**4. Pet diets and animal well‐being**

Some scientific evidence has pointed out the efficacy of selected ingredients, as part of a commercially available diet, in relieving inflammatory conditions in pets by means of an immune modulatory and antioxidant activity [49–57].

Pasquini et al. [53] demonstrated that *dogs* fed a specific diet (F10 Maxi Maintenance®) based on maize, fish meal (20%), maize oil, fish oil, brewer's yeast, beet pulp, minerals, MOS, FOS, *Elaeis guineensis*, *Yucca schidigera*, *C. papaya*, *Ananas* spp., *P. granatum*, *Panax ginseng*, and *Rosmarinus officinalis* was able to influence gender, age, and breed‐derived lipid metabolism alterations in healthy *dogs* by significantly decreasing C‐tot, C‐high‐density lipoprotein (HDL), and C‐low‐density lipoprotein (LDL) (\* *p* < 0.05) [57].

Further studies have then clearly demonstrated the synergic efficacy of selected ingredients in modulating several inflammatory conditions, which commonly affect pets, especially *dogs* [49–52, 56, 58].

An inflammatory condition can also occur during food allergy reactions, which usually takes place after the intake of a harmless dietary component [59]. Generally, food‐allergic reactions in pets include cutaneous (flush, itching, dandruff, skin malodor, dry fur, and skin lesions) and gastrointestinal manifestations (dehydration, appetite loss, regurgitation, emesis, abdominal pain, flatulence, borborygma, diarrhea, weight loss, stool consistency, blood, and mucus presence in the stool) [56].

Based on these observations, we conducted two different clinical evaluations aimed to validate two different commercially available formulas for aforementioned dermatological and gastrointestinal issues. For instance, a mixture of fish, potato, *A. vera, Arctium lappa, Malva sylvestris,* and *Ribes nigrum* (FORZA10 Dermo ActiveTM) resulted particularly effective in halving the intensity of cutaneous symptoms (flush, itch, dandruff, skin malodor, dry fur, and skin lesions) in 71 *dogs* affected by atopic dermatitis (\*\*\**p* < 0.001) [56]. On the other hand, a specific diet consisting of a mixture of milk enzymes, *Origanum vulgare*, chestnut, *Plantago psyllium*, MOS, FOS, electrolytes, and *Rosa canina*, significantly reduced the intensity of symptoms (dehydration, appetite loss, regurgitation, emesis, abdominal pain, flatulence, borborygma, diarrhea, weight loss, stool consistency, blood, and mucus presence in the stool) in 60 *dogs* with evident gastrointestinal issues (\*\*\**p* < 0.001) [56]. Obviously, an inflammatory condition may also occur in other segments of the gastrointestinal apparatus, such as the mouth. In this case, the main clinical manifestation is halitosis, which is generally the sum of metabolic anomalies, poor oral hygiene, hyposalivation, dental appliances, as well as gingival, mucosal, and periodontal disease [60]. In this regard, pets can be affected by halitosis and a correct dietary approach becomes fundamental to manage and treat such complaints [50, 61]. We compared the efficacy of a specific nutraceutical diet and a standard diet in significantly reducing the concentration of three specific volatile sulfur compounds, hydrogen sulfide, methyl mercaptans, and dimethyl sulfide, in 16 *dogs* suffering from chronic halitosis [50]. More in detail, the diet consisted of a mixture of fish meal, rice carbohydrates, propolis, *Salvia officinalis*, lysozyme, bioflavonoids, *Thymus vulgaris*, *R. nigrum,* and an Omega 3/6 ratio of 1:4. Then, by means of a portable gas chromatograph (OralChroma™), a syringe to collect the breath and specific software, a significant reduction in halitosis, were observed after 30 days from the beginning of the nutraceutical diet supplementation (\* *p* < 0.05). Further, a long‐lasting effect was still observed even 20 days after the diet interruption.

**4. Pet diets and animal well‐being**

and C‐low‐density lipoprotein (LDL) (\*

[49–52, 56, 58].

presence in the stool) [56].

immune modulatory and antioxidant activity [49–57].

332 Superfood and Functional Food - An Overview of Their Processing and Utilization

Some scientific evidence has pointed out the efficacy of selected ingredients, as part of a commercially available diet, in relieving inflammatory conditions in pets by means of an

Pasquini et al. [53] demonstrated that *dogs* fed a specific diet (F10 Maxi Maintenance®) based on maize, fish meal (20%), maize oil, fish oil, brewer's yeast, beet pulp, minerals, MOS, FOS, *Elaeis guineensis*, *Yucca schidigera*, *C. papaya*, *Ananas* spp., *P. granatum*, *Panax ginseng*, and *Rosmarinus officinalis* was able to influence gender, age, and breed‐derived lipid metabolism alterations in healthy *dogs* by significantly decreasing C‐tot, C‐high‐density lipoprotein (HDL),

*p* < 0.05) [57].

Further studies have then clearly demonstrated the synergic efficacy of selected ingredients in modulating several inflammatory conditions, which commonly affect pets, especially *dogs*

An inflammatory condition can also occur during food allergy reactions, which usually takes place after the intake of a harmless dietary component [59]. Generally, food‐allergic reactions in pets include cutaneous (flush, itching, dandruff, skin malodor, dry fur, and skin lesions) and gastrointestinal manifestations (dehydration, appetite loss, regurgitation, emesis, abdominal pain, flatulence, borborygma, diarrhea, weight loss, stool consistency, blood, and mucus

Based on these observations, we conducted two different clinical evaluations aimed to validate two different commercially available formulas for aforementioned dermatological and gastrointestinal issues. For instance, a mixture of fish, potato, *A. vera, Arctium lappa, Malva sylvestris,* and *Ribes nigrum* (FORZA10 Dermo ActiveTM) resulted particularly effective in halving the intensity of cutaneous symptoms (flush, itch, dandruff, skin malodor, dry fur, and skin lesions) in 71 *dogs* affected by atopic dermatitis (\*\*\**p* < 0.001) [56]. On the other hand, a specific diet consisting of a mixture of milk enzymes, *Origanum vulgare*, chestnut, *Plantago psyllium*, MOS, FOS, electrolytes, and *Rosa canina*, significantly reduced the intensity of symptoms (dehydration, appetite loss, regurgitation, emesis, abdominal pain, flatulence, borborygma, diarrhea, weight loss, stool consistency, blood, and mucus presence in the stool) in 60 *dogs* with evident gastrointestinal issues (\*\*\**p* < 0.001) [56]. Obviously, an inflammatory condition may also occur in other segments of the gastrointestinal apparatus, such as the mouth. In this case, the main clinical manifestation is halitosis, which is generally the sum of metabolic anomalies, poor oral hygiene, hyposalivation, dental appliances, as well as gingival, mucosal, and periodontal disease [60]. In this regard, pets can be affected by halitosis and a correct dietary approach becomes fundamental to manage and treat such complaints [50, 61]. We compared the efficacy of a specific nutraceutical diet and a standard diet in significantly reducing the concentration of three specific volatile sulfur compounds, hydrogen sulfide, methyl mercaptans, and dimethyl sulfide, in 16 *dogs* suffering from chronic halitosis [50]. More in detail, the diet consisted of a mixture of fish meal, rice carbohydrates, propolis, *Salvia officinalis*, lysozyme, bioflavonoids, *Thymus vulgaris*, *R. nigrum,* and an Omega 3/6 ratio of 1:4. As previously stated, *cats* and *dogs* can be affected by adverse food reactions, which can involve apparatuses including the gastroenteric and cutaneous but can be extended also to otological, ocular, urinary, and respiratory systems [49, 62]. We recently ascertained the effectiveness of a combined use of a nutraceutical diet and current pharmacological therapy in 15 adult *dogs* affected by chronic bilateral otitis externa [49]. The diet, which was composed of fish proteins, rice carbohydrates, *Melaleuca alternifolia, Tilia platyphyllos scapoli et cordata, Allium sativum L., Rosa canina L.* and Zinc and an Omega 3/6 ratio of 1:4, and the drug (Otomax®) significantly reduced the mean intensity of all clinical symptoms (occlusion of ear canal, erythema, discharge quantity, and odor) within 90 days (\*\*\**p* < 0.001). This study can be considered a further example of the importance of the selection of substances endowed with anti‐inflam‐ matory and antioxidant activity in a pet food diet.

In some cases, substances endowed with anti‐inflammatory as well as immune‐modulatory activity can drastically influence the clinical outcome of lethal pathologies, that is, *Leishmania* [63].

A study conducted by Cortese et al. investigated the effect of an immune‐modulating diet, based on fish‐ and vegetable‐hydrolyzed proteins, minerals, *A. nodosum, C. melo, C. papaya, A. vera, Astaxanthin, C. longa, C. sinensis, P. granatum, P. nigrum, Poligonum* spp*., E. purpurea, G. frondosa, G. max* and an Omega 3/6 ratio of 1:1 along with an anti‐*Leishmania* pharmacological therapy (meglumine antimoniate, and allopurinol) in 20 naturally infected *dogs* over a period of 12 months [63]. The diet results were particularly effective in restoring regulatory T cells and decreasing T helper cell percentage (\*\*\**p* < 0.001).

In other cases, the selection of substances with a remarkable antioxidant activity also acquires a pivotal role in other clinical conditions, which are not strictly related to adverse food reactions, that is, cognitive impairment, as a consequence of aging or pathologies such as Alzheimer's and Parkinson's disease [52, 64, 65].

In this regard, we studied the effect of a nutraceutical diet based on fish proteins, rice carbo‐ hydrates, *G. frondosa, C. longa, C. papaya, P. granatum, A. vera, P. cuspidatum, Solanum lycopersi‐ cum, Vitis vinifera, R. officinalis,* and an Omega 3/6 ratio of 1:0.8 on cognitive decline of nine elderly *dogs* over a period of 6 months [52]. Specifically, derivatives of reactive oxygen metabolites, biological antioxidant potential levels, and brain‐derived neurotrophic factor were evaluated in *dogs*' plasma samples at the beginning and at the end of the dietary regime. Results showed a significant decrease of dROMs (\* *p* < 0.05) and a significant increase in brain‐ derived neurotropic factor (BDNF) (\* *p* < 0.05) serum levels.

A recent study has also raised the possible key role of some selected ingredients (fish proteins, rice carbohydrates, *P. granatum, Valeriana officinalis, R. officinalis, Tilia* spp*.,* tea extract, and l‐ tryptophan, with an Omega‐3:‐6 ratio of 1:0.8) in modulating behavioral disturbances in 12 *dogs* with chronic anxiety and stress caused by intense and restless activity over a period of only 10 days [58]. By means of a sophisticated and extremely sensitive sensor, a mobile phone app, and a wireless router, it was possible to induce and monitor significant improvements in the time spent in activity and at rest (\*\**p* < 0.01 and \* *p* < 0.05, respectively). Last but not least, all *dogs* also showed an overall significant improvement in clinical (dandruff, itchiness, flush, seborrhea, fur opacity, vomiting, diarrhea, flatulence, lachrymation, and anal sac repletion) and behavioral (marking, anxiety, diffidence, irregular biorhythm, reactivity, activation, irritability, alertness, environmental exploration, and attention requirement) symptoms.

#### **5. The "market stand" of functional pet foods**

The interest into the adequacy and safety of commercially available pet foods has been growing worldwide [66]. Functional foods such as prebiotics, for example, inulin, gluco‐oligosacchar‐ ides, and galacto‐oligosaccharides have shown to induce beneficial effects on biochemical parameters improving satiety and reducing postprandial glucose and insulin concentrations, thus reducing diabetes‐related disorders [67–69]. Inulin and oligofructose, but also dietary fibers, can also modify the intestinal microflora in pets and humans by promoting commensal bacteria growth [70–72]. However, many *in vitro* studies highlighted other hidden properties of dietary fibers such as gastric emptying, gastric transit time and decrease in blood cholesterol concentrations, increase in satiety, glucose uptake rate, and fecal excretion as well as dilution in diet calorie density [73–76]. Another valuable fiber source is represented by corn fiber due to the lack of detrimental effects on palatability and nutrient digestibility, and the glycemic response lowering in adult *dogs* [71, 77]. Based on these novel and unexpected activities of dietary fibers, many commercially available pet diets moved to an accurate use of these along with novel sources of carbohydrates including cereal grains, which represents almost 90% of animal diet content, and whole grains [78]. These latter, whose main source are wheat, corn, oats, barley, and rye [79], are rich in dietary fibers, trace minerals, and vitamins B and E [80]. Furthermore, whole grains have bioactive compounds, for example, tocotrienols, lignans and polyphenols, lipotropes and methyl donors, such as choline, methionine, betaine, inositol and folate and antinutrients, that is, compounds that interfere with the absorption of nutrients such as phytic acid, tannins, and saponins endowed with antioxidant and anti‐carcinogenic effect [79–82]. As to rice bran, the vitamin‐rich outer layer that surrounds the endosperm of whole grain brown rice has bioactive molecules such as tocopherols, tocotrienols, polyphenols including ferulic acid and α‐lipoic acid, phytoesterols, γ‐oryzanol and carotenoids such as carotene, lycopene, lutein, and zeaxanthin, which was endowed with antioxidant, anti‐ inflammatory, and chemopreventive activity [83]. Rice bran is also an excellent source of essential amino acids (especially sulfur‐containing amino acids) and micronutrients such as magnesium, manganese, and B‐vitamins (especially B9 and B12) [83, 84]. It is worth noting that during pet food heat processing, known as the Maillard reaction, that is, a nonenzymatic browning and flavoring reaction, a reduction of essential amino acids, such as lysine, bioa‐ vailability occurs [85, 86]. Therefore, many pet diets might be at the risk of supplying less lysine than the animal may require. Hence, the understanding of nutritional benefits of functional foods currently available is of key importance for the owners to provide their pets with the correct diet. Nevertheless, great attention has to be paid to pet food palatability along with adequacy and safety. For instance, Spears and coworkers examined the palatability and its effect on digestion of stabilized rice bran in a dry canine diet determining fecal characteristics, food intake, selected immune mediators, and blood lipid characteristics [87]. They observed that dry pet food containing 12% stabilized rice bran was well tolerated by *dogs* with no detrimental effect on nutrient digestibility, fecal characteristics, and changes in inflammatory/ immune mediators. Moreover, the rice bran diet presented greater palatability compared to the defatted rice bran diet. Vitamin A (retinol), whose safe upper limit in complete diets for *dogs* ranges from 5.24 to 104.80 mmol, is an essential fat‐soluble vitamin at the center of investigations in *dogs* in the context of immune stimulation, vision‐supporting functions, reproduction, bone growth, and cellular differentiation [88–90]. In *dogs*, unlike humans where retinyl esters are only detected in plasma in cases of intoxication or following a vitamin A‐rich meal [91], vitamin A is present in the plasma predominantly in the form of retinyl esters, in both adequate and vitamin A‐deprived states [92]. Moreover, in *dogs* [93], which excrete vitamin A in the urine [91] along with retinyl esters [94], retinol concentrations are unaffected by dietary vitamin A intake (1.2 ± 0.03 vs. 1.0 ± 0.03 mg/l, respectively), whereas serum retinyl esters parallel the concentrations of vitamin A in the diet [91].

#### **5.1. The role of microbiota**

only 10 days [58]. By means of a sophisticated and extremely sensitive sensor, a mobile phone app, and a wireless router, it was possible to induce and monitor significant improvements in

all *dogs* also showed an overall significant improvement in clinical (dandruff, itchiness, flush, seborrhea, fur opacity, vomiting, diarrhea, flatulence, lachrymation, and anal sac repletion) and behavioral (marking, anxiety, diffidence, irregular biorhythm, reactivity, activation, irritability, alertness, environmental exploration, and attention requirement) symptoms.

The interest into the adequacy and safety of commercially available pet foods has been growing worldwide [66]. Functional foods such as prebiotics, for example, inulin, gluco‐oligosacchar‐ ides, and galacto‐oligosaccharides have shown to induce beneficial effects on biochemical parameters improving satiety and reducing postprandial glucose and insulin concentrations, thus reducing diabetes‐related disorders [67–69]. Inulin and oligofructose, but also dietary fibers, can also modify the intestinal microflora in pets and humans by promoting commensal bacteria growth [70–72]. However, many *in vitro* studies highlighted other hidden properties of dietary fibers such as gastric emptying, gastric transit time and decrease in blood cholesterol concentrations, increase in satiety, glucose uptake rate, and fecal excretion as well as dilution in diet calorie density [73–76]. Another valuable fiber source is represented by corn fiber due to the lack of detrimental effects on palatability and nutrient digestibility, and the glycemic response lowering in adult *dogs* [71, 77]. Based on these novel and unexpected activities of dietary fibers, many commercially available pet diets moved to an accurate use of these along with novel sources of carbohydrates including cereal grains, which represents almost 90% of animal diet content, and whole grains [78]. These latter, whose main source are wheat, corn, oats, barley, and rye [79], are rich in dietary fibers, trace minerals, and vitamins B and E [80]. Furthermore, whole grains have bioactive compounds, for example, tocotrienols, lignans and polyphenols, lipotropes and methyl donors, such as choline, methionine, betaine, inositol and folate and antinutrients, that is, compounds that interfere with the absorption of nutrients such as phytic acid, tannins, and saponins endowed with antioxidant and anti‐carcinogenic effect [79–82]. As to rice bran, the vitamin‐rich outer layer that surrounds the endosperm of whole grain brown rice has bioactive molecules such as tocopherols, tocotrienols, polyphenols including ferulic acid and α‐lipoic acid, phytoesterols, γ‐oryzanol and carotenoids such as carotene, lycopene, lutein, and zeaxanthin, which was endowed with antioxidant, anti‐ inflammatory, and chemopreventive activity [83]. Rice bran is also an excellent source of essential amino acids (especially sulfur‐containing amino acids) and micronutrients such as magnesium, manganese, and B‐vitamins (especially B9 and B12) [83, 84]. It is worth noting that during pet food heat processing, known as the Maillard reaction, that is, a nonenzymatic browning and flavoring reaction, a reduction of essential amino acids, such as lysine, bioa‐ vailability occurs [85, 86]. Therefore, many pet diets might be at the risk of supplying less lysine than the animal may require. Hence, the understanding of nutritional benefits of functional foods currently available is of key importance for the owners to provide their pets with the

*p* < 0.05, respectively). Last but not least,

the time spent in activity and at rest (\*\**p* < 0.01 and \*

334 Superfood and Functional Food - An Overview of Their Processing and Utilization

**5. The "market stand" of functional pet foods**

Pet's well‐being and health also depend on gut microbiota, whose composition and activity is correlated to several diseases [95–97]. *Cats* and *dogs* harbor several bacterial species (with *Firmicutes*, *Bacteroidetes*, *Proteobacteria*, *Fusobacteria,* and *Eubacterium* as the predominant phyla [98–102]), which differ from each other but also from the same species [103]. As shown in human studies, gut microbiota also plays a key role in immune and food intake regulation [104, 105]. However, only a few studies have examined the impact of the diet on canine gut microbial population [106–108]. For example, Kerr and coworkers evidenced the lack of negative alterations of the microbiome in healthy *dogs* of different species (*Golden Retriever, Hound Mix, Pitbull Mix, Mixed, St. Bernard, Australian Cattle, Dalmatian, Pointer, Standard Poodle, Terrier*) fed on a cooked navy bean powder [109]. Cooked beans decreased *Actinobacteria* and *Fusobacteria* and increased *Firmicutes*. Therefore, it is reasonable to think that future dietary products for *dogs* will modulate the gut microbial population in order to treat or prevent some food‐related diseases (dysbiosis, leaky gut, intestinal bowel disease, irritable bowel syndrome, coeliac disease, sepsis, renal failure, autoimmune disease, peritonitis, and intestinal obstruc‐ tion). Recently, Park and coworkers monitored healthy *dogs* for 6 months [110]. The first group was fed *ad libitum* on commercial food, while the second was fed on a restricted amount of the same commercial food. Animals fed *ad libitum* resulted in obesity with high levels of trigly‐ cerides and cholesterol. The microbiota presented differences between the two groups of animals, while *Actinobacteria* and *Bacteroidetes* were the predominant microflora in animals receiving a restricted amount of food, animals fed ad libitum presented *Firmicutes*, *Fusobacteria,* and *Actinobacteria*. Thus, a targeted diet may promote changes in gut microbiota affecting the activity of specific beneficial microbes resulting in benefits for the overall dog's health. In this sense, a targeted diet might also be based on the use of prebiotics, for example, chicory, fructooligosaccharides, pectin, and polydextrose. Zentek and coworkers demonstrated that nine adult healthy *beagles* fed a diet supplemented with 3% chicory had more consistent stools with increased levels of *bifidobacteria* and decreased *Clostridium perfringens* and a lower fecal pH with respect to a protein‐rich diet [111]. Further, *cats* fed diet supplemented with FOS (4% of diet) showed increased concentrations of *bifidobacteria* and reduced count of *Escherichia coli*, while pectins (4% of diet) increased *C. perfringens* and *lactobacilli* concentrations [112]. Conversely, *dogs* fed a diet with low level of dietary fiber (beet pulp) for 2 weeks decreased *Fusobacteria* and increased *Firmicutes* [107]. Interestingly, feline diets, particularly rich in animal proteins and low‐carbohydrate plant‐based additives, promoted fecal *Clostridium*, *Faecalibac‐ terium*, *Ruminococcus*, *Blautia*, and *Eubacterium* growth affecting circulating hormones and metabolites of key importance in satiety and host metabolism [113, 114]. As to polydextrose, its consumption by *dogs* resulted in an increase of fecal acetate, propionate, and total SCFA concentrations, while fecal pH, indole, and *C. perfringens* population decreased [106]. Thus, incorporating prebiotics in pets' diet may beneficially modulate gut microbiota and intestinal health and possibly protect the animals from enteric infections.
