Chemical Compositions and Physico-chemical Properties of Three Varieties Essential oils of Cymbopogon giganteus Growing to the Spontaneous State in Benin

Introduction

The study of aromatic plants is still of current events considering the fact that the bioactive compounds of their essential oils focus in recent times,and effectively, theattention of researchersin the world ofmodern medicine. In Africaand in Benin inparticular, thevegetationhas a richand diversepanel ofaromaticplants speciespoorly exploredfor the developmentof plant biotechnologyandhigh reachapplicationsincosmetics,pharmacyandfood industries.Cymbopogonis an important genusofthe Poaceae familyand containing120 speciesdistributedin several varieties [1]. The three varieties, Cymbopogongiganteus(Hochst.)Chiov.,Cymbopogonnardus (L.) RendleetCymbopogonschoenanthus (L.) Sprend. Ssp. Proximus (Hochtst. Ex A. Rich.)Maire&Weiler [2], are aromatic botanical species which grow in the savannas of the tropical regions of Africa. The literature indicates thatCymbopogongiganteusis aherbaceous,sustainable,large(2 to 2.5mhigh), growing in tufts [2, 3]. Its leaves,long from 3 to40 cm and wide2 to 2.5cm,arebanded, glaucous, sheathing and having arounded baseand anarrowedpeak[2].AlphaThe edges arerough to the touch. The youngestare covered with awhitish pubescencedescribed by someauthorsas floury[4].Theaqueousdecoctionofstems with leaves ofC.giganteusassociated withthose ofOcimumbasilicumis used to treatsickle cell disease.Usedonlyin the state ofdecoction, C. giganteuscalmquiet the epileptic fits[2]. Its aroma, acharacteristicpeppery flavor,isenjoyedthroughthe foodinBenin countrysides). Cymbopogonnardus, meanwhile,is a perennial herbgrowing in thewild.It growsindensetuftsand sometimes reaches1.5 m in heightwith stemstrimmed, purple color.Itsspikeletsgeminateare more or lessdifferent according totheir shape and their sex, one sessile, the other stalked, inserted on thearticulatedspine.The floral handle of this planthas manybranchesending ingreenishpelletsears. Concerning Cymbopogonschoanenthus, several studies have reported that itis a widespread species, especially in the dry areas oftropical Africa.Bushyperennialreachinga height of90 cm,Cymbopogonschoanenthuscarrieslinear leaves, fragrant andscabrouson margins.Inflorescencesarepaniclesdense andcontracted.The whole plantcrushed,mixed withleaves of Vitexsimplicifoliais used in thetreatment of the madness in the form of aqueous decoctionorally, in West Africa and in Ethiopia [2].Several recent studies havefocused on identifyingof the chemical profileof the essential oilextracted from eachof these plants. The works realized  byNyamadoret al.[5] showed that the leaves ofCymbopogongiganteuscollectedin Togoare mainly composedof limonene(23.0%) followed by p-mentha-2,8-dien-1-ol split between the trans(5,63%) and the cis(14.3%)forms. Kétohet al.identified a chemical composition of the essential oil ofCymbopogonschoanenthusleaves of Togo and the major components of this essential oil (piperitone: 68.0%, δ-2-carene: 16.48%) [6] weresimilars to those(piperitone: 60.0%, δ-2-carene: 15%, elemol: 8.4%)  identified by Ayedounet al. in Benin [7]. In 2003, Nakahara et al. scrutinized the chemical composition of Cymbopogonnardus leaves essential oil from Tsukuba (Japan) [8]. The chromatographical analyzes performed by adding mass spectrometry showed six major oxygenates components: geraniol (35.7%), trans-citral (22.7%), cis-citral (14.2%), geranyl acetate (9.7%), citronellal (5.8%) and citronellol (4.6%) [8]. Vijender and Ali had also brought back from the essential oil of the leaves of New Dehli (India) the citronellal (29.7%), geraniol (24.2 %), ȣ-terpineol (9.2 %), cis-sabinene hydrate (3.8%), (E)-nerolidol (4.8%), β-caryophyllene (2.2%) and germacren-4-ol (1.5%) [9]. All these volatile extracts, by means of the synergic effects of the constituents of their totum, had showed different biological effects [6, 10, 11, 12]. The present workaims tostudy thechemical composition andphysicochemical characteristicsof the volatileextractsofC.giganteus, C. nardusandC.schoenanthusharvestedin several villages ofBenin.

Experimental

Plant Material Andessential Oil Extraction

Cymbopogongiganteus andCymbopogonnardusleaves were collected in several localities in the south of Benin (Gbakpodji: F₁, Kétou: F₂andF₅,Tinou:F₃, Sèto: F₄,Ifangni: F₆ andF₇, Boukoumbé: F₈). The leaves ofCymbopogonschoenanthus were collected inanother region (Boukounbé) in northern of Benin. A voucher specimen of these diversearomatic plantsaredeposited in Abomey-Calavi University National Herbarium. They were keptin the laboratory between18 and 20°Cin the shade during all the extractions period.Essential oils wereextractedby hydrodistillation of the leaves(250g) for 3 to 4 hoursusing aClevenger according to the methoddescribed in britishpharmacopoeia[13].The volatileextractscollectedwere dried overanhydrous sodium sulfateand analyzed byGC/MS.

Physico-Chemical Propertiesof Essential Oils

Physical parametersof the essential oilsextracted fromleaves Cymbopogonspeciesof were determinedusingthe methods described byAFNOR[5, 6]. These parameters arethe density, refractive index, refractive index, rotatory power and acid Index.

Density At 20°C

Thedensity measurewascarried out usinga micro-pycnometeranda precision balance.

Refractive Index At 20 ° C

The refractive index was determined by means of the refractometer CARL ZEISS JENA 234678.

Rotatory Power At 20 ° C

Themeasurement was madebyCarl Zeisspolarimeter128291.

Acid Index Ia

The material used to determine the acid index was constituted by phenolphtalein, neutralized ethanol, potassium hydroxide (0.05N) and a graduated burette. The index acid calculation was done using the following formula

Ia = 5.61xV / m.

V = Volume in mL of the ethanolic solution of potassium hydroxide

m = Mass measured in gram of essential oil charged.

Volatile Components Analysis

GC/FID

The extracts were analysed on a Hewlett-Packard gas chromatograph Model 6890, equiped with a DB5 MS column (30 m x 0.25 mm, 0.25 µm), programming from 50°C (5 min) to 300°C at 5°C/min, 5 min hold. Hydrogen as carrier gas (1.0 mL/min); injection in split mode (1:60); injector and detector temperature: 280 and 300°C respectively. Each extract is diluted in hexane: 1/30.

GC/MS

The extracts compositions were analysed on a Hewlett-Packard gas chromatograph Model 5890, coupled to a  Hewlett-Packad MS model 5871, equipped with a DB5 MS column (30 m x 0.25 mm, 0.25 µm), programming from 50°C (5 min) to 300°C at 5°C/min, 5 min hold. Helium as carrier gas (1.0 mL/min); injection in split mode (1:30); injector and detector temperature, 250 and 280°C respectively. The MS working in electron impact mode at 70 eV; electronmultiplier: 2500 eV; ion source temperature: 180°C; mass spectra data were acquired in the scan mode in m/z range 33-450.

The compounds assayed by GC in the different essential oils were identified by comparing their retention indices with those of reference compounds in the literature and confirmed by GC-MS by comparison of their mass spectra with those of reference substances [14, 15, 16].

Results and Discussion

The tableI shows thevalues​​of the fourphysico-chemical factorsmeasured.

 Table 1: physico-chemical properties of F₂, F₃, F₄, F₆, F₈ essential oilssamples.

	Density	refractive Index  (at20°C)	Rotary power (at20°C)	Ia (mg de KOH/g)
F2	0.943	1.4845	– 44.87	2.562
F3	0.941	1.4865	+ 21.70	5.452
F4	0.948	1.4880	– 62.74	2.441
F6	0.897	1.4759	– 3.70	0.805
F8	0.928	1.4847	+ 37.20	2.026
F2 = Kétou (11-07-07). F3 = Tinou (15-07-07). F4 = Sèto (02-07-06). F6= Ifangni (25-04-07). F8 = Boukoumbé (28-09-06). Ia= Indice d’acide

In the table IIare presentedthe results ofchromatographic analysis ofessential oils extracted fromleaves of Cymbopogongiganteus,CymbopogonnardusandCymbopogonschoenanthus from Benin.

Table 2: yield andchemical compositionof essential oils fromleaves of C.giganteus, C. nardusandC.schoenanthus.

The results ofphysico-chemical properties (density, refractive index, rotary power andacid index) of essential oils ofCymbopogon(F₂, F₃, F₄, F₆, F₈) presented in Table II revealed a major differences in accordence with sampling localities and variety.Theless densefractions ofessential oilsanalyzedis fromtheleaves ofCymbopogonschoenanthus. The values​​of the rotatory power, refractive indexandacidindex aredifferent from a variety ofCymbopogonto another.In the case ofCymbopogonschoenanthus, physical parametersdetermined(density,refractive index,rotary power) are very different from thosefound byOnadjaet al. in thevolatileextractof the same plantin Burkina Faso [17].

The three species ofCymbopogoneach producedvaried quantities ofessential oil(TableII). The results indicate thatCymbopogonnardus(1.61-2.66%) contains moreessential oilfollowed by Cymbopogonschoenanthus(1.20%)andCymbopogongiganteus(0.03-0.17%). The values supplied by the calculation of volatile extract yield of the Cymbopogongiganteusleaves are lower than those obtained by Alitonou [18] from the leaves collected atSavalouand Sèto in Benin [18]. The yield of essential oil of C. giganteus collected at Sèto in 2006 is also lower compared to that obtained by Alitonou[18]. These differences should be due to the factors such as the influence of the harvest place and period, the composition, the harvest period, the vegetative stage of the plant and theclimatical factors inherent to the collectionzone.Different compounds (23 to 32)were identifiedand representing77.4 to 97.3% of theessential oils(TableII).In fact, theyare rich inoxygenated monoterpenes(57.2-78.4%) known for theirbiologicalefficiencies. In these volatileextracts, it was also noteda highoccurrence ofmonoterpenoic compoundsdominated bya high rate ofoxygenated monoterpenes(57.2-75.3%). However, theessential oilfromleaves growingin Setocontains less than60.0%of oxygenated monoterpenes. Sesquiterpenehydrocarbonwas not detectedinplant speciesF₁, F₂, F₃andF₄.Similarly,volatileextractsfrom leavescollected to Gbakpodji, Kétou and Tinoudid not containoxygenated sesquiterpenes. On the over hand, a hardly unimportant percentage (0.2%)ofoxygenated sesquiterpeneswas notedin the sample ofessential oilfromSetowitha relatively high proportion(19.9%)ofhydrocarbonmonoterpenes. However,this low rate ofoxygenated sesquiterpenes(0.2%)would enhance thebiological efficacy ofsampleF₄.The proportions ofmonoterpenehydrocarbonsinF₁, F₂, F₃volatileextracts samplesare respectively8.2%, 13.5% and 11.4%.

Themajor compoundsof the volatileextractsanalyzed, regardless of their origin,were composedof limonene(7.8-19.4%), trans-para-mentha-2,8-dien-1-ol (12.0-17.4%), cis-limonene oxide(19.2%), cis-para-mentha-2,8-dien-1-ol (8.3-8.9%),cis-verbenol(9.6%), trans-verbenol(8.9%), dihydrocarveol(6.2%), cis-mentha-1(7),8-dien-2-ol (18.4%), myrtenol(5.3-11.9%), cis-dihydrocarvone (10.1-17.2%), trans-piperitol(5.4%), trans-carveol(5.1%), cis-carveol(6.4%), (Z)-ocimenone(5.2%), trans-mentha-1 (7),8-dien-2-ol (17.0-19.9%) and carvotanacetone(17.9%). These componentsare mostlyoxygenated compoundsskeletonmenthadienelikethose studied bySahouo[19] andAlitonou[18]as well as thoseextracted from flowers, leaves and stems acclimatedin Cameroon[20].

25 in 38 compounds were identified representing 64.6 to 99.7% of essential oils extracted from the leaves of Cymbopogonnardus collected at Kétou andIfangni (Table II). A high rate of oxygenated monoterpenes (> 75.0%) characterized the essential oils obtained from leaves of Kétou(F₅) and from Ifangni(F₆, F₇). To Ifangni, the rate is 42.4% and the major compounds of this essential oil, remained similar, in percentage, to those noted in F₁ and F₂. The main componentsaregeraniol(29.9- 34.5%), citronellal (27.9- 32.3%), citronellol(10.1 -11.7%)andelemol(6.6- 7.4%). This group compounds is similarto that determinedby Nakaharaet al.in Japan, which contained, except the geraniol(35.7%), citronellal(5.8%)and citronellol(4.6%). The isomers (cis: 14.2%andtrans: 22.7%) of citralandgeranylacetate(9.7%)were also identified[18]. The same observationwas made byBaranauskiéné[21] in the essential oil fromleavesof Cymbopogonnardus collected at Lithuania and also byKoba in Lomé(Togo) [22].Bycons, these majors compoundswere quite differentfrom those identified(a-pinene4.4%, camphene 8.2%, limonene, 11.0% and geraniol: 18.0%)byParanagamaet al.in the volatile extracts of C.nardus leaves of Kelaniya inSriLanka [23].

Moreover,the analysis of results presented in the tableIIindicates that theabundant components identified in the essential oilof Cymbopogonschoanenthuscollected atBoukoumbéarepiperitone(62.9%),δ-2-carene (14.4%),elemol(5.0%). Also, Ayédounet al. have reported in 1997, in Benin, these same monoterpenoiccompounds(piperitone: 60.0%;δ-2-carene: 15.0%;elemol: 8.4%)punctuated withsomedifferences between proportions[7].In Togo,nearby country ofBenin,Kobaet al.were reported twomajor componentsinrelatively higher proportions(piperitone: 68.0%),δ-2-carene: 16.48%) in the essential [10]. On the other hand, the analysis by GC/MS and¹³C NMRof the essential oilfrom thisplant studiedin Tunisiain 2008byKhadriet al.[24]revealedmajor compoundsdifferentsto thoseobtained during thecurrentinvestigation. Indeed,limonene (10.5-27.3%), β-phellandrene(8.2-16.3%),δ-terpinene(4.3-21.2%) and α-terpineol(6.8-11.0%) were the main componentsof this volatile extract[24].

Conclusion

The chemical compositions ofessential oils studiedhavevaried according to theplace of collection ofplant speciesand the varietyof Cymbopogonstudied.The volatile extractsamples of Cymbopogonnardusexploredcontain lessthan 5.0% ofhydrogenatedterpenes. In Cymbopogonschoenanthusessential oil, hydrogenatedsesquiterpenesare poorly represented(2.4%), while no trace ofthemwas observedin Cymbopogongiganteusvolatile extractsinvestigated.In general,essential oilsof the threeCymbopogonvarietiesinvestigatedare stronglydominated bymonoterpenoiccompounds. Theevaluation of physico-chemical factorshad completed the chemical profiles of thesethree varietiesof essential oil. The estimated physico-chemical factors will help in a better characterization of these three varieties of Cymbopogon essential oil.

References

1. Shahi A.K. and Tava A., Essential Oil Composition of three Cymbopogon Species of Indian Thar Desert, J. Essent. Oil Res., 5, 639-643(1993).
2. Adjanohoun E.J., Adjakidjè V., Ahyi M.R.A., AkeAssi L., Akoègninou A., d’Ameida J., Apovo F., Boukef K., Chadaré M., Cusset G., Dramane K., Eymé J., Gassita J.N.,
3. Gbaguidi N., Goudoté E., Guinko S., Houngnon P., Issa Lo, Keita A., Kiniffo H.V., Koné- Bamba D., MusampaNseyya A., Saadou M., Sodogandji Th., de Souza S., Tchabi A., Dossa C.Z., Zohoun Th., Médecine traditionnelle et Pharmacopée, Contribution aux études ethnobotaniques et floristiques en République Populaire du Bénin, ACCT, Paris, (1989).
4. Dougall H.W. etBogdan A.V., Browse plants of Kenya: with special reference to those occurring in south Baringo, The East African Agricultural Journal, 236-245 (1958).
5. Kerharo J., Adam J.G., Pharmacopée sénégalaise traditionnelle, Plantes médicinales et toxiques, Vigot et Frères, 644-645 (1974).
6. Nyamador W.S., Ketoh G.K., Amévoin K., Nuto Y., Koumaglo H.K., Glitho I.A., Variation in the susceptibility of two Callosobruchus species to essential oils, Journal of Stored Products Research, 46, 48–51(2010).
7. Ketoh G.K., Koumaglo H.K., Glitho I.A., Huignard J., Comparative effects of Cymbopogonschoenanthus essential oil and piperitone on Callosobruchusmaculatus development, Fitoterapia, 77, 506–510 (2006).
8. Ayedoun M.A., Sohounhloué D.K., Menut C., Lamaty G., Bessiere J.M., Composition chimique des huiles essentielles de deux espèces de Cymbopogon du Bénin exploitables industriellement, Bulletin africain Bio ressources. Energie. Développement. Environnement, 8 (1997).
9. Nakahara K., Alzoreky N.S., Yoshihashi T., Nguyen H.T.T. and Trakoontivakorn G., Chemical Composition and antifungal Activity of Essential Oil from Cymbopogonnardus (Citronella Grass), JARQ, 37(4), 249-252 (2003).
10. VijenderS. Mahalwal and Mohd.Ali, Volatile constituents of Cymbopogonnardus(Linn.) Rendle, FlavourFragr. J., 18, 73–76 (2003).
11. Koba K., Sanda K., Raymaud C., Mandin D., Millet J., Chaumont J.P., Activité antimicrobienne des huiles essentielles de Cymbopogoncitratus L.(DC) staff; Cymbopogonnardus L. rendle et Cymbopogonschoanenthus L. spreng., Journal de mycologie médicale, 13(4), 175-180 (2003).
12. JavedIqbal Sultan, Inam-Ur-Rahim, Haq Nawaz, Muhammad Yaqoob and IjazJaved, Mineral composition, palatability and digestibility of free rangeland grasses of northern grasslands of Pakistan, Pak. J. Bot., 40(5), 2059-2070 (2008).
13.  GuévaraNonviho, Valentin D. Wotto, Jean-Pierre Noudogbessi, FélicienAvlessi, Martin Akogbeto, Dominique C. K. Sohounhloué, Insecticidal activities of essential oils extracted from three species of poaceae on Anopheles gambiaespp, major vector of malaria, Scientific Study & Research, 11(4), 11–420 (2010).
14. British Pharmacopoeia, 11. P. A. HMSO: London, (1980).
15. Rösch P., PoppJ., KieferW..Raman and SERS Investigations on Lamiaceae.J. Mol. Struct.,121, 480-481 (1999).
16. Adams R.P. Identification of essential oils by ion trap mass spectrometry. Academy Press, Inc, New-York, 1989.British Pharmacopoeia, 11. P. A. HMSO: London, (1980).
17. Swigar A.A., Silverstein R.M., Monoterpènes, Infrared, Mass, NMR Spectra and Kovats Indices, Aldrich Chem. Co. Milwaukee, WI, USA., (1981).
18. OnadjaYentema, OuedraogoAlioune and SamateAbdoulDorosso,Chemical Composition and Physical Characteristics of the Essential Oil of Cymbopogonschoenanthus(L.) Spreng of Burkina Faso, Journal of Applied Sciences, 7, 503-506 (2007).
19. Alitonou G.A., Huiles essentielles extraites de plantes aromatiques acclimatées au Bénin: étude chimique, évaluation biologique et applications potentielles, Thèse de Doctorat en cotutelle, UAC et Univ. Montpellier II, (2006).
20. Sahouo B.G., Tonzibo Z.F., Boti B., Chopard C., Mahy J.P., N’Guessan Y.T., Anti-
21. inflammatory and analgesic activities: chemical constituents of essential oils of Ocimumgratissimum, Eucalyptus citriodora and Cymbopogongiganteus inhibited lipoxygenase L-1 and cyclooxygenase of PGHS, Bull. Chem. Soc. Ethiop., 17(2), 191-197 (2003).
22. Jirovetz L., Buchbauer G., Eller G., Ngassoum M.B., Maponmetsem P. M.,
23. Composition and antimicrobial activity ofCymbopogongiganteus (Hochst.)Chiov. Essential
24. flower, leaf and Stem oils from Cameroon, J. Essent. Oil Res., 19(5), 485-489 (2007).
25. RenataBaranauskiéné, PetrasRimantasVenskutonis, KoenDewettinck, Roland Verhé, Properties of oregano (Origanumvulgare L.), citronella (Cymbopogonnardus G.) and marjoram (Majoranahortensis L.) flavors encapsulated into milk protein-based matrices, Food Research International, 39, 413–425 (2006).
26. Koba K., Sanda K., Reynaud C., Nenonene Y.A., Millet J., Chaumont J.P., Activités
27. antimicrobiennes d’huiles essentielles de trois Cymbopogonsp. africains vis-à-vis de germes
28. pathogènes d’animaux de compagnie, Ann. Méd. Vét., 148, 202-206 (2004).
29. PriyaniParanagama, ThusharaAbeysekera, Lionel Nugaliyadde and KrishanthiAbeywickrama, Effect of the essential oils of Cymbopogoncitratus, C. nardus and Cinnamomumzeylanicum on pest incidence and grain quality of rough rice (paddy) stored in an enclosed seed box, Food, Agriculture & Environment, 1(2), 134-136 (2003).
30. AydaKhadri, SerralheiroM.L.M., NogueiraJ.M.F., NeffatiM., SmitiS., AraujoM.E.M., Antioxidant and antiacetylcholinesterase activities of essential oils from Cymbopogonschoenanthus L. Spreng. Determination of chemical composition by GC–mass spectrometry and 13C NMR, Food Chemistry, 109, 630–637 (2008).

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