User:Roberta Diaz Jimenez/Notebook/CHEM 471 Experimental Lab/2015/09/15

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Objective

For this lab session, our group (GRP 3) will be doing oceanography and prepping samples with our assigned protease: trypsin. After completing oceanography readings, we will also redo stock solutions to have gold fibers instead of nanoparticles, and prepare more trypsin samples for later use.

Procedure

  • Oceanography
  1. Centrifuged eppindorf tubes containing gold fibers.
  2. Removed water with plastic pipette.
  3. Prepare the sample that goes in the 3mL cuvette that will be used for ocean optics.
  4. The sample must contain
    1. 1mL of gold fibers
    2. 0.3mL of buffer
    3. 0.0573mL of trypsin, calculations are below
    4. The rest of the volume was filled with water: 1 - (1mL fibers + 0.3mL buffer + 0.0573mL trypsin)= 1.643mL water.
    5. The trypsin was added last, at the second reading such that second reading becomes time 0.
  5. We ran our sample for 2hrs30mins.
  6. Readings were taken every 2 minutes and uploaded directly to DropBox for later analysis.

Calculations for volume of trypsin:

    C1·V1 = C2·V2,
     where C1 = 52.36µM
           C2 = 1µM 
           V2 = 3mL
    V1 = 57.3µL 
  • Preparation of stock solution containing fibers for analysis
  1. We followed Dr. Hartings' modified protocol for making the fiber samples
  • Preparation of trypsin samples.
  1. Weigh 15 eppendorf tubes and record their mass.
  2. To each tube, add approximately 1mg of trypsin, weigh and record.
  3. Calculate the concentration in each tube if 1mL of liquid were added.

Calculations to

Data

Table 1. Measurements taken while making the trypsin sample tubes

Number of Eppi Tube Mass of Eppi Tube (g) Mass of Trypsin (g) Concentration (µM)
1 1.02691 0.00108 46.35
2 1.02622 0.00148 63.52
3 1.0224 0.00117 50.21
4 1.01944 0.00169 72.33
5 1.02922 0.00181 77.68
6 1.02625 0.00128 54.93
7 1.01909 0.00165 70.82
8 1.02036 0.00182 78.11
9 1.01922 0.00128 54.93
10 1.02024 0.00122 52.36
11 1.02097 0.00154 66.09
12 1.02422 0.00175 75.11
13 1.02669 0.00124 53.22
14 1.02936 0.00148 63.52
15 1.02923 0.00155 66.52

Table 2. Raw data for the graph following it. The graph is a Concentration vs. Absorbance of the class's compiled UV/Vis data at 280 nm.

Group Concentration A(280nm)
Group 1 2 0.052
5 0.16
8 0.239
10 0.292
15 0.444
Group 3 0.7875 0.026
2.625 0.091
5.25 0.186
7.5 0.252
15 0.502
Group 4 0.965 0.032
1.93 0.069
3.86 0.123
7.225 0.261
15.445 0.517
Group 5 1.56 0.052
3.13 0.108
6.25 0.221
12.5 0.441
15 0.54

Graph 1. 571ClassesUVVISGRAPHMABC.png

  • Ocean Optics:

Graph 2.

Ocean Optics TrypsinandAUfibers 15Sept.png

Graph 3. Ocean Optics TrypsinandAUfibers at533nm 15Sept.png