Microencapsulated antioxidant stevia fraction fortifies whey protein and enhances its antidiabetic activity.

Whey protein was fortified with a microencapsulated fraction of Stevia rebaudiana, in the proportion 1:4 (w/w), with maltodextrin from the elite variety of Stevia UEM-13, rich in antioxidant compounds, and evaluated its antioxidant and antidiabetic potential in vitro. The fraction in ethyl acetate, the microencapsulated fraction, the whey protein obtained by membrane and a commercial whey protein were characterized and were also investigated solubility, microencapsulation efficiency and stability and digestion in vitro. In addition, these products and two formulations of the icroencapsulated fraction with the obtained whey protein were tested for their potential to inhibit the α-amylase and α-glucosidase enzyme (antidiabetic activity). The microencapsulated fraction (0.5%) and the supplement fortified with the 20% fraction microencapsulated showed inhibitory potential for the enzyme. As for the α-glucosidase enzyme, all products tested showed inhibition, with the formulation with 1.6% microencapsulated fraction added to whey protein being significantly higher. The microencapsulated fraction showed better solubility and stability, including in vitro digestion analysis, and showed antioxidant and antidiabetic capacity. A sensory evaluation was performed with panelists who regularly consume whey protein supplements and products with stevia and the supplement formulation with 1.6 g microencapsulated stevia per 100 g of whey protein have good sensory acceptance.

Whey protein enriched with Stevia rebaudiana fraction restores the pancreatic function of streptozotocin induced diabetic rats.

The study aims to analyse the treatment of whey protein enriched with Stevia rebaudiana fraction in insulin secretion and its role mitigating streptozotocin-induced hyperglycemia in rats. Thus, diabetic animals were treated with whey protein enriched with S. rebaudiana fraction or with only the protein isolate or only the Stevia fraction. Insulin level in plasma was measured by radioimmunoassay and the viability of ß cells was detected by immunohistochemistry. The results showed that diabetic animals treated with whey protein enriched with S. rebaudiana fraction had a greater recovery from insulinemia, with plasma levels similar to non-diabetic animals (~ 0.13 ng/mL). In addition, the same group showed a higher number of insulin-positive pancreatic B cells (~ 66%) in immunohistochemistry analysis, while the diabetic groups treated with only the fraction of stevia or whey protein showed 38 and 59% of positive cells, respectively. These results show that the treatment may have restored the viability of streptozotocin-injured pancreatic B cells, and consequently increased insulin secretion, suggesting whey protein enriched with S. rebaudiana fraction can be used an adjunct/supplement in diabetic treatment.

Fortification of the whey protein isolate antioxidant and antidiabetic activity with fraction rich in phenolic compounds obtained from Stevia rebaudiana (Bert.). Bertoni leaves.

A stevia fraction (ASF) free of steviol glycosides was extracted from Stevia rebaudiana leaves (Stevia UEM-13). ASF essentially constitutes phenolic compounds (52.42%), which were identified by liquid chromatography tandem mass spectrometry (LC-MS/MS) as caffeic acid, quercetin-3-o-glycoside, cyanidin-3-glucoside, kaempferol, quercetin, apigenin, rozmarinic acid, chlorogenic acid and dicaffeoylquinic acid. ASF was used as a multi-functional source of phenolic compounds to fortify the whey protein isolate (WPI) obtained by membrane separation. WPI fortified with 0.2% ASF showed an 80% increase in its antioxidant activity and more pronounced antidiabetic effects than the unfortified WPI, mainly in the glycemic control of diabetic animals induced by streptozotocin. The in vitro and in vivo antioxidant effects of ASF may enhance the effects of WPI. Indeed, this pioneering study revealed that ASF can be used to enrich the antioxidant and antidiabetic properties of WPI.

Whey protein sweetened with Stevia rebaudiana Bertoni (Bert.) increases mitochondrial biogenesis markers in the skeletal muscle of resistance-trained rats.

BACKGROUND: A combination of resistance training and whey protein supplementation is a common practice among athletes and recreational exercisers to enhance muscle growth and strength. Although their safety as food additives is controversial, artificial sweeteners are present in whey protein supplements. Thus, natural sweeteners extracted from the leaves of Stevia rebaudiana are a potential alternative, due to their safety and health benefits. Here, we investigated the effects of whey protein sweetened with S. rebaudiana on physical performance and mitochondrial biogenesis markers in the skeletal muscle of resistance-trained rats. METHODS: Forty male Wistar rats were distributed into four groups: sedentary rats, trained rats, trained rats receiving whey protein and trained rats receiving whey protein sweetened with S. rebaudiana leaf extracts. Resistance training was performed by climbing a ladder 5 days per week, during 8-weeks. The training sessions consisted of four climbs carrying a load of 50, 75, 90, and 100% of the maximum load-carrying capacity which we determined before by performing a maximum load-carrying test for each animal. After this period, we collected plasma and tissues samples to evaluate biochemical, histological and molecular (western blot) parameters in these rats. RESULTS: Dietary supplementation with whey protein sweetened with S. rebaudiana significantly enhanced the maximum load-carrying capacity of resistance-trained rats, compared with non-sweetened whey protein supplementation. This enhanced physical performance was accompanied by an increase in the weight of the gastrocnemius and soleus muscle pads. Although the muscle pad of the biceps brachii was not altered, we observed a significant increase in PGC-1α expression, which was followed by a similar pattern in TFAM protein expression, two important mitochondrial biogenesis markers. In addition, a higher level of AMPK phosphorylation was observed in these resistance-trained rats. Finally, supplementation with whey protein sweetened with S. rebaudiana also induced a significant decrease in retroperitoneal adipocyte diameter and an increase in the weight of brown adipose tissue pads in resistance-trained rats. CONCLUSION: The addition of Stevia rebaudiana leaf extracts to whey protein appears to be a potential strategy for those who want to increase muscular mass and strength and also improve mitochondrial function. This strategy may be useful for both athletes and patients with metabolic disorders, such as obesity and type 2 diabetes.

Cannabidiol improves metabolic dysfunction in middle-aged diabetic rats submitted to a chronic cerebral hypoperfusion.

Cannabidiol (CBD), a compound obtained from Cannabis sativa, has wide range of therapeutic properties, including mitigation of diabetes and neurodegeneration. Cerebral ischemia and consequent learning disabilities are aggravated in elderly diabetic subjects. However, there are no studies showing the effect of CBD treatment in elderly diabetes patients suffering cerebral ischemia. The present work tested the hypothesis that CBD treatment improves metabolic dysfunctions in middle-aged diabetic rats submitted to chronic cerebral hypoperfusion. In this work, 350-day-old male Wistar streptozotocin-induced diabetic rats were used. To induce cerebral ischemia was used a chronic cerebral hypoperfusion (CCH), surgically, via the four-vessel occlusion/internal carotid artery (4-VO/ICA). Four diabetic groups were established: Non-CCH Treated Diabetic (DNT), CCH Treated Diabetic (DCT), Non-CCH Vehicle Diabetic (DNV), and CCH Vehicle Diabetic (DCV). Vehicle groups were not treated with CBD. The animals were treated during 30 days with 10 mg CBD/Kg bw/day. After treatment, the animals were euthanized, and blood levels of glucose, insulin, total cholesterol, high density lipoprotein (HDL), low density lipoprotein (LDL), triglycerides, fructosamine, aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were evaluated. DCT group presented reduction of hyperglycemia and an increase of insulinemia. Also was observed lower fructosamine, LDL, HDL, triglycerides and total cholesterol levels. AST and ALT concentration were reduced in CBD treated groups. CBD may be used as therapeutic tool to protect metabolism against injuries from diabetes aggravated by cerebral ischemia.

Pretreatment with ethanol as an alternative to improve steviol glycosides extraction and purification from a new variety of stevia.

Leaves of a new variety of Stevia rebaudiana with a high content of rebaudioside A were pretreated with ethanol. The ethanolic extract showed high antioxidant potential and 39 compounds were identified, by UPLC/HRMS, among them one not yet mentioned in the literature for stevia leaves. From the in natura leaves and pretreated leaves, the conditions of aqueous extraction of steviol glycosides were investigated using response surface methodology. The aqueous extracts obtained were purified by ion exchange chromatography techniques and membrane separation methods. The recuperation of steviol glycosides was 4.02g for pretreated leaves and 2.20g for in natura leaves. The level of purity was, respectively, 87% and 84.8%. The results obtained demonstrate that pretreatment increases the yield and purity level of stevia sweeteners by the use of environmentally friendly methodologies and the final product presented acceptable sensory characteristics.

Stevia Nonsweetener Fraction Displays an Insulinotropic Effect Involving Neurotransmission in Pancreatic Islets.

Stevia rebaudiana (Bert.) Bertoni besides being a source of noncaloric sweeteners is also an important source of bioactive molecules. Many plant extracts, mostly obtained with ethyl acetate solvent, are rich in polyphenol compounds that present insulinotropic effects. To investigate whether the nonsweetener fraction, which is rich in phenolic compounds isolated from Stevia rebaudiana with the solvent ethyl acetate (EAF), has an insulinotropic effect, including interference at the terminals of the autonomic nervous system of the pancreatic islets of rats. Pancreatic islets were isolated from Wistar rats and incubated with EAF and inhibitory or stimulatory substances of insulin secretion, including cholinergic and adrenergic agonists and antagonists. EAF potentiates glucose-stimulated insulin secretion (GSIS) only in the presence of high glucose and calcium-dependent concentrations. EAF increased muscarinic insulinotropic effects in pancreatic islets, interfering with the muscarinic receptor subfamily M3. Adrenergic inhibitory effects on GSIS were attenuated in the presence of EAF, which interfered with the adrenergic α2 receptor. Results suggest that EAF isolated from stevia leaves is a potential therapy for treating type 2 diabetes mellitus by stimulating insulin secretion only in high glucose concentrations, enhancing parasympathetic signal transduction and inhibiting sympathetic signal transduction in beta cells.

Functional properties and sensory testing of whey protein concentrate sweetened with rebaudioside A / Propriedades funcionais e teste sensorial de concentrado proteico do soro do leite adoçado com rebaudiosídeo A

ABSTRACT Objective: To develop a natural dietary product with functional benefits for diabetic patients. Whey protein concentrate was obtained through the separation membrane processes and sweetened with rebaudioside A. This product was submitted to sensory testing in humans and used to evaluate possible functional properties in male Wistar rats models with diabetesMellitus induced by streptozotocin. Methods: Two concentrates were produced. Only the second showed protein content of 74.3 and 17.3% of lactose was used as supplementation in induced diabetic rats. This concentrate was obtained from the concentration by reverse osmosis system (180 k Daltons), followed by nanofiltration in a 500 k Daltons membrane and spray drying at 5.0% solution of the first concentrate developed. The concentrate was sweetened with rebaudioside A (rebaudioside A 26 mg/100 g concentrate). All procedures were performed at the Center for Studies in Natural Products, at the Universidade Estadual de Maringá. Three experimental groups were established (n=6): two groups of diabetic animals, one control group and one supplemented group; and a control group of normal mice (non-diabetic). The supplemented group received concentrates sweetened with rebaudioside A in a dose of 100 mg/kg bw/day by an esophageal tube for 35 days. Fasting, the fed state and body weight were assessed weekly for all groups. At the end of the supplementation period, the following were analyzed: plasma parameters of glucose, total cholesterol, triglycerides and fructosamine; the serum levels of aspartate aminotransferase and alanine aminotransferase, water and food intake. Organs and tissues were removed and weighed to assess mass and anatomical changes. Results: The product presented 74% of proteins and 17% of lactose and showed satisfactory sensory testing by the addition of 26 mg of rebaudioside A/100 g concentrate. Supplementation of the product reduced hyperglycemia, plasma fructosamine levels, triglycerides and total cholesterol, and improved body weight gain of streptozotocin-induced diabetic rats. Conclusion: Whey protein concentrate with substantial content of protein (above 70%) and low lactose was obtained through the membrane separation processes. The addition of rebaudioside A at the concentration of 26 mg/100 g rebaudioside A proved to be as sweet as sucralose with satisfactory sensory testing, which indicates that this is a non-caloric natural sweetener that can replace artificial sweeteners. The product (whey protein concentrate sweetened with rebaudioside A) presented important functional properties and reduced the metabolic disorders caused by the syndrome.

RESUMO Objetivo: Obtenção de um concentrado proteico do soro do leite por meio de processos de separação por membranas, adoçado com rebaudiosídeo A, análise sensorial em humanos e avaliação de propriedades funcionais em modelos de ratos induzidos por estreptozotocina. Métodos: Foram produzidos dois concentrados, mas apenas o segundo, que apresentou teor proteico de 74,3 e 17,3% de lactose, foi utilizado na suplementação de animais diabéticos. Esse concentrado foi obtido a partir da concentração em sistema de osmose reversa (180 Daltons), seguida de nanofiltração em membrana de 500 Daltons e secagem emspray dryer de uma solução a 5% do primeiro concentrado desenvolvido. O concentrado foi adoçado com rebaudiosídeo A (26 mg de rebaudiosídeo A/100 g de concentrado) obtido por meio da extração, separação e purificação das folhas de Stevia rebaudiana. Todos os processos foram realizados no Núcleo de Estudos em Produtos Naturais, da Universidade Estadual de Maringá. Foram estabelecidos três grupos experimentais (n=6): dois de animais diabéticos, um controle e outro suplementado; e um grupo de animais não diabéticos controle. O grupo suplementado recebeu o concentrado adoçado com rebaudiosídeo A na dose de 100 mg/kg peso corporal/dia, por sonda esofágica, por um período de 35 dias. Em todos os grupos, foram avaliadas semanalmente as glicemias de jejum e no estado alimentado, e o peso corporal. Ao final do período de suplementação, os parâmetros plasmáticos de glicose, colesterol total, triglicérides e da frutosamina; os valores séricos de aspartato aminotransferase e alanina aminotransferase e a ingestão hídrica e alimentar foram analisados. Órgãos e tecidos foram retirados e pesados a fim de se avaliarem alterações de massa e anatômicas. Resultados: O produto formulado apresentou 74% de proteínas e 17% de lactose e apresentou perfil sensorial satisfatório por meio da adição de 26 mg de rebaudiosídeo A/100 g de concentrado. A suplementação do produto reduziu a hiperglicemia, os níveis plasmáticos de frutosamina, triglicérides e colesterol total e ainda melhorou o ganho de peso corporal dos ratos diabéticos induzidos. Conclusão: Os processos de separação por membranas utilizados neste estudo proporcionaram a obtenção eficiente de um concentrado proteico do soro do leite, com teor proteico importante, superior a 70% e com baixo teor de lactose. A adição de rebaudiosídeo A ao produto na concentração de 26 mg/100 g de rebaudiosídeo A proporcionou dulçor equivalente ao da sucralose, com perfil sensorial satisfatório, o que indica que esse edulcorante natural não calórico tem potencialidade para substituir os artificiais. O produto obtido (concentrado proteico do soro do leite adoçado com rebaudiosídeo A) apresentou propriedades funcionais importantes, reduzindo alguns distúrbios metabólicos decorrentes dessa síndrome.