Selected coffee (Coffea arabica L.) extracts inhibit intestinal α-glucosidases activities in-vitro and postprandial hyperglycemia in SD Rats.

One of the therapeutic approaches for decreasing postprandial hyperglycemia is to retard absorption of glucose by the inhibition of carbohydrate hydrolyzing enzymes, α-amylase, and α-glucosidases, in the digestive organs. Coffee consumption has been reported to beneficial effects for controlling calorie and cardiovascular diseases, however, the clear efficacy and mode of action are yet to be proved well. Therefore, in this study we evaluated in- vitro rat intestinal α-glucosidases and porcine α-amylase inhibitory activities as well as in vivo (Sprague-Dawley rat model) blood glucose lowering effects of selected coffee extracts. The water extracted Sumatra coffee (SWE) showed strong α-glucosidase inhibitory activity (IC50, 4.39 mg/mL) in a dose-dependent manner followed by Ethiopian water extract (EWE) (IC50, 4.97) and Guatemala water extract (GWE) (IC50, 5.19). Excepted for GWE all the coffee types significantly reduced the plasma glucose level at 0.5 h after oral intake (0.5 g/kg-body weight) in sucrose and starch-loaded SD rats. In sucrose loading test SWE (p < 0.001) and EWE (p < 0.05) had significantly postprandial blood glucose reduction effect, when compared to control. The maximum blood glucose levels (Cmax) of EWE administration group were decreased by about 18% (from 222.3 ± 16.0 to 182.5 ± 15.4, p < 0.01) and 19% (from 236.2 ± 25.1 to 191.3 ± 13.2 h·mg/dL, p < 0.01) in sucrose and starch loading tests, respectively. These results indicate that selected coffee extract may improve exaggerated postprandial spikes in blood glucose via inhibition of intestinal sucrase and thus delays carbohydrate absorption. These in vitro and in vivo studies therefore could provide the biochemical rationale for the benefit of coffee-based dietary supplement and the basis for further clinical study.

Anti-Obesity and Anti-Adipogenic Effects of Administration of Arginyl-Fructose-Enriched Jeju Barley (Hordeum vulgare L.) Extract in C57BL/6 Mice and in 3T3-L1 Preadipocytes Models.

In our previous study, we reported that arginyl-fructose (AF), one of the Amadori rearrangement compounds (ARCs) produced by the heat processing of Korean ginseng can reduce carbohydrate absorption by inhibiting intestinal carbohydrate hydrolyzing enzymes in both in vitro and in vivo animal models. This reduced absorption of carbohydrate might be helpful to control body weight gain due to excessive carbohydrate consumption and support induced calorie restriction. However, the weight management effect, except for the effect due to anti-hyperglycemic action, along with the potential mechanism of action have not yet been determined. Therefore, the efforts of this study are to investigate and understand the possible weight management effect and mechanism action of AF-enriched barley extracts (BEE). More specifically, the effect of BEE on lipid accumulation and adipogenic gene expression, body weight gain, body weight, plasma lipids, body fat mass, and lipid deposition were evaluated using C57BL/6 mice and 3T3-L1 preadipocytes models. The formation of lipid droplets in the 3T3-L1 treated with BEE (500 and 750 µg/mL) was significantly blocked (p < 0.05 and p < 0.01, respectively). Male C57BL/6 mice were fed a high-fat diet (30% fat) for 8 weeks with BEE (0.3 g/kg-body weight). Compared to the high fat diet control (HFD) group, the cells treated with BEE significantly decreased in intracellular lipid accumulation with concomitant decreases in the expression of key transcription factors, peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer-binding protein alpha (CEBP/α), the mRNA expression of downstream lipogenic target genes such as fatty acid binding protein 4 (FABP4), fatty acid synthase (FAS), and sterol regulatory element-binding protein 1c (SREBP-1c). Supplementation of BEE effectively lowered the body weight gain, visceral fat accumulation, and plasma lipid concentrations. Compared to the HFD group, BEE significantly suppressed body weight gain (16.06 ± 2.44 g vs. 9.40 ± 1.39 g, p < 0.01) and increased serum adiponectin levels, significantly, 1.6-folder higher than the control group. These results indicate that AF-enriched barley extracts may prevent diet-induced weight gain and the anti-obesity effect is mediated in part by inhibiting adipogenesis and increasing adiponectin level.

Evaluation of a Witch Hazel Extract for the Potential Prebiotic and Protective Effect on Select Lactiplantibacillus plantarum (Prev. Lactobacillus plantarum) Strains.

Witch hazel extract has been evaluated in prior studies demonstrating the phenolic-mediated biofilm inhibition, toxin production inhibition, and growth inhibition in Staphylococcus aureus. In this study, we are evaluating the possible prebiotic and protective effect of witch hazel extract on select probiotic Lactiplantibacillus plantarum strains, namely L. plantarum LP 10241 and L. plantarum LPBAA-793. When the prebiotic effect was evaluated, we observed that the tested extract had prebiotic effect at the higher tested dose (0.5%) on LPBAA-793 strain (8.7 log CFU/mL after 18 h compared to 5.1 log CFU/mL with the control) and on LP 10241 strain (7.7 log CFU/mL after 18 h compared to 4.4 log CFU/mL with the control). For the evaluation of the protective effect of witch hazel extract on the select strains, we subjected nutrient depletion stress under aerobic conditions and monitored the cell death with and without addition of witch hazel extract. We observed that the tested extract had a significant protective effect on LPBAA-793 strain (4 log CFU/mL after 12 days, compared to no growth with control) and a slighter protective effect against LP 10241 strains (6.3 log CFU/mL in day 2 compared to 4.3 log CFU/mL with control). The results from this research provide for the first time the rationale that while witch hazel extract has significant antimicrobial, anti-toxin production and anti-biofilm activities on pathogenic microorganisms, it might play an important and positive role on health-beneficial probiotic bacteria.

The Effect of Tannin-Rich Witch Hazel on Growth of Probiotic Lactobacillus plantarum.

Probiotic bacteria help maintain microbiome homeostasis and promote gut health. Maintaining the competitive advantage of the probiotics over pathogenic bacteria is a challenge, as they are part of the gut microbiome that is continuously exposed to digestive and nutritional changes and various stressors. Witch hazel that is rich in hamamelitannin (WH, whISOBAXTM) is an inhibitor of growth and virulence of pathogenic bacteria. To test for its effect on probiotic bacteria, WH was tested on the growth and biofilm formation of a commercially available probiotic Lactobacillus plantarum PS128. As these bacteria are aerotolerant, the experiments were carried out aerobically and in nutritionally inadequate/poor (nutrient broth) or adequate/rich (MRS broth) conditions. Interestingly, despite its negative effect on the growth and biofilm formation of pathogenic bacteria such as Staphylococcus epidermidis, WH promotes the growth of the probiotic bacteria in a nutritionally inadequate environment while maintaining their growth under a nutritionally rich environment. In the absence of WH, no significant biofilm is formed on the surfaces tested (polystyrene and alginate), but in the presence of WH, biofilm formation was significantly enhanced. These results indicate that WH may thus be used to enhance the growth and survival of probiotics.

Immune Modulatory Activities of Arginyl-Fructose (AF) and AF-Enriched Natural Products in In-Vitro and In-Vivo Animal Models.

The immune system plays an important role in maintaining body homeostasis. Recent studies on the immune-enhancing effects of ginseng saponins have revealed more diverse mechanisms of action. Maillard reaction that occurs during the manufacturing processes of red ginseng produces a large amount of Amadori rearrangement compounds (ARCs), such as arginyl-fructose (AF). The antioxidant and anti-hyperglycemic effects of AF have been reported. However, the possible immune enhancing effects of non-saponin ginseng compounds, such as AF, have not been investigated. In this study the effects of AF and AF-enriched natural product (Ginofos, GF) on proliferation of normal mouse splenocytes were evaluated in vitro and male BALB/c mice models. The proliferation of splenocytes treated with mitogens (concanavalin A, lipopolysaccharide) were further increased by addition of AF (p < 0.01) or GF (p < 0.01), in a dose dependent manner. After the 10 days of oral administration of compounds, changes in weights of spleen and thymus, serum immunoglobulin, and expression of cytokines were measured as biomarkers of immune-enhancing potential in male BALB/c mice model. The AF or GF treated groups had higher weights of the thymus (0.94 ± 0.25 and 0.86 ± 0.18, p < 0.05, respectively) than that of cyclophosphamide treated group (0.59 ± 0.18). This result indicates that AF or AF-enriched extract (GF) increased humoral immunity against CY-induced immunosuppression. In addition, immunoglobulin contents and expression of cytokines including IgM (p < 0.01), IgG (p < 0.05), IL-2 (p < 0.01), IL-4 (p < 0.01), IL-6 (p < 0.01), and IFN-γ (p < 0.05) were also significantly increased by supplementation of AF or GF. These results indicate that AF has immune enhancing effects by activation of adaptive immunity via increase of expression of immunoglobulins and cytokines such as IgM, IgG, IL-2, IL-4, IL-6 and thereby proliferating the weight of thymus. Our findings provide a pharmacological rationale for AF-enriched natural products such as ginseng and red ginseng that can possibly have immune-enhancement potential and should be further evaluated.

Anti-Obesity and Anti-Adipogenic Effects of Chitosan Oligosaccharide (GO2KA1) in SD Rats and in 3T3-L1 Preadipocytes Models.

Excess body weight is a major risk factor for type 2 diabetes (T2D) and associated metabolic complications, and weight loss has been shown to improve glycemic control and decrease morbidity and mortality in T2D patients. Weight-loss strategies using dietary interventions produce a significant decrease in diabetes-related metabolic disturbance. We have previously reported that the supplementation of low molecular chitosan oligosaccharide (GO2KA1) significantly inhibited blood glucose levels in both animals and humans. However, the effect of GO2KA1 on obesity still remains unclear. The aim of the study was to evaluate the anti-obesity effect of GO2KA1 on lipid accumulation and adipogenic gene expression using 3T3-L1 adipocytes in vitro and plasma lipid profiles using a Sprague-Dawley (SD) rat model. Murine 3T3-L1 preadipocytes were stimulated to differentiate under the adipogenic stimulation in the presence and absence of varying concentrations of GO2KA1. Adipocyte differentiation was confirmed by Oil Red O staining of lipids and the expression of adipogenic gene expression. Compared to control group, the cells treated with GO2KA1 significantly decreased in intracellular lipid accumulation with concomitant decreases in the expression of key transcription factors, peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT/enhancer-binding protein alpha (CEBP/α). Consistently, the mRNA expression of downstream adipogenic target genes such as fatty acid binding protein 4 (FABP4), fatty acid synthase (FAS), were significantly lower in the GO2KA1-treated group than in the control group. In vivo, male SD rats were fed a high fat diet (HFD) for 6 weeks to induced obesity, followed by oral administration of GO2KA1 at 0.1 g/kg/body weight or vehicle control in HFD. We assessed body weight, food intake, plasma lipids, levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) for liver function, and serum level of adiponectin, a marker for obesity-mediated metabolic syndrome. Compared to control group GO2KA1 significantly suppressed body weight gain (185.8 ± 8.8 g vs. 211.6 ± 20.1 g, p < 0.05) with no significant difference in food intake. The serum total cholesterol, triglyceride, and low-density lipoprotein (LDL) levels were significantly lower in the GO2KA1-treated group than in the control group, whereas the high-density lipoprotein (HDL) level was higher in the GO2KA1 group. The GO2KA1-treated group also showed a significant reduction in ALT and AST levels compared to the control. Moreover, serum adiponectin levels were significantly 1.5-folder higher than the control group. These in vivo and in vitro findings suggest that dietary supplementation of GO2KA1 may prevent diet-induced weight gain and the anti-obesity effect is mediated in part by inhibiting adipogenesis and increasing adiponectin level.

whISOBAXTM Inhibits Bacterial Pathogenesis and Enhances the Effect of Antibiotics.

As bacteria are becoming more resistant to commonly used antibiotics, alternative therapies are being sought. whISOBAX (WH) is a witch hazel extract that is highly stable (tested up to 2 months in 37 °C) and contains a high phenolic content, where 75% of it is hamamelitannin and traces of gallic acid. Phenolic compounds like gallic acid are known to inhibit bacterial growth, while hamamelitannin is known to inhibit staphylococcal pathogenesis (biofilm formation and toxin production). WH was tested in vitro for its antibacterial activity against clinically relevant Gram-positive and Gram-negative bacteria, and its synergy with antibiotics determined using checkerboard assays followed by isobologram analysis. WH was also tested for its ability to suppress staphylococcal pathogenesis, which is the cause of a myriad of resistant infections. Here we show that WH inhibits the growth of all bacteria tested, with variable efficacy levels. The most WH-sensitive bacteria tested were Staphylococcus epidermidis, Staphylococcus aureus, Enterococcus faecium and Enterococcus faecalis, followed by Acinetobacter baumannii, Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa, Streptococcus agalactiae and Streptococcus pneumoniae. Furthermore, WH was shown on S. aureus to be synergistic to linezolid and chloramphenicol and cumulative to vancomycin and amikacin. The effect of WH was tested on staphylococcal pathogenesis and shown here to inhibit biofilm formation (tested on S. epidermidis) and toxin production (tested on S. aureus Enterotoxin A (SEA)). Toxin inhibition was also evident in the presence of subinhibitory concentrations of ciprofloxacin that induces pathogenesis. Put together, our study indicates that WH is very effective in inhibiting the growth of multiple types of bacteria, is synergistic to antibiotics, and is also effective against staphylococcal pathogenesis, often the cause of persistent infections. Our study thus suggests the benefits of using WH to combat various types of bacterial infections, especially those that involve resistant persistent bacterial pathogens.

Witch Hazel Significantly Improves the Efficacy of Commercially Available Teat Dips.

Bovine intramammary infections (IMIs) are the main cause of economic loss in milk production. Antibiotics are often ineffective in treating infections due to antimicrobial resistance and the formation of bacterial biofilms that enhance bacterial survival and persistence. Teat dips containing germicides are recommended to prevent new IMIs and improve udder health and milk quality. IMIs are often caused by staphylococci, which are Gram-positive bacteria that become pathogenic by forming biofilms and producing toxins. As a model for a teat dip (DIP), the BacStop iodine-based teat dip (DIP) was used. Witch hazel extract (whISOBAX (WH)) was tested because it contains a high concentration of the anti-biofilm/anti-toxin phenolic compound hamamelitannin. We found that the minimal inhibitory or bactericidal concentrations of DIP against planktonic S. epidermidis cells increased up to 160fold in the presence of WH, and that DIP was 10-fold less effective against biofilm cells. While both DIP and WH are effective in inhibiting the growth of S. aureus, only WH inhibits toxin production (tested for enterotoxin-A). Importantly, WH also significantly enhances the antibacterial effect of DIP against Gram-negative bacteria that can cause IMIs, like Escherichia coli and Pseudomonas aeruginosa. Put together, these results suggest that the antibacterial activity of DIP combined with WH is significantly higher, and thus have potential in eradicating bacterial infections, both in acute (planktonic-associated) and in chronic (biofilm-associated) conditions.

In-Vitro Inhibition of Staphylococcal Pathogenesis by Witch-Hazel and Green Tea Extracts.

whISOBAX (WH), an extract of the witch-hazel plant that is native to the Northeast coast of the United States, contains significant amounts of a phenolic compound, Hamamelitannin (HAMA). Green tea (GT) is a widely consumed plant that contains various catechins. Both plants have been associated with antimicrobial effects. In this study we test the effects of these two plant extracts on the pathogenesis of staphylococci, and evaluate their effects on bacterial growth, biofilm formation, and toxin production. Our observations show that both extracts have antimicrobial effects against both strains of S. aureus and S. epidermidis tested, and that this inhibitory effect is synergistic. Also, we confirmed that this inhibitory effect does not depend on HAMA, but rather on other phenolic compounds present in WH and GT. In terms of biofilm inhibition, only WH exhibited an effect and the observed anti-biofilm effect was HAMA-depended. Finally, among the tested extracts, only WH exhibited an effect against Staphylococcal Enterotoxin A (SEA) production and this effect correlated to the HAMA present in WH. Our results suggest that GT and WH in combination can enhance the antimicrobial effects against staphylococci. However, only WH can control biofilm development and SEA production, due to the presence of HAMA. This study provides the initial rationale for the development of natural antimicrobials, to protect from staphylococcal colonization, infection, or contamination.

The Postprandial Anti-Hyperglycemic Effect of Pyridoxine and Its Derivatives Using In Vitro and In Vivo Animal Models.

In the current study, we investigated the inhibitory activity of pyridoxine, pyridoxal, and pyridoxamine, against various digestive enzymes such as α-glucosidases, sucrase, maltase, and glucoamylase. Inhibition of these enzymes involved in the absorption of disaccharide can improve post-prandial hyperglycemia due to a carbohydrate-based diet. Pyridoxal (4.14 mg/mL of IC50) had the highest rat intestinal α-glucosidase inhibitory activity, followed by pyridoxamine and pyridoxine (4.85 and 5.02 mg/mL of IC50, respectively). Pyridoxal demonstrated superior inhibition against maltase (0.38 mg/mL IC50) and glucoamylase (0.27 mg/mLIC50). In addition, pyridoxal showed significant higher α-amylase inhibitory activity (10.87 mg/mL of IC50) than that of pyridoxine (23.18 mg/mL of IC50). This indicates that pyridoxal can also inhibit starch hydrolyzing by pancreatic α-amylase in small intestine. Based on these in vitro results, the deeper evaluation of the anti-hyperglycemic potential of pyridoxine and its derivatives using Sprague-Dawley (SD) rat models, was initiated. The post-prandial blood glucose levels were tested two hours after sucrose/starch administration, with and without pyridoxine and its derivatives. In the animal trial, pyridoxal (p < 0.05) had a significantly reduction to the postprandial glucose levels, when compared to the control. The maximum blood glucose levels (Cmax) of pyridoxal administration group were decreased by about 18% (from 199.52 ± 22.93 to 164.10 ± 10.27, p < 0.05) and 19% (from 216.92 ± 12.46 to 175.36 ± 10.84, p < 0.05) in sucrose and starch loading tests, respectively, when compared to the control in pharmacodynamics study. The pyridoxal administration significantly decreased the minimum, maximum, and mean level of post-prandial blood glucose at 0.5 h after meals. These results indicate that water-soluble vitamin pyridoxine and its derivatives can decrease blood glucose level via the inhibition of carbohydrate-hydrolyzing and absorption-linked enzymes. Therefore, pyridoxal may have the potential to be used as a food ingredient for the prevention of prediabetes progression to type 2 diabetes.

Evaluation of Phenolic Phytochemical Enriched Commercial Plant Extracts on the In Vitro Inhibition of α-Glucosidase.

Green tea (GT), cranberry (CR), and tart cherry extracts were evaluated for their ability to inhibit yeast α-glucosidase, relevant to glucose uptake. The total phenolic content (TPC), antioxidant activity, and in vitro inhibitory activity of yeast α-glucosidase were examined for the extracts in the present study. GT had higher TPC and antioxidant activity, but CR demonstrated a greater α-glucosidase inhibitory activity, on phenolic basis. CR was fractionated using LH-20 column chromatography into two fractions: 30% methanol (CME) and 70% acetone (CAE). TPC, antioxidant activity, and yeast α-glucosidase inhibitory activity were determined for the fractions. CAE had a greater TPC and antioxidant activity than CME, but the two fractions had a synergistic effect when inhibiting yeast α-glucosidase. Our findings suggest that CR has the greatest potential to possibly manage post-prandial blood glucose levels via the inhibition of α-glucosidase, and that the effect is through synergistic activity of the extract's phenolic compounds.

Antidiabetic effect of chitosan oligosaccharide (GO2KA1) is mediated via inhibition of intestinal alpha-glucosidase and glucose transporters and PPARγ expression.

We have previously reported that administration of low molecular weight chitosan oligosaccharide (GO2KA1) significantly suppressed postprandial blood glucose rise with increased plasma adiponectin and HbA1c levels in animals and humans. However, the cellular mechanisms whereby GO2KA1 exerts antihyperglycemic effects still remain to be determined. Using intestinal Caco-2 cells and 3T3-L1 cells, here we show that GO2KA1 has dual modes of antidiabetic action by (1) inhibiting intestinal α-glucosidase as well as glucose transporters SGLT1 and GLUT2 that were distinct from the acarbose effect; (2) enhancing adipocyte differentiation, PPARγ expression and its target genes, such as FABP4, adiponectin, and GLUT4, whereas the effects were abolished by co-treatment with BADGE, a PPARγ antagonist. Moreover, GO2KA1 significantly increased glucose uptake, which was reduced in the presence of BADGE. Our data show that GO2KA1 may prevent hyperglycemia by inhibiting intestinal glucose digestion and transport and also enhance glucose uptake, at least in part, by upregulating adiponectin expression through PPARγ in adipocytes. These findings may provide potential molecular modes of action for the antidiabetic effects of chitosan oligosaccharide observed in clinical and animal studies. © 2016 BioFactors, 43(1):90-99, 2017.

In vitro and in vivo reduction of post-prandial blood glucose levels by ethyl alcohol and water Zingiber mioga extracts through the inhibition of carbohydrate hydrolyzing enzymes.

BACKGROUND: Type 2 diabetes is a serious problem for developed and developing countries. Prevention of prediabetes progression to type 2 diabetes with the use of natural products appears to be a cost-effective solution. Zingiber mioga has been used as a traditional food in Asia. Recent research has reported the potential health benefits of Zingiber mioga, but the blood glucose reducing effect has not been yet evaluated. METHODS: In this study Zingiber mioga extracts (water and ethanol) were investigated for their anti-hyperglycemic and antioxidant potential using both in vitro and animal models. The in vitro study evaluated the total phenolic content, the oxygen radical absorbance capacity (ORAC) and the inhibitory effect against carbohydrate hydrolyzing enzymes (porcine pancreatic α-amylase and rat intestinal sucrase and maltase) of both Zingiber mioga extracts. Also, the extracts were evaluated for their in vivo post-prandial blood glucose reducing effect using SD rat and db/db mice models. RESULTS: Our findings suggest that the ethanol extract of Zingiber mioga (ZME) exhibited the higher sucrase and maltase inhibitory activity (IC50, 3.50 and 3.13 mg/mL) and moderate α-amylase inhibitory activity (IC50, >10 mg/mL). Additionally, ZME exhibited potent peroxyl radical scavenging linked antioxidant activity (0.53/TE 1 µM). The in vivo study using SD rat and db/db mice models also showed that ZME reduces postprandial increases of blood glucose level after an oral administration of sucrose by possibly acting as an intestinal α-glucosidase inhibitor (ZME 0.1 g/kg 55.61 ± 13.24 mg/dL) CONCLUSION: The results indicate that Zingiber mioga extracts exhibited significant in vitro α-glucosidase inhibition and antioxidant activity. Additionally, the tested extracts demonstrated in vivo anti-hyperglycemic effects using SD rat and db/db mice models. Our findings provide a strong rationale for the further evaluation of Zingiber mioga for the potential to contribute as a useful dietary strategy to manage postprandial hyperglycemia.

Effect of supplementation of low-molecular-weight chitosan oligosaccharide, GO2KA1, on postprandial blood glucose levels in healthy individuals following bread consumption.

The effect of chitosan oligosaccharide (GO2KA1) administration on postprandial blood glucose levels of subjects with normal blood glucose levels was evaluated following bread consumption. Postprandial blood glucose levels were determined for 2 h after bread ingestion with or without 500 mg of GO2KA1. GO2KA1 significantly lowered the mean, maximum, and minimum levels of postprandial blood glucose at 30 min after the meal. Postprandial blood glucose levels were decreased by about 25% (from 155.11±13.06 to 138.50±13.59, p<0.01) at 30 min when compared to control. Furthermore, we observed that the area under the concentration-time curve (AUCt) was decreased by about 6% (from 255.46±15.43 to 240.15±14.22, p<0.05) and the peak concentration of blood glucose (C max) was decreased by about 11% (from 157.94±10.90 to 140.61±12.52, p<0.01) when compared to control. However, postprandial the time to reach C max (Tmax) levels were the same as those found in control. Our findings suggest that GO2KA1 limits the increase in postprandial blood glucose levels following bread consumption.

Effect of Black Tea and Black Tea Pomace Polyphenols on α-Glucosidase and α-Amylase Inhibition, Relevant to Type 2 Diabetes Prevention.

This study evaluates the potential mechanism of action and bioactivity of black tea and black tea pomace for type 2 diabetes prevention via inhibition of carbohydrate hydrolyzing enzymes. Black tea leaves were extracted in hot water and black tea pomace was extracted in 70% acetone. The phenolic content of the water extract (WBT) and pomace acetone extracts (AOBT) were 5.77 and 8.9 mg/mL, respectively, both based on the same concentration of solid tea in the extract. The water extract was subjected to C18 extraction and the resulting hydrophobic fraction (HBBT) was further subjected to LH-20 extraction to recover a low molecular weight phenolic enriched fraction (LMW) and a high molecular weight enriched fraction (HMW). The phenolic content of the LMW and HMW fraction were 1.42 and 2.66 mg/mL, respectively. Among water extracts the HMW fraction was most bioactive against α-glucosidase (IC50 = 8.97 µg/mL) followed by HBBT fraction (IC50 = 14.83 µg/mL). However, the HBBT fraction was the most bioactive fraction against α-amylase (IC50 = 0.049 mg/mL). The black tea pomace (AOBT) had significant α-glucosidase inhibitory activity (IC50 = 14.72 µg/mL) but lower α-amylase inhibitory activity (IC50 = 0.21 mg/mL). The phenolic profiles for LMW and HMW fractions were evaluated using HPLC and the differences between the two profiles were identified. Further research is underway to identify and evaluate the phenolic compounds that are present in the HMW fraction. Our findings suggest that black tea and black tea pomace has potential for carbohydrate hydrolyzing enzyme inhibition and this activity depends on high molecular weight phenolic compounds.

Effect of long-term supplementation of low molecular weight chitosan oligosaccharide (GO2KA1) on fasting blood glucose and HbA1c in db/db mice model and elucidation of mechanism of action.

BACKGROUND: Type 2 diabetes is a serious problem for developed countries. Prevention of prediabetes progression to type 2 diabetes with the use of natural products appears to a cost-effective solution. Previously we showed that enzymatically digested low molecular weight chitosan-oligosaccharide with molecular weight (MW) below 1,000 Da (GO2KA1) has potential for hyperglycemia management. METHODS: In this study we evaluated the effect of long-term supplementation of GO2KA1 on hyperglycemia using a db/db mice model. Additionally, we evaluated the effect of GO2KA1 on sucrase and glucoamylase activities and expression, using the same db/db mice model. RESULTS: After 42 days we observed that GO2KA1 supplementation reduced both the blood glucose level and HbA1c in a similar manner with a known anti-diabetic drug, acarbose. When the sucrase and glucoamylase activities of GO2KA1 and control mice were evaluated using enzymatic assay, we observed that GO2KA1 significantly inhibited sucrase in all 3 parts of the intestine, while glucoamylase activity was significantly reduced only in the middle and lower part. When the sucrase-isomaltase (SI) complex expression on mRNA level was evaluated, we observed that GO2KA1 had minimal inhibitory effect on the upper part, more pronounced inhibitory effect on the middle part, while the highest inhibition was observed on the lower part. Our findings suggest that long-term GO2KA1 supplementation in db/db mice results to significant blood glucose and HbA1c reduction, to levels similar with those of acarbose. Furthermore, our findings confirm previous in vitro observations that GO2KA1 has inhibitory effect on carbohydrate hydrolysis enzymes, namely sucrase, maltase and SI complex. CONCLUSIONS: Results from this study provide a strong rationale for the use of GO2KA1 for type 2 diabetes prevention, via inhibition of carbohydrate hydrolysis enzymes. Based on the findings of this animal trial, clinical trials will be designed and pursued.

Molecular weight dependent glucose lowering effect of low molecular weight Chitosan Oligosaccharide (GO2KA1) on postprandial blood glucose level in SD rats model.

This research investigated the effect of enzymatically digested low molecular weight (MW) chitosan oligosaccharide on type 2 diabetes prevention. Three different chitosan oligosaccharide samples with varying MW were evaluated in vitro for inhibition of rat small intestinal α-glucosidase and porcine pancreatic α-amylase (GO2KA1; <1000 Da, GO2KA2; 1000-10,000 Da, GO2KA3; MW > 10,000 Da). The in vitro results showed that all tested samples had similar rat α-glucosidase inhibitory and porcine α-amylase inhibitory activity. Based on these observations, we decided to further investigate the effect of all three samples at a dose of 0.1 g/kg, on reducing postprandial blood glucose levels in Sprague-Dawley (SD) rat model after sucrose loading test. In the animal trial, all tested samples had postprandial blood glucose reduction effect, when compared to control, however GO2KA1 supplementation had the strongest effect. The glucose peak (Cmax) for GO2KA1 and control was 152 mg/dL and 193 mg/dL, respectively. The area under the blood glucose-time curve (AUC) for GO2KA1 and control was 262 h mg/dL and 305 h mg/dL, respectively. Furthermore, the time of peak plasma concentration of blood glucose (Tmax) for GO2KA1 was significantly delayed (0.9 h) compared to control (0.5 h). These results suggest that GO2KA1 could have a beneficial effect for blood glucose management relevant to diabetes prevention in normal and pre-diabetic individuals. The suggested mechanism of action is via inhibition of the carbohydrate hydrolysis enzyme α-glucosidase and since GO2KA1 (MW < 1000 Da) had higher in vivo effect, we hypothesize that it is more readily absorbed and might exert further biological effect once it is absorbed in the blood stream, relevant to blood glucose management.

Antimicrobial activity of an Amazon medicinal plant (Chancapiedra) (Phyllanthus niruri L.) against Helicobacter pylori and lactic acid bacteria.

The potential of water extracts of the Amazon medicinal plant Chancapiedra (Phyllanthus niruri L.) from Ecuador and Peru for antimicrobial activity against Helicobacter pylori and different strains of lactic acid bacteria such as Lactobacillus acidophilus, Lactobacillus casei and Lactobacillus plantarum was investigated. H. pylori was inhibited by both water extracts in a dose dependent manner, whereas lactic acid bacterial growth was not affected. Both extracts contained ellagic acid and hydroxycinnamic acid derivatives and exhibited high free radical scavenging linked-antioxidant activities (89%). However, gallic acid was detected only in the Ecuadorian extract. Preliminary studies on the mode of action of Chancapiedra against H. pylori revealed that inhibition may not involve proline dehydrogenase-based oxidative phosphorylation inhibition associated with simple mono-phenolics and could involve ellagitannins or other non-phenolic compounds through a yet unknown mechanism. This study provides evidence about the potential of Chancapiedra for H. pylori inhibition without affecting beneficial lactic acid bacteria.

Seasonal variation of phenolic antioxidant-mediated α-glucosidase inhibition of Ascophyllum nodosum.

Ascophyllum nodosum is a brown seaweed that grows abundantly in the US Northeast coastal region. This study examined the seasonal variation of A. nodosum in phenolic contents and subsequent antioxidant, α-glucosidase and α-amylase inhibitory activities. A. nodosum was harvested monthly and extracted in hot water and the resulting extracts were spray-dried. The results indicate a clear seasonal variation in terms of phenolic content, with June and July being the highest (36.4 and 37 mg/g, respectively) and May the lowest (21.8 mg/g). The antioxidant activities, in terms of 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging activity, correlated with the phenolic contents observed (r = 0.81), with the month of July being the highest (58%) and April the lowest (26%). Similarly in terms of Trolox equivalent, July had the highest activity (15.53 µM) and April and May the lowest (8.40 and 8.27 µM, respectively). α-glucosidase inhibitory activity exhibited a pattern similar to the phenolic contents observed with July having the highest inhibitory activity (IC(70) 2.23 µg) and April the lowest (IC(70) 26.13 µg), resulting in an inverse correlation between IC(70) values and total phenolic content (r = -0.89). Such seasonal variation is believed to be caused by temperature-related stress considering that A. nodosum is a cold water species.

In vitro and in vivo antihyperglycemic effect of 2 amadori rearrangement compounds, arginyl-fructose and arginyl-fructosyl-glucose.

During the heat processing of raw ginseng to produce red ginseng, amino acid derivatives such as arginyl-fructose (AF) and arginyl-fructosyl-glucose (AFG) are formed at high levels, through amadori rearrangement, the early step of Maillard reaction, from arginine and glucose or maltose, respectively. However, very limited information is available about the effect of the structural difference between AF and AFG on various biological activities. This is the first report of the mode of action and effect of AF and AFG on the type 2 diabetes management related inhibition of postprandial hyperglycemia in vitro and in animal model. In our previous study, standards AF and AFG were chemically synthesized and in this study their inhibitory activities against rat intestinal α-glucosidases and porcine pancreatic α-amylase were investigated in vitro. The IC(50) value of the in vitro inhibitory activity of AF and AFG on rat intestinal sucrase was high and in similar levels (6.40 and 6.20 mM, respectively). Additionally, a mild pancreatic α-amylase inhibitory activity was observed, with IC(50) values 36.30 and 37.60 mM for AF and AFG, respectively. The effect of AF and AFG on the postprandial blood glucose increase after meal was investigated in Sprague Dawley rats fed on starch or sucrose meals. Both amadori compounds significantly reduced the postprandial blood glucose levels after starch or sucrose loading. These results indicate that AF and AFG, Maillard reaction products, may have antidiabetic effect by suppressing carbohydrate absorption in the gastrointestinal level, and thereby reducing the postprandial increase of blood glucose.