User:Javier Vinals Camallonga/Notebook/Javier Vinals notebook/2014/03/26: Difference between revisions

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==Objective==
==Objective==
Today we'll be determining the molar absorptivities of two different molecules, [http://en.wikipedia.org/wiki/Adenosine adenosine] and [http://en.wikipedia.org/wiki/Inosine inosine]. The data that we generate today will be important when we study [http://en.wikipedia.org/wiki/Adenosine_deaminase adenosine deaminase] (ADA), which converts adenosine to inosine. The difference between these two molecules is that adenosine contains a primary amine whereas inosine contains a carboxy group. Overexpression of this protein causes anemia in humans. A shortage of this protein can lead to severe immuno-defficiency.  
* Run UV-Vis and AA on room temperature samples from 3/25/14.
* Synthesize more 30:1 lysozyme-AuNPs to repeat testing from yesterday.


Adenosine and inosine have different absorption spectra. We will be observing changes in UV-Vis spectra to determine changes in concentration of both adenosine and inosine. In order to do this, we will need to know the molar absorptivity (ε) of both of these molecules. Just as each molecule has a characteristic absorption at each wavelength, this (per-wavelength) absorption can be quantified by a molar absorptivity. Or ... for a given concentration a molecule will absorb a very specific amount of light at a precise wavelength. A molecule doesn't have just one molar absorptivity; there is a molar absorptivity to describe each wavelength in a molecular absorbance spectrum.
==Procedure==
===Lysozyme-AuNPs===
'''Creating Stock Solutions of Gold and Lysozyme'''
* As seen below, each stock solution was created following the calculations in the table.
* Each solution was created using a volumetric flask and deionized water.  
 
[[Image:3.26.stock.png|650px|]]
 
 
'''Synthesizing Lysozyme-AuNPs'''
* As seen below, each solutions was created using the following calculations.
* We made 66 tubes total.
* Note: Instead of adding each amount into the tubes individually, we added the total volumes for 66 tubes into a beaker. We then measured 5 mL, which were transferred to glass test tubes.
 
[[Image:3.26.data.png|650px|]]
 
===Atomic Absorption===
 
'''Creating the Gold Stock Solutions'''
# Add 50 μL of HAuCl<sub>4</sub>·3H<sub>2</sub>O and 4950 μL of distilled water to a Falcon tube, for a final concentration of 10 μg/mL Au.
# Add 100 μL of HAuCl<sub>4</sub>·3H<sub>2</sub>O and 4900 μL of distilled water to a Falcon tube, for a final concentration of 20 μg/mL Au.
# Add 150 μL of HAuCl<sub>4</sub>·3H<sub>2</sub>O and 4850 μL of distilled water to a Falcon tube, for a final concentration of 30 μg/mL Au.
# Add 200 μL of HAuCl<sub>4</sub>·3H<sub>2</sub>O and 4800 μL of distilled water to a Falcon tube, for a final concentration of 40 μg/mL Au.
# Add 250 μL of HAuCl<sub>4</sub>·3H<sub>2</sub>O and 4750 μL of distilled water to a Falcon tube, for a final concentration of 50 μg/mL Au.
 
 
 
'''Atomic Absorption Samples'''
 
Solutions with the following Au:lysozyme ratio were run on the AA:
* 30:1 lysozyme-AuNP with 0.03 M MgCl<sub>2</sub>, CaCl<sub>2</sub>, NaCl, KCl, MES, citric acid (0.0002316 M 2,2 bipyridine)
* 30:1 lysozyme-AuNP with 0.06 M MgCl<sub>2</sub>, CaCl<sub>2</sub>, NaCl, KCl, MES, citric acid (0.0004544 M 2,2 bipyridine)
* 30:1 lysozyme-AuNP with 0.09 M MgCl<sub>2</sub>, CaCl<sub>2</sub>, NaCl, KCl, MES, citric acid (0.0006772 M 2,2 bipyridine)
 
==Figures==
[[Image:3.26.aa.png|750px|]]
*This graph shows relatively constant amounts of gold present for all variables added. MES overall had the least amount of gold present in solution in comparison to other variables. Citric acid was the only variable that a visual reduction in the amount of gold in solution as the concentration of variable increased.
 
 
 
 
[[Image:Screen_Shot_2014-03-31_at_11.28.08_PM.png]]
[[Image:Screen_Shot_2014-03-31_at_11.28.28_PM.png]]
*No 30:1 lysozyme-AuNP control was run because no samples were leftover from this batch of AuNP's. A single tube was made, but a mistake was made in its preparation causing fibers to form.
*This UV-Vis corroborates all of the data presented in the AA by showing a change in absorbance as concentrations of gold change.




Revision as of 10:42, 2 April 2014

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Entry title

  • Insert content here...

Objective

  • Run UV-Vis and AA on room temperature samples from 3/25/14.
  • Synthesize more 30:1 lysozyme-AuNPs to repeat testing from yesterday.

Procedure

Lysozyme-AuNPs

Creating Stock Solutions of Gold and Lysozyme

  • As seen below, each stock solution was created following the calculations in the table.
  • Each solution was created using a volumetric flask and deionized water.


Synthesizing Lysozyme-AuNPs

  • As seen below, each solutions was created using the following calculations.
  • We made 66 tubes total.
  • Note: Instead of adding each amount into the tubes individually, we added the total volumes for 66 tubes into a beaker. We then measured 5 mL, which were transferred to glass test tubes.

Atomic Absorption

Creating the Gold Stock Solutions

  1. Add 50 μL of HAuCl4·3H2O and 4950 μL of distilled water to a Falcon tube, for a final concentration of 10 μg/mL Au.
  2. Add 100 μL of HAuCl4·3H2O and 4900 μL of distilled water to a Falcon tube, for a final concentration of 20 μg/mL Au.
  3. Add 150 μL of HAuCl4·3H2O and 4850 μL of distilled water to a Falcon tube, for a final concentration of 30 μg/mL Au.
  4. Add 200 μL of HAuCl4·3H2O and 4800 μL of distilled water to a Falcon tube, for a final concentration of 40 μg/mL Au.
  5. Add 250 μL of HAuCl4·3H2O and 4750 μL of distilled water to a Falcon tube, for a final concentration of 50 μg/mL Au.


Atomic Absorption Samples

Solutions with the following Au:lysozyme ratio were run on the AA:

  • 30:1 lysozyme-AuNP with 0.03 M MgCl2, CaCl2, NaCl, KCl, MES, citric acid (0.0002316 M 2,2 bipyridine)
  • 30:1 lysozyme-AuNP with 0.06 M MgCl2, CaCl2, NaCl, KCl, MES, citric acid (0.0004544 M 2,2 bipyridine)
  • 30:1 lysozyme-AuNP with 0.09 M MgCl2, CaCl2, NaCl, KCl, MES, citric acid (0.0006772 M 2,2 bipyridine)

Figures

  • This graph shows relatively constant amounts of gold present for all variables added. MES overall had the least amount of gold present in solution in comparison to other variables. Citric acid was the only variable that a visual reduction in the amount of gold in solution as the concentration of variable increased.



  • No 30:1 lysozyme-AuNP control was run because no samples were leftover from this batch of AuNP's. A single tube was made, but a mistake was made in its preparation causing fibers to form.
  • This UV-Vis corroborates all of the data presented in the AA by showing a change in absorbance as concentrations of gold change.



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