寄生隐丛赤壳菌Cryphonectria parasitica致病毒素Cp-I的分离纯化和结构分析

寄生隐丛赤壳菌Cryphonectria parasitica菌株经液体培养,石油醚萃取其发酵液获得对板栗带叶嫩枝具有致萎活性的粗提物,以氯仿:石油醚:甲醇(6:2:2)作洗脱剂,粗提物经硅胶色谱分离,共得到3组纯组份,其中第1组份(Cp-I)对板栗幼苗致萎活性较高.质谱、核磁共振和红外光谱测定表明Cp-I分子量为278,化学式为C<,16>H<,22>O<,4>.     

白肉榕叶提取物的生物活性及化学成分研究

本文以白肉榕叶为研究对象,采用药剂浸泡法和平板涂布法分别测定了白肉榕乙醇粗提物对蚕豆蚜、致倦库蚊和福寿螺的毒杀活性以及对蜡样芽胞杆菌、大肠杆菌、伤寒杆菌、痢疾志贺氏菌和金黄色葡萄球菌5种供试菌的抑菌活性。结果表明,白肉榕乙醇粗提物对蚕豆蚜、致倦库蚊和福寿螺有生物活性,而对上述5种供试菌没有抑菌活性。采用液-液萃取法对白肉榕乙醇粗提物进行萃取分离,并对得到的石油醚萃取物、乙酸乙酯萃取物和水相3种不同极性部位进行生物活性测定。生物测定结果表明,白肉榕叶的活性成分主要存在于石油醚萃取物和乙酸乙酯萃取物中。从乙酸乙酯萃取物中进一步分离得到1个活性成分,经核磁共振氢谱(~1H NMR)和碳谱(~(13)C NMR)鉴定为桦木醇(betulin)。该化合物对致倦库蚊3龄幼虫具有较好的杀虫活性,24 h的LC50值为16.44 mg/L。     

大丽轮枝菌毒素的多糖组份对棉花致萎作用的研究

用β-葡萄糖苷酶水解大丽轮枝菌(Verticillium dahliae Kleb.)毒素中的多糖组份。发现经酶处理后的5个不同致病力类型的棉花黄萎病菌(大丽轮枝菌)菌株毒素都不能影响整个毒素复合物对棉花的致萎作用,表明毒素中的多糖组份在棉花的致萎作用中不占有重量地位。     

小桐子提取物除草活性的生物测定

为全面了解小桐子(Jatropha curcas L. )提取物的除草活性,以萝卜(Raphanus sativus L. )、苋(Amaranthus tricolor L. )、苏丹草[Sorghum sudanense (Piper) Stapf]和黑麦草(Lolium perenne L. )为实验材料,对小桐子果壳和枝叶的水、乙醇(体积分数95%)、正丁醇、乙酸乙酯、氯仿和石油醚粗提物的除草活性进行了生物测定,并从中筛选出抑制作用最强的水粗提物进行进一步的活性组分分离及其除草活性的生物测定.测定结果显示,小桐子果壳和枝叶的6种溶剂提取物(10 g·L~(-1))对供试的4种植物幼苗的根长和茎高均有不同程度的抑制作用,其中水粗提物和乙醇(体积分数95%)粗提物的抑制作用较强,且水粗提物对供试的4种植物幼苗的根长和茎高的抑制作用均在75%以上,显著高于其他溶剂粗提物(P<0.05);石油醚粗提物的抑制作用最小,均在10%以下.小桐子果壳和枝叶水粗提物的石油醚、氯仿、乙酸乙酯、正丁醇和水萃取物(10 g·L~(-1))对萝卜和苏丹草幼苗的根长和茎高均有不同程度的抑制作用,其中水、正丁醇和乙酸乙酯萃取物的抑制作用显著高于氯仿和石油醚萃取物,抑制率均在70%以上;水萃取物的抑制作用最强,抑制率均在80%以上;石油醚萃取物的抑制作用最小,抑制率均在10%以下.研究结果表明,小桐子果壳和枝叶的水粗提物具有一定的除草活性,其有效成分为极性较大的组分.     

一株红茄苳内生烟曲霉菌的分离鉴定及其发酵产物抗氧化活性

【目的】红树林内生真菌生长于特殊的生境,有些菌株能分泌出结构新颖和具有生物活性的物质。从红树林植物红茄苳(Rhizophora mucronata)中分离获得一株内生真菌HQD24,对其进行生物学鉴定和抗氧化活性评价。【方法】综合运用形态结构及ITS r DNA序列分析,HQD24被鉴定为曲霉属(Aspergillus)的烟曲霉(Aspergillus fumigatus)。采用抗氧化多活性模型,2-2′二苯基-1-三硝基苯肼(DPPH)自由基测定法、2′-联氨-双-3-乙基苯并噻唑啉-6-磺酸(ABTS)自由基测定法、超氧自由基测定法、还原Fe~(3+)能力、螯合Fe~(2+)能力对HQD24麦麸发酵提取物的石油醚萃取物、二氯甲烷萃取物、乙酸乙酯萃取物和正丁醇萃取物进行抗氧化活性评价。【结果】经抗氧化活性实验发现不同有机溶剂萃取成分都具有清除DPPH自由基、ABTS自由基、超氧自由基和还原Fe~(3+)、螯合Fe~(2+)的能力,尤其是清除ABTS自由基和超氧自由基,抗氧化活性的能力都随着浓度的增高而增强。当不同有机溶剂萃取粗提物浓度为1.5 g/L时,二氯甲烷萃取物清除ABTS自由基率为71.92%,正丁醇萃取物清除超氧自由基率为62.36%,综合评价,HQD24的发酵产物中具有抗氧化活性的物质主要分布于二氯甲烷萃取粗提物和正丁醇萃取粗提物,其次分布于乙酸乙酯萃取粗提物,石油醚萃取粗提物中抗氧化活性物质含量最少。【结论】研究结果预示红树林内生烟曲霉菌是产生抗氧化活性物质的重要资源,HQD24菌株可以作为进一步实验研究的对象。     

小桐子枝叶提取物对蚜虫的毒杀活性

以水、乙醇、正丁醇、乙酸乙酯、氯仿和石油醚为溶剂,采用冷浸法对小桐子(Jatropha curcas L. )枝叶进行了粗提并采用喷雾法测定了各粗提物对豌豆长管蚜[Acyrthosiphon pisum (Harris)]和桃蚜[Myzus persicae (Sulzer)]的毒力,从中筛选出毒力最高的粗提物进行进一步的活性组分分离及毒杀活性测定.结果显示,随溶剂极性的减小,小桐子枝叶不同溶剂粗提物的提取率降低,其中水和乙醇粗提物的提取率较高,分别为18.76%和11.94%;6种溶剂粗提物对豌豆长管蚜和桃蚜都具有一定的毒力,其中,乙醇粗提物的毒力显著高于其他溶剂粗提物,LC50分别为2.693 3和2.565 6 g·L-1.乙醇粗提物的水、正丁醇、乙酸乙酯、氯仿和石油醚萃取物均对桃蚜有一定的毒杀活性,其中石油醚萃取物的萃取率最高(74.27%),对桃蚜的毒杀活性均显著高于其他溶剂萃取物,8 g·L-1石油醚萃取物对桃蚜的24 h校正死亡率高达94.5%.研究结果表明,小桐子枝叶的灭蚜活性组分存在于其乙醇粗提物的低极性组分中.根据实验结果初步确定了小桐子枝叶灭蚜活性组分的提取流程.     

松生拟层孔菌乙醇提取物抗肿瘤作用及其机制的实验研究

目的:探讨松生拟层孔菌抗肿瘤作用及其机制.方法:采用乙醇提取与硅胶柱层析分离,并对其乙醇粗提物、石油醚相、乙酸乙酯相提取物分别进行了体内(小鼠移植性肿瘤),外(采用Alamar Blue法)抗肿瘤活性研究.结果:松生拟层孔菌的乙醇粗提物,石油醚相,乙醇乙酯相在100ug/ml时,体外对NCI-H460细胞株具有增殖抑制作用,100mg/kg时对在体的H22瘤细胞具有显著的抗肿瘤活性,并能诱导H22瘤细胞凋亡,显著提高荷瘤鼠血清中IL-2、TNF-a的水平.结论:松生拟层孔菌乙醇提取物具有抗肿瘤作用,并能增强机体的免疫功能.     

石榴皮抗氧化活性成分的提取及其组分的研究

以DPPH抑制率为指标,研究石榴皮中抗氧化活性成分。采用超声波法进行提取,通过正交实验确定了最佳提取工艺为:乙醇浓度60%、pH3、料液比(m/v)1∶15、提取3次,每次40 min。石榴皮粗提物依次用石油醚、氯仿、乙酸乙酯、正丁醇、水萃取,经D101大孔吸附树脂纯化,并采用液相色谱-质谱联用法(LC-MS)对石榴皮抗氧化成分进行鉴定。结果表明,石榴皮粗提物中,正丁醇部位对DPPH的抑制率最高,达62.68%,液-质联用鉴定出安石榴苷的两种异构体为石榴皮中最主要的抗氧化成分。     

枯盘斑多毛孢Pf-毒素组分的研究I.活性组分I的结构分析

以正丁醇:水:甲醇(4:2:1)作洗脱剂,通过硅胶H60型柱反复柱层析,将3种有致病活性的物质充分纯化,在-40℃下冷冻干燥后,它们为褐色深浅不一的蓬松状物质,且极易吸潮。活性组分I(Rf0.83)、活性组分II(Rf0.79)和活性组分III(Rf0.80)对马尾松切根幼苗和湿地松切根幼苗针叶都有致萎作用,通过质谱(MS)、核磁共振谱(1HNMR、)和红外光谱(IR)等分析手段确定出所分离的活性组分I的化学组成为C5H11O5N(M=165)。     

地龙蛋白对人增生性瘢痕成纤维细胞的作用研究

目的:研究鲜地龙提取物对人增生性瘢痕成纤维细胞(HSF)增殖作用,及从其分离纯化的活性蛋白对基质金属蛋白酶(MMP-1、9)表达影响。方法:地龙提取物依次通过DEAE离子柱层析、葡聚糖凝胶柱层析分离纯化,应用HSF细胞模型,以活性为导向得到活性组份。获得样品组份进行部分理化性质实验,及对HSF细胞基质金属蛋白酶(MMP-1、9)表达影响。结果:地龙样品组份明显抑制HSF细胞增殖;样品经化学反应定性为蛋白,SDS-PAGE电泳法测定分子量为38k Da,p H值的平均值为7.82。分离纯化的DB-2样品使基质金属蛋白酶表达量下调,P0.05。结论:地龙样品组份能够显著抑制HSF细胞增殖,具有开发成为治疗瘢痕药物的潜力。     

瓜果腐霉菌丝中除草活性物质的初步研究

将瓜果腐霉进行液体发酵,其菌丝用等体积乙酸乙酯和甲醇进行依次萃取,甲醇萃取液旋转蒸发去溶剂后进行硅胶柱层析,以乙酸乙酯和石油醚(V/V=3:1和V/V=2:1)的混合液进行梯度洗脱,每50 mL收集为一个馏分,共收集到40个馏分。生物测定结果表明,以乙酸乙酯和石油醚(V/V=2:1)洗脱得到的馏分21 24对供试杂草马唐表现出了较强的活性,其对马唐的生长抑制作用均为4级。合并馏分21 24,以乙酸乙酯和石油醚(V/V=2:1)的混合液为展开剂进行等度洗脱,每50 mL收集为一个馏分,共收集到20个馏分。生物测定结果表明,馏分3对马唐有较强的抑制活性。HPLC分析发现,该馏分主要含有3个组分,其保留时间分别为12.7、14.0和30.5 min。     

生物破乳菌的筛选及发酵条件的优化

生物破乳剂的开发可以降低油田乳状液对石油工业和生态环境的负面影响并减少化学破乳剂的使用量。本研究建立了一套高效、便捷的破乳菌筛选方法, 并对破乳菌的特性进行研究。利用大庆油田受石油污染土壤作为菌种来源, 将Tween 60-water(0.072%, V/V)和Span 60-oil (0.028%, V/V)以6.5:3.5体积比配置出可以稳定200 h以上的O/W型乳状液, 用于破乳菌破乳效能的评价。经过分离纯化、血平板试验、排油试验和破乳试验最终筛选出2株24 h平均排油率在80%以上的优势破乳菌, 初步鉴定为芽孢杆菌属(Bascillus)。通过该破乳菌发酵条的件优化得到, 当温度为25°C, 摇床转数为160 r/min, pH值为9, 接菌量为20%时对该破乳菌生长速率最快, 积累发酵产物的量最多; 当温度为35°C, 摇床转数为120 r/min, pH值为9, 接菌量为2%时该破乳菌代谢产物的破乳活性最高。     

Acaricidal activity of petroleum ether extract of neem (Azadirachta indica) oil and its four fractions separated by column chromatography against Sarcoptes scabiei var. cuniculi larvae in vitro

The petroleum ether extract of neem oil and its four fractions separated by column chromatography was diluted at different concentrations with liquid paraffin. The acaricidal bioassay was conducted using a dipping method. The results indicated that the median lethal concentration (LC50) of the petroleum ether extract (at the concentration of 500.0ml/l) was 70.9ml/l, 24h after treatment. At concentrations of 500.0, 250.0, 125.0, 62.5 and 31.2ml/l, the median lethal times (LT50) of the petroleum ether extract were 8.7, 8.8, 10.8, 11.5 and 13.1h, respectively. Thin-layer chromatography (TLC) showed that the petroleum ether extract of neem oil separated into four fractions (F1-F4). Acaricidal activity of 68.3% and 100.0% in the F2 and F4 was confirmed. These results suggest that petroleum ether extracts of neem oil and its four fractions possess useful acaricidal activity in vitro.     

Acyl-coenzyme A: cholesterol acyltransferase assay: silica gel column separation of reaction products

Acyl-coenzyme A:cholesterol acyltransferase (ACAT) assays are usually performed by incubation of the enzyme with a labeled substrate followed by thin-layer chromatography separation and subsequent quantification of cholesteryl esters (CE) formed. Herein, a method is described for rapid separation of CE from other lipids, by elution from a silica gel column with a solvent mixture of petroleum ether/diethyl ether (98:2, v/v). Silica gel column chromatography is reliable and more rapid and safer than TLC. The best results were obtained when the reaction was stopped by Dole extraction followed by CE separation on a silica gel column. Assays for ACAT from rat intestinal microsomes showed that the specific activity values obtained using this method were reproducible and in good agreement with those obtained by conventional TLC method.     

An improved method for the extraction of cholesterol and phospholipids from the bovine spinal cord

A pharmacopoeial grade of cholesterol and a separate phospholipid fraction can be obtained by this simple extraction procedure of bovine spinal cord. The tissue is collected in 5% (Wt/V) formalin. It is then separated from the formalin solution, minced and dried in a hot-air oven at 105 degrees C. Total lipids are extracted with petroleum ether. The cholesterol is separated from the phospholipids by extraction with a mixture of petroleum ether and acetone. The method allows for not only the recovery of the phospholipids but also a high yield of cholesterol without using saponification or tedious purification procedures.     

高效液相色谱法测定毛豆中吡虫啉残留量

采用高效液相色谱法测定毛豆中吡虫啉农药残留。试样用二氯甲烷超声波提取,中性氧化铝柱净化,石油醚去除脂类杂质,以0.2%冰乙酸水溶液-乙腈(70:30,体积比)为流动相,配备Agilent TC-c18柱、高效液相色谱紫外检测器检测(HPLC-UV),外标法定量。实验表明,毛豆样品中吡虫啉添加回收率为84.0%~102.8%,相对标准偏差(n=6)为6.8%~10.9%,样品中吡虫啉最低检测浓度为0.003 mg/kg。     

The acyl lipid composition of wheat leaves and moss protonemata using a new, non-carcinogenic extraction solvent system

As chloroform has proved to be carcinogenic we were looking for an alternative solvent system for chloroform:methanol widely used in plant lipid investigations. The lipids from leaves of wheat ( Triticum aestivum L. cv. Vakka) and from protonemata of the moss Ceratodon purpureus (Hedw.) Brid. were extracted with two petroleum ether:methanol solvent systems. The polar lipids were separated by two-dimensional thin-layer chromatography and the amounts of each lipid class were compared with those obtained from chloroform:methanol (2:1, v/v) extractions. The significantly higher amounts of phosphatidylinositol observed in petroleum ether:methanol (1:1, v/v) extraction suggest that the small amounts reported earlier in plants may be an artefact relating to the solvent system used. As petroleum ether:methanol (1:1, v/v) proved to be at least as good a solvent system as chloroform:methanol (2:1, v/v) we propose it as an alternative extractant for plant polar lipids.     

侧柏乙醇提取物对21种植物病原真菌的抑菌活性

采用菌丝生长速率法测定了侧柏(Platycladus orientalis)叶、小枝、球果和种子4个不同部位乙醇提取物对21种植物病原真菌的抑菌活性。结果显示:(1)在供试浓度为50g/L(相当于干样)时,侧柏各部位乙醇提取物对4种供试植物病原真菌均具有较好抑制作用,其中侧柏叶提取物的抑菌效果最好,对供试葡萄白腐病菌(Conio-thyrium diplodiella)、葡萄黑痘病菌(Elsinoe ampelina)、番茄绵腐病菌(Phytophthora melongenae)和青霉病菌(Penicilliu mexpansum)的EC50分别为:5.424、3.186、8.913和19.000g/L。(2)侧柏叶乙醇提取物的石油醚萃取物和乙酸乙酯萃取物抑菌活性均较好,在供试浓度为0.5g/L时,石油醚萃取物对苹果腐烂病菌(Valsa mali)和葡萄黑痘病菌(E.ampelina)的抑菌率分别为80.35%和60.23%;乙酸乙酯萃取物对以上2种植物病原菌的抑菌率分别为81.88%和64.06%。结果表明:侧柏叶、小枝、球果和种子乙醇提取物均具有一定抑菌活性,叶乙醇提取物的活性最好,活性成分主要集中在石油醚萃取物和乙酸乙酯萃取物中。     

Antioxidant activity of phenylpropanoid esters isolated and identified from Platycodon grandiflorum A. DC

Platycodon grandiflorum A. DC (Campanulaceae) is used as a traditional oriental medicine and also as a food in Korea. Here we investigated its antioxidant activity, and isolated and identified its active compounds. Petroleum ether extracts from the whole root of P. grandiflorum were fractionated by silica gel column chromatography using a solvent gradient (petroleum ether:diethyl ether, v/v; 9:1-5:5). The 8:2 fraction showed a higher radical scavenging activity than the other fractions, and active compounds were purified from this fraction by reversed-phased HPLC. Two active compounds were identified as coniferyl alcohol esters of palmitic and oleic acids by FAB-MS, UV, IR and NMR spectroscopy. The antioxidant activities of these two compounds, which were evaluated by 1,1-diphenyl-2-picrylhydrazyl (DPPH), superoxide and nitric oxide radical scavenging capacity, were found to be as high as those of BHT or BHA.