The blank test tube will be added with 0.20ml 85% Sodium chloride Solution. Te blank test tube will be used as reference test tube. Tis will be the test tube for comparison purposes with the other test tubes containing the various reagents. Te blank test tube will be the first tube to be read in the spectrophotometer that will set back the reading to zero. Al the other test tubes would be read in comparison with the blank test tube’s reading to determine significant results and readings. Te the test tubes would be added with certain volume of reagents such as Biuret reagent and Folin and Ciucalteu’s reagent, mxed and allowed to stand for different holding times.
Te contents of the test tubes would then be read with the absorbance with the blank solution as reference. Te protein concentration of every diluted sample would be determined. Te use of Biuret Method is greatly used for protein analysis. Buret is a compound formed from combination of urea through heat producing amide groups. Apeptide bond results with the amide the biuret reagent and the carboxyl group.
Tis would result to blue color solution observable through spectrophotometer in the presence of copper ion complex (Biuret Protein Assay, 2010). Te assay would greatly result to colored solution in alkali environment (Detection and Assay Methods, 1995). Oe of the reagents used in the experiment is the Folin and Ciocalteu’s Phenol reagent. Tis mixture of phosphomolybdate and phosphototungstate (MP Biomedicals, 2014) is useful a reagent commonly used for portein concentration determination through the Lowry method. Te addition of this reagent to previously diluted sample generates chromogens that increase absorbance to 550-750 nm.
Clor intensity is affected by the presence of tryphtophan and tyrosine (Sigma-Aldrich, 2014). Uder alkaline conditions, amonovalent ion and radical groups of tryhtophan and tyrosine reacts with the Folin reagent turning the solution into tungsten blue that can be viewed on spectrophotometer (Caprette, 2012). Bsed on the results it can be observed that the absorbance and protein concentration is linear in nature. Wth the use of various reagents, i can be deduced that the methodology used getting data protein content can be translated into graphical representation.
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