Effects of reduced crude protein diets while maintaining essential amino acid concentrations on growth performance, nitrogen output, ammonia production, and meat yield

Abstract

An experiment was conducted to determine the effect of feeding reduced crude protein (CP) diets to Ross × Ross 708 male broilers while providing adequate essential amino acid (AA) concentrations on growth performance, nitrogen (N) and ammonia output, and carcass characteristics from d 1 to 33 post hatch. Birds received 1 of 6 dietary treatments (10 replicate pens per treatment) varying in CP content. Diet 1 (control) was formulated with DL-Met, L-Lys, and L-Thr (23.2, 20.7, and 19.1% CP) in the starter (1–14 d of age), grower (15–25 d of age), and finisher (26–33 d of age) periods, respectively. Dietary L-Val, Gly (only in starter period), L-Ile, L-Arg, and L-Trp were sequentially supplemented in the order of limitation in Diets 2 through 6. Dietary CP was reduced gradually across the dietary treatments resulting in a CP reduction in Diets 1 to 6 by 3.4, 3.4, and 2.3% points in the starter, grower, and finisher periods, respectively. At d 14, 25, and 33 posthatch, feed conversion decreased (P < 0.05) with L-Val addition (Diet 2) and increased (P < 0.01) with L-Val to L-Trp addition (Diet 6) to the control. Dietary treatments did not alter weights and yields of carcass, breast, drum, or thighs. Dietary CP reduction with added L-Val (Diet 2), L-Val to L-Arg (Diet 5), or L-Val to L-Trp (Diet 6) increased abdominal fat (P < 0.01) compared with control. Nitrogen excretion (g/bird; P = 0.003) and equilibrium ammonia concentration (mg/kg; P = 0.041) at day 33 reduced by 16% and 48% respectively in birds fed reduced-CP diets with L-Val to L-Trp (Diet 6) compared with control-fed birds. This study indicated that sequential addition of supplemental AA in the order of limitation from DL-Met to L-Arg allowed reduction of dietary CP beyond 2%-point without depressing growth performance and meat yield of broilers from day 1 to 33 while reducing nitrogen excretion and ammonia emissions.

INTRODUCTION

From a sustainability and environmental standpoint, utilizing reduced crude protein (CP) diets for broiler chickens holds significant promise in reducing production costs, nitrogen (N) excretion (Hernández et al., 2012), and ammonia emissions (Ferguson et al., 1998). Nonetheless, prior research has revealed that decreasing CP levels by more than 2% comes with trade-offs in terms of performance goals and meat yield outcomes (Namroud et al., 2008; Hernández et al., 2012; Kriseldi et al., 2018). These compromises have been attributed to various factors, including suboptimal quantities and balances of essential amino acids (AA) in the diet, alterations in dietary electrolyte balance, insufficient nonspecific N for nonessential AA synthesis, and changes in the ratios of essential to nonessential AA (Aftab et al., 2006).

The emergence of feed-grade AA has empowered nutritionists to formulate cost-effective diets while upholding acceptable performance benchmarks. This development is of significance as it allows for more substantial reductions in CP levels with adequate supplementation of AA that are less limiting, beyond Met, Lys, and Thr. Kriseldi et al. (2017, 2018) demonstrated that feeding reduced CP diets (up to a 2% reduction) while supplementing with DL-Met, L-Lys, L-Thr, L-Val, L-Ile, L-Arg, and L-Trp did not compromise growth performance (days 1–21) or meat yield (days 15–35) compared with diets supplemented only with DL-Met, L-Lys, and L-Thr. Other studies have also highlighted the conditional essentiality of Gly in broilers fed reduced CP diets, suggesting a need for a minimum total Gly concentration during the starter period, in addition to meeting essential AA requirements, to achieve optimal growth performance (Corzo et al., 2004; Dean et al., 2006; Waguespack et al., 2009; Awad et al., 2015; Kriseldi et al., 2017, 2018).

While these investigations have shown the potential of feeding reduced CP diets with sequential addition of essential AA beyond Thr during the starter and grower phases, there is a paucity of research that examines this effect from hatch to slaughter. Consequently, an experiment was conducted to assess the consequences of feeding reduced CP diets to broiler chickens while maintaining essential AA concentrations, and a minimum Gly concentration in the starter period, on growth performance, carcass yield, N output, and ammonia production throughout a 5-wk production period.

MATERIALS AND METHODS

Protocols of animal experiments were reviewed and approved by the Auburn University Institutional Animal Care and Use Committee.

Bird Husbandry

One thousand five hundred Ross × Ross 708 male broiler chicks (Aviagen North America, Huntsville, AL) were obtained from a commercial hatchery. Chicks were vaccinated for Marek's disease, infectious bronchitis, and Newcastle disease at the hatchery. Broilers were randomly distributed into 60 floor pens (25 birds per pen) at 1 d of age and reared in a solid-sided house designed with a cross-ventilation system. The experimental facility was equipped with vent boards, exhaust fans, evaporative cooling pads, forced-air heaters, and an electronic controller to maintain optimum ventilation and temperature. Each pen contained a hanging feeder, a nipple drinker line, and pine shavings. Birds received feed and water ad libitum. Broilers were placed in 1.83 m × 1.07 m pens, which allowed for an initial stocking density of 0.08 m²/ bird. Lighting was set at 30 lux at placement and subsequently dimmed to 10 lux, and 5 lux at d 7 and day 14 posthatch, respectively. Photoperiod was set to 23L:1D from placement to d 7, and then set to 20L:4D for the remainder of the experiment. Light intensity settings were verified at bird level (30 cm) using a photometric sensor with National Institute of Standards and Technology-traceable calibration (I-250A Light Meter, LI-COR Bioscience, Lincoln, NE). Ambient temperature was maintained to ensure bird comfort with the set point 33°C at placement and decreased gradually with advancing bird age to a final set point of 20°C at the end of the experiment. Birds and feed were weighed at d 1, 14, 25, and 33 for the determination of body weight (BW) gain, feed intake, and feed conversion ratio (FCR). The incidence of mortality was recorded daily.

Dietary Treatments

Experimental diets were formulated to meet the amino acid guidelines of the Evonik AMINO Chick 2.0 and nutrient recommendations of the Brazilian Tables for Poultry and Swine (Rostagno, 2017). Diets were formulated on digestible AA ratios and standardized AA digestibility coefficients of feed ingredients were used from AMINODat 5.0. Starter diets were provided from 1 to 14 d of age (Table 1), grower diets from 15 to 25 d of age (Table 2), and finisher diets were provided from 26 to 33 d of age (Table 3). For all growth periods, Diet 1 was formulated with added DL-Met, L-Lys, and L-Thr (control diet). Free AA were sequentially supplemented in the order of limitation (L-Val, Gly (starter only), L-Ile, L-Arg, L-Trp) to maintain adequate essential AA concentrations while simultaneously decreasing dietary CP contents in Diets 2 to 6. Dietary CP was reduced in Diets 1 to 6 by 3.4, 3.4, and 2.3% points in the starter, grower, and finisher periods, respectively. In addition, starter diets (1–14 d of age) were formulated to contain 1.70 total Gly + Ser to digestible Lys ratio, but a minimum of Gly + Ser was not employed in the grower and finisher diets. This was based on previous recommendations (Corzo et al., 2004; Waguespack et al., 2009; Yuan et al., 2012) and results from our lab (Kriseldi et al., 2018). Diets were provided in crumble form during the starter period and as whole pellet during subsequent growth periods. After mixing of the experimental grower and finisher diets, representative diet samples were collected to determine pellet integrity using standard procedures (American Society of Agricultural Engineers, 1993).

Table 1.

Ingredient and nutrient composition of dietary treatments fed to Ross × Ross 708 broilers from 1 to 14 d of age, as-fed.

	Dietary treatment
Ingredient, %	1	2	3	4	5	6
Corn	53.15	56.35	56.17	58.02	61.34	67.93
Soybean Meal	40.69	37.83	37.85	36.11	32.90	26.50
Vegetable Oil	2.92	2.31	2.36	1.99	1.33	—
Limestone	1.05	1.06	1.06	1.06	1.07	1.09
Dicalcium Phosphate	1.02	1.05	1.05	1.07	1.10	1.16
DL-Met1	0.32	0.35	0.35	0.36	0.38	0.43
Salt	0.32	0.32	0.32	0.32	0.32	0.32
L-Lys Sulphate (54.6%)1	0.16	0.28	0.28	0.36	0.49	0.76
L-Thr1	0.11	0.14	0.14	0.16	0.20	0.28
L-Val	—	0.04	0.04	0.07	0.12	0.22
Gly	—	—	0.11	0.17	0.29	0.52
L-Ile	—	—	—	0.03	0.08	0.18
L-Arg	—	—	—	—	0.09	0.26
L-Trp	—	—	—	—	—	0.03
Mineral Premix2	0.10	0.10	0.10	0.10	0.10	0.10
Vitamin Premix3	0.10	0.10	0.10	0.10	0.10	0.10
Choline Chloride	0.05	0.06	0.06	0.07	0.08	0.11
Phytase4	0.01	0.01	0.01	0.01	0.01	0.01
Calculated composition, % (unless otherwise noted)
AMEn, kcal/kg	3,000	3,000	3,000	3,000	3,000	3,000
Crude protein	23.23	22.31	22.44	21.95	21.23	19.81
Digestible Lys	1.24	1.24	1.24	1.24	1.24	1.24
Digestible Met + Cys	0.94	0.94	0.94	0.94	0.94	0.94
Digestible Met	0.64	0.65	0.65	0.66	0.67	0.69
Digestible Thr	0.84	0.84	0.84	0.84	0.84	0.84
Digestible Val	0.97	0.97	0.97	0.97	0.97	0.97
Digestible Ile	0.89	0.84	0.84	0.84	0.84	0.84
Digestible Arg	1.42	1.35	1.35	1.30	1.30	1.30
Digestible Trp	0.25	0.24	0.24	0.23	0.21	0.21
Total Lys	1.38	1.37	1.37	1.37	1.36	1.34
Total Met + Cys	1.04	1.04	1.04	1.04	1.03	1.02
Total Met	0.67	0.67	0.67	0.68	0.70	0.72
Total Thr	0.98	0.98	0.98	0.97	0.96	0.95
Total Val	1.09	1.09	1.09	1.08	1.07	1.05
Total Ile	1.00	0.95	0.95	0.95	0.94	0.92
Total Arg	1.55	1.47	1.47	1.42	1.42	1.40
Total Trp	0.28	0.27	0.27	0.27	0.24	0.24
Total Leu	1.92	1.84	1.84	1.80	1.72	1.56
Total His	0.62	0.59	0.59	0.57	0.54	0.48
Total Phe	1.16	1.10	1.10	1.07	1.00	0.88
Total Gly + Ser	2.10	2.10	2.11	2.11	2.11	2.11
Total Gly+Ser : Dig Lys	1.69	1.69	1.70	1.70	1.70	1.70
Ca	0.92	0.92	0.92	0.92	0.92	0.92
Non-Phytate P	0.45	0.45	0.45	0.45	0.45	0.45
Na	0.18	0.18	0.18	0.18	0.18	0.18
Analyzed Composition, %
Crude Protein	23.02	22.21	21.83	22.17	21.74	19.98
Total Lys	1.37	1.34	1.32	1.33	1.32	1.29
Total Met + Cys	0.93	0.92	0.97	1.02	0.93	0.95
Total Met	0.61	0.60	0.65	0.70	0.63	0.68
Total Thr	0.96	0.94	0.93	0.94	0.90	0.89
Total Val	1.06	1.03	1.01	1.02	1.00	0.99
Total Ile	0.98	0.91	0.88	0.89	0.87	0.85
Total Arg	1.63	1.45	1.42	1.40	1.36	1.36
Total Leu	1.87	1.77	1.73	1.67	1.62	1.49
Total His	0.59	0.55	0.54	0.52	0.49	0.45
Total Phe	1.10	1.05	1.02	1.00	0.94	0.84
Total Gly+Ser	2.09	1.96	2.03	2.05	2.00	2.15

¹Sources of DL-Met, L-Lys sulphate, and L-Thr, are MetAAMINO, Biolys, ThreAMINO, respectively (Evonik Nutrition and Care GmbH, Germany).

²Mineral premix include per kg of diet: Mn (manganese sulfate), 120 mg; Zn (zinc sulfate), 100 mg; Fe (iron sulfate monohydrate), 30 mg; Cu (tri-basic copper chloride), 8 mg; I (ethylenediamine dihydriodide), 1.4 mg; and Se (sodium selenite), 0.3 mg.

³Vitamin premix includes per kg of diet: Vitamin A (Vitamin A acetate), 18,739 IU; Vitamin D3 (cholecalciferol), 6,614 IU; Vitamin E (DL-alpha tocopherol acetate), 66 IU; menadione (menadione sodium bisulfate complex), 4 mg; Vitamin B12 (cyanocobalamin), 0.03 mg; folacin (folic acid), 2.6 mg: D-pantothenic acid (calcium pantothenate), 31 mg; riboflavin (riboflavin), 22 mg; niacin (niacinamide), 88 mg; thiamin (thiamin mononitrate), 5.5 mg; biotin (biotin), 0.18 mg; and pyridoxine (pyridoxine hydrochloride), 7.7 mg.

⁴Quantum Blue, AB Vista Feed Ingredients, Marlborough, UK

Table 2.

Ingredient and nutrient composition of dietary treatments fed to Ross × Ross 708 broilers from 15 to 25 d of age, as-fed.¹

	Dietary treatment
Ingredient, %	1	2	3	4	5	6
Corn	60.56	63.18	63.18	64.74	67.14	74.13
Soybean Meal	33.09	30.75	30.75	29.35	27.13	20.67
Vegetable Oil	3.36	2.86	2.86	2.55	2.06	0.63
Limestone	0.92	0.93	0.93	0.93	0.94	0.95
Dicalcium Phosphate	0.99	1.01	1.01	1.02	1.05	1.11
DL-Met2	0.25	0.27	0.27	0.28	0.30	0.35
Salt	0.32	0.32	0.32	0.32	0.32	0.33
L-Lys Sulphate (54.6%)2	0.18	0.28	0.28	0.34	0.43	0.70
L-Thr2	0.08	0.11	0.11	0.12	0.15	0.23
L-Val	—	0.03	0.03	0.06	0.09	0.19
L-Ile	—	—	—	0.02	0.06	0.15
L-Arg	—	—	—	—	0.06	0.23
L-Trp	—	—	—	—	—	0.03
Mineral Premix3	0.10	0.10	0.10	0.10	0.10	0.10
Vitamin Premix4	0.08	0.08	0.08	0.08	0.08	0.08
Choline Chloride	0.06	0.07	0.07	0.08	0.08	0.11
Phytase5	0.01	0.01	0.01	0.01	0.01	0.01
Calculated composition, % (unless otherwise noted)
AMEn, kcal/kg	3,100	3,100	3,100	3,100	3,100	3,100
Crude protein	20.69	19.95	19.95	19.52	18.95	17.31
Digestible Lys	1.08	1.08	1.08	1.08	1.08	1.08
Digestible Met	0.54	0.55	0.55	0.56	0.57	0.59
Digestible Met + Cys	0.82	0.82	0.82	0.82	0.82	0.82
Digestible Thr	0.73	0.73	0.73	0.73	0.73	0.73
Digestible Val	0.86	0.86	0.86	0.86	0.86	0.86
Digestible Ile	0.78	0.75	0.75	0.75	0.75	0.75
Digestible Arg	1.23	1.17	1.17	1.13	1.13	1.13
Digestible Trp	0.21	0.20	0.20	0.19	0.18	0.18
Total Lys	1.20	1.19	1.19	1.19	1.18	1.16
Total Met + Cys	0.91	0.90	0.90	0.90	0.90	0.89
Total Met	0.57	0.58	0.58	0.58	0.59	0.61
Total Thr	0.86	0.85	0.85	0.85	0.84	0.82
Total Val	0.97	0.96	0.96	0.96	0.95	0.94
Total Ile	0.87	0.83	0.83	0.83	0.82	0.81
Total Arg	1.35	1.28	1.28	1.24	1.24	1.22
Total Leu	1.77	1.72	1.72	1.69	1.64	1.49
Total His	0.55	0.53	0.53	0.51	0.49	0.43
Total Phe	1.03	0.98	0.98	0.96	0.92	0.79
Total Gly + Ser	1.86	1.78	1.78	1.73	1.65	1.43
Total Gly+Ser : Dig Lys	1.72	1.65	1.65	1.60	1.53	1.32
Ca	0.84	0.84	0.84	0.84	0.84	0.84
Non-Phytate P	0.43	0.43	0.43	0.43	0.43	0.43
Na	0.18	0.18	0.18	0.18	0.18	0.18
Analyzed Composition, %
Crude Protein	19.78	20.58	20.03	20.03	19.16	17.44
Total Lys	1.13	1.16	1.19	1.17	1.17	1.17
Total Met + Cys	0.90	0.88	0.89	0.87	0.85	0.86
Total Met	0.59	0.57	0.58	0.56	0.55	0.59
Total Thr	0.80	0.82	0.84	0.82	0.82	0.80
Total Val	0.91	0.93	0.93	0.94	0.95	0.90
Total Ile	0.80	0.81	0.82	0.80	0.81	0.77
Total Arg	1.29	1.27	1.29	1.24	1.26	1.19
Total Leu	1.60	1.62	1.63	1.60	1.61	1.43
Total His	0.50	0.50	0.50	0.49	0.48	0.42
Total Phe	0.93	0.92	0.93	0.91	0.89	0.76
Total Gly+Ser	1.75	1.74	1.76	1.71	1.67	1.46

¹Pellet durability index of diets 1 to 6 was 92.86, 91.96, 90.38, 88.00, 90.83, 90.44%, respectively.

²Sources of DL-Met, L-Lys sulphate, and L-Thr are MetAMINO, Biolys, and ThreAMINO, respectively (Evonik Nutrition and Care GmbH, Germany).

³Mineral premix include per kg of diet: Mn (manganese sulfate), 120 mg; Zn (zinc sulfate), 100 mg; Fe (iron sulfate monohydrate), 30 mg; Cu (tri-basic copper chloride), 8 mg; I (ethylenediamine dihydriodide), 1.4 mg; and Se (sodium selenite), 0.3 mg.

⁴Vitamin premix includes per kg of diet: Vitamin A (Vitamin A acetate), 18,739 IU; Vitamin D3 (cholecalciferol), 6,614 IU; Vitamin E (DL-alpha tocopherol acetate), 66 IU; menadione (menadione sodium bisulfate complex), 4 mg; Vitamin B12 (cyanocobalamin), 0.03 mg; folacin (folic acid), 2.6 mg: D-pantothenic acid (calcium pantothenate), 31 mg; riboflavin (riboflavin), 22 mg; niacin (niacinamide), 88 mg; thiamin (thiamin mononitrate), 5.5 mg; biotin (biotin), 0.18 mg; and pyridoxine (pyridoxine hydrochloride), 7.7 mg.

⁵Quantum Blue, AB Vista Feed Ingredients, Marlborough, UK

Table 3.

Ingredient and nutrient composition of dietary treatments fed to Ross × Ross 708 broilers from 26 to 33 d of age, as-fed.¹

	Dietary treatment
Ingredient, %	1	2	3	4	5	6
Corn	64.09	66.01	66.01	67.12	69.17	73.12
Soybean Meal	29.02	27.30	27.30	26.32	24.44	20.79
Vegetable Oil	4.08	3.71	3.71	3.49	3.08	2.27
Limestone	0.95	0.96	0.96	0.96	0.96	0.97
Dicalcium Phosphate	0.76	0.78	0.78	0.79	0.80	0.84
DL-Met2	0.23	0.24	0.24	0.25	0.26	0.29
Salt	0.33	0.33	0.33	0.33	0.33	0.33
L-Lys Sulphate (54.6%)2	0.21	0.28	0.28	0.32	0.40	0.55
L-Thr2	0.08	0.10	0.10	0.11	0.13	0.17
L-Val	—	0.03	0.03	0.04	0.07	0.12
L-Ile	—	—	—	0.01	0.04	0.10
L-Arg	—	—	—	—	0.05	0.15
L-Trp	—	—	—	—	—	0.02
Mineral Premix3	0.10	0.10	0.10	0.10	0.10	0.10
Vitamin Premix4	0.08	0.08	0.08	0.08	0.08	0.08
Choline Chloride	0.06	0.07	0.07	0.07	0.08	0.09
Phytase5	0.01	0.01	0.01	0.01	0.01	0.01
Calculated composition, % (unless otherwise noted)
AMEn, kcal/kg	3,180	3,180	3,180	3,180	3,180	3,180
Crude protein	19.13	18.58	18.58	18.28	17.80	16.87
Digestible Lys	1.00	1.00	1.00	1.00	1.00	1.00
Digestible Met	0.50	0.51	0.51	0.51	0.52	0.53
Digestible Met + Cys	0.76	0.76	0.76	0.76	0.76	0.76
Digestible Thr	0.68	0.68	0.68	0.68	0.68	0.68
Digestible Val	0.80	0.80	0.80	0.80	0.80	0.80
Digestible Ile	0.72	0.69	0.69	0.69	0.69	0.69
Digestible Arg	1.12	1.08	1.08	1.05	1.05	1.05
Digestible Trp	0.19	0.18	0.18	0.18	0.17	0.17
Total Lys	1.11	1.10	1.10	1.10	1.09	1.08
Total Met + Cys	0.84	0.84	0.84	0.84	0.83	0.83
Total Met	0.52	0.53	0.53	0.53	0.54	0.55
Total Thr	0.79	0.79	0.79	0.78	0.78	0.77
Total Val	0.90	0.89	0.89	0.89	0.88	0.87
Total Ile	0.80	0.77	0.77	0.77	0.76	0.75
Total Arg	1.23	1.18	1.18	1.15	1.15	1.14
Total Leu	1.67	1.63	1.63	1.61	1.56	1.48
Total His	0.51	0.51	0.51	0.48	0.47	0.43
Total Phe	0.95	0.92	0.92	0.90	0.86	0.79
Total Gly + Ser	1.71	1.69	1.69	1.62	1.55	1.42
Total Gly+Ser : Dig Lys	1.71	1.69	1.62	1.62	1.55	1.42
Ca	0.79	0.79	0.79	0.79	0.79	0.79
Non-Phytate P	0.38	0.38	0.38	0.38	0.38	0.38
Na	0.18	0.18	0.18	0.18	0.18	0.18
Analyzed Composition, %
Crude Protein	18.89	18.31	18.33	18.22	18.26	16.73
Total Lys	1.12	1.10	1.09	1.11	1.12	1.13
Total Met + Cys	0.83	0.79	0.80	0.79	0.80	0.79
Total Met	0.54	0.50	0.51	0.50	0.51	0.53
Total Thr	0.77	0.76	0.77	0.77	0.79	0.75
Total Val	0.86	0.86	0.85	0.86	0.88	0.86
Total Ile	0.77	0.76	0.75	0.75	0.77	0.72
Total Arg	1.19	1.17	1.16	1.15	1.18	1.11
Total Leu	1.59	1.60	1.59	1.57	1.59	1.44
Total His	0.48	0.47	0.47	0.47	0.46	0.41
Total Phe	0.88	0.88	0.87	0.85	0.86	0.75
Total Gly+Ser	1.66	1.63	1.61	1.60	1.60	1.40

¹Pellet durability index of diets 1 to 6 was 83.01, 83.93, 81.94, 89.17, 83.33, 81.82%, respectively.

²Sources of DL-Met, L-Lys sulphate, L-Thr, and L-Trp were MetAMINO, Biolys, and ThreAMINO, respectively (Evonik Nutrition and Care GmbH, Germany).

³Mineral premix include per kg of diet: Mn (manganese sulfate), 120 mg; Zn (zinc sulfate), 100 mg; Fe (iron sulfate monohydrate), 30 mg; Cu (tri-basic copper chloride), 8 mg; I (ethylenediamine dihydriodide), 1.4 mg; and Se (sodium selenite), 0.3 mg.

⁴Vitamin premix includes per kg of diet: Vitamin A (Vitamin A acetate), 18,739 IU; Vitamin D3 (cholecalciferol), 6,614 IU; Vitamin E (DL-alpha tocopherol acetate), 66 IU; menadione (menadione sodium bisulfate complex), 4 mg; Vitamin B12 (cyanocobalamin), 0.03 mg; folacin (folic acid), 2.6 mg: D-pantothenic acid (calcium pantothenate), 31 mg; riboflavin (riboflavin), 22 mg; niacin (niacinamide), 88 mg; thiamin (thiamin mononitrate), 5.5 mg; biotin (biotin), 0.18 mg; and pyridoxine (pyridoxine hydrochloride), 7.7 mg.

⁵Quantum Blue, AB Vista Feed Ingredients, Marlborough, UK.

Dietary CP content was determined as N × 6.25 using Leco FP-2000 (Leco Corp., St. Joseph, MI) analyzer. Diets were analyzed for total AA concentrations by ion-exchange chromatography with postcolumn derivatization with ninhydrin. Amino acids were oxidized with performic acid, which was neutralized with sodium metabisulfite (Llames and Fontaine, 1994; Commission Directive, 1998). Amino acids were liberated from the protein by hydrolysis with 6 N HCL for 24 h at 110°C. Amino acids were quantified with the internal standard method by measuring the absorption of reaction products with ninhydrin at 570 nm. Tryptophan was determined by HPLC with fluorescence detection (extinction 280 nm, emission 356 nm), after alkaline hydrolysis with barium hydroxide octahydrate for 20 h at 110°C (Commission Directive, 2000). Tyrosine was not determined.

Litter Nitrogen and Ammonia Measurements

The amount of bedding material before bird placement and the litter at the end of experimentation from each pen were weighed. The average shavings weight across all pens at day 0 was 10.93 kg +/- 0.03 and had 0.17% nitrogen content. Litter samples from each pen were collected for determination of N content. In each pen, 10 sub-samples of litter were collected at different locations and mixed to obtain a composite sample. At each interval, samples were picked up by a gloved hand and changed between treatments. The samples were subsequently stored at –20°C until analysis. Diet and litter samples were lyophilized using VirTis Genesis 25ES freeze dryer (SP Industries, Inc., Warminster, PA). Duplicate samples of 250 mg dried diet and litter from each pen were placed into an aluminum foil for N analysis (average moisture content was 41.3% +/- 3.9). The analysis was performed using combustion analyzer rapid N cube (Elementar, Hanau, Germany) according to Dumas method (method 968.06; AOAC International, 2006). Subsequently, the total N intake was calculated as the amount of N consumed per bird (g/bird) throughout the 3 growth periods, and N excretion was the amount of litter N at day 33 or expressed as a proportion of N intake (%), BW gain (g/kg) or total meat (g/kg). The average litter weight across all pens at the end of experimentation was 27.32 kg +/- 1.48 and contained nitrogen content of 2.28% +/- 0.23.

Ammonia volatilization from the litter was assessed by measuring equilibrium ammonia concentrations (Gates et al., 1997) with a dynamic flux chamber (Woodbury et al., 2006) placed on the litter on d 33 after birds were removed from the pens and allowed to stabilize for 10 min before taking readings. The dynamic flux chamber method was used to measure equilibrium ammonia concentrations, as all pens shared a common airspace and emissions could not be measured directly. All measurements were taken near the center of each pen to avoid areas near the drinker line and feeder that typically have increased litter moisture and may increase data variability.

Carcass Characteristics

On 33 d of age, feed and water were removed for 12 h prior to processing. On 34 d of age, 10 birds were randomly selected per pen to assess carcass characteristics. Birds were electrically stunned, euthanized by exsanguination, and processed in the Auburn University Poultry Science Department Pilot Processing Plant. Stunning was accomplished by means of immersion of the bird's head and neck in a water bath designed for poultry processing that is electrified with a minimum current of 120 mA. Carcasses were chilled in ice water for a period of 3 h and then drained of excess water for approximately 3 min. Carcass (without abdominal fat), pectoralis major (breast fillets) and pectoralis minor (tenders), wings, thighs, drumsticks, and abdominal fat pad weights were determined after processing to assess yield per bird. Tender and breast fillet weights were combined for the analysis of total breast meat weight. Meat yield percentages were calculated relative to the 33-d live weight.

Statistical Analyses

Dietary treatments were randomized within each block with pen location as the blocking factor. Each treatment comprised 10 replicate pens with pen serving as experimental unit. Analysis of variance was performed using the MIXED procedure of SAS Institute (2017) by the following mixed-effects model:

$$Y_{ij}=\mu ..+{\rho }_i+{\tau }_j+{\epsilon }_{ij}$$

Where $\mu ..$ is the overall mean; the ${\rho }_i$ are identically and independently normally distributed random block effects with mean 0 and variance ${{\sigma }^2}_{\rho }$; the ${\tau }_j$ are fixed factor level effects corresponding to the j^th dietary treatment (Diets 1–6) such that $\sum {\tau }_j$ = 0; and the ${\epsilon }_{ij}$ are identically and independently normally distributed random errors with mean 0 and a variance ${\sigma }^2.$ Mortality data were arcsine transformed before analysis. Preplanned orthogonal contrasts were used to detect differences on growth performance, carcass characteristics, ammonia concentrations, and N balance between each dietary treatment and the control diet (Diet 1). In addition, means were separated for differences determination by Tukey's HSD. Statistical significance was considered at P ≤ 0.05.

RESULTS AND DISCUSSION

Growth Performance

Broilers fed reduced-CP diets with L-Arg (21.2 % CP) or L-Arg and L-Trp (19.8% CP) had decreased (P < 0.01) BW gain compared with birds fed the control diet during the initial 14 d (Table 4). Birds fed reduced-CP diets supplemented with L-Val (22.3% CP) or, L-Ile to L-Trp (Diets 4–6; 21.9–19.8% CP) had increased (P < 0.05) FCR compared with control-fed birds. Dietary treatment influenced BW gain (P = 0.03) from 15 to 25 d of age (Table 5). Birds fed reduced-CP diets supplemented with L-Val (Diet 2) had increased BW gain (P < 0.05) compared with L-Val to Trp (Diet 6). However, there was no difference observed between the control and other treatments. In addition, dietary treatments influenced FCR (P < 0.001) from 15 to 25 d of age. Relative to the control diet, FCR decreased (P = 0.03) with L-Val addition (Diet 2, 19.95% CP) but FCR increased (P < 0.001) with L-Val to L-Trp supplementation (Diet 6, 17.31% CP). The FCR of birds fed Diets 3 to 5 was similar to the control at day 15 to 25.

Table 4.

Growth performance characteristics of Ross × Ross 708 male broilers fed reduced crude protein diets while maintaining essential amino acid concentrations from 1 to 14 d of age.¹

Dietary treatments	Calculated CP, %	BW, kg	BW gain, kg	Feed intake, kg	FCR2, kg/kg	Mortality, %
1) Control (DL-Met, L-Lys, L-Thr)	23.23	0.465a	0.426a	0.505	1.186c	0.8
2) Control + L-Val	22.31	0.459ab	0.420ab	0.506	1.204b	0.4
3) Control + L-Val, Gly	22.44	0.457ab	0.418ab	0.501	1.198bc	0.8
4) Control + L-Val, Gly, L-Ile	21.95	0.454ab	0.415ab	0.502	1.208ab	0.4
5) Control + L-Val, Gly, L-Ile, L-Arg	21.23	0.444b	0.406b	0.495	1.221ab	2.4
6) Control + L-Val, Gly, L-Ile, L-Arg, L-Trp	19.81	0.441b	0.402b	0.494	1.231a	0.0
Polled standard error		0.005	0.005	0.005	0.007	0.6
Preplanned orthogonal contrasts				Probabilities
Treatment 1 vs. 2		0.43	0.40	0.91	0.044	0.58
Treatment 1 vs. 3		0.25	0.26	0.57	0.15	1.00
Treatment 1 vs. 4		0.10	0.13	0.65	0.012	0.58
Treatment 1 vs. 5		0.004	0.004	0.18	< 0.001	0.12
Treatment 1 vs. 6		< 0.001	< 0.001	0.13	< 0.001	0.28
Diet		0.006	0.006	0.45	< 0.001	0.156

¹Values represent least-square means for 10 replicate pens.

²Corrected for mortality.

^a-cMeans within columns with different superscripts are significant (P < 0.05).

Table 5.

Growth performance characteristics of Ross × Ross 708 male broilers fed reduced crude protein diets while maintaining essential amino acid concentrations from 15 to 25 d of age.¹

Dietary treatment	Calculated CP2, %	Day 25 BW, kg	BW gain, kg	Feed intake, kg	FCR3, kg/kg	Mortality, %
1) Control (DL-Met, L-Lys, L-Thr)	23.23, 20.69	1.282ab	0.817ab	1.172	1.436b	0.4
2) Control + L-Val	22.31, 19.95	1.320a	0.861a	1.200	1.394c	0.4
3) Control + L-Val, Gly4	22.44, 19.95	1.294ab	0.837ab	1.183	1.414bc	0.0
4) Control + L-Val, Gly, L-Ile4	21.95, 19.52	1.303ab	0.850ab	1.192	1.404bc	0.8
5) Control + L-Val, Gly, L-Ile, L-Arg4	21.23, 18.95	1.262ab	0.818ab	1.150	1.405bc	0.0
6) Control + L-Val, Gly, L-Ile, L-Arg, L-Trp4	19.81, 17.31	1.242b	0.802b	1.170	1.460a	0.4
Pooled standard error		0.016	0.014	0.018	0.009	0.5
Preplanned orthogonal contrasts				Probabilities
Treatment 1 vs. 2		0.11	0.11	0.33	0.030	0.98
Treatment 1 vs. 3		0.61	0.61	0.79	0.39	0.44
Treatment 1 vs. 4		0.36	0.35	0.57	0.21	0.77
Treatment 1 vs. 5		0.40	0.41	0.29	0.54	0.42
Treatment 1 vs. 6		0.09	0.09	0.67	< 0.001	0.99
Diet		0.02	0.03	0.41	< 0.001	0.82

¹Values represent least-square means for 10 replicate pens.

²Calculated CP in the starter (1–14 d of age) and grower (15–25 d of age) periods, respectively.

³Corrected for mortality.

⁴Glycine was added to diets only from day 1 to 14.

^a-cMeans within columns with different superscripts are significant (P < 0.05).

Cumulatively, feeding reduced-CP diets affected BW gain (P = 0.02) and FCR (P < 0.001) from d 1 to 25 (Table 6). However, compared to the control, there was no difference observed in birds fed reduced-CP diets for BW gain. Interestingly, birds fed reduced-CP diets supplemented with L-Val (Diet 2) had increased BW gain (P < 0.05) compared to diets with L-Val to Trp (Diet 6). On the other hand, FCR decreased (P = 0.03) with L-Val addition (Diet 2), remained unchanged from Diet 3 to 5 but increased (P < 0.001) with L-Val to L-Trp supplementation (Diet 6), relative to the control diet. At d 26 to 33 (Table 7), BW gain improved (P = 0.045) in birds fed reduced-CP diet supplemented with L-Val, relative to the control. In addition, birds fed reduced-CP diet with L-Val to L-Ile (Diet 4) had a 0.8% greater mortality rate (P = 0.03) compared with the control-fed birds. Cumulative final BW, BW gain and FCR (1–33 d of age, Table 8) improved (P < 0.05) in birds fed reduced-CP diet supplemented with L-Val, relative to the control or control with added L-Val to L-Trp (Diet 6). Conversely, birds fed reduced-CP diets containing Gly, L-Ile and L-Arg (Diets 3 to 5) had comparable cumulative BW gain and FCR (1–33 d of age) with the control-fed birds. Dietary treatments did not influence feed intake or the incidence of mortality throughout the experimental period (1–33 d of age). It is important to note that mortality was relatively low, remaining below 3% throughout the study duration.

Table 6.

Growth performance characteristics of Ross × Ross 708 male broilers fed reduced crude protein diets while maintaining essential amino acid concentrations from 1 to 25 d of age.¹

Dietary treatment	Calculated CP2, %	Day 25 BW, kg	BW gain, kg	Feed intake, kg	FCR3, kg/kg	Mortality, %
1) Control (DL-Met, L-Lys, L-Thr)	23.23, 20.69	1.282ab	1.243ab	1.676	1.349b	1.2
2) Control + L-Val	22.31, 19.95	1.320a	1.282a	1.705	1.331c	0.8
3) Control + L-Val, Gly4	22.44, 19.95	1.294ab	1.255ab	1.685	1.342bc	0.4
4) Control + L-Val, Gly, L-Ile4	21.95, 19.52	1.303ab	1.265ab	1.693	1.339bc	1.2
5) Control + L-Val, Gly, L-Ile, L-Arg4	21.23, 18.95	1.262ab	1.224ab	1.645	1.344bc	1.8
6) Control + L-Val, Gly, L-Ile, L-Arg, L-Trp4	19.81, 17.31	1.242b	1.204b	1.664	1.380a	1.2
Pooled standard error		0.016	0.016	0.020	0.006	0.8
Preplanned orthogonal contrasts				Probabilities
Treatment 1 vs. 2		0.11	0.11	0.33	0.030	0.48
Treatment 1 vs. 3		0.61	0.61	0.79	0.39	0.38
Treatment 1 vs. 4		0.36	0.35	0.57	0.21	0.79
Treatment 1 vs. 5		0.40	0.41	0.29	0.54	0.77
Treatment 1 vs. 6		0.09	0.09	0.67	< 0.001	0.79
Diet		0.02	0.02	0.36	< 0.001	0.85

¹Values represent least-square means for 10 replicate pens.

²Calculated CP in the starter (1–14 d of age) and grower (15–25 d of age) periods, respectively.

³Corrected for mortality.

⁴Glycine was added to diets only from day 1 to 14.

^a-cMeans within columns with different superscripts are significant (P < 0.05).

Table 7.

Growth performance characteristics of Ross × Ross 708 male broilers fed reduced crude protein diets while maintaining essential amino acid concentrations from 26 to 33 d of age.¹

Dietary treatment	Calculated CP2, %	Day 33 BW, kg	BW Gain, kg	Feed intake, kg	FCR3, kg/kg	Mortality, %
1) Control (DL-Met, L-Lys, L-Thr)	23.23, 20.69, 19.13	2.020b	0.739b	1.154	1.562	0.0
2) Control + L-Val	22.31, 19.95, 18.58	2.091a	0.771a	1.189	1.543	0.0
3) Control + L-Val, Gly4	22.44, 19.95, 18.58	2.063ab	0.770ab	1.178	1.532	0.0
4) Control + L-Val, Gly, L-Ile4	21.95, 19.52, 18.28	2.072ab	0.768ab	1.196	1.559	0.8
5) Control + L-Val, Gly, L-Ile, L-Arg4	21.23, 18.95, 17.80	2.017bc	0.755ab	1.176	1.559	0.0
6) Control + L-Val, Gly, L-Ile, L-Arg, L-Trp4	19.81, 17.31, 16.87	1.984c	0.742ab	1.159	1.562	0.0
Pooled standard error		0.022	0.011	0.013	0.016	0.2
Preplanned orthogonal contrasts				Probabilities
Treatment 1 vs. 2		0.033	0.045	0.14	0.41	0.83
Treatment 1 vs. 3		0.19	0.054	0.44	0.20	0.83
Treatment 1 vs. 4		0.12	0.07	0.17	0.90	0.03
Treatment 1 vs. 5		0.92	0.31	0.87	0.88	0.79
Treatment 1 vs. 6		0.27	0.83	0.89	0.99	0.83
Diet		0.01	0.14	0.11	0.69	0.08

¹Values represent least-square means for 10 replicate pens.

²Calculated CP in the starter (1–14 d of age), grower (15–25 d of age), and finisher (26–33 d of age) periods, respectively.

³Corrected for mortality.

⁴Glycine was added to diets only from day 1 to 14.

^a-cMeans within columns with different superscripts are significant (P < 0.05).

Table 8.

Growth performance characteristics of Ross × Ross 708 male broilers fed reduced crude protein diets while maintaining essential amino acid concentrations from 1 to 33 d of age.¹

Dietary treatment	Calculated CP2, %	Day 33 BW, kg	BW Gain, kg	Feed intake, kg	FCR3, kg/kg	Mortality, %
1) Control (DL-Met, L-Lys, L-Thr)	23.23, 20.69, 19.13	2.020b	1.982b	2.829	1.428b	1.2
2) Control + L-Val	22.31, 19.95, 18.58	2.091a	2.052a	2.895	1.411c	0.8
3) Control + L-Val, Gly4	22.44, 19.95, 18.58	2.063ab	2.025ab	2.862	1.414bc	0.4
4) Control + L-Val, Gly, L-Ile4	21.95, 19.52, 18.28	2.072ab	2.033ab	2.889	1.421bc	2.0
5) Control + L-Val, Gly, L-Ile, L-Arg4	21.23, 18.95, 17.80	2.017bc	1.979bc	2.821	1.426b	2.8
6) Control + L-Val, Gly, L-Ile, L-Arg, L-Trp4	19.81, 17.31, 16.87	1.984c	1.946c	2.822	1.451a	1.2
Pooled standard error		0.022	0.022	0.030	0.005	0.9
Preplanned orthogonal contrasts				Probabilities
Treatment 1 vs. 2		0.033	0.033	0.14	0.028	0.49
Treatment 1 vs. 3		0.19	0.19	0.44	0.07	0.39
Treatment 1 vs. 4		0.12	0.12	0.17	0.37	0.85
Treatment 1 vs. 5		0.92	0.93	0.87	0.79	0.46
Treatment 1 vs. 6		0.27	0.27	0.89	0.004	0.75
Diet		0.01	0.01	0.30	< 0.001	0.59

¹Values represent least-square means for 10 replicate pens.

²Calculated CP in the starter (1–14 d of age), grower (15–25 d of age), and finisher (26–33 d of age) periods, respectively.

³Corrected for mortality.

⁴Glycine was added to diets only from day 1 to 14.

^a-cMeans within columns with different superscripts are significant (P < 0.05).

The findings from the current study align with previous research indicating that feeding broiler chickens diets with approximately 2 to 3 percentage point reductions in CP has either none or negligible effects on final bird performance parameters (Belloir et al., 2017; Kriseldi et al., 2018; Lambert et al., 2022). However, it is noteworthy that FCR deteriorated only by 2 percentage points in birds fed diets with more than a 3-percentage point reduction in CP (Diet 6; during the 33-d cumulative period). This observation agrees with previous studies (Si et al., 2004; Waldroup et al., 2005, Chrystal et al., 2020) emphasizing that free AA may not completely substitute for CP in diets, underscoring the concept of less limiting essential amino acids and non-essential nitrogen significantly affects performance. Additionally, these present data provide further substantiation for the importance of maintaining adequate Gly concentration during the starter period in birds consuming reduced CP diets to mitigate impaired growth performance. Previous research reported that Gly may be essential early in life and nonessential later in life (Dean et al., 2006; Kriseldi et al., 2018). Supplemental Gly was incorporated to reduced CP diets during the starter period, and this improved growth performance (in Diets 3–5; <3% CP reduction) similar to control fed birds and the response carried through to 33 d of age. Siegert and Rodehutscord (2019) proposed that Gly is likely to become limiting in low-CP plant-based diets and a deficiency may impair broiler performance. Other authors have indicated that the positive response of Gly on growth performance may be attributed to its role in biological processes such as feather development, creatine formation, mucin synthesis, enterocyte development and bone formation (Stilborn et al., 1997; Yuan et al., 2012; Ospina-Rojas et al., 2013; Wang et al., 2014).

Carcass Characteristics

Feeding broiler chickens with reduced CP diets did not alter the weights or yields of carcasses, total breast meat, drums, or thighs when compared to the control fed group (Table 9). However, birds fed reduced CP diets containing L-Trp (Diet 6) exhibited decreased (P = 0.01) wing yield in comparison with the control. There was increased wing weight in birds fed reduced CP diets with supplemental L-Val to L-Ile (P < 0.05) in Diet 4 relative to diets with L-Val to L-Trp (Diet 6). Birds fed reduced CP diets with supplemental L-Val (Diet 2) or L-Val to L-Arg and L-Trp (Diets 5 and 6, with Gly during the starter period) had increased (P < 0.01) abdominal fat percentage when compared with the control. Conversely, birds fed reduced CP diets with L-Val addition and supplemental Gly during the starter period (Diet 3) showed no significant change in abdominal fat percentage when compared with the control, or diets with added L-Val to L-Ile or L-Val to L-Arg (Diet 4 and 5, respectively).

Table 9.

Carcass characteristics of Ross × Ross 708 male broilers fed reduced crude protein diets while maintaining essential amino acid concentrations at 33 d of age.¹

Dietary treatments	Carcass		Total breast		Wings		Drums		Thigh		Abdominal fat
	Weight, g	Yield, %	Weight, g	Yield, %	Weight, g	Yield, %	Weight, g	Yield, %	Weight, g	Yield, %	Weight, g	Yield, %
1) Control (DL-Met, L-Lys, L-Thr)	1,521	72.35	493	23.42	165ab	7.85a	195	9.29	204	9.69	24c	1.15c
2) Control + L-Val	1,574	72.41	516	23.72	169ab	7.76ab	198	9.12	204	9.53	27ab	1.26ab
3) Control + L-Val, Gly2	1,563	72.49	517	23.95	169ab	7.82ab	197	9.12	205	9.51	25bc	1.14c
4) Control + L-Val, Gly, L-Ile2	1,574	72.97	516	23.90	170a	7.81ab	199	9.11	207	9.61	26ab	1.21bc
5) Control + L-Val, Gly, L-Ile, L-Arg2	1,537	72.82	496	23.46	165ab	7.81ab	195	9.26	207	9.78	28a	1.31ab
6) Control + L-Val, Gly, L-Ile, L-Arg, L-Trp2	1,494	72.23	481	23.11	161b	7.73b	191	9.19	195	9.38	28a	1.37a
Pooled standard error	23	0.29	10	0.23	2	0.05	2	0.07	4	0.13	1	0.04
Preplanned orthogonal contrasts	Probabilities
Treatment 1 vs. 2	0.09	0.88	0.13	0.37	0.16	0.22	0.31	0.09	0.99	0.35	0.001	0.008
Treatment 1 vs. 3	0.16	0.74	0.11	0.11	0.16	0.66	0.64	0.10	0.84	0.28	0.41	0.88
Treatment 1 vs. 4	0.08	0.14	0.12	0.15	0.051	0.55	0.19	0.08	0.50	0.62	0.033	0.13
Treatment 1 vs. 5	0.59	0.23	0.83	0.90	0.87	0.55	1.00	0.75	0.61	0.59	< 0.001	< 0.002
Treatment 1 vs. 6	0.38	0.77	0.42	0.34	0.08	0.010	0.15	0.32	0.12	0.07	< 0.001	< 0.001
Diet	0.07	0.44	0.08	0.11	0.01	0.62	0.11	0.32	0.27	0.23	< 0.001	< 0.001

¹Values represent least-square means of 10 replicate pens with 10 birds per pen selected for processing at day 33.

²Glycine was added to diets only from day 1 to 14.

^a-cMeans within columns with different superscripts are significant (P < 0.05).

In the current study, a reduction in dietary CP of approximately 3% points did not result in reduced weight or yields of total carcass, breast meat, drums, and thighs. This finding agrees with other reports (Belloir et al., 2017; van Harn et al., 2019; Brink et al., 2022; Lambert et al., 2022) showing that reduced CP diets have no negative consequences on carcass yield as long as essential AA requirements are maintained. Feed intake was unchanged in response to reduced CP content. Therefore, the observed effects on carcass yield in birds fed reduced CP diets, relative to the control, cannot be attributed to differences in energy or essential amino acid intake. However, birds fed L-Trp supplemented diet (Diet 6) had a decrease (–1.5%) in wing yield compared with the control-fed birds, which is consistent with a previous report (van Harn et al., 2019). Ospina-Rojas et al. (2014) reported that it is important that Gly and Arg are added to low protein diets, accompanied by the supplementation of Val and Ile, besides Lys, Met and Thr to maintain carcass yields. In the present research, the contents of essential AA, including Gly, of the reduced CP diets met amino acid recommendations (Rostagno, 2017).

Abdominal fat deposition increased with lower dietary CP concentrations (Bregendahl et al., 2002; Belloir et al., 2017; Chrystal et al., 2020), presumably due to impaired muscle deposition, leading to excess energy deposited as abdominal fat (Namroud et al., 2008; Belloir et al., 2017). In our study, abdominal fat increased by 8.7% in birds fed L-Val supplemented, reduced-CP diet relative to the control. Dietary Val is an important regulator of muscle protein synthesis (Yoshizawa, 2004; Baracos and Mackenzie, 2006). Although breast yield was unaffected by Val addition in the present study or in any other dietary treatments, increased abdominal fat seems more attributable to non-essential nitrogen (Collin et al., 2003; Namroud et al., 2008; Yalçin et al., 2010; Kriseldi et al., 2017) and not a direct consequence of supplemental Val (Tavernari et al., 2013; Ospina-Rojas et al., 2017). However, Gly addition during the starter phase resulted in a significant decrease in abdominal fat at slaughter age in L-Val and L-Val with L-Ile supplemented reduced-CP diets (–9.5 and -4.9%, respectively), restoring both diets to a level comparable to the control. This response may be partly explained by Gly being a precursor for creatine synthesis (Smith, 1968; Miller and Kifer, 1970), which is the main source for energy in the muscle, favoring more meat yield relative to fat. However, further CP reduction of approximately 1.3 to 2.6 percentage points (Diets 5 and 6) increased abdominal fat yield (+12% in L-Arg and +16% in L-Trp supplemented diets) at slaughter age compared to the control fed birds, regardless of Gly addition during the starter phase. Hence, further investigation is warranted to mitigate abdominal fat deposition when broilers are fed lower CP diets.

Nitrogen Balance

The amount of excreted N, expressed in various metrics including g/bird, as a proportion of intake (%), g/kg BW gain or g/kg total meat yield was not different (Table 10) between birds fed the control diet and reduced CP diets containing L- Val to L-Arg (Diets 2–5). However, birds fed reduced-CP diet containing L-Trp (Diet 6) had decreased N excretion (g/kg BW gain and total meat yield; P < 0.05) relative to other treatments. Additionally, birds fed reduced-CP diets with added L-Val to L-Arg (Diets 2–5) showed no significant change in litter ammonia concentration relative to the control-fed birds. Conversely, birds fed reduced-CP diet with L-Trp addition (Diet 6) resulted in a 48% decrease (P = 0.041) in litter ammonia volatilization relative to the control.

Table 10.

Nitrogen intake, excretion, and litter ammonia concentrations of Ross × Ross 708 male broilers fed reduced crude protein diets while maintaining amino acid concentrations from 1 to 33 d of age.¹

Dietary treatments	Calculated CP2, %	N Intake, g/bird	N excretion				NH3, mg/kg5
			g/bird3	% Intake	g/kg BW gain	g/kg total meat4
1) Control (DL-Met, L-Lys, L-Thr)	23.23, 20.69, 19.13	79.22a	21.42a	27.00	10.87a	20.24a	93.68a
2) Control + L-Val	22.31, 19.95, 18.58	81.47a	21.70a	26.61	10.57ab	19.94ab	86.25ab
3) Control + L-Val, Gly6	22.44, 19.95, 18.58	79.40a	20.86ab	26.30	10.31ab	19.22ab	76.42ab
4) Control + L-Val, Gly, L-Ile6	21.95, 19.52, 18.28	80.36a	21.23ab	26.54	10.47ab	19.69ab	70.94ab
5) Control + L-Val, Gly, L-Ile, L-Arg6	21.23, 18.95, 17.80	76.81b	20.31ab	26.38	10.24ab	19.06ab	56.35ab
6) Control + L-Val, Gly, L-Ile, L-Arg, L-Trp6	19.81, 17.31, 16.87	70.21c	18.00b	25.68	9.27c	17.51c	48.85b
Pooled standard error		0.83	1.04	1.25	0.48	0.86	16.15
Preplanned orthogonal contrast				Probabilities
Treatment 1 vs. 2		0.06	0.80	0.77	0.55	0.75	0.73
Treatment 1 vs. 3		0.88	0.61	0.60	0.27	0.27	0.42
Treatment 1 vs. 4		0.34	0.87	0.74	0.47	0.58	0.29
Treatment 1 vs. 5		0.044	0.31	0.64	0.22	0.21	0.09
Treatment 1 vs. 6		< 0.001	0.003	0.32	0.003	0.005	0.041
Diet		< 0.001	0.02	0.95	0.06	0.07	0.34

¹Values represent least-square means for 10 replicate pens.

²Calculated CP content in the starter (1–14 d of age), grower (15–25 d of age), and finisher (26–33 d of age) diets, respectively.

³Litter N concentration at the conclusion of the experiment (day 33) was subtracted by litter N concentration before bird placement (day 0).

⁴Total meat includes the sum of wings, drums, thighs, and total breast meat.

⁵Values were determined using temperature as a covariate. Temperature was included as a covariate to account for any ammonia concentration variation as ammonia is temperature dependent and there was some variation in the room temperature while equilibrium concentrations were recorded.

⁶Glycine was added to diets only from day 1 to 14.

^a-cMeans within columns with different superscripts are significant (P < 0.05).

The current study demonstrated that reducing dietary CP by approximately 3 percentage points led to a notable reduction in excreted N and litter ammonia concentration, resulting in an enhanced N retention efficiency. This observation aligns with previous findings (Ferguson et al.,1998; Kamran et al., 2010; Ospina-Rojas et al., 2012; Belloir et al., 2017; de Rauglaudre et al., 2023). Importantly, the reduction in N excretion and litter ammonia volatilization did not compromise breast meat weight or yield. When expressed relative to gain or carcass yield, this reduction amounted to approximately 14% reduction in N excreted per kilogram of BW gain and kilogram of total meat produced. This finding holds significant importance because previous reports in literature typically provide a reduction in N excretion without establishing a relationship to BW gain or total meat produced. Evaluating the impact of reducing excess nitrogen excretion per unit of meat produced is vital for ensuring the sustainability of reduced-protein diet utilization in modern poultry operations. Furthermore, it is worth noting that the reduction in dietary CP content had a stronger effect on reducing ammonia volatilization per percentage point CP reduction compared to N excretion (19.8% vs. 7%), corroborating previous findings (Lemme et al., 2019).

Collectively, this study demonstrates that dietary CP can be reduced by approximately 2.0 percentage points beyond L-Thr inclusion without adversely affecting growth performance and meat yield of broilers when essential amino acid concentrations are maintained. Reducing dietary CP content can lower nitrogenous excretion and emissions without negative impacting meat yield and potentially lowering dietary cost with the use of supplemental amino acids. Future research should evaluate broilers fed reduced CP diets for broilers raised to 4.0 kg.