Dead volume in possible excess tubing [August 13, 2004]
Posted: Sun Aug 29, 2004 6:51 pm
By DeimerlyC on Friday, August 13, 2004 - 07:34 am:
I have a system that uses post-column derivitization with ninhydrin to detect amino acids. The way the system works is after the eluents have been separated ninhydrin is introduced into the system at a tee, and then the tubing is heated at 125°C.
My problem: I am looking at the instrument manufacturer's tubing and it is PEEK blue, 0.010" ID, from the tee to the detector. From the eluent pump outlet through the injector to the tee the tubing is PEEK yellow, 0.007" ID. The reagent pump is the same. The heated tubing is very long and coiled up inside an oven to give the compounds time to react.
Would you suppose that the manufacturer choose a larger diameter tubing because two flows are going into it? Could the larger tubing be causing band broadening due to its size and length?
Thanks in advance
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By Uwe Neue on Friday, August 13, 2004 - 04:03 pm:
Ninhydrin reactions are not very fast. 125 degrees is improving the speed, but it is still slow. How long is your tubing, and how long is the reaction time? What is your flow rate? There may be better ways to do this than with a coiled tubing, but it depends on your LC conditions.
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By DeimerlyC on Monday, August 16, 2004 - 08:57 am:
Well, it's really hard to tell just exactly how long the coil is. My hunch is it's over 22 feet long (seriously, there's a lot of coil there and it looks like it should go from one end of our lab to the other!). For a flow rate I'm using 0.22ml/min.
Today I'm raising the temperature to 135°C to discern the effect.
I'm used to using a reactor column in the Beckman 6300. It's short and apparently filled with "diamond dust," or so I'm told. Whatever it is, it contains some inert material that helps the amino acids and ninhydrin react well together in a short span.
I'll probably try to splice in one of these reactor columns into my equipment and see the effect, if any, sometime soon.
I have a system that uses post-column derivitization with ninhydrin to detect amino acids. The way the system works is after the eluents have been separated ninhydrin is introduced into the system at a tee, and then the tubing is heated at 125°C.
My problem: I am looking at the instrument manufacturer's tubing and it is PEEK blue, 0.010" ID, from the tee to the detector. From the eluent pump outlet through the injector to the tee the tubing is PEEK yellow, 0.007" ID. The reagent pump is the same. The heated tubing is very long and coiled up inside an oven to give the compounds time to react.
Would you suppose that the manufacturer choose a larger diameter tubing because two flows are going into it? Could the larger tubing be causing band broadening due to its size and length?
Thanks in advance
-------------------------------------------------------------------------------------------------------
By Uwe Neue on Friday, August 13, 2004 - 04:03 pm:
Ninhydrin reactions are not very fast. 125 degrees is improving the speed, but it is still slow. How long is your tubing, and how long is the reaction time? What is your flow rate? There may be better ways to do this than with a coiled tubing, but it depends on your LC conditions.
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By DeimerlyC on Monday, August 16, 2004 - 08:57 am:
Well, it's really hard to tell just exactly how long the coil is. My hunch is it's over 22 feet long (seriously, there's a lot of coil there and it looks like it should go from one end of our lab to the other!). For a flow rate I'm using 0.22ml/min.
Today I'm raising the temperature to 135°C to discern the effect.
I'm used to using a reactor column in the Beckman 6300. It's short and apparently filled with "diamond dust," or so I'm told. Whatever it is, it contains some inert material that helps the amino acids and ninhydrin react well together in a short span.
I'll probably try to splice in one of these reactor columns into my equipment and see the effect, if any, sometime soon.