA has been found to increase the oleic acid content in egg yolks (Milinsk et al. 2003). Oleic acid is not essential for humans and to produce ƩPUFA and Ʃn-3 design eggs CAO may not be the best choice. On the other hand, Ʃn-9 has been defined as a beneficial fatty acid for human health (Galán-Arriero et al. 2017) and CAO can be used in the diet of laying hens to improve the MUFA content in the egg yolk. The enrichment effect of dietary HSO on α-linolenic acid, DHA, and PUFA content in egg yolk was impressive. HSO caused the highest content of linolenic acid in the egg yolk. In humans, linolenic acid can be converted to a limited extent to EPA and DHA (Fraeye et al. 2012) and many studies have shown that higher intakes of omega-3 PUFAs, especially EPA and DHA, are linked to lower chronic diseases incidence (Djuricic and Calder 2021). Our previous study also confirmed that HSO in the diet significantly increases ƩPUFA and Ʃn-3 content in quail egg yolk compared to SBO and SFO (Göçmen et al. 2021). In another study, HSO in the diet was reported to significantly increase n-3 PUFA content in egg yolk (Goldberg et al. 2012). A recent meta-analysis confirmed that table eggs from chickens fed whole hemp seeds or their by-products containing HSO in varying amounts had higher levels of long-chain fatty acids ALA (α-linolenic acid), DHA and n-3 (Fabro et al. 2021). We concluded that it is possible to use HSO at 3.7% in the diet of laying hens without differences or negative effects on performance and egg quality parameters. SBO, SFO, and CO helped to increase the ƩPUFA and Ʃn-6 content of the yolk fat and improve the pigmentation of the yolk. Dietary CAO may contribute to the ƩMUFA, and Ʃn-9 content of the egg but may reduce the redness of the yolk. HSO is promising for the design of Ʃn-3-rich eggs, but changes in yolk colour should be considered when using HSO in the diet. On the other hand, HSO is a newer lipid source for animal nutrition, and further research on its use in laying poultry nutrition is needed.

Declarations

Statement of Animal Rights: In this study, the animal experiment was conducted according to the guidelines of the local ethics committee of Selcuk University, which were prepared following the "Directive 2010/63/EU is the European Union (EU) legislation". All procedures in this study complied with the ethical principles of animal welfare.

Funding Statement: The authors declare that no funds, grants, or other support were received during the preparation of this manuscript

Competing Interests: The authors declare they have no financial interests.

Author Contributions: All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Gülşah Kanbur, Rabia Göçmen and Yusuf Cufadar. The first draft of the manuscript was written by Gülşah Kanbur and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

Conflict of Interest: The authors declare that they have no conflict of interest.

Data Availability Statement: The data that support the findings of this study are available from the corresponding author, [Gülşah Kanbur], upon reasonable request.

References

1. Abbasi, S. et al., 2019. Productive performance, blood biochemical variables and immune responses of laying hens fed diets containing various fat sources, South African Journal of Animal Science, 49, 675–683
2. Alagawany, M. et al., 2019. Omega-3 and omega-6 fatty acids in poultry nutrition: effect on production performance and health, Animals, 9, 573
3. Ayyildiz, H.F. et al., 2015. Evaluation of fatty acid composition, tocols profile, and oxidative stability of some fully refined edible oils, International Journal of Food Properties, 18, 2064–2076
4. Balevi, T., and Coskun, B., 2000. Effects of some dietary oils on performance and fatty acid composition of eggs in layers, Revue de Médecine Vétérinaire, 151, 847–854
5. Batkowska, J. et al., 2020. Fatty acids profile, cholesterol level and quality of table eggs from hens fed with the addition of linseed and soybean oil, Food chemistry, 334, 127612
6. Batkowska, J. et al., 2021. Fatty acids profile, cholesterol level and quality of table eggs from hens fed with the addition of linseed and soybean oil, Food chemistry, 334, 127612
7. Bohnsack, C. et al., 2002. Performance of commercial layers when fed diets with four levels of corn oil or poultry fat, Journal of Applied Poultry Research, 11, 68–76
8. Ceylan, N. et al., 2011. Influence of different dietary oil sources on performance and fatty acid profile of egg yolk in laying hens, Journal of Animal and Feed Sciences, 20, 71–83
9. Dei, H., 2011. Soybean as a Feed Ingredient for Livestock and Poultry, Recent Trends for Enhancing the Diversity and Quality of Soybean Products. 2011, (IntechOpen, India), 536
10. Djuricic, I., and Calder, P.C., 2021. Beneficial outcomes of omega-6 and omega-3 polyunsaturated fatty acids on human health: An update for 2021, Nutrients, 13, 2421
11. Fabro, C., Romanzin, A., and Spanghero, M., 2021. Fatty acid profile of table eggs from laying hens fed hempseed products: A meta-analysis, Livestock Science, 254, 104748
12. Faitarone, A.B.G. et al., 2016. Yolk color and lipid oxidation of the eggs of commercial white layers fed diets supplemented with vegetable oils, Brazilian Journal of Poultry Science, 18, 9–16
13. Fraeye, I. et al., 2012. Dietary enrichment of eggs with omega-3 fatty acids: A review, Food Research International, 48, 961–969
14. Franke, S. et al., 2010. Analysis of carotenoids and vitamin E in selected oilseeds, press cakes and oils, European Journal of Lipid Science and Technology, 112, 1122–1129
15. Galán-Arriero, I. et al., 2017. The role of Omega-3 and Omega-9 fatty acids for the treatment of neuropathic pain after neurotrauma, Biochimica et Biophysica Acta (BBA)-Biomembranes, 1859, 1629–1635
16. Galea, F., 2011. Nutrition and food management and their influence on egg quality. Proccedings XLVIII Simposio Científico de Avicultura, Santiago de Compostela, 2011,
17. Gao, Z. et al., 2021. Effect of Oils in Feed on the Production Performance and Egg Quality of Laying Hens, Animals, 11, 3482
18. Goldberg, E.M. et al., 2012. Fatty acid profile and sensory characteristics of table eggs from laying hens fed hempseed and hempseed oil, Journal of food science, 77, S153-S160
19. Göçmen, R., Kanbur, G., and Cufadar, Y., 2021. The Use of Different Fat Sources on Performance, Egg Quality and Egg Yolk Fatty Acids Content in Laying Quails, Turkish Journal of Agriculture-Food Science and Technology, 9, 1413–1418
20. Güçlü, B., Uyanık, F., and İşcan, K., 2008. Effects of dietary oil sources on egg quality, fatty acid composition of eggs and blood lipids in laying quail, South African Journal of Animal Science, 38, 91–100
21. Gül, M. et al., 2012. The effect of different levels of canola oil on performance, egg shell quality and fatty acid composition of laying hens, International Journal of Poultry Science, 11, 769
22. Jakobsen, M.U. et al., 2009. Major types of dietary fat and risk of coronary heart disease: a pooled analysis of 11 cohort studies, The American Journal of Clinical Nutrition, 89, 1425–1432
23. Jensen, L.S. et al., 1958. Evidence for an unidentified factor necessary for maximum egg weight in chickens, The Journal of Nutrition, 65, 219–233
24. Kang, H.-K. et al., 2006. Effects of dietary fats on the cholesterol content and fatty acid composition of egg yolk, Food Science of Animal Resources, 26, 517–524
25. Kovalcuks, A., and Duma, M., 2014. Solvent extraction of egg oil from liquid egg yolk. 9th Baltic Conference on Food Science and Technology–Food for Consumer Well-Being: Foodbalt, 2014, 253–256
26. Küçükersan, K., and Küçükersan, S., 2010. Influence of different dietary oil sources on performance and cholesterol content of egg yolk in laying hens, J. Biol. Environ. Sci., 12, 117–122
27. Leeson, S., and Summers, J.D., 2009. Commercial poultry nutrition, (Nottingham University Press)
28. Milinsk, M. et al., 2003. Fatty acid profile of egg yolk lipids from hens fed diets rich in n-3 fatty acids, Food chemistry, 83, 287–292
29. NRC, 1994. Nutrient requirements of poultry. 1994, (National Academies Press, USA)
30. Oliveira, D. et al., 2010. Effects of lipid sources in the diet of laying hens on the fatty acid profiles of egg yolks, Poultry Science, 89, 2484–2490
31. Omidi, M., Rahimi, S., and Torshizi, M.A.K., 2015. Modification of egg yolk fatty acids profile by using different oil sources. Veterinary Research Forum, 2015, (Faculty of Veterinary Medicine, Urmia University, Urmia, Iran, 137
32. Oomah, B.D. et al., 2002. Characteristics of hemp (Cannabis sativa L.) seed oil, Food chemistry, 76, 33–43
33. Reda, F. et al., 2020. Does the use of different oil sources in quail diets impact their productive and reproductive performance, egg quality, and blood constituents?, Poultry Science, 99, 3511–3518
34. Shutze, J., Jensen, L., and McGinnis, J., 1958. Effect of different dietary lipids on egg size, (Poultry science assocation)
35. Silversides, F., and Lefrancois, M., 2005. The effect of feeding hemp seed meal to laying hens, British poultry science, 46, 231–235
36. Simopoulos, A.P., 2001. N – 3 fatty acids and human health: Defining strategies for public policy, Lipids, 36, 83–89
37. Treat, C. et al., 1960. Effect of animal fat and mixtures of animal and vegetable fats containing varying amounts of free fatty acids on performance of cage layers, Poultry Science, 39, 1550–1555
38. Whitehead, C., Bowman, A., and Griffin, H., 1993. Regulation of plasma oestrogen by dietary fats in the laying hen: relationships with egg weight, British poultry science, 34, 999–1010
39. Yang, H. et al., 2018. Effects of three positively buoyant dietary supplements on the buoyancy of feces, growth and intestinal health of Tilapia, Oreochromis niloticus× O. aureus, Aquaculture and fisheries, 3, 72–78
40. Yang, X. et al., 2006. Fatty acids and coccidiosis: effects of dietary supplementation with different oils on coccidiosis in chickens, Avian Pathology, 35, 373–378