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Objective: The aim of this study was to investigate the anti-hyperglycemia effect and mechanism of aqueous extract from Gynostemma pentaphyllum (GP) leaves on STZ-induced diabetic rats. Methods: Diabetic rat models were established by intraperitoneal injection of streptozotocin (STZ, 50 mg/kg). A total of 21 successful SD male rats were randomly divided into model group (STZ). G. pentaphyllum leaves aqueous extract low dose group (GP•H2O-L, 100 mg/kg) and high dose group (GP•H2O-H, 500 mg/kg), another seven normal rats were taken as the control group. Blood samples were taken from the 2nd and 3rd weekends to detect plasma glucose and triglyceride concentrations; Real-time PCR was used to detect the expression of TNF-α and GLUT-4 mRNA in skeletal muscle; Western blotting was used to detect GLUT-4 total protein in skeletal muscle; The expression of GLUT-4 protein on skeletal muscle sarcolemma was observed by immunofluorescence staining. Results: The results showed that the food intake and water intake of the STZ group were significantly increased compared with the control group, while the body weight and skeletal muscle weight were obviously decreased; Plasma triglyceride and blood glucose concentrations and the expression of TNF-α mRNA in skeletal muscle were significantly increased, while the expression of GLUT-4 mRNA, GLUT-4 total protein and GLUT-4 protein in skeletal muscle sarcolemma was obviously decreased. Compared with the STZ group, the high-dose aqueous extract of G. pentaphyllum leaves significantly reduced the blood glucose of STZ-induced diabetic rats, and reversed the expression of TNF-α mRNA and GLUT-4 protein in skeletal muscle. Conclusion: The aqueous extract from G. pentaphyllum leaves could reduce hyperglycemia in STZ-induced diabetic rats, and its mechanism may be related to increasing the expression of GLUT-4 protein on skeletal muscle sarcolemma and inhibiting skeletal muscle inflammation.
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The present study attempts to study alcohol metabolizing and antioxidant properties of Gynostemma pentaphyllum(Thunb.) Makino distillate (GPD) and combination effects with Hovenia dulcis Thunb. extract (HDE) on these activities.The alcohol-metabolizing activity of GPD with/without HDE was determined by assessing alcohol dehydrogenase(ADH) and acetaldehyde dehydrogenase (ALDH) activities. To define the effect of GPD with/without HDE on alcoholmetabolism, antioxidant activities and total phenolic content of GPD with/without HD extract were evaluated using2-diphenyl-1-picrylhydrazyl free radical scavenging, ferrous chelating assays, and the Folin–Ciocalteu method.Cytotoxicity against human normal liver CHANG cells was also evaluated using the 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. GPD treatment alone or in combination with HDE significantly increased ADHand ALDH activities; combined treatment was most effective. Total phenolic and flavonoid contents were greater incombination than the level found in GPD alone. GPD revealed a synergistic antioxidant effect when combined withHDE. GPD and/or HDE had no antiproliferative activity against the normal liver cell line. These results suggest thatGPD-HDE combination is the possible natural resource for the management of alcohol-induced liver injury.
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Objective: The fingerprint of Gynostemma pentaphyllum was established with liquid chromatograph-mass spectrometer (LC-MS), and the main chromatographic peaks were preliminary identified, and combined with multi-variable analysis pattern recognition method to identify G. pentaphyllum from different origins. Methods: Inertsil ODS-SP (250 mm × 4.6 mm, 5 μm) C18 column was eluted with acetonitrile-0.1% formic acid as mobile phase gradient. The detection wavelength was 203 nm. The mass spectrometry equipped with electrospray ionization (ESI) was used as detector and operated under the negative ion mode. A total of 21 batches of G. pentaphyllum were analyzed by cluster analysis, principal component analysis, and corresponding analysis. Results: The fingerprints of G. pentaphyllum were established by LC-MS, and nine common peaks and 10 characteristic peaks were calibrated. According to the mass spectrometry information and literature comparison, 16 chromatographic peaks were qualitatively analyzed. Combined with multivariate analysis, G. pentaphyllum from different habitats was distinguished. It would establish the foundations for quality control and comprehensive evaluation of G. pentaphyllum. Conclusion: The LC-MS and pattern recognition analysis can be used to distinguish G. pentaphyllum from different regions and the targeting compounds of interest would be separated.
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AIM To establish a UFLC-QTRAP-MS/MS method for the simultaneous content determination of phenylalanine,isoleucine,methionine,valine,proline,tyrosine,aspartic acid,histidine,arginine,lysine and glutamic acid in Gynostemma pentaphyllum (Thunb.) Makino.METHODS The analysis of aqueous extract of G.pentaphyllum was performed on a Waters XBridge Amide column (2.1 mm × 100 mm,3.5 μm),with the mobile phase comprising of water-acetonitrile (containing 0.2％ formic acid) flowing at 0.6 mL/min in a gradient elution manner.RESULTS Eleven amino acids showed good linear relationships within their own ranges (r ＞0.998 9),whose average recoveries were 90.74％-103.05％ with the RSDs of 0.61％-5.00％.CONCLU-SION This accurate,stable and reproducible method can be used for the quality control of G.pentaphyllum.
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Objective To establish a high performance liquid chromatography(HPLC) method for the simultaneous determination of eight components in Gynostemma pentaphyllum(Thunb.) Makino (ginsenosides Rb1, Rb3, Rd, Re, F2, and gypenosides A, XLIX and XVII). Methods HPLC was performed on Agilent Eclipse XDB-C18(4.6 mm × 250 mm, 5 μm) chromatographic column with 0.3％ formic acid solution (A)- acetonitrile (B) as the mobile phase by gradient elution, flow rate was 1.5 mL·min-1, detection wavelength was 203 nm, and column temperature was set at 50 ℃. Results There was a good linear relationship between peak area and concentration of eight components (r ≥ 0.999) , the recovery was in the range of 97％ to 100％. Conclusion The established method is simple, rapid, accurate, reliable, and reproducible, and can be used for the quality control of Gynostemma pentaphyllum(Thunb.) Makino.
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Dammarane saponins, classified as tetracyclic triterpenoid saponins, are major components of Panax ginseng, P. notoginseng, P. quinquefolium, and Gynostemma pentaphyllum. The results of recent research have demonstrated that the dammarane saponins have a significant effect on regulating blood glucose, which provides the important value in the prevention and treatment of diabetes and its complications. In this study, we summarized the progress in the research of hypoglycemic effect of dammarane saponins in ginseng, American ginseng, notoginseng, and gynostemma pentaphyllum, providing theoretical foundation for further researching.
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Objective: To clone Gynostemma pentaphyllum squalene synthase gene (GpSS1) and analyze its sequence and expression pattern as well as its regulation in response to MeJA. Methods: Primers were designed based on the sequence of GpSS (GenBank accession numbers: FJ906799) and GpSS1 was cloned by using RT-PCR method. Physicochemical properties and transmembrane regions of the deduced GpSS1 protein were predicted via Protparam and Tmpred programs, respectively. Conserved domains involved in catalytic activity of SS enzyme were identified using MotifScan and multiple sequence alignment was achieved using the BioEdit software. Expression pattern of GpSS1 and its regulation by MeJA were analyzed by quantitative real-time RT-PCR. Results: GpSS1 contains an open reading frame of 1 254 bp encoding a putative protein of 417 amino acids. The protein has an aspartate-rich motif and an endoplasmic reticulum membrane anchoring region in addition to three conserved domains involved in catalytic activity of SS enzyme. The expression level of GpSS1 in young leaves was the highest, followed by that in old leaves, and the lowest was in rhizomes. The expression of GpSS1 was significantly upregulated in G. pentaphyllum leaves sprayed with different concentration of MeJA. The greatest upregulation of GpSS1 occurred in G. pentaphyllum leaves treated with 50 μmol/L MeJA. In both young and old leaves, the transcription of GpSS1 gradually increased and then decreased to varying degrees at 6-96 h after MeJA treatment. However, the expression level of GpSS1 was always higher in young leaves than that in old leaves. Conclusion: The cloning of GpSS1 and analysis on the expression regulated by MeJA will be helpful for further elucidating the function of GpSS1 and its mechanism regulated by MeJA, as well as for improving the quality and content of gypenosides.
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Objective: To study the chemical constituents of Gynostema pentaphyllum. Methods: The chemical constituents were isolated and purified by silica gel, polyamide, and Sephadex LH-20 chromatography. Their structures were elucidated by physicochemical properties and spectral analysis. Results: Seventeen compounds were isolated and identified as dodecanoic acid (1), β-sitosterol (2), 3, 3', 5-trihydroxy-4', 7-dimethoxyflavanone (3), 1-2-benzenediol (4), 3'-O-methyltaxifolin (5), quercetin (6), rhamnetin (7), α-spinasterol-3-O-β-D-glucopyranoside (8), 3, 4-dihydroxy benzoic acid (9), narcissoside (10), L-rhamnosemono-hydrate (11), malonic acid (12), β-ethoxy-rutinoside (13), rutin (14), ombuoside (15), ginsenoside Rb1 (16), and β-daucosterol (17). Conclusion: Compound 1, 3-5, 7, 9, 10, and 12 are obtained from G. pentaphyllum for the first time.
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Objective: To optimize the extracting process of total flavonoids from Gynostemma pentaphyllum. Methods: The total flavonoids were extracted with ethanol 95% after the cellulose treatment to G. pentaphyllum, and the effects of enzyme dosage, enzymatic treatment temperature, enzymatic treatment time, and pH value on the total flavonoids yield was studied with response surface method. Results: The optimal enzymatic extracting process of total flavonoids was as follows: enzyme dosage 120 U/mL, enzymatic temperature 49 °C, enzymatic time 150 min, and pH value 3.9, and the total flavonoids yield reached 0.357%. Conclusion: Cellulase is suitable for the total flavonoid extraction from G. pentaphyllum.
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Objective To construct a stably-performing and high-efficiency regeneration system of Gynostemma pentaphyllum and lay a foundation for the gene transformation of it.Methods The blades of five-leaf G.pentaphyllum were used as the explants and cultured in MS media with different portions of hormone to induce fascicled-bud,root and plantlet regeneration.Results The medium suitable for inducing the blade differentiation of G.pentaphyllum was MS+6-BA 1.0 mg/L and for the frequency of blade differentiation could reach 40%.The cultural medium MS+6-BA 1.0 mg/L was suitable for the sub-multiplication of fascicled-bud and the medium 1/2 MS for root inducement and the plantlet regeneration.Conclusion A stably-performing acceptor system of the direct differentiation for Agrobacterium tumefaciens mediated gene transformation of G.pentaphyllum blades is constructed.
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Object To explore the protoplast fusion of Dendrobium candidum Wall ex. Lindl. and Gynostemma pentaphyllum. (Thunb) Makino. Methods The two kinds of protoplasts were fused by PEG method, and then cultured in the modified liquid medium containing 2 mg/L BA and 1 mg/L NAA. Results High yield, viability, and pure mesophyll protoplasts were isolated from D. candidum and G. pentaphyllum. Conclusion The first cell division occurres within three days after fusion. And some of the cells are divided three times.