5.2 Aroma Analysis 353by chromatography on silica gel, as shown in Ta-
ble 5.11 for coffee aroma. To localize the aroma
substances each of the four fractions is ana-
lyzed by gas chromatography and olfactometry.
Some volatile compounds are aroma active in
such low concentrations that even enrichment by
column chromatography does not allow identifi-
cation, e. g., 3-methyl-2-butenethiol (Fraction A
in Table 5.11) and the two methoxypyrazines
(Fraction B) in coffee. In most cases, further
concentration is achieved with the help of multi-
dimensional gas chromatography (MGC). The
fraction which contains the unknown aroma
Table 5.11. Column chromatographic preliminary
separation of an aroma extract of roasted coffee
FractionaAroma substance
A 2-Methyl-3-furanthiol, 2-furfurylthiol,
bis(2-methyl-3-furyl)disulfide,
3-methyl-2-butenethiol
B 2,3-Butanedione, 3-methylbutanal,
2,3-pentanedione, trimethylthiazole,
3-mercapto-3-methylbutylformiate,
3-isopropyl-2-methoxypyrazine,
phenylacetaldehyde,
3-isobuty1-2-methoxypyrazine,
5-methyl-5(H)-cyclopentapyrazine,
p-anisaldehyde,
(E)-β-damascenone
C Methional, 2-ethenyl-3,5-dimethylpyrazine,
linalool, 2,3-diethyl-5-methylpyrazine,
guaiacol, 2-ethenyl-3-ethyl-
5-methylpyrazine,
4-ethylguaiacol, 4-vinylguaiacol
D 2-/3-Methylbutyric acid, trimethylpyrazine,
3-mercapto-3-methyl-1-butanol,
5-ethyl-2,4-dimethylthiazole,
2-ethyl-3,5-dimethylpyrazine, 3,4-dimethy1-
2-cyclopentenol-1-one, 4-hydroxy-2,5-
dimethyl-3(2H)-furanone, 5-ethyl-4-
hydroxy-2-methyl-3(2H)-furanone,
3-hydroxy-4,5-dimethy1-2(5H)-furanone,
5-ethyl-3-hydroxy-4-methyl-
2(5H)-furanone, vanillin
aChromatography at 10–12◦C on a silica gel column
(24×1 cm, deactivated with 7% water); elution with
mixtures of pentane-diethylether (50 ml, 95+5, v/v,
fraction A; 30 ml, 75×25, v/v, Fraction B; 30 ml,
1 +1, v/v, Fraction C) and with diethylether (100 ml,
Fraction D).
substance is first subjected to preliminary sepa-
ration on a polar capillary. The eluate containing
the substance is cut out, rechromatographed on
a non-polar capillary and finally analyzed by
mass spectrometry. The MGC is also used in
quantitative analysis for the preliminary purifica-
tion of analyte and internal standard (cf. 5.2.6.1).5.2.4 ChemicalStructure......................................
In the structure elucidation of aroma substances,
mass spectrometry has become an indispensable
tool because the substance amounts eluted by gas
chromatography are generally sufficient for an
evaluable spectrum. If the corresponding refer-
ence substance is available, identification of the
aroma substance is based on agreement of the
mass spectrum, retention indices on at least two
capillary columns of different polarity, and of
odor thresholds, which are compared by gas chro-
matography/olfactometry. If the reference sub-
stance is not available, the following procedure
is suitable for the identification of the odorant:
The analyte is concentrated until a^1 H-NMR
spectrum and, if necessary, a^13 C-spectrum can
be measured. An example is the identification of
the characteristic odorant of roasted hazelnuts.
The mass spectrum of this substance (Fig. 5.11a)
indicates an unsaturated carbonyl compound with
a molar mass of 126. In conjunction with the
structural elements shown by the^1 H-NMR spec-
trum (Fig. 5.11b), it was proposed that the odor-
ant is 5-methyl-(E)-2-hepten-4-one (Filbertone).
It goes without saying that the synthesis of the
proposed aroma substance was part of the identi-
fication. It was also guaranteed that its chromato-
graphic and sensory properties correspond with
those of the unknown odorant.5.2.5 EnantioselectiveAnalysis.................................
In the case of chiral aroma substances, elu-
cidation of the absolute configuration and
determination of the enantiomeric ratio, which is
usually given as the enantiomeric excess (ee), are
of especial interest because the enantiomers of
a compound can differ considerably in their odor
quality and threshold. The compound 3a,4,5,7a-