1. Introduction
pioids exert their pharmacological effects by activating opioid receptors [1]. As a μ receptor agonist, morphine is the most important opium alkaloid. So far, many pharmacological properties of morphine have been reported [2]. By affecting μ receptors, morphine inhibits the release of various neurotransmitters such as noradrenaline and acetylcholine [3]. Oxidative stress is the impairment of oxidant-antioxidant balance. Biosystems have defense mechanisms that help protect against cellular damage caused by free radicals [4]. Ziziphus jujuba, commonly known as Jujube, has antioxidant activity due to its compounds such as saponins, flavonoids, and jujubosides [7, 6, 5]. This study aims to evaluate the effect of jujube extract on the activity of Acetylcholinesterase (AChE) enzyme and oxidative stress in the cortex and serum of morphine-treated male rats.
2. Materials and Methods 
In this experimental study, 42 male Wistar rats in the weight range of 200-250 g were purchased from the Pasteur Institute of Amol Research Center and kept in standard conditions (12 hours of light and 12 hours of darkness, a humidity of 40% -50% and a temperature of 25 °C). All experiments were performed according to the guidelines of the Bioethics Committee of the University of Mazandaran. Jujube fruit was collected, dried and powdered. Then 500 ml of ethanol was added to100 g of the powder and put in a shaker for 4 days in darkness. The mixture was then filtered by using Whatman filter paper. The obtained extract was dissolved and concentrated by a rotary evaporator [8].
Animals were divided into six groups of 7 including: Control group (received normal saline by gavage intraperitoneally), morphine group (received normal saline by gavage plus 0.5 mg/kg morphine 30 minutes later intraperitoneally) [8], jujube 100 and 200 groups (received 100 and 200 mg/kg body weight of jujube extract by gavage plus normal saline 30 minutes later intraperitoneally) [9], jujube 100 + morphine and jujube 200 + morphine groups (received 100 and 200 mg/kg body weight of jujube extract by gavage plus morphine 30 minutes later intraperitoneally). All treatments were performed for 30 days. The last gavage for each rat was 24 hours before anesthesia. At the end of treatment (day 31), following blood collection, separation of serum from blood samples, and homogenization of brain tissue, the activity of Catalase (CAT), AChE, and Superoxide Dismutase (SOD) was assessed. In order to statistically analyze the collected data and compare groups, one-way ANOVA and Tukey’s post hoc test were used considering a significance level of P<0.05. 
3. Results
The results of ANOVA showed significant changes in the serum and cortical AChE in the study groups (P<0.001). Serum AChE activity significantly decreased in the morphine group compared to the control group (P<0.001). Administration of 100 and 200 mg/kg extracts of jujube plus morphine injection increased the activity of serum AChE compared to when only morphine was injected, but this increase was not significant. However, administration of 100 mg/kg (P=0.004) and 200 mg/kg (P=0.036) extracts showed significant changes compared to the control group. Cortical AChE activity showed a significant decrease in the morphine group (7.02±0.61) compared to the control group (13.56±0.73) (P<0.001). Administration of 100 and 200 mg/kg jujube extracts plus morphine injection increased the activity of cortical AChE compared to the morphine group, but the increase was significant only at a dose of 200 mg/kg (P=0.002).
Cortical CAT activity was significantly decreased in the morphine group (8.09±0.91) compared to the control group (20.79±1.31) (P<0.001). Administration of 100 and 200 mg/kg extracts of jujube plus morphine injection increased the cortical CAT activity compared to the morphine group but this increase was not significant (P>0.05). However, significant changes were observed compared to the control group (P=0.005 and 0.027). Serum CAT activity in the morphine group significantly decreased compared to the control group (P<0.001). Administration of 100 mg/kg (P= 0.015) and 200 mg/kg (P=0.017) extracts of jujube plus morphine injection increased the serum CAT activity compared to when only morphine was injected, but this increase was not significant. However, compared to the control group, the differences were significant.
Finally, the results of ANOVA showed significant changes in the serum and cortical SOD activities in the study groups (P= 0.001). Cortical and serum SOD activities in the morphine group significantly decreased compared to the control group (P=0.001 and 0.011, respectively). Administration of 100 and 200 mg/kg jujube extracts alone increased the activity of cortical SOD compared to when only morphine was injected (P<0.01). The morphine group after receiving jujube extracts increased the inhibition of cortical SOD, but the increase in SOD activity was significant only in the jujube 200 + morphine group (P=0.013). 
4. Discussion and Conclusion
Intraperitoneal injection of morphine increases oxidative stress and reduces the activity of CAT, SOD, and AChE. Administration of jujube extracts improves morphine-induced oxidative stress and increases AChE activity. Therefore, jujube extract improves the activity of CAT, SOD, and AChE in morphine-treated male rats based on its antioxidant properties. The consumption of jujube fruit can prevent the reduction of oxidative stress and its complications.

Ethical Considerations
Compliance with ethical guidelines
This study was approved by the Bioethics Committee of the University of Mazandaran (Code: IR.UMZ.REC.1398.003).

Funding
This article was extracted from the MA. thesis of first author at the Department of Biology, Faculty of Basic Sciences, University of Mazandaran.

Authors' contributions
Conceptualization: Bagher Seyedalipour, Farhad Valizadegan; Methodology, investigation, resource, writing-original draft: Zahra Haratian; Visualization, data analysis, writing–review & editing, supervision, project administration: Bagher Seyedalipour, Farhad Valizadegan.

Conflicts of interest
The authors declare no conflict of interest.

Acknowledgements
 The authors would like to thank the Vice Chancellor for Research of Mazandaran University for his financial support and Ms. Rezaei for his experimental support. 


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