A Modular Strategy for the Synthesis of Dothideopyrones E and F, Secondary Metabolites from an Endolichenic Fungus

Endolichenic fungi are a rich source of natural products with a wide range of potent bioactivities. Herein, syntheses of the two naturally occurring α-pyrones dothideopyrone E and F are presented. These natural products were isolated from a culture of the endolichenic fungus Dothideomycetes sp. EL003334. The outlined strategy includes a Fu–Suzuki akyl–alkyl cross-coupling, a MacMillan α-oxyamination, and a Sato’s pericyclic cascade process to construct the 4-hydroxy-2-pyrone ring system. All the obtained data on the synthesized compounds matched with that of the isolated material.

N atural products from endolichenic fungi have attracted attention due to their bioactivity and new structural motifs. Several structural classes including alkaloids, steroids, peptides, and pyrones are represented and have found applications in agrochemical and pharmaceutical industries. 1 Dothideopyrones E (1) and F (2) are recent examples of natural products isolated from cultures of the endolichenic fungus Dothideomycetes sp. EL003334, obtained from the lichen Stereocaulon tomentosum. 2 Together with dothideopyrones A (3), B (4), C (5), and D (6), dothideopyrones E (1) and F (2) belong to a small class of naturally occurring αpyrones. 3 The structures of these compounds are characterized by a 4-methoxy-2-pyrone core substituted with a hydroxymethyl group at C-3 and an aliphatic side chain with one or two secondary alcohols. Natural α-pyrones present a range of antifungal, cytotoxic, neurotoxic, and phytotoxic properties. 4 Additionally, several naturally occurring α-pyrones have been investigated for treatment of high cholesterol and Alzheimer's disease. 5 Among the dothideopyrones, compound 6 displayed cytotoxic activity on cancer cell lines, while dothideopyrone F (2) inhibited nitric oxide (NO) and prostaglandin E 2 (PGE 2 ) production in lipopolysaccharide (LPS)-induced BV2 microglial cells. 2 Additionally, compound 2 demonstrated the ability to decrease the transcript levels of IL-1β, IL-6, and TNF-α in a dose-dependent manner on BV2 cells stimulated with LPS. Activated microglia cells produce neuroinflammatory factors, including NO, PGE 2 , and TNF-α, as a response to danger in the central nervous system (CNS). 6 However, uncontrolled neuroinflammatory factors contribute to neurodegeneration, leading to changes in the CNS and contribute to diseases such as Alzheimer's and Parkinson's disease. 7 Hence, the control of neuroinflammation is a suitable pharmacologic target for neurodegenerative disease. 8 In this context, the dothideopyrones, and especially dothideopyrone F (2), have been highlighted as a promising therapeutic lead agent to prevent neurodegenerative diseases. Owing to our interest in naturally occurring compounds, especially related to anti-inflammatory properties, this class of α-pyrones attracted our attention. Herein we present our synthetic effort to synthesize dothideopyrones E (1) and F (2).

■ RESULTS AND DISCUSSION
The retrosynthetic analysis applied to the structure of dothideopyrone E (1) is outlined in Scheme 1. The 4hydroxy-2-pyrone ring system was planned to be constructed using the pericyclic cascade approach developed by Sato,9 allowing, thereafter, the attachment of the needed substitutions onto the α-pyrone system. For the introduction of the secondary alcohol adjacent to the pyrone ring, the enantioselective, organocatalytic α-oxyamination developed by the MacMillan group was deemed ideal. 10 The Fu−Suzuki cross-coupling 11,12 was chosen to forge the indicated 4′−5′ carbon−carbon bond, with readily available (R)-(−)-4-penten-2-ol (9) planned transformed into the organoborane component to be reacted with 1-bromo-5-chloropentane (8).
The synthesis commenced with hydroboration of TBS (tertbutyldimethylsilyl eter)-protected 10, 13 which was thereafter coupled with alkyl bromide 8. Initially, the general procedure developed by the Fu group was applied. 11,12 This entailed the use of 1.2 equiv of the organoboron compound and 4 mol % Pd(OAc) 2 , 8 mol % PCy 3 , and K 3 PO 4 ·H 2 O in tetrahydrofuran (THF). 11 While these standard conditions worked well, the homocoupled biproduct formed as a result of the initial Pd 2+ → Pd 0 reduction by the organoborane derived from 10 proved difficult to remove from the desired product 11, thereby complicating the purification process. Additionally, it was also deemed desirable to avoid using a 20% excess of the comparably more valuable enantioenriched fragment in this specific case.
One established alternative is the application of Pd(PCy 3 ) 2 , which has been found to be comparable in effectiveness to the above-mentioned catalytic system, and this approach has previously been applied in the context of synthesis of bioactive natural products. 14 The Pd(PCy 3 ) 2 catalyst is air-sensitive and quite labile, however, and hence typically requires handling in a glovebox for optimal results. After some experimentation, we settled on using 4 mol % of the easily handled Buchwald fourth-generation 15,16 PCy 3 -Pd-G4 as well as 4 mol % of HPCy 3 ·BF 4 , 17 with the reasoning that this should furnish Pd(PCy 3 ) 2 in situ under the basic conditions and, importantly, without the formation of the undesired byproduct given the activation mechanism for the palladacycle precatalyst. This approach led to the smooth union of the two fragments and furnished the coupled product 11 in 77% yield. Next, using the Kornblum oxidation, 18 the alkyl chloride functionality was transformed into the corresponding aldehyde 12 in 69% yield by simply heating 11 to 115−120°C in DMSO together with NaHCO 3 and NaI.
The resulting aldehyde 12 was then subjected to the MacMillan α-oxyamination conditions using 10 mol % Dproline and nitrosobenzene in CHCl 3 . 19 In the telescoped sequence, 20 this was followed by two reduction steps employing NaBH 4 and Zn/AcOH before purification using column chromatography, giving the 1,2-diol 13 in 75% yield and 56:1 d.r. (HPLC analysis).
It should also be pointed out that given the formation of the dioxazinaneol intermediate depicted in Scheme 2, two equivalents of the aldehyde are effectively consumed in this reaction. Consequently, an excess of aldehyde is generally used in order to achieve full consumption of nitrosobenzene: typically three equivalents or more when the aldehyde is readily available and affordable. In this case, however, the number of equivalents was lowered to two.
At this stage, the primary alcohol in the 1,2-diol system in 13 was selectively protected as the sterically hindered pivaloyl ester, and the remaining secondary alcohol was reacted with excess TBS triflate together with a catalytic amount of DMAP (4-dimethylaminopyridine) to yield 14 in 79% yield. 19 The ester moiety was then reductively cleaved with DIBAL-H (diisobutylaluminum hydride) in hexane, giving access to the primary alcohol 15, which was subsequently oxidized to the corresponding aldehyde 16 in 84% yield over two steps. This sensitive intermediate was rapidly taken forward in a Scheme 1. Overview of the Key Retrosynthetic Disconnections Made for Dothideopyrone E (1) vinylogous Mukaiyama aldol reaction (VMAR) with 17 and BF 3 ·OEt 2 as the Lewis acid. 21 To aid in the purification process, the resulting crude aldol product 18 was first oxidized, giving ketone 19 in 72% yield over two steps after column chromatography. Adding 19 dropwise to a boiling solution of toluene set in motion a retro-hetero Diels−Alder reaction, expelling acetone, followed by tautomerization and finally electrocyclization to furnish the 4-hydroxy-2-pyrone intermediate 20 in 76% yield (Scheme 3). 9 The method of Moreno-Manãs was used for the introduction of the sulfide functionality, 22 installed as a temporary substitute for the hydroxymethyl group attached to the 2-pyrone moiety of dothideopyrone E (1). Thus, by subjecting 20 to Knoevenagel conditions, employing paraformaldehyde, acetic acid, and piperidine in EtOH, a highly reactive Michael acceptor intermediate presumably forms, which is subsequently trapped by thiophenol also added to the reaction mixture. 23 This procedure afforded the thioether product in 79% yield. Thereafter, dimethyl sulfate was used to methylate the 4-hydroxy group present in the depicted and dominant tautomer form, giving 21 in 78% yield. The next objective was to convert the thioether into the corresponding primary alcohol, and this was accomplished with the abnormal Pummerer rearrangement, which with this specific system will furnish the alcohol rather than the aldehyde functionality. 24 The sequence was initiated by careful m-CBPA (metachloroperoxybenzoic acid) oxidation to prepare the sulfoxide 23, and, after rapid purification, the obtained material was immediately treated with TFAA (trifluoroacetic anhydride). Finally, aqueous sodium hydroxide was added to hydrolyze any TFA ester formed, giving 24 in 56% yield over two steps.
The ultimate step involved the removal of the two TBS protecting groups. Given the highly hydrophilic nature of 1, a procedure that avoided aqueous workup was desirable. Employing five equivalents of TBAF (tetra-n-butylammonium fluoride) in THF eventually led to full consumption of the starting material 24 (Scheme 4). After addition of acetic acid and removal of the solvent, the crude material was purified using column chromatography with the aim of removing as much of the tetrabutylammonium salts as possible. Fractions containing product were stored at −20°C in order to induce precipitation of the final product. Subsequently, another round of column chromatography then afforded dothideopyrone E (1) in 62% yield and >96% chemical purity (Supporting Information). NMR ( 1 H, 13 C), MS, UV, and optical rotation data were all in accordance with the structure of dothideopyrone E (1). 2 Furthermore, comparison between synthetic and authentic material, using the original NMR data from the isolation and characterization work, showed a clear match (Supporting Information).
During the course of the synthesis of dothideopyrone E (1), no sign of epimerization of the carbinol chiral center adjacent to the pyrone ring system was observed in any of the synthetic intermediates. This observation, coupled with the success of the synthetic approach described above, as well as the interesting biological activity of dothideopyrone F (2), led to the initiation of a campaign toward 2 following essentially the same strategy. The lack of a secondary alcohol in the 8′position, however, meant that readily available and affordable decanal could be used as the starting point (Scheme 5).
The organocatalytic α-oxyamination was again used to introduce the secondary alcohol present in the 1′-position of dothideopyrone F (2) in 83% yield and in >94% ee. The absolute configuration was confirmed by comparison of the optical rotation value of the synthetically prepared (S)-decane-1,2-diol (26) to that of literature values (Supporting Information). From there on, the same sequence of events used to prepare dothideopyrone E (1) was employed to furnish dothideopyrone F (2) in 12 steps and 7% overall yield (Supporting Information). Also in this case, all experimental characterization data were in accordance with the structure of dothideopyrone F (2). 2 Furthermore, comparison between synthetic and authentic material, using the original NMR data from the isolation and characterization work, showed a clear match (Supporting Information).
In summary, the first total syntheses of dothideopyrone E (1) and F (2) have been achieved. The key transformations include a MacMillan enantioselective and organocatalytic αoxyamination, the Fu−Suzuki alkyl−alkyl cross-coupling reaction, and a Sato pericyclic cascade approach to build the pyrone nucleus. Regarding the Fu−Suzuki coupling, it was found that a constellation of catalytic amounts of the palladacycle precatalyst PCy 3 -Pd-G4 and additional ligand precursor HPCy 3 ·BF 4 were both convenient and effective for achieving the desired cross-coupling without the formation of a homocoupled byproduct associated with the reduction of Pd(OAc) 2 and without the use of a glovebox.
The original elucidation work made use of the modified Mosher analysis 25 to establish the absolute configurations of the two carbinol atoms at the 1′ and 8′ positions. In the synthetic preparation of dothideopyrone E (1), commercially available (R)-(−)-4-penten-2-ol (10) served as the origin for the secondary alcohol in the 8′ position, while the MacMillan α-oxyamination, with its well-established and reliable mode of stereoinduction, was used to set the absolute configuration of the carbinol atom in the 1′ position of both dothideopyrones E (1) and F (2).