Name: cercosporin-photocatalyzed [4+1]- and [4+2]-annulations of azoalkenes under mild conditions
Uploaded: 2020-07-17T13:00:00
Description: Watch this Scientific Journal Video about Cercosporin-Photocatalyzed [4+1]- and [4+2]-Annulations of Azoalkenes Under Mild Conditions at JoVE.com

We thank for the National Key R&#38;D Program of China (2018YFA0901700), Natural Science Foundation of Jiangsu Province (Grants No. BK20160167), the Thousand Talents Plan (Young Professionals), the Fundamental Research Funds for the Central Universities (JUSRP51712B), the National First-class Discipline Program of Light Industry Technology and Engineering (LITE2018-14) and Postdoctoral Foundation in Jiangsu Province (2018K153C) for the funding support.

NOTE: &#945;-Halo-N-acyl-hydrazones were prepared according to a published procedure60. All the solvents and other chemical reagents were obtained from commercial sources without further purification. We first described the synthesis of &#945;-Halo-N-acyl-hydrazones and the biosynthesis of cercosporin as a metal-free photocatalyst. Next, we illustrated the protocols of the cercosporin-photocatalyzed reactions for the synthesis of 1,2,3-thiadiazoles and 1,4,5,6-t...

Nitrogen-containing heterocycles are important motifs for many new drugs and were traditionally synthesized through thermal cycloaddition reactions. Due to great interest, a new photocatalytic method for the synthesis of these compounds is highly desired. To take advantage of the excellent photosensitization properties of cercosporin, we applied cercosporin as a metal-free photocatalyst in two categories of annulation reactions to synthesize nitrogen-containing heterocycles.
First, we reported...

Nitrogen-containing heterocycles have drawn much attention since they are not only important skeletons for a wide range of natural products with bioactivities, but also the synthetic precursors for agrochemicals and drug molecules1,2. Among the various N-heterocycles, 1,2,3-thiadiazoles3,4 and 1,4,5,6-tetrahydropyridazines5,6 are the most important molecules, which are utilized as versatile intermediates in the synthetic chemistry (Figure 1). Since modification of their functional groups always induces distinctive pharmacological activities, extensive efforts have been devoted to developing effective strategies for the synthesis of nitrogen-containing heterocycles and they were mostly synthesized through thermal cycloaddition reactions7,8,9,10. Nowadays, to meet the requirements of sustainable development and green chemistry, photocatalysis has exerted great importance and advantages11,12,13,14, which includes effectiveness15,16,17,18,19 and avoidance of stoichiometric reagents for the activation20,21. The powerful and versatile four-unit intermediates, azoalkenes (1,2-diaza-1,3-dienes)22,23,24,25,26,27,28,29, have been employed as precursors in metal-based Ru(bpy)3Cl2-photocatalyzed reactions with high efficiency for the annulation of halogeno hydrazine and ketocarbonyls30. Furthermore, it was also used in the metal-free Eosin Y photocatalyzed system, but affording the desired product in only 7% yield. Since metal-free photocatalysts show great advantage over transition metal-based photocatalysts, regarding to the environmental factor as well as the cheaper prices18,19, it is highly important to develop new metal-free photocatalytic systems for the synthesis of N-heterocycles.
Cercosporin31,32,33,34,35, hypocrellin36,37,38,39,40, elsinochrome41 and phleichrome42,43 (Figure 2) belong to perylenequinonoid pigments (PQPs) in nature and are produced by endophytic fungi, which have been widely investigated regarding to their photophysical and photobiological properties, and applied in photodynamic therapy and photophysical diagnosis, due to their strong absorption in UV-vis region and unique properties of photosensitization36,44,45,46,47. Upon irradiation, those PQPs can be prompted to excited state and then generate active species through energy transfer (EnT) and electron transfer (ET)35,38,44,48,49,50,51,52,53,54. Thus, we envisioned that these natural PQPs may be utilized as &#34;metal-free&#34; photocatalysts to drive organic reactions, which have rarely been investigated55,56,57,58,59.
Herein, we report the protocol for the biosynthesis of cercosporin from liquid fermentation and then apply it as a metal-free photocatalyst for the [4+1] annulation reaction of azoalkenes and KSCN, as well as the [4+2] cyclodimerization of azoalkenes, which supply 1,2,3-thiadiazoles and 1,4,5,6- tetrahydropyridazines with high efficiency under mild conditions, respectively (Figure 3).

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cercosporin-photocatalyzed [4+1]- and [4+2]-annulations of azoalkenes under mild conditions

The interest on nitrogen-containing heterocycles has expanded rapidly in the synthetic community since they are important motifs for new drugs. Traditionally, they were synthesized through thermal cycloaddition reactions, whereas today, photocatalysis is preferred due to the mild and efficient conditions. With this focus, a new photocatalytic method for the synthesis of nitrogen-containing heterocycles is highly desired. Here, we report a protocol for the biosynthesis of cercosporin, which could function as a metal-free photocatalyst. We then illustrate cercosporin-photocatalyzed protocols for the synthesis of nitrogen-containing heterocycles 1,2,3-thiadiazoles through annulation of azoalkenes with KSCN, and synthesis of 1,4,5,6-tetrahydropyridazines [4+2] through cyclodimerization of azoalkenes under mild conditions, respectively. As a result, there is a new bridge between the microbial fermentation method and organic synthesis in a mild, cost-effective, environmentally friendly and sustainable manner.

Synthesis of &#945;-Halo-N-acyl-hydrazones: They are synthesized according to Protocol 1.
Synthesis of cercosporin: It was synthesized and purified according to Protocol 2. 1H NMR (400 MHz, CDCl3): &#948; ppm 14.82 (s, 2H, ArH), 7.06 (s, 2H, ArH), 5.57 (s, 2H, CH2), 4.20 (s, 6H, 2OCH3), 3.62-3.57 (m, 2H, CH2), 3.42-3.37 (m,...

Watch this Scientific Journal Video about Cercosporin-Photocatalyzed [4+1]- and [4+2]-Annulations of Azoalkenes Under Mild Conditions at JoVE.com

Cercosporin-Photocatalyzed [4+1]- and [4+2]-Annulations of Azoalkenes Under Mild Conditions

New routes for the synthesis of nitrogen-containing heterocycles utilizing cercosporin as a metal-free photocatalyst were developed.

The interest on nitrogen-containing heterocycles has expanded rapidly in the synthetic community since they are important motifs for new drugs. ...

This protocol can be used for the biosynthesis of cercosporins through microbial fermentation, which in turn can be used to synthesize nitrogen-containing heterocycles in the green and mild conditions. An advantage of this technique is that it can be used as a bridge between the microbial fermentation methods and the organic synthesized processes. Nitrogen-containing heterocycles are not only important skeleton for many natural product with bioactivities, but they are synthetic precursors for many agrochemicals and drugs.Demonstrating the procedure with Philippe Icyishaka will be Yan Zhang, a technician from my laboratory. For alpha-Halo-N-acyl-hydrazone preparation, weigh out 10 millimolar of ketone and 10 millimolar of benzoyl hydrazine into a flask. Add 20 milliliters of methanol to the flask, and equip the flask with a stir bar and a rubber stopper.Slowly inject 250 microliters of hydrochloric acid into the mixture, and incubate the flask in air at room temperature. After one hour, collect the precipitate after reaction by filtration, and wash the product with acetone. Then dry the precipitate by vacuum, and identify the product by NMR.To prepare cercosporin, charge a three-liter shake flask with one liter of S-7 medium, and inoculate the cercosporin-producing strain into the shake flask. Culture the mixture under light conditions at 135 revolutions per minute and 25 degrees Celsius for two weeks. At the end of the incubation, use a vacuum pump to subject the fermentation broth to vacuum filtration, and extract the pellet and the supernatant separately with three 50-milliliter volumes of dichloromethane.Collect the pellet for drying in a freezer dryer, and combine the organic phases. Wash the organic phases with water two to three times, and concentrate the solution under vacuum. Redissolve the residue with analytical methanol, and filter the product through a 18-micrometer, organic microfiltration membrane.Then purify the cercosporin with a Sephadex LH-20 column, and identify the product by HPLC according to standard protocols. To prepare 1, 2, 3-thiadiazole, add the entire volume of the alpha-Halo-N-acyl-hydrazone, one milligram of cercosporin, 27 milligrams of potassium tert-butoxide, and 39 milligrams of potassium thiocyanate into a 10-millimeter Schlenk tube equipped with a rubber stopper and a stirring bar. Purge the Schlenk tube with oxygen three times before injecting two milliliters of dry acetonitrile into the tube.Subject the tube to a five-watt, blue LED from the bottom for 16 hours before washing four times with 15 milliliters of saturated sodium chloride solution. After the last wash, combine the aqueous phase, and re-extract the aqueous phase with four 15-milliliter volumes of ethyl acetate. Combine the organic phase, and dry with anhydrous sodium sulfate.Remove the solvent with a vacuum evaporator. To purify the product with thin-layer chromatography, draw a straight line on the silica plate, and add the dissolved product to the line. Put the silica plate in the chromatography cylinder with 10 to one PE to EA.After, dissolve the silica in DCM.Remove the silica from the product by filtration, and dry the DCM with a rotary evaporator. Then identify the compound by NMR according to standard protocol. To prepare 1, 4, 5, 6-tetrahydropyridazine, add the entire volume of the alpha-Halo-N-acyl-hydrazone, 2.7 milligrams of cercosporin, and 195 milligrams of cesium carbonate into a 10-milliliter Schlenk tube equipped with a rubber stopper and a stirring bar.Purge the Schlenk tube three times with nitrogen, and inject two milliliters of acetonitrile in water into the tube. Subject the Schlenk tube to a five-watt, blue LED from the bottom for 16 hours followed by four washes with 15 milliliters of saturated sodium chloride per wash. Combine the aqueous phase, and re-extract the aqueous phase with four 15-milliliter volumes of ethyl acetate.Combine the organic phase, and dry with anhydrous sodium sulfate. Remove the solvent with a vacuum evaporator. To purify the product with thin-layer chromatography, draw a straight line on the silica plate, and add the dissolved product to the line.Put the silica plate in the chromatography cylinder with 10 to one PE to EA.After, dissolve the silica in DCM. Remove the silica from the product with filtration, and dry the DCM with a rotary evaporator. Then identify the compound by NMR according to standard protocol.These representative results demonstrate how 4-aryl-1, 2, 3-thiadiazoles and 1, 4, 5, 6-tetrahydropyridazines can be conveniently synthesized by cercosporin-catalyzed photocatalytic reactions from alpha-Halo-N-acyl-hydrazone. The 4-aryl-1, 2, 3-thiadiazoles acquisition procedure is suitable for substrates bearing both electron-donating and electron-accepting groups in the phenyl ring, providing the desired products with moderate to good yields. Always remember to use nitrogen and oxygen for two different categories of the reactions.

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