|Year : 2011 | Volume
| Issue : 2 | Page : 163-166
Effect of N-phthaloyl gamma-aminobutyric acid on lipid peroxidation, antioxidants and liver markers in constant light exposed rats
Selvaraju Subash, Perumal Subramanian
Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalainagar, Tamilnadu, India
|Date of Submission||01-Jan-2011|
|Date of Acceptance||30-Mar-2011|
|Date of Web Publication||23-Aug-2011|
Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalainagar - 608 002, Tamil Nadu
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Introduction & Materials and Methods : The effect of N-Phthaloyl GABA on the levels of circulatory lipid peroxidation products such as TBARS (thiobarbituric acid reactive substances), antioxidants: SOD (superoxide dimutase), CAT (catalase), GSH (reduced glutathione), and GPx (glutathione peroxidase), liver markers such as AST (aspartate transaminase), ALT (alanine transaminase), and ALP (alkaline phosphatase), were studied for its protective effect during constant light (LL) exposure in rats. Results: A significant increase in the levels of circulatory TBARS, AST, ALT, and ALP were found in LL exposed rats. These changes were significantly decreased in N-Phthaloyl GABA and LL-exposed rats. The levels of antioxidants were significantly decreased in LL-exposed rats and administration of GABA normalized these changes. Conclusions: Our results indicated that NPhthaloyl GABA offered protection by influencing the levels of the lipid peroxidation products, antioxidants, and liver markers during LL-exposed stress. This might be due to the antistress, anticonvulsant, antinociceptive, and antiulcer activities of N-Phthaloyl GABA. The exact mechanism remains to be explored and further research is needed.
Keywords: Antioxidants, Constant light, gamma-aminobutyric acid, Liver markers, thiobarbituric acid reactive substances
|How to cite this article:|
Subash S, Subramanian P. Effect of N-phthaloyl gamma-aminobutyric acid on lipid peroxidation, antioxidants and liver markers in constant light exposed rats. Int J Nutr Pharmacol Neurol Dis 2011;1:163-6
|How to cite this URL:|
Subash S, Subramanian P. Effect of N-phthaloyl gamma-aminobutyric acid on lipid peroxidation, antioxidants and liver markers in constant light exposed rats. Int J Nutr Pharmacol Neurol Dis [serial online] 2011 [cited 2019 Jun 15];1:163-6. Available from: http://www.ijnpnd.com/text.asp?2011/1/2/163/84208
| Introduction|| |
Physiological and anatomical evidence suggests that amino acid transmitters carry the major excitatory / inhibitory commands in the central nervous system (CNS) and the other types of CNS transmitters play more diffuse modulatory roles. Gamma-aminobutyric acid (GABA) and glycine are the principal inhibitory transmitters, whereas, glutamate and aspartate are presumed to be the excitatory transmitters.  GABA plays a critical role in regulating synaptic activity within the CNS.  Evidence for GABA in the suprachiasmatic nucleus (SCN),  retina,  and ventral lateral geniculate nuclei  has been reported for a variety of mammals. It was earlier demonstrated that N-phthaloyl g-aminobutyric acid (P-GABA), a GABA mimic agent, possesses anticonvulsant,  antinociceptive,  and antiulcer activities , Many GABA analogs have been shown to prevent several manifestations of stress in experimental studies, for example, the GABAergic system in antistress activities. 
Light exposure at night seems to be associated with a number of serious behavioral as well as health problems, including cancer.  Results from the human experiments  have reported that night shift work (exposure to light at night) was associated with an increased risk of breast cancer as well as other cancers.  It was reported that constant light (LL) exposure disrupts the circadian rhythms in rodents, and results in increasing the circadian period, arrhythmicity (rats), and splitting of wheel-running activity rhythms (in hamsters). Previous studies show that the continuous exposure of bright light strongly suppresses circadian rhythms of the sleep-wake cycle, drinking, locomotion, and body temperature. 
To our knowledge, no biochemical investigations have been carried out on the effect of P-GABA on circulatory lipid peroxidation, liver markers, and antioxidant status in LL-exposed experimental rats. Therefore, we made an attempt to bridge the information gap; the present study was undertaken to investigate the effect of P-GABA on circulatory lipid peroxidation products, liver markers, and the antioxidant status in LL-exposed rats.
| Materials and Methods|| |
Adult male albino Wistar rats, weighing 160 - 180 g, bred in the Central Animal House, Rajah Muthiah Medical College, Annamalai University, were used. The animals were housed in polycarbonate cages in an experimental room, with 12 : 12 hours day-night cycle, a temperature of 22 ± 2°C, and humidity of 45 - 64%. The animals were fed a standard pellet diet (Hindustan Lever Ltd., Mumbai, India) and water ad libitum. The animal experiments were approved by the ethical committee (Vide. No. 244 / 2004), Annamalai University, and were in accordance with the guidelines of the National Institute of Nutrition (NIN), Indian Council of Medical Research (ICMR), Hyderabad, India. GABA and phthalic anhydride were obtained from Sigma Chemical Co., USA. All other chemicals used in this study were of analytical grade.
The animals were randomized and divided into four groups (n = 6 in each group). Group 1: Normal (untreated rats); Group 2: Constant light [12 : 12 hours (LL)]-exposed rats; Group-III: LL-exposed + P-GABA treated (50 mg / kg body Wt);  and Group - IV: P-GABA treated. At the end of the experimental period (21 days), the animals were killed by decapitation and blood samples were collected for the estimation of circulatory TBARS (thiobarbituric acid reactive substances),  reduced glutathione (GSH),  glutathione peroxidase (GPx),  superoxide dismutase (SOD),  catalase (CAT),  aspartate transaminase (AST), alanine transaminase (ALT),  and alkaline phosphatase (ALP). 
Data analysis was carried out by analysis of variance (ANOVA) and the groups were compared using the Duncan's Multiple Range Test (DMRT).
Preparation of N-Phthaloyl GABA (P-GABA)
An intimate mixture of 6.18 g GABA and 8.95 g of finely grounded phthalic anhydride was heated for 30 minutes, with stirring, in an oil bath at 145°-150°C. After cooling the solid material was dissolved in hot methanol. The filtrate was diluted with 20 ml of water and the product was allowed to crystallize. The product was identified by (i) measuring the melting point, (ii) UV absorption, and (iii) IR spectroscopy. 
| Results|| |
[Table 1] shows the levels of circulatory lipid peroxidation products (plasma TBARS) and liver markers (serum AST, ALT, and ALP) of the control and experimental groups. The levels of TBARS, AST, ALT, and ALP were significantly (P < 0.05) higher in LL-exposed rats. Treatment with P-GABA (50 mg / kg body Wt) given to LL-exposed rats, for 21 days, significantly (P < 0.05) decreased the levels of TBARS, AST, ALT, and ALP, when compared with LL-exposed rats.
|Table 1: Effect of P-GABA on changes in the levels of circulatory liver markers and TBARS in normal and experimental rats|
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[Table 2] shows the effect of P-GABA on circulatory antioxidants (SOD, CAT, GPx, and GSH) in normal and LL-exposed rats. Rats exposed to LL, exhibited a significant decrease in the levels of antioxidants (SOD, CAT, GPx, and GSH) when compared with normal rats. Administration of P-GABA to LL rats significantly (P < 0.05) increased the levels of SOD, CAT, GPx, and GSH in the circulation, when compared with LL-induced rats.
|Table 2: Effect of P-GABA on changes in the levels of circulatory antioxidants in normal and experimental rats|
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| Discussion|| |
Evidences point out that LL exposure leads to oxidative stress and free radical production-mediated lipid peroxidation.  Marked elevations in the concentration of TBARS have been observed in LL-exposed rats, in our study. It has been reported that continuous light can cause a formation of reactive radicals.  Previous studies have shown that healthy animals exposed to environmental factors such as oxygen or constant light display elevated malonaldialdehyde levels in the brain, liver, and kidney,  suggesting a generalized increase in free radicals under LL, prompted by the formation of photo-oxidants. It has been reported that continuous light enhances the release of glutamate, which activates NMDA receptors. This leads to neuronal degeneration and death, and involves the increase in Ca 2+ concentration in the postsynaptic neuron. Ca 2+ binds to calmodulin and activates nitric oxide synthase, increasing the formation of nitric oxide (NO), and other toxic radicals (hydroxyl radical, etc.) as well as thiobarbituric acid-positive compounds in the brain by inducing nitric oxide synthase. , This may result in increased lipid peroxidation in the circulation. The higher plasma lipid peroxide concentrations (in group II rats) in our study may be due to leakage of the lipid peroxidation products from the organ or tissues into the blood stream, under constant light condition. 
Administration of P-GABA significantly decreases the levels of TBARS in group III rats, which may be due to the anti-lipid peroxidative property of P-GABA.
In several organs, cell damage is followed by the release of a number of cytoplasmic enzymes into the blood, a phenomenon that provides the basis for clinical diagnosis.  In our study, the increased activities of AST, ALT, and ALP in the serum, obviously indicate that the liver is susceptible to LL-induced toxicity, and also this may be due to the liver damage caused by the LL-induced free radical generation. Serum AST, ALT, and ALP have been widely used as indices of liver dysfunction.  In group II rats, the significant elevation of serum AST, ALT, and ALP may be related to increased oxidative stress developed during LL exposure. P-GABA may stabilize the hepatic cellular membrane and protect the hepatocytes against the toxic effects of LL exposure, which may decrease the leakage of enzymes into the blood stream. The level of lipid peroxidation in the cells is controlled by various cellular defense mechanisms consisting of enzymatic and non-enzymatic scavenger systems,  the levels of which are altered in LL-exposed conditions.  This may have decreased the levels of antioxidants such as, SOD, CAT, GPx, and GSH in the circulation of LL-exposed rats. This may be due to the overutilization of antioxidants to scavenge the LL-mediated lipid peroxidation. In our investigations, the levels of antioxidants, which declined in LL-exposed animals, were significantly restored on treatment with P-GABA. , Significant elevation in the levels of these antioxidants, in continuous light-exposed rats treated with P-GABA, shows the potential oxidant defense of P-GABA.
| Conclusions|| |
Due to LL exposure, stress-mediated free radical generation can occur, which can cause increased TBARS, liver markers, and depleted antioxidants. Administration of P-GABA to LL-exposed rats normalizes the levels of the lipid peroxidation products, liver marker enzymes, and antioxidants. This may be due to the antistress, antiperoxidative, anticonvulsant, antinociceptive, and antiulcer activities, and the antioxidant properties of P-GABA. The exact mechanism remains to be explored and further research is needed.
| References|| |
|1.||McGeer PL, Eccles JC, McGeer EG. Molecular neurobiology of the mammalian Brain. New York: Plenum Press; 1987. p. 553-94. |
|2.||Curtis DR, Johnston GA. Amino acid transmitters in the mammalian central nervous system. Ergebn Physiol 1974;69:97-188. |
|3.||Card JP, Moore RY. Neurotransmitter and peptide localization in rat suprachiasmatic nucleus. Neurosci 1983;9:1069. |
|4.||Brandon C. Retinal GABA neurons: Immunocytochemical localization using a new antiserum against rat brain glutamate decarboxylase. Soc Neurosci 1983;9:800. |
|5.||Hendrickson AE, Ogren MP, Vaughn Je, Barber RP, Wu JY. Light and electron microscope immunocytochemical localization of glutamic acid decarboxylase in monkey geniculate complex: Evidence for GABaergic neurons and synapses. J Neurosci 1983;3:1245-62. |
|6.||Bhowmick S, Pal M, Pel SP. Synthesis and anticonvulsant activity of N-Phthaloyl GABA- a new GABA derivative. Indian J Exp Biol 1989;27:805-8. |
|7.||Banerji S, Bhowmick S, Bera M, Pal M, Pal SP. Antinociceptive action of GABA-mimetic agent-N-Phthaloyl GABA. Indian J Exp Biol 1991;6:538-42. |
|8.||Bhowmick S, Bose R, Pal M, Pal SP. Antiulcer activity of N-phthaloyl GABA-a new GABA mimetic agent. Indian J Exp Biol 1990;28:190-2. |
|9.||Banerji S, Habibuddin M, Pal SP. Antiulcer and gastric secretary activity of N-Phthaloyl gamma-aminobutyric acid. Eur J Pharmacol 1992;25:211-5. |
|10.||Kulkarni SK, Kunchandy J. Brain and psychophysiology of stress, In: Sharma KN, Selvamurthy W and Bhattacharya V, editors. Indian Council of Medical Research, New Delhi. 1998;191:1998. |
|11.||Levi. Circadian chronotheraphy for human. Cancer oncology 2001;2:307-15. |
|12.||Rafnssion V, Tulinius H, Jonasson JG, Hrafnkelsson J. Risk of breast cancer in female flight attendants: A population - based study (Iceland). Cancer Causes Control 2001;12:95-101. |
|13.||Kerengi NA, Pandula E, Feuer GM. Oncostatic effects of the pineal gland. Drug Metabol Drug Interact 1990;8:313-9. |
|14.||Depres-Brummer P, Levi F, Metzger G, Touitou Y. Light-induced suppression of the rat circadian system. Am J Physiol 1995;268:1111-6. |
|15.||Subramanian P, Sundaresan S, Balamurugan E. Temporal oscillations of phosphatases in N-Phthaloyl gamma-aminobutyric acid treated rats. Indian J Exp Biol 1998;36:1141-3. |
|16.||Nichans WG, Samuelson B. Formation of malondialdehyde from phospholipid arachidonate during microsomal lipid peroxidation. Eur J Biochem 1986;6:126-30. |
|17.||Ellman GL. Tissue sulphydryl groups. Arch Biochem Biophys 1959;82:70-7. |
|18.||Rotruck JT, Pope AL, Ganther HE, Swanson AB, Hafeman BD, Hoekstra WG. Selenium: Biochemical roles as components of glutathione peroxidase. Science 1973;179:588-90. |
|19.||Kakkar P, Das B, Viswanathan PN. A modified spectrophotometric assay of superoxide dismutase. Ind J Biochem Biophys 1984;22:130-2. |
|20.||Sinha KA. Colorimetric assay of catalase. Anal Biochem 1972;47:389-94. |
|21.||Reitman S, Frankel S. A colorimetric method for the determination of serum glutamic oxalacetic and glutamic pyruvic transaminases. Am J Clin Pathol 1957;28:56-63. |
|22.||King EJ, Armstrong AR. Determination of serum and bile phosphatase activity. Canad Med Assoc J 1934;31:56-63. |
|23.||Baydas G, Ercel E, Canatan H, Donder E, Akyol A. Effect of melatonin on oxidative status of rat brain, liver and kidney tissues under constant light exposure. Cell Biochem Funct 2001;19:37-41. |
|24.||Hardeland R, Poeggelar B, Balzer I, Behrmann G. A hypothesis on the evolutionary origins of photoperiodism based on circadian rhythmicity of melatonin in phylogenetically distant organisms. Chronobiology and chronomedicine, In: Guttenbrunner C, Hidtebrant G, Moog R, editors. Frankfurt: Peter Long verlag; 1993. p. 113-20. |
|25.||Ding JM, Chen D, Weber Et, Faiman LE, Rea MA, Gillette MU. Resetting the biological clock: Mediation of nocturnal circadian shifts by glutamate and NO. Science 1994;266:1713-6. |
|26.||Kosenko E, Kaminsky A, Valencia M, Lee L, Hermenegildo C, Felipo V. Superoxide production and antioxidant enzymes in ammonia intoxication in rats. Free Radic Res 1997;27:637-44. |
|27.||Sundberg A, Appelkwist EL, Dallner G, Nilsson R. Glutathione transferase in urine: Sensitive methods for detection of kidney damage induced by nephrotoxic agents in human. Environ Health Perspect 1994;102:293-6. |
|28.||Lee DH, Blomhoff R, Jacobs JD. Is serum gamma glutamyltransferase a marker of oxidative stress? Free Radic Res 2004;38:535-9. |
|29.||Halliwell B, Gutterridge JM. Lipid peroxidation, oxygen radicals, cell damage and antioxidant therapy. Lancet 1994;1:1396-7. |
|30.||Subash S, Subramanian P, Sivaperumal R, Manivasagam T, Mohamed Essa M. Constant light influences the circadian oscillations of circulatory lipid peroxidation, antioxidants and some biochemical variables in rats. Biol Rhythm Res 2006;37:471-7. |
|31.||Subash S, Subramanian P. Influence of N-Phthaloyl GABA on the circadian rhythms of lipid peroxidation and antioxidants in Wistar rats under constant light. Iran J Pharmacol Therap 2007;6:115-8. |
|32.||Subash S, Subramanian P. Influence of N-phthaloyl gamma-aminobutyric acid on circadian rhythms of lipid peroxidation products and antioxidants. Biol Rhythm Res 2008;39:3-7. |
[Table 1], [Table 2]