Name: field collection and laboratory routine identification of rhodiola crenulata
Uploaded: 2023-10-27T13:20:00.000+00:00
Duration: 6 min 44 s
Description: Watch this Scientific Journal Video about Field Collection and Laboratory Routine Identification of Rhodiola crenulata at JoVE.com

本研究得到了国家自然科学基金（81973569、82274207、82104533）、成都中医药大学星林学者研究推进项目（XKTD2022013）和宁夏市重点研发计划（2023BEG02012）的支持。

红景天 采集自中国四川省甘孜藏族自治州甘孜县卓达雪山（北纬31.44570°，东经99.96086°，海拔4892米）。这些植物由成都中医药大学民族医学院的张毅教授鉴定为真品。
1. 红景天的收集
<ol><li>拍摄 红景天的栖息地地图。</li>
	<li>拍摄红景天的整个植物、叶子、花萼和根茎。</li>
	<li>用铁锹清除红 景天 1 m 范围内的杂草和碎石，以确保随后的顺利开采。</li>
	<li>用锄头挖土，直到看到整个根际并收集主根。 注:用于药用部位的 红景天 的根和根茎应在秋季花茎枯萎时采集。</li>
</ol>2.特征识别
<ol><li>用肉眼观察 红景天 的外观特征:圆柱形和短主根和根茎，棕色表面，带有粉红色图案的膜黄色表皮以及橙红色或勃艮第切片。</li>
	<li>通过气味识别它:靠近鼻子时会散发出芬芳的气味。</li>
	<li>按味辨:取一小块根放入口中，先啜饮后咀嚼，略带苦味，后甜。</li>
</ol>3.药粉中淀粉颗粒的显微鉴定
<ol><li>用刷子去除 红景天 表面的土壤，将其放入40°C的烤箱中，每24小时翻动一次草药。 注意:药材的易碎性被认为是干燥水分的标准。</li>
	<li>用粉剂机将干燥的药材粉化，并用药用的3号筛子过滤药粉（见 材料表）。</li>
	<li>取一个干净的载玻片（见 材料表），用解剖针挖出粉末（见 材料表），并将其均匀地放置在载玻片的三分之一处，在2毫米以内。</li>
	<li>使用玻璃滴管（见 材料表）向粉末中加入一滴去离子水。使用镊子（见 材料表）握住盖玻片的一端（见 材料表），快速接触液面并覆盖粉末。 注意: 使用解剖针轻轻混合水和药粉，以确保样品混合均匀。载玻片、粉末和盖玻片之间不应有气泡。</li>
	<li>打开显微镜（参见 材料表）并将步骤3.4中的载玻片放在平台上以固定它。调整光源和粗焦螺旋以查看粉末。调整细小的旁焦点螺旋，直到清楚地看到组织。切换到40倍物镜并观察淀粉颗粒。 注意:对呈现为单个或多个颗粒的谷物进行淀粉，脐带点呈人字形或裂缝状。</li>
</ol>4.药粉中导管、软木细胞、纤维、木实质细胞、色素团的显微鉴定
<ol><li>取一张干净的载玻片（见 材料表），用解剖针挖出粉末（见 材料表），并将其放置在载玻片的三分之一处。</li>
	<li>使用玻璃滴管（见 材料表）向粉末中加入一滴水合氯醛（见 材料表）。用镊子取载玻片（见 材料表），在酒精灯中加热三次，每次加热1秒。 注意: 加热过程中应避免气泡。液体保持不流动，表明渗透已完成。</li>
	<li>使用玻璃滴管加入一滴甘油（见 材料表）。用镊子握住盖玻片的一端，快速接触液位。</li>
	<li>打开显微镜并将载玻片放在平台上以固定它。调整光源和粗焦螺旋以查看粉末。调整细小的旁焦点螺旋，直到清楚地看到组织。通过切换到 40 倍物镜观察导管、软木细胞、纤维、木实质细胞和颜料块。 注:软木细胞多边形或长多边形，螺旋状血管具有明显的螺旋结构，木质部薄壁组织含有草酸钙砂晶，红色或棕红色色素块。</li>
</ol>5. 红景天薄层色谱（TLC）样品的制备及其对照品
<ol><li>将称重纸放在天平上（见 材料表）并称取 3 克 红景天粉末。</li>
	<li>将粉末放入 100 mL 锥形瓶中（参见 材料表），并用大腹移液管加入 25 mL 甲醇（参见 材料表）。将锥形瓶放入超声波仪器中。将功率设置为 250 W，频率设置为 40 kHz，时间设置为 30 分钟（参见 材料表），然后打开仪器。 注:超声仪器的目的是确保 红景天 粉末完全溶解，而不影响后续薄层色谱实验的结果。</li>
	<li>取出锥形瓶，用流水冲洗外瓶至室温 （RT）。</li>
	<li>用 1 mL 注射器吸出步骤 5.3 中制备的 800 μL 液体。用0.22μm微孔滤膜过滤（参见 材料表），并在色谱样品瓶中收集400μL 红景天 中游样品溶液。</li>
	<li>称取并分别加入 2 mg 水景苷、酪醇和没食子酸（参见 材料表）到 3 个单独的 100 mL 锥形瓶中。用大肚移液管向每个锥形瓶中加入 25 mL 甲醇。</li>
	<li>将锥形瓶放入超声波仪器中，将功率设置为250 W，频率设置为40 kHz，时间设置为30分钟，然后重复步骤5.3（见 材料表）。</li>
	<li>用 1 mL 注射器吸出步骤 5.6 中制备的 800 μL 液体。用0.22μm微孔滤膜过滤（见 材料表），并在相应的色谱样品瓶中收集400μL中游水景苷，酪醇和没食子酸标准溶液。</li>
</ol>6. TLC鉴定
<ol><li>移液器三氯甲烷 （5 mL）、乙酸乙酯 （4 mL）、甲醇 （2 mL） 和甲酸 （0.5 mL）（参见 材料表）。加入双罐层析筒的一侧（见 材料表），摇匀并混合。盖住上气缸盖。</li>
	<li>将没食子酸，水景苷，酪醇标准溶液和 红景天 溶液放在样品架上的位置A1-A4。</li>
	<li>将 5 cm x 10 cm 硅胶片（参见 材料表）放在采样台上。启动自动取样机（见 物料表），打开气控阀。</li>
	<li>打开visionCATS软件（参见材料表）。单击"新建">"新建文件夹（名为红景天样本测试）">"确定"。单击"新方法"（名称 Rhodiola crenulata 样品测试）> OK > ATS 4。</li>
	<li>单击 "完成步骤定义"（Finish Step Definition）。单击 "跟踪分配 "以编辑描述（没食子酸、水景苷、酪醇和 红景天 样品）。</li>
	<li>单击 HPTLC 步骤。设置薄层参数 （5 cm x 10 cm）。选择 应用类型 （波段），设置参数（表1），单击 "确定 "按钮。</li>
	<li>开放式 灌装/冲洗质量。选中 仅填充编程音量。将样品瓶底部液位 （mm） 设置为 0.5，然后单击 OK 按钮。</li>
	<li>单击 "执行方法"。</li>
	<li>单击 "跟踪分配 "按钮，选中 "居中"，然后设置参数（表 2）。</li>
	<li>单击 "继续 "按钮进行自动采样。</li>
	<li>关闭自动取样机和空气阀。从自动取样机上取下硅胶片。</li>
	<li>在步骤6.1中将硅胶片放入双罐层析筒的另一侧，盖上上筒盖，预饱和20分钟。</li>
	<li>用镊子轻轻夹住薄层板的上端，迅速将硅胶片放入显影剂中，盖上上气缸盖。 注意: 当展开的前边缘距离薄层板上端 0.5-1.0 厘米时，取出硅胶片。</li>
	<li>有机溶剂蒸发后，在室温下将显色溶液喷洒到硅胶片表面，以获得显色效果。 注:显色溶液是含有 2% FeCl3 和 1% K3[Fe（CN）6] 的水溶液。</li>
</ol>

红景天月状物的显微表征和基于 TLC 的鉴定

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作者没有什么可透露的。

世界上有90多种红景天植物，其中60%以上的物种分布在我国，常见的有红景天、红景天、红景天和红景天17。《中国药典》（2020年）第一部分记载的红景天是一种生长在高海拔地区的传统藏药。红景天的市场需求每年都在增加，因此确保正确使用来源是确保安全使用的关键。特别是，从现场拣选到简单快速的实验室常规鉴定的标准化不容忽视。据报道，高效液相色谱、质谱和简单序列重复序列可以准确地将红景天与其他红景天物种区分开来，这既费时又复杂，而且成本高昂18,19,20。同时，我们建立了感官识别（E-nose和颜色分析）和HPLC法的多维评价方法，以区分红景天21,22。
每种药材都有其独特的生长环境、微观结构特征和指标成分。该协议提供了一种鉴定 红景天 的综合方法，从现场鉴定到实验室显微镜检查和使用薄层色谱法进行验证。 红景天 主要生长在海拔3000米以上、低温、低氧、高紫外线辐射地区23。 红景天 是一种多汁的草本植物，这是其直观的视觉特征。其粉末呈红褐色，有香味。根据其习性、植物形态、花和叶， 红景天 与该领域的其他红景天物种区分开来。药材显微结果显示，存在淀粉颗粒、木质薄壁细胞（包括草酸钙砂晶）、软木细胞、导管（主要是螺纹导管）和大量红色素。TLC是一种用于分离多组分样品的色谱分离技术，常用于中药材的鉴定。没食子酸、酪醇和水景苷通常被确定为 红景天24 的指示成分。薄层色谱结果表明， 红景天 溶液在对应位置（没食子酸、酪醇和水景苷）处呈现出与对照相同的色斑。它表明 红景天 含有没食子酸、酪醇和水利果苷。结合对生长环境的了解和显微结果，可以初步鉴定 红景天 。
值得注意的是，独特的生长环境决定了在海拔3000 m以下几乎不可能有野生 红景天 。此外，不建议在开花期间收获 红景天 的根和根茎。对于实验室显微镜鉴定，干燥根部和筛选粉末是成功制备显微样品的先决条件。确保载玻片、药粉和盖玻片之间没有气泡也是在显微镜下观察特征成分的关键。对于薄层色谱法，硅胶板的预饱和度、合理的显影剂、样品浓度等是成功分离供试品中不同组分的重要因素。与传统的人工采样相比，该协议的自动采样过程无疑提高了结果的准确性和实验的可重复性。香气和味觉识别的主观性太强，可能导致误判。与高效液相色谱、氢核磁共振和质谱相比，薄层色谱无法定量分析药材中化合物的含量25。尽管DNA条形码在草药鉴定方面具有较高的准确性，但其高昂的价格决定了它在鉴定药材方面并不普遍26。此外，从现场到实验室的鉴定和显微镜检查与本方案中提供的薄层色谱技术相结合，几乎适用于所有药用材料。这是一个廉价、简单、快速的过程，用于识别任何药材。

草药在中国有着悠久的历史和丰富的应用经验，是神农草药经典1中的第一个系统记录。青蒿素应用于疟疾的发现，推动了草药的发展进入一个新的阶段1。利用现代科学技术揭示草药的确切机理，提高了草药的利用率和需求，为其开辟了新的国际市场2,3,4。然而，这导致了一系列负面影响。非专业人士对草药的特性认识模糊，这使得草药的使用面临巨大的安全风险5。
红景天是红景天属植物之一，主要分布于西藏、云南西北部和四川西部（图1）6,7。红景天含有黑景天、酪醇、没食子酸等化合物，具有"活气促血、清脉、镇静哮喘"等功能，用于治疗缺氧相关疾病8,9,10,11。野外调查表明，在海拔4,000-5,600 m的高山距骨带、沟壑斜坡和岩石缝隙中可以找到红景天。它的生长环境寒冷，阳光充足，辐射强烈，属于高寒草甸生态系统。红景天根据生长地形可分布在层状和点状种群中，通过异花授粉进行基因流动。
红景天属的花粉流产、非法挖掘和生态环境的退化使红景天成为濒危物种6,12。鉴于红景天的高药用价值，预计假冒产品将流入市场。本文介绍了红景天的栖息地和一些方便的实验室鉴定方法。首先，我们观察了红景天的生长环境及其药用特性。其次，通过显微镜观察药粉的微观结构;最后一步是关键点。根据红景天在某种物质中的不同吸附或溶解特性，对红景天的代表性成分进行分离和鉴定。基于DNA的药用植物鉴定或代谢组学分析方法复杂且昂贵13.这些基本、方便、经济的方法可以快速识别红景天。

<table><tbody><tr><td>0.22 &amp;mu;m millipore filter</td><td>Millipore</td><td>SLGP033RB</td><td></td></tr><tr><td>Automatic sampling machine</td><td>CAMAG</td><td>ATS 4</td><td></td></tr><tr><td>Chloral hydrate</td><td>Fuzhou Brunei Technology Co., Ltd</td><td>ST1002</td><td></td></tr><tr><td>Chromatographic&amp;nbsp;sample&amp;nbsp;bottles</td><td>Zhejiang ALWSCI Technology Co., Ltd</td><td>C0000008</td><td></td></tr><tr><td>Conical flask</td><td>Sichuan Shubo Co., Ltd</td><td>1121</td><td></td></tr><tr><td>Cover glass</td><td>Citotest Labware Manufacturing Co., Ltd</td><td>10211818c</td><td></td></tr><tr><td>Dissecting needle</td><td>Shanghai Bingyu Fluid Technology Co., Ltd</td><td>BY-5026</td><td></td></tr><tr><td>Electronic balance</td><td>SHIMADZU</td><td>ATX124</td><td></td></tr><tr><td>Ethyl acetate</td><td>Chengdu Kelong Chemical Co., Ltd</td><td>2022120901</td><td></td></tr><tr><td>Formic acid</td><td>Chengdu Kelong Chemical Co., Ltd</td><td>2021110801</td><td></td></tr><tr><td>Gallic acid</td><td>Chengdu Herbpurify Co., Ltd</td><td>M-017</td><td></td></tr><tr><td>Glycerol</td><td>Sinopharm Chemical Reagent Co., Ltd</td><td>10010618</td><td></td></tr><tr><td>High speed&amp;nbsp; crusher</td><td>Beijing Zhongxingweiye Instrument Co., Ltd</td><td>FW-100</td><td></td></tr><tr><td>Methanol</td><td>Chengdu Kelong Chemical Co., Ltd</td><td>20230108</td><td></td></tr><tr><td>Microscope</td><td>Chongqing Oprec Nistrument Co.,&amp;nbsp; Ltd</td><td>B203</td><td></td></tr><tr><td>Microscope slide</td><td>Citotest Labware Manufacturing Co., Ltd</td><td>7105P-G</td><td></td></tr><tr><td>Oven</td><td>Shanghai Yuejin Medical Equipment Co., Ltd</td><td>DHG-8145</td><td></td></tr><tr><td>Pharmacopoeia sieve</td><td>Hangzhou Xingrun sieve factory</td><td>572423281330</td><td></td></tr><tr><td>Pipette</td><td>Changde BKMAM Biotechnology Co., Ltd</td><td>120302008</td><td></td></tr><tr><td>Salidroside</td><td>Chengdu Herbpurify Co., Ltd</td><td>H-040</td><td></td></tr><tr><td>Saturate tank&amp;nbsp;</td><td>Yancheng Liegu Technology Co., Ltd</td><td>10*20 P-1</td><td></td></tr><tr><td>Silica gel plate</td><td>Yantai Jiangyou Silica Gel Development&amp;nbsp; Co., Ltd</td><td>HSG20211227</td><td></td></tr><tr><td>Trichloromethane</td><td>Chengdu Kelong Chemical Co., Ltd</td><td>20221013-1</td><td></td></tr><tr><td>Tweezer&amp;nbsp;</td><td>Changde BKMAM Biotechnology Co., Ltd</td><td>130302027</td><td></td></tr><tr><td>Tyrosol</td><td>Chengdu Herbpurify Co., Ltd</td><td>L-042</td><td></td></tr><tr><td>Ultrasound equipment</td><td>Ningbo Xinyi Ultrasonic Equipment Co., Ltd</td><td>SB-8200DTS</td><td></td></tr><tr><td>Volumetric pipet</td><td>Changde BKMAM Biotechnology Co., Ltd</td><td>120301006</td><td></td></tr></tbody></table>

field collection and laboratory routine identification of rhodiola crenulata

药材鉴定是药品安全的前提和保障。大多数科研人员必然会青睐简单、快速、有效、廉价的草药鉴定过程。 红景天 是一种生长在高海拔地区的传统藏药，主要分布于中国西藏、云南和四川地区。 红景天 具有抗炎、抗缺氧、抗氧化等多种生物活性，具有很大的发展潜力。随着市场需求的增加和资源含量的迅速减少，大量混淆的 红景天 产品一直困扰着人们。因此，该协议引入了在现场鉴定 红景天 的标准过程，并结合常规实验室测试。生境、微观特征和薄层色谱法的结合，无疑将快速、高效、经济地鉴定 红景 天，为藏医药的不断发展和药材的质量控制做出贡献。

该实验方案描述了在田间识别和收集 红景天 。 红景天 倾向于生活在高山距骨带、沟壑斜坡和高海拔的岩石缝隙中。 红景天 的生境、全株、花和叶如 图2所示。 红景天 具有红褐色的根茎（图3A）。药粉的代表性图像如 图3B所示。根据上述实验方案，显微结果可列举如下:1）棕黄色或无色软木细胞，外观大，多边形或长多边形，壁稍厚（图3C）;2）淀粉粒表现为单个或多个粒，脐点呈人字形或裂缝状（图3D）;3）主要是紧密排列的螺旋容器（图3E）;4）呈片状的无色和椭圆形木质部薄壁组织，含有草酸钙砂晶（图3F）;5）形状不规则的棕红色色素块（图3G）。薄层分离结果表明，红 景天 样品（A4）在没食子酸（A1）、水稻苷（A2）、酪醇标准溶液（A3）的色谱图对应位置出现相同颜色的斑点（图3H）。没食子酸、水景苷和酪醇是 红景天的主要和代表性成分。这些结果表明，通过方案中讨论的测试可以初步鉴定 红景天 。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/65947/65947fig01.jpg"> 图1:红景天分布图。 （A） 红景天主要分布于中国、印度、尼泊尔和不丹14.（B）红景天主要分布于中国西藏、青海、四川、云南、贵州等地（软ArcGis 10.6生产）。统计数据来自植物学研究所的网站15,16。<a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/65947/65947fig01large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/65947/65947fig02.jpg"> 图2: 红景天 植物的图片。 （A） 红景天的生物群落。（B） 红景天的特写镜头。（C）红景天整株。（D） 红景天花。（E） 红景天的叶子呈锯齿状。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/65947/65947fig02large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/65947/65947fig03.jpg"> 图3: 红景天的显微特征和薄层色谱图。 （A） 红景天的药用根。（B） 红景天的药用材料功效。（C） 软木细胞。（四）淀粉粒。（E） 螺旋容器。（F）木质部薄壁组织（草酸钙晶体）。（g） 颜料。（H）薄层色谱分离水稻苷（A1）、没食子酸（A2）、酪醇（A3）和 红景 天（A4）。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/65947/65947fig03large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always"><tbody><tr><td>参数</td>
			<td>选择</td>
		</tr><tr><td>应用位置 Y （mm）</td>
			<td>10</td>
		</tr><tr><td>第一轨道位置 （mm）</td>
			<td>10</td>
		</tr><tr><td>轨道距离 （mm）</td>
			<td>10</td>
		</tr><tr><td>应用长度 （mm）</td>
			<td>5</td>
		</tr><tr><td>应用宽度 （mm）</td>
			<td>0.5</td>
		</tr></tbody></table>表1:自动采样位置的参数设置。
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always"><tbody><tr><td>样品瓶 ID</td>
			<td>描述</td>
			<td>体积 （μL）</td>
			<td>位置</td>
			<td>类型</td>
		</tr><tr><td>1</td>
			<td>沙利果苷</td>
			<td>3</td>
			<td>答1</td>
			<td>参考</td>
		</tr><tr><td>11</td>
			<td>没食子酸</td>
			<td>1</td>
			<td>答2</td>
			<td>参考</td>
		</tr><tr><td>12</td>
			<td>酪醇</td>
			<td>2</td>
			<td>答3</td>
			<td>参考</td>
		</tr><tr><td>13</td>
			<td>红景天 样品</td>
			<td>2</td>
			<td>答4</td>
			<td>样本</td>
		</tr></tbody></table>表 2:自动采样顺序的参数设置。

Watch this Scientific Journal Video about Field Collection and Laboratory Routine Identification of Rhodiola crenulata at JoVE.com

作者聚焦:开发 红景天 鉴定方法并研究其资源分布和药理作用

在这里，我们从栖息地、植物形态、药用特性、微观特征和薄层色谱法中描述了红 景天 的鉴定。

红景天的现场采集和实验室常规鉴定

The identification of medicinal materials is the premise and guarantee of drug safety. The majority of scientific researchers are bound to favor the ...

我们的研究范围集中在进行民族医学的资源调查。具体而言，本研究旨在开发一种快速方便的红景天鉴定方法。我们提供的方案提供了一种具有成本效益、直接和快速的方法来识别各种药材。我们的实验室将把未来的研究工作集中在两个主要领域的调查上，即与红景天相关的研究分布和该物种的药理学影响。

field-collection-laboratory-routine-identification-rhodiola

Universidad San Sebastian

Research

JoVE Journal

Medicine

Field Collection and Laboratory Routine Identification of Rhodiola crenulata

559 次观看.  Chengdu University of Traditional Chinese Medicine. 我们的研究范围集中在进行民族医学的资源调查。具体而言，本研究旨在开发一种快速方便的红景天鉴定方法。我们提供的方案提供了一种具有成本效益、直接和快速的方法来识别各种药材。我们的实验室将把未来的研究工作集中在两个主要领域的调查上，即与红景天相关的研究分布和该物种的药理学影响。

视频: 作者聚焦:开发 红景天 鉴定方法并研究其资源分布和药理作用

Colletotrichum fioriniae Development in Water and Chloroform-based Blueberry and Cranberry Floral Extracts

To accurately monitor the phenology of the bloom period and the temporal dynamics of floral chemical cues on fungal fruit rotting pathogens, floral extraction methods and coverslip bioassays were developed utilizing Colletotrichum fioriniae. In blueberry and cranberry, this pathogen is optimally controlled by applying fungicides during the bloom period because of the role flowers play in the initial stages of infection. The protocol detailed here describes how floral extracts (FE) were obtained using water-, chloroform-, and field rainwater-based methods for later use in corresponding glass coverslip bioassays. Each FE served to provide a different set of information: response of C. fioriniae to mobilized floral chemical cues in water (water-based), pathogen response to flower and fruit surface waxes (chloroform-based), and field-based monitoring of collected floral rainwater, moving in vitro observations to an agricultural setting. The FE is broadly described as either water- or chloroform-based, with an appropriate bioassay described to compensate for the inherent differences between these two materials. Rainwater that had run off flowers was collected in unique devices for each crop, alluding to the flexibility and application of this approach for other crop systems. The bioassays are quick, inexpensive, simple, and provide the ability to generate spatiotemporal and site-specific information about the presence of stimulatory floral compounds from various sources. This information will ultimately better inform disease management strategies, as FE decrease the time needed for infection to occur, thus providing insight into changing risks for pathogen infection over the growing season.

Colletotrichum fioriniae Development in Water and Chloroform-based Blueberry and Cranberry Floral Extracts

Here, bioassays designed to monitor the development of a fungal pathogen, Colletotrichum fioriniae, in the presence of blueberry or cranberry floral extracts on glass coverslips are described. Water-, chloroform-, and field rainwater- based floral extraction techniques are detailed as well as insight into how this information can be applied.

Fioriniae水和氯蓝莓和越橘花提取物的研究进展

To accurately monitor the phenology of the bloom period and the temporal dynamics of floral chemical cues on fungal fruit rotting pathogens, floral ...

科研

环境科学

High-throughput Siderophore Screening from Environmental Samples: Plant Tissues, Bulk Soils, and Rhizosphere Soils

Siderophores (low-molecular weight metal chelating compounds) are important in various ecological phenomenon ranging from iron (Fe) biogeochemical cycling in soils, to pathogen competition, plant growth promotion, and cross-kingdom signaling. Furthermore, siderophores are also of commercial interest in bioleaching and bioweathering of metal-bearing minerals and ores. A rapid, cost effective, and robust means of quantitatively assessing siderophore production in complex samples is key to identifying important aspects of the ecological ramifications of siderophore activity, including, novel siderophore producing microbes. The method presented here was developed to assess siderophore activity of in-tact microbiome communities, in environmental samples, such as soil or plant tissues. The samples were homogenized and diluted in a modified M9 medium (without Fe), and enrichment cultures were incubated for 3 days. Siderophore production was assessed in samples at 24, 48, and 72 hours (h) using a novel 96-well microplate CAS (Chrome azurol sulphonate)-Fe agar assay, an adaptation of the traditionally tedious and time-consuming colorimetric method of assessing siderophore activity, performed on individual cultivated microbial isolates. We applied our method to 4 different genotypes/Lines of wheat (Triticum aestivum L.), including Lewjain, Madsen, and PI561725, and PI561727 commonly grown in the inland Pacific Northwest. Siderophore production was clearly impacted by the genotype of wheat, and in the specific types of plant tissues observed. We successfully used our method to rapidly screen for the influence of plant genotype on siderophore production, a key function in terrestrial and aquatic ecosystems. We produced many technical replicates, yielding very reliable statistical differences in soils and within plant tissues. Importantly, the results show the proposed method can be used to rapidly examine siderophore production in complex samples with a high degree of reliability, in a manner that allows communities to be preserved for later work to identify taxa and functional genes.

We present a protocol for rapid screening of environmental samples for siderophore potential contributing to micronutrient bioavailability and turnover in terrestrial systems.

从环境样品中筛选高通量的安德森菌: 植物组织、散装土壤和根际土壤

Siderophores (low-molecular weight metal chelating compounds) are important in various ecological phenomenon ranging from iron (Fe) biogeochemical cycling ...

Comparative Karyotype Analysis of Lycoris aurea Herb Populations Using Fluorochrome Banding and 45S and 5S rDNA-FISH

To understand karyotype variation in eight populations, detailed karyotypes were meticulously established using chromosomal measurements, fluorescence bands, and rDNA FISH signals. The number of 45S rDNA sites varies from one to five pairs per population, with the most common number per karyotype being four pairs. The 45S rDNA locus is predominantly located in the short arms and terminal regions of chromosomes, while the 5S rDNA locus is found mainly in the short arm and the terminal or proximal regions. Populations HBWF, HNXN, HBBD, and HNZX showed a similar distribution of 45S rDNA sites, as did GXTL, HBFC, and SCLS, indicating a close relationship between populations with similar 45S rDNA site distributions. The karyotypes of all studied populations are symmetrical, comprising stable and metastable centromeres or exclusively stable centromeres. Scatter plots of MCA and CVCL effectively distinguish their karyotypic structures. The analysis includes six quantitative parameters (x, 2n, TCL, MCA, CVCL, CVCI). Additionally, the results indicate that PCoA based on these six parameters is a robust method for determining biological karyotype relationships among the eight populations. The chromosome number in Lycoris populations is x = 6-8. Based on the current study and literature, genomic differentiation of these populations is discussed in terms of genome size, heterochromatin, 45S and 5S rDNA sites, and karyotype asymmetry.

This protocol investigates the heterochromatin distribution in eight populations of Lycoris aurea (L&#8242;Her) Herb, using combined PI and DAPI chromosome staining, fluorescence in situ hybridization (FISH) with 18S-5.8-26S rRNA gene regions (45S rDNA regions), and 5S ribosomal DNA probes.

To understand karyotype variation in eight populations, detailed karyotypes were meticulously established using chromosomal measurements, fluorescence ...

遗传学

HPLC Coupled with Chemical Fingerprinting for Multi-Pattern Recognition for Identifying the Authenticity of Clematidis Armandii Caulis

A method for identifying Chinese medicinal materials and their related adulterants was constructed by taking Clematidis Armandii Caulis (Chuanmutong, a universally used traditional Chinese medicine) as an example. Ten batches of genuine Chuanmutong varieties and five batches of related adulterants were analyzed and compared based on the high-performance liquid chromatography (HPLC) fingerprints combined with chemometrics, including cluster analysis (CA), principal component analysis (PCA), and orthogonal partial least-squares discrimination analysis (OPLS-DA). In addition, the content of &#946;-sitosterol was determined. The control chemical fingerprint of Chuanmutong was established, and 12 common peaks were identified. The similarity between the fingerprint of 10 batches of genuine Chuanmutong varieties and the control fingerprint was 0.910-0.989, while the similarity of five batches of adulterants was only 0.133-0.720. Based on the common peaks in the chromatogram, 15 batches of samples were classified into three content levels by PCA, and were aggregated into four categories by CA, achieving a clear distinction between authentic Chuanmutong and adulterants of Chuanmutong. Further, seven differential components that can effectively identify authentic Chuanmutong and adulterants of Chuanmutong were found through OPLS-DA. The &#946;-sitosterol content of 10 batches of genuine Chuanmutong varieties was 97.53-161.56 &#181;g/g, while the &#946;-sitosterol content of the five batches of adulterants varied greatly, among which the &#946;-sitosterol content of Clematis peterae Hand.-Mazz. and Clematis gouriana Roxb. Var. finetii Rehd. et Wils. was significantly lower than that of authentic varieties of Chuanmutong. The HPLC index component content and chemical fingerprint multi-pattern recognition method established in this study provide a new strategy for effectively identifying authentic Chinese medicinal materials and related adulterants.

HPLC Coupled with Chemical Fingerprinting for Multi-Pattern Recognition for Identifying the Authenticity of Clematidis Armandii Caulis

Here, we present a protocol to establish high-performance liquid chromatography (HPLC), coupled with chemical fingerprint multi-pattern recognition, which provides a new strategy for effectively identifying the genuine varieties of Clematidis Armandii Caulis and its adulterants.

高效液相色谱结合化学指纹图谱进行多模式识别，鉴定金线莲真伪

A method for identifying Chinese medicinal materials and their related adulterants was constructed by taking Clematidis Armandii Caulis (Chuanmutong, a ...

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化学

Ultra High-Performance Liquid Chromatography-Mass Spectrometry and Self-established Database Analysis of Chinese Herbal Medicine Components

Chinese herbal medicine is complex and has numerous unknown compounds, making qualitative research crucial. Ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) is the most widely used method in qualitative analysis of compounds. The method includes standardized and programmed protocols for sample pretreatment, MS tune, MS acquisition, and data processing. The sample pretreatments include collection, pulverization, solvent extraction, ultrasound, centrifugation, and filtration. Data post-processing was described in detail and includes data importing, self-established database construction, method establishment, data processing, and other manual operations. The above-ground part of the alpine yarrow herb, Achillea millefolium L., is used to treat inflammation, gastrointestinal disturbances, and pain and its 3-oxa-guaianolides could be useful leads for anti-inflammatory drug development. Three representative compounds in AML were identified, combining TOF-MS with a self-established database. Moreover, the differences from existing literature, liquid-phase parameter optimization, scan mode selection, ion source suitability, collision energy adjustment, isomer screening, method limitation, and possible solutions were discussed. This standardized analysis method is universal and can be applied to identify complex compounds in Chinese herbal medicine.

We describe a general protocol and systematic design that could be applied to separate and recognize complex components in alpine yarrow herb, Achillea millefolium L., a Chinese herbal medicine.

Chinese herbal medicine is complex and has numerous unknown compounds, making qualitative research crucial. Ultra-high-performance liquid chromatography ...

<meta charset="utf8"></head><body>通过 UPLC-TOF 质谱法鉴定 双色 Gynura bicolor 的化学成分

High-resolution Tandem Mass Spectrometry for Studying Chemical Constituents of Gynura bicolor DC

The separation and analysis of the desired chemical components are important subjects for the fundamental research of traditional Chinese medicine (TCM). Ultra-high-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS) has gradually become a leading technology for the identification of TCM ingredients. Gynura bicolor DC. (BFH), a perennial stemless herb used for medicine and food in China has medicinal effects such as clearing heat, moistening the lung, relieving cough, dispersing stasis, and relieving swelling. Polyphenols and flavonoids contain numerous isomers, which hinder the identification of the complex compounds in BFH. This paper presents a systematic protocol for studying chemical constituents of BFH based on solvent extraction and integrated data via UPLC-Q-TOF-MS.
The method described here includes systematic protocols for sample pretreatment, MS calibration, MS acquisition, data processing, and analysis of results. Sample pretreatment includes collection, cleaning, drying, crushing, and extraction. MS calibration consists of multipoint and single-point correction. Data processing includes data importing, method establishment, analysis processing, and result presentation. Representative results of the typical fragmentation pattern of phenolic acids, esters, and glycosides in Gynura bicolor DC. (BFH) are presented in this paper. In addition, organic solvent selection, extraction, data integration, collision energy selection, and method improvement are discussed in detail. This universal protocol can be widely used to identify complex compounds in TCM.

<meta charset="utf8"></head><body>用于高分辨率串联质谱分析的 Gynura bicolor DC 样品的制备

We describe a general protocol that integrates high-resolution mass spectrometry analysis and molecular docking in traditional Chinese medicine research.

用于研究双色 DC 化学成分的高分辨率串联质谱法

The separation and analysis of the desired chemical components are important subjects for the fundamental research of traditional Chinese medicine (TCM). ...

An In Vitro Dissolution Determination of Multi-Index Components in Tibetan Medicine Rhodiola Granules

The composition of the Tibetan medicine Rhodiola granules (RG) is complex, and the overall quality of RG is difficult to determine. Therefore, establishing a method to determine the multi-component in vitro dissolution of RG is of great significance for quality control. This study uses the second paddle method of the fourth general rule 0931 from the Chinese Pharmacopoeia (2020 edition), compliant with apparatus 2 of the United States Pharmacopeia (USP). The dissolution apparatus was set to a rotation speed of 100 rpm with ultrapure water as the dissolution medium. A sample volume of 1 mL was collected at each timepoint. Furthermore, the cumulative dissolution of gallic acid, salidroside, and ethyl gallic acid in RG at different time points was determined by high-performance liquid chromatography (HPLC). Finally, the dissolution curves were drawn, and the curves were fitted to the GompertzMod, the Gompertz, the Logistic, and the Weibull equations. The results showed that the cumulative dissolution of gallic acid in RG was over 80% at 1 min, the cumulative dissolution of salidroside and ethyl gallic acid was over 65% at 5 min, and the cumulative dissolution of each index component decreased after 30 min. The curve fitting demonstrated that the GompertzMod equation was the best-fitting model for each index component of RG. In conclusion, the dissolution test method described in this protocol is simple, accurate, and reliable. It can characterize the dissolution behavior of the index components in RG in vitro, which provides a methodological reference for quality control of RG and quality evaluation of other ethnic compounds.

An In Vitro Dissolution Determination of Multi-Index Components in Tibetan Medicine Rhodiola Granules

Here, we test the dissolution of Rhodiola granules (RG) in vitro, draw dissolution curves of salidroside, gallic acid, and ethyl gallate in ultrapure water, and fit the curves to different mathematical models. This protocol provides information and guidance for in vivo bioequivalence and in vivo-in vitro correlation studies of RG.

藏药红景天颗粒中多指标组分的 体外 溶出度测定

The composition of the Tibetan medicine Rhodiola granules (RG) is complex, and the overall quality of RG is difficult to determine. Therefore, ...

医学

A Plate Competition Assay As a Quick Preliminary Assessment of Disease Suppression

The goal was to develop and optimize a simple, affordable, and effective bioassay to detect disease suppressive ability of a specific compost against soilborne fungus Rhizoctonia solani. R. solani is a pathogen of a wide range of plant hosts worldwide. The fungus survives in soils as a saprophyte and grows rapidly on simple water agar media. The plate assay is a rapid method to compare composts for their ability to slow the growth of R. solani. The assay also correlates well with suppression of other soilborne fungal pathogens that survive as saprophytes in soils such as Alternaria early blights, Fusarium wilt, Phytophthora root rot, and Pythium root rot.

Presented is a protocol for a plate competition assay to identify whether a specific compost is likely to contain bacteria and fungi that suppress growth of Rhizoctonia solani.

一种快速的疾病抑制初步评估的板块竞争分析法

The goal was to develop and optimize a simple, affordable, and effective bioassay to detect disease suppressive ability of a specific compost against ...

红景天的现场采集和实验室常规鉴定

副本

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Fioriniae水和氯蓝莓和越橘花提取物的研究进展

从环境样品中筛选高通量的安德森菌: 植物组织、散装土壤和根际土壤

Comparative Karyotype Analysis of Lycoris aurea Herb Populations Using Fluorochrome Banding and 45S and 5S rDNA-FISH

高效液相色谱结合化学指纹图谱进行多模式识别，鉴定金线莲真伪

Ultra High-Performance Liquid Chromatography-Mass Spectrometry and Self-established Database Analysis of Chinese Herbal Medicine Components

用于研究双色 DC 化学成分的高分辨率串联质谱法

用于研究双色 DC 化学成分的高分辨率串联质谱法

用于研究双色 DC 化学成分的高分辨率串联质谱法

藏药红景天颗粒中多指标组分的体外溶出度测定

一种快速的疾病抑制初步评估的板块竞争分析法

红景天的现场采集和实验室常规鉴定

副本

摘要

探索更多视频

此视频中的章节

Fioriniae水和氯蓝莓和越橘花提取物的研究进展

从环境样品中筛选高通量的安德森菌: 植物组织、散装土壤和根际土壤

Comparative Karyotype Analysis of Lycoris aurea Herb Populations Using Fluorochrome Banding and 45S and 5S rDNA-FISH

高效液相色谱结合化学指纹图谱进行多模式识别，鉴定金线莲真伪

Ultra High-Performance Liquid Chromatography-Mass Spectrometry and Self-established Database Analysis of Chinese Herbal Medicine Components

用于研究双色 DC 化学成分的高分辨率串联质谱法

用于研究双色 DC 化学成分的高分辨率串联质谱法

用于研究双色 DC 化学成分的高分辨率串联质谱法

藏药红景天颗粒中多指标组分的 体外 溶出度测定

一种快速的疾病抑制初步评估的板块竞争分析法

藏药红景天颗粒中多指标组分的体外溶出度测定