Feeding a reduced protein diet with a near ideal amino acid profile improves amino acid efficiency and nitrogen utilization for milk production in sows

Abstract

Fifty-four lactating multiparous Yorkshire sows were used to test the hypothesis that feeding a reduced protein diet with a near ideal AA (NIAA) profile increases the biological utilization efficiency of N and essential AA (EAA) for milk production in part as a result of reduced dietary Leu concentration. Sows were fed 1 of 3 isocaloric diets containing the following concentration of CP (% as-fed, analyzed): 18.74 (Control: CON), 13.78 (Optimal: OPT), and 14.25 (Optimal + Leu: OPTLEU). The OPT and OPTLEU diets contained the same concentration of crystalline AA (CAA) to meet requirements of the limiting AA. Crystalline Leu was added to OPTLEU to contain the same SID Leu concentration as that of CON. Sows were weighed on days 1 and 21 of lactation and piglets on days 1, 4, 8, 14, 18, and 21 of lactation. Nitrogen retention was measured for 48 or 72 h between days 4 and 8 (early) and days 14 and 18 (peak) of lactation. Sow BW change and ADFI did not differ between diets. Litter growth rate (LGR) during early lactation did not differ between diets. At peak lactation, LGR was higher in sows fed OPT compared with CON (P < 0.05) and lower in sows fed OPTLEU compared with OPT (P < 0.05). In early and peak lactation, total N retention, and milk N output efficiency were greater in OPT (P < 0.01) and OPTLEU (P < 0.05) than CON. Compared with CON, overall biological efficiency of N, Arg, His, Ile, Leu, Phe, and Trp were greater (P < 0.05), whereas those of Lys, Met, Thr, and Val did not differ in sows fed OPT and OPTLEU, except for Leu which did not differ between OPTLEU and CON. Compared with OPT, only Leu and Met efficiency were lower (P < 0.01) and tended to be lower (P = 0.10), respectively, in sows fed OPTLEU. Reducing CP with a NIAA profile to attain the minimum Leu requirement maintained overall lactation performance, improved utilization efficiency of N, Arg, His, Ile, Leu, Phe + Tyr, and Trp for milk production, and maximized efficiency of Ile, Leu, Lys, Met + Cys, Phe + Tyr, Thr, Trp, and Val. Addition of Leu did not reduce N and EAA utilization efficiency. This study provides revised and novel maximum biological efficiency value (MBEV) for Ile (65.4), Leu (75.1), Lys (63.2), Met + Cys (78.2), Phe + Tyr (69.5), Thr (71.0), Trp (70.1), and Val (57.0). These MBEV can be used to more accurately predict the requirement for those AA during lactation.

INTRODUCTION

The breeding herd contributes to as much as 11.8 × 10⁶ metric tons of fresh manure produced annually in the United States (Koelsch et al., 2005). Therefore, small change in the efficiency of dietary N utilization in lactating sows can have major impacts on N excretion at the global scale. Determination of individual essential AA (EAA) biological efficiency value at near maximal biological potential is needed to accurately predict the requirement of each EAA. Underestimation of efficiency leads to overestimation of requirement and increase N losses to the environment. Except for Lys, maximum biological efficiency value (MBEV) of individual EAA reported by NRC (2012) was not empirically determined, nor has been validated. Furthermore, it is unclear why feeding individual EAA at or near minimum requirement in a low CP diet improves efficiency. It may be due to reduction in intake of the said EAA alone or in competitive inhibition with other AA present in excess of requirements. Previous work from the same lab (Guan et al., 2004; Manjarín et al., 2012) suggested that there is competition among AA, in particular between Leu and Lys utilization for milk production. Thus, Lys utilization even when present at its minimum requirement may not be maximized in the presence of excessive concentration of N or other specific EAA.

We hypothesized that reducing CP to meet the minimum SID Leu requirement increases efficiency of individual EAA. We further hypothesized that the relatively high Leu:Lys in a conventional corn-soybean meal-based diet (1.63:1) compared with that in a reduced CP diet (1.14:1) meeting minimum SID Leu requirement reduces Lys efficiency for milk protein production. The objectives were to 1) estimate MBEV of EAA in lactating sows fed a diet meeting the minimum SID requirement for Leu (NRC, 2012) and 2) determine whether the corresponding decrease in Leu concentration in reduced CP diet affects Lys efficiency.

MATERIALS AND METHODS

The experimental protocol was approved by the Michigan State University Institutional Animal Care and Use Committee (AUF # 05/16-091-00) and followed the American Association for Laboratory Animal Science guidelines.

Animals, Feeding, and Experimental Design

The study was conducted at the Michigan State University Swine Teaching and Research Center, using 54 purebred multiparous (parity 2+) Yorkshire sows. Sows were moved to conventional farrowing crates between days 105 and 107 of gestation, grouped by parity, and randomly assigned to 1 of 3 dietary treatments within parity groups (Control, n = 18; Optimal, n = 19; Optimal + Leu, n = 17). The study was conducted over 4 blocks of time, with 12 to 18 sows per block. Litters were standardized to 11 piglets within the first 24 h after farrowing with the aim of weaning 10 piglets per sow. Sows were adapted to the experimental diets (2.2 kg/d) 4 to 6 d before the expected farrowing date. After farrowing, sows feed allowance was progressively increased from 1.88 kg/d at day 1 to 7.44 kg/d at day 21 of lactation, according to the NRC (2012) model, with targeted ADFI of 6.0 kg/d during the whole lactation period. Feed was provided daily in 3 equal meals (0700, 1300, and 1900) with feed intake and refusal recorded daily before the morning meal. Water was freely accessible to sows and piglets. Injection of iron and surgical castration were conducted on days 1 and 7, respectively. No creep feed was supplied to the piglets.

Sows and piglets were weighed on day 1 (i.e., 24 h postpartum) after standardization of litter size and 21. Sow BW was only recorded on days 1 and 21 due to high variability and labor intensive between short period of time. Sow back fat thickness was measured (Lean-meater, series 12, Renco Corp., Golden Valley, MN) on days 1 and 21. Corn oil was applied as an ultrasound enhancing agent, and the probe was placed perpendicularly on the back 6 to 8 cm from the midline at the last rib. Two separate measurements were taken on each side of the midline and averaged. Litters were also weighed on days 4, 8, 14, and 18 of lactation to estimate milk yield (Theil et al., 2002) between days 4 and 8 and days 14 and 18, representing early and peak lactation periods, respectively.

Dietary Treatment

Ingredient and calculated nutrient composition of the diets are presented in Table 1. Analyzed total (hydrolysate) and free AA of the diets are presented in Table 2. The NRC (2012) model was used to estimate requirements for AA, NE, Ca, and P for sows. The requirements were based on the swine herd performance at the Michigan State University Swine Teaching and Research Center, including sow BW of 210 kg, sow parity number of 2 and above, sow ADFI of 6 kg/d, litter size of 10, piglet BW gain of 280 g/d over a 21-d lactation period, and ambient temperature of 20 °C. The model predicted a minimum sow BW loss of 7.5 kg and the protein: lipid was adjusted to the minimum allowable value of near zero. All diets were formulated to contain the same SID Lys (0.90%) and NE (2,580 kcal/kg) concentrations. The control diet (CON) was formulated using corn and soybean meal as the only sources of Lys to meet NRC (2012) SID Lys requirement (0.90%) and consequently contained 18.74% CP. Valine met near SID requirement (NRC, 2012; 0.77% vs. 0.79%). All other EAA SID concentrations were in excess relative to NRC (2012). A second diet balanced to reach a near ideal AA (NIAA) profile was formulated. In this article, we chose to the term “near ideal AA profile” in lieu of the conventional “ideal AA profile” because the “ideal AA profile” is conceptual rather than biologically factual. Our rationale is further based on the notions that an “ideal AA profile” 1) cannot be limited to the relative contribution of only 2 AA pools (i.e., milk and maintenance), 2) needs an accurate characterization of the maintenance AA pool for the lactating sow, and 3) should include AA for which dietary essentiality in known lactating sows (i.e., Arg and His). The NIAA diet was designed by reducing soybean meal relative to corn to meet the minimum SID Leu requirement, which corresponded to a CP concentration of 13.78%. Then, supplemental crystalline source of l-Lys, l-Val, l-Thr, l-Phe, dl-Met, l-Ile, l-His, l-Trp, and l-Leu were added to meet the minimum SID requirement for those AA in the NIAA diet. dl-Met was added to meet the requirement of Met + Cys. This diet is referred to as the optimal diet (OPT) throughout the remainder of the manuscript. A third diet was formulated to be the same as OPT with added crystalline l-Leu to equate the SID Leu concentration of CON and referred to as optimal + Leu diet (OPTLEU). Sugar food product (International Ingredient Corporation, St. Louis, MO) was included in all 3 diets at 5% to increase diet palatability. Titanium dioxide was included at 0.10% as indigestible marker in all experimental diets.

Table 1.

Ingredient composition and nutrient content of experimental diets (as-fed)

	Control	Optimal	Optimal + Leu
Ingredient composition, %
Corn, yellow dent	59.17	61.45	61.21
Soybean meal, 48% CP	30.00	14.00	14.00
Soy hulls	0	10.57	10.57
Sugar food product1	5.00	5.00	5.00
Beef tallow	3.35	5.02	4.81
l-Lys·HCl	0	0.47	0.47
l-Val	0	0.29	0.29
l-Thr	0	0.20	0.20
l-Phe	0	0.13	0.13
dl-Met	0	0.11	0.11
l-Ile	0	0.08	0.08
l-His	0	0.07	0.07
l-Trp	0	0.05	0.05
l-Leu	0	0	0.45
Limestone	1.18	0.93	0.93
Dicalcium phosphate	0.45	0.78	0.78
Sodium chloride	0.50	0.50	0.50
Vitamin and mineral premix2	0.25	0.25	0.25
Titanium dioxide	0.10	0.10	0.10
Total	100.00	100.00	100.00
Calculated nutrient concentration3
NE, kcal/kg	2,580	2,580	2,580
CP, %	19.24	14.00	14.34
Fermentable fiber, %	11.58	11.58	11.57
SID AA, %4
Arg	1.17	0.71	0.71
His	0.47	0.37	0.37
Ile	0.71	0.52	0.52
Leu	1.47	1.03	1.47
Lys	0.90	0.90	0.90
Met5	0.27	0.30	0.30
Met + Cys	0.54	0.49	0.49
Phe	0.84	0.67	0.67
Phe + Tyr	1.38	1.03	1.03
Thr	0.61	0.58	0.58
Trp	0.21	0.17	0.17
Val	0.77	0.79	0.79
N	2.63	1.88	1.93
Total Ca, %6	0.65	0.65	0.65
STTD P, %6	0.23	0.23	0.23

¹Supplied per kg: NE 2,842 kcal; fermentable fiber 0.05%; CP 1.00% (International Ingredient Corporation, St. Louis, MO).

²Sow micro 5 and Se-yeast PIDX15 (Provimi North America, Inc., Brookville, OH).

³Based on nutrient concentrations in feed ingredients according to NRC (2012).

⁴SID = standardized ileal digestible (NRC, 2012).

⁵Met concentration in OPT and OPTLEU is higher than CON because Met was added to meet Cys requirement (Met + Cys).

⁶Concentrations of Ca and P were based on phytase activity from the premix.

Table 2.

Analyzed and calculated concentration of N, total EAA and free EAA in experimental diets¹ (as-fed)

	Control		Optimal		Optimal + Leu
	Analyzed	Calculated2	Analyzed	Calculated	Analyzed	Calculated
Total, %
DM	88.76	—	88.95	—	89.15	—
N	3.00	3.08	2.20	2.24	2.28	2.29
Arg	1.23	1.26	0.75	0.78	0.80	0.78
His	0.49	0.53	0.39	0.43	0.40	0.43
Ile	0.85	0.81	0.61	0.60	0.64	0.60
Leu	1.65	1.67	1.14	1.19	1.59	1.64
Lys	1.11	1.04	1.08	1.01	1.11	1.01
Met	0.27	0.31	0.27	0.33	0.31	0.33
Met + Cys	0.56	0.63	0.48	0.57	0.52	0.57
Phe	0.98	0.96	0.75	0.76	0.77	0.76
Phe + Tyr	1.60	1.59	1.19	1.20	1.23	1.20
Thr	0.72	0.73	0.64	0.68	0.66	0.68
Trp	0.25	0.23	0.18	0.19	0.18	0.19
Val	0.94	0.90	0.89	0.89	0.92	0.89
Free AA, %
Arg	0.03	0.00	0.01	0.00	0.01	0.00
His	0.00	0.00	0.07	0.07	0.07	0.07
Ile	0.01	0.00	0.08	0.08	0.08	0.08
Leu	0.01	0.00	0.01	0.00	0.43	0.45
Lys	0.02	0.00	0.36	0.37	0.37	0.37
Met3	0.00	0.00	0.07	0.11	0.07	0.11
Met + Cys	0.00	0.00	0.07	0.11	0.07	0.11
Phe	0.00	0.00	0.12	0.13	0.12	0.13
Phe + Tyr	0.01	0.00	0.12	0.13	0.12	0.13
Thr	0.02	0.00	0.20	0.20	0.20	0.20
Trp4	—	0.00	—	0.05	—	0.05
Val	0.00	0.00	0.27	0.29	0.27	0.29

¹Analyzed values represent average across 3 blocks (feed mixes).

²Calculated values for the total AA are based on the AA concentration in feed ingredients according to NRC (2012), and calculated values for the free AA correspond to the dietary inclusion rate in crystalline form.

³Addition of dl-Met was omitted in one of the 3 blocks, thus reducing the overall free Met concentration across all 3 blocks. The average free Met concentration between blocks 1 and 3 was 0.11 and was zero in block 2. Therefore, across blocks 1, 2, and 3, average free Met was 0.07.

⁴Analysis of free Trp was not performed.

Nitrogen Balance

For the N balance study, sows with an actual feed intake relative to predicted feed intake of 75% or above were used. Nitrogen balance was conducted during early lactation (between days 4 and 8) and peak lactation (between days 14 and 18) on a subset of sows from blocks 2 (n = 10), 3 (n = 12), and 4 (n = 12) for a total of 34 sows. During the N balance period, sow overall activity and appetite were carefully monitored, along with measurements of rectal temperature before the morning and afternoon feeding to ensure that sows were healthy with no signs of urinary tract infections. The urinary catheter was removed for any sows showing signs of depression or increase in rectal temperature. Urine collection was performed for a minimum of 48 h and a maximum of 72 h. Balance studies were conducted in either early lactation or late lactation to minimize urinary tract irritation and follow animal care guidelines; hence, the number of sows in early and peak lactation differed. Total urine collection and fecal grab sampling methods were as described in Huber et al. (2015) and Möhn and de Lange (1998), respectively. Briefly, Foley urinary catheters (BARDEX I.C., 2-way, 30cc balloon, 18FR, Bard Medical, Covington, GA) were aseptically inserted into the bladder before feeding in the morning at 0600. The distal end of the catheter was connected to a sterilized polyvinyl tubing secured with electrical tape and long enough to reach a 5-gallon bucket set behind the sow and outside of the crate. The tubing was maintained in place through a rubber stopper inserted into the bucket cover. The urine collection bucket contained 30 mL of H₂SO₄ to acidify the urine and maintain pH of less than 3. Urine was removed and weighed daily at 0700, and 2 subsamples (45 mL) were collected and frozen at −20 °C. Urinary catheters were removed before feeding at 0700 on the last day of the N balance (either 48 or 72 h). Fresh feces were collected by rectal digital stimulation on days 10 and 11, pooled and frozen at −20 °C.

Milk Sampling

Milk was collected after each N balance (days 8 and 18). For milk collection, piglets were separated from the sows for approximately 1 h, and sows were administered 1 mL of oxytocin i.m. (20 IU/mL oxytocin, sodium chloride 0.9% w/v, and chlorobutanol 0.5% w/v, VetTek, Blue Springs, MO). A total of 100-mL milk was manually collected across all glands and stored in 2 separate 50-mL tubes (polypropylene centrifuge tubes with screw cap, Denville Scientific). Piglets were immediately returned to sows to complete nursing.

Nutrient and Titanium Analyses

Approximately 50 g of subsampled feed was ground using a commercial coffee grinder and sent to the Agricultural Experiment Station Chemical Laboratories (University of Missouri–Columbia, Columbia, MO) for AA analyses (AOAC Official Method 982.30 E (a,b,c), 45.3.05, 2006) to verify accuracy of feed mixing. Both hydrolysate and free AA concentrations were analyzed to verify the accuracy of crystalline AA (CAA) inclusion during feed mixing (Table 2). The DM content of diets was measured via oven drying at 135 °C for 2 h according to AOAC (1997; Method 930.15). Fecal samples were homogenized, oven dried at 65 °C for 4 d, and ground using a commercial coffee grinder. Feed, fecal, and urinary N concentration was measured based on the Hach method (Hach et al., 1987). Milk samples were submitted to the Michigan Dairy Herd Improvement Association (NorthStar Cooperative, Lansing, MI) for analyses of fat, true protein, lactose, and milk urea N using infrared spectroscopy. Titanium concentration in feed and feces were analyzed based on Myers et al. (2004). Absorbance of standards and samples was measured by spectrophotometry (Beckman DU-7400; Beckman Instruments, Inc., Fullerton, CA) at 408 nm.

Calculations

Sow milk yield was estimated based on piglet ADG (g/d) during early (days 4 to 8) and peak (days 14 to 18) lactation (Theil et al., 2002) as follows (Eq. 1 and 2, respectively):

$$\begin{array}{l}\begin{array}{ll}\text{Daily milk yield }(\text{g}/\text{d},\text{ days }4\text{ to }8)=\hfill & \text{Litter size}\hfill \\ \hfill & \times (317+1.168\times \text{ ADG}\hfill \\ \hfill & +0.00425\times {\text{ADG}}^2)\hfill \\ \hfill & \hfill \end{array}\hfill \end{array}$$ (1)

$$\begin{array}{l}\begin{array}{ll}\text{Daily milk yield }(\text{g}/\text{d},\text{ days }14\text{ to }18)=\hfill & \text{Litter size}\hfill \\ \hfill & \times (582+1.168\times \text{ ADG}\hfill \\ \hfill & +0.00425\times {\text{ADG}}^2)\hfill \\ \hfill & \hfill \end{array}\hfill \end{array}$$ (2)

For all calculations pertaining to the N balance, the analyzed N concentration in each respective diet and corresponding block was used to calculate N intake. Daily total N retention (N maternal retention + N milk) and N maternal retention were calculated as follows (Eq. 3 and 4, respectively):

$${\displaystyle \begin{array}{ll}{\displaystyle \mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}(\mathrm{g}/\mathrm{d})=}& \mathrm{N}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}(\mathrm{g}/\mathrm{d})\\ & -[\mathrm{f}\mathrm{e}\mathrm{c}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t}(\mathrm{g}/\mathrm{d})\\ & +\mathrm{u}\mathrm{r}\mathrm{i}\mathrm{n}\mathrm{a}\mathrm{r}\mathrm{y}\mathrm{N}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t}(\mathrm{g}/\mathrm{d})]\end{array}}$$ (3)

$${\displaystyle \begin{array}{ll}{\displaystyle \mathrm{M}\mathrm{a}\mathrm{t}\mathrm{e}\mathrm{r}\mathrm{n}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}(\mathrm{g}/\mathrm{d})=}& \mathrm{N}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}(\mathrm{g}/\mathrm{d})\\ & -[\mathrm{f}\mathrm{e}\mathrm{c}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t}(\mathrm{g}/\mathrm{d})\\ & +\mathrm{u}\mathrm{r}\mathrm{i}\mathrm{n}\mathrm{a}\mathrm{r}\mathrm{y}\mathrm{N}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t}(\mathrm{g}/\mathrm{d})\\ & +\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}\mathrm{N}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t}(\mathrm{g}/\mathrm{d})]\end{array}}$$ (4)

Actual daily feed intake and analyzed N concentration of the diets (Table 2) were used to calculate daily N intake in each respective block. Apparent total tract digestibility (ATTD) of N was estimated using analyzed titanium dioxide concentration in feed and feces (Eq. 5) according to Zhu et al. (2005), and fecal N output was calculated based on the estimated N digestibility and N intake, as follows (Eq. 6).

$$\begin{array}{ll}\hfill & \text{Apparent total tract}\hfill \\ \hfill & \text{digestibility coefficient of N}=1-\frac{{\text{TiO}}_2\text{ \% in feed }\times \text{ N \% in feces}}{{\text{TiO}}_2\text{ \% in feces}\times \text{ N \% in feed}}\hfill \end{array}$$ (5)

$${\displaystyle \begin{array}{ll}{\displaystyle \mathrm{F}\mathrm{e}\mathrm{c}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t}(\mathrm{g}/\mathrm{d})=}& (1-\mathrm{A}\mathrm{T}\mathrm{T}\mathrm{D}\mathrm{o}\mathrm{f}\mathrm{N})\\ & \times \mathrm{N}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}(\mathrm{g}/\mathrm{d})\end{array}}$$ (6)

Daily urine weight and urinary N concentrations were used to calculate daily urinary N output. Daily milk N output was calculated based on the sum of analyzed milk true protein N and milk urea N concentrations multiplied by the predicted daily milk yield. Apparent efficiency of dietary N utilization was expressed as efficiency of total N retention (maternal + milk) and of N secreted in milk, relative to N intake or N absorbed, as follows (Eq. 7 and 8, respectively):

$${\displaystyle \begin{array}{l}{\displaystyle \begin{array}{c}\mathrm{A}\mathrm{p}\mathrm{p}\mathrm{a}\mathrm{r}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{e}\mathrm{f}\mathrm{f}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{c}\mathrm{y}\\ \mathrm{o}\mathrm{f}\mathrm{t}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\end{array}=\frac{\begin{array}{c}\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{N}\\ \mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}(\mathrm{g}/\mathrm{d})\end{array}}{\begin{array}{c}\mathrm{N}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}\mathrm{o}\mathrm{r}\mathrm{N}\\ \mathrm{a}\mathrm{b}\mathrm{s}\mathrm{o}\mathrm{r}\mathrm{b}\mathrm{e}\mathrm{d}(\mathrm{g}/\mathrm{d})\end{array}}\times 100\mathrm{\%}}\end{array}}$$ (7)

$${\displaystyle \begin{array}{l}{\displaystyle \begin{array}{c}\mathrm{A}\mathrm{p}\mathrm{p}\mathrm{a}\mathrm{r}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{e}\mathrm{f}\mathrm{f}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{c}\mathrm{y}\\ \mathrm{o}\mathrm{f}\mathrm{N}\mathrm{s}\mathrm{e}\mathrm{c}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{d}\mathrm{i}\mathrm{n}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}\end{array}=\frac{\begin{array}{c}\mathrm{N}\mathrm{s}\mathrm{e}\mathrm{c}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{d}\\ \mathrm{i}\mathrm{n}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}(\mathrm{g}/\mathrm{d})\end{array}}{\begin{array}{c}\mathrm{N}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}\mathrm{o}\mathrm{r}\\ \mathrm{N}\mathrm{a}\mathrm{b}\mathrm{s}\mathrm{o}\mathrm{r}\mathrm{b}\mathrm{e}\mathrm{d}(\mathrm{g}/\mathrm{d})\end{array}}\times 100\mathrm{\%}}\end{array}}$$ (8)

For calculations pertaining to true efficiency estimation of N and individual EAA utilization, an adjustment was made to account for any discrepancy between the analyzed and calculated dietary AA concentrations. Relying on calculated SID N and EAA intake alone may either underestimate or overestimate true efficiency values. Therefore, the SID N or individual SID EAA concentrations were adjusted by multiplying the calculated SID N or SID EAA concentration with the ratio of analyzed to calculated N or EAA concentrations (as-fed basis) in each of the respective block (Eq. 9):

$$\text{Adjusted SID N or EAA concentration =\hspace{0.17em}Calculated SID N or EAA (\%) }\times \frac{\text{Analyzed N or EAA (\%, as fed)}}{\text{Calculated N or EAA (\%, as fed)}}$$ (9)

Daily individual SID N or EAA intake was then calculated from the actual sow ADFI and adjusted dietary SID N or EAA as follows (Eq. 10):

$${\displaystyle \begin{array}{l}{\displaystyle \begin{array}{ll}{\displaystyle \mathrm{S}\mathrm{I}\mathrm{D}\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}(\mathrm{g}/\mathrm{d})=}& \mathrm{S}\mathrm{o}\mathrm{w}\mathrm{f}\mathrm{e}\mathrm{e}\mathrm{d}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}(\mathrm{g}/\mathrm{d})\\ & \times \mathrm{a}\mathrm{d}\mathrm{j}\mathrm{u}\mathrm{s}\mathrm{t}\mathrm{e}\mathrm{d}\mathrm{S}\mathrm{I}\mathrm{D}\mathrm{N}\mathrm{o}\mathrm{r}\\ & \mathrm{E}\mathrm{A}\mathrm{A}(\mathrm{g}/100\mathrm{g})\\ & {\displaystyle }\end{array}}\end{array}}$$ (10)

True efficiency values of N and individual EAA secreted in milk were determined by correcting for N or EAA mobilized from body protein and used for maintenance, as follows (Eq. 11):

$${\displaystyle \begin{array}{l}{\displaystyle \begin{array}{l}\mathrm{T}\mathrm{r}\mathrm{u}\mathrm{e}\mathrm{e}\mathrm{f}\mathrm{f}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{c}\mathrm{y}\\ \mathrm{o}\mathrm{f}\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\\ \mathrm{s}\mathrm{e}\mathrm{c}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\mathrm{i}\mathrm{n}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}\end{array}=\frac{\begin{array}{c}\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{o}\mathrm{u}\mathrm{p}\mathrm{u}\mathrm{t}\mathrm{i}\mathrm{n}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}(\mathrm{g}/\mathrm{d})\\ -\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{m}\mathrm{o}\mathrm{b}\mathrm{i}\mathrm{l}\mathrm{i}\mathrm{z}\mathrm{e}\mathrm{d}\\ \mathrm{f}\mathrm{r}\mathrm{o}\mathrm{m}\mathrm{b}\mathrm{o}\mathrm{d}\mathrm{y}\mathrm{p}\mathrm{r}\mathrm{o}\mathrm{t}\mathrm{e}\mathrm{i}\mathrm{n}(\mathrm{g}/\mathrm{d})\end{array}}{\begin{array}{c}\mathrm{S}\mathrm{I}\mathrm{D}\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{k}\mathrm{e}(\mathrm{g}/\mathrm{d})\\ -\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{f}\mathrm{o}\mathrm{r}\mathrm{m}\mathrm{a}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{a}\mathrm{n}\mathrm{c}\mathrm{e}(\mathrm{g}/\mathrm{d})\end{array}}}\end{array}}$$ (11)

where the N or EAA output in milk in early and peak lactation periods were calculated from estimated milk yield (Theil et al., 2002) for early lactation and peak lactation period, respectively, and the average N or EAA concentration in mature milk protein (NRC, 2012), as follows (Eq. 12):

$${\displaystyle \begin{array}{l}{\displaystyle \begin{array}{ll}{\displaystyle \mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t}\mathrm{i}\mathrm{n}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}(\mathrm{g}/\mathrm{d})=}& \mathrm{M}\mathrm{i}\mathrm{l}\mathrm{k}\mathrm{y}\mathrm{i}\mathrm{e}\mathrm{l}\mathrm{d}(\mathrm{g}/\mathrm{d})\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\\ & \times \mathrm{i}\mathrm{n}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}\mathrm{p}\mathrm{r}\mathrm{o}\mathrm{t}\mathrm{e}\mathrm{i}\mathrm{n}(\mathrm{g}/100\mathrm{g})\\ & {\displaystyle }\end{array}}\end{array}}$$ (12)

Daily N mobilized from body protein and partitioned to milk was estimated by multiplying the negative maternal N retention with the efficiency of N secretion in milk from mobilized body N of 0.87 (NRC, 2012), as follows (Eq. 13):

$${\displaystyle \begin{array}{l}{\displaystyle \begin{array}{ll}{\displaystyle \mathrm{N}\mathrm{m}\mathrm{o}\mathrm{b}\mathrm{i}\mathrm{l}\mathrm{i}\mathrm{z}\mathrm{e}\mathrm{d}(\mathrm{g}/\mathrm{d})=}& \mathrm{M}\mathrm{a}\mathrm{t}\mathrm{e}\mathrm{r}\mathrm{n}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}(\mathrm{g}/\mathrm{d})\\ & \times \mathrm{e}\mathrm{f}\mathrm{f}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{c}\mathrm{y}\mathrm{o}\mathrm{f}\mathrm{b}\mathrm{o}\mathrm{d}\mathrm{y}\mathrm{N}\mathrm{m}\mathrm{o}\mathrm{b}\mathrm{i}\mathrm{l}\mathrm{i}\mathrm{z}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\\ & \times \mathrm{t}\mathrm{o}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}\mathrm{N}\mathrm{s}\mathrm{e}\mathrm{c}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}(0.87)\\ & {\displaystyle }\end{array}}\end{array}}$$ (13)

Daily individual EAA mobilized from body protein and partitioned to milk was estimated from the product of the negative maternal N retention and the EAA concentration in body protein (NRC, 2012), multiplied by the efficiency of N secretion in milk from mobilized body N of 0.87, as follows (Eq. 14):

$${\displaystyle \begin{array}{ll}{\displaystyle \mathrm{E}\mathrm{A}\mathrm{A}\mathrm{m}\mathrm{o}\mathrm{b}\mathrm{i}\mathrm{l}\mathrm{i}\mathrm{z}\mathrm{e}\mathrm{d}(\mathrm{g}/\mathrm{d})=}& \mathrm{M}\mathrm{a}\mathrm{t}\mathrm{e}\mathrm{r}\mathrm{n}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}(\mathrm{g}/\mathrm{d})\\ & \times 6.25\times \mathrm{E}\mathrm{A}\mathrm{A}\mathrm{i}\mathrm{n}\mathrm{b}\mathrm{o}\mathrm{d}\mathrm{y}\mathrm{p}\mathrm{r}\mathrm{o}\mathrm{t}\mathrm{e}\mathrm{i}\mathrm{n}(\mathrm{g}/100\mathrm{g})\\ & \times \mathrm{e}\mathrm{f}\mathrm{f}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{c}\mathrm{y}\mathrm{o}\mathrm{f}\mathrm{b}\mathrm{o}\mathrm{d}\mathrm{y}\mathrm{N}\mathrm{m}\mathrm{o}\mathrm{b}\mathrm{i}\mathrm{l}\mathrm{i}\mathrm{z}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\\ & \mathrm{t}\mathrm{o}\mathrm{m}\mathrm{i}\mathrm{l}\mathrm{k}\mathrm{N}\mathrm{d}\mathrm{e}\mathrm{p}\mathrm{o}\mathrm{s}\mathrm{t}\mathrm{i}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}(0.87)\end{array}}$$ (14)

Daily SID N and SID EAA were calculated as described above in Eq. 9.

Maintenance requirement for N or individual EAA was calculated as the sum of basal endogenous gastrointestinal tract (GIT) and integumental N or EAA losses (NRC, 2012), and the efficiency of N or EAA utilization for maintenance (NRC, 2012), as follows (Eq. 15):

$${\displaystyle \begin{array}{l}{\displaystyle \mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{f}\mathrm{o}\mathrm{r}\mathrm{m}\mathrm{a}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{a}\mathrm{n}\mathrm{c}\mathrm{e}=\frac{\begin{array}{c}\mathrm{B}\mathrm{a}\mathrm{s}\mathrm{a}\mathrm{l}\mathrm{e}\mathrm{n}\mathrm{d}\mathrm{o}\mathrm{g}\mathrm{e}\mathrm{n}\mathrm{o}\mathrm{u}\mathrm{s}\mathrm{G}\mathrm{I}\mathrm{T}\mathrm{N}\mathrm{o}\mathrm{r}\\ \mathrm{E}\mathrm{A}\mathrm{A}\mathrm{l}\mathrm{o}\mathrm{s}\mathrm{s}(\mathrm{g}/\mathrm{d})\\ +\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{e}\mathrm{g}\mathrm{u}\mathrm{m}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{N}\\ \mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{l}\mathrm{o}\mathrm{s}\mathrm{s}(\mathrm{g}/\mathrm{d})\end{array}}{\begin{array}{c}\mathrm{N}\mathrm{o}\mathrm{r}\mathrm{E}\mathrm{A}\mathrm{A}\mathrm{e}\mathrm{f}\mathrm{f}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{c}\mathrm{y}\\ \mathrm{f}\mathrm{o}\mathrm{r}\mathrm{m}\mathrm{a}\mathrm{i}\mathrm{n}\mathrm{t}\mathrm{e}\mathrm{n}\mathrm{a}\mathrm{n}\mathrm{c}\mathrm{e}\end{array}}}\end{array}}$$ (15)

Statistical Analyses

Statistical analyses were conducted using SAS 9.4 (SAS Inst. Inc., Cary, NC). The homogeneity of residual variance among dietary treatments (minimum P = 0.088 for milk protein output) and normality of residuals were confirmed by using Mixed Procedures and Univariate Procedures, respectively.

Data were analyzed by ANOVA using the Glimmix procedures model as follows:

$${\displaystyle \begin{array}{ll}{\displaystyle {\mathrm{Y}}_{ijklm}=}& \mathrm{\mu }+a_i+b_j+p_k+t_l+d_{m(ij)}\\ & +{(\mathrm{a}\mathrm{b})}_{ij}+(\mathrm{a}\mathrm{p}{)}_{ik}+(\mathrm{a}\mathrm{t}{)}_{il}+e_{ijklm}\end{array}}$$

where Y_ijklm is the response on animal m of parity k for treatment i in block j at period l, μ is the treatment mean, a_i is the fixed effect of dietary treatment i, p_k is the fixed effect of parity k (e.g., early [P 2 to 3] vs. late parity [P 4 to 6]), t_l is the fixed effect of lactation period l (e.g., early vs. peak lactation), b_j is the random effect of block j, d_m(ij) is the random effect of animal m nested within treatment i block j, (ap)_ik is the fixed interactive effect of treatment i with parity k, (at)_il is the fixed interactive effect of treatment i with period l, (ab)_ij is the random interactive effect of treatment i with block j, and e_ijklm is the random error on animal m of parity k for treatment i in block j at period l. When appropriate, a reduced model was used. Specifically, effects of parity and parity × treatment were not significant (minimum P = 0.18 and P = 0.13, respectively) and therefore were excluded in the reduced model for analyses of all lactation performance and N balance data, and individual EAA efficiency values. Pairwise comparisons (OPT vs. CON, OPTLEU vs. CON, and OPTLEU vs. OPT) were carried out for different period of lactation (early, peak, and 21-d overall lactation) using the slice option in SAS and Tukey adjustment. Effects were declared significant at P ≤ 0.05, and tendencies at 0.05 ≤ P ≤ 0.10.

RESULTS

Dietary Amino Acid Analyses

Analyzed N and individual EAA concentration values agreed closely with their calculated values derived from selected NRC (2012) feed ingredients (Table 2). Analyzed values were within a minimum of 96% of the expected calculated values. Of note however was Met, with analyzed to calculated values of 87%, 82%, and 94% in CON, OPT, and OPTLEU diets, respectively. The discrepancy between calculated and analyzed values of Met was attributed to the omission of supplemental dl-Met in block 2 of the nitrogen balance studies, as revealed from the free AA analysis report (see Table 2 footnote). In addition, the lower analyzed relative to calculated Met concentration value in the CON diet may have been attributed to a lower Met concentration in soybean meal in NRC (2012) than that of the actual concentration in soybean meal used for this study. As described earlier in Methods, because individual SID EAA intake was calculated with an adjustment to account for any discrepancy between analyzed and calculated EAA concentrations, albeit very small for the majority of EAA, there was no difference in Met efficiency between blocks.

Performance

Lactation performance data of all sows are presented in Table 3. Sow feed intake, BW, and back fat loss did not differ between dietary treatments. Sow BW and back fat loss differed from zero (P = 0.025) for sows fed OPT and did not differ from zero in sows fed CON and OPTLEU. The interaction between dietary treatments and lactation period for litter growth rate (LGR) and ADG was significant (P < 0.05). Litter growth rate during early lactation period and over the 21 d of lactation period did not differ across dietary treatments. At peak lactation, compared with CON, LGR of sows fed OPT was greater (P < 0.05) and that of sows fed OPTLEU did not differ. Compared with OPT, sows fed OPTLEU had lower LGR (P < 0.05).

Table 3.

Lactation performance of all sows fed Control (CON; 18.74% CP), Optimal (OPT; 13.78% CP), or Optimal + Leucine (OPTLEU; 14.25% CP) over a 21-d lactation period

	Diet				P-value
Item	CON	OPT	OPTLEU	SEM1	OPT vs CON	OPTLEU vs. CON	OPTLEU vs. OPT
No. of sows	18	19	17
Parity	3.4	3.5	3.3
Sow ADFI, kg/d2
Overall, days 1 to 21	5.30	5.18	5.23	0.22	0.809	0.923	0.970
Early, days 4 to 8	4.73	4.39	4.45	0.25	0.341	0.494	0.969
Peak, days 14 to 18	6.27	6.28	6.23	0.25	0.999	0.987	0.981
Sow initial BW, kg	246	249	252	7	0.921	0.787	0.953
Sow BW change3, kg	−1.6	−8.4*	−0.6	3.0	0.282	0.969	0.216
Sow initial back fat, mm	16.9	18.8	18.8	1.4	0.432	0.445	1.000
Sow back fat change3, mm	−1.2	−3.6*	−1.6	0.9	0.188	0.932	0.310
Litter size
Day 14	10.3	10.3	10.2	0.24
Day 21	9.6	10.0	9.9	0.27
Litter growth rate, kg/d2
Overall, days 1 to 21	2.45	2.59	2.35	0.13	0.541	0.700	0.208
Early, days 4 to 8	2.33	2.35	2.44	0.18	0.990	0.854	0.911
Peak, days 14 to 185	2.71	3.28	2.65	0.18	0.026	0.963	0.016
Piglet ADG, g/d2
Overall, days 1 to 21	253	259	237	9	0.896	0.485	0.291
Early, days 4 to 8	233	234	244	15	1.000	0.877	0.885
Peak, days 14 to 185	278	329	264	16	0.047	0.797	0.011

¹Maximum value of the SE of the least squares means.

²The main effect of period (early vs. peak) was significant (P < 0.01) for feed intake, LGR, and ADG. Interaction of treatment × period for LGR (P = 0.035) and ADG (P = 0.033). LGR = litter growth rate.

^3,*Body weight and back fat change were different from 0 (P = 0.025 and P = 0.005, respectively).

⁴Litter size after standardization (within 24 h after parturition).

⁵One litter (OPTLEU) was excluded for LGR and ADG due to a negative growth rate.

Lactation performance, and milk nutrient concentration and output are presented in Table 4. In early lactation, piglet ADG, estimated daily milk yield, milk true protein, lactose and fat concentration and output did not differ between diets. At peak lactation, piglet ADG of sows fed OPTLEU was lower (P < 0.05) compared with that of sows fed OPT. Estimated daily milk yield of sows fed OPT tended to be greater than CON (P = 0.06) and that of OPTLEU did not differ from CON and was lower (P < 0.05) than OPT. Milk true protein and lactose concentration did not differ between dietary treatments. Sows fed OPT tended to have higher (P = 0.08) milk fat concentration than CON, and those fed OPTLEU did not differ from CON or OPT. Milk true protein output did not differ between dietary treatments. Lactose output of sows fed OPT tended to be greater (P = 0.107) than that of CON, but did not differ between OPTLEU and CON, and was lower (P < 0.05) in sows fed OPTLEU compared with OPT. Milk fat output of sows fed OPT was higher (P < 0.05) than CON and did not differ for sows fed OPTLEU when compared with CON or OPT. In both early and peak lactation periods, milk urea N of sows fed OPT and OPTLEU was lower (P < 0.01) compared with CON and did not differ between OPTLEU and OPT.

Table 4.

Performance and milk nutrient composition and yield in early and peak lactation periods of sows selected for the N balance studies and fed Control (CON; 18.74% CP), Optimal (OPT; 13.78% CP), or Optimal + Leucine (OPTLEU; 14.25% CP) diets

	Diet				P-value
Item	CON	OPT	OPTLEU	SEM1	OPT vs CON	OPTLEU vs. CON	OPTLEU vs. OPT
Early lactation (days 4 to 8)2
No. of sows	12	11	11
Sow ADFI, kg/d	4.93	4.64	4.58	0.23	0.390	0.268	0.957
Litter size	10.3	10.3	10.2	0.3
Piglet ADG, g/d	248	248	255	21	1.000	0.962	0.957
Estimated milk yield, kg/d3	8.76	8.84	8.79	0.94	0.996	0.999	0.999
Milk nutrient concentration
True protein, %	4.49	4.25	4.25	0.14	0.315	0.335	1.000
Urea nitrogen, mg/dL	12.30	3.81	3.51	0.82	<0.001	<0.001	0.949
Lactose, %	5.52	5.49	5.60	0.20	0.952	0.738	0.560
Fat, %	6.93	7.89	6.97	0.50	0.342	0.998	0.378
Milk nutrient output, g/d
True protein output	390.5	375.3	387.4	39.0	0.954	0.998	0.971
Lactose output	484.6	486.7	494.5	53.4	0.999	0.981	0.988
Fat output	606.3	701.4	621.0	89.4	0.730	0.993	0.800
Peak lactation (days 14 to 18)2
No. of sows	11	11	11
Sow ADFI, kg/d	6.83	6.65	6.38	0.23	0.722	0.125	0.422
Litter size	9.9	10.2	9.9	0.3
Piglet ADG, g/d	262	311	238	22	0.173	0.648	0.031
Estimated milk yield, kg/d3	11.62	13.90	11.01	0.98	0.059	0.809	0.016
Milk nutrient concentration
True protein, %	4.41	4.35	4.39	0.14	0.934	0.994	0.966
Urea nitrogen, mg/dL	15.51	4.84	5.85	0.82	<0.001	<0.001	0.572
Lactose, %	5.65	5.69	5.62	0.20	0.888	0.965	0.755
Fat, %	6.23	7.76	7.00	0.50	0.083	0.510	0.510
Milk nutrient output, g/d
True protein output	512.3	607.3	530.7	39.8	0.195	0.935	0.333
Lactose output	655.5	767.2	619.8	55.0	0.107	0.793	0.030
Fat output	725.9	1077.7	841.2	90.0	0.026	0.637	0.165

¹Maximum value of the SE of the least squares means.

²The main effect of period (early vs. peak) was significant except for ADG, milk fat, protein, lactose, and milk N output/N intake.

³Estimated milk yield was based on piglet ADG.

Nitrogen Balance

Nitrogen absorption, retention, and utilization efficiency are presented in Table 5.

Table 5.

Nitrogen utilization for milk in early and peak lactation periods in sows selected for the N balance studies and fed Control (CON; 18.74% CP), Optimal (OPT; 13.78% CP), or Optimal + Leucine (OPTLEU; 14.25% CP) diets¹

	Diet				P-Value
Item	CON	OPT	OPTLEU	SEM2	OPT vs CON	OPTLEU vs. CON	OPTLEU vs. OPT
Early lactation (days 4 to 8)3
No. of sows	12	11	11
Body weight, kg4	245.4	255.8	246.3	7.4	0.440	0.994	0.493
N intake, g/d	152.1	112.9	106.0	4.7	<0.001	<0.001	0.482
N absorbed, g/d	137.4	93.8	95.7	3.8	<0.001	<0.001	0.936
Dry fecal output, kg/d	0.523	0.538	0.586	0.057	0.979	0.713	0.826
Urine weight, kg/d	10.1	4.7	5.5	1.5	0.047	0.103	0.920
Urinary N, g/kg	4.91	3.22	3.23	0.64	0.161	0.166	0.999
N excretion, g/d
Fecal N	14.59	15.47	16.56	1.49	0.909	0.625	0.864
Urinary N	37.84	14.29	14.93	4.39	<0.001	<0.001	0.993
Milk N	61.7	59.0	62.1	5.4	0.927	0.999	0.909
Total N retention, g/d	99.6	79.6	80.1	5.1	0.009	0.011	0.948
Maternal N retention, g/d	37.8*	20.7*	20.0*	8.2	0.308	0.286	0.998
Apparent N utilization efficiency
Total N retention, % of N intake	65.5	72.3	75.8	3.0	0.050	0.005	0.315
Total N retention, % of N absorbed	72.4	84.6	84.1	3.0	0.002	0.003	0.882
Milk N output, % of N intake	41.1	54.7	58.3	3.2	0.005	<0.001	0.417
Milk N output, % of N absorbed	45.4	63.6	66.1	3.8	0.006	0.002	0.890
Peak lactation (days 14 to 18)3
No. of sows	11	11	11
Body weight, kg4	249.4	249.3	250.0	7.5	0.999	0.998	0.996
N intake, g/d	210.0	151.5	145.7	4.4	<0.001	<0.001	0.565
N absorbed, g/d	189.3	130.3	122.3	3.8	<0.001	<0.001	0.311
Dry fecal output, kg/d	0.72	0.74	0.81	0.06	0.971	0.461	0.600
Urine weight, kg/d	13.2	5.6	6.1	1.5	0.005	0.009	0.969
Urinary N, g/kg	4.06	3.30	3.17	0.64	0.683	0.593	0.988
N excretion, g/d
Fecal N	20.3	21.2	22.9	1.5	0.901	0.429	0.689
Urinary N	36.9	17.7	18.6	4.5	0.006	0.008	0.984
Milk N	81.7	99.4	85.5	5.4	0.064	0.871	0.168
Total N retention, g/d	149.8	112.7	109.6	5.2	<0.001	<0.001	0.671
Maternal N retention, g/d	68.3*	13.4	17.8*	8.2	<0.001	<0.001	0.922
Apparent N utilization efficiency
Total N retention, % of intake	71.4	74.5	73.4	3.0	0.363	0.556	0.756
Total N retention, % of absorbed	79.2	86.6	87.2	3.0	0.050	0.037	0.862
Milk N output, % of N intake	39.5	62.9	58.4	3.6	<0.001	<0.001	0.328
Milk N output, % of N absorbed	43.9	73.2	69.5	3.8	<0.001	<0.001	0.780

¹Nitrogen balance was conducted between days 4 and 8 or days 14 and 18 for either 48 or 72 h.

²Maximum value of the standard error of the least squares means.

³The main effect of period was significant for all variables, except BW, UN output, maternal N retention, NB/N intake, milk N/N intake, milk N/N absorb.

⁴Body weight of days 1 and 21 were used as reference for early and peak lactation.

*Maternal N retention was different from 0 (P < 0.05).

Early lactation.

Milk N excretion did not differ between sows fed OPT and CON, as well as between OPTLEU and OPT. Compared with sows fed CON, urine output was lower (P < 0.05) in OPT and tended to be lower (P = 0.10) in OPTLEU. Maternal N retention was positive (P < 0.05) and did not differ between diets.

Peak lactation.

Milk N excretion of sows fed OPT tended to be greater (P = 0.06) than those fed CON, and did not differ between OPTLEU and OPT. Sows fed OPT and OPTLEU had lower (P < 0.01) maternal N retention compared with those fed CON.

Early and peak lactation.

Nitrogen intake, N absorbed, urinary N excretion, and total N retention were lower (P < 0.05), and apparent efficiency of N utilization for milk N secretion was greater (P < 0.05) in sows fed OPT and OPTLEU compared with sows fed CON, and did not differ between OPTLEU and OPT.

True Nitrogen and Essential Amino Acid Efficiencies for Milk N and EAA Deposition

True dietary N and EAA efficiency for milk production are presented in Table 6. Individual EAA efficiency did not differ between early and peak lactation periods. In early, peak, and overall lactation period, compared with CON, N, Arg, His, Ile, Leu, Phe, and Trp efficiency were greater (P < 0.05) and those of Lys, Met, and Val did not differ in sows fed OPT or OPTLEU. In early lactation, compared with CON, Thr efficiency in sows fed OPT or OPTLEU did not differ. At peak lactation, compared with CON, Thr efficiency tended to be greater (P = 0.054) in sows fed OPT, but did not differ in sows fed OPTLEU. Individual EAA efficiency did not differ between OPTLEU and OPT, except for that of Leu and Met. Utilization efficiency of Leu in sows fed OPTLEU was lower (P < 0.01) compared with sows fed OPT and did not differ from that of sows fed CON. Utilization efficiency of Met was lower (P < 0.05) and tended to be lower (P = 0.10) in sows fed OPTLEU compared with those fed OPT during peak and overall lactation period, respectively.

Table 6.

True dietary AA utilization efficiency estimated based on maternal N retention for milk protein production of sows fed Control (CON; 18.74% CP), Optimal (OPT; 13.78% CP), or Optimal + Leucine (OPTLEU; 14.25% CP) diets between days 4 and 8 of lactation (early lactation) and between days 14 and 18 of lactation (peak lactation)

	Diet					P-value
Item	CON	OPT	OPTLEU	NRC (2012) 1	SEM2	OPT vs. CON	OPTLEU vs. CON	OPTLEU vs. OPT
Early lactation (days 4 to 8)3
No. of sows4	12	10	11
Arg	32.8	58.4	54.9	—	3.7	<0.001	<0.001	0.728
His	54.1	74.3	72.5	—	5.1	0.002	0.004	0.755
Ile	41.7	61.9	59.4	—	4.5	0.001	0.002	0.853
Leu	45.2	71.2	48.1	—	3.5	<0.001	0.491	<0.001
Lys	57.3	60.1	58.5	—	3.6	0.823	0.960	0.944
Met	62.4	64.6	56.0	—	4.3	0.885	0.368	0.197
Met+Cys	59.8	74.3	68.2	—	5.6	0.035	0.274	0.520
Phe	36.9	50.6	49.5	—	3.7	0.006	0.010	0.955
Phe+Tyr	45.8	65.8	63.9	—	4.4	0.002	0.004	0.926
Thr	58.7	67.4	66.4	—	4.6	0.252	0.314	0.984
Trp	44.5	66.1	66.7	—	6.0	0.010	0.008	0.996
Val	50.4	54.2	52.7	—	3.9	0.645	0.846	0.934
N	50.7	75.4	72.7	—	4.1	<0.001	0.002	0.882
EAA5	50.1	63.4	58.8	—	4.2	0.026	0.167	0.607
Peak lactation (days 14 to 18)3
No. of sows4	9	10	9
Arg	33.8	63.8	57.5	—	3.8	<0.001	<0.001	0.399
His	55.7	82.2	75.9	—	5.3	<0.001	0.004	0.308
Ile	42.9	68.8	62.3	—	4.6	<0.001	0.003	0.391
Leu	46.3	79.1	50.7	—	3.7	<0.001	0.366	<0.001
Lys	58.9	66.2	61.3	—	3.8	0.325	0.893	0.581
Met	64.5	71.3	58.3	—	4.5	0.375	0.460	0.041
Met+Cys	61.8	82.2	71.0	—	5.8	0.005	0.287	0.148
Phe	38.1	56.3	51.8	—	3.8	<0.001	0.011	0.542
Phe+Tyr	47.0	73.2	67.1	—	4.6	<0.001	0.004	0.497
Thr	60.5	74.5	69.3	—	4.7	0.054	0.302	0.628
Trp	45.7	74.2	69.6	—	6.1	0.001	0.007	0.792
Val	51.8	59.9	55.0	—	4.1	0.191	0.764	0.531
N	51.9	82.7	75.9	—	4.3	<0.001	0.002	0.497
EAA5	51.6	70.2	61.6	—	4.4	0.003	0.149	0.212
Overall lactation6
Arg	33.3	61.1	56.2	81.6	3.2	0.004	0.008	0.469
His	54.9	78.3	74.2	72.2	4.4	0.016	0.030	0.679
Ile	42.3	65.4	60.8	69.8	4.0	0.007	0.016	0.500
Leu	45.7	75.1	49.4	72.3	2.9	0.002	0.574	0.004
Lys	58.1	63.2	59.9	67.0	2.9	0.451	0.889	0.688
Met	63.4	67.9	57.2	67.5	3.8	0.523	0.333	0.100
Met+Cys	60.8	78.2	69.6	66.2	5.1	0.034	0.224	0.230
Phe	37.5	53.4	50.6	73.3	3.2	0.016	0.030	0.677
Phe+Tyr	46.4	69.5	65.5	70.5	3.7	0.010	0.021	0.620
Thr	59.6	71.0	67.9	76.4	3.9	0.123	0.254	0.773
Trp	45.0	70.1	68.1	67.4	5.5	0.029	0.038	0.941
Val	51.1	57.0	53.8	58.3	3.4	0.297	0.719	0.650
N	51.3	79.1	74.3	75.9	3.3	0.009	0.017	0.600
EAA5	50.8	66.8	60.2	—	3.6	0.029	0.141	0.293

¹Efficiency values of AA for lactation were reported by NRC (2012) only for the whole lactation period.

²Maximum value of the standard error of the least squares means.

³The main effect of period was not significant (EAA period effect: CON, P = 0.740; OPT, P = 0.128; OPTLEU, P = 0.537).

⁴Only sows consuming at least 75% of the predicted feed intake over the entire 4-d periods (i.e., 4 to 8 d and 14 to 18 d) were included in the estimation of efficiency values.

⁵EAA is the average efficiency values of all the EAA listed above excluding Arg.

⁶Mean values between early and peak.

DISCUSSION

Objectives of the Present Study

The goal of the study was in part to determine the MBEV of N and EAA in lactating sows by feeding a diet containing an NIAA profile. We first formulated a diet limiting in all EAA down to the minimum SID Leu requirement. Because Arg is synthesized de novo, and its essentiality has not been characterized for the lactating sow, it was not possible to create a practical diet limiting in Arg, and therefore, MBEV for Arg was not determined. To generate MBEV for practical prediction of EAA requirement, each limiting EAA was supplemented in their crystalline form to meet their minimum SID requirement (NRC, 2012) and to attain an NIAA profile. Several previous studies reported that similar dietary strategies to the current work either maintained or increased milk yield, casein yield and LGR (Manjarín et al., 2012; Chamberlin et al., 2015a,b; Huber et al., 2015). In the present study, the overall lactation performance was unaffected however sow fed OPT had greater BW and back fat loss. In contrast, at peak lactation, sows fed OPT had greater LGR and milk fat output and tended to have greater milk yield. Our results corroborate with those of Huber et al. (2015) who suggested that ameliorating dietary AA balance may facilitate nutrient partitioning toward milk protein synthesis. Although sows fed a NIAA profile diet had greater milk N production at peak lactation, neither milk true protein concentration nor true protein yield differed. What was noticeably greater was the milk fat yield. Estimation of body lipid mobilization in future work will be important to further understand the potential impact of feeding a NIAA profile on nutrient partitioning. A second objective was to determine whether the corresponding decrease in Leu concentration in reduced CP diet (OPT) affects the efficiency of Lys utilization. The only difference between OPT and OPTLEU was the additional LEU in the OPTLEU diet whereby SID Leu:SID Lys was 1.14:1 and 1.63:1, respectively. The SID Leu:SID Lys was identical between OPTLEU and CON. As initially hypothesized, the addition of Leu to the OPT diet reduced milk yield at peak lactation to similar level as that of CON, potentially indicating an AA imbalance and interaction between Leu and other EAA utilization for milk production. Sows fed OPTLEU and CON did not lose appreciable BW and were in positive maternal N balance. Supplementary Leu has been reported to improve muscle (Escobar et al., 2006) and visceral (Torrazza et al., 2010) protein synthesis in piglets, thus Leu in CON and OPTLEU may have played a role in nutrients partitioning away from the mammary gland and toward maternal body.

Lactation Performance and N Balance

It is clear that the reduced CP diets not only maintain lactation performances compared with nonreduced CP diets, but also greatly improve the global efficiency of N utilization. Feeding either OPT or OPTLEU diets led to dramatic decrease in urinary N excretion and increase in overall apparent N utilization efficiency for milk N production up to 73% and true N utilization efficiency of up to 82.7%. Urine weight decreased by 58% and urinary N excretion by up to 60%. Difference in daily quantitative urinary N excretion between CON and low protein diets (OPT or OPTLEU) was attributed in this study to both urine volume and urinary N concentration. Others have also reported that reducing dietary CP concentrations can lead to lower urine volume in horses (Wickens, 2003), lactating sows (Huber et al., 2015), and growing pigs (Shaw et al., 2006). Additionally, the lower milk urea N secretion parallels the urinary N excretion, suggesting less AA catabolism in OPT than CON diets (Huber et al., 2015). Across diets, sows were in a positive maternal N balance in early lactation, whereas sows fed OPT ended up at maternal N equilibrium during peak lactation. The apparent discrepancy between average maternal N retention (17 ± 8 g/d, Table 5) and BW loss (400 ± 143 g/d, Table 3) of OPT fed sows may be explained in part by the contribution from fat loss rather than from body protein loss. Furthermore, 400 g BW loss per day translates into 2 ± 3 g N/d when accounting for water and protein mass (NRC, 2012). In addition, there may have been some degree of overestimation of N retention (MacRae et al., 1993).

Biological Efficiency of Amino Acid for Lactation

Level of dietary CP reduction and CAA inclusion, and the practical implementation of thereof for lactating sows are dependent on feed and AA costs, and whether environmental constraints are in place. A major focus of the present study was to determine MBEV for individual EAA and to assess whether Leu affects efficiency of EAA. Accurate prediction of dietary AA requirement using the factorial approach is directly dependent on MBEV, a fundamental focus of the modeling approach employed by NRC (2012). The reported MBEV in NRC (2012) however were not experimentally determined except for that of Lys, which was later validated by Huber et al. (2015). Thus, reduced CP diet with an NIAA profile is a powerful tool to experimentally generate MBEV of EAA. In the study reported by Huber et al. (2016), MBEV was only estimated for Lys because all other EAA were present in excess of requirement in the reduced CP diets. When predicting efficiency of EAA using the available literature data (White et al., 2016), the majority of efficiency of EAA are grossly underestimated relative to those of NRC (2012). The majority of available studies have focused on assessing the minimum requirement for Lys which corresponds to the point of near maximum biological utilization. Therefore, Lys is the only EAA for which reliable efficiency value can be predicted (NRC, 2012; White et al., 2016) and a close estimation of Lys requirement for milk production exist.

Dependency of lysine efficiency on amino acid balance.

This study aimed at assessing whether MBEV of Lys is independent of N and EAA concentration because NRC (2012) estimated MBEV of Lys in diets containing N and all of the other AA in excess of their requirements. Similarly, Huber et al. (2015) validated MBEV of Lys in sows fed reduced CP diet and containing the other EAA in excess of their requirement. Utilization efficiency of 66.2% for Lys at peak lactation in the present study was similar to that of Huber et al. (2016) at 67.6% and NRC (2012) at 67.0%. As mentioned earlier, separate MBEV for early and peak lactation may be potentially relevant if phase feeding is implemented in lactation. Both Huber et al. (2016) and NRC (2012) used calculated SID Lys values to estimate efficiency. Here, if calculated values are used, overall lactation Lys MBEV (data not shown) aligns perfectly with that of NRC (2012). Instead, the calculated AA values were adjusted based on the analyzed values to account for discrepancy because a minor discrepancy can have a large impact on efficiency estimation.

Efficiency of amino acid for early vs. peak lactation.

In the present study, the OPT diet formulated to contain an NIAA profile was used to estimate MBEV of individual EAA. There were noticeable changes in efficiency values from early to peak lactation between diets; however, the limited number of sows and the relatively high SEM precluded drawing strong conclusions pertaining to the impact of lactation stage. Nonetheless, because trends were very consistent for each individual EAA, N, and averaged EAA, additional work is clearly warranted to ascertain the individual MBEV of EAA in early and peak lactation with a larger number of sows. Results herein are pointing to possible larger differences in EAA requirements between early and peak lactation, which is not captured in the current NRC (2012) because only one MBEV was estimated for the entire lactation period.

Efficiency of amino acid for Optimal vs. Control.

Consistent and significantly greater efficiency of use for Arg, His, Ile, Leu, Met + Cys, Phe, Phe + Tyr, and Trp in sows fed OPT relative to those fed CON indicate that these AA were in excess of requirements in the CON diet during both early and peak lactation. As well, except for Arg, these EAA reached their MBEV in the OPT diet because this diet was definitively limiting in Ile, Leu, Met + Cys, Phe, Phe + Tyr, and Trp. For Thr, the noticeable trend from early to peak lactation between diets is indicative that Thr was in excess of requirement in early lactation and near requirement in the OPT diet at peak lactation. On the other hand, efficiency values of Val and Met did not differ between OPT and CON. It is therefore likely that both Met and Val were near their minimum requirement and were at maximum biological efficiency in the CON diet. Such low MBEV for Val is supported by several studies. For instance, Val uptake by the sow mammary gland relative to its output in milk is the largest amongst the EAA (Trottier et al., 1997; Lei et al., 2012). Previous in vivo isotope tracer research conducted in our lab (Guan et al., 2002) showed that the net Val output to net Val uptake ratio by mammary gland was 0.56 in sows fed a diet with Val: Lys of 1.04, and 0.45 in sows fed a diet with Val to Lys ratio of 1.37. In this study, Val MBEV was 57% in sows fed Val: Lys of 0.88 (OPT), closely agreeing with Guan et al. (2002). The net Val output:net Val uptake determined with tracer approach is essentially a true mammary efficiency value because it is independent of Val used for maintenance and Val from body protein mobilization. Therefore, the study by Guan et al. (2002) validates the calculations used herein and by others (NRC, 2012; Huber et al., 2016) for estimating efficiency of EAA utilization. We propose to adopt the word “true” when estimating efficiency using such approach. Moreover, Val requirement for swine lactation has been reported as 44.3 g/d by Guan et al. (2004) based on maximal mammary uptake of EAA, which is higher than a predicted 38.5 g/d based on NRC (2012) model. Xu et al. (2017) suggested a higher Val:Lys requirement ratio (88% to 113%) than 85% previously reported by NRC (2012) based on minimum back fat loss and maximum piglet growth rate, suggesting that Val MBEV from NRC (2012) may be slightly overestimated, and as such, underestimating Val requirement. Metabolic pathways of Val utilization in mammary gland are unknown. Trottier (1995) proposed that Val is retained by the mammary gland for remodeling of in situ mammary proteins. Valine was also reported to be used for the synthesis of glutamate AA family (Li et al., 2009). Our data point to Val among the top 4 limiting EAA, as previously suggested by others (Kim et al., 2001; Xu et al., 2017).

Efficiency of amino acid for Optimal vs. NRC.

For several EAA, the overall MBEV derived from the OPT diet agree with those of NRC (2012), except for Arg and Phe. Estimated efficiency values for Arg and Phe were noticeably lower than those reported in NRC (2012), i.e., 61.1% vs. 81.6% and 53.4% vs. 73.3%, respectively. The amount of Arg taken up by the mammary glands greatly exceeds Arg output in milk (Trottier et al., 1997; O’Quinn et al., 2002); therefore, its efficiency of use for milk protein should be relatively low. Furthermore, since Arg is synthesized de novo via the intestinal-renal axis (Tomlinson et al., 2011; Marini et al., 2017), it is recognized as a conditionally essential AA (NRC, 2012). It is likely that the NRC (2012) reported value of 81.6% is a gross overestimation and a true MBEV for Arg may not be estimable. With regard to Phe, it is unknown whether its low efficiency is indicative that Phe was in excess in the OPT diet. On the other hand, mammary metabolic pathways for Phe are unknown but it is possible that there is a high rate of Phe hydroxylation to Tyr in mammary tissue. For instance, total aromatic EAA efficiency value from OPT compared with NRC (2012) was very close, i.e., 69.5% vs. 70.5%. Threonine MBEV between the present study and NRC (2012) was lower than expected with 71.0% vs. 76.4%. As observed for Lys, Thr MBEV at peak lactation was 74.5% which is in closer agreement with that of NRC (2012) value for the overall lactation of 76.4%.

Efficiency of amino acid for Optimal + Leucine vs. Optimal.

We previously suggested the presence of competitive inhibition between AA for their utilization by the mammary gland, potentially between Lys and Leu (Guan et al., 2004; Manjarín et al., 2012). Earlier, high concentration of Leu was reported to inhibit Lys uptake in rat mammary explants (Shennan et al., 1994; Calvert and Shennan, 1996). Reduced CP diet with CAA inclusion increased mammary extraction efficiency of Lys and Arg (Manjarín et al., 2012). We questioned whether increase in efficiency of EAA in a reduced CP diet was related in part to a reduction in Leu. Addition of Leu to the OPT did not affect efficiency of Lys or the majority of EAA. However, the efficiency of Met was reduced as a result of Leu supplementation. This response was unexpected but offers an insight into potential interaction between crystalline Leu and Met utilization by the mammary gland via common transporter systems (Manjarín et al., 2014).

To summarize, MBEV for individual EAA were estimated for Ile, Leu, Lys, Met, Met + Cys, Phe + Tyr, Thr, Trp, and Val by feeding a diet that met the minimum SID requirement for Leu. Generating efficiency estimates for Arg and potentially His may not be biologically relevant given de novo synthesis of Arg and possible mammary excretion of His (Trottier et al., 1997). Valine MBEV is low relative to other EAA and agrees with that of NRC (2012) and previous work which supports a low efficiency of Val utilization for milk production. Nonetheless, testing OPT diets with limiting Val as low as 50% of NRC (2012) are critically needed to further validate this low efficiency value. In addition, the MBEV of other EAA, in particular Thr and Phe should be validated using the same approach but with graded levels of inclusion from 30% below to 30% above NRC (2012) requirements using a similar OPT diet as used in this study. Leucine did not reduce efficiency of N, Lys, and other EAA utilization; therefore, Leu concentration in conventional diets is unlikely to be directly affecting the global utilization of N as proposed in our earlier work. Feeding an NIAA diet not only maintained overall milk production and litter growth, but increased litter growth between days 14 and 18 of lactation, corroborating results from previous studies. The increase in performance was accompanied by greater milk fat yield and a tendency to increase milk production, reduction in maternal N retention, and loss in BW and back fat, indicating possible nutrient repartitioning toward the mammary glands. Leucine therefore may be playing a role in maternal N retention and sow body condition during lactation rather than interacting with Lys utilization for milk production, as initially hypothesized. In fact, it is unknown whether feeding reduced CP diets to lactating sows over multiple lactations affect sow body condition and longevity. As mentioned earlier, practical implementation of such diets will depend on feed and CAA availability and costs and on environmental constraints. Continued testing of such diets to generate and validate the MBEV of EAA is critical to refine future models for prediction of EAA requirements. The increase in several EAA efficiency with reduction in dietary protein and improvement of AA balance suggest a need to establish a dynamic model to predict EAA requirement under different scenarios of dietary protein concentrations and crystalline AA inclusion rates.