Column certificate


(Ashish Bhatnagar) #1

In HPLC column certificate for C18 column why certificates always mention uracil toluene, benzene etc. Peaks


(Jp Ganesh) #2

Uracil – Void Volume
Toluene –Hydrophobic Retention
Naphthalene –Hydrophobic Retention
Biphenyl –Overall Hydophobicity, Asymmetry and Efficiency


(Jp Ganesh) #3

Uracil is a very polar and rarely retained at all by reversed phase columns. For that reason, uracil is used to measure void volume. Be aware also that the internal volume of the instrument between the injection valve and detector also contributes to this volume, so you will likely see slight differences between different instruments. Columns are tested at Restek using configurations that minimize dead volume in tubing and connections, so chances are that your void volume will be slightly larger.
Toluene and naphthalene are used to monitor hydrophobic retention. You can get a good idea of column retention, as well as get a rough idea of resolution between the two at a quick glance.
Since it is the latest eluting peak, the retention time of biphenyl is used to measure overall hydrophobicity, column efficiency, and asymmetry. Hydrophobicity is the overall retention of the column, while efficiency is the number of theoretical plates. Asymmetry is a measurement of how much the shape varies from a perfect Gaussian curve, which is neither tailing nor fronting.
The best way to use a test mixture like this in your lab is to inject the mixture on your new column after equilibration. Doing so will allow you to gauge the combined performance of the LC system and the column. It is important to note that we take great strides to eliminate external factors and measure only the column performance for the QA Report. Because of the potential influence of the LC system, it can be expected that results will not match precisely. The test mixture can be analyzed again periodically, or when troubleshooting performance. Comparing the first results to any subsequent results may be useful in indicating issues with the LC system or column. Keep in mind that some changes are expected over the lifetime of a column when samples are being injected, but you may be able to recognize changes more readily by doing this.
Another helpful piece of information that should be collected from the above analysis is the pressure. You will most likely see some difference in pressure versus what it was when tested at Restek due to mobile phase differences and system pressure. However, if you notice a significant change after equilibration, you can go back to the original conditions and compare the pressure to what it was when you first installed it onto your system. Remember that the flow rate and solvent composition has an effect on pressure.
Getting back to the original question “Why do you need to keep the certificate?”, there are several reasons why this might be helpful. First of all, the certificate records when the column was tested by Restek, it also contains the catalog number, the lot number of packing material used, and the serial number. Should you have any questions or experience any issues, all of this information will be needed to supply you with the best possible service. You may also need this information for your records, as it may be required by regulatory bodies or you may need it to accurately distinguish one column from another.


(Ashish Bhatnagar) #4

Thanks a lot for solving my query


(yasob) #5

Continuing the discussion from Column certificate:

What is the difference between column x-terra C18 and column water symmetry C18 ?