GC-Mass and Phytochemical Investigation of Cymbopogon citratus

 

Zahraa Abdulelah Al Naqqash^1*, Huda Khalil Al-Bazaz¹, Fatima Mahdi Salh², Shahad Qusay Ibraheem²

¹Department of Pharmacognosy & Medical Plants, College of Pharmacy,

Mustansiriyah University, Baghdad, Iraq

²(5^thstage students) Department of Pharmacognosy & Medical Plants, College of Pharmacy,

Mustansiriyah University, Baghdad, Iraq

 

ABSTRACT:

As medicinal plants known for their importance in different uses especially pharmaceutical usage due to presence of the active compounds represented by terpens, phenols, alkaloids…etc. and one of these important medicinal plants is our plant under study which is Cymbopogon citratus locally known as lemon grass that belongs to a Poaceae family. It is an aromatic tropical perennial herb, known in Egypt, Arabian Peninsula and North and West tropical Africa. In this study lemon grass was collected from  Baghdad Province Al-Yarmuk area then extracted in two ways; first one was soxhlet extraction method so we got an alcoholic extract and identify the chemical compounds found in it by using general tests which shown the presence of alkaloids, phenols, tannins and flavonoids while cardiac glycosides and saponins were absent; and the other method was hydrodistillation by using Clevenger apparatus and the sample was identified using GC-Mass which revealed the presence of the following compounds((E)-3(10)-Caren-4-ol.7,7-Dimethyl-4-methylenebicyclo[4.1.0]heptan-3-o  and its percentage in plant sample was 15.07% and the other compound (2,6-Octadienal, 3,7-dimethyl-, (E)- .alpha.-Citral  (E)-Citral. trans-Citral trans-3,7-Dimethyl-2,6-octadienal) with percentage 42.27%. We can separate some pure phytopharmaceutical, which in turn can be used as lead molecule for producing the novel agent having good therapeutic activity. This study was conducted to determine the phytoconstituents presents in Cymbopogon citratus plant collected from Baghdad Province Al-Yarmuk area.

 

 

 

INTRODUCTION:

For thousands of years now, natural products mainly from plants have been explored by man in treatment of various diseases [1]. Tradomedicine, over the past decade, has become a subject matter of global importance, playing an essential role on both healthcare system and world economy [2]. In recent times, medicinal plants has maintained their pivotal role in healthcare system of many persons universally.

 

This is particularly true in developing countries, where tradomedicine has a long and uninterrupted history of use [3].

 

Due to increased demand of natural products from medicinal plants in healthcare serves worldwide, herbal plant producers have commenced the use of the most apt extraction methods in order to see and detach the chemical compounds present in them [4]. The main objectives of this study were to determine the active substances content, and to identify the chemical composition of lemongrass essential oil cultivated under southern Nile delta conditions (Bilbeis, Sharqia, Egypt). Lemon grass is one of the most important medicinal plants. The scientific name of lemongrass is Cymbopogon citratus. The name Cymbopogon is derived from the Greek words "kymbe" (boat) and pogon (beard), denoting to the flower spike arrangement [5] which is a genus of about 55 species of grasses [6] while the word citratus originates from the ancient Latin, meaning lemon-scented leaves [7]. Lemongrass is grass that belongs to a Poaceae family. It is an aromatic herb, known in Egypt, in Arabian Peninsula and in the North and West tropical Africa [8]. It is tropical perennial herbs that grow from 60-120 cm high with its leaves green, long and slats and have pleasant aroma and taste (Figure 1) which grows spontaneously around the world, mainly in the tropical and savanna regions. The common names of this grass include; lemongrass, barbed wire grass, silky heads, citronella grass, chadedartigalongue, fever grass, tanglad, hierbaluisa or gavatichaha amongst many others. Cymbopogon citratus is a native of (Southwest Asia) South India but present in many part of the world growing in dense clumps.

 

 

Figure1.1 Cymbopogon citratus leaves

 

In the folk medicine of Brazil and Mexico, C. citratus is believed to have anxiolytic, hypnotic and anticonvulsant potentials. Laboratory studies have shown that C. citratus possesses cytoprotective, hypocholesterolemic, antioxidant, anti-inflammatory antimalarial, antimutagenic, antiprotazoan activities as well as antifungal properties [5-9]. It has been shown to be effective in the treatment of fever and infection, digestive disorder, headaches, menstrual disorder, joint pain, stomach aches and rheumatic pain [10]. However, the leaves of lemongrass are known to have phenolcompounds which can act as antioxidants [6]. The major terpenes in C. citratus contain citral-α or geranial (10%–48%) and citral-β or neral (3%–43%), borneol (5%), geraniol (2.6%–40%), geranyl acetate (0.1%–3.0%), linalool (1.2%–3.4%), and nerol (0.8%–4.5%). There are dribbles of camphene, camphor, α-camphorene, caryophyllene oxide, ∆-3-carene, caryophyllene, 1,8-cineole, citronellal, citronellol, n-decyldehyde, β elemene, elemol, farnesal, farnesol, fenchone, α,β-dihydropseudoionone, dipentene, furfural, iso-pulegol, iso-valeraldehyde, limonene, linalyl acetate, menthol, menthone, methyl heptenol, ocimene, α- oxobisabolene, α-pinene, β-pinene, β-phellandrene, terpineol, terpinolene, neral, nerolic acid, 2 undecanone and geranic acid [11].

 

MATERIALS AND METHODS:

Plant sample:

Cympobogon citratus leaves were collected from house garden in Baghdad Province Al-Yarmuk area in the absence of rain, and then dried under shade so the plant compounds won't be affected by sun heat and later grinded to be used in the extraction methods.

 

Extraction Method:

Extraction by Soxhlet apparatus:

Soxhlet apparatus was used to extract 25 gram. of dried leaves of Cympobogon citratus in 250 ml of solvent (Methanol) for several hours.

 

Clevenger apparatus method (Hydrodistillation):

The Clevenger apparatus was called from its inventor, Joseph Franklin Clevenger, who printed in 1928. A few models be present. The most public one is a part of specific glassware, as can be observed above the rotund bottom flask. The flask, of inconstant size, contains water which is heated as well as the plant to be extracted. The steam increases in the association to a condenser and the condensate drops into the small burette on the right (Figure 3). Oil floats on the water, which for its part is progressively regained to the heated flask over the diagonal conduit. After few hours of extraction, the oil volume together in the burette can be straightly measured [12]. The plant sample weight was about 200 gram sunken in 400 ml of distilled water. After 3 hour about 3 ml of oil was obtained.

 

Chemical Tests:

Alkaloids:

2ml of extract was taken distinctly with 5 ml of 1.5%v/v aqueous hydrochloric acid and filtered. The consequential acidic solution tested with Mayer's Wagner and Dragendorff's precipitate was observed on addition of reagents, indicate the presence of alkaloids. Development of orange sedimentation on addition of Dragendroff's reagent is the positive examination for alkaloid [13].

 

Flavanoids:

2ml of alcoholic extract with few drops of KOH give yellow color and then added few drops of  H₂SO₄ give colorless.

 

Tannin:

0.5ml of alcoholic extracts were diluted with 1ml of water and 2-3 drops of dilute ferric chloride solution was added .Development of a black/green color indicate the presence of tannin.

 

Phenols:

A few drops of acidified ferric chloride 5% solution were additional to the extract. The existence of blue, green or brown coloration indicates the presence of phenolic compound in the sample.

 

Cardiac glycosides:

1ml of glacial acetic acid was added to 2ml of methanol extract in test tube. In this combination few ml of ferric chloride followed by 2 drops of concentrated H2SO4 were added. Green blue color indicates the existence of cardiac glycosides (Killer-Killiani test)

Saponins:

2ml of the alcoholic extract was shaken strongly for 10 seconds and permitted to stand. The formation of determined honeycomb like froth is the positive examination for the presence of saponin.

 

RESULTS AND DISCUSSION:

Chemical Investigation Results:

Table 3.1shows the results for chemical tests

Class of Compound	Result
Alkaloids	+
Flavonoids	+
Tannin	+
Phenols	+
Cardiac glycosides	-
Saponin	-

(+ Presence, - absence)

 

Phytochemicals analysis of samples results showed that the extract of lemon grass contains alkaloids, tannins, flavonoids and phenols, except saponin and cardiac glycosides (Table 1). Results of this study was supported by research [6] which states that the methaolic extract of leaves of lemongrass contains alkaloids, tannins, flavonoids and phenols. The study also showed that the methanolic extract of the leaves of lemongrass does not contain saponin and cardiac glycosides which was contradictory to the results of  another study [14].

 

 

 

Volatile oil obtained Clevenger and Gc-mass Results:

Distillation is the primary technique for extracting the essential oil from plants for commercial produces. The benefit over traditional extraction procedures including solvent is the lack of nonvolatiles in the extract such as chlorophyll and fatty acids. This means that no cleaning procedure is required, which may lead to the damage of product [15].

 

GC-mass analysis:

The sample was analyzed in Ministry of Science and Technology/ Department of Water & Ecology/ Ecology research center.

 

Principle of GC-MS: GC/MS-a combination of two altered analytical techniques, Gas Chromatography (GC) and Mass Spectrometry (MS), is used to investigate complex organic and biochemical mixes [16]. The GC-MS instrument consists of two main components. The gas chromatography portion splits altered compounds in the sample into pulses of pure chemicals according to their vaporization [17]. By influx an inert gas (mobile phase), which transports the sample, over a stationary phase stable in the column [16]. Spectra of compounds are together as they leaving a chromatographic column by the mass spectrometer, which recognizes and quantifies the chemicals depended on their mass-to-charge ratio (m/z). These spectra can then be stored on the computer and analyzed [17].

The result of the GC-MS is shown below:

 

 

Figure 3.1 GC-Mass of  Cymbopogon citratus essential oil

 

 

 

 

 

Compound 5 Line 5

 

 

Compound 1 Line 7

 

As shown from the GC/MS results, we can identify a number of volatile oils in the leaves of our Iraqi plant (C. citratus) which are monoterpenes (C10) carbon skeletons, with a good quantity of oils in these leaves. Citral is a combination of geranial (α-citral) and neral (β-citral) isomeric aldehydes, and it is used for several chemical syntheses [18, 19]

 

Table 3.2 Chemical Composition (%) of cymbopogon citratus essential oil

Peak	R.Time	Area%	Name
5	12.384	15.07	(E)-3(10)-Caren-4-ol.7,7-Dimethyl-4-methylenebicyclo[4.1.0] heptan-3-o
7	12.946	42.27	2,6-Octadienal, 3,7-dimethyl-, (E)- .alpha.-Citral  (E)-Citral. trans-Citral  trans-3,7-Dimethyl-2,6-octadienal

 

Alpha-citral represent the major compound in the oil with 42.27% is and this result is comparable to this study [20] result which identified a number of volatile oil in lemon grass with α-citral being the major compound with 40.8%. The quality of lemongrass oils determines by citral content [18, 19]. It also contains caren-4-ol in a good quantity. There are many altered classes of naturally be founding compounds. Terpenoids also form a group of naturally be founding compounds common of which find in plants, a few of them have also been gotten from other sources. Terpenoids are volatile substances that provide plants and flowers their fragrance. They find widely in the leaves and fruits of higher plants, citrus, conifers and eucalyptus [21]. The term ‘terpene’ was originally employed to designate a mixture of isomeric hydrocarbons of the molecular formula C₁₀H₁₆ finding in the essential oils attained from sap and tissue of plants, and trees. But there is an affinity to use more common term ‘terpenoids’ which contain hydrocarbons and their oxygenated derivatives. Nevertheless, the term terpene is being used these days by some authors to characterize terpenoids [22, 23]. Isoprene rule thermal breakdown of terpenoids offer isoprene asone of the product. Otto Wallach mentioned that terpenoids can be synthesize of isoprene unit [24]. Isoprene rule found that the terpenoid molecules are built from two or more isoprene unit. Additional Ingold  proposed that isoprene units are linked in the terpenoid via ‘head to tail’fashion. Distinctive isoprene rule states that the terpenoid molecule is synthesized of two or more isoprene units linked in a ‘head to tail’fashion [25].

 

 

Figure 3.2 Isoprene unit

 

Citral, and other monoterpenes from lemongrass, exhibits in-vivo anti-inflammatory using carrageenan prompted paw edema and peritonitis in model rat. Paw edema was supposedly reduced by application of citral (100 and 200 mg/kg body weight) and peritonitis was also reduced as leukocyte transformation to peritoneal cavity was mitigated [26].

 

CONCLUSION:

Cymbopogon citratus encloses many phytoconstituents such as flavonoids, alkaloid, tannin and phenolic compounds, terpenoids and which may be accountable for the altered biological activities of this plant. Therefore, we can separate some pure phytopharmaceuticals, which in turn can be used as lead molecules for producing the novel agents having good therapeutic action. The essential oil of C. citratus is mostly composed of monoterpenes hydrocarbons, of which, citral is the chief component. GC-MS investigates show that the major components were citral (42.27%), caren (15.07%). With respect to the development of quality herbal medicine adjustment of the extracts, phytopharmacology of altered extracts, isolation and representation of active phytopharmaceuticals, illumination of the mechanism of action of the isolated compounds and clinical features of the compounds are much required. In the altering global scenario, the interest concerning plants with medicinal value is aggregate substantially in the primary healthcare system both in the developed and in the developing countries. Thus, the data will help scientists and researchers to investigate the compounds responsible for different bioactivities and to illuminate the molecular mechanism of action.

 

ACKNOWLEDGMENT:

I would like to be grateful Mustansiriyah University (www.uomustansiriyah.edu.iq) Baghdad – Iraq of its support in the current work.

 

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Accepted on 03.11.2018        © RJPT All right reserved

Research J. Pharm. and Tech 2019; 12(1): 67-73.