Combining Conventional Organic Solvent Extraction, Ultrasound-Assisted Extraction, and Chromatographic Techniques to Obtain Pure Betanin from Beetroot for Clinical Purposes.

Red beetroot extract (E162) is a natural colorant that owes its color to betanin, its major red pigment. Betanin displays remarkable antioxidant, anti-inflammatory, and chemoprotective properties mediated by its structure and influence on gene expression. However, the betanin employed in most preclinical assays is a beetroot extract diluted in dextrin, not pure betanin, as no isolated compound is commercially available. This makes its use inaccurate concerning product content estimates and biological effect assessments. Herein, a combination of conventional extraction under orbital shaking and ultrasound-assisted extraction (UAE) to purify betanin by semi-preparative HPLC was performed. The employed methodology extracts betalains at over a 90% yield, achieving 1.74 ± 0.01 mg of pure betanin/g beetroot, a 41% yield from beetroot contents increasing to 50 %, considering the betalains pool. The purified betanin exhibited an 85% purity degree against 32 or 72% of a commercial standard evaluated by LC-MS or HPLC methods, respectively. The identity of purified betanin was confirmed by UV-Vis, LC-MS, and 1H NMR. The combination of a conventional extraction, UAE, and semi-preparative HPLC allowed for betanin purification with a high yield, superior purity, and almost three times more antioxidant power compared to commercial betanin, being, therefore, more suitable for clinical purposes.

A Critical Review on Vasoactive Nutrients for the Management of Endothelial Dysfunction and Arterial Stiffness in Individuals under Cardiovascular Risk.

Pathophysiological conditions such as endothelial dysfunction and arterial stiffness, characterized by low nitric oxide bioavailability, deficient endothelium-dependent vasodilation and heart effort, predispose individuals to atherosclerotic lesions and cardiac events. Nitrate (NO3-), L-arginine, L-citrulline and potassium (K+) can mitigate arterial dysfunction and stiffness by intensifying NO bioavailability. Dietary compounds such as L-arginine, L-citrulline, NO3- and K+ exert vasoactive effects as demonstrated in clinical interventions by noninvasive flow-mediated vasodilation (FMD) and pulse-wave velocity (PWV) prognostic techniques. Daily L-arginine intakes ranging from 4.5 to 21 g lead to increased FMD and reduced PWV responses. Isolated L-citrulline intake of at least 5.6 g has a better effect compared to watermelon extract, which is only effective on endothelial function when supplemented for longer than 6 weeks and contains at least 6 g of L-citrulline. NO3- supplementation employing beetroot at doses greater than 370 mg promotes hemodynamic effects through the NO3--NO2-/NO pathway, a well-documented effect. A potassium intake of 1.5 g/day can restore endothelial function and arterial mobility, where decreased vascular tone takes place via ATPase pump/hyperpolarization and natriuresis, leading to muscle relaxation and NO release. These dietary interventions, alone or synergically, can ameliorate endothelial dysfunction and should be considered as adjuvant therapies in cardiovascular diseases.

Microencapsulated and Ready-to-Eat Beetroot Soup: A Stable and Attractive Formulation Enriched in Nitrate, Betalains and Minerals.

Beetroot is a tuber rich in antioxidant compounds, i.e., betanin and saponins, and is one of the main sources of dietary nitrate. The aim of the present study was to microencapsulate a ready-to-eat beetroot soup by lyophilization using different encapsulating agents, which supply the required amount of bioactive nutrients. Particle size distributions ranged from 7.94 ± 1.74 to 245.66 ± 2.31 µm for beetroot soup in starch and from 30.56 ± 1.66 to 636.34 ± 2.04 µm in maltodextrin. Microparticle yields of powdered beetroot soup in starch varied from 77.68% to 88.91%, and in maltodextrin from 75.01% to 80.25%. The NO3- and total betalain contents at a 1:2 ratio were 10.46 ± 0.22 mmol·100 g-1 fresh weight basis and 219.7 ± 4.92 mg·g-1 in starch powdered beetroot soup and 8.43 ± 0.09 mmol·100 g-1 fresh weight basis and 223.9 ± 4.21 mg·g-1 in maltodextrin powdered beetroot soup. Six distinct minerals were identified and quantified in beetroot soups, namely Na, K, Mg, Mn, Zn and P. Beetroot soup microencapsulated in starch or maltodextrin complied with microbiological quality guidelines for consumption, with good acceptance and purchase intention throughout 90 days of storage. Microencapsulated beetroot soup may, thus, comprise a novel attractive strategy to offer high contents of bioaccessible dietary nitrate and antioxidant compounds that may aid in the improvement of vascular-protective effects.

Acute Effects of Dietary Nitrate on Central Pressure and Endothelial Function in Hypertensive Patients: A Randomized, Placebo-Controlled Crossover Study. / Efeitos Agudos do Nitrato Dietético na Pressão Central e Desempenho Cardíaco em Hipertensos: Estudo Cruzado, Randomizado e Placebo-Controlado.

BACKGROUND: The diet's inorganic nitrate (NO3-) may provide a physiological substrate for reducing nitrate (NO2-) to NO independent of the endothelium. Studies suggest that inorganic NO3- has beneficial effects on cardiovascular health. OBJECTIVE: This study evaluated the acute effects of 500 mL nitrate-rich beetroot juice (BRJ; containing 11.5mmol NO3-) on blood pressure and endothelial function in treated hypertensive patients. METHODS: A randomized, placebo-controlled, crossover study was conducted in treated hypertensive patients (n=37; women=62%) who underwent clinical and nutritional evaluation and assessment of central hemodynamic parameters and microvascular reactivity. The significance level was p<0.05. RESULTS: The mean age was 59±7 years, and mean systolic and diastolic blood pressures were 142±10/83±9mmHg. There was a significant increase in the subendocardial viability ratio (SEVR; 149±25 vs. 165±30%, p<0.001) and reduction in ejection duration (ED; 37±4 vs. 34±4%, p<0.001) in the beetroot phase but no significant SEVR difference in the control phase. The % increase in perfusion (155 vs. 159 %, p=0.042) was significantly increased in the beetroot phase, which was not observed in the control phase. In the beetroot phase, the change in SEVR showed a significant correlation with the change in the area under the curve of post-occlusive reactive hyperemia (AUC-PORH) (r=0.45, p=0.012). The change in ED showed a significant correlation with the post-intervention perfusion peak (r=-0.37, p=0.031) and AUC-PORH (r=-0.36, p=0.046). CONCLUSIONS: The acute ingestion of BRJ by hypertensive patients resulted in an improvement of endothelial function, which was associated with higher subendocardial viability and performance in myocardial contraction.

FUNDAMENTO: O nitrato inorgânico (NO3­) da dieta pode fornecer substrato fisiológico para reduzir o nitrito (NO2­) a óxido nítrico (NO) independente do endotélio. Estudos sugerem que o NO3­ inorgânico tem efeitos benéficos na saúde cardiovascular. OBJETIVOS: Este estudo avaliou os efeitos agudos de 500 mL de suco de beterraba rico em nitrato (SB; contendo 11,5mmol NO3­) na pressão arterial e na função endotelial em pacientes hipertensos tratados. MÉTODOS: Estudo cruzado, randomizado, controlado por placebo foi realizado em pacientes hipertensos tratados (n=37; mulheres=62%) que foram submetidos à avaliação clínica e nutricional, avaliação dos parâmetros hemodinâmicos centrais e reatividade microvascular. O nível de significância foi p<0,05. RESULTADOS: A média de idade foi 59±7 anos e das pressões sistólica e diastólica foi de 142±10/83±9 mmHg. Houve aumento significativo na taxa de viabilidade subendocárdica (RVSE; 149±25 vs. 165±30%, p<0,001) e redução na duração da ejeção (DE; 37±4 vs. 34±4%, p<0,001) na fase beterraba, mas nenhuma diferença significativa de RVSE na fase controle. O % de aumento na perfusão (155 vs. 159%, p=0,042) cresceu significativamente na fase beterraba, o que não foi observado na fase controle. Na fase beterraba, a alteração da RVSE apresentou correlação significativa com a alteração da área sob a curva de hiperemia reativa pós-oclusiva (ASC-HRPO) (r=0,45, p=0,012). A mudança na DE mostrou uma correlação significativa com pico de perfusão pós-intervenção (r=-0,37, p=0,031) e ASC-HRPO (r=-0,36, p=0,046). CONCLUSÃO: A ingestão aguda de SB por pacientes hipertensos resultou em melhora da função endotelial, que foi associada à maior viabilidade subendocárdica e desempenho na contração miocárdica.

Efeitos Agudos do Nitrato Dietético na Pressão Central e Desempenho Cardíaco em Hipertensos: Estudo Cruzado, Randomizado e Placebo-Controlado / Acute Effects of Dietary Nitrate on Central Pressure and Endothelial Function in Hypertensive Patients: A Randomized, Placebo-Controlled Crossover Study

Resumo Fundamento O nitrato inorgânico (NO3-) da dieta pode fornecer substrato fisiológico para reduzir o nitrito (NO2-) a óxido nítrico (NO) independente do endotélio. Estudos sugerem que o NO3- inorgânico tem efeitos benéficos na saúde cardiovascular. Objetivos Este estudo avaliou os efeitos agudos de 500 mL de suco de beterraba rico em nitrato (SB; contendo 11,5mmol NO3-) na pressão arterial e na função endotelial em pacientes hipertensos tratados. Métodos Estudo cruzado, randomizado, controlado por placebo foi realizado em pacientes hipertensos tratados (n=37; mulheres=62%) que foram submetidos à avaliação clínica e nutricional, avaliação dos parâmetros hemodinâmicos centrais e reatividade microvascular. O nível de significância foi p<0,05. Resultados A média de idade foi 59±7 anos e das pressões sistólica e diastólica foi de 142±10/83±9 mmHg. Houve aumento significativo na taxa de viabilidade subendocárdica (RVSE; 149±25 vs. 165±30%, p<0,001) e redução na duração da ejeção (DE; 37±4 vs. 34±4%, p<0,001) na fase beterraba, mas nenhuma diferença significativa de RVSE na fase controle. O % de aumento na perfusão (155 vs. 159%, p=0,042) cresceu significativamente na fase beterraba, o que não foi observado na fase controle. Na fase beterraba, a alteração da RVSE apresentou correlação significativa com a alteração da área sob a curva de hiperemia reativa pós-oclusiva (ASC-HRPO) (r=0,45, p=0,012). A mudança na DE mostrou uma correlação significativa com pico de perfusão pós-intervenção (r=-0,37, p=0,031) e ASC-HRPO (r=-0,36, p=0,046). Conclusão A ingestão aguda de SB por pacientes hipertensos resultou em melhora da função endotelial, que foi associada à maior viabilidade subendocárdica e desempenho na contração miocárdica.

Abstract Background The diet's inorganic nitrate (NO3-) may provide a physiological substrate for reducing nitrate (NO2-) to NO independent of the endothelium. Studies suggest that inorganic NO3- has beneficial effects on cardiovascular health. Objective This study evaluated the acute effects of 500 mL nitrate-rich beetroot juice (BRJ; containing 11.5mmol NO3-) on blood pressure and endothelial function in treated hypertensive patients. Methods A randomized, placebo-controlled, crossover study was conducted in treated hypertensive patients (n=37; women=62%) who underwent clinical and nutritional evaluation and assessment of central hemodynamic parameters and microvascular reactivity. The significance level was p<0.05. Results The mean age was 59±7 years, and mean systolic and diastolic blood pressures were 142±10/83±9mmHg. There was a significant increase in the subendocardial viability ratio (SEVR; 149±25 vs. 165±30%, p<0.001) and reduction in ejection duration (ED; 37±4 vs. 34±4%, p<0.001) in the beetroot phase but no significant SEVR difference in the control phase. The % increase in perfusion (155 vs. 159 %, p=0.042) was significantly increased in the beetroot phase, which was not observed in the control phase. In the beetroot phase, the change in SEVR showed a significant correlation with the change in the area under the curve of post-occlusive reactive hyperemia (AUC-PORH) (r=0.45, p=0.012). The change in ED showed a significant correlation with the post-intervention perfusion peak (r=-0.37, p=0.031) and AUC-PORH (r=-0.36, p=0.046). Conclusions The acute ingestion of BRJ by hypertensive patients resulted in an improvement of endothelial function, which was associated with higher subendocardial viability and performance in myocardial contraction.

Toxic Metals and Metalloids in Infant Formulas Marketed in Brazil, and Child Health Risks According to the Target Hazard Quotients and Target Cancer Risk.

Children are highly vulnerable to chemical exposure. Thus, metal and metalloid in infant formulas are a concern, although studies in this regard are still relatively scarce. Thus, the presence of aluminum, arsenic, cadmium, tin, mercury, lead, and uranium was investigated in infant formulas marketed in Brazil by inductively coupled plasma mass spectrometry, and the Target Hazard Quotients (THQ) and Target Cancer Risk (TCR) were calculated in to assess the potential risk of toxicity for children who consume these products continuously. Aluminum ranging from 0.432 ± 0.049 to 1.241 ± 0.113 mg·kg-1, arsenic from 0.012 ± 0.009 to 0.034 ± 0.006 mg·kg-1, and tin from 0.007 ± 0.003 to 0.095 ± 0.024 mg·kg-1 were the major elements, while cadmium and uranium were present at the lowest concentrations. According to the THQ, arsenic contents in infant formulas showed a THQ > 1, indicating potential health risk concerns for newborns or children. Minimal carcinogenic risks were observed for the elements considered carcinogenic. Metabolic and nutritional interactions are also discussed. This study indicates the need to improve infant formula surveillance concerning contamination by potentially toxic and carcinogenic elements.

Macrominerals and Trace Minerals in Commercial Infant Formulas Marketed in Brazil: Compliance With Established Minimum and Maximum Requirements, Label Statements, and Estimated Daily Intake.

Infant formulas are the main nutritional source for infants when breastfeeding is not possible or recommended. The daily need for specific nutrients, such as essential minerals, in early stages of a child's life is high because of rapid infant growth and development, which impose metabolic flux increases on these pathways to support growth, physical activity, and defense against infections. In this context, this research aimed to determine macromineral and trace mineral contents in starting (phase 1) and follow-up (phase 2) infant formulas marketed in Brazil (n = 30) by inductively coupled plasma-mass spectrometry, calculate estimated daily intakes, and compare them to reference values regarding adequate intake and tolerable upper intake levels. The highest concentrations of macrominerals were observed in Ca, K, P, and Na, and trace minerals in Fe, Zn, Mn, and Cu. Certain homogeneity only to trace mineral contents was observed when analyzing inter-batch values from same manufacturers. In general, all phase 1 and phase 2 infant formula brands and batches met or exceeded Fe, Zn, Cu, Mo, and Se contents when compared to maximum limits established by Codex Alimentarius. In addition, Zn contents in eight phase 1 and in four phase 2 infant formulas were above the contents established by the tolerable upper intake level for children aged 0-6 and/or 7-12 months, respectively. These findings highlight the need to expand regular infant formula inspection concerning nutritional quality, as some composition aspects of these foods must be improved to follow international guidelines, since ideal requirements for infant formula composition, quality, and safety interfere in child development and adult health.

Bioactive Compounds from Pale Ale Beer Powder Attenuate Experimental Colitis in BALB/c Mice.

Phenolic compounds (PCs) present in foods are associated with a decreased risk of developing inflammatory diseases. The aim of this study was to extract and characterize PCs from craft beer powder and evaluate their potential benefits in an experimental model of inflammatory bowel disease (IBD). PCs were extracted and quantified from pure beer samples. BALB/c mice received either the beer phenolic extract (BPE) or beer powder fortified with phenolic extract (BPFPE) of PCs daily for 20 days by gavage. Colon samples were collected for histopathological and immunohistochemical analyses. Dextran sodium sulfate (DSS)-induced mice lost more weight, had reduced colon length, and developed more inflammatory changes compared with DSS-induced mice treated with either BPE or BPFPE. In addition, in DSS-induced mice, the densities of CD4- and CD11b-positive cells, apoptotic rates, and activation of NF-κB and p-ERK1/2 MAPK intracellular signaling pathways were higher in those treated with BPE and BPFPE than in those not treated. Pretreatment with the phenolic extract and BPFPE remarkably attenuated DSS-induced colitis. The protective effect of PCs supports further investigation and development of therapies for human IBD.

Betanin as a multipath oxidative stress and inflammation modulator: a beetroot pigment with protective effects on cardiovascular disease pathogenesis.

Oxidative stress is a common physiopathological condition enrolled in risk factors for cardiovascular diseases. Individuals in such a redox imbalance status present endothelial dysfunctions and inflammation, reaching the onset of heart disease. Phytochemicals are able to attenuate the main mechanisms of oxidative stress and inflammation and should be considered as supportive therapies to manage risk factors for cardiovascular diseases. Beetroot (Beta vulgaris L.) is a rich source of bioactive compounds, including betanin (betanidin-5-O-ß-glucoside), a pigment displaying the potential to alleviate oxidative stress and inflammantion, as previously demonstrated in preclinical trials. Betanin resists gastrointestinal digestion, is absorbed by the epithelial cells of intestinal mucosa and reaches the plasma in its active form. Betanin displays free-radical scavenger ability through hydrogen or electron donation, preserving lipid structures and LDL particles while inducing the transcription of antioxidant genes through the nuclear factor erythroid-2-related factor 2 and, simultaneously, suppressing the pro-inflammatory nuclear factor kappa-B pathways. This review discusses the anti-radical and gene regulatory cardioprotective activities of betanin in the pathophysiology of endothelial damage and atherogenesis, the main conditions for cardiovascular disease. In addition, betanin influences on these multipath cellular signals and aiding in reducing cardiovascular disorders is proposed.

Protein Quality in Infant Formulas Marketed in Brazil: Assessments on Biodigestibility, Essential Amino Acid Content and Proteins of Biological Importance.

Infant formulas, designed to provide similar nutritional composition and performance to human milk, are recommended when breastfeeding is not enough to provide for the nutritional needs of children under 12 months of age. In this context, the present study aimed to assess the protein quality and essential amino acid content of both starting (phase 1) and follow-up (phase 2) formulas from different manufacturers. The chemical amino acid score and protein digestibility corrected by the amino acid score were calculated. The determined protein contents in most formulas were above the maximum limit recommended by FAO and WHO guidelines and at odds with the protein contents declared in the label. All infant formulas contained lactoferrin (0.06 to 0.44 g·100 g-1) and α-lactalbumin (0.02 to 1.34 g·100 g-1) below recommended concentrations, whereas ĸ-casein (8.28 to 12.91 g·100 g-1), α-casein (0.70 to 2.28 g·100 g-1) and ß-lactoglobulin (1.32 to 4.19 g·100 g-1) were detected above recommended concentrations. Essential amino acid quantification indicated that threonine, leucine and phenylalanine were the most abundant amino acids found in the investigated infant formulas. In conclusion, infant formulas are still unconforming to nutritional breast milk quality and must be improved in order to follow current global health authority guidelines.

Increasing the Power of Polyphenols through Nanoencapsulation for Adjuvant Therapy against Cardiovascular Diseases.

Polyphenols play a therapeutic role in vascular diseases, acting in inherent illness-associate conditions such as inflammation, diabetes, dyslipidemia, hypertension, and oxidative stress, as demonstrated by clinical trials and epidemiological surveys. The main polyphenol cardioprotective mechanisms rely on increased nitric oxide, decreased asymmetric dimethylarginine levels, upregulation of genes encoding antioxidant enzymes via the Nrf2-ARE pathway and anti-inflammatory action through the redox-sensitive transcription factor NF-κB and PPAR-γ receptor. However, poor polyphenol bioavailability and extensive metabolization restrict their applicability. Polyphenols carried by nanoparticles circumvent these limitations providing controlled release and better solubility, chemical protection, and target achievement. Nano-encapsulate polyphenols loaded in food grade polymers and lipids appear to be safe, gaining resistance in the enteric route for intestinal absorption, in which the mucoadhesiveness ensures their increased uptake, achieving high systemic levels in non-metabolized forms. Nano-capsules confer a gradual release to these compounds, as well as longer half-lives and cell and whole organism permanence, reinforcing their effectiveness, as demonstrated in pre-clinical trials, enabling their application as an adjuvant therapy against cardiovascular diseases. Polyphenol entrapment in nanoparticles should be encouraged in nutraceutical manufacturing for the fortification of foods and beverages. This study discusses pre-clinical trials evaluating how nano-encapsulate polyphenols following oral administration can aid in cardiovascular performance.

Short-Term Betanin Intake Reduces Oxidative Stress in Wistar Rats.

Oxidative stress is a common condition described in risk factors for cardiovascular disease. Betanin, a bioactive pigment from red beetroot demonstrates anti-inflammatory and antioxidant properties. The main aim of this study was to evaluate the short-term intake of betanin against oxidative stress in a rodent model, a common condition described in several risk factors for cardiovascular disease. Oxidative stress was induced in Wistar rats by a hyperlipidemic diet for 60 days, followed by betanin administration (20 mg·kg-1) through oral gavage for 20 days. Plasma biochemical parameters and antioxidant enzyme activities were evaluated. Lipid peroxidation and histopathological changes were determined in the liver. The hyperlipidemic diet caused hyperglycemia, hyperinsulinemia, insulin resistance, and increases in alanine transaminase and aspartate transaminase levels. Oxidative stress status was confirmed by reduction of antioxidant enzyme activities, increased lipid peroxidation, and liver damage. Purified betanin regulated glucose levels, insulin, and insulin resistance. Hepatic damage was reversed as evidenced by the reduction in alanine transaminase and aspartate transaminase levels and confirmed by histological analyses. Betanin reduced hepatic malondialdehyde and increased superoxide dismutase, catalase, and glutathione peroxidase activities. Short-term betanin intake modulated biochemical parameters, reversed hepatic tissue damage, and attenuated oxidative stress in Wistar rats.

Polyphenols from Root, Tubercles and Grains Cropped in Brazil: Chemical and Nutritional Characterization and Their Effects on Human Health and Diseases.

Throughout evolution, plants have developed the ability to produce secondary phenolic metabolites, which are important for their interactions with the environment, reproductive strategies and defense mechanisms. These (poly)phenolic compounds are a heterogeneous group of natural antioxidants found in vegetables, cereals and leguminous that exert beneficial and protective actions on human health, playing roles such as enzymatic reaction inhibitors and cofactors, toxic chemicals scavengers and biochemical reaction substrates, increasing the absorption of essential nutrients and selectively inhibiting deleterious intestinal bacteria. Polyphenols present in some commodity grains, such as soy and cocoa beans, as well as in other vegetables considered security foods for developing countries, including cassava, taro and beetroot, all of them cropped in Brazil, have been identified and quantified in order to point out their bioavailability and the adequate dietary intake to promote health. The effects of the flavonoid and non-flavonoid compounds present in these vegetables, their metabolism and their effects on preventing chronic and degenerative disorders like cancers, diabetes, osteoporosis, cardiovascular and neurological diseases are herein discussed based on recent epidemiological studies.

Beetroot juice increase nitric oxide metabolites in both men and women regardless of body mass.

The nitrate (NO3(-)) present in beetroot juice (BJ) has been studied for its effect on the cardiovascular system by converting to nitric oxide (NO). In the present study, we evaluated the effect of BJ on the excretion of NO metabolites and its relationship with body mass in both men and women. NO metabolites - urinary NO3(-), nitrite (NO2(-)) and NOx were analyzed by using a high-performance liquid chromatography system. There were significant increases in urinary NO3(-), NO2(-) and NOx in BJ as compared to PLA (BJ without NO3(-)). No significant difference between men and women was observed in NO metabolites after BJ at any time point. There were no significant relationships between urinary NO3(-), NO2(-) and NOx and body mass in BJ intervention for both men and women. In conclusion, urinary NO metabolites after BJ consumption increases in similar manner between sexes regardless of body mass.