HPLC sample result quantification

Basic questions from students; resources for projects and reports.

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Hello,

I'm not quite sure on how to determine the real sample concentration of a component. here is how I did the experiment, I use soxhlet extractor to extract antioxidant from flax seed. Before HPLC sample analysis, I first prepare a calibration curve (concentration Vs area) for the component. And then I took 100ul of oil from soxhlet extraction and dissolved in 900ul of alcohol. Then based on my calibration curve I got concentration of 3mg/ml. So, my question is, is this result (3 mg/ml) same as the real sample concentration? or do I have to consider a dilution factor for this? how do I calculate concentration of the real sample? and also i appreciate if you can suggest any material on this regard.

Thanks in advance,

Pardize
3mg/ml is clearly not the result you're looking for, since you want to quantify an amount of antioxidant in a solid sample.

You've diluted the extract 10 times, this means that the extract had a concentration of 30 mg/ml.

To go back to the sample, you need to know the amount of sample you extracted to the amount of solvent.
Thanks a lot Rndirk for your help,

I used 200ml of organic solvent and after rotary evaporation i get 5ml extract.
from this I use 100ul for HPLC analysis.

So, does this mean that the extract sample concentration would be 6mg/ml?

Thanks again,
No. The concentration of the 5ml after rotary evaporation is 30mg/ml.

What was the original concentration of the 200ml then? Hint: it's less.

To go back to your sample you need the weight of flax seeds you extracted to 200ml!
Hmm,
I use 10g of flaxseed.

I really appreciate your assist.
Thanks,
15mg of antioxidant in 1 g of flaxseed
Your analysis showed a concentration of 3mg/ml.
This is the concentration of the sample you injected, so the 3mg came from the 100µl of oil you used to prepare the sample. This means the concentration of this oil is 30mg/ml
The total volume of oil after soxlet is 5ml, so it contains 150mg of anti-oxidant.
This 150mg came from the 10g of flax-seeds you used.
150mg/10g => 15mg/gram of seeds.
Thanks a lot, guys!
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