User:Moira M. Esson/Notebook/CHEM-571/2013/10/23

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(HRP-AuNP and Myoglobin-AuNP)
 
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#Synthesize new HRP-AuNP solutions and Myoglobin-AuNP solutions for future catalytic activity testing.
#Synthesize new HRP-AuNP solutions and Myoglobin-AuNP solutions for future catalytic activity testing.
<br>
<br>
-
==
+
==HRP-AuNP and Myoglobin-AuNP==
 +
*The protein-AuNP solutions prepared on [[User:Moira M. Esson/Notebook/CHEM-571/2013/10/22|10/22/2013]] did not produce fibers. As such, new protein-AuNP solutions with increased ratios of protein-AuNP were prepared in order to determine the optimal concentration of Au to protein to synthesize fibers.
 +
*The general protocol for the synthesis of HRP-AuNP and Pepsin-AuNP follows the general protocol for the preparation of protein wrapped Au-NPs described on [[User:Moira M. Esson/Notebook/CHEM-571/2013/08/28|08/28/2013]].
 +
*5mL of each solution was prepared.
 +
<br>
 +
Table 1. Stock solutions used in the preparation of Protein-AuNP solutions
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<br>
 +
{| {{table}}
 +
| align="center" style="background:#f0f0f0;"|'''Concentration HRP Day 2(M)'''
 +
| align="center" style="background:#f0f0f0;"|'''Concentration Gold Stock (M)'''
 +
| align="center" style="background:#f0f0f0;"|'''Myoglobin (M)'''
 +
|-
 +
| 0.0000015||0.0025||0.0000165
 +
|}
 +
<br>
 +
Table 2. Preparation of HRP-AuNP solutions
 +
<br>
 +
{| {{table}}
 +
| align="center" style="background:#f0f0f0;"|'''Ratio'''
 +
| align="center" style="background:#f0f0f0;"|'''Amount of gold added (mL)'''
 +
| align="center" style="background:#f0f0f0;"|'''Concentration HRP (M)'''
 +
| align="center" style="background:#f0f0f0;"|'''Volume HRP (mL)'''
 +
| align="center" style="background:#f0f0f0;"|'''Volume H<sub>2</sub>O added (mL)'''
 +
|-
 +
| 230||0.5||1.08696E-06||3.623188406||0.876811594
 +
|-
 +
| 250||0.5||0.000001||3.333333333||1.166666667
 +
|-
 +
| 270||0.5||9.25926E-07||3.086419753||1.413580247
 +
|-
 +
| 290||0.5||8.62069E-07||2.873563218||1.626436782
 +
|-
 +
| 310||0.5||8.06452E-07||2.688172043||1.811827957
 +
|-
 +
| 330||0.5||7.57576E-07||2.525252525||1.974747475
 +
|-
 +
| 350||0.5||7.14286E-07||2.380952381||2.119047619
 +
|-
 +
| 370||0.5||6.75676E-07||2.252252252||2.247747748
 +
|-
 +
| 390||0.5||6.41026E-07||2.136752137||2.363247863
 +
|-
 +
| 410||0.5||6.09756E-07||2.032520325||2.467479675
 +
|}
 +
<br>
 +
Table 3. Preparation of Myoglobin-AuNPs
 +
<br>
 +
{| {{table}}
 +
| align="center" style="background:#f0f0f0;"|'''Ratio'''
 +
| align="center" style="background:#f0f0f0;"|'''Amount of gold added (mL)'''
 +
| align="center" style="background:#f0f0f0;"|'''Concentration Myoglobin (M)'''
 +
| align="center" style="background:#f0f0f0;"|'''Volume of myogloblin(mL)'''
 +
| align="center" style="background:#f0f0f0;"|'''Volume Water Myoglobin(mL)'''
 +
|-
 +
| 30||0.5||8.33333E-06||2.525252525||1.974747475
 +
|-
 +
| 50||0.5||0.000005||1.515151515||2.984848485
 +
|-
 +
| 70||0.5||3.57143E-06||1.082251082||3.417748918
 +
|-
 +
| 90||0.5||2.77778E-06||0.841750842||3.658249158
 +
|-
 +
| 110||0.5||2.27273E-06||0.688705234||3.811294766
 +
|-
 +
| 130||0.5||1.92308E-06||0.582750583||3.917249417
 +
|-
 +
| 150||0.5||1.66667E-06||0.505050505||3.994949495
 +
|-
 +
| 170||0.5||1.47059E-06||0.445632799||4.054367201
 +
|-
 +
| 190||0.5||1.31579E-06||0.398724083||4.101275917
 +
|-
 +
| 210||1.19048E-06||0.360750361||4.139249639
 +
|}
 +
<br>
 +
==UV/vis of Pepsin-AuNP and HRP-AuNP==
 +
'''General Protocol''':
 +
# Place 1mL of each ratio in an 1.5mL eppendorf tube.
 +
# Centrifuge at maximum speed for 30 minutes.
 +
# Take a UV-Vis spectrum of each sample from 200nm to 800nm.
 +
# After taking the spectrum, store the sample in a fresh eppy tube in an upright position.
 +
<br>
 +
Figure 1. Corrected Absorbance spectra of pepsin-AuNPs
 +
<br>
 +
[[Image:Pepsin Nano.png]]
 +
<br>
 +
*Both 30:1 and 70:1 pepsin-AuNP had an absorbance peak at 518nm.
 +
 +
==Notes==
 +
*The remaining UV/vis samples will be run during the next lab period.
 +
*Although neither the HRP or the pepsin solutions formed fibers, a black residue and color change(see figure 2) was observed for the pepsin solutions. The black residue will be tested in the future for identification.
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Objectives

  1. Run UV-vis on the 20 protein-Au nanoparticle samples prepared on 10/22/2013.
  2. Synthesize new HRP-AuNP solutions and Myoglobin-AuNP solutions for future catalytic activity testing.


HRP-AuNP and Myoglobin-AuNP

  • The protein-AuNP solutions prepared on 10/22/2013 did not produce fibers. As such, new protein-AuNP solutions with increased ratios of protein-AuNP were prepared in order to determine the optimal concentration of Au to protein to synthesize fibers.
  • The general protocol for the synthesis of HRP-AuNP and Pepsin-AuNP follows the general protocol for the preparation of protein wrapped Au-NPs described on 08/28/2013.
  • 5mL of each solution was prepared.


Table 1. Stock solutions used in the preparation of Protein-AuNP solutions

Concentration HRP Day 2(M) Concentration Gold Stock (M) Myoglobin (M)
0.00000150.00250.0000165


Table 2. Preparation of HRP-AuNP solutions

Ratio Amount of gold added (mL) Concentration HRP (M) Volume HRP (mL) Volume H2O added (mL)
2300.51.08696E-063.6231884060.876811594
2500.50.0000013.3333333331.166666667
2700.59.25926E-073.0864197531.413580247
2900.58.62069E-072.8735632181.626436782
3100.58.06452E-072.6881720431.811827957
3300.57.57576E-072.5252525251.974747475
3500.57.14286E-072.3809523812.119047619
3700.56.75676E-072.2522522522.247747748
3900.56.41026E-072.1367521372.363247863
4100.56.09756E-072.0325203252.467479675


Table 3. Preparation of Myoglobin-AuNPs

Ratio Amount of gold added (mL) Concentration Myoglobin (M) Volume of myogloblin(mL) Volume Water Myoglobin(mL)
300.58.33333E-062.5252525251.974747475
500.50.0000051.5151515152.984848485
700.53.57143E-061.0822510823.417748918
900.52.77778E-060.8417508423.658249158
1100.52.27273E-060.6887052343.811294766
1300.51.92308E-060.5827505833.917249417
1500.51.66667E-060.5050505053.994949495
1700.51.47059E-060.4456327994.054367201
1900.51.31579E-060.3987240834.101275917
2101.19048E-060.3607503614.139249639


UV/vis of Pepsin-AuNP and HRP-AuNP

General Protocol:

  1. Place 1mL of each ratio in an 1.5mL eppendorf tube.
  2. Centrifuge at maximum speed for 30 minutes.
  3. Take a UV-Vis spectrum of each sample from 200nm to 800nm.
  4. After taking the spectrum, store the sample in a fresh eppy tube in an upright position.


Figure 1. Corrected Absorbance spectra of pepsin-AuNPs
Image:Pepsin Nano.png

  • Both 30:1 and 70:1 pepsin-AuNP had an absorbance peak at 518nm.

Notes

  • The remaining UV/vis samples will be run during the next lab period.
  • Although neither the HRP or the pepsin solutions formed fibers, a black residue and color change(see figure 2) was observed for the pepsin solutions. The black residue will be tested in the future for identification.


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