Top 2 tips for HPLC method development

Рет қаралды 13,286

Күн бұрын

Check out our in person and virtual courses:
axionlabs.com/lc-gc-bootcamp-...
axionlabs.com/lc-gc-bootcamp/
axionlabs.com/advanced-hplc/
axionlabs.com/advanced-gc/
axionlabs.com/lc-gc-bootcamp-...
Develop HPLC methods with two tips from Axion Labs:
1. Don't reinvent the wheel - check out working methods in an HPLC column catalog.
2. Perform a scouting run.
Check out axionlabs.com/ for upcoming course dates.

Пікірлер: 17

@shivamverma4478 Жыл бұрын

The way you explain the things, i fall in love with HPLC, you are amazing please Continue it 😊

@TCPLab Жыл бұрын

I didn’t know method development was regarded as the hardest part of doing LC, A while ago I developed a method pretty fast. As you said, just from 10 to 100% of ACN and then you work from there. I just ended making a steep gradient up to 40% in a couple of minutes then a more gentle slope up to 72% and back again to a steeper gradient up to 100% to get junk out. I was expecting 3 peaks but was getting 4. Turns out one compound is a weak acid so I just increased the amount of the phase that contains trifluoroacetic acid and bam! Two peaks became one! Then I just increased the flow rate a bit because I could and that’s it. I still don’t understand why some methods use MetOH and ACN at the same time though, plus water in another bottle and water and TFA in yet another one. All four at the same time. What is really hard for me is troubleshooting the system. I’m trying to bring back to life an Agilent 1100 that was abandoned in a corner for a couple of years and it failed every single test in the Chemstation’s diagnosis tab. Bubbles everywhere, all kinds of baseline noise mixed all together, plus an eternal downwards drift, pressure fluctuations, peaks coming out in pure water injections… It’s driving me insane!

@axionlabs Жыл бұрын

How's it going with your 1100? Were you able to figure it out?

@TCPLab Жыл бұрын

@@axionlabs I did figure it out, everything was dirty, especially the DAD and FLD flow cells, so I washed everything with IPA, changed the degasser and the MCGV, which were not working properly, the outlet ball valve was sticking so I sonicated it, and the solvent inlet glass filters were completely clogged. This was the source of the bubbles, the pump was pulling gas out of the solvents themselves, so I placed them in a hot nitric acid solution for an hour and that's it, I was then finally able to run my 400 samples and finish this project once and for all.

@sorooshsaghebi9465 2 жыл бұрын

Hey there, I was wondering if you have any tips or tricks when it comes to analyzing syrups and stuff with xanthan gum or other thickeners. I have to deal with oral solutions at work everyday and the thickeners mess with my column, my guard and my chromatograms…

@chjawad8776 Жыл бұрын

hi is there any method for carotenoids determination by using hplc (UV-Vis)

@dexpuma Жыл бұрын

any idea on what is a good flow rate to start with for a new method?

@mohamedislambenghaname133 2 ай бұрын

Is there any good method for erythropoietin in albumin serum ?

@ramyabilla466 2 жыл бұрын

the base deactivated columns life time is very less sir can u tell me any process to increase the life of the columns? since the normal HPLC columns can be run upto 4000 injections where as the base deactivated columns like eclipse columns gives a very less life of below 1000 injections.

@gedask98t 2 жыл бұрын

It really depends on what samples you planning to run. If you planning to run biological samples I advise to take extra care during sample preparation. To avoid any proteins which will precipitate on the column. In chemical synthesis world try to avoid running strong oxidators or strong alkaline solutions.