Extraction, Purification and Kinetic Study of Lactate Dehydrogenase of Male Chicken from Ebocha-oil Exploration Area, Nigeria

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Chimdi E. Esonu
G. O. C. Onyeze
Kizito M. E. Iheanacho
Linus N. Nwaogu
Simon-Peter Odirichukwu


Aim: This study focused on the extraction, purification and kinetic studies of lactate dehydrogenase of male chickens from Ebocha oil exploration area, Imo state, Nigeria.

Methods: Twenty-one apparently healthy mature (6-9 months) male chickens (Gallus domesticus) from Ebocha oil exploration area, Imo State, Nigeria were screened for lactate dehydrogenase activity, thus accessing the level of chronic cell exposure to gas flaring. Their thigh muscle tissues were severed and investigated for lactate dehydrogenase activity using the standard method and sodium pyruvate as the substrate. Lactate dehydrogenase was isolated and purified by ammonium sulphate precipitation, desalted by dialysis and then gel filtration.

Results: The enzyme activity increased with advancement in the purification steps and was maximum using dialysis. The values for the lactate dehydrogenase activities were 103.43±3.27 U/L, 279.50±5.38 U/L, 318.16±13.08 U/L, 100.47±2.59 U/L, with a purification fold of 1, 3.7, 6.24 and 2.55 for the purification steps respectively. Also, the values of the protein concentrations were 0.071 mg/ml, 0.050 mg/ml, 0.035 mg/ml and 0.027 mg/ml (values for the crude enzyme, ammonium sulphate precipitation, dialysis and gel filtration respectively). The enzyme showed optimal activity at pH range of 5.5-6.5 and temperature of 30ºC-40ºC. Using sodium pyruvate as the substrate, with a fixed enzyme volume, an increase in the concentration of substrate resulted in increase in enzyme activity until a saturation point 0.3mM was reached. The apparent Km and Vmax values obtained were 0.01 mM and 0.12 U/mg/min. The Lineweaver-burk plot of the partially purified enzyme gave real Km and Vmax values of 0.20 mM and 0.16 U/mg/min respectively.

Conclusion: Partial purification procedures and biochemical properties of lactate dehydrogenase, from the muscle tissues of male chickens of Ebocha origin, gives room for more investigation on the metabolic shift caused by chronic exposure of the environment, humans and livestock to gas flaring and petroleum exploration.

Lactate dehydrogenase, Gallus domesticus, purification, kinetic studies, Ebocha.

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How to Cite
E. Esonu, C., C. Onyeze, G. O., M. E. Iheanacho, K., N. Nwaogu, L., & Odirichukwu, S.-P. (2019). Extraction, Purification and Kinetic Study of Lactate Dehydrogenase of Male Chicken from Ebocha-oil Exploration Area, Nigeria. Asian Journal of Biochemistry, Genetics and Molecular Biology, 2(4), 1-12. https://doi.org/10.9734/ajbgmb/2019/v2i430065
Original Research Article


Omoniyi OA, Ubale M. The impact of gas flaring in Nigeria. International Journal of Science, Technology and Society. 2015;3 (2):40-50.

Human Development Report.
Available:http://hdr.undp.org/en/ [Accessed: 29.7.2012]

Ayoola TJ. Gas flaring and its Implication for Environmental Accounting in Nigeria. J. Sustainable Develop. 2011;4(5):244‐248.

Barmidele J, Agboidi M. Toxicology of Odidi petroleum oil and its water soluble fraction on three aquatic macrophytes. Nigerian Journal of Science and Environment. 2000;2:113-114.

Lawanson TO. Challenges of sustainability and urban development in Nigeria: Reviewing the Millenium development goals. Africa Insight. 2006;1-19.

Kori-Siakpere O. Petroleum induced attuation in the African catfish (Clarias gariepinus). Nigerian Journal of Science and Environment. 1998;49:55-59.

Odiete WO. Environmental physiology of animals and pollution Diversified Resources Ltd, Lagos. 1997;50-52

Oluwasoye PM, Ugonna CN. The impacts of gas flaring and climate risks: An appraisal of Nigerian petroleum industry. International Journal of Scientific & Engineering Research. 2014;5(4):2229-5518.

Nwaogu LA, Onyeze CE, Alisi CS, Ijeh II, Onyeze GOC. Petroleum hydrocarbon induced changes in tissues of the native chicken (Gallus domesticus) following chronic exposure. Nigerian Journal of Biochemistry and Molecular Biology. 2008; 23(1):42-46.

Nwaogu LA, Onyeze GOC. Effect of chronic exposure to petroleum hydrocarbon pollution on oxidative stress parameters and histology of liver tissues of native chicken (Gallus domesticus). International Journal of Biochemistry Research & Review. 2014;4(3):233-242.

Drent M, Cobben NAM, Henderson RF, Wouters EFM, Dieijen VM. Usefulness of lactate dehydrogenase and its isoenzymes as indicators of lung damage or inflammation. Magazine. 1996;50-53.

Airaodion AI, Ogbuagu U, Ekenjoku JA, Ogbuagu EO, Airaodion EO, Okoroukwu VN. Hepato-protective efficiency of ethanol leaf extract of Moringa oleifera against hydrocarbon exposure. International Journal of advances in Herbal and Alternative Medicine. 2019;3(1):32-41.

Yannis K. Purification and characterization of lactate dehydrogenase: An undergraduate laboratory experiment. Advances in Biochemistry. 2014;2(1):14-23.

Wingfield P. Protein precipitation using ammonium sulfate. Current protocols in protein science. Appendix. 2001;3(1): Appendix3F.

Brooks DE, Haynes CA Hritcu D. Size exclusion chromatography does not require pores. Proceedings of the National Academy of Sciences of the United States of America. 2000;97(13):7064–7067.

Danson M, Eisenthal R. Enzyme assays: A practical approach. Oxford. Oxford University Press; 2002.


Bergmeyer HU. Methods of Enzymatic analysis. 4. New York: Academic Press. 1974;2066–72.
[ISBN 0-89573-236-X]

Airaodion AI, Akinmolayan JD, Ogbuagu EO, Esonu CE, Ogbuagu U. Preventive and therapeutic activities of methanolic extract of Talinum triangulare leaves against ethanol-induced oxidative stress in Wistar rats. International Journal of BioScience and BioTechnology. 2019;11 (7):85-96.

Theresa P. Specific activity and its importance in protein isolation.

Burgess RR. Protein precipitation techniques. Methods in Enzymology. 2009; 463:331–342.

Diao M, Aekoue B, Dibala CI, Dabonne S, Dicko MH. Purification and charaterisation of sweet potato peroxidase. Journal of Animal and Plant Sciences. 2014;22(2): 3419-3432.

Zia MA, Kousar M, Ahmed I, Igbal HMN, Abbas RZ. Comparative study of peroxidase purification from apple and orange seeds. African Journal of Biotechnology. 2011;10(33):6300-6303.

Pandey VP, Dwivedi N. Purification and characterization of peroxidase from L. leucocephala, a tree legume. Journal of Molecular Catalysis B. Enzymatic. 2011; 68:168-173.

Duong-Ly KC, Gabelli SB. Salting out of proteins using ammonium sulfate precipitation. Methods in Enzymology. 2014;541:85–94.

Onyeze GOC, Nwaogu LA, Awa ME. Chronic gas flaring pollution-induced alterations in kinetic parameters of lactate dehydrogenase in Gallus Domesticus Native to Ebocha, Niger Delta, Nigeria. Futojnls. 2016;2(2):98–109.

Daniel RM, Peterson ME, Danson MJ. The molecular basis of the effect of temperature on enzyme activity. Biochem. J. 2010;425(2):353–360.

Martinek R. Practical clinical enzymology. J. Am. Med. Tech. 1969;31:162.

Mitchell P, Yen HC, Mathemeier PF. Properties of lactate dehydrogenase in a psychrophilic marine bacterium. Applied and Environmental Microbiology. 1985;49 (5):1332-1334.

Yang CP, Schwert GW. Lactate dehydrogenase. The Journal of Biological Chemistry. 1970;245(19):4886-4893.

Masood HJ, Syed MI, Abida NH, Asifa A, Mohammed I. Purification and characterization of lactate dehydrogenase from Varanus liver. Experimental and Molecular Medicine. 1997;29(1):25-30.

Arriaga DD, Soler J, Cadenas E. Influence of pH on the allosteric properties of lactate dehydrogenase activity of Phycomyces blakesleeanus. Biochemical Journal. 1982; 203(2):392-400

Armstrong J, Coates JH, Morton RT. Characterization of lactate dehydrogenase from baker’s yeast. Biochem J. 1963;88: 266.

Cox MM, Nelson DL. Chapter 14: Glycolysis, Gluconeogenesis and the Pentose Phosphate Pathway. Lehninger Principles of Biochemistry (5th ed.). W H Freeman & Co. 2008;527-568.