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1. sss 17 3 2 3 Rawdabd control oco ior Een ORAN ma dh i dd 17 3 24 Processing and Automatic Quality control of data 18 3 3 Filter pad absorption curre nente ca x Eee cont on dees ke eU ati sated aamevad teens 19 3 3 1 Instrumentation checks and calibrations susesss 20 3 3 3 Documentation of deployment parameters and sample preparation 22 3 3 3 Raw data controlere eiiiai a M UR dd MM MH I 22 3 3 4 Processing and Automatic Quality control of data 23 3 4 Laboratory CDOM 4 enie aeaaeae ct x Een aon dante ee san vene eaters dee 23 3 4 1 Instrumentation checks and calibrations ssssssss 24 3 4 2 Documentation of deployment parameters and sample preparation 24 3 4 3 Raw data Conttol coo co e pr Wo a tM op de uM duc d 25 3 4 4 Processing and Automatic Quality control of data 25 3 5 Suspended particulate matter by gravimetric methods 26 3 5 1 Instrumentation checks and calibrations sussss 26 3 5 2 Documentation of deployment parameters sess 26 3 5 3 Raw data control siue seo ee rio ta ER bc ndi aa NS epi sm uM ax 27 3 5 4 Quality control of data som etr ER bcr r p er tir lm p Mte S ieu 27 3 6 Phytoplankton Pigments using HPLC
2. Alternative payments terms are available at an Customer additional charge Prices shown on this sheet are exclusive of VAT and any local taxes 60057 60020 AUTOSAMPLER UHP ACCELA 83492 Coverage UNITY ESSENTIAL SUPPORT PLAN CHROM LC 1 376 96 GBP 60057 60010 PUMP UHP ACCELA DISC USE 60057 60111 83303 Coverage UNITY ESSENTIAL SUPPORT PLAN MSPEC LC MS 1 682 72 GBP 60057 60050 DETECTOR ACCELA PDA 5CM LIGHTPIPE 81687 Coverage UNITY ESSENTIAL SUPPORT PLAN CHROM LC 54912 GBP Support Plan Sub Total 3 608 80 GBP Support Plan Total 3 608 80 GBP Thermo Fisher Scientific looks forward to providing service on those instruments ified above subject to the terms and conditions stated on the attached document If you have questions please call 4100888 to contact your Support Plan specialist ACCEPTANCE OF SUPPORT PLAN Plymouth Marine Laboratory Signature Date apan cc Thermo SCIENTIFIC Thermo Fisher Scientific ts the trading name of Thermo Electron Manufacturing Lid Page 2 of 4 30 Appendix 7 3 List of standards available from DHI 1 amp hex fucoxanthin PPS 19HEX 25m 1 175 157 74 a carotene PPS ACAR 25m 1 175 157 74 a cryptoxanthin PPS ACRYP 25m 1 042 139 91 Alloxanthin PPS ALLO 25m 1 042 139 91 Antheraxanthin PPS ANTH 25 mi 2 911 390 69 Aphanizophyll PPS APHAN 25mi 1 175 157 74 Astaxanthin PPS ASTA 25m 1 042 139 91 p Carotene PPS BCAR 25mi 1 175 157 74 B Cryptoxanthin PPS BCRYP 25mi 1 042 1
3. esses 14 Deployment a oaa eu eire eos Sas cabs cee eG Sault othe bach Ne MIS a dep qutt Ca a a Kou RUE 14 gie ino MM TE RS EERS 14 Upkeep and Maintenance ud eo deor tp i esempi Tesi ee paul a ia pem dE geben ue dp Rada deae 15 Data PLOCOSSIIE s te da ocn Ret ebd tane deis ed Todes a face AR od edendi da dla eed 15 Attenuation coupling s etos e e oU oes dices REIN aE AAA APR IRR FEMME T qUEMS 15 jsuis PEDEM MM tem 15 In situ spectral Beam Attenuation coefficient c z Ini obs eee 16 In situ spectral Absorption Coefficient a z A m ELE MES 16 Instrument Calibration and Quality Assurance eseeeeeeeeeee n 16 Mounting and Deployment of the instrument esee 16 Field p re Wwat rcalibration dude teu de ob RR nhu qoibns nid e Toi UE 16 Methodology and processing description sseeeesssseessesessessesressesresserressesresrersesressesees 17 Temperature and salinity corrections iie irren inpr ee ia nt era pou ai peer ene 17 Scattering corrections of the absorption coefficient sse 18 IESCA satellite validation protocols 07 027 FR ISECA 2 Primary Quality Checks eure vetro D epatis rette ene RA URL ed beasties pee ete os 18 Calibration coefficients soe Lerner i OP c EE rte tu oo ERE E pec E A EE 18 Limitations eee costs obithadaards ceder bes n dcr tiae bes papi rede d De a FIRES ADDE Dee 19 Aia RM P 19 Coloured dissolved organic material m TI DUE TEL
4. esssssesseeeerenennen nennen 28 REFERENCES ineen aT EEE E E E AETA a EE a aaa 28 1 INTRODUCTION Investigations of marine environment often require complex and large national and international research programmes Such programmes need a data management plan which includes details about the data quality control in addition to a scientific measurement plan The objective of data quality control is to ensure the data consistency within a single data set and within a collection of data sets and to ensure that the quality and errors of the data are apparent to the user so that there is sufficient information to assess its suitability for a task This quality control procedure guides data providers and managers to rigorously test the quality of the data that is sent to the database Only after these tests should the data be included in a database or distributed to users via international or national data exchange This document is an extension of previous work within the EU project REVAMP 1 and follows the general recommendations from IOC IODE 2 Specific quality control procedures in bio optical data have been obtained from existing literature on NASA databases like SeaBASS 3 and NOMAD 4 databases Other procedures have been obtained from the scientific literature and in particular from the synthesis in the UNESCO book on monitoring coastal harmful algal blooms 5 The current report is divided in two parts first a general overview
5. e If samples are to be run more than 24 hrs after collection then samples should be flash frozen and stored in flat containers e g petri dishes petri slides in liquid nitrogen Dry shippers are favored for the transportation of samples but dry ice will suffice for short distances 36 hr duration For further details on sample storage see section on HPLC p Measurements procedure The methodology is described in Tassan and Ferrari 1995 with the following modifications e The Autozero of the instrument should be made with free entrance ports using high grade perfectly balanced reflecting plates on the exit ports these can be replaced by standard spectralon plates for the following measurements Performing the Autozero with filters on the entrance ports is not considered a good practice because of the difference that may occur in filter transmittances Baseline flatness using integrating sphere should be at least 0 004 A units e Depigmentation using NaClO is recommended Bleaching by Methanol is not advised as phycobilins and eukaryotic pigments are not extracted and some loss of the sample can occur The bleaching concentration of NaClO can be 1 active chlorine Tassan amp Ferrari 1995 or 0 1 active chlorine Tassan et al 2000 The choice of active chlorine solution depends on the dominant particles or species in the sample If the sample has a high detritus content 0 1 active chlorine is recommended since a 1 96 ac
6. 0 25 0 245 0 24 T T T 04 01 01 04 10 06 07 09 04 12 12 14 Figure 6 Monitoring with time the change in absorbance A at different wavelengths at PML measurements of the F2 cell lowest intensity The range of change per year is between 0 16 and 0 01 per year for the PML spectrophotometer The lowest absorbance filter F2 shows the greatest change and spectral differences are also observed with the greater rates of change in the blue 20 o ma o o O c an o e R t Ic o9 v9 o 2 ra o un 2 lt q o v9 oD o 4 o x o o 500 550 Wavelength nm o o e Figure 7 Spectral variation of the of change of Absorbance per year for the three standards used at PML In addition to the calibration absolute and tracking some quality check of the instrument should be done previous and during sampling A baseline should be recorded and checked for spectral trends and or variations in absorbance greater than 0 005 If this happens additional autozero of the instrument and possibly checking the alignment of the mirrors around the integrating sphere A INSTRUMENT METHOD CHECK 1 Sensor output check Before measurement Y N NOTES After measurement Y N NOTES Instrument checked Y N NOTES Instrument model and Model Serial N NOTES Serial N 2 Calibration Date of factory calibration dd mm yyyy NOTES Last labor
7. Date prepared Name Do this in an externally venting fume cupboard Wear gloves and glasses Use all solvents at room temp Preparation of eluent B Take a 250 mL measuring cylinder and rinse with methanol Measure 200 mL methanol and transfer to a pre rinsed 1L HPLC flask Measure 600 mL acetonitrile and add to flask Measure 200 mL acetone and add to flask Stopper flask and mix by inversion Add HPLC cap and seal with foil Label with B MeOH ACN acetone 20 60 20 Date prepared Name Do this in a fume cupboard Wear gloves and glasses Any modifications to this protocol must be approved and recorded 21 6 0 Spreadsheets 6 1 Spreadsheet 1 RS tracker MgDVP Chl c2 Viola Hex Zeax Lut Dv chl a Chl a 11 10 2012 10 12 2012 11 12 2012 17 01 2013 18 01 2013 31 01 2013 01 02 2013 25 04 2013 25 04 2013 23 05 2013 PMd 10 12 PMd 10 12 PMd 10 12 PMd10 12b PMd10 12b PMd10 12b PMd10 12c PMd10 12d PMd10 12e pigmixdil RT 10 40 Width 0 59 RT 10 88 Width 0 62 Rs 0 79 RT 20 78 Width 0 45 RT 21 21 Width 0 49 Rs 0 91 RT 27 40 Width 0 26 RT 27 60 Width 0 28 Rs 0 74 RT 33 60 Width 0 28 RT 33 89 Width 0 37 Rs 0 91 9 94 0 57 10 39 0 60 0 76 20 20 0 44 20 62 0 52 0 88 27 09 0 29 27 30 0 30 0 74 33 61 0 30 93102 0 39 0 90 NMP 9 98 0 61 10 43 0 60 0 74 20 25 0 45 20 67 0 53 0 87 27 14 0 28 27 36 0 31 0 73 33 65 0 30 33 97 0 39 0 91 6 2
8. 0 39 20 30 0 47 0 84 27 00 0 30 2722 0 32 0 71 33 45 0 28 33 74 0 36 0 92 Mean 0 74 0 88 0 71 0 90 08 10 2012 09 10 2012 11 10 2012 10 12 2012 11 12 2012 17 01 2013 18 01 2013 31 01 2012 01 02 2012 25 04 2013 25 04 2013 23 05 2013 pigmix dil pigmix dil pigmix_dil PMd10 12 PMd10 12 PMd10 12 PMd10 12b PMd10 12b PMd10 12b PMd10 12c PMd10 12d PMd10 12e 7 27 9 92 10 36 10 83 13 4 14 08 17 33 18 21 19 22 19 95 20 75 21 18 21 92 23 3 24 66 25 82 26 7 27 39 27 59 29 02 29 58 32 17 32 49 33 6 33 89 34 25 36 03 36 82 37 01 10 94 5 09 4 09 6 50 3 12 7 32 9 97 10 41 10 89 13 47 14 16 17 41 18 29 19 29 20 02 20 83 21 26 22 00 23 38 24 73 25 88 26 76 27 44 27 64 29 07 29 58 32 22 32 54 33 66 33 96 34 33 36 08 36 92 37 12 10 97 5 09 4 06 6 52 3 16 7 32 9 94 10 40 10 88 13 45 14 13 17 37 18 24 19 25 19 97 20 78 21 21 21 93 23 32 24 68 25 83 26 71 27 40 27 60 29 03 29 49 32 17 32 49 33 60 33 89 34 26 36 03 36 82 37 01 10 93 5 08 4 08 6 50 3 12 NMP 6 97 9 52 9 94 10 39 12 89 13 53 16 77 17 65 18 66 19 39 20 20 20 62 21 35 22 75 24 12 25 35 26 35 27 09 27 30 28 81 29 49 32 10 32 44 33 61 33 92 34 31 36 20 37 04 37 24 10 68 5 10 4 34 6 83 3 33 7 05 9 59 9 98 10 43 12 92 13 58 16 82 17 70 18 70 19 44 20 25 20 67 21 43 22 81 24 17 25 41 26 40 27 14 27 36 28 87
9. 2001 In situ studies have shown that the different instruments compare well with measurements of the whole volume scattering function Berthon et al 2007 More recently several commercial backscatter meters have been developed and are available from Hobilabs and WETLABS The HydroScat 6 manufactured by Hobilabs measures scattering at centroid angle of 140 and at many fixed wavelengths The ECO VSF 3 manufactured by WETLABS measures 0A at single wavelengths 450 530 650 nm but at three centroid scattering angles 100 120 150 Both of these instruments will be utilized during the ISECA contract These sensors measure a weighted integral of radiance scattered from a working volume defined by the intersection of illumination source beam and angular field of view of the detector Mueller et al 2000 The backscattering coefficient m is calculated from b A 2z b0 a sinaa6 z 2 The Hobilabs instrument The following information has been taken from Hydroscat 8 Manual 2010 Instrument description The Hobilabs Hydroscat 6 is a hyperspectral instrument Fig 2 that measures p 02 at six wavelengths and at 140 It also makes auxiliary measurements of fluorescence The beam from the LED goes through a lens to adjust its divergence then through a prism that bends the beam before it enters the water IESCA satellite validation protocols 07 027 FR ISECA 11 Figure 2 Hobilabs Hydroscat 6 Multispectral backscatt
10. 29 38 32 14 32 48 33 65 33 97 34 37 36 26 37 10 37 30 10 65 5 11 4 33 6 83 3 34 NMP 7 08 9 59 10 01 10 47 12 93 13 60 16 81 17 69 18 70 19 43 20 22 20 64 21 40 22 78 24 14 25 36 26 35 27 09 27 30 28 79 29 49 32 01 32 33 33 46 33 76 34 13 36 26 36 73 36 92 10 61 5 09 4 31 6 68 3 15 7 04 9 58 10 02 10 47 12 95 13 62 16 86 17 73 18 75 19 48 20 28 20 70 21 48 22 83 24 20 25 41 26 39 27 12 27 33 28 82 29 30 32 03 32 36 33 50 33 80 34 18 36 30 36 80 37 00 10 69 5 10 4 29 6 68 3 20 23 NPC 7 18 9 74 10 16 10 62 13 14 13 79 17 05 17 93 18 46 19 66 20 47 20 90 21 66 23 03 24 40 25 58 26 51 27 21 27 42 28 88 29 49 32 09 32 42 33 58 33 88 34 26 36 12 36 95 37 15 10 74 5 10 4 19 6 67 3 28 NMP SMP 7 08 7 84 9 58 10 61 10 03 11 06 10 49 11 55 13 00 14 26 13 67 14 96 16 92 18 23 17 80 19 10 18 79 20 11 19 53 20 83 20 34 21 61 20 77 22 07 21 53 22 90 22 90 24 16 24 26 25 49 25 47 26 41 26 43 27 13 27 15 27 74 27 36 27 91 28 83 29 28 29 49 29 49 32 00 32 39 32 33 32 73 33 44 33 92 33 73 34 24 34 10 34 46 35 90 36 64 36 69 37 48 36 84 37 70 10 72 1126 5 10 5 06 4 25 3 58 6 58 6 50 3 10 3 46 7 79 10 56 11 04 11 53 14 21 14 92 18 19 19 06 20 08 20 80 21 58 22 04 22 88 24 13 25 46 26 40 27 14 27 75 27 92 29 93 29 49 32 39 32 75 33 93 34 24 34 46 36 63 37 44 37 65 1
11. Chl a absorbance Zero absorbance reading Chl a concentration spec g L Chl a concentration spec ng L Chl a concentration spec ng uL Baseline data WS data Date analysed by HPLC HPLC Method HPLC peak areas Injection Number Std dev 96std dev Injection volume sample water uL vol mixer uL vol sample uL ng injected ng adjusted purity Response factor ng adj peak area Calibration value 96 change from Calibration value Chl a stock protocol Chl a working standard protocol Quantification of working standard Determination of RF protocol Chl a concentration calculation Solvent 25 04 2013 10096 acetone 25 04 2013 9096 acetone 25 04 2013 Perkin Elmer Lamda 800 G18 9096 Acetone Wavelength nm 0 0259 663 0 0002861 750 0 00030 298245 0 298 d pigments 20130425 90aceton d pigments 20130425 chlaws 25 04 2013 ZAPATA Peak area 440 nm Area other peaks 440 nm 425074 3960 424796 4022 423581 4179 412567 4161 435332 3936 427710 3931 424843 4032 7356 1 73 25 80 200 5 33 5 28 1 242E 05 1 196E 05 3 82 Labbook4 P104 Labbook4 P105 Labbook4 p106 Labbook4 p91 Labbook4 p106 24 Lab book page 4 104 4 105 4 106 Purity 99 07 99 05 99 01 98 99 99 10 99 08 99 05 6 4 Spreadsheet 4 Injection reproducibility Date Peak 24 07 2012 Int std 24 07 2012 chl a ws 26 07 2012 chla ws 27 07 2012 chla ws 30 07 2012 chla ws 07 08 2012 chla ws 09 08 2012 chla ws 13 08
12. Gould R Kahru M Kishino M Maske H Moisan T Moore L Nelson N Phimney D Reynodls R Sosik H Stramski D Tassan S Trees C Weideman A Wieland J Vodacek A 2000 Determination of spectral absorption coefficients of particles dissolved material and phytoplankton for discrete water samples NASA Tech Memo 209966 in GS Fargion and JL Mueller Eds Ocean Optics Protocols for Satellite Ocean Colour IESCA satellite validation protocols 07 027 FR ISECA 21 Sensor Validation Revision 2 NASA Goddard Space Flight Center Greenbelt Maryland pp 125 153 Mueller J L and R W Austin 1995 Ocean Optics Protocols for SeaWiFS Validation Revision 1 NASA Tech Memo 104566 Vol 25 S B Hooker and E R Firestone Eds NASA Goddard Space Flight Center Greenbelt Maryland 67pp Pegau W S and Zaneveld J R V 1993 Temperature dependent absorption of water in the red and near infrared portions of the spectrum Limnol Oceanogr 38 188 192 IESCA satellite validation protocols 07 027 FR ISECA 22 Pigments Concentration by High Performance Liquid Chromatography mg m or ug r The high performance liquid chromatography HPLC method described here JGOFS 1994 aims at separating the following phytoplankton pigments chlorophyll a chlorophyll b chlorophyll c chlorophyllide a fucoxanthin 19 butanoyloxyfucoxanthin 9 hexanoyloxyfucoxanthin zeaxanthin alloxanthin peridinin diadinoxanthin diatoxanthin carotene The methods
13. Instrumentation See section on In vivo Absorption Spectra of pigmented and non pigmented Particulate Matter p 4 Instrument Calibration and quality assurance e Spectra are visually checked for high background such as high absorption values in the red part of the spectra and abnormal slopes e Pure water such as Millipor MilliQ Alpha Q and Barnstead Nanopore is recommended Ensure when carrying out optical density measurements of CDOM at sea that this water 1s available otherwise preparation of pure water prior to field work is recommended e The response of the spectrophotometer should be verified with Holmium Oxide filters especially at 412 amp 443 nm Filtration and Storage It is essential to minimize contamination of the samples by organic materials and the samples should be protected from light to reduce sample degradation e Wash hands with soap and water to avoid contamination of samples e Use 0 2 um polycarbonate filters Whatman Nucleopore are recommended e Filtration apparatus all glass a funnel flask and borosilicate filter support and clenching aluminium pliers Individual vacuum control of each sample for accurate pressure regulation and direct filtration to clean bottles is required e Mount filters on funnel and filter 100 mls of purified water through filter and discard water e Sea water should be collected into all glass brown bottles direct from Niskin bottles or equivalent Pre wash dark bottle three time
14. J P De Blauwe E D Vreker P Y Deschamps M Knockaert B Nechad A Pollentier P Roose D Saudemont and D v Tuyckom 2002 Preliminary validation of MERIS water products for Belgian coastal waters Envisat Validation workshop 9 13th December 2002 Frascati European Space Agency Ruddick K De Cauwer V Park Y J 2006 Seaborne measurements of near infra red water leaving reflectance The similarity spectrum for turbid waters Limnology and Oceanography 51 1167 1179 IESCA satellite validation protocols 07 027 FR ISECA 42 SIMBADA method The MERIS reflectance p A as defined by L 4 Po EA is calculated from sequential above water measurements of the vertically polarised component of radiance from the water surface L A and sun radiance L A L A is calculated from Z A after correction for residual air sea interface reflection and downwelling irradiance E A is calculated from L 4 using an atmospheric model This method corresponds to Method 3 of Mueller et al 2000 Full details of the method and processing can be obtained from the Laboratoire d Optique Atmosph rique of the University of Lille France Results of the method as used for MERIS Validation are presented in Ruddick et al 2002 Figure 11 View of SIMBADA showing foreoptics Instrument description As described in the SIMBADA User s Guide http www loa univ lillel fr recherche ocean color src the SimbadA
15. Sample 12 Sample 13 Sample 14 Sample 15 Sample 16 Sample 17 Sample 18 14 5 3 Protocol 3 Determining the resolution of critical pairs in the mixed standard and updating retention time tracker Determining the resolution of critical pairs in the mixed standard Notes and updating retention time tracker In Accela offline open the chromatogram of the mixed standard Open the processing method from today s analysis folder From the top menu select analysis and analyse BW N R Open the peak table and enter the retention time of each peak from the chromatogram Ui Save the processing method Re analyse the chromatogram 6 Copy the retention times from the peak table to the retention time tracker spreadsheet Update the average std deviation and 96 change columns Print sheet and stick in lab book 7 Look at the integration of the peaks MgDVP Chl c2 viola hex zeax lut DVChl a and Chl a Ensure peak width is accurately defined If integration changed select Analyse 8 Change chromatogram annotation to retention time name and width Note retention time and width of critical pairs and input data to RS tracker spreadsheet Update average std deviation and 96 change columns If change is greater than 5 for more than 2 sets of critical pairs or is greater than 9 for one critical pair take corrective action to improve chro
16. The windows should be periodically inspected for contamination Pressure transducer If your HydroScat 6 is equipped with an oil filled pressure reservoir and capillary tube check the tube occasionally to see that it contains oil It need not be completely full but the oil meniscus should be visible For the HydroScat 6 without oil reservoirs the pressure transducer is located under a black plastic cap flush with the rear endcap with four small drain holes Rinse the sensor with fresh water by gently spraying it into the drain holes Data Processing The HydroScat software HydroSoft allows you to save calibrated data automatically at the time you collect or download data Raw data files can also be processed by converting raw hexadecimal data to decimal form without calibrating them An IDL program has been designed in PML to implement the sigma correction that takes into account the effect of the attenuation of the backscattered light from the particle This includes the use of absorption in situ and attenuation from a Wetlabs ac9 deployed simultaneously to the Hobilabs Hydroscat 6 Calibration coefficients HydroScat data are transmitted in a partially processed hexadecimal form which must be converted to calibrated units The coefficients required for this conversion are unique to each instrument and may be revised from time to time when the instrument is recalibrated HydroSoft requires an appropriate calibration to be loaded before i
17. interpolation and binning to an equally spaced grid is recommended L2 For the WCO application one sample is defined by the measurement binned at 0 5 m This definition may change according to the research application envisaged for instance it may be different for thin layers studies This allows Product confidence flags to be attributed on a sample by sample basis Section 3 Product confidence flags mark data that should not be used automatically For the WCO application two flags are defined e QC 1 flag no negative values in the spectra Taking into account the uncertainty of the measurement negative values less than 0 005 mare flagged e QC2 flag absorption only spectral shape raises a flag if a 510 or 532 or 555 or 650 or 676 or 715 a 412 or 440 or 488 An example of the application of the product confidence flags is shown in Figure 3 Some spectra are removed after the application of each filter 13 406 506 660 700 800 G G 0 1 0 2 0 3 0 4 6 5 Wavelength nm apg t m o o apq 1 m 0 1 0 0 M 406 506 660 806 C G 0 1 0 2 0 3 0 4 6 5 Wavelength nm apgl tm QC1 and QC2 0 5 ja 10 20 03 E 30 2 02 t y T 40 0 1 50 0 0 Y 60 4060 500 600 700 800 60 001 02 203 Of Q5 Wavelength nm apg 1 m Figure 3 Example of the application of automatic quality control procedures to total absorption spectra at L4 for 06 08 2012 Spectra a c and e are coloured accordi
18. 2 minute stop at the matching depths of water samples from CTD If following the above sampling protocol is not possible due to operational ship restrictions the scientist in charge should note it in the QAD The sampling profile can also be obtained a posteriori by plotting the time vs depth of the cast A deviation from the above routine should be checked and noted down in the QAD Only the upcast should be used for data analysis as it contains the data after the instrument has warmed up according to the manufacturer protocol and bubbles should have been eliminated by pressure However depending on the aim of the study e g correlation to physical structures downcast may be preferred Whether the down or up cast has been chosen should be detailed in the QAD 11 B DOCUMENTATION OF DEPLOYMENT PARAMETERS Deployment following Protocol Yes No Notes 3 1 3 Raw data control Ideally cruise deployment of in situ bio optical instrumentation should be examined during deployment with on screen monitoring of values This would allow for manoeuvring the ship s winch to remove bubbles or do start stop cycles of the instrument s pump However this situation seldom occurs due to operational constrains e g winch time limits or lack of suitable cable lengths Therefore alternative blind deployments are often performed during cruises The following QC guidelines address this second type of deployment i e wh
19. European Regional Development Fund Disclaimer The document reflects the author s views The INTERREG IVA 2 Seas Programme Authorities are not liable for any use that may be made of the information contained therein Contents Page 1 0 Introduction 4 2 0 Components of Pigment Analysis 4 2 1 Sample collection 4 2 2 Sample Storage 4 2 3 HPLC instrumentation and performance 4 2 4 Sample Extraction 5 2 5 Description of Standards 6 2 6 Performance Management Summary 6 2 6 1 Pigment Resolution and Retention time precision daily 6 2 6 2 Injection precision monthly 6 2 6 3 Method Uncertainty and Chl a Calibration Accuracy monthly and daily 6 2 6 4 Repipette accuracy and precision 7 2 6 5 Sample Extract Analysis Precision 7 2 6 6 Method Precision 7 2 6 7 Chl a Linearity 7 2 7 Data Processing 7 2 8 Reporting 8 2 9 HPLC Calibration 8 3 0 Summary of HPLC method and procedures 10 4 0 References 11 5 0 Protocols 12 5 1 Protocol 1 Setting up for HPLC analysis 13 5 2 Protocol 2 Pigment analysis QC sheet 14 5 3 Protocol 3 Determining the resolution of critical pairs in the mixed standard and updating retention time tracker 15 5 4 Protocol 4 Pigment extraction from filters 16 5 5 Protocol 5 Preparation of stock solution of internal standard and internal standard extraction solution 17 5 6 Protocol 6 Preparation of chlorophyll a stock solution 5 7 Protocol 7 Preparation of chlorophyll a working standar
20. NASA ESA IOC IODE SeaBASS Meaning National Administration of Space and Aeronautics European Space Agency International Oceanographic Comitee International Oceanographic Data and Information Exchange SeaWiFS Bio optical Archive and Storage System 29 Source Document J opticsdatabasev00 L4 yyyy L4_yyyymmdd Quality Assurance Document ISECA WEC 2007 2013 data Y N Parameter L4 A INSTRUMENT METHOD CHECK 1 Sensor output check Before measurement N After measurement N Previous cleaning N Previous intercalibration N 2 Calibration Date of factory calibration N Factory calibration used Y Last water calibration date 16 09 2013 Water calibration tracking Y B DOCUMENTATION OF DEPLOYMENT PARAMETERS 1 Information provided on Deployment following Protocol Y Meter wash down Y Instrument cleaned Y C RAW DATA CONTROL 1 Raw data Q C tests Plot Y Check data bounds max min Y D PROCESSING OF DATA 1 Method of processing check LO Extraction Y L1 Watercal T amp S corr scatt corr Y L2 Interpolation Y 2 Processed data tests Plot Y QC1 Automatic check for no negative values QC2 0 discards spectra QC2 1 valid X QC2 Automatic check for spectral shape raise flag if a 510 or 532 or 555 or 650 or 676 or 715 gt a 412 or 440 or 488 QC2 0 discards spectra QC2 1 valid Y E OCEANOGRAPHIC ASSESSMENT Y 1 Assessment checks Seasonal interanua
21. Section 3 3 4 are implemented here for CDOM Additional measures could include the goodness of the fit to an exponential shape D PROCESSING OF DATA 1 Method of processing check L1 Convertion used to compute Y N Absorbance into absorption DIRECTORY FILE 2 Processed data tests 25 QC1 Automatic check for no negative Y N values DIRECTORY FILE QC2 Automatic check spectral shape Y N raises a flag if a 510 or 532 or 555 or 650 or 676 or 715 gt a 412 or 440 or 488 DIRECTORY FILE 3 5 Suspended particulate matter by gravimetric methods Suspended matter quantification is based on a simple principle however accurate measurements are not trivial to obtain and QC procedures are still subject of active research 14 15 3 5 1 Instrumentation checks and calibrations The instrument should be checked following the recommendations from the National Physical Laboratory 16 Regular absolute calibrations from manufacturer and monitoring of standard balance weights should be used and the dates noted A INSTRUMENT METHOD CHECK 1 Sensor output check Before measurement Y N After measurement Y N Instrument checked Y N 2 Calibration Date of factory calibration dd mm yyyy Last laboratory calibration date dd mm yyyy 3 5 2 Documentation of deployment parameters Similarly to Section 3 3 2 water samples need to be document
22. a A 1 Amtsb A Amts A Amts 7 15 3 by assuming no absorption at 715 nm and no spectral dependence of scattering 2 Amtsb A Amts A E Cmts A amts A I 4 by assuming the error as a constant proportion of scattering Typically s 0 14 but can vary between 0 08 phytoplankton dominated and 0 3 sediment dominated 3 Amtsb A Aamts A i Cmts A Amts A ams 7 15 Cmts 7 15 amts 7 1 5 5 by using a reference wavelength 715 nm to determine the proportion of scattering and also assuming no absorption at this wavelength Although method 3 is reputed the most accurate and used as default here the data provider is let free to propose the most appropriate method for his site Primary Quality Checks Quality checks are performed after temperature salinity and scattering corrections and when depth binning the data level 2 results are written in the log file In particular the following criteria must be respected E Cmtsbl 2amtsb A 20 gt number of points within the binning layer 1 meter per default gt 1 depth centroid of data comprised within a layer lt 25 of the binning layer nominal central depth Calibration coefficients The calibration coefficients adopted are Coe0 c Coel kt IESCA satellite validation protocols 07 027 FR ISECA 18 Limitations The use of deployment speeds higher than 0 3 m sj may reduce the possibility of resolving the vertical structures in wat
23. are immediately filtered through 25 mm GF F filters nominal pore size 0 7m e The goals for filtration of particulate samples are to minimize contamination and particle degradation maximize retention and concentrate an adequate amount of particles on the filters to permit accurate spectrophotometric measurements Muller and Austin 1995 The filtration procedure should therefore be performed as follows Rinse the filtration equipment with distilled water Filter a convenient volume of seawater 500 2000ml The filtration should be carried under low vacuum pressure below 120mmHg to prevent particle breakage and pigment degradation e One pair of blank filters for each sample date should be prepared for the subsequent analysis The blank consists of filters through which 0 22 um pre filtered seawater has been passed The pre filtered seawater volume should match or be similar to the sample volume e Ensure that for both sample and blank GF F filters that the same side of the filter is used For GFF filters there is a striated and smooth side to the filter The striated shows more scattering than the smooth side and if the sample and blank side are not IESCA satellite validation protocols 07 027 FR ISECA 7 equally matched then differences in compensation between sample and blank may arise See Appendix A pp 61 63 Sample storage e Optical density spectra of the sample filters should be measured as soon after filtering as possible
24. at 440 nm by selecting baseline icon 16 Record starting conditions back pressure back pressure SD and noise amplitude at 440 nm in pigment analysis QC sheet 17 If baseline satisfactory noise amplitude 0 06 stop baseline save method save sequence and start sequence Note sequence start time on pigment analysis QC sheet 18 Any modifications to this protocol must be approved and recorded 13 5 2 Protocol 2 Pigment analysis QC sheet Pigment analysis QC sheet Extractions performed by name Notes Date of extraction HPLC lamps switched on at time Autosampler thermostat switched on at time Column thermostat switched on at time Solvents refilled New solvent prepared A B Date pyridine solution Baseline checked at time Starting conditions back pressure Back pressure SD 440 nm noise amplitude First vials in autosampler at time Sequence start time Rs and tp of mixed standard approved to proceed with extraction Mass 2mLs internal standard solution room temp 1 Av Calculated volume internal std soln mass 0 8119 Solvent added to filters at time Samples added to autosampler gt 40 mins prior to inj Sample 1 Sample 2 Sample 3 Sample 4 Sample 5 Sample 6 Sample 7 Sample 8 Sample 9 Sample 10 Sample 11
25. containing 900 mL milli Q pH meter buffers for calibration acetic acid glacial pyridine HPLC grade acetone HPLC grade methanol HPLC grade acetonitrile HPLC grade gloves pasteur pipettes and teats 2 clean 1 L HPLC eluent bottles small bottle of milli Q 2 small beakers 3x250mL measuring cylinders Eluent A methanol acetonitrile aqueous pyridine soln 50 25 25 Eluent B methanol acetonitrile acetone 20 60 20 All handling of pyridine and pyridine containing solutions to be carried out in an externally venting fume cupboard Wear gloves Use methanol acetonitrile and acetone in a fume cupboard Preparation of aqueous pyridine solution Add 10 mL of acetic acid and 20 mL of pyridine to 900 mL of milli Q water in a 1L flask Mix using a magnetic stirrer Add acetic acid dropwise until pH is 5 0 Dilute the mixture to 1000 mL with water and recheck the pH Label Aq pyridine solution 0 248M pH preparation date Name Keep in fridge Do this in an externally venting fume cupboard Wear gloves and glasses Preparation of eluent A Take a 250 mL measuring cylinder and rinse with methanol Measure 2x250 mL aliquots of methanol and transfer to a pre rinsed 1L HPLC flask Measure 250 mL acetonitrile and add to the flask Measure 250 mL of the aqueous pyridine solution and add to the flask Stopper and mix by inversion Add HPLC cap and seal with foil Label with A MeOH ACN aqueous pyridine 50 25 25
26. each measurement e Inspect the cuvettes Cuvette should be cleaned with MilliQ and lint free wipes If surface contamination still persists soak overnight in 10 HCl and clean with copious amounts of MilliQ e Allow the spectrophotometer to warm up for 30 mins Confirm that the optical windows are clean If necessary clean with MilliQ followed by ethanol HPLC grade and dry thoroughly with a lint free laboratory tissue The instrument scan speed should be 120 and slit width 4 Run an air vs air baseline Record the baseline The baseline should be spectrally flat with 0 0005 A units e Place one empty cuvette in the spectrophotometer and scan relative to air e Perform an autozero from 350 to 800nm as follows place a cuvette filled with MilliQ water in the sample cell and nothing in the reference cell Record the spectrum e Discard the MilliQ from the cuvette and rinse it three times with 5 to 10 ml of the next sample Then fill the cuvette with the sample and repeat the scan e RunaMilliQ scan between every sample to check the stability of the instrument Data Processing The MilliQ spectra is subtracted from the sample spectra No scattering offset correction should be performed The spectral absorption coefficient of dissolved organic matter is calculated from the measured absorbance as follows Ays A 2 303 Ays A 1 Where is the cuvette pathlength References Mitchell GB Bricaud A Carder K Cleveland J Ferrari G
27. from clouds or haze passing in front of the sun e Avoid measurements during strong temporal fluctuations of Ly 4 arising mainly from variable cloudiness in the sky viewing direction e Avoid outliers of L A e Avoid measurements with high tilt or roll greater than five degrees Five scans of E A La A and L A are used for further processing 9 sky Data Processing The water leaving radiance is calculated by L L 5 p sky Ly where the air sea interface reflection coefficient is estimated for sunny conditions from Figure 9 of Mobley 1999 as function of wind speed in m s W Psy 0 0256 0 00039 W 0 000034 w The reflectance o A is then calculated for each scan and the mean and standard deviation over the five scans are calculated and plotted Postprocessing Quality Checks Reflectance spectra are inspected subjectively to ensure e limited variability over scans comparing standard deviation with mean e internal consistency of spectra in red and near infrared positive reflectances with reflectance ratios given approximately by the inverse ratio of pure water absorption Measurements outside the range 400 900nm are not used for scientific analysis because of high uncertainty and instrument noise Limitations e Measurement uncertainties associated with the air sea interface reflection correction become significant in conditions of cloudy sun and to a lesser extent cloudy sky in the
28. instrument is an above water radiometer designed and manufactured by the Laboratoire d Optique Atmosph rique of the University of Lille France It measures water leaving radiance and aerosol optical thickness in 11 spectral bands each bandwidth of 10nm centered at 350 380 410 443 490 510 565 620 670 750 870nm by viewing the sun and the ocean surface sequentially The same optics with a field of view of about 3 the same interference filters and the same detectors are used in both ocean viewing and sun viewing mode A different electronic gain high and low is used for each mode respectively The optics IESCA satellite validation protocols 07 027 FR ISECA 43 are fitted with a vertical polarizer to reduce reflected skylight when the instrument is operated in ocean viewing mode Pressure temperature and viewing angles are also acquired automatically An integrated GPS antenna acquires automaticaly the geographic location at the time of measurement and a display indicates various information Instrument Calibration and Quality Assurance The instruments are calibrated at the Universit de Lille detailed calibration histories for each instrument can be found on the SIMBADA web site Methodology and Processing Description Deployment of the instrument The instrument is operated from the deck of a ship using the measurement sequence Dark with lens cap on 3 Sun 3 Sea 3 Sun Dark The ship is manoeuvred on station
29. is given on the principles the minimum requirements division of tasks and summary of the quality control checks in a Quality Assessment Document QAD The second part of the report describes application of the quality control QC methodology applied to the ISECA bio optical data set as an example for construction of similar datasets 2 VALIDATION OF METOCEAN DATA The four major aspects of metocean data validation are a Instrumentation checks and calibrations which include calibration checks of sensor response tests on instrument or system electronics and checks on data processing and recording equipment b The documentation of deployment parameters which includes definition of the location and duration of the measurements method of deployment of the instrumentation and sampling scheme used for the measurements c Automatic quality control of data which comprises a series of tests on the data to identify erroneous and anomalous values in order to establish whether the data have been corrupted in any way either during initial measurement or in copying or transmission to a user d Oceanographic assessment which includes an assessment of the results of conditions a to c and an assessment of the oceanographic and meteorological reasonableness of the data comprising checks on expected patterns or trends and comparisons with other data sources The four aspects of data quality control i e instrument calibration deploymen
30. observation of a sharp increase after day 1080 from the manufacturer calibration Data collected during the period of sharp increase were flagged as suspicious Hence it is very important to be able to check in real time the drift of the instrument to detect any malfunction The date of the last checked value of water calibration should be recorded in the QAD ABSORPTION OFFSET Values 0 10 E 0 05 o p o L S be gt oof I L o li uL u L o L of 0 05 0 10 L Lus a l ho Moon 0 400 600 800 1000 1200 Days from WL calibration d 200 Figure 1 Example of water calibration tracking of a Wetlabs ac9 instrument 10 A INSTRUMENT METHOD CHECK 1 Sensor output check Before measurement Y N After measurement Y N Instrument wash down Y N Instrument cleaned Y N 2 Calibration Date of factory calibration Y N Factory calibration used Y N Air tracking Y N Last water calibration date dd mm yyyy Water calibration trend dd mm yyyy Directory file 3 1 2 Documentation of deployment parameters Deployment should be as follow e Switch on the instruments and deploy in water e Lower the instrument to 10 m and leave for 5 to eliminate bubbles e Raise the instrument to 1 m below the surface and lower to maximum depth at 15 m min e Raise the instrument from the maximum depth at a speed of 7 m min or lesser If feasible allow for 1 or
31. of Chla in these waters For Case 2 waters where high suspended particulate material TSM and coloured dissolved organic material CDOM causes a de coupling of phytoplankton absorption and the underwater light field accurate retrieval of Chla is far more complex and as yet remains unresolved Optical and bio optical protocols have been well documented for Case 1 waters for validating SeaWiFS data Mueller amp Austin 1992 Fargion amp Mueller 2000 but require modification for the more complex Case 2 waters and for validating data from more recent sensors such as MODIS and MERIS Defining the contribution of CDOM living and non living matter to the optical properties of the upper water column and the development of reliable and robust methodologies for Case 2 waters is fundamental for remote sensing research The following protocols document draws on the experience of NASA s SeaWiFS project and the EU Colors project Coastal region long term measurements for colour remote sensing development and validation MAS3 CT97 0087 funded by the EU Marine Science and Technology Programme MAST III Startegic Marine Research and the EU FP5 project REVAMP Regional validation of MERIS chlorophyll products in North Sea coastal waters EVGI CT 2001 00049 Tilstone et al 2003 The protocols should be used in parallel with Protocols for the Validation of MERIS water products Doerffer 2002 which documents MERIS water products validation strategies and s
32. on a frame lowered into the sea by means of a winch A small pump brings the water through the ac9 flow tubes flow rate through the tubes should be kept above 1 liter minute All tubing is black or covered with black tape at least the 20 cm at the flow inlet and outlet to avoid direct light into the tubes The lowering speed for a frequency of acquisition of 6Hz should be about 0 1 0 2 m s Air bubbles passing through or even remaining trapped into the flow tubes when the instrument is at surface and or in the first meters according to sea state can affect measurements and induce differences between down and up cast values profiles Assuming that putting the instrument at depth at least 10 meters may help purging the system for bubbles only the up cast profiles are considered here Simultaneous profiles of in situ temperature and salinity are collected for post correcting the data see 4 Data Post Processing Field pure water calibration The instrument must regularly once per day of measurement if possible be calibrated in the field with pure water milli Q water is recommended in its deployment configuration in order to remove the effects of small misalignments of the optical system and or to track possible long term drift See also the air calibration procedure in WetLabs protocol The calibration is performed by making milli Q water pass through IESCA satellite validation protocols 07 027 FR ISECA 16 the flow tubes
33. out other routine tasks as a pilot test for future deployment Unsupervised sampling of above water radiometric quantities was done using a hyperspectral Satlantic HyperSAS system composed of three sensors measuring simultaneously downwelling irradiance Eg sky radiance Li and water leaving radiance L This system also included a Satlantic tilt heading and roll sensor THR and GPS The three sensors were mounted on a pole on the bow of the vessel at 5 m off the water surface Lt was measured pointing to the water surface with an angle of 40 from the nadir and the crew was instructed to measure away from the sun azimuth at 135 when possible during other routine operations 9 A new algorithm to filter and process R from radiance spectra was tested 10 Results have been presented in a paper 11 3 DATASETS IMPORTED IN THE WAS Data were collected by ISECA partners through their interactions with relevant national agencies which hold data repositories Tables 1 and 2 provide the time span and describes the sources of the different data variables Table 1 Data sources and reference web sites per project partner Time Link to data source Main person of contact Span PML UK 1988 http www westernchannelobservatory org Victor Martinez Vicente 2012 uk vmv pmil ac uk IFREMER 1998 http wwz ifremer fr lerpc Activites et Francis Gohin FR 2013 Missions Surveillance REPHY Francis Gohin ifremer fr NI
34. seawater should be filtered through pre washed pre ashed pre weighed 0 7 um filters The volume of seawater filtered is dependent on the amount of material present in the water and should be sufficient to detect weights to 5 significant figures e Water samples should be filtered immediately on collection If this is not possible it is recommended that 1 ml of 4 formalin per litre of sea water is added to the water sample Multiple replicates should be taken to quantify sample variability A blank filter should be used for each sample to calculate the handling error of the sample e After filtration leave the filter on the glass frit and the filtration apparatus standing Filter at least 50 mls of distilled water through the filtration apparatus to remove any salt Repeat this procedure three times With the vacuum pressure still on carefully remove the filtration cup and using a wash bottle gently wash the outer edge unfiltered area of the filter The filters should then be dried in an IESCA satellite validation protocols 07 027 FR ISECA 34 oven at 75 C for 24 hrs after which they are stored in a dessicator before weighing See Van der Linde 1998 TSM concentration is deduced from the difference between original filter weight minus sample filter weight divided by filtration volume Limitations Non accurate washing of filters could induce very large errors in the derived TSM values References J D H Strickland and T R Parso
35. specifications characterization and calibration overview Petzold T J 1972 Volume scattering functions for selected natural waters Scripps Institution of Oceanography Visibility Laboratory San Diego CA SIO Ref 71 78 HydroScat 6 Spectral backscattering sensor USER S MANUAL 2010 Rev J edited p 42 HOBILabs Inc Berthon J F et al 2007 Measurements and modeling of the volume scattering function in the coastal northern Adriatic Sea Applied Optics 46 5189 5203 IESCA satellite validation protocols 07 027 FR ISECA 15 In situ spectral Beam Attenuation coefficient c z A m In situ spectral Absorption Coefficient a z A m Both at wavelengths412 440 488 510 555 630 650 676 715 nm Instrument Description AC9 Dual Path Absorption and Attenuation Meter WET Labs Inc USA The ac 9 concurrently determines the spectral beam attenuation and spectral absorption of water over nine wavelengths Optical Specifications Bandpass 10 nm channel Pathlength 25 cm Beam cross section diameter 8 mm Receiver Acceptance Angle 0 7 deg in water Fig 4 AC 9 unit WETlabsInc USA Instrument Calibration and Quality Assurance The protocol proposed by Wet Labs ac9 Protocol document Revision B is followed The salient points regarding deployment and calibration are highlighted below Mounting and Deployment of the instrument The instrument is deployed preferably upright
36. sun pointing References Fougnie B R Frouin P Lecomte and P Y Deschamps 1999 Reduction of skylight reflection effects in the above water measurement of diffuse marine reflectance Applied Optics 38 18 3844 3856 IESCA satellite validation protocols 07 027 FR ISECA 44 Mueller J L C Davis R Arnone R Frouin K Carder Z P Lee R G Steward S Hooker C D Mobley and S McLean 2000 Above water radiance and remote sensing reflectance measurements and analysis protocols Ocean Optics protocols for satellite ocean color sensor validation Revision 2 Greenbelt Maryland National Aeronautical and Space Administration 98 107 Ruddick K V De Cauwer Y Park G Becu J P De Blauwe E D Vreker P Y Deschamps M Knockaert B Nechad A Pollentier P Roose D Saudemont and D v Tuyckom 2002 Preliminary validation of MERIS water products for Belgian coastal waters Envisat Validation workshop 9 13th December 2002 Frascati European Space Agency IESCA satellite validation protocols 07 027 FR ISECA 45 INTERREG IVA 2 Mers Seas Zeeen Cross border Cooperation Programme 2007 2013 Deliverable 2 1 Guidelines for Quality Control of bio optical measurements in Case 2 European Waters Contract 07 02 7 FR ISECA V Martinez Vicente G H Tilstone and R Airs 2013 Plymouth Marine Laboratory PML UK seca INFORMATION SYSTEM ON THE EUTROPHICATION u e OF OUR COASTAL ARE
37. to point to a ship heading of 135 with respect to sun Sun measurements are made from anywhere offering a clear view of the sun Sea measurements are made from the prow of the ship pointing forwards at relative azimuth angle of 135 with respect to sun and zenith angle of approximately 40 The complete measurement sequence lasts approximately 5 minutes During measurements wind speed atmospheric pressure cloud cover and type and sea and sky state conditions are noted Any variability in illumination e g clouds passing near sun requires a restart of the measurement sequence Measurements can also be made underway for a ship heading of 135 relative to sun During such measurements visual checks are made of the sea surface for variability such as fronts or floating material and the ship heading is monitored Description of processing techniques and quality checks Data is processed at the Universit de Lille Reflectances are given for 10 wavelengths excluding the 870nm band The processing method is outlined on the SIMBADA web site The use of a polarizer to reduce air sea interface reflection is discussed in Fougnie et al 1999 Limitations e Measurements can only be in clear sun and clear sky cloud cover lt 2 8 conditions to ensure accurate calculation of E 4 Cloud cover is estimated subjectively e Measurements from small ships e g Rigid Inflatable Boats are limited to calm sea state e g Bf lt 3 to ensure accurate
38. using 2 Open method C UVWINLAB METHSOO ChI_std2 msc 3 Go to My computer D drive pigments Create folder with todays date eg 20120419 4 Run air blank ie press autozero to zero spectrophotometer on air 5 Run air scan ie press start to run scan on air Resulting spectrum baseline should be 0 0 005 If not inform G Tilstone 6 Use matched cuvettes from lab G13 pigment drawer Ensure cuvettes clean Fill both cuvettes with 9096 acetone Insert into sample and reference cuvette holders Go to instrument tab in software Select baseline filename 90aceton and press AUTOZERO At Do you want to perform background correction prompt select Yes 7 Run solvent scan Leave 9096 acetone in both cuvettes and press start 8 When scan complete In graph window save file File save as d pigments 20120419 90aceton 9 Goto sample tab in software Remove sample cuvette Rinse and fill Make sure chl ws is at with chl a WS Wipe surfaces and replace in cuvette holder room temp Take portion from freezer in advance and keep in dark 10 Press START 11 When scan complete In graph window save file File save as d pigments 20120419 chla_ws 12 In graph window open file Can open new graph window and remove current spectra Use vertical cursor icon to get cursor on screen and record absorbance at 663 750 and 700 nm 13 Input data to results sheet chl a standard 14 Calculate chl concentration using following formula d
39. which are identified by a combination of retention time and UV vis spectra The pigment analysis method is relatively complex The whole process includes multiple stages comprising filtration sample storage extraction clarification analysis by HPLC and data processing All component procedures can contribute to errors and quality assurance procedures are essential to produce reliable results Each component of the process will be detailed below with reference to documented protocols 2 0 Components of Pigment Analysis 2 1 Sample Collection Phytoplankton samples for pigment analysis are obtained by filtering seawater through GF F filters For discussion of filter types see Chapter 10 of Jeffrey et al 1997 and Appendix A in Roy et al 2011 Volumes of between 1 and 4L are typically filtered depending on the water type eg 1L is sufficient at L4 in the spring summer 2 L at L4 in the winter 3 4 L in the most oligotrophic waters during AMT Seawater is filtered through GF Fs via gentle vacuum Filtration should occur as soon as possible after sampling If samples cannot be filtered immediately they should be stored in a cool dark environment prior to filtering During filtration filters should be removed from the filter rig as soon as the last of the water passes through the filter Filters should not be left on the rig longer than this due to risk of oxidation Once removed from the rig with forceps filters should be folded in half inserted
40. 1 Chlide a 0 05 0 16 8 23E 06 N Calc from chl a 1 98 0 002 MgDVP 0 02 0 06 3 07E 06 N Chl c2 0 74 0 001 Chl c2 0 02 0 06 3 07E 06 Y 0 74 0 001 Perid 0 04 0 13 6 81E 06 Y 1 64 0 002 But fuco 0 03 0 10 4 97E 06 Y 1 19 0 001 Fuco 0 03 0 10 4 98E 06 Y 1 20 0 001 Neo 0 02 0 07 3 45E 06 Y 0 83 0 001 Pras 0 03 0 09 4 89E 06 Y 1 18 0 001 4 ketohex 0 03 0 09 4 78E 06 N Hex fuco 1 15 0 001 Viola 0 02 0 06 3 25E 06 Y 0 78 0 001 Hex fuco 0 03 0 09 4 78E 06 Y 1 15 0 001 Asta 0 02 0 07 3 59E 06 N B caro 0 86 0 001 Diad 0 02 0 06 3 33E 06 Y 0 80 0 001 Allo 0 02 0 07 3 64E 06 Y 0 87 0 001 Dlato 0 02 0 07 3 55E 06 Y 0 85 0 001 Zea 0 02 0 07 3 72E 06 Y 0 89 0 001 Lut 0 02 0 06 3 36E 06 Y 0 81 0 001 Canth 0 02 0 07 3 59E 06 N B caro 0 86 0 001 Gyro 0 02 0 06 3 14E 06 Y 0 75 0 001 Croco 0 02 0 06 3 33E 06 N Diad 0 80 0 001 Chl b 0 07 0 23 1 18E 05 Y 2 84 0 003 Chl b 0 07 0 23 1 18E 05 N Chl b 2 84 0 003 Chl c2 0 04 0 13 6 62E 06 N Calc from chl c2 1 59 0 002 MGDG DVChl a 0 04 0 13 6 77E 06 Y 1 63 0 002 Chl a 0 07 0 23 1 196E 05 Y 2 87 0 003 Chl a 0 07 0 23 1 196E 05 N Chl a 2 87 0 003 Phea 0 07 0 23 1 20E 05 N 2 87 0 003 A caro 0 02 0 07 3 59E 06 N B caro 0 86 0 001 B caro 0 02 0 07 3 59E 06 Y 0 86 0 001 Table 2 LOD s LOQ s reponse factors and effective LOD s for pigments quantified by HPLC at PML from 2012 calibration 3 0 Summary of HPLC method and procedures Waters C8 Symm
41. 1 Perkin Elmer Lambda 800 spectrophotometer IESCA satellite validation protocols 07 027 FR ISECA 6 Recommended baseline noise from 350 to 800 nm for GF F s is 0 005 A and for 10 cm quartz cuvettes with purified water is 0 0005 A Analytical procedure e Warm up the spectrophotometer for at least 30 minutes Check the specific instrument warm up guidelines to meet photometric and baseline accuracy e If samples and blank are frozen place in petri dish on filtered water to ensure hydration and allow to thaw for at least 5 minutes Store in a refrigerator until analysis e Both sample and blank filters will dry out over time and must be re hydrated regularly after every measurement If the absorbance signal deviates greater than 0 02 absorbance from zero between 750 800 nm this indicates a drying of the sample Mitchell et al 2000 Instrument Calibration and Quality Assurance Spectra should be visually and or automatically checked in particular for e The presence of a significant peak around 665 nm in agp A spectra which indicates non complete bleaching of the sample e abnormal lt 1 ratio of app 443 apn 665 Methodology Sample collection and filtration e Filtration volume should be adjusted to keep the samples in the optical density range that is ideal for the path length amplification corrections see below e After collection water samples are transferred to black polyethylene bottles e The samples
42. 1 28 5 07 3 62 6 50 3 41 6 97 9 44 9 88 10 32 12 70 13 38 16 51 17 40 18 50 19 17 19 94 20 30 21 13 22 48 23 58 25 10 26 20 27 00 27 22 28 78 29 30 32 01 32 33 33 45 33 74 34 11 35 89 36 65 36 85 10 43 5 08 4 52 6 74 3 11 Stddev RSD 0 04 5 91 0 04 4 31 0 04 5 37 0 02 1 80 stddev Mean r 0 2937 7 24 0 3917 9 84 04054 10 27 04200 1074 051001 1328 0 5254 13 95 05479 1749 0 5441 1807 0 5654 1904 05394 1981 0 5351 20 60 05551 2108 05700 2180 05330 2345 0 5548 24 49 04147 2567 03053 2659 02519 2729 02352 2750 0 3254 2901 0 0907 29 47 01366 3214 0 1439 3247 016923 338 0 1698 3392 0 1243 3427 0 2444 3620 027255 3695 02840 3745 01372 1077 0 0087 5 09 0 1147 4 22 0 1285 6 64 0 0911 3 20 change from mean 2 90984 4 438873 0 63473 3 18844 CV or RSD change 4 06 2 30 3 98 2 62 3 95 2 35 3 91 2 29 3 84 2 08 3 77 2 00 3 19 1 56 3 01 1 50 2 97 1 33 2 72 1 38 2 60 1 27 2 64 1 24 2 62 1 23 2 30 1 12 2 27 0 93 1 62 0 79 1 15 0 60 0 92 0 52 0 86 0 48 1 12 0 62 0 31 0 09 0 43 0 45 0 44 0 46 0 48 0 53 0 50 0 55 0 36 0 48 0 68 0 82 0 74 0 71 0 76 0 84 1 27 0 46 0 17 0 11 2 72 0 92 1 93 0 89 2 85 2 96 6 3 Spreadsheet 3 Chl a standard RESULTS SHEET CHL A STANDARD Date chl stock prepared Date w s prepared Date w s measured spec Spectrophotometer used Baseline
43. 2012 chla ws 10 12 2012 Int std 11 01 2013 chla ws 17 01 2013 Int std 18 01 2013 Int std 31 01 2013 chla ws 25 04 2013 chl ws Peak areas Inj 1 510121 584953 589591 584424 582255 565911 554941 570073 112473 390041 106707 83888 372804 425074 Inj 2 503148 584534 579497 580843 580104 572966 570894 572332 108257 385601 107965 83566 378828 424796 Inj 3 493914 565722 573025 577626 565198 574831 554388 554015 111290 372015 110548 85096 370320 423581 6 5 Spreadsheet 5 Method uncertainty Chl a ws concentration Spectrophotometry ng injected Measurement 1 HPLC Measurement 2 HPLC Measurement 3 HPLC Measurement 4 HPLC Measurement 5 HPLC Measurement 6 HPLC Average SD Precision SD Avg 100 Accuracy 96 5 17 5 28 5 26 5 13 5 39 5 36 5 27 528 0 09 175 2 15 Inj 4 481615 573498 569067 553397 570784 556834 553591 111314 373867 412567 5 34 5 32 5 24 5 35 5 30 0 06 1 09 0 71 Inj 5 501796 572520 567534 575911 567674 380363 435332 5 54 5 70 5 50 5 47 5 56 0 12 2 25 0 33 10 12 2012 11 10 2012 09 10 2012 08 10 2012 5 54 5 90 5 70 5 77 5 79 0 10 1 71 4 49 25 r r Inj 6 491318 567586 570661 572536 567759 364361 427710 02 10 2012 5 54 5 60 5 73 5 66 0 09 1 63 2 21 std dev 10114 8236 8327 10944 9289 3848 7468 10089 1804 9453 1958 807 4375 7356 Mean CV o
44. 39 91 Canthaxanthin PPS CHAN 25mi 1 175 157 74 Capsanthin PPS CAPSA 25mi 1 042 139 91 Chlorophyll a PPS CHLA 25mi 1 042 139 91 Chlorophyll b PPS CHLB 25mi 1 042 139 91 Chlorophyll c2 PPS CHLC2 25mi 1 175 157 74 Chlorophyll c3 PPS CHLC3 25mi 1 175 157 74 Chlorophyllide a PPS CHLIA 25mi 2911 390 69 Crocoxanthin PPS CROC 25m 2 911 390 69 Diadinoxanthin PPS DIAD 25mi 1 175 157 74 Diatoxanthin PPS DIAT 25m 2 911 390 69 Dinoxanthin PPS DINO 25mi 2 911 390 69 Divinyl Chlorophyll a PPS DVCHLA 25mi 1 175 15774 Divinyl Protochiorophyllide PPS MgDVP 25mi 1 175 157 74 Echinenone PPS ECHI 25mi 1 175 157 74 Fucoxanthin PPS FUCO 25mi 1 042 139 01 Gyroxanthin diester PPS GYRO 25mi 2 911 390 69 Lutein PPS LUTE 25mi 1 042 139 91 Lycopene PPSLYCO 25 mi 1 042 139 91 Mixed Pigments PPS MIX 1 imi E 58 60 Mixed Pigments PPS MIX 2 1m c 58 60 Mutatoxanthin PPS MUTA 25mi 1 042 139 91 Myxoxanthophyll PPS MYXO 25mi 1 175 157 74 Neoxanthin PPS NEOX 25mi 1 175 157 74 Peridinin PPS PERI 25mi 1 175 157 74 Pheophorbide a PPS PHBA 25mi 1 042 139 91 Pheophythin a PPS PHAE 25mi 1 175 15774 Prasinoxanthin PPS PRAS 25mi 1 042 139 91 Violaxanthin PPS VIOL 25mi 1 175 157 74 Zeaxanthin PPS ZEAX 25mi 1 175 157 74 EURO prices may vary with exchange rates S units minimum sale 31
45. 440878 11 01 2013 390041 377708 4 49 1 19E 05 0 61 New working standard prepared Harris Airs 385601 372015 373867 380363 364361 17 01 2013 399762 391768 4 49 1 15E 05 4 17 393461 382082 18 01 2013 375587 373760 4 49 1 20E 05 0 44 374149 371545 31 01 2013 372804 373984 4 49 1 20E 05 0 38 378828 370320 01 02 2013 387055 384967 4 49 1 17E 05 2 48 385671 382176 25 04 2013 425074 424843 5 28 1 24E 05 3 91 New working standard prepared Cummings Harris Airs 424796 423581 412567 435332 427710 26 04 2013 420288 410967 5 28 1 28E 05 7 42 Above 5 prepare new working standard 409414 403200 06 06 2013 438414 428433 5 25 1 23E 05 2 46 New working standard prepared 418390 417442 429212 430918 436219 10 06 2013 439655 431670 5 25 1 22E 05 1 69 433200 422156 26 7 0 Appendices Appendix 7 1 Common pigments consumables and suppliers Item Supplier P N Cost Ea 2012 HPLC consumables Symmetry C8 HPLC column Waters WAT106011 328 00 Symmetry C8 guard column Waters WAT106128 110 00 Guard holder kit Waters WAT097958 102 30 Vials amber 1000 pk Kinesis STV12 O2LA 89 25 Vials clear 1000 pk Kinesis STV12 02L 72 75 Caps 1000 pk Kinesis SCCO9 04B 138 00 Lamps Thermo tbc Standards Mixed sample DHI 16 19 Full set calibration stds DHI 2 802 54 Dry ice delivery costs DHI 166 62 Calibration stds 6 month check DHI 62
46. 596 of the calibration value the system is recalibrated Every time pigment analysis is performed three injections of the chlorophyll a working standard are performed The response factor is determined and the change from the calibration value is calculated This must be within 5 or a new standard is prepared see spreadsheet 6 2 6 4 Repipette accuracy and precision The accuracy of the pipette used for pigment extractions is determined before use by performing three weighings of the repipette volume Three replicate weights of extraction solution are taken by pipetting the internal standard extraction solution at room temperature The average weight is used to calculate the pipette volume using the specific gravity of 9096 acetone The data are recorded on the pigment analysis QC chart see protocol 2 2 6 5 Sample Extract Analysis Precision The first sample extracted is split between two vials which become the first and last analysed of that daily batch of samples and therefore represents the minimum and maximum time the vials reside in the autosampler The precision for TChla and PPig major pigments only can be calculated The measurements indicate analytical precision even if for example sampling of duplicates is unreliable It is also an aid for QC decisions eg whether reanalysis of samples is required if autosampler cooling fails 2 6 6 Method Precision Method precision is determined by analysis of duplicate filters at the freq
47. 6 E Green 6 E 06 i i A aRed 4 E 06 A a A ry 2 E 06 0 E 00 0 200 400 600 800 1000 Days since manufacturer calibration Figure 4 Change of calibration scaling factors S as a function of time for the three channels for a Wetlabs backscatter meter ECO BB3 monitored at PML A INSTRUMENT METHOD CHECK 1 Sensor output check Before measurement Y N After measurement Y N Instrument wash down Y N Instrument cleaned Y N 2 Calibration Date of factory calibration Y N 16 Factory calibration used Y N Last laboratory calibration date dd mm yyyy Dark count calibration trend dd mm yyyy Directory file S calibration trend dd mm yyyy Directory file 3 2 2 Documentation of deployment parameters Because the backscattermeter is often deployed with the optical package containing the absorption and attenuation meter same procedures apply Section 3 1 2 Only the upcast should be used for data analysis as it contains the data after the instrument has warmed up according to the manufacturer protocol and bubbles should have been eliminated by pressure B DOCUMENTATION OF DEPLOYMENT PARAMETERS Deployment following Protocol Yes No Notes 3 2 3 Raw data control The data logging of the Wetlabs backscatter meter is usually done with the other Wetlabs sensors so the same procedure applies Section 3 1 3 A raw data control that
48. 6 30 Chl a Sigma C6144 1MG 133 50 Internal standard trans B Apo 8 carotenal Sigma 10810G 1G 59 00 Solvents Methanol 2 5L Fisher M 4056 17 20 57 Acetone 2 5 L Fisher A 0606 17 39 15 Acetonitrile 4x2 5L Fisher A 0626 pk4 374 74 Pyridine 500 mL Fisher P 7960 08 78 89 Acetic acid 500 mL Fisher A 0406 PBO8 32 10 Filters tubes etc Cryovials 1000 pk Fisher FB74405 163 81 Extraction tubes 500 pk 25 00 Microcentrifuge tubes 500 pk Fisher FB55921 21 45 Filters GF F 100 pk Fisher 11754083 18 53 Syringe filters 0 2uM Teflon 17mm 100 pk DHI 69 00 Syringes 1mL 100 box Medisave SYR113 6 62 Pipettes long 4x250 Fisher 11566963 35 36 28 Appendix 7 2 Copy of Thermo Maintenance contract 2013 Thermo Fisher Scientific Stafford House Boundary Way SUPPORT PLAN QUOTATION m c QUOTATION Number 202856707 40090654 Te 01449 233555 Coverage Start Date Dril 01 2013 Fax 01442 233667 Coverage End Date March 31 2014 Quotation Expires October 18 2012 Quote Created On 20 2012 Dr Ruth Airs Plymouth Marine Laboratory Prospect Place Poole PL13DH Phone Dear Dr Airs Plan on your Thermo Scientific instrument is near to expiring Please find attached a quotation to provide coverage and help you continue to gain the greatest value Md iere When purchased your instrument a partner that is committed to your term success We understand tha every Fetument issue i important and our re
49. AS Investing in your future Crossborder cooperation programme 2007 2013 Part financed by the European Union European Regional Development Fund Disclaimer The document reflects the author s views The INTERREG IVA 2 Seas Programme Authorities are not liable for any use that may be made of the information contained therein Contents 1 INTRODUCTION E 5 2 VALIDATION OF METOCEAN DATA ssssssseseseeeeeneneeen enne ener nnne enne nennen nnn 5 2 4 Data quality control measures for data providers eesesesss 6 2 2 Data quality control measures for data managers sse 7 2 3 Quality assurance documents sse ennt enne nennen nenne 7 3 DATA QUALITY PROCEDURES FOR BIO OPTICAL MEASUREMENTS 8 3 1 In water absorption and attenuation essent 9 3 11 Instrumentation checks and calibrations suusss 9 3 1 2 Documentation of deployment parameters sss 11 33 Raw data COllPOl es c ori rd D PED M ERN ENIM PD NM UM 12 3 1 4 Processing and Automatic Quality control of data 13 3 2 In water backscattering sssssssessssseseeeseeeeee enne tentent enses nni 15 3 21 Instrumentation checks and calibrations uussss 15 3 2 2 Documentation of deployment parameters
50. ESCA satellite validation protocols 07 027 FR ISECA 3 DOT nA ENET hearer E E EE E Ease ee eee va tovee torres eE resesevsee eaque yo tee eve dege 33 didi m E 33 Total Suspended Matter TSM g m sss 34 Instr ment desSCEIDHOT dao disi iot rt ta Rame eroe ii dide EAE operare gU A i see Pd dpud an ad 34 JB iter ERE NM 34 Instrument calibration and quality assurance esee 34 M th dolo BY ete cM C 34 Filter preparation d esee eei v rite bdbria Erates eo a ab Diana a ue IU EHE as dida REM ER d 34 laicis ohe nia bes Duet E E A E E EEA E AO EE 34 TSM c ncentrati m o e aA NEE E EA baut E E EG 35 Limitation E 35 IARE RM 35 Above water Water Leaving Radiance Lw Wm nm sr and Downwelling Irradiance Es W mnm NEN KE ERN NENNT RENE 36 Instt ment DesSctiptlolic scissor dc ahead cae aiite uen ede u asia sea edes 36 QA and data processing detail ss uso iene cesis Eoi tuse y iu Senec e eut d dae 36 Instrument Calibration and Quality Assurance eeseeeeeenee n 97 Methodology and Processing Description eese 37 EREMO RETI TUN E 37 REfETENCE So catus aa aa a a a a aa a e aa 37 Above water MERIS reflectance pw 4 dimensionless 39 TAOS meth id ooo ode die oap itv er pepe A daba eds 39 Instrument desSetlpli Oll eoe ch re etat ip a tei RU seat a pa dimus ie Cin vH dee Madrina 39 Instrument Calibration and Quality Assurance seeeeeseseeeseese
51. INTERREG IVA 2 Mers Seas Zeeen Cross border Cooperation Programme 2007 2013 ISECA Deliverable 2 1 A description of the in situ measurements the attached protocols and the quality control Contract 07 027 FR ISECA V Martinez Vicente G H Tilstone and S Groom Plymouth Marine Laboratory PML UK 2 Mers Seas Zee n J FRANCE ENGLAND VL REN NEDERLAND Iseca INFORMATION SYSTEM ON THE EUTROPHICATION OF OUR COASTAL AREAS Investing in your future Crossborder cooperation programme 2007 2013 Part financed by the European Union European Regional Development Fund Disclaimer The document reflects the author s views The INTERREG IVA 2 Seas Programme Authorities are not liable for any use that may be made of the information contained therein INTERREG IVA 2 Mers Seas Zeeen Cross border Cooperation Programme 2007 2013 ISECA Deliverable 2 1 Description of the in situ measurements Contract 07 02 7 FR ISECA V Martinez Vicente and G H Tilstone Plymouth Marine Laboratory PML UK 2 Mers Seas Zee n FRANCE ENGLAND EN 4 DERLAND Iseca INFORMATION SYSTEM ON THE EUTROPHICATION OF OUR COASTAL AREAS Investing in your future Crossborder cooperation programme 2007 2013 Part financed by the European Union European Regional Development Fund Disclaimer The document reflects the author s views The INTERREG IVA 2 Seas Programme Authorities are not liab
52. ISECA region Phaeocystis globosa and the sum of all Phaeocystis were selected In addition to the basic set of variables an extended range of variables was monitored by PML following recommendations from users D 1 2 User Requirements for the Remote sensing of Eutrophication in the 2Seas coastal waters The first part of this report summarises the overall sampling techniques and basic data processing steps for the new data collected during ISECA by PML Detailed methods description and quality control are described in other parts of this Deliverable Protocols and Quality Control Guidelines The second part of this report summarises the cross border data that were used in the WAP 2 BIOLOGICAL AND BIO OPTICAL SAMPLING DURING ISECA In situ sampling activities at L4 and E1 have two different strategies One is long term monitoring of the vertical biological and optical properties of the water column Another is the ad hoc above water reflectance sampling attempting to obtain matching data with satellite overpasses The time series approach is intended as a part of a long term effort to monitor trends in ecosystem behaviour and to test in water algorithms The opportunistic above water sampling is focused on obtaining a dataset useful to test adjacency effect algorithms and atmospheric correction schemes A summary of the available datasets the sampling methodology and processing is summarised in this report and is based on precedent published st
53. OZ NL 1990 http live waterbase nl waterbase wns cfm Jacco Krokamp 2011 wbwns1 en Jacco Kromkamp nioz nl VITO BE 2003 http www vliz be vmdcdata midas Francisco Hernandez 2013 francisco hernandez VLIZ fr Overall 25 stations were selected for inclusion in the WAS covering a period of over 20 years Spatial coverage of the selected stations is shown in Figure 2 51 N 49 N 4 W 2 W 0 2 E 4 E 6 E Figure 2 Location of the stations selected for input into the WAS Extended documentation on the methodology used for each variable by each data provider can be found in the relevant internet link or by contacting the person provided in Table 1 Table 2 Data parameters and availability per partner shown by shadowed cells IFREMER NIOZ Temperature Salinity Chl a SPM Turbidity Nutrients Phaeocystis spp Phaeocystis all Data from individual data providers were formatted into a common data format at PML using IDL code An example of the common data format agreed with VITO for ingestion into the WAS is provided here DI WEC bc seabass hit No File Edt Format View Help 38 44 2014 jarrian cente claire widdiconbe Malcolm woodward Ruth Airs Peter Land Denise cummings Tysouth Mar inet abo c stationeL4 data file name WEC bgc seabass txt documents nof ile txt calibration files nofile txt data type under way dat
54. Spreadsheet 2 RT tracker Pkno Name 1 Chl c3 2 Chlorophyllide a 3 MgDVP 4 Chl c2 5 Peridinin 6 Peri isomer 7 19 Butfucoxanthin 8 Fucoxanthin 9 Neoxanthin 10 Prasinoxanthin 11 Violaxanthin 12 19 Hexfucoxanthin 13 Astaxanthin 14 Diadinoxanthin 15 Antheraxanthin 16 Alloxanthin 17 Diatoxanthin 18 Zeaxanthin 19 Lutein 20 Canthaxanthin 21 Int std 22 Chl b 23 Chl b 24 Di vinylchl a 25 Chl a 26 Chl a 27 pheophytin a 28 alpha carotene 29 Betacarotene RRT c3 fuco RRT fuco diadino RRT diad zeax RRT zeax chl a RRT chl a Bcar 10 01 0 60 10 47 0 57 0 78 20 22 0 44 20 64 0 53 0 86 27 09 0 30 27 30 0 30 0 71 33 46 0 30 33 76 0 39 0 87 NMP 10 02 0 60 10 47 0 64 0 73 20 28 0 47 20 70 0 55 0 83 27 12 0 30 27 33 0 30 0 71 33 50 0 30 33 80 0 38 0 88 10 16 0 65 10 62 0 74 0 66 20 47 0 46 20 90 0 54 0 86 PUPAL 0 30 27 42 0 31 0 69 33 58 0 31 33 88 0 38 0 88 NPC 10 03 0 61 10 49 0 64 0 73 20 34 0 42 20 77 0 48 0 94 27 15 0 27 27 36 0 29 0 76 33 44 0 28 33 73 0 37 0 91 NMP NPC new precolumn NMP new mobile phase SMP swapped mobile phase 11 06 0 63 11 55 0 80 0 68 21 61 0 47 22 07 0 54 0 91 27 74 0 23 27 91 0 28 0 67 33 92 0 31 34 24 0 41 0 89 SMP 11 04 0 62 11 53 0 64 0 78 21 58 0 47 22 04 0 53 0 92 27 75 0 24 27 92 0 30 0 63 33 93 0 31 34 24 0 40 0 89 9 88 0 56 10 32 0 60 0 76 19 94
55. Surface Downwelling Spectral Irradiance Es 4 W m nm Meee 28 Instrument descripIotm ecd teer einen Ges anita situs dee cu eme Eta puss du acia dde 28 Instrument Calibration and Quality Assurance eeeeeeeeeee n 29 Methodology and Processing Description seseeseeeeeeeeeeeeneen eene 29 Deployment Of the tHSEICUTTIEBL iiiter nire onn enge retos teque cera eee dpa ud uoa 29 Description of processing techniques employed esee 29 Primary Quality Bec kso eerte ys cad auc tee sa ag uss adea co Riu ete aiat R 30 Primary Processing orae ae Sean amat qe enne A A E exui ARRA A oll pdg di 30 Calibration Coefficients iarain Soo ae teen To a teat ette quaudo iude vos Fui ud nde 30 bona PP CE 30 Reference Srono MEE 30 Spectral Sky Radiance Lsky eee 3l Spectral Direct Sun Irradiance E 4 A eese 31 Instrument description CIMEL 318 Sun Photometer eee 31 Instrument Calibration and Quality Assurance esee 32 Methodology and Processing Description essere 32 Deployment of the instrument cane rut hp ede eati Steg ilu tas deba e dee npn suE date 32 Methodology for sample collection 2 1 nre Ler reta ennt stt petet eoa tt aded eoeanpins 32 Primary quality checks before submission of Level 1 data s 32 Calibration coefficients caepit tee a a ae a a Rass 33 T
56. UTR EA E 20 DTA ON EEEE EEA M T E Un sbeatecvtebe 20 InstrithettattoD uten e retro PE epu pap ea t aD E E E R veu uel Dos 20 Instrument Calibration and quality assurance eese 20 Filtration and Storage a pm tentent d er ebeecec tom scent teta Ee Died eo eei oes ede ca CORE ERE t dee 20 Measurement procedures ror ete I tete eO PE HERE NE E LSSRESEEE NOR EU PME ES e ATUS 21 Dato PROCESS IM ous eme tO Bt tUe b a pubs esl ba teet E x xps tatiyseesuteaeents 21 giis ARCEM Dat icun cant caunesns dua seecveh aanpatseousealie EE E E RS 21 Pigments Concentration by High Performance Liquid Chromatography mg m or Lg r e M 23 Instrument descriptions x soot di apie eet ERO HE R peser atra tuere EK RR 23 Instrument Calibration and Quality Assurance eeeeeeeeeeee e 24 Determination of pigment response factors seen 24 Methodology and Processing Description eeeeeseeeeee eren 24 Pigment extraction and sample prepatatiOt uius asia entree orato embed aan drea eh ae 25 Analysis DEOS ADV uinum aie hate A eae ARE agit aby feu obierat de 25 PROCESSING CSCI OM iicet ua m dp ra Peto tt vasa e ERR EH R E t fric esr apibus Ou ce 25 Quali ASSUFTII COS UD eet ots Piet utt Dedit optet ee 26 Sample SIOFdBE sc iine i thv HEU IO Sa ais uer Dial oru s 26 Tanarionsoss cese dann iMt ted pat p diebns pS btts 26 PRE TROT COS otof o MIL Ue Hc A Reni E E Hagb ett Lie ubl Dei E 26
57. W Inc 8 Wetlabs ECO BB User s Guide Revision AH 9 J M Sullivan et al Measuring optical backscattering in water in Light Scattering Reviews 72013 Springer Berlin Heidelberg p 189 224 10 G Dall Olmo et al Particulate optical scattering coefficients along an Atlantic Meridional Transect Opt Express 20 21532 21551 2012 11 G Dall Olmo et al Significant contribution of large particles to optical backscattering in the open ocean Biogeosciences 6 947 967 2009 12 G Tilstone and V Martinez Vicente Protocols for the Validation of Ocean Colour Satellite data in Case 2 European Waters ISECA 13 S Tassan and G M Ferrari An alternative approach to absorption measurements of aquatic particles retained on filters Limnol Oceanogr 40 1358 1368 1995 14 G Neukermans et al Optimization and quality control of suspended particulate matter concentration measurement using turbidity measurements Limnology and Oceanography Methods 10 1011 1023 2012 15 D W Van der Linde Protocol for the determination of total suspended matter in oceans and coastal zones Ispra Italy 16 S Davidson M Perkin and M Buckley The measurement of mass and weight Measurement good practice guide Teddington UK 17 S Roy et al Phytoplankton pigments Characterization Chemotaxonomy and Applications in Oceanography2011 Cambridge UK Cambridge University Press 845 28 TABLE of ACRONYMS Acronym
58. a foil covered sealed duran Wear gloves when flask at 20 C handling acetone 3 Any modifications to this protocol must be approved and recorded 5 6 Protocol 6 Preparation of chlorophyll a stock solution Preparation of chlorophyll a stock solution Notes 1 Pure chlorophyll a 1mg supplied by Sigma source Anacystis Lot BCBG31290 Code C6144 1MG 128 50 Stored in freezer in G13 Received Mar2012 Stored 20 2 Take a clean 100mL volumetric flask and rinse with HPLC grade Use acetone in a fume acetone cupboard Wear gloves and safety glasses 3 Take a small clean beaker and rinse with HPLC grade acetone 4 Turn off lights in lab Maintain minimum light required for safe working 5 Open chlorophyll ampoule and transfer contents to beaker Rinse Ampoule usually made ampoule with HPLC grade acetone and transfer rinsings to beaker of glass Break carefully at fracture line Beware of splinters 6 Add acetone to beaker so chlorophyll dissolves 7 Pour chlorophyll solution into volumetric flask Use funnel if necessary 8 Rinse beaker with acetone and transfer rinsings to volumetric flask 9 Add acetone to volumetric so bottom of meniscus is level with volume indicator line 10 Stopper flask and mix gently by inverting 11 Label as follows Chl a stock solution approx 0 1mg L Preparation date Expiration date Name Lab book number
59. a light beam passing through particles retained on a filter to derive the absorbance Apm A on the filter and is then transformed to give the equivalent absorption coefficient apm A m in suspension Instrument description A dual beam spectrophotometer provided with a Spectralon coated barium sulphate degrades with seawater integrating sphere attachment is ideal In dual beam instruments the correction for the difference in the beam efficiencies is automatically performed Tassan and Ferrari 1995 Single beam instruments are not recommended as it is difficult to characterize the baseline and spectral performance of the instrument Mitchell et al 2000 Before sample measurements are performed baseline and spectral noise should be well documented using air air scans to check instrument performance each time the spectrophotometer is switched on Measurements are performed in the spectral range 350 800 nm with a 1nm resolution The instrument photometric accuracy should be at least 0 003A or 0 08 T at 1A 0 002A or 0 05 T at 0 05 A measured with NBS 930 filters Perkin Elmer Lambda specifications Systems with variable slit widths are preferred from 4 nm to below 4 nm A NASA workshop recommended the use of Cary 100 Mitchell et al 2000 and EU FP5 REVAMP workshop showed that the Perkin Elmer range of spectrophotometers higher than Lambda 800 Fig 1 also shows quality optical performance comparable to the Cary range Figure
60. a statussvl art COMMENTS iphvtoplantkon counts protocols in http w westernchannelobservatory org uk data php phytoplankton counts zero values do not mean absolute absence fr amples and may instead reflect the uncertainties of enumerating Phaeocystis in 50 mt subamples This is particularly important when Phaeocystis forms colonies and bhyroplanicon counts cells ml tor y og uk data php e data from 4 nutrients txt org uk data php c ww westernchannelobser vatory org uk data php HPLC m iChla HMPLC wCO sips jgwenrta1998 201 vw c5 code prowco chla Proso YET Ment struct v4 proWco SP struct prowco phaeocyst is struct prowco cosbine pr int pro_header_bgc_wec_seabassv2 expanded_arrays pro 999 0 Er metadata Jabels_end 154 SETIO Type Tine Lon Lar Teper abate aoe Rage Bes PSAL TSM Tot Ch a NO2 NO2 NO3 Ni Si 85 03 1311 000 00 5 oM4 PO4 Phaeocystis globosa Phaeocystis all 00 999 000 Sia 999 000 999 00000 999 00000 00 999 000 55 999 000 999 00000 00 00 999 000 9 99 00 4 8 nnnnnnnnnnnnnnn REFERENCES 1 S B Groom et al The western English Channel observatory Optical characteristics of station L4 Journal of Marine Systems 15 20 50 2009 2 V Martinez Vicente et al Particulate scattering and backscattering related to water constituents and seasonal changes in the Western English Channel Journal of Plankton Re
61. alculated and checked to be within 596 of calibration value Up to 20 samples filters are analysed per day so maximum time of samples in autosampler is 24h Autosampler is maintained at 4 C Reporting Where a pigment is not detected the effective LOD is reported These values are marked in red in the spreadsheet Higher order products eg TChl a are calculated before reporting pigments not detected as effective LOD s Abbreviations recommended in Phytoplankton Pigments 2012 Roy Llewellyn Egeland and Johnsen Eds Cambridge are used The unknowns are reported This is equal to peak area unknowns total peak area all pigments x 100 Reporting rules Retention time and spectra need to match the pigment in question for it to be reported Sample storage Filters are stored at 80 C or in liquid nitrogen until analysed 10 4 0 References Van Heukelem L and Hooker S B 2011 The importance of a quality assurance plan for method validation and minimizing uncertainties in the HPLC analysis of phytoplankton pigments In Roy S Llewellyn C A Egeland E S and Johnsen G Phytoplankton Pigments Characterisation Chemotaxonomy and Applications in Oceanography Cambridge University Press Jeffrey S W Mantoura R F C and Wright S W eds 1997 Phytoplankton pigments in oceanography Guidelines to modern methods Paris UNESCO Sosik H M 1999 Storage of marine particulate samples for light absorption measurem
62. ampling criteria This document builds on MERIS protocols to give more detailed guide lines for the determination of apparent and inherent optical properties of Case 2 waters of the INTERREG 2 Seas area References Doerffer R 2002 Protocols for the validation of MERIS water products European Space Agency Doc No PO TN MEL GS 0043 Fargion G S Mueller J L 2000 Ocean Optics Protocols for Satellite Ocean Colour Sensor Validation Revision 2 NASA Goddard Space Flight Center Greenbelt Maryland pp 125 153 Mueller J L Austin R W 1992 Ocean Optics Protocols for SeaWiFS Validation SeaWiFS Technical Report Series NASA Tech Memo 104566 Tilstone GH Moore GF Sorensen K Doerffer R Rottgers R Ruddick KG Pasterkamp R 2003 Protocols for the validation of MERIS products in Case 2 waters Proceedings from ENVISAT MAVT Conference 20 24 October 2003 Frascatti Italy European Space Agency http envisat esa int workshops mavt_2003 MAVT 2003 802 REVAMPprotocols3 pdf IESCA satellite validation protocols 07 027 FR ISECA 5 In vivo Absorption Spectra of pigmented and non pigmented Particulate Matter ay A m Definition The light transmission of aquatic particles retained on filter Introduction The light transmission measurement of aquatic particles retained on a filter is considered a standard method for the determination of the in vivo particle absorption The analysis consists of measuring the fraction of
63. and page number 12 Wrap flask in foil Label actual flask and foil covering 13 Store top shelf freezer G13 Shelf life 3 months Dispose of old stock solution in fume cupboard 14 Any modifications to this protocol must be approved and recorded 17 5 7 Protocol 7 Preparation of chlorophyll a working standard solution Preparation of chlorophyll a working standard solution Safety Notes 1 Take a clean 100mL volumetric flask and rinse with HPLC grade 90 Use acetone in a fume acetone cupboard Wear gloves and safety glasses 2 Take a small clean beaker and rinse with HPLC grade 9096 acetone Invert on blue paper to dry 3 Turn off lights in lab Maintain minimum light required for safe working 4 Take chlorophyll stock solution from top shelf freezer G13 Check expiry date Mix gently by inverting Decant a few mL into pre washed beaker cover with foil and leave to equilibrate to room temperature Return chlorophyll stock solution to freezer 5 Transfer 3mL stock solution to pre washed volumetric flask 6 Add 9096 acetone to volumetric so bottom of meniscus is level with volume indicator line 7 Stopper flask and mix gently by inverting 8 Label as follows Chl a working standard solution in 9096 acetone Preparation date Expiration date Name Lab book number and page number 9 Wrap flask in foil Label actual fla
64. at all the files needed for data processing have been collected and note the directory of the raw LO data C RAW DATA CONTROL Data collected Y N Software and version LO Data Y N DIRECTORY 22 3 3 4 Processing and Automatic Quality control of data Processing should be documented including both the reference used and the source code location Product confidence flags for particulate absorption are QC 1 flag no negative values in the spectra Taking into account the uncertainty of the measurement negative values less than 0 005 mare flagged QC 2 flag spectral shape raises a flag if a 510 or 532 or 555 or 650 or 676 or 715 a 412 or 440 or 488 QC 3 flag for apny only spectral shape raises a flag if apny 443 apny 665 lt 1 QC 4 flag for age only spectral shape raises a flag if the age 676 aaa 600 ag 710 x 600 676 600 710 21 indicating that there is still a residual peak from pigments D PROCESSING OF DATA 1 Method of processing check a flag if the adet 676 ag amp 600 aga 7 10 x 600 676 600 710 21 L1 Xfactor Beta water and pathlength att Y N DIRECTORY FILE 2 Processed data tests QC1 Automatic check for no negative Y N values DIRECTORY FILE QC2 Automatic check spectral shape Y N raises a flag if a 510 or 532 or 555 or 650 or 676 or 715 a 412 or 440 or 488 DIRECTORY FILE QC3 A
65. ation In a time series as in WCO this should be done every day after washing down with fresh water e The meter should be cleaned if fouling is suspected for buoy deployments e Profiling in very clean waters where signal changes are on the order of 0 01 m may require more frequent cleaning It is important to note whether the instrument has been cleaned or not in the QAD Another important aspect of the optical instruments maintenance is calibration In addition to the absolute calibration performed by the manufacturer there are air tracking and water calibration procedures The methods recommended by the manufacturer are detailed in the instrument manuals and will not be repeated here 7 Both methods are useful to check functioning and stability of the instrument however it is important that the operator is able to perform the calibration with a known repeatability tested by repeating by triplicate the whole procedure including cleaning of the instrument and the setup Repeatability between 0 005 and 0 002 m should be achieved for water calibration at all channels In addition drift in regular calibrations should be monitored and noted Manufacturer acknowledges drifts of 0 01 m in the blue channels However smaller drifts can also be indicative of problems An example of drift is shown in Figure 1 In this example although the calculated slope for the period was 0 001 m d the instrument was sent back to calibration after
66. atory calibration NOTES date dd mm yyyy 3 Measurement check Baseline 0 005 Y N NOTES 21 3 3 2 Documentation of deployment parameters and sample preparation Water samples need to be documented around aspects that may affect the final results of the measurement like light exposure or temperature In particular some aspects should be documented sample collection concentration method and preservation Concerning water sampling important characteristics to note are method of sampling i e Niskin bottle or underway system sampling time position and depth of sample Details on the concentration method should include time before filtration water stored in dark bottles and at what temperature during that period type and brand of filter used and filtering vacuum pressure Sample preservation should contain information on whether the samples were flash frozen in liquid N the length of time in storage before analysis and the temperature of storage Not all the details should be included for each sample in the QAD If the same method is used for all the samples it suffices to reference the method documentation in the QAD B DOCUMENTATION OF DEPLOYMENT PARAMETERS Metadata collected Y N Document Sample collection method Niskin UW Concentration method Document Preservation method Document 3 3 3 Raw data control Verify that the data have been recorded in a file and th
67. by the measurement binned at 0 5 m This 18 definition may change according to the research application envisaged for instance it may be different for thin layers studies This allows Product confidence flags to be attributed on a sample by sample basis Section 3 Product confidence flags mark data that should not be used automatically For the backscattering coefficient in WCO application the flags defined are e QC 1 flag no negative values in the spectra Taking into account the uncertainty on the measurement values less than 0 002 m are flagged D PROCESSING OF DATA 1 Method of processing check L1 Xfactor Beta water and pathlength att Y N DIRECTORY FILE L2 Interpolation Y N DIRECTORY FILE 2 Processed data tests Spectral Plots Y N DIRECTORY FILE QC1 Automatic check for no negative Y N values QC1 0 discards spectra QC1 1 valid DIRECTORY FILE Additional checks on the spectral shape could be included however the uncertainty on spectral shape of the particulate backscattering coefficient remains high therefore this test is not implemented 3 3 Filter pad absorption As opposed to the measurements presented in Sections 3 1 and 3 2 the filter pad absorption measurement is done on a bench based instrument in the laboratory with pre concentrated samples The specific method that will be addressed here are those described in the ISECA protocols 12 wh
68. d must be chosen in such a way that pigments and standard peak areas are comparable After extraction the sample is micro centrifuged for 2 minutes The extract is then injected through a 100 ul loop into the HPLC system Analysis program The solvent systems used are as follows solvent A 70 30 methanol 1M ammonium acetate solvent B methanol The flow rate is 1 ml min with the following gradient Time min A B 0 0 75 25 1 0 50 50 20 0 30 70 25 0 0 100 30 0 0 100 30 1 75 25 39 0 13 25 Processing description Detection wavelengths are 440 nm for chlorophylls and carotenoids and 667 nm for phaeo pigments The chromQuest software automatically outputs integrated peak areas and assign pigment identities but these are checked manually for all samples and re assigned re integrated when necessary using retention times and absorption spectra Individual pigment concentrations Cpi in ng L are calculated as one i Where i is the peak area of the pigment RF is the response factor for the pigment Ba is the ammonium acetate buffer dilution 2 Vi is the volume injected 0 0122 mL V refers to the extraction volume 2 mL IESCA satellite validation protocols 07 027 FR ISECA 25 V refers to the volume of water filtered usually 1 L Ams is the mean peak area of six internal standard injection run with each batch and A is the peak area of internal standard in the sample Quality Assu
69. d solution 5 8 Protocol 8 Preparation of mixed pigments standard 5 9 Protocol 9 Quantification of chlorophyll a working standard solution by spectrophotometry 5 10 Protocol 10 Determination of response factor 5 11 Protocol 11 Preparation of Zapata et al 2000 eluent 6 0 Spreadsheets 6 1 Spreadsheet 1 RS tracker 6 2 Spreadsheet 2 RT tracker 6 3 Spreadsheet 3 Chl a standard 6 4 Spreadsheet 4 Injection reproducibility 6 5 Spreadsheet 5 Method uncertainty 6 6 Spreadsheet 6 Chl ws tracker 2013 7 0 Appendices 7 1 Common pigments consumables and suppliers 7 2 Copy of Thermo Maintenance contract 2013 7 3 List of standards available from DHI 17 18 18 19 20 21 22 23 23 24 25 25 26 27 28 29 31 1 0 Introduction The pigment analysis facility at PML provides fundamental measurements important to remote sensing modelling in situ optics primary production and biogeochemistry It plays an important role in providing quality assured results for National Capability programmes such as the Western Channel Observatory and AMT for PML research program projects and CR funded projects eg ISECA Once collected pigment samples are extracted according to a strict protocol and analysed by High Performance Liquid Chromatography Knowing with accuracy and precision the pigments that are present in phytoplankton is fundamental to many aspects of PML science HPLC can separate upwards of 30 pigments
70. dards The following standards need to be prepared prior to analysis Standard Compound Order Solvent Storage Shelf life Protocol code number Internal standard Trans B Apo 8 9096 20 C 1 month 5 extraction solution carotenal acetone Internal standard stock Trans B Apo 8 10810G 90 20 C 36 5 solution carotenal 1G acetone months Sigma Chlorophyll a working Chlorophyll a 90 20 C 1 month 7 standard acetone Chlorophyll a stock Chlorophyll a C6144 100 20 C 3 6 solution 1MG acetone months Sigma Mixed pigments standard Mixed pigments From Internal 20 C Prepared 8 DHI standard daily extraction solution The internal standard extraction solution and chlorophyll a working standard are prepared from the internal standard stock solution and chlorophyll stock solution respectively The chlorophyll working standard is quantified according to protocol 9 The HPLC response factor of the freshly made chlorophyll standard is determined according to protocol 10 Data from the chlorophyll working standard quantification and response factor determination are recorded in spreadsheet Chl a standard Spreadsheet 3 Four injections of chlorophyll working standard three injections of internal standard extraction solution and one injection of mixed pigments standard are made on each day of analysis 2 6 Performance Management Summary 2 6 1 Pigment Resolution and Retention ti
71. e tests for values outside the normal boundaries of the dataset or very different from other datasets of similar characteristics i e same location but different years or nearby locations The data with this flag could be used with the awareness of the tests that have failed In summary the approach proposed in this document is to apply a series of objective procedures to quantify the quality of data This does not preclude the use of scientific knowledge but rather attempts to incorporate it into some explicit form on the routine data processing 3 1 In water absorption and attenuation This section expands the tests that are then summarised in the individual QAD Appendix 3 11 Instrumentation checks and calibrations A record of the checks and maintenance of the instrument should be kept Regular maintenance after or before deployment should be performed according to the manufacturer protocol 7 e Washing down the exterior of the meter regularly with fresh water reduces possible effects of corrosion This should be done after every cast if possible In a time series as in WCO it should be done once the instrument returns to the laboratory e Ideally the instrument should be cleaned after every cast However during a cruise operations may allow only once per day which should be the minimum frequency This could occur after the last cast of the day or before the first cast of the day It can also correspond with a field calibr
72. e teaa 8 Data Processing was fesse das epatis der tesi ro rie Ye such dd qun S RENE Iba EA aded qe deci E AEA AREST 9 Pathlength Wavelength Correction D i e e patto edente qae 9 RETSPONCES e 10 Backscatter coefficient ACOA m 1 eee 11 Introduce Hbi cidit EE vigo Gel iube a fede feste E oe o LE oam EET 11 The Hobilabs instrumentso isserrat 11 Instrom nt descripto nae aeae reer ea dpi usa dela a SA E EEOAE eut Eene SaR 11 Methodology and data quality control eese 12 Deployment dawn testuetist iss rco vert deus lta A aE a tuteuthea Sela p e duo blu bus o 12 WHO WS cost Pass Gui erp et a Eat Ru tatebsatatezdbvbsiraauadaebeatie gal aa ote eod bg etus 12 PRECAUTIONS and Maintenance feurn te an ede a tei essei du iere bita fe oibus eb sagt Lieb bote pans 13 General cleanin on ceterae eener erret eMe ofr ttf ete bte eds ronde c doutes 13 Pressure transducer ss atout or ecibuncer ta ono ree postea va dob eoa beta uso eu e a aars 13 Data PROCESSING s ois insera a a e grate Pots eee a aa aaa 13 Calibration Coefficient Seni ia a srme lesan aaa e Oa iaa e E oE Saa 13 Calibration eenn Cm 13 The WETLABS iste utente iseis sac cessbesandauncievesaesdpesdcususetundeasbeanteas 14 Instrument HOSEL ROI renea tase snen nent a iens aSa a ia ae ia ETNAS 14 Instrument Calibration and quality assurance eese 14 Methodology and processing description
73. ed around aspects that may affect the final results of the measurement like light exposure or temperature In 26 particular some aspects should be documented sample collection concentration method and preservation B DOCUMENTATION OF DEPLOYMENT PARAMETERS Metadata collected Y N Document Sample collection method Niskin UW Filtration method Document Preservation method Document 3 5 3 Raw data control Data are recorded manually from a visual reading of the balance at PML They are noted on a laboratory notebook when stable for at least 10 seconds It is important to record the location of the raw data C RAW DATA CONTROL Data collected Y N Instrument brand model amp serial N LO Data Y N DIRECTORY 3 5 4 Quality control of data One method to obtain Quality Check the suspended matter data is through the calculation of blank correction B 15 B is a difference between the initial weight of the blank filter and the weight of the blank filter at the moment of final weighing if the B is greater than 0 0003 g check the performance of the balance temperature and humidity near the balance In case no errors are found the QC1 flag should be raised Product confidence If B is greater than 0 0001 g then it should be used to correct the suspended matter data and the QC2 flag should be raised Product science D PROCESSING OF DATA 1 Method of pr
74. ed at some distance from the instrument at least 50 cm It is recommended to take Esky and Es measurements in sequence Description of processing techniques employed Primary quality control includes data screening for any rapid change in Es A and ensuring that profiles are smooth in log linear scale IESCA satellite validation protocols 07 027 FR ISECA 29 The data presented for the level 1 archive must be corrected for dark The most recent calibration factors available should be included in the level 1 file If any consistent change in calibration is found during field work activities then the data should be re submitted to the level 1 archive with a modified calibration date Primary Quality Checks Stability of skylight Removal of records with bad tilt roll higher than five degrees Removal of records below instrument noise Primary Processing Normalization of Ed z A and Lu z making use of Es Calculation of Kd A and K1 A Calculation of Rrs 0X making use of Lu 0 4 and Ed 07 4 Spectral consistency of Kd A and K1 Calibration coefficients Calibration and quality assurance as per NIST Limitations Sensor tilt induced by ship roll should produce significant errors on normalized values of Lu and Ed Surface effects induced by rough sea can induce significant noise in Lu and Ed measurements Non stable illumination during the sequential measurements of Esky and Es could induce erroneous values of r Re
75. eed to be documented around aspects that may affect the final results of the measurement like light exposure or temperature In particular some aspects should be documented sample collection concentration method and preservation Details on the concentration method should include time before filtration water stored in the dark and at what temperature during that period MQ available or not type there are several types that have a nominal 0 2um pore size and brand of filter used and filtering vacuum pressure 24 Sample preservation should contain information on whether the samples were or not preserved with Sodium Azide NaN3 12 Not all the details should be included for each sample in the QAD If the same method is used for all the samples it suffices to reference the method documentation in the QAD and note any deviations on individual samples B DOCUMENTATION OF DEPLOYMENT PARAMETERS Metadata collected Y N Document Sample collection method Niskin UW Filtration method Document Preservation method Document 3 4 3 Raw data control Verify that the data have been recorded in a file and that all the files needed for data processing have been collected and note the directory of the raw LO data C RAW DATA CONTROL Data collected Y N Software and version LO Data Y N DIRECTORY 3 4 4 Processing and Automatic Quality control of data Similar procedures to
76. en scientists can only examine data after instrument deployment Measurements collected should be extracted processed and visualised as soon as possible so that faults can be quickly identified In a time series as in WCO the data should be downloaded after the outside of the instrument and its tubes have been cleaned Section 3 1 2 A raw data control that ensures the existence of data and an initial processing producing quick views plots should be done within 24 hours of the data collection Figure 2 120411unf a440 120411unf a488 depth rn depth r 6 0 0 2 0 4 0 6 0 8 1 0 0 0 0 2 0 4 0 6 0 8 1 0 depth m depth m 12 Figure 2 Example of a quick look plot of an absorption profile at four wavelengths i e 440 488 510 and 555 nm at L4 ona given date 11 04 2012 C RAW DATA CONTROL Data downloaded Y N Software and version LO Extraction Y N Software and version Quick plots Y N dd mm yyyy 3 1 4 Processing and Automatic Quality control of data Once the raw data control has been done and the initial QC plots show that data have been collected within the real range i e non negative the data should be put through the automatic processing and quality check chain The initial step is to follow the recommended processing from the manufacturers 7 This applies the temperature and salinity water calibration and scattering corrections to the data producing L1 data After this
77. ensures the existence of data and an initial processing producing quick views plots should be done within 24 hours of the data collection Figure 5 17 120411flt bbp 470nm 10 20 30 Depth m 40 50 60 0 000 0 002 0 004 i 6 008 0 010 Depth m LA o uw a 60 bp mr 120411flt bbp 880nm _ _ SSS Depth m 0 000 0 002 0 004 G G06 6 008 0 010 Figure 5 Example of quick looks of vertical profiles not binned for the three channels of a Wetlabs bbp mr backscatter meter ECO BB3 120411flt bbp 532nm 0 000 0 002 0 004 0 006 6 6008 4 010 bbp nr C RAW DATA CONTROL Data downloaded Y N Software and version LO Extraction Y N Software and version Quick plots Y N dd mm yyyy 3 2 4 Processing and Automatic Quality control of data Once the raw data control has been done and the initial QC plots show that data have been collected within the real range i e non negative the data should be put through the automatic processing and quality check chain The initial step is to follow the recommended processing from the manufacturers 8 This applies the x factor subtracts volume scattering function values of seawater and applies pathlength attenuation correction to the raw backscattering data to produce L1 data After this interpolation and binning to an equally spaced grid is recommended L2 For the WCO application one sample is defined
78. ents Limnol Oceanogr 44 1139 41 Roy S Llewellyn C A Egeland E S and Johnsen G 2011 Phytoplankton Pigments Characterisation Chemotaxonomy and Applications in Oceanography Cambridge University Press Zapata M Rodriguez F and Garrido J L 2000 Separation of chlorophylls and carotenoids from marine phytoplankton a new HPLC method using a reversed phase C8 column and pyridine containing mobile phases Mar Ecol Prog Ser 195 29 45 11 5 0 Protocols 12 5 1 Protocol 1 Setting up for HPLC analysis Setting up HPLC for analysis Notes Switch on LC modules at instrument if not already on Go to My computer D Data and create folder with todays date eg 20131130 Copy methods from last analysis to todays folder Zapatashutdown shutdown method date seq sequence file date Pig12zapatav1 HPLC method file date Pigzapataproc data processing method Pigzap40b ape pretreatment method Change dates of filenames to todays date Open Chromquest software if not already open Double click on Accela to open instrument control Load todays HPLC method file from todays folder Go to Control menu and select Instrument status Go to PDA tab and switch lamps on Note lamps on time on pigment analysis QC sheet Ensure lamps on for 1 hour prior to starting analysis Check system visually check sufficient solvent in solvent reservoirs No visible particles in tubes or bottl
79. er The presence of air bubbles in the measurement chambers may irreparably affect measurements References Home Page of Oregon State University College of Oceanic and Atmospheric Sciences Environmental Optics 1999 http photon oce orst edu ocean instruments ac9 ac9 html Pegau W S and J R V Zaneveld Temperature dependent absorption of water in the red and near infrared portions of the spectrum Limnol Oceanogr 38 188 192 1993 Twardowski M S J M Sullivan P L Donaghay and J R Zaneveld 1999 Microscale quantification of the absorption by dissolved and particulate material in coastal waters with an ac 9 Journal of Atmospheric and Oceanic Technology 16 691 707 WET Labs 2011 ac 9 Protocol Document Revision Q 10 April 2011 Zaneveld J R V J C Kitchen A Bricaud and C Moore 1992 Analysis of in situ spectral absorption meter data Ocean Optics XI G D Gilbert Ed Proc SPIE 1750 187 200 Zaneveld J R V J C Kitchen and C C Moore 1994 Scattering error correction of reflecting tube absorption meters Ocean Optics XII S Ackleson Ed Proc SPIE Vol 2258 44 55 IESCA satellite validation protocols 07 027 FR ISECA 19 Coloured dissolved organic material m Also known as Yellow substance Chromophoric dissolved organic material Gelbstoff Definition Coloured dissolved organic material is defined as the fraction of organic matter which passes through 0 22 um pore size filter
80. er a reflective bottom and under bright solar illumination light reflected into the windows may cause high noise levels or in extreme cases saturation In such situations it may be advantageous to mount the sensor horizontally so that the backscattering receivers do not face the bottom Windows HydroScats have acrylic windows that are easily scratched Minor scratches will not seriously compromise the measurements but the windows must be treated carefully to avoid abrasion Do not use acetone or abrasive cleaners Do not over clean the windows Unless the windows become visibly dirty during use it is usually sufficient to clean them once daily with soap or alcohol and a soft cloth then rinse them with clean water whenever they are removed from the water IESCA satellite validation protocols 07 027 FR ISECA 12 Precautions and maintenance e The instrument windows should always be protected Ensure that the instrument face is covered whenever the instrument is not in use Do not use acetone to clean any part of the instrument Thoroughly clean the HydroScat with fresh water before storing it Avoid letting the sensor sit in direct sunlight on deck If the water temperature is very different from the temperature on deck let the instrument stabilize in the water for 10 minutes before collecting data General cleaning After deploying the instrument rinse it thoroughly with fresh water and rinse the windows with distilled or deionized water
81. erating program calibration factors and the capacity for storing over measurement files reside on the standard 256 Kbyte PCMCIA card Personal Computer Memory Card International Association The PR 650 incorporates Automatic Adaptive Sensitivity that optimizes the detector signal to noise for accurate measurement regardless of the signal level QA and data processing details The radiance measurement of the reflectance standard is used to calculate above water L downwelling irradiance E z where p is the reflectance of the reflectance panel standard 99 The standard is measured under an angle of 45 degrees panel L The MERIS reflectance can than be calculated as o Tis where Lw the water ad leaving radiance is calculated as L L p L With p is the effective Fresnel reflection coefficient for the wind roughened sea surface IESCA satellite validation protocols 07 027 FR ISECA 36 Instrument Calibration and Quality Assurance The absolute radiometric response for each radiometer is determined at the start and end of the project using an NIST standard 1000W lamp A Photo Research near Lambertian calibrated spectralon reflectance standard 5 cm of about 99 reflectance is used as a reflective standard to calibrate the instrument Because reflectance is a relative quantity the absolute radiometric calibration has no influence on the accuracy of the derived water leaving reflectance provided
82. ering meter The HydroScat 6 has six independent channels each sensitive to a different narrow range of optical wavelengths Hobilabs will configure the instrument to 3 wavelength pairs For the REVAMP project the following wave bands have been selected 420 442 488 550 671 850 plus fluorescence excited by 442 and emitted to 671 The source produces a beam of light in the water and the detector collects a portion of the light that is scattered out of that beam by the water Each source beam originates from a light emitting diode LED selected to match the desired measurement Methodology and data quality control Deployment The HydroScat can be suspended vertically from the metal eye on the connector endcap or strapped to another support If mounting it to another structure the finish on the case should be protected from direct metal contact To ensure that the HydroScat does not detect reflections from any other objects It is best to keep a clear 30 cone in front of the detection windows for at least 1 meter Even objects that appear very non reflective or are well out of the nominal sampling volume can create substantial offsets in the backscattering measurement The operator should manually check that readings are not unnecessarily elevated by interference from other reflective objects The sensor should normally face directly down in the water to minimize the effect of background illumination However in shallow water ov
83. erived from Beer Lambert s Law Chl a conc gL abs 662 abs 750 extinction coefficient Use extinction coefficient for chl a in 9096 acetone 88 67 Lg cm Jeffrey et al 1997 If absorbance at 750nm is non negligible eg gt 0 002 absorbance reading at 700nm may be used 15 Any modifications to this protocol must be approved and recorded 19 5 10 Protocol 10 Determination of response factor Determination of response factor Safety Notes e Perform 6 injections of chl a ws on HPLC N Integrate areas of chl peaks and any impurities 440 nm chromatogram Input data to results sheet chl a standard Fill in remaining information in results sheet chl a standard Print chl a standard sheet and stick in chl standards lab book our BR w If change from calibration value is gt 5 prepare and quantify new chl a working standard and repeat determination of response factor If change from calibration value is still gt 5 prepare new chl a stock solution and new chl a working standard and repeat determination of response factor Any modifications to this protocol must be approved and recorded 20 5 11 Protocol 11 Preparation of Zapata et al 2000 eluent Preparation of Zapata et al 2000 eluent Safety Notes Equipment list Stirrer plate magnetic flea remover 10 mL measuring cylinder 50 mL measuring cylinder 1L flask
84. ers scattering by particles 2 750 nm is not negligible since scattering and absorption by detritus increase with decreasing wavelength Tassan amp Ferrari 1995 The experimental and data processing methods of Tassan and Ferrari 1995 equations 11 to 14 are recommended with some modifications to convert the measured absorbance of the filter retained particles into the equivalent particle suspension absorption Four measurements are therefore required for each sample two transmission and two reflectance The instrument baseline for the integrating sphere should be recorded The data is processed by fitting the detrital curve to an exponential with an offset which takes into consideration the baseline The particulate absorbance spectra is scaled to the exponent of the detrital curve t is defined as the ratio of 1 Tsd 1 Tsp where Tsd is the transmission of diffuse light through the filter and Tsp is the transmission of parallel light The following routine is used to calculate c t 1 171 0 2615 0 00013 Equation 1 where a is the absorption in transmission mode either of the pigmented or de pigmented sample given as follows 1 a log B Equation 2 s where st is the sample transmission The wavelength specific absorption coefficient is calculated from the absorbance of the material in suspension Asus a A e Equation 3 where X is the ratio of the filtered volume to the filter clearance area and C is t
85. es View syringe bubble free Check waste level in HPLC solvent waste Prime pump open purge valve attach syringe In pump tab of instrument status set to 100 A 1000 uL min and set to flow Allow minimum of 4 mL to fill syringe then switch to 10096 B and allow to fill further 4 mL Empty in between if necessary stop flow before removing syringe Stop flow close purge valve remove syringe and dispose of solvent Wear gloves and glasses Mobile phase contains pyridine and acetonitrile dispose of in labelled solvent waste in fume cupboard Set pump flow to 200 uL min 100 A Start flow Select baseline icon to download method Then select stop icon to stop viewing baseline Note time on pigment analysis QC sheet Downloading method switches on thermostats for column compartment and autosampler 10 Dispose of old sample vials from autosampler 11 Empty wash and refill reservoir 1 RV1 with fresh MQ 12 Place colourless empty vials in positions 1 3 5 etc 13 Put chl a working standard 4 separate vials internal standard 3 separate vials and diluted mixed pigments 1 vial in amber vials and insert in autosampler in this order chl wsd mixed pigments chl ws1 3 internal std 1 3 Note time on pigment analysis QC sheet 14 Load sequence and edit ie correct methods datafiles and sample names 15 When lamps have been on for at least 45 mins monitor baseline
86. es retained on filters Limnol Oceanogr 40 1358 1368 Tassan S and G M Ferrari 1998 Measurement of light absorption by aquatic particles retained on filters determination of the optical pathlength amplification by the transmittance reflectance method J Plankt Res 20 1699 1709 Tassan S Ferrari GM Bricaud A Babin M 2000 Variability of the amplification factor of light absorption by filter retained aquatic particles in the coastal environment Journal of Plankton Res 22 659 668 Tassan S and G M Ferrari 2002 A sensitivity analysis of the Transmittance Reflectance method for measuring light absorption by aquatic particles J Plankt Res 24 757 774 IESCA satellite validation protocols 07 027 FR ISECA 10 Backscatter coefficient B 6 m 1 Introduction Few historic data exist on the variation in shape of the volume scattering function B 0X in the backward direction The most widely published data are those of Petzold 1972 and Balch et al 1994 who used a general angle scattering meter Mueller et al 2000 to measure B 8A for marine hydrosols Recent studies have shown however that a relationship between the measurement of the volume scattering function in one angle and the total backscattering coefficient exists and can be simplified with the use of a constant value Boss and Pegau 2001 Several values for the different angles of measurement have been proposed Maffione and Dana 1992 Boss and Pegau
87. etry 150 x 2 1 mm 3 5 um particle size Column thermostated at 25 C Mobile phase A methanol acetonitrile aqueous pyridine 0 25M pyridine 50 25 25 B methanol acetonitrile acetone 20 60 20 v v v Flow rate 200 uL min See protocol for instructions for preparing mobile phase Internal standard Trans B Apo 8 carotenal Stock solution of internal standard prepared by dissolving 0 01 g of trans B Apo 8 carotenal in 100 mL of 9096 acetone The stock solution is stored in a foil covered sealed flask at 20 C The internal standard extraction solution is prepared by adding 100 pL of stock solution to 250 mL of 90 acetone The internal standard extraction solution is stored in a foil covered sealed flask at 20 C The internal standard extraction solution is used at room temperature 21 C Pipette accuracy is determined by three weighings of internal standard extraction solution 2 mL prior to addition to samples 2 mLs of internal standard extraction solution is added to each filter in a screw cap centrifuge tube The centrifuge tubes are maintained tightly sealed on ice in the dark Disruption method and time Sonication probe 35 seconds Total soak time 1hr Clarification procedure Centrifugation then filtration 0 20 um 17 mm Teflon syringe filter straight into vial Injection procedure and Pretreatment program Pigzap40b ape volume Autosampler injection procedure Draw 200 uL extract draw 40 uL water deposit 240 uL in e
88. every 10s During measurements wind speed is recorded and sea sun and sky state conditions are noted especially if variable because of cloud movement or floating matter The ship position and orientation are monitored for drift Lens caps are used to protect all three sensors except during the 10 minute measurement sequence Measurements can also be made underway for a ship heading of 135 relative to sun providing a transect of reflectance spectra For such measurements the lenses are inspected at the end of the transect and any spray droplets are noted During such measurements visual checks are made of the sea surface for variability such as fronts or floating material and the ship heading is monitored Description of processing techniques employed Data is acquired with the MSDA software v1 94 in 2001 2002 using the file recorder function and calibrated radiometrically using nominal calibration constants Dark values are removed with the dynamic offset function which uses blocked photodiode array channels Calibrated data for E A L A and Lwy A is interpolated to 2 5nm intervals IESCA satellite validation protocols 07 027 FR ISECA 40 and exported to Excel for recalibration to the MERIS Validation Team standard and for further processing Preprocessing Quality Checks The multitemporal dataset is screened to e Remove dropout incomplete spectra e Avoid measurements during temporal fluctations of E A arising mainly
89. ferences Mueller J L amp Austin R W 1992 Ocean Optics Protocols for SeaWiFS Validation SeaWiFS Technical Report Series NASA Tech Memo 104566 5 43 pp Walker J H Saunders R D Jackson J K amp McSparron D A 1987 NBS Measurement Service Spectral Irradiance Calibrations Report NBS SP 250 20 National Bureau of Standards Gaithersburg MD 20899 USA IESCA satellite validation protocols 07 027 FR ISECA 30 Spectral Sky Radiance La4 A Spectral Direct Sun Irradiance E A Both at wavelengths 440 670 870 940 and 1020nm Instrument description CIMEL 318 Sun Photometer Figure 8 CE 318 Sun photometer The CIMEL Paris France CE 318 Sun photometer is a radiometer designed to perform atmospheric studies specifically to determine the optical characteristics of the aerosols It is made up of three parts a programmable box that controls the measurement sequences a mobile device with two rotational axes azimuthal and zenithal asensor head fixed on the mobile device The instrument is powered with solar panels and rechargeable batteries The optical part of the instrument includes at least five filters four to study the aerosols characteristics 440 670 870 1020 nm 10 nm wide and one to determine the water vapour 940 nm 10 nm wide The filter wheel includes a dark mask which is used to determine the dark current Between the filter wheel and the electronic part there are two collimato
90. follow the recommendations given in Jeffrey at al 1997 and revised in Roy et al 2011 D D D D D D u A Figure 5 Agilent system diode array detector and pumping system for High performance liquid chromatography Instrument description Current NASA protocols recommend Agilent Technologies Beckman ThermoQuest Waters Associates HPLC systems for the determination of phytoplankton pigment IESCA satellite validation protocols 07 027 FR ISECA 23 concentrations for ocean colour satellite validation Mueller et al 2003 The minimum requirement for the HPLC system is A Diode array detector 190 800nm pumping system vacuum degasser system controller A Reverse phase column A Computer equipped with hardware and software e g ChromQuest A 100 ul sample loop e g Rheodyne An Air compressor A Centrifuge A temperature controlled autosampler is optional but highly recommended for increasing the through put of samples The C method of Wright et al 1991 is recommended by SCOR and separates more than 50 chlorophylls carotenoids and their derivatives using a ternary gradient system Instrument Calibration and Quality Assurance Determination of pigment response factors The HPLC system is calibrated with the pigment standard obtained from VKI Concentrations of the pigment standard are given from VKI but are also checked using a spectrophotometer The extinction coefficients
91. g Routine 8 13 Semiquantitative 5 8 Quantitative 3 5 State of the art lt 2 lt 3 Table 1 Values of average precision for TChl a and PPig and corresponding performance ratings as described in Van Heukelem and Hooker 2011 2 9 HPLC Calibration Calibration is performed at least annually for the following pigments Chl c3 Chl c Peri But fuco Fuco c Neo Pras Viola Hex fuco Diadino Diato Allo Zea Lut Gyro de Chl b DVChl a Chl a BB Car In Oct 2012 calibration curves were carried out for fourteen pigments and single point calibration carried out for five pigments For single point calibrations four injections of each standard were performed For calibration curves the standards were used to prepare a dilution series comprising three solutions bracketing the LOQ and three bracketing the expected sample concentration New calibration values are expected to agree with the previous calibration within 5 96 providing there have been no changes to the HPLC method The LOD s LOQ s reponse factors and effective LOD s are given in Table 2 Pigment LOD LOQ RF RF If no which used Effective LOD for 2 mL extraction determined volume and 3 L filter volume ng injected ng L ug L Chl a 0 07 0 23 1 196E 05 Y 2 87 0 003 Chl c3 0 03 0 09 4 55E 06 Y 1 09 0 001 MV chlc3 0 03 0 09 4 55E 06 N Chl c3 1 09 0 00
92. g term used in the determination of transmittance may cause errors in the measured optical density spectra of mineral particles They therefore revised the methods by conducting the transmission and reference measurements not referenced to a blank filter then measuring the optical transmission of the blank filter separately in the same way Tassan and Ferrari 2002 References Mitchell GB Bricaud A Carder K Cleveland J Ferrari G Gould R Kahru M Kishino M Maske H Moisan T Moore L Nelson N Phimney D Reynodls R Sosik H Stramski D Tassan S Trees C Weideman A Wieland J Vodacek A 2000 Determination of spectral absorption coefficients of particles dissolved material and phytoplankton for discrete water samples NASA Tech Memo 209966 in GS Fargion and JL Mueller Eds Ocean Optics Protocols for Satellite Ocean Colour Sensor Validation Revision 2 NASA Goddard Space Flight Center Greenbelt Maryland pp 125 153 Mueller J L and R W Austin 1995 Ocean Optics Protocols for SeaWiFS Validation Revision 1 NASA Tech Memo 104566 Vol 25 S B Hooker and E R Firestone Eds NASA Goddard Space Flight Center Greenbelt Maryland 67pp Roesler CS Theoretical and experimental approaches to improve the accuracy of particulate absorption coefficients derived from the quantitative filter technique 43 1649 1660 1998 Tassan S and G M Ferrari 1995 An alternative approach to absorption measurements of aquatic particl
93. gravimetrically or by pressurizing the tank and measuring the resulting offsets Calibration can simultaneously be done for both a andc pressurizing the tank is then recommended or for each one successively Milli Q water is stocked into a clean tank polycarbonate carboy for example at least 12h before the measurement to allow for degassing Water can be checked for particles by pointing a helium neon laser through a glass beaker in the dark and looking for light flashes that indicate particles big flashes or air bubbles small flashes Again all tubing must be black or covered with black tape at least the 20 cm at the flow inlet and outlet The instrument flow tubes and optical windows is cleaned using soap water and methanol Water temperature must be recorded several times during the calibration if necessary for post correction see 4 Data Post Processing Measurements are taken for about 30 seconds with the WETVIEW software the measured offset must be stable within 0 005 for each wavelength Average a portion of stable data Such a sequence is repeated 2 times opening and cleaning the instrument each time and the measured offsets must not differ by more than 0 005 In particular during the calibration one has to check for bubbles that can induce large spikes in the data recorded After correction for temperature effect see 4 Data Post Processing the resulting mean offsets are averaged and subtracted from the i
94. he QAD headed Data Tape and Documentation for Banking lies with the authority which is archiving the data since these aspects refer to the data tape or disc submitted for banking 3 DATA QUALITY PROCEDURES FOR BIO OPTICAL MEASUREMENTS Optically active components and inherent optical properties are relevant for satellite validation Data quality measures introduced in Section 2 i e instrumentation checks and calibrations documentation of deployment parameters automatic quality control and oceanographic assessment are applied to this category of data hereafter In particular the following measurements will be addressed e In water bio optical variables o Total absorption coeff a o Total scattering coeff b o Total backscatter coeff b e Discrete bio optical variables o Particulate absorption coeff a part o Yellow substance or Dissolved CDOM absorption coeff YS e Discrete biogeochemical variables o Chl a concentration by HPLC spectrophotometric fluorometric CHL o Total suspended matter TSM Data quality Samples failing the quality control tests are not removed from the dataset rather flagged with two types of flags similar to Earth Observation flags 6 e Product confidence flags this flag is raised if a measurement fails any of the tests for values within physical boundaries of the variable The data with this flag should not be used e Product science flags this flag is raised if a measurement fails any of th
95. he particle concentration Absorbance of the material retained on the filter is converted to absorbance of the material in suspension using a pathlength wavelength correction factor see below Pathlength Wavelength Correction f e The amount of sample filtered should yield an optical density at 675 nm of between 0 05 amp 0 25 A and with a blue absorption lt 0 4 A High suspension absorbance leads to increasing errors when applying p Mitchell et al 2000 e Few p values have been reported for Case 2 waters Tassan amp Ferrari 1998 For the purpose of data storage B is set equal to 2 Roesler 1998 which is based on the assumption that for GF F filters the diffuse absorption of a sample is twice the volume of absorption coefficient IESCA satellite validation protocols 07 027 FR ISECA 9 e Specific D correction should be calculated for specific areas and phytoplankton assemblages and the method of p correction should be recorded Accuracy The overall error of the filter retained optical particle optical density is 0 002 with an error of 0 015 associated with the variability in physical properties of the GF F filter The corresponding error of the optical density of suspended particles showed that the error increased with increasing optical density from 0 0015 at an OD of 0 05 to 0 027 at 0 59 OD Whilst the T R method detritus rich coastal waters in mineral laden waters Tassan and Ferrari 2002 reported that the light scatterin
96. hen data are transferred from the originating group to a national or international data centre it is sometimes required that the data are transformed into a standard exchange format used between data centres The general experience of data centres is that the processing of data sets into standard exchange format is best carried out by the data centre itself and the originator is only required to provide the data in a well documented format which is acceptable to both the originator and the data centre This avoids the introduction of further errors by requiring data originators to use unfamiliar software and formats 2 2 Data quality control measures for data managers It is assumed that data managers have no previous knowledge on a specific discipline i e bio optics in this particular case hence most of the checks are for consistency and completeness of data They have to be ensured by following the procedures below Test of format coding Check of incoming data set against location and identification errors Tests of fixed and computed limits Tests according to climatological standards e g Levitus Asheville climatology Visual inspection Duplicates check Parameter screening Oceanographic and meteorological assessment 2 3 Quality assurance documents Quality Assurance Documents QADs summarise the data validation procedures applied to metocean data sets They are essentially check lists indicating the procedu
97. here A refers to wavelength was calculated from the optical density and the cuvette pathlength and baseline offset was subtracted from aCDOM Data have been processed using published methods 8 BOX 1 Historical summary of variables sampling Table and recent sampling per year Bar Charts Measurement 1999 2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011 2012 2013 2014 BHPLC a SPM B Optical casts aL d 2008 2009 2010 2011 2012 2008 2009 2010 2011 2012 Year Year In situ absorption and backscattering coefficients In situ optics at L4 combined a WETLabs ac 9 to derive the particle scattering bp and total absorption a and a WETLabs VSF 3 to measure particle backscattering bpp The ac 9 measures absorption a and attenuation c at nine wavelengths 412 440 488 510 555 630 650 676 and 715 nm with a spectral resolution of 5nm and a measurement accuracy 0 005 m 1 The calibration has been checked using pure water calibrations and by the manufacturer The data processing included the correction of measurements using the pure water offsets the temperature and salinity correction using data from the SeaBird CTD and the scattering correction using the Zaneveld method following the recommendations from the manufacturer by can then be obtained by subtraction of the absorption fro
98. iance Coe2 sun_radiances_cal Coe3 sky_radiances_cal Limitations Cloudless conditions are required References University du Littoral C te d Opale March 1998 Ground based atmospheric measurements during the COLORS experiment Report Version 1 0 Holben et al 1999 AERONET A Federated Instrument Network and Data Archive for Aerosol Characterization Remote Sensing of Environment 66 1 16 1998 IESCA satellite validation protocols 07 027 FR ISECA 33 Total Suspended Matter TSM g m Also known as Suspended particulate material Instrument description Electro balance Definition The net weight of material collected on a GF F by sea water filtration Units mg I g m Instrument calibration and quality assurance The electrobalance should be accurate to at least 10g The electro balance zero should be checked before weighing Methodology Filter preparation e GH F filters 0 7 um are pre ashed at 450 C for 1 hr e Filters are then pre washed in MilliQ to remove friable fractions that can be dislodged during filtration Soak not more than 20 filters at a time together for 5 mins in 0 5 1 of MilliQ Place the filters on the shiny surface of clean aluminium foil Dry the filters in a hot air oven at 75 C for 1 hr Store filters in a dessicator with dry silica gel Pre weigh dry filters to 5 significant figures noting the temperature and humidity in the weighing chamber Filtration e A volume of
99. ich are based on the work from Tassan and Ferrari 13 In practice the analytical method comprises the direct measurement of the particulate absorption coefficient apart and depigmented particulate absorption or non algal particles absoption anap and indirectly the phytoplankton absorption apny coefficients 19 3 3 4 Instrumentation checks and calibrations The instrument should have regular absolute calibration and tracking for drifts The absolute calibration is made by the engineer from the manufacturing company The dates of these absolute calibrations should be recorded In addition the tracking of the instrument response is recommended At PML the tracking is done using Holmium Oxide filters that provide a known absorbance A at different wavelengths and three intensities Regular monitoring of the A from standards allow for the construction of time series Figure 6 and the derivation of spectral values of change year Figure 7 0 268 0 26 A 440 A 590 0 266 h ad 0 255 0 25 OS 0 262 ee 636 w 0 245 0 258 0 24 4 0 264 04 01 01 04 10 06 07 09 04 12 12 14 04 01 01 04 10 06 07 09 04 12 12 14 0 24 A 465 0 235 2 0 23 e e 0 225 0 22 T 04 01 01 04 10 06 07 09 04 12 12 14 0 24 A 546 0 235 0 23 0 225 0 22 T T 04 01 01 04 10 06 07 09 04 12 12 14 0 26 A 635 0 255
100. into a labeled cryovial and flash frozen in liquid nitrogen before being transferred to storage in liquid nitrogen or at 80 C 2 2 Sample Storage Filters for phytoplankton pigment analysis should be stored at 80 C or in liquid nitrogen Storage at this temperature has been assessed for up to 1 year and found to be satisfactory Sosik 1999 2 3 HPLC instrumentation and performance HPLC instrumentation needs to be set up and performance checked prior to extraction of samples Samples must be analysed within 24 hours of extraction so instrument problems need to be diagnosed before samples are extracted The HPLC instrument should be properly serviced and maintained HPLC set up includes ensuring sufficient solvent available waste bottles are empty priming pump to ensure solvent lines are bubble free giving detector lamps sufficient time to warm up equilibrating column and preparing mixing vials and milliQ reservoir Detailed instructions for HPLC set up are given in the protocols section Protocol 1 Details of time given for lamps to warm up back pressure and stability etc are recorded on the Pigment analysis QC sheet Protocol 2 The first injection of the day is a sample of the chlorophyll working standard This run is discarded ie not used for data The second analysis of the day is a standard of mixed pigments This sample is used to check the performance of the instrument in terms of resolution 4 The resolution of four sets
101. ity Assurance The calibration methodology is fully described in the NASA SeaWiFS protocols Mueller amp Austin 1992 In summary the irradiance sensors are calibrated using an FEL 1000W lamp traceable to the NIST scale Walker et al 1987 while the radiance sensors can be calibrated with an integrating sphere or with an FEL 1000W lamp and a reference 99 reflectance plaque The sensors are referenced to the JRC NIST traceable standard lamp through a reference set of sensors maintained by JRC On each deployment the actual offset is determined by taking a dark reading immediately before deployment Methodology and Processing Description Deployment of the instrument The optical measurements should be taken in stable illumination conditions The Ed and Lu sensors must be deployed towards the sun or the brightest part of the sky i e the ship or the platform should not shade the instrument The lowering and raising speed of the in water profiling system used for Ed and Lu measurements should be adequate for Case II waters There should be 100 samples for each optical depth when the Kd 490 is 0 25m and for SATLANTIC instruments this corresponds to 0 3 m s Where waters are more turbid a lower speed should be used The pressure sensor should be checked prior to deployment to remove the effect of on barometric pressure changes Esky measurements are taken by shading the direct sun irradiance to the Es sensor making use of the small disc locat
102. l trends checked Y Trend on position Y F FILE INFORMATION AND DOCUMENTATION FOR BANKING 1 File Data in format as specified Duplicates check Coherence of linked values Test according to climatological standard Reference file name Y 2 Documentation standard documentation provided G REPORTING AND DATA PRESENTATION 1 Report interim final 2 Data presentation interim final 3 Data submitted for banking Optics group Calibration Filesvac9calsvac90277Wetlabs cal gYoutput historyV J opticsdatabasev00 L4 yyyy L4_yyyymmdd Wetlabs WAPv4 27 J opticsdatabasev00 L4 1_raw_data_proc_sources J opticsdatabasev00 L4 2_database_creation_sources J opticsdatabasev00 L4 3_DatasetQC_sources pro_plotter_QC pro J opticsdatabasev00 L4 3_DatasetQC_sources QC_spectral_values_v3 pro J opticsdatabasev00 L4 3_DatasetQC_sources QC_spectral_values_v3 pro Notes Eliminatory test Automatic Flag code Not eliminatory test Contact Person vmv andy perkin QUEST vmv vmv vmv vmv ISECA Guidelines for Quality Control of bio optical measurements in Case 2 European Waters APPENDIX The Pigments Manual A Complete Description of Pigments Analysis at PML Version 1 July 2013 Ruth Airs seca INFORMATION SYSTEM ON THE EUTROPHICATION Investing in your future OF OUR COASTAL AREAS Crossborder cooperation programme 2007 2013 Part financed by the European Union
103. lane procedure the CE 318 points at the sun and takes measurements at different zenith angles in the sun plane During the Sun procedure the CE 318 points at the sun and takes irradiance measurements for each wavelength these measurements are repeated 3 times to check the stability Methodology for sample collection CE 318 data are regularly transmitted to the AERONET server at NASA GSFC through a Satellite link Holben et al 1998 Data are then downloaded twice a month by ftp to the LISE ULCO laboratory to produce calibrated data Several aerosol high level products i e scattering phase function aerosol downward fluxes are generated ULCO 1998 Primary quality checks before submission of Level 1 data e Screening data for rapid variability temporal and angular of measurements taken in the principal plane off solar views e Screening data for rapid variability temporal and angular of measurements taken in the almucantar e Checking the symmetry of the almucantar versus the solar plane e Checking the variability of the triplet of sun irradiance measurements e Screening data for very rapid temporal variability of the optical thickness e Thresholding of the sun irradiances with boundary values e Checking the spectral dependency of the optical thickness IESCA satellite validation protocols 07 027 FR ISECA 32 Calibration coefficients The calibration coefficients adopted are as follows Coe1 sun_exoatmospheric _ irrad
104. le for any use that may be made of the information contained therein Contents 1 SUMMARY 4 2 BIOLOGICAL AND BIO OPTICAL SAMPLING DURING ISECA essent 4 v PMEM Aic T 5 2 1 1 Sampling of a time series at L4 and E1 ou cccccsssssseceeeeeceesessaeseeeeseesseseeaeeeeeessesseseaaees 5 2 1 2 Sampling at a transactio dete tenir ere couse ae EEE E Vosa iv en en R ed ta 8 3 DATASETS IMPORTED IN THE WAS ssseesseeseeeeen ene nnee nnne nnne entente tnnt entres inset sn rese tests nenne 8 REFERENCES m M 10 Description of in situ data in ISECA 1 SUMMARY The in situ data in the ISECA Web Application Server WAS are the result of cross border collaboration within the project Overall they constitute a combination of historical data pre ISECA and data collected during the project life 2011 2014 In order to provide a comparable tool across the different partners that at the same time fulfil the user requirements for Eutrophication detection a sub set of variables has been selected to be included in the WAS The variables selected by the consortium for eutrophication monitoring and detection were temperature salinity phytoplankton chlorophyll a concentration total suspended matter concentration dissolved nutrients NO2 NO2 NO3 NH4 SiOH4 PO4 and phytoplankton counts of species indicating of eutrophication For the
105. lts of the method as used for MERIS Validation are presented in Ruddick et al 2002 and Ruddick et al 2006 Figure 10 left System of two radiance and one irradiance sensor installed on steel frame right As installed at prow of ship with irradiance sensor mounted separately to reduce optical interference from mast Instrument description The measurement system consists of three hyperspectral spectroradiometers either TriOS RAMSES or SATLANTIC OCR two measuring radiance and one measuring downwelling irradiance with a cosine collector The sensors measure over the wavelength range 350 950nm with sampling approximately every 3 3nm with spectral width of about 1Onm The sensors are based on the Carl Zeiss IESCA satellite validation protocols 07 027 FR ISECA 39 Monolithic Miniature Spectrometer MMS incorporating a 256 channel silicon photodiode array Integration time varies from 4ms to 8s and is automatically adjusted to measured light intensity The data stream from all three instruments is integrated by a IPS 104 power supply and interface unit and logged on a PC via a RS232 connection The radiance sensors have a field of view of 7 A two axis tilt sensor is incorporated inside the downwelling irradiance sensor The instruments are mounted on a steel frame similar in concept to that used by Hooker and Lazin 2000 The frame is fixed to the prow of the ship facing forwards to minimise ship shadow and reflection and 1 8m above
106. m microscopy cell counts Journal of Plankton Research 27 103 119 Mantoura R F C Wright S W Jeffrey S W Barlow R G Cummings D E 1997 in Jeffrey S W Mantoura RFC Wright SW eds Phytoplankton pigments in Oceanography guidelines to modern methods SCOR UNESCO Mueller J L Giulietta S Fargion and C R McClain Editors J L Mueller R R Bidigare C Trees W M Balch J Dore D T Drapeau D Karl L Van Heukelem and J Perl 2003 Ocean Optics Protocols For Satellite Ocean Color Sensor Validation Revision 5 Volume V Biogeochemical and Bio Optical Measurements and Data Analysis Protocols National Aeronautical and Space administration USA TM 2003 IESCA satellite validation protocols 07 027 FR ISECA 26 Roy S Llewellyn C A Egeland E S Johnsen G 2011 Phytoplankton Pigments Characterization Chemotaxonomy and Application in Oceanography Environmental Chemistry Cambridge UK IESCA satellite validation protocols 07 027 FR ISECA 27 Surface Downwelling Spectral Irradiance Es A W m nm Subsurface Downwelling Spectral Irradiance Ed z A W m nm Subsurface Upwelling Spectral Radiance Lu z A W m nm sr Surface Downwelling Diffuse Spectral Irradiance over Direct Spectral Irradiance r A Es A is normally measured at the nominal MERIS visible bands Lu z A measurements are taken to derive the subsurface upwelling radiance Lu 0 A Ed z XA measurements are taken in order to derive the diffu
107. m the attenuation The VSF 3 measures the volume scattering function B 0 at three angles 100 125 and 150 and three wavelengths 470 530 and 660 nm The processing of the data from the VSF meter was done in three steps Firstly conversion of digital counts into B 6 done using the calibration parameters supplied by the manufacturer Secondly a pathlength correction in turbid or very absorbing water c gt 5m 1 Given the geometry of the sensor and the characteristics of the water sampled the pathlength correction was neglected implying an error not greater than 5 of the measurement Finally calculation of bb from 8 at three angles This was done by fitting a third order polynomial through all the measurements points of 2mB sin 6 including 02 mn where f 8 sin m 0 Then the area under the polynomial was integrated using the Newton method To obtain bpp seawater backscatter bbw was subtracted to measured bp Only the upcast of each deployment was selected and all data were median filtered to eliminate salt and pepper noise and binned to 0 5m After a visual quality control and elimination of the individual profiles following manufacturer s guidelines data presented here correspond to a depth of 5m 2 1 2 Sampling at a transect Transect sampling took place on the Plymouth Quest between 5 7 Sept 2012 The objective was to collect radiometric data in a transect from offshore to the harbour as the vessel carried
108. matography 9 Print sheet and stick in lab book 10 Any modifications to this protocol must be approved and recorded 15 5 4 Protocol 4 Pigment extraction from filters Pigment extraction from filters 25 mm Notes Take internal standard extraction solution from freezer and allow to come to room temperature Protect from light and keep tightly sealed If insufficient solution to perform todays extractions prepare a new batch Transfer internal standard to four vials and place in autosampler Check system performance resolution of critical pairs in mixed standard and baseline noise amplitude before extracting samples Check pipette accuracy by performing three weighings of internal standard extraction solution 2 mL G13 room temperature prior to addition to samples Record on pigment analysis QC sheet Calculate average and actual volume and record Wear gloves when handling acetone Label centrifuge tubes 1 20 Dim lights in lab Take samples from freezer liquid nitrogen and fill in sample names on pigment analysis QC sheet Transfer filters 1 20 to corresponding centrifuge tube and place on ice Transfer cryovials to storage box Add 2 mLs internal standard extraction solution to each tube and tightly cap Keep samples on ice in dark Note time solvent added to filters on pigment analysis QC sheet Sonicate each solution for 35 sec using sonic probe amplit
109. me precision daily Inject extract from mixed pigment standard containing pigments to be quantified including critical pairs Inject at beginning of sequence before samples and use to calculate resolution of critical pairs If resolution starts to decrease take remedial action Record and monitor retention times of peaks in mixed standard see protocol 3 and spreadsheets 1 and 2 2 6 2 Injection precision monthly Perform repeat injections 6 of the same standard usually internal standard extraction solution of chlorophyll a working standard and calculate relative standard deviation of the peak area Also record the average RSD with time see spreadsheet 4 2 6 3 Method Uncertainty and Chl a Calibration Accuracy monthly and daily When a new chlorophyll working standard is prepared and quantified by spectrophotometry six injections are performed using the HPLC The calibration response factor is then used to determine the concentration of the standard from the HPLC peak areas and compared to the value from spectrophotometry to determine the uncertainty of the HPLC method see spreadsheet 5 In addition the response factor for chl a is determined from the HPLC peak areas and the concentration determined by spectrophotometry The response factor must be within 5 of the calibration value see spreadsheet 6 If not within 5 a new chlorophyll a working standard is prepared from a new stock solution If the value is still not within
110. mpty vial Draw 40 uL water deposit into vial mix in vial Inject 25uL Actual volume of sample injected 17 86 pL Injection procedure includes wash steps Calibration procedure Calibration performed for the following pigments Chl c3 Chl c Peri But fuco Fuco c Neo Pras Viola Hex fuco Diadino Diato Allo Zea Lut Gyro de Chl b DVChl a Chl a BB Car In Oct 2012 calibration curves were carried out for fourteen pigments and single point calibration carried out for five pigments For single point calibrations four injections of each standard were performed For calibration curves the standards were used to prepare a dilution series comprising three solutions bracketing the LOQ and three bracketing the expected sample concentration Source for standards DHI Absorption coefficients used as provided by DHI Chl a and internal standard from Sigma Water content in the filters Internal standard used to take into account volume of water in filters Estimated uncertainties of the Dec 2012 Average precision of method for Chl a was 1 44 and average accuracy was 2 0196 method Validation summary First run of the day is discarded A sample of mixed pigments is run prior to any samples to check retention times and resolution of critical pairs These data are recorded Three samples of chlorophyll working standard and of internal standard extraction solution are analysed with each sample set The response factor of the working standard is c
111. n situ measurements corrected for temperature salinity and scattering Methodology and processing description Temperature and salinity corrections After collection raw data must be corrected for the in situ temperature and salinity effects to correct for differences between the absorption coefficient of the optically pure water used as a reference when calibrating the instrument and the absorption coefficient of the water in which the measurements are performed These effects are removed by applying to the measured Cm and a A the following algorithms Cmts A Cm A w A T T eat Woe A S S cai 1 Amts A Aam A un w A T T eat Wsa A S S cai 2 Where T and S are the temperature and salinity of the water during measurement respectively and Tea and S 4 are the temperature and salinity in principle 0 of the water during calibration respectively The y and y coefficients used are the following WetLabs ac9 Protocol Document Revision Q April 2011 for c and a IESCA satellite validation protocols 07 027 FR ISECA 17 Scattering corrections of the absorption coefficient The portion of the scattered light not collected by the reflecting tube absorption meter causes the instrument to overestimate the absorption coefficient Presently three methods mainly are available in order to perform a correction of the measured absorption with methods 2 and 3 implying that c A be measured simultaneously with
112. ng to depth with the same colour scale as profiles b d and f where red is deeper blue is shallower 14 Product confidence flag based on spectral shapes could also be implemented for attenuation assuming that spectrally the attenuation follows a power law if the r of the fit to the power law was too low a spectra could be flagged in this category D PROCESSING OF DATA 1 Method of processing check L1 Watercal T amp S corr scatt corr Y N DIRECTORY FILE L2 Interpolation Y N DIRECTORY FILE 2 Processed data tests Spectral Plots Y N DIRECTORY FILE QC1 Automatic check for no negative Y N values QC2 0 discards spectra QC2 1 valid DIRECTORY FILE QC2 Automatic check for spectral shape Y N raise flag if a 510 or 532 or 555 or 650 or 676 or 715 a 412 or 440 or 488 QC2 0 discards spectra QC2 1 valid DIRECTORY FILE 3 2 In water backscattering 3 21 Instrumentation checks and calibrations Similarly to the in situ absorption and attenuation meter Section 3 1 1 a record of the checks and maintenance of the instrument should be kept Regular maintenance after or before deployment should be performed according to the manufacturer protocol 8 It is important to note whether the instrument has been cleaned or not in the QAD Another important aspect of the optical instruments maintenance is calibration It is possible to perform laboratory calibra
113. ns 1972 A practical Handbook of seawater Analysis 8 181 184 D Van der Linde Protocol for Total Suspended Matter estimate JRC Technical Note June 1998 IESCA satellite validation protocols 07 027 FR ISECA 35 Above water Water Leaving Radiance Lw Wm nm sr and Downwelling Irradiance Es W mnm Instrument Description The PR 650 640 is a hand held portable battery powered spectroradiometer manufactured by Photo Research The instrument measures radiance within a 1 aperture angle in 101 wavelength bands from 380 780nm in 4 nm steps Full width half mean is 8 nm The detector integration time is varied automatically to provide the necessary dynamic range Fig 9 Figure 9 PR650 instrument PR 650 640 SpectraColorimeter System A 1 field of view is used with the PR650 for measuring L A 0 9 the radiance emanating from the water surface and the sky radiance Lj 4 0 9 The downwelling irradiance is measured from a calibrated Lambertian reflectance panel Alternatively a cosine collector can be used with the PR640 to measure the incident spectral irradiance E A Photometric and colorimetric accuracy is assured by virtue of the fact that the PR 650 640 measures sources spectrally by diffracting the visible simultaneously over the 128 Regardless of the spectral distribution of the source be it a CRT or an incandescent lamp the correct luminance and color values displayed without special calibration The op
114. o processed data alternatively the output is in a simple ASCII format that may be processed by a spreadsheet Attenuation coupling Many scattering sensors require a subsequent attenuation correction for pathlength coupling of the transmitted and scattered light This is typically a function of the propagation distances of the light as well as the magnitude of the water attenuation Because the ECO VSF 3 incorporates very short pathlengths and scattering volumes in its measurements it is relatively immune to this pathlength coupling Figure 7 For attenuation coefficients up to approximately 5 m no data correction is required If you are operating the meter in waters with greater turbidity a different configuration is required Determination of primary angular coefficients The primary angular coefficients for each angle of backscattering can be applied upon raw data downloaded from the instruments Determination is made by subtracting the clean water offset from the measured value and multiplying the result by the scaling factors provided in the calibration sheet References Boss E Pegau W S 1997 Relationship of light scattering at an angle in the backward direction to the backscattering coefficient Applied Optics 40 30 5503 5507 2001 Maffione R A Dana D R 1997 Instruments and methods for measuring the backward scattering coefficient of ocean water Applied Optics 36 6057 6067 Mueller J L 2000 Instrument
115. ocessing check QC1 If B gt 0 0003 g Y N QC2 If 0 0001 B 0 0003 g Y N 27 3 6 Phytoplankton Pigments using HPLC The analysis of phytoplankton pigments using High Performance Liquid Cromatography is a complex laboratory based analytical technique 17 The protocols for analytical Quality Assurance and Quality Control will be summarised separately in an Annex section by Dr Ruth Airs REFERENCES 1 V Martinez Vicente and G Tilstone Data quality control guide for bio optical measurements EU REVAMP 2 IOC IODE Manual of quality control procedures for validation of oceanographic data M IOC IODE and CED DGXII UNESCO manual and guides Paris 3 P J Werdell and S J Bailey The SeaWiFS Bio Optical Archive and Storage System SeaBASS Current Architecture and Implementation G S Fargion and C McClain Greenbelt 4 P J Werdell and SJ Bailey An improved in situ bio optical data set for ocean color algorithm development and satellite data product validation Remote Sens Environ 98 122 140 2005 5 M Babin C Roesler and J J Cullen Real time Coastal Observing systems for marine ecosystem dynamics and harmful algal blooms Oceanographic Methodology Series ed M Babin C Roesler and J J Cullen2008 Paris UNESCO 6 C Brockmann Scope of water products the quality and science flags Proc MERIS User Workshop Frascati Italy 7 Wetlabs ac meter protocol Revision P
116. of critical pairs pairs of pigments that are challenging to separate from the chromatogram are determined and recorded A deterioration in resolution of more than one set of critical pairs results in remedial action eg changing precolumn The protocol for determining the resolution is described in protocol 3 The resolution data are recorded in the spreadsheet RS tracker Spreadsheet 1 Retention time data are recorded in the spreadsheet RT tracker spreadsheet 2 An example chromatogram of mixed pigments standard is shown in Figure 1 Figure 1 HPLC chromatogram of mixed pigments standard DHI 2 4 Sample Extraction Once the performance of the HPLC system has been verified extractions can commence Pigment filters are extracted in 90 acetone containing an internal standard trans B Apo 8 carotenal by sonication Extraction conditions are detailed in protocol 4 Important components for quality control are verifying the performance of volumetric pipette before use ensuring the internal standard extraction solution is allowed to warm up to room temperature before use and tightly sealing lids on extraction tubes to prevent evaporation of volatile solvent For pigment extraction from 25 mm GF Fs 2 mL extraction solvent is used For pigment extraction from 47 mm GF Fs 5 mL extraction solvent is used sufficient to cover the filter in extraction tube Twenty filters can be extracted and analysed within 24 hours 2 5 Description of Stan
117. on A peak area of internal standard injected directly onto column A peak area of internal standard in sample Vm extraction volume in pL V filter volume in L C ng of pigment on column V volume of sample extract injected in uL 2 8 Reporting For a pigment to be reported the retention time and spectrum need to match the pigment in question Where a pigment is not detected the effective LOD is reported Table These values are marked in red in the results spreadsheet The effective LOD is calculated from the LOD for the pigment and the typical filter volume extraction volume and injection volume used for the sample set A list of effective LODs is provided with the data as an ancillary file Values for precision from analysis of replicate filters are also reported including the average precision for TChl a and PPigs These values enable the data user to gauge the quality of the data according to Van Heukelem and Hooker 2011 Table 1 All abbreviations used in the results spreadsheet are as recommended in Phytoplankton Pigments 2012 Roy Llewellyn Egeland and Johnsen Eds Cambridge The percentage of unknowns in each sample is also reported This is equal to peak area unknowns total peak area all pigments x 100 The percentage of unknowns enables the data user to gauge how well the presented data represents the pigments present in the extract Average precision 96 Level of performance TChl a PPi
118. on of angular coefficients through direct measurement of suspensions of NIST traceable standard spherical beads which are serially diluted The dilutions are extrapolated to zero hence the VSF calibration does not include the angular scattering of pure water Methodology and processing description Deployment The ECO VSF 3 requires no pumps to assure successful operation Once power is supplied the unit is ready for submersion and subsequent measurements The sensor faces should not be pointed directly into the sun or other bright lights Precautions e When lowering the instrument ensure that the mounting brackets are not damaging the unit casing e Avoid obstructing the sensors optical paths The sensor will detect an object directly in front of its optics IESCA satellite validation protocols 07 027 FR ISECA 14 Upkeep and Maintenance After each cast or exposure of the instrument to natural water flush the instrument with clean fresh water paying careful attention to the sensor face Use soapy water to cut any grease or oil accumulation Gently wipe clean with a soft cloth The sensor face is composed of ABS plastic and optical epoxy and can easily be damaged or scratched Do not use acetone or other solvents to clean the sensor At the end of an experiment the instrument should be rinsed thoroughly air dried and stored in a cool dry place Data Processing ECO Host will convert raw data obtained during a deployment t
119. r RSD 496985 574802 574896 574123 575852 571123 562082 562503 110834 377708 108407 84183 373984 424843 4 49 4 63 4 58 4 42 4 44 4 52 4 33 4 49 0 11 247 2 04 1 43 1 45 1 91 1 61 0 67 1 33 1 79 1 63 2 50 1 81 0 96 1 17 1 73 5 28 5 08 5 08 5 07 4 93 5 21 5 12 5 08 0 09 1 73 27 09 2012 21 09 2012 18 09 2012 11 01 2013 25 04 2013 5 54 5 54 5 54 5 57 5 67 5 67 571 5 66 5 78 5 76 5 64 5 67 5 74 0 10 0 01 0 06 1 81 0 22 1 06 177 226 3 54 0 07 3 77 Avg CV AVG 1 57 0 08 1 57 1 22 6 6 Spreadsheet 6 Chl ws tracker 2013 Date Peak areas AVG peak area ng adj inj on column RF value change from calibration value Notes 10 08 2012 1 196E 05 Multipoint calibration See file LOD and working range4 zapata ftz 13 08 2012 570073 562503 6 59 1 172bE 05 2 04 572332 554015 553591 12 09 2012 473919 479610 5 54 1 155E 05 3 42 New stock and working std prepared DJS 483638 481272 21 09 2012 474447 473699 5 54 1 170E 05 2 21 472950 27 09 2012 465379 r 471427 5 54 1 175E 05 1 74 477475 02 10 2012 468004 473457 5 54 1 170E 05 2 16 478910 08 10 2012 493073 483986 5 54 1 145E 05 4 29 476834 482051 09 10 2012 476692 464735 5 54 1 192E 05 0 33 460102 457411 11 10 2010 458466 447933 5 34 1 192E 05 0 32 New working std prepared DJS 437960 447374 10 12 2012 441324 442016 5 17 1 17E 05 2 20 New working standard prepared DJS 439956 428560 450409 450967
120. rance Use an internal standard pigment standards are authenticated by VKI Quasimeme membership Sample Storage If filters are not analyzed immediately they should be flash frozen and stored in cryovials or petri dishes in liquid nitrogen Mantoura et al 1997 found that liquid nitrogen is the best form of sample preservation The storage of filters in ultra cold freezers 90 C also achieves excellent pigment recovery with minimum degradation Long term storage of samples in 20 C freezers is not recommended but can suffice for short term 1 wk storage Freeze drying causes rapid loss and extensive degradation of chlorophylls and carotenoids and is therefore not recommended Limitations The detection limit of this technique is about 0 001 ugT Divinyl chlorophyll a and b are distinguished using reverse phase C 8 HPLC and the methods described in Barlow et al 1997 References Barlow R G D G Cummings and S W Gibb 1997 Improved resolution of mono and divinyl chlorophylls a and b and zeaxanthin and lutein in phytoplankton extracts using reverse phase C 8 HPLC Mar Ecol Prog Ser 161 303 307 Jeffrey S W Mantoura RFC Wright SW 1997 Phytoplankton pigments in Oceanography guidelines to modern methods SCOR UNESCO Llewellyn C A J R Fishwick and J C Blackford 2005 Phytoplankton community assemblage in the English Channel a comparison using chlorophyll a derived from HPLC CHEMTAX and carbon derived fro
121. red for calculating MERIS reflectance but can be used to derive the fraction diffuse total downwelling irradiance which serves as input in numerical radiative transfer code such as Hydrolight Limitations Foam caused by waves Low sun heights can cause high contributions of sun glint The PR650 cannot be operated under rainy conditions because the instrument is not water proof References Fargion G S J L Mueller 2000 Ocean Optics Protocols For Satellite Ocean Color Sensor Validation Revision 2 NASA TM 2000 209966 Goddard Space Flight Space Center Greenbelt Maryland USA 184 p IESCA satellite validation protocols 07 027 FR ISECA 37 Mobley C D 1999 Estimation of the remote sensing reflectance from above surface measurements Appl Opt Vol 38 No 36 p 7442 7455 Mueller and Austin 1995 Volume 25 SeaWiFs Techn Rep Ser Chapter 6 2 IESCA satellite validation protocols 07 027 FR ISECA 38 Above water MERIS reflectance p A dimensionless SATLANTIC and TriOS methods The MERIS reflectance p A as defined by is calculated from simultaneous above water measurements of downwelling irradiance E A radiance from the water surface L 2 and sky radiance Ls A The latter two measurements are used to calculate the intermediate parameter L A the water leaving radiance after removal of air sea interface reflection This method corresponds to Method 1 of Mueller et al 2000 Resu
122. reflectance measurements Remote Sens Environ 135 202 212 2013 11 V Martinez Vicente et al Above water reflectance for the evaluation of adjacency effects in Earth observation data initial results and methods comparison for near coastal waters in the Western Channel UK Journal of European Optical Society Rapid Publications 2013 8 DOI 10 2971 jeos 2013 13060 10 11 INTERREG IVA 2 Mers Seas Zeeen Cross border Cooperation Programme 2007 2013 ISECA Protocols for the Validation of Ocean Colour Satellite data in Case 2 European Waters Contract 07 027 FR ISECA G H Tilstone and V Martinez Vicente 2012 Plymouth Marine Laboratory PML UK Based on NASA and ESA protocols IESCA satellite validation protocols 07 027 FR ISECA INTRODUCTION TO REVAMP PROTOCOLS 5 IB Stir iu PON TE E E EAT R 6 Introd ct oneei hera a e a a E bu beta ae aa 6 Instr mentdeseript on a e etd tesis ik rout isbe tolto ib utem a OON r ba e eaS O EAE 6 Analytical BEOGOQUEe viuis rete a folc bun rele toten bro aa e sit stars deos b fefe festo ona T Instrument Calibration and Quality Assurance eese 7 MCIBOdOIOS S 6x ser ed vf gssten ihr ponts tuedafttedtut a dv tst Sete fresca rir fetta e UR 7 Sample collection and filtration seco ice tes quss iate Get de vetare aot eheu 7 AMPLE storages MESH ET TRE EMEN 8 MeaSut nients pr ced re d ase tis retrato etat bu rrr pb ot tiet aa uini
123. res which have been undertaken in validating metocean data and the source documents to which reference can be made for details of these procedures In addition any significant comments relating to the procedures can be stated They therefore allow a rapid assessment to be made of the level to which data validation have been applied to a particular data set A QAD filled as necessary should be appended to each individual metocean data set upon the completion of the data validation by the data gatherer This QAD should then accompany this data set wherever it is transferred since it provides a definitive summary of the data validation applied to the data Any subsequent validation procedures which are applied can then be incorporated into the QAD and referenced An example of QAD is shown in Table 1 Table 1 Example of a QAD table for non directional wave data set IOC IOD 1993 NOTE Tick Box Y N only if specified action or check has been undertaken otherwise leave blank Initial responsibility for completing the QAD lies with the data gatherer although it is the responsibility of the project co ordinator or chief scientist to ensure that it has been filled in correctly Responsibility for incorporating any subsequent validation undertaken e g by a programme data manager lies with the analyst performing those validation procedures and these procedures must be adequately referenced Finally responsibility for completing section F of t
124. rs one used for sky radiance measurements SKY collimator the other used for both sky measurements and direct sun measurement SUN collimator IESCA satellite validation protocols 07 027 FR ISECA 31 Instrument Calibration and Quality Assurance The CE 318 calibration for radiance measurements is performed with an integrating sphere Inter calibration with a portable radiometer calibrated with the same integrating sphere is occasionally performed Independent calibration is also performed at 440 and 670 nm using the Rayleigh scattering calibration technique The CE 318 calibration for irradiance measurements is performed every two months using the Langley Bouguer method weather permitting applied to data from the measurement sites Inter calibration with the portable radiometer is occasionally performed for the direct sun irradiance measurements The Quality Assurance of CE 318 data is mostly addressed to remove contamination by cirrus following the methodology used in AERONET Holben et al 1998 Methodology and Processing Description Deployment of the instrument The sky measurements are made using two different procedures I the Almucantar procedure and ii the Principal Plane procedure The sun measurements are made using the Sun procedure During the Almucantar procedure the CE 318 points at the sun and than takes measurements with fixed sun zenith angle at different azimuth angles over 360 degrees During the Principal P
125. s with seawater and collect 200 ml of seawater e Blank preparation Filter 75 ml of MilliQ or bi distilled water into glass storage bottle and discard the filtrate Filter a further 75 ml of pure water for use as blank e Sample preparation Filter 75 ml of sample into clean bottles at a vacuum pressure of 120 mm Hg Shake bottles and discard water Repeat Filter at least 250 ml of seawater into glass bottles Cap the bottles and store in the dark e Sample Storage Samples can be stored for up to 4 hrs at room temperature before being analyzed Samples can be stored 4 to 24 hrs in a refrigerator Mitchell et al 2000 For longer storage 0 5 ml solution of 10g l of NaN3 per 100 ml of sample Ferrari et al 1996 can be added to prevent degradation of IESCA satellite validation protocols 07 027 FR ISECA 20 CDOM and sample bottles should be kept upright in a refrigerator 4 C However NaN3 adds to the absorption of the sample It is recommended that CDOM samples should be run fresh whenever possible If NaN3 is added for prolonged storage the sample should be flagged in the meta data base Measurement procedure e Ifsamples have been refrigerated allow them to warm up so that sample and blank are at the same temperature before scanning the samples Temperature differences between reference water and sample can lead to strong spectral absorption features Pegau amp Zaneveld 1993 Temperature of reference and sample should be recorded for
126. se attenuation co efficient Kd z A and the subsurface downwelling irradiance Ed 0 The ratio r A between the Surface Downwelling Diffuse Spectral Irradiance and the Direct Spectral Sun Irradiance is computed from Esky A Es A Esky A where Esky A is the Diffuse Sky Irradiance and Direct Sun Irradiance Attitude measurement of the Es A sensor is recommended when the instrument is installed on non stable platforms i e ships The attitude of the Ed z A and Lu z A sensors must be measured during profiles Sensor depth must also be determined with high accuracy Instrument description The measurement system consists of a compact seven channel analog sensor capable of 16 bit performance The analogue signals are digitized by a 16 bit a d unit DATA 100 Data is transferred by the DATA 100 as RS232 or RS422 The data acquisition rates are fully programmable but the normal data stream uses the default of 8 Hz sampling Physically the Es sensor is mounted on a pole clear of any shading structures The Ed and Lu sensors are mounted on a profiling rig designed to minimize any shading from close devices Figure 6 Satlantic sensor head for Es A measurements IESCA satellite validation protocols 07 027 FR ISECA 28 NO Y rs ON A ex cy r ee AN UN NN Y SAKA ON AN ANN S AV wee N N M Figure 7 Free falling profiler unit Satlantic Halifax Instrument Calibration and Qual
127. search 32 603 619 2010 3 G H Tilstone et al Variability in specific absorption properties and their use in a semi analytical ocean colour algorithm for MERIS in North Sea and Western English Channel Coastal Waters Remote Sens Environ 118 320 338 2012 4 C A Llewellyn J Fishwick and J C Blackford Phytoplankton community assemblage in the English Channel a comparison using chlorophyll a derived from HPLC CHEMTAX and carbon derived from microscopy cell counts Journal of Plankton Research 27 103 119 2005 5 S W Jeffrey et al Phytoplankton pigments in oceanography guidelines to modern methods1997 UNESCO Publishing 6 G H Tilstone et al Phytoplankton composition photosynthesis and primary production during different hydrographic conditions at the Northwest Iberian upwelling system Marine Ecology Progress Series 252 89 104 2003 7 Y Xie et al Effect of increases in temperature and nutrients on phytoplankton community structure and photosynthesis in the western English Channel Marine Ecology Progress Series in press 8 S Tassan and G M Ferrari An alternative approach to absorption measurements of aquatic particles retained on filters Limnol Oceanogr 40 1358 1368 1995 9 C D Mobley Estimation of the remote sensing reflectance from above surface measurements Applied Optics 38 7442 7455 1999 10 S G H Simis and J Olsson Unattended processing of shipborne hyperspectral
128. sk and foil covering 10 Store top shelf freezer G13 Shelf life one month Dispose of old stock solution in fume cupboard 11 Any modifications to this protocol must be approved and recorded 02 07 12 5 8 Protocol 8 Preparation of mixed pigments standard Preparation of mixed pigments standard Notes 1 Take a vial of mixed pigments standard supplied by DHI and the internal standard extraction solution from the freezer Allow to come to room temperature If it is a new vial of mixed pigments write the month and year first used 2 Using a gastight syringe transfer 400 uL of internal standard extraction solution to an amber vial Add 100 uL of mixed pigment standard and mix using the syringe 3 Label the sample according to the month the mixed pigments vial Maintain minimum light was first used and the number of dilutions eg mpd 06 13d mixed required for safe pigments diluted vial first used June 2013 fourth dilution from this working vial 4 Return stock standards to freezer 11 Any modifications to this protocol must be approved and recorded 18 5 9 Protocol 9 Quantification of chlorophyll a working standard solution by spectrophotometry Quantification of chlorophyll a working standard solution by spectrophotometry Notes 1 Switch on spectrophotometer Ensure correct cuvette holders Leave spec to warm up fitted for 45 min before
129. sky viewing direction and high wind Such uncertainties are relatively more important for clearer waters e Measurement uncertainties increase for underway measurements because of increased tilt roll and possible contamination of lenses by spray e Underway measurements from small ships e g Rigid Inflatable Boats are limited to calm sea state e g Bf lt 3 to avoid excessive tilt and roll IESCA satellite validation protocols 07 027 FR ISECA 41 References Balch W M D T Drapeau B C Bowler E Lyczskowski E S Booth and D Alley 2011 The contribution of coccolithophores to the optical and inorganic carbon budgets during the Southern Ocean Gas Exchange Experiment New evidence in support of the Great Calcite Belt hypothesis J Geophys Res 116 COOF06 doi 10 1029 2011JC006941 Hooker S B and G Lazin 2000 The SeaBOARR 99 Field Campaign Greenbelt Maryland NASA 46 Mobley C D 1999 Estimation of the remote sensing reflectance from above surface measurements Applied Optics 38 7442 7455 Mueller J L C Davis R Arnone R Frouin K Carder Z P Lee R G Steward S Hooker C D Mobley and S McLean 2000 Above water radiance and remote sensing reflectance measurements and analysis protocols Ocean Optics protocols for satellite ocean color sensor validation Revision 2 Greenbelt Maryland National Aeronautical and Space Administration 98 107 Ruddick K V De Cauwer Y Park G Becu
130. sponse wil fluence the produciviy your laboratory Whether it is a engineers and applications specialists will J ee AD m Saves you precious time and improves instrument uptime asa efficient E ation Thermo Scientific Plans offer for ee bicagir ly service guaranteed aceon A an e RR n EE The Critical Essential includes specified consumable items in accordance with our standard instrument lar maintenance will extend the life of your instrument improve the quality of your results lower your pem rem aere ph annual maintenance costs To accept this quotation and continue support ign and return the official acceptance form ther with a order marked for ire anenton of fhe undersigned Upon receipt we wil issue an voice and that Support Plan has been set up If we can be of any further assistance please do not hesitate to contact us Yours sincerely Matthew Wolfenden 0870 4100888 contracts cmd_hemek thermofisher com Thermo Fisher Scientific is the trading name of Thermo Electron Manufacturing Lid Page 1 of 4 29 Boundary SUPPORT PLAN QUOTATION a HP 7GE QUOTATION Number 20285670 40090654 Tel Q1429 233555 Coverage Start Date Drill 01 2013 Fax 01442 233667 Coverage End Date March 31 2014 Quotation Expires October 18 2012 Dr Ruth Airs Quote Created On 20 2012 Plymouth Marine Laboratory Prospect Place Poole PL1 3DH Phone Fax
131. sseeressresesrresrreresrsreesereees 40 Methodology and Processing Description eese enne 40 Deployment of the Wiis CUDITIE uoa i roro ptas ERI RE Pt edat bauen edid ete EE uds ine 40 Description of processing techniques employed sese 40 Preprocessing Quality Checks oae eap fesce rone EC ESRN UR IU SH EPHIS MOR REP ERES EOD 41 Pata reso IN PA M E 41 Postprocessine Quality Checks marire eter tpi e a p eE E E EES 41 Limitation PLU a A p E E E A E R AE ES 41 igicur io t EE ar a E E EEE E ETE E ERR 42 SIMBADA method 4 teenaa aina a iea iaaa tutus 43 Instrument descripti i siennes asee i e ape EaR AEE a RERS 43 Instrument Calibration and Quality Assurance seseeeseeeeeseseessresessrrsesrrrseseresrsseesresees 44 Methodology and Processing Description s seeesssssresessesrrsresresresresrrsrerresresresresresreeees 44 Deployment of the instrument eseesseeeessesesesesesertsrssresrrstesresresresresesresresrerseseessesees 44 Description of processing techniques and quality checks sess 44 Eimitation S EIDEM MR CMM E 44 jars PERMET a a Ea oaae aa 44 TESCA satellite validation protocols 07 027 FR ISECA 4 INTRODUCTION TO ISECA PROTOCOLS In Case 1 waters Chlorophyll a Chla determined from Ocean Color is closely related to the absorption of light by phytoplankton pigments Algorithms based on blue blue green reflectance ratios are reliable for the derivation
132. t documentation automatic quality control oceanographic assessment can be divided between a the originator of data to improve the data consistency within the data set b the data manager to improve the data consistency within a data bank or in a multisource data set Crossover of the tasks can occur if the bio optical data have been collected for different areas by the same originator then the originator has to ensure consistency among different areas that may have different optical properties Hence the data originator has to follow sections a d This report will focus on sections a to c 2 1 Data quality control measures for data providers Regarding the data quality control measures the originator is responsible for the following use of documented or international recommended standard measurement methods and equipment national and international calibration of measurement methods and instruments data validation according to results of calibration and intercalibration as well as in comparison with standard methods information on temporal and spatial sampling tests of fixed and computed limits gaps and constant values detection correction and flagging of spikes detection correction and flagging of errors in position and time documentation of the process of data sampling and validation including any algorithm applied documentation of QC checks carried out and their results in QAD W
133. t can plot or store calibrated data from an instrument or raw data file Calibration The weighting function can be measured by moving a spectralon reflective target through the working volume Maffione and Dana 1997 IESCA satellite validation protocols 07 027 FR ISECA 13 The WETLABS instrument Instrument description The ECO VSF 3 measures the optical scattering at three distinct angles 100 125 and 150 degrees at three wavelengths thus providing the shape of the Volume Scattering Function VSF throughout its angular domain the three angle measurement allows determination of specific angles of backscattering through interpolation Conversely it also can provide the total backscattering coefficient by integration and extrapolation from 90 to 180 degrees using a 3 order polynomial according to the VSF manual Figure 3 The ECO VSF 3 backscattering meter The optics include three sets of three LED based transmitters that couple to three receivers The transmitters and receiver are located to establish centroid light scattering angles of approximately 100 125 and 150 degrees respectively For each angle the region of intersection encompasses a full width half maximum FWHM bandwidth of about 18 degrees Each sensor head operates at one wavelength Presently there are three wavelengths available 450 nm 530 nm and 650 nm Instrument Calibration and quality assurance Calibration of the ECO VSF involves the determinati
134. that L Lsky and E are measured with the same instrument Methodology and Processing Description The PR650 from an altitude of 2 4 m above the sea is pointed towards the sea surface 135 degrees azimuth away from the sun with a viewing angle of 35 40 degrees The downwelling irradiance is measured from a calibrated Lambertian reflectance panel or Es is simultaneously measured with a PR640 looking straight upwards cosine At each station reflectance is measured at least three times as quick as possible to reduce effects of changing water masses and illumination conditions Preferable position on the ship is on the bow to minimize surface wave effects and shading and or reflectance from the ship s superstructure In general each reflectance measurement consists of four radiance measurements l radiance emanating from the water surface L 2 radiance from the sky Ly 3 radiance from the reflectance standard L or simultaneously with a PR640 4 optionally radiance from the shaded reflectance standard L Each radiance measurement is an average of five readings internally averaged by the radiometer The sky radiance is measured to correct the total surface radiance for sky radiance reflected at the sea surface to yield water leaving radiance L L p L where p is the effective Fresnel reflection coefficient for the wind roughened sea surface Fargion and Mueller 2000 The measurement of the shaded reflectance panel is not requi
135. the water surface Where necessary to avoid optical interference the downwelling irradiance sensor is mounted separately elsewhere on the ship Instrument Calibration and Quality Assurance The instruments are calibrated twice per year at NIST traceable facilities in the framework of MERIS Validation Team workshops Methodology and Processing Description Deployment of the instrument The instruments are mounted on a steel frame which can be fixed to the bow of the ship The sensors should face forward to minimize ship shadow and reflection Before measurements the frame is levelled horizontally and the sea and sky viewing angles are fixed at 40 with respect to zenith and viewing in the same azimuth angle In this way the sky is viewed in the direction from which light will enter the sea viewing sensor after reflection at a flat sea surface The radiance sensor lenses and the irradiance sensor collector are inspected manually before each measurement and are cleaned of spray and dust when necessary The ship is manoeuvred on station to point the radiance sensors at a relative azimuth angle of 135 with respect to sun New platforms have been developed which automatically track the position of the sun so that continuous quality assured measurements can be taken whilst the ship is steaming Balch et al 2011 When the correct position and angle are achieved measurements are started and continue for 10 minutes taking a scan of the three instruments
136. tions to track the instrument performance with time The methods recommended by the manufacturer are detailed in the instrument manuals and will not be repeated here 7 Several additional quality control measures during calibration and sources of uncertainty are summarized in recent publications 9 11 It is important that the operator is able to perform the calibration with a known repeatability tested by repeating by triplicate the whole procedure including cleaning of the instrument and the setup The slope or scaling factor obtained from calibrations should have an RMSE 1 not more than 2 9 In addition drift in regular calibrations should be monitored and noted There are two spectral parameters to be monitored for stability the dark counts and the scaling factor Monitoring 15 of the dark counts has shown drifts of 1 counts over 1 month and 2 counts over 8 years The scaling factors change at different rates with time for each channel e Blue 896 y e Green 1 296 y e Red 3 496 y An example of the scaling factor S in sr m counts drift is shown in Figure 4 In this example the calculated change per year was 4 5 and 2396y for the blue green and red channels respectively The greater increase on the slope was observed after the second year The date of the last checked value of dark count and S should be recorded in the QAD 1 E 05 gt es 1 E 05 gt m ig 1 E 05 HB gs E F Blue S 8 E 0
137. tive chloride solution may cause excessive bleaching of the detrital fraction which would result in higher phytoplankton absorption coefficients If a 1 solution is used the NaCIO can be applied to the filter as 4 to 5 drops as described in Tassan and Ferrari 1995 and ensure that the NaClO spreads over the whole of the filtration area If 0 1 active chlorine NaCIO is used the sample filter should be placed on the filtration port and stood in 5 ml of NaClO for up to 15 mins Disappearance of the peak at 675 nm in the bleached sample and evidence of a concave shape of the OD spectrum near to 440 nm can be considered evidence of complete filter bleaching Mitchell et al 2000 For both 0 1 amp 1 active chlorine treatment 5 ml of MilliQ should be re filtered through the treated GFF filter to remove any residual NaClO Tassan et al 2000 Blank filters should also be bleached and re filtered using the same procedure e Ensure that both sample and blank filters do not dry out Dry filters adversely affect the optical density of the sample IESCA satellite validation protocols 07 027 FR ISECA 8 Data processing In Case I waters a zero offset from the baseline may occur which is presumed to be the product of scattering throughout spectrum Hence a spectral region is identified where phytoplankton absorption is assumed to be negligible typically 750 to 800nm and the scattering observed is due to non phytoplankton material However in Case 2 wat
138. ude 4 Dip sonic probe in beaker of acetone and wipe dry in between samples Recap each solution after sonication and keep on ice in dark Leave for 30 mins total soak time 1 hr Wear ear protection and glasses when using sonic probe 10 Filter each extract through syringe filter 0 2 um PTFE 17mm into prelabelled amber sample vial Samples can be centrifuged first if necessary Cap immediately and transfer to autosampler 11 Edit sequence to include sample information 12 Stick Pigment analysis QC sheet into lab book Print sequence and stick into lab book 13 Any modifications to this protocol must be approved and recorded 16 5 5 Protocol 5 Preparation of stock solution of internal standard and internal standard extraction solution Preparation of stock solution of internal standard and internal standard extraction solution Notes 1 Stock solution of internal standard Trans B Apo 8 carotenal Wear gloves when Sigma 10810G 1G prepared by dissolving 0 01 g of trans B Apo 8 handling acetone carotenal in 100 mL of 90 acetone The stock solution is stored ina Prepare and use solution foil covered sealed duran flask at 20 C under dim light 2 The internal standard extraction solution is prepared by adding 100 Prepare and use solution uL of stock solution to 250 mL of 90 acetone The internal under dim light standard extraction solution is stored in
139. udies 1 3 The area of study is off the Plymouth coast UK Figure 1 Figure 1 A Map showing the position of the L4 and E1 stations as well as the opportunistic above water transects dark blue line B Electronic in situ optical measurements deployment cage C Above water reflectance measurements in transect 2 1 METHODS 2 1 1 Sampling of a time series at L4 and E1 In situ sampling was undertaken on board RV Plymouth Quest weekly at station L4 approximately monthly at E1 and comprised vertical profiles of hydrographic biological and optical parameters A summary of the samples collected since 1989 and with a focus on 2008 2012 years is given in Box 1 Water for laboratory analysis was collected near surface in 10 L carboys and returned to the laboratory in a cool box Samples for coloured dissolved organic matter CDOM determination were kept in 0 5 L dark glass bottles also transported in the cool box Hydrography Vertical temperature salinity and fluorescence profiles were measured with a SeaBird SBE19 CTD coupled with a Chelsea Technology MINITracka fluorometer Phytoplankton pigments Phytoplankton pigments have been measured using High Performance Liquid Chromatography HPLC systematically at the surface at L4 since 2000 Since 2007 at L4 pigments have been also collected at depth 0 10 25 and 50 m At E1 pigments have been analysed at O 10 20 30 40 and 60m since 2002 On board approximately 1 2 L of sea
140. uency of 1 duplicate per 20 samples This describes the overall method precision from filtering to analysis Average precisions for TChl a and PPig are reported with results see Reporting section 2 6 7 Chl a Linearity Annually or if column method is changed Five or more chlorophyll standards within the working range are prepared in 9096 acetone using gas tight calibrated glass syringes and class A volumetric glassware Prior to analysis the mixed standard is injected to evaluate resolution and retention performance One injection is performed per standard and the data are used for a linear regression The y intercept must be near zero and well below the lowest point of the working range The slope should be within 3 296 of average from previous determinations Over time the normalised response factor can be plotted as a function of amount of Chl a injected to compute warning and control limits 2 7 Data Processing Data processing is semi automated Assignments and integration in each chromatogram are checked manually The total peak area of unknowns is recorded and expressed as a percentage of the total peak area of detected peaks at 440 nm Once the data have been exported the peak areas are multiplied by the response factors for the individual pigments to determine the ng of pigment on column The concentration of individual pigments are then calculated as follows Cp AA X Vm V X C V lt Where C pigment concentrati
141. ulated following Tilstone et al 2003 6 and normalized to chl a and the curves were then fitted using the equation given by Platt et al 1980 PB PBs 1 exp a PBs exp bI PBs 1 where a is the light limited slope b is the parameter representing the reduction by photoinhibition and the maximal light photosynthetic rate PBm is calculated as follows PBm PBs a a b b a b b a 2 Full details of this method and analysis of the results have been recently published in Xie in press 7 Particulate phytoplankton detrital and coloured dissolved organic matter absorption CDOM coefficients Measurements of absorption coefficients have been made of L4 surface water since 2001 The absorption coefficients of total particulate and detrital material retained on 25 mm GF F filters were measured before and after pigment extraction using NaClO 1 active chloride from 350 to 750 nm at a 1 nm bandwidth using a dual beam Perkin Elmer Lambda 2 spectrophotometer retro fitted with an integrating sphere Concerning CDOM replicate seawater samples were filtered through 47 mm diameter 0 2 um Whatman Anopore membrane filters using pre ashed glassware The first two 0 25 L of the filtered seawater were discarded The absorption properties of the third sample were determined immediately on the spectrophotometer and a 10 cm quartz cuvette from 350 to 750 nm relative to a bi distilled MilliQ reference blank Spectral CDOM absorption aCDOM A w
142. used are given by VKI International Agency for C determination VKI Water Quality Institute Agern All 11 DK 2970 H rsholm Denmark Pigments standard concentrations Cp are calculated as follows Cp A1 A750 E1cm 1 10 Cp pigment concentration of standard ug I A3 absorbance at wavelength nm Table I Ajs0 absorbance at 750 nm to correct for light scattering Eien extinction coefficient Ejem 1 g cm Table I l cuvette pathlength cm 10 conversion factor g to ug A recalibration of the HPLC with pigments standard is recommended every 3 4 months The recalibration with respect to internal standard should be performed every day Methodology and Processing Description Methodology of Sample Processing Sampling collection and storage For each seawater sample 1 5 to 2 liters are immediately filtered after collection through a Niskin bottle or other using 25 mm GF F filter The filter is then folded in half twice and placed into a labeled cryovial and stored in liquid nitrogen until laboratory analysis TESCA satellite validation protocols 07 027 FR ISECA 24 Pigment extraction and sample preparation For pigment extraction 2 ml of 90 acetone is added to the filter which is ultrasonicated using an ultrasonic probe for 20 secs as described in Llewellyn et al 2005 The extracting solvent also has an internal standard typically Apo 8 Carotenal trans The concentration of internal standar
143. utomatic check for for apny only Y N spectral shape raises a flag if Aphyl443 Apny 665 lt 1 DIRECTORY FILE QCA4 for aga only spectral shape raises Y N DIRECTORY FILE 3 4 Laboratory CDOM Similar to the measurements presented in Section 3 3 the CDOM coloured dissolved organic matter absorption measurement is done on a bench based instrument in the 23 laboratory with pre concentrated samples The specific method that will be addressed here are those described in the ISECA protocols 12 In practice the analytical method comprises the direct measurement of the absorption coefficient of the dissolved fraction CDOM or ays The dissolved fraction is defined operationally by the substances that pass through a 0 2um pore size filter 3 4 1 Instrumentation checks and calibrations Similar procedures should be used as in Section 3 3 1 It is worth noting that the nominal precision of the method for baseline control is 0 0005 A or ten times higher than for the particulate absorption method A INSTRUMENT METHOD CHECK 1 Sensor output check Before measurement Y N After measurement Y N Instrument checked Y N 2 Calibration Date of factory calibration dd mm yyyy Last laboratory calibration date dd mm yyyy 3 Measurement check Baseline 0 0005 Y N 3 4 2 Documentation of deployment parameters and sample preparation Similarly to Section 3 3 2 water samples n
144. water was filtered onto a GF F and stored in liquid nitrogen until analysis Pigments were extracted into 2 mL methanol containing an internal standard apo carotenoate Sigma Aldrich Company Ltd using an ultrasonic probe 30 S 50 W following the standard PML methods 4 Pigments were identified using retention time and spectral match using PDA 5 and pigment concentrations calculated using response factors generated from calibration using a suite of pigment standards DHIWater and Environment Denmark Phytoplankton primary production Phytoplankton photosynthetic parameters were calculated from photosynthesis irradiance P E curves measured using linear photosynthetrons illuminated with 50 W tungsten halogen lamps following the methods described by Tilstone et al 2003 6 For each depth 15 aliquots of 70 ml seawater within polycarbonate bottles Nalgene were inoculated with 5 to 10 uCi of 14C labelled bicarbonate Incubations were maintained at in situ temperature for a 1 5 h period after which the samples were filtered onto GF F under a vacuum pressure no greater than 27 kPa The filters were then exposed to 37 fuming hydrochloric acid for 12 h and immersed in 4 ml scintillation cocktail for 24 h and beta activity was counted on a TriCarb 2910 scintillation counter PerkinElmer Correction for quenching was performed using the external standard and the channel ratio methods Total inorganic carbon fixation within each sample was calc

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