Brewer’s Grains on Growth Performance, Nutrient Digestibility, Blood Metabolites, and Fecal Microbiota in Simmental Crossbred Cattle Finished in Feedlot

Abstract

This study was conducted to evaluate the impacts of brewer’s grains (BG) on growth performance, apparent nutrient digestibility, immunity, antioxidant capacity, and fecal microbiota of Simmental crossbred cattle and the economic benefits. A completely randomized design was adopted in our study. Twenty-four 15-month-old finishing Simmental crossbred male cattle (body weight, 433.43 ± 32.47 kg) were randomly assigned to three groups: control group (basal diet), 10% BG group (supplemented with 10% BG on a dry matter basis), and 15% BG group (supplemented with 15% BG on a dry matter basis). The trial lasted for 48 days, with serum samples collected on days 24 and 48 and fecal samples collected from days 46 to 48. Diets did not influence the average daily gain, dry matter intake, feed efficiency, and serum antioxidant parameters (p > 0.05). The 15% BG group showed significantly higher acid detergent fiber digestibility (p < 0.01) and elevated serum albumin levels on day 48 (p = 0.047) compared with the control group. As for fecal microbiota, there was a lower Chao index (p = 0.040) and a higher abundance of Romboutsia in the 15% BG group (p = 0.025). Moreover, the feed costs of cattle fell by 9.34% and 14.66% after 10% and 15% BG supplementation, respectively. On the whole, BG supplementation demonstrated no significant effects on growth performance or animal health in finishing cattle. The 15% inclusion level demonstrated the greatest cost reduction potential. We, therefore, recommend adopting 15% BG supplementation as the optimal strategy to enhance economic returns in cattle production systems.

1. Introduction

Soybean meal (SBM) has traditionally served as the primary protein supplement in cattle diets due to its high crude protein content (44–48% dry matter basis) and favorable amino acid profile containing 2.8–3.2% lysine [1,2]. However, the increasing price volatility of SBM in global markets has created an urgent need to identify cost-effective alternative protein sources for sustainable cattle production.

Brewer’s grains (BG), a major by-product of the beer brewing industry [3], represent a promising nutritional and economical alternative. BG contains substantial protein content [4], with approximately 50% existing as rumen-undegradable protein (RUP) that bypasses microbial fermentation in the rumen [5,6]. RUP exhibits greater resistance to microbial degradation in the rumen, enabling its subsequent digestion in the abomasum and small intestine, thereby enhancing the utilization efficiency of BG-derived protein. Furthermore, BG is rich in dietary fiber, with a reported content of ~59% (natural matter basis) [7]. Rumen microorganisms can degrade dietary fiber into low-molecular-weight compounds. More importantly, BG is particularly rich in bioactive phenolic compounds, especially hydroxycinnamic acids—a structurally diverse class including ferulic, p-coumaric, caffeic, and sinapic acids—which demonstrate well-documented antioxidant, anticarcinogenic, antiatherogenic, and anti-inflammatory properties [8].

With China’s substantial beer production reaching 35.7 billion liters in 2022 (National Bureau of Statistics), BG is consistently available at prices significantly lower than SBM, making it both a nutritionally dense and economically viable feed ingredient [9].

Existing research has established BG’s effects on production parameters. Multiple studies confirm that dietary wet BG supplementation maintains the meat quality of finishing cattle [10] and preserves desirable carcass characteristics in Angus × Simmental cattle [4]. Meanwhile, Belon et al. [11] confirmed that early inclusion of wet BG supported the growth and carcass performance of Simmental-Angus beef calves. However, several critical knowledge gaps remain regarding BG’s functional properties.

The immunomodulatory effects appear inconsistent, with Moriel et al. [12] reporting that decreasing wet BG supplementation frequency or rate for recently weaned beef heifers restrained vaccine-induced antibody production against pathogens associated with bovine respiratory disease during a 42-day preconditioning period. As for antioxidant capacity, evidence from poultry studies indicates that distillers dried grains with solubles, a cognate brewing by-product, enhances systemic antioxidant responses in broilers [13], suggesting the antioxidant capacity of BG warranting validation in cattle.

Furthermore, although rumen microbiome studies are abundant, analysis of fecal microbiota—which provides crucial insights into hindgut fermentation efficiency and overall gastrointestinal health [14]—reveals that 20% inclusion of reduced-fat dried distillers’ grains with solubles increases microbial diversity metrics (richness, evenness, and Shannon index) while maintaining community structure in dairy cattle [15], suggesting potential gut health benefits of brewing by-products.

We hypothesize that the inclusion of 10–15% BG as an alternative protein source in diets for Simmental crossbred cattle improves the health of confined animals and reduces production costs. This study systematically evaluates the effects of 10% and 15% BG dietary inclusion on the following: (1) growth performance, (2) nutrient digestibility, (3) immune function, (4) antioxidant capacity, and (5) fecal microbiome composition in finishing Simmental crossbred cattle. Through comprehensive economic analysis, we aim to establish evidence-based recommendations for partial or complete replacement of SBM with BG in finishing diets.

2. Materials and Methods

2.1. Animal Ethics Statement

All experimental procedures were approved by the Institutional Animal Care and Use Committee of Fujian Agricultural and Forest University (Fuzhou, Fujian, China). (Approval ID: PZCASFAFU23042).

2.2. Animals, Treatments, and Management

The animal experiment was conducted in the winter of 2023 at Zhangzhou Kanghong Agriculture and Animal Husbandry Co., Ltd., Fujian, China.

The experiment was conducted using a completely randomized design. Twenty-four 15-month-old healthy Simmental crossbred male cattle (body weight, 433.43 ± 32.47 kg) were randomly assigned to three treatment groups with eight replicates per treatment. The 3 treatment groups were designated as follows: control group (CON) (fed basal ration), 10% BG group (BG10) (fed basal ration with 10% BG on a dry matter basis), and 15% BG group (BG15) (fed basal ration with 15% BG on a dry matter basis). The 10% and 15% BG levels were chosen through preliminary trials: 10% represented a conservative substitution rate aligning with industry practices, while 15% tested the upper limit before observed palatability decline (dry matter intake reduction > 8% in pre-trial). The trial lasted 48 days after a 3-day pre-feeding period.

The wet BG was provided by Anheuser-Busch InBev Sedrin Brewery Co., Ltd. (Zhangzhou, China), and the nutrient levels of the wet brewer’s grains are listed in

Table 1. Nutrient levels of brewer’s grains (%, dry matter basis).

Items	Nutrient Levels
Dry matter	17.83
Crude protein	21.30
Neutral detergent fiber	38.83
Acid detergent fiber	21.84
Calcium	0.31
Phosphorus	0.39

.

All cattle were provided diets formulated to meet nutritional requirements, administered twice daily at 08:30 and 16:00 h, with ad libitum access to fresh water throughout the trial period. Cattle received 27.5–32.5 kg of feed (as-fed basis) daily per animal, with residual orts accounting for 5–10% of the offered feed. Dry matter intake (DMI) was calculated based on the difference between offered feed and orts adjusted for dry matter content. Cattle were vaccinated against foot-and-mouth disease virus and bovine respiratory syncytial virus, with individual animals identified through ear tags to enable systematic health monitoring throughout the trial.

The basal diet was formulated to meet the nutritional requirements specified in the Chinese Feeding Standard of Beef Cattle (NY/T 815–2004) [16] for finishing cattle with a target body weight of 450 kg and a daily gain of 1.2 kg.

Peanut straw and wheat straw were included as roughage sources in the diet and mechanically crushed to a particle size of 2–3 cm using a hammer mill prior to feeding. The composition and nutrient levels (dry matter basis) of experiment diets are described in

Table 2. Ingredients and nutrient levels of experiment diets (%, dry matter basis).

Items	CON	BG10	BG15
Ingredients
Peanuts straw	31.20	30.20	25.90
Wheat straw	15.30	9.80	9.90
Ground corn	41.70	44.40	46.30
Wet brewer’s grains	0.00	10.00	15.00
Soybean meal	8.90	2.70	0.00
Sodium bicarbonate	1.00	1.00	1.00
Calcium hydrogen phosphate	0.20	0.20	0.20
Salt	0.30	0.30	0.30
Premix 1	1.40	1.40	1.40
Total	100.00	100.00	100.00
Nutrient levels 2
NEmf/(MJ/d)	65.88	65.31	64.73
Crude protein	12.54	12.92	12.86
Neutral detergent fiber	46.77	48.03	48.44
Acid detergent fiber	24.52	25.96	26.94
Calcium	0.69	0.71	0.66
Phosphorus	0.24	0.27	0.26

Note: CON = control group; BG10 = 10% BG group; BG15 = 15% BG group; NE_mf = combined net energy. ¹ The premix provided the following per kilogram of the basal ration: vitamin A, 4400 IU; vitamin D₃, 550 IU; tocopherol acetate, 180 mg; Fe, 50 mg; Zn, 30 mg; Mn, 20 mg; Cu, 10 mg; I, 0.5 mg; Se, 0.15 mg; Co, 0.1 mg. ² NE_mf was a calculated value, while the others were measured values. NE_mf was calculated as follows [16]: NE_mf = DE × (K_m × K_f × 1.5)/(K_f + 0.5 × K_m), where DE is the digestive energy of the feed (kJ/kg); K_m is the efficiency of converting digestive energy to net energy for maintenance; K_f is the efficiency of converting digestive energy to net energy for gain; and 1.5 is the feeding level coefficient.

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2.3. Growth Performance and Economic Benefit Analysis

Individual body weights (BWs) were measured following a 12 h fasting period at trial initiation and termination. Average daily gain (ADG) was calculated as total body weight gain (TWG, final BW minus initial BW) divided by the 48-day experimental period. DMI was calculated by subtracting residual orts from daily feed offerings, with quantities accurately measured on a dry matter basis. The feed conversion ratio (FCR) was computed as the ratio of DMI to ADG.

Economic benefit analysis was conducted by calculating total feed cost and cost-to-gain ratios. The feed price (dry matter basis) for each treatment was determined using real-time market prices of dietary ingredients and their inclusion rates during the trial period. The following formulas were applied:

Total feed cost (CNY per cattle/d) = feed price (CNY/kg) × DMI (kg/d)

Cost-to-gain ratio (CNY/kg/d) = total feed cost (CNY per cattle/d)/ADG (kg/d)

2.4. Sample Collection

Fecal samples were continuously collected over 24 h periods during the last 3-day period of the trial using harness-equipped collection bags, ensuring complete excreta recovery without temporal interruption. Following collection, daily fecal outputs were weighed, recorded, homogenized, and subsampled (20% of fresh weight per animal). Subsamples were stored at −20 °C for subsequent analysis. A total of 10 g of each fecal sample was collected and preserved at −80 °C in sterile cryovials for microbial 16S rRNA sequencing. Following trial completion, frozen samples were thawed and lyophilized at 65 °C for 48 h until constant mass was achieved, following established protocols [17]. BG, diet samples, and dried excreta were ground to pass through a 1 mm sieve for apparent nutrient digestibility analysis.

Following a 12 h fasting period, jugular venipuncture was performed using 10 mL evacuated tubes on all animals at trial termination. Blood samples were centrifuged by the model TGL-16B centrifuges (Shanghai Anting Scientific Instrument Factory, Shanghai, China) at 3000 rpm/min and 4 °C for 15 min, and then the supernatant was separated to obtain serum. Clarified serum was aliquoted into 1.5 mL microtubes and archived at −20 °C for serum biochemical parameters, immunity, and antioxidant capacity analysis.

2.5. Nutrient Analysis and Apparent Nutrient Digestibility Analysis

The nutrient composition of BG, diets, and fecal samples was analyzed following AOAC International standard protocols [18] and fiber analysis methods. Dry matter (DM) content was determined by oven drying at 105 °C (AOAC 934.01). Crude protein (CP) content was quantified via Kjeldahl nitrogen analysis (AOAC 984.13). Calcium (Ca) and phosphorus (P) concentrations were measured using atomic absorption spectroscopy (AOAC 968.08) and vanadomolybdate colorimetry (AOAC 965.17), respectively. Neutral detergent fiber (NDF) and acid detergent fiber (ADF) contents were quantified using an Ankom 2000 fiber analyzer (Ankom Technology, Macedon, NY, USA) following the sequential detergent protocol of Van Soest et al. [19].

Total fecal collection (i.e., the total collection method) was employed to calculate apparent nutrient digestibility. On day 46 of the main experimental period, a 3-day digestibility trial was initiated. Ingested dry matter (IDM) during the trial was calculated based on daily feed consumption data, and individual IDM per animal was determined. Additionally, fecal dry matter (FDM) content was quantified throughout the digestibility assessment period. The apparent nutrient digestibility was calculated as follows:

$$\mathrm{Apparent} \mathrm{nutrient} \mathrm{digestibility} (\%)={\displaystyle \frac{(IDM\times Nd)-(FDM\times Nf)}{IDM\times Nd}}\times 100$$

(1)

where IDM is the ingested dry matter (g), representing the total dry weight of feed consumed; Nd is the nutrient content in diet dry matter (%); FDM is the fecal dry matter output (g), representing the total dry weight of feces excreted; and Nf is the nutrient content in fecal dry matter (%).

2.6. Serum Biochemical Parameters Analysis

The serum biochemical parameters, including total protein (TP), albumin (ALB), glucose (GLU), blood urea nitrogen (BUN), triglyceride (TG), total cholesterol (TCHO), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), alanine transaminase (ALT), and aspartate transaminase (AST) were analyzed using the model BS-280 automatic biochemical analyzer (Mindray Bio-Medical Electronics Co., Ltd., Shenzhen, Guangdong, China). The content of globulin (GLB) was calculated as follows [20]:

GLB (g/L) = TP (g/L) − ALB (g/L)

2.7. Serum Immunity Indexes Analysis

Serum concentrations of interferon-γ (IFN-γ), immunoglobulin A (IgA), immunoglobulin G (IgG), tumor necrosis factor-α (TNF-α), interleukin-2 (IL-2), and interleukin-10 (IL-10) were quantified using bovine-specific ELISA kits (Shanghai Enzyme-linked Biotechnology Co., Ltd., Shanghai, China) according to manufacturer protocols. Kit catalog numbers were as follows: IFN-γ (IFN-γ-ID: YJ002465), IgA (IgA-ID: YJ542063), IgG (IgG-ID: YJ330698), TNF-α (TNF-α-ID: YJ077389), IL-2 (IL-2-ID: YJ633561), and IL-10 (IL-10-ID: YJ506601).

Briefly, 50 µL of calibrators and test samples were pipetted in duplicate into antibody-coated wells, followed by the addition of 100 µL horseradish peroxidase (HRP)-conjugated detection antibody. Plates were emptied and washed 5 times after incubating at 37 °C for one hour. Subsequently, 100 µL of the chromogenic substrate was dispensed per well, with plates incubated in light-protected conditions for 15 min at room temperature. The reaction was terminated with 50 µL 2 M sulfuric acid per well, and optical density (OD) values were immediately measured at 450 nm using a Synergy-H1 microtitre plate reader (BioTek Instruments, Inc., Winooski, VT, USA). Analyte concentrations were determined via four-parameter logistic regression of standard curves using BioTek Gen5 software (v3.11).

2.8. Serum Antioxidant Indexes Analysis

Serum total antioxidant capacity (T-AOC) and malondialdehyde (MDA) concentrations, along with glutathione peroxidase (GSH-Px) and total superoxide dismutase (T-SOD) activities, were quantified using bovine-specific commercial assay kits (Nanjing Jiancheng Bioengineering Institute, Nanjing, Jiangsu, China). Kit catalog numbers were as follows: T-AOC (T-AOC-ID: A015-1-2), MDA (MDA-ID: A003-1-1), GSH-Px (GSH-Px -ID: A005-1-2), and T-SOD (T-SOD-ID: A001-1-1).

To ensure measurement accuracy, pre-experimental optimization was performed to establish appropriate sample dilution ratios. Specifically, T-AOC was assessed via the ABTS [2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)] radical cation decolorization assay. MDA quantification employed the thiobarbituric acid reactive substances (TBARS) method. GSH-Px activity was determined by monitoring glutathione oxidation coupled with 5,5′-dithiobis (2-nitrobenzoic acid) (DTNB) reduction at 412 nm. T-SOD activity was evaluated through the hydroxylamine oxidation inhibition method. Assays were run following the kits’ instructions. T-AOC plates were read at 520 nm, MDA plates were read at 532 nm, GSH-Px plates were read at 412 nm, and T-SOD plates were read at 550 nm, both using a model UV-1800PC UV–Visible spectrophotometer (Shanghai Mapada Instruments Co., Ltd., Shanghai, China).

2.9. Bacterial DNA Extraction and 16S rRNA Gene Sequencing

Fecal bacterial genomic DNA was extracted using Stool DNA Kit (D4015-01, Omega Bio-tek, Norcross, GA, USA). The hypervariable V3–V4 regions of bacterial 16S rRNA genes were amplified using primer pair 338F (5′-ACTCCTACGGGAGGCAGCAG-3′) and 806R (5′-GGACTACHVGGGTWTCTAAT-3′) on an ABI GeneAmp 9700 PCR thermocycler (Foster City, CA, USA) and subjected to 2 × 250 bp paired-end sequencing on an Illumina NextSeq 2000 platform (Illumina, San Diego, CA, USA) following the manufacturer’s sequencing-by-synthesis protocol, with library preparation and bioinformatics processing performed by Majorbio Bio-Pharm Technology Co. Ltd. (Shanghai, China).

Raw sequences were denoised into Amplicon Sequence Variants (ASV) at 99% identity using DADA2′s divisive partitioning algorithm. Alpha diversity metrics (Ace, Chao1, Shannon) were computed in Mothur v1.30.1 [21], while beta diversity patterns were visualized via Bray–Curtis-based principal coordinate analysis (PCoA) ordination. Inter-group microbiota dissimilarities were statistically validated through Analysis of Similarities (ANOSIM). Taxon-specific abundance variations (phylum/genus) were identified via Kruskal–Wallis tests with Tukey’s correction (p < 0.05).

All raw sequencing data have been deposited in the NCBI Sequence Read Archive under the BioProject PRJNA1184030.

2.10. Statistical Analysis

The experimental data were initially processed using Excel 2021 and analyzed by SPSS statistical software (Version 26.0 for Windows). To account for potential confounding effects of baseline body weight, growth performance parameters were assessed via analysis of covariance (ANCOVA) using initial body weight as a covariate, with adjusted outcomes expressed as estimated marginal means with standard error of the mean (SEM). The statistical model is expressed as follows:

$$Y_{\mathrm{ij}}=\mathsf{\mu }+{\mathsf{\alpha }}_i+\mathsf{\beta } (x_{\mathrm{ij}}-\overline{x})+e_{ij}$$

where Y_ij is the dependent variable for the j-th cattle in the i-th treatment group; μ is the overall mean; α_i is the fixed effect of the i-th dietary treatment; β is the regression coefficient for the covariate; (x_ij − $\overline{x}$) is the covariate adjusted for its overall mean to center the data; and e_ij is the random error term.

Except for growth performance parameters, a one-way analysis of variance (ANOVA) was conducted for other response variables, followed by Duncan’s multiple-range tests for post hoc comparisons. Statistical significance was set at p < 0.05, with trends toward significance defined as 0.05 ≤ p < 0.10. All data are expressed as means with SEM in the respective tables unless otherwise stated. The statistical model is expressed as follows:

Y_ij = μ + α_i + e_ij

where Y_ij is the dependent variable for the j-th cattle in the i-th treatment group; μ is the overall mean; α_i is the fixed effect of the i-th treatment; and e_ij is the random error term.

3. Results

3.1. Growth Performance and Economic Benefit

Following covariance adjustment (ANCOVA) with initial body weight as a covariate, no significant differences (p > 0.05) were detected in TWG, ADG, DMI, or FCR among the three treatment groups (

Table 3. Effects of brewer’s grains on growth performance and economic benefit of Simmental crossbred cattle.

Items	CON	BG10	BG15	SEM	p-Value
Growth performance
TWG/kg	78.85	67.99	87.03	7.21	0.266
ADG/(kg/d)	1.64	1.42	1.77	0.15	0.343
DMI/(kg/d)	14.02	14.00	13.76	0.15	0.479
FCR	8.84	10.14	8.35	0.83	0.371
Economic benefit
Feed price/(CNY/kg)	2.08	1.89	1.82	-	-
Total feed cost/(CNY per cattle/d)	29.18 c	26.42 b	24.91 a	0.41	<0.01
Cost-to-gain ratio/(CNY/kg/d)	18.35	19.04	15.17	0.85	0.143

Note: CON = control group; BG10 = 10% BG group; BG15 = 15% BG group; SEM = standard error of the mean; TWG = total weight gain; ADG = average daily gain; DMI = dry matter intake; FCR = feed conversion ratio = DMI/ADG. ^a–c mean values within the row, unlike superscript letters, were significantly different (p < 0.05).

).

According to the market conditions during the trial period, the price of feedstuffs was as follows: BG: CNY 500 per ton; soybean meal: CNY 4000 per ton; peanuts straw: CNY 1000 per ton; wheat straw: CNY 950 per ton; corn: CNY 2800 per ton. Based on the prices of all feed ingredients, the feed prices of 0%, 10%, and 15% BG diet were, on average, CNY 2.08, CNY 1.89, and CNY 1.82 kg (DM basis), respectively. While BG supplementation showed no significant effects on cost-to-gain ratio across treatment groups (p > 0.05), it induced dose-dependent reductions in total feed cost. The BG10 and BG15 group formulations demonstrated 9.46% and 14.63% lower feed cost, respectively, compared to the CON group (p < 0.01), with BG15 achieving the most economical pricing profile (Table 3).

3.2. Apparent Nutrient Digestibility

There were no obvious differences in the digestibilities of DM, CP, NDF, and Ca with increasing supplementation levels of BG (p > 0.05); notably, phosphorus digestibility exhibited a trend toward improvement in the BG10 group relative to the CON group (p = 0.069). The digestibility of ADF in the BG15 group was elevated as compared to the CON group (p < 0.01) (

Table 4. Effects of brewer’s grains on apparent nutrient digestibility of Simmental crossbred cattle.

Items	CON	BG10	BG15	SEM	p-Value
Dry matter/%	57.08	57.17	57.02	0.49	0.993
Crude protein/%	43.67	44.65	45.76	0.66	0.458
Neutral detergent fiber/%	50.15	50.86	52.46	0.60	0.296
Acid detergent fiber/%	50.90 a	55.64 ab	60.95 b	1.32	<0.01
Calcium/%	45.59	40.14	51.48	3.00	0.334
Phosphorus/%	48.66	57.45	53.13	1.65	0.069

Note: CON = control group; BG10 = 10% BG group; BG15 = 15% BG group; SEM = standard error of the mean. ^a–c mean values within the row, unlike superscript letters, were significantly different (p < 0.05).

).

3.3. Serum Biochemical Parameters

Compared with the CON group, the content of ALB in the BG10 group and BG15 group were elevated on day 24 (p = 0.028) and day 48 (p = 0.047), respectively. However, the contents of serum GLU, TG, TCHO, HDL-C, LDL-C, BUN, AST, ALT, TP, and GLB were similar between three groups on both day 24 and day 48 (p > 0.05) (

Table 5. Effects of brewer’s grains on serum biochemical parameters of Simmental crossbred cattle.

Items	CON	BG10	BG15	SEM	p-Value
Day 24
GLU/(mmol/L)	4.06	4.36	4.34	0.08	0.168
TG/(mmol/L)	0.21	0.23	0.23	0.02	0.897
TCHO/(mmol/L)	2.93	3.26	3.50	0.17	0.422
HDL-C/(mmol/L)	1.37	1.34	1.40	0.09	0.974
LDL-C/(mmol/L)	0.51	0.66	0.70	0.05	0.298
BUN/(mmol/L)	3.53	3.46	3.18	0.20	0.773
AST/(U/L)	88.17	92.00	92.60	2.37	0.726
ALT/(U/L)	25.00	26.80	25.00	0.97	0.722
TP/(g/L)	76.40	77.30	75.20	1.70	0.901
ALB/(g/L)	29.28 a	32.40 b	30.66 ab	0.51	0.028
GLB/(g/L)	47.12	44.90	44.54	1.81	0.832
Day 48
GLU/(mmol/L)	4.14	4.28	4.29	0.08	0.705
TG/(mmol/L)	0.28	0.30	0.27	0.01	0.543
TCHO/(mmol/L)	3.19	3.22	3.75	0.13	0.145
HDL-C/(mmol/L)	1.41	1.30	1.58	0.06	0.157
LDL-C/(mmol/L)	0.63	0.70	0.77	0.04	0.271
BUN/(mmol/L)	3.64	3.24	3.59	0.20	0.711
AST/(U/L)	81.75	86.00	76.14	2.54	0.314
ALT/(U/L)	31.50	31.57	30.43	0.83	0.837
TP/(g/L)	77.15	74.79	77.86	0.97	0.432
ALB/(g/L)	29.31 a	30.87 ab	32.24 b	0.50	0.047
GLB/(g/L)	47.84	43.91	45.61	1.07	0.332

Note: CON = control group; BG10 = 10% BG group; BG15 = 15% BG group; SEM = standard error of the mean; GLU = glucose; TG = triglyceride; TCHO = total cholesterol; HDL-C = high-density lipoprotein cholesterol; LDL-C = low-density lipoprotein cholesterol; BUN = blood urea nitrogen; AST = aspartate aminotransferase; ALT = alanine transaminase; TP = total protein; ALB = albumin; GLB = globulin. ^a–c mean values within the row, unlike superscript letters, were significantly different (p < 0.05).

).

3.4. Serum Immunity Indexes

No significant differences in IFN-γ, Ig-A, Ig-G, IL-2, IL-10, and TNF-α contents were observed between the three groups (p > 0.05) on day 24. By day 48, however, serum IFN-γ levels in the BG15 group exhibited a tendency toward reduction (p = 0.069) (

Table 6. Effects of brewer’s grains on immunity of Simmental crossbred cattle.

Items	CON	BG10	BG15	SEM	p-Value
Day 24
IFN-γ/(pg/mL)	1101.26	1210.41	1098.12	28.91	0.202
IgA/(g/L)	11.89	12.76	11.56	0.28	0.208
IgG/(g/L)	22.91	25.06	22.22	0.72	0.262
IL-2/(pg/mL)	240.76	243.16	255.03	6.14	0.625
IL-10/(pg/mL)	25.80	26.69	27.97	0.65	0.416
TNF-α/(pg/mL)	134.29	146.35	131.63	3.61	0.215
Day 48
IFN-γ/(pg/mL)	924.55	960.82	775.20	35.60	0.069
IgA/(g/L)	14.78	16.92	14.44	0.54	0.109
IgG/(g/L)	23.46	23.00	22.33	0.46	0.629
IL-2/(pg/mL)	210.50	258.79	223.14	13.49	0.337
IL-10/(pg/mL)	29.23	30.40	25.15	1.42	0.304
TNF-α/(pg/mL)	117.96	116.23	127.90	3.53	0.368

Note: CON = control group; BG10 = 10% BG group; BG15 = 15% BG group; SEM = standard error of the mean; IFN-γ = interferon-γ; IgA = immunoglobulin A; IgG = immunoglobulin G; IL-2 = interleukin 2; IL-10 = interleukin 10; TNF-α = tumor necrosis factor alpha. ^a–c mean values within the row, unlike superscript letters, were significantly different (p < 0.05).

).

3.5. Serum Antioxidant Indexes

The contents of serum T-AOC, MDA, GSH-Px, and T-SOD were not affected by BG treatments on both day 24 and day 48 (p > 0.05) (

Table 7. Effects of brewer’s grains on antioxidant capacity of Simmental crossbred cattle.

Items	CON	BG10	BG15	SEM	p-Value
Day 24
T-AOC/(U/mL)	1.95	1.94	1.90	0.05	0.931
MDA/(nmol/mL)	1.60	1.64	1.34	0.17	0.759
GSH-Px/(U/mL)	266.19	306.88	268.19	18.60	0.631
T-SOD/(U/mL)	68.95	71.62	69.96	1.94	0.866
Day 48
T-AOC/(U/mL)	2.05	2.22	0.84	0.27	0.108
MDA/(nmol/mL)	1.07	1.31	1.47	0.16	0.628
GSH-Px/(U/mL)	349.26	264.90	219.79	43.11	0.479
T-SOD/(U/mL)	69.33	68.75	75.18	1.27	0.105

Note: CON = control group; BG10 = 10% BG group; BG15 = 15% BG group; SEM = standard error of the mean; T-AOC = total antioxidant capacity; MDA = malondialdehyde; GSH-Px = glutathione peroxidase; T-SOD = total superoxide dismutase. ^a–c mean values within the row, unlike superscript letters, were significantly different (p < 0.05).

).

3.6. Fecal Microbiota Community

The sequence coverage reached 99.60%, 99.79%, and 99.76% for the CON, BG10, and BG15 groups, respectively, demonstrating adequate sequencing depth.

As displayed in Figure 1A, the alpha diversity analysis of fecal microbiota revealed that the BG15 group exhibited a tendency toward reduced Ace index compared to the CON group (p = 0.064), along with a significantly lower Chao index (p = 0.040). Additionally, the Shannon index in the BG15 group showed a marginal decrease relative to the BG10 group (p = 0.053). As indicated in Figure 1B, there was overlap among the three groups through the beta diversity analysis of cattle fecal samples in each group by PCoA, and the CON group fecal microbiota community was quite different. The contribution rate was 25.84% for the first principal component (PC1) and 20.58% for the second principal component (PC2).

At the phylum level (Figure 1C), the fecal microbiota community in cattle was dominated by Firmicutes, Actinobacteria, Proteobacteria, Bacteroidota, and Verrucomicrobiota and the proportions of these five phyla accounted for more than 99%. At the genus level (Figure 1D), Lactobacillus, Romboutsia, Paeniclostridium, Clostridium_sensu_stricto_1, Turicibacter, Weissella, Bifidobacterium, Candidatus_Saccharimonas, and Prevotella were dominant genera.

The relative abundance of Romboutsia in the BG15 group was significantly higher than in the CON group (p = 0.025). Meanwhile, the relative abundance of Clostridium_sensu_stricto_12 (p = 0.041) and Rummeliibacillus (p = 0.020) in the BG10 group was significantly higher than in the BG15 group (

Table 8. Effects of brewer’s grains on fecal bacterial community of Simmental crossbred cattle.

Items	CON	BG10	BG15	SEM	p-Value
Romboutsia/%	10.04 a	12.76 ab	20.29 b	1.65	0.025
Clostridium_sensu_stricto_12/%	0.74 ab	1.97 b	0.46 a	0.26	0.041
Rummeliibacillus/%	0.45 ab	0.69 b	0.11 a	0.21	0.020

Note: CON = control group; BG10 = 10% BG group; BG15 = 15% BG group; SEM = standard error of the mean. ^a–c mean values within the row, unlike superscript letters, were significantly different (p < 0.05).

).

4. Discussion

4.1. Growth Performance and Economic Benefits

Feed intake, a vital indicator of animal growth performance, is affected by many factors, such as palatability, feeding management, and environment. A multitude of studies have substantiated that the aroma emitted from beer grains can improve feed palatability and elevate the dry matter intake (DMI) of animals [22], both heifers [23] and lactating cows [24]. However, the impact of brewer’s grains (BG) on feed intake is diverse, stemming from variations in dosage, sources, nutrient levels, moisture content, and processing methods of BG.

Similar to our results, no notable differences in DMI of dairy cows were reported by Belibasakis and Tsirgogianni [25] and Wen-Shyg Chiou et al. [26] when supplementing 16% or 10% wet BG, while the outcome was unfavorable once the dosage reached 40% or more. Davis et al. [27] observed a decrease in DMI when 40% pressed BG was fed to lactating dairy cows. Meanwhile, Homm et al. [4] reported significantly lower DMI in beef heifers fed 45% BG compared to 15–30% inclusion groups.

This phenomenon may be attributable to the combined effects of the high moisture content (above 70%) [28], high polysaccharide, and protein content in diets when high-dose wet BG was supplemented, which promotes rapid microbial proliferation and feed spoilage [29], thus resulting in DMI decrease. On the other hand, the unbalance of amino acids and minerals in BG also hinders the high-dose addition of BG as a feedstuff in ruminant production [30,31]. BG includes leucine, valine, alanine, serine, glycine, glutamic acid, and aspartic acid in the largest amounts, but there is a lack of lysine, the first limiting essential amino acid [30]. Accordingly, meticulous consideration of the dosage is imperative when administering BG.

Additionally, our data demonstrated that there were no obvious differences not only in DMI but also in average daily gain (ADG) and feed conversion ratio (FCR) when feeding 0%, 10%, and 15% BG to finishing cattle. Consistent with our study, Parmenter et al. [32] observed that ADG did not show an obvious difference when 8% of BG was fed to finishing cattle. Analogously, Manthey et al. [33] reported that no differences in FCR were observed when limit-fed distillers dried grains at 0.8% of body weight were fed to dairy heifers. All the above results suggested that low-dose BG (at least no more than 15%) in diets could not affect the growth performance of cattle.

From an economic standpoint, feeding diets with 15% BG to cattle would be more economical than diets with conventional protein feedstuffs like soybean meal. The market price of BG during the trial was around one-eighth of soybean meal, causing the cost of the 10% and 15% BG diet lower than the CON diet in our study.

Compared to the control (CON) group, the BG10 and BG15 groups reduced dietary costs by 9.46% (CNY 29.18 vs. CNY 26.42 per cattle/d) and 14.63% (CNY 29.18 vs. CNY 24.91 per cattle/d), respectively. Notably, the BG15 group achieved a 17.33% lower cost-to-gain ratio than CON (CNY 15.17 vs. CNY 18.35/kg/d) and a 20.33% reduction compared to BG10 (CNY 15.17 vs. CNY 19.04/kg/d), establishing it as the most economically viable formulation.

4.2. Apparent Nutrient Digestibility

We further detected the digestibility of nutrients, which is a direct reflection of the feed’s utilization by animals and serves as a crucial index for evaluating the nutritional value of the feed. Our experiment found no differences in the digestibility of dry matter (DM), crude protein (CP), and neutral detergent fiber (NDF) with the BG supplementation. Gurung et al. [34] also observed that the digestibilities of DM and NDF were affected when 12.7%, 25.4%, and 38.1% distillers dried grains with solubles (DDGS) in diets were fed to Boer × Spanish castrated male goats.

Notably, phosphorus (P) digestibility showed a marginally significant improvement tendency in the BG10 group compared to the CON group, which means a higher utilization rate and less excretion of P with the dietary supplementation of BG in cattle. P is mostly located in bones and teeth and is a macro mineral supplemented to beef cattle for growth, production, and reproduction, involved in the maintenance of acid–base and osmotic balance [35] and participating in energy metabolism as a component of ATP, ADP, and AMP [36].

It is noteworthy that the 15% BG treatment could improve the digestibility of acid detergent fiber (ADF) in our trial. ADF comprises recalcitrant structural components, including cellulose, lignin, and silica [37]. The elevated ADF digestibility observed in the BG15 group may be attributed to the structural composition of BG, which contains fermentable fiber components (e.g., lignin and cellulose) that promote microbial degradation in the rumen.

Emerging evidence indicates that Romboutsia possesses fibrolytic capacity to degrade complex carbohydrates into volatile fatty acids (VFAs) while also fermenting amino acids such as histidine and threonine to support intestinal homeostasis [38]. This aligns with our findings of increased Romboutsia abundance (a genus linked to fiber utilization) in the fecal microbiota of the BG15 group, suggesting enhanced hindgut fermentation efficiency.

4.3. Serum Biochemical Parameters, Immunity, and Antioxidant Capacity

Serum biochemical parameters serve as biomarkers for systemic metabolic homeostasis and tissue/organ dysfunction, providing critical insights into physiological health status [39]. The concentrations of serum total protein (TP), albumin (ALB), globulin (GLB), glucose (GLU), and blood urea nitrogen (BUN) are crucial to trace protein and energy metabolism, while triglyceride (TG), total cholesterol (TCHO), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) are remarkable indicators of lipid digestion and metabolism [40].

Gurung et al. [41] observed that the addition of DDGS did not affect the concentrations of serum BUN in goats. Another study reported that no discrepancies were observed in ALB, GLU, TCHO, TG, LDL-C, aspartate aminotransferase (AST), and alanine transaminase (ALT) when Korean rice wine residue was fed to Hanwoo steers [42]. Consistent with these studies, no significant differences in most biochemical parameters were found between the three groups in our study, indicating that BG supplementation did not modify the body metabolism of cattle in general.

The ALB content on day 24 and day 48 rose after 10% and 15% BG treatments. ALB regulates plasma oncotic pressure, intravascular volume, and hydrophobic ligand transport. Meanwhile, as a negative acute-phase protein, hypoalbuminemia signals hepatic dysfunction [43]; however, the ALB levels in each group were in the normal range, suggesting no abnormality in liver metabolism.

We further detected immunologic and antioxidant indexes as distillers grains were reported to enhance the immunity and antioxidant capacity of animals according to existing studies [44,45]. Our results demonstrated that serum immunoglobulin A (IgA), immunoglobulin G (IgG), interleukin 2 (IL-2), interleukin 10 (IL-10), and tumor necrosis factor alpha (TNF-α) levels were not influenced by BG treatments; however, interferon-γ (IFN-γ) concentrations in the BG15 group exhibited a downward trend (p = 0.069) by day 48 of supplementation.

IFN-γ, a proinflammatory cytokine secreted by Th1 lymphocytes, activates macrophage function and orchestrates cell-mediated immune responses [46]. In line with our results, Alizadeh et al. [47] discovered that serum IFN-γ dropped notably when DDGS was fed to broiler chickens. Whilst Weber et al. (2011) [48] demonstrated elevated plasma concentrations of IgA and IgG in distillers dried grains with solubles (DDGS)-fed swine. Earlier work by the same research group (Whilst Weber et al., 2008) [49] revealed that 7-day DDGS supplementation upregulated ileal gene expression of IL-10, IL-6, and IL-1β in pigs, while TNF-α and IFN-γ expression remained unaltered. Therefore, the observed reduction in serum IFN-γ concentrations in cattle points to a potential immunomodulatory effect of BG, potentially mediated through Th1 pathway modulation. While this speculative association warrants validation, targeted mechanistic studies should investigate BG’s influence on cytokine networks and lymphocyte populations in Simmental crossbred cattle.

The imbalance between the generation of radical species and the antioxidant defense system will result in the progression of oxidative stress, causing damage to the structure and function of cells and increasing the risk of animal infection [50,51]. Total antioxidant capacity (T-AOC), quantifying oxidant neutralization per liter of biofluid, serves as a key metric for oxidative stress severity [52]. Malondialdehyde (MDA) is the end product of lipid peroxidation and occurs during the formation of free oxygen radicals or when arachidonic acid is present. While some antioxidative biomarkers, such as glutathione peroxidase (GSH-Px) and total superoxide dismutase (T-SOD), can inhibit lipid peroxidation by scavenging excessive free radicals in the body [53,54].

Up to now, research about BG’s effect on the antioxidant capacity of cattle remains scarce, merely with a few studies focusing on DDGS. Li et al. [1] reported significantly lower MDA levels in dairy cows fed pumpkin seed cake and DDGS versus soybean meal-based diets. However, the diet supplemented with DDGS was reported to decrease serum T-AOC and T-SOD levels but heighten serum MDA content in 21-day-old broilers [55]. In our study, the contents of serum T-AOC, MDA, GSH-Px, and T-SOD did not differ between groups, implying that BG did not exert any impact on the antioxidant capacity of Simmental crossbred cattle.

4.4. Fecal Microbiota Community

We subsequently evaluated the effects of BG on the fecal microbiota of cattle. Except for predominant rumen fermentation, the hindgut contributes approximately 27% of daily cellulose digestion and 40% of hemicellulose degradation, and the resultant VFA production can account for 8 to 17% of the total produced daily [14].

Our study confirmed that the alpha diversity of fecal microbiota was influenced by BG treatments, and the Chao index decreased with increasing supplementation levels of BG. However, different from our results, Shen et al. [56] observed that replacing soybean meal with DDGS did not affect the alpha diversity of fecal microbiota in growing Hu lambs. Alpha diversity metrics quantify microbial community complexity and stability, serving as biomarkers for host–microbiome homeostasis [57]. The disparity in microbiota community structure among different studies may be due to differences in the addition level of distillers grains and the chemical composition of the diets [58].

Although the results of alpha diversity were different from others, the microbiome of cattle was dominated by Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes in our study, which is consistent with previous reports [59]. Among them, Firmicutes occupied the preponderant share, and the proportions of this phyla accounted for 70 to 80%. In ruminants, Firmicutes promotes the degradation of fiber and cellulose into VFA, thereby facilitating food digestion and promoting animal growth [60].

At the genus level, Romboutsia was the dominant genus in our study, and its relative abundance in the BG15 group was higher than in the CON group. Romboutsia, a Gram-positive coccoid genus, is associated with intestinal mucosal health in humans [61]. Recent studies uncover that Romboutsia mainly uses monosaccharides and disaccharides to produce VFAs and can ferment amino acids, like histidine, threonine, and serine, to regulate the luminal health of cattle [62,63].

In our study, there was a higher relative abundance of Clostridium_sensu_stricto_12 and Rummeliibacillus in the BG10 group than in the BG15 group. Clostridium_sensu_stricto_12 is a bacterium with transferases of acetyl-coenzyme and butyrate kinase, which can generate n-butyric acid through the transformation of intracellular and extracellular acetate [64]. Rummeliibacillus, a facultatively aerobic Firmicutes member [65], demonstrates potent fibrinolytic activity [66]. Genomic predictions suggest Rummeliibacillus utilizes glycogen phosphorylase, glycogen branching protein, glycogen biosynthesis protein, and 4-alphaglucan branching enzyme to convert cellulose into glucose via glycogen intermediates [67]. In addition, Rummeliibacillus was also found to produce VFAs in accordance with Clostridium_sensu_stricto_12 [68].

Although hindgut fermentation contributes to plant fiber digestion, this process accounts for only a minor proportion (≤ 27% cellulose; ≤ 40% hemicellulose) compared to rumen-based degradation [14]. Direct comparisons between ruminal and fecal microbiota in cattle have revealed significantly greater microbial diversity and functional complexity in the rumen ecosystem [69,70]. Therefore, future studies should prioritize rumen microbiota analysis alongside fecal assessments to holistically evaluate BG’s effects on gastrointestinal health.

5. Conclusions

Dietary inclusion of brewer’s grains at 10–15% dry matter maintained growth performance and did not compromise health status in finishing cattle. Notably, 15% inclusion provided the greatest cost reduction. Hence, we recommend the inclusion of 15% brewer’s grains inclusion as the optimal strategy for maximizing economic returns in cattle production.