作者要感谢汤姆帕特森和陶氏 AgroSciences 的分析技术小组使用他们的实验室设施和他们的导师。这项工作的资金部分是通过凯洛格公司和陶氏 AgroSciences 的礼物, 并通过美国农业部舱口赠款, ILLU-802-354 奖。学生支持 CJBW 是由伊利诺伊州杰出的奖学金和威廉 B 和南茜·刘汉铨的作物科学奖学金。

1. 生产煮熟的面粉
<ol><li>在电加热板上放置一个15升罐装压力锅。</li>
	<li>将自来水1升放入罐装压力锅, 加热至100摄氏度。</li>
	<li>当水加热时, 在1夸脱罐装罐中放置100克工业剥落粒或剥落砂砾材料 (12% 含水量, 湿基)7的样品。 注: 本研究的代表性结果是基于威严 macke等的剥落砂砾材料。9采用 Rausch等的实验室尺度干铣工艺规程。7
</li>
	<li>添加糖盐溶液, 由200毫升蒸馏水、2克盐、6克粒状白糖和2克液态麦芽萃取物组成。 注: 可以一次分析多个样品, 虽然样品的确切数量将取决于罐头压力锅的大小。</li>
	<li>用玻璃搅拌棒将溶液与剥落砂砾材料混合。</li>
	<li>在罐装压力锅中的水开始沸腾后, 添加1升的自来水以冷却罐装压力锅中的水。</li>
	<li>把罐头罐放在罐装压力锅里, 这样它们就能彼此等距, 从罐装压力锅的墙上。</li>
</ol><img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/57809/57809fig1.jpg"> 图 1: 在罐装压力锅中放置罐装罐.罐头罐应该被放置相等从彼此和从罐头压力锅的边, 保证甚而烹调和避免损坏罐头罐。<a href="https://www.jove.com/files/ftp_upload/57809/57809fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<ol start="8"><li>让水沸腾。把盖子放在罐装压力锅上。</li>
	<li>在 15 psi 上烹调大的剥落的面粉或剥落的砂砾材料为一 h. 允许罐装压力锅在打开前完全冷却和减压。</li>
	<li>使用耐热手套从罐装压力锅上取下盖子。</li>
	<li>使用钳子从罐装压力锅中取出罐装罐。把罐子放在耐热的表面上。 注: 所产生的中间产品在这一点上是熟的面粉。</li>
	<li>如果使用 Rausch 所概述的协议所产生的剥落砂砾材料 。7,烹调后用刮刀除去非胚乳材料。如果使用工业剥落磨料, 跳过这一步。</li>
</ol><img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/57809/57809fig2.jpg"> 图 2: 去除非胚乳材料.(a) 在烹调过程中, 在去除非胚乳材料之前, 煮熟的砂砾样品。(b) 去除非胚乳材料后的熟砂样品。<a href="https://www.jove.com/files/ftp_upload/57809/57809fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<ol start="13"><li>将30克熟的面粉 (每加工样品) 放在一条重量船上, 在烤箱中干燥65摄氏度, 12 小时。烘干后, 用咖啡机研磨熟砂样品, 并将其贮存在阴凉干燥处进行酚类分析。</li>
</ol>2. 生产烘烤的面粉
<ol><li>
把剩下的熟的面粉放在铝箔衬里的烘烤板上。
	<ol>
<li>要提高吞吐量, 请同时烘烤两个样本。要做到这一点, 在烘烤板上创建两个箔船。这消除了样品间交叉污染的可能性。</li>
	</ol>
</li>
</ol><img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/57809/57809fig3.jpg"> 图 3: 在烘烤板上放置熟的面粉.在烘烤前, 两个不同的熟砂样品放在单独的箔船上烘烤托盘上。照片上的小船上贴着绿胶带。这增加了议定书的吞吐量, 同时确保没有发生交叉污染。<a href="https://www.jove.com/files/ftp_upload/57809/57809fig3large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<ol start="2"><li>
将两个样品放在预热的对流烤箱中, 放置在107.2 摄氏度 (225°F) 50 分钟的烘烤盘中。
	<ol>
<li>搅拌样品25分钟后烘烤, 以确保甚至烘烤。</li>
	</ol>
</li>
	<li>在50分钟的结束时间内, 取出含有前两个样品的烘烤盘, 并允许在室温下冷却30分钟。</li>
	<li>在冷却期结束时, 从烤粒中间产品中取30克样品。把这个样品放在一个重量船在一个烤箱在65°c 12 小时。烘干后, 用咖啡机和商店进行植物分析, 将烘烤后的砂砾试样研磨成细粉。</li>
</ol>3. 生产最终烤玉米片产品
<ol><li>
用玉米饼印刷机把烤过的面粉卷起来。
	<ol>
<li>从铝箔衬里的烤盘中取出剩余的烤粒, 将这些面粉放在大约1米长的羊皮纸纸上。 注: 为了增加吞吐量, 将羊皮纸纸长度折叠成袋是有帮助的。这将减少在随后滚动阶段丢失的样本量。</li>
		<li>通过玉米饼印刷机慢慢地在袋中喂出被烘烤的砂砾样。注意不要在印刷机上捏手指。</li>
	</ol>
</li>
</ol><img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/57809/57809fig4.jpg"> 图 4: 羊皮纸纸袋.(a) 羊皮纸纵向折叠。(b) 邮袋的长而开的一侧折叠起来。(c) 长边以10°的角度再次折叠。(d) 邮袋的短、开边被折叠。这将是通过玉米饼印刷机喂养的邮袋的一侧。<a href="https://www.jove.com/files/ftp_upload/57809/57809fig4large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<ol start="2"><li>将轧制的烤粒切成2.5 厘米 x 2.5 厘米 (1 英寸2) 方块。使用一个工具, 如比萨饼切割机, 以削减/评分的滚动面团通过羊皮纸纸。</li>
</ol><img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/57809/57809fig5.jpg"> 图5。把卷成的面团切成薄片.轧面团是通过羊皮纸的纸。<a href="https://www.jove.com/files/ftp_upload/57809/57809fig5large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<ol start="3"><li>
打开羊皮纸, 并允许轧制, 切割, 烘烤的面粉在室温下干燥12小时。
	<ol>
<li>为了提高吞吐量, 在室温下将干燥样品储存在箔覆盖的重量船上, 直到多个样品 (通常为24或更多) 准备好进行烘烤。</li>
	</ol>
</li>
	<li>预加热对流烤箱到204.4 °c (400 °F)。将干燥的 untoasted 鳞片样品放在平烤板上。传播样本, 使样本的最小重叠发生。这甚至可以保证烘烤。</li>
	<li>把样品放在烤箱里 60–90, 直到它达到合适的颜色 (见图 6)。</li>
</ol><img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/57809/57809fig6.jpg"> 图 6: 最终烤玉米片的正确颜色.图片左侧的玉米片被烘烤了适当的时间。图片右侧的玉米片烤得太久了。<a href="https://www.jove.com/files/ftp_upload/57809/57809fig6large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<ol start="6"><li>允许样品在室温下冷却约5分钟。这就产生了最后的烤玉米片。</li>
	<li>用咖啡机将烤好的玉米片样品研磨成细粉。</li>
</ol>4. 植物和统计分析
注意: 根据植物的确切兴趣和实验室设备, 这些分析协议可能会发生变化。
<ol><li>使用植物中概述的协议 (如 Wilmsmeyer等) 来确定内容。3遵守议定书规定的所有安全程序。</li>
	<li>使用适当的统计模型分析数据。 注: 这些示例数据是用一个 RCBD 的分割图来分析的, 其中整个地块单元是收获谷物的场图, 次要单元是加工阶段。对 SAS (9.3 版) 混合过程进行了分析, 并在 R 中生成了数据。</li>
</ol>

<ol>
	<li>United States Department of Agriculture. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> National Agricultural Statistics Service. , (2017).</li><li>USDA. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> ERS. , (2017).</li><li>Butts-Wilmsmeyer, C. J., Mumm, R. H., Bohn, M. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Concentration+of+Beneficial+Phytochemicals+in+Harvested+Grain+of+U.S.+Yellow+Dent+Maize+(Zea+mays+L.)+Germplasm.">Concentration of Beneficial Phytochemicals in Harvested Grain of U.S. Yellow Dent Maize (Zea mays L.) Germplasm.</a> Journal of Agricultural and Food Chemistry. 65 (38), 8311-8318 (2017).</li><li>Butts-Wilmsmeyer, C. J., 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=Changes+in+phenolic+acid+content+in+maize+during+food+product+processing.">Changes in phenolic acid content in maize during food product processing.</a> Journal of Agricultural and Food. , (2018).</li><li>Dewanto, V., Wu, X. Z., Liu, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Processed+sweet+corn+has+higher+antioxidant+activity.">Processed sweet corn has higher antioxidant activity.</a> Journal of Agricultural and Food Chemistry. 50 (17), 4959-4964 (2002).</li><li>Fast, R. B., Caldwell, E. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Breakfast+cereals+and+how+they+are+made.">Breakfast cereals and how they are made.</a> Breakfast cereals and how they are made. , (2000).</li><li>Rausch, K. D., 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=Laboratory+measurement+of+yield+and+composition+of+dry-milled+corn+fractions+using+a+shortened,+single-stage+tempering+procedure.">Laboratory measurement of yield and composition of dry-milled corn fractions using a shortened, single-stage tempering procedure.</a> Cereal Chemistry. 86 (4), 434-438 (2009).</li><li>Kandhola, 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> Processing and Genetic Effects on Resistant Starch in Corn Flakes. , (2015).</li><li>Macke, J. A., Bohn, M. O., Rausch, K. D., Mumm, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+factors+underlying+dry-milling+efficiency+and+flaking-grit+yield+examined+in+us+maize+germplasm.">Genetic factors underlying dry-milling efficiency and flaking-grit yield examined in us maize germplasm.</a> Crop Science. 56 (5), 2516-2526 (2016).</li><li>Somavat, P., 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=A+new+lab+scale+corn+dry+milling+protocol+generating+commercial+sized+flaking+grits+for+quick+estimation+of+coproduct+yield+and+composition.">A new lab scale corn dry milling protocol generating commercial sized flaking grits for quick estimation of coproduct yield and composition.</a> Industrial Crops and Products. 109, 92-100 (2017).</li><li>Adam, A., 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=The+bioavailability+of+ferulic+acid+is+governed+primarily+by+the+food+matrix+rather+than+its+metabolism+in+intestine+and+liver+in+rats.">The bioavailability of ferulic acid is governed primarily by the food matrix rather than its metabolism in intestine and liver in rats.</a> Journal of Nutrition. 132 (7), 1962-1968 (2002).</li><li>Rumpagapom, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Structural Features of Cereal Bran Arabinoxylans Related to Colon Fermentation Rate. , (2011).</li><li>Wong, J. M. W., de Souza, R., Kendall, C. W. C., Emam, A., Jenkins, D. J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Colonic+health:+Fermentation+and+short+chain+fatty+acids.">Colonic health: Fermentation and short chain fatty acids.</a> Journal of Clinical Gastroenterology. 40 (3), 235-243 (2006).</li></ol>

作者没有什么可透露的。

在整个加工过程中, 玉米基食品的营养含量的变化很可能是由于去除了增益成分和热应力5,10。然而, 在本议定书4、8的发展之前, 对处理如何影响各种营养素的确切方法进行了比较详细的研究。此外, 由于大多数实验室处理方案的规模较大, 对玉米籽粒感官和营养特性的遗传基础的研究往往是不可能的.8。在这里, 我们提出了一个微型实验室方法来研究玉米的营养和感官性状在整个食品加工。
该协议允许取样发生在剥落的砂砾阶段, 烹调后, 烘烤后, 并在轧制过程中遇到的剪切力。因此, 通过对收获玉米籽粒的补充分析, 该议定书有利于分析初始阶段基质, 以及最终的食品和加工中间阶段, 以阐明与营养成分的变化。该协议的这一关键功能使营养和感官特征在整个处理过程中得到分析, 同时也使研究人员能够选择用于这些特定分析的分析化学协议。该协议的另一个主要特点是这种微型协议的效率。首先, 这个协议使用一个小样本, 这是适当的植物育种设置 (表 3)。一公斤谷物往往产生约0.3 公斤大的剥落砂砾成分, 大约有三个大的剥落砂成分生产需要处理。其次, 该协议允许实验室处理每天约16个样本, 这比以前的要求大样本量6的协议效率高得多。
该协议可以很容易地修改, 以模仿其他加工的玉米食品产品的生产。例如, 除了即食早餐谷物9, 大型剥落的面粉还用于生产各种快餐食品。生产这些快餐食品的实验室协议可以可以预见包括调整烹调时间和烹调方法或调整烘烤时间。还有可能将本议定书的修改版本用于研究其他谷物及其各自加工的产品。加工过的谷物产品通常包括烹调、烘烤或烘烤加工阶段, 可以使用此处介绍的协议修改版本进行模仿。
此协议的一个重要限制是它的停止点很少,即一旦处理步骤开始, 它和后续步骤必须完成 (表 3)。有一个单一的停止点后, 生产的熟粒从剥落的面粉。只有在必要的时候, 熟的面粉可以放在一个密封的容器 (如密封罐) 和冷藏至多两天。然而, 储存熟的面粉较长的时间内似乎改变样品。此外, 一旦烘焙开始, 就没有停止点, 直到烤砂面团已轧制, 切割和干燥。
结论
通过这些例子结果 (见 Wilmsmeyer等。4有关更多信息), 我们证明在整个加工过程中可以监测营养成分。此外, 确定了营养变化发生的关键处理阶段。此外, 该处理协议所需的小样本大小使得在植物育种计划的范围内对多种混合动力进行了研究。利用这些杂交种, 我们确定了在整个加工过程中, 哪组混合体保持了最高浓度的不溶性的阿魏酸和 p-香豆酸。这些特征是最终烤玉米片生命起源前潜力的重要标志。11,12,13这些结果可直接用于帮助植物育种者建立繁殖种群, 以改善加工玉米产品的生命起源前潜力。
这种处理协议的主要优点之一是它不限制可以进行的营养分析。如果植物协议存在于谷物分析中, 则可用于对加工产品进行研究。此外, 由于这种处理协议能够独立进行实验室规模的食品加工和营养分析, 因此可以研究多种植物化学物质。然而, 由于使用实验室尺度处理协议产生的中间和最终加工产品很少, 因此研究植物含量的分析协议应该使用小样本量。

玉米 (玉蜀黍) 是美国最广泛栽培的粮食作物1。在 2016年, 711.2亿公斤 (28亿蒲式耳) 玉米用于人类消费2, 表明玉米在美国饮食中的重要性。玉米谷物的最大好处之一是它是一种相对廉价的商品, 但它也含有有益的植物化学物质, 如酚类物质、不饱和脂肪酸和蛋白质3。因此, 以玉米为基础的食品可能是人类有益植物化学物质的相对廉价来源。
然而, 玉米必须在人类消费之前进行加工。因此, 加工活动往往影响最终加工食品4的营养价值。例如, 在生产快餐食品和即食早餐谷物 (即冷谷物) 的过程中, 玉米籽粒是干磨的, 以生产大型剥落的面粉。在干磨过程中, 麸皮和胚芽被物理去除, 只留下胚乳物质。由于许多植物化学物质主要位于麸皮或胚芽 (例如酚类和不饱和脂肪酸) 中, 这可能导致加工食品4的营养价值显著下降。反之, 下游加工步骤可提高营养价值。例如, 许多食品加工技术包括烹调、烘烤或烘烤。在这些阶段所遇到的热应力可以提高有益植物化学物质的生物利用度5。
从食品科学和人类营养的角度来看, 知道加工如何影响到加工食品的营养价值, 可以预见, 以及如何调整加工参数可能会影响其他感官, 这将是有趣的。品质, 包括颜色, 质地和味道。一项允许在整个加工过程中监测这种品质的协议, 可用于选择玉米品种, 以改善最终加工的玉米食品产品。过去分析这些特征的两个主要障碍是可用协议的规模和吞吐量。例如, 在生产用于实验室分析的早餐谷物时, 快速和考德威尔6建议使用45.4 公斤大的剥落的面粉。这种大量的大剥落的粒度远远超过了大量的大型剥落的面粉或大型剥落砂砾材料7 , 可以生产的小地块现场试验, 是典型的植物育种项目。因此, 制定加工食品生产的微型实验室协议, 可使 (1) 植物育种者改善对食品加工者具有重要意义的营养和感官特性的玉米品种, (2) 加工者有效地设计和测试替代处理策略。
在这篇手稿中, 我们描述了一个高通量修改 Kandohla8所描述的微型处理协议, 用于生产烤玉米片从大的剥落砂砾材料。本文给出了用该处理协议研究玉米中不溶性阿魏酸和对香豆酸变化的实例实验结果。我们在这项具体研究中的目标是确定 (1) 玉米的酚酸含量如何在生产即食早餐谷物时发生变化, (2) 在哪些加工阶段发生这些变化, (3) 是否我们的实验杂交种对加工应力反应不同, 该协议可与高通量分析化学协议结合, 有效地分析营养性状。该议定书还可进行调整, 以模仿其他谷物生产的其他加工玉米食品或加工食品产品。

<table border="0" cellpadding="0" cellspacing="0" width="1480">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:259px;">Canning pressure cooker</td>
			<td style="width:315px;">Wisconsin Aluminum Foundry Co.</td>
			<td style="width:108px;">Model 921</td>
			<td style="width:799px;">Any can be used, but it should be large enough to accommodate multiple canning jars</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Single burner or large hot plate</td>
			<td>Waring Professional</td>
			<td>Model SB30</td>
			<td>Any can be used, but it should be large enough so that canning pressure cooker can securely be placed on burner or hot plate</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">1 quart wide mouth canning jars</td>
			<td>Ball</td>
			<td>1440096258</td>
			<td>Any can be used, but they should be wide mouthed quart jars</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">1 L Beaker</td>
			<td>Fisher Scientific</td>
			<td>09-841-104</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Stir plate</td>
			<td>Corning</td>
			<td>6796420D</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Magnetic stir bar</td>
			<td>Fisher Scientific</td>
			<td>14-513-67</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">1 L Graduated cylinder</td>
			<td>Kimble</td>
			<td>20027500</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Spatula</td>
			<td>Wal-Mart</td>
			<td>552145280</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Hot pads</td>
			<td>Wal-Mart</td>
			<td>556501140</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Scale</td>
			<td>Any</td>
			<td>NA</td>
			<td>Mettler Toledo Model MS105DU or Similar</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Weigh boats</td>
			<td>Fisher Scientific</td>
			<td>08-732-113</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Sugar</td>
			<td>Wal-Mart</td>
			<td>9259244</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Salt</td>
			<td>Morton (Purchased at Wal-Mart)</td>
			<td>9244849</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Liquid malt extract</td>
			<td>By the Cup (Purchased on Amazon)</td>
			<td>NA</td>
			<td>https://www.amazon.com/Barley-Malt-Extract-Syrup-Bottle/dp/B01N4SK72C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Labeling tape&#160;</td>
			<td>Fisher Scientific</td>
			<td>15966</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Permanent marker</td>
			<td>Wal-Mart</td>
			<td>55529894</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Convection oven</td>
			<td>Wal-Mart</td>
			<td>1598495</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Baking pan (usually included with oven)</td>
			<td>Wal-Mart</td>
			<td>1598495</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Cooking foil</td>
			<td>Wal-Mart</td>
			<td>564264789</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tortilla press</td>
			<td>E&#38;A Hotel &#38; Restaurant Equipment and Supplies</td>
			<td>CTM-2000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Parchment paper</td>
			<td>Reynolds (Purchased at Wal-Mart)</td>
			<td>551219672</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Pizza cutter</td>
			<td>Farberware (Purchased at Wal-Mart)</td>
			<td>&#160;553012200</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Cooling racks</td>
			<td>Flytt (Purchased on Amazon)</td>
			<td>NA</td>
			<td>https://www.amazon.com/dp/B075HQY627/ref=sspa_dk_detail_7?psc=1&#38;pd_rd_i=B075HQY627&#38;pd_rd_wg=WaJol&#38;pd_rd_r=SF07KCHMP753WAPG6ED4&#38;pd_rd_w=2BOwf</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">SAS Version 9.4</td>
			<td>SAS Institute</td>
			<td>Version 9.4</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">R</td>
			<td>R Foundation for Statistical Computing</td>
			<td>Version 3.4.0</td>
			<td></td>
		</tr>
	</tbody>
</table>

high-throughput, microscale protocol for the analysis of processing parameters and nutritional qualities in maize (zea mays l.)

玉米是美国和世界上重要的粮食作物。然而, 在人类消费之前必须处理玉米谷物。此外, 玉米杂交种的整体籽粒组成和加工特性各不相同, 对最终加工产品的质量有影响。因此, 要想从玉米生产出更健康的加工食品, 就必须知道如何优化特定种质资源的加工参数, 以此来解释籽粒组成和加工特性的差异。这包括更好地了解当前的加工技术如何影响最终加工食品的营养质量。在这里, 我们描述了一个微型协议, 既模拟处理管道生产玉米片从大的剥落颗粒, 并允许处理多个谷物样品同时。作为高通量分析管道的一部分, 可对薄片颗粒、中间加工产品或最终加工产品以及玉米籽粒本身进行营养成分分析。本程序是专门为纳入玉米育种研究项目而开发的, 可用于其他粮食作物的改造。为分析玉米中不溶性阿魏酸和香豆酸含量提供了一个实例。样品在五个不同的加工阶段被采取了。我们证明, 在微型加工过程中, 取样可以在多个阶段进行, 这种加工技术可以在专门的玉米育种项目的背景下使用, 在我们的例子中, 大部分的营养成分都丢失了。在食品加工过程中。

该协议允许对加工后的玉米食品、玉米片进行抽样和营养分析, 从大剥落的面粉开始, 并通过加工的中间阶段持续到最终产品。该议定书与 Rausch等所概述的议定书相结合。7生产混合颗粒样品中的剥落砂砾成分。介绍了全谷物、大粒砂、熟砂、烤砂、烤玉米片加工阶段对混合样品的营养含量的信息。无论在评价的杂交品种, 大部分不溶性的阿魏酸和 p-香豆酸被删除在干铣削 (图 7)。在烹调过程中, 不溶性的阿魏酸和 p-香豆酸的另一减少。在烹调过程中观察到的不溶性阿魏酸和 p-香豆酸含量的减少, 可能是由于去除了大量的非胚乳材料, 而这些物质仍然存在于大的剥落砂砾材料中。多自由度对比表明, 阿魏酸和 p-香豆酸含量保持稳定, 在其余的处理, 无论混合 (表 1)。
此外, 杂交品种在其不溶性的阿魏酸含量和 p-香豆酸含量方面的初步排名在最后加工阶段没有表明混合动力的排名 (表 2和图 8)。换言之, 在整个内核中的初始内容并不表明在加工结束时, 混合体中哪一种杂交将拥有最不溶性的阿魏酸或 p-香豆酸。因此, 为了研究加工食品营养特性的遗传特征, 必须利用微尺度过程来研究玉米籽粒。
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/57809/57809fig7.jpg"> 图 7: 在整个加工过程中改变不溶性的酚酸含量.(a) 在整个加工过程中改变不溶性的阿魏酸含量。(b) 在整个加工过程中改变不溶性的香豆酸含量。周: 全仁, 成品: 剥落砂砾, CG: 熟的砂砾, BG: 烤的砂砾, 给: 烤玉米片。不同的颜色点代表不同的杂交种。图最初发表在补充信息的烟头-Wilmsmeyer等。4<a href="https://www.jove.com/files/ftp_upload/57809/57809fig7large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/57809/57809fig8.jpg"> 图 8: 交互图的混合处理阶段相互作用.(a) 不溶性的阿魏酸含量的相互作用图。(b) 不溶性香豆酸含量的相互作用图。相交的线表示一个变化的秩相互作用, 这意味着无论是不溶性的阿魏酸和不溶性的香豆酸含量的最终烤玉米片可以预测的基础上, 其中任一的初始内容整个内核中的植物化学物质。周: 全仁, 成品: 剥落砂砾, 对: 烤玉米片。图最初发表在补充信息的烟头-Wilmsmeyer等。4<a href="https://www.jove.com/files/ftp_upload/57809/57809fig8large.jpg" target="_blank">请单击此处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></td>
			<td colspan="2" style="text-align: center;">阿魏酸</td>
			<td colspan="2" style="text-align: center;">对羟基香豆酸</td>
		</tr>
<tr>
<td>混合</td>
			<td style="text-align: center;">F值</td>
			<td style="text-align: center;">p值</td>
			<td style="text-align: center;">F值</td>
			<td style="text-align: center;">p值</td>
		</tr>
<tr>
<td>B73xMO17</td>
			<td style="text-align: center;">0.07</td>
			<td style="text-align: center;">0.93</td>
			<td style="text-align: center;">0.34</td>
			<td style="text-align: center;">0.72</td>
		</tr>
<tr>
<td>B73xPHG47</td>
			<td style="text-align: center;">0.02</td>
			<td style="text-align: center;">0.98</td>
			<td style="text-align: center;">0.61</td>
			<td style="text-align: center;">0.55</td>
		</tr>
<tr>
<td>LH1xMO17</td>
			<td style="text-align: center;">0.08</td>
			<td style="text-align: center;">0.93</td>
			<td style="text-align: center;">0.14</td>
			<td style="text-align: center;">0.87</td>
		</tr>
<tr>
<td>PHJ40xLH123HT</td>
			<td style="text-align: center;">0.32</td>
			<td style="text-align: center;">0.73</td>
			<td style="text-align: center;">0.74</td>
			<td style="text-align: center;">0.48</td>
		</tr>
<tr>
<td>PH207xPHG47</td>
			<td style="text-align: center;">0.15</td>
			<td style="text-align: center;">0.86</td>
			<td style="text-align: center;">0.24</td>
			<td style="text-align: center;">0.79</td>
		</tr>
<tr>
<td>PHJ40xMO17</td>
			<td style="text-align: center;">0.01</td>
			<td style="text-align: center;">0.99</td>
			<td style="text-align: center;">0.31</td>
			<td style="text-align: center;">0.74</td>
		</tr>
<tr>
<td>PHG39xPHZ51</td>
			<td style="text-align: center;">0.06</td>
			<td style="text-align: center;">0.94</td>
			<td style="text-align: center;">0.07</td>
			<td style="text-align: center;">0.93</td>
		</tr>
</tbody></table>表 1: 多自由度对比试验了熟粒、烤粒和烤玉米片中不溶性的酚酸含量的差异。
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always"><tbody>
<tr>
<td></td>
			<td colspan="2" style="text-align: center;">阿魏酸</td>
			<td colspan="2" style="text-align: center;">对羟基香豆酸</td>
		</tr>
<tr>
<td></td>
			<td style="text-align: center;">F值</td>
			<td style="text-align: center;">p 值</td>
			<td style="text-align: center;">F值</td>
			<td style="text-align: center;">p 值</td>
		</tr>
<tr>
<td>杂交种</td>
			<td style="text-align: center;">7.15</td>
			<td style="text-align: center;">0.001</td>
			<td style="text-align: center;">8。7</td>
			<td style="text-align: center;">< 0.001</td>
		</tr>
<tr>
<td>自交系</td>
			<td style="text-align: center;">4.07</td>
			<td style="text-align: center;">0.007</td>
			<td style="text-align: center;">6.57</td>
			<td style="text-align: center;">< 0.001</td>
		</tr>
<tr>
<td colspan="5">注: 所有按基因分型的按加工阶段 intereactions 在α = 0.05 不显著。</td>
		</tr>
</tbody></table>表 2: 按加工阶段相互作用的基因型的意义。
<table align="center" border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always"><tbody>
<tr>
<td style="text-align: center;">协议步骤</td>
			<td style="text-align: center;">关键信息</td>
			<td style="text-align: center;">故障 排除</td>
			<td style="text-align: center;">高通量建议</td>
		</tr>
<tr>
<td style="text-align: center;">1.2 和1。6</td>
			<td style="text-align: center;">这两个步骤的耦合, 使水加热, 而不打破罐装罐。</td>
			<td style="text-align: center;">那</td>
			<td style="text-align: center;">在添加罐装罐之前, 加热一半的水会增加吞吐量。</td>
		</tr>
<tr>
<td style="text-align: center;">1。4</td>
			<td style="text-align: center;">那</td>
			<td style="text-align: center;">那</td>
			<td style="text-align: center;">预测量成分。 给定的量是一个罐子, 所以乘以数量或大量的配料所需的罐子的数目, 使用的步骤1.4.2。将产生的混合物均匀地分在罐罐中。</td>
		</tr>
<tr>
<td style="text-align: center;">1.4 注</td>
			<td style="text-align: center;">不要让罐子接触到罐装压力锅的边缘。他们将打破, 样品将丢失。</td>
			<td style="text-align: center;">那</td>
			<td style="text-align: center;">那</td>
		</tr>
<tr>
<td style="text-align: center;">1.9 和1.10</td>
			<td style="text-align: center;">在烹调之前, 水应该达到沸腾的沸点。</td>
			<td style="text-align: center;">如果在一个 h 后未彻底煮熟, 则检查以确保压力重量设置为 15 psi, 水是达到充分煮沸之前, 盖子放在罐头压力锅和定时器设置。</td>
			<td style="text-align: center;">那</td>
		</tr>
<tr>
<td style="text-align: center;">1.13</td>
			<td style="text-align: center;">除去在烹调过程中漂浮到顶端的非胚乳材料。 这将扭曲任何植物结果, 如果在样本中留下。</td>
			<td style="text-align: center;">如果非胚乳物质在烹调过程中不上升到顶端, 那么面粉就不会煮彻底。 请参见有关步骤1.9 和1.10 的信息。</td>
			<td style="text-align: center;">那</td>
		</tr>
<tr>
<td style="text-align: center;">1.15、2.4 和3。7</td>
			<td style="text-align: center;">研磨样品到细粉。</td>
			<td style="text-align: center;">如果植物分析似乎没有工作, 确保样品已研磨到细粉, 这样就有一个更大的表面积暴露在溶剂。</td>
			<td style="text-align: center;">那</td>
		</tr>
<tr>
<td style="text-align: center;">2。1</td>
			<td style="text-align: center;">不要让样品互相接触。 它们将成为交叉污染。</td>
			<td style="text-align: center;">那</td>
			<td style="text-align: center;">同时烘烤两个样品, 在一张烹饪纸上制作各自的箔船。</td>
		</tr>
<tr>
<td style="text-align: center;">2.2。1</td>
			<td style="text-align: center;">搅拌样品后25分钟, 以确保甚至烘烤。</td>
			<td style="text-align: center;">如果样品似乎没有均匀烘烤, 在更频繁的间隔 (例如每15分钟) 搅拌。</td>
			<td style="text-align: center;">那</td>
		</tr>
<tr>
<td style="text-align: center;">3。1</td>
			<td style="text-align: center;">将烘烤的砂砾面团放在羊皮纸纸袋中。 这样可以确保样品在冲压过程中不会丢失。</td>
			<td style="text-align: center;">如果样品开始出来的羊皮纸纸袋的末尾, 使邮袋更长。我们发现1米似乎是足够的。</td>
			<td style="text-align: center;">那</td>
		</tr>
<tr>
<td style="text-align: center;">3。2</td>
			<td style="text-align: center;">把羊皮纸纸袋收起来。</td>
			<td style="text-align: center;">如果切割工具通过羊皮纸, 使用一个更枯燥的工具。</td>
			<td style="text-align: center;">我们发现, 一个比萨切割机是最好的工具, 以削减烘烤的面粉成方形。 我们没有用这个工具剪掉羊皮纸, 但是烤的面粉仍然能够很快地被切成正方形。</td>
		</tr>
<tr>
<td style="text-align: center;">3。5</td>
			<td style="text-align: center;">变得非常舒适的颜色和不烤太久。</td>
			<td style="text-align: center;">如果样品变得太黑, 减少用于敬酒的时间。</td>
			<td style="text-align: center;">在个别箔覆盖的重量船上储存多个干燥的烘烤砂砾样品, 直到多个样品准备好进行烘烤。</td>
		</tr>
</tbody></table>表 3:关键步骤表、疑难解答步骤和建议.

Watch this Scientific Journal Video about High-throughput, Microscale Protocol for the Analysis of Processing Parameters and Nutritional Qualities in Maize Zea mays L. at JoVE.com

High-throughput, Microscale Protocol for the Analysis of Processing Parameters and Nutritional Qualities in Maize Zea mays L.

在这里, 我们提出了处理谷物样品的微型协议, 并将这种微型方法纳入高通量分析管道。这是对当前可用协议的更高吞吐量适应性。

玉米加工参数和营养品质分析的高通量、微尺度协议 (玉蜀黍)

Maize is an important grain crop in the United States and worldwide. However, maize grain must be processed prior to human consumption. Furthermore, whole ...

high-throughput-microscale-protocol-for-analysis-processing

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JoVE Journal

Environment

High-throughput, Microscale Protocol for the Analysis of Processing Parameters and Nutritional Qualities in Maize (Zea mays L.)

6.8K 次观看.  University of Illinois at Urbana-Champaign. 在这里, 我们提出了处理谷物样品的微型协议, 并将这种微型方法纳入高通量分析管道。这是对当前可用协议的更高吞吐量适应性。

视频: High-throughput, Microscale Protocol for the Analysis of Processing Parameters and Nutritional Qualities in Maize Zea mays L.

Determination of Microbial Extracellular Enzyme Activity in Waters, Soils, and Sediments using High Throughput Microplate Assays

Much of the nutrient cycling and carbon processing in natural environments occurs through the activity of extracellular enzymes released by microorganisms. Thus, measurement of the activity of these extracellular enzymes can give insights into the rates of ecosystem level processes, such as organic matter decomposition or nitrogen and phosphorus mineralization. Assays of extracellular enzyme activity in environmental samples typically involve exposing the samples to artificial colorimetric or fluorometric substrates and tracking the rate of substrate hydrolysis. Here we describe microplate based methods for these procedures that allow the analysis of large numbers of samples within a short time frame. Samples are allowed to react with artificial substrates within 96-well microplates or deep well microplate blocks, and enzyme activity is subsequently determined by absorption or fluorescence of the resulting end product using a typical microplate reader or fluorometer. Such high throughput procedures not only facilitate comparisons between spatially separate sites or ecosystems, but also substantially reduce the cost of such assays by reducing overall reagent volumes needed per sample.

Microplate based procedures are described for the colorimetric or fluorometric analysis of extracellular enzyme activity. These procedures allow for the rapid assay of such activity in large numbers of environmental samples within a manageable time frame.

在使用高通量微孔板测定水，土壤，沉积物和微生物测定胞外酶活性的

Much of the nutrient cycling and carbon  processing in natural environments occurs through the activity of extracellular  enzymes released by ...

科研

环境科学

High-throughput Fluorometric Measurement of Potential Soil Extracellular Enzyme Activities

Microbes in soils and other environments produce extracellular enzymes to depolymerize and hydrolyze organic macromolecules so that they can be assimilated for energy and nutrients. Measuring soil microbial enzyme activity is crucial in understanding soil ecosystem functional dynamics. The general concept of the fluorescence enzyme assay is that synthetic C-, N-, or P-rich substrates bound with a fluorescent dye are added to soil samples. When intact, the labeled substrates do not fluoresce. Enzyme activity is measured as the increase in fluorescence as the fluorescent dyes are cleaved from their substrates, which allows them to fluoresce. Enzyme measurements can be expressed in units of molarity or activity. To perform this assay, soil slurries are prepared by combining soil with a pH buffer. The pH buffer (typically a 50 mM sodium acetate or 50 mM Tris buffer), is chosen for the buffer&#39;s particular acid dissociation constant (pKa) to best match the soil sample pH. The soil slurries are inoculated with a nonlimiting amount of fluorescently labeled (i.e. C-, N-, or P-rich) substrate. Using soil slurries in the assay serves to minimize limitations on enzyme and substrate diffusion. Therefore, this assay controls for differences in substrate limitation, diffusion rates, and soil pH conditions; thus detecting potential enzyme activity rates as a function of the difference in enzyme concentrations (per sample).
Fluorescence enzyme assays are typically more sensitive than spectrophotometric (i.e. colorimetric) assays,&#160;but can suffer from interference caused by impurities and the instability of many fluorescent compounds when exposed to light; so caution is required when handling fluorescent substrates. Likewise, this method only assesses potential enzyme activities under laboratory conditions when substrates are not limiting. Caution should be used when interpreting the data representing cross-site comparisons with differing temperatures or soil types, as in situ soil type and temperature can influence enzyme kinetics.

To measure potential rates of soil extracellular enzyme activities, synthetic substrates that are bound to a fluorescent dye are added to soil samples. Enzyme activity is measured as the fluorescent dye is released from the substrate by an enzyme-catalyzed reaction, where higher fluorescence indicates more substrate degradation.

潜在土壤胞外酶活性的高通量荧光测量

Microbes in soils and other environments produce extracellular enzymes to depolymerize and hydrolyze organic macromolecules so that they can be ...

Optimized Negative Staining: a High-throughput Protocol for Examining Small and Asymmetric Protein Structure by Electron Microscopy

Structural determination of proteins is rather challenging for proteins with molecular masses between 40 - 200 kDa. Considering that more than half of natural proteins have a molecular mass between 40 - 200 kDa1,2, a robust and high-throughput method with a nanometer resolution capability is needed. Negative staining (NS) electron microscopy (EM) is an easy, rapid, and qualitative approach which has frequently been used in research laboratories to examine protein structure and protein-protein interactions. Unfortunately, conventional NS protocols often generate structural artifacts on proteins, especially with lipoproteins that usually form presenting rouleaux artifacts. By using images of lipoproteins from cryo-electron microscopy (cryo-EM) as a standard, the key parameters in NS specimen preparation conditions were recently screened and reported as the optimized NS protocol (OpNS), a modified conventional NS protocol 3 . Artifacts like rouleaux can be greatly limited by OpNS, additionally providing high contrast along with reasonably high&#8208;resolution (near 1 nm) images of small and asymmetric proteins. These high-resolution and high contrast images are even favorable for an individual protein (a single object, no average) 3D reconstruction, such as a 160 kDa antibody, through the method of electron tomography4,5. Moreover, OpNS can be a high&#8208;throughput tool to examine hundreds of samples of small proteins. For example, the previously published mechanism of 53 kDa cholesteryl ester transfer protein (CETP) involved the screening and imaging of hundreds of samples 6. Considering cryo-EM rarely successfully images proteins less than 200 kDa has yet to publish any study involving screening over one hundred sample conditions, it is fair to call OpNS a high-throughput method for studying small proteins. Hopefully the OpNS protocol presented here can be a useful tool to push the boundaries of EM and accelerate EM studies into small protein structure, dynamics and mechanisms.

More than half of proteins are small proteins (molecular mass &#60;200 kDa) that are challenging for both electron microscope imaging and three-dimensional reconstructions. Optimized negative staining is a robust and high-throughput protocol to obtain high contrast and relatively high resolution (~1 nm) images of small asymmetric proteins or complexes under different physiological conditions.

优化负染:通过电子显微镜检查小和不对称蛋白结构的高通量协议

Structural determination of proteins is rather challenging for proteins with molecular masses between 40 - 200 kDa. Considering that more than half of ...

Semi-High Throughput Screening for Potential Drought-tolerance in Lettuce (Lactuca sativa) Germplasm Collections

This protocol describes a method by which a large collection of the leafy green vegetable lettuce (Lactuca sativa L.) germplasm was screened for likely drought-tolerance traits. Fresh water availability for agricultural use is a growing concern across the United States as well as many regions of the world. Short-term drought events along with regulatory intervention in the regulation of water availability coupled with the looming threat of long-term climate shifts that may lead to reduced precipitation in many important agricultural regions has increased the need to hasten the development of crops adapted for improved water use efficiency in order to maintain or expand production in the coming years. This protocol is not meant as a step-by-step guide to identifying at either the physiological or molecular level drought-tolerance traits in lettuce, but rather is a method developed and refined through the screening of thousands of different lettuce varieties. The nature of this screen is based in part on the streamlined measurements focusing on only three water-stress indicators: leaf relative water content, wilt, and differential plant growth following drought-stress. The purpose of rapidly screening a large germplasm collection is to narrow the candidate pool to a point in which more intensive physiological, molecular, and genetic methods can be applied to identify specific drought-tolerant traits in either the lab or field. Candidates can also be directly incorporated into breeding programs as a source of drought-tolerance traits.

Semi-High Throughput Screening for Potential Drought-tolerance in Lettuce Lactuca sativa Germplasm Collections

This protocol was developed to screen a large germplasm collection of the leafy vegetable lettuce (Lactuca sativa L.) for drought-tolerance, in order to identify a small candidate pool of lettuce for use in physiological, molecular, and genetic studies to identify underlying drought-tolerance traits along with breeding programs.

半高通量筛选潜在的抗干旱的生菜（<em&gt;莴苣</em&gt;）种质资源

This protocol describes a method by which a large collection of the leafy green vegetable lettuce (Lactuca sativa L.) germplasm was screened for likely ...

Use of Chironomidae (Diptera) Surface-Floating Pupal Exuviae as a Rapid Bioassessment Protocol for Water Bodies

Rapid bioassessment protocols using benthic macroinvertebrate assemblages have been successfully used to assess human impacts on water quality. Unfortunately, traditional benthic larval sampling methods, such as the dip-net, can be time-consuming and expensive. An alternative protocol involves collection of Chironomidae surface-floating pupal exuviae (SFPE). Chironomidae is a species-rich family of flies (Diptera) whose immature stages typically occur in aquatic habitats. Adult chironomids emerge from the water, leaving their pupal skins, or exuviae, floating on the water&#8217;s surface. Exuviae often accumulate along banks or behind obstructions by action of the wind or water current, where they can be collected to assess chironomid diversity and richness. Chironomids can be used as important biological indicators, since some species are more tolerant to pollution than others. Therefore, the relative abundance and species composition of collected SFPE reflect changes in water quality. Here, methods associated with field collection, laboratory processing, slide mounting, and identification of chironomid SFPE are described in detail. Advantages of the SFPE method include minimal disturbance at a sampling area, efficient and economical sample collection and laboratory processing, ease of identification, applicability in nearly all aquatic environments, and a potentially more sensitive measure of ecosystem stress. Limitations include the inability to determine larval microhabitat use and inability to identify pupal exuviae to species if they have not been associated with adult males.

Use of Chironomidae Diptera Surface-Floating Pupal Exuviae as a Rapid Bioassessment Protocol for Water Bodies

Rapid bioassessment protocols using benthic macroinvertebrates are often used to monitor and assess water quality. An efficient protocol involves collections of Chironomidae surface-floating pupal exuviae (SFPE). Here, techniques for field collection, laboratory processing, slide mounting, and identification of Chironomidae SFPE are described.

使用摇蚊科（双翅目）的表面浮蛹蜕为快速生物议定书水体

Rapid bioassessment protocols using benthic macroinvertebrate assemblages have been successfully used to assess human impacts on water quality. ...

Protocol for Microplastics Sampling on the Sea Surface and Sample Analysis

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of scientific publications address microplastic pollution of the sea surface. The protocol below describes the methodology for sampling, sample preparation, separation and chemical identification of microplastic particles. A manta net fixed on an &#187;A frame&#171; attached to the side of the vessel was used for sampling. Microplastic particles caught in the cod end of the net were separated from samples by visual identification and use of stereomicroscopes. Particles were analyzed for their size using an image analysis program and for their chemical structure using ATR-FTIR and micro FTIR spectroscopy. The described protocol is in line with recommendations for microplastics monitoring published by the Marine Strategy Framework Directive (MSFD) Technical Subgroup on Marine Litter. This written protocol with video guide will support the work of researchers that deal with microplastics monitoring all over the world.

The protocol below describes the methodology for: microplastics sampling on the sea surface, separation of microplastic and chemical identification of particles. This protocol is in line with the recommendations for microplastics monitoring published by the MSFD Technical Subgroup on Marine Litter.

协议塑料微粒采样的海面和样品分析

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of ...

Extraction of Organochlorine Pesticides from Plastic Pellets and Plastic Type Analysis

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment during manufacturing and transport. Because of their environmental persistence, they are widely distributed in the oceans and on beaches all over the world. They can act as a vector of potentially toxic organic compounds (e.g., polychlorinated biphenyls) and might consequently negatively affect marine organisms. Their possible impacts along the food chain are not yet well understood. In order to assess the hazards associated with the occurrence of plastic pellets in the marine environment, it is necessary to develop methodologies that allow for rapid determination of associated organic contaminant levels. The present protocol describes the different steps required for sampling resin pellets, analyzing adsorbed organochlorine pesticides (OCPs) and identifying the plastic type. The focus is on the extraction of OCPs from plastic pellets by means of a pressurized fluid extractor (PFE) and on the polymer chemical analysis applying Fourier Transform-InfraRed (FT-IR) spectroscopy. The developed methodology focuses on 11 OCPs and related compounds, including dichlorodiphenyltrichloroethane (DDT) and its two main metabolites, lindane and two production isomers, as well as the two biologically active isomers of technical endosulfan. This protocol constitutes a simple and rapid alternative to existing methodology for evaluating the concentration of organic contaminants adsorbed on plastic pieces.

Microplastics act as vector of potentially toxic organic contaminants with unpredictable effects. This protocol describes an alternative methodology for assessing the levels of organochlorine pesticides adsorbed on plastic pellets and identifying the polymer chemical structure. The focus is on pressurized fluid extraction and attenuated total reflectance Fourier transform infrared spectroscopy.

从塑料颗粒中提取有机氯农药和塑料类型分析

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment ...

Dispersion of Nanomaterials in Aqueous Media: Towards Protocol Optimization

The sonication process is commonly used for de-agglomerating and dispersing nanomaterials in aqueous based media, necessary to improve homogeneity and stability of the suspension. In this study, a systematic step-wise approach is carried out to identify optimal sonication conditions in order to achieve a stable dispersion. This approach has been adopted and shown to be suitable for several nanomaterials (cerium oxide, zinc oxide, and carbon nanotubes) dispersed in deionized (DI) water. However, with any change in either the nanomaterial type or dispersing medium, there needs to be optimization of the basic protocol by adjusting various factors such as sonication time, power, and sonicator type as well as temperature rise during the process. The approach records the dispersion process in detail. This is necessary to identify the time points as well as other above-mentioned conditions during the sonication process in which there may be undesirable changes, such as damage to the particle surface thus affecting surface properties. Our goal is to offer a harmonized approach that can control the quality of the final, produced dispersion. Such a guideline is instrumental in ensuring dispersion quality repeatability in the nanoscience community, particularly in the field of nanotoxicology.

Here, we present a step-wise protocol for the dispersion of nanomaterials in aqueous media with real-time characterization to identify the optimal sonication conditions, intensity, and duration for improved stability and uniformity of nanoparticle dispersions without impacting the sample integrity.

纳米材料在水介质中的分散: 对协议优化的研究

The sonication process is commonly used for de-agglomerating and dispersing nanomaterials in aqueous based media, necessary to improve homogeneity and ...

A 3-dimensional (3D)-printed Template for High Throughput Zebrafish Embryo Arraying

The zebrafish is a globally recognized fresh water organism frequently used in developmental biology, environmental toxicology, and human disease related research fields. Thanks to its unique features, including large fecundity, embryo translucency, rapid and simultaneous development, etc., zebrafish embryos are often used for large scale toxicity assessment of chemicals and drug/compound screening. A typical screening procedure involves adult zebrafish spawning, embryos selection, and arraying the embryos into multi-well plates. From there, embryos are subjected to exposure and the toxicity of chemical, or the effectiveness of the drugs/compounds can be evaluated relatively quickly based on phenotypic observations. Among these processes, embryos arraying is one of the most time-consuming and labor-intensive steps that limits the throughput level. In this protocol, we present an innovative approach that makes use of a 3D-printed arraying template coupled with vacuum manipulation to speed up this laborious step. The protocol herein describes the overall design of the arraying template, a detailed experimental setup and step-by-step procedure, followed by representative results. When implemented, this approach should prove beneficial in a variety of research applications using zebrafish embryos as testing subjects.

A 3-dimensional 3D-printed Template for High Throughput Zebrafish Embryo Arraying

Here, we present a protocol to design and fabricate a zebrafish embryo arraying template, followed by a detailed procedure on the use of such template for high throughput zebrafish embryo arraying into a 96-well plate.

3维 (3 d) 印制的高吞吐量斑马鱼胚胎排列模板

The zebrafish is a globally recognized fresh water organism frequently used in developmental biology, environmental toxicology, and human disease related ...

Isolation and Analysis of Microbial Communities in Soil, Rhizosphere, and Roots in Perennial Grass Experiments

Plant and soil microbiome studies are becoming increasingly important for understanding the roles microorganisms play in agricultural productivity. The purpose of this manuscript is to provide detail on how to rapidly sample soil, rhizosphere, and endosphere of replicated field trials and analyze changes that may occur in the microbial communities due to sample type, treatment, and plant genotype. The experiment used to demonstrate these methods consists of replicated field plots containing two, pure, warm-season grasses (Panicum virgatum and Andropogon gerardii) and a low-diversity grass mixture (A. gerardii, Sorghastrum nutans, and Bouteloua curtipendula). Briefly, plants are excavated, a variety of roots are cut and placed in phosphate buffer, and then shaken to collect the rhizosphere. Roots are brought to the laboratory on ice and surface sterilized with bleach and ethanol (EtOH). The rhizosphere is filtered and concentrated by centrifugation. Excavated soil from around the root ball is placed into plastic bags and brought to the lab where a small amount of soil is taken for DNA extractions. DNA is extracted from roots, soil, and rhizosphere and then amplified with primers for the V4 region of the 16S rRNA gene. Amplicons are sequenced, then analyzed with open access bioinformatics tools. These methods allow researchers to test how the microbial community diversity and composition varies due to sample type, treatment, and plant genotype. Using these methods along with statistical models, the representative results demonstrate there are significant differences in microbial communities of roots, rhizosphere, and soil. Methods presented here provide a complete set of steps for how to collect field samples, isolate, extract, quantify, amplify, and sequence DNA, and analyze microbial community diversity and composition in replicated field trials.

Excavation of plant roots from the field as well as processing of samples into endosphere, rhizosphere, and soil are described in detail, including DNA extraction and data analysis methods. This paper is designed to enable other laboratories to use these techniques for the study of soil, endosphere, and rhizosphere microbiomes.

多年生草试验中土壤、根际和根系微生物群落的分离与分析

Plant and soil microbiome studies are becoming increasingly important for understanding the roles microorganisms play in agricultural productivity. The ...