PHARMACOGNOSTIC, PHYTOCHEMICAL, AND PHARMACOLOGICAL INVESTIGATION ON BARK OF THUJA ORIENTALIS LINN (CUPRESSACEAE)

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1 wjpmr, 2017,3(3), SJIF Impact Factor: WORLD JOURNAL OF PHARMACEUTICAL AND MEDICAL RESEARCH Research Article ISSN WJPMR PHARMACOGNOSTIC, PHYTOCHEMICAL, AND PHARMACOLOGICAL INVESTIGATION ON BARK OF THUJA ORIENTALIS LINN (CUPRESSACEAE) Malode Sunanda* 1, Bodile Shubhangi 2, Niranjan Bhadakwade 2, Omkar Shikare 2, More Akshaya 2 and Malode Manisha 2 1 Department of Pharmacognosy, M. V. P. Samaja s College of Pharmacy, Gangapur Road, Nashik , Maharashtra, India. 2 Kasturi Shikshan Santha s College of Pharmacy, Shikrapur, Pune. *Corresponding Author: Malode Sunanda Department of Pharmacognosy, M. V. P. Samaja s College of Pharmacy, Gangapur Road, Nashik , Maharashtra, India. Article Received on 23/02/2017 Article Revised on 16/03/2017 Article Accepted on 05/04/2017 ABSTRACT The present research was conducted to investigate the analgesic activity of Ethyl acetate soluble alcoholic (ETA) extracts of plants of Thuja orientalis. (Family: Cupressaceae) The analgesic activity of the ethyl acetate soluble alcoholic extracts of the Thuja orientalis at the dose of 100mg/kg body weight was evaluated against the standard drug Pentazocineat a dose of 10 mg/kg body weight. Adult swiss albino mice of either sex of six numbers in each group, was undertaken for study and evaluated by Hot Plate Method and Tail Immersion Test. The Ethyl acetate soluble alcoholic extracts of plants of Thuja orientalis showed greater analgesic activity when compare with the standard drug. Result of present studies suggest that ETA extract of Thuja orientalis significant (P value < 0.001) analgesic activity. KEYWORDS: Thuja orientalis, Analgesic Activity, Hot Plate Method, Tail Immersion Test. INTRODUCTION Thuja orientalis is a common ornamental evergreen tree that is originally native to Northwest China belonging to family Cupressaceae. It is highly aromatic and resinous shrub that widely cultivated in gardens located in temperate and semi-temperate areas. [13] Thuja orientalis is hardy, large evergreen shrub or small to medium sized-tree rarely exceeding 20 m in nature. It has a dense, pyramidal shape, but often exhibits a more open and spreading form. It prefers moist, well-drained soil and full sun. The bark is gray with brown highlights and has thin but deep furrows. The bark has a rugged charm about it, especially on large mature specimens. Younger bark is a reddish-brown color and exfoliates in long, thin strips. [10] It is endemic to northwestern China. It is also now naturalized as an introduced species elsewhere in Asia: eastward to Korea and Japan; southward to northern India; and westward to northern Iran. The common name 'arbor-vitae' is from Latin, 'Tree of life', and is based on its association with long life and vitality in Buddhist thought in China. This is probably based on the tree's unchanging evergreen nature in the cold dry climate of northwest China, and its longevity; some of the larger specimens planted around Buddhist temples in China are said to be in excess of 1,000 years old. [11] Pain is a symptom of many diseases requiring treatment with analgesics. Pain is a disabling accompaniment of many medical conditions and pain control is one of the most important therapeutic priorities. 1 Pain has been officially defined as an unpleasant sensory and emotional experience associated with actual or potential tissue damage. It is always a warning signal and primarily protective in nature but often causes a lot of discomfort and lead to many adverse effects. 2 Analgesics are drugs used to treat or reduce pain and the classical analgesic drugs notably opiates and nonsteroidal anti-inflammatory drugs have their origin in natural products but many synthetic compounds that act by the same mechanism have been developed and are associated with serious adverse effects such as ulceration, gastrointestinal bleeding, additive potential, respiratory distress, drowsiness, nausea etc. [7] MATERIALS AND METHODS Collection and identification of plant material The fresh plants of Thuja orientalis Linn were collected in the months of July-August from Hadapsar, Pune, Maharashtra, India, and authenticated by P. Laksharasimhan, Scientist E and HOD, Botanical Survey of India, Pune

2 Preparation of extracts The bark of Thuja orientalis was collected, washed and dried at room temperature. Bark were grinded into the fine powder, extracted with different solvents in decreasing order of solvent polarity i,e n- Hexane, Chloroform, Alcohol and Ethyl acetate. The extract was dried in a vacuum oven to obtained constant weight. Phytochemical evaluation The Ethyl acetate soluble alcoholic extract was used to analyze qualitatively various phytoconstituents such as alkaloids, glycosides, steroids, phenolic compounds, tannins, flavonoids and carotenoids using standard procedures. [7] Table 1: Characterization of extracts by Chemical Tests. Sr. No. Chemical Test Bark Extract n-hexane Chloroform Alcohol 1 Test for sterol Present Test for Glycosides - - Present 3 Test for Alkaloids - Present - 4 Test for Flavonoids - - Present 5 Test for Tannins - - Present 6 Test for Carotenoids Present - - Analgesic activity Chemical and extract Pentazocine, Carboxy methylcellulose (CMC), n- Hexane, Ethyl acetate soluble alcoholic extracts of Thuja orientalis bark Animal used Studies were conducted in pharmacology laboratory using male mice gm. The mice were grouped and housed in polyacrylic cages and maintain under standard laboratory condition (temperature ±20 ᵒc) with dark/ light cycle (14/10 hrs). They were allowed free access to standard dry pellet diet and water ad. libitum. The animals were acclimatized to laboratory condition for 10 days before commencement of the experiment. Drug adistration Preparation of control 0.5% w/v Carboxy methylcellulose solution was adistered orally (1mg/kg) to the Group I. Preparation of Standard group The Group II animals received the Standard drug Pentazocine (10mg/kg), intraperitoneally. Preparation of Test group A dose of 100 mg /kg body weight of extracts (n- Hexane, ETA) of Thuja orientalis bark was adistered orally to groups III, and IV of animals respectively. 1) Eddy s Hot Plate Method Principle The paws of mice are very sensitive to heat at temperatures which are not damaging the skin. The responses are jumping, withdrawal of the and licking of the paws. The time until these responses occur is prolonged after adistration of centrally acting analgesics, whereas peripheral analgesics do not generally affect these responses. [13] Procedure Swiss albino mice (20-30 gm body weight) are used. They were randomly selected and divided into four groups designated as group-i, group-ii, group-iii and group-iv consisting of six mice in each group for Control, Standard and Test sample group respectively. Each group received a particular treatment i.e. Control (0.5 % CMC solution in water, 1mg/kg, orally), Standard (Pentazocine, 10 mg/kg, intraperitoneally) and Test sample (n-hexane and Ethyl acetate soluble alcoholic extract 100 mg/keg, orally respectively). The animals were positioned on Eddy s hot plate kept at a temperature of 55 to 56 0 C. The reaction time until either licking or jumping occurs was recorded by a stopwatch. Preparation of Test drug The Carboxy methylcellulose of 0.5% w/v was prepared by dissolving the 0.5gm Carboxy methylcellulose in 100ml water. The 0.5% w/v solution of Carboxy methylcellulose was used as surfactant for dissolving the test drug. It was used for preparing varying doses of test drug. 1gm of each extract was dissolved in 50ml of 0.5% w/v Carboxy methylcellulose to get the dose of 100mg/kg body weight to prepare 5ml dose

3 R e a c tio n T im e (s e c ) Table 2: Observation table of Analgesic activity of Thuja orientalis bark extract by Hot Plate Method. Groups Drug Animals Group I Control Vehicle (1mg/kg) 6 Group II Standard Pentazocine (10mg/kg) 6 Group III Test I n-hexane (100 mg/kg) 6 Group IIV Test II Ethyl acetate soluble alcoholic extract (100 mg/kg) 6 Table 3: Analgesic activity of Thuja orientalis bark extract by Hot Plate Method Sr. no Groups 1 Control 2 Standard 3 n- Hexane extract 4 Ethyl acetate soluble alcoholic extract Reaction Time (sec) Min Mean SEM SD Mean SEM SD Mean SEM SD Mean SEM SD * C o n tr o l s ta n d a r d n -H e x a n e E T A T im e (m in ) ** ** C o n tro l s ta n d a rd n -H exane E T A Figure 1: Analgesic activity of Thuja orientalis bark extract by Hot Plate method. Statistical analysis The results were analyzed for statistical significance using one way ANOVA followed by Dunnet test. A P value < was significant. tail-withdrawal reflex in rats induced by immersing the end of the tail in warm water of 55 C. [5] Procedure Swiss albino mice (20-30 gm body weight) were used. They were placed into individual restraining cages leaving the tail hanging out freely. The animals were allowed to adapt to the cages for 30 before testing. The lower 5 cm portion of the tail was marked. This part of the tail is immersed in a cup of freshly filled water of exactly 55 C. Within a few seconds the rat reacts by withdrawing the tail. The reaction time was recorded in 0.5 s units by a stopwatch. After each deteration the tail was carefully dried. The reaction time was detered before and periodically after either oral or subcutaneous adistration of the test substance, e.g., after 0.5, 1, 2, 3, 4 and 6 h. The cut off time of the immersion is 15 s. The withdrawal time of untreated animals was between 1 and 5.5 s. A withdrawal time of more than 6 s therefore is regarded as a positive response. 2) Tail immersion test Principle The method has been developed to be selective for morphine- like compounds. The procedure is based on the observation that morphine-like drugs are selectively capable of prolonging the reaction time of the typical 247

4 R e a c tio n T im e (s e c ) Table 4: Analgesic activity of Thuja orientalis bark extract by Tail Immersion Test. Sr.no. Groups 1 Control 2 Standard 3 n- Hexane 4 Ethyl acetate soluble alcoholic extract Reaction Time (sec) hr 2 Hr 4 hr Mean SEM SD Mean SEM SD Mean SEM SD Mean SEM SD * * *** C o n tro l s ta n d a rd n -H exane E T A CONCLUSION From all the experiments done on bark of Thuja orientalis. It is concluded that the Thuja orientalis plant shows presence of sterols, carotenoids, alkaloids, flavonoids, tannins. The phytochemical studies showed the presence of most of the biologically active compounds in the plant. 8(17), 13- Labdine- 16, 15 olide-18-atewas foundas compound in ethyl acetate soluble alcoholic extract. 0 C o n tr o l s ta n d a r d n -H e x a n e E T A T im e (m in ) Figure 2: Analgesic activity of Thuja orientalis bark extract by Tail immersion. Statistical analysis The results were analyzed for statistical significance using one way ANOVA followed by Dunnet test. A P value < was significant. RESULTS AND DISCUSSION Ethyl acetate soluble alcoholic extract of bark of Thuja orientalis was screened for Analgesic Activity. The fraction isolated from ethyl acetate soluble alcoholic extract of bark is 8(17), 13- Labdine- 16, 15 olide-18-ate. The analgesic activity of extract of bark of Thuja orientalis was performed. Analgesic activity of the extract was compared with the standard drug pentazocine. Results were analyzed for statistical significance with help of one-way ANOVA followed by Dunnet test. A P value < was significant. Analgesic activity of ETA extract is significant as compared to standard pentazocine. ETA and n-hexane extract have same analgesic activity in Hot plate method. ETA extract have more analgesic activity than n-hexane extract in Tail immersion test. Ethyl acetate soluble alcoholic extract of Thuja orientalis bark was found to be significant in Analgesic activity. ACKNOWLEDGMENT Author is thankful to all dear Friends who involved in research work and also thankful to M.V.P.Samaja s College of Pharmacy, Nashikand Kasturi Shikshan Santha s College of Pharmacy, Shikrapur, Punefor giving continuous support for doing this work. REFERENCES 1. Khandelwal K.P., Practical Pharmacognosy, 10 th Edition, Nirali Prakashan, Pune, 2003; Pg. no Kadbadi S.S., Deore S.L. and Baviskar B.A., Experimental Phytopharmacognosy, Nirali Publication, Pune, 2011; Pg. no. 4.8, 4.9, Mukherjee Pulok K., Quality Control of Herbal Drugs: An Approach to Evaluation of Botanicals.1 st edition, Business Horizons Publication, New Delhi, 2002; Pg. no Harborn J.B., Phytochemical Method A Guide to Modern Techniques of Plant Analysis, 3 rd Edition, Chapman and Hall, 2005; Pg. no , 74-90, Vogel H.G, Drug Discovery and Evaluation Pharmacology assays, Spinger publication, 2002; Pg. no 669, ,

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