23


propanediol-bonded silica) or for the HILIC mode (triazole, amide...) have been shown to provide adequate retention for polar compounds [11-14]


. An example separation with


seventeen polar pesticides analysed on a cyanopropyl-bonded phase is shown in Figure 1. Some column manufacturers now favour the production of SFC-devoted stationary phases. Ethylpyridine is one of the favourite phases of SFC chromatographers and now proposed by several manufacturers [15-17]


. Octadecylbonded-


silica stationary phases have found use on some occasions, but they need to have some polar function (as polar embedded groups or hydrophilic endcapping groups) in order to allow for sufficient retention of polar compounds.


Table 1: Polar neutral compounds successfully analysed with packed column SFC


development is more LC-like with mobile phase composition gradients generally favoured over pressure gradients. True, in these conditions column efficiency is much less than in GC, but usually more than in HPLC, with identical columns but at higher flow rates.


Common practice nowadays is to use a high back-pressure, above the critical pressure of the mobile phase and a temperature below the critical temperature of the mobile phase, thus the resulting fluid is referred to as ‘subcritical’. Whatever the composition of the mobile phase employed, its real state of matter should be of little concern, as it was shown in the past that the advantageous features of the fluid remain unchanged as there is a continuum of properties when moving from a supercritical fluid to a liquid. The instrumentation used is no different, only the name of ‘supercritical fluid’ chromatography would not be appropriate. However, changing now the name of a well- established method might cause confusion.


Surprisingly, the opposite misconceptions to the ones that existed initially are now prevailing, in that most chromatographers who are inexperienced with carbon dioxide-based mobile phases a prioribelieve that most compounds are too polar to be analysed in SFC. Although SFC is generally viewed as a normal-phase technique, only non-polar compounds are supposedly amenable to carbon dioxide – based mobile phases. In the following, significant examples will be presented to question this opinion.


What are polar compounds and how to analyse them?


The definition of polarity is rather vague, and highly dependent on the chemist and the reference in which they are accustomed to working. We must first distinguish between polar neutral compounds, which in our definition might comprise any compound with an octanol-water partition coefficient log Po/w lower than 2, and ionisable or ionic species, regardless of the hydrophilicity of the non- ionic moiety. As a rule-of-thumb, it was suggested that any compound soluble to at least 1 mg/mL in methanol should be amenable to SFC [4]


usually considered inappropriate to analyse water-soluble compounds [5]


. However, the


possible direct injection of aqueous formulations in the supercritical mobile phase has been demonstrated [6-8]


, which should be


encouraging as regards the possible polarity range amenable to SFC.


To analyse polar compounds, there are two pre-requisites: the stationary phase must be polar enough to retain them, and the mobile phase must be polar enough to allow for a good compound solubility.


SFC stationary phases for polar compound retention


The stationary phase should thus preferably be a polar surface[9] straightforward[10]


. Bare silica would be the most , but a number of other


bonded-silica stationary phases can be of use. Stationary phases designed for normal-phase HPLC (aminopropyl-, cyanopropyl- or


Figure 2: SFC of metoprolol and some related analogues. Kromasil 100-5NH2, 8% methanol, 60 °C, 250 bar, 1.5 ml/min. Reprinted from [61], with permission from Elsevier.


The charge state of ionisable ligands is an important parameter, since the pH control of the carbon dioxide – based mobile phase is impossible. It is however believed that carbon dioxide – alcohol mixtures are acidic, because they react to form alkylcarbonic acid. The estimated pH might be close to 4-5 [18-19]


. In


such acidic conditions, it is likely that ionisable groups of the stationary phase could be charged: for instance, amino or pyridine bonded ligands could be partly cationic, while residual silanol groups may be partly anionic. Poor robustness might thus be expected.


Permanently charged stationary phases are also the object of current research. Zheng et al. [18]


. On the other hand, SFC is


reported on the use of a strong anion- exchange stationary phase with propyltrimethylammonium ligands, while others investigated ionic liquid stationary phases based on phosphonium, pyridinium or imidazolium ligands[20-21]


. Adequate retention


and peak shape was demonstrated for neutral, acidic and basic compounds. According to Zheng et al. [18]


by Jessop et al. [22]


, and based on previous works , it is probable that stationary


phases containing basic ligands such as aminopropyl or ethylpyridine might result in an ionic liquid through reaction with the carbon dioxide – alcohol mixture.


SFC mobile phases for


Page 1  |  Page 2  |  Page 3  |  Page 4  |  Page 5  |  Page 6  |  Page 7  |  Page 8  |  Page 9  |  Page 10  |  Page 11  |  Page 12  |  Page 13  |  Page 14  |  Page 15  |  Page 16  |  Page 17  |  Page 18  |  Page 19  |  Page 20  |  Page 21  |  Page 22  |  Page 23  |  Page 24  |  Page 25  |  Page 26  |  Page 27  |  Page 28  |  Page 29  |  Page 30  |  Page 31  |  Page 32  |  Page 33  |  Page 34  |  Page 35  |  Page 36  |  Page 37  |  Page 38  |  Page 39  |  Page 40  |  Page 41  |  Page 42  |  Page 43  |  Page 44  |  Page 45  |  Page 46  |  Page 47  |  Page 48  |  Page 49  |  Page 50  |  Page 51  |  Page 52  |  Page 53  |  Page 54  |  Page 55  |  Page 56  |  Page 57  |  Page 58  |  Page 59  |  Page 60