ELECTROCHEMISTRY

IN CLINICAL CHEMISTRY

BASIC CONCEPTS, INSTRUMENTATION

AND APPLICATION

By

Dr. BASIL, B MBBS (Nigeria),

Department of Chemical Pathology/Metabolic
Medicine,
Benue State University Teaching Hospital,
 OUTLINE
Introduction
Electrochemical Cells
Potentiometry
Ion-Selective Electrodes
Types
Basic concepts
Instrumentation
Preanalytical Considerations
Quality Control
Merits and Limitations
Applications
Voltametry/Amperometry
Coulometry
Conductometry
 INTRODUCTION
Electrochemistry involves the
measurement of electrical signals
associated with chemical systems.
Electrochemical analyses used in the
clinical laboratory include: Potentiometry,
Amperometry/Voltametry, Coulometry and
Conductometry
The two basic electrochemical cells
involved in these analyses are Galvanic
and Electrolytic cells
 ELECTROCHEMICAL CELLS

Consist of two half-cells and a salt bridge saturated with

electrolytes, or in a solution which serves as the salt bridge.
Electrochemical activity of one half-cell cannot be measured,
both must be coupled, and one compared with the other
To rate a half-cell reaction, a specific electrode reaction is
arbitrarily assigned 0.00V.
Any other reaction coupled to it is either positive or negative
depending on the relative affinity for electrons
The electrode defined as 0.00V is the standard hydrogen
electrode: H2 gas at 1atm.
 The hydrogen electrode coupled with a Zn half-cell is cathodic,
and with Cu half-cell is anodic
The potential generated by the hydrogen-gas electrode is used
to rate the electrode potential of metals in 1mol/L solution
GALVANIC CELL:
There is spontaneous flow of electrons from the electrode with
the lower electron affinity (oxidation e.g silver) as the
electrodes are connected
The electrons pass through the external meter to the cathode
(reduction), where OH- are liberated
This reaction continues until one of the chemical components
is depleted, thus, the cell becomes dead and cannot produce
electrical energy to the external meter
ELECTROLYTIC CELL:
When an external electromotive force, E, is applied > electrons
will flow in the reverse direction
Thus a Galvanic cell can be built from an Electrolytic cell by
turning off the external E
 POTENTIOMETRY
Measurement of the electrical potential difference btw two
electrodes in a galvanic electrochemical cell
These electrodes Reference and Indicator(measuring)
electrodes generate an EMF when the cell current is zero
The EMF is measured using a potentiometer e.g pH meter
which does not alter the cells composition or draw current
One of the electrolyte solutions is the sample containing
the analyte(s) of interest can be replaced by a reference
solution for calibration purposes
The potential gradients have been classified as
Redox potentials, Membrane potentials or Diffusion potentials
The cell can be devised such that all the gradients are kept
constant except one which is related to the activity a
specific ion of interest
The activity or EMF generated is then related to the
concentration of the ion (analyte) in the solution (sample).
Types of potentiometric electrodes used for clinical
applications include:
Redox electrodes
Ion-selective electrodes, and
pCO2 electrodes

Schematic diagram of an electrochemical cell of

 ION-SELECTIVE
PotentiometricELECTRODES
methods of analysis involve the direct
measurement of electrical potential due to the
activity of free ions
ISEs are designed to be selectively sensitive toward
individual ions
The potential produced at the membrane-sample-
solution interface is proportional to the log of the
ionic activity or concentration
Ion-Selective membranes typically consist of glass,
crystalline, or polymeric materials
The chemical composition of the membrane is
designed to achieve an optimal permselectivity
toward the ion of interest
Other ions exhibit slight interaction with membrane
sites and display some degree of interferences
Clinically, a correction is required when the
 TYPES OF ISE:
Ion-selective membrane is the heart of the ISE as it controls
the selectivity of the electrode. They include:
Inert metal electrodes in contact with a Redox couple e.g
Standard Hydrogen electrode
Metal electrodes participating in a redox reaction, e.g Ag/AgCl
electrode
and Membrane electrodes
The membrane can be solid (e.g glass), liquid (e.g ion-
exchange electrodes), or special membrane (e.g compound
electrodes) such as Gas-sensing and Enzyme electrodes
The ISEs that are commonly used in clinical chemistry are
the membrane (esp Glass and Polymer) electrodes.
GLASS MEMBRANE ELECTRODE:
Used to measure pH and Na+ and as an internal transducer
for pCO2 sensors
They are formulated from melts of Si and/or Al-oxide mixed
with other oxides
 By varying the glass composition, electrodes with selectivity
for H+, Na+, K+, Li+, Rb+, Cs+, Ag+, Tl+, and NH4+ have
been produced with only those of H+ and Na+ having
sufficient selectivity
A Glass ISE universally used in the clinical lab is the pH
electrode
The pH Electrode (Prototype):
Basic components of the pH meter include:
Indicator Electrode:
Silver wire coated with AgCl immersed into an internal solution
of 0.1mmol/L HCl, and placed into a tube containing special
glass membrane tip
The membrane is only sensitive to H+ and consist of specific
quantities of Li, Cesium, La, Ba, or Al oxides in silicate
When placed into the sample solution, H+ move to the tip of
the electrode and produce a potential difference btw internal
solution and the sample solution which is measured as pH
The glass continually dissolves very slowly from the surface
(lasts for several yrs)
 Ionic exchange occurs only in the gel layer there is
no penetration of H+ through the glass
It is highly selective for H+, but other cations in high
concentration, esp Na+, can interfere
Reference Electrode:
Commonly used is the calomel electrode.
Calomel a paste of mainly Hg2Cl2 in direct contact
with metallic Hg in an electrolyte solution of KCl
At constant electrolyte conc. and temp, a stable
voltage is generated at the interphase of the mercury
and its salt
A filling hole is needed for adding KCl and a tiny
opening at the bottom for completion of electric
contact btw the 2 electrodes
The liquid junction consist of a fiber or ceramic plug
that allows a small flow of electrolyte filling solution
 Hg/HgCl as a reference electrode is
slow to reach stable voltage following voltage change, and
unstable above 80oC
Ag/AgCl as a reference electrode
can be used at high temp up to 275oC, and
the AgCl-coated Ag wire make a more compact electrode than Hg
In measurements where Chloride contamination must be
avoided, Hg-sulfate and K-sulfate reference electrode can be
used
Reference electrodes generally consist of a metal and its salt,
in contact with a solution containing the same anion.
Liquid Junctions:
Allows a slow flow of electrolyte from the tip of the reference
electrode, thus, establishing electrical connection btw the
indicator and reference electrode
A junction potential set up at the boundary btw two different
solutions because of positive and negative ions diffusing
across the boundary at unequal rates
This may increase or decrease the potential of the reference
electrode, thus, it should be kept at a minimum reproducible
value when the reference electrode is in solution
Readout Meter:
Zero potential indicates that both electrodes are generating
same voltage (assuming there is no liquid junction potential)
Isopotential temp change has no effect on the response of
the cell
Achieved by making midscale (pH 7.0) correspond to 0V at all
temps via the use of an internal buffer that alters with
response to temp changes
Nernst equation:
Describes the EMF generated by H+ at the glass tip
= pH x (RT ln 10)/F = pH x 0.059V
Where, = EMF, F = Faraday constant, R = molar gas constant, and T
= temp in kelvin
As temp increases, H+ activity increases and the potential
generated increases
Most pH meters are manufactured for greatest accuracy in the
10 60oC range
 pH Combination Electrode:
Has both indicator and reference electrodes in one
small probe
Consists of Ag/AgCl internal reference electrode
sealed in a narrow glass cylinder with a pH-sensitive
glass tip

LIQUID MEMBRANE ELECTRODE (POLYMER

MEMBRANE ISE):
Porous membrane impregnated with solution (2-
ethylhexyl phosphoric acid dissolved in dioctyl
phenyl phosphate) and mounted at the end of an
electrode body
This was later modified into a plasticized poly(vinyl
chloride) membrane Polymer Membrane ISE it is a
more rugged and convenient to use.
 They are the dominant class of potentiometric
electrodes used in modern clinical analysis
instruments
Calcuim ISE:
Dioctylphenyl phosphate (an ion-selective carrier)
dissolved in an inert water-insoluble solvent, diffuses
though a porous membrane
Since the solvent is insoluble in water, test samples
cannot cross the membrane but Ca2+ are exchanged
Potassium ISE:
Use antibiotic valinomycin (major breakthrough),
which show great selectivity for K+ over Na+, as the
carrier
The K+ ISE based on valinomycim is extensively used
today for the routine measurement of K+ in blood and
urine due to its wide linear range of over three orders
of magnitude
 Are recharged every few months to replace the liquid ion
exchanger membrane and the porous membrane
Interference from other cations, seen as deviation from
linearity, is not apparent at K+ activities >10 -4 mol/L
Other less selective polymembrane ISEs (Mg2+ and Li+) are
subject to interferences from Ca2+/Na+ and Na+
(respectively)
SPECIAL MEMBRANES:
Gas-Sensing Electrodes:
Similar to pH electrodes but are designed to detect specific
gases (e.g CO2 and NH3) in solutions and are usually
separated from the solution by a thin, gas-permeable
hydrophobic membrane
CO2 permselectively diffuses into a thin film of NaCO3 soln
The diffusion changes the pH of the soln and that change
is detected by a pH electrode within it
pCO2 electrode = pH electrode + CO2 membrane
 In the NH3 gas electrode, the NaCO3 soln is replaced
by NH4Cl
Other gas-sensing electrodes function on the basis of
an amperometric principle e.g pO2, Glucose, and
Peroxidase
Glucose determination is based on reduction in pO2 during
glucose oxidase reaction with glucose and O2
Peroxidase electrode is dependent on oxidation of H 2O2 to
release O2 which is detected
Enzyme Electrodes:
ISE covered with immobilized enzymes that can
catalyze a specific chemical reaction and the reaction
product detected.
They include:
Urease in Urea electrode with an ISE selective for NH3 or
NH4+
Glucose oxidase used in combination with a pH electrode
The pCO2 electrode
 PRE-ANALYTICAL AND
ANALYTICAL CONSIDERATIONS
SAMPLE COLLECTION:
Blood Samples:
Serum or plasma specimen should be collected in
heparinised bottles
Capillary blood (microsample tubes, capillary tubes or
directly from fingerstick) to some POC devices
Heparinized whole blood (arterial or venous) for blood gases
should be read promptly
Samples should be centrifuged unopened and the plasma
separated immediately
Grossly lipaemic samples should be ultra centrifuged before
analysis
Urine Samples:
Assessment of urine sodium, potassium or chloride
Samples for gases should be layered with oil
 Faeces, aspirates and fluids drained from portions of
the GIT can be analyzed for electrolytes
ANALYTICAL VARIATIONS:
Interferences from undesired ions (most serious
problem limiting ISEs) no ISE is completely ion
specific
Sensitive to other ions having similar properties e.g.
Nitrate electrode may have ionic interferences from
perchlorate, iodide, chloride and sulfate
Blockage of the selective membranes with time by
repeated coating with proteins
Leakages in the membranes would lead to
reduced/lack of analytical selectivity
Exposure of electrodes and galvanometer to cold
temperatures would reduce sensitivity
 QUALITY CONTROL
Periodic changing of the selective
membranes
Ensure appropriate temperature control
protocols
Ensure adequate and appropriate power
supply
Ensure MANUAL is read thoroughly by
all USERS
All mechanical faults should be promptly
reported to the appropriate channels
Equipment should be recalibrated
 MERITS AND DEMERITS OF
Advantages:
POTENTIOMETRY
Relatively inexpensive and simple to use and have an
extremely wide range of applications and wide concentration
range.
They can be used very rapidly and easily.
ISEs can measure both positive and negative ions.
They are unaffected by sample colour or turbidity.
Non-destructive: no consumption of analyte.
Non-contaminating.
Short response time: in sec. or min. useful in industrial
applications.
Limitations:
Precision is rarely better than 1%.
Electrodes can be affected by proteins or other organic
solutes.
Interference by other ions.
Electrodes are fragile and have limited shelf life.
 APPLICATIONS OF POTENTIOMETRY
Clinical Chemistry
Ion-selective electrodes are important sensors for clinical
samples because of their selectivity for analytes.
The most common analytes are electrolytes, such as Na +,
K+, Ca2+,H+, and Cl-, and dissolved gases such as CO2.
Environmental Chemistry
For the analysis of CN-, F-, NH3, and NO3- in water and
wastewater.
Potentiometric Titrations
pH electrode used to monitor the change in pH during the
titration.
For determining the equivalence point of an acidbase
titration.
Possible for acidbase, redox, and precipitation titrations, as
well as for titrations in aqueous and non-aqueous solvents.
 Agriculture
NO3, NH4, Cl, K, Ca, I, CN in soils, plant
material, fertilizers.
Detergent Manufacture
Ca, Ba, F for studying effects on water quality
Food Processing
NO3, NO2 in meat preservatives
Salt content of meat, fish, dairy products,
fruit juices, brewing solutions.
F in drinking water and other drinks.
Ca in dairy products and beer.
K in fruit juices and wine making.
Corrosive effect of NO3 in canned foods.
 OTHER ELECTROCHEMICAL TECHNIQUES

VOLTAMMETRY:
Measures current in an electrochemical cell as a function
of the applied potential
Assortment of electrodes allows for a very specific
analysis of different chemicals
Types:
Polarography, Hydrodynamic voltammetry, Stripping
voltammetry, and Amperometry
Amperometry:
apply a constant potential to the electrode and measure the
resulting current.
most often used in the construction of chemical sensors for the
quantitative analysis of single analytes.
Example is the Clark O2electrode, which responds to the
concentration of dissolved O2in solutions such as blood and
water.
COULOMETRY:
Measures electrical charge passing btw two electrodes
Based on Faradays law:
the total charge passed during an electrolysis is proportional to the
amount of reactants and products in the redox reaction
If the electrolysis is 100% efficient (i.e only the analyte is
oxidized or reduced), then we can use the total charge or
current to determine the amount of analyte in a sample
Types:
In controlled-potential coulometry a constant potential is applied
and the resulting current is measured as a function of time
In controlled-current coulometry current is held constant and the
time required to completely oxidize or reduce the analyte is
measured
Gold standard for determination of chloride in serum/plasma
It is also a mode of transduction for some biosensors
Interferences: anions with greater affinity for Ag + e.g Br-
CONDUCTOMETRY:
Determines the quantity of an analyte in a mixture by
measuring its effect on the electrical conductivity of the
mixture
The current is directly proportional to the solutions
conductance
Used in the measurement of the volume fraction of
erythrocytes in whole blood (hematocrit)
They are insulators because of their lipid-based membrane
composition
Interference: abnormal protein conc and insufficient mixing of
sample will change plasma conductivity
Applied in the electronic counting of blood cells in
suspension Coulter principle
Cells are forced through a tiny orifice flanked by two electrodes
(btw which a constant current is established) which generate
signal (increased resistance) with the passage of cells between
them
The pulses are amplified and counted
 REFERENCES
Tietz Textbook of Clinical Chemistry and
Molecular Diagnostics.
Clinical Chemistry; Principles, Techniques
and Correlations, 7th ed., by Bishop et al.
http://chemwiki.ucdavis.edu/Analytical_Ch
emistry/Analytical_Chemistry_2.0/11_Elect
rochemical_Methods/11E
%3A_Summary_and_Problems
http://www.cma-science.nl
www2.vernier.com/booklets/ise.pdf
http://www.sfu.ca/chemistry/groups/Li/che
m215/selective.PDF