Chromatography Forum

Hello

I need some help understanding the theoretical plates calculation of merck columns, when I got the column it cames with a certificate thats saids

minimun theoretical plates 80000 the certificate saids it haves 101569 under these conditiones

Acetonitrile/water (75/25 v/v)
1.3 ml/min
1 ul injection volume
UV 254 nm
Detector cell volumen: 3.2 uL
Extra-column volume: 25 uL

I made my theoretical plate determination and I got 16500 under these conditions

Acetonitrile/water (75/25 v/v)
1.3 ml/min
1 ul injection volume
UV 254 nm
Detector cell volumen: 13 uL

I returned the column for test and some time later I ask merck to solve the issue they gave to me a the test made to the column they got 62000 theoretical plates under these conditions


Acetonitrile/water (75/25 v/v)
1.3 ml/min
10 ul injection volume
UV 254 nm
Detector cell volumen: 13 uL

The injection volume is highly incresed, I ask for an explanation but the only thing they told me is that I will never get the theoretical plates from factory because of the detector cell volume.

It´s possible that the detector cell volumen is so critical in the determination that i am getting way less than they get on factory or they are trying to just to gave explanations to don´t accept my column is bad?

Unfortunately you don't tell us anything about the column dimension and particle size.

There may also be a mix-up of units. Are the numbers from Merck "per meter" whereas your number is "per column"?

Also essential is the diameter and length of the tubing to detector.

You may want to read the topic in the Waters "Beginners Guide to UPLC"
http://www.waters.com/waters/en_US/Chro ... =134803614

Thanks a lot for your reply.

The column is a Purospher Star RP-18 endcapped, 250mm lenght, 4.6 mm ID, 5um particle size.

The Column was analyzed on fabric with Separation Efiicency (N/m), Peak symmetry (AS and T-USP).

The theoretical plates on lab where determine with USP method, On a HPLC Hitachi Chromaster with DAD Detector.

Ok I got it what you said about per Meter and per column with the data of the column I gave to you. it can be determine also still doesn´t meet specs or I do have to make another correction?

I don't know if Merck calculates the "real" meaning column-only plate count whereas you can only measure the total plate count as the sum of plate and system.
Then your system is not optimized to determine column performance but doing it's daily job of separation. That's what the customer service is telling you.
Comparing your count and the one of the customer service they are pretty close if taken as per meter.
Personally, as rule of thumb I'm ok with a reduced plate height of 3-4 (particle diameters/plate) on an HPLC system.
That's what your column delivers.
Also the flow of 1.3 ml/min seems a bit high too me for a 4.6mm, 5 um column. Maybe with a slower flow the efficiency may even increase.

If your result of 16500 was calculated per column and the result of 62000 was calculated per meter by the column manufacture, then the two are essentially the same.