User:Sara K. Mattson/Notebook/Chemistry 671/2016/09/27: Difference between revisions

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==Purpose==
==Purpose==
Use fluorescence to examine the rate at which myoglobin unfolds at pH4
Use fluorescence to examine the rate at which myoglobin unfolds at pH4
==Procedure==
#Sample was made Myoglobin: 0.25mM Au, 12.5uM Myoglobin, 1.0mM HCl 3mL total
##Au 4.57mM
###(0.25mM)(3.0mL)=(XmL)(4.57mM)
###XmL= .176mL= 176uL Au
##Myoglobin 160uM
###(12.5uM)(3.0mL)=(XmL)(160uM)
###X=0.234mL= 234uL Myoglobin
##HCl 1.0mM
###(3.0mL)(1.0mM)=(1.0mM)(XmL)
###XmL=3.0mL= 300uL HCl
##Water
###Volume of water= 3000uL- 176uL-234uL-300uL= 2290uL
#Absorbance measurements were taken and saved every 3 minutes for 3 hours
#The instrument was set at 80 degrees Celsius
#The scan range parameters were start at 310nm and ending at 540nm
#The emission and exit slit were set at 10nm
==Solutions for UV-VIS preparation==
Stock Concentrations:
#Au 4.57mM
#Myoglobin 160mM
#Fructose 40.9mM
{|border="1" cellpadding="2"
|+ 0.25 mM Fructose
! scope="col" width="50" | pH
! scope="col" width="100" | μL of 1 mM HCl<sub>stock</sub>
! scope="col" width="100" | μL of 1 mM NaOH<sub>stock</sub>
! scope="col" width="100" | μL of 1 M NaOH<sub>stock</sub>
! scope="col" width="100" | μL of AuCl<sub>3stock</sub>
! scope="col" width="100" | μL of Fructose<sub>stock</sub>
! scope="col" width="100" | μL of Myoglobin<sub>stock</sub>
! scope="col" width="100" | μL of Water
|-
| 4 || 500 || 0 || 0 || 293 || 31 || 388 || 3788
|-
| 5 || 50 || 0 || 0 || 293 || 31 || 388 || 4238
|-
| 6 || 5 || 0 || 0 || 293 || 31 || 388 || 4283
|-
| 7 || 0 || 0 || 0 || 293 || 31 || 388 || 4288
|-
| 8 || 0 || 5 || 0 || 293 || 31 || 388 || 4283
|-
| 9 || 0 || 50 || 0 || 293 || 31 || 388 || 4238
|-
| 10 || 0 || 500 || 0 || 293 || 31 || 388 || 3788
|-
| 11 || 0 || 0 || 5 || 293 || 31 || 388 || 4283
|-
| 12 || 0 || 0 || 50 || 293 || 31 || 388 || 4238
|}
{|border="1" cellpadding="2"
|+ 0.75 mM Fructose
! scope="col" width="50" | pH
! scope="col" width="100" | μL of 1 mM HCl<sub>stock</sub>
! scope="col" width="100" | μL of 1 mM NaOH<sub>stock</sub>
! scope="col" width="100" | μL of 1 M NaOH<sub>stock</sub>
! scope="col" width="100" | μL of AuCl<sub>3stock</sub>
! scope="col" width="100" | μL of Fructose<sub>stock</sub>
! scope="col" width="100" | μL of Myoglobin<sub>stock</sub>
! scope="col" width="100" | μL of Water
|-
| 4 || 500 || 0 || 0 || 293 || 92 || 388 || 3727
|-
| 5 || 50 || 0 || 0 || 293 || 92 || 388 || 4177
|-
| 6 || 5 || 0 || 0 || 293 || 92 || 388 || 4222
|-
| 7 || 0 || 0 || 0 || 293 || 92 || 388 || 4227
|-
| 8 || 0 || 5 || 0 || 293 || 92 || 388 || 4222
|-
| 9 || 0 || 50 || 0 || 293 || 92 || 388 || 4177
|-
| 10 || 0 || 500 || 0 || 293 || 92 || 388 || 3727
|-
| 11 || 0 || 0 || 5 || 293 || 92 || 388 || 4222
|-
| 12 || 0 || 0 || 50 || 293 || 92 || 388 || 4177
|}
{|border="1" cellpadding="2"
|+ 1.25 mM Fructose
! scope="col" width="50" | pH
! scope="col" width="100" | μL of 1 mM HCl<sub>stock</sub>
! scope="col" width="100" | μL of 1 mM NaOH<sub>stock</sub>
! scope="col" width="100" | μL of 1 M NaOH<sub>stock</sub>
! scope="col" width="100" | μL of AuCl<sub>3stock</sub>
! scope="col" width="100" | μL of Fructose<sub>stock</sub>
! scope="col" width="100" | μL of Myoglobin<sub>stock</sub>
! scope="col" width="100" | μL of Water
|-
| 4 || 500 || 0 || 0 || 293 || 159 || 388 || 3666
|-
| 5 || 50 || 0 || 0 || 293 || 159 || 388 || 4116
|-
| 6 || 5 || 0 || 0 || 293 || 159 || 388 || 4161
|-
| 7 || 0 || 0 || 0 || 293 || 159 || 388 || 4166
|-
| 8 || 0 || 5 || 0 || 293 || 159 || 388 || 4161
|-
| 9 || 0 || 50 || 0 || 293 || 159 || 388 || 4116
|-
| 10 || 0 || 500 || 0 || 293 || 159 || 388 || 3666
|-
| 11 || 0 || 0 || 5 || 293 || 159 || 388 || 4161
|-
| 12 || 0 || 0 || 50 || 293 || 159 || 388 || 4116
|}
==Notes==
==Notes==
pH4 was already done on [[User:Sara K. Mattson/Notebook/Chemistry 671/2016/08/31|08/31/2016]] but the prepared solution had incorrect concentrations of myoglobin and gold NPs
pH4 was already done on [[User:Sara K. Mattson/Notebook/Chemistry 671/2016/08/31|08/31/2016]] but the prepared solution had incorrect concentrations of myoglobin and gold NPs

Revision as of 09:47, 28 September 2016

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Fluorescence

Purpose

Use fluorescence to examine the rate at which myoglobin unfolds at pH4

Procedure

  1. Sample was made Myoglobin: 0.25mM Au, 12.5uM Myoglobin, 1.0mM HCl 3mL total
    1. Au 4.57mM
      1. (0.25mM)(3.0mL)=(XmL)(4.57mM)
      2. XmL= .176mL= 176uL Au
    2. Myoglobin 160uM
      1. (12.5uM)(3.0mL)=(XmL)(160uM)
      2. X=0.234mL= 234uL Myoglobin
    3. HCl 1.0mM
      1. (3.0mL)(1.0mM)=(1.0mM)(XmL)
      2. XmL=3.0mL= 300uL HCl
    4. Water
      1. Volume of water= 3000uL- 176uL-234uL-300uL= 2290uL
  2. Absorbance measurements were taken and saved every 3 minutes for 3 hours
  3. The instrument was set at 80 degrees Celsius
  4. The scan range parameters were start at 310nm and ending at 540nm
  5. The emission and exit slit were set at 10nm


Solutions for UV-VIS preparation

Stock Concentrations:

  1. Au 4.57mM
  2. Myoglobin 160mM
  3. Fructose 40.9mM
0.25 mM Fructose
pH μL of 1 mM HClstock μL of 1 mM NaOHstock μL of 1 M NaOHstock μL of AuCl3stock μL of Fructosestock μL of Myoglobinstock μL of Water
4 500 0 0 293 31 388 3788
5 50 0 0 293 31 388 4238
6 5 0 0 293 31 388 4283
7 0 0 0 293 31 388 4288
8 0 5 0 293 31 388 4283
9 0 50 0 293 31 388 4238
10 0 500 0 293 31 388 3788
11 0 0 5 293 31 388 4283
12 0 0 50 293 31 388 4238
0.75 mM Fructose
pH μL of 1 mM HClstock μL of 1 mM NaOHstock μL of 1 M NaOHstock μL of AuCl3stock μL of Fructosestock μL of Myoglobinstock μL of Water
4 500 0 0 293 92 388 3727
5 50 0 0 293 92 388 4177
6 5 0 0 293 92 388 4222
7 0 0 0 293 92 388 4227
8 0 5 0 293 92 388 4222
9 0 50 0 293 92 388 4177
10 0 500 0 293 92 388 3727
11 0 0 5 293 92 388 4222
12 0 0 50 293 92 388 4177


1.25 mM Fructose
pH μL of 1 mM HClstock μL of 1 mM NaOHstock μL of 1 M NaOHstock μL of AuCl3stock μL of Fructosestock μL of Myoglobinstock μL of Water
4 500 0 0 293 159 388 3666
5 50 0 0 293 159 388 4116
6 5 0 0 293 159 388 4161
7 0 0 0 293 159 388 4166
8 0 5 0 293 159 388 4161
9 0 50 0 293 159 388 4116
10 0 500 0 293 159 388 3666
11 0 0 5 293 159 388 4161
12 0 0 50 293 159 388 4116

Notes

pH4 was already done on 08/31/2016 but the prepared solution had incorrect concentrations of myoglobin and gold NPs


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