# Can someone help? - (Aug/04/2006 )

Hi there

I am unsure about this question

If the extinction coefficient (ε) of substance X at wavelength λ=560 nm is 2500 M-1 cm-1, what is the molar concentration of a sample with an absorption of 2 ?

-0.9992 M

-1250 M

-0.0008 M

-0.0002 M

so using c=A/el

c=2/(2500x0.000056)

=14.29 but its obviously wrong! where have I ogne wrong I changed nm into cm so??????

Thanks

biology_06er

Absorbance = (molar extinction coefficient * molar concentration * sample thickness)

Thus:

2 = 2500 * x * 1

2 = 2500x

x = 2/2500

x = 0.0008

Homebrew is dead right.

0.8 mM (assuming 1 cm light path)

However, look back at your question. If they have not given you the length of the light path then technically you cannot calculate the concentration.

Most people always use 1 cm light paths but I have, on occassion, used a 0.2 cm light path.

so your mistake was to integrate the wavelength in the formula.

In the formula, the cm refers to the distance covered by the light, i.e. the size of the cuve.

They do not tell you the size, that's why you should not answer. however you can assume that the distance is 1 cm, which is quite usual.

About the wavelength, you need to know the wavelength, because the coefficient absorption is always referred to a specific wave length, and you will calibrate the reader at the rigth wavelength.

. and obtain an absorption at this specific wavelength. But you don't use it in the formula.

I did assume that the wavelength and light path distance used to obtain the absorbance of 2 was the same as that used to express the extinction coefficient.

Hey there guys

Thanks for the help...much appreciated....so when Homebrew said it was sample thickness and Missele said it was the distance covered by the light does it just mean that the light travelled through a thickness of X?

Thanks

biology_06er

Thanks for the help...much appreciated....so when Homebrew said it was sample thickness and Missele said it was the distance covered by the light does it just mean that the light travelled through a thickness of X?

Thanks

biology_06er

Yes, just worded differently. Most of the time you'll see Beer's Law written with the words, length of light path (in cm). This basically means the distance the light travels through your sample, i.e., the thickness of the sample or the width of your cuvette (minus the walls).