Name: synthesis of strong adhesive hydrogel, gelatin o-nitrosobenzaldehyde
Uploaded: 2022-11-11T14:50:00
Description: Watch this Scientific Journal Video about Synthesis of Strong Adhesive Hydrogel, Gelatin O-Nitrosobenzaldehyde at JoVE.com

C57BL/6小鼠购自浙江大学医学院邵逸夫医院。新西兰兔是从浙江大学购买的。动物在自然的明暗循环条件下饲养，并自由提供食物和饮用水。所有实验程序均通过浙江大学伦理委员会标准指南（ZJU20200156）和浙江大学医学院邵逸夫爵士医院动物护理和使用委员会的机构指南，符合NIH实验动物护理和使用指南（SRRSH202107106）。
1. NB-COOH的合成
<ol><li>根据?...

<ol>
	<li>Tam, R. Y., Smith, L. J., Shoichet, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Engineering+cellular+microenvironments+with+photo-+and+enzymatically+responsive+hydrogels:+toward+biomimetic+3D+cell+culture+models.">Engineering cellular microenvironments with photo- and enzymatically responsive hydrogels: toward biomimetic 3D cell culture models.</a> Accounts of Chemical Research. 50 (4), 703-713 (2017).</li><li>Xu, X., 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=Bioadhesive+hydrogels+demonstrating+pH-independent+and+ultrafast+gelation+promote+gastric+ulcer+healing+in+pigs.">Bioadhesive hydrogels demonstrating pH-independent and ultrafast gelation promote gastric ulcer healing in pigs.</a> Science Translational Medicine. 12 (558), (2020).</li><li>Zheng, 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=Directed+self-assembly+of+herbal+small+molecules+into+sustained+release+hydrogels+for+treating+neural+inflammation.">Directed self-assembly of herbal small molecules into sustained release hydrogels for treating neural inflammation.</a> Nature Communications. 10 (1), 1604 (2019).</li><li>Seif-Naraghi, S. B., 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=Safety+and+efficacy+of+an+injectable+extracellular+matrix+hydrogel+for+treating+myocardial+infarction.">Safety and efficacy of an injectable extracellular matrix hydrogel for treating myocardial infarction.</a> Science Translational Medicine. 5 (173), (2013).</li><li>Mao, Q., 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=GelNB+molecular+coating+as+a+biophysical+barrier+to+isolate+intestinal+irritating+metabolites+and+regulate+intestinal+microbial+homeostasis+in+the+treatment+of+inflammatory+bowel+disease.">GelNB molecular coating as a biophysical barrier to isolate intestinal irritating metabolites and regulate intestinal microbial homeostasis in the treatment of inflammatory bowel disease.</a> Bioactive Materials. 19, 251-267 (2022).</li><li>Nan, 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=A+highly+elastic+and+fatigue-resistant+natural+protein-reinforced+hydrogel+electrolyte+for+reversible-compressible+quasi-solid-state+supercapacitors.">A highly elastic and fatigue-resistant natural protein-reinforced hydrogel electrolyte for reversible-compressible quasi-solid-state supercapacitors.</a> Advanced Science. 7 (14), 2000587 (2020).</li><li>Matsumoto, K., Sakikawa, N., Miyata, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thermo-responsive+gels+that+absorb+moisture+and+ooze+water.">Thermo-responsive gels that absorb moisture and ooze water.</a> Nature Communications. 9 (1), 2315 (2018).</li><li>Liu, R., 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=resilient,+adhesive,+and+anti-freezing+hydrogels+cross-linked+with+a+macromolecular+cross-linker+for+wearable+strain+sensors.">resilient, adhesive, and anti-freezing hydrogels cross-linked with a macromolecular cross-linker for wearable strain sensors.</a> ACS Applied Materials &amp; Interfaces. 13 (35), 42052-42062 (2021).</li><li>Hong, Y., 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+strongly+adhesive+hemostatic+hydrogel+for+the+repair+of+arterial+and+heart+bleeds.">A strongly adhesive hemostatic hydrogel for the repair of arterial and heart bleeds.</a> Nature Communications. 10 (1), 2060 (2019).</li><li>Yang, Y., 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=Tissue-integratable+and+biocompatible+photogelation+by+the+imine+crosslinking+reaction.">Tissue-integratable and biocompatible photogelation by the imine crosslinking reaction.</a> Advanced Materials. 28 (14), 2724-2730 (2016).</li><li>Ofner, C. M., Bubnis, W. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+and+swelling+evaluations+of+amino+group+crosslinking+in+gelatin+and+modified+gelatin+matrices.">Chemical and swelling evaluations of amino group crosslinking in gelatin and modified gelatin matrices.</a> Pharmaceutical Research. 13 (12), 1821-1827 (1996).</li><li>Zhang, Y., 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+long-term+retaining+molecular+coating+for+corneal+regeneration.">A long-term retaining molecular coating for corneal regeneration.</a> Bioactive Materials. 6 (12), 4447-4454 (2021).</li><li>Liang, Y., Li, Z., Huang, Y., Yu, R., Guo, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dual-dynamic-bond+cross-linked+antibacterial+adhesive+hydrogel+sealants+with+on-demand+removability+for+post-wound-closure+and+infected+wound+healing.">Dual-dynamic-bond cross-linked antibacterial adhesive hydrogel sealants with on-demand removability for post-wound-closure and infected wound healing.</a> ACS Nano. 15 (4), 7078-7093 (2021).</li></ol>

粘接材料是一类新型材料。越来越多的研究人员致力于各类胶粘剂材料的合成，并试图在生物技术、组织工程、再生医学等领域寻找其应用，这导致了近年来的蓬勃发展。除了关注粘合材料的强附着力外，研究人员还更加关注其他性能，如注射性、自愈性、止血性、抗菌性、控制去除等13。这些新应用极大地扩大了胶粘剂材料的应用范围，具有广阔的应用前景。
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水凝胶是一种由水溶胀形成的三维聚合物。特别是，来源于细胞外基质的水凝胶因其优异的生物相容性和治疗效果而广泛应用于生物合成和再生医学领域1。水凝胶已被报道用于治疗胃溃疡、神经炎、心肌梗塞2、3、4 和其他疾病。此外，已经证明明胶-NB可以促进炎症性炎症性肠病（IBD）的结果5。传统的水凝胶包括结冷胶、明胶、透明质酸、聚乙二醇（PEG）、层状疏水/亲水、海藻酸盐/聚丙烯酰胺、双网络和多两性水凝胶6，均具有良好的组织相容性和机械性能。然而，这些传统的水凝胶容易受到环境中的水分和空气的影响。如果长时间暴露在空气中，会失水变干;如果它们长时间浸泡在水中，它们会吸收水分并膨胀7，从而降低它们的柔韧性和机械功能。此外，保持传统水凝胶的组织粘附力是一项重大挑战8。
基于此，我们设计并合成了一种纳米级水凝胶明胶-NB，这是一种用NB修饰生物明胶形成的新型水凝胶（图1）。NB对-NH2 在组织上的粘附能力很强，可以形成大量的C=N键，从而增加了水凝胶-组织界面的粘附性。这种强的附着力可以使水凝胶牢固地粘附在组织表面，从而形成纳米级的分子涂层。在团队先前的研究中，已经证实这种改性水凝胶涂层提高了组织粘附性9;能稳定地粘附在角膜和肠道器官组织上，起到抗炎、屏障隔离、再生促进作用。目的是在这里详细介绍明胶-NB的具体合成过程，让明胶-NB应用于损伤修复的更多场景。此外，我们鼓励其他研究人员进一步加强和扩展这种材料的性质，以适应更多的应用场景。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1-(3Dimethylaminopropyl)-3-ethylcarbodimide hydrochloride (EDC)</td>
			<td>Aladdin</td>
			<td>L287553</td>
			<td></td>
		</tr>
		<tr>
			<td>4-Hydroxy-3-(methoxy-D3) benzaldehyde</td>
			<td>Shanghai Acmec Biochemical Co., Ltd</td>
			<td>H946072</td>
			<td></td>
		</tr>
		<tr>
			<td>DCM</td>
			<td>Aladdin</td>
			<td>D154840</td>
			<td></td>
		</tr>
		<tr>
			<td>Dichloromethane</td>
			<td>Sigma-Aldrich</td>
			<td>270997</td>
			<td></td>
		</tr>
		<tr>
			<td>Dimethyl sulfoxide (DMSO)</td>
			<td>Sigma-Aldrich</td>
			<td>20-139</td>
			<td></td>
		</tr>
		<tr>
			<td>dimethylformamide (DMF)</td>
			<td>Sigma-Aldrich</td>
			<td>PHR1553</td>
			<td></td>
		</tr>
		<tr>
			<td>gelatin</td>
			<td>Sigma-Aldrich</td>
			<td>1288485</td>
			<td></td>
		</tr>
		<tr>
			<td>magnesium sulfate</td>
			<td>Sigma-Aldrich</td>
			<td>M7506</td>
			<td></td>
		</tr>
		<tr>
			<td>MeOH</td>
			<td>Sigma-Aldrich</td>
			<td>1424109</td>
			<td></td>
		</tr>
		<tr>
			<td>methyl 4-(4-formyl-2-methoxyphenoxy methoxyphenyl) butanoic acid methyl ester</td>
			<td>chemsrc</td>
			<td>141333-27-9</td>
			<td></td>
		</tr>
		<tr>
			<td>methyl 4-bromobutyrate</td>
			<td>Aladdin</td>
			<td>M158832</td>
			<td></td>
		</tr>
		<tr>
			<td>NaBH4</td>
			<td>Sigma-Aldrich</td>
			<td>215511</td>
			<td></td>
		</tr>
		<tr>
			<td>N-hydroxysuccinimide (NHS)</td>
			<td>Aladdin</td>
			<td>D342712</td>
			<td></td>
		</tr>
		<tr>
			<td>nitric acid</td>
			<td>Sigma-Aldrich</td>
			<td>225711</td>
			<td></td>
		</tr>
		<tr>
			<td>potassium carbonate</td>
			<td>Sigma-Aldrich</td>
			<td>209619</td>
			<td></td>
		</tr>
		<tr>
			<td>SEM (Nova Nano 450)</td>
			<td>Thermo FEI</td>
			<td>17024560</td>
			<td></td>
		</tr>
		<tr>
			<td>THF/EtOH</td>
			<td>Aladdin</td>
			<td>D380010</td>
			<td></td>
		</tr>
		<tr>
			<td>trifluoroacetic acid (TFA)</td>
			<td>Sigma-Aldrich</td>
			<td>8.0826</td>
			<td></td>
		</tr>
	</tbody>
</table>

synthesis of strong adhesive hydrogel, gelatin o-nitrosobenzaldehyde

粘接材料已成为生物医学和组织工程领域流行的生物材料。在我们之前的工作中，我们提出了一种新材料 - 明胶邻亚硝基苯甲醛（明胶-NB） - 主要用于组织再生，并已在角膜损伤和炎症性肠病的动物模型中得到验证。这是一种通过用邻亚硝基苯甲醛（NB）修饰生物明胶而形成的新型水凝胶。明胶-NB通过激活NB-COOH的羧基，并通过1-（3-二甲氨基丙基）-3-乙基碳二亚胺盐酸盐（EDC）和N-羟基琥珀酰亚胺（NHS）与明胶反应合成。将得到的化合物纯化以生成最终产品，其可稳定储存至少18个月。NB对组织上的-NH2 有很强的粘附性，可以形成许多C=N键，从而增加了明胶-NB对组织界面的粘附。制备过程包括合成NB-COOH基团、修饰基团、合成明胶-NB和纯化化合物的步骤。目的是详细描述明胶-NB的具体合成过程，并演示明胶-NB在损伤修复中的应用。此外，提出该议定书是为了进一步加强和扩大科学界为更适用的情况制作的材料的性质。

图2A 显示了明胶-NB合成中涉及的主要化学反应的示意图，明胶-NB通过将NB基团嫁接到明胶上来促进组织整合。 图2B 显示，明胶-NB水凝胶的O-硝基苯在紫外线照射后立即转化为NB基团，然后活性醛基团可以与氨基交联形成席夫碱。 图2C 表明，不同比例的NB基团会导致明胶-NB的不同交联结构。
同时，还对明胶-NB的?...

Watch this Scientific Journal Video about Synthesis of Strong Adhesive Hydrogel, Gelatin O-Nitrosobenzaldehyde at JoVE.com

Synthesis of Strong Adhesive Hydrogel, Gelatin O-Nitrosobenzaldehyde (Video) | JoVE

这里介绍的方案显示了强粘性水凝胶明胶O-亚硝基苯甲醛（明胶-NB）的合成。明胶-NB具有快速高效的组织粘附能力，可以形成强大的物理屏障来保护伤口表面，因此有望应用于损伤修复生物技术领域。

强粘性水凝胶明胶O-亚硝基苯甲醛的合成

Adhesive materials have become popular biomaterials in the field of biomedical and tissue engineering. In our previous work, we presented a new material - ...

我们的协议详细描述了一种新的粘合水凝胶明胶邻亚硝基苯甲醛的合成。它主要解决如何最大化水凝胶附着力的问题。在我们的技术中，氢中的化学基团被修饰，从而与动物组织表面的化学基团发生化学反应，以实现紧密弯曲和关注。通过我们的技术合成的产品可以在动物组织表面保持强大的长期关注，从而有效地促进损伤修复和组织再生。这种方法可以提供对再生医学的见解，包括生物系统的损害。我们之前已经报道了用这种方法合成的产品在角膜损伤和IBD中的应用。首先，将手稿中描述的制备化合物溶解在40毫升N，N-二甲基甲酰胺中，并在环境温度下搅拌16小时。然后在混合物中加入200毫升零摄氏度的水，使混合物沉淀，即得粗品。反复将粗品溶解在N，N-二甲基甲酰胺中，然后沉淀5个循环。粗品沉淀后，在80摄氏度下干燥两小时，即得早期产品。通过将9.4克4-丁酸甲酯缓慢加入70%硝酸的预冷溶液中，并在零下2摄氏度下搅拌3小时，进行4-丁酸甲酯的ipso取代。然后沉淀混合物。用200毫升零摄氏度水过滤，在N，N-二甲基甲酰胺中纯化，析出固体产品。将固体产物在90摄氏度的三氟乙酸中水解，并将其放入旋转蒸发器中，然后在烤箱中干燥。中间产物溶解于四氢呋喃中后，在零摄氏度下缓慢加入硼氢化钠1.43克。三小时后，在真空下除去所有溶剂，并将残留物悬浮在一对一的水/二氯甲烷溶液中。制备二氯甲烷以从水层中提取产品，同时去除有机层并用硫酸镁干燥。然后使用二氯甲烷甲醇以10：1的比例通过硅胶柱色谱纯化粗产物。最后，得到5.31克相对纯的淡黄色粉末NB羧基。通过将5克明胶溶解在100毫升去离子水中并储存在37摄氏度进行一批改性来制备均匀的明胶溶液。按照手稿中所述确定进料比例后，将1， 060毫克NB羧基溶解在5毫升二甲基亚砜中。为了活化NB羧基的羧基，将746毫克1-3-乙基碳酰亚胺盐酸盐加入NB羧甲基亚砜溶液中，搅拌五分钟。1-3-乙基碳酰亚胺盐酸盐溶解后，加入448毫克N-羟基琥珀酰亚胺，搅拌5分钟。使用滴液漏斗以每分钟0.5毫升的速度将混合物缓慢滴入溶解的明胶溶液中，剧烈搅拌，在45摄氏度下反应四小时。将明胶NB溶液与过量的去离子水透析至少三天后，收集，冷冻并冻干以获得明胶-NB泡沫。将泡沫放在黑暗中的干燥剂中以备将来使用。使用前立即将冷冻干燥的明胶-NB 泡沫溶解在 37 摄氏度的去离子水中。获得受损角膜表面明胶和明胶-NB-4蛋白包衣处理角膜表面的SEM图像，显示明胶-NB-4蛋白的形态稳定粘附在角膜表面。然而，受伤的角膜表面没有经过任何处理或明胶处理，看起来很光滑。由明胶和明胶-NB分子涂层标记的小鼠结肠表面的荧光图像显示，荧光标记的明胶-NB具有粘附在肠道组织并形成致密涂层的能力。然而，明胶基团的荧光强度非常弱，表明它不能牢固地粘附在肠壁上。标记明胶和明胶-NB分子涂层处理的胺化板在零和24小时的荧光图像显示，两者都可以最初粘附在板上。在倒入PBS并每四小时更换一次胺化板24小时后，只有明胶-NB保持强荧光，表明其粘附性强。明胶-NB-4键合的X射线光子光谱表明，角膜表面和用明胶处理的表面的光谱是相同的。然而，在400电子伏特处出现一个额外的峰，表明用紫外线活化明胶-NB处理后在组织中形成许多CN键。要记住的最重要的一点是，在不同应用条件下合成明胶-NB时，需要设置实验前FR梯度，以探索最佳的注意力效果。这项技术可能为开发生物合成和再生医学铺平道路。研究人员可以参考视频来修改该组以满足生物医学需求。

Research

JoVE Journal

Bioengineering

Synthesis of Strong Adhesive Hydrogel, Gelatin O-Nitrosobenzaldehyde

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科研

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Natural polymers over the years have gained more importance because of their host biocompatibility and ability to interact with cells in vitro and in ...

Creating Adhesive and Soluble Gradients for Imaging Cell Migration with Fluorescence Microscopy

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创建的胶粘剂和可溶性梯度荧光显微镜成像细胞迁移

Cells can sense and migrate towards higher concentrations of adhesive cues such as the glycoproteins of the extracellular matrix and soluble cues such as ...

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Microelectrode  arrays for neural interface devices that are made of biocompatible thin-film  polymer are expected to have extended functional lifetime ...

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内含肽介导的人工蛋白水凝胶的合成

We present the synthesis of a highly stable protein hydrogel mediated by a split-intein-catalyzed protein trans-splicing reaction. The building blocks of ...

Using Adhesive Patterning to Construct 3D Paper Microfluidic Devices

Using Adhesive Patterning to Construct 3D Paper Microfluidic Devices (Video) | JoVE

使用粘性图案构建3D纸微流体装置

We demonstrate the use of patterned aerosol adhesives to construct both planar and nonplanar 3D paper microfluidic devices. By spraying an aerosol ...

Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation

Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation (Video) | JoVE

Bioprintable 藻酸盐/明胶水凝胶 3D体外模型诱导细胞球形形成

The cellular, biochemical, and biophysical heterogeneity of the native tumor microenvironment is not recapitulated by growing immortalized cancer cell ...

Fabricating a Kidney Cortex Extracellular Matrix-Derived Hydrogel

Fabricating a Kidney Cortex Extracellular Matrix-Derived Hydrogel (Video) | JoVE

肾皮质细胞外基质衍生水凝胶的制备

Extracellular matrix (ECM) provides important biophysical and biochemical cues to maintain tissue homeostasis. Current synthetic hydrogels offer robust ...

Rat Model of Adhesive Capsulitis of the Shoulder

Rat Model of Adhesive Capsulitis of the Shoulder (Video) | JoVE

肩上粘 Capsulitis 大鼠模型的建立

This proposal aims to create an in vivo rat model of adhesive capsulitis for researching potential treatment options for this condition and other ...

Wet-spinning-based Molding Process of Gelatin for Tissue Regeneration

Wet-spinning-based Molding Process of Gelatin for Tissue Regeneration (Video) | JoVE

明胶湿纺成型工艺促进组织再生

This article presents an inexpensive method to fabricate gelatin, as a natural polymer, into monofilament fibers or other appropriate forms. Through the ...