INTRODUCTION:
Since the time in immemorial, our traditional system of medicine and folklore claming that medicinal plants as a whole or their parts are being used in all types of skin diseases successfully including antibacterial and antifungal. As we know very well, nowadays the medicinal preparation available in the market from which most of them either not effective up to the mark or has to develop resistance resulting in reoccurrence again. Plant derived drug serve as a prototype to develop more effective and less toxic medicines1.
Ougeinia oojeinensis (Roxb.) Hochr (Fabaceae) known in Hindi as Tinsa and in Sanskrit as Ratha is a deciduous trees , found in the outer Himalayas and sub-Himalayan tracts from Jammu to Bhutan up to an altitude of 1500m and extending through the whole of northern and central India into the greater part of Deccan peninsula2,3. The extract of the whole plant Ougeinia oojeinensis were scientifically evaluated for analgesic activity4. The 50% of ethanolic extract of stem bark showed antispasmodic action. Phytochemical investigated of O. oojeinensis showed the presence of lupeol, hydroxlupeol, betulin and isoflavanones such as dalbergioidin, homoferreirin and ougenin5-7. A survey of literature showed that no systematic approach has been made to study anti-inflammatory activity in this plant by in-vitro method. The present study is an attempt to study the anti-inflammatory activity of O. oojeinensis using ethanolic and aqueous extracts.
Plant material
The bark of O. oojeinensis were collected from the Lamber forest Raipur, Chhattisgarh in the month of May. The collected material was authenticated by Dr. P. Jayaraman, Botanist, Plant Anatomy Research Centre (PARC), Chennai. The plant was also compared with herbarium specimen maintained at Minor Forest Produce (Trading and Development) Co-Op. Fed. Ltd., Shankar Nagar, Raipur, Chhattisgarh, by Expert Medicinal Plant, Mr. S. N. Khotele.
Preparation of extract
Dried powder of bark was exhaustively extracted successively in soxhlet apparatus, using petroleum ether, choloroform, ethylacetate, ethanol and distilled water repectively. The extracts were then made to powder by using rotary evaporator under reduced pressure. Bark of O. oojeinensis yielded 0.62%, 0.45%, 0.57%, 4.50% and 3.7% w/w powdered extract with petroleum ether, choloroform, ethylacetate, ethanol and distilled water repectively.
Anti-inlammatory activity
The HRBC membrane stabilization has been used as method to study the anti-inflammatory activity. Blood was collected from healthy volunteers. The collected blood was mixed with equal volume of sterilized Alsever solution (2% dextrose, 0.8% sodium citrate, 0.5% citric acid and 0.42% sodium chloride in water).
The blood was centrifuged at 3000 rpm and packed cell were washed with isosaline (0.85%, pH 7.2) and a 10 %( v/v) suspension was made with isosaline.
The assay mixture contained the drug (concentration as mentioned in Table 1), 1 ml of phosphate buffer (0.15M, pH 7.4), 2 ml of hyposaline (0.36%) and 0.5 ml of HRBC suspension. Diclofenac was used as the reference drug. Instead of hyposaline 2 ml of distilled water was used in the control. All the assay mixture were incubated at 37°C for 30 min and centrifuged. The hemoglobin content in the supernatant solution was estimated using spectrophotometer at 560 nm. The percentage hemolysis was calculated by assuming the hemolysis produced in the presence of distilled water as 100%. The percentage of HRBC membrane stabilization or protection was calculated using the formula 8-11.
% Protection = 100- O. D. of drug treated sample
O. D. of control * 100
RESULT AND DISCUSSION:
The lysosomal enzymes released during inflammation produced a variety of disorders. The extracellular activity of these enzymes is said to be related to acute or chronic inflammation. The diclofenac drugs act either by inhibiting these lysosomal enzymes or by stabilizing the lysosomal membrane.
Since HRBC membrane is similar to lysosomal membrane components, the prevention of hypotonicity-induced HRBC membrane lysis is taken as a measure of anti-inflammatory activity of drugs. The results are reported in Table 1. Both the extracts of the bark of O. oojeinensis showed biphasic effects on HRBC membrane stabilization. They increasing activity at low concentration levels but decreasing activity with high concentrations. They have a critical concentration (50 mg/ml) at which their activities are maximum. The activities of both extracts are comparable to that of Diclofenac at concentration of 50 mg/ml. The variation of activity with time was studied at 10 mg/ml concentration, the activities in general decreased with time (Table 2).
Further work is in progress to isolate and identity the compounds responsible for activity.
Table. 1
Effect of concentration on the activity of both extracts of O. oojeinensis
|
Concentration (mg/ml) |
Activity (prevention of lysis %) |
||
|
Ethanolic extract |
Aqueous extract |
Diclofenac |
|
|
10 |
42.12±0.35 |
38.01±0.51 |
------ |
|
50 |
51.23±1.01 |
48.36±0.41 |
52.22±0.05 |
|
100 |
35.13±0.03 |
34.65±0.21 |
------ |
|
200 |
30.32±0.02 |
31.31±0.18 |
----- |
Data are expressed as mean ± S.E., n = 6
Table. 2
Variation of activity with time (drug concentration 10 mg/ml in all)
|
Concentration (mg/ml) |
Activity (prevention of lysis %) |
||
|
Ethanolic extract |
Aqueous extract |
Diclofenac |
|
|
10 |
54.36±0.23 |
49.53±0.12 |
68.25±0.36 |
|
20 |
44.12±0.05 |
43.25±1.02 |
64.20±0.04 |
|
50 |
36.34±0.10 |
38.12±0.24 |
59.31±0.13 |
|
100 |
34.78±0.03 |
35.35±0.03 |
51.63±0.52 |
|
200 |
32.36±1.01 |
31.46±0.56 |
48.16±0.02 |
Data are expressed as mean ± S.E., n = 6
REFERENCES:
1. Kothari A, Shrivastava N. Indian Drugs 2005; 42(3): 133-135.
2. Kirtikar and Basu. Indian Medicinal Plant. Vol. I: 1998. Dehradun, p. 756.
3. Anonymous. The Wealth of India. Raw Material, Vol. VII: C.S.I.R. New Delhi, 1997. p. 193 –197.
4. Khare CP. Encyclopedia of Indian Medicinal Plant. Springer, 2004. p. 343.
5. Mukherjee DK, Barua AK, Bose PK. Chemical Investigation of Ougeinia dalbergioides Benth. Science and Culture 1963; 29: 151–152.
6. Ghosh AC, Dutta NL. Chemical Investigation of Ougeinia dalbergioides Benth. Journal of Indian Chemical Society 1965; 42(12): 831– 835.
7. Balakrishna S, Ramanathan JD, Seshadri TR, FRS, Venkataramani B. Special Chemical Components of the Heartwood of Ougeinia dalbergioides Benth. Proc. Royal Society London 1962; 268A: 1.
8. Gandhidasan R, Thamarraichelvan A, Baburaj S. Anti-inflammatory action of lannea coromandelica by HRBC membrane stabilization. Fitoterapia 1991; LXII: No. 1: 81-83.
9. Seeman P, Weinstein J. Biochem. Pharmacol 1968; 17: 269.
10. Goodman, Gilman. The Pharmacological basis of therapeutics: 6th edition, MacMillan Publishig company, New York:1982.
11. Geisler AJ, Bekemeier H, Hirschelmann R, Bakathir HA. Pharmacology, Biochemistry and Immunology of Inflammatory Reaction:Bekemeier H. Hirschelmann R. (Eds.), Martin Luther University: p. 682.
Received on 12.03.2008 Modified on 15.03.2008
Accepted on 29.03.2008 © RJPT All right reserved
Research J. Pharm. and Tech. 1(1): Jan.-Mar. 2008; Page 57-58