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## Kinetics

General Protocol:

1. Prepare a 40μM solution of adenosine in 50mM phosphate pH 7.4 buffer.
• Note: This solution was prepared via dilution of a 0.0123M ADA stock solution
2. Add 3mL of 40μM adenosine to a cuvette with a stir bar
• Note: All analysis should be carrier out in a temperature controlled environment. The temperature was kept at 25°C
3. Allow the solution to reach a constant 25°C
4. Begin taking absorbance spectra (take a spectrum every 15seconds)
6. Allow the reaction to run for 10 minutes, taking a spectrum every 15seconds.

Sample Preparation:
Table 1. Preparation of 0.0123M stock solution

 Mass of adenosine (g) 0.0165 Volume of buffer(mL) 5

Table 2. Preparation of 40μM adenosine solution

 Volume of stock(mL) 0.0098 Volume of buffer(mL) 2.9902

UV-vis:
Figure 1. Corrected Absorbance spectra of the conversion of adenosine to inosine by ADA

• Using the molar absorptivity of inosine and adenosine determined on 2013/09/03, the conversion of adenosine to inosine can be represented as concentrations.The molar absorptivity of adenosine and inosine was also determined for all wavelengths.

Table 3. Molar Absorptivity of Adenosine at 260nm and Inosine at 250nm

 Wavelength Adenosine Inosine 250 13333 11937.5 260 16866.7 7312.5

Table 4. Concentration of Adenosine during the conversion to inosine

 Time (s) Concentration(M) 15 0.000039 30 45 60 75 90 105 120 135 150 165 180 195 210 225 240 255 270 285 300 315 330 345 360 375 390 405 420 435 450 465 480 495 510 525 540 555 570 585 600

Table 5. Monitoring the concentration of inosine during the conversion of adenosine to inosine by ADA

 Time (s) Concentration(M) 15 0.0000 30 0.0000 45 0.0000 60 0.0000 75 90 105 120 135 150 165 180 195 210 225 240 255 270 285 300 315 330 345 360 375 390 405 420 435 450 465 480 495 510 525 540 555 570 585 600