Cascade N-Alkylation/Hemiacetalization for Facile Construction of the Spiroketal Skeleton of Acortatarin Alkaloids with Therapeutic Potentiality in Diabetic Nephropathy

The concise building of the spiroketal core of acortatarin-type alkaloids as potential therapeutic agents in diabetic nephropathy was established in four steps, through a tandem N-alkylation/hemiacetalization between pyrrole units and the corresponding halo alcohols generated by convenient halomethylation of chiral lactones from natural aldoses. The Acortatarin alkaloids exhibit a promising perspective against diabetic nephropathy and other ROS-linked diseases. Facile construction of the dioxaspirocycle motif was realized through tandem N-alkylation/hemiacetalization between pyrrole units and the corresponding halo alcohols, which were derived in turn from convenient halomethylation of chiral lactones in natural carbohydrates pool.

however starting from expensive D-thymidine, using stoichiometric amounts of toxic mercury salt in the key step for acortatarin A. Recently, Kuwahara and coworkers [8] reported their expeditious route for acortatarin A derived from elaborately protected chiral olefin, nevertheless, associated with higher cost and harsher deprotection/ cyclization steps. This particular architecture which appears simple comprises a significant and attractive synthetic challenge indeed. Herein, we report our efforts towards facile building of the spiroketal core by extending the application of halo alcohols, which were generated by convenient halomethylation of chiral lactones from natural aldoses. An interesting tandem N-alkylation/hemiacetalization with pyrrole units was established to provide an alternative route for the target compounds.

Results and Discussion
The synthetic entries mentioned above are mainly based on S N 2 reaction between the cyclic or acyclic ketose-type derivatives (epoxides [4], bromides [6,8] or iodides [7]) and the pyrrole fragments, only with one example employing Maillard-type condensation [5]. Based upon the proposed biogenesis of acortatarins [2], we envisaged the chemoselective Amadori rearrangement to get the furanoid amino sugars, but our attempts did not give acceptable preparative yields (see supporting information). The work of Morin [9] and Wadouachi [10] made us recognize that the iodomethyl group acts as an interchanging hydroxymethyl or aminomethyl synthetic equivalent (Scheme 1). At the meanwhile, we noticed that the regioselective protection of fructofuranosyl acceptors directly from D-fructose needs time-consuming long process [11]. Araújo [12] described an efficient approach for the furanoside originating from arabinose, taking advantage of the fixed stereochemistry of the arabinolactone, which was easily prepared by the oxidation of the anomeric position with bromine in water. The one-carbon elongation of aldoses to ketoses in these cases used lithium reagent for the key homologation step. Enlightened by Araújo's successful access to furanoside [12], an alternative retrospective route was outlined to obtain the key intermediates 3a and 3b from the readily available 2-deoxy-D-ribose (4a) [13] and D-arabinose (4b) [14], respectively. Similar to the protocol of Tan [7], the subsequently direct S N 2 coupling with pyrrole partner 5 [10] could bridge the two subunits (Scheme 2). We reasoned that this plan might offer additional strategic advantages in that all the requisite stereochemistry of the hydroxyl groups for 1 and 2 would be attained or kept along with the initial natural aldoses from the beginning to the end. The versatility of assembling other unnatural acortatarin-type alkaloids was to succeed in conjunction with the implementation of one-step haloalkylation on natural chirons.
With the goal of devising a concise synthesis of this uncommon core, the exploration was initiated with the preparation of lactones 7a and 7b by the oxidation of 4a and 4b buffered in an aqueous K 2 CO 3 solution (Scheme 3) [14], then subjected to one-pot protection without further purification to get TBDMS ethers 8a and 8b in approximately 75 % overall yield for both (Scheme 3) [15]. Previously employed iodomethylation [9] for the conversion of 8a to 9 was proved to be unfavorable in this case, because it was very difficult to separate the iodide 9 from starting material recovered by chromatography along with liable decomposition in chloroform (or CDCl 3 ). The focus shifted to the more stable bromomethylating compounds prepared by BrCH 2 Li (in situ generation from CH 2 Br 2 and n-BuLi in the presence of LiBr) [16]. Unfortunately, an uncharacterized side product was observed after purification with slight decomposition by treatment of 8a with these combinational reagents. We were gratified to really furnish 10a as inseparable anomeric isomers (about 1:1 ratio) after venturously removing the additive LiBr. The yield (31 %) was modest, and the major byproduct, dihydrofuranone 10a 0 [17] was delivered in 36 % yield. All attempts to enhance the ratio by changing the solvent (THF, toluene, DME), the amount of BrCH 2 Li (1, 1.5, 2 equivalents), reaction temperature (-65 or -78°C), running-time (0.5, 1, 2 h), or work-up in acidic medium (quenching by saturated ammonium chloride solution or equivalent acetic acid) failed (Scheme 3).
Paal-Knorr condensation of diketones 19 with NH 4 OAc were carried out under a variety of buffered or acidic conditions (HOAc-NaOAc, p-TsOH, CSA, I 2 ). Only polymeric products or decomposition resulted, probably because the acid-catalyzed decomposition of the pyrrolic products is competitive with their formation. Ultimately, after screening dioxane, DME, THF, MeCN, EtOH, MeOH and H 2 O as solvents, one mild neutral buffering system worked only in EtOH (NH 4 OAc/NaOAc = 3.5/1.2 equivalents) to afford pyrrole unit 20 quantitatively. The direct coupling between iodide 14b and 20 proved ineffective. Moreover, the unpredictability of oxidative cleavage of olefins to aldehydes and the additive protection/deprotection for hydroxyl group in 20 put us in a dilemma. At this juncture, the reductive deoxygenation of lactol to benzopyran achieved by exposure to NaBH 4 [23] or triethylsilane (Et 3 SiH) [24] in the presence of TFA attracted our attention. Thus we could continue and complete the expected total synthesis of acortatarin A as illustrated in Scheme 6.
N-alkylation of pyrrole 11 with iodide 14b could deliver hemiacetal 12b (Cs 2 CO 3 /MeCN [6], nearly 1/1 diastereoisomers, 60 % yield without further optimization). The reduction of lactol 12b to create morpholine motif 21 would not be problematic though the free aldehyde could not be kept. The oxidation to 22 and deprotection of silyl ether for acortatarin A and its epimer could be finished according to the known literature methods. To minimize the synthesis steps and maximize the overall efficiency of our strategy, one alternative route was proposed to introduce the second aldehyde group by formylation of 24 at a later stage as depicted.

Conclusion
We hypothesized that the thermodynamic stability of the final products as the driving force in the cascade N-alkylation and concomitant semiacetalization. The spontaneous spiro-hemiacetalization was induced by the deprotonation of hydroxyl group adhering to the anomeric carbon at the ring junction position, followed by sequential attack of the resulting oxygen anion aiming at neighboring aldehyde group. This crucial semiketal carbon serves as one incorporating ''middle hinge'' necessary for the simultaneous shutting of the crab-claw-shaped morpholine ring (Scheme 6). It may supply one alternative route for the troublesome coupling of pyrrole units bearing sterically hindered THP or TBDMS protected hydroxyl groups [4]. For the importance of deoxyribonucleotide chemistry, 2-deoxy-D-ribose derivatives were found to be more advantageous. But the stepwise improved halomethylating condition is adaptable within wider scope. The present experiences were envisioned to allow the assembly of the target molecules efficiently, including the construction of all the subunits in the near future. The key elements in this synthesis sequence include the optimized halomethylation for lactones and tandem nucleophilic substitution. The other important features lie in prompting the utility of natural sugars as the chiral pool inspired by the biogenetic hypothesis. Obviously, when combined with the unnatural formylated pyrrole analogues with para-substitutions (C-2/5) (Scheme 5), such a synthetic strategy is also readily performed for other novel acortatarins. The pursuit towards these goals is currently underway, and the results will be reported in due course.

Experiment Section
Melting points were measured on an SGW X-4 melting instrument and uncorrected. MS were recorded on a Waters AutoSpec Premier P776 or API STAR Pulsar instrument. NMR spectra were recorded on a Bruker AM-400 or DRX-500 spectrometer and calibrated using residual solvent peaks as internal reference, for example CDCl 3 solutions. TLC analyses were performed on commercial glass plates bearing 0.25 mm layer of Merck Silica gel 60F 254 . Silica gel (200-300 mesh, Qingdao Marine Chemical Co. Ltd., People's Republic of China) was used for general column chromatography. Spots on TLC plates were detected under UV radiation and by heating after spraying with 10 % H 2 SO 4 in EtOH or dilute solutions of KMnO 4 . Microwave irradiation reactions were carried out in a CEM Discover SP system. Tetrahydrofuran (THF), diethyl ether and toluene were distilled over sodium/benzophenone. Other reagents were obtained from commercial sources and used without further purification unless otherwise stated.

2-Deoxy
To a solution of 2-deoxy-D-ribose 4a (1.0 g, 7.5 mmol) in 6 mL water, K 2 CO 3 (1. by the addition of 88 % formic acid, the solvent from the aqueous fraction was evaporated to give a pale brown syrup 7a, which was heated at 50°C under reduced pressure for 1 h. EtOH (20 mL) was added, and the solvent was repeatedly evaporated after each addition of EtOH. Without further purification, the crude product 7a was dissolved in 20 mL of anhydrous DMF, to which imidazole (2.5 g, 37.3 mmol) and tert-butyldimethylsilyl chloride (4.5 g, 29.8 mmol) were added. The resulting solution was stirred at rt for 24 h and quenched by addition of water. The water layer was extracted with ethyl acetate (30 mL 9 3), and the organic layers were combined, washed with brine, and dried over anhydrous Na 2 SO 4 . The crude product was concentrated in vacuo. Flash chromatography afforded 8a as a white solid: mp 72-73°C. Yield: 2.03 g (75 % over two steps) [14,15]. 1

1H-Pyrrole-2,5-dicarbaldehyde (11)
A solution of pyrrole-2-aldehyde 6 (0.48 g, 5 mmol), ethyl cyanoacetate (0.68 g, 6 mmol) and piperidine (0.05 mL) in ethanol (5 mL) was stirred at rt for 1 h then heated to reflux for 2 h under the protection of nitrogen. The reactants were cooled to rt, the orange solids precipitated were filtrated and washed by petroleum ether, after removal of the volatiles in vacuo, 15 was obtained in quantitative yield. 3 mL DMF was cooled below 5°C in a round-bottom flask equipped with a stir bar, under protection of nitrogen, oxalyl chloride (0.76 g, 6 mmol) was added under vigorous stirring, the cooling bath was removed, 30 min later, the mixture was cooled to 0°C again. A solution of compound 15 in 2 mL of DMF was added and the resulting mixture was allowed to stir in an ice bath for 45 min and at rt for 2 h. The flask was then quickly fitted with a short condenser pipe protected by calcium chloride drying tube and heated by microwaves at 100°C for 12 min in a CEMmicrowave reactor. After cooling to rt, a solution of saturated aqueous NaHCO 3 was added carefully to adjust the pH to slightly basic, and the mixture was heated for 15-30 min under reflux. After cooling, the crude precipitate 16 was filtered off and added to a solution of 15 mL 3 M NaOH, the mixture was refluxed for 2 h under nitrogen, cooled, neutralized with 6 M hydrochloride acid in an ice bath, the precipitate resulted was filtered. The aqueous phase was extracted by ethyl acetate (20 mL 9 3), the total organic level was dried over anhydrous Na 2 SO 4 , after removal of the solvents in vacuo, the solids filtrated previously and the organic phase were combined and subjected to flash column chromatography on silica gel to afford 11 as pale yellow solid: mp 120-122°C. Yield: 0.32 g (52 % over three steps) [18,19]. 1  in dry THF (2 mL) was cautiously added. After 4 h at rt, the mixture was quenched with a saturated solution of NH 4 Cl, extracted with CH 2 Cl 2 (4 mL 9 2). The combined organic phase was washed with brine, dried over Na 2 SO 4 , filtered and evaporated to dryness. After purification by flash chromatography on silica gel, the lactol 12a (nearly 1/1 diastereoisomers) was provided as pale yellow oil [10].  The S N 2 coupling between 11 and 14b was similar to the above formation of 12a, however, using Cs 2 CO 3 as mild base [6]. Assigned by comparison with compound 12a, part of assignments were in some cases interchangeable. The modified procedure for bromomethylation was as same as that of 10a [9,16], 14a (nearly 1/1 diastereoisomers) was afforded as colorless oil. Yield: 32 mg (36 % Assigned by comparison with compound 10a, part of assignments were in some cases interchangeable.  13  Assigned by comparison with compound 10a, part of assignments were in some cases interchangeable. (18) To a solution of cinnamic acid (1.48 g, 10 mmol) in CH 2 Cl 2 (4 mL) cooled to 0°C, was added triethylamine (2.02 g, 20 mmol). After stirring 10 min at rt, chloroacetonitrile (1.13 g, 15 mmol) was added and the mixture was stirred overnight. The reaction mixture was quenched with ice-H 2 O mixture (3 mL) and the aqueous phase was extracted with CH 2 Cl 2 (4 mL 9 2). Combined organic layers were dried over anhydrous Na 2 SO 4 and filtered. After evaporation of the solvents, the residue was purified