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Application of Hydrophilic Interaction Liquid Chromatography for the Quantification of Flavonoids in Genista tinctoria Extract.

Sentkowska A, Biesaga M, Pyrzynska K - J Anal Methods Chem (2016)

Bottom Line: In the case of bare silica column more or less apparent dual retention mechanism was observed, depending on the water component content in the mobile phase.ZIC-HILIC showed better selectivity (in comparison to silica column) with the detection limit of 0.01 mg/L (only for rutin was 0.05 mg/L).Finally, this chromatographic procedure was validated and applied for the determination of some flavonoids in Genista tinctoria L. extract.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland.

ABSTRACT
Hydrophilic interaction chromatography (HILIC) was employed to investigate chromatographic behavior of selected flavonoids from their different subgroups differing in polarity. Chromatographic measurements were performed on two different HILIC columns: unmodified silica (Atlantis-HILIC) and zwitterionic sulfoalkylbetaine (SeQuant ZIC-HILIC). Separation parameters such as content and type of organic modifier were studied. On ZIC column retention factors were observed to be inversely proportional to the buffer content in the mobile phase, which is the typical partitioning mechanism. In the case of bare silica column more or less apparent dual retention mechanism was observed, depending on the water component content in the mobile phase. ZIC-HILIC showed better selectivity (in comparison to silica column) with the detection limit of 0.01 mg/L (only for rutin was 0.05 mg/L). Finally, this chromatographic procedure was validated and applied for the determination of some flavonoids in Genista tinctoria L. extract.

No MeSH data available.


Related in: MedlinePlus

The retention factor as a function of ACN or MeOH content in the mobile phase. (a) Bare silica column; (b) ZIC column.
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fig1: The retention factor as a function of ACN or MeOH content in the mobile phase. (a) Bare silica column; (b) ZIC column.

Mentions: The effect of organic solvent content in the mobile phase on the retention of flavonoids was investigated using acetonitrile and methanol. The study was carried out in the presence of ammonium formate at pH 7.0, which refers to the aqueous portion of the mobile phase. It should be noticed that the apparent pH value of the mobile phase containing high percentage of organic solvent is different from that of the water component [23, 24]. At the acidic pH, the acidity of such mobile phase is likely higher than the aqueous acid or buffer solution applied for eluent preparation. Mobile phase pH plays a key role in retention due to its impact on ionization state of both analytes and stationary phase. Silanol groups are weak acids and become deprotonated (negatively charged) at higher pH values. Positively charged compounds can have electrostatic interaction with such groups and can result in stronger retention. Therefore, it is important to consider the impact of mobile phase pH on the stationary phase and the ionization state of the analytes. The pKa values of our analytes are in range of 6.4–7.1. Thus at pH 7.0 they exist in the equilibrium between the charged and the neutral forms. The obtained retention factors were plotted against the organic solvent content in the range of 40–95% (v/v) as shown in Figure 1.


Application of Hydrophilic Interaction Liquid Chromatography for the Quantification of Flavonoids in Genista tinctoria Extract.

Sentkowska A, Biesaga M, Pyrzynska K - J Anal Methods Chem (2016)

The retention factor as a function of ACN or MeOH content in the mobile phase. (a) Bare silica column; (b) ZIC column.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4940558&req=5

fig1: The retention factor as a function of ACN or MeOH content in the mobile phase. (a) Bare silica column; (b) ZIC column.
Mentions: The effect of organic solvent content in the mobile phase on the retention of flavonoids was investigated using acetonitrile and methanol. The study was carried out in the presence of ammonium formate at pH 7.0, which refers to the aqueous portion of the mobile phase. It should be noticed that the apparent pH value of the mobile phase containing high percentage of organic solvent is different from that of the water component [23, 24]. At the acidic pH, the acidity of such mobile phase is likely higher than the aqueous acid or buffer solution applied for eluent preparation. Mobile phase pH plays a key role in retention due to its impact on ionization state of both analytes and stationary phase. Silanol groups are weak acids and become deprotonated (negatively charged) at higher pH values. Positively charged compounds can have electrostatic interaction with such groups and can result in stronger retention. Therefore, it is important to consider the impact of mobile phase pH on the stationary phase and the ionization state of the analytes. The pKa values of our analytes are in range of 6.4–7.1. Thus at pH 7.0 they exist in the equilibrium between the charged and the neutral forms. The obtained retention factors were plotted against the organic solvent content in the range of 40–95% (v/v) as shown in Figure 1.

Bottom Line: In the case of bare silica column more or less apparent dual retention mechanism was observed, depending on the water component content in the mobile phase.ZIC-HILIC showed better selectivity (in comparison to silica column) with the detection limit of 0.01 mg/L (only for rutin was 0.05 mg/L).Finally, this chromatographic procedure was validated and applied for the determination of some flavonoids in Genista tinctoria L. extract.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland.

ABSTRACT
Hydrophilic interaction chromatography (HILIC) was employed to investigate chromatographic behavior of selected flavonoids from their different subgroups differing in polarity. Chromatographic measurements were performed on two different HILIC columns: unmodified silica (Atlantis-HILIC) and zwitterionic sulfoalkylbetaine (SeQuant ZIC-HILIC). Separation parameters such as content and type of organic modifier were studied. On ZIC column retention factors were observed to be inversely proportional to the buffer content in the mobile phase, which is the typical partitioning mechanism. In the case of bare silica column more or less apparent dual retention mechanism was observed, depending on the water component content in the mobile phase. ZIC-HILIC showed better selectivity (in comparison to silica column) with the detection limit of 0.01 mg/L (only for rutin was 0.05 mg/L). Finally, this chromatographic procedure was validated and applied for the determination of some flavonoids in Genista tinctoria L. extract.

No MeSH data available.


Related in: MedlinePlus