Replacement of thiocarbonyl

Rhodanine-like compounds are known as promiscuous binders to a range of molecular tar-gets.^[83] An analysis of rhodanine derivatives in protein X-ray structures have shown that the exo-cyclic thiocarbonyl group is involved in multiple molecular interactions such as hydrogen bonding and electrostatic effects.^[83] Although thiocarbonyl hydrogen bonds are less favoured compared to carbonyl hydrogen bond interactions, it has been suggested that water disso-ciation from thiocarbonyl residues is facilitated and this is a major factor in the formation of hydrogen bonds in thiocarbonyl groups.^[83]Therefore, substitution of the thiocarbonyl to a car-bonyl group could increase the selectivity towards anti-parasitic targets through the formation of specific hydrogen bond interactions with the carbonyl group, possibly reducing unwanted interactions. The substitution will also reveal the importance of a exo-cyclic thiocarbonyl group for anti-parasitic activity. The synthetic strategy towards N-acetic acid thiazolidine-2,4-dione analogues is outlined in Scheme 6.

N S

Scheme 7: Synthesis of thiazolidine-2,4-dione ester derivatives 7a–k; (i) EtOH, 80^◦C, for R¹ -R³see Table 4.

Initially, thiazolidine-2,4-dione was subjected to alkylation conditions with iodo- and bromo-acetic acid sodium salts (X=Br,I and R=Na) in the presence of sodium hydride in DMF (Scheme 6 Method 1). The reaction protocol was adapted from Bhat et al. The solvent was substituted with DMF for better solubility of the starting thiazolidine-2,4-dione (technical grade, 90 %).^[165]

However, the alkylation did not occur under these reaction conditions and only starting material was recovered. Using the adapted protocol with ethyl-2-bromoacetate as the alkylating agent yielded the desired ethyl ester7. The yield was dependant on the use of anhydrous solvents, varying from 86-90 % after column chromatography. At the same time a different protocol was explored for the synthesis of the ethyl ester7 (Scheme 6 Method 2). Treating thiazolidine-2,4-diones with potassium hydroxide in ethanol resulted in the formation of the potassium salt 8 after simple filtration in quantitative yields.^[166] The following alkylation was carried out in the presence of ethyl-2-bromoacetate to afford the product7 in excellent yields (>98 %) after sim-ple extraction. Both methods gave access to the ester derivative7, but the synthesis of the free carboxylic acid was not achieved. Instead of hydrolysing the ester under basic or acidic condi-tions,7 was chosen for subsequent Knoevenagel reactions with various aldehydes. The ester 7 can be seen as the pro-drug of the free carboxylic acid. Similar ester-modified compounds have been shown to improve membrane diffusion and consequently improve anti-parasitic ac-tivity.^[167] Within the parasite, the ester derivative could be hydrolysed and release the active free carboxylic acid, or the ester derivative binds directly to the desired target. The capability of hydrolysing ester analogues to free acids has previously been shown in T. brucei.^[167]

Derivatives of 7a–l were synthesised via Knoevenagel condensation with various ben-zaldehydes and sodium acetate as the base (Scheme 7). The synthesised analogues 7a–l showed improved solubility in ethanol. Because of this improved solubility only7j and 7a were afforded after simple filtration from ethanol, all other derivatives (7c, 7d, 7e, 7f, 7l, 7i, and 7k) were purified by column chromatography, explaining the particularly low yields of 31 % for7k.

The synthesis of the para-trifluoromethyl substituted derivative7g did not succeed under these

3 Rhodanine-N-acetic acid derivatives

Table 4: Synthesised thiazolidine-2,4-dione ester derivatives 7a–k including yields and NMR chemical shifts of CH signal (both ¹H and ¹³C); R¹-R³; yield; δ_C for 7k in MeOD otherwise all δ in CDCl₃; n.a.: not available.

[%] NMR shifts of CH

# R¹ R² R³ yield δ_H δ_C

7a H H H 46 7.95 n.a.

7b CH3 H H 69 8.07 132.8

7c H CH₃ H 71 7.91 135.1

7d H H CH₃ 71 7.75 134.8

7e CF3 H H 87 8.20 130.6

7f H CF₃ H 52 7.94 132.7

7g H H CF₃ no reaction

7h H OBn H 88 7.86 134.6

7i H H OBn 50 7.89 134.7

7j H OBn OBn 67 7.79 134.8

7k H OH OH 31-58 7.78 136.2

conditions. TLC showed complete consumption of starting7, but also the formation of multiple decomposition products. All products were afforded as the Z-isomer, which was confirmed by the chemical shifts of the methylene signal in¹H- and ¹³C-NMR experiments. The chemical shifts of the CH signal were in the same range as for the corresponding rhodanine derivatives 9. A particularly interesting compound with respect to the configuration of the double bond was the ortho-trifluoromethyl substituted derivative7e. The chemical shift in the ¹H-NMR ex-periment for the CH-signal was further downfield at 8.20 ppm, while all other derivatives were in the range of 7.75-8.07 ppm. The closest chemical shift was that of7b, with a chemical shift of 8.07 ppm. Both structures are related through substitution of the ortho-position in the 5-benzylidene moiety. Thus, indicating that the ortho-substituent is in proximity of the CH-group, deshielding it to appear further downfield compared to meta- and para-substituted analogues.

The three dimensional coordinates of 7e were prepared with the MOE2009.10 software package (Merck molecular force field 94x (MMFF94x)) and distances between the CH group and the trifluoromethyl group were calculated. The fluorine atoms of the trifluoromethyl group are only 0.2 Å further apart from the CH-group than the carbonyl group (Figure 20). This re-sults suggested that the trifluoro-methyl group was indeed responsible for the change of the chemical shift in the CH-signal in the¹H- and¹³C-NMR experiments. But even more remark-able was the observation of a⁵JH,F-coupling constant of 1.9 Hz, between the proton of the CH-group and the fluorines in the trifluoromethyl (CF3) group. The intramolecular distance between the CH-proton and the trifluoromethyl group was estimated to be 2.8 Å and therefore just out of the van der Waals-radii of the participating fluorine and hydrogen atoms (2.7 Å).^[168] However,

N S

Scheme 8: Synthesis of elongated N-3 sidechain linkers in rhodanine-N-acetic acid analogues 12 and 13; (i) NaOH_aq, rt; (ii) HCl, 100^◦C.

the distance measurement was only estimated, assuming a slightly shorter intramolecular dis-tance within the van der Waals-radii would allow the overlap of the 1s orbital of the proton and 2p orbital of the fluorine, generating a one-center molecular orbital (Figure 21).^[169–171]All atomic orbitals (the bonding and anti-bonding) will be filled, so that covalent bonds cannot be formed. These orbital interactions might explain the observed⁵J_H,F coupling of 1.9 Hz. How-ever, the exact mechanism of this long range coupling constant is unknown, but has been shown to be dependant on the intramolecular distance between both coupling partners.^[172]

Furthermore such coupling constants have previously been used to reveal intramolecular interactions, such as hydrogen-bonds of participating groups.^[173] For example fluorine-groups participating in hydrogen-bonding to an intermolecular amide (NH) showed⁵J_H,F coupling con-stants in the range of 1.7-2.0 Hz.^[173] It can be concluded that a value of 1.9 Hz for the⁵J_H,F coupling constant in7e was dependant on the through space distance and or intermolecular interaction of the hydrogen (CH) and fluorine group (CF₃), thus suggesting possible hydrogen-bonding interaction between the trifluoromethyl group and the CH moiety. Interestingly, the

13C-NMR experiment showed a similar long range⁴JC,F coupling constant of 1.9 Hz between the CH and the CF3group.