Alteration in hepatic enzyme activity of Tilapia mossambica Vol. 9 (1), July 2018 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 30 R A D S J . B i o l . R e s . A p p l . S c i . 30 Op en Ac ces s F u l l L e n g t h A r t i c l e Alteration in Hepatic Enzyme Activity of Tilapia mossambica upon Exposure to Fluoride Farha Aziz*, Afshan Zeeshan Wasti and Farah Jabeen Department of Biochemistry, Jinnah University for Women, Karachi, Pakistan A B S T R A C T Fluoride, as the super reactive element fluorine, is found naturally throughout earth's crust. It has been identified as a strong, persistent powerful cumulative toxic agent, commonly distributed in the rivers, lakes, seas of earth. It is highly mobile and biologically active element in aquatic systems. Fish are considered as the excellent and valuable bioindicator of ecosystem pollution. The present study was designed to estimate acute fluoride toxicity on enzyme activity of liver of freshwater fish Tilapia mossambica. The major enzymes of Carbohydrate - Protein Metabolic pathways are Alkaline phosphatase (ALP), Alanine transaminase (ALT), Aspartate transaminase (AST) in association with carbohydrate, protein, lipid in the liver of Tilapia mossambica, from Kalri Lake (Keenjhar Lake), Sindh, Pakistan at low amount of fluoride (sub-lethal) was estimated by using UV- Visible Spectrophotometer. Results showed ALP, AST and ALT enzymes present in the liver tissue were significantly changed (p < 0.001). Finally, it is concluded that fluoride produces the adverse poisonous effect on liver functioning which may be associated with altered or elevated enzyme activity of protein-carbohydrate metabolism. Keywords: Fluoride, Carbohydrate-Protein Metabolic pathways, Alkaline phosphatase (ALP), Alanine transaminase (ALT), Aspartate transaminase (AST), Tilapia mossambica, Liver, Keenjhar Lake. *Address of Correspondence: farahtariq2@hotmail.com, farah786star@yahoo.com Article info. Received: April 18, 2018 Accepted: April 23, 2018 Cite this article: Aziz F, Wasti AZ, Jabeen F. Alteration in hepatic enzyme activity of Tilapia mossambica upon exposure to fluoride. RADS J. Biol. Res. Appl. Sci. 2018; 9(1): 30-34. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Funding Source: Nil Conflict of Interest: Nil I N T R O D U C T I O N The liver is a chief central metabolic organ, essential for life in vertebrates and invertebrates. The liver plays a main role in the metabolism of carbohydrate, protein, amino acid, and lipid. It acts as a detoxifying organ which may accumulate bio-transform and excretes a variety of toxicants or their by-products1,2. Toxicants can produce disorders in the physiology of animal in association with altered enzyme activity. Several studies reported the changes in enzyme activity in the freshwater fish when exposed to various pollutants3-6. Fluoride is highly mobile and biologically active element in aquatic systems. Fish are considered as an excellent and valuable bioindicator of ecosystem pollution. A number of cellular and biological processes including enzymatic reactions, cessation of protein secretion and synthesis, production of reactive oxygen species (ROS), and variation of gene expression can be affected by fluoride7,8. Fluoride as an anabolic agent or toxicant promotes cell proliferation and acts as an enzyme inhibitor for lipases, phosphatases and esterases, ATP production cycle and cellular respiration at the sublethal concentrations. Fluoride showed a significant alteration in enzymes activities of Alkaline phosphatase (ALP), Alanine transaminase (ALT), Aspartate transaminase (AST) in fish gills at the sublethal concentration of fluoride9,10. O R I G I N A L A R T I C L E Alteration in hepatic enzyme activity of Tilapia mossambica Vol. 9 (1), July 2018 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 31 R A D S J . B i o l . R e s . A p p l . S c i . 31 M A T E R I A L A N D M E T H O D S Normal alive Tilapia mossambica (weight 90.2g and length 8.7cm) were collected from the Keenjhar Lake, with the help of local fisherman. Fish were divided into two groups under normal condition. Group I was considered as control (non-treated) while group II marked as experimental treated with 1.5 gm NaF / 70 L of water. Each group contains twelve fishes and acclimated in a glass aquarium, filled with tap water. Aquarium was aerated by using air pumps and filters. The Commercial pellet as the fish food was used for both groups (I and II) per a day. Aquarium water was changed after 2 or 3 days. Chemical study of water was performed according to approved procedures of APHA11. Study of biochemical constituents and enzymatic assays Fishes of both groups (control and treated) were dissected and livers of all fishes were removed and cleaned with water. A glass homogenizer was used to homogenized tissues in cold solution of saline (0.89% NaCl) and centrifuged in a refrigerated laboratory centrifuge. A clear supernatant was used for Alkaline phosphatase (ALP) estimation by p-nitro phenol method However Aspartate transaminase (AST), Alanine transaminase (ALT) were measured by Randox Kit methods. Estimation of glucose was done by GOP-PAP method and protein by Lowry et al. method12,13. Estimation of total lipid by sulphophospho-vanilline (SPV) method14. Data were represented as Mean ± S.E.M. A paired student t-test at 95% confidence interval of the difference was applied to find out the level of significance. P value < 0.05 was considered significant. R E S U L T S A N D D I S C U S S I O N The effect of fluoride on important and essential enzymes of Carbohydrate - Protein metabolism were measured in the liver of Tilapia mossambica, at 1st, 2nd, 3rd and 4th week upon exposure to fluoride. Aminotransferases are important group of enzymes participating in the shifting of the amino group from alpha-amino acids to the alpha-keto acid without the liberation of ammonia. The alpha-keto acid enters into the citric acid cycle for the release of energy. In this process alpha-keto acids changed into amino acids might be used in protein synthesis and take part in carbohydrate-protein metabolism regulation 10, 15, 16. Alkaline phosphatase (ALP) removing phosphate groups from nucleotides, proteins, and alkaloids, a process called dephosphorylation. The results showed significant increase in the enzymatic action of Alkaline phosphatase (ALP), Alanine transaminase (ALT), Aspartate aminotransferase (AST) (Table 1, Fig. 1) in treated liver under fluoride when compare to control fish (p < 0.001) due to utilization of glucose and amino acid in carbohydrate-protein metabolism (Table 2)10,17-21. High levels of AST, ALT, ALP upon long-term exposure (4th week) to fluoride might be due to altered liver function or liver dysfunction under stress22, 23. Table 1: Enzymatic activity of AST, ALT, ALP in the liver of Tilapia mossambica at 1st, 2nd, 3rd and 4th week upon exposure to fluoride (1.5g/70L). Biochemical parameter (U/L) Control Fish Treated Fish 1st week Treated Fish 2nd week Treated Fish 3rd week Treated Fish 4th week AST 633.65±0.077 823.54±0.009 876.42±0.013 856.14±0.008 734.12±0.051 ALT 755.6±0.068 890.4±0.073 974.6±0.005 1005.6±0.009 798.34±0.034 ALP 308.4±0.090 677.13±0.007 790.85±0.007 859.26±0.007 467.47±0.074 Values expressed as Mean ± S.E.M; *: Significant at P< 0.05; **: Highly significant at P<0.01; ***; Very highly significant (p<0.001) compared with control Alteration in hepatic enzyme activity of Tilapia mossambica Vol. 9 (1), July 2018 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 32 R A D S J . B i o l . R e s . A p p l . S c i . 32 Fig. 1: Fluoride Effect (1.5g/70L) on the activity of ASP, ALT, ALP enzymes in the liver of Tilapia mossambica at 1st, 2nd, 3rd and 4th week. Table 2: Biochemical constituents Protein, Lipid, Cholesterol, Glucose in the liver of Tilapia massombica at 1st, 2nd, 3rd and 4th week upon exposure to fluoride (1.5g/70L). Biochemical parameter (mg/g) Control Fish Treated Fish 1st week Treated Fish 2nd week Treated Fish 3rd week Treated Fish 4th week Protein 6.745±0.089 4.553±0.098 3.497±0.068 3.300±0.067 2.756±0.023 Lipid 86.063±0.018 72.55±0.008 48.4±0.105 52.637±0.056 49.327±0.016 Cholesterol 82.805±0.078 308.612±0.066 177.6±0.077 131.625±0.055 101±0.078 Glucose 63.418±0.084 37.418±0.037 32.495±0.050 12.947±0.008 10.650±0.097 Values expressed as Mean ± S.E.M; **: Highly significant at P<0.01; ***; Very highly significant (p<0.001) compared with control Fig. 2: Fluoride Effect (1.5g/70L) on Protein, Lipid, Cholesterol Glucose in the liver of Tilapia mossambica at 1st, 2nd, 3rd and 4th week. Alteration in hepatic enzyme activity of Tilapia mossambica Vol. 9 (1), July 2018 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 33 R A D S J . B i o l . R e s . A p p l . S c i . 33 Quantitative measurements of biochemical constituents in the liver of freshwater fish Tilapia massombica at 1st, 2nd, 3rd and 4th week are presented in Table 2 and Fig. 2. Results showed decreased in protein-carbohydrate and lipid contents due to changes in carbohydrate–protein metabolism (p < 0.001). This condition might be due to the decrease oxidation of glucose while decreased in protein might be associated with energy production. The whole process is needed to overwhelm the tension for the endurance of fish24. Decreased lipid content in fish liver exposed to fluoride may be due to the decrease in enzyme acyl co-A synthetase activity, play an essential role in fatty acid synthesis. An increase in the cholesterol level in the liver due to fluoride dose, measurements of these important clinical biochemical parameters serve as excellent biomarkers of the aquatic environmental condition. 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