Directory UMM :Data Elmu:jurnal:B:Biochemical Systematics and Ecology:Vol29.Issue1.Jan2001:

Biochemical Systematics and Ecology 29 (2001) 77}104

Chemical variability of peel and leaf essential oils
of 15 species of mandarins
Marie-Laure Lota, Dominique de Rocca Serra, FeH lix Tomi,
Joseph Casanova*
Universite& de Corse - Equipe Chimie et Biomasse, URA CNRS 2053, Route des Sanguinaires, 20000 Ajaccio,
France
Received 1 December 1999; received in revised form 24 February 2000; accepted 28 February 2000

Abstract
Peel and leaf oils of 58 mandarin cultivars, belonging to 15 di!erent species were obtained
from fruits and leaves collected on mandarin-trees submitted to the same pedoclimatic and
cultural conditions. Their chemical composition was investigated by capillary GC, GC/MS and
13C NMR and the results were submitted to a cluster analysis and a discriminant analysis.
Three major chemotypes, limonene, limonene/c-terpinene and linalyl acetate/limonene, were
distinguished for peel oils while three other chemotypes, sabinene/linalool, c-terpinene/linalool
and methyl N-methylanthranilate, were observed for leaf oils. ( 2000 Elsevier Science Ltd. All
rights reserved.
Keywords: Rutaceae; Citrus; Mandarin; Peel oil; Leaf oil; Essential oil composition; GC; GC/MS;
13C NMR; Statistical analysis

1. Introduction
Mandarins (Rutaceae family, Citrus genus) predominate with oranges the fresh fruit
market. According to Tanaka (1961), they are classi"ed into more than 30 species,
comprising from one to several tens of varieties. Cultivars of mandarin present a great
diversity of morphological and horticultural characters.

* Corresponding author. Tel.: 33-4-95-52-41-21; fax: 33-4-95-52-41-42.
E-mail address: casanova@vignola.univ-corse.fr (J. Casanova).
0305-1978/00/$ - see front matter ( 2000 Elsevier Science Ltd. All rights reserved.
PII: S 0 3 0 5 - 1 9 7 8 ( 0 0 ) 0 0 0 2 9 - 6

78

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

As for most of the Citrus, mandarin peel oil and leaf or `petit graina oil can be
obtained, respectively, by cold pressing and hydrodistillation of the fresh material.
Studies concerning the chemical composition of peel and leaf oils of mandarins have
been reviewed (Shaw, 1979; Lawrence, 1995a, b, 1996; Mondello et al., 1997). Nevertheless, in most cases, the species and/or the varieties remained unspeci"ed and the

results are not useful from the taxonomic point of view.
The aim of our work was to study the inter- and intraspeci"c chemical variability of
the large mandarin group. We divided our work into two parts:
f the "rst concerned 41 cultivars from C. reticulata Blanco, (Lota et al., 2000);
f the second dealt with 15 other species of mandarin, including C. clementina Hort. ex
Tan.
We report our results on the chemical variability of these 15 species represented by
58 cultivars of mandarin. We will compare the chemical composition of peel oils on
the one hand and that of leaf oils on the other. We will discuss chemical variability
including our previous results concerning cultivars from C. reticulata Blanco (Lota
et al., 2000).

2. Materials and methods
2.1. Plant materials
Clonal propagated trees, grafted on Troyer citrange rootstock, were 12 years old
and grown in the same pedoclimatic and cultural conditions in the germplasm
collection orchard of the `Station de Recherches Agronomiquesa of INRA-CIRAD,
located at San Ghjulianu (Corsica, France). The Citrus varieties collection of
INRA-CIRAD in Corsica is one of the FAO recognized Citrus collection in the
world. In this arboretum, each tree has a computerized identi"cation number. Geographic and climatic characteristics were: average per year: rainfall 840 mm and

temperature 15.23C, soil derived from alluvial deposits and classi"ed as fersiallitic, pH
range 5.0}5.6. Trees were in good vigor, disease-free and without visible insect
infestation.
2.2. Sampling, peel and leaf essential oil
For each cultivar of mandarin, about 500 g of leaves from the last autumn leaf #ush
and at least 30 ripe fruits were collected from many parts round the same tree, early in
the morning and only by dry weather during the period November 1996 to April 1997.
The peel of fresh fruits was cold-pressed and then the essential oil was separated
from the crude-extract by centrifugation (10 min at 15000 rpm). Fresh leaves were
subjected to hydrodistillation for 3 h using a Clevenger-type apparatus. Yield ranged
between 0.05 and 0.60%.

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

79

2.3. GC, GC/MS and 13C NMR analyses
Identi"cation of components and data processing were carried out as previously
reported for peel and leaf oils from C. reticulata (Lota et al., 2000). All peel and leaf oils
were investigated by GC. Five peel oils and seven leaf oils were analysed by GC/MS

while 23 peel oils and 25 leaf oils were analysed by 13C NMR, following a methodology "rst reported by FormaH cek and Kubeczka (1982), developed in our laboratory
(Tomi et al., 1995) and well-suited for chemical polymorphism studies (Salgueiro et al.,
1997; Corticchiato et al., 1998; Castola et al., 2000). Samples submitted to GC/MS
and/or 13C NMR analysis were selected on the basis of their chromatographic pro"le.
Note that 56 peel oils instead of 58 were analysed because the mandarin-trees of
`ougona and `shekwashaa cultivars do not produce fruits in the climatic conditions of
San Ghjulianu.
2.4. Data analyses
The data (components *1% for peel and leaf oils, respectively) were processed by
cluster analysis using hierarchical clustering (Ward`s technique and Euclidean distance measure) and were submitted to discriminant analysis. These processing were
performed with the xlStat-pro software.

3. Results
The 58 following cultivars, belonging to 15 species, were investigated. For the
convenience of comparison of the present results with the previous ones for 41
varieties from C. reticulata (Lota et al., 2000), we numbered the samples of this study
from 42 to 99.
Citrus clementina Hort. ex Tan. species : MA3 (no. 42), Nules (no. 43), MA2 (no. 44),
Hernandina (no. 45), Tardia Villareal (no. 46), Reina (no. 47), Ca$n (no. 48), MacBean
(no. 49), Oroval (no. 50), Monreal (no. 51), Bruno (no. 52), Tomatera (no. 53),

Commune (no. 54), Marisol (no. 55), Ragheb (no. 56), Guillermina (no. 57),
C. deliciosa Ten. species : Late Emperor (no. 58), Empress (no. 59), Emperor (no. 62),
Peau rugueuse (no. 63), Peau lisse (no. 76), Commune (no. 79), de Chios (no. 92),
Avana Apireno (no. 93), Willow leaf (no. 90), Tardivo di Ciaculli (no. 94),
C. nobilis Lour. species : Geleking (no. 74), Yellowking (no. 75), King of Siam (no.
80), Du Japon (no. 85), King (no. 86), Rode king (no. 87), Kunembo (no. 89),
C. tangerina Hort. ex Tan. species : Vohangisahy (no. 60), Beauty of Glen Retreat
(no. 61), Brickaville (no. 64), Dancy (no. 66), Redskin (no. 68), Swatow (no. 78),
C. unshiu Mac. Mark. species : Wase (no. 65), Clausellina (no. 70), URSS (no. 71),
Owari (no. 72),
C. suhuiensis Hort. ex Tan. species : Sihue Gan (no. 73), Szibat (no. 91), Szinkom
(no. 96);
C. temple Hort. ex Y. Tan. species : Temple Sue Linda (no. 81), Temple]Temple
(no. 83), Temple (no. 84),

80

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

C. paratangerina Hort. ex Tan. species : Ladu (no. 69), Ladu Ordinary (no. 77),

C. amblycarpa Hassk. Ochse. species : Nasnaran (no. 88),
C. depressa Hay. species : Shekwasha (no. 98),
C. erythrosa Hort. ex Tan. species : Fuzhu (no. 67),
C. reshni Hort. ex Tan. species : Cleopatra (no. 82),
C. suavissima Hort. ex Tan. species : Ougon (no. 99),
C. sunki Hort. ex Tan. : Sunki (no. 95),
C. yatsushiro Hort. ex Tan. : Yatsushiro (no. 97).
In order to simplify the discussion, we will "rst describe our results for C. clementina
Hort. ex Tan. All other taxa will be reported in Section 3.2.
3.1. Citrus clementina Hort. ex Tan.
3.1.1. Peel oils
The chemical composition of the 16 investigated samples are presented in Table 1.
The 30 identi"ed components accounted for 97.3}99.5% of the total amount of oil.
Peel oils consisted almost exclusively of hydrocarbons with limonene as the major
component (89.1}95.5%) with sabinene (0.3}4.0%) and myrcene (1.4}2.0%). a-Pinene,
b-phellandrene, b-pinene, (E)-b-ocimene, 3-carene and c-terpinene were identi"ed in
almost all samples at low amounts (tr-0.6%). The oxygenated fraction was made up of
linalool (0.6}2.3%), octanal, decanal, citronellal, a-terpineol, a-sinensal and b-sinensal
()0.7% for each one).
The homogeneous composition of our 16 samples is similar to that reported in the

literature (percentage of limonene: 92}97%) for `communea and `nulesa cultivars
from Uruguay (Verzera et al., 1998), for `orovala, `monreala and `communea cultivars
from Italy (Calabria) (Verzera et al., 1997) and for unspeci"ed cultivars from Italy
(Calabria, Sicily) (Mondello et al., 1995), Algeria (Baaliouamer et al., 1992) and
unspeci"ed cultivars from unspeci"ed origin (Calvarano et al., 1974; Huet, 1991;
Gazea et al., 1998; Ruberto et al., 1993, 1994, 1997).
3.1.2. Leaf oils
The 45 identi"ed components accounted for 96.1 to 99.8% of the total amount of
oil (Table 2). All samples exhibited a high sabinene/linalool composition
(33.1}49.8%/16.6}24.7%). The other main components of the ole"nic fraction
(20.9}28.2%) were limonene, 3-carene, (E)-b-ocimene, myrcene, b-pinene, c-terpinene,
a-pinene, terpinolene, a-terpinene and b-phellandrene (0.8}6.9% each). Terpinen-4-ol,
a-terpineol, trans-sabinene hydrate, citronellal, citronellol, geranyl acetate, a-sinensal
and b-sinensal were also identi"ed in almost all samples (0.1}4.8%). The oxygenated
fraction represented less than 40% of the whole oil.
The composition of a few leaf oils from clementin are reported in the literature.
Three samples (unspeci"ed cultivar) from Italy were characterized by a bpinene/linalool composition (approximately 45%/15%) (Di Giacomo et al., 1982).
Spanish oils from 12 cultivars `rufatinaa, `clemennullesa, `clemenvillaa, `esbala,
`"naa, `guillermaa, `hernandinaa, `monreala, `orovala and `tomateraa exhibited
a major component of unknown structure (19.2}48.8%) associated with linalool

Table 1
Chemical composition of clementin peel oils!
BP-20

BP-1

42

43

44

45

46

47

48

49

50

51

52

53

54

55

56

57

a-Pinene

b-Pinene
Sabinene
3-Carene
Myrcene
Limonene
b-Phellandrene
c-Terpinene
(E)-b-Ocimene
Octanal
trans-Sabinene hydrate
Citronellal
a-Copaene
Decanal
Linalool
trans-a-Bergamotene
(E)-b-Farnesene
a-Terpineol
Germacrene-D
Geranial
d-Cadinene

b-Sinensal
a-Sinensal

1021
1109
1119
1145
1157
1199
1208
1241
1245
1285
1458
1473
1484
1492
1539
1580
1654
1688
1704
1742
1746
2225
2323

930
971
964
1005
978
1021
1021
1048
1035
978
1053
1130
1379
1183
1082
1432
1448
1172
1480
1242
1516
1673
1726

0.3
tr
0.3
*
1.6
95.5
0.3
tr
tr
0.1
*
tr
tr
0.2
0.8
*
*
0.1
tr
*
tr
tr
0.1

0.4
tr
0.7
tr
1.7
95.0
0.3
tr
tr
0.1
tr
tr
tr
0.2
0.6
*
*
tr
tr
*
tr
0.1
0.3

0.3
tr
0.4
0.1
1.6
95.0
0.3
tr
tr
tr
*
*
tr
0.1
0.8
*
*
tr
tr
tr
tr
tr
0.2

0.3
tr
0.6
tr
1.4
94.6
0.2
*
tr
0.1
tr
tr
tr
0.1
0.8
tr
tr
0.1
tr
tr
*
0.1
0.1

0.4
tr
1.0
tr
1.6
94.1
0.3
tr
tr
0.3
tr
tr
tr
0.2
1.2
*
*
0.1
tr
tr
tr
tr
0.2

0.3
tr
0.9
tr
1.6
94.1
0.3
tr
tr
0.2
tr
0.1
tr
0.3
0.9
tr
tr
0.1
tr
*
*
0.1
0.3

0.3
tr
0.9
tr
1.7
94.1
0.3
tr
0.2
0.2
tr
tr
tr
0.2
0.8
tr
tr
tr
tr
*
*
0.2
0.4

0.4
t r
0.6
0.1
1.6
93.9
0.3
0.1
tr
*
tr
tr
0.1
0.2
1.0
0.1
*
0.1
tr
0.1
0.1
*
0.4

0.3
tr
0.9
0.1
1.6
93.5
0.3
tr
tr
0.2
tr
0.1
tr
0.2
1.5
tr
*
0.1
tr
*
tr
0.1
0.4

0.5
0.1
1.3
0.1
1.8
92.9
0.3
0.1
0.1
0.2
tr
tr
tr
0.2
0.8
*
*
tr
tr
0.1
tr
0.1
0.3

0.5
0.1
1.8
0.1
1.8
92.7
0.3
tr
0.1
0.3
tr
tr
tr
0.2
1.0
*
*
0.1
tr
*
tr
0.1
0.4

0.5
0.1
1.8
0.1
1.8
92.2
0.3
0.1
0.1
0.3
tr
tr
tr
0.2
1.1
*
*
0.1
tr
*
*
0.1
-

0.5
0.1
1.2
*
1.7
91.8
0.3
tr
tr
0.3
*
tr
tr
0.2
0.8
tr
tr
*
*
*
*
0.1
0.3

0.3
0.1
1.8
tr
1.6
91.2
0.3
tr
tr
0.3
0.1
0.1
tr
0.2
2.3
tr
tr
0.1
tr
*
tr
0.1
0.5

0.6
0.2
4.0
0.1
2.0
90.1
0.3
tr
0.1
0.2
tr
tr
tr
0.2
0.7
tr
tr
0.1
tr
0.1
*
0.2
0.4

*
0.1
2.8
0.1
1.6
89.1
0.3
tr
0.1
0.1
0.2
0.1
0.1
0.5
1.9
*
0.1
0.2
0.1
*
0.1
0.3
0.7

99.3

99.4

98.8

98.4

99.4

99.2

99.3

99.1

99.3

98.9

99.5

98.8

97.3

99.0

99.3

98.5

Total

81

!Other compounds ( tr): p-cymene, a-phellandrene, terpinolene, nonanal, cis-limonene-1,2-oxide, trans-limonene-1,2-oxide, citronellol.
Cultivars : MA3 (42), Nules (43), MA2 (44), Hernandina (45), Tardia Villareal (46), Reina (47), Ca$n (48), Mac Bean (49), Oroval (50), Monreal (51), Bruno
(52), Tomatera (53), Commune (54), Marisol (55), Ragheb (56), Guillerma (57).
Order of elution and percentages of components are given on BP-20 column. All the components were identi"ed by GC-RI on polar and apolar columns. All
the compounds of samples no. 49 and 54 were also identi"ed by GC/MS. The major components (bold letters) of samples no. 51, 53, 54, 56 and 57 were identi"ed
by 13C NMR.

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Constituents

82

Constituents

BP-20

BP-1

56

43

47

53

46

52

55

45

44

48

51

49

54

42

50

57

a-ThujeneH
a-PineneH
Camphene
b-Pinene
Sabinene
3-Carene
Myrcene
a-Phellandrene
a-Terpinene
Limonene
b-Phellandrene
(Z)-b-Ocimene
c-Terpinene
(E)-b-Ocimene
p-Cymene
Terpinolene
6-Methylhept-5en-2-one
trans-Sabinene hydrate
Citronellal
Decanal
Linalool
cis-p-Menth-2-en-1-ol

1021
1021
1066
1109
1119
1145
1157
1162
1179
1199
1208
1228
1241
1245
1267
1278
1332

922
930
944
971
964
1005
978
997
1009
1021
1021
1024
1048
1035
1012
1078
963

0.3
1.3
tr
1.7
33.1
4.5
2.8
0.5
1.3
6.9
0.8
0.2
2.1
3.8
tr
1.2
tr

0.3
1.6
tr
1.8
35.1
6.5
3.3
0.7
1.3
2.5
0.9
0.2
1.9
5.5
tr
1.5
0.1

0.3
1.4
tr
1.7
34.3
6.5
3.2
0.6
0.9
2.7
0.8
0.2
1.5
5.6
0.3
1.4
tr

0.3
1.4
0.1
1.8
35.2
6.2
3.2
0.6
1.2
3.1
0.8
0.2
1.9
4.8
tr
1.5
0.1

0.4
1.1
tr
1.9
38.1
4.8
3.0
0.5
1.0
2.8
0.8
0.2
1.6
4.8
0.2
1.2
tr

0.4
1.5
tr
1.9
37.9
4.4
3.1
0.5
1.4
2.9
0.8
0.2
2.2
5.4
tr
1.2
*

0.3
0.7
tr
1.8
36.9
5.1
3.0
0.5
1.0
4.3
0.8
0.2
1.5
5.4
0.1
1.2
tr

0.3
1.6
tr
2.2
40.1
3.8
3.1
0.4
1.2
4.8
0.8
0.1
1.9
3.8
tr
1.1
*

0.3
1.5
tr
1.9
39.9
3.8
3.0
0.4
1.0
3.8
0.8
0.1
1.5
3.9
tr
1.0
*

0.3
1.5
tr
2.0
41.8
2.6
3.0
0.3
1.2
3.3
0.8
0.2
2.0
4.2
tr
0.9
-

0.4
0.8
tr
2.1
43.5
3.3
3.0
0.4
1.2
3.6
0.8
0.1
2.0
3.6
tr
1.0
*

0.4
1.7
tr
2.1
42.5
3.8
3.2
0.4
1.2
2.9
0.8
0.2
1.9
4.4
tr
1.1
*

0.4
1.5
tr
2.1
42.0
4.1
3.3
0.4
1.2
3.5
0.8
0.2
1.9
4.5
0.1
1.1
*

0.4
1.6
tr
2.0
41.9
3.8
3.2
0.4
1.0
4.0
0.8
0.2
1.6
4.6
tr
1.0
*

0.3
1.5
tr
2.2
47.0
2.4
3.0
0.3
0.9
4.2
0.8
0.1
1.5
3.8
tr
0.8
*

0.4
1.2
tr
2.3
49.8
3.0
3.2
0.4
1.1
3.0
0.9
0.1
1.7
2.5
0.1
1.0
tr

1458
1473
1492
1539
1555

1053
1130
1183
1082
1107

0.7
3.4
tr
23.5
0.3

0.6
1.6
tr
19.4
0.3

1.0
3.7
*
21.7
0.2

0.9
3.0
tr
22.9
0.3

1.4
1.4
*
24.7
0.3

0.9
1.2
tr
22.7
0.3

1.0
2.2
tr
21.8
0.2

0.8
1.5
tr
22.6
0.3

1.2
1.5
tr
23.9
0.2

0.9
0.9
tr
24.2
0.3

0.9
1.8
*
21.5
0.3

1.0
1.6
*
20.5
0.3

1.2
0.9
tr
21.0
0.3

1.0
1.1
0.1
21.6
0.2

1.1
0.3
0.1
22.0
0.2

1.3
1.7
tr
16.6
0.3

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Table 2
Chemical composition of clementin leaf oils!

Total

1588
1595
1618

1420
1162
1123

0.1
4.6
0.2

0.1
4.6
0.2

0.1
3.6
0.1

0.1
3.7
0.1

0.1
4.3
0.2

0.1
4.8
0.2

0.1
3.6
0.1

0.1
4.0
0.1

0.1
3.5
0.1

0.1
4.3
0.1

tr
4.6
0.2

0.1
4.2
0.1

0.1
4.1
0.1

0.2
3.6
0.1

0.1
3.3
0.1

tr
4.5
0.1

1654
1674
1688
1725
1727
1742
1748
1756
1790
1837
2031
2225
2323

1448
1213
1172
1340
1494
1242
1358
1207
1207
1232
1547
1673
1726

tr
0.2
1.1
0.1
0.1
0.3
0.7
0.7
0.2
0.1
0.1
2.0
0.4

tr
0.6
1.4
*
0.1
0.8
0.4
0.3
0.2
0.5
tr
1.2
0.6

tr
0.4
1.2
*
*
0.6
0.5
0.7
0.2
0.1
0.1
2.2
1.1

*
0.4
1.2
0.1
*
0.6
0.4
0.5
0.2
0.1
0.1
1.8
0.9

tr
0.1
1.3
*
*
0.2
0.2
0.3
0.1
tr
0.1
1.6
0.8

*
0.2
1.0
0.3
*
0.1
0.2
0.3
0.1
0.1
0.1
1.9
1.0

tr
0.3
1.0
0.4
*
*
1.0
0.6
0.1
0.2
0.1
2.0
1.0

*
tr
0.8
*
*
*
0.1
0.4
0.1
tr
0.1
2.1
0.7

0.1
0.1
1.1
*
*
0.2
0.1
0.4
0.2
tr
0.1
2.1
0.8

*
tr
0.7
0.1
*
0.1
0.1
0.3
0.1
tr
0.1
2.5
0.8

tr
0.1
0.7
*
*
0.1
0.2
0.4
0.1
tr
tr
1.8
0.6

tr
tr
0.7
tr
0.1
*
0.2
0.3
0.1
tr
0.1
2.2
0.7

*
0.1
0.8
*
tr
0.1
0.1
0.3
0.1
tr
0.1
2.2
1.0

0.1
tr
0.8
*
tr
*
0.1
0.3
0.1
tr
0.1
2.6
1.0

tr
tr
0.7
*
*
tr
0.1
0.1
tr
tr
0.1
2.2
0.9

*
0.1
1.1
0.2
*
0.1
0.2
0.5
0.1
tr
tr
0.8
0.3

99.2

96.1

98.9

99.8

99.5

99.3

98.5

98.9

98.6

99.7

99.1

98.8

99.8

99.5

99.6

98.6

!Other compounds (tr): octanal, nonanal, citronellyl acetate, a-humulene, (E,E)-a-farnesene, d-cadinene, thymol.
Cultivars : Ragheb (56), Nules (43), Reina (47), Tomatera (53), Tardia Villareal (46), Bruno (52), Marisol (55), Hernandina (45), MA2 (44), Ca$n (48), Monreal
(51), Mac Bean (49), Commune (54), MA3 (42), Oroval (50), Guillerma (57).
Order of elution and percentages of components are given on BP-20 column, except compounds with an asterisk (percentages given on BP-1 column). All the
components were identi"ed by GC-RI on polar and apolar columns. All the compounds of samples no. 47, 54 and 56 were also identi"ed by GC/MS. The major
components (bold letters) of samples no. 50, 53, 54 and 56 were identi"ed by 13C NMR.

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

(E)-Caryophyllene
Terpinen-4-ol
trans-p-Menth2-en-1-ol
(E)-b-Farnesene
Neral
a-Terpineol
Neryl acetate
Bicyclogermacrene
Geranial
Geranyl acetate
Citronellol
Nerol
Geraniol
(E)-Nerolidol
b-Sinensal
a-Sinensal

83

84

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

(16.7}27.1%) (Ortiz Marcide et al., 1983). It is likely if not certain that the unidenti"ed
major component reported by Ortiz Marcide is sabinene. The sabinene/linalool
chemotype, found in our sampling, is con"rmed unambiguously for the "rst time for
C. clementina leaf oils in this study.
3.2. The other species of mandarin
3.2.1. Peel oils
The total of the 44 identi"ed components accounted for 95.8}99.7% of the oil
(Table 3). Even though the composition was dominated by limonene (55.8}96.7%) for
39 samples over 40, the content of the major components varied considerably from
sample to sample. Several other monoterpene hydrocarbons were frequently identi"ed at appreciable contents: c-terpinene (tr-19.9%), p-cymene (tr-12.0%), myrcene
(0.7}24.0%), b-pinene (tr-14.2%), sabinene (0.1}8.7%), a-pinene (0.2}2.2%) and bphellandrene (0.2}0.8%). Among the oxygenated compounds, linalool was present in
all the samples (0.1}10.7%) whereas percentages of octanal, a-terpineol and decanal
were not over 0.5%. The contents of citronellal (9.9%) and of linalyl acetate (48.7%)
were important, respectively, in two samples (no. 88, `Nasnarana cultivar from C.
amblycarpa species and no. 97, `Yatsushiroa cultivar from C. yatsushiro species).
The dendrogram, obtained from cluster analysis and reported in Fig. 1, suggested
the existence of three principal clusters (I, II and III) within the essential oil of the
individuals of mandarin. Discriminant analysis (Fig. 2) con"rmed this clustering with
respect to the contents of limonene, c-terpinene and linalyl acetate (Fig. 3).
Limonene chemotype : most of the peel oils (no. 58}87) were characterized by a very
high amount of limonene (87.1}96.7%). This chemotype (cluster I) could be divided
into two subgroups on the basis of the content of c-terpinene, which was very low
()0.9%) in the subgroup IA (samples no. 79}87) and higher (3.5}5.8%) in the
subgroup IB (samples no. 58}78)
Limonene/c-terpinene chemotype: nine samples (no. 88}96) belonged to this chemotype (cluster II). The composition was dominated by limonene (55.8}79.0%) associated with c-terpinene (0.1}19.9%) and p-cymene (0}12.0%). Three samples (no. 88,
89 and 96), which belonged to the same group, exhibited quantitative di!erences in
their composition. Indeed, the sample no. 88 was discriminated by higher contents of
b-pinene (14.2% vs 0.2}1.6%), sabinene (8.7% vs 0.1}2.1%) and citronellal (9.9% vs
tr-0.1%). Similarly, samples no. 89 and 96 were distinguished, respectively, by important amounts of myrcene (24.0% vs 1.2}1.7%) and p-cymene (12.0% vs 0}6.9%).
Linalyl acetate/limonene chemotype: only one sample (no. 97) belonged to the cluster
III and was characterized by the predominance of linalyl acetate (48.7%) over
limonene (22.8%), c-terpinene (5.6%) and b-pinene (5.4%).
A few studies are reported on the chemical composition of peel mandarin oils from
species other than C. reticulata and C. clementina. Essential oils of C. unshiu were
reviewed by Lawrence (1989) and divided into two chemotypes: (i) limonene
(78}96%) from `praecoxa cultivar from Japan (Lawrence, 1989), unspeci"ed cultivar
from Japan (Yamanishi et al., 1968; Yajima et al., 1979), from Georgia, Russia and
from unspeci"ed origin (Lawrence, 1989), (ii) limonene/p-cymene (42}79%/4}27%)

Table 3
Chemical composition of peel mandarin oils!
Constituents

Total

BP-1

IB
58

59

60

61

62

63

64

65

66

67

68

69

70

71

1021
1021
1109
1119
1157
1179
1199
1208
1241
1245
1267
1278
1285
1451

922
930
971
964
978
1009
1021
1021
1048
1035
1012
1078
978
1120

0.1
0.7
0.3
0.2
1.6
tr
91.4
0.3
3.4
tr
0.2
0.1
0.2
tr

0.1
0.6
0.3
0.2
1.6
0.1
90.8
0.2
4.0
0.1
0.3
0.2
tr
tr

0.2
0.9
0.4
0.1
1.7
0.1
90.9
*
3.9
0.1
0.4
0.2
0.1
tr

0.1
0.6
0.3
0.1
1.6
0.1
90.2
0.2
4.3
0.2
0.2
0.2
0.1
*

0.1
0.6
0.3
0.3
1.6
0.1
90.4
0.3
4.2
0.1
0.4
0.2
0.1
tr

0.1
0.4
0.2
0.1
1.5
0.1
90.5
0.3
4.3
0.2
0.5
0.2
*
tr

0.1
0.5
0.3
0.1
1.5
0.1
90.2
0.3
4.7
0.2
0.2
0.2
0.1
*

0.1
0.5
0.4
0.1
1.7
0.1
90.3
0.3
4.3
tr
0.1
0.2
tr
*

0.1
0.5
0.3
0.1
1.5
0.1
87.1
0.2
5.2
0.2
0.2
0.2
0.2
*-

0.2
0.6
0.4
0.1
1.6
0.1
88.4
0.2
4.9
0.1
0.1
0.2
0.2
*

0.1
0.7
0.3
0.1
1.6
0.1
89.2
0.2
4.4
0.1
0.3
0.2
0.1
*

0.2
0.9
0.4
0.1
1.7
tr
87.6
0.3
4.6
0.2
0.4
0.2
0.2
*

0.1
0.5
0.4
0.1
1.8
0.1
88.0
0.2
5.3
0.1
0.2
0.3
*
*

0.1
0.5
0.4
0.1
1.6
0.1
88.0
0.2
5.8
0.1
0.2
0.3
tr
*

1473
1539
1549
1586
1654
1688
1721
1725
1744
1748
1756
2082

1130
1082
1239
1388
1448
1172
1500
1340
1500
1358
1207
1385

*
0.5
*
*
*
0.1
*
*
*
tr
*
*

tr
0.6
*
tr
*
0.3
*
*
*
tr
tr
*-

*
0.4
*
*
*
tr
*
*
*
*
*
*

tr
0.7
*
*
*
0.1
*
tr
*
*
*
*

*
0.6
*
*
*
tr
*
*
*
tr
*
*

*
0.4
*
*
*
0.1
*
*
*
tr
tr
*

tr
0.6
*
*
*
0.1
*
*
*
*
*
*

tr
0.2
*
0.2
*
tr
*
*
0.3
*
*
*

tr
2.2
tr
0.1
*
0.1
0.1
*
*
tr
tr
*

tr
1.9
tr
tr
*
0.1
*
*
*
tr
*
*

*
1.7
*
*
*
0.1
*
*
*
*
*
*

tr
0.8
tr
*
*
0.1
*
*
*
tr
*
*

*
0.3
*
0.6
*
tr
*
*
0.7
*
*
*

tr
0.2
*
0.4
*
tr
*
*
0.6
*
*
*

2323

1726

tr

tr

0.1

0.2

0.2

0.2

0.2

0.1

0.1

0.2

*

*

99.3

99.5

99.6

99.3

99.2

99.5

98.7

99.3

99.4

98.0

99.2

99.1

*
99.5

*
99.1

85

*continued

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

a-ThujeneH
a-PineneH
b-Pinene
Sabinene
Myrcene
a-Terpinene
Limonene
b-Phellandrene
c-Terpinene
(E)-b-Ocimene
p-Cymene
Terpinolene
Octanal
trans-Limonene1,2-oxide
Citronellal
Linalool
Linalyl acetate
b-Elemene
(E)-b-Farnesene
a-Terpineol
b-Bisabolene
Neryl acetate
(E,E)-a-Farnesene
Geranyl acetate
Citronellol
Methyl Nmethylanthranilate
a-Sinensal

BP-20

86

Table 3*continued
IB

IA

BP-20

BP-1

72

73

74

75

76

77

78

79

80

81

82

83

84

85

a-ThujeneH
a-PineneH
b-Pinene
Sabinene
Myrcene
a-Terpinene
Limonene
b-Phellandrene
c-Terpinene
(E)-b-Ocimene
p-Cymene
Terpinolene
Octanal
trans-Limonene1,2-oxide
Citronellal
Linalool
Linalyl acetate
b-Elemene
(E)-b-Farnesene
a-Terpineol
b-Bisabolene
Neryl acetate
(E,E)-a-Farnesene
Geranyl acetate
Citronellol
Methyl Nmethylanthranilate
a-Sinensal

1021
1021
1109
1119
1157
1179
1199
1208
1241
1245
1267
1278
1285
1451

922
930
971
964
978
1009
1021
1021
1048
1035
1012
1078
978
1120

0.2
0.8
0.4
0.1
1.8
0.1
88.8
0.3
5.5
tr
0.2
0.3
*
*

0.2
0.7
0.4
0.1
1.7
0.1
89.1
0.2
5.3
0.2
0.2
0.2
0.1
*

0.1
0.6
0.3
0.2
1.6
0.1
89.2
0.3
5.1
0.5
0.2
0.2
0.3
*

0.1
0.5
0.3
0.2
1.6
0.1
89.2
0.3
5.1
0.6
0.2
0.2
tr
*

0.1
0.5
0.3
0.1
1.5
0.1
89.6
0.3
4.7
0.2
0.4
0.2
*
tr

0.2
0.8
0.4
0.1
1.7
tr
89.6
0.2
4.8
0.2
0.1
0.2
0.1
*

0.1
0.6
0.3
0.2
1.6
0.1
89.6
0.3
5.2
tr
0.1
0.2
0.1
tr

*
0.6
1.5
0.3
1.8
*
92.6
0.3
*
0.3
*
*
0.1
*

tr
0.9
0.2
0.4
1.5
tr
93.1
0.8
0.9
0.1
0.3
0.1
*
*

tr
0.5
tr
1.0
1.8
*
93.3
0.2
*
0.1
*
tr
0.4
*

*
0.3
tr
1.1
1.7
*
93.6
0.3
tr
0.1
*
tr
0.1
*

*
0.5
tr
0.4
1.7
*
93.9
0.3
tr
0.1
*
tr
0.3
*

tr
0.5
tr
0.9
1.7
*
90.7
0.2
tr
0.1
*
tr
0.4
*

*
0.4
tr
0.2
1.6
*
95.4
0.3
tr
*
*
*
0.4
*

1473
1539
1549
1586
1654
1688
1721
1725
1744
1748
1756
2082

1130
1082
1239
1388
1448
1172
1500
1340
1500
1358
1207
1385

tr
0.1
*
0.3
tr
*
*
*
*
*
*
*

tr
0.8
*
*
*
0.1
*
*
*
tr
*
*

tr
0.2
*
*
*
0.1
*
tr
tr
*
*
*

tr
0.2
*
*
*
0.1
*
*
*
*
*
*

tr
0.6
tr
*
*
0.1
*
*
*
*
tr
*

*
0.7
*
*
*
0.2
*
0.1
*
tr
*
*

tr
0.6
*
tr
*
0.1
*
tr
*
tr
tr
*

tr
0.1
*
*
tr
tr
tr
*
*
0.2
*
*

*
0.2
0.2
*
0.1
0.1
tr
*
*
*
*
*

tr
1.7
*
*
*
tr
*
*
*
*
*
*

*
1.0
*
*
*
0.1
*
*
*
*
*

tr
1.1
*
*
*
0.1
*
0.1
*
*
*
*

tr
4.0
tr
*
*
0.1
*
0.2
*
*
*
*

0.1
0.6
*
*
0.1
tr
*
*
*
*
*
*

2323

1726

*

*

*

0.3

0.1

tr

*

*

*

*

0.1

*

99.3

99.0

99.2

99.5

99.3

98.5

98.3

98.5

98.7

Total

99.5

0.1
99.6

tr
99.3

99.2

99.4

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Constituents

Constituents

Total

II

III

BP-20

BP-1

86

87

88

89

90

91

92

93

94

95

96

97

1021
1021
1109
1119
1157
1179
1199
1208
1241
1245
1267
1278
1285
1451

922
930
971
964
978
1009
1021
1021
1048
1035
1012
1078
978
1120

*
0.4
tr
0.1
1.7
*
96.7
0.3
tr
tr
tr
*
0.1
*

*
0.5
tr
0.1
1.7
*
96.7
0.3
tr
tr
*
*
0.1
*

0.1
1.3
14.2
8.7
1.7
*
55.8
0.5
0.1
0.2
*
tr
0.5
*

tr
0.2
0.2
0.1
24.0
0.1
63.7
0.3
5.8
*
0.2
0.3
*
*

0.5
1.3
1.3
2.1
1.6
0.2
65.3
0.2
15.9
*
6.9
0.7
0.1
tr

0.9
2.2
1.6
0.2
1.6
0.4
68.9
0.2
19.9
*
1.2
0.8
0.1
tr

0.6
1.9
1.3
0.2
1.7
0.4
73.1
0.2
17.3
*
0.5
0.8
0.1
*

0.6
0.7
1.3
0.2
1.7
0.3
75.3
0.2
16.7
tr
0.5
0.7
0.1
*

0.5
1.8
1.3
0.2
1.5
0.1
74.5
0.3
13.7
tr
2.8
0.5
tr
0.1

0.1
0.4
0.5
0.1
1.3
0.1
79.0
0.2
11.5
*
1.3
0.4
0.1
tr

0.4
1.7
1.3
0.3
1.2
*
75.0
0.4
0.7
*
12.0
tr
0.2
0.7

0.2
0.7
5.4
0.9
0.7
0.1
22.8
0.2
5.6
0.3
0.1
0.2
tr
*

1473
1539
1549
1586
1654
1688
1721
1725
1744
1748
1756
2082

1130
1082
1239
1388
1448
1172
1500
1340
1500
1358
1207
1385

*
0.1
*
*
tr
tr
*
*
tr
0.1
*
*

tr
0.1
*
*
tr
tr
*
*
*
0.1
*
*

9.9
0.4
tr
0.3
*
*
*
*
1.5
*
0.7
*

0.1
0.1
*
*
0.4
0.1
*
0.9
*
1.6
0.2
*

tr
0.6
*
tr
*
0.4
*
tr
*
*
*
1.3

tr
0.2
*
*
*
tr
*
*
0.9
tr
tr
*

tr
0.2
*
*
*
0.2
*
*
*
*
tr
0.4

tr
0.2
*
*
*
0.2
*
*
*
*
*
0.3

tr
0.2
tr
tr
*
0.2
*
*
*
tr
tr
*

0.1
1.4
*
0.3
*
0.2
*
0.1
*
0.2
tr
*

tr
0.8
*
*
*
0.3
0.3
*
*
tr
*
*

*
10.7
48.7
*
0.1
*
0.4
0.5
*
0.3
*
*

2323

1726

*

*

0.2

0.2

0.3

99.6

99.7

99.3

99.3

98.2

0.8
97.9

*
98.4

0.3
99.0

*
99.3

0.1

*
97.7

95.8

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

a-ThujeneH
a-PineneH
b-Pinene
Sabinene
Myrcene
a-Terpinene
Limonene
b-Phellandrene
c-Terpinene
(E)-b-Ocimene
p-Cymene
Terpinolene
Octanal
trans-Limonene1,2-oxide
Citronellal
Linalool
Linalyl acetate
b-Elemene
(E)-b-Farnesene
a-Terpineol
b-Bisabolene
Neryl acetate
(E,E)-a-Farnesene
Geranyl acetate
Citronellol
Methyl Nmethylanthranilate
a-Sinensal

IA

*
99.1
87

*¹able footnote continued overleaf

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

!Other compounds (tr-0.3%): camphene, a-phellandrene, (Z)-b-ocimene, nonanal, cis-limonene-1,2-oxide, trans-sabinene hydrate, octyl acetate, a-copaene,
decanal, trans-a-bergamotene, (E)-caryophyllene, terpinen-4-ol, a-humulene, neral, a-terpinyl acetate, germacrene-D, bicyclogermacrene.
Cultivars : Late Emperor (58, C. deliciosa), Empress (59, C. deliciosa), Vohangisahy (60, C. tangerina), Beauty of Glen Retreat (61, C. tangerina), Emperor (62,
C. deliciosa), Peau rugueuse (63, C. deliciosa), Brickaville (64, C. tangerina), Wase (65, C. unshiu), Dancy (66, C. tangerina), Fuzhu (67, C. erythrosa), Redskin (68,
C. tangerina), Ladu (69, C. paratangerina), Clauselina (70, C. unshiu), URSS (71, C. unshiu), Owari (72, C. unshiu), Sihue Gan (73, C. suhuiensis), Geleking (74, C.
nobilis), Yellowking (75, C. nobilis), Peau lisse (76, C. deliciosa), Ladu Ordinary (77, C. paratangerina), Swatow (78, C. tangerina), Commune (79, C. deliciosa),
King of Siam (80, C. nobilis), Temple Sue Linda (81, C. temple), Cleopatra (82, C. reshni), Temple x Temple (83, C. temple), Temple (84, C. temple), Du Japon (85,
C. nobilis), King (86, C. nobilis), Rode king (87, C. nobilis), Nasnaran (88, C. amblycarpa), Kunembo (89, C. nobilis), Willow leaf (90, C. deliciosa), Szibat (91, C.
suhuiensis), de Chios (92, C. deliciosa), Avana Apireno (93, C. deliciosa), Tardivo di Ciaculli (94, C. deliciosa), Sunki (95, C. sunki), Szinkom (96, C. suhuiensis),
Yatsushiro (97, C. yatsushiro).
Order of elution and percentages of components are given on BP-20 column, except compounds with an asterisk (percentages given on BP-1 column). All the
components were identi"ed by GC-RI on polar and apolar columns. All the compounds of samples no. 88, 96 and 97 were also identi"ed by GC/MS. The major
components (bold letters) of samples no. 65, 66, 67, 69, 75, 77, 78, 80, 82, 83, 84, 88, 90, 92, 93, 94, 96 and 97 were identi"ed by 13C NMR.

88

Table 3*footnote continued

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

89

Fig. 1. Dendrogram obtained from the cluster analysis of 56 mandarin peel oils. Samples are clustered
using Ward's technique with an Euclidean distance measure.

for four samples from an unspeci"ed japanese cultivar (Lawrence, 1989). To our
knowledge, only one study concerns C. deliciosa species. Two samples (unspeci"ed
variety) from Italy and Spain were characterized by the limonene/c-terpinene chemotype (72}77%/14}19%) (Boelens and Jimenez, 1989). It should be pointed out that
a very large number of commercial oils from Italy of certainly known but not reported
species and variety (400 samples, Dugo, 1994), as well as oils from Argentina
(Retamar, 1986) or from unspeci"ed origin (Lawrence, 1996) exhibited this last
chemotype.
3.2.2. Leaf oils
The 58 identi"ed components accounted for 97.1}99.9% of the total amount of oil
(Tables 4). We observed an important chemical variability with the occurrence of
sabinene (0.1}57.3%), c-terpinene (0.1}67.4%), linalool (tr-59.3%) and methyl Nmethylanthranilate (0}78.7%). For most samples (29 over 42), the content of monoterpene hydrocarbons was important. The best represented components were sabinene,
c-terpinene, myrcene, limonene and (E)-b-ocimene. Conversely, for the 13 other
samples, the oxygenated fraction was dominant with high contents of linalool, thymol
or methyl N-methylanthranilate.
The dendrogramm (Fig. 4) suggested the existence of three principal groups. The
discriminant analysis con"rmed this repartition with respect to the contents of
sabinene, c-terpinene, linalool and methyl N-methylanthranilate (Figs. 5 and 6).

90

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Fig. 2. Discriminant analysis scatterplott of 56 mandarin peel oils. (h) Cluster I; (L) Cluster II, (])
Cluster III.

Fig. 3. Discriminant analysis scatterplot of the peel oil constituents.

Table 4
Chemical composition of leaf mandarin oils!

a-ThujeneH
a-PineneH
b-Pinene
Sabinene
3-Carene
Myrcene
a-Terpinene
Limonene
b-Phellandrene
(Z)-b-Ocimene
c-Terpinene
(E)-b-Ocimene
p-Cymene
Terpinolene
p-Cymenene
trans-Sabinene hydrate
Citronellal
Decanal
Linalool
cis-p-Menth-2-en-1-ol
b-Elemene
(E)-Caryophyllene
Thymyl methyl ether
Terpinen-4-ol

BP-20

1021
1021
1109
1119
1145
1157
1179
1199
1208
1228
1241
1245
1267
1278
1432
1458
1473
1492
1539
1555
1586
1588
1589
1595

BP-1

922
930
971
964
1005
978
1009
1021
1021
1024
1048
1035
1012
1078
1072
1053
1130
1183
1082
1107
1388
1420
1214
1162

IB

IA

74

75

86

87

80

83

82

63

84

62

59

78

81

58

0.6
1.7
2.6
46.5
tr
3.5
1.7
1.8
0.7
0.5
4.3
16.0
0.6
0.8
0.1
0.9
*
*
5.4
0.5
0.5
0.2
*
7.4

0.8
2.1
2.9
47.8
tr
3.6
1.9
1.9
0.7
0.5
4.4
14.0
0.7
0.8
0.1
0.9
*
tr
3.4
0.5
0.4
0.2
*
7.4

0.5
0.9
3.4
55.5
tr
3.5
1.5
7.7
0.7
0.2
2.4
6.5
tr
0.6
tr
0.8
*
*
6.7
0.3
0.6
0.2
*
5.4

0.5
0.9
3.3
57.3
tr
3.8
1.6
5.6
0.7
0.2
2.8
6.6
tr
0.7
tr
0.7
*
*
5.3
0.4
0.8
0.2
*
6.1

0.5
1.3
3.5
56.0
tr
3.6
1.6
8.6
0.7
0.2
2.7
5.9
0.1
0.7
*
0.8
*
tr
3.1
0.4
0.7
0.2
*
6.6

0.5
2.2
2.7
55.6
tr
3.4
1.2
1.7
0.8
0.3
2.1
7.1
0.2
0.5
*
0.7
*
*
10.6
0.3
tr
0.3
*
4.5

0.5
1.9
2.7
49.7
tr
3.6
1.8
1.6
0.6
0.3
3.1
6.9
*
0.7
tr
0.7
*
tr
13.0
0.5
0.2
0.3
*
7.9

0.6
1.0
2.8
48.3
tr
3.2
1.2
2.9
0.5
0.4
2.8
6.9
0.4
0.6
0.4
0.7
tr
0.1
17.0
0.3
0.1
*
1.0
3.8

0.4
1.9
2.5
51.5
1.5
3.2
1.2
2.0
0.8
0.2
1.9
5.0
0.1
0.7
*
0.9
0.2
tr
17.2
0.3
*
0.2
*
4.3

0.5
1.9
2.3
37.9
tr
2.8
1.2
2.3
0.4
0.4
2.7
9.1
0.4
0.6
0.3
0.7
tr
tr
26.2
0.3
0.1
*
1.1
4.3

0.5
2.0
2.4
40.5
tr
2.9
0.9
1.8
0.4
0.4
2.3
9.0
0.4
0.5
0.4
0.6
tr
tr
25.5
0.2
0.1
*
1.2
3.2

1.2
1.4
2.3
37.8
tr
2.8
1.5
2.4
0.5
0.3
3.3
5.6
0.5
0.7
0.3
0.9
tr
0.1
25.6
0.4
0.1
*
1.0
6.2

0.3
1.4
1.9
39.0
2.2
2.7
1.1
1.4
0.7
0.3
1.8
6.2
0.1
0.7
*
0.8
0.5
tr
27.1
0.3
tr
*
*
4.2

0.5
1.8
2.2
37.4
tr
2.7
0.9
1.9
0.4
0.4
2.2
7.7
0.4
0.5
0.3
0.8
tr
tr
31.0
0.2
0.1
*
1.0
3.2

91

*continued

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Constituents

92

Table 4*continued

trans-p-Menth-2-en-1-ol
Citronellyl acetate
a-Humulene
Neral
a-Terpineol
a-Bisabolene
Neryl acetate
Bicyclogermacrene
Geranyl acetate
Citronellol
Nerol
Geraniol
Methyl Nmethylanthranilate
Thymol
b-Sinensal
a-Sinensal
Total

BP-20

BP-1

IB

IA

74

75

86

87

80

83

82

63

84

62

59

78

81

58

0.1
*
tr
*
0.2
0.4
*
*
*
*
tr
*
*

0.1
*
tr
tr
0.4
*
*
*
*
0.2
0.6
tr
*

0.1
*
tr
*
0.4
0.5
*
*
*
*
tr
*
*

0.1
*
*
0.1
0.3
0.4
*
*
*
*
*
*
*

0.2
*
tr
*
0.5
0.4
*
*
*
*
*
tr
*

0.1
tr
tr
0.3
0.7
*
0.3
*
*
0.2
0.8
0.1
*

0.1
*
tr
*
0.5
*
*
0.4
*
*
*
*
*

0.1
1.3
0.9

*
1.1
0.8

0.1
1.5
0.6

*
1.3
0.6

0.1
1.2
0.6

*
1.7
0.9

0.2
1.4
0.7

98.4

99.5

99.1

98.3

98.2

98.7

99.4

1618
1654
1660
1674
1688
1724
1725
1727
1748
1756
1790
1837
2082

1123
1332
1453
1213
1172
1496
1340
1494
1358
1207
1207
1232
1385

0.3
*
0.1
*
0.3
*
*
*
*
*
0.1
*
*

0.3
*
0.1
tr
0.3
*
*
0.1
tr
*
0.1
*
tr

0.2
*
0.1
tr
0.2
*
*
0.1
*
*
*
*
tr

0.2
*
0.1
*
0.2
*
*
0.1
*
*
*
*
tr

0.2
*
0.1
*
0.2
*
*
*
*
*
*
*
0.1

0.2
tr
tr
*
0.3
0.1
*
*
*
0.1
tr
tr
*

*
*
0.1
*
0.6
*
*
0.1
*
*
tr
tr
*

2189
2225
2323

1266
1673
1726

0.4
0.5
0.9

0.3
0.5
0.8

0.2
tr
tr

0.1
tr
*

0.1
tr
tr

*
1.5
1.6

*
*
*

99.3

97.8

98.7

98.7

98.3

99.1

97.2

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Constituents

Table 4*continued
Constituents

1021
1021
1109
1119
1145
1157
1179
1199
1208
1228
1241
1245
1267
1278
1432
1458
1473
1492
1539
1555
1586
1588
1589
1595

BP-1

922
930
971
964
1005
978
1009
1021
1021
1024
1048
1035
1012
1078
1072
1053
1130
1183
1082
1107
1388
1420
1214
1162

IA

IIB

68

85

88

89

87

70

71

65

72

67

95

91

96

98

0.5
1.6
2.4
43.1
tr
2.9
1.3
2.0
0.4
0.3
2.9
6.0
0.5
0.7
0.5
0.7
tr
tr
22.1
0.4
0.1
*
0.9
6.1

0.4
1.6
2.2
42.8
3.1
3.4
1.4
2.2
0.7
0.4
2.3
7.8
0.1
1.1
*
0.7
0.3
*
16.7
0.3
0.6
*
0.2
5.2

0.2
0.8
3.0
21.2
tr
1.5
0.4
2.2
0.3
0.1
0.7
1.2
tr
0.2
*
0.3
47.8
*
6.6
0.1
0.1
0.2
*
1.5

1.2
2.6
8.1
1.1
*
15.5
0.3
15.9
0.2
0.1
31.9
2.4
6.6
1.0
tr
0.1
1.2
0.4
2.0
*
0.6
0.1
*
0.2

1.2
2.4
8.0
1.2
*
19.4
0.5
7.7
0.2
0.1
32.4
3.3
1.5
1.2
*
tr
2.8
0.2
3.3
*
0.7
0.1
*
0.2

1.5
4.2
17.1
2.5
*
1.1
0.8
4.7
0.3
0.1
39.0
4.2
14.3
2.1
1.7
*
*
*
1.6
tr
1.5
0.4
*
0.3

1.9
4.0
14.5
2.2
*
1.1
0.9
5.2
0.2
0.1
38.8
3.3
16.0
2.1
2.0
*
*
0.1
4.4
tr
0.8
0.2
*
0.3

1.3
3.3
13.7
1.8
tr
1.0
0.9
4.8
0.3
0.1
37.5
3.4
15.0
2.1
1.6
*
tr
0.1
5.8
tr
2.1
0.5
*
0.6

1.5
3.9
15.4
2.0
*
1.0
1.0
4.5
0.4
0.1
35.8
3.4
16.6
2.1
1.7
*
*
*
5.3
tr
1.9
0.5
*
0.6

2.1
4.7
4.2
0.3
*
1.2
0.3
4.0
tr
0.7
11.7
10.3
3.2
1.9
2.1
tr
0.2
tr
27.6
tr
0.2
*
10.8
0.2

2.1
4.3
4.2
0.4
*
1.1
0.7
4.1
tr
0.3
28.3
7.5
4.9
1.9
0.6
tr
*
*
19.5
tr
*
*
17.3
0.2

1.8
3.5
4.5
0.4
*
1.0
0.8
4.1
tr
0.1
34.3
1.2
6.4
2.2
1.5
0.1
tr
0.1
15.0
tr
*
tr
*
0.3

2.6
5.3
5.3
0.4
*
1.4
1.5
4.6
tr
0.1
67.4
3.4
1.3
2.8
*
*
*
*
0.3
*
tr
0.3
*
0.1

2.2
4.3
5.2
0.5
*
1.1
1.1
4.1
0.1
0.1
53.4
*
4.1
2.2
0.1
0.1
*
tr
11.4
*
*
*
8.0
0.3

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

a-ThujeneH
a-Pinene*
b-Pinene
Sabinene
3-Carene
myrcene
a-Terpinene
Limonene
b-Phellandrene
(Z)-b-Ocimene
c-Terpinene
(E)-b-Ocimene
p-Cymene
Terpinolene
p-Cymenene
trans-Sabinene hydrate
Citronellal
Decanal
Linalool
cis-p-Menth-2-en-1-ol
b-Elemene
(E)-Caryophyllene
Thymyl methyl ether
Terpinen-4-ol

BP-20

*continued
93

94

Constituents

trans-p-Menth-2-en-1-ol
Citronellyl acetate
a-Humulene
Neral
a-Terpineol
a-Bisabolene
Neryl acetate
Bicyclogermacrene
Geranyl acetate
Citronellol
Nerol
Geraniol
Methyl Nmethylanthranilate
Thymol
b-Sinensal
a-Sinensal
Total

BP-20

BP-1

IA

IIB

68

85

88

89

87

70

71

65

72

67

95

91

96

98

1618
1654
1660
1674
1688
1724
1725
1727
1748
1756
1790
1837
2082

1123
1332
1453
1213
1172
1496
1340
1494
1358
1207
1207
1232
1385

0.2
*
*
0.1
0.3
*
0.3
*
0.1
*
tr
tr
0.1

0.2
*
*
*
0.6
*
0.3
*
tr
0.3
0.5
0.1
tr

*
0.1
tr
tr
0.3
*
*
0.1
0.1
7.7
*
0.3
*

*
0.3
0.1
0.4
0.1
*
1.9
*
1.4
0.1
0.3
0.2
tr

tr
0.5
0.1
1.5
0.2
*
2.8
*
3.2
0.2
0.6
0.5
*

tr
*
0.2
*
0.1
0.1
*
*
*
*
*
*
*

tr
*
0.1
*
0.1
0.1
*
*
*
*
*
*
*

tr
*
0.3
0.1
0.3
*
*
*
*
*
tr
tr
*

*
*
0.2
*
0.5
0.1
*
*
*
*
*
*
*

*
*
*
*
0.3
*
*
0.6
*
*
tr
0.1
*

tr
*
*
*
0.4
*
*
*
*
*
tr
tr
*

tr
*
tr
0.1
0.5
0.1
*
*
*
*
0.1
*
*

*
*
tr
*
0.1
0.3
*
*
*
*
tr
*
*

tr
*
*
*
0.6
0.1
*
*
*
*
*
*
*

2189
2225
2323

1266
1673
1726

0.1
1.2
0.5

tr
1.5
*

*
*
0.2

*
*
*

*
*
*

0.1
*
tr

tr
*
*

*
*
*

0.1
*
*

9.5
0.2
0.8

0.4
*
*

20.5
*
*

*
*
1.3

0.1
*
*

98.6

97.6

97.2

97.5

99.3

98.6

99.1

97.6

97.1

97.7

98.4

98.8

98.7

99.6

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Table 4*continued

Table 4*continued

a-ThujeneH
a-PineneH
b-Pinene
Sabinene
3-Carene
Myrcene
a-Terpinene
Limonene
b-Phellandrene
(Z)-b-Ocimene
c-Terpinene
(E)-b-Ocimene
p-Cymene
Terpinolene
p-Cymenene
trans-Sabinene hydrate
Citronellal
Decanal
linalool
cis-p-Menth-2-en-1-ol
b-Elemene
(E)-Caryophyllene
Thymyl methyl ether
Terpinen-4-ol

BP-20

1021
1021
1109
1119
1145
1157
1179
1199
1208
1228
1241
1245
1267
1278
1432
1458
1473
1492
1539
1555
1586
1588
1589
1595

BO-1

922
930
971
964
1005
978
1009
1021
1021
1024
1048
1035
1012
1078
1072
1053
1130
1183
1082
1107
1388
1420
1214
1162

IIB

IIA

III

73

77

64

76

60

61

69

66

99

93

92

90

79

94

2.0
3.7
4.7
0.5
*
1.1
1.4
4.3
tr
0.1
59.7
2.4
1.4
2.4
tr
tr
tr
*
13.7
tr
*
0.2
*
0.3

1.3
2.8
2.9
0.2
*
0.8
0.3
2.5
tr
0.5
15.2
9.4
3.0
1.4
1.2
tr
*
*
37.6
*
tr
0.6
0.3
0.2

1.1
2.7
2.7
0.2
*
0.7
0.3
4.3
tr
0.5
14.5
9.3
3.2
1.1
0.5
tr
*
*
42.9
*
*
0.4
0.4
0.1

1.1
2.5
2.6
0.2
*
0.7
0.3
2.5
tr
0.5
15.7
10.3
3.5
1.1
0.6
*
*
*
42.4
*
*
0.4
0.4
0.1

1.2
2.4
2.7
0.3
*
0.8
0.3
2.6
0.5
0.6
12.0
11.0
2.7
1.2
0.8
tr
*
*
41.4
0.2
*
0.3
0.4
0.1

1.3
2.7
2.9
0.3
*
0.8
0.3
2.8
tr
0.6
13.4
10.1
3.6
1.2
1.4
tr
*
0.1
40.9
*
*
*
0.4
0.1

1.2
2.4
2.6
0.3
*
0.8
0.3
2.1
tr
9.8
10.7
0.5
2.9
1.1
1.2
tr
*
*
47.0
tr
tr
0.3
0.3
0.2

0.3
0.7
1.1
0.3
*
0.4
0.1
4.5
0.1
0.3
3.0
4.1
3.1
0.7
1.5
tr
0.1
*
59.3
*
tr
*
8.8
0.1

tr
0.2
0.1
1.4
*
0.4
tr
11.8
tr
0.7
0.1
1.7
*
tr
*
tr
*
0.1
tr
*
*
0.4
*
0.2

0.7
1.5
1.8
0.2
*
0.5
0.2
4.6
tr
0.2
21.4
0.6
3.4
0.7
0.1
tr
*
tr
0.6
*
*
0.1
0.1
0.1

0.6
1.3
1.3
0.1
*
0.5
0.2
8.9
tr
0.1
16.5
0.5
2.2
0.5
0.1
*
*
tr
0.3
*
*
0.4
*
0.1

2.8
1.6
2.5
0.2
*
0.7
0.2
8.3
0.1
0.2
28.6
0.5
3.9
0.8
0.1
*
*
tr
0.2
*
0.1
0.4
*
0.1

0.9
2.0
2.0
0.9
tr
0.5
0.3
6.3
tr
0.1
22.6
0.5
2.7
0.7
0.1
tr
*
*
0.9
*
*
0.1
0.1
0.2

1.0
2.3
2.2
0.2
*
0.6
0.3
7.9
tr
0.2
24.1
0.5
3.7
0.7
0.1
*
*
tr
0.4
tr
*
0.3
*
0.2

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Constituents

95

Table 4*continued
96

Constituents

BP-20

Total

IA

IIB

68

85

88

89

87

70

71

65

72

67

95

91

96

98

1123
1332
1453
1213
1172
1496
1340
1494
1358
1207
1207
1232
1385

tr
*
tr
*
0.4
*
0.1
*
*
*
tr
*
*

tr
*
0.1
*
0.3
0.2
*
*
tr
*
tr
tr
*

tr
*
*
*
0.1
*
*
0.2
*
*
*
tr
*

*
*
*
*
0.1
0.1
*
*
*
*
*
tr
*

*
*
tr
*
0.3
0.1
*
*
*
*
*
0.2
*

0.1
*
tr
*
0.2
*
*
0.1
*
tr
0.1
tr
0.1

tr
*
tr
*
0.3
0.1
*
*
*
*
tr
tr
*

*
tr
*
*
0.2
*
*
*
*
*
*
tr
*

*
*
tr
*
*
tr
*
*
*
*
*
*
78.7

*
*
*
*
0.2
*
*
*
*
*
*
*
62.3

*
*
*
tr
0.1
tr
*
*
*
*
tr
*
66.1

*
*
tr
0.1
0.1
tr
*
*
*
*
*
*
48.0

*
*
*
*
0.2
*
*
*
*
*
*
*
57.7

*
*
tr
0.1
0.2
tr
*
*
*
*
tr
*
54.5

1266
1673
1726

*
*
0.6

16.2
*
1.4

12.2
*
1.4

12.5
*
1.5

15.3
*
1.8

13.0
*
0.6

14.1
*
0.6

8.4
*
0.5

0.1
2.7
*

0.3
*
*

0.1
*
*

0.3
*
*

0.1
*
*

0.1
*
*

99.1

98.6

98.9

99.2

99.4

97.4

98.9

98.1

98.8

99.6

99.9

99.8

98.2

99.6

!Other compounds ( tr-0.3%): camphene, a-phellandrene, octanal, 6-methylhept-5-en-2-one, nonanal, cis-limonene-1,2-oxide, trans-limonene-1,2-oxide,
(E)-b-farnesene, germacrene-D, (E,E)-a-farnesene, d-cadinene, caryophyllene oxide, (E)-nerolidol, elemol, spathulenol, q-cadinol, a-cadinol, manoyl oxide.
Cultivars : Geleking (74, C. nobilis), Yellow King (75, C. nobilis), King (86, C. nobilis), Rode king (87, C. nobilis), King of Siam (80, C. nobilis), Temple
] Temple (83, C. temple), Cleopatra (82, C. reshni), Peau rugueuse (63, C. deliciosa), Temple (84, C. temple), Emperor (62, C. deliciosa), Empress (59, C. deliciosa),
Swatow (78, C. tangerina), Temple Sue Linda (81, C. temple), Late Emperor (58, C. deliciosa), Redskin (68, C. tangerina), Du Japon (85, C. nobilis), Nasnaran (88,
C. amblycarpa), Kunembo (89, C. nobilis), Yatsushiro (97, C. yatsushiro), Clausellina (70, C. unshiu), URSS (71, C. unshiu), Wase (65, C. unshiu), Owari (72, C.
unshiu), Fuzhu (67, C. erythrosa), Sunki (95, C. sunki), Szibat (91, C. suhuiensis), Szinkom (96, C. suhuiensis), Shekwasha (98, C. depressa), Sihue Gan (73, C.
suhuiensis), Ladu Ordinary (77, C. paratangerina), Brickaville (64, C. tangerina), Peau lisse (76, C. deliciosa), Vohangisahy (60, C. tangerina), Beauty of Glen
Retreat (61, C. tangerina), Ladu (69, C. paratangerina), Dancy (66, C. tangerina), Ougon (99, C. suavissima), Avana Apireno (93, C. deliciosa), de Chios (92, C.
deliciosa), Willow leaf (90, C. deliciosa), Commune (79, C. deliciosa), Tardivo di Ciacculli (94, C. deliciosa).
Order of elution and percentages of components are given on BP-20 column, except compounds with an asterisk (percentages given on BP-1 column). All the
components were identi"ed by GC-RI on polar and apolar columns. All the compounds of samples no. 58, 62, 97 and 93 were also identi"ed by GC/MS. The
major components (bold letters) of samples no. 75, 78, 81, 82, 83, 65, 67, 72, 88, 89, 91, 95, 96, 97, 66, 69, 76, 79, 92, 93 and 94 were identi"ed by 13C NMR.

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

trans-p-Menth-2-en-1-ol
1618
Citronellyl acetate
1654
a-Humulene
1660
Neral
1674
a-Terpineol
1688
a-Bisabolene
1724
Neryl acetate
1725
Bicyclogermacrene
1727
Geranyl acetate
1748
Citronellol
1756
Nerol
1790
Geraniol
1837
Methyl N-methylanthrani-2082
late
Thymol
2189
b-Sinensal
2225
a-Sinensal
2323

BP-1

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

97

Fig. 4. Dendrogram obtained from the cluster analysis of 58 mandarin leaf oils. Samples are clustered using
Ward's technique with an Euclidean distance measure.

Sabinene/linalool chemotype: 17 cultivars (no. 58,59,62,63,68,74,75,78,80}88) belonged to this group. Essential oils were characterized by a high content of sabinene
(21.2}57.3%), associated with linalool (3.1}31.0%) and terpinen-4-ol (1.5}7.9%). This
cluster could be divided into three subgroups. Subgroups IA and IB are di!erentiated
on the basis of sabinene content: 21}43% for seven samples (no. 58,59,62,68,78,81,85,
subgroup IA) and 46}57% for the nine others (no. 63,74,75,80,82-84,86,87, subgroup
IB). For most samples of subgroup IA, linalool exhibited a higher percentage
(17}31%) that those of subgroup IB (3}17%). It is noticeable that the samples no. 74
and 75 exhibited higher content of (E)-b-ocimene (14.0 and 16.0% vs 5.0}7.1%). The
remaining sample (no. 88, subgroup IC) exhibited an atypical composition dominated
by citronellal (47.8%) with an appreciable amount of sabinene (21.2%) and moderate
contents of citronellol (7.7%) and linalool (6.6%).
c-terpinene/linalool chemotype: 19 essential oils (samples no. 60,61,64}67,
69}73,76,77}89,91,95}98) belonged to this chemotype (cluster II). This cluster could be
readily divided into two subgroups on the basis of linalool/c-terpinene ratio. The "rst
subgroup (IIA), to which belonged seven samples (no. 60,61,64,66,69,76,77) exhibited
linalool as major component (37.6}59.3%) with appreciable contents of c-terpinene

98

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Fig. 5. Discriminant analysis scatterplott of 58 mandarin leaf oils.

Fig. 6. Discriminant analysis scatterplot of the leaf oil constituents.

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

99

(3.0}15.7%) and thymol (8.4}16.2%). The second subgroup (IIB), to which belonged
the 12 other samples (no. 65,67,70}73,89,91,95}98) exhibited an appreciable amount of
c-terpinene (28.3}67.4%) except for the oil no. 67 (11.4%). The content of linalool was
not over 27.6%.
We noticed that several components were sometimes important:
f (Z)-b-ocimene (9.8% vs 0.3}0.6%) associated with a low content of (E)-b-ocimene
(0.5% vs 4.1}11.0%) in the sample no. 69 (subgroup IIA);
f b-pinene (13.7}17.1% vs 4.2}8.1%) and p-cymene (14.3}16.6% vs 1.3}6.6%) in the
oils no. 65, 70}72 (subgroup IIB);
f myrcene (15.5}19.4% vs 1.0}1.4%) and limonene (7.7}15.9% vs 4.0}5.2%) associated with a slightly higher content of oxygenated acyclic monoterpenes: citronellal, neral, neryl acetate and geranyl acetate in the oils no. 89 and 97 (subgroup IIB);
f thymol (20.5% vs 0}9.5%) in the sample no. 67 (subgroup IIB);
f thymyl methyl ether (8.0}17.3% vs 0}0.4%) in the sample no. 66 of subgroup IIA
and samples no. 67, 95 and 98 of subgroup IIB.
Methyl N-methylanthranilate chemotype: only six oils (no. 79, 90, 92}94 and 99) were
dominated by methyl N-methylanthranilate (48.0}78.7%) with appreciable amounts
of c-terpinene (0.1}28.6%) and limonene (4.6}11.8%). It is noticeable that the sample
no. 99 (C. suavissima) exhibited a higher percentage of methyl N-methylanthranilate
(78.7%), a very low content of c-terpinene (0.1%) and an appreciable amount of
b-sinensal (2.7%).
In the literature, three di!erent compositions were described: (i) cis- and translinalool oxide (12}60%), associated with linalool (14}36%) for `miyacawaa, `okitsua
(C. unshiu) and `dahongpaoa cultivars (C. tangerina) from China (Lin and Hua, 1992);
(ii) c-terpinene (11}38%) and p-cymene (14}41%) associated with b-pinene (6}14%)
(Kamiyama, 1968; Lawrence, 1995b) or linalool (23%) (Kamiyama, 1967) for samples
from Japan, or associated with b-elemene (11}25%) and linalool (3}15%) (Lawrence,
1995a) for the samples from unspeci"ed cultivars (C. unshiu); (iii) methyl N-methylanthranilate (42}52%) associated with c-terpinene (24}29%) for commercial oils (Dugo
et al., 1996; Mondello et al., 1996a, b, 1997). The c-terpinene/p-cymene/b-pinene
composition described for our samples no. 65, 70}72 from C. unshiu (`wasea, `clausellinaa, `URSSa, and `owaria cultivars) was close to that reported in the literature for
three samples from the same species (Kamiyama, 1968).

4. Discussion
To our knowledge, this is the "rst time that the chemical composition of peel and
leaf oils of nearly 100 cultivars of mandarin from 16 species has been reported.
Peel oils of mandarin belonged to three major chemotypes: limonene, limonene/cterpinene and linalyl acetate/limonene. Fig. 7 shows that the limonene chemotype was
more widespread than the limonene/c-terpinene chemotype (80 varieties from 11
species vs 16 varieties from 6 species). The monovarietal C. yatsushiro exhibited an
atypical composition dominated by linalyl acetate and limonene.

100

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

Fig. 7. Inter- and intraspeci"c di!erentiation on the basis of three chemotypes distinguished for peel
mandarin oils.

For "ve species (C. clementina, C. tangerina, C. unshiu, C. temple and C. paratangerina), all the varieties investigated exhibited the limonene chemotype. Conversely,
a chemical intraspeci"c variability was observed for C. reticulata, C. deliciosa, C.
nobilis and C. suhuensis. Concerning C. reticulata and C. deliciosa (41 and 10 samples,
respectively), the limonene and limonene/c-terpinene compositions appeared at a ratio 5 and 3. C. nobilis is dominated by the limonene chemotype while two samples over
2
1
three, belonging to C. suhuiensis, exhibited the limonene/c-terpinene composition.
Concerning the monovarietal species, the situation appears more variable. The oils
from C. erythrosa and C. reshni exhibited limonene as a major component whereas
the samples from C. amblycarpa and C. sunki on the one hand, the sample from C.
yatsushiro on the other hand showed limonene/c-terpinene and linalyl acetate/
limonene compositions respectively.
It has to be pointed out that three samples, belonging to the limonene/c-terpinene
chemotype, were distinguished by atypical compositions: limonene/b-pinene for the
sample no. 88 from the monovarietal C. amblycarpa, limonene/myrcene for the oil no.
89 from C. nobilis and limonene/p-cymene for the sample no. 96 from C. suhuiensis.
In our sampling, domination of the oils by limonene was common. This has been
reported before for the oils of `dancya, `malvasioa, `ortaniquea, `ellendalea and
`cravoa cultivars from C. reticulata (Ashoor and Bernhard, 1967; Moshonas and

M.-L. Lota et al. / Biochemical Systematics and Ecology 29 (2001) 77}104

101

Fig. 8. Inter- and intraspeci"c di!erentiation on the basis of three chemotypes distinguished for leaf
mandarin oils.

Shaw, 1974; Koketsu et al., 1983; Dellacassa et al., 1992; Calvarano et al., 1989),
`communea, `nulesa, `orovala, `monreala cultivars from C. clementina (Verzera et al.,
1997, 1998), `praecoxa cultivar from C. unshiu (Lawrenc