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175_0010 - DECADE Elite user manual - rev 01

1. CHAPTER 1 Introduction DECADE Elite Back side Lad 9 TT nte i 15 Description Description 1 Instrument rear panel 7 USB connector USB B 2 Type label pn sn etc 8 Fuse amp power rating 3 Digital I O connector 25 pins sub D fem 9 Mains switch inlet 4 Analog data 9 pins sub D fem 10 Grounding stud 5 Valve connector 9 pins sub D male 11 Fuse compartment 6 LAN connector RJ45 jack 12 Ventilation holes 16 DECADE Elite Oven compartment 175 0010 DECADE Elite user manual rev 01 it Description Description 1 Cell cabinet 8 Column clamp 2 Cell connector 9 pins sub D fem 9 Mounting hole for cell clamp M4 3 Top fan heater intake 10 Bottom fan heater exhaust 4 Door sensor 11 Mounting hole for column clamp M3 5 Mounting plate for cells amp col 12 Fuse compartment umns 6 Door lock 13 Door panel rear 7 Flow cell clamp for SenCell 14 Type label CHAPTER 2 Installation 17 CHAPTER 2 Installation Site Preparation Requirements For a successful onsite installation of the instrument please arrange the following requirements at your location in advance Personal Computer In case the instrument is used via remote control by PC software Dia logue Clarity or firmware FW update needs to be performed a desktop computer is required with the following require
2. DECADE Elite controller 53 Parameter Screen Description Type programmed the next run is started Cannot be smaller than smallest time in time file 0 01 min Is therefore never smaller than 0 01 min File events setup Selected time file number In the DC mode file numbers 1 5 are available in the pulse mode file numbers 6 9 can be selected The time files remain stored in RAM also after switching off the DECADE Elite Time files can be uploaded via LAN Hold resume run scan stat Toggle holds or resumes execu tion of time file or scan HOLD 0 1 run scan stat Holds or continues execution of time file or scan Toggles between 1 and 0 Pressing hold again con tinues time file or scan were it has been hold prog Board identifier for multi cell pur pose Indicates for which boards time file settings apply Binary coded Offs run Displays percentage offset during execution of a time file Outp prog Control of four output functions in EVENTS Is open high if 0 is closed low if 1 AUX1 0001 AUX2 0010 relay 1 0100 relay 2 1000 Combinations are possi ble QUIT run Aborts the time file and returns to the EVENTS SETUP screen The cycle counter Cy is reset to 1 Outputs Aux 1 and 2 and Re lays 1 and 2 are reset status 0000 RUN events setup Enters RUN screen System waits w
3. An example of an electrochemical reaction is shown in Fig 31 norepi nephrine is converted into a quinone by oxidation at the WE Two elec trons are transferred at the WE resulting in an electrical current that is am plified by the controller Chapter 5 Detection and Parameters 85 Hydrodynamic and scanning voltammogram Hydrodynamic voltammogram A hydrodynamic voltammogram is constructed when the pure analyte is not available and separation over an analytical column is required Further more under real chromatographic conditions reliable information about the S N ratio is obtained The peak heights obtained from the sequence of chromatograms are plotted against the working potential used Also the background current I cell is plotted Icell nA gi ie uw Ecell V Fig 32 Hydrodynamic voltammogram of norepinephrine A at a glassy carbon working electrode and the current of the baseline B At E the electrochemical signal becomes diffusion limited 86 175 0010 DECADE Elite user manual rev 01 Scanning voltammogram An alternative for the chromatographic construction of an I E relationship is the application of scanning voltammetry In a scanning voltammetry experi ment the working electrode potential is ramped up and down between two preset potentials E1 and E2 and the current is measured while the ana lyte is continuously flushed through the flow cell This is repeated as many times as desired The
4. Antec Industrieweg 12 2382 NV Zoeterwoude The Netherlands DECADE Elite User Manual 175 0010 Edition 1 2015 ISO 9001 T 31715813333 F 31715813334 info myantec com www myantec com certified Copyright 2015 Antec The Netherlands Contents of this publication may not be reproduced in any form or by any means including electronic storage and retrieval or translation into a foreign language without prior agreement and written consent from the copyright of the owner Antec reserves the right to make changes to the design and specifications of the instrument and this manual without prior notice ROXY ALEXYS DECADE DECADE II DECADE Elite DECADE Lite INTRO Flexcell ReactorCell ISAAC HyREF Sen Cell and SynthesisCell are trademarks of Antec Whatman word and device and Whatrnan word only are trademarks of Whatman International Ltd SOLVENT IFD and AQUEOUS IFD are trademarks of Arbor Technologies Inc Clarity DataApex are trademarks of DataApex Ltd Microsoft and Windows are trademarks of Microsoft Corporation Excel is a registered trademark of the Microsoft Corporation All other trademarks are the property of their respective owners The software and the information provided herein is believed to be reliable Antec shall not be liable for errors contained herein or for incidental or consequential damages in connection with the furnishing performance or use of software or this manual All use of
5. Installation 19 See check list below for reference 1 Delivery is in accordance with order O 2 Delivery is undamaged O 3 All items on checklist s are included O 4 Certificates of performance are included detector O flow cell s O 5 User manual s is are included on USB stick O Note that flow cells are not part of the DECADE Elite detector and have to be ordered separately To unpack the DECADE Elite lift it from its box by both hands Fig 1 Never lift the DECADE Elite at its front door but at its sides lift here Fig 1 Lift instructions DECADE Elite With both hands under the instrument lift the DECADE Elite to its operation location Install the detector in an area which meets the environmental conditions Fan Fig 2 Location of power supply fan DECADE Elite 20 WARNING 175 0010 DECADE Elite user manual rev 01 Remove the protective tape from the DECADE Elite LCD screen Leave the instrument to adopt ambient temperature for at least half an hour in the place of installation Use the detector indoors only Place the detector upright on its instrument 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 connectio
6. Valve configuration A Vici electrically actuated 2 position valve is automatically detected dur ing start up of the instrument Automatically the Valve parameter in the IO menu is set to Valve Present Contrast vout source i Di master 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 case of a manual Valve configuration has to be done manually by set ting the Valve parameter in the IO menu to Valve Present Valve connector layout In the table below the connector layout is shown for the Valve connector Table V Valve connector lay out GND TRANSMIT TxD RECEIVE RxD GND SWITCH Hand valve INJECT MARKER 7 2 3 4 5 6 7 8 9 102 175 0010 DECADE Elite user manual rev 01 For a manual valve the status LOAD INJ read out is established by con necting the LOAD INJ sensor wires leads of a manual valve to pin 5 and 6 contact closure CHAPTER 11 Rear panel I O 103 Inject marker A connected valve enables the inject marker pin 9 The contact is high when the valve is in load position 3 3V and low OV in the inject posi tion On an inject event the contact will go to OV low for 2000 ms It can for example be used to start the integration software when injection is done Valve control In stand alone mode the valve pos
7. Dig Input Device DECADE Elite Number 1 to enable an external start trigger CHAPTER 2 Installation 29 e Incase an ALIAS SINEAS AS 110 auto sampler is used to trigger a run digitally via RS232 LAN USB etc please refer to the corresponding Clarity instruction manual of the specific control module e Type OK and login to your Clarity instrument to connect and the DECADE Elite is ready for use HPLC fluid connections In this section the installation amp priming of all relevant fluidic connections are described to be able to use the DECADE Elite for the analysis of sub stances with U HPLC ECD When working with HPLC solutions and mo bile phases take the following precautions Use proper eye and skin protection when working with solvents The solvents used may be flammable toxic or corrosive Organic sol vents are toxic above a certain concentration Ensure that work areas are always well ventilated Use of open fire in the vicinity of this sys TOXIC HAZARD tem must be strictly prohibited Do not install the system in the 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 DECADE Elite Detector The equipment connected to the system sho
8. In the above example a half scan is programmed sweeping the potential from O 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 time 92 175 0010 DECADE Elite user manual rev 01 A convenient way to record a scanning voltammogram is by direct infusion of analyte in the flow cell using a syringe pump In the figure below a scan ning voltammetry set up is shown a4 a i Antec f Fig 40 DECADE Elite scanning voltammetry set up with a syringe pump The Antec dual syringe infusion pump pn 188 0035 which can be ob tained as accessory has as an advantage that it can be controlled in Elite Dialogue software as well r al gt Elite Dialogue DECADE ELITE See File Tools Options Events Window Help Monitor Detector Events S pump Log Monitor Pump started to infuse Start Stop Control Status Direction Infuse Withdraw Volume 638 347 ul Reset Flow rate 2500 uL min Target vol 0 000 ful Jon Units Syringe ulmin uL hr Diameter 461 mm mL min mL hr Selector vw Fig 41 Programming the Antec infusion pump in Elite Dialogue Chapter 5 Detection and Parameters 93 In the example below a half scan is shown at a flow rate of 10 uL min of a 20 uM Serotonin 5 HT standard in mobile phase zi X
9. Weight Installation Power requirements Mains fuse 10 40 C indoor use only 25 50 C 20 80 RH According to EC directives Emission Group Class A CMETus approved 1 II 2 43 D x 22 W x 44 H cm 16 9 D x 8 7 W x 17 3 H max 14 4 kg 82 lbs without flow cell and column Install upright on flat amp smooth surface keep space under the detector free risk of blocking power supply fan 110 240 VAC 50 60 Hz 260 VA auto sens ing 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 Antec authorized personnel Only use manufacturer supplied fuses For optimum analytical performance it is recommended that the ambient temperature of the laboratory be between 20 25 C and be held constant to within 2 C throughout the entire working day Note that for optimal performance of the oven the oven temperature should be set at least 7 degrees higher as ambient temperature 96 General Operating modes Other mode Sensors Autozero Max current compensa tion Autozero Offset PC control Embedded software Oven Rear panel connectors Analog output DAC Analog output 1 E Digital O HW Programmable I O func tionality Valve control DC mode Range Filter ADF Potential Ec Data Rate Noise 175 0010 DECADE Elite user manu
10. pulse damper column mobile phase etc 175 0010 DECADE Elite user manual rev 01 Dummy cell test External dummy cell An external dummy flow cell pn 250 0040 is shipped with every DECADE Elite instrument for troubleshooting purposes and maintenance checks A successful dummy cell test confirms that the controller including the cell cable functions properly If the result of the noise measurement with the dummy cell is within specs the controller is excluded in a troubleshooting procedure R i C WORK AUX REF RED BLUE BLACK Fig 49 Left schematics of external dummy flow cell Right photo of the external dummy flow cell on 250 0040 The dummy flow cell consists of a resistor R of 300 MOhm and a capaci tor C of 0 47 uF in parallel The current is measured over the resistor ac cording to Ohm s law V I x R hence with a working potential of 800 mV the current drawn will be about 2 67 nA Slight differences as to this ideal value are due to the tolerance of the resistor 1 The capacitor func tions as a noise generator and in fact resembles the capacitance of a well functioning VT 03 flow cell in an ideal U HPLC set up The noise generated via the dummy should be less than 2 pA if the filter of the con troller is set to off provided that the dummy is within the fully closed Fara day shield at the same position as the flow cell Table XI Dummy cell test settings Parameter Setting
11. the flow cell early in the morning by means of a timer Two of the inputs are configurable pin 12 pin 15 cell on and off respectively in the IO menu see previous chapter about the overload output The configuration settings of these inputs are 1 2 3 4 5 and all In case all is selected all cells present in the DECADE Elite will be switched on or off when the corresponding input is triggered Autozero The DECADE Elite has 3 TTL inputs pin 18 20 available to autozero the cell current of the cell s Triggering these inputs enables external acti vation of the auto zero command This function is active only when the I cell is displayed One autozero input is configurable pin 18 in the IO menu see previous chapter about the overload output The configuration settings of this input are 1 2 3 4 5 and all In case all is selected the cell current of all cells present in the DECADE Elite will be zeroed when the input is triggered Start The DECADE Elite has 4 TTL inputs pin 21 24 available to start data acquisition and or start a scan One start input is configurable pin 21 in the IO menu and can be used for example to start the data acquisition of all cells synchronously using only one trigger input when the setting All is selected The manufacturer will not accept any liability for damage direct or indirect caused by connecting this instrument to devices that do n
12. 0 025 Hz 3 0 4 0 5 0 min 6 0 Fig 21 Typical chromatogram with peak widths between 10 100 s Noise in chromatography can come from different sources Pump pulsa tions are typically shown as a very regular noise pattern while electronic 175 0010 DECADE Elite user manual rev 01 noise has a more random character This is illustrated in Fig 22 where a noise trace is shown with an overlay of a 10 and 0 4 Hz sine 20 sine f 10 Hz 10 7 sine f 0 4 Hz 30 31 32 33 34 35 36 37 38 39 40 Fig 22 Typical random noise in chromatography lower trace Both fre quencies 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 collection of more or less random frequencies Low pass noise filters The way noise filters work is by suppressing certain frequencies in the ac quired signal Typically low pass filters allow chromatographic peaks low frequency to pass while high er frequency noise is attenuated No matter how advanced it is impossible to use a low pass filter successfully if there is no difference in frequency of signal and noise Analogue filters are made of hardware from capacitors resistors and am plifiers opamps Digital filters are mathematical routines to process an ac quired signal Traditionally in many detectors for chr
13. 45 nA PUL E 1 0 50V REM compeo r t 252 gt 30 C Remote Control 44 175 0010 DECADE Elite user manual rev 01 SCAN mode SCC Remote Control Chapter 4 DECADE Elite controller 45 CONFIG menu Antec DECADE EC Detector CONFIG PULSE SCAN Pp contrast 20 CONFI G Vout sources DAC fi D1 master no 46 175 0010 DECADE Elite user manual rev 01 DIAG menu 42 1 30 SYS TE Ms 3 80 lt 7 42312415 Chapter 4 DECADE Elite controller 47 Parameters Explanation Type S is status F is function and C is control Parameter Screen Description Type 28 gt 30 C dc stat Displays the actual left value S pulse stat and the pre set oven temperature scan stat right value run AZERO dc stat run Sets the output voltage to 0 V or F pulse stat to the offset voltage Control scan stat Comp off changes to Comp on If cell current exceeds the max compensation a message cell current exceeds max com pensation appears In that case max compensation will be ap plied which may not be the 0 Volt level but higher Boot system Displays boot firmware version S CELL ON dc stat pulse Toggles between cell ON and F OFF stat scan OFF Confirmation is required setup scan Switch cell on off Switching stat on resets the clock to 0 00 Pulse mode pulsation occurs as long as the cell is on irrespective which screen is
14. A 4 B and 8 C pole Bessel filter In Fig 24 it is shown that the number of poles is important a filter behaves more ideal with increasing number of poles In a hardware filter the num ber of poles is the number of filter circuits that are placed in series 0 01yF stage 1 k 0 0990 k 0 040 stage 3 k 0 0834 k 1 023 Fig 25 Analogue 6 pole Bessel filter Chapter 5 Detection and Parameters A digital filter does not have poles but it is characterized by the number of input data points used to calculate a new output data point For example a 9 point digital filter Savitzky Golay is given as 0 090909091 0 168831169 0 255411255 0 168831169 0 090909091 Note that the sum of coefficients is exactly 1 Y n is the output data point X n are input data points Generally spoken the performance of a digital filter improves with more input data points but also more processor capac EN LD SD eae EP 060606061 233766234 233766234 060606061 ity is required for the large number of calculations 175 0010 DECADE Elite user manual rev 01 Applying ADF in chromatography If noise frequencies in LC EC differ from the frequency of the signal noise can be suppressed Using the right filter setting cut off frequency will spe cifically attenuate noise and improve the signal to noise S N ratio No matter how advanced a filter is it is only possible to apply low
15. C Johnson D Dobberpuhl R Roberts and P Vandeberg Review Pulsed amperometric detection of carbohydrates amines and sulphur spe cies 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 at gold and plati num electrodes Anal Chem 62 1990 589A 597A 3 W R LaCourse en D C Johnson Optimization of waveforms for pulsed amperometric detection of carbohydrates following separation by LC Car bohydrate Research 215 1991 159 178 4 W R LaCourse en D C Johnson Optimization of waveforms for pulsed amperometric detection of carbohydrates based on pulsed voltammetry Anal Chem 65 1993 50 55 82 175 0010 DECADE Elite user manual rev 01 Chapter 5 Detection and Parameters 83 CHAPTER 9 Optimization of the working potential Introduction A current voltage I E relationship or voltammogram characterizes an analyte It gives information on the optimum working potential which can be used to improve detection sensitivity and selectivity There are several ways to obtain a voltammogram e A hydrodynamic voltammogram is obtained in the DC mode by running several chromatograms at different working potentials Both peak height and background current are plotted against the working potential A hydrodynamic voltammogram has as an ad vantage that the I E relationship of all analytes of interest can be obtained simultaneously in one se
16. Cell potential 800 mV Oven 35 C stable Filter off Range 1 nA V CHAPTER 12 Troubleshooting 115 Test criteria e cell 2 67 0 05 nA e Noise lt 2 pA The results cell current and noise of the dummy cell test should be within the above mentioned test criteria If the current value Icell and the noise are not within the criteria it is an indication that something could be wrong with the detector hardware Please consult your local representative CAUTION Internal dummy cell The DECADE Elite 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 ex ternal 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 after 5 min stabilisation auto zero is activated and the dummy cell test is ready Noise of the internal dummy cell can be measured at the output As with the external dummy cell the noise should be better than 2 pA Detector settings in the NOISE screen are the same as in the external dummy cell test with the exception of the oven tempera ture Temperature is switched off Please wait stabilizing cell current t ime remaining 05 00 Stop flow test The stop flow test is a basic test to check if the problems are related to ei ther the flow cell
17. Control ii Software Chapter 5 Detection and Parameters 61 Fig 12 Typical configuration for parallel mode detection Two independent HPLC systems with dual channel support from OR 110 DECADE Elite and Clarity software Fig 13 DECADE Elite with 2 columns and 2 SenCell flow cells for parallel detection Navigation in dual cell menu All menus for a dual flow cell system are similar to a single cell system with 2 exceptions First in the top right corner a number is visible which indi cates the active cell in display Toggle with the and buttons between 62 175 0010 DECADE Elite user manual rev 01 sensor boards If the board number does not change it means that the sec ond sensor board is not installed or not properly recognized Second a new status screen is available in dual cell systems which indicates the sta tus of both cells in a single screen However for convenience it is advisa ble to use PC control from the Clarity Chromatography software when working with 2 flow cells Antec ELITE EC Detector PULSE SCAN Fig 14 DECADE Elite main menu top with active cell indicator in top right corner Multi STAT screen showing cell 1 DC mode and cell 2 PULSE mode Parameters Operational parameters are controlled from the SETUP screens in the DECADE II Parameters are filter cell potential and offset Temperature is set in CONFIG menu Fig 15 Selection of parameters in the DC SE
18. Elite Dialogue software and Clarity Chromatography software AUX The DECADE Elite has 4 free programmable TTL outputs AUX1 AUX4 pin 7 10 These contacts are default high 3 3V inactive when active the status is low OV Out of range CURRENT TME Fig 47 Example of a chromatogram were the cell current exceeds the maximum current level and the signal is out of range 108 CAUTION 175 0010 DECADE Elite user manual rev 01 Overload The overload output pin 11 can be used to monitor if the cell current goes out of range during a chromatographic run An Out of range error appears when the cell current Ice exceeds the limit of the current range at which the measurement is performed See figure below It is important to recognize an out of range overload situation because it may lead to erratic results when quantifying analyte concentrations in samples If for instance the cell current goes in out of range during the recording of an analyte peak it can be in most cases easily recognized by a flat top of the peak and a very abrupt transition to the flat top at the edges By default the status of the overload output is high 3 3V When the cell current has the status out of range the overload output will change status to low 0 Volt until the cell current returns to a value within the measure ment range again The overload output pin 11
19. TIT 192 168 5 1 70080 A3 9C 85 40 DECADE II SDC 192 168 5 129 00 80 A3 A3 64 79 ROXY 192 168 5 221 00 80 A3 A3 69 5F ROXY HC Syringe pump Connect Bite via USB 26 175 0010 DECADE Elite user manual rev 01 e When a DECADE Elite is available it is automatically detected and the IP address is shown in the port settings box If not press port scan or type in the default IP address 192 168 5 1 e The pull down field shows all responding devices with their IP and MAC address In case of doubt check the device unique MAC address on the rear panel IO connector e Type OK The instrument will connect and is ready for use Clarity Chromatography software In the case an ALEXYS Analyzer was purchased which includes preconfig ured Clarity software installation files please refer to the installation infor mation shipped with the ALEXYS system otherwise see instruction below to install the software e Open the official installation DVD of the Clarity Chromatography software e For disk space requirements and detailed installation information see the official Clarity Getting Started manual It is available on the installation DVD e Select the INSTALL EXE file and double click it if installation does not automatically start e The software installation wizard will guide you through the installation process including creation of a Clarity menu in the Windows start menu and a Clarity quick start icon on the PC d
20. V CRC 32 Check V Valve Connected Z Temperature 35 sc Digital Inputs Analogue Output pac Digital Outputs Coc cance ee The instrument has a CRC 32 check on the data communication to correct for transmission errors It is advised to have the CRC 32 check switched on For use of the analog start trigger to start a run using an external device open the Digital Inputs and select Sensor 1 start in 28 175 0010 DECADE Elite user manual rev 01 Digital Inputs Connect the relay contact closure of the external device which functions as inject trigger to the digital IO connector on the rear panel of the detector to pin 21 start 1 and pin 25 ground For sensor board 2 or 3 use pin 22 start 2 and pin 23 start 3 re spectively I s Number of Instruments 1 AA Instrument 1 x Instrument 2 3t Instrument 3 Instrument 4 Instrument Type hare Tenet Image for Closed Instrument Image for Opened Instrument Instrument 1 Aa Cel 1 DECADE Elite Thermostat Valve Fraction Collector Auxiliary Device Number Ext Start Dig Input DECADE Elite zj Ready Dig Output DECADE Eite gt oc Cera C re Add the DECADE Elite device to the Clarity instrument by clicking the right arrow button encircled and select Ext Start
21. embedded software controlling the DECADE Elite electronics In this section the installation and configuration of the DECADE Elite is shortly described Note this is by no means a replacement of the installation doc umentation available for the software packages Please refer to this docu mentation for details CHAPTER 2 Installation 25 Ensure you have Administrator access rights in your system before you start with the installation of the software packages Elite Dialogue and Clarity users must have read write access to all software folders and subfolders Elite Dialogue To install the software e Download the latest version of the Elite Dialogue from the Antec website www myantec com register to get access e Double click on the setuo exe file to start the installation wizard e Follow the instructions of the installation wizard for successful installation of the software e Insert the Elite Dialogue license dongle to get full access to the software without dongle it will operate in demo mode e Make sure that the LAN connection is configured and the LAN cable is connected e Power up the detector by means of the mains switch on the rear panel e Start the program Elite Dialogue from the Windows start menu e During start up the Select devices menu will pop up as shown below a Select devices X Select devices Devices Port settings pos V DECADE Bite DECADE Bite 192 168 5 1 v a DECADE II
22. flow cell x 14 Control board SRAM error 20 Checksum error Please contact your local supplier if one of the above errors occur for fur ther instructions In the case the instrument does not power up at all check the following remedies 112 WARNING 175 0010 DECADE Elite user manual rev 01 No detector response Possible cause Remedy No power Check line voltage setting plug in power cord Power switch off Turn this switch ON at the rear panel Faulty fuse Replace fuse Divergent mains voltage Check line voltage Make sure the DECADE Elite is connected to a grounded power source with a line voltage which is within the specified ratings If the DECADE Elite does not respond a fuse in the mains inlet may be blown Furthermore the following messages can be displayed on the LCD screen during a measurement Table X Messages Message Advice 01 Out of range Output is either above 1 0V or below 1 0V Pressing AZERO may give an adequate read out again If the message remains after pressing AZERO the autozero function is unable to com pensate the background cell current Advice use a less sensitive range in the SETUP menu 02 PAD overload Charging current in pulse mode out of range Pressing AZERO may give an adequate read 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 Ou
23. is one of the configurable I O s The configurable I O s can be programmed in the IO menu which is a sub menu of the CONFIG menu By default the overload output is assigned to cell 1 P11 OVLD 1 This means that only when the cell current of cell 1 is out of range the status of the overload output will change to low OV For all other cells present in the DECADE Elite in case of a DCC or TCC version of the instrument an out of range situation will not trigger a re sponse on pin 11 The following options can be selected for the configura tion of pin 11 P11 OVLD 1 Overload output active for cell 1 only P11 OVLD 2 Overload output active for cell 2 only P11 OVLD 3 Overload output active for cell 3 only P11 OVLD 4 Overload output active for cell 4 only P11 OVLD 5 Overload output active for cell 5 only P11 OVLD Overload output inactive P11 OVLD All Overload output active for all cells present CHAPTER 11 Rear panel I O 109 When this option is selected the overload output will be active for all cells present in the DECADE Elite If the cell current of either one of those cells will go Out of range the overload input pin 11 will become active Cell on Cell off The DECADE Elite 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 stabilise
24. network IP address of the LAN network card is now set up for communication with the DECADE Elite 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 DECADE Elite as depicted on the photograph 9 Switch on the DECADE Elite Set the detector temperature to 35 C if a Performance Qualification PQ will be executed or set it to the tempera ture at which your application is running Allow the instrument to stabilize for at least one hour before starting analysis The PQ and OQ are de scribed in separate manuals available on the Antec website 24 175 0010 DECADE Elite user manual rev 01 switch crossover UTP cable Ta standard UTP cables Fig 3 Example of LAN connection between PC and DECADE Elite A di rect connection requires the special crossover UTP cable With a switch any regular UTP cable can be used Software The DECADE Elite can be used in combination with PC control software Currently there are two software packages available for control and data acquisition of the DECADE Elite electrochemical detector e Elite Dialogue software from Antec The Netherlands e Clarity chromatography software Version 6 x and higher from DataApex Czech The Elite Dialogue software is also required for the upload of new firmware
25. or the rest of our U HPLC system Perform the following steps to execute the stop flow test 116 175 0010 DECADE Elite user manual rev 01 e Switch off the U HPLC pump e Disconnect the tubing connection from the column outlet see figure below e Disconnect the outlet tubing from the flow cell e Connect the other end of the tubing of the tubing connected to the inlet of the flow cell to the outlet of the flow cell e The fluidic path of the flow cell is now completely isolated from the rest of the LC system e Record a run without injection of sample to measure evaluate the background cell current Icell and noise e Compare 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 electrochemically active contami nants in the mobile phase column bleeding leaking pulse damper This causes can be systematically eliminated by replacing the mobile phase or disconnecting column or pulse damper etcetera and re evaluate the cell current CHAPTER 12 Troubleshooting 117 Stop Flow Significant drop in Icell Flow cell Contamination in F
26. oven compartment Do not allow flammable and or toxic solvents to accumulate Follow a regulated approved waste Disposal 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 are within An acceptable range and evaluate precision from day to day and run to run Data collected when quality control samples are out of range may not be valid Do not report this data until you are certain that system Performance is acceptable Apart from use of quality control samples we Recommend that you use blanks The blanks will help you assess whether carry over is within an acceptable range and monitor the integrity of your data Using the DECADE Elite in other ways than indicated in the manual or defined by good laboratory practice may result in erratic or unsafe operation 12 175 0010 DECADE Elite user manual rev 01 CHAPTER 1 Introduction 13 CHAPTER 1 Introduction Congratulations on your purchase of the DECADE Elite This detector ena bles you to perform all U HPLC applications using electrochemical detec tion The DECADE Elite includes a highly stable Faraday shielded oven compartment accommodating column and flow cell This flow cell has sur prised
27. procedure or practice which if not adhered to could re sult in damage or destruction of parts or all of the equip ment and or erratic results Do not proceed beyond a cautions sign until the indicated conditions are fully un derstood and met The biohazard sign draws attention to the fact that use of biological materials viral samples may carry a signifi cant health risk The toxic hazard sign draws attention to the fact that use of toxic solvents or samples may carry a significant health risk 175 0010 DECADE Elite user manual rev 01 The attention sign signals relevant information Read this information The note sign signals additional information It provides advice or a suggestion that may support you in using the equipment CHAPTER 1 Table of contents 9 Safety instructions Adhere to the following standard quality control procedures and the follow ing equipment guidelines when using the DECADE Elite detector The fol lowing safety practices are intended to ensure safe operation of the instru ment Working environment amp safety The intended use of the instrument is to detect electroactive substances in liquid samples in combination with a U HPLC system in a GLP approved environment Operators using the system should have the appropriate ed ucation an extensive understanding of GLP rules and be skilled in the art WANES Use this system ONLY for the intended use Use of the system for any other purpose
28. rate of voltage change over time is defined as the scan rate V s Scanning Voltammetry Waveform Working Potential V to ty to Time Seconds Fig 33 Scanning voltammetry potential waveform The current is plotted against the working potential to give a voltammo gram I E curve An example is shown in Fig 34 A difference with a hy drodynamic In scanning voltammetry no HPLC separation is involved The signal is the sum of all EC active substances It takes only a few minutes to con struct a scanning voltammogram This is an advantage especially when a number of analytes have to be characterized However it is a prerequisite to have the pure analyte dissolved 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 Chapter 5 Detection and Parameters 87 As can be seen in both Fig 32 and Fig 34 when the working potential is increased the electrochemical reaction is enhanced hence the signal in creases At a certain potential the I E curve flattens All analyte molecules that reach the working electrode are converted at such a high rate that the analyte supply becomes the limiting factor At the working electrode sur face a stagnant double layer exists where molecular transport takes place by diffusion only Therefore the current at and beyond this potential is called the diffusion limited cur
29. selected Scan mode potential E1 is applied Checksum system Displays checksum S Comp dc stat pulse Toggles between ON and OFF IC stat releases auto zero offset Switches ON if AZERO is pressed Affects auto zero com pensation only not the offset CONFIG main Enters config screen F Contrast config Sets the contrast of display C Cyc scan setup Controls the nature of the cycle C half full and continuous Half means that the cell potential runs from E1 to E2 and stops at E2 Full means that the cell poten tial runs from E1 to E2 and back to E1 and then stops Cont 48 175 0010 DECADE Elite user manual rev 01 Parameter Screen Description Type means that the cell potential runs from E1 to E2 and back to E1 continuously A V Press ing STOP or finishing the cycle sets the potential to E1 DIAG main Enters Diag screen F DISPL test Enters DISP screen for display F test E1 E2 E3 pulse setup2 Controls the cell potential settings C E4 E5 pulse setup3 of the pulse Can be set between 2 50 and 2 50 V with 10 mV steps Can only be set or changed after confirmation with the enter button Ec prog dc only Controls the cell potential canbe C events setup set between 2 50 and 2 50 V dc only dc with 10 mV steps Can only be set setup or changed after confirmation with
30. stat Displays the total duration of one pulse t1 t2 t3 t4 15 t1 12 13 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 Temp config Controls the temperature of the oven Range off 15 60 C se lectable in 1 C steps The oven is stable from 7 C above ambient oven temperature Toven dc setup pulse setup1 Controls the temperature of the oven Range off 15 60 C se lectable in 1 C steps The oven is stable from 7 C above ambient ts pulse setup2 Controls the duration of the sam pling time in the pulse mode The time can be set between 20 ms and maximum t1 60ms with 20 ms increments Tsensor system Displays active temperature sen sor Valve config User confirmation whether a man ual valve is connected to phone jack C on rear panel If present INJ I or INJ L appears in DC Pulse Status screen Vout stat dc pulse scan events setup run noise Displays output signal Vout source config Sets the output source from the analog data output DAC pro cessed digital signal after 16 bit 52 175 0010 DECADE Elite user manual rev 01 Parameter Screen Description Type AD conversion or I E true ana log signal from the I E c
31. the Analog data output 1 V full scale on the rear panel Refer to the documentation of the third party software controlling the A D converter for detailed instruction how to set up a measurement with the A D device Your system is now ready for use The DECADE Elite has been developed for continuous operation For maximum stability it is advised to leave the system ON continuously If preferred the flow cell may be switched OFF at night CHAPTER 3 Maintenance amp Shutdown 37 CHAPTER 3 Maintenance amp Shutdown Maintenance In this paragraph all maintenance is described which can be performed by the end user all other maintenance amp service procedures may only be per formed by authorized service engineers only Periodic check for leakage Perform leak checks on LC tubing flow cells and connections on a daily basis and check if the drain on the bottom of the oven compartment is not blocked or closed Do not allow flammable and or toxic solvents to accu mulate Follow a regulated approved waste disposal program Empty and clean waste container regularly Never dispose of such products through the municipal sewage system Check daily that the mobile phase bottles contain enough mobile phase for the number of analysis planned to be Executed Periodic check of the oven temperature The operator should perform regular checks to verify if the actual oven temperature is in accordance with the set temperature of the DECAD
32. the enter button Ec run dc only Reflects the set cell potential Dis S scan stat dur plays the actual cell potential in ing scanning the scan mode Events dc setup Enter events menu F pulse set upup2 Filt dc setup dc Filter settings RAW 100 Hz Off C DC mode stat prog 10 Hz and 1 Hz to 0 001 Hz cut off frequency in 1 2 5 steps Filt pulse setup Filter settings Off and 0 5 Hz to C PULSE pulse stat 0 001 Hz cut off frequency in 1 2 mode prog 5 steps Fcut off filter coeffi cients based on 1 Hz input fre quency in pulse mode Filt run Reflects the actual filter setting S Firmware system Displays firmware version S Hold run scan stat Toggle holds or resumes execu F resume tion of scan HOLD 0 1 run scan stat Holds or continues execution of F scan Toggles between 1 and 0 Pressing hold again continues scan were it has been hold Chapter 4 DECADE Elite controller 49 Parameter Screen Description Type Ic stat dc Displays the true non compen S pulse scan sated cell current unaffected by events setup auto zero or offset run noise ID1 master config Sets sensor board 1 as master C When this setting is set to yes all parameter settings from sensor board 1 are automatically cop ied transferred to all other sensor boards present IO Config Enter IO menu F INJ I L dc stat pulse Displays o
33. the software shall be entirely at the user s own risk WARNING C Declaration of conformity We Antec Leyden B V Zoeterwoude The Netherlands declare that the product DECADE Elite Electrochemical Detector type 175 and 176 to which this declaration relates is in conformity with the following CE directives Low Voltage Directive 2014 35 EU Safety requirements for electrical equipment for measurement control and laboratory use Part 1 General Requirements 1EC61010 1 3 4 2010 Part 2 010 Particular requirements for laboratory 1EC61010 2 010 3 2014 equipment for the heating of materials EMC Directive 2004 108 EC Electrical equipment for measurement control and laboratory use EMC requirements Part 1 General Requirements IEC61326 1 2 2012 Electromagnetic Compatibility CISPR11 group 1 Class A Group 1 ISM equipment group 1 contains all ISM equipment in which there is intentionally generated and or used conductively coupled radio frequency energy which is necessary for the internal functioning of the equip ment itself Class A equipment is equipment suitable for use in all establishments other than domestic and those directly connected to a low voltage power supply network which supplies buildings used for domestic purposes WEEE directive 2002 96 EC Waste Electrical and Electronic Equipment RoHS directive 2011 65 EU Restriction of Hazardous Substances This device complies wi
34. this way is not pos sible and the chromatography has to be optimized Electrochemical detection differs from most other LC detection methods in that a reaction takes place in the detection cell Due to reaction kinetics an increased temperature speeds up the oxidation reduction reaction How ever this not only holds for the analyte but also for the background current and possible interferences An elevated temperature will therefore not au tomatically lead to a better detection A constant temperature is of para mount importance for a stable baseline and reproducible detection condi tions 4 Er E E E Fig 35 Selectivity in LC EC of compound X and Y is optimised by choos ing the working potential with the largest difference in peak height Chapter 5 Detection and Parameters 89 Electrochemical reactions are pH sensitive Fig 36 For norepinephrine the I E curve is shifted to a lower potential at higher pH When the working potential is high Ez and the signal is diffusion limited an increase in pH will result only in a small increase of the peak height When 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 in creases at a higher pH norepinephrine background 0 E 0 5 E EW 1 0 Fig 36 At a higher pH the I E curve of norepinephrine is shifted to the left Reaction kinetics predict that ele
35. 2 free sodium hydroxide is available from several suppliers as a 50 solution Chapter 5 Detection and Parameters 79 19 2 mol l NaOH pellets are not recommended because of their high CO gt z content Organic modifiers acetonitrile strongly attenuate the signal of most carbohydrates in PAD and are therefore not recommended Pulse settings In PAD of carbohydrates a series of potentials is applied in a continuous cyclic manner During time interval t1 the detection potential is applied The data collection occurs within t1 during time interval ts sampling time The time difference t1 ts is the stabilization time 1 t3 t2 0 0 1 0 2 0 3 0 4 0 5 0 6 time sec Fig 29 Potential steps in pulsed amperometric detection During t1 the detection potential is applied detection occurs during ts Steps t2 t3 and t4 are for regenerating the electrode This process repeats itself continu ously as soon as the cell is on During the next time intervals t2 t4 the electrode is cleaned by reductive and oxidative potential steps Optimization of wave forms LaCourse and Johnson 2 4 have published several papers on optimiza tion of wave forms in PAD Several considerations are important for the choice of the pulse duration Optimization is depending on the working electrode material the sample constituents and the required detection fre quency The impression may arise that the number of variables 5 poten
36. 2 or 3 is switched OFF when active or ALL cells P18 AZERO Programmable input can be con figured that the signal of cell 1 2 or 3 is zeroed when active or ALL cells P21 START Programmable input can be con figured that the data acquisition on sensor board 1 2 or 3 is started when active or on ALL sensor boards Range dc setup dc stat prog pulse setup pulse stat scan setup scan stat Range setting varying from 10 pA to 200 UA full scale in 1 2 and 5 steps In the pulse and scan mode 10 nA to 200 UA full scale can be used C scan setup Scan speed can be set from 1 100 mV s in 1 2 5 steps SPD scan stat Scan speed can be set from 1 100 mV s in 1 2 5 steps START run scan stat In DC and pulse mode toggle be tween STOP and START execu tion of a time file Starts a scan in scan mode STOP run scan stat Scan mode STOP aborts scan and resets cell potential to E1 DC and pulse mode toggle be tween STOP and START to con trol execution of a time file Press ing STOP aborts this run cycle Chapter 4 DECADE Elite controller 51 Parameter Screen Description Type counter Cy is reset to 1 STOP also deactivates the outputs Aux 1 and 2 and Relays 1 and 2 status 0000 and sets the electric valve to load if present SYSTEM diag Enter SYSTEM menu t pulse setup2 pulse
37. A 25 LA 25 LA 25 LA 2 5 HA 2 5 HA 2 5 HA A maximum offset of 50 and 50 in 5 steps can be set For exam ple 20 is a 200 mV offset when the maximum output is 1 0 Volt Polarity 64 PO ALX A HOA 175 0010 DECADE Elite user manual rev 01 The polarity of the output can be inversed Oxidative and reductive anal yses 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 A B 7 315 0 16 0 lt 313 0 e m 311 0 11 0 0 00 2 00 4 00 6 00 0 00 2 00 4 00 6 00 Time min Time min Fig 16 Signal to noise ratio is improved using a filter A vs B The DECADE Elite is equipped with ADF Advanced Digital Filter as a tool to filter the acquired signal and improve the sensitivity of the analysis Sig nal to noise ratio In the next chapter the filter setting is explained includ ing detailed background information about filtering DC mode In the tables below the available filter settings for the DC mode are listed with the corresponding data rate of the output Data rate is expressed as number of data points per second Hz In the DC mode the data rate is not an adjustable parameter but is coupled to th
38. E Elite In case the actual temperature exceeds 70 C switch off the detector and contact the manufacturer or its representatives for service CAUTION Flow cell Check the performance of the detector amp flow cell on a daily basis by eval uating 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 is required on the reference electrode REF of the flow cell If necessary perform mainte nance on the flow cell Detailed information about flow cell maintenance in structions are described in the user manual supplied with the specific flow cell SenCell VT 03 or FlexCell purchased in combination with your de tector 38 CAUTION CAUTION 175 0010 DECADE Elite user manual rev 01 Cleaning In general the DECADE Elite needs very little maintenance The outside of the detector may be cleaned with a non aggressive cleaning liquid Do not use any organic solvents to clean the exterior of the detector because this may lead to damage of the paint layer In case of leakage in the cell cabinet tubing connectors cell column etc remove the spilled mobile phase or other solutions as soon as possible be cause this may damage the paint layer or result in the deposition of salt crusts in case of buffered mobile phases which could block the drain in the bottom
39. TUP screen Temperature is set in CONFIG menu Range Range selection is done in the SETUP or STAT screen in DC PULSE and SCAN mode A number of ranges can be selected the maximum cur rent that can be compensated for using auto zero and offset differs The high sensitivity ranges 10 pA 5 nA have the best noise specifications In fact there is a trade off between best noise specification at sensitive Chapter 5 Detection and Parameters 63 ranges and maximum compensation at the less sensitive ranges This is an inevitable consequence of the tremendous dynamic range that is cov ered by electrochemical detection Table Il DC ranges and maximum compensation Range FS Max comp 200 uA 100 pA 50 LA 20 UA 10 HA 5 HA 2 UA 1 uA 500 nA 200 nA 100 nA 50 nA 2 5 mA 2 5 mA 2 5 mA 2 5 mA 2 5 mA 2 5 mA 25 pA 25 UA 25 pA 25 UA 25 UA 2 5 HA Range FS Max comp 20 nA 10 nA 5 nA 2 nA 1 nA 500 pA 200 pA 100 pA 50 pA 20 pA 10 pA 2 5 HA 2 5 HA 250 nA 250 nA 250 nA 250 nA 250 nA 25 nA 25 nA 25 nA 25 nA In the PULSE and SCAN mode current is much higher than in DC mode Therefore it is not possible to select pA ranges Table III PAD ranges and maximum compensation Range FS Max comp 200 LA 100 LA 50 LA 20 UA 10 pA 5 pA 2 UA 1 uA Offset 2 5 mA 2 5 mA 2 5 mA 2 5 mA 2 5 mA 2 5 mA 25 UA 25 UA Range FS Max comp 500 nA 200 nA 100 nA 50 nA 20 nA 10 n
40. V in the 200 nA range cor responds with an actual cell current of 250 1000 200 nA 50 nA I E output The I E output is the unprocessed analog signal from the I E converter cir cuit This signal is a true analog signal which is as close as practically pos sible to the working electrode WE The maximum output voltage of the I E output is 2 5V under all conditions and is independent of the detec tor range setting The signal in mV is related to the selectable I E resistor 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 fol lowing formula le Ampere Vou Volt Rive Ohm x 10 So for example an I E output signal of 250 mV in the 5 nA range Rve 10 MQ corresponds to an actual cell current of 0 25V 10 x 1E7Q 2 5E 9A 2 5 nA In the table below the I E resistor value is listed for every range setting 106 175 0010 DECADE Elite user manual rev 01 Table VI DECADE Elite I E converter resistors Current Ranges IE Resistor Ohm 10pA 20pA 50pA 100pA 200pA 500pA and 1nA 2nA 5nA 10nA 20nA 50nA 100nA 200nA 500nA 1pA 2uA SHA 10uA 20uA 50A 100A 200A In the detector accessory kit a dedicated Output cable D9 male open 2m pn 250 0128A is supplied It is advised to use this manufacturer sup plied cable for this type of measurements For reference the layout of the analog data out connector i
41. aiting for the START input trigger external or keyboard to start a run 54 175 0010 DECADE Elite user manual rev 01 Parameter Screen Description Type SCROLL prog Scrolls through a time file F Temp events setup Controls the temperature of the oven is stored with time file Tem perature of active time file temper ature overrules other temperature setting after selecting START C Time prog Controls the time to execute a data line in a time file can be set with 0 01 min resolution Maxi mum time is 999 99 min The time to stop the execution of a time file must be programmed by EndCycleTime Valve prog Controls the electrically actuated injector during execution of a time file Forces this valve to load LD or inject INJ Chapter 4 DECADE Elite controller Clarity DECADE Elite control module 55 Full control and data acquisition of the DC and Pulse measurement mode is supported in the Clarity chromatography software amp AN gt P De Gre 2 Copyright C by DataApex Ltd Build Mar 27 2014 S N 011 43477 006 01 http www dataapex com il rsion 5 0 3 180 oe Carty What s new Readme System Fies Method Setup dumey cell 100Hz NERE Select Detector cet r Enabled Dataapex DECADE Elite Detector Method Det Status _ Mai
42. al rev 01 DC PULSE SCAN Lite DC mode only CONFIG DIAG and SERVICE Up to 3 flow cells Lite single cell only triggered by keyboard rear panel TTL or re mote PC control LAN 25 nA 2 5 mA in DC and PULSE mode de pendent on range setting 50 to 50 of max output voltage 5 steps Parametric control and data acquisition via LAN port USB service port Flash technology upgradeable via PC USB 7 C above ambient to 60 C accuracy 0 5 stability 0 1 C accommodates column and flow cell s 1x IEC inlet Mains 1x USB B 1x RJ45 LAN 1x 9 pins sub D Male Valve 1x 9 pins sub D Female Analog output 1x 25 pins sub D Female Digital I O 1 to 1 V full scale via 16 bit D A converter 2 5 to 2 5 V full scale unprocessed I E con verter signal 2x Relay 5x TTL outputs CMOS 3 3V logic 13 TTL inputs programmable 1x GND Cell on Cell off Autozero Start Overload Relay Auxiliary VICI valco 2 pos electrically actuated valve E2CA EHCA via serial cable Manual valve 1x inject marker output 10 pA 200 UA in 1 2 5 increments 10 0 001 Hz in 1 2 5 increments RAW and OFF for unprocessed data 2 50 V to 2 50V with 10 mV increments 1 100 Hz in 1 2 5 increments dependent on filter setting lt 2 pA with dummy cell load of 300 MQ 470 pF in 1 nA range filter off Ec 800mV and temperature of 35 C CHAPTER 10 Specifications DECADE Elite 97 PULSE mode Ran
43. cal detection limit of 100 pmol L or lower for catecholamines is no exception Improving detection limits will al ways be limited by the weakest link in an LC EC system In daily practice a couple of rules must be obeyed to fully exploit the incredible linear dy namic range and low detection limits of an EC detector These are not only hardware related but also refer to mobile phase composition degassing temperature and pH stability and several other issues Mobile phase requirements e Electrochemically clean HPLC grade or better e lon strength 20 200 mmol L e Buffer pH near pKa e In line 0 2 um filter amp degassing of mobile phase e EDTA for trapping of metal ions Mobile phase must be clean as it is in direct contact with the working elec trode in EC detection High purity chemicals including water is a pre requi site In some applications EDTA is added to the mobile phase to traps electrochemically metals such as Fe by forming an inactive complex 32 175 0010 DECADE Elite user manual rev 01 However at higher working potentials typically gt 1 2 V vs salt bridge AgCl ref also EDTA can become electrochemically active and is not recom mended In that case only a passivation step with 15 HNOs is recom mended see installation guide Electrolytes assure contact between 3 electrodes in an electrochemical flow cell Low ion strength destabilises an EC system and noise will in crease Extremely high buffer c
44. ctrochemical detection is mass flow de pendent When the LC flow is stopped in LC EC the analyte will be oxi dized completely and the signal decreases rapidly This means that the flow rate not only affects temporal peak width and analysis time but also peak height Also the background signal is sensitive towards fluctuations in the flow rate Therefore it is important to use a pulse free solvent deliv ery system like in the Antec ALEXYS LC EC analyzer 90 175 0010 DECADE Elite user manual rev 01 Construction of a hydrodynamic voltammogram Before a hydrodynamic voltammogram can be obtained the chromato graphic conditions should be optimized Then the following steps are taken 1 A solution of the analyte at a concentration between 1 100 umol l is prepared in mobile phase 2 The electrochemical detector is stabilized in the DC mode at a high potential After stabilization the background current is read from the display of the detector I cell and the noise is measured 3 The run is 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 derivatization 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 37 5 The peak heights and the background currents are plotted against
45. ds EV time hin t min Fig 42 Scan cycle half of a 20 uM Serotonin in mobile phase at a glassy carbon working electrode Scan speed 20 mV s The following procedure is advised to record scanning voltammogram of analytes e Use a voltammetry set up as shown in figure 48 preferably in combination with the Elite Dialogue software and a syring pump for direct infusion of analyte in the flow cell If Dialogue is not used connect an A D converter to the analog output of the detector to record the cell current Set the A D converter sampling rate to 1 Hz This is the same frequency as the voltage steps during the scan If a higher sampling frequency is chosen a typical stepwise pattern may appear Note that with such set up only I versus t curves can be obtained e Prepare a solution of the pure compound dissolved in preferably the HPLC buffer with a concentration in the range of 10 100 uM e Set the lower E1 and upper potential E2 the scan rate Spd range and scan cycle in the SCAN SETUP menu or under the 94 X axis 175 0010 DECADE Elite user manual rev 01 detector tab in Dialogue Typical scan settings to start with E1 0 mV E2 1000 mV scan speed 10 mV s range 5 A V Optimize the settings if required for your specific compounds Prime a 1 mL glass syringe with analyte solution and install it in the syringe holder of the pump Program the syringe diameter and flow rate in
46. e filter setting except for RAW RAW is special the incoming data are not filtered and a data rate between 1 and 100 Hz can be selected Chapter 5 Detection and Parameters 65 Table IV DC mode filter setting and corresponding data rate Filter setting DC mode Hz Data rate Hz 100 default 50 20 10 5 2 1 100 OFF 0 5 0 2 0 1 0 05 0 02 0 01 0 005 0 002 0 001 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 continuously regenerated by a series of potential steps in a cyclic manner Data is pro cessed differently and the data rate is defined by the total duration of the 5 potential steps in a pulse t1 t2 t3 t4 t5 Usually the duration is be tween 0 5 and 2s data rate between 2 0 5 Hz Filter settings in the pulse mode are therefore between 0 5 and 0 001 Hz and OFF Scan mode has no filter at all 66 175 0010 DECADE Elite user manual rev 01 Chapter 5 Detection and Parameters 67 CHAPTER 6 Noise suppression ADF Introduction Besides for its tremendous linear dynamic range and selectivity electro chemical detection is well known for its very low limits of detection To fur ther improve these detection limits the Antec engineers have developed ADF Advanced Digital Filter and the DECADE Elite has been equ
47. el connected to a recorder or an external A D converter Potential l E converter Voltage clamp Vout LCin LC out AUX Fig 9 Schematic representation of an electrochemical cell with a three electrode configuration Essentially for the oxidation or reduction reaction it would be sufficient to use only two electrodes However the three electrode configuration has several advantages over a two electrode configuration If the working po tential would be applied only over an AUX versus the WE without REF the working potential would continuously change due to polarization effects at the electrodes resulting in highly unstable working conditions If the working potential would be applied only over the REF versus the WE without AUX the working potential would be very well defined However the potential of a REF is only well defined if the current drawn is extremely low pico amperes resulting in a very limited dynamic range A three electrode configuration combines the best of both electrodes The REF stabilizes the working potential and the AUX can supply high cur rents This results in the tremendous dynamic range of a three electrode system Chapter 5 Detection and Parameters 59 Internal organization At the working electrode WE in the electrochemical flow cell the electron transfer takes place due to an oxidation or reduction reaction The result ing electrical current is amplified by the current potent
48. esktop e When asked during the installation fill in the license key code of the Clarity license dongle purchased To be able to control the DECADE Elite you need to have at least the following Clarity modules single instrument SW module the LC control module e Insert the license dongle e Make sure that the LAN connection is configured and the LAN cable is connected e Power up the detector by means of the mains switch on the rear panel e Start Clarity by double clicking on the quick start icon on the desktop e Open the Configuration menu from the system pull down menu in the Clarity main window e Add button bottom left field the DECADE Elite control module listed under detectors CHAPTER 2 Installation 27 The detector set up menu will open see figure on the next page Select LAN communication checkbox and fill in the IP address 192 168 5 1 By clicking Auto detect the instrument will be automatically connected and the number of available sensor boards shown Set the desired heater temperature For valve control check the valve connected box F nl Antec Detector Setup RS232 Communication No port available IP Address 192 168 5 1 Find Device Port 10001 Autodetect Check whether hardware is connected Instrument Type DECADE ELITE z Number of Sensors 1 J Signal 1 Cell 1 Signal 2 Cell 2 Signal 3 Cell 3 Signal 4 Cell 4 Signal 5 Cell 5
49. essing the signal will result in a delay This is inevitable as the mathematics of digital signal processing re quires a number of previous data points to process a new data point The filter characteristic in DSP is often named after the scientist who in vented the mathematics behind the signal processing routine Well known names in this field are Bessel Chebychev Savitsky Golay Hamming and many others Amplitude response plot Amplitude filter block 0 1 2 3 4 5 Frequency Hz Fig 23 Amplitude response plot of an ideal low pass filter with a cut off frequency of 1 Hz There are several ways to describe the filter characteristics An amplitude response plot gives important information on filter behavior Suppose our signal of interest has a frequency between 0 1 Hz and all higher frequen cies are noise An ideal filter is shown in Fig 23 where signal frequencies 72 175 0010 DECADE Elite user manual rev 01 between 0 1 Hz completely pass while frequencies of higher than 1 Hz are completely blocked In practice filters behave a bit different from the ideal situation Amplitude response plot shows a more gradual attenuation profile at higher fre quency This cut off frequency is where the output signal amplitude is 70 of the input signal also known as 3 dB point Amplitude Frequency Hz Fig 24 An amplitude response plot of a low pass filter with a cut off fre quency of 1 Hz Itis a 2
50. f a series of po tential changes This is particularly useful for certain applications where he working electrode is rapidly fouled due to adsorption of insoluble reaction products A well known application area of PAD is the analysis of carbohy drates Fig 28 1 Glucose Lactose v nd So FS fe 9 5 2 5 uw Ww I T T T T T T 1 Minutes Fig 28 Carbohydrate analysis in the pulse mode 78 175 0010 DECADE Elite user manual rev 01 Pulse vs DC mode The pulse mode is quite different from the DC mode Instead of a constant potential a series of potential steps is applied in a cyclic manner The sig nal is sampled during a fraction of the total pulse cycle During the sam pling 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 higher 100 1000 nA than in the DC mode Therefore only nano and microampere ranges are available in the pulse mode Typically the background current is between 0 1 2 pA When the frequency of the data acquisition system integrator is higher than the pulse frequency a typical stepwise pattern may appear in the chromatogram This is called oversampling and these steps are usually only visible after considerable magnification o
51. f the chromatogram It is recommended to keep the data acquisition a 1 Hz High pH of mobile phase In carbohydrate analysis another special consideration has to be taken into account Detection is done in the pulse mode using a flow cell with gold electrode For separation an anion exchange column is used with a mobile phase of around 20 100 mmol L sodium hydroxide The pH is between 12 and 14 This puts some demands on the HPLC system and flow cells After prolonged use of the flow cell with a gold working electrode WE in the pulse mode the gold oxide which is generated at the WE precipitates on the auxiliary electrode AUX This gold oxide coating may electrically isolate the AUX and result in an increase of the noise Cleaning the AUX electrode with metal wool is a way to remove this coating The reference electrodes of the Ag AgCl type are not suitable for carbohydrate analysis Due to silver 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 gt 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 hydroxide should be used since carbonate anions may disturb the ion exchange chromatography It forms CO3 which is a strong modifier The CO
52. ge 10 nA 200 pA in 1 2 5 increments Filter ADF 0 5 0 001 Hz in 1 2 5 increments OFF for unprocessed data Potential Ec 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 SCAN mode Range 10 nA 200 pA 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 98 175 0010 DECADE Elite user manual rev 01 CHAPTER 11 Rear panel I O 99 CHAPTER 11 Rear panel I O In this chapter all rear panel functionality is described The DECADE Elite has besides the mains inlet in total 5 connectors on the rear panel for com munication data output and I O A photo of the rear panel connectors is shown below for reference DIGITAL I O ANALOG DATA O lo gt n V 100 240V FUSE RATING 50 60Hz E 260VA 25AT 250V Fig 44 DECADE Elite rear panel 100 175 0010 DECADE Elite user manual rev 01 USB B connector USB type B connector for serial instrument control over USB for service use only e Based on USB to serial UART interface using the FT232R chip from FTDI Future Technology Devices International Ltd e FT232R is fully compliant with USB 2 0 specifications e Fixed communication baud ra
53. gh salt concentrations in organic solvent first wash out salts with water if necessary CAUTION e Flush and store the system with 50 water acetonitrile or methanol Switch the injector valve between load and inject a few times Make sure all tubing filters etc are flushed so no traces of salt are left that could precipitate and clog the system e Remove the flow cell from the system by disconnecting the inlet and outlet capillary e Open the cell flush with water use some tissues to carefully dry the cell Be careful not to damage the spacer in the case of a VT 03 or FlexCell the SenCell does not have a spacer e 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 KCI solution and close the vial with a cap e Switch off the detector and other LC equipment via the mains switch switch to position 0 on the rear panel 40 175 0010 DECADE Elite user manual rev 01 Chapter 4 DECADE Elite controller 41 CHAPTER 4 DECADE Elite controller Introduction The DECADE Elite has been designed for maximum functionality and ease of use The control of ECD parameters is such that without reading this chapter it should be possible to operate the detector This chapter is in tended as a reference guide in case questions arise during operation The information shown in the numerous screens is
54. gular evenly spaced alterations a sine 400000 300000 200000 100000 0 1 00000 200000 t min 300000 400000 Fig 18 Sine of Fig 17 A full period is 0 41 min 25 s which corresponds to a frequency of 1 25 0 04 Hz Frequency of signal and noise Also a chromatographic peak can be expressed in terms of frequencies The way to determine this frequency is the same The duration of the full peak is measured and expressed in Hz 540000 440000 S 408 lt 340000 240000 140000 40000 f F F F 3 0 3 2 3 4 3 6 3 8 4 0 4 2 4 4 46 48 5 0 Fig 19 Frequency tells how often something happens 1 peak in about 0 25 min 15 s f 1 15 0 07 Hz Chapter 5 Detection and Parameters 69 This is further illustrated by an overlay of the same chromatographic peak with a sine of 0 07 Hz Fig 20 540000 440000 340000 240000 140000 5 0 Fig 20 Overlay of a chromatographic peak with 0 07 Hz sine Typically in chromatography narrow peaks are in front of a chromatogram while peaks with longer retention times get wider As a consequence fre quencies are not constant but vary between 0 1 0 01 Hz which corre sponds to 10 100 s peak width 698000 2000 ADF 598000 498000 398000 298000 198000 98000 10s 0 10 Hz NN 40s
55. h amount of Fe in buffer Remedy Check line voltage setting plug in power cord Turn this switch ON at the rear panel Replace fuse Check line voltage Check connection Check connection Clean WE Remedy Replace buffer do not recycle the buffer Optimise potential if possible use smaller WE diameter Refill with wetted KCI crystals Wait for elution of these very broad peaks Replace column Add EDTA to buffer rinse metal parts with 15 HNOs CHAPTER 12 Troubleshooting 119 Noisy baseline Possible cause Salt bridge in REF not satu rated Air bubble in REF or in cell Slow temperature fluctua tions Fouled WE Leaking REF or cell Remedy Refill with saturated KCI add wetted KCI crystals Remove air bubble continuously degas the mobile phase Isolate detector cell set oven temperature Clean WE Tighten connections with care Decreased sensitivity low S N ratio Possible cause Fouled WE by dirty sam ples Cell potential too low Contaminated buffer high Icen Saturation of output Possible cause Damaged REF Damaged WE Cell incorrectly connected Cell potential too high Base line oscillations Possible cause Malfunctioning pump regu lar pattern Over tightened cell bolts Air bubbles in cell or REF Temperature oscillations Contaminated buffer high Icen Fouled WE Fe in buffer Remedy Clean WE if possible dilute samples Optimise potential Replace buffer do not
56. ial 1 E converter Fig 10 Azero Offset compensation VE converter VE analog output 24 bit ADC 16 bit DAC DAC analog output 1 VFS Digital output LAN Fig 10 DECADE Elite 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 set to a 1 V full scale ana log data output by default Output ADC Also the true analog signal from the I E converter before AD conversion is available via the Analog data output connector This output can be selected in the CONFIG menu by setting the parameter Output I E Dual flow cell control The DECADE Elite electronics are located on 2 different PCB s printed circuit boards The contro board and the sensor board The control board is dedicated to communication with PC LAN and keyboard amp display It has a processor with a so called event handler that takes care of all user commands and hardware interrupts The sensor board is fully dedicated to data acquisition and flow cell control By using this architecture it is possi ble to extend the functionality of the DECADE Elite to more than one flow cell by simply adding a sensor board The control board and other hard ware is prepared for mo
57. ience in the safe handling of toxic and corro sive chemicals and knowledge of the application of fire prevention measures prescribed for laboratories Information on safety practices is provided with your instrument and opera tion manuals Before using your instrument or accessories you must thor oughly read these safety practices This manual is written for laboratory technicians who use the DECADE Elite detector for U HPLC analysis Unskilled improper or careless use of this instrument can create fire hazards or other hazards which can cause death serious injury to personnel or severe damage to equipment and property Observe all relevant safety practices at all times Only use the device for applications that fall within the scope of the specified intended use Else the protective and safety equipment of the device could fail ISO 9001 certified 175 0010 DECADE Elite user manual rev 01 WEEE directive All equipment of Antec Leyden which are subjected to the WEEE directive shipped after August 13 2005 are compliant with the WEEE marking re quirements Such products are labelled with the crossed out wheelie de picted on the left site The symbol on the product indicates that the product must not be disposed as unsorted municipality waste Collection amp recycling information Please ship the instrument back to the manufacturer Antec Leyden the Netherlands at the end of life time of the pr
58. ies Lo e Networking General Connect using Contro You can get IP settings assigned automatically if your network supports fg Fei PCn Gee Faiy sa this capability Otherwise you need to ask your network administrator for the appropriate IP settings This connection uses the following items Obtain an IP address automatically M OM Client for Microsoft Networks Use the following IP address CI Y QoS Packet Scheduler IP address 192 168 5 10 al File and Printer Sharing for Microsoft Networks M Intemet Protocol Version 6 TCP IPv6 Subnet mask 255 255 255 0 F Intemet Protocol Version 4 TCP IPv4 Default gateway Link Layer Topology Discovery Mapper 1 0 Driver Se s Link Layer Topology Discovery Responder Obtain DNS server address automatically N Use the following DNS server addresses Install Uninstal Properties Preferred DNS server Description Transmission Control Protocol Intemet Protocol The default Alternate DNS server wide area network protocol that provides communication across diverse interconnected networks E Validate settings upon exit 4 Open the menu Internet Protocol Version 4 TCP IPv4 menu double click 5 Configure the network IP address and subnet mask as depicted in the screen dump below IP 192 168 5 10 Subnet mask 255 255 255 0 Gate way and DNS fields are not filled 6 Close the menu s by clicking the OK buttons The
59. ipped with it as a standard The improvement factor in signal to noise S N ratio depends on the frequency relation of signal and baseline noise S N im provements from a factor 5 up to more than 100 have been obtained To understand how a digital filter works first the importance of frequencies in chromatographic analysis will be explained Then we will look at peak width filter settings cut off frequency amplitude response plots and finally at a few chromatograms before and after applying ADF Frequency A scientific definition of frequency is the number of completed alterations 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 Alternatively the counts can run in an irregular manner within the specified unit of time If the latter happens frequencies would vary if broken down into t smaller units of time In the example of Fig 17 a signal is shown with a frequency of 12 altera tions in 5 minutes To express its frequency in a more scientific way a full period is precisely determined and expressed in Hertz or s It is a sine wave with a frequency of 0 04 Hz Fig 78 68 175 0010 DECADE Elite user manual rev 01 400000 SALAD SVT TTT 300000 o amg 400000 Fig 17 Example of a signal with re
60. is shipped with a complete set of dedicated tubing assemblies LC connec tion kits tailor made for the application the analyzer was specifically de fined for Please note that the tubing length length L below required to make a good connection differs for each brand of connection If length L is not cor rect this will result in faulty peaks and carry over Essentially when you create a connection the ferrule on the tubing is compressed into the valve to make sure that the connection is leak tight Take the following into ac count 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 e Part of the tubing may end up in the connection port internals e Internals of the port may be damaged ferrule does not seal the connection CHAPTER 2 Installation 31 If Lis too short this may result in e Leakage e Dead volume at the end of the ferrule a mixing chamber tubing does not touch end of the connection resulting in dead volume Every ferrule type needs an appropriate length of tubing for connecting it to the type of connection port depending on the depth of the connection port Refer to the information provided by the manufacturer for specific infor mation Mobile Phase Electrochemical detection is a sensitive detection technique characterized by extremely low detection limits A typi
61. it must be flushed away immedi ately Provide equipment such eye wash stations and safety showers as close to system as possible Use proper eye and skin protection when working with solvents Additional safety requirements or protection may be necessary depending on the chemicals used in combination with this equipment Make sure that you understand the hazards associated with the chemicals used and take appropriate measures with regards to safety and protection Sample containers vials should be sealed to minimize any risks related to solvent vapor CHAPTER 1 BIOHAZARD CAUTION WARNING Table of contents 11 Biological Hazard When you analyze biological fluids you need possible precautions and treat all specimens as potentially infectious Always wear protective And gloves when handling toxic or biologically infectious samples to pre vent bio hazards or hazards while working with the DECADE Elite If nec essary the instrument must be decontaminated before decommissioning or shipment of the instrument for repair to Antec or its representatives When shipped to Antec every instrument has to be accompanied with a decon tamination form which should be completely filled in and signed by the cus tomer Without this decontamination form the instrument will not be pro cessed by Antec either repaired or disposed Waste disposal Perform periodic leak checks on LC tubing and connections Do not close or block the drain in the
62. ition can be controlled via the F5 func tion button in the STAT screen For example see the screen dumps of the STAT screen for a DECADE Elite SCC top and TCC detector INJ L rep resents the LOAD position position A on the actuator control module 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 Make sure that the valve flow path is connected correctly and LOAD corresponds with position A and INJECT with position B See example in the figure below for a 6 port valve POSITION A POSITION B SAMPLE IN SAMPLE IN VENT y VENT WASTE WASTE gt COLUMN gt COLUMN SAMPLE SAMPLE LOOP LOOP U PUMP Fig 45 Example of the valve flow path of a 6 port valve 104 175 0010 DECADE Elite user manual rev 01 In the case of a manual valve the information above the F5 button INJ L in the STAT screen shows the status position of the valve only no valve control is possible with this type of valve For information about valve control of the electrically actuated VICI valves in the Clarity chromatography software or Elite Dialogue please refer to the corresponding user manuals ANALOG DATA connector The DECADE Elite is equipped with an analog data output connector to provide the measured signals in millivolts mV for users who work stand alone without PC control with the instrument The ANALOG DATA c
63. ld up of static electricity The power source should exhibit minimal power transients and fluctuations If necessary connect to a filtered mains socket WARNING RISK OF FIRE REPLACE FUSE AS MARKED AVERTISSEMENT RISQUE DE FEU REMPLACEZ LE FUSIBLE COMME INDIQU V 100 240V FUSE RATING 50 60Hz 260VA 2 5AT 250V Replace blown fuses with fuses of proper type and rating as indicated on the rear panel and as listed in the list of accessories and spares appendix D The fuse holder is integrated in the mains connector Ensure that the instrument is never put in operation with fuses of a different type This could cause fire Only use manufacturer supplied I O cable s to connect with other devices Thoroughly connect shielding to common Manufac turer will not accept any liability for damage direct or indirect caused by connecting this instrument to devices and with cables which do not meet relevant safety standards Solvents The solvents used may be flammable toxic or corrosive The room in which the system is installed should be well ventilated to prevent that sol vent 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 sys tem in the same room with any other equipment that emits or could poten tially emit sparks Provide protective equipment near the instrument when solvent gets into the eyes or on the skin
64. ll outlet tubing connection from cell to waste make sure that the outlet is positioned on the top side to prevent entrapped air bubbles 4 WE contact red 5 AUX contact blue 6 REF contact black Top right electrical connections of WE red con nector and AUX electrode blue connector Bottom right SenCell with SaltBridge reference electrode 9 Before switching ON the flow cell make sure that the mobile phase contains sufficient electrolyte buffer ions A stable baseline will never be obtained if the cell is switched ON with only 36 175 0010 DECADE Elite user manual rev 01 water or another non conducting mobile phase Also be sure that no air bubbles are trapped in the flow cell 10 The outlet tubing from the flow cell 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 83 for optimization of the potential switch ON the flow cell and allow the system to stabilize for approximately 30 min A good stabilization curve shows a mono exponential decline without jumps and or spikes 12 Connect the DECADE Elite to the PC control software Clarity or Elite Dialogue In the case the instrument is used stand alone connect an external A D converter or recorder to
65. low path Inspection Service Mobile Phase Replace Flow cell Column Pulse damper Tubing Filter Pump Noise If a significant drop of the noise only is observed it could be signa ture of for example pump problem check valves air in pump head com pressibility issues or leaking seals No Yes Significant drop in Noise Flow cell Check Pump Pulse damper Replace Flow cell Air bubbles Contamination see Icell In case no significant drop in noise or cell current is observed service or replace the cell In case you still cannot solve the problem contact your local representative 118 175 0010 DECADE Elite user manual rev 01 Please bear in mind that analytical problems may also be caused by exter nal Influences like temperature unstable samples Make sure that the applica tion 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 fur ther help No detector response Possible cause No power Power switch off Faulty fuse Divergent mains voltage Cell disconnected or switched off Output disconnected Fouled WE High cell current Possible cause Contaminated buffer High WE potential Salt bridge in REF not satu rated Retained peaks from previ ous runs Column is bleeding Hig
66. ments e Free LAN port onboard PCI PCI express or PCI X e Free USB port required for FW updates e Microsoft Windows 7 Operating System or higher Detailed requirements for use of the DECADE Elite in combination with Clarity chromatography software are listed in document 195 7000 Com puter requirements which can be downloaded from the Antec website www myantec com Laboratory requirements Your instrument is intended for indoor use only in an industrial or commer cial environment EN55011 group 1 class A ISM equipment It is suitable for the following categories Installation category II Pollution degree 2 equipment class I Table I Environmental specifications Parameter Requirement Operating temperature 10 40 C 50 104 F Operating humidity 20 80 non condensing 18 175 0010 DECADE Elite user manual rev 01 For optimum analytical performance it is recommended that the ambient temperature of the laboratory be between 20 25 C and be held constant to within 2 C throughout the entire working day Note that for optimal temperature stability of the cell cabinet the oven temperature should be set to a temperature at least 7 degrees higher as ambient temperature Requirements for the laboratory bench on which the instrument will be in stalled e Stabile clean flat and smooth surface e Enough mechanical strength to hold at least the weight of the detector a full dressed detector
67. n Output Time Table 7 Cell On Mode Pulse Mi Sensor Rate a pulse duration 2 00 Hz E1 0 1 M ti 0 1 s E2 0 5 M t2 0 1 i E 03 M 8 01 Il ts 20 gt my E4 0 5 M t4 0 1 s E5 0 3 M t5 01 s L Status Not Ready Cell off Method has not been sent Fromoet i Event Table Measurement Acquisition Integration Calculation Advanced Se a ee en kh Fig 8 DECADE Elite method window in Clarity On the main tab the main measurement conditions can be set controlled measurement mode amp po tential settings Within a Clarity method it is possible to execute timed event by program ming time lines under the Time table tab Running a Time table enables time based automated parametric control of the electrochemical detection ECD during a run This is particularly useful when during a run settings have to be changed such as the sensitivity auto zero or control of external equipment The SCAN mode is not supported in Clarity Scanning Voltammetry is supported in the Elite Dialogue software 56 175 0010 DECADE Elite user manual rev 01 Chapter 5 Detection and Parameters 57 CHAPTER 5 Detection and Parameters Introduction One of the characteristics of electrochemical detection is its tremendous dynamic range In amperometric detection peak heights may vary from mi cro amperes down to the pico ampere range The DECADE Elite covers such a wide range from 200 LA d
68. n can be reached easily to disconnect it from the mains power by removing the Mains power cable Do not block the ventilation holes at the back of the instrument and do not block the fan located at the bottom of the detector Fig 2 Blocking the fan will impair the cooling capability of the power supply Do not place object instruments on top of the DECADE Elite Objects can be placed on any side of the detector however make sure these objects are placed at a distance of 5 cm from the DECADE Elite if objects are placed at only one side of the instrument and 10 cm from the DECADE Elite if objects are placed on more than one side of the instruments Mains connection CAUTION ELECTRICAL SHOCK Check that fuses and voltage range on the rear side of the instrument match that for the power outlet to be used This device complies with the requirements of EN 55011 group 1 class A ISM equipment In a domestic environment this product may cause radio interference in which case the user may be required to take adequate measures on his own expenses Understanding power surges Power surges line spikes and transient energy sources can impede instrument operation Ensure that the instrument s electrical supply is adequately protected from these conditions and properly grounded If necessary connect the instrument to a filtered mains inlet Power supply and protective earth The system must be connected to a suitable mains
69. nce a one time degassing step of the HPLC buffer is almost never sufficient If the DECADE II 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 Helium and the use of stainless steel tubing impermeable for oxygen 8 Consult your flow cell manual for detailed information about the installation of the flow cell See figure 11 on the next page for reference Connect the flow cell to the corresponding cell connector in the oven compartment All cell connectors are marked with a label for identification In case of a DECADE Elite CHAPTER 2 Installation 35 SCC connect the flow cell to the cell connector on the left side marked Cell 1 The cell connector inside the oven compartment is ESD sensitive Make sure that the flow cell is OFF when removing or connecting the cell cable Never switch ON the flow cell when 1 the cell cable is not correctly connected 2 the cell is only partly or not at all filled with mobile phase 3 the outside of the flow cell is wet particularly the part between the auxiliary and working electrode connection because substantial damage to the working electrode or electronics may occur Fig 5 Left SenCell with ISAAC reference mounted under an angle of ap proximately 45 in the detector 1 Cell clamp 2 Cell Inlet tubing connec tion from column to cell 3 Ce
70. oduct The manufac turer will take care of the proper disposal and recycling of the instru ment at its facilities Shipping address for the end of life products Antec Leyden B V Industrieweg 12 2382NV Zoeterwoude The Netherlands In case of questions or if further information is required about the collec tion amp recycling procedure please contact your local distributor ROHS directive The DECADE Elite is ROHS compliant and in conformity with Directive 2011 65 EU Restricted use of Hazardous Substances in electrical and electronic Equipment ROHS Antec Leyden is an ISO 9001 2008 certified company CHAPTER 1 Warning Symbols Table of contents The following symbols are used in this guide gt ELECTRICAL SHOCK gt WARNING gt CAUTION BIOHAZARD TOXIC HAZARD This sign warns about the risk of electric shock It calls attention to a procedure or practice which if not adhered to could result in loss of life by electrocution Do not pro ceed beyond a danger sign until the indicated conditions are fully understood and met The warning sign denotes a hazard It calls attention to a procedure or practice which if not adhered to could re sult in severe injury loss of life or damage or destruction of parts or all of the equipment Do not proceed beyond a warning sign until the indicated conditions are fully under stood and met The caution sign denotes a hazard It calls attention to a
71. of the cell cabinet Remove any dust on the protective screens that cover the fans in the oven compartment Replacement of fuses Replace blown fuses with fuses of proper type and rating as stipulated on the rear panel and specified in the installation section of this manual The fuse holder is integrated in the mains connector Ensure that the instru ment is never put in operation with fuses of a different type This could cause fire V 100 240 FUSE RATING WARNING RISK OF FIRE REPLACE FUSE AS MARKED AVERTISSEMENT RISQUE DE FEU 2 5AT 250V REMPLACEZ LE FUSIBLE COMME INDIQU In case the fuses blow out repetitively contact Antec or its representatives for instructions and or service of the instrument CHAPTER 3 Maintenance amp Shutdown 39 Shutting down the system There are a couple of steps to take to switch off an LC system with electro chemical detector for a longer period of time Shutting down is not different from most other HPLC systems Perform the following procedure e Switch off the flow cell using the keyboard standalone or via the software Clarity or Elite Dialogue e Check the column s documentation for the appropriate storage liquid apply this and make sure the column is properly flushed A reversed phase C18 column is usually stored with 50 Acetonitrile water e Take out the column mount the corresponding end caps and store the column in an appropriate place Avoid precipitation of hi
72. omatography an analogue low pass filter is applied rise time filter A passive RC filter consists of resistors and capacitors An active higher order filter can be considered as a num ber of these RC filters in series In a 4 order filter the signal coming from the first filter is filtered again in a second third and fourth filter During these steps loss of signal occurs simply because of all the resistors that are applied Operational amplifiers which are active components are ap plied in each stage to restore the signal to its original value With the availability of powerful processors digital signal processing has become an excellent alternative for hardware filters In its most simple Chapter 5 Detection and Parameters 71 form a running 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 80 2180 x 81 x 82 x 83 x 84 5 Each input data point has the same weighting factor of 1 5 In more ad vanced digital signal processing a more complicated equation is used to calculate the output data point y n yin agx n a x n 1 a x n 2 a x a 3 In contrast to the previous equation each data point has a different weighting factor a Sum of these weighting factors ao n will always be 1 Characteristic of noise filters is that proc
73. on nector D9 female can either be connected to an X Y recorder or A D con verter The ANALOG DATA output can supply either a non manipulated signal directly from the I E converter or the processed data signal by the CPU of the DECADE Elite The type of output can be selected from the CONFIG screen by setting the parameter Vout source to either DAC or I E i contrast vout source Di master VE analog output Flow cell O DAC analog output 1 VFS VE converter CHAPTER 11 Rear panel I O 105 Fig 46 Top CONFIG screen Bottom DECADE Elite signal processing from electrochemical flow cell to output R is a selectable IE resistor of 100M 10M 1M 100K or 1K in the I E converter circuit DAC output The DAC output is the processed signal by the DECADE Elite s CPU and is identical to that of the digital cell current signal obtain via data acquisi tion over LAN using the Elite Dialogue or the Clarity Chromatography soft ware The signal in mV from this output is directly related to the range setting of the DECADE Elite So for example in the case the 200 UA measurement range is selected Range 200 HA the DAC signal on the analog data connector corresponds with 1000 mV 200 HA and 1000 mV 200 UA so 1V full scale To convert the signal in mV to the actual cell cur rent in nA use the following calculation Ic nA Vout mV x Range setting nA 1000 mV So for example a signal on the output of 250 m
74. oncentrations cause problems of salt for mation Therefore concentrations between 20 and 200 mmol L are recom mended 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 are phosphate carbonate acetate and citrate Modifiers such as methanol propanol and acetonitril can be used without problems in DC amperometry but not in pulsed amperometric detection as peaks are strongly attenuated In our experience the quality and expiration date of or ganic modifiers can be a problem resulting in increased noise levels Metal inlet frits in mobile phase bottles are not advised as metal ions are introduced that increase baseline noise Instead 0 2 um nylon inlet filters are recommended Vacuum degassing Considerable amounts of the gases N2 O2 and CO2 may be dissolved in HPLC mobile phases Whenever the temperature changes solvents are mixed or a pressure reduction occurs these gases may show up as very small air bubbles To avoid noisy baselines an in line vacuum degasser has been integrated in the ALEXYS analyzer ROXY EC LC system It has a low dead volume less than 0 5 mL and high de gassing efficiency The advantage over helium degassing is that the de gasser does not change the mobile phase composition Helium degassing Degassing using helium is an effective and universally applicable method but only recommended when working in reduc
75. onverter Time file commands Parameter Screen Description Type ADD prog Adds the active data line to the time file Confirmation is asked for if an existing time is overwritten As time 0 00 always exists changing this time results in an overwrite warning Azero prog Controls auto zero which can be programmed in a time file Tog gles between set and not Cyc run Displays the cycle counter If a time file has to be executed more than once Cycles gt 1 this is the number of times a time file has been started RESET external or QUIT sets Cy to 1 and returns to EVENTS SETUP screen Cycles events setup Controls the number of times a time file has to be repeated This number can be 1 999 or continu ous DEL prog Deletes the current data line from the time file Deleting time 00 00 results in deleting the complete time file Confirmation is required DEL prog Deletes the current data line from the time file Deleting time 00 00 results in deleting the complete time file Confirmation is required EVENTS dc setup pulse setup2 Enters EVENTS EVENTS SETUP screen for editing and running a time file EndCycle prog Enters a screen to set EndCycleTime Controls duration of a time file max 999 99 min When this time is reached the ex ecution of the time file stops If Chapter 4
76. or these and other DECADE Elite parts or flow cells contact your local supplier 122 175 0010 DECADE Elite user manual rev 01 Index Index 123 ADF 67 Amplitude response 71 auxiliary electrode 57 Buffer pH 31 buttons DECADE II keyboard 41 cleaning of detector 38 compensation 62 cut off frequency 72 degassing mobile phase 34 digital filter 70 DSP 71 dual flow cell control 59 dual flow cell navigation 61 dummy cell external 114 internal 115 E1 control 48 E2 control 48 EDTA 31 electrochemical reaction 84 filter 48 64 70 frequency 67 fuses replacement of 10 38 I E converter 57 59 I E relationship 83 inject marker trigger 103 installation DECADE Il 17 121 integrator connection of trigger 103 lon strength 31 keyboard DECADE II 41 maintenance cleaning of detector 38 maximum compensation 62 messages 111 112 mobile phase requirements 31 noise 34 36 78 90 119 noise filters 70 offset 63 output 124 175 0010 DECADE Elite user manual rev 01 frequency 80 pulse mode 80 passivation with nitric acid 34 polarity 63 potential optimisation 83 pulse mode characteristics 78 literature references 81 optimisation 79 output frequency 80 potential settings 79 working electrode 80 pulsed amperometric detection 77 range 62 reference electrode 57 rotor Tefzel 78 rotor Vespel 78 S N ratio 90 119 scan mode example 91 scanning voltammogram 86 specifica
77. ot meet the relevant safety standards 110 175 0010 DECADE Elite user manual rev 01 Table VIII DIGITAL I O connector layout Pin Type I O Function Configurable Description default I O 1 2 3 Relay 1 Relay 1 Contact between 3 com mon and 1 default acti vated by time file Outp 0100 or RS232 command 47 4 5 6 Relay 2 Relay 2 Contact between 6 com mon and 4 default acti vated by time file Outp 0001 or RS232 command 47 7 TTLOUT AUX1 Free programmable TTL output activated by time file Outp 0001 RS232 or com mand 47 8 TTLOUT AUX2 Free programmable TTL output activated by time file Outp 0010 RS232 or com mand 47 9 TTLOUT AUX3 Free programmable TTL output activated by RS232 command 47 10 TTLOUT AUX 4 Free programmable TTL output activated by RS232 command 47 11 TTL OUT Overload A 1 5 Active in case of signal over load 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 cell on 15 TTL IN Cell off 1 A 1 5 Trigger to switch the cell off 16 TTL IN Cell off 2 Trigger to switch the cell off 17 TTL IN Cell off 3 Trigger to switch the cell off 18 TTL IN Autozero 1 A 1 5 Trigger to zero null the cell current compensation Ic 19 TTLIN Autozero 2 Trigger to zero null the cell current compensation Ic 20 TTLIN Au
78. oting 111 Instrument errors 111 Analytical troubleshooting 113 Dummy cell test 114 Stop flow test 115 Detector accessories 121 Index 122 CHAPTER 1 WARNING Table of contents 5 Intended use The DECADE Elite Electrochemical Detector is used in combination with Ultra High Performance Liquid Chromatography for the electrochemical detection of suitable analytes in liquid samples With this technique the amount of electroactive substances in mobile phase can be quantified The instrument can be used for the chromatographic analysis of a wide range of electroactive analytes in the fields of for example Bioanalytical analyses Food analyses Pharmaceutical analyses Environmental analyses For research purposes only While clinical applications may be shown this instrument is not tested by the manufacturer to comply with the In Vitro Diagnostics Directive Operation of an electrochemical detector can involve the use of hazardous materials including corrosive fluids and flammable liquids The instrument should only be operated by users with the following expertise Completed degree as chemical laboratory technician or compara ble vocational training e Fundamental knowledge of liquid chromatography Participation in an installation of the system performed by the man ufacturer or a company authorized by the manufacturer and suitable train ing on the system and chromatography software Knowledge and exper
79. own to 10 pA full scale without being lim ited by electronic noise For this reason the DECADE Elite is equipped with a 24 bit ADC and 16 bit DAC for analogue data output One of the key features is that data can be sampled with data collection rates up to 100 Hz 100 pts sec in DC mode which assures that fast peak responses typical for UHPLC can be detected with sufficient resolution Three electrode configuration The circuitry of the DECADE Elite detector is designed for operation with electrochemical flow cells with a three electrode configuration Fig 9 The working potential is set between the working electrode WE and the auxil iary 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 the electrochemical reaction takes place i e electrons are transferred at the WE This results in an electrical current to the I E converter which is a special type of operational amplifier The output voltage of the I E converter is digitized in the instru ment by means of a 24 bit A D converter and processed and the resulting output current Ic can be acquired digitally by PC control software Elite Di alogue or Clarity or analog using the Analog Data output on the rear 175 0010 DECADE Elite user manual rev 01 pan
80. pass filter ing if noise frequencies are higher than the frequency of the signal 400000 300000 200000 100000 0 400000 300000 200000 100000 04 400000 300000 200000 100000 04 0 Fig 26 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 0 005 Hz whereas wider peaks show hardly any deformation see peak at t 13 min Attenu ation of noise is shown in the close up on the right Prerequisite for a good noise filter for data acquisition in liquid chromatog raphy is that it improves the S N ratio without significant distortion of the Chapter 5 Detection and Parameters 75 signal of interest This is particularly difficult if the frequency of the signal is close to the frequency of the noise The DECADE Elite has a number of filter settings to optimize for best pos sible signal to noise ratio The width of the peaks of interest is important because wider peaks allow stronger filter settings simply because of the lower frequency of such peaks Advised filter setting to start further optimi zation is given as Filter setting 1 2 peak width So at a 10 s peak width a 0 05 Hz filter setting is advised If peaks are 50 s a 0 01 Hz filter is advised to start with Note that if a chromatogram has in teresting peaks of 10
81. power supply with a correctly installed protective earth conductor Never use the system without a properly connected protective earth conductor CHAPTER 2 Installation 21 Leave the instrument powered off until specifically mentioned in the proce dure below PC connection Follow the instructions in this paragraph when the instrument is used with PC control over LAN using the Elite Dialogue or Clarity Chromatography Software This section can be skipped if the instrument is used stand alone To be able to communicate over LAN a computer is required with a free PCI PCI Express or PCI X LAN port The DECADE Elite has a fixed IP address 192 168 5 1 with subnet mask 255 255 255 0 Gateway and DNS are not filled in The instrument is standard delivered with a special crossover LAN UTP cable pn 250 0170 which is part of the DECADE Elite accessory kit pn 175 0200 If the instrument is delivered as a part of a complete ALEXYS U HPLC ECD system an additional LAN connectivity kit on 250 0180 which consist of a set of network cables and a pre configured broadband router is available To insure stable and error free communication only use the manufacturer supplied LAN connectivity kit to connect the DECADE Elite to LAN Create a small dedicated local area network to connect the DECADE Elite to the PC Do not connect the DECADE Elite over a company Local Area Network If needed a second network adapter with CAUTION a diffe
82. presented in alphabeti cal order For each item an explanation is given together with the item s nature and the screen s of appearance The nature of an item can be 1 Control parameters with a cursor box O 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 top right corner of each screen the name of the present screen is displayed If available the bottom left function button displays a previous screen and the bottom right one the next screen Vout U Ic 2 661nA DCH BRange inf QEc 8 V STAT OFilt OFF OCome 0N 35935 C 003 30 MARK AZERO NEXT PREV CELL ON Fig 6 DECADE Elite keyboard The cursor is on Range which allows changes using the value buttons and The Enter button is only used to confirm changes in potential Ec and range 42 175 0010 DECADE Elite user manual rev 01 Overview of DECADE Elite screens DC mode SCC 23 45nA 0 50V off 25 gt 30 Remote Control tec 2 EC Detect sc u 0 05 23 45nA Range 50nA E c 0 50V STAT Fiut 002Hz a k pe 26 gt 30 G 211 20 RE CELL AZERO NE mM Chapter 4 DECADE Elite controller 43 Pulse mode Ie 423
83. r switches the position F S stat of the injection valve toggles be tween inject 1 and load L If a manual injector with position sen sor is applied it echoes the posi tion of the injector If an electri cally actuated injector is used op tional it is possible to switch the injector with this function button KEYB test Enters KEYB screen for key F board test Press 2x F1 to leave MARK dc stat pulse Triggers a marker signal on out F stat put MaxComp dc setup Maximum cell current that canbe S pulse setup1 compensated for using auto zero Next several Enter next screen F screens NOISE test Enters NOISE screen for perfor F mance test Offs dc setup dc Percentage offset can be set be C stat prog tween 50 and 50 pulse setup pulse stat scan setup scan stat POLAR dc setup Inverts output polarity toggle be F pulse setup2 tween and Requires confirma tion PREV several Return to previous screen F screens 50 175 0010 DECADE Elite user manual rev 01 Parameter Screen Description Type P114 OVLD IO Programmable output can be configured that the overload OVLD signal of cell 1 2 or 3 only is present on pin 11 when active or ALL cells C P12 C ON Programmable input can be con figured that only cell 1 2 or 3 is switched ON when active or ALL cells P15 C OFF Programmable input can be con figured that only cell 1
84. re than one sensor board Typically a two flow cell configuration can be used in serial or parallel mode detection 175 0010 DECADE Elite user manual rev 01 Serial mode detection In serial mode one LC system is used with 2 flow cells in series For data acquisition 2 data channels are applied with the same time base Serial mode detection is especially suitable for OX RED or RED OX applications examples are analysis of vitamin K and nitro tyrosine using micro HPLC The first flow cell is a FlexCell that converts the analyte of interest in a de tectable substance The second flow cell is a SenCell or VT 03 cell which is used for detection Note that it is necessary to work with micro HPLC be cause the conversion rate of the reactor cell is too small when using stand ard HPLC Clarity instrument 1 Channel f Clarity instrument 1 Channel 1 DECADE Elite 22 i i with i Clarity Chromatography OR 110 3 Dual Cell Control Software Fig 11 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 parallel mode 2 HPLC systems are used with 2 flow cells In fact the DECADE Elite is operated as if 2 independent detectors are in one hous ing Clarity instrument 1 Channel 1 Clarity instrument 2 Channel 1 DECADE Elite with i Clarity Chromatography a ond i Dual Cell
85. recycle the buffer Remedy Check with spare REF replace if necessary Replace cell block Check connections REF black WE red AUX blue Optimise cell potential Remedy Check pump seals valves Adjust cell bolts check pump pressure Maintenance REF Set oven temperature Replace buffer do not recycle the buffer Clean WE Add EDTA passivate metal parts with HNO3 120 175 0010 DECADE Elite user manual rev 01 CHAPTER 13 CHAPTER 13 Detector accessories 121 Detector accessories The electrochemical detector is shipped together with a number of parts The listing in Table below may not be complete see check list of delivery for complete listing Table XII Accessory kit on 175 0200 DECADE Elite detector Part number 250 0040 Component External dummy flow cell 250 0107 250 0102 250 0113 250 0170 250 0175 250 0131 250 0128A 250 0116 250 0118 250 0126A Column clamp 12 mm Cell clamp kit 40 mm Fuse 2 5 AT 250 V LAN UTP cable crossed 3 meter USB cable A B 3 meter I O cable D25 male open 2m Output cable D9 male open 2m Mains cable Europe Mains cable USA Elite cell cable for the VT 03 flow cell pn 250 0102 Cell clamp kit 40 mm is available in the accessory lit For a Vici Valco electrically actuated 2 position valve with an E2CA EHCA actuator a serial cable is available for control via the DECADE Elite detector pn 250 0190 Serial valve cable Valco 2m F
86. rent A 10 _ B 5 a 2 0 0 0 0 4 0 8 1 2 Ecell V Fig 34 Scanning voltammetry of 1 0 umol l norepinephrine A at a glassy carbon working electrode at a scan speed of 10 mV s Scan B is the blank solvent With respect to sensitivity a high working potential is important However at higher working potentials more analytes are detectable So as to selec tivity a low working potential will be favorable Working at a potential on the slope of the I E curve will result in less repro ducibility in HPLC A small fluctuation in the applied potential or any change in the system like for instance a pH change may result in a differ ences in measured peak height In practice the choice of the working po tential is a compromise between sensitivity selectivity and reproducibility In the example of Fig 32 a working potential E1 of 0 8 V is chosen 175 0010 DECADE Elite user manual rev 01 Optimization using a voltammogram Sometimes when interfering peaks appear in the chromatogram it is pos sible to optimize the method with regard to selectivity If the interfering 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 35 the selectivity for compound X is improved consid erably by decreasing the potential to E2 or E1 Obviously if compound Y is the compound of interest optimization of selectivity in
87. rent unique IP address range can be applied In the following section the procedure to connect the instrument to the PC using the crossover UTP cable is described Configure the IP address of the PC LAN network card by executing the following steps in this example the Windows 7 OS is used in Windows 8 the example screens may look different 1 Right click on the Network icon in the bottom right of the Windows taskbar and open the menu Open Network and Sharing Center Alternative open Windows start menu open control panel and goto Open Network and Sharing Center 22 175 0010 DECADE Elite user manual rev 01 Troubleshoot problems Open Network and Sharing Center s v HER a 13 44 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 and click on properties to open the Network card setting GOF Net Network C gt File Edit View Tools Advanced Help Organize v Disable this network device Fr Ff o A Local Area Connection 7 N Unidentified network GE Realtek PCle GBE Family Controller Disable Status Diagnose Bridge Connections vies Create Shortcut Delete E Rename CHAPTER 2 E Local Area Connection Properties Installation 23 m r r Internet Protocol Version 4 TCP IPv4 Propert
88. researchers for its unsurpassed S N ratio and therefore you now possess the best possible combination for extremely sensitive EC anal yses The DECADE Elite has 3 operational measurement modes DC SCAN and PULSE mode Furthermore a Service Diag nostics and Config ura tion mode are available In addition crucial parameters can be controlled by either relays or TTL The DECADE Elite can support up to 4 flow cells optional which makes it possible to perform 4 independent measure ments with one detector The DECADE Elite is available in different colors The DECADE Lite is a light version of the Elite it is for single flow cell operations in DC mode only This manual covers the installation set up and operation of the DECADE Elite only Detailed operation instructions for other peripheral LC equipment and parts such as flow cells pumps autosamplers valves column heaters etc are given in the manuals accompanying those accessories 14 175 0010 DECADE Elite user manual rev 01 Instrument description DECADE Elite Front side o DECADE Flite 2 DECADE ELITE EC Detector ex a 3 DHF IG OC PULSE SCAN DIAG OOOO 4 eo 5 6 7 8 9 ay Description Description Instrument housing 7 and value keys LC tubing inlet outlet 8 Cursor keys Instrument door panel 9 Door handle for opening door 4 x 40 Ch LCD display Function keys lt Enter gt key ooaRWHD
89. s as well as 50 s it may not be possible to work with one filter setting In that case it is advisable 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 fre quency After optimization do not change the cut off frequency setting during anal ysis of a calibration sequence Use the same settings for analysis of sam ples and calibration standards ADF off ADF 0 02 amp 0 001 Hz programmed Blank ADF on ipa AANA TITTEN 0 02Hz __ ___ 0 001 Hz a OS RS RI 5 10 15 20 25 Fig 27 Analysis of 20 pmol L 5 hydroxytryptophan 5 HIAA and 5 HT us ing ADF for improving detection limits The S N improvement depends on the composition of the frequency spec trum Improvement up to a factor 100 may be obtained compared to an un filtered signal As high frequency noise is suppressed remaining noise 76 175 0010 DECADE Elite user manual rev 01 components will be in the same frequency range as chromatographic peaks As suppressing noise will always result in some suppression of signal it is advised to switch the DECADE Elite to the highest acceptable sensitivity Chapter 5 Detection and Parameters 77 CHAPTER 8 Pulsed Amperometric Detection PAD Introduction In pulsed amperometric detection PAD the working electrode WE is re generated at a frequency of 0 5 3 Hz by the application o
90. s shown in the table below Table VII Analog data output connector lay out To measure for example the analog signal of cell 1 with an external A D converter e Connect the signal wire lead of pin 1 Vout1 to the analog measurement channel of the A D converter e Connect the GND wire lead of pin 6 or 7 9V to the corresponding analog ground connection of the A D converter Digital I O connector 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 CHAPTER 11 Rear panel I O 107 TTL inputs amp outputs Both the TTL inputs and outputs are default high 3 3 Volt The TTL in puts are level triggered the contacts require a minimum TTL low pulse du ration of 100 ms If multiple activations are required the next pulse should be given after 100 ms TTL high When the input is kept low only one acti vation will occur Relays The DECADE Elite has 2 free programmable contact closure outputs o Normally open NO Common ie a em Normally closed NC e Relay1 pin 1 normally closed pin 2 normally open pin 3 common e Relay2 pin 4 normally closed pin 5 normally open pin 6 common The maximum rating for these contact closure outputs are 24 VDC switch ing voltage and 0 25 A The relays can be controlled in the
91. t of experiments boundary con dition all analytes should be sufficiently separated under the ap plied LC conditions Furthermore under real chromatographic conditions reliable information about the S N ratio is obtained e A scanning voltammogram is obtained in the so called scan mode of the DECADE Elite the voltage runs between two pre set poten tial 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 voltammo gram is described Electrochemical reactions In electrochemical detection ECD a reaction of the analyte at an elec trode surface is monitored This distinguishes ECD from most other detec tion techniques where detection is based on the physical properties of an analyte i e mass spectrometry molecular mass absorbance detection molar absorptivity For electrochemically active compounds the potential between reference electrode REF and working electrode WE deter mines the reactivity of the analyte at the WE The potential difference sup 84 175 0010 DECADE Elite user manual rev 01 plies the energy level needed to initiate or enhance the electrochemical re action Different analytes may have different oxidation or reduction poten tials which determines the selectivity of ECD OH OH HON A CH CH NH Ox CH CH NH Z l 2H 2e FSR o Fig 31 Oxidation reduction reaction of norepinephrine
92. t of range error appears when the cell current Ice exceeds the limit of the current range at which the measurement is performed See fig ure below CHAPTER 12 Troubleshooting 113 Out of range CURRENT TIME Fig 48 Example of a chromatogram were the cell current exceeds the maximum current level and the signal is out of range It is important to recognize an out of range overload situation because it may lead to erratic results when quantifying analyte concentrations in samples CAUTION Analytical troubleshooting Analytical problems like loss of signal increase in noise level high cell cur rent loss in sensitivity etc may occur in any U JHPLC ECD system It may be hard to find the cause and several checks need to be performed to eliminate the source of the problem The first step is to determine whether the problem is caused by the DECADE Elite electrochemical detector or the rest of the U HPLC system For that purpose two basic checks should be performed by the end user e Dummy cell test e Stop flow test Dummy cell test The outcome of the dummy cell test which is described in the next paragraph will give an indication if the problems are caused by the detector hardware electronics Stop flow test The stop flow test will eliminate if the problems are caused by the electrochemical flow cell or is originating from the rest of the U HPLC system pump autosampler
93. te 921600 bps e Communication over USB is used for software FW update of the instrument only using Antec boot loader FW upload utility LAN connector RJ 45 bus for serial instrument control over LAN e 10Base T or 100Base TX Auto Sensing serial to ethernet connectivity e Network configuration of Xport via Lantronix device installer software utility e Fixed communication baud rate 921600 bps e Communication over LAN is used for parametric instrument control and data acquisition To establish communication over LAN the LAN cable must be connected when starting the detector using the mains power switch on the rear panel If no communication cable either LAN or USB is connected during start up communication via USB is enabled default VALVE connector Serial D9 9 pins subD male valve control connector for electrical Vici and manual valves e 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 CHAPTER 11 Rear panel I O 101 e Automatic detection of electrically actuated Vici valve valve control in STAT screen and RS232 command ID 30 0 load 1 inject e Manual valve status load inj read out via pin 5 and 6 contact closure e Inject Marker pin 9 TTL output Default high 3 3V load on inject event low 0 V for 2000 ms
94. th the requirements of CISPR11 group 1 class AISM equipment Ina domestic environment this product may cause radio interference in which case the user may be required to take adequate measures on his own expenses Only use manufacturer supplied cable s to connect with other devices Thoroughly connect shielding to common Manufacturer will not accept any liability for damage direct or indirect caused by connecting this instrument to devices and with cables which do not meet relevant safety standards Zoeterwoude The Netherlands March 2 2015 175 0010 DECADE Elite user manual rev 01 Table of contents Table of contents 4 Warning Symbols 7 Safety instructions 9 Introduction 13 Instrument description 14 Installation 17 Site Preparation Requirements 17 Unpacking 18 Mains connection 20 PC connection 21 Software 24 HPLC fluid connections 29 Maintenance amp Shutdown 37 Maintenance 37 Shutting down the system 39 DECADE Elite controller 41 Introduction 41 Overview of DECADE Elite screens 42 Parameters 47 Clarity DECADE Elite control module 55 Detection and Parameters 57 Introduction 57 Three electrode configuration 57 Internal organization 59 Dual flow cell control 59 Filter 64 Introduction 77 Hydrodynamic and scanning voltammogram 85 Specifications DECADE Elite 95 Rear panel I O 99 USB B connector 100 LAN connector 100 VALVE connector 100 ANALOG DATA connector 104 Digital I O connector 106 Troublesho
95. the syring pump settings menu In case a syringe pump is used which can be con trolled in Dialogue the settings can be programmed under the S pump tab A typical flow rate to start with is 10 uL min Optimize the flow rate if required during the scan experiments Start the syringe pump and before scanning assure that the flow cell is sufficiently primed with analyte solution A scan can be started stand alone by pressing the START but ton in the SCAN STAT menu or by starting a single run in Dia logue press the F5 button or click start single run under the Op tions menu To record a background blank scan repeat the experiment with the pure HPLC buffer in which the analyte was dissolved When scanning with the Elite Dialogue software continuous all scans are displayed and can be selected individually 5 wo g HA 8 br A i CSICSICSICSICSICSICSICSICSICSICSICSICSICSICSICSICSICS oe 900 000 Em T T 1 420 000 60 000 540 000 1020 000 1500 000 gt time min E mV Fig 43 Example of multiple scans continuous of Amodiaquine CHAPTER 10 Specifications DECADE Elite 95 CHAPTER 10 Specifications DECADE Elite Environmental dimensions weight amp power requirements Working temperature Storage temperature Humidity Safety and EMC Equipment class Installation category Pollution degree Dimensions
96. the working potential Fig 32 The working potential which gives the best sensitivity is obtained by plotting the signal to noise ratio against the working potential Fig 37 Construction of a hydrodynamic voltammogram for norepinephrine Chromatograms are obtained at cell potentials ranging from 1 0 V back to 0 4 V front with 100 mV steps Chapter 5 Detection and Parameters 91 Construction of a scanning voltammogram A scanning voltammogram can be recorded using the DECADE Elite scan mode The scan mode is programmed in the SCAN SETUP screen of the DECADE Elite Depending on the data acquisition software that is used and the experimental set up a full half or continuous scan cycle can be chosen Fig 38 Programming the scan mode in the SCAN SETUP screen With the Elite Dialogue control software a scanning voltammogram can be programmed under the Detector tab in the main window hl gt Elite Dialogue DECADE ELITE Sax Fie Tools Options Events Window Help Monitor Detector Events S pump Log Cell 1 X On F Enabled Mode Scan X Dev status Output Potential Range 2 pA v E1 000 M Offset 0 B E2 100 M Filter 0 001 Hz scan rate 20 v mV s Data rate 1 Hz sweep half X Polarity l Compensation F On Autozero Temperature Run time Oven 7 On t 0 833 min Set 3 IC Measured 35 5 rG Fig 39 Programming a scan in the Elite Dialogue software
97. tial 80 175 0010 DECADE Elite user manual rev 01 steps and 6 time settings may lead to a time consuming optimization pro cedure 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 mode is the fre quency of the output signal In the DC mode the signal has a 1 100 Hz fre quency in the pulse mode the frequency is determined by the duration of the pulse Once every cycle the ts signal is sent to the output If a stepwise pattern in the chromatogram is seen when zooming in it means that the data acquisition has an unnecessarily high sampling fre quency This leads to large data files but certainly not to a better chroma togram Usually data acquisition at 1 HZ is sufficient D E Fig 30 A detailed part of a chromatogram acquired at different data fre quencies The data rate is A 5x B 2 5x C 1 2x D 0 6x and E 0 3x the frequency of the pulse C is 1 Hz data rate Working electrode material Gold and platinum are used as working electrodes for PAD Glassy carbon appears to be unsuitable due to the high electric capacitance of this mate rial Furthermore resurfacing of the noble metal working electrode is Chapter 5 Detection and Parameters 81 based on formation and removal of a metal oxide layer This is impossi ble with glassy carbon References 1 D
98. tions DECADE II 95 99 Symbols 7 t1 control 51 Tefzel rotor 78 three electrode configuration 57 ts 78 Vespel rotor 78 voltage clamp 57 voltammogram construction 85 90 introduction 83 warning sign 7 working electrode 57 material 80
99. tive elec trochemical detection and pulsed electrochemical detection analysis of carbohydrates using anion exchange chromatography with NaOH as mo bile phase All gases except helium are removed completely Helium is not EC active and does not change the mobile phase properties signifi cantly To prevent mobile phase contamination only high purity helium must be used Installation amp start up Please carefully follow the next steps for a successful installation and start up CHAPTER 2 Installation 33 The installation of the flow cell and column is shown in Fig 4 If a manual injector is applied with position sensor the sensor cable must be connected to the VALVE connector on the rear panel to enable INJECT LOAD functions In the CONFIG screen the valve option should be set to Valve present manually An electrically actuated valve of Vici Valco type E2CA EHCA can be connected amp controlled by the DECADE Elite For that purpose a Serial valve cable is required The serial cable should be connected to the VALVE connector on the rear panel of the instrument to enable INJECT LOAD functions This presence of this valve is automatically detected and no further configuration has to be performed Fig 4 Installation of flow cell and column in the DECADE Elite 34 CAUTION CAUTION 175 0010 DECADE Elite user manual rev 01 4 Prior to connection of the HPLC system to the detector all metal parts sho
100. tozero 3 Trigger to zero null the cell current compensation Ic 21 TTL IN Start 1 A 1 5 Trigger to start a Timefile Scan and Data acauisition 22 TTL IN Start 2 Trigger to start a Timefile Scan and Data acquisition 23 TTL IN Start 3 Trigger to start a Timefile Scan and Data acquisition 24 TTL IN Start 4 Trigger to start a Timefile Scan and Data acquisition 25 GND GND Ground connection A All boards In the detector accessory kit a dedicated I O cable D25 male open 2m pn 250 0131 is supplied CHAPTER 12 Troubleshooting 111 CHAPTER 12 Troubleshooting Maintenance Even though great care was taken in the design of the DECADE Elite problems may occur during operation of the instrument The information in this chapter may help you to identify and solve the source of the problems Errors can be categorized in two types e Instrument errors e Analytical problems In the next sections both types of errors are described In the event that the problems cannot be solved after following the instructions in this trou bleshooting section contact your local supplier for further assistance Instrument errors Incidental fault conditions may occur in any instrument The DECADE Elite will generate an error message containing an error number with a short de scription for several hardware fault conditions Table IX Error messages Error Message 11 Checksum error 12 Temperature sensor 1 error 13 Disconnect
101. uld be specifically designed for use in Ultra High Performance Liquid Chromatography and capable of delivering flow rates typically in the range between 1 uL min up to 5 mL min The manufacturer will not accept any liability for damage direct or indirect caused by connecting this instrument to devices that do not meet the relevant safety standards WARNING Tubing connectors For optimal operation it is of the utmost importance that all tubing connec tions on the injection valve columns and flow cells of the injection valves are made without introducing dead volumes to minimize peak broadening carry over etc CAUTION 30 175 0010 DECADE Elite user manual rev 01 Use only the original polymeric fingertights supplied by Antec with the purchased flow cell to make LC connections on the flow cells inlet and outlet Do not use metal tubing on the flow cell because it may lead to damage or incorrect operation of the flow cell Use PEEK PEEKsil or Fused Silica tubing with FEP sleeves For columns and injection valves etc use only nuts ferrules or finger tights recommend by the respective manufacturer of the parts So for Vici Valco valves use Valco nuts and ferrules for Rheodyne valves use Rheodyne valves etc The use of unsuitable connectors may lead to damage of the parts or the introduction of dead volumes In case the DECADE Elite was purchased as a part of an ALEXYS HPLC ECD analyzer the instrument
102. uld preferably be passivated with 15 nitric acid during 20 min The acid is flushed through the pump the pump tubing the dampener the injector in load and inject position and to waste Make sure that all parts that are not acid resistant such as nylon inlet filters column and flow cell are not connected during this step 5 After flushing with nitric acid the system must be thoroughly flushed with demi water Make sure that no traces of nitric acid are left in the tubing or pulse dampener check with pH paper Flush the system with HPLC buffer If an ISAAC reference electrode is used make sure that the mobile phase contains at least 2 mmol l chloride KCI or NaCl ions 6 Before connecting a new column read the manufacturer s instructions Our experience is that thorough pre conditioning of a column is always required Only a pre conditioned column is electrochemically clean If not the background current may be unacceptably high and substantial fouling of the working electrode occurs For reversed phase columns flushing with 50 methanol in water for 3 days at a low flow rate is highly recommended Before switching to mobile phase flushing with water 10 column volumes is recommended to prevent precipitation of buffer salts 7 Passage of air bubbles through the flow cell will lead to unacceptable noise levels and spikes Therefore the use of an in line degasser is strongly recommended In our experie
103. will cause unsafe situations System Operation To assure optimal performance keep of the detector we recommend that the instrument is checked regularly and maintenance procedures are car ried out Preventive maintenance contracts are available for that Purpose Please contact your local dealer or the nearest sales office for TEA 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 before disassembly ELECTRICAL SHOCK WARNING RISK OF ELECTRIC SHOCK A DISCONNECT POWER BEFORE SERVICING AVERTISSEMENT RISQUE DE CHOC ELECTRIQUE COUPER L ALIMENTATION AVANT LA MAINTENANCE Untrained personnel should not open the instrument this may only be done by authorized service engineers Replace or repair faulty insula tion on power cords immediately after discovery of the fault Check that the actual power voltage is the same as the voltage for which the instruments are wired Make sure power cords are connected to correct voltage sources grounded AC power source line voltage 100 240 VAC The in strument should be connected to a protective earth via a ground socket The DECADE Elite must only be used with appliances and power sources WARNING 10 WARNING 175 0010 DECADE Elite user manual rev 01 with proper protective grounding to prevent damage through bui
104. with flow cells columns and valves can weigh up to 20 kg e A detector has the following dimensions 44 D x 22 W x 43 H cm 17 3 D x 8 7 W x 16 9 H Take into account that additional space is necessary on all sides at least 10 cm to prevent obstruction of ventilation holes and allow sufficient heat dissipation Chemicals Mobile phase and flush storage solutions must be clean as it is in direct contact with the working electrode in EC detection High purity chemicals including water is a prerequisite So all chemicals should be electrochemi cally clean HPLC grade or better For water used for the preparation of mobile phases a water purification apparatus is advised which is able to supply high purity deionized water with resistivity of gt 18 MOhm cm and low TOC level lt 10 ppb Unpacking Inspect the transport box for possible damage as it arrives Immediately in form the transport company in case of damage otherwise she may not ac cept any responsibility Keep the transport box as it is designed for opti mum protection during transport and it may be needed again Carefully un pack the system and inspect it for completeness and for possible damage Contact your supplier in case of damage or if not all marked items on the checklist are included Prior to shipment your detector has been thor oughly inspected and tested to meet the highest possible demands The results of all tests are included CHAPTER 2

175_0010 - DECADE Elite user manual - rev 01

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