ABSTRACTIn this study, the bioactive compounds of Nardostachys jatamansi have been evaluated using gas chromatography-mass spectrometry (GC-MS). Thechemical composition of the rhizome of N. jatamansi, petroleum ether, chloroform, and ethanol extract was investigated using agilent 7890 GC-MSinstrument. N. jatamansi extract contains 61 compounds, i.e., actinidine (11.2%), indane (28.7%), aristolene (7.2%), gurjuenen (5.5%), valencene(8.9%), globulol (8.2%), betapatachoulene (8.4%), etc. Actinidine and indane are the major compounds in the ethanol extract along with few minorcompounds.Keywords: Chloroform extract, Gas chromatography-mass spectrometry analysis, Nardostachys jatamansi

ME, VEENA

N Achur, Rajeshwara

P, NIRANJANA

English

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Vol 9, Issue 3, 2016Online - 2455-3891 Print - 0974-2441GAS CHROMATOGRAPHY-MASS SPECTROMETRY ANALYSIS OF BIOACTIVE COMPONENTS FROM THE RHIZOME EXTRACT OF NARDOSTACHYS JATAMANSI DCVEENA ME, NIRANJANA P*, RAJESHWARA N ACHURDepartment of Biochemistry, Kuvempu University, Jnana Sahyadri, Shankaraghatta - 577 451, Karnataka, India. Email: bpniru@gmail.comReceived: 18 December 2015, Revised and Accepted: 02 January 2016ABSTRACTIn this study, the bioactive compounds of Nardostachys jatamansi have been evaluated using gas chromatography-mass spectrometry (GC-MS). The chemical composition of the rhizome of N. jatamansi, petroleum ether, chloroform, and ethanol extract was investigated using agilent 7890 GC-MS instrument. N. jatamansi extract contains 61 compounds, i.e., actinidine (11.2%), indane (28.7%), aristolene (7.2%), gurjuenen (5.5%), valencene (8.9%), globulol (8.2%), betapatachoulene (8.4%), etc. Actinidine and indane are the major compounds in the ethanol extract along with few minor compounds.Keywords: Chloroform extract, Gas chromatography-mass spectrometry analysis, Nardostachys jatamansi.AQ1AQ2INTRODUCTIONNardostachys jatamansi DC is an important plant of the family Valerianaceae found growing from Himalayan regions to southwest China and has been used as crude drugs in traditional medicines. It is commonly known as Indian spikenard. It is an erect perennial herb, 10-60 m height with long stout, woody rootstock found in the alpine the Himalayas from Punjab to Sikkim and Bhutan at an altitude of 3000-5000 m. The bitter tonic obtained from the rhizomes of the Jatamansi, also called Bhutajata, Jatahilaa, Thapaswini can be used as a neuroprotective, sunscreen, stimulant, antispasmodic, repellent, antipyretic, antioxidant, as well as to treat herpes infection, leprosy, various neuropsychiatric illnesses, and excessive thirst. The plants are known to be rich in sesquiterpenes [14], which have been found to exhibit antimalarial, antinociceptive, and cytotoxic activities, as well as to enhance the action of nerve growth factor [15-17]. It also has other ayurvedic applications such as in complexion, kidney stones, jaundice, removes blood impurities, spasmodic hysteria and other nervous convulsive ailments, heart palpitations, nervous headache, flatulence, epilepsy, convulsions, respiratory and digestive diseases, skin diseases, typhoid, gastric disorders.The plant is reported for hypolipidemic effects in triton-induced hyperlipidemic rats, for inhibitory activity of methanolic and water extracts against acetyl cholinesterase, anticonvulsant, and neurotoxicity profile in rats, as well as protective effect in rat cerebral ischemia [4-6]. Comparative study of the volatile compounds in the N. jatamansi DC and N. chinesis was done from the commercially available oil. However, the detailed study of phytochemicals in different extracts of N. jatamansi was not done. Hence, the objective of the present study is to identify the phytochemical constituents with the gas chromatography-mass spectrometry (GC-MS) technique.METHODSPlant material and extractionRhizomes of N. jatamansi were purchased from local shops of Udupi, Karnataka, India and authenticated. The dried rhizomes were pulverized to powder in a mechanical grinder. The dried powder was weighed and subjected to Soxhlet extraction procedure in the increasing order of their polarity of solvents, i.e. petroleum ether, chloroform, and ethanol. The extract was concentrated under reduced pressure using rotary evaporator. The final residue thus obtained was then subjected to GC-MS analysis.GC-MS analysisGC combined with mass spectroscopy is a preferable methodology for routine analysis of compounds. 1 Ul of petroleum ether extract, chloroform extract, and ethanol extract was injected into the GC-MS instrument.GC analyzes were carried out on an Agilent 7890 instrument equipped with a hydrogen flame ionization detector and HP-5 capillary column (30 m × 0.32 mm × 0.25 μm, J and W Scientific). Nitrogen was used as carrier gas at a flow rate of 1 mL/minute. Initially, the column temperature was maintained at 60°C for 2 minutes, followed by a temperature gradient from 60°C to 120°C at 2°C/minutes and held constant for 5 minutes at 120°C, then raised to a temperature of 150°C at 1°C/minutes and finally to a temperature of 200°C with a 5°C/minutes rise and maintained for 10 minutes at 200°C. The injector and detector temperatures were maintained at 250°C and operated in split mode (split ratio 1:10). GC-MS was performed on an Agilent 5975C mass selective detector interfaced with an Agilent 7890A gas chromatograph. GC-MS analyzes were performed under similar conditions using an HP-5-MS capillary column (30 m × 0.32 mm × 0.25 μm, J and W Scientific).Thin-layer chromatography was performed on silica gel G-coated plates (0.25 mm for analytical) developed with hexane (system I), hexane–ethyl acetate (98.5:1.5; system II), dichloromethane (system III), and dichloromethane–methanol (98.5:1.5; system IV). Compounds were visualized by spraying with a solution of 3.0% anisaldehyde, 2.8% H2SO4, and 2% acetic acid in ethanol followed by heating for 1-2 minutes.Identification of compoundsInterpretation of the mass spectrum GC-MS was conducted using the database of National Chemical Laboratory (NCL), Pune. The spectrums of the unknown compounds were compared with the spectrum of known compounds stored in the NCL library.RESULT AND DISCUSSIONGC-MS analysis of the phytochemicals present in different extracts (pet ether, chloroform, and ethanol) of N. jatamansi showed the presence of 61 compounds. The active principles with their retention time (RT) probability, R-match, and relative percentages and compound name are presented in the Tables 1-3. The GC-MS chromatogram of the petroleum Research Article116 Veena et al. Asian J Pharm Clin Res, Vol 9, Issue 3, 2016, 115-118Table 1: Compounds detected in the petroleum ether extract of N. jatamansiSerial number Rt Probability R-match Percent Compound1 7.359 39.7 835 0.22 3-Methylbutanoic acid2 10.899 15.3 920 0.054 (-)-β-Pinene3 13.634 11.9 801 0.016 1,8-Cineol4 26.569 3.07 837 0.11 3-Methyl-3-hexen-2-ol5 26.962 9.41 765 0.024 Cyclopentaneacetaldehyde, 2-formyl-3-methyl-α-methylene-6 27.078 2.71 884 0.018 Bicyclo|5.1.0|octane, 8-(1-methylethylidene)-7 28.097 3.87 800 1.18 Beta-pinenoxide8 28.594 40.4 870 0.22 Cyclopentaneacetaldehyde, 2-formyl-3-methyl-α-methylene-9 29.316 8.86 796 0.113 D-Verbenone10 29.628 9.58 879 0.08 Carveol11 30.072 5.2 769 0.049 7’-Oxaspiro|cyclopropane-1,4’-tricyclo|3.3.1.0 (6,8)|nonan-2’-one|12 30.75 18.5 812 0.017 (+)-β-Guaiene13 30.888 18.5 866 0.037 (R)-(+)-Limonene14 32.345 10.3 854 0.221 τ-Gurjunene15 33.018 17.9 914 0.18 (-)-β-Elemene16 33.636 9.52 851 0.099 (-)-α-Gurjunene17 33.805 6.9 918 1.672 β-Maaliene18 34.191 12.3 934 1.163 (+)-9-Aristolene19 34.664 202 32.8 0.178 β-Vatirenene20 34.916 26.7 953 7.917 (+)-1[10]-Aristolene or (+)-Calarene21 35.016 11.6 884 0.176 Eudesma-3,7[11]-diene22 35.257 53.6 972 2.691 Seychellene23 35.35 20.8 928 0.314 α-Guaiene24 35.586 9.25 920 5.466 (-)-α-Gurjunene25 35.954 39.6 916 2.569 α-Patchoulene26 36.292 15.1 950 0.444 (+)-Aromadendrene27 36.419 22.3 928 3.961 (-)-α-Gurjunene28 36.565 12.1 743 0.478 5β,7βH,10α-Eudesm-11-en-1α-ol29 37.527 6.43 912 1 τ-Gurjunene30 37.705 14.6 843 0.636 (E)-β-Ionone31 37.981 26.3 953 8.887 (+)-Valencene32 38.246 5.71 873 0.469 Seychellene33 38.566 25.7 819 2.05 8-Camphenemethanol34 39.06 28.5 910 3.677 Eudesma-3,7[11]-diene35 39.448 36.3 920 0.272 (-)-β-Cadinene36 39.572 4.78 843 0.479 Globulol37 40.438 5.42 856 0.548 β-Vatirenene38 41.15 3.34 785 0.611 Thujopsene39 41.77 21.6 835 0.671 2-Butenal, 2-methyl-4-[2,6,6-trimethyl-cyclohexen-1-yl]-40 42.034 8.58 900 4.367 Seychellene41 42.284 9.25 865 1.433 Isoledene42 42.727 7.4 859 2.156 Cubenol43 43.039 3.51 876 1.259 (-)-Alloaromadendrene44 44.309 6.93 863 6.067 (+)-Aromadendrene45 45.388 4.16 892 8.402 β-Patchoulene46 46.121 74.7 946 4.679 (+)-Valeranone or 1[2H]-Naphthalenone, octahydro-4a, 8a-dimethyl-7-[1-methylethyl]-,|4aR-[4aα,7β,8aα]|-47 48.202 15.2 947 2.014 Valerenal48 50.27 18.6 865 0.773 2,4a-Methanonaphthale-7[4aH]-one, 1,2,3,4,5,6-hexahydro-1,1,5,5-tetramethyl-,[2s-cis]-49 50.538 10.1 840 8.197 Globulol50 50.759 35.1 782 0.934 Methylhinokiate51 51.231 39.1 747 1.898 Butanal, 3-hydroxy-2-methyl-4-|4-t-butyl|-52 51.617 12 955 1.233 9-Acetyl-2,6-dimethyl-10-hydroxybicyclo|4.4.0|deca-1,4-dien-3-one53 52.546 3.8 826 2.755 6-Methyl-5-[1-methylethylidene]-3,6,9-decatrien-2-one54 53.603 25.7 823 0.493 6-[1-Hydroxymethyl-vinyl]-4,8a-dimethyl-3,5,6,7,8,8a- hexahydro-1H-naphthalen-2-one55 53.981 5.35 828 0.275 2[1H] Naphthalenone, 3,5,6,7,8,8a-hexahydro-4,8a-dimethyl-6-[1-methylethenyl]-56 54.151 7.21 832 0.221 Phenol, 2-methoxy-4-[1-propenyl]-57 55.241 5.87 868 0.224 Verrucarol58 59.412 21.3 722 0.982 3-[6,6-Dimethyl-5-oxohept-2-enyl]-cycloheptanone59 60.902 22.1 861 0.73 4,4-Dimethyl-3-[3-methylbut-3-enylidine]-2- methylenebicyclo|4.1.0|heptane60 61.194 21.7 873 1.133 Dehydroaromadendrene61 67.263 4.91 822 0.807 1,4-Butanediol, 1-phenyl-RT: Retention time, N. jatamansi: Nardostachys jatamansi117 Veena et al. Asian J Pharm Clin Res, Vol 9, Issue 3, 2016, 115-118Table 2: Compounds detected in the chloroform extract of N. jatamansiSerial number Rt Probability R-match Percent Compound1 8.287 98.5 978 16.589 1,1,2,2-Tetrachloroethane2 10.871 85.5 866 0.262 Pentachloroethane3 13.513 20.1 760 0.868 1,1,2-Trichloro-2-methylpropane4 13.863 62.1 767 0.27 2,3-Dichloro-2-methylbutane5 15.655 98.9 979 3.413 1,1,1,2,2,2-Hexachloroethane6 17.194 1.45 919 0.37 4-Chloro-2-[chloromethyl]-1-butene7 18.888 96.4 927 2.859 1,1,2,3,3-Pentachloropropane8 25.009 57.6 679 2.855 1,1,2,2,3,3-Hexachloropropane9 29.528 33.1 861 2.22 (S)-(-)-Actinidine10 30.066 5.25 780 2.715 1-Isopropenyl-3-propenyl-cyclopentane11 32.28 11.3 903 1.166 β-Patchoulene12 32.616 27.5 784 0.297 4-Iodo-2-oxaadamantan-6-one13 32.828 7.18 774 0.143 Bicyclo|4.1.0|heptane, 7-bicyclo|4.1.0|hept-7-ylidene14 34.161 17.1 818 0.478 β-Guaiene15 34.594 3.48 853 0.532 (-)-Alloaromadendrene16 34.689 6.91 813 1.037 Cyclopentaneacetaldehyde, 2-formyl-3-methyl-α-methylene-17 34.853 7.5 775 3.824 8-Camphenemethanol18 35.009 28.7 875 1.171 β-Vatirenene19 35.187 31.6 943 1.973 Seychellene20 35.903 5.47 854 2.272 Viridiflorol21 36.358 5.48 870 2.233 Cyperene22 37.093 2.63 804 0.267 α-Selinene23 37.195 11.5 793 0.167 (+)-β-Cedrene24 37.427 7.38 889 0.86 τ-Gurjunene25 37.494 8.22 856 0.918 (+)-Aromadendrene26 37.649 21.8 922 0.867 Seychellene27 37.846 13.7 842 4.78 (±)-Cadinene28 38.115 60.9 731 2.04 Dehydrocyclolongifolene oxide29 39.436 37.6 962 1.404 15-Methyltricyclo|6.5.2[13,14].0[7.15]|pentadeca-1,3,5,7,9,11,13-heptene30 39.923 33.8 806 0.396 4,5,9,10-Dehydroisolongifolene31 40.158 4.1 703 0.82 (+)-Longifolene32 40.604 11 786 0.587 Nerolidol-epoxyacetate33 41.736 14.1 806 0.459 2-Butenal, 2-methyl-4-[2,6,6-trimethyl-1-cyclohexen-1-yl]-34 42.699 8.77 760 4.431 Caryophyllene oxide35 42.86 12.6 813 0.828 1-Nonadecene 36 43.005 18.4 731 0.863 Bicyclo|4.2.0|oct-1-ene, 2-methyl-7-endo-phenyl37 44.215 14.2 834 4.095 τ-Himachalene38 44.356 5.71 866 3.282 Isoledene39 45.293 3.64 876 1.421 Seychellene40 45.415 12.9 855 0.716 β-Vatirenene41 46.028 42.9 923 3.385 (+)-Latamansone or (+)-Valeranone42 46.629 2.96 912 2.726 (+)-Valencene43 48.039 6.08 909 2.674 β-Patchoulene44 48.655 3.25 900 3.763 9-Acetyl-2,6-dimethyl-10-hydroxybicylco|4.4.0|deca-1,4-dien-3-one45 48.973 8.03 817 0.839 Murolan-3,9[11]-diene-10-peroxy46 50.111 8.17 792 0.849 2,2,7,7-Tetramethyltricyclo|6.2.1.0 (1,6)|undec-4-en-3-one47 50.71 4.51 860 0.475 Verrucarol48 51.555 19 774 1.535 1-Chloro-7-heptadecene49 59.474 2.28 869 1.212 1-Eicosene50 70.653 14.9 824 5.795 4-AllylphenolRT: Retention time, N. jatamansi: Nardostachys jatamansiTable 3: Compounds detected in the ethanol extract of N. jatamansiSerial number Rt Probability R-match Percent Compound1 10.271 44 907 3.737 n-Propylbenzene2 10.622 26.3 937 27.892 m-Ethyltoluene3 10.927 15.8 940 2.08 1,3,5-Trimethylbenzene4 11.399 19.2 927 8.213 p-Ethyltoluene5 12.032 22.4 946 9.219 1,3,5-Trimethylbenzene6 13.371 16.9 879 1.467 1,2,4-Trimethylbenzene7 13.962 17.6 928 28.787 Indane8 29.426 14.1 938 11.729 (±)-Actinidine9 46.023 4.19 850 1.957 (+)-Valeranone10 59.532 73.7 816 1.897 Ethylpalmitate11 65.522 23.4 784 1.092 Ethyllinoleate12 65.749 11.1 795 1.929 EthyloleateRT: Retention time, N. jatamansi: Nardostachys jatamansi118 Veena et al. Asian J Pharm Clin Res, Vol 9, Issue 3, 2016, 115-118ether extract, chloroform extract, ethanol extract is shown in the Figs. 1-3 respectively.About 61 compounds are found in the petroleum ether extract. The results revealed that valencene (8.8%), globulol (8.2%), beta-patacholene (8.4%), aristolene (7.9%), and alpha-gurjunene (5.4%) were found as major components in petroleum ether extract. A total of 50 compounds found in chloroform extract in which tetrachloroethane (16.6%), allylphenol (5.8%), and valeranone (3.4%) are the major compounds. 11 compounds are reported from ethanol extract out of which m-ethyl toluene (27.9%), indane (28.8%), and actinidine (11.8%) are major compounds.The bioactive compounds of N. jatamansi have been evaluated using GC-MS. The chemical composition of the N. jatamansi was investigated using aglient 7890 GC-MS instrument. It has been reported that secondary metabolites exert a wide range of biological activities on physiological systems. Among the identified chemicals globulol, gurjunene, and aristolene have a role in antioxidants and antimicrobial activities. Similarly, the presence of alkaloids such as indane and actinidine in the ethanol extract of N. jatamansi are reported to have a role in the antidiabetic and anticancer activity. In traditional medicine, the rhizome of N. Jatamansi used to treat epilepsy, hysteria, coryza, convulsions, internal colic, palpitations of the heart, and mental disorders. Thus, this type of GC-MS analysis is the fruitful step toward understanding active principles in medicinal plants, and this type of study will be helpful for further detailed study and isolation of active components present in the plant extract.CONCLUSIONThe quality of herbal medicine can be evaluated by the construction of chromatographic fingerprints. In the present study, 60 compounds have been identified from the rhizome of N. jatamansi DC by GC-MS analysis. There for GC-MS method is a direct and fast analytical approach for identification of terpenoids and steroids and only a few grams of plant material is required [18]. The presence of various bioactive compounds with different biological activity such has anti-inflammatory, antipyretics, antibacterial, antifungal, skin conditioning have been identified, justifies the use of the whole plant for various ailments by traditional practitioners.ACKNOWLEDGMENTWe would like to thank Dr. Thulasiram Senior Scientist National Chemical Laboratory, Pune, for providing all the facilities to carry out the work.REFERENCES1. Gowda KM, Krishna AP, Kumari NS, Naveen PG, Sharada V. Ethanolic extract of Nardostachys jatamansi potentiates haemapoietic system in albino rats. Nitte Univ J Health Sci 2013;3(1):25-9.2. Rekha K, Rao RR, Pandey R, Prasad KR, Babu KS, Vangala JR, et al. Two new sesquiterpenoids from the rhizomes of Nardostachys jatamansi. J Asian Nat Prod Res 2013;15(2):111-6.3. 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Itokawa H, Masuyama K, Morita H, Takeya K. Cytotoxic sesquiterpenes from nardostachys chinensis. Chem. Pharm Bull (Tokyo) 1993;41(6):1183-4.17. Li P, Matsunaga K, Yamakuni T, Ohizumi Y. Nardosinone, the first enhancer of neurite outgrowth-promoting activity of staurosporine and dibutyryl cyclic AMP in PC12D cells. Brain Res Dev Brain Res 2003;145(2):177-83.18. Liang YZ, Xie P, Chan K. Quality control of herbal medicines. J Chromatogr B Analyt Technol Biomed Life Sci 2004;812(1-2):53-70.AQ3Author Queries???AQ1: Kindly check the city name.AQ2: Kindly provide structure abstract subheading as per journal (Objective, Methods, Results and Conclusion)AQ3: Kindly cite references 1-3 and 7-13 in the text part and also we found the duplicate references and highlighted (3 and 13). Kindly cite the reference citation in chronological order.Fig. 1: Gas chromatography-mass spectrometry chromatogram of the petroleum ether extract of the Nardostachys jatamansiFig. 2: Gas chromatography-mass spectrometry chromatogram of the chloroform extract of the Nardostachys jatamansiFig. 3: Gas chromatography-mass spectrometry chromatogram of the ethanol extract of the Nardostachys jatamansi

GAS CHROMATOGRAPHY-MASS SPECTROMETRY ANALYSIS OF BIOACTIVE COMPONENTS FROM THE RHIZOME EXTRACT OF NARDOSTACHYS JATAMANSI DC

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GAS CHROMATOGRAPHY-MASS SPECTROMETRY ANALYSIS OF BIOACTIVE COMPONENTS FROM THE RHIZOME EXTRACT OF NARDOSTACHYS JATAMANSI DC

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