INTRODUCTION:

Acorus calamus Linn. (Commonly called as ‘Sweet flag’) of family Araceae, is a semi-aquatic, perennial, aromatic herb with creeping rhizomes, sword shaped leaves and spadix inflorescence. A. calamus grows either as wild or cultivated crop throughout India ascending upto 1800 m in the Himalayas.⁽¹⁾

Acorus calamus is now also found across Europe, in southern Russia, northern Asia Minor, southern Siberia, China, Indonesia, Japan, Burma, Sri Lanka, Australia, as well as southern Canada and the northern United States where it may be confused with diploid Acorus americanus. (The morphological distinction between the Acorus species is made by the number of prominent leaf veins. Acorus calamus has a single prominent midvein and then on both sides slightly raised secondary veins (with a diameter less than half the midvein) and many fine tertiary veins. This makes it clearly distinct from Acorus americanus)⁽²⁾

^{Traditional uses}

In Asia, Sweet flag has been used for at least the last 2000 years. The ancient peoples of China used it to lessen swelling and for constipation. The parts used are leaves, root (rhizome) and stem. In India, Ayurvedic medicinal practice has used the rhizomes to cure several diseases like fever, asthma and bronchitis, and as a sedative.⁽⁴⁾The Sioux used the whole plant, making aromatic garlands from the leaves and using the root as a tea for bowel pains, or rubbed the chewed root on the skin for a general illness cure.⁽⁵⁾

For the Penobscot people this was a very important root. One story goes that a sickness was plaguing the people. A muskrat spirit came to a man in dream, telling him that he (the muskrat) was a root and where to find him. The man awoke, found the root, and made a medicine which cured the people. In Penobscot homes, pieces of the dried root were strung together and hung up for preservation. Steaming it throughout the home was thought to "kill" sickness. While traveling, a piece of root was kept and chewed to ward off illness.The Potawatomi people powdered the dried root and placed this up the nose to cure catarrh.⁽²⁾

Medicinal uses

Sweet flag has a very long history of medicinal use in Chinese and Indian herbal traditions.⁽⁷⁾ It is widely employed in modern herbal medicine as its sedative, laxative, diuretic,and carminativeproperties.⁽³⁾ It is used in Ayurveda to counter the side effects of all hallucinogens.⁽⁸⁾Both roots and leaves of A. calamus have shown antioxidant,⁽¹⁰⁾antimicrobial and insecticidal activities.⁽⁶⁾Acorus Calamus was also known to many early American settlers and used for a number of diseases. Walt Whitman even wrote poetry about his beloved herb in "Leaves of grass".⁽⁹⁾

Active Constituents

Volatile Oil (principal constituents of the Volatile oil are Asamyl alcohol, Eugenol and Asarone), also contains a bitter principle Acorin (Glucoside), Starch and Tannin.⁽¹⁾

The objective of the present study was to establish various Pharmacognostic standards and to evaluate preliminary phytochemical and physicochemical analysis that can facilitate identification and assist in the preparation of monograph of the plant.

MATERIALS AND METHODS:

Plant Material

The dried rhizome of Acorus calamus was collected from local market of Jaipur in the month of October. The shade dried powder was used for the determination of macroscopic, microscopic, physicochemical parameters and phytochemical screening.

Macroscopical studies

The dried rhizome of Acorus calamus were subjected to macroscopical studies which comprised of organoleptic characteristics of the drug viz., size, colour, odour, taste, shape.

Microscopical studies

Qualitative microscopic evaluation was carried out by taking transverse sections of rhizome of Acorus calamus. Free hand section of softened rhizome were boiled with chloral hydrate to remove all the coloring matter and then carefully stained with phloroglucinol and hydrochloric acid. The sections were transferred to mounted (glycerin) on a slide and a cover slip was placed over it. Powder characteristics of Acorus calamus rhizome powder were also studied using reported method.⁽¹³⁾

Physicochemical parameters

Various physicochemical parameters such as total ash, water soluble ash, acid insoluble ash, sulphated ash, water extractable matter, alcohol extractable matter, foreign matter, moisture content, were calculated ^(12,13).

Preliminary phytochemical screening

The aqueous and alcoholic extracts were subjected to qualitative chemical examination for the identification of various plant constituents. Following tests were performed.

Tests for Carbohydrates and Glycosides

200 mg of aqueous extract was dissolved in 5ml of distilled water and this solution was subjected to Molisch test for the detection of carbohydrates. Small portion of the extract was hydrolyzed with dilute hydrochloric acid for few hours in water bath and was subjected to Liberman-Burchard's test, Legal's and Borntager's test to detect the presence of different glycosides. Another small portion of extract was treated with Fehling's reagent, Barfoed reagent to detect the presence of various sugars. For the detection of saponin glycosides, Foam test and Hemolytic tests were carried out.^(12,13).

Tests for protein and free amino acids

A small quantity of alcoholic extract was dissolved in few ml of water and was subjected to Millon's test, Biuret test and Ninhydrine test ^{(12, 13)}.

Tests for phenolic compounds and tannins

Small quantities of alcoholic extracts were treated with 5 % FeCl₃solution, 1 % of gelatin containing 10 % NaCl, 10 % lead acetate and aqueous bromine solution for the detection of phenolic compounds and tannins ^(12,13).

Tests for alkaloids

The small portion of dried alcoholic extract was stirred with a few drops of dilute Hydrochloric acid and was filtered. The filtrate was tested with various alkaloidal reagents such as Mayer's reagent, Dragendroffs reagent, Hager's reagent, Wagner's reagent.^(12,13).

RESULTS:

Macroscopical studies

The macroscopic character was useful in quick identification of plant material and also serves as an important standardization parameter. The Acorus calamus rhizome are brownish in colour with 12-14.5cm in length and 1- 2cm thick, cylindrical and branched shape, aromatic odour, tasteless in taste (Figure.1). The organoleptic evaluation of Acorus calamus rhizome were showed in (Table1).

Figure 1: Dried rhizome of Acorus calamus

Table 1: Organoleptic evalution of Acorus calamus rhizome

Shape Cylindrical and branched

Colour Brownish

Odour Aromatic

Taste Tasteless

Size 12-14.5 cm length 1-2 cm thick

Microscopic study

Figure 2(a)

Figure 2(b)

Figure 2(c)

Figure 2: T.S. of rhizome of Acorus Calamus

Figure 3: Powder Microscopy of Acorus calamus rhizome:- Fibre, Starch grain, Vessel.

PHYSICOCHEMICAL PARAMETER

Table 1: physicochemical parameter of Acorus calamus rhizome

Physicochemical parameters	Observed value	Standard value(API)
Total ash	5%	NMT 7%
water soluble ash	3%
Acid insoluble ash	0.69%	NMT 1%
Sulphated ash	4.13%
Foreign matter	0.25%	NMT 1%
Moisture content(100-105⁰C)	7.2%
Water soluble extractive	25%	NMT 16%
Alcohol soluble extractive	14.3%	NMT 9%

PHYTOCHEMICAL SCREENING

Table 2: phytochemical screening of Acorus calamus

S. No.	Test	Aqueous extract	Ethanolic extract
1.	Alkaloids	positive	Positive
2.	Carbohydrate	positive	Positive
3.	Protein	negative	Negative
4.	Glycoside	negative	Positive
5.	Phenolic compound	positive	Negative
6.	Tannin	negative	Positive

Thin layer chromatography (TLC) of methanolic extract

Extract - methanolic

Solvent system –Toluene : ethylacetate : acetic acid

(5 :4.2 : 0.8)

Standard - Eugenol oil Rf value - 0.61,0.9

DISCUSSION:

The T.S of rhizome shows single layered epidermis; cortex composed of spherical to oblong,thin-walled cells of various sizes, cells towards periphery, smaller, somewhat collenchymatous, fibro-vascular bundles having light yellowish-brown contents, present in this region; endodermis distinct; composed of round,parenchymatous cells. The powder microscopy shows fibre,starch grain and vessel.The extractive values are useful to evaluate the chemical constituents present in the crude drug and also help in estimation of specific constituents soluble in a particular solvent. In the present investigation water-soluble extractive is more when compared to alcohol soluble extractive values. Preliminary phytochemical analysis indicated presence of steroids, phenolic compounds, flavanoids and tannins.

CONCLUSION:

In present investigation various standardization parameters such as macroscopical, microscopical, physicochemical parameters and phytochemical screening of Acorus calamus Linn was carried out. In this scenario, anatomical studies have become mandatory for proper identification. Thus, our study is an important landmark in correct identification of Acorus calamus.

REFERENCES:

1. Ayurvedic Formulary of India, 2nd vol. Part-1.New Delhi; Govt. of India, Ministry of health and family welfare. Department of Indian system of Medicine and Homeopathy;2003. 177-179.

2. Balakumbahan, R.; K. Rajamani and K. Kumanan (2010-12-29). "Acorus calamus: An overview". Journal of Medicinal Plants Research 4 (25): 2740–2745

3. WHO. Quality control methods of medicinal plants materials, Geneva, 1998.

4. Gualtiero Simonetti (1990). Stanley Schuler. ed. Simon & Schuster's Guide to Herbs and Spices. Simon & Schuster.

5. Modern Phytomedicine. Turning medicinal plants into drugs, Published by John Wiley and Sons; 2007: 25-57

6. Srinivasan, D., Nathan, S., Suresh, T., Perumalsamy, O. (2001): Antimicrobial activity of certain Indian medicinal plants used in folkloric medicine. J. Ethnopharmacol. 74:217-220.

7. Erichsen-Brown, Charlotte (1989). Medicinal and Other Uses of North American Plants: A Historical Survey with Special Reference to the Eastern Indian Tribes. Dover Publications. pp. 231–232.

8. Mukherjee P.K., Kumar V., Mal M., Houghton P.J. "Acorus calamus: Scientific validation of ayurvedic tradition from natural resources"Pharmaceutical Biology 2007 45:8 (651-666)

9. Johnson, Derek; Linda Kershaw, Andy MacKinnon, Jim Pojar (1995). Plants of the Western Boreal Forest & Aspen Parkland.Lone Pine Publishing

10. S. Asha Devi; Deepak Ganjewala ,"Antioxidant Activities of Methanolic Extracts of Sweet-Flag (Acorus calamus) Leaves and Rhizomes Journal of Herbs, Spices & Medicinal Plants Volume 17, Issue 1, 2011; 1 – 11

11. Ghosh S, Sharma AK, Kumar S, Tiwari SS, Rastogi S, Srivastava S, Singh M, Kumar R, Paul S, Ray DD, Rawat AK "In vitro and in vivo efficacy of Acorus calamus extract against Rhipicephalus (Boophilus) microplus." Parasitol Res. 2011 Feb;108(2):361-70

12. Khanderwal K.R. Practical Pharmacognostic Techniques and Experiments, 19 ^thed. Pune: Nirali Prakashan; 2008; 149-156.

13. Kokate C, Purohit A, Gokhale S. Practical Pharmacognosy. 10^thed. New delhi, India: Vallabh prakashan; 1994, 112-114.

14. Subha T S, Gnanamani A, Mandal AB,” Pharamcognostic Evaluation of Acorus calamus L.” Phcog J., july 2011,vol 3, 23-27.

Received on 29.01.2012 Accepted on 10.03.2012

Asian J. Pharm. Res. 2(1): Jan.-Mar. 2012; Page 39-42

ABSTRACT: