Syntheses of racemic and non-racemic silicon- and germanium-containing α-amino acids of the formula type H2NCH(CH2ElR3)COOH (El=Si, Ge; R=organyl) and incorporation of D-H2NCH(CH2SiMe3)COOH and D-H2NCH(CH2GeMe3)COOH into biologically active decapeptides: A study on C/Si/Ge bioisosterism

Article abstract of DOI:10.1016/S0022-328X(01)00783-5

Two novel efficient methods for the synthesis of racemic silicon- and germanium-containing α-amino acids of the formula type rac-H₂NCH(CH₂ElR₃)COOH (El=Si, Ge; R=organyl), starting from 3,6-diethoxy-2,5-dihydropyrazine, have been developed. Racemic α-amino acids synthesized: rac-H₂NCH(CH₂SiMe₃)COOH (rac-2), rac-H₂NCH(CH₂GeMe₃)COOH (rac-3), rac-H₂NCH(CH₂SiMe₂Ph)COOH (rac-4), rac-H₂NCH(CH₂GeMe₂Ph)COOH (rac-5), and rac-H₂NCH(CH₂SiMe₂CH=CH₂)COOH (rac-6). Preparative liquid-chromatographic resolution of rac-2 and rac-3 [CHIROBIOTIC T (glycopeptide Teicoplanin covalently linked to spherical silica gel) as the stationary phase] yielded the α-amino acids (R)-2, (S)-2, (R)-3, and (S)-3. The (R)- and (S)-enantiomers of β-(trimethylsilyl)alanine [(R)- and (S)-2] and β-(trimethylgermyl)alanine [(R)- and (S)-3] are sila-analogs and germa-analogs, respectively, of the antipodes of the non-proteinogenic α-amino acid β-tert-butylalanine [(S)- and (R)-H₂NCH(CH₂CMe₃)COOH; (S)- and (R)-1]. Starting from the N-Fmoc-protected C/Si/Ge-analogous (D-configurated) α-amino acids (R)-1, (S)-2, and (S)-3, the C/Si/Ge-analogous decapeptides 7-9 [Ac-D-Nal¹-4-Cl-D-Phe²-D-Pal³-Ser ⁴-N-Me-Tyr⁵-D-Hci⁶-Nle⁷-Arg ⁸-Pro⁹-D-Me₃El-Ala¹⁰-NH₂ (7, El=C; 8, El=Si; 9, El=Ge)] were prepared by sequential solid-phase synthesis. The decapeptides 7-9 were studied in vitro in a functional assay using a recombinant cell line expressing the human GnRH receptor (agonist Triptorelin). Compounds 7-9 behaved as medium-potent GnRH antagonists, the antagonistic potencies of these three C/Si/Ge analogs being very similar.

Full text of DOI:10.1016/S0022-328X(01)00783-5

Journal of Organometallic Chemistry 628 (2001) 183–194
www.elsevier.nl/locate/jorganchem
Syntheses of racemic and non-racemic silicon- and
germanium-containing a-amino acids of the formula type
H2NCH(CH2ElR3)COOH (El=Si, Ge; R=organyl) and
incorporation of
D-H2NCH(CH2SiMe3)COOH and
D-H2NCH(CH2GeMe3)COOH into biologically active decapeptides:
a study on C/Si/Ge bioisosterism
Markus Merget a, Kurt Gu¨nther b, Michael Bernd c, Eckhard Gu¨nther c,
Reinhold Tacke a,*
a Institut fu¨r Anorganische Chemie, Uni6ersita¨t Wu¨rzburg, Am Hubland, D-97074 Wu¨rzburg, Germany
b Infracor GmbH, Postfach 1345, D-63403 Hanau, Germany
c ASTA Medica AG, Konzernforschung, Weissmu¨llerstrasse 45, D-60315 Frankfurt, Germany
Received 19 December 2000; accepted 7 March 2001
Abstract
Two novel efficient methods for the synthesis of racemic silicon- and germanium-containing a-amino acids of the formula type
rac-H2NCH(CH2ElR3)COOH (El=Si, Ge; R=organyl), starting from 3,6-diethoxy-2,5-dihydropyrazine, have been developed.
Racemic a-amino acids synthesized: rac-H2NCH(CH2SiMe3)COOH (rac-2), rac-H2NCH(CH2GeMe3)COOH (rac-3), rac-
H2NCH(CH2SiMe2Ph)COOH (rac-4), rac-H2NCH(CH2GeMe2Ph)COOH (rac-5), and rac-H2NCH(CH2SiMe2CHꢀCH2)COOH
(rac-6). Preparative liquid-chromatographic resolution of rac-2 and rac-3 [CHIROBIOTIC T (glycopeptide Teicoplanin covalently
linked to spherical silica gel) as the stationary phase] yielded the a-amino acids (R)-2, (S)-2, (R)-3, and (S)-3. The (R)- and
(S)-enantiomers of b-(trimethylsilyl)alanine [(R)- and (S)-2] and b-(trimethylgermyl)alanine [(R)- and (S)-3] are sila-analogs and
germa-analogs, respectively, of the antipodes of the non-proteinogenic a-amino acid b-tert-butylalanine [(S)- and (R)-
H2NCH(CH2CMe3)COOH; (S)- and (R)-1]. Starting from the N-Fmoc-protected C/Si/Ge-analogous (
acids (R)-1, (S)-2, and (S)-3, the C/Si/Ge-analogous decapeptides 7–9 [Ac-
-Nal1-4-Cl- -Phe2- -Pal3-Ser4-N-Me-Tyr5-
Nle7-Arg8-Pro9- -Me3El-Ala10-NH2 (7, El=C; 8, El=Si; 9, El=Ge)] were prepared by sequential solid-phase synthesis. The
D
-configurated) a-amino
D
D
D
D
-Hci6-
D
decapeptides 7–9 were studied in vitro in a functional assay using a recombinant cell line expressing the human GnRH receptor
(agonist Triptorelin). Compounds 7–9 behaved as medium-potent GnRH antagonists, the antagonistic potencies of these three
C/Si/Ge analogs being very similar. © 2001 Elsevier Science B.V. All rights reserved.
Keywords: a-Amino acids; C/Si/Ge bioisosterism; Germanium; GnRH antagonists; Peptides; Silicon
containing a-amino acids [1d,2–4] and peptides [1d,4]
(for further publications dealing with silicon-containing
amino acids and their derivatives, see Refs. [5–15]). We
have now succeeded in developing efficient methods for
the synthesis of racemic silicon- and germanium-con-
taining a-amino acids of the formula type H2NCH-
(CH2SiR3)COOH and H2NCH(CH2GeR3)COOH (R=
organyl) and for the preparative chromatographic reso-
1. Introduction
In the course of our systematic studies on
bioorganosilicon and bioorganogermanium chemistry
[1], we became interested in silicon- and germanium-
* Corresponding author. Tel.: +49-931-8885250; fax: +49-931-
8884609.
E-mail address: r.tacke@mail.uni-wuerzburg.de (R. Tacke).
lution of the a-amino acids - and
L
D-H2NCH(CH2-
SiMe3)COOH as well as - and D-H2NCH(CH2GeMe3)-
L
0022-328X/01/$ - see front matter © 2001 Elsevier Science B.V. All rights reserved.
PII: S0022-328X(01)00783-5

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M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
(for reviews dealing with CetrorelixINN, see Ref. [16]).
This decapeptide bears an (S)-tyrosine residue in posi-
tion 5 [instead of (S)-N-methyltyrosine], an (R)-cit-
rulline residue in position 6 [instead of (R)-homo-
citrulline], an (S)-leucine residue in position 7 [instead
of (S)-neoleucine], and an (R)-alanine residue in posi-
tion 10 [instead of the Me3El-Ala (El=C, Si, Ge)
COOH. We report here on the syntheses of the a-amino
acids rac-2–rac-6 and on the chromatographic resolu-
tions of rac-2 and rac-3 to yield the respective a-amino
acids (R)-2, (S)-2, (R)-3, and (S)-3. b-(Trimethylsi-
lyl)alanine (2) and b-(trimethylgermyl)alanine (3) are
sila- and germa-analogs of the non-proteinogenic a-
amino acid b-tert-butylalanine (1).
Furthermore, we report on the syntheses of the de-
capeptides 7–9 and their antagonistic potencies at the
human GnRH receptor. These peptides contain the
residues derived from (R)-1, (S)-2, and (S)-3]. The
pharmacological studies of 7–9 presented here were
carried out with special emphasis on C/Si/Ge
bioisosterism.
On the other hand, compounds 7–9 were also syn-
thesized as part of ASTA Medica’s search for potent
C/Si/Ge-analogous
D-configurated amino acid residues
Me3C-Ala, Me3Si-Ala, or Me3Ge-Ala in position 10 of
their backbone. The C/Si/Ge analogs 7–9 are struc-
turally related to the GnRH antagonist CetrorelixINN

M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
185
peptide analogs of CetrorelixINN and its derivative D-
63153. While CetrorelixINN (Cetrotide®) has already
been approved by EU authorities as the first GnRH
antagonist for marketing in controlled ovarian stimula-
tion for assisted reproduction (COS/ART), the de-
capeptide D-63153 is currently in advanced preclinical
development due to its favorable pharmacological and
physicochemical properties, such as a high water solu-
bility [17]. A high water solubility is of special impor-
tance for the development of GnRH antagonist depot
Table 2
Yields of the a-amino acids rac-2–rac-6 obtained by hydrolysis of the
respective a-amino acid esters rac-15, rac-16, and rac-25–rac-27 as
well as overall yields
Product Yield (educt)
(%)
Overall yield
(%) a
Overall yield (%) b
rac-2
rac-3
rac-4
rac-5
rac-6
84 (rac-15)
85 (rac-16)
63 (rac-25)
43 (rac-26)
42 (rac-27)
42
30
21
47
51
38
22
12
a Monoalkylation of the dihydropyrazine 10 (Method A); yields
relative to 10.
b Dialkylation of the dihydropyrazine 10 (Method B); yields rela-
tive to the two-fold molar amount of 10.
formulations for clinical use in hormone-dependent
cancer therapy [18]. The decapeptides 7–9 are deriva-
tives of D-63153 bearing a Me3El-Ala residue [El=C,
Si, Ge; derived from (R)-1, (S)-2, or (S)-3] in position
10 instead of an (R)-alanine residue.
2. Results and discussion
2.1. Syntheses of the h-amino acids
The silicon- and germanium-containing a-amino
acids rac-2 and rac-3 were prepared by three- (Method
A) or four-step (Method B) syntheses, starting from the
dihydropyrazine 10 (Scheme 1).
Metalation of the dihydropyrazine 10 with n-butyl-
lithium and subsequent treatment with ClCH2SiMe3
(11) or ClCH2GeMe3 (12) yielded mixtures of the
respective dihydropyrazines rac-13 and rac-14, along
with the non-reacted educt 10 (Method A). After treat-
ment of these mixtures (rac-13 and rac-14 not isolated
and characterized) with hydrochloric acid at 0°C and
subsequent separation of the byproduct glycine ethyl
ester by extraction, the respective a-amino acid ethyl
esters rac-15 and rac-16 were obtained (for the yields,
see Table 1). Hydrolysis of these esters in boiling hy-
drochloric acid and subsequent treatment of the result-
ing a-amino acid hydrochlorides with propylene oxide
finally yielded the a-amino acids rac-2 and rac-3 (for
the yields, see Table 2).
Scheme 1.
Table 1
Yields of the a-amino acid ethyl esters rac-15, rac-16, and rac-25–
rac-27 obtained by hydrolysis of the respective dihydropyrazines
10/rac-13, 10/rac-14, 10/rac-22, 10/rac-24, rac-13/17, rac-14/18, rac-
22/28, and rac-23/29
Product
Yield (educt) (%) a
Yield (educt) (%) b
The overall yields and the atom economy [19] for the
syntheses of rac-2 and rac-3 could be improved by
two-fold alkylation of the dihydropyrazine 10 (Method
B). After metalation of 10 with n-butyllithium and
subsequent treatment with ClCH2SiMe3 (11) or
ClCH2GeMe3 (12), this metalation/alkylation proce-
dure was repeated to yield the dihydropyrazines 17 and
18 (stereochemistry not analyzed), along with the re-
spective dihydropyrazines rac-13 and rac-14. Treatment
rac-15
rac-16
rac-25
rac-26
rac-27
60 (10/rac-13)
42 (10/rac-14)
35 (10/rac-22)
66 (rac-13/17)
71 (rac-14/18)
64 (rac-22/28)
60 (rac-23/29)
35 (10/rac-24)
a Monoalkylation of the dihydropyrazine 10 (Method A); yields
relative to 10.
b Dialkylation of the dihydropyrazine 10 (Method B); yields rela-
tive to the two-fold molar amount of 10.

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M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
of these mixtures (rac-13, rac-14, 17, and 18 not iso-
lated and characterized) with hydrochloric acid at 0°C
and separation of the byproduct glycine ethyl ester by
extraction gave the a-amino acid esters rac-15 and
rac-16 (for the yields, see Table 1), which were then
converted into the a-amino acids rac-2 and rac-3 (for
the yields, see Table 3) in analogy to Method A.
The structurally related a-amino acids rac-4–rac-6
were prepared according to Scheme 2. Thus, metalation
of the dihydropyrazine 10 with n-butyllithium and sub-
sequent treatment with ClCH2SiMe2Ph (19) or
ClCH2SiMe2CHꢀCH2 (21) yielded mixtures of the re-
spective dihydropyrazines rac-22 and rac-24, along
with the non-reacted educt 10 (Method A). Treatment
of these mixtures (rac-22 and rac-24 not isolated and
characterized) with hydrochloric acid at 0°C and subse-
quent separation of the byproduct glycine ethyl ester by
extraction gave the respective a-amino acid ethyl esters
rac-25 and rac-27 (for the yields, see Table 1). Attempts
to prepare the a-amino acids rac-4 and rac-6 by hydrol-
ysis of these esters in boiling hydrochloric acid failed:
under the conditions applied to the hydrolysis of rac-15
and rac-16, SiꢁC (SiꢁPh, SiꢁCHꢀCH2) bond cleavage
was observed. However, this problem was solved by
avoiding acidic conditions. Thus, treatment of the a-
amino acid esters rac-25 and rac-27 with lithium hy-
droxide (molar ratio 1:1) in a water–dioxane mixture,
followed by a special workup (including an Li+/H+
exchange; see Ref. [4]), yielded the respective a-amino
acids rac-4 and rac-6.
Table 3
Compound rac-4 was prepared alternatively by start-
ing with a two-fold alkylation of the dihydropyrazine
10 (Method B). The same procedure was also used for
the synthesis of the a-amino acid rac-5. Thus, after
deprotonation of 10 with n-butyllithium and subse-
quent treatment with ClCH2SiMe2Ph (19) or
ClCH2GeMe2Ph (20), this metalation/alkylation proce-
dure was repeated to yield the dihydropyrazines 28 and
29 (stereochemistry not analyzed), along with the re-
spective dihydropyrazines rac-22 and rac-23. Treatment
of these mixtures (rac-22, rac-23, 28, and 29 not iso-
lated and characterized) with hydrochloric acid at 0°C
and subsequent separation of the byproduct glycine
ethyl ester by extraction gave the respective a-amino
acid ethyl esters rac-25 and rac-26 (for the yields, see
Table 1). To avoid SiꢁC (SiꢁPh) and GeꢁC (GeꢁPh)
bond cleavage, the conversions of these a-amino acid
esters into the respective a-amino acids rac-4 and rac-5
were carried out in analogy to Method A by treatment
with lithium hydroxide in a water–dioxane mixture,
followed by a special workup.
Antagonistic potencies (IC50 values) of the decapeptides
CetrorelixINN, D-63153, and 7–9 at the human GnRH receptor a
Decapeptide
IC50 (nM)
CetrorelixINN
D-63153
7
8
9
0.28
0.26
14
27
15
a For details, see Section 2.
The a-amino acids rac-2–rac-6 were isolated as col-
orless crystalline solids, whereas rac-15, rac-16, and
rac-25–rac-27 were obtained as colorless liquids. The
identities of all compounds were established by elemen-
tal analyses (C, H, N), NMR-spectroscopic studies (1H,
13C, 29Si), and mass-spectrometric investigations.
2.2. Preparati6e liquid-chromatographic resolutions
The Si/Ge analogs rac-2 and rac-3 were resolved by
preparative liquid chromatography using CHIROBIO-
TIC T (glycopeptide Teicoplanin covalently linked to
spherical silica gel) as the chiral stationary phase and
methanol or methanol–water [90:10 (v/v)] as the elu-
ents [yields (relative to the racemic a-amino acid):
(R)-2, 36%; (S)-2, 23%; (R)-3, 32%; (S)-3, 21%]. The
enantiomeric purities (see Section 2.3) of the resolved
antipodes were as follows: (R)-2, 99% ee; (S)-2, 99% ee;
Scheme 2.

M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
187
enantiomers are as follows: (R)-2, 20.8 min; (S)-2, 22.1
min. The (R)- and (S)-enantiomers of 3 were separated
by analytical liquid chromatography (HPLC) using
CHIROBIOTIC T (glycopeptide Teicoplanin cova-
lently linked to spherical silica gel) as the chiral station-
ary phase (see Fig. 2). The retention times of the
enantiomers are as follows: (R)-3, 9.2 min; (S)-3, 11.2
min. The assignment of absolute configurations to the
antipodes of 2 and 3 was made by comparison with
authentic samples of (R)-2 [4] and (R)-3 [4].
2.4. Syntheses of the decapeptides
To incorporate the a-amino acids (R)-1, (S)-2, and
(S)-3 into the decapeptides 7–9, the N-(fluoren-9-
yl)methoxycarbonyl (N-Fmoc) derivatives of these
amino acids were prepared. The N-Fmoc derivatives
(R)-30, (S)-31, and (S)-32 were synthesized according
to Scheme 3 the treatment of (R)-1, (S)-2, or (S)-3 with
(fluoren-9-yl)methyl chloroformate (Fmoc-Cl) and were
isolated as colorless solids [yields: (R)-30, 80%; (S)-31,
68%; (S)-32, 59%].
The decapeptides 7–9 were prepared by solid-phase
syntheses, using standard methods [20] for the sequen-
tial synthesis of oligopeptides and starting with the
C-terminal attachment of (R)-30, (S)-31, or (S)-32. The
decapeptides 7–9 were purified by high-performance
liquid chromatography, and their identities were estab-
lished by mass-spectrometric studies (ESI MS and ESI
MS/MS).
Fig. 1. Quantitative analytical gas-chromatographic determination of
the molar ratio of the (R)- and (S)-enantiomers of the chemically
modified a-amino acid 2. Gas chromatograms: (a) 50:50 mixture of
(R)-2/(S)-2 obtained by hydrolysis of rac-15 (analysis of the crude
product); (b) enantiomerically pure (99% ee) isomer (R)-2 obtained
by preparative liquid-chromatographic separation; (c) enantiomeri-
cally pure (99% ee) isomer (S)-2 obtained by preparative liquid-chro-
matographic separation. For details, see Section 3.
(R)-3, 99% ee; (S)-3, 93% ee. To the best of our
knowledge, the resolutions of rac-2 and rac-3 are the
first examples of preparative liquid-chromatographic
separations of the antipodes of chiral silicon- and ger-
manium-containing a-amino acids. The enantiopure a-
amino acids (R)-2 and (R)-3 have already been
prepared by asymmetric syntheses, along with the
byproducts (S)-2 and (S)-3 [4]. The method described
in this paper (synthesis of compounds rac-2 and rac-3
and their chromatographic resolution) represents an
efficient alternative approach for the preparation of
(R)-2, (S)-2, (R)-3, and (S)-3.
2.5. Pharmacological characterization of the
decapeptides
The decapeptides 7–9 were studied in vitro in a
functional assay using a recombinant cell line express-
ing the human GnRH receptor. Triptorelin (D
-Trp6-
GnRH) served as the agonist in these studies.
Compounds 7–9 were found to be medium-potent
GnRH antagonists. As can be seen from Table 3, the
antagonistic potencies (IC50 values) of 7–9 are quite
similar, but significantly reduced compared with that of
CetrorelixINN and D-63153. The results obtained clearly
indicate strongly pronounced bioisosteric relationships
between the C/Si/Ge-analogous decapeptides 7–9. This
is in line with the findings of a series of further biolog-
ical studies with other C/Si/Ge-analogous drugs [1d].
2.3. Determination of the enantiomeric purities and
assignment of absolute configurations
To monitor the preparative liquid-chromatographic
resolutions of rac-2 and rac-3 and to determine the
enantiomeric purities of the antipodes of 2 and 3,
analytical chromatographic methods were developed.
The (R)- and (S)-enantiomers of 2 were separated, after
conversion into the respective N-(trifluoroaceto)amino
acid propyl esters (see Section 3), by analytical capillary
3. Experimental
3.1. Syntheses
3.1.1. General procedures
gas chromatography using a Chiralsil-
(Fig. 1). The retention times of the chemically modified
D
-Val column
Unless otherwise indicated, the reactions were carried
out under dry nitrogen. THF was dried and purified

188
M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
according to standard procedures and stored under
nitrogen. The 1H, 13C, and 29Si NMR spectra were
recorded at (room temperature) r.t. on a Bruker DRX-
300 (1H, 300.1 MHz; 13C, 75.5 MHz; 29Si, 59.6 MHz) or
Bruker AMX-400 NMR spectrometer (1H, 400.1 MHz;
13C, 100.6 MHz). Chemical shifts (ppm) were deter-
mined relative to internal CHCl3 (1H, l 7.24, solvent
CDCl3), CDCl3 (13C, l 77.05, solvent CDCl3), HDO
(1H, l 4.82, solvent D2O), and external Me4Si (13C, l 0,
solvent D2O; 29Si, l 0, solvents CDCl3 and D2O). The
1H spin systems of the a-amino acids and the a-amino
acid esters were analyzed by simulations using the
WIN-DAISY software package (version 4.0, Bruker). As-
signment of the 13C-NMR data was supported by
DEPT 135 experiments. Mass spectra were obtained
with a Varian MAT-711 (EI MS, 70 eV), Finnigan
MAT-8430 (EI MS, 70 eV; CI MS, NH3 as reactant
gas), Finnigan MAT-8200 (EI MS, 70 eV), or Thermo-
Quest TRIO 1000 mass spectrometer (EI MS, 70 eV).
1
The selected m/z values given refer to the isotopes H,
12C, 14N, 16O, 28Si, 35Cl, and 74Ge. IR spectra were
obtained with a Bruker Equinox 55 FT-IR spectrome-
ter. The ion exchanger (Aldrich, Art. 21,739-5) was
activated by washing with 0.5 M hydrochloric acid (50
ml) and water (3×50 ml).
3.1.2. (R)-2-Amino-4,4-dimethylpentanoic acid
[(R)-i-tert-butylalanine; (R)-1]
This compound was commercially available (Bachem
AG).
3.1.3. rac-2-Amino-3-(trimethylsilyl)propionic acid
[rac-i-(trimethylsilyl)alanine; rac-2]
A solution of rac-15 (13.5 g, 71.3 mmol) in hydro-
chloric acid (6 M, 200 ml) was heated under reflux for
1 h. After the mixture was cooled to r.t., the solvent
was removed in vacuo (0.1 mbar, 20°C) and the residue
dried at 40°C/0.01 mbar for 1 h and then dissolved in
EtOH (250 ml). After addition of propylene oxide (180
ml), the mixture was heated under reflux for 10 min.
The resulting precipitate was isolated by centrifugation,
washed with EtOH (2×25 ml) and Et2O (2×25 ml),
and then dried in vacuo (0.01 mbar, 40°C, 36 h) to give
rac-2 in 84% yield as a colorless crystalline solid (9.69
g, 60.1 mmol). The NMR and MS data of the product
were identical with those described for (R)-2 in Ref. [4].
Anal. Found: C, 44.2; H, 8.9; N, 8.6. Calc. for
C6H15NO2Si: C, 44.68; H, 9.37; N, 8.68%.
Fig. 2. Quantitative analytical liquid-chromatographic determination
of the molar ratio of the (R)- and (S)-enantiomers of the a-amino
acid 3. Liquid chromatograms: (a) 50:50 mixture of (R)-3/(S)-3
obtained by hydrolysis of rac-16 (analysis of the crude product); (b)
enantiomerically pure (99% ee) isomer (R)-3 obtained by preparative
liquid-chromatographic separation; (c) enantiomerically enriched
(93% ee) isomer (S)-3 obtained by preparative liquid-chromato-
graphic separation. For details, see Section 3.
3.1.4. (R)-2-Amino-3-(trimethylsilyl)propionic acid
[(R)-i-(trimethylsilyl)alanine; (R)-2]
This compound was obtained by preparative liquid-
chromatographic separation (see Section 3.1.27). The
IR, NMR, and MS data of the product were identical
with those already described for (R)-2 (prepared by an
alternative synthesis) in Ref. [4]. Anal. Found: C, 44.8;
H, 9.5; N, 8.7. Calc. for C6H15NO2Si: C, 44.68; H, 9.37;
N, 8.68%.
3.1.5. (S)-2-Amino-3-(trimethylsilyl)propionic acid
[(S)-i-(trimethylsilyl)alanine; (S)-2]
This compound was obtained by preparative liquid-
chromatographic separation (see Section 3.1.27). The
IR, NMR, and MS data of the product were identical
with those already described for (R)-2 (prepared by an
alternative synthesis) in Ref. [4]. Anal. Found: C, 44.3;
H, 9.3; N, 8.5. Calc. for C6H15NO2Si: C, 44.68; H, 9.37;
N, 8.68%.
Scheme 3.

M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
189
3.1.6. rac-2-Amino-3-(trimethylgermyl)propionic acid
[rac-i-(trimethylgermyl)alanine; rac-3]
under reduced pressure (0.1 mbar, 20°C) and the residue
dried in vacuo (0.01 mbar, 20°C, 1 h) and then dissolved
in EtOH (10 ml). After addition of propylene oxide (3
ml), the mixture was heated under reflux for 10 min. The
resulting precipitate was isolated by centrifugation,
washed with EtOH (2×5 ml) and Et2O (2×5 ml), and
then dried in vacuo (0.01 mbar, 40°C, 36 h) to give rac-4
in 63% yield as a colorless crystalline solid (377 mg, 1.69
mmol). The NMR and MS data of the product were
identical with those described for (R)-4 in Ref. [4]. Anal.
Found: C, 59.0; H, 7.5; N, 6.3. Calc. for C11H17NO2Si:
C, 59.16; H, 7.67; N, 6.27%.
This compound was prepared analogously to the
synthesis of rac-2 by heating a solution of rac-16 (3.53
g, 15.1 mmol) in hydrochloric acid (6 M, 70 ml). After
treatment of the solution of the residue (rac-3·HCl) in
EtOH (40 ml) with propylene oxide (10 ml), rac-3 was
isolated in 85% yield as a colorless crystalline solid (2.65
g, 12.9 mmol). The NMR and MS data of the product
were identical with those described for (R)-3 in Ref. [4].
Anal. Found: C, 35.2; H, 7.3; N, 6.7. Calc. for
C6H15GeNO2: C, 35.02; H, 7.35; N, 6.81%.
3.1.7. (R)-2-Amino-3-(trimethylgermyl)propionic acid
[(R)-i-(trimethylgermyl)alanine; (R)-3]
3.1.10. rac-2-Amino-3-[dimethyl(phenyl)germyl]-
propionic acid [rac-i-[dimethyl(phenyl)germyl]-
alanine; rac-5]
This compound was obtained by preparative liquid-
chromatographic separation (see Section 3.1.27). The
IR, NMR, and MS data of the product were identical
with those already described for (R)-3 (prepared by an
alternative synthesis) in Ref. [4]. Anal. Found: C, 35.2;
H, 7.5; N, 7.0. Calc. for C6H15GeNO2: C, 35.02; H, 7.35;
N, 6.81%.
This compound was prepared analogously to the
synthesis of rac-4 starting from rac-26 (408 mg, 1.38
mmol). The product was isolated in 43% yield as a
colorless crystalline solid (159 mg, 594 mmol). The
NMR and MS data of the product were identical with
those described for rac-5 in Ref. [4]. Anal. Found: C,
48.9; H, 6.2; N, 5.1. Calc. for C11H17GeNO2: C, 49.32;
H, 6.40; N, 5.23%.
3.1.8. (S)-2-Amino-3-(trimethylgermyl)propionic acid
[(S)-i-(trimethylgermyl)alanine; (S)-3]
This compound was obtained by preparative liquid-
chromatographic separation (see Section 3.1.27). The
IR, NMR, and MS data of the product were identical
with those already described for (R)-3 (prepared by an
alternative synthesis) in Ref. [4]. Anal. Found: C, 34.9;
H, 7.2; N, 6.9. Calc. for C6H15GeNO2: C, 35.02; H, 7.35;
N, 6.81%.
3.1.11. rac-2-Amino-3-[dimethyl(6inyl)silyl]propionic
acid [rac-i-[dimethyl(6inyl)silyl]alanine; rac-6]
This compound was prepared analogously to the
synthesis of rac-4 starting from rac-27 (634 mg, 2.52
mmol). The product was isolated in 42% yield as a
colorless crystalline solid (184 mg, 1.06 mmol). The
NMR and MS data of the product were identical with
those described for rac-6 in Ref. [4]. Anal. Found: C,
48.5; H, 8.6; N, 8.0. Calc. for C7H15NO2Si: C, 48.52; H,
8.72; N, 8.08%.
3.1.9. rac-2-Amino-3-[dimethyl(phenyl)silyl]propionic
acid [rac-i-[dimethyl(phenyl)silyl]alanine; rac-4]
Water (2.5 ml) and an aqueous lithium hydroxide
solution (1 M, 2.69 ml; 2.69 mmol LiOH) were added to
a solution of rac-25 (676 mg, 2.69 mmol) in dioxane (7
ml). After the mixture was stirred at r.t. for 16 h, the
solvent was removed in vacuo. The residue was washed
with n-pentane (10 ml) and then dried in vacuo (0.01
mbar, 20°C, 2 h) to give the lithium salt of rac-4 in
quantitative yield as a colorless solid (617 mg, 2.69
mmol). A mixture of this lithium salt (617 mg, 2.69
mmol) and an activated ion exchanger (11 g) in water
(35 ml) was shaken at r.t. for 30 min. After the pH value
was adjusted to pH 10–11 by the addition of an 0.1 M
aqueous diethylamine solution, the mixture was shaken
for a further 30 min. The ion exchanger was filtered off,
washed with water (20 ml), and then resuspended in 0.5
M hydrochloric acid (35 ml). After the mixture was
shaken at r.t. for 5 min, the ion exchanger was filtered
off and resuspended in 0.5 M hydrochloric acid (35 ml),
and the mixture was shaken once again at r.t. for 5 min.
After separation of the ion exchanger by filtration, the
solvent of the combined aqueous phases was removed
3.1.12. Acetyl-
phenylalanyl-
-homocitrullyl-neoleucyl-arginyl-prolyl-
alaninamide (Ac-
-Nal1-4-Cl- -Phe2- -Pal3-Ser4-N-
Me-Tyr5- -Hci6-Nle7-Arg8-Pro9- -Me3C-Ala10-NH2;
7), acetyl- -2-naphthylalanyl- -4-chlorophenylalanyl-
-3-pyridylalanyl-seryl-N-methyltyrosyl- -homocitrul-
lyl-neoleucyl-arginyl-prolyl- -(trimethylsilyl)-
alaninamide (Ac-
-Nal1-4-Cl- -Phe2- -Pal3-Ser4-N-
Me-Tyr5- -Hci6-Nle7-Arg8-Pro9- -Me3Si-Ala10-NH2;
8), and acetyl- -2-naphthylalanyl- -4-chloro-
phenylalanyl- -3-pyridylalanyl-seryl-N-methyltyrosyl-
-homocitrullyl-neoleucyl-arginyl-prolyl- -(trime-
-Phe2-
-Hci6-Nle7-Arg8-Pro9-
D
-2-naphthylalanyl-
-3-pyridylalanyl-seryl-N-methyltyrosyl-
-tert-butyl-
D-4-chloro-
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
D
thylgermyl)alaninamide (Ac-
D
-Nal1-4-Cl-
D
D-
Pal3-Ser4-N-Me-Tyr5-
D
D-
Me3Ge-Ala10-NH2; 9)
Compounds 7–9 were prepared by solid-phase syn-
thesis using a Labortec SP650 semiautomatic peptide
synthesizer. [(Fluoren-9-yl)methoxycarbonyl]-4-meth-

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M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
HNCO)++], 740 [42%, Y5¦+], 656 [11%, B+4 ], 569 [8%,
B+3 or Y¦4+], 456 [2%, Y¦3+].
oxy-4%-(g-carboxypropyloxy)benzhydrylamine
resin
with a capacity of 0.55 mmol g−1 (Fmoc-MBHA resin;
Bachem AG, D 1600) was used as polymeric support,
starting with 3.5 g of resin for the synthesis of 7 and 2.0
g of resin each for the synthesis of 8 and 9. The
sequential synthesis was performed as follows. The
respective N-Fmoc-protected a-amino acid (R)-30, (S)-
31, or (S)-32 [derived from (R)-1, (S)-2, and (S)-3]
(C-terminal) was covalently linked to the unprotected
MBHA resin by using a two-fold molar excess of both
diisopropylcarbodiimide (DIC) and 1-hydroxybenzotri-
azole (HOBt) in a mixture of CH2Cl2 and DMF [1:1
(v/v)]. Removal of the Fmoc group was performed with
a 20% solution of piperidine in DMF as standard
routine. Chain elongation according to the respective
amino acid sequence was accomplished with a two-mo-
lar excess of each N-Fmoc-amino acid, DIC, and HOBt
within 90 min of coupling time. Completion of acyla-
tion was checked via the chloroanil test. Cleavage of
the decapeptides from the polymeric support was ac-
complished by treatment with a mixture of AcOH–
2,2,2-trifluoroethanol–CH2Cl2 [1:1:8 (v/v/v)] at 40°C
for 2.5 h. The solvents were removed under reduced
pressure, and the residue was washed with Et2O. The
precipitate was filtered off and dried at r.t. for 12 h.
The decapeptides were purified by high-performance
liquid chromatography using a Nucleoprep RP-C-18
silica gel column. The experimental conditions were as
follows: HPLC pump, Shimadzu LC 8A; detector,
Shimadzu SPD 68; column (50 mm i.d.×250 mm);
eluent, water–MeCN–2,2,2-trifluoroacetic acid [A:
97:3:1 (v/v/v); B: 30:70:1 (v/v/v); 45% B“90% B within
50 min]; injection volume, 20 ml; flow rate, 60.0
ml min−1. After removal of the solvents under reduced
pressure, the isolated decapeptides were lyophilized.
The yields of the products (white fluffy residues) were
as follows: 7, 1.17 g; 8, 940 mg; 9, 1.33 g.
3.1.13. 3,6-Diethoxy-2,5-dihydropyrazine (10)
Synthesis as described in Ref. [21].
3.1.14. (Chloromethyl)trimethylsilane (11)
This compound was commercially available
(Aldrich).
3.1.15. (Chloromethyl)trimethylgermane (12)
This compound was synthesized from trichloro-
(chloromethyl)germane [22] according to Ref. [23].
3.1.16. rac-2-Amino-3-(trimethylsilyl)propionic acid
ethyl ester (rac-15)
Method A. A 1.6 M solution of n-butyllithium in
n-hexane (3.00 ml, 4.80 mmol n-BuLi) was added drop-
wise at −10°C within 10 min to a stirred solution of 10
(6.79 g, 39.9 mmol) in THF (50 ml). After the reaction
mixture was cooled to −70°C, a 1.6 M solution of
n-butyllithium in n-hexane (22.0 ml, 35.2 mmol n-BuLi)
was added dropwise over a period of 20 min. The
resulting mixture was stirred at −70°C for 15 min and
a solution of 11 (4.89 g, 39.9 mmol) in THF (10 ml)
was added dropwise at this temperature within 30 min.
After the mixture was stirred at −70°C for 2 h, it was
warmed to r.t. within 12 h, followed by addition of
Et2O (50 ml) and water (50 ml). The organic phase was
separated and the aqueous layer extracted with Et2O
(2×25 ml), and the combined organic extracts were
dried over anhydrous Na2SO4. The solvent was re-
moved under reduced pressure and the oily residue
distilled in a Kugelrohr apparatus (oven temperature
140°C, 0.01 mbar) to give a 23:77 mixture (8.70 g) of 10
and rac-13. Hydrochloric acid (3 M, 10 ml) was added
dropwise at 0°C within 15 min to a stirred solution of
10/rac-13 in EtOH (25 ml). After the mixture was
stirred at 0°C for 2 h, the solvent was removed in vacuo
(0.1 mbar, 20°C), the residue dissolved in CH2Cl2 (25
ml), and the resulting solution extracted with a satu-
rated aqueous Na2CO3 solution (2×25 ml). After the
organic layer was dried over anhydrous Na2SO4, the
solvent was removed under reduced pressure (rotary
evaporator) and the residue purified by distillation in a
Kugelrohr apparatus (oven temperature 80°C/0.01
mbar) to give rac-15 in 60% yield as a colorless liquid
(4.52 g, 23.9 mmol). The IR, NMR, and MS data of the
product were identical with those described for (R)-15
in Ref. [4]. Anal. Found: C, 50.5; H, 10.1; N, 7.4. Calc.
for C8H19NO2Si: C, 50.75; H, 10.12; N, 7.40%.
Data for 7. ESI MS: 1514 [9%, (M+H)+], 758
[100%, (M+2 H)++]. ESI MS/MS [758; (C76H104
-
ClN17O14+2 H)++]: 1273 [4%, B8+], 1117 [3%, B+7 ],
1004 [2%, B+6 ], 859 [10%, Y6¦+], 815 [48%, (B5−
H2O)+], 736 [100%, (M+2 H−HNCO)++], 682 [42%,
Y5¦+], 656 [12%, B4+], 398 [3%, Y3¦+].
Data for 8. ESI MS: 1530 [28%, (M+H)+], 765
[100%, (M+2 H)++]. ESI MS/MS [765; (C75H104
-
ClN17O14Si+2 H)++]: 1273 [6%, B8+], 1117 [4%, B+7 ],
1110 [1%, Y8¦+], 1004 [2%, B6+], 962 [2%, Y7¦+], 875
[12%, Y¦6+], 815 [52%, (B5−H2O)+], 744 [100%, (M+
2 H−HNCO)++], 698 [40%, Y5¦+], 656 [13%, B+4 ], 569
[2%, B+3 ], 527 [1%, Y4¦+], 412 [2%, Y3¦+].
Data for 9. ESI MS: 1576 [22%, (M+H)+], 789
Method B. A 1.6 M solution of n-butyllithium in
n-hexane (5.00 ml, 8.00 mmol n-BuLi) was added drop-
wise at −10°C within 10 min to a stirred solution of 10
(10.2 g, 59.9 mmol) in THF (250 ml). After the reaction
mixture was cooled to −70°C, a 1.6 M solution of
[100%, (M+2 H)++]. ESI MS/MS [787; (C75H104
-
Cl70GeN17O14+2 H)++]: 1273 [4%, B+8 ], 1117 [2%,
B7+], 1152 [1%, Y¦8+], 1004 [3%, B6+ or Y7¦+], 915 [12%,
Y6¦+], 815 [53%, (B5−H2O)+], 765 [100%, (M+2 H−

M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
191
n-butyllithium in n-hexane (32.5 ml, 52.0 mmol n-BuLi)
was added dropwise over a period of 20 min. The
resulting mixture was stirred at −70°C for 15 min and
a solution of 11 (7.35 g, 59.9 mmol) in THF (15 ml)
was added dropwise at this temperature within 30 min.
After the mixture was stirred at −70°C for 2 h, it was
warmed to r.t. within 12 h. The mixture was cooled
again to −70°C and then treated dropwise with a 1.6
M solution of n-butyllithium in n-hexane (37.5 ml, 60.0
mmol n-BuLi). After the mixture was stirred at −70°C
for 30 min, a solution of 11 (7.35 g, 59.9 mmol) in THF
(15 ml) was added dropwise at this temperature within
30 min. The resulting mixture was stirred at −70°C for
2 h and then warmed to r.t. within 12 h, followed by
the addition of Et2O (500 ml) and water (500 ml). The
organic phase was separated and the aqueous layer
extracted with Et2O (2×250 ml), and the combined
organic extracts were dried over anhydrous Na2SO4.
The solvent was removed under reduced pressure and
the oily residue distilled in a Kugelrohr apparatus (oven
temperature 180°C, 0.01 mbar) to give a 16:84 mixture
(16.6 g) of rac-13 and 17. Hydrochloric acid (3 M, 150
ml) was added dropwise at 0°C within 15 min to a
stirred solution of rac-13/17 in EtOH (100 ml). After
the mixture was stirred for 2 h at 0°C, the solvent was
removed in vacuo (0.1 mbar, 20°C), the residue dis-
solved in CH2Cl2 (150 ml), and the resulting solution
extracted with a saturated aqueous Na2CO3 solution
(75 ml). After the organic layer was dried over anhy-
drous Na2SO4, the solvent was removed under reduced
pressure (rotary evaporator) and the residue purified by
distillation in a Kugelrohr apparatus (oven temperature
80°C/0.01 mbar) to give rac-15 in 66% yield (relative to
the two-fold molar amount of 10) as a colorless liquid
(14.9 g, 78.7 mmol). The IR, NMR, and MS data of the
product were identical with those described for rac-15
in Ref. [4]. Anal. Found: C, 50.6; H, 10.0; N, 7.2. Calc.
for C8H19NO2Si: C, 50.75; H, 10.12; N, 7.40%.
(R)-16 in Ref. [4]. Anal. Found: C, 41.1; H, 8.2; N, 6.0.
Calc. for C8H19GeNO2: C, 41.09; H, 8.19; N, 5.99%.
Method B. This compound was prepared analogously
to the synthesis of rac-15 (Method B) by addition of a
1.6 M solution of n-butyllithium in n-hexane (7.38 ml,
11.8 mmol n-BuLi) to a solution of 10 (2.01 g, 11.8
mmol) in THF (50 ml), followed by treatment with a
solution of 12 (1.97 g, 11.8 mmol) in THF (15 ml).
After the metalation/alkylation procedure was repeated
[metalation with a 1.6 M solution of n-butyllithium in
n-hexane (7.38 ml, 11.8 mmol n-BuLi); alkylation with
a solution of 12 (1.97 g, 11.8 mmol) in THF (15 ml)],
the crude product was purified by Kugelrohr distillation
(oven temperature 190°C, 0.01 mbar) to give a 11:89
mixture (4.38 g) of rac-14 and 18. Hydrolysis of this
mixture was carried out by treatment of a solution of
rac-14/18 in EtOH (30 ml) with hydrochloric acid (3 M,
50 ml). After extractive separation of glycine ethyl
ester, rac-16 was isolated in 71% yield (relative to the
two-fold molar amount of 10) as a colorless liquid (3.91
g, 16.7 mmol). The IR, NMR, and MS data of the
product were identical with those described for (R)-16
in Ref. [4]. Anal. Found: C, 41.1; H, 8.2; N, 6.0. Calc.
for C8H19GeNO2: C, 41.09; H, 8.19; N, 5.99%.
3.1.18. (Chloromethyl)dimethyl(phenyl)silane (19)
This compound was commercially available
(Aldrich).
3.1.19. (Chloromethyl)dimethyl(phenyl)germane (20)
Synthesis as described in Ref. [4].
3.1.20. (Chloromethyl)dimethyl(6inyl)silane (21)
Synthesis as described in Ref. [24].
3.1.21. rac-2-Amino-3-[dimethyl(phenyl)silyl]propionic
acid ethyl ester (rac-25)
Method A. This compound was prepared analogously
to the synthesis of rac-15 (Method A) by addition of a
1.6 M solution of n-butyllithium in n-hexane (31.3 ml,
50.1 mmol n-BuLi) to a solution of 10 (8.51 g, 50.0
mmol) in THF (25 ml), followed by treatment with a
solution of 19 (9.24 g, 50.0 mmol) in THF (10 ml). The
crude product was purified by Kugelrohr distillation
(oven temperature 160°C, 0.01 mbar) to give a 45:55
mixture of 10 and rac-22. Hydrolysis of this mixture
was carried out by treatment of a solution of 10/rac-22
in EtOH (25 ml) with hydrochloric acid (3 M, 10 ml).
After extractive separation of glycine ethyl ester, rac-25
was isolated in 35% yield as a colorless liquid (4.35 g,
17.3 mmol). The IR, NMR, and MS data of the
product were identical with those described for (R)-25
in Ref. [4]. Anal. Found: C, 62.0; H, 8.4; N, 5.6. Calc.
for C13H21NO2Si: C, 62.11; H, 8.42; N, 5.57%.
3.1.17. rac-2-Amino-3-(trimethylgermyl)propionic acid
ethyl ester (rac-16)
Method A. This compound was prepared analogously
to the synthesis of rac-15 (Method A) by addition of a
1.6 M solution of n-butyllithium in n-hexane (13.3 ml,
21.3 mmol n-BuLi) to a solution of 10 (3.61 g, 21.2
mmol) in THF (25 ml), followed by treatment with a
solution of 12 (3.54 g, 21.2 mmol) in THF (10 ml). The
crude product was purified by Kugelrohr distillation
(oven temperature 150°C, 0.01 mbar) to give a 14:86
mixture (5.89 g) of 10 and rac-14. Hydrolysis of this
mixture was carried out by treatment of a solution of
10/rac-14 in EtOH (25 ml) with hydrochloric acid (3 M,
10 ml). After extractive separation of glycine ethyl
ester, rac-16 was isolated in 42% yield as a colorless
liquid (2.06 g, 8.81 mmol). The IR, NMR, and MS data
of the product were identical with those described for
Method B. This compound was prepared analogously
to the synthesis of rac-15 (Method B) by addition of a

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M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
1.6 M solution of n-butyllithium in n-hexane (22.1 ml,
35.4 mmol n-BuLi) to a solution of 10 (6.01 g, 35.3
mmol) in THF (150 ml), followed by treatment with a
solution of 19 (6.52 g, 35.3 mmol) in THF (15 ml).
After the metalation/alkylation procedure was repeated
[metalation with a 1.6 M solution of n-butyllithium in
n-hexane (22.1 ml, 35.4 mmol n-BuLi); alkylation with
a solution of 19 (6.52 g, 35.3 mmol) in THF (15 ml)],
the crude product was purified by Kugelrohr distillation
(oven temperature 210°C, 0.01 mbar) to give a 33:67
mixture of rac-22 and 28. Hydrolysis of this mixture
was carried out by treatment of a solution of rac-22/28
in EtOH (100 ml) with hydrochloric acid (3 M, 150 ml).
After extractive separation of glycine ethyl ester, rac-25
was isolated in 64% yield (relative to the two-fold molar
amount of 10) as a colorless liquid (11.4 g, 45.3 mmol).
The IR, NMR, and MS data of the product were
identical with those described for (R)-25 in Ref. [4].
Anal. Found: C, 61.8; H, 8.2; N, 5.5. Calc. for
C13H21NO2Si: C, 62.11; H, 8.42; N, 5.57%.
(1.51 g) of 10 and rac-24. Hydrolysis of this mixture
was carried out by treatment of a solution of 10/rac-24
in EtOH (25 ml) with HCl (3 M, 10 ml). After extrac-
tive separation of glycine ethyl ester, rac-27 was iso-
lated in 35% yield as a colorless liquid (548 mg, 2.72
mmol). The IR, NMR, and MS data of the product
were identical with those described for (R)-27 in Ref.
[4]. Anal. Found: C, 53.0; H, 9.3; N, 7.1. Calc. for
C9H19NO2Si: C, 53.69; H, 9.51; N, 6.96%.
3.1.24. (R)-N-[(Fluoren-9-yl)methoxycarbonyl]-
2-amino-4,4-dimethylpentanoic acid [(R)-30]
A solution of (fluoren-9-yl)methyl chloroformate
(3.93 g, 15.2 mmol) in dioxane (40 ml) was added
dropwise at 0°C within 30 min to a stirred solution of
(R)-1 (2.00 g, 13.8 mmol) and Na2CO3 (2.93 g, 27.6
mmol) in water–dioxane [3:5 (v/v)] (80 ml). After the
mixture was stirred at 10°C for 1 h, it was warmed to
r.t. within 12 h. The solvents were removed under
reduced pressure (0.1 mbar, 20°C) and the residue
acidified with hydrochloric acid (6 M, pH 2). The
aqueous solution was extracted with EtOAc (3×5 ml),
and the combined organic extracts were dried over
anhydrous Na2SO4. After the solution was concen-
trated under reduced pressure, the resulting precipitate
was isolated by filtration, washed with petroleum ether
(40–60°C) (2×10 ml) and then dried in vacuo (0.1
mbar, 20°C, 2 h) to give (R)-30 in 80% yield as a
colorless solid (4.04 g, 11.0 mmol). The IR, NMR, and
MS data of the product were identical with those
described for (S)-30 in Ref. [4]. Anal. Found: C, 71.3;
H, 6.7; N, 3.8. Calc. for C22H25NO4: C, 71.91; H, 6.86;
N, 3.81%.
3.1.22. rac-2-Amino-3-[dimethyl(phenyl)germyl]-
propionic acid ethyl ester (rac-26)
This compound was prepared analogously to the
synthesis of rac-15 (Method B) by addition of a 1.6 M
solution of n-butyllithium in n-hexane (2.34 ml, 3.74
mmol n-BuLi) to a solution of 10 (637 mg, 3.74 mmol)
in THF (25 ml), followed by treatment with a solution
of 20 (857 mg, 3.74 mmol) in THF (15 ml). After the
metalation/alkylation procedure was repeated [metala-
tion with a 1.6 M solution of n-butyllithium in n-hex-
ane (2.34 ml, 3.74 mmol n-BuLi); alkylation with a
solution of 20 (857 mg, 3.74 mmol) in THF (15 ml)],
the crude product was purified by Kugelrohr distillation
(oven temperature 250°C, 0.01 mbar) to give a 25:75
mixture (1.62 g) of rac-23 and 29. Hydrolysis of this
mixture was carried out by treatment of a solution of
rac-23/29 in EtOH (10 ml) with hydrochloric acid (3 M,
15 ml). After extractive separation of glycine ethyl
ester, rac-26 was isolated in 60% yield (relative to the
two-fold molar amount of 10) as a colorless liquid (1.32
g, 4.46 mmol). The IR, NMR, and MS data of the
product were identical with those described for (R)-26
in Ref. [4]. Anal. Found: C, 52.8; H, 7.1; N, 4.4. Calc.
for C13H21GeNO2: C, 52.76; H, 7.15; N, 4.73%.
3.1.25. (S)-N-[(Fluoren-9-yl)methoxycarbonyl]-2-
amino-3-(trimethylsilyl)propionic acid [(S)-31]
This compound was prepared analogously to the
synthesis of (R)-30 starting from (S)-2 (1.04 g, 6.45
mmol). The product (S)-31 was isolated in 68% yield as
a colorless solid (1.68 g, 4.38 mmol). The IR, NMR,
and MS data of (S)-31 were identical with those de-
scribed for (R)-31 in Ref. [4]. Anal. Found: C, 65.5; H,
6.6; N, 3.6. Calc. for C21H25NO4Si: C, 65.77; H, 6.57;
N, 3.65%.
3.1.23. rac-2-Amino-3-[dimethyl(6inyl)silyl]propionic
acid ethyl ester (rac-27)
3.1.26. (S)-N-[(Fluoren-9-yl)methoxycarbonyl]-2-
amino-3-(trimethylgermyl)propionic acid [(S)-32]
This compound was prepared analogously to the
synthesis of (R)-30 starting from (S)-3 (973 mg, 4.73
mmol). The product (S)-32 was isolated in 59% yield as
a colorless solid (1.20 g, 2.80 mmol). The IR, NMR,
and MS data of (S)-32 were identical with those de-
scribed for (R)-32 in Ref. [4]. Anal. Found: C, 58.8; H,
5.8; N, 3.3. Calc. for C21H25GeNO4: C, 58.93; H, 5.89;
N, 3.27%.
This compound was prepared analogously to the
synthesis of rac-15 (Method A) by addition of a 1.6 M
solution of n-butyllithium in n-hexane (4.92 ml, 7.87
mmol n-BuLi) to a solution of 10 (1.34 g, 7.87 mmol) in
THF (25 ml), followed by treatment with a solution of
21 (1.06 g, 7.87 mmol) in THF (10 ml). The crude
product was purified by Kugelrohr distillation (oven
temperature 150°C, 0.01 mbar) to give a 20:80 mixture

M. Merget et al. / Journal of Organometallic Chemistry 628 (2001) 183–194
193
3.1.27. Preparati6e liquid-chromatographic resolution of
rac-2 and rac-3
0.2 ml-sample of this solution was injected into the gas
chromatograph.
The (R)- and (S)-enantiomers of 2 and 3 were sepa-
rated by liquid-chromatographic resolution of rac-2
and rac-3, respectively, using CHIROBIOTIC T (gly-
copeptide Teicoplanin covalently linked to spherical
silica gel, 40 mm; Advanced Separation Technologies,
Inc.) as the chiral stationary phase. The experimental
conditions were as follows: HPLC pump, Shimadzu
LC8-A; detector, Knauer VW monitor; integrator, Shi-
madzu C-R3A; column (50 mm i.d.×500 mm for
rac-2; 50 mm i.d.×250 mm for rac-3), Merck Super-
formance; eluent, MeOH for rac-2 and MeOH–water
[90:10 (v/v)] for rac-3; detection, 215 nm; temperature,
21–23°C; injection volume, 150 ml (2 mg of the sample
material in 1 ml of the eluent) for rac-2 and 30 ml (2
mg of the sample material in 1 ml of the eluent) for
rac-3. The solvents of the respective fractions obtained
[(R)-2, first fraction, (S)-2, second fraction; (R)-3, first
fraction, (S)-3, second fraction] were removed immedi-
ately after the chromatographic separation (rotary
evaporator, 35°C, 100 mbar), and the respective
residues were combined and then stored at r.t. The
yields and enantiomeric purities (for the analytic meth-
ods, see below) of the separated enantiomers (colorless
solids) were as follows: (R)-2, 36%, 99% ee; (S)-2, 23%,
99% ee; (R)-3, 32%, 99% ee; (S)-3, 21%, 93% ee.
3.1.30. Determination of the enantiomeric purities of
(R)-3 and (S)-3 by liquid chromatography
The (R)- and (S)-enantiomers of 3 were separated by
liquid chromatography using CHIROBIOTIC T (gly-
copeptide Teicoplanin covalently linked to spherical
silica gel, 5 mm; Advanced Separation Technologies,
Inc.) as the chiral stationary phase. The experimental
conditions were as follows: HPLC pump, Shimadzu
LC6-A; detector, Soma S-3702 UV–vis; integrator, Shi-
madzu C-R3A; column, 4.6 mm i.d.×250 mm, Merck
Superformance; eluent, MeOH–water [90:10 (v/v)];
flow rate, 600 ml min−1; pressure, 58 bar; detection, 215
nm; temperature, 20°C; injection volume, 20 ml (2 mg of
the sample material dissolved in 1 ml of the eluent).
The retention times were as follows: (R)-3, 9.2 min;
(S)-3, 11.2 min.
3.1.31. Determination of the antagonistic potencies of
the decapeptides 7–9 at the human GnRH receptor
The antagonistic potencies of the decapeptides 7–9 at
the human GnRH receptor were studied in vitro in a
functional assay as described in Ref. [4].
Acknowledgements
3.1.28. Determination of the enantiomeric purities of
(R)-2 and (S)-2 by capillary gas chromatography
The (R)- and (S)-enantiomers of 2 were separated,
after chemical transformation (see Section 3.1.29), by
capillary gas chromatography. The experimental condi-
tions were as follows: gas chromatograph, HP 6890;
column (0.25 mm i.d.×25 m, film thickness 0.12 mm),
Chrompack 7496; carrier gas, hydrogen; pressure, 0.55
bar; temperature program, 50°C (1 min) to 105°C (0
min) with 3°C min−1 to 200°C (10 min) with
4°C min−1; injector temperature, 250°C; split, 1:100;
detector, FID; detector temperature, 250°C. The reten-
tion times of the derivatives of (R)-2 and (S)-2 were as
follows: (R)-2, 20.8 min; (S)-2, 22.1 min.
Financial support of this work by Degussa AG and
the Fonds der Chemischen Industrie is gratefully
acknowledged. M.M. thanks the Fond der Chemischen
Industrie for a postgraduate fellowship.
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