continual innovation and achievement: 40 years of ion ... · reagent-free ion chromatography (rfic)...

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The world leader in serving science

25th Annual Good Laboratory Practice Conference

Virginia AWWA / VWEA

Laboratory Practice Committee

Buddy Goodnight

NA COE IC Support Specialist

Continual Innovation and Achievement: 40 Years of Ion Chromatography

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Agenda

• Define Ion Chromatography

• History of Ion Chromatography

• Typical IC Configuration

• Markets

• Role of Chemical Eluent Suppression

• Typical Applications

• (Reagent Free Ion Chromatography) Technology

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Ion Chromatography

Ion Chromatography is an analytical technique that

utilizes ion exchange mechanisms to separate ionic

substances followed by detection utilizing conductivity,

amperometry, UV/Vis, fluorescence, and mass

spectrometry.

Analyte classes include:

Anions

Cations

Organic Acids

Amines

Transition Metals

Carbohydrates & Amino Acids

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Definitions http://en.wikipedia.org/wiki/Ion

• Ion chromatography (or ion-exchange chromatography) is a process

that allows the separation of ions (or ionizable) and polar molecules

based on the charge properties of the molecules. It can be used for

almost any kind of charged molecule including carboxylic acids,

amines, large proteins, carbohydrates, small nucleotides and amino

acids.

• The solution to be injected is usually called a sample, and the

individually separated components are called analytes. It is often used

in water analysis, quality control, and protein purification.

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Definitions

• Mobile phase or eluent

• The phase that travels through the column

• Typically a liquid

• Its purpose is to transport the analytes and permit interaction with the

stationary phase

• Stationary phase or Column

• Generally an inert solid or gel to which various functional groups are

attached

• The stationary phase interacts with the compounds being analyzed and

slows their passage through the column

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• Limited automated instrumentation

• Most ion analysis was colorimetric, turbidimetic or ISE

• Ion-exchange resins were used but only as a sample prep tool

for wet chemical analysis methods

• Data handling: Do you remember “chart recorders” and “integrators”?

• Reports: Typed with carbon paper on an electric typewriter and mailed with

a stamp

Disadvantages of Wet Chemistry Methods

• Large matrix effects, color interference, low sensitivity,

labor intensive

Chromatography Goal: Water as a mobile phase

• Tried and discarded

The Landscape Before Ion Chromatography

Ionic analysis was a desert, a wet desert

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Essential Elements of an Ion Chromatography System

IC was a disruptive technology.

What was missing for today’s IC?

• Manufacturing process for ion-exchange stationary phases

• Suitable mobile phase

• Aqueous solutions had electrical conductance due to high ionic content

• Device to strip ionic content from the mobile phase

• Low electrical conductance. Water is the perfect background (goal)

• Detector for analytes that do not have chromophores

• A conductivity cell for conductivity detection

• Instrument to run the separation and detection methods

• “Integrator” or method to process or present the data

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• Defined “ion chromatography”

exclusively as a combination of

1) Ion-exchange separation

2) Eluent suppression

3) Electrical conductance detection

Fathers of Ion Chromatography

• Hamish Small, Bill Bauman and Tim Stevens

(Dow Chemical)

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The Very First Ion Chromatograph – Dow Chemical

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Cations Came First: 26 min for Three Cations!

• 10 mM lithium, sodium, and potassium

as chloride salts

• Only monovalent cations

• Column: a lightly sulfonated

styrene–divinylbenzene (DVB) polymer, 50 µm

• Eluent: 20 mM HCl

• Injection Vol.: 100 µL

• Dowex 1 “Stripper” (suppressor): (hydroxide form)

Hamish laboratory notebook,

November 9, 1971

Hamish Small, LCGC April 02, 2013

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In 1975, Established the Foundations of IC

• Licensing agreements: Dow Chemical and Durrum Instruments

• Durrum/Dow ION EXchange = DIONEX

• Formation of one company

– Dionex Corporation (1978)

• Column Process: Surface-sulfonated

styrene–DVB resin with a suspension of a

colloidal anion-exchange resin

• Eluent system: Carbonate based buffers

• Suppressor: Switching packed bed

• Detector: Suppressed conductivity

• First commercial IC: Dionex Model 10

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Sample Inject

(Autosampler)

Eluent

Non-

Metallic

High-

Pressure

Pump

Separation Detection

Data

Management

Ion-Exchange

Analytical Column

Eluent

Suppressor

Conductivity

Detector

Principles of Ion Chromatography

Main Menu

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Ion Chromatography answers a Wide Diversity of Application Areas and the Markets it Supports

• Environmental

• Chemical & Petrochemical

• Pharmaceutical & Biotechnology

• Power & Energy Production

• Electronics, Semiconductors & Plating

• Pulp & Paper

• Foods & Beverages

• Personal Care & Household Products

• Agricultural Products

• Forensics

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Typical Ion Chromatographic System - Anion Analysis

Eluent Bottle (CO3/HCO3 or NaOH)

Pump

Guard Column

Analytical Column

Suppressor

Regen In (H2SO4)

Conductivity Cell

Chromatography Software Ion

Exchange Separation

Post-

Suppression Conductivity

Data Handling and Instrument Control

Sample Injection

Regen Out (H2SO4)

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Chemical Suppression

Analytical Column (Anion

Exchanger )

Anion Suppress

or (Cation

Exchanger)

HF, HCI, H2SO4 in H2CO3

NaF, NaCI, Na2SO4 in Na2CO3

Waste

H+

Eluent (Na2CO3)

Sample F–, CI–, SO4

2–

F–

SO42

– CI –

Time

µS

With Suppression

H2CO3

Without Suppression Counterions

µS

Time

F–

SO42

– CI–

Na2CO3 Background

Main Menu

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1980s: Developing Continuous Suppression

• Switching packed bed suppressors required periodic regeneration

• Disadvantages: defined capacity (4 to 8 hr)

• Periodically replace with another suppressor or

• Wait while suppressor was being regenerated off line

• Led to development of continuous tubular suppressor (Tim Stevens –

Dow; Sandy Dasgupta –Texas Tech)

• In 1985 Dionex Corp. introduced flat membrane continuous suppressor,

the MicroMembrane Suppressor (MMS)

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Suppressor Innovations Inspire Column Innovations

• Goal: NaOH as anion eluent because it suppresses to water

Efficient suppressors to neutralize high concentrations of strong base

• Thermo Scientific™ Dionex™ MMS™ MicroMembrane suppressor has sufficient

suppression capacity

Sparking Column Innovations

• Optimized polymer branching in stationary phases to include hydroxide

selectivity

• Result: increased selectivity, more control over resolution

• Launched with the introduction of Dionex IonPac AS5A column in 1985

Chris Pohl: Stationary Phases and Suppressors in Ion Chromatography, 1975-2000

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Selection of Eluent Driven by Column Innovations

• Goal: NaOH as anion eluent because it suppresses to water

• Limitations

• Hydroxide has low affinity for 1970’s anion-exchange columns

• Columns require high eluent concentrations to elute analytes

• High hydroxide concentrations strain the suppressor capacity

• Compromise: Carbonate eluents

• Converts to weak and low conducting carbonic acid

• Compatible with the early suppressors

• Issue: the baseline conductivity is ~10–22 µSiemens

• Eluent family for more than 20 yrs

circa 1974

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1990’s: Fixing Carbonate Eluent Limitations

• Conductivity is 10–20x higher than pure water which impacts sensitivity

• Nonlinear responses at lower analyte levels

• Gradient separations were difficult

• Ramping

• Baseline disturbances

• Slow buffering

Solution: Hydroxide based eluent

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Hamish Small et al.

Publish First IC paper

Switching Packed Bed Suppressor

Fiber suppressor (continuous suppression)

Micromembrane suppressor

(high capacity continuous

suppression) I

Cation Suppression Introduced

Self-regenerating micromembrane

suppressor (electrolytic

regeneration)

1975 1981

1985

1992 1978

2007

Thermo

Scientific™

Dionex™

SRS™ 300

Suppressor

Continuous development to extend lifetime,

pressure range, and robustness

2010

Cap IC

Thermo

Scientific™

Dionex™ CES™

300 Suppressor

2013

Thermo

Scientific™

Dionex™ CRS™

500 Suppressor

Suppressor Development

Thermo

Scientific™

Dionex™ ERS™

500 Suppressor

2015

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Altering Selectivity in Ion Chromatography

Mobile phase parameters

• Eluent Ion

• Eluent concentration

• Eluent pH

• Organic additives

• Temperature

Mainly retention changes Little control on selectivity

Better control of selectivity

Stationary phase parameters

• Material

• Functional group

• Capacity

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Supermacroporous

(SMP) Microporous

Resin Technology

• Resin: polystyrene-divinylbenzene

• Bead type: microporous, macroporous, supermacroporous

• Size: down to 4 µm i.d.

Hyperbranched Polymer

Grafting

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Pellicular Anion-Exchange Bead (Latex)

SO– 3

SO– 3

SO– 3

SO– 3

SO– 3

SO– 3

0.1-mm Diameter Latex Core Ion-Exchange

Surface

R3N+

R3N+

N+R3

N+R3

N+R3

R3N+

R3N+

N+R3

N+R3

R3N+

R3N+

N+R3

N+R3

R3N+

R3N+

N+R3

N+R3

SO–3

3183-03

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Latex Agglomerated Supermacroporous Resin

Anion Exchange Latex Coating

65-nm Diameter Microbeads

Ethylvinylbenzene-Divinylbenzene Core

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+

Cl–

Cl–

Cl–

Cl–

Positively

Charged

Polymeric

Particle

Anion Exchange Mechanism

Stationary Phase

OH– < BO3– < HCO3

– << CO32–

+

+

+

+

+

+ +

Mobile Phase

Cl–

Eluent

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Ion-Exchange Chromatography Principles

Ion separation is determined by the analytes different

affinities towards the stationary phase.

----> High affinity for the functional group results in longer retention

Ion properties determining retention:

● Ion’s charge: single charge = weak retention, double charge = strong retention

● Ion’s size: large solvated ion = shielded charge = weak retention

● Ion’s polarizability (hydrophobicity): no or poor solvation = very strong retention

General isocratic elution order for anions:

F– Cl– Br– NO3– HPO4

2– SO42–

For cations:

Li+ Na+ NH4+ K+ Mg2+ Ca2+ Sr2+ Ba2+

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Total Conductivity (Σ ) Detection

Σ = λanion + λcation

where λ = equivalent conductance

Anions Cations

OH- 198 H+ 350

F- 54 Li+ 39

Cl- 76 Na+ 50

NO3- 71 K+ 74

Acetate- 41 CH3NH3+ 58

Benzoate- 32 N(CH3CH2)4+ 33

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1° Amines

R-NH2

2° Amines

R’

R-N-H

4° Amines R4N

+

Functional Groups Detected with Suppressed Conductivity

3° Amines

R’

R-N-R”

Suppressed Conductivity Detection

Inorganic Anions

e.g., F_,

CI_,

NO2

_,

NO3

_,

SO42_

,

PO43_

, I_,

Br

_, CIO3

_,

SCN

_,

S2 O3 2_

Inorganic Cations

Alkali Metals:

Li+, Na+, K+,

Rb+, Cs+,

Alkaline Earths:

Mg2+, Ca2+

Sr2+, Ba2+ ,

Ammonia:

NH4+

Phosphates

O

R-O-P-OH

OH

Sulfates

R-O-S-OH

O

O

Sulfonates

R-S-OH

O

O

Phosphonates

R-P-OH

O

OH

Carboxylics

R-C-OH

O

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Column: IonPac™AG4A-SC/AS4A-SC Eluent: 1.7 mM Sodium bicarbonate/ 1.8 mM sodium carbonate Flow Rate: 2.0 mL/min Injection: 50 µL Detection: Suppressed conductivity, ASRS™ ULTRA Anion Self Regenerating Suppressor, recycle mode

Peaks: 1. Fluoride 2 mg/L 2. Chloride 3 3. Nitrite 5 4. Bromide 10 5. Nitrate 10 6. Phosphate 15 7. Sulfate 15

0 2 4 6 8

Minutes

0

µS

10

10

1

2 3 4 5

6

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IC Written into U.S. EPA Method 300.0 (A) (Drinking Water Analysis)

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Column: IonPac® AG14A, AS14A, 5µm, 3 x 150 mm

Eluent: 8.0 mM sodium carbonate 1.0 mM sodium bicarbonate

Flow Rate: 0.8 mL/min

Temperature: 30 °C

Inj. Volume: 5 µL

Detection: Suppressed conductivity,

ASRS® ULTRA II, 2 mm, AutoSuppression® recycle mode

Peaks: 1. Fluoride 5 mg/L (ppm) 2. Acetate 20 3. Chloride 10 4. Nitrite 15 5. Bromide 25 6. Nitrate 25 7. Phosphate 40 8. Sulfate 30

Isocratic Anion Separation EPA 300.0

0 1 2 3 4 5 6

Minutes

0

12

1 3

4 5

7

2

µS

6 8

15907

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Reagent-Free Ion Chromatography (RFIC) Systems

● RFIC™ systems are ion chromatography systems

that utilize electrolysis and/or electrolytic devices

to generate (EG) or regenerate (ER) eluents in

the separation processes and produce

regenerant ions in the suppression process.

●RFIC-EG™ systems (2003)

●RFIC-ER™ systems (2008)

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Electrolytic Eluent Generation and Suppression

Eluent

Generation

(KOH)

Anion Suppressor

Anion Column

HF, HCI, H2SO4 in H2O

KF, KCl, K2SO4 in KOH

Waste

Time

µS

With Suppression

Waste H+

K+

F–, CI–, SO42 –

F–

CI– SO42–

Sample

H2O

Detector

Without Suppression

Counterions

µS

Time

F– CI– SO42 –

KOH

Main Menu

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Essential Elements of RFIC

Electrolytic functions built on the simple electrolysis of water

• Electrolytic eluent generation and purification

• Electrolytic suppression of eluents

“Just Add Water”

Cathode 2e– + 2H2O 2OH– + H2 (g)

Anode H2O 2 H+ + ½ O2 (g) + 2e– + OXIDATION

REDUCTION -

RFIC = Reagent-Free Ion Chromatography

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Benefits of RFIC-EG™ Systems

• Use EGC cartridges to generate high purity acid, base, or carbonate eluents on-line using deionized water as the carrier

• Produce eluents of precise and reproducible concentrations through the convenient control of electrical current

• Improve retention time reproducibility for both isocratic and gradient separations

• Compatible with a wide range of high-performance detection methods including conductivity, UV-Vis, electrochemical, and MS

• Improve the ease of use, sensitivity, and performance of IC methods (day-to-day and lab-to-lab) for the determination of target analytes in a wide variety of sample matrices

• Simplify system operation and reduce overall operating cost

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Separation of Common Anions and Oxyhalides:

(A) IonPac AS9-HC (Isocratic)

0 5 10 15 20 25

0

10

µS

1

2 3

4

5

7

6

8

11

10

Minutes

–1

40

1 2 3

4 5 6 7 8

9

10

11

0 5 10 15 20 25 30

µS

(B) IonPac AS19 (Gradient)

Column: (A) IonPac AG9-HC, AS9-HC (B) IonPac AG19, AS19, 4 mm

Eluent: (A) 9 mM sodium carbonate (B) Potassium hydroxide: 10 mM 0 to 10 min, 10–45 mM 10 to 25 min

Eluent Source: (B) ICS2100 EG with CR-ATC

Temperature: (B) 30 °C

Flow Rate: 1.0 mL/min

Inj. Volume: 25 µL

Detection: ASRS® ULTRA II, 4 mm, recycle mode, 130 mA

Peaks: 1. Fluoride 3 mg/L 2. Chlorite 10 3. Bromate 20 4. Chloride 5 5. Nitrite 15 6. Chlorate 25 7. Bromide 25 8. Nitrate 25 9. Carbonate — 10. Sulfate 30 11. Phosphate 40

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RFIC™ System for Anions Using EPA Method 300.0 (A)

Column: IonPac® AG18, AS18, 4 mm

Eluent: 22–40 mM KOH from 7–8 min

Eluent source: ICS-2000 with CR-ATC

Temperature: 30 °C

Flow rate: 1.0 mL/min

Inj. volume: 25 µL

Detection: ASRS® ULTRA, 4 mm recycle mode, 100 mA

Peaks (A) (B) 1. Fluoride 2 0.07 mg/L (ppm) 2. Chloride 5 45.3 3. Nitrite 10 0.07 4. Carbonate — --- 5. Bromide 20 0.03 6. Sulfate 10 58.7 7. Nitrate 20 2.88 8. Phosphate 30 1.44

Sample: Sunnyvale, CA, drinking water

0 4 8 12 16 0

µS

20

1

2 3

4

5

6 7

8

Standard

Minutes 0 4 8 12 16

0

14

1

2

3 4 5

6

7 8

µS

Municipal Drinking

Water

(A)

(B)

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Eluent

Autosampler

High-Pressure

Nonmetallic

Pump

Thermo Scientific

Dionex

IonPac™ NG1

Thermo Scientific

Dionex

IonPac AS7

Post column

Reagent

System Configuration for Hexavalent Chromium by U.S. EPA Method 218.6

Mixing

Tee

UV-vis

Detector

Knitted Reaction

Coil

Waste

Sample Loop

17901

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Chromate Detection

Column: IonPac AS7 (2 × 50 mm), AS7 (2×250)

Eluent: 250 mM (NH4)2 SO4,

100 mM NH4OH

Flow: 0.36 mL/min

Inj. Vol: 1000 µL

Postcolumn Reagent: 2 mM diphenylcarbazide

10% methanol

1N sulfuric acid

Reaction Coil: 125 µL

Flow Cell: Standard (PEEK), 11 µL

0 1 2 3 4 5 6 7 8 9 10 -0.5

2.5

Minutes

mAU

A. DI water blank

B. 0.005 µg/L Cr(VI) in DI water

B

A

Blank and low-level chromate Comparison of chromate in high ionic-strength

water (HIW) and Sunnyvale tap water

Column: IonPac™AS7 (2 × 50 mm),

AS7 ( 2 × 250 mm)

Eluent: 250 mM (NH4)2 SO4,

100 mM NH4OH

Flow: 0.36 mL/min

Inj. Vol: 1000 µL

Postcolumn Reagent: 2 mM diphenylcarbazide

10% methanol

1N sulfuric acid

Reaction Coil: 125 µL

Flow Cell: Standard (PEEK), 11 µL

A. 0.1 µg/L Cr(VI) in DI water

B. 0.1 µg/L Cr(VI) in HIW

C. 0.1 µg/L Cr(VI) spiked in

Sunnyvale, CA tap water

1 ppt Detection can be achieved!

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• IC is extremely sensitive and can measure down to 1ppt MDL

• Cr(VI) is very stable in most waters

• Holding time in EPA 218.7 is 14 days

• Cr(VI) contamination has been found in chemicals used for eluents and preservation

buffers

• EPA 218.7 allows easier pH adjustment to pH 8.0

• Also uses weaker buffers to prevent column overloading

• Free chlorine will oxidize Cr(III) to Cr(VI)

• Need to minimize impact with NH4SO4 addition to form chloramines

• Chlorine must be <0.1 ppm prior to collection

• Filtering samples is not required

• Preservative is prepared once every 30 days

• Can add preservative to bottles prior to sample addition

• Chilling of samples is not required.

Analytical Issues involving Cr(VI) determination in water

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Maintaining a Healthy IC

• In this workshop session, we will provide tips and tricks to maintain your IC at optimum performance for a more practical, lean, and green operation.

• Four Steps to IC Troubleshooting 1. Identify the symptom

Know your systems vital signs Check for leaks and alarms

2. Identify the consequences 3. Isolate the cause of the problem

Start with the basics Change only one variable at a time Record the changes to the system and the results of the changes

4. Repair the problem

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Know Your System’s Vital Signs

Backpressure

Background

Baseline noise

Retention times

Peak shape

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ICS Components That Impact System Wellness

Eluent

Pump

Injector

Column

Suppressor

Detector

“Keep it Simple”

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M & T: Backpressure

Measured by the pump transducer

Too much backpressure may cause… Not enough backpressure may cause…

Leaks

Damage to components

Noise

Performance problems

Trapped bubbles

Noise

Unstable flowrate

Performance problems

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M & T: Background Conductivity

Symptoms

Background too high Background too low

Eluent too strong

Contaminated eluent

No current

Bad water

Eluent too weak

Wrong eluent

Bad flow cell

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M & T: Retention Times

Possible causes of retention time shift:

Contamination

Incorrect eluent strength

Gradient problem

Inconsistent flowrate

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M & T: Noise

Possible causes for baseline noise:

Small leak

Air

Pump problems

Turbulent flow path

Contaminated eluent

Temperature instability

Faulty electronics

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Pumps—Theory

• Piston

• Draws and expels liquid

• Check valves

• Control direction of flow

• Piston seal

• Allows piston to travel in and out

• Blocks liquid from leaving

• Series pumps

• Only the primary head has check

valves

• Sides collaborate & increase efficiency

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Check Valves

Check valve assemblies have removable

cartridges

Inlet and outlet assemblies are different

Inlet and outlet cartridges are identical

Purpose:

Controls direction of flow

FLOW

Check Valve Cartridge

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Suppressor Troubleshooting

• Alarms

• Spikes

• Increased noise

• High background conductivity

• Leaks

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M & T: Peak Shape

Peak shape variation may be a symptom of a problem

Peak broadening

Peak tailing

Peak fronting

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Tips for Column Care / Cleanup

• Always use a guard column to retain contaminants and protect the analytical column. Replace or clean guard when capacity is 50% from new.

• When unsure of the appropriate column cleaning solution, use 10x to 100x the eluent concentration.

• During column cleanup, place the guard column after the analytical column.

• Place after pressure transducer for minimum contamination.

• Use low flow rate: 0.5 mL/min for 4 mm columns.

• Always pump cleaning solution through the columns in the same direction as the eluent flow.

• Column manuals contain detailed information on operation, maintenance, and troubleshooting.

• Organic contaminants, use solvents; transition metals, use 0.2 M oxalic acid. Wash with deionized water in-between.

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Column Troubleshooting Summary

• Remake eluent

• Check for leaks

• Check if pump is running properly

• Check capacity on guard column

• Remove or replace the guard column; recheck backpressure and

retention time of standard

• Check bed supports for discoloration

• Clean or replace column

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To contact your local Thermo Fisher Scientific Sales Representative

• Central-Eastern Virginia/Eastern NC: Dan Ciminelli

• Washington/DC-Beltway Maryland/North VA: Donna Zwirner

• Charlotte, Greensboro, Fayetteville, and parts of Western NC and

Western Virginia: Samantha Stikeleather

• Your representative will be determined by your company location and

your zip code

• I can be contacted by email: buddy.goodnight@thermofisher.com

• http://www.thermoscientific.com/dionex

• https://appslab.thermofisher.com/

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Thank You!

Thank you for your attention!