Sample Preparation Strategies for Water Analysis - AmChamamcham.dk/dl/events/ESAC/Presentation3.pdf · ©2007 Waters Corporation 3 For high sensitivity analyses, such as those employing

©2007 Waters Corporation

Sample Preparation Strategies Sample Preparation Strategies for Water Analysisfor Water Analysis

Hannah WhiteHannah White

Waters Business Development ManagerWaters Business Development Manager

©2007 Waters Corporation 2

OutlineOutline

Introduction— Why sample Prep— Considerations

o Choices of tools— Why SPE

o Pre-Treatment

Strategies— Traditional approaches— Modern approaches

o Mixed Modeo Reverse phase

SummaryAppendix


For high sensitivity analyses, such as those employing LC/MS/MS, proper sample preparationcan be critical for minimizing matrix effects and concentrating analytes of interest.

Why Sample Prep?Why Sample Prep?

60% of the work activity and operating cost is spent on sample preparation for introduction into the analytical system

Three Purposes: – Removes interferences from sample matrix– Concentrating analytes of interest– Improving analytical system performances


Sample Preparation TechniquesSample Preparation Techniques

Sample Preparation- The simplification of sample matrix and enrichment of target analyte(s)

Types of Sample Prep include:— Dilution

— Centrifugation

— Filtration

— Liquid/Liquid Extraction

— Solid Phase Extraction


Some ConsiderationsSome Considerations

Solid samples — usually start with organic or aqueous extract of tissue or soil— initial extract is adjusted for optimal SPE enrichment and/or cleanup

o pH adjustmento solvent adjustment

• acetone/acetonitrile/IPA – suitable for aqueous dilution, Reversed-Phase and Mixed-Mode SPE

• ethyl acetate/DCM/MTBE – can be exchanged to hexane for normal-phase SPE

Aqueous samples (water, beverage, plasma/urine)— pretreatment may be appropriate

o pH adjustmento filtration/centrifugationo protein precipitation

— Usually, aqueous samples can be analyzed using Oasis® Reversed-Phase or Mixed-Mode SPE


PrePre--Treatment Prior to SPETreatment Prior to SPE

Pre treatment:

Solid samples (soil, tissue, etc.)— shake, sonicate or soxhlet

o extract with polar organic solvent (methanol, acetonitrile); polars

o extract with organic solvent + drying agent (DCM, acetone); non-polars, multi-residue

Non aqueous Liquido if water soluble, dilute with water for reversed-phase (or mixed-

mode) SPE

o if hexane soluble, dilute with or exchange to hexane for NP-SPE

Wastewater— filter or centrifuge as necessary

o filtered solids and filter may require analysis as solids


Why Solid Phase ExtractionWhy Solid Phase Extraction

Isolation of the analyte(s) of interest from the matrix

Sample Cleanup — removal of matrix interference

— Increased sensitivity

o Increased system uptime

o Longer column lifetime

Enrichment of analyte(s) of interest o Increased sensitivity

Exchange to LC or GC compatible solvent

SPE is also faster and more suitable for automation compared with liquid-liquid extraction


Short List of Sorbent Types Short List of Sorbent Types for SPEfor SPE

Normal-Phase Sorbents (polar sorbents)— Silica, Alumina, Florisil®, Aminopropyl silica, Diol silica, GCB

Reversed-Phase Sorbents (non-polar sorbents)— Oasis® HLB — C18, C8 etc (alkyl silica's)— Carbon based sorbents

Ion Exchange— Accell Plus™ CM, QMA

Mixed Mode (ion-exchange/reversed phase) — Oasis® MAX, Oasis WAX (strong and weak anion-exchange)— Oasis® MCX, Oasis WCX (strong and weak cation-exchange)


OutlineOutline



o Pre-Treatment



SummaryAppendix


SPE StrategiesSPE Strategies

1. Approach #1Retention, cleanup, elution

2. Approach #2Pass-through

3. Approach #3Dispersion


1. load 2. wash 3. elute

1. Sample is loaded onto SPE sorbent

• Analyte(s) of interest are retained on sorbent

2. Matrix interferences are washed off sorbent

3. Analytes are eluted from sorbent

SPE Strategy 1SPE Strategy 1RetentionRetention--CleanupCleanup--ElutionElution


SPE Strategy 2SPE Strategy 2PassPass--Through CleanupThrough Cleanup

1. Sample is passed through sorbent and collected• no sample

enrichment

2. Matrix interferences are retained on sorbent

pass through


SPE Strategy 3SPE Strategy 3Dispersion CleanupDispersion Cleanup

Bulk sorbent is added to sample with agitation

Sample is filtered or centrifuged

Supernatant is collected for analysis

This is similar to pass-through cleanup, but less effective

- Dispersion SPE is a one stage (one theoretical plate) cleanup

- Pass-through SPE is a multi-stage cleanup


OutlineOutline



o Pre-Treatment



SummaryAppendix


IonIon--Exchange and MixedExchange and Mixed--ModeMode

Many compounds of environmental interest are weak acids (i.e. dinoseb) or weak bases (i. e. aniline). — weak acids can be ionized at high pH

— weak bases can be ionized at low pH

Some compounds are strong acids (i.e. PFOA) or strong bases (i.e. chlorhexidine) that are ionic except at extreme pH values

A few of these compounds are quaternary amines (i.e. paraquat), ionic at all pH

Ionizable Compounds


Why MixedWhy Mixed--Mode?Mode?

Mixed-Mode SPE extends pH range for good retention of acids or bases

Retention can be by reversed-phase, ion-exchange or both— Chose retention mode by adjusting pH

— ion-exchange allows for good retention in strong solvent

o acids can be retained by anion-exchange while bases/neutrals are washed off with strong solvent

o bases can be retained on cation-exchange while acids/neutrals are washed off with strong solvent

For environmental analysis, mixed-mode SPE allows simultaneous retention of acids and bases


OasisOasis®® Family of MixedFamily of Mixed--Mode Mode Sorbents:Sorbents:ReversedReversed--Phase Retention and Ion ExchangePhase Retention and Ion Exchange


Oasis MixedOasis Mixed--Mode Sorbents Mode Sorbents

PFOS, PFOA (perfluoroacids and related compounds)— Oasis WAX

Acidic Herbicides— Oasis MAX

Quats— Oasis WCX

Pharmaceuticals/pesticides (organic bases)— Oasis MCX

Strategies for Isolation and Enrichment of Individual Compounds or Compound Classes

Oasis® 2x4 method


IntroductionIntroduction

Perfluorinated compounds (PFCs) such as perfluorooctanesulfonate and perfluorooctanoic acid are persistent organic pollutants (POPs)

PFCs have been identified in environmental samples worldwide— PFOS can be detected at low PPT levels in most humans

— PFOS commonly found in arctic fauna

There is need for reliable analytical methods for PFCs in food, drinking water, tissue, plasma and blood

In this presentation we will discuss sample preparation for UPLC-MS determination of PFCs in water and tissue samples


UPLCUPLC--MSMS--MS SystemMS System

ACQUITY Ultra Performance LC™— Using 1.7μm particles, and at elevated pressures up to 15,000

psi

Shorter Analysis Time

Higher Resolution

Broad selectivity options

Quattro Premier™ XE— Fast acquisition rates

— Sensitive detection

Oasis sorbents— Cleaner samples


GoalsGoals

Develop an Acquity UPLCTM separation based on a recently published method*

Adapt or modify the SPE protocol for UPLC— River Water sample

— Chicken Liver tissue sample

Lower the quantification limits to under 1 ppb in Chicken Liver tissue, and Low ppt level in River Water sample

*S. Taniyasu et. al.J. Chrom. A., 1093 (2005) pp89-97


Structures of PFOS and PFOAStructures of PFOS and PFOA

F3CO

OH

F

F

F

F

F

F

F

F

F

FF

F

perfluorooctanoic acidPFOA

pKa ~ 1

perfluorooctanesulfonatePFOS

pKa<<1

F

F

F

F

F

F

F

F

F

FF

F

S

O

O

O-F3C

F

F

PFOA and PFOS are Persistent Organic Pollutants of high interest worldwide.


OasisOasis®® 2x4 Method 2x4 Method For Acids, Bases, and NeutralsFor Acids, Bases, and Neutrals

Neutrals

For Bases:pKa 2-10

Use Oasis® MCX

For Strong AcidspKa <1.0

Use Oasis® WAX

For Strong BasespKa >10

Use Oasis® WCX

For AcidspKa 2-8

Use Oasis® MAX

Prepare Sample

Condition/EquilibrateLoad Sample

Wash:5% NH4OH

Elute 1:100% MeOH

Elute 2:2% Formic Acid in MeOH

Protocol 2Prepare Sample


Wash:2% Formic acid

Elute 1:100% MeOH

Elute 2:5% NH4OH in MeOH

Protocol 1

Bases Strong Acids

Strong Bases Acids


Optimized SPE ProtocolOptimized SPE Protocolfor River Waterfor River Water

Conditions for Oasis® WAX 3 cc 60mg cartridges

Prepare SamplepH 3

Condition2 mL methanol/2 mL water

Load200 mL

Wash #11 mL 2% Formic acid

Elute 1 (Wash #2)2 mL methanol

Elute 22 mL 1% conc. ammonia in

10:90 methanol/MTBE

Oasis® WAXOptimized Protocol 1

Oasis® WAX sorbent was selected for these analytes

Logic: PFOA pKa ~1PFOS pKa < 1

Oasis® WAX

N O

N

N

N

NH

H

H+

+

H

mixed-mode weak anion-exchangepKa ~6

Samples were evaporated and reconstituted in 0.15 mL mobile phase


SPE Protocol SPE Protocol

Prepare SamplepH 3

Condition2 mL methanol/2 mL water

Load200 mL

Wash #11 mL 2% formic acid

Wash #22 mL methanol

Elute 22 mL 1% conc. ammonia in

10:90 methanol/MTBE

Oasis® WAXOptimized Protocol 1

@ pH 3 Sorbent, and analytes are fully charged(assures mixed-mode retention)

Maximum load for good recovery of C3, C4 and PFBS

Assures sorbent is charged

Removes neutrals and bases retainedby reversed-phase

MTBE based eluent minimizes elution of any retained humic material


200mL river water 6cc WAX _200uL recon _BK

1.25 1.50 1.75 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75

%

0

100

1.25 1.50 1.75 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75

%

0

100

PFOS_082306AQC21x50C18_3 Sm (SG, 1x1) 2: MRM of 3 Channels ES-TIC

1.34e4


1.34e4

PFBS/PFOS in River WaterPFBS/PFOS in River Water100 ng/L (ppt)100 ng/L (ppt)

Blank

Spiked River WaterPFBS

PFOS



0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00 4.20

%

0

100

0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00 4.20

%

0

100


9.38e4


9.38e4

Blank

Spiked River Water

C3C3--C7 in River WaterC7 in River Water100 ng/L (ppt)100 ng/L (ppt)

C5

C4C3

C7

C6



2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00 6.50 7.00 7.50 8.00

%

0

100

2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00 6.50 7.00 7.50 8.00

%

0

100

PFOS_082306AQC21x50C18_3 1: MRM of 5 Channels ES-TIC

1.61e5

PFOS_082306AQC21x50C18_4 1: MRM of 5 Channels ES-TIC

1.61e5

C8C8--C12 in River WaterC12 in River Water100 ng/L (ppt)100 ng/L (ppt)

Blank

Spiked River Water

C12

C8 C10C9

C11


River Water RecoveriesRiver Water Recoveries

Spike Levelμg/L PFBS PFOS C3 C4 C5 C6 C7 C8 C9 C10 C11 C120.10 122 109 108 119 97 154 107 83 121 101 101 1000.30 110 117 95 132 105 110 119 126 137 118 94 950.70 102 98 91 107 93 118 100 78 103 126 119 1211.0 113 94 128 106 98 130 100 88 100 110 117 874.0 104 86 101 99 99 102 102 92 115 99 84 6810 104 100 98 101 100 87 89 82 103 99 101 66

Spike Levelμg/L PFBS PFOS C3 C4 C5 C6 C7 C8 C9 C10 C11 C120.10 122 109 108 119 97 107 83 121 101 101 1000.30 110 117 95 132 105 110 119 126 137 118 94 950.70 102 98 91 107 93 118 100 78 103 126 119 1211.0 113 94 128 106 98 130 100 88 100 110 117 874.0 104 86 101 99 99 102 102 92 115 99 84 6810 104 100 98 101 100 87 89 82 103 99 101 66

Spike Levelμg/L PFBS PFOS C C4 C5 C6 C7 C8 C9 C10 C11 C120.10 122 109 108 119 97 107 83 121 101 101 1000.30 110 117 95 132 105 110 119 126 137 118 94 950.70 102 98 91 107 93 118 100 78 103 126 119 1211.0 113 94 128 106 98 130 100 88 100 110 117 874.0 104 86 101 99 99 102 102 92 115 99 84 6810 104 100 98 101 100 87 89 82 103 99 101 66


Observations/RecommendationsObservations/Recommendations

Fluorocarbon parts, tubing, etc. are potential sources of interferences— UPLC fluidic lines were conditioned with 2% TFA in propanol

followed with 4% conc. ammonia in water (4 hours each step)Polypropylene (PP) lab ware may be best for sample prep— Do not use Teflon!! (possible positive interference)— Analytes may adsorb to glass (possible negative interference)C3-C5 analytes are highly volatile— Evaporative losses are possible, much more so at very low pH

Samples in glass vials may show loss of some analytes with time — Analyze within 24 hrs of sample prep


ConclusionsConclusions

• Oasis® WAX SPE method is effective for isolation and enrichment of C4-C8 perfluorosulfonic acids and C3-C12 perfluorocarboxylic acids from water and tissue

• Acquity UPLC™ provides significantly reduced analysis time and improved chromatographic behavior for these compounds compared with traditional HPLC

The Quattro Premier XE™ API mass spectrometer, operated in MRM mode, provides outstanding sensitivity and selectivity for these compounds


Acidic HerbicidesAcidic Herbicides

Cl

OCH2COOHCl

2,4-D

These herbicides, such as 2,4-D, are used in cultivated agriculture, in pasture and rangeland applications, forest management and home and garden. Also in aquatic applications.

Step 1 – characterize analytes they are acids pKa 3-6

For AcidspKa 2-8Select

Oasis® MAX


OasisOasis®® 2x42x4 MethodMethod::Starting Protocols For Acids and BasesStarting Protocols For Acids and Bases

Neutrals

For Bases:pKa 2-10

Use Oasis® MCX


Use Oasis® WAX


Use Oasis® WCX

For AcidspKa 2-8

Use Oasis® MAX

Prepare Sample


Wash:5% NH4OH

Elute 1:100% MeOH




Wash:2% Formic acid

Elute 1:100% MeOH


Protocol 1

Bases Strong Acids

Strong Bases Acids


OasisOasis®® 2x42x4SMSM MethodMethodChoose Starting ProtocolChoose Starting Protocol

The Oasis MAX cartridge was chosen for retention of acid herbicides

Logic: 2,4-D and other acid herbicides

pKa 3-6

For AcidspKa 2-8

Use Oasis® MAX

Prepare Sample


Wash:5% NH4OH*

Elute 1:100% MeOH


Protocol 2

Acids

Cl

OCH2COOHCl

2,4-D


Conditions for 6 cc cartridges

OasisOasis®® MAX SPE MethodMAX SPE MethodAcidic Herbicides 1Acidic Herbicides 1µµg/kgg/kg in River Waterin River Water

Prepare SampleCondition

3 mL methanol/ 3 mL water

Load300 mL sample

Wash #13 mL 5% NH4OH

Elute 1 (Wash #2)3 mL methanol

Elute 24 mL 2% Formic Acid in MeOH

Evaporate and Reconstitute

Oasis MAXProtocol 2

Waters XTerra™MS C18, 2.1 x 100 mmA: 15mM ammonium formate (pH 3.5), B: acetonitrile25% B to 60% B in 9 min, hold 5 min, to 90% B in 16 min

Waters ZQ, ESI-, SIR mode

20 min

12

3

4

5

67,8

9

10

11

1213

14

15

1 ppb in river water

1. picloram 2. chloramben3. 4-nitrophenol4. bentazon5. 2,4-D6. MCPA7. dichlorprop8. 2,4,5-T

9. MCPP10. DCB11. acifluorfen12. 2,4,5-TP13. 2,4-DB14. dinoseb15. pentachlorophenol


Paraquat/DiquatParaquat/Diquat

For Quats

SelectOasis® WCX

N N CH3CH3++

N N CH3CH3++ +

+N

N+

+N

N

paraquat diquat

The Oasis WCX cartridge was chosen for these analytes

Logic: quats are cationic at all pH values

quats can be eluted from Oasis WCX with acidic solvent


0 1 2 3 4 5 6 7 8 9 10 11 12 13 14pH

Ret

entio

n Fa

ctor

(k’)

Oasis® WCX

Oasis® MCX

Retention

Elution

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14pH of elution solvent (80:20 acetonitrile/water)

% E

lute

d

Oasis® WCX

Oasis® MCX

note: quats are eluted from Oasis WCX at low pH

Retention and Elution of Retention and Elution of ParaquatParaquaton Mixedon Mixed--Mode SorbentsMode Sorbents


Paraquat/DiquatParaquat/DiquatOptimized Oasis MCX ProtocolOptimized Oasis MCX Protocol

Prepare Sample

Condition/Equilibrate

Load Sample

Wash1:5% NH4OH in water

Wash2:100% MeOH

Elute :1.5 mL ACN/water/TFA 84:14:2

Protocol 2

optimized elution solventacetonitrile/water/TFA

For Quats

SelectOasis® WCX


Optimized SPE ProtocolOptimized SPE ProtocolParaquat/DiquatParaquat/Diquat

Oasis® WCX SPE MethodParaquat/Diquat

50:1 sample enrichment

Prepare Sampleadjust to pH 7

Condition1mL methanol/ 1 mL water

Loadup to 25 mL sample

Wash 1 mL pH 7 buffer/1mL methanol

Elute1.5 mL ACN/water/TFA 84:14:2



0.5 mL mobile phase

Prepare Sample

Condition1mL methanol/ 1 mL water

Loadup to 25 mL sample

Wash 1 mL pH 7 buffer/1mL methanol

Elute1.5 mL ACN/water/TFA 84:14:2



0.5 mL mobile phase


LCLC--MS ConditionsMS Conditionsparaquat/diquatparaquat/diquat

MS ConditionsInstrument:MS ConditionsInstrument: Waters Quattro micro API™

Paraquat:Paraquat: cone 40 VMRM

cone 40 VMRM 171 → 77 (CID 35 eV)

171 155 (CID 35 eV77 (CID 35 eV)

171 → 155 (CID 35 eVcone 15 V

MRMcone 15 V

MRM 93* 93* 77 (CID 30 eV) 77 (CID 30 eV)

Diquat:Diquat: cone 40 VMRM

cone 40 VMRM 183

183 183 →183 →

157 (CID 30 eV)168 (CID 35 eV)157 (CID 30 eV)168 (CID 35 eV)

cone 15 VMRM

cone 15 VMRM 92* 92* 85 (CID 30 eV)

→

→

LC ConditionsColumn: Waters Atlantis™ HILIC, 2.1 x 150 mmFlow: 0.4 mL/minMobile Phase: 40% acetonitrile

60% aqueous buffer pH 3.7(200 mM ammonium formate)

Column Temp: 30 oCSample Temp: 5 oCInjection: 10 µL

0.20 µg/L Spiked Sample

diquat

paraquat

1 2 3 4 5 6 7 81

1001

100


0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

0 2 4 6

r2 = 0.998

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

0 2 4 6

r2 = 0.998

ValidationValidation

Performance was demonstrated from 0.1 to 5 µg/L using 20 mL samples of Sudbury River water.

Paraquat Intraday Results (1 µg/L)Day 1 1.08 µg/L (8.1% RSD)Day 4 1.10 µg/L (8.0% RSD)Day 5 0.95 µg/L (7.1% RSD)

Overall (n=15) 1.04 µg/L (9.8% RSD)


Advantage of OasisAdvantage of Oasis®® WCXWCXfor for Paraquat/DiquatParaquat/Diquat

No Salts required for elution— Eluent can be evaporated and reconstituted in

mimimal volume— Method is more compatible with API mass

spectrometry— Method is more compatible with ion-pair

chromatography— Method is more compatible with on-line SPE


OasisOasis®® 2x42x4 MethodMethod::Starting Protocols For Acids and BasesStarting Protocols For Acids and Bases

Neutrals

For Bases:pKa 2-10

Use Oasis® MCX


Use Oasis® WAX


Use Oasis® WCX

For AcidspKa 2-8

Use Oasis® MAX

Prepare Sample


Wash:5% NH4OH

Elute 1:100% MeOH




Wash:2% Formic acid

Elute 1:100% MeOH


Protocol 1

Bases Strong Acids

Strong Bases Acids


Pharmaceuticals/Pesticides/Industrial Pharmaceuticals/Pesticides/Industrial ChemicalsChemicals((Organic Bases, Organic Bases, pKapKa 22--10)10)

Example: Aniline (pKa ~ 4)

For BasespKa 2-10Select

Oasis® MCX

NH2 NH3

pH 2

+

Prepare Sample


Wash:2% Formic acid

Elute 1:100% MeOH

Elute :5% NH4OH in MeOH

Protocol 1

for GC, use 90:10MTBE/methanolic ammoniafor elute 2

To recover acids and neutrals, analyze Elute 1


Minutes0 10 20 30 40 50

1

2

3

9

10

11

13

16

17

19 20

2223

2425

26

27

28

29

30

3132

33

NPD

Pharmaceuticals/Pesticides/Industrial Pharmaceuticals/Pesticides/Industrial ChemicalsChemicals((Organic Bases, Organic Bases, pKapKa 22--10)10)

COMPOUND % RECOVERY ±RSD (20 µg/L Tap Water)

1. pyridine 61 (17)2. picoline 77 (16)3. aniline 90 (11)

o-toluidine 82 (12)10. phentermine 73 (18)11. chloroaniline 82 (11)13. phenylenediamine 93 (15)

16. 2 -nitroaniline 95 (7.2) 17. 3 -nitroaniline 103 (8.5)19. 1 -aminonaphthalene 87 (5.1)20. 2 -aminonaphthalene 88 (8.5) 22. 2 -methyl -5-nitroaniline 104 (6.2) 23. 4 -nitroaniline 106 (8.7)24. diphenylamine 93 (4.4)

26. aminobiphenyl 105 (4.2)30. dimethylaminoazobenzene 100 (3.9) 31. dimethylbenzidine 64 (8.9)33. dichlorobenzidine 111 (6.0)

9.


1. pyridine 61 (17)2. picoline 77 (16)3. aniline 90 (11)

o-toluidine 82 (12)10. phentermine 73 (18)11. chloroaniline 82 (11)13. phenylenediamine 93 (15)

16. 2 -nitroaniline 95 (7.2) 17. 3 -nitroaniline 103 (8.5)19. 1 -aminonaphthalene 87 (5.1)20. 2 -aminonaphthalene 88 (8.5) 22. 2 -methyl -5-nitroaniline 104 (6.2) 23. 4 -nitroaniline 106 (8.7)24. diphenylamine 93 (4.4)

26. aminobiphenyl 105 (4.2)30. dimethylaminoazobenzene 100 (3.9) 31. dimethylbenzidine 64 (8.9)33. dichlorobenzidine 111 (6.0)

9.

GC-NPD ConditionsAgilent 5890 series II30 m x 0.25 mm (ID) RTX 5 (0.25 µm)EPA 8270C bases, 20 ug/L 200 mL tap water/Oasis MCX protocol2 uL inject


SummarySummary

Sample Preparation is necessary to obtain the best analytical results

SPE is a very versatile and cost efficient sample preparation technique for environmental samples.

Waters provides strategies which combine sorbents, formats and methodologies resulting in optimal SPE protocols.

Whether for analysis by LCMS or GCMS; Waters analytical solutions, including SPE, cover a wide range of sample matrices and compounds classes


OasisOasis®® MixedMixed--Mode Sorbents Mode Sorbents

Mixed-Mode strong ion-exchange sorbents (Oasis MCX and Oasis MAX) can simultaneously retain polar acids and bases better thanthe best reversed-phase sorbents such as Oasis HLB

— Oasis® MCX, sample adjusted to low pHo acids/neutrals retained by reversed-phase o bases retained by mixed-mode cation-exchange

— Oasis® MAX, sample adjusted to high pHo acids retained by mixed-mode anion-exchangeo bases/neutrals retained by reversed-phase

Strategies for Multiresidue Isolation and Enrichment(acids, bases and neutrals together)


Consider:

Aniline, phenol and benzyl alcohol on Reversed-Phase SPE

SPE of Acids and Base/NeutralsSPE of Acids and Base/NeutralsReversedReversed--Phase LogicPhase Logic

NH2 OH OH

At pH 2: Aniline is cation – not retainedPhenol is protonated – retainedBenzyl alcohol is neutral – retained

At pH 11 Aniline is neutral – retainedPhenol is ionized – not retainedBenzyl alcohol is neutral – retained


Consider:

Aniline, phenol and benzyl alcohol on Mixed-Mode SPE

SPE of Acids and Base/NeutralsSPE of Acids and Base/NeutralsMixedMixed--Mode LogicMode Logic

NH2 OH OH

At pH 2 on Oasis® MCX: Aniline is cation –retainedPhenol is neutral – retainedBenzyl alcohol is neutral – retained

At pH 11 on Oasis® MAX: Aniline is neutral – retainedPhenol is anion – retainedBenzyl alcohol is neutral – retained


Multi residue AnalysisMulti residue AnalysisOasisOasis®® MCX Method for GCMCX Method for GC

prepare reagent using anhydrous ammonia in methanol (Aldrich)

Prepare SamplepH 2

Condition2 mL DCM, 2 mL methanol, 2 mL water

Load250 mL sample

Wash2 mL 5 % MeOH/water

Elute4 mL of 0.7 M NH4OH in 90:10 DCM/MeOH

Oasis® MCXOptimized Protocol

Dry over Sodium SulfateEvaporate to Final Volume

Micro K-D


Minutes0 10 20 30 40 50

1

2

3

9

10

11

13

16

17

19 20

2223

2425

26

27

28

29

30

3132

33

4

5

6

7,8

12

14

1518

21

FID

NPD

bases, acids, neutrals

SPE for Base/Neutrals and Acids SPE for Base/Neutrals and Acids OasisOasis®® MCX GC ProtocolMCX GC Protocol


1. pyridine 61 (17)2. picoline 77 (16)3. aniline 90 (11)4. phenol 65 (14)5. benzyl alcohol 75 (25)6. o-cresol 91 (8.6)

7,8. m,p-cresol 91 (8.9)9. o-toluidine 82 (12)10. phentermine 73 (18)11. chloroaniline 82 (11)12. dichlorophenol 57 (6.2)13. phenylenediamine 93 (15)

14. 2-methylnaphthalene 81 (8.0)15. trichlorophenol 54 (10)16. 2-nitroaniline 95 (7.2) 17. 3-nitroaniline 103 (8.5)18. dibenzofuran 80 (5.4)19. 1-aminonaphthalene 87 (5.1)

20. 2-aminonaphthalene 88 (8.5) 21. tetrachlorophenol 35 (17)22. 2-methyl-5-nitroaniline 104 (6.2) 23. 4-nitroaniline 106 (8.7)24. diphenylamine 93 (4.4) 25. phenacetin 85 (7.3)

26. aminobiphenyl 105 (4.2)27. dinoseb 90 (7.1)28. nitroquinoline oxide 100 (6.5)29. methapyrilene 105 (5.5)30. dimethylaminoazobenzene 100 (3.9) 31. dimethylbenzidine 64 (8.9)

32. acetamidofluorene 135 (5.4)33. dichlorobenzidine 111 (6.0)

Documents

Sample Preparation Strategies for Water Analysis - AmChamamcham.dk/dl/events/ESAC/Presentation3.pdf · ©2007 Waters Corporation 3 For high sensitivity analyses, such as those employing