Skip to main content
PMC home page
PLOS ONE logoLink to PLOS ONE
. 2013 Jan 10;8(1):e52797. doi: 10.1371/journal.pone.0052797

Chemometric Profile of Root Extracts of Rhodiola imbricata Edgew. with Hyphenated Gas Chromatography Mass Spectrometric Technique

Amol B Tayade 1, Priyanka Dhar 1, Jatinder Kumar 1, Manu Sharma 2, Rajinder S Chauhan 3, Om P Chaurasia 1,*, Ravi B Srivastava 1
Editor: Gianfranco Pintus4
PMCID: PMC3542346  PMID: 23326358

Abstract

Rhodiola imbricata Edgew. (Rose root or Arctic root or Golden root or Shrolo), belonging to the family Crassulaceae, is an important food crop and medicinal plant in the Indian trans-Himalayan cold desert. Chemometric profile of the n-hexane, chloroform, dichloroethane, ethyl acetate, methanol, and 60% ethanol root extracts of R. imbricata were performed by hyphenated gas chromatography mass spectrometry (GC/MS) technique. GC/MS analysis was carried out using Thermo Finnigan PolarisQ Ion Trap GC/MS MS system comprising of an AS2000 liquid autosampler. Interpretation on mass spectrum of GC/MS was done using the NIST/EPA/NIH Mass Spectral Database, with NIST MS search program v.2.0g. Chemometric profile of root extracts revealed the presence of 63 phyto-chemotypes, among them, 1-pentacosanol; stigmast-5-en-3-ol, (3β,24S); 1-teracosanol; 1-henteracontanol; 17-pentatriacontene; 13-tetradecen-1-ol acetate; methyl tri-butyl ammonium chloride; bis(2-ethylhexyl) phthalate; 7,8-dimethylbenzocyclooctene; ethyl linoleate; 3-methoxy-5-methylphenol; hexadecanoic acid; camphor; 1,3-dimethoxybenzene; thujone; 1,3-benzenediol, 5-pentadecyl; benzenemethanol, 3-hydroxy, 5-methoxy; cholest-4-ene-3,6-dione; dodecanoic acid, 3-hydroxy; octadecane, 1-chloro; ethanone, 1-(4-hydroxyphenyl); α-tocopherol; ascaridole; campesterol; 1-dotriacontane; heptadecane, 9-hexyl were found to be present in major amount. Eventually, in the present study we have found phytosterols, terpenoids, fatty acids, fatty acid esters, alkyl halides, phenols, alcohols, ethers, alkanes, and alkenes as the major group of phyto-chemotypes in the different root extracts of R. imbricata. All these compounds identified by GC/MS analysis were further investigated for their biological activities and it was found that they possess a diverse range of positive pharmacological actions. In future, isolation of individual phyto-chemotypes and subjecting them to biological activity will definitely prove fruitful results in designing a novel drug.

Introduction

To identify and evaluate the therapeutic potential of medicinal herbs, isolation of active components and structural elucidation of these compounds is very essential in medicinal chemistry and natural product research. In recent years a lot of attention has been given towards the study of organic compounds from medicinal herbs and to elucidate their pharmacological activities. Numerous extraction techniques and analytical systems like spectrophotometry, capillary electrophoresis, high performance liquid chromatography (HPLC), high performance thin layer chromatography (HPTLC), gas chromatography (GC) with flame ionization detection (FID), gas chromatography/mass spectrometry (GC/MS) have been developed for the analysis and characterization of active compounds from medicinal plants. GC/MS has become an ideal technique for qualitative and quantitative analysis of volatile and semi-volatile compounds of plant origin. It has the unique combination of a perfect separation system (GC) with the excellent identification and confirmation technique (MS) which has made it the best suited analytical system for plant compound characterization. Additionally, for rapid extraction and precise analysis of these active phyto-compounds, the experimental design should also be optimized to obtain enhanced recoveries, low solvent consumption, and reduced extraction time [1]–[5].

Rhodiola imbricata Edgew. (Rose root/Arctic root/Golden root/Shrolo), belonging to the family Crassulaceae, is an important food crop and medicinal plant in the high altitude region of Indian trans-Himalayan cold desert. It is a popular medicinal plant in Pakistan, Nepal, India, Tibet, China, and many other countries and is widely used as food and traditional medicine around the world. A number of metabolites like phenylpropanoids, phenylethanol derivatives, flavanoids, terpenoids, and phenolic acids have been found in good quantity from these Rhodiola species and extracts of these plant species, particularly those from roots, have been shown to possess pharmacological activities. A survey of the literature showed that Rhodiola species influence a number of physiological functions including neurotransmitter levels, central nervous system activity, and cardiovascular function. It is being used to stimulate the nervous system, decrease depression, enhance work performance, eliminate fatigue, and prevents high-altitude sickness. Most of these effects have been ascribed to constituents such as salidrosides (rhodiolosides), rosavins, and p-tyrosol. Many pharmacological studies on R. imbricata have demonstrated that this plant exhibits cardioprotective, anti-inflammatory, antistress, dermal wound healing, and adaptogenic activities. It has also been found to possess antioxidant, antiaging, immuno-stimulant, radioprotective, and anticarcinogenic properties [6]–[24]. All these reports validate its use in traditional system of medicine.

However, the phytochemistry of the most important plant part having the medicinal and therapeutic potential, the root of R. imbricata has not been studied in considerable details. Hence, aim of the present investigation was to identify and quantify the chemotypes extracted successively in different solvents such as n-hexane, chloroform, dichloroethane, ethyl acetate, methanol, and 60% ethanol, from roots of R. imbricata from trans-Himalayan cold desert of Ladakh, India, by hyphenated GC/MS technique.

Materials and Methods

Chemicals

n-Hexane, chloroform, dichloroethane, ethyl acetate, methanol, ethanol, and water CHROMASOLV HPLC grade and all other chemicals used were of analytical grade and purchased from Sigma-Aldrich (St. Louis, MO, USA).

Ethics statement

All necessary permits were obtained for the described field studies. The permit was issued by Dr. B. Balaji (IFS), Divisional Forest Officer, Leh Forest Division, Jammu & Kashmir, India.

Plant materials and extraction

R. imbricata roots were collected from the trans-Himalayan region (Chang-La Top, altitude = 5330 m above mean sea level, Indus valley, Ladakh) of India in the month of October, 2011 after the period of senescence, with the prior permission from the local authorities. The plant roots were washed thoroughly and cut into small pieces and shade dried at room temperature for 15 days. Then they were finely powdered and used for extraction. The root powder (20 gm) was taken for the sequential extraction in six solvent systems with increasing polarity viz. n-hexane, chloroform, dichloroethane, ethyl acetate, methanol, and 60% ethanol by Soxhlet apparatus (Borosil GlassWorks Limited, Worli, Mumbai, India) at 40°C. The extracted fractions were concentrated under vacuum and reduced pressure (BUCHI Rotavapor R-205, BUCHI Labortechnik AG CH-9230, Flawil, Switzerland) at 40°C by circulation of cold water using thermostat maintained at 4°C in order to minimize the degradation of thermolabile compounds. The dry extracts were then stored in a −80°C freezer till further analysis.

Preparation of sample for GC/MS analysis

The 25 mg of concentrated n-hexane, chloroform, dichloroethane, ethyl acetate, methanol, and 60% ethanol root extracts were redissolved in the respective solvents, vortexed properly and filtered through 0.22 µm syringe filter (Millipore Corp., Bedford, MA, USA). One microlitre aliquot of the sample solution was injected into the GC/MS MS system for the requisite analysis.

Instrumentation and chromatographic conditions

GC/MS analysis was carried out on a Thermo Finnigan PolarisQ Ion Trap GC/MS MS system comprising of an AS2000 liquid autosampler (Thermo Finnigan, Thermo Electron Corporation, Austin, TX, USA). The gas chromatograph was interfaced to a mass spectrometer instrument employing the following conditions viz. Durabond DB-5 ms column (30 m×0.25 mm×0.25 µm), operating in electron impact [electron ionisation positive (EI+)] mode at 70 eV, helium (99.999%) was used as carrier gas at a constant flow of 1 ml/min, an injection volume of 0.5 EI was employed (split ratio of 10∶1), injector temperature 280°C, and transfer line temperature 300°C. The oven temperature was programmed from 50°C (isothermal for 2 min), with gradual increase in steps of 10°C/min, to 300°C. Mass spectra were taken at 70 eV, a scan interval of 0.5 s, and full mass scan range from 25 m/z to 1000 m/z. The data acquisition was performed on Finnigan Xcalibur data acquisition and processing software version 2.0 (ThermoQuest, LC and LC/MS Division, San Jose, California, USA).

Identification of components

Interpretation of mass spectrum of GC/MS was done using the NIST/EPA/NIH Mass Spectral Database (NIST11), with NIST MS search program v.2.0g [National Institute Standard and Technology (NIST), Scientific Instrument services, Inc., NJ, USA]. The mass spectrum of the unknown component was compared with the spectrum of the known components stored in the NIST library. The name, molecular weight, and structure of the components of the test materials were ascertained.

Results

GC/MS chromatograms of n-hexane (Fig. 1), chloroform (Fig. 2), dichloroethane (Fig. 3), ethyl acetate (Fig. 4), methanol (Fig. 5), and 60% ethanol (Fig. 6) root extracts of R. imbricate as per the experimental procedure discussed above, showed various peaks indicating the presence of different chemotypes in the respective extracts.

Figure 1. GC/MS chromatogram of n-hexane root extract of R. imbricata.

Figure 1

Open in a new tab

Figure 2. GC/MS chromatogram of chloroform root extract of R. imbricata.

Figure 2

Open in a new tab

Figure 3. GC/MS chromatogram of dichloroethane root extract of R. imbricata.

Figure 3

Open in a new tab

Figure 4. GC/MS chromatogram of ethyl acetate root extract of R. imbricata.

Figure 4

Open in a new tab

Figure 5. GC/MS chromatogram of methanol root extract of R. imbricata.

Figure 5

Open in a new tab

Figure 6. GC/MS chromatogram of 60% ethanol root extract of R. imbricata.

Figure 6

Open in a new tab

GC/MS chemometric profile

n-Hexane root extract

The n-hexane root extract revealed the presence of 22 different chemotypes which were characterized and identified (Table 1, Fig. 1) by comparison of their mass fragmentation patterns with the similar in NIST database library. Of these 22 chemotypes, 1-pentacosanol (28.21%), stigmast-5-en-3-ol, (3β,24S) (13.40%), 1-teracosanol (9.23%), 1-henteracontanol (8.53%), 17-pentatriacontene (7.01%), and 13-tetradecen-1-ol acetate (6.40%) were found to be major constituents whereas 1-hentriacontane (3.66%), 1-heptacosane (3.47%), 1-tericosanol (2.51%), 13-docosan-1-ol, (Z) (2.12%), eicosen-1-ol, cis-9 (1.99%) 1,30-triacontanediol (1.49%), stigmast-4-en-3-one (1.28%), bis(2-ethylhexyl) phthalate (1.20%), hexadecanoic acid (1.16%), 1-tetrateracontane (0.90%), campesterol (0.90%), α-Tocopherol-β-D-mannoside (0.73%), stigmastanol (0.71%), 1-pentatriacontane (0.48%), and 3-methoxy-5-methylphenol (0.46%) were found to be present in trace amount.

Table 1. Phyto-chemotypes identified in the n-hexane root extract of R. imbricata by GC/MS.
S. No. Peak RT (min) Peak area Peak area (%) Compound detected Hit SI RSI Prob CAS No Mol. Formula Mol. Wt.
1 13.66 3489208 0.46 3-Methoxy-5-methylphenol 1 838 874 72.98 3209-13-0 C8H10O2 138
2 22.84 9266063 1.16 Hexadecanoic acid 1 801 842 73.13 57-10-3 C16H32O2 256
3 24.91 34516141 4.16 Ethyl linoleate 1 839 876 17.81 544-35-4 C20H36O2 308
4 26.69 6791237 0.9 1-Tetratetracontane 1 827 829 10.27 7098-22-8 C44H90 618
5 27.74 3769798 0.48 1-Pentatriacontane 1 823 838 11.01 630-07-9 C35H72 492
6 28.77 30375043 3.66 1-Hentriacontane 1 848 855 14.33 630-04-6 C31H64 436
7 29.07 9600422 1.2 Bis(2-ethylhexyl) phthalate 3 855 866 18.65 117-81-7 C24H38O4 390
8 29.82 20564432 2.51 1-Tricosanol 4 807 823 6.18 05-01-3133 C23H48O 340
9 30.08 16467246 1.99 Eicosen-1-ol, cis-9 1 810 847 10.25 112248-30-3 C20H40O 296
10 30.7 56408242 7.01 17-Pentatriacontene 1 799 800 7.41 6971-40-0 C35H70 490
11 31.95 74961756 9.23 1-Tetracosanol 2 804 821 12.41 506-51-4 C24H50O 354
12 33.31 16764245 2.12 13-Docosen-1-ol, (Z) 1 798 820 9.17 629-98-1 C22H44O 324
13 33.17 68435403 8.53 1-Hentetracontanol 1 848 870 37.4 40710-42-7 C41H84O 592
14 35.01 229932016 28.21 1-Pentacosanol 1 812 823 31.85 26040-98-2 C25H52O 368
15 35.5 12157246 1.49 1,30-Triacontanediol 1 769 786 7.65 36645-68-8 C30H62O2 454
16 36.4 29211487 3.47 1-Heptacosane 3 781 825 10.3 593-49-7 C27H56 380
17 36.74 5914760 0.73 α-Tocopherol-β-D-mannoside 1 814 865 57.26 CID 597057 C35H60O7 592
18 37.95 54630744 6.4 13-Tetradecen-1-ol acetate 5 743 801 4.89 56221-91-1 C16H30O2 254
19 38.08 6914671 0.9 Campesterol 1 771 794 53.27 474-62-4 C28H48O 400
20 39.39 107745880 13.4 Stigmast-5-en-3-ol, (3β,24S) 1 848 855 45.49 83-47-6 C29H50O 414
21 39.62 5803953 0.71 Stigmastanol 1 720 732 58.69 19466-47-8 C29H52O 416
22 41.64 10789041 1.28 Stigmast-4-en-3-one 1 699 874 18.43 1058-61-3 C29H48O 412
Open in a new tab

Chloroform root extract

GC/MS chemometric profile of chloroform root extract showed the presence of 18 different chemotypes (Table 2, Fig. 2). Amongst these, stigmast-5-en-3-ol, (3β,24S) (24.30%), methyl tri-butyl ammonium chloride (14.64%), bis(2-ethylhexyl) phthalate (11.50%), 7,8-dimethylbenzocyclooctene (7.97%), ethyl linoleate (4.75%), 3-methoxy-5-methylphenol (4.16%), and hexadecanoic acid (4.13%) were found to constitute major amount while, campesterol (3.94%), 1-pentacosanol (3.82%), 17-pentariacontene (3.38%), benzene sulfonic acid, 4-amino-3-nitro (3.21%), orcinol (2.93%), benzenemethanol, 3-hydroxy, 5-methoxy (2.62%), 1-hentetracontanol (2.54%), 1-tetracosanol (1.86%), stigmast-4-en-3-one (1.82%); and α-tocopherol (1.31%), and eicosen-1-ol, cis-9 (1.13%) were found to be present in trace quantity.

Table 2. Phyto-chemotypes identified in the chloroform root extract of R. imbricata by GC/MS.
S. No. Peak RT (min) Peak area Peak area % Compound detected Hit SI RSI Prob CAS No Mol. Formula Mol. Wt.
1 11.69 9872792 14.64 Methyl tri-butyl ammonium chloride 1 792 797 56.37 56375-79-2 C13H30ClN 235
2 13.75 2364627 4.16 3-Methoxy-5-methylphenol 1 808 858 67.78 3209-13-0 C8H10O2 138
3 15.14 1927265 2.93 1,3-Benzenediol, 5-methyl 1 708 865 22.77 504-15-4 C7H8O2 124
4 18.35 1520584 2.62 Benzenemethanol, 3-hydroxy-5-methoxy 1 811 860 85.91 30891-29-3 C8H10O3 154
5 20.55 5041042 7.97 7,8-Dimethylbenzocyclooctene 1 770 849 30.04 99027-76-6 C14H14 182
6 22.89 2762363 4.13 Hexadecanoic acid 1 749 813 60.75 57-10-3 C16H32O2 256
7 25.01 3042142 4.75 Ethyl linoleate 4 736 854 17.03 544-35-4 C20H36O2 308
8 28.8 2066650 3.21 Benzene sulfonic acid, 4-amino-3-nitro 4 582 652 2.29 616-84-2 C6H6N2O5S 218
9 29.11 7589941 11.5 Bis(2-ethylhexyl) phthalate 4 793 806 9.51 117-81-7 C24H38O4 390
10 30.12 670458 1.13 Eicosen-1-ol, cis-9 1 810 847 10.25 112248-30-3 C20H40O 296
11 30.74 2150315 3.38 17-Pentariacontene 1 799 800 7.41 6971-40-0 C35H70 490
12 31.98 1129772 1.86 1-Tetracosanol 2 804 821 12.41 506-51-4 C24H50O 354
13 33.23 1665168 2.54 1-Hentetracontanol 1 848 870 37.4 40710-42-7 C41H84O 592
14 34.99 2475797 3.82 1-Pentacosanol 8 616 763 0.8 26040-98-2 C25H52O 368
15 36.83 898525 1.31 α-Tocopherol 3 571 702 11.35 59-02-9 C29H50O2 430
16 38.17 2267712 3.94 Campesterol 1 682 779 20.92 474-62-4 C28H48O 400
17 39.51 16343303 24.29 Stigmast-5-en-3-ol, (3β,24S) 1 798 823 43.31 83-47-6 C29H50O 414
18 41.82 1236875 1.82 Stigmast-4-en-3-one 1 540 625 8.51 1058-61-3 C29H48O 412
Open in a new tab

Dichloroethane root extract

GC/MS chemometric profile of dichloroethane root extract illustrated the presence of 25 different chemotypes (Table 3, Fig. 3). Among these, camphor (17.78%), stigmast-5-en-3-ol, (3β,24S) (15.42%), ethyl linoleate (9.95%), 1,3-dimethoxybenzene (8.15%), hexadecanoic acid (6.55%), and thujone (4.73%) were present in major amount, whereas, benzene sulfonic acid, 4-amino-3-nitro (3.96%), campesterol (3.88%), methanol, (4-carboxymethoxy) benzoyl (3.27%), stigmast-4-en-3-one (2.84%), 1-hentetracontanol (2.74%), oleic acid (2.18%), bis(2-ethylhexyl) adipate (2.16%), bacteriochlorophyll-c-stearyl (2.11%), eucalyptol (1.95%), ethanone, 1-(2,6-dihydroxy-4-methoxyphenyl) (1.66%), 1-dotriacontane (1.58%), linalyl isovalerate (1.49%), 3-methoxy-5-methylphenol (1.47%), 1-chloro-2,4-dimethoxybenzene (1.44%), borneol (1.22%), 4-chlorothiophenol (1.02%), phenol, 2,4-bis(1,1-dimethylethyl) (0.94%), fenchyl alcohol (0.91%), and stigmast-3,5-dien-7-one (0.60%) were found to be present in trace.

Table 3. Phyto-chemotypes identified in the dichloroethane root extract of R. imbricata by GC/MS.
S. No. Peak RT (min) Peak area Peak area % Compound detected Hit SI RSI Prob CAS No Mol. Formula Mol. Wt.
1 8.19 1187393 1.95 Eucalyptol 2 800 829 51.1 470-82-6 C10H18O 154
2 9.64 2910856 4.73 Thujone 1 835 845 23.35 546-80-5 C10H16O 152
3 10.44 11188301 0.85 Camphor 1 837 854 25.13 76-22-2 C10H16O 152
4 10.91 742204 1.22 Borneol 1 866 878 28.4 464-45-9 C10H18O 154
5 11.32 552901 0.91 β-fenchyl alcohol 1 765 836 10.05 470-08-6 C10H18O 154
6 13.06 631861 1.02 Benzenethiol, 4-chloro 1 599 670 34.81 106-54-7 C6H5ClS 144
7 13.66 898266 1.47 3-Methoxy-5-methylphenol 1 759 851 50.29 3209-13-0 C8H10O2 138
8 15.88 2126709 3.27 Methanol, (4-carboxymethoxy) benzoyl 1 694 746 16.43 80099-44-1 C10H10O5 210
9 16.5 571000 0.94 Phenol, 2,4-bis(1,1-dimethylethyl) 2 842 865 22.66 96-76-4 C14H22O 206
10 17.36 885243 1.44 1-Chloro-2,4-dimethoxybenzene 1 635 756 24.81 7051-13-0 C8H9ClO2 172
11 17.56 926151 1.49 Linalyl isovalerate 1 751 812 14.87 50649-12-2 C15H26O2 238
12 19.33 953645 1.58 1-Dotriacontane 1 787 803 45.91 544-85-4 C32H66 450
13 20.49 975679 1.66 Ethanone, 1-(2,6-dihydroxy-4-methoxyphenyl) 1 661 835 45.49 7507-89-3 C9H10O4 182
14 22.81 4154976 6.55 Hexadecanoic acid 1 801 855 70.5 57-10-3 C16H32O2 256
15 23.18 1089256 2.18 Oleic acid 1 772 798 49.32 112-80-1 C18H34O2 282
16 24.28 1364685 2.11 Bacteriochlorophyll-c-stearyl 1 755 767 13.97 CID5367801 C52H72MgN4O4 840
17 24.94 6245279 9.95 Ethyl linoleate 1 783 883 10.73 544-35-4 C20H36O2 308
18 27.61 1330635 2.16 Hexanedioic acid, bis(2-ethylhexyl) ester 1 695 773 29.62 103-23-1 C22H42O4 370
19 28.74 2480706 3.96 Benzene sulfonic acid, 4-amino-3-nitro 6 590 655 1.81 616-84-2 C6H6N2O5S 218
20 29.1 5070293 8.15 1,3-Dimethoxybenzene
21 33.12 1747914 2.74 1-Hentetracontanol 4 675 828 4.63 40710-42-7 C41H84O 592
22 38.04 2440601 3.88 Campesterol 1 652 759 13.55 474-62-4 C28H48O 400
23 39.36 9725435 15.42 Stigmast-5-en-3-ol, (3β,24S) 1 814 853 63.99 83-47-6 C29H50O 414
24 40.9 366122 0.6 Stigmast-3,5-dien-7-one 1 512 760 71 2034-72-2 C29H46O 410
25 41.65 1869647 2.84 Stigmast-4-en-3-one 1 628 856 22.86 1058-61-3 C29H48O 412
Open in a new tab

Ethyl acetate root extract

Nineteen different chemotypes were identified in ethyl acetate extract (Table 4, Fig. 4). Amongst these 19 chemotypes, 1,3-dimethoxybenzene (27.61%), 1,3-benzenediol, 5-pentadecyl (16.90%), 3-methoxy-5-methylphenol (10.11%), 1,3-benzenediol, 5-methyl (8.40%), benzenemethanol, 3-hydroxy, 5-methoxy (5.75%), cholest-4-ene-3,6-dione (5.75%), and dodecanoic acid, 3-hydroxy (4.46%) were found to constitute major amount, whereas, 7,8-dimethylbenzocyclooctene (3.57%) 3,5-dimethoxyphenyl acetate (3.44%), α-D-glucopyranoside, O-α-D-glucopyranosyl-(1.fwdarw.3)-β-D-fructofuranosyl (2.95%), stigmast-5-en-3-ol, (3β,24S) (2.12%), eicosen-1-ol, cis-9 (2.12%), hexadecanoic acid (1.84%), oleic acid (1.34%), bacteriochlorophyll-c-stearyl (1.14%), phenol, 2,4-bis(1,1-dimethylethyl) (0.85%), 1-pentatricontene (0.72%), 1-dodecanol, 3,7,11-trimethyl (0.61%), and stigmast-4-en-3-one (0.32%) were found to be present in trace.

Table 4. Phyto-chemotypes identified in the ethyl acetate root extract of R. imbricata by GC/MS.
S. No. Peak RT (min) Peak area Peak area % Compound detected Hit SI RSI Prob CAS No Mol. Formula Mol. Wt.
1 10.11 18961417 4.16 3-Methoxy-5-methylphenol 1 880 881 78.08 3209-13-0 C8H10O2 138
2 14.63 17158116 8.4 1,3-Benzenediol, 5-methyl 1 909 929 70.69 504-15-4 C7H8O2 124
3 16.5 1721777 0.85 Phenol, 2,4-bis(1,1-dimethylethyl) 1 860 887 30.17 96-76-4 C14H22O 206
4 17.78 1213849 0.61 1-Dodecanol, 3,7,11-trimethyl 1 696 721 3.99 6750-34-1 C15H32O 228
5 18.07 11340896 5.75 Benzenemethanol, 3-hydroxy-5-methoxy 1 866 868 75.89 30891-29-3 C8H10O3 154
6 19.54 7034263 3.44 Phenol, 3,5-dimethoxy acetate 7 636 829 4.41 23133-74-6 C10H12O4 196
7 20.4 7323033 3.57 7,8-Dimethylbenzocyclooctene 1 803 870 51.6 99027-76-6 C14H14 182
8 20.6 4129209 2.12 Eicosen-1-ol, cis-9 1 750 771 5.19 629-96-9 C20H40O 296
9 21.22 6328149 2.95 α-D-glucopyranoside, O-α-D-glucopyranosyl-(1.fwdarw.3)-β-D-fructofuranosyl 1 720 759 37.89 597-12-6 C18H32O16 504
10 22.8 3545343 1.84 Hexadecanoic Acid 1 795 837 71.82 57-10-3 C16H32O2 256
11 23.17 2659100 1.34 Oleic acid 1 779 794 11.53 112-80-1 C18H34O2 282
12 24.97 9179686 4.46 Dodecanoic acid, 3-hydroxy 1 674 706 35.46 1883-13-2 C12H24O3 216
13 25.52 2261098 1.14 Bacteriochlorophyll-c-stearyl 1 722 739 6.88 CID5367801 C52H72MgN4O4 840
14 27.68 1352685 0.72 17-Pentatriacontene 1 720 737 46.41 6971-40-0 C35H70 490
15 29.08 76996880 27.61 1,3-Dimethoxybenzene 2 712 767 11.52 151-10-0 C8H10O2 138
16 30.51 29595091 16.9 1,3-Benzenediol, 5-pentadecyl 1 664 797 23.45 3158-56-3 C21H36O2 320
17 32.06 11993388 5.75 Cholest-4-ene-3,6-dione 1 710 773 32.12 984-84-9 C27H42O2 398
18 39.36 4575754 2.12 Stigmast-5-en-3-ol, (3β,24S) 1 757 821 54.07 83-47-6 C29H50O 414
19 41.64 684162 0.32 Stigmast-4-en-3-one 1 508 755 16.81 1058-61-3 C29H48O 412
Open in a new tab

Methanol root extract

The methanol root extract revealed the presence of 18 different chemotypes (Table 5, Fig. 5). Among the identified chemotypes, stigmast-5-en-3-ol, (3β,24S) (21.91%), octadecane, 1-chloro (17.01%), ethanone, 1-(4-hydroxyphenyl) (11.07%), α-tocopherol (8.42%), ascaridole (5.92%), and campesterol (4.98%) were found to present in major amount, while, linolein, 2-mono (3.99%), hexadecanoic acid (3.67%), 1,3-dimethoxybenzene (3.57%), ethyl linoleate (3.35%), 1-dotriacontane (2.21%), linolein, 1-mono (1.74%), methyl palmitate (1.73%), stigmast-4-en-3-one (1.55%), 1-dodecane (0.66%), δ-tocopherol (0.56%), and 3-methoxy-5-methylphenol (0.43%) were found to be present in trace.

Table 5. Phyto-chemotypes identified in the methanol root extract of R. imbricata by GC/MS.
S. No. Peak RT (min) Peak area Peak area % Compound detected Hit SI RSI Prob CAS No Mol. Formula Mol. Wt.
1 13.73 224745 0.43 3-Methoxy-5-methylphenol 1 730 810 72.09 3209-13-0 C8H10O2 138
2 15.78 5548712 11.07 Ethanone, 1-(4-hydroxyphenyl) 1 884 910 60.39 99-93-4 C8H8O2 136
3 17.87 345721 0.66 1-Dodecane 1 699 751 16.38 112-40-3 C12H26 170
4 19.33 1157289 2.21 1-Dotriacontane 1 777 785 14.73 544-85-4 C32H66 450
5 20.69 8405167 17.01 Octadecane, 1-chloro 1 734 738 22.3 386-33-2 C18H37Cl 288
6 22.32 905705 1.73 Hexadecanoic acid, methyl ester 1 793 866 61.21 112-39-0 C17H34O2 270
7 22.89 1901742 3.67 Hexadecanoic acid 4 660 762 11.87 57-10-3 C16H32O2 256
8 24.36 1592833 3.99 9,12-Octadecadienoic acid (Z,Z)-, 2-hydroxy-1-(hydroxymethyl)ethyl ester 5 807 833 6.93 3443-82-1 C21H38O4 354
9 24.44 821776 1.74 9,12,15-Octadecatrienoic acid, 2,3-dihydroxypropyl ester, (Z,Z,Z) 2 804 825 32.86 18465-99-1 C21H36O4 352
10 24.96 1495805 3.35 Ethyl linoleate 10 681 835 2.99 544-35-4 C20H36O2 308
11 29.06 1614826 3.57 1,3-Dimethoxybenzene 34 471 675 0.33 151-10-0 C8H10O2 138
12 30.91 3109377 5.92 Ascaridole 1 619 700 57.05 512-85-6 C10H16O2 168
13 31.92 3533697 7.23 Unknown - - - - - - -
14 33.77 174824 0.56 δ-Tocopherol 1 667 763 89.78 119-13-1 C27H46O2 402
15 36.71 4419092 8.42 α-Tocopherol 1 731 853 46.36 59-02-9 C29H50O2 430
16 38.04 2615057 4.98 Campesterol 1 726 810 24.4 474-62-4 C28H48O 400
17 39.36 11245680 21.91 Stigmast-5-en-3-ol, (3β,24S) 1 813 850 67.6 83-47-6 C29H50O 414
18 41.38 902689 1.55 Stigmast-4-en-3-one 45 395 709 0.74 1058-61-3 C29H48O 412
Open in a new tab

60% Ethanol root extract

GC/MS chemometric profile of 60% ethanol root extracts illustrated the presence of 12 different chemotypes (Table 6, Fig. 6). Amongst the identified chemotypes, dotriacontane (5.69%), and heptadecane, 9-hexyl (5.44%) were found to be present in major amount, whereas, bis(2-ethylhexyl) phthalate (3.58%), hexadecanoic acid, methyl ester (2.27%), and dibutyl phthalate (1.23%) were found to be present in trace.

Table 6. Phyto-chemotypes identified in the 60% ethanol root extract of R. imbricata by GC/MS.
S. No. Peak RT (min) Peak area Peak area % Compound detected Hit SI RSI Prob CAS No MF MW
1 19.37 258681 5.69 1-Dotriacontane 1 769 805 39.82 544-85-4 C32H66 450
2 20.72 276232 5.44 Heptadecane, 9-hexyl 1 696 719 24.93 55124-79-3 C23H48 324
3 21.56 62567 1.23 Dibutyl phthalate 5 799 875 5.32 84-74-2 C16H22O4 278
4 22.36 115053 2.27 Hexadecanoic acid, methyl ester 1 624 700 20.11 112-39-0 C17H34O2 270
5 23.22 296422 5.84 Unknown - - - - - - -
6 24.48 194215 3.83 Unknown - - - - - - -
7 25.41 56168 1.11 Unknown - - - - - - -
8 29.1 181582 3.58 Bis(2-ethylhexyl) phthalate 2 697 786 11.81 117-81-7 C24H38O4 390
9 34.21 644518 12.7 Unknown - - - - - - -
10 39.51 1832781 35.53 Unknown - - - - - - -
11 39.95 550771 10.86 Unknown - - - - - - -
12 40.92 604576 11.92 Unknown - - - - - - -
Open in a new tab

Discussion

We have conducted the present investigation to identify the major volatile and semivolatile components in the root of R. imbricata. The presence of various bioactive compounds justifies the use of the plant by traditional practitioners of ‘Amchi’ system of medicine in trans-Himalayan Ladakh region. Also, extensive pharmacological studies were conducted by different researchers with the plant root extracts [6]–[23] and the results were very promising to justify the use of this plant as therapeutic agent.

However, the phytochemical profiling of the plant root still remains to be unexplored and to the best of our knowledge, this is the first ever study of its kind on the GC/MS chemometric profiling of the root extracts. In medicinal chemistry, it is very essential to ascertain the chemotyping of medicinal plant parts that are responsible for its numerous pharmacological properties and by this technique we may be able to scientifically determine and validate the traditional uses, pharmacological activities, and therapeutic potential of these plant parts. Profiling of metabolites in plant extracts permits the complete phenotyping of genetically or environmentally adapted plant systems and such investigations draw on simple extraction procedures that have been shown to be very robust and have permitted broad range of high-throughput applications in plant metabolomics. [25]–[27].

The major phytochemical groups in n-hexane, ethyl acetate, and 60% ethanol extracts were saturated alcohol (50%), phenols (40%), and alkanes (61%) respectively. On the other hand, phytosterols were the major group in chloroform (31%), dichloroethane (27%), and methanol (40%) extracts. The total of various volatile and semi volatile groups present in different root extracts of R. imbricata had the following distribution order: phytosterols (122%), alkanes (83%), phenols (69.46%), esters (48%), ethers (44%), fatty acid esters (43%), fatty acids (33%), terpenoids (18%), arenes (16%), alkyl ammonium halide salt (15%), alkenes (10%), sulfonic acid (8%), unsaturated alcohols (8%), organic acids (4%), saturated alcohols (4%), glycosides (3%), photosynthetic pigments (3%), steroidal glycoside (1%). The order of extraction capacities of different polarity solvents for phytosterols, phenols, fatty acids, alkanes, esters, fatty acid esters, ethers, unsaturated alcohols, arenes, terpenoids, alkenes, sulfonic acid, photosynthetic pigment, and saturated alcohols was as follows:

  1. Phytosterols: methanol (40%), chloroform (31%), dichloroethane (27%), n-hexane (16%), ethyl acetate (8%)

  2. Phenols: ethyl acetate (40%), methanol (13%), chloroform (10%), dichloroethane (6%), n-hexane (0.46%)

  3. Fatty acids: dichloroethane (10%) = methanol (10%), ethyl acetate (8%), chloroform (4%), n-hexane (1%)

  4. Alkane: 60% ethanol (61%), n-hexane (9%), chloroform (8%), dichloroethane (5%)

  5. Esters: 60% ethanol (26%), chloroform (12%), n-hexane (8%), dichloroethane (2%)

  6. Fatty acid esters: dichloroethane (15%), 60% ethanol (13%), methanol (6%), chloroform (5%), n-hexane (4%)

  7. Ethers: ethyl acetate (28%), dichloroethane (12%), methanol (4%)

  8. Unsaturated alcohols: n-hexane (4%), ethyl acetate (3%), chloroform (1%)

  9. Arenes: chloroform (8%) = ethyl acetate (8%)

  10. Terpenoids: dichloroethane (12%), methanol (6%)

  11. Alkenes: n-hexane (7%), chloroform (3%)

  12. Sulfonic acid: dichloroethane (5%), chloroform (3%)

  13. Photosynthetic pigment: dichloroethane (2%), ethyl acetate (1%)

  14. Saturated alcohols: methanol (3%), ethyl acetate (1%)

The steroidal glycoside, alkyl ammonium halide salt, organic acids, and glycoside were found only in n-hexane (1%), chloroform (15%), dichloroethane (4%), and ethyl acetate (3%), respectively. Eventually, in the present study we have found phytosterols, terpenoids, fatty acids, fatty acid esters, alkyl halides, phenols, alcohols, ethers, alkanes, and alkenes as the major group of phyto-chemotypes in the different root extracts of R. imbricate (Fig. 7, Table 7). All these compounds identified by GC/MS analysis (Fig. 8) were further investigated for their biological activities [28] and most of them were found to possess a diverse range of positive pharmacological actions (Table 8).

Figure 7. Estimation of major phytochemical groups in different root extracts of R. imbricata, a) n-hexane extract, b) chloroform extract, c) dichloroethane extract, d) ethyl acetate extract, e) methanol extract, f) 60% ethanol extract.

Figure 7

Open in a new tab

Table 7. Distribution of phyto-chemotypes in different root extracts of R. imbricatea.

Phyto-chemotypes Root extracts
n-Hexane Chloroform Dichloroethane Ethyl acetate Methanol 60% Ethanol
1,30-Triacontanediol √ – – – – –
1,3-Benzenediol, 5-methyl – √ – √ – –
1,3-Benzenediol, 5-pentadecyl – – – √ – –
13-Docosen-1-ol, (Z) √ – – – – –
13-Tetradecen-1-ol acetate √ – – – – –
17-Pentatriacontene √ √ – √ – –
1-Chloro-2,4-dimethoxybenzene – – √ – – –
1-Dodecanol, 3,7,11-trimethyl – – – √ – –
Eicosen-1-ol, cis-9 – – – √ – –
1-Hentetracontanol √ – √ – – –
1-Pentacosanol √ √ – – – –
1-Tetracosanol √ √ – – – –
1-Tricosanol √ – – – – –
3-Methoxy-5-methylphenol √ √ √ √ √ –
7,8-Dimethylbenzocyclooctene – √ – √ – –
9,12,15-Octadecatrienoic acid, 2,3-dihydroxypropyl ester, (Z,Z,Z) – – – – √ –
9,12-Octadecadienoic acid (Z,Z)-, 2-hydroxy-1-(hydroxymethyl)ethyl ester – – – – √ –
Ascaridole – – – – √ –
Bacteriochlorophyll-c-stearyl – – √ √ – –
Benzene, 1,3-dimethoxy – – – √ √ –
Benzenemethanol, 3-hydroxy-5-methoxy – √ – √ – –
Benzene sulfonic acid, 4-amino-3-nitro – √ √ – – –
Benzenethiol, 4-chloro – – √ – – –
Bis(2-ethylhexyl) phthalate √ √ – – – √
Borneol – – √ – – –
Campesterol √ √ √ – √ –
Camphor – – √ – – –
Cholest-4-ene-3,6-dione – – – √ – –
Stigmast-5-en-3-ol, (3β,24S) √ √ √ √ √ –
Di-butyl phthalate – – – – – √
1-Dodecane – – – – √ –
Dodecanoic acid, 3-hydroxy – – – √ – –
1-Dotriacontane – – √ – √ √
Ethanone, 1-(2,6-dihydroxy-4-methoxyphenyl) – – √ – – –
Ethanone, 1-(4-hydroxyphenyl) – – – – √ –
Ethyl linoleate √ √ √ – √ –
Eucalyptol – – √ – – –
α-D-Glucopyranoside, O-α-D-glucopyranosyl-(1.fwdarw.3)-β-D-fructofuranosyl – – – √ – –
1-Hentetracontanol – √ – – – –
1-Hentriacontane √ – – – – –
1-Heptacosane √ – – – – –
Heptadecane, 9-hexyl – – – – – √
Hexadecanoic acid √ √ √ √ √ –
Hexadecanoic acid, methyl ester – – – – √ √
Hexanedioic acid, bis(2-ethylhexyl) ester – – √ – – –
Linalyl isovalerate – – √ – – –
1,3-Dimethoxybenzene – – √ – – –
Methanol, (4-carboxymethoxy)benzoyl – – √ – – –
Octadecane, 1-chloro – – – – √ –
Oleic acid – – √ √ – –
1-Pentatriacontane √ – – – – –
Phenol, 2,4-bis(1,1-dimethylethyl) – – √ √ – –
Phenol, 3,5-dimethoxy, acetate – – – √ – –
Stigmast-4-en-3-one √ √ √ √ √ –
Stigmast-3,5-dien-7-one – – √ – – –
Stigmastanol √ – – – – –
1-Tetratetracontane √ – – – – –
Thujone – – √ – – –
Methyl tri-butyl ammonium chloride – √ – – – –
α-Tocopherol √ √ – – √ –
β-Fenchyl alcohol – – √ – – –
δ-Tocopherol – – – – √ –
α-Tocopherol-β-D-mannoside √ – – – – –
Open in a new tab

√ Present; – Absent.

Figure 8. Phyto-chemotypes identified in different root extracts of R. imbricata.

Figure 8

Open in a new tab

1: 1,30-triacontanediol; 2: 1,3-benzenediol, 5-methyl; 3: 1,3-benzenediol, 5-pentadecyl; 4: 13-docosen-1-ol, (Z); 5: 13-tetradecen-1-ol acetate; 6: 17-pentatriacontene; 7: 1-chloro-2,4-dimethoxybenzene; 8: 1-dodecanol, 3,7,11-trimethyl; 9: eicosen-1-ol, cis-9; 10: 1-hentetracontanol; 11: 1-pentacosanol; 12: 1-tetracosanol; 13: 1-tricosanol; 14: 3-methoxy-5-methylphenol; 15: 7,8-dimethylbenzocyclooctene; 16: 9,12,15-octadecatrienoic acid, 2,3-dihydroxypropyl ester, (Z,Z,Z); 17: 9,12-octadecadienoic acid (Z,Z)-, 2-hydroxy-1-(hydroxymethyl)ethyl ester; 18: ascaridole; 19: bacteriochlorophyll-c-stearyl; 20: benzene, 1,3-dimethoxy; 21: benzenemethanol, 3-hydroxy-5-methoxy; 22: benzene sulfonic acid, 4-amino-3-nitro; 23: benzenethiol, 4-chloro; 24: bis(2-ethylhexyl) phthalate; 25: borneol; 26: campesterol; 27: camphor; 28: cholest-4-ene-3,6-dione; 29: stigmast-5-en-3-ol, (3β,24S); 30: di-butyl phthalate; 31: 1-dodecane; 32: dodecanoic acid, 3-hydroxy; 33: 1-dotriacontane; 34: ethanone, 1-(2,6-dihydroxy-4-methoxyphenyl); 35: ethanone, 1-(4-hydroxyphenyl); 36: ethyl linoleate; 37: eucalyptol; 38: α-D-glucopyranoside, O-α-D-glucopyranosyl-(1.fwdarw.3)-β-D-fructofuranosyl; 39: 1-hentetracontanol; 40: 1-hentriacontane; 41: 1-heptacosane; 42: heptadecane, 9-hexyl; 43: hexadecanoic acid; 44: hexadecanoic acid, methyl ester; 45: hexanedioic acid, bis(2-ethylhexyl) ester; 46: linalyl isovalerate; 47: 1,3-dimethoxybenzene; 48: methanol, (4-carboxymethoxy)benzoyl; 49: octadecane, 1-chloro; 50: oleic acid; 51: 1-pentatriacontane; 52: phenol, 2,4-bis(1,1-dimethylethyl); 53: phenol, 3,5-dimethoxy acetate; 54: stigmast-4-en-3-one; 55: stigmast-3,5-dien-7-one; 56: stigmastanol; 57: 1-tetratetracontane; 58: thujone; 59: methyl tri-butyl ammonium chloride; 60: α-tocopherol; 61: β-fenchyl alcohol; 62: δ-tocopherol; 63: α-tocopherol-β-D-mannoside.

Table 8. Biological activities of active principles present in different root extracts of R. imbricate.

Phyto-chemotypes Biological activity
Eicosen-1-ol, cis-9 Antimalarial, antifungal, antioxidant
1-Tricosanol Antibacterial, antifungal
9,12,15-Octadecatrienoic acid, 2,3-dihydroxypropyl ester, (Z,Z,Z) 5-Alpha-reductase inhibitor, antiMS, antiacne, antialopecic, antianaphylactic, antiandrogenic, antiarteriosclerotic, antiarthritc, anticoronary, antieczemic, antifibrinolytic, antigranular, antihistaminic, antiinflammatory, antileukotriene-D4, antimenorrhagic, antiprostatitic, cancer-preventive, carcinogenic, comedolytic, hepatoprotective, hypocholesterolemic, immunomodulator, insectifuge, metastatic, nematicide, propecic
9,12-Octadecadienoic acid (Z,Z)-, 2-hydroxy-1-(hydroxymethyl)ethyl ester Antiinflammatory, hypocholesterolemic, cancer preventive, hepatoprotective, nematicide, insectifuge, antihistaminic, antieczemic, antiacne, 5-alpha reductase inhibitor antiandrogenic, antiarthritic, anticoronary, insectifuge
Ascaridole Analgesic, ancylostomicide, anthelmintic, antiflatulent, antimalarial, carcinogenic, carminative, fungicide, nematicide, pesticide, plasmodicide, sedative, transdermal, trypanocide, vermifuge
Borneol (-)-Chronotropic, (-)-inotropic, allelochemic, analgesic, antiacetylcholine, antibacterial, antibronchitic, antiescherichic, antifeedant, antiinflammatory, antiotitic, antipyretic, antisalmonella, antispasmodic, antistaphylococcic, antiyeast, CNS-stimulant, CNS-toxic, candidicide, choleretic, flavor; fungicide, hepatoprotective, herbicide, herbicide, inhalant, insect-repellent, insectifuge, irritant, myorelaxant, nematicide, perfumery, pesticide, sedative, tranquilizer
Campesterol Antioxidant, hypocholesterolemic
Camphor Allelopathic, analgesica, anesthetic, antiacne, antidiarrheic, antidysenteric, antiemetic, antifeedant, antifibrositic, antineuralgic, antipruritic, antiseptic, antispasmodic, CNS-stimulant, cancer preventive, carminative, convulsant, cosmetic, counterirritant, decongestant, deliriant, ecbolic, emetic, epileptigenic, expectorant, fungicide, herbicide, insect-repellent, insectifuge, irritant, nematicide, occuloirritant, P450-2B1-inhibitor, pesticide, respirainhibitor, respirastimulant, rubefacient, stimulant, transdermal, verrucolytic, vibriocide
Stigmast-5-en-3-ol, (3β,24S) Androgenic, angiogenic, anorexic, antiadenomic, antiandrogenic, antibacterial, anticancer (breast), anticancer (cervix), anticancer (lung), antiedemic, antiestrogenic, antifeedant, antifertility, antigonadotrophic, antihyperlipoproteinaemic, antiinflammatory, antileukemic, antilymphomic, antimutagenic, antiophidic, antioxidant, antiprogestational, antiprostaglandin, antiprostatadenomic, antiprostatitic, antipyretic, antitumor (breast), antitumor (cervix), antitumor (lung), antiviral, apoptotic, artemicide, cancer-preventive, candidicide, caspase-8-inducer, estrogenic, febrifuge, gonadotrophic, hepatoprotective, hypocholesterolemic, hypoglycemic, hypolipidemic, pesticide, spermicide, ubiquiot, ulcerogenic
Di-butyl phthalate Antimicrobial, Antifouling
Dodecanoic acid, 3-hydroxy Flavor
Eucalyptol Anesthetic, anthelmintic, antibacterial, antihalitosic, antiseptic, antitussive, decongestant, expectorant, hypotensive, insectifuge, irritant, pesticide, vermicide
α-D-Glucopyranoside, O-α-D-glucopyranosyl-(1.fwdarw.3)-β-D-fructofuranosyl Preservative
Hexadecanoic acid Antioxidant, hypocholesterolemic, nematicide, pesticide, lubricant, antiandrogenic, flavor, hemolytic 5-alpha reductase inhibitor
Hexadecanoic acid, methyl ester Antioxidant, nematicide, pesticide, lubricant, antiandrogenic, flavor, hemolytic 5-alpha reductase inhibitor, hypocholesterolemic
Linalyl isovalerate Fragrance
Oleic acid 5-Alpha-reductase-inhibitor, allergenic, anemiagenic, antialopecic, antiandrogenic, antiinflammatory, antileukotriene-D4; cancer-preventive, choleretic, dermatitigenic, flavor, hypocholesterolemic, insectifuge, irritant, percutaneostimulant, perfumery, propecic
1-Pentatriacontane Herbistat
Stigmast-4-en-3-one Antiprostatitic
Stigmast-3,5-dien-7-one Antifertility
Thujone Abortifacient, anthelmintic, antibacterial, antiseptic, antispasmodic, cerebrodepressant, convulsant, counterirritant, emmenagogue, epileptigenic, hallucinogenic, herbicide, neurotoxic, perfumery, pesticide, respirainhibitor, toxic
β-Fenchyl alcohol Antimicrobial, antioxidant, flavor
δ-Tocopherol 5-HETE-inhibitor, allergenic, analgesic, antiMD, antiMS, antiPMS, antiaggregant, antiaging, antialzheimeran, antianginal, antiarteriosclerotic, antiarthritic, antiatherosclerotic, antibronchitic, anticancer (breast), anticariogenic, anticataract, antichorea, antichoreic, anticonvulsant, anticoronary, antidecubitic, antidementia, antidermatitic, antidiabetic, antidysmenorrheic, antiepitheleomic, antifibrositic, antiglycosation, antiherpetic, antiinfertility, antiinflammatory, antiischemic, antileukemic, antileukotriene, antilithic, antilupus, antimaculitic, antimastalgic, antimelanomic, antimyoclonic, antineuritic, antineuropathic, antinitrosaminic, antiophthalmic, antiosteoarthritic, antioxidant, antiparkinsonian, antiproliferant, antiradicular, antiretinopathic, antirheumatic, antisenility, antisickling, antispasmodic, antisterility, antistroke, antisunburn, antisyndrome-X, antithalassemic, antithrombotic, antithromboxane-B2, antitoxemic, antitumor; antitumor (breast), antitumor (colorectal), antitumor (prostate), antitumor (stomach), antiulcerogenic, apoptotic, calcium-antagonist, cancer-preventive, cardioprotective, cerebroprotective, circulatory-stimulant, circulotonic, hepatoprotective, hypocholesterolemic, hypoglycemic, immunomodulator, immunostimulant, insulin-sparing, lipoxygenase-inhibitor, NO-inhibitor, ornithine-decarboxylase-inhibitor, P21-inducer, phospholipase-A2-inhibitor, protein-kinase-C-inhibitor, vasodilator
Open in a new tab

Most of the pharmacological studies were conducted with the aqueous, ethanol, and hydro-alcoholic root extracts of this plant and it was found to have numerous biological activities such as anti-stress, adaptogenic, anti-hypoxic, immune-stimulatory, anti-cancer, cytoprotective, radioprotective, anti-hemolytic, anti-inflammatory, and wound healing potential [6]–[23]. Our investigations conclude that the compounds present in the ethanol and water extracts have the potential to perform these functions. Though, the root extracts of the plant obtained by polar solvent extraction have been investigated for their pharmacological actions in considerable detail, non polar root extracts were not studied till date. Hence, our primary objective in the present work was to find the bioactive constituents present in the non polar extraction of root of this herb. These findings will definitely usher in new directions in pharmacological and therapeutic investigations with the root extracts obtained from non polar solvent extraction such as n-hexane, chloroform, dichloroethane, and ethyl acetate.

Conclusion

In the present study, sixty three phyto-chemotypes have been identified from n-hexane, chloroform, dichloroethane, ethyl acetate, methanol, and 60% ethanol root extracts of R. imbricata by GC/MS analysis. It showed the existence of various bioactive principles that confirm the application of R. imbricata for various ailments in traditional system of medicine. However, isolation of individual phyto-chemotypes and subjecting them to biological activity will definitely give fruitful results to find a novel drug. It could be concluded that R. imbricata contains various bioactive phyto-chemotypes having phyto-pharmaceutical importance. However, further studies will need to be undertaken to ascertain its bioactivity, toxicity profile, effect on the ecosystem, and agricultural products.

Acknowledgments

Authors are thankful to Dr. Bassant Ballabh, STA ‘C’, Defence Institute of Bio-Energy Research, Defence Research & Development Organisation, Goraparao, PO-Arjunpur, Haldwani, PIN - 263 139, Uttarakhand, India, for critical review of the manuscript. Authors also acknowledge our colleague Mr. Ritendra Mishra for copyediting of the manuscript.

Funding Statement

Funding provided by Defence Research & Development Organization, Ministry of Defence, Government of India. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

References

  • 1. Gherman C, Culea M, Cozar O (2000) Comparative analysis of some active principles of herb plants by GC/MS. Talanta 53: 253–262. [DOI] [PubMed] [Google Scholar]
  • 2. Carro N, Garcia CM, Cela R (1997) Microwave-assisted extraction of monoterpenols in must samples. Analyst 122: 325–329. [Google Scholar]
  • 3. Robards K, Antolovich M (1997) Analytical chemistry of fruit bioflavonoids. Analyst 122: 11–34. [Google Scholar]
  • 4. Pallado P, Tassinato G, D'Alpaos D, Traldi P (1997) Gas chromatography/mass spectrometry in aroma chemistry: a comparison of essential oils and flavours extracted by classical and supercritical techniques. Rapid Commun Mass Spectrom 11: 1335–1341. [Google Scholar]
  • 5. Iordache A, Culea M, Gherman C, Cozar O (2009) Characterization of some plant extracts by GC–MS. Nuclear Instruments and Methods in Physics Research B 267: 338–342. [Google Scholar]
  • 6. Mishra KP, Ganju L, Singh SB (2012) Anti-cellular and immunomodulatory potential of aqueous extract of Rhodiola imbricata rhizome. Immunopharmacol Immunotoxicol 34: 513–518. [DOI] [PubMed] [Google Scholar]
  • 7. Tulsawani R, Meena DK, Shukla H, Sharma P, Meena RN, et al. (2011) Ninety days of repeated gavage administration of Rhodiola imbricata extract in rats. J Appl Toxicol doi:10.1002/jat.1739. [DOI] [PubMed] [Google Scholar]
  • 8. Chawla R, Jaiswal S, Kumar R, Arora R, Sharma RK (2010) Himalayan bioresource Rhodiola imbricata as a promising radioprotector for nuclear and radiological emergencies. J Pharma and Bioallied Sci 2: 213–219. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9. Gupta V, Lahiri SS, Sultana S, Tulsawani RK, Kumar R (2010) Anti-oxidative effect of Rhodiola imbricata root extract in rats during cold, hypoxia and restraint (C-H-R) exposure and post-stress recovery. Food Chem Toxicol 48: 1019–1025. [DOI] [PubMed] [Google Scholar]
  • 10. Mishra KP, Chanda S, Shukla K, Ganju L (2010) Adjuvant effect of aqueous extract of Rhodiola imbricata rhizome on the immune responses to tetanus toxoid and ovalbumin in rats. Immunopharmacol Immunotoxicol 32: 141–146. [DOI] [PubMed] [Google Scholar]
  • 11. Kumar R, Phani Kumar G, Chaurasia OP (2010) In vitro antioxidant activity of methanolic extract of Rhodiola imbricata Edgew. Phcog J 2: 157–161. [Google Scholar]
  • 12. Kumar R, Tayade A, Chaurasia OP, Hota S, Singh SB (2010) Evaluation of anti-oxidant activities and total phenol and flavonoid content of the hydro-alcoholic extracts of Rhodiola sp. Phcog J 2: 431–435. [Google Scholar]
  • 13. Gupta V, Lahiri SS, Sultana S, Kumar R (2009) Mechanism of action of Rhodiola imbricata Edgew during exposure to cold, hypoxia and restraint (C-H-R) stress induced hypothermia and post stress recovery in rats. Food Chem Toxicol 47: 1239–1245. [DOI] [PubMed] [Google Scholar]
  • 14. Mishra KP, Ganju L, Chanda S, Karan D, Sawhney RC (2009) Aqueous extract of Rhodiola imbricata rhizome stimulates Toll-like receptor 4, granzyme-B and Th1 cytokines in vitro . Immunobiology 214: 27–31. [DOI] [PubMed] [Google Scholar]
  • 15. Gupta V, Saggu S, Tulsawani RK, Sawhney RC, Kumar R (2008) A dose dependent adaptogenic and safety evaluation of Rhodiola imbricata Edgew, a high altitude rhizome. Food Chem Toxicol 46: 1645–1652. [DOI] [PubMed] [Google Scholar]
  • 16. Mishra KP, Padwad YS, Dutta A, Ganju L, Sairam M, et al. (2008) Aqueous extract of Rhodiola imbricata rhizome inhibits proliferation of an erythroleukemic cell line K-562 by inducing apoptosis and cell cycle arrest at G2/M phase. Immunobiology 213: 125–131. [DOI] [PubMed] [Google Scholar]
  • 17. Arora R, Singh S, Sagar RK, Chawla R, Kumar R, et al. (2008) Radiomodulatory and free-radical scavenging activity of the fractionated aquo-alcoholic extract of the adaptogenic nutraceutical (Rhodiola imbricata)-A comparative in vitro assessment with ascorbate. J Diet Suppl 5: 147–163. [DOI] [PubMed] [Google Scholar]
  • 18. Gupta A, Kumar R, Upadhyay NK, Pal K, Kumar R, et al. (2007) Effects of Rhodiola imbricata on dermal wound healing. Planta Med 73: 774–777. [DOI] [PubMed] [Google Scholar]
  • 19. Kumar R, Gupta V, Saggu S, Tulswania RK, Divekar HM (2007) Adaptogenic and safety evaluation of Rhodiola imbricata root extract: A dose dependent study. Ind J Clin Biochem 22(Supl.)20S3.1: 171. [Google Scholar]
  • 20. Mishra KP, Padwad YS, Jain M, Karan D, Ganju L, et al. (2006) Aqueous extract of Rhodiola imbricata rhizome stimulates proinflammatory mediators via phosphorylated IkappaB and transcription factor nuclear factor-kappaB. Immunopharmacol Immunotoxicol 28: 201–212. [DOI] [PubMed] [Google Scholar]
  • 21. Goel HC, Bala M, Prasad J, Singh S, Agrawala PK, et al. (2006) Radioprotection by Rhodiola imbricata in mice against whole-body lethal irradiation. J Med Food 9: 154–160. [DOI] [PubMed] [Google Scholar]
  • 22. Kanupriya, Prasad D, Sai Ram M, Kumar R, Sawhney RC, et al. (2005) Cytoprotective and antioxidant activity of Rhodiola imbricata against tert-butyl hydroperoxide induced oxidative injury in U-937 human macrophages. Mol Cell Biochem 275: 1–6. [DOI] [PubMed] [Google Scholar]
  • 23. Arora R, Chawla R, Sagar R, Prasad J, Singh S, et al. (2005) Evaluation of radioprotective activities Rhodiola imbricata Edgew - a high altitude plant. Mol Cell Biochem 273: 209–223. [DOI] [PubMed] [Google Scholar]
  • 24. Khanum F, Bawa AS, Singh B (2005) Rhodiola rosea: A Versatile Adaptogen. Comprehensive Reviews in Food Science and Food Safety 4: 55–62. [DOI] [PubMed] [Google Scholar]
  • 25. Shellie RA, Poynter SD, Li J, Gathercole JL, Whittock SP, et al. (2009) Varietal characterization of hop (Humulus lupulus L.) by GC–MS analysis of hop cone extracts. J Sep Sci 32: 3720–3725. [DOI] [PubMed] [Google Scholar]
  • 26. Roessner U, Luedemann A, Brust D, Fiehn O, Linke T, et al. (2001) Metabolic profiling allows comprehensive phenotyping of genetically or environmentally modifiedplant systems. Plant Cell 13: 11–29. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 27. Gullberg J, Jonsson P, Nordstrom A, Sjostrom M, Moritz T (2004) Design of experiments: an efficient strategy to identify factors influencing extraction and derivatization of Arabidopsis thaliana samples in metabolomic studies with gas chromatography/mass spectrometry. Anal Biochem 331: 283–295. [DOI] [PubMed] [Google Scholar]
  • 28.Dr. Duke's Phytochemical and Ethnobotanical Databases. http://www.ars-grin.gov/duke/ Accessed 2012 Aug 10.

Articles from PLoS ONE are provided here courtesy of PLOS

RESOURCES