Can anyone suggest a suitable solvent system in HPLC (using column of Licrhospher 100 RP-18 5um, Merck, Darmstadt, Germany) to determine Ginkgolide A and B ?
also, take into account the possibility to add something in your mobile phase (mainly acidic additives). you should look at the pKa (or generally physical-chemical properties) of your solutes.
not so many choices in RP: water/methanol or water/acetonitrile. if you have a binary pump you can easily change mobile phase composition and find the better retention factor suitable for you application, and see if you reach the wanted separation and resolution between the two compounds. Look first in literature!!
also, take into account the possibility to add something in your mobile phase (mainly acidic additives). you should look at the pKa (or generally physical-chemical properties) of your solutes.
As some collegues has said you can use water:methanol or ACN:water
Dear Conan,
I use UV/ visible (Waters 2489) detector.
Dear Sanjay, Yuelin and Nicola,
Thank you for your suggestion.
You should check Chem. Pharm. Bull. 2001, 49(9), 1170-1173. It should be available for free on the web : https://www.jstage.jst.go.jp/article/cpb/49/9/49_9_1170/_pdf.
Depends on detection, UV or MS. For UV detection at low wavelength (e.g. 220 nm) I would suggest ACN / water. If you use LC-MS, try water / methanol, e.g. 15-min gradient separation consisted of 10–70% aqueous methanol (J Mass Spectrom. 2005 March; 40(3): 373–379).
I would agree with either water:ACN or water:MeOH. If you know the max absorption and it is lower than 215nm I would prefer ACN over methanol. If you do not know your max absorption then I would advise you scan your analytes on a standard UV/Vis first and then choose the best wavelength.
If it is very low (les than 210nm) you will be restricted using a gradient elution.
As nicola said, using a dilute acid:water is a good idea. For terpenoids, often formic or phosphoric acid is used but you will need to check the literature.
You can try with water-Acetonitrile (90:10, 80:20 and 90:10 plus 0.1% TFA)
Dear Conan,
Thank you very much for your explanation and your suggestion.
Hi!! May be you could be try this method: LiChrocart Lichrospher RP 18 250 x 4 (5 mcm). Flow 1.0 mL/min; 270 nm; Vol Iny 10 mcL; mobile phase = H3PO4 1% v/v : MeOH(48:52). Solver for the sample = MeOH: H2O: HCl conc (50:20:8). I wish the best resoult!! Take care, see you...
There are some papers by Kanfer and Dubber in the literature. There is a PhD thesis by Dubber located at the Rhodes University Library e-repository that may be of assistance
It will be better to use mobile phase without any additives to start with.If problems occurs in peak shape or retention repeatability,then one can switch over to buffers/modifiers etc.Also without buffers/additives column will show less back pressure as well as good life for injections.
As per USP procedure, Methanol & Water itself are sufficient. We can try with various ratios of this.
Pl. refer www.merckmillipore.com/chromatography website for initial applications.I will revert back to you.
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