Column Care

[mordacious]

Hello, great forum here!

I'm very new to rpHPLC and fairly new to GC. I recently took a position analyzing combustion products of biodiesel. I have gotten zero training and I have so many questions that I hardly know where to start

So, column care. I just purchased a Deltabond Fast AK column, 100x4.6mm, 5µm which I use to analyze DNPH derivatives exclusively. My mobile phase is ACN/water.

I use HPLC everyday, how should I store the column overnight? As of right now I'm following a procedure written by the previous analyst which just says to turn the flow down to 0 mL/min and turn the instrument off. Is this sufficient?

Also, how can I monitor any degradation of the column?

Thanks

[Bruce Hamilton]

Gosh, that takes me back. In 1983, I spent most fo the year analysing DNPH derivatives of carbonyl products from engine exhaust gases from engines using alternative fuels such as tallow biodiesel.

The only issue may be the cleanliness of the samples, especially if using impinger sampling and direct injection of the impinger solution into the HPLC. It's a good idea to have a guard column, and I'd get about 500-1000 injections before having to replace the guard due to high pressure - from particulates, rather than gunk.

If you are using an acid, such as perchloric, with your DNPH, then you should ensiure you inject a couple of larger volumes of blank solvent ( without acid ) at the end of the day to remove the acid traces. You don't need to do anything special for close down, if you use good quality water and acetonitrile as mobile phase.

You can check the performance of the column against the CoA supplied with the column, and also by comparing the symmetry and shape of your internal standard ( if you have one ).

Alternatively, you can create standard mixture of derivatives and analyse them each day. Compare the peak retention, resolution, and symmetry/tailing.

Often the DNPH or acetonitrile produce small early-eluting peaks from trace impurities that can also be used as an indication of the column condition if your method doesn't have an internal standard.

Please keep having fun,

Bruce Hamilton

[mordacious]

It looks like I'll be doing this for the next 5 years or, so if they offer me a GRA in the fall, so I'll be here a lot I'm sure

I don't have a column guard as of right now but perhaps I should spend the $100 or so and just get one. I don't use acids so I guess the current shutdown procedure is okay.

Often the DNPH or acetonitrile produce small early-eluting peaks from trace impurities that can also be used as an indication of the column condition if your method doesn't have an internal standard.

I've actually had this problem already. There are 2 or 3 small peaks eluting before formaldehyde in my samples but not the standard. I ran a blank on the DNPH cartridges (from the same lot), along with the ACN I've been using, and they were not there. On Monday I'm working with the MS department to figure out what these are.

Thanks for the help!
Ashley

edit: Did you do any GC work with the exhaust by any chance?

[Uwe Neue]

General rule on column storage: for overnight storage without runs, you can keep the column on the instrument in the mobile phase that you are using. For storage for longer than a standard weekend, put the column into 100% acetonitrile. This is a bit of work, but the column will last longer.

With very few exceptions, peaks don't come from the column, but from sample or injector or mobile phase. For this assay, I cannot see how the peaks could come from the column.

[Bruce Hamilton]

As those peaks are only in your samples, they are likely to be another component of the exhaust reacting with DNPH, most likely NOx or similar. I was able to quantify NOx as well as carbonyls using the DNPH impinger solutions. At the time, I didn't find any literature on the technique, as Chemiluminescence detectors were commonly used for NOx.

I used GC for other components, such as HCs, halogenated compounds. the DNPH-HPLC method is so good for carbonyls that I didn't even try the DNPH-GC methods that were around then.

Around the same time, researcher up the road from me was flying around in a large reseach aircraft ( NASA?) with a huge churning water wheel of DNHP solution passing huge volumes of air to measure atmospheric profiles of carbonyls. That would have been fun..

Uwe has given you the definitive abvice on storage.

Bruce Hamilton

[mordacious]

General rule on column storage: for overnight storage without runs, you can keep the column on the instrument in the mobile phase that you are using. For storage for longer than a standard weekend, put the column into 100% acetonitrile. This is a bit of work, but the column will last longer.

With very few exceptions, peaks don't come from the column, but from sample or injector or mobile phase. For this assay, I cannot see how the peaks could come from the column.

Sounds good, thanks

As those peaks are only in your samples, they are likely to be another component of the exhaust reacting with DNPH, most likely NOx or similar. I was able to quantify NOx as well as carbonyls using the DNPH impinger solutions. At the time, I didn't find any literature on the technique, as Chemiluminescence detectors were commonly used for NOx.

I used GC for other components, such as HCs, halogenated compounds. the DNPH-HPLC method is so good for carbonyls that I didn't even try the DNPH-GC methods that were around then.

Bruce Hamilton

I'll keep NOx compounds in mind tomorrow.

I also use GC for HCs. I'm having trouble finding literature that will give me clues as to what to look for in the exhaust. Everything I find dealing with diesel or biodiesel emissions is on THC concentrations. I can't find much of anything on specific combustion products.

Instead of guessing, buying standards and hoping I can gain some insight we are paying the MS department to do an analysis in a few weeks. This will get very expensive though as we are interested in unburned HCs which requires analyzing the biodiesel itself and buying a column to do this, method development, etc. In order to analyze the emission samples I have to drag my ATD over there to actually introduce the samples via tenax tubes. In other words, it's turning into a much larger project than I expected!

Do you have an tips on maximizing the data I can get from GC before this project starts?

Sorry for all of the questions but I work with a bunch of mech. engineers

Thanks

[Bruce Hamilton]

There' should be heaps of information on the individual HC species in exhaust emissions from biodiesel. Virtually every major manufacturer of Ci engines will have speciated the exhaust profile.

A quick search on the WWW found a Cummins reports at www.biodiesel.org database, and I'm sure there will be others.
Most of the literature will be in environmental journals and regulatory databases, and most auto companies will be happy to share the contents of their reports, if not already published.

The search terms I used were " hydrocarbon emissions speciation biodiesel exhaust " Typical data sources would be :-

1. regulatory databases, eg epa, Carb.
http://www.epa.gov/otaq/regs/toxics/420r06005.pdf
2. Diesel engine companies
3. Southwest Research Institute, esp Chris Sharp's publications
4. SAE Reports ( www.sae.org )
5. WWW.biodiesel.org
6. Enviromental Science journals.
.
There should be plenty of detailed GC data that should give reasonable guidance on what to expect, and contacting a lead author, such as Chris Sharp ( used to be at SWRI ), should give you plenty of suggestions.

There is little point in trying to characterise the diesel ( probably close to 1000 compounds ), as the volatile emssions ( resulting from thermal breakdown and composition ) are usually more significant than the condensable components ( except for PNAs, which are well characterised ).

About the only good to come from the USA's current infatuation with Biodiesel will be the extensive sales of analytical toys and keeping lots of researchers and analysts employed. Not since the introduction of exhaust catalysts and the oil supply crisis in the early 1970s have so many people been tinkering with ICE fuels and pollution. It's very unlikely the planet will benefit.

Please keep having fun,

Bruce Hamilton

[mordacious]

Great, thanks again!