The synergistic effect of Cordyceps sinensis and Gymnema inodorum (Lour.) Decne. extracts on amelioration of glucose intolerance and dyslipidemia in high-fat diet-fed mice.

Cordyceps sinensis (C. sinensis) and Gymnema inodorum (Lour.) Decne. (G. inodorum) have been widely used for treating various illnesses. The study focused on exploring the effects of C. sinensis extract (CSE), G. inodorum extract (GIE), using alone and combined (COM) on ameliorating glucose intolerance, dyslipidemia, and obesity in mice fed with a high-fat diet (HFD). The results revealed that the oral glucose tolerance test (OGTT), total cholesterol (TC), triglycerides (TG), and LDL-cholesterol (LDL) exhibited a significant decrease in all groups treated with CSE, GIE, and COM compared to the control (p < 0.05). Obviously, CSE plus GIE exhibited a synergistic effect on amelioration of OGTT, TC, TG, and LDL, which is also the first report. Furthermore, the extracts showed no toxicity in the mice's vital organs. These results suggest that CSE, GIE, and their combined could have the potential as complementary therapeutic approaches for managing hyperglycaemia and dyslipidemia.

Free radical scavenging and anti-isolated human LDL oxidation activities of Butea superba Roxb. extract.

BACKGROUND: Butea superba Roxb. (B. superba), is an herbal plant traditionally used for rejuvenation. Additionally, there have been reports on its antioxidant properties. Low-density lipoproteins (LDL) oxidation is the leading cause of cardiovascular diseases (CVDs). Natural products with antioxidant properties have the potential to inhibit LDL oxidation. However, no work has been done about the anti-isolated human LDL oxidation of B. superba extract (BSE). This study aimed to investigate the antioxidant potential of BSE and its ability to prevent isolated human (LDL) oxidation induced by free radical agents. METHODS: The antioxidant properties were investigated by antioxidant assays, including 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2-azinobis-(3-ethylbenzothiazoline)-6-sulfonic acid (ABTS), ferric reducing ability power (FRAP), nitric oxide (NO) and peroxynitrite scavenging assay. More so, anti-isolated human LDL oxidation activities were evaluated by 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) and 3-morpholinosydnonimine hydrochloride (SIN-1) induced LDL oxidation assay. RESULTS: BSE exhibited a significant (p < 0.05) antioxidant activity in all the test systems, demonstrating its potential as a potent free radical scavenger. It displayed scavenging effects on DPPH (p < 0.05; IC50 = 487.67 ± 21.94 µg/ml), ABTS (p < 0.05; IC50 = 30.83 ± 1.29 µg/ml). Furthermore, it generated significantly (p < 0.05) increased antioxidant capacity in a dose-dependent manner in FRAP assay and exhibited significantly (p < 0.01) higher percent NO scavenging activity than gallic acid. Besides, BSE at 62.5 µg/ml exhibited a considerable percent peroxynitrite scavenging of 71.40 ± 6.59% after a 2 h period. Moreover, BSE demonstrated anti-isolated human LDL oxidation activity induced by AAPH and SIN-1 (p < 0.05) and revealed scavenging activity similar to ascorbic acid (p > 0.05). Identifying the main constituents of BSE revealed the presence of genistein, daidzein, and biochanin A through Liquid Chromatography-Mass Spectrometer/Mass Spectrometer (LC-MS/MS) analysis. CONCLUSION: This is the first report that the presence of isoflavones in BSE could play an important role in its antioxidation and isolated human LDL oxidation scavenging properties. These findings suggest the potential for developing antioxidant herbal supplements. However, further studies must be investigated, including efficacious and safe human dosages.

Butea superba Roxb. Extract Ameliorates Scopolamine-Induced Cognitive and Memory Impairment in Aged Male Rats.

Butea superba Roxb. (B. superba) is a herb that has been used for rejuvenation, to improve sexual performance, or to prevent erectile dysfunction function. Alzheimer's disease (AD) is a chronic neurodegenerative disorder that is the main cause of progressive dementia. This study aimed to investigate the amelioration for cognitive and memory dysfunction of B. superba ethanolic extract (BSE), a possible mechanism of action, and its toxicity. The results from the Y-maze test, novel object recognition test, and passive avoidance test exhibited that the administration of BSE at 50 mg/kg (BSL) and 200 mg/kg (BSH) could ameliorate scopolamine-induced cognitive impairment in all behavior testing. Moreover, BSE could prevent the cognitive deficit in a dose-dependent manner in a passive avoidance test. Furthermore, BSE inhibited acetylcholinesterase's (AChE) ex vivo activity in the cerebral cortex and hippocampus. Also, the in vitro and ex vivo antioxidative effects of BSE revealed that BSE had free radical scavenging activities in both DPPH and FRAP assay. Furthermore, male rats treated with BSE at 200 mg/kg/day for two weeks could significantly increase serum testosterone compared with control (P < 0.05). The GC-MS analysis and previous studies revealed that BSE contained propanoic acid, 3,3'-thiobis-, didodecyl ester, oleic acid, gamma-sitosterol, and stigmasterol which may play an important role in cognitive and memory impairment prevention. The toxicity test of BSE in rats at 50 and 200 mg/kg/day for two weeks showed that relative organ weight, serum creatinine, ALT, ALP, and CBC levels of both treated groups were not significantly different compared to the CON (P > 0.05). These results suggest that BSE may not be toxic to the vital organ and blood. In conclusion, BSE has the potential to be developed as a health supplement product or medicine for AD prevention and treatment.

The Effect of Pluchea indica (L.) Less. Tea on Blood Glucose and Lipid Profile in People with Prediabetes: A Randomized Clinical Trial.

Objective: Diabetes mellitus and dyslipidemia are currently increasing dramatically, and conventional medicine in the treatment of them has limited efficacies and serious adverse effects. Pluchea indica (L.) Less. tea (PIT) is widely consumed as a health-promoting drink in Southeast Asia. This study aimed to investigate whether P. indica tea has antidyslipidemic and antihyperglycemic effects and toxicity in humans. Design: A randomized clinical trial. Setting/Location: Nakhonratchasima, Thailand. Participants: Forty-five participants with prediabetes. Interventions: Participants were randomized to receive placebo tea, 1.5 g of PIT, and 1.5 g Camellia sinensis tea (green tea, CST) once daily for 12 weeks. Outcome Measurements: The oral glucose tolerance test (OGTT), total cholesterol, triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), blood urea nitrogen (BUN), creatinine, alanine transaminase (ALT), alkaline phosphatase (ALP), and complete blood count (CBC) before and after treatment were investigated. Results: The results showed that PIT significantly ameliorated hyperglycemia and significantly lower serum TG (109.22 ± 5.21 mg/dL) and LDL-C (122.20 ± 3.67 mg/dL) than placebo (145.56 ± 8.18 and 142.07 ± 8.58 mg/dL, respectively) (p < 0.05). Moreover, PIT exhibited serum TG (109.22 ± 5.21 mg/dL) significantly lower than CST (124.38 ± 4.70 mg/dL) (p < 0.05). In addition, the serum HDL-C of PIT (57.56 ± 3.05 mg/dL) was significantly higher than the placebo (46.44 ± 2.47 mg/dL) (p < 0.05). Furthermore, the toxicity testing showed that no significant difference in BUN, creatinine, ALT, ALP, and CBC of PIT-treated group compared with the placebo (p > 0.05). Conclusions: These results suggest that PIT may ameliorate hyperglycemia and dyslipidemia in prediabetes people. It may not be toxic to the kidney, liver, and blood. So, PIT has the potential to develop to be a health-promoting tea or herbal medicine for hyperglycemia and dyslipidemia prevention.

The Free Radical Scavenging and Anti-Isolated Human LDL Oxidation Activities of Pluchea indica (L.) Less. Tea Compared to Green Tea (Camellia sinensis).

Tea is one of the most popular beverages in the world. Camellia sinensis tea (CST) or green tea is widely regarded as a potent antioxidant. In Thailand, Pluchea indica (L.) Less. tea (PIT) has been commercially available as a health-promoting drink. This study focused on free radical scavenging activities of PIT, and its ability to protect isolated human low-density lipoproteins (LDL) from oxidation by chemical agents. A preliminary study to investigate the antioxidant nature of PIT was undertaken. These included common antioxidant assays involving 2,2-Diphenyl-1-picrylhydrazyl (DPPH), 2,2-azinobis-(3-ethylbenzothiazoline)-6-sulfonic acid (ABTS), hypochlorous acid (HOCl), and its potential to scavenge peroxynitrite. In separated experiments, isolated human LDL was challenged with either 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH), copper (Cu2+), or 3-Morpholinosydnonimine hydrochloride (SIN-1) to induce LDL oxidation. PIT exhibited antioxidant activity in all test systems and performed significantly better than CST in both DPPH (P < 0.05; IC50PIT = 245.85 ± 15.83 and CST = 315.41 ± 24.18 µg/ml) and peroxynitrite scavenging assays. PIT at 75 µg/ml almost fully prevented the peroxynitrite over a 5 h period. Moreover, it displayed similar properties to CST during the antioxidation of isolated human LDL using AAPH, Cu2+, SIN-1, and hypochlorous acid scavenging assays. However, it revealed a significantly lower ABTS scavenging activity than CST (P < 0.05; IC50PIT = 30.47 ± 2.20 and CST = 21.59 ± 0.67 µg/ml). The main constituents of the PIT were identified using LC-MS/MS. It contained 4-O-caffeoylquinic acid (4-CQ), 5-O-caffeoylquinic acid (5-CQ), 3,4-O-dicaffeoylquinic acid (3,4-CQ), 3,5-O-dicaffeoylquinic acid (3,5-CQ), and 4,5-O-dicaffeoylquinic acid (4,5-CQ). In conclusion, caffeoyl derivatives in PIT could play an important role in potent antioxidant properties. So, it may be further developed to be antioxidant beverages for preventing atherosclerosis and cardiovascular diseases associated with oxidative stress.

Pluchea indica (L.) Less. Tea Ameliorates Hyperglycemia, Dyslipidemia, and Obesity in High Fat Diet-Fed Mice.

Pluchea indica (L.) Less. (P. indica) tea has been used for a health-promoting drink, especially in Southeast Asia. The effect of P. indica tea (PIT) on amelioration of hyperglycemia; dyslipidemia that was total cholesterol (TC), LDL-cholesterol (LDL-C), HDL-cholesterol (HDL-C), and triglyceride (TG); and obesity in high fat diet-induced (HFD) mice was investigated. Oral glucose tolerance test (OGTT) displayed that PIT at 400 and 600 mg/kg orally ameliorated hyperglycemia with a dose-dependent manner compared to the untreated group. Moreover, PIT at these dosages exhibited significantly lower TC, LDL-C, TG, and perigonadal fat weight in HFD treated mice compared to HFD mice (P < 0.05) with a dose-dependent manner. In contrast, HDL-C was higher than in the HFD group, but not a significant difference (P > 0.05). The PIT chemical analysis results demonstrated that PIT contained total phenolic content (TPC), total flavonoid content (TFC), 4-O-caffeoylquinic acid (4-CQ), 5-O-caffeoylquinic acid (5-CQ), 3,4-O-dicaffeoylquinic acid (3,4-CQ), 3,5-O-dicaffeoylquinic acid (3,5-CQ), 4,5-O-dicaffeoylquinic acid (4,5-CQ), beta-caryophyllene, and gamma-gurjunene that may play an important role in inhibiting hyperlipidemia and hyperglycemia. Also, histological analysis expressed that the mean area and amount of perigonadal fat adipocytes of PIT treated groups were significantly lower and higher than the HFD group (P < 0.05), respectively. The toxicity test of PIT at 600 mg/kg/day in mice showed that serum creatinine, alanine transaminase (ALT), alkaline phosphatase (ALP), and complete blood count (CBC) levels of HFD and PIT treated groups were not significantly different compared to the normal control diet group (NCD) (P > 0.05). These results suggest that PIT does not become toxic to the kidney, liver, and blood. In conclusion, PIT has the potential to develop into healthy food supplement or medicine for the prevention and treatment of hyperglycemic, hyperlipidemic, and obese patients.

The Effects of Cordyceps sinensis (Berk.) Sacc. and Gymnema inodorum (Lour.) Decne. Extracts on Adipogenesis and Lipase Activity In Vitro.

This study aimed to investigate the effects of Cordyceps sinensis extract (CSE) and Gymnema inodorum extract (GIE), used alone and combined, on antiadipogenesis in 3T3-L1 cells. Oil Red O staining was used to examine the effects of these extracts on inhibition of intracellular lipid accumulation in 3T3-L1 adipocytes and on lipid droplet morphology. Fourier transform-infrared (FTIR) microspectroscopy was used to examine biomolecular changes in 3T3-L1 adipocytes. The pancreatic lipase assay was used to evaluate the inhibitory effects of CSE and GIE on pancreatic lipase activity. Taken together, the results indicated that CSE, GIE, and their combination suppressed lipid accumulation. The FTIR microspectroscopy results indicated that CSE, GIE, and their combination had inhibitory effects on lipid accumulation in the adipocytes. Compared with the untreated adipocytes, the signal intensity and integrated areas of glycogen and other carbohydrates, the acyl chain of phospholipids, and the lipid/protein ratios of the CSE, GIE, alone, and combined treated adipocytes were significantly lower (p < 0.05). Combination treatment resulted in a synergistic effect on lipid accumulation reduction in the adipocytes. Principal component analysis of the biomolecular changes revealed six distinct clusters in the FTIR spectra of the sample cells. The pancreatic lipase assay results indicated that CSE and GIE inhibited the pancreatic lipase activity in a dose-dependent manner (mean ± standard error of the mean IC50 values, 2312.44 ± 176.55 µg mL-1 and 982.24 ± 44.40 µg mL-1, resp.). Our findings indicated that FTIR microspectroscopy has potential application for evaluation of the effectiveness of medicinal plants and for the development of infrared biochemical obesity markers useful for treating patients with obesity. These results suggested that use of CSE and GIE alone and in combination may be efficacious as a complementary therapy for hyperlipidemia and obesity management. However, clinical trials in animals and humans must first be completed.

The Effect of Pluchea indica (L.) Less. Tea on Adipogenesis in 3T3-L1 Adipocytes and Lipase Activity.

Obesity and hyperlipidemia are a major problem in the world. Pluchea indica (L.) Less. tea (PIT) is a beverage that has various indications. This study focused on the effect of the PIT on inhibiting adipogenesis of 3T3-L1 cells and pancreatic lipase enzyme activity. The viability of 3T3-L1 cells was not significantly decreased after exposure to 200 to 1000 µg mL-1 PIT compared to controls (p > 0.05). The PIT at 750 to 1000 µg mL-1 exhibited a significantly reduced lipid accumulation compared to the control (p < 0.05). The inhibitory effects of the PIT at 250 to 1000 µg mL-1 on lipase activity were significantly increased compared to control (p < 0.05). The FTIR results showed that the integrated areas of lipids, proteins, nucleic acids, glycogen, and carbohydrates of the PIT-treated 3T3-L1 adipocytes were significantly lower than the untreated 3T3-L1 adipocytes (p < 0.05). These findings may indicate that the PIT is not only capable of inhibiting lipids and carbohydrate accumulation in adipocytes but also has a potential to inhibit pancreatic lipase activity. So, the PIT may be further developed to the novel lipid-lowering herbal supplement for the management of overweight or obesity.

The synergy effect of daidzein and genistein isolated from Butea superba Roxb. on the reproductive system of male mice.

Butea superba Roxb. (BS) has been used in Thai men as an aphrodisiac, and prevent erectile dysfunction. Nevertheless, the active ingredients, dosages, have not been cleared. Hence, this study was to investigate the effect of compounds from the BS on the reproductive parameters of male mice. The results revealed that BS was extracted to afford biochanin A and genistein, which were first reported on BS, and daidzein. The mice were treated by daidzein and genistein alone and in combination. The results showed that the sperm number and motility, cholesterol and testosterone level of all isoflavones-treated groups were significantly higher than controls (p < 0.01). Obviously, daidzein plus genistein exhibited a synergistic effect, which is also the first report, and resulted in significantly displayed higher levels of these parameters compared to others. So, the synergistic activity of these isoflavones may be useful in improving libido, erectile capacity and assist infertility of poor spermatozoa in men.

Synergism and the mechanism of action of the combination of α-mangostin isolated from Garcinia mangostana L. and oxacillin against an oxacillin-resistant Staphylococcus saprophyticus.

BACKGROUND: Globally, staphylococci have developed resistance to many antibiotics. New approaches to chemotherapy are needed and one such approach could be to use plant derived actives with conventional antibiotics in a synergestic way. The purpose of this study was to isolate α-mangostin from the mangosteen (Garcinia mangostana L.; GML) and investigate antibacterial activity and mechanisms of action when used singly and when combined with oxacillin against oxacillin-resistant Staphylococcus saprophyticus (ORSS) strains. The isolated α-mangostin was confirmed by HPLC chromatogram and NMR spectroscopy. The minimum inhibitory concentration (MIC), checkerboard and killing curve were determined. The modes of action of these compounds were also investigated by enzyme assay, transmission electron microscopy (TEM), confocal microscopic images, and cytoplasmic membrane (CM) permeabilization studies. RESULTS: The MICs of isolated α-mangostin and oxacillin against these strains were 8 and 128 µg/ml, respectively. Checkerboard assays showed the synergistic activity of isolated α-mangostin (2 µg/ml) plus oxacillin (16 µg/ml) at a fractional inhibitory concentration index (FICI) of 0.37. The kill curve assay confirmed that the viability of oxacillin-resistant Staphylococcus saprophyticus DMST 27055 (ORSS-27055) was dramatically reduced after exposure to isolated α-mangostin (2 µg/ml) plus oxacillin (16 µg/ml). Enzyme assays demonstrated that isolated α-mangostin had an inhibitory activity against ß-lactamase in a dose-dependent manner. TEM results clearly showed that these ORSS-27055 cells treated with this combination caused peptidoglycan and cytoplasmic membrane damage, irregular cell shapes and average cell areas were significantly larger than the control. Clearly, confocal microscopic images confirmed that this combination caused considerable peptidoglycan damage and DNA leakage. In addition, the CM permeability of ORSS-27055 was also increased by this combination of actives. CONCLUSIONS: These findings provide evidence that isolated α-mangostin alone has not only some activity but also shows the synergistic activity with oxacillin against ORSS-27055. The chromone and isoprenyl structures could play a significant role in its action. This synergistic activity may involve three mechanisms of action. Firstly, potential effects of cytoplasmic membrane disruption and increases permeability. Secondly, inhibit ß-lactamase activity. Finally, also damage to the peptidoglycan structure. We proposes the potential to develop a novel adjunct phytopharmaceutical to oxacillin for the treatment of ORSS. Future studies require clinical trials to establish if the synergy reported can be translated to animals and humans.

Synergistic activity and mechanism of action of Stephania suberosa Forman extract and ampicillin combination against ampicillin-resistant Staphylococcus aureus.

BACKGROUND: Ampicillin-resistant S. aureus (ARSA) now poses a serious problem for hospitalized patients, and their care providers. Plant-derived antibacterial that can reverse the resistance to well-tried agents which have lost their original effectiveness are the research objectives of far reaching importance. To this aim, the present study investigated antibacterial and synergistic activities of Stephania suberosa extracts (SSE) against ARSA when used singly and in combination with ampicillin. RESULTS: The majority chemical compounds of SSE were alkaloid (526.27 ± 47.27 mg/1 g of dried extract). The Minimum inhibitory concentration (MICs) for ampicillin and SSE against all ARSA strains were >512 µg/ml and 4 mg/ml, respectively. Checkerboard assay revealed synergistic activity in the combination of ampicillin (0.15 µg/ml) and SSE (2 mg/ml) at fractional inhibitory concentration index (FICI) <0.5. The killing curve assay had confirmed that the viability of ARSA was dramatically reduced from 5 x 10(5) cfu/ml to 10(3) cfu/ml within 6 h after exposure to SSE (2 mg/ml) plus ampicillin (0.15 µg/ml) combination. Electron microscopic study clearly revealed that these ARSA cells treated with this combination caused marked morphological damage, peptidoglycan and cytoplasmic membrane damage, and average cell areas significant smaller than control. Obviously, Immunofluorescence staining and confocal microscopic images confirmed that the peptidoglycan of these cells were undoubtedly disrupted by this combination. Furthermore, the CM permeability of ARSA was also increased by this combination. Enzyme assay demonstrated that SSE had an inhibitory activity against ß-lactamase in concentrations manner. CONCLUSIONS: So, these findings provide evidence that SSE has the high potential to reverse bacterial resistance to originate traditional drug susceptibility of it and may relate to three modes of actions of SSE: (1) inhibits peptidoglycan synthesis, resulting in morphological damage, (2) inhibits ß-lactamases activity, and (3) increases CM permeability. It is widely recognized that many types of drugs are derived from alkaloids. So, this SSE offers the prominent potential to develop a novel adjunct phytopharmaceutical to ampicillin for the treatment of ARSA. Further active ingredients study, toxicity of it, and the synergistic effect on blood and tissue should be performed and confirmed in an animal test or in humans.