Basics of Method Development Developing a Temperature Program


Using a Linear Temperature Program as a Starting Point If previous analysis information is not available to use as a guide, the first program development step is to try a simple, linear temperature program. This provides information on the retention characteristics of the solutes. Start with an initial temperature of 50°C (or 10°C below the boiling point of your sample solvent), a ramp rate of 10°/min, a final temperature equal to the isothermal temperature limit of the column, and a final hold time of approximately 30 minutes. The long final hold time is used to ensure all of the solutes elute from the column. The program can be stopped several minutes after the last solute has eluted from the column. This may occur before the final temperature is reached (Figure 11). After obtaining a chromatogram using the simple, linear temperature program, the next steps are to adjust the various program components to obtain adequate resolution and the shortest analysis time.


Figure 11: Simple, Linear Temperature Program


Column: DB-1, 15 m x 0.25 mm, 0.25 µm Carrier: Helium at 30 cm/sec Oven:


50-130°C at 10°/min


1. 3-Heptanone 2. 2-Heptanone 3. Cyclohexanone 4. 1,3-Dichlorobenzene 5. 1,4-Dichlorobenzene 6. 1,2-Dichlorobenzene 7. Iodobenzene 8. Naphthalene 9. 3-Nitrobenzene


2 1 3 4 5 8 9 6,7


01 23


4 Time (min.) Adjusting the Initial Temperature and Hold Time


To improve the resolution of earlier eluting peaks, decrease the initial temperature or increase the initial hold time. Decreasing the initial temperature usually results in the largest resolution improvement, but analysis times are substantially increased (Figure 12a). In addition, cool down times between runs can be significantly increased especially when cooling below 50°C. It is often impossible to cool a GC oven below 35°C in most laboratory environments without using cryogenic oven cooling. The resolution of the later eluting peaks are minimally affected by lowering the initial temperature especially for longer length columns. If excessive resolution is obtained with the original linear temperature program, increase the initial temperature to reduce the resolution and analysis time. The resolution of later eluting peaks may also be reduced upon increasing the initial temperature.


5


6


7


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