Tuesday, July 5, 2011

Calculations in Analytical Methods(HPLC and GC)

I want to write this topic for giving an idea about how to calculate the Assay, related substances and Residual solvents.

Calculations for Assay Method (Tirtimetry)

Titire value(Sample - Blank) x M x Factor x 100 x 100
                            Ws x (100-LOD)

M= Molarity of the volumetric solution 
Ws= Weight of the sample 
Factor = Factor for Drug substance 
LOD is Loss on drying or MC is water content 

Calculations for Assay Substances Method (HPLC)
                    At x Ws x P x (100-WC of std)
Assay % = ----------------------------------
                    As x Wt x (100-WC of sample)

At = Area due to sample Preparation 
As= Area due to working standard solution 
Wt= Weight of the sample in mg
Ws= Weight of the working standard
P=Potency  or Assay of the standard 
WC=% water content of the sample or standard 

Note: WC may be replaced by LOD based on the drug substance 

Calculations for Related Substances Method (HPLC)
% of Known Impurity = ---x 100 X RF

% of unknown Impurity = ---x 100

Total Impurities = Sum of all known and unknown impurities 

ri =Area of each impurity Peak in the chromatogram of the sample solution preparation
rs=Sum of areas of Main drug and all impurity Peaks in the chromatogram of the sample solution preparation
RF= Response factor 

Calculations for Residual solvents Method (GC)
    At        Volume of solvent taken x density     0.1 ml       dilution of sample(ml)
= ------x -------------------------------------------x ---------x-------------------------------x 10 ^ 6
    As                     10 ml                                                                   10 ml        Weight of the sample

At= Area of the solvent in the sample - Blank
As= Area of the solvents in the standard -blank 

Note: Volume of the solvents taken should be mentioned in microliters 

As we are multiplying with 10 power 6 we can get the results in PPM 

Note: Refer Merck index for solvent densities


  1. This comment has been removed by the author.

  2. In the calculation the total value must be multiplied by 100 for percentage

    1. For Calculation for Assay substances method (By HPLC)

    2. Hello,

      Can you please clarify about the potency..

      is the Potency is on 'as-is' basis or 'Anhydrous' basis in the Calculation for Assay...

    3. Hey Chandra,

      i think your query makes no sense, as if there is any LOD in standard then there is every chance for Over Assay, so everytime the standard to be dried[if required] till the LOD becomes zero/ until it comes into specified limit.

      Pharma Engineering

  3. Hello how can i calculate the potency of a peak from a chromatogram. i have an agilent 1100. i have the standards and my sample. Any suggestions?

    1. spl area/std area*std weight/spl weight*%potency*(100-LOD or water)%*100, in case of tablet put average weight and divide by its label claim * 100

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  5. can anybody suggest me why loss on drying or water content of drug substance is not considered while calculating the assay percentage of API in drug product, only standard potency and water or LOD is considered. why is so? please anyone

  6. can u give me the formula for calculation of assay of tablet, for example with potency of 98.55 % and water content 10 %, water of sample = 10.25 %, average weight of tablet = 210 mg

  7. This is too good to learn it here, good explanation, thanks for sharing.

    Pharma Engineering

  8. Nice Information and Very Useful for me and also Useful for Who wants more Information About HPLC GC.

  9. can you please tell me tge formula for calculating assay of suspension?

  10. Wt is label claim?