Lab #3 info

# Lab #3 info - Laboratory III Students must bring in their...

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Laboratory III Students must bring in their Laptop to plot the graph for λ max of p-nitrophenol in the lab Title: : Pipetting Techniques and Photometry Introduction In previous experiments, we learned the relationship between 1) pH and ionization 2) color , absorbance. and wavelength. The degree of ionization of weak acids is dependent on pH. As pH increases, ionization of a weak acid increases. Some weak acids such as pH indicators have colored ions. The maximum absorbance of a colored solution lies within the wavelength of its complimentary color. The linear relationship between absorbance and the concentration is given by Beer’s law : A = ε bc or A = abc where A is the absorbance, ε is the molar extinction coefficient, a is the molar absorptivity , b is the length (in cms) through which the light passes in the solution, and c is the molar concentration of the solution. The concentration of an sample can be obtained using a standard curve The standard curve is a graph plotted by measuring absorbance (A) using known concentrations of standard solutions. The linear relationship between absorbance (A) and concentration is limited. Very low and very high concentrations will not obey Beer’s law. Only unknown concentration that lie within the linear range of the graph can be estimated accurately. The molar extinction coefficient is a fundamental unit in spectrophotometry. The molar extinction coefficient is a constant value for a molecule when absorbance is measured at the predetermined solvent, pH, temperature and λ max. Molar extinction coefficient, ( ε29 and molar absorptivity (a) differ in units. The concentration for molar absorptivity is units other Molar (example expressed in gms/L) p-Nitrophenol (p-NP) is a weak acid. It is colorless in a unionized form and yellow in its ionized form. As the pH increases, there will a proportionate increase absorbance at λ max due to increased ionization. The wavelength of maximum absorbance ( λ max) of a yellow solution is in the violet range. The pKa of p-Nitrophenol is 7.15. It is completely un-ionized ( HA ) at pH 4, partially ionized ( HA + A - ) at pH 7 and completely ionized at pH 10 ( A - ). In today’s experiment, you will calculate the pKa of p-nitrophenol. You will determine λ max at intervals of 10 nm and use the λ max to determine the absorbance (A) of p-nitrophenol at different pH. You will calculate molar absorptivity (a) using the following formula.

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Lab #3 info - Laboratory III Students must bring in their...

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