Concise Synthesis of Both Enantiomers of Pilocarpine

Furan-2-carboxylic acid was used as a starting material for the synthesis of dehydro-homopilopic acid. Esterification, hydrogenation and enzymatic hydrolysis followed by the reduction of Weinreb amides and a single-step attachment of a 1-methyl-imidazole residue allowed for the concise synthesis of both enantiomers of pilocarpine.


Results
In our own synthetic strategy, the introduction of the 1-methylimidazole residue should be the last step of the synthesis, since this residue-as previously shown [42] -can be introduced in a single step and under very mild conditions using a Leusen imidazole synthesis [58][59][60][61]. However, a prerequisite for carrying out this planned synthetic sequence is that (homo)-pilopaldehyde or (homo)-pilopic acid must be present in pure enantiomeric and diastereomeric forms. Furthermore, these intermediates should be accessible in high yields. Since we were interested in the synthesis of both pilocarpine enantiomers, a synthesis to racemic (homo)-pilopic acid with a subsequent separation of the enantiomers seemed to be a reasonable strategy. A good starting material should be readily accessible furan-2-carboxylic acid (2).
Thus, 2 (Scheme 1) was photo-oxidized [62] in the presence of oxygen and the sensitizer bengal rosa via an intermediately formed hydroperoxide to yield 3. Thereby, 4 was formed as the by-product (by acetalisation). Since acetals are easy to introduce on the one hand, but also easy to cleave off on the other, they should be our preferred protecting groups in the next steps. Hereby, the methoxymethylation of the free hydroxyl groups using formaldehyde dimethyl acetal seems particularly suitable, since these reactions are known to proceed mostly with very high yields. In previous experiments, furfural [63][64][65] had been oxidized as an alternative to 2, but polymerization reactions were observed to a high extent during its photo-oxidation.
The reaction of 3 with formaldehyde dimethyl acetal in the presence of solid P 4 O 10 [66] produced an almost quantitative yield of methoxymethylated 5. Interestingly enough, applying the same conditions from 2, ester 6 was formed together with 7 as a minor byproduct. These reaction conditions can therefore be used not only to very efficiently introduce a protecting group on a hydroxyl group but also to protect the carboxylic acids. 2-Ethylmalonic acid (8) was also neatly transformed into its bis(methoxymethylester) 9. The Michael addition of the latter with 5 produced 10 in a 70% isolated yield and 15% of 11 as a by-product; the latter product was formed by transacetalisation. A Stobbe condensation [47] of 10 for 2 days produced dehydrohomopilopic acid (12) together with traces of a side product 13. To elucidate the formation and structure of 13, a mixture of 10 and 11 was heated under reflux with aqueous HBr (48%) for 4 days, and 13 was obtained as colorless crystals. The results from the mass spectrometry showed 13 to hold a bromine substituent, and from the interpretation of 1 H and the 13 C-NMR spectra, 13 was assigned the structure of a 3a-(bromomethyl)-3-ethyl-dihydro-[2,3-b]furan-(3H, 4H)-2,5-dione. For verification of this structure, suitable crystals were grown that were subjected to a single crystal X-ray analysis, whose results are depicted in Table 1 and Figure 2. The formation of 13 remains unclear but should start by a cleavage of the ester moieties of 11 by HBr followed by a decarboxylation of the substituted malonic acid and intramolecular lactonization. Details of the data collection and refinement of the crystal structure of compound 13 are collected in Table 1. Compound 13 ( Figure 2) crystallizes in the monoclinic space group P2 1 /n with four formula units per unit cell. Due to its crystallographic symmetry, the crystal structure contains both enantiomers of compound 13 as racemate. Moreover, the enantiomeric pairs are linked by weak C-H . . . O hydrogen bonds (Figure 2, right). Compound 13 exhibits C-C, C-O and C-Br bond lengths that are in the expected range. Both the lactone rings C1-C2-C3-O1-C4 and C1-C5-C6-O3-C4 exhibit envelope conformation with the central C1 atom at the flap position. However, C1 differs only 8.2 and 11.1 pm, respectively, from the corresponding mean planes. The interplanar angle between both lactone rings is 64.4 • .  It seems convenient to perform a resolution of the racemate at this stage. An enantioselective hydrolysis of an ester with a suitable enzyme appeared to be particularly attractive. Thus, 12 was activated using thionyl chloride (Scheme 2) to afford in situ the acid chloride 14, the reaction of which with n-hexanol produced ester 15. and 95% of (-)-19; (h) CH 3 NH 2 , TosMic, DCM, benzene, NEt 3 , 7 d, 23 • C, 59% (of (+1)-1 and 60% of (-)-1; Hex stands for n-hexyl.

Conclusions
Furan-2-carboxylic acid served as a valuable starting material for the straightforward synthesis of dehydro-homopilopic acid. Esterification, hydrogenation and enzymatic hydrolysis followed by the reduction of Weinreb amides and the single-step attachment of a 1-methyl-imidazole residue allowed for the convenient synthesis of both enantiomers of pilocarpine in good overall yields.

Experimental
NMR spectra were recorded using the Varian spectrometers (Darmstadt, Germany) DD2 and VNMRS (400 and 500 MHz, respectively). MS spectra were taken on a Advion expression L CMS mass spectrometer (Ithaca, NY, USA); positive ion polarity mode, solvent: methanol, solvent flow: 0.2 mL/min, spray voltage: 5.17 kV, source voltage: 77 V, APCI corona discharge: 4.2 µA, capillary temperature: 250 • C, capillary voltage: 180 V, sheath gas: N 2 ). Thin-layer chromatography was performed on pre-coated silica gel plates supplied by Macherey-Nagel (Düren, Germany). IR spectra were recorded on a Spectrum 1000 FT-IRspectrometer from Perkin Elmer (Rodgau, Germany). The UV/Vis-spectra were recorded on a Lambda 14 spectrometer from Perkin Elmer (Rodgau, Germany); optical rotations were measured at 20 • C using a JASCO-P2000 instrument (JASCO Germany GmbH, Pfungstadt, Germany) The melting points were determined using the Leica hot stage microscope Galen III (Leica Biosystems, Nussloch, Germany) and are uncorrected. The solvents were dried according to usual procedures. Microanalyses were performed with an Elementar Vario EL (CHNS) instrument (Elementar Analysensysteme GmbH, Elementar-Straße 1, D-63505, Langenselbold, Germany). The crystal structure of compound 13 was solved by direct methods (SHELXS) and refined with the SHELXL (2008) program [73]. OLEX2 (2021) was used as graphical user interface [74]. The hydrogen atoms were positioned geometrically using a riding model. The crystal structure drawings were generated with DIAMOND (4.4.0) [75]. CCDC 2086056 contains the supplementary crystallographic data for this paper. These data can be obtained free of charge from The Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/structures.

Data availability Statement:
The data presented in this study are available on request from the corresponding author.