Calibration Examples
Calibrating without Standard Sample

If you calibrate with an internal standard substance, i.e., an Internal Standard, you may as well perform the calibration without standard samples. The prerequisite is that you know the relative Response Factor of the respective substance for the ISTD. The following entries are required:

Sample List

Assign all samples the Unknown sample type. Enter the amount for the ISTD in the ISTD Amount column:

QNT Method/General Tab

On the General tab page, select Fixed as Calibration Mode.

QNT Method/Peak Table Tab

The internal standard substance is defined in the peak table. If all three samples contain two relevant substances only as in the example, make the following entries in the Standard column, using the F8 input box.

• For substance 1, select Use this peak as Internal Standard and enable the Use sample amount as reference (Variable Internal Standard).

• In the F8 input box of substance 2, select Internal. In the Associated ISTD Peak field, select the standard substance serving as internal standard (here: Subst. 1).

• If necessary, repeat this operation for every additional substance that should be calibrated based on the internal method.

After completing the input, the substance 2 is labeled Var. Internal Subst. 1 in the Standard column. In addition, substance 1 is labeled ISTD Var. Internal to indicate that substance 1 is used as internal standard for the internal calibration:

QNT Method/Amount Table Tab

Define the corresponding calibration curve in the Amount table of the QNT Editor:

• Enter =1.000 as C1 value for both the ISTD and the unknown substance.

 Tip:

Do not recalibrate afterwards by clicking Calibrate on the General tab page because this action overwrites the values for the Calibration Coefficient.

• Enter the known response factor. This factor automatically refers to the ISTD because the ISTD is used for calibration. It is not necessary to define the ISTD as reference.

Amount Calculation

Chromeleon automatically calculates the amount values for the substance 2. The calculation is performed according to the formula for amount calculation (rel. to ISTD) (for more information, refer to Formula for Amount Calculation (Rel. to ISTD) in the Glossary section). In case of internal calibration, the external calibration of the internal standard substance is adapted to the corresponding sample, using the area ratio of the peak to the ISTD in the corresponding sample. In addition, in this special case, the response factor is used to take the absorption of substance 2 in relation to the ISTD (here: Subst. 1) into account.

Chromeleon calculates the amount for substance 2 adapted by the deviation of substance 1.

For detailed information about how to calculate the amount for unknown substance using internal calibration, refer to  Internal Calibration: Calculation.

For an overview of the different calibration possibilities with internal standard substances provided by Chromeleon, refer to Calibrating  Calibrating with an Internal Standard Substance.