How can I calculate concentation (mg/kg) of sample by using area of peak during analyzing the pesticide residues by HPLC?
Known concentration/unknown concentration=known sample peak area/unknown sample peak area.
Muhammad Nauman Hanif
you could use an external calibration curve then applying (or not) the recovery factor to the results. Anyway, you should declare your choice.
Otherwise, you could use and internal standard and again a calibration curve.
There are other techniques, but the first is the easiest.
If you know area of peck of known concentration than you can calculate unknown. The better way for this to have linear correlation between concentration and area which can named as calibration.
Using the same HPLC method, a multi-level calibration table must be created using proper standards for comparison.
Assuming that you are using a UV detector then you have a relatively linear response versus concentration curve according to Beer's law (you should look that up; it is useful). So, you make a calibration by plotting the area of the peaks of known standards against the concentration of the known standards. This is your "calibration curve". Most HPLC software will have this built into it so you don't have to plot the curve by hand, but you should do it by hand at least once so that you understand how it works. You make your standards from pure pesticides - you can buy those from many different sources.
The process is the same for other detectors only depending upon the detector you might not get a linear response. You have to understand how the detector works to know what to expect in your calibration.
To calculate the concentration (mg/kg) of a pesticide residue in a sample using the area of the peak obtained from an HPLC analysis, you'll need the following information:
1. Calibration Curve: First, you need to establish a calibration curve for the pesticide of interest. This involves analyzing standard solutions of known concentrations of the pesticide using the same HPLC method. From this, you can plot a calibration curve relating peak area to concentration.
2.Peak Area: During the analysis of your sample, you'll obtain a peak corresponding to the pesticide residue. The area of this peak can be obtained from the chromatogram.
3.Sample Preparation Details: You need to know the details of your sample preparation, such as the volume of the sample extracted and any dilutions performed.
Once you have this information, you can use the following formula to calculate the concentration of the pesticide residue in your sample:
Concentration (mg/kg)=Peak area of standard×Sample volume (L)Peak area of sample×Concentration of standard (mg/L)×Dilution factor
Where:
- Peak area of sample: Area of the peak corresponding to the pesticide residue in your sample.
- Concentration of standard (mg/L): Concentration of the pesticide in the standard solution used to generate the calibration curve.
- Peak area of standard: Area of the peak corresponding to the pesticide residue in the standard solution.
- Sample volume (L): Volume of the sample extracted for analysis.
- Dilution factor: If any dilutions were performed on the sample before analysis, you need to account for this by including the dilution factor in the calculation.
By plugging in the appropriate values, you can calculate the concentration of the pesticide residue in your sample in mg/kg. Ensure that all units are consistent throughout the calculation.
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