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1. 6 The Software Pyro Oxygen Logger This chapter describes all functions of the Pyro Oxygen Logger software excluding the sensor calibration which is described in detail in the chapters 7 and 9 6 1 Coupling of Piccolo2 and PiccoTHP If the Piccolo2 should be used in combination with the climate module PiccoTHP both devices might have to be coupled at the first usage by the following procedure e Connect both the Piccoloz and the PiccoTHP to the computer It is important that no other Piccolo2 or PiccoTHP devices are connected to the computer e Start the Oxygen Logger software and the following dialog should appear An uncoupled PiccoTHP was detected Should this PiccoTHP be coupled to the oxygen meter Piccolo2 e Click Yes Now this specific PiccoTHP is programmatically coupled to the specific Piccolo2 shown by the message Coupled to PiccoTHP visible below the Pyro Oxygen Logger logo Pyro Oxygen Logger Coupled to PiccoTHP pyro M The coupling is stored in the internal flash memory of the PiccoTHP Therefore the coupling is also effective when the devices are used with another PC 22 6 2 Main Window After start of the Pyro Oxygen Logger software the following main window is shown O Pyro Oxygen Logger 28 Pyro Oxygen Logger Ma pyro V 3 201 c 2014 by Robust Oxygen Probe Compensation gem vevews pyro scence com normal range 22
2. Po2 actual partial pressure Patm actual barometric pressure T actual temperature Ph20 T saturated water vapor pressure at temperature T Dissolved O2 concentration C pmol L Definition C umol L A a s 100 x C T P S Used in water phase with C T P S interpolation formula for dissolved oxygen concentration in units of umol L at temperature T atmospheric pressure P and Salinity S see chapter 11 7 Dissolved O2 concentration C mg L ppm Definition C mg L C umol L x 32 1000 Used in water phase Dissolved O2 concentration C mL L Definition C mL L C umol L x 0 02241 Used in water phase 86 11 7 Table of Oxygen Solubility The following Table shows the equilibrium oxygen concentration Cioo T P 1013mbar S in units of umol L at standard atmospheric pressure of 1013 mbar as a function of water temperature in units of C and salinity in units of PSU practical salinity unit g L In order to correct these values for the actual atmospheric pressure Patm the following formula has to be applied Cio0 T P S Cio0 T P 1013mbar S X Pam 1013mMbar References Garcia HE and Gordon LI 1992 Oxygen solubility in seawater Better fitting equations Limnol Oceanogr 37 1307 1312 Millero FJ and Poisson A 1981 International one atmosphere equation of state of seawater Deep Sea Res 28A 625 629 87 Sal PSU Temp C 0 5 10 15 20 25 30 35 88 40 200 4
3. easy and precise 1 click calibration of the oxygen sensor More information concerning our products can be found at www pyro science com or contact us under info pyro science com We would be pleased to serve you concerning all needs for high precision oxygen sensing with state of the art technology Your PyroScience Team 2 Safety Guidelines The Piccolo2 and the PiccoTHP are laboratory instruments to be used with optical oxygen sensors optodes from PyroScience In order to guarantee an optimal performance of the Piccolo2 please follow these operation instructions and safety guidelines The Piccolo2 the PiccoTHP and the sensors should be used in the laboratory by qualified personal only following the operation instructions and safety guidelines of this manual Please follow the appropriate laws and guidelines for safety like EEC directives for protective labor legislation national protective labor legislation safety regulations for accident prevention and safety data sheets from manufacturer of chemicals used during measurements When used in the field extreme environmental conditions like high humidity dust and exposure to salt water or intense solar radiation may cause damage to the Piccolo2 and PiccoTHP Before using the Piccolo2 and the PiccoTHP and the sensors read carefully the instructions and user manuals In case of problems or damage disconnect the instrument and mark it to prevent any furth
4. By clicking on the small rectangular button next to the color control it is possible to show or hide the respective graph This show hide functionality is especially useful if e g a single graph should be inspected in detail while all other graphs are hidden The graph in the Overview panel can be cleared with the button Clear Graph This will not affect the saved data in the data file O Pyro Oxygen Logger xs Pyro Oxygen Logger My Channel 1 Overview Coupled to PiccoTHP pyro DES V 3 201 c 2014 by Oxygen Oxygen Oxygen Oxygen ext Temp Pressure Humidity a oS 9 189 26 65 1009 2 59 16 Geert mg l ppm c mbar Measurement HI ES Oma Ces e 10 27 1014 64 ae gt 3 75 26 9 1013 63 5 single data point 26 8 1012 63 continuous 9 5 duration 26 7 1011 62 5 H 00 01 00 aul 26 6 gi 1010 SEI HH MM SS 9 mi 1009 das H Sample Interval s SZ kb f 8 75 26 4 G 1008 61 J1 i 26 3 1007 60 5 Data Smoothing SS 3 26 2 1005 60 8 25 26 1 1005 59 5 PSS SL SS E 1004 59 120 150 175 200 225 250 275 300 325 350 375 400 425 450 475 500 525 550 575 600 625 650 675 700720 WARNING Data are not 4 ed cl S Seconds s HAR logged into Autoscroll Time Scale a data file Log to File Sme ad So E GD Setup Setup Logo Data 40 6 5 Raw Data Window The Raw Data Window is mostly intended for trouble shooting and advanced users During standard measurements it
5. 10 raw value m AM 20 00 C Li Firmware 3 19 Signal TT C 221 p 22 9 Measurement sto 22 7 single data point continuous 22 6 duration 2 00 01 00 S 22 5 HH MM SS 22 4 Sample Interval s set actual SH 1 22 3 D Data Smoothing 22 2 LG pen J 3 22 1 a 1 1 D D 1 D 1 D H 1 D H D D H D D H 1 H 1 0 25 50 75 100 125 150 175 200 225 250 275 300 325 350 375 400 425 450 475 500 525 550 575 600 WARNING sm Data are not lt FI Cc e J Seconds s HAR into D Se _ Calibrate o Log to File save toad Fah E lt r Setup Setup Logo Data Last Setup SC R The default oxygen sensor readings show uncalibrated sensor readings raw value which give only a qualitative information of the actual oxygen level Note that placing the mouse on many elements of the window will show a short description tool tip By clicking on the right mouse button and selecting Description and Tip a more detailed description might be available additionally 23 S D UD CC TDT Pyro mjt Com pensation Temperature 20 00 tes range 22 10 raw value Dons D O d a ETET HH MH 55 Sampie Interval ei iH atual J h I 25 50 75 100 125 150 175 200 225 250 275 300 32h aen 375 400 425 450 475 500 525 550 575 600 WARNING i Ta P eh Seconds 3 FE Sal HAR logged into Teng Dosia a data fike Log to File c WD SCB C The sensor readings are displayed in a numeric di
6. Background dphi phase shift in can be adjusted refer to chapter 9 2 for more details These two values are only relevant if a background compensation for measurements with contactless sensors has been activated in the Advanced Settings of the respective channel 66 8 Calibration Standards 8 1 The Air Calibration Standard The Air Calibration standard can be e ambient air e water vapor saturated air e air saturated water 100 air saturation All air calibration standards described in the following rely on the virtually constant oxygen content in the earth s atmosphere of about 20 95 O2 in dry air Slight deviations might be given in closed rooms occupied by many people or e g candles combustion engines consuming the oxygen So if in doubt ensure a good ventilation of the room with fresh air e g by opening a window for some minutes Furthermore the relative humidity of the air causes deviations from the ideal value of 20 95 O2 Simply speaking the water vapor in humid air replaces a fraction of the oxygen resulting ina diminished oxygen level of e g 20 7 O2 For temperatures around and below 20 C this effect causes fortunately only a maximum deviation of about 0 5 02 However for higher temperatures at 30 C or even 40 50 C the humidity of the air gets a significant influence on the actual oxygen level For example ambient air at body temperature 37 C with 100 relative humidity contains only 19 6
7. activated By default only the y axis oxygen is in Autoscale mode The button Clear Graph offers the option to clear the graph Note that this will not affect the saved data in the data file All data recorded can be displayed in the chart recorder by moving the bar along the scroll bar SCB Switching off the Autoscroll button will allow inspection of older data which are not displayed anymore in the actual graph e g during longtime measurements The Plot Style of the chart can be changed by clicking with the right mouse button onto the black field of the respective color control CC M above the chart recorder opening a pop up menu With Common Plots Color Line Style and Width Interpolation and Point Style the chart appearance can be changed By clicking on the small rectangular button of the color control CC it is possible to show or hide the respective graph Calibrate e NOT CALIBRATED If a sensor is not yet calibrated i i i E E E E the warning Not Calibrated is EE E shown on the right hand side of pasau Stn Sa oa A i the Calibrate button As long as the sensors are not calibrated the data are shown in units of raw value reflecting only qualitatively the measured oxygen levels In order to switch to quantitative oxygen units a calibration has to be performed by clicking on the button Calibrate The calibration procedure is explained in detail in chapter 7 p 7 Low Signal l l Signal too high The warning
8. and 1 m in length C e a label L attached to the other cable end protected by a black cap instrument end IE ae The attached label L indicates the instrument end of the robust probe to be inserted into the sensor port of the Piccolo2 Instrument It contains also the Sensor Code SC which needs to be entered into the Settings of the Pyro Oxygen Logger software see chapter 6 3 12 SE The instrument end IE of the robust probe needs to be inserted into the Piccolo2 meter see chapter 5 6 Here the bare optical fiber protrudes ca 1 mm from the fiber jacket At the sensor end SE of the robust probe which needs to be inserted into the sample the oxygen sensing surface is surrounded by the steel tubing and covered with a black coating optical isolation to prevent interferences with ambient light The robust probe for the Piccolo2 can be used for standard laboratory applications and for long term measurements in gases and liquids The sensor end and the cable are completely submersible in water and specified for longtime submersion in liquid media Due to the dimensions of the robust probes diffusion limitation in liquid samples of oxygen towards the 3mm tip has a measurable effect on the response time For optimum operation conditions it is therefore recommended to utilize these robust probes in permanently stirred liquids this diffusion limitation is negligible in gas samples 13 5 2 Dippi
9. and PICROD3 please use a Fiber Length of o m 6 3 2 Advanced Settings If Advanced Sensor Settings are chosen more complex setting controls get visible Please ensure that the correct sensor code attached to the sensor has been entered in the field Sensor Code The Advanced Measuring Parameters eee ior nila ers eee comprise the LED Intensity for x A 7 532 207 excitation of the REDFLASH indicator Sensor Settings Basic Advanced in and the Amplification of the oxygen measuring Time ms sensor signal As a rule of thumb the a 1 2 4 E E B amp IB LED Intensity should be 10 30 for LED Intensity Amplification robust dipping probes but can be 2 S10 40 sie Fiber Length im EP increased up to 100 for contactless sensors sensor spots flow through cells respiration vials The Amplification should be typically chosen as 80x 200x or 400x Note that varying the LED Intensity and the Amplification has direct influence on the signal intensity and therefore on the signal to noise ratio The Oxygen Measuring Time defines the integration time for the acquisition of a single data point Shorter measuring times provide 33 low long term drift whereby longer measuring times assure less noise The maximal possible value is 128 ms The Fiber Length m of the connected robust probe OPROB3 or of the connected optical fiber P CFIB2 for contactless sensors must be entered typ is 1m refer to chapte
10. automatic temperature compensation for measurements in a sample the option External Temperature Sensor has to be selected in the Conditions in the Samples in the Piccolo2 Settings window The temperature readings are also saved into the data file 78 11 Appendix 11 1 Specifications of the Piccolo2 Power Supply SVDC from USB port ca 3 5mA average current consumption with max 40mA peaks with ca 10 200ms duration during an oxygen measurement Supported operating systems Windows 2000 XP VISTA 7 8 but not Windows RT Max relative humidity Non condensing conditions Oxygen measuring principle lifetime detection of REDFLASH indicator luminescence Excitation wavelength 620 nm orange red Detection wavelength 760 nm NIR Max sample rate A samples per second Max sample rate with enabled high Ca 10 20 samples per second speed sampling in Settings gt Options ext temperature sensor not activated Please note that the oxygen sensors have a different temperature range typ o 50 C specified 20 C to 70 C not specified Tbe 11 2 Specifications of the PiccoTHP Dimensions ssassn Supported operating systems Windows 2000 XP VISTA 7 8 but not Windows RT Operating temperature o to 50 C Max relative humidity Non condensing conditions External Temperature Sensors 4 wire PT100 Range Resolution Accuracy 30 C to 150 C 0 02 C 0 3 C Internal Temperature Sensor Range Re
11. compensated for pressure changes e g due to weather changes Alternatively it is also possible to enter a fixed pressure see section before Salinity The Salinity g L of the environmental sample in which the measurements are performed needs to be adjusted e g in case of saline water For measurements in gas samples this value has no relevance and is not active For measurements in liquid samples the salinity is only relevant if a concentration unit for oxygen was selected e g mg L or UM L 36 6 3 4 Temperature Settings only with PiccoTHP The External Temperature Sensor of the PiccoTHP and the Internal Temperature Sensor inside the PiccoTHP can be activated in the panel Temperature Both temperature sensors can be activated even if the oxygen measurements are performed at ae Fixed Temperature Channel 1 Temperature options and hence are not Ste affected by the 23 External Temperature Sensor temperature sensor Temperature Measuring Time L Fee TEE measurements The high Default 240ms low noise measured temperature Manual Offset CC 7000 vote is then displayed in the aaa Overview panel see chapter 6 4 of the main window and saved into the data file After activation of a temperature sensor in the Channel 1 tab of the Piccolo2z Settings see 6 3 3 the measured temperature is displayed in the main window right hand side y axis and in the Overview panel see chapter 6 4 a
12. flushed with nitrogen gas for several minutes or directly in nitrogen gas 8 z S The air calibration standard and the calibration conditions need to be defined and entered as described for the 1 Point calibration see chapter 7 1 2 Place now the oxygen sensor into the air calibration standard Wait for steady state until the sensor reading is stable by observing the graph Ensure also stable temperature conditions during the calibration Note that the button Set Air will be highlighted as soon as the oxygen readings are within the expected range for the connected sensor type the latter does not apply for a custom air calibration with 420 95 02 48 If the oxygen sensor readings have reached their steady state click on Set Air and the actual oxygen sensor reading is taken for the air calibration If the oxygen reading seems to be out of the expected range a warning will be shown offering the possibility to repeat the calibration A completed air calibration is indicated by the green indicator Calibrated at Air Subsequently insert the oxygen sensor into the o calibration standard Wait for steady state until the sensor reading is stable by observing the graph Ensure also stable temperature conditions during the calibration Note that the button Set 0 will be highlighted as soon as the oxygen readings are within the expected range for the connected sensor type 4 Calibrate If the oxygen sensor readi
13. is done by a simple running average e g with Data Smoothing s5 always the last 5 sampled data points are averaged However for single data point measurements and for continuous or duration measurements with sample intervals gt 10s the data smoothing is done by averaging repetitive measurements e g with Data Smoothing 3 for each data point 3 oxygen measurements are performed as fast as possible sequentially and the average of these 3 measurements is displayed as the new data point IMPORTANT By default the displayed data are not automatically saved to a data file To activate data saving click on the red start button SB of Log to File Select a file name in the appearing file dialog The saved data files are simple text files with the file extension txt which can be easily imported into common spreadsheet programs Thereafter the indicators Comment and File Path are additionally shown 29 In the field Comment the measure Comment ments can be commented and this comment is then saved together Log to File with the next data point into the LI 2 data file During data logging the data file can be displayed and opened by clicking on the button Show File The data saving is indicated by the light green arrow in the grey Log to File button and can be stopped by clicking this button again 30 6 3 Settings To open the dialog window Piccolo2 Settings click on the Settings button in the Main Window Q
14. is in general not needed After clicking on the Raw Data button in the main window the following Oxygen Sensor Raw Data window opens O Oxygen Sensor Raw Data lee GUSuz S 2 0 00 0 00 0 00 0 00 0 00 The panel of the oxygen channel Chan 1 shows the phase shift as delta phi dphi in dphi is the actual measured raw value which is used for the internal calculation of the oxygen concentration Oxygen uM Oxygen partial pressure hPa Oxygen in air saturation air sat and Oxygen O2 see also chapter 11 4 The Signal Intensity in mV gives a measure of the quality of the oxygen measurement which is also displayed in the horizontal bar indicator in the main window 41 Ambient Light in mV gives a measure of the ambient light entering the sensor from outside At too high ambient light levels the detector of the Piccolo2 might get saturated giving the warning Signal too high in this window and in the warning display of the main window In combination with a connected climate module PiccoTHP the temperature measured by the external temperature sensor External Temp C is displayed as well as the readings of the internal sensors inside the PiccoTHP Internal Temperature Pressure mbar and Humidity sensors On the left side the Status and different warnings can be indicated by the software concerning the signal and reference intensity too low too high and the detection of the
15. measure and adjust manually these parameters The PiccoTHP can be operated in parallel to the Piccolo2 and allows to measure temperature atmospheric pressure and relative humidity automatically ensuring a straightforward calibration procedure and eliminating many possible error sources This chapter covers only the necessary steps during the calibration procedure For details regarding the preparation of the calibration standards refer to chapter 8 Please note that during the oxygen sensor calibration the Sample Interval is automatically set to 0 5 s and the Data Smoothing to 5 ensuring a fast determination of a precise mean value during the sensor calibration After finalization of the calibration the program returns automatically to the former settings 43 7 1 Calibration without PiccoTHP To calibrate a sensor click on the button Calibrate in the main window Note that during data logging this button cannot be used until Log to File was stopped A dialog window Oxygen Sensor Calibration opens in which the calibration mode can be selected Factory calibration for a quick rough calibration taking the o and the air calibration values from the sensor code advised only for rough measurements 1 Point taking the 0 value from the sensor code and the air calibration value from a manual calibration for precise measurements around 21 O2 2 Point taking the 0 and the air calibration value from a manual calibration for
16. modulated emission in the NIR The Piccolo2 measures this phase shift termed dphi in the software The phase shift is then converted into oxygen units based on the Stern Vollmer Theory 83 11 5 Operating several Piccolo2 in parallel The fiber optic oxygen meter Piccolo2 offers one oxygen channel In order to realize extendable multichannel systems with higher channel numbers several Piccolo2 can be easily operated at a single PC as described in the following Connect each Piccolo2 to a free USB port of your PC If the PC does not provide a sufficient number of USB ports you can use an external USB hub The Pyro Oxygen Logger software has now to be started separately for each connected Piccolo2 So if you want to operate e g 4 different Piccolo2 you have to start the Pyro Oxygen Logger software 4 times which will open 4 Pyro Oxygen Logger windows on your desktop The different windows operate completely independent from each other and are associated to exactly one Piccolo2 In order to check which window is associated to a specific Piccolo2 simply press the Flash Logo button in the main window of the Pyro Oxygen Logger software which causes a short flashing 1 2 sec of the red status LED at the associated Piccolo2 followed by a LED flashing at the optionally coupled PiccoTHP When closing the Pyro Oxygen Logger software all settings and all current calibration data are saved in a setup file which is automati
17. readings indicated at Temperature Sensor C Note that the button Set Air will be highlighted as soon as the oxygen readings are within the expected range for the connected sensor type If all readings have reached their steady state click on Set Air and the actual oxygen sensor reading is taken for the air calibration If the oxygen reading seems to be out of the expected range a warning will be shown offering the possibility to repeat the calibration A completed calibration is indicated by the green indicator Calibrated at Air Subsequently insert the oxygen sensor and the external temperature sensor if used into the o calibration standard Also for the o calibration standard either a Fixed 4 Calibrate dphi100 20 000 dphid 52 000 Finish Temperature is adjusted manually or the temperature is read from the external temperature sensor by selecting External Temperature Sensor 58 Wait until the oxygen sensor reading is stable by observing the graph If External Temperature Sensor was selected ensure also stable temperature readings indicated at Temperature Sensor C Note that the button Set 0 will be highlighted as soon as the oxygen readings are within the expected range for the connected sensor type If all readings have reached their steady state click on Set 0 and the actual oxygen sensor reading is taken for the o calibration If the oxygen reading seems to be out of the expected r
18. selected Using the 63 internal humidity sensor requires several precautions please refer to chapter 7 2 2 for more details For precision calibrations it is generally advised to prepare calibration standards with 100 RH refer to chapter 8 which eliminates any possible error source by the usage of the internal humidity sensor The actual Pressure mbar in the calibration standard can be entered manually in Pressure mbar If the calibration standard is exposed to the ambient atmospheric pressure which is given in most typical applications the Internal Pressure Sensor of the PiccoTHP can be selected alternatively This option will read the ambient atmospheric pressure automatically from the integrated pressure sensor in the PiccoTHP The temperature of the calibration standard needs to be determined Either a Fixed Temperature is adjusted manually or the temperature is read from the temperature sensor connected to the PiccoTHP by selecting External Temperature Sensor Place now the oxygen sensor connected to the Piccolo2 and the temperature sensor connected to the PiccoTHP into the calibration standard Wait until the oxygen sensor reading is stable by observing the graph If External Temperature Sensor was selected ensure also stable temperature readings indicated at Temperature Sensor C 4 Calibrate If all readings have reached their steady state click on seen e f Set Air Set Air and the actual oxygen s
19. submersion in liquid samples Further the PiccoTHP has integrated sensors for measuring the atmospheric pressure and the relative humidity of the ambient air The combination of the Piccolo2 with the PiccoTHP makes automatic temperature and pressure compensation of the oxygen measurement possible Additionally it simplifies the calibration procedure of the oxygen sensor 1 click sensor calibration 10 4 Software Installation System requirements e PC or tablet with Windows XP Vista 7 8 but not Windows RT and min 200 MB free disk space Installation steps e download the installer package for the newest version of the Pyro Oxygen Logger software from the PyroScience homepage www pyro science com downloads html e unzip and start the installer and follow the instructions e connect the Piccolo2 to a free USB port of the computer e optionally connect also the PiccoTHP to the computer After the successful installation a new program group Pyro Oxygen Logger is added to the start menu and a short cut named Oxygen Logger can be found on the desktop 11 For an overview of all available oxygen sensor types please visit also the website from PyroScience http www pyroscience com 5 1 Robust Probe The robust probe for the Piccolo2 item no OPROB3 consists of e a stainless steel tubing 3mm in diameter and 30 mm in length protected by a black cap sensor end SE e afiber cable with a diameter of 3 mm
20. the near infrared NIR 760 790 nm red light excitation 02 REDFLASH sensor NIR emission O red light excitation NIR emission Principle red light excited REDFLASH indicators show luminescence in the near infrared NIR which decreases with increasing oxygen quenching effect A high NIR emission at low oxygen and B low NIR at high oxygen The REDFLASH technology impresses by its high precision high reliability low power consumption low cross sensitivity and fast response times The red light excitation significantly reduces 82 interferences caused by autofluorescence and reduces stress in biological systems The REDFLASH indicators show much higher luminescence brightness than competing products working with blue light excitation Therefore the duration of the red flash for a single oxygen measurement could be decreased from typically 100 ms to now typically 10 ms significantly decreasing the light dose exposed to the measuring setup Further due to the excellent luminescence brightness of the REDFLASH indicator the actual sensor matrix can be now prepared much thinner leading to fast response times of the PyroScience oxygen sensors K red light source infrared detector syringe needle infrared emission optical fiber red excitation REDFLASH dye The measuring principle is based on a sinusoidally modulated red excitation light This results in a phase shifted sinusoidally
21. the tips of the sensors are immersed in the water Otherwise follow the calibration procedures given in chapter 7 Please consider that streaming air through water may cause cooling of the water Ensure a correct temperature determination 7O 8 2 The o Standard The o calibration standard can be e water mixed with a strong reductant e water flushed with nitrogen gas N2 e nitrogen gas N2 8 2 1 Water Mixed with a Strong Reductant Fill an appropriate amount of water into a glass flask e g Duran flask with a lid prepared with holes for inserting the oxygen sensor and a temperature sensor Add a strong reductant like sodium dithionite Na S 0 or sodium sulfite Na SO at a concentration of 30 g L creating oxygen free water by chemical reaction It is not recommended to use saline water e g sea water for this because the high salinity of the water might prevent a proper dissolution of the reductant Stir the solution until the salt is completely dissolved and let the solution stand for about 15 minutes Insert the oxygen sensor and optionally also the temperature sensor into the flask and ensure that the sensor tips are completely immersed into the water Otherwise follow the calibration procedures given in chapter 7 8 2 2 Water Flushed with Nitrogen Gas Fill water into a glass flask e g Duran flask with a lid prepared with holes for inserting the oxygen sensor and a temperature sensor Close it and s
22. tips of the oxygen and the temperature sensor are completely dry Wet sensor tips will cause undefined humidity levels around the sensor tips And even worse the evaporation of water drops would cool down the sensor tips causing undefined temperatures 69 8 1 2 Water Vapor Saturated Air Enclose wet cotton wool into a flask e g DURAN flask with a lid prepared with holes for the oxygen sensor connected to the Piccolo2 and a temperature sensor Typically about 1 3 to 1 2 of the flask volume is filled with the wet cotton wool while the other volume fraction is left free for inserting the tip of the oxygen sensor and optionally also the temperature sensor Otherwise follow the calibration procedures given in chapter 7 8 1 3 Air Saturated Water Fill an appropriate amount of water into a flask e g Duran flask with a lid prepared with holes for inserting the oxygen sensor anda temperature sensor Stream for about 10 minutes air through the water with an air stone connected to an air pump available as commercial equipment for fish aquaria Alternatively if no air pump is available fill water into the flask leaving gt 50 air in the head space close it with a lid and shake the flask strongly for about 1 minute Open the lid shortly for ventilating the headspace with fresh air Close it again and shake the flask for 1 more minute Insert the oxygen sensor and optionally also the temperature sensor into the flask and ensure that
23. to the Piccolo2 are exposed to identical environmental conditions 53 Ensure also that the correct sensor code has been entered in the Settings If the sensor code displayed in the Factory o Calibration is not correct click on Finish go to the Settings enter the correct Sensor Code and repeat the calibration Calibration of Channel 1 2 Adjust Calibration Conditions Ambient Air 22 Fixed Temperature C 120 0 o External Temperature Sensor 1 Point in Water S i Lao Place the sensor at the ambient air close to the dphi100 19 900 or Humid Air FireSting device Ensure for a dry sensor tip p 2 Point in Ambient Air 2 Point in Water or Humid Air 2j a 468480 500 520 540 560 588 Factory 0 Calibration Oxygen 02 21 36 Temperature Sensor CC l 20 58 Sensor Code YA7 547 199 Then the air temperature needs to be determined Either a Fixed Temperature is adjusted manually or the temperature is read from the temperature sensor connected to the PiccoTHP by selecting External Temperature Sensor Wait now until the sensor reading is stable by observing the graph and the numerical display of the oxygen sensor reading If External Temperature Sensor was selected ensure also stable temperature readings indicated at Temperature Sensor C Note that the button Set Air will be highlighted as soon as the oxygen readings are within the expected range for the connected sensor type 54 If al
24. 120 16 External Temperature Sensor Place the sensor in water mixed with sodium Custom Mode sulfite or in water flushed with ntrogen gas for several minutes or directly in nitrogen gas Note that this calibration mode will automatically read the atmospheric pressure from the internal pressure sensor in the PiccoTHP The calibration standards must be therefore exposed to the same atmospheric pressure as in typical applications For the air calibration value Air with 100 Humidity or Air Saturated Water place the oxygen sensor connected to the Piccolo2 and the external temperature sensor of the PiccoTHP if used into the flask containing either water vapor saturated air or air Saturated water 100 air saturation Then the temperature of the air calibration standard needs to be determined Either a Fixed Temperature is adjusted manually or the temperature is read from the temperature sensor connected to the PiccoTHP by selecting External Temperature Sensor Wait until the oxygen sensor reading is stable by observing the graph If External Temperature Sensor was selected ensure also stable temperature readings indicated at Temperature Sensor C Note that the button Set Air will be highlighted as soon as the 3 In rare cases and for special applications the calibration standards might be exposed to different pressures than the ambient air where the FireStingO2 is positioned In this case you have to choose the
25. 198 3 196 3 194 3 192 3 190 3 188 4 186 5 184 6 182 7 180 8 179 0 177 1 175 3 173 5 171 7 170 0 168 2 166 5 164 8 163 1 11 8 Explanation of the Sensor Code The oxygen sensors are delivered with an attached sensor code which has to be entered in the Settings refer to chapter 6 3 The following figure gives a short explanation about the information given in the sensor code Example Code XB7 532 205 Sensor Type o LED Intensity Amplification Factory Calibration Co Factory Calibration C100 Sensor Type Robust Dipping Oxygen Probe normal range Oxygen Sensor Spot FTC normal range fast Robust Dipping Oxygen Probe trace range Oxygen Sensor Spot FTC trace range ACHE X Oxygen Sensor Spot FTC normal range LED Intensity A 10 E 40 B 15 F 60 3 20 G 80 D 30 H 100 Amplification 40X 6 200X 5 80x 7 400X 89 Co Factory Calibration at 0 02 dphio Co 10 C100 Factory Calibration at 100 02 dphi1oo C100 10 The values of the factory calibration are valid for the following calibration conditions Partial Volume of Oxygen 02 20 95 Temperature at both calibration points C 20 0 Air Pressure mbar 1013 Humidity RH O DO
26. Custom Mode chapter 7 2 6 where the calibration pressure can be entered manually 60 oxygen readings are within the expected range for the connected sensor type If all readings have reached their steady state click on Set Air and the actual oxygen sensor reading is taken for the air calibration If the oxygen reading seems to be out of the expected range a warning will be shown offering the possibility to repeat the calibration A completed calibration is indicated by the green indicator Calibrated at Air Subsequently insert the oxygen sensor connected to the Piccolo2 and the temperature sensor connected to the PiccoTHP if used into the o calibration standard Also for the o calibration standard either a Fixed Temperature is adjusted manually or the temperature is read from the temperature sensor connected to the PiccoTHP by selecting External Temperature Sensor Wait until the oxygen sensor reading is stable by observing the graph If External Temperature Sensor was selected ensure also stable temperature readings indicated at Temperature Sensor C Note that the button Set 0 will be highlighted as soon as the oxygen readings are within the expected range for the connected sensor type If all readings have reached their steady state click on Gabrated a A dphi100 20 000 4 Calibrate Set 0 and the actual oxygen sensor reading is taken for the 0 calibration If the oxygen reading seems to be out of the
27. ERAL PICCOLO2 INPARALLEL 84 11 6 DEFINITION OF OXYGEN UNITe 85 11 7 TABLE OF OXYGEN SOLUBILITY ees 87 11 8 EXPLANATION OF THE SENSOR CODE 89 1 Overview The ultra compact Piccolo2 is a fiber optic oxygen meter integrated in a small USB stick housing Despite its small size it features the proven high precision REDFLASH technology from PyroScience see Appendix 11 4 for more details The Piccolo2 can be used in combination with a variety of optical oxygen sensors like robust probes dipping probes or contactless sensors sensor spots respiration vials flow through cells The optimized optics of the Piccolo2 enables contactless oxygen measurements up to a window thickness of 20mm All sensors can be used in water samples dissolved oxygen DO as well as in the gas phase gaseous oxygen O2 Simply plug the Piccolo2 into an USB port of your Windows PC or Windows tablet connect the oxygen sensor of your choice and start measuring with the comfortable logging software Pyro Oxygen Logger niin Serpagerieae f SCH beggen Pyro Oxygen Logger puro CG 7 ies a ET Ae WOR SEEN MEMEO Gg comme Se e The Piccolo2 can be optionally operated in combination with the climate module PiccoTHP The PiccoTHP offers temperature atmospheric pressure and relative humidity sensors which can be used for automatic temperature and pressure compensation of the oxygen measurements Further the PiccoTHP allows performing an
28. Mode 1 Point without PiCCOTHP c 0000008 45 7 1 3 Calibration Mode 2 Point without DiccofHp 48 7 1 4 Calibration Mode Custom without PiCCOTHP 0 c 000008 50 7 2 CALIBRATION WITH PICCOTAP 3 ocd edercnalasiaciedaieacnslacleaS dabeecesiatnea 51 7 2 1 Calibration Mode Factory ccccceceeeceeeeeeentteeeeeeeeeeeeeeeeeeeees 53 7 2 2 Calibration Mode 1 Point in Ambient Air 53 7 2 3 Calibration Mode 1 Point in Water or Humid Air 55 7 2 4 Calibration Mode 2 Point in Ambient Air 57 7 2 5 Calibration Mode 2 Point in Water or Humid Air 59 7 2 6 Calibration Mode Custom Mode 62 7 3 ADVANCED ADJUSTMENTS nisi eegen 65 8 CALIBRATION STANDARDS eeh 67 8 1 THE AIR CALIBRATION STANDARD EE 67 Gar tee 69 8 1 2 Water Vapor Saturated Aif ee EE E 70 813 We 70 8 2 REES ege eege 71 8 2 1 Water Mixed witha Strong Reductont 71 8 2 2 Water Flushed with Nitrogen EE 71 623 E de RE 72 9 CALIBRATION OF CONTACTLESS SENSORS cccseeeeeees 73 9 1 CALIBRATION PROCEDUR Eesaia E E E 73 9 2 MANUAL BACKGROUND COMPENSATION 74 10 PICCOTHP Gisin 76 10 1 EXTERNAL TEMPERATURE GENSOR 76 10 2 INTERNAL TEMPERATURE PRESSURE AND HUMIDITY SENSORS 77 10 3 AUTOMATIC TEMPERATURE COMPENSATION 78 ti APPENDIX gia aaa atte 79 11 1 SPECIFICATIONS OF THE PICCOLO2 E 79 11 2 SPECIFICATIONS OF THE PICCOTAP tee 80 SE SE EE ee te 81 Ti MEASURING PRINGIPLE icsceeinssacdsistastaindands a a a EE aE 82 11 5 OPERATING SEV
29. O2 compared to dry air with 20 95 O2 During the calibration of oxygen sensors there are two possibilities to take the humidity into account 67 1 The relative humidity and the temperature of the ambient air must be determined during the calibration The Pyro Oxygen Logger software calculates then automatically the real oxygen level under these conditions bh 2 The calibration standard is prepared in a closed vessel either filled with water or partly filled with e g wet cotton wool or a wet sponge This ensures a constant humidity of 100 RH and there is no need to measure the humidity Option 1 is utilized in the calibration standard Ambient Air when the calibration is performed in combination with the PiccoTHP see section 8 1 1 Option 2 is utilized for the calibration standards Water Vapor Saturated Air see section 8 1 2 and Air Saturated Water see section 8 1 3 Another parameter even more important for the air calibration standard is the atmospheric pressure The principle parameter measured by oxygen sensors is not the partial volume i e 02 but the partial oxygen pressure i e mbar see also appendix 11 6 So an oxygen level of e g 20 7 02 determined as described above by a given humidity and temperature is converted internally by the Pyro Oxygen Logger software into a partial pressure of oxygen essentially by multiplying the relative oxygen level with the atmospheric pressur
30. Piccolo2 Settings A XA 7 53 2 206 Settings can only be adjusted if data logging is not active In the settings the user has to define 1 the Sensor Settings and 2 also the Conditions in the Sample under investigation In the Channel 1 tab the oxygen sensor can be activated de activated by clicking on the button Activate Activation is indicated by a change from dark to light green of the arrow in the button A text describing the connected sensor type appears on the right hand side of this button after the Sensor Code see label on sensor has been entered This description will be shown in the description display DD of the main window and in the data file Please take care that the sensor code of the sensor connected to the Piccolo2 is entered into the field Sensor Code in the Channel 1 tab of the window Piccolo2 Settings It includes information for 31 optimal sensor settings and for calibration data needed for the factory and 1 point calibration The first letter of the sensor code defines the sensor type A detailed explanation of the sensor code is given in chapter 11 8 The oxygen units can be selected by the selector Units The selectable units include raw value default air saturation O mL LU umol LU mg L ppm hPa mbar mmHg Torr dphi and ug LU ppb For measurements in a Gas Phase only the units raw value 0 hPa mbar mmHg Torr and dphi can be selected whereas for measurements o
31. USB driven fiber optic oxygen meter for usage in the laboratory It is compatible to a variety of oxygen sensors from PyroScience like robust probes dipping probes and contactless sensors including sensor spots respiration vials and flow through cells These sensors are available in versions for the full range advised range 0 50 O2 max range o 100 O2 and selected sensor types are additionally available for the trace range advised range 0 10 O2 max range o 21 O2 The Piccolo2 utilizes a measuring principle based on red light excitation and lifetime detection in the near infrared using unique luminescent oxygen indicators REDFLASH technology see Appendix 11 4 for more details The Piccolo2 can be connected directly with the USB plug to a PC with a Windows operation system providing the energy supply and data exchange with the PC If a greater distance between the PC and the Piccolo2 is needed the delivered USB extension cable can be used Comfortable calibration and logging functions are provided by the logging software Pyro Oxygen Logger available as a free download DC gt oxygen sensor If you would like to perform oxygen measurements under changing temperature conditions the Piccolo2 can be operated in parallel with the optional climate module PiccoTHP The PiccoTHP offers an external temperature sensor with 1m cable length The cable of the temperature sensor is Teflon coated and intended for long term
32. also possible to enter values for the Background and dphi manually into the field Manual and subsequently clicking on Take Manual Values If you enter manually zero for Background no background compensation is performed After the background compensation is finished the window closes and the program proceeds with the main oxygen sensor calibration window see chapter 7 It is important that for the subsequent calibration process the Optical Fiber is again attached to the sensor spot position e g by connecting the adapter end again to the spot adapter adapter ring or to the flow through cell Please ensure that during the background compensation the Optical Fiber is not connected to the contactless sensor Please ensure that during the subsequent calibration process the Optical Fiber is again attached to the contactless sensor Remind that the position of the spot adapter or adapter ring should not be changed after calibration of the sensor spot otherwise it has to be re calibrated 75 10 PiccoTHP The Piccolo2 oxygen meter can be operated in parallel with the USB driven climate module PiccoTHP offering several advanced functionalities for sensor calibration and oxygen measurements with the Piccoloz The PiccoTHP has integrated atmospheric pressure and humidity sensors for a precise and easy sensor calibration but also for automatic pressure compensation of the oxygen measurements The PiccoTHP has a perma
33. and the internal Humidity Sensor inside the PiccoTHP can be independently activated by clicking on the respective buttons and are displayed in the Overview panel see chapter 6 4 and saved into the data file If the Pressure Sensor of the PiccoTHP was activated in the Channel 1 tab see 6 3 3 it is automatically activated in the Options tab 38 Q Piccolo2 Settings Advanced Options only for advanced users The USB communication speed can be adjusted e g for improving the maximum sampling rate default 57600 Activation of the button Enable High Speed Sampling will enable the adjustment of a Sample Interval lt o 25 s in the main window and disable Max Data Points in Graphs The Data Export is only relevant for OEM modules or third party software 39 6 4 Overview Panel only with PiccoTHP The Overview panel is only visible in combination with the climate module PiccoTHP The sensor readings of the oxygen sensor left y axis external temperature sensor internal pressure and humidity sensor right y axis are displayed here Each sensor reading is shown also as a numerical value in the on top of the overview graph The plot style of each channel in the chart can be changed by clicking on the color control With Common Plots Color Line Style and Width Interpolation and Point Style each plot in the chart can be changed The items Bar Plots Fill BaseLine and Y Scale are not appropriate for this application
34. andard see chapter 8 1 1 a good air circulation of the ambient air into the dry setup is important ensuring that the relative humidity within the setup is identical to the relative humidity outside the setup at the PiccoTHP However for typical applications around room temperature and below a falsely determined relative humidity gives a maximum relative error of 1 for the air calibration refer also to chapter 8 1 So if this precision is sufficient Ambient Air should be the preferred option for the air calibration standard For precision applications without the possibility to ensure a dry setup for the calibration procedure the alternative air calibration standards Water Vapor Saturated Air see also chapter 8 1 2 or Air Saturated Water see also chapter 8 1 3 should be preferred In the first case some part of the inner volume of the setup can be filled with e g wet cotton wool ensuring 100 RH around the oxygen sensor position In the latter case the inner volume of the setup is simply filled with air saturated water prepared as described in chapter 8 1 3 Ensure that the oxygen sensor is completely covered with the air saturated water 73 9 2 Manual Background Compensation The calibration of contactless sensors i e sensor spots flow through cells and respiration vials includes a compensation of potential background fluorescence from the fiber optic cable connecting the Piccolo2 with the contactless o
35. ange a warning will be shown offering the possibility to repeat the calibration A completed calibration is indicated by the green indicator Calibrated at 0 Finally click on Finish for returning to the main window 7 2 5 Calibration Mode 2 Point in Water or Humid Air In the calibration mode 2 Point in Water or Humid Air both the air calibration value and the 0 calibration value are determined from a manual calibration Air saturated water or water vapor saturated air is used for determining the air calibration value and a specially prepared o calibration standard for determining the o calibration value The preparation of appropriate calibration standards is explained in chapter 8 59 P Oxygen Sensor Calibration Calibration of Channel 1 1 Choose Calibration Mode 2 Adjust Calibration Conditions 3 Wait for Steady State 4 Calibrate Air with 100 Humidity or Air Saturated Water Set Air 1 Point in Ambient Air Fixed Temperature C d H 20 0 External Temperature Sensor _ Not Calbrated 1 Point in Water OPTION 1 dphi100 19 900 or Humid Air Place sensor in a closed chamber half filled with wet cotton ensuring 100 humidity S 5 2 Point in eet Ambient Air ith air for several Ge r and 50 air shaken vividly for several minutes Set 0 21 e 1 103 120 140 160 180 200 Z l l 0 Calibration Oxygen 02 21 42 dphid 54 700 Faxed Temperature C A Ian Temperature Sensor CC
36. aseous chlorine Cl2 can interfere with the oxygen sensor reading No cross sensitivity is found for pH 1 14 CO2 CH4 H2S and any ionic species A signal drift of the sensor can indicate photobleaching of the oxygen sensitive REDFLASH indicator depending on the ambient light intensity as well as the intensity of the excitation light and the sample frequency This can necessitate a recalibration of the sensor and eventually also a readjustment of the Sensor Settings LED intensity see also chapter 6 3 2 In case of sensor spots a re positioning of the spot fiber on the sensor spot and a subsequent new calibration might be necessary If the signal intensity is getting too low typically after several millions of data points as indicated by the horizontal indicator bar in the Pyro Oxygen Logger software and by the respective warning see chapter 6 the sensor needs to be replaced After finalization of the measurements the sensor tip of the robust dipping probe should be rinsed carefully with demineralized water Especially after application in seawater it is recommended to clean the sensor thoroughly with demineralized water Put on carefully the protective caps onto both ends of the probe For sensor spots and respiration vials wet cotton swabs can be used for carefully wiping over the sensing surface Rinse the sensing surface sufficiently with demineralized water afterwards to remove small particles and let it dry before storage
37. ate 0 calibration standards is explained in chapter 8 O Oxygen Sensor Calibration Calibration of Channel 1 1 Choose Calibration Mode 2 Adjust Calibration Conditions 3 Wait for Steady State 4 Calibrate Ambient Air Factory 22 Set Air 1 Point in ad Ambient Air Fixed Temperature CC J 20 0 External Temperature Sensor 1 Point in Water Place the sensor at the ambient air close to the or Humid Air FireSting device Ensure for a dry sensor tip 2 Point in Ambient Air 2 Point in Water Set 0 or Humid Air Too n 140 160 180 19 es l 0 Calibration Oxygen 02 21 43 tot citata dphi0 54 700 Fixed Temperature C 20 0 Temperature Sensor CC 120 16 _ Not Calbrated dphi100 19 900 Oxygen 02 External Temperature Sensor Place the sensor in water mixed with sodium Custom Mode sulfite or in water flushed with nitrogen gas for several minutes or directly in nitrogen gas Note that this calibration mode will automatically read the atmospheric pressure and humidity from the internal pressure and humidity sensor in the PiccoTHP 5 For the air calibration value Ambient Air place the oxygen sensor connected to the Piccolo2 and the temperature sensor if used of the PiccoTHP close to the air holes at the side of the PiccoTHP Wait until the oxygen sensor reading is stable by observing the graph If External Temperature Sensor was selected ensure also stable temperature
38. ature readings indicated at Temperature Sensor C Note that the button Set Air will be highlighted as soon as the oxygen readings are within the expected 4 Calibrate range for the connected sensor type If all readings have reached their steady state click on Set Air and the actual oxygen sensor reading is taken dphi100 21 650 2 In rare cases and for special applications the calibration standard might be exposed to different pressures than the ambient air where the PiccoTHP is positioned In this case you have to choose the Custom Mode chapter 7 2 6 where the calibration pressure can be entered manually 56 for the air calibration If the oxygen reading seems to be out of the expected range a warning will be shown offering the possibility to repeat the calibration A completed calibration is indicated by the green indicator Calibrated at Air Finally click on Finish for returning to the main window 7 2 4 Calibration Mode 2 Point in Ambient Air NOTE The calibration 2 Point in Ambient Air is only possible if the correct Sensor Code has been entered in the Settings see chapter 6 3 In the calibration mode 2 Point in Ambient Air both the air calibration value and the 0 calibration value are determined from a manual calibration Ambient air is used for determining the air calibration value and a specially prepared 0 calibration standard for determining the 0 calibration value The preparation of appropri
39. bration Calibration of Channel 1 1 Choose Calibration Mode 2 Adjust Calibration Conditions 3 Wait for Steady State 4 Calibrate Air Calibration Oxygen 02 Water DO gt pe of 20 95 o Gas Phase air 20 95 Fixed Temperature CC a 22 0 External Temperature Sensor Pressure mbar 1013 Elevation m 200 dphi100 20 700 5 Humidity 9RH A 100 0 a a A a ala aE 60 80 100 120 0 40 l Factory 0 Calibration Oxygen 02 20 58 Factory Calibration Temperature Sensor CC NaN The factory calibration is taken pov 0 that 02 calibration point Ensure correct sensor code is entered in the Custom Mode Sing window Sensor Code XA7 532 207 The oxygen level in the calibration standard can be freely chosen in Oxygen 02 If the air calibration standard is based on ambient air or air saturated water then this value should be kept at 20 95 O2 default representing the standard oxygen volume fraction in ambient air However other values can be adjusted if custom calibration gases are used of e g 5 02 which might be useful when using trace oxygen sensors The temperature of the calibration standard needs to be determined and adjusted manually Fixed Temperature Also the actual atmospheric pressure in the calibration standard can be entered manually in Pressure mbar Normal conditions refer to 1013 mbar default setting If the actual atmospheric pressure cannot be determin
40. cally loaded at the next startup This setup file is saved specifically for each Piccolo2 serial number i e each Piccolo2 in the above described setup keeps its own settings and calibration data 84 11 6 Definition of Oxygen Units phase shift dphi The phase shift dphi is the fundamental unit measured by the optoelectronics in the Piccolo2 see chapter 11 4 Please note that dphi is not at all linearly dependent on the oxygen units and increasing oxygen levels correspond to decreasing dphi values and vice versa As athumb of rule anoxic conditions will give about dphi 53 whereby ambient air will give about dphi 20 raw value raw value Definition raw value 02 uncalibrated The unit raw value is the default unit for uncalibrated sensors and shows only qualitative oxygen sensor readings partial pressure po hPa mbar Used in gas and water phase For a calibrated sensor the partial oxygen pressure po in units of hPa equivalent to mbar is the fundamental oxygen unit measured by the Piccoloz2 partial pressure po Torr Definition PoalTorr po hPa x 759 96 1013 25 Used in gas or water phase volume percent py 02 Definition Pv Po2 hPa Patm X 100 Used in gas with Daten actual barometric pressure air saturation A a s Definition A a s 100 x po P1000 Used in water phase 85 with P1009 0 2095 Patm PxH20 T Py20 T 6 112mMbar x exp 17 62 T C 243 12 T C
41. display WD can show the Soe following warnings 26 Low signal The sensor is either not connected or needs to be replaced by a new one In case of contactless sensors it might indicate that the distance between the optical fiber and the sensor spot is too large For advanced users increase the LED intensity and or the amplification in the Advanced Settings Signal too high There might be too much ambient light on the sensor tip or the sensor spot Avoid direct sun light exposure or strong direct illumination with a lamp For advanced users decrease the LED intensity and or the amplification in the Advanced Settings Bad reference This indicates internal problems of the Piccoloz Please contact PyroScience for support Refer also to the Troubleshooting in chapter 11 3 The adjustment of the Settings using the button Settings is described in detail in chapter 6 3 The button Save Setup can be used to save the current settings and calibration data of a Piccolo2 They can be reloaded anytime by pressing the button Load Setup This allows e g to switch between different laboratory setups with a single Piccolo2 This function might be also useful if different computers are used for the calibration and for the actual measurements You might calibrate the sensors with the first computer save the configuration with Save Setup By transferring this file and also the oxygen meter Piccolo2 to a second computer you can load the
42. e SG Piccolo2 consists of a black fiber cable 3mm diameter 1m length with a label L attached to it The attached label L indicates the hi instrument end IE of the optical fiber Instrument which needs to be inserted into the sensor port of the Piccolo2 see chapter 5 6 Here the optical fiber protrudes ca 1 mm from the fiber jacket At the adapter end AE of the optical fiber P CF B2 the optical fiber protrudes ca 8 mm from the fiber jacket The AE needs to be inserted into the basic spot adapter at the spot position Alternatively the optical fiber rod item no P CROD2 can be connected to the basic spot adapter It consists of a stainless steel tubing 3mm in diameter and 40mm total length At the instrument end IE the optical fiber protrudes ca 1 mm from the steel tubing which needs to be inserted into the sensor port of the Piccolo2 see chapter 5 6 EE The adapter end AE of the N optical fiber rod is marked by a black ring and needs to be Optical Fiber Rod pyroscience inserted into the basic spot ED adapter at the sensor spot position At the adapter end the m IE PICROD2 AE fiber tip protrudes ca 8 mm from the steel tubing Both ends are protected with caps when delivered 16 For read out of sensor spots through transparent windows 10 20mm_ in thickness the optical fiber rod P CROD3 including an adapter is available with ca 50mm total length and a diamete
43. e of e g 990 mbar see chapter 11 6 0 207 xX g90mbar 205mbar giving a partial oxygen pressure of e g 205 mbar This is the essential calibration value used internally by the Pyro Oxygen Logger software The atmospheric pressure can be influenced 1 by weather changes e g varying between ca 990 and 1030 at sea level and 2 by the elevation above sea level e g at 1000 m 68 elevation the typical atmospheric pressure is about 900 mbar compared to 1013 mbar at sea level Thus in summary there are three important parameters to be known for the air calibration standard e Temperature C e Relative Humidity RH e Atmospheric Pressure mbar The Piccolo2 and the Pyro Oxygen Logger software will guide the user through all calibration steps not requiring the theoretical knowledge given above And in combination with the PiccoTHP the built in humidity and pressure sensors together with the external temperature sensor will measure these parameters automatically for most calibration types see also chapter 7 8 1 1 Ambient Air If ambient air is used as the air calibration standard there is no need for preparation The dry oxygen sensor connected to the Piccolo2 optionally together with the dry temperature sensor connected to the PiccoTHP is simply exposed to the ambient air Otherwise follow the calibration procedures given in chapter 7 For precise calibrations in ambient air it is important that the measuring
44. ed on site it is alternatively possible to enter the actual Elevation m in meters above sea level For this click on Elevation and enter the actual elevation Please note that this option takes only the elevation dependent pressure change into account but not the variations due to the actual weather conditions Therefore determining the actual atmospheric pressure with a barometer gives more precise results 46 The relative Humidity RH of the gas phase e g air needs to be adjusted If a calibration standard with water vapor saturated air is used see chapter 8 1 2 it must be adjusted to 100 RH Otherwise the humidity must be determined with a hygrometer and entered If the current Humidity cannot be determined a value of 40 is a good estimate under conditions around 20 C an incorrectly entered humidity will cause a max calibration error of ca 1 deviation Place now the oxygen sensor into the air calibration standard NOTE Ensure constant calibration conditions Wait for steady state until the sensor reading is stable by observing the graph Ensure also a stable temperature during calibration Note that the button Set Air will be highlighted as soon as the oxygen reading is within the expected range for the connected sensor type the latter does not apply for a custom air calibration with 420 95 02 4 Calibrate If the oxygen readings have reached their steady state click on Set Air and the actual oxygen
45. ensor reading is taken for the air calibration A completed calibration is calibrated atai dphi100 20 000 indicated by the green indicator Calibrated at Air 64 Consequently the o calibration should be performed If the calibration type o Factory Calibration was selected no further steps are necessary refer also to chapter 7 2 1 If o calibration was selected follow the instructions given in chapter 7 2 4 Click on Finish for reverting to the main window 7 3 Advanced adjustments This section is only for advanced users For very advanced applications it is possible to manipulate all internal calibration parameters of the Piccolo2 manually This option is accessible by selecting Custom Mode and subsequently clicking on Advanced Adjust Manually which opens a separate window showing all internal calibration parameters OF Adjust Calibration Manually ass sta J220 ar 1013 Ao Aoo Here the Upper Calibration Point default Partial Volume of Oxygen 20 95 O2 can be defined as well as the calibration conditions Temperature Air Pressure Humidity as described for the Custom Calibration see chapter 7 2 6 The Temperature C at o needs to be determined and entered 65 Further the phase shift dphi dphi see chapter 11 4 for the 0 calibration standard dphi 0 in and for the air calibration standard dphi 100 in can be adjusted manually The Background Amplitude in mV and the
46. er use Consult PyroScience for advice There are no serviceable parts inside the device Please note that opening the housing will void the warranty The Piccolo2 and PiccoTHP are not watertight are sensitive to corrosive conditions and to changes in temperature causing condensation Avoid any condition e g direct sun light causing a heating of the device above 50 C 122 F or below o C Avoid any elevated humidity causing condensing conditions Handle the sensors with care especially after removal of the protective cap Prevent mechanical stress to the fragile sensing tip Avoid strong bending of the fiber cable Calibration and application of the sensors are on the user s authority as well as data acquisition treatment and publication The sensors and the oxygen meter Piccolo2 and PiccoTHP are not intended for medical diagnostic therapeutic or military purposes or any other safety critical applications The sensors must not be used for applications in humans and should not be brought in direct contact with foods intended for consumption by humans The sensors should be used in the laboratory only by qualified personal following the user instructions and the safety guidelines of the manual as well as the appropriate laws and guidelines for safety in the laboratory Keep the sensors the oxygen meter Piccolo2 and the PiccoTHP outside the reach of children 3 Introduction to the Piccolo2 The Piccolo2 is a miniature
47. expected range a warning will be shown offering the possibility to repeat the calibration Calibrated ai i dphid 52 000 A completed calibration is indicated by the green indicator Calibrated at 0 Finally click on Finish for returning to the main window 61 7 2 6 Calibration Mode Custom Mode The custom calibration mode is selected by clicking on the button Custom Mode This mode allows the user to combine freely all possible calibration types for the air calibration and the o calibration The air calibration type can be selected by clicking on the Air Calibration Selector The 0 calibration type can be selected by clicking on the o Calibration Selector Air Calibration Selector Calibration of Channel 1 2 Adjust Calibration Conditions 3 Wait for Steady State 4 Calibrate J Custom Calibration l as Set Air 1 Point in water oo Coen 02 Ambient Air o Gas Phase J2 r ST r G H Zon 1 Point m Water Fxed Temperature C 20 0 getut G0 19 900 11003 2 Point in Ambient Air 2 Point in Water 2 or Humid Ap 21 182 200 220 240 260 280 302 ated 0 Calibration Oxygen 02 21 41 dohi 54 700 o Fixed Temperature C SEI Temperature Sensor CC 20 20 External Temperature Sensor dphe 19 743 csr o Calibration Selector The following air calibration types can be chosen e Factory Air Calibration refer to chapter 7 2 1 e Air with 100 Hum
48. f dissolved oxygen in a Water sample DO all units except O can be selected For detailed information please refer to chapter 11 6 NOTE The Chart is automatically cleared in the main window after the Settings have been modified Re adjustments in the Settings might require also a re calibration of the sensor If the changes of the Settings require a sensor recalibration a warning Not Calibrated appears right hand side of the Calibrate button in the main window arseen preamen 6 3 1 Basic Settings The Sensor Settings can be adjusted in a Basic or an Advanced mode The first time user is advised Sensor Code see label on sensor to work with the Basic Settings X A7 532 207 Please ensure that the correct sensor erer olease Advanced code attached tothe sensorhasbeen SS Measuring Mode entered in the field Sensor Code 1 2 4 8 6 R o e The Measuring Mode can be mot defaut low noise adjusted gradually between low drift Fiber Length m 279 1 and low noise 128 of the sensor signal by moving the arrow with the mouse along the scale thereby changing the oxygen 32 measuring time For standard applications an intermediate mode 8 or 16 is advisable The Fiber Length m of the connected robust probe OPROB3 or of the connected optical fiber P CFIB2 for contactless sensors must be entered typ is 1m refer to chapter 9 2 for more details For the dipping probe OPD P2o and for the optical fiber rods PICROD2
49. he air calibration value from a manual calibration for precise measurements over the full range 0 100 dissolved O2 DO This mode uses air saturated water or water vapor saturated air for determining the air calibration value For advanced users and applications a Custom Mode can be selected allowing the user to combine freely the possible calibration types for the air and the 0 calibration 52 7 2 1 Calibration Mode Factory If the calibration mode Factory Calibration is chosen ensure that the correct sensor code has been entered in the Settings as displayed in 2 Adjust Calibration Conditions of the Oxygen Sensor Calibration window If the sensor code displayed is not correct click on Finish go to the Settings enter the correct Sensor Code and repeat the Factory Calibration If the Sensor code is correct the factory calibration is completed by clicking on Finish thereby returning to the main window 7 2 2 Calibration Mode 1 Point in Ambient Air The calibration mode 1 Point in Ambient Air is selected by clicking on the button 1 Point in Ambient Air This mode uses the ambient air for determining the air calibration value The o calibration value is taken from the Sensor Code Because this calibration mode will automatically read the atmospheric pressure and the relative humidity from the internal pressure and humidity sensors it is important that both the PiccoTHP and the oxygen sensor connected
50. iccoTHP 19 5 6 Connecting the Sensors and Optical Fibers The robust probe OPROB3 the dipping probe OPDIP2o0 and the optical fiber P CF B2 or the optical fiber rod P CROD2 PICROD3 are connected to the sensor port of the Piccolo2 with their instrument end IE At the instrument end IE the core of the optical fiber protrudes ca 1 mm out of the fiber jacket For the OPROB3 the OPDIP2o and the PICFIB2 the instrument end IE is marked with gt Instrument on the attached label First remove the protective caps from both ends of the optical probe fiber rod Then slightly unscrew and loosen the nut at the sensor port of the Piccolo2 The nut must not be removed from the sensor port Typically unscrewing the nut with a single turn 180 is sufficient Insert now the instrument end IE of the robust dipping probe or optical fiber rod carefully into the sensor port of the Piccolo2 see pictures with insertion of a dipping probe and b optical fiber rod Fix the optical probe fiber rod by screwing down tight the nut onto the sensor port of the Piccoloz wm B gt mj ee 20 5 7 Cleaning and Maintenance of the Sensors All oxygen sensors for the Piccolo2 can be sterilized with ethylene oxide EtO and can be cleaned with ethanol peroxide 3 H202 or soap solution They can be applied in the gas phase in aqueous solutions as well as in ethanol methanol or isopropanol Other organic solvents and g
51. iccolo2 and optional PiccoTHP more details in chapter 11 5 Clicking on Raw Data opens a pop up MSB _ single data point continuous duration J 00 01 40 HH MM SS window Raw Data which is described in chapter 6 5 DUD A Measurement is started by clicking on the EE measurement start button MSB on the fon right hand side The arrow in the button Data Smoothing SP turns from dark green to light green indicating that a measurement is in progress Clicking again on it will stop the Dota are not a data file measurement Measurement Measurement Log to File 28 The mode of Measurement can be chosen as single data point acquisition continuous sampling default setting or as continuous sampling limited to a defined time interval The duration of the time interval can be adjusted in the duration display DUD shown as hour HH minutes MM seconds SS The Sample Interval s for continuous sampling can be defined in the field designed with set Setting the sample interval to 0 25 will give the maximal possible scan rate The exact maximal rate depends on the settings The actual sample interval is shown in the display actual and is displayed in red if the actual is not equal the set sample interval The acquired data can be smoothed by a Data Smoothing range 1 10 default 3 a value of 1 means no data smoothing For continuous or duration measurements with a sample interval lt 10s data smoothing
52. idity or Air Saturated Water refer to chapter 7 2 3 e Ambient Air refer to chapter 7 2 2 e Custom Calibration more details below And for the 0 calibration the following types can be selected e Factory 0 Calibration refer to chapter 7 2 1 e o Calibration refer to chapter 7 2 4 62 Most calibration types have been explained already in the preceding chapters see references in the above list Only the air calibration type Custom Calibration is a unique feature only available in the Custom Mode The Custom Calibration offers the most flexible options for performing an air calibration as explained in the following r Custom Calibration Custom Calibration The oxygen level in the calibration standard can be freely chosen in Oxygen 02 If the air calibration standard is based on ambient air or air saturated water then this value should be kept at 20 95 02 representing the standard oxygen volume fraction in ambient air However other values can be adjusted if custom calibration gases are used of e g 5 02 which might be useful when using trace oxygen sensors Depending on the calibration standard used select either Water DO for dissolved oxygen or Gas Phase In the latter case the of relative Humidity RH of the gas needs to be determined with a hygrometer and entered If the calibration is performed in the same gas phase in which the PiccoTHP is placed the Internal Humidity Sensor of the PiccoTHP can be
53. l readings have reached their steady state click on Set Air and the actual oxygen sensor reading is taken for the air 4 Calibrate calibration If the oxygen reading seems to be out of the expected range a warning will be shown offering the possibility to repeat the calibration A completed ang calibration is indicated by the green indicator 4 Calibrated at Air Finally click on Finish for returning to the main window 7 2 3 Calibration Mode 1 Point in Water or Humid Air NOTE The calibration 1 Point in Water or Humid Air is only possible if the correct Sensor Code has been entered in the Settings see chapter 6 3 The calibration mode 1 Point in Water or Humid Air uses air saturated water or water vapor saturated air for determining the O Oxygen Sensor Calibration Calibration of Channel 1 1 Choose Calibration Mode 2 Adjust Calibration Conditions 3 Wait for Steady State 4 Calibrate Air with 100 Humidity or Air Saturated Water 1 Point in Ambient Air Fixed Temperature CC J 120 0 a ae a External Temperature Sensor 1 Point in Water OPTION 1 or Air Place sensor in a closed chamber half filled with wet cotton ensuring 100 humidity dphi100 19 900 Oxygen 02 2 Point in Ambient Air and 50 air shaken vividly for ral 2 Point in Water w ES or Humid Air 28540 560 580 600 620 Factory 0 Calibration Oxygen 02 21 36 Factory Calibration Temperature Sensor CC l 20 59 The facto
54. nd saved into the data file The temperature is displayed in units of degree Celsius C The warning Bad Temperature appears DEST below the chart recorder in the main window if the temperature sensor is faulty iili eoni A Manual Offset of the External Temperature Sensor might be entered for possible re calibration of the temperature sensor default o 37 6 3 5 Options In the panel Options it is possible to designate a specific name to the connected Piccolo2 in Device Name e g water sample 12 This device name is then displayed in the top line of the main window This option is especially useful if several Piccolo2 devices are operated in parallel in order to distinguish the opened logger windows O Piccolo2 Settings Channel1 Options Device Name water sample 12 d NOTE Changing will clear the graphs Max Data Points in Graphs 10800 ZS Values much larger than 10000 might vi slow down the max sample rate Advanced Options USB speed lt 57600 Default 57600 Enable High Speed Sampling The maximum number of data points kept in the graphs can be changed by the selector Max Data Points in Graphs default 10800 A change of the number will clear the graphs and high values gt gt 10000 might decrease the maximum sample rate If a PiccoTHP is operated in parallel to the Piccolo2 the internal sensors can be activated in the Options tab The internal Pressure Sensor
55. nently mounted external temperature sensor enabling automatic temperature compensation see chapter 10 3 of the oxygen measurements and of the oxygen sensor calibration Besides this external temperature sensor the PiccoTHP has also an internal temperature sensor integrated into the housing for temperature measurements in the ambient air gas phase For the measuring range precision and absolute accuracy of the sensors refer to chapter 11 2 10 1 External Temperature Sensor The PiccoTHP has a permanently mounted submersible temperature sensor specified for long term submersion into aquatic samples The complete sensor cable is Teflon coated and can be submersed 76 If inserted into the calibration standard or the sample the external temperature sensor can be used for automatic temperature compensation of the calibration see chapter 7 2 and or of the oxygen measurements with the Piccolo2 see chapter 6 3 4 Furthermore even if oxygen measurements are performed at a fixed temperature the external temperature sensor can be used for independent temperature measurements if activated in the Temperature panel of the Piccolo2 Settings window see chapter 6 3 4 and displayed in the Overview panel see chapter 6 4 10 2 Internal Temperature Pressure and Humidity Sensors For most applications usage of the external temperature sensor should be the preferred option The PiccoTHP has an additional internal temperature sens
56. ng Probe The oxygen dipping probe for the Piccolo2 item no OPDIP20 consists of a stainless steel tubing 3mm in diameter and 200mm in length with a label attached to it and protective caps on both ends The attached label L indicates the instrument end IE of the dipping probe Instrument which needs to be inserted into the sensor port of the Piccolo2 see chapter 5 6 At the IE the bare optical fiber protrudes ca 1mm from the steel tubing L IE SE SC XA7 534 206 49 13 2PCRT 140038 The label L contains also the Sensor Code SC which needs to be entered into the Settings of the Pyro Oxygen Logger software see chapter 6 3 The sensor end SE of the dipping probe has to be inserted into the sample and CO is covered with a black coating optical isolation to prevent interferences with ambient light The full length of the dipping probe can be dipped into gaseous or liquid samples But it is important that no part of the Piccolo2 device itself is submersed into liquid samples For optimal performance and response time it is recommended to utilize the dipping probe in permanently stirred liquids this diffusion limitation is negligible in gas samples 14 5 3 Sensor Spots PyroScience offers a range of contactless oxygen sensors comprising sensor spots respiration vials and flow through cells The contactless sensors can be used with the Piccolo2 in combination with an optical fiber e g item
57. ngs have reached their steady state click on Set 0 and the actual oxygen sensor reading is taken for the o calibration If the es Calbrated at Ajij oxygen reading is out of the expected range a avon 20 000 warning will be shown offering the possibility to repeat the calibration A completed o calibration is indicated by the green indicator Calibrated at 0 Finally click on Finish for returning to the main caibrated at o9 window dek 52 000 49 7 1 4 Calibration Mode Custom without PiccoTHP The custom calibration mode is selected by clicking on the button Custom Mode This mode allows the user to combine freely all possible calibration types for the air calibration and the o calibration The air calibration type can be selected by clicking on the Air Calibration Selector The o calibration type can be selected by clicking on the o Calibration Selector Air Calibration Selector Calibration of Channel 1 3 Wait for Steady State J Air Calibration Set Air water oo Gg 02 SE oou srny et Calibrated d o Feed Temperature CO 122 0 get 00 20 700 External Temperature Sensor Pressure mbar Aio Elevation m i 200 Humidity RH Zoos 20 1 21 140 160 180 200 220 241 Oxygen 02 20 63 Temperature Sensor C Nan dphc 20 644 g Advance Aapa Harah gpho 53 200 o Calibration Selector The following air calibration types can be selected e Factory Air Calib
58. no P CFIB2 or alternatively with an optical fiber rod e g item no PICROD2 with the unique REDFLASH indicator with PET foil as weg dt i source carrier material and can be optical fiber K used for measurements in The sensor spots are coated pa etector infrared emission closed sample containers The itakata sensor spots have a diameter transparent of 5 mm with an optional DESEN sensor spot with REDFLASH dye optical isolation black e Coating They have a rough sensing surface ew whereas their backside is green smooth and shiny VW e The sensor spots can be glued with their backside on transparent clean and dry inner container walls plastic or glass wall thickness 0 20 mm using an appropriate adhesive e g transparent silicone based on acetic acid item no SPGLUE After the glue has dried the gaseous or liquid sample has to be filled into the container so that the sensing surface of the spot is completely covered and in contact with the sample For windows with a wall thickness of lt 10mm the Basic Spot Adapter ca 10x10x8 mm plastic cube ic item no SPADBAS needs to be fixed tightly or v i GP glued to the outer container wall at the spot position 15 The sensor signal can be read out using the optical fiber or optical fiber rod e g item no P CFIB2 or PICROD2 connecting the basic spot adapter with the Piccoloz2 pyroscie AL The optical fiber P CF B2 for th
59. or integrated behind the air holes of the housing for temperature measurements of the ambient air If used during calibration or during temperature compensation of the oxygen readings in a gas sample or air ensure that the temperature surrounding the PiccoTHP equals the ambient temperature in the gas sample air Further the PiccoTHP has integrated sensors for atmospheric pressure mbar and relative humidity of the ambient air RH These sensors allow a precise and easy sensor calibration see chapter 7 2 Furthermore the integrated pressure sensor can be used for automatic pressure compensation of the oxygen measurements see chapter 6 3 3 which is especially useful during long term experiments to account for weather changes and or for measurements above or below sea level These internal sensors can be activated also independently in the Options tab of the Piccolo2 Settings window see chapter 6 3 5 71 10 3 Automatic Temperature Compensation The external temperature sensor can be simply used for recording the temperature in the measuring setup However the temperature sensor is especially useful in order to compensate automatically the oxygen sensor signals for temperature variations in the setup The temperature compensation is needed due to two reasons e the luminescence of the REDFLASH indicator is temperature dependent and e the conversion of some oxygen units is dependent on the temperature In order to activate
60. precise measurements over the full range 0 21 O2 or 0 100 dissolved O2 DO For advanced users and applications a Custom Mode can be selected allowing the user to combine freely the possible calibration types for the air and the 0 calibration 4h 7 1 1 Calibration Mode Factory without PiccoTHP If the calibration mode Factory Calibration is chosen ensure that the correct sensor code has been entered in the Settings as displayed in 2 Adjust Calibration Conditions of the Oxygen Sensor Calibration window If the sensor code displayed is not correct click on Finish go to the Settings enter the correct Sensor Code and repeat the Factory Calibration After clicking on Finish the factory calibration is completed thereby returning to the main window 7 1 2 Calibration Mode 1 Point without PiccoTHP The calibration mode 1 point is selected by clicking on the button 1 Point This mode uses a manual calibration in an air calibration standard for adjusting the air calibration value The o calibration value is taken from the Sensor Code The preparation of appropriate air calibration standards is described in chapter 8 1 Depending on the air calibration standard used select Water DO for dissolved oxygen or Gas Phase The air calibration standard see also chapter 8 1 can be e ambient air of known humidity e water vapor saturated air or e air Saturated water 100 air saturation 45 P Oxygen Sensor Cali
61. puroscience sensor technology Piccolo amp PiccoTHP FIBER OPTIC OXYGEN METER amp CLIMATE MODULE USER MANUAL Document Version 2 03 Refers to Pyro Oxygen Logger Software version gt 3 2 The Piccolo2 is manufactured by PyroScience GmbH Hubertusstr 35 52064 Aachen Germany Phone 49 0 241 4004 555 Fax 49 0 241 4004 558 Email info pyro science com Internet www pyro science com Registered Aachen HRB 17329 Germany TABLE OF CONTENT RAA 5 SAFETY GUIDELINE gees 7 INTRODUCTION TO THE BICCOL OD EEN 9 SOFTWARE INSTALLATION 0ccccecceeeceeeeseeeeeeseeaeeeseeaeeneees 11 OXYGEN SENSOR TYPES isamaa a 12 5 1 ROBUSTIRROBE E 12 5 2 EE 14 5 3 SENSOR OPOTS E 15 5 4 FLOW THROUGH CES ceppe nn nn Re EEO 17 5 5 RESPIRATION VIALS eienn E 19 5 6 CONNECTING THE SENSORS AND OPTICAL FER 20 5 7 CLEANING AND MAINTENANCE OF THE DENSORS 21 THE SOFTWARE PYRO OXYGEN LOGGER ask 22 6 1 COUPLING OF PICCOLO2 AND PICCOTHP E 22 6 2 RER 23 6 3 EE etc ahaa ce chant dena isa 31 Deh i BOSIC DS CLUNGS ee 32 e Nee E 33 6 3 3 Conditions in the Sample eeben 35 6 3 4 Temperature Settings only with PiCCOTHP cccccceseeeee 37 G35 ee 38 6 4 OVERVIEW PANEL ONLY WITH DiccCOTHD 40 6 5 RAW DATA VINO eer 41 CALIBRATION OF OXYGEN SENSORS NENNEN 43 Ji CALIBRATION WITHOUT PICCOTHP 0 ccccseecceceeceecaeeceesaeeceesanes 44 7 1 1 Calibration Mode Factory without PiCCOTHP 22 22ccc000 45 7 1 2 Calibration
62. r 9 2 for more details For the dipping probe OPD P2o and for the optical fiber rods PICROD2 and PICROD3 please use a Fiber Length of o m Alternatively it is possible to Sensor Code ss label on sensor select Manual Background E A 7 5 3 0 2 0 o Compensation which is described in detail in chapter 9 2 Sensor Settings Basic 2 Advanced Finally the background Oxygen Measuring Time ms compensation can be completely i lS de activated by selecting No ee a dee SS Y g ger Amplification Background Compensation 0 vio 400x ip Fiber Length m 139 ea e Ne Bac cee 34 6 3 3 Conditions in the Sample The next step is the determination of the Conditions in the Sample which can be Water for dissolved oxygen DO or a Gas Phase Units umol l Conditions in the Sample External Temperature Sensor i Internal Temperature Sensor o Fixed Temperature CC 20 0 Temperature without PiccoTHP Internal Pressure Sensor The sample temperature has to be 2 Fied Pressure mbar 1013 j i Elevation 0 entered in Fixed Temperature z m 200 Salinity 9 1 0 0 Temperature with PiccoTHP It can be chosen whether the sample units 7 umol I temperature will be determined by the External Temperature Sensor of the Conditions in the Sample Water DO Gas Phase PiccoTHP or whether the measurements Po Ektemal temperature sensor Internal Temperature Sensor are performed at a Fixed Temperat
63. r of 3mm Due to its thicker diameter the PICROD3 cannot be used in combination with the basic spot adapter SPADBAS only with the included spot adapter sap Alternatively a custom fixation at the window of the sample container should be built to fix the P CROD3 at the sensor spot position 5 4 Flow Through Cells The flow through cells have an integrated oxygen sensor allowing online monitoring of oxygen in a gaseous or liquid sample pumped through the cell The flow through cells are available as a small version OXFTC and as a large version Z OXFTC2 The flow through cells are 17 equipped with luer lock connectors on both tubing ends allowing connection of gas tight tubings with ca 1 5 3 0 mm inner diameter to the OXFTC and with ca 4 0 5 5 mm to the OXFTC2 via the included luer lock adapters IE The flow through cell can be connected directly with an optical fiber item no PICFIB2 or PICROD2 to the Piccoloz AE oxygen meter The optical fiber P CF B2 consists of a black fiber cable 3 mm diameter 1m length with a label L attached to it The label L indicates the instrument end IE Instrument which needs to be inserted into the sensor port of the Piccolo2 see chapter 5 6 The adapter end AE of the optical fiber P CF B2 needs to be inserted into the flow through cell and fixed with the white clamping screw Pyroscience 18 5 5 Respiration Vials w Based on the contac
64. ration refer to chapter 7 1 1 e Air Calibration refer to chapter 7 1 2 And for the 0 calibration the following types can be selected e Factory 0 Calibration refer to chapter 7 1 1 e o Calibration refer to chapter 7 1 3 50 7 2 Calibration with PiccoTHP To calibrate a sensor click on the button Calibrate in the main window Note that during data logging this button cannot be used until Log to File was stopped A dialog window Oxygen Sensor Calibration opens in which the calibration mode can be selected Five standard modes can be chosen for the calibration Factory calibration for a quick rough calibration taking the 0 and the air calibration values from the sensor code advised only for rough measurements 51 1 Point in Ambient Air taking the o calibration value from the sensor code and the air calibration value from a manual calibration in ambient air for precise measurements around 21 O2 1 Point in Water or Humid Air taking the 0 calibration value from the sensor code and the air calibration value from a manual calibration in air saturated water or water vapor saturated air for precise measurements around 100 air saturation 2 Point in Ambient Air taking the o and the air calibration value from a manual calibration for precise measurements over the full range 0 21 O2 This mode uses ambient air for determining the air calibration value 2 Point in Water or Humid Air taking the o and t
65. re again this configuration with Load Setup giving you calibrated sensors ready for the measurement Note that for this procedure both computers must operate with the same logger version Save Setup and Load Setup might be also useful if e g a Piccolo2 should be used repeatedly with several oxygen sensors Initially 27 each sensor needs to be calibrated only once and the configuration of each sensor is saved with Save Setup If later on a measurement should be performed with a specific sensor it is only necessary to load the configuration for this sensor with Load Setup All current settings and calibration data are automatically saved when closing the Pyro Oxygen Logger software and are automatically loaded again at the next startup The present setup loaded is shown under the Settings button The button Flash Logo causes a short flashing ca 1 2 sec of the status LED of the Piccolo2 followed by a short LED flashing of an optionally coupled PiccoTHP Several Piccolo2 meters can be connected to the PC in parallel and multiple measurements can be performed by opening the Pyro Oxygen Logger software a number of times corresponding to the number of connected Piccolo2 meters The different windows operate completely independent of each other and are assigned to exactly one Piccolo2 This allows measurements in different setups at the same time The flashing of the status LEDs can help to assign a specific logger window to the corresponding P
66. ry calibration is taken for the 0 O2 calibration point Ensure that the r code is entered in the Custom Mode Sensor Code YA7 547 199 air calibration value The preparation of appropriate air calibration standards is explained in chapter 8 The o calibration value is taken from the Sensor Code Note that this calibration mode will automatically read the atmospheric pressure from the internal pressure sensor in the 55 PiccoTHP The calibration standard with the air saturated water must be therefore exposed to the same atmospheric pressure which is given in typical applications Insert the oxygen sensor connected to the Piccolo2 and the temperature sensor connected to the PiccoTHP into the flask containing the air saturated water or water vapor saturated air Ensure that the correct sensor code has been entered in the Settings If the sensor code displayed in the Factory o Calibration is not correct click on Finish go to the Settings enter the correct Sensor Code and repeat the calibration Then the temperature of the air saturated water or water vapor saturated air needs to be determined Either a Fixed Temperature is adjusted manually or the temperature is read from the temperature sensor connected to the PiccoTHP by selecting External Temperature Sensor Wait now until the oxygen sensor reading is stable by observing the graph If External Temperature Sensor was selected ensure also stable temper
67. sensor reading is taken for the air calibration If the oxygen 9 7 LE reading seems to be out of the expected range a usa warning will be shown offering the possibility to repeat the calibration A completed air calibration is indicated by the green indicator Calibrated at Air For the Factory 0 Calibration no further steps are necessary refer also to chapter 7 1 3 Ensure that the correct sensor code has been entered in the Settings 47 7 1 3 Calibration Mode 2 Point without PiccoTHP The calibration mode 2 point is selected by clicking on the button 2 Point In this mode both the air calibration value and the 0 calibration value are determined in specially prepared calibration standards The preparation of appropriate o and air calibration standards is explained in chapter 8 O Oxygen Sensor Calibration me Calibration of Channel 1 1 Choose Calibration Mode 2 Adjust Calibration Conditions 3 Wait for Steady State 4 Calibrate Air Calibration 21 en 02 Water oo T97E lt s Serie _ Not calirated air 20 95 Fixed Temperature CC S 122 0 dphi100 20 700 External Temperature Sensor Pressure mbar A 1013 Elevation m 200 l o Humidity RH J 100 0 re 60 80 100 l 0 Calibration Oxygen 02 20 61 Net colada dphi0 53 200 U Fixed Temperature CC 200 Temperature Sensor CC Nah External Temperature Sensor Place the sensor in water mixed with sodium sulfite or in water
68. settings see chapter 6 3 2 The actual Compensation Temperature i e the temperature used for calculating the oxygen values is shown in the temperature display TD in units of degree Celsius C The display of the data in the charts can be changed by different chart tools arranged underneath the chart recorder Le S Seconds s poe poe Ha R Autoscroll Time Scale The button with the magnifying glass offers different zoom options After clicking the button with the hand the user has the possibility to click onto the chart and move the whole area while keeping the mouse button pressed The unit of the x axis can be changed with the selector Time Scale The time scale can be displayed in Seconds s Minutes min Hours h Relative Time HH MM SS Absolute Time HH MM SS Absolute Time and Date and numbered Data Points The scales of the y and x axis can be adjusted by clicking on Adjust Scales opening a pop up window The upper Maximum and lower limits Minimum and the Increment of the Y Scales Oxygen and of the X Scale Time can be changed by clicking on the 25 respective selectors or by double clicking directly onto the field and entering the values manually changing these parameters will automatically de activate the autoscaling Autoscaling for all axes can be activated with the Autoscale button The arrow in the button turns from dark green to light green indicating that Autoscale is
69. solution Accuracy 40 to 125 C 0 02 C 0 3 C Internal Pressure Sensor 300 to 1100 mbar 0 06 mbar Range Resolution Accuracy typ 3 mbar Internal Humidity Sensor o to 100 rel humidity RH Range Resolution Accuracy 0 04 RH typ 0 2 RH Ho 11 3 Troubleshooting How to respond to the warnings shown in the Pyro Oxygen Logger Signal Too High Too much ambient light exposed to the sensor or amplification is too high or LED intensity is too high darken the surrounding and or decrease Amplification in the Advanced settings and or decrease LED Intensity in the Advanced settings Low Signal Sensor signal is too low check whether the sensor cable is connected increase Amplification in the Advanced settings and or increase LED Intensity in the Advanced settings replace sensor the tip might be broken bleached Bad Reference Internal problem of the electronics contact PyroScience 81 11 4 Measuring Principle The REDFLASH technology is based on the unique oxygen sensitive REDFLASH indicator showing excellent brightness The measuring principle is based on the quenching of the REDFLASH indicator luminescence caused by collision between oxygen molecules and the REDFLASH indicator immobilized on the sensor tip or surface The REDFLASH indicators are excitable with red light more precisely orange red at a wavelength of 610 630 nm and show an oxygen dependent luminescence in
70. splay D and in a chart recorder C in the chosen oxygen unit UD The color and appearance of each graph can be changed by clicking on the color control CC The description of the sensor as defined in the Settings by the Sensor Code is shown in the description display DD The Signal Intensity SI of the oxygen sensor is shown as a horizontal indicator bar just underneath the numeric display D A reasonable oxygen sensor shows signal intensities well above 20 typically 50 500 If the signal intensity drops below 50 the indicator bar SI turns gradually from grey to red indicating that the sensor might get degraded soon Most sensors will still work even at a signal intensity of ca 10 yet with increased noise But at even lower signal intensities the warning Low signal will appear in the warning display WD see below and chapter 11 3 At a signal Note Exceptions are trace oxygen sensors During the 100 calibration at 21 O2 these sensors show naturally a very low signal intensity as low as 10 But the signal intensity will strongly increase when a trace oxygen sensor is applied within its specified range of o 10 O2 24 intensity lt 5 the display changes to NaN Not a Number indicating that the signal is too low and the sensor needs to be replaced or moved to another position on the sensor spot Note that the signal intensity can be optimized by varying the LED Intensity or the Amplification in the advanced
71. temperature sensor On the right hand side of the channel tab a graph can be activated showing the dphi and Signal Intensity mV in the graph default setting Plotting of additional parameters can be activated by clicking on the small rectangular button next to the color control of the respective parameter dphi 20 881 Oxygen uM 284 984 stoohigh Oxygen hPa 208 322_ temp sensor intiized Oxygen air sat 100 465_ Oxygen 02 20 564 Signal Intensity mV 552 68 Ambient Light mv 68 82 External Temp CC mm Internal Temp CC Inn Pressure mbar 0 00 Humidity 0 00 Analog In mV 0 00 0 o we 150 200 250 300 350 400 450 50 550 60 552 76 A 348 98 D Hew Data Points TAE ITTI SI ag d OG i A le A fd dy 313 89 B 135 63 E w ia pA 49 584 c 28 293 F H KA Show Configuration 42 7 Calibration of Oxygen Sensors This chapter describes the possible calibration modes for oxygen sensors connected to the Piccolo2 using the logger software Pyro Oxygen Logger Please note that the possible calibration modes change if the climate module PiccoTHP is connected in parallel to your PC For many calibration modes the temperature in the calibration standard the atmospheric pressure and the relative humidity of the ambient air are important parameters ensuring a high precision of the calibration Without a connected PiccoTHP it is on the user s authority to
72. tless sensor technology from PyroScience ready assembled respiration vials with an integrated optically isolated oxygen sa sensor stripe are available The glass respiration vials are available with 4ml or 25 l 20ml volume item no OXVIAL4 OXVIAL20 Separately available adapter rings item no ADVIAL4 ADVIAL20 allow an easy fixation of an optical fiber item no PICFIB2 which connects the respiration Sech PICFIB2 aL vial to the Piccolo2 pss E e S A label L attached to the optical fiber PICFIB2 indicates the instrument end IE of the optical fiber 4Instrument which AE needs to be inserted into the sensor port of the Piccolo2 see chapter 5 6 The adapter end AE of the optical fiber P CFIB2 needs to be inserted into the respective adapter ring and fixed with the clamping screw allows The built in sensor stripe oxygen measurements at different p Pyroscience heights within the vial by simply sliding picriez the adapter ring up and down Therefore it is e g possible to measure oxygen both in the liquid and in the gaseous J headspace within the same vial pyroscience Forhigh precision respiration rate py measurements the respiration vials n would be typically placed within a water bath with a regulated constant temperature at 0 1 C precision If higher temperature fluctuations are expected the oxygen measurement should be temperature compensated using the climate module P
73. tream for about 10 minutes nitrogen gas through the water You might speed up this process by first boiling the water and thereby removing all dissolved gases and then stream the nitrogen gas during cooling through it Insert the 71 oxygen sensor and the temperature sensor into the flask let it equilibrate and perform the calibration as described in chapter 7 8 2 3 Nitrogen Gas Flush 100 nitrogen gas through a glass flask e g Duran flask with a lid prepared with holes for inserting the oxygen sensor and a temperature sensor Ensure that all air has been replaced by the nitrogen gas before performing the calibration Insert the oxygen sensor and optionally also the temperature sensor into the flask let it equilibrate and perform the calibration as described in chapter 7 72 9 Calibration of Contactless Sensors For preparing a setup with contactless oxygen sensors please refer to the chapters 5 3 5 5 9 1 Calibration Procedure In general the calibration procedure for contactless sensors e g sensor spots flow through cells respiration vials is the same as for the robust and dipping probe described in the chapters 7 and 8 However if a 1 point or a 2 point calibration should be performed the calibration standards have to be filled directly into the vessel containing the sensor spot into the tubing of the flow through cell or into the respiration vial If Ambient Air is used for the air calibration st
74. ure en 0 Fixed Temperature C 20 0 If External Temperature Sensor is o Internal Pressure Sensor selected automatic temperature xed presure mba 101 compensation of the oxygen sensor Elevation m 200 readings is activated see chapter 10 3 Salinity oi joo If Fixed Temperature C is chosen the temperature of the environmental sample has to be determined with an external thermometer kept constant and entered manually This Compensation Temperature either entered manually at constant temperature conditions or measured by the external 35 temperature sensor of the PiccoTHP will be displayed in the main window TD Pressure without PiccoTHP The atmospheric pressure e g read from a separate barometer can be entered manually at Fixed Pressure mbar Normal conditions refer to 1013 mbar default If the actual atmospheric pressure cannot be determined on site it is also possible to enter the Elevation m above sea level For this click on Elevation and enter the actual elevation in meters This procedure will only calculate the average atmospheric pressure for this elevation therefore this option is less precise than measuring the actual atmospheric pressure Pressure with PiccoTHP In combination with the PiccoTHP it is adviced to select Internal Pressure Sensor then the actual atmospheric pressure is measured before each single oxygen measurement and the oxygen measurement is automatically
75. xygen sensor Based on the Fiber Length m entered in the Settings see chapters 6 3 1 and 6 3 2 a background signal for compensation is estimated automatically by the Pyro Oxygen Logger software So the user usually does not notice the background compensation at all For normal applications this should be the preferred procedure But for precision applications and especially for low signal intensities e g lt 20mV a manual background compensation can be performed alternatively by the user For this Manual Background Compensation must be selected in the Advanced Settings see chapter 6 3 2 After opening the calibration window by clicking on Calibrate a separate Background Compensation window will open automatically Q Background Compensation Here the background fluorescence of the connected optical fiber can be compensated For this it is important that 74 The instrument end of the Optical Fiber is connected to the Piccolo2 see chapters 5 3 5 5 and the adapter end of the Optical Fiber is not attached to the sensor i e disconnect the adapter end from the spot adapter adapter ring or from the flow through cell Then wait for steady state and press the button Take Actual Values Alternatively the button Keep Last Values can be used if the sensor spots are re calibrated with the same optical fiber which was background compensated before Then the last values for the background compensation are kept It is

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