Split-feeding as a sustainable feeding strategy for improving egg production and quality, nutrient digestibility, and environmental impact in laying hens

Abstract

This study evaluated the effects of split feeding on production performance, egg quality, nutrient digestibility, odor emissions, and economic benefits in laying hens. A total of 468 Hy-Line Brown hens, aged 63 weeks, were divided into three dietary treatments over 13 weeks. The control group received a single basal diet, while two split-feeding groups (TRT1 and TRT2) received diets differing in nutrient composition between the morning and afternoon. TRT1 and TRT2 contained 10 % and 15 % less crude protein (CP), respectively, compared to CON. TRT1 maintained the same levels of metabolizable energy, standardized ileal digestibility amino acids, calcium, and phosphorus as CON, while TRT2 had 5 % lower metabolizable energy, standardized ileal digestibility amino acids, and calcium, and 15 % lower phosphate. Results showed that TRT1 reduced the incidence of downgraded eggs by 19.3 % in the later phase of the experiment (9–12 weeks) compared to CON (P = 0.025, 0.043, 0.022, 0.011). Both TRT1 and TRT2 reduced diet costs by 6 % and 15 %, respectively. Additionally, split feeding improved nutrient digestibility, reduced harmful gas emissions, and resulted in a 4.5 % feed cost savings per kilogram of salable egg production compared to CON. These findings suggest that split feeding (TRT1) could enhance egg quality, reduce environmental impact, and contribute to the sustainability of poultry farming

Introduction

Nutritional management is a critical aspect of commercial egg production, as laying hens require precise nutrient intake to maintain high productivity and egg quality. The process of eggshell formation takes place mainly during the hours of the night, with the most intense period occurring about 12-18 h after an egg is laid. This period of heightened activity peaks around 18 h post-laying and gradually diminishes before the formation of the next egg (Araújo et al., 2011). Traditional feeding practices, which often involve providing a single diet composition throughout the day, may not fully align with the hens' physiological needs during the different stages of egg formation. In response to this mismatch, between the timing of eggshell formation and nutrient availability, farmers sometimes have implemented strategies such as top-feeding coarse limestone in the afternoon to address the Calcium (Ca) requirements for eggshell formation. While effective for Ca supplementation, this approach primarily addresses Ca supplementation, making it difficult to optimize other critical nutrients.

Split feeding, which involves adjusting nutrient composition and limestone particle size at different times of the day, offers a more holistic solution to better match the birds' nutrient requirements during the egg formation process (Hervo et al., 2022). Molnár et al. (2018) determined that traditional feeding was significantly less effective than all split feeding strategies, resulting in egg mass (36.56 g, P ≤ 0.001) and an increased feed conversion ratio (FCR; 2.482, P <0.05). Prior research also indicated that the standardized ileal digestibility (SID) of Ca is lower for fine limestone compared to coarse limestone (Li et al., 2021). This supports the inclusion of coarse limestone in the afternoon diet for laying hens as it better aligns with their Ca needs during the nighttime egg production phase.

Recent studies suggest that split feeding strategies not only improve productivity and egg quality but also hold promise for addressing the growing environmental concerns associated with poultry farming. Reducing Crude protein (CP) content in laying hens’ diets effectively reduces total Nitrogen (N) excretion and ammonia (NH₃) emissions in poultry facilities. Additionally, split feeding optimizes nutrition provision to satisfy the requirements of laying hens, minimizing nutrient wastage and improving nutrient absorption (Jahan et al., 2024). Studies on low-protein diets have shown that N intake in laying hens consuming split feeds is lower than that of hens on a traditional diet, leading to a 7 % decrease in daily N excretion in the split feed group compared to the control group (Horváth et al., 2024). As the poultry industry faces increasing pressure to reduce feed costs and minimize the environmental impact of production, strategies such as split feeding provide an opportunity to optimize nutrient delivery while simultaneously mitigating harmful emissions. By optimizing nutrient delivery according to the hens' physiology, split feeding presents an opportunity to enhance both productivity and sustainability in commercial egg production (Ferket et al., 2002).

This study aims to investigate the impact of split feeding on laying hens, specifically focusing on its effects on egg production, egg quality, nutrient digestibility, and environmental outcomes. By comparing a traditional single-diet approach with two split-feeding strategies that adjust nutrient density and limestone particle size, this research seeks to determine whether split feeding can improve production performance and environmental effects.

Materials and methods

Experimental design, diets and animals

A research proposal was submitted to Dankook University (DK-1-2405) Animal care and Use committee. After approval, the experiment was conducted over 13 weeks, involving 468 Hy-line Brown hens aged 63 weeks at the research farm of Dankook University, located in Sejong City, South Korea. The hens were randomly assigned to three dietary treatments with 13 replicates of 12 birds each. The treatments included: The CON received a basal diet throughout the day, providing consistent levels of metabolizable energy (ME, 2,720 Kcal/Kg), SID lysine (0.70 %), Ca, phosphorus (P), and CP (15.5 %). This diet, offered as a single feed with coarse limestone, served as the reference diet. In contrast, TRT1 (S100) followed a split feeding regimen, with 40 % of the diet provided in the morning and 60 % in the afternoon. The nutrient density of the morning and afternoon feeds ensured that the total daily intake of ME, SID amino acids, Ca, and P matched the levels in the control diet, though TRT1 was formulated with 10 % less CP overall. The morning feed included fine limestone, while coarse limestone was provided in the afternoon to meet the hens' Ca needs at specific times of the day. Similarly, TRT2 (S95) employed a split feeding strategy (40 % morning, 60 % afternoon) but introduced further nutrient reductions compared to the control diet, including 95 % ME, 95 % SID amino acids, 12 % less CP, 6 % less Ca, 13 % less P, and 32 % less available P.

Diet composition

The diets were formulated in accordance with the nutrient specifications delineated in the Hy-Line Brown Management Guide (2023), utilizing corn and soybean meal as primary ingredients. Detailed formulations are presented in Table 2. Limestone particle size was adjusted for each treatment: the CON utilized coarse limestone exclusively, while the split-feeding treatments (TRT1 and TRT2) incorporated fine limestone in the morning feed and coarse limestone in the afternoon feed. Nutrient composition was calculated, analyzed at the Lab of Dankook University, and recorded.

Table 2.

Feed composition of laying hen diet (as fed basis).

Item	Experimental diet
	CON	S100M	S100A	S95M	S95A
Ingredients (%)
Corn	58.60	59.68	66.63	56.87	62.62
Wheat bran	-	-	-	6.19	8.35
Soybean meal (CP 44 %)	21.96	19.48	15.69	12.67	13.30
Corn DDGS	5.30	11.57	1.45	18.60	-
Tallow	1.83	1.41	0.29	0.04	-
MDCP	1.10	1.25	0.98	0.55	0.30
Limestone	10.10	5.30	13.48	3.70	13.95
Salt	0.25	0.25	0.25	0.25	0.25
Amino Acid mix	0.26	0.46	0.0.63	0.53	0.63
Vitamin mix1	0.20	0.20	0.20	0.20	0.20
Mineral mix2	0.20	0.20	0.20	0.20	0.20
Choline (50 %)	0.10	0.10	0.10	0.10	0.10
Phytase	0.05	0.05	0.05	0.05	0.05
CTCzyme	0.05	0.05	0.05	0.05	0.05
Total	100.00	100.00	100.00	100.00	100.00
Calculated value
ME, kcal/kg	2720	2800	2667	2670	2525
Crude Protein, %	15.50	16.30	12.50	16.00	12.00
Calcium, %	4.16	2.35	5.38	1.60	5.45
Phosphorus, %	0.46	0.52	0.43	0.45	0.36
Available Phosphorus, %	0.31	0.35	0.29	0.25	0.18
SID Lys, %	0.70	0.76	0.66	0.72	0.63
SID Met, %	0.35	0.38	0.33	0.36	0.32
Linoleic acid, %	1.95	2.17	1.47	2.20	1.41
Choline, mg/kg	1775	1834	1620	1809	1563
Analyzed value
Crude Protein, %	15.51	16.55	12.68	16.28	12.11
Calcium, %	4.26	2.48	5.48	1.61	5.60
Phosphorus, %	0.52	0.55	0.48	0.49	0.38
Lysine, %	0.82	0.87	0.74	0.84	0.71
Methionine, %	0.40	0.42	0.35	0.38	0.37

¹Provided per kg of diet: vitamin A, 8,000 IU; vitamin D3, 3,300 IU; vitamin E, 20 g; vitamin K3, 2.5 g; vitamin B1, 2.5 g; vitamin B2, 5.5 g; vitamin B6, 4 g; vitamin B12, 23 mg; biotin, 75 mg; folic acid, 0.9 g; niacin, 30 g; d-calcium pantothenate, 8 g.

²Provided per kg of diet: Fe, 40 g as ferrous sulfate; Cu, 8 g as copper sulfate; Mn, 90 g as manganese oxide; Zn, 80 g as zinc oxide; 1.2 g as potassium iodide; and Se, 0.22 g as sodium selenite.

*S100: Split feeding with total nutrient intake equivalent to 100 % of the control diet; S95: Split feeding with total nutrient intake equivalent to 95 % of the control diet. M: Morning diet; A: Afternoon diet.

Experiment flock management

The feeding trial was conducted at Dankook University Research Farm, located in Sejong City, South Korea. All treatment groups—CON, TRT1, and TRT2— were subjected to a one-week adaptation period before the start of the experiment, during which all birds were provided the same diet as the CON group. Each hen was housed in an individual cage (50 cm x 40 cm x 38 cm), with each treatment group receiving 40 % of the daily feed in the morning and 60 % in the afternoon. Water was provided ad libitum. Additionally, the lighting schedule was adjusted according to the age recommendations in the Hy-Line Brown Management Guide (2023).

Productivity and laying rate

Body weight was measured by replicate at the end of the adaptation period (1 week), at the start of the trial, and weeks 4, 8, and the end of the trial (12 weeks). Average daily feed intake (ADFI) was calculated by subtracting the remaining feed from the total feed provided and dividing by the number of birds per replicate. FCR was calculated by dividing feed intake by egg mass. The laying rate was expressed as a percentage, calculated by dividing the total number of eggs by the number of hens in each treatment group during the trial period. The feed cost per kilogram of salable egg mass was calculated by dividing the total feed cost during the trial period by the total egg mass produced, after subtracting the egg mass from downgraded abnormal eggs to obtain the salable egg mass. Downgraded abnormal eggs were identified based on criteria such as shell defects (e.g., cracks, thin shells, misshapen shells), cleanliness (e.g., presence of stains or adhering dirt), and internal defects (e.g., blood spots, meat spots), following the standards outlined by the USDA Egg-Grading Manual.

Egg quality

Egg quality measurements were taken at weeks 4, 8, and at the end of the trial (12 weeks), with 30 eggs randomly selected from each treatment group for the egg quality evaluation. Egg weight was determined using an egg multi tester (Touhoku Rhythm Co. Ltd., Tokyo, Japan). Eggshell breaking strength was measured with an eggshell force gauge (Model II, Robotmation Co., Ltd., Tokyo, Japan). Eggshell thickness was recorded using a dial pipe gauge (Ozaki MFG. Co., Ltd., Tokyo, Japan), based on the average thickness of the rounded end, pointed end, and equatorial region of the egg, excluding the inner membrane. Yolk color, albumen height, and Haugh unit (HU) were analyzed with an egg multi-tester (Touhoku Rhythm Co., Ltd., Tokyo, Japan). Eggshell color was evaluated using an eggshell color fan (Dacho Ltd., Hwaseong, Korea).

Nutrient digestibility

To measure nutrient digestibility, 0.2 % chromium oxide (Cr₂O₃) was added to the feed as an indigestible marker at weeks 4, 8, and at the end of the trial (12 weeks). Fecal samples were collected over a 7-day period using trays positioned beneath the cages. The collected feces were pooled by replicate and stored at −20°C until analysis.​ Prior to chemical analysis, fecal samples were thawed, dried in a forced-air oven at 60°C for 72 h, and finely ground using a Wiley mill equipped with a 1 mm screen. The concentrations of N, P, and Ca in both feed and fecal samples were determined following AOAC (2000) methods. Cr₂O₃concentrations in feed and fecal samples were quantified using UV absorption spectrophotometry (Shimadzu, UV-1201, Japan), following the method described by Williams et al. (1962). Apparent nutrient digestibility coefficients were calculated using the indicator method with Cr₂O₃ serving as the indigestible marker

Odorous gas emission in excreta

To analyze odorous gases in the excreta, samples were collected at weeks 4, 8, and the end of the trial (12 weeks). The fresh excreta (300 g) were placed in a 2,600 mL sealed plastic container and allowed to ferment at room temperature for 7 days. ​Following the methodology detailed by Hossain et al. (2024), concentrations of NH₃, hydrogen sulfide (H₂S), methyl mercaptan, acetic acid, and carbon dioxide (CO₂) were measured using a multi-gas detector (MultiRAE Lite model PGM-6208, RAE, USA).

Statistical analysis

All data were analyzed using the General Linear Model procedure of SAS (2013), and the significance of differences between means was tested by Duncan's multiple range test (Duncan, 1955). Significant value was held as P < 0.05.

Results

Body weight growth

The effects of nutrient composition, limestone particle size, and split feeding on body weight growth are shown in Table 3. From the start to the end of the experiment, no significant differences in body weight growth were observed among the experimental groups.

Table 3.

The effects of split feeding on growth performance in laying hens^1.

Items	CON	TRT1	TRT2	SEM2	P-value3
Body weight, g
Initial (Adaptation Week)	1917	1917	1918	1	0.859
Week 4	1932	1930	1930	4	0.920
Week 8	1945	1943	1942	6	0.950
Week 12	1960	1958	1957	7	0.951

¹Abbreviation: CON, Basal diet (one diet, conventional feeding with coarse limestone); TRT1, SF-100 (same nutrition density of day total consumption with control group, but different ratio of nutrients and particle size of limestone with split feeding; morning time fine limestone / afternoon time coarse limestone); TRT2, SF-95 (95 % nutrient density of ME and SID amino acid, 90 % density in available phosphorus and calcium of day- total consumption compared with basal diet; morning time fine limestone / afternoon time coarse limestone).

Productivity and egg production cost

Table 4 presents the impact of nutrient composition, limestone particle size, and split feeding on productivity metrics, including egg production rate, downgraded abnormal eggs, FCR, feed intake, and feed cost per kilogram of salable eggs. Since no significant differences were observed during weeks 1 to 6, these data have been moved to Supplementary Table 1, while Table 4 focuses on weeks 7 to 12, where significant treatment effects were detected. There was a general trend of lower downgraded abnormal egg rates in TRT1, followed by CON and TRT2 over the entire experimental period. From weeks 9 to 10, TRT1 exhibited a significantly lower downgraded abnormal egg ratio compared to TRT2 (P = 0.025, 0.043), while from weeks 11 to 12, the downgraded abnormal egg ratio of TRT1 was significantly lower than that of CON and TRT2 (P = 0.022, 0.011). At weeks 11 and 12 (end of the trial), both CON and TRT2 had significantly higher cracked egg rates than TRT1 (P = 0.022, 0.011). In terms of laying rates, there was a trend of higher laying rates in CON, followed by TRT1 and TRT2 throughout the trial. However, TRT1, which had a 10 % lower daily CP intake, did not show a significant reduction in laying rate compared to CON. At weeks 12, CON showed a significantly higher laying rate compared to TRT2 (P = 0.024). In the CON group, although the laying rate was higher compared to the other groups, the proportion of downgraded abnormal eggs was also higher than in the TRT1 group. Additionally, the diet unit cost was also 5 % higher, resulting in a 4.5 % higher feed cost per kilogram of salable eggs on average over weeks 1 to 12. While differences were observed in downgraded abnormal eggs and laying rates among the treatments, ADFI and FCR remained unaffected, with no significant differences detected between groups.

Table 4.

The effects of Split feeding on the production performance.

Items	CON	TRT1	TRT2	SEM2	P-value3
Week 7
Downgraded egg, %	2.1	1.8	2.3	0.4	0.656
Egg production, %	83.1	82.3	81.8	0.7	0.487
Feed Cost (Won)/ Salable Egg Kg	943.7	902.5	899.8	12.2	0.257
FCR	2.05	2.07	2.09	0.02	0.344
ADFI, g	109.3	109.5	109.7	0.3	0.639
Week 8
Downgraded egg, %	2.3	1.9	2.5	0.3	0.310
Egg production, %	82.3	81.4	80.7	0.9	0.478
Feed Cost (Won)/ Salable Egg Kg	953.8	910.1	914.5	13.1	0.193
FCR	2.06	2.08	2.12	0.03	0.285
ADFI, g	109.1	109.4	109.5	0.3	0.573
Week 9
Downgraded egg, %	2.4ab	2.0b	2.7a	0.2	0.025
Egg production, %	81.6	80.4	79.6	1.0	0.356
Feed Cost (Won)/ Salable Egg Kg	961.4	922.1	928.2	16.0	0.218
FCR	2.08	2.10	2.15	0.03	0.355
ADFI, g	108.9	109.1	109.4	0.7	0.887
Week 10
Downgraded egg, %	2.6ab	2.1b	2.8a	0.2	0.043
Egg production, %	80.8	79.5	78.5	0.7	0.118
Feed Cost (Won)/ Salable Egg Kg	973.4b	932.1ab	943.4a	12.9	0.059
FCR	2.10	2.12	2.18	0.03	0.323
ADFI, g	108.8	109.1	109.3	0.6	0.829
Week 11
Downgraded egg, %	2.8a	2.2b	3.0a	0.2	0.022
Egg production, %	79.9a	78.7ab	77.4b	0.8	0.114
Feed Cost (Won)/ Salable Egg Kg	984.7b	942.5ab	959.5a	15.1	0.049
FCR	2.11	2.14	2.20	0.03	0.138
ADFI, g	108.7	109.0	109.3	0.7	0.843
Week 12
Downgraded egg, %	3.0a	2.4b	3.3a	0.2	0.011
Egg production, %	79.2a	78.1ab	76.5b	0.7	0.024
Feed Cost (Won)/ Salable Egg Kg	997.2b	948.9b	973.6a	17.0	0.055
FCR	2.13	2.15	2.23	0.03	0.132
ADFI, g	108.6	108.9	109.2	1.0	0.901

¹Abbreviation: CON, Basal diet (one diet, conventional feeding with coarse limestone); TRT1, S100 (same nutrition density of day total consumption with control group, but different ratio of nutrients and particle size of limestone with split feeding; morning time fine limestone / afternoon time coarse limestone); TRT2, S95 (95 % nutrient density of ME and SID amino acid, 90 % density in available phosphorus and calcium of day- total consumption compared with basal diet; morning time fine limestone / afternoon time coarse limestone).

²Standard error of means.

^a,b,3Means in the same row with different superscript differ significantly (P < 0.05).

Egg quality

Egg quality parameters were influenced by nutrient composition, limestone particle size, and split feeding, as shown in Table 5. At week 8, TRT1 exhibited significantly higher eggshell thickness and strength compared to TRT2 (P = 0.046, 0.047). By the end of the experiment (week 12), TRT1 also had significantly higher eggshell thickness and strength compared to both CON and TRT2 (P = 0.001).

Table 5.

The effects of split feeding on egg quality in laying hens^1.

Items	CON	TRT1	TRT2	SEM2	P-value3
Week 4
Egg Shell color	9.3	9.2	9.2	0.3	0.949
Haugh Unit	81.5	81.3	81.0	0.7	0.858
Egg weight, g	64.1	64.3	64.1	0.3	0.833
Yolk color	7.4	7.5	7.4	0.4	0.972
Albumen height, mm	7.3	7.1	7.0	0.3	0.875
Eggshell Strength, kg/cm2	3.96	4.00	3.84	0.10	0.530
Eggshell Thickness, mm-2	37.5	37.9	36.4	1.2	0.651
Week 8
Egg Shell color	9.0	9.0	9.0	0.4	0.990
Haugh Unit	81.0	80.7	80.4	0.7	0.819
Egg weight, g	64.3	64.6	64.2	0.8	0.960
Yolk color	7.3	7.3	7.2	0.3	0.977
Albumen height, mm	7.3	7.0	6.9	0.4	0.669
Eggshell Strength, kg/cm2	3.83ab	3.94a	3.64b	0.08	0.046
Eggshell Thickness, mm-2	36.8ab	37.4a	35.3b	0.6	0.047
Week 12
Egg Shell color	8.8	8.9	8.8	0.5	0.990
Haugh Unit	80.7	80.3	79.9	1.0	0.844
Egg weight, g	64.5	65.0	64.3	1.2	0.891
Yolk color	7.1	7.1	7.1	0.4	0.997
Albumen height, mm	7.2	7.0	6.8	0.4	0.789
Eggshell Strength, kg/cm2	3.58b	3.86a	3.32b	0.09	0.001
Eggshell Thickness, mm-2	35.2b	37.0a	33.6b	0.6	0.001

¹Abbreviation: CON, Basal diet (one diet, conventional feeding with coarse limestone); TRT1, S100 (same nutrition density of day total consumption with control group, but different ratio of nutrients and particle size of limestone with split feeding; morning time fine limestone / afternoon time coarse limestone); TRT2, S95 (95 % nutrient density of ME and SID amino acid, 90 % density in available phosphorus and calcium of day- total consumption compared with basal diet; morning time fine limestone / afternoon time coarse limestone).

²Standard error of means.

^a,b,3Means in the same row with different superscript differ significantly (P < 0.05).

Nutrient digestibility

At week 8, TRT1 showed significantly higher digestibility of P and Ca compared to TRT2 (P = 0.039, 0.043). By the end of the experiment (week 12), TRT1 had significantly higher P and Ca digestibility than both CON and TRT2 (P = 0.009). Detailed findings are presented in Table 6.

Table 6.

The effects of split feeding on nutrient digestibility in laying hens^1.

Items, %	CON	TRT1	TRT2	SEM2	P-value3
Week 4
Nitrogen	45.01	45.46	44.79	0.92	0.878
Phosphorus	30.33	30.77	29.89	0.84	0.767
Calcium	58.43	58.80	57.97	0.83	0.787
Week 8
Nitrogen	43.32	44.04	43.11	0.60	0.552
Phosphorus	28.96ab	29.71a	28.08b	0.34	0.039
Calcium	56.58ab	57.25a	55.69b	0.33	0.043
Week 12
Nitrogen	42.07	43.00	41.73	0.72	0.483
Phosphorus	26.43b	27.79a	25.24b	0.38	0.009
Calcium	54.33b	55.62a	53.28b	0.34	0.009

¹Abbreviation: CON, Basal diet (one diet, conventional feeding with coarse limestone); TRT1, S100 (same nutrition density of day total consumption with control group, but different ratio of nutrients and particle size of limestone with split feeding; morning time fine limestone / afternoon time coarse limestone); TRT2, S95 (95 % nutrient density of ME and SID amino acid, 90 % density in available phosphorus and calcium of day- total consumption compared with basal diet; morning time fine limestone / afternoon time coarse limestone).

²Standard error of means.

^a,b,3Means in the same row with different superscript differ significantly (P < 0.05).

Odorous gas emissions in excreta

As shown in Table 7, odorous gas emissions were affected by nutrient composition, limestone particle size, and split feeding. At week 8, CON exhibited significantly higher NH₃ levels than TRT1 (P = 0.015), and by the end of the experiment (week 12), CON had significantly higher NH₃ levels than both TRT1 and TRT2 (P = 0.005)

Table 7.

The effects of split feeding on gas emission in laying hens^1.

Items, ppm	CON	TRT1	TRT2	SEM2	P-value3
Week 4
NH3	29.88	29.00	29.63	0.69	0.674
H2S	6.20	5.93	6.13	0.51	0.926
Methyl mercaptans	12.50	12.88	12.75	0.95	0.961
Acetic acid	10.75	10.88	10.50	0.77	0.942
CO2	2475	2463	2500	160	0.986
Week 8
NH3	32.63a	30.75b	31.63ab	0.31	0.015
H2S	6.98	6.60	6.83	0.71	0.932
Methyl mercaptans	14.13	14.75	14.50	0.44	0.627
Acetic acid	11.88	11.63	11.50	0.49	0.865
CO2	2675	2625	2675	158	0.967
Week 12
NH3	34.75a	31.88b	32.50b	0.40	0.005
H2S	8.13	7.65	7.98	0.23	0.387
Methyl mercaptans	15.25	15.38	15.00	0.99	0.964
Acetic acid	12.88	12.88	12.75	0.84	0.993
CO2	2900	2875	2875	91	0.975

¹Abbreviation: CON, Basal diet (one diet, conventional feeding with coarse limestone); TRT1, S100 (same nutrition density of day total consumption with control group, but different ratio of nutrients and particle size of limestone with split feeding; morning time fine limestone / afternoon time coarse limestone); TRT2, S95 (95 % nutrient density of ME and SID amino acid, 90 % density in available phosphorus and calcium of day- total consumption compared with basal diet; morning time fine limestone / afternoon time coarse limestone).

²Standard error of means.

^a,b,3Means in the same row with different superscript differ significantly (P < 0.05).

Discussion

The results of this study suggest that split feeding, which involves providing different feed compositions at various times of the day, can enhance laying hen productivity and egg quality, while also contributing to improved nutrient digestibility. The findings align with previous research indicating that hens may benefit from tailored nutrient intake that matches their physiological needs throughout the day (Tunç and Cufadar, 2015). Split feeding (TRT1) led to a lower proportion of downgraded eggs and improved eggshell quality compared to the control group in this study. The use of fine limestone in the morning and coarse limestone in the afternoon in TRT1 could provide an optimal Ca supply for eggshell formation, effectively meeting the high Ca demand in the evening time. Araujo et al. (2011) mentioned that coarse dietary Ca in laying hen's diet can stay longer in the gizzard and can be absorbed more to provide adequate Ca to egg formation. This resulted in a significant reduction in downgraded abnormal eggs, ultimately increasing the proportion of salable eggs. Studies have similarly shown that proper Ca timing improves eggshell strength and reduces egg losses (Keshavarz, 2003; Van Emous and Mens, 2021). Additionally, the improved eggshell quality observed in TRT1 may have implications for food safety. Stronger eggshells are less prone to cracking, potentially reducing the risk of bacterial contamination and extending shelf life (De Reu et al., 2009). This aspect could be particularly valuable for the egg industry, as it may lead to reduced losses during transportation and storage.

TRT2, which had a more aggressive reduction in nutrient density (5 % lower energy and amino acids, 12 % lower CP, 6 % lower Ca, and 12 % lower P), resulted in lower productivity and eggshell quality compared to both CON and TRT1 groups. In contrast, TRT1, which maintained the same total daily intake of energy, amino acids, Ca, and P as CON but distributed these nutrients across morning and afternoon diets, outperformed CON in most production parameters and reduced feed cost per kilogram of salable eggs. These findings suggest that split feeding with precise nutrient management can enhance both productivity and economic efficiency. However, the lower nutrient levels in TRT2 resulted in less favorable performance outcomes, highlighting the need for further research to determine the optimal level of nutrient reductions that can sustain productivity and cost-effectiveness without compromising overall performance.

Akter et al. (2025) reported that an optimized AM/PM split-feeding regimen, which adjusts nutrient composition according to the diurnal physiological needs of laying hens, can enhance protein and calcium digestibility. This improvement in nutrient digestibility indicates that split feeding can facilitate better nutrient utilization, thereby reducing nutrient excretion and improving overall dietary efficiency. A key environmental benefit of split feeding is the reduction in harmful gas emissions. Horváth et al. (2024) demonstrated that split feeding reduces nitrogen intake, leading to decreased urinary excretion of NH₄⁺-N and uric acid-N, which in turn lowers NH₃ emissions. Our results indicate that both TRT1 and TRT2 groups showed decreased NH₃ levels, indicating that split feeding can lower N excretion through improved nutrient utilization, thus contributing to reduced air pollution.

However, the observed reduction in NH₃ emissions may also be attributed to the lower crude protein content in the TRT1 and TRT2 diets. Given that both treatment groups received 10 % and 15 % less CP than the control, the decrease in N excretion could primarily be a result of reduced protein intake rather than the feeding strategy itself. While split feeding may improve nutrient utilization, further studies with controlled CP levels across feeding strategies are needed to isolate its specific effects on N metabolism. Nevertheless, the observed reduction in NH₃ emissions in TRT1 and TRT2 suggests that split feeding may contribute to lower nitrogen excretion, potentially improving overall environmental sustainability. The better CP intake and optimized nutrient digestibility not only decreased N emissions but also minimized N and P excretion in the manure, which is crucial for mitigating water pollution (Nahm, 2007). Reduced P in manure, for instance, could help mitigate eutrophication in nearby water bodies when the manure is used as fertilizer. These benefits of lower N emissions for improved air quality and reduced N and P for water protection could reinforce the potential of split feeding to support sustainability goals in poultry farming by benefiting both air and water conservation.

Improved nutrient digestibility, particularly for Ca and P, was observed in TRT1. This led to better nutrient absorption, reducing the need for excessive dietary supplementation and improving feed efficiency. The precise feed formulation used in split feeding reduced feed costs, particularly with TRT1 reducing the feed cost per kilogram of salable egg by 4.5 % compared to the control group (Tables 1 and Table 4). This is crucial for the economic viability of poultry operations, as lowering feed costs without compromising productivity offers significant financial benefits (Vries et al., 2015).

Table 1.

Nutritional specification and cost of treatment diet.

Item	CON (One diet)	S100			S95
		Morning	Afternoon	Average	Morning	Afternoon	Average
ME, kcal/kg	2720	2800	2667	2720	2670	2525	2583
SID Lys, %	0.70	0.76	0.66	0.70	0.72	0.63	0.67
Crude Protein, %	15.50	16.30	12.50	14.02	16.00	12.00	13.60
Calcium, %	4.16	2.35	5.38	4.17	1.60	5.45	3.91
Phosphorus, %	0.46	0.52	0.43	0.47	0.45	0.36	0.40
Avail. P %	0.31	0.35	0.29	0.31	0.25	0.18	0.21
Diet Price/kg	447	458	404	423	438	401	416

*ME: Metabolizable energy; * Amino acids and other than SID lysine was designed according to the recommended ratio relative to SID lysine for the HY-Line brown breed; * The diet price was calculated based on each ingredient in the Korean market as of October 2024; * The daily average cost was calculated based on allocating 40 % to the morning diet and 60 % to the afternoon diet.

Although the results are promising, the scale of this experiment may not be sufficient to confidently predict the same outcomes in larger, fully commercialized poultry farms. Inconsistent feeding schedules, environmental stressors such as heat stress, quality of feed raw material, and poor ventilation systems could significantly affect the performance of split feeding. These factors must be considered when applying this feeding strategy at the production flock level. Further large-scale studies are needed to validate these findings under field conditions in layer farms to determine whether split feeding can reliably enhance productivity and reduce costs in a commercial context.

Conclusion

This study suggests that split feeding may enhance egg production, egg quality, and nutrient digestibility in laying hens. Specifically, TRT1, which adjusted nutrient intake throughout the day, demonstrated improved performance and reduced feed cost compared to the control group. Additionally, split feeding significantly reduced NH₃ emissions and improve Ca and P digestibility. These findings suggest that aligning nutrient delivery with hens' physiological needs could address economic and environmental challenges in poultry farming. However, the scale of this experiment may not be sufficient to confidently predict the same outcomes in larger, fully commercialized poultry farms. Inconsistent feeding schedules, environmental stressors such as heat stress, quality of feed raw materials, and poor ventilation systems could significantly affect the performance of split feeding. Therefore, further research is necessary to confirm these results and to explore the long-term effects of split feeding on nutrient absorption, environmental sustainability, and economic viability in diverse poultry production systems.

Disclosures

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.