Hello everyone, today let’s talk about the influence of temperature on gas phase analysis in detail. How to choose and optimize the method in the constant temperature analysis and temperature programming of the gas phase?

Whether it is temperature-programmed or constant-temperature analysis, the ultimate goal is to achieve baseline separation of multiple compounds in the sample, and to present a chromatographic peak with a sharp peak shape and a Gaussian distribution. Resolution is a crucial parameter for measuring column efficiency in chromatographic analysis, so what are the factors that affect resolution? First, let’s look at the separation equation.

1.Separation Equation

In the above formula, Rs represents resolution, N represents column efficiency, α represents selectivity, and K represents retention factor.

Retention factor K: For compounds, the effect of temperature on retention time is mainly as follows: the higher the column temperature, the weaker the retention and the shorter the retention time; the lower the column temperature, the stronger the retention and the longer the retention time. In general, retention time decreases by 50% for every 30°C increase in column temperature.

Selectivity α: Lower column temperature helps to improve resolution.

Column efficiency N: The column temperature also has a certain influence on the column efficiency of the chromatographic column. Generally, as the column temperature increases, the retention time is shortened, the peak width becomes smaller, and the column efficiency increases.

2.Constant temperature analysis and temperature programmed analysis in gas analysis methods

In constant temperature analysis, the column temperature of the gas chromatography column is constant, which is mainly suitable for the analysis of compounds with little difference in boiling point and similar retention properties.

However, when the retention properties of the components to be tested differ greatly, the chromatographic peaks of the compounds with late eluting peaks have various problems such as wider peak width, poor peak shape, and low sensitivity; the chromatographic peaks of compounds eluting earlier will appear again. The phenomenon of poor separation. In this case, the temperature-programmed method is required.

In the temperature-programmed method, the initial temperature of the column is lower, which is beneficial to improve the resolution of the earlier eluting chromatographic peaks. Then, as the column temperature rises and the method is kept at the highest temperature for a period of time, the retention time of the later peak compounds can be moved forward, which improves the work efficiency.

3.Optimization of the heating program

In general, the following heating gradients can be used as a starting point for method optimization.

■ The initial temperature should not be too high. It is recommended to set it to 15°C higher than the laboratory room temperature.
■ It is recommended to set the heating rate to 10℃/min.
■ Set the maximum temperature to about 20°C below the upper limit of the constant temperature of the chromatographic column, and then keep it for 10 minutes to observe the separation state of the components to be tested in the chromatogram.
■ According to the obtained chromatogram, further optimize the heating conditions.

(1). Choice of heating method

In the chromatogram, if the “window” of the peak time of the analyte (all compounds) is less than 1/4 of the whole temperature program time, then isothermal analysis is recommended. At this time, it is recommended to set the constant temperature to 45°C lower than the peak temperature of the last peak, and then adjust it up and down by about 10°C until the desired degree of separation is achieved.

In addition, if the “window” of the peak time of all compounds is greater than 1/4 of the entire temperature-programmed time, or if the isothermal analysis method cannot achieve a satisfactory resolution, in this case, a temperature-programmed analysis method is required.

(2). During temperature programming, the setting of the initial temperature

■ For splitless injection, generally set the initial temperature to be 10-20°C lower than the boiling point of the solvent, and maintain it for 1 min. The purpose is to reduce the solvent effect, because during splitless injection, the sample expansion volume is large, the carrier gas flow rate is relatively small, and the vaporized sample cannot enter the chromatographic column instantaneously. Setting the initial temperature to 10-20°C lower than the boiling point of the solvent and maintaining it for 1 min is beneficial to the condensation and focusing of the solvent, resulting in narrower peak broadening and improved resolution.

■ In case of split injection, the flow rate of carrier gas in the liner is relatively fast, and the components to be tested can enter the chromatographic column quickly. So there is no need to perform condensation focusing on the sample, and there are no special requirements for the setting of the initial temperature.

In addition, the resolution of earlier eluting chromatographic peaks can sometimes be improved by slightly lowering the initial temperature and increasing the hold time.

(3). Setting of heating rate

In general, the faster the heating rate, the shorter the retention time of the overall chromatographic peaks, and the faster the peaks appear, which can improve work efficiency, but at the same time reduce the resolution of chromatographic peaks. Therefore, we hope to obtain the best separation effect with the shortest analysis time. The recommended heating rate can be calculated by the above formula: T₀ is the dead time, which refers to the time when the peak value of the components not retained by the stationary phase occurs from the beginning of the injection.

(4). Setting of the constant temperature platform

When we use the same heating rate, the chromatographic peaks of some compounds still cannot reach a satisfactory resolution. We can set a constant temperature platform at about 45°C below the peak temperature of these compounds, maintain this temperature for 2-5min, and then continue warming up. If there are many types of components to be tested, in order to achieve better separation, it may be necessary to set up multiple constant temperature platforms.

(5). End temperature setting

As for the end temperature, it is generally set to be about 20°C higher than the peak temperature of the last chromatographic peak. If the sample matrix is complex or splitless injection, the end temperature needs to be set slightly higher, the purpose is to bake the gas column at high temperature to remove the possible high-boiling impurities. But at the same time, it should be noted that the set temperature cannot exceed the upper limit of the operating temperature of the chromatographic column.

Well, today’s introduction to how to choose between gas chromatography constant temperature analysis and temperature programmed analysis, and how to optimize the method. In addition, Welch Materials has been focusing on the development and production of gas chromatography columns for many years, with the advantages of high inertness, low bleed, high column efficiency and long life.

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