Small‐Molecule Mechanism of Action Studies in Caenorhabditis elegans

A general protocol for exogenous small‐molecule pull‐down experiments with Caenorhabditis elegans is described; it provides a link between small‐molecule screens in worms and existing mutant and RNAi technologies, thereby enabling organismal mechanism of action studies for the natural product clovanemagnolol. Forward chemical genetic screens followed by mechanism of action studies with C. elegans, when coupled with genetic validation of identified targets to reproduce the small molecule's phenotypic effects, provide a unique platform for discovering the biological targets of compounds that affect multicellular processes. First, the use of an immobilized FK506 derivative and soluble competition experiments with optimally prepared soluble C. elegans proteome successfully identified interactions with FK506 binding proteins 1 to 6. This approach was used to determine an unknown mechanism of action for clovanemagnolol, a small molecule that promotes axonal branching in both primary neuronal cultures and in vivo in C. elegans. Following the synthesis of an appropriately functionalized solid‐phase reagent bearing a clovanemagnolol analogue pull‐down experiments employing soluble competition identified kinesin light chain‐1 (KLC‐1), a protein involved in axonal cargo transport, as a putative target. This was corroborated through the use of mutant worms lacking klc‐1 and possessing GFP neuronal labeling, reproducing the axonal branching phenotype induced by the small molecule clovanemagnolol.     

肿柄菊粗提物对线虫毒杀活性的研究

为评价肿柄菊粗提物毒杀线虫的潜力,利用石油醚、乙酸乙酯、二氟甲烷浸泡粉碎的肿柄菊枝叶混合物,经旋转蒸发获得粗提物,通过浸渍法检测了不同粗提物的毒杀线虫活性。结果表明：当浓度为10mg·mL^-1时,石油醚、乙酸乙酯和二氟甲烷粗提物24～72h内对线虫的毒杀效果较弱;当浓度为20mg·mL^-1时,二氯甲烷粗提物毒杀线虫活性最好,处理72h的线虫校正死亡率可达50．6％,乙酸乙酯粗提物和石油醚粗提物毒杀线虫活性均较弱;当浓度为40mg·mL^-1时,毒杀线虫活性大小依次是乙酸乙酯粗提物〉石油醚粗提物〉二氯甲烷粗提物,其中乙酸乙酯粗提物处理72h的线虫校正死亡率达73．6％。表明,对肿柄菊粗提物进行合理的处理、分离、纯化,有望开发出高效低毒、绿色环保的新型杀线虫农药。     

The lin-4 Gene Controls Fat Accumulation and Longevity in Caenorhabditis elegans

Previous studies have determined that lin-4, which was the first miRNA to be discovered, controls the timing of cell fate determination and life span in Caenorhabditis elegans. However, the mechanism of lin-4 involvement in these processes remains poorly understood. Fat storage is an essential aspect of the life cycle of organisms, and the function of lin-4 in fat accumulation is not clear. In this study, we showed that the fat content is reduced remarkably in C. elegans lin-4 mutants. Quantitative RT-PCR analysis revealed a considerable decrease in the levels of SBP-1 and OGA-1 mRNA in lin-4 mutants. We also showed that lin-4 mutants have a significantly shorter life span than wild-type worms. DCF assay experiments showed that the reactive oxygen species (ROS) levels increased and mitochondrial DNA (mtDNA) copy number decreased in loss-of-function lin-4 mutants. These mutants also showed attenuation of locomotion. Taken together, our findings suggest that lin-4 may play an important role in regulating fat accumulation and locomotion and that lin-4 may control the life span of C. elegans by mediating ROS production.     

紫玉米芯花青素的射频冷等离子体辅助提取及体内外抗氧化活性

采用射频冷等离子体辅助提取技术从紫玉米芯中提取花青素,通过单因素和响应面优化实验,考察放电功率、处理时间和工作压力对紫玉米芯花青素提取量的影响。利用超高效液相色谱仪-串联质谱仪（UPLC-MS）对紫玉米芯花青素的成分进行分析和鉴定,使用秀丽隐杆线虫和双氧水分别对提取的紫玉米芯花青素进行抗氧化性能评价。结果表明,冷等离子体预处理的最佳条件为：放电功率127 W、处理时间75 s、工作压力150 Pa,此条件下紫玉米芯花青素提取量为(4.05±0.05) mg/g(玉米芯),较溶剂提取法和超声辅助提取法的提取量分别提高49.45%和28.98% 。紫玉米芯花青素主要由天竺葵素-3-O-葡糖苷、矢车菊素-3-O-葡糖苷、矢车菊素和矮牵牛素-3-O-葡糖苷组成。与溶剂提取法和超声辅助提取法相比,冷等离子体辅助提取的花青素成分中矢车菊素-3-O-葡糖苷含量提高,天竺葵素-3-O-葡糖苷含量降低。紫玉米芯花青素清除1,1-二苯基-2-三硝基苯肼（DPPH）自由基、2,2-联氮-二(3-乙基-苯并噻唑-6-磺酸)二铵盐（ABTS+）自由基的半抑制质量浓度（IC50）分别为0.2972、0.2770 mg/mL,接近同样条件下抗坏血酸对DPPH自由基和ABTS+自由基的清除能力。采用紫玉米芯花青素喂养的秀丽隐杆线虫,平均寿命延长29.26%;较H2O2处理组,其抗氧化能力提高了26.25%,体内活性氧水平降低了60.83%。     

Benzofuran–Chalcone Hybrids as Potential Multifunctional Agents against Alzheimer’s Disease: Synthesis and in vivo Studies with Transgenic Caenorhabditis elegans

In the search for effective multifunctional agents for the treatment of Alzheimer’s disease (AD), a series of novel hybrids incorporating benzofuran and chalcone fragments were designed and synthesized. These hybrids were screened by using a transgenic Caenorhabditis elegans model that expresses the human β‐amyloid (Aβ) peptide. Among the hybrids investigated, (E)‐3‐(7‐methyl‐2‐(4‐methylbenzoyl)benzofuran‐5‐yl)‐1‐phenylprop‐2‐en‐1‐one ( 4 f ), (E)‐3‐(2‐benzoyl‐7‐methylbenzofuran‐5‐yl)‐1‐phenylprop‐2‐en‐1‐one ( 4 i ), and (E)‐3‐(2‐benzoyl‐7‐methylbenzofuran‐5‐yl)‐1‐(thiophen‐2‐yl)prop‐2‐en‐1‐one ( 4 m ) significantly decreased Aβ aggregation and increased acetylcholine (ACh) levels along with the overall availability of ACh at the synaptic junction. These compounds were also found to decrease acetylcholinesterase (AChE) levels, reduce oxidative stress in the worms, lower lipid content, and to provide protection against chemically induced cholinergic neurodegeneration. Overall, the multifunctional effects of these hybrids qualify them as potential drug leads for further development in AD therapy.     

以离子液体密度为例的分子性质预测模型建模方法探讨

分子性质预测模型是针对特定应用需求筛选设计化学品的有力工具,然而诸多相关建模过程中的测试集划分、交叉验证、算法选择等关键环节普遍存在严谨性不足的问题,模型真实预测性能难以保证。以基团贡献法预测离子液体密度为例,探讨了分子性质预测模型建模过程中数据集划分和交叉验证的重要性,提出了自动基团划分方法并研究了数据集中基团涉及分子个数对预测精度的影响。通过对比五种回归算法（多重线性回归、岭回归、随机森林、支持向量机、神经网络）,基于岭回归的基团贡献模型预测性能最佳,在由1078种离子液体、共计23034个数据点组成的数据集上得到的平均相对误差为1.88%。