There are many possible explanations for retention time increases. We will need more information before we can determine the cause.
How much does the retention time increase each time? Is it a constant percentage, or is it increasing or decreasing?
What concentration are you injecting? Is the sample filtered?
Do you see an increase in pressure after each injection?
Please provide more information on the column. What is the packing in the column? Silica? Bonded phase?
Some possible explanations?
1. These normal phase systems for separating lecithins are known to be very slow to equilibrate. Some methods suggest a three day equilibration time! Maybe you have to pump mobile phase through the column for a longer time. If the retention time changes are smaller after more injections, then maybe your column is not equilibrated.
2. Lecithin is often a mixture of many components. Sometimes some of these compounds stick to the stationary phase and do not elute. With every injection you are creating a new stationary phase, which has different eluting properties.
3. If the pressure is increasing with each injection, you may have a solubility problem; something in the sample is not soluble in the mobile phase. If your flow decreases when the pressure increases, then you have a pump problem. Check your pump seals and check valves.
If these suggestions do not help, write back and we will try again.
