MEPSMicro Extraction by Packed Sorbent

Jade AntonioSGE Europe Ltd

What is MEPS?

MEPS is a technique which combines sample preparation by solid phase

extraction with syringe based sample injection.

Objectives of Sample Preparation

• Make sure the analytes of interest are in solution

• Remove any interferences found in the sample matrix

• Concentrate the analytes to a level which can be detected

History of Sample Preparation

1941 – First use of Liquid:Liquid extraction

1966 – First paper on HPLC

1974 – First reported use of SPE

1993 – Solid Phase Micro Extraction (SPME)

2007 – MEPS

Impact of Sample Preparation

• An estimated 75% of labor time in an analytical labs are spent preparing and processing samples

• Any reduction in this time will have a big impact on a laboratory costs and increase profit.

Liquid-Liquid Extraction (LLE)

• Liquid-liquid extraction is used to separate compounds based on how easily they dissolve in two different liquids that will not mix.

• These liquids are usually water and an organic solvent.

• The extraction moves the compound from one liquid phase to the other.

Liquid Liquid Extraction

LLE has three main draw backs

• It requires large quantities of solvent

• It is not automated (labor intensive)

• Operator Inconsistency is enhanced

Solid Phase Extraction

• Used to isolate analytes of interest from a wide variety of matrices, including urine, blood, water samples

• Separation is based on the affinity of the solutes with the stationary phase and impurities are washed away

• Analytes that are retained on the stationary phase are then eluted from the cartridge and analysed

SPE advantages over LLE

• Smaller volumes of solvent and sample are required

• Processing times are much faster

• The process allows some steps to be automated but in most cases the entire process could not be automated.

• Requires a transfer step before sample introduction into analytical equipment

How Similar are SPE and MEPS?

• MEPS and SPE function in the same manner and use the same phases

• MEPS and SPE are physically very similar

• Methods created for SPE simply require scaling down of the volumes used for use with MEPS

MEPS Advantages

• MEPS combines the sample preparation of SPE with the sample injection capability of autosamplers

• Uses 10 - 100x less solvent and sample

• Increases through put from 10 to 15 minutes per sample to 1 to 2 minutes

• Ability to concentrate the analytes onto the sorbent material

MEPS Application benefits

• MEPS allows existing SPE methodology and applications to be applied to small sample volumes.

• Provides labs a method for priority handling for critical samples.

• Samples maybe concentrated by cycling multiple aliquots through the sorbent bed

• Ability to automate SPE without requiring expensive dedicated on-line systems

MEPS Product

MEPS Products available

• Volumes– 100µL for CTC, Thermo, HTA, Varian 8400 Autosamplers– 250µL for CTC , Thermo, HTA, Varian 8400 Autosamplers

• Phases– C18 end capped, 45µm, 60Å– C8, 45µm, 60Å– C2, 45µm, 60Å– C8/SCX (cation exchange), 45µm, 60Å– Silica, 45µm, 60Å

MEPS operation

MEPS Animation

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Performance

Performance

Calibration Curves 5-2000 ng/mL

Compound 1 name: BusCoefficient of Determination: 0.996893Calibration curve: 3.55292e-8 * x 2̂ + 0.00177372 * x + 0.00148196Response type: Internal Std ( Ref 2 ), Area * ( IS Conc. / IS Area )Curve type: 2nd Order, Origin: Exclude, Weighting: 1/x, Axis trans: None

0.0 200.0 400.0 600.0 800.0 1000.0 1200.0 1400.0 1600.0 1800.0 2000.0ng0

3.83

Response

Repeated 3x per/day

Assay The coefficients of determination(R2)

Day 1 0.9969Day 2 0.9997Day 3 0.9998

Assay The coefficients of determination(R2)

Day 1 0.9969Day 2 0.9997Day 3 0.9998

Assay The coefficients of determination(R2)

Assay The coefficients of determination(R2)

Day 1 0.9969Day 2 0.9997Day 3 0.9998

Day 1 0.9969Day 2 0.9997Day 3 0.9998

Carry Over

• All classes of compound influence MEPS and conventional SPE chemistry in exactly the same way.

• The small quantity of sorbent used and incorporation of MEPS into automation provides for multiple washes of the bed that are not practical for full scale SPE.

• One paper on this technique states washing 4 times with solvent followed by 4 washes with water reduced carry over to 0.02%

Carry Over and Extraction Efficiency

Comparing MEPS, SPE and SPME

Factor MEPS SPE SPME

Amount sorbent 0.5-2 mg 50-2000 mg thickness 150 mm

Sample prep. time 1-2 min 10-15 min 10-40 min

Usage 100 extractions once 50-70 extractions

Recoveries good good low

Sensitivety good good low

Correlation between MEPS & LLE

0 250 500 750 1000 1250 1500 17500

250

500

750

1000

1250

1500

1750

ng/mL (MEPS)

ng/m

L (P

P)

Good correlation between MEPS & LLE with patient sample,but 1 min vs. 45 min, plus automation with MEPS

Bisulphan concentration

ng/m

L (L

LE)

Market Sectors

• Environmental

– Pesticide – Herbicides – POP’s– Water screening

• Food (liquid), Flavors and Fragrances

– Trace Analysis– POP’s– Essential Oils

Market Sectors

• Pharmaceutical

– Bioanalysis– Drug Metabolism & Pharmacokinetics (DMPK). – High Through-put System

• Important: users in high throughput laboratories will require full support and specialist advice (hand holding) for adoption of MEPS as part of their procedure.

• Clinical

– Urine & Plasma Screening– Drugs of Abuse

Market Sectors

• Forensic– Drugs of abuse (Human and Animal)

• SPE methods have been developed for many drugs of abuse.

– Amphetamine and Barbiturates– Benzodiazepines (Valium®)– Flunitrazepam (Rohypnol®)

– Tetrahydrocannabinol (THC)– Cocaine

Who are users of MEPS?

• Anyone who is currently using any of the following techniques in their lab is a potential customer for MEPS:

– Solid Phase extraction (SPE) – Solid Phase Micro Extraction (SPME) – Liquid-Liquid Extraction (LLE) – Protein Precipitation

Which MEPS Sorbent?

C18 BIN’s are like 5% phenyl GC columns – 70% or more of all analysis are made using a C18 packed bed cartridge. This is because most

samples are water based and the analytes need to be transferred from the water matrix into a

non-polar solvent for analysis.

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Which Sorbent?

• For aqueous based fluid, i.e. water, biological fluids, use a reversed phase ‘BIN’ – C18, C8 or C2

• For non-polar organic solvent based samples, hexane, dichloromethane, chloroform, acetonitrile , use a normal phase ‘BIN’ – Silica

• For aqueous or polar organic eluent samples with basic analytes, i.e. some drugs and amines, use a cation ion-exchange ‘BIN’ – M1 (C8/SCX)

Miniaturising your method

SPE Method MEPS Method

Sorbent C18, (500mg sorbent - 6mL volume) Sorbent C18, (2mg sorbent – 7µL bed

volume)

Conditioning 5mL Methanol, 5mL HPLC grade Water Conditioning 2 x 50µL Methanol, 2 x 50µL

HPLC grade Water

Sample 20mL (4mL/minute) Sample 50uL (10-20uL/sec)

Wash 5mL of HPLC grade water Wash 2 x 50µL HPLC grade water

Elution 1mL Methanol/Water 95:5 (v/v) Elution 25µL Methanol/Water 95:5 (v/v)

Benefits of MEPS

• Extraction and injection combined in one process

• On-Line and fully automated (for LC and GC).

• Reduce the time to prepare and inject samples from hours to minutes

• Sample volumes as small as 3.6µL

• Minimum solvent required is 15µL for elution

• 'BIN' maybe reused 40 to 100 times (or greater)

SGE would like to acknowledge an agreement for the development and commercialisation of MEPS with:

• Mohamed Abdel-Rehim AstraZeneca Södertälje, Sweden

• Lars BlombergDepartment of ChemistryKarlstad UniversityKarlstad, Sweden

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