<p class="jove_content">The authors thank the funding of the National Natural Science Foundation of China (82001945), the Shanghai Pujiang Program (20PJ1410700), and the starting grant of ShanghaiTech University. The authors thank the Centre for High-resolution Electron Microscopy (ChEM), School of Physical Science and Technology, ShanghaiTech University (No. EM02161943) for the material characterization support. The authors thank the Analytical Instrumentation Center (#SPST-AIC10112914), School of Physical Science and Technology, ShanghaiTech University for the spectral test support and XRD test support. The authors also thank Prof. Jianfeng Li for the help with the material characterizations.</p>

<p class="jove_title"><strong>1. Synthesis of Zn</strong><strong><sub>2</sub>GeO<sub>4</sub>: Mn PLNPs</strong></p>
<ol>
	<li>Prepare 2 M/L sodium hydroxide solution by dissolving 10 mM sodium hydroxide in 5 mL of deionized water.</li>
	<li>Prepare 0.4 M/L sodium germanate solution by adding 2 mM of germanium oxide into 5 mL of sodium hydroxide solution, and then stir at room temperature for about 30 min.</li>
	<li>Add 4 mM of zinc chloride, 0.01 mM of manganese nitrate and 600 &#181;L of nitric acid (65%-68%, wt) to a 100 mL small beaker containing 22 mL of deionized water.<br />
	CAUTION: The addition of nitric acid should be strictly carried out in a fume hood and ensure that there are no open flames or heating around.</li>
	<li>Stir vigorously until the solution of step 1.3 is completely dissolved.</li>
	<li>Slowly add 2 mM sodium germanate solution to the solution of step 1.4. Add 1 mL of polyethylene glycol (PEG; 300, Mw) to the solution.</li>
	<li>Put the calibrated pH meter probe into the solution to monitor the pH value of the reaction system. Set a relatively gentle stirring to avoid splashing of the solution and collision between the stirring bar and the probe.</li>
	<li>Add ammonium hydroxide with a mass fraction of 25%-28% to the solution drop by drop, and adjust the pH of the solution to 6.0, 8.0 or 9.4 depending on the luminescence property to be studied. Be sure to add ammonium hydroxide slowly and monitor the pH changes of the solution at all times to prevent the system from being too acidic or too alkaline, so as not to affect the morphology and luminescent properties of nanomaterials.</li>
	<li>Cover the beaker with sealing film and stir the solution at room temperature for 1 h. Try not to expose the system to air to prevent dust from entering and causing solvent volatilization, while stirring at a constant speed so that the liquid level of the system will not splash when the system is fully mixed.</li>
	<li>Transfer the solution to a Teflon-lined autoclave and place it in an electric thermostatic drying oven at 220 &#176;C for 4 h.<br />
	NOTE: The appropriate Teflon-lined autoclave should be selected according to the volume of the system, and the reactor should be kept clean. The volume of the added raw materials should not exceed 1/3<sup>rd</sup> of the volume of the autoclave. At the same time, make sure the autoclave is completely closed before placing it into the electric thermostatic drying oven.</li>
	<li>Turn OFF the electric thermostatic drying oven when the reaction is completed and wait for the system to cool down to room temperature to take out the reactor. Be sure to wait until the reactor is completely cooled and the pressure is reduced to a safe range before proceeding to the next step to avoid direct contact between high temperature and skin.</li>
	<li>Slowly open the reactor and transfer the reaction solution to two 50 mL centrifuge tubes. Rinse the reactor with 40 mL of ethanol, and subsequently transfer the ethanol solution to the same centrifuge tubes.</li>
	<li>Vortex for 30 s so that the solution can be mixed evenly, then centrifuge the sample at 4000 x <em>g</em> for 15 min at room temperature and remove the supernatant.</li>
	<li>Add 10 mL of deionized water to each centrifuge tube and sonicate for 5 min (240 W, 40 kHz) to redisperse the product.</li>
	<li>Add 20 mL of ethanol into each centrifuge tube and vortex for 30 s to mix the solution evenly.</li>
	<li>Continue to centrifuge the product according to the setting mentioned before (4000 x <em>g</em>, 15 min) at room temperature and discard the supernatant.</li>
	<li>Ultrasonicate for 5 min to disperse the product in 2 mL of methanol solution, seal the sample with sealing film and store it in a 4 &#176;C refrigerator to prevent sample contamination and solvent evaporation for future illumination applications.</li>
</ol>
<p class="jove_title"><strong>2. Synthesis of ZnGa</strong><strong><sub>2</sub>O<sub>4</sub>: Cr PLNPs</strong></p>
<ol>
	<li>Dissolve 12 mM Ga(NO<sub>3</sub>)<sub>3</sub>.xH<sub>2</sub>O, 7.2 mM ZnCl<sub>2</sub> and 0.024 mM Cr(NO<sub>3</sub>)<sub>3</sub>.9H<sub>2</sub>O in 30 mL of deionized water.</li>
	<li>Add 1 mL of PEG (300, Mw) to the solution. Add ammonium hydroxide (25%-28% wt) to the solution, stir gently to reach a pH of 9.0-9.4. Be sure to control the stirring speed so that the solution can mix thoroughly without splashing onto the pH meter.</li>
	<li>Cover the beaker with a sealing film and stir the solution at room temperature for 1 h. Try to minimize the exposure of the system to air to prevent dust from entering and solvent evaporation. At the same time, control the stirring speed so that the system does not splash while mixing thoroughly.</li>
	<li>Transfer the solution to a Teflon-lined autoclave and run at 220 &#176;C for 6 h. Take out the container after the temperature drops to room temperature. Ensure that the reaction vessel has fully cooled down, and the pressure has dropped to a safe range before proceeding with subsequent operations, as well as avoid direct contact of high temperatures with the skin.</li>
	<li>Transfer the reaction solution to two 50 mL centrifuge tubes. Rinse the reactor with 40 mL of ethanol, and then transfer the ethanol solution to the same centrifuge tubes.</li>
	<li>Vortex for 30 s to mix the solution and then centrifuge the sample at 4000 x <em>g</em> for 15 min at room temperature and remove the supernatant.</li>
	<li>Add 10 mL of deionized water to each centrifuge tube and sonicate for 5 min to redisperse the product.</li>
	<li>Add 20 mL of ethanol into each centrifuge tube and vortex for 30 s to mix the solution evenly. Continue centrifuging the product at room temperature as previously mentioned (4000 x <em>g</em>, 15 min) and discard the supernatant.</li>
	<li>Ultrasonicate for 5 min to disperse the product in 2 mL of deionized water and seal the sample with a sealing film for storage.</li>
</ol>
<p class="jove_title"><strong>3. Purification for raw materials</strong></p>
<ol>
	<li>Purify methyl methacrylate (MMA) by column chromatography as described below.
	<ol>
		<li>Fill half of the column with alkaline aluminum oxide (100-200 mesh) and compact lightly with a glass rod. When filling the column with aluminum oxide, pay attention to even distribution and uniform compaction of the filler to improve separation efficiency.</li>
		<li>Add a small amount of MMA and open the PTFE throttle piston below. Once the solvent layer wets the entire aluminum oxide and liquid flows out, add more MMA and repeat this process multiple times. The time when mass ratio of the entire MMA added to the basic aluminum oxide is: 1:50 represents the end of the process.</li>
		<li>Place the final collected MMA sample into a glass bottle, seal it with a sealing film and store at 4 &#176;C.<br />
		CAUTION: The entire process should be carried out in a fume hood due to the strong volatility of MMA. At the same time, operators should wear masks and lab coats.</li>
	</ol>
	</li>
	<li>Purify azobisisobutyronitrile (AIBN) by recrystallization as described below.
	<ol>
		<li>Prepare a 50 mL of mixed solution with a volume ratio of 7:3 of ethanol and distilled water and heat the solution.</li>
		<li>Add 5 g of AIBN when the solution is boiling and stir to mix the solution evenly.</li>
		<li>Remove insoluble impurities by hot filtration as described below.
		<ol>
			<li>Place the filter paper snugly against the inner wall of the triangular funnel and ensure that the filter paper is below the edge of the funnel.</li>
			<li>Place the glass rod against the three-layer section of the filter paper. If the glass rod is not placed against the three-layer section of the filter paper, it may puncture the filter paper, leading to inefficient filtration.</li>
			<li>Place the tip of the beaker containing the solution close to the glass rod and pour it while it is hot. The purpose of this step is to prevent splashing liquid droplets.</li>
			<li>Rinse the beaker with 10 mL of cold distilled water, and again perform the above filtration process; repeat 3x.</li>
		</ol>
		</li>
		<li>The solution becomes supersaturated due to the decrease in solubility during cooling, resulting in the precipitation of crystals. Put the collected solution in a refrigerator at 4 &#176;C for cooling and crystallization. The sample will appear in the state of white needle-like crystals.</li>
		<li>Seal the sample with aluminum foil and store at 4 &#176;C.<br />
		CAUTION Protective measures should be taken during the operation due to the toxicity of AIBN, while also avoiding contact with open flames, high temperatures, and oxidizing agents.</li>
	</ol>
	</li>
</ol>
<p class="jove_title"><strong>4. Copolymerization of methyl methacrylate (MMA)</strong></p>
<ol>
	<li>Set the water bath temperature to 80 &#176;C.<br />
	NOTE: The temperature of the water has a severe influence on the rate of polymerization and thus affects the final product formation. Therefore, the temperature of the water bath should be strictly guaranteed not to be too high.</li>
	<li>Weigh 20 g of MMA into a 100 mL eggplant-shaped bottle. Keep the container dry before the experiment.<br />
	NOTE: The eggplant-shaped bottle is chosen to facilitate the water bath heating and the replacement of air with nitrogen in the system. Try to weigh the sample in a well-ventilated environment while wearing a mask.</li>
	<li>Add the pre-prepared methanol solution of Zn<sub>2</sub>GeO<sub>4</sub>: Mn into the reaction vessel.</li>
	<li>Thoroughly dissolve the sample in MMA with the help of ultrasound for about 10 min (240 W, 40 kHz) at room temperature. Keep the reaction vessel sealed to prevent solvent evaporation and avoid excessively high temperatures during the ultrasonication process.</li>
	<li>Add 0.012 g of AIBN to the solution and mix the solution completely.<br />
	NOTE: AIBN should be used under anhydrous conditions and ensure that there is no open flame around the experimental operation. Be sure to wear protective gear.</li>
	<li>Place the flask in an 80 &#176;C water bath and purge the air from the reaction system with N<sub>2</sub> for approximately 35 min. When the reaction is about to end, gently shake the reaction container. If the solution does not shake vigorously, it proves that the reaction is successful.<br />
	NOTE: The reaction time will change with the temperature of the water bath. Make sure that the temperature of the water bath reaches 80 &#176;C and start timing for 35 min.</li>
	<li>After the reaction is over, quickly transfer the reaction vessel to an ice bath to cool it down quickly.<br />
	NOTE: This process should be as fast as possible to avoid excessive pre-polymerization of MMA, and the ice bath can be prepared in advance during the reaction interval.</li>
	<li>Slowly pour the solution into a two-dimensional or three-dimensional mold, put the mold into an electric thermostatic drying oven at 40 &#176;C for 10 h, 70 &#176;C for 8 h, and 100 &#176;C for another 2 h to obtain target material.</li>
	<li>Close the electric thermostatic drying oven after the reaction stops, and let it cool down to room temperature. Open the electric thermostatic drying oven to take out the mold after the reaction system is sufficiently cooled to avoid skin burns caused by direct contact between high temperature and body.</li>
	<li>Carefully remove the mold and expose the polymerized PMMA sample (ZGO: Mn-PMMA) to a UV lamp for about 3 min. For example, when exposing a transparent ZGO: Mn-PMMA film to ultraviolet light through a black cardboard cutout in the shape of the letter H, a corresponding pattern of green phosphorescent emission is obtained. The pattern can be erased after 5 min. Subsequently, the process can be repeated by using another black cardboard cutout in the shape of different letters, generating new luminescent patterns.</li>
</ol>

Hydrothermal Synthesis of Persistent Luminescent Nanoparticles

<p class="jove_content">There is nothing to disclose.</p>

<p class="jove_content">This article introduces a synthesis method for persistent luminescent nanomaterials and polymerization for color rendering applications. The materials showed extremely stable optical properties and a visible afterglow after ceasing excitation of ultraviolet light. A persistent luminescent nanomaterial (Zn<sub>2</sub>GeO<sub>4</sub>: Mn) was prepared using a hydrothermal method with different pH (<strong class="xfig">Figure 1A</strong>). The TEM image showed that ZGO: Mn PLNPs whose pH is 9.4 were rod-shaped with an avera...

<p class="jove_content">Persistent luminescence (PL) is a unique optical process that can store energy from ultraviolet light, visible light, X-rays, or other excitation sources and then release it in the form of photon emission for seconds, minutes, hours, or even for days<sup class="xref">1</sup>. The discovery of continuous luminous phenomenon originated from the Song dynasty in ancient China 1000 years ago when a painter accidentally discovered a painting that glowed in the dark. It was later found that some natural raw materials and minerals could absorb sunlight and then glow in the dark and can even be made into fascinating glowing pearls<sup class="xref">2</sup>. However, the first adequate record of persistent phosphors needed to be traced back to the discovery of PL emission from Bologna stone in the early 17<sup>th</sup> century, which gave off a yellow to orange afterglow in the dark<sup class="xref">1</sup><sup>,</sup><sup class="xref">2</sup><sup>,</sup><sup class="xref">3</sup><sup>,</sup><sup class="xref">4</sup>. Later, it was discovered that the natural impurities of Cu<sup>+</sup> in BaS played an important role in this persistent luminescence phenomenon<sup class="xref">1</sup><sup>,</sup><sup class="xref">4</sup>. Until the mid-1990s, the production of persistent phosphors was largely limited to sulfides<sup class="xref">5</sup>. In 1996, Matsuzawa et al. reported a new metal oxide (SrAl<sub>2</sub>O<sub>4</sub>:Eu<sup>2+</sup>, Dy<sup>3+</sup>) phosphor showing extremely bright afterglow, which greatly stimulated the expansion of persistent luminescence research<sup class="xref">6</sup>.</p>
<p class="jove_content">The unique properties of persistent luminescent materials are mainly derived from two kinds of active centers: emission centers and trap centers<sup class="xref">1</sup><sup>,</sup><sup class="xref">7</sup><sup>,</sup><sup class="xref">8</sup>. Among them, the former determines the emission wavelength, while the sustained intensity and time are mainly determined by the trap centers. Therefore, the design of PL materials should take both aspects into consideration in order to achieve the desired emission wavelength and long-lasting luminescence<sup class="xref">9</sup><sup>,</sup><sup class="xref">10</sup>. The emission centers can be lanthanide ions with 5d to 4f or 4f to 4f transitions, transition metal ions with d to d transitions, or post-transition metal ions with p to s transitions<sup class="xref">1</sup><sup>,</sup><sup class="xref">11</sup><sup>,</sup><sup class="xref">12</sup><sup>,</sup><sup class="xref">13</sup>. On the other hand, trap centers are formed by lattice defects or various co-dopants<sup class="xref">14</sup><sup>,</sup><sup class="xref">15</sup>, which usually do not emit radiation but instead store the excitation energy for a while and then gradually release it to the emitting center through thermal or other physical activation<sup class="xref">16</sup><sup>,</sup><sup class="xref">17</sup>. Many phosphors with different hosts and dopant ions have been reported. So far, inorganic metal compounds<sup class="xref">18</sup>, metal-organic frameworks<sup class="xref">8</sup>, certain organic composites<sup class="xref">19</sup>, and polymers<sup class="xref">20</sup> have been found to have PL properties. In recent years, deep trap persistent luminescent materials with controllable energy storage and photon release properties have shown great potential applications in information storage<sup class="xref">21</sup>, multi-layer anti-counterfeiting<sup class="xref">22</sup>, and advanced displays<sup class="xref">23</sup>.</p>
<p class="jove_content">Based on the above composition, PLNPs with various matrices have been successfully designed and synthesized, such as BaZrSi<sub>3</sub>O<sub>9</sub><sup class="xref">7</sup>, Y<sub>2</sub>O<sub>2</sub>S<sup class="xref">24</sup>, Ca<sub>14</sub>Mg<sub>2</sub>(SiO<sub>4</sub>)<sub>8</sub><sup class="xref">25</sup>, CaAl<sub>2</sub>O<sub>4</sub><sup class="xref">26</sup>, SrAl<sub>2</sub>O<sub>4</sub><sup class="xref">26</sup><sup>,</sup><sup class="xref">27</sup> , and Sr<sub>2</sub>MgSi<sub>2</sub>O<sub>7</sub><sup class="xref">28</sup> with multi-doped luminescent centers, in which the luminescence centers strongly depend on the crystal field effect of the host lattice, while the defects generated or improved by different doping serve as auxiliary centers to control the afterglow intensity and duration. In addition to co-doping, long-lasting emission can also be observed in the case of only one activator, such as heterogeneous PLNPs with the matrix of Y<sub>3</sub>Al<sub>2</sub>Ga<sub>3</sub>O<sub>12</sub><sup class="xref">29</sup>, BaGa<sub>2</sub>O<sub>4</sub><sup class="xref">30</sup>, Ca<sub>2</sub>SnO<sub>4</sub><sup class="xref">31</sup>, CdSiO<sub>3</sub><sup class="xref">32</sup> , and Zn<sub>3</sub>Ga<sub>2</sub>Ge<sub>2</sub>O<sub>10</sub><sup class="xref">33</sup>. Germanate-based ternary oxides include Ca<sub>2</sub>Ge<sub>7</sub>O<sub>16</sub>, Zn<sub>2</sub>GeO<sub>4</sub>, BaGe<sub>4</sub>O<sub>9</sub>, etc., which are typical wide-bandgap semiconductor materials with tunable emission, reproducible and stable luminescence, high quantum yield, environmental friendliness and wide availability<sup class="xref">34</sup><sup>,</sup><sup class="xref">35</sup><sup>,</sup><sup class="xref">36</sup>. These advantages make it a good activator-type photoluminescent carrier. In the past few years, germanates with various microstructures<sup class="xref">35</sup><sup>,</sup><sup class="xref">37</sup>, have been prepared by conventional solid-state reactions or chemical solution methods, and these characteristics make Zn<sub>2</sub>GeO<sub>4</sub> useful in sterilize<sup class="xref">38</sup>, anti-counterfeiting<sup class="xref">39</sup>, catalysis<sup class="xref">40</sup>, light diodes<sup class="xref">41</sup> , biosensing<sup class="xref">42</sup>, battery anodes<sup class="xref">43</sup>, detectors<sup class="xref">44</sup><sup>,</sup><sup class="xref">45</sup>, etc.</p>
<p class="jove_content">In order to expand the application of PL materials, the controllable synthesis of uniform and persistent luminescent nanoparticles has been developed. A decade ago, persistent phosphors were synthesized by solid-state synthesis<sup class="xref">46</sup>. However, the long reaction time and high annealing temperature during the synthesis process resulted in large and irregular phosphors, which limited their application in other fields such as biomedicine. In 2007, Chermont et al. used sol-gel approach to synthesize nanoparticles for the first time and prepared Ca<sub>0.2</sub>Zn<sub>0.9</sub>Mg<sub>0.9</sub>Si<sub>2</sub>O<sub>6</sub>: Eu<sup>2+</sup>, Dy<sup>3+</sup>, Mn<sup>2+</sup>, which opened the era of PLNPs<sup class="xref">47</sup>. However, the top-down synthesis strategy is accompanied by problems such as uncontrollable size and morphology, so researchers have done a lot of work in the development of controllable bottom-up synthesis of PLNPs. Since 2015, various synthesis methods have emerged one after another, such as the template synthesis method, hydrothermal/solvent thermal method, sol-gel method and other wet chemical synthesis methods for the synthesis of uniform and controllable PLNPs<sup class="xref">47</sup><sup>,</sup><sup class="xref">48</sup><sup>,</sup><sup class="xref">49</sup><sup>,</sup><sup class="xref">50</sup>. Among them, hydrothermal synthesis is one of the most commonly used methods for preparing nanomaterials, which can provide an adjustable and mild synthetic method to prepare compounds or materials with special structures and properties<sup class="xref">51</sup>.</p>
<p class="jove_content">Here, we present a detailed experimental procedure for synthesizing Zn<sub>2</sub>GeO<sub>4</sub>: Mn PLNPs with 1D nanorods morphology via the hydrothermal method and providing them with a rigid environment for further illumination applications. It was found that the luminescence properties of PLNPs, including emission wavelength and afterglow decay curve, can be changed by adjusting the pH value of the precursor. On the other hand, to emphasize the versatility of this method, we also synthesize PLNPs with Cr as the luminescent center using ZnGa<sub>2</sub>O<sub>4</sub> as the matrix (ZnGa<sub>2</sub>O<sub>4</sub>: Cr), which exhibits afterglow emission (697 nm) in the near-infrared region after being excited by ultraviolet light (365 nm). This article mainly focuses on Zn<sub>2</sub>GeO<sub>4</sub>: Mn whose pH value of precursor solution is 9.4 for two-dimensional and three-dimensional artworks production and visualization. Zn<sub>2</sub>GeO<sub>4</sub>: Mn is a type of nanomaterial with Mn ions as the luminescent center which obtains strong green light emission (~ 537 nm) under the excitation of 365 nm ultraviolet light. At the same time, the continuous green light can still be seen after stopping excitation. In order to promote the polymerization of PLNPs in methyl methacrylate, ligands (Poly-ethylene glycol) were added during the hydrothermal synthesis process, and then PLNPs were polymerized with methyl methacrylate (MMA) in a two-dimensional or three-dimensional mold so that it can form glowing artwork while smoothly demolding.</p>
<p class="jove_content">This protocol provides a feasible method for the hydrothermal synthesis, polymerization reactions, and luminescent applications of PLNPs in advanced color rendering. Any differences in pH, temperature, and chemical reagents during nanocrystal growth will affect the size and optical properties of PLNP nanostructures. This detailed protocol aims to help new researchers in the field to improve the reproducibility of PLNPs using a hydrothermal method for further wider applications.</p>

<table><tbody><tr><td>azobisisobutyronitrile (99%)</td><td>Macklin</td><td>A800354</td><td>Further purification required</td></tr><tr><td>methyl methacrylate(99%)</td><td>Sigma-Aldrich</td><td>M55909</td><td>Further purification required</td></tr><tr><td>deionized water</td><td>Merck</td><td>ZEQ7016T0C</td><td>Milli-Q&amp;nbsp;Direct Water Purification System</td></tr><tr><td>alkaline aluminum oxide (100-200 mesh)</td><td>Macklin</td><td>A800033</td><td></td></tr><tr><td>&amp;nbsp;ammonium hydroxide&amp;nbsp; (25%-28%, wt)</td><td>Macklin</td><td>A801005</td><td></td></tr><tr><td>beaker&amp;nbsp;</td><td>Synthware</td><td>B220100</td><td></td></tr><tr><td>chromium(III) nitrate nonahydrate (99.95%)</td><td>Aladdin</td><td>C116448</td><td></td></tr><tr><td>centrifuge</td><td>ThermoFisher Scientific</td><td>75004250</td><td></td></tr><tr><td>column</td><td>Synthware</td><td>C184464CR</td><td></td></tr><tr><td>digital camera</td><td>&amp;nbsp;Canon</td><td>EOS M50 Mark II</td><td></td></tr><tr><td>electric thermostaticdrying oven</td><td>Longyue</td><td>LDO-9036A</td><td></td></tr><tr><td>ethanol (99.7%)</td><td>Greagent</td><td>1158566</td><td></td></tr><tr><td>gallium nitrate hydrate(99.9%)</td><td>Aladdin</td><td>G109501</td><td></td></tr><tr><td>germanium oxide (99.99%)</td><td>Sinopharm Chemical ReagentCo., Ltd</td><td>51009860</td><td></td></tr><tr><td>glass rod</td><td>Sinopharm Chemical ReagentCo., Ltd</td><td>91229401</td><td></td></tr><tr><td>powder X-Ray Diffractometer</td><td>D2 PHASER DESKTOP XRD</td><td>BRUKER</td><td></td></tr><tr><td>manganese nitrate (98%)</td><td>Macklin</td><td>M828399</td><td></td></tr><tr><td>methanol (99.5%)</td><td>Greagent</td><td>1226426</td><td></td></tr><tr><td>nitric acid (65.0-68.0%, wt)</td><td>Sinopharm Chemical ReagentCo., Ltd</td><td>10014508</td><td></td></tr><tr><td>pH meter</td><td>Shanghai Leici Sensor Technology Co., Ltd</td><td>PHS-3C</td><td></td></tr><tr><td>polyethylene glycol (300, Mw)</td><td>Adamas</td><td>01050882(41713A)</td><td></td></tr><tr><td>sealing film</td><td>Parafilm</td><td>2025722</td><td></td></tr><tr><td>sodium hydroxide (GR)</td><td>Sinopharm Chemical ReagentCo., Ltd</td><td>10019764</td><td></td></tr><tr><td>spectrometer</td><td>Horiba</td><td>Fluorolog-3&amp;nbsp;</td><td></td></tr><tr><td>transmission electron microscope</td><td>JEOL&amp;nbsp;</td><td>JEM-1400 Plus</td><td></td></tr><tr><td>transmission electron microscope</td><td>JEOL</td><td>2100 Plus&amp;nbsp;</td><td></td></tr><tr><td>triangular funnel</td><td>Synthware</td><td>F181975</td><td></td></tr><tr><td>ultrasound machine</td><td>centrifuge</td><td>JP-040S</td><td></td></tr><tr><td>zinc chloride (98%)</td><td>Greagent</td><td>01113266/G81783A</td><td></td></tr></tbody></table>

synthesis of persistent luminescent nanoparticles for rewritable displays and illumination applications

<p class="jove_content">Persistent luminescent nanoparticles (PLNPs) possess the capabilities to maintain extended longevity and robust emission even after the excitation has ceased. PLNPs have been widely used across various domains, including information displays, data encryption, biological imaging, and artistic decoration with sustained and vivid luminosity, providing boundless possibilities for a variety of innovative technology and artistic projects. This protocol focuses on an experimental procedure for the hydrothermal synthesis of PLNPs. The successful synthesis of enduring luminescent nanomaterials with Mn<sup>2+</sup> or Cr<sup>3+</sup> serving as a luminescent center in Zn<sub>2</sub>GeO<sub>4</sub>: Mn (ZGO: Mn) or ZnGa<sub>2</sub>O<sub>4</sub>: Cr highlights the universality of this synthetic method. On the other hand, the optical properties of ZGO: Mn can be changed by adjusting the pH of precursor solutions, demonstrating the tunability of the protocol. When charged with ultraviolet (UV) at a wavelength of 365 nm for 3 min and then stopped, PLNPs exhibit the remarkable capacity to generate afterglow efficiently and consistently, which makes them ideal for making two-dimensional rewritable displays and three-dimensional transparent, luminous artworks. This protocol outlined in this paper provides a feasible method for the synthesis of persistent luminescent nanoparticles for further illumination and imaging applications, opening up novel prospects for the fields of science and art.</p>

<p class="jove_content">The synthesis diagram of Zn<sub>2</sub>GeO<sub>4</sub>: Mn (ZGO: Mn) PLNPs is shown in <strong class="xfig">Figure 1</strong>. The amphiphilic polymer Poly-ethylene glycol (PEG) is added to modify the ligand-free Zn<sub>2</sub>GeO<sub>4</sub>: Mn (ZGO: Mn) nanorods to better dissolve in MMA medium. First, the transmission electron microscopy (TEM), high-resolution transmission electron microscopy (HRTEM) images of ZGO: Mn whose pH is 9.4 are collected (<strong class="xfig">Figure 1</strong>), and then dynamic light scatt...

Author Spotlight: Advancing Bioimaging and Therapy with Functional Nanomaterials

<p class="jove_content">A protocol is presented for the synthesis of persistent luminescent nanomaterials (PLNPs) and their potential applications in rewritable displays and artistic processing utilizing the afterglow effect under ultraviolet light (365 nm) irradiation. </p>

Synthesis of Persistent Luminescent Nanoparticles for Rewritable Displays and Illumination Applications

Persistent luminescent nanoparticles (PLNPs) possess the capabilities to maintain extended longevity and robust emission even after the excitation has ...

synthesis-persistent-luminescent-nanoparticles-for-rewritable

Research

JoVE Journal

Chemistry

1.8K Views.  Shanghai Tech University. A protocol is presented for the synthesis of persistent luminescent nanomaterials (PLNPs) and their potential applications in rewritable displays and artistic processing utilizing the afterglow effect under ultraviolet light (365 nm) irradiation. 

Video: Author Spotlight: Advancing Bioimaging and Therapy with Functional Nanomaterials

A 'Plug and Play' Method to Create Water-dispersible Nanoassemblies Containing an Amphiphilic Polymer, Organic Dyes and Upconverting Nanoparticles

<p class='jove_content'>In this protocol, we first describe a procedure to synthesize lanthanide doped upconverting nanoparticles (UCNPs). We then demonstrate how to generate amphiphilic polymers <em>in situ</em>, and describe a protocol to encapsulate the prepared UCNPs and different organic dye molecules (porphyrins and diarylethenes) using polymer shells to form stable water-dispersible nanoassemblies. The nanoassembly samples containing both the UCNPs and the diarylethene organic dyes have interesting photochemical and photophysical properties. Upon 365 nm UV irradiation, the diarylethene group undergoes a visual color change. When the samples are irradiated with visible light of another specific wavelength, the color fades and the samples return to the initial colorless state. The samples also emit visible light from the UCNPs upon irradiation with 980 nm near-infrared light. The emission intensity of the samples can be tuned through alternate irradiation with UV and visible light. Modulation of fluorescence can be performed for many cycles without observable degradation of the samples. This versatile encapsulation procedure allows for the transfer of hydrophobic molecules and nanoparticles from an organic solvent to an aqueous medium. The polymer helps to maintain a lipid-like microenvironment for the organic molecules to aid in preservation of their photochemical behavior in water. Thus this method is ideal to prepare water-dispersible photoresponsive systems. The use of near-infrared light to activate upconverting nanoparticles allows for lower energy light to be used to activate photoreactions instead of more harmful ultraviolet light. </p>

<p class='jove_content'>Organic dye molecules and oleic acid coated upconverting nanoparticles are not water-soluble. This protocol describes a &#8216;plug and play&#8217; method that enables the transfer of organic dye molecules and upconverting particles from their initial hydrophobic solvent to water. </p>

In this protocol, we first describe a procedure to synthesize lanthanide doped upconverting nanoparticles (UCNPs). We then demonstrate how to generate ...

Low-energy Cathodoluminescence for (Oxy)Nitride Phosphors

<p class="jove_content">Nitride and oxynitride (Sialon) phosphors are good candidates for the ultraviolet and visible emission applications. High performance, good stability and flexibility of their emission properties can be achieved by controlling their composition and dopants. However, a lot of work is still required to improve their properties and to reduce the production cost. A possible approach is to correlate the luminescence properties of the Sialon particles with their local structural and chemical environment in order to optimize their growth parameters and find novel phosphors. For such a purpose, the low-voltage cathodoluminescence (CL) microscopy is a powerful technique. The use of electron as an excitation source allows detecting most of the luminescence centers, revealing their luminescence distribution spatially and in depth, directly comparing CL results with the other electron-based techniques, and investigating the stability of their luminescence properties under stress. Such advantages for phosphors characterization will be highlighted through examples of investigation on several Sialon phosphors by low-energy CL.</p>

Low-energy Cathodoluminescence for OxyNitride Phosphors

<p class="jove_content">An excellent chemical and luminescence stabilities of (oxy)nitride phosphors present it as an promising alternative to currently used sulfide and oxide phosphors. In this paper, we present the way to investigate its local luminescence properties using low-energy cathodoluminescence (CL).</p>

Nitride and oxynitride (Sialon) phosphors are good candidates for the ultraviolet and visible emission applications. High performance, good stability and ...

Facile Synthesis of Worm-like Micelles by Visible Light Mediated Dispersion Polymerization Using Photoredox Catalyst

<p class='jove_content'>Presented herein is a protocol for the facile synthesis of worm-like micelles by visible light mediated dispersion polymerization. This approach begins with the synthesis of a hydrophilic poly(oligo(ethylene glycol) methyl ether methacrylate) (POEGMA) homopolymer using reversible addition-fragmentation chain-transfer (RAFT) polymerization. Under mild visible light irradiation (&#955; = 460 nm, 0.7 mW/cm<sup>2</sup>), this macro-chain transfer agent (macro-CTA) in the presence of a ruthenium based photoredox catalyst, Ru(bpy)<sub>3</sub>Cl<sub>2</sub> can be chain extended with a second monomer to form a well-defined block copolymer in a process known as Photoinduced Electron Transfer RAFT (PET-RAFT). When PET-RAFT is used to chain extend POEGMA with benzyl methacrylate (BzMA) in ethanol (EtOH), polymeric nanoparticles with different morphologies are formed <em>in situ</em> according to a polymerization-induced self-assembly (PISA) mechanism. Self-assembly into nanoparticles presenting POEGMA chains at the corona and poly(benzyl methacrylate) (PBzMA) chains in the core occurs <em>in situ</em> due to the growing insolubility of the PBzMA block in ethanol. Interestingly, the formation of highly pure worm-like micelles can be readily monitored by observing the onset of a highly viscous gel <em>in situ</em> due to nanoparticle entanglements occurring during the polymerization. This process thereby allows for a more reproducible synthesis of worm-like micelles simply by monitoring the solution viscosity during the course of the polymerization. In addition, the light stimulus can be intermittently applied in an ON/OFF manner demonstrating temporal control over the nanoparticle morphology. </p>

<p class='jove_content'>This article describes a process for producing polymeric self-assembled nanoparticles using visible light mediated dispersion polymerization. Using low energy visible light to control the polymerization allows for the reproducible formation of self-assembled worm-like micelles at high solids content.</p>

Presented herein is a protocol for the facile synthesis of worm-like micelles by visible light mediated dispersion polymerization. This approach begins ...

Fabrication of White Light-emitting Electrochemical Cells with Stable Emission from Exciplexes

<p class="jove_content">The authors present an approach for fabricating stable white light emission from polymer light-emitting electrochemical cells (PLECs) having an active layer which consists of blue-fluorescent poly(9,9-di-n-dodecylfluorenyl-2,7-diyl) (PFD) and &#960;-conjugated triphenylamine molecules. This white light emission originates from exciplexes formed between PFD and amines in electronically excited states. A device containing PFD, 4,4&#39;,4&#39;&#39;-tris[2-naphthyl(phenyl)amino]triphenylamine (2-TNATA), Poly(ethylene oxide) and K<sub>2</sub>CF<sub>3</sub>SO<sub>3</sub> showed white light emission with Commission internationale de l&#39;&#233;clairage (CIE) coordinates of (0.33, 0.43) and a Color Rendering Index (CRI) of Ra = 73 at an applied voltage of 3.5 V. Constant voltage measurements showed that the CIE coordinates of (0.27, 0.37), Ra of 67, and the emission color observed immediately after application of a voltage of 5 V were nearly unchanged and stable after 300 sec.</p>

<p class='jove_content'>The authors present a method for fabricating stable white-light-emitting electrochemical cells utilizing emission from exciplexes formed between a blue-emitting fluorene polymer and aromatic amines.</p>

The authors present an approach for fabricating stable white light emission from polymer light-emitting electrochemical cells (PLECs) having an active ...

Engineering

Synthesis of Core-shell Lanthanide-doped Upconversion Nanocrystals for Cellular Applications

<p class="jove_content">Lanthanide-doped upconversion nanocrystals (UCNs) have attracted much attention in recent years based on their promising and controllable optical properties, which allow for the absorption of near-infrared (NIR) light and can subsequently convert it into multiplexed emissions that span over a broad range of regions from the UV to the visible to the NIR. This article presents detailed experimental procedures for high-temperature co-precipitation synthesis of core-shell UCNs that incorporate different lanthanide ions into nanocrystals for efficiently converting deep-tissue penetrable NIR excitation (808 nm) into a strong blue emission at 480 nm. By controlling the surface modification with biocompatible polymer (polyacrylic acid, PAA), the as-prepared UCNs acquires great solubility in buffer solutions. The hydrophilic nanocrystals are further functionalized with specific ligands (dibenzyl cyclooctyne, DBCO) for localization on the cell membrane. Upon NIR light (808 nm) irradiation, the upconverted blue emission can effectively activate the light-gated channel protein on the cell membrane and specifically regulate the cation (<em>e.g.</em>, Ca<sup>2+</sup>) influx in the cytoplasm. This protocol provides a feasible methodology for the synthesis of core-shell lanthanide-doped UCNs and subsequent biocompatible surface modification for further cellular applications.</p>

<p class="jove_content">A protocol is presented for the synthesis of core-shell lanthanide-doped upconversion nanocrystals (UCNs) and their cellular applications for channel protein regulation upon near-infrared (NIR) light illumination.</p>

Lanthanide-doped upconversion nanocrystals (UCNs) have attracted much attention in recent years based on their promising and controllable optical ...

Scale-up Chemical Synthesis of Thermally-activated Delayed Fluorescence Emitters Based on the Dibenzothiophene-S,S-Dioxide Core

<p class="jove_content">We report a procedure to linearly scale-up the synthesis of 2,8-bis(3,6-di-tert-butyl-9H-carbazol-9-yl)dibenzothiophene-S,S-dioxide (compound 4) and 2,8-bis(10H-phenothiazin-10-yl)dibenzothiophene-S,S-dioxide (compound 5) using Buchwald-Hartwig cross-coupling reaction conditions. In addition, we demonstrate a scaled-up synthesis of all non-commercially available starting materials that are required for the amination cross-coupling reaction. In the present article, we provide the detailed synthetic procedures for all of the described compounds, alongside their spectral characterization. This work shows the possibility to produce organic molecules for optoelectronic applications on a large scale, which facilitates their implementation into real world devices.</p>

<p class="jove_content">Scale-up synthesis of highly efficient thermally activated delayed fluorescence emitters is described in the presented article.</p>

We report a procedure to linearly scale-up the synthesis of 2,8-bis(3,6-di-tert-butyl-9H-carbazol-9-yl)dibenzothiophene-S,S-dioxide (compound 4) and ...

A Step Beyond BRET: Fluorescence by Unbound Excitation from Luminescence (FUEL)

<p class="jove_content">Fluorescence by Unbound Excitation from Luminescence (FUEL) is a radiative excitation-emission process that produces increased signal and contrast enhancement <em>in vitro</em> and <em>in vivo</em>. FUEL shares many of the same underlying principles as Bioluminescence Resonance Energy Transfer (BRET), yet greatly differs in the acceptable working distances between the luminescent source and the fluorescent entity. While BRET is effectively limited to a maximum of 2 times the F&ouml;rster radius, commonly less than 14 nm, FUEL can occur at distances up to &micro;m or even cm in the absence of an optical absorber. Here we expand upon the foundation and applicability of FUEL by reviewing the relevant principles behind the phenomenon and demonstrate its compatibility with a wide variety of fluorophores and fluorescent nanoparticles. Further, the utility of antibody-targeted FUEL is explored. The examples shown here provide evidence that FUEL can be utilized for applications where BRET is not possible, filling the spatial void that exists between BRET and traditional whole animal imaging.</p>

A Step Beyond BRET: Fluorescence by Unbound Excitation from Luminescence FUEL

<p class="jove_content">Expanding the foundation and applicability of Fluorescence by Unbound Excitation from Luminescence (FUEL) by surveying the relevant principles and demonstrating its compatibility with a multitude of fluorophores and antibody-targeted conditions.</p>

Fluorescence by Unbound Excitation from Luminescence (FUEL) is a radiative excitation-emission process that produces increased signal and contrast ...

Bioengineering

Production and Characterization of Vacuum Deposited Organic Light Emitting Diodes

<p class="jove_content">A method for producing simple and efficient thermally-activated delayed fluorescence organic light-emitting diodes (OLEDs) based on guest-host or exciplex donor-acceptor emitters is presented. With a step-by-step procedure, readers will be able to repeat and produce OLED devices based on simple organic emitters. A patterning procedure allowing the creation of personalized indium tin oxide (ITO) shape is shown. This is followed by the evaporation of all layers, encapsulation and characterization of each individual device. The end goal is to present a procedure that will give the opportunity to repeat the information presented in cited publication but also using different compounds and structures in order to prepare efficient OLEDs.</p>

<p class="jove_content">A protocol for the production of simple structured organic light-emitting diodes (OLEDs) is presented.</p>

A method for producing simple and efficient thermally-activated delayed fluorescence organic light-emitting diodes (OLEDs) based on guest-host or exciplex ...

Enhanced Electron Injection and Exciton Confinement for Pure Blue Quantum-Dot Light-Emitting Diodes by Introducing Partially Oxidized Aluminum Cathode

<p class="jove_content">Stable and efficient red (R), green (G), and blue (B) light sources based on solution-processed quantum dots (QDs) play important roles in next-generation displays and solid-state lighting technologies. The brightness and efficiency of blue QDs-based light-emitting diodes (LEDs) remain inferior to their red and green counterparts, due to the inherently unfavorable energy levels of different colors of light. To solve these problems, a device structure should be designed to balance the injection holes and electrons into the emissive QD layer. Herein, through a simple autoxidation strategy, pure blue QD-LEDs which are highly bright and efficient are demonstrated, with a structure of ITO/PEDOT:PSS/Poly-TPD/QDs/Al:Al<sub>2</sub>O<sub>3</sub>. The autoxidized Al:Al<sub>2</sub>O<sub>3</sub> cathode can effectively balance the injected charges and enhance radiative recombination without introducing an additional electron transport layer (ETL). As a result, high color-saturated blue QD-LEDs are achieved with a maximum luminance over 13,000 cd m<sup>-2</sup>, and a maximum current efficiency of 1.15 cd A<sup>-1</sup>. The easily controlled autoxidation procedure paves the way for achieving high-performance blue QD-LEDs.</p>

<p class="jove_content">A protocol is presented for fabricating high-performance, pure blue ZnCdS/ZnS-based quantum dots light-emitting diodes by employing an autoxidized aluminum cathode.</p>

Stable and efficient red (R), green (G), and blue (B) light sources based on solution-processed quantum dots (QDs) play important roles in next-generation ...

Synthesis of Luminol

<p>Source: Lara Al Hariri and Ahmed Basabrain at the University of Massachusetts Amherst, MA, USA</p>
<ol class="note-items">
	<li data-note-item="1" data-parent-collapse="">Synthesis of 3-Nitrophthalhydrazide<button class="expand">Expand</button>
	<p>In this lab, you&#39;ll synthesize 3-aminophthalhydrazide, which is also called luminol, in a 2-step process. The first step is a condensation reaction between 3-nitrophthalic acid and hydrazine.</p>
	<p>During this step, the carboxylic acid groups are substituted with NH groups to produce 3-nitrophthalhydrazide and two water molecules. You must work in a fume hood throughout this lab because the reactions produce toxic vapors and gases.</p>
	<ul class="lab-items">
		<li data-sub-note-item="1-3">
		<div class="lab-step">Before you get started, put on a lab coat, splash-proof safety glasses, and nitrile gloves. <strong>Note:</strong> Hydrazine is highly toxic and flammable, so avoid touching it and keep anything containing hydrazine closed or covered when you transport it between fume hoods.</div>
		</li>
		<li data-sub-note-item="1-4">
		<div class="lab-step">Now measure 10 mL of deionized water and pour it into a 25-mL Erlenmeyer flask. Clamp the flask on a hot plate and place a thermometer in it.</div>
		</li>
		<li data-sub-note-item="1-5">
		<div class="lab-step">Then, start heating the water. You&#39;ll want the temperature of the water to be 80 &#176;C, so set the hot plate setting to slightly higher.</div>
		</li>
		<li data-sub-note-item="1-6">
		<div class="lab-step">While it heats, set up a Bunsen burner under a clamp fixed on a lab stand.</div>
		</li>
		<li data-sub-note-item="1-7">
		<div class="lab-step">Obtain a 25-mL test tube and adjust the clamp so that it will hold the test tube about 5 &#8211; 7 cm above the flame. Then, remove the test tube and place it in a 400-mL beaker.</div>
		</li>
		<li data-sub-note-item="1-8">
		<div class="lab-step">Once the water reaches 80 &#176;C, adjust the heat setting to keep the temperature stable.</div>
		</li>
		<li data-sub-note-item="1-9">
		<div class="lab-step">Then, weigh 0.6 g of 3-nitrophthalic acid in a tared weighing boat. Pour the 3-nitrophthalic acid into your test tube. Then, bring the test tube and a test tube stopper to the solvent hood.</div>
		</li>
		<li data-sub-note-item="1-10">
		<div class="lab-step">Use the provided volumetric pipette to carefully measure 0.9 mL of the 10% v/v hydrazine solution and transfer it to the test tube. Remember to cap the bottle of hydrazine solution and stopper your test tube before you return to your hood.</div>
		</li>
		<li data-sub-note-item="1-11">
		<div class="lab-step">Clamp the test tube over the Bunsen burner and remove the stopper. Then, pour ~2 mL of triethylene glycol into a graduated cylinder and set it out of the way in your hood.</div>
		</li>
		<li data-sub-note-item="1-12">
		<div class="lab-step">Now, add 2 or 3 boiling chips to the test tube. Place a high-temperature thermometer in the test tube and ignite the Bunsen burner. Heat the mixture with a medium flame until the 3-nitrophthalic acid dissolves.</div>
		</li>
		<li data-sub-note-item="1-13">
		<div class="lab-step">Then, add the triethylene glycol and increase the heat of the flame. <strong>Note:</strong> Once the water from the hydrazine solution has boiled off, the temperature will continue increasing.</div>
		</li>
		<li data-sub-note-item="1-14">
		<div class="lab-step">When the reaction mixture reaches 210 &#176;C, turn down the heat and keep the mixture between 210 &#176;C and 220 &#176;C for 3 &#8211; 4 min.</div>
		</li>
		<li data-sub-note-item="1-15">
		<div class="lab-step">Then, extinguish the flame and wait for the mixture to cool to 100 &#176;C.</div>
		</li>
		<li data-sub-note-item="1-16">
		<div class="lab-step">Turn off the hotplate, remove the thermometer, and unclamp the flask. Use tongs to hold the flask and measure 8 mL of hot water with the graduated cylinder. Pour this hot water into the test tube and remove the thermometer.</div>
		</li>
		<li data-sub-note-item="1-17">
		<div class="lab-step">Obtain a piece of plastic paraffin film and cover the mouth of the test tube. Then, use tongs to transfer the test tube to the holding beaker.</div>
		</li>
		<li data-sub-note-item="1-18">
		<div class="lab-step">At a lab sink, hold the test tube under running tap water to cool it, being careful not to let water into it.</div>
		</li>
		<li data-sub-note-item="1-19">
		<div class="lab-step">Then, dry the outside of the tube and bring it back to your hood. Clamp the tube upright and remove the paraffin film. Let the yellow 3-nitrophthalhydrazide precipitate in the tube for 15 min.</div>
		</li>
		<li data-sub-note-item="1-20">
		<div class="lab-step">While you wait, add ~10 mL of deionized water to a 25-mL Erlenmeyer flask. Prepare an ice bath in a 250-mL beaker and start chilling the water in it.</div>
		</li>
		<li data-sub-note-item="1-21">
		<div class="lab-step">Then, set up for vacuum filtration using a 250-mL filter flask, a B&#252;chner funnel, and a piece of circular filter paper.</div>
		</li>
		<li data-sub-note-item="1-22">
		<div class="lab-step">Once the product has precipitated for at least 15 min, obtain a Pasteur pipette and wet the filter paper with a few drops of cold water.</div>
		</li>
		<li data-sub-note-item="1-23">
		<div class="lab-step">Then, turn on the vacuum and pour the contents of the test tube into the B&#252;chner funnel. Rinse the remaining solid into the funnel with 1 &#8211; 2 mL of cold water.</div>
		</li>
		<li data-sub-note-item="1-24">
		<div class="lab-step">Remove the boiling chips with tweezers and rinse them with a few drops of cold water to finish collecting the product.</div>
		</li>
		<li data-sub-note-item="1-25">
		<div class="lab-step">Turn off the vacuum pump once no more liquid is dripping into the flask from the funnel.</div>
		</li>
	</ul>
	</li>
	<li data-note-item="2" data-parent-collapse="">Synthesis of 3-Aminopthalhydrazide<button class="expand">Expand</button>
	<p>In the second step of the luminol synthesis, you&#39;ll reduce the nitro group of 3-nitrophthalhydrazide using sodium dithionite in a basic solution of NaOH. This will produce 3-aminophthalhydrazide dianion.</p>
	<p>After the reduction, you&#39;ll add acetic acid to protonate the dianion, forming luminol. Sodium dithionite is highly reactive and degrades quickly, so you&#39;ll use a slight excess of it. After the reduction, you&#39;ll protonate the luminol dianion to decrease its solubility in water. Lastly, you&#39;ll precipitate and collect luminol as a yellow solid.</p>
	<ul class="lab-items">
		<li data-sub-note-item="2-3">
		<div class="lab-step">Place a clean 25-mL test tube in a beaker and use a clean spatula to transfer your 3-nitrophthalhydrazide to the test tube.</div>
		</li>
		<li data-sub-note-item="2-4">
		<div class="lab-step">Set the B&#252;chner funnel aside and clamp the test tube over the Bunsen burner. Leave the filter flask in place for later.</div>
		</li>
		<li data-sub-note-item="2-5">
		<div class="lab-step">Now, use a 10-mL graduated cylinder to obtain 3 mL of approximately 3 M NaOH. Pour the sodium hydroxide solution into the test tube and stir the mixture with a glass rod until the 3-nitrophthalhydrazide dissolves.</div>
		</li>
		<li data-sub-note-item="2-6">
		<div class="lab-step">Now, weigh 2 g of sodium dithionite in a tared weighing boat and add it to the test tube. Obtain 3 mL of deionized water and pour it into the tube.</div>
		</li>
		<li data-sub-note-item="2-7">
		<div class="lab-step">Stir the mixture well to dissolve as much of the solid as you can. Then, add a few clean boiling chips to the test tube and ignite the Bunsen burner.</div>
		</li>
		<li data-sub-note-item="2-8">
		<div class="lab-step">Heat the mixture to boiling over a medium flame and then let it boil for 5 min.</div>
		</li>
		<li data-sub-note-item="2-9">
		<div class="lab-step">While you wait, measure 1.5 mL of glacial acetic acid and bring it to your hood.</div>
		</li>
		<li data-sub-note-item="2-10">
		<div class="lab-step">Once the reaction mixture has boiled for 5 min, shut off the Bunsen burner and add the acetic acid to the tube. Let the mixture cool to room temperature, which usually takes 10 &#8211;15 minutes. Meanwhile, prepare an ice bath in a 600-mL beaker.</div>
		</li>
		<li data-sub-note-item="2-11">
		<div class="lab-step">Once the mixture has cooled, place it in the ice bath and wait 15 min for the luminol to precipitate.</div>
		</li>
		<li data-sub-note-item="2-12">
		<div class="lab-step">Reassemble the vacuum filtration setup using a clean B&#252;chner funnel and a new piece of filter paper, and start cooling another 10 mL of deionized water in an ice bath.</div>
		</li>
		<li data-sub-note-item="2-13">
		<div class="lab-step">Once the luminol has been in the ice bath for at least 15 min, wet the filter paper with cold water. Then, collect the solid luminol by vacuum filtration.</div>
		</li>
		<li data-sub-note-item="2-14">
		<div class="lab-step">Retrieve the boiling chips and rinse the luminol with more cold water. Once liquid stops dripping from the funnel, turn off the vacuum and transfer the luminol to a 50-mL beaker.</div>
		</li>
		<li data-sub-note-item="2-15">
		<div class="lab-step">Now, prepare the combined filtrate for disposal. First, label a 250-mL beaker &#8216;hydrazine and dithionite waste&#8217;. Transfer the filtrate to the beaker and rinse the flask with deionized water to remove residual hydrazine and dithionite.</div>
		</li>
		<li data-sub-note-item="2-16">
		<div class="lab-step">Next, dilute the filtrate with 10 mL of deionized water. Then, measure 10 mL of 1 M sodium carbonate and add it to the combined filtrates.</div>
		</li>
		<li data-sub-note-item="2-17">
		<div class="lab-step">Bring a 50-mL graduated cylinder to the waste hood. Measure 40 mL of 10% sodium hypochlorite and pour it into the filtrate mixture. Place the beaker on the hot plate, add a stir bar, and secure a thermometer in the beaker.</div>
		</li>
		<li data-sub-note-item="2-18">
		<div class="lab-step">Heat the mixture to 50 &#176;C while stirring. The mixture must stay at 50 &#176;C for 1 h to oxidize leftover hydrazine and dithionite, so make sure that the temperature is stable and set a timer for 1 h before starting the last part of the lab.</div>
		</li>
	</ul>
	</li>
	<li data-note-item="3" data-parent-collapse="">Chemiluminescence and&#160;Fluorescence<button class="expand">Expand</button>
	<p>For the last part of the lab, you&#39;ll mix some of the luminol you made with dimethyl sulfoxide over solid potassium hydroxide. The hydroxyl ions deprotonate the two amine groups. Then, ambient oxygen oxidizes the luminol dianion to 3-aminophthalate, or 3-APA, in an excited state. This unstable complex will quickly relax to the ground state, releasing energy as visible light. Since the excited 3-APA was the product of a chemical reaction, the emitted light is called chemiluminescence.</p>
	<p>Once you have seen the chemiluminescence from the oxidation reaction, you&#39;ll add fluorescein, a fluorescent molecule, to the mixture. Some excited 3-APA will transfer energy directly to fluorescein rather than emitting light, giving you a solution with two different light-emitting compounds.</p>
	<ul class="lab-items">
		<li data-sub-note-item="3-3">
		<div class="lab-step">Now, measure about 0.2 g of the luminol you made and place it in a 15-mL Erlenmeyer flask. It&#39;s OK if the solid is still damp.</div>
		</li>
		<li data-sub-note-item="3-4">
		<div class="lab-step">Obtain 5 &#8211; 7 pellets of potassium hydroxide and pour them into the flask. Then, bring the flask, a stopper, and a 10-mL graduated cylinder to the solvent hood.</div>
		</li>
		<li data-sub-note-item="3-5">
		<div class="lab-step">Measure 2 mL of DMSO, add it to the flask, and stopper it. Put the graduated cylinder in your fume hood and bring the flask to a dark room.</div>
		</li>
		<li data-sub-note-item="3-6">
		<div class="lab-step">Shake the stoppered flask vigorously until you see chemiluminescence and then put the flask down. The light fades quickly because both the oxidation and relaxation are fast processes.</div>
		</li>
		<li data-sub-note-item="3-7">
		<div class="lab-step">Shake the flask and let the light fade 6 &#8211; 10 more times without opening the flask. The light will be progressively dimmer as the oxygen in the flask is consumed. <strong>Note:</strong> To make it brighter, leave the flask open in your fume hood for a minute to replenish the oxygen.</div>
		</li>
		<li data-sub-note-item="3-8">
		<div class="lab-step">Close the flask, return to the dark room, and shake the flask again to see the effect.</div>
		</li>
		<li data-sub-note-item="3-9">
		<div class="lab-step">Now, measure 0.2 g of fluorescein dye and add it to your flask. Stopper the flask and return to the dark room. Shake the flask vigorously until the mixture is glowing brightly and then set it down to observe the difference in color from luminol alone.</div>
		</li>
		<li data-sub-note-item="3-10">
		<div class="lab-step">When you&#39;re finished, check on the filtrate. Once it has been at 50 &#176;C for at least 1 h, turn off the hotplate and wait for the beaker to cool to room temperature.</div>
		</li>
		<li data-sub-note-item="3-11">
		<div class="lab-step">Then, add about 10 mL of tap water to the filtrate and flush it down the drain.</div>
		</li>
		<li data-sub-note-item="3-12">
		<div class="lab-step">Dispose of the remaining chemical and glass waste in the appropriate containers, clean your glassware and tools, and put away your lab equipment.</div>
		</li>
	</ul>
	</li>
	<li data-note-item="4" data-parent-collapse="">Results<button class="expand">Expand</button>
	<p>Under your oxidation reaction conditions, excited 3-APA emits blue-green light in a broad range around 500 nm when it relaxes, and fluorescein is excited by absorbing light at 480 &#8211; 490 nm and 515 &#8211; 525 nm.</p>
	<p>When you add fluorescein to the luminol oxidation reaction mixture, excited 3-APA can transfer energy that fluorescein would absorb as light directly to a nearby fluorescein molecule in a special interaction called nonradiative energy transfer. The resulting excited fluorescein emits yellow-green light when it relaxes.</p>
	<p>Excited 3-APA is highly unstable, so if no fluorescein is close enough for nonradiative energy transfer, it will relax by emitting light as usual. Thus, a spectrum of your final glowing mixture would show contributions from both blue-green and yellow-green light.</p>
	<ul class="lab-items">
	</ul>
	</li>
</ol>

Synthesis of Luminol and Chemiluminescence - Procedure

Source: Lara Al Hariri and Ahmed Basabrain at the University of Massachusetts Amherst, MA, USA

	Synthesis of 3-NitrophthalhydrazideExpand
	In this lab, ...