Synthesis of Novel Nitrogen Mustards Containing ...

Journal of Life Sciences and Technologies Vol. 3, No. 2, December 2015

Synthesis of Novel Nitrogen Mustards Containing Amino Alcohol Derivatives Aïcha. Amira, Malika. Berredjem, and Nour-Eddine. Aouf Laboratory of Applied Organic Chemistry (LAOC), Bioorganic Chemistry Group, Chemistry Department, Sciences Faculty, Badji-Mokhtar-Annaba University, Box 12, 23000 Annaba, Algeria Email: [email protected], {malika.berredjem, noureddine.aouf}@univ-annaba.dz

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Chlorethamine [2], Cyclophosphamide [3] and Chlorambucil [4] (Fig. 1) are the most potent and effective drugs react as bifunctional alkylating agents. N,N-bis(2-chloroethyl)amine pharmacophore, is believed to exert the antitumoral activity through interstrand cross-linking in the major groove of DNA and this linkage represents the highest toxicity of all alkylation events [5]-[7]. Development of resistance against the existing anticancer drugs keeps research window open in search of newer anticancer molecules. Here we report an attempt to achieve new potent antitumor agents by the synthesis, characterization of novel nitrogen mustards containing amino alcohols derivatives. Many bioactive natural products and medical molecules consist of amino alcohols structural motifs due to their importance in asymmetric synthesis [8], peptide and pharmaceutical chemistry [9], resolution of racemic mixtures [10], synthesis of insecticidal compounds [11], and others.

Abstract—Nitrogen mustards are an extremely active class of alkylating agents that have widespread clinical application in the treatment of various tumors. A Novel series of 1,1-bis(2-chloroethyl)-3-(2-hydroxy ethyl)urea 3(a-c) having different substituent on C-2 have been synthesized from a number of simple and chiral amino alcohols derivatives prepared by reduction of amino acids. Nitrogen mustard motif is introduced by acylation of bis (2chloroethyl) amine with ethyl chloroform ate in water. The aminolysis of ethyl ester with a variety of amino alcohols in solvent-free conditions is the key step to give the desired products. All structures of the compounds were analyzed by different spectroscopic methods. Index Terms—nitrogen mustard, acylation, amino alcohol, aminolysis

I.

INTRODUCTION

Cancer is major health hazard for the world surpassing heart diseases. Each year, tens of millions of people are diagnosed with cancer around the world, and more than half of the patients eventually die from it. Surgery, radiotherapy, and chemotherapy are the most common types of cancer treatment. The last category include several class based on their biochemical structure and mechanism of action. Alkylating agents are the oldest group of chemotherapeutics in use today, and nitrogen mustards are an important subtype of this class widely used in the treatment of a variety of cancer. [1]

Figure. 2. Synthesis of nitrogen mustards containing amino alcohols derivatives

The direct transformation of ester to amide is a potentially important step in this route, the amide bond is among the most common chemical function present in natural or synthetic organic molecules, is generally carried out under harsh conditions requiring high temperatures and extended reaction times [12]. An

Figure 1. Nitrogen mustards agents. Manuscript received March 1, 2015; revised August 25, 2015.

© 2015 Journal of Life Sciences and Technologies doi: 10.18178/jolst.3.2.65-68

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Hz, JBX = 5.2 Hz, Jgem =13.5 Hz, CH2-Ar), 3.12 (m, 1H, *CH), 3.38-3.64 (2dd, ABX system, H A-HB, JAX = 7.3 Hz, JBX=3.9 Hz, Jgem =10.7 Hz, CH2-OH), 7.25 (m, 5H, CHAr) ppm; 13C NMR (100 MHz, CDCl3): δ 38.3 (CH2-Ar), 57.3 (*CH), 63.9 (CH2-OH), 127.0, 128.5, 129.2 (CHAr), 135.9(CAr) ppm. MS (ESI): 100% (152.1, [M+H]+); Anal. Calcd for C9H13NO (151.21).

efficient method under solvent-free conditions to novel molecules from a variety of amino alcohols and ethyl ester of bis (2-chloroethyl)amine is reported (Fig. 2). II.

MATERIALS AND METHODS

Melting points were determined in open capillary tubes on an electrothermal apparatus. Infrared spectra were recorded on a Perkin–Elmer FT-600 spectrometer. Proton nuclear magnetic resonance was determined on JEOL EX-400 or Brucker AC 250 spectrometer using CDCl3 as a solvent and TMS as an internal standard. Chemical shifts are reported in δ units (ppm). All coupling constants (J) are reported in Hertz. Multiplicity is indicated as s (singlet), d (doublet), t (triplet), m (multiplet) and combination of these signals. Mass spectral analysis was performed using electrospray ionization mass spectrometry (ESI-MS) (Thermo Scientific, France). All reactions were monitored by TLC on silica Merck 60 F254 (Art. 5554) precoated aluminum plates visualized using UV light and were developed by spraying with ninhydrin solution.

B.

Representative Procedure for the Acylation of bis(2chloroethyl) Amine. To an aqueous solution of bis (2-chloroethyl) amine hydrochloride 1, are added 2 equivalents of K2CO3 and 1 equivalent of ClCO2Et at 0°C. The mixture is stirred at room temperature for 3h. The product was then extracted with ethyl acetate; the organic phase is dried over MgSO4, filtered and concentrated. Ethyl bis(2-chloroethyl)carbamate (2, C7H13Cl2NO2, oil, 98% yield), 1H NMR (400 MHz, CDCl3):δ 1.25 (t, 3H, J = 7.0 Hz, CH3), 3.62 (m, 8H, 4CH2), 4.1 (q, 2H, J = 7.3 Hz, CH2) ppm; 13C NMR (100 MHz, CDCl3): δ 14.6 (CH3), 41.9 (CH2), 42.1 (CH2), 50.7 (CH2-Cl), 51.1 (CH2Cl), 61.9 (CH2), 155.9 (C=O) ppm.

A.

Representative Procedure for the Synthesis of Compounds 2’(a–d). To a solution of amino acid (1 equiv) 1’(a-d), and NaBH4 (2.4 equiv) in anhydrous THF (80 mL) at 0°C was added drop wise a solution of I2 (1 equiv) dissolved in the same solvent. The reaction mixture is refluxed for 18h. A volume of methanol was added to the mixture cooled at 0°C until the solution becomes clear. After 30 min, the solvent was evaporated and the residue was dissolved in a solution of KOH (20%), the resulting mixture was stirred for 4 hours and was then extracted with ethyl acetate (3×75mL). The organic extracts were combined and dried over anhydrous sodium sulfate. The solvent was removed to give the compounds 2’(a-d). 2-Aminoethanol (2’a, C2H7NO, Oil, 97% yield), 1H NMR (250 MHz, CDCl3): δ 3.25 (t, 2H, J = 5.2 Hz, CH2NH), 3.65 (t, 2H, J = 5.1 Hz, CH2-OH) ppm; 13C NMR (62 MHz, CDCl3): δ 43.6 (CH2-NH), 62.5 (CH2-OH) ppm. (S)-2-Amino-3-methylbutan-1-ol (2’b, C5H13NO, Oil, 90% yield), 1H NMR (250 MHz, CDCl3): δ 0.92 (d, 6H, J = 8.5 Hz, 2CH3), 1.86 (m, 1H, CH), 3.20 (m, 1H, *CH), 3.36-3.62 (2dd, ABX system, H A-HB, JAX=7.5 Hz, JBX=5.2 Hz, Jgem=12.6 Hz, CH2) ppm; 13C NMR (62 MHz, CDCl3):δ 21.1 (CH3), 21.3 (CH3), 24.8 (CH), 50.2 (*CH), 63.4 (CH2) ppm. (S)-2-Amino-4-methylpentan-1-ol (2’c, C6H15NO, Oil, 90% yield), 1H NMR (400 MHz, CDCl3): δ 0.90 (2d, 6H, J= 8.2 Hz, 2CH3), 1.25 (t, 2H, CH2) 1.69 (m, 1H, CH), 3.00 (m, 1H, *CH), 3.30-3.62 (2dd, ABX system, HA-HB, JAX=7.2 Hz, JBX=4.5 Hz, Jgem=11.7 Hz, CH2) ppm; 13C NMR (100 MHz, CDCl3): δ 22,2 (CH3), 22.9 (CH3), 24.7 (CH), 41.2 (CH2), 55.0 (*CH), 65.2 (CH2-OH) ppm. MS (ESI): 100% (118.1, [M+H]+); Anal. Calcd for C6H15NO (117.19). (S)-2-Amino-3-phenylpropan-1-ol (2’d, C9H13NO, white powder, Mp 91°C, 95% yield), 1H NMR (400 MHz, CDCl3): δ 2.54-2.78 (2dd, ABX system, H A-HB, JAX = 8.5

© 2015 Journal of Life Sciences and Technologies

C.

Representative Procedure for the Aminolysis of Ethyl bis(2-chloroethyl) Carbamate. In a round flask equipped with a distillation system, are placed 1 equivalent of aminoalcool 2’(a-d), 1 equivalent of the ethyl bis(2-chloroethyl)carbamate 1 and 1 equivalent of K2CO3, the reaction mixture is heated to 135 °C under magnetic stirring. Upon complete distillation of ethanol formed during the reaction, the TLC analysis shows the complete disappearance of the starting material and the appearance of a new product. After cooling, the reaction medium is diluted in dichloromethane, the base is filtered and then the reaction crude is purified by chromatography on silica gel column with dichloromethane: methanol (90:10). 1,1-bis(2-chloroethyl)-3-(2-hydroxyethyl)urea (3a, C7H13Cl2NO2, Oil, 68% yield), 1H NMR (400 MHz, CDCl3) δ 2.25 (bs, 1H, OH), 2.76 (t, 2H, J = 4.9 Hz, NCH2), 2.80 (t, 2H, J = 6.0 Hz, N-CH2), 3.36 (t, 2H, J = 6.0 Hz, CH2-Cl), 3.59 (t, 2H, J = 7.8 Hz, CH2-NH), 3.63 (t, 2H, J = 4.9 Hz, CH2-Cl), 4.32 (t, 2H, J = 8.4 Hz, CH2OH) ppm; 13C NMR (100 MHz, CDCl3) δ 44.2 (CH2-Cl), 44.9 (CH2-Cl), 46.7 (CH2-N), 51.0 (CH2-N), 61.0 (CH2NH), 61.9 (CH2-OH), 159.2 (C=O) ppm. (S)-1,1-bis(2-chloroethyl)-3-(1-hydroxy3methylbutan-2-yl)urea (3b, C10H20Cl2N2O2, Oil, 72% yield), NMR 1H (400 MHz, CDCl3) δ 0.88 (2d, 6H, J = 7.4 Hz, 2CH3), 1.58 (m, 1H, CH), 2.62 (m, 1H, *CH), 2.81 (t, 2H, J=7.6 Hz N-CH2), 2.95 (t, 2H, J=7.6 Hz, NCH2), 3.39-3.50 (2dd, ABX system, HA-HB, JAX = 10.0 Hz, JBX = 4.6 Hz, Jgem = 13.2 Hz, CH2-OH), 3.61(t, 2H, J=8.0 Hz, CH2-Cl), 3.74 (t, 2H, J=8.0 Hz, CH2-Cl) ppm; NMR 13C (100 MHz, CDCl3) δ 21.9 (CH3), 22.1 (CH3), 25.0 (CH), 44.5 (CH2-Cl), 44.9 (CH2-Cl), 46.1 (*CH), 58.0 (N-CH2), 60.0 (N-CH2), 64.2 (CH2-OH), 159.6 (C=O) ppm. (S)-1,1-bis(2-chloroethyl)-3-(1-hydroxy4methylpenta -2-yl)urea (3c, C11H22Cl2N2O2, Oil, 75%

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yield), NMR 1H (400 MHz, CDCl3) δ 0.90 (d, 6H, J = 6.4 Hz, 2CH3), 1.22-1.32 (m, 2H, CH2-CH), 1.62 (m, 1H, CH), 2.71 (m, 1H, *CH), 2.78 (t, 2H, J=7.8 Hz, N-CH2), 2.91 (t, 2H, J=7.8 Hz, N-CH2), 3.39 (m, 2H, CH2-OH), 3.61(t, 2H, J=8.6 Hz CH2-Cl), 4.34 (t, 2H, J=8.6 Hz, CH2-Cl) ppm; NMR 13C (100 MHz, CDCl3) δ 22.6 (CH3), 23.1 (CH3), 25.0 (CH), 40.9 (CH2), 44.1 (CH2-Cl), 44.7 (CH2-Cl), 45.1 (*CH), 57.0 (N-CH2), 62.0 (N-CH2), 63.2 (CH2-OH), 159.1 (C=O) ppm; IR (KBr, ʋ) 786 (2 C-Cl), 1737 (C=O), 3275 (NH), 3403 (OH) cm-1. (S)-1,1-bis(2-chloroethyl)-3-(1-hydroxy-3-phenyl propan-2-yl)urea (3d, C14H20Cl2N2O2, Oil, 70% yield), 1 H NMR δ 2.55-2.65 (2dd, système ABX, HA-HB, JAX = 6.0 Hz, JBX = 7.5 Hz, Jgem = 12.9 Hz, CH2), 3.10 (m, 4H, 2 ×CH2-N), 3.29 (m, 4H, 2 ×CH2-Cl), 3.75 (m, 2H, CH2), 4.22 (m, 1H, *CH), 7.21 (m, 5H, CHAr) ppm; NMR 13C (100 MHz, CDCl3) δ 38.1 (CH2-Ar), 44.4 (2×CH2-Cl), 40.8 (*CH), 60.2 (N-CH2), 61.9 (N-CH2) 62.7 (CH2-OH), 126.5, 128.6, 129.1 (CHAr), 138.5 (CAr) 159.0 (C=O) ppm. III.

ACKNOWLEDGMENT This work was generously supported by the (Direction Générale de la Recherche Scientifique et du Développement Technologique, DGRS-DT) Algerian Ministry of Scientific Research. REFERENCES [1]

J. Hansson, R. Lewensohn, U. Ringborg, and B. Nilsson, “Formation and removal of DNA cross-links induced by melphalan and nitrogen mustard in relation to drug-induced cytotoxicity in human melanoma cells,” Cancer Res., vol. 47, pp. 2631-2637, May 1987. [2] L. S Goodman, M. M. Wintrobe, W. Dameshek, M. J. Goodman, A. Gilman, and M. Mclennan, “Nitrogen mustard therapyuse of methyl-bis(beta-chloroethyl) amine hydrochloride and tris(betachloroethyl)amine hydrochloride for hodgkin's disease, lymphosarcoma, leukemia and certain allied and miscellaneous disorders,” J. Am. Med. Assoc., vol. 132, pp. 126-132, September. 1946. [3] A. Emadi, R. J. Jones, and R. A. Brodsky, “Cyclophosphamide and cancer: Golden anniversary,” Nat. Rev. Clin. Oncol., vol. 6, pp. 638-47, September 2009. [4] P. Sienkiewicz, K. Bielawski, A. Bielawska, and J. Palka, “Amidine analogue of chlorambucil is a stronger inhibitor of protein and dna synthesis in breast cancer MCF-7 cells than is the parent drug,” J. Eur. J. Pharmacol., vol. 492, pp. 95-101, May 2004. [5] S. R. Rajski and R. M. Williams, “DNA cross-linking agents as antitumor drugs,” Chem Rev., vol. 98, pp. 2723–2795, December 1998. [6] W. A. Denny, “DNA minor groove alkylating agents,” Med. Chem, vol. 8, pp. 533-544, April 2001. [7] S. M. Rink and P. B. Hopkins, “Direct evidence for DNA intrastrand cross-linking by the nitrogen mustard mechlorethamine in synthetic oligonucleotides,” Bioorg. Med. Chem. Lett., vol. 5, pp. 2845–2850, December 1995. [8] G. M, Coppola and H. F. Schuster, “Asymmetric synthesis construction of chiral molecules using amino acids,” in Asymmetric Synthesis Construction of Chiral Molecules using Amino Acids, John Wdey& Sons, NewYork, 1987. [9] R. E. J, TenBrink, “A method for the preparation of stereochemically defined. psi. [CH2O] pseudodipeptides,” Org. Chem., vol. 52, pp. 418-422, August 1987. [10] T. Sawayama, M. Tnukamoto, T. Sasagawa, S. Naruto, J. Matsumoto, and H. Uno, “An improved optical resolution of 3acetylthio-2-methylpropionic acid by use of a new chiral amine, N isopropyl (phenylalaninol),” Chem. Pharm. Bull., vol. 37, pp. 1382-1383, 1989. [11] S. Wu, R. Takeya, M. Eto, and C. Tomizawa, “Insecticidal activity of optically active 1, 3, 2-Oxazaphospholidine 2-Sulfides and 1, 3, 2-Benzodioxaphosphorin 2-Sulfides,” J. Pestic. Sci., vol. 12, pp. 221-227, August 1987. [12] A. L. J. Beckwith, In the Chemistry of Amides: Synthesis of Amides, J. Zabicky, Ed., New York: Interscience, 1970, pp. 96.

RESULTS AND DISCUSSION

The synthetic route for the new nitrogen mustard derivatives is outlined in Fig. 2. N,N-bis(2chloroethyl)amine 1 was treated with ethyl chloroformate and K2CO3 in water to give the ethyl bis(2chloroethyl)carbamate 2 after 3h with a good yield. Sodium borohydride and iodine are used to generate βamino alcohols 2’(a-d) from their corresponding acids in excellent yields (90-97%). Synthesis of compounds 3(a-d) was accomplished by solvent-free ethyl ester aminolysis with amino alcohols having different derivatization on C-2 was achieved with K2CO3 at 135°C. The good yields were obtained in all cases. The structures of all the synthesized compounds were established on the basis of spectroscopic methods. For a compounds 2, the appearance of IR band at 1737cm-1 for C=O stretching vibration of ethoxycarbonyl group confirmed that the acylation has been occurred. In their 1H NMR spectra, it showed a triplet to 1.25 ppm and a quadruplet to 4.14 ppm indicating the presence of ethoxy group. The IR spectra of compounds 3(a-d) confirmed by appearance of OH band at 3403±10 cm-1 and NH band at 3275±10 cm-1, and 1H NMR and 13C spectra showed disappearance of the characteristic signal of ethoxy group. IV.

CONCLUSION

In conclusion, we have developed novel nitrogen mustards containing amino alcohol derivatives by selective monoamidation of ethyl ester with primary amino alcohols mediated by K2CO3 under solvent-free conditions. This simple procedure provides the corresponding amides in good yields. The study of the antitumor activity of novel compounds is currently in progress.

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Aïcha Amira was born in Annaba, town of Algeria on 1989. She graduated from Badji Mokhtar Annaba-university. She pursued her PhD in organic chemistry under the supervision of Prof Nour-Eddine Aouf researching greener methods for protecting groups of different functions, and synthetic methodology for preparation of heterocycles, also the development of new alkylating agents.

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