IN VIVO ANTIINFLAMMATORY EFFECT OF EMU OIL (DROMAIS NOVAEHOLLANDIAE) AND VIRGIN COCONUT OIL (COCOS NUCIFERA) ON PHORBOL ESTER INDUCED ACUTE INFLAMMATORY MODEL

Received on 15 October, 2013; received in revised form, 25 November, 2013; accepted, 11 February, 2014; published 01 March, 2014

IN VIVO ANTIINFLAMMATORY EFFECT OF EMU OIL (DROMAIS NOVAEHOLLANDIAE) AND VIRGIN COCONUT OIL (COCOS NUCIFERA) ON PHORBOL ESTER INDUCED ACUTE INFLAMMATORY MODEL

P. Logeswari*¹, V. Dinesh Kumar ¹, P.T.A. Usha ¹ and Prathap Kumar ²

Department of Veterinary Pharmacology and Toxicology, College of Veterinary and Animal Sciences ¹, Mannuthy, Thrissur- 680 651, Kerala, India

Department of Safety Assessment, Advinus Therapeutics Ltd. ², Peenya Industrial Area, Bangalore- 560058, Karnataka, India

ABSTRACT: Inflammation is a cellular reaction to injury and plays a central role in pathophysiology of a variety of diseases.Oil derived from Emu (Dromais novaehollandiae) fat, is reported to have healing property. The present investigation was performed to demonstrate in vivo anti-inflammatory effect of Emu oil using phorbol ester induced acute inflammatory ear edema model. Also, the efficacy of Emu oil was compared with Virgin Coconut Oil extracted from coconut (Cocos nucifera) which is traditionally being used in India. Acute inflammation was induced applying 20 μl of 2% Croton oil (a phorbol ester) in Swiss Albino mice inner pinna. After 2 hours, ears were treated with single application of 20 μl of Emu oil and Virgin Coconut oil. Pinna thickness was measured pretreatment and 6 and 24 hours of post treatment. After 24 hours, animals were euthanized and ear punches were collected and weighed. Peak edema was evidenced at 2 hours of croton oil application. Emu oil and Virgin Coconut oil significantly attenuated croton oil induced inflammation. Inhibition of edema was observed starting from 2 hours of post treatment and Emu oil exhibited peak reduction than Virgin Coconut oil with the maximum efficacy at 6 hours post treatment. Virgin Coconut oil showed maximum reduction around 24 hours post treatment. There were no significant differences between weights of ear punches between treatments. Our findings support in-vivo anti-inflammatory effect of Emu oil and Virgin coconut oil and also validate their use in treating local inflammatory conditions.

Emu oil, Virgin coconut oil, phorbol ester, pinna thickness, ear punch

INTRODUCTION:Inflammation is an integral part of the complex biological local response of living tissue to harmful stimuli and a body defense shot in order to get rid of stimuli. Acute inflammation is of short duration which signifies the early response of body and resolve quickly and is generally followed by healing.

An escalation in release of pro-inflammatory cytokines, including interleukins, tumor necrosis factor, transcription factor, interferon, are bring about peripheral inflammation ¹.

Clinically available steroidal and non-steroidal anti-inflammatory (NSAID) drugs are demonstrating undesired side effects and have their own limitations ^{2 and 3}. Thus, it has become indispensible to revisit the importance of the traditional remedies.

Emu (Dromais novaehollandiae), flightless bird indigenous to Australia and its fat (adipose tissue) being used topically by Australian aboriginals since ages to treat chronic musculoskeletal disorders⁴. Oil extracted from subcutaneous and retroperitoneal adipose tissues by maceration and series of filtration processes ⁵. Prime component of emu oil is fatty acids which includes oleic acid (43–46 %), linoleic acid (9.6 %), palmitic acid (23.5 %) and linolenic acid (0.6 %) ⁴. It is reported to have natural antioxidants such as carotenoids and flavones ¹¹. Modern investigations have been focused on the protective effects of emu oil on adjuvant induced arthritis ⁶, dermal inflammation ^{7 and 8}, wound healing ⁹ and chemotherapy induced inflammation ¹⁸. Emu oil shown to have cosmetic pharmaceutical property as a moisturizer ¹⁰ and orally active in cancer chemotherapy induced mucositis ¹¹.

Virgin coconut oil is obtained from fresh coconut (Cocos nucifera) without chemical or high temperature process. Wet processing of coconut meal, reported to preserve the beneficial unsaponifiable components to maximum extend ¹². Virgin coconut oil is reported to have beneficial therapeutic effects including wound healing ¹³ and anti-inflammatory property ¹⁴.

In this present study, we evaluated the effect of Emu oil on acute ear inflammation induced by croton oil. Croton oil contains 12-o-tetracanoilphorbol-13-acetate, a phorbol ester that elicits inflammatory reaction ¹⁵. Croton oil induces inflammation by release of moderate amount of inflammatory eicosanoids mediated by by cyclooxygenase and 5- lipoxygenase enzymes along with primary pro-inflammatory mediators ¹⁵. So, drug Diclofenac sodium topical gel, a cyclooxygenase inhibitor employed as reference drug. Also, the efficacy of Emu oil was compared with Virgin coconut oil.

The study was conducted in 24 Swiss Albino mice of either sex, weighing 15-20 g. Animals were procured from Small Animal Breeding Station, College of Veterinary and Animal Sciences, Thrissur, Kerala. The animals were housed in appropriate clean polypropylene cages in a well-ventilated room with temperature (21-24° C), relative humidity (65-68 % ) with 12 hours daylight and 12 hours dark cycle. All animals were maintained under identical feeding and management practices in the laboratory.

All procedures involving the use of experimental animals were in accordance to the committee for the purpose of control and supervision of experiments on Animals (CPCSEA) and were approved by the Institutional Animal Ethics Committee (No. Acad [3] 6554/ 04).

Emu oil was procured from Tamilnadu Veterinary and Animal Sciences University, Tamilnadu, India. Virgin Coconut oil was purchased from Patson International, Mumbai, India and Croton oil was purchased from Otto Chemie Pvt Ltd, Mumbai, India.

The animals were randomized into four groups comprising of six animals in each group were treated as indicated in below in table 1.

TABLE 1: EXPERIMENTAL DESIGN WITH TREATMENT DETAILS

Anti-inflammatory activity was measured by inducing acute inflammation using croton oil ¹⁶. Pinna thickness was recorded initially just before the induction of inflammation using an electronic digital micrometer. Inflammation was induced at 10 a.m. to avoid the influence of circadian disparity in corticosteroid levels on the inflammatory responses. Croton oil 2% was prepared by adding 30μl of croton oil in 1500 μl of acetone solution.

Twenty μl of 2% Croton oil was applied on right inner pinna of all animals. Left pinna was smeared with acetone and kept as uninflammed control. Group I animals were served as croton oil control and left untreated. After 2 hours of induced inflammation group II and III animals were topically treated with 20 μl of Emu oil and Virgin Coconut oil respectively.

Group IV animals were treated with diclofenac gel. The Pinna thickness was measured at 6 and 24 hours after treatment. All mice were euthanized by cervical dislocation, after 24 hours and ear punches of 6 mm diameter were taken using biopsy punch and weighed.

The experimental data obtained were analyzed using SPSS 17.0 software (Statistical Package for Social Sciences) by two-way ANOVA test to determine the significant difference among experimental groups. Results were expressed as mean ± standard error. The value of P ≤ 0.05 was considered statistically significant.

Inflammatory reactions such as increases vascular permeability and edema were set around 2 hours of croton oil application. The treatment groups (Emu oil, Virgin coconut oil and Diclofenac gel) shown significant reduction in inflammation when compared to inflammatory control. Animals treated with emu oil evidenced reduced pinna thickness when compared to animals treated with Virgin coconut oil and Diclofenac gel.

Efficacy of emu oil was high at 6 hours of post application from then swelling reduction rate was maintained same [Figure 1]. This might be due to action of emu oil on lessening the pro-inflammatory mediators. Topically applied emu oil has been shown to reduce the levels of TNFα and other pro inflammatory cytokines ¹⁷. Repeated application of emu oil might be needed to prolong the anti-inflammatory effect beyond 6 hours. Our study is in consistent with previous study which concluded that emu oil has more anti-inflammatory property than flax and olive oil in CD-I mice ⁷.

However, Virgin coconut oil found to have moderate effect after 6 hours and further reduction in magnitude of swelling was noticed till 24 hours but not higher than the effect of emu oil. Similar finding has been reported using Virgin coconut oil in ethyl phenyl propiolate-induced ear edema in rats ¹⁴.

After 24 hours, collected ear punches from inflamed and treated pinna (right pinna) were weighed more when compared to uninflammed (left pinna) ear punches which are due to inflammatory response.

There was no significant difference in ear punch weight between treated groups [Figure 2]. This may be explained by the fact that normal body defense to resolves inflammation which induced for short duration.

FIGURE 1:  PINNA THICKNESS (mm) IN DIFFERENT TIME PERIODS

FIGURE 2: EAR PUNCH WEIGHT (G) AT 24 HOURS Right- Inflamed and Treated Pinna; Left – Un-inflamed Pinna

Hence, the results of the current preliminary study demonstrate that single application of Emu oil and Virgin coconut oil has promising anti-inflammatory property for minimum period and repeated application is necessary to enhance the effect. Also, Emu oil has high efficacy at 6 hours and elicit potent topical anti-inflammatory property when compare to Virgin coconut oil which is moderately effective at 6 hours. Nonetheless, a profound study is necessary to find out the endogenous corticosteroid response on induced inflammation and isolation, structural elucidation of active moiety to corroborate the exact mechanism of action.

ACKNOWLEDGEMENT: Authors are thankful to Dean, College of Veterinary and Animal Sciences, Thrissur, Kerala for having provided necessary facilities to carry out the research work.

REFERENCES:

1. Hanada T and Yoshimura A: Regulation of cytokine signaling and inflammation.          Cytokine and Growth Factor Rev  2002;13(4–5): 413–421
2. Kirchrath MJ and Schror K: Cyclooxygenase-2 inhibition and side-effects of non-  steroidal anti-inflammatory drugs in the gastrointestinal tract. Curr Med Chem 2000;7(11):1121-1129
3.  Suleyman H, Demircan B and Karagoz Y: Anti-inflammatory and side effects    of cyclooxygenase inhibitors. Pharmacol Rep 2007; 59:247-258.
4. Whitehouse MW, Turner AG, Davis CK and Roberts MS: Emu oil(s): a source of non-toxic transdermal anti-inflammatory agents in aboriginal medicine.  Inflammopharmacol 1998; 6: 1–8.
5.  Beckerbauer LM, Cooper RT, Ahn DU, Sell JL, Parrish FC and Beitz DC:  Influence of two dietary fats on the composition of emu oil and meat. Poultry Sci 2001;80: 187–194.
6. Snowden JM and Whitehouse MW: Anti-inflammatory activity of emu oils in rat.  Inflammopharmacol 1997; 5:  127-132.
7. Yoganathan S, Nicolosi R, Wilson T, Handelman G, Scollin P, Tao R, Binfordd P and Orthoefere F: Antagonism of croton oil inflammation by topical emu oil in CD- 1 mice. Lipids 2003; 38: 603–607.
8.  Lopez A, Sims DE,  Ablett RF, Skinner RE, Leger LW, Lariviere CM,  Burnes LAM and Zawadzka GG: Effect of emu oil on auricular inflammation induced with croton oil in mice.  Am J Vet Res 1991; 60: 1558–1561.
9. Politis, MJ and Dmytrowich A: Promotion of Second Intention Wound Healing by Emu Oil Lotion: Comparative Results with Furasin, Polysporin, and Cortisone.  Plast Reconstr Surg. 1998; 102 (7): 2404-2407.
10.   Zemstov A, Gaddis M, and Montalvo LVM: Moisturizing and cosmetic properties of emu oil: A pilot double blind study.  Aust J Dermatol 1996; 37: 159– 162.
11.  Lindsay RJ, Geier MS, Yazbeck R, Butler RN and Howarth GS: Orally administered emu oil decreases acute inflammation and alters selected small intestinal parameters in a rat model of mucositis.  Br J Nutr 2010; 104: 513–519.
12. Nevin KG and Rajamohan T: Wet and dry extraction of coconut oil: impact on   lipid metabolic and antioxidant status in cholesterol coadministered rats.  Can J               Physiol Pharmacol. 2009; 87(8): 610-616.
13. Srivastava P and Durgaprasad S: Burn wound healing property of Cocos nucifera: An appraisal.  Indian J Pharmacol 2008; 40: 144-146.
14. Intahphuak S, Khonsung P and Panthong A: Anti-inflammatory, analgesic,   and antipyretic activities of virgin coconut oil. Pharmaceutical Bio 2010; 48: 151-157.
15. Vogel HG, Wolfganga HV, Bernward AS, Jurgen S, Gunter M and Wolfganga FV: Drug discovery and evaluation pharmacological assays. Springer, Berlin, Germany,       Edition 3,2002: 725-771.
16. Coruzzi G, Pozzoli C, Adami M,  Grandi D, Guido N, Smits R, Esch DI and Leurs R:  Strain-dependent effects of the histamine H4 receptor antagonist        JNJ7777120  in a murine model of acute skin inflammation. Experimental Dermatology 2012; 21: 32-37.
17. Zhou FH, Foster BK, Sander G and Xian CJ: Expression of proinflammatory cytokines and growth factors at the injured growth plate cartilage in young rats. Bone 2004; 35: 1307–1315.
18. Nadhanan RR, Abimosleh SM, Su YW, Scherer MA, Howarth GS and   Xian CJ: Dietary emu oil supplementation suppresses 5-fluorouracil chemotherapy-induced inflammation, osteoclast formation, and bone loss. Am J Physiol Endocr Metab 2012; 302: 1440- 1449.
    

    ---

How to cite this article:

Logeswari P, Kumar VD, Usha PTA and Kumar P: In vivo anti-inflammatory effect of emu oil (Dromais novaehollandiae) and virgin coconut oil (Cocos nucifera) on phorbol ester induced acute inflammatory model. Int J Pharm Sci Res 2014; 5(3): 896-99.doi: 10.13040/IJPSR.0975-8232.5(3).896-99

All © 2013 are reserved by International Journal of Pharmaceutical Sciences and Research. This Journal licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License.