568 Notes and brief articles

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MASON, P. A. (1980). Aseptic synthesis of sheath ing
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U .K. : Blackwell Scientific Publications.
MOLINA, R. (1979). Pure culture synthesis and host
specificity of red alder mycorrhizae. Canadian Journal
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25 1-254.

ODOUR COMPOSITION OF THE SUMMER TRUFFLE, TUBER AESTIVUM

BY C. BELLINA-AGOSTINONE AND M. D'ANTONIO

Laboratorio Multizonale Igiene e Profilassi, USSL, L'Aquila, Italy

AND G. PACIONI
Dipartimento Scienze Ambientali, Uniuersita, L'Aquila, Italy

Volatile substances from Tuber aestioum have been studied by gas chromatography coupled
to a mass spectrometer (G C-M S). Thiobismethane is the main compound, mixed with
oxygen-containing organic compounds with molecular masses between 46 and 88. The
qualitative composition of odour could be useful as a taxonomic character.

During their evolution, hypogeous fungi have lost
the ability for active discharge of spores, but they
can attract animals that feed on their fruit bodies
spreading spores with faeces. Probably this attrac-
tion is mediated by volatile substances or odours.
Some hypogeous fungi, called truffles, are very
attractive to human beings too, and are amongst
the most expensive foods in the world. Truffle
odours have both ecological and economic interest;
in spite of this they are generally unknown.
Summer truffle (Tuber aestivum Vitt. sensu lato)
is the most widespread edible truffle, growing in
many European countries and in Algeria, and it
is possible that several distinct species may be
included within this binomial.
Several fresh specimens of Tuber aestivum sensu
stricto from seven collections, collected during
different periods of the year under both oak and
beech trees in central Italy, have been studied with
a gas chromatograph-mass spectrometer (G C-M S),
Hewlett-Packard Model 5986, computerized with
HP 9825B, using a 6 ft 0'1 % SPI000 on 80/100
Carbopack-C column programmed from 40 to
220°C, rate 10°Imino Chromatogram time was
30 min. Single compounds have been identified
using computer methods for comparing the
normalized mass spectra of specific peaks with data
recorded in computer memory and those available
Trans. Br . mycol. Soc. 88 (4), (1987) Printed in Great Britain
in the literature. Merck's standards for all possible
compounds, except ethanol, have been used.
Samples were kept in small airtight bottles.
Volatile substances were studied using head-space
analysis, i.e, sampling the air in the bottles with a
gas syringe at room temperature and then heating
quickly to a temperature of 50°, with the aim of
increasing the peaks of higher-boiling substances.
The sampled air containing volatile substances was
injected into the helium stream going into the
analyser system.
The flavour was shown to contain a mixture of
several substances. In the tail of the head-space
peak, the presence of ethanol is evident. Then, at
a retention time of about 3 min, we have found a
large peak (sometimes 50 % of the total amount) of
thiobismethane, which seems to be mainly respon-
sible for the smell. Thiobismethane has been
reported as a product of the fungal metabolism
resulting in the reduction of inorganic sulphates
(Birkinshaw, Findlay & Webb, 1942; Challenger &
Charlton, 1947).
Another organic sulphide, bis-rnethylthio-
methane, has been identified as the main compound
of the flavour impact of the white truffle ( Tuber
magnatum Pico) (F iecch i et al., 1967).
A number of oxygen-containing organic com-
pounds, probably produced via the pyruvate
 Notes and brief articles
Table 1. Volatile substances from fresh sporocarps of Tuber aestivum
Name" Structure Retention timet
Ethanol C "H.OH 0'5
Thiobismethane CH,SCH, 1"0
z-Methylpropanal (CH,)"CH . CHO 1·6
z-Butanone CH,CO .CHzCH , 1'9
z-Methyl-r-propenol (CH 3 )zC H . CH"OH 2'3
z-Methylbutanal CH,CH"CH(CH 3) . CHO 3'0
3- M ethylbutanal (CH , )"CH . CH"CHO 3'1
2- Methyl-a-butanol CH,CH"CH(CH 3) · CH"OH 3'5
3- M ethyl -1-butanol (CH 3 )"CH . CH"CH"OH 37
" From Chemical A bstracts. t Retention time related to thiobismethane (3 min).

pathway or leucine metabolism (Vickery & Vickery ,

1981) also appear. They give a pungent smell to
the summer truffle taste. An additional substance,
sometimes as plentiful as thiobismethane, is
z-butanone. Other substances identified include:
2-methylpropanal, 2 methyl-r-propanol, z-rnethyl-
butanal, 3-methylbutanal, z-rnethyl-r-butanol,
3-methyl-1-butanol. These results are summarized
in Table 1.
Repeating the GC-MS analyses several times we
obtained the same qualitative results, with some
variation only in the relative amounts.
Our results show that the odour of mature truffle
is a constant character that might be used for
taxonomic purposes.
REFERENCES
BIRKINSHAW, J. H., FINDLAY, W . P. K. & WEBB, R. A.
(1942). Biochemistry of the wood-rotting fungi. 3. The
production of methyl mercaptan by Schizophyllum
commune Fr. Biochemical Journal 36, 526-529.
CHALLENGER, F. & CHARLTON, P. T. (1947). Studies on
biol ogical methylation. Part X . The fission of the mono-
and di-sulphide links by moulds. Journal of the
Chemical Society 424-429.
FIECCHI, A., GALLI-KIENLE, M., SCALA, A. & CABELLA, P.
(1967). Bis-methylthiornethane, an odorous substance
from white truffle, Tuber magnatum Pico. Tetrahedron
Letters 18, 1681-1682.
VICKERY, M. L. & VICKERY, B. ( 1981). Secondary Plant
M etabolism. London : The Macmillan Press.

A NOTE ON USE AND INTERPRETATION OF NITRITE ASSIMILATION

TESTS IN PENICILLIUM SYSTEMATICS

BY P . D. BRIDGE
CAB International Mycological Institute, Ferry Lane, Ke w, Surrey, TW9 lAF

The effects of nitrite concentration and pH on the results of nitrite assimilation tests were
investigated with three strains of Penicillium. Apparent nitrite assimilation may not be directly
related to nitrite utilization but may be due in part to the uptake of nitrite. Although two
strains may both give positive reactions for nitrite assimilation, the underlying mechanisms
may be different.
Physiological and biochemical characters have been to nitrite and then subsequent reductions to
included in a number of taxonomic schemes for the ammonia (Pateman & Kinghorn, 1976), the failure
genus Penicillium, with the ability to utilize nitrite of nitrate-utilizing fungi to utilize nitrite generally
as sole nitrogen source being considered by several being attributed to the toxicity of the nitrite ion
authors (Abe, 1956; Frisvad, 1981; Bridge et aI., (Cochrane, 1958; Pateman & Kinghorn, 1976). It
1986). This feature may be a useful taxonomic is reasonable to assume that tests for nitrite
criterion, for although the majority of species in the utilization in nitrate-utilizing fungi may well
genus can utilize nitrate as the sole nitrogen source depend upon this toxicity, rather than true
(Domsch, Gams & Anderson, 1980; Onions, utilization. In this case a number of factors will
Allsopp & Eggins, 1981), relatively few species are contribute to the final result, such as pH and nitrite
reported to utilize nitrite (Abe, 1956; Frisvad, concentration. These factors have varied in taxo-
1981). nomic studies with nitrite concentrations varying
Nitrate utilization involves a reduction of nitrate from 22 mM (Abe, 1956) to 43 mM (Frisvad, 1981),
T rans. Br. mycol. S oc. 88 (4), (1987) Printed in Great Britain