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In Vivo Analysis of Bisphenol Asian Journal of Pharmacy and Pharmacology 2019; 5(S1): 28-36 28 Research Article In vivo analysis of bisphenol A-induced sub-chronic toxicity on reproductive accessory glands of male mice and its amelioration by quercetin Sanman Samova, Hetal Doctor, Dimple Damore, Ramtej Verma Department of Zoology, BMTC and Human Genetics, School of Sciences, Gujarat University, Ahmedabad, India Received: 20 December 2018 Revised: 1 February 2019 Accepted: 25 February 2019 Abstract Objective: Bisphenol A is an endocrine disrupting chemical, widely used as a material for the production of epoxy resins and polycarbonate plastics. Food is considered as the main source of exposure to BPA as it leaches out from the food containers as well as surface coatings into it. BPA is toxic to vital organs such as liver kidney and brain. Quercetin, the most abundant flavonoid in nature, is present in large amounts in vegetables, fruits and tea. The aim of the present study was to evaluate the toxic effects of BPA in prostate gland and seminal vesicle of mice and its possible amelioration by quercetin. Material and methods: Inbred Swiss strain male albino mice were orally administered with BPA (80, 120 and 240 mg/kg body weight/day) for 45 Days. Oral administration of BPA caused significant, dose-dependent reduction in absolute and relative weights of prostate gland and seminal vesicle. Results and conclusion: Biochemical analysis revealed that protein content reduced significantly, whereas acid phosphatase activity increased significantly in prostate gland and reduction in fructose content was observed in seminal vesicle. Oral administration of quercetin (30, 60 and 90 mg/kg body weight/day) alone with high dose of BPA (240 mg/kg body weight/day) for 45 days caused significant and dose-dependent amelioration in all parameters as compared to BPA along treated group. Histopathological alterations were also ameliorated by quercetin. BPA causes toxicity in prostate gland and seminal vesicle of mice and quercetin is potent enough to ameliorate BPA induced toxicity. Keywords: Bisphenol A, male accessory glands, prostate gland, seminal vesicle, biochemical analysis, histopathology Introduction 2003; Bindhumol et al., 2003; Sangai et al., 2012) According Bisphenol A (2, 2-bis 4-hydroxyphenyl propane) is a to studies, even at very low doses, BPA mimic estrogen xenoestrogenic organic endocrine-disrupting chemical (Dodds resulting in an array of health maladies including prostate and Lawson, 1936) able to bind estrogen receptors (Takayanagi (Prins et al., 2007) as well as breast cancers (Pupo et al., et al., 2006; Chapin et al., 2008). It is one of the highest volume 2012), genotoxic effect (Karim and Husain, 2010) and other chemical produced worldwide with a total of 2.8 million metric health problems. (Vom Saal et al., 2007). tons manufactured in 2002 that increased to 5.5 million metric Quercetin, the most abundant flavonoid in nature, is present tons in 2011 (Rochester, 2013). A large body of data has shown in large amounts in vegetables, fruits, tea and olive oil, and that BPA, is mainly used for the production of polycarbonate contains a number of phenolic hydroxyl groups. (Wach et al., plastics, epoxy resins and non-polymer additives (Kuruto-Niwa 2007) It was reported that quercetin has many beneficial et al., 2007; Grumetto et al., 2008). Food is considered as a main effects in human health, including cardiovascular protection, source of exposure to BPA as a consequence of its migration (Grassi et al., 2010) anti-canceractivity, (Sak, 2014) anti- from food containers. (Ranjit et al., 2010) Bisphenol-A is toxic ulcer effects, (Suzuki et al., 1998) anti-allergy activity, to vital organs such as liver, kidney and brain. (Kabuto et al., (Bellik et al., 2013) cataract prevention, (Cornish et al., 2002) antiviral activity and anti-inflammatory effects. (Guardia et *Address for Corresponding Author: al., 2001) These effects of quercetin are due to its antioxidant Sanman Samova properties of scavenging free radicals directly. (Afanas' et al., Department of Zoology, University School of Sciences, Gujarat University, Ahmedabad- 1989; Robak et al., 1988) 380009, India The majority of studies on BPA have focused on the potential E-mail: [email protected] adverse effects on vital organs. However, there is insufficient DOI: https://doi.org/10.31024/ajpp.2019.5.s1.2 2455-2674/Copyright © 2019, N.S. Memorial Scientific Research and Education Society. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). www.ajpp.in Asian Journal of Pharmacy and Pharmacology 2019; 5(S1): 28-36 29 data available on the effect of BPA on the accessory reproductive Dose selection organs especially of males in mammals. The aim of the present Different doses of BPA was selected on the bases of LD50 study was to evaluate the toxic effects of BPA in prostate gland value. (MSDS 2004) Animals of BPA-treated groups and seminal vesicle of mice and its possible amelioration by received three different doses of BPA, which is 1/10th , 1/20 th quercetin. th and 1/30 of LD50 (240,120 and 80 mg/kg bw/day Materials and methods respectively) for 45 days for the amelioration of BPA- Chemicals induced toxicity different doses of quercetin (30, 60 and 90 mg/kg bw/day) were selected on the basis of the study of Bisphenol A and quercetin was purchased from Hi Media Sangai and Verma (Sangai et al., 2012). Laboratories Pvt. Ltd., Mumbai, India and was of analytical grade. Olive oil was obtained from Figaro, Madrid, Spain. All Experimental design the other chemicals used in the present study were of AR grade Experimental protocol is shown in table 1. Mice were and procured from Hi Media Laboratories Pvt. Ltd., Sisco randomly divided into nine groups, each containing10 Research Laboratories Pvt. Ltd., Mumbai, India and Sigma- animals. Treatment schedule of the animals was as follows. Aldrich St. Louis, MO, USA. Animals from group I (untreated control) were kept Experimental animals untreated and given free access to feed and water. Group II (vehicle control) animals were administered with olive oil In the experiment, inbred healthy adult Swiss strain male albino (0.2 ml/animal/day), which has been used as vehicle to mice weighing 35-40 gm obtained from Cadila pharmaceutical dissolve BPA. Group III (antidote control) animals were Center, Ahmedabad, India, were kept in the Animal House of treated with quercetin (90mg/kg bodyweight/day), which Zoology Department of Gujarat University, Ahmedabad, India. has been used for amelioration of BPA-induced toxicity. They were housed in an air-conditioned room at a temperature of 25±2ºC and 50-55% relative humidity with a 12 h light/dark Animals of group IV, V and VI received three different cycle throughout the experiment. Animals were fed with doses of BPA (80, 120 and 240 mg/kg bw/day) for 45 days. certified pelleted rodent feed supplied by Amrut Feeds, Pranav However animals of group VII, VIII and IX received three Agro Industries Ltd., Pune, India and potable water ad libitum. different doses of quercetin (30, 60 and 90 mg/kg bw/day) All the experimental protocols were approved by the Committee along with high dose of BPA (240 mg/kg bw/day). All for the Purpose of Control and Supervision of Experiments on treatments were given orally using a feeding tube attached Animals (Reg. – 167/1999/CPCSEA), New Delhi, India. to hypodermic syringe. Animals were handled According to the guidelines published by On completion of treatment, the animals were sacrificed by Indian National Science Academy, New Delhi, India (1991). using anesthetic ether. The prostate gland and seminal Table 1. Experimental protocol Experimental Groups Numbers of animal Duration of treatment (Days) Day of Necropsy Control groups th I Untreated control 10 45 46 th II Vehicle control (0.2 ml olive oil/animal/day) 10 45 46 th III Antidote control (90 mg quercetin /kg body 10 45 46 weight/day) BPA -treated groups th IV BPA-Low dose (80 mg/kg body weight/day) 10 45 46 th V BPA-Medium dose (120mg /kg body weight/day) 10 45 46 th VI BPA-High dose (240 mg/kg body weight/day) 10 45 46 HD BPA (240 mg/kg body weight/day) + quercetin treated groups th VII BPA-HD + Q30 (30 mg/kg body weight/day) 10 45 46 th VIII BPA-HD + Q60 (60 mg/kg body weight/day 10 45 46 th IX BPA-HD + Q90 (90 mg/kg body weight/day) 10 45 46 www.ajpp.in Asian Journal of Pharmacy and Pharmacology 2019; 5(S1): 28-36 30 vesicle were isolated, blotted free from blood, weighted and used Statistical analysis for the biochemical analysis such as protein and ACP in prostate The results were expressed as the means ± SEM. The data and Fructose in seminal vesicle. Tissue was stored in 10% were statistically analyzed using one way Analysis of formalin for histopathological study. Variance (ANOVA) followed by Turkey's post hoc test in Analysis of biochemical parameters Graph pad prism 5 (graph pad, software, USA). Statistical Protein content in prostate gland was estimated by the method of significance was accepted with p< 0.05. Correlation Lowry et al. (1951) and expressed as mg/100 mg tissue weight. Coefficient was measured to estimate the strength of linear The acid phosphatase (ACP) activity in the prostate gland was association between two Variables. Pearson's correlation assayed by the method as described in Sigma Technical Bulletin analysis was used to find the correlation between vehicle (Sigma Technical Bulletin, MO, USA. The enzyme activity was control and BPA toxicant-treated groups. expressed as μmoles p-nitrophenol released/mg protein/30 min. Results The concentration of fructose in tissue of seminal vesicle was Absolute and relative weights estimated by the method of Foreman et al.
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