EFFECT OF VITAMIN E ON ATORVASTATIN …ijtpls.com/.../2015/12/IJTPLS-2015-Vol-15-PRASANNA-KUMAR-7-600-612.pdfPrasanna Kumar B et al. Int J Trends in Pharm & Life Sci. 2015: 1(5);600-612

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Prasanna Kumar B et al. Int J Trends in Pharm & Life Sci. 2015: 1(5);600-612 . 600

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International Journal of Trends in Pharmacy and Life Sciences Vol. 1, Issue: 5, 2015: 600-612

EFFECT OF VITAMIN E ON ATORVASTATIN INDUCED SELECTED

BEHAVIORAL AND BIOCHEMICAL ALTERATIONS IN NORMAL RATS B. Prasanna Kumar, K. Sravanth & D. Rama brahma Reddy

Nalanda institute of Pharmaceutical Sciences, Siddharth Nagar, Kantepudi(Village),

Sattenapalli(Mandal), Guntur(Dist)-522438 E.Mail: [email protected]

ABSTRACT

Atorvastatin is synthetic compound containing a heptanoic acid side chain that forms a structural

analog of the HMG-CoA intermediate. Atorvastatin, like all statins, is a selective, competitive inhibitor of

HMG-CoA reductase. The aim of the Study is to find out the Possible Role of Vitamin E on Atorvastatin

Induced Behavioural and Selected biochemical alterations in Normal Rats. The results of the present study

showed that, chronic treatment with Atorvastatin produce Neuro pharmacological and Biochemical

alterations such as decreased body weight, exploratory behaviour, loco motor activity, motor co –

ordination, muscle grip strength and depression . Decrease in levels of antioxidant enzymes. Vitamin E

Supplementation decreased Atorvastatin induced Neuro pharmacological and severe side effects the

increasing anti-oxidant enzymes SOD, CAT, gultathion, Dose dependently.

*Corresponding Author:

Mr.B. Prasanna Kumar,

Nalanda institute of Pharmaceutical Sciences, Siddharth Nagar,

Kantepudi(Village),

Sattenapalli(Mandal), Guntur(Dist)-522438 E.Mail: [email protected]

INTRODUCTION

Over the past decade, there has been substantial interest in oxidative stress and its potential role in

epilepsy, development of diabetic complications, atherosclerosis and associated cardiovascular disease.

Consequences of oxidative stress are damage to DNA, lipids, proteins, resulting in the disruption of cellular

homeostasis. 3-hydroxy-3-methyl glutaryl co-enzyme A (HMG – CoA reductase) inhibitors are the most

widely prescribed drugs in the world and the United States and contributed substantially to prescription drug

cost increases in 2001. [1].Atorvastatin in most widely prescribed statin drug. It is drug of choice for

treatment of elevated cholesterol and congestive heart diseases and obesity .Cholesterol is a vital substance

of the body, at the lower levels of cholesterol mortality is increased due to non-cardiovascular deaths, like

cancer, accidents and depression [2].

Atorvastatin is synthetic compound containing a heptanoic acid side chain that forms a structural

analog of the HMG-CoA intermediate. Atorvastatin, like all statins, is a selective, competitive inhibitor of

HMG-CoA reductase.

Vitamin E is the principle membrane-associated antioxidant molecule in mammals. It plays a major

role in preventing oxidative damage to membrane lipids by scavenging free radicals. [3,4].

Received: 22/10/2015

Revised: 20/11/2015

Accepted: 28/11/2015



The aim of the Study is to find out the Possible Role of Vitamin E on Atorvastatin Induced Behavioural

and Selected biochemical alterations in Normal Rats.

MATERIALS AND METHODS

Experimental Animals

Pathogen free adult male albino rats weighing 150-200 g were used. Male rats were chosen to avoid

fluctuations due to oestrous cycle (Sugioka et al., 1987). The rats were housed in polypropylene cages (Five

per cage) under standard laboratory conditions with 12 hours light / dark cycle. The rats were fed with

standard laboratory chow (NATIONAL INSTITUTE OF NUTRITION) and Water ad libitum.

Study Protocol

Grouping of Animals

The animals were divided in to six groups of 5 each

1. Group-I : Normal control

2. Group-II : Vitamine E group

3. Group-III : Atorvastatin group (10 mg/kg)

4. Group-IV : Atorvastatin + Low dose of vitamin E 50 mg/kg

5. Group-V : Atorvastatin + Middle dose of vitamin E 100 mg/kg

6. Group-VI : Atorvastatin + highest dose of vitamin E 200 mg/kg

Drugs and treatment schedule

Atorvastatin at a dose of 10 mg/kg diluted with 0.2% CMC was administered by oral gauge daily

between 8.00 to 9.00 am for 4-6 weeks [5, 6, 7,8]

Vitamin E, a lipid soluble antioxidant, at doses of 50, 100, 200 mg/kg was suspended in 1% of tween

80 and was administered orally once (one hour before Atorvastatin) treatment Control group

received 1% solution of tween 80 orally daily for 4-6 weeks.

Behavioural Parameters: On every 15 day, 24 hours after drug treatment all behavioural parameters were

studied. All animals were handled for at least for 7 days before testing behavioural parameters.

Motor Co-ordination test: Motor co-ordination test was conducted in groups of rats using a Rota rod

Apparatus (Inco Ambala). The animals were placed on the Moing rod prior to the treatment and the rats that

stayed on the rod without falling for 120 sec chosen for the study [9]

Exploratory Behaviour test: This test was done using Hole Board. The hole Board consisted of a 0.5 m3

wooden board with 16 holes (3 cm in diameter). Each rat was placed singly on the board for a period of 6

minutes. The number of exploratory movements performed within the last 4 minutes of this period was

noted for each animal [10]

Locomotor Activity: The locomotor activity can be easily measured using an Actophotometer which

operates on photo electric cells which are connected in circuit with a counter. When the beam of light



falling on photo cell is cut off by the animal, a count is recorded. An Actophotometer could have either

circular (or) square area in which the animal moves. Both rats and mice may be used for testing in this

equipment.

Body Weight Analysis: On the first day of the experiment each animal weight noted and expressed in

grams. Weight analysis is done for 15 days up to 45 days.

Biochemical Analyses: Blood samples were collected at the end of the experiment from retro-orbital plexus

under light ether anesthesia. Blood samples were kept for 30-60 minutes in dry test tubes without any

disturbance, citrated blood is used for obtaining plasma. Then centrifuged for 10 minutes at 3000 rpm to

separate the serum and plasma.

Blood was analyzed for aspartate transminase (AST) [11].

On 45th

day blood samples were collected from retro-orbital plexus and estimated for

superoxide dismutase, catalase [12] glutathione [13]

Statistical Analysis

All the values are expressed as mean ± SD. The statistical analysis was carried out using Two way

analysis of variance (ANOVA) followed by Tukey’s mutiple comparison test. In all tests, the criterion for

statistical significance was p< 0.05, p<0.01, p<0.001 were considered as significant.

RESULTS AND DISCUSSION

Body Weight: -Chronic Atorvastatin 10mg/kg treatment significantly decreased (p<0.01) the body weight

in all treatment groups except vitamin E treated group on 15th

day when compared to the control group (fig.

1). Vitamin E 50 mg/kg treatment did not increase the Atorvastatin decreased body weight when compared

to Atorvastatin treatment group. Vitamin E 100 and 200mg/kg treatment significantly increase (P<0.05 and

P.01) the Atorvastatin decreased body weight on 30th

day when compared to the Atorvastatin treatment

group (fig . 1). And maintained the body weight when compared to the control group (fig 1 ).

Table 1: Effect of chronic treatment of Atorvastatin, vitamin E and Atorvastatin + Vitamin E on body

weight

Group

No.

Treatment mg/kg Body weight (days)

0 15 30 45

1. Control 176 ± 25.61 182.4±26.34 188.3 ± 28.310 194 ± 29.81

2. Atorvastatin (10 mg/kg) 180 ± 31.62 176.2 ± 34.89 164.4 ± 31.958 152.2 ± 31.26 b

3. Vitamin E (100 mg/kg) 172.8 ±

32.05

178.6 ± 30.163 182.8± 29.819 189.8 ± 30.27 1a

4. Atorvastatin (10 mg/kg) +

vitamin E

(50 mg/kg)

182.6 ±

27.83

175.8 ± 26.3 186 ± 26.623 1a

196.4 ± 27.53 1a


vitamin E

(100 mg/kg)

175.8 ± 26.3 197.2 ± 23.511a, a

199.4 ± 23.44 1b

205 ± 23.17 a 1b


vitamin E

(200 mg/kg)

171 ± 24.

597

176.4 ± 22.963 182.4 ± 23.201 187.8 ± 23.541 1a



Fig.1: Effect of chronic treatment of Atorvastatin, vitamin E and Atorvastatin + Vitamin E on body weight

Note: Values expressed as mean ± SD (N=5)

a, p<0.05; b p<0.01; c p<0.001 vs control group

1a, p<0.05; 1b p<0.01; 1c p<0.001 vs atorvastatin group

2a, p<0.05; 2b p<0.01; 2c p<0.001 vs vitamin E group

3a, p<0.05; 3b p<0.01; 3c p<0.001 vs ator + vitamin E (50 mg/kg)



(2 WAY ANOVA) followed by TUKEY-KRAMER multiple comparison test

Hole board test: Chronic Atorvastatin at a dose of 10mg/kg significantly decreased (P<0.001) the number

of exploratory movements in all treatment groups except Vitamin E on 30th

day onwards when compared to

the control group (fig-2).

Vitamin E 50mg/kg treatment did not increase the Atorvastatin decreased exploratory movements when

compared to Atorvastatin treatment group. Whereas Vitamin E 100 and 200-mg/kg treatment significantly

increased (P<0.01) and (P<0.001) Atorvastatin decreased exploratory movements on 40th

day and 30th

day

respectively when compared to the Atorvastatin treatment group (fig-2).



Table 2: Effect of chronic treatment of Atorvastatin, vitamin E and Atorvastatin + Vitamin E on Hole

Board

Group

No.

Treatment mg/kg Hole Board

0 15 30 45

1. Control 16.4 ± 6.542 17.2 ± 3.136 18.6 ± 2.178 20.06 ± 3.876

2. Atorvastatin (10 mg/kg) 16.4 ± 6.542 8.6 ± 2.408 6.8 ± 2.168 5.2 ± 1.924

3. Vitamin E (100 mg/kg) 20.8 ± 4.817 14.6 ± 2.408 15.8 ± 2.28 1b

18.2 ± 2.387 1c


vitamin E

(50 mg/kg)

17.8 ± 3.271 15.4 ± 2.881 11.6 ± 5.55 1a

12.8 ± 3.421 1b


vitamin E

(100 mg/kg)

22.2 ± 4.919 13.4 ± 3.578 15.6 ± 3.209 1b

20.6 ± 3.975 1c


vitamin E

(200 mg/kg)

24 ± 3.536 14.2 ± 2.864 17.6 ± 4.037 1c

21 ± 3.536 1c,3a

Fig.2: Effect of chronic treatment of Atorvastatin, vitamin E and Atorvastatin + Vitamin E on Hole Board











Loco motor activity:

Chronic Atorvastatin 10 mg/kg treatment significantly decreased (P<0.001). The Locomotion in all

treatment groups Except Vitamin E treated group on 30th day when compared to the control group (fig-3).

Vitamin E 50mg/kg treatment did not increase the Atorvastatin decreased Locomotion when compared to

the Atorvastatin treatment group. Vitamin E 100 and 200mg/kg treatment , significantly increased

(P<0.01) the Atorvastatin decreased Locomotion on 45th

day and 30th

day of treatment respectively when

compared to the Atorvastatin treatment group (fig -3).

Table 3: Effect of chronic treatment of Atorvastatin, vitamin E and Atorvastatin + Vitamin E on

Spontaneous motor activity

Group

No.

Treatment mg/kg SMA Count

0 15 30 45

1. Control 167.2 ± 23.25 2a

184 ± 40.97 184 ± 36.73 176 ± 29.31

2. Atorvastatin (10 mg/kg) 144.8 ± 28.38 2a

130.1 ± 36.28 a 112 ± 41.34

a 96.2 ± 11.47

c

3. Vitamin E (100 mg/kg) 181.8 ± 29.21 b 192.8 ± 34.65

1b

187.2 ± 24.26 208.2 ± 23.79 1a

4. Atorvastatin (10 mg/kg)

+ vitamin E

(50 mg/kg)

158.4 ± 19.922b

197.8 ± 27.62 b 139.8 ± 48.42

b, 1a, 2a

122.2 ± 34 b,2a


+ vitamin E

(100 mg/kg)

170.2 ± 34.10 2b

172.6 ± 57.47 1a

175.6 ± 58.19 a 215 ± 36.1

3b


+ vitamin E

(200 mg/kg)

172.0 ± 31.23 a 174.0 ± 38.70

b, 1b, 2a

186.8 ± 32.19 a,

2a

187.2 ± 34. a, 4a, 2a



Fig.3: Effect of chronic treatment of Atorvastatin, vitamin E and Atorvastatin + Vitamin E on Spontaneous

motor activity (SMA)









MOTOR CO- ORDINATION TEST:- Chronic Atorvastatin 10mg/kg treatment significantly impaired

(P<0.001) the Rota rod performance from 15th

day of the treatment in all treatment groups except Vitamin

E treated group and from 30th

day of treatment and maintain rota rod performance when compared to the

control group (fig. 4).

Atorvastatin 50mg/kg did not maintain the Atorvastatin decreased Rota rod performance when compared to

Atorvastatin treatment group. Vitamin E 100 and 200mg/kg treatment significantly increased (P<0.01 and P

< 0.001) , maintained the Atorvastatin impaired Rota rod performance from 30th

day when compared to the

Atorvastatin treatment group (fig.4 ).



Table 4: Effect of chronic treatment Atorvastatin, vitamin E and Atorvastatin + vitamin E on Rota

Rod performance

Group

No.

Treatment mg/kg Rota Rod Performance (days)

0 15 30 45

1. Control 120 ±

0

120 ± 0 120 ± 0 120 ± 0

2. Atorvastatin (10 mg/kg) 120 ±

0

111.6 ± 9.343 112 ± 16.793c 2b 3b

90 ± 10.886 c2c

3. Vitamin E (100 mg/kg) 120 ±

0

120 ± 0c 120 ± 0

1b 120 ± 0

4. Atorvastatin (10 mg/kg) + vitamin

E

(50 mg/kg)

120 ±

0

59.8 ± 24.284 70.6 ± 34.480c 96.2 ± 15.023

b 2a

1c


E

(100 mg/kg)

120 ±

0

90 ± 10.886c 112 ± 16.973

a 1c

3b

120 ± 0 1c 3b


E

(200 mg/kg)

120 ±

0

114.4 ±

12.522a

116.2 ± 6.943 1b

120 ± 0 1c

Fig. 4: Effect of chronic treatment ofAtorvastatin, vitamin E and Atorvastatin + vitamin E on Rota Rod

performance











SUPEROXIDE DISMUTASE:

Chronic Atorvastatin 10mg/kg treatment significantly decreased (P<0.001) the Super oxide dismutase levels

in all treatment groups except Vitamin E treatment group when compared to control group (fig - 5).

Vitamin E 50mg/kg treatment did not increased the Atorvastatin decreased Super oxide dismutase levels

when compared to the Atorvastatin treatment group. Where as Vitamin E 100 and 200mg/kg treatment

significantly increased (P<0.01( and (P< 0.01). Super oxide dismutase levels respectively when compared to

the Atorvastatin group. (fig - 5).

Table 5: Effect of Chronic Treatment of Atorvastatin, Vitamin-E and Atorvastatin and Vitamin-E on Super

Oxide Dismutase Levels

S.No Groups SOD (IU/ml)

1 Control 33.43.782 2 2a 1c 3c 4c

2 Vitamin E 100 mg/kg 25.83.114 a1c3b

3 Atorvastatin 10mg/kg 7.82.049 c2c 4b 5c

4 Atorvastatin +50mg/kg vitamin E 54.583

c 2b 5c

5 Atorvastatin +100mgkg vitamin E 19.43.286 c 1b 5a

6 Atorvastatin+200mg/kg vitamin E 27.64.879 1c 3c 4a

Fig 5: Effect of Chronic Treatment of Atorvastatin, Vitamin-E and Atorvastatin and Vitamin-E on Super

Oxide Dismutase Levels

Note: Values expressed as mean SD (N=5)

a, p<0.05; b p<0.01; c p<0.001 vs Control Group

1a, p<0.05; 1b p<0.01; 1c p<0.001 vs Atorvastatin Group



2a, p<0.05; 2b p<0.01; 2c p<0.001 vs Vitamin – E Group

3a, p<0.05; 3b p<0.01; 3c p<0.001 vs Ator + Vit – E (50 mg/kg)




CATALASE:

Chronic Atorvastatin 10mg/kg treatment significantly decreased (P<0.001) the catalase levels in all

treatment groups except Vitamin E treatment group when compared to control group. (fig- 6).Vitamin E

50mg/kg treatment did not increase the Atorvastatin decreased Catalase levels when compared to the

Atorvastatin treatment group. Whereas Vitamin E 100 and 200mg/kg treatment significantly (P<0.001) the

Catalase levels respectively when compared to Atorvastatin group. (fig-6).

Table 6: Effect Chronic treatment of Atorvastatin, Vitamin-E and Atorvastatin + Vitamin -E on Catalase

Levels

S.No Groups Catalase(µmoles/mg/min)

1 Control 5.944 ± 0.4451 3c 4b 5a

2 Vitamin E 100mg/kg 4.38 ± 0.7338 b 1c 3c

3 Atorvastatin 10mg/kg 1.2244 ± 0.3558 c 2c 4c 5c

4 Atorvastatin+50mg/kg vitamin E 2.145 ±0.6971 c 2c 4b 5c

5 Atorvastatin +100mg/kg vitamin E 3.492 ± 0.4798 c 1c 3b 5b

6 Atorvastatin +200mg/kg vitamin E 4.876 ± 0.4451 a 1c 3c 4b

Fig 5: Effect of Chronic Treatment of Atorvastatin, Vitamin-E and Atorvastatin and Vitamin-E on Catalase

Levels











GLUTATHIONE:

Atorvastatin 10mg/kg treatment significantly decreased (P<0.001) the Glutathione level in all treatment

groups except Vitamin E treatment group. When compared to control group. (fig-7 ).

Vitamin E 50mg/kg treatment did not increase the Atorvastatin decreased Glutathione levels when compared

to Atorvastatin treatment group. Whereas Vitamin E 100mg/kg and 200mg/kg treatment significantly

increased (P<0.01) and (P<0.001) the Glutathione levels respectively when compared to the Atorvastatin

treatment group (fig-7 ).

Table 7: Effect Chronic treatment of Atorvastatin, Vitamin-E and Atorvastatin + Vitamin -E on Glutathione

Levels

S.No Groups Glutathione (mg/dl)

1 Control 199 ± 57.922 1c 3c 4a

2 Vitamin E 100 mg/kg 229.8 ± 58.611 1c 3c 4b

3 Atorvastatin 10mg/kg 11.6 ± 1.700 c 2c 4a 5c

4 Atorvastatin+50mg/kg vitamin E 16.38 ± 2.574 c 2c 4a 5c

5 Atorvastatin +100mg /kg vitamin E 105.4 ± 23.458 a 1a 2a 3a

6 Atorvastatin+200mg/kg vitamin E 228 ± 55.082 1c 2c 3c 4b 5b

CONCLUSION

The results of the present study showed that, chronic treatment with Atorvastatin produce

Neuropharmacological and Biochemical alterations such as decreased body weight, exploratory behavior,

loco motor activity, motor co –ordination, muscle grip strength and depression . Decrease in levels of

antioxidant enzymes. This may support the proposed contribution of oxidative injury in exacerbating

Neuropharmacological, Biochemical alterations induced by atorvastatin. The results of the present study

indicate thatAtorvastatin prescription requires monitoring or cholesterol levels, as it may lead to

Neuropharmacological alterations. Vitamin E Supplementation decreased Atorvastatin induced

Neuropharmacological and severe side effects the increasing anti-oxidant enzymes SOD, CAT, gultathion,

Dose dependently. Vitamin E used as nutritional supplement, it is a powerful biological antioxidant and is

the principle membrane-associated antioxidant molecule in mammals. It plays a major role in preventing

oxidative damage by scavenging free radicals.











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EFFECT OF VITAMIN E ON ATORVASTATIN …ijtpls.com/.../2015/12/IJTPLS-2015-Vol-15-PRASANNA-KUMAR-7-600-612.pdfPrasanna Kumar B et al. Int J Trends in Pharm & Life Sci. 2015: 1(5);600-612