Cognitive Enhancing Effect of Murraya koengii Leaves by Radial Arm Maze Test in Rats

 

Kadiri Sunil Kumar*, S. Mounica, Rajesh Behera, K. Vijay Kumar, Y. Shanmukh Sai, R. Suthakaran

Vijaya College of Pharmacy, Munaganoor – 501511, Hyderabad, Telangana, India

*Corresponding Author E-mail: sunil.cology@gmail.com

 

ABSTRACT:

The study was designed to assess the cognitive enhancing potential of Murraya koengii leaves by using radial arm maze test in albino rats. The radial arm maze test consists of a total 7 days session. During first 6 days training session (habituation phase) rats were placed individually in the central hub and were allowed to choose the arm containing the food freely for a period of 5 minutes. The time taken by each rat to find the food was considered to assess RAM performance. Latency to find the food was noted as an index of memory. The same procedure was repeated with test and standard treated rats after one hour of administration of murraya koengii chloroform extract 100 mg/kg and Piracetam 200 mg/kg. After 6 days training session, 7th day session includes induction of short term memory loss in control rats by administration of propofol 0.5 ml/200g (amnesia inducer)  followed by latency to find the food is noted for a period of 5 minutes for each rat. Repeat the same propofol treatment on 7th day for test and standard treated rats after 1 hr of administration of extract (Murraya koenigii 100mg/kg) and piracetam (200mg/kg) and the latency to find was recorded for 5 minutes.  It was found that the latency to find food on 7th day of radial arm maze test in control rats after administration of propofol is 2.59 seconds, whereas latency to find food with test extract treated rats on 7th day was found to be 2.20 seconds. This suggests that decrease in the latency for food (sec) with Murraya koengii leaves extract has significantly increased the memorizing ability of rats to find food. Hence it indicates that Murraya koenigii leaves at a dose of 100mg/kg produces significant cognitive enhancing potential in albino rats. However the memory enhancing potential of Murraya koenigii leaves was found to be lesser that standard drug piracetam (latency for food 2.05 seconds) in Radial arm maze test.

 

KEYWORDS: Cognitive enhancing, Murraya koenigii, latency for food, radial arm maze test.

 

 


INTRODUCTION:

Cognition is the process in which information is encoded, stored and retrieved1. Cognition is the mental ability of developing knowledge and understanding through thought, experience and the senses 2. Poor memory, slow recall and lower retention are common problems in today’s world3. Memory declines mostly under stress and fatigue4. Several memory enhancing drugs are available in the modern medicine but with potentially toxic adverse effects5.

The Murraya koenigii plant is widely used as herb, spice, condiments and also used to treat various types of ailments in Indian traditional system. World’s about 80% population relies upon herbal products, because they have been considered as safe, effective and economical. The various parts of this plant are widely used by different tribal communities. The leaves of plant are use as tonic, stomachic, carminative, internally in dysentery, vomiting6,7. Used as antihelminthic, analgesic, cures piles, allays heat of the body, thirst, inflammation and itching 8, 9. The present study was designed to investigate the memory enhancing activity of Murraya koenigii leaves by using radial arm maze test in rats.

 

MATERIALS AND METHODS:

For the present study, the leaves of Murraya koenigii were collected from the surrounding gardens of the Vanasthalipuram, Hayathnagar (Mandal), Ranga Reddy dist, Telangana, India. After the fresh leaves were authenticated by Dr. N. Sivaraj, Senior Scientist (Eco Botany), National Bureau of Plant Genetic Resources, Rajendranagar, Hyderabad, leaf specimens have been deposited at the museum of the college. Fresh mature leaves were shade dried at room temperature, coarse powdered with electric grinder and further extracted with chloroform by maceration for a period of 5 days. Thereafter, the extract was concentrated by evaporation. The percentage yield of the leaf extract was found to be 12.7%.  The extract was stored in airtight container in refrigerator below 10ºc.  Appropriate concentration of stock solution of extract were prepared using span 80 and used for the following studies.

 

Preliminary phytochemical screening:10

Preliminary phytochemical tests were performed for the chloroform extract of Murraya koenigii leaves to detect the presence of phytochemicals by following the standard methods described in the Practical Pharmacognosy of Kokate and Khandelwal. The results have been tabulated in table I.

 

Experimental animals:

Albino rats (180-225g) were used in the present research. They were procured from Sainath Agencies, Musheerabad (282/99/CPCSEA). After randomization into various groups and before initiation of experiment, the rats were acclimatized for a period of 10 days. Animals were housed in polypropylene cages and maintained under standard environmental conditions such as temperature (26 ± 2ºc), relative humidity (45-55%) and 12hr dark/light cycle. The animals were fed with rodent pellet diet (Golden Mohur Lipton India Ltd.) and water ad libitum. The study protocol was approved from the institutional animal ethics committee (IAEC) before commencement of experiment IAEC (1292/ac/09/CPCSEA).

Determination of acute toxicity (OECD guidelines 423):11

The Murraya koenigii chloroform extract was studied for acute toxicity study at a dose of 100 mg/kg, 200mg/kg, 400 mg/kg, 800 mg/kg and 2000 mg/kg p.o in albino rats (6 rats in each group). All the rats were observed for toxicity signs for 24hrs followed by 14 days. On 8th day and14th day body weight of rats were recorded. The rats were found toxic at the dose of 2000mg/kg i.e., rats exhibited signs of convulsions, drowsiness and ataxia. Hence 1000 mg/kg was selected as safe dose and 1/10 of 1000mg/kg i.e., 100mg/kg of Murraya koenigii leaves was selected for the studies.

 

Effect of Chloroform extract of Murraya koenigii leaves on memory by radial arm maze test in rats:

The experiment was performed on albino rats (150-250gms) of either sex procured from Sainath Agencies, Musheerabad. The animals were housed in colony cages at an ambient temperature of 26±20C and relative humidity (45-55%) with a 12h/12h light dark cycle and access to food and water ad libitum. Food was restricted during experiments. Stock solutions of piracetam (200mg/kg) was prepared in 2% acacia suspension and Chloroform extract of Murraya koenigii leaves (100mg/kg) was prepared in SPAN 80.  Divide the animals in to three groups control(C), test (T), and standard(S) each consisting of three rats. All the animals were subjected to radial arm maze test apparatus assessment of nootropic levels. The radial arm maze apparatus is composed of 8 arms (33cm L, 13cm W) with each arm number, octagonal central chamber (Diameter 37cm) and there are hidden platforms in each arm at a distance of 30cm from central hub where the food is placed in  2 fixed arms (1 and5). The rats were trained for RAM performance by conducting daily training trial for 6 days12. The radial arm maze test includes 6 days training session (habituation phase) in which rats were placed individually in the central hub and were allowed to choose the arm freely to get the food within 5 minutes. The time taken by each rat to find the food was considered to assess RAM performance. Latency to find the food was noted in the habituation phase. Repeat the same procedure with test and standard treated rats after one hour of administration of murraya koengii chloroform extract 100 mg/kg and Piracetam 200 mg/kg. After 6 days training session, 7th day session includes administration of propofol 0.5 ml/ 200 gm which induces short term amnesia to control rats and latency to find the food is noted for a period of 5 minutes for each rat13. Repeat the same propofol treatment for test and standard treated rats after 1 hr of administration of extract (Murraya koenigii 100mg/kg) and piracetam (200mg/kg) and the latency to find the food is noted for a period of 5 minutes.

 

 

 

 

 

Figure I: Rats Performing Radial Arm Maze Test to Find Food

 

STATISTICAL ANALYSIS:

The values are represented as mean ± S.E.M and statistical significance between treated and control groups was analyzed using One way ANOVA, followed by Dunnett’s test where P<0.001, P<0.01 and P<0.05 was considered statistically significant.

 

RESULTS AND DISCUSSION:

Preliminary Phytochemical screening of chloroform extract of Murraya koenigii 100mg/kg leaves reveals the presence of flavonoids, triterpenoids, saponins, traces of tannins, carbohydrates, steroids, glycosides, alkaloids, proteins, resins, phytosterols, gum and mucilages. It is evident from the literature that alkaloids have potent memory enhancing ability14. Murraya koenigii leaves contains significant amounts of  carbazole alkaloids such as koenimbine, o-methyl murrayamine, o-methyl mahanine, isomahanine, bismahanine and bispyrayafoline.15 In radial arm maze nootropic assessment of Murraya koenigii leaf extract, it was found that the latency for food shown by control rats is 2.59 seconds. Whereas the latency to find food by test treated rats was found to be 2.20 seconds. This indicates that the test treated rats have a quick tendency (in seconds) to find food when compared to control rats. This clearly shows that the Murraya koenigii leaves treated rats exhibits memory enhancing potential. However Piracetam 200 mg/kg treated rats has shown latency for food of 2.05 seconds which is much lesser than the test extract treated rats. This indicates that standard piracetam is a potent nootropic drug than the test extract.

 

Table I: Preliminary Phytochemical screening of chloroform extract of Murraya koenigii leaves.

Phytoconstituents

Chloroform leaf extract

Carbohydrates

Steroids

Glycosides

Flavonoids

Alkaloids

Tannins

proteins

Triterpeniods

Resins

Phytosterols

Gum and mucilages

   +

   -

  +

  +

  +

  -

  +

  +

  +

  +

  - 

-   Absent; +   Present

 


 

 

 

 

Table II:  Latency to find food (seconds) by rats during 6 days training period (habituation phase)

GROUP

DAY 1

DAY 2

DAY 3

DAY 4

DAY 5

DAY 6

Control

3.48

2.86

2.63

2.41

2.20

1.84

Test

3.46

2.62

2.26*

2.36

1.97

1.55*

standard

3.32

2.46

2.09**

2.17*

1.46**

0.90**

                      P<0.001***, P<0.01** and P<0.05* was considered statistically significant.


 

 

 

 

Table III: Latency to find food (seconds) by rats on 7th day after induction of Propofol.

GROUP

Latency for food (seconds) on 7th day

Control

2.59

Test extract 100 mg/kg

2.20*

 Standard Piracetam     200 mg/kg

2.05**

 

                 P<0.001***, P<0.01** and P<0.05* was considered statistically significant

 

 


 

 

Figure II: latency for food (sec) on 7th day with control, test and standard treated rats after propofol treatment.

 

 

CONCLUSION:

The administration of anesthetic agent propofol produces transient memory deficit. The data obtained from the present nootropic study by using radial arm maze model in rats indicates that the latency for food with Murraya koenigii leaves at 100 mg/kg was lesser than the latency for food with control rats.  As the latency for food determines how quickly the rats recognizes the arm containing the food and enters in that particular arm. Hence decrease in latency for food with test extract indicates that that test treated rats retains the ability to memorize the arms containing the food.  This suggests the nootropic potential of Murraya koenigii leaves. It may be hypothesized that Murraya koenigii leaves may have a neuroprotective mechanism on brain cholinergic neurons.  However further phytochemical isolation of alkaloids has to be done to know the exact alkaloids involved in cognition enhancing activity of Murraya koenigii leaves with their precise mechanism of action.

 

ACKNOWLEDGEMENTS:

The authors are grateful to the management of Vijaya College of Pharmacy, Hyderabad for providing the facilities for our Research.

 

REFERENCES

1.     Baddeley, A. D. (1966). The influence of acoustic and semantic similarity on long-term memory for word sequences. Quart. J. Exp. Psychol 18 (4): 302–9.

2.     Parke, R. D., and Gauvain, M. (2009). Child psychology: A contemporary viewpoint (7th Ed.). Boston, MA: McGraw-Hill.

3.     Atkins CM, Selcher JC, Petraitis JJ, Trzaskos JM, Sweatt JD 1998. The MAPK cascade is required for mammalian associative learning. Nat Neurosci 1:602–609.

4.     Petkov VD, Y on kov D, Mosharoff A, Kambourova T, Petkov VV, and Todorov I. 1986. Effects of alcohol aqueous extract  from Rhodiola rosea L. roots on learning and memory. Acta Physiol Pharmacol Bulg 12(1):3–16.

5.     Joint Formulary Committee (2004). British National Formulary (47th ed.). London: BMA and the Royal Pharmaceutical Society of Great Britain. ISBN 085369-584-9.

6.     Gupta S, George M, Singhal M, Sharma GN, Garg V. Leaves extract of Murraya koenigii Linn. for anti-inflammatory and analgesic activity in animal models. Journal of Advanced Pharmaceutical Technology and Research 2010; 1:68-77.

7.     Khan BA, Abraham A, Leelamma S. Hypoglycemic action of Murraya koenigii curry leaf and Brassica juncea mustard: mechanism of action. Indian Journal of Biochemistry and Biophysics 1995; 32:106-108.

8.     Goswami RB, Prabhat Khare, Sukhwant Singh, Neelima Goswami, Princey Thomas, Uma Devi P et al. Studies on antigenotoxic of Murraya koenigii. International Journal of Pharma Recent Research 2010; 2 2 :65- 68.

9.     Khuntia TK, Panda DS. Evaluation of antibacterial, antifungal and anthelmintic activity of Murraya koenigii Spreng. An International Journal of Pharmaceutical Sciences 2011; 2 2 :105-110.

10.   Kokate CK. Practical Pharmacognosy, 4th edition. Vallabh Prakashan Publications, New Delhi; 1994. 

11.   Roll R., Höfer-Bosse Th.  and Kayser D. (1986). New Perspectives in Acute Toxicity Testing of Chemicals. Toxicol. Lett., Suppl. 31, 86.

12.   G.L. Gupta and S.S. Nigam. Chemical examination of the leaves of Murraya koenigii. Planta Med. 19: 83 (1970)

13.   R.L. Khosa, S.P. Sen and S.N. Dixit. Studies on Murraya paniculata. Indian J. Pharm. 32: 65 (1970).

14.   Pramodine D. Kulkarni, Mahesh M. Ghaisas, Niranjan M. Chivate, Poournima. S. Sankpal. Memory enhancing activity of Cissampelos pariera. International Journal of Pharmacy and Pharmaceutical Sciences 2011.3(2), 206-211.

15.   Vandana Jain, Munira Momin, Kirti laddha. Murraya Koenigii : An updated review. International Journal of Ayurvedic and Herbal Medicine. 2012.2 (4), 607-627.

 

 

 

 

Received on 02.05.2016       Modified on 19.05.2016

Accepted on 11.06.2016      ©A&V Publications All right reserved

Research J. Pharmacology & Pharmacodynamics.2016; 8(2): 71-74

DOI: 10.5958/2321-5836.2016.00013.6: