The fatty acid profile in the yolk and yolk sac from incubated goose eggs depends on the breeder age and laying period

ABSTRACT

1. The study analysed the content of fatty acids in the lipids of the yolk and yolk sac of hatching eggs obtained from geese in four reproductive flocks and three laying periods at different incubation dates.

2. A total of 1080 hatching eggs were used in the study (90 eggs from each age group in three laying periods). The geese were kept on one farm under the same conditions.

3. On days 0, 16, 22, and 28 of incubation, the yolk/yolk sac was sampled. Saturated and unsaturated (mono- and poly-) fatty acids were determined, including myristic acid, palmitic acid, palmitoleic acid, margaric acid, stearic acid, oleic acid, linoleic acid, α-linolenic acid, behenic acid, eicosapentaenoic acid. The ratio of unsaturated to saturated fatty acids was calculated.

4. Embryo fatty acid utilisation in eggs from different age groups of geese was similar. The fatty acid profile depended mostly on the laying period. The different proportions of fatty acids in the yolk during incubation indicated changes in the activity of various enzymatic processes in the membrane of the yolk sac of embryos from the beginning and at the end of the laying period.

5. When analysing the interactions between the age of the parent flock and the laying period, the most significant effect on the fatty acid composition was found in fresh eggs. On d 16 of lay the myristic, stearic, linoleic, and behenic acids and PUFA; on d 22 of lay linolenic acid, and on day 28th palmitoleic and margaric acids were involved in this interaction.

KEYWORDS:

• Goose
• hatching eggs
• lipid
• fatty acid
• yolk
• reproduction
• embryo
• incubation

Introduction

An egg must contain appropriate nutrients as a being environment for the development of avian embryos (Givisiez et al. 2020). The genotype of the birds determines the chemical composition of the egg. This might be altered by breeding or artificially modifying dietary components. The level of individual nutrients depends on the intensity of laying and the age of birds (Hamidu et al. 2007; Yadgary et al. 2010). Such variation of the components could be because the size of the egg (and hence its components) which varies significantly with age (Salamon and Kent 2013; Salamon 2020) as well as within the laying season (Mróz and Lepek 2003; Salamon and Kent 2013).

Further, with this variation in egg size, there is a variation in egg components (Adamski et al. 2016). The nutrients of the yolk are absorbed and metabolised into other compounds (lipoproteins, carbohydrates, amino acids and fatty acids). The yolk is a crucial energy source during embryonic development and the only source of lipids (Cherian 2015; Guo et al. 2021; Ding et al. 2022). The oxygen supply to the embryos depends on the lipids contained in the egg, which is related to the oxidation process (Hamidu et al. 2007; Nasri et al. 2020). According to some authors, higher amounts of lipids in the egg increased the oxygen consumption of the embryos (Hamidu et al. 2007; Ovideo-Rondon et al. 2008). The embryos in larger eggs may be exposed to hypoxia, resulting in hatching abnormalities (Ovideo-Rondon et al. 2008). Egg yolk lipids are rich in fatty acids (FA). In fertilised egg yolks, polyunsaturated FA are higher in the early stages of development and support the development of neurons and the embryo’s brain. As they develop, FAs are absorbed by the embryo (Cherian 2015; Liu et al. 2021).

Changes in the FA composition of the egg yolk depend on the age and laying period in hens (Lešić et al. 2017; Baykalir, Simsek, and Yilmaz 2020). This is related to the absorption and utilisation of FA by developing embryos (Yadgary et al. 2010; Nangsuay et al. 2011). Other trials (Okruszek et al. 2006; Yilmaz-Dikmen and Sahan 2009; Hu 2013) have studied changes in fresh chicken and duck eggs (yolk) and during incubation (yolk sac). Hu (2013) showed that the proportion of oleic and linolenic acid was higher in the yolks of fresh eggs obtained from broiler breeders at 51 weeks old compared to breeders at 32 weeks of age. At the same time, a lower proportion of palmitic and palmitoleic acid was reported. These changes were further demonstrated during egg incubation (on d 11, 14, 17 and 20). On d 14 of incubation, the yolks in eggs from younger hens were characterised by a higher concentration of palmitic, stearic, and linoleic acids. Yilmaz-Dikmen and Sahan (2009) showed that the hatchability of broiler breeder eggs depended on the FA content. They found a negative correlation between the content of myristic acid and linoleic acid in the yolk and the age of the layers. Their conclusions indicated that these changes might be correlated with embryonic death, and, additionally, Grochowska et al. (2019) discussed that FA content in egg yolks might affect hatchability, as this is related to the embryos’ effective mobilisation and assimilation of yolk sac lipids. However, data for geese are scarce.

The following study was conducted to record changes in the yolk/yolk sac of hatching eggs obtained from reproductive geese. The study analysed FA in the yolk of fresh eggs obtained from geese from parent flocks at different ages and laying periods, as well as in the yolk sac of hatching eggs during incubation.

Material and methods

Ethical approval

The research was carried out with the consent of the Local Ethical Committee in Bydgoszcz No. 30/2015, Bydgoszcz University of Science and Technology, Poland.

Egg collection and incubation

Hatching eggs were obtained during each laying season from White Kołuda® geese kept in parent flocks. They were divided, depending on the age of the parent flock, into four experimental groups: I (1-year-old), II (2-year-old), III (3-year-old), and IV (4-year-old). Each flock contained ganders, which were the same age as the female geese, to take into account any changes in fertility. The flocks consisted of 1,500 birds and the sex ratio was 4/5:1 (female: male) and the laying period was from January to June.

Eggs were collected during three laying periods – at the beginning (4^th week of laying), at peak (13^th week of laying, April) and the end (20^th week of laying). The experimental material was collected from each age group simultaneously three days before incubation. After collecting, the eggs were disinfected with 0.5% Virkon solution. Virkon was diluted with warm water (room temperature) and the solution sprinkled onto the eggs.

In total, 90 goose eggs from each experimental group (1080 in total) were obtained for incubation during the three laying periods. The eggs were stored at 10°C and 70–75% relative humidity. Before incubation, the eggs were disinfected again, as above. The material was marked with a group symbol and a sequential number. Incubation was carried out in a single-stage incubator (Jarson, Gostyń, Poland). The air temperature in the incubator was 37.7°C, and relative humidity was 55%, while in the hatcher, it was 37.4°C with 75% humidity.

On the second day of incubation, cooling was started for 20 min twice a day. This treatment was repeated until the eggs were transferred to the hatcher. From d 9, the egg airing and sprinkling with water for cooling outside the incubator in a well-ventilated hall were carried out. From d 16 of incubation, eggs were manually rotated 180° along the long axis (Salamon and Kent 2016). Rotation and airing were performed daily. On d 27, the eggs were set into the hatcher. During the last 3–4 days of incubation, the hatcher was ventilated for about 20 min daily. Incubation lasted 30–31 d.

Fatty acid analyses

Samples were taken from the separated whole yolks (without vitelline membrane) into polypropylene containers. The contents from the yolk sac were sampled on incubation d 16, 22 and 28. Samples were frozen in air under free convection at −21°Cand then subjected to freeze-drying. Lyophilisation of the frozen samples was carried out in an Alpha 1–4 LD plus lyophiliser (Martin Christ Gefriertrocknungsanlagen GmbH, Osterode am Harz, Germany). Fat extraction was performed using a chloroform-methanol (2: 1) v/v extraction mixture, and FA methyl esters were prepared following the PN-EN ISO 12 966–2:2011 method (Polish Committee for Standardisation, Warsaw, Poland). Using a type 7890B gas chromatograph with a MSD5977 A detector and autosampler (Agilent Technologies, Inc., Santa Clara, CA, U.S.A), FA methyl ester analysis was performed (Kowalska et al. 2021). The following fatty acids were determined: myristic acid (C14:0), palmitic acid (C16:0), palmitoleic acid (C16:1), margaric acid (C17:0), stearic acid (C18:0), oleic acid (C18:1n9), linoleic acid (C18:2n6, LA), α-linolenic acid (C18:3n3, ALA), behenic acid (C22:0), eicosapentaenoic acid (C20:5n3, EPA). For data analysis, the acids were divided into saturated fatty acids (SFA: myristic, palmitic, stearic, and behenic acids), unsaturated fatty acids (UFA: palmitoleic, LA, ALA, and EPA acids), monounsaturated fatty acids (MUFA: palmitoleic) and polyunsaturated fatty acids (PUFA: LA, ALA, and EPA acids). The ratio of unsaturated fatty acids to saturated fatty acids (UFA/SFA) was calculated.

Statistical analysis

The data were statistically processed using Statistica 12.5 PL (Statsoft, TIBCO, Kraków, Poland). The mean values of all examined features and their standard deviations (±SD) were calculated. A two-way analysis of variance was used in the calculations. The Kolmogorov-Smirnov test verified the normality of the distribution, and the post-hoc test was used to identify statistically significant differences, assuming significance at P < 0.05. The interactions between the factors (parent flock age and laying season) were verified by two-way ANOVA, and P values were obtained from the function one-dimensional results for each dependent variable (P-value < 0.05). For the interactions, 12 groups were compared, based on parent age and incubation time.

Results

The FAs content in fresh goose egg yolks is presented in Tables 1 and 2. Considering the age of the geese, the highest content of margaric acid, linoleic acid (LA), α-linolenic acid (ALA), and eicosapentaenoic acid (EPA) was found in the egg yolks of the youngest geese. The FA proportion decreased with each successive season (bird age). In the fourth season, margaric acid content decreased by 10%, LA by 23%, α-ALA by 25% and EPA by 39%. The higher UFA content in the egg yolks of young geese was related to higher PUFA content. The proportion in the yolk lipids was 4.61% in the one-year-old and 3.22% in 4-year-old geese (P < 0.05). Unlike the other periods, eggs from the beginning of laying were characterised by the lowest content of myristic, palmitic, margaric and stearic acids. The highest content was found at the peak of lay.

Table 1. The fatty acid content in fresh goose egg yolk lipids.

Factor	Group^1	Fatty acids^2
		C14:0	C16:0	C16:1	C17:0	C18:0	C18:1n9	C18:2n6	C18:3n3	C22:0	C20:5n3
Parent flock age	I	0.40 ± 0.19	43.27 ± 5.66	1.66 ± 0.34	0.10^ab ± 0.04	9.90 ± 4.22	37.22 ± 9.47	4.18^a ± 0.84	0.20^a ± 0.08	2.84 ± 0.98	0.23^a±0.12
	II	0.46 ± 0.19	45.54 ± 3.96	1.71 ± 0.23	0.11^a ± 0.05	10.80 ± 3.06	35.15 ± 7.25	3.68^ab ± 0.80	0.19^ab ± 0.06	2.19 ± 0.48	0.17^ab ± 0.06
	III	0.48 ± 0.22	44.95 ± 4.82	1.69 ± 0.41	0.09^b ± 0.03	10.56 ± 2.22	36.35 ± 6.82	3.38^b ± 0.60	0.16^ab ± 0.05	2.16 ± 0.80	0.16^b ± 0.09
	IV	0.43 ± 0.22	43.89 ± 4.80	1.52 ± 0.38	0.09^b ± 0.03	10.77 ± 3.27	37.35 ± 7.71	3.22^b ± 0.53	0.15^b ± 0.04	2.44 ± 1.07	0.14^b ± 0.03
	P-value	0.206	0.133	0.355	0.022	0.177	0.345	0.001	0.013	0.053	0.009
Laying period	Beginning (B)	0.22^c ± 0.04	39.68^b ± 2.15	1.51 ± 0.30	0.06^c ± 0.02	7.37^c ± 0.94	44.77^a ± 2.63	3.17^a ± 0.64	0.18 ± 0.05	2.86^a ± 0.84	0.18^ab ± 0.07
	Peak (P)	0.61^a ± 0.12	47.79^a ± 1.46	1.74 ± 0.33	0.13^a ± 0.03	13.32^a ± 1.63	29.67^c ± 3.19	4.05^b ± 0.68	0.18 ± 0.05	2.30^ab ± 0.90	0.20^a ± 0.11
	End (E)	0.50^b ± 0.15	45.98^a ± 5.18	1.69 ± 0.37	0.10^b ± 0.03	10.97^b ± 7.67	34.71^b ± 7.67	3.67^b ± 0.77	0.18 ± 0.08	2.05^b ± 0.76	0.14^b ± 0.06
	P-value	< 0.001	< 0.001	0.089	< 0.001	< 0.001	< 0.001	< 0.001	0.878	0.006	0.029
Interaction^3	I × B	0.23 ± 0.04	40.76^BC ± 1.57	1.44 ± 0.21	0.09^CDE ± 0.02	7.56^C ± 0.90	42.70^A ± 1.01	3.77 ± 0.53	0.21^AB ± 0.04	3.09^AB ± 0.44	0.17^B ± 0.04
	II × B	0.25 ± 0.02	40.71^BC ± 1.46	1.78 ± 0.22	0.06^DE ± 0.01	7.10^C ± 0.32	44.15^A ± 2.48	3.14 ± 0.73	0.20^AB ± 0.07	2.45^AB ± 0.58	0.16^B ± 0.05
	III × B	0.21 ± 0.04	39.03^C ± 2.59	1.41 ± 0.32	0.05^DE ± 0.01	7.84^C ± 1.06	45.00^A ± 3.30	2.96 ± 0.53	0.16^B ± 0.07	3.04^AB ± 0.85	0.24^AB ± 0.11
	IV × B	0.18 ± 0.05	38.20^C ± 2.15	1.42 ± 0.35	0.05^DE ± 0.01	6.97^C ± 1.23	47.21^A ± 1.20	2.81 ± 0.43	0.15^B ± 0.03	2.85^AB ± 1.34	0.15^B ± 0.04
	I × P	0.55 ± 0.11	48.54^A ± 1.25	1.61 ± 0.30	0.14^AB ± 0.02	14.97^A ± 1.36	25.75^B ± 2.96	4.51 ± 0.73	0.13^B ± 0.04	3.46^A ± 0.99	0.34^A ± 0.13
	II × P	0.61 ± 0.16	47.69^A ± 1.81	1.68 ± 0.35	0.17^A ± 0.04	13.72^AB ± 1.20	29.28^B ± 2.43	4.24 ± 0.99	0.21^AB ± 0.06	2.19^AB ± 0.28	0.22^AB ± 0.08
	III × P	0.65 ± 0.12	47.66^A ± 1.35	1.91 ± 0.40	0.12^ABC ± 0.02	12.35^AB ± 1.07	31.46^B ± 1.11	3.78 ± 0.42	0.19^AB ± 0.03	1.75^B ± 0.34	0.13^B ± 0.04
	IV × P	0.61 ± 0.08	47.28^A ± 1.62	1.74 ± 0.30	0.11^BCD ± 0.01	12.34^AB ± 1.39	32.12^B ± 0.67	3.70 ± 0.32	0.18^AB ± 0.01	1.79^B ± 0.30	0.14^B ± 0.02
	I × E	0.42 ± 0.23	40.52^B ± 7.49	1.94 ± 0.32	0.08^CDE ± 0.05	7.17^C ± 3.39	43.22^A ± 11.27	4.24 ± 1.13	0.28^A ± 0.09	1.99^AB ± 0.84	0.18^B ± 0.08
	II × E	0.56 ± 0.04	48.64^A ± 0.74	1.66 ± 0.13	0.11^BCD ± 0.02	12.16^AB ± 1.15	30.86^B ± 1.28	3.77 ± 0.34	0.16^B ± 0.03	1.93^AB ± 0.45	0.15^B ± 0.05
	III × E	0.57 ± 0.09	48.16^A ± 2.62	1.74 ± 0.37	0.10^BCDE ± 0.02	11.51^B ± 0.72	32.57^B ± 3.04	3.41 ± 0.61	0.14^B ± 0.03	1.69^B ± 0.16	0.11^B ± 0.07
	IV × E	0.51 ± 0.16	46.78^AB ± 2.42	1.37 ± 0.47	0.10^BCDE ± 0.03	13.55^AB ± 1.78	31.57^B ± 1.59	3.15 ± 0.39	0.11^B ± 0.03	2.75^AB ± 1.15	0.13^B ± 0.02
	P-value	0.594	0.003	0.101	0.025	< 0.001	< 0.001	0.995	< 0.001	0.022	0.005

Values are presented as mean ± standard deviation (SD); ^a,b mean values with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ± values mean standard deviation; ¹1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese;²fatty acids: myristic acid (C14:0), palmitic acid (C16:0), palmitoleic acid (C16:1), margaric acid (C17:0), stearic acid (C18:0), oleic acid (C18:1n9), linoleic acid (C18:2n6, LA), α-linolenic acid (C18:3n3, ALA), behenic acid (C22:0), eicosapentaenoic acid (C20:5n3, EPA); ³the interaction (^{A, B}) between two factors: parent flock age and laying period; P < 0.05.

Table 2. Fatty acid groups content in fresh goose egg yolk lipids.

Factor	Group^1	Fatty acids^2
		SFA	UFA	UFA/SFA	MUFA	PUFA
Parent flock age	I	56.52 ± 10.06	43.49 ± 10.07	0.82 ± 0.34	38.88 ± 10.51	4.61^a ± 0.85
	II	59.09 ± 6.73	40.90 ± 6.73	0.71 ± 0.21	36.86 ± 7.28	4.04^ab ± 0.87
	III	58.24 ± 6.20	41.73 ± 6.18	0.73 ± 0.20	38.03 ± 6.57	3.70^b ± 0.62
	IV	57.62 ± 7.32	42.38 ± 7.31	0.76 ± 0.24	38.87 ± 7.63	3.52^b ± 0.54
	P-value	0.178	0.174	0.102	0.375	< 0.001
Laying period	Beginning (B)	50.18^c ± 2.04	49.80^a ± 2.08	0.99^a ± 0.08	46.27^a ± 2.57	3.53^b ± 0.69
	Peak (P)	64.15^a ± 2.74	35.83^c ± 2.74	0.56^c ± 0.06	31.41^c ± 3.27	4.42^a ± 0.75
	End (E)	59.61^b ± 7.77	40.39^b ± 7.77	0.71^b ± 0.29	36.40^b ± 7.73	3.98^ab ± 0.83
	P-value	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001
Interaction^3	I × B	51.73^B ± 1.35	48.29^A ± 1.36	0.93^A ± 0.05	44.14^A ± 0.99	4.15 ± 0.57
	II × B	50.57^B ± 1.62	49.43^A ± 1.62	0.98^A ± 0.06	45.93^A ± 2.27	3.50 ± 0.81
	III × B	50.16^B ± 2.64	49.77^A ± 2.75	1.00^A ± 0.11	46.41^A ± 3.29	3.36 ± 0.61
	IV × B	48.25^B ± 0.84	51.75^A ± 0.84	1.07^A ± 0.04	48.63^A ± 1.26	3.12 ± 0.45
	I × P	67.65^A ± 2.78	32.34^B ± 2.78	0.48^B ± 0.06	27.36^B ± 3.14	4.98 ± 0.74
	II × P	64.37^A ± 1.41	35.63^B ± 1.42	0.55^B ± 0.03	30.96^B ± 2.37	4.67 ± 1.10
	III × P	62.53^A ± 0.82	37.47^B ± 0.82	0.60^B ± 0.02	33.38^B ± 0.96	4.10 ± 0.47
	IV × P	62.13^A ± 0.48	37.87^B ± 0.48	0.61^B ± 0.01	33.86^B ± 0.51	4.02 ± 0.31
	I × E	50.17^B ± 10.55	49.86^A ± 10.52	1.06^A ± 0.42	45.16^A ± 11.24	4.70 ± 1.12
	II × E	63.39^A ± 1.22	36.61^B ± 1.22	0.58^B ± 0.03	32.52^B ± 1.32	4.08 ± 0.37
	III × E	62.03^A ± 2.12	37.97^B ± 2.13	0.61^B ± 0.06	34.31^B ± 2.74	3.66 ± 0.65
	IV × E	63.68^A ± 1.27	36.32^B ± 1.26	0.57^B ± 0.03	32.93^B ± 1.49	3.39 ± 0.44
	P-value	< 0.001	< 0.001	< 0.001	< 0.001	0.988

Values are presented as mean ± standard deviation (SD); ^a,bvalues with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ± values mean standard deviation; ¹I, 1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese;²fatty acids: SFA, saturated fatty acids; UFA, unsaturated fatty acids; UFA/SFA, unsaturated to saturated fatty acids ratio; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids; ³the interaction ^{(A, B)} between two factors: parent flock age and laying period, P < 0.05.

Content decreased in the subsequent weeks of production in the yolk lipid. At the beginning of laying the behenic acid proportion was the highest (2.86%) and was lowest (2.05%) at the end of production. The highest proportion of oleic and LA acids was recorded in egg yolks at the beginning lay (P < 0.05). The highest EPA content was found at the beginning and at peak of lay, and the lowest at the end of the laying period.

The proportion of SFA, UFA, MUFA, and PUFA in goose egg yolk fat depended significantly on the laying period. As production continued, the proportion of SFA showed an upward trend. However, the content of UFA significantly decreased at the end of the laying period (approximately 19% compared to the initial value). The proportion of MUFA was in the following order: peak <end <beginning of lay. The highest proportion of PUFA was found at the peak of laying.

In fresh eggs, the interaction of both factors was observed in the proportion of palmitic acid (the highest seen in all groups at the peak, and II and IV at the end of laying), margaric (the highest – I and II groups at the beginning of laying), stearic (I group at the peak of lay), oleic (group I-V at the beginning and the group I at the end of lay), as well as ALA (group I at the end of lay), behenic and EPA (group I at the beginning of lay) (P < 0.05). For all FA, except PUFA, the interaction of factors was significant in fresh eggs (P < 0.05). The highest proportion of SFA was found in the I, II, III, and IV groups at the peak and II-IV at the end of lay. For UFA, UFA/SFA, and MUFA – in groups I – IV at the beginning and in group I at the end of lay.

On d 16 of incubation, a significant effect of the age of the females on the content of stearic, oleic, behenic, and EPA acids was found (Table 3). The highest proportion of stearic and behenic acid (11.41 and 2.99%, respectively) was found in the yolk sac lipids from 1-year-old laying goose eggs and lowest in 4-year-old laying goose eggs (10.05% and 2.25% respectively). The highest oleic acid content was recorded in group IV (36.62%) and the lowest in groups I (34.73%) and II (34.33%) (P ≤ 0.05).

Table 3. The fatty acid content in lipids of the yolk sac of goose eggs on day 16 of incubation.

Factor	Group^1	Fatty acids^2
		C14:0	C16:0	C16:1	C17:0	C18:0	C18:1n9	C18:2n6	C18:3n3	C22:0	C20:5n3
Parent flock age	I	0.41 ± 0.19	45.88 ± 7.32	1.36 ± 0.17	0.12 ± 0.05	11.41^a ± 3.26	34.73^b ± 7.65	3.91 ± 0.67	0.18 ± 0.05	2.99^a ± 1.01	0.21^ab ± 0.06
	II	0.40 ± 0.19	46.08 ± 4.98	1.57 ± 0.35	0.11 ± 0.05	10.40^bc ± 3.59	34.33^b ± 9.41	3.81 ± 0.94	0.18 ± 0.07	2.87^a ± 0.95	0.25^a ± 0.10
	III	0.40 ± 0.20	44.72 ± 5.41	1.54 ± 0.31	0.11 ± 0.09	11.16^ab ± 3.71	35.70^ab ± 8.96	3.56 ± 0.57	0.17 ± 0.05	2.49^ab ± 0.53	0.18^b ± 0.06
	IV	0.40 ± 0.18	45.06 ± 6.12	1.58 ± 0.26	0.08 ± 0.04	10.05^c ± 3.65	36.62^a ± 9.67	3.62 ± 0.72	0.17 ± 0.04	2.25^b ± 0.61	0.18^b ± 0.06
	P-value	0.966	0.393	0.107	0.110	< 0.001	0.030	0.340	0.988	0.006	0.049
Laying period	Beginning (B)	0.18^b ± 0.02	38.13^b±1.52	1.61 ± 0.27	0.05^b ± 0.02	6.28^c ± 0.73	46.84^a ± 2.24	3.35^b ± 0.62	0.21^a ± 0.05	3.17^a ± 0.80	0.20 ± 0.07
	Peak (P)	0.49^a ± 0.11	49.12^a±2.01	1.44 ± 0.22	0.12^a ± 0.04	12.07^b ± 1.40	30.31^b ± 3.12	3.87^a ± 0.56	0.17^ab ± 0.03	2.22^b ± 0.54	0.20 ± 0.07
	End (E)	0.54^a ± 0.11	49.22^a±4.03	1.47 ± 0.34	0.15^a ± 0.07	14.11^a ± 0.99	28.59^b ± 2.94	3.96^a ± 0.86	0.14^b ± 0.05	2.54^b ± 0.86	0.21 ± 0.10
	P-value	< 0.001	< 0.001	0.128	< 0.001	< 0.001	< 0.001	0.004	< 0.001	< 0.001	0.683
Interaction^3	I × B	0.17^C ± 0.02	37.71 ± 0.93	1.32 ± 0.18	0.06 ± 0.01	7.11^E ± 0.75	45.01 ± 1.73	3.98^AB ± 0.63	0.21 ± 0.04	4.22^A ± 0.70	0.21 ± 0.08
	II × B	0.18^C ± 0.02	39.95 ± 1.20	1.66 ± 0.16	0.06 ± 0.02	5.92^E ± 0.31	45.75 ± 2.36	3.17^AB ± 0.63	0.23 ± 0.06	2.84^BC ± 0.66	0.25 ± 0.08
	III × B	0.17^C ± 0.03	37.75 ± 1.34	1.60 ± 0.29	0.05 ± 0.02	6.37^E ± 0.32	47.47 ± 1.33	3.26^AB ± 0.42	0.21 ± 0.06	2.92^ABC ± 0.31	0.19 ± 0.03
	IV × B	0.19^C ± 0.02	37.09 ± 0.96	1.84 ± 0.19	0.04 ± 0.01	5.71^E ± 0.59	49.11 ± 0.95	2.98^B ± 0.40	0.18 ± 0.04	2.71^BC ± 0.42	0.17 ± 0.05
	I × P	0.58^AB ± 0.11	48.55 ± 0.43	1.46 ± 0.18	0.16 ± 0.03	13.24^ABC ± 0.91	28.87 ± 0.76	4.40^A ± 0.63	0.18 ± 0.04	2.31^BC ± 0.42	0.24 ± 0.05
	II × P	0.51^AB ± 0.05	49.85 ± 1.27	1.59 ± 0.25	0.13 ± 0.01	12.36^C ± 0.95	28.73 ± 1.56	3.90^AB ± 0.20	0.17 ± 0.02	2.56^BC ± 0.39	0.22 ± 0.04
	III × P	0.47^AB ± 0.13	48.53 ± 2.68	1.40 ± 0.26	0.10 ± 0.06	12.43^BC ± 0.69	30.77 ± 3.78	3.64^AB ± 0.61	0.17 ± 0.03	2.30^BC ± 0.55	0.19 ± 0.09
	IV × P	0.40^B ± 0.07	49.53 ± 2.92	1.33 ± 0.18	0.09 ± 0.02	10.24^D ± 0.99	32.86 ± 3.86	3.55^AB ± 0.35	0.17 ± 0.03	1.69^C ± 0.48	0.14 ± 0.05
	I × E	0.49^AB ± 0.05	51.38 ± 7.50	1.30 ± 0.13	0.14 ± 0.03	13.89^ABC ± 0.93	30.30 ± 1.22	3.35^AB ± 0.28	0.13 ± 0.01	2.45^BC ± 0.33	0.19 ± 0.05
	II × E	0.53^AB ± 0.15	49.03 ± 2.29	1.42 ± 0.60	0.15 ± 0.04	13.55^ABC ± 1.25	27.03 ± 5.65	4.48^A ± 1.40	0.13 ± 0.09	3.29^AB ± 1.64	0.28 ± 0.18
	III × E	0.55^AB ± 0.14	47.87 ± 1.57	1.61 ± 0.37	0.19 ± 0.12	14.68^A ± 1.17	28.84 ± 1.69	3.79^AB ± 0.64	0.13 ± 0.04	2.24^BC ± 0.47	0.16 ± 0.06
	IV × E	0.60^A ± 0.07	48.55 ± 1.36	1.56 ± 0.07	0.12 ± 0.03	14.20^AB ± 0.50	27.89 ± 1.94	4.32^A ± 0.63	0.17 ± 0.06	2.34^BC ± 0.48	0.24 ± 0.05
P-value		0.021	0.490	0.126	0.280	0.002	0.111	0.007	0.452	0.014	0.221

Values are presented as mean ± standard deviation (SD); ^a,bvalues with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ±values mean standard deviation; ¹I, 1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese; ²fatty acids: myristic acid (C14:0), palmitic acid (C16:0), palmitoleic acid (C16:1), margaric acid (C17:0), stearic acid (C18:0), oleic acid (C18:1n9), linoleic acid (C18:2n6, LA), α-linolenic acid (C18:3n3, ALA), behenic acid (C22:0), eicosapentaenoic acid (C20:5n3, EPA); ³the interaction (^{A, B}) between two factors: parent flock age and laying period, P < 0.05.

The change in the proportion of some FA contributed to the change in the total content of SFA and UFA on d 16 of incubation (Table 4). The highest percentage of UFA and MUFA was recorded in yolk sac lipids from embryos from the oldest geese and the lowest in group II. The analysis of the FA profile, depending on the laying period, showed a significant effect of this factor on the content of FA in the yolk sac lipids of developing embryos (Table 3). On d 16 of incubation the proportion of SFA (myristic, palmitic, margaric, and stearic) significantly increased with the laying duration. The exception was behenic acid, where the concentration was higher at the beginning of lay. The proportion of UFA (oleic and ALA) decreased significantly in successive weeks of the laying period (P < 0.05). A 15% increase in LA was recorded at the end of the laying period. The proportion of SFA increased by 28% and PUFA by 13% compared to the percentage of SFA found at the beginning of lay (Table 4). The proportion of UFA and MUFA decreased with the duration of lay by 34 and 40%, respectively.

Table 4. Fatty acid groups content in lipids of the yolk sac of goose eggs on day 16 of incubation.

Factor	Group^1	Fatty acids^2
		SFA	UFA	UFA/SFA	MUFA	PUFA
Parent flock age	I	60.81 ± 9.80	40.38^ab ± 7.63	0.70^b ± 0.24	36.08^ab ± 7.61	4.30 ± 0.74
	II	59.85 ± 8.78	40.12^b ± 8.81	0.71^b ± 0.27	35.89^b ± 9.49	4.23 ± 0.99
	III	58.87 ± 7.81	41.14^ab ± 8.69	0.74^ab ± 0.28	37.23^ab ± 8.96	3.91 ± 0.62
	IV	57.83 ± 9.24	42.17^a ± 9.24	0.78^a ± 0.31	38.20^a ± 9.83	3.97 ± 0.76
	P-value	0.062	0.012	0.001	0.021	0.290
Laying period	Beginning (B)	47.80^c ± 1.87	52.19^a ± 1.87	1.09^a ± 0.08	48.44^a ± 2.34	3.75^b ± 0.68
	Peak (P)	64.00^b ± 2.61	35.99^b ± 2.59	0.56^b ± 0.06	31.75^b ± 3.02	4.24^ab ± 0.61
	End (E)	66.55^a ± 4.69	34.38^c ± 2.15	0.51^b ± 0.05	30.07^b ± 2.93	4.31^a ± 0.94
	P-value	< 0.001	< 0.001	< 0.001	< 0.001	0.021
Interaction^3	I × B	49.27 ± 1.25	50.73 ± 1.25	1.03 ± 0.05	46.33 ± 1.73	4.40^AB ± 0.72
	II × B	48.94 ± 1.71	51.06 ± 1.71	1.05 ± 0.07	47.42 ± 2.38	3.65^AB ± 0.74
	III × B	47.27 ± 1.25	52.74 ± 1.25	1.12 ± 0.06	49.07 ± 1.16	3.66^AB ± 0.47
	IV × B	45.74 ± 0.79	54.27 ± 0.79	1.19 ± 0.04	50.94 ± 0.91	3.32^B ± 0.38
	I × P	64.83 ± 0.66	35.16 ± 0.67	0.54 ± 0.02	30.34 ± 0.90	4.83^A ± 0.68
	II × P	65.41 ± 1.40	34.61 ± 1.31	0.53 ± 0.03	30.32 ± 1.40	4.29^AB ± 0.19
	III × P	63.83 ± 2.99	36.17 ± 2.99	0.57 ± 0.08	32.17 ± 3.62	4.00^AB ± 0.69
	IV × P	61.94 ± 3.51	38.05 ± 3.51	0.62 ± 0.10	34.19 ± 3.86	3.86^AB ± 0.38
	I × E	68.34 ± 8.74	35.26 ± 0.97	0.52 ± 0.07	31.59 ± 1.25	3.67^AB ± 0.31
	II × E	66.55 ± 4.22	33.34 ± 4.40	0.51 ± 0.10	28.45 ± 5.80	4.90^A ± 1.52
	III × E	65.52 ± 0.99	34.53 ± 0.90	0.53 ± 0.02	30.45 ± 1.42	4.08^AB ± 0.73
	IV × E	65.82 ± 1.56	34.19 ± 1.56	0.52 ± 0.04	29.45 ± 1.95	4.74^AB ± 0.71
P-value		0.962	0.275	0.149	0.109	0.010

Values are presented as mean ± standard deviation (SD); ^a,bvalues with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ± values mean standard deviation; ¹I, 1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese;²fatty acids: SFA, saturated fatty acids; UFA, unsaturated fatty acids; UFA/SFA, unsaturated to saturated fatty acids ratio; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids; ³the interaction (^{A, B…}) between two factors: parent flock age and laying period; P < 0.05.

When analysing the interactions between the age of the parent flock and laying duration, on d 16 the highest proportion of myristic acid was in group IV and the lowest in groups I to V at the beginning of lay, stearic acid in the III group at the end of lay and the lowest in groups I to V at the beginning of lay, LA in group I at the beginning, II, and IV at the end of lay and for behenic the highest was in group I at the beginning of lay. A significantly higher proportion of PUFA was seen in group II at the end compared to group IV at the beginning of lay (P = 0.010).

With increasing age of the laying geese, the proportion of myristic acid decreased by 20% in eggs from the youngest females on d 22 of incubation (Table 5). The opposite tendency was seen in the other two FA cases, whereby the proportion of behenic acid and EPA in the yolk sac increased with the age of the female by 0.63% and 0.08% respectively, between groups I and IV. Changes in the FA profile of the embryo yolk sac depended on the laying, which were recorded on d 22 of incubation (Table 5). With increasing duration of laying, the proportion of myristic, palmitic, margaric, and stearic acids increased significantly, and similar differences were demonstrated for palmitoleic acid, oleic acid, ALA, and EPA, whereby content in the yolk sacs decreased in subsequent weeks of laying.

Table 5. The fatty acid content in lipids of the yolk sac of goose eggs on day 22 of incubation.

Factor	Group^1	Fatty acids^2
		C14:0	C16:0	C16:1	C17:0	C18:0	C18:1n9	C18:2n6	C18:3n3	C22:0	C20:5n3
Parent flock age	I	0.40^a ± 0.24	45.66 ± 7.60	1.65 ± 0.40	0.09 ± 0.05	8.74 ± 3.06	37.68 ± 9.36	3.58 ± 0.64	0.19 ± 0.05	1.98^b ± 0.86	0.11^c ± 0.03
	II	0.34^ab ± 0.19	44.33 ± 6.97	1.46 ± 0.46	0.08 ± 0.04	8.78 ± 3.12	39.33 ± 8.49	3.37 ± 0.95	0.21 ± 0.16	1.97^b ± 0.73	0.13^bc ± 0.05
	III	0.31^b ± 0.14	44.25 ± 6.12	1.52 ± 0.43	0.08 ± 0.04	9.09 ± 3.20	38.99 ± 9.45	3.21 ± 1.03	0.17 ± 0.08	2.25^ab ± 0.72	0.16^ab ± 0.06
	IV	0.32^b ± 0.17	43.64 ± 6.51	1.45 ± 0.40	0.07 ± 0.03	9.42 ± 3.15	38.79 ± 8.59	3.37 ± 0.72	0.16 ± 0.04	2.61^a ± 0.79	0.19^a ± 0.05
	P-value	0.014	0.103	0.426	0.152	0.317	0.286	0.639	0.283	0.016	< 0.001
Laying period	Beginning (B)	0.18^c ± 0.03	35.71^b ± 2.02	1.73^a ± 0.35	0.04^c ± 0.02	5.90^c ± 0.85	49.98^a ± 2.63	3.26^ab ± 0.95	0.24^a ± 0.13	2.84^a ± 0.78	0.18^a ± 0.06
	Peak (P)	0.28^b ± 0.07	48.77^a ± 3.33	1.20^b ± 0.32	0.08^b ± 0.02	8.33^b ± 1.02	36.13^b ± 3.06	3.12^b ± 0.73	0.15^b ± 0.05	1.82^b ± 0.69	0.13^b ± 0.06
	End (E)	0.56^a ± 0.14	48.93^a ± 1.54	1.62^a ± 0.40	0.12^a ± 0.03	12.79^a ± 1.30	29.97^c ± 1.26	3.75^a ± 0.73	0.16^b ± 0.05	1.95^b ± 0.51	0.14^b ± 0.05
	P-value	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001	0.036	0.002	< 0.001	0.006
Interaction^3	I × B	0.17 ± 0.02	35.59 ± 2.07	1.67 ± 0.21	0.05 ± 0.01	5.81 ± 0.56	49.98 ± 2.75	3.60 ± 0.74	0.24^AB ± 0.05	2.95 ± 0.69	0.13 ± 0.02
	II × B	0.18 ± 0.03	35.19 ± 2.50	1.72 ± 0.60	0.04 ± 0.02	5.69 ± 1.22	50.19 ± 1.20	3.86 ± 1.33	0.35^A ± 0.21	2.59 ± 0.49	0.16 ± 0.06
	III × B	0.19 ± 0.04	36.63 ± 2.43	1.76 ± 0.28	0.04 ± 0.02	5.79 ± 0.89	50.11 ± 4.70	2.71 ± 0.74	0.20^AB ± 0.07	2.40 ± 0.94	0.20 ± 0.06
	IV × B	0.18 ± 0.03	35.41 ± 1.23	1.79 ± 0.31	0.04 ± 0.01	6.31 ± 0.70	49.65 ± 1.25	2.88 ± 0.53	0.15^B ± 0.05	3.41 ± 0.72	0.21 ± 0.05
	I × P	0.35 ± 0.10	51.71 ± 1.56	1.50 ± 0.50	0.09 ± 0.03	7.94 ± 0.84	33.51 ± 2.04	3.16 ± 0.49	0.15^B ± 0.04	1.50 ± 0.48	0.10 ± 0.02
	II × P	0.27 ± 0.04	49.45 ± 2.22	1.05 ± 0.10	0.07 ± 0.02	8.01 ± 0.59	36.95 ± 2.36	2.63 ± 0.55	0.13^B ± 0.06	1.37 ± 0.77	0.09 ± 0.04
	III × P	0.25 ± 0.06	46.68 ± 2.88	1.14 ± 0.20	0.08 ± 0.02	8.69 ± 1.11	37.48 ± 3.92	3.02 ± 0.74	0.13^B ± 0.03	2.38 ± 0.59	0.15 ± 0.07
	IV × P	0.26 ± 0.04	47.25 ± 4.15	1.12 ± 0.18	0.07 ± 0.02	8.67 ± 1.43	36.59 ± 2.72	3.67 ± 0.85	0.18^B ± 0.05	2.02 ± 0.52	0.16 ± 0.06
	I × E	0.68 ± 0.12	49.69 ± 1.60	1.77 ± 0.48	0.14 ± 0.04	12.47 ± 1.71	29.54 ± 1.26	3.96 ± 0.50	0.17^B ± 0.02	1.48 ± 0.33	0.11 ± 0.03
	II × E	0.58 ± 0.08	48.34 ± 1.20	1.61 ± 0.25	0.13 ± 0.02	12.64 ± 0.93	30.85 ± 0.84	3.61 ± 0.24	0.16^B ± 0.03	1.95 ± 0.27	0.12 ± 0.02
	III × E	0.48 ± 0.09	49.44 ± 1.82	1.67 ± 0.49	0.11 ± 0.04	12.78 ± 1.73	29.36 ± 1.36	3.89 ± 1.30	0.18^B ± 0.11	1.97 ± 0.66	0.13 ± 0.03
	IV × E	0.51 ± 0.17	48.27 ± 1.38	1.44 ± 0.39	0.11 ± 0.01	13.28 ± 0.91	30.12 ± 1.31	3.54 ± 0.62	0.14^B ± 0.03	2.39 ± 0.32	0.19 ± 0.05
P-value		0.083	0.082	0.650	0.746	0.992	0.492	0.098	0.038	0.083	0.792

Values are presented as mean ± standard deviation (SD); ^a,bvalues with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ±values mean standard deviation; ¹I, 1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese;²fatty acids: SFA, saturated fatty acids; UFA, unsaturated fatty acids; UFA/SFA, unsaturated to saturated fatty acids ratio; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids; ³the interaction (^{A, B…}) between two factors: parent flock age and laying period; P < 0.05.

The palmitoleic acid and LA proportions were the highest at the beginning and end of the laying period. The SFA was higher by 30% at the end compared to the beginning of lay (Table 6). At the end of lay, the percentage of the UFA and MUFA decreased by 36 and 39%, respectively, and an interaction for ALA was seen at d 22 of incubation. In group II (at the beginning), a higher proportion of acid was found than in groups III (at the beginning) and in groups I to IV at the peak and end of lay (P = 0.038).

Table 6. Fatty acid groups content in lipids of the yolk sac of goose eggs on day 22 of incubation.

Factor	Group^1	Fatty acids^2
		SFA	UFA	UFA/SFA	MUFA	PUFA
Parent flock age	I	56.87 ± 9.25	43.20 ± 9.34	0.81 ± 0.34	39.32 ± 9.35	3.88 ± 0.67
	II	55.50 ± 8.98	44.50 ± 8.96	0.85 ± 0.33	40.79 ± 8.85	3.71 ± 1.11
	III	55.97 ± 8.93	44.05 ± 8.95	0.83 ± 0.32	40.51 ± 9.49	3.54 ± 1.07
	IV	56.06 ± 8.39	43.95 ± 8.40	0.82 ± 0.29	40.24 ± 8.74	3.71 ± 0.77
	P-value	0.374	0.420	0.634	0.357	0.758
Laying period	Beginning (B)	44.66^c ± 2.44	55.39^a ± 2.42	1.24^a ± 0.12	51.71^a ± 2.60	3.68 ± 1.06
	Peak (P)	59.27^b ± 2.73	40.73^b ± 2.73	0.69^b ± 0.07	37.33^b ± 2.90	3.39 ± 0.80
	End (E)	64.35^a ± 0.82	35.64^c ± 0.82	0.55^c ± 0.02	31.59^c ± 1.01	4.05 ± 0.76
	P-value	< 0.001	< 0.001	< 0.001	< 0.001	0.061
Interaction^3	I × B	44.57 ± 2.82	55.63 ± 2.67	1.26 ± 0.14	51.65 ± 2.85	3.98 ± 0.78
	II × B	43.70 ± 1.80	56.28 ± 1.77	1.29 ± 0.10	51.91 ± 0.80	4.37 ± 1.53
	III × B	45.05 ± 3.79	54.99 ± 3.86	1.23 ± 0.19	51.87 ± 4.69	3.11 ± 0.84
	IV × B	45.35 ± 0.93	54.68 ± 0.92	1.21 ± 0.05	51.44 ± 1.11	3.25 ± 0.59
	I × P	61.58 ± 1.62	38.43 ± 1.62	0.62 ± 0.04	35.01 ± 1.84	3.41 ± 0.51
	II × P	59.16 ± 2.18	40.86 ± 2.17	0.69 ± 0.06	38.01 ± 2.39	2.85 ± 0.64
	III × P	58.08 ± 3.56	41.92 ± 3.56	0.73 ± 0.10	38.62 ± 3.86	3.30 ± 0.77
	IV × P	58.28 ± 2.40	41.72 ± 2.41	0.72 ± 0.07	37.71 ± 2.56	4.01 ± 0.96
	I × E	64.45 ± 0.90	35.55 ± 0.89	0.55 ± 0.02	31.31 ± 1.08	4.24 ± 0.52
	II × E	63.65 ± 0.77	36.36 ± 0.78	0.57 ± 0.02	32.46 ± 0.82	3.90 ± 0.26
	III × E	64.77 ± 0.88	35.23 ± 0.87	0.54 ± 0.02	31.04 ± 0.89	4.19 ± 1.39
	IV × E	64.55 ± 0.39	35.45 ± 0.39	0.55 ± 0.01	31.57 ± 0.94	3.88 ± 0.63
P-value		0.272	0.249	0.531	0.573	0.096

Values are presented as mean ± standard deviation (SD); ^a,bvalues with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ± values mean standard deviation; ¹I, 1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese;²fatty acids: SFA, saturated fatty acids; UFA, unsaturated fatty acids; UFA/SFA, unsaturated to saturated fatty acids ratio; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids; ³the interaction (^{A, B…}) between two factors: parent flock age and laying period; P < 0.05.

On d 28, the proportion of individual FAs in the yolk sacs of embryos from groups I, III, and IV were similar (Table 7). The yolk sacs in eggs from two-year-old geese differed from the eggs of the remaining groups only in the content of palmitic acid, the proportion of which was, on average, higher by 2.4%. On d 28 of incubation, the content of SFA (myristic, palmitic, margaric, and stearic) in yolk sacs was increased (Table 7). Moreover, the proportion of oleic acid, ALA, behenic acid and EPA decreased.

Table 7. The fatty acid content in lipids of the yolk sac of goose eggs on day 28 of incubation.

Factor	Group^1	Fatty acids^2
		C14:0	C16:0	C16:1	C17:0	C18:0	C18:1n9	C18:2n6	C18:3n3	C22:0	C20:5n3
Parent flock age	I	0.37 ± 0.21	40.56^b ± 6.84	1.54 ± 0.36	0.10 ± 0.03	8.39 ± 2.84	42.76 ± 8.91	4.10 ± 1.15	0.25 ± 0.14	1.83 ± 0.64	0.13 ± 0.06
	II	0.33 ± 0.14	41.16^a ± 6.41	1.57 ± 0.38	0.09 ± 0.04	8.75 ± 3.58	43.08 ± 8.58	3.14 ± 0.38	0.20 ± 0.06	1.58 ± 0.71	0.14 ± 0.09
	III	0.35 ± 0.17	40.89^b ± 7.15	1.71 ± 0.33	0.08 ± 0.03	8.43 ± 3.22	42.76 ± 9.47	3.52 ± 0.75	0.21 ± 0.09	1.92 ± 0.78	0.16 ± 0.09
	IV	0.32 ± 0.19	39.03^b ± 8.61	1.37 ± 0.48	0.09 ± 0.03	8.74 ± 2.84	44.92 ± 10.81	3.27 ± 1.01	0.19 ± 0.08	1.94 ± 0.94	0.16 ± 0.08
	P-value	0.369	0.023	0.103	0.304	0.994	0.059	0.098	0.668	0.285	0.391
Laying period	Beginning (B)	0.19^c ± 0.04	32.37^c ± 3.18	1.56 ± 0.35	0.06^c ± 0.02	5.86^c ± 0.82	53.46^a ± 4.13	3.59 ± 1.12	0.27^a ± 0.12	2.48^a ± 0.75	0.23^a ± 0.06
	Peak (P)	0.27^b ± 0.06	42.44^b ± 3.09	1.43 ± 0.41	0.09^b ± 0.02	7.42^b ± 1.17	43.07^b ± 3.05	3.49 ± 0.63	0.21^ab ± 0.05	1.46^b ± 0.41	0.12^b ± 0.06
	End (E)	0.53^a ± 0.12	46.89^a ± 2.45	1.60 ± 0.43	0.12^a ± 0.03	11.85^a ± 1.88	33.91^c ± 2.28	3.46 ± 0.90	0.17^b ± 0.06	1.39^b ± 0.44	0.09^b ± 0.04
	P-value	< 0.001	< 0.001	0.436	< 0.001	< 0.001	< 0.001	0.944	0.003	< 0.001	< 0.001
Interaction^3	I × B	0.22 ± 0.02	33.38 ± 2.54	1.61^ABC ± 0.24	0.08^BCD ± 0.03	5.51 ± 0.49	51.58 ± 3.06	4.66 ± 1.40	0.35 ± 0.17	2.46 ± 0.42	0.20 ± 0.05
	II × B	0.20 ± 0.01	33.68 ± 1.54	1.67^AB ± 0.12	0.06^D ± 0.02	6.07 ± 0.72	52.63 ± 3.27	3.30 ± 0.31	0.25 ± 0.04	2.05 ± 0.92	0.22 ± 0.08
	III × B	0.21 ± 0.03	33.11 ± 3.57	1.84^A ± 0.14	0.06^D ± 0.02	5.56 ± 0.55	52.72 ± 5.14	3.41 ± 0.80	0.25 ± 0.10	2.67 ± 0.55	0.26 ± 0.06
	IV × B	0.13 ± 0.05	28.57 ± 2.64	1.04^D ± 0.33	0.05^D ± 0.01	6.41 ± 1.35	57.80 ± 2.74	2.82 ± 1.03	0.21 ± 0.11	2.80 ± 1.04	0.23 ± 0.03
	I × P	0.23 ± 0.08	40.57 ± 3.18	1.12^CD ± 0.30	0.09^ABCD ± 0.02	8.24 ± 1.69	44.67 ± 2.71	3.25 ± 0.88	0.17 ± 0.07	1.57 ± 0.20	0.10 ± 0.03
	II × P	0.32 ± 0.07	45.65 ± 2.02	1.75^AB ± 0.66	0.07^CD ± 0.01	6.41 ± 1.10	41.25 ± 2.19	3.13 ± 0.57	0.20 ± 0.06	1.13 ± 0.24	0.09 ± 0.01
	III × P	0.30 ± 0.02	43.25 ± 1.93	1.57^ABCD ± 0.17	0.10^ABCD ± 0.01	7.42 ± 0.39	41.68 ± 2.18	3.96 ± 0.55	0.25 ± 0.04	1.35 ± 0.17	0.12 ± 0.03
	IV × P	0.25 ± 0.04	40.31 ± 2.50	1.29^BCD ± 0.11	0.10^ABCD ± 0.02	7.62 ± 0.85	44.65 ± 4.34	3.61 ± 0.36	0.22 ± 0.02	1.78 ± 0.66	0.16 ± 0.11
	I × E	0.62 ± 0.09	47.73 ± 0.63	1.71^AB ± 0.32	0.12^AB ± 0.03	11.35 ± 1.05	32.79 ± 1.04	4.05 ± 0.80	0.19 ± 0.05	1.35 ± 0.44	0.09 ± 0.04
	II × E	0.47 ± 0.09	45.95 ± 2.22	1.37^BCD ± 0.33	0.14^A ± 0.02	12.84 ± 1.81	34.64 ± 1.45	2.99 ± 0.32	0.15 ± 0.03	1.37 ± 0.38	0.09 ± 0.03
	III × E	0.52 ± 0.13	47.25 ± 2.91	1.65^AB ± 0.50	0.10^ABC ± 0.02	11.92 ± 2.12	33.44 ± 2.58	3.36 ± 0.85	0.16 ± 0.07	1.51 ± 0.60	0.08 ± 0.02
	IV × E	0.51 ± 0.12	46.64 ± 3.53	1.67^AB ± 0.55	0.10^ABC ± 0.02	11.28 ± 2.44	34.77 ± 3.38	3.42 ± 1.28	0.16 ± 0.08	1.34 ± 0.46	0.10 ± 0.05
P-value		0.135	0.088	0.037	0.035	0.334	0.331	0.282	0.324	0.785	0.509

Values are presented as mean ± standard deviation (SD); ^a,bvalues with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ± values mean standard deviation; ¹I, 1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese;²fatty acids: SFA, saturated fatty acids; UFA, unsaturated fatty acids; UFA/SFA, unsaturated to saturated fatty acids ratio; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids; ³the interaction (^{A, B…}) between two factors: parent flock age and laying period; P < 0.05.

The highest percentage of SFA was recorded in the yolk sac of embryos at the end of lay and was during in the initial production (Table 8). The content of UFA and MUFA decreased with the duration of laying, although, on d 28, significant differences (and interactions) were seen in the proportion of palmitoleic and margaric acids. The highest proportion was in group III at the beginning (palmitoleic acid, P = 0.037) and in group II at the end of lay (margaric acid, P = 0.035). For the other fatty acids and their groups, there was no statistically significant relationship in terms of interactions between the age of the parent flock and the laying duration (P > 0.05).

Table 8. Fatty acid groups content in lipids of the yolk sac of goose eggs on day 28 of incubation.

Factor	Group^1	Fatty acids^2
		SFA^2	UFA	UFA/SFA	MUFA	PUFA
Parent flock age	I	51.24 ± 9.03	48.77 ± 9.04	1.01 ± 0.36	44.29 ± 8.80	4.48 ± 1.29
	II	51.90 ± 8.79	48.14 ± 8.85	0.98 ± 0.36	44.65 ± 8.67	3.48 ± 0.46
	III	51.68 ± 9.43	48.36 ± 9.49	1.00 ± 0.39	44.46 ± 9.52	3.89 ± 0.84
	IV	50.12 ± 9.90	49.90 ± 10.11	1.08 ± 0.45	46.28 ± 10.46	3.61 ± 1.06
	P-value	0.057	0.052	0.058	0.078	0.131
Laying period	Beginning (B)	40.96^c ± 3.13	59.11^a ± 3.16	1.46^a ± 0.18	55.02^a ± 3.89	4.08 ± 1.22
	Peak (P)	51.68^b ± 2.54	48.31^b ± 2.54	0.93^b ± 0.10	44.49^b ± 2.83	3.81 ± 0.70
	End (E)	60.77^a ± 1.46	39.2^c ± 1.46	0.64^c ± 0.03	35.51^c ± 2.06	3.71 ± 0.96
	P-value	< 0.001	< 0.001	< 0.001	< 0.001	0.554
Interaction^3	I × B	41.64 ± 2.18	58.39 ± 2.14	1.41 ± 0.13	53.18 ± 2.83	5.21 ± 1.54
	II × B	42.05 ± 2.85	58.07 ± 2.91	1.39 ± 0.15	54.29 ± 3.24	3.78 ± 0.38
	III × B	41.61 ± 4.19	58.49 ± 4.26	1.43 ± 0.25	54.57 ± 5.01	3.92 ± 0.93
	IV × B	37.96 ± 1.74	62.09 ± 1.86	1.64 ± 0.13	58.84 ± 2.52	3.26 ± 1.11
	I × P	50.70 ± 1.46	49.31 ± 1.46	0.97 ± 0.06	45.79 ± 2.43	3.51 ± 0.97
	II × P	53.57 ± 1.21	46.42 ± 1.21	0.87 ± 0.04	43.00 ± 1.65	3.42 ± 0.64
	III × P	52.41 ± 2.32	47.59 ± 2.33	0.91 ± 0.08	43.26 ± 2.35	4.33 ± 0.59
	IV × P	50.06 ± 3.88	49.94 ± 3.88	1.01 ± 0.16	45.95 ± 4.31	3.99 ± 0.44
	I × E	61.17 ± 0.77	38.84 ± 0.77	0.64 ± 0.02	34.50 ± 1.25	4.33 ± 0.88
	II × E	60.76 ± 1.24	39.24 ± 1.24	0.65 ± 0.03	36.01 ± 1.37	3.23 ± 0.33
	III × E	61.31 ± 1.93	38.69 ± 1.93	0.63 ± 0.05	35.09 ± 2.32	3.60 ± 0.92
	IV × E	59.88 ± 1.66	40.12 ± 1.66	0.67 ± 0.05	36.44 ± 2.90	3.68 ± 1.37
P-value		0.658	0.668	0.432	0.469	0.286

Values are presented as mean ± standard deviation (SD); ^a,bvalues with different letters in columns differ significantly between groups (parent flock age or laying period), with P-value < 0.05; ± values mean standard deviation; ¹I, 1-year-old geese; II, 2-year-old geese; III, 3-year-old geese; IV, 4-year-old geese;²fatty acids: SFA, saturated fatty acids; UFA, unsaturated fatty acids; UFA/SFA, unsaturated to saturated fatty acids ratio; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids; ³the interaction (^{A, B…}) between two factors: parent flock age and laying period; P < 0.05.

Discussion

The significant influence of goose age on the content of FAs in egg yolk lipids was previously reported by Kalaycı and Yılmaz (2011), who showed a similar proportion of these FA in goose egg yolks. The current research showed that the age of laying geese significantly influenced the content of UFA: LA, ALA, and EPA. The largest proportion of these FA were seen in eggs from the youngest females. Razmaitė, Šveistienė, and Švirmickas (2014) showed significantly higher content of palmitic, oleic, LA, behenic and EPA acids in the group of three-year-old Lithuanian geese compared to one-year-old females. However, age did not affect the content of SFA, MUFA and PUFA although it varied in both age groups of females in the current trial. This contradicted the data provided by Mazanowski (2012), where the conclusion was that neither the age nor breed significantly affected the quantitative composition of FA in yolks.

The current research revealed a higher susceptibility to changes in the FA profile in the lipids in yolks during the laying period. During the production of goose eggs, the proportion of SFA increased, including myristic, palmitic, stearic, and margaric, and decreased UFA; oleic, LA, EPA. Okruszek et al. (2006) noted similar trends in duck eggs and found that, in the 22^nd week of laying, there was an increase in palmitic and stearic acid in the yolk lipids compared to the 6^th week, by 7.3 and 27%, respectively. The FA in the yolk differ between the laying phases in different species, which has been related to the size of the egg, yolk and even protein content (Razmaitė, Šveistienė, and Švirmickas 2014). It should be noted that nutrition influences yolk composition (fatty acid content) (Chen et al. 2014). Therefore, it may be suggested that such differences could be influenced by the degree of nutrient utilisation by laying geese, which varies with age and laying season. Gu et al. (2021) showed changes in both digestive function and egg quality in laying hens regarding age.

In the case of most UFAs, a decrease in yolk content was noted in line with laying duration. These results were identical to the data obtained in the experiment in which the influence of the laying period on the FA profile in hen eggs was assessed. Mickey et al. (1998) found that the content of palmitic and stearic acid was higher in the yolks of eggs from 51- and 64-week-old hens than in the eggs of young hens aged 36 weeks. With increasing laying duration, the proportion of SFA increased, and the percentage of UFA, MUFA, and PUFA decreased in the current research, which contradicted other published data. In the case of goose and duck eggs (Tilki and Inal 2004; Okruszek et al. 2006), these FA groups remained similar throughout the laying period. Koppenol et al. (2014) compared layers in different laying periods (28-, 43-, and 58-weeks-old), and found that, as laying continued, the content of SFA and PUFA decreased by 3.1 and 13.1%, respectively, and MUFA increased by an average of 9.4%.

The absorption and use of FA of the yolk by embryos depends on hen age. In the research of Koppenol et al. (2014), the composition of the yolk sacs of chicks from eggs laid in different periods after 28 d of incubation was similar, and differences were seen only in the content of palmitic acid, the largest proportion of which was found in the group of two-year-old females.

The current research showed that laying duration had a higher impact on the FA profile during incubation than the reproductive season (parent flock age). On d 28, a lower proportion of myristic, palmitic, margaric and stearic acids and a higher content of behenic acid, EPA and UFA with 18 carbon atoms were recorded in yolk sacs of goose eggs obtained at the beginning of the lay compared to the peak and the end of production. Comparing hens aged 36- and 52-weeks-old, Şahan, Ipek, and Sozcu (2014) showed that, at the end of incubation, yolk sacs from eggs laid by younger hens contained more myristic, palmitoleic and oleic acid. The yolk sacs from eggs laid by older hens were characterised by a higher proportion of stearic acid, LA and arachidonic acid. When examining the composition of the yolk sacs of embryos from three age groups of chickens (28-, 35- and 49-weeks-old) in the last phase of incubation, Boonsinchai (2015) found that the proportion of myristic, palmitic, palmitoleic, stearic, oleic, dihomo-y-linolenic, LA, EPA acids was higher in those from the oldest hens. Yalçin et al. (2008) showed that the chick yolk sacs did not differ in MUFA content, although the eggs analysed came from different laying periods. Thes authors showed that eggs at the beginning of lay were characterised by a higher proportion of UFA and a lower PUFA content compared to the final production period. In previous studies, there has been a lack of unanimity regarding FA profiles in the last incubation period. This indicated several additional influencing factors, including hatching phase, the time in the hatcher, temperature and the humidity in the incubator and hatcher (Burnham et al. 2001; Malec et al. 2004; Yalçin et al. 2008).

Comparing the FA composition of fresh eggs to yolk sacs at the end of incubation, Mickey et al. (1998) found that the most significant changes in the proportion up to d 18 of embryogenesis were seen in oleic and arachidonic acid, proving embryonic use of lipids. Between d 18 of incubation and hatching, oleic acid content decreased by 4.19% in eggs from older hens, while it increased by 1.05% in eggs from 36-week-old hens. The current research showed similar changes in oleic acid content. From the d 1to d 28 of incubation, oleic acid increased by 8.7 and 13.4%, respectively, in eggs obtained at the beginning and peak of lay and decreased by 0.8% in eggs from the final laying period. According to Mickey et al. (1998), the level of stearic acid decreased in the yolk sacs from eggs laid by young hens, which corresponded to the results of the current research. Mickey et al. (1998) found that stearic acid was converted to oleic acid through the Δ9 desaturase enzyme, and showed that palmitoleic acid decreased 3.5-fold during incubation of eggs laid by young hens. In older females, there were no such differences in the content of this acid between fresh yolks and the content of the yolk sac taken after hatching. The observed change in the proportion of palmitoleic, oleic and arachidonic acid, and, to a lesser extent, LA, in eggs from young hens may be due to changes in the activity of various enzymatic processes occurring in the yolk sac membrane and in the transport of lipoproteins. Levels of corticotropin hormone and cortisol may influence this. Şahan, Ipek, and Sozcu (2014) stated it was not entirely clear how the age of the female influenced the change in the proportion of FA in the yolk sac during incubation. This may be due to the different absorption coefficients or the activity of enzymatic processes in the yolk sac membrane in eggs from different age groups of hens. Future trials should therefore consider factors such as bird origin, egg weight and incubation conditions. Donaldson (1964) suggested that embryonic fatty acid deposition mainly occurs in structural (early-stage) fats. In older embryos, the deposition of FA in adipose tissue causes a dilution of the structural fats so that the overall composition is similar to the content seen in the yolk.

The layer geese were kept in a semi-intensive system (access to the enclosure) in the current trial. As these are seasonal birds, their adaptation to conditions and proper functioning depends on the environment, which is related to hormonal balance (Shi et al. 2008). It can be suggested that goose age and laying period interactions may have caused the birds to react differently to changing environmental conditions (temperature, humidity, access to grass in the enclosure) during the various laying periods. This might have resulted in a different proportion of FA being present in the hatching eggs. The content of PUFA in egg yolks decreases with increasing age of the parent geese, and duration of lay affected the content of the majority of FA in yolk lipids. As the production period increased, the proportion of UFA decreased, while SFA increased compared to the beginning of lay. The use of FA by developing embryos in eggs from different age groups of females was similar. The FA profile in the lipids of the embryo’s yolk sac depends on the date the eggs were obtained during laying. The different FA profiles in the yolk content during incubation indicated changes in the activity of various enzymatic processes of the embryonic yolk sac membrane from the beginning, peak and end of production.

The most significant impact in terms of interactions was demonstrated for fresh eggs. Mostly, these results varied amongst individual fatty acids. The results showed intensive changes in the embryonic development of geese, so it appears crucial to adjust the conditions of reproduction to the age, duration or laying season of geese to obtain high-quality eggs with good hatchability.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This research was part of the doctoral dissertation of the first author. Thesis: Joanna Kucharska-Gaca. 2018. The analysis of embryonic and post-embryonic development of geese depends on the age of the parent flock. Publisher: UTP - University of Science and Technology in Bydgoszcz. The APC was paid from the internal fund of the Faculty of Animal Breeding and Biology of the Bydgoszcz University of Science and Technology, Poland.