Ethnobotanical Leaflets 13: 83-88. 2009.

 

 

Inter- Generic Relationship of Ocimum and Origanum Based on GC-MS Volatile Oils Data using Software NTSPSpc Version 2.0

 

M. Maridass

 

Animal Health Research Unit, St. Xavier’s College (Autonomous),

Palayamkottai-627002, Tamil Nadu, South India

Email: orchideyadass@yahoo.com

 

Issued 04 January 2009

Abstract

To study of inter- generic relationships of Origanum vulgare L, Origanum applii (Domin) Borus, Ocimum gratissimum L. and Ocimum basilicum L essential oils peaks based on GC-MS methods. The result of the essential oils of Origanum vulgare L, Origanum applii (Domin) Borus, Ocimum gratissimum L. and Ocimum basilicum were constructed the phenogram formed in one cluster and two sub cluster. The inter- generic relationship of 26% formed in main clusters both genus of Origanum sp and Ocimum sp. The sub cluster I, Ocimum gratissimum L. and O. basilicum was very close relationship of 40% and 34% of sub cluster II Origanum vulgare L, and O. applii respectively.

Introduction

The storage of essential oils in higher plants is not restricted to specialized plant parts. Essential oils occur in both roots, stems, leaves, flowers and seeds, or in the plant as a whole. Both epidermal or mesophyll tissue can function as the site of terpene biosynthesis in general, whereas typical storage cells or cell structures characterize the taxonomic group of aromatic plants: Oil cells, secretory glands, ducts and canals and the Labiatae-typical glandular trichomes (capitate and peltate glands). Depending on morphological structures, varying secondary metabolism and thus, determined signalling and defence functions of essential oils in plant organs, the pronounced alteration gives the ability to obtain essential oil qualities that are quite different from one and the same plant.

Infraspecific and intervarietal differences can be observed in both morphology and chemical structures, which establish the basis for determining important chemically defined populations or chemotypes (Hay and Svoboda,1993). Despite aromatherapeutic demands, which require that the medicinal value of an essential oil be based on its complete composition rather than its constituent parts (Franchomme et al., 1990), one still has to consider the chemical specification of Essential oils with regard to toxic concentrations of single constituents (Tisserand and Balacs, 1995; Price and Price, 1999).

Knowledge about genetic diversity and population genetic structure is a good baseline for formulating effective conservation plans, and can often provide novel, conservation-relevant insights. An effective conservation strategy for a species can be made only after detailed population genetic information becomes available. Estimates of genetic similarity using genetic fingerprinting data are a useful tool in plant breeding, allowing breeders to make informed decisions regarding the selection of germplasm to be used in crossing schemes. Fingerprints themselves are also useful to breeders for every day for protection of their own varieties and to seed producers, growers and end users for checking the identity and purity of their produce (Milbourne et al., 1997).

Materials and Methods

Genetic materials

Origanum vulgare L. (UEC 121.409), O. applii (Domin) Borus (UEC 121.410), Ocimum gratissimum L. (UEC 121.407) and O. basilicum L. (UEC 121.408) were chosen to the present study. The aromatic plants were collected from CPQBA/UNICAMP experimental field, between 9:00 and 10:00 am, in the first week on March, in full flowering, except to O. vulgare L. in vegetative stage (Sartoratto et al., 2004).

Essential oil extraction

The oil extraction was obtained from 40g fresh plants by steam distillation using Clevenger system, during 3h. The aqueous phase was extracted with dichloromethane (3x50mL). The organic phase was dried with sodium sulphate, filtered and the solvent evaporated until dryness. The oil was solubilized in ethyl acetate for gas chromatography and mass spectrometry analysis (Sartoratto et al., 2004).

Chromatography conditions

Sartoratto et al., (2004) work done on the identification of essential oil constituents was conducted by gas-chromatography in Hewlett-Packard 5890 Series II (Palo Alto, CA, USA) equipment, with selective mass detector HP-5971 in the electron impact (EI) ionization mode (70 eV), injector split/splitless, capillary column HP-5 (25 m x 0.2 mm x 0.33 µm). Temperature: injector = 220ºC, column = 60ºC, 3ºC.min-1, 240ºC (7 min). Carrier gas (He) = 1.0 mL.min-1. Retention indices (RI) have been obtained according to the method (Van den Dool, and Kratz, 1963).

Data analysis

The chromatogram peaks were converted into a “1” and “0” matrix, to indicate the presence or absence of a peak, respectively. Genetic similarities (GS) were estimated for all comparisons of each samples according to Nei (1972) as GS=2nxy/(nx+ny) in which nx and ny are the total numbers of peaks in the chromatograms of the samples x and y, respectively, and nxy is the number of peaks shared by the two samples. To examine the inter- genetic relationships between four species populations, a dendrogram was constructed by an unweighted paired group method of cluster analysis using arithmetic averages (UPGMA) option of the NTSYSpc-2.0 software.

Results and Discussion

Table-1 identified with 60 peaks, with the assumption that peaks with the same retention time on different chromatograms was the same compounds. The data on these 60 peaks for four plant samples such as Origanum vulgare L, O. applii (Domin) Borus, Ocimum gratissimum L. and O. basilicum L. (Sartoratto et al., (2004). The phenogram constructed based on common peaks of essential oils identification from the Ocimum gratissimum, O. basilicum, Origanum vulgare L, and O. applii shown in fig-1. The inter generic relationship of 26% formed in main clusters both genus of Origanum sp and Ocimum sp. The graphic phenogram (Fig. 1) distantly placed Origanum vulgare L, and O. applii was 34% similarity.  And Ocimum gratissimum L. and O. basilicum was 40% similarity. According to literature reported the distantly placed C. martinii (var. motia and var. so.a) from the rest of the taxa sharing 54% similarity. C. pendulus was distantly placed from rest of the accessions but was closer to the subclusters containing C. nardus var. nardus, C. nardus var. Java II and C. winterianus (49, 44, 52% similarity, respectively) on one hand and to the subcluster containing C. exuosus and C. citratus (52, 43% similarity, respectively (Khanuja et al.,2005). In conclusion of the present study, essential oils constituents can be used as an additional tool to assist in identification of similar and closely related species of Ocimum sp and Origanum sp.

Acknowledgements

I would like to thank A. Sartoratto, A. L. M. Machado, Camila Delarmelina; Glyn Mara Figueira, Marta Cristina T. Duarte,  and Vera Lúcia G. Rehder, Centro Pluridisciplinar de Pesquisas Químicas, Biológicas e Agrícolas – Universidade Estadual de Campinas, Campinas, SP, Brasil, and also Brazilian Journal Microbiology for  providing of Research article.

References

Hay RKM, Svoboda KP. Botany. In Volatile oil crops: their biology, biochemistry and production, Hay, R.K.M. and Waterman, P.G. (eds.), Longman Scientific & Technical, Harlow;1993: 5-22.

 

Franchomme P, Jollois R, Penoel D. L'aromathérapie exactement: Encyclopédie de l'utilisation thérapeutique des huiles essentielles - Fondements, démonstration, illustration et applications d'une science médicale naturelle. Jollois, Limoges; 1990: 446.

 

Tisserand RB, Balacs T. Essential oil safety: A guide for health care professionals. Churchill Livingstone, London; 1995: 279.

 

Price S, Price N. Aromatherapy for health professionals. Churchill Livingstone, London; 1999:390.

 

Milbourne D, Meyer R, Bradshaw JE, Baird E, Bonar N, Provan J, Powell W, Waugh, R. Comparison of PCR-based marker system for the analysis of genetic relationships in cultivated potato. Mol. Breed, 1997; 3: 127–136.

 

Sartoratto A, Machado ALM., Delarmelina C, Figueira GM,  Duarte MCT, Rehder VLG. Composition And Antimicrobial Activity Of Essential Oils From Aromatic Plants Used In Brazil. Brazilian Journal of Microbiology,2004; 35:275-280.

 

Van den Dool H, Kratz  PDA. Generalization of the retention index system including linear temperature programmed gas-liquid partition chromatography. J. Chrom, 1963:11:463-471.

 

Nei M. Genetic distance between populations. Am.Nat. 1972; 106: 283–292.

 

Khanuja SPS,  Shasany AK,  Pawar A, Lal RK,  Darokar MP,  Naqvi AA,  Rajkumar S,  Sundaresan V,  Nirupama Lal, Kumar S. Essential oil constituents and RAPD markers to establish species relationship in Cymbopogon Spreng. (Poaceae) Biochemical Systematics and Ecology, 2005; 33: 171-186.

 

Table 1. Inter- generic chemotypical variation of Ocimum and Origanum species (Sartoratto et al., 2004).

 

Composition

RRI

Ocimum gratissimum

O. basilicum

Origanum. vulgare

O. applii

1-octen-3-ol

3-octanol

p-cimene

limonene

1,8-cineol

trans--ocimene

-terpinene

isopentyl n-butirate

-terpinolene

fenchone

linalool

cis-p-menth-2-en-1-ol

trans-p-menth-2-en-1-ol

camphor

borneol

Terpin-4-ol

cimen-8-ol

-terpineol

trans-piperitol

thymol methyl ether

neral

carvacrol methyl ether

carvone

geranial

thymol

carvacrol

eugenol

piperitenone oxide

Geranyl acetate

-bourbonene

-elemene

Cedrene <1,7-di-epi-alpha>

trans-caryophillene

-gurjunene

-bergamotene

-guaiene

-farnesene

-humulene

allo-aromadendrene

Germacrene D

Curcumene <AR>

-muurolene

zingiberene

-guaiene

-bisabolene

-cadinene

-cadinene

espatulenol

caryophillene oxide

epi--muurolol

-eudesmol

-cadinol

978

998

1024

1028

1031

1035

1057

1068

1083

1088

1093

1122

1140

1150

1169

1176

1182

1188

1208

1231

1238

1241

1241

1270

1294

1298

1358

1373

1379

1385

1393

1397

1422

1430

1435

1438

1454

1455

1461

1482

1483

1484

1495

1506

1508

1516

1524

1580

1586

1643

1655

1656

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

0.26

-

-

-

-

-

-

-

-

-

-

93.9

-

-

-

-

-

1.08

-

-

-

-

-

-

4.23

-

-

-

-

-

-

-

-

-

-

-

0.16

-

-

-

-

1.05

-

-

-

-

0.53

32.06

-

-

10.1

-

0.99

-

3.90

-

-

-

-

-

-

-

-

28.1

-

-

0.23

1.92

-

2.00

-

1.66

0.29

-

0.77

-

5.49

-

-

-

0.68

-

1.27

0.29

-

-

5.81

0.24

0.38

0.68

-

1.50

-

-

0.39

1.99

-

0.31

-

-

1.47

0.86

-

2.52

33.3

-

4.25

0.16

0.24

-

1.33

-

-

38.0

-

-

-

0.33

-

-

-

2.66

-

-

-

-

-

-

1.47

-

-

-

-

1.05

-

-

1.44

1.07

-

-

0.29

-

-

-

-

-

-

-

-

-

-

4.92

-

-

-

-

3.08

0.64

-

-

1.47

-

5.91

-

-

64.5

-

-

-

-

1.35

-

-

-

0.36

-

-

-

-

0.28

4.79

-

-

-

-

1.98

-

-

1.62

-

0.72

-

1.53

 

Table 2. Volatile data converted on inter-generic chemotypical variation of Ocimum and Origanum species.

 

Composition

RRI

Ocimum gratissimum

O. basilicum

Origanum vulgare

O. applii

1-octen-3-ol

3-octanol

p-cimene

limonene

1,8-cineol

trans--ocimene

-terpinene

isopentyl n-butirate

-terpinolene

fenchone

linalool

cis-p-menth-2-en-1-ol

trans-p-menth-2-en-1-ol

camphor

borneol

Terpin-4-ol

cimen-8-ol

-terpineol

trans-piperitol

thymol methyl ether

neral

carvacrol methyl ether

carvone

geranial

thymol

carvacrol

eugenol

piperitenone oxide

Geranyl acetate

-bourbonene

-elemene

Cedrene <1,7-di-epi-alpha>

trans-caryophillene

-gurjunene

-bergamotene

-guaiene

-farnesene

-humulene

allo-aromadendrene

Germacrene D

Curcumene <AR>

-muurolene

zingiberene

-guaiene

-bisabolene

-cadinene

-cadinene

espatulenol

caryophillene oxide

epi--muurolol

-eudesmol

-cadinol

978

998

1024

1028

1031

1035

1057

1068

1083

1088

1093

1122

1140

1150

1169

1176

1182

1188

1208

1231

1238

1241

1241

1270

1294

1298

1358

1373

1379

1385

1393

1397

1422

1430

1435

1438

1454

1455

1461

1482

1483

1484

1495

1506

1508

1516

1524

1580

1586

1643

1655

1656

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

1

0

0

0

0

0

0

0

0

0

0

1

0

0

0

0

0

1

0

0

0

0

0

0

1

0

0

0

0

0

0

0

0

0

0

0

1

0

0

0

0

1

0

0

0

0

1

1

0

0

1

0

1

0

1

0

0

0

0

0

0

0

0

1

0

0

1

1

0

1

0

1

1

0

1

0

1

0

0

0

1

0

1

1

0

0

1

1

1

1

0

1

0

0

1

1

0

1

0

0

1

1

0

1

1

0

1

1

1

0

1

0

0

1

0

0

0

1

0

0

0

1

0

0

0

0

0

0

1

0

0

0

0

1

0

0

1

1

0

0

1

0

0

0

0

0

0

0

0

0

0

1

0

0

0

0

1

1

0

0

1

0

1

0

0

1

0

0

0

0

1

0

0

0

1

0

0

0

-

1

1

0

0

0

0

1

0

0

1

0

1

0

1

 

 

Table 3. The average of taxonomical similarities of Ocimum sp. and Origanum sp.

Population

Ocimum

gratissimum

O. basilicum

Origanum

 vulgare

O. applii

Ocimum gratissimum

0.0000

 

 

 

O. basilicum

0.4

0.0000

 

 

Origanum vulgare

0.31

0.2

0.0000

 

O. applii

0.32

0.24

0.34

0.0000

 

Fig.1. Phenogram based on 60 essential oils peaks on Ocimum sp and Origanum sp.