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Volatile methyl esters of medium chain length from the bacterium Chitinophaga Fx7914

DOI: 10.1002/cbdv.201000190

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Chemistry and biodiversity p. 2228 - 2253 (2010)

Update date:2022-07-29

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Authors:

Nawrath, ThorbenNawrath, Thorben

Gerth, KlausGerth, Klaus

Mueller, RolfMueller, Rolf

Schulz, StefanSchulz, Stefan

Read Full Text PDF DownLoad Join now for total 90,000,000 free articles

Article abstract of DOI:10.1002/cbdv.201000190

The analysis of the volatiles released by the novel bacterial isolate Chitinophaga Fx7914 revealed the presence of ca. 200 compounds including different methyl esters. These esters comprise monomethyl- and dimethyl-branched, saturated, and unsaturated fatty acid methyl esters that have not been described as bacterial volatiles before. More than 30 esters of medium C-chain length were identified, which belong to five main classes, methyl (S)-2-methylalkanoates (class A), methyl (S)-2,(ω-1)-dimethylalkanoates (class B), methyl 2,(ω-2)-dimethylalkanoates (class C), methyl (E)-2-methylalk-2-enoates (class D), and methyl (E)-2,(ω-1)-dimethylalk-2- enoates (class E). The structures of the compounds were verified by GC/MS analysis and synthesis of the target compounds as methyl (S)-2-methyloctanoate (28), methyl (S)-2,7-dimethyloctanoate ((S)-43), methyl 2,6-dimethyloctanoate (49), methyl (E)-2-methylnon-2-enoate (20a), and methyl (E)-2,7-dimethyloct-2- enoate (41a). Furthermore, the natural saturated 2-methyl-branched methyl esters showed (S)-configuration as confirmed by GC/MS experiments using chiral phases. Additionally, the biosynthetic pathway leading to the methyl esters was investigated by feeding experiments with labeled precursors. The Me group at C(2) is introduced by propanoate incorporation, while the methyl ester is formed from the respective carboxylic acid by a methyltransferase using S-adenosylmethionine (SAM).

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Full text of DOI:10.1002/cbdv.201000190

2228  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
Volatile Methyl Esters of Medium Chain Length from the Bacterium  
Chitinophaga Fx7914  
by Thorben Nawratha), Klaus Gerthb), Rolf Mꢀllerb)c), and Stefan Schulz*a)  
a) Institut fꢀr Organische Chemie, Technische Universitꢁt Braunschweig, Hagenring 30,  
D-38106 Braunschweig (phone: þ49-531-3917353; e-mail: stefan.schulz@tu-bs.de)  
b) Helmholtz-Zentrum fꢀr Infektionsforschung, Inhoffenstrasse 7, D-38124 Braunschweig  
c) Institut fꢀr Pharmazeutische Biotechnologie, Universitꢁt des Saarlandes, Gebꢁude 2.3,  
D-66123 Saarbrꢀcken  
The analysis of the volatiles released by the novel bacterial isolate Chitinophaga Fx7914 revealed the  
presence of ca. 200 compounds including different methyl esters. These esters comprise monomethyl- and  
dimethyl-branched, saturated, and unsaturated fatty acid methyl esters that have not been described as  
bacterial volatiles before. More than 30 esters of medium C-chain length were identified, which belong to  
five main classes, methyl (S)-2-methylalkanoates (class A), methyl (S)-2,(wꢀ1)-dimethylalkanoates  
(class B), methyl 2,(wꢀ2)-dimethylalkanoates (class C), methyl (E)-2-methylalk-2-enoates (class D),  
and methyl (E)-2,(wꢀ1)-dimethylalk-2-enoates (class E). The structures of the compounds were  
verified by GC/MS analysis and synthesis of the target compounds as methyl (S)-2-methyloctanoate (28),  
methyl (S)-2,7-dimethyloctanoate ((S)-43), methyl 2,6-dimethyloctanoate (49), methyl (E)-2-methyl-  
non-2-enoate (20a), and methyl (E)-2,7-dimethyloct-2-enoate (41a). Furthermore, the natural saturated  
2-methyl-branched methyl esters showed (S)-configuration as confirmed by GC/MS experiments using  
chiral phases. Additionally, the biosynthetic pathway leading to the methyl esters was investigated by  
feeding experiments with labeled precursors. The Me group at C(2) is introduced by propanoate  
incorporation, while the methyl ester is formed from the respective carboxylic acid by a methyltransfer-  
ase using S-adenosylmethionine (SAM).  
Introduction. – Fatty acids are one of the ubiquitous building blocks of biological  
systems. The typical chain length varies in bacteria between 14 and 20, with the most  
common fatty acids containing 16 or 18 C-atoms. In addition to these compounds, many  
bacteria also produce methyl-branched fatty acids, carrying an additional Me group  
often in the iso- or anteiso-position [1]. These acids are usually conjugated to other  
compounds forming the various lipid classes of living organisms. In bacteria, a second  
group of acids can be produced, short-chain, often amino-acid derived, volatile acids  
with up to six C-atoms [2]. Fatty acids with a chain length between these two classes are  
known, but are encountered relatively rarely in bacteria, often present only in low  
amounts compared to the more common acids with a longer C-chain.  
While short-chain fatty acids are volatile and have a distinct odor, the long-chain  
fatty acids possess a much lower vapor pressure. Some bacteria increase the volatility  
by transformation of the fatty acids into the methyl ester, making the compound also  
unavailable for other biosynthetic transformations.  
Methyl esters of intermediate chain length have only rarely been reported from  
bacteria. Methyl 5-methylhexanoate and methyl 9-methyldecanoate have been  
ꢂ 2010 Verlag Helvetica Chimica Acta AG, Zꢀrich  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2229  
detected in volatiles released by Stigmatella aurantiaca [3], while methyl 4-methyl-  
pentanoate is produced by different actinomycetes [4]. Furthermore, methyl octanoate  
was found in the odor of different Pecorino cheeses, most likely originating from  
bacteria [5].  
Chitinophaga belongs to the sphingobacteria and is a gliding bacterium not well  
investigated for the production of secondary metabolites. The antibiotic elansolide is  
the only natural compound known to be produced by these bacteria [6]. We became  
interested in the volatiles released by Chitinophaga Fx7914 which proved to be very  
complex. Beside several other components like sulfur compounds, amino acid  
derivatives, and diterpenes, a complex set of saturated and unsaturated fatty acid  
methyl esters of medium chain length was identified, all of them carrying a Me branch  
at C(2). Many of these compounds have not been reported from bacteria or other  
natural sources, beside a few exceptions [7–10]. The identification of these esters by  
GC and GC/MS, the syntheses of representative examples as well as the determination  
of the absolute configuration by stereoselective synthesis, followed by GC/MS on chiral  
phases, will be presented here. Furthermore, some aspects of the biosynthesis of these  
compounds were probed by feeding experiments using deuterium (2H1)-labeled  
precursors. Finally, the bouquet of volatiles released by Chitinophaga Fx7914 is  
described.  
Results and Discussion. – Liquid cultures of Chitinophaga Fx7914 were analyzed  
using the closed-loop-stripping methodology (CLSA) adopted to microbiological work  
as described in [11][12]. The volatiles emitted by the bacteria were trapped on  
activated charcoal. Elution with CH2Cl2 furnished an extract that was analyzed by GC/  
MS. Chitinophaga cultures release a large variety of compounds which were identified  
by comparison of mass spectra and GC retention indices with library data and with  
synthetic standards (Fig. 1 and Table 1).  
Fig. 1. Total ion chromatogram of a headspace extract of a liquid culture of Chitinophaga Fx7914  
Characteristic compounds, repeatedly detected in large amounts in different  
experiments, were 2-methyl- and 3-methylbutanoic acid (1 and 2, resp.), 2-methylte-  
trahydrothiophenone (3), as well as the corresponding alcohol 4, 2-phenylethanol (5),  
methyl phenylacetate (6), sesquiterpenes of the cadinane family such as g-cadinene (7)  
and a-cadinol (8), several alcohols, ketones, and lactones, as well as diterpenoids of  
unknown structure. The biosynthesis of 3 has recently been investigated by us [13].  
2230  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
Table 1. Volatiles Released by Chitinophaga Fx7914  
Ia)  
Entry  
Compound  
Ident.b)  
Conc.c)  
1
2
3
4
5
6
7
8
9
Methyl 2-methylpropionate  
4-Methylpentan-2-one  
Methyl 3-methylbutanoate  
3-Hydroxypentan-2-one  
2-Hydroxypentan-3-one  
5-Methylhexan-3-one  
4-Hydroxy-4-methylpentan-2-one  
2-Hydroxy-4-methylpentan-3-one  
Ethylbenzene  
MSd)  
MS  
MS  
synf)  
syn  
MS  
MS, RIg)  
MS  
xxxe)  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xxx  
xi)  
x
809  
816  
839  
846  
853  
860  
862  
868  
870  
877  
892  
898  
903  
908  
910  
913  
914  
916  
923  
931  
948  
953  
957  
958  
962  
964  
968  
976  
983  
990  
999  
1000  
1011  
1013  
1013  
1018  
1020  
1025  
1027  
1030  
1032  
1035  
1039  
1044  
1050  
1054  
MS, RI  
10  
11  
12  
13  
14  
15  
16  
17  
18  
19  
20  
21  
22  
23  
24  
25  
26  
27  
28  
29  
30  
31  
32  
33  
34  
35  
36  
37  
38  
39  
40  
41  
42  
43  
44  
45  
46  
47  
48  
49  
Unknown 55, 73, 98h)  
m-Xylene  
syn  
Methyl 2-methylbut-2-enoate  
2-Methoxy-1-methylethyl acetate  
o-Xylene  
2-Hydroxyhexan-3-one  
3-Methylbutanoic acid (2)  
2-Methylbutanoic acid (1)  
2-Butoxyethanol  
2,5-Dimethylpyrazine  
Butano-4-lactone  
But-2-eno-4-lactone  
MS, RI  
MS  
syn  
MS  
syn  
syn  
MS  
syn  
syn  
MS  
syn  
x
x
x
x
Dimethylsulfone  
Hexane-2,5-dione  
Unknown 41, 73, 100  
Pentano-4-lactone  
MS  
xxx  
x
syn  
MS, RI  
syn  
6-Methylheptan-2-one  
xxj)  
xx  
Benzaldehydek)  
Unknown 82, 132  
Dimethyl trisulfide  
Unknown 45, 88  
Aniline  
2-Methyltetrahydrothiophen-3-one (3)  
2,4,6-Trimethylpyridinek)  
Methyl 4-methylhexanoate  
3-Methylpenten-2-eno-4-lactone  
2,4-Dimethylpent-2-eno-4-lactone  
Unknown 43, 111, 126  
Unknown 43, 69, 113  
Mesitylene  
xx  
xx  
x
x
xxx  
xxx  
xxx  
xxx  
x
x
x
x
x
x
xxx  
x
MS, RI  
syn  
syn  
MS  
MS, RI  
MS  
MS  
MS, RI  
Unknown 43, 83, 98, 101  
Unknown 68, 100  
2-Methyltetrahydrothiophen-3-ol (4)  
syn  
MS  
syn  
syn  
2-Ethylhexan-1-olk)  
Benzyl alcohol  
xx  
2-Methyltetrahydrothiophen-3-ol (4)  
Unknown 43, 109  
xxx  
xxx  
x
x
x
Methyl 2-ethylhexanoatek)  
7-Methyloctan-3-one  
Unknown 66, 132  
MS  
MS, RI  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2231  
Table 1 (cont.)  
Entry  
Compound  
Ia)  
Ident.b)  
Conc.c)  
50  
51  
52  
53  
54  
55  
56  
57  
58  
59  
60  
61  
62  
63  
64  
65  
66  
67  
68  
69  
70  
71  
72  
73  
74  
75  
76  
77  
78  
79  
80  
81  
82  
83  
84  
85  
86  
87  
88  
89  
90  
91  
92  
93  
94  
95  
96  
97  
98  
99  
Unknown 84, 129  
1-Phenylethanol  
Acetophenone  
N-Methylaniline  
Methyl 2-methylheptanoate (A1)  
2-Ethyl-4,6-dimethylpyridinek)  
Methylthiobenzene  
2-Phenylpropan-2-ol  
o-Guaicol  
Unknown 71, 95, 110, 128  
Unknown 43, 57, 83  
2-Phenylethanol (5)  
Heptano-4-lactone  
Methylundecene  
8-Methylnonan-2-one  
Methyl 2-methyloctanoate (A2)  
Unknown 43, 77, 105  
Unknown 91, 119, 134  
Methyl 2-phenylacetate (6)  
Decanal  
1056  
1059  
1062  
1062  
1064  
1067  
1079  
1082  
1086  
1103  
1106  
1109  
1150  
1154  
1156  
1161  
1164  
1168  
1175  
1204  
1215  
1224  
1226  
1248  
1251  
1255  
1257  
1260  
1269  
1271  
1277  
1279  
1281  
1286  
1300  
1311  
1324  
1328  
1348  
1353  
1356  
1358  
1360  
1373  
1389  
1394  
1400  
1408  
1417  
1423  
xx  
xx  
xxx  
xxx  
xxx  
x
x
xxx  
x
x
x
xx  
x
x
x
xx  
x
x
xxx  
x
xxx  
xxx  
x
x
x
syn  
syn  
syn  
syn  
MS  
MS  
MS  
MS  
syn  
MS, RI  
MS  
MS, RI  
syn  
syn  
syn  
MS, RI  
syn  
syn  
Benzothiazole  
Methyl 2,7-dimethyloctanoate (B1)  
Methyl 2,6-dimethyloctanoate (C1)  
Methyl (E)-2-methyloct-2-enoate (D1)  
9-Methyldecan-3-one  
Octano-4-lactone  
Benzyl propanoate  
Methyl 2-methylnonanoate (A3)  
Unknown 58, 87, 95  
Undecan-6-one  
Unknown 77, 106, 135, 159  
4-(Methylsulfanyl)thiophenol  
Nonano-5-lactone  
Ethyl 4-methylbenzoate  
Tridecane  
Methyl (E)-2,7-dimethyloct-2-enoate (E1)  
Methyl geranate  
Methyl 3-methylalkanoate  
Methyl (E)-2-methylnon-2-enoate (D2)  
Unknown 111, 184  
Methyl 2-methyldecenoate  
Unknown 55, 70, 84, 124, 142  
Methyl 2-methyldecanoate (A4)  
2-Vinylnaphthalenek)  
Dodecan-3-one  
Dodecan-2-one  
Tetradecane  
Methyl (E)-2,8-dimethylnon-2-enoate (E2)  
Methyl 2,9-dimethyldecanoate  
Methyl 2,9-dimethyldecenoate (B2)  
syn  
MS, RI  
MS, RI  
MS  
x
x
x
x
syn  
MS, RI  
x
xxx  
xxx  
x
x
x
xxx  
x
x
xx  
x
x
x
xx  
x
x
x
x
x
x
xxx  
MS  
MS, RI  
MS  
MS, RI  
syn  
MS, RI  
MS  
syn  
MS  
syn  
MS  
MS, RI  
MS, RI  
MS, RI  
MS, RI  
MS  
MS, RI  
2232  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
Table 1 (cont.)  
Entry  
Compound  
Ia)  
Ident.b)  
Conc.c)  
100  
101  
102  
103  
104  
105  
106  
107  
108  
109  
110  
111  
112  
113  
114  
115  
116  
117  
118  
119  
120  
121  
122  
123  
124  
125  
126  
127  
128  
129  
130  
131  
132  
133  
134  
135  
136  
137  
138  
139  
140  
141  
142  
143  
144  
145  
146  
147  
148  
Methyl 2,8-dimethyldecanoate (C2)  
1429  
1440  
1448  
1453  
1460  
1474  
1493  
1499  
1510  
1512  
1517  
1521  
1523  
1526  
1547  
1552  
1559  
1563  
1567  
1571  
1573  
1576  
1597  
1600  
1600  
1609  
1613  
1618  
1621  
1626  
1629  
1638  
1640  
1644  
1647  
1652  
1659  
1662  
1664  
1677  
1699  
1707  
1710  
1717  
1733  
1747  
1750  
1753  
1759  
MS, RI  
MS  
syn  
MS  
syn  
MS, RI  
MS, RI  
MS, RI  
MS, RI  
MS, RI  
x
x
xx  
xx  
x
x
x
x
xxx  
xxx  
x
x
x
x
x
xx  
xx  
x
1,2-Dihydro-2,2,4-trimethylquinolinek)  
Methyl (E)-2-methyldec-2-enoate (D3)  
Methyl 2-methylundecenoate  
Methyl 2-methylundecanoate (A5)  
Dodecan-1-ol  
Decano-5-lactone  
Tridecan-2-ol  
Methyl (E)-2,9-dimethyldec-2-enoate (E3)  
g-Cadinene (7)  
Sesquiterpene  
Sesquiterpene  
Sesquiterpene  
Methyl 3-methylalkanoate  
(E)-Methyl 2-methylundec-2-enoate (D4)  
Unknown 153, 237  
Methyl 2-methyldodecanoate (A6)  
Nerolidol  
12-Methyltridecan-2-ol  
Undecano-4-lactone  
Sesquiterpene  
MS  
syn  
syn  
MS, RI  
MS, RI  
MS, RI  
x
x
x
x
Tridecan-1-ol  
Unknown 41, 73, 88, 243  
Sesquiterpene  
MS, RI  
x
xx  
xx  
x
x
x
xxx  
x
x
xx  
xx  
x
xxx  
x
xx  
x
xx  
xxx  
x
xxx  
xxx  
xx  
x
xxx  
xx  
x
Hexadecane  
MS, RI  
MS, RI  
MS  
Methyl (E)-2,10-dimethylundec-2-enoate (E4)  
Methyl 2,11-dimethyldodacenoate  
Sesquiterpene  
Methyl 2,11-dimethyldodecanoate (B3)  
1-Epicubenol  
Methyl 2,10-dimethyldodecanoate (C3)  
t-Cadinol  
Sesquiterpene  
syn  
MS, RI  
MS, RI  
MS, RI  
d-Cadinol  
MS, RI  
syn  
MS, RI  
syn  
Methyl (E)-2-methyldodec-2-enoate (D5)  
a-Cadinol (8)  
Methyl 2-methyltridecanoate (A7)  
Unknown 88, 101, 256 (ester)  
13-Methyltetradecan-3-ol  
Tetradecan-1-ol  
MS, RI  
MS, RI  
MS  
MS  
syn  
MS  
MS  
syn  
MS  
Methyl alkanoate  
2-Ethylhexyl benzoatek)  
Methyl (E)-2,11-dimethyldodec-2-enoate (E5)  
Methyl alkenoate  
Methyl 2-methyltetradecenoate  
Methyl (E)-2-methyltridec-2-enoate (D6)  
Methyl 2-methyltetradecenoate  
Unknown 105, 126  
Methyl 2-methyltetradecanoate (A8)  
syn  
xx  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2233  
Table 1 (cont.)  
Entry  
Compound  
Ia)  
Ident.b)  
MS  
Conc.c)  
149  
150  
151  
152  
153  
154  
155  
156  
157  
158  
159  
160  
161  
162  
163  
164  
165  
166  
167  
168  
169  
170  
171  
172  
173  
174  
175  
176  
177  
178  
179  
180  
181  
182  
183  
184  
185  
186  
187  
188  
189  
190  
191  
192  
193  
194  
195  
196  
Alcohol  
Methyl farnesoate  
1779  
x
x
1784  
1791  
1795  
1800  
1805  
1811  
1814  
1822  
1829  
1833  
1845  
1848  
1855  
1858  
1859  
1863  
1869  
1881  
1889  
1894  
1900  
1905  
1906  
1911  
1914  
1918  
1922  
1929  
1935  
1935  
1941  
1944  
1949  
1951  
1957  
1966  
1971  
1980  
1995  
2003  
2058  
2066  
2071  
2079  
2115  
2125  
2133  
Diterpene  
Methyl alkenoate  
Octadecane  
Diterpene  
Methyl 2,13-dimethyltetradecenoate  
Diterpene  
Methyl 2,13-dimethyltetradecanoate (B4)  
Methyl 2,12-dimethyltetradecanoate (C4)  
Methyl alkanoate  
Hexahydrofarnesylacetone  
Methyl (E)-2-methyltetradec-2-enoate (D7)  
Diterpene  
Diterpene  
Methyl 2-methylpentadecanoate (A8)  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Nonadecane  
Diterpene  
Diterpene  
Methyl (E)-2,13-dimethyltetradec-2-enoate (E6)  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Methyl ester  
Diterpene  
Diterpene  
Methyl (E)-2-methylpentadec-2-enoate (D8)  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
xxx  
xx  
x
xxx  
xxx  
xxx  
xxx  
x
MS  
MS, RI  
MS  
MS, RI  
MS, RI  
MS  
syn  
syn  
x
xx  
xxx  
x
x
x
syn  
x
xxx  
xxx  
xxx  
x
x
xx  
xx  
xxx  
xx  
xxx  
xxx  
xx  
x
MS, RI  
xxx  
xxx  
x
x
x
xxx  
xx  
xx  
xx  
xx  
xxx  
xx  
xx  
xxx  
xxx  
xxx  
xxx  
syn  
2234  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
Table 1 (cont.)  
Entry  
Compound  
Ia)  
Ident.b)  
Conc.c)  
197  
198  
199  
200  
Diterpene  
Diterpene  
Diterpene  
Diterpene  
2141  
2153  
2173  
2186  
xxx  
xx  
xxx  
xxx  
a) I¼GC Retention index. b) Ident.¼Identification method. c) Conc.¼concentration. d) MS¼Mass  
spectrum. e) xxx¼Main compound (> 0.1%). f) syn: Synthetic compound. g) RI¼Retention index.  
h) Numbers indicate characteristic ions in the mass spectra of unknown compounds. i) x¼Trace  
compound (< 0.04% of total area of the TIC). j) xx¼Minor compound (0.04–0.1%). k) Medium  
constituent.  
In addition to these compounds, a series of esters was detected featuring  
characteristic ions at m/z 88 and 101 (Fig. 2). The ion corresponding to m/z 88 is  
generated by a McLafferty rearrangement of the ester function, while the ion ascribed  
to m/z 101 is caused by a primary H shift from C(6) to the CO group, followed by H-  
transfer and cleavage of the C(3)ꢀC(4) bond [14]. Characteristic for ethyl esters,  
usually associated with the ions corresponding to m/z 88 and 101, are those at m/z 73  
and [Mꢀ45]þ that arise by cleavage next to the CO group. The latter ions were  
missing; instead, m/z 59 and [Mꢀ31]þ were present, indicating the presence of a  
methyl ester functional group. In this case, peaks at m/z 88 and 101 indicate the  
presence of a Me branch at C(2), as shown in Scheme 1. These methyl 2-  
methylalkanoates were accompanied by compounds which exhibit a similar mass  
spectrum but a molecular-ion 2 amu lower than the parent compound, indicating the  
presence of a C¼C bond equivalent. A derivatization of the extract with dimethyl  
disulfide (MeSSMe) to locate the position of the C¼C bond in the chain furnished no  
derivatized esters. This behavior strongly suggested the presence of a C¼C bond at  
C(2), because a CO-conjugated C¼C bond does not react with MeSSMe due to its  
lower reactivity [15–17]. The occurrence of ions with peaks at m/z 88 and 101 in the  
unsaturated methyl esters, at first hand surprising when a C¼C bond is located at C(2),  
might be explained by the migration of the C¼C bond out of conjugation before  
fragmentation [18]. A diagnostic ion peak of considerable intensity in the spectra of  
some unsaturated esters is at m/z 127. Its formation can be explained by the mechanism  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2235  
Fig. 2. Mass spectra of a) methyl 2-methylnonanoate (29; A3); b) methyl 2,7-dimethyloctanoate (43; B1);  
c) methyl 2,7-dimethylnonanoate (49; C1); d) methyl 2-methylnon-2-enoate (20a; D2); and e) methyl 2,7-  
dimethyloct-2-enoate (41a; E1)  
Scheme 1. Characteristic Mass-Spectral Fragmentations of Saturated and Unsaturated Methyl Esters  
2236  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
shown in Scheme 1 [19]. Transfer of a H-atom of C(6) to the CO group is followed by  
loss of the g-alkyl group leading to an extended conjugated C¼C bond system. This  
reaction can only occur in a,b-unsaturated methyl esters, pointing again to a C¼C bond  
at C(2).  
More than 30 different methyl esters containing between 9 to 17 C-atoms were  
detected in the natural samples (Tables 1 and 2). The determination of the GC  
retention indices (RIs) revealed the presence of five different subtypes of these esters  
[20]. The saturated methyl esters had RIs ending around 60, 22, and 28 (classes A, B,  
and C), while the unsaturated ones exhibited values around 48 and 10 (class D and E).  
The RIs of aliphatic compounds with a terminal functional group (FG) can be  
calculated using an empirical RI model developed by us and used successfully in several  
projects [3][21–24]. The functional group (FG) increment for methyl esters is 331,  
calculated from the retention index of 1331 of methyl decanoate. The FG of a Me group  
in a-position to a CO group was determined to be 30 [25]. The compound class A can  
thus be identified to constitute methyl 2-methylalkanoates, exemplified by identifica-  
tion of compound A3, the most prominent member of this class. The molecular-ion  
peak at m/z 186 in the mass spectrum (Fig. 2,a) indicates a methyl ester with ten C-  
atoms in the acid part. The fragmentation pattern establishes the position of a Me  
group at C(2). The calculated retention index RIcalc for methyl 2-methylnonanoate  
(A3) is therefore 900þ331þ30¼1261, in good agreement with the observed value of  
1259 (Table 2) of the synthetic material (see below). The class-B compound B1 (mass  
spectrum Fig. 2,b) has the same molecular mass as A3, but is eluted earlier in GC. The  
RI of 1224 indicates an additional Me branch in the chain. By use of the known  
increments for a Me branch at various positions along the chain, a wꢀ1 Me group  
seems to be most plausible [21]. The value RIcalc ¼800þ331þ30þ60¼1221 is very  
close to the observed value. The second best position would be wꢀ3 with RIcalc ¼800þ  
331þ30þ56¼1217. The mass spectra of the class-B compounds were very similar to  
those of the class A compounds, but showed a small intensity increase in the [Mꢀ43]þ  
ion, typical for iso-esters [26]. Methyl esters branched at wꢀ3 also show a [Mꢀ43]þ  
ion, but with higher intensity [26]. In addition to the classes A and B, the class C esters  
proved to have a very similar mass spectrum (Fig. 2,c), also with a molecular-ion peak  
at m/z 186 in case of C1. The class C esters are eluted directly after the corresponding  
class B esters, pointing to a second Me branch in a different position than wꢀ1. Only  
the anteiso-esters show a higher RI value compared to the iso-esters [21]. Therefore,  
the second Me branch in the class-C compounds is located at the wꢀ2 position. The  
corresponding calculated value for C1 is RIcalc ¼800þ331þ30þ73¼1234 which is  
close to the observed value of 1226. The mass spectra of the class-C esters (Fig. 2,c)  
also showed an enhanced formation of [Mꢀ29]þ which can be explained the loss of the  
Et group at the end of the chain. The unsaturated esters of class D and E showed a RI  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2237  
Table 2. Different Methyl Esters from Chitinophaga Fx7914  
b
c
d
Compound  
Peaka)  
RInat  
)
RIcalc  
)
RIexp  
)
Methyl 2-methylheptanoate (27)  
Methyl 2-methyloctanoate (28)  
Methyl 2-methylnonanoate (29)  
Methyl 2-methyldecanoate (30)  
Methyl 2-methylundecanoate (31)  
Methyl 2-methyldodecanoate (32)  
Methyl 2-methyltridecanoate (33)  
Methyl 2-methyltetradecanoate (34)  
Methyl 2-methylpentadecanoate (35)  
Methyl 2,7-dimethyloctanoate (43)  
Methyl 2,9-dimethyldecanoate  
Methyl 2,11-dimethyldodecanoate (44)  
Methyl 2,13-dimethyltetradecanoate  
Methyl 2,6-dimethyloctanoate (49)  
Methyl 2,8-dimethyldecanoate  
A1  
A2  
A3  
A4  
A5  
A6  
A7  
A8  
A9  
B1  
B2  
B3  
B4  
C1  
C2  
C3  
C4  
D1  
D2  
D3  
D4  
D5  
D6  
D7  
D8  
E1  
E2  
E3  
E4  
E5  
E6  
1064  
1161  
1259  
1360  
1460  
1559  
1659  
1759  
1859  
1224  
1423  
1621  
1822  
1226  
1429  
1629  
1829  
1248  
1348  
1448  
1547  
1647  
1747  
1848  
1949  
1311  
1408  
1510  
1609  
1710  
1912  
1061  
1161  
1261  
1361  
1461  
1561  
1661  
1761  
1861  
1221  
1421  
1621  
1821  
1234  
1434  
1634  
1834  
1251  
1351  
1451  
1551  
1651  
1751  
1851  
1951  
1311  
1411  
1511  
1611  
1711  
1911  
1060  
1161  
1258  
1359  
1459  
1559  
1660  
1759  
1859  
1223  
1620  
1229  
Methyl 2,10-dimethyldodecanoate  
Methyl 2,12-dimethyltetradecanoate  
Methyl (E)-2-methyloct-2-enoate (19a)  
Methyl (E)-2-methylnon-2-enoate (20a)  
Methyl (E)-2-methyldec-2-enoate (21a)  
Methyl (E)-2-methylundec-2-enoate (22a)  
Methyl (E)-2-methyldodec-2-enoate (23a)  
Methyl (E)-2-methyltridec-2-enoate (24a)  
Methyl (E)-2-methyltetradec-2-enoate (25a)  
Methyl (E)-2-methylpentadec-2-enoate (26a)  
Methyl (E)-2,7-dimethyloct-2-enoate (41a)  
Methyl (E)-2,8-dimethylnon-2-enoate  
Methyl (E)-2,9-dimethyldec-2-enoate  
Methyl (E)-2,10-dimethylundec-2-enoate  
Methyl (E)-2,11-dimethyldodec-2-enoate (42a)  
Methyl (E)-2,13-dimethyltetradec-2-enoate  
1245  
1346  
1446  
1547  
1648  
1746  
1848  
1949  
1313  
1710  
a) Peak: see Fig. 1. b) RInat: Retention index of the naturally occurring ester. c) RIcalc: Calculated  
retention index. d) RIexp: Retention index of the synthetic standard.  
value 90 units higher than that of the respective saturated compounds of class A and B.  
This retention behavior is typical for a,b-unsaturated CO compounds and point  
together with the MS data to a C¼C bond at C(2) (Fig. 2,d and e). In conclusion, class-  
D and -E compounds were suggested to be analogs of the class A and B compounds  
with an additional C¼C bond at C(2). Besides these esters, several other methyl esters  
of low abundance occurred in the headspace extract, but their concentration was to low  
to confirm their structure (Table 1).  
Several of the methyl esters of classes A–E were then synthesized to verify the  
proposed structures and to clarify the configuration of the C¼C bond in the  
unsaturated esters.  
Synthesis of Methyl Esters. Different aldehydes 9–17, ranging from pentanal (9) to  
tridecanal (17), were transformed by a Horner–Wadsworth–Emmons reaction with  
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CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
methyl 2-(dimethoxyphosphoryl)propanoate to the unsaturated methyl esters 18a/  
18b–26a/26b, respectively, as diastereoisomeric mixtures in varying ratios (Scheme 2)  
[27]. NMR Analysis verified the C¼C bond configuration of the product diaster-  
eoisomers, and GC/MS comparison with the natural esters established that the class-D  
esters are constituted of methyl (E)-2-methylalk-2-enoates.  
Scheme 2. Synthesis of Unsaturated and Saturated Methyl 2-Methylalkanoates  
a) NaH, methyl 2-(dimethoxyphosphoryl)propanoate; 30–78%. b) H2, PtO2; 40–100%.  
In a final reaction step, the diastereoisomer mixtures of the unsaturated esters were  
hydrogenated in MeOH with PtO2 as catalyst, giving racemic mixtures of the  
corresponding saturated mono-Me-branched methyl esters 27–35 (Scheme 2) [28].  
Comparison of the synthetic material with the natural extract proved that the class-A  
esters were indeed methyl 2-methylalkanoates.  
The iso-esters of classes B and E were synthesized as shown in Scheme 3. Isobutyl  
bromide (36) was transformed into 5-methylhexan-1-ol (37) or 9-methyldecan-1-ol  
(38) by a Li2CuCl4-mediated chain elongation with 3-bromopropan-1-ol or 6-  
bromohexan-1-ol, respectively [29]. The alcohols 37 and 38 were then oxidized with  
PCC to furnish 5-methylhexanal (39) and 9-methyldecanal (40) [30]. The trans-  
formation to the unsaturated esters as just described afforded methyl 2,7-dimethyloct-  
2-enoate (41a and 41b) and 2,11-dimethyldodec-2-enoate (42a and 42b), respectively,  
as mixtures of diastereoisomers. After separation of the isomers, the (E)-isomers  
showed the same mass spectrum and RIs as the corresponding volatiles from  
Chitinophaga Fx7914, verifying the suggested structures for the class-E compounds.  
Scheme 3. Synthesis of Iso-Branched Unsaturated and Saturated Methyl 2-Methylalkanoates  
a) Mg, bromoalcohol, Li2CuCl4; 85% (37), 68% (38). b) Pyridinium chlorochromate (PCC); 69% (39),  
86% (40). c) NaH, methyl 2-(dimethoxyphosphoryl)propanoate; 32% (41), 44% (42). d) H2, PtO2; 66%  
(43), 61% (44).  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2239  
Finally, hydrogenation of 41 or 42 provided racemic methyl 2,7-dimethyloctanoate  
(43) and methyl 2,11-dimethyldodecanoate (44), respectively (Scheme 3), confirming  
the proposed structure for the class-B compounds of Chitinophaga Fx7914.  
The synthesis of the anteiso-esters of class C started from 4-methylhexanoic acid  
(45) that was reduced with LiAlH4 to give 4-methylhexan-1-ol (46; Scheme 4) [22].  
Following oxidation with PCC afforded the corresponding aldehyde 47 that was  
transformed as described into methyl (E)- and (Z)-2,6-dimethyloct-2-enoate (48)  
[27][30]. In a final reaction, 48 was hydrogenated in MeOH using PtO2 as catalyst [28]  
to yield methyl 2,6-dimethyloctanoate (49) which proved identical to the natural  
compound C1.  
Scheme 4. Synthesis of Anteiso-Branched Saturated Methyl 2-Methylalkanoates  
a) LiAlH4; 75% (46). b) PCC (47). c) NaH, methyl 2-(dimethoxyphosphoryl)propanoate; 38% (over  
two steps) (48). d) H2, PtO2; quant. (49).  
Although not all esters listed in Table 2, i.e., A1–E6, were synthesized, the  
structures of B2, B4, C2, C3, C4, E2, E3, E4, and E6 were confirmed by their MS data  
and RIs, showing the expected class values.  
Determination of the Absolute Configuration. The mono- and dimethyl-branched  
saturated methyl esters both contained a stereogenic center at C(2). Therefore, a  
representative of class-A and -B esters was synthesized stereoselectively for the  
determination of their configuration by GC/MS on chiral phases. Methyl (S)-2-  
methyloctanoate ((S)-28) was synthesized as depicted in Scheme 5.  
Octanoyl chloride (50) was transformed with the Evans chiral auxiliary (4S)-4-  
benzyl-1,3-oxazolidin-2-one to the corresponding derivative 51 [31]. The following  
stereoselective methylation with MeI under basic conditions furnished compound 52 in  
a high diastereoisomeric ratio of 98 :2, which was purified by column chromatography  
to give only one diastereoisomer [32]. The chiral auxiliary was cleaved off with H2O2  
and LiOH, providing (S)-2-methyloctanoic acid (53) [32]. Finally, an acid-catalyzed  
esterification with methanol afforded enantiomerically pure methyl (S)-2-methylocta-  
noate ((S)-28) [33].  
The methyl ester (S)-28 and rac-28 were used for GC analyses using a chiral b-Dex  
225 GC phase (Fig. 3,a). The analyses confirmed the ee value of synthetic (S)-28  
to be 100%, and proved that the natural compound also shows this configuration.  
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CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
Scheme 5. Stereoselective Synthesis of Methyl (S)-2-Methyloctanoate ((S)-28)  
a) BuLi, (4S)-4-benzyl-1,3-oxazolidin-2-one; 84%. b) Sodium hexamethyldisilazane (NaHMDS), MeI;  
66%. c) H2O2, LiOH; 100%. d) MeOH, HCl; 92%.  
Furthermore, natural 28 is produced as single enantiomer. This results suggest that all  
esters of class A possess an (S)-configuration.  
Fig. 3. Determination of the absolute configuration of a) methyl (S)-2-methyloctanoate ((S)-28) and  
b) methyl 2,7-dimethyloctanoate ((S)-43) on a chiral b-Dex 225 phase. a) 1) Synthetic rac-28. 2) Synthetic  
(S)-28. 3) Coinjection of rac- and (S)-28. 4) Natural extract of Chitinophaga Fx7914. 5) Coinjection of  
rac-28 and natural extract. 6) Coinjection of (S)-28 and natural sample. b) 1) Synthetic rac-43. 2)  
Synthetic (S)-43. 3) Coinjection of rac- and (S)-43. 4) Natural extract of Chitinophaga Fx7914. 5)  
Coinjection of rac-43 and natural extract. 6) Coinjection of (S)-43 and natural sample.  
To extend these investigations to the dimethyl-branched saturated methyl esters of  
class B, methyl (S)-2,7-dimethyloctanoate ((S)-43) was prepared (Scheme 6). Pre-  
viously prepared 7-methyloctanoic acid (54) was converted to the corresponding acid  
chloride 55 that was transformed to the Evans oxazolidinone 56 [17][31][34].  
Methylation led to compound 57 with a diastereoisomeric ratio of 96 :4 that gave  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2241  
Scheme 6. Stereoselective Synthesis of Methyl (S)-2,7-Dimethyloctanoate ((S)-43)  
a) Oxalyl chloride; 80%. b) BuLi, (4S)-4-benzyl-1,3-oxazolidin-2-one; 48%. c) NaHMDS, MeI; 46%. d)  
H2O2, LiOH; 89%. e) MeOH, 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride (EDC),  
4-(dimethylamino)pyridine (DMAP); 66%.  
enantiomerically pure 57 after purification [32]. The following cleavage of the  
oxazolidinone produced acid 58 [32]. Finally, EDC-assisted esterification with MeOH  
furnished the target compound (S)-43 that was used for GC/MS analyses on a chiral b-  
Dex 225 column (Fig. 3,b) [35].  
The natural methyl 2,7-dimethyloctanoate ((S)-43) is also produced exclusively as  
pure (S)-enantiomer. Most likely, all class-B compounds possess (S)-configuration as  
the class A esters do. The configuration of the two stereogenic centers in the class-C  
compounds was not determined because of their low natural abundance.  
Biosynthesis of the Methyl Esters. It is well known that a (wꢀ1)-Me group in fatty  
acids containing an even number of C-atoms in the chain originates from 3-  
methylbutyryl-CoA, derived from the amino acid leucine, while, in case of an odd  
number of C-atoms, isobutyryl-CoA, derived from valine, serves as starter compound.  
Similarly, the (wꢀ2)-Me group occurs only in fatty acids with an even number of C-  
atoms in the chain and is formed from 2-methylbutyryl-CoA derived from isoleucine  
[24][36][37]. The Me groups in the esters of Chitinophaga located at the end of the  
chain are most likely biosynthetically formed via these pathways. In contrast, the  
biosynthetic origin of the Me group at C(2) was unknown. Possibilities are either the  
addition of the Me group to a C¼C bond via (S)-adenosylmethionine (SAM) or  
incorporation of methylmalonate in the carboxylic acid chain extension by fatty acid  
synthases [27][38][39].  
Liquid cultures of Chitinophaga Fx7914 were supplemented with either  
[1’-2H3]methionine as precursor of SAM or with sodium [1-13C]propanoate as precursor  
of methylmalonate to investigate whether one of the proposed biosynthetic pathways is  
operating. The emitted headspace volatiles were then analyzed by GC/MS [11][12].  
The labeled precursors were incorporated in both experiments. Upon feeding of  
[1’-2H3]methionine the molecular ion of methyl 2,7-dimethyloctanoate (43) shifted  
from m/z 186 to 189, while the characteristic ions at m/z 88 and 101 were replaced by  
the corresponding values at m/z 91 and 104 respectively (Fig. 4,a), indicating the  
formation of [2H3]methyl 2,7-dimethyloctanoate ([2H3]-43) (incorporation rate  
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CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
>95%). Furthermore, the [Mꢀ31]þ ion was not present, but instead a [Mꢀ34]þ  
2
fragment appeared. This shift indicated incorporation of the H label only into the  
MeO group of the ester, but not into the Me group at C(2).  
Fig. 4. Mass spectra of methyl 2,7-dimethyloctanoate (43) from Chitinophaga Fx7914 after feeding of  
a) [1’,1’,1’-2H3]methionine and b) sodium [1-13C]propanoate  
The feeding of sodium [1-13C]propanoate furnished labeled esters (incorporation  
rate 20%). Contrary to 2H isotopomers that have shorter GC retention times compared  
to their parent compounds [3][22][24], 13C-labelled compounds show no retention-  
time shift. In the mass spectrum of labeled methyl 2,7-dimethyloctanoate (43) an  
intensity increase of the ion peaks at m/z 89, 102, 144, 156, 172, and 187, compared to  
the spectrum of unlabelled 43, was observed, indicating the incorporation of labeled  
propanoate into 43 (Fig. 4,b). The mass spectrum showed the position of the label at  
C(1), thus proving that the Me branch at C(2) originates from methylmalonate and not  
from SAM.  
On the basis of the known fatty acid biosynthesis and the labeling experiments, a  
biosynthetic pathway for the formation of the different methyl-ester classes can be  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2243  
postulated (Scheme 7). Starting either with acetyl-CoA, propionyl-CoA (class A and  
D), 3-methylbutyryl-CoA derived from leucine (class B and E), 2-methylbutyryl-CoA  
derived from isoleucine (class C), or isobutyryl-CoA derived from valine (class E),  
methyl-branched saturated and unsaturated methyl esters are synthesized.  
Scheme 7. Biosynthetic Pathway for the Formation of the Different Methyl Ester Classes  
The chain is extended from the starter molecules via malonyl-SCoA [2][22][40].  
The odd or even chain length of the esters can be explained by use of the various  
starters, but an a-oxidation process, leading to a loss of one C-atom during or after  
chain extension, cannot be ruled out [24][37]. The feeding experiments with  
[13C]propanoate did not clarify whether a propionyl starter is really used, because  
the few ions which contain the alkyl end of the esters are of low abundance in the mass  
spectra. A definite proof for a propionyl-CoA starter would be the enhanced intensity  
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CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
of the [Mþ2]þ ion, requiring incorporation of two propanoate units. The abundance of  
such ions was too low for a definite conclusion.  
In the final chain-extension step methylmalonyl-CoA derived from propanoate is  
added to the chain, leading to the Me branch at C(2). Hydrolysis from the acyl carrier  
protein (ACP) before reduction and final methylation with SAM leads to the  
unsaturated esters, while reduction of the C¼C bond with NADPH, followed by  
hydrolysis from ACP and SAM mediated methylation, affords the saturated esters. The  
methylation of free fatty acids with SAM, established here by the labeling experiments,  
has been described before by the action of a methyltransferase in Mycobacterium phlei  
and the myxobacterium S. aurantiaca, and also in the rat hypothalamus [41–43].  
As discussed in the introduction, methyl esters of medium chain length have only  
occasionally been reported from bacteria. Most of the compounds described in Table 2  
have not been reported before from nature. The related unsaturated fatty acids are also  
rare. In mycobacteria, 2-methylalk-2-enoates of longer chain length have been found as  
constituents of complex lipids [44][45]. In contrast, simple 2-methylalkanoic acids are  
found in bacteria, plants, and animals. The parent acids of the 2,X-dimethyl-branched  
esters reported here are also not known from nature, except for 2,7-dimethyloctanoic  
acid which occurs bound to glycerol in epidermal glands of the reptile, Sphenodon  
punctatus [46].  
In summary, the identification, biosynthesis, and synthesis of novel Me-branched  
aliphatic methyl alkanoates and methyl alkenoates of medium chain length is de-  
scribed. Similar compounds have not been reported before as a major volatile class  
released by bacteria. Why these esters are formed is unknown, but they might function  
as signals or spacing compounds, inhibiting microbial growth next to Chitinophaga cells,  
thus contributing to the success of these gliding bacteria.  
We thank the Deutsche Forschungsgemeinschaft for financial support (DFG Schu 984-6).  
Experimental Part  
General. Chemicals were purchased from Sigma–Aldrich (Germany), Fluka (Switzerland), Acros  
(Belgium), Merck (Germany), Cambridge Isotope Laboratories (USA), and Deutero (Germany). All  
solvents were purified by distillation and dried according to standard methods. TLC: 0.2-mm pre-coated  
plastic sheets (Polygram Sil G/UV254; Macherey-Nagel). Column chromatography (CC): Merck silica gel  
60 (SiO2; 70–200 mesh); solvent mixtures as used for the determination of the Rf values in TLC;  
compounds were detected using a molybdatophosphoric acid soln. (5% in EtOH) or a KMnO4 soln.,  
followed by heat-gun treatment. NMR Spectra: Bruker AVII-300, DRX-400, or AVIII-400 spectrom-  
eters; chemical shifts d in ppm relative to Me4Si as internal standard and coupling constants J in Hz.  
Media and Growth Conditions. T2 Medium of the following composition was used to study the  
production of metabolites and to perform feeding experiments under defined conditions: glutamine,  
0.2% (w/w); MgSO4 ꢁ7 H2O, 0.02% (w/w); Fe-EDTA, 8 mg/l; MnSO4, 5 mg/l; ZnCl2, 0.25 mg/l; HEPES,  
12 g/l; XAD 16 adsorber resin (Rohm and Haas), 2%; pH adjusted to 7.0. 5 ml of a 20% (w/w) autoclaved  
sucrose soln. and 1 ml of a 7% (w/w) KH2PO4 autoclaved soln. were added to 100 ml of the above  
medium. Liquid cultures were started in 106 ml T2 medium in 250-ml Erlenmeyer flasks inoculated with  
2% of a preculture and incubated on a rotary shaker at 308 for 4 d. The feeding experiments were carried  
out by the addition of [2H3]methionine (50 mg, 5.5 mm final concentration) or sodium [1-13C]propanoate  
(200 mg, 31.2 mm final concentration) to well grown cultures of Chitinophaga Fx7914.  
Sampling of Volatiles. Volatile org. compounds emitted by bacterial cultures were collected by the  
closed-loop stripping analysis (CLSA) using the mentioned 250-ml Erlenmeyer flasks [11][12]. The  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2245  
volatiles were adsorbed on charcoal (Chromtech; Precision Charcoal Filter, 5 mg) for 24 h and extracted  
with 30 ml of CH2Cl2. These extracts were immediately analyzed by GC/MS and then stored at ꢀ308.  
GC/MS Analysis. GC/MS Analyses were carried out on a HP-6890 GC system connected to a HP-  
5973 mass-selective detector fitted with a BPX5 fused-silica cap. column (25 m, 0.22 mm i.d., 0.25 mm  
film; SGE, Australia) or on an Agilent 7890A GC system connected to an Agilent 5975C inert mass  
detector fitted with a HP-5MS fused silica cap. column (30 m, 0.25 mm i.d., 0.25 mm film; J&W Scientific,  
USA). Conditions for the HP-6890/HP-5973 system were: inlet pressure: 77.1 kPa, 23.3 ml He minꢀ1  
;
injection volume: 1 ml; transfer line: 3008; electron energy: 70 eV. GC Program: 5 min at 508, increasing  
with 108 minꢀ1 to 3208, operated in either split or splitless mode (60 s valve time); for the Agilent 7890A/  
Agilent 5975C system, the following conditions were used: inlet pressure: 77.1 kPa, 23.3 ml He minꢀ1  
;
injection volume: 2 ml; transfer line: 3008; electron energy: 70 eV. GC Programm: 5 min at 408, increasing  
with 38 minꢀ1 to 3208, operated in either split or splitless mode (60 s valve time); He carrier gas at 1 ml  
minꢀ1 (HP-6890) or 1.2 ml minꢀ1 (Agilent 7890A).  
RI Values were determined from a homologous series of n-alkanes (C8 –C35) [20]. Identification of  
compounds was performed by comparison of mass spectra to the Wiley-6 Library and the Essential Oils  
Library (Massfinder) and by comparison of RI data from the literature or with synthetic standards.  
GC/MS on Chiral Stationary Phase. Enantiomer separations were carried out on the Agilent 7890A/  
5975C system equipped with a b-Dex 225 fused-silica cap. column (30 m, 0.32 mm i.d., 0.25 mm film;  
Supelco/Sigma–Aldrich, Germany). The carrier gas was He at 1.6 ml minꢀ1 (valve time: 60 s, spitless  
injection), injection volume: 1 ml. The mono-methyl esters were separated with the following temp.  
program: 50 min at 608, then with 108 minꢀ1 to 2108. The dimethyl esters were separated with the  
following temp. program: 120 min at 508, then with 208 minꢀ1 to 2108.  
Derivatization with Dimethyl Disulfide (DMDS). According to the method of Leonhardt and  
DeVilbiss [16], a soln. of the natural extract (10 ml), DMDS (50 ml), and an I2 soln. (5 ml, 5% in Et2O) was  
heated to 608 for 8 h. The soln. was diluted with pentane, and excess I2 was removed by washing with sat.  
aq. Na2S2O3 soln. The org. layer was separated, dried (MgSO4), concentrated, and analyzed by GC/MS.  
Preparation of Methyl 2-(Dimethoxyphosphoryl)propanoate. A mixture of methyl 2-bromopropa-  
noate (18.37 g, 0.11 mol) and (MeO)3P (12.40 g, 0.1 mol) was stirred and heated to 105–1108 for 2 h [47].  
The MeBr formed during the reaction was removed continuously, finally furnishing the crude product.  
This preparation was purified on SiO2 using Et2O as eluent. The purified product still contained small  
amounts of irremovable side product. Rf (Et2O) 0.27. GC (BPX5): I 1293. 1H-NMR (400 MHz, CDCl3):  
1.42–1.50 (m, 3 H); 3.02–3.14 (m, 1 H); 3.76–3.77 (m, 3 H); 3.78–3.80 (m, 3 H); 3.81–3.83 (m, 3 H).  
13C-NMR (100 MHz, CDCl3): 11.6 (Me); 38.6 (d, J¼134.5, CH); 52.6 (Me); 53.3 (Me); 53.4 (Me); 170.0  
(C). EI-MS (70 eV): 196 (1, Mþ ), 181 (21), 165 (40), 151 (4), 141 (15), 137 (31), 127 (15), 109 (100), 93  
(12), 79 (27), 55 (53).  
Preparation of Unsaturated Methyl Esters. According to Dickschat et al. [27], NaH (1.3 equiv., 60% in  
mineral oil) was suspended in THF (1 ml/mmol) and cooled to 08. Methyl 2-(dimethoxyphosphoryl)-  
propanoate (1.3 equiv.) dissolved in THF (0.5 ml/mmol) was added dropwise to this suspension. The  
resulting mixture was stirred for 1 h at 408. After cooling to 08 the respective aldehyde (1–1.3 equiv.) in  
THF (0.5 ml/mmol) was added dropwise. The mixture was stirred under reflux for 24 h, and the reaction  
was quenched by the addition of 2m HCl. The aq. layer was extracted three times with Et2O, the  
combined org. phases were dried with MgSO4 and finally concentrated under vacuum. Column  
chromatography on SiO2 using a gradient of pentane/Et2O (200 :1 to 100 :1) gave the respective ester as a  
diastereoisomeric mixture.  
Methyl 2-Methylhept-2-enoate (18). Diastereoisomeric ratio (crude product): (E)/(Z) 2 :1. Yield:  
42% (328 mg, 2.10 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.61; (E): 0.49. GC (HP-5MS): (Z): RI 1088;  
(E): RI 1145. 1H-NMR (400 MHz, CDCl3): (E): 0.85–0.93 (m, 3 H); 1.24–1.46 (m, 4 H); 1.83–1.84 (m,  
3 H); 2.17 (q, J¼7.1, 2 H); 3.73 (s, 3 H); 6.77 (tq, J¼1.5, 7.5, 1 H). 13C-NMR (100 MHz, CDCl3): (E): 12.3  
(Me); 13.8 (Me); 22.4 (CH2); 28.3 (CH2); 30.7 (CH2); 51.6 (Me); 127.4 (C); 142.7 (CH); 168.7 (C). EI-  
MS (70 eV): (Z): 156 (46, Mþ ), 127 (100), 101 (33), 95 (47), 88 (27), 81 (10), 67 (27), 55 (55), 41 (29);  
(E): 156 (35, Mþ ), 127 (63), 114 (7), 101 (92), 88 (65), 81 (17), 69 (29), 55 (100), 54 (11), 41 (40).  
Methyl 2-Methyloct-2-enoate (19). Diastereoisomeric ratio (crude product): (E)/(Z) 3 :1. Yield:  
30% (257 mg, 1.48 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.64; (E): 0.46. GC (HP-5MS): (Z): RI 1189;  
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CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
(E): RI 1245. 1H-NMR (400 MHz, CDCl3): (E): 0.89 (t, J¼6.9, 3 H); 1.27–1.37 (m, 4 H); 1.40–1.49 (m,  
2 H); 1.83–1.84 (m, 3 H); 2.17 (dq, J¼0.8, 7.4, 2 H); 3.73 (s, 3 H); 6.77 (tq, J¼1.4, 7.5, 1 H). 13C-NMR  
(100 MHz, CDCl3): (E): 12.3 (Me); 14.0 (Me); 22.5 (CH2); 28.2 (CH2); 28.6 (CH2); 31.5 (CH2); 51.7  
(Me); 127.4 (C); 142.8 (CH); 168.8 (C). EI-MS (70 eV): (Z): 170 (42, Mþ ), 139 (19), 127 (100), 101 (58),  
95 (42), 88 (45), 83 (21), 81(14), 67 (25), 55 (18), 53 (20), 41 (63); (E): 170 (19, Mþ ), 139 (27), 127 (45),  
101 (100), 95 (21), 88 (69), 82 (21), 69 (53), 59 (20), 55 (56), 41 (38).  
Methyl 2-Methylnon-2-enoate (20). Diastereoisomeric ratio (crude product): (E)/(Z) 1.3 :1. Yield:  
78% (714 mg, 3.88 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.65; (E): 0.50. GC (HP-5MS): (Z): RI 1282;  
(E): RI 1346. 1H-NMR (400 MHz, CDCl3): (E): 0.89 (t, J¼6.9, 3 H); 1.21–1.36 (m, 6 H); 1.40–1.47 (m,  
2 H); 1.82–1.83 (m, 3 H); 2.17 (dq, J¼0.9, 7.4, 2 H); 3.73 (s, 3 H); 6.74–6.79 (m, 1 H). 13C-NMR  
(100 MHz, CDCl3): (E): 12.3 (Me); 14.1 (Me); 22.6 (CH2); 28.5 (CH2); 28.7 (CH2); 29.0 (CH2); 31.7  
(CH2); 51.6 (Me); 127.4 (C); 142.8 (CH); 168.8 (C). EI-MS (70 eV): (Z): 184 (34, Mþ ), 153 (20), 127  
(100), 101 (55), 95 (37), 88 (43), 83 (14), 69 (27), 55 (34), 41 (28); (E): 184 (10, Mþ ), 153 (23), 127 (37),  
101 (100), 88 (74), 82 (17), 69 (39), 55 (45), 41 (32).  
Methyl 2-Methyldec-2-enoate (21). Diastereoisomeric ratio (crude product): (E)/(Z) 2 :1. Yield:  
52% (514 mg, 2.60 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.64; (E): 0.51. GC (HP-5MS): (Z): RI 1381;  
(E): RI 1446. 1H-NMR (400 MHz, CDCl3): (E): 0.88 (t, J¼6.9, 3 H); 1.23–1.48 (m, 9 H); 1.60–1.67 (m,  
1 H); 1.82–1.83 (m, 3 H); 2.13–2.19 (m, 2 H); 3.73 (s, 3 H); 6.77 (tq, J¼1.4, 7.5, 1 H). 13C-NMR  
(100 MHz, CDCl3): (E): 12.3 (Me); 14.1 (Me); 22.6 (CH2); 28.6 (CH2); 29.0 (CH2); 29.1 (CH2); 29.3  
(CH2); 31.8 (CH2); 51.6 (Me); 127.4 (C); 142.9 (CH); 168.8 (C). EI-MS (70 eV): (Z): 198 (29, Mþ ), 167  
(18), 127 (100), 114 (12), 101 (67), 95 (33), 88 (52), 83 (13), 69 (24), 55 (32), 41 (29); (E): 198 (7, Mþ ),  
167 (19), 127 (29), 101 (100), 95 (17), 88 (71), 83 (17), 69 (29), 55 (36), 41 (29).  
Methyl 2-Methylundec-2-enoate (22). Diastereoisomeric ratio (crude product): (E)/(Z) 2 :1. Yield:  
74% (601 mg, 2.84 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.64; (E): 0.50. GC (HP-5MS): (Z): RI 1480;  
(E): RI 1547. 1H-NMR (400 MHz, CDCl3): (E): 0.88 (t, J¼6.8, 3 H); 1.20–1.52 (m, 11 H); 1.56–1.68 (m,  
1 H); 1.82–1.83 (m, 3 H); 2.16 (dq, J¼0.8, J¼7.4, 2 H); 3.73 (s, 3 H); 6.77 (tq, J¼1.4, 7.5, 1 H). 13C-NMR  
(100 MHz, CDCl3): (E): 12.3 (Me); 14.1 (Me); 22.6 (CH2); 28.7 (CH2); 29.2 (CH2); 29.3 (CH2); 29.4  
(CH2); 29.6 (CH2); 31.8 (CH2); 51.6 (Me); 127.4 (C); 142.8 (CH); 168.8 (C). EI-MS (70 eV): (Z): 212  
(22, Mþ ), 181 (17), 127 (100), 114 (13), 101 (77), 95 (32), 88 (57), 83 (13), 81 (13), 69 (25), 55 (31), 41  
(30); (E): 212 (6, Mþ ), 181 (16), 127 (27), 101 (100), 95 (17), 88 (73), 83 (15), 69 (28), 55 (33), 41 (28).  
Methyl 2-Methyldodec-2-enoate (23). Diastereoisomeric ratio (crude product): (E)/(Z) 2 :1. Yield:  
76% (665 mg, 2.94 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.61; (E): 0.48. GC (HP-5MS): (Z): RI 1580;  
(E): RI 1648. 1H-NMR (400 MHz, CDCl3): (E): 0.88 (t, J¼6.8, 3 H); 1.20–1.52 (m, 13 H); 1.55–1.69 (m,  
1 H); 1.82–1.83 (m, 3 H); 2.13–2.19 (m, 2 H); 3.73 (s, 3 H); 6.77 (tq, J¼1.6, 7.5, 1 H). 13C-NMR  
(100 MHz, CD3OD): (E): 12.3 (Me), 14.1 (Me), 22.7 (CH2), 28.7 (CH2), 29.3 (CH2), 29.4 (2 CH2), 29.5  
(CH2), 29.6 (CH2), 31.9 (CH2), 51.6 (Me), 128.6 (C), 142.8 (CH), 168.7 (C). EI-MS (70 eV): (Z): 226  
(23, Mþ ), 195 (16), 127 (100), 115 (17), 101 (66), 95 (32), 88 (59), 69 (23), 55 (29), 41 (30); (E): 226 (5,  
Mþ ), 195 (16), 127 (25), 101 (100), 95 (17), 88 (77), 83 (17), 69 (26), 55 (31), 41 (27).  
Methyl 2-Methyltridec-2-enoate (24). Diastereoisomeric ratio (crude product): (E)/(Z) 1:1. Yield:  
60% (554 mg, 2.31 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.61; (E): 0.48. GC (HP-5MS): (Z): RI 1677;  
(E): RI 1746. 1H-NMR (400 MHz, CDCl3): (E): 0.88 (t, J¼6.8, 3 H); 1.23–1.34 (m, 14 H); 1.36–1.47 (m,  
2 H); 1.82–1.83 (m, 3 H); 2.13–2.20 (m, 2 H); 3.73 (s, 3 H); 6.77 (dq, J¼1.4, 7.5, 1 H). 13C-NMR  
(100 MHz, CDCl3) (E): 12.3 (Me); 14.1 (Me); 22.6 (CH2); 28.6 (CH2); 29.3 (2 CH2); 29.4 (2 CH2); 29.6  
(2 CH2); 31.9 (CH2); 51.6 (Me); 127.3 (C); 142.8 (CH); 168.7 (C). EI-MS (70 eV): (Z): 240 (22, Mþ ), 209  
(15), 143 (9), 127 (100), 114 (14), 101 (69), 95 (31), 88 (63), 83 (15), 81 (15), 69 (23), 55 (29), 41 (31);  
(E): 240 (5, Mþ ), 209 (15), 127 (25), 114 (9), 101 (100), 95 (17), 88 (82), 83 (16), 81 (14), 69 (24), 55  
(30), 41 (28).  
Methyl 2-Methyltetradec-2-enoate (25). Diastereoisomeric ratio (crude product): (E)/(Z) 1:1. Yield:  
73% (715 mg, 2.81 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.59; (E): 0.48. GC (HP-5MS): (Z): RI 1778;  
(E): RI 1848. 1H-NMR (400 MHz, CDCl3): (E): 0.88 (t, J¼6.9, 3 H); 1.23–1.33 (m, 16 H); 1.36–1.45 (m,  
2 H); 1.82–1.83 (m, 3 H); 2.13–2.20 (m, 2 H); 3.73 (s, 3 H); 6.77 (dq, J¼1.4, 7.5, 1 H). 13C-NMR  
(100 MHz, CDCl3): (E): 12.3 (Me); 14.1 (Me); 22.7 (CH2); 28.6 (CH2); 29.3 (2 CH2); 29.4 (2 CH2); 29.6  
(3 CH2); 31.9 (CH2); 51.6 (Me); 127.4 (C); 142.8 (CH); 168.8 (C). EI-MS (70 eV): (Z): 254 (22, Mþ ), 223  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
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(14), 157 (9), 143 (21), 127 (100), 114 (14), 101 (69), 95 (30), 88 (65), 83 (14), 81 (15), 69 (23), 55 (29), 41  
(31); (E): 254 (4, Mþ ), 223 (12), 143 (5), 127 (24), 114 (8), 101 (100), 95 (17), 88 (91), 83 (16), 81 (14),  
69 (26), 55 (31), 41 (28).  
Methyl 2-Methylpentadec-2-enoate (26). Diastereoisomeric ratio (crude product): (E)/(Z) 1:1.  
Yield: 65% (667 mg, 2.49 mmol). Rf (pentane/Et2O 10 :1): (Z): 0.60; (E): 0.48. GC (HP-5MS): (Z): RI  
1877; (E): RI 1949. 1H-NMR (400 MHz, CDCl3): (E): 0.88 (t, J¼6.8, 3 H); 1.23–1.33 (m, 18 H); 1.35–  
1.45 (m, 2 H); 1.82–1.83 (m, 3 H); 2.13–2.19 (m, 2 H); 3.73 (s, 3 H); 6.77 (dq, J¼1.4, 7.5, 1 H). 13C-NMR  
(100 MHz, CDCl3): (E): 12.3 (Me); 14.1 (Me); 22.7 (CH2); 28.6 (CH2); 29.3 (2 CH2); 29.4 (2 CH2); 29.6  
(4 CH2); 31.9 (CH2); 51.6 (Me); 127.3 (C); 142.8 (CH); 168.7 (C). EI-MS (70 eV): (Z): 268 (22, Mþ ), 237  
(14), 171 (9), 157 (23), 127 (100), 114 (14), 101 (73), 95 (30), 88 (69), 83 (15), 81 (15), 69 (24), 55 (30),  
41 (32); (E): 268 (5, Mþ ), 237 (12), 157 (5), 127 (23), 114 (8), 101 (100), 99 (6), 98 (12), 95 (16), 88 (83),  
83 (15), 81 (14), 69 (23), 55 (29), 41 (25).  
Methyl (2E)-2,7-Dimethyloct-2-enoate (41a). Yield: 13% (91 mg, 0.49 mmol). Rf (pentane/Et2O  
1
10 :1) 0.40. GC (HP-5MS): RI 1313. H-NMR (400 MHz, CDCl3): 0.88 (d, J¼6.5, 6 H); 1.17–1.24 (m,  
2 H); 1.44 (dt, J¼7.6, 15.6, 2 H); 1.49–1.58 (m, 1 H); 1.83–1.84 (m, 3 H); 2.12–2.18 (m, 2 H); 3.74 (s,  
3 H); 6.74–6.79 (m, 1 H). 13C-NMR (100 MHz, CDCl3): 12.3 (Me); 22.5 (2 Me); 26.4 (CH2); 27.9 (CH);  
28.9 (CH2); 38.3 (CH2); 51.6 (Me); 127.4 (C); 142.8 (CH); 168.8 (C). EI-MS (70 eV): 184 (1, Mþ ), 152  
(68), 141 (4), 124 (25), 113 (10), 101 (75), 95 (23), 88 (100), 83 (21), 82 (65), 81 (26), 69 (71), 55 (42), 41  
(45).  
Methyl (2Z)-2,7-Dimethyloct-2-enoate (41b). Yield: 19% (138 mg, 0.75 mmol). Rf (pentane/Et2O  
10 :1) 0.53. GC (HP-5MS): RI 1246. EI-MS (70 eV): 184 (9, Mþ ), 153 (14), 152 (32), 141 (6), 127 (67),  
114 (11), 101 (55), 95 (48), 88 (67), 83 (22), 82 (59), 81 (33), 69 (66), 55 (60), 41 (100), 39 (50).  
Methyl (2E)-2,11-Dimethyldodec-2-enoate (42a). Yield: 25% (228 mg, 0.95 mmol). Rf (pentane/  
Et2O 10 :1) 0.58. GC (HP-5MS): RI 1710. 1H-NMR (400 MHz, CDCl3): 0.86 (d, J¼6.8, 6 H); 1.12–1.17  
(m, 2 H); 1.25–1.35 (m, 8 H); 1.40–1.46 (m, 2 H); 1.48–1.55 (m, 1 H); 1.82–1.83 (m, 3 H); 2.13–2.19 (m,  
2 H); 3.73 (s, 3 H); 6.77 (tq, J¼1.4, 7.5, 1 H). 13C-NMR (100 MHz, CDCl3): 12.3 (Me); 22.6 (2 Me); 27.3  
(CH2); 27.9 (CH); 28.6 (CH2); 28.7 (CH2); 29.4 (CH2); 29.5 (CH2); 29.8 (CH2); 39.0 (CH2); 51.6 (Me);  
127.4 (C); 142.8 (CH); 168.7 (C). EI-MS (70 eV): 240 (5, Mþ ), 209 (11), 208 (10), 127 (24), 114 (8), 101  
(100), 95 (22), 88 (72), 83 (17), 81 (17), 69 (32), 55 (31), 41 (35).  
Methyl (2Z)-2,11-Dimethyldodec-2-enoate (42b). Yield: 19% (174 mg, 0.73 mmol). Rf (pentane/  
Et2O 10 :1) 0.69. GC (HP-5MS): RI 1643. EI-MS (70 eV): 240 (26, Mþ ), 209 (15), 127 (100), 114 (14),  
101 (67), 95 (36), 88 (59), 83 (15), 81 (17), 69 (27), 55 (30), 43 (33), 41 (36).  
Methyl 2,6-Dimethyloct-2-enoate (48). Diastereoisomeric ratio (crude product): (E)/(Z) 6 :1. Yield:  
38% (203 mg, 1.10 mmol, over two steps). Rf (pentane/Et2O 10 :1): (Z): 0.74; (E): 0.62. GC (BPX5):  
(Z): RI 1247; (E): RI 1317. 1H-NMR (400 MHz, CDCl3): (E): 0.75–0.84 (m, 6 H); 1.04–1.42 (m, 4 H);  
1.50 (m, 1 H); 1.75–1.78 (m, 3 H); 2.04–2.14 (m, 2 H); 3.66 (s, 3 H); 7.19 (s, 1 H). 13C-NMR (100 MHz,  
CDCl3): (E): 11.3 (Me); 12.3 (Me); 19.0 (Me); 26.3 (CH2); 29.3 (CH2); 34.1 (CH); 35.3 (CH2); 51.7  
(Me); 127.2 (C); 143.0 (CH); 168.8 (C). EI-MS (70 eV): (Z): 184 (19, Mþ ), 127 (100), 101 (21), 95 (28),  
88 (7), 67 (15), 55 (1541 (17); (E): 184 (4, Mþ ), 153 (14), 127 (23), 114 (10), 101 (100), 95 (20), 88 (20),  
84 (33), 69 (37), 55 (30), 41 (31).  
Preparation of Saturated Methyl Esters. According to Suzukamo et al. [28], the respective  
unsaturated ester (1 mmol) was dissolved in 10 ml of dry MeOH, and 1 mol-% PtO2 was added. The  
resulting suspension was stirred, while H2 was passed through it, and the progress of the reaction was  
checked with GC. After the reaction was completed, column chromatography on SiO2 with pentane/Et2O  
10 :1 as eluent afforded the pure product.  
Methyl 2-Methylheptanoate (27). Yield: quant. (332 mg, 2.10 mmol). Rf (pentane/Et2O 10 :1) 0.59.  
GC (HP-5MS): RI 1060. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H); 1.25–  
1.46 (m, 7 H); 1.59–1.70 (m, 1 H); 2.44 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz, CDCl3):  
14.0 (Me); 17.0 (Me); 22.5 (CH2); 26.9 (CH2); 31.7 (CH2); 33.8 (CH2); 39.4 (CH); 51.4 (Me); 177.4 (C).  
EI-MS (70 eV): 143 (2, [Mꢀ15]þ ), 127 (6), 115 (8), 101 (25), 88 (100), 69 (5), 57 (28), 41 (12).  
Methyl 2-Methyloctanoate (28). Yield: 57% (138 mg, 0.80 mmol). Rf (pentane/Et2O 10 :1) 0.53. GC  
(HP-5MS): RI 1161. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H); 1.22–1.33  
(m, 8 H); 1.36–1.47 (m, 1 H); 1.61–1.69 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR  
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CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
(100 MHz, CDCl3): 14.0 (Me); 17.1 (Me); 22.6 (CH2); 27.2 (CH2); 29.2 (CH2); 31.7 (CH2); 33.8 (CH2);  
39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 172 (1, Mþ ), 143 (3), 141 (4), 129 (4), 115 (6), 101 (24),  
88 (100), 71 (7), 57 (16), 41 (11).  
Methyl 2-Methylnonanoate (29). Yield: 40% (176 mg, 0.95 mmol). Rf (pentane/Et2O 10 :1) 0.53. GC  
(HP-5MS): RI 1258. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H); 1.21–1.33  
(m, 10 H); 1.36–1.45 (m, 1 H); 1.60–1.69 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR  
(100 MHz, CDCl3): 14.0 (Me); 17.0 (Me); 22.6 (CH2); 27.2 (CH2); 29.1 (CH2); 29.5 (CH2); 31.8 (CH2);  
33.8 (CH2); 39.4 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 186 (1, Mþ ), 171 (1), 157 (3), 143 (8), 129  
(4), 115 (3), 101 (27), 88 (100), 69 (5), 55 (9), 41 (11).  
Methyl 2-Methyldecanoate (30). Yield: 80% (285 mg, 1.43 mmol). Rf (pentane/Et2O 10 :1) 0.52. GC  
(HP-5MS): RI 1359. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼7.1, 3 H); 1.14 (d, J¼6.8, 3 H); 1.19–1.46  
(m, 13 H); 1.60–1.69 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz, CDCl3): 14.1  
(Me); 17.1 (Me); 22.7 (CH2); 27.3 (CH2); 29.3 (CH2); 29.5 (2 CH2); 31.9 (CH2); 33.8 (CH2); 39.5 (CH);  
51.4 (Me); 177.4 (C). EI-MS (70 eV): 200 (2, Mþ ), 169 (3), 157 (7), 143 (9), 115 (3), 101 (30), 88 (100),  
69 (6), 57 (11), 41 (10).  
Methyl 2-Methylundecanoate (31). Yield: 76% (237 mg, 1.11 mmol). Rf (pentane/Et2O 10 :1) 0.53.  
GC (HP-5MS): RI 1459. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H); 1.20–  
1.34 (m, 14 H); 1.35–1.45 (m, 1 H); 1.60–1.69 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR  
(100 MHz, CDCl3): 14.1 (Me); 17.1 (Me); 22.7 (CH2); 27.2 (CH2); 29.3 (CH2); 29.5 (3 CH2); 31.9 (CH2);  
33.8 (CH2); 39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 214 (2, Mþ ), 183 (3), 171 (5), 157 (9), 143  
(7), 115 (4), 101 (32), 88 (100), 69 (6), 57 (11), 41 (10).  
Methyl 2-Methyldodecanoate (32). Yield: 71% (332 mg, 1.46 mmol). Rf (pentane/Et2O 10 :1) 0.52.  
GC (HP-5MS): RI 1559. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H); 1.22–  
1.33 (m, 16 H); 1.35–1.44 (m, 1 H); 1.60–1.70 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR  
(100 MHz, CDCl3): 14.1 (Me); 17.0 (Me); 22.7 (CH2); 27.2 (CH2); 29.3 (CH2); 29.5 (2 CH2); 29.6  
(2 CH2); 31.9 (CH2); 33.8 (CH2); 39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 228 (3, Mþ ), 197 (2),  
185 (5), 171 (6), 157 (6), 143 (7), 115 (3), 101 (34), 98 (2), 88 (100), 69 (6), 57 (10), 41 (10).  
Methyl 2-Methyltridecanoate (33). Yield: 76% (226 mg, 0.93 mmol). Rf (pentane/Et2O 10 :1) 0.53.  
GC (HP-5MS): RI 1660. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.8, 3 H); 1.14 (d, J¼6.8, 3 H); 1.21–  
1.47 (m, 19 H); 1.57–1.70 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz, CDCl3):  
14.1 (Me); 17.0 (Me); 22.7 (CH2); 27.2 (CH2); 29.3 (CH2); 29.5 (2 CH2); 29.6 (3 CH2); 31.9 (CH2); 33.8  
(CH2); 39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 242 (4, Mþ ), 211 (2), 199 (6), 185 (6), 157 (7),  
143 (9), 115 (3), 101 (36), 88 (100), 69 (7), 55 (10), 41 (10).  
Methyl 2-Methyltetradecanoate (34). Yield: 73% (336 mg, 1.31 mmol). Rf (pentane/Et2O 10 :1) 0.57.  
GC (HP-5MS): RI 1759. 1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H); 1.21–  
1.46 (m, 21 H); 1.59–1.70 (m, 2 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz, CDCl3):  
14.1 (Me); 17.1 (Me); 22.7 (CH2); 27.2 (CH2); 29.4 (CH2); 29.5 (2 CH2); 29.6 (3 CH2); 29.7 (CH2); 31.9  
(CH2); 33.8 (CH2); 39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 256 (5, Mþ ), 225 (2), 213 (6), 199  
(8), 157 (8), 143 (9), 115 (3), 101 (38), 88 (100), 69 (7), 55 (10), 41 (10).  
Methyl 2-Methylpentadecanoate (35). Yield: 83% (250 mg, 0.93 mmol). Rf (pentane/Et2O 10 :1)  
1
0.56. GC (HP-5MS): RI 1859. H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H);  
1.21–1.46 (m, 23 H); 1.59–1.70 (m, 2 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz,  
CDCl3): 14.1 (Me); 17.1 (Me); 22.7 (CH2); 27.2 (CH2); 29.4 (CH2); 29.5 (2 CH2); 29.6 (3 CH2); 29.7  
(2 CH2); 31.9 (CH2); 33.8 (CH2); 39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 270 (7, Mþ ), 239 (2),  
227 (5), 213 (8), 199 (2), 171 (2), 157 (9), 143 (8), 101 (38), 88 (100), 69 (8), 55 (11), 41 (10).  
Methyl 2,7-Dimethyloctanoate (43). Yield: 66% (153 mg, 0.82 mmol). Rf (pentane/Et2O 10 :1) 0.58.  
GC (HP-5MS): RI 1223. 1H-NMR (400 MHz, CDCl3): 0.86 (d, J¼6.6, 6 H); 1.12–1.18 (m, 5 H); 1.24–  
1.35 (m, 4 H); 1.36–1.56 (m, 2 H); 1.60–1.70 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR  
(100 MHz, CDCl3): 17.1 (Me); 22.6 (2 Me); 27.3 (CH2); 27.5 (CH2); 27.9 (CH); 33.9 (CH2); 38.8 (CH2);  
39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 186 (1, Mþ ), 171 (2), 155 (3), 143 (13), 129 (3), 115 (4),  
101 (40), 88 (100), 69 (13), 57 (14), 55 (13), 43 (15), 41 (18).  
Methyl 2,11-Dimethyldodecanoate (44). Yield: 61% (156 mg, 0.64 mmol). Rf (pentane/Et2O 10 :1)  
0.59. GC (HP-5MS): RI 1620. 1H-NMR (300 MHz, CDCl3): 0.86 (d, J¼6.6, 6 H); 1.12–1.46 (m, 10 H);  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
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1.47–1.56 (m, 1 H); 1.58–1.71 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (75 MHz,  
CDCl3): 17.1 (Me); 22.6 (2 Me); 27.2 (CH2); 27.4 (CH2); 28.0 (CH); 29.5 (2 CH2); 29.6 (CH2); 29.9 (CH2);  
33.8 (CH2); 39.0 (CH2); 39.5 (CH); 51.4 (Me); 177.4 (C). EI-MS (70 eV): 242 (4, Mþ ), 211 (3), 199 (11),  
185 (7), 157 (8), 143 (10), 101 (41), 88 (100), 69 (9), 55 (13), 43 (15), 41 (13).  
Methyl 2,6-Dimethyloctanoate (49). Yield: quant. (183 mg, 0.99 mmol). Rf (pentane/Et2O 10 :1)  
1
0.55. GC (HP-5MS): RI 1229. H-NMR (400 MHz, CDCl3): 0.83–0.94 (m, 6 H); 1.07–1.15 (m, 5 H);  
1.20–1.44 (m, 6 H); 1.54–1.70 (m, 1 H); 2.39–2.49 (m, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz, CDCl3):  
11.4 (Me); 17.1 (Me); 19.1 (Me); 24.7 (CH2); 29.4 (CH2); 34.1 (CH2); 34.2 (CH); 36.4 (CH2); 39.5 (CH);  
51.4 (Me); 177.4 (C). EI-MS (70 eV): 186 (2, Mþ ), 157 (14), 129 (9), 115 (3), 101 (46), 88 (100), 69 (15),  
55 (28), 41 (24).  
Preparation of (wꢀ1)-Me-Branched Alcohols. According to the procedure of Fꢀrstner et al. [29], a  
soln. of i-BuMgBr (40 mmol, 2.5 equiv.) in THF (12.5 ml), freshly prepared from Mg (40 mmol,  
2.5 equiv.) and i-BuBr (40 mmol, 2.5 equiv.), was added to an ice-cooled soln. of the respective 1-  
bromoalcohol (16 mmol, 1 equiv.) and Li2CuCl4 (0.1m soln. in THF, 3 mmol, 0.19 mmol). The mixture  
was stirred for 1.5 h at 08 and was then allowed to warm up to r.t. Then, conc. HCl (10–15 ml) was added.  
The aq. layer was separated and extracted three times with Et2O. The combined org. extracts were  
successively washed with sat. NaHCO3 and brine, dried (MgSO4), and then concentrated under vacuum.  
The crude product was purified by CC on SiO2 with pentane/Et2O (5 :1) as eluent to yield the pure  
alcohol.  
5-Methylhexan-1-ol (37). Yield: 85% (1.571 g, 13.55 mmol). Rf (pentane/Et2O 5 :1) 0.07. GC  
(BPX5): RI 942. 1H-NMR (400 MHz, CDCl3): 0.88 (d, J¼6.5, 6 H); 1.16–1.22 (m, 2 H); 1.31–1.38 (m,  
2 H); 1.51–1.59 (m, 3 H); 3.61–3.69 (m, 2 H). 13C-NMR (100 MHz, CDCl3): 22.5 (2 Me); 23.5 (CH2);  
27.9 (CH); 32.9 (CH2); 38.7 (CH2); 62.9 (CH2). EI-MS (70 eV): 98 (1, [Mꢀ18]þ ), 83 (21), 70 (40), 69  
(42), 56 (90), 55 (100), 43 (73), 42 (30), 41 (85).  
9-Methyldecan-1-ol (38). Yield: 68% (1.161 g, 6.75 mmol). Rf (pentane/Et2O 5 :1) 0.07. GC (BPX5):  
1
RI 1338. H-NMR (400 MHz, CDCl3): 0.86 (d, J¼6.8, 6 H); 1.12–1.17 (m, 2 H); 1.24–1.38 (m, 9 H);  
1.46–1.60 (m, 4 H); 3.63 (t, J¼6.7, 2 H). 13C-NMR (100 MHz, CDCl3): 22.6 (2 Me); 25.7 (CH2); 27.4  
(CH2); 27.9 (CH); 29.4 (CH2); 29.6 (CH2); 29.8 (CH2); 32.8 (CH2); 39.0 (CH2); 63.0 (CH2). EI-MS  
(70 eV): 139 (2, [Mꢀ43]þ ), 126 (5), 111 (12), 98 (13), 97 (12), 83 (34), 69 (78), 57 (63), 56 (100), 55  
(84), 43 (86), 41 (91).  
Preparation of 4-Methylhexan-1-ol (46). The alcohol was prepared according to known methods by  
LiAlH4 reduction of 4-methylhexanoic acid (0.5 g. 3.85 mmol) [22]. Aqueous workup and removal of the  
solvents furnished pure 46. Yield: 75% (0.335 g, 2.89 mmol). Rf (pentane/Et2O 5 :1) 0.09. GC (BPX5):  
RI 954. 1H-NMR (300 MHz, CDCl3): 0.84–0.89 (m, 6 H); 1.08–1.23 (m, 2 H); 1.28–1.42 (m, 3 H); 1.45–  
1.67 (m, 2 H); 2.37 (s, 1 H); 3.60 (t, J¼6.7, 2 H). 13C-NMR (75 MHz, CDCl3): 11.7 (Me); 19.5 (Me); 29.8  
(CH2); 30.6 (CH2); 32.9 (CH2); 34.6 (CH); 63.6 (CH). EI-MS (70 eV): 98 (4, Mþ ), 83 (8), 70 (100), 69  
(88), 57 (27), 56 (32), 55 (40), 43 (18), 42 (19), 41 (74), 39 (17).  
Preparation of (wꢀ1)- and (wꢀ2)-Me-Branched Aldehydes. The aldehydes were synthesized from  
the corresponding alcohols with PCC according to the standard procedure of Corey and Suggs [30].  
Purification was performed by CC on SiO2 using pentane/Et2O in the ratio mentioned for the respective  
aldehyde.  
5-Methylhexanal (39). Yield: 69% (1.017 g, 8.92 mmol). Rf (pentane/Et2O 3 :1) 0.61. GC (BPX5):  
RI 869. 1H-NMR (400 MHz, CDCl3): 0.89 (d, J¼6.6, 6 H); 1.18–1.24 (m, 2 H); 1.50–1.68 (m, 3 H); 2.31–  
2.43 (m, 2 H); 9.77 (t, J¼1.9, 1 H). 13C-NMR (100 MHz, CDCl3): 19.9 (CH2); 22.4 (2 Me); 27.8 (CH);  
38.3 (CH2); 44.1 (CH2); 203.0 (CH). EI-MS (70 eV): (33, [Mꢀ18]þ ), 86 (8), 81 (38), 71 (49), 55 (81), 43  
(100), 41 (90), 39 (48).  
9-Methyldecanal (40). Yield: 86% (0.958 g, 5.64 mmol). Rf (pentane/Et2O 2 :1) 0.75. GC (BPX5): RI  
1267. 1H-NMR (400 MHz, CDCl3): 0.86 (d, J¼6.8, 6 H); 1.12–1.17 (m, 2 H); 1.24–1.35 (m, 8 H); 1.44–  
1.57 (m, 1 H); 1.63 (quint., J¼7.3, 2 H); 2.42 (dt, J¼1.9, 7.3, 2 H); 9.77 (t, J¼1.9, 1 H). 13C-NMR  
(100 MHz, CDCl3): 22.1 (CH2); 22.6 (2 Me); 27.3 (CH2); 27.9 (CH); 29.2 (CH2); 29.4 (CH2); 29.7 (CH2);  
39.0 (CH2); 43.9 (CH2); 202.9 (CH). EI-MS (70 eV): 152 (1, [Mꢀ18]þ ), 142 (3), 137 (6), 124 (9), 109  
(25), 96 (35), 95 (39), 82 (49), 81 (41), 69 (60), 57 (89), 43 (94), 41 (100).  
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CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
4-Methylhexanal (47). Rf (pentane/Et2O 3 :1) 0.60. 1H-NMR (200 MHz, CDCl3): 0.84–0.92 (m,  
6 H); 1.09–1.55 (m, 4 H); 1.57–1.76 (m, 1 H); 2.30–2.49 (m, 2 H); 9.78 (t, J¼1.9, 1 H). 13C-NMR  
(50 MHz, CDCl3): 11.9 (Me); 19.5 (Me); 29.2 (CH2); 29.8 (CH2); 34.7 (CH); 42.4 (CH2); 203.6 (CH). EI-  
MS (70 eV): 96 (13, Mþ ), 81 (17), 71 (29), 70 (100), 57 (63), 55 (63), 43 (31), 42 (19), 41 (78), 39 (29).  
Preparation of 7-Methyloctanoyl Chloride (55). According to the procedure of Spessard et al. [34], 7-  
methyloctanoic acid (54) (140 mg, 0.89 mmol) was dissolved in 5 ml of dry Et2O and cooled to 08. Oxalyl  
chloride (225 mg, 1.77 mmol) was added dropwise, and the mixture was stirred for 24 h at r.t. The solvent  
and excess oxalyl chloride were removed under vacuum to produce pure 55 (125 mg, 0.71 mmol, 80%). Rf  
(pentane/Et2O 2 :1) 0.93. GC (BPX5): RI 1202. 1H-NMR (400 MHz, CDCl3): 0.87 (d, J¼6.5, 6 H); 1.14–  
1.19 (m, 2 H); 1.26–1.38 (m, 4 H); 1.52 (non., J¼6.6, 1 H); 1.63 (quint., J¼7.3, 2 H); 2.88 (t, J¼7.3, 2 H).  
13C-NMR (100 MHz, CDCl3): 22.6 (2 Me); 25.1 (CH2); 26.8 (CH2); 27.9 (CH); 28.7 (CH2); 38.6 (CH2);  
47.1 (CH2); 173.9 (C). EI-MS (70 eV): 141 (15, [Mꢀ35/37]þ ), 125 (18), 123 (17), 107 (16), 98 (33), 97  
(42), 96 (30), 84 (55), 69 (48), 55 (100), 43 (93), 41 (73).  
Preparation of Evans Chiral Auxiliary Acid Derivatives. As described by Gage and Evans [31], a  
soln. of (4S)-4-benzyl-1,3-oxazolidin-2-one (1 equiv.) in THF (3 ml/mmol) was cooled to ꢀ788. Then,  
BuLi (1.6m in hexane, 1 equiv.) was added, and the resulting mixture was stirred for 30 min under these  
conditions. The respective octanoyl chloride (1.1 equiv.) was added dropwise, and the stirring was  
continued for 4 h at ꢀ788. The mixture was allowed to warm up to r.t., and the reaction was quenched by  
the addition of sat. NH4Cl soln. Then, the org. solvents were removed under vacuum, and the resulting aq.  
layer was extracted three times with CH2Cl2. The combined org. layers were successively washed with 1m  
NaOH soln. and brine, dried (MgSO4), and concentrated under vacuum. Purification of the crude  
product by CC on SiO2 with pentane/Et2O (5 :1) furnished the pure product.  
(4S)-4-Benzyl-3-octanoyl-1,3-oxazolidin-2-one (51). Yield: 84% (2.54 g, 8.38 mmol). Rf (pentane/  
Et2O 5 :1) 0.14. GC (BPX5): RI 2459. 1H-NMR (400 MHz, CDCl3): 0.89 (t, J¼6.9, 3 H); 1.25–1.42 (m,  
8 H); 1.63–1.77 (m, 2 H); 2.77 (dd, J¼9.7, J¼13.4, 1 H); 2.86–3.01 (m, 2 H); 3.30 (dd, J¼3.4, 13.4, 1 H);  
4.14–4.22 (m, 2 H); 4.64–4.70 (m, 1 H); 7.20–7.35 (m, 5 H). 13C-NMR (100 MHz, CDCl3): 14.1 (Me);  
22.6 (CH2); 24.2 (CH2); 29.0 (CH2); 29.1 (CH2); 31.7 (CH2); 35.5 (CH2); 37.9 (CH2); 55.1 (CH); 66.1  
(CH2); 127.3 (CH); 128.9 (2 CH); 129.4 (2 CH); 135.3 (C); 153.4 (C); 173.4 (C). EI-MS (70 eV): 303 (6,  
Mþ ), 232 (3), 219 (8), 212 (18), 178 (4), 127 (100), 117 (10), 109 (4), 91 (17), 57 (32), 41 (9).  
(4S)-4-Benzyl-3-(7-methyloctanoyl)-1,3-oxazolidin-2-one (56). Yield: 48% (96 mg, 0.31 mmol). Rf  
(pentane/Et2O 5 :1) 0.12. GC (BPX5): RI 2521. 1H-NMR (400 MHz, CDCl3): 0.87 (d, J¼6.8, 6 H); 1.15–  
1.21 (m, 2 H); 1.29–1.41 (m, 4 H); 1.48–1.58 (m, 1 H); 1.66–1.74 (m, 2 H); 2.77 (dd, J¼9.6, 13.1, 1 H);  
2.85–3.01 (m, 2 H); 3.30 (dd, J¼3.3, 13.4, 1 H); 4.15–4.22 (m, 2 H); 4.64–4.69 (m, 1 H); 7.20–7.36 (m,  
5 H). 13C-NMR (100 MHz, CDCl3): 22.6 (2 Me); 24.3 (CH2); 27.2 (CH2); 27.9 (CH); 29.4 (CH2); 35.6  
(CH2); 38.0 (CH2); 38.8 (CH2); 55.2 (CH); 66.1 (CH2); 127.3 (CH); 129.0 (2 CH); 129.4 (2 CH); 135.3  
(C); 153.5 (C); 173.5 (C). EI-MS (70 eV): 317 (13, Mþ ), 302 (2), 226 (33), 219 (19), 178 (18), 141 (100),  
123 (78), 117 (27), 91 (37), 81 (28), 71 (15), 67 (14), 57 (20), 55 (23), 43 (28), 41 (20).  
Preparation of Methylated Evans Chiral Auxiliary Acid Derivatives. According to the procedure of  
Siebum et al. [32], a soln. of the acylated Evans oxazolidinone (1 equiv.) in dry THF (3 ml/mmol) was  
added dropwise to a cold (ꢀ 788) soln. of NaHMDS (1.0m soln. in THF, 2.5 equiv.) in THF (3 ml/mmol).  
The resulting mixture was stirred for 1 h at ꢀ788, and then MeI (2 equiv.) was added. The mixture was  
stirred for 4 h at ꢀ788 and then slowly allowed to warm up to r.t. Workup was carried out by the addition  
of sat. NH4Cl soln., H2O, and 1m H2SO4 to adjust the pH to 1–2. The aq. layer was first separated and then  
extracted three times with Et2O. The combined org. extracts were successively washed with sat. NaHCO3,  
sat. Na2S2O3, and brine, dried (MgSO4), and then concentrated applying vacuum. The crude product  
(diastereoisomeric ratio 98 :2 (52) and 96 :4 (57), resp.) was purified by CC on SiO2 with pentane/Et2O  
(gradient from 10 :1 to 7:1) as eluent. The obtained products were diastereoisomerically almost pure.  
(4S)-4-Benzyl-3-[(2S)-2-methyloctanoyl]-1,3-oxazolidin-2-one (52). Diastereoisomeric ratio: 99 :1.  
Yield: 66% (859 mg, 8.38 mmol). Rf (pentane/Et2O 10 :1) 0.13. GC (BPX5): RI 2442. 1H-NMR  
(400 MHz, CDCl3): 0.87 (t, J¼6.8, 3 H); 1.22 (d, J¼6.8, 3 H); 1.24–1.35 (m, 6 H); 1.36–1.46 (m, 2 H);  
2.77 (dd, J¼9.6, 13.4, 1 H); 3.27 (dd, J¼3.3, 13.1, 1 H); 3.71 (sext., J¼6.8, 1 H); 4.14–4.22 (m, 2 H);  
4.64–4.70 (m, 1 H); 7.20–7.35 (m, 5 H). 13C-NMR (100 MHz, CDCl3): 14.0 (Me); 17.3 (Me); 22.6 (CH2);  
27.2 (CH2); 29.3 (CH2); 31.7 (CH2); 33.4 (CH2); 37.7 (CH); 37.9 (CH2); 55.3 (CH); 66.0 (CH2); 127.3  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
2251  
(CH); 128.9 (2 CH); 129.4 (2 CH); 135.3 (C); 153.0 (C); 177.3 (C). EI-MS (70 eV): 317 (5, Mþ ), 233  
(18), 226 (20), 178 (14), 141 (100), 134 (7), 117 (21), 113 (22), 91 (39), 86 (21), 71 (65), 57 (57), 43 (27),  
41 (32).  
(4S)-4-Benzyl-3-[(2S)-2,7-dimethyloctanoyl]-1,3-oxazolidin-2-one (57). Diastereoisomeric ratio:  
1
100 :0. Yield: 46% (70 mg, 0.22 mmol). Rf (pentane/Et2O 5 :1) 0.12. GC (BPX5): RI 2494. H-NMR  
(400 MHz, CDCl3): 0.86 (d, J¼6.8, 6 H); 1.12–1.19 (m, 2 H); 1.22 (d, J¼6.8, 3 H); 1.25–1.33 (m, 4 H);  
1.37–1.56 (m, 2 H); 1.70–1.79 (m, 1 H); 2.77 (dd, J¼9.7, 13.4, 1 H); 3.27 (dd, J¼3.1, 13.4, 1 H); 3.67–  
3.75 (m, 1 H); 4.15–4.22 (m, 2 H); 4.65–4.70 (m, 1 H); 7.21–7.35 (m, 5 H). 13C-NMR (100 MHz,  
CDCl3): 17.4 (Me); 22.6 (2 Me); 27.4 (CH2); 27.5 (CH2); 27.9 (CH); 33.5 (CH2); 37.7 (CH); 37.9 (CH2);  
38.8 (CH2); 55.4 (CH); 66.0 (CH2); 127.3 (CH); 128.9 (2 CH); 129.4 (2 CH); 135.3 (C); 153.1 (C); 177.4  
(C). EI-MS (70 eV): 331 (5, Mþ ), 316 (2), 240 (19), 233 (23), 178 (22), 155 (100), 134 (8), 133 (8), 127  
(22), 117 (22), 91 (30), 85 (34), 71 (50), 57 (30), 43 (29), 41 (23).  
Preparation of Carboxylic Acids from Acylated Evans Oxazolidinones. As reported by Siebum et al.  
[32], a H2O2 soln. (30% in H2O, 5 equiv.) and LiOH (2.5 equiv.), dissolved in H2O (0.5 ml/mmol), were  
added at 08 to a soln. of the respective oxazolidinone (1 equiv.) in THF (10 ml/mmol) and H2O (5 ml/  
mmol). The mixture was stirred for 2 h under these conditions. Sodium sulfite in H2O (1.9 equiv., 4 ml/  
mmol) was added, and the mixture was stirred for 15 min at 08. The soln. was adjusted to pH 9–10 by the  
addition of sat. NaHCO3 soln. THF was evaporated, and the residual aq. layer was extracted two times  
with CH2Cl2. The combined org. phases were dried (MgSO4) and then concentrated under vacuum to  
give the recovered auxiliary (4S)-4-benzyl-1,3-oxazolidin-2-one. The aq. soln. was acidified to pH 1–2  
with 1m H2SO4 soln. and extracted three times with Et2O. The combined org. extracts were dried  
(MgSO4), and the solvents were evaporated to furnish the respective pure carboxylic acid.  
(2S)-2-Methyloctanoic Acid (53). Yield: quant. (300 mg, 1.90 mmol). GC (BPX5): RI 1269.  
1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.18 (d, J¼6.8, 3 H); 1.24–1.37 (m, 8 H); 1.39–1.47  
(m, 1 H); 1.64–1.73 (m, 1 H); 2.46 (sext., J¼6.9, 1 H); 10.52 (br. s, 1 H). 13C-NMR (100 MHz, CDCl3):  
14.0 (Me); 16.8 (Me); 22.6 (CH2); 27.1 (CH2); 29.2 (CH2); 31.7 (CH2); 33.5 (CH2); 39.4 (CH); 183.4 (C).  
EI-MS (70 eV): 158 (1, Mþ ), 129 (5), 115 (6), 101 (8), 87 (30), 74 (100), 69 (5), 55 (11), 43 (14), 41 (17).  
(2S)-2,7-Dimethyloctanoic Acid (58). Yield: 89% (32 mg, 0.19 mmol). GC (BPX5): RI 1337.  
1H-NMR (400 MHz, CDCl3): 0.86 (d, J¼6.6, 6 H); 1.13–1.23 (m, 5 H); 1.24–1.35 (m, 4 H); 1.39–1.56  
(m, 2 H); 1.64–1.73 (m, 1 H); 2.46 (sext., J¼7.0, 1 H). 13C-NMR (100 MHz, CDCl3): 16.8 (Me); 22.6  
(2 Me); 27.3 (CH2); 27.4 (CH2); 27.9 (CH); 33.6 (CH2); 38.8 (CH2); 39.3 (CH); 183.0 (C). EI-MS (70 eV):  
172 (1, Mþ ), 157 (4), 129 (21), 115 (6), 101 (4), 87 (56), 74 (100), 69 (20), 57 (16), 55 (21), 43 (25), 41  
(28).  
Preparation of Methyl (2S)-2-Methyloctanoate ((S)-28). Similar to the procedure of Wipf et al. [33],  
the acid 53 (258 mg, 1.63 mmol) was dissolved in an excess of dry MeOH (16 ml), and a few drops of  
conc. HCl were added. The mixture was stirred under reflux for 4 h, concentrated, and then filtered  
through SiO2 with Et2O as eluent. After drying (MgSO4), all solvents were removed under vacuum to  
give the pure product (257 mg, 1.49 mmol, 92%). Rf (pentane/Et2O 10 :1) 0.53. GC (HP-5MS): RI 1161.  
1H-NMR (400 MHz, CDCl3): 0.88 (t, J¼6.9, 3 H); 1.14 (d, J¼7.0, 3 H); 1.22–1.33 (m, 8 H); 1.36–1.45  
(m, 1 H); 1.61–1.69 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz, CDCl3): 14.0  
(Me); 17.0 (Me); 22.6 (CH2); 27.2 (CH2); 29.1 (CH2); 31.7 (CH2); 33.8 (CH2); 39.4 (CH); 51.4 (Me);  
177.4 (C). EI-MS (70 eV): EI-MS (70 eV): 172 (1, Mþ ), 143 (5), 141 (4), 129 (4), 115 (6), 101 (25), 88  
(100), 71 (6), 57 (17), 41 (12).  
Preparation of Methyl (2S)-2,7-Dimethyloctanoate ((S)-43). According to Patel et al. [35],  
compound 58 (16 mg, 0.09 mmol) was dissolved in 3.5 ml of dry CH2Cl2. A few drops of MeOH were  
added to the soln. before it was cooled to 08. DMAP (15 mg, 0.01 mmol) and EDC were added. The  
mixture was stirred for 1 h under these conditions and for 1.5 h at r.t. Then, Et2O was added, and the org.  
layer was successively washed with H2O and sat. NaHCO3 soln. The combined org. layers were dried  
(MgSO4) and concentrated under vacuum. Purification on SiO2 with pentane/Et2O 2 :1 as eluent  
afforded the pure product (11 mg, 0.57 mmol, 61%). Rf (pentane/Et2O 10 :1) 0.58. GC (HP-5MS): RI  
1223. 1H-NMR (400 MHz, CDCl3): 0.86 (d, J¼6.6, 6 H); 1.12–1.18 (m, 5 H); 1.24–1.35 (m, 4 H); 1.36–  
1.56 (m, 2 H); 1.60–1.70 (m, 1 H); 2.43 (sext., J¼7.0, 1 H); 3.67 (s, 3 H). 13C-NMR (100 MHz, CDCl3):  
17.1 (Me); 22.6 (2 Me); 27.3 (CH2); 27.5 (CH2); 27.9 (CH); 33.9 (CH2); 38.8 (CH2); 39.5 (CH); 51.4 (Me);  
2252  
CHEMISTRY & BIODIVERSITY – Vol. 7 (2010)  
177.4 (C). EI-MS (70 eV): 186 (1, Mþ ), 171 (2), 155 (3), 143 (14), 129 (3), 115 (4), 101 (41), 88 (100), 69  
(13), 57 (14), 55 (13), 43 (14), 41 (17).  
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Received July 2, 2010  

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