Antioxidant, anti-inflammatory and anti-obesity effects of onion and its by-products in high-fat fed rodents: a systematic review

Abstract The effects of onion and its by-products on metabolic changes induced by excessive consumption of a high fat diet have been the focus of many studies. The aim of this study was to systematically review the effects of onion and its by-products antioxidant, anti-inflammatory and anti-obesity in rats exposed to a high-fat diet. Five databases were used: Pubmed, EMBASE, Science Direct, Web of science and Scopus until June 2020 updated December 1, 2022. Research of the articles was carried out by two reviewers, searching and selecting studies after an initial reading of the titles and abstracts. In total, 2,448 papers were found and, after assessing against the inclusion and exclusion criteria, 18 papers were selected for this review. The findings of this review show the beneficial effects of onion and its by-products on inflammatory parameters, obesity, cardiovascular disease, thermogenesis and hepatic alterations generally associated with the consumption of a high-fat diet.


Introduction
Obesity is a multifactorial disease that is growing in the world's population (Popkin et al., 2012;Popkin, 2015).Among the factors associated with this pathology, modern eating habits, characterized mainly by the excessive consumption of ultra-processed foods with a high saturated fat content, have been associated with the appearance of metabolic alterations resulting in the development of diseases (WHO, 2003;Popkin et al., 2012;Popkin, 2015).Obesity induced by the excessive dietary intake of fat is associated with abdominal fat accumulation, adipocyte size expansion, and the triggering of inflammatory processes, and may also predispose the individual to the development of insulin resistance, and non-alcoholic fatty liver disease (NAFLD), among other things (Lam et al., 2012;Dinh et al., 2015;Umekawa et al., 2015;Perez et al., 2015;

Search strategy and study selection
The evaluation of titles, abstracts, and complete articles was performed between May and June 2020 updated December 1, 2022, following the steps of identification, screening, and eligibility with respect to the inclusion criteria, with no limitation of the year of publication.
The paper research was performed in the electronic databases Pubmed, EMBASE, ScienceDirect, Web of Science and Scopus, using a combination of the terms MeSH (Medical Subject Headings): high fat diet, high-fat diet, occidental diet, western diet, onion, Allium cepa, rat, mouse, mice.The boolean operators "AND" and "OR" were used to cross terms: (high fat diet OR high-fat diet OR occidental diet OR western diet) AND (onion OR Allium cepa) AND (rat OR mouse OR mice).
Research of the articles was carried out by two reviewers, searching and selecting studies after an initial reading of the titles and abstracts.In cases of disagreement, a third reviewer was used to verify whether the study was eligible.

Quality and risk-of-bias assessment
The quality of the articles used in this review was evaluated according to the guidelines of ARRIVE -Animals in Research: Reporting In Vivo Experiments (Percie Du Sert et al., 2020).To evaluate the adequacy of the papers according to the guidelines, a scoring system (0 -no; 1 -yes) was used for the 20 items listed in the ARRIVE guidelines.Information about each study analyzed is summarized in Table 2.
To assess the risk of bias, the Systematic Review Center for Laboratory animal Research (SYRCLE) (Hooijmans et al., 2014), risk of bias tool was used for the 18 studies selected for this review.The "yes" judgment indicates that the risk of bias is low and the "no" judgment indicates a high degree of risk of bias.If the details of the report are insufficient to assess the appropriate risk of bias, the judgment will be "unclear" Table 3.

Results
A total of 2,448 papers were selected from the five databases searched.Of these, 347 were excluded due to duplicated titles and 2,064 after reading the abstracts, as (Pimentel et al., 2013;Zou et al., 2017;Porras et al., 2017;Barroso et al., 2019).These substances, present in vegetables and fruits, are part of a large group of bioactive compounds that are divided into subclasses, flavonoids being the most studied, especially quercetin (Slimestad et al., 2007;Hamauzu et al., 2011;Lee et al., 2012;Moon et al., 2013;Nabavi et al., 2015;Forney et al., 2018;Fraga et al., 2019;Grzelak-Błaszczyk et al., 2020).Among vegetables, the onion (Allium cepa L.), a popular consumer product that is rich in quercetin and also in other phenolic compounds such as kampferol, protocatechuic acid, anthocyanin and dietary fiber, has been the focus of some studies due to its effects antioxidant, anti-inflammatory and anti-obesity characteristics (Slimestad et al., 2007;Hamauzu et al., 2011;Lee et al., 2012;Moon et al., 2013;Nabavi et al., 2015;Forney et al., 2018;Fraga et al., 2019;Grzelak-Błaszczyk et al., 2020).Experimental studies have shown the beneficial effects of the consumption of onion peel powder, bulb, or even tea, on NAFLD, inflammation, cardiovascular disease, body weight, and insulin resistance induced by the consumption of an excessive high-fat diet (Hamauzu et al., 2011;Moon et al., 2013;Emamat et al., 2016Emamat et al., , 2018)).
However, despite the evidence already established with regard to the benefits of using and dose onion or its by-products to reduce the consequences of a high-fat diet intake, no systematic review has been performed to date.Thus, the aim of this study was to systematically review the effects of onion and its by-products antioxidant, anti-inflammatory and anti-obesity in rats subjected to a high-fat diet.

Methods
The systematic review was carried out following the recommendations of the Preferred reporting items for Systematic reviews and MetaAnalysis (PRISMA) (Moher et al., 2009).Registered the protocol in PROSPERO having as registration number is CDR42020188172.The record can be accessed at https://www.crd.york.ac.uk/PROSPERO.

Eligibility criteria
The research question was defined according to the PICOS approach (Participants / Population, Interventions / Exposure, Comparison, Outcomes and Study design) Table 1.
Studies that investigated the effect of the high-fat diet and supplementation of the onion or its by-products on any result related to the health of rats or mice were included.The comparison groups were defined as follows: use of a diet rich in fat versus supplemented with onion and its by-products (in any dose or quantity and any duration of consumption / exposure).In addition, to be included, studies needed to report any health-related results.No restrictions for language were applied.Reviews, human studies, and studies with drugs or other substances unrelated to the use of onion or its by-products were excluded.The identified studies were evaluated according to the inclusion criteria: studies with rats or mice that were exposed to a high-fat diet in the early stages or during adulthood.= no statistically significant effect.they did not fit the defined inclusion criteria.After the full-text reading, of the 37 remaining articles, 18 studies were considered eligible and were included in the review (Figure 1).
The results tabulated in the present study were: identification of the work (main author and year); species of animal that served as the study model; age and gender; high-fat diet composition; period of exposure to the diet; onion type and experimental methodology used; exposure period of onion, bulb, oil or tea or peel powder; dosage used of onion, bulb, oil or tea or peel powder; and main results (Table 2).
In 83% of the articles, male animals were used, 1 study used female animals and 2 studies did not report the gender of the animal.In 66.6% of the studies Wistar or Sprague-Dawley rats were used and in 33.3% C57BL/6 and Balb/c mice, aged three to nine weeks, were considered young animals.The percentage of fat in the high-fat diets varied between 5.1% and 60% and the period of exposure to this diet varied between 4 and 18 weeks.The period of exposure to onion or its by-products varied between 4 and 12 weeks.
Of the studies selected for this review, 13% showed an increase in the total concentration of short chain fatty acids (SCFA), of intestinal enzymes such as β-glucosidase and β-glucuronidase and of propionic and butyric acid in groups supplemented with onion or its by-products (Grzelak-Błaszczyk et al., 2018, 2020).
In addition, it should be noted that the papers that composed this review reached 50% to 85% of the items recommended by the ARRIVE protocol (Table 2).In assessing the risk of bias, it was observed that 50% of the questions in the SYRCLE protocol, were positively filled by all selected Figure 1.Flowchart of the search result in the information sources, the selection and inclusion of the original articles in the systematic review (Moher et al., 2009).
articles, considering a low risk of bias.However, all studies showed inadequacies in 20% of the questions related to Allocation Concealment and Performance Blinding, presenting a high risk of bias.It was also observed that 30% of the questions related to Random outcome assessment, Blinding of outcome assessment and Selective outcome reporting, were unclear in all studies in this review (Table 3).

Discussion
Excessive consumption of high saturated fat diets is associated with obesity, predisposing the individual to the appearance of metabolic and inflammatory changes.One way to mitigate the negative effects caused by the consumption of a high-fat diet is the addition of onion or its by-products in the diet, as onion contains elements such as quercetin, a well-studied flavonoid that may contribute to the prevention or treatment of diseases.The most studied forms of quercetin are aglycone (free form), quercetin monoglycosides, and quercetin diglycosides.The onion peel has a higher concentration of quercetin aglycone compared to the whole onion, which may be related to the hydrolysis of quercetin glycosides during the formation of the peel (Grzelak-Błaszczyk et al., 2018;Xu et al., 2019).This vegetable and its by-products are rich in fiber and also contain other flavonoids, such as kampferol and protocatechuic acid, both of which exert their own health benefits (Slimestad et al., 2007;Hamauzu et al., 2011;Benítez et al., 2012;Lee et al., 2012;Moon et al., 2013;Matsunaga et al., 2014;Emamat et al., 2016Emamat et al., , 2018;;Grzelak-Błaszczyk et al., 2018;Yang et al., 2018).This systematic review evaluated a number of studies that investigated the effects of onion or its by-products singly or in combination with garlic oil, either at different doses or exposure periods, but always added to a high-fat diet.Studies have also shown that onion or its by-products, reduced the deleterious health effects of high-fat diet (Figure 2).
All selected papers in this review used male or female rats or mice, aged three to nine weeks, being considered young animals.These animals were exposed to a highfat diet with onion peel powder, bulb, oil, leaves or tea added at different periods, ranging from 4 to 18 weeks.The amount of onion peel and leaves powder also varied between the authors studied, from 0.2% to 20%, onion extract varying in mg (17mg) and percentage (0.21% to 0.44%) and the onion peel tea varied from 46.3 mg/kg to 92 mg/kg animal weight.Independent of the amount of onion or its by-products used, the introduction of this vegetable brought benefits to the supplemented animals.
To analyze the quality of the papers, we used the ARRIVE protocol, specific for experimental studies (Percie Du Sert et al., 2020).Most articles in this review were found to meet more than 50% of the ARRIVE checklist with most of the criteria not met being related to the absence of facilities description, animal veterinary conditions, and the description of scientific implications such as study limitations.
Figure 2. Graphical Abstract.Onions and their by-products mitigated the effects of consuming a high-fat diet in different organs, tissues, and cells.In the intestine, there was an increase in intestinal enzymes, short-chain fatty acids, and a decrease in digestive pH.In the liver, the degree of steatosis was reduced, as well as liver enzymes and the expression of TNF-α, a pro-inflammatory cytokine.In the heart, there was a reduction in the rates of atherosclerosis I and II, in the modulation and elasticity of the aorta, and an improvement in the lipid profile.At the mitochondrial level, there was an increase in Pgc-1 and a decrease in mitochondrial genes with quercetin supplementation but not with onion extract.There was an increase in genes specific for brown adipose tissue in white adipose tissues, genes such as UCP-1, PRDM16, and CIDEA.Thus, the use of these products may have additional implications for the prevention of pathologies related to obesity.
There was a risk of bias in the selected studies when related to Allocation Concealment and Performance Blinding.Some studies were evaluated with unclear risk for evaluation Random outcome assessment, Blinding of outcome assessment and Selective outcome reporting.However, most articles had a low risk of bias in 50% of the questions in the SYRCLE assessment.
Onion is known for its antioxidant, anti-obesity, antiinflammatory, and prebiotic powers.These attributes are mainly due to the presence of quercetin, a highly studied flavonoid with important functional characteristics (Moon et al., 2013;Nabavi et al., 2015;Forney et al., 2018;Fraga et al., 2019).This flavonoid is capable of eliminating free radicals due to its effect on the potentiation of antioxidant enzymes such as glutathione (GSH) and superoxide dismutase (SOD) (Hayek et al., 1997;Erden Inal and Kahraman, 2000;Kobori et al., 2015).In this process, the donation of electrons contributes to stabilizing these free radicals, thus reducing the oxidative stress and consequent associated diseases (Hayek et al., 1997;Erden Inal and Kahraman, 2000;Kobori et al., 2015).Quercetin, for example, can reduce the oxidation of LDL molecules due to the elimination of free radicals, thus preventing diseases such as atherosclerosis (Ji et al., 2019).
In this review, 22% of the selected articles showed an improvement in the lipid profile, especially in LDL levels, of animals supplemented with onion oil compared to the group that was submitted only to a high-fat diet, confirming the protective effect of onion (Yang et al., 2018).Also, studies have demonstrated improvements in all lipid profiles, with a decreased serum determination of total cholesterol and TG, and an increase in HDL-C, confirming that animals benefited from the powder, tea, oil or extracts of onion concentrated of protocatechuic acid (Benítez et al., 2012;Matsunaga et al., 2014;Emamat et al., 2018;Yang et al., 2018;Grzelak-Błaszczyk et al., 2020;Yu et al., 2021;Chang et al., 2022;Momoh et al.,2022).
Cardiovascular protection associated with the consumption of powder, oil or extracts of onion concentrated with protocatechuic acid and quercetin monoglycosides was also observed in the heart structure of rats, which showed benefits in the atherosclerosis index, as well as decreasing the elasticity of the aorta of these animals, even when combined with a high-fat diet (Hamauzu et al., 2011;Yang et al., 2018;Grzelak-Błaszczyk et al., 2018, 2020).
Another important characteristic of onion is its anti-inflammatory effect.Inflammation may occur due to various stimuli, one of which may be the excessive consumption of a high saturated fat diet.This eating habit may predispose the individual to intestinal dysbiosis, with an increase in gram-negative bacteria, which has in its outermost layer the endotoxin called lipopolysaccharide (LPS).LPS binds to toll-like 4 membrane receptors (TLR4), increasing intestinal permeability, triggering inflammatory processes, and releasing pro-inflammatory cytokines such as TNFα and IL-6, as well as elevating nuclear factor kappa B (NF-Kb) levels (Poggi et al., 2007;Manco et al., 2010;Guo et al., 2013;Ma et al., 2017).Quercetin, present in onion, significantly inhibits these inflammatory effects.This was confirmed in this review, with 20% of the studies showing that the onion, rich in this flavonoid, reduces the excretion of inflammatory cytokines, mainly TNFα and IL-6 (Emamat et al., 2016(Emamat et al., , 2018;;Forney et al., 2018).
Increased intestinal permeability, induced by the consumption of a high fat-diet, may be associated with the development of NAFLD, due to the increase of gramnegative bacteria that release lipopolysaccharides LPS (Poggi et al., 2007;Manco et al., 2010;Guo et al., 2013).Increased LPS levels in the liver inhibit the fasting-induced adipose factor (Fiaf) that regulates lipoprotein lipase, whose function is to store fat in organs such as the liver and adipose tissue (Poggi et al., 2007;Aguirre and Venema, 2015;Larsen et al., 2019).Studies using powder, oil or extracts of onion concentrated with quercetin monoglycosides showed an improvement in the activity scores of NAFLD, improving the degree of steatosis, decreasing hepatic ALT and AST enzymes and lobular and portal inflammation (Emamat et al., 2016(Emamat et al., , 2018;;Yang et al., 2018;Grzelak-Błaszczyk et al., 2018;Yu et al., 2021;Chang et al., 2022;Momoh et al.,2022).
In addition to the benefits of quercetin, pectin, a soluble dietary fiber, is another important component present in onions, which may contribute to the balance of intestinal microbiota and the reduction of cardiovascular risks (Hamauzu et al., 2011;Benítez et al., 2012).This is because pectin is easily degraded by commensal bacteria in the gut, generating energy from the fermentation process and producing SCFA, primarily butyrate, which serve as an energy source for epithelial cells (Habinowski and Witters, 2001;Hamauzu et al., 2011;Guo et al., 2013).The formation of SCFA can inhibit the development of pathogenic bacteria and assist in intestinal immunity, improving their integrity and favoring adhesion of probiotic strains in the intestinal epithelium (Habinowski and Witters, 2001).
Fiber intake can provide benefits, such as reduced energy intake, increased satiety due to its viscosity, and gel formation, which may favor decreased intestinal absorption of food components such as fat, thereby reducing serum lipids and total cholesterol (Habinowski and Witters, 2001;Hamauzu et al., 2011;Benítez et al., 2012).In addition, the consumption of fiber-rich foods may benefit intestinal transit time with a decreased caecal pH due to fermentation by commensal bacteria producing SCFA, and thus improving constipation (Benítez et al., 2012;Grzelak-Błaszczyk et al., 2018).In this review, an increase in SCFA was observed in animals supplemented with onion extract concentrated in quercetin and quercetin monoglycosides, with an increase in the concentrations of propionic and butyric acid (Grzelak-Błaszczyk et al., 2018, 2020).There was also an increase in intestinal enzymes such as β-glucosidase and β-glucuronidase, which can be associated with bacterial adaptation to higher levels of quercetin glycosides present in the diet (Grzelak-Błaszczyk et al., 2018, 2020).
The anti-obesity effect is another functional characteristic of quercetin, present in onion, and some authors claim that this flavonoid has an antiadipogenic effect due to the activation of adenosine monophosphate-activated protein kinase (AMPK) (Berg et al., 2001;Devarshi et al., 2017).Quercetin can reduce adipocyte hyperplasia which is an important process for the development of metabolic syndrome (Berg et al., 2001;Devarshi et al., 2017).
Brazilian Journal of Biology, 2023, vol.83, e266108 15/18 Antioxidant, anti-inflammatory and anti-obesity effects of onion and its by-products In this review, than 38.8% of the studies found benefits from the addition of onion powder, tea, oil, or extract concentrated in quercetin and monoglycoside quercetin to the diet, including reduced body weight and weight of epididymal, perirenal, mesenteric, inguinal, and retroperitoneal adipose tissues (Lee et al., 2012;Benítez et al., 2012;Kim et al., 2012;Moon et al., 2013;Matsunaga et al., 2014;Henagan et al., 2015;Forney et al., 2018;Yang et al., 2018;Yu et al., 2021;Chang et al., 2022).With regard to food intake, the animals that were exposed to onion presented an increased food efficiency index and decreased consumption, possibly related to the satiety provided by the dietary fiber intake, which may contribute to a reduction in weight (Benítez et al., 2012;Kim et al., 2012).Also observed was an increased expression of adiponectin (in mesenteric adipose tissue), an adipokine secreted by adipocytes that exerts some protective effects which are anti-atherogenic and anti-inflammatory (Berg et al., 2001;Kim et al., 2012).The decreased expression of inflammatory genes such as Cd11b and Cd68 in inguinal adipose tissue and of McP-1 and IL-6 genes in epididymal adipose tissue in groups supplemented with onion extract attenuated the inflammatory effect of the high-fat diet (Forney et al., 2018).
The positive effects of onion can also be observed with the increased expression of the uncoupling protein 1 (UCP-1), important in the thermogenesis process, suggesting that this vegetable can stimulate lipid catabolism, promoting fat burning (Moon et al., 2013;Lee et al., 2017).This process can occur by the action of quercetin on white and brown adipose tissue, in which this flavonoid activates AMPK which in turn activates PPARγ (Lee et al., 2012;Kim et al., 2012;Moon et al., 2013).This subsequently interacts with the gamma 1 peroxisome proliferator-activated receptor (Pgc-1α), its coactivator increases UCP-1 transcription, contributing to the browning of white adipose tissue and the activation of brown adipose tissue (Moon et al., 2013;Devarshi et al., 2017;Lee et al., 2017;Choi et al., 2018).This browning can be observed by the presence of multilocular adipocytes in white adipose tissue presenting morphology and functions similar to those found in brown adipose tissue (Forney et al., 2018).
Other genes specific to brown adipose tissue showed greater expression in retroperitoneal and epididymal white adipose tissue, genes such as PRDM16 transcriptional coregulator necessary for the development of brown and beige adipocytes, and CIDEA an important protein in the formation of lipid droplets brown and beige adipose tissue (Harms et al., 2014;Barneda et al., 2015;Lee et al., 2017).
The onion extract was also able to increase the number of mitochondria in skeletal muscle, improving fat metabolism, insulin sensitivity, and increasing energy expenditure (Henagan et al., 2015;Emamat et al., 2016Emamat et al., , 2018)).There was also a reduction in mitochondrial genes Nd3, Nd4 (subunits of complex I), Cox1 (a subunit of complex IV) and Atp8 (a subunit of ATPsintase) in groups supplemented with quercetin, but not in those supplemented with onion extract, than can indicate changes in the oxidative phosphorylation system (OXPHOS) which is located in the inner mitochondrial membrane (Henagan et al., 2015;Bouchez and Devin, 2019).
The anti-obesity effect of onion and its by-products may be justified by the reduction of adipogenesis through pathways involved in lipid metabolism (Moon et al., 2013).Authors have shown in their experimental studies that the addition of onion peel extract reduced PPARγ expression as an adipogenesis transcription factor, thus contributing to decreased adipocyte proliferation (Moon et al., 2013).Also, PPARγ regulates the enzymes fatty acid synthase (FAS) and acetyl CoA carboxylase (ACC), consequently reducing the expression of these genes (Moon et al., 2013;Lee et al., 2017).
The studies that make up this review show methodological differences, such as the use of different by-products, dosages, and administration periods, which made it impossible to carry out a meta-analysis due to the heterogeneity of the data.This review, demonstrated the importance of consuming onion and its by-products.Benefits related to the inflammatory parameters, obesity, cardiovascular disease, thermogenesis, and liver disorders were observed, associated with the consumption of a high-fat diet.
In conclusion, was observed the beneficial effects of onion or its by-products on inflammatory parameters, obesity, cardiovascular diseases, thermogenesis and hepatic alterations associated with the consumption of a high-fat diet.Thus, the use of these products may have further implications in preventing obesity-related pathologies.Despite these beneficial effects, the minimum amount sufficient to bring benefits or the limit amount /excessive that were able of causing damage to health were not shown in the studies.Therefore, further studies are needed to determine these issues.

Table 1 .
PICOS criteria for inclusion and exclusion of studies.

Table 2 .
Described variables of the selected articles regarding the analysis of the effects of onion and its by-products in animals submitted to a high-fat diet.

Composition of the high fat diet high fat diet exposure period Onion characteristics and methodology used Period of exposure to onion or its by-products Quantity used of onion or its by-products Main results Final score of adequacy by ARRIVE
↓ statistically significant decrease.↑ statistically significant increase.TG: triglyceride; HDL-C: high density lipoproteins; LDL-C: low density lipoproteins; ALT: alanine aminotransferase; AST: aspartate aminotransferase; GGT: geranylgeranyltransferase; TNF-α: Tumor Necrosis Factor alpha; Pgc-1α: gamma peroxisome proliferator-

Table 3 .
Risk of bias of the included animal studies, assessed according to the SYRCLE guideline.