Name: thin-layer chromatographic (tlc) separations and bioassays of plant extracts to identify antimicrobial compounds
Uploaded: 2014-03-27T22:00:00
Description: Watch this Scientific Journal Video about Thin-layer Chromatographic TLC Separations and Bioassays of Plant Extracts to Identify Antimicrobial Compounds at JoVE.com

我们感谢已故的博士。规范泰勒，系植物与土壤科学在肯塔基大学，允许我们使用的样品从他的红三叶草地块为这项研究。该项目是由农业部美国农业部。

1。植物提取物的制备<ol><li>见卡根和Flythe 10从红车轴草提取CV酚类化合物。 Kenland。 </li><li>要提取其他化合物在其它植物中，检查（很多）里描述的植物化学分析文献植物或代谢物的具体提取方法，或者寻找协议，如那些KHURRAM 等。7,8的其中分离许多化合物具有广泛极性范围。 </li></ol> 2。薄层板的制备<ol><li>清洁TLC板通过在一个或多个极性，中性...

<ol>
	<li>Stahl, E., Ashworth, M. R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Thin-layer chromatography. , (1969).</li><li>Marston, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thin-layer+chromatography+with+biological+detection+in+phytochemistry.">Thin-layer chromatography with biological detection in phytochemistry.</a> J. Chromatogr. A. 1218 (19), 2676-2683 .</li><li>Homans, A. L., Fuchs, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+bioautography+on+thin-layer+chromatograms+as+a+method+for+detecting+fungitoxic+substances.">Direct bioautography on thin-layer chromatograms as a method for detecting fungitoxic substances.</a> J. Chromatogr. 51, 327-329 .</li><li>Lund, B. M., Lyon, G. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detection+of+inhibitors+of+Erwinia+carotovora+and+E.+herbicola+on+thin-layer+chromatograms.">Detection of inhibitors of Erwinia carotovora and E. herbicola on thin-layer chromatograms.</a> J. Chromatogr. 110, 193-196 (1975).</li><li>Betina, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioautography+in+paper+and+thin-layer+chromatography+and+its+scope+in+the+antibiotic+field.">Bioautography in paper and thin-layer chromatography and its scope in the antibiotic field.</a> J. Chromatogr. A. 78, 41-51 (1973).</li><li>Weltzien, H. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ein+biologischer+Test+f&#252;r+fungizide+Substanzen+auf+dem+Papierchromatogramm.">Ein biologischer Test f&#252;r fungizide Substanzen auf dem Papierchromatogramm.</a> Naturwissenschaften. 45, 288-289 (1958).</li><li>Khurram, M., Khan, A. M., Hameed, A., Abbas, N., Quayum, A., Inayat, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antibacterial+activities+of+Dodonaea+viscosa+using+contact+bioautography+technique.">Antibacterial activities of Dodonaea viscosa using contact bioautography technique.</a> Molecules. 14 (3), 1332-1341 (2009).</li><li>Khurram, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+anticandidal+potential+of+Quercus+baloot+Griff.+using+contact+bioautography+technique.">Evaluation of anticandidal potential of Quercus baloot Griff. using contact bioautography technique.</a> Afr. J. Pharm. Pharmacol. 5 (12), 1538-1542 (2012).</li><li>Robinson, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Organic Constituents of Higher Plants. , (1963).</li><li>Kagan, I. A., Flythe, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+affecting+the+separation+and+bioactivity+of+red+clover+(Trifolium+pratense)+extracts+assayed+against+Clostridium+sticklandii,+a+ruminal+hyper+ammonia-producing+bacterium.">Factors affecting the separation and bioactivity of red clover (Trifolium pratense) extracts assayed against Clostridium sticklandii, a ruminal hyper ammonia-producing bacterium.</a> Nat. Prod. Commun. 7 (12), 1605-1608 (2012).</li><li>Mattila, P., Kumpulainen, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+free+and+total+phenolic+acids+in+plant-derived+foods+by+HPLC+with+diode-array+detection.">Determination of free and total phenolic acids in plant-derived foods by HPLC with diode-array detection.</a> J. Agric. Food Chem. 50 (13), 3660-3667 (2002).</li><li>Hamburger, M. O., Cordell, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+direct+bioautographic+TLC+assay+for+compounds+possessing+antibacterial+activity.">A direct bioautographic TLC assay for compounds possessing antibacterial activity.</a> J. Nat. Prod. 50 (1), 19-22 (1987).</li><li>Flythe, M., Kagan, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antimicrobial+effect+of+red+clover+(Trifolium+pratense)+phenolic+extract+on+the+ruminal+hyper+ammonia-producing+bacterium,+Clostridium+sticklandii.">Antimicrobial effect of red clover (Trifolium pratense) phenolic extract on the ruminal hyper ammonia-producing bacterium, Clostridium sticklandii.</a> Curr. Microbiol. 61, 125-131 .</li><li>Rahalison, L., Hamburger, M., Hostettmann, K., Monod, M., Frenk, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+bioautographic+agar+overlay+method+for+the+detection+of+antifungal+compounds+from+higher+plants.">A bioautographic agar overlay method for the detection of antifungal compounds from higher plants.</a> Phytochem. Anal. 2 (5), 199-203 (1991).</li><li>Chomnawang, M. T., Trinapakul, C., Gritsanapan, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+antigonococcal+activity+of+Coscinium+fenestratum+stem+extract.">In vitro antigonococcal activity of Coscinium fenestratum stem extract.</a> J. Ethnopharmacol. 122, 445-449 (2009).</li><li>Stahl, E., Kaldewey, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spurenanalyse+physiologisch+aktiver,+einfacher+Indolderivate.">Spurenanalyse physiologisch aktiver, einfacher Indolderivate.</a> Hoppe-Seyler&#8217;s Z. Physiol. Chem. 323, 182-191 .</li><li>Kagan, I. A., Hammerschmidt, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Arabidopsis+ecotype+variability+in+camalexin+production+and+reaction+to+infection+by+Alternaria+brassicicola.">Arabidopsis ecotype variability in camalexin production and reaction to infection by Alternaria brassicicola.</a> J. Chem. Ecol. 28 (11), 2121-2140 (2002).</li><li>Kline, R. M., Golab, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+technique+in+developing+thin-layer+bioautographs.">A simple technique in developing thin-layer bioautographs.</a> J. Chromatogr. 18, 409-411 (1965).</li><li>Wedge, D. E., Nagle, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+2D-TLC+bioautography+method+for+the+discovery+of+novel+antifungal+agents+to+control+plant+pathogens.">A new 2D-TLC bioautography method for the discovery of novel antifungal agents to control plant pathogens.</a> J. Nat. Prod. 63 (8), 1050-1054 (2000).</li><li>Beck, A. B., Knox, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+acylated+isoflavone+glycosides+from+subterranean+clover+and+red+clover.">The acylated isoflavone glycosides from subterranean clover and red clover.</a> Aust. J. Chem. 24 (7), 1509-1518 (1971).</li><li>Kahn, R. A., Bak, S., Svendsen, I., Halkier, B. A., M&#248;ller, B. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+reconstitution+of+cytochrome+P450ox+and+in+vitro+reconstitution+of+the+entire+biosynthetic+pathway+of+the+cyanogenic+glucoside+dhurrin+from+sorghum.">Isolation and reconstitution of cytochrome P450ox and in vitro reconstitution of the entire biosynthetic pathway of the cyanogenic glucoside dhurrin from sorghum.</a> Plant Physiol. 115 (4), (1997).</li><li>Peterson, C. A., Edgington, L. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+estimation+of+the+fungicide+benomyl+using+a+bioautograph+technique.">Quantitative estimation of the fungicide benomyl using a bioautograph technique.</a> J. Agr. Food Chem. 17 (4), 898-899 (1969).</li></ol>

这个协议描述了一种用于提取物中分离到的化合物的子集，并通过接触生物自显影法测定的那些子集的简单方法。该方法非常类似于1使用Chomnawang 等人 15来筛选植物代谢产物抑制到淋病的细菌。生物自显影来屏幕抗菌植物化合物的类型取决于许多因素，包括微生物检验，实验室的设置，而该人（次）进行生物测定的喜好。联系生物自显影，在这项研究中所使用的方法，已经被批评?...

在植物中测定抗微生物化合物需要分离的植物提取物的组分，露出试验微生物的那些部件，并确定由任何所述化合物的微生物的生长是否受到抑制。分离由纸或薄层色谱法（PC或TLC）是方便的，因为很多化合物可在平面表面上进行分离。分离是基于极性，与一些化合物的结合紧密的吸附剂（纤维素在PC机的情况下，以及各种在薄层的情况下，吸附剂）和迁移小于其他1， 图1提供的相对位置的一个例子在硅胶TLC板上分离后极性和非极性的酚类化合物。 <img alt="图1" fo:content-width="5in" fo:src="/files/ftp_upload/51411/51411fig1highres.jpg" src="/files/ftp_upload/51411/51411fig1.jpg" /> 图1。示出在硅胶薄层色谱（TLC）板分离后不同极性的化合物的分布。红三叶草（ 红三叶 L.）的酚类化合物被用作一个例子。极性化合物，如clovamide，有一种极性吸附剂像二氧化硅有很强的亲和力，并保持靠近原点（OR），而极性较小的化合物，如靠近溶剂前沿的3异黄酮（SF），分区更容易进入溶剂（除非水，酸，碱或包含它们比二氧化硅极性较小）和迁移更远了板。 的提取物在TLC板上分离后，试验微生物可以暴露于板上的所有化合物中，从而加速的提取物2的活性成分的鉴定。如果是真菌或细菌培养物暴露于色谱，会发生微生物生长无处不在，除了以上与生长抑制剂领域Ÿ化合物。抑制区则可以通过观察菌丝生长和生长无区域之间的对比，如果真菌已经应用3或者通过与该改变颜色由活细胞4降低或水解时的化合物喷雾可视化。虽然使用纸或薄层色谱的抗菌试验首先被施加到抗生素5和杀菌剂3,6，植物提取物，现在经常筛选出抗微生物化合物用这种方法，通常被称为生物自显影。本文中所描述的协议适用于薄层色谱生物自显影。 TLC被广泛使用，因为它是相对快速的，并且可以在不同的吸附剂（ 如硅胶，淀粉，氧化铝），以及提供良好的分辨率和灵敏度1进行。 植物提取物可用于TLC以多种方式来制备。常用的方法包括提取植物材料的ALCohol -水的混合物，如80％乙醇7,8，可能与另外的酸或碱9。以下在这些溶剂，其中含有一些水，并且可能是酸性或碱性的提取，提取物，必须进行浓缩，以便它们可以应用到TLC板在最小的体积。的醇-水提取物中的浓度可以通过与水不混溶的有机溶剂的8或与此类溶剂，如乙酸乙酯-乙醚（1:1，V / V）10,11的混合物隔开来实现。异植物代谢产物提取到不同的有机溶剂，这取决于它们的极性。以确保植物有机的酸或碱都在此阶段萃取到有机溶剂中，醇 - 水提取物的pH可以升高或降低与水溶性酸或碱解离的分析物转化为它们的nondissociated的形式，然后将它们溶于中性有机溶剂9。然后将有机相可以通过电子邮件在减压或在氮气氛下vaporated并调整为薄层所需的音量。该提取物的pH不太可能是致命的，因为分析物的分割成中性溶剂，小终体积，并在TLC板上分离前的提取物蒸发生物测定微生物。 这两种真菌和细菌被用作试验的微生物在植物生物自显影提取2。一些真菌，如黑星病的孢子发芽在TLC板（除了与抑制性化合物的区域），如果在一个营养液喷洒到板并孵育在潮湿的环境中几天3。 C.暗菌丝体星病的noninhibitory区提供了一个鲜明的对比，免费菌丝生长的区域。虽然细菌已被施加到薄层色谱（TLC）板以相同的方式4,12，细菌也被倾倒在TLC板面在琼脂覆盖13,14。酵母，如白色念珠菌 ，可在琼脂覆盖层被应用，以及14。此外，TLC板可以向下放置脸上琼脂上接种细菌10,15或酵母8，称为接触性生物自显影2的方法。 我们描述了一种接触生物自显影筛选抗菌酚类化合物从红三叶草（ 红三叶品种。Kenland）。该测试微生物是梭菌sticklandii，一个超瘤胃氨产生菌（HAB）和专性厌氧菌。尽管所使用的分离不能解决提取物的所有组成部分，它们有助于确定抗菌活性的区域，从而缩小可能的抗菌化合物的游泳池。该协议采用标准程序TLC 1。该协议还介绍了一些需要培养obli技术门厌氧菌对这样的测定，接触生物自显影15和可视化方法的具有四唑鎓盐，其染色活细胞2,4的 ​​使用。

<table border="0" cellpadding="0" cellspacing="0" width="739">
	<tbody>
		<tr>
			<td>Silica F254 TLC plates, aluminum-backed, 0.2 mm thickness, 20 &#215; 20 cm</td>
			<td>EMD Chemicals&#160;</td>
			<td>5554/7</td>
			<td>These plates are coated with silica that contains an indicator fluorescing at 254 nm.&#160; Compounds absorbing at that wavelength appear dark on a fluorescent green background.&#160; Alternative sources include Analtech, Selecto Scientific, Fluka.&#160; Adsorbents other than silica may be needed.&#160; Plastic-backed plates may be suitable, depending on the solvents to be used.&#160;&#160;</td>
		</tr>
		<tr>
			<td>Sharp, heavy-duty scissors&#160;</td>
			<td>any sewing supply company</td>
			<td>similar to Fiskars&#160; 175800-1002</td>
			<td>For cutting TLC plates.&#160; A paper cutter with a sharp blade can be used as well.&#160; Do not inhale silica dust.</td>
		</tr>
		<tr>
			<td>Drying oven at 100 &#176;C (mechanical convection)</td>
			<td>Thermo Scientific</td>
			<td>PR305225M</td>
			<td>Quincy Lab, Inc, Chicago, IL (www.quincylab.com); Cascade Technical Sciences, Hillsboro, OR (www.cascadetek.com)</td>
		</tr>
		<tr>
			<td>TLC chamber</td>
			<td>Kimble Chase&#160;</td>
			<td>416180-0000</td>
			<td>Alternative sources:&#160; Aldrich. Pyrex beakers or preserving jars can be used for small plates (i.e. 5 &#215; 10 cm).&#160; Cover with aluminum foil (jar lids may contain material extractable by solvent vapors).</td>
		</tr>
		<tr>
			<td>50-&#181;L syringe with flat needle tip</td>
			<td>Hamilton</td>
			<td>80965</td>
			<td>For loading amounts of standard or sample exceeding 5-10 &#181;L.&#160; Alternative sources are equivalent.</td>
		</tr>
		<tr>
			<td>micropipets</td>
			<td>Drummond</td>
			<td>2-000-001</td>
			<td>For loading small amounts of standards or samples.&#160; Alternative sources:&#160; VWR.&#160; Also, Pasteur pipets can be stretched to a thinner diameter with a butane torch.&#160;&#160;</td>
		</tr>
		<tr>
			<td>Filter paper (#1 grade)</td>
			<td>Whatman</td>
			<td>1001 917</td>
			<td>Serves as a chamber wick.&#160; Other grades of filter paper are OK.&#160; This size can be trimmed for the chambers holding 20 &#215; 20 cm plates.&#160;&#160;&#160;&#160;</td>
		</tr>
		<tr>
			<td>Beaker tongs</td>
			<td>Fisher Scientific</td>
			<td>15-186</td>
			<td>For putting plates in and out of a large TLC chamber.&#160; Alternate sources: VWR&#160;</td>
		</tr>
		<tr>
			<td>Flat-edge forceps&#160;</td>
			<td>Fisher Scientific</td>
			<td>10-275</td>
			<td>For putting plates in and out of a small chamber.&#160;&#160; Alternate sources: VWR&#160;</td>
		</tr>
		<tr>
			<td>Small portable UV lamp with 4-Watt or 6-Watt bulbs for short- and long-wave UV light illumination (254 and 365 nm, respectively)</td>
			<td>Ultraviolet Products&#160;</td>
			<td>95-0271-01</td>
			<td>Alternate sources: Spectronics Corporation (www.spectroline.net)</td>
		</tr>
		<tr>
			<td>Viewing cabinet for use with hand-held UV lamp</td>
			<td>Ultraviolet Products&#160;</td>
			<td>Chromato-Vue C-10E</td>
			<td>UV-active bands are more easily circled if plates can be set in here.&#160; Alternate sources: Spectronics Corporation.&#160;</td>
		</tr>
		<tr>
			<td>Photodocumentation system with overhead UV lamp and visible lamp</td>
			<td>Kodak&#160;</td>
			<td>Gel Logic 200</td>
			<td>&#160;Alternate sources: Ultraviolet Products (www.uvp.com).&#160; See protocol for homemade alternative.</td>
		</tr>
		<tr>
			<td>Anaerobic Chamber, Type A, Vinyl</td>
			<td>Coy&#160;</td>
			<td>7150000</td>
			<td>This chamber is appropriate for anaerobic bacteria, like Clostridium sticklandii, as described. &#160;However, growth conditions must be tailored to organism used in the assay.&#160; A biosafety cabinet and other precautions should be taken if pathogenic organisms are used. Alternate sources: Anaerobe Systems, BioRad, Plas Labs, others&#160;</td>
		</tr>
		<tr>
			<td>Tetrazolium red</td>
			<td>Sigma-Aldrich</td>
			<td>T8877</td>
			<td>Alternate sources: MP Biomedicals, Santa Cruz Biotechnology, Alfa Aesar</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td>Ingredients for HAB media</td>
		</tr>
		<tr>
			<td>Pyridoxamine &#183; 2 HCl</td>
			<td>Sigma-Aldrich</td>
			<td>P9380</td>
			<td>For this and for all the other reagents in this table, alternative sources are equivalent.</td>
		</tr>
		<tr>
			<td>Riboflavin</td>
			<td>Sigma-Aldrich</td>
			<td>R4500</td>
			<td></td>
		</tr>
		<tr>
			<td>Thiamine HCl</td>
			<td>Sigma-Aldrich</td>
			<td>T3902</td>
			<td></td>
		</tr>
		<tr>
			<td>Nicotinamide</td>
			<td>Sigma-Aldrich</td>
			<td>N3376</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcium D-Pantothenate</td>
			<td>Sigma-Aldrich</td>
			<td>C8731</td>
			<td></td>
		</tr>
		<tr>
			<td>Lipoic Acid&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>T5625</td>
			<td></td>
		</tr>
		<tr>
			<td>p-Aminobenzoic acid&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>A9878</td>
			<td></td>
		</tr>
		<tr>
			<td>Folic acid</td>
			<td>Sigma-Aldrich</td>
			<td>F8798</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotin</td>
			<td>Sigma-Aldrich</td>
			<td>B4639</td>
			<td></td>
		</tr>
		<tr>
			<td>Cobalamine&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>C3607</td>
			<td></td>
		</tr>
		<tr>
			<td>Pyridoxal HCl</td>
			<td>Sigma-Aldrich</td>
			<td>P9130</td>
			<td></td>
		</tr>
		<tr>
			<td>Pyridoxine</td>
			<td>Sigma-Aldrich</td>
			<td>P5669</td>
			<td></td>
		</tr>
		<tr>
			<td>EDTA</td>
			<td>Sigma-Aldrich</td>
			<td>&#160;E6758</td>
			<td></td>
		</tr>
		<tr>
			<td>Iron sulfate &#183; 7 H2O</td>
			<td>Sigma-Aldrich</td>
			<td>&#160;F8263</td>
			<td></td>
		</tr>
		<tr>
			<td>Zinc sulfate &#183; 7 H2O</td>
			<td>Sigma-Aldrich</td>
			<td>Z0251</td>
			<td></td>
		</tr>
		<tr>
			<td>Manganese chloride &#183; 4 H2O</td>
			<td>Sigma-Aldrich</td>
			<td>M8054</td>
			<td></td>
		</tr>
		<tr>
			<td>Boric acid</td>
			<td>Sigma-Aldrich</td>
			<td>B6768</td>
			<td></td>
		</tr>
		<tr>
			<td>Cobalt chloride &#183; 6 H2O</td>
			<td>Sigma-Aldrich</td>
			<td>&#160;C8661</td>
			<td></td>
		</tr>
		<tr>
			<td>Copper chloride &#183; 2 H2O</td>
			<td>Sigma-Aldrich</td>
			<td>459097</td>
			<td></td>
		</tr>
		<tr>
			<td>Nickel chloride &#183; 6 H2O</td>
			<td>Sigma-Aldrich</td>
			<td>203866</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium molybdate &#183; 2 H2O</td>
			<td>Sigma-Aldrich</td>
			<td>331058</td>
			<td></td>
		</tr>
		<tr>
			<td>&#160;Trypticase (Pancreatic digest of casein)</td>
			<td>Thermo Fisher</td>
			<td>B11921</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium phosphate monobasic anhydrous</td>
			<td>Thermo Fisher</td>
			<td>P284</td>
			<td></td>
		</tr>
		<tr>
			<td>sodium carbonate &#183; H2O&#160;</td>
			<td>Thermo Fisher</td>
			<td>S636</td>
			<td></td>
		</tr>
		<tr>
			<td>Agar</td>
			<td>Thermo Fisher</td>
			<td>50841063</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium sulfate &#183; 6 H2O</td>
			<td>Thermo Fisher</td>
			<td>7791-18-6</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcium chloride &#183; 2 H2O</td>
			<td>Thermo Fisher</td>
			<td>BP510</td>
			<td></td>
		</tr>
		<tr>
			<td>Cysteine HCl</td>
			<td>Thermo Fisher</td>
			<td>19464780</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium phosphate dibasic anhydrous</td>
			<td>Thermo Fisher</td>
			<td>P290</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium chloride</td>
			<td>Thermo Fisher</td>
			<td>BP358</td>
			<td></td>
		</tr>
	</tbody>
</table>

thin-layer chromatographic (tlc) separations and bioassays of plant extracts to identify antimicrobial compounds

对植物的抗微生物化合物的常用屏幕包括由纸或薄层色谱法（PC或TLC）分离的植物提取物，露出色谱微生物悬浮液（ 例如，在液体培养基真菌或细菌或琼脂），允许时间为微生物生长中的潮湿环境中，并且可视区与无微生物生长。该筛选方法，被称为生物自显影的效果，取决于色谱分离的质量和用微生物培养条件下所采取的照顾。本文介绍了一种新型的应用氨基酸发酵菌的TLC和接触生物自显影技术的标准协议。将提取物上分离柔性（铝背）硅胶TLC板，并频带下的紫外线（UV）光可视化。区域被切出并孵育面朝下于琼脂接种试验微生物。抑制带被染色的琼脂平板可视化s的四氮唑红。该方法适用于红三叶草（ 红三叶品种。Kenland）分离酚类化合物及其筛查对梭状芽孢杆菌sticklandii，一个超氨产生菌（HAB）是原产于牛瘤胃活动。在TLC的方法适用于许多类型的植物提取物和其它的细菌菌种（好氧或厌氧），以及真菌的，可以作为测试生物体，如果培养条件进行修改，以适应植物的生长要求。

红三叶草（ 红三叶品种。Kenland）提取物，含有酚类化合物，有代表性的二氧化硅薄层色谱分离见图2。红车轴草提取物中乙酸乙酯-正己烷分离（9:1，V / V），超过850厘米，导致五阶，一是从原点不完全解析（ 图2A）。然而， 图2B表明，大约两倍多的条带，表明当红三叶草提取物（来自同一品种，但在一个单独的情节在同一农场种植）不同的样本中分离?...

Watch this Scientific Journal Video about Thin-layer Chromatographic TLC Separations and Bioassays of Plant Extracts to Identify Antimicrobial Compounds at JoVE.com

Thin-layer Chromatographic TLC Separations and Bioassays of Plant Extracts to Identify Antimicrobial Compounds

薄层色谱（TLC）分离和植物提取物的生物测定识别抗菌化合物

方法描述为薄层色谱（TLC）分离植物提取物和生物自显影接触，以确定抗菌代谢产物。该方法适用于红三叶草酚类化合物抑制超氨生产菌（HAB），原产于牛瘤胃的筛选。

A common screen for plant antimicrobial compounds consists of separating plant extracts by paper or thin-layer chromatography (PC or TLC), exposing the ...

Research

JoVE Journal

Chemistry

Thin-layer Chromatographic (TLC) Separations and Bioassays of Plant Extracts to Identify Antimicrobial Compounds

Profiling the Triacylglyceride Contents in Bat Integumentary Lipids by Preparative Thin Layer Chromatography and MALDI-TOF Mass Spectrometry

用制备薄层色谱和MALDI-TOF质谱分析的三酸甘油酯含量的蝙蝠外皮血脂

The mammalian integument includes sebaceous glands that secrete an oily material onto the skin surface. Sebum production is part of the innate immune ...

科研

化学

Methods to Identify the NMR Resonances of the 13C-Dimethyl N-terminal Amine on Reductively Methylated Proteins

Methods to Identify the NMR Resonances of the 13C-Dimethyl N-terminal Amine on Reductively Methylated Proteins

Nuclear magnetic resonance (NMR) spectroscopy is a proven technique for protein structure and dynamic studies. To study proteins with NMR, stable magnetic ...

Analysis of Volatile and Oxidation Sensitive Compounds Using a Cold Inlet System and Electron Impact Mass Spectrometry

挥发和氧化敏感的化合物使用冷进气系统和电子轰击质谱分析

This video presents a protocol for the mass spectrometrical analysis of volatile and oxidation sensitive compounds using electron impact ionization. The ...

Sequencing of Plant Wall Heteroxylans Using Enzymic, Chemical (Methylation) and Physical (Mass Spectrometry, Nuclear Magnetic Resonance) Techniques

Sequencing of Plant Wall Heteroxylans Using Enzymic, Chemical Methylation and Physical Mass Spectrometry, Nuclear Magnetic Resonance Techniques

使用酶法，化学（甲基化）和物理（质谱，核磁共振）技术，植物墙Heteroxylans测序

This protocol describes the specific techniques used for the characterization of reducing end (RE) and internal region glycosyl sequence(s) of ...

On-line Analysis of Nitrogen Containing Compounds in Complex Hydrocarbon Matrixes

在线含硫化合物在复杂油气矩阵氮分析

The shift to heavy crude oils and the use of alternative fossil resources such as shale oil are a challenge for the petrochemical industry. The ...

Using Capillary Electrophoresis to Quantify Organic Acids from Plant Tissue: A Test Case Examining Coffea arabica Seeds

Using Capillary Electrophoresis to Quantify Organic Acids from Plant Tissue: A Test Case Examining Coffea arabica Seeds

利用毛细管电泳从植物组织量化有机酸：一个测试用例检查<em&gt;小粒种咖啡</em&gt;种子

Carboxylic acids are organic acids containing one or more terminal carboxyl (COOH) functional groups. Short chain carboxylic acids (SCCAs; carboxylic ...

Synthesis of Plant Phenol-derived Polymeric Dyes for Direct or Mordant-based Hair Dyeing

对于直接或媒介为基础的头发染色厂苯酚衍生的高分子染料的合成

Effective hair dyeing through in situ incubation of keratin hair with the products of fungal laccase-catalyzed polymerization of plant phenols has been ...

Raman and IR Spectroelectrochemical Methods as Tools to Analyze Conjugated Organic Compounds

拉曼光谱和红外光谱电化学法作为分析共轭有机化合物的工具

In the presented work, two spectroelectrochemical techniques are discussed as tools for the analysis of the structural changes occurring in the molecule ...

Using Cyclic Voltammetry, UV-Vis-NIR, and EPR Spectroelectrochemistry to Analyze Organic Compounds

利用循环伏安法、紫外-近红外光谱和 EPR 光谱电化学对有机化合物进行分析

Cyclic voltammetry (CV) is a technique used in the analysis of organic compounds. When this technique is combined with electron paramagnetic resonance ...

Electrophoretic Crystallization of Ultrathin High-performance Metal-organic Framework Membranes

超薄高性能金属-有机骨架膜的电泳结晶

We report the synthesis of thin, highly intergrown, polycrystalline metal-organic framework (MOF) membranes on a wide range of unmodified porous and ...