-
- Posts: 64
- Joined: Sat Dec 14, 2013 4:14 pm
Hello:
I am running an 1100 with a RP Column and Isocratic flow.
Water: ACN :: 92:8 + TEA and pH Adjustment
I recently made a new very precise solvent mix. After degassing, I started up let it run for about 20 mins and then injected the sample. The retention time of the main peak and impurities was as expected. A single impurity peak was more or less flat but acceptable.
Now in the second injection, the main peak took almost 2wice as long to get eluted. The main peak, however was still very crisp (not spread out). In the second injection, I also got very crisp impurity peak.
Now I observe that there is a small reduction in the pressure, this could be due to improper degassing (?), however how could this lead to crisp impurity peaks and delayed main peak. This has happened to me earlier also. Back then we were testing the method out so, we didnt give much attention.
Since this is a USP method, the retention time of the main peak is approximately known.
I have another question: I have a binary pump so, some times I run the water in one pump and ACN in the other. This seems to have provided very stable pumping. However since this is an isocratic method. I mix the 2 some times and pump in one pump.
I always thought, it is better to mix earlier and pump. However in this case, it seems to be leading to retention time variations. Inspite of vigerous shaking and then degassing, is it possible that a pre-mix solvent is not homogeneous ? Nothing else could logically explain the sudden shift in retention time ?
Thanks for your thoughts.
I am running an 1100 with a RP Column and Isocratic flow.
Water: ACN :: 92:8 + TEA and pH Adjustment
I recently made a new very precise solvent mix. After degassing, I started up let it run for about 20 mins and then injected the sample. The retention time of the main peak and impurities was as expected. A single impurity peak was more or less flat but acceptable.
Now in the second injection, the main peak took almost 2wice as long to get eluted. The main peak, however was still very crisp (not spread out). In the second injection, I also got very crisp impurity peak.
Now I observe that there is a small reduction in the pressure, this could be due to improper degassing (?), however how could this lead to crisp impurity peaks and delayed main peak. This has happened to me earlier also. Back then we were testing the method out so, we didnt give much attention.
Since this is a USP method, the retention time of the main peak is approximately known.
I have another question: I have a binary pump so, some times I run the water in one pump and ACN in the other. This seems to have provided very stable pumping. However since this is an isocratic method. I mix the 2 some times and pump in one pump.
I always thought, it is better to mix earlier and pump. However in this case, it seems to be leading to retention time variations. Inspite of vigerous shaking and then degassing, is it possible that a pre-mix solvent is not homogeneous ? Nothing else could logically explain the sudden shift in retention time ?
Thanks for your thoughts.
