First chemoenzymatic stereodivergent synthesis of both enantiomers of promethazine and ethopropazine

Summary Enantioenriched promethazine and ethopropazine were synthesized through a simple and straightforward four-step chemoenzymatic route. The central chiral building block, 1-(10H-phenothiazin-10-yl)propan-2-ol, was obtained via a lipase-mediated kinetic resolution protocol, which furnished both enantiomeric forms, with superb enantioselectivity (up to E = 844), from the racemate. Novozym 435 and Lipozyme TL IM have been found as ideal biocatalysts for preparation of highly enantioenriched phenothiazolic alcohols (up to >99% ee), which absolute configurations were assigned by Mosher’s methodology and unambiguously confirmed by XRD analysis. Thus obtained key-intermediates were further transformed into bromide derivatives by means of PBr3, and subsequently reacted with appropriate amine providing desired pharmacologically valuable (R)- and (S)-stereoisomers of title drugs in an ee range of 84–98%, respectively. The modular amination procedure is based on a solvent-dependent stereodivergent transformation of the bromo derivative, which conducted in toluene gives mainly the product of single inversion, whereas carried out in methanol it provides exclusively the product of net retention. Enantiomeric excess of optically active promethazine and ethopropazine were established by HPLC measurements with chiral columns.

S3 position 48, 19 mm hole depth, for circular top Hot plate stirrer. Melting points were obtained with an MPA100 Optimelt SRS apparatus and are uncorrected. Thin-layer chromatography was carried on TLC aluminum plates with silica gel Kieselgel 60 F 254 (Merck, Germany) (0.2 mm thickness film) using UV light as a visualizing agent. Preparative separations were carried out by: (i) column chromatography using Merck silica gel (230-400 mesh), with grain size 40-63 μm or by (ii) PLC PSC-Fertigplatten Kieselgel 60 F 254 (20 × 20 cm with 2 mm thickness layer) glass plates purchased from Merck, Germany. The gas chromatographic (GC) analyses were performed with an HP Series II 5890 instrument (Maryland, United States) equipped with a flame ionization detector (FID) and fitted with HP-50+ (30 m) semi-polar column; the GC injector was maintained at 250 °C; Column temperature programs are given for respective analytes in the compound characterization data paragraphs; Helium (2 mL/min) was used as carrier gas; retention times (t R ) are given in minutes under these conditions. The enantiomeric excesses (% ee) of optically active compounds were determined by high performance liquid chromatography (HPLC) analyses performed on Shimadzu CTO-10ASV chromatograph (Shimadzu Corporation, Japan) equipped with STD-20A UV detector and Chiralcel OD-H (Daicel Chemical Industries Ltd., Japan) chiral column using mixtures of nhexane/iso-propyl alcohol or Chiralcel OJ (Daicel Chemical Industries Ltd., Japan) chiral column using mixtures of n-hexane/ethyl alcohol or n-hexane/tert-butanol/triethyl amine as mobile phase in appropriate ratios given in experimental section (see later for further details for each individual compound); both chiral columns were analytical type, dimension 250 mm x 4.6 mm; the HPLC analyses were executed in an isocratic manner if not stated otherwise; flow (f) is given in mL/min; racemic compounds were used as standards; HPLC conditions and retention times (t R ) are given in Table S1. Optical rotations were measured with a P20 polarimeter (Belligham & Stanley Ltd.) in a 2 dm long cuvette using the sodium D line (589 nm); [α] D are given in units of: deg dm −1 cm 3 g −1 ; the concentration c is in g/100 mL. UV spectra were measured with a Varian Cary 3 UV-Visible Spectrophotometer (Varian, Inc., USA). 1 H and 13 C NMR spectra were measured with a Varian Mercury 400BB spectrometer (Varian, Inc., USA) operating at 400 MHz for 1 H and 100 MHz for 13 C nuclei; chemical shifts (δ) are given in parts per million (ppm) on the delta scale related to the solvent peak used as reference value; signal multiplicity assignment: s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; coupling constant (J) are given in hertz (Hz); All samples were recorded as solutions in fully deuterated chloroform (CDCl 3 ), methanol (CD 3 OD) or benzene (C 6 D 6 ), respectively. All NMR reports for Supporting Information document were created by ACD/NMR Processor Academic Edition 12.0. (Freeware software provided by ACD/Labs, USA & Canada) and show only delta range where signals were present. Mass spectra were S4 recorded on a Micro-mass ESI Q-TOF spectrometer in ESI mode; sample of analyte (2 mg) was prepared by dissolving it in methanol (1 mL). The spectrometer had an electrospray ion source and a linear ion trap analyzer. FT-IR spectra of neat samples were recorded on a Perkin Elmer System 2000 FTIR Spectrometer equipped with a Pike Technologies GladiATR attenuated total reflectance (ATR) accessory with a monolithic diamond crystal stage and a pressure clamp; FT-IR spectra were recorded in transmittance mode in the 300-4000 cm -1 range, in ambient air at room temperature, with 2 cm -1 resolution, 0.5 cm -1 interval and accumulation of 32 scans; unit are given in %T.
Compound characterization data and representative synthetic procedures:

Synthesis of 1-(10H-phenothiazin-10-yl)propan-2-ol (±)-3
Method A: To a stirred solution of 10H-phenothiazine 1 (2 g, 10 mmol) in dry THF (70 mL) at -78 °C n-butyllithium (9.4 mL of 1.6 M solution in hexane, 15 mmol) was added dropwise in gentle flow of argon. After 1 h of stirring, propylene oxide 2 (1.16 g, 20 mmol, 1.4 mL) was added dropwise, then cooling bath was removed, and the reaction mixture was stirred overnight at room temperature. The reaction mixture was subsequently quenched by adding finely crushed ice (10 g), then extracted with Et 2 O (3 × 20 mL). The combined organic layer was washed with water (100 mL), combined and dried over anhydrous MgSO 4 . Evaporation of the solvent gave crude product as an oily residue, which was purified by column chromatography on silica gel using mixture of hexane/AcOEt (80:20, v/v) as an eluent. After high-vacuum drying the desired alcohol crystallized as light-gray solid (1.84 g, 7.15 mmol, 71%).

Method B:
To a stirred solution of 10H-phenothiazine 1 (5 g, 25.1 mmol) in dry THF (120 mL) at -78 °C n-butyllithium (23.5 mL of 1.6 M solution in hexane, 37.6 mmol) was added dropwise in gentle flow of argon. After 1 h of stirring, propylene oxide 2 (2.91 g, 50.2 mmol, 3.5 mL) was added dropwise, then cooling bath was removed, and the reaction mixture was stirred overnight at room temperature. The reaction mixture was subsequently quenched by adding finely crushed ice (50 g), then extracted with Et 2 O (3 × 50 mL). The combined organic layer was washed with water (220 mL), isolated and dried over anhydrous MgSO 4 . After filtration and solvent evaporation under reduced pressure, an oily residue was purified by vacuum distillation thus obtaining amorphous non-crystalline solid (4.94 g, 19.2 mmol, 77%), which solidified after drying under high-vacuum. Attention: since phenothiazine 1 is lightsensitive reagent the above-mentioned reactions have been performed in flasks covered with aluminum folic.

General procedure for analytical-scale kinetic resolution of (±)-3 -acyl donor screening
The reaction mixture containing racemic (±)-3 (100 mg, 0.39 mmol), MTBE (2 mL Table 3 in the main manuscript; physical, spectroscopic and analytical data are identical as for the corresponding racemic standard compounds).

General procedure for gram-scale (Lipozyme TL IM)-catalyzed KR of (±)-3
The reaction was carried out under identical conditions as in the case of gram-scale synthesis

Crystal structure determination of (S)-(+)-5
Colorless single crystals, suitable for X-ray diffraction studies, were grown by slow diffusion of hexane (0.5 mL) into a concentrated solution of the (S)-(+)-5 (50 mg) in AcOEt (1 mL Software [4]. The structure was solved by direct methods using the SHELXS-97 structure solution program and refined by full-matrix least-squares against F 2 with SHELXL-2013 [5] and OLEX2 [6] programs. All non-hydrogen atoms were refined with anisotropic displacement parameters. Hydrogen atoms attached to carbon atoms were added to the structure model at geometrically idealized coordinates and refined as riding atoms with Uiso(H) = 1.2Ueq(CH and CH 2 ) or Uiso(H) = 1.5Ueq(CH 3 ). The hydrogen atom of the hydroxyl group was refined with a restraint of O-H = 0.84 (2)Å. An absolute (S)-configuration for the compound molecule (+)-5 was successfully determined using anomalous dispersion effects. Flack parameter [7] calculated from 991 selected quotients (Parsons' method) [8] equals 0.007 (4). Further analysis of the absolute structure was performed using likelihood methods with PLATON [9]. A total of 999 Bijvoet pairs (coverage of 1.00) were included in the calculations. The resulting value of the Hooft parameter [10] was 0.002 (4), with a P3 probability for an inverted structure smaller than 1×10 -100 . These results indicated that the absolute structure has been correctly assigned. Crystal Data for ( email: deposit@ccdc.cam.ac.uk]. ORTEP drawing was made using Ortep3 for Windows [11]. S11 Table 1 Crystal data and structure refinement parameters for (S)-(+)-5.