What is a good test mix for normal phase HPLC in Hexane:Ethyl Acetate?

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Markus Christ

Hi Sasha,

here are some test mixtures from different vendors:

http://www.perkinelmer.de/CMSResources/Images/44-152609APP_Normal_Phase%20_Test_Mix_%20Brownlee_Analytical_Silica.pdf

http://www.restek.com/catalog/view/6111

https://books.google.de/books?id=6huy4XaToD8C&pg=PA98&lpg=PA98&dq=test+mixture+silica+normal+phase&source=bl&ots=Iwmx6uROPj&sig=EO5MK3uYi7ej_TXBLno_UwH-1qw&hl=de&sa=X&ved=2ahUKEwiXzIrq1MbcAhUDU1AKHeY2AoE4ChDoATAPegQICRAB#v=onepage&q=test%20mixture%20silica%20normal%20phase&f=false

When you are doing NP-HPLC be careful with the water content of your solvents. Fluctuating water content can lead to unreproducible results. Therefore, check the water content regularly with KF titration. Water traces can be removed quantitatively with 2,2'-dimethoxypropane. It reacts with water to methanol and acetone.

Regards Markus

Grzegorz Boczkaj

O-,m-,p- nitroanilines.

Mixture of aromatic hydrocarbons with different number of benzene rings + saturated hydrocarbon+ olefin.

Regards

William Letter

Ethyl acetate does not absorb 100% at 300 nm. It is relative to your reference. It usually leaves too little dynamic headroom at 256 nm and below for use in analyzing samples. We use it for TLC and in some Prep LC applications, but it sees very limited use as a mobile phase for NP analytical LC applications where UV/VIS detection is used. Best to select a more appropriate mobile phase solvent. Be sure and consult a proper table of HPLC solvent properties first: Link: " LC SOLVENT PROPERTIES " http://www.hplctools.com/lcsolvent.htm

"HPLC COLUMN TEST MIXTURE SOLUTIONS; NP & RP EXAMPLES"; https://hplctips.blogspot.com/2017/07/hplc-column-test-mixture-solutions-np.html

Hello Sasha. Yes, in simple terms, it refers to the remaining "space" available that can actually be used. In this context, it refers to the fact that ethyl acetate does absorb a lot of light in the low UV range (hexane does not). This has the effect that very little light is transmitted through the flow cell of the UV/VIS detector. The usable range of most UV/Vis detectors is under 1.4 AU (this value varies, but is approximate). If the ethyle acetate absorbs 1 AU of this, then the remaining linear range may only be 0.4 AU.