樟芝产生三萜类化合物的液体培养条件的响应面法优化

樟芝(Antrodia cinnamomea)是一种非褶菌目、多孔菌科的多年生蕈菌类,樟芝子实体中含有多种生理活性物质包括三萜类化合物,但野生樟芝只生长在台湾特有的牛樟树树干腐朽的心材内壁和枯死倒伏的牛樟树表面,生长极其缓慢,很难获得,价格昂贵。本研究的目的是通过液体培养樟芝的菌丝体以产生三萜类化合物,用响应面法优化了樟芝产生三萜类化合物的液体发酵条件。首先对樟芝的液体发酵的培养基成分及培养条件进行了单因素优化,研究发现樟芝液体发酵产生三萜类化合物的最佳碳源和氮源分别为葡萄糖和酵母提取物,最佳浓度分别为100 g/L和9 g/L,优化的培养基配方为(g/L):葡萄糖100.0,酵母提取物9.0,Mg SO4·7H2O 0.5,KH2PO41.0。优化的培养条件为:初始pH值6.0,温度28℃,接种量10%(v/v),摇床转速180 r/min。在此基础上,用响应面法对樟芝液体发酵产生三萜类化合物影响最大的三个因素即碳源、氮源和培养温度进行了优化,结果优化的三种因素条件为:葡萄糖104.71 g/L,酵母提取物9.93 g/L,温度28.42℃。用优化的培养条件液体培养樟芝,其三萜类化合物的产量达到391.45 mg/L,比优化前的产量223.39 mg/L提高了75.23%,为进一步研究液体培养樟芝产生三萜类化合物奠定了基础。     

Adenosine as an active component of Antrodia cinnamomea that prevents rat PC12 cells from serum deprivation-induced apoptosis through the activation of adenosine A(2A) receptors

Antrodia cinnamomea (formerly named Antrodia camphorata) is a rare medicinal fungus. We previously reported that it exhibits antioxidative, vasorelaxative, anti-inflammatory, and anti-angiogenic effects. When serum deprivation-induced apoptosis in neuronal-like PC12 cells was used as a stress model, the extract of A. cinnamomea displayed effectiveness in preventing serum-deprived apoptosis. Since our previous data show that the extract of A. cinnamomea contains adenosine (ADO), we attempt to investigate if the active component is ADO and to identify its targeting site in this study. After pre-incubation with ADO deaminase, neither ADO nor the extract of A. cinnamomea exerted any protection, demonstrating that the active component of A. cinnamomea is ADO. Furthermore, an ADO A(2A) receptor (A(2A)-R) antagonist was used and was able to block the protective effects of ADO and the extract of A. cinnamomea, demonstrating that the ADO targeting site in this model is A(2A)-R. Taken together, the protective effect of A. cinnamomea is owed to its active component, ADO, which acts through activation of A(2A)-R to prevent serum deprivation-induced PC12 cell apoptosis.     

响应面法优化深层发酵樟芝总三萜的提取工艺(英文)

采用响应面法对樟芝深层培养中总三萜的提取工艺进行优化。根据Box-Benhnken的中心组合实验设计原理,在单因素实验的基础上,选取溶剂浓度、提取温度和液固比为变量,应用响应面法进行三因素三水平的实验设计。以总三萜得率作为响应值,建立了樟芝总三萜提取的回归模型,对其提取条件进行进一步优化。结果表明,优化的总三萜提取条件为乙醇浓度86%,提取温度75℃,液固比37。进一步的实验也验证了该模型的有效性。     

均匀设计法优化烟管菌产漆酶培养基

应用均匀设计、二次多项式逐步回归分析对烟管菌(Bjerkandera adusta)WZFF.W-Y11产漆酶液态发酵培养基进行优化。结果表明,培养基组成为麸皮水解液1%、淀粉24.0 g/L、葡萄糖24.0 g/L、豆饼粉4.8 g/L、NH₄Cl 3.2g/L、KH₂PO₄ 3.2 g/L、MgSO₄·7H₂O 0.2 g/L、CuSO₄ ·5H₂O 0.006 g/L,起始pH6.5,在28℃、150r/min、250mL的摇瓶培养条件下可以稳定地获得9672U/L的漆酶活力。     

红根草三萜类化学成分的研究

该研究利用大孔吸附树脂D101、MCI树脂、正相硅胶、ODS、Sephadex LH-20和反相HPLC等多种色谱分离方法,对红根草95%乙醇提取物的乙酸乙酯萃取部位进行了分离。结果表明:通过分离共得到了10个化合物,经波谱数据分析鉴定为3-表-白桦脂酸(1)、白桦脂醇(2)、齐墩果酸(3)、3β,6β,23-三羟基-12-烯-28-齐墩果酸(4)、高根二醇(5)、2α,3β,23-三羟基-12-烯-28-乌苏酸(6)、2α,3β,19α-三羟基-12-烯-28-乌苏酸(7)、2α,3α,24-三羟基-12-烯-28-乌苏酸(8)、2α,3α,24-三羟基-12,20(30)-二烯-28-乌苏酸(9)、山奈酚-3-O-葡萄糖苷(10)。其中,化合物1为首次从鼠尾草属植物中分离得到,化合物2、4、5、6、7、8、9、10为首次从该植物中分离得到。     

南丹参三萜类化学成分的研究

为研究南丹参根中三萜类化学成分,该文采用95％乙醇浸泡提取,依次应用D101大孔吸附树脂、MCI、硅胶柱色谱、ODS及反相高效液相色谱方法进行分离纯化,根据理化性质及波谱数据鉴定其化合物的结构.结果表明:从南丹参根提取物中分离得到9个化合物,分别鉴定为齐墩果酸(1)、2α,3α-二羟基-12-烯-28-齐墩果酸(2)、...     

Triterpenoids from Gentiana scabra

Five triterpenoids, (20S)-dammara-13(17),24-dien-3-one, (20R)-dammara-13(17),24-dien-3-one, chirat-16-en-3-one, chirat-17(22)-en-3-one and 17beta,21beta-epoxyhopan-3-one, were isolated from the rhizomes and roots of Gentiana scabra together with five known ones, chiratenol, hop-17(21)-en-3-one, hop-17(21)-en-3beta-ol, lupeol and alpha-amyrin. The structures of new compounds were elucidated on the basis of spectroscopic studies.     

A Shortcut to the Optimization of Cellulase Production Using the Mutant Trichoderma reesei YC-108

Trichoderma reesei YC-108, a strain isolated by a kind of newly invented plate was found to over produce cellulase and it was then used as a cellulase producer. To get the maximum amount of cellulase, the combination of the medium ingredients, which has a profound influence on metabolic pathway was optimized using response surface methodology. The optimum composition was found to be 24.63 g/L wheat bran, 30.78 g/L avicel, and 19.16 g/L soya-bean cake powder. By using the optimized medium, the filter paper activity (FPA) increased nearly five times to 15.82 IU/mL in a 30 L stirred fermenter, carboxymethyl cellulase activity (CMCase) was increased from 83.02 to 628.05 IU/mL and the CMCase/FPA ratio was nearly doubled compared with the parent strain at initial medium.     

Optimization of Inoculation for In Vivo Production of Entomopathogenic Nematodes

Entomopathogenic nematodes are potent biopesticides that can be mass-produced by in vitro or in vivo methods. For in vivo production, consistently high infection rates are critical to efficiency of the process. Our objective was to optimize in vivo inoculation of Steinernema carpocapsae and Heterorhabditis bacteriophora in Galleria mellonella and Tenebrio molitor by determining effects of inoculation method, nematode concentration, and host density. We found immersing hosts in a nematode suspension to be approximately four times more efficient in time than pipeting inoculum onto the hosts. The number of hosts exhibiting signs of nematode infection increased with nematode concentration and decreased with host density per unit area. This is the first report indicating an effect of host density on inoculation efficiency. We did not detect an effect of nematode inoculum concentration on nematode yield per host or per gram of host. Yield was affected by host density in one of the four nematode-host combinations (S. carpocapsae and T. molitor). We conclude that optimization of inoculation parameters is a necessary component of developing an in vivo production system for entomopathogenic nematodes.     

Optimization of Antimicrobial Production by a Marine Actinomycete Streptomyces afghaniensis VPTS3-1 Isolated from Palk Strait, East Coast of India

Totally 25 marine soil samples were collected from the region of Palk Strait of Bay of Bengal, Tamil Nadu, and were subjected to the isolation of actinomycetes. Sixty-eight morphologically distinct isolates were obtained and 37% (25) of them had antimicrobial activity. The potential producer was named as Streptomyces sp. VPTS3-1 and the phylogenetic evaluation on the basis of 16S rDNA sequence further categorized the organism as Streptomyces afghaniensis VPTS3-1. Further, the antimicrobial compound was extracted from the isolate using various solvents and the antimicrobial efficacies were tested against bacterial and fungal pathogens. In addition, in vitro optimization of parameters for the antimicrobial compound production revealed that the suitable pH as 7–8, the period of incubation as 9 days, temperature (30°C), salinity (2%), and starch and KNO₃ as the suitable carbon and nitrogen sources respectively in starch–casein medium.     

Molecular characterization and expression analysis of Acmago and AcY14 in Antrodia cinnamomea

Mago nashi (Mago) and Y14 proteins, highly conserved among eukaryotes, participate in mRNA localization and splicing, and as such play important roles in oogenesis, embryogenesis and germ-line sex determination during animal development. Here we identified mago (Acmago) and Y14 (AcY14) homologues derived from Antrodia cinnamomea. Acmago encodes 149 amino acids and AcY14 encodes 168 amino acids. Multiple amino acid sequence alignment as well as secondary and tertiary structure prediction showed that AcMago and AcY14 have similar protein structure to the reported crystal structures of other Mago and Y14 proteins. During fungal development both Acmago and AcY14 genes were abundantly expressed in natural basidiomes. This is the first report of the molecular characterization and expression analysis of the mago and Y14 genes from fungi.     

Triterpenoids from Sanguisorba officinalis

Seven triterpenoids, i.e., 3beta-[(alpha-L-arabinopyranosyl)oxy]-19beta-hydroxyurs-12,20(30)-dien-28-oic acid (1), 3beta-[(alpha-L-arabinopyranosyl)oxy]-urs-11,13(18)-dien-28-oic acid beta-D-glucopyranosyl ester (2), 2alpha,3alpha,23-trihydroxyurs-12-en-24,28-dioic acid 28-beta-D-glucopyranosyl ester (3), 3beta-[(alpha-L-arabinopyranosyl)oxy]-urs-12,19(20)-dien-28-oic acid (4), 3beta-[(alpha-L-arabinopyranosyl)oxy]-urs-12,19(29)-dien-28-oic acid (5), 3beta-[(alpha-L-arabinopyranosyl)oxy]-19alpha-hydroxyolean-12-en-28-oic acid (6), 2alpha,3beta-dihydroxy-28-norurs-12,17,19(20),21-tetraen-23-oic cid (7), together with three known ones (8-10), were isolated from the roots of Sanguisorba officinalis. Their structures were determined by spectroscopic and chemical methods. Compounds 7 and 10 showed marginal inhibition activity against the growth of tumor cell lines.     

Triterpenoids from lipids of Rhodomicrobium vanniellii

Triterpenoids have been isolated from the lipids of the anaerobic, photosynthetic bacterium, Rhodomicrobium vannielii. These compounds constitute various hopanoids, including hydrocarbons, 22-, 29-and 3-hydroxy, 3-keto-, C-3 and C-21 methyl-triterpenoids, as well as fernenes.     

Protection of oxidative damage by aqueous extract from Antrodia camphorata mycelia in normal human erythrocytes

Antrodia camphorata (A. camphorata) is well known in Taiwan as a traditional Chinese medicine. The purpose of this study was to evaluate the ability of aqueous extract from A. camphorata mycelia to protect normal human erythrocytes against oxidative damage in vitro. Oxidative hemolysis and lipid/protein peroxidation of erythrocytes induced by the aqueous peroxyl radical [2,2'-Azobis(2-amidinopropane) dihydrochloride, AAPH] were suppressed by A. camphorata mycelia in a time-and concentration-dependent manner. A. camphorata mycelia also prevented the depletion of cytosolic antioxidant glutathione (GSH) and ATP in erythrocytes. Moreover, cultured human endothelial cell damage induced by AAPH was suppressed by A. camphorata mycelia. Interestingly, A. camphorata mycelia exhibited significant cytotoxicity against leukemia HL-60 cells but not against cultured human endothelial cells. These results imply that A. camphorata mycelia may have protective antioxidant and anticancer properties.     

Optimization of Process Variables for Lipase Biosynthesis from <Emphasis Type="Italic">Rhizopus oligosporus</Emphasis> NRRL 5905 Using Evolutionary Operation Factorial Design Technique

Extracellular lipase was produced from Rhizopus oligosporus NRRL 5905 through solid state fermentation (SSF). To provide an optimum fermentation conditions for maximum lipase yield, five process variables (temperature, liquid–solid ratio, pH, incubation period and spore concentration) were optimized using evolutionary operation (EVOP) factorial design technique taking into account the interaction between the process variables. Optimization through EVOP resulted in around 3 fold increase in lipase activity (77 U gds⁻¹) at a liquid–solid ratio of 1.5:1, fermentation temperature of 35°C, initial fermentation pH 6, incubation period 5 days and a spore concentration of 10⁸ spores ml⁻¹.     

牛樟芝发酵液提取物抗菌活性研究

牛樟芝作为一种珍稀食用和药用菌,具有极大的开发潜力。该研究以麦芽浸粉肉汤液体培养基(BD,美国BD公司)对牛樟芝菌丝体进行摇床培养60 d后,收获发酵液并用乙酸乙酯对其进行萃取,浓缩至干获得提取物;同时,采用抑菌圈法评价培养物对13种致病细菌抗菌活性(蜡样芽孢杆菌、缓慢芽孢杆菌、无乳链球菌、短小芽孢杆菌、福氏志贺氏菌、枯草芽孢杆菌、金黄色葡萄球菌、藤黄微球菌、副溶血性弧菌、溶血性葡萄球菌、铜尿假单胞菌、乙型副伤寒沙门氏菌、大肠埃希菌),并检测相应致病细菌的最低抑制浓度(MIC)。结果表明:牛樟芝麦芽浸粉肉汤发酵液提取物对供试的13种致病菌均有抑菌活性;在供试的13种致病菌中,提取物对缓慢芽孢杆菌、短小芽孢杆菌、枯草芽孢杆菌、副溶血性弧菌、藤黄微球菌5种致病菌的最低抑制浓度值均小于80μg·m L~(-1),其中对藤黄微球菌的最低抑制浓度最低为66.5μg·m~(-1);随着培养时间的增加,提取物的抗菌活性也增加。这说明牛樟芝菌丝体在液体培养条件下,能够产生广谱高效抑菌活性的次生代谢产物。该研究结果为牛樟芝进一步的有效利用开发奠定了理论基础。     

Optimization and Utilization of Agrobacterium-mediated Transient Protein Production in Nicotiana

Agrobacterium-mediated transient protein production in plants is a promising approach to produce vaccine antigens and therapeutic proteins within a short period of time. However, this technology is only just beginning to be applied to large-scale production as many technological obstacles to scale up are now being overcome. Here, we demonstrate a simple and reproducible method for industrial-scale transient protein production based on vacuum infiltration of Nicotiana plants with Agrobacteria carrying launch vectors. Optimization of Agrobacterium cultivation in AB medium allows direct dilution of the bacterial culture in Milli-Q water, simplifying the infiltration process. Among three tested species of Nicotiana, N. excelsiana (N. benthamiana × N. excelsior) was selected as the most promising host due to the ease of infiltration, high level of reporter protein production, and about two-fold higher biomass production under controlled environmental conditions. Induction of Agrobacterium harboring pBID4-GFP (Tobacco mosaic virus-based) using chemicals such as acetosyringone and monosaccharide had no effect on the protein production level. Infiltrating plant under 50 to 100 mbar for 30 or 60 sec resulted in about 95% infiltration of plant leaf tissues. Infiltration with Agrobacterium laboratory strain GV3101 showed the highest protein production compared to Agrobacteria laboratory strains LBA4404 and C58C1 and wild-type Agrobacteria strains at6, at10, at77 and A4. Co-expression of a viral RNA silencing suppressor, p23 or p19, in N. benthamiana resulted in earlier accumulation and increased production (15-25%) of target protein (influenza virus hemagglutinin).     

Optimization of Lipase Production from <Emphasis Type="Italic">Aspergillus terreus</Emphasis> by Response Surface Methodology and Its Potential for Synthesis of Partial Glycerides Under Solvent Free Conditions

Aspergillus terreus produces lipase 7.01 IU/ml in 96 h after optimization by one variable at a time method. Using the significant factors i.e. corn oil (A), sodium nitrate (B), casein (C), agitation rate (D) and incubation period (E) RSM was carried out resulting in 19.65 IU/ml from the combination +1(A), −1(B), −1(C), +1(D) and 0(E). The interactions between sodium nitrate, casein and agitation with corn oil were most significant. Scale up of production from 250 ml shake flask to 30 l bioreactor resulted in increased productivity of 0.52 IU/ml/h as against 0.2 IU/ml/h obtained in shake flasks. This lipase could carryout solvent free synthesis of partial glycerides of oleic acid with 96% efficiency in 12 h.     

Triterpenoids from the orchids Agrostophyllum brevipes and Agrostophyllum callosum

Agrostophyllinol and agrostophyllinone, two new triterpenoids, were isolated from the orchid Agrostophyllum brevipes. Agrostophyllinone was also isolated from another orchid Agrostophyllum callosum. The structures of agrostophyllinol and agrostophyllinone were established as 24-methylene-lanosta-9(11)-en-3beta-ol (5a) and 24-methylene-lanosta-9(11)-en-3-one (5c), respectively, from spectral and chemical evidence. The above triterpenoids are of considerable biogenetic importance.     

Optimization of date syrup for enhancement of the production of citric acid using immobilized cells of Aspergillus niger

Date syrup as an economical source of carbohydrates and immobilized Aspergillus niger J4, which was entrapped in calcium alginate pellets, were employed for enhancing the production of citric acid. Maximum production was achieved by pre-treating date syrup with 1.5% tricalcium phosphate to remove heavy metals. The production of citric acid using a pretreated medium was 38.87% higher than an untreated one that consumed sugar. The appropriate presence of nitrogen, phosphate and magnesium appeared to be important in order for citric acid to accumulate. The production of citric acid and the consumed sugar was higher when using 0.1% ammonium nitrate as the best source of nitrogen. The production of citric acid increased significantly when 0.1 g/l of KH₂PO₄ was added to the medium of date syrup. The addition of magnesium sulfate at the rate of 0.20 g/l had a stimulating effect on the production of citric acid. Maximum production of citric acid was obtained when calcium chloride was absent. One of the most important benefits of immobilized cells is their ability and stability to produce citric acid under a repeated batch culture. Over four repeated batches, the production of citric acid production was maintained for 24 days when each cycle continued for 144 h. The results obtained in the repeated batch cultivation using date syrup confirmed that date syrup could be used as a medium for the industrial production of citric acid.