Disposable electrochemical flow cells for catalytic adsorptive stripping voltammetry -LRB- CAdSV -RRB- at a bismuth film electrode -LRB- BiFE -RRB- Catalytic adsorptive stripping voltammetry -LRB- CAdSV -RRB- has been demonstrated at a bismuth film electrode -LRB- BiFE -RRB- in an injection-moulded electrochemical micro-flow cell . The polystyrene three-electrode flow cell was fabricated with electrodes moulded from a conducting grade of polystyrene containing 40 % carbon fibre , one of which was precoated with Ag to enable its use as an on-chip Ag/AgCl reference electrode . CAdSV of Co -LRB- II -RRB- and Ni -LRB- II -RRB- in the presence of dimethylglyoxime -LRB- DMG -RRB- with nitrite employed as the catalyst was performed in order to assess the performance of the flow cell with an in-line plated BiFE . The injection-moulded electrodes were found to be suitable substrates for the formation of BiFEs . Key parameters such as the plating solution matrix , plating flow rate , analysis flow rate , solution composition and square-wave parameters have been characterised and optimal conditions selected for successful and rapid analysis of Co -LRB- II -RRB- and Ni -LRB- II -RRB- at the ppb level . The analytical response was linear over the range 1 to 20 ppb and deoxygenation of the sample solution was not required . The successful coupling of a microfluidic flow cell with a BiFE , thereby forming a `` mercury-free '' AdSV flow analysis sensor , shows promise for industrial and in-the-field applications where inexpensive , compact , and robust instrumentation capable of low-volume analysis is required . Introduction Mercury electrodes , normally in the form of mercury films for flow systems , have been widely used for performing stripping voltammetry as a result of their high sensitivity and reproducibility , but , due to increasing concern and legislation regarding the toxicity of mercury , alternative working electrode -LRB- WE -RRB- materials have been investigated . Metals such as Au -LSB- 1 , 2 -RSB- , Ag -LSB- 2 , 3 -RSB- , Ir -LSB- 4 -RSB- and even W -LSB- 5 -RSB- as well as bare and coated carbon electrodes -LSB- 6 , 7 -RSB- have all been investigated for stripping analysis of metals . In 2000 , bismuth-film electrodes -LSB- 8 -RSB- were mooted as a replacement for mercury-film electrodes , as bismuth exhibits negligible toxicity and is more environmentally friendly in comparison . Mercury use is increasingly regulated , motivating the search for mercury-free methods . For example , Sweden banned the sale of switches , apparatuses and instruments containing mercury in 1993 -LRB- Ordinance SFS 1991:1290 -RRB- and instructed the Swedish Chemicals Inspectorate -LRB- KemI -RRB- to look into a total ban on the handling of mercury in Sweden , including chemicals used for analysis . The Inspectorate proposed that mercury for chemical analysis should be exempted from the general ban until 31st December 2008 in order to enable the development and implementation of mercury-free analysis methods -LSB- 9 -RSB- . In 2005 the EU commission proposed a ban on mercury exports by 2011 as part of a strategy to reduce mercury emissions and protect against exposure -LSB- 10 -RSB- , and recently MEPs voted to bring this date forward to 2010 -LSB- 11 -RSB- . A variety of substrates have been used for bismuth films . While platinum -LSB- 12 -- 14 -RSB- and gold -LSB- 13 -RSB- have both been investigated , the majority of films have been deposited on carbon substrates . Most studies have used glassy carbon -LSB- 8 , 13 , 15 -- 32 -RSB- , though wax-impregnated graphite -LSB- 22 , 24 -RSB- , pencil-lead graphite -LSB- 33 -RSB- , carbon paste -LSB- 13 , 34 -- 36 -RSB- , screen-printed carbon inks -LSB- 21 , 37 , 38 -RSB- , carbon fibres -LSB- 8 , 13 , 14 -RSB- and boron-doped diamond -LSB- 39 -RSB- have all been employed as substrates . The use of Bi2O3 containing pastes -LSB- 34 , 40 -RSB- and a bismuth bulk electrode -LSB- 41 -RSB- have also been reported . The use of BiFE has recently been reviewed -LSB- 42 -RSB- . Bismuth-film electrodes have mainly been applied to anodic stripping voltammetry -LRB- ASV -RRB- and adsorptive stripping voltammetry -LRB- AdSV -RRB- techniques for metal ion analysis , although cathodic detection of nitrophenols -LSB- 17 , 27 , 41 -RSB- and amperometric detection of glucose -LSB- 12 -RSB- have also been reported . Cd -LSB- 28 -RSB- , Pb -LSB- 18 , 37 -RSB- , Zn -LSB- 31 , 32 -RSB- and mixtures of all three ions -LSB- 8 , 20 , 22 , 23 , 33 , 36 , 41 -RSB- have been the most extensively studied by ASV . Tl , Cu -LSB- 16 -RSB- , In -LSB- 16 , 43 -RSB- , Mn -LSB- 39 -RSB- and Sn -LSB- 44 -RSB- have also been analysed . Fewer reports have been made employing AdSV , though Ni -LSB- 15 -RSB- and Co -LSB- 28 -- 30 -RSB- and Ni with Co -LSB- 24 , 26 -RSB- have been examined using dimethylglyoxime -LRB- DMG -RRB- . Cr -LSB- 21 , 25 , 45 -RSB- , U -LSB- 46 , 47 -RSB- , Al -LSB- 48 -RSB- , V -LSB- 49 -RSB- and Mo -LSB- 50 -RSB- analysis by AdSV on bismuth-film electrodes has also been performed . Stirred solutions have normally been used , though Economou et al. have reported the use of a rotating disc electrode -LSB- 22 -- 25 -RSB- and a Nafion-coated BiFE in a thin layer flow cell -LRB- 0.5 mm spacer -RRB- for sequential injection-ASV of metals -LSB- 51 , 52 -RSB- . Hutton et al. generated an in-situ BiFE in a thin layer -LRB- 0.3 mm spacer -RRB- flow cell -LRB- Bioanalytical Systems Inc. , West Lafayette , IN , USA -RRB- for flow injection analysis of nitrophenols with cathodic amperometric detection -LSB- 27 -RSB- . Adsorptive stripping voltammetry of DMG-complexed Co and Ni has found widespread use in trace metal analysis . The enhancement of the Co-DMG response using nitrite , i.e. catalytic adsorptive stripping voltammetry -LRB- CAdSV -RRB- , has also been the subject of several publications , for example van den Berg et al. -LSB- 53 , 54 -RSB- and the review of catalytic systems by Bobrowski and Zarębski -LSB- 55 -RSB- . CAdSV of Co on a BiFE was reported by Krolicka et al. -LSB- 29 , 30 -RSB- using an ex situ plated film on glassy carbon . Most BiFEs have been plated from acetic buffers , although Krolicka et al. used a plating solution of Bi -LRB- NO3 -RRB- 3 -LRB- 0.02 M -RRB- , LiBr -LRB- 0.5 M -RRB- , and HCl -LRB- 1 M -RRB- with a plating potential of − 0.25 V for 45 s -LSB- addition of bromide and strong acid enabled a highly concentrated Bi -LRB- III -RRB- plating solution to be produced -RSB- . Stripping was typically carried out in 0.1 M ammonia buffer -LRB- pH 9.2 -RRB- containing DMG -LRB- 1 × 10 − 4 M -RRB- and NaNO2 -LRB- 0.5 M -RRB- . Voltammograms were recorded in the differential pulse mode and a 15-fold enhancement of the Co-DMG stripping signal was reported . Recently the authors reported the use of 40 % carbon fibre-filled nylon 6/6 injection-moulded electrodes as working electrodes for the determination of Cu -LRB- II -RRB- by CV and ASV techniques -LSB- 56 -RSB- . In this work , the electrodes were moulded from a 40 % carbon fibre-filled polystyrene , which was also found to be suitable for performing electrochemical detection . This paper presents the first report of the use of CAdSV at an in-line plated BiFE in an integrated three-electrode injection-moulded plastic micro flow cell . Experimental Instrumentation Voltammetric measurements were performed using an Autolab electrochemical system -LRB- PSTAT 10 , Eco Chemie B.V. , Utrecht , The Netherlands -RRB- controlled with GPES software -LRB- v4 .6 , Eco Chemie -RRB- running on a PC . Rotating disc electrode experiments The rotating disc electrode -LRB- RDE -RRB- was a glassy carbon electrode -LRB- 6.1204.000 , Metrohm , Buckingham , UK -RRB- 3 mm in diameter controlled with a variable speed -LRB- 500 -- 3000 rpm -RRB- electrode rotator -LRB- Part No. 628-10 , Metrohm -RRB- . The disc electrode was polished with an alumina slurry -LRB- 0.3 μm Al2O3 , Metrohm -RRB- before use . A standard -LRB- 12 mm in diameter -RRB- glass double junction Ag/AgCl/KCl -LRB- 3M -RRB- reference electrode was used -LRB- CR4/DJ/AG , Thermo Electron , Auchtermuchty , Fife , UK -RRB- with a Pt wire auxiliary 0.5 mm in diameter -LRB- 99.9 % , Aldrich , Gillingham , UK -RRB- . The electrodes were mounted in a small-volume 50-ml beaker used as an electrochemical cell . Flow cell experiments A gravity-fed flow system was used comprising a reservoir formed from a disposable syringe body -LRB- 20 ml -RRB- , and narrow-bore PTFE tubing -LRB- 0.032 in . i.d. , Lee Products , Gerrards Cross , UK -RRB- . The flow was controlled with a manual two-way valve -LRB- Part No. 2420 , Omnifit , Cambridge , UK -RRB- . Barbed -LRB- 1/16 ″ -RRB- polypropylene female Luer fittings -LRB- Cole-Parmer , Hanwell , London , UK -RRB- enabled connection of the flow system to the injection-moulded flow cell . Electrical connection to the Autolab electrochemical system was made using crocodile clips . Chip fabrication The injection-moulded flow cells were produced in-house . Figure 1 depicts the fabricated device showing the three integrated electrodes -LRB- WE , reference and auxiliary -RRB- . Solid models created using AutoCAD -LRB- Mechanical Desktop 2004 , Autodesk , San Jose , CA , USA -RRB- were converted into macro commands for a CNC milling machine -LRB- Datron CAT3D M6 , Datron Technology , Milton Keynes , UK -RRB- to produce the milled aluminium moulds for the injection-moulding machines -LRB- Babyplast 6/6 or 6/10 , Cronoplast SL , Barcelona , Spain -RRB- . A four-stage process was required to manufacture a ready-to-use device . The polymer electrodes were injection moulded -LRB- moulding temperature of 220 °C -RRB- from a conducting polymer -LRB- 40 % carbon-fibre-filled high-impact polystyrene -LRB- HIPS -RRB- , RTP 487 , RTP Company -LRB- UK -RRB- Plastics Ltd. , Bury , UK -RRB- . The reference electrodes were coated with Ag -LRB- Ag evaporation slug , 99.99 % , Aldrich -RRB- using an e-beam evaporator -LRB- Auto500 with EB3 e-beam unit , Edwards High Vacuum International , Crawley , West Sussex , UK -RRB- to a thickness of 125 nm . The WE , reference and auxiliary electrodes were then incorporated into the flow cell by an overmoulding procedure . The mould cavity for the flow cell design comprises a base plate into which the preformed electrodes are inserted in the appropriate recesses and a top plate with a relief pattern that forms the flow channel and integrated fluidic connectors . The flow cell was moulded -LRB- moulding temperature of 220 °C -RRB- from crystal polystyrene -LRB- Northern Industrial Plastics Ltd. . Chadderton , UK -RRB- . Using a 500-W ultrasonicwelder -LRB- Mini delta 9500 , FFR Ultrasonics Ltd. , Queniborough , Leicestershire , UK -RRB- , with a weld time of 1 s and a hold time of 2 s , the top and base plates were sealed together to form a complete and ready-to-use device . The flow channel dimensions were 1 mm wide , 1 mm deep and 30 mm long . The working and auxiliary electrodes were 5 mm wide , the reference was 2 mm wide and the spacing between them was 2 mm . Fig. 1The injection-moulded electrochemical micro flow cell . Here -LRB- A -RRB- is the Ag-coated electrode , which forms the reference -LRB- Ag/AgCl -RRB- element ; -LRB- B -RRB- is the working electrode , on which the bismuth film electrode is formed ; -LRB- C -RRB- is the auxiliary electrode ; -LRB- D -RRB- is the device inlet and -LRB- E -RRB- is the device outlet . The flow channel is 1 mm × 1 mm and 30 mm long Reagents and solutions All aqueous solutions were prepared using > 18MΩ water -LRB- Elga Maxima Ultra Pure , Vivendi , High Wycombe , UK -RRB- . The acetate buffer was prepared from acetic acid -LRB- glacial 99.8 % AnalaR grade , BDH , Poole , UK -RRB- and sodium acetate -LRB- 99 + % ACS grade , Aldrich , Gillingham , UK -RRB- . The ammonium buffer was prepared from ammonium chloride -LRB- 99.8 % AnalaR grade , BDH -RRB- and ammonium hydroxide -LRB- 28 -- 30 % NH3 ACS grade , Aldrich -RRB- . Bismuth plating solutions were prepared with bismuth -LRB- III -RRB- nitrate pentahydrate -LRB- 99.999 % , Aldrich , -RRB- either in acetate buffer or with lithium bromide -LRB- 99 + % , Aldrich -RRB- in hydrochloric acid -LRB- 1 M volumetric standard , Riedel-de Haën , Gillingham , UK -RRB- , as described in the `` Procedure '' section . The mercury plating solution was prepared in nitric acid from a 1000 ppm mercury -LRB- II -RRB- chloride -LRB- 98 + % , Aldrich -RRB- stock solution , as described in the `` Procedure '' section . Sample solutions were made from cobalt -LRB- 995 μg/l , Aldrich -RRB- and nickel -LRB- 1 g/l , Fluka , Gillingham , UK -RRB- atomic absorption standards with dimethylglyoxime -LRB- 99 + % , Acros , Loughborough , UK -RRB- and sodium nitrite -LRB- 97 + % , Aldrich -RRB- . The saturated AgCl solution was prepared from silver nitrate -LRB- 99.9 % AnalaR grade , BDH -RRB- in potassium chloride -LRB- 99.5 % AnalaR grade , BDH -RRB- . Nitric acid -LRB- 2 M volumetric standard , Riedel-de Haën -RRB- was used for electrode cleaning , as described in the `` Procedure '' section . Procedure Formation of the Ag/AgCl miniaturised reference electrode Using an external auxiliary platinum wire located in the inlet of the microdevice , the Ag electrode was coated in AgCl -LRB- in saturated KCl solution that was also saturated with AgCl through the addition of a small quantity of AgNO3 -RRB- at 2 V for 3 s. Ex situ electroplating of the BiFE on the rotating disc electrode -LRB- RDE -RRB- Two electroplating solutions were investigated , -LRB- A -RRB- and -LRB- B -RRB- . Acetate buffer Bi -LRB- III -RRB- plating solution . 100 ppm Bi -LRB- III -RRB- in 1 M acetate buffer -LRB- pH 4.5 -RRB- . Plated for 5 mins at − 1.0 V in a quiescent solution.LiBr / HCl Bi -LRB- III -RRB- plating solution . 0.02 M Bi -LRB- III -RRB- in 0.5 M LiBr and 1 M HCl . Plated for 60 s at − 0.28 V in a quiescent solution . The RDE was then carefully rinsed before transferring it to the analysis solution . In-line electroplating of the BiFE on the flow cell working electrode Two electroplating solutions were investigated , -LRB- A -RRB- and -LRB- B -RRB- . Acetate buffer Bi -LRB- III -RRB- plating solution . 100 ppm Bi in 0.1 M acetate buffer -LRB- pH 4.5 -RRB- . Plated for 5 mins at − 1.0 V at a flow rate of 37.1 μl/s . LiBr / HCl Bi -LRB- III -RRB- plating solution . 0.02 M Bi -LRB- III -RRB- in 0.5 M LiBr and 1 M HCl . Plated for 120 s at − 0.28 V at a flow rate of 37.1 μl/s . After formation of the BiFE , the plating flow stream was flushed with water and switched to the analysis stream . Bismuth film cleaning procedure The Bi -LRB- III -RRB- film was cleaned of the remaining adsorbed complexes by holding the electrode at a potential of − 1.3 V for 30 s in 0.1 M HCl . Bismuth film removal Bi -LRB- III -RRB- films were removed by applying a conditioning potential at +0.3 V , − 0.5 V and 0 V for 2 × 50 s in a flowing solution of 0.1 M HNO3 . In-line plating of MFE on the flow cell working electrode The Hg plating solution was 200 ppm in 0.1 M HNO3 . Plated for 120 s at − 1 V at a flow rate of 9 μl/s . After film formation the cell was flushed with water and switched to the analysis stream . Mercury film cleaning procedure The film was cleaned at a potential of − 1.4 V for 10 s in 0.1 M HNO3 . Mercury film removal Films were removed at +0.8 V in a flowing solution of 0.1 M HNO3 for 60 s. Results and discussion The in-line prepared BiFE was assessed for its suitability in performing CAdSV of Co -LRB- II -RRB- and Ni -LRB- II -RRB- . Various parameters , such as flow rates , solution composition , plating parameters and square-wave conditions , were studied to investigate the electrochemical behaviour of the Bi -LRB- III -RRB- films prepared on the injection-moulded carbon fibre electrodes . Investigation of the effect of flow rate during the plating and accumulation stages Using the acetate buffer-based plating solution , i.e. the 100 ppm Bi -LRB- III -RRB- in 0.1 M acetate buffer , the effect of varying the flow rate during BiFE plating procedure was examined at 20 , 25 and 37 μl/s by varying the height of the reservoir in the gravity-fed flow system -LRB- with sample conditions as given in Fig. 2 -RRB- . The maximum response was obtained for the stripping peaks at a flow rate of 37 μl / s and so this rate was subsequently employed in this study . Fig. 2Influence of flow rate during the accumulation stage on the Ni -LRB- II -RRB- and Co -LRB- II -RRB- CAdSV peaks , where the flow rates are : -LRB- a -RRB- 5.8 , -LRB- b -RRB- 15.7 , -LRB- c -RRB- 25.6 , -LRB- d -RRB- 37.1 , -LRB- e -RRB- 42.9 and -LRB- f -RRB- 55.5 μl/s respectively . Carried out with a 10 ppb sample of Co -LRB- II -RRB- and Ni -LRB- II -RRB- in 0.1 M ammonium buffer -LRB- pH 9.2 -RRB- with 0.1 mM DMG and 0.5 M NaNO2 . SW conditions : Conditioning potential − 1.3 V -LRB- 10 s -RRB- , deposition potential − 0.8 V -LRB- 120 s -RRB- , equilibration time 5 s , pulse frequency 15 Hz , pulse amplitude 40 mV and step potential 4 mV . Scan : initial − 0.8 V to end potential − 1.3 V. Data were smoothed with a level-2 Savitzky -- Golay filter The effect of varying the flow rate during the accumulation stage on the peak heights of Co -LRB- II -RRB- and Ni -LRB- II -RRB- was studied over the range of 5.8 -- 55.5 μl/s ; the results are depicted in Fig. 2 . The response for Co -LRB- II -RRB- increased linearly with increasing flow rate with a slope of − 0.036 μAs / μl and an intercept of − 1.26 μA with a correlation coefficient -LRB- R2 = 0.96 -RRB- , whereas that for Ni -LRB- II -RRB- did not show significant variation . A flow rate of 37 μl/s was selected , as this gave an acceptable response without adverse peak broadening and was compatible with the BiFE plating step flow rate , thereby simplifying the instrumentation . Effect of composition of the BiFE plating solution Reported methods for AdSV of Co -LRB- II -RRB- and Ni -LRB- II -RRB- have generally employed acetate buffer -LRB- pH 4.5 -RRB- containing 100 mg/l Bi -LRB- III -RRB- -LSB- 15 , 24 , 26 , 28 -RSB- . However , Królicka et al. -LSB- 29 , 30 -RSB- suggested the use of a bromide-modified plating solution . Hence , both plating solutions were assessed using the RDE and flow cell setups . Figure 3 depicts the differing performances of the two plating solutions . The LiBr-containing plating solution gave the best stripping responses on both the RDE and the flow cell and was therefore used for subsequent studies . Fig. 3Comparison of different Bi -LRB- III -RRB- plating solutions . a Effect of -LRB- a -RRB- 100 ppm Bi -LRB- III -RRB- in acetate buffer -LRB- 1 M , pH 4.5 -RRB- and -LRB- b -RRB- 0.02 M Bi -LRB- III -RRB- in 0.5 M LiBr - and 1M HCl-based preplating solutions on RDE at 500 rpm . b Effect of -LRB- c -RRB- the acetate buffer -LRB- 0.1 M , pH 4.5 -RRB- and -LRB- d -RRB- LiBr/HCl preplating solutions on the flow cell . c Comparision of acetate buffer pH 4.5 Bi -LRB- III -RRB- preplating solution on -LRB- e -RRB- flow cell -LRB- 0.1 M -RRB- and -LRB- f -RRB- RDE -LRB- 1 M -RRB- . d Comparison of LiBr preplating solution on -LRB- g -RRB- flow cell and -LRB- h -RRB- RDE . Carried out with a 10 ppb sample of Co -LRB- II -RRB- and Ni -LRB- II -RRB- in 0.1 M ammonium buffer -LRB- pH 9.2 -RRB- with 0.1 mM DMG and 0.5 M NaNO2 . SW conditions : Conditioning potential − 1.3 V -LRB- 10 s -RRB- , deposition potential − 0.8 V -LRB- 120 s -RRB- , equilibration time 10 s , pulse frequency 25 Hz , pulse amplitude 50 mV and step potential 5 mV -LRB- except trace -LRB- e -RRB- which was at 15 Hz , 40 and 4 mV respectively -RRB- . Scan : initial − 0.8 V to end potential − 1.3 V Effect of solution parameters on the system behaviour Buffer concentration The effect of changing the ammonium buffer concentration over the range 0 -- 0.5 M is shown in Fig. 4a . Increasing the buffer concentration over the range 0 -- 0.1 M had a marked effect on the height of the Co -LRB- II -RRB- peak , but increasing the concentration further did not improve the peak height due to the increased background signal . As Co -LRB- II -RRB- and Ni -LRB- II -RRB- were sufficiently resolved at 0.1 M , this concentration was selected for all further experiments . Fig. 4Effect of solution parameters on the CAdSV behaviour of a 10 ppb Co -LRB- II -RRB- and Ni -LRB- II -RRB- sample : -LRB- a -RRB- ammonium buffer -LRB- pH 9.2 -RRB- concentration over the range 0 -- 0.5 M , -LRB- b -RRB- DMG concentration over the range 0 -- 0.35 M and -LRB- c -RRB- nitrite concentration over the range 0 -- 0.8 M. Supporting electrolyte conditions were : -LRB- b and c -RRB- 0.1 M ammonium buffer -LRB- pH 9.2 -RRB- , -LRB- a and c -RRB- 0.1 mM DMG and -LRB- a and b -RRB- 0.5 M NaNO2 . SW conditions were : conditioning potential − 1.3 V -LRB- 10 s -RRB- , deposition potential − 0.8 V -LRB- 120 s -RRB- , equilibration time 10 s , pulse frequency 25 Hz , pulse amplitude 40 mV -LRB- except c : 50 mV -RRB- and step potential 4 mV -LRB- except c : 5 mV -RRB- . Scan : initial − 0.8 V to end potential − 1.3 V Effect of DMG concentration Figure 4b shows the effect of increasing DMG concentration , over the range 0.01 -- 0.35 mM , on the peak current . The Co -LRB- II -RRB- signal increases almost linearly with increasing DMG concentrations , whereas the signal for Ni -LRB- II -RRB- steadily decreased to 0.2 mM whereupon it levelled off . The signal for the Co -LRB- II -RRB- peak shifted to a slightly more negative potential -LRB- − 4 mV -RRB- than that for Ni -LRB- II -RRB- with increasing concentration . A concentration of 0.3 mM was employed in the subsequent studies . Effect of nitrite concentration Figure 4c shows the influence of nitrite concentration on the sample signal . The Co -LRB- II -RRB- signal shows a rapid increase in signal to 0.3 M and a slower increase thereafter , whereas the Ni -LRB- II -RRB- peak height remained level to a concentration of 0.6 M. Concentrations of greater than 0.6 M lead to increased background current and so a nitrite concentration of 0.5 M was subsequently employed . Figure 5 shows the results obtained from the addition of nitrite to the sample system . Addition of nitrite to separate samples of Ni -LRB- II -RRB- and Co -LRB- II -RRB- results in increases in response for both metals , Fig. 5a and b , whereas the addition of nitrite to a mixture of 10 ppb Co -LRB- II -RRB- and Ni -LRB- II -RRB- results in an almost unchanged Ni -LRB- II -RRB- peak height whilst that for Co -LRB- II -RRB- has increased elevenfold , Fig. 5c . Figure 5d shows the reduction -LRB- 16.6 % -RRB- of the Co -LRB- II -RRB- peak height upon the addition of 10 ppb Ni -LRB- II -RRB- , while Fig. 5e displays the reduction -LRB- 51.9 % -RRB- of the Ni -LRB- II -RRB- peak height upon the addition of 10 ppb Co -LRB- II -RRB- . Fig. 5a Effect of nitrite on stripping peak for 10 ppb Ni -LRB- II -RRB- when -LRB- a -RRB- no NaNO2 is present ; -LRB- b -RRB- 0.5 M NaNO2 is present . b Effect of nitrite on stripping peak for 10 ppb Co -LRB- II -RRB- when -LRB- c -RRB- no NaNO2 is present ; -LRB- d -RRB- 0.5 M NaNO2 is present . c Effect of nitrite on the stripping peaks for 10 ppb Co -LRB- II -RRB- and Ni -LRB- II -RRB- when -LRB- e -RRB- no NaNO2 is present ; -LRB- f -RRB- 0.5 M NaNO2 is present . d Effect of addition of 10 ppb Ni -LRB- II -RRB- on the peak height for 10 ppb Co -LRB- II -RRB- in the presence of 0.5 M NaNO2 when -LRB- h -RRB- Co -LRB- II -RRB- only is present and -LRB- g -RRB- both Co -LRB- II -RRB- and Ni -LRB- II -RRB- are present . e Effect of addition of 10 ppb Co -LRB- II -RRB- on the peak height for 10 ppb Ni -LRB- II -RRB- in the presence of 0.5 M NaNO2 when -LRB- j -RRB- Ni -LRB- II -RRB- only is present and -LRB- k -RRB- Co -LRB- II -RRB- and Ni -LRB- II -RRB- are present . Carried out in 0.1 M ammonium buffer -LRB- pH 9.2 -RRB- with 0.1 mM DMG . SW conditions : conditioning potential − 1.3 V -LRB- 10 s -RRB- , deposition potential − 0.8 V -LRB- 120 s -RRB- , equilibration time 5 s , pulse frequency 25 Hz , pulse amplitude 50 mV and step potential 5 mV . Scan : initial − 0.8 V to end potential − 1.3 V Effect of varying the square-wave -LRB- SW -RRB- stripping parameters Frequency The effect of frequency was studied in the range 12.5 -- 100 Hz , Fig. 6a depicts the results obtained . The peak heights for both metals increased with increasing SW frequency -LRB- as expected from increasing the effective scan rate -RRB- and shifted towards more negative values . However , increasing the frequency leads to insufficient time for the background current to decay and leads to an increase in the contribution to the signal and hence a distortion in peak shape . A frequency of 25 Hz was generally the maximum employed . Fig. 6Effect of varying the SW stripping parameters on the CAdSV peaks of a 10 ppb Co -LRB- II -RRB- and Ni -LRB- II -RRB- sample . The variables are -LRB- a -RRB- frequency -LRB- 12.5 -- 100 Hz -RRB- , -LRB- b -RRB- step potential -LRB- 0.15 -- 7 mV -RRB- , -LRB- c -RRB- amplitude -LRB- 5 -- 70 mV -RRB- , -LRB- d -RRB- deposition potential -LRB- − 0.3 to − 0.9 V -RRB- , -LRB- e -RRB- deposition time -LRB- 30 -- 210 s -RRB- . Supporting electrolyte : 0.1 M ammonium buffer -LRB- pH 9.2 -RRB- with 0.1 mM DMG and 0.5 M NaNO2 . SW conditions were -LRB- unless being investigated as above -RRB- : conditioning potential − 1.3 V -LRB- 10 s -RRB- , deposition potential − 0.8 V -LRB- 120 s -RRB- , equilibration time 5 s , frequency 25 Hz , pulse amplitude 50 mV -LRB- except b and e : 40 mV -RRB- and step potential 5 mV -LRB- except e : 4 mV -RRB- . Scan : initial − 0.8 V to end potential − 1.3 V Step potential Figure 6b shows the effect of the step potential on the peak heights . The response for Co -LRB- II -RRB- rose rapidly with rising step potentials -LRB- i.e. with an increase in effective scan rate -RRB- up to 3 mV and thereafter levelled off , whereas the response for Ni -LRB- II -RRB- increased slowly to 7 mV . Peak resolution remained the same across this step potential range , with both peaks shifting towards more negative potentials . Values of 4 or 5 mV were generally found to be ideal for adequate peak definition . Amplitude The effect of the SW pulse amplitude was studied over the range 5 -- 80 mV . Both peaks shifted to more positive values , see Fig. 6c and they both initially increased rapidly in height -LRB- 5 -- 20 mV -RRB- whereupon the responses for Co -LRB- II -RRB- and Ni -LRB- II -RRB- levelled off as a result of the increase in contribution from the background current . A SW pulse amplitude of either 40 or 50 mV was employed . Deposition potential Figure 6d shows the influence of the deposition potential on the peak currents . At potentials less than − 0.4 V the Ni -LRB- II -RRB- signal is completely suppressed and the Co -LRB- II -RRB- signal was markedly reduced due to the oxidation of the bismuth film -LRB- at around − 0.46 V in alkaline solutions -RRB- . Both peaks gradually increase with potential in the range of − 0.4 V to a maximum at − 0.8 V and then exhibit a sharp reduction in adsorption at a more negative potential of − 0.9 V resulting from further reduction of the analyte complex . A deposition potential of − 0.8 V was selected , as this gave the maximum response for both metals . Accumulation time The influence of the accumulation time on the stripping signal was examined . Figure 6e depicts the response in peak currents for 10 ppb each of Ni -LRB- II -RRB- and Co -LRB- II -RRB- . For both Co -LRB- II -RRB- and Ni -LRB- II -RRB- the response increases with accumulation time up to 150 s for Co -LRB- II -RRB- and 180 s for Ni -LRB- II -RRB- , after which the response levels off , indicating saturation of the surface . An accumulation time of 120 s was used for subsequent measurements as this gave the best compromise between sensitivity and analysis time . Analytical characterisation The simultaneous determination of Co -LRB- II -RRB- and Ni -LRB- II -RRB- over the concentration range of 1 -- 20 ppb was investigated . The determination was carried out in 0.1 M ammonium buffer -LRB- pH 9.2 -RRB- with 0.1 mM DMG and 0.5 M NaNO2 . SW conditions : conditioning potential − 1.3 V -LRB- 10 s -RRB- , deposition potential − 0.8 V -LRB- 120 s -RRB- , equilibration time 5 s , pulse frequency 25 Hz , pulse amplitude 50 mV and step potential 5 mV . Scan : initial − 0.8 V to end potential − 1.3 V . The BiFE exhibited a well-defined response to changing concentration and the peak current response -LRB- y , μA -RRB- was linear over the concentration range -LRB- x , ppb -RRB- of 1 -- 20 ppb , as expressed in the calibration plots for Co -LRB- II -RRB- , y = − 11.4 x − 85.5 -LRB- R2 = 0.96 -RRB- , and Ni -LRB- II -RRB- , y = − 1.64 x -- 7.20 -LRB- R2 = 0.92 -RRB- , respectively . The stability of the BiFE formed on the injection-moulded substrates was investigated by performing replicate runs on the same film . Ten replicate runs gave an average peak height for Ni -LRB- II -RRB- of − 1.05 ± 0.10 × 10 − 5 A -LRB- rsd : 9.6 % -RRB- and for Co -LRB- II -RRB- − 8.19 ± 0.82 × 10 − 5 A -LRB- rsd : 10.0 % -RRB- . To compare the working electrode behaviour , AdSV and CAdSV of Co -LRB- II -RRB- and Ni -LRB- II -RRB- were also performed at a MFE in the flow cell . The solutions needed to be deaerated before analysis on the MFE . Neither AdSV or CAdSV of Co -LRB- II -RRB- on a MFE was possible in these particular flow cells due to a significant shift in the stripping peak to an extreme cathodic potential -LRB- < − 1.4 V vs Ag/AgCl -RRB- , which led to analytical uncertainty at the edge of the supporting electrolyte potential window . However , AdSV of Ni -LRB- II -RRB- was possible , as illustrated in Fig. 7 . It has been noted previously by this group -LSB- 57 , 58 -RSB- that the main drawback with MFEs is the difficulty involved in regenerating the active mercury surface after use , necessitating the inclusion of cleaning and mercury film renewal procedures . In contrast to the MFEs , the BiFEs formed in these flow cells enabled the generation of reproducible , quantitative analytical data , due to the enhanced stability of the bismuth films during the cleaning cycles , and the reduced cathodic potential for the cobalt stripping peak . Fig. 7AdSV of Ni -LRB- II -RRB- on a mercury film electrode . Here decimal numerals indicate the film number and roman numerals indicate replicate analyses carried out on the same film . Carried out with a 200 ppb sample in 0.1 M ammonium buffer -LRB- pH 9.3 -RRB- with 0.1 mM DMG -LRB- deaerated -RRB- at a flow rate of 16 μl/s . SW conditions : conditioning potential − 1.4 V -LRB- 10 s -RRB- , deposition potential − 0.5 V -LRB- 200 s -RRB- , equilibration time 5 s , pulse frequency 25 Hz , pulse amplitude 10 mV and step potential 4 mV . Scan : − 0.5 V to end potential − 1.4 V. Data were smoothed with a level-2 Savitzky -- Golay filter Conclusions The application of a BiFE for the determination of Co -LRB- II -RRB- and Ni -LRB- II -RRB- by square-wave CAdSV under flow conditions in a non-deaerated solution has been demonstrated . The injection-moulded carbon-fibre-filled polystyrene electrodes have proven to be suitable supports for the formation of bismuth films . The LiBr-containing plating solution was found to offer superior performance for both the RDE and the flow cell under the alkaline conditions required for CAdSV . The application of a flow stable BiFE in an inexpensive sensor system permits the possibility of autonomous on-site industrial and environmental monitoring . The inherent disposability of the devices coupled with their mercury-free status should advance the scope of the sensor to point-of-care clinical applications .

O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O B O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O

cobalt
nickel
dimethylglyoxime
electrochemical detection
flow cell
channel
sensor

square-wave adsorptive stripping voltammetry