The etiology of chronic non-communicable diseases such as obesity is multifactorial, including genetics and the environment (1). These diseases are the world’s leading cause of mortality (2, 3). Data from the World Health Organization (WHO) estimate the worldwide prevalence of 1.9 billion overweight adults [body mass index (BMI) ≥25 kg/m²] and more than 650 million people with obesity (BMI ≥30 kg/m²) (3). Recently, during the critical phase of the pandemic, the prevalence of chronic non-communicable diseases was closely related to higher mortality in hospitalization due to coronavirus disease 2019 (COVID-19) (4). These alarming data highlight the high and growing prevalence of obesity that represents a risk factor for the development and aggravation of other metabolic diseases.

Considering this, the incessant search for nutritional intervention strategies is essential to pursue the mitigation of obesity and its comorbidities. In this sense, the use of natural products and derivatives as non-pharmacological strategies can be an alternative in the prevention and treatment of acute or chronic diseases. These tools could contribute to preventing or reducing obesity-related negative outcomes, promoting longer life spans and better-quality lives (5). Thus, the use of food supplements, nutraceuticals, and natural integrative therapies has grown compared to conventional medications, which are still necessary and indispensable for the adequate treatment of some conditions (6, 7). Indeed, non-pharmacological therapies seem to be promising strategies for the prevention and treatment of metabolic–inflammatory diseases, such as obesity, metabolic syndrome, dyslipidemia, and type 2 diabetes (8–10).

Nevertheless, in this study, we also intend to contribute to the construction of knowledge for clinical practice using anthropometric assessment and serum tests reachable to practitioners’ daily routines. Previous studies have shown that increased neck circumference in obese adults is related to increased cardiovascular and inflammatory risk, acting as an anthropometric predictor of cardiovascular and inflammatory risk (11). Also, the waist-to-height ratio (WHtR), a measure with significant statistical representation, works as a good predictor of metabolic syndrome and appears to be directly proportional to inflammatory levels and correlated with poor eating habits (12–14).

Several cellular mechanisms are involved in obesity pathologies such as the metabolic pathways activated by directly influencing the insulin signaling in insulin-target tissues such as the liver, eventually leading to insulin resistance (15). Thus, nonalcoholic fatty liver disease (NAFLD) is frequently considered the hepatic outcome of type 2 diabetes. Indeed, insulin resistance also plays a central role in the development of the fatty liver (16), since hepatic fatty acid oxidation is reduced by mitochondrial dysfunctions triggered by obesity, resulting in hepatic accumulation of triacylglycerols (17, 18). Another common metabolic alteration in obesity is persistent and systemic low-grade inflammation (19, 20), which has been termed metainflammation. Metainflammation is closely linked to insulin resistance mechanisms that feedback each other, perpetuating long-term metabolic dysfunction (21). The combination of low-grade inflammation processes and insulin resistance establishment is an important link between obesity and its comorbidities.

The increased incidence and prevalence of obesity-related comorbidities is a concern still unsolved by worldwide public health regulators. Despite efforts to mitigate obesity, little effect has been noticed on the increase of all diseases associated with it (22). Considering this, the benefits of several natural compounds have been investigated in the obesity research field nowadays. In this sense, the effect of isolated prebiotic compounds, such as fructooligosaccharides (FOSs) (23), galactooligosaccharides (GOSs) (24), and yeast β-glucans (25, 26), has been related to improved metabolic markers recovering the health status. Also, supplementation with minerals such as magnesium (27), zinc (28), and selenium (29) and plant-derived compounds such as Silybum marianum (silymarin) (30) has displayed positive effects in obesity models. Compounds such as FOS and GOS display a potential effect of modulating the gut microbiome, which is closely linked to inflammatory modulation (31). The same aspect might be related to yeast β-glucans (32), magnesium (27), zinc (28), and selenium (33), which are known for the immune system-strengthening action. Likewise, silymarin, the bioactive compound obtained from milk thistle (S. marianum), has been used over the centuries to primarily treat liver diseases and protect the liver against obesity injury, contributing to the restoration of hepatic function (34). The main bioactive compound of silymarin is the component called silybin. Silybin is a flavonolignan that has different activities such as antioxidant and anti-inflammatory properties (35). In this sense, preclinical protocols testing a combination of these compounds have shown their benefits in the improvement of mitochondrial activity and key inflammatory molecules involved in the pathogenesis of several non-communicable chronic diseases such as type 2 diabetes, obesity, and cardiovascular diseases (9, 10). The preclinical model compared the effect of the supplement components separately and in combination, and findings suggest that the nutraceutical compounds might have a stronger synergic effect, representing an innovative tool for obesity and insulin resistance management.

Therefore, in this study, we aim to apply our previously developed supplement composition containing yeast β-glucan, prebiotics, minerals, and S. marianum (milk thistle or silymarin) (9, 10) as a tool to modulate inflammatory and metabolic responses in a population with obesity. The main hypothesis is that the supplements can improve and supply essential micronutrients such as zinc, magnesium, and selenium, which are usually deficient in populations with obesity (36), providing an improvement of fatty liver diseases through metabolic and endocrine pathways.

The application of natural products or their derivatives for the prevention or treatment of several pathologies represents a reasonable and accessible way for clinical practitioners and the general population to use natural resources as viable alternative therapies for health promotion and disease intervention. The development of new products containing different molecules and natural derivatives that interact synergistically has shown beneficial potential, acting holistically and producing few adverse effects on the human body (5). Our recently published findings show the synergistic effects between yeast β-glucan, prebiotic, minerals (selenium, zinc, magnesium), and S. marianum of an innovative supplement with nutraceutical properties in a preclinical model of diet-induced obesity (9, 10). These preclinical investigations have shown that after 4 weeks of supplementation, the new nutraceutical reduced fasting glycemia, insulin, HOMA-IR, HOMA-β, dyslipidemia, ectopic fat deposition, and hepatic fibrosis levels. It was probably mediated by molecules such as Peroxisome proliferator- activated receptor gamma (PPARg) coactivator 1 α (PGC-1α), interleukin-6 (IL-6), and IL-10, since our previous preclinical study has shown the modulation of this pathway by gene and protein expression changes (9, 10). The glutathione-S-transferase activity status was also modulated by the supplement displaying antioxidant effects. The preclinical studies also display enhanced metabolism promoted by molecules such as PPARα, adenosine monophosphate-activated protein kinase (AMPK)-PGC-1α, and Mitochondrial transcription factor A (TFAM) signaling pathways. Notably, the cortisol/C-reactive protein ratio, a well-characterized marker of the hypothalamic–pituitary–adrenal axis immune interface status, was found to be modulated by the supplement (9, 10).

Based on these preclinical studies and aiming to build up the evidence pyramid in this rapidly growing area of research, we have initiated a double-blind randomized clinical trial applying this supplement formula to participants who were overweight or obese. In this clinical trial, we began by focusing on evaluating clinical, biochemical, and anthropometric parameters. This is relevant because it brings scientific criteria for the investigation and analysis of parameters affordable to clinical practitioners who work in the area of obesity prevention and treatment. In this way, we hope to generate knowledge and scientific support for the advancement and updating of clinical practice in this research area.

Regarding the results of our randomized double-blind clinical trial, it is important to note that there was no indication of dietary interventions, encouragement of physical activity, or sort of a change in the participants’ lifestyle. Therefore, our results reflect the effects that can be directly and exclusively attributed to the consumption of the nutraceutical formula applied in the study.

The characterization of the anthropometric and demographic data shows that there are no significant differences between the groups at the beginning of the study. This is relevant to state the similarity of both groups in our sample to guarantee the reliability of data. Analyzing the anthropometrics among the supplementation groups at different time points, it is noteworthy that the WC reduction in both groups was promoted by the supplementation throughout the time. Despite the lack of weight loss, the reduction of WC represents a marker of decreased cardiovascular risk for the participants. It is well established that high WC reflects abdominal fat deposition, and it is strongly associated with the development of non-communicable chronic diseases (42). The indicators of abdominal obesity are better to discriminate inflammatory status and metabolic risk than the indicator and generalized obesity as BMI. In this sense, considering the cutoff points of WHtR and WHR preconized by health guidelines, it is possible to note that both markers decreased from the T0 time point to T180 time point, returning to low-risk values. The WHtR and WHR are easily collected anthropometric measures and reliable indicators for the diagnosis of obesity mainly related to increased visceral adiposity. Thus, they also represent a relevant clinical parameter to assess cardiovascular risk that is strongly related to visceral fat deposition. When increased, WHtR is a good predictor of metabolic syndrome and appears to be directly proportional to the inflammatory indicators and directly correlated with poor eating habits (12–14). Also, the neck circumference was slightly reduced by both supplements at the end of the treatment compared to the beginning; this finding is related to the natural course of obesity, since the volunteers were instructed not to change their lifestyle throughout the supplementation period, in addition to not having nutritional intervention and lifestyle changes in this study. Results from previous studies show that increased neck circumference in obese adults is related to increased cardiovascular and inflammatory risks, acting as an anthropometric predictor of cardiovascular and inflammatory risks for this population (11).

In our data, the supplements with a combination of minerals, fibers, and bioactive compounds were able to restore low cardiovascular risk levels without the need for medicines or dietary interventions. It might be related to high metabolic rates and an increase in energy expenditure promoted by the supplement compounds that are described to enhance molecules related to fatty acid metabolism activation (9). In a review, (43) described the effect of reducing adiposity by intake of bioactive natural compounds in obese populations. In particular, compounds such as FOSs (44) and GOSs (45) have shown an adiposity-reducing effect through activation of β-oxidation markers, such as acetyl CoA-carboxylase (ACC) inactivation, AMPK activation, and mitochondrial oxidation via allosteric regulation of carnitine palmitoyl transferase 1 (CPT-1) (43). In this sense, studies that used acute bioactive compound supplementation found similar results without changes in body weight, but in other specific biomarkers (46, 47). These changes may not yet be evident in anthropometric data; however, these results allow us to believe that long-term consumption could exert more pronounced effects on weight loss or parameters such as WHtR and BMI. Thus, in clinical practice, these anthropometric changes might be a good sign of improvement in cardiometabolic status.

Considering the biochemical parameters analyzed in serum samples collected at the end of the experimental protocol, there were no significant changes in serum concentrations of total cholesterol and its fractions or TGs. The improvement of the serum lipoprotein profile through the consumption of nutraceutical compounds such as fibers and antioxidants is recognized in the literature as a strategy to control dyslipidemia and prevent the onset of cardiovascular diseases, among other metabolic changes (48). Preclinical models of obesity that chronically consumed different bioactive compounds showed improvement in serum lipids (TG, HDL-cholesterol, and LDL-cholesterol) and insulin sensitivity associated with weight (49–52). Specifically, the supplement formula with silymarin can improve insulin sensitivity and reduce pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-α) and IL-6 in mice (9, 10). It might be attributed to silymarin compounds such as silibinin, which has been proven to be an immune modulator in vivo, inhibiting hepatic activation of Nuclear Factor-kappaB and consequently modulating the production of pro-inflammatory cytokines such as TNF-α, interferon-gamma, IL-4, IL-2, and Inducible nitric oxide synthase (iNOS) and upregulating anti-inflammatory cytokines such as IL-10 (35). However, these effects may also be related to the increased intake of daily fiber from FOSs and GOSs and antioxidant compounds by the supplement tested. Furthermore, these findings were not confirmed in the clinical study, showing the limitation of preclinical studies that are not able to mimic factors ranging from preexisting diseases and lifestyle, genetic predispositions and variability, and even factors such as the composition of the intestinal microbiota that can influence the response to supplement consumption.

To investigate other markers related to NAFLD, serum concentrations of AST and ALT as well as the AAR were evaluated. AST, ALT, and gamma-GT are accessible clinical markers to check for liver damage. It is known that ALT is found mainly in the hepatocyte cytoplasm, while AST is present in the mitochondria. Therefore, in cases of mild liver damage, ALT is altered in the serum, while in severe liver damage, there is an increase in AST in the bloodstream (53). Despite the absence of statistical differences in gamma-GT, it might represent an indicator of risk for advanced fibrosis in NAFLD and must be carefully watched by clinical practitioners (40). Based on this, our results allow us to suggest that there was no severe liver damage despite the mild liver damage displayed by both groups at all time points. It is important to consider that obesity and overweight are a strong risk factor for NAFLD per se, as the participants started the study with a high level of ALT, which was maintained until the end of the study. It is also noteworthy that the new supplement had its consumption safety reinforced, since, over time, there were no worsening or severe liver damage signs in this study. In fact, the AAR was reduced by the supplementation over time. These data are interesting, since, in alcoholic liver disease, the AAR is usually higher than 2, while in NAFLD, this index tends to be between 1 and 2. Recently, some studies have proposed the use of the AAR as the main laboratory parameter for the diagnosis of NAFLD. NAFLD is a chronic inflammatory liver disease with the evolutionary potential to cirrhosis. Currently, cases of cirrhosis in obese patients, previously diagnosed as cryptogenic, have been attributed to the evolution of NAFLD (41). Thus, AAR is an extremely relevant clinical parameter that reflects and helps in the diagnosis of hepatic complications. The supplement recovers the AAR, promoting hepatoprotective effects. This effect might be promoted by the supplement and can be explained by its rich composition. In this sense, the application of silymarin extract stands out by its hepatoprotective effects, enhancement of insulin receptor (IR) sensitivity, and immune modulation, besides its beneficial effects against cardiovascular and neurological diseases and some types of cancer among other positive effects (34). Also, the yeast β-glucan is an immunomodulatory molecule and might also contribute to the hepatoprotective effect (54). Notably, there are many drug therapies available in the pipeline that have been shown to be effective in the treatment of NAFLD/Nonalcoholic steatohepatitis (NASH) (55). However, nutraceutical supplements and integrative therapies should be considered as options for clinical frontline professionals as complementary tools and not substitutes for well-established drug treatments.

In obesity, the classic chronic low-grade inflammatory state, also known as metainflammation, is associated with increased inflammatory processes; the reactive oxygen species (ROS) is also increased. We did not find alterations in the C-reactive protein; however, this can be considered a nonspecific inflammatory marker and therefore not very sensitive to the discreet changes promoted by the metainflammation of obesity. Hyperglycemia and hyperinsulinemia are also known to trigger an inflammatory process and oxidative stress environment (56). In parallel, insulin resistance is also established by visceral fat deposition that increases the activation of the lipolysis process (57, 58). In turn, an increased flow of portal vein free fatty acids (FFAs) occurs, exposing the liver to a greater influx of FFAs modifying hepatic metabolic patterns, contributing to the maintenance of systemic hyperinsulinemia and insulin resistance (59). As observed in our results, insulin resistance comes alongside overweight/obesity and represents a persistent metabolic issue with a major concern for practitioners due to its complex management.

Accessing the endocrine profile, we found a significant reduction in cortisol levels promoted by the new supplement tested. Cortisol is a hormone well known for its key role in responding to stressful stimuli. Despite its physiological role, in obesity, it is known that high levels of cortisol promote body weight gain and increase appetite with a preference for energy-dense food, contributing to a high cardiometabolic risk. Also, chronic exposure to glucocorticoids such as cortisol is known to promote the specifically abdominal obesity metabolic syndrome and eventually cardiovascular diseases. In addition, the reduction in cortisol is related to the improvement in sleep quality, which is also directly linked to the prevalence of obesity (60). Our results demonstrate an improvement in cortisol levels, contributing to a reduction in abdominal obesity and a better metabolic prognosis without any lifestyle change intervention. In this way, the new supplement proves to be a tool capable of modulating endocrine pathways of cortisol production.

In this way, our primary clinical data allow us to conclude that the new supplement has an innovative combination of nutrients and bioactive compounds capable of promoting the improvement of biochemical and endocrine parameters in overweight/obesity. Its effect was pronounced, since it was able to promote metabolic improvement without the association of lifestyle change therapies such as diet intervention and physical activity. Although the nutrients of this supplement are accessible through a healthy diet, epidemiological data show that the consumption of these nutrients is usually deficient in a large part of the population. Thus, supplementation proves to be an important alternative to maintain adequate intake levels when the diet does not meet this demand. Therefore, the supplementation of beneficial compounds can be a valuable tool for preventing the onset of chronic diseases in the long term, avoiding the need for medication treatments. Nevertheless, these data represent an initial study of this new supplement. For more robust conclusions to be obtained, further studies are necessary with a larger sample size of both genders equally distributed. Further research also needs to address refined and in-depth biomolecular analyses that would provide a basis for a discussion on the reported clinical findings of robust cellular pathways, which might be considered another limitation of the present study. Nonetheless, the present results allow us to suggest that the parameters analyzed in this study can also be evaluated in clinical practice as an indication of endocrine and metabolic changes, bringing the research for the development of new nutraceuticals closer to the clinical practice of medical professionals, which has proven to be a major barrier between scientific research and practice application.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

The studies involving human participants were reviewed and approved by the HC-FMUSP Research Ethics Committee under the CAAE number 39984320.5.0000.0068 and registered as a Clinical Trial under identification number NCT04810572 (ClinicalTrials.gov). The patients/participants provided their written informed consent to participate in this study.

Conceptualization, AS and AP; Methodology, JF, DM and AP; Methodology, JF and DM; Investigation, VN and AP; Resources, VN; Data curation, ET, FP-B, and AP; Writing—original draft preparation, AS, JF and AP; writing—review, editing and visualization, AE-G, ES, and DO; Supervision, AP and JO; Project administration, AP; funding acquisition, VN. All authors contributed to the article and approved the submitted version.