|Year : 2013 | Volume
| Issue : 1 | Page : 83-86
Peritoneal mast cell stabilization potential of Pothos scandens L.
Saurabh Gupta1, B Duraiswamy1, MN Satishkumar2
1 Department of Pharmacognosy, J.S.S. College of Pharmacy, Ootacamund, Tamil Nadu, India
2 Department of Pharmacology, J.S.S. College of Pharmacy, Ootacamund, Tamil Nadu, India
|Date of Submission||18-Jan-2012|
|Date of Decision||28-Sep-2012|
|Date of Acceptance||29-Oct-2012|
|Date of Web Publication||24-Jan-2013|
Department of Pharmacognosy, J.S.S. College of Pharmacy, Ootacamund, Tamil Nadu
Source of Support: Authors are grateful to JSS College of Pharmacy
and JSS University, Mysore for kind support., Conflict of Interest: None
Objective: To investigate the peritoneal mast cell stabilization activity of Pothos scandens extracts
Materials and Methods: Pothos scandens L. (family- Araceae) aerial part was successively extracted with ethanol and aqueous to prepare extract of the plant. The extracts of P. scandens were evaluated for stabilization of mast cell in rat allergic models. The extract of P. scandens ethanolic, 50% aqueous ethanolic and aqueous (1, 10 and 100 μg/ml) was studied for peritoneal mast cell stabilization activity in rat mesenteric preparation induced by C 48/80.
Result: Preliminary phytochemical analysis revealed the presence of carbohydrates, fixed oil, proteins, alkaloids, glycosides, flavonoids and phenolic compounds. The ethanolic, 50% aqueous ethanolic and aqueous extracts of P. scandens L. showed dose dependent increase in the number of intact cells when compare with C48/80 at the concentration of 10 and 100 μg/ml. It virtues further work towards the isolation of phytoconstituents from this plant.
Conclusion: This finding provides evidence that the P. scandens L. inhibits mast cell-derived immediate-type allergic reactions and mast cell degranulation. P. scandens has a potential as allergic anti- asthmatic agent.
Keywords: Compound 48/80, ethanolic extract, mast cell, Pothos scanden
|How to cite this article:|
Gupta S, Duraiswamy B, Satishkumar M N. Peritoneal mast cell stabilization potential of Pothos scandens L. Indian J Pharmacol 2013;45:83-6
| » Introduction|| |
Pothos scandens L. (family-Araceae) is a climbing shrub having adventitious acrid roots. The leaves are obovate, lanceolate and coriaceous having a bright green colour.
The leaves are traditionally used to treat skin disorders. An ethnobotanical survey carried out among the ethnic groups (Kani/Kanikaran) in Southern Western Ghats of India revealed the use of P. scandens leaf mixed with the fruits of Capsicum annum and rhizome of Allium sativum. The mixture is ground into a paste with coconut oil and applied topically on affected places to heal wounds created during delivery. , Sri Lankan tribal people use leaves of P. scandens to reduce swelling speedily in trauma area.  In China the plants are used as blood coagulant, wounds, tumors and drinking for anti-cough.  In India, the infusion of the leaves of this plant as a bath, is used for curing convulsions and epilepsy. Apart from that, the stem is also reportedly used to treat asthma, after being cut with camphor and smoked like tobacco. 
The previous literature reported that the phytochemical investigation of P. scandens leaf extracts showed the presence of chemical compounds such as alkaloid, catachin, coumarin, tannin, saponin, flavonoid, phenol, sugar, glycoside andxanthoprotein.  The GC-MS analysisof ethanolic extract of P. scandens leaf detected nineteen compounds. The major compounds are dodecanoic acid, tetradecanoic acid, 3, 7, 11, 15-tetramethyl-2-hexadecan-1-ol, n-Hexadecanoic acid, phytol, 9,12-Octadecadienoic acid (Z,Z), 9, 12, 15-octadecatrienoic acid (Z,Z,Z), 1,2-Benzenedicarboxylic acid, diisooctylester. 9,12-octadecadienoic acid (Z,Z)- and 9, 12,15-Octadecatrienoic acid (Z,Z,Z)- have the anti-inflammatory and anti-arthritic property. Among the identified phytochemicals, dodecanoic acid, tetradecanoic acid and n-hexadecanoic acid have the antioxidant property. 
Mast cells are found in the skin and in mucosal tissues at homeostasis and their numbers are elevated in asthmatic lungs and inflammatory bowel disease of gastrointestinal tract. Mast cells were first described by Ehrlich in his 1878 doctoral thesis on the basis of their unique staining characteristics and large granules, that gave them their name, "Mastzellen" which means well-fed cells, because their cytoplasm was stuffed with granular material. Mast cells are now considered to be part of the immune system.  The mast cell was identified as a mesenchymal cell which is stained metachromatically with O-toludine blue and it was recognized several years later that these cells contained in their granules the majority of the body's histamine. Mast cells play a central role in inflammatory and immediate allergic reactions.  The release of potent inflammatory mediators, such as histamine, proteases, chemotactic factors, cytokines and metabolites of arachidonic acid act on the vasculature, smooth muscle, connective tissue, mucous glands and inflammatory cells. Histamine is not only released when the body encounters a toxic substance, but also released when mast cells detect injury.  The present study was undertaken to evaluate the mast cell stabilization property of the extracts of the aerial part of P. scandens on rat mesentery.
| » Material and Methods|| |
P. scandens L. aerial parts were collected in the month of August, 2010, from Tirupati district, Andhra Pradesh, India. Dr. K. Madhava Chetty, Botanist, Department of Botany, Sri Venkateswara University, Tirupati authenticated the collected plant. Voucher specimen has been preserved in our laboratory (SVU/SC/09/25/10-11) for future reference.
Ethanol, NaCl, KCl, CaCl 2 , NaHCO 3 , Dextrose, Xylene, Acetone and Anesthetic ether were purchased from E-Merck (India) Ltd., Mumbai, India. Compound C40/80 and O-toludine blue were purchased from sigma Pvt. Ltd. etc. All chemicals used were of analytical grade.
Healthy male albino rats of wistar strain (180-220 g) were obtained from the animal house, J.S.S. College of Pharmacy, Ootacamund, India, and were maintained under standard environmental conditions (22-28 °C, 60-70% relative humidity,12-h dark:12-h light cycle) and were fed with standard rat feed (M/S Hindustan Lever Ltd., Bangalore, India) and water ad libitum. The experiments were conducted as per the guidelines of CPCSEA, Chennai, India (Approval no. JSSCP/IAEC/Ph.D/P.Cog/02/2011-12).
Preparation of Extracts
The P. scandens aerial part was cleaned thoroughly with water to remove any unwanted matter, dried in shade, ground to a coarse powder with a mechanical grinder and passed through sieve no. 40. Further, it was extracted with cold maceration process using absolute ethanol by intermittent shaking for 10 days, filtered and the dried marcleft was macerated with 50% aqueous ethanolic for 10 days, filtered and again the resultant residue was macerated with aqueous for another 10 days with intermittent shaking. The solvent was removed by distillation under reduced pressure and the resulting semisolid mass was vacuum dried using rotary flash evaporator (Rota vapor, R-210/215, Buchi, Switzerland). The concentrated semi solid material was kept in a desiccator for drying.
Qualitative Phytochemical Screening
The P. scandens L. extracts were subjected for phytochemical tests to find the presence of major phytochemical constituents such as carbohydrates, proteins, amino acids, alkaloids, glycosides, saponins, sterols, flavonoids, phenolic compounds, fixed oil and fat, gum and mucilage, phytosterols and tannins according to standard methods. 
Mast Cell Stabilization Activity
The overnight fasted male Wister rats were sacrificed with excess dose of anesthetic ether. The abdomen was cut open to expose the intestine. Pieces of mesentery with connecting lobes of fat and blood vessels were rapidly dissected out. A small pieces of the mesentery were cut and placed in a beaker containing Ringer Locke (in mM: NaCl 154, KCl 5.6, CaCl 2 2.2, NaHCO 3 6.0 and dextrose 5.5) solution for 30 ± 1 min containing different concentration of plant extracts. Later, the tissues were exposed to compoundC48/80 (C 48/80 at 0.8 μg/ml to promote mast cell degranulation) and the tissue was incubated for further 30 ± 1 min. The pieces of mesentery were removed and placed in a clean slide. Excess fatty layers and adhering small intestine tissues were removed. The trimmed tissue was dipped in 4% formaldehyde solution containing 0.1 % O-toludine blue for 20-30 min and then the tissue was transferred through acetone and then xylene (2 changes each) for 5 ± 1 min. Six pieces of mesentery were used for each concentration of the test substance. The stained mesentery pieces were focused through a digital light microscope (M/s. Motic, Japan) at 100x magnification. 100 mast cells were counted, starting from the left hand side of the field and then proceeding clockwise. The number of intact and fragmented or disrupted mast cells was noted. A mast cell was considered disrupted if 4 or 5 granules were found around the mast cells. The percentage of mast cell fragmented or disrupted and of intact mast cells was calculated. 
The experimental groups for were as follows:
Group I: Vehicle control (tissues exposed to Ringer Locke solution only)
Group II: Negative control (tissues exposed to 0.8 μg/ml of C 48/80 only)
Group III: Positive control (tissues exposed to disodium cromogylate-DSCG, 1 mg/ml)
Group IV: Tissues exposed in ethanolic extract of P. scandens (PSE) 1 μg/ml
Group V: Tissues exposed in ethanolic extract of P. scandens (PSE) 10 μg/ml
Group VI: Tissues exposed in ethanolic extract of P. scandens (PSE) 100 μg/ml
Group VII: Tissues exposed in 50% aqueous ethanolic extract of P. scandens (50% PSE) 1 μg/ml
Group VIII: Tissues exposed in 50% aqueous ethanolic extract of P. scandens (50%PSE) 10 μg/ml
Group VIIII: Tissues exposed in 50% aqueous ethanolic extract of P. scandens (50%PSE) 100 μg/ml
Group X: Tissues exposed in aqueous extract of P. scandens aqueous (PSA) 1 μg/ml
Group XI: Tissues exposed in aqueous extract of P. scandens aqueous (PSA) 10 μg/ml
Group XII: Tissues exposed in aqueous extract of P. scandens aqueous (PSA) 100 μg/ml
All the groups except group I and II were later exposed to 0.8 μg/ml of C 48/80.
Statistical analysis was done by using one-way Analysis of Variance (ANOVA) followed by Turkey's multiple comparison tests. P value < 0.05 was considered to be statistically significant. The analysis was carried out using Graph Pad Prism software V.5.04.
| » Results|| |
Extract Recovery Percent
The maximum yield was obtained in 50% aqueous ethanolic compared to other solvents used. The extractive values were found to be: 2.98, 9.73 and 5.39% w/w for ethanolic, 50% aqueous ethanolic and aqueous respectively.
Qualitative Phytochemical Screening
The phytochemical screening on P. scandens revealed the presence of primary metabolites such as, carbohydrates, fixed oil and proteins the secondary metabolites such as alkaloids, glycosides, flavonoids and phenolic compounds [Table 1].
Mast Cell Stabilization Activity
Compound 48/80, a known mast cell degranulation agent, produced a significant (P < 0.001) reduction in intact mesenteric mast cells, 16.8 ± 1.5, when compared to the mesentery exposed to the vehicle, Ringer Locke's solution, alone 83.2 ± 2.4. At the concentration of 10 and 100 μg/ml, ethanolic, 50% aqueous ethanolic and aqueous extracts of P. scandens produced dose dependent and showed significant (P < 0.001) increase in the number of intact cells when compared with C48/80 treated tissues [Figure 1]. Among the different extracts 50% aqueous ethanolic 10 μg/ml (52.2 ± 4.9) and 100 μg/ml (67.8 ± 2.9) showed significant protection of cell as compared other two extracts. The mast cell stabilization activity of different extracts was as follows; 50% aqueous ethanolic > aqueous > ethanolic.
|Figure 1: Effect of P. scandens extracts on mast cell degranulation induced by compound 48/80 in the rat mesentery. # P < 0.001 vs. control; *P < 0.001 vs. C 48/80; One way ANOVA followed by Tukey's multiple comparison test (F = 78.88; df = 71; P < 0.001). DSSGdisodium cromoglycate 1mg/mL, P. scandens extract (ethanolic-PSE, 50% aqueous ethanolic -50% PSE and aqueous-PSA)|
Click here to view
| » Discussion|| |
It is well known that compound 48/80 (acondensation product of N-methyl-p methoxyphenethylamine with formaldehyde) has the potential to secrete allergy-related factors from mast cells as stimulators.  The compound C48/80 has been used to study allergies and anaphylaxis, because it can vigorously activate the release of histamine via the mechanism of cellular exocytosis. Several flavonoids have been shown to possess smooth muscle relaxant and bronchodilator activity.  The phytochemical screening on P. scandens revealed the presence of such as alkaloids, flavonoids and phenolic compounds. Flavonoids also inhibited the histamine release induced by C48/80.  The various phytochemical compounds are present in plant like dodecanoic acid, tetradecanoic acid, 3, 7, 11, 15-tetramethyl-2-hexadecan-1-ol, 1,2-Benzenedicarboxylic acid, diisooctylester.9,12- octadecadienoic acid (Z,Z)- and 9, 12,15 Octadecatrienoic acid (Z,Z,Z)- have the anti-inflammatory property. Among the identified phytochemicals, dodecanoic acid, tetradecanoic acid and n-hexadecanoic acid have the antioxidant property.  Based on present investigation, it can be concluded that P. scandens stabilizes mast cells in rat mesenteric tissue. As the mast cell play a major role in Type I hypersensitivity-mediated diseases like allergic asthma and rhinitis.  This could be attributed due to the presence of rich chemical constituents in P. scandens.
| » Conclusion|| |
This finding provides evidence that the P. scandens L. inhibits mast cell derived immediate type allergic reactions and mast cell degranulation. P. scandens will be the potential candidate for allergic anti-asthmatic activity. Studies are underway to evaluate the efficacy of P. scandens due to its mast stabilization property in these animal allergic models.
| » References|| |
|1.||Haneefa MK, Hanan SK, Saraswathi R, Guru MP, Nayar C. Formulation and evaluation of herbal gel of P. scandens Linn. Asia Pac J Trop Med 2010;3:988-92. |
|2.||Ayyanar M, Ignacimuthu S. Traditional knowledge of Kanitribals in Kouthalai of Tirunelveli hills, Tamil Nadu, India. J Ethnopharmacol 2005;102:246-55. |
|3.||Ayyanar M, Ignacimuthu S. Herbal medicines for wound healing among tribal people in Southern India: Ethnobotanical and Scientific evidences. Int J Applied Res Nat Prod 2009;3:29-42. |
|4.||Ediriweeraa ER, Grerub DD. Traditional medical practices of Sri Lanka in orthopaedic treatment. AYU 2009;30:147-52. |
|5.||Boyce PC. A review of Pothos L. (Araceae: Pothoideae: Pothoeae) for Thailand. Thai Forest Bull (Bot) 2009;37:15-26. |
|6.||Bhandary MJ, Chandrashekar KR, Kaveriappa KM. Medical ethanobotany of Siddis of Uttara Kannada district, Karnataka, India. J Ethnopharmacol 1995;47:149-58. |
|7.||Lalitharani, Mohan VR, Maruthupandian A. Pharmacognostical and phytochemical studies on P. scandens L. Int J Phytomed 2010;2:277-83. |
|8.||Lalitharani S, Mohan VR, Regini GS, Kalidass C. GC-MS analysis of ethanolic extract of P. scandens leaf. J Herb Med Toxicol 2009;2:159-60. |
|9.||Kawa A. The role of mast cells in allergic inflammation. Respir Med 2012;106:9-14. |
|10.||Peachell P. Targeting the mast cell in asthma. Curr Opin Pharmacol 2005;5:251-6. |
|11.||Skidgel RA, Erdos EG. Histamine, bradykinin and their antagonists In: Brunton LL, Lazo JS, Parker KL, editors. Goodman and Gilman's: The Pharmacological Basis of Therapeutics. 11 th ed. New York: McGraw Hill Companies, Inc; 2006. p. 629-5. |
|12.||Kokate CK. Practical pharmacognosy, 2 nd ed, New Delhi: Vallabhprakasham; 1988. p. 142-59. |
|13.||Norton S. Quantitative determination of mast cell fragmentation by compound 48/80. Br J Pharmacol 1954;9:494-7. |
|14.||Allansmith MR, Baird RS, Ross RN, Barney NP, Bloch KJ. Ocular anaphylaxis induced in the rat by topical application of compound 48/80. Dose response and time course study. Acta Ophthalmol Suppl 1989;192:145-53. |
|15.||Hazekamp A, Verpoorte R, Panthong A. Isolation of bronchodilator flavonoid from the Thia medicinal plant Clerodendrumpetasites. J Ethnopharmacol 2001;78:45-9. |
|16.||Bellanti JA. Mechanism of tissue injury produced by immunologic reactions. Immunology. Asian ed. Tokyo: W.B. Saunders Co; 1971. p. 184. |