HETEROCYCLES

Short Paper | Regular issue | Vol. 89, No. 12, 2014, pp. 2745-2759
Received, 15th October, 2014, Accepted, 6th November, 2014, Published online, 14th November, 2014.
DOI: 10.3987/COM-14-13106

■ Reaction of 5-Methylene-hydantoins and Their Chemical Modification to Twin-Drug Type Symmeteical Molecules

Fumiko Fujisaki, Ryou Fujiwara, Miriko Okabe, Ayumi Naito, Enko Fukami, Hatsumi Aki, Nobuhiro Kashige, Fumio Miake, and Kunihiro Sumoto*

Faculty of Pharmaceutical Sciences, Fukuoka University, 8-19-1 Nanakuma, Jonan-ku, Fukuoka 814-0180, Japan

Abstract

We report chemical modifications of 5-methylenehydantoins (3a-e) with various amine nucleophiles to 5-substituted hydantoins and transformations of the obtained hydantoins to new symmetrical twin-drug type bivalent molecules 12 and 13. We also report 5-methylenehydantoin derivatives (3a) with some symmetrical diamines such as piperadine, ethylene-1,2-diamine, and 1,2-phenylenediamine. From trials using diamines such as ethylene-1,2-diamine and 1,2-phenylenediamine, hydantoin ring-opened products 10 and 11 were isolated predominantly.

INTRODUCTION
Considering the molecular geometry of bioactive compounds, small C2- or C3-symmetrical molecules frequently appear in various synthetic compounds and those molecules are usually constructed on symmetrical templates. 1-4 We have already reported the synthesis of a few interesting symmetrical candidates with significant biological activities.5-13
As one of our projects to investigate new antibacterial active compounds,5, 7, 8, 11, 14 we have already reported a few symmetrical molecules. In our previous paper,5 we reported that treatment of 5-methylene- hydantoins (3) without a solvent with primary or secondary aliphatic dialkylamines regioselectively gave predominantly 5-alkylaminomethylhydantoin derivatives (4) and that the reaction conditions with CH2Cl2 resulted in the formation of isomeric 5-alkylamino-5-methyl-hydantoins (5). Furthermore, among the reported hydantoin derivatives, we found that some twin-drug type symmetrical molecules containing two 5-substituted hydantoin nuclei in the molecules showed high levels of antibacterial activity against a Gram-positive strain (S. aureus). We previously reported that identical twin-drug type symmetrical molecule A5, 15 (Figure 1) shows a considerably high level of antibacterial activity, and we also confirmed by calorimetric experiments that this bivalent molecule had an interesting binding property to sulfated glucosaminoglycans such as heparan sulfate and dermatan sulfate.16

In this article, we report regioselective chemical modification of 5-methylene-hydantoins with various amines to new 5-substituted hydantoins and preparation of new symmetrical twin-drug type bivalent molecules from the obtained hydantoins in order to find new antibacterial active leads.

RESULT AND DISCUSSION
Reaction of 5-Methylene-3-arylhydantoins (3) and Modifications of Reaction Products to Symmetrical Bivalent Molecules.
We prepared 5-metylene-3-arylhydantoins (3a-e) by deamination of 5-dialkylaminomethylhydantoin derivatives (2) readily obtained by addition of β-aminoalanines (1) to arylisocyanates, in a manner similar to that reported previously.5, 14, 17, 18 Synthesis of the compounds (3a and 3b) have already been reported.5 Preparation of new derivatives (3c, 3d and 3e) and their physical and spectroscopic data are described in the Experimental section. The overall reaction for 5-methylene-3-arylhydantoins (3a-e) as starting　materials19, 20 is shown in Scheme 1.

In our previous paper,5 we reported that treatment of 5-methylene-hydantoins (3) without a solvent (neat) (Method 1) with primary or secondary aliphatic dialkylamines regioselectively gave predominantly 5-alkylaminomethylhydantoin derivatives (4). On the other hand, the reaction conditions with CH2Cl2 (Method 2) resulted in the formation of 5-alkylamino-5-methylhydantoins (5) in moderate to good yields. For example, reactions of 5-methylenehydantoins (3) and various benzylamines without a solvent resulted in the formation of the corresponding 5-alkylaminomethyl or 5-dialkylaminomethylhydantoin derivatives (4a-d) in moderate to good yields, as shown in Sheme 2. Reactions of 3a with primary or secondary aliphatic amines such as pyrrolidine or benzylamine in CH2Cl2 also gave predominantly 5-alkylamino-5-methylhydantoin (5a or 5b) in good yield.5 We considered that tautomeric isomerization (A⇔B), as shown in Scheme 2, is a crucial intermediate for these regioselective additions of amine nucleophiles.18 In fact, both reactions of pyrrolidine with N(1)-methyl derivatives of 5-methylene-hydantoins such as compounds 3d and 3e, which could not afford an imine B-type tautomeric isomer under the conditions of Method 1 or 2, resulted in the selective formation of 5-prrrolidinomethyl-hydantoin derivatives (6d and 6e) (Scheme 2). These results also reconfirm the importance of tautomeric isomerization for the orientation of nucleophilic addition of amines to the 5-methylene moiety.

As an extension of this regioselective transformation, our previous method with CH2Cl2 as a reaction solvent was slightly modified. When using a solid monoamine such as anisidine, the homogenized residue obtained after evaporation of the mixture of 5-methylene-hydantoin (3) and an amine in CH2Cl2 was warmed up to an appropriate temperature (70 °C) to complete the reaction (Method 3). The results obtained by this procedure were similar to those reported previously. Thus, when using a monoamine such as solid anisidine or N-methylbenzylamine in the above reaction conditions (Method 3), the isolated adducts were 5-amino-5-methylhydantoin derivatives (7 and 8) in 35% and 70% yields, respectively. In order to obtain symmetrical twin-drug type derivatives containing two 5-amino-5-methylhydantoin moieties, we further examined the reaction of 5-methylene derivative 3a with a symmetrical diamine such as piperazine in a molar ratio of 3a: piperazine = 2:1 under the conditions of Method 3 (see Experimental). However, this trial for obtaining twin-drug type 5-amino-5-methylhydantoin derivatives was unsuccessful and resulted in predominant formation of the structural isomer (9a). The targeted symmetrical compound (9b) was not isolated (Scheme 3).

In some other attempts using the above reaction conditions (Method 3), reactions of compound 3a with the diamines ethylene-1,2-diamine and 1,2-phenylenediamine afforded predominantly hydantoin ring-opened products (10) and (11) in 23% and 53% yields, respectively (Scheme 4). Formation of the 1,4-diazepane derivative 10 was probably initiated by nucleophilic attachment of an amine moiety to 5-exo ethylene carbon in compound 3a and internal ring closure of an intermediate (a) with cleavage of

the hydantoin N3-C4 bond to afford compound 10. In contrast, the reaction of 5-methylenehydantoin with 1,2-phenylenediamine was initiated by nucleophilic access of an amine group to the C5-hydantoin ring carbon to give a ring-opened imine (b) and then a similar internal ringclosure of the imine with cleavage of the hydantoin N3-C4 bond, resulting in the formation of compound 11 together with the formation of N-phenylurea (Scheme 4). We consider that formation of bivalent symmetrical compounds, such as compound 9b, containing two 5-amino-5-methylhydantoin moieties in the molecules may require more defined conditions for the preparation of such compounds.
Among the above regioselective chemical transformations of 5-methylenehydantoins with amine nucleophiles, we are particularly interested in the formation of secondary amine type 5-alkylaminomethylhydantoin derivatives and in the formation of N-acyl derivatives by acylation reaction. In order to confirm the validity of this conventional procedure for the preparation of new bivalent 5-acylaminomethylhydantoin derivatives, we examined various diacylation reactions of primary amine adducts (4a-b) with aryl diisocynates (or aryl diisothiocyanates) or with dicarboxylic acid dichloride. For targeted bivalent symmetrical molecules having two hydantoin rings in the molecule, we designed symmetrical twin-drug type molecules (a mixture of C2-symmetrical and Cs-symmetrical compounds)21 shown as a general structure (12 and 13) starting with benzylamine adducts (4a-b) (Scheme 5).

In this modification, we used mainly less flexible platform linkers such as phenyl, biphenyl and diphenyl- methane moieties for target bivalent symmetrical derivatives because we have already observed that flexible methylene-chain linkers [(-CH2)n-: n = 4~12] in previously reported examples (symmetrical molecule A analogs) were not preferred linker structures, resulting in no antibacterial active compounds at a concentration of less than 128 μg/mL. In order to compare the bioactivities of the twin- drug type compounds, we also prepared additional examples having flexible chain-linkers in the molecule (for example, see 13b).
The symmetrical structures of the synthesized compounds (12 and 13) were easily confirmed by NMR spectroscopic analysis. All of the compounds except for compound 13b showed magnetically equivalent spectroscopic signal patterns, indicating a symmetrical molecular feature in solution21 (see Experimental). The structures and yields of these new bivalent symmetrical hydantoin derivatives (12 and 13) are summarized in Table 1, and the protocol for the preparation for these new bivalent symmetrical derivatives and their spectroscopic data are shown in detail in the Experimental section.

All of the synthesized bivalent symmetrical hydantoin derivatives (see Table 1) are now under antibacterial evaluation with gram-positive (S. aureus) and gram-negative (E. coli) strains. The results of evaluation of antibacterial activities and details of a structure-activity relationship (SAR) study will be described separately. For compounds with high levels of antibacterial activity in this article, results of thermodynamic experiments will be presented in the following papers.

EXPERIMENTAL
Melting points are uncorrected. IR spectra were measured by a Shimadzu FT/IR-8100 spectrometer. The　1H- and 13C-NMR spectra were obtained by a JEOL JNM A-500 at 35 °C. Chemical shifts are expressed in δ ppm downfield from an internal tetramethylsilane (TMS) signal. The signal assignments were confirmed by 1H - 1H two-dimensional (2D) correlation spectroscopy (COSY), 1H -13C heteronuclear multiple quantum coherence (HMQC), and 1H -13C heteronuclear multiple-bond connectivity (HMBC) spectra. High FAB-MS spectra were obtained by a JEOL JMS-HX110 mass spectrometer. The following abbreviations in parentheses were used for pyrrolidine ring (Pyr), 1-naphthyl ring (Np), hydantoin ring (Hyd), decandiamide (DD) and 1,4-diazepane ring (DAP).
5-Methylene-3-(naphthalen-1-yl)imidazolidine-2,4-dione (3c)
This compound was prepared from 1-naphthyl isocyanate and methyl 2-amino-3-(pyrrolidin-1-yl)propanoate dihydrochloride (1a: R1 = H) by the method described previously.5 Yield was 73%, mp 194 °C (dec). IR (KBr) cm-1: 3265, 1789, 1728, 1778, 1662. FAB-MS (positive) m/z: 239 (M+H)+. 1H-NMR (DMSO-d6) δ: 5.02 and 5.31 (each 1H, d, J = 1.5 Hz, =CH2), 7.33-7.73 (5H, m, Np H), 8.03-8.12 (2H, m, Np H), 10.89 (1H, br s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 95.0 (=CH2), 122.3, 125.5, 126.5, 127.1, 127.2 (Np C), 128.0 (Np C-8a), 128.2, 129.4 (Np C), 129.7 (Np -4a), 133.6 (Np C-1), 135.3 (Hyd C-5), 153.2 (Hyd C-2), 162.6 (Hyd C-4). Anal. Calcd for C114H10N2O2 • 0.3 H2O: C, 69.01; H, 4.39; N, 11.50. Found. C, 69.23; H, 4.29; N, 11.66.
3-(4-Chlorophenyl)-1-methyl-5-methyleneimidazolidine-2,4-dione (3d)
This compound was prepared from 1a (R1 = Me) and 4-chlorophenyl isocyanate by the method described previously. 5, 18 Yield was 65%, mp 146-147 °C. IR (KBr) cm-1: 1783, 1732, 1664. FAB-MS (positive) m/z: 237 (M+H)+. HR-FAB-MS (positive) m/z: 237.0434 (Calcd for C11H9N2O2Cl: 237.0431). 1H-NMR (DMSO-d6) δ: 3.12 (3H, s, NMe), 5.09 and 5.35 (each 1H, d, J = 2.5 Hz, =CH2), 7.46 (2H, d, J = 8.8 Hz, Ar H), 7.56 (2H, d, J = 8.8 Hz, Ar H). 13C-NMR (DMSO-d6) δ: 26.5 (NMe), 94.6 (=CH2), 128.2, 128.8, (Ar C), 130.5 (Ar C-4), 132.4 (Ar C-1), 136.4 (Hyd C-5), 152.4 (Hyd C-2), 160.9 (Hyd C-4). Anal. Calcd for C11H9N2O2Cl: C, 55.83; H, 3.83; N, 11.84. Found. C, 55.71; H, 3.82; N, 11.91.
1-Methyl-5-methylene-3-(naphthalen-1-yl)imidazolidine-2,4-dione (3e)
This compound was prepared from 1b (R1 = Me) and 1-naphthyl isocyanate by the method described previously. 5, 18 Yield was 63%, mp 173-174 °C (dec). IR (KBr) cm-1: 1774, 1729, 1661. FAB-MS (positive) m/z: 253 (M+H)+. 1H-NMR (DMSO-d6) δ: 3.19 (3H, s, NMe), 5.15 and 5.40 (each 1H, d, J = 2.1~2.4 Hz, =CH2), 7.55-7.65 (4H, m, Np H), 7.71-7.73 (1H, m, Np H), 8.04-8.09 (2H, m, Np H). 13C-NMR (DMSO-d6) δ: 26.6 (NMe), 94.7 (=CH2), 122.5, 125.5, 126.5, 127.0, 127.1 (Np C), 128.0 (Np C-8a), 128.2, 129.5 (Np C), 129.7 (Np C-4a), 133.6 (Np C-1), 136.9 (Hyd C-5), 153.1 (Hyd C-2), 161.7 (Hyd C-4). Anal. Calcd for C15H12N2O2: C, 71.42; H, 4.79; N, 11.10. Found. C, 71.32; H, 4.93; N, 11.03.
Preparation of Compounds 4, and 6-11. [Methods 1-3 shown below were used for the Preparation of 4, and 6-11].
[Method 1] 5 A mixture of methylenehydantoin 3 and an appropriate molar (2~4 times molar) amount of an amine was allowed to stand at rt for several hours. The resulting reaction mixture was purified by washing with Et2O or by column chromatography.
[Method 2] 5 A solution of methylenehydantoin 3 and an appropriate molar (2~5 times molar) amount of an amine in CH2Cl2 was stirred for several hours. After evaporation of the solvent, the resulting material was purified by washing with Et2O or by centrifugal chromatography (silica gel).
[Method 3] A solution of methylenehydantoin 3 and an appropriate molar (2~5 times molar) amount of an amine in CH2Cl2 was concentrated under reduced pressure. The homogenized residue was warmed up at ca 70 °C in a water bath and kept for several hours. The obtained product was purified by washing with Et2O or by centrifugal chromatography (silica gel).
3-Phenyl-5-((((R)-1-phenylethyl)amino)methyl)imidazolidine-2,4-dione　(4c)
This compound was obtained by using method 1. Thus, a mixture of methylenehydantoin 3a (55 mg, 0.29 mmol) and R(+)-α-methylbenylamine (140 mg, 1.16 mmol) was kept at rt for 6 h. The resulting solid material was purified with centrifugal chromatography (silica gel) using AcOEt as a solvent to afford viscous product 4c (54 mg, 60%). The compound 4c was separated as a diastereomeric mixture (ca 2:3) as indicated by 1H and 13C-NMR data shown below. IR (KBr) cm-1: 1779, 1718. FAB-MS (positive) m/z: 310 (M+H)+, HR-FAB-MS (positive) m/z: 310.1557 (Calcd for C18H20N3O2; 310.1556). 1H-NMR (DMSO-d6) δ: 1.26 (3H, d, J = 6.5 Hz, NHCH(Me)Ph), 2.1-2.3 (1H, br, NHCH(Me)Ph), 2.68-2.80 (2H, m, Hyd-CH2-N=), 3.72-3.78 (1H, m, NHCH(Me)Ph), 4.19 (0.4H, dd, J = 6.0, 4.0 Hz, Hyd H-5), 4.26 (0.6H, dd, J = 6.0, 4.0 Hz, Hyd H-5), 7.20-7.49 (10H, m, Ar H), 8.32 (0.4H, br s, Hyd H-1), 8.36 (0.6H, br, Hyd H-1). 13C-NMR (DMSO-d6) δ: 24.0, 24.4 (Me), 47.7, 48.8 (Hyd-CH2-NH), 56.95, 56.98 (Hyd C-5 or NHCH(Me)Ph), 57.3, 58.5 (NHCH(Me)Ph or Hyd C-5), 126.2, 126.3, 126.4, 126.48, 126.51, 126.53, 127.51, 127.54, 128.1, 128.2, 128.57, 128.58 (Ar C),　132.2, 132.2 (Ar C-1 in Hyd-Ph), 145.5, 145.7 (Ar C-1 in NHCH(Me)Ph), 155.82, 155.83 (Hyd C-2), 172.4, 172.5 (Hyd C-4).
5-((Benzyl(methyl)amino)methyl)-3-phenylimidazolidine-2,4-dione (4d)
This compound was obtained from the reaction of 3a and N-methylbenzylamine in 68% yield by using method 1. Yield was 68%; mp 133-136 °C, IR (KBr) cm-1: 1772, 1710. FAB-MS (positive) m/z: 310 (M+H)+. 1H-NMR (DMSO-d6) δ: 2.18 (3H, s, NMe), 2.83 (1H, dd, J = 13.5, 3.0 Hz, Hyd-CHH-N=), 2.91 (1H, dd, J = 13.5, 6.0 Hz, Hyd-CHH-N=), 3.53 and 3.69 (each 1H, d, J = 13.5 Hz, Ph-CH2), 4.38-4.40 (1H, m, Hyd H-5), 7.23-7.49 (10H, m, Ar H), 8.53 (1H, s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 42.4 (NMe), 56.5 (Hyd-CH2-NH), 62.1 (NMe-CH2-Ph), 126.4, 126.8, 127.5, 128.0, 128.5, 128.6 (Ar C), 132.2 (Ar C-1 in Hyd-Ph), 138.8 (Ar C-1 in NMe-CH2-Ph), 155.8 (Hyd C-2), 172.4 (Hyd C-4). Anal. Calcd for C18H19N3O2: C, 69.88; H, 6.19; N, 13.58. Found. C, 69.75; H, 6.26; N, 13.62.
Reaction of N-Methyl-5-methylene Hydantoins 3d and 3e with Pyrrolidine by Method 1 and Method 2.
3-(4-Chlorophenyl)-1-methyl-5-(pyrrolidin-1-ylmethyl)imidazolidine-2,4-dione (6d)　
[Method 1] A mixture of methylenehydantoin (3d) (100 mg, 0.42 mmol) and pyrrolidine (40 mg, 0.55 mmol) was allowed to stand for 5 h at rt. After addition of Et2O to the reaction mixture, the precipitated material was filtered to give compound 6d (43 mg, 33%). The obtained product was identical to an authentic sample. 22
[Method 2] A solution of methylenehydantoin (3d) (100 mg, 0.42 mmol) and pyrrolidine (40 mg, 0.55 mmol) in CH2Cl2 was stirred for 5 h and kept for 18 h at rt. After concentration of the solvent, the residue was purified by centrifugal chromatography (silica gel) using AcOEt to give 3-(4-chlorophenyl)-5-methyl-5-pyrrolidinylhydantoin 6d (30 mg, 23%). The obtained product was identical to an authentic sample.22
1-Methyl-3-(naphthalen-1-yl)-5-(pyrrolidin-1-ylmethyl)imidazolidine-2,4-dione (6e)
[Method 1] A mixture of methylenehydantoin (3e) (30 mg, 0.12 mmol) and pyrrolidine (20 mg, 0.28 mmol) was allowed to stand for 1 h at rt. Et2O was added to the reaction mixture and then the precipitated material was filtered to give compound 6e (17 mg, 45%). A ratio of the two rotational isomers (major/minor) of 7:3 was indicated by the 1H-NMR spectrum in DMSO-d6 at 34.6 °C. Mp 143-145 °C. IR (KBr) cm-1: 1772, 1713. FAB-MS (positive) m/z: 324 (M+H)+. HR-FAB-MS (positive) m/z: 324.1717 (Calcd for C19H21N2O2: 324.1712). 1H-NMR (DMSO-d6) δ: 1.71-1.81 (4H, m, Pyr H-3, H-4), 2.54-2.73 (4H, m, Pyr H-2, H-5), 3.01 (2.1H, s, NMe), 3.03 (0.9H, s, NMe), 3.00-3.12 (1H, m, Pyr-CHH), 3.20-3.23 (1H, m, Pyr-CHH), 4.23-4.33 (0.7H, m, Hyd H-5), 4.47-4.49 (0.3H, m, Hyd H-5), 7.46-7.56 (7H, m, Np H). 13C-NMR (DMSO-d6) δ: 23.5 (Pip C-3, C-4), 27.7 (NCH3), 52.8 (Pyr C-2, C-5), 55.2 (CH2-Pip), 62.8 (Hyd C-5), 129.0 (Np C-8a), 130.0 (Np C-4a), 133.7 (Np C-1), 155.7 (Hyd C-2), 171.8 (Hyd C-4). These signals are ascribable to the major rotational isomer. δ: 23.45 (Pip C-3, C-4), 28.0 (NCH3), 53.8 (Pyr C-2, C-5), 54.4 (CH2-Pip), 61.4 (Hyd C-5), 129.0 (Np C-8a), 130.0 (Np C-4a), 133.7 (Np C-1), 155.6 (Hyd C-2), 171.9 (Hyd C-4). These signals are ascribable to the minor rotational isomer. δ: 122.7, 122.9, 125.4, 125.5, 125.5, 126.39, 126.42, 126.42, 126.7, 127.0, 128.1, 128.1, 129.1, 129.1 (Other Np C, These signals are for both rotational isomers). Anal. Calcd for C19H21N3O2 • 0.15 H2O: C, 69.98; H, 6.58; N, 12.96. Found. C, 69.98; H, 6.62; N, 12.91.
[Method 2] A mixture of methylenehydantoin (3e) (30 mg, 0.12 mmol) and pyrrolidine (30 mg, 0.42 mmol) in CH2Cl2 was stirred for 20 h at rt. After evaporation of the solvent, Et2O was added to the residue and then the precipitated material was filtered to give compound 6e (18 mg, 47%). The product obtained by this method contained a small amount of the starting 3e (by 1H-NMR analysis).
5-((4-Methoxyphenyl)amino)-5-methyl-3-phenylimidazolidine-2,4-dione (7)
This compound was prepared by using method 3. A mixture of methylenehydantoin (3a) (70 mg, 0.37 mmol) and p-anisidine (33 mg, 0.27 mmol) in CH2Cl2 (5 mL) was concentrated under reduced pressure. The resulting residue was warmed up to 70 °C in a water bath and kept of that temperature for 6 h. The reaction mixture was purified by centrifugal chromatography (silica gel) using AcOEt/n-hexane (1:1) as a solvent to give compound 7 in 35% yield (29 mg); a white solid; mp 138-140 °C. IR (KBr) cm-1: 1715. FAB-MS (positive) m/z: 312 (M+H)+. 1H-NMR (DMSO-d6) δ: 1.66 (3H, m, Hyd 5-Me), 3.27 (3H, s, OMe), 5.87 (1H, s, Ar-NH), 6.68 (2H, dd, J = 6.7, 2.4 Hz, Ar H-2, H-6 in anisidine), 6.78 (2H, dd, J = 6.7, 2.4 Hz, Ar H-3, H-5 in anisidine), 7.19-7.22 (2H, m, Ar H-2, H-6 in Hyd-Ph), 7.36-7.39 (1H, m, Ar H-4 in Hyd-Ph), 7.44-7.47 (2H, m, Ar H-3, H-5 in Hyd-Ph), 8.79 (1H, s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 25.3 (Hyd C(5)-Me), 55.1 (OMe), 72.2 (Hyd C-5), 114.3 (Ar C-2, C-6 in anisidine), 118.2 (Ar C-3, C-5 in anisidine), 126.5 (Ar C-2, C-6 in Hyd-Ph), 127.8 (Ar C-4 in Hyd-Ph), 128.7 (Ar C-3, C-5 in Hyd-Ph), 131.8 (Ar C-1 in Hyd-Ph), 138.1 (Ar C-1 in anisidine), 153.2 (Ar C-4 in anisidine), 153.7 (Hyd C-2), 173.7 (Hyd C-4). Anal. Calcd for C17H17N3O3: C, 65.58; H, 5.50; N, 13.50. Found: C, 65.38; H, 5.47; N, 13.51.
5-(Benzyl(methyl)amino)-5-methyl-3-phenylimidazolidine-2,4-dione (8)
This compound was prepared by using method 3. From the reaction of methylenehydantoin (3a) (50 mg, 0.27 mmol) and N-methylbenzylamine (70 mg, 0.58 mmol) in CH2Cl2 (5 mL), compound 8 was obtained in 70% yield (57 mg) as a white solid. Mp 132-135 °C. IR (KBr) cm-1: 1776, 1717. FAB-MS (positive) m/z: 310 (M+H)+. 1H-NMR (DMSO-d6) δ: 1.67 (3H, m, Hyd C(5)-Me), 2.15 (3H, s, NMe), 3.47 and 3.71 (each 1H, d, J = 13.5 Hz, N-CH2-Ph), 7.25-7.52 (10H, m, Ar H), 8.87 (1H, s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 22.5 (Hyd C(5)-Me), 32.1 (NMe), 54.2 (N-CH2-Ph), 77.0 (Hyd C-5), 126.7, 126.9, 127.8, 128.1, 128.4, 128.7 (Ar C), 131.8 (Ar C-1 in Hyd-Ph), 138.9 (Ar C-1 in N(Me)CH2-Ph), 154.2 (Hyd C-2), 173.2 (Hyd C-4). Anal. Calcd for C18H19N3O2 • 0.3 H2O: C, 68.68; H, 6.28; N, 13.35. Found: C, 68.88; H, 6.31; N, 13.12.
5,5’(Piperazine-1,4-diylbis(methylene)bis(3-phenylimidazolidine-2,4-dione) (9a)
This compound was prepared by using method 3. From the reaction of methylenehydantoin (3a) (101 mg, 0.54 mmol) and piperazine (23 mg, 0.27 mmol) in CH2Cl2 (5 mL), compound 9a was obtained in 34% yield (42 mg) as a white solid; mp 203 °C (dec). IR (KBr)cm-1:1776, 1721. FAB-MS (positive) m/z: 463 (M+H)+. 1H-NMR (DMSO-d6) δ: 2.42-2.43, 2.50-2.57 (each 4H, m, piperazine ring), 2.68-2.78 (4H, m, Hyd- CH2- N=), 4.30 (2H, s, Hyd H-5), 7.31-7.39 (6H, m, Ar H), 7.46-7.49 (4H, m, Ar H), 8.41 (2H, s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 53.7 (piperazine ring), 56.2, 56.3 (Hyd C-5), 58.19, 58.22 (Hyd- CH2-N=), 126.5, 127.6, 128.6 (Ar C), 132.2 (Ar C-1), 155.8 (Hyd C-2), 172.5 (Hyd C-4). Anal. Calcd for C24H26N6O4 • 0.5 H2O: C, 61.13; H, 5.77; N, 17.82. Found: C, 61.23; H, 5.64; N, 17.90.
Reaction of Methylene-Hydantoin 3a and Ethylene-1,2-diamine
By using method 3, from the reaction of methylenehydantoin (3a) (117 mg, 0.62 mmol) and ethylene-1,2-diamine (34 mg, 0.57 mmol) in CH2Cl2 (2 mL), we isolated 1-(5-oxo-1,4-diazepan-6-yl)-3-phenylurea 10 in 23% yield (33 mg) as a white solid; mp 160 °C (dec). IR (KBr) cm-1: 3338, 3220, 1665, 1636. FAB-MS (positive) m/z: 249 (M+H)+, HR-FAB-MS (positive) m/z: 249.1351 (Calcd for C12H17N4O2; 249.1352). 1H-NMR (DMSO-d6) δ: 2.42-2.46 (2H, m, DAP H-2 and H-7), 2.50-2.92 (1H, br, DAP H-1), 2.94-2.99 (1H, m, DAP H-2), 3.01-3.09 (2H, m, DAP H-3 and H-7), 3.26-3.31 (1H, m, DAP H-3), 4.38 (1H, d, J = 6.7 Hz, DAP H-6), 6.53 (1H, d, J = 5.5 Hz, phenylurea H-1), 6.89 (1H, d, J = 7.0 Hz, Ar H-4), 7.21 (2H, t, J = 7.6 Hz, Ar H-2 and H-6), 7.38 (2H, t, J = 7.6 Hz, Ar H-3 and H-5), 7.93 (1H, br s, DAP H-4), 8.95 (1H, br s, phenylurea H-3). 13C-NMR (DMSO-d6) δ: 43.8 (DAP C-3), 49.1 (DAP C-2), 50.8 (DAP C-7), 55.6 (DAP C-6), 117.4 (Ar C-2 and C-6), 120.9 (Ar C-4), 128.5 (Ar C-3 and C-5), 140.4 (Ar C-1) 154.1 (phenylurea C-2), 174.7 (DAP C-6). Anal. Calcd for C12H16N4O2 • 0.6 H2O: C, 55.63; H, 6.69; N, 21.62. Found: C, 55.91; H, 6.55; N, 21.32.
Reaction of Methylene-Hydantoin 3a and 1,2-Phenylenediamine
By using method 3, from the reaction of methylenehydantoin (3a) (117 mg, 0.62 mmol) and 1,2-phenylenediamine (29 mg, 0.27 mmol) in CH2Cl2 (2 mL), we obtained compound 11 (CAS 14003-34-0) in 53% yield (23 mg) as a white solid. N-Phenylurea was also isolated (35 mg). The obtained compound 11 showed mp 180 °C. The NMR data were identical to those of an authentic sample.23 Anal. Calcd for C9H8N2O: C, 67.49; H, 5.03; N, 17.49. Found: C, 67.42; H, 5.01; N, 17.48.
Preparation of Bivalent Twin-Drug Type Molecules (12a-12d).
1,1'-(1,4-Phenylene)bis(3-benzyl-3-((2,5-dioxo-1-phenylimidazolidin-4-yl)methyl)urea) (12a)
A solution of 1,4-diisocyanatebenzene (35 mg, 0.22 mmol) in CH2Cl2 was added to a solution of 5-((benzylamino)methyl)-3-phenylimidazolidine-2,4-dione　(4a) (100 mg, 0.34 mmol) in CH2Cl2, and the resulting mixture was stirred for 1 d at rt. After concentration of the solvent, the obtained solid material was purified by centrifugal chromatography (silica gel) using AcOEt as a solvent to afford a white solid 12a (66 mg, 52%). Mp 135-145 °C. IR (KBr) cm-1: 3412, 1779, 1718, 1644. FAB-MS (positive) m/z: 751 (M+H)+. 1H-NMR (DMSO-d6) δ: 3.73-3.74 (4H, m, Hyd-CH2-N=), 4.52 (2H, t, J = 6.0 Hz, Hyd H-5), 4.68, 4.74 (each 2H, d, J = 17.0 Hz, CH2-Ph), (2H, d, J = 17.0 Hz, CHH-Ph), 7.25-7.47 (24H, m, Ar H), 8.36 (2H, br s, NHCON=), 8.55 (2H, br s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 47.5 (Hyd-CH2-N=), 50.1 (CH2-Ph), 55.6 (Hyd C-5), 120.4 (Ar C in NH-Ph-NH), 126.5, 126.87, 126.91, 127.6, 128.4, 128.5 (Ar C), 132.1 (Ar C-1 in Hyd-Ph), 134.7 (Ar C-1 in NH-Ph-NH), 138.1 (Ar C-1 in CH2-Ph), 155.5 (NHCON=), 155.7 (Hyd C-2), 171.9 (Hyd C-4). Anal. Calcd for C42H38N8O6 • 0.5 H2O: C, 66.39; H, 5.17; N, 14.75. Found: C, 66.36; H, 5.43; N, 14.95.
1,1'-(1,4-Phenylene)bis(3-benzyl-3-((1-(4-chlorophenyl)-2,5-dioxoimidazolidin-4-yl)methyl)urea) (12b)
This compound was obtained from the reaction of 4b and 1,4-diisocyanatebenzene in 87% yield by a method similar to that for 12a as a white solid. Mp 236-237 °C. IR (KBr) cm-1: 1771, 1717, 1648. FAB-MS (positive) m/z: 819 (M+H)+. 1H-NMR (DMSO-d6) δ: 3.72 (4H, S, Hyd-CH2-N=), 4.51 (2H, t, J = 6.0 Hz, Hyd H-5), 4.68 and 4.74 (each 2H, d, J = 17.0 Hz, CH2-Ph), 7.26-7.28 (8H, m, Ar H-2, H-3, H5, H-6 in CH2-Ph), 7.32 (4H, s, Ar H in NH-Ph-NH), 7.35-7.36 (2H, m, Ar H-4 in CH2-Ph), 7.40, 7.50 (each 4H, d, J = 9.0 Hz, Ar H in p-Cl-Ph), 8.35 (2H, br s, NHCON=), 8.61 (2H, br s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 47.5 (Hyd-CH2-N=), 50.2 (CH2-Ph), 55.5 (Hyd C-5), 120.4 (Ar C in NH-Ph-NH), 126.8, 126.9, 128.1, 128.4, 128.5 (Ar C), 131.0 (Ar C-4 in p-Cl-Ph), 131.9 (Ar C-1 in p-Cl-Ph), 134.7 (Ar C-1 in NH-Ph-NH), 138.1 (Ar C-1 in CH2-Ph), 155.4 (NHCON=), 155.5 (Hyd C-2), 171.7 (Hyd C-4). Anal. Calcd for C42H36N8O6Cl2 • H2O: C, 60.22; H, 4.57; N, 13.38. Found: C, 60.24; H, 4.39; N, 13.45.
1,1'-(Methylenebis(4,1-phenylene))bis(3-benzyl-3-((2,5-dioxo-1-phenylimidazolidin-4-yl)methyl)-urea) (12c)
This compound was obtained from the reaction of 4a and bis(4-isocyanatophenyl)methane in 95% yield by a method similar to that for 12a as a white solid. Mp 128-145 °C. IR (KBr) cm-1: 1779, 1717. FAB-MS (positive) m/z: 841 (M+H)+. 1H-NMR (DMSO-d6) δ: 3.73-3.74 (4H, d, J = 6.0 Hz, Hyd-CH2-N=), 3.80 (2H, br s, Ph-CH2-Ph), 4.51 (2H, t, J = 6.0 Hz, Hyd H-5), 4.68 and 4.74 (each 2H, d, J = 17.0 Hz, CH2-Ph), 7.05-7.47 (28H, m, Ar H), 8.40 (2H, br s, NHCON=), 8.54 (2H, br s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 40.0 (Ph-CH2-Ph), 47.5 (Hyd-CH2-N=), 50.2 (CH2-Ph), 55.5 (Hyd C-5), 120.1, 126.5, 126.9, 126.9, 127.1, 128.3, 128.4, 128.5 (Ar C), 132.1, (Ar C-1 in Ph-Hyd), 135.2 (Ar C-4 in Ph-CH2-Ph) 138.0 (Ar C-1 in Ph-CH2-Ph), 138.1 (Ar C-1 in CH2-Ph), 155.4 (NHCON=), 155.7 (Hyd C-2), 171.9 (Hyd C-4). Anal. Calcd for C49H44N8O6 • 0.4H2O: C, 69.39; H, 5.32; N, 13.21. Found: C, 69.40; H, 5.45; N, 12.98.
1,1'-(Methylenebis(4,1-phenylene))bis(3-benzyl-3-((2,5-dioxo-1-phenylimidazolidin-4-yl)methyl)-thiourea) (12d)
A solution of bis(4-isothiocyanatophenyl)methane (42 mg, 0.15 mmol) in CH2Cl2 was added to a solution of　(4a) (100 mg, 0.34 mmol) in CH2Cl2, and the resulting mixture was stirred for 4 h at rt. After concentration of the solvent, the obtained solid material was washed with EtOH to give 12d (100 mg, 77%) as a white solid. Mp 132-136 °C. IR (KBr) cm-1: 1779, 1717. FAB-MS (positive) m/z: 873 (M+H)+. 1H-NMR (DMSO-d6) δ: 3.89 (2H, br s, Ph-CH2-Ph), 4.10-4.14 (2H, m, Hyd-CHH-N=), 4.26-4.30 (2H, m, Hyd-CHH-N=), 4.80 (2H, t, J = 6.5 Hz, Hyd H-5), 5.11 and 5.20 (each 2H, bd, J = 17.0 Hz, CH2-Ph), 7.14-7.47 (28H, m, Ar H), 8.59 (2H, s, Hyd H-1), 9.23 (2H, s, NHCSN=). 13C-NMR (DMSO-d6) δ: 40.0 (Ph-CH2-Ph), 52.0 (Hyd-CH2-N=), 53.9 (CH2-Ph), 54.8 (Hyd C-5), 126.4, 126.59, 126.62, 127.0, 127.6, 128.1, 128.4, 128.5 (Ar C), 132.1, (Ar C-1 in Ph-Hyd), 136.7 (Ar C-1 in CH2-Ph), 138.0 (Ar C-4 in Ph-CH2-Ph), 138.5 (Ar C-1 in Ph-CH2-Ph), 155.7 (Hyd C-2), 171.7 (Hyd C-4), 182.3 (C=S). Anal. Calcd for C49H44N8O4S2: C, 67.41; H, 5.08; N, 12.83. Found: C, 67.17; H, 5.16; N, 12.76.
N4,N4'-Dibenzyl-N4,N4'-bis((2,5-dioxo-1-phenylimidazolidin-4-yl)methyl)-[1,1'-biphenyl]-4,4'-dicarboxamide (13a)
A solution of [1,1'-biphenyl]-4,4'-dicarbonyl dichloride (42 mg, 0.15 mmol) in CH2Cl2 was added to a solution of compound 4a (100 mg, 0.34 mmol) and TEA (29 mg, 0.29 mmol) in CH2Cl2 (20 mL). The mixture was stirred for 1 h at rt and precipitated material was filtered off. The filtrate was concentrated under reduced pressure. A small amount of MeOH was added to the oily residue to give 13a (72 mg, 60%) as a white solid. Mp 132-138 °C. IR (KBr) cm-1: 1632. FAB-MS (positive) m/z: 797 (M+H)+. 1H-NMR (DMSO-d6) δ: 3.72-3.81 (4H, m, Hyd-CH2-N=), 4.66-4.73 (6H, m, CH2-Ph + Hyd H-5), 7.19-7.80 (28H, m, Ar H), 8.73 (2H, br s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 46.2 (Hyd-CH2-N=), 53.3 (Ph-CH2), 54.3 (Hyd C-5), 126.6, 126.7, 127.1, 127.4, 127.6, 128.2, 128.5, 128.7 (Ar C), 132.1, (Ar C-1 in Ph-Hyd), 135.4 (Ar C-4 in COPh-), 136.8 (Ar C-1 in Ph-CH2), 140.1 (Ar C-1 in COPh-), 155.8 (Hyd C-2), 171.5 (CON or Hyd C-4), 171.6 (Hyd-C-4 or CON). Anal. Calcd for C48H40N6O6 • 0.3H2O: C, 71.86; H, 5.10; N, 10.48. Found: C, 71.87; H, 5.18; N, 10.35.
N1,N10-Dibenzyl-N1,N10-bis((2,5-dioxo-1-phenylimidazolidin-4-yl)methyl)decanediamide (13b)
A solution of decanedioyl dichloride (17 mg, 0.07 mmol) in CH2Cl2 was added to a solution of compound 4a (50 mg, 0.17 mmol) and TEA (15 mg, 0.15 mmol) in CH2Cl2 (20 mL). The mixture was stirred for 1 h at rt and concentrated in vacuo. The residue was washed with water to give 13b (42 mg, 78%) as a white solid. Mp 86-90 °C. IR (KBr) cm-1: 1778, 1721. FAB-MS (positive) m/z: 757 (M+H)+. 1H-NMR (DMSO-d6) δ: 1.13-1.47 (8H, m, DD H-4, H-5), 1.49-1.54 (4H, m, DD H-3), 2.27-2.29, 2.46-2.50 (each 2H, m, DD H-2), 3.62-3.77 (4H, m, Hyd-CH2-N=), 4.48-4.74 (6H, m, CH2-Ph + Hyd H-5), 7.23-7.56 (20H, m, Ar H), 8.49 (1.3H, br s, Hyd H-1), 8.67(0.7H, br s, Hyd H-1). 13C-NMR (DMSO-d6) δ: 24.5, 24.8 (DD C-3), 28.5, 28.6, 28.6, 28.7 (DD C-4, C-5), 32.15, 32.22 (DD C-2), 47.4, 51.8 (CH2-Ph), 47.5, 47.5 (Hyd-CH2-N=), 54.7, 55.5 (Hyd C-5), 126.1, 126.49, 126.53, 126.8, 127.1, 127.2, 127.5, 127.7, 128.3, 128.5, 128.55, 128.61 (Ar C), 132.0, 132.1 (Ar C-1 in Hyd-Ph), 137.6, 137.7 (Ar C-1 in CH2-Ph), 155.6, 155.6 (Hyd C-2), 171.6, 171.7 (Hyd C-4), 172.8, 173.3 (CH2-CO-N). Anal. Calcd for C44H48N6O6 • 2.3H2O: C, 66.20; H, 6.64; N, 10.53. Found: C, 66.19; H, 6.68; N, 10.83.

References

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16. Regarding heat of binding between the twin-drug type compound A and sulfated glycosaminoglycans, we previously used dermatan sulfate (GAG-DS01) and heparan sulfate (GAG-HS01) as sulfated glycosaminoglycans.5 Shortly afterward, we obtained an additional result of exothermic binding reaction of compound A with scallop heparan sulfate (GM01/01). In isothermal titration experiments, the binding reaction with GM01/01 [Hepgm] was also exothermic and compound A showed thermodynamic parameters of K = 4.10 x 104 1/M and ΔH = -9.98 kJ/mol. Thermodynamic parameters for dermatan sulfate (GAG-DS01) [Der: K=1.11 x 103 1/M and ΔH = -10.9 kJ/mol] and heparan sulfate (GAG-HS01) [Hep: K = 2.75 x 104 1/M and ΔH = -9.46 kJ/mol] were shown in our previous paper.5 The calculated titration fitting curves of the above three kinds of sulfated glycosaminoglycans are shown together in the Figure.
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19. Recently, it has been reported that chemical modifications of 1,3-dibenzyl-5-methylene-hydantoin with a few reagents to 5-substituted hydantoin derivatives (see reference 20).
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21. All of the obtained compounds except for compound 13b exhibited very simple symmetrical 13C-NMR spectra in DMSO-d6, indicating little difference with respect to the signal assignable to two substituted hydantoin rings and a linker moiety. From a stereochemical viewpoint, obtained products 12-13 can be considered to be a mixture of three twin-drug type bivalent molecules, i.e., two C2-symmetrical molecules that have the same absolute configuration (R,R or S,S) regarding two chiral hydantoin rings in the molecules and a Cs-symmetrical meso compound having different absolute configurations (R,S). We previously found the presence of three stereoisomers in the free base of compound A by the HPLC method.15 In the case of compounds 13b, we consider that the diastereomeric mixture gave rise to slightly different non-equivalent magnetic resonance patterns. We used isomeric mixtures for biological prescreening (antibacterial activity) and for calorimetric experiments.
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