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http://matcmadison.edu/biotech/ THE MEASUREMENT OF pH

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Page 1: ppt on pH meter

http://matcmadison.edu/biotech/

THE MEASUREMENT OF pH

Page 2: ppt on pH meter

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General Principles

• Convenient way to express hydrogen ion concentration, or acidity

pH = - log[ H+] • Where concentration is expressed in moles/liter

Page 3: ppt on pH meter

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Ph SCALE

• Is logarithmic Example, pH 5.0 solution has ten times

more hydrogen ions than pH 6.0 solution • As hydrogen ion concentration, or acidity,

increases, the pH value decreases

Page 4: ppt on pH meter

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Consider Ph OfPure Water

• The [H+] of pure water is 1 X 10-7 mole/L What is pH?

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pH = - log[ H+]

• The log of 1 X 10-7 is -7• The negative log of 10-7 is -(-7) = 7 • The pH of pure water is 7, which we call

neutral

Consider Ph OfPure Water Cont…

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Questions

• What is the pH of a solution with an H+ ion concentration of 10-4 mole/L?

• What is the pH of solution with an H+ ion concentration of 5.0 X 10-6 mole/L?

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Answer

pH = - log [H+] = - log 10-4 = -(-4) = 4

pH = - log [H+] = - log 5.0 X 10-6 =

-(-5.3) = 5.3

Page 8: ppt on pH meter

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Question

What is the concentration of H+ ions in a solution with a pH of 9.0?

Page 9: ppt on pH meter

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Answer

pH = - log [H+] 9.0 = - log [H+]- 9.0 = log [H+]antilog (-9.0) = 1 X 10-9 mole/L

Page 10: ppt on pH meter

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Design OfpH Meter/ElectrodeMeasuring Systems

• pH meter systems measure hydrogen ion concentration; electrochemical

Page 11: ppt on pH meter

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Page 12: ppt on pH meter

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Animations

• http://www.umd.umich.edu/casl/natsci/slc/slconline/PHM/theory.html

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Characteristics & Types of Electrodes & Ph Meters

• Many types of electrodes and meters• Meters:

Conventional Portable Solid state Microprocessor controlled

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Electrodes

• Heart of the system• Can buy separate measuring and reference

electrodes or• Combination; both electrodes are in one

housing• Combination almost always found in biology

labs But not necessarily right

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Separate Electrodes

• Use them for “difficult” samples that:1. Require different type of glass electrode

or2. Require a different type of junction

a) Junction is small hole from which filling solution slowly flows out of electrode

b) Different types of junctions have different flow rates

c) Tendency to clog

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ReferenceElectrodes

• Two major types: Ag/AgCl and calomel (mercury)

• General purpose usually silver/silver chloride

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• Calomel electrodes are said to be more accurate for measuring pH of Tris buffers

But calomel electrodes cannot be used in solutions above 80o C and contain mercury

• Both types are available in combination electrodes or separately

ReferenceElectrodes Cont…

Page 18: ppt on pH meter

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ProperOperation; Avoiding Error

• Consider electrodes• Consider sample• Consider meter operation

Page 19: ppt on pH meter

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Electrodes

• Maintain electrodes properly Filling Solution. Filling solution, often

saturated KCl or AgCl/KCl1. Refillable electrodes are periodically

filled to nearly the top2. Filling hole must be open to pH and

closed for storage3. Gel-filled electrodes contain gelled filling

solution, never refilled

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Page 21: ppt on pH meter

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Electrodes

• Storage. Consult the manufacturer's instructions

Don’t store electrodes in distilled water New combination electrodes often stored

dry

1. Then are conditioned before use by soaking at least 8 hours in pH 7 buffer

Page 22: ppt on pH meter

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Sample

• Important part of the system• Must be homogenous• May be chemical changes in sample• May be temperature effects

Page 23: ppt on pH meter

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Difficult Samples

• Non-aqueous solvents• High purity water. High purity water does not

readily conduct current and it absorbs CO2 from the atmosphere

• High salt samples. Sample ions compete with the reference filling solution ions

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• Sample-electrode compatibility. Tris buffer, sulfides, proteins, Br-, and I- , can complex with silver in Ag/AgCl electrodes leading to a clogged junction. A calomel electrode is often recommended

• Slurries, sludges, viscous and colloidal samples. Use a fast flowing junction. Keep measuring electrode bulb clean

Difficult Samples Cont…

Page 25: ppt on pH meter

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Operation;Calibration

• User calibrates frequently• Two buffers of known pH; forms calibration

line. Try this in lab• Older pH meter first calibration buffer should

be pH 7.00 For acidic samples, second is pH 4.00 For basic samples, second is pH 10.00

Page 26: ppt on pH meter

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• Newer, microprocessor-controlled meters: Any two standard buffers that bracket pH of

samples may be used Some microprocessor-controlled meters

allow use of more than two standard buffers

Operation;Calibration Cont…

Page 27: ppt on pH meter

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The Relationship Between pH andMillivolt Readings

The X axis is pH and the Y axis is millivolts

Page 28: ppt on pH meter

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Calibration Buffers

• Accuracy of pH meter depends on calibration buffers

• Some buffers react with CO2 from air Keep buffer containers closed Throw away buffer after use pH 10.0 buffer is particularly sensitive to

CO2

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Buffers

• For best accuracy, do not use buffers after their expiration date

• Avoid contamination• pH of a buffer will change as its temperature

changes

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Temperature And pH

• Temperature has two important effects: Measuring electrode's response to pH is

affected by the temperature pH of solution that is being measured

change as its temperature changes

Page 31: ppt on pH meter

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Effect of Temperature on pH Readings

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CompensationFor Temperature

• Meter needs to "know" temperature of solution• Can measure solution temperature with

thermometer and "tell" pH meter• Alternatively, use ATC (automatic temperature

compensating) probes that is connected to the pH meter

• Compact devices may have a temperature probe built into the electrode housing

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Temperature

• The pH of some solutions changes with temperature

• Usually measure pH of samples at the temperature at which they will be used

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A ConventionalMethod or Measuring pH

1. Warm-up meter

2. Open filling hole; check filling solution level

3. If meter has a "standby” mode, use it when the electrodes are not immersed in sample

Page 35: ppt on pH meter

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4. Calibrate each day or before use

A. Adjust temperature setting.

B. Rinse electrodes and blot dry.

C. Don’t wipe.Immerse electrodes in pH 7.00 calibration buffer. Junction must be immersed and level of sample must be below filling solution level. Allow reading to stabilize.

A ConventionalMethod or Measuring pH cont…

Page 36: ppt on pH meter

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D. Adjust meter to read 7.00

E. Remove electrodes, rinse, blot dry.

F. Place electrodes in second standardization buffer Adjust meter to pH. Remove, rinse, and blot.

A ConventionalMethod or Measuring pH cont…

Page 37: ppt on pH meter

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G. Older meters: Recheck pH 7.00 buffer and readjust as necessary. Recheck second buffer and readjust as necessary.

H. Readjust as needed up to three times. If readings are not within 0.05 pH units of what they should be, electrode probably needs cleaning.

A ConventionalMethod or Measuring pH cont…

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Quality Control

5. Optional: Quality control checks.Try this in lab.

A. Linearity Check. Take reading of a third calibration buffer. For example, if you calibrated with pH 7.00 and 10.00 buffers, check pH 4.00 buffer. B. Allow the reading to stabilize and record value. Do not readjust meter. If reading is outside proper range, service electrodes.

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Quality Control

C. Check pH of a control buffer whose pH is known and that has a pH close to the pH of the sample.

Common to set maximum allowable error of control buffer to + 0.10 pH units.

Do not adjust meter to pH of control buffer.

If pH of control buffer is not within required tolerance, service electrodes.

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Sample

6. Set meter to temperature of sample or use ATC probe.

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7. Place electrodes in sample; allow reading to stabilize.

A. Wait too long, pH of some samples will change due to air, chemical reactions, etc.

B. Difficult solutions may require longer to stabilize.

C. Many new pH meters have an “autoread” feature.

Sample

Page 42: ppt on pH meter

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9. Record all relevant information.

10. Remove electrodes from sample, rinse and blot, store properly with filling hole closed (refillable electrodes only).

Sample

Page 43: ppt on pH meter

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Iv Trouble-shooting

• First step is to know you have trouble.• Symptoms of pH system problems include:

Reading drifts and won't stabilize. Reading fluctuates. Meter cannot be adjusted to both

calibration buffers. pH value for a buffer or sample seems

wrong.

Page 44: ppt on pH meter

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Trouble-shooting Tips

• Look for and correct simple (embarrassing) mistakes:

Electrode measuring bulb and junction are not immersed in sample.

Meter is not turned on or plugged in, or the electrode cables are not connected to meter.

Page 45: ppt on pH meter

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Trouble-shootingTips Cont.

• Reference electrode is not filled with electrolyte.

• Reference electrode filling hole is closed.• Sample is not well-stirred.• Calibration buffers are not good.• Electrode is cracked or broken.

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Next

• What is most likely cause of problems?• Problems can arise in:

The reference electrode The measuring electrode The calibration buffers The sample The meter

Page 47: ppt on pH meter

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Reference Electrode

• Reference electrode junction most common source of problems.

• Occluded junction causes long stabilization time; reading drifts slowly towards correct pH.

• If junction is completely plugged, reading may never stabilize.

This is also caused by broken electrodes and by some problems within meter.

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• Slow equilibration also caused by changes in sample temperature, reactions in sample, or sample-electrode incompatibility.

Reference Electrode cont…

Page 49: ppt on pH meter

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• Poor calibration buffers will cause inaccuracy. This may be detected if solution of known pH is checked.

• If sample is not homogenous, or if its temperature is unstable, then pH readings will fluctuate or drift.

• Difficult samples may be slow to equilibrate or give incorrect readings.

Reference Electrode cont…

Page 50: ppt on pH meter

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• Complete lack of response likely caused by problems with meter, but meter is least likely component to cause problems.

• To check which component is at fault substitute in a new reference or combination electrode.

Reference Electrode cont…