The fresh leaves of Loniceraligustrina were collected and authenticated by botanist Dr. Madhavachetty, Assistant professor,Department of Botany in S.V. University, Tirupathi, and voucher specimen number (1189).

Fresh leaves of Loniceraligustrina were cleaned and dried under shade in clean dust free environment, grinded and stored in an air-tight container. The total 50 g of coarse powder of LL was used & extracted with 250ml of 90% ethanol in a soxhlet apparatus[7] at 60- 75°C for 48 hrs. The extract was concentrated by evaporation. The yield was about 5.0% and stored at 4˚C for future use. The above extracted yield was taken to perform Phytochemical screening & the results are tabulated below in Table 2.

Table1: Preliminary-phytochemical screening

Table2:Preliminary-Phytochemical screening[8-10].

The solid EELL was dissolved by using 1 % v/v Tween-80 as a vehicle for i.padministration [2-4].

Swiss albino mice (15-20)g, was maintained under the standard conditions of (27 + 20C; relative humidity 60 + 5%, light dark cycle for 12 hrs) and standard pellet diet feed and water adlibidum were used for present study[5] . All the experimental protocols were duly approved by Institutional Animal Ethics Committee (Reg. No: 1477/Re/s/11/CPCSEA), Dhanvanthri College of Pharmaceutical sciences, Mahaboobnagar, Telangana, India.

Swiss albino mices are taken under the study of limit test. Three animals were kept for overnight fasting prior to drug administration and treated with 2000mg/kg as a single dose of Loniceraligustrina leaf extract. Via i.p. route as per the limit test of OECD guide lines 423 and the food was with held for further 3-4 hours and the mices were individually observed individually atleast once prior to 30minutes after dosing & periodically during the first 24 hours (with special attention given during the first 4 hours), and daily thereafter, for the total of 14days, for neurological, behavioral, and autonomic profiles (skin and fur, eye and mucous membranes, and also, circulatory, respiratory autonomic and central nervous systems, and somatomotor activity and behavioral pattern.

Attention should be directed to observe the salivation, convulsions, tremors, lethargy, sleep,coma& diarrhea) and for any lethality, morbidity and mortality. As per animal body weight required amount of standard drug (25 mg/kg) of Diclofenac sodium taken and dissolved in distilled water Loniceraligustrina leaf extract was used for trituration with 1ml of Tween 80 and 9ml of water added to form a suspension and dose was calculated as per animal body weight.

Swiss albino mice weighing around 150-200g. were divided in five groups, each group consisting of about six animals is treated as described below in table 3

Table 3:

This test was performed by using a tail immersion method, where swiss albino mice of either sex were divided in to four groups, each containing six animals. The basal reaction timeof all animals were recorded. A water bath was maintained at 510C (+ 0.5) using thermostatic setting provided which is provided with an instrument.

A thick paper rolled in to make a cone with a hole in the bottom & each animal were taken in cone and allowed to extend tail out through bottom hole. 2cm length of a tail was marked using marker pen and was dipped to note the response. Tail flick out by the mice of the hot water and body jerks were taken as response by using stop watch starting from the time of dipping to the time of flick, responses were recorded. A cut off period of 60sec was observed to avoid damage to tail.

The latency period was recorded at 30 min interval up to 90 min for control, standard and test drug administration.

The tail flick test was used to calculate an analgesic activity by following the method defined by D’amour and Smith 1941 [6], with minor alterations in a given procedure. This tail flick method was utilized to study analgesic activity in mice.

A radiant heat automatic tail flick analgesiometer was applied to measure reaction latencies. Basal reaction time was recorded for the radiant heat by observing the tip (last 1-2 cm) of the tail on radiant heat source. The tail removal from the radiant warmth was taken as end point. To avoid injury of tail by radiant heat ,cut off time of 15 seconds was used. Miceswere divided into five groups, each contains six mice. Treated with standard (25 mg/kg), control, and extract (150, and 300 mg/kg)was followed.Tail-flick response with the latent period was determined at 30min, 45min, 60min, 75min, and 90 minutes after drugs administration.

Table 2:Data of EELL (Ethanolic extract of LoniceraLigustrina) by Tail Immersion Method

Table 3: Mean data of Analgesic activity of EELL by Tail immersion method on Swiss Albino Mice.

Values are expressed as mean + SEM (n=6); Statistical Analysis of data was carried out by Student t-test comparision test, *p< 0.05, **p <0.01, ***p < 0.001 compared to distilled water treated group; EELL=Ethanolic Extract of Loni

ceraligustrina.

Fig 1: Graphical representation by Tail immersion method using Swiss albino mice.

Table 4:Tail Flick Method

Table 5 : Data of Analgesic Activity of EELL by Tail Flick Method on Swiss Albino Mice

Values are expressed as mean + SEM (n=6); Statistical Analysis of data

was carried out by Student t-test comparision test, *p<0.05, **p<0.01, ***p<0.001 compared to distilled water treated group; EELL=Ethanolic Extract of Lonicera ligustrina

Fig 3: Graphical representation by Tail flick method using Swiss albino mice.