Microwave-enhanced solution coupling of the α,α-dialkyl amino acid, Aib

Article abstract of DOI:10.1016/S0040-4039(01)00806-1

The difficult coupling of α-aminoisobutyric acid (Aib), during the synthesis of dipeptides (1-6), was carried out using PyBOP/HOBt and HBTU/HOBt reagents by application of microwave energy in the presence of solvent. Room temperature, conventional heating

Full text of DOI:10.1016/S0040-4039(01)00806-1

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 42 (2001) 5171–5173
Microwave-enhanced solution coupling of the a,a-dialkyl amino
acid, Aib†
Vincenzo Santagada,a,* Ferdinando Fiorino,a Elisa Perissutti,a Beatrice Severino,a
Vincenzo De Filippis,b Beniamino Vivenzioa and Giuseppe Caliendoa
aDipartimento di Chimica Farmaceutica e Tossicologica, Universita` di Napoli, Federico II, Via D. Montesano, 49,
80131 Naples, Italy
bDipartimento di Scienze Farmaceutiche, Universita` di Padova, Via F. Marzolo 5, 35131 Padua, Italy
Received 11 April 2001; revised 14 May 2001; accepted 15 May 2001
Abstract—The difficult coupling of a-aminoisobutyric acid (Aib), during the synthesis of dipeptides (1–6), was carried out using
PyBOP/HOBt and HBTU/HOBt reagents by application of microwave energy in the presence of solvent. Room temperature,
conventional heating (oil bath) and microwave irradiation of the reactions are compared. Synthesis by microwave irradiation gave
the desired compounds in higher yields and in shorter reaction times than those obtained by conventional heating or at room
temperature. © 2001 Elsevier Science Ltd. All rights reserved.
a-Aminoisobutyric acid (Aib), the prototype of a,a-di-
substituted glycines, is a non-coded amino acid found
in membrane channel-forming peptides of microbial
origin.1 The presence of the extra-methyl group at the
Ca atom dramatically restricts the conformational space
accessible to Aib and forces the peptide chain into a
left- or right-handed helical conformation.2–7 Due to
the stringency of the steric requirements, as well as to
the high stability8 and crystallizability2 of its peptide
derivatives, Aib has been often used to study the rela-
tionships existing between structure, stability and func-
tion in bioactive peptides,9–11 long polypeptide chains,12
and even proteins.13 However, the incorporation of Aib
into even short peptides has been strongly limited by
poor yields in both solution and solid-phase synthesis
using standard coupling reagents (i.e. DCC/HOBt),
even for prolonged reaction times.14,15 Other methods
have been used, i.e. active esters,16 mixed anhy-
drides,17,18 but none of them gives consistently satisfac-
tory results. Better results were obtained using BOP,
PyBOP, PyBroP and BrOP.19
These limitations prompted us to explore novel experi-
mental conditions, in order to improve Aib coupling
efficiency and reduce reaction times. In recent years, the
application of microwaves to organic synthesis has
become increasingly popular,20–27 by shortening reac-
tion times and/or increasing yields. In the present
paper, we report the results of a systematic study on the
application of microwaves in the coupling of Aib to
sterically hindered natural (i.e. Val, Ile) or non-coded
(i.e. Aib) amino acids, using PyBOP/HOBt and HBTU/
HOBt as coupling agents28 (Table 1). Coupling yields
and reaction times, obtained at room temperature and
conventional heating (oil bath), were compared to those
obtained by microwave irradiation.
Keywords: Aib; difficult couplings; microwave irradiation in solution.
* Corresponding author. Tel.: 0039-81-678648; fax: 0039-81-678649;
e-mail: santagad@unina.it
† Abbreviations and symbols follow the recommendations of the
IUPAC-IUB Joint Commission on Biochemical Nomenclature
(Eur. J. Biochem. 1984, 138, 9). In addition the following abbrevia-
tions are used: PyBOP, (1H-1,2,3-benzotriazol-1-yloxy)-tris(pyrro-
lidino)-phosphonium hexafluorophosphate; HBTU, O-benzotriazol-
1-yl-N,N,N%,N%-tetramethyluronium hexafluorophosphate; BOP,
(1H-1,2,3-benzotriazol-1-yloxy)-tris(dimethylamino)-phosphonium
hexafluorophosphate; BroP, bromo-tris(dimethylamino)-phospho-
nium hexafluorophosphate; PyBroP, bromo-tris(pyrrolidino)-phos-
phonium hexafluorophosphate; DCC, dicyclohexyl-carbodiimide;
HOBt, 1,2,3-benzotriazole-1-hydroxide; DIEA, diisopropylethyl-
amine.
The synthetic procedure by microwave irradiation was
performed using a microwave oven (ETHOS 1600,
Milestone) specifically designed for organic synthesis.21
The results obtained with the coupling reagents
PyBOP/HOBt and HBTU/HOBt are summarized in the
Table 1, together with the experimental conditions
employed. For compounds 1–4 we obtained high yields
(76 to 87% in 1 h reaction at rt and 55°C, respectively)
0040-4039/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(01)00806-1

Table 1. Comparison of the coupling yields for the synthesis of Aib-containing dipeptides at 23 and 55°C by conventional heating versus microwave irradiationa
Peptide
Rt (2392°C)
Conventional heatingb
PyBOP/HOBt HBTU/HOBt
Microwave Irradiationc
PyBOP/HOBt HBTU/HOBt
PyBOP/HOBt
Yield (%) Time (h)
HBTU/HOBt
Yield (%) Time (h)
Yield (%)
84
Time (h)
1
Yield (%)
77
Time (h)
16
Yield (%)
96
Time (min)
30
Yield (%)
94
Time (min)
30
1
80
82
76
78
84
80
1
1
74
35
40
45
86
50
16
16
16
16
16
16
Z-Aib-Val-
OCH3
2
3
4
5
6
87
85
80
86
88
1
1
45
50
53
90
60
16
16
16
16
16
94
90
94
92
88
30
30
30
30
30
95
93
90
90
86
30
30
30
30
30
/
Z-Val-Aib-
OCH3
1
Z-Aib-Ile-
OCH3
1
1
Boc-Ile-Aib-
OCH3
4
)
16
16
16
16
Z-Aib-Aib-
OCH3
5
Boc-Aib-Aib-
OCH3
5
a All dipeptides were synthesized by standard protocols,27 unless otherwise specified. The reaction yields were determined by RP-HPLC analysis assuming a similar extinction coefficients at 220 nm for
reagents and products. The values reported correspond to the average of the values obtained from three experiments under identical experimental conditions.
b Oil-bath at 55°C.
c Power 200 W for 15 min at 55°C and then 300 W for 15 min at 60°C.

V. Santagada et al. / Tetrahedron Letters 42 (2001) 5171–5173
5173
using PyBOP/HOBt, while with HBTU/HOBt the
yields were lower (35–77% for 1 h reaction at rt and
55°C, respectively). The results were identical whether
Aib was in the C- or N-terminal position except for
compound 1 when HBTU/HOBt method was used
(both at rt or 55°C). Compounds 5 and 6 involved the
difficult coupling of two Aib residues. For the HBTU/
HOBt mediated coupling of Na-protected Aib-OH to
H-Aib-OMe (compounds 5 and 6) the coupling yields
varied significantly whether Z- or Boc-protecting
groups were used. These differences were not observed
when PyBOP/HOBt activation method was used.
Instead, coupling under microwave irradiation of solu-
tions gave, in any case, the desired dipeptides in higher
yields (86–96% in 40 min) and cleaner reactions than
those obtained by conventional heating or at rt. The
overall reaction times were dramatically reduced, from
16 h to 30 min. Moreover, the coupling yields were not
influenced by the particular activator employed
(PyBOP and HBTU), or there were no appreciable
differences in the coupling yields between the two cou-
pling reagents, or by the Na-protection (Z- or Boc) of
the incoming residue (compounds 5 and 6).
Temussi, P. A.; Selinger, Z.; Raider, F.; Chorev, M.
Biopolymers 1993, 33, 915–926.
12. De Filippis, V.; De Antoni, F.; Frigo, M.; de Laureto
Polverino, P.; Fontana, A. Biochemistry 1998, 37, 1686–
1696.
13. Ellman, J. A.; Mendel, D.; Schultz, P. G. Science 1992,
255, 197–200.
14. Leibfritz, D.; Haupt, E.; Dubischar, N.; Lachmann, H.;
Oekonomopoulos, R.; Jung, G. Tetrahedron 1982, 38,
2165–2181.
15. Balasubramanian, T. M.; Kendrick, N. C. E.; Taylor, M.;
Marshall, G. R.; Hall, J. E.; Vodyanov, I.; Reusser, F. J.
Am. Chem. Soc. 1981, 103, 6127–6132.
16. Mc Gahren, W. J.; Goodman, M. Tetrahedron 1967, 23,
2017–2030.
17. Leplawy, M. T.; Jones, D. S.; Kenner, G. W.; Sheppard,
R. C. Tetrahedron 1960, 11, 39–51.
18. Jones, D. S.; Kenner, G. W.; Preston, J.; Sheppard, R. C.
J. Chem. Soc. 1965, 6227.
19. Frerot, E.; Coste, J.; Pantaloni, A.; Dufour, M. N.;
Jouin, P. Tetrahedron 1991, 47, 259–270.
20. Varma, R. S. Green Chem. 1999, 43–55.
21. Santagada, V.; Perissutti, E.; Fiorino, F.; Vivenzio, B.;
Caliendo, G. Tetrahedron Lett. 2001, 42, 2397–2400.
22. Loupy, A.; Petit, A.; Hamelin, J.; Texier-Boullet, F.;
Jacquault, P.; Mathe, D. Synthesis 1998, 1213–1234.
23. Bose, A. K.; Banik, B. K.; Lavlinskaia, N.; Jayaraman,
M.; Manhas, M. S. Chemtech. 1997, 27, 18–24.
24. Galema, S. A. Chem. Soc. Rev. 1997, 26, 233–238.
25. Majetich, G.; Hicks, R. Radiat. Phys. Chem. 1995, 45,
567–579.
In conclusion, we have shown that the application of
microwave irradiation improves coupling yields and
significantly reduces reaction times in the solution syn-
thesis of Aib-containing dipeptides. Our findings will be
useful in the solution and possibly solid-phase peptide
synthesis, when the incorporation of Aib or other steri-
cally hindered amino acid residues into peptide chains
is required.
26. Caddick, S. Tetrahedron 1995, 51, 10403–10432.
27. Strauss, C. R.; Trainor, R. W. Aust. J. Chem. 1995, 48,
1665–1692.
References
28. Coupling Methods: PyBOP/HOBt and HBTU/HOBt
General procedure: To a DMF mixture of 1 mmol of
N-protected acid component (Z- or Boc-Xaa-OH), 1.1
mmol of coupling reagent (HBTU/HOBt or PyBOP/
HOBt), and 1.1 mmol of the C-protected amino acid
1. Nagaraj, R.; Balaram, P. Acc. Chem. Res. 1991, 14,
356–363 and references cited herein.
2. Karle, I. L.; Balaram, P. Biochemistry 1990, 29, 6747–
6756.
(H-Xaa-OCH3),
2
mmol of diisopropylethylamine
3. Toniolo, C.; Benedetti, E. Macromolecules 1991, 24,
4004–4009.
4. Pavone, V.; Benedetti, E.; Di Blasio, B.; Pedone, C.;
Santini, A.; Bavoso, A.; Toniolo, C.; Crisma, M. J. Mol.
Biol. 1990, 214, 633–635.
5. Toniolo, C.; Benedetti, E. Trends Biochem. Sci. 1991, 16,
350–353.
6. Burgess, A. W.; Leech, S. Biopolymers 1973, 12, 2599–
2605.
7. Paterson, Y.; Rumsey, S. M.; Benedetti, E.; Nemethy, G.;
Scheraga, H. A. J. Am. Chem. Soc. 1981, 103, 2947–2955.
8. Augsburger, J. D.; Bindra, V. A.; Scheraga, H. A.; Kuki,
A. Biochemistry 1995, 34, 2566–2576.
9. Cotton, R.; Giles, M. G.; Miller, L.; Shaw, J. S.; Timms,
D. Eur. J. Pharmacol. 1984, 97, 331–332.
10. Samanen, J.; Cash, T.; Narindray, D.; Brandeis, E.;
Adams, W.; Weideman, H.; Yellin, T. J. Med. Chem.
1991, 34, 3036–3043.
(DIPEA, 3 mmol if amine salt was present) was added.
The reaction was stirred at rt or by conventional heating
or by microwave irradiation according to than reported
in Table 1. The reaction mixture was separated from the
solvent, the crude residue was taken up in ethyl acetate
and washed successively three times with citric acid (5%),
sodium bicarbonate (5%), and a saturated solution of
sodium chloride. All dipeptides obtained were purified by
preparative RP-HPLC and the homogeneity of the
purified products was assessed by analytical RP-HPLC
with a Vydac C18-column (5 mm, 4.6×250 mm, spherical).
Analytical determinations were carried out by two sol-
vent systems: A: 10%, (v/v), acetonitrile in 0.1% TFA, B:
60% (v/v), acetonitrile in 0.1% TFA (linear gradient from
100 A to 100% B over 25 min, UV detection at 220 nm,
flow rate 1 mL/min). The final compounds were charac-
terized by mass spectrometry (LCQ Thermoquest-Ion
Trap) and the data were consistent with the considered
structures.
11. Tallon, M.; Ron, D.; Halle, D.; Amodeo, P.; Saviano, G.;
.