Methyl 6,7-Difluoro-2-[(4-fluorobenzyl)sulfanyl]-4-hydroxyquinoline-3-carboxylate

Abstract

A convenient synthesis of a novel fluoroquinolone precursor, methyl 6,7-difluoro-2-[(4-fluorobenzyl)sulfanyl]-4-hydroxyquinoline-3-carboxylate, at a 78% yield starting from 3,4-difluorophenyl isothiocyanate was developed. The structure of the product was established by ¹H, ¹³C, and ¹⁹F NMR spectroscopy, mass spectrometry, IR spectroscopy and confirmed by elemental analysis. The title compound, containing the pharmacophoric 4-fluorobenzyl group, will be used in the synthesis of novel fluoroquinolone derivatives.

1. Introduction

Quinolones are an important group of quinoline derivatives that have a carbonyl group at the 4-position. Historically, nalidixic acid [1] (1-ethyl-7-methyl-4-oxoquinoline-3-carboxylic acid) has been considered to be the first of the synthetic quinolone antibiotics. However, nalidixic acid, which is classified as the first-generation quinolone antibiotic, does not contain a fluorine atom in its structure. The first generation also included other quinolone drugs, such as oxolinic acid and cinoxacin, which were introduced in the 1970s. Extensive research, including the study of the biological activity of various functionalized quinolone derivatives, has led to the development of numerous fluoroquinolone drugs [1,2,3,4,5,6]. The introduction of a fluorine atom at the C-6 position of the quinolone ring distinguishes later-generation fluoroquinolones from the first generation.

The first drugs containing the fluorine atom belong to second-generation quinolone antibiotics [1,2,3,4]. Almost all quinolone drugs widely used in medicine today are fluoroquinolones, which have a 4-quinolone core structure and a fluorine atom usually located at the 6-position. Fluoroquinolone antibiotics are broad-spectrum bactericidal drugs that are effective against both Gram-negative and Gram-positive bacteria and are used in human and veterinary medicine, including poultry and livestock, for the treatment of bacterial infections [1,2,3,4,5].

Examples of widely used fluoroquinolone antibiotics such as ciprofloxacin (one of the most widely used antibiotics in the world), levofloxacin (the third-generation class of fluoroquinolone antibiotics), temofloxacin (the third generation), and prulifloxacin (the fourth generation) are presented in Figure 1. It is worth noting that prulifloxacin [7], the antibiotic of the fourth generation, has a sulfur atom at the 2-position of the quinolone ring (Figure 1).

The 4-carbonyl and 3-carboxylate groups are considered important for the manifestation of antibacterial activity. These two positions on the quinolone ring are essential for gyrase binding and bacterial transport. It is known that the introduction of the fluorine atom at the 6-position had markedly improved antimicrobial activity compared to the analogous quinolone compounds and gave rise to clinically successful and widely used fluoroquinolone antibiotics [2,3,4,5,6].

Fluoroquinolones, introduced into medical practice since the mid-1990s (III-IV generations), including prulifloxacin [7], are characterized by higher activity against Gram-positive bacteria (primarily pneumococci), intracellular pathogens, and anaerobes (IV generation) [5,6,7,8].

It has been recently found that some quinolones exhibit high antivirus activity, e.g., 2-aminoquinolone acid derivatives have been discovered to act as potent inhibitors of the SARS-CoV-2 virus [9].

A number of synthetic approaches to fluoroquinolones involve the preparation of 3-carboxy-6-fluoro-4-hydroxyquinoline derivatives as fluoroquinolone precursors. Developing a convenient synthesis of these compounds is a decisive factor in fluoroquinolone preparation since their conversion to fluoroquinolone derivatives by alkylation on the nitrogen atom of the quinoline ring or by other reactions is usually straightforward [1,2,3,4,10,11,12,13].

The development of efficient methods for the synthesis of new derivatives of nitrogen/sulfur heterocycles with practically valuable properties, including biological activity, is within the scope of our scientific interests [14,15,16]. An efficient synthesis of a number of water-soluble quinoline derivatives fused with thiazine and thiazole heterocycles ([1,4]thiazino[2,3,4-ij]quinolin-4-ium and [1,3]thiazolo[3,2-a]quinolin-10-ium derivatives) has been developed [15,16]. Some of the obtained products, [1,4]thiazino[2,3,4-ij]quinolin-4-ium derivatives, exhibit high antibacterial activity [15].

2. Results and Discussion

We developed a convenient and efficient synthesis of a novel fluoroquinolone precursor, methyl 6,7-difluoro-2-[(4-fluorobenzyl)sulfanyl]-4-hydroxyquinoline-3-carboxylate 2, at a 78% yield from 3,4-difluorophenyl isothiocyanate 1, dimethyl malonate, and 4-fluorobenzyl chloride (Scheme 1).

The starting compound 1 was prepared by a known method [10] using the reaction of 3,4-difluoroaniline with carbon disulfide in triethylamine followed by the conversion of triethyl ammonium 3,4-difluorophenyl dithiocarbamate to 3,4-difluorophenyl isothiocyanate by its reaction with ethyl chloroformate.

The deprotonation of dimethyl malonate was carried out with potassium tert-butylate in acetonitrile. After the reaction of dimethyl malonate carbanion with isothiocyanate 1, 4-fluorobenzyl chloride was added as an alkylating reagent. The reaction pathway includes the nucleophilic addition of the (MeOOC)₂CH⁻ anion to the thiocyanate group, the benzylation of the intermediate 3 with 4-fluorobenzyl chloride, and cyclization of the formed 4-fluorobenzyl sulfide 4, which was used without purification in the next stage (Scheme 2).

Cyclization can be considered as an intramolecular electrophilic substitution reaction of the Friedel–Crafts type with an ester group, which usually occurs under harsh conditions. The cyclization stage was carried out by heating in diphenyl ether at 145–148 °C with stirring under nitrogen. These conditions allowed for the removal of methanol, which was formed during the reaction. The target compound 2 was obtained at a 78% yield based on isothiocyanate 1 taken.

The development of an efficient and convenient synthesis of 3-carboxy-6-fluoro-4-hydroxyquinoline derivatives, fluoroquinolone precursors, is of great importance because these compounds can be readily converted to corresponding 3-carboxy-6-fluoro-4-quinolones [1,2,3,4,10,11,12,13]. The sulfur atom is present at position 2 of the quinolone ring of prulifloxacin, the fourth-generation antibiotic [7]. However, in general, very few synthetic methods are known for the preparation of sulfur-containing quinolones, given that the sulfur atom can be located in various positions in the quinolone molecule.

Based on product 2, we plan to obtain a series of new fluoroquinolone derivatives [2-(4-fluorobenzylsulfanyl)-6-fluoro-4-oxo-7-piperazino-3-quinolinecarboxylic acids] with the substitution of the fluorine atom in the 7-position by the piperazine heterocycle and the introduction of various functional substituents at the nitrogen atom of the quinoline core (the relevant methods of which are known [10,11,12,13]). Compounds of this series are unknown, but they may exhibit high antibacterial activity. It is worth noting that the 4-fluorobenzyl moiety is considered a pharmacophoric group, which often increases biological activity, including antibacterial properties [17,18,19,20].

The structural assignments of compound 2 were made using ¹H, ¹³C, and ¹⁹F NMR spectroscopy, mass spectrometry, and IR spectroscopy and confirmed by elemental analysis. The complete assignment of ¹H and ¹³C NMR chemical shifts of compound 2 was carried out by the analysis of values of spin–spin coupling constants (including ¹J_C-F, ²J_C-F, ³J_C-F, ⁴J_C-F, and ⁵J_C-F) and a combination of 1D (¹H and ¹³C NMR) and 2D (¹H-¹H COSY and ¹H-¹³C HMBC) NMR experiments.

Compound 2 is supposed to exist in equilibrium between the enol and keto forms of the bicyclic quinoline system, with the enol form predominating (Scheme 3). The hydroxyl group appears downfield as a broad signal at 13.16 ppm in the ¹H NMR spectrum of compound 2. The carbon signals of the C-OH and carbonyl (MeOC=O) groups are observed in the downfield region at 167.73 and 171.04 ppm, respectively, in the ¹³C NMR spectrum of compound 2.

The ¹⁹F NMR spectrum of compound 2 shows three signals at −137.06 (C⁶F), −127.02 (C⁷F), and –115.23 (C¹⁵F) ppm. Several carbon–fluorine spin–spin coupling constants (¹J_C-F, ²J_C-F, ³J_C-F, ⁴J_C-F, and ⁵J_C-F) are observed in the ¹³C NMR spectrum of compound 2. The values of direct carbon–fluorine spin–spin coupling constants (¹J_C-F) are 245.5–257.5 Hz, while those for ²J_C-F are considerably lower (15.2–21.4 Hz).

In the mass spectrum of compound 2, the molecular ion 379 (29.4%) and maximum ion (109) corresponding to the 4-fluorobenzyl group are observed.

3. Experimental Section

3.1. General Information

The ¹H (400 MHz), ¹³C (100 MHz), and ¹⁹F (376 MHz) NMR spectra (see Supplementary Materials) were recorded on a Bruker DPX-400 spectrometer (Bruker BioSpin GmbH, Rheinstetten, Germany) in CDCl₃ solutions and referred to CFCl₃ (¹⁹F NMR, external) and to the residual solvent peaks of CDCl₃ (δ = 7.27 and 77.16 ppm for ¹H and ¹³C NMR, respectively). The mass spectrum was recorded on a Shimadzu GCMS-QP5050A (Shimadzu Corporation, Kyoto, Japan) with electron impact (EI) ionization (70 eV). Elemental analysis was performed on a Thermo Scientific Flash 2000 Elemental Analyzer (Thermo Fisher Scientific Inc., Milan, Italy). The melting point was determined on a Kofler Hot-Stage Microscope PolyTherm A apparatus (Wagner & Munz GmbH, München, Germany). The organic solvents were dried and distilled according to standard procedures. Silica gel (Alfa Aesar, Haverhill, MA, USA, 70–230 mesh) was used for column chromatography.

3.2. Synthesis of Compound 2

A solution of dimethyl malonate (0.264 g, 2 mmol) in MeCN (0.5 mL) was added dropwise to a suspension of t-BuOK (0.24 g, 2.14 mmol) in MeCN (3.5 mL) with stirring and the mixture was stirred for 3 h at room temperature. A solution of isothiocyanate 1 (0.342 g, 2 mmol) in MeCN (1 mL) was added with stirring, and the mixture was stirred overnight (16 h) at room temperature. Then, a solution of 4-fluorobenzyl chloride (0.32 g, 2.2 mol) in 1 mL of MeCN was added dropwise, and the mixture was stirred for 4 h at room temperature. The mixture was filtered, and the solvent was removed from the filtrate on a rotary evaporator. Diphenyl ether (3 mL) was added to the residue, and the mixture was heated at 145−148 °C while stirring for 5 h in a flask under nitrogen bubbling in such a way as to remove methanol that was formed during the reaction. The resulting product was subjected to column chromatography on silica gel (eluent: hexane → hexane/chloroform 3:1) to give compound 2 at a 78% yield (0.592 g) based on the isothiocyanate 1 taken.

Methyl 6,7-difluoro-2-[(4-fluorobenzyl)sulfanyl]-4-hydroxyquinoline-3-carboxylate (2), 78% yield, white solid, m.p. 152–153 °C.

¹H NMR (400 MHz, CDCl₃): 4.05 (s, 3H, OCH₃), 4.46 (s, 2H, CH₂S), 7.00 (dd, 2H, C¹⁴H, C¹⁶H, ³J_H-F = 8.3, ³J_H-H = 8.6 Hz), 7.43 (dd, 2H, C¹³H, C¹⁷H, ⁴J_H-F = 5.6, ³J_H-H = 8.6 Hz), 7.57 (dd, 1H, C⁸H, ³J_H-F = 11.0, ⁴J_H-F = 7.3 Hz), 7.92 (dd, 1H, C⁵H, ³J_H-F = 10.2, ⁴J_H-F =9.0 Hz), 13.16 (s, 1H, OH).

¹³C NMR (100 MHz, CDCl₃): 34.95 (C¹¹), 52.83 (MeO), 102.32 (C³), 110.28 (dd, C⁵, ²J_C-F = 19.2, ³J_C-F = 2.1 Hz), 114.34 (d, C⁸, ²J_C-F = 16.9 Hz), 114.73 (d, C¹⁰, ³J_C-F = 7.2 Hz), 115.42 (d, C^14,16, ²J_C-F = 21.4 Hz), 131.02 (d, C^13,17, ³J_C-F = 8.0 Hz), 133.70 (d, C¹², ³J_C-F = 3.3 Hz), 146.32 (d, C⁹, ³J_C-F = 10.9 Hz), 149.10 (dd, C⁶, ¹J_C-F = 250.9, ²J_C-F = 15.2 Hz), 154.71 (dd, C⁷, ¹J_C-F = 257.5, ²J_C-F = 15.7 Hz),. 160.17 (d, C², ⁵J_C-F = 2.2 Hz), 162.08 (d, C¹⁵, ¹J_C-F = 245.5 Hz), 167.73 (dd, C⁴, ⁴J_C-F = 4.4, ⁵J_C-F = 1.1 Hz), 171.04 (C=O).

¹⁹F NMR (376 MHz, CDCl₃): −115.08–−115.37 (m, C¹⁵F), −126.88–−127.15 (m, C⁷F), −136.90–−137.19 (m, C⁶F).

IR (KBr, ν/cm⁻¹): 3350, 2920, 2850, 1658, 1594, 1506, 1449, 1315, 1214, 1139, 997, 949, 864, 753, 592, 524.

MS, m/z: 379 (M⁺, 29.4%), 347 (41.2%); 314 (16.4%); 286 (8.9%); 270 ([M−FC₆H₄CH₂]⁺, 3.5%); 208 (4.7%); 181 (23.1%); 166 (10.6%); 139 (10.1%); 109 ([FC₆H₄CH₂]⁺, 100%); 83 (22.0%); 57 (5.4%); 45 (8.2%).

Anal. calcd for C₁₈H₁₂F₃O₃NS (379.35): C 56.99, H 3.19, N 3.69, S 8.45%. Found: C 57.24, H 3.31, N 3.82, S 8.57%.