Analytica Chimica Acta 727 (2012) 1–7 Contents lists available at SciVerse ScienceDirect Analytica Chimica Acta journal homepage: www.elsevier.com/locate/aca Automated capillary electrophoresis with on-line preconcentration by solid phase extraction using a sequential injection manifold and contactless conductivity detection Thanh Duc Mai a,b , Benjamin Bomastyk a , Hong Anh Duong b , Hung Viet Pham b,∗∗ , Peter C Hauser a,∗ a b University of Basel, Department of Chemistry, Spitalstrasse 51, 4056 Basel, Switzerland Centre for Environmental Technology and Sustainable Development (CETASD), Hanoi University of Science, Nguyen Trai Street 334, Hanoi, Viet Nam a r t i c l e i n f o Article history: Received February 2012 Received in revised form 15 March 2012 Accepted 22 March 2012 Available online April 2012 Keywords: Solid phase extraction Capillary electrophoresis Sequential injection analysis Capacitively coupled contactless conductivity detection Drug residues a b s t r a c t An extension of a capillary electrophoresis instrument coupled to a sequential injection analysis manifold was developed for automated measurements with on-line solid-phase extraction preconcentration An in-house built capacitively coupled contactless conductivity detector was employed for sensitive detection with narrow capillaries of 25 m internal diameter The system was assembled into standardized 19 in frames and racks for easy transport and mobile deployment The system can be left running unattendedly without manual intervention with good operation stability To demonstrate the application of the system, a method for the determination of four drugs, namely ibuprofen, diclofenac, naproxen and bezafibrate, was developed with enrichment factors of up to several hundreds Detection of the drug residues down to the nM-scale was found possible and the method was found suitable for the detection of ibuprofen in the waste water of a hospital in Hanoi © 2012 Elsevier B.V All rights reserved Introduction In capillary electrophoresis (CE), detection sensitivities are generally not as good as those in liquid chromatography, mainly due to the by necessity small injection volumes in the nano-liter range To improve the sensitivity of CE several on- and off-line preconcentration strategies have been applied, of which the most commonly employed are electrokinetic techniques (isotachophoresis, stacking, sweeping and dynamic pH junction methods) and solid-phase extraction (SPE) methods For an overview over the two approaches see for example these reviews [1–8] The electrophoretic methods, although inherently being on-line and having the general advantage of relative simplicity, have limitations such as restriction of the primary sample volume, undesired discrimination between analytes of different electrophoretic mobilities, the need of a low background conductivity and the possible requirement of a preceding matrix clean-up step for complex samples The SPE-methods, on the other hand, allow the stripping of large sample volumes, much exceeding the internal volume of the entire separation capillary, and thus can achieve high preconcentration factors These ∗ Corresponding author Tel.: +41 61 267 1003; fax: +41 61 267 1013 ∗∗ Corresponding author Tel.: +84 3858 7964; fax: +84 3858 8152 E-mail addresses: vietph@hn.vnn.vn (H.V Pham), Peter.Hauser@unibas.ch (P.C Hauser) 0003-2670/$ – see front matter © 2012 Elsevier B.V All rights reserved http://dx.doi.org/10.1016/j.aca.2012.03.035 techniques also efficiently remove components not retained on the trapping material, which is useful in the analysis of samples with complex matrices So far, most of the coupled SPE–CE systems reported have been based on liquid delivery with a chromatography pump or a flow injection manifold coupled, often via a robotic arm, to a commercial electrophoresis instrument employing detection based on UV-absorption, laser induced fluorescence or mass-spectrometry [4–9] These setups are thus relatively complex and expensive and not suited for on-site deployment in automated monitoring tasks An attractive alternative approach in the construction of a CEinstrument is the use of a simple sequential-injection analysis (SIA) setup based on a syringe pump and a multi-position valve The SI-manifold can be employed as an alternative means for sample injection and for flushing of the separation capillary and applications of the combination are summarized in [10] Recently Mai et al [11] used such a SIA–CE system for an unattended on-site and on-line monitoring application and also for pressure-assisted CE [12,13] However, to our knowledge, the exploration of the potential of a coupled SI-manifold for extended sample pretreatment prior to analysis by CE has only been reported once Horstkotte et al [14] reported a multi-syringe SPE-CE setup for the determination of nitrophenols by UV-detection A further simplification is possible by employing capacitively coupled contactless conductivity detection (C4 D) This is based on a simple measuring cell consisting of a pair of short tubular T.D Mai et al / Analytica Chimica Acta 727 (2012) 1–7 electrodes placed on the outside of the separation capillary; C4 D is fully electronic and less demanding in construction and power consumption than the common optical detection methods employing UV-radiation Discussions of applications of C4 D for CE can be found in recent reviews [15–19] and fundamental details are given in [20–24] and earlier work cited therein The combination of SPE with CE–C4 D has been reported by Ding and Rogers [25] for the determination of haloacetic acids in swimming pool water, but the preconcentration was carried out off-line in a manual fashion Herein, the implementation of automated SPE on a SIA–CE–C4 D system is described Experimental 2.1 Chemicals and materials All chemicals were of analytical or reagent grade and purchased from Fluka (Buchs, Switzerland) or Merck (Darmstadt, Germany) Stock solutions of ibuprofen, diclofenac, naproxen and bezafibrate in the form of sodium salts (1 mmol L−1 ) were used for the daily preparation of the standards Before use, the capillary was preconditioned with M NaOH for 10 and deionized water for 10 prior to being flushed with the BGE solution at the appropriate pH (for h) Deionized water purified using a system from Millipore (Bedford, MA, USA) was used for the preparation of all solutions The water samples were filtered with 0.02 m PTFE membrane filters (Chromafil O-20/15 MS, Macherey-Nagel, Oensingen, Switzerland), spiked with the selected drug residues as needed, and ultra-sonicated for for degassing 2.2 Instrumentation A dual polarity high voltage power supply (Spellman CZE2000, Pulborough, UK) with ±30 kV maximum output voltage and polyimide coated fused silica capillaries of 365 m OD and 25 m ID (from Polymicro, Phoenix, AZ, USA) were used for all experiments Detection was carried out with a C4 D-system built in-house; details can be found elsewhere [26] An e-corder 201 data acquisition system (eDAQ, Denistone East, NSW, Australia) was used for recording the detector signals The SIA section consisted of a syringe pump (Cavro XLP 6000) fitted with a mL syringe and a 9-port channel selection valve (Cavro Smart Valve) (both purchased from Tecan, Crailsheim, Germany) The isolation and 3-gate valves used were obtained from NResearch (HP225T021 and HP225T031, Gümligen, Switzerland) Commercial SPE cartridges (52602-U, Supelco, Buchs, Switzerland) containing 100 mg of packing material (octadecyl bonded silica particles of 50 m diameter) were fitted into the system with the help of two tubing adapters (57020-U, Supelco) The programming package LabVIEW (version 8.0 for Windows XP, from National Instruments, Austin, TX, USA) was used to write the control code Further details on the instrumentation can be found in [11] Results and discussion 3.1 System design and operation A simplified diagram of the instrument is given in Fig Several extensions and modifications have been made to the earlier SIA–CE–C4 D design [11] in order to incorporate the preconcentration procedure into the fully automated operation For electrophoretic separation without preconcentration, the previously reported system [11] relies on a combination of a stepper motor-driven 2-way syringe and a multi-port selector valve for delivery of solutions, and on a SI–CE interface as well as blocking valves for hydrodynamic injection and flushing of the capillary For operation with SPE preconcentration, the essential change is the inclusion of a second holding coil (HC2) between the multi-port valve and the interface The two holding coils play different roles in the task of liquid handling The conventional holding coil (HC1), situated between the pump and the multi-selector valve, is utilized for aspiration of sample and standards (for separation without preconcentration) or eluent (for elution from the trap prior to separation) while the other coil (HC2) serves as a reservoir to hold the solution following elution from the cartridge before it is pumped to the CE-interface for hydrodynamic injection A Y-shape tubing coupler is employed to divert the fluid to HC2 either from the multiselector valve (for non-preconcentration operations) or from the cartridge (for the elution step in the procencentration procedure) The employment of a 3-gate valve positioned after the cartridge allows the passing of solution either to waste (during loading of sample, flushing and regeneration of the cartridge) or to HC2 during elution In order to allow large sample volumes to be passed through the trapping cartridge repetitive loading was employed In this mode, the entire syringe is filled with sample and then emptied through the preconcentration column The syringe is then filled again with sample for the subsequent loading These steps are repeated until the desired sample volume (typically much larger than the fixed volume of the syringe) is completely passed through the cartridge With this setup there is no upper limit for the loading volume The sample also needs to be acidified before loading onto the trap In order to achieve complete mixing with the acid, which would be difficult to assure in this approach if aspirated consecutively, the auxiliary was merged in from a separate stream This may be accomplished using a separate syringe pump [14], but a more simple approach was used here which is based on a split inlet to allow simultaneous aspiration of two separate solutions A graduated needle valve was employed to adjust the mixing ratio to the desired value All electronic as well as the fluidic parts were assembled into a standard 19 in rack The electronic parts were arranged in two chassis One of these holds the power supplies with the different requisite DC-voltages for the syringe pumps, the valves and the high voltage unit, and the second the control and interface electronics for the different modules Switches, controls as well as displays for the voltage and current of the high voltage unit are accessible on the front panels These rack-mounted cases can be easily withdrawn for modifications All fluidic components, including the pump, valves, holding coils, connecting tubing and liquid containers, are fixed onto a panel situated above the two electronic rack inserts The details of a typical sequence of operations are given in Table All steps are fully automated and can be performed unattended as controlled by the software program running on the personal computer Note that the movement of the syringe pump is determined by setting the desired volume to be aspirated or dispensed and the flow rate The protocol starts with rinsing of the cartridge (step 1) prior to repeated loading of sample onto the preconcentration column (step 2) The loaded cartridge is then rinsed again with water (step 3) In preparation for the electrophoretic separation a flushing of the SI–CE interface (step 4) is then carried out by pumping the buffer through the interface on simultaneous opening of both stop-valves (designated as V1 and V2 in Fig 1) The capillary itself is then flushed (step 5) by slowly advancing the syringe pump while both V1 and V2 are closed in order to push all of the dispensed fluid through the separation tubing Elution of the trapped analyte is then implemented (step 6) by passing eluent through the cartridge while the switching valve is at position Once the eluted solution has been collected in the holding coil HC2, hydrodynamic injection takes place (step 7), followed by electrophoretic separation (steps 8, 9) The hydrodynamic T.D Mai et al / Analytica Chimica Acta 727 (2012) 1–7 Capillary: 25 µm, 60 cm C 4D Selection Valve Safety switch DI Water Eluent Regenerating solution HCl 0.1 M Pt HV +/- Y- Coupler Electrolyte Solution 5000 µL Syringe Needle-Valve Pt HC1 (500 µL) HV-interface W M NaOH Cartridge (50 µm C18 particles) Grounded HC2 interface (500 µL) W V1 V2 Safety case W W Switching Valve W HCl 0.1 M Sample Needle-Valve Fig Schematic drawing of the SIA–CE–C4 D-system for automated electrophoretic separation with on-line SPE preconcentration C4 D: contactless conductivity detector; HV: high-voltage power supply; Pt: platinum electrode; W: waste; V1, V2: stop valves; HC: holding coil split-injection is carried out by pumping a sample plug past the capillary inlet in the SIA–CE interface while pressurizing the manifold by closing only V2 The splitting ratio is very large, as only a small volume in the nanoliter range can be injected into the capillary, but not known as the setup is done empirically by changing the positioning of the needle valve until a good compromise between sensitivity and peak resolution is obtained More details on this procedure can be found in a previous publication [11] Separation is implemented by application of the high voltage of appropriate polarity from the detector end, with the injection end being grounded Rinsing of the manifolds, interface and capillary with buffer (steps 10–12) is then carried out on completion of the electrophoretic separation Finally, the cartridge is regenerated and flushed thoroughly (steps 13–16) to ready it for the next preconcentration operation Note that after each solution delivery, the syringe is also rinsed with deionized water (steps 3, 14, 16, requiring 36 s) before continuing with delivery of another solution Separations with or without preconcentration can be selected from the computer For separation without preconcentration, the sample is transferred directly to the SI–CE interface instead of being loaded onto the cartridge and steps 1, 2, 3, 4, 6, 13, 14, 15 and 16 are omitted from the protocol 3.2 Determination of pharmaceuticals In order to demonstrate the system a method for the determination of drug residues was implemented Many pharmaceuticals are classified as environmental contaminants due to their low Table Typical operation sequence with preconcentration Step Description Position of selection valve Volume dispensed (L) Flow rate (L s−1 ) Position of V1 Position of V2 Position of switching valve 1a 1b 2a* 2b* 3a 3b 4a 4b 5a 5b 5c 6a 6b 9** 10 11 12 13a 13b 14a 14b 15a 15b 16a 16b Pick up of water Flushing of the cartridge with water Pick up of sample Dispensing of sample to cartridge Pick up of water Flushing of the cartridge with water Buffer aspiration Flushing of syringe and HC1 with buffer Buffer aspiration Flushing of the SI–CE interface Flushing of the capillary Pick up of eluent Dispensing of eluent through cartridge to HC2 Sample injection Flushing of the SI–CE interface Electrophoretic Separation Flushing of the SI–CE interface Flushing of the capillary Empty syringe Pick up of regenerating solution Dispensing of regenerating solution to cartridge Pick up of water Flushing of the cartridge with water Pick up of 0.1 M HCl solution Dispensing of HCl to cartridge Pick up of water Flushing of the cartridge with water 8 – – 7 7 7 8 8 5000 5000 5000 5000 5000 5000 3000 3000 3000 1000 25 500 500 500 500 500 25 – 5000 5000 5000 5000 500 500 5000 5000 140 140 140 140 140 140 167 167 167 167 140 25 167 167 – 167 167 167 140 140 140 140 140 140 140 140 Open Open Open Open Open Open Open Open Open Open Closed Open Open Open Open Closed Open Closed Open Open Open Open Open Open Open Open Open Open Open Open Open Open Open Open Open Open Open Closed Open Open Closed Open Closed Open Closed Open Open Open Open Open Open Open Open Open 1 1 1 1 1 1 1 1 1 1 1 1 1 * ** Steps 2a and 2b are repeated n times until the desired sample volume has been loaded The high voltage is automatically turned on for step 4 T.D Mai et al / Analytica Chimica Acta 727 (2012) 1–7 biodegradability, and the fact that their bioactive properties might lead to adverse effect on human beings and ecosystems [27–30] The determination of drug residues in water with CE using UV or mass-spectrometric detection has been reported repeatedly with and without using electrokinetic stacking preconcentration, see for example [31–34] Quek et al [35] also reported the determination of 13 pharmaceuticals deemed potential pollutants by CE–C4 D No preconcentration methods were employed and detection limits of typically M were obtained In this study, four widely used pharmaceuticals, namely ibuprofen, diclofenac, naproxen and bezafibrate, were selected as exemplary analytes 0.2 V (a) (b) 3.2.1 Separation buffer For the electrophoretic separation the background electrolyte (BGE) based on the one reported by Quek et al [35] consisting of a Tris/lactic acid buffer at pH and containing hydroxypropyl-cyclodextrin (HP--CD) was employed The pH value assures deprotonation and thus anionic ionization of the compounds Cyclodextrin HP--CD was added into the BGE as a complexing reagent for separation of ibuprofen and diclofenac It does not possess a charge, and thus does not contribute to the conductivity of the BGE As discussed below, the release of the analytes from the trapping material requires an eluent with a pH which leads to ionization as well as the inclusion of a substantial fraction of an organic solvent For this reason the injection of standards in a Tris/lactic acid buffer with an acetonitrile content of 37.5% (v/v) was tested As shown in electropherogram (b) of Fig 2, broad peaks with low sensitivity and poor resolution were obtained, whereas the injection of a purely aqueous solution (no buffer) of the four species led to the good separation of electropherogram (a) It was assumed that the sharper peaks of this top trace were due to a transient stacking effect due to the low conductivity of the sample plug In order to induce a similar performance with the background of the given eluent the composition of the BGE was adjusted, i.e the conductivity of the latter was increased by increasing the buffer concentration As seen in electropherogram (c) of Fig 2, indeed a good performance could also be obtained with these conditions Buffers of even higher concentrations were also tried, but led to noisy baselines, presumably due to Joule heating Thus there is a limit to this approach The performance data for the direct determination without preconcentration of the four analgesics on the CE system are given in Table The detection limits achieved are in the range from of 0.8 to 1.5 M Calibrations were carried out for the range from 2.5 M to 100 M (6 concentrations) and linearity was obtained 3.2.2 Sample loading The four investigated drugs are carboxylates, and thus are negatively charged in water of neutral pH For effective adsorption onto C18 sorbent particles, the sample needs to be acidified in order to render the compounds in their neutral form The addition of 0.1 M HCl was found to be suitable for sample acidification since a minor fraction (less than 1%, v/v) of this strong inorganic acid solution is sufficient to adjust the pH of the samples below the pKa of these carboxylates (50%, v/v) for good elution Tetrahydrofuran, which possesses the highest elution strength among all commonly used organic solvents miscible with water, though offering very efficient desorption even at low concentrations (less than 25%, v/v), was found to disturb the resolution of the peaks of naproxen and ibuprofen in the electrophoretic separation Acetonitrile, with intermediate elution strength, was therefore chosen as the organic additive The effect of the concentration of acetonitrile on elution efficiency is shown in Fig The measurements were repeated twice, and the results were within Peak area (V·s) Fig Effect of acetonitrile concentration on efficiency of elution from the trap The eluent was prepared by addition of acetonitrile to an aqueous solution of mM Tris and mM lactic acid Peak areas were obtained from electropherograms with the following CE conditions: E = 400 V cm−1 , feedback resistance for C4 D = M , capillary of 25 m inner diameter and 41 cm effective length, BGE composed of 36 mM Tris/5 mM lactic acid/1 mM HP--CD, hydrodynamic split-injection with a dispensed sample volume of 0.5 mL and a setting of 0.11 on the micrometer screw of the needle valve 0.5 20 30 40 50 (B) 0.4 Diclofenac 0.3 0.2 Naproxen 0.1 0.0 10 20 30 40 50 Run Number Fig Reproducibility of peak areas for the preconcentrated pharmaceuticals during continuous operation of 10 h per day for successive days For each run 15 mL solution of standard mixtures of 0.5 M in deionized water was loaded onto a C18 cartridge followed by an elution with 0.5 mL eluent (9 mM Tris/5 mM lactic acid (62.5%, v/v) + CH3 CN (37.5%, v/v)) Peak areas were obtained for a preconcentration factor of 30 Other CE conditions as for Fig ±1.5% The four retained drugs are eluted with different efficiencies, with diclofenac being the compound which is most easily desorbed Complete elution occurs for acetonitrile concentrations of 37.5% (v/v) or higher in an eluent containing mM Tris and mM lactic acid Note that no significant elution was observed when only acetonitrile in water was employed as the eluent even at acetonitrile concentrations higher than 50% (v/v) Following the analyte elution the cartridge was regenerated by first passing a relatively large volume of mL of the mM Tris/5 mM lactic acid buffer mixed with acetonitrile (50%, v/v) in order to assure efficient removal of any organic species that may still be retained on the cartridge after preconcentration The trap was subsequently rinsed with 0.5 mL of 0.1 M HCl to dissolve any precipitates that may have formed on the surface of the sorbent particles during the preceding operations at higher pH Finally, the cartridge was rinsed with deionized water (5 mL) before the next trapping sequence 3.3 Performance and sample analysis In order to evaluate the potential for unattended operation, the system was set up for a test run for a period of 10 h per day over continuous days, in which repeated preconcentrations from 15 mL of sample solution with an enrichment factor of 30 (as follows from the volume ratios for efficient trapping and elution) and CE measurements of the pharmaceuticals (0.5 M prepared in T.D Mai et al / Analytica Chimica Acta 727 (2012) 1–7 0.30 100 (A) 0.25 80 Recovery (%) Peak Area (V·s) Ibuprofen 0.20 0.15 0.10 Bezafibrate 0.00 Diclofenac Ibuprofen Naproxen Bezafibrate 40 80 120 160 200 Loading volume (mL) (B) Fig Recoveries of the four pharmaceuticals spiked in Rhein river water at different preconcentration factors The spiked concentrations were varied from 25 nM to 250 nM depending on the loading volume in order to keep the concentrations after enrichments fixed at 10 M Other conditions as for Fig 0.25 Peak Area (V·s) 40 20 0.05 0.30 60 Diclofenac 0.20 0.15 0.10 Naproxen 0.05 0.00 Run Number Fig Reproducibility of peak areas for the preconcentration and CE determination of the pharmaceuticals spiked in tap water For each operation, 375 mL solution of standard mixtures of 0.01 M prepared in filtered tap water was loaded onto a C18 cartridge followed by an elution with 0.5 mL eluent (9 mM of Tris/5 mM lactic acid (62.5%, v/v) + CH3 CN (37.5%, v/v)) Peak areas were obtained for an enrichment factor of 750 Other CE conditions as for Fig deionized water) were carried out The preconcentration process for this enrichment factor requires min, whereas it takes about 25 for the entire protocol including preconcentration, analysis and all flushing operations to be completed The results for peak areas are shown in Fig The maximum deviation is about ±8%, which is deemed acceptable considering that this deviation is due to the accumulation of errors of all operations, i.e sample loading, elution, injection and separation Without the preconcentration procedure, deviations of up to ±4% were observed for automated CE separations [11] No bias of peak areas was recorded after more than 50 continuous runs in days, which demonstrates the suitability of the system for unattended operation The operational performance was further evaluated by carrying out preconcentrations under more severe conditions, in which solutions of the pharmaceuticals were prepared directly in a tap water matrix instead of deionized water The loading of a large volume of 375 mL of tap water spiked with pharmaceuticals (10 nM) and desorption with 0.5 mL eluent was repeated several times with the same cartridge The enrichment factor in this case is 750, which is very large for SPE preconcentration The results for peak areas are shown in Fig The measurements were repeated twice and the results could be reproduced within ±4% It is apparent that after successive preconcentrations, the peak areas of all pharmaceuticals decreased significantly This is thought to be due to the sorbent material becoming permanently saturated with strongly bound species present in the matrix in minor quantities which are not released in the elution step Thus, for these conditions a total loading volume of 1000 mL should not be exceeded before replacing the trap Further tests were carried out by spiking water from the river Rhine (Basel, Switzerland) with the pharmaceuticals as it was expected that this effect would be more pronounced for the more complex matrix This was indeed the case The results are given in Fig 6, where the recovery in dependence of the loading volume is shown The measurements were repeated twice and the results could be reproduced within ±4.5% For equivalent comparison of recovery, the concentration in the donor solution was varied (25–250 nM) according to the volume passed through the trap (from 200 mL to 20 mL) so that the nominal concentration in the eluting solution for complete extraction was fixed at 10 M The data of Fig demonstrates that a significant reduction in recovery for river water spiked with the pharmaceuticals occurred when a sample volume of 50 mL was exceeded This indicates a general limitation of the method which has to be carefully assessed for a task at hand The system employing the optimized SPE–CE–C4 D conditions was then used to analyze a water sample taken from the outlet of a wastewater treatment plan of a hospital in Hanoi, Vietnam Electropherograms for the sample with and without enrichment are shown in Fig Ibuprofen, which is barely discerned without preconcentration, can be clearly observed after enrichment Some other minor peaks also appear in the electropherogram for the enrichment factor of 40, but no effort was made to identify these species 200 mV Without preconcentration Enrichment factor = 20 Enrichment factor = 40 260 280 300 320 340 360 380 400 Migration time (s) Fig Analysis of a water sample taken from a wastewater treatment plan of a hospital in Hanoi, Vietnam Other conditions as for Fig T.D Mai et al / Analytica Chimica Acta 727 (2012) 1–7 Conclusions The automated SPE technique could be readily implemented on the SIA–CE–C4 D system and provides a robust and straightforward means to extend the detection limit of CE–C4 D Enrichment factors of up to 750 could be demonstrated For the pharmaceuticals tested a lower limit of quantification (LOQ) of about M is possible by direct CE–C4 D analysis without preconcentration Consequently, with preconcentration LOQs in the low nM-range are possible if the sample volume is sufficient In practice the sample matrix may also impose a limit to the volume of sample which can be passed through the trap, and thus the highest preconcentration factor that can be achieved Due to the employment of electrophoretic separation and conductivity detection the construction of the entire instrument is relatively simple The components for the standardized 19-in system are widely available commercially, the fluidic section is based on standard parts, and only a few special components had to be manufactured for purpose The compact all-in-one design of the overall instrument allows easy transport and deployment for automated on-site monitoring applications is readily possible 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volume of 375 mL of tap water spiked with pharmaceuticals (10 nM) and desorption with. .. conditions was then used to analyze a water sample taken from the outlet of a wastewater treatment plan of a hospital in Hanoi, Vietnam Electropherograms for the sample with and without enrichment are... was carried out off-line in a manual fashion Herein, the implementation of automated SPE on a SIA–CE–C4 D system is described Experimental 2.1 Chemicals and materials All chemicals were of analytical