So the question is: what is the optical density of the original sample?

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SUMMARY

The discussion centers on determining the optical density (OD) of a diluted algae sample using the Beer-Lambert Law. The original sample had an OD greater than 4.5, necessitating dilution for measurement. After diluting the sample by a factor of 7 and measuring the OD at 1.58, the original OD was calculated incorrectly by one participant as 11.06, while the correct calculation using logarithmic principles yielded an OD of 2.43. The conversation emphasizes the importance of understanding the linear relationship between absorbance and concentration in the context of optical density measurements.

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  • Understanding of the Beer-Lambert Law
  • Familiarity with optical density and absorbance concepts
  • Knowledge of dilution factors and their application in measurements
  • Experience with spectrophotometry techniques
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  • Study the Beer-Lambert Law in detail, focusing on its mathematical implications
  • Learn about the calibration of spectrophotometers for accurate OD measurements
  • Explore advanced dilution techniques for high-density samples
  • Investigate the impact of sample matrix on optical density readings
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Researchers in bioenergy, laboratory technicians working with algae samples, and anyone involved in optical density measurements in spectrophotometry.

Arjan.
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The problem: how to determine the optical density of a diluted sample
Hi, i work on algae and bioenergy and i have a question about the OD.

I have to determine the optical density of about 400 samples. Some samples however are too "dense" for the photospectrometer, that is they have an optical density greater than the range of the spectrometer (>4,5 OD)

In order to determine the optical density of these samples I diluted them.

So for example:
The original sample has an OD > 4,5, which is too high for the machine
So it was diluted.
0,5 ml of the sample were taken and 3,0 ml distilled H2O were added.
The OD of the diluted sample was measured
The OD of this sample is 1,58.

Question
What is the optical density of the original sample?

Homework Equations



http://upload.wikimedia.org/wikipedia/commons/0/04/Beer_lambert.png

The Lamber-Beer equation says:

Absorbance = extinctioncoefficient x length of path x concentration

Optical density = log ( I0/I1)
Absorbance = log ( I1/I0)

Links: http://en.wikipedia.org/wiki/Optical_density
http://en.wikipedia.org/wiki/Beer-Lambert_law

The Attempt at a Solution



In the example above i diluted 7 times, so the optical density is 7 x 1,58 = 11,06
 
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Your answer is correct. The OD of the original sample is the OD of the diluted sample * the dilution factor.
 
This doesn't make sense to me. OD is a base-10 logarithm scale. So for example, if you dilute by 10x then the diluted OD will be 1 less than (not 1/10th) of the undiluted OD.

The original OD should be
1.58 + log(7)
= 1.58 + 0.85
=2.43

However, you should have been able to measure OD 2.43 in the original sample.
 
According to the Beer Lambert Law, the absorbance of a sample (aka optical density) varies linearly with the concentration for sufficiently dilute samples.
 
Deleted post ... just realized an error in how I was thinking about all this.
 
Last edited:
Ygggdrasil said:
varies linearly with the concentration for sufficiently dilute samples.

And this word "sufficiently" is the only reason why I have not posted earlier that Arjan procedure was correct...
 

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