Increasing dietary proportion of wheat grain in finishing diets containing distillers’ grains: impact on nitrogen utilization, ruminal pH, and digestive function

Abstract

Because of its high crude protein (CP) content, dietary inclusion of corn dried distillers’ grains with solubles (DDGS) in finishing cattle diets can increase the ruminal loss of ammonia-nitrogen (NH₃-N), which ends up excreted as urine urea-N (UUN). Increasing dietary fermentable energy supply can enhance ruminal use of N; however, it could also lead to acidotic conditions that compromise digestive function and animal performance. We evaluated the effects of partially replacing dietary corn grain with 20% or 40% (dry matter [DM] basis) wheat grain in finishing diets containing 15% corn DDGS on N utilization, ruminal pH, and digestive function. Nutrient intake and digestion, ruminal fermentation characteristics, microbial protein synthesis, route of N excretion, and blood metabolites were measured. Six ruminally fistulated crossbred beef heifers (initial body weight ± SD; 797 ± 58.8 kg) were used in a replicated 3 × 3 Latin square design with 28-d periods. Dietary treatments were either corn (73% of diet DM; CON), 53:20 corn:wheat blend (20W), or 33:40 corn:wheat blend (40W) as the major fermentable energy source. Dry matter intake (DMI) tended to be lower for heifers fed the 40W than CON and 20W diets. Feeding diets containing wheat grain led to an increase (P = 0.04) in neutral detergent fiber (NDF) intake. However, there was no diet effect (P ≥ 0.60) on apparent total tract DM and NDF digestibility. Feeding wheat grain led to a decrease (P ≤ 0.03) in mean and minimum pH, an increase (P = 0.04) in pH < 5.8 duration, and a tendency for an increase in the area and acidosis index for pH < 5.8 and 5.5. Nitrogen intake, which was lower (P = 0.04) for 40W than 20W heifers did not differ between CON and 20W heifers. There was no diet effect (P = 0.80) on ruminal NH₃-N concentration and estimated microbial N flow. However, feeding diets containing wheat grain led to a decrease (P = 0.045) in UUN excretion (% total urine N). Fecal and total N excretion (% of N intake) increased (P < 0.01) following the addition of wheat grain to the diet. Apparent N retention was lower (P = 0.03) for 40W than CON and 20W heifers. In summary, although it led to a desirable decrease in UUN excretion, feeding wheat grain in corn DDGS-containing diets increased acidotic conditions in the rumen, which possibly led to the tendency for a decrease in DMI. The negative apparent N retention at the 40% wheat grain inclusion also suggests a decrease in nutrient supply, which could compromise feedlot performance.

Introduction

A major challenge when formulating cattle diets, which contain high protein byproducts like dried distillers’ grains with soluble (DDGS), is achieving a dietary inclusion concentration that lowers feed costs substantially, while preventing overfeeding of nitrogen (N), which increases N excretion (Barros et al., 2017). Excreted N, especially urinary N, is readily converted to reactive forms like ammonia-N (NH₃-N) that compromise air and water quality. In the United States, finishing rations typically contain between 15% and 25% (dry matter [DM] basis) distiller’s grains (Samuelson et al., 2016). However, feeding as low as 7% to 14% distiller’s grains (DM basis) increased ruminal NH₃-N and blood urea-N concentrations (Beliveau and McKinnon, 2009; Li et al., 2013) and urinary excretion of N (Hales et al., 2013). Because NH₃-N emissions increased even at low dietary inclusion levels, there is a need for feeding strategies that limit N wastage when DDGS are added to cattle diets (Legesse et al., 2018).

A major factor that influences ruminal and, ultimately, whole-body N use, is dietary carbohydrate supply (Nocek and Russell, 1988; Hristov et al., 2005). When ruminal fermentable energy supply is inadequate, microbial sequestration of diet-derived peptide-, amino-, and NH₃-N is limited, thereby increasing the loss of ruminal NH₃-N into blood and, ultimately, urinary excretion of urea-N (UUN). Therefore, increasing ruminal fermentable energy supply when feeding DDGS could enhance N utilization. However, to date, most of the research on fermentable energy supply has primarily focused on grain processing (DiLorenzo and Gaylean, 2010). Steam-flaking of corn grain increases ruminal starch digestibility by up to 24% (Harrelson et al., 2019). Therefore, several studies (Corrigan et al., 2009; Swanson et al., 2014) have evaluated the effects of feeding steam-flaked compared with dry-rolled corn in finishing diets containing DDGS on growth performance. However, measures of N utilization, including urinary excretion of N and urea-N were not evaluated in those studies.

Besides processing method, grain type also influences ruminal fermentable energy supply. Corn and wheat are the major grains fed to feedlot cattle in the United States (Samuelson et al., 2016). Although it typically contains less starch, the rate and extent of ruminal starch degradation (RSD) is greater for wheat than corn grain (Huntington, 1997). As a result, RSD can be up to 25% greater for wheat than corn grain depending on numerous factors including processing method (Ferraretto et al., 2013). Although it remains to be evaluated in DDGS-containing finishing diets, this difference in RSD, could favorably alter N utilization. Zinn (1992) and Plascencia et al. (2018) reported an increase in ruminal and post-ruminal N digestion, microbial N supply, and microbial efficiency when feeding steam-flaked wheat (74% DM) compared with steam-flaked corn grain. However, the recommendation is for dietary inclusion of wheat grain to not exceed 40% to 50% of diet DM due to concerns related to ruminal acidosis, which can compromise digestive function, growth performance, and carcass quality (Lardy and Dhuyvetter, 2000; Yang et al., 2014). Therefore, we examined effects of partially replacing dry-rolled corn grain with up to 40% (DM basis) dry-rolled wheat grain in finishing diets containing 15% corn DDGS on nutrient intake and digestion, ruminal fermentation characteristics, measures of N utilization, and blood metabolites. We hypothesized that partial replacement of corn grain with up to 40% (DM) wheat grain in diets containing corn DDGS enhances N utilization without compromising nutrient intake and digestive function.

Materials and Methods

Animal use for this experiment was approved by the Institutional Animal Care and Use Committee at the University of Idaho (Protocol # 2018-23).

Animals, experimental design, and treatments

Six ruminally fistulated crossbred beef heifers (797 ± 58.8 kg; mean body weight [BW] ± SD at the start) were randomly assigned to one of three dietary treatments in a replicated 3 × 3 Latin Square design balanced for residual effects. Experimental and sample collection periods were 28 d in length with 21 d for dietary adaptation and 7 d for measurements. Prior to the start of the study, heifers were transitioned to a diet containing 73% corn grain DM.

The three dietary treatments were either dry-rolled corn grain (73% of diet DM; CON), 53:20 corn:wheat grain blend (20W), or 33:40 corn:wheat grain blend (40W) as the major cereal grain source (Table 1). The club wheat used in the study was dry-rolled to a processing index (ratio of volume weight of processed grain to the original volume weight) of 80% (Yang et al., 2014; Moya et al., 2015). The three diets also contained corn DDGS (15%), grass hay (10%), mineral–vitamin supplement (1.16%), and varying amounts of urea to ensure they were isonitrogenous. Monensin (Rumensin premix, Elanco Animal Health, Ontario, Canada) was added to all diets (28 mg/kg DM). The diets were formulated to provide enough energy, protein, minerals, and vitamins to exceed the nutrient requirements of animals gaining 1.5 kg/d (NASEM, 2016). Total mixed rations (TMR) were prepared and offered once daily at 0700 hours for ad libitum intake. During the first 12 d of each period, the proportion of corn or wheat grain was adjusted upward or downward as appropriate using 4-d steps.

Table 1.

Dietary ingredient and chemical composition of experimental diets

Items	Diet1
	CON	20W	40W
Ingredient, % of DM
Hay	10.0	10.0	10.0
Corn DDGS2	15.0	15.0	15.0
Corn	73.2	53.2	33.2
Wheat	0.0	20.0	40.0
Urea	0.30	0.10	0.00
Mineral vitamin mix3	1.16	1.16	1.16
Chemical composition
Dry matter (DM), %	88.4 ± 0.03	89.0 ± 0.34	89.7 ± 0.10
Organic matter, % of DM	97.1 ± 0.01	97.0 ± 0.33	97.0 ± 0.27
Crude protein, % of DM	12.4 ± 0.38	12.1 ± 0.49	11.7 ± 0.75
Starch, % DM	51.2 ± 2.01	50.7 ± 1.45	50.1 ± 0.91
Acid detergent fiber, % of DM	7.59 ± 0.50	8.27 ± 0.47	8.40 ± 0.47
Neutral detergent fiber (NDF), % of DM	21.4 ± 1.38	23.2 ± 2.39	26.8 ± 1.10
Indigestible NDF, % of DM	10.7 ± 0.01	11.7 ± 0.02	12.2 ± 0.02

¹Experiental diets: corn (73% of diet DM, CON), 53:20 corn:wheat blend (20W), and 33:40 corn:wheat blend (40W).

²Dried distillers grains with solubles.

³Supplement DM contained CP, 51.3%; crude fat, 0.48%; salt, 12.3%, Ca, 19.7%; P, 0.07%; Mg, 0.55%; K, 0.10%, S, 0.15%, Fe, 12.5%, Mn, 1,230 ppm; Zn, 2,050 ppm; organic Zn, 1,025 ppm; Cu, 615 ppm; organic Cu, 205 ppm; Co, 31.2 ppm; I, 175 ppm; Se, 13.5 ppm; Selenium yeast, 4.51 ppm; Vitamin A, 27,948 IU/kg; Vitamin D, 2,795 IU/kg; Vitamin E, 93.1 IU/kg; and Rumensin 90 (Elanco Animal Health, Greenfield, IN), 1,128 g/ton.

Although statistical analysis could not be conducted due to the use of single lots of dietary ingredients, corn grain had a greater starch but lower crude protein (CP), neutral detergent fiber (NDF), and acid detergent insoluble N (ADIN) content than wheat grain (Table 2). Similar observations were made in other studies (Moate et al., 2017; Alvarez-Hess et al., 2019) comparing the two cereal grains. In situ ruminal starch disappearance at 4 and 8 h of incubation was 20% to 36% greater for wheat than corn grain in the present study (Table 2). This is in agreement with others (Philippeau et al., 1999; McAllister and Sultana, 2011), with the greater RSD attributed to the greater solubility of the protein matrix adhered to starch granules for wheat than corn grain (Zinn et al., 2002).

Table 2.

Chemical composition and in situ ruminal starch disappearance of corn and wheat grain

Items	Corn grain	Wheat grain	SEM	P-value
Chemical composition
Dry matter (DM), %	87.6 ± 0.75	87.5 ± 0.75	–	–
Organic matter, % of DM	98.4 ± 0.20	98.2 ± 0.08	–	–
Crude protein (CP), % of DM	6.62 ± 0.57	9.16 ± 0.72	–	–
Acid detergent insoluble nitrogen, % of CP	2.64 ± 0.58	0.40 ± 0.09	–	–
Neutral detergent insoluble nitrogen, % of CP	3.21 ± 0.49	6.49 ± 0.48	–	–
Acid detergent fiber, % of DM	3.57 ± 1.09	3.47 ± 0.33	–	–
Neutral detergent fiber, % of DM	10.4 ± 0.86	20.2 ± 3.70	–	–
Starch, % of DM	70.1 ± 2.75	67.5 ± 0.44	–	–
In situ starch ruminal disappearance, % of total starch
4 h	58.4	69.8	3.63	0.05
8 h	64.8a	87.8b	1.36	<0.01

^a,bMeans with different superscripts differ (P < 0.05).

Measurements

All heifers were weighed prior to morning feeding on two consecutive days at the beginning of each experimental period and at the end of the study. To determine dry matter intake (DMI), total mixed ration (TMR) offered, and orts were recorded daily. Weekly TMR and feed ingredient samples were collected over two consecutive days, whereas refusals samples were collected over five consecutive days. The TMR and feed ingredient samples were composited by week. Refusal samples were composited by animal and week, with their nutrient composition later used when calculating nutrient intake.

To measure diurnal changes in ruminal NH₃-N concentration, digesta was collected on day 23 at 0630, 0800, 0900, 1000, 1300, 1600, 1900, and 2300 hours, and on day 24 at 0300 hours. Ruminal pH was measured continuously from days 22 to 28 of each period using indwelling pH data loggers (Lethbridge Research and Development Centre pH data logger system, Dascor, Escondido, CA; Penner et al., 2006). At the beginning of each measurement period, the loggers were standardized in pH 4 and 7 buffers and programmed to record pH every minute.

Grab fecal samples were collected on day 26 (0900, 1500, and 2100 hours), day 27 (0300, 1200, and 1800 hours), and day 28 (0000 and 0600 hours) and stored at −20 °C for later apparent total tract nutrient digestibility and N balance determination (Morris et al., 2018). Spot urine samples were also collected at each fecal sampling time (Lee et al., 2019). A subsample (80 mL) of the collected urine was immediately mixed with 5 mL of 2M H₂SO₄ to a pH <2.5 and placed on dry ice to prevent the loss of NH₃-N. Thereafter, 1 mL of the acidified urine was diluted 1:10 with distilled H₂O, composited by animal and period, and frozen (−20 °C) for later analysis of total N, urea-N, creatinine, and purine derivatives (PD). To measure ruminal NH₃-N and short-chain fatty acid (SCFA) concentration over a 24-h feeding cycle, approximately 500 mL of ruminal digesta was also collected at the same time as grab fecal samples. In addition, to measure diurnal changes in ruminal NH₃-N concentration, digesta was also collected on day 23 at 0630, 0800, 0900, 1000, 1300, 1600, 1900, and 2300 hours, and on day 24 at 0300 hours. At each sampling point, approximately 500 mL of digesta was collected from the cranial ventral, caudal ventral, central, and cranial dorsal regions of the rumen. Samples were strained through polyester monofilament fabric (350 µm mesh opening; ELKO Filtering Co, LLC, Fort Lauderdale, FL). For later NH₃-N analysis, a 5-mL aliquot of ruminal fluid was collected and mixed with chilled 1% H₂SO₄, prior to storage (−20 °C). For later SCFA analysis, a 5-mL ruminal fluid aliquot was collected, mixed with chilled 25% (wt/vol) meta-phosphoric acid (H₂PO₄), and stored at −20 °C.

On the last day of each period (d 28), blood samples were collected via jugular venipuncture 3 h post-feeding into a 10-mL tubes containing 158 IU lithium heparin (Becton Dickinson, Franklin Lakes, NJ). The lithium heparin tubes were stored briefly on ice before centrifugation (2,400 × g; 20 min; 4 °C), and then harvest and subsequent storage (−20 °C) of plasma in cryogenic vials for later analysis of plasma urea-N (PUN) and glucose concentrations.

Laboratory analyses

Composited TMR, feed ingredient, and refusals (collected during the last week of each period) and daily fecal samples were oven-dried at 55 °C for 72 h for chemical analysis. Dried fecal samples were pooled based on their respective DM content to obtain a representative composite sample by animal within period. Fecal, dried TMR, feed ingredient, and refusals samples were sequentially ground through 4- and 1-mm screens (Retsch Cutting Mill SM 200, Retsch). All ground samples were then analyzed for analytical DM by drying at 135 °C for 2 h (AOAC, 2005; method 930.15). Samples were combusted at 600 °C for at least 5 h to determine ash content, and the organic matter (OM) content was calculated by difference (DM − ash; AOAC, 2005; method 942.05). Samples were analyzed for acid detergent fiber (ADF) and NDF, with amylase and sodium sulfite used during NDF determination (AOAC, 2005; method 2002.04). The Kjeldahl procedure (Foss Analytics; Hillerød, Denmark; AOAC, 1990; method 976.05) was used for CP analysis. The neutral detergent insoluble N (NDIN) and ADIN content were determined by analyzing the NDF (obtained using the above NDF procedure without the use of sodium sulfite) and ADF residues, respectively, for N. The indigestible NDF (iNDF) content of TMR, refusal, and fecal samples were determined as described by Valente et al. (2011). Briefly, samples (0.6 g) were weighed into F57 bags (Ankom Technology; Macedon, NY) that were then incubated for 288 h in the rumen of two cows. After incubation, the residues were analyzed for NDF as previously described. In situ ruminal starch degradability for corn and wheat grain was determined as described by Mojahedi et al. (2018). Briefly, 5 g samples were weighed in duplicate into 10 × 18 cm polyester bags (R1020, ANKOM Technology, Macedon, NY; 50-μm porosity) before incubation in 3 ruminally fistulated heifers for 4 and 8 h. The starch content of the corn and wheat grain, and 4 and 8 h residues were then determined by enzymatic hydrolysis of α-linked glucose polymers as described by Koenig et al. (2013).

Ruminal fluid samples were thawed at room temperature, mixed thoroughly, and centrifuged (12,000 × g for 10 min at 4 °C). The supernatant from samples preserved with H₂SO₄ were analyzed for NH₃-N using a phenol-hypochlorite assay (Broderick and Kang, 1980). The supernatant from samples preserved in H₂PO₄ was collected and centrifuged again (16,000 × g for 10 min at 4 °C). The resultant supernatant was then filtered through a 0.2 μm Nylon filter and diluted 1:1 with distilled water. The concentration of SCFA was subsequently determined using a gas chromatograph fitted with a flame-ionization detector (GC-FID; 6890 Series, Hewlett-Packard; Palo Alto, CA) as described by Coats et al. (2012).

Acidified urine composites were thawed and analyzed for total N using the Kjeldahl procedure (Foss Analytics; Hillerød, Denmark; AOAC, 1990; method 976.05). Commercial kits (Arbor Assays; Ann Arbor, MI) were used for the analysis of urine creatinine and urea-N, and PUN. Urine allantoin and uric acid concentrations were determined using a method adapted from Stentoft et al. (2014). Briefly, quantification was carried out using HPLC/MS (Waters Corporation, Milford, MA) fitted with a reversed-phase column (C18, 5µm particle size, 2 mm × 250 mm; Phenomenex, Torrance, CA) and a 5% methanol mobile phase.

Calculations

Ruminal pH data for each heifer were summarized by period as daily minimum, mean, and maximum. Additionally, the duration (h/d), area under the curve (AUC; pH × h), and acidosis index (pH × h/kg DMI; Chibisa et al., 2015) were calculated using pH thresholds of 5.8 (mild acidosis; Zebeli et al., 2012), 5.5 (subacute acidosis; Schwaiger et al., 2013), and 5.2 (acute acidosis; Owens et al., 1998).

Apparent total tract digestibility of DM, OM, CP, NDF, and ADF was calculated as follows:

$${\displaystyle \begin{array}{l}{\displaystyle \begin{array}{l}100-100\times [(\mathrm{i}\mathrm{N}\mathrm{D}\mathrm{F}\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{c}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\mathrm{i}\mathrm{n}\mathrm{f}\mathrm{e}\mathrm{e}\mathrm{d}/\mathrm{i}\mathrm{N}\mathrm{D}\mathrm{F}\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{c}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\mathrm{i}\mathrm{n}\mathrm{f}\mathrm{e}\mathrm{c}\mathrm{e}\mathrm{s})\\ \times (\mathrm{n}\mathrm{u}\mathrm{t}\mathrm{r}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{c}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\mathrm{i}\mathrm{n}\mathrm{f}\mathrm{e}\mathrm{c}\mathrm{e}\mathrm{s}/\mathrm{n}\mathrm{u}\mathrm{t}\mathrm{r}\mathrm{i}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{c}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}\mathrm{i}\mathrm{n}\mathrm{f}\mathrm{e}\mathrm{e}\mathrm{d})].\end{array}}\end{array}}$$

Urine output was estimated using the concentration of creatinine measured in urine, and BW and creatinine constant of 29 mg/kg BW per day (Valadares et al., 1999) according to the following equation:

$${\displaystyle \begin{array}{l}{\displaystyle \mathrm{U}\mathrm{r}\mathrm{i}\mathrm{n}\mathrm{e}\mathrm{o}\mathrm{u}\mathrm{t}\mathrm{p}\mathrm{u}\mathrm{t},\mathrm{k}\mathrm{g}/\mathrm{d}=(29\times \mathrm{B}{\mathrm{W}}^{0.96})/\mathrm{c}\mathrm{r}\mathrm{e}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{n}\mathrm{i}\mathrm{n}\mathrm{e}\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{c}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n},\mathrm{m}\mathrm{g}/\mathrm{L}.}\end{array}}$$

Apparent N balance was calculated as the difference between N intake and excretion (fecal + urine).

The excretion of allantoin and uric acid was used to estimate the total absorption of PD as described by Chen and Gomes (1992) according to the following equation:

$${\displaystyle \begin{array}{l}{\displaystyle \mathrm{P}{\mathrm{D}}_{\mathrm{e}\mathrm{x}\mathrm{c}\mathrm{r}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{d}},\mathrm{m}\mathrm{m}\mathrm{o}\mathrm{l}/\mathrm{d}=0.85(\mathrm{P}{\mathrm{D}}_{\mathrm{a}\mathrm{b}\mathrm{s}\mathrm{o}\mathrm{r}\mathrm{b}\mathrm{e}\mathrm{d}})+(0.385\mathrm{B}{\mathrm{W}}^{0.75}),}\end{array}}$$

where 0.85 is the recovery of absorbed purines as PD and 0.385 BW^0.75 is representative of purine excretion from endogenous sources. The flow of microbial N was calculated using the following equation:

$${\displaystyle \begin{array}{l}{\displaystyle \mathrm{M}\mathrm{i}\mathrm{c}\mathrm{r}\mathrm{o}\mathrm{b}\mathrm{i}\mathrm{a}\mathrm{l}\mathrm{N},\mathrm{g}/\mathrm{d}=70(\mathrm{P}{\mathrm{D}}_{\mathrm{a}\mathrm{b}\mathrm{s}\mathrm{o}\mathrm{r}\mathrm{b}\mathrm{e}\mathrm{d}})/(0.116\times 0.83\times 1,000),}\end{array}}$$

where 70 represents the N content of purines, 0.83 is the digestibility of those purines, and the ratio of purine-N:total N in rumen microbes is 11.6:100.

Statistical analysis

All data on nutrient intake and apparent total tract digestion, ruminal fermentation characteristics, and measures of N utilization, and were analyzed using the MIXED procedure of SAS (SAS 9.4; SAS Inst. Inc., Cary, NC) for a replicated 3 × 3 Latin square. Square, period, period within square, and treatment (CON, 20W, and 40W) were considered fixed effects in the model, whereas cow within square was included as a random effect. Temporal ruminal NH₃-N data was analyzed accounting for repeated measures through the inclusion of an additional term for time (hour) and treatment × time interaction in the model described previously. Cow within square was the subject, and the variance–covariance structure of the repeated measures was modeled separately with an appropriate structure fitted using the lowest values of the fit statistics based on the Bayesian information criteria. Residual distributions were evaluated for normality and homoscedasticity prior to analysis. Data are presented as least square means. Significance was declared at P < 0.05 and trends at 0.05 < P ≤ 0.10.

Results

Nutrient intake and apparent total tract digestibility

DMI tended to be lower (P = 0.06) for heifers fed the 40W compared with CON and 20W diets (Table 3). CP intake, which was lower (P = 0.04) for 40W than 20W heifers, did not differ between CON and 20W heifers. However, feeding wheat grain resulted in an increase (P = 0.04) in NDF intake. Apparent total tract CP digestibility, which was lower (P = 0.02) for 40W than CON heifers, did not differ between CON and 20W heifers. However, there was no diet effect (P ≥ 0.60) on apparent total tract DM, OM, and NDF digestibility.

Table 3.

Nutrient intake and apparent total tract digestion for heifers fed corn (73% of diet DM, CON), 53:20 corn:wheat blend (20W), and 33:40 corn:wheat blend (40W) in diets containing 15% corn dried distillers’ grains with solubles

Variables	Diet			SEM	P-value
	CON	20W	40W
Nutrient intake, kg/d
Dry matter	15.1	15.9	13.9	0.48	0.06
Organic matter	14.7	15.3	13.6	0.47	0.11
Crude protein	1.74ab	1.87a	1.35b	0.12	0.04
Acid detergent fiber	1.19	1.38	1.25	0.06	0.11
Neutral detergent fiber	3.63a	4.17b	4.32b	0.18	0.04
Nutrient digestibility, %
Dry matter	63.4	61.4	59.1	5.03	0.60
Organic matter	65.6	63.8	62.2	4.90	0.72
Crude protein	54.4a	50.6ab	36.7b	6.82	0.02
Acid detergent fiber	28.4a	12.6b	29.5a	5.00	0.02
Neutral detergent fiber	32.2	26.0	32.7	7.52	0.64

^a,bMeans with different superscripts differ (P < 0.05).

Ruminal fermentation

Feeding wheat grain led to a decrease (P < 0.01) in mean and minimum ruminal pH (Figure 1). Similarly, there was an increase (P = 0.04) in duration pH < 5.8 for 20W and 40W compared with CON heifers (Table 4). There was a tendency for an increase (0.057 ≤ P ≤ 0.096) in pH < 5.5 duration, and area and acidosis index for pH < 5.8 and 5.5 following the addition of wheat grain to the diet. However, there was no dietary treatment effect (P ≥ 0.15) on maximum pH, and pH < 5.2 duration, area, and acidosis index. Feeding increasing amounts of wheat grain had no impact (P ≥ 0.15) on total ruminal SCFA concentration, the molar proportions of propionate, butyrate, valerate, and total BCFA, and acetate:propionate ratio. However, the molar proportion of ruminal acetate, which was greater (P = 0.04) for 20W than CON heifers, did not differ between 20W and 40W heifers.

Figure 1.

Diurnal changes in ruminal pH during the 6-d measurement period for heifers fed corn (73% of diet dry matter [DM]; CON), 53:20 corn:wheat blend (20W), and 33:40 corn:wheat blend (40W) in diets containing 15% corn dried distillers’ grains with solubles. Feeding time was 0700 hours each day. Data were plotted by hour to smooth the curves. There was no diet effect (P = 0.89) on maximum pH. However, there was a decrease in mean (P < 0.01) and minimum pH (P = 0.01) when wheat grain was added to the diet. Although there was no day effect (P ≥ 0.20) on mean, minimum, and maximum pH, and day × diet effect (P ≥ 0.63) on mean and maximum pH, there was a day × diet interaction (P = 0.04) for minimum pH. Ruminal pH data averaged across the 6-d measurement period to represent a 24-h feeding cycle is reported in Table 4.

Table 4.

Ruminal fermentation characteristics for heifers fed corn (73% of diet DM, CON), 53:20 corn:wheat blend (20W), and 33:40 corn:wheat blend (40W) in diets containing 15% corn dried distillers’ grains with solubles

Variables	Diet			SEM	P-value
	CON	20W	40W
pH
Mean	6.01a	5.83b	5.80b	0.14	0.03
Minimum	5.41a	5.18b	5.18b	0.14	0.02
Maximum	6.65	6.63	6.60	0.09	0.84
Duration of pH, min/d
<5.8	434a	693b	655b	185	0.04
<5.5	243	461	426	164	0.07
<5.2	72	161	203	82.0	0.18
Area under curve, pH × min/d
<5.8	153	289	295	106	0.06
<5.5	54.3	114	133	50.1	0.096
<5.2	6.7	17.4	37.0	12.5	0.18
Acidosis index1
<5.8	9.9	17.8	20.9	6.51	0.057
<5.5	3.50	7.05	9.45	3.121	0.090
<5.2	0.43	1.07	2.64	0.837	0.15
SCFA2
Total, mM	79.7	84.9	87.0	3.01	0.15
Acetate, mol/100 mol	48.6a	52.7b	49.9ab	3.18	0.04
Propionate, mol/100 mol	29.6	31.6	35.6	4.47	0.34
Butyrate, mol/100 mol	9.02	8.80	9.07	1.24	0.96
Valerate, mol/100 mol	1.14	1.05	1.31	0.13	0.20
Total BCFA3, mol/100 mol	2.40	2.28	3.51	0.71	0.33
Acetate:propionate	1.81	1.75	1.64	0.41	0.72

¹Area under curve/kg of DMI.

²Short-chain fatty acid.

³Branched chain fatty acid = isobutyrate + isovalaerate.

^a,bMeans with different superscripts differ (P < 0.05).

Nitrogen utilization

Nitrogen intake, which was lower (P = 0.04) for 40W than 20W heifers did not differ between CON and 20W heifers (Table 5). There was no dietary treatment effect (P ≥ 0.44) on absolute (g/d) fecal, urine, and total N excretion. Similarly, urine N excretion (% of N intake) did not differ (P = 0.28) across dietary treatments. However, feeding wheat grain led to an increase (P < 0.01) in fractional excretion (% of N intake) of fecal and total N. Urinary excretion of urea-N (% of total urine N) also decreased (P = 0.045) following the addition of wheat grain to diets. In addition, apparent N retention was lower (P = 0.03) for 40W than CON and 20W heifers. There was no diet effect or diet × time interaction for ruminal NH₃-N concentration (Figure 2). Urinary excretion of PD and estimated microbial N flow also did not differ (P ≥ 0.69) across dietary treatments. However, PUN concentration tended to increase (P = 0.06) following the addition of wheat grain to the diet.

Table 5.

Nitrogen intake, fecal N excretion, urine N and purine derivative excretion, microbial N supply and rumen ammonia-N, and plasma metabolite concentrations for heifers fed corn (73% of diet DM; CON), 53:20 corn:wheat blend (20W), and 33:40 corn:wheat blend (40W) in diets containing 15% corn dried distillers’ grains with solubles

Variables	Diet			SEM	P-value
	CON	20W	40W
Intake
Nitrogen (N), g/d	278ab	299a	217b	18.8	0.04
Fecal excretion
Dry matter, kg/d	5.64	6.15	5.69	0.85	0.77
N, g/d	132	146	141	23.1	0.79
N, % of N intake	45.6a	49.4a	67.1b	7.47	<0.01
Urinary excretion
Total output, kg/d	10.5	8.91	10.5	1.59	0.45
N, g/d	121	115	132	30.3	0.80
Urea-N, g/d	84.5	71.5	81.0	18.2	0.69
Urea-N, % of total urine N	75.5a	64.1b	64.0b	5.00	0.045
Total N, % N intake	47.3	38.0	62.7	11.7	0.28
Allantoin, mmol/d	88.9	93.1	105	23.0	0.74
Uric acid, mmol/d	74.7	80.9	62.5	20.9	0.69
Total purine derivatives, mmol/dL	119	132	121	47.8	0.95
Microbial N flow, g/d	86.5	95.7	87.7	34.8	0.94
Total N excretion
g/d	254	261	281	29.6	0.44
% N of intake	92.8a	87.4a	132.2b	10.6	<0.01
Apparent N retention, g/d	24.8a	38.3a	−62.3b	26.3	0.03
Rumen ammonia-N, mg/dL	6.00	6.80	5.05	1.02	0.80
Plasma urea-N, mg/dL	9.40	12.4	15.0	1.59	0.06
Plasma glucose, mg/dL	70.6	74.7	74.1	4.02	0.36

a,bMeans with different superscripts differ (P < 0.05).

Figure 2.

Diurnal changes in ruminal ammonia-N concentration for heifers fed corn (73% of diet dry matter [DM]; CON), 53:20 corn:wheat blend (20W), and 33:40 corn:wheat blend (40W) in diets containing 15% corn dried distillers’ grains with solubles. Feeding time was at 0700 hours. Treatment, P = 0.83; time, P < 0.01; treatment × time interaction, P = 0.43. The error bars reflect the SEM associated with time. The error bars reflect the SEM associated with time.

Discussion

Enhancing ruminal use of N, which maximizes microbial protein synthesis and limits urinary excretion of urea-N, is one of the keys to ensuring continued sustainability of the beef industry (Hristov et al., 2005). This is especially important when feeding byproducts such as corn DDGS, as their upcycling to high-quality animal protein must be done in an economically and environmentally desirable manner. Because it is a major factor that impacts ruminal and, ultimately, whole-body N metabolism, the major goal of this study was to evaluate the effects of altering ruminal fermentable energy supply by partially replacing corn with wheat grain on measures N utilization in finishing cattle fed corn DDGS as the major protein source.

As expected, in situ RSD was greater for wheat than corn grain in the present study. The observed longer duration and greater area for pH < 5.8 and 5.5 following the partial substitution of corn with wheat grain is suggestive of an increase in ruminal fermentable energy supply. Therefore, we anticipated enhanced ruminal capture and use of NH₃-N for microbial growth (Hristov et al., 2005). In addition, an increase in energy supply also upregulates microbial sequestration of peptide- and amino-N, thereby limiting their breakdown to NH₃-N. In the present study, both ruminal NH₃-N concentration and estimated microbial N supply did not differ across dietary treatments. Others (Liu et al., 2016; Alvarez-Hess et al., 2019) also did not observe changes in ruminal concentration of NH₃-N following the substitution of corn grain with wheat grain (up to 66% of diet DM). On the other hand, feeding wheat instead of corn grain (43% of diet DM) led to a decrease in ruminal NH₃-N concentration (Moate et al., 2017, 2018). Moreover, others (Philippeau et al., 1999; Plascencia et al., 2018) also reported an increase in duodenal flow of microbial N, which is suggestive of enhanced ruminal use of N for microbial growth. The reasons for the lack of a diet effect on ruminal NH₃-N concentration and microbial N supply in the present study are not clear. However, there are indications that compromised fiber digestion and DMI due to acidotic conditions in the rumen when fermentable energy supply increases can limit microbial protein synthesis (Firkins et al., 2001).

It has been suggested that a ruminal NH₃-N concentration of 5.0 to 11.8 mg/dL is needed to maximize microbial growth (Russell and Strobel, 1987; Reynal and Broderick, 2005). This wide range in requirements possibly reflects differences in dietary factors, including fermentable energy supply. For instance, in a study by Olde and Schaefer (1987), a greater NH₃-N concentration (12.5 vs. 6.1 mg/dL) was needed to match the faster in situ fractional degradation rate of barley compared to corn grain (0.036 vs. 0.024/h). In the present study, ruminal NH₃-N concentration averaged 5.9 mg/dL for heifers fed the wheat grain-containing diets. Therefore, although peptide- and amino-N supply were not evaluated, a NH₃-N deficiency could have also prevented the anticipated increase in microbial N supply. This is supported by the predicted RDP balance (Beef Cattle Nutrient Requirements Model; NASEM, 2016), which was negative across diets (−0.31, −0.30, and −0.24 kg/d for CON, 20W, and 40W diets, respectively). It is worthwhile noting that urea was added to the CON and 20W diets. Although this was to ensure that diets were isonitrogenous, it could have impacted ruminal N metabolism. However, small amounts of urea were used (0.1% and 0.3% of diet DM for 20W and CON) and, more importantly, predicted RDP balance was comparable across the three dietary treatments.

Increased ruminal loss of NH₃-N when feeding DDGS upregulates the activity of hepatocyte ornithine-cycle enzymes, which increases the synthesis, and subsequent release of urea-N into blood (Salim et al., 2015). In the present study, because there was no dietary treatment effect on ruminal NH₃-N concentration, we expected the PUN concentration to not differ. However, feeding the 40W compared with the CON and 20W diets led to a tendency for an increase in the PUN concentration. Although reasons for this are not clear, this discrepancy could be due to differences in sampling times; blood sampling was at 3 h post-feeding, whereas ruminal fluid sampling was at set time intervals over a 24-h feeding cycle. Apparent N retention was also negative for 40W heifers, which is suggestive of a metabolizable protein (MP) deficit that can upregulate mobilization of skeletal muscle protein (Roche et al., 2013). Because microbial N flow did not differ across diets, the tendency for a decrease in DMI for 40W than CON and 20W heifers possibly led to a decrease in RUP and, thus, MP supply. Although not measured, feeding wheat grain could have also reduced duodenal RUP digestibility given the observed increase in fecal N excretion (% of N intake). Therefore, a MP deficit, which increased hepatocyte deamination of skeletal muscle-derived amino acids, could account for the increase in the PUN concentration for the 40W heifers (Drackley et al., 2001; Jorritsma et al., 2003). Measures of N utilization including apparent N retention were determined using spot urine and grab fecal samples in the present study. Although useful in making treatment comparisons, there could be errors associated with accuracy of obtained estimates (Morris et al., 2018; Lee et al., 2019). Although steps to increase accuracy, including collection of at least six evenly spaced samples in a 24-h feeding cycle were followed in our study, caution is still warranted when interpreting results.

From an environmental standpoint, UUN is the major concern as its conversion by microbial urease to NH₃-N is so rapid that the process can be complete within 96 h of excretion (Hristov et al., 2011). The released NH₃-N, which can be up to 72% of fed N in feedlot cattle, and other reactive forms generated from subsequent transformations of NH₃-N, compromise air and water quality (Waldrip et al., 2015). In the present study, excreted UUN, which was within the reported range of 56% to 93% of urine N (Bristow et al., 1992), was lower when diets contained wheat grain. This was despite the tendency for an increase in PUN concentration. Between 40% and 80% of hepatic urea-N output can be recycled back to the gut, a process modulated by several factors including dietary fermentable energy supply (Lapierre and Lobley, 2001; Reynolds and Kristensen, 2008). For instance, shifting carbohydrate digestion from the small intestine to the rumen via steam-flaking compared with dry-rolling of sorghum grain resulted in an upregulation of urea-N recycling to the gut (Theurer et al., 2002). Therefore, although not quantified in this study, partial substitution of wheat with corn grain could have upregulated urea-N recycling to the gut, thereby contributing to the observed decrease in urinary excretion of urea-N as a percent of urine N. A variable amount of urea-N recycled back to the rumen can be used for anabolic purposes including supporting microbial growth. However, despite the potential increase in urea-N recycling to the gut on the wheat grain diets, there were no changes in microbial N flow. This possibly could be due to the acidotic conditions in the rumen that can compromise microbial growth. However, further studies evaluating the impact of feeding wheat grain in corn DDGS-containing diets on N transactions across the portal drained viscera, microbial capture of recycled urea-N and, ultimately, animal performance, are warranted.

Fermentation acid production in the rumen is a key determinant of pH (Aschenbach et al., 2011). In previous studies (Martin et al., 1999; Philippeau et al., 1999), increasing RSD (48% to 87% of DM) by completely replacing cracked corn with wheat grain (67% to 75% of diet DM) increased ruminal amylolytic enzyme (amylase and α-d-glucosidase) activity and total SCFA concentration, which led to a decrease in mean ruminal pH. Similarly, Russo et al. (2018) reported a decrease in mean, minimum, and maximum pH with every kg of DM of crushed wheat grain added (up to 40% DM) to lactating cow diets. Russo et al. (2018) also noted a strong negative correlation (r = −0.95) between total ruminal SCFA concentration and pH. In the present study, total SCFA concentration did not differ across dietary treatments. However, mean and minimum pH were lower for heifers fed wheat grain. The discrepancy between SCFA concentration and pH could be due to differences in the timing of measurements; pH was measured every minute over a 6-d period, whereas ruminal fluid for SCFA analysis was collected at 6-h intervals over 3 d to obtain a 24-h composite representative of a feeding cycle. This was unlike Russo et al. (2018) who concurrently measured ruminal pH and SCFA concentration over the first 4 h post-feeding. However, total SCFA production was not measured in the present study, and the study by Russo et al. (2018).

A pH-related impairment of digestive function, animal health and welfare, and production performance is a major concern when feeding wheat grain to beef cattle, and this has led to recommendations to limit the inclusion level to < 40% of diet DM (Yang et al., 2014). Fiber digestion in the rumen is compromised when pH < 5.8 (Russell and Wilson, 1996), and the occurrence of health disturbances increases as the duration pH < 5.8 extends beyond 5 to 6 h/d (Zebeli et al., 2012). A decrease in pH to < 5.6 for > 1 h initiates early inflammatory responses (Gozho et al., 2006), whereas the exposure of epithelial cells to a pH of 5.1 for only 2 h impairs barrier and absorptive functions (Meissner et al., 2017). These negative outcomes have in part led to the use of benchmark pH thresholds of 5.5 to 5.8 to diagnose subacute ruminal acidosis and 5.2 to 5.0 for acute acidosis (Aschenbach et al., 2011). In the present study, the duration and area and acidosis index for pH < 5.8 and 5.5 were greater for heifers fed wheat compared to corn grain. Although not evaluated in this study, feeding wheat than corn grain could have resulted in a greater inflammatory response and compromised health and welfare. In addition, it could have also led to microbial dysbiosis. For instance, substituting wheat for corn grain in lactating cow diets reduced the ruminal protozoa population (Moate et al., 2017), which compromises fiber digestion (Newbold et al., 2015). Martin et al. (1999) also reported a pH-related decrease in bacterial xylanase activity when cracked wheat grain completely replaced corn grain in finishing diets. However, the inflammatory response, ruminal fibrolytic enzyme activity, and fiber digestion were not evaluated in the current study.

Acidotic conditions in the rumen can compromise DMI by suppressing fiber digestion, thereby increasing fill effects (Allen, 2000). Therefore, the decrease in ruminal pH as wheat was added to the diet could account for the tendency for a decrease in DMI in our study. Besides physical regulation, hepatic oxidation of metabolic fuels including fermentation acids also impacts DMI (Allen, 2000). However, the ruminal concentration of propionate, whose hypophagic effects are well documented in ruminants, did not differ across dietary treatments in the present study. Feeding increasing amounts (35%, 55%, 75%, and 90% DM) of coarse-rolled wheat in place of corn grain also caused a decrease in DMI (Fulton et al., 1979). In that study, finishing steers fed wheat grain altered feeding behavior to cope with the wider fluctuations in ruminal pH by gradually consuming feed during the later parts of the day. Moate et al. (2017, 2018) also noted suppressed DMI in lactating cows fed rolled or crushed wheat (43.5% of diet DM). However, others (Martin et al., 1999; Philippeau et al., 1999) did not observe changes in DMI when diets contained up to 75% wheat grain. Numerous factors including the differences in dietary fiber content and fermentability, and animal-to-animal variation in responses to a decrease in pH, impact outcomes and could account for the discrepancies in DMI across studies (Pittroff and Kothman, 2001; Aschenbach et al., 2011). CP intake followed the same trend as DMI in the present study, and this decrease in dietary nutrient supply could compromise animal performance and carcass characteristics.

Conclusions

In the current study, partial replacement of dry-rolled corn grain with wheat grain (20% and 40% of diet DM) in finishing diets containing corn DDGS (15% DM) resulted in a desirable decrease in urinary excretion of urea-N as a percent of urine N. This was despite the tendency for the PUN concentration to increase when feeding wheat grain, and this is suggestive of a potential increase in the transfer of hepatic urea-N output to the gut relative to excretion in urine. However, feeding wheat grain also led to acidotic conditions in the rumen that possibly limited DM and CP intake at the 40% inclusion level. Apparent N retention was also negative for heifers fed the 40W diet, possibly reflecting a decrease in nutrient supply, which can compromise growth performance and carcass quality. Therefore, additional research evaluating the impact of feeding wheat grain in diets containing corn DDGS on N transactions across the portal drained viscera, anabolic use recycled urea-N, and growth performance and carcass characteristics of feedlot cattle is warranted.

Glossary

Abbreviations

ADF

acid detergent fiber

ADIN

acid detergent insoluble nitrogen

BW

body weight

CP

crude protein

DDGS

dried distillers’ grains with solubles

DM

dry matter

DMI

dry matter intake

iNDF

indigestible neutral detergent fiber

N

nitrogen

NDF

neutral detergent fiber

NDIN

neutral detergent insoluble nitrogen

NH₃-N

ammonia-nitrogen

OM

organic matter

PD

purine derivatives

PUN

plasma urea-nitrogen

SCFA

short-chain fatty acid

TMR

total mixed ration

UUN

urine urea-nitrogen

Conflict of interest statement

The authors declare no real or perceived conflicts of interest.