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(Atomic Absorption of Au/BSA Solutions)
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Revision as of 21:22, 6 December 2012

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Objectives

  • Analyze Au/BSA solutions made on 2012/10/03 with atomic absorption.
  • Calculate activity of HRP from the UV-vis data obtained from the HRP-AAP assay on 2012/09/18 and 2012/09/19.

Atomic Absorption of Au/BSA Solutions

  • The AA was calibrated with standard solutions at concentrations of 5, 8, 10, 15, 20, 25, 30, and 40 ppm made by Dr. Miller on 2012/10/08 and blanked with 1 N HCl.
  • Deionized water was run through the AA between samples.
  • Au/BSA solutions containing fibers at Au:BSA mole ratios of 120 through 170 were filtered with dry 0.45 μM pore-size nylon filter paper into a clean, dry flask. The filtrate was then decanted into a Falcon tube for testing.
    • Note that the fibers on the filter paper were not washed. The fibers on the filter paper were dark purple in color.
    • Colored solutions at Au:BSA mole ratios of 120, 128, and 130 that contained fibers produced clear filtrate after being filtered and the AA detected that they contained little to no gold (please refer to Table 1). This indicates that either the gold was of large enough size to be filtered by the filter paper, or that the pores in the filter paper were blocked with fibers and thus did not filter the gold. It would be incorrect to assume that the solutions did not contain any free gold, as literature states that the purple color of the solutions before filtering is due to the optical properties of gold nanoparticles.


  • Table 1: Change in Au Concentration (ppm) of Solutions of Au/BSA
Au:BSA Mole Ratio Theoretical Conc. Au [ppm] Actual Conc. Au [ppm] Δ Au Conc. [ppm] Percent Change Conc. Au [%]
60 40.9690424 50.1489 -9.1798576 -22.40681515
80 40.9690424 50.5319 -9.5628576 -23.34166737
100 40.9690424 52.6358 -11.6667576 -28.47700829
120 40.9690424 -0.77 41.7390424 101.8794679
128 40.9690424 -0.3222 41.2912424 100.7864475
130 40.9690424 -0.3654 41.3344424 100.8918929
132 40.9690424 35.2115 5.7575424 14.05339755
133 40.9690424 35.0928 5.8762424 14.34312851
134 40.9690424 33.8251 7.1439424 17.43741611
136 40.9690424 -0.8617 41.8307424 102.1032954
138 40.9690424 -0.7862 41.7552424 101.9190099
140 40.9690424 -0.8185 41.7875424 101.99785
160 40.9690424 -0.6891 41.6581424 101.6820017
170 40.9690424 -0.2845 41.2535424 100.6944268


Measured Au ppm.jpg

Change in Au ppm.jpg

  • Note that as negative concentrations of Au are experimentally impossible, the values are likely due to improper zeroing of the AAS. For the purposes of data analysis, it will be assumed that the concentration of Au in those solutions is zero.
  • According to Table 1, Au/BSA solutions at mole ratios of 60, 80, 100, 132, 133, and 134 contained Au. These solutions likely successfully formed AuNPs, corroborated by visual observation of their purple color. The remainder of the solutions likely did not form AuNPs, as no Au was detected by the AAS. Thus, it can be assumed that BSA encapsulated the Au added to the solutions and thereby prevented Au from being present in solution. However, it is also possible that filtration removed AuNPs from solution, as in the case of solutions of Au:BSA mole ratios of 136 through 170.


  • Table 2: Change in Moles of Au of Solutions of Au/BSA
Au:BSA Mole Ratio Theoretical Mol Au [mol] Actual Conc. Au [M] Actual Mol Au [mol] Δ Mol Au [mol] Percent Change Mol Au [%]
60 2.08E-06 0.000254606 2.54606E-06 -4.66E-07 -22.40681515
80 2.08E-06 0.000256551 2.56551E-06 -4.86E-07 -23.34166737
100 2.08E-06 0.000267232 2.67232E-06 -5.92E-07 -28.47700829
120 2.08E-06 -3.90929E-06 -3.90929E-08 2.12E-06 101.8794679
128 2.08E-06 -1.63581E-06 -1.63581E-08 2.10E-06 100.7864475
130 2.08E-06 -1.85514E-06 -1.85514E-08 2.10E-06 100.8918929
132 2.08E-06 0.000178769 1.78769E-06 2.92E-07 14.05339755
133 2.08E-06 0.000178166 1.78166E-06 2.98E-07 14.34312851
134 2.08E-06 0.00017173 1.7173E-06 3.63E-07 17.43741611
136 2.08E-06 -4.37485E-06 -4.37485E-08 2.12E-06 102.1032954
138 2.08E-06 -3.99154E-06 -3.99154E-08 2.12E-06 101.9190099
140 2.08E-06 -4.15553E-06 -4.15553E-08 2.12E-06 101.99785
160 2.08E-06 -3.49856E-06 -3.49856E-08 2.11E-06 101.6820017
170 2.08E-06 -1.44441E-06 -1.44441E-08 2.09E-06 100.6944268


Change in Mol Au.jpg

Activity of HRP Based on the HRP-AAP Assay

  • Note that data obtained on 2012/09/18 was used to calculate the activity of HRP.
  • The ratio of the change in time to the change in absorbance was calculated to be the rate of the reaction. The inverse of the rate was plotted against the inverse of the concentration of whichever reactant was varied, either AAP or H2O2.


Varying AAP.jpg

  • Note that the concentration of H2O2 was fixed at 1.7 mM.


Varying H2O2.jpg

  • Note that the concentration of AAP was fixed at 2.5 mM.