植物穿心莲营养成分的研究

对植物穿心莲的营养成分进行了分析,结果表明,穿心莲中含有多种营养成分,丰富的矿质元素和维生素以及β-胡萝卜素。穿心莲中至少含有17种氨基酸。旨在为开发利用植物资源提供科学依据。     

Antifertility effect of Andrographis paniculata (Nees) in male albino rat

Dry leaf powder of A. paniculata, when fed orally to male albino rats, at a dose level of 20 mg powder per day for 60 days, resulted in cessation of spermatogenesis, degenerative changes in the seminiferous tubules, regression of Leydig cells and regressive and/or degenerative changes in the epididymis, seminal vesicle, ventral prostate and coagulating gland. There was reduction in the weight and fluid content of the accessory glands. The treatment also resulted in accumulation of glycogen and cholesterol in the testis, and increased activities of lactate dehydrogenase in testis and alkaline phosphatase in testis and ventral prostate. The results suggest antispermatogenic and/or antiandrogenic effect of the plant.     

Proteomic Analysis of Salt-Responsive Proteins in the Leaves of Mangrove Kandelia candel during Short-Term Stress

Salt stress is a major abiotic stress that limits crop productivity in many regions of the world. A comparative proteomic approach to identify salt stress-responsive proteins and to understand the molecular mechanisms was carried out in the woody halophyte Kandelia candel. Four-leaf-old K. candel seedlings were exposed to 150 (control), 300, 450, and 600 mM NaCl for 3 days. Proteins extracted from the leaves of K. candel seedlings were separated by two-dimensional gel electrophoresis (2-DE). More than 900 protein spots were detected on each gel, and 53 differentially expressed protein spots were located with at least two-fold differences in abundance on 2-DE maps, of which 48 were identified by matrix-assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF-TOF/MS). The results showed that K. candel could withstand up to 450 mM NaCl stress by up-regulating proteins that are mainly involved in photosynthesis, respiration and energy metabolism, Na⁺ compartmentalization, protein folding and assembly, and signal transduction. Physiological data, including superoxide dismutase (SOD) and dehydroascorbate reductase (DHAR) activities, hydrogen peroxide (H₂O₂) and superoxide anion radicals (O₂ ⁻) contents, as well as Na⁺ content and K⁺/Na⁺ ratios all correlated well with our proteomic results. This study provides new global insights into woody halophyte salt stress responses. Identification of differentially expressed proteins promotes better understanding of the molecular basis for salt stress reduction in K. candel.     

Determination and variation of three active diterpenoids in Andrographis paniculata (Burm.f.) Nees

Simple and rapid methods have been developed for the extraction and simultaneous determination of the three active diterpenoids, andrographolide (AP1), 14-deoxy-11,12-didehydroandrographolide (AP3) and neoandrographolide (AP4), from various samples of Andrographis paniculata (Burm.f.) Nees. Methanol extracts from the dried leaves, stems and crude products were analysed by isocratic HPLC using a methanol and water mobile phase with monitoring at 220 nm. There was a large variation of the three active diterpenoids in different A. paniculata products obtained from Thai markets. The results indicated that the amounts of these active compounds consumed, based on the recommended daily doses, from materials obtained from the different suppliers will be different. In addition, the stability of these three active compounds was also examined in dry herbs stored at room temperature. The results showed that andrographolide was more stable than the others. In contrast, the content of 14-deoxy-11,12-didehydroandrographolide increased and the neoandrographolide content fluctuated during storage time. The combination of different levels of these compounds in the source materials and the changes during storage could have a significant effect on the efficacy of this traditional herbal medicine in clinical treatment.     

Hepatoprotective and antioxidant property of Andrographis paniculata (Nees) in BHC induced liver damage in mice

Andrographis paniculata (AP) treatment prevents BHC induced increase in the activities of enzymes y-Glutamyl transpeptidase, glutathione-S-transferase and lipid peroxidation. The activities of antioxidant enzymes like superoxide dismutase, catalase, glutathione peroxidase and the levels of glutathione were decreased following BHC effect. Administration of AP showed protective effects in the activity of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase as well the level of glutathione. The activity of lipid peroxidase was also decreased. The result indicate antioxidant and hepatoprotective action of A. paniculata.     

Proteomic Analysis of Specific Proteins in the Root of Salt-Tolerant Barley

Comparative two-dimensional electrophoresis showed six proteins, which were significantly produced in the root of salt-tolerant barley. These proteins were identified as stress/defense-related proteins that do not scavenge reactive oxygen species directly, suggesting that salt-tolerant barley develops not only an antioxidative system, but also physical and biochemical changes to cope with salt stress.     

Andrographis paniculata Leaf Extract Prevents Thioacetamide-Induced Liver Cirrhosis in Rats

This study investigated the hepatoprotective effects of ethanolic Andrographis paniculata leaf extract (ELAP) on thioacetamide-induced hepatotoxicity in rats. An acute toxicity study proved that ELAP is not toxic in rats. To examine the effects of ELAP in vivo, male Sprague Dawley rats were given intraperitoneal injections of vehicle 10% Tween-20, 5 mL/kg (normal control) or 200 mg/kg TAA thioacetamide (to induce liver cirrhosis) three times per week. Three additional groups were treated with thioacetamide plus daily oral silymarin (50 mg/kg) or ELAP (250 or 500 mg/kg). Liver injury was assessed using biochemical tests, macroscopic and microscopic tissue analysis, histopathology, and immunohistochemistry. In addition, HepG2 and WRL-68 cells were treated in vitro with ELAP fractions to test cytotoxicity. Rats treated with ELAP exhibited significantly lower liver/body weight ratios and smoother, more normal liver surfaces compared with the cirrhosis group. Histopathology using Hematoxylin and Eosin along with Masson’s Trichrome stain showed minimal disruption of hepatic cellular structure, minor fibrotic septa, a low degree of lymphocyte infiltration, and minimal collagen deposition after ELAP treatment. Immunohistochemistry indicated that ELAP induced down regulation of proliferating cell nuclear antigen. Also, hepatic antioxidant enzymes and oxidative stress parameters in ELAP-treated rats were comparable to silymarin-treated rats. ELAP administration reduced levels of altered serum liver biomarkers. ELAP fractions were non-cytotoxic to WRL-68 cells, but possessed anti-proliferative activity on HepG2 cells, which was confirmed by a significant elevation of lactate dehydrogenase, reactive oxygen species, cell membrane permeability, cytochrome c, and caspase-8,-9, and, -3/7 activity in HepG2 cells. A reduction of mitochondrial membrane potential was also detected in ELAP-treated HepG2 cells. The hepatoprotective effect of 500 mg/kg of ELAP is proposed to result from the reduction of thioacetamide-induced toxicity, normalizing reactive oxygen species levels, inhibiting cellular proliferation, and inducing apoptosis in HepG2 cells.     

Biosynthesis of andrographolide in Andrographis paniculata

Andrographolide, a diterpene lactone, is isolated from Andrographis paniculata which is well known for its medicinal properties. The biosynthetic route to andrographolide was studied using [1-¹³C]acetate, [2-¹³C]acetate and [1,6-¹³C₂]glucose. The peak enrichment of eight carbon atoms in the ¹³C NMR spectra of andrographolide suggested that deoxyxylulose pathway (DXP) is the major biosynthetic pathway to this diterpene.The contribution of the mevalonic acid pathway (MVA) is indicated by the observed ¹³C-labeling pattern, and because the labeling patterns indicate a simultaneous contribution of both methyl erythritol phosphate (MEP) and MVA pathways it can be deduced that cross-talk occurs between plastids and cytoplasm.     

An Investigation of Factors which Influence the Production of Abnormal Terpenoids by Callus Cultures of Andrographis paniculata Nees

Callus and suspension cultures of Andrographis paniculata havebeen shown to produce three new sesquiterpene lactones whichhave been named paniculides. A, B, and C and not the andrographolides(diterpenes) produced by the intact plant. Cultures derived from leaves, stems, hypocotyls, roots, andembryos accumulated paniculides, but no andrographolides weredetected. Conversely andrographolides, but no paniculides weredetected in extracts from leaves, stems and roots of intactseedlings. Modifications of the medium including the use of different syntheticauxins and the replacement of coconut milk with kinetin andcasein hydrolysate did not influence paniculide production bythe hypocotyl cultures. In addition an established root isolatecultured for 9 months on a simple synthetic medium without auxin,meso-inositol or coconut milk still synthesized significantquantities of paniculides. Diurnal fluctuations of temperature and light did not influencethe pattern of compounds produced although cultures grown incomplete darkness produced the individual paniculides in differentproportions. Possible explanations for these findings are discussed.     

穿心莲ent-柯巴基焦磷酸合酶基因的克隆与生物信息学分析

旨在克隆穿心莲CPS的编码基因,并进行序列分析.通过RT-PCR和cDNA末端快速扩增的技术从穿心莲叶片中获得目的基因;并利用生物信息学的方法对其编码蛋白进行功能分析.结果显示,获得了全长2 499 bp的cDNA序列,编码一个833aa的蛋白.序列同源性比较表明,该蛋白与野甘草、咖啡等其他植物来源的CPS具有较高的同源性.保守功能结构域分析显示,该蛋白包含一个富含Asp残基的植物萜类合酶的共有保守功能域“DXDD”,归属于萜类生物合成酶Ⅰ类超级家族和顺式聚异戊二烯焦磷酸合酶超级家族.初步推测克隆得到了穿心莲CPS的编码基因(命名为ApCPS,GenBank登录号为JN216843.1).     

Intra-specific hybridization: generator of genetic diversification and heterosis in Andrographis paniculata Nees. A bridge from extinction to survival

Andrographis paniculata (AP) has been stated as a low-diverse, endangered and red-listed plant species. Self-pollinated mating system, being an introduced species and experiencing a bottleneck as well as over exploitation cause such a consequence. Inter and intra-specific hybridizations have been suggested as essential techniques for generating genetic diversity. To test the effect of intra-specific hybridization on diversification and heterosis of AP, seven accessions were outcrossed manually in all 21 possible combinations. Three types of markers including morphological, phytochemical and RAPD markers were employed to evaluate the mentioned hypothesis. The results revealed that hybridization acted as a powerful engine for diversification of AP as it caused heterotic expression of the studied traits, simultaneously. Initially, it seems that additive and non-additive gene effects both can be considered as the genetic basis of heterosis in AP for the investigated traits. Agronomic and morphological traits were differentiated from each other, while positive heterosis was recorded mainly for agronomic traits but not for the morphological traits. Intra-specific hybridization increased the genetic diversity in AP population. Nevertheless, a part of this variation could also be attributed to the negative heterosis. The current exploration demonstrated the first ever conducted manual intra-specific hybridization among AP accessions in a mass scale. However, the 17 RAPD primers produced a monomorph pattern, but perhaps increasing the number of markers can feature a new genetic profile in this plant.     

ABA and GA3 affect the growth and pigment composition in Andrographis paniculata Wall.ex Nees., an important folk herb

In this study, 5 μmol·L-1 abscisic acid (ABA) and gibberellic acid (GA3) were used to study the effect of both growth regulators on the morphological parameters and pigment composition of Andrographispaniculata. The growth regulators were applied by means of foliar spray during morning hours. ABA treatment inhibited the growth of the stem and internodal length when compared with control, whereas GA3 treatment increased the plant height and internodal length. The total number of leaves per plant decreased in the ABA-treated plants, but GA3 treatment increased the total number of leaves when compared with the control. Both growth regulators (ABA and GA3) showed increased leaf area. ABA and GA3 treatments slightly decreased the total root growth at all the stages of growth. The growth regulator treatments increased the whole plant fresh and dry weight at all stages of growth. ABA enhanced the fresh and dry weight to a larger extent when compared with GA3. An increase in the total chlorophyll content was recorded in ABA and GA3 treatments. The chlorophyll-a, chlorophyll-b, and carote-noids were increased by ABA and GA3 treatments when compared with the control plants. The xanthophylls and anthocyanin content were increased with ABA and GA3 treatments in A. Paniculata plants.     

Some Characteristic Relaxant Effects of Aqueous Leaf Extract of Andrographis paniculata and Andrographolide on Guinea pig Tracheal Rings

The ethnomedicinal uses of the aqueous leaf extract of Andrographis paniculata Nees (AP) include treatment of pain and inflammation, malaria, asthma and common cold. We designed this study to characterize some effects of AP and those of its andrographolide constituent. Guinea pig tracheal rings suspended in organ baths containing PSS were precontracted with histamine or carbachol and then exposed to cumulative concentrations of AP, andographolide or theophylline. The effect of AP was tested in Ca2+-depleted tracheal rings stimulated with the EC50 of histamine in Ca2+-free PSS. IC50 and Emax values were calculated for each relaxant. Results showed that both AP and andrographolide possessed relaxant effects on the tracheal smooth muscle. While AP was more effective on histamine-induced contraction, andrographolide and theophylline were more effective on carbachol-induced contraction. The IC50 values of andrographolide were significantly higher than those of theophylline in the two contractile agents. The presence of AP significantly attenuated the contractile force produced by 6.4 x 10-3 M Ca2+ in Ca2+-depleted rings. It is concluded that andographolide contributes at least in part to the relaxant action of AP on tracheal smooth muscles. The mechanism of action is related to inhibition of Ca2+ influx into tracheal smooth muscle cells.     

烟草叶片衰老期过程中的蛋白质组学分析

大田烟叶生产过程中因打顶打叉的处理,改变了烟叶正常的衰老模式。为研究这一特殊的衰老机制,我们自旺长期开始,对‘云烟87’不同发育阶段烟株的中部叶片,进行形态观测、生理生化分析及蛋白质组学检测。结果显示：随着烟叶的逐渐成熟和衰老,烟草的叶色逐渐变黄,叶片逐渐变短、变窄,厚度减少;解剖结构清晰看到栅栏组织和海绵组织从最初的整齐排列到逐渐排列紊乱,组织细胞间轮廓不明显,细胞间隙明显增大;亚显微观测表明,淀粉粒在叶绿体中逐渐积累,类囊体片层结构被挤散,叶绿体膜被撑破。生理与生化分析表明衰老过程伴随着光合作用速率下降,光合色素降解加速,呼吸代谢的增加,这可能与衰老叶片中叶绿体逐渐崩塌和细胞膜透性增加相一致。iTRAQ标记方法共检测到不同发育阶段432个差异表达蛋白质,其中注释到308个与多种生命过程相关。蛋白差异富集分析表明,烟草叶片衰老过程中与光合作用等合成代谢相关蛋白多下调表达,而逆境反应及呼吸作用等分解代谢相关蛋白多上调表达。     

Chemical fingerprinting of Andrographis paniculata using HPLC, HPTLC and densitometry

An attempt has been made to develop a method by which to determine the chemical fingerprint of Andrographis paniculata (Acanthaceae). High-performance thin layer chromatography (HPTLC) was used to analyse hexane, chloroform, methanol and water extracts of leaves of A. paniculata. A computerised densitometer was applied to the two-dimensional spectrographic image analysis of the HPTLC plates. An HPLC equipped with a photodiode array detector was used for the analyses of these different extracts. The analyses showed that andrographolide and neoandrographolide are absent in the hexane extract but are present in greater amounts in the methanol extract as compared with the other extracts. These chromatograms may serve as a chemical fingerprint of the drug A. paniculata for quality control purposes and in the preparation of formulations based on the drug.     

Oviposition deterrent, ovicidal and gravid mortality effects of ethanolic extract of Andrographis paniculata Nees against the malarial vector Anopheles stephensi Liston (Diptera: Culicidae)

Oviposition is an important phenomenon of mosquitoes and has recently become the focus in the concept of integrated vector control management. In the present study, we evaluated oviposition deterrent, ovicidal and mortality effects of ethanolic extract of Andrographis paniculata Nees against gravid and oviposited females of Anopheles stephensi Liston. Water treated with the ethanolic extract had a high deterrent activity in ovipositing females: oviposition activity index values for the test species were –0.28, –0.45, –0.49 and –0.59 for extract concentrations of 29, 35, 41 and 46 p.p.m., respectively. High degrees of mortality were observed with various concentrations of extract: 1.12 (control) to 11.70 for gravid females, and 0.65 (control) to 10.25 for oviposited females. The highest mortality in both gravid and oviposited females was observed soon after they came in contact with oviposition medium treated with the extract, and this was found to be significant at doses higher than 35 p.p.m., suggesting possible contact toxicity of the extract. The extract caused moderate ovicidal activity against various age groups of A. stephensi, but it inflicted delayed effects such as high larval, pupal and adult mortality. The age of the eggs and the duration of the extract treatment influenced the ovicidal activity observed. It is clear that ethanolic extract of A. paniculata Nees can affect the oviposition cycle of A. stephensi Liston, thereby suppressing the vector population and adversely influencing transmission of the disease pathogen.     

RNA-Sequencing Analysis Reveals Critical Roles of Hormone Metabolism and Signaling Transduction in Seed Germination of Andrographis paniculata

Journal of Plant Growth Regulation - Andrographis paniculata is a traditional medicinal plant widely used in South-east Asia. It is cultivated from seed; however, seed germination—even of...     

Proteomic Analysis of Osmotic Stress-Responsive Proteins in Sugarcane Leaves

Osmotic stress-related proteins in sugarcane were identified using proteomics approach based on two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS). Sugarcane settlings were subjected to osmotic stress in the nutrient solution containing 10% (w/v) PEG 6000 for 14 h. Total proteins were extracted from leaves, and separated by 2-DE. Four typical spots exhibited significant changes in PEG treatment compared to control, which were identified using MALDI-TOF-MS successfully. The drought inducible 22 kDa protein and Rubisco small subunit were up-regulated while isoflavone reductase-like (IRLs, related to antioxidant defense system) protein and delta chain of ATP synthase were down-regulated by the osmotic stress. Analysis of the results showed that the most differential proteins under osmotic stress were acidic, unstable and transmembrane proteins, enriched with hydrophobic amino acids such as leucine and alanine which are extremely important for structural stabilization of proteins by hydrophobic interaction. However, the drought inducible 22 kDa protein was a hydrophile and non-transmembrane protein enriched with glutamic acid. These results provide new insight into the part of regulatory mechanism of adaptations to osmotic stress through differential expression of specific proteins and implicate several previously unrecognized proteins to osmotic stress.     

Proteomic Analysis of Protease Resistant Proteins in the Diabetic Rat Kidney

Glycation induced protein aggregation has been implicated in the development of diabetic complications and neurodegenerative diseases. These aggregates are known to be resistant to proteolytic digestion. Here we report the identification of protease resistant proteins from the streptozotocin induced diabetic rat kidney, which included enzymes in glucose metabolism and stress response proteins. These protease resistant proteins were characterized to be advanced glycation end products modified and ubiquitinated by immunological and mass spectrometry analysis. Further, diabetic rat kidney exhibited significantly impaired proteasomal activity. The functional analysis of identified physiologically important enzymes showed that their activity was reduced in diabetic condition. Loss of functional activity of these proteins was compensated by enhanced gene expression. Aggregation prone regions were predicted by in silico analysis and compared with advanced glycation end products modification sites. These findings suggested that the accumulation of protein aggregates is an inevitable consequence of impaired proteasomal activity and protease resistance due to advanced glycation end products modification.One of the foremost causes of diabetic complications is formation of sugar-derived substances called advanced glycation end products (AGEs),¹ which affect target cell through altered protein structure- function, matrix-matrix/matrix-cell interaction, and by activation of receptor for AGE (RAGE) signaling pathway (1). Although the accumulation of AGEs is a slow process in healthy individuals, their formation is markedly accelerated in diabetes because of hyperglycemia (2). AGE-modified proteins are thermostable and resistant to denaturation. The stability of proteins is believed to be because of additional negative charge (highly oxidized state) brought by AGE modification of proteins, which may contribute to protease resistance (3). Glycation induced protease resistance has been studied in collagen (4–6) and amyloid (7). In addition to glycation, impairment in the proteasomal function may facilitate accumulation of protease resistant protein aggregates in diabetes. Proteasome mediated protein degradation is a central quality control mechanism in the cell. Activity of proteasome is affected during aging (8) and physiological disorders like diabetes (9) resulting in accumulation of ubiquitinated protein aggregates. In muscle extract of diabetic rats, accumulation of toxic glycated proteins was observed because of decreased proteasomal activity (6–9). This proteolytic system is of particular importance in protecting cells against adverse conditions, such as heat shock, glycation, or oxidative stress. However, when the generation of damaged proteins exceeds the capacity of the cell to degrade them, they are progressively accumulated leading to cytotoxicity (10). Severely aggregated, cross-linked, and oxidized proteins are poor substrates for degradation and inhibit the proteasomal activity (11).The kidney is one of the main organs affected in diabetes caused by accumulation of AGEs. Proteins of extracellular matrix, kidney, as well as proteins from circulation, get AGE modified and trapped in the kidney (12). Both intracellular and extracellular AGEs have been observed in the diabetic kidney. Extracellular AGEs interact with the RAGE leading to apoptosis and inflammation (13), whereas intracellular AGEs are formed because of various dicarbonyls. Eventually, both types of the AGEs contribute to kidney damage (14). Furthermore, methyl glyoxal, a highly reactive dicarbonyl covalently modifies the 20S proteasome, decreasing its activity in the diabetic kidney (15). Together AGE modification and decreased proteasomal function may be responsible for the accumulation of protease resistant proteins (PRPs) in the diabetic kidney. In our previous study, we have reported the presence of AGE modified proteins in the kidney of the streptozotocin (STZ) induced diabetic rat (12). The current work is inspired by a DARTS (drug affinity responsive target stability) approach, wherein the drug targets are relatively less susceptible to protease action on drug binding (16). A similar approach was adopted here to identify protease resistant proteins from the diabetic kidney. These proteins were characterized to be AGE modified and ubiquitinated by Western blot analysis and mass spectrometry. Functional characterization and expression analysis of some of the identified proteins was performed to gain insight into the consequences of these modifications in diabetes. Further, aggregation prone regions in these proteins were predicted by the in silico approach. These findings shed light on the role of identified PRPs in diabetic complications.     

In vitro alpha-glucosidase and alpha-amylase enzyme inhibitory effects of Andrographis paniculata extract and andrographolide

There has been an enormous interest in the development of alternative medicines for type 2 diabetes, specifically screening for phytochemicals with the ability to delay or prevent glucose absorption. The goal of the present study was to provide in vitro evidence for potential inhibition of alpha-glucosidase and alpha-amylase enzymes, followed by a confirmatory in vivo study on rats to generate a stronger biochemical rationale for further studies on the ethanolic extract of Andrographis paniculata and andrographolide. The extract showed appreciable alpha-glucosidase inhibitory effect in a concentration-dependent manner (IC(50)=17.2+/-0.15 mg/ml) and a weak alpha-amylase inhibitory activity (IC(50)=50.9+/-0.17 mg/ml). Andrographolide demonstrated a similar (IC(50)=11.0+/-0.28 mg/ml) alpha-glucosidase and alpha-amylase inhibitory activity (IC(50)=11.3+/-0.29 mg/ml). The positive in vitro enzyme inhibition tests paved way for confirmatory in vivo studies. The in vivo studies demonstrated that A. paniculata extract significantly (P<0.05) reduced peak blood glucose and area under curve in diabetic rats when challenged with oral administration of starch and sucrose. Further, andrographolide also caused a significant (P<0.05) reduction in peak blood glucose and area under the curve in diabetic rats. Hence alpha-glucosidase inhibition may possibly be one of the mechanisms for the A. paniculata extract to exert antidiabetic activity and indicates that AP extract can be considered as a potential candidate for the management of type 2 diabetes mellitus.