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Table of contents

AZURA® Detector ECD 2.1 Instructions, V6685

8. Pulsed Amperometric Detection (PAD)

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8.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

9. Optimization of the working potential

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9.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

9.2 Electrochemical reactions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

9.3 Hydrodynamic and scanning voltammogram . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

9.4 Optimization using a voltammogram . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

9.5 Construction of a hydrodynamic voltammogram . . . . . . . . . . . . . . . . . . . . . . . . . . . .

9.6 Construction of a scanning voltammogram . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

10. Technical data

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10.1 Environmental, dimensions, weight & power requirements . . . . . . . . . . . . . . . . . . .

10.2 General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

10.3 DC mode . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

10.4 PULSE mode . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

10.5 SCAN mode . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

11. Rear panel I/O

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11.1 USB B connector. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

11.2 LAN connector . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

11.3 VALVE connector . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

11.4 ANALOG DATA connector . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

11.5 Digital I/O connector . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

11.6 Chassis grounding stud. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

12. Functionality tests

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12.1 Installation Qualification (IQ) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

12.2 Operation Qualification (OQ) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

13. Troubleshooting

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13.1 Instrument errors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

13.2 Analytical troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

13.3 Dummy cell test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

13.4 Stop flow test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

14. Reorders

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15. Index

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Содержание Azura ECD 2.1

Страница 1: ...HPLC Detector ECD 2 1 Instructions Document No V6685 ...

Страница 2: ...x 49 30 8015010 Internet www knauer net E Mail info knauer net Document number V6685 Version number 1 0 Release date 2019 10 23 Original edition The information in this document is subject to change without prior notice For the latest version of the instructions visit our website www knauer net library For the printed versions of our instructions we use environmentally friendly paper from sustaina...

Страница 3: ...nnections 17 4 Maintenance shutdown 23 4 1 Maintenance 23 4 2 Shutting down the system 24 5 ECD 2 1 controller 26 5 1 Introduction 26 5 2 Overview of ECD 2 1 screens 27 5 3 Parameters 31 5 4 ClarityChrom ECD 2 1 control module 34 5 5 Dialogue Elite 35 6 Detection and Parameters 36 6 1 Introduction 36 6 2 Three electrode configuration 36 6 3 Internal organization 37 6 4 Dual flow cell control 37 6 ...

Страница 4: ...gram 60 10 Technical data 64 10 1 Environmental dimensions weight power requirements 64 10 2 General 65 10 3 DC mode 65 10 4 PULSE mode 66 10 5 SCAN mode 66 11 Rear panel I O 67 11 1 USB B connector 67 11 2 LAN connector 68 11 3 VALVE connector 68 11 4 ANALOG DATA connector 73 11 5 Digital I O connector 76 11 6 Chassis grounding stud 83 12 Functionality tests 84 12 1 Installation Qualification IQ ...

Страница 5: ...training Fundamental knowledge of liquid chromatography Participation in an installation of the system performed by the manu facturer or a company authorized by the manufacturer and suitable training on the system and chromatography software Knowledge and experience in the safe handling of toxic and corrosive chemicals and knowledge of the application of fire prevention measu res prescribed for la...

Страница 6: ...nformation Please ship the instrument back to the manufacturer KNAUER Wis senschaftliche Geräte GmbH Germany at the end of life time of the product The manufacturer will take care of the proper disposal and recycling of the instrument at its facilities Shipping address for the end of life products KNAUER Wissenschaftliche Geräte GmbH Hegauer Weg 38 14163 Berlin Germany In case of questions or if f...

Страница 7: ...ood and met The caution sign denotes a hazard It calls attention to a procedure or practice which if not adhered to could result in damage or destruction of parts or all of the equipment and or erratic results Do not proceed beyond a cautions sign until the indicated conditions are fully understood and met The biohazard sign draws attention to the fact that use of biological materials viral sample...

Страница 8: ... checked regularly and maintenance procedures are carried out Preventive maintenance contracts are available for that purpose Please contact your local distributor or the nearest sales office for more information Electrical safety The removal of protective panels on the instrument can result in exposure to potentially dangerous voltages Therefore disconnect the instrument from all power sources be...

Страница 9: ... solvents used may be flammable toxic or corrosive The room in which the system is installed should be well ventilated to prevent that solvent vapors cause poisoning or ignite and cause a fire Use of open fire in the vicinity of this system must be strictly prohibited Do not install the system in the same room with any other equipment that emits or could potentially emit sparks Provide protective ...

Страница 10: ... or toxic solvents to accumulate Follow a regulated approved waste dis posal program Never dispose of flammable and toxic solvents through the municipal sewage system Applications quality control It is recommended that you routinely run several quality control samples Quality control samples should represent low average and high levels of a compound Make sure that quality control sample results ar...

Страница 11: ...ers can be controlled by either relays or TTL The ECD 2 1 can support up to 4 flow cells optional which makes it possible to perform 4 independent mea surements with one detector These instructions cover the installation set up and operation of the AZURA Detector ECD 2 1 only Detailed operation instructions for other peripheral LC equipment and parts such as flow cells pumps auto samplers valves c...

Страница 12: ...artment bn Ventilation holes Fig 2 Rear view Legend 1 Cell cabinet 2 Cell connector 9 pins sub D fem 3 Top fan heater intake 4 Door sensor 5 Mounting plate for cells columns 6 Door lock 7 Flow cell clamp for SenCell 8 Column clamp 9 Mounting hole for cell clamp M4 bl Bottom fan heater exhaust bm Mounting hole for column clamp M3 bn Fuse compartment bo Door panel rear bp Type label Fig 3 Oven compa...

Страница 13: ...s and in the list of supported drivers and software packages for all KNAUER devices document no V1667 Web site www knauer net softwarecontrol Laboratory requirements Your instrument is intended for indoor use only in an industrial or com mercial environment EN55011 group 1 class A ISM equipment It is suitable for the following categories Installation category II Pollution degree 2 equipment class ...

Страница 14: ...ed which is able to supply high purity deionized water with resistivity of 18 MOhm cm and low TOC level 10 ppb 3 2 Unpacking Inspect the transport box for possible damage as it arrives Immediately inform the transport company in case of damage otherwise it may not accept any responsibility Keep the transport box as it is designed for op timum protection during transport and it may be needed again ...

Страница 15: ...y Place the detector upright on its ins trument feet on a stable flat and smooth surface Do not place the instrument in an area subject to excessive dust or shocks Do not place it near a source of heat or in direct sun light as this may influence the heating capabilities of the instrument Make sure the detector is placed in such a way that the mains power connec tion can be reached easily to disco...

Страница 16: ...able mains power supply with a correctly installed protective earth conductor Never use the system without a properly connected protective earth conductor Leave the instrument powered off until specifically mentioned in the pro cedure below 3 4 PC connection Follow the instructions in this paragraph when the instrument is used with PC control over LAN using the Dialogue Elite or ClarityChrom Chrom...

Страница 17: ...bottom right of the Windows taskbar and open the menu Open Network and Sharing Center Alternative open Windows start menu open control panel and go to Open Network and Sharing Center Fig 6 Open Network and Sharing Center 2 Open the menu Change adapter settings on the left panel of the Open Network and Sharing Center window 3 Right click on the Local Area Connection icon of the LAN card in your PC ...

Страница 18: ... ECD 2 1 7 Connect the crossover UTP cable to the RJ45 Jack of the LAN card of your PC typically located on the backside of a desktop PC 8 Connect the other end of the crossover LAN UTP cable to the LAN port on the rear panel of the ECD 2 1 as depicted on the photograph 9 Switch on the ECD 2 1 Set the detector temperature to 35 C if a Performance Verification PV will be executed or set it to the t...

Страница 19: ... Netherlands ClarityChrom Chromatography software Version 6 x and higher from DataApex Czech The Elite Dialogue software is also required for the upload of new firm ware embedded software controlling the ECD 2 1 electronics In this section the installation and configuration of the ECD 2 1 is shortly described Note this is by no means a replacement of the installation documentation available for th...

Страница 20: ... the LAN connection is configured and the LAN cable is connected Power up the detector by means of the mains switch on the rear panel Start the program Dialogue Elite from the Windows start menu During start up the Select devices menu will pop up as shown in Fig 10 Fig 10 Dialogue Elite Select devices When a ECD 2 1 detector is available it is automatically detected and the IP address is shown in ...

Страница 21: ...same room with any other equipment that emits or could potentially emit sparks Wear protective gloves safety glasses and other relevant protective clothing when working on the device With respect to third party U HPLC equipment such as LC pumps auto samplers injection valves column heaters etc used in combination with the ECD 2 1 detector The equipment connected to the system should be specificall...

Страница 22: ...is compressed into the valve to make sure that the connection is leak tight Take the following into account when creating the connection If L is too long the ferrule cannot form a seal in the connection This may cause irreparable damage port of the valve column or other part as Part of the tubing may end up in the connection port internals Internals of the port may be damaged Fig 11 Too long tubin...

Страница 23: ...trodes in an electrochemi cal flow cell Low ion strength destabilises an EC system and noise will increase Extremely high buffer concentrations cause problems of salt formation Therefore concentrations between 20 and 200 mmol L are recommended Also constant pH is important for baseline stability and reproducible re sults Stability of pH is best when close to pKa of a buffer ion Often used buffers ...

Страница 24: ... the rear panel to ena ble INJECT LOAD functions In the CONFIG screen the valve option should be set to Valve present manually 3 An electrically actuated valve of VICI Valco type E2CA EHCA can be connected controlled by the ECD 2 1 For that purpose a serial val ve cable is required The serial cable should be connected to the VAL VE connector on the rear panel of the instrument to enable INJECT LOA...

Страница 25: ...hrough the flow cell will lead to unaccepta ble noise levels and spikes Therefore the use of an in line degasser is strongly recommended In our experience a one time degassing step of the HPLC buffer is almost never sufficient If the ECD 2 1 is used for reductive ECD at a negative working potential additional steps should be taken to remove oxygen from the mobile phase These include degassing with...

Страница 26: ...should lead to a reservoir that is at a higher level than the flow cell This ensures a small back pressure which prevents air bubble entrapment The outlet tubing should be under the liquid level to avoid electrical noise induced by dripping of mobile phase 11 Set the cell potential see page 54 for optimization of the poten tial switch ON the flow cell and allow the system to stabilize for approxim...

Страница 27: ...tual temperature exceeds 70 C switch off the detector and contact the manufacturer or its representatives for service Flow cell Check the performance of the detector flow cell on a daily basis by eva luating background current noise and signal An increase in background current noise and or loss of sensitivity may be a sign of contamination of the working electrode WE and or a sign that maintenance...

Страница 28: ...an LC system with electrochemical detector for a longer period of time Shutting down is not different from most other HPLC systems Perform the following procedure Switch off the flow cell using the keyboard standalone or via the soft ware ClarityChrom or Dialogue Elite Check the column s documentation for the appropriate storage liquid apply this and make sure the column is properly flushed A reve...

Страница 29: ...ase of a VT 03 or Flex Cell the SenCell does not have a spacer Close the cell and store dry In case of salt bridge REF store the REF separately with a cap on it to prevent drying out Alternatively put the sb REF in a 10 mL vial under a KCl solution and close the vial with a cap Switch off the detector and other LC equipment via the mains switch switch to position 0 on the rear panel ...

Страница 30: ...e and the screen s of appearance The nature of an item can be 1 Control parameters with a cursor box can be attained via cursor buttons and changed by the value button 2 Status without a cursor box a parameter reflects the current status 3 Functions parameters in CAPITALS are commands accessible via function buttons F1 F5 4 The Enter button is only used to accept changes in cell potential In the t...

Страница 31: ...27 ECD 2 1 controller AZURA Detector ECD 2 1 Instructions V6685 5 2 Overview of ECD 2 1 screens DC mode ...

Страница 32: ...28 ECD 2 1 controller AZURA Detector ECD 2 1 Instructions V6685 Pulse mode ...

Страница 33: ...29 ECD 2 1 controller AZURA Detector ECD 2 1 Instructions V6685 Scan mode CONFIC menu ...

Страница 34: ...30 ECD 2 1 controller AZURA Detector ECD 2 1 Instructions V6685 DIAG menu ...

Страница 35: ... zero offset Switches ON if AZERO is pressed Affects auto zero com pensation only not the offset C CONFIG main Enters config screen F Contrast config Sets the contrast of display C Cyc scan setup Controls the nature of the cycle half full and continuous Half means that the cell potential runs from E1 to E2 and stops at E2 Full means that the cell potential runs from E1 to E2 and back to E1 and the...

Страница 36: ...gles between inject I and load L If a manual injector with position sensor is applied it echoes the position of the injector If an electrically actuated injector is used optional it is possible to switch the injector with this function button F S KEYB test Enters KEYB screen for keyboard test Press 2x F1 to leave F MARK dc stat pulse stat Triggers a marker signal on output F MaxComp dc setup pul s...

Страница 37: ...1 and 2 and Relays 1 and 2 status 0000 and sets the electric valve to load if present F SYSTEM diag Enter SYSTEM menu F t pulse setup2 pulse stat Displays the total duration of one pulse t1 t2 t3 t4 t5 S t1 t2 t3 t4 t5 pulse setup2 pulse setup3 Duration of potential step E1 E2 E3 E4 and E5 Time can be set between 0 t2 t5 or 100 t1 and 2000 ms in 10 ms increments Maximum pulse duration is 9999 ms C...

Страница 38: ... be set controlled measurement mode potential settings Within a ClarityChrom method it is possible to execute timed event by programming time lines under the Time table tab Running a Time table enables time based automated parametric control of the electrochemical detection ECD 2 1 during a run This is particularly useful when during a run settings have to be changed such as the sensitivity auto z...

Страница 39: ...wizard OQ PQ and dummy cell measurements device calibration There are no tools for further data analysis therefore Dialogue cannot be considered as fully featured chromatography software package The free demo version of this program has already all functionality implemented functionality is unlocked by a license dongle The free version is primarily used for controlling all operational parame ters ...

Страница 40: ...ls with a three electrode configuration see Fig 18 The working potential is set between the working electrode WE and the auxiliary electrode AUX The AUX is kept at a precisely defined reference electrode REF potential by means of the so called voltage clamp This is an electronic feedback circuit that compensates for polarization effects at the electrodes At the WE which is kept at virtual ground t...

Страница 41: ...t is amplified by the current potential I E con verter Fig 19 Fig 19 ECD 2 1 signal processing from electrochemical flow cell to output The signal from the I E converter can be compensated with auto zero or offset and is digitized using a 24 bits ADC In the CPU the signal is pro cessed for example noise filtering or more complex data processing in PAD Finally after the 16 bits DAC the signal is se...

Страница 42: ...ond flow cell is a SenCell or VT 03 cell which is used for detection Note that it is necessary to work with micro HPLC because the conversion rate of the reactor cell is too small when using standard HPLC Fig 20 Typical configuration for serial mode detection Cell 1 is a FlexCell cell 2 is a SenCell for detection Channel 1 and 2 use the same time base of system 1 Parallel mode detection In paralle...

Страница 43: ... second sensor board is not installed or not properly recognized Second a new status screen is available in dual cell systems which indica tes the status of both cells in a single screen However for convenience it is advisable to use PC control from the ClarityChrom Chromatography software or Dialogue Elite software when working with 2 flow cells Fig 23 ECD 2 1 main menu top with active cell indic...

Страница 44: ...mum compensation at the less sensitive ranges This is an inevi table consequence of the tremendous dynamic range that is covered by electrochemical detection Table II DC ranges and maximum compensation Range FS Max comp Range FS Max comp 200 µA 2 5 mA 20 nA 2 5 µA 100 µA 2 5 mA 10 nA 2 5 µA 50 µA 2 5 mA 5 nA 250 nA 20 µA 2 5 mA 2 nA 250 nA 10 µA 2 5 mA 1 nA 250 nA 5 µA 2 5 mA 500 pA 250 nA 2 µA 25...

Страница 45: ...y of the output can be inversed Oxidative and reductive analy ses generate opposite currents For data acquisition traditionally chroma tographic peaks have a positive amplitude Therefore selection of polarity is useful Filter High frequency noise is efficiently removed and chromatographic peaks can be detected with better signal to noise ratio Fig 25 Signal to noise ratio is improved using a filte...

Страница 46: ...e Hz Data rate Hz RAW 100 default 50 20 10 5 2 1 10 100 5 50 2 1 20 OFF 10 0 5 0 2 0 1 10 0 05 5 0 02 2 0 01 0 005 0 002 0 001 1 Filter OFF is also a special case The data rate is fixed to 10 Hz and the data is not filtered Setting OFF is therefore the same as RAW at 10 Hz Pulse mode In the pulse mode the working electrode is dynamically and continuous ly regenerated by a series of potential steps...

Страница 47: ... and finally at a few chromatograms before and after applying ADF 7 2 Frequency A scientific definition of frequency is the number of completed altera tions per unit time It has two dimensions count and time Frequency is usually expressed in Hz which is counts per second The counts themselves can run in a regular evenly spaced manner as with sine waves whose curve shapes do not change Alternativel...

Страница 48: ...pressed in Hz Fig 28 Frequency tells how often something happens 1 peak in about 0 25 min 15 s f 1 15 0 07 Hz This is further illustrated by an overlay of the same chromatographic peak with a sine of 0 07 Hz Fig 29 Fig 29 Overlay of a chromatographic peak with 0 07 Hz sine 400000 300000 200000 100000 0 100000 200000 300000 400000 4 0 4 1 4 2 4 3 4 4 4 5 4 6 4 7 4 8 4 9 5 0 t min 4 56 4 15 40000 14...

Страница 49: ...andom noise in chromatography lower trace Both frequen cies 0 4 and 10 Hz can be recognised amongst others Looking closely to the lower noise trace both frequencies and others can be recognised This is typical for noise in chromatography a collec tion of more or less random frequencies Low pass noise filters The way noise filters work is by suppressing certain frequencies in the acquired signal Ty...

Страница 50: ...ning average filter takes the average of n data points to create a new data point For example in a 5 points running average filter output data point y 80 is calculated from measured data points x 80 x 84 as Each input data point has the same weighting factor of 1 5 In more advanced digital signal processing a more complicated equation is used to calculate the output data point y n In contrast to t...

Страница 51: ...litude is 70 of the input signal also known as 3 dB point Fig 33 An amplitude response plot of a low pass filter with a cut off frequen cy of 1 Hz It is a 2 A 4 B and 8 C pole Bessel filter In Fig 33 it is shown that the number of poles is important a filter beha ves more ideal with increasing number of poles In a hardware filter the number of poles is the number of filter circuits that are placed...

Страница 52: ...ilter is it is only possible to apply low pass filtering if noise frequencies are higher than the frequency of the signal Fig 35 From top to bottom filter setting of 0 5 0 02 and 0 002 Hz Narrow peaks in front of the chromatogram are deformed at 0 002 Hz where as wider peaks show hardly any deformation see peak at t 13 min Attenuation of noise is shown in the close up on the right Prerequisite for...

Страница 53: ...le to switch to a stronger filter setting for the second half of the chromatogram using a timed event To optimize for the best S N ratio use the lowest acceptable cut off frequency After optimization do not change the cut off frequency setting during analysis of a calibration sequence Use the same settings for analysis of samples and calibration standards Fig 36 Analysis of 20 pmol L 5 hydroxytryp...

Страница 54: ...is applied in a cyclic manner The signal is sampled during a fraction of the total pulse cycle During the sampling time ts the signal is collected and this value is sent to the detector output The frequency of data output is determined by the pulse duration t1 t2 t3 t4 t5 Usually the duration is between 0 5 and 2s data rate between 2 0 5 Hz The background or cell current is usually considerably hi...

Страница 55: ...er oxide formation they require regular monthly maintenance HyREF reference electrodes are maintenance free under these conditions and are therefore particularly suited If a mobile phase is used with a high pH pH 10 carbohydrate analysis the standard Vespel rotors from the injection valve should be replaced by Tefzel rotors which are pH resistant For carbohydrate analysis only CO2 free sodium hydr...

Страница 56: ...the sample constituents and the required detection frequency The impression may arise that the number of variables 5 potential steps and 6 time settings may lead to a time consuming optimi zation procedure In practice the pulse mode is more straightforward and published in several excellent review papers and application notes Output frequency An important difference between the DC and the pulse mo...

Страница 57: ... of a metal oxide layer This is impos sible with glassy carbon References 1 D C Johnson D Dobberpuhl R Roberts and P Vandeberg Review Pulsed amperometric detection of carbohydrates amines and sul phur species in ion chromatography the current state of research J Chromatogr 640 1993 79 96 2 D C Johnson en W R LaCourse LC with pulsed ECD 2 1 at gold and platinum electrodes Anal Chem 62 1990 589A 597...

Страница 58: ...runs between two preset potential values E1 and E2 and scan speed in mV s and the current is measured Optimization of the working potential and the construction of a voltam mogram is described 9 2 Electrochemical reactions In electrochemical detection ECD a reaction of the analyte at an electro de surface is monitored This distinguishes ECD from most other detec tion techniques where detection is ...

Страница 59: ...nt Icell is plotted Fig 41 Hydrodynamic voltammogram of norepinephrine A at a glassy car bon working electrode and the current of the baseline B At E1 the electrochemical signal becomes diffusion limited Scanning voltammogram An alternative for the chromatographic construction of an I E relationship is the application of scanning voltammetry In a scanning voltammetry experiment the working electro...

Страница 60: ...in buffer A scan of the buffer blank should be used to distinguish between solvent peaks and analyte peaks Any contamination in the buffer may lead to artefacts As can be seen in both Fig 41 and Fig 43 when the working potential is increased the electrochemical reaction is enhanced hence the signal increases At a certain potential the I E curve flattens All analyte molecu les that reach the workin...

Страница 61: ...hen interfering peaks appear in the chromatogram it is possible to optimize the method with regard to selectivity If the interfe ring compound has a higher oxidation potential a working potential is chosen that gives the best selectivity i e the largest difference in peak height In the example of Fig 44 the selectivity for compound X is im proved considerably by decreasing the potential to E2 or E...

Страница 62: ...the working potential is lower E1 and the signal is not diffusion limited the signal will strongly increase at higher pH In both cases the background current increases at a higher pH Fig 45 At a higher pH the I E curve of norepinephrine is shifted to the left Reaction kinetics predict that electrochemical detection is mass flow de pendent When the LC flow is stopped in LC EC the analyte will be ox...

Страница 63: ... started by injecting the compound When at the high working potential no signal is obtained it may be concluded that the compound is not electrochemically active In such a case derivatiza tion of the compound may be an option 4 If a peak is measured the working potential is decreased by 50 or 100 mV and step 2 to 4 is repeated until the lowest potential setting Fig 46 5 The peak heights and the ba...

Страница 64: ...oltammogram can be programmed under the Detector tab in the main window Fig 48 Programming a scan in the Elite Dialogue software In the above example a half scan is programmed sweeping the poten tial from 0 V to 1 0 V A full scan would include the reverse scan i e from 0 V to 1 0 V and back to 0 V In the continuous mode the voltage is swept up and down between both potentials for a pre defined run...

Страница 65: ...ump The Antec dual syringe infusion pump which can be obtained as accesso ry has as an advantage that it can be controlled in Elite Dialogue software as well Fig 50 Programming the Antec infusion pump in Elite Dialogue In the example in Fig 51 a half scan is shown at a flow rate of 10 µL min of a 20 µM Serotonin 5 HT standard in mobile phase ...

Страница 66: ...up only I versus t curves can be obtained Prepare a solution of the pure compound dissolved in preferably the HPLC buffer with a concentration in the range of 10 100 µM Set the lower E1 and upper potential E2 the scan rate Spd range and scan cycle in the SCAN SETUP menu or under the detector tab in Dialogue Typical scan settings to start with E1 0 mV E2 1000 mV scan speed 10 mV s range 5 µA V Opti...

Страница 67: ...ing the START button in the SCAN STAT menu or by starting a single run in Dialogue press the F5 button or click start single run under the Options menu To record a background blank scan repeat the experiment with the pure HPLC buffer in which the analyte was dissolved Fig 52 Example of multiple scans continuous of Amodiaquine ...

Страница 68: ...t smooth surface keep space under the detector free risk of blo cking power supply fan Power requirements 100 240 VAC 50 60 Hz 260 VA auto sensing Mains fuse 2 5 AT 250V 5x20 mm IEC 60127 2 For safety reasons any other internal fuse or circuit breaker is not operator accessible and should be replaced only by authorized personnel by the manufacturer Only use manufacturer supplied fuses For optimum ...

Страница 69: ...ins sub D Male Valve 1x 9 pins sub D Female Analog output 1x 25 pins sub D Female Digital I O Analog output DAC 1 to 1 V full scale via 16 bit D A converter Analog output I E 2 5 to 2 5 V full scale unprocessed I E con verter signal Digital I O HW 2x Relay 5x TTL outputs CMOS 3 3V logic 13 TTL inputs programmable 1x GND Programmable I O functionality Cell on Cell off Autozero Start Overload Relay ...

Страница 70: ...2 50 V to 2 50 V with 10 mV increments Data Rate 1 pulse duration Hz Waveform Max 5 potential steps Pulse times t1 t5 t1 100 ms 2000 ms t2 t3 t4 t5 0 2000 ms in 10 ms increments Sampling times ts 20 ms t1 60 ms 10 5 SCAN mode Range 10 nA 200 µA in 1 2 5 increments Potential Ec 2 50 V to 2 50V with 10 mV increments Data Rate 1 Hz Scan rate 1 100 mV s in 1 2 5 increments Cycle Half Full Continuous ...

Страница 71: ...l connectors is shown below in Fig 53 for reference Fig 53 ECD 2 1 rear panel 11 1 USB B connector USB type B connector for serial instrument control over USB for service use only Based on USB to serial UART interface using the FT232R chip from FTDI Future Technology Devices International Ltd FT232R is fully compliant with USB 2 0 specifications Fixed communication baud rate 921600 bps Communicati...

Страница 72: ...on for details about configuration and set up of communication over LAN 11 3 VALVE connector Serial D9 9 pins subD male valve control connector for electrical VICI and manual valves Serial RS232 valve control of VICI 2 position electrically actuated valves via pin 1 3 see VICI technical note tn413 pdf on the official www vici com web site Compatible with the E2CA and EHCA actuator control module A...

Страница 73: ...on A VICI electrically actuated 2 position valve is automatically detected du ring start up of the instrument Automatically the Valve parameter in the ECD 2 1 IO menu is set to Valve Present Fig 54 Valve parameter Always restart the detector by re powering via the mains switch on the rear panel when connecting the valve for correct initialization of an electrically actuated VICI valve In the case ...

Страница 74: ...a manual valve the valve present parameter has to be set manu ally Configuration of a manual valve can be done via the LCD screen or the Dialogue Elite software LCD display Configuration of a manual valve can be done by setting the Valve parameter in the IO menu manually to Valve Present Dialogue Elite Go to the device settings and the EC device tab and check the manual valve check box ...

Страница 75: ...ically actuated valve and INJ I represents the INJECT position B on the actuator control module The LOAD position is the default position at start up of the instrument in the case of an electrically actuated valve Fig 58 Valve status In stand alone mode the valve position can be controlled via the F5 func tion button in the STAT screen In the case of a manual valve the infor mation above the F5 bu...

Страница 76: ...e Monitor tab with valve status position displayed An electrically actuated VICI valve can also be controlled via Dialogue Elite under the detector tab see Fig 56 Note that the valve will switch immediately after a change from Load to Inject checkboxes or vice ver sa no Send to device action is required An electrically actuated VICI valve can also be controlled via timed events under the events ta...

Страница 77: ...ing instructions 11 4 ANALOG DATA connector The ECD 2 1 is equipped with an analog data output connector to provi de the measured signals in millivolts mV for users who work stand alone without PC control with the instrument The ANALOG DATA connector D9 female can either be connected to an X Y recorder or A D converter The ANALOG DATA output can supply either a non manipulated signal directly from...

Страница 78: ...b analog Output setting DAC output The DAC output is the processed signal by the ECD 2 1 s CPU and is iden tical to that of the digital cell current signal obtained via data acquisition over LAN using the Elite Dialogue or the ClarityChrom chromatography software The signal in mV from this output is directly related to the range setting of the ECD 2 1 So for example in the case the 200 µA measurem...

Страница 79: ...r of 100M 10M 1M 100K or 1K in the I E converter circuit The actual cell current can be calculated from the analog I E output in Volt using the following formula Ic Ampere Vout Volt RI E Ohm x 10 So for example an I E output signal of 250 mV in the 5 nA range RI E 10 MΩ corresponds to an actual cell current of 0 25V 10 x 1E7Ω 2 5E 9A 2 5 nA In the table below the I E resistor value is listed for e...

Страница 80: ...ector The detector has one 25 pins digital I O connector which enables control of or by external equipment The IO connector contains 18 TTL contacts 5 outputs and 13 inputs 3 3V CMOS logic 2 RELAYS contact closure and 1 ground GND connection TTL inputs outputs Both the TTL inputs and outputs are default high 3 3 Volt The TTL inputs are level triggered the contacts require a minimum TTL low pul se ...

Страница 81: ...h the measurement is performed see Fig 65 Fig 65 Example of a chromatogram were the cell current exceeds the maxi mum current level and the signal is out of range It is important to recognize an out of range overload situation be cause it may lead to erratic results when quantifying analyte concen trations in samples If for instance the cell current goes in out of range during the recording of an ...

Страница 82: ...ells will go Out of range the overload input pin 11 will become active Cell on Cell off The ECD 2 1 has 3 TTL inputs to switch on cells pin 12 14 and also 3 inputs to switch off cells pin 15 17 AUX1 AUX4 pin 7 10 This input command can be used for example to switch on and stabilize the flow cell early in the morning by means of a timer Two of the inputs are confi gurable pin 12 pin 15 cell on and ...

Страница 83: ...s hidden and not shown in the monitor tab To be able to see the status and control buttons for the I O go to the Device settings window settings tab and check the Show status outputs checkbox under the controls section Fig 67 Dialogue Elite Detector tab with I O status and control buttons The status of the Relays and TTL inputs can be set to active or inactive The selected state of the outputs wil...

Страница 84: ...RA Detector ECD 2 1 Instructions V6685 Fig 68 Dialogue Elite Event Editor window Programming of output A Fig 69 Dialogue Elite Event table window with an example of programming timed events for output A in this case Relay 1 ...

Страница 85: ...time file Outp 0010 Command 47 9 TTL OUT AUX 3 Free programmable TTL output activated by Command 47 10 TTL OUT AUX 4 Free programmable TTL output activated by Command 47 11 TTL OUT Overload A 1 5 Active in case of signal overload OUT OF RANGE PAD OVLD 12 TTL IN Cell on 1 A 1 5 Trigger to switch the cell on 13 TTL IN Cell on 2 Trigger to switch the cell on 14 TTL IN Cell on 3 Trigger to switch the ...

Страница 86: ...tart a Scan and Data acquisition 25 GND GND Ground connection A All boards In the detector accessory kit a dedicated I O cable is supplied with an I O connector board with screw terminal connections for easy connection of open ended electrical wiring for I O ECD 2 1 I O conn Board ECD 2 1 I O cable 25M 25M 1 8m For use connect the I O cable to the 25 pins I O connector on the detec tor rear panel ...

Страница 87: ...int of the instrument can be used for shielding purposes For example to shield the flow cell outlet from external electrical interferences troubleshooting noise issues or to shield the flow cell from other equipment which might be coupled in series with the electrochemical detector such as an MS Fig 70 Left ECD 2 1 rear panel grounding stud Right example of shielding the flow cell by grounding the...

Страница 88: ...he functionality of the device 12 2 Operation Qualification OQ The Operation Qualification includes an extensive functionality test accor ding to KNAUER standard OQ documents The Operation Qualification is a standardized document and free of charge It is not part of the delivery Please contact the Customer Support in case of request The Operation Qualification includes the following definition of ...

Страница 89: ...sage containing an error number with a short de scription for several hardware fault conditions which will be shown in the instruments LCD display Table IX Error messages Error Message 11 Checksum error 12 Temperature sensor 1 error 13 Disconnect flow cell x 14 Control board SRAM error 20 Checksum error Please contact your local supplier if one of the above errors occur for further instructions In...

Страница 90: ...02 PAD overload Charging current in pulse mode out of range Pressing AZERO may give an adequate re ad out again If not it is advisable to change the pulse settings increase t1 or use a less sensitive range Make sure that maintenance is performed on a regular basis An Out of range error appears when the cell current Icell exceeds the limit of the current range at which the measurement is performed ...

Страница 91: ...m pump autosampler pulse damper column mobile phase etc 13 3 Dummy cell test External dummy cell An external dummy flow cell is shipped with every ECD 2 1 instrument for troubleshooting purposes and maintenance checks The dummy cell test can be performed standalone via the LCD display in combination with an A D converter or via the Dialogue Elite ClarityChrom software A successful dummy cell test ...

Страница 92: ...nternal dummy cell The ECD 2 1 also has the option to run a so called internal dummy cell test This exclusively checks the performance of the electronic circuit boards amplifier circuitry only so it excludes the cell cables and the external dummy flow cell From the MAIN screen DIAG can be selected to enter the DIAG screen followed by selecting NOISE This activates a timer in the NOISE screen and a...

Страница 93: ...rent generated with the internal dummy cell The internal dummy cell current and noise level should fall within the criteria specified on the previous page Make sure to uncheck the Internal dummy cell option again from the device settings menu after the test Fig 75 Right Device settings with internal dummy cell option checked Left side internal dummy cell current shown in the monitor tab 13 4 Stop ...

Страница 94: ...are the obtained values of the cell current and noise with the values observed before the stop flow with the pump on Results Icell In case a significant drop in Icell is observed for example a drop of more than 50 it is an indication that the problem is not flow cell related but originates from other parts of the LC system The most obvious rea sons for a high background current are electrochemical...

Страница 95: ...ytical problems may also be caused by ex ternal Influences like temperature unstable samples Make sure that the application was running trouble free before and that no changes have been made to the system A number of causes and possible solutions for analytical problems is listed below Contact your local representative if you need further help No detector response Possible cause Remedy No power Ch...

Страница 96: ...ed Refill with saturated KCl add wetted KCl crystals Air bubble in REF or in cell Remove air bubble continuously degas the mobile phase Slow temperature fluctuations Isolate detector cell set oven temperature Fouled WE Clean WE Leaking REF or cell Tighten connections with care Decreased sensitivity low S N ratio Possible cause Remedy Fouled WE by dirty samples Clean WE if possible dilute samples C...

Страница 97: ...ar pattern Check pump seals valves Over tightened cell bolts Adjust cell bolts check pump pressure Air bubbles in cell or REF Maintenance REF Temperature oscillations Set oven temperature Contaminated buffer high Icell Replace buffer do not recycle the buffer Fouled WE Clean WE Fe in buffer Add EDTA passivate metal parts with HNO3 ...

Страница 98: ...7 Column clamp 12 mm 250 0102 Cell clamp kit 40 mm 2 500 113 Fuse 2 5 AT 250 V 2 500 170 LAN UTP cable crossed 3 meter 2 500 175 USB cable A B 3 meter 250 0131B I O conn board 250 0131C I O cable 25M 25M 1 8 meter 250 0128A Output cable D9 male open 2 meter 2 500 116 Mains cable Europe 2 500 118 Mains cable USA 250 0126A Cell cable For a VICI Valco electrically actuated 2 position valve with an E2...

Страница 99: ...hemical reactions 56 Elite Dialogue software 15 EN 55011 group 1 class A ISM equipment 12 Environmental specifications 9 Errors 87 F Filter 43 settings 33 Filters low pass noise 47 Flow cell 25 Frequency 45 Fuse replacement 5 26 H Helium degassing 21 I I E converter 38 39 output 77 relationship 56 Inject marker 71 Installation 9 Installation Qualification IQ 86 Instrument description 7 Integrator ...

Страница 100: ...ime 52 Scan mode example 62 scanning voltammogram 57 Serial mode detection 40 Signs 3 S N ratio decreased sensitivity 94 filter settings 50 Solvents 5 Stop flow test 91 System operation 4 T t1 35 Technical data 66 Tefzel rotor 53 Three electrode configuration 38 Time table 36 Transport box 10 ts sampling time 52 U USB connector 8 69 V Vacuum degassing 21 Valve configuration electrical 71 manual 72...

Страница 101: ...AUER instructions online www knauer net library KNAUER Wissenschaftliche Geräte GmbH Hegauer Weg 38 14163 Berlin Phone Fax E Mail Internet 49 30 809727 0 49 30 8015010 info knauer net www knauer net KNAUER 2019 ...

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