WATERS AquaAnalysis System. Online analiza voda LC/MS-MS

Transcription

1 WATERS AquaAnalysis System Online analiza voda LC/MS-MS

2 Kompletno resenje A B C priprema uzoraka (SPE) Hromatografija (LC) Masena spektrometrija (MS or MS/MS)

3 Pocetak? Priprema Masena uzoraka Hromatografija Spektrometrija Ne pripremljen uzorak : Polarnost: - Vd Voda za pice Silica- C 18, C 8, C 4, C 2 - Povrsinske vode Hybrid- C 18, C 8, C 4, C 2 - Otpadne vode Polymer- C 18, C 8, C 4, C 2 - Sedimenti Embedded polar group Cyano, Phenyl Velicina cestice: 1.8, 2.5, 3.5, 5 or 8 µm Extracted sample For LC/MS/MS Unutrasnji dijametar kolone : 4.6, 3.9, 2.1, 1.,.32 mm and 75 m Ideally, the final sample should be as clean as a non-extracted Duzina kolone: standard. 15,, 5, 3, 2 mm Izvor: ESI APcI Nano-ESI Maseni analizator: magnetic sectors electric sectors time of flight quadrupole ion trap FT-ICR

4 Moguca resenja???? ACQUITY UPLC 1525µ 2777 Sample Manager Acquity SQD ZQ single quadrupole LCT Premier ToF Quattro Premier XE

5 Hemija kolona?

6 SPE?

7 Cetri osnovna pitanja Kompleksnost matriksa tik Drinking water Ground water Waste Water Raspolozivost zapremine 1 ul 5 ul ul 5 ul 1 ml 5 ml 1 ml 5 ml ml 5 ml 1 L Pre-clinical clinical environment LOQ 1 pg/ml 1 pg/ml pg/ml 1 ng/ml 1 ng/ml ng/ml 1 ug/ml 1 ug/ml ug/ml 1 mg/ml1 mg/ml # uzoraka MRM SIR SCAN / UV Syringe / Barrels 96-well plate On-line column / cartridge Integrated platform

8 Unselective retention/elution Selective elution Selection retention/elution Extraction Methods in Organic Analysis A.J. Handley 1998

9 Off line SPE protocol Volume Availability Condition/Equilibrate 1 ul 5 ul ul 5 ul 1 ml 5 ml 1 ml 5 ml ml 5 ml 1 L Load wash 1 x x x Elute Evaporate and Reconstitute Final extract volume 1mL inject 3 5 µl Target LOQ x x 1 x Method LOQ 1 pg/ml 1 pg/ml pg/ml 1 ng/ml 1 ng/ml ng/ml 1 ug/ml 1 ug/ml ug/ml 1 mg/ml 1 mg/ml Instrument LOQ

10 On line/ Off line Sample Mass Preparation Chromatography Spectrometry Off-line SPE ON-line SPE

11 ON Line SPE Format ON line analiza je tehnikakoja redukuje kolicinu manualnog rada sve do analizabilnog uzorka za to LC/MS/MS. Typical ON line SPE Method Kondicioniranje/ekvilibracija Nanosenje uzorka pranje 1, 2, 3.. N Direktno ispiranje u HPLC 4.6 x x x 2 Analiza Extraction protocol fully automated

12 ON line SPE Volume Equivalency Volume Availability 1 ul 5 ul ul 5 ul 1 ml 5 ml 1 ml 5 ml ml 5 ml 1 L Condition/Equilibrate Load wash Elute Final extract volume is (ml, L, nl?) Analytes are trapped on Extraction Column Direct elution to HPLC Column Off Line SPE Extraction 5 ml (starting volume) 1 ml final (elution volume) 5:1 enrichment LC/MS Analysis 3-5 µl (injection volume) 3 µl 1/33 th of original volume 15.1 ml 5 µl 1/2 th of original volume 25. ml

13 OFF-line Vs ON-line Sample Volume ml Container 2 ml vial Autosampler rack

14 Waters AquaAnalysis System (2-1 configuration) Parallel N SPE (2-1) configuration Flush 515 Rec 515A Rec B Flush: 5/5 Acetonitrile/Acetone Rec A: 2/8 Acetone/Ethyl acetate Rec B: 2/8 Acetone/Hexane extraction column A Loading Pump waste Line A: H 2 O + Additive Line B: MeOH Elution Pump Line A: H2O + Additive Line B: 7/3 ACN/MeOH +Additive extraction column B C 18 Column Mass Spectrometer.

15 Waters AquaAnalysis (2-1 configuration) Autosampler Isocratic pumps MS/MS Binary pumps

16 Waters AquaAnalysis System (2-1 configuration) Sequential and Parallel Events loading Acquisition Start Elution Re-equilibration 2min per 5mL SPE wash 5 95 Extraction Column A Short Gradient 2.1 x 5 mm C Flush Recon Flush Re-equilibration Extraction Column B

17 AquaAnalysis System (2-1 configuration) Sequential and Parallel Events loading Acquisition Start Elution Re-equilibration 2min per 5mL SPE wash 5 95 Extraction Column A Long Gradient 2.1 x mm C Flush Recon Flush Re-equilibration 2 nd Injection loading Extraction Column B

18 Triazines Mix 1 ml Loading Volume R N 1,4 R N N R MRM of 9 Channels ES+ TIC 1.88e6 Triazine Mix at ppt 1- Deisopropylatrazine 2- Simazine - * 3- Simetryne 4- Atrazine 5- Prometon 6- Ametryn 7- Propazine 8- Cyanazine 9- Terbutryn

19 Simazine Loading Volume ml 1 ppt Area: ml 1 ppt Area: ml ppt Area: ml ppt Area: mL ppt Area: Average of 1 injections - CV > 1

20 Loading Conditions Low, Neutral and High ph Simazine Cyanazine Propazine Average of 1 injections CV > 1 Area: 439 Area: 582 Area: ph H3 2 Formic Acid 4 ml/min 5 ml at ppt Area: 367 Area: 721 Area: ph 7 No Addition 4 ml/min 5 ml at ppt Area: 355 Area: 591 Area: ph 1 2 NH 4 OH 4 ml/min 5 ml at ppt

21 Eluting Conditions Low and High ph.5 Ammonium Hydroxide MRM of 1 Channels ES+ TIC 9.22e3 High ph load MRM of 1 Channels ES TIC 5.83e5 3,8 1,4, mm Ammonium Formate ph MRM of 1 Channels ES TIC e , Formic acid

22 Eluting Conditions Low and High ph Breakthrough? Partial ilelution? Partial loading? 1.52 Area: (88 reduction) MRM of 1 Channels ES > e3 Area.5 Ammonium Hydroxide MRM of 1 Channels ES > e5 Area Area: (2 reduction) Prometon 5mL at ppt 5 mm Ammonium Formate ph MRM of 1 Channels ES > e5 Area Area: Formic acid

23 Infusion Triazines Mix at 1 ppm : Low and High ph.5 Ammonium Hydroxide Scan ES+ 5.28e9 5 (-88) 9 (-98) 5 mm Ammonium Formate ph (-74) Formic acid (-75) (-55) (-2) 4 (-73) (-51) 7 (-73) (-7) (-49) 9 Scan ES+ 5.28e9 Scan ES+ 5.28e9 m/z

24 Organophosphorous 15 ml Water Sample 1 ppt 6.93 Dicrotophos 1 ppt S:N 139:1 MRM of 3 Channels ES > e4 High ph load Low ph elution Malation 1 ppt S:N 273:1 MRM of 3 Channels ES > e MRM of 3 Channels ES > e3 Ethion 1 ppt S:N 8:

25 Carbamates 15 ml Water Sample 1 ppt Thiobencarb 3: MRM of 1 Channel ES+ TIC 5.67e : MRM of 4 Channels ES Thiodicarb TIC 4.54e4 9.9 Propoxur Aldicarb 8.74 Carbaryl : MRM of 2 Channels ES TIC 2.37e5 Propamocarb 6.76 Pirimicarb High ph loading Low ph elution

26 Robustness 1 ml volume at ppt Injection # TIC 1.8e TIC 2.95e TIC 4.59e Injection # 5 C.V. A: 3.5 C.V. Rt: 1.1 Propamocarb TIC 1.4e7 Propoxur TIC 2.65e6 CVA:93 C.V. 9.3 C.V. Rt: TIC 4.84e5 Thiobencarb C.V. A: 3.9 C.V. Rt:

27 Extraction Column Lifetime Organo-nitrogen 15 ml water at ppt 4.35 MRM of 3 Channels ES+ TIC 3.3e5 Siduron 4.3 Tebuthioron Linuron

28 Column Lifetime Organo-nitrogen 1 th injection 5 th injection 4.39 Linuron 4.39 MRM of 3 Channels ES > e Siduron MRM of 3 Channels ES > e Tebuthioron MRM of 3 Channels ES > e5 Loading Pressure 45 psi Loading Pressure 47 psi

29 Zakljucak - Pt Potpuno automatizovan ti sistem it - Limiti detekcije ng/ml LOQ s - Minimalizovana i priprema - Redukovan manualni rad - Kompatibilnost SPE i hromatografskih kolona Reproduktivnost i robusnost.