Urea Cycle Lab Report

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Abstract: Ammonia is toxic to the human body and the complex process of removing it from the body is called the urea cycle. This experiment focuses on the intermediates and enzymes within the urea cycle This was done through spectrophotometry and assay techniques which helps show to us the varying intermediates and key steps involved in the urea cycle and allowed us to determine the activities of the proteins and enzymes and identify the proteins that do cause the production of urea in the cycle. Introduction: Ammonia and Amonium (NH3- and NH4+) are produced within the body and are highly toxic, especially to the brain and very reactive and so our bodies need a way of quickly processing this into a form for excretion, it does this by converting…show more content…
These five steps can be summarized as follows: 1. Formation of Carbamoyl Phosphate: Ammonia undergoes a condensation reaction with bicarbonate ions resulting in the formation of carbamoyl phosphate this reaction is then catalysed by the enzyme carbamoyl phosphate synthetase I, this step also requires Mg2+ and N-acetyl glutamate. 2. Synthesis of Citrulline Carbmoyal phosphate combines with ornithine by the transfer of a carbamoyl group to ornithine to form citrulline this is in the presence of enzyme citrulline synthase or ornithine transcarbamoylase and then diffuses through the mitochondrial membrane and into the cytosol. Phosphate is produced as a byproduct. 3. Synthesis of Argino-succinate: Once in the cytosol of the hepatocyte citrulline will comine with the ammino acid aspartate forming aginino-succinate this is catalysed by the enzyme arginine-succinate synthase. This process requires energy in the form of ATP which will be hydrolysed resulting in AMP and the use of the cofactor Mg2+ 4. Cleavage of…show more content…
Cleavage of Arginine: Arginine is hydrolysed to form urea and ornithine, under the enzyme arginase. Through this step Ornithine is regenerated and the urea cycle is completed with the formation of Urea. The overall reaction of the urea cycle is as follows: 2NH3 + CO2 + Aspartate + 3ATP → Urea + Fumarate + 2ADP + AMP + PPi + 2Pi The main methods used were that of spectrophotometry, using light and standard curves. A urea enzyme assay was performed via separating the proteins so as to observe which of these proteins were specifically causing an effect on the absorbance and produce urea. Another technique used in the experiment was the generation of a protein standard so as to determine the concentration of protein in the liver extract mixture The objective of this experiment is that the urea cycle substrates are amino acids integrated in cycle and that specific proteins and enzymes must be present to continue on and produce urea. This lab was based on the assumption that all enzymes were present and functional and so ultimately we were looking at the intermediates in the cycle and how well the enzymes moved them through the cycle and produced urea. Materials and
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