Sharma Surendra Kumar et al / IJRAP 2011, 2 (3) 978-982

International Journal of Research in Ayurveda & Pharmacy, 2(3), 2011 978-982

Research Article Available online through www.ijrap.net ISSN 2229-3566

STANDARDIZATION AND PRELIMINARY PHYTOCHEMICAL SCREENING OF

NARDOSTACHYS JATAMANSI DC. RHIZOME Sharma Surendra Kumar* and Singh Ajay Pal

Department of Pharmaceutical Sciences, Guru Jambheshwar University of Science and Technology, Hisar, Haryana, India

Received on: 05/04/2011 Revised on: 27/05/2011 Accepted on: 15/06/2011

ABSTRACT The plant Nardostachys jatamansi DC. (Valerianaceae) is a critically endangered rhizome bearing medicinal herb. It is restricted to specialized habitats in high altitudes of Himalaya ranging from 3000 to 5000m. Nardostachys jatamansi has been traditionally employed in treatment of wide range of disorders including nervous system, digestive system, circulatory system, respiratory system, urinary system, reproductive system and skin diseases. It shows marked tranquillizing activity, as well as hypotensive, hypolipidemic, hepatoprotective, neuroprotective, anti-ischemic, antiarrhythmic, anticonvulsant activities. The present study was aimed to evaluate the parameters to determine the quality of Nardostachys jatamansi rhizome and its standardization. These studies comprise macroscopy, powder analysis, physiochemical parameters, fluorescence characteristics and preliminary phytochemical screening. These studies will provide referential information for the correct identification of the crude drug. KEYWORDS: Nardostachys jatamansi, Valerianaceae, Physicochemical parameters, Phytochemical screening, Fluorescence analysis. *Corresponding author Dr. Surendra Kr. Sharma, Professor and Chairman, Department of Pharmaceutical Sciences, Guru Jambheshwar University of Science and Technology, Hisar-125001, Haryana, India Email: prof.sharmask@gmail.com INTRODUCTION Nardostachys jatamansi DC (Family Valerianaceae), commonly known as ‘bhutjata’, ‘nalada’ or ‘spikenard’, is an erect perennial herb, 10-60m high with long, stout, woody rootstock1. It is found in the alpine Himalayas from Punjab to Sikkim and Bhutan at altitude of 3000- 5000m2. It is described in Indian traditional system of medicine (Ayurveda), for its use in mental disorder, insomnia, hyperlipidemia, hypertension and heart disease3. It has protective effect in epilepsy, parkinsonism, cerebral ischemia, liver damage4, 5. The rhizomes of jatamansi are used as a bitter tonic, stimulant and antispasmodic as well as to treat hysteria, palpitations and convulsion6, 7. Oil from its rhizome promotes the growth and blackness of hair. It is also recommended in scorpion string8. This plant is a major component of “Karpuradyarka”, an ayurvedic formulation used for brain disorder9. Phytochemical investigations have shown the presence of esters, phenolic compounds, alkaloids, sesquiterpene ketone (jatamansone)10, valeranone, valeranal, nordal,

jatamansin, β-sitosterol etc11,12. The present investigation was undertaken to standardize the rhizome of Nardostachys jatamansi with a view to develop its quality parameters, so as to curb substitution and adulteration. MATERIAL AND METHODS Plant material The plant material was collected at its flowering stage from Kullu district, Himachal Pradesh, July 2008. The plant was identified and authenticated by the Dr. H. B. Singh, Head, Raw Materials Herbarium & Museum, NISCAIR, New Delhi, Ref. No. NISCAIR/ RHMD/ Consult/06/741/58. The fresh rhizomes were taken for macroscopic and microscopic powder studies. The dried rhizomes were ground to a coarse powder and subjected to physicochemical analytical parameters like ash values, extractive values and phytochemical screening. Macroscopy The organoleptic features of crude drugs were observed with sensory organs and analyzed for colour, odour and

Sharma Surendra Kumar et al / IJRAP 2011, 2 (3) 978-982

International Journal of Research in Ayurveda & Pharmacy, 2(3), 2011 978-982

taste. The external features of rhizome were observed under dissecting microscope. Powder analysis The shade dried rhizomes were ground with a wood grinder and sifted through 40 mesh sieve. The ingredients of powder were observed under microscope. The powder was treated with different chemical reagents and observations were made. A small quantity of drug mounted in different reagents was observed under UV light and fluorescence was recorded13. Histochemical colour reactions The histochemical colour reactions on the rhizome of Nardostachys jatamansi were performed according to standard procedures reported. The colour tests were performed for the identification of major cell components14. Physicochemical parameters The determination of various physicochemical parameters such as total ash, acid insoluble ash, water soluble ash, sulphated ash, water soluble extractive and alcohol soluble extractive were determined and estimated in percentage by using method as recommended by Indian Pharmacopoeia15. Successive soxhlet extractives of the drug were carried out with various solvents and weight, colour/consistency of extractives were observed16. Loss on drying for drug was also determined17, 18. Preliminary phytochemical screening The different extractives were subjected to preliminary phytochemical investigation for the presence of various phytoconstituents like alkaloids, glycosides, flavonoids, steroids, saponins, phenolic compounds, etc. following usual testing procedures19, 20. Fluorescence analysis The physical and chemical parameters, when felt inadequate, as it often happens with the powdered drugs, the plant material may be identified from their adulterants on basis of fluorescence study. The fluorescence analyses of the rhizome powder was carried out by treating with different chemical reagents and observed under UV light and daylight21. RESULTS AND DISCUSSION The organoleptic characters of Nardostachys jatamansi rhizome are shown in (Table1). The rhizomes are dark brown, cylindrical and covered with fibers (Figure 1). The various diagnostic characters of powder are coarse, dark brown with strong aromatic odour and bitter taste. Microscopic examination of powder shows the presence of cork cells, starch, vessels and fibers (Figure 2). The histochemical colour reactions, which are important for

identification of major cell components, are shown in (Table 2). The physicochemical parameters are important for identifying adulterants and improper handling of drugs. The (Table 3) reveals the result of physicochemical parameters of powdered drug, carried out by using standard procedures. viz. ash values used to determine quality and purity of drug; the extractive values are useful to evaluate the chemical constituents present in crude drugs and also help in estimation of specific constituents soluble in particular solvent. The result of extractive values of powdered drug in different solvent obtained by successive extraction is given in (Table 4). Loss on drying is shown in (Table 5). The (Table 6) reveals the behavior analysis of different solvent extract of Nardostachys jatamansi rhizome, under visible light, short and long UV. All the extracts obtained by successive extraction were subjected to qualitative chemical tests and the results are shown in (Table 7). Such preliminary phytochemical screening is helpful in prediction of the nature of drugs and also useful for detection of different constituents in different polarity solvent. The fluorescence analysis of powdered drug in day light, short UV and long UV were examined by reported method21. The observations are given in (Table 8). CONCLUSION The organoleptic studies confirm shape, size, colour, odour and taste of Nardostachys jatamansi rhizome. The histochemical studies of transverse section with different reagents showed the various secondary plant metabolites. The standard behavior has been shown by different extracts under visible and U.V light. The information obtained from preliminary phytochemical investigation will be useful in finding out genuity of drug. Ash values, extractive values can be used as reliable aid for detection of adulteration. Thus we can conclude that the morphoanatomical characters, physicochemical values and pharmacognostical studies will serve as standard reference for identification and distinguishing Nardostachys jatamansi rhizome from its substituent and adulterants, and assist in future drug evaluation. This is first such report on pharmacognostical studies of Nardostachys jatamansi rhizome. REFERENCES 1. Jadhav VM, Throat RM, Kadam VJ, Kamble SS. Herbal

Anxiolyte: Nardostachys jatamansi. J Pharm Res 2009; 2(8): 1208-1211.

2. Anonymous, Indian Medicinal Plants. Vol-4. Madras: Orient Longman; 1997. p. 104

3. Subashini R, Ganapragasam A, Senthil SK, Yogeeeta S, Devaki T. Protective effect of Nardostachys jatamansi (Rhizomes) on mitochondrial respiration and lysosomal

Sharma Surendra Kumar et al / IJRAP 2011, 2 (3) 978-982

International Journal of Research in Ayurveda & Pharmacy, 2(3), 2011 978-982

hydrolases during doxorubicin induced myocardial injury in rats. J Health Science 2007; 29: 67-72

4. Ali S, Ansari KA, Jafery MA, Kabeer H, Diwakar G. Nardostachys jatamansi protects against liver damage induced by thioacetamide in rats. J Ethnopharm 2000;71: 359-363

5. Anonymous, The Wealth of India Vol. VII New Delhi: National Institute of Science Communication, CSIR; 2001. p. 3-4

6. Bagchi A, Oshima Y, Hikino H. Neolignans and lignans of Nardostachys jatamansi roots. Planta Med 1991; 57: 96-97

7. Nadkarni AK, Nadkarni KM. Indian Materia Medica. Bombay: Popular Book Depot; 1994. p. 235

8. Chopra IC, Jamwal KS, Khajuria BN. Pharmacological action of some common essential oil bearing plants used in indigenous medicine. Part II. Pharmacological action of Alpinia galangal, Pistacia integrima, Piper betel and Nardostachys jatamansi. Indian J Med Res 1954; 42: 385

9. Anonymous, The Ayurvedic Formulary of India, Part I. New Delhi: Govt. of India, Controller of Publication; 1978. p. 21-22

10. Govindachari TR, Rajadurai S, Pai BR. Structure of Jatamansone. Chem Ber 1958; 91: 908

11. Sastry SD, Maheshwari ML, Chakravati KK, Bhattacharya SC. Terenoids-CVI The Structure of Nardostachnone. Tetrahedrone 1967c; 23: 1997

12. Pai PP, Chakravarti KL, Paknikar SK. Revised structure for nardostachnone, Idetify with 1,8,9,10-tetrahydroaristolane. Indian J Chem 1971; 9: 729

13. Trease GE, Evans WC. Pharmacognosy. 15th Ed. New Delhi: WB Saunders Company Ltd.; 2009. p. 101-105

14. Harborne JB. Phytochemical Methods. 2nd Ed. London: Chapman and Hall; 1973. p. 5-6

15. Anonymous, Indian Pharmacopoeia. Vol-II. New Delhi: Govt. of India, Controller of Publication; 1996. A-53-55

16. Mukherjee PK. Quality Control of Herbal Drug. 1st Ed. New Delhi: Business Horizons Pharmaceutical Publishers; 2010. p. 184-191

17. Quality control methods for medicinal plants. World Health Organization, Geneva; 1998

18. Yadav N, Vasudeva N, Sharma SK, Singh S. Pharmacognostical and Phytochemical Studies on Chenopodium album Linn. root. Hamdard Medicus 2007; 50(1): 95-102

19. Khandelwal KR. Practical Pharmacognosy. 19th Ed. Pune: Nirali Prakashan; 2009. p. 149-156

20. Kokate CK. Practical Pharmacognosy. 4th Ed. New Delhi: Vallabh Prakashan; 2008. p. 107-111

21. Kokoski CJ, Kokoski RJ, Salma FJ. Fluorescence of powdered vegetable drugs under UV radiation. J Am Pharm Assoc 1958; 47: 715-717 .

Table 1: Morphological Characters of Nardostachys jatamansi Rhizome

Table 2: Histochemical Colour Reactions of the T.S. of Nardostachys jatamansi Rhizome

S. No. Reagents Test for Nature of change Degree of change

1 Aniline sulphate & H2SO4 Lignin Pink colour + 2 Phloroglucinol + HCl Lignin Pink colour ++ 3 Iodine solution followed by H2SO4 Cellulose Brown colour ++ 4 Million’s reagent Protein Yellow + 5 FeCl3 solution Tannins Black ++ 6 Caustic alkali + HCl Calcium oxalate - - 7 Acetic anhydride + H2SO4 Steroids Black + 8 Dragendroff’s reagent Alkaloids Orange + 9 Weak iodine solution Starch Blue-black ++

++ = High, + = Moderate, - = Absent

Table 3: Physicochemical Parameters of Rhizomes Powder of Nardostachys jatamansi Physicochemical parameters Value* (% W/W)

Ash values Total ash 8.5

Acid insoluble ash 3.8 Water soluble ash 0.5

Sulphated ash 8.7 Extractive values

Water soluble extractive 7.3 Alcohol soluble extractive 5.2

*Data are mean of 3 values

S. No Rhizome Characters 1 Shape Cylindrical, covered with reddish brown fibers forming a network which are

skeletons of sheathing leaf bases 2 Size 3 to 7 cm long, 0.5 to 1.5 cm in diameter 3 Colour Dark Brown externally and Reddish Brown internally 4 Fracture Brittle 5 Surface Fibrous 6 Odour Strongly aromatic 7 Taste Acrid, slightly bitter and aromatic

Sharma Surendra Kumar et al / IJRAP 2011, 2 (3) 978-982

International Journal of Research in Ayurveda & Pharmacy, 2(3), 2011 978-982

Table 4: Yield of Extract by Successive Solvent Extraction

Sr. No. Extract* % W/W 1. Petroleum Ether 3.7 2. Chloroform 1.1 3. Methanol 2.3 4. Water 3.1

*Extraction period is 24 hrs

Table 5: Loss on Drying

Wt. of the crucible Wt. of crucible+ Drug Final wt. %W/W

36.58gm 37.89gm 37.80gm 6.87% 37.79gm 7.63% 37.79gm 7.63%

Table 6: Fluorescence Nature of Different Extracts of Nardostachys jatamansi by Visible and Ultra-violet (UV) Radiations

S. No. Extract

Observations

Visible Short UV (254 nm) Long UV (366nm)

1. Petroleum ether Light yellow Yellowish Fluorescent 2. Chloroform Brown Greenish brown Light brown 3. Ethanol Brown Greenish brown Brown

4. Water Black Greenish brown Black

Table 7: Preliminary Phytochemical Screening of Extract of Nardostachys jatamansi S.

No. Plant Constituents Pet. Ether Extract Chloroform Extract Ethanol Extract Water Extract

1. Alkaloids - - + + 2. Glycosides - - - - 3. Protein & Amino

acids - - + +

4. Carbohydrates - - + + 5. Tannins & Phenolics - + + + 6. Flavonoids - - + + 7. Steroids + + - - 8. Saponins - - + + 9. Fixed oil + - - -

+: Positive; -: Negative Table 8: Fluorescent Nature of Nardostachys jatamansi Rhizome Powder

S. No. Treatment Observations Visible Short UV (254 nm) Long UV (366nm)

1. Powder as such Brown Green Dark brown 2. Powder + 1N HCl Black Green Light brown 3. Powder + 1N NaOH Greenish Brown Green Dark brown 4. Powder + 50% HNO3 Black Greenish black Light brown 5. Powder + 50% H2SO4 Brown Greenish brown Dark brown 6. Powder + Methanol Brown Chocolate Dark brown 7. Powder + Acetic acid Brown Greenish brown Dark brown

8. Powder + Picric acid Yellowish brown Yellowish brown Dark brown

9. Powder + 1N NaOH in methanol Brownish black Greenish black Brown 10. Powder + Nitrocellulose in amyl acetate Brown Green Yellowish brown

11. Powder + 1N NaOH in methanol+ nitrocellulose in amyl acetate Brown Green Yellowish brown

12. Powder + 1N HCl + Nitrocellulose in amyl acetate Brown Green Brown 13. Powder + 1N NaOH + Nitrocellulose in amyl acetate Brown Green Brown

Sharma Surendra Kumar et al / IJRAP 2011, 2 (3) 978-982

International Journal of Research in Ayurveda & Pharmacy, 2(3), 2011 978-982

Figure 1: Nardostachys jatamansi rhizome

Figure 2: Powder microscopy of Nardostachys jatamansi rhizome showing Ck = Cork, Ph. f = Phloem fibre, V = Vessel, S = Starch

Source of support: Nil, Conflict of interest: None Declared