GC-MS ANALYSIS OF LEAF EXTRACT OF GYNOCHTHODES UMBELLATA (L.) RAZAFIM. AND B. BREMER (RUBIACEAE)

GC-MS ANALYSIS OF LEAF EXTRACT OF GYNOCHTHODES UMBELLATA (L.) RAZAFIM. AND B. BREMER (RUBIACEAE)

S. Anjusha, A. Gangaprasad * and P. M. Radhamany

Plant Tissue Culture and Molecular Biology Lab, Department of Botany, University of Kerala, Thiruvananthapuram, Kerala - 695581, India

ABSTRACT: The present investigation was carried out to determine the presence of volatile, bioactive, phytocomponents in the leaf ethanolic extract of Gynochthodes umbellata (Syn. Morinda umbellata) belonging to the family Rubiaceae,  using Gas Chromatography- Mass Spectrometry (GC-MS) analysis. Gynochthodes umbellata is an important medicinal plant widely used in traditional system of medicine. Literature survey revealed that till date, no work has been reported on GC-MS analysis of ethanol extract of Gynochthodes umbellata leaves. The mass spectrum of the compounds found in the extract was matched with the National Institute of Standards and Technology (NIST) library. GC-MS analysis revealed the presence of 32 compounds. Sterols, stanols and Terpenoids were the most prominent compounds detected in leaf ethanolic extract of this plant. The major compounds detected were 9,19-Cyclolanost-24-en-3-ol (16.09%), 2,5,5-Triphenyl-4-methoxyimidazole (10.87%), β-Sitosterol (8.81%), Stigmast-4-en-3-one (7.26%), Phytol (5.62%). The compounds were identified by comparing their retention time and peak area with that of literature and by interpretation of mass spectra

Gynochthodes umbellata,

Morinda umbellata, Ethanolic extract, GC-MS, Rubiaceae

INTRODUCTION: Since ancient times, people have been exploring the nature particularly plants in search of new drugs which resulted in the use of large number of medicinal plants to treat various diseases. The medicinal plants appear to be rich in secondary metabolites, widely used in traditional medicine to combat and cure various ailments ¹. The study of plants continues principally for the discovery of novel secondary metabolites. Many higher plants are major sources of useful secondary metabolites which are used in pharmaceutical, agrochemical, flavour and aroma industries.

These secondary metabolites can be derived from any part of the plant like bark, leaves, flowers, seeds, etc i.e., any part of the plant may contain active components ². The screening of plant extracts is an innovative strategy to find medicinally important compounds, which will help to develop new drugs ³. Gas chromatography (GC) is used to separates mixtures into individual components using a temperature-controlled capillary column. Mass spectrometry (MS) is used to identify the various components from their mass spectra. Gas Chromatography (GC) and Mass Spectroscopy (MS) associated with particular detection techniques has become a sophisticated means for analysis of various compounds ⁴.

Gynochthodes umbellata (Syn: Morinda umbellata) belonging to the family Rubiaceae, a rare medicinal plant was selected for the present study. G. umbellata is a pretty woody climber with bright orange fruits having potent medicinal properties. Traditionally it is used for treating dysentery and diarrhoea. The leaves are used to expel intestinal worms. Literature survey revealed that till date, no work has been reported on GC-MS analysis of ethanol extract of Gynochthodes umbellata leaves. A preliminary phytochemical screening has shown that many phytochemicals are present in the ethanolic extracts of the different parts of the plant⁵.Therefore, it is important to identify and characterize bioactive phytochemical compounds from ethanolic extract of the plant with the help of GC-MS.

MATERIALS AND METHODES:

Collection of plant material:

GC-MS analysis of leaf of G. umbellata was done in the present work. Fresh leaves of G. umbellata were collected from naturally growing plants in Kariyavattom Campus, University of Kerala, Thiruvananthapuram, Kerala. The plant was properly identified with the help of authentic literature and documented with their characteristic features. The voucher specimens are deposited in the herbarium of Department of Botany, University of Kerala, Kariyavattom (KUBH 7000).

Preparation of the plant extract:

The fresh leaves of G. umbellata were collected, washed and shade dried at room temperature. The dried leaves were pulverized to powder in a mechanical grinder and stored in airtight bottles until future use. 10gm of the leaf powder was subjected to Soxhlet extraction using 200ml ethanol. The extracts were concentrated under reduced pressure in a rotary evaporator (Superfit, Rotavap) and stored in the refrigerator until further use. Two microlitres of the extract was employed in GC-MS analysis.

GC – MS analysis:

The analysis of the extract was performed using GC-MS (Model: GC-MS-QP 2010, Shimadzu, Tokyo, Japan) equipped with a VF 5ms fused silica capillary column of 30m length, 0.25mm diameter and 0.25μm film thickness. For GC-MS detection, an electron ionization system with ionization energy of 70eV was used. The carrier gas was Helium (99.99%) used at a constant flow rate of 1.51 ml/min. injector and mass transfer line temperature were set at 200°C and 240°C respectively. The oven temperature was set from 70 to 220°C at 10°C/min, held isothermal for three minutes and finally raised to 300°C at 10°C/min. Two microlitres of the sample was injected in a split mode with a scan range of 40 – 1000 m/z. The total running time of GC-MS was 35 min. The relative percentage of the extract was expressed as percentage with peak area normalization.

Identification of the components:

Interpretation of mass spectrum GC-MS was conducted using data base of National Institute Standard and Technology ⁶ and Wiley Spectra Libraries ⁷. Spectrum of the unknown component was compared with the spectrum of known components stored in the NIST library.

RESULTS AND DISCUSSION:

The phytocomponents present in the leaf ethanolic extract of G. umbellata were identified by GC-MS analysis, GC-MS running time is 35min. The GC-MS chromatogram of ethanolic extract of leaf of G. umbellata is presented in Fig.1. The active compounds in the leaf ethanolic extract of the plant, their retention time (RT), molecular formula, molecular weight and concentration is provided in Table 1.

FIG. 1: GC-MS CHROMATOGRAM OF G. UMBELLATA LEAF ETHANOL EXTRACT

TABLE 1: PHYTOCOMPONENTS IN THE LEAF ETHANOLIC EXTRACT OF G. UMBELLATA

The gas chromatogram shows the relative concentrations of various compounds getting eluted as a function of retention time. The heights of the peaks indicate the relative concentrations of the components present in the plant ⁸. GC-MS analysis revealed the presence of 32 compounds. Sterols, stanols and Terpenoids were the most prominent compounds detected. The major compounds detected were 9,19-Cyclolanost-24-en-3-ol (16.09%), 2,5,5-Triphenyl-4-methoxyimidazole (10.87%), Beta-Sitosterol (8.81%), Stigmast-4-en-3-one (7.26%), Phytol (5.62%), Bicyclo[3.1.1] heptane, 2,6,6-trimethyl-, (1.alpha.,2.beta.,5.alpha.) (4.34%), Acetamide, 2-(1-isoquinolinylthio) -N,N-dimethyl- (4.20%), Cholest-4-en-3-one (4.06%),

Campesterol (3.97%), Humulane-1,6-dien-3-ol (3.84%). 9,19-Cyclolanost-24-en-3-ol or cycloartenol, is an important type of stanol found in plants. Cycloartenol is an important precursor of steroid biosynthesis and is a component of making K-liquid chlorophyll, which is a health beverage preparation, which helps detoxify and reduce toxins in the body. Drink supplements that contain efficacious cycloartenol also increase nutrient intake in the blood to increase oxygen in the blood helping the regeneration of red blood cell ⁹. 2,5,5-Triphenyl-4-methoxyimidazole is an another important compound eluted, which possessing wide range of biological activities.

β-sitosterol is a waxy substance which is white in colour which possessing antimicrobial, anti oxidant, anti inflammatory, antiasthma, anti arthritic and hepato protective activities ¹⁰. β-sitosterol present in plant oil is reported to lower down the cholesterol level in blood ¹¹, this is due to the property of β-sitosterol to inhibit the absorption of cholesterol in the body ¹². Beta-sitosterol is used for heart disease and to reduce high cholesterol.

It is also used for boosting the immune system.  Stigmast-4-en-3-one is an important steroid compound present in plants, which shows antimicrobial, antioxidant and antiarthritic properties ¹³. It is important to note that steroids are found in large number with very high concentration as compared to other chemical constituents. Phytosterols, which cannot be synthesized by human and therefore all plant sterols and stanols in the human body are of diet origin ¹⁴. Another important compound detected was phytol. It is a diterpene, which posses antibacterial and anti-inflammatory activities ^{15, 16}.

This study revealed that this important woody climber have many important phytoconstituents which posses various biological activities. Thus, this type of GC-MS analysis is the first step towards understanding the nature of active principles in this medicinal plant and this type of study will be helpful for further detailed study.

 CONCLUSION: This is the first report on the GC-MS analysis of G. umbellata. From the GC – MS analysis result, it could be concluded that G. umbellata leaf contains various important bioactive compounds and therefore, it is recommended as a plant of phytochemical and pharmaceutical importance. Further studies are needed to isolate active principle of the extract as well as to elucidate their exact mechanism of action in various diseases.

ACKNOWLEDGEMENT: The authors wish to thank Kerala State Council for Science Technology and Environment, Government of Kerala  for funding this research work and express our  thanks to Professor and  Head, Department of Botany, University of Kerala, Kariyavattom, Thiruvananthapuram, Kerala for providing facilities for doing this work.

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    How to cite this article:

    Anjusha S, Gangaprasad A and Radhamany PM: GC-MS Analysis of Leaf Extract of Gynochthodes Umbellata (L.) Razafim. and B. Bremer (Rubiaceae). Int J Pharm Sci Res 2015; 6(11): 4826-29.doi: 10.13040/IJPSR.0975-8232.6(11).4826-29.

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