Capillary Electrophoresis Vanessa CHEM6200

7/25/2019 Capillary Electrophoresis Vanessa CHEM6200

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CAPILLARY

ELECTROPHORESIS:

TECHNIQUE AND

APPLICATIONBy Vanessa Hobbs


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CAPILLARY ELECTROPHORESIS


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DEVELOPMENT OF CAPILLARYELECTROPHORESIS

1803 F.F. Reuss Clay Slab

1886 O. Lodge Zone Electrophoresis

1937 A. Tiselius Electrophoretic Cell

1967 S. Hjerten Rotating tubes (300 um)1970 V. Neuhoff PAG filled tubes

1979 Mikkers, Everaerts, VerheggenFZE

1981 Jorgenson and Lukags 75 um cap.

1983 Micellar Electrokinetic Chrom.

1988 Commercial introduction


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ELECTROOSMOTIC FLOW

http://www.electrokinetic.co.uk/images/tech1.gif


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MOVEMENT OF ANALYTEAnalyte = E

= velocity = electrophoretic mobility E = Electric field

Electrophoretic mobility

= q/[6r] q = charge = solutio viscosity r = radius

Electroosmotic flowE!"= [#/$]%E

# = dielectric &ostat % = 'eta potetialFlow of migration = [()E!* )e+,]/-

, = potetial - = legth of capillary

Forensic Science International77 (1996) 211 - 229


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INJECTION OF SAMPLE

Current Analytical Chemistry. 2005,1http://www.calstatela.edu/dept/chem/gomez/pubs-pdf/flow-injection.pdf


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INJECTION OF SAMPLE

Injection is difficult due to sample size

Electrokinetic InjectionDiffers by analyte

HydrodynamicMany parameters

Anal. Chem., 1997, 69 (15), pp 29522954


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INJECTION OF SAMPLES

Anal. Chem.2001, 73,1974-1978


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INJECTION OF SAMPLE

Current Analytical Chemistry. 2005,1http://www.calstatela.edu/dept/chem/gomez/pubs-pdf/flow-injection.pdf


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CAPILLARY ZONEELECTROPHORESIS

Separated by mass to charge ratio Based on Electroosmotic Flow Detectors:

UV Detector Beers Law

Laser Fluorescence Deriv. MS - electrospray Chemiluminescence Diode Array Detector Indirect

Refractive Index Compare with HPLC and GC Neutral Compounds Chiral Compounds


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INCREASING PATH LENGTH

http://www.chem.agilent.com/Library/technicaloverviews/Public/5989-


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MICELLAR ELECTROKINETICCHROMATOGRAPHY

UVNeutral compoundsComprable to HPLC


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Capillary Electrochromatography

Packed column with no pressure applied, onlyelectroosmotic pressure.


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CAPILLARY GELELECTROPHORESIS

Crosslinked vs. non crosslinked

DNA sequencing

Protein analysis

Chirality possibleEOF less desirable


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CAPILLARY GELELECTROPHORESIS

http://www1.qiagen.com/Images/Catalog/2134.jpg


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CAPILLARY ISOELECTRICFOCUSING

http://www.targetdiscovery.com/~tdidocs/App_Note_5_200405.pdf


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CAPILLARY ISOELECTRICFOCUSING

pH gradient

Sample focusing and detection

Movement of gradient towards the detector

Zone broadeningNot useful for chiral compounds


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CAPILLARY ISOTACHOPHORESIS

Two buffers form ionic zones

Anions and Cations seperately

Neutral compounds

Used for concentrationEOF less desirable


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APPLICATIONSCE and Analysis of Illicit Drugs


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HPLC Heroin Analysis


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HPLC Analysis of Heroin (SPE)

Fig. 2. (a) Representative total ionchromatograms of all quantifiable

analytes spiked at LLQ level in

human plasma ( ng!mL). "he

intensity of the deuterated analytes

#as above 2$$ %cps&. (b)

Representative total ionchromatograms of random chosen

patient' plasma sample. (c) "otal ion

chromatogram of a plasma sample

of a nondrug using volunteer. ()

*+, and *+,d+- () morphine

and morphined+- (/) *0,- (1) 0**- () heroin and heroind0-

(F) 3 methadone and methadone

d4- (,) *15- (6) cocaine- (7)

ben8oylecgonine.

1791 RR: ;1 "R75L *

Capillary Electrophoresis Vanessa CHEM6200

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