User:Patrick Hampson/Notebook/chem471/2016/09/06

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Ocean Optics 201696

Description/ Objective

Sample Preparation

BSA Ocean Optics Final Sample Solution Concentrations:

  1. 3 mL total
  2. 0.25 mM Au
  3. 3.125 μM BSA
  4. HCl or NaOH appropriate for your pH

Stock solution Concentrations:

  1. 5.08 mM AuCl3
  2. 34 mM BSA
  3. 1 mM HCl
  4. DI H20

Solution Transfer Volumes:

  1. 2278 uL H20
  2. 147 uL AuCl3
  3. 275 uL BSA
  4. 300 uL HCl

Protocol

  1. Extra Steps
    1. Do you need temperature control or need stirring?
      1. See the protocol for controlling the cuvette controller Quantum Northwest Cuvette Holder
  2. Turn on the jaz spectrophotometer (red on-off button)
  3. Turn on the Lamps (Ocean Optics DH-2000)
    1. Turn on the switch in the back of the controller box
    2. Press the blue "Deuterium" button and wait for the light to stay on
    3. Press the red "Halogen" button
    4. The lamps need 15 minutes to warm up before you are ready to proceed to the next step.
  4. Starting the software
    1. Open the OceanView software.
      1. Click on the "Create New Spectroscopy Application" icon (looks like a bunch of shark fins stacked on one another)
        1. Click on "Spectroscopy"
        2. Select JAZA1666 and click "Next"
        3. Select "Absorbance" and click "Next"
        4. Set Acquisition Parameters
          1. Set "Integration Time" to 1 ms
          2. Set "Scans to Average" to 100
          3. Click "Next"
        5. Store Reference Spectrum
          1. Make sure that the toggle on the light source is switched to "Open"
          2. Click on the light bulb icon
            1. A spectrum should appear in the middle display
          3. Click "Next"
        6. Store Background Spectrum
          1. Make sure that the toggle on the light source is switched to "Closed"
          2. Click on the light bulb icon
            1. A spectrum should appear in the middle display
          3. Click "Finish"
        7. Make sure that the toggle on the light source is switched to "On"
  5. Set up your data acquisition schedule
    1. Click on the "Save graph to file/files" icon (the floppy disc icon on the lower icon panel)
    2. Click "Yes" for "You must configure save parameters first. Do you want to configure now?"
    3. Set Save Options
      1. Choose the "Between saved scans, wait at least" option
      2. Set the time to whatever the experiment requires
        1. For most nanoparticle syntheses, this will be 2 minutes
      3. Choose the "Stop after this amount of time" option
        1. For most nanoparticle syntheses, this will be 3 hours
    4. Set File Options
      1. Save to Directory: C:\Users\Lab Admin\DropBox\ 2016\Ocean Optics\Year\Month\Date
      2. Select Open
      3. Set Basename to something that will be descriptive of your data for example Lysozyme_AuNP_Ratio45_80C
      4. Click "Apply"
      5. Click "Exit"
  6. Start data collection
    1. After you have your sample, cuvette, and cuvette holder properly prepared perform the following:
      1. Click the "Save graph to file/files" icon (the floppy disc icon on the lower icon panel)
      2. Data files should be showing up in your selected folder. If it isn't, you have done something wrong.
  7. Shutting down
    1. When your data collection is finished, perform the following steps
      1. Shut down the OceanView software
      2. Turn off the jaz spectrophotometer
      3. Turn off the lamps (main switch in the back)
      4. Shut down the Quantum Northwest Cuvette Holder if necessary
  8. Analyzing Data
    1. Open data compilation folder.
    2. Open copy command.txt document.
    3. In data compilation folder shift right click and choose open command window here.
    4. Copy the first line from the copy command.txt document and paste into the command window. Hit enter.
    5. It will prompt “location?” Copy the location of the saved scans, paste it into the command window, and hit enter.
    6. Then copy the second line from the copy command.txt document, paste into the command window, and hit enter.
    7. It will prompt “number of files.” Enter the number of files that it has copied which will be immediately above the prompt.
    8. It will prompt “number of lines per file.” Enter 2048.
    9. It will prompt “number of lines before data.” Enter 17.
    10. It will prompt “Are you sure (Y/N)?” Enter Y.
    11. In the data compilation there will a document titled out.txt. Open out.txt and save as an excel file in the folder with the saved scans and rename to match the folder.
    12. Delete out.txt if still saved in data compilation folder.

Conducting Protocol

  1. After the Ocean Optics instrument was set up and the samples were prepared, they were placed in the machine.
  2. The machine was run, and recorded data every two minutes.
  3. The recorded data was uploaded immediately to dropbox.

Data

The data that was gathered from the Ocean optics machine, which was uploaded to dropbox, was assembled in an excel file. The maximum absorbance and maximum wavelength over the time duration of the experiment were calculated and analyzed. Max abs over time.PNG

This graph illustrates an increase in absorbance of the solution over time. The increase in absorbance is accounted for by the formation of gold nano-particles during the course of the experiment.

Max wavelength over time.PNG

This graph illustrates that the maximum wavelength achieved by the nano-particles shifted. The maximum wave length was stable at a consisitent 453.4nm until approximately 3000 seconds into the experiment, when it jumped to 536.5nm.

The absorbance at wavelength 535.15 was also analyzed over time. A353 over time.PNG

This graph illustrates the obvious increase in absorbance over time.

The group's raw data can be found here: File:BSA AuNP pH4.xlsx

Analysis

Change in the absorbance over time is to be accounted for by the presence of gold nano-particles. A decrease in absorbance would suggest that the nano-particles are disappearing, whereas an increase in the absorbance of the solution signifies the formation of gold nano-particles.

A shift in wavelength could signify changes in the structure, formation, or concentration of the gold-nanoparticles.