Question for NH2P-50 4E Column

Record: 915

Category: Column Problem
Product: NH2P-50
Sample:

Question
The column only washed at the end of the whole sequence.

The standard solution is reproducible after five injections. I see the same results. Only the first injection of six showed lower peak area sometimes.

Answer
What is the sample matrix? Are you injecting samples in 65% ACN or are they in water?

When you write: “Only the first injection of six showed lower peak area sometimes.” Do you mean that the first injection after the water injections is lower?

Perhaps the blank water injections are causing the column to need extra time to equilibrate. Our tech support says the solubility of water and ACN is not good, and that it takes a longer time to make the base line stable when the ratios have been changed immediately before. In this case can you run a “blank” sample with eluent (65% ACN) instead of water.

 
 
Additional Comments
I have described your experimental procedure to Japan as follows:
8 minutes 5 mL/min 65% ACN
1-3 Water
4 Mannitol Control in water
5-10 sample in water
Wash with 100% ACN.

Using this procedure our tech support in Japan has made the following comments:
She understood the first step as equilibration for the system only because the flow rate is not the usable condition for NH2P-50 4E column. The maximum flow rate for the column is 1.5 mL/min. She thinks the column has not equilibrated from 100% ACN to 65 % ACN in the short amount of time. She also recommends storing the column in 65 % ACN to eliminate the need for long equilibration times.

However since there was no problem for the analysis of mannitol control, she think the matrices in the sample solution might be the cause. She recommends adding some ACN into the sample solution to obtain the better shape for all of the HILIC analyses. She also recommends to dissolve the mannitol standard in the sample matrix and confirm the reproducibility.

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