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==Objectives==
 
==Objectives==
 
* Analyze AuADA solutions made on [[User:Melissa_Novy/Notebook/CHEM-571/2012/11/27|2012/11/27]] with UV-vis and AA spectroscopy.
 
* Analyze AuADA solutions made on [[User:Melissa_Novy/Notebook/CHEM-571/2012/11/27|2012/11/27]] with UV-vis and AA spectroscopy.
 +
** Please refer to [[User:Keyun_Wang/Notebook/Experimental_Biological_Chemistry_I/2012/11/28|Keyun Wang's entry]] for observations on and photographs of the solutions.
 
* Resuspend AuADA fibers in solutions made on [[User:Melissa_Novy/Notebook/CHEM-571/2012/11/27|2012/11/27]] with Tris buffer.
 
* Resuspend AuADA fibers in solutions made on [[User:Melissa_Novy/Notebook/CHEM-571/2012/11/27|2012/11/27]] with Tris buffer.
  
 
==UV-Vis Analysis of AuADA Solutions==
 
==UV-Vis Analysis of AuADA Solutions==
 
* The spectra of AuADA solutions were collected over 200 nm to 800 nm.
 
* The spectra of AuADA solutions were collected over 200 nm to 800 nm.
* 3 mL of AuADA solution was pipetted into a quartz cuvette and then placed in the UV-vis spectrophotometer.
+
* Exactly 3 mL of AuADA solution was pipetted into a quartz cuvette and then placed in the UV-vis spectrophotometer.
 
[[Image:AuADA_11272012.png]]
 
[[Image:AuADA_11272012.png]]
 
* A peak at around 520 nm was visible in the spectra of the following mole ratio Au:ADA solutions: 60 through 130.  This peak is likely due to the presence of GNPs in these solutions.
 
* A peak at around 520 nm was visible in the spectra of the following mole ratio Au:ADA solutions: 60 through 130.  This peak is likely due to the presence of GNPs in these solutions.
 +
* Purple fibers were observed in all of the solutions.  The liquid in each test tube was transparent purple.
 +
* Whereas [[User:Melissa_Novy/Notebook/CHEM-571/2012/11/14|undialyzed ADA]] did not form GNPs, these solutions using dialyzed ADA appeared to form GNPs.  It is possible that either the decrease in concentration of buffer salts between undialyzed and dialyzed ADA or the use of plastic test tubes to contain the Au and dialyzed ADA solutions contributed to the formation of GNP-ADA fibers.
 +
<br>
 +
[[Image:AuADA_Abs_at_520nm.jpg]]
 +
* The absorbance of 60 AuADA was the highest and the absorbance decreased as the mole ratio of the solutions increased.  Overlooking any experimental error, which is discussed below, this indicates that 60 AuADA created the most GNPs in solution and that this mole ratio was the best at forming the maximum number of GNPs out of the solutions tested.
 +
* One source of experimental error is resuspension of AuADA fibers that contain GNPs.  If they were resuspended, the GNPs in these fibers would be released into solution and contribute to the absorbance at 520 nm.
  
==Resuspension of ADA Fibers==
+
==Analysis of AuADA AAS Data==
 +
* Please refer to [[User:Keyun_Wang/Notebook/Experimental_Biological_Chemistry_I/2012/11/28|Keyun Wang's entry]] for the concentration of Au in AuADA solutions obtained from AAS, and the calibration curve used to calibrate the AAS with standard solutions of HAuCl<sub>4</sub> in 1 N HCl.
 +
* There were no deviations from the standard [[AU_Biomaterials_Design_Lab:Equipment/Shimadzu_AA-6200|protocol]] for analyzing solutions with AAS.
 +
<br>
 +
{| {{table}}
 +
| align="center" style="background:#f0f0f0;"|'''Au/ADA ratio'''
 +
| align="center" style="background:#f0f0f0;"|'''ADA added [uL]'''
 +
| align="center" style="background:#f0f0f0;"|'''HAuCl4 [uL]'''
 +
| align="center" style="background:#f0f0f0;"|'''Water [uL]'''
 +
| align="center" style="background:#f0f0f0;"|'''Mol Au [mol]'''
 +
| align="center" style="background:#f0f0f0;"|'''Mass Au [mg]'''
 +
| align="center" style="background:#f0f0f0;"|'''Initial [Au] [ppm]'''
 +
| align="center" style="background:#f0f0f0;"|'''Final [Au] [ppm]'''
 +
| align="center" style="background:#f0f0f0;"|'''Change in [Au] [ppm]'''
 +
| align="center" style="background:#f0f0f0;"|'''Percent Change [Au] [%]'''
 +
|-
 +
| 60||137.1||45.71||7817.2||4.79955E-07||0.094535296||11.81691206||1.3572||10.45971206||88.51476601
 +
|-
 +
| 70||137.1||53.3||7809.6||5.5965E-07||0.110232582||13.77907269||-0.2608||14.03987269||101.8927253
 +
|-
 +
| 80||137.1||60.9||7802||6.3945E-07||0.125950548||15.74381852||3.6762||12.06761852||76.64988328
 +
|-
 +
| 90||137.1||68.6||7794.4||7.203E-07||0.14187533||17.73441626||-1.0337||18.76811626||105.8287794
 +
|-
 +
| 100||137.1||76.2||7786.8||8.001E-07||0.157593297||19.69916209||-1.331||21.03016209||106.7566326
 +
|-
 +
| 110||137.1||83.8||7779.1||8.799E-07||0.173311263||21.66390791||-1.2499||22.91380791||105.7695038
 +
|-
 +
| 120||137.1||91.4||7771.5||9.597E-07||0.18902923||23.62865374||-1.3351||24.96375374||105.6503431
 +
|-
 +
| 130||137.1||99||7763.9||1.0395E-06||0.204747197||25.59339956||-1.4923||27.08569956||105.8308002
 +
|-
 +
| 140||137.1||106.7||7756.3||1.12035E-06||0.220671978||27.58399731||-1.2172||28.80119731||104.4127034
 +
|-
 +
| 150||137.1||114.3||7748.7||1.20015E-06||0.236389945||29.54874313||-1.4464||30.99514313||104.8949629
 +
|}
 +
<br>
 +
[[Image:AuADA_AAS_Percent.jpg]]
 +
<br>
 +
* The lowest percent change in Au ppm was observed in the 80 AuADA solution.  It can be assumed that this mole ratio is the best at forming the most GNPs relative to the amount of Au initially added.  As all solutions formed purple fibers and the supernatant was transparent purple in color, it can be assumed that GNPs were formed in all solutions.  However, in the solutions 90 through 150 AuADA, it can be inferred from the AAS and percent change [Au] data that the GNPs were entrapped in ADA fibers.
 +
 
 +
==Resuspension of AuADA Fibers==
 
* Tris buffer was made on [[User:Melissa_Novy/Notebook/CHEM-571/2012/09/04|2012/09/04]].
 
* Tris buffer was made on [[User:Melissa_Novy/Notebook/CHEM-571/2012/09/04|2012/09/04]].
* Please refer to [[User:Keyun_Wang/Notebook/Experimental_Biological_Chemistry_I/2012/11/28|Keyun Wang's entry]] for volumes and concentrations of Tris buffer added to  
+
* Please refer to [[User:Keyun_Wang/Notebook/Experimental_Biological_Chemistry_I/2012/11/28|Keyun Wang's entry]] for volumes and concentrations of Tris buffer added to the AuADA fibers.  Six solutions of mole ratio Au:ADA from 60 to 110 were chosen arbitrarily.
 +
** Note that the mole ratios of Au to ADA were irrelevant for these experiments, because the goal was to determine the effect of buffer pH and concentration on resuspending an arbitrary amount of AuADA fibers.
 +
** Within seconds after the addition of Tris buffer, all AuADA solutions became transparent purple in color and the fibers disappeared.
 +
*** Please refer to [[User:Keyun_Wang/Notebook/Experimental_Biological_Chemistry_I/2012/11/28|Keyun Wang's entry]] for a table of approximate time taken to resuspend the fibers after adding Tris buffer.
 +
* According to the data, it took a shorter amount of time to resuspend AuADA fibers in pH 10 Tris buffer.  This corresponds to protein theory, as any variance from a protein's optimal pH will denature or unfold it.  The optimal pH of ADA is between 6 and 7.  A pH of 10 has the greatest difference between the protein's optimal pH and therefore may have caused ADA to unfold more quickly.
  
 
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Latest revision as of 18:50, 23 September 2017

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Objectives

  • Analyze AuADA solutions made on 2012/11/27 with UV-vis and AA spectroscopy.
  • Resuspend AuADA fibers in solutions made on 2012/11/27 with Tris buffer.

UV-Vis Analysis of AuADA Solutions

  • The spectra of AuADA solutions were collected over 200 nm to 800 nm.
  • Exactly 3 mL of AuADA solution was pipetted into a quartz cuvette and then placed in the UV-vis spectrophotometer.

AuADA 11272012.png

  • A peak at around 520 nm was visible in the spectra of the following mole ratio Au:ADA solutions: 60 through 130. This peak is likely due to the presence of GNPs in these solutions.
  • Purple fibers were observed in all of the solutions. The liquid in each test tube was transparent purple.
  • Whereas undialyzed ADA did not form GNPs, these solutions using dialyzed ADA appeared to form GNPs. It is possible that either the decrease in concentration of buffer salts between undialyzed and dialyzed ADA or the use of plastic test tubes to contain the Au and dialyzed ADA solutions contributed to the formation of GNP-ADA fibers.


AuADA Abs at 520nm.jpg

  • The absorbance of 60 AuADA was the highest and the absorbance decreased as the mole ratio of the solutions increased. Overlooking any experimental error, which is discussed below, this indicates that 60 AuADA created the most GNPs in solution and that this mole ratio was the best at forming the maximum number of GNPs out of the solutions tested.
  • One source of experimental error is resuspension of AuADA fibers that contain GNPs. If they were resuspended, the GNPs in these fibers would be released into solution and contribute to the absorbance at 520 nm.

Analysis of AuADA AAS Data

  • Please refer to Keyun Wang's entry for the concentration of Au in AuADA solutions obtained from AAS, and the calibration curve used to calibrate the AAS with standard solutions of HAuCl4 in 1 N HCl.
  • There were no deviations from the standard protocol for analyzing solutions with AAS.


Au/ADA ratio ADA added [uL] HAuCl4 [uL] Water [uL] Mol Au [mol] Mass Au [mg] Initial [Au] [ppm] Final [Au] [ppm] Change in [Au] [ppm] Percent Change [Au] [%]
60 137.1 45.71 7817.2 4.79955E-07 0.094535296 11.81691206 1.3572 10.45971206 88.51476601
70 137.1 53.3 7809.6 5.5965E-07 0.110232582 13.77907269 -0.2608 14.03987269 101.8927253
80 137.1 60.9 7802 6.3945E-07 0.125950548 15.74381852 3.6762 12.06761852 76.64988328
90 137.1 68.6 7794.4 7.203E-07 0.14187533 17.73441626 -1.0337 18.76811626 105.8287794
100 137.1 76.2 7786.8 8.001E-07 0.157593297 19.69916209 -1.331 21.03016209 106.7566326
110 137.1 83.8 7779.1 8.799E-07 0.173311263 21.66390791 -1.2499 22.91380791 105.7695038
120 137.1 91.4 7771.5 9.597E-07 0.18902923 23.62865374 -1.3351 24.96375374 105.6503431
130 137.1 99 7763.9 1.0395E-06 0.204747197 25.59339956 -1.4923 27.08569956 105.8308002
140 137.1 106.7 7756.3 1.12035E-06 0.220671978 27.58399731 -1.2172 28.80119731 104.4127034
150 137.1 114.3 7748.7 1.20015E-06 0.236389945 29.54874313 -1.4464 30.99514313 104.8949629


AuADA AAS Percent.jpg

  • The lowest percent change in Au ppm was observed in the 80 AuADA solution. It can be assumed that this mole ratio is the best at forming the most GNPs relative to the amount of Au initially added. As all solutions formed purple fibers and the supernatant was transparent purple in color, it can be assumed that GNPs were formed in all solutions. However, in the solutions 90 through 150 AuADA, it can be inferred from the AAS and percent change [Au] data that the GNPs were entrapped in ADA fibers.

Resuspension of AuADA Fibers

  • Tris buffer was made on 2012/09/04.
  • Please refer to Keyun Wang's entry for volumes and concentrations of Tris buffer added to the AuADA fibers. Six solutions of mole ratio Au:ADA from 60 to 110 were chosen arbitrarily.
    • Note that the mole ratios of Au to ADA were irrelevant for these experiments, because the goal was to determine the effect of buffer pH and concentration on resuspending an arbitrary amount of AuADA fibers.
    • Within seconds after the addition of Tris buffer, all AuADA solutions became transparent purple in color and the fibers disappeared.
      • Please refer to Keyun Wang's entry for a table of approximate time taken to resuspend the fibers after adding Tris buffer.
  • According to the data, it took a shorter amount of time to resuspend AuADA fibers in pH 10 Tris buffer. This corresponds to protein theory, as any variance from a protein's optimal pH will denature or unfold it. The optimal pH of ADA is between 6 and 7. A pH of 10 has the greatest difference between the protein's optimal pH and therefore may have caused ADA to unfold more quickly.