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- Posts: 84
- Joined: Wed Jan 12, 2005 6:17 pm
- Location: Georgia Southern University
I use a Waters Delta 600 system with 717plus A/S to do size exclusion chromatography.
I have been working on a purely analytical scale so far. I work with environmental samples of dissolved organic matter, which are extremely complex mixtures. Since molarities can't be defined for these mixtures the concentrations are in mgC/L. My samples are currently about 40 mg/L. I certainly spread the material out according the to molecular size of the solution components. My initial work with small injections and columns has been successful and we now are seeking to collect fractions as they elute off of the SEC column for hopeful analysis by other analytical techniques. We want to study some properties (like fluorescence) of the fractions as a function of molecular weight. At this time I get barely enough material coming off for any analyses. You could say I want to repeat the experiments on larger, preparative scale so I can get fractions containing more workable amounts of materials.
I will simultaneously move towards 1) injecting greater quantities of sample and and 2) increasing the amount of stationary phase in the flow path.
In the area of issue 1) I have two options.. I can either use more concentrated samples (up to 700 mgC/L) and continue to use the same configuration of my autosampler (100 microliter loop) or I can go to a 2.5 mL loop (with corresponding larger syringe) and keep my sample concentration at about 40 mgC/L.
Hence, my question to the chromatography community is this: Which manner of injecting more sample is chromatographically preferable? Small injections of highly concentrated sample, or larger injections of relatively lower concentration sample?
I am aware of one potential pitfall to increasing my loop size. I was told by Waters tech support that if I went to the larger loop/syringe that the relative variability on injection volumes would increase for small injections.
Also what do you think I should keep in mind as I explore larger columns? Similarly, I see two ways to increase the amount of stationary phase: Increase column inner diameter while keeping length the same or increasing column length while keeping ID constant. Any ideas about pros/cons of these two approaches?
Thanks