Synthesis and reactions of new chiral linear and macrocyclic tetraand penta-peptide candidates

Article abstract of DOI:10.5560/ZNB.2012-0116

A series of linear and macrocyclic pentapeptide derivatives have been prepared via the coupling of pyridine-2,6-dicarboxylic acid (1) or pyridine-2,6-dicarbonyl dichloride (2) with appropriate amino acid methyl esters. The coupling of 1 or 2 with aminoacid methyl esters gave the corresponding pyridine dipeptide methyl esters 3, which were hydrolyzed with sodium hydroxide to the corresponding acids 4. The latter compounds 4 were coupled with other amino acid methyl esters to afford the corresponding tetrapeptide esters 5, which were hydrolyzed with sodium hydroxide to the corresponding acids 6. Cyclization of tetrapeptide acids with L-lysine methyl ester or with aliphatic diamide derivatives afforded the corresponding cyclic pentapeptide methyl ester derivatives 7 and cyclic tetrapeptide diamines 8, respectively. Finally, hydrolysis with 1 N sodium hydroxide or hydrazinolysis with hydrazine hydrate of methyl esters 7 afforded the corresponding acids 9a - e and hydrazides 10a - e, respectively.

Full text of DOI:10.5560/ZNB.2012-0116

Synthesis and Reactions of New Chiral Linear and Macrocyclic Tetra-
and Penta-peptide Candidates
Mohamed H. Abo-Ghaliaa, Mohamed Abd El-Hamidb, Mohamed A. Zweila, Abd El-Galil
E. Amrc,d, and Shimaa A. Moafia
a
Peptide Chemistry Department, National Research Center, Cairo, Dokki, Egypt
Pharmaceutical Chemistry Departmenty, Faculty of Pharmacy, Ain-Shams University, Abasia,
b
Egypt
c
Drug Exploration & Development Chair (DEDC), College of Pharmacy, King Saud University,
Riyadh 11451, Saudi Arabia
Applied Organic Chemistry Department, National Research Center, Cairo, Dokki, Egypt
d
Reprint requests to Prof. Dr. Abd El-Galil E. Amr. Fax: +966-1-4676220.
E-mail: aamr1963@yahoo.com
Z. Naturforsch. 2012, 67b, 806 – 818 / DOI: 10.5560/ZNB.2012-0116
Received April 24, 2012
A series of linear and macrocyclic pentapeptide derivatives have been prepared via the coupling of
pyridine-2,6-dicarboxylic acid (1) or pyridine-2,6-dicarbonyl dichloride (2) with appropriate amino
acid methyl esters. The coupling of 1 or 2 with aminoacid methyl esters gave the corresponding pyri-
dine dipeptide methyl esters 3, which were hydrolyzed with sodium hydroxide to the corresponding
acids 4. The latter compounds 4 were coupled with other amino acid methyl esters to afford the cor-
responding tetrapeptide esters 5, which were hydrolyzed with sodium hydroxide to the corresponding
acids 6. Cyclization of tetrapeptide acids with L-lysine methyl ester or with aliphatic diamide deriva-
tives afforded the corresponding cyclic pentapeptide methyl ester derivatives 7 and cyclic tetrapeptide
diamines 8, respectively. Finally, hydrolysis with 1 N sodium hydroxide or hydrazinolysis with hy-
drazine hydrate of methyl esters 7 afforded the corresponding acids 9a – e and hydrazides 10a – e,
respectively.
Key words: Pyridine-2,6-dicarbonyl Dichloride, Amino Acids, Linear Piptides, Macrocyclic
Pentapeptides
Introduction
didates from dipicolinic acid with amino acids and
the screening of their biological activity [11 – 16].
Among the different areas of supramolecular and On the other hand, the synthesis of chemosensors
macrocyclic chemistry, the synthesis and complexing is an interesting approach providing accurate ana-
properties of azacrown compounds have been a sub- lytical tools in different analytical fields. In particu-
ject of intensive exploration [1 – 7]. Synthesis of chem- lar, 2,6-peptido-pyridines exhibited a general poten-
ical modifications of existing antibacterial agents in tial as ionophors [17] and were used for inventing
order to generate novel macromolecules with bet- novel thiocyanate-selective membrane sensors [18].
ter therapeutic properties is necessary because of the Recently, some new heterocyclic and peptide deriva-
emergence of multidrug-resistant bacteria [8]. Peptides tives have been studied with respect to their anti-
rarely function well as drugs due to their low bioavail- HIV [19], anti-inflammatory [20], anticoagulant [21],
ability and rapid degradation within cells [9]. The con- analgesic and anticonvulsant [22], anticancer [23],
version of these active peptides into peptidomimet- and antimicrobial activities [24 – 26]. In view of these
ics has been a successful approach for making new observations and as continuation of our previous
biologically active compounds [10]. In addition, we work [11 – 26] in macrocyclic and heterocyclic chem-
reported on the synthesis of some macrocyclic can- istry, we have synthesized some new macrocyclic
c
ꢀ 2012 Verlag der Zeitschrift fu¨r Naturforschung, Tu¨bingen · http://znaturforsch.com

M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
807
pentapeptides containing amino acid and pyridine respectively (Scheme 1). The IR spectra of 3a – c
moieties. confirmed the presence of an aromatic ring, aliphatic
The versatile biological activities of the synthesized hydrogens and an amide linkage in addition to the ester
diasteromeric compounds will be a subject of future re- group. The amide linkage was confirmed by its three
ports. For attributing the biological activity exclusively characteristic IR bands in the regions ν = 1680 – 1625,
to one active diaesteromer, a resolution of the mixtures 1548 – 1517 and 1316 – 1240 cm−1 (amide I, II and
into their individual components will then be manda- III, respectively). The presence of the ester group is
tory. Alternatively, synthesis and comparative studies supported by a band in the regions 1753 – 1740 cm−1
of all L- or all D-peptide candidates could equally be ν (C=O), ester). In addition, an absorption band was
envisioned.
observed at 3370 – 3335 cm−1, attributed to hydrogen
bonded amide ν (NH). Also, the IR spectra of 4a – c
showed the absence of ν (C=O, ester), and instead the
presence of a band at 1738 – 1726 cm−1 for ν (C=O,
Results and Discussion
The synthesis of Nα -dipicolinoyl-bis-(amino acid).
acid) methyl esters 3a – c was based on 2,6-
The synthesis of Nα -dipicolinoyl-bis[tetrapeptide
pyridinedicarbonyl dichloride (2), which was obtained methyl ester] derivatives 5a – e was based on the dipep-
by conversion of 2,6-pyridine-dicarboxylic acid (1) via tide acids 4a – c. Their treatment with amino acid
the reaction with thionyl chloride [27]. This acid chlo- methyl ester hydrochlorides in the presence of ethyl
ride was then coupled, at low temperature, with amino chloroformate in dichloromethane afforded the corre-
acid methyl esters in the presence of triethylamine sponding Nα -dipicolinoyl-bis[tetrapeptide methyl es-
as organic base. Bis-esters 3a – c were also prepared ter] derivatives 5a – e, which were hydrolyzed with
from 2,6-pyridinedicarboxylic acid (1) and amino acid methanolic sodium hydroxide to afford the corre-
esters in the presence ethyl chloroformate. Hydrolysis sponding Nα -dipicolinoyl-bis[tetrapeptide] derivatives
of the dipeptide methyl esters 3a – c with 1 N sodium 6a – e (Scheme 2). The 1H NMR spectra of 5 revealed
hydroxide in methanol afforded the corresponding the presence of a singlet (6H) at δ = 3.6 – 3.7 ppm for
Nα -dipicolinoyl-bis-amino acid derivatives 4a – c, the ester-CH3 protons. The IR spectra of 6 showed the
Scheme 1. Synthetic routes for compounds 3a–c and 4a–c.

808
M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
Scheme 2. Synthetic routes for compounds 5a–e and 6a–e.
absence of ν (C=O, ester), and the presence of a band Also, tetrapeptides 6a were cyclized with aliphatic
1
at 1738 – 1726 cm−1 for ν (C=O, acid). Also, the H diamines in the presence of ethyl chloroformate
NMR spectra revealed the disappearance of the singlet (mixed anhydride method) or in the presence of DCC
(6H) at δ = 3.6 – 3.7 ppm for ester-CH3 protons, and (Method B) to afford the corresponding cyclo-(Nα -
the appearance of a singlet (2H) at δ = 9.5 – 9.8 ppm dipicolinoyl)-bis[L-Leu-DL-Nva]-aliphatic
diamine
1
for carboxylic (OH) protons which are exchangeable 8a, b (Scheme 3). The IR and H NMR spectra of
with D2O.
7 supported the presence of the ester group by the
Cyclization of the tetrapeptides 6a – e with L- observation of a band in the region 1753 – 1740 cm−1
lysine methyl ester by different methods afforded ν (C=O) and the presence of a singlet (3H) at
the corresponding cyclic pentapeptide esters 7a – e. δ = 3.6 – 3.7 ppm for ester-CH3. Also, the IR spectra

M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
809
Scheme 3. Synthetic routes for compounds 7a–e and 8a, b.
of 8 showed the absence of ν (C=O, acid), and the responding acid derivatives 9a – e. Also, hydrazinol-
1H NMR spectra revealed the disappearance of the ysis of 7a – e with hydrazine hydrate in methanol af-
singlet (2H) at δ = 9.5 – 9.8 ppm for the carboxylic forded the corresponding cyclic pentapeptiedie hy-
protons.
drazide derivatives 10a – e (Scheme 4). The IR spec-
Finally, the methyl groups of the L-Lys-OMe es- tra of 9 showed the absence of ν (C=O, ester) and the
ters of the cyclic pentapeptides 7a – e were converted presence of a band at 1738 – 1726 cm−1 for ν (C=O,
to carboxylic acid groups or hydrazides. Hydrolysis of acid). The IR spectra of 10 showed the NH stretch-
the pentapeptide methyl ester derivatives 7a – e with ing vibrations of the amide and hydrazide groups as
1 N sodium hydroxide in methanol afforded the cor- a broad band centered at 3370 – 3335 cm−1
.

810
M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
Scheme 4. Synthetic routes for compounds 9a–e and 10a–e.
spectrometer, using the electron impact technique (EI). Ana-
lytical thin layer chromatography (TLC) was performed on
silica gel aluminum sheets, 60 F254 (E. Merck). The final
compounds were purified on manually prepared silica gel
glass plates using Fluka silica gel GF254, with 13% calcium
sulfate as a binder. Specific optical rotations were measured
with a A. Krawss, Optronic, P8000a polarimeter, in a 1 dm
length observation tube, at the indicated conditions, and ac-
cording to the equation: [α]TD = 100. α/(cl), where: α =
observed rotation angle, D = sodium line (λ = 589 nm),
c = concentration (g/100 mL), l = path length in dm and
T = temperature (◦C). The following solvent systems (by
volume) were used as eluents for the development of the
Experimental
Melting points were determined in open glass capillary
tubes with an “Electro Thermal” Digital melting point ap-
paratus, (model: IA9100) and are uncorrected. Elemental
micro-analysis for carbon, hydrogen and nitrogen (Microan-
alytical Unit, NRC) was found within the acceptable lim-
its of the calculated values. Infrared spectra (KBr) were
recorded on a Nexus 670 FTIR Nicolet, Fourier Transform
infrared spectrometer. Proton nuclear magnetic resonance
(1H NMR) spectra were run in [D6]DMSO on Jeol 270
MHz or 500 MHz instruments. Chemical shifts δ are given in
ppm. Mass spectra were run on a MAT Finnigan SSQ 7000

M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
811
plates: S: chloroform-methanol-acetic acid (85 : 10 : 5); S1: 1747 (C=O, ester), 1652, 1533, 1252 (C=O, amide I, II and
S-petroleum ether (b. p. 40 – 60 ◦C) (1 : 1); S2: S-petroleum III) cm−1. – 1H NMR (500 MHz, [D6]DMSO): δ = 8.75
ether (b. p. 40 – 60 ◦C) (3 : 2); S3: S-petroleum ether (b. p. (s, 2H, 2 NH, D2O exchangeable), 8.40 – 8.35 (d, 2H, 3,5-
40 – 60 ◦C) (1 : 2) and S4: butanol-water-acetic acid-pyridine pyridyl-H), 8.31 – 8.25 (t, 1H, 4-pyridyl-H), 4.60 – 4.55 (t,
(120 : 48 : 12 : 40).
2H, 2 CHNH), 3.75 (s, 6H, 2 OCH3), 1.90 – 1.80 (m,
It is generally known that basic reaction media enhance 4H, 2CH2CH2CH3), 1.40 – 1.30 (m, 4H, 2CH2CH2CH3),
racemization. However, under the reaction conditions em- 1.05 – 0.95 (m, 6H, 2 CH2CH2CH3). – MS (EI, 70 eV): m/z
ployed in this work, especially short reaction times and tem- (%) = 394 (14) [M+1]+, 334 (100), 274 (40), 177 (3), 147
peratures below 0 ◦C, only negligible racemization was ob- (2), 134 (8). – C19H27N3O6 (393): calcd. C 58.01, H 6.87, N
served.
10.68; found C 57.95, H 6.76, N 10.55.
3c: Yield, %: 53 [A], 80 [B]; m. p. 115 – 117 ◦C. –
[α]2D5 = −146.6 (c = 0.5, MeOH). – IR (KBr): ν = 3338
(NH str.), 3028 (CH-Ar), 2945 (CH-aliph.), 1735 (C=O, es-
Synthesis of Nα -dipicolinoyl-bis[amino acid methyl
esters] 3a – c
ter), 1647, 1532, 1275 (C=O, amide I, II and III) cm−1
.
Method A: acid chloride method
–
1H NMR (500 MHz, [D6]DMSO): δ = 8.57 (s, 2H, 2
NH, D2O exchangeable), 8.35 – 8.30 (d, 2H, 3,5-pyridyl-H),
8.25 – 8.20 (t, 1H, 4-pyridyl-H), 7.45 – 7.33 (m, 10H, 2 Ar-
H), 4.70 – 4.65 (t, 2H, 2 CHNH), 3.68 (s, 6H, 2 OCH3 ),
3.40 – 3.35 (dd, 4H, 2 CH2Ph). – MS (EI, 70 eV): m/z (%)
= 489 (7) [M]+, 430 (25), 398 (30), 327 (22), 162 (30), 91
(100). – C27H27N3O6 (489): calcd. C 66.26, H 5.52, N 8.59;
found C 66.05, H 5.33, N 8.14.
2,6-Pyridinedicarbonyl dichloride (2) [27] (0.02 g,
1 mmol) was added drop by drop to a cold (−15 ◦C) and
stirred dichloromethane solution (20 mL) of the correspond-
ing free amino acid methyl ester (2 mmol), obtained by the
addition of an equivalent amount of N-methylmorpholine
(0.3 mL) to the amino acid methyl ester hydrochloride in
stirred and cold (−15 ◦C) dichloromethane (20 mL). The
reaction mixture was stirred for additional 3 h at the same
temperature, then for 12 h at room temperature, washed with
water, 1 N sodium bicarbonate, 1 N potassium hydrogen
sulfate and water, and then dried over anhydrous sodium
sulfate. The solvent was evaporated under reduced pressure
to dryness, and the obtained residue was solidified by
trituration with n-hexane. The obtained solid was filtered off
and crystallized from ethanol to give the esters 3a [28] and
3b, c, respectively.
Synthesis of Nα -dipicolinoyl-bis[amino acids] 4a – c
To a stirred and cold methanolic solution (−5 ◦C, 20 mL)
of the corresponding dipeptide ester 3a – c (1 mmol), 1 N
sodium hydroxide (25 mL) was gradually added. The reac-
tion mixture was stirred for 2 h at the same temperature, then
for 3 h at room temperature. The solvent was concentrated
under reduced pressure, and the remaining aqueous solution
was cooled and acidified with 1 N hydrochloric acid to pH
∼ 3. The obtained solid was filtered off, washed with water,
dried and crystallized from ethanol-water to give the corre-
Method B: mixed anhydride method
Ethyl chloroformate (0.2 mL, 2 mmol) was added sponding dipeptides 4a [28] and 4b,c, respectively.
to
a
stirred and cold (−15 ◦C) solution of 2,6-
4b: Yield, 70%; m. p. 90 – 93 ◦C. – IR (KBr): ν = 3307
pyridinedicarboxylic acid (1) (0.17 g, 1 mmol) and (NH, str.), 3048 (CH-Ar), 2960 (CH-aliph.), 1720 (C=O,
N-methylmorpholine (0.2 mL, 2 mmol) in dichloromethane acid), 1658, 1542, 1270 (C=O, amide I, II and III) cm−1. –
(20 mL). The reaction mixture was stirred for additional 1H NMR (500 MHz, [D6]DMSO): δ = 9.36 (s, 2H, 2 OH,
10 min, then the free amino acid methyl ester (2 mmol), D2O exchangeable), 8.48 (s, 2H, 2 NH, D2O exchange-
dissolved in dichloromethane (20 mL, −15 ◦C) was added. able), 8.15 – 8.05 (m, 3H, pyridyl-H), 4.55 – 4.50 (m, 2H, 2
Stirring was maintained for 3 h at −15 ◦C, then for 12 h at CHNH), 1.90 – 1.80 (m, 4H, 2CH2CH2CH3), 1.40 – 1.30 (m,
room temperature. The reaction mixture was then washed 4H, 2CH2CH2CH3), 1.10 – 0.98 (m, 6H, 2 CH2CH2CH3). –
with water, 1 N sodium bicarbonate, 1 N potassium hydrogen MS (EI, 70 eV): m/z (%) = 362 (3) [M–3]+, 350 (4), 334
sulfate and water, and dried over anhydrous sodium sulfate. (100), 274 (42), 134 (7), 78 (6). – C17H23N3O6 (365): calcd.
The solvent was evaporated to dryness, and the obtained C 55.89, H 6.30, N 11.51; found C 55.34, H 6.05, N 11.22.
oily residue was solidified by trituration with n-hexane. The
4c: Yield: 87%; m. p. 120 – 122 ◦C. – [α]D25 = −89.6
obtained solid was collected by filtration and crystallized (c = 0.5, MeOH). – IR (KBr): ν = 3339 (NH, str.), 3030
from ethanol-n-hexane to give esters 3a [28] and 3b, c as (CH-Ar), 2929 (CH-aliph.), 1727 (C=O, acid), 1648, 1534,
identified by melting point and TLC in comparison with 1225 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz,
authentic samples prepared according to method A.
[D6]DMSO): δ = 9.42 (s, 2H, 2 OH, D2O exchangeable),
3b: Yield, %: 55 [A], 87 [B]; m. p. 101 – 102 ◦C. – IR 8.65 (s, 2H, 2 NH, D2O exchangeable), 8.25 – 8.15 (m, 3H,
(KBr): ν = 3300 (NH, str), 3033 (CH-Ar), 2960 (CH-aliph.), pyridyl-H), 7.25 – 7.10 (m, 10H, 2 Ar-H), 4.75 – 4.70 (t, 2H,

812
M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
2 CHNH), 3.30 – 3.25 (dd, 4H, 2 CH2Ph). – MS (EI, 70 eV):
5d: Yield: 80%; m. p. 92 – 94 ◦C. – [α]D25 = −140
m/z (%) = 460 (10) [M–1]+, 373 (48), 313 (30), 148 (62), (c = 0.5, MeOH). – IR (KBr): ν = 3302 (NH, str.),
105 (36), 91 (100). –C25H23N3O6 (461): calcd. C 65.08, H 3040 (CH-Ar), 2959 (CH-aliph.), 1744 (C=O, ester), 1658,
4.99, N 9.11; found C 64.91, H 4.86, N 9.03.
1531, 1208 (C=O, amide I, II and III) cm−1. – 1H
NMR (500 MHz, [D6]DMSO): δ = 8.65 (s,4H,4NH,D2O ex-
changeable), 8.40 – 8.30 (m, 3H, pyridyl-H), 7.40 – 7.30 (m,
10H, 2Ar-H), 4.30 – 4.20 (t, 2H, 2 CHNH), 4.05 – 3.95 (t,
2H, 2 CHNH), 3.61 (s, 6H, 2 OCH3), 3.20 – 3.10 (dd, 4H, 2
CH2Ph), 1.70 – 1.60 (m, 4H, 2CH2CH2CH3), 1.40 – 1.30 (m,
4H, 2 CH2CH2CH3), 1.10 – 0.95 (m, 6H, 2 CH2CH2CH3).
– MS (EI, 70 eV): m/z (%) = 687 (8) [M]+, 540 (33), 382
(100), 322 (38), 91 (48), 78 (30). – C37H45N5O8 (687):
calcd. C 64.63, H 6.55, N 10.19; found C 64.45, H 6.25, N
10.03.
Synthesis of Nα -dipicolinoyl-bis[dipeptide methyl ester]
derivatives 5a – e by the mixed anhydride method
Ethyl chloroformate (0.2 mL, 2 mmol) was added to
a stirred and cold (−15 ◦C) dichloromethane solution
(20 mL) of the corresponding Nα -dipicolinoyl-bis[amino
acid] 4a – c (1 mmol), containing N-methylmorpholine
(0.2 mL, 2 mmol). The reaction mixture was stirred for addi-
tional 10 min, then a cold dichloromethane solution (20 mL)
of the free amino acid methyl ester of L-Leu, DL-Nva, or L-
Phe (2 mmol), was added. Stirring was maintained for 3 h
at −15 ◦C, then for 12 h at room temperature. The reaction
mixture was washed with water, 1 N sodium bicarbonate, 1 N
potassium hydrogen sulfate and water, and dried over anhy-
drous sodium sulfate. The solvent was evaporated under re-
duced pressure to dryness, and the obtained oily residue was
solidified by trituration with a dry ether-n-hexane mixture.
The obtained solid was collected by filtration and crystal-
lized from ethanol-n-hexane to give the corresponding esters
5a [29] and 5b – e, respectively.
5e: Yield: 64%; m. p. 142 – 144 ◦C. – [α]D25 = −127.4
(c = 0.5, MeOH). – IR (KBr): ν = 3289 (NH, str.), 3028
(CH-Ar), 2946 (CH-aliph.), 1741 (C=O, ester), 1656, 1531,
1213 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz,
[D6]DMSO): δ = 8.85 and 8.65 (2s, 4H, 4 NH, D2O ex-
changeable), 8.40 – 8.35 (t, 1H, 4-pyridyl), 8.20 – 8.15 (d,
2H, 3,5-pyridyl), 7.45 – 7.35 (m, 20H, 4Ar-H), 4.80 – 4.70 (t,
2H, 2 CHNH), 4.60 – 4.50 (t, 2H, 2 CHNH), 3.65 (s, 6H, 2
OCH3), 3.10 – 2.95 (m, 8H, 4CH2Ph). – MS (EI, 70 eV): m/z
(%) = 783 (2) [M]+, 749 (6), 489 (50), 398 (38), 168 (26),
91 (100). – C45H45N5O8 (783): calcd. C 68.97, H 5.75, N
8.94; found C 68.15, H 5.42, N 8.65.
5b: Yield: 66%; m. p. 95 – 97 ◦C. – [α]D25 = −58.6
(c = 0.5, MeOH). – IR (KBr): ν = 3309 (NH, str.),
3028 (CH-Ar), 2957 (CH-aliph.), 1743 (C=O, ester),
1660,1530,1214 (C=O, amide I, II and III) cm−1. – 1H
NMR (500 MHz, [D6]DMSO): δ = 8.80, 8.52 (2s, 4H, 4
NH, D2O exchangeable), 8.25 – 8.20 (t, 1H, 4-pyridyl-H),
8.15 – 8.10 (d, 2H, 3,5 pyridyl-H), 7.20 – 7.10 (m, 10H,
2Ar-H), 4.50 – 4.40 (m, 4H, 4 CHNH), 3.64 (s, 6H, 2 of the corresponding tetrapeptide ester 5a – e (1 mmol),
OCH3), 2.80 – 2.70 (dd, 4H, 2 CH2Ph), 1.80 – 1.70 (m, 2H, sodium hydroxide (1 N, 25 mL) was gradually added. The
2CH2CH(CH3)2), 1.50 – 1.40 (m, 4H, 2CH2CH(CH3)2), reaction mixture was stirred for 2 h at the same temperature
0.90 – 0.80 (m, 12H, 2CH2CH(CH3)2). – MS (EI, 70 eV): then for 3 h at room temperature. The solvent was distilled
m/z (%) = 716 (2) [M+1]+, 508 (7), 376 (47), 302 (74), off under reduced pressure, and the remaining aqueous so-
162 (100), 78 (68). – C39H49N5O8 (715): calcd. C 65.45, H lution was cooled and acidified with 1 N hydrochloric acid
Synthesis of Nα -dipicolinoyl-bis[dipeptide]
derivatives 6a – e
To a stirred and cold methanolic solution (−5 ◦C, 20 mL)
6.85, N 9.79; found C 65.12, H 6.56, N 9.44.
to pH ∼ 3. The obtained solid was filtered off, washed with
5c: Yield: 78%; m. p. 99 – 100 ◦C. – [α]D25 = −18.6 water, dried and crystallized from ethanol–water to give the
(c = 0.5, MeOH). – IR (KBr): ν = 3297 (NH str.), 3032 (CH- corresponding tetrapeptides 6a [29] and 6b – e.
Ar), 2960 (CH-aliph.), 1746 (C=O, ester), 1653, 1539, 1208
6b: Yield: 73%; m. p. 124 – 126 ◦C. – [α]D25 = −66.2
(C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz, (c = 0.5, MeOH). – IR (KBr): ν = 3310 (NH, str.), 3035
[D6]DMSO): δ = 8.58 (s, 4H, 4 NH, D2O exchange- (CH-Ar), 2957 (CH-aliph.), 1720 (C=O, acid), 1655, 1528,
able), 8.40 – 8.30 (m, 3H, pyridyl-H), 4.60 – 4.55 (t, 2H, 1218 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz,
2 CHNH), 4.30 – 4.25 (t, 2H, 2 CHNH), 3.65 (s, 6H, 2 [D6]DMSO): δ = 10.53 (s, 2 H, 2 OH, D2O exchange-
OCH3), 1.90 – 1.80 (m, 2H, 2CH2CH(CH3)2), 1.70 – 1.60 able), 9.15 and 9.05 (2s, 4H, 4 NH, D2O exchangeable),
(m, 4H, 2CH2), 1.55 – 1.45 (m, 4H, 2 CH2), 1.30 – 1.20 (m, 8.35 – 8.25 (m, 3H, pyridyl-H), 7.20 – 7.10 (m, 10H, 2Ar-H),
4H, 2 CH2CH2CH3), 0.90 – 0.80 (m, 18H, 2 CH2CH2CH3, 4.50 – 4.45 (t, 2H, 2 CHNH), 4.40 – 4.35 (t, 2H, 2 CHNH),
2CH2CH(CH3)2). – MS (EI, 70 eV): m/z (%) = 619 (2) 3.10 – 3.05 (dd, 2H, CH2Ph), 2.90 – 2.80 (dd, 2H, CH2Ph),
[M]+, 447 (81), 348 (60), 302 (18), 274 (95), 158 (100). – 1.80 – 1.70 (m, 2H, 2 CH2CH(CH3)2), 1.50 – 1.40 (m, 4H, 2
C31H49N5O8 (619): calcd. C 60.09, H 7.91, N 11.31; found CH2CH(CH3)2), 0.90 – 0.80 (m, 12H, 2 CH2CH(CH3)2). –
C 59.88, H 7.14, N 11.02.
MS (EI, 70 eV): m/z (%) = 687 (5) [M]+, 634 (2), 489 (70),

M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
813
327 (48), 162 (36), 91 (100). – C37H45N5O8 (687): calcd. C 2 mmol). The reaction mixture was stirred for additional
64.62, H 6.55, N 10.19; found C 64.15, H 6.38, N 10.03.
20 min, then a cold (−15 ◦C) dichloromethane solution
6c: Yield: 75%; m. p. 118 – 120 ◦C. – [α]D25 = −20.8 (20 mL) of the free L-lysine methyl ester (1 mmol) was
(c = 0.5, MeOH). – IR (KBr): ν = 3315 (NH, str.), 3048 added. Stirring was maintained for 3 h at −15 ◦C, then for
(CH-Ar), 2961 (CH-aliph.), 1722 (C=O, acid), 1653, 1531, 12 h at room temperature. The reaction mixture was washed
1236 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz, with water, 1 N sodium bicarbonate, 1 N potassium hydrogen
[D6]DMSO): δ = 9.53 (s, 2H, 2 OH, D2O exchange- sulfate and water, and then dried over anhydrous sodium sul-
able), 8.75 and 8.62 (2s, 4H, 4 NH, D2O exchangeable), fate. The solvent was evaporated under reduced pressure to
8.40 – 8.30 (m, 3H, pyridyl-H), 4.70 – 4.40 (m, 4H, 4CHNH), dryness, and the obtained oily residue was solidified by trit-
1.90 – 1.80 (m, 2H, 2 CH2CH(CH3)2), 1.70 – 1.60 (m, 4H, uration with dry ether-n-hexane mixture. The crude product
2CH2), 1.55 – 1.50 (m, 4H, 2 CH2), 1.40 – 1.30 (m, 4H, 2 was purified by preparative thin layer chromatography using
CH2CH2CH3), 0.90 – 0.80 (m, 18H, 2 CH2CH2CH3 and 2 S3 as eluent to give the corresponding cyclic pentapeptide
CH2CH(CH3)2). – MS (EI, 70 eV): m/z (%) = 588 (2) [M]+, methyl esters 7a [29] and 7b – e.
447 (8), 348 (14), 274 (8), 158 (100), 78 (12). – C29H45N5O8
(591): calcd. C 58.88, H 7.61, N 11.84; found C 58.49, H
7.53, N 11.45.
Method B: DCC method
A cold (−5 ◦C) tetrahydrofuran solution (20 mL) of the
free L-lysine methyl ester (1 mmol) was added to a stirred dry
tetrahydrofuran solution (−5 ◦C, 20 mL) of the correspond-
ing Nα -dipicolinoyl-bis[dipeptide] 6a – e (1 mmol). Dicyclo-
hexylcarbodiimide (0.42 g, 2 mmol) was then added in por-
tions to the reaction mixture over 20 min at the same tem-
perature. Stirring was maintained for 20 h at room tempera-
ture. The reaction mixture was then diluted with acetonitrile
(20 mL), and the formed dicyclohexylurea was filtered off
and washed with acetonitrile (2 × 10 mL). The filtrate was
kept in the refrigerator overnight, and the newly formed dicy-
clohexylurea was then filtered off. Tetrahydrofuran was evap-
orated to dryness, and the obtained residue was dissolved in
dichloromethane, washed with 1 N sodium bicarbonate, 1 N
potassium hydrogen sulfate and water, and then dried over
anhydrous sodium sulfate. The solvent was evaporated to
dryness, and the obtained oily residue was solidified by tritu-
ration with dry ether-n-hexane mixture. In case of 7c and 7d,
the obtained solid was collected by filtration and crystallized
from ethanol-n-hexane, while for 7a, b and e, the crude prod-
ucts were purified by preparative thin layer chromatography
(S3). The cyclic pentapeptide methyl esters 7a – e were iden-
tified by melting point and TLC in comparison with authentic
samples prepared according to method A.
6d: Yield: 76%; m. p. 113 – 115 ◦C. – [α]D25 = −64.6
(c = 0.5, MeOH). – IR (KBr): ν = 3317 (NH, str.), 3043
(CH-Ar), 2961 (CH-aliph.), 1727 (C=O, acid), 1656, 1530,
1232 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz,
[D6]DMSO): δ = 9.55 (s, 2H, 2 OH, D2O exchangeable),
8.85 and 8.80 (2s, 4H, 4 NH, D2O exchangeable), 8.20 – 8.10
(m, 3H, pyridyl-H), 7.40 – 7.30 (m, 10H, 2 Ar-H), 4.75 – 4.70
(t, 2H, 2 CHNH), 4.65 – 4.60 (t, 2H, 2 CHNH), 4.20 – 4.10
(dd, 4H, 2 CH2Ph), 1.80 – 1.70 (m, 4H, 2 CH2CH2CH3),
1.40 – 1.30 (m, 4H, 2 CH2CH2CH3), 1.10 – 0.95 (m, 6H,
2 CH2CH2CH3). – MS (EI, 70 eV): m/z (%) = 659 (4)
[M]+, 582 (52), 408 (36), 241 (60), 131 (52), 56 (100). –
C35H41N5O8 (659): calcd. C 63.73, H 6.22, N 10.62; found
C 63.52, H 6.13, N 10.39.
6e: Yield: 65%; m. p. 116 – 118 ◦C. – [α]D25 = −108.4
(c = 0.5, MeOH). – IR (KBr): ν = 3309 (NH, str.), 3028
(CH-Ar), 2948 (CH-aliph.), 1725 (C=O, acid), 1655, 1528,
1219 (C=O, amide I, II and III) cm−1. – 1H NMR
(500 MHz, [D6]DMSO): δ = 9.85 (s, 2H, 2 OH, D2O ex-
changeable), 8.85 and 8.80 (2s, 4H, 4 NH, D2O exchange-
able), 8.35 – 8.30 (d, 2H, 3,5-pyridyl-H), 8.05 – 8.00 (t, 1H,4-
pyridyl-H), 7.50 – 7.30 (m, 20H, 4 Ar-H), 4.75 – 4.70 (t, 2H,
2 CHNH), 4.45 – 4.40 (t, 2H, 2 CHNH), 3.10 – 2.85 (m, 8H,
4 CH2Ph). – MS (EI, 70 eV): m/z (%) = 752 (3) [M–3]+,
681 (2), 549 (4), 284 (8), 168 (7), 91 (100). – C43H41N5O8
(755): calcd. C 68.34, H 5.43, N 9.27; found C 68.05, H 5.28,
N 9.17.
Method C: active ester method
To a stirred cold (−5 ◦C) dry tetrahydrofuran solu-
tion (20 mL) of Nα -dipi-colinoyl-bis-[L-Leu-DL-Nva] (6a)
(0.59 g, 1 mmol) containing N-hydroxysuccinimide (0.24 g,
2 mmol), dicyclohexylcarbodiimide (0.42 g, 2 mmol) was
added to the reaction mixture in portions over 20 min at the
same temperature. Free L-lysine methyl ester (1 mmol) was
Synthesis of
cyclo-(Nα -dipicolinoyl)-bis-[dipeptide]-L-Lys-OMe
(cyclic pentapeptide methyl esters) 7a – e
Method A: mixed anhydride method
Ethyl chloroformate (0.2 mL, 2 mmol) was added to then added. Stirring was maintained for 20 h at room temper-
a stirred and cold (−15 ◦C) dichloromethane solution ature. The reaction mixture was then diluted with acetoni-
(20 mL) of the corresponding Nα -dipicolinoyl-bis[dipeptide] trile (20 mL), and the formed dicyclohexylurea was filtered
6a – e (1 mmol), containing N-methylmorpholine (0.2 mL, off and washed with acetonitrile (2×10 mL). The filtrate was

814
M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
kept in the refrigerator overnight, and the newly formed di- 2CH2CH(CH3)2), 1.70 – 1.60 (m, 4H, 2 CH2), 1.50 – 1.40
cyclohexulurea was filtered off. Tetrahydrofuran was evapo- (m, 8H, 4 CH2), 1.30 – 1.20 (m, 6H, 3 CH2), 1.05 – 0.90
rated to dryness, and the obtained residue was dissolved in (m, 18H, 2 CH2CH2CH3 and 2CH2CH(CH3)2). – MS (EI,
dichloromethane, washed with 1 N sodium bicarbonate, 1 N 70 eV): m/z (%) = 716 (5) [M+1]+, 571 (11), 475 (60),
potassium hydrogen sulfate and water, and then dried over 302 (20), 156 (25), 91 (100). – C36H57N7O8 (715): calcd.
anhydrous sodium sulfate. The solvent was evaporated to C 60.42, H 7.97, N 13.71; found C 60.15, H 7.69, N 13.28.
7b: Yield 18% [A], 55 [B]; m. p. 108 – 110 ◦C. –
dryness, and the obtained oily residue was solidified by trit-
[α]2D5 = −52 (c = 0.5, MeOH). – IR (KBr): ν = 3306 (NH,
uration with dry ether-n-hexane mixture. The crude product
was purified by preparative thin layer chromatography (S3)
to give the corresponding cyclic pentapeptide methyl ester 7a
as identified by melting point and TLC in comparison with
an authentic sample prepared according to method A.
str.), 3045 (CH-Ar), 2955 (CH-aliph.), 1743 (C=O, ester),
1656, 1525, 1220 (C=O, amide I, II and III) cm−1. – 1H
NMR (500 MHz, [D6]DMSO): δ = 8.60 (s, 6H, 6 NH, D2O
exchangeable), 8.30 – 8.20 (m, 3H, pyridyl-H), 7.15 – 6.95
(m, 10H, 2Ar-H), 4.80 – 4.70 (m, 4H, 4 CHNH), 4.10 – 4.05
(t, 1H, NHCH2CH2CH2-CH2CHNH), 3.65 (s, 3H,
OCH3), 3.40 – 3.35 (m, 2H, NHCH2CH2CH2CH2CHNH),
3.10 – 3.00 (dd, 4H, 2CH2Ph), 1.90 – 1.80 (m, 2H,
2CH2CH(CH3)2), 1.70 – 1.60 (m, 4H, 2 CH2), 1.45 – 1.40
(m, 4H, 2 CH2), 1.30 – 1.20 (m, 2H, CH2), 0.95 – 0.85 (m,
12H, 2CH2CH(CH3)2). – MS (EI, 70 eV): m/z (%) = 811
(2) [M]+, 554 (3), 509 (35), 302 (92), 186 (64), 91 (100). –
C44H57N7O8 (811): calcd. C 65.10, H 7.02, N 12.08; found
C 64.95, H 6.89, N 11.93.
Method D: azide method
To
a
stirred methanolic solution (20 mL) of Nα -
(5a) (1.24 g,
dipicolinoyl-bis[L-Leu-DL-Nva-OMe]
2 mmol), anhydrous hydrazine hydrate (0.7 mL, 20 mmol)
was added. The reaction mixture was refluxed for 3 h, after
which the solvent was evaporated. The obtained residue
was triturated with ether, filtered off and crystallized from
methanol-ether to afford the corresponding dihydrazide
derivative (yield 80%, m. p. 130 – 132 ◦C). A cold mixture
(−15 ◦C) of the dihydrazide derivative (0.62 g, 1 mmol) in
hydrochloric acid (6 N, 2 mL) and glacial acetic acid (1 mL)
was stirred for 10 min, then an aqueous solution of sodium
nitrite (5 M, 2 mL) was added. Stirring was maintained for
30 min at the same temperature, after which the reaction
mixture was extracted with ether (60 mL), washed with cold
water, 5% sodium bicarbonate and water, and then dried over
anhydrous sodium sulfate. The cold ethereal azide solution
(−15 ◦C) was added to free L-lysine methyl ester (1 mmol).
Stirring was maintained for 5 h at the same temperature,
then for 20 h at room temperature. The reaction mixture
was washed with water, 5% potassium hydrogen sulfate and
water, and then dried over anhydrous sodium sulfate. Ether
was evaporated to dryness, and the obtained oily residue
was solidified by trituration with a dry ether-n−hexane
mixture. The crude product was purified by preparative thin
layer chromatography (S3) to give the corresponding cyclic
7c: Yield, 15% [A], 64 [B]; m. p. 95 – 98 ◦C.
–
[α]2D5 = −46 (c = 0.5, MeOH). – IR (KBr): ν = 3315
(NH, str.), 3035 (CH-Ar), 2958 (CH-aliph.), 1742 (C=O,
ester), 1655, 1533, 1236 (C=O, amide I, II and III) cm−1
.
–
1H NMR (500 MHz, [D6]DMSO): δ = 8.85 and 8.00
(2s, 6H, 6 NH, D2O exchangeable), 8.30 – 8.20 (m, 3H,
pyridyl-H), 4.50 – 4.40 (m, 4H,4CHNH), 4.15 – 4.10 (t,
1H, NHCH2CH2CH2CH2CHNH), 3.65 (s, 3H, OCH3),
3.30 – 3.25
(m,
2H,
NHCH2CH2CH2-CH2CHNH),
2.00 – 1.90 (m, 2H, 2CH2CH(CH3)2), 1.70 – 1.60 (m,
4H, 2CH2), 1.50 – 1.40 (m, 8H, 4 CH2), 1.30 – 1.20 (m,
6H, 3 CH2), 1.05 – 0.90 (m, 18H, 2 CH2CH2CH3 and
2CH2CH(CH3)2). – MS (EI, 70 eV): m/z (%) = 716 (5)
[M+1]+, 595 (7), 302 (10), 268 (32), 91 (100), 78 (40). –
C36H57N7O8 (715): calcd. C 60.42, H 7.97, N 13.71; found
C 60.21, H 7.69, N 13.45.
7d: Yield 13% [A], 40 [B]; m. p. 117 – 119 ◦C. –
pentapeptide methyl ester 7a as identified by melting point [α]2D5 = −61 (c = 0.5, MeOH). – IR (KBr): ν = 3313
and TLC in comparison with an authentic sample prepared (NH, str.), 3052 (CH-Ar), 2959 (CH-aliph.), 1736
according to method A.
(C=O, ester), 1655, 1527, 1233 (C=O, amide I, II
7a: Yield 20% [A] (lit. [29]: 60), 61 [B], 45 [C], 17 [D] and III) cm−1. – 1H NMR (500 MHz, [D6]DMSO):
(lit. [35]: 29); m. p. 145 – 147 ◦C [lit. [29]: 148 – 150oC]. δ = 8.90 and 8.00 (2s, 6H, 6 NH, D2O exchangeable),
– [α]2D5 = −70 (c = 0.5, MeOH). – IR (KBr): ν = 3254 8.30 – 8.20 (m, 3H, pyridyl-H), 7.30 – 7.20 (m, 10H,
(NH, str.), 3033 (CH-Ar), 2931 (CH-aliph.), 1751 (C=O, 2Ar-H), 4.10 – 4.00 (m, 4H, 4CHNH), 3.95 – 3.90 (t, 1H,
ester), 1644, 1530, 1235 (C=O, amide I, II and III) cm−1
.
NHCH2CH2CH2CH2CHNH), 3.67 (s, 3H, OCH3),
–
1H NMR (500 MHz, [D6]DMSO): δ = 8.65 and 8.00 3.45 – 3.40
(m, 2H, NHCH2CH2CH2CH2CHNH),
(2s, 6H, 6NH, D2O exchangeable), 8.30 – 8.20 (m, 3H, 3.20 – 3.10 (dd, 4H, 2 CH2Ph), 1.50 – 1.40 (m, 8H, 4
pyridyl-H), 4.50 – 4.40 (m, 4H, 4CHNH), 4.25 – 4.20 (t, CH2), 1.30 – 1.20 (m, 6H, 3 CH2), 1.15 – 0.95 (m, 6H, 2
1H, NHCH2CH2CH2CH2CHNH), 3.55 (s, 3H, OCH3), CH2CH2CH3). – MS (EI, 70 eV): m/z (%) = 782 (25)
3.25 – 3.20 (m, 2H, NHCH2CH2), 1.90 – 1.80 (m, 2H, [M+1]+, 632 (20), 503 (45), 349 (40), 91 (100), 78 (40). –

M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
815
C42H53N7O8 (783): calcd. C 64.37, H 6.77, N 12.52; found was filtered off. Tetrahydrofuran was evaporated to dryness,
C 64.22, H 6.43, N 12.09.
and the obtained residue was dissolved in dichloromethane,
washed with 1 N sodium bicarbonate, 1 N potassium hydro-
7e: Yield 16% [A], 48 [B]; m. p. 140 – 142 ◦C. – [α]D25
=
−58 (c = 0.5, MeOH). – IR (KBr): ν = 3299 (NH, str.), gen sulfate and water, and then dried over anhydrous sodium
3028 (CH-Ar), 2956 (CH-aliph.), 1745 (C=O, ester), 1655, sulfate. The solvent was evaporated to dryness, and the ob-
1529, 1216 (C=O, amide I, II and III) cm−1. – 1H NMR tained oily residue was solidified by trituration with dry
(500 MHz, [D6]DMSO): δ = 8.85 and 8.00 (2s, 6H, 6 ether-n-hexane mixture. The obtained solid was collected by
NH, D2O exchangeable), 8.30 – 8.20 (m, 3H, pyridyl-H), filtration and crystallized from ethanol-n−hexane to give the
7.20 – 7.05 (m, 20H, 4 Ar-H), 4.90 – 4.80 (m, 4H, 4 CHNH), cyclic derivatives 8a,b as identified by melting point and
4.25 – 4.20 (t,1H, NHCH2CH2CH2CH2CHNH), 3.75 (s, 3H, TLC in comparison with authentic samples prepared accord-
OCH3), 3.40 – 3.35 (m, 2H, NHCH2CH2CH2CH2CHNH), ing to method A.
3.20 – 3.10 (m, 8H, 4 CH2Ph), 1.45 – 1.40 (m, 4H, 2 CH2),
8a: Yield 14% [A], 46 [B]; m. p. 92 – 93 ◦C. – [α]D25
=
1.30 – 1.25 (m, 2H, CH2). – MS (EI, 70 eV): m/z (%) = 879 −41 (c = 0.5, MeOH). – IR (KBr): ν = 3310 (NH, str.), 3028
(0.5) [M]+, 759 (2), 263 (44), 120 (54), 91 (100), 78 (22). – (CH-Ar), 2959 (CH-aliph.), 1661, 1529, 1247 (C=O, amide
C50H53N7O8 (879): calcd. C 68.26, H 6.03, N 11.15; found I, II and III) cm−1. – 1H NMR (500 MHz, [D6]DMSO):
C 68.15, H 5.97, N 10.98.
δ = 8.85 and 8.00 (2s, 6H, 6NH D2O exchangeable),
8.40 – 8.30 (m, 3H, pyridyl-H), 4.10 – 4.00 (m, 4H, 4
CHNH), 3.45 – 3.30 (m, 4H, NHCH2CH2CH2CH2NH),
1.90 – 1.80 (m, 2H, 2 CH2CH(CH3)2), 1.70 – 1.60 (m,
4H, 2 CH2), 1.40 – 1.30 (m, 8H, 4 CH2), 1.30 – 1.20 (m,
4H, 2 CH2), 1.05 – 0.85 (m, 18H, 2 CH2CH2CH3 and 2
CH2CH(CH3)2). – MS (EI, 70 eV): m/z (%) = 647 (2)
[M+3]+, 606 (5), 302 (3), 267 (100), 185 (25), 78 (30). –
C33H53N7O6 (643): calcd. C 61.59, H 8.24, N 15.24; found
C 61.34, H 8.12, N 15.03.
Synthesis of
cyclo-(Nα -dipicolinoyl)-bis[L-Leu-DL-Nva]-aliphatic
diamines 8a, b
Method A: mixed anhydride method
Ethyl chloroformate (0.2 mL, 2 mmol) was added to
a stirred and cold (−15 ◦C) dichloromethane (20 mL) so-
lution of Nα -dipicolinoyl-bis-[L-Leu-DL-Nva] (6a) (0.59 g,
1 mmol) containing N-methylmorpholine (0.2 mL, 2 mmol).
The reaction mixture was stirred for additional 20 min,
then a cold dichloromethane solution (20 mL) of 1,4-
diaminobutane (0.09 g, 1 mmol) or 1,6-diaminohexane
(0.12 g, 1 mmol) was added. Stirring was maintained for 3 h
at −15 ◦C, then for 12 h at room temperature. The reaction
mixture was washed with water, 1 N sodium bicarbonate, 1 N
potassium hydrogen sulfate and water, and then dried over
anhydrous sodium sulfate. The solvent was evaporated to
dryness, and the obtained oily residue was solidified by tritu-
ration with a dry ether-n-hexane mixture. The crude product
was purified by preparative thin layer chromatography using
(S3) as eluent to give the corresponding cyclic derivatives
8a,b.
8b: Yield 18% [A], 49 [B]; m. p. 136 – 138 ◦C. –
[α]2D5 = −20.2 (c = 0.5, MeOH). – IR (KBr): ν = 3315
(NH, str.), 3045 (CH-Ar), 2959 (CH-aliph.), 1660, 1530,
1249 (C=O, amide I, II and III) cm−1. – 1H NMR
(500 MHz, [D6]DMSO): δ = 8.75 and 8.00 (2s, 6H,
6NH, D2O exchangeable), 8.40 – 8.30 (m, 3H, pyridyl-
H), 4.20 – 4.10 (m, 4H, 4 CHNH), 3.40 – 3.30 (m, 4H,
NHCH2CH2CH2CH2CH2CH2NH), 1.90 – 1.80 (m, 2H, 2
CH2CH(CH3)2), 1.70 – 1.60 (m, 4H, 2 CH2), 1.40 – 1.30 (m,
8H, 4 CH2), 1.30 – 1.20 (m, 8H, 4 CH2), 1.00 – 0.80 (m, 18H,
2 CH2CH2CH3 and 2 CH2CH(CH3)2). – MS (EI, 70 eV):
m/z (%) = 671 (2) [M]+, 601 (3), 302 (5), 143 (45), 78 (2),
56 (100). – C35H57N7O6 (671): calcd. C 62.59, H 8.49, N
14.61; found C 62.28, H 8.32, N 14.19.
Method B: DCC method
Synthesis of cyclo-(Nα -dipicolinoyl)-bis
[dipeptide]-L-Lys(cyclic pentapeptides) 9a – e
A cold (−5 ◦C) tetrahydrofuran solution (20 mL) of
1,4-diaminobutane (0.09 g, 1 mmol) or 1,6-diaminohexane
(0.12 g, 1 mmol) was added to a stirred dry tetrahydrofu-
To a stirred and cold methanolic solution (−5 ◦C, 20 mL)
ran solution (−5 ◦C, 20 mL) of Nα -dipicolinoyl-bis[L-Leu- of the corresponding cyclic pentapeptide methyl ester 7a – e
DL-Nva] (6a) (0.59 g, 1 mmol). Dicyclohexyl- carbodiimide (1 mmol), sodium hydroxide (1 N, 25 mL) was gradually
(0.42 g, 2 mmol) was then added to the reaction mixture, added. The reaction mixture was stirred for 2 h at the same
in portions, over 20 min at the same temperature. Stirring temperature, then for 3 h at room temperature. The solvent
was maintained for 20 h at room temperature. The reaction was distilled off under reduced pressure, and the remain-
mixture was then diluted with acetonitrile (20 mL), and the ing aqueous solution was cooled and acidified with 1 N hy-
formed dicyclohexylurea was filtered off and washed with drochloric acid to pH ∼ 3. The obtained solid was filtered off,
acetonitrile (2 × 10 mL). The filtrate was kept in the re- washed with water, dried and crystallized from ethanol-water
frigerator overnight, and the newly formed dicyclohexylurea to give the corresponding cyclic pentapeptides 9a – e.

816
M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
9a: Yield: 71%; m. p. 155 – 158 ◦C. – [α]D25 = −113.2 NH D2O exchangeable), 8.40 – 8.30 (m, 3H, pyridyl-H),
(c = 0.5, MeOH). – IR (KBr): ν = 3323 (NH, str.), 3045 7.30 – 7.20 (m, 10H, 2 Ar-H), 4.40 – 4.30 (m, 4H, 4
(CH-Ar), 2959 (CH-aliph.), 1722 (C=O, acid), 1653, 1534, CHNH), 4.25 – 4.20 (t, 1H, CH2CHNH), 3.45 – 3.40 (m,
1240 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz, 2H, NHCH2CH2CH2CH2CHNH), 3.00 – 2.90 (dd, 4H, 2
[D6]DMSO): δ = 12.50 (s, 1H, OH, D2O exchangeable), CH2Ph), 1.60 – 1.50 (m, 8H, 4 CH2), 1.30 – 1.20 (m, 6H, 3
8.85 and 8.20 (2s, 6H, 6NH, D2O exchangeable), 8.40 – 8.35 CH2), 0.95 – 0.85 (m, 6H, 2 CH2CH2CH3). – MS (EI, 70 eV):
(m, 3H, pyridyl-H), 4.70 – 4.60 (m, 4H,
4
CHNH), m/z (%) = 768 (5) [M–1]+, 688 (10), 524 (23), 304 (10), 143
4.25 – 4.20 (t, 1H, NHCH2CH2CH2CH2CHNH), 3.15 – 3.10 (27), 56 (100). – C41H51N7O8 (769): calcd. C 63.98, H 6.63,
(m, 2H, NHCH2CH2CH2CH2CHNH), 1.90 – 1.80 (m, 2H, 2 N 12.74; found C 63.15, H 6.43, N 12.31.
CH2CH(CH3)2), 1.70 – 1.60 (m, 4H, 2 CH2), 1.45 – 1.40 (m,
9e: Yield: 71%; m. p. 151 – 153 ◦C. – [α]D25 = −16.6
8H, 4 CH2), 1.30 – 1.20 (m, 6H, 3 CH2), 0.95 – 0.85 (m, 18H, (c = 0.5, MeOH). – IR (KBr): ν = 3325 (NH, str.), 3028
2 CH2CH2CH3 and 2 CH2CH(CH3)2). – MS (EI, 70 eV): (CH-Ar), 2929 (CH-aliph.), 1720 (C=O, acid), 1656, 1528,
m/z (%) = 701 (7) [M]+, 572 (12), 430 (44), 304 (68), 199 1230 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz,
(22), 125 (100). – C35H55N7O8 (701): calcd. C 59.91, H [D6]DMSO): δ = 8.75 and 8.10 (2s, 6H, 6 NH, D2O ex-
7.85, N 13.98; found C 59.57, H 7.63, N 13.18.
changeable), 8.40 – 8.35 (m, 3H, pyridyl-H), 7.20 – 7.10 (m,
20H, 4 Ar-H), 4.70 – 4.60 (m, 4H, 4 CHNH), 4.15 – 4.10
(t,1H, CH2CHNH), 3.55 – 3.50 (m, 2H, NHCH2CH2CH2-
CH2CHNH), 3.00 – 2.90 (m, 8H, 4 CH2Ph), 1.50 – 1.40 (m,
4H, 2 CH2), 1.30 – 1.25 (m, 2H, CH2). – MS (EI, 70 eV): m/z
(%) = 865 (20) [M]+, 498 (52), 370 (13), 205 (21), 91 (100).
– C49H51N7O8 (865): calcd. C 67.97, H 5.89, N 11.33; found
C 67.48, H 5.12, N 11.01.
9b: Yield: 56%; m. p. 146 – 148 ◦C. – [α]D25 = −2.8
(c = 0.5, MeOH). – IR (KBr): ν = 3326 (NH, str.),
3033 (CH-Ar), 2930 (CH-aliph.), 1721 (C=O, acid), 1653,
1537, 1230 (C=O, amide I, II and III) cm−1. – 1H NMR
(500 MHz, [D6]DMSO): δ = 12.55 (s, 1H, OH, D2O ex-
changeable), 8.65 and 8.22 (2s, 6H, 6 NH, D2O ex-
changeable), 8.35 – 8.30 (m, 3H, pyridyl-H), 7.30 – 7.20 (m,
10H, 2Ar-H), 4.60 – 4.50 (m, 4H,4 CHNH), 4.10 – 4.05
(t, 1H, NHCH2CH2CH2-CH2CHNH), 3.55 – 3.50 (m,
2H, NHCH2CH2CH2CH2CHNH), 3.20 – 3.10 (dd, 4H,
2CH2Ph), 1.95 – 1.85 (m, 2H, 2 CH2CH(CH3)2), 1.80 – 1.70
(m, 4H, 2 CH2), 1.45 – 1.40 (m, 4H, 2 CH2), 1.35 – 1.30 (m,
2H, CH2), 0.95 – 0.85 (m, 12H, 2 CH2CH(CH3)2). – MS
(EI, 70 eV): m/z (%) = 797 (5) [M]+, 755 (20), 477 (15),
302 (60), 231 (26), 91 (100). – C43H55N7O8 (797): calcd. C
64.74, H 6.90, N 12.29; found C 64.62, H 6.45, N 12.03.
9c: Yield: 60%; m. p. 140 – 143 ◦C. – [α]D25 = −85
(c = 0.5, MeOH). – IR (KBr): ν = 3326 (NH, str.), 3044
(CH-Ar), 2930 (CH-aliph.), 1721 (C=O, acid), 1653, 1537,
1230 (C=O, amide I, II and III) cm−1. – 1H NMR (500 MHz,
[D6]DMSO): δ = 12.50 (s, 1H, OH, D2O exchangeable),
8.85 and 8.00 (2s, 6H, 6 NH, D2O exchangeable), 8.40 – 8.30
Synthesis of
cyclo-(Nα -dipicolinoyl)-bis[dipeptide]-L-Lys-NHNH2
(cyclic pentapeptide hydrazides) 10a – e
To a stirred methanolic solution (20 mL) of the corre-
sponding cyclic pentapeptide methyl ester 7a – e (1 mmol),
anhydrous hydrazine hydrate (0.35 mL, 10 mmol) was
added. The reaction mixture was refluxed for 3 h, after which
the solvent was evaporated. The obtained residue was tritu-
rated with ether, filtered off and crystallized from methanol-
ether to afford the corresponding cyclic hydrazides 10a – e.
10a: Yield: 66%; m. p. 165 – 167 ◦C. – [α]D25 = −24.8
(c = 0.5, MeOH). – IR (KBr): ν = 3318 (NH, str.), 3028
(CH-Ar), 2931 (CH-aliph.), 1657, 1531, 1240 (C=O, amide
I, II and III) cm−1. – 1H NMR (500 MHz, [D6]DMSO):
δ = 8.85, 8.75 and 8.00 (3s, 7H, 7NH, D2O exchange-
able), 8.30 – 8.25 (m, 3H, pyridyl-H), 4.70 – 4.60 (m, 4H, 4
CHNH), 4.15 – 4.10 (t, 1H, NHCH2CH2-CH2CH2CHNH),
3.45 (bs, 2H, NH2, D2O exchangeable), 3.20 – 3.15 (m,
2H, NHCH2CH2CH2-CH2CHNH), 1.90 – 1.80 (m, 2H, 2
CH2CH(CH3)2), 1.70 – 1.60 (m, 4H, 2 CH2), 1.50 – 1.40 (m,
8H, 4 CH2), 1.30 – 1.20 (m, 6H, 2 CH2), 1.05 – 0.85 (m, 18H,
2 CH2CH2CH3 and 2CH2-CH(CH3)2). – MS (EI, 70 eV):
m/z (%) = 716 (2) [M+1]+, 603 (5), 266 (12), 185 (100),
(m, 3H, pyridyl-H), 4.60 – 4.50 (m, 4H,
4 CHNH),
4.25 – 4.20 (t, 1H, NHCH2CH2CH2CH2CHNH), 3.20 – 3.15
(m, 2H, NHCH2CH2CH2CH2CHNH), 1.90 – 1.80 (m, 2H, 2
CH2CH(CH3)2), 1.70 – 1.60 (m, 4H, 2 CH2), 1.45 – 1.35 (m,
8H, 4 CH2), 1.25 – 1.15 (m, 6H, 3 CH2), 0.95 – 0.85 (m, 18H,
2 CH2CH2CH3 and 2 CH2CH(CH3)2). – MS (EI, 70 eV):
m/z (%) = 701 (1) [M]+, 678 (2), 318 (85), 236 (45), 154
(66), 56 (100). – C35H55N7O8 (701): calcd. C 59.91, H 7.85,
N 13.98; found C 59.76, H 7.48, N 13.29.
9d: Yield: 60%; m. p. 148 – 150 ◦C. – [α]D25 = −45.4 102 (18), 78 (5). – C35H57N9O7 (715): calcd. C 58.74, H
(c = 0.5, MeOH). – IR (KBr): ν = 3325 (NH, str.), 7.97, N 17.62; found C 58.35, H 7.43, N 17.36.
3028 (CH-Ar), 2930 (CH-aliph.), 1723 (C=O, acid),
1655, 1528, 1241 (C=O, amide I, II and III) cm−1
1H NMR (500 MHz, [D6]DMSO): δ = 12.55 (s, 1H, str.), 3035 (CH-Ar), 2959 (CH-aliph.), 1657, 1530,
OH, D2O exchangeable), 9.25 and 8.00 (2s, 6H, 6 1242 (C=O, amide I, II and III) cm−1. – 1H NMR
10b: Yield: 52%; m. p. 130 – 132 ◦C. [α]D25
– =
−22.2 (c = 0.5, MeOH). – IR (KBr): ν = 3325 (NH,
.
–

M. H. Abo-Ghalia et al. · Chiral Linear and Macrocyclic Tetra- and Penta-peptide Candidates
817
(500 MHz, [D6]DMSO): δ = 9.50 and 8.20 (2s, 7H, 7 7 NH, D2O exchangeable), 8.30 – 8.25 (m, 3H, pyridyl-
NH, D2O exchangeable), 8.40 – 8.35 (m, 3H, pyridyl- H), 7.30 – 7.20 (m, 10H, 2 Ar-H), 4.90 – 4.80 (m, 4H, 4
H), 7.20 – 7.10 (m, 10H, 2 Ar-H), 4.50 – 4.40 (m, 4H, 4 CHNH), 4.15 – 4.10 (t, 1H, NHCH2CH2CH2CH2CHNH),
CHNH), 4.20 – 4.015 (t, 1H, NHCH2CH2CH2CH2CHNH), 3.45 (bs, 2H, NH2, D2O exchangeable), 3.30 – 3.25 (m,
3.45 (bs, 2H, NH2, D2O exchangeable), 3.30 – 3.25 (m, 2H, NHCH2CH2CH2CH2CHNH), 3.15 – 3.10 (dd, 4H, 2
2H, NHCH2CH2CH2CH2CHNH), 2.90 – 2.80 (dd, 4H, 2 CH2Ph), 1.50 – 1.40 (m, 8H, 4 CH2), 1.30 – 1.20 (m, 6H, 3
CH2Ph), 1.80 – 1.70 (m, 2H, 2 CH2CH(CH3)2), 1.60 – 1.50 CH2), 0.95 – 0.85 (m, 6H, 2 CH2CH2CH3). – MS (EI, 70 eV):
(m, 4H, 2 CH2), 1.45 – 1.40 (m, 4H, 2 CH2), 1.35 – 1.30 (m, m/z (%) = 784 (6) [M+1]+, 706 (7), 316 (33), 276 (50), 149
2H, CH2), 0.95 – 0.85 (m, 12H, 2 CH2CH(CH3)2). – MS (EI, (100), 78 (80). – C41H53N9O7 (783): calcd. C 62.83, H 6.77,
70 eV): m/z (%) = 811 (9) [M]+, 720 (7), 373 (10), 187 (60), N 16.09; found C 62.51, H 6.65, N 15.89.
131 (46), 91 (100). – C43H57N9O7 (811): calcd. C 63.62, H
7.03, N 15.53; found C 63.42, H 6.91, N 15.23.
10e: Yield: 74%; m. p. 184 – 186 ◦C. – [α]D25 = −53.8
(c = 0.5, MeOH). – IR (KBr): ν = 3298 (NH stretch-
10c: Yield: 67%; m. p. 116 – 118 ◦C. – [α]D25 = −11.0 ing), 3028 (CH-Ar), 2930 (CH-aliph.), 1657, 1528, 1230
(c = 0.5, MeOH). – IR (KBr): ν = 3300 (NH, str.), 3053 (C=O amide I, II and III) cm−1. – 1H NMR (500 MHz,
(CH-Ar), 2958 (CH-aliph.), 1657, 1529, 1240 (C=O, amide [D6]DMSO): δ = 8.90, 8.80 and 8.00 (3s, 7H, 7 NH, D2O
I, II and III) cm−1. – 1H NMR (500 MHz, [D6]DMSO): exchangeable), 8.40 – 8.35 (m, 3H, pyridyl-H), 7.20 – 7.10
δ = 8.85 and 8.10 (2s, 7 H, 7 NH, D2O exchange- (m, 20H, 4 Ar-H), 4.80 – 4.70 (m, 4H, 4 CHNH), 4.10 – 4.05
able), 8.35 – 8.30 (m, 3H, pyridyl-H), 4.50 – 4.40 (m, 4H, 4 (t, 1H, NHCH2CH2CH2CH2CHNH), 3.55 – 3.50 (m, 2H,
CHNH), 4.05 – 4.00 (t, 1H, NHCH2CH2CH2-CH2CHNH), NHCH2CH2CH2CH2CHNH), 3.45 (bs, 2H, NH2 D2O
3.45 (bs, 2H, NH2, D2O exchangeable), 3.05 – 3.00 (m, exchangeable), 3.30 – 3.20 (m, 8H, 4 CH2Ph), 1.50 – 1.40
2H, NHCH2CH2CH2CH2-CHNH), 1.90 – 1.80 (m, 2H, (m, 4H, 2CH2), 1.30 – 1.25 (m, 2H, CH2). – MS (EI, 70 eV):
2CH2CH(CH3)2), 1.60 – 1.50 (m, 4H, 2 CH2), 1.40 – 1.30 m/z (%) = 879 (3) [M]+, 749 (4), 279 (19), 192 (50), 91
(m, 8H, 4 CH2), 1.20 – 1.10 (m, 6H, 3 CH2), 0.95 – 0.85 (47), 56 (100). – C49H53N9O7 (879): calcd. C 66.89, H 6.03,
(m, 18H, 2 CH2CH2CH3 and 2 CH2CH(CH3)2). – MS (EI, N 14.33; found C 66.49, H 5.92, N 14.06.
70 eV): m/z (%) = 715 (6) [M]+, 654 (5), 348 (26), 330 (75),
225 (82), 78 (100). – C35H57N9O7 (715): calcd. C 58.74, H
Acknowledgement
7.97, N 17.62; found C 58.23, H 7.35, N 17.21.
10d: Yield: 58%; m. p. 136 – 138 ◦C.
– =
[α]2D5
The authors extend their appreciation to the Dean-
ship of Scientific Research at King Saud University for
funding the work through the research group project
no. RGP-VPP-172.
−47.6 (c = 0.5, MeOH). – IR (KBr): ν = 3302 (NH,
str.), 3045 (CH-Ar), 2932 (CH-aliph.), 1658, 1528,
1233 (C=O, amide I, II and III) cm−1. – 1H NMR
(500 MHz, [D6]DMSO): δ = 8.90, 8.85 and 8.00 (3s, 7H,
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