For example i have a solution of a compound having lambda max at 307nm. I want to know the concentration of this solution. Can i calculate by using UV Spectrophotometer? Please help.
Hi Salman,
It is very easy. At first, you need to prepare a number of solutions with known concentration of that compound i.e. 2 mg/mL, 5mg/mL, 8mg/mL,10 mg/mL, 15 mg/mL, 20mg/mL, 25 mg/mL etc. and measure the uv-vis absorbence. Plot them by using concentration in X axis and absorbance values in Y axis. Then, do the linear fit which will give you an equation like y= a + bx ( If you get the r square value ~0.99 then that calibration curve is quite reliable to figure out the unknown concentration). Then, measure the uv-absorbance of your sample solution. Calculate the concentration by placing the absorbance value in that equation.
Regards,
Tasnuva
Hi Salman,
It is very easy. At first, you need to prepare a number of solutions with known concentration of that compound i.e. 2 mg/mL, 5mg/mL, 8mg/mL,10 mg/mL, 15 mg/mL, 20mg/mL, 25 mg/mL etc. and measure the uv-vis absorbence. Plot them by using concentration in X axis and absorbance values in Y axis. Then, do the linear fit which will give you an equation like y= a + bx ( If you get the r square value ~0.99 then that calibration curve is quite reliable to figure out the unknown concentration). Then, measure the uv-absorbance of your sample solution. Calculate the concentration by placing the absorbance value in that equation.
Regards,
Tasnuva
Hello! You have to prepare 6-7 solutions of known concentration in small volumetric flasks and measure the adsorption of each solution. Do not prepare high concentrated solutions. You have to measure these known concentration solutions because you need to make a CALIBRATION CURVE. This curve must be linear with R square=0,99. The y-axis is the absorbance that you measured from the instrument and the x-axis is the known concentration of the solution (be careful! not the concentration of the solution in the flask, but the concentration of the solution that you added in the UV-cell. The calibration curve is based on the Beer-Lambert law:
A=e b C
So by making the calibration curve you find the coefficient e.
Then if you measure the absorbance of the unknown solution you will again use the above equation.
A(absorbance) is known becaused you have measured it, e coefficient is known from the calibration curve and b is usually 1cm (b is the cell length). So you can calculate C (concentration).
Regards, MP
take known concentration of analyte make a calibration curve than take the unknown you will get absorbance and that A means what conc. you will get from your calibration curve. for better understanding on how to make calibration curve read spectrophotometric determination of iron in dissolved water.
I think this link is very useful,
http://www.instanano.com/characterization/theoretical/concentration-calculation-from-uv-vis-absorbance/
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