New use of bolo-rou extract and pet food additive

Abstract

The application discloses a new use of a fumaria officinalis extract and a pet food additive. The new use of the fumaria officinalis extract is used as a pet food additive for improving the first mouth preference of pet food and reducing the peculiar smell of pet feces, and the pet includes dogs and cats. The addition amount of the fumaria officinalis extract in the feed is 300 g / t; and the addition amount of glucose oxidase in the feed is 300 g / t. The addition in the daily ration can improve the abundance of probiotics in the intestinal tract of pets and reduce the abundance of pathogenic bacteria. The application can improve the first mouth preference of pets, dogs and cats, and the two have a certain synergistic effect, can effectively reduce the generation of fecal odor substances in the feces of the pets, dogs and cats, effectively improve the intestinal flora structure of the pets, dogs and cats, improve the intestinal health level of the pets, dogs and cats, improve the intestinal health condition of the pets, dogs and cats, and the product has the advantages of stable quality, high safety, simple preparation method and suitability for industrialized production.

Description

Technical Field

[0001] This invention relates to the field of feed additive technology, specifically to new uses of Boletus edulis extract and its application as a pet food additive. Background Technology

[0002] In modern life, many pet owners focus excessively on their pets' appearance, behavior, and apparent health indicators, neglecting their gut health. Some pet owners choose to feed their pets human food, processed food, and foods unsuitable for pet digestion. The additives, preservatives, and flavorings in these foods can negatively impact a pet's gut, leading to indigestion, gastroenteritis, allergies, and other illnesses. A pet's gut health is crucial to their overall well-being. A pet's gut contains a large number of beneficial bacteria that help them digest food, absorb nutrients, and maintain a balanced gut microbiota.

[0003] Improving a pet's gut health can reduce their risk of gastrointestinal diseases and boost their immunity, further reducing their chances of getting sick. In addition, improved gut health can also improve a pet's appetite and weight control, increase their energy levels, and enhance their overall well-being.

[0004] Because dog and cat food is high in protein and fat, and has a low absorption rate, pet excrement often has a strong odor. Pet excrement contains gases such as ammonia, hydrogen sulfide, methane, skatole, and amines, which can affect the indoor environment and people's affection for their pets. In addition, improper management by some pet owners, such as allowing their pets to overeat or ingesting spoiled food, can impair the normal function of their digestive system, sometimes leading to vomiting, diarrhea, constipation, bad breath, and foul-smelling feces. Many products on the market address these issues, but most contain antibiotics and high doses of zinc oxide to treat and prevent pet diarrhea. However, long-term use can lead to antibiotic resistance and secondary diarrhea, exacerbating the fecal odor. Furthermore, probiotics, prebiotics, and complex enzyme preparations have been reported to be used in pet food. These ingredients often have the effects of regulating intestinal flora, maintaining intestinal health, enhancing disease resistance, and reducing fecal odor. For example, Chinese patent applications CN111838444A, CN112568332A, CN112568333A, CN109527208A, and CN116515687A all disclose the relevant applications of probiotics, prebiotics, and compound enzyme preparations as suitable pet food additives.

[0005] However, existing reports do not mention any technology that uses extracts of *Gynostemma pentaphyllum* to reduce the odor of pet feces. Summary of the Invention

[0006] In order to overcome the shortcomings of the prior art, the purpose of this invention is to provide new uses for extracts of *Botrytis cinerea* and as a pet food additive.

[0007] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0008] A novel use of *Lysimachia christinae* extract as a pet food additive to enhance first-bite preference in pet food while reducing fecal odor in pets, including dogs and cats.

[0009] The amount of the extract of *Botrytis cinerea* added to the feed is 300 grams per ton.

[0010] The new use also includes glucose oxidase, which is added to feed at a rate of 300 grams per ton.

[0011] The new use involves feeding twice a day, replacing the old food with an arithmetic progression of 20% per day.

[0012] A novel use of *Gnaphalium affine* extract as a pet food additive, wherein adding 300 g / ton each of *Gnaphalium affine* extract and glucose oxidase to the diet increases the abundance of probiotics and reduces the abundance of pathogenic bacteria in the pet's gut, including dogs and cats.

[0013] The probiotics include Lactococcus lactis, Akkermansia muciniphila, Subdoligranulum and / or Cetobacterium; the pathogens include Klebsiella, Desulfovibrio and / or Escherichia-Shigella.

[0014] A feed additive that enhances the first bite preference of pet food while reducing the odor of pet feces, wherein the feed additive is a berberine extract, and the pets include dogs and cats.

[0015] The feed additives also include glucose oxidase.

[0016] The amount of the *Botrytis cinerea* extract added to the feed is 300 g / ton; the amount of glucose oxidase added to the feed is 300 g / ton.

[0017] The beneficial effects of this invention are:

[0018] 1. The pet food additives of this invention increase the first taste preference of dogs and cats, which directly affects the subsequent effects of the pet food additives of this invention in the pet's body, especially in dogs and cats.

[0019] 2. The pet food additive of this invention can increase the pet's food intake, while regulating the intestinal flora of dogs and cats, enhancing antioxidant and immune capabilities, preventing intestinal diseases, and improving the pet's intestinal health.

[0020] 3. The pet food additive of this invention can increase the levels of nutrients in the pet's intestines. Lactococcuslactis , Akkermansiamuciniphila , Subdoligranulum , Cetobacterium Increase the abundance of probiotics and reduce Klebsiella , Desulfovibrio , Escherichia-Shigella The abundance of pathogenic bacteria.

[0021] 4. In summary, the pet food additive of the present invention, starting from the first step of feeding, improves the first bite preference, increases feed intake, effectively improves the composition of intestinal microbiota, maintains intestinal flora balance, and enhances intestinal health and immunity in a synergistic way, ultimately effectively reducing fecal odor substances in pet feces. Attached Figure Description

[0022] Figure 1 The effect of *Pterocarya stenoptera* extract plus glucose oxidase on average daily feed intake in dogs.

[0023] Figure 2 This indicates the dog's initial preference for a diet containing Boletus extract and glucose oxidase.

[0024] Figure 3 The effect of *Pterocarya stenoptera* extract plus glucose oxidase on average daily food intake in cats.

[0025] Figure 4 This indicates the cat's initial preference for a diet containing Boletus extract and glucose oxidase.

[0026] Figure 5 The effect of glucose oxidase on the average daily food intake of dogs.

[0027] Figure 6 This indicates the dog's first preference for a diet containing glucose oxidase.

[0028] Figure 7 The effect of glucose oxidase on the average daily food intake of cats.

[0029] Figure 8 This indicates the cat's first preference for a glucose oxidase-rich diet.

[0030] Figure 9 To investigate the effect of *Pleurotus ostreatus* extract on the average daily feed intake of dogs.

[0031] Figure 10 The dog's first preference for a diet containing Boletus extract.

[0032] Figure 11 To investigate the effect of *Pleurotus ostreatus* extract on the average daily food intake of cats.

[0033] Figure 12 The cat's first preference for a diet containing Boletus extract. Detailed Implementation

[0034] The present invention will be further described below with reference to the embodiments and accompanying drawings.

[0035] Glucose oxidase is an enzyme that breaks down glucose into pyruvate and lactate, primarily found in the gut microbiota of animals. Its mechanism of action involves breaking down glucose into lactate and pyruvate, generating ATP energy to power intestinal cells. Furthermore, glucose oxidase promotes the growth of beneficial bacteria in the gut, maintains intestinal microecological balance, and enhances intestinal immunity, thus having a significant impact on animal gut health.

[0036] Boletus extract is a plant-based alkaloid, primarily derived from the plant *Boletus* genus, and is a natural product of plant origin. The mechanism of action of Boletus extract mainly involves inhibiting the growth and reproduction of harmful bacteria in the intestines while promoting the growth and reproduction of beneficial bacteria, thereby improving the balance of intestinal flora and maintaining intestinal health. An examination of existing reports has not found that Boletus extract or its extract components can directly reduce fecal odor or related substances. They are commonly used to prevent gastrointestinal inflammation in animals, inhibit the growth and reproduction of harmful pathogenic bacteria, and improve intestinal health. Regarding the application of Boletus extract, the Boletus extract injection listed in veterinary drug quality standards is a sterile aqueous solution of alkaloid sulfate prepared by extraction and separation of Boletus extract fruit. It has antibacterial and anti-inflammatory functions and is mainly used to treat white and yellow scours in piglets. Traditional Chinese veterinary medicine Boletus extract and its preparations are benzo[a]phenanthridine alkaloids prepared by extraction and separation of Boletus extract pods and / or leaves, possessing antibacterial, anti-inflammatory, and growth-promoting effects (CN101530475A).

[0037] The applicant (inventor) has accidentally discovered that adding one or both of glucose oxidase and *Polygonum hydropiper* extract in appropriate amounts to a pet's daily diet and feeding it regularly can effectively reduce the odor in pet feces. The following examples are intended to illustrate the invention and not to further limit it.

[0038] The extract of *Hemiberlesia lingua* of this invention can be prepared using the method described in invention patent CN101849994B of Hunan Mekeda Bioresources Co., Ltd., which extracts benzophenanthridine alkaloids from the natural plant *Hemiberlesia lingua*, mainly sanguisorbin and celandine, into a powder. Alternatively, those skilled in the art can use other existing methods for extraction and preparation, or directly purchase commercially available finished products.

[0039] Example 1: Palatability test of glucose oxidase and *Polygonum hydropiper* extract in dogs and cats.

[0040] Pet food supplemented with glucose oxidase and *Polygonum hydropiper* extract was prepared and added to the finished feed at a rate of 300 grams per ton. Twenty-four British Shorthair Blue cats and twenty-four Poodles without soft stools or diarrhea were selected for the experiment and divided into four groups of six cats / dogs each.

[0041] Specific groupings: Control group, fed a basal diet (Group J); Experimental group 1, fed a diet supplemented with 600g / ton of *Bletilla striata* extract and glucose oxidase (Group BP, 300g / ton each of *Bletilla striata* extract and glucose oxidase); Experimental group 2, fed a diet supplemented with 300g / ton of glucose oxidase (Group P); Experimental group 3, fed a diet supplemented with 300g / ton of *Bletilla striata* extract (Group B). The dietary supplementation was 100g / cat / day and 100g / dog / meal, with dogs fed twice daily, morning and evening. A 3-day observation period was conducted to monitor for any abnormalities in the cats and dogs. Adjustments were made based on the observation results, with a 4-day transition period using a 20% arithmetic gradient. The formal trial period was 5 days, for a total of 12 days.

[0042] Specific arrangements:

[0043] Day 1 of the observation period: Observe the cat's feces and eating habits, and weigh 100 g of old food (100 g / meal for dogs).

[0044] Day 2 of the observation period: Weigh the remaining amount, and proceed as above.

[0045] Day 3 of the observation period: Same as above

[0046] Day 1 of food transition: Weigh out 80 g of old food and 10 g each of food from the control group and the sample group (80 g: 20 g per meal for dogs).

[0047] Day 2 of the food transition period: Weigh out 60 g of old food and 20 g each of food from the control group and the sample group (60 g: 40 g per meal for dogs).

[0048] Day 3 of the food transition period: Weigh out 40 g of old food, and 30 g each of food from the control group and the sample group (40 g: 60 g per meal for dogs).

[0049] Day 4 of the food transition period: Weigh out 20 g of old food and 40 g each of food from the control group and the sample group (20:80 per meal for dogs).

[0050] Days 1-5 of the experiment: The palatability of pet food was assessed using the double-bowl method, comparing the control group and the sample group. In the same treatment group, two types of food, A and B, were distributed daily, 100 g each in two separate bowls (100 g / meal for dogs). The bowls were rotated daily, maintaining a distance of approximately 10 cm between them, and presented to the cat / dog simultaneously. The first type of food to be encountered and consumed was observed and recorded, along with the daily intake of each type of food. After recording the first bite, the observers quickly left to eliminate human interference. The frequency of first-bite preference and the consumption rate of each food in each group were calculated. The consumption rate was calculated using the formula: Consumption rate of food A = A food intake / (A food intake + B food intake) × 100%.

[0051] Figure 1 and Figure 2 The results showed that *Botrytis cinerea* extract combined with glucose oxidase significantly improved feed intake and first-bite preference in dogs, while its effect on cats was... Figure 3 and Figure 4 The study showed that average daily feed intake decreased slightly, while first-bite preference increased significantly. Figure 5 and Figure 6 The results showed that the addition of glucose oxidase significantly increased the average daily feed intake and first-served food preference in dogs. Figure 7 and Figure 8 The data showed that the average daily food intake of cats decreased, but their preference for the first bite increased significantly. Figure 9 and Figure 10 The results showed that *Botrytis cinerea* extract significantly increased average daily food intake and first-paste preference in dogs, while the results in cats were similar. Figure 11 and Figure 12 The study showed that extracts of *Botrytis cinerea* significantly increased average daily feed intake and slightly improved the degree of improvement in first-bite preference.

[0052] In conclusion, the addition of substances such as *Clerodendrum trichotomum* extract and glucose oxidase to the feed can increase the first-meal preference of dogs and cats. Specifically, the feed intake of dogs significantly increased after the addition of *Clerodendrum trichotomum* extract and glucose oxidase, or both.

[0053] Example 2: Effects of glucose oxidase and *Euphorbia lathyris* extract on the feeding of pet dogs and cats.

[0054] Twenty-four British Shorthair Blue cats and twenty-four Poodles without soft stools or diarrhea were selected for the experiment and divided into four groups of six cats / dogs each.

[0055] Specific groupings: Control group, fed a basal diet (Group J); Experimental group 1, fed a diet supplemented with *Gynostemma pentaphyllum* extract and glucose oxidase at 600 g / ton (Group BP); Experimental group 2, fed a diet supplemented with glucose oxidase at 300 g / ton (Group P); Experimental group 3, fed a diet supplemented with *Gynostemma pentaphyllum* extract at 300 g / ton (Group B). The diet needed to meet the nutritional requirements of AAFCO (2017). The diet transition period was 20% arithmetic change, lasting 4 days, with a formal experimental period of 14 days, for a total of 18 days. Throughout the experimental period, food intake was recorded daily; fecal scores (FS) were performed daily; fasting weight and body condition scores (BCS) were performed on the first day of diet transition and the last day of the experiment, along with fecal scores, blood samples, and fresh feces were collected.

[0056] Necessary immunizations and deworming treatments were administered prior to the experiment, and the animals were not given any medications (such as antibiotics) that could alter their gut microbiota within one month prior to the start of the experiment. Daily cleaning and disinfection were carried out to maintain the cleanliness of the pet enclosures. Each animal was fed a fixed amount of food at 8:30 AM and 5:00 PM daily, and the amount fed, the amount remaining the following day, and the stool pattern were accurately recorded each day. Clean water was provided free of charge to the animals throughout the experiment.

[0057] As shown in Tables 1 and 2, the feed intake of the Bo Luo Hui extract + glucose oxidase group, the glucose oxidase group, and the Bo Luo Hui extract group was higher than that of the basal diet group. There were no significant differences in fecal scores and body weight changes, and the feces of the experimental animals in each group were in a normal state.

[0058] Table 1. Dog's food intake, fecal score, and weight

[0059] project Group J BP group Group P Group B Feed intake (g) 80.88 92.95 86.23 94.49 Stool score 2.58 2.53 2.51 2.51 Weight change (kg) 0.09 0.025 0.11 0.24

[0060] Note: Stool score (FS): 1 ≤ FS < 2 indicates constipation, 2 ≤ FS ≤ 3 indicates normal stool, 3 < FS < 4 indicates soft stool, and 4 ≤ FS ≤ 5 indicates diarrhea.

[0061] Table 2. Cat food intake, fecal score, and weight

[0062] project Group J BP group Group P Group B Feed intake (g) 50.03 58.22 63.54 57.30 Stool score 2.47 2.49 2.50 2.50 Weight change (kg) -0.07 -0.17 0.01 0.03

[0063] Note: Stool score (FS): 1 ≤ FS < 2 indicates constipation, 2 ≤ FS ≤ 3 indicates normal stool, 3 < FS < 4 indicates soft stool, and 4 ≤ FS ≤ 5 indicates diarrhea.

[0064] As shown in Table 3, compared with the basal diet group, group B significantly reduced the levels of indole, cadaverine, and skatole in dogs' feces; compared with the basal diet group, group BP significantly reduced the levels of indole and skatole in dogs' feces, and group P significantly reduced the levels of indole and skatole in dogs' feces.

[0065] As shown in Table 4, compared with the basal diet group, the BP group and the B group reduced the indole content in cat feces; compared with the basal diet group, the BP group, the B group and the P group significantly reduced the putrescine content in cat feces; compared with the basal diet group, the BP group significantly reduced the spermidine content in feces, and the BP group and the B group significantly reduced the skatole content in cat feces.

[0066] Table 3. Effects of *Gynostemma pentaphyllum* extract and glucose oxidase on canine odor compounds.

[0067] project Group J BP group Group P Group B Indole 576.95a 331.20b 404.23b 425.10b Cadaniamine 4742.30a 4116.92ab 2622.97ab 2070.11b Spermine 5.18b 5.98ab 8.67a 5.13b skatole 250.84a 199.99b 48.60b 137.84b

[0068] Note: All indicators are in ng / g; different subtitles in the same data indicate significant differences (P<0.05), while the same subtitle indicates no significant differences (P>0.05).

[0069] Table 4. Effects of *Cephalotaxus fortunei* extract and glucose oxidase on cat odor compounds.

[0070] project Group J BP group Group P Group B Indole 26618.09a 24314.09ab 15190.67b 16193.32ab putrescine 123377.11a 41531.75b 11666.29b 47308.18b spermidine 353.32b 309.45ab 169.66b 203.20b skatole 257.73a 165.98b 193.88ab 165.29b

[0071] Note: All indicators are in ng / g; different subtitles in the same data indicate significant differences (P<0.05), while the same subtitle indicates no significant differences (P>0.05).

[0072] As shown in Table 5, compared with group B, group BP significantly increased the serum albumin-globulin ratio in dogs, indicating that it has an effect on improving the immune level of the animal body; compared with the basal diet group, there were no significant differences in serum biochemical indicators among the other three groups.

[0073] As shown in Table 6, compared with groups P and B, group BP significantly increased the serum total protein content of cats, indicating that it has an effect on improving the immune level of the animal body; compared with the basal diet group, there were no significant differences in serum biochemical indicators among the other three groups.

[0074] Table 5. Effects of *Botrytis cinerea* extract and glucose oxidase on serum biochemical parameters in dogs.

[0075] project Group J BP group Group P Group B Albumin (g / L) 33.02 35.57 33.80 34.20 Total protein (g / L) 67.93 67.17 69.20 71.75 Globulin (g / L) 34.92 31.63 35.40 36.53 White ball ratio 1.01ab 1.14a 1.05ab 0.97b Aspartate aminotransferase (U / L) 35.40 37.67 40.83 37.50 Alanine aminotransferase (U / L) 27.67 28.83 36.67 37.17 Amylase (U / L) 613.67 621.17 646.17 656.83 Creatine kinase (U / L) 168.75 151.00 161.67 142.00 Creatinine (µmol / L) 60.82 64.42 67.30 69.86 Blood urea nitrogen (mmol / L) 4.64 4.90 5.42 4.90 Blood urea nitrogen / creatinine 80.54 76.40 81.17 101.74 Glucose (mmol / L) 4.71 4.60 4.83 4.36 Triglycerides (mmol / L) 0.68 0.77 0.75 0.76 Calcium (mmol / L) 2.34 2.36 2.38 2.35 Inorganic phosphorus (mmol / L) 1.15 1.01 0.96 0.98

[0076] Note: Different annotated labels in the same data indicate significant differences (P<0.05), while identical annotated labels indicate no significant differences (P>0.05).

[0077] Table 6. Effects of *Botrytis cinerea* extract and glucose oxidase on feline serum biochemical parameters.

[0078] project Group J BP group Group P Group B Albumin (g / L) 25.95 24.64 25.26 24.23 Total protein (g / L) 77.88ab 83.22a 76.35b 76.17b Globulin (g / L) 51.93 54.68 50.16 51.93 White ball ratio 0.50 0.52 0.51 0.51 Aspartate aminotransferase (U / L) 24.40 22.17 24.83 23.33 Alanine aminotransferase (U / L) 58.00 47.50 53.50 52.00 Amylase (U / L) 939.33 1047.67 951.83 1076.83 Creatine kinase (U / L) 148.25 132.60 175.33 158.00 Creatinine (µmol / L) 102.33 96.90 106.80 94.92 Blood urea nitrogen (mmol / L) 6.76 5.66 6.25 6.57 Blood urea nitrogen / creatinine 66.84 68.01 74.44 69.44 Glucose (mmol / L) 4.90 5.86 4.84 4.70 Triglycerides (mmol / L) 0.43 0.42 0.42 0.56 Calcium (mmol / L) 2.28 2.18 2.25 2.27 Inorganic phosphorus (mmol / L) 1.50 1.60 1.51 1.53

[0079] Note: Different annotated labels in the same data indicate significant differences (P<0.05), while identical annotated labels indicate no significant differences (P>0.05).

[0080] Table 7 shows the fecal microbial composition of dogs after the experimental treatment. In this experiment, 23 phyla were identified, with Firmicutes, Actinobacteriota, Proteobacteria, Fusobacteriota, and Bacteroidota being the dominant phyla, accounting for over 99%. Compared to the basal diet (J) group, the relative abundance of Proteobacteria was significantly decreased in the glucose oxidase (P) group, while the relative abundance of Campylobacterota was significantly increased in the group supplemented with 300 g / t Campylobacterota extract (B). The relative abundance of Firmicutes in the glucose oxidase (P) group was significantly higher than that in the Campylobacterota extract + glucose oxidase (BP) group, while the relative abundance of Proteobacteria was significantly lower than that in the Campylobacterota extract + glucose oxidase (BP) group. At the genus level, Holdemanella , Collinsella , Ruminococcus , Allobaculum , Erysipelatoclostridium , Streptococcus It is the dominant genus of bacteria.

[0081] Compared with the basal diet (J) group, the relative abundance of Akkermansia and Lactococcus was significantly increased in the group receiving Boluhui extract + glucose oxidase (BP).

[0082] Compared with the basal diet (J) group, the relative abundance of Akkermansia and Ligilactobacillus was significantly increased in the glucose oxidase (P) group, while the relative abundance of Romboutsia and Klebsiella was significantly decreased.

[0083] Compared with the basal diet (J) group, the relative abundance of Akkermansia and Dubosiella species was significantly increased in the Boluhui extract (B) group, while the relative abundance of Staphylococcus and Acinetobacter species was significantly decreased.

[0084] Table 7. Microbial differences in feces of different groups of Poodles at the phylum and genus levels.

[0085] project Group J BP group Group P Group B 3.15a 5.23a 1.55b 3.16ab 0a 0.02ab 0.00ab 0.01b 77.76ab 69.33b 84.10a 78.96ab 0.05c 0.10b 0.10b 0.15a 0.01b 0.02a 0.01b 0c 0.07b 0.11ab 0.15a 0.07b 0.89a 1.17ab 0.05b 5.46a 0.17a 0.13a 0.06b 0.09ab 0.13a 0.09ab 0.06b 0.00c 0.07a 0.07ab 0.02b 0.02b 0.00b 0.00b 0.00b 0.08a

[0086] Table 8 shows the community composition of cat feces microorganisms after experimental treatment. In this experiment, a total of 29 phyla were identified, with Firmicutes, Actinobacteriota, Proteobacteria, Bacteroidota, and Verrucomicrobiota being the dominant phyla, accounting for more than 99%.

[0087] Compared with the basal diet (J) group, the relative abundance of Bacteroidota was significantly reduced in the Bacteroidota-Bacteroidota group with Bacteroidota extract and glucose oxidase (BP); the relative abundance of Firmicutes was significantly increased in the glucose oxidase (P) group (P=0.02), while the relative abundance of Actinobacteriota was significantly reduced.

[0088] At the genus level, Megasphaera, Collinsella, Subdoligranulum, Catenibacterium, Bifidobacterium, Dialister, Prevotella_9, Megamonas, and Escherichia-Shigella are the dominant genera.

[0089] Compared with the basal diet (J) group, the relative abundance of Subdoligranulum in the Boluoke extract + glucose oxidase (BP) group was significantly increased;

[0090] Compared with the basal diet (J) group, the relative abundance of *Cetobacterium* was significantly increased and the relative abundance of *Desulfovibrio* was significantly decreased in the glucose oxidase (P) group;

[0091] Compared with the basal diet (J) group, the relative abundance of Escherichia-Shigella and Desulfovibrio, and the relative abundance of Klebsiella were significantly reduced in the Bo Luo Hui extract (B) group.

[0092] Table 8. Microbial differences in feces of British Shorthair Blue cats at the phylum and genus levels among different groups.

[0093] project Group J BP group Group P Group B 14.07a 0.55b 3.16ab 0.97ab 46.53b 61.05b 76.43a 63.01b 32.47a 31.18a 13.13b 30.46a 3.47b 12.67a 8.95ab 10.02ab 0.02b 0.16ab 0.34a 0.00c 0.39a 0.17a 0.05b 0.00c 2.13ab 2.25a 1.51b 0.01c 0.71a 0.64a 0.50a 0.12b

[0094] Existing research indicates that *Lactococcus lactis* possesses anti-inflammatory and immunomodulatory capabilities, enhancing growth performance; *Akkermansia muciniphila* acts as a probiotic in metabolic regulation, immune regulation, and gut health protection; many strains of *Ligilactobacillus* exhibit beneficial functional properties, such as antibacterial activity, immune effects, and the ability to regulate the gut microbiota; *Romboutsia* is a potential pathogen in ulcerative colitis, and the expression levels of pro-inflammatory central genes (such as CXCL1, CXCL9, and CCL7) are positively correlated with the abundance of *Romboutsia*; *Klebsiella* infection can induce clinical diseases in immunodeficient animals, especially those with severe immunodeficiency or congenital immune system deficiencies; *Acinetobacter* is an opportunistic pathogen that easily causes infection when the body's resistance is lowered, leading to respiratory infections, sepsis, meningitis, endocarditis, wound and skin infections, and genitourinary tract infections. Severe cases can lead to death, and are more common in canines. The abundance of Dubosiella and Romboutsia is negatively correlated with body weight, while Dubosiella is positively correlated with butyrate levels, which may help alleviate colitis. Subdoligranulum is one of the producers of butyrate, which is crucial for gut health. Cetobacterium is considered a major group of gut bacteria in many freshwater fish, and its members contribute to anaerobic metabolism, which helps maintain glucose homeostasis. Desulfovibrio is a type of anaerobic bacterium that reduces sulfate and produces H2S. Endogenous H2S can toxicize intestinal epithelial cells, causing intestinal sensitivity, leaky gut, or abdominal pain. Multiple clinical studies have confirmed that an increase in the number of Desulfovibrio is an important feature of polyps and ulcerative colitis. Studies have shown that Desulfovibrio is significantly higher in Parkinson's patients than in healthy controls, and reducing its relative abundance may help prevent the disease. Escherichia-Shigella is a factor associated with gut microbiota dysbiosis and is significantly positively correlated with the pro-inflammatory factor IL-1β, which can promote intestinal damage and affect amino acid metabolism.

[0095] After adding Lactococcus lactis extract or glucose oxidase, the abundance of probiotics such as Lactococcus lactis, Akkermansia muciniphila, Subdoligranulum, and Cetobacterium in the intestines of pet dogs and cats increased significantly, indicating that this additive has a significant effect on improving the animal's immunity, anti-infection, and intestinal health. Meanwhile, the abundance of pathogenic bacteria such as Klebsiella, Desulfovibrio, and Escherichia-Shigella in the intestines of pet dogs and cats decreased significantly after using this additive, indicating that the additives of this invention can effectively reduce the abundance of pathogenic bacteria, reduce the probability of infection and inflammation caused by these pathogenic bacteria in the intestines, maintain the intestinal microecological flora structure and microbial regulatory function, and improve the intestinal health of pets.

[0096] In summary, adding glucose oxidase or *Bletilla striata* extract to the diet can increase feed intake in dogs and cats, while effectively reducing odorous substances in their feces. It has no adverse effects on serum biochemical indicators. By regulating the intestinal flora, it enhances antioxidant and immune capabilities, preventing intestinal diseases. Addressing the issue that high protein in current pet foods leads to increased odorous substances in dog and cat feces, the additives used in this invention can effectively reduce this phenomenon. Simultaneously, it can effectively regulate the intestinal flora, increase its diversity, improve the immune regulation capacity of the pet's gut, enhance intestinal health, and prevent intestinal diseases.

[0097] The embodiments described above can be further combined or replaced, and these embodiments are merely descriptions of preferred embodiments of the present invention, not limitations on the concept and scope of the present invention. Various changes and improvements made to the technical solutions of the present invention by those skilled in the art without departing from the inventive concept are all within the protection scope of the present invention. The protection scope of the present invention is given by the appended claims and any equivalents.

Claims

1. The use of *Botrytis cinerea* extract in the preparation of pet food additives that improve the first bite preference of pet food while reducing the odor of pet feces, characterized in that, The pets mentioned are dogs and cats; The amount of the extract of *Botrytis cinerea* added to the feed is 300 grams per ton; The pet food additive also includes glucose oxidase, which is added to the feed at a rate of 300 grams per ton. Replace old food with an arithmetic gradient of 20% per day.

2. The use of *Polygonum hydropiper* extract in the preparation of pet food additives that improve the abundance of probiotics in the pet's gut, characterized in that... Adding 300 grams / ton each of *Bombyx mori* extract and glucose oxidase to the diet of dogs and cats can increase the abundance of probiotics in the gut. When the pet is a dog, the probiotics are Lactococcus lactis and Akkermansia muciniphila; When the pet is a cat, the probiotic is Subdoligranulum.

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