**Author details**

side, gardenoside, geniposidic acid and loganic acid. All were commercially purchased and only loganin, loganic acid and gardenoside were described for the Apocynaceae family. Several conditions of analyses were studied, including different pH, surfactants, concentra‐ tions of sodium borate and the addition of cyclodextrins (CD) to the buffer, and it was concluded that the less polar DM-*β*-CD added to 50 mM borate solution was the most suitable running buffer. In this condition, only aucubin and catalpol could not be separated, even with the addition of organic solvents and/or valine, urea and barium ion. The greatest advantage of capillary electrophoresis compared to HPLC analyses (the most commonly used technique)

According to [132], capillary electrophoresis can be used to analyze a mixture of eleven iridoid glycosides: unedoside, harpagide, methyl catalpol, morroniside, asperuloside, griselinoside, catalpol, ketologanin, verbenalin, loganin and 10-cinnamoyl catalpol. Only loganin was found in the Apocynaceae family. For the analyses, iridoids were diluted in purified water. A Hewlett-Packard 3DCE system coupled to a diode array detector and equipped with an aircooling device was used. The fused-silica capillary tube measured 80 cm in length, 50 µm in I.D. and 375 µm in O.D. Distance to the detector was 71.5 cm only for UV detection (197 nm, 235 nm, 239 nm and 283 nm). When coupled to a mass spectrometer system (Bruker ESQUIRE) with an electrospray ionization source, the drying gas was nitrogen at 200 ºC and flow-rate 100 L/h. In this case, the distance between injector and UV detector was 20 cm. Other condi‐ tions: sample injection at 50 mbar for 5 s (only UV detection) or 25 s (with MS system); voltage, +20 kV; cartridge temperature, 25 °C; electrolyte solution, 20 mM ammonium acetate with 100 mM sodium dodecyl sulfate (SDS), pH 9.5; sheath liquid, 1 mM lithium acetate mixture to water/methanol (1:1 v/v) at a flow rate of 200 µL/h. When the MS system was used: scan range, 100-550 *m/z*; cut-off, 80 *m/z*; glass capillary exit, 95 V; skimmer, 32 V; electrospray voltage for the capillary, -4.0 kV; for the cylinder, -1,8 kV; for the end plate, -3.5 kV. In the comparison among the counterions sodium dodecyl sulfate (SDS), ammonium dodecyl sulfate and lithium dodecyl sulfate, diluted in water and running buffer, the best resolution for separating iridoid glucosides, lower noise in the MS system, and better repeatability and sensitivity were found with SDS in the running buffer. The volatility of ammonium acetate in buffer enables MS analyses, and concentrations higher than 20 mM did not represent better resolution. Quite the contrary, higher SDS concentrations furnished better results. In the study of the influence of pH, the best one was 9.5, although its influence in the range of 8.7-10.0 was lower than the SDS effect. Good linearity was observed for all the iridoids glucosides analyzed, but in different

The literature describes chromatographic techniques related to the characterization, isolation and purification of iridoids. Most reports show the open column technique as the principal technique used to isolate this class. Also, there have been few studies on counterflow and capillary electrophoresis chromatographies. In general, there has been little scientific invest‐ ment in the area of obtaining iridoids of the Apocynaceae family, despite the great pharma‐

is its speed.

172 Column Chromatography

ranges.

cological importance of this class of constituents.

Ana Cláudia F. Amaral1 , Aline de S. Ramos1 , José Luiz P. Ferreira1,2, Arith R. dos Santos1 , Deborah Q. Falcão2 , Bianca O. da Silva4 , Debora T. Ohana1,5 and Jefferson Rocha de A. Silva3

\*Address all correspondence to: acamaral@fiocruz.br

1 Laboratório de Plantas Medicinais e Derivados, Depto de Produtos Naturais, Farmanguinhos – FIOCRUZ, Manguinhos, Brazil

2 Faculdade de Farmácia – UFF, Niterói, Brazil

3 Laboratório de Cromatografia – Depto. de Química – UFAM, Japiim, Manaus, Brazil

4 Instituto de Pesquisas Biomédicas, Hospital Naval Marcílio Dias, Brazil

5 Fac. de Ciências Farmacêuticas – UFAM, Manaus, Brazil
