We are presently using a strong cation exchange gradient method with the following mobile phase composition and gradient conditions:

Mobile Phase 1: 0.1 M phosphate, pH 3.3: ACN (1128:872, w:w)
Mobile Phase 2: 0.1 M phosphate, pH 3.3, 0.2 M sodium perchlorate: ACN (1128:872, w:w)

time MP1 MP2
0 100 0
5 100 0
45 40 60
50 0 100
52 0 100
54 100 0
65 100 0

flow rate: 1.2 mL/min
column temp: 40C
detection: 214 nm

Instead of getting a positive or rising baseline, we in most cases get a negative or decreasing baseline. Several people believe that it is do to the change in the acetonitrile concentration, even though they are prepared by weight, they believe that the specific gravity of the buffers are different thus resulting in different acetonitrile concentrations throughout the run. I believe that since they are both prepared by weight the acetonitrile concentration in both should be the same, and thus constant throughout the run.

If someone could help me answer this dilemma of the decreasing baseline I would greatly appreciate it. Thanks.

Chris,

I think the drifting baseline is much more likely due to cation composition of your system since this is a SCX column. What is the cation used with your pH 3.3 phosphate buffer?

Chris,

It doesn't seem like your observation could be explained by density differences between your two eluents. I did a quick check and it looks like your eluent containing perchlorate has a density 2-3% higher than the other eluent. If you argue that the UV absorbing component responsible for the drift arises from the common components in the two eluents than it would follow that these are actually more concentrated in the eluent containing perchlorate since eluent delivery is on a volume basis. Thus, one would expect a rising baseline in the case where density differences are responsible for the problem. However, in retrospect it seems equally unlikely that the problem is due to UV absorbing cations in your buffer since the increasing cation concentration in your perchlorate containing eluent should have the opposite effect on the background (i.e. as the sodium from the sodium perchlorate elutes such a contaminant, the background should be elevated, not reduced). If you argue that the UV absorbing cation is a contaminant in the perchlorate, this should also cause a rising baseline.

It seems like a possible explanation is a refractive index effect. Perchlorate has significantly higher refractive index than many commonly used eluent components. Why are you using a perchlorate salt in a cation exchange separation? Perhaps you should try substituting 0.1 molar sodium sulfate to see if this helps since sulfate has significantly lower specific refractivity than perchlorate.