Synthesis and Characterization of 2-Thiophenemethanamine-²H₅ Hydrochloride

Abstract

Synthesis of the key building block, 2-thiophenemethanamine-²H₅ hydrochloride, was achieved using mild conditions and purification methods in three steps from commercially available thiophene-²H₄, with an overall yield of 61.6% and an overall isotopic enrichment of 87.6%. 2-thiophenemethanamine-²H₅ hydrochloride has the potential to be a useful intermediate in the synthesis of isotopically labelled compounds of pharmaceutical interest.

1 |. Introduction

2-Thiophenemethanamine is a useful reagent in the preparation of potential pharmaceutical compounds and is readily commercially available at a low cost. A couple of examples of potential pharmaceuticals that contain the 2-thiophenemethanamino moiety are shown in Figure 1. Compound A [1] is described in a patent application for the treatment of cancer, and Compound B is described in a patent application for the treatment of drug-resistant malaria [2].

FIGURE 1 |.

Literature compounds containing a 2-thiophenemethanamino moiety.

A deuterium labelled version of 2-thiophenemethanamine would be useful for the synthesis of stable isotopically labelled standards of pharmaceuticals. This paper describes the synthesis and characterization of 2-thiophenemethanamine-²H₅ hydrochloride from the commercially available starting material thiophene-²H₄, in three synthetic steps.

2 |. Results and Discussion

During our studies, it was determined that a deuterium labelled version of 2-thiophenemethanamine would be useful for the synthesis of stable isotopically labelled standards of potential pharmaceuticals our group was researching. Indeed, isotopically labelled 2-thiophenemethanamines, containing one or two deuterium atoms, are known (CAS: 3050653-11-4, 1621713-89-0, 2587191-56-6), with deuterium substitution either at the methylene carbon or at the 5-position of the thiophene ring. For use as a bioanalytical standard, maximum mass increase above the molecular weight of native material provides clear analytical differentiation, so for our studies, we desired a molecule that contained isotopic labelling of plus 5 Da. It was realized that synthesizing 2-thiophenemethanamine-²H₅ would allow us to access our molecule of interest with the desired molecular weight increase of 5 Da. Our methodology for the synthesis of 2-thiophenemethanamine-²H₅ is shown in Figure 2.

FIGURE 2 |.

Synthesis of 2-thiophenemethanamine-²H₅.

Our strategy began with bromination of commercially available thiophene-²H₄ at the 2 position. Multiple bromination techniques were investigated. Cyanation of the resulting 2-bromo thiophene-²H₃ and reduction to 2-thiophenemethanamine-²H₅ with the introduction of two deuterium atoms on the resulting methylene carbon completed the strategy.

Initially, several bromination conditions were tried on non-deuterated thiophene, as a model system (HBr, hydrogen peroxide, ether) [3], (N-bromosuccinimide [NBS], acetic acid, chloroform), (NBS, ultrasound, heptane) [4]. These reactions, however, were not selective and gave the poly brominated products 2,5-dibromothiophene, 2,3,4-tribromothiophene, and 2,3,5-tribromothiophene or gave no brominated products at all.

Finally, NBS with a solvent screen of N,N-dimethylformamide, dichloromethane, acetonitrile, tetrahydrofuran, and dimethylsulfoxide was attempted. NBS in 50 volumes of acetonitrile, at 50°C, emerged as giving the best ratio of 2-bromothiophene to 2,5-dibromothiophene (approx. 3.7:1) and was selected as the preferred bromination method.

Purification of the reaction mixture by vacuum distillation caused 2-bromothiophene to co-distil with acetonitrile. Removal of the acetonitrile under atmospheric pressure distillation caused significant degradation of the 2-bromothiophene with the subsequent reduction in yield, due to prolonged heating.

It was shown that pure 2-bromothiophene could be obtained by flash chromatography, but significant amounts of product were lost during rotary evaporation of the collected fractions due to co-distillation with the eluent solvents.

A purification process using high vacuum under cold conditions was developed to allow removal of the volatile reaction products and solvents from the succinimide residues. After the reaction was complete, the reaction mixture was cooled to room temperature and then frozen in a liquid nitrogen bath, under argon. The frozen mixture was then placed under high vacuum (4–13 Pa) with a transfer line leading to a cold finger trap installed between the frozen flask and the vacuum pump. The cold finger trap was cooled in liquid nitrogen and the frozen flask was allowed to warm to room temperature. During the warming phase, all the volatile solvent and brominated thiophene components distilled over to the cold finger trap and solidified. Once all the material had been transferred, the material in the cold finger trap was allowed to warm to room temperature to give a mixture of brominated thiophene compounds as a solution in acetonitrile.

When the bromination reaction was performed on thiophene-²H₄ (1) in acetonitrile, it was shown by mass spectrometry that some deuteron/proton exchange was occurring. This exchange was mitigated by performing the reaction in acetonitrile-²H₃. The cold transfer purification process allowed the expensive acetonitrile-²H₃ to be recovered later in the process and potentially recycled. The resulting 2-bromothiophene-²H₃ (2) was used as is, without purification, due to its inability to be separated from the solvent by evaporation, having a high volatility, and due to the possibility of degradation, as described above. Purity was determined to be 75% using GC/MS analysis. The structure was confirmed using GC/MS and NMR analysis. GC/MS comparison with an unlabelled standard confirmed the same retention time and expected molecular weight. Proton decoupled ¹³C NMR comparison with an unlabelled standard confirmed the same chemical shifts, without the splitting associated with deuterium-carbon coupling.

The reaction of 2-bromothiophene-²H₃ (2), as a solution in acetonitrile-²H₃, with sodium cyanide in the presence of nickel ferrite and potassium carbonate in N,N-dimethylformamide [5] did not give the desired product. Zinc cyanide in the presence of zinc, tri (dibenzylideneacetone)dipalladium, and 1,1-bis (diphenylphosphino)ferrocene in N,N-dimethylacetamide [6] did, however, give the desired 2-thiophenecarbonitrile-²H₃ (3).

Once the reaction to form 2-thiophenecarbonitrile-²H₃ (3) had progressed to completion, the acetonitrile-²H₃ was able to be recovered by atmospheric pressure distillation, and the concentrated residue—a mixture of 2-thiophenecarbonitrile-²H₃ (3), 2,5-thiophenedicarbonitrile-²H₂, and other minor impurities—was able to be purified using normal phase flash chromatography. 2-Thiophenecarbonitrile-²H₃ (3) is less volatile than 2-bromothiophene-²H₃ (2) and was able to be isolated from the chromatography fractions after rotary evaporation, without loss due to co-distillation, giving 2-thiophenecarbonitrile-²H₃ (3) in 54% yield, over the 2 telescoped steps. The structure was confirmed using GC/MS and NMR analysis. GC/MS comparison with an unlabelled standard confirmed the same retention time and expected molecular weight. Proton decoupled ¹³C NMR comparison with an unlabelled standard confirmed the same chemical shifts, without the splitting associated with deuterium-carbon coupling.

2-Thiophenecarbonitrile-²H₃ (3) was treated with lithium aluminum deuteride as the reducing agent. We realized that isolation as the hydrochloride salt provided a convenient purification and isolation method after filtration, and so 2-thiophenemethanamine-²H₅ hydrochloride (4) was obtained in 79.5% yield after workup and salt formation. The structure was confirmed using LC/MS and NMR analysis. LC/MS comparison with an unlabelled standard confirmed the same retention time and expected molecular weight. Proton decoupled ¹³C NMR comparison with an unlabelled standard confirmed the same chemical shifts, without the splitting associated with deuterium-carbon coupling.

2-Thiophenemethanamine-²H₅ hydrochloride (4) did contain small amounts of partially protonated analogs. The chemical shifts of these protons corresponded with the ¹H NMR of an unlabelled standard. Only singlet peaks in the ¹H NMR spectrum suggested that there was no bis-protonation or tris-protonation occurring on adjacent ring carbons. The highest proportion of protonation occurred at the 5-position on the thiophene ring. The overall isotopic enrichment was calculated to be 87.6%.

3 |. Conclusion

Previously unreported, fully deuterium labelled, 2-thiophenemethanamine-²H₅ hydrochloride was synthesized from commercially available thiophene-²H₄, in three steps with an overall yield of 61.6% and an overall isotopic enrichment of 87.6%.

4 |. Materials and Instrumentation

Gas chromatography (GC/MS) analyses were performed using Agilent 6890 GC and Rtx-5 Sil MS, 30M, 0.25 mmID, 0.5 μm df column; splitless injection mode; 250°C inlet temperature; 1 μL injection volume, 1.5 mL/min (constant flow) helium, carrier gas flow; gradient of 35°C (2 min hold), 20°C/min to 330°C (5 min hold). High-resolution mass spectra were obtained using Agilent MSD 5975 Mass Spectrometer; column Rtx-5 Sil MS, 30M, 0.25 mmID, 0.5 μm df;1 μL injection volume; injector temperature 150°C; detector temperature 230°C; (EI) ~70 eV.

Ultra-performance liquid chromatography (UPLC-UV/MS) analyses were performed using Waters Acquity Ultra Performance LC and Waters Symmetry C18 (4.6 × 75 mm, 3.5 μm); temperature 30°C; wavelength, 254; injection 5 μL; flow 1.0 mL/min; run time 10 min; gradient A: 0.1% formic acid in water, B: 0.1% formic acid in acetonitrile, t₀: 10% B, t₁ 10% B, t_1–6 90% B, t_6–7 90% B, t_7–7.1 10% B, t_7.1–10 10% B, unless indicated otherwise. High resolution mass spectra were obtained using Agilent HP 6890 Mass Spectrometer; column Agilent DB-1 (30 mm × 0.32 mm ID, 0.25 μm film thickness); injection 1 μL; injector temperature 150°C; detector temperature 250°C; (EI) ~70 eV. NMR spectra were recorded with a Bruker 400 MHz spectrometer using deuterated solvents, as stated. Thiophene-[²H₄] (98 atom % ²H) was purchased from Arctom Scientific (Westlake Village, CA, USA); Lithium Aluminum Deuteride-²H₄ (LAD) (99.4 atom % ²H) was purchased from Oakwood Chemicals. Remaining reagents were purchased from Oakwood Chemicals (Estill, SC, USA).

5 |. Experimental

5.1 |. 2-Bromothiophene-²H₃ (2)

Thiophene-²H₄ (1) (10 g, 113.4 mmol, 1 eq), N-bromosuccinimide (25 g, 140.5 mmol, 1.2 eq), and acetonitrile-²H₃ (500 mL) were heated together at 50°C for 16 h. The reaction was cooled to room temperature and then frozen in liquid nitrogen. A vacuum was applied (4 Pa) to the frozen reaction mixture, and all the volatile components distilled out of the flask and were condensed, and solidified, in a liquid nitrogen cooled cold-finger trap. During the transfer, the pressure rose to 13 Pa. After being allowed to melt, the resulting solution free of succinimide byproducts was used, as is, in the next reaction. The mixture contained 2-bromothiophene-²H₃ (2) and 2,5-dibromothiophene-²H₂ in an approximately 3.7:1 ratio in approximately 500 mL of acetonitrile-²H₃, as shown by GC/MS analysis. ¹³C NMR (CD₃CN, 100 MHz) δ: 131.3 (t, J = 26 Hz), 129.0 (t, J = 16 Hz), 128.8 (t, J = 18 Hz), 112.4 (t, J = 10 Hz).

Observed mass [MH+] 167.00, calculated mass 166.05, C₄D₃BrS.

5.2 |. 2-Thiophenecarbonitrile-²H₃ (3)

The crude 2-bromothiophene-²H₃ and acetonitrile-²H₃ mixture (500 mL of acetonitrile-²H₃) from the previous reaction was charged with tris (dibenzylideneacetone)dipalladium (3.2 g, 3.5 mmol, 0.03 eq), 1,1′-ferrocenediyl-bis (diphenylphosphine) (3.8 g, 6.9 mmol, 0.06 eq), zinc powder (1.3 g, 19.9 mmol, 0.18 eq), zinc cyanide (17.5 g, 149 mmol, 1.3 eq), and anhydrous N,N-dimethylacetamide (100 mL). The mixture was heated to reflux for 3 h. The reaction was monitored by thin layer chromatography (normal phase silica plate, 40% ethyl acetate in heptane eluent, 254 nm UV lamp). Once all the starting material had been consumed, the reaction was cooled to room temperature and filtered. The filtrate was concentrated by distillation at atmospheric pressure, to remove excess volatile solvents, allowing recovery of the acetonitrile-²H₃. The concentrated residue was partitioned between water (500 mL) and dichloromethane (100 mL). The organic phase was separated, dried over anhydrous sodium sulfate, filtered, and evaporated. The residue was purified using a normal phase 120-g disposable silica gel column, eluting with a linear gradient of 100% heptane to 40% dichloromethane in heptane to afford 2-thiophenecarbonitrile-²H₃ (3) as a clear, yellow oil. (6.8 g, 61 mmol, 100% purity by GC/MS, 53.8% yield over the two telescoped steps). ¹³C NMR (THF-d₈, 100 MHz) δ: 138.0 (t, J = 26 Hz), 133.5 (t, J = 29 Hz), 128.0 (t, J = 27 Hz), 114.2 (s), 110.4 (s).

Observed mass [MH+] 113.10, calculated mass 112.16, C₅D₃NS.

5.3 |. 2-Thiophenemethanamine-²H₅ Hydrochloride (4)

Lithium aluminum deuteride (4.7 g, 112 mmol, 1.5 eq) was charged to a dry flask and flushed with argon for 15 min. Anhydrous tetrahydrofuran (100 mL) was added, and the mixture was cooled to 0°C in an ice bath. 2-Thiophenecarbonitrile-²H₃ (3) (8.2 g, 73.2 mmol, 1 eq) dissolved in anhydrous tetrahydrofuran (23 mL) was added, dropwise, over 10 min. The reaction was stirred at 0°C for a further 10 min then allowed to warm to room temperature. The reaction was monitored using thin layer chromatography (normal phase silica plate, ethyl acetate eluent, 254 nm UV lamp), which confirmed that all the 2-thiophenecarbonitrile-²H₃ had been consumed. The reaction mixture was cooled again to 0°C then quenched by the slow addition of 2-propanol (25 mL), water (13 mL), and then 5-M aqueous sodium hydroxide solution (100 mL). The resulting slurry was filtered and the filtrate was extracted with tert-butyl methyl ether (6 × 50 mL). The combined organic extracts were dried over anhydrous sodium sulfate and filtered, and the filtrate was acidified with 2-M hydrogen chloride in diethyl ether. The resulting slurry was filtered, and the solids were dried at the pump to afford 2-thiophenemethanamine-²H₅ hydrochloride (4) as an off-white powder (5.8 g, 58.2 mmol, 79.5% yield, 98.95% purity by UPLC-UV/MS). ¹³C NMR (D₂O, 100 MHz) δ: 133.3 (s), 129.2 (t, J = 20 Hz), 127.6 (t, J = 27 Hz), 127.3 (t, J = 26 Hz), 36.8 (p, J = 23 Hz).

Observed mass [M-NH₂] 102.03, calculated mass 118.21, C₅H₂D₅NS (freebase).

Data Availability Statement

Author elects not to share data.