Application of Accelerated Solvent Extraction Technique for Analysis of Active Components in Traditional Chinese Medicinal Herbs

以两种药材为研究实例,对加速溶剂萃取法(ASE)在中药材有效成分提取研究中的应用进行了简要介绍。采用正交试验法考察了提取丹参中丹酚酸B的提取条件(萃取温度、静态萃取时间、萃取溶剂以及料液比),确定了较好的实验条件。比较了ASE、水蒸气蒸馏法、超声波提取法及索氏提取法对木香挥发油的提取效果,结果表明ASE对木香挥发油的提取效果最好。The application of accelerated solvent extraction(ASE) technique for the Analysis of active components in traditional Chinese medicinal herbs was introduced by using two kinds of herbs as examples.The conditions including extraction temperature,static extraction time,the ratio of material to solvent and solvent of ASE for extraction of salvianolic acid B in Salvia miltiorrhiza were optimized by orthogonal experiments,and the optimal conditions were obtained.Different extraction methods(ASE,steam distillation,ultrasonic wave and Soxhlet extraction) were used to extract volatile oil in Aucklandia lappa Decne.Results of the comparative experiments indicated that ASE was the most effective method in this case.All the results from these studies demonstrate that ASE is indeed a powerful tool in the preparation of herbal extracts for downstream chromatographic analysis.青岛“2004将才计划”(04-3-JJ-11);; 共建生物医药研发测试中心(LS-05-KJZX-76)资

The Changes in Ca~(2+) level and ultrastructure in the Leaf Cells of Garyota urens L. under Low Temperature Stress

用焦锑酸钙沉淀的电镜细胞化学方法 ,研究了低温胁迫下董棕 (GaryotaurensL .)幼苗叶肉细胞内Ca2 + 水平的变化。研究结果表明 ,未经低温处理的董棕幼苗叶肉细胞 ,焦锑酸钙沉淀颗粒大量出现在液泡和细胞间隙中 ,细胞壁中也可见少量沉淀 ,而细胞基质中则看不到焦锑酸钙沉淀 ;经 2℃ 48h低温处理后 ,细胞基质和细胞膜上焦锑酸钙沉淀增加 ,而液泡和细胞间隙中的焦锑酸钙沉淀则显著减少 ,并且超微结构已初步显示出寒害的特征 ,叶绿体外膜部分破损 ,类囊体片层稀疏且排列不规则 ,光合速率明显下降等 ;经 2℃ 1 2 0h低温处理后 ,细胞间隙内的焦锑酸钙沉淀极少 ,有的也紧贴在细胞外壁上 ,而细胞基质和细胞膜上则分布有非常多的焦锑酸钙沉淀 ,在核基质和液泡中也可见到少量的焦锑酸钙沉淀 ,并且超微结构遭到了显著破坏 ,叶绿体结构完全被破坏 ,核膜与液泡膜严重破损 ,内部结构模糊 ,细胞只表现为呼吸作用 ,不进行光合作用。表明Ca2 + 的区域性分布的变化与植物抗寒性存在一定关系。The changes in Ca 2+ localization in the leaf cells of Garyota urens L. under chilling stress were investigated with calcium antimonate precipitate_electromicroscopic_cytochemical methods. When Garyota urens L. grew on the normal temperature, it was shown that the deposits of calcium antimonate being the indicator for Ca 2+ localization mainly concentrated within the vacuoles and intercellular spaces and there was also some Ca 2+ deposits in cell walls. But when Garyota urens L. was treated by the temperature of 2℃ for 48 h, the level of Ca 2+ increased in cytoplasm and plasma membrane, but decreased in vacuoles and intercellular spaces considerably. At the same time, the ultrastructure of chloroplasts suffered from chilling: the membrane of chloroplasts had been damaged, the layer of thylakoids was exiguous and unclear, the photosynthetic rate decreased evidently. And when Garyota urens L. was treated by the temperature of 2℃ for 120 h, the deposits of Ca 2+ mainly concentrated within the cytoplasm, nucleus and plasma membrane and there was also some Ca 2+ deposits in vacuoles, and the ultrastructure of some cells was simultaneously damaged severely: Chloroplasts structure, vacuole membrane and nuclear membrane had been damaged fully, the structure within the cell had become unclear, and the cell only have respiration. Thus it can be seen that there are some relations between the changes in Ca 2+ distribution within the cell and plant cold_hardiness.国家建设部及厦门市建委资助项

Content of gentiopicroside and loganic acid in Radix gentianae and their fingerprints

To develop a HPLC-DAD-ESI-TOF/MS analysis method for the determination of gentiopicroside and loganic acid in Radix gentianae samples and for the research of their fingerprints.The samples were extracted using ASE for 10 min under 100 ℃ and 9.65 MPa,and divided into water phase and chloroform phase and analyzed them with HPLC-DAD-ESI-TOF/MS method respectively.Based on this method,the HPLC fingerprints of Radix gentianae were established.Comparing the spectrogram and mass spectrum of the chromatogram peak with the reference value,three compounds in water phase were identified as gentiopicroside,asafetida acid and loganic acid.There is no report of the compounds in chloroform phase.The content of gentiopicroside and loganic acid in samples of different groups were determined,separately.The fingerprints were compared by the software of the similarity evaluation system for chromatographic fingerprint.The water phase fingerprint congruence coefficients of samples from six different areas were above 0.90,however,the chloroform phase fingerprint congruence coefficients were within 0.62-0.99.This method can be used for determination of potent component in Radix gentianae and its quality control.Radix gentianae from different producing areas have the largest diversities,and the diversities embodied in the content of chloroform phase compounds.青岛“2004将才计划”(04-3-JJ-11);; 国家海洋局青年基金资助项目(2005602);; 崂山区政府区校共建生物医药研发测试中心资助(LS-05-KJZX-76)

Determination of Gentiopicroside and Loganic Acid in Radix Gentianae by MEKC and MEEKC Mode

目的建立胶束电动毛细管色谱(MEKC)和微乳液毛细管电动色谱(MEEKC)分析龙胆药材中龙胆苦苷和马钱子苷酸含量的方法。方法采用加速溶剂萃取法(ASE)对龙胆药材进行提取,萃取温度:100℃,压力:9.65MPa,萃取时间:10min。采用未涂层熔融石英毛细管(内径75μm,有效长度50cm)。分别考察了两种分离模式下电泳介质的构成和电泳过程中的各操作参数对样品分离过程的影响,优化了MEKC和MEEKC的分析条件,在各自对应的缓冲液体系下,MEKC和MEEKC分离电压分别为30和22kV,柱温均为25℃,检测波长均为238nm。结果在选定的工作条件下,龙胆苦苷和马钱子苷酸与其他组分达到了基线分离,两种成分的浓度与其响应信号值之间具有较好的线性相关性,加标回收率在96.3%～105.1%之间,检测限均低于10mg·L-1,对6处不同产地的龙胆药材进行了分析,并对测定结果进行了t检验,结果表明,两种模式下,测定结果之间不存在显著性差异,而不同产地的龙胆药材的龙胆苦苷和马钱子苷酸含量之间存在较大差异。结论本方法简便,准确,快速,重现性较好,可用于龙胆药材有效成分的含量测定和质量控制。OBJECTIVE To develop MEKC and MEEKC modes for the determination of gentiopicroside and loganic acid in extracts of Radix Gentianae. METHODS The analyte was extracted from Radix Gentianae samples by accelerated solvent extraction, and the extraction conditions were optimized. Separation and determination were carried out on a bared fused silica capillary(50 cm×75 μm) with corresponding buffer. The run voltage of MEKC and MEEKC were 30kV and 22 kV respectively. Detection wavelength of DAD was at 238 nm and column temperature was 25 ℃. RESULTS The developed ASE-HPCE method was simple and reliable for the determination of gentiopicroside and loganic acid in Radix Gentianae samples with a broad linear dynamic range, a recovery range of 96.3%～105.16%,and the detection limit was below 10 mg·L-1. The contents of gentiopicroside and loganic acid in six samples from different regions were determined by the developed method in two modes. T-test value indicated that the determined contents of gentiopicroside and loganic acid by MEKC and MEEKC were consistent. CONCLUSION The method is simple,accurate,rapid and with good reproducibility.It can be used to determine active components in Radix Gentianae.国家自然科学基金重点项目(20235020);; 青岛“2004将才计划”(04-3-JJ-11);; 崂山区校区共建生物医药研发测试中心资助(LS-05-KJZX-76

大孔吸附树脂分离纯化龙胆药材中龙胆苦苷和马钱子苷酸的研究

目的:建立利用大孔吸附树脂对龙胆药材中龙胆苦苷和马钱子苷酸进行富集和分离纯化的方法。方法:采用加速溶剂萃取法对龙胆药材中两种有效成分进行高效提取,比较了D301,AB-8,D101,XDA-1四种大孔树脂对龙胆苦苷和马钱子苷酸的吸附性能,最终确定采用D301型大孔对脂对二者进行富集吸附,对其工艺参数进行优化,全程采用高效液相色谱进行目标化合物浓度检测。结果:化优后的工艺参数为:上样浓度:0.2 g/mL,最大上样量:0.25 g龙胆药材/g树脂,最佳静态吸附时间:8 h,采用8%和55%的乙醇溶液对龙胆苦苷和马钱子苷酸分别进行洗脱;龙胆苦苷和马钱子苷酸分别富集在8%和55%的乙醇洗脱液中,洗脱液浓缩后冷冻干燥,可得到纯度分别为74.3%和80.9%的粗产物,龙胆苦苷和马钱子苷酸的回收率分别为70.11%和67.82%。结论:此法效率较高,操作简便,即可用于实验室制备少量的难以购置的标准品,也可进行放大研究,用于工业生产。国家自然科学基金重点项目(20235020);; 青岛“2004将才计划”(04-3-JJ-11);; 共建生物医药研发测试中心(LS-05-KJZX-76)资

Effects of Moxibustion on Gastric Stress Ulcer Rats' Apoptosis Protein Phosphorylation

目的:研究艾灸对应激性胃溃疡大鼠胃黏膜细胞凋亡蛋白质磷酸化的影响,探讨艾灸促进胃黏膜损伤修复的信号转导机制。方法:将大鼠随机分为正常组、模型组、胃经穴组和对照点组,采用束缚冷应激法制作应激性胃溃疡大鼠模型,肉眼观察大鼠胃黏膜损伤程度,APOPTOSIS MICrOArrAy SlIdES芯片检测胃黏膜细胞凋亡蛋白质磷酸化水平。结果:与模型组比较,胃经穴组和对照点组大鼠胃黏膜损伤指数值均显著降低(P<0.05);与对照点组比较,胃经穴组大鼠胃黏膜损伤指数显著降低(P<0.05);APOPTOSIS MICrOArrAy SlIdES芯片检测结果显示:与模型组比较,胃经穴组大鼠胃黏膜细胞10种蛋白质磷酸化水平上调,其中bCl-Xl、MCl-1、bCl-2、IAPS 4种蛋白磷酸化水平差异有统计学意义(P<0.01,P<0.05);18种蛋白质磷酸化水平下调,其中Tnf、fAS、APAf-1、CASPASE-3、CASPASE-9、bAX 6种蛋白质磷酸化水平差异有统计学意义(P<0.01,P<0.05)。结论:艾灸可促进胃黏膜的损伤修复,调节多种凋亡相关信号蛋白质的磷酸化水平,并且存在一定的经脉脏腑相关性。Objective: To study the effects of moxibustion on apoptosis protein phosphorylation in rats with gastric stress ulcer and to explore the signal transduction mechanisms promoting gastric mucosal injury and repairing moxibustion.Methods: The rats were randomly divided into normal group,model group,stomach meridian group and the control point groups and each group had 10 rats.The stress ulcer rat model was established by using restraint cold stress method.We observed the rat gastric mucosa injury degree and Apoptosis Microarray Slides microarray was used to observe the gastric apoptosis protein phosphorylation levels.Results: Compared with the model group,the gastric mucosa injury index values of control point and meridian groups were significantly lower( P < 0.05).Compared with the control point group,a significant reduction in rat gastric mucosa injury index was in meridian group( P < 0.05).Apoptosis Microarray Slides microarray results showed that 10 kinds of protein phosphorylation levels increased,among which Bcl- XL,Mcl- 1,Bcl-2 and IAPs protein phosphorylation levels were statistically significant( P < 0.01,P < 0.05) and 18 kinds of protein phosphorylation levels decreased,among which TNF,Fas,Apaf- 1,Caspase- 3,Caspase- 9 and Bax protein phosphorylation levels were statistically significant( P < 0.01,P < 0.05).Conclusion: Moxibustion can promote gastric mucosal injury and repair and regulate phosphorylation of several proteins related to apoptotic signals and there is a certain correlation between the meridians organs.国家自然科学基金项目(30960484;81260556

Analysis of Alkaloids in Coptis chinensis Franch by High Performance Capillary Electrophoresis-Electrospray Ionization Time of Flight Mass Spectrometry

建立了高效毛细管电泳-电喷雾飞行时间质谱联用(HPCE-ESI-TOF/MS)快速定性分析黄连中生物碱类化合物的分析方法.使用未涂层石英毛细管,以50mmol/L乙酸铵-0.5%甲醇溶液(用氨水调至pH=7.2)作为运行缓冲液,分离电压为25kV;鞘液组成为50%甲醇-49.5%水-0.5%乙酸,鞘液流速为4μL/min;质谱选用正离子模式,碰撞电压(Fragmentor)为100V.结果表明,通过各色谱峰紫外光谱和质谱测得精确分子量结果,结合文献,对黄连中7种生物碱进行了鉴定.表明本方法简便、快速,是黄连中生物碱类化合物快速分离、鉴别的有效方法.A new method for the analysis of alkaloids in Coptis chinensis Franch was established by high performance capillary electrophoresis-electrospray ionization time of flight mass spectrometry (HPCE-ESI-TOF/MS). The real samples were separated by an uncoated capillary. 50 mmol/L ammonium acetate containing 0.5% methanol (pH=7.2) was used as the running buffer, and separation voltage was 25 kV. A coaxial sheath flow interface was used as the CE-MS interface, and a 50% methanol-49.5% water- 0.5% acetic acid mixture was used as the sheath liquid with a flow rate of 4 μL/min. The lens voltages in a positive ion mode with a collision induced dissociation (CID) voltage of 100 V were used for ESI-TOF/MS analysis. Seven alkaloids in Coptis chinensis Franch methanol extracts were separated and identified by CE-DAD and CE-ESI-TOF/MS. The coupling of HPCE separation with accurate mass measurement capability of ESI-TOF/MS provides an attractive tool for the identification of alkaloid compounds in Coptis chinensis Franch.国家自然科学基金重点项目(No.20235020);; 青岛“2004将才计划”(No.04-3-JJ-11);; 共建生物医药研发测试中心(No.LS-05-KJZX-76)资助项目

Analysis of alkaloids in Coptis chinensis Franch by high performance capillary electrophoresis-electrospray ionization time of flight mass Spectrometry

A new method for the analysis of alkaloids in Coptis chinensis Franch was established by high performance capillary electrophoresis-electrospray ionization time of flight mass spectrometry (HPCE-ESI-TOF/MS). The real samples were separated by an uncoated capillary. 50 mmol/L ammonium acetate containing 0.5% methanol (pH=7.2) was used as the running buffer, and separation voltage was 25 kV. A coaxial sheath flow interface was used as the CE-MS interface, and a 50% methanol-49.5% water- 0.5% acetic acid mixture was used as the sheath liquid with a flow rate of 4 mu L/min. The lens voltages in a positive ion mode with a collision induced dissociation (CID) voltage of 100 V were used for ESI-TOF/MS analysis. Seven alkaloids in Coptis chinensis Franch methanol extracts were separated and identified by CE-DAD and CE-ESI-TOF/MS. The coupling of HPCE separation with accurate mass measurement capability of ESI-TOF/MS provides an attractive tool for the identification of alkaloid compounds in Coptis chinensis Franch

Room-temperature quantum interference in single perovskite quantum dot junctions

钙钛矿材料由于其高量子产率、载流子迁移率和独特的光致发光特性而在光电材料领域存在诸多潜在的重要应用。研究钙钛矿材料在纳米尺度下电荷输运的独特尺寸效应对钙钛矿光电器件的设计和开发具有重要的指导意义。洪文晶教授课题组基于机械可控裂结技术自主研发了具有皮米级位移调控灵敏度和飞安级电学测量精度的精密科学仪器，对南开大学李跃龙副教授团队合成的钙钛矿量子点进行了深入表征，研究工作成功将量子干涉的研究体系拓展至在光电领域具有重要应用的钙钛矿材料领域，为未来制备基于量子干涉效应的新型钙钛矿器件提供了一种全新的思路。 这一跨学科国际合作研究工作是在化学化工学院洪文晶教授、英国Lancaster 大学物理系Colin J. Lambert教授以及南开大学电子信息与光电工程学院李跃龙副教授的共同指导下完成的。化工系硕士研究生郑海宁、Lancaster University大学Songjun Hou博士、南开大学硕士研究生辛晨光为论文第一作者。博士后林禄春，博士研究生谭志冰、郑珏婷，硕士研究生蒋枫、张珑漪，本科生何文翔、李庆民等参与了论文的研究工作。刘俊扬特任副研究员、师佳副教授和萨本栋微纳米研究院杨扬副教授也参与了部分指导工作。The studies of quantum interference effects through bulk perovskite materials at the Ångstrom scale still remain as a major challenge. Herein, we provide the observation of roomtemperature quantum interference effects in metal halide perovskite quantum dots (QDs) using the mechanically controllable break junction technique. Single-QD conductance measurements reveal that there are multiple conductance peaks for the CH3NH3PbBr3 and CH3NH3PbBr2.15Cl0.85 QDs, whose displacement distributions match the lattice constant of QDs, suggesting that the gold electrodes slide through different lattice sites of the QD via Auhalogen coupling. We also observe a distinct conductance ‘jump’ at the end of the sliding process, which is further evidence that quantum interference effects dominate charge transport in these single-QD junctions. This conductance ‘jump’ is also confirmed by our theoretical calculations utilizing density functional theory combined with quantum transport theory. Our measurements and theory create a pathway to exploit quantum interference effects in quantum-controlled perovskite materials.This work was supported by the National Key R&D Program of China (2017YFA0204902, 2014DFE60170, 2018YFB1500105), the National Natural Science Foundation of China (Nos. 21673195, 21503179, 21490573, 61674084, 61874167), the Open Fund of the Key Laboratory of Optical Information Science & Technology (Nankai University) of China, the Fundamental Research Funds for the Central Universities of China (63181321, 63191414, 96173224), and the 111 Project (B16027), the Tianjin Natural Science Foundation (17JCYBJC41400), FET Open project 767187—QuIET, the EU project BAC-TO-FUEL and the UK EPSRC projects EP/N017188/1, EP/M014452/1. 该工作得到国家重点研发计划课题（2017YFA0204902）、国家自然科学基金（21673195、21503179、21490573）、厦门大学“人工智能分析引擎”双一流重大专项等项目的资助，也得到了固体表面物理化学国家重点实验室、能源材料化学协同创新中心的支持

Desalting of Fermentation Wastewater with High Concentration of Salt by Electrodialysis

发酵工业废水处理是困扰发酵生产企业的一大难题。在有机酸和氨基酸的发酵生产及提取过程中，需加入大量无机酸碱调节剂调剂pH值，导致产生含高盐浓度无机盐(浓度大于100 g·L-1)的发酵废水，此类废水难以直接采用传统的生化方法进行处理，解决高盐限制是处理此类废水的关键。随着传统电渗析技术的发展和双极膜电渗析技术的出现，为高盐浓度发酵工业废水脱盐提供了新的方法。 本论文以吸附交换法提取柠檬酸后的高盐废液和浓缩等电法提取谷氨酸后的高盐废液为例，根据高盐废液的性质和成分差异，分别采用双极膜电渗析脱盐同时再生酸碱和传统电渗析单纯脱盐的方法进行处理，并对脱盐的问题提出了改进方案、优化了操作条件，以降低脱盐能耗。主要结论如下： (1)针对柠檬酸吸交废液成分简单、杂质少、色度浅的特点，以硫酸铵溶液模拟柠檬酸吸交废液，采用双极膜电渗析技术对其进行脱盐，同时再生硫酸和氨水。结果显示，模拟废液中硫酸根脱除率可以达到99%，整个电渗析过程膜通量为4.38 mol·m-2·h-1，能耗为3.10 kWh·kg-1 SO42-。为了进一步减小膜两侧溶液中的盐浓度梯度，降低能耗，采用分级的方法对电渗析操作进行了优化。与单级双极膜电渗析相比，二级和三级双极膜电渗析全过程的膜通量分别提高了20%和9%，能耗分别降低了10%和4%。 (2)测定了浓缩等电提取谷氨酸后的废液中各物质的浓度，针对其成分复杂、杂质含量高，色度深的特点，采用传统电渗析对浓缩废液进行脱盐。比较了三种不同膜组合的电渗析脱盐过程及性能指标，结果显示使用GeCM-1109-2+GeAM-1109-2膜组合时废液电渗析脱盐的效果最好。选择该膜组合为后续实验用膜。 (3)采用调节谷氨酸浓缩废液初始pH值的方法减少电渗析脱盐过程中有机酸和氨基酸的迁移。结果显示，先调节废液初始pH值至2.00再进行电渗析脱盐，当废液中硫酸根脱除率达到85%时，乳酸、谷氨酸的迁移率分别不调节pH值时(pH 2.67)降低了53%和66%，但能耗略有升高。 (4)考察了脱色预处理对降低谷氨酸浓缩废液电渗析脱盐能耗的效果。分别采用活性炭和30%过氧化氢溶液对废液进行脱色处理，对脱色温度、脱色剂用量和脱色时间进行了优化。以活性炭为脱色剂，当用量为100 g·L-1废液和200 g·L-1废液时，在60 ℃条件下处理30 min，废液的脱色率分别可达61%和75%，脱色后废液电渗析脱盐能耗分别降低了19%和25%；以过氧化氢为脱色剂，当30%过氧化氢溶液与废液体积比为1:3时，处理温度60 ℃条件下氧化处理120 min，废液脱色率可以达到81%，电渗析脱盐能耗降低29%