A start-up grant from Ariel University and an ISF Individual Research Grant (1914/15) to AMS is gratefully acknowledged.

1. Preparation of the Enolonium Species
Caution: Before carrying out the protocol, consult the MSDS for all reagents and solvents.
NOTE: All new reagents were used as received from the commercial source. If the boron trifluoride etherate has been stored, distill it before use.
<ol>
	<li>In a dry round bottomed flask equipped with a septum and a magnet for magnetic stirring, add Koser&#39;s reagent (1.5 equiv.) and flush the flask with nitrogen or argon.</li>
	<li>Add dry dichloromethane&#160;to give a suspension of 0.234 mol/L formal concentration.</li>
	<li>Cool the suspension to -78 &#176;C using a dry ice/acetone bath or a cold finger instrument/acetone bath.</li>
	<li>Add neat BF3OEt2 (1.5 equiv.) slowly.</li>
	<li>Warm the heterogeneous mixture to room temperature until the formation of yellow solution. Typically, this happens within 5 min.</li>
	<li>Cool the solution to -78 &#176;C.</li>
	<li>To the cooled solution, add the trimethylsilyl-enolether (1 equiv., 0.313 mol/L) in dry dichloromethane dropwise over 2-10 min (depending on the scale). After the addition of silyl enol ether is complete, the formation of the enolonium species is complete. 
	NOTE: The solution of enolonium species can be left at -78 &#176;C for at least 30 min with no deterioration in yield. The enolonium species is stable during this time as indicated by reported NMR studies6.</li>
</ol>
2. Functionalization of the Enolonium Species
<ol>
	<li>Chlorination with the chloride anion
	<ol>
		<li>To the prepared solution of enolonium species, add benzyl-dimethyl-decylammonium chloride (2.0 equiv., 1.25 mol/L) in dry dichloromethane in a drop wise manner. Add this solution at a rate such that the temperature remains below -55 &#176;C. At the 0.5-2 mmol scale, addition over 5 min is satisfactory.</li>
		<li>Leave the reaction mixture at -78 &#176;C for 5 min.</li>
		<li>Remove the cooling bath and allow the reaction mixture to reach room temperature.</li>
		<li>Leave the reaction at room temperature for 20 min.</li>
		<li>Add water to the reaction mixture (half the volume of dichloromethane used in the preparation of the enolonium species).</li>
		<li>Extract thrice with dichloromethane. Typically, use 2-3 times the reaction volume in each extraction at the 0.5-2 mmol small scale.</li>
		<li>Wash the combined organic layers twice with brine. Typically, use the same volume of brine as the combined reaction volume.</li>
		<li>Dry with anhydrous sodium sulfate for 30 min.</li>
		<li>Filter off the sodium sulfate (e.g., through a Celite plug).</li>
		<li>Remove the solvent on a rotary evaporator at reduced pressure and at 40 &#176;C.</li>
		<li>Purify the crude product by column chromatography on silica gel using hexane and ethyl acetate eluents to afford, after removal of the solvents, the pure corresponding &#945;-azido ketone. 
		Note: At scales of 0.5 mmol to 2 mmol of trimethylsilyl enolate, carry out column chromatography on a glass column of 2 cm diameter using standard silica gel 60 at a height (length) of 15 cm. The volume will need to be varied for other scales.</li>
	</ol>
	</li>
	<li>Azidation with TMS-Azide 
	Caution: Organic-azides in general are explosive and care should be taken in handling and preparing the products. TMS-azide is toxic. Consult MSDS before use.
	<ol>
		<li>To the prepared solution of enolonium species at -78 &#176;C, add neat azidotrimethylsilane (2.5 equiv.) in a dropwise fashion. Add this solution at a rate so that the temperature remains below -55 &#176;C. At the 0.5-2 mmol scale, addition over 2-3 min is satisfactory.</li>
		<li>Stir the reaction mixture for 15 min at -78 &#176;C.</li>
		<li>Heat the reaction mixture to -55 &#176;C and leave at this temperature for 2 to 3 h.</li>
		<li>Add water to the reaction mixture (half the volume of dichloromethane used in the preparation of the enolonium species).</li>
		<li>Extract thrice with dichloromethane. Typically, use 2-3 times the reaction volume in each extraction at the 0.5-2 mmol small scale.</li>
		<li>Wash the combined organic layers twice with brine. Typically, use the same volume of brine as the combined reaction volume.</li>
		<li>Dry the extracts with anhydrous sodium sulfate for 30 min.</li>
		<li>Filter off the sodium sulfate.</li>
		<li>Remove the solvent on a rotary evaporator at reduced pressure and at 40 &#176;C.</li>
		<li>Purify the crude product by column chromatography on silica gel using hexane and ethyl acetate eluents to afford, after removal of the solvents, the pure corresponding &#945;-azido ketone.</li>
	</ol>
	</li>
	<li>Reaction with azoles
	<ol>
		<li>To the prepared solution of enolonium species at -78 &#176;C, add azole (4 to 5 equiv., 1 mol/L) dissolved in 5 mL of dichloromethane in a dropwise fashion. At the 0.5-2 mmol scale, addition over 5 min is satisfactory. 
		Note: In the case of poorly soluble azoles such as tetrazoles, use acetonitrile at a concentration of 0.5 mol/L instead of dichloromethane. Add this solution at a rate so that the temperature remains below -55 &#176;C.</li>
		<li>Stir the reaction mixture for 15 min at -78 &#176;C.</li>
		<li>Heat the reaction mixture to -55 &#176;C and leave at this temperature for 4 to 8 h.</li>
		<li>Add water to the reaction mixture (half the volume of organic solvents used in the preparation of the enolonium species).</li>
		<li>Extract thrice with dichloromethane. Typically, use 2-3 times the reaction volume in each extraction at the 0.5-2 mmol small scale.</li>
		<li>Wash the combined organic layers twice with brine. Typically, use the same volume of brine as the combined reaction volume.</li>
		<li>Dry the extracts with anhydrous sodium sulfate for 30 min.</li>
		<li>Filter off the sodium sulfate.</li>
		<li>Remove the solvent on a rotary evaporator at reduced pressure and at 40 &#176;C.</li>
		<li>Purify the crude product by column chromatography on silica gel using hexane and ethyl acetate eluents to afford, after removal of the solvents, the pure corresponding &#945;-azole ketone.</li>
	</ol>
	</li>
	<li>Allylation, crotylation, cinnamylation, and prenylation using allyl-silanes
	<ol>
		<li>Add neat allyl,-, crotyl-, cinnamyl-, or prenyl-trimethylsilane (2 equiv.) slowly at -78 &#176;C. Add this solution at a rate so that the temperature remains below -55 &#176;C. At a 0.5-2 mmol scale, addition over 2-3 min is satisfactory.</li>
		<li>Stir the reaction mixture for 10 min at -78 &#176;C.</li>
		<li>Allow the reaction mixture to warm slowly to room temperature by removing the cooling bath. Leave the reaction at room temperature for 20 min.</li>
		<li>Add water to the reaction mixture (half the volume of dichloromethane used in the preparation of the enolonium species).</li>
		<li>Extract thrice with dichloromethane. Typically, use 2-3 times the reaction volume in each extraction at the 0.5-2 mmol small scale.</li>
		<li>Wash the combined organic layers twice with brine. Typically, use the same volume of brine as the combined reaction volume.</li>
		<li>Dry the extracts with anhydrous sodium sulfate for 30 min.</li>
		<li>Filter off the sodium sulfate.</li>
		<li>Remove the solvent on a rotary evaporator at reduced pressure and at 40 &#176;C.</li>
		<li>Purify the crude product by column chromatography on silica gel using hexane and ethyl acetate eluents to afford, after removal of the solvents, the pure corresponding &#945;-allyl product.</li>
	</ol>
	</li>
	<li>Arylation 
	Note: For arylation, use 3 equivalents of BF3OEt2 during the preparation of the enolonium species in order to avoid tosylation of the enolonium species as a major side reaction. In general, only 1.6 equivalent of the aromatic substrate is needed. However, if the aromatic substrate is a pyrane, thiophene, or pyrrole, the best results are achieved using 5 equivalents of the aromatic substrate.
	<ol>
		<li>To the solution of prepared enolonium species add a solution of aromatic substrate in dry dichloromethane (1.6 equiv., 0.5 mol/L) in a dropwise manner. Add this solution at a rate so that the temperature remains below -55 &#176;C. At the 0.5-2 mmol scale, addition over 5-10 min is satisfactory.</li>
		<li>After the addition of the aromatic substrate is complete, increase the temperature of the mixture to -55 &#176;C and leave the mixture at this temperature for 20 min.</li>
		<li>Add water to the reaction mixture (half the volume of dichloromethane used in the preparation of the enolonium species).</li>
		<li>Extract thrice with dichloromethane. Typically, use 2-3 times the reaction volume in each extraction at the 0.5-2 mmol small scale.</li>
		<li>Wash the combined organic layers twice with brine. Typically, use the same volume of brine as the combined reaction volume.</li>
		<li>Dry the extracts with anhydrous sodium sulfate for 30 min.</li>
		<li>Filter off the sodium sulfate.</li>
		<li>Remove the solvent on a rotary evaporator at reduced pressure and at 40 &#176;C.</li>
		<li>Purify the crude product by column chromatography on silica gel using hexane and ethyl acetate eluents to afford, after removal of the solvents, the pure corresponding &#945;-arylated ketone.</li>
	</ol>
	</li>
</ol>

<ol>
	<li>Mizar, P., Wirth, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flexible+stereoselective+functionalizations+of+ketones+through+umpolung+with+hypervalent+iodine+reagents.">Flexible stereoselective functionalizations of ketones through umpolung with hypervalent iodine reagents.</a> Angewandte Chemie International Edition. 53 (23), 5993-5997 (2014).</li><li>Yoshimura, A., Zhdankin, V. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Advances+in+synthetic+applications+of+hypervalent+iodine+compounds.">Advances in synthetic applications of hypervalent iodine compounds.</a> Chemical Reviews. 116 (5), 3328-3435 (2016).</li><li>Zhdankin, V. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Hypervalent Iodine Chemistry: Preparation, Structure, and Synthetic Applications of Polyvalent Iodine Compounds. , (2013).</li><li>Wirth, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Topics in Current Chemistry. 373, (2016).</li><li>Merritt, E. A., Olofsson, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=&#945;-functionalization+of+carbonyl+compounds+using+hypervalent+iodine+reagents.">&#945;-functionalization of carbonyl compounds using hypervalent iodine reagents.</a> Synthesis. 4 (4), 517-538 (2011).</li><li>Arava, S., 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=Enolonium+Species-Umpoled+Enolates.">Enolonium Species-Umpoled Enolates.</a> Angewandte Chemie International Edition. 56 (10), 2599-2603 (2017).</li><li>Parida, K. N., Maksymenko, S., Pathe, G. K., Szpilman, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cross-Coupling+of+Dissimilar+Ketone+Enolates+via+Enolonium+Species+to+afford+Nonsymmetrical+1,4-Diketones.">Cross-Coupling of Dissimilar Ketone Enolates via Enolonium Species to afford Nonsymmetrical 1,4-Diketones.</a> Beilstein Journal of Organic Chemistry. 14, 992-997 (2018).</li><li>Zhdankin, V. V., 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=Carbon-carbon+bond+formation+in+reactions+of+PhIO&#183;HBF4-silyl+enol+ether+adduct+with+alkenes+or+silyl+enol+ethers.">Carbon-carbon bond formation in reactions of PhIO&#183;HBF4-silyl enol ether adduct with alkenes or silyl enol ethers.</a> Journal of Organic Chemistry. 54 (11), 2605-2608 (1989).</li><li>Maksymenko, S., 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=Transition-metal-free+intermolecular+&#945;-arylation+of+ketones+via+enolonium+species.">Transition-metal-free intermolecular &#945;-arylation of ketones via enolonium species.</a> Organic Letters. 19 (23), 6312-6315 (2017).</li><li>Vita, M. V., Waser, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Azidation+of+&#946;-keto+esters+and+silyl+enol+ethers+with+a+benziodoxole+reagent.">Azidation of &#946;-keto esters and silyl enol ethers with a benziodoxole reagent.</a> Organic Letters. 15 (13), 3246-3249 (2013).</li><li>More, 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=&#945;-N-Heteroarylation+and+&#945;-azidation+of+ketones+via+enolonium+species.">&#945;-N-Heteroarylation and &#945;-azidation of ketones via enolonium species.</a> Journal of Organic Chemistry. 83, 2442-2447 (2018).</li><li>Xie, L., 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=Gold-catalyzed+hydration+of+haloalkynes+to+&#945;-halomethyl+ketones.">Gold-catalyzed hydration of haloalkynes to &#945;-halomethyl ketones.</a> Journal of Organic Chemistry. 78 (18), 9190-9195 (2013).</li><li>Patonay, T., Juh&#225;sz-T&#243;th, &. #. 2. 0. 1. ;., B&#233;nyei, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Base-induced+coupling+of+&#945;-azido+ketones+with+aldehydes+&#8722;+An+easy+and+efficient+route+to+trifunctionalized+synthons+2-azido-3-hydroxy+ketones,+2-acylaziridines,+and+2-acylspiroaziridines.">Base-induced coupling of &#945;-azido ketones with aldehydes &#8722; An easy and efficient route to trifunctionalized synthons 2-azido-3-hydroxy ketones, 2-acylaziridines, and 2-acylspiroaziridines.</a> European Journal of Organic Chemistry. 2002 (2), 285-295 (2002).</li><li>Li, C., Breit, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rhodium-catalyzed+chemo-+and+regioselective+decarboxylative+addition+of+&#946;-ketoacids+to+allenes:+Efficient+construction+of+tertiary+and+quaternary+carbon+Centers.">Rhodium-catalyzed chemo- and regioselective decarboxylative addition of &#946;-ketoacids to allenes: Efficient construction of tertiary and quaternary carbon Centers.</a> Journal of the American Chemical Society. 136 (3), 862-865 (2014).</li></ol>

We have nothing to disclose.

The successful preparation of enolonium species from TMS-enolates is dependent on a number of factors. The major side reaction in the preparation step is the homo coupling of the starting material by reaction of a molecule of formed enolonium species with a molecule of TMS-enolate. Thus, the requirement of the reaction conditions is to avoid this dimerization by ensuring fast reaction of the Lewis acid activated hypervalent iodine reagent with added TMS-enolate relative to the rate of dimerization. This is achieved in th...

Enolates are classical carbon nucleophiles in organic chemistry and among the most widely used. Umpolung of enolates to create electrophilic enolonium species allows valuable alternative ways to produce &#945;-functionalized ketones as well as to enable novel reactions not possible via classical enolate chemistry. Enolonium species have been proposed as intermediates in numerous reactions, in particular reactions involving hypervalent iodine reagents. These reactions include &#945;-halogenation, oxygenation, and amination1 as well as other reactions2,3,4,5.
However, the scopes of these reactions were always limited by the transient nature of the reactive enolonium species. This transiency required any nucleophile to be present in the reaction mixture during the reaction of the carbonyl enolates with the strongly oxidizing hypervalent iodine reagent. Thus, any nucleophile prone to oxidation, such as electron rich aromatic compounds (heterocycles) and alkenes, could not be used.
In the last year, we have overcome these limitations by developing conditions in which the enolonium species is formed as a discrete intermediate in one step followed by addition of the nucleophile in a second step. This protocol allows not only the classical type of functionalization such as chlorination6, but also the use of oxidizable carbon nucleophiles, such as allylsilanes6,8, enolates1,6,7, and electron rich aromatic compounds9, resulting in C-C bond formation. The allylation method is amenable to the formation of quaternary and tertiary centers. The ketone arylation method constitutes formal C-H functionalization of the aromatic compound without the need for a directing group9. Recently, we have reported the addition of azoles and azides10 as well11. The detailed presentation of the protocol is expected to assist in the introduction of these methods into the day to day tool box of the synthetic organic chemist.

<table>
	<tbody>
		<tr>
			<td>Chlorotrimethylsilane, 98+%</td>
			<td>Alfa Aesar</td>
			<td>A13651</td>
			<td>TMS-Cl</td>
		</tr>
		<tr>
			<td>Boron trifluoride diethyl etherate, 98+%</td>
			<td>Alfa Aesar</td>
			<td>A15275</td>
			<td>BF3*Et2O</td>
		</tr>
		<tr>
			<td>2-Methylindole, 98+%</td>
			<td>Alfa Aesar</td>
			<td>A10764</td>
			<td>2-Me-indole</td>
		</tr>
		<tr>
			<td>Hydroxy(tosyloxy) iodobenzene, 97%</td>
			<td>Alfa Aesar</td>
			<td>L15701</td>
			<td>Koser&#39;s reagent</td>
		</tr>
		<tr>
			<td>Acetophenone, &#62;98%</td>
			<td>Merck</td>
			<td>800028</td>
			<td></td>
		</tr>
		<tr>
			<td>n-Butyllithium solution 1.6M in hexanes</td>
			<td>Aldrich</td>
			<td>186171</td>
			<td>nBuLi</td>
		</tr>
		<tr>
			<td>BIS(ISOPROPYL)AMINE</td>
			<td>Apollo</td>
			<td>OR1090</td>
			<td>DIPA</td>
		</tr>
		<tr>
			<td>Trimethylsilyl azide, 94%</td>
			<td>Alfa Aesar</td>
			<td>L00173</td>
			<td>TMS-N3</td>
		</tr>
	</tbody>
</table>

a two-step protocol for umpolung functionalization of ketones via enolonium species

&#945;-Functionalization of ketones via umpolung of enolates by hypervalent iodine reagents is an important concept in synthetic organic chemistry. Recently, we have developed a two-step strategy for ketone enolate umpolung that has enabled the development of methods for chlorination, azidation, and amination using azoles. In addition, we have developed C-C bond&#8211;forming arylation and allylation reactions. At the heart of these methods is the preparation of the intermediate and highly reactive enolonium species prior to addition of a reactive nucleophile. This strategy is thus reminiscent of the preparation and use of metal enolates in classical synthetic chemistry. This strategy allows the use of nucleophiles that would otherwise be incompatible with the strongly oxidizing hypervalent iodine reagents. In this paper we present a detailed protocol for chlorination, azidation, N-heteroarylation, arylation, and allylation. The products include motifs prevalent in medicinally active products. This article will greatly assist others in using these methods.

Representative results, achieved following the protocol, are given in Figure 1 and are discussed in the discussion section. Notably, a very large range of different ketones may be used successfully in the reaction to give the products in good yields as may be seen for the azidation11. The scope of the reaction for introducing azoles in the &#945;-position of ketones includes most of the common mono-cyclic and bicyclic nitrogen containi...

Watch this Scientific Journal Video about A Two-Step Protocol for Umpolung Functionalization of Ketones Via Enolonium Species at JoVE.com

A Two-Step Protocol for Umpolung Functionalization of Ketones Via Enolonium Species

A two step one-pot protocol for the umpolung of ketone enolates to enolonium species and addition of a nucleophile to the &#945;-position is described. Nucleophiles include chloride, azide, azoles, allyl-silanes, and aromatic compounds.

&#945;-Functionalization of ketones via umpolung of enolates by hypervalent iodine reagents is an important concept in synthetic organic chemistry. ...

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9.9K Views.  Ariel University. A two step one-pot protocol for the umpolung of ketone enolates to enolonium species and addition of a nucleophile to the α-position is described. Nucleophiles include chloride, azide, azoles, allyl-silanes, and aromatic compounds.

Video: A Two-Step Protocol for Umpolung Functionalization of Ketones Via Enolonium Species

Synthesis of Antiviral Tetrahydrocarbazole Derivatives by Photochemical and Acid-catalyzed C-H Functionalization via Intermediate Peroxides (CHIPS)

The direct functionalization of C-H bonds is an important and long standing goal in organic chemistry. Such transformations can be very powerful in order to streamline synthesis by saving steps, time and material compared to conventional methods that require the introduction and removal of activating or directing groups. Therefore, the functionalization of C-H bonds is also attractive for green chemistry. Under oxidative conditions, two C-H bonds or one C-H and one heteroatom-H bond can be transformed to C-C and C-heteroatom bonds, respectively. Often these oxidative coupling reactions require synthetic oxidants, expensive catalysts or high temperatures. Here, we describe a two-step procedure to functionalize indole derivatives, more specifically tetrahydrocarbazoles, by C-H amination using only elemental oxygen as oxidant. The reaction uses the principle of C-H functionalization via Intermediate PeroxideS (CHIPS). In the first step, a hydroperoxide is generated oxidatively using visible light, a photosensitizer and elemental oxygen. In the second step, the N-nucleophile, an aniline, is introduced by Br&#248;nsted-acid catalyzed activation of the hydroperoxide leaving group. The products of the first and second step often precipitate and can be conveniently filtered off. The synthesis of a biologically active compound is shown.

Synthesis of Antiviral Tetrahydrocarbazole Derivatives by Photochemical and Acid-catalyzed C-H Functionalization via Intermediate Peroxides CHIPS

A two-step procedure for the synthesis of pharmaceutically active indole-derivatives by C-H functionalization with anilines is described, using photo- and Br&#248;nsted acid catalysis.

The direct functionalization of C-H bonds is an important and long standing goal in organic chemistry. Such transformations can be very powerful in order ...

A Protocol for the Production of Gliadin-cyanoacrylate Nanoparticles for Hydrophilic Coating

This article presents a protocol for the production of protein-based nanoparticles that changes the hydrophobic surface to hydrophilic by a simple spray coating. These nanoparticles are produced by the polymerization reaction of alkyl cyanoacrylate on the surface of cereal protein (gliadin) molecules. Alkyl cyanoacrylate is a monomer that instantly polymerizes at RT when it is applied to the surface of materials. Its polymerization reaction is initiated by the trace amounts of weakly basic or nucleophilic species on the surface, including moisture. Once polymerized, the polymerized alkyl cyanoacrylates show a strong affinity with the object materials because nitrile groups are in the backbone of poly (alkyl cyanoacrylate). Proteins also work as initiator for this polymerization because they contain amine groups that can initiate the polymerization of cyanoacrylate. If aggregated protein is used as an initiator, protein aggregate is surrounded by the hydrophobic poly(alkyl cyanoacrylate) chains after the polymerization reaction of alkyl cyanoacrylate. By controlling the experimental condition, particles in the nanometer range are produced. The produced nanoparticles readily adsorb to the surface of most materials including glass, metals, plastics, wood, leather, and fabrics. When the surface of a material is sprayed with the produced nanoparticle suspension and rinsed with water, the micellar structure of nanoparticle changes its conformation, and the hydrophilic proteins are exposed to the air. As a result, the nanoparticle-coated surface changes to hydrophilic.

This article presents a protocol for the production of protein-based nanoparticles that changes the hydrophobic surface to hydrophilic. The produced nanoparticle is an assembly of gliadin-cyanoacrylate diblock copolymers. Spray coating with the produced nanoparticle changes the surface of target material to a hydrophilic surface.

This article presents a protocol for the production of protein-based nanoparticles that changes the hydrophobic surface to hydrophilic by a simple spray ...

Rapid High-throughput Species Identification of Botanical Material Using Direct Analysis in Real Time High Resolution Mass Spectrometry

We demonstrate that direct analysis in real time-high resolution mass spectrometry can be used to produce mass spectral profiles of botanical material, and that these chemical fingerprints can be used for plant species identification. The mass spectral data can be acquired rapidly and in a high throughput manner without the need for sample extraction, derivatization or pH adjustment steps. The use of this technique bypasses challenges presented by more conventional techniques including lengthy chromatography analysis times and resource intensive methods. The high throughput capabilities of the direct analysis in real time-high resolution mass spectrometry protocol, coupled with multivariate statistical analysis processing of the data, provide not only class characterization of plants, but also yield species and varietal information. Here, the technique is demonstrated with two psychoactive plant products, Mitragyna speciosa (Kratom) and Datura (Jimsonweed), which were subjected to direct analysis in real time-high resolution mass spectrometry followed by statistical analysis processing of the mass spectral data. The application of these tools in tandem enabled the plant materials to be rapidly identified at the level of variety and species.

A method for species identification of botanical material by direct analysis in real time-high resolution mass spectrometry and multivariate statistical analysis is presented.

We demonstrate that direct analysis in real time-high resolution mass spectrometry can be used to produce mass spectral profiles of botanical material, ...

A Protocol for Safe Lithiation Reactions Using Organolithium Reagents

Organolithium reagents are powerful tools in the synthetic chemist&#39;s toolbox. However, the extreme pyrophoric nature of the most reactive reagents warrants proper technique, thorough training, and proper personal protective equipment. To aid in the training of researchers using organolithium reagents, a thorough, step-by-step protocol for the safe and effective use of tert-butyllithium on an inert gas line or within a glovebox is described. As a model reaction, preparation of lithium tert-butyl amide by the reaction of tert-butyl amine with one equivalent of tert-butyl lithium is presented.

The safe and proper use of organolithium reagents is described.

Organolithium reagents are powerful tools in the synthetic chemist&#39;s toolbox. However, the extreme pyrophoric nature of the most reactive reagents ...

Protocol for the Solid-phase Synthesis of Oligomers of RNA Containing a 2'-O-thiophenylmethyl Modification and Characterization via Circular Dichroism

Solid-phase synthesis has been used to obtain canonical and modified polymers of nucleic acids, specifically of DNA or RNA, which has made it a popular methodology for applications in various fields and for different research purposes. The procedure described herein focuses on the synthesis, purification, and characterization of dodecamers of RNA 5&#39;-[CUA CGG AAU CAU]-3&#39; containing zero, one, or two modifications located at the C2&#39;-O-position. The probes are based on 2-thiophenylmethyl groups, incorporated into RNA nucleotides via standard organic synthesis and introduced into the corresponding oligonucleotides via their respective phosphoramidites. This report makes use of phosphoramidite chemistry via the four canonical nucleobases (Uridine (U), Cytosine (C), Guanosine (G), Adenosine (A)), as well as 2-thiophenylmethyl functionalized nucleotides modified at the 2&#39;-O-position; however, the methodology is amenable for a large variety of modifications that have been developed over the years. The oligonucleotides were synthesized on a controlled-pore glass (CPG) support followed by cleavage from the resin and deprotection under standard conditions, i.e., a mixture of ammonia and methylamine (AMA) followed by hydrogen fluoride/triethylamine/N-methylpyrrolidinone. The corresponding oligonucleotides were purified via polyacrylamide electrophoresis (20% denaturing) followed by elution, desalting, and isolation via reversed-phase chromatography (Sep-pak, C18-column). Quantification and structural parameters were assessed via ultraviolet-visible (UV-vis) and circular dichroism (CD) photometric analysis, respectively. This report aims to serve as a resource and guide for beginner and expert researchers interested in embarking in this field. It is expected to serve as a work-in-progress as new technologies and methodologies are developed. The description of the methodologies and techniques within this document correspond to a DNA/RNA synthesizer (refurbished and purchased in 2013) that uses phosphoramidite chemistry.

Protocol for the Solid-phase Synthesis of Oligomers of RNA Containing a 2'-O-thiophenylmethyl Modification and Characterization via Circular Dichroism

This article provides a detailed procedure on the solid-phase synthesis, purification, and characterization of dodecamers of RNA modified at the C2&#39;-O-position. UV-vis and circular dichroism photometric analyses are used to quantify and characterize structural aspects, i.e., single-strands or double-strands.

Solid-phase synthesis has been used to obtain canonical and modified polymers of nucleic acids, specifically of DNA or RNA, which has made it a popular ...

Fabrication and Testing of Catalytic Aerogels Prepared Via Rapid Supercritical Extraction

Protocols for preparing and testing catalytic aerogels by incorporating metal species into silica and alumina aerogel platforms are presented. Three preparation methods are described: (a) the incorporation of metal salts into silica or alumina wet gels using an impregnation method; (b) the incorporation of metal salts into alumina wet gels using a co-precursor method; and (c) the addition of metal nanoparticles directly into a silica aerogel precursor mixture. The methods utilize a hydraulic hot press, which allows for rapid (&#60;6 h) supercritical extraction and results in aerogels of low density (0.10 g/mL) and high surface area (200-800 m2/g). While the work presented here focuses on the use of copper salts and copper nanoparticles, the approach can be implemented using other metal salts and nanoparticles. A protocol for testing the three-way catalytic ability of these aerogels for automotive pollution mitigation is also presented. This technique uses custom-built equipment, the Union Catalytic Testbed (UCAT), in which a simulated exhaust mixture is passed over an aerogel sample at a controlled temperature and flow rate. The system is capable of measuring the ability of the catalytic aerogels, under both oxidizing and reducing conditions, to convert CO, NO and unburned hydrocarbons (HCs) to less harmful species (CO2, H2O and N2). Example catalytic results are presented for the aerogels described.

Here we present protocols for preparing and testing catalytic aerogels by incorporating metal species into silica and alumina aerogel platforms. Methods for preparing materials using copper salts and copper-containing nanoparticles are featured. Catalytic testing protocols demonstrate the effectiveness of these aerogels for three-way catalysis applications.

Protocols for preparing and testing catalytic aerogels by incorporating metal species into silica and alumina aerogel platforms are presented. Three ...

The MPLEx Protocol for Multi-omic Analyses of Soil Samples

Mass spectrometry (MS)-based integrated metaproteomic, metabolomic, and lipidomic (multi-omic) studies are transforming our ability to understand and characterize microbial communities in environmental and biological systems. These measurements are even enabling enhanced analyses of complex soil microbial communities, which are the most complex microbial systems known to date. Multi-omic analyses, however, do have sample preparation challenges, since separate extractions are typically needed for each omic study, thereby greatly amplifying the preparation time and amount of sample required. To address this limitation, a 3-in-1 method for the simultaneous extraction of metabolites, proteins, and lipids (MPLEx) from the same soil sample was created by adapting a solvent-based approach. This MPLEx protocol has proven to be both simple and robust for many sample types, even when utilized for limited quantities of complex soil samples. The MPLEx method also greatly enabled the rapid multi-omic measurements needed to gain a better understanding of the members of each microbial community, while evaluating the changes taking place upon biological and environmental perturbations.

A protocol is presented for simultaneously extracting metabolites, proteins, and lipids from a single soil sample, allowing reduced sample preparation times and enabling multi-omic mass spectrometry analyses of samples with limited quantities.

Mass spectrometry (MS)-based integrated metaproteomic, metabolomic, and lipidomic (multi-omic) studies are transforming our ability to understand and ...

Separation of Aldehydes and Reactive Ketones from Mixtures Using a Bisulfite Extraction Protocol

The purification of organic compounds is an essential component of routine synthetic operations. The ability to remove contaminants into an aqueous layer by generating a charged structure provides an opportunity to use extraction as a simple purification technique. By combining the use of a miscible organic solvent with saturated sodium bisulfite, aldehydes and reactive ketones can be successfully transformed into charged bisulfite adducts that can then be separated from other organic components of a mixture by the introduction of an immiscible organic layer. Here, we describe a simple protocol for the removal of aldehydes, including sterically-hindered neopentyl aldehydes and some ketones, from chemical mixtures. Ketones can be separated if they are sterically unhindered cyclic or methyl ketones. For aliphatic aldehydes and ketones, dimethylformamide is used as the miscible solvent to improve removal rates. The bisulfite addition reaction can be reversed by basification of the aqueous layer, allowing for the re-isolation of the reactive carbonyl component of a mixture.

Here, we present a protocol to remove aldehydes and reactive ketones from mixtures by a liquid-liquid extraction protocol directly with saturated sodium bisulfite in a miscible solvent. This combined protocol is rapid and facile to perform. The aldehyde or ketone can be re-isolated by the basification of the aqueous layer.

The purification of organic compounds is an essential component of routine synthetic operations. The ability to remove contaminants into an aqueous layer ...

Surface Functionalization of Metal-Organic Frameworks for Improved Moisture Resistance

Metal-organic frameworks (MOFs) are a class of porous inorganic materials with promising properties in gas storage and separation, catalysis and sensing. However, the main issue limiting their applicability is their poor stability in humid conditions. The common methods to overcome this problem involve the formation of strong metal-linker bonds by using highly charged metals, which is limited to a number of structures, the introduction of alkylic groups to the framework by post-synthetic modification (PSM) or chemical vapour deposition (CVD) to enhance overall hydrophobicity of the framework. These last two usually provoke a drastic reduction of the porosity of the material. These strategies do not permit to exploit the properties of the MOF already available and it is imperative to find new methods to enhance the stability of MOFs in water while keeping their properties intact. Herein, we report a novel method to enhance the water stability of MOF crystals featuring Cu2(O2C)4&#160;paddle-wheel units, such as HKUST (where HKUST stands for Hong Kong University of Science &#38; Technology), with the catechols functionalized with alkyl and fluoro-alkyl chains. By taking advantage of the unsaturated metal sites and the catalytic catecholase-like activity of CuII ions, we are able to create robust hydrophobic coatings through the oxidation and subsequent polymerization of the catechol units on the surface of the crystals under anaerobic and water-free conditions without disrupting the underlying structure of the framework. This approach not only affords the material with improved water stability but also provides control over the function of the protective coating, which enables the development of functional coatings for the adsorption and separations of volatile organic compounds. We are confident that this approach could also be extended to other unstable MOFs featuring open metal sites.

Robust functional catechol coatings were produced in one step by direct reaction of the material known as HKUST with synthetic catechols under anaerobic conditions. The formation of homogeneous coatings surrounding the entire crystal is ascribed to the biomimetic catalytic activity of Cu(II) dimers on the external surface of the crystals.

Metal-organic frameworks (MOFs) are a class of porous inorganic materials with promising properties in gas storage and separation, catalysis and sensing. ...

Automation of a Positron-emission Tomography (PET) Radiotracer Synthesis Protocol for Clinical Production

The development of new positron-emission tomography (PET) tracers is enabling researchers and clinicians to image an increasingly wide array of biological targets and processes. However, the increasing number of different tracers creates challenges for their production at radiopharmacies. While historically it has been practical to dedicate a custom-configured radiosynthesizer and hot cell for the repeated production of each individual tracer, it is becoming necessary to change this workflow. Recent commercial radiosynthesizers based on disposable cassettes/kits for each tracer simplify the production of multiple tracers with one set of equipment by eliminating the need for custom tracer-specific modifications. Furthermore, some of these radiosynthesizers enable the operator to develop and optimize their own synthesis protocols in addition to purchasing commercially-available kits. In this protocol, we describe the general procedure for how the manual synthesis of a new PET tracer can be automated on one of these radiosynthesizers and validated for the production of clinical-grade tracers. As an example, we use the ELIXYS radiosynthesizer, a flexible cassette-based radiochemistry tool that can support both PET tracer development efforts, as well as routine clinical probe manufacturing on the same system, to produce [18F]Clofarabine ([18F]CFA), a PET tracer to measure in vivo deoxycytidine kinase (dCK) enzyme activity. Translating a manual synthesis involves breaking down the synthetic protocol into basic radiochemistry processes that are then translated into intuitive chemistry &#34;unit operations&#34; supported by the synthesizer software. These operations can then rapidly be converted into an automated synthesis program by assembling them using the drag-and-drop interface. After basic testing, the synthesis and purification procedure may require optimization to achieve the desired yield and purity. Once the desired performance is achieved, a validation of the synthesis is carried out to determine its suitability for the production of the radiotracer for clinical use.

Automation of a Positron-emission Tomography PET Radiotracer Synthesis Protocol for Clinical Production

Positron-emission tomography (PET) imaging sites that are involved in multiple early clinical research trials need robust and versatile radiotracer manufacturing capabilities. Using the radiotracer [18F]Clofarabine as an example, we illustrate how to automate the synthesis of a radiotracer using a flexible, cassette-based radiosynthesizer and validate the synthesis for clinical use.

The development of new positron-emission tomography (PET) tracers is enabling researchers and clinicians to image an increasingly wide array of biological ...