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Chemistry Forums for Students => Analytical Chemistry Forum => Topic started by: STM on June 21, 2015, 11:55:55 AM

Title: UV Visible Spectroscopy calculation: Beer-Lambart's law
Post by: STM on June 21, 2015, 11:55:55 AM
Dear all,
Please I need clarification on this calculation procedure.

I did a UV-Visible measurement of Conjugated diene in lipid by measuring absorbance at 234 nm. I got an Absorbance = 3.31. Using molar absorbtivity, e, = 24000 M-1 cm-1, and path length = 1 cm.

From the formula A= eCL, C=A/eL = 3.310/ 24000x1 =0.0001379 M (137.9uM).

The weight of lipid (Oil) sample taken was 0.018g and was dissolved in 3.5 ml methanol as medium for the UV-Vis measurement.

I am expected to express the answer as umole/g (micromole/g). Is it just the 137.9 uM/0.018g or will there be need for other parameters to come into the calculation.

Please I need you advice on how to get to umole/ g. I have done some calculations already but I just need some clarifications from you all.

Thank you.
Title: Re: UV Visible Spectroscopy calculation: Beer-Lambart's law
Post by: Borek on June 21, 2015, 01:50:04 PM
137.9 μM is the concentration of the diene in the solution, not its amount in the original sample.
Title: Re: UV Visible Spectroscopy calculation: Beer-Lambart's law
Post by: Babcock_Hall on June 22, 2015, 09:53:52 AM
Maybe I am missing something here and I am not an analytical chemist, but an absorbance of 3.31 is out of the linear range of most spectrophotometers, due to stray light.  If stray light is a problem, there is an apparent negative deviation from the Beer-Lambert law.
Title: Re: UV Visible Spectroscopy calculation: Beer-Lambart's law
Post by: marquis on June 22, 2015, 10:29:40 PM
Babcock Hall has a good point.

Typically, with Beer-Larmbert, linearity has to be proven.  It
can't be assumed.

If you want a quick test, do a 1:10 dilution and rerun the test.
See if the answer comes in at one tenth the value.  If it does, all is
good with the world.
Title: Re: UV Visible Spectroscopy calculation: Beer-Lambart's law
Post by: STM on June 23, 2015, 12:28:02 PM
Thank you all for your response. I am grateful. The points on the linearity range for Beer-Lamberts law is noted. Although, I do not intend to use the measurement for any form of calibration. Nevertheless, the result of the 10-fold dillution was still consistent.

Also, I will like to check this conversion>

If an oil sample has peroxide value of 5 milliequivalent of Oxygen per Kg, density = 0.889 g/ml. Its conversion to mmol/L is?

If 1 mmol of O2 = 2 meq, therefore, 5 meq = 2.5 mmol

5 meq/kg  x 1/2 = 2.5 mmol/kg.       2.5 mmol/Kg x 1 kg/1000 g x 0.889 g/ml x 1000 ml/L = 2.22 mmol/ L (2.22 mM)

Can you please confirm this conversion?

Thank you