Industrial Crops and Products 45 (2013) 7–19

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Industrial Crops and Products

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Exploring compositional diversity in the essential oils of 34 Ocimum taxa from

Indian flora
Ram S. Verma a,∗ , Rajendra C. Padalia a , Amit Chauhan a , Sanjog T. Thul b,1
a
CSIR-Central Institute of Medicinal and Aromatic Plants, Research Centre, Pantnagar, P.O. Dairy Farm Nagla, Udham Singh Nagar, Uttarakhand 263149, India
b
CSIR-Central Institute of Medicinal and Aromatic Plants, P.O. CIMAP, Lucknow 226015, India

a r t i c l e i n f o a b s t r a c t

Article history: To explore diversity in the morphological features, essential oil yield, and chemical composition of Oci-
Received 24 October 2012 mum taxa growing in foot and mid-hills of northern India; 34 populations representing five Ocimum
Received in revised form 2 December 2012 species were collected and investigated. Essential oil yield (%v/w) was found to vary from 0.10 to 0.74% in
Accepted 3 December 2012
different populations. Analyses of the essential oils by GC/FID and GC/MS and the subsequent classification
by statistical analysis resulted in seven clusters with significant variations in their terpenoid compositions
Keywords:
(cluster I: methyl chavicol/linalool; cluster II: eugenol/(Z)-␤-ocimene/␤-elemene/(E)-caryophyllene;
Ocimum spp
cluster III: camphor/limonene; cluster IV: eugenol/(E)-caryophyllene/␤-bisabolene/␤-elemene/methyl
Diversity
Morphology
chavicol/1,8-cineole; cluster V: linalool; cluster VI: methyl eugenol/(E)-caryophyllene/␤-elemene; and
Essential oil yield cluster VII: citral/nerol/geraniol type). Comparison of present results with earlier reports revealed the
Composition existence of four new chemotypes in Ocimum taxa from this region.
New chemotypes © 2012 Elsevier B.V. All rights reserved.

1. Introduction shown antioxidant and antimicrobial activities due to its phenolic

Ocimum spp (Lamiaceae), commonly known as ‘Basil’, are annual
or perennial, highly aromatic, branched herb or shrub native to the
tropical and subtropical regions of Asia, Africa, and Central South
America (Labra et al., 2004). The genus Ocimum comprises more
than 150 species and is considered as one of the largest genera
of family Lamiaceae (Pushpangadan and Bradu, 1995). However,
according to another classification, Ocimum had only 65 species
and other attributions should be considered as synonyms (Paton
et al., 1999). Ocimum tenuiflorum L. (syn. Ocimum sanctum L.), Oci-
mum gratissimum L., Ocimum americanum L. (syn. Ocimum canum
Sims), Ocimum basilicum L., Ocimum kilimandscharicum Gurke, and
Ocimum micranthum Willd. are examples of important species of
the genus Ocimum, which are known to have several medicinal
properties (Prakash and Gupta, 2005). In Ayurveda, O. tenuiflorum
(Tulsi or holy basil) has been well documented for its therapeu-
tic potentials and described as antiasthmatic and antikaphic drugs
(Sirkar, 1989). The oils of basil have been used principally in the
food and cosmetic industries (Vina and Murillo, 2003). Basil has
∗ Corresponding author. Tel.: +91 5944 234445; fax: +91 5944 234712.
E-mail addresses: rs.verma@cimap.res.in, rswaroop1979@yahoo.com
(R.S. Verma).
1
Present address: CSIR-National Environmental Engineering Research Institute,
Nehru Marg, Nagpur 440020, India.
and aromatic compounds (Lee and Scagel, 2009).
The genus Ocimum is characterized by a great chemical and
morphological variability. The ease of crosspollination leads to a
large number of subspecies, varieties, and forms, which differ in
essential oil composition and morphological characters (Krishnan,
1981; Simon et al., 1990). Previous studies show that there are
large numbers of species and varieties fall in this genus (Labra
et al., 2004; Mondello et al., 2002; Padalia and Verma, 2011;
Vieira et al., 2001; Vina and Murillo, 2003). In a study of essen-
tial oils of different geographical origins, Lawrence (1988) found
that the main constituents of the essential oils of basil are pro-
duced by two different biochemical pathways, viz. shikimic acid
pathway (phenylpropanoids) and mevalonic acid pathway (ter-
penes). Apart from chemotypic and genetic factors, other factors
such as harvesting season, plant phenological stages, and plant
part also affect chemical composition of basil oils (Verma et al.,
2012).
Among the various species of basil, O. basilicum is considered
most important for its sweet aromatic oil and cultivated in differ-
ent regions. On the basis of essential oil composition of O. basilicum,
Lawrence (1988) established four chemotypes (methyl chavicol,
linalool, methyl eugenol, and methyl cinnamate) and numerous
subtypes. In addition, other species of the genus have also been
explored for essential oil composition from different parts of the
world (Gupta, 1996; Gupta and Tava, 1997; Jirovetz et al., 2003;
Koba et al., 2009; Kothari et al., 2005; Padalia and Verma, 2011;
Runyoro et al., 2010).

0926-6690/$ – see front matter © 2012 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.indcrop.2012.12.005
8 R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19
However, to the best of our knowledge no systematic chemical
investigations have been carried out on essential oil composition
of Ocimum spp growing wild or for the medico-religious purposes
in foot and mid-hill regions of north India. Therefore, aim of the
present study was to investigate the chemical diversity among the
Ocimum spp growing in north India (foot and mid-hills). This study
will contribute to the knowledge of chemical diversity of an Oci-
mum that could improve the use of Indian Ocimum spp.
2. Materials and methods
2.1. Plant materials
A total of thirty-four populations belong to five Ocimum spp,
viz. O. basilicum L., O. tenuiflorum L., O. kilimandscharicum Gurke,
O. gratissimum L., and O. americanum L. were collected by three
authors (R.S. Verma, R.C. Padalia, and Amit Chauhan) in the month
of August–September (2009–2012), at flowering stage, from foot
and mid-hill regions of north India. Seeds of one population of O.
basilicum (O13) were obtained from Andhra Pradesh and grown
under foot hill conditions. Botanical authentication of the plant
materials was carried out at taxonomy department of CIMAP
Research Centre Pantnagar by one of the author (Dr. Amit Chauhan).
The voucher specimens of the investigated populations were pre-
pared and maintained at taxonomy department, CIMAP Research
Centre Pantnagar. Origin/locality, essential oil yield, and abbre-
viation of the investigated Ocimum populations are described in
Table 1. The representative pictures of the collected Ocimum spp
are shown in Fig. 1.
2.2. Isolation of essential oils
The fresh plant materials (aerial parts) of different Ocimum spp
were hydrodistilled in a Clevenger’s type apparatus for 3 h to isolate
essential oils. The essential oil content (%v/w) was estimated on
a fresh weight basis. The oil samples obtained were dehydrated
over anhydrous Na2 SO4 and kept in a cool and dark place before
analyses.
2.3. Gas chromatography (GC/FID)
GC analysis of the essential oil samples was carried out on a
Nucon gas chromatograph model 5765 equipped with flame ioniza-
tion detector (FID) and DB-5 (coated with 5% phenyl polysiloxane,
30 m length × 0.32 mm internal diameter; 0.25 ␮m film coating)
fused silica capillary column. Hydrogen was used as a carrier gas at
1.0 ml/min. Temperature programming was done from 60 to 230 ◦ C
at 3 ◦ C/min with final hold time of 10 min. Injector and detector
temperatures were 220 ◦ C and 230 ◦ C, respectively. Injection size
was 0.02 ␮L neat (syringe: Hamilton 1.0 ␮L capacity, Alltech USA)
and split ratio was 1:40.
2.4. Gas chromatography–mass spectrometry (GC/MS)
GC/MS analysis of the essential oil samples was carried out
on a PerkinElmer AutoSystem XL GC interfaced with a Turbomass
Quadrupole mass spectrometer fitted with a DB-5 fused-silica cap-
illary column (60 m × 0.32 mm i.d., film thickness 0.25 ␮m). The
oven temperature programme was from 70 to 250 ◦ C, at 3 ◦ C/min,
with initial and final hold time of 2 min; injector, transfer line and
source temperatures were 250 ◦ C; injection size 0.03 ␮L neat; split
ratio 1:30; carrier gas He at 1.0 ml/min; ionization energy 70 eV;
mass scan range 40–450 amu.
2.5. Identification of essential oil constituents
Identification of the essential oil constituents was done on the
basis of retention time (tR ), retention index (RI), calculated using
a homologous series of n-alkanes (C8 –C30 , Supelco Bellefonte, PA,
USA) under identical experimental conditions, co-injection with
standards or known essential oil constituents, mass spectra library
search (NIST/EPA/NIH version 2.1 and Wiley registry of mass spec-
tral data 7th edition), and by comparing the mass spectral and
retention data with literature (Adams, 2007). The relative amounts
of individual components were calculated based on the GC peak
area (FID response) without using a correction factor.
2.6. Statistical analysis
To examine phytochemical diversity based on the content (%)
of chemical constituents in essential oil among the studied 34
populations; these were subjected for statistical analysis based on
Euclidean Distance Scaling model using SPSS statistics 17.0 soft-
ware (SPSS, Inc.). In this analysis, Euclidean distance model was
used which generally used to compute raw data by employing
ALSCAL algorithm. This algorithm optimizes the total variables that
measures the distances between similar values and fits the vari-
ables accordingly than to the dissimilarities. The value measured
depicts the proportion of variance of the scaled data and get par-
titioned which is accounted for by their corresponding distances.
These distances are used to produce a joint configuration of ele-
ments and mapped in the form of a plot. In this case, the derived
multidimensional scaling plot depicts the grouping of individual
populations as per their chemical constituents. The derived plot
was further used for classifying the populations as chemotypes
based on their major chemical components.
3. Results and discussion
The morphological characteristics and essential oil yields
observed in 34 populations of five Ocimum taxa (15 populations
of O. basilicum; 07 of O. tenuiflorum; 09 of O. kilimandscharicum;
02 of O. gratissimum, and 01 of O. americanum) are summerized in
Table 1. Essential oil yield was found to vary from 0.20 to 0.70% in
different populations of O. basilicum (O1–O15). Maximum oil yield
was found in population O8 (0.70%), followed by O6 (0.65%), O15
(0.65%), and O14 (0.56%). The oil yield varied from 0.10 to 0.53%
in O. tenuiflorum populations (O16–O22) with the highest in pop-
ulation O18. Further, the oil yield was found to vary from 0.14 to
0.74% in different populations of O. kilimandscharicum (O23–O31).
The promising populations of O. kilimandscharicum group were O24
(0.74%), O23 (0.68%), and O26 (0.67%). On the other hand, essential
oil yield was found to be 0.40% and 0.63% in populations O32 and
O33, respectively of O. gratissimum. The only collected population,
O34, of O. americanum was found to possess 0.30% essential oil.
The results from essential oil yield clearly indicated the existence
of huge variation between and within the species. These variations
may be linked to numerous intrinsic as well as extrinsic factors.
Essential oils isolated from different Ocimum populations were
analyses by GC/FID and GC/MS. Altogether, ninety-six constituents,
representing 84.3–98.6% of total oil compositions were identified
in different populations of ‘basilicum group’ (O1–O15). In ‘tenuiflo-
rum group’ (O16–O22), a total of sixty-seven constituents, forming
88.2–98.0% of total oil compositions were characterized. Further,
in ‘kilimandscharicum group’ (O23–O31) a total of eighty-two con-
stituents, representing 83.1–97.8% of the total oil compositions
were identified. Moreover, in ‘gratissimum group’ (O32, O33) a
total of forty-eight constituents, forming 93.6–95.1% of the total
oil compositions were identified. However, in ‘americanum group’,
Table 1
Origin and essential oil yield of 34 investigated Ocimum spp populations.

Ocimum taxa Character Voucher no. Code Origin/locality EO (%v/w)

O. basilicum – CIMPANT-221 O1 Dehradun, Uttarakhand 0.38

O. basilicum Annual, up to 50 cm tall; inflorescence purple; stems ciliate; corolla and calyx ciliate CIMPANT-222 O2 Forest nursery, Purara, 0.52
Bageshwar, UK
O. basilicum Perennial; leaves glabrous, leathery, large, entire, lanceolate, surface wrinkled; inflorescence green, flowers CIMPANT-242 O3 Haldwani, Nainital, UK 0.45
white, bracts large
O. basilicum Annual, up to 50 cm tall; inflorescence purple, corolla and calyx ciliate CIMPANT-244 O4 Vajol, Almora, UK 0.52
O. basilicum Annual, up to 50 cm tall; inflorescence purple, corolla and calyx ciliate CIMPANT-245 O5 Bagwalipokhar, Almora, UK 0.41
O. basilicum Perennial, up to 1.0 m tall; inflorescence purple, corolla and calyx ciliate CIMPANT-246 O6 Kamedi, Bageshwar, UK 0.65
O. basilicum Perennial, up to 1.0 m tall; inflorescence green, flowers white, corolla and calyx ciliate CIMPANT-247 O7 Khatima, U.S. Nagar, UK 0.45
O. basilicum Perennial, up to 1.0 m tall; inflorescence purple, corolla and calyx ciliate CIMPANT-248 O8 Aashon, Bageshwar, UK 0.70
O. basilicum Annual, up to 50 cm tall; leaves small, lemon scented; inflorescence green, flowers white, corolla and calyx CIMPANT-249 O9 Chatti-Mankote, 0.45

R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19

ciliate Bageshwar, UK
O. basilicum Annual, up to 50 cm tall; leaves small, lemon scented; inflorescence green, flowers pink, corolla and calyx CIMPANT-243 O10 Banvasa, Champawat, UK 0.35
ciliate
O. basilicum Perennial, stem greenish-purple, glabrous; leaves glabrous, somewhat wrinkled; inflorescence purple CIMPANT-317 O11 Sitarganj, U.S. Nagar, UK 0.20
O. basilicum Perennial, up to 50 cm tall; stem purple; leaves ovate, green; inflorescence purple CIMPANT-318 O12 Pantnagar, U.S. Nagar, UK 0.30
O. basilicum Perennial, up to 50 cm tall; stem purple; leaves lanceolate, purplish on dorsal side; inflorescence purple CIMPANT-347 O13 Tirupati Balaji, AP 0.32
O. basilicum Perennial, up to 50 cm tall; stem purple; leaves ovate, light green; inflorescence purple CIMPANT-348 O14 Akhtyarpur, 0.56
Lakhimpur-Kheri, UP
O. basilicum Perennial, up to 1.0 m tall; inflorescence purple, corolla and calyx ciliate CIMPANT-250 O15 Roonikhet, Bageshwar, UK 0.65
O. tenuiflorum ‘CIM-Ayu’ Perennial, leaves light green, chartaceous, minute hairs above, hirsute below and on nerves, oblong to CIMPANT-223 O16 Purara, Bageshwar, UK 0.36
oblanceolate, serrate, subacute-acuminate; inflorescence green
O. tenuiflorum ‘CIM-Angna’ Morphologically same as O16 except being dark purple pigmented on stem, leaves and inflorescence CIMPANT-224 O17 Purara, Bageshwar, UK 0.15
O. tenuiflorum ‘CIM-Kanchan’ Morphologically similar to other O. tenuiflorum; pigmentation in between O16 and O17 CIMPANT-225 O18 Purara, Bageshwar, UK 0.53
O. tenuiflorum Perennial; leaves light green, smaller as compared to O16, chartaceous, minute hairs above, hirsute below CIMPANT-226 O19 Teetbazar, Bageshwar, UK 0.18
and on nerves, oblong to oblanceolate, serrate, subacute-acuminate; inflorescence green
O. tenuiflorum Perennial; leaves light green, smaller as compared to O16, chartaceous, minute hairs above, hirsute below CIMPANT-319 O20 Nagla, US Nagar, UK 0.20
and on nerves, oblong to oblanceolate, serrate, subacute-acuminate; inflorescence green
O. tenuiflorum Perennial; leaves purple, smaller as compared to O16, chartaceous, minute hairs above, hirsute below and CIMPANT-257 O21 Katerniaghat WLS, 0.10
on nerves, oblong to oblanceolate, serrate, subacute-acuminate; inflorescence green Baharaich, UP
O. tenuiflorum Perennial; leaves light green, smaller as compared to O16, chartaceous, minute hairs above, hirsute below CIMPANT-320 O22 CRC, Pantnagar, UK 0.17
and on nerves, oblong to oblanceolate, serrate, subacute-acuminate; inflorescence green
O. kilimandscharicum Perennial; up to 1 m tall, oil glands dense, leaves ciliate between veins dorsally, margin veins shallow CIMPANT-227 O23 Vajulla, Bageshwar, UK 0.68
O. kilimandscharicum Perennial; up to 1.5 m tall; bushy; leaves green laceolate; inflorescence green CIMPANT-228 O24 Takula, Almora, UK 0.74
O. kilimandscharicum Plants ca. 50–60 m tall; leaves ovate, veins very prominent, glabrous between veins CIMPANT-251 O25 Pilkholi, Almora, UK 0.35
O. kilimandscharicum Perennial; up to 1.0 m tall, leaves ciliate between veins dorsally, margin veins shallow, glands dense CIMPANT-252 O26 Berinag, Pithoragarh, UK 0.67
O. kilimandscharicum Plants ca. 50–60 m tall; leaves ovate, veins very prominent, leaves glabrous between veins dorsally CIMPANT-253 O27 Bahuli, Bageshwar, UK 0.25
O. kilimandscharicum Plants ca. 50–60 m tall; leaves ovate, veins very prominent, leaves glabrous between veins dorsally CIMPANT-229 O28 Kausani, Almora, UK 0.23
O. kilimandscharicum Plants ca. 50–60 m tall; leaves ovate, veins very prominent, leaves glabrous between veins dorsally CIMPANT-254 O29 Pilkholi, Almora, UK 0.32
O. kilimandscharicum Plants ca. 50–60 m tall; leaves ovate, veins very prominent, leaves glabrous between veins dorsally CIMPANT-255 O30 Niglat, Almora, UK 0.14
O. kilimandscharicum Perennial; up to 1 mt tall, leaves ciliate between veins dorsally, margin veins shallow, glands dense CIMPANT-321 O31 CRC, Pantnagar, UK 0.47
O. gratissimum Perennial; woody shrubs, throat of fruiting calyx closed by the upcurved 2 median teeth of anterior lip CIMPANT-258 O32 Aampadav, Nainital, UK 0.40
O. gratissimum Perennial; woody shrubs, throat of fruiting calyx closed by the upcurved 2 median teeth of anterior lip CIMPANT-322 O33 Santipuri, US Nagar, UK 0.63
O. americanum Annual; dwarf, 20–30 cm tall; reproductive parts much smaller than O. basilicum plants, flowers white, calyx CIMPANT-256 O34 Ramapur, 0.30
and corolla green, ciliate Lakhimpur-Kheri, UP

EO, essential oil; UK, Uttarakhand; AP, Andhra Pradesh; UP, Uttar Pradesh.

9
10 R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19

represented by solitary population (O34), altogether, sixty-three
constituents, representing 99.1% of the total oil composition were
characterized. The detailed results pertaining to essential oil com-
position are summerized in Table 2 (populations O1–O17) and
Table 3 (populations O18–O34).
Considerable variations were observed at chemical level among
the investigated populations of ‘basilicum group’ (O1–O15), which
revealed the existence of eight distinct chemotypes/sub-groups.
Populations O3, O11, and O13 were dominated by methyl chavicol
(>83.0%; 83.9–88.2%). Populations O1, O4, and O5 were char-
acterized by presence of higher amounts of methyl chavicol
(64.5–73.8%), and linalool (16.8–27.1%). The populations O6,
O8, and O15 were rich in methyl chavicol (46.5–62.1%), methyl
eugenol (0.8–14.9%), camphor (8.2–9.1%), and linalool (5.4–8.6%).

Fig. 1. Inter- and intraspecific variation in genus Ocimum from India: A–G (O. basilicum), H–I (O. tenuiflorum), J–M (O. kilimandscharicum), N (O. gratissimum), and O (O.
americanum).
 R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19 11

Fig. 1. (Continued)

Further, the population O2 was dominated by linalool (74.3%) with
lesser amounts of eugenol (4.7%), and ␤-selinene (4.4%). However,
the population O7 was found to be rich in ␤-bisabolene (25.6%),
1,8-cineole (17.5%), ␣-cadinene (11.9%), methyl chavicol (7.6%),
and eugenol (5.9%). Moreover, the population O9 was dominated
by citral (64.5%; neral 27.6% + geranial 36.9%), nerol (9.7%), and
geraniol (6.8%). Characteristic volatile constituents of population
O10 were linalool (35.5%), citral (24.9%), (E)-␣-bergamotene (5.9%)
and (E)-caryophyllene (4.9%); whereas populations O12 and O14
were dominated by linalool (36.3–45.0%), (E)-methyl cinnamate
(33.3–38.7%), (Z)-methyl cinnamate (5.8–6.1%), and ␣-cadinol
(2.5–3.4%).
A total of seven investigated populations of ‘tenuiflorum group’
(O16–O22) could be classified in to three chemotypes. Populations
O16, O17, O19, O20, and O22 were characterized by the presence
of higher amounts of eugenol (54.9–77.3%), ␤-elemene (4.1–15.0%),
12 R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19
and (E)-caryophyllene (3.0–8.4%). Further, the population O18 was
dominated by methyl eugenol (79.7%), (E)-caryophyllene (5.0%),
and ␤-elemene (4.5%). However, the characteristic constituents
of population O21 were (E)-caryophyllene (36.2%), ␤-elemene
(23.4%), and eugenol (18.9%).
Populations belonging to ‘kilimandscharicum group’ (O23–O31)
could be arranged as three distinct chemotypes. Among these,
O23 was rich in linalool (84.1%), camphor (6.0%), and (E)-
caryophyllene (2.0%). However, the populations O24–O26 and O31
were dominated by camphor (43.5–64.9%), limonene (8.7–29.8%),
and camphene (0.0–6.4%). Further, the populations O27–O30
were characterized by higher amounts of eugenol (4.5–52.4%),
methyl chavicol (7.7–23.3%), ␤-bisabolene (4.5–22.9%), 1,8-cineole
(14.4–20.9%), and (E)-␣-bisabolene (3.0–10.9%).
Furthermore, the ‘gratissimum group’ (O32, O33) repre-
sented by only one chemotype, which was characterized by the
presence eugenol (66.3–66.4%), (Z)-␤-ocimene (11.0–15.5%), ger-
macrene D (4.8–5.3%), and (E)-caryophyllene (1.6–2.3%) as main
constituents.
On the other hand, essential oil of population O34, belongs to
‘americanum group’ was dominated by camphor (45.6%), longip-
inanol (7.4%), limonene (6.3%), ␣-pinene (4.9%), camphene (4.6%),
␣-guaiene (4.2%), ␤-selinene (4.0%), ␣-selinene (3.0%), and (E)-
caryophyllene (2.9%).
Above chemotypic differentiation of the Ocimum popula-
tions was also validated statitistically. To evaluate whether the
identified essential oil constituents may be useful in reflecting
similarity and differences within the investigated populations,
thirty-three components (amount detected ≥3.0% in any popu-
lation) of Tables 2 and 3 were subjected for statistical analysis
based on Euclidean distance scaling model. Further, the derived
distance matrix yielded multidimensional scaling (MDS) plot
depicting seven clusters of thirty-four populations for thirty-three
compounds, with individual compounds expressed as percentage
of the total fraction, as shown in Fig. 2. Based on MDS analysis
the thirty-four populations could be divided into seven clusters,
first with nine populations (I: O1, O3, O4, O5, O6, O8, O11, O13,
O15); second with 7 populations (II: O16, O17, O19, O20, O22,
O32, O33); third with five populations (III: O24, O25, O26, O31,
O34); fourth with six populations (IV: O7, O21, O27, O28, O29,
O30); fifth with five populations (V: O2, O10, O12, O14, O23); and
last two with solitary population (VI: O18; VII: O9). Further, the
populations of cluster ‘I’ were characterized by the presence of
higher amount of methyl chavicol (46.5–88.2%), followed by small
quantity of linalool (1.0–27.1%), camphor (0.0–9.1%), and methyl
eugenol (0.0–14.9%); cluster ‘II’ was characterized by higher
percentages of eugenol (54.9–77.3%), ␤-elemene (0.2–15.0%), (Z)-
␤-ocimene (0.0–15.5%), (E)-caryophyllene (1.6–8.4%), germacrene
D (0.2–5.3%), and methyl eugenol (0.1–4.7%); cluster ‘III’ was found
to be dominated by camphor (43.5–64.9%), limonene (6.3–29.8%),
and camphene (0.0–6.4%); cluster ‘IV’ was characterized by the
presence of multiple major components, viz. eugenol (4.5–52.4%),
(E)-caryophyllene (<0.05–36.2%), ␤-bisabolene (0.0–25.6%),
␤-elemene (<0.05–23.4%), methyl chavicol (0.0–23.3%), 1,8-
cineole (0.05–20.9%), ␣-cadinene (0.0–11.9%), (E)-␣-bisabolene
(0.0–10.9%), and (E)-␤-ocimene (0.0–3.5%). Further, the cluster ‘V’
was found to be rich primarily in linalool (35.5–84.1%); however,
next major constituents were different in different populations
of this cluster. For instance, eugenol (4.7%) and ␤-selinene (4.4%)
were other major constituents in population O2; citral (24.9%),
(E)-␣-bergamotene (5.9%), and (E)-caryophyllene (4.9%) were
other major constituents of population O10; (E)-methyl cinna-
mate (33.3–38.7%), and (Z)-methyl cinnamate (5.8–6.1%) were
other major constituents in populations O12 and O14; however,
camphor (6.0%), and (E)-caryophyllene (2.0%) were noticed to be
other major constituents in population O23. Further, the cluster
‘VI’ was characterized by presence of higher amounts of methyl
eugenol (79.7%), (E)-caryophyllene (5.0%), ␤-elemene (4.5%), and
eugenol (1.5%); however, the cluster ‘VII’ denoted as citral type
containing citral (64.5%), nerol (9.7%), and geraniol (6.8%) as
principal constituents.
The existence of chemical diversity within the species and
similarity with the populations of other species could easily be
seen from the distribution pattern of the Ocimum populations in
different clusters (Fig. 2). The fifteen populations of ‘basilicum
group’ (O1–O15) were distributed in four clusters (I: 09 popu-
lations; IV: 01; V: 04 and VII: 01). However, seven populations
of ‘tenuiflorum group’ (O16–O22) were scattered in three clus-
ters (II: 05 populations; IV: 01 and VI: 01) due to their diverse
chemical characters. Further, nine populations of ‘kilimandschar-
icum group’ (O23–O31) were found to be divided in three clusters
(III: 04 populations; IV: 04 and V: 01), whereas, two populations
of ‘gratissimum group’ (O32, O33) were found placed in cluster
‘II’ only due to similar chemical characters. Finally, the ‘ameri-
canum group’ found place in cluster ‘III’ owing to its similarity
with other camphor rich populations. These results clearly showed
that the certain taxonomically different species were grouped in
same cluster because of the presence of similar major chemi-
cal constituents. However, they could be differentiated on the
basis of second/third major constituents present in their essen-
tial oils. For example, eugenol rich populations of O. tenuiflorum
(O16, O17, O19, O20, and O22) were clustered together with the O.
gratissimum populations (O32, O33). However, these two species
could be distinguished by the second/third major constituents of
their essential oils (␤-elemene/(E)-caryophyllene in O. tenuiflorum
and (Z)-␤-ocimene/germacrene D in O. gratissimum). Likewise, the
linalool chemotypes of O. basilicum (O2) and O. kilimandscharicum
(O23) could easily be distinguished by their next major constituents
of the respective oils (eugenol in O. basilicum and camphor in O.
kilimandscharicum).
Compositional analysis of the essential oils of Ocimum taxa has
revealed a comprehensive diversity in the oil components, and the
different chemovarieties have been reported from various regions
of the world. The chemotypes described for O. basilicum are mainly
methyl chavicol (Koba et al., 2009; Verma et al., 2012), methyl
chavicol/linalool (Alves et al., 2007; Verma et al., 2012), methyl
cinnamate (Gupta, 1996; Jirovetz et al., 2003; Vina and Murillo,
2003), linalool (Alves et al., 2007; Dambolena et al., 2010), trans-␣-
bergamotene/linalool (Yavari et al., 2011), menthone/methyl chav-
icol (Hassanpouraghdam et al., 2010), 1,8-cineole (Runyoro et al.,
2010), (E)-myroxide/caryophyllene oxide (Runyoro et al., 2010),
camphor/linalool (Dambolena et al., 2010), citral (Dambolena et al.,
2010; Telci et al., 2006), methyl eugenol (Koba et al., 2009), (E)-
anethol (Koba et al., 2009), methyl eugenol/(E)-anethol (Koba et al.,
2009), geraniol/linalool (Alves et al., 2007), and eugenol (Gupta,
1996) types. In present study, methyl chavicol (O3, O11, O13);
methyl chavicol/linalool (O1, O4, O5); methyl chavicol/methyl
eugenol/camphor/linalool (O6, O8, O15); linalool (O2); ␤-
bisabolene/1,8-cineole/␣-cadinene/methyl chavicol/eugenol (O7);
citral (O9); linalool/citral (O10); linalool/methyl cinnamate (O12,
O14) rich populations have been identified for O. basilicum.
Out of these eight chemovariants, two chemovariants, viz. ␤-
bisabolene/1,8-cineole/␣-cadinene/methyl chavicol/eugenol type
(O7); and linalool/citral type (O10) were described for the first time
for O. basilicum from this region. Further, the chemovariant, methyl
chavicol/methyl eugenol/camphor/linalool (O6, O8, O15) was also
somewhat different from earlier reported methyl chavicol chemo-
types (methyl chavicol >80.0%; and methyl chavicol/linalool) due
to presence of methyl eugenol and camphor as the second and third
major constituents, respectively in their essential oil.
The taxa belonging to O. tenuiflorum have been explored for
volatile oils from different places. Chemotypes reported till date
Table 2
Compositional variation in the essential oils of Ocimum populations (O1–O17).

S. no. Compounda RIb RIc Content (%)d

O1 O2 O3 O4 O5 O6 O7 O8 O9 O10 O11 O12 O13 O14 O15 O16 O17

1 2-Methyl butanoic acid 842 841 – – – – – – 0.1 – – – – – – – – – –

2 Ethyl isovalerate 845 849 – – – – – – 0.7 – – – 0.1 – 0.2 – – – –
3 (Z)-3-Hexenol 856 859 – – – – – – t – – t – 0.1 – 0.1 – – –
4 n-Hexanol 860 863 – – – – – – t – – – – – – – – – –
5 Isopentyl acetate 874 876 – – – – – – t – – – – – – – – – –
6 ␣-Thujene 928 924 t t – – – – t – t t – – – – t t –
7 ␣-Pinene 935 932 0.1 t t t – 0.2 0.5 0.2 0.3 0.2 – – 0.1 0.1 0.2 0.5 0.2
8 Camphene 942 946 0.1 – – t – 0.4 t – – t t – – 0.1 – 0.2 0.3
9 Sabinene 967 969 t – – – – – 0.1 – 0.2 – 0.1 t t 0.1 – – –
10 1-Octen-3-ol 975 974 – – – – – – – – – – – 0.2 – 0.4 – – –
11 ␤-Pinene 975 974 t t t t – 0.2 1.4 – 0.4 t – – 0.2 t – 0.1 0.2
12 3-Octoanone 980 979 – – – – – – t – – – – – t – – – –
13 6-Methyl-5-hepten-2-one 983 981 – – – – – – – – t 0.5 – – – – – – –
14 Myrcene 988 988 t 1.1 0.4 0.1 0.1 0.2 0.2 0.3 t 0.1 0.3 0.3 0.7 0.4 0.3 t t
15 3-Octanol 989 988 – – – – – – t – – – – – – – – – –

R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19

16 ␦-2-Carene 1000 1001 – – – – – – – – 0.1 – – – – – – – –
16 ␣-Phellandrene 1002 1002 – – – – – t 0.1 t 0.2 – t t t – t t t
18 (Z)-3-Hexenyl acetate 1005 1004 – t – – – – – – – 0.2 – – – – – – –
19 ␣-Terpinene 1015 1014 t t – – – – t – – 0.1 – – – – – – t
20 p-Cymene 1023 1020 – 0.2 – – – 0.1 0.2 – t 0.1 t t – – – 0.1 0.1
21 Limonene 1025 1024 t 0.8 t – t – 0.4 – t 0.3 0.1 t 0.2 0.2 t 0.2 0.2
22 1,8-Cineole 1028 1026 t t 0.9 t 0.2 1.2 17.5 0.6 – 0.1 0.4 0.6 1.1 2.1 0.7 – –
23 (Z)-␤-Ocimene 1030 1032 0.3 – – – – – 0.1 – – 0.8 0.1 t 0.1 0.1 – – –
24 (E)-␤-Ocimene 1043 1044 – 0.9 1.0 0.2 0.2 0.1 3.5 2.0 0.4 t 1.5 0.9 3.3 2.9 2.4 0.4 0.2
25 ␥-Terpinene 1058 1054 t t t – – t 0.1 1.7 0.1 0.6 t – – – 1.9 t t
26 cis-Sabinene hydrate 1062 1065 – t – – – – 0.2 – – 1.2 – t t 0.1 t – –
27 cis-Linalool oxide (furanoid) 1072 1067 – t – – – – – t t t – t – t – – –
28 Fenchone 1079 1083 – – – – – – – – – – – – – – – t –
29 trans-Linalool oxide (furanoid) 1080 1084 – t – – – – – – 0.1 0.2 0.1 t – – – – –
30 Terpinolene 1084 1086 – t – – – – t – – – – – 0.5 0.3 – – –
31 Linalool 1099 1095 27.1 74.3 1.3 16.8 24.7 5.4 0.9 8.6 0.1 35.5 2.0 45.0 1.0 36.3 5.5 0.4 1.7
32 trans-Sabinene hydrate 1102 1098 – – – – – – – – t t – – – – – – –
33 1-Octen-3-yl acetate 1111 1110 – t – – – – – – – 0.3 t – – – – – –
34 Camphor 1144 1141 t 0.2 t – – 8.2 0.1 9.1 0.5 2.0 – t 0.3 2.3 9.1 – –
35 Citronellal 1148 1148 – – – – – – – – 0.4 – – – – – – – –
36 ␦-Terpineol 1161 1162 – – – – – – 0.3 – – – – – – – – – –
37 Borneol 1163 1165 0.1 1.4 t – – 0.1 – – – 0.2 – 0.2 0.1 0.5 – t t
38 Terpinen-4-ol 1177 1174 0.2 t – t – t 0.3 0.2 1.5 4.8 t 0.1 t t 0.1 t –
39 ␣-Terpineol 1186 1186 0.7 3.3 – – – – 1.0 0.1 0.1 0.2 – 0.1 t 0.5 0.1 0.3 0.4
40 Methyl chavicol 1200 1195 64.5 – 88.2 73.8 67.1 46.5 7.6 54.7 – t 85.2 0.3 83.9 0.8 62.1 – –
41 Nerol 1227 1227 0.2 t – t t – – t 9.7 0.6 t – – – – – –
42 Neral (cis-citral) 1232 1235 0.8 – – – – – – t 27.6 10.6 t t – t t – –
43 p-Anisaldehyde 1245 1247 – – t – – – – – – – t – 0.1 – – – –
44 Geraniol 1250 1249 0.3 0.5 – 0.2 0.2 0.9 – t 6.8 0.7 t – – – t – –
45 Linalyl acetate 1254 1254 – 0.4 – 0.5 0.4 t – t – t t 0.1 t 0.2 0.5 – –
46 Geranial (trans-citral) 1264 1264 – t – – – – – – 36.9 14.3 – – – t – – –
47 Bornyl acetate 1282 1284 t t 1.2 t t t t – – t 0.3 t t 0.3 – 0.1 t
48 Thymole 1286 1289 – – – – – – – – – – – – – – – – t
49 Geranyl formate 1296 1298 – – – – – – – – 0.2 – – – – – – – –
50 Carvacrol 1298 1298 – – – – – – – t – – t – – – – – t
51 (Z)-Methyl cinnamate 1303 1299 – – – – – – – – – – – 5.8 – 6.1 – – –
52 Undec-(9Z)-en-1-al 1323 1322 – – – – – – – – – 0.1 – – – – – – –
53 ␣-Cubebene 1346 1345 – – t 0.2 0.3 0.4 – 0.1 – – t – – – t t 0.3
54 Citronellyl acetate 1350 1350 – – – – – – – – 0.6 – – – – – – – –

13
 14
Table 2 (Continued)

S. no. Compounda RIb RIc Content (%)d

O1 O2 O3 O4 O5 O6 O7 O8 O9 O10 O11 O12 O13 O14 O15 O16 O17

55 Eugenol 1355 1356 0.4 4.7 – t – t 5.9 t – – – t t 0.1 t 68.0 54.9

56 Neryl acetate 1361 1359 – – – – – – – – 0.7 0.1 – – – – – – –
57 Cyclosativene 1367 1369 – – – – – – – – – – – – – – – – –
58 ␣-Copaene 1376 1374 t – t t t t 0.3 1.9 t – t t – t 0.6 0.1 0.9
59 (E)-Methyl cinnamate 1380 1376 – – – – – – – – – – – 38.7 – 33.3 – – –
60 Geranyl acetate 1380 1379 – – – – – – – – 0.2 t – – – – – – –
61 (3Z)-Hexenyl-(3Z)-hexenoate 1384 1383 – – – – – – – – – 0.2 – – – – – – –
62 ␤-Cubebene 1389 1387 – – – – – – – – – – 0.1 – – – – t –
63 ␤-Bourbonene 1390 1387 – – – – – – 0.1 – – – – – – – – – –
64 ␤-Elemene 1391 1389 0.1 t – t – t 0.2 0.1 – – 0.8 t 0.2 0.5 0.1 4.1 10.2
65 Methyl eugenol 1403 1403 0.1 – 0.5 t – 14.9 – 0.8 – – 0.5 t 1.2 0.2 4.3 4.7 0.3
66 (Z)-Isoeugenol 1405 1406 – – – – – – – – – – – – – – – – t
67 ␣-Gurjunene 1411 1409 – – – – – – 0.1 – t t – – – – – – –
68 (E)-Caryophyllene 1420 1417 t 0.6 0.3 1.1 0.6 0.9 2.3 5.2 1.3 4.9 0.7 t 1.5 1.3 1.6 8.4 4.0
69 ␤-Copaene 1425 1430 – – 0.2 0.4 0.5 – 0.2 – – – – t 0.1 0.1 – t 0.2
70 trans-␣-Bergamotene 1429 1432 t t – – – – 2.2 2.3 0.7 5.9 t t – 0.5 t t t

R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19

71 Aromadendrene 1439 1439 – – – – – 0.2 0.7 – – 0.3 0.4 t – – 0.2 0.1 t
72 (Z)-␤-Farnesene 1441 1440 – – – – – – – – 0.1 – t 0.1 0.1 – – – –
73 ␣-Humulene 1452 1452 – 0.2 0.1 0.5 0.4 t 1.3 0.2 t 2.7 0.2 0.1 0.4 0.1 0.2 t 0.1
74 (E)-␤-Farnesene 1456 1454 – – t – – t 1.9 0.2 – t t 0.2 0.1 0.3 0.2 – t
75 Allo-Aromadendrene 1458 1458 – – – – – – t – – – 0.1 t – 0.2 – – –
76 ␥-Muurolene 1472 1478 t – 0.1 0.6 0.6 0.2 0.1 0.1 – – t – 0.2 – 0.9 – –
77 Germacrene D 1476 1484 1.9 – – 0.1 0.1 t 2.6 – 0.9 0.1 0.7 0.5 – 0.6 – 0.3 5.3
78 ␤-Selinene 1490 1489 t 4.4 0.3 0.2 0.1 – – 1.7 – – – t – 0.1 – 0.8 0.6
79 ␣-Selinene 1497 1498 t 1.5 0.5 t t 0.4 0.1 – – 0.2 – 0.2 0.7 0.7 t 1.1 1.4
80 Bicyclogermacrene 1498 1500 0.6 – 0.6 0.1 0.3 0.1 – 0.4 – – 0.9 0.4 0.2 0.3 0.2 – –
81 (E,E)-␣-Farnesene 1505 1505 – – – – – – – – – 0.3 – – – – – – –
82 ␤-Bisabolene 1510 1505 – – – – – – 25.6 – 0.2 – 0.7 – – – t – –
83 ␥-Cadinene 1515 1513 – – – t t 0.3 – 0.7 – 0.1 t – 0.4 – 0.4 0.6 1.4
84 Eugenol acetate 1520 1521 – – – – – – – – – – – – – – – t 0.5
85 ␤-Seaquiphellandrene 1522 1521 – – – – – – 0.3 – – t – – – – – – –
86 ␦-Cadinene 1520 1522 – – – – t 0.1 t 0.5 – t – t – 1.1 0.4 0.1 1.1
87 (E)-␥-Bisabolene 1533 1529 – – – – – – – – 3.9 – t – 0.1 – – – –
88 ␣-Cadinene 1542 1537 – – – 1.6 1.6 t 11.9 – – – t – – – – t t
89 (E)-Nerolidol 1563 1561 – – 0.1 0.2 0.2 0.1 – 0.1 t 0.4 t – – – t – t
90 Spathulenol 1574 1577 – – 0.1 0.1 t 0.5 – 0.6 – t 0.1 0.1 t 0.7 0.4 0.8 0.9
91 Caryophyllene oxide 1577 1582 t t – 0.1 0.1 0.4 1.1 0.1 0.5 – 0.1 0.1 t 0.1 0.1 0.1 t
92 Guaiol 1591 1600 – – – – – – – 0.1 – – – – t – – 0.1 0.1
93 Humulene epoxide-II 1600 1608 – – 0.2 t t 0.1 – 0.1 0.2 – t – – – t 0.1 0.4
94 ␥-Eudesmol 1626 1630 – – – – – – – 1.9 – – 1.5 – t – – 0.1 t
95 epi-␣-Cadinol 1640 1638 – t 1.8 t t 1.3 – – – 0.1 – – – – – – –
96 epi-␣-Muurolol 1643 1640 – – – – – – – – – – – – – – – t –
97 ␣-Muurolol 1645 1644 – – – – t t t – – – – – 0.1 – – t 0.3
98 ␤-Eudesmol 1648 1649 1.1 2.5 0.2 0.4 0.3 0.8 – – – – t – 0.1 – 0.7 1.2 1.6
99 ␣-Cadinol 1652 1652 – – – – – – – 1.2 t 0.1 0.1 2.5 t 3.4 t – –
100 ␤-Bisabolol 1677 1674 – – – 0.1 0.1 0.1 t 0.5 – 0.3 t 0.1 t 0.2 0.4 0.1 0.4
101 epi-␣-Bisabolol 1682 1683 – – – – – – – – 0.3 – – – – – – – –

Total identified 98.6 97.0 98.0 97.3 98.1 84.3 92.2 96.3 95.2 89.4 97.1 96.6 97.2 97.7 93.6 93.1 88.2

Note: for abbreviation of different Ocimum accessions please see Table 1.

a
Compounds were identified by comparison of their RI (determined rel. to n-alkanes (C8 –C30 ) and mass spectra with those of authentic compounds or with databases) (see Exper. part).
b
RI: retention index on the non-polar GC column DB-5.
c
RI literature (Adams, 2007).
d
The contents (%) of the individual components were calculated based on the peak area (FID response) without using a correction factor. t, trace (<0.05%).
e
Tentative identification.
Table 3
Compositional variation in the essential oils of Ocimum populations (O18–O34).

S. no. Compounda RIb RIc Content (%)d

O18 O19 O20 O21 O22 O23 O24 O25 O26 O27 O28 O29 O30 O31e O32 O33 O34

1 Tricyclene 924 921 – – – – – – – – – t – – – – – – 0.1

2 ␣-Thujene 928 924 – – – – – t 0.1 0.1 0.1 t – t – 0.2 0.2 t 0.1
3 ␣-Pinene 935 932 0.4 0.2 0.3 t t 0.1 0.8 0.4 1.1 0.2 0.4 0.4 0.7 1.4 – 0.3 4.9
4 ␣-Fenchene 940 945 – – – – – – – – – – – – – – – – t
5 Camphene 942 946 – t t – t 0.4 4.3 5.1 – t t 0.1 – 6.4 – – 4.6
6 Thuja-2,4(10)-diene 949 953 – – – – – – – – – – – – – – – – 0.1
7 Verbenene 959 961 – – – t – – – – – t t – 1.7 – – – –
8 Sabinene 967 969 – – – – – – – – – 0.9 0.8 t – – – – –
9 1-Octen-3-ol 975 974 – – – – – – – – – – – – – – – – 1.4
10 ␤-Pinene 975 974 t t t t 0.2 0.1 1.4 0.1 – 0.2 – 0.9 – t 0.4 0.5 0.4
11 3-Octoanone 980 979 – – – t – – – – – t t – – – – – 0.1
12 Myrcene 988 988 t t – – – 0.2 0.9 0.9 0.9 t 0.1 0.2 0.3 1.2 0.2 0.2 1.3
13 3-Octanol 989 988 – – – – – – – – – – – – – – – – 0.1

R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19

14 n-Octanal 999 998 – – – 0.1 0.1 – – – – – – – – – – – –
15 n-Decane 999 1000 – – – – – – – – – t t – – – – – –
16 (3E)-Hexenyl acetate 1000 1001 – – – – – – – – – – – – – – – – t
16 ␣-Phellandrene 1002 1002 – – – – t – – – – t 0.6 – – – – – 0.2
18 ␣-Terpinene 1015 1014 – – – – – t t 0.1 0.1 t – – – 0.5 0.1 0.1 0.1
19 p-Cymene 1021 1020 t t – – t t – – t t 0.2 t t – 0.2 0.1 1.1
20 Limonene 1025 1024 t t t 0.2 t 0.4 22.7 10.3 29.8 t t t t 8.7 – t 6.3
21 1,8-Cineole 1028 1026 – – – t 0.2 – t t t 14.4 16.7 15.0 20.9 0.7 – – 0.2
22 (Z)-␤-Ocimene 1030 1032 – – – – – – – – – t t t t – 11.0 15.5 t
23 (E)-␤-Ocimene 1043 1044 0.1 t – – – 1.0 1.1 0.3 1.8 1.2 0.2 3.1 2.1 3.0 1.2 1.1 t
24 ␥-Terpinene 1054 1054 – t – – – – 0.2 1.6 t t 0.1 0.7 0.5 0.6 0.1 0.2 0.3
25 cis-Sabinene hydrate 1062 1065 – – – 0.1 0.1 t 0.2 – 0.2 0.9 t – – 1.4 t 0.2 0.4
26 cis-Linalool oxide 1072 1067 – – – – – – – 1.0 1.8 – – – – – – – –
27 Fenchone 1079 1083 – – – – – t – – – – t – – – – – –
28 trans-Linalool oxide 1080 1084 0.1 – – – – t – – – – – – – – – – –
29 Terpinolene 1084 1086 – – – – – – – – – t – 0.3 0.5 1.1 0.4 0.3 0.3
30 Linalool 1099 1095 t t t t t 84.1 0.8 3.3 0.9 0.3 0.3 0.1 – 1.4 0.3 t –
31 trans-Sabinene hydrate 1102 1098 – – – t t – – – – t t – – – – – 0.7
32 3-Octanol acetate 1120 1120 – – – – – – – – – – – – – – – – t
33 Camphor 1144 1141 – t – – 0.3 6.0 56.9 55.5 43.5 t t t – 64.9 – – 45.6
34 Isopulegol 1147 1145 – – – – – – – – – – – – – – – – 0.1
35 Citronellal 1148 1148 – – – – – – – – – t t 0.3 – – – – –
36 Iso-isopulegol 1155 1155 – – – – – – – – – – – – – – – – 0.3
37 Borneol 1163 1165 – t 0.2 t t t 0.9 2.6 t – 0.3 – 0.3 t 0.3 0.6 1.0
38 Menthol 1170 1167 – – – – – – – – – t t – – – – – –
39 Terpinen-4-ol 1177 1174 t t – – t 0.1 t 0.4 0.7 t 0.5 – – 0.6 t t 0.3
40 p-Cymen-8-ol 1180 1179 – – – – – – – – – – – – – – – – t
41 ␣-Terpineol 1186 1186 0.3 1.0 – – t 0.1 0.7 1.6 0.8 – 1.1 0.6 – 2.7 0.1 0.1 0.1
42 Myrtenol 1195 1194 – – – – – – – – – – – – – – – 0.1 2.8
43 Methyl chavicol 1200 1195 – – – – – – – – – 23.3 7.7 8.4 15.9 – – – –
44 Verbenone 1208 1204 – – – – – – – – – – – – – – – – 0.1
45 Nerol 1227 1227 – – – – – t – – – – – – – – – – –
46 Geraniol 1250 1249 – – – – – 0.1 – – – – – – – – – – –
47 Linalyl acetate 1254 1254 – – – – – t – – – – – – – – – – –
48 Bornyl acetate 1282 1284 t – 0.2 0.1 0.1 t t t 0.1 t t – 0.1 – t t –
49 ␣-Cubebene 1346 1345 t t t – 0.2 – 0.1 0.1 t – t – – – 0.1 t –
50 Eugenol 1355 1356 1.5 75.9 77.3 18.9 74.3 t – – – 30.0 52.4 9.5 4.5 – 66.3 66.4 0.1

15
 16
Table 3 (Continued)

S. no. Compounda RIb RIc Content (%)d

O18 O19 O20 O21 O22 O23 O24 O25 O26 O27 O28 O29 O30 O31e O32 O33 O34

51 Cyclosativene 1367 1369 – – – – – t t 0.4 t – – – 0.1 – – – t

52 ␣-Copaene 1376 1374 t t 0.2 0.2 t 0.1 t 0.1 0.1 t t 0.2 0.1 – 1.2 0.4 0.3
53 Geranyl acetate 1376 1379 – – – – – – – – – t t 0.5 – – – – –
54 2-epi-␣-Funebrene 1378 1380 – – – – – – – – – t t – – – – – –
55 (3Z)-Hexenyl-(3Z)-hexenoate 1384 1383 – – – – – – – – – – – – 0.2 – – – t
56 ␤-Cubebene 1389 1387 0.2 – 0.6 – 1.1 – – – – – – – – – – – –
57 ␤-Elemene 1391 1389 4.5 15.0 10.0 23.4 10.7 t t t 0.2 t t t t t 0.3 0.2 0.7
58 Methyl eugenol 1403 1403 79.7 0.1 0.1 t 0.1 t – – 0.2 0.3 0.1 0.2 t – 0.2 0.2 0.2
59 (Z)-Isoeugenol 1405 1406 – t – – – – – – – – – – – – – – –
60 (Z)-Caryophyllene 1410 1408 – – – 2.3 t – – – – – – – 1.9 – – – 0.1
61 ␣-Gurjunene 1411 1409 – – – – – – – – – – – – – – – – 0.1
62 cis-␣-Bergamotene 1412 1411 – – t 0.2 0.1 – – – – t t t t – – – –

R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19

63 (E)-Caryophyllene 1420 1417 5.0 3.0 4.9 36.2 8.2 2.0 t t 0.2 2.0 0.4 2.5 t 1.2 2.3 1.6 2.9
64 ␤-Copaene 1425 1430 – – – t – t t 0.1 – 0.9 – 0.2 2.0 – t t –
65 trans-␣-Bergamotene 1429 1432 – – – – – – – – – t 0.7 2.3 – – t – 0.7
66 ␣-Guaiene 1438 1437 – – t – t – – – – – – – – – – – 4.2
67 Aromadendrene 1441 1439 t t – 0.3 – – – – t 0.3 0.2 – – – – – –
68 (Z)-␤-Farnesene 1443 1440 – – – – t – – – – 3.3 0.9 0.9 0.5 – – – –
69 Seychellene 1444 1444 – – – – – – – – – – – – – – – – 0.6
70 ␣-Humulene 1452 1452 0.4 1.0 0.3 2.4 0.5 0.2 t 0.2 – – – 1.7 – – t t 0.2
71 (E)-␤-Farnesene 1456 1454 – – – – t – – t 0.3 t t 1.5 0.9 0.2 t t t
72 Allo-Aromadendrene 1458 1458 – – – – – – – – – t t – – – – – t
73 cis-Muurola-4(14), 5-diene 1464 1465 – – 0.1 t – – – – – – – – – – – – –
74 ␥-Gurjunene 1474 1475 – – – – – – – – – t t – – – – – t
75 ␥-Muurolene 1476 1478 t – t 0.1 t – – 1.9 – – – – 1.6 – – – –
76 ar-Curcumene 1479 1479 – – – – – – – – – t t t – – – – –
77 Germacrene D 1481 1480 0.1 0.5 0.9 0.7 0.2 1.3 t t 1.5 0.1 – 0.5 0.3 0.7 4.8 5.3 0.1
78 ␤-Selinene 1490 1489 0.1 t 0.1 0.7 0.2 – – 2.0 – – – – 0.1 – – – 4.0
79 ␣-Selinene 1497 1498 0.1 t – 2.6 – 1.1 t 0.3 – t 0.3 – t – 0.2 0.2 3.0
80 Bicyclogermacrene 1498 1500 – – – t – t – 0.1 0.1 – – – t – t t –
81 Germacrene A 1507 1508 – – – 0.1 – – – – – – – – – – – – –
82 ␤-Bisabolene 1510 1505 0.5 – 1.1 – 1.1 – – – – 8.5 4.5 22.9 14.7 – – t 0.1
83 ␥-Cadinene 1515 1513 0.1 t 0.1 t t – t 0.3 0.3 – – – 0.2 – 0.1 t –
84 Cubebol 1516 1514 – – – – – – – – – – – – – – – – 0.1
85 Sesquicineole 1517 1515 – – – – – – – – – 0.1 t 0.3 – – – – –
86 7-epi-␣-Selinene 1520 1520 – – – – – – – – – – – – – – – – 0.1
87 Eugenol acetate 1522 1521 – t – – – – – – – – – – – – – 0.2 –
88 ␤-Seaquiphellandrene 1522 1521 – – – – – – – – – t t – t – – – –
89 ␦-Cadinene 1524 1522 t t – 0.3 – t – t – – – – – – 0.6 – 0.5
90 (E)-␣-Bisabolene 1542 1537 – – – – – t t t – 6.7 3.0 10.9 9.0 – t – t
91 (E)-Nerolidol 1563 1561 t – – t – 0.1 t 0.5 – t – 0.6 0.1 – 0.1 – 0.1
92 Longipinanol 1570 1567 – – – – – – – – – – – – – – – – 7.4
93 Germacrene D-4-ol 1576 1574 – – – – 0.1 – – – – – – – – – – 0.1 0.1
94 Spathulenol 1579 1577 – t 0.1 – 0.1 0.1 t 0.2 0.3 0.2 t – 1.1 – t 0.7 0.1
95 Caryophyllene oxide 1584 1582 0.8 0.4 t 3.6 0.1 t t 0.1 0.1 0.2 0.4 1.4 0.4 0.1 1.2 t 0.4
96 Guaiol 1591 1600 – t – – – – – – – 0.1 t – – – 0.1 – –
97 Humulene epoxide-II 1600 1608 0.1 t 0.1 t t – – – t 0.1 0.8 t 0.1 – t – –
 R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19 17

for this species are eugenol type (Mondello et al., 2002; Padalia and

99.1
O34
Verma, 2011), methyl eugenol type (Jirovetz et al., 2003; Kothari

–
–
–

–
–
–
–
–
t
et al., 2005), and methyl chavicol type (Brophy et al., 1993). Present
study found three chemotypes, viz. eugenol, methyl eugenol, and

95.1
O33

0.4

0.1
(E)-caryophyllene/␤-elemene/eugenol types for this species. Out
–
–
–
–

–
–

–
of these three chemotypes, the population rich in sesquiterpenes
and deficient in eugenol content is a new report to this region.

93.6
O32

0.1
0.2
0.2
0.1
0.5

0.5
However, O. tenuiflorum chemotype rich in ␤-caryophyllene is

–
–

–
reported earlier from other region (Simon et al., 1990).
O. kilimandscharicum populations grown in different parts of the
O31e

97.0
world have also been investigated by many researchers. In addi-
–
–
–
–
–
–
–
–
–
tion to the camphor chemotype (Padalia and Verma, 2011; Runyoro

Compounds were identified by comparison of their RI (determined rel. to n-alkanes (C8 –C30 ) and mass spectra with those of authentic compounds or with databases) (see Exper. part).
et al., 2010), other chemotypes of O. kilimandscharicum have also
83.1
O30

0.1

0.6
0.9

0.7

been found. Essential oil rich in linalool/camphor is reported from

–

–
t
t

Greece (Charles and Simon, 1992). An oil of O. kilimandschar-

icum grown in Rwanda (Central Africa) was reported to contain
86.8
O29

0.4
0.2

1,8-cineole as the major constituent and only traces of camphor

–
–
–
–
–

–
–

(Ntezurubanza et al., 1984). Further, present study identified three

chemotypes for O. kilimandscharicum, in which two (camphor and
93.0
O28

0.1
0.1

0.1

–
–
–

linalool/camphor types) were common with earlier reports; how-

t

ever, third one, rich in multiple components, viz. eugenol/methyl

94.6
O27

chavicol/␤-bisabolene/1,8-cineole/(E)-␣-bisabolene was reported

0.4
–
–
–
–

–
t

for the first time.

Literature survey revealed that the essential oil composition
86.6
O26

of O. gratissimum have been investigated in detail. Chemotypes

1.1

0.1

0.3
–
–
–

–
t

described so far for this species are eugenol (Dambolena et al., 2010;
The contents (%) of the individual components were calculated based on the peak area (FID response) without using a correction factor.

Jirovetz et al., 2003; Vieira et al., 2001), thymol/␥-terpinene/p-

90.2
O25

cymene (Tchoumbougnang et al., 2006), and geraniol (Vieira et al.,

0.1

0.5
–
–

–
–
–
t

2001) types. The O. gratissimum populations investigated in present

study belong to eugenol/(Z)-␤-ocimene chemotype.
91.5
O24

0.4

Studies conducted earlier on essential oil composition of O.

–

–
–
–
–
–
t
t

americanum showed that the number of chemotypes exists for this

species. Reported chemotypes of this species are citral (Mondello
97.8
O23

0.3

et al., 2002), 1,8-cineole/(Z)-␤-ocimene (Tchoumbougnang et al.,

–

–
–
–
t
t
t

2006), (E)-methyl cinnamate/linalool (Vina and Murillo, 2003), (Z)-

methyl cinnamate/(E)-methyl cinnamate (Jirovetz et al., 2003),
98.0
O22

(E)-caryophyllene/methyl eugenol (Vina and Murillo, 2003), cam-

–
–
–
–
–
–
–
–
–

phor/limonene (Mondello et al., 2002), and methyl chavicol

(Gupta, 1996). Moreover, present study showed that cam-
92.9
O21

0.4

phor/longipinanol/limonene chemotype of O. americanum was

–
–
–
–
–
–
–
t

prevalent in Tarai region of north India.

According to the biosynthetic origins of major compounds,
96.6
O20

–
–
–
–
–
–
–

Lawrence (1988) classified them as chemotypes with single or

t

dual biosynthetic pathways. In a study of essential oils of different

97.1

geographical origins, Lawrence (1988) found that the main con-

O19
Content (%)d

Note: for abbreviation of different Ocimum accessions please see Table 1.

–

–
–
–

–
t
t
t

stituents of essential oil of basil are produced by two different

biochemical pathways; the phenylpropanoids, viz. methyl chavi-
94.0
O18

col, eugenol, methyl eugenol, and methyl cinnamate are produced

–
–
–
–
–
–
–
–
–

by the shikimic acid pathway, and the terpenes, viz. linalool, cit-
RI: retention index on the non-polar GC column DB-5.

ral, geraniol etc. by the mevalonic acid pathway. It showed that,

1630
1638
1640
1644
1649
1652
1671
1674
1742

the Ocimum populations investigated in present study are produc-

RIc

ing essential oils either by shikimic acid pathway (e.g., O3, O11, and
Padalia and Verma (2011). t, trace (<0.05%).

O13; O6, O8, and O15; O16, O17, O19, O20, O22, O32, and O33; O18)
or by mevalonic acid pathway (e.g., O2 and O23; O9; O10; O24-O26
1626
1640
1643
1645
1648
1652
1674
1677
1743
RIb

and O31; O34). However, Ocimum populations dominated by both

kind of compounds, viz. phenylpropanoids and terpenes (e.g., O12
and O14; O21) followed dual biosynthetic pathways.
(6R,7R)-Bisabolone

RI literature (Adams, 2007).

The taxonomy of Ocimum is complex due to interspecific

epi-␣-Muurolol

Total identified
n-Tetradecanol
epi-␣-Cadinol

hybridization and polyploidy of the species in the genus (Paton

␤-Eudesmol
␥-Eudesmol
Compounda

␣-Muurolol

␤-Bisabolol
␣-Cadinol

et al., 1999; Pushpangadan and Bradu, 1995; Simon et al., 1990).

Further, the occurrence of huge chemical variations among Oci-
Table 3 (Continued)

mum populations collected from diverse localities seems to be

due to the divergent climatological and geographical conditions
as well as different genetical factors such as subspecies, natu-
S. no.

98
99
100
101
102
103
104
105
106

ral hybridization and chemovariety (Krishnan, 1981; Simon et al.,

a

1990).
18 R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19
Fig. 2. MDS plot based on Euclidean distance scaling depicting phytochemical proximities among the studied 34 Ocimum spp populations. For the abbreviations of the
population name, see Table 1.
4. Conclusions
The results of present study clearly indicated that there were
large amount of morphological and biochemical diversity among
the investigated populations of genus Ocimum. Populations with
similar chemotypic characters differed considerably in morpho-
logical features (e.g., O3, O11, and O13; O2 and O23 etc.).
Therefore, chemical constituents could not be necessarily cor-
related with their taxonomical category. From small (20–30 cm
tall) annual to shrubby (∼1.0 m) perennial Ocimum taxa, with
several chemotypic forms, were found to grow in this region,
which were rich in potential phytomolecules, viz. methyl chav-
icol, linalool, methyl cinnamate, citral, eugenol, methyl eugenol,
and camphor. These chemotypes offers the great opportunity
for production of Ocimum to meet the market requirements
of specific essential oils or individual compounds. Further,
present study of Ocimum taxa revealed the existence of four
new essential oils/chemotypes which were not described before
from this region. The newly explored compositions were two
chemovariants for O. basilicum, viz. ␤-bisabolene/1,8-cineole/␣-
cadinene/methyl chavicol/eugenol type, and linalool/citral type;
one chemovariant for O. tenuiflorum, viz. (E)-caryophyllene/␤-
elemene/eugenol; and one chemovariant for O. kilimandscharicum,
rich in multiple components, viz. eugenol/methyl chavicol/␤-
bisabolene/1,8-cineole/(E)-␣-bisabolene. Moreover, this study has
contributed to the primary knowledge of Ocimum populations from
an under-explored region. The genetic resources of the genus grow-
ing wild or for the medico-religious purposes in this region could
be utilized in further research programme to ensure the production
of high value fragrance chemicals for industrial use.
Acknowledgements
Authors are thankful to the Director, CSIR-Central Institute of
Medicinal and Aromatic Plants (CIMAP) for providing necessary
facilities and encouragement.
References
Adams, R.P., 2007. Identification of Essential Oil Components by Gas Chromatogra-
phy/Mass Spectrometry. Allured Publishing Corporation, Carol Stream, IL, USA.
Alves, P.B., Filho, P.S.F., Moraes, V.R.S., Blank, A.F., Filho, J.L.S.C., Arrigoni-Blank, M.F.,
Oliva, G., Thiemann, O.H., 2007. Chemical composition of essential oil from seven
Ocimum basilicum L. accessions, Brine Shrimp Lethality bioassay and inhibitory
activities against GAPDH and APRT. J. Essent. Oil Res. 19 (1), 89–92.
Brophy, J.J., Goldsack, R.J., Clarkson, J.R., 1993. The essential oil of Ocimum tenuiflorum
L. (Lamiaceae) growing in Northern Australia. J. Essent. Oil Res. 5 (4), 459–461.
Charles, D.J., Simon, J.E., 1992. Essential oil constituents of Ocimum kilimandschar-
icum Guerke. J. Essent. Oil Res. 4 (2), 125–128.
Dambolena, J.S., Zunino, M.P., Lopez, A.G., Rubinstein, H.R., Zygadlo, J.A., Mwangi,
J.W., Thoithi, G.N., Kibwage, I.O., Mwalukumbi, J.M., Kariuki, S.T., 2010. Essential
oils composition of Ocimum basilicum L. and Ocimum gratissimum L. from Kenya
and their inhibitory effects on growth and fumonisin production by Fusarium
verticillioides. Innov. Food Sci. Emerg. Technol. 11, 410–414.
Gupta, S.C., 1996. Variation in herbage yield, oil yield and major component of var-
ious Ocimum species/varieties (chemotypes) harvested at different stages of
maturity. J. Essent. Oil Res. 8 (3), 275–279.
Gupta, S.C., Tava, A., 1997. Chemical composition of an improved hybrid strain of
Ocimum canum Sims. (RRL-OC-11). J. Essent. Oil Res. 9 (3), 375–377.
Hassanpouraghdam, M.B., Hassani, A., Shalamzari, M.S., 2010. Menthone and
Estragole rich essential oil of cultivated Ocimum basilicum L. from Northwest
Iran. Chemija 21 (1), 59–62.
Jirovetz, L., Buchbauer, G., Shafi, M.P., Kaniampady, M.M., 2003. Chemotaxonomical
analysis of the essential oil aroma compounds of four different Ocimum species
from southern India. Eur. Food Res. Technol. 217, 120–124.
Koba, K., Poutouli, P.W., Raynaud, C., Chaumont, J.P., Sanda, K., 2009. Chemical com-
position and antimicrobial properties of different basil essential oils chemotypes
from Togo. Bangladesh J. Pharmacol. 4, 1–8.
Kothari, S.K., Bhattacharya, A.K., Ramesh, S., Garg, S.N., Khanuja, S.P.S., 2005. Volatile
constituents in oil from different plant parts of methyl eugenol-rich Ocimum
tenuiflorum L.f. (syn. O. sanctum L.) grown in South India. J. Essent. Oil Res. 17,
656–658.
Krishnan, R., 1981. Natural out crossing in sweet basil – Ocimum basilicum L. Indian
Perfumer 25 (3 and 4), 74–77.
Labra, M., Miele, M., Ledda, B., Grassi, F., Mazzei, M., Sala, F., 2004. Morphologi-
cal characterization, essential oil composition and DNA genotyping of Ocimum
basilicum L. cultivars. Plant Sci. 167, 725–731.
Lawrence, B.M., 1988. In: Lawrence, B.M., Mookheyee, B.D., Willis, B.J. (Eds.), Devel-
opments in Food Sciences, Flavors and Fragrances: A World Perspective. Elsevier,
Amsterdam.
Lee, J., Scagel, C.F., 2009. Chicoric acid found in basil (Ocimum basilicum L.) leaves.
Food Chem. 115, 650–656.
Mondello, L., Zappia, G., Cotroneo, A., Bonaccorsi, I., Chowdhury, J.U., Yusuf, M., Dugo,
G., 2002. Studies on the essential oil bearing plants of Bangladesh, Part VIII.
Composition of some Ocimum oils O. basilicum L. var. purpurascens, O. sanctum
 R.S. Verma et al. / Industrial Crops and Products 45 (2013) 7–19
L. green, O. sanctum L. purple, O. americanum L. citral type, O. americanum L.
camphor type. Flavour Frag. J. 17, 335–340.
Ntezurubanza, L., Scheffer, J.J.C., Looman, A., Svendsen, A.B., 1984. Composition of
essential oil of Ocimum kilimandscharicum grown in Rwancla. Planta Med. 50,
385–388.
Padalia, R.C., Verma, R.S., 2011. Comparative volatile oil composition of four Ocimum
species from northern India. Nat. Product Res. 25 (6), 569–575.
Paton, A., Harley, R.M., Harley, M.M., 1999. Ocimum: an overview of relationships
and classification. In: Holm, Y., Hiltunen, R. (Eds.), Medicinal and Aromatic Plants
and Industrial Profiles. The Genus Ocimum. Harwood Academic Press, Amster-
dam.
Prakash, P., Gupta, N., 2005. Therapeutic uses of Ocimum sanctum Linn (Tulsi) with
a note on eugenol and its pharmacological actions: a short review. Indian J.
Physiol. Pharmacol. 49 (2), 125–131.
Pushpangadan, P., Bradu, B.L., 1995. Basil. In: Chadha, K.L., Gupta, R. (Eds.), Advances
in Horticulture, Medicinal and Aromatic Plants, vol. 11. Malhotra Publishing
House, New Delhi, pp. 627–657.
Runyoro, D., Ngassapa, O., Vagionas, K., Aligiannis, N., Graikou, K., Chinou, I., 2010.
Chemical composition and antimicrobial activity of the essential oils of four
Ocimum species growing in Tanzania. Food Chem. 119 (1), 311–316.
Simon, J.E., Quinn, J., Murray, R.G., 1990. Basil: a source of essential oils. In: Janik,
J., Simon, J.E. (Eds.), Advances in New Crops. Timber Press, Portland, OR, pp.
484–489.
19
Sirkar, N.N., 1989. Pharmacological basis of ayurvedic therapeutics. In: Atal, C.K.,
Kapoor, B.M. (Eds.), Cultivation and Utilization of Medicinal Plants. PID, CSIR,
New Delhi.
Tchoumbougnang, F., Zollo, P.H.A., Avlessi, F., Alitonou, G.A., Sohounhloue, D.K.,
Ouamba, J.M., Tsomambet, A., Okemy-Andissa, N., Dagne, E., Agnaniet, H.,
Bessiére, J.M., Menut, C., 2006. Variability in the chemical compositions of the
essential oils of five Ocimum species from tropical African area. J. Essent. Oil Res.
18 (2), 194–199.
Telci, I., Bayram, E., Yılmaz, G., Avci, B., 2006. Variability in essential oil composition
of Turkish basils (Ocimum basilicum L.). Biochem. Syst. Ecol. 34, 489–497.
Verma, R.S., Padalia, R.C., Chauhan, A., 2012. Variation in the volatile terpenoids of
two industrially important basil (Ocimum basilicum L.) cultivars during plant
ontogeny in two different cropping seasons from India. J. Sci. Food Agric. 92,
626–631.
Vieira, R.F., Grayer, R.J., Paton, A., Simon, J.E., 2001. Genetic diversity of Ocimum
gratissimum L. based on volatile oil constituents, flavonoids and RAPD markers.
Biochem. Syst. Ecol. 29, 287–304.
Vina, A., Murillo, E., 2003. Essential oil composition from twelve varieties of basil
(Ocimum spp) grown in Colombia. J. Braz. Chem. Soc. 14 (5), 744–749.
Yavari, M., Mirdamadi, S., Masoudi, S., Tabatabaei-Anaraki, M., Larijani, K., Rustaiyan,
A., 2011. Composition and antibacterial activity of the essential oil of a green
type and a purple type of Ocimum basilicum L. from Iran. J. Essent. Oil Res. 23 (1),
1–3.