Rhodanine derived enethiols react to give 1,3-dithiolanes and mixed disulfides

Rhodanines have been characterised as ‘difficult to progress’ compounds for medicinal use, though one rhodanine is used for diabetes mellitus treatment and others are in clinical development. Rhodanines can undergo hydrolysis to enethiols which are inhibitors of metallo-enzymes, such as metallo β-lactamases. We report that in DMSO, rhodanine derived enethiols undergo dimerisations to give 1,3-dithiolanes and mixed disulfides. The results highlight the potential of rhodanines and enethiols to give multiple products. They suggest that where possible DMSO should be avoided as a storage solvent for rhodanines/enethiols and highlight the need for further research on biologically relevant enethiols/mixed disulfides.


General Experimental Conditions
The enethiols (1-8b) were prepared using reported methods and conditions. 2,3Starting materials and products formed were monitored by NMR and characterized by 1D and 2D NMR techniques.Samples were recorded at 298 K, unless stated otherwise, in a 5 mm tube, using a Bruker AVII 500 equipped with a TXI H/F/C probe, Bruker AVIII HD 500 equipped with a BBF probe, Bruker AVIII HD 600 equipped with a BB-F/H N2 CryoProbe or a Bruker AVIII HD 700 equipped with a TCI H/N/C He CryoProbe, at their respective resonances.Chemical shifts (δ) are reported in parts per million (ppm) referenced to the solvent resonance.Coupling constants (J) are reported to the nearest 0.5 Hz.Multiplicities are reported as singlet (s), doublet (d), triplet (t), multiplet (m).δC values are derived from 2D 1 H- 13 C heteronuclear single quantum coherence spectroscopy (HSQC) and 1 H- 13 C heteronuclear multiple bond correlation (HMBC) experiments.When specified, liquid chromatography mass spectrometry (LC-MS) was carried out using a Waters LCT Premier bench-top orthogonal acceleration time-of-flight (TOF) LC-MS system, equipped with a Acquite PDA detector.Reactions under anaerobic conditions (<2 ppm O2) were performed in an anaerobic chamber (Belle technology, UK), under a N2 atmosphere.
The standard reaction conditions for monitoring enethiol reactions were as follows.The solid enethiol was dissolved in DMSO-d6 and diluted to give a solution (450 μL, 5 or 11 mM) of the desired concentration.The solution was transferred into an NMR tube (5 mm) and monitored over time.Reactions of enethiols in different solvents were studied in an analogous manner using the appropriate deuterated solvent, that is CD3OD, 1,4-dioxane-d8, DMF-d7, THF-d8, or acetone-d6.For reactions carried out under anaerobic conditions, the solid enethiol, DMSO-d6 (1 mL), and a J Young valve NMR tubes (5 mm, Norell, USA) were transferred to an anaerobic chamber (<2 ppm O2, Belle Technology, UK) and left to equilibrate for 24 hours, prior to use.The solid enethiol was then dissolved in DMSO-d6 and diluted appropriately (450 μL, 5 mM) before being transferred into the J Young valve NMR tube and monitored by 1 H NMR over time.

Figure S1: 1 H
Figure S1: 1 H-13 C HSQC and HMBC NMR (700, 176 MHz, 298 K) characterisation of 1c, the major observed product when 1b (5 mM) reacts in DMSO-d6 for 10 hours at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.Dashed lines represent through bond HMBC couplings, indicated by arrows in the structure of 1c.

Figure S2: 1 H
Figure S2: 1 H-13 C HSQC and HMBC NMR (700 MHz, 298 K) characterisation of 2d, the observed product when 2b (50 mM) reacts in DMSO-d6 for 2 hours at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.Dashed lines represent through bond HMBC couplings, which are labelled by arrows in the structure of 2d.The dashed lines and arrows are colour coded to match the couplings they represent.*Splitting of the 13 C signal as a result of 19 F coupling.

Figure S3: 1 H
Figure S3: 1 H NMR (600 MHz, 298 K) and 19 F NMR (565 MHz, 298 K) analyses of 2b (5 mM) in DMSO-d6 after reacting for 2 hours at room temperature to give 2d.Comparison of the integrals for H-2 and H-12, and F-9 and F-18 indicates a near 1 : 1 stoichiometry of the two 'components' forming the mixed disulfide 2d.

Figure S6: 1 H
Figure S6: 1 H NMR (600 MHz, 298 K) analysis of the reaction of 1b (5 mM) in methanol-d4 for 72 hours at room temperature.No evidence for substantial formation of 1c or 1d was observed.*Indicates potential formation of the symmetric unsaturated disulfide.

Figure S7: 1 H
Figure S7: 1 H NMR (600 MHz, 298 K) analysis of the reaction of 1b (5 mM) in THF-d8 for 72 hours at room temperature.No evidence for substantial formation of 1c or 1d was observed.

Figure S8: 1 H
Figure S8: 1 H NMR (600 MHz, 298 K) analysis of the reaction of 1b (5 mM) in acetone-d6 for 72 hours at room temperature.No evidence for substantial formation of 1c or 1d was observed.

Figure S9: 1 H
Figure S9: 1 H NMR (600 MHz, 298 K) analysis of the reaction of 1b (5 mM) in DMF-d7 for 72 hours at room temperature.Evidence for formation of 1c as indicated by the peaks (red box), albeit incomplete, was observed after 72 hours.

Figure S10: 1 H
Figure S10: 1 H NMR (600 MHz, 298 K) analysis of the reaction of 1b (5 mM) in 1,4-dioxane-d8 for 72 hours at room temperature.No evidence for substantive formation of 1c or 1d was observed.*Indicates potential formation of a symmetric unsaturated disulfide.

Figure S16 :
Figure S16: Incorporation of 2 H from 2 H2O into 1c based on relative 1 H NMR integrals.Percentages are derived using integrals from the 1 H NMR (600 MHz, 298 K) spectra, comparing the resonance corresponding to the indicated protons and non-overlapping aromatic resonances.

Figure S18 :
Figure S18: Incorporation of 2 H into 2d based on relative 1 H NMR integrals.Percentages are derived using relative integrals from the 1 H NMR (600 MHz, 298 K) spectra, comparing the resonance corresponding to the indicated protons and a non-overlapping aromatic resonance.

Figure S19: 1
Figure S19: 1 H-13 C HSQC and HMBC NMR (700, 176 MHz, 298 K) characterisation of 3c the major product formed when 3b (5 mM) reacts in DMSO-d6 for 1 hour at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.Dashed lines represent through bond HMBC couplings, which are labelled by arrows in the structure.The arrows represent couplings and correspond to the dashed lines which highlight couplings used to assign 3c.* 13 C signal split as a result of 19 F coupling.

Figure S20: 1
Figure S20: 1 H-13 C HSQC and HMBC NMR (700, 176 MHz, 298 K) characterisation of 4c and 4d, the major products formed when 4b (5 mM) reacts in DMSO-d6 for 20 minutes at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.Dashed lines represent through bond HMBC couplings, which are labelled by arrows in the structure.The arrows represent couplings and correspond to the colour matched dashed lines which highlight couplings used to assign 4c and 4d.* 13 C signal split as a result of 19 F coupling.

Figure S21: 1
Figure S21: 1 H-13 C HSQC and HMBC NMR (700, 176 MHz, 298 K) characterisation of 5d, the major product formed when 5b (5 mM) reacts in DMSO-d6 for 11 hours at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.Dashed lines represent through bond HMBC couplings, which are labelled by arrows in the structure.The arrows represent couplings and correspond to the dashed lines which highlight couplings used to assign 5d.

Figure S22: 1 H
FigureS22: 1 H-13  C HSQC and HMBC NMR (700, 176 MHz, 298 K) characterisation of 6d, the major product formed when 6b (5 mM) reacts in DMSO-d6 for 24 hours at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.The arrows represent couplings and correspond to the dashed lines which highlight couplings used to assign 6d.

Figure S23: 1 H
FigureS23: 1 H-13  C HSQC and HMBC NMR (700, 176 MHz, 298 K) characterisation of 7c and 7d, the major products formed when 7b (5 mM) reacts in DMSO-d6 for 24 hours at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.The arrows represent couplings and correspond to the dashed lines which highlight couplings used to assign 7c and 7d.

Figure S24: 1 H
FigureS24: 1 H-13  C HSQC and HMBC NMR (700, 176 MHz, 298 K) characterisation of 8c and 8d the major products formed when 8b (5 mM) reacts in DMSO-d6 for 24 hours at room temperature.Cross peaks of the phase sensitive HSQC are in blue and red.Green peaks represent HMBC through bond correlations.The arrows represent couplings and correspond to the colour matched dashed lines which highlight couplings used to assign 8c and 8d.

Figure S33 :
Figure S33: Comparison of the 1,3-dithiolane ring δC values reported by Castiñeiras et.al. for 9c and those for 1c assigned in this study. 1a) Structure of 1,3-thiolane ring and comparison of δC (ppm) values.b) Overlay of HSQC (blue and red cross peaks) and HMBC (green cross peaks); couplings establishing the 1,3-thiolane ring structure are highlighted.Note that 1c is used as an example for chemical shifts of the 1,3-dithiolane ring; similar chemical shifts are found of the other 1,3-dithiolane rings reported in this study.