I need the formula for estimation of thiamine content in plant extract after measuring the absorbance? If thiamine standard curve is needed then how can that be constructed?
Dear Ali,
The best way to go about this is to make a thiamine standard curve from the absorbance values obtained from a serially diluted concentration of thiamine. You may wish to express it in mg/ml or ug/ml. Whichever you adopt, please note that you first make a stock solution of higher concentration and use C1V1=C2V2 to get diluted concentration.
As a guide, dissolving 1mg (0.001g) of thiamine in 1 ml of solvent gives 1mg/ml which equals 1000µg/ml.
Since, measuring 0.001g thiamine will be difficult, I will recommend you measure 0.010g (10mg) and then dissolve in 10 ml of solvent which gives you same concentration as 1mg/ml or 1000µg/ml stock.
From here, you can make a dillution of
500µg/ml (the same as 0.50mg/ml),
250µg/ml (0.25mg/ml),
125µg/ml (0.125mg/ml)
62.5µg/ml (0.0625mg/ml)
31.25µg/ml (0.03125mg/ml)
After you make the dilutions, you apply same treatment you gave the unknown sample to it and read the absorbance. It is always good you make readings of the sample and diluted standards same time using same blank.
After reading, please make the standard curve and obtain the equation of the curve. Y=Mx+C, where Y = Absorbance, x = concentration. You can make x the subject and substitute the absorbance of the unknown sample. Once you determine the concentration, remember it will be in µg/ml or mg/ml depending on the one you used.
To determine the exert concentration say in 100g of sample, please apply the following formula:
thiamine (mg/100g) = (Concentration determined from curve x total volume of extract in ml x 100)/(Volume of aliquot used in ml x weight of sample)
I hope this helps.
With regards,
Egbuna, C.
https://egbunac.com
Hi, you can use the Beer-Lambert Law :
https://chem.libretexts.org/Core/Physical_and_Theoretical_Chemistry/Spectroscopy/Electronic_Spectroscopy/Electronic_Spectroscopy_Basics/The_Beer-Lambert_Law
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