Supporting Information
Synthesis of some novel class of peptides from α-amino nitrones and their
potential biological activities
Bhaskar Chakraborty1*, Amalesh Samanta2, Govinda Prasad Luitel1, Neelam Rai1,
Debmalya Mitra2
1Organic
2Division
Chemistry Laboratory, Sikkim Government College, Gangtok, Sikkim-737102, India
of Microbiology, Department of Pharmaceutical Technology, Jadavpur University, Kolkata-700032, India
E-mail: bhaskargtk@yahoo.com
Experimental
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H NMR spectra were recorded with a Bruker DRX 300 spectrometer (300 MHz, FT NMR) using
TMS as internal standard. 13C NMR spectra were recorded on the same instrument at 75 MHz. The
coupling constants (J) are given in Hz. IR spectra were obtained with a Perkin-Elmer RX 1-881
machine as film or as KBr pellets for all the products. MS spectra were recorded with a Jeol SX102 (FAB) instrument. All the reactions were monitored by TLC using 0.25 mm silica gel plates
(Merck 60F254 UV indicator) while column chromatography was performed with silica gel
(E.Merck India) 60–200 mesh. All other reagents and solvents were purified after receiving from
commercial suppliers. N-Benzylhydroxylamine, N-Phenyl maleimide, N-Methyl maleimide,
starting materials, reagents used in the reactions were obtained commercially from Aldrich,
Lancaster, Fluka while CDMT, DCC were obtained from Sigma-Aldrich, Switzerland and were
used without purification, unless otherwise indicated. Characterization of the novel peptides have
been confirmed on the basis of spectral data. The microorganisms used in this experiment includes
15 bacterial strains of both Gram positive & Gram negative varieties. These were obtained from
Division of Microbiology, Dept of Pharmaceutical Technology, Jadavpur University, Kolkata.
Representative experimental procedure for peptide synthesis (Peptide 4, Table 1; entry 1):
A mixture of isoxazolidine (1 mmol), Fmoc-glycine (I equivalent) and CDMT (1 equivalent) were
added to DMF (10 ml). The mixture was at kept at RT with constant stirring for 180 min. After the
completion of reaction as monitored by TLC ((Rf = 0.48), the solvent (DMF) was evaporated and
the residue was extracted in to ETOAc, and the organic layer was washed with 10% HCl, 5%
Na2CO3, water and brine. The organic phase was dried over anhydrous Na2SO4 and evaporated to
afford crude product 4. Deprotection of Fmoc was accomplished with piperidine/DMF (2:8 ; 2x3
1
min, 1x4 min, and 1x5 min). Washings of Fmoc in deprotection was carried out with 10 ml of
solvent for each wash. The peptide was then subjected to silica gel column chromatography to
obtain the pure product (Table 1, entry 1) as white crystals (yield 88%, m.p. 94 0C).
Selected spectroscopic data of Peptides:
O
O
C6H5
N
N
CH3
H
O
H
N
C
O
H
C
ee = 96%
H
NH2
2-amino-N-((3S)-hexahydro-2-methyl-4,6-dioxo-5-phenyl-2H-pyrrolo[3,4d]isoxazol-3-yl)acetamide
Spectroscopic data for peptide 4 (Table 1, entry 1): Yield : 88%, White crystals, M.P: 94 0C; IR
(KBr): υmax 3380 (s, N-H), 3220 (br, -NH2), 3152 (aromatic C-H protons), 1736 (s, C=O), 1685 (s, NHC=O), 1660 (s, C=O), 1423 (m, isoxazolidine ring C-C), 786 (s, aromatic C-C) cm-1. 1H NMR
(300 MHz, CDCl3): δ 8.12 (d, J = 6.60 Hz, -NH), 7.64-7.12 (m, 5H, C6H5 protons), 4.80 (d, 1H, J =
6.10 Hz, C5H), 4.46 (dd, 1H, J = 3.26, 3.22 Hz, C3H), 3.63 (dd, 1H, J = 6.04, 6.06 Hz, C4H), 3.52
(s, 3H, -CH3), 2.48 (br, s, 2H, NH2), 2.10 (s, 2H, CH2 protons).
C NMR (75 MHz, CDCl3): δ
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173.55, 173.00 (carbonyl carbons), 165.30 (-CONH), 135.06, 134.70, 134.48, 133.60 (aromatic
carbons), 84.43 (C5), 75.70 (C3), 55.24 (C4), 26.12 (–CH3), 25.66 (CH2 carbon). EI-MS (m/z): 304
(M)+, 289, 231, 230, 216, 77, 73. Anal. Calcd. for C14H16O4N4: C, 55.24; H, 5.29; N, 18.42%.
Found: C, 55.15; H, 5.18; N, 18.30%.
O
O
H3C
N
N
C6H5
H
O
H
N
H
C
ee = 93 %
O
C
H
NH2
2-amino-N-((3R)-hexahydro-5-methyl-4,6-dioxo-2-phenyl-2H-pyrrolo[3,4-d]isoxazol3-yl)acetamide
2
Spectroscopic data for peptide 4 (Table 1, entry 2): Yield : 87%, Pale yellow crystals, M.P: 121
0
C ; IR (KBr): υmax 3376 (s, N-H), 3216 (br, -NH2), 3148 (aromatic C-H protons), 1730 (s, C=O),
1680 (s, -NHC=O), 1660 (s, C=O), 1420 (m, isoxazolidine ring C-C), 780 (s, aromatic C-C) cm-1.
1
H NMR (300 MHz, CDCl3): δ 8.06 (d, J = 6.44 Hz, -NH), 7.47-7.30 (m, 5H, C6H5 protons), 4.72
(d, 1H, J = 6.64 Hz, C5H), 4.35 (dd, 1H, J = 2.88, 2.84 Hz, C3H), 3.52 (dd, 1H, J = 6.12, 6.10 Hz,
C4H), 3.50 (s, 3H, -CH3), 2.44 (br, s, 2H, NH2), 2.24 (s, 2H, CH2 protons).
13
C NMR (75 MHz,
CDCl3): δ 174.22, 174.10 (carbonyl carbons), 167.40 (-CONH), 134.44, 134.13, 133.96, 133.75
(aromatic carbons), 80.52 (C5), 77.36 (C3), 56.50 (C4), 28.18 (–CH3), 25.40 (CH2 carbon). EI-MS
(m/z): 304 (M)+, 289, 231, 230, 216, 139, 77, 73. Anal. Calcd. for C14H16O4N4: C, 55.24; H, 5.29;
N, 18.42%. Found: C, 55.19; H, 5.21; N, 18.35%.
O
O
C6H5
N
N
CH3
H
O
H
N
C
ee = 91%
O
H
C
H3C
NH2
2-amino-N-((3S)-hexahydro-2-methyl-4,6-dioxo-5-phenyl-2H-pyrrolo[3,4d]isoxazol-3-yl)propanamide
Spectroscopic data for peptide 5 (Table 1, entry 4): Yield : 84%, Gray paste; IR (KBr): υmax 3368
(s, N-H), 3235 (br, -NH2), 3150 (aromatic C-H protons), 1730 (s, C=O), 1678 (s, -NHC=O), 1664
(s, C=O), 1420 (m, isoxazolidine ring C-C), 784 (s, aromatic C-C) cm-1. 1H NMR (300 MHz,
CDCl3): δ 8.06 (d, J = 6.08 Hz, -NH), 7.26-6.86 (m, 5H, C6H5 protons), 4.75 (d, 1H, J = 6.02 Hz,
C5H), 4.36 (dd, 1H, J = 3.90, 3.92 Hz, C3H), 3.54 (dd, 1H, J = 6.52, 6.56 Hz, C4H), 3.42 (s, 3H,
CH3), 2.30 (br, s, 2H, NH2), 1.82 (d, 3H, J = 1.68 Hz, -CHCH3), 1.64 (dd, 1H, J = 2.40, 2.48 Hz,
CH proton). 13C NMR (75 MHz, CDCl3): δ 173.38, 172.14 (carbonyl carbons), 166.86 (-CONH),
132.55, 132.48, 132.39, 132.15 (aromatic carbons), 82.80 (C5), 75.26 (C3), 57.66 (C4), 28.42,
28.24 (2xCH3 carbons), 23.27 (CHCH3 carbon). EI-MS (m/z): 318 (M)+, 303, 231, 216, 87, 77.
Anal. Calcd. for C15H18O4N4: C, 56.57; H, 5.69; N, 17.60%. Found: C, 56.52; H, 5.60; N, 17.55%.
3
C 6H 5
O
N
C 6H 5
H
H3COOC
H
N
H
C
O
ee = 83%
C
H
NH2
Spectroscopic data for peptide 4 (Table 1, entry 10): Yield : 78%, White paste ; IR (KBr): υmax
3362 (s, N-H), 3234 (br, -NH2), 3146 (aromatic C-H protons), 1752 (C=O of ester), 1724 (s, C=O),
1674 (s, -NHC=O), 1660 (s, C=O), 1426 (m, isoxazolidine ring C-C), 788 (s, aromatic C-C) cm-1.
1
H NMR (300 MHz, CDCl3): δ 8.16 (d, J = 5.22 Hz, -NH), 7.38-7.06 (m, 2x5H, C6H5 protons),
4.16 (d, 1H, J = 3.48 Hz, C3H), 3.48 (s, 3H, -COOCH3), 2.26 (br, s, 2H, NH2), 1.53 (s, 2H, CH2
protons). 13C NMR (75 MHz, CDCl3): δ 175.42 (carbonyl carbon), 170.72 (ester carbon), 165.90 (CONH), 137.33, 137.15, 137.10, 137.05, 132.54, 132.46, 132.40, 132.17 (aromatic carbons), 80.70
(C5), 77.44 (C3), 54.46 (C4), 25.40 (CH2 carbon), 23.27. EI-MS (m/z): 353 (M)+, 280, 276, 203, 77,
73. Anal. Calcd. for C19H19O4N3: C, 64.56; H, 5.41; N, 11.89%. Found: C, 64.43; H, 5.38; N,
11.72%.
O
O
H3CH2C
N
N
CH2C6H5
H
O
H
N
H
C
ee = 92%
O
C
H3C
NH2
2-amino-N-((3S)-2-benzyl-5-ethyl-hexahydro-4,6-dioxo-2H-pyrrolo[3,4d]isoxazol-3-yl)propanamide
Spectroscopic data for peptide 5 (Table 1, entry 6): Yield : 81%, Yellow solid, M.P: 84 0C ; IR
(KBr): υmax 3360 (s, N-H), 3230 (br, -NH2), 3148 (aromatic C-H protons), 1730 (s, C=O), 1724 (s,
C=O), 1672 (s, -NHC=O), 1660 (s, C=O), 1434 (m, isoxazolidine ring C-C), 1180 (C-C), 785 (s,
aromatic C-C) cm-1. 1H NMR (300 MHz, CDCl3): δ 8.10 (d, J = 5.22 Hz, -NH), 7.20-7.12 (m, 5H,
C6H5 protons), 4.80 (d, 1H, J = 6.32 Hz, C5H), 4.26 (dd, 1H, J = 2.70, 2.76 Hz, C3H), 3.34 (dd,
4
1H, J = 6.12, 6.16 Hz, C4H), 2.50 (t, J = 1.30 Hz, -CH2CH3), 2.30 (s, 2H, CH2C6H5), 2.22 (br, s,
2H, NH2), 2.14 (d, 3H, J = 2.02 Hz, CH3), 1.90 (dd, 1H, J = 2.12, 2.16 Hz, CH proton of
CHCH3NH2), 1.80 (dd, 2H, J = 1.82, 1.76 Hz, -NCH2CH3). 13C NMR (75 MHz, CDCl3): δ 172.74,
172.60 (carbonyl carbons),165.90 (-CONH), 134.50, 134.46, 134.30, 134.16 (aromatic carbons),
84.55 (C5), 76.40 (C3), 56.70(C4), 30.12, 30.04 (2xCH3), 25.43, 24.71 (CH2 carbons). EI-MS (m/z):
346 (M)+, 259, 255, 226, 168, 91, 87, 77. Anal. Calcd. for C17H22O4N4: C, 58.93; H, 6.39; N,
16.18%. Found: C, 58.80; H, 6.28; N, 16.12%.
References
1 (a). Sureshbabu, V.V.; Narendra, N.; Hemantha, H.P.; Chennakrisna, Reddy G. Protein Pept
Lett. 2010, 17, 499-503. (b). Jones, J. Amino acid and Peptide synthesis; Oxford University Press
(2nd ed)-USA, 2002. (c) Deshong, P.; Li, W.; Kennington, J.W.; Ammon, H.L. J.Org.Chem. 1991,
56, 1364-1369. (d). Lloyd-Williams, P.; Albericio, F.; Giralt, E. Chemical Approaches to the
Synthesis of Peptides and Proteins, CRC, Boca Raton, FL, USA, 1997.
Note : All the Spectra of the Peptides are available with author.
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