Quantification Methods

This page covers calibration and quantitation setup in GCsolution for the Shimadzu GC-2025. You should already have a working method file — see Method_Development if not.

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External Standard Calibration

How it works: Prepare standard solutions at known concentrations. Inject a constant volume of each. Plot a calibration curve (concentration vs. peak area/height). Match unknown sample peak area to the curve.

Limitations:

• Highly sensitive to injection volume variations
• Requires strictly identical conditions between standards and unknowns
• Non-repeatable if injection volume varies

📋 From course materials

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Internal Standard Calibration

How it works: Add a constant, known amount of a reference compound (Internal Standard) to every sample. Plot the calibration curve using ratios: (target peak area ÷ IS peak area) vs. (target concentration ÷ IS concentration). This automatically corrects for injection volume errors and instrument drift.

Advantages:

• Highly accurate quantitation
• Compensates for injection volume and instrument variations

Selecting an Internal Standard:

• Must be stable and chemically different from the target
• Close in chemical structure and properties to the target
• Similar concentration to the target
• Elutes near the target (similar retention time)
• Completely separated from all other sample components

Time-saving tip: If all samples (standards + unknowns) contain the same IS concentration, enter “1.0” for all IS amounts. The ratio-based calculation works correctly regardless of actual concentration.

📋 From course materials

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Setting Up Calibration in GCsolution

Step 1: Configure the Method File

1. Open method file → Quantitative tab
2. Select quantitative method: External Standard or Internal Standard
3. Set # of Calib. Levels (number of standard concentrations)

For External Standard: click Compound tab → enter target concentrations in [Conc.1], [Conc.2], etc.

For Internal Standard:

1. Click Compound tab → add target compounds
2. Add IS compound → set [Type] to ISTD
3. Ensure target and IS share the same ISTD Group number
4. Enter known IS amount in [Conc.1]

📋 From manual

Step 2: Build the Batch Table

Open Batch Processing on the Assistant Bar. Register samples as follows:

Sample	Sample Type	Level #	Purpose
First standard	1:Standard:(I)	1	Initialize calibration (clears old data)
Subsequent standards	1:Standard	2, 3, 4…	Add calibration levels
Unknowns	0:Unknown	—	Quantitate against curve

Level # must correspond to the concentration level defined in the Compound table.

📋 From manual

Step 3: ISTD Amount for Unknowns

• Standard samples: software auto-pulls IS concentration from the Method file — no entry needed
• Unknown samples: you must enter the IS mass in the ISTD Amount column of the Batch Table

Quick fill method: Right-click the ISTD Amount column header → Input Col. Data… → specify row range → enter value → ensure Auto-increment is unchecked → OK.

Simplified approach: If all samples have the same IS concentration, enter “1.0” everywhere (both in Method’s Compound table and in the Batch Table). The ratio math works correctly.

📋 From manual

Step 4: Run

1. Save batch: File → Save Batch File As
2. Click Start (green play icon)
3. System automatically injects standards, builds calibration curve, then quantifies unknowns

For full batch workflow details, see Batch_Processing.

📋 From manual

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See Also

• Method_Development — Creating method files
• Single_Run_Analysis — Running one sample
• Batch_Processing — Running automated sequences
• Sample_Preparation — Preparing standards and unknowns