Standard equation of FeSO4 is y=4.5682x-0.0347.
Different concentration used for the preparation of std curve are 60, 120, 180, 240, 300 mg/L.
I took 100ul of sample having concentration 1mg/ml and to it added 100 ul of water.
So, I used total of 200 ul of sample after assay reading were noted at 593 nm.
For my extract absorbance value comes out to be 1.567.
Now how to calculate mg Fe (II) equivalent/g of dry weight with this reading.
- please help
You must to dilute your sample in order to get an absorbance value that fits in the midle of the absorbances of the standard curve (e.g. if the 180 mg/L FeSO4 is 0,300 nm your samples must have absorbances around 0,3 nm). If the aborbances of the standard curve do not reach 1.567, you can not calculate the value of your sample.
After the diluition and assay, you have to substitute the "y" value in the given standard equation and then calculate it using the concentration of the sample in the assay. For example: If your initial concentration is 1mg/mL or 1g/L (you have to transform it to g/L, since your curve is in mg/L). Thus, you have 1g/L and you will dilute it 2 times, now you have 0,5 g/L.
Supposing that the value of "x" of your sample after the calculation with the equation was 190mg/L you must to divide 190 mg/L by 0,5g/L of sample and the result will be 380 mg FeSO4 by g of sample.
I hope that it helps.
First thing is that you need to dilute (4X) your sample again because your absorbance sholuld fall between 0.1 to 0.9 absorbance units (yours is 1.567). By putting the value of absorbance in your standard curve equation you can obtain the value for x i.e. concentration in mg/L (as your standard curve is prepared in mg/L) multiply it by 1/2 (dilution factor in this case) will give you the desired value for Fe (II) in mg/L.
You must to dilute your sample in order to get an absorbance value that fits in the midle of the absorbances of the standard curve (e.g. if the 180 mg/L FeSO4 is 0,300 nm your samples must have absorbances around 0,3 nm). If the aborbances of the standard curve do not reach 1.567, you can not calculate the value of your sample.
After the diluition and assay, you have to substitute the "y" value in the given standard equation and then calculate it using the concentration of the sample in the assay. For example: If your initial concentration is 1mg/mL or 1g/L (you have to transform it to g/L, since your curve is in mg/L). Thus, you have 1g/L and you will dilute it 2 times, now you have 0,5 g/L.
Supposing that the value of "x" of your sample after the calculation with the equation was 190mg/L you must to divide 190 mg/L by 0,5g/L of sample and the result will be 380 mg FeSO4 by g of sample.
I hope that it helps.
I have the similar questions to Dr. Batista. I took approximately 0.5-0.8 g field moist sample and measured Fe2+ from soil after extraction with 6M Hcl and ferrozine method and measured the absorbance. The absorbance varied from 1.5-3.01 at 562 nm. How can I calculate Fe2+ in my sample and what should be the unit (e.g. mg/g dry soil or mM). Please help me.
Dear coleague,
You should analyze a linear standard curve using Fe (II) according to the same method. The absorbance of the samples must fit around the average absorbance of the standard curve. The unit used for the standard curve should be the one for the sample. e. g. If you have a mg/L curve and your extracts is in g/L, you can divide the result of the linear regression in mg/L by the concentration of the extract in g/L and the result will be then given in mg/g. I don't have experience with the ferrozine method, but I hope it helps.
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