Effects of Regular Brazil Nut (Bertholletia excelsa H.B.K.) Consumption on Health: A Systematic Review of Clinical Trials

The Brazil nut (BN) is a promising food due to its numerous health benefits, but it is still necessary to systematically review the scientific evidence on these benefits. Thus, we examined the effects of regular BN consumption on health markers in humans according to the health state (with specific diseases or not) of the subjects. PubMed, Embase®, and Scielo databases were used to search for clinical trials. The PRISMA guideline was used to report the review, and the risk of bias for all studies was assessed. Twenty-four studies were included in the present review, of which fifteen were non-randomized. BNs were consumed in the context of a habitual free-living diet in all studies. Improvement in antioxidant status through increased levels of selenium and/or glutathione peroxidase activity in plasma, serum, whole blood, and/or erythrocytes was observed in all studies that evaluated antioxidant status, regardless of the health state of the sample. In addition, healthy subjects improved lipid markers and fasting glucose. Subjects with obesity had improvement in markers of lipid metabolism. Subjects with type 2 diabetes mellitus or dyslipidemia improved oxidative stress or DNA damage. Subjects undergoing hemodialysis benefited greatly from BN consumption, as they improved lipid profile markers, oxidative stress, inflammation, and thyroid function. Older adults with mild cognitive impairment improved verbal fluency and constructional praxis, and controversial results regarding the change in a marker of lipid peroxidation were observed in subjects with coronary artery disease. In conclusion, the benefits of BN consumption were found in different pathways of action and study populations.


Introduction
The Brazil nut (BN) (Bertholletia excelsa H.B.K.) is an edible seed of the Brazil nut tree native to the Amazonia biome. According to data provided by International Nut & Dried Fruit (INC), Bolivia is the main producer of BNs, followed by Peru and Brazil. The world consumption of BNs, considering a total supply of BNs minus ending stock, was 26,250 tons in 2020/2021 [1]. Like other nuts such as walnuts, hazelnuts, macadamia, pistachios, and almonds, several bioactive compounds are also present in BNs [2]. Unsaturated fats, minerals, vitamins, fibers, and phytochemicals give nuts potential and recognized beneficial health effects [3]. The BN contains smaller amounts of protein compared to plant-based protein sources such as tropical vegetables; however, in 100 g of BNs, 14.3 g is from protein.
Although small doses of BNs are recommended daily due to the selenium (Se) content (depending on the growing area, one unit is sufficient to reach the recommended daily intake of Se), BNs can be a potential alternative source of protein in diets [4]. However, unlike other nuts, the BN is the richest source of selenium (Se), a key element in forming antioxidant defense systems, modulation of the immune system, and helping to prevent ageing-related diseases [5]. In 2017, Cardoso et al. revised the health benefits of BNs and addressed their composition, safety, planting, processing, and use [3]. One study regarding BN safety and toxicity showed the absence of health risks regarding the presence of aflatoxins and radioactivity in 30 samples of BN originating from the Amazon [6]. In addition to the heavy metal barium, other elements may be present in BNs, such as strontium. However, the forms and levels of these elements in BNs do not seem harmful to human health [3,7,8]. Other review articles on BNs are also available. However, they have specific objectives and do not provide the most current findings of all studies that explore the effects of regular BN consumption on human health [3,[7][8][9][10][11]. Hence, we aim to carry out a systematic review regarding the effects of regular BN consumption on human health. We also discussed the mechanisms involved in these effects and critically assessed available literature and future perspectives on nutrition science.

Protocol and Registration
This systematic review was carried out per the "Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA)" guidelines [12] and was registered in PROSPERO (https://www.crd.york.ac.uk/prospero/, accessed on 11 September 2022), registration number CRD42021198965.

Literature Search
We identified studies by searching the following electronic databases: MEDLINE/PubMed (https://pubmed.ncbi.nlm.nih.gov/, accessed on 11 September 2022), Embase ® , and Scielo. The descriptors used were based on Medical Subject Headings (MeSH) (Appendix A Table A1). We conducted an exhaustive literature review using "intervention" from PICOS (population/intervention/comparator/outcome/study design) search criteria to identify all studies with the BN in any health outcome.
Filters were used to select studies on humans and clinical trials when available. We also searched for articles on this theme in the reference list of previously published review articles [3,[7][8][9][10][11]. Table A1 lists the search terms used. The last search was conducted in March 2022. The search was performed independently by two authors (AS and BKSS). First, titles and abstracts were selected. The articles were then read in full, and eligible studies were selected. Any disagreement between authors was resolved by consensus. Duplicate articles were removed using Mendeley ® software.

Eligibility Criteria
The following eligibility criteria were considered to answer our research question, "What are the effects of regular BN consumption on human health?" (i). Original clinical trials, randomized or not, controlled or not. (ii). Studies that evaluated the effects of regular BN consumption on any human health marker. (iii). Studies with any dose and time of intervention. (iv). Studies in which BNs were consumed as whole seed.
(v). Studies evaluating any health marker, anthropometrics, or body composition and those examining lipid, glucose, kidney, liver, and other markers. (vi). Studies with any subject profile (healthy subjects or subjects with comorbidities), and with any sex.
The following exclusion criteria were applied: (i). Postprandial studies.
(ii). Studies with children or animals, observational designs, reviews, congress abstracts, letters, protocol articles, notes, and in vitro analyses. (iii). Studies that did not investigate the effects of the BN on human health markers (iv). Interventions that supplemented BNs with minerals/vitamins or other nutritional enhancements. Studies with children were not considered due to different Se requirements compared to other ages.

Bias Risk Assessment
Two authors (AS and BVMF) independently assessed the risk of bias, following the Joanna Briggs Institute (JBI) Reviewer's Manual. This appraisal assesses methodological quality and determines the extent to which a study has addressed the possibility of bias in its design, conduct, and analysis. Thirteen questions for randomized clinical trials and nine questions for non-randomized clinical trials were answered for each study included in the systematic review. The answers to these questions were classified as yes, no, unclear, or not applicable [13]. A third author (BKSS) resolved disagreements.

Study Selection
We identified 463 records and removed 27 duplicates. During the titles and abstracts screening, 404 articles were excluded because they did not meet the eligibility criteria. Thirty-two articles remained for full-text evaluation. However, eight were excluded. Postprandial studies (n = 2), abstract congress (n = 1), unavailable articles (n = 1), intervention with granulated BN flour (n = 3), and child studies (n = 1) were excluded. Eleven articles were revised [3]. As result, 24 articles were incorporated per the eligibility criteria ( Figure 1). Tables 1 and 2 present the characteristics of these studies.

BNs Consumed by Healthy Subjects (Subjects without Diagnosed Diseases)
Regular (15 days to 12 weeks) consumption of BNs (1-11 units/day; 48 µ g to 862.65 µ g of Se) improved the antioxidant statuses (increased Se in BNs (whole seed) were consumed in a free-living habitual diet in all studies. The Se concentration in BNs varied from 48 µg in six units [18] to 1.261 µg only in one unit [19].

BNs Consumed by Healthy Subjects (Subjects without Diagnosed Diseases)
Regular (15 days to 12 weeks) consumption of BNs (1-11 units/day; 48 µg to 862.65 µg of Se) improved the antioxidant statuses (increased Se in plasma and erythrocytes, GPx, GPx3, SELENOP, and SELENOP mRNA expression), and lipid profiles (decreased TC and increased HDL-c cholesteryl reception) of healthy subjects and reduced their fasting glucose [16,17,21,30,32,38] (Figure 3). Similarly, acute consumption of 1-10 units/day of BNs (156 µg to 1560 µg of Se) associated with a normocaloric diet improved the antioxidant statuses (increased Se in plasma) and lipid profiles (increased HDL-c, decreased LDL-c, and atherogenic indices) in 24 postprandial hours up to 30 days after a single day of consumption [38,39]. Considering that the recommended plasma Se range is between 60-100 µL, only one study with healthy subjects observed that the participants were Sedeficient at baseline ( Figure 4A). In this study, Se-deficient subjects displayed an increased reception to cholesteryl esters by HDL-c after BN consumption [24].   Oxidative stress and related diseases, such as cardiovascular diseases (CVDs), are favored when antioxidant status is inferior to oxidizing agents in the body. Considering the importance of primary health care for disease prevention and health promotion, including BNs in the diet can help maintain the antioxidant status equilibrium by Se and related proteins [40,41]. In response to BN consumption, increased plasma SELENOP [34] and its expression [18] were observed in healthy subjects, confirming that SELENOP levels are a biomarker of Se status. Furthermore, an increase in GPx activity in the plasma and whole blood was observed in adult New Zealanders after 12 weeks, indicating the importance of Se in the production of antioxidant enzymes [14].
There is no evidence concerning the mechanisms involved in the beneficial effects of BN consumption on human lipid profiles. However, in experimental studies, Se supplementation downregulated Apo B and 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase expression in hypercholesterolemic rats [42]. This finding may potentiate a beneficial effect of BN consumption on the lipid profile, as HMG-CoA participates in the cholesterol biosynthesis pathway, while Apo B is the main protein component of LDLc. In another experimental study, co-supplementing rats fed a high-fat diet with Se and magnesium decreased blood and liver TG levels and TC/HDL-c and TG/HDL-c ratios while increasing antioxidant enzymes. Furthermore, co-supplementation inhibited hepatic lipogenesis gene expression, decreased HMG-CoA reductase, and increased cholesterol 7α-hydroxylase (CYP7A1) and lecithin cholesterol acyltransferase (LCAT) in the liver [43]. LCAT removes cholesterol from the blood and tissues and may help explain the increase in HDL-c and cholesteryl ester reception by HDL-c after BN consumption.
BN consumption decreased fasting glucose levels in healthy subjects [32]. In a study of non-diabetic subjects, supplementation with 200 µg of Se (as Se yeast) for six weeks reduced glycated hemoglobin but not fasting glucose [44]. In an observational study of participants with adequate Se status, a 10 µg/L increase in Se was associated with a 1.5% increase in insulin and a 1.7% increase in HOMA-IR [45]. Se status is an important determinant of the effect that Se supplementation will have on the body. In contrast, pistachio consumption showed beneficial effects on glucose metabolism markers in prediabetic subjects. The authors hypothesized that a synergistic effect of unsaturated fats, polyphenols, and carotenoids in the food matrix improved insulin sensitivity through the PI3K-AKT pathway [46]. Some studies have also explored the effect of polymorphisms in specific genes on the responses of healthy subjects to BN consumption, highlighting the influence of genetics on food consumption responses. TC decreased in the GA + AA and GG groups in subjects with polymorphism rs7579 in the SELENOP1 gene. However, allele A carriers tended to have higher cholesterol levels after consumption, mainly after four weeks of intervention. In addition, allele A carriers of polymorphism rs3877899 in the SELENOP1 gene had lower glucose levels at baseline and after eight weeks of intervention [32]. T allele (CT + TT) of the rs1050450 polymorphism in the GPx1 gene tended to have lower GPx activity than CC carriers. In contrast, subjects with polymorphism rs7579 in the SELENOP1 gene had increased plasma SELENOP1 concentrations after eight weeks of intervention, but with no difference between the GG and GG + AA alleles [34]. GPx1 mRNA expression increased in subjects with the CC genotype for rs1050450 after BN consumption, while it did not change in T allele carriers. SELENOP mRNA expression was higher in allele A carriers for rs7579 than in GG subjects after BN consumption [33,34]. SELENOS and SELENOF mRNA expression levels remained unchanged after BN consumption.

BN Consumption by Subjects with Obesity
Different effects were found in subjects with obesity who consumed BN depending on the amount of Se present in the BN. Consumption of three-five units/day of BNs (mean Se: 108.5 µg) increased serum Se levels compared to baseline. TC, TGs, and ox-LDL levels reduced after BN consumption, while red blood cell velocity increased compared to a placebo in adolescents with obesity after 16 weeks of consumption (Figure 3). No changes were observed in anthropometric and glucose metabolism markers, CRP, GPx-3, and 8-epi-PGF2α levels [15]. Another study with women with obesity observed an improvement in plasma Se, GPx activity, and HDL-c, and a decrease in atherogenic indices after participants consumed one unit/day of BN (290 µg of Se) for eight weeks [27]. However, the BNs had higher Se contents compared to the study performed by Maranhão et al. (2011) [15], and the subjects were Se-deficient at baseline ( Figure 4B). Moreover, improvements in plasma Se and GPx activity were observed in the Pro/Pro, Pro/Leu, and Leu/Leu genotypes of the Pro198Leu glutathione peroxidase polymorphism. In contrast, DNA damage decreased only in Pro/Pro genotype carriers versus the baseline, which was significantly lower than that in carriers of the Leu/Leu genotype [26].
As previously discussed, no human studies have explained the relationship between Se and lipid metabolism. However, animal studies have noted a reduction in HMG-CoA reductase after Se supplementation and consequently decreased endogenous cholesterol production [42]. The BN is a food matrix and, in addition to Se, also contains unsaturated fats, such as monounsaturated fatty acids (MUFAs), which have recognized functions in TC, TGs, and LDL-c in healthy subjects [47,48]. The benefits of the BN in reducing ox-LDLs, known oxidative stress marker and a risk factor for atherosclerosis [49], are easier to explain because Se is a non-enzymatic antioxidant and contributes to the selenoproteins' formation, adding to the redox balance by scavenging free radicals [50]. Excessive concentrations of these free radicals in the body can oxidize LDL-c, stimulating inflammation and leading to endothelial dysfunction in the intimal artery.
Weight and BMI did not decrease after regular BN consumption in subjects with obesity [15], suggesting that BN consumption cannot modulate weight when consumed in the context of a habitual diet, without energy restriction, for example. In addition, weight and BMI also did not increase. Studies have shown that despite the high-calorie density and high-fat content of nuts, habitually consuming nuts is not related to weight gain [51].
On the other hand, despite an increase in antioxidant status markers, women with obesity supplemented with one unit/day of BN containing approximately three times the upper limit (UL) of Se (400 µg) for eight weeks had increased pro-inflammatory cytokine expression and decreased GPx1 expression compared to the control group. Furthermore, increased miR-454-3p and miR-584-5p expression levels were observed [19,20]. The findings of this study reinforce that Se deficiencies and overload are harmful to health. It has been hypothesized that excessive Se consumption may play a pro-oxidant role by inducing reactive oxygen species (ROS), activating the Akt-NF-κB pathway, and further stimulating leukocytes and pro-inflammatory cytokine genes [52]. Simultaneously, the decrease in GPx expression can be explained by the fact that from a specific concentration of Se in the blood, GPx activity reaches its maximum [53]. Moreover, these studies note the beneficial health effects of consuming BN with Se contents below the tolerable UL for this population with high adiposity, reinforcing the need for attention regarding the choice of BN consumed based on Se content.
In summary, improvements in antioxidant status and lipid metabolism markers were observed after the minimal consumption of BNs containing an average Se of 108.5 µg in subjects with obesity ( Figure 3). Se doses above the UL have shown adverse effects in women with obesity.

BN Consumption by Subjects with Dyslipidemia, Type 2 Diabetes, or Coronary Artery Disease
An increase in inflammatory cytokines and oxidative stress is expected in dyslipidemia, diabetes mellitus, and CVD etiology and progression. In this context, Se deficiency is related to an increased risk of CVDs and mortality, possibly due to reduced selenoprotein synthesis and antioxidant capacity feeding back inflammation [54,55].
BN is a source of Se, MUFAs, polyphenols, and phytochemicals. However, a single unit of BN does not contain significant amounts of MUFAs and bioactive compounds. The nutrient that stands out in its composition is the Se because a single unit of BN can contribute five times more than the daily requirement for this mineral [10]. For this reason, we believe that most of the benefits achieved from BN intake are related to Se content (Table A2).
Consumption of one unit/day of BN (290 µg of Se) for 24 weeks improved antioxidant statuses (increased GPx activity and Se in the plasma and erythrocytes and reduced MDA and creatine kinase) in subjects using statins, while SELENOP mRNA expression was not affected [36] (Figure 3). Statins inhibit cholesterol synthesis via the mevalonate pathway, an important pathway for selenoprotein synthesis. Therefore, subjects who use statins are at an increased risk of developing Se deficiency and oxidative stress [56,57]. BN is a source of Se and may help maintain the nutritional status and oxidative balance of Se in patients using statins. Despite being instructed to maintain their usual diet, these subjects had reduced body weights and BMI. It is unlikely that the reductions in body weight and BMI were related to BN consumption. The authors reported that the effect of Se on BMI occurred under suboptimal Se conditions, which was not the case for the participants [36]. In addition, caloric restriction is necessary for weight loss, regardless of Se intake. However, studies have shown a relationship between selenium and adipogenesis [58], including increased lean mass and muscle mass [59]. Regarding the presence of polymorphisms, BN consumption increased GPX1 mRNA expression only in subjects with the rs1050450 CC genotype. SELENOP mRNA expression was significantly lower in subjects with the rs7579 GG genotype before and after the intervention.
No significant changes in Nrf2, NF-κB, NQO1, or peroxisome proliferator-activated receptor (PPARβ/δ) mRNA expression levels or markers of lipid metabolism and inflammation were found in subjects with coronary artery disease after three months of intervention with one unit/day of BNs (~290.5 µg of Se) compared to the control group. Controversial results regarding changes in TBARS were observed in the two articles on the effect of BN consumption in individuals with coronary artery disease [21,22] (Figure 3). The authors did not provide the baseline values of plasma Se values. This missing information is important to clarify the discussion because subjects with a normal Se status may not benefit from Se supplementation [55,60]. Another hypothesis is that these patients had multiple metabolic disorders and, therefore, required higher Se doses or extended interventions to benefit from BN consumption.
The consumption of one unit/day of BN (213.67 µg of Se) increased serum Se and decreased DNA damage ex vivo in subjects with type 2 diabetes treated with oral glucoselowering medication and insulin [35] (Figure 3). Again, BN consumption had benefits related to oxidative stress in a different population and was closely associated with Se status and selenoprotein synthesis.
Nevertheless, the subjects had increased waist circumferences and fasting blood glucose levels compared to the baseline, but the HbA1c levels remained unaltered [35]. Plasma Se and fasting glucose levels did not correlate. Considering that all the subjects were overweight and instructed to maintain their regular diet, these results can be attributed to the unbalanced diet. BN consumption was unable to improve glucose metabolism markers. Other nuts, especially pistachios, have been associated with better glucose control. This nut can modulate the insulin signaling pathway (via PI3K-AKT), favoring glycemic control [46]. However, higher doses (>40 g/day or 20% of total energy) may be needed [61].
Indeed, few interventions regarding BN intake have included participants with diabetes, dyslipidemia, or coronary artery disease. Hence, the evidence is limited. In this population, as in others, the effects of daily BN intake seem to be related to Se metabolism. Therefore, people with low plasma Se levels probably benefit more from BN consumption. Further investigations are necessary to understand the pathways involved.

BN Consumption by Subjects Undergoing Hemodialysis
A range of benefits could be observed in patients undergoing hemodialysis who regularly consumed one unit/day of BN (290 µg of Se) for three months. These benefits included enhanced non-enzymatic (increased Se in plasma and erythrocytes) and enzymatic (increased GSH-Px activity in erythrocytes and GPx activity in plasma) antioxidant defense systems, in line with the increased Nrf2 and NQO1 mRNA expression. In addition, the studies also showed improvements in inflammation (reduced Nf-κB mRNA expression, TNF, and IL-6 levels), oxidative stress (reduced malondialdehyde and 8-isoprostanes), and DNA damage (decreased 8-OHdG). There were also improved lipid profiles with an increased HDL-c and decreased atherogenic indices. Furthermore, elevated T3 and free thyroxine (FT4) levels were observed [19,34,35,37] (Figure 3). The same group of researchers in Brazil published all of these results. As Figure 4C presents, subjects undergoing hemodialysis were Se-deficient before BN supplementation (Figure 4).
Dialysis is a treatment for end-stage renal disease caused by chronic kidney disease, a debilitant disease associated with altered immune response, systemic inflammation, and oxidative stress. In addition to high oxidant production, subjects with chronic kidney disease have impaired antioxidative systems, favoring an oxidizing state [62]. These processes are responsible for the metabolic disorders observed in subjects with chronic kidney disease, such as dyslipidemias, who become susceptible to developing atherosclerosis, CVDs, and other complications [63]. Faced with a weakened state of health, subjects undergoing hemodialysis benefited greatly from BN consumption, a food matrix with a valuable nutritional profile, especially antioxidant compounds such as Se.
Although inflammation and oxidative stress are natural mechanisms in the body, they lead to the overproduction of pro-inflammatory mediators and oxidant agents when unregulated. For example, uremic toxins, such as indoxyl sulfate, produced by subjects with chronic kidney disease, are one of the agents responsible for activating NF-κB. Thus, proinflammatory cytokines, such as IL-6 and IL-1, are produced, and TNF, oxygen, and reactive hydrogen species feed a vicious cycle of stimulating the production of pro-inflammatory molecules and consequent inflammation exacerbation. Additionally, uremic toxins are linked to malondialdehyde and peroxynitrite production and also advanced glycation end products. These are related to NF-κB activation and a reduction in the activity of the transcription factor Nrf2 and, consequently, the underregulated expression of antioxidant enzymes [62]. These events favor a state of oxidative stress, making DNA susceptible to oxidative damage, as marked by an increase in 8-OH-dG.
Subjects with chronic kidney disease and end-stage renal disease are deficient in Se due to reduced intestinal absorption, loss during hemodialysis, and other causes. This worsens the depleting state of antioxidant agents. Therefore, BN supplementation seems to be interesting, improving the antioxidant, inflammatory, and oxidative stress status. However, 8-isoprostanes, 8-OHdG, IL-6, and TNF levels were significantly increased, whereas Se levels and GPx activity decreased 12 months after the intervention was interrupted [28,30], highlighting the importance of regular BN consumption. In addition, the kidneys participate in thyroid hormones' metabolism, which tends to be impaired in subjects with chronic kidney disease. As Se participates in thyroid hormone formation, BN supplementation increased T3 and FT4 levels in hemodialysis subjects.

BN Consumption by Subjects with MCIs
Consumption of one unit/day of BN (288.75 µg of Se) for 24 weeks increased Se levels in plasma and erythrocytes, plasma GPx activity, verbal fluency, and construction praxis in older adults with MCIs ( Figure 3). In addition, Se status was positively correlated with the Consortium to Establish a Registry for Alzheimer's Disease (CERAD) score, with higher scores indicating better cognitive performance [16].
Cognitive impairments and neurodegenerative diseases are related to oxidative stress caused by damage to the central nervous system. For this reason, Se deficiency can compromise the redox balance and reduce cognitive performance [64]. The authors have pointed out that most of the sample had Se deficiency at baseline, which seems to be a determining factor in achieving benefits from Se supplementation. However, in a population of older adults with adequate Se intake (85% of all participants), plasma Se levels were not associated with cognitive performance or neurotrophic factors [65].
Genetic factors also influence the responses to BN intake. For example, single nucleotide polymorphisms (SNPs) in genes encoding selenoproteins seem to negatively contribute to the protective effect against oxidative stress. In this sense, in subjects with MCIs, consuming BN daily (288.75 µg of Se) for 24 weeks increased GPx1 mRNA expression in carriers of a variant allele (CT + TT) for rs105045. Conversely, GPx1 mRNA expression was significantly reduced in rs7579 A-carriers and rs3877899 GG carriers [17].
The SNPs also affected SELENOP mRNA expression in MCI patients. While carriers of a variant allele (CT + TT) for rs1050450 showed increased SELENOP mRNA expression after treatment, no differences were observed for CC carriers [17]. Further studies are needed to clarify how polymorphisms would influence Se markers.
Studies on the effect of BN intake on MCIs are scarce. However, the available evidence shows superior benefits in people with low plasma Se values. In addition, Se deficiency can be a risk factor for cognitive impairment and polymorphisms that affect Se metabolism and utilization.

Future Perspectives
Currently, we know that BN consumption benefits human health, and seems to vary according to different routes of action and the health status of the sample evaluated. In addition, these findings refer to BN consumption in a free-living habitual diet without restrictions or adaptations. We emphasized above that individuals on hemodialysis benefited greatly from BN consumption. Many markers were evaluated in this sample of subjects undergoing hemodialysis, which does not mean that other studies did not observe the same benefits. This suggests that more studies are required to evaluate such markers also in other samples, with or without diseases. Considering that caloric restriction is one of the most reputable approaches in the literature for weight management, studies evaluating the effects of nut consumption in the context of caloric restriction are scarce. A recent study showed the superior effects of consuming of a mix of Brazilian nuts (30 g cashew nuts + 15 g BN) with a 500-calorie restricted diet in improving body composition and reducing the soluble adhesion molecule vascular cell adhesion protein 1 (VCAM-1) [66]. Thus, it would be interesting to conduct studies in this field using BN. In addition, as Se is a powerful antioxidant, studies evaluating longevity markers are of great relevance alongside those examining the intestinal microbiota and their response to adequate Se blood levels compared to inadequate levels. In view of the evidence regarding the interaction between nutrients and the intestinal microbiota, nuts have been investigated as modulators of the microbiota, since it is a food matrix rich in nutrients. However, to date, no study has investigated the effect of BN on the microbiota. Studies comparing the effect of BN consumption with isolated Se supplementation would also be of great interest because of limited evidence in this field.

Conclusions
Nowadays, scientific evidence supports the health benefits of regular BN consumption in a free-living habitual diet. This seems to vary according to the health status of the subjects, the Se content of the BNs, the Se levels of the participants before the intervention, and the presence of some polymorphisms. The benefits of BN consumption were found in different action pathways.    Composition of Brazil nut (BN) was evaluated through Association of Official Analytical Chemists (AOAC) and carbohydrate obtained by difference. Selenium concentrations were determined through inductively coupled plasma mass spectrometry.