Application of Highly Digestible Fermented Corn Protein Powder in Fecal Low-Odor Adult Dog Feed

Abstract

This study aimed to develop fecal low-odor adult dog feed using processed corn protein powder as the primary raw material and evaluate its effectiveness through feeding experiments. The objectives included analyzing the fundamental nutritional indicators, comparing the quality of fecal low-odor adult dog feed with commercially available dog feed, and assessing the changes in the eating behavior, physical condition, and physiological and biochemical indicators before and after feeding on the fecal low-odor adult dog feed. This study involved formulating dog feed using processed fermented corn protein powder and conducting nutritional analyses to compare moisture, crude protein, ash, crude fat, and starch contents. Physical properties such as hardness and cohesion were also evaluated. Feeding experiments were conducted with adult dogs to monitor the changes in the eating behavior and physical condition. Physiological and biochemical indicators, including ammoniacal nitrogen and putrefactive odor in feces, were measured both before and after feeding to assess the impact of the fecal low-odor adult dog feed. The analysis showed no significant difference in moisture (p > 0.05), crude protein (p > 0.05), ash (p > 0.05), crude fat (p > 0.05), and starch (p > 0.05) contents between the fecal low-odor adult dog feed and commercially available dog feed. Similarly, no significant difference was noted in the hardness (p > 0.05) and cohesion (p > 0.05) between the two types of dog feed. Both types of dog feed contained rich volatile compounds with varying compositions. The commercially available dog feed had a sour odor, whereas the fecal low-odor adult dog feed had a barbecue and resin-like smell. After feeding with the fecal low-odor adult dog feed, the liquid nitrogen content significantly decreased (p < 0.01). Also, the indole content, a major contributor to fecal odor, based on the data, decreased after feeding.

1. Introduction

During pet feeding, metabolites often contain many putrid gases such as ammonia and hydrogen sulfide, which contribute to the unpleasant odor of feces [1]. The incomplete utilization of nitrogen in pet feed not only reduces its nutritional value, but also causes environmental pollution [2]. The primary contributors to fecal odor are compounds such as indole, organic nitrogen, and acids [3]. Improving the diet structure of pets can enhance digestion and nutrient utilization, thereby reducing the unused nitrogen content and odor production [4]. This also improves nutrient efficiency and reduces environmental pollution [5]. The digestibility of pet feed reflects how well pets can absorb and utilize the nutrients in their feed. High-digestibility feeds result in small, well-formed, and less frequent feces. In contrast, low-digestibility feeds contain many undigested components that reach the large intestine, where they undergo bacterial fermentation, producing odorous gases and resulting in loose stools or constipation [6]. Adding fermented corn protein powder to animal diets improved the growth performance and feed efficiency. For instance, it enhanced the daily weight gain and feed efficiency in weaned calves and promoted rumen fermentation and lamb growth [7]. Fermented corn protein powder is often used in poultry feed to improve protein digestibility and the amino acid balance. This leads to better growth rates, enhanced muscle development, and improved overall health of chickens [8]. In swine nutrition, fermented corn protein powder helps in improving the feed efficiency and growth performance. Pigs fed fermented corn protein exhibited better weight gain and feed conversion ratios [9]. Fermented corn protein powder is increasingly used in aquaculture feeds. It provides a high-quality, plant-based protein source which is more digestible than non-fermented alternatives for species such as tilapia, salmon, and shrimp. This results in better growth rates, higher survival rates, and the improved overall health of aquatic animals [10]. Despite these findings, research on the application of fermented corn protein powder in dog feed is limited. This study aimed to explore the use of fermented corn protein powder as the primary ingredient in dog feed and evaluate its impact on the nutritional value and health of pets. It analyzed the characteristics and benefits of this formulation, providing valuable insights for the pet feed industry, helping to meet the nutritional needs of dogs.

2. Materials and Methods

2.1. Materials

2.1.1. Preparation of Corn Protein Powder

Corn protein powder is the main by-product of corn wet starch processing. It is rich in various amino acids, such as alanine, serine, and leucine. It is a high-quality, high-yield protein raw material with high comprehensive utilization value. It is made from the crude starch milk obtained by wet grinding of corn kernels. The protein water, gluten water, is separated by a starch separator. It is concentrated by a concentration centrifuge or a sedimentation tank, and then dehydrated and dried. It is commonly known as yellow powder. The corn protein powder was subjected to composite treatment with papain (added at the dose of 400 U/g, temperature of 55 °C, and pH 6.0, for 3 h), acidic protease (added at a temperature of 40 °C, dose of 800 U/g, and pH 3.0, for 3 h), and Lactobacillus (fermentation at 37 °C, inoculation amount 4%, time 24 h, and solid–liquid ratio 1:4) to obtain corn protein powder through synergistic fermentation of bacteria and enzymes. After the composite treatment and fermentation of corn protein powder with papain, acidic protease, and Lactobacillus, a low-temperature drying process is used to preserve the activity of microorganisms. The mixture is first separated to remove excess liquid, resulting in a concentrated protein paste. This paste is then freeze-dried, a process that involves rapidly freezing the paste. This gentle method maintains the integrity of the active ingredients and microorganisms. The dried protein powder is then cooled to room temperature, milled into a fine powder, sieved for uniform particle size, and finally packaged in airtight containers to protect it from moisture and contamination. This process ensures the corn protein powder retains its nutritional and functional properties, making it suitable for applications requiring active microorganisms.

2.1.2. Preparation of Fecal Low-Odor Adult Dog Feed

Based on the requirements outlined in the national standard GB/T31216-2014 [11] for dog feed requirements, the dog feed formula was designed as follows: Fermented corn protein powder 40%, barley 8%, fish meal 10%, chicken 10%, wheat bran 7.2%, cooked soybean meal 14%, sheep bone meal 6%, salt 0.5%, multi-dimensional mineral—vitamins 0.3%, and fish oil 4%. All ingredients were mixed in a granulator (CPM 7932-8 California Pellet Mill, Crawfordsville, IN, USA) to prepare dog feed.

Commercially available dog feed (my foodie brand) was purchased from a pet store. The raw materials of commercially available dog feed include chicken meal (20%), oats (15%), cod (15%), cooked soybean meal (14%), fish oil (5%), whole-wheat bran (14%), beef bone meal (5%), sheep bone meal (6%), corn meal (3%), beet pulp (2%), salt (0.8%), and vitamins (0.2%).

2.2. Methods

2.2.1. Quality Evaluation of Fecal Low-Odor Adult Dog Feed and Commercially Available Dog Feed

Analysis of texture characteristics: Two types of dog feed with the same size and shape were analyzed using a texture analyzer. The analysis involved applying a 100 N force with a 75 mm-diameter disk compression probe. The pre-test speed was 0.25 mm/s, the test speed was 0.25 mm/s, and the post-test speed was 1 mm/s. The dwell time between the two compressions was 6 s, and the deformation was 60%. Texture analyzer, which records force, displacement, and energy values during sample compression, was used for quantitative analysis.

GC-MS (Gas Chromatography-Mass Spectrometry) analysis of volatile components in dog feed: Using a DB-WAX (Bonded Polyethylene Glycol) column (30 m × 0.25 mm × 0.25 μm), 0.35 g of the sample was placed in a 20 mL headspace bottle and sealed with a lid. The analysis utilized helium gas at a constant flow rate of 1 mL/min. The inlet temperature was set at 260 °C. The heating program began at 40 °C for 5 min, followed by an increase of 5 °C/min to 220 °C and then a rapid increase to 250 °C at 20 °C/min, which was maintained for 2.5 min. The GC-MS interface temperature was 260 °C, and the ion source temperature was 230 °C. Electron ionization (EI+) was used at 70 eV, with a full scan mode covering a mass range of 20–400 m/z. The data were analyzed using the NIST12014 spectral library to identify the volatile compounds.

2.2.2. Fecal Low-Odor Adult Dog Feed Experiment

Eight adult Chinese pastoral dogs (four male and four female, aged 3–4 years, weight: 4.5 ± 0.8 kg/dog) were selected for feeding and categorized into groups, each comprising two males and two females.

Each day, a dedicated person set out the enclosure for half an hour of activity and provided sufficient sunlight. A fixed amount of artificial feed (calculated as 4% of body weight) and free drinking water were provided twice daily (at 9:00 a.m. and 4:30 p.m.) for a 30-day feeding experiment. The use of animals for this study was approved by the Ethics Committee on Animal Use of Qiqihar University, China (012/2021).

2.2.3. Determination of Ammoniacal Nitrogen Content in Feces

The indophenol blue spectrophotometric method was used to determine the ammoniacal nitrogen content in feces. Reagents and solutions were prepared as follows: 0.3622 g of sodium nitroferricyanide was dissolved in deionized water to make a solution, and 5 g of phenol was dissolved in 400 mL of deionized water, then 2.0 mL of sodium nitroferricyanide solution was added and diluted to 500 mL (Solution A). A mixture of 2.50 g of NaOH, 2.0 g of trisodium citrate and 3.5 mL of NaClO was dissolved in deionized water and diluted to 500 mL (Solution B). Fecal samples were homogenized and 1 g of each sample was placed in a test tube. To each test tube, 5 mL of Solution A and 5 mL of Solution B were added and mixed thoroughly. The test tubes were incubated at room temperature for 15 min. The optical density (OD) of each sample was measured at a wavelength of 800 nm using a spectrophotometer. The concentration of ammoniacal nitrogen in each fecal sample was determined by comparing the OD values to a standard curve prepared using known concentrations of ammonium chloride solutions. Results were expressed as mg of ammoniacal nitrogen per gram of feces.

2.2.4. Determination of Apparent Digestibility of Grain Nutrients

Before concluding the feeding experiment, a metabolic test was conducted using the complete fecal collection method. And nitrogen fixation was performed on the feces using 10% dilute sulfuric acid. Dry matter was measured according to HJ1222-2021 [12], crude protein was measured according to GB/T24318-2009 [13], crude fat was measured according to GB/T6433-2006 [14], and neutral detergent fiber was measured according to NY/T1459-2022 Determination of Acid Washing Fiber [15], GB5009.92-2016 Determination of Calcium [16], and GB5009.87-2016 Determination of Phosphorus Index [17]. The apparent digestibility of nutrients was calculated.

2.2.5. Analysis of Fecal Spoilage Odor

GC-MS was used to analyze the changes in the fecal spoilage odor of experimental dogs. Feces from eating fecal low-odor adult dog feed group were taken on the morning of the first day and the last day of the experiment. The feces of all dogs participating in the experiment were mixed, and approximately 0.3 g was taken for analysis. They were heated in a boiling water bath for 15 min. The feces were centrifuged at 10,000 rpm for 10 min. Then, 1 mL of the supernatant was taken and divided into different centrifuge tubes. The contents were frozen in liquid nitrogen for 15 min and stored at an ultra-low temperature. The metabolites were separated and detected using Waters 2D UPLC (Ultra-Performance Liquid Chromatography, Milford, MA, USA) in series with a QExactive high-resolution mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA).

2.2.6. Statistical Analysis

The obtained data and images were processed using Excel 2019 and GraphPad Prism 10 software, and the data were subjected to repaired sample t-test using IBM SPSS Statistics 29 (p < 0.05). Each group of data was analyzed in triplicate.

3. Results

3.1. Basic Nutritional Index Analysis of Fecal Low-Odor Adult Dog Feed and Commercially Available Dog Feed

The contents of the main components of fecal low-odor adult dog feed and commercially available dog feed are shown in

Table 1. Analysis of main component content of fecal low-odor adult dog feed and commercially available dog feed.

Sample	Moisture (%)	Crude Protein (%)	Ash (%)	Crude Fat (%)	Starch (%)
Fecal low-odor adult dog feed	6.51 ± 0.14	46.27 ± 0.80	5.78 ± 0.07	10.56 ± 0.22	30.12 ± 0.32
Commercially available dog feed	6.77 ± 0.09	45.67 ± 0.29	5.66 ± 0.25	11.06 ± 0.49	30.66 ± 0.01

The p-values > 0.05 indicate no significant differences. No significant difference in the nutritional characteristics was found between fecal low-odor adult dog feed and commercially available dog feed.

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3.2. Quality Evaluation of Fecal Low-Odor Adult Dog Feed and Commercially Available Dog Feed

3.2.1. Analysis of Texture Characteristics

Texture characteristics are essential indicators for evaluating the taste of dog feed. This includes hardness, cohesion, chewiness, and elasticity. The hardness directly affects the taste of dog feed and is also related to cleaning dog teeth. The analysis of the texture characteristics of fecal low-odor adult dog feed and commercially available dog feed is presented in

Table 2. Analysis of texture characteristics of fecal low-odor adult dog feed and commercially available dog feed.

Sample	Hardness (N)	Cohesion (Ratio)	Chewiness (mj)	Elasticity (mm)
Fecal low-odor adult dog feed	45.40 ± 3.05	0.07 ± 0.01	13.57 ± 0.60	2.73 ± 0.02 *
Commercially available dog feed	43.66 ± 3.21	0.06 ± 0.06	16.79 ± 0.08 *	0.77 ± 0.03

*: Indicates a statistically significant difference (p < 0.05) between the two groups for that measurement.

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3.2.2. Gc-Ms Analysis of Volatile Components in Dog Feed

GC-MS analysis was conducted on the collected samples. For this, 0.35 g each of fecal low-odor adult dog feed and commercially available dog feed were taken for odor analysis, the volatile components in the samples were identified, and the peak area was used to represent the relative content of each component. The total ion chromatogram and the content of volatile components are shown in Figure 1 and

Table 3. Analysis of volatile components in fecal low-odor adult dog feed and commercially available dog feed.

Category	Name of Volatile Components	Aroma	Absolute Peak Area
			Commercially Available Dog Feed	Fecal Low-Odor Adult Dog Feed
Aldehyde	3-Methylbutanal	Barbecue flavor, malt flavor	107.24	431.55
	Hexanal	Avocado flavor	636.7	440.87
	Benzaldehyde	Almond flavor	34.15	141.54
	2-Heptanenal	Oil flavor, green fragrance	397.51	198.12
	1-Hexanol	Resin flavor, floral fragrance	18.28	130.97
Alcohols	1-Pentanol	Fruit aroma, tomato aroma	51.02	134.69
	2,3-Butanediol	Special fragrance	0.00	51.20
	2,3-Hexanediol	Special fragrance	0.00	62.51
	Polypropylene glycol	Special fragrance	91.81	12.89
Acids	Caproic acid	Sour	636.75	440.87
	4-Methylvaleric acid	Sour	7.75	79.89
	Propionic acid	Sour	460.38	37.87
Ketones	2 (5H)-furanone	–	5.82	69.55
	1-Hydroxy-2-acetone	–	3.75	592.57
	3-Hydroxy-2-butanone	–	0.00	198.65
	2-Methyl-3-hydroxy-γ-pyranone	Malt flavor	0.00	75.16
Alkanes	3-Methylhexane	Alkane odor	39.13	0.00
	2-Methylpentane	Alkane odor	630.69	0.00
	2,4-Dimethylheptane	Alkane odor	371.58	0.00
	2,6-Dimethyldecane	Alkane odor	0.00	39.11
	Furfural	Sweet and roasted taste	77.89	326.77
Other heterocycles	2-Pentylfuran	Green bean flavor, butter flavor	0.00	106.61
	2,3-Dihydro-3,5-dihydroxy-6-methyl-4(H)-pyran-4-one	–	0.90	43.10

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Based on the provided graph and data, the peak intensities illustrated the relative abundance of various volatile components before and after feeding. For commercially available dog feed (brown line), the five most abundant volatile components and their retention times were as follows: hexanal, with a retention time of around 5–6 min and an absolute peak area of 636.74, giving an avocado flavor; caproic acid, appearing around 6–7 min with an absolute peak area of 636.75, known for its sour odor; 2-methylpentane, appearing around 13–14 min with an absolute peak area of 630.69, contributing an alkane odor; 2,4-dimethylheptane, peaking around 14–15 min with an absolute peak area of 371.58, also with an alkane odor; and 2-heptanal, appearing around 7–8 min with an absolute peak area of 397.51, providing an oil flavor and green fragrance. In contrast, for fecal low-odor adult dog feed (green line), the five most abundant volatile components were as follows: 3-methylbutanal, with a retention time of around 4–5 min and an absolute peak area of 431.55, contributing a barbecue and malt flavor; hexanal, also appearing around 5–6 min with an absolute peak area of 440.87; 1-hydroxy-2-acetone, peaking at around 9–10 min with an absolute peak area of 592.57, with no specific odor characteristics; furfural, appearing around 10–11 min with an absolute peak area of 326.77, providing a sweet and roasted taste; and caproic acid, peaking at around 10–12 min with an absolute peak area of 440.87, with sour characteristics. These observations indicated a distinct variation in the composition and concentration of volatile components between commercial dog feed and low-fecal-odor dog feed.

This comprehensive analysis highlighted how the composition of volatile components differed significantly between the two types of dog feed, reflecting their respective odor profiles and potential impacts on canine sensory experiences. For instance, the higher levels of hexanal and caproic acid in commercial dog feed suggested a stronger avocado and sour odor profile. In contrast, there is a presence of 3-methylbutanal in low-fecal-odor dog feed. The stronger barbecue flavor indicates that the main aromatic compounds of the two are different.

3.2.3. Fecal Low-Odor Adult Dog Feed Feeding Experiment

During the experiment, the dogs fed with fecal low-odor adult dog feed were lively, vocal, and exhibited no abnormal behavior. After the experiment, their fur color became brighter, with no hair loss, skin diseases, or other issues. The physiological signs included no abnormal secretions in the five sense organs, anus, and other areas. Their body temperature remained average. The dogs exhibited healthy growth, showing no signs of distress or mortality. Their appetite was normal throughout the experiment, with no instances of overeating. Their feces were normal, with no loose stools. Blood routine tests and blood biochemistry analyses were essential indicators of the health status of the dogs. From these two graphs, Figure 2 and Figure 3, it can be concluded that the physiological and blood indicators of the dogs show no significant changes before and after the experiment. This suggests that the experimental process and the test feed did not have any noticeable adverse effects on the overall health status of the dogs.

3.2.4. Changes in Body Weight of Test Dogs before and after the Experiment

The feeding rate of the experimental dogs after 30 days of feeding resulted in a weight gain of 1.11 kg (Figure 4).

3.2.5. Analysis of Ammoniacal Nitrogen Content in Feces

The putrid odor in feces was mainly attributed to indole, hydrogen sulfide, ammonia, and other substances produced during animal metabolism. The ammoniacal nitrogen in feces generally refers to the ammonia gas and ammonium ions converted from urea. The change in the ammoniacal nitrogen content is also one of the factors influencing the odor of spoilage. The changes in the ammoniacal nitrogen content of feces before and after the experiment are shown in Figure 5.

3.2.6. Analysis of the Effect of Dog Feed on the Apparent Digestibility of Nutrients in Experimental Dogs

The changes in the apparent digestibility of various nutrients in commercially available dog feed and fecal low-odor adult dog feed are shown in

Table 4. Effects of dog food on nutrient apparent digestibility of dog.

Nutrient Apparent Digestibility (%)	Commercial Dog Feed	Fecal Low-Odor Adult Dog Feed
Dry Matter	80.01 ± 3.14	85.65 ± 1.55
Crude Protein	80.88 ± 4.38	89.01 ± 5.25
Crude Fat	70.47 ± 1.23	65.79 ± 2.31
Neutral Detergent Fiber	62.33 ± 2.51	62.41 ± 2.01
Acid Detergent Fiber	55.99 ± 0.18	50.26 ± 3.33
Calcium	40.93 ± 1.11	41.03 ± 2.30
Phosphorus	41.60 ± 0.58	41.26 ± 1.03

The p-values > 0.05, indicating no significant differences.

. It can be seen that there is no significant difference in the content of dry matter, crude protein, crude fat, neutral detergent fiber, acid detergent fiber, calcium, and phosphorus between the commercially available dog feed and the fecal low-odor adult dog feed.

3.2.7. Analysis of Fecal Spoilage Odor

GC-MS analysis was performed on the collected samples to obtain the mass spectra. These mass spectra were then compared and retrieved using the NIST (National Institute of Standards and Technology) library to identify the volatile components in the samples. The relative content of each component was represented by its peak area. The total ion chromatogram and the content of volatile odor components are shown in Figure 6 and

Table 5. Analysis of the content of fecal spoilage components before and after feeding on fecal low-odor adult dog feed.

Volatile Component	Odor Characteristics	Absolute Peak Area
		Before Feeding	After Feeding
Acetic acid	Spicy vinegar flavor	48.62	24.51
Caproic acid	Spicy vinegar flavor	12.95	0.00
Butyrate	Stimulating yogurt, buttery flavor	0.78	0.31
Valeric acid	Vinegar flavor	54.62	58.55
Heptanoic acid	Fatty and rancid taste	92.03	152.19
Cyclohexadiene	–	118.36	103.95
Octadecanoic acid	Vinegar flavor	0.96	0.35
Indole	Fecal odor	245.03	100.75
Isovaleraldehyde	Meat spoilage odor	68.45	6.95
3-Methylbutanoic acid	Sour and foul-smelling foot sweat odor	12.43	34.91
2-Ethyl hexanol	2-Ethyl hexanol	73.59	20.03
Phenol	–	20.48	18.30
4-Methylindole	Fecal odor	126.34	123.19

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The description of the five most abundant volatile components before and after feeding is as follows:

Before feeding (brown line): indole, with a retention time of around 16–17 min and an absolute peak area of 245.03, providing a fecal odor; cyclohexadiene, with a retention time of around 14–15 min and an absolute peak area of 118.36, which had no odor characteristics; heptanoic acid, with a retention time of around 12–13 min and an absolute peak area of 92.03, known for its fatty and rancid taste; 2-ethyl hexanol, with a retention time of around 21–22 min and an absolute peak area of 73.59, which contributed to an alcoholic odor; and 4-methylindole, with a retention time of around 18–19 min and an absolute peak area of 126.34, which also had a fecal odor.

After feeding (green line): heptanoic acid, with a retention time of around 12–13 min and an absolute peak area of 152.19, known for its fatty taste; cyclohexadiene, with a retention time of around 14–15 min and an absolute peak area of 103.95, with no odor characteristics; indole, with a retention time of around 16–17 min and an absolute peak area of 100.75, providing a fecal odor; 4-methylindole, with a retention time of around 18–19 min and an absolute peak area of 123.19, also contributing a fecal odor; and valeric acid, with a retention time of around 13–15 min and an absolute peak area of 58.55, known for its sour vinegar flavor.

The graphs show that the peak values and retention times of different volatile components varied significantly before and after feeding. For instance, indole had the highest peak value before feeding, whereas heptanoic acid had the highest peak value after feeding. These changes indicated that feeding considerably impacted the concentration and distribution of these volatile components.

4. Discussion

The chewiness of fecal low-odor adult dog feed was lower than that of commercially available dog feed. Chewability refers to the effort required to chew feed in the mouth until it can be swallowed [18]. The lower the chewiness, the less effort the animal needs to make during chewing, making the feed more convenient to consume [19].

The elasticity of fecal low-odor adult dog feed is higher than that of commercially available dog feed. The lower the elasticity of dog feed, the easier it is to break. Higher elasticity ensures that the dog feed remains intact during handling and chewing. This durability can help maintain the nutritional content and structure of the feed [20].

Based on the variations in volatile components, it can be argued that fecal low-odor adult dog feed has advantages over commercially available dog feed. It boasts a wider array of aromatic compounds, including barbecue, malt, almond, resin, and floral fragrances, enhancing the attractiveness and palatability of the feed. In contrast, commercially available dog feed contains more acidic, alkane, and sweet flavors, potentially limiting its appeal to both pets and owners [5]. These differences suggest that fecal low-odor adult dog feed may be advantageous in terms of its palatability and attractiveness. Considering the volatile components, fecal low-odor adult dog feed appears to be more advantageous.

In our study, we identified several key volatile compounds in fecal low-odor adult dog feed that potentially enhance its attractiveness, including 3-methylbutanal with its appealing barbecue and malt flavor. These compounds likely enhance the feed’s overall aroma and attractiveness to dogs. However, further research with controlled trials is needed to confirm their roles in influencing feed preferences. These benefits were not directly measured in this study [21].The number of white blood cells and lymphocytes in the blood routine examination served as indicators to determine whether the experimental dogs were infected by bacteria. A significant decrease in the numbers indicated that the body was infected by bacteria [22].

The number of red blood cells and hematocrit were used to determine whether the experimental dogs had anemia. Hemoglobin is mainly used to diagnose degenerative hemoglobinemia and hematological diseases and can better reflect the degree of anemia [23].

Despite the variations observed in various indicators during this experiment, which remained within the normal fluctuation range, the results indicated that feeding fecal low-odor adult dog feed did not cause anemia, bacterial infection, or any other complications in the experimental dogs, nor did it affect their physical performance [24].

γ-Glutathione aminotransferase, alkaline phosphatase, and alanine aminotransferase are essential indicators for evaluating the liver function of experimental dogs [25]. The levels of various enzymes in the blood of experimental dogs fluctuated within the normal range before and after feeding fecal low-odor adult dog feed, indicating that fecal low-odor adult dog feed had no physiological impact on the experimental dogs. The levels of calcium and phosphorus in serum serve as indicators for judging the health of bones and teeth in the body [26]. Normal levels of calcium and phosphorus before and after this experiment indicated that the parathyroid function of the experimental dogs was as expected, the bone tissue was healthy, the vitamins in the dog feed were suitable, and the calcium-to-phosphorus ratio was appropriate. The level of creatinine indicated whether the kidney of the experimental dogs was diseased [27]. The average creatinine level before and after this experiment indicated that the experimental dogs had no renal diseases.

The total protein comprises globulin and albumin, which are related to the immune functions in the body. It can also maintain normal colloid osmotic pressure and regulate body substance metabolism, coagulation, and anticoagulation [28]. Although the total protein, albumin, and globulin contents fluctuated before and after this experiment, they were within the normal range, indicating that the immune and coagulation functions of experimental dogs were normal. The low-odor fecal adult dog feed did not affect the experimental dogs. The liquid nitrogen content in dog feces decreased from 1.60 to 1.26 mg/g.

The protein raw materials in dog feed underwent co-treatment with papain, acidic protease, and lactic acid bacteria. This process converted the protein molecules into more easily digestible small-molecule proteins and peptides, leading to a decrease in the liquid nitrogen content in feces. Additionally, the lactic acid bacteria introduced through the synergistic fermentation of bacteria and enzymes could help other beneficial bacteria in the intestine in using nitrogen to produce amino acids and proteins, thus reducing the liquid nitrogen content in feces [29].

The volatile odor content in the feces of the experimental dogs before and after feeding low-odor adult dog feed showed a slight decrease in the contents of acetic acid, octadecanoic acid, indole, isovaleraldehyde, 2-ethylhexanol, phenol, and 4-methylindole. Among these, the indole content, which had the most significant impact on the fecal spoilage odor, decreased significantly [30]. For humans, the reduction in unpleasant odors indicates potential improvements in the living environment. However, this implication requires further validation through additional studies focused on human perception and experience.

5. Conclusions

This study demonstrated no significant differences in nutritional characteristics and quality between fecal low-odor adult dog feed and commercially available dog feed. Feeding experiments showed that the fecal low-odor adult dog feed did not cause significant changes in the physiological and biochemical indicators of dogs. However, it effectively reduced the content of volatile nitrogen compounds and putrid odor in the feces. It remains unclear whether these effects are primarily due to the 40% inclusion rate of a non-meat compound or the specific properties of the processed plant protein. Nonetheless, this innovation provides a viable solution for pet owners aiming to minimize unpleasant odors and improve their pets’ well-being.