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Cymbopogon flexuosus is widely cultivated for indispensable oil which is holding assorted wellness benefits apart from perfumery utilizations. Microwave radiation combined with hydrodistillation was investigated for efficient and rapid extraction of indispensable oil from lemon grass. Output and citral content were dependent on assorted parametric quantities like weight of natural stuff, volume of H2O, rehydration clip, extraction clip and applied power. Based on these parametric quantities, optimized conditions were found at 100 g of natural stuff, 500 milliliter of H2O, 3 H of rehydration clip, 45 min of extraction clip and 850 W applied power. Essential oil was found to incorporate higher per centum of oxygenated compounds, chiefly citral ( & gt ; 72 % ) . Microscopic analysis provided the footing to understand the extraction mechanism. The result of this survey recommends that hydrodistillation utilizing microwave may farther be utilized for extraction of from assorted medicative workss and scale up.

Introduction

Cymbopogon flexuosus ( lemon grass ) , normally known as lemon grass, is a tall, harsh perennial grass incorporating 1-2 % indispensable oil on a dry footing extracted from the foliages ( Husain et al. , 1988, Nikos ) . Citral, mixture of neral and geranial isomers ( Figure 1 ) ( Rauber, Guterres & A ; Schapoval 2005 ) , is the major compound found in the lemon grass oil ( Schaneberg and Khan, 2002 ) and used in synthesis of ionone, vitamin A and beta-carotene [ 2 ] . Several surveies reported that lemon grass oil possess disinfectant ( ) , fungicidal and antibacterial activity against a diverse scope of beings. Besides, the indispensable oil was proved to be superior fungicide compared to synthetic one ( Nikos ) and found to be non-phytotoxic in nature ( Naik ) . It is possible to handle musculus achings, acne, flatulency, jock ‘s pes, inordinate sweat, itchs and oily clamber utilizing lemongrass oil [ 4 ] . Besides, the oil is utile as anxiolytic, ataractic or anticonvulsive agent ( Blanco, Costa, Freire, Santos Jr. & A ; Costa, 2009 ) . Lemongrass oil was found to be utile in suppressing the activity of I?-glucuronidase and some tried homo, works and animate being pathogens ( Saleem et al. , 2003 ) . Apart from assorted medicative applications, indispensable oil is utile for perfumery because of easy solubility with other indispensable oils and intermixing agents [ 3, 4 ] .

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Driven by the possible applications of indispensable oil extracted from lemon grass, oil was extracted utilizing conventional method like solvent extraction ( Sargenti & A ; Lancas, 1997 ) , steam distillment ( Marongiu, Piras, Porcedda & A ; Tuveri, 2006 ; Sargenti & A ; Lancas, 1997 ; Silou, Malanda & A ; Loubaki, 2004 ) , hydrodistillation ( Marongiu, Piras, Porcedda & A ; Tuveri, 2006, Parikh and Desai, 2011 ) and fresh technique like supercritical fluid extraction ( Marongiu, Piras, Porcedda & A ; Tuveri, 2006 ; Sargenti & A ; Lancas, 1997 ) . Though the conventional techniques are good established, they have energy and environmental issues. The elevated temperature for longer extraction clip can do chemical modii¬?cation of the oil constituents and a loss of the most volatile molecules in instance of steam distillment and hydrodistillation. For solvent extraction, it is vey hard to hold solvent free merchandise and this procedure, besides, leads to loss of the most volatile compounds. Although supercritical fluid extraction is known to be a clean engineering giving acceptable outputs and pureness, its major disadvantage lies in its high operating force per unit area and cost. These drawbacks have led to seek for ‘green ‘ extraction technique in order to hold non merely environment friendly but besides energy efficient method.

In recent old ages, microwave extraction is used for extraction of assorted natural merchandises from works matrices because of big decrease in solvent ingestion [ 7 ] and extraction clip [ 7 ] with better efficiencies compared to conventional techniques. Besides, fewer chemical alterations of original works constituents were observed in instance of microwave extraction compared to hydrodistillation [ 19, 20 ] . Essential oil obtained by microwave irradiation has increased antimicrobic every bit good as antioxidant activities compared to oil obtained by hydrodistillation [ 23 ] . Major advantages of microwave extraction prevarication in its ability for volumetric warming [ 7 ] and maximal incursion of electromagnetic energy in biological tissues [ 8 ] . A elaborate reappraisal of microwave extraction for natural merchandises and its comparing with assorted conventional every bit good as novel techniques were given by Desai and Parikh ( 2010 ) . The comparing showed that microwave outputs better extraction in less clip. Besides, fresh techniques supercritical fluid extraction and pressurized fluid extraction involve high operating force per unit area making safety concern and necessitate higher investing compared to the centrist of microwave extraction.

Extraction of indispensable oil from Cymbopogon flexuosus ( lemon grass ) utilizing microwave extraction has non been reported yet. In the present survey, micro-cook irradiation is combined with hydrodistillation to speed up the extraction of indispensable oil from lemon grass. The purpose of this survey is to analyze the feasibleness of microwave extraction as an efficient technique for the extraction of indispensable oil. The parametric quantities impacting the output of indispensable oil ( weight of works stuffs, volume of H2O, rehydration clip and irradiation power ) are studied in order to optimise the experimental conditions.

Materials and Methods

Materials

Leafs of Cymbopogon flexuosus ( lemon grass ) were collected from Navsari Agriculture University, Navsari in the month of May, 2010. The foliages were dried at room temperature under shed for 2 yearss and stored in wet free environment at room temperature.

Methods

Microwave Extraction

Microwave extraction ( ME ) was performed in a pit of 50 cubic decimeters supplied by M/s Falcon Microwave Technology, Mumbai, India. The unit is equipped with two magnetrons, 850 W each. The clip, temperature and power were controlled by Indusoft Software bundle. The rehydrated works stuff was kept in a flask of 2 fifty capacity along with H2O. The captive H2O molecules shacking in the secretory organs of foliages get vaporized because of microwave irradiation which in bend causes ecstasy of the cell walls. During this procedure, indispensable oil gets vaporized and carried along with H2O vapour towards the capacitor, kept above the microwave pit. The condensed indispensable oil and H2O are so decanted and separated. The oil, therefore, collected is dehydrated and stored at 2 oC. The maximal temperature of the system was boiling point of H2O and system was operated at atmospheric force per unit area.

Assorted parametric quantities like weight of natural stuff, volume of H2O, rehydration of natural stuff, size of natural stuff and power may impact the extraction procedure and, therefore, output. In hydrodistillation survey, it was observed that crunching had an inauspicious consequence on the output of the indispensable oil, hence, leaves of size 2.5 centimeter have been used for the survey ( Communicated paper ) .

Analysis by GC-MS

The indispensable oils obtained by microwave extraction were analyzed by gas chromatography-mass spectroscopy ( Clarus 600 GC-MS, Perkin Elmer ) utilizing Rxi-5Sil-MS column holding arylene stabilized equivalent of 5 % diphenyl ( 30 m x 0.25 millimeters x 0.25 Aµm movie thickness ) . The conditions maintained for obtaining GC -MS spectra were: Carrier gas He with a flow rate of 0.7 ml/min ; split 1:50 ; injection volume 0.1 Aµl ; injection temperature 250 oC ; oven temperature advancement from 80 to 200 oC at 5 oC/min and from 200 to 290 oC at 20 oC/min ; the ionisation manner used was electronic impact at 70 electron volt.

Designation of constituents was carried out by comparing of the mass spectral atomization forms with those stored in MS database ( National Institute of Standards and Technology ) .

Scaning Electron Microscope ( SEM )

Surface morphology of specimen was studied utilizing SEM ( ESEM EDAX XL-30, Philips ) . The scrutiny was carried under vacuity ( 0.8 millimeter of mercury ) and speed uping electromotive force of 30 V.

Result and Discussion

Consequence of Weight of Raw Material

Natural stuffs of different weights were studied to optimise the solid burden ( Figure 5.2 ) . For lower solid burden, the output was found to be changeless. However, citral content increased which might be due to either lessening in loss of volatiles or less extraction of higher molecular weight compounds. The lessening in output every bit good as citral content was observed after 100g. With addition in weight of natural stuff, distribution of microwave radiation per atom lessenings at a fixed applied power which in bend consequences in low dielectric warming and, therefore, a decreased consequence of microwave radiation. Besides, the stuff near the surface of vas will hold higher soaking up of microwaves compared the stuff residing in the interior portion of the vas [ 26 ] . This may supply non unvarying warming of the stuff. Hence longer extraction clip may necessitate for accomplishing same degree of extraction for increased weight of natural stuff. Similar consequences wert obtained for extraction of curcuminoids [ 27 ] , piperine and RRP when solid burden was increased.

Figure 5.2: Consequence of weight of natural stuff ( Extraction conditions: Volume of H2O: 500 milliliter ; Rehydration clip: 1 hour ; Power applied: 850 W ; Extraction clip: 45 min )

Consequence of Volume of Water

Volume of H2O was varied from 0 – 1000 milliliter. As shown in Figure 5.3, with addition in volume, lessening in output every bit good as citral was observed after 500 milliliter. Initially, H2O content was less, resulted in instantaneous warming, which may take to loss of volatile compounds or extraction of compounds which are otherwise hard to acquire extracted. Water, holding polar nature, absorbs important part of microwaves which decreases the incursion deepness for microwave radiation. For this ground, higher H2O sum can take to uncomplete extraction of indispensable oil and lesser sum of citral since more sum of heat is required to make the boiling point of the H2O. This was, besides, observed in the extraction of triterpenoid saponins from Ganoderma atrum [ 28 ] .

Figure 5.3: Consequence of volume of H2O ( Extraction conditions: Weight of natural stuff: 100 milliliter ; Rehydration clip: 1 hour ; Power applied: 850 W ; Extraction clip: 45 min )

Consequence of Rehydration Time

Since microwaves are absorbed the most by polar compound, experiments were carried out with H2O soaked leaves for different rehydration clip. The H2O soaked natural stuff was kept for few proceedingss to run out extra H2O. The sum of H2O consumption for 50 g natural stuff at different rehydration clip is given in Table 5.3. Water consumption was about changeless after 3 H.

Table 5.3: Sum of H2O uptake at different rehydration clip

Rehydration clip, H

60

120

180

240

24

Sum of H2O consumption, milliliter

74

103

124

125

128

Figure 5.4 shows the consequence of different rehydration clip on the output of indispensable oil. The output was found to be changeless. However, the extraction clip was reduced greatly from 60 min to 35 min with addition in rehydration clip. Extraction clip was found to be changeless for rehydration clip of 3 H and onwards. The captive H2O molecule gets heated really quickly on exposure to micro-cook radiation which in bend vaporizes and ruptures the cell. With addition in rehydration clip, sum of H2O molecules absorbed besides increased taking to coevals of more vapour, therefore, leaving more force per unit area on the cell wall and, eventually, rupture of cell wall in lesser clip. Citral content was found to be about changeless over full scope of survey except 24 h. The rehydration caused betterment in extraction efficiency of assorted natural merchandises [ 12, 27 ] .

( a: Extraction status: Weight of natural stuff: 100 milliliter ; Volume of H2O: 500 milliliter ; Power applied: 850 W )

Consequence of Extraction Time

The extraction clip was varied to detect the consequence of radiation on indispensable oil output and citral content. As shown in Figure 5.4, the output increased with addition in extraction clip. The rate of extraction was really fast upto 35 min and so it decreased for the remainder of extraction. However, output was found to be changeless after 45 min of extraction bespeaking terminal of extraction. Citral content was found to increase with addition in extraction clip and so a autumn was observed which may be because of dilution consequence i.e. extraction of compounds which require more clip to acquire vaporized. Similar tendency was observed for extraction of triterpenoid saponins and gly acid where no farther extraction was achieved after some clip.

Figure 5.5: Consequence of extraction clip ( Extraction conditions: Weight of natural stuff: 100ml ; Volume of H2O: 500 milliliter ; Rehydration clip: 1 hour ; Power applied: 850 W )

Consequence of Power Applied

With addition in power degree, microwave soaking up by polar compounds becomes really rapid which enhances warming rate and, finally, rate of extraction. The extraction was carried out at 850 W and 1700 W. In both the instances, the output and the citral content were found to be about changeless and the extraction clip reduced from 45 min to 35 min by increasing power from 850 W to 1700 W. However, the decrease in extraction clip was merely 10 min bespeaking more energy ingestion at higher power. In instance of curucminoids and piperine extraction, higher output was achieved with reduced pureness may be due to extraction of other drosss at higher power.

Composition of Essential Oil

The entire ion chromatogram of indispensable oil is given in Figure 6. Compositions of indispensable oil obtained for different experimental status are given in Table 2. Figure 7 shows the fluctuation of hydrocarbons and oxygenated compounds with regard to different experimental conditions. As seen from the tabular array and figure, oxygenated compounds dominated the composing lending above 90 % of entire composing. Oxygenated compounds are found to be odoriferous and responsible for the characteristic olfactory property of indispensable oil ( Ferhat, 2006 ) . Among oxygenated compounds, citral was the major compound nowadays, runing from 73 – 88 % of entire oxygenated compounds. Citral is the mixture of neral and geranial. Percentage part of geranial ( 52 – 60 % ) was found to be higher compared to that of neral ( 40 – 48 % ) in citral.

It is hard to state that a clear tendency demoing the fluctuation of hydrocarbons and oxygenated compounds with alteration in experimental conditions was observed. However, presence of caryophyllene oxide has affected the per centum of citral content by bring oning dilution consequence. At lower sum of natural stuff, lower volume of H2O, higher rehydration clip, higher extraction clip and higher power, per centum of caryophyllene oxide increased may be because of utmost operating conditions.

Surface morphology of foliages

Adaxial side of lemon grass foliages was examined by SEM. The comparing of micrographs of untreated ( Figure ) and microwave treated ( Figure ) leaves shows that the microwave radiation caused physical alterations in the construction of foliages. The cells incorporating indispensable oil were found to be present next to non photosynthetic tissues ( Lewinsohn ) . After microwave extraction, the cells were empty while the non photosynthetic tissues were merely dried. Since the polar compounds have absorbed the microwave radiation expeditiously, instantaneous energy transportation and vaporisation of the compounds occurred which lead to rupture of cell walls. This phenomenon caused the release of volatiles from the cell wall, therefore, emptying the cells. Similar morphological alterations were observed by ( Ferhat and Reila )

Decisions

Microwave extraction was employed to pull out indispensable oil from Cymbopogon flexuosus for the first clip utilizing unfastened vas setup. By analyzing the consequence of assorted parametric quantities on microwave extraction, optimum conditions for extraction of indispensable oil from lemon grass were: weight of natural stuff 100 g, volume of H2O 500 milliliter, rehydration clip 3 H, extraction clip: 45 min and power applied: 850 W. Essential oil was found to be rich with oxygenated compounds ( & gt ; 90 % ) , chiefly citral. The citral content varied from 72 – 80 % with alteration in operating conditions. Surface morphology of lemon grass foliages revealed that microwave radiation caused rupture of cell wall because of localised warming. The method utilized was found to be rapid and efficient and provided green attack for extraction of indispensable oil from lemon grass. The consequences suggest that the combination of microwave with hydrodistillation can farther be utilized for extraction of volatile compounds from works stuffs and graduated table up.

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