Analysis of aminofluorescein-fatty acid derivatives by capillary electrophoresis with laser-induced fluorescence detection at the attomole level: application to mycobacterial fatty acids

In the present work, a new method of purification for antithrombin was developed using an expanded bed chromatography technique. Milk fat was removed by centrifugation and caseins were precipitated selectively by addition of zinc chloride. Crude skim milk was then directly fed to an expanded bed column containing the ion-exchange matrix. The use of a cation-exchanger (P-11) resulted in 100% adsorption and 13% recovery whereas the use of an anion-exchanger (DE-52) resulted in 100% adsorption and 84% recovery and up to five-fold purification of antithrombin. The buffer, 25 mM Tris-HCl pH 8.0; the eluting agent, 2 M (NH4)2SO4; and 100% expansion of settled bed were determined to be the optimum conditions for the purification of antithrombin by ion-exchange expanded bed chromatography. A comparison of column diameters revealed that the elution yields increase by two-fold while the column diameter increases from 1 to 2.5 cm. However, antithrombin III was concentrated to a higher degree by using the column with an internal diameter of 1 cm.     

Enhanced performance of expanded bed chromatography on rigid superporous adsorbent matrix

Rigid spherical macroporous adsorbent beads with surface hydroxyl groups were prepared by cross-linking of cellulose. These beads had diameter in the range 100-200 microm and a mean pore size of about 3 microm with about 60% pore volume. The matrix (bulk density approximately 1600 kg m(-3)) could be expanded into a stable bed and used for protein chromatography. Chromatographic runs were performed on a 10 mm diameter column under non-retaining and retaining conditions on the prepared matrix (called Celbeads) and performance of the runs was measured in terms of the height equivalent to a theoretical plate (HETP). The HETP curves in both packed and expanded bed modes followed profiles typical of macroporous adsorbents, i.e. increasing and levelling with velocity. Unimpaired performance of the matrix at increasing flow-rates permitted expanded bed elution of adsorbed solutes without loss of efficiency in terms of purification factor and product concentration. As a model system, Celbeads was used to purify lactate dehydrogenase from porcine muscle homogenate by dye-affinity chromatography. The prepared matrix provided about 100 theoretical plates per meter for the enzyme system at a linear flow velocity of 1.27 cm x min(-1) in an expanded bed elution mode, and gave enzyme yields of 100% with a purification factor of 31 using an optimized procedure. The adsorbent could be cleaned in place with 5 M urea and used repeatedly without loss of performance.     

Affinity purification of proteins using expanded beds.

The use of expanded beds of affinity adsorbents for the purification of proteins from feedstocks containing whole or broken cells is described. It is demonstrated that such feedstocks can be applied to the bed without prior removal of particulate material by centrifugation or filtration thus showing considerable potential for this approach in simplifying downstream processing flow-sheets. A stable, expanded bed can be obtained using simple equipment adapted from that used for conventional packed bed adsorption and chromatography processes. Circulation and mixing of the adsorbent particles is minimal and liquid flow through the expanded bed shows characteristics similar to those of plug flow. Frontal analysis performed with the highly selective affinity system involving the adsorption of human polyclonal immunoglobulin G onto Protein A Sepharose Fast Flow indicate that the adsorption performance of the expanded bed is similar to that achieved when the same amount of adsorbent is used in a packed configuration at the same volumetric flow-rate. The adsorption performance of the expanded bed was not diminished when adsorption was carried out in the presence of intact yeast cells. Batch adsorption experiments also indicated that the adsorption characteristics of the affinity system were not greatly altered in the presence of cells in contrast to results from a less selective ion-exchange system. An expanded bed of Cibacron Blue Sepharose Fast Flow was used to purify phosphofructokinase from feedstock of disrupted yeast prepared by high pressure homogenisation without the need for prior removal of particulate material. The potential for the use of expanded beds in large scale purification systems is discussed.     

Separation of sympathomimetic amines of abuse and related compounds by micellar electrokinetic chromatography.

Separation of twelve sympathomimetic amines and related compounds by micellar electrokinetic chromatography (MEKC) with UV absorbance detection is described. These amines were well separated within 25 min using 50 mM sodium tetraborate solution containing 15 mM sodium dodecylsulfate (SDS) of pH 9.3 as a running solution and detected at 210 nm. MEKC was performed with an applied voltage of 13 kV at 25 degrees C using a fused-silica capillary (50 cm x 75 mm i.d.) with effective length of 37.5 cm. The detection limits of these compounds were in the range from 4 to 97 fmol/injection at a signal-to-noise ratio (S/N) of 3. The reproducibility of the method expressed as relative standard deviation (RSD) for within-day (n=6) and between-day (n=5) assays was less than 4.8 and 8.8%, respectively. The proposed method could be applied to the determination of an anorectic drug, phentermine, in Chinese tea with a detection limit of 99 microg/g (105 fmol/injection, S/N=3).     

Evaluation of new high-density ion exchange adsorbents for expanded bed adsorption chromatography

New adsorbents Q HyperZ and CM HyperZ composed of hydrogel-filled porous zirconium oxide particles were evaluated for expanded bed adsorption applications in the present work. The HyperZ adsorbents have wet density of 3.16 g ml(-1), particle size of 44.5-100.8 microm and average sphere diameter of 67 microm. The bed expansion as the function of flow velocity and fluid viscosity was measured and correlated with Richardson-Zaki equation. The suitable expansion factor was considered less than 2.5, while the corresponding flow velocity was about 450 cmh(-1). Liquid mixing in the bed was determined to evaluate the stability of expanded bed. The Bodenstein numbers tested were higher than 40 and the axial mixing coefficients (D(ax)) were between 0.5 and 9.7x10(-6)m(2)s(-1), which demonstrated that a stable expanded bed could be formed under suitable operation conditions. Bovine serum albumin (BSA) and lysozyme were used as model proteins to estimate the adsorption capacities of Q and CM HyperZ, respectively. The maximum equilibrium adsorption of Q and CM HyperZ could reach 45.7 and 27.2 mg g(-1) drained adsorbents, respectively. It was found that yeast cells had little influence on the adsorption capacities of the two adsorbents tested. The dynamic adsorption capacity of BSA at 10% breakthrough with Q HyperZ was 35.9 mg g(-1) drained adsorbent at flow velocity of 100 cm h(-1) for packed bed adsorption. The values for expanded bed adsorption were 34.4 mg g(-1) drained adsorbent at flow velocity of 200 cm h(-1), 33.6 mg g(-1) drained adsorbent at 300 cm h(-1) and 31.7 mg g(-1) drained adsorbent 400 cm h(-1). The results demonstrated that Q HyperZ and CM HyperZ are suitable for expanded bed adsorption of biomolecules.     

Superporous agarose beads as a hydrophobic interaction chromatography support

Superporous agarose beads were used as a support for hydrophobic interaction chromatography. These beads have large connecting flow pores in addition to their normal diffusion pores. The flow pores, which are approximately one fifth of the overall diameter of the superporous agarose beads, were earlier shown to give the beads improved mass transfer properties relative to homogeneous agarose beads (Gustavsson and Larsson, J. Chromatogr. A, 734 (1996) 231–240). Superporous agarose beads and homogeneous agarose beads of the same particle size range (106–180 μm) were derivatized with phenyl groups. The properties of the superporous beads were then compared with the homogeneous beads in the separation of a mixture of three model proteins (ribonuclease A, lysozyme and bovine serum albumin) at various superficial flow velocities from 30 to 600 cm/h. The superporous beads gave satisfactory separation at flow velocities five times higher than was possible for homogeneous beads. The performance of the two types of beads was also compared in the purification of lactate dehydrogenase from a beef heart extract at a superficial flow velocity of 150 cm/h. The superporous beads performed considerably better, leading to twice the purification factor and twice the concentration of the desired product. The results were interpreted using the theoretical treatment given by Carta and Rodrigues (Carta and Rodrigues, Chem. Eng. Sci., 48 (1993) 3927).     

DNA binding during expanded bed adsorption and factors affecting adsorbent aggregation

DNA-induced aggregation and contraction of expanded bed adsorption chromatography beds have been examined using strong anion exchanger Q HyperZ and calf thymus DNA in buffers containing added NaCl. Two batches of adsorbent with different ionic capacities were used allowing the effects of different ligand densities to be examined. Very high dynamic binding capacities at 10% breakthrough were found in the absence of added salt. However, the highest binding capacities ( approximately 10 and approximately 19mgDNAml(-1) gel) were found in buffers containing added salt at concentrations of either 0.25 or 0.35M, for the low and high ligand density adsorbents, respectively. Bed contraction was observed, but did not correlate with dynamic binding capacity or with the amount of DNA loaded. No differences in bed contraction were seen by varying the concentration of DNA loaded in the range of 20-80mugml(-1) even though the dynamic binding capacity was reduced as DNA concentration was increased. The extent of bed contraction during DNA loading was found to be a function of added salt concentration and ligand density of the adsorbent. The results imply that ligand density significantly affects the salt tolerance of anion exchangers when binding DNA. However, more importantly, with the adsorbents examined here, attempts to reduce bed aggregation by feedstock conditioning with added salt may increase DNA binding leading to a reduction in expanded bed adsorption performance compromising protein capture in real feedstocks.     

Expanded Bed Adsorption Chromatography for Recovery of Phycocyanins from the Microalga Spirulina Platensis

We carried out the purification of C-phycocyanin and allophycocyanin from Spirulina platensis taking advantage of the adsorption properties of the expanded beds. Initially, phycobiliproteins were released from the microalga cells by osmotic shock. Next, phycocyanins were recovered by applying the centrifuged cell suspension directly to the anion exchanger Streamline-DEAE using expanded bed columns, equilibrated with 50 mM sodium phosphate buffer, pH 7.0. After adsorption, washing was carried out in the expanded-bed mode. Having removed unbound proteins and cellular debris, the bed was allowed to sediment and phycocyanins rich solution was eluted with a downward flow of 500 mM sodium phosphate buffer, pH 7.0. Finally, we utilized conventional gel filtration and ion exchange chromatography methods for separation and purification of C-phycocyanin and allophycocyanin. The purification steps were monitored using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the purity of recovered phycocyanins was confirmed by absorption and emission spectroscopy. The main advantage of this new method is the high yield achieved in the steps of product extraction and adsorption by expanded bed adsorption, so reducing both processing times and costs.     

Use of expanded bed adsorption to purify flavonoids from Ginkgo biloba L.

Three techniques (liquid–liquid extraction, packed bed adsorption and expanded bed adsorption) have been compared for the purification of flavonoids from the leaves of Ginkgo biloba L. A crude Ginkgo extract was obtained by refluxing with ethanol for 3 h. The yield of flavonoids achieved by this crude extraction was about 19% (w/w) and the purity of flavonoids in the concentrated extract was between 1.9 and 2.3% (w/w). The crude extract was then dissolved in deionized water and centrifuged where necessary to prepare clarified feedstock for further purification. For the method using liquid–liquid extraction with ethyl acetate, the purity, concentration ratio and yield of flavonoids were 25.4–31.0%, 16–18 and >98%, respectively. For the method using packed bed adsorption, Amberlite XAD7HP was selected as the adsorbent and clarified extract was used as the feedstock. The dynamic adsorption breakthrough curves and elution profiles were measured. For a feedstock containing flavonoids at a concentration of 0.25 mg/mL, the appropriate loading volume to reach a 5% breakthrough point during the adsorption stage was estimated to be 550–600 mL for a packed bed of volume 53 mL and a flow rate of 183 cm/h. The results from the elution stage indicated that the majority of impurities were eluted by ethanol concentrations of 40% (v/v) or below and efficient separation of flavonoids from the impurities could be achieved by elution of the flavonoids with 50–80% ethanol reaching an average purity of ∼25%. The recovery yield of flavonoids using the packed bed purification method was about 60% of the flavonoids present in the clarified feedstock (corresponding to around 30% for the total flavonoids in the unclarified crude extract). For the method using expanded bed adsorption also conducted with Amberlite XAD7HP as the adsorbent, the optimal operation conditions scouted during the packed bed experiments were used but unclarified crude extract could be loaded directly into the column. For an expanded bed with a settled bed height of 30 cm, the loss of flavonoids in the column flow-through was about 30%. The two-step elution protocol again proved to be effective in separating the adsorbed impurities and flavonoids. More than 96% of the bound impurities were completely removed by 40% ethanol in the first elution stage and less than 4% remained in the final product eluted by 90% ethanol in the second elution stage. Also, ∼74% of the adsorbed flavonoids on column (corresponding to 51% of the total flavonoids in the unclarified feedstock) were recovered in the product. In addition to higher recovery yield, the average process time to obtain the same amount of product was decreased in the expanded bed adsorption (EBA) process. The results suggest that the adoption of EBA procedures can greatly simplify the process flow sheet and in addition reduce the cost and time to purify flavonoids from Ginkgo biloba. These results clearly demonstrate the potential for the use of EBA to purify pharmaceuticals from plant sources.     

Particle size and density distributions of two dense matrices in an expanded bed system

The size and density distributions of two commercial media, that is, Streamline particles and 6% agarose coated steel beads (6AS), in an expanded bed system has been studied with a glass column (26 mm I.D.) modified by side ports. The Streamline particles have a broad size distribution but a relatively uniform density, while the 6AS beads have both broad size and density distributions. The effect of liquid-phase flow velocity, liquid viscosity and settled bed height on the particle size and density distributions is investigated. It is found that the radial mean particle size and density of the two matrices are uniform, while axial classifications are obvious in the expanded beds. For the Streamline, the volume-weighted mean particle size decreases linearly with increasing expanded bed height. For the 6AS beads, however, the mean particle size is even in the axial direction, but the particle density decreases exponentially with the increase of bed height. Moreover, the mean particle size of the Streamline or the density of the 6AS beads is well expressed as a function of the normalized bed height (that is, the ratio of the distance from bed bottom to the expanded bed height). The liquid flow-rate, liquid viscosity and settled bed height influence the mean axial size or density distribution by affecting the expanded bed height.     

Determination of bisphenol A and 10 alkylphenols in serum using SDS micelle capillary electrophoresis with gamma-cyclodextrin.

A new micelle capillary electrophoresis based on cyclodextrin micellar electrokinetic chromatography (MEKC) for the determination of bisphenol A and 10 alkylphenols in rat serum is reported. Several surfactants and dextrins were studied. Bisphenol A and alkylphenols were separated using a 50 microm x 50 cm capillary with 20 mM borate phosphate buffer (pH 8.0) containing 20 mM sodium dodecylsulfate and 5 mM gamma-cyclodextrin as carrier. The method could determine 0.6-2000 microg/mL of phenols in 100 microL serum by photometric detection at 214 nm. Using 2.0 mL serum, 1.0 ng/mL of phenols could be determined. The relative standards deviations were 6.3-7.7% at 10 microg/mL in serum. The recoveries were 91.8-93.0% with 10 microg/mL serum samples.     

Adsorption Strategy of Plasmid DNA Nanoparticulate: Preparative Purification by a Simple Custom Expanded Bed Column

This paper summarizes the critical examination of the hydrodynamic performance of the NBG expanded bed contactor operated with streamline-DEAE adsorbent under various operating conditions for expanded bed adsorption of plasmid DNA nanoparticles from alkaline lysate. The purification process is not RNase-free. In this study, a rapid and efficient scaleable purification protocol obtaining, plasmid DNA nanoparticles (average size of 40 nm) with a high purity level for use as therapeutic agent in customized NBG expanded bed columns was developed. This technique allows efficient levels of binding to the column media and vector purification without centrifugation or filtration steps. Residence time distribution (RTD) studies were exploited to achieve the optimal condition of plasmid DNA nanoparticle (pDNA) recovery upon anion exchange adsorbent in this contactor. In addition, the purification experiments were carried out in the expanded bed columns with settle bed height of 6.0 ± 0.2 cm. NaCl gradient elution enabled the isolation of supercoiled plasmid from low-Mr RNA, cDNA and plasmid variants. Subsequently dynamic binding capacity of the adsorbent was calculated while these values decreased with increase in flow velocity. Moreover, the effect of pH upon the performance of this recovery process and the feedstock volume upon the expanded bed anion exchange purification was investigated. The results demonstrated that separation of low-Mr RNA from plasmid DNA isoforms in the range of pH between 5.5 and 7.5 is achievable in this column. The yield of recovery of pDNA in optimal condition was higher than 88.51% which was a superior result in one-pass frontal chromatography. The generic application of simple customized NBG expanded bed column and its potential for the purification and recovery of plasmid DNA as a nanoparticulate bioproduct is strongly discussed.     

Influence of column design on process-scale ion-exchange chromatography

The performance of two new designs of pump-packed axial flow process chromatography columns have been evaluated for the preparative anion-exchange chromatography of hen egg-white proteins using Whatman Express-Ion Exchanger Q. A 16 1 Side-Pack column and a 24 1 IsoPak column containing Express-Ion Q were used in this study. In each case ca. 20 1 feedstock containing 5-7 g protein/l, was applied per litre packed bed at flow-rates of ca. 150 and 300 cm/h. In each case the ovalbumin binding capacity was ca. 70 g/l packed bed with ca. 100% (w/w) recovery of applied protein. A clean-in-place procedure involving storage in 0.5 M NaOH was effective in maintaining chromatographic performance in all cases. These data were consistent with our previous work using the more traditionally configured slurry-packed axial flow columns. Each of these column designs were easy to use facilitating rapid packing with this adsorbent and in the case of IsoPak rapid pump unpacking. The introduction of these column designs significantly improves the task of column packing, hitherto a labour intensive, physically demanding and potentially unreproducible process.     

Combination of integrated expanded bed adsorption chromatography and countercurrent chromatography for the direct extraction and purification of pseudohypericin and hypericin from St. John's wort (Hypericum perforatum L.)

St. John's wort has attracted particular attention because of its beneficial effects as an antidepressant, antiviral, and anticancer agent. A method for the combination of integrated expanded bed adsorption chromatography and countercurrent chromatography for the simultaneous extraction and purification of pseudohypericin and hypericin from the herb is presented in this paper. Firstly, the constituents were extracted and directly adsorbed by expanded bed adsorption chromatography under optimal conditions. The stepwise elution was then performed by expanded bed adsorption chromatography that enriched the targets with higher purities and recoveries compared to other methods. Secondly, the eluent fractions from expanded bed adsorption chromatography were further separated by two‐step high‐speed countercurrent chromatography. A two‐step high‐speed countercurrent chromatography method with a biphasic solvent system composed of n‐hexane/ethyl acetate/methanol/water with a volume ratio of 1:2:1:2 was performed by stepwise changing the flow rate of the mobile phase. Consequently, 5.6 mg of pseudohypericin and 2.2 mg of hypericin with purities of 95.5 and 95.0%, respectively, were successfully obtained from 40 mg of crude sample.     

Direct recovery of recombinant nucleocapsid protein of Nipah virus from unclarified Escherichia coli homogenate using hydrophobic interaction expanded bed adsorption chromatography

A direct recovery of recombinant nucleocapsid protein of Nipah virus (NCp-NiV) from crude Escherichia coli (E. coli) homogenate was developed successfully using a hydrophobic interaction expanded bed adsorption chromatography (HI-EBAC). The nucleic acids co-released with the recombinant protein have increased the viscosity of the E. coli homogenate, thus affected the axial mixing in the EBAC column. Hence, DNase was added to reduce the viscosity of feedstock prior to its loading into the EBAC column packed with the hydrophobic interaction chromatography (HIC) adsorbent. The addition of glycerol to the washing buffer has reduced the volume of washing buffer applied, and thus reduced the loss of the NCp-NiV during the washing stage. The influences of flow velocity, degree of bed expansion and viscosity of mobile phase on the adsorption efficiency of HI-EBAC were studied. The dynamic binding capacity at 10% breakthrough of 3.2 mg/g adsorbent was achieved at a linear flow velocity of 178 cm/h, bed expansion of two and feedstock viscosity of 3.4 mPa s. The adsorbed NCp-NiV was eluted with the buffer containing a step gradient of salt concentration. The purification of hydrophobic NCp-NiV using the HI-EBAC column has recovered 80% of NCp-NiV from unclarified E. coli homogenate with a purification factor of 12.5.     

Cytotoxic activity of polyacetylene compounds in Panax ginseng C. A. Meyer

The effects of the three polyacetylene compounds, panaxynol, panaxydol and panaxytriol, on in vitro-cell growth were studied. These compounds are much different in their water-solubility. In order to increase water-solubility, solid complexes of polyacetylene compounds with alpha-cyclodextrin (CD) were prepared. Accurate concentrations of the active compounds in a culture medium were determined by gas chromatography. All of these CD-complexes inhibited the growth of various kinds of cultured cell lines in a dose-dependent fashion. The cell growth inhibitory activity of these complexes was much stronger against malignant cells than against normal cells. A continuous contact between the compounds and target cells was not necessarily required for growth inhibition. And the inhibition was cytotoxic at high concentrations and cytostatic at low concentrations. These findings indicate that these polyacetylene compounds' mode of action is more dose-dependent than time-dependent. The panaxynol, panaxydol and panaxytriol contents in red ginseng powder were 250, 297 and 320 micrograms/g, respectively.     

A cation exchange resin bead-based microscale electrolytic suppressor for capillary ion chromatography

A cation exchange resin (CER) bead-based microscale electrolytic suppressor for capillary ion chromatography (CIC) has been fabricated and evaluated. Relative to common ion exchange membrane, the use of CER beads presents a simple way to fabricate a microscale suppressor due to its small surface area and spherical shape. The internal volume of the device is ∼600 nL, which matches the requirements by CIC. Continuous online regeneration is readily achieved using pure water or diluted sulfuric acid as the regenerant. The use of a cation exchange functionalized monolithic segment inserted between two CER beads greatly reduces the electrical resistance and improves the suppression efficiency. The device works well only in the electrolytic mode, chemical regeneration alone is ineffective. The suppressed background is ∼3 μS/cm for 10 mM KOH eluent generated online by a microscale KOH electrolytic generator operated at the flow rate of 3 μL/min. The preliminary application for capillary ion chromatography has been demonstrated.     

Production of cyclodextrins in a fluidized-bed reactor using cyclodextrin-glycosyl-transferase

Cyclodextrin-glycosyl-transferase (EC2.4.1.19), produced by Wacker (Munich, Germany), was purified by biospecific affinity chromatography with β-cyclodextrin (β-CD) as ligand, and immobilized into controlled pore silica particles (0.42 mm). This immobilized enzyme (IE) had 4.7 mg of protein/g of support and a specific activity of 8.6 μmol of β-CD/(min·g_IF) at 50°C, pH 8.0. It was used in a fluidized-bed reactor (FBR) at the same conditions for producing cyclodextrins (CDs) with 10% (w/v) maltodextrin solution as substrate. Bed expansion was modeled by the Richardson and Zaki equation, giving a good fit in two distin ctranges of bed porosities. The minimum fluidization velocity was 0.045 cm/s, the bed expansion coefficient was 3.98, and the particle terminal velocity was 2.4 cm/s. The FBR achieved high productivity, reaching in only 4 min of residence time the same amount of CDs normally achieved in a batch reactor with free enzyme after 24h of reaction, namely, 10.4 mM β-CD and 2.3 mM γ-CD.     

Nd-Fe-B alloy-densified agarose gel for expanded bed adsorption of proteins

Novel dense composite adsorbents for expanded bed adsorption of protein have been fabricated by coating 4% agarose gel onto Nd-Fe-B alloy powder by a water-in-oil emulsification method. Two composite matrices, namely Nd-Fe-B alloy-densified agarose (NFBA) gels with different size distributions and densities, NFBA-S (50-165 microm, 1.88 g/ml) and NFBA-L (140-300 microm, 2.04 g/ml), were produced. Lysozyme was used as a model protein to test the adsorption capacity and kinetics for the NFBA gels modified by Cibacron blue 3GA (CB-NFBA gels). Liquid-phase dispersion behavior in the expanded beds was examined by measurements of residence time distributions, and compared with that of Streamline SP (Amersham-Pharmacia Biotech, Sweden). The dependence of axial mixing in the expanded beds on flow velocity, bed expansion degree. settled bed height, and viscosity of liquid phase was investigated. Breakthrough curves of lysozyme in the expanded beds of the CB-NFBA gels were also examined. The dynamic binding capacity at 5% breakthrough was 23.3 mg/ml matrix for the CB-NFBA-S gels, and 16.7 mg/ml matrix for the CB-NFBA-L, at a flow velocity of 220 cm/h. The results indicate that the NFBA gels are promising for expanded bed adsorption of proteins.     

Characterization and evaluation of a macroporous adsorbent for possible use in the expanded bed adsorption of flavonoids from Ginkgo biloba L.

The suitability of the use of macroporous adsorbent Amberlite XAD7HP in expanded bed adsorption processes for the isolation of flavonoids from crude extracts of Ginkgo biloba L. has been assessed. The expansion and hydrodynamic properties of expanded beds were investigated and analyzed. The bed expansion as a function of operational fluid velocity was measured and correlated with the Richardson–Zaki equation. Theoretical predictions of the correlation parameters (the terminal settling velocity u_t and exponent n) were improved by modifying equations in the literature. Residence time distributions (RTDs) were studied using acetone as a tracer. Three measures of liquid phase dispersion (the height equivalent of theoretical plate, Bodenstein number and axial distribution coefficient) were investigated and compared to values previously obtained with commercial EBA adsorbents developed for protein purification. A suitable bed expansion ratio was found to be 1.25 times the settled bed height, which occurred at a corresponding flow velocity of 183 cm/h. For an initial settled bed height of 42 cm, the mean residence time of liquid in the expanded bed was around 28 min. Under these flow conditions, the axial mixing coefficient D_ax was 7.54 × 10⁻⁶ m²/s and the Bodenstein number was 28; the number of theoretical plates (N) was 19 and the height equivalent of a theoretical plate (HETP) was 2.77 cm. Rutin trihydrate was used as a model flavonoid for the characterization of the adsorption properties of Amberlite XAD7HP. Adsorption was observed to reach equilibrium within 3 h with 70% of the adsorption capacity being achieved within 30 min. The estimated maximum equilibrium adsorption capacity for rutin was estimated to be 43.0 mg/(g resin) when the results were fitted to Langmuir isotherms. The adsorption performance was not seriously impaired by the physical presence of G. biloba leaf powders. Assessment of the kinetics of the adsorption of rutin revealed that the rate constant for adsorption was only reduced by 15% in the presence of leaf powders at a concentration of 50 mg/mL. The results demonstrated that Amberlite XAD7HP should be suitable for expanded bed adsorption of flavonoids from crude extracts of G. biloba L.