pH MEASUREMENT

• The chemical balance in the body can be determined by the pH value of blood and other
body fluids.

• pH is defined as the hydrogen ion concentration of a fluid.

• It is the logarithm of the reciprocal value of h+ concentration.

pH= - log10 [H + ] = log10 1/[H+ ]

• pH is the measure of acid- base balance in a fluid,

• A neutral solution has the pH value as 7. Solutions with pH value less than 7 are acidic
and above 7 are basic. Most of the body fluids are slightly basic in nature.
pH Electrode
• For easier ph measurement combination electrodes are used.

• In this type both the active glass electrode and reference electrode are
present in the same meter.

• The glass electrodes are suitable only to measure ph values around 7.

• After every measurement the pH meter is washed with 20%

ammonium biflouride solution, for accurate results.

• The Ph meter with hydroscopic glass absorbs water readily and

provides best pH value.
 pO2 MEASUREMENT
• po2 is defined as the partial pressure of oxygen respectively.

• The effective functioning of both respiratory and cardiovascular

system can be by po2 measurement.

• The partial pressure of a gas is proportional to the quantity of that gas

present in the blood.
pO2 Electrode
The platinum wire, which is an active electrode, is embedded in glass for
insulation and only its tip is exposed.

It is kept in the electrolyte solution in which the oxygen is allowed to diffuse.

The reference electrode is made up of silver-silver chloride (Ab/AgCl).

A voltage of 0.7 is applied between the platinum wire and the reference
electrode.

The negative terminal is connected to the active electrode through a micro

ammeter and the positive terminal is given to the reference electrode.
Due to the negative terminal, the oxygen reduction takes place at the
platinum cathode.

Finally the oxidation reduction current proportional to the partial

pressure of oxygen diffused into the electrolyte can be measured in
the micro ammeter.

The electrolyte is generally scaled in the electrode chamber by means

of a membrane through which the oxygen can diffuse from the blood
or sample solution
• Two types of pO2 measurement.

 Vitro measurement

 Vivo measurement

Vitro Measurements
 In this method the blood sample is taken and the measurement for oxygen saturation is
made in the laboratory. The electrode is placed in the sample blood solution and the
pO2 value is determined.

In Vivo Measurements

 In this method the oxygen saturation is determined while the blood is flowing in the
circulatory system
 pCO2 MEASUREMENT
 pco2 is defined as the partial pressure of carbon dioxide.

 The partial pressure of a gas is proportional to the quantity of that gas present in the blood.

 The partial pressure of carbon dioxide can be measured with the help of pCO2 electrodes.

 Since there is a linear relationship between the logarithm of pCO2 and pH of a solution.

 The pCO2 measurement is made by surrounding a pH electrode with a membrane selectively

permeable to CO2.

 The modern improved pCO2 electrode is called as severinghous electrode.

pHCO3 Measurement
pCO2 Electrode
• In this electrode the membrane permeable to CO2 is made up of Teflon which is not

permeable to other ions which affects the pH value.

• The space between the Teflon and glass contains a matrix layer which allows only the

CO2 gas molecules to diffuse through it.

• The demerits in older CO2 electrode is, it requires a length of time for the CO2

molecules to diffuse through the membrane.

• The modern CO2 electrode is designed in such a way to overcome this demerit. Here

the CO2 molecules diffuse rapidly through the membrane and the measurement can

be done easily
Blood gas analyzers are used to measure the content of pH, pCO and
PO  from the blood
2

Two gases of accurately known O  and CO  percentages are required

2 2

for calibrating the analyzer in pO  and pCO  modes.

2 2

These gases are used with precision regulators for flow and pressure
control.
• Two standard buffers of known pH are required for calibration of the analyzer in the pH mode.
• Input signal to the calculator is obtained from the outputs of the pH and pCO2 amplifiers
• The outputs are adjusted by multiplying with a constant and are given to an adder circuit
• The output of adder is passed to antilog generators circuit. Then it is passed to A/D converter for
display. Resistance R is used to adjust zero at the output.
• Total CO2 is calculated by summing the output signals of the calculators and the output of the pCO2
amplifier
Base excess calculator consists of three stages.
 
 In the first stage, the output of pH amplifier is inverted in an operational amplifier, whose gain is
controlled by a potentiometer.

 The output of HCO3—calculator is inverted in the second stage.

 The third stage is a summing amplifier A3 whose output is given to A/D converter, that gives a
digital read out.
 CALORIMETER
• Measures the color concentration of a substance in a solution by
detecting the color light intensity passing through a sample containing
the substance and a reagent

• Optical color filters are used to detect the color wavelength of interest.
E.g., urine passes yellow light and absorbs blue and green

• Laser LEDs are preferred if their wavelength is suitable due to purity of

the monochromatic color
CALORIMETER
• A solution of concentration ‘c’ is placed in a cuvette with a length of light path “L”.
Light ray of particular wavelength is passed through a filter F.
• Io is the initial intensity of the light entering the cuvette. With the part of
intensity being absorbed in the solution the light leaving the cuvette has a lower
intensity, I1. The transmittance is

• The light leaving the second cuvette has the intensity I2 which is lower than that
of I1. The light is transmitted through successive cuvette decreases om the same
manner. The absorbance can be measured
 PHOTOMETER
• Measures the color intensity of a flame supported by O2 and a
specific substance.
• Sample’s emission of light is measured (rather than the absorbance of
light).
• Typically used to determine the conc. of pure metals and/or Na+, K+,
Li+ and Ca++
• In this method, fine droplets of the sample is aspirated into gas flame that burns in a chimney.

• A known amount of lithium salt is added to the sample, as a reference. As a result, red light is
emitted by the lithium and yellow and violet beam are emitted due to sodium and potassium
respectively.

• These diffracted colors are made to incident on photodiodes. The photo detector circuits consists of
a reverse biased diode in which the current flow increases as intensity of incident light increases.

• A calibration potentiometer is used in every channel. Since the lithium is used as a standard
reference, the output of sodium and potassium channel are calibrated in terms of differences with
the known lithium.

• The output can be compared with the spectral illustration.

SPECTROPHOTOMETER
• Depending on the substance being measured, the wavelength used is typically in the ultraviolet (200-400 nm), visible (400-700nm)

or infrared (700 to 800 nm) range.

• Spectrophotometer can be used to determine the entity of an unknown substance, or the concentration of a number of known

substances. The type of source / filters used typically determines the type of the spectrophotometer.

• Rays of light bend around sharp corners, where the amount of bending depends on the wavelength. This results in separation of

light into a spectrum at each line.

• In spectrophotometer, selection filter of colorimeter is replaced by a monochromator. Monochromatic uses a diffraction grating G

to disperse light from the lamp. Light falls through the slit S0 into its spectral components.

• Slit S1 is used for selecting a narrow band of the spectrum which is used to measure the absorption of a sample in the cuvette.

• The light from the cuvette is given to photo detector. It converts light into a electrical signal. This electrical signal is amplified by

using an amplifier. The output from the amplifier is given to meter which shows absorbance.

• Light absorption is varied when the wavelength is varied. Mirror M is used to reduce the size of the instruments.
AUTO ANALYZER