522 RECUEIL 74 (1955)

545.37 ; 546.13 I POTENTIOMETRIC TITRATION OF CHLORIDES IN

BIOLOGICAL FLUIDS BY

ARTHUR JORDAN, (Sheffield).

The object of the work was to develop a method for the direct titration of chlorides in biological fluids using silver nitrate, the end-point being detected by means of a direct-reading potentiometer. This has been achieved.

The principle of potentiometric titrations is well-known. The curve ordinarily obtained is illustrated in fig. 1. The abscissa

rerepresents the amount of silver nitrate solution added: the ordinate represents the potential - expressed in milli-volts - between the titration half-cell and a calomel half-cell. A silver electrode dips into the titration half-cell and a sulphate bridge joins the two half-cells. The titration half-cell must contain enough acid to prevent precipitation of phosphates or of protein. These determinations are very accurate but are tedious and time-consuming.

It was pointed out by Jones 1) that the calomel half-cell could be replaced by a reference half-cell composed of chloride exactly titrated by silver nitrate, whereupon the end-point of the titration would be indicated by the disappearance of any potential difference between the two half-cells. Jones demonstrated an apparatus permitting titrations using this principle. Accurate measurement of potential differences became unnecessary and a galvanometer was used to appreciate the end-point. In fact Jones used two galvanometers: a less sensitive one for the early part of the titration and a sensitive galvano- meter for the exact end-point. Polarisation was avoided by having a push button in the circuit, which had to be depressed to make a reading after each addition of silver nitrate. The electrolyte bridge consisted of a single glass tube plugged at each end with cotton wool. Stirring was effected with a glass rod by hand.

1) E. R. Jones, Demonstration to the Association of Clinical Pathologists, beds, July 1950.

Pofenfiornefric fitration of chforides, efc. 74 (1955) RECUEIL 523

Jon,e.s's technique had several disadvantages. The reading was not continuous which made it more time-consuming. The electrolyte bridge had to be cleaned and prepared after each titration. Stirring with a glass rod proved slow and inadequate.

.

I 1 3 4 5 6 7

r n t AgNo, N/SO

Fig. 1 . Curve of a normal potentiometric titration. The end-point corresponds to the point on the curve where the increment of mV is maximal for a given

addition of silver nitrate.

524 Arthur Jordan,

It was obvious that some from of efficient, continuous stirring was essential. Various methods were tried, such as overhead stirring and rotating the titration vessel. Ultimately a magnetic stirrer was found to be the simplest and most effective.

The need to clean the electrolyte bridge was obviated by the use of the simple device shown in fig. 2, which represents one end of the

FROM RtslIRVOlR AND SALT BRIDG*

PAWFFIN WAX. Tb WASTE-

Fig. 2. Terminal part of electrolyte bridge. At the end of each titration fresh elctrolyte from the reservoir passes down the polythene tube and displaces

the electrolyte in contact with the sintered glass filter disc.

electrolyte bridge: the other end is similar. The two are joined by a glass tube which also communicates by means of a T-piece with a tap funnel which acts as a reservoir of electrolyte. At the end of each titration the outside of each end of the bridge is rinsed with distilled water from a wash-bottle, and the electrolyte in contact with the sintered glass plate is run to waste by opening the tap of the reservoir

Potentiornettic titration of chlorides. efc. 74 (1955) RECUEIL 525

I I

of electrolyte. When the tap of the reservoir is opened, electrolyte passes down the fine polythene tube onto the sintered glass filter disc and then passes upwards outside the polythene tube and so to waste.

Various .possible electrolytes were studied for use in the electrolyte bridge. Some gave more stable potentials than others. Some diffusion must occur at the sintered discs and it is possible that interface potentials might perhaps account for the lack of stable potential differences. Electrolytes containing nitric acid were noticed to give unstable potentials. Saturated potassium sulphate in water was found to be most satisfactory. It has the great disadvantage of "creeping" between glass . and rubber, and coating everything with a layer of crystals.

Titrations are carried out with N/50 AgN03. To follow the progress of the titration a mirror galvanometer in

series with a resistance was used initially (fig. 3 ) . With the galvano-

.--I I I

potentiometric titrations. The resistance should be of the order of half to one million ohms.

meter used, 1 pamp. corresponded to 126 mm scale deflection. With this arrangement, satisfactory titrations can be carried out although some polarisation occurs. Some idea of the extent of the polarisation is given by fig. 4. The galvanometer was in series with a resistance of 470000 D and the output was measured without any addition of silver nitrate. As titrations are completed in about two minutes and furthermore, the progress of the titration reduces the potential, and hence the current, and so delays the onset of ,polarisation, the curve

526 Arthur Jordan,

cat. be,

LH

I00

60

10

20

shown in fig. 4 suggests a less satisfactory state of affairs than prevails in practice. Nevertheless, it seemed that a method which took virtually no current would be desirable. This would be particularly so if very small quantities of chloride were to be titrated.

CURREFIT civrr~’ 6I I mC S E a w i~ 10 mt Marit i 3 m f

WNO, 4 ntru SILVER aucruMm, K ~ S O ~ ~eia+r RtdD rirRRr- NaCC R s Rf feaaeE WW-C&U,

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Potenfiometric titration of chlorides, efc. 74 (1955) RECUEIL 527

trodes broken, the two circuits of valve 6SN7 are balanced by adjusting the variable resistances shown on the right until the micro- ammeter reads zero. The two input leads are then connected to the two electrodes, The potential developed by the two half-cells destroys

STeb,U CLR

Fig. 5. Diagram of circuit to detect the presence of a difference of potential with continuous reading.

the balance and titration is continued until the meter once more reads zero. The use of this apparatus permits the replacement of a delicate and costly galvanometer by a robust and inexpensive apparatus. The apparatus is home-made and the microammeter used costs about one tenth of what a galvanometer costs.

An outside view of the apparatus is given in fig. 6 . Below on the left is a switch with two positions “Read” and “Set Zero”. In the middle is a knob for adjusting the zero. O n the right is a switch giving various sensitivities. Above on the left is a light indicating when the apparatus is switched on, in the middle is the microammeter and on the right is the switch for switching the apparatus on and off. The apparatus operates from the mains electric supply.

The modus operandi is as follows: the sensitivity switch is set to the lowest sensitivity and the left hand knob is set to “Set Zero”. The apparatus is switched on and allowed to warm up (a t least ten minutes). When it is desired to carry out the titration, the titration

528 Arthur Jordan,

L - - - - Fig. 6. External appearance of the continuous-reading apparatus for detecting

the existance of a potential between two electrodes.

mixture and the electrolyte bridge are set up and the burette is filled. The titration beaker normally contains 10 ml water, 3 ml H N 0 3 N/1 and 1 ml of the serum for analysis. The zero is adjusted and then the left hand knob is turned to "Read" and titration begun. When the microammeter reading falls to about one third of the initial reading, the sensitivity switch is turned to the next higher sensitivity and zero once more adjusted by switching the left hand knob to "Set Zero" and adjusting zero: the titration is then continued in the same way This series of operations is continued until the end-point is reached when the microammeter reads zero with the apparatus switched to maximum sensitivity. The method gives good reproducibility. A series of four titrations of the same serum gave the values:

100 m-equiv./l, 101 m-equiv./l, 101 m-equiv./l, 101 m-equiv./l. Control experiments have shown that the protein does not affect the readings and that all the chloride is determined. Comparisons with other methods give satisfactory agreement.

In principle the method can be used for any amount of serum. In practice, dilution causes the end point to lose its sharpness (fig, 7). If small amounts of serum were to be titrated, it were better to develop a micro-apparatus and to use the same concentration of silver nitrate from a microburette.

Potenfiometric tifration of chlorides, etc. 74 (1955) RECUEIL 529

. Q

0

IL. 0

0 0 I

. 0

0 Q

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Fig. 7. Effect of dilution on the curve of a potentiometric titration.

Acknowledgements. I am very much indebted to Mr. A. B. Kesfeven for designing and

making the valve bridge circuit apparatus. Without his continual advice and assistance, the method would not have been developed. M y own staff have used the apparatus continuously for four years and their comments and experience have been invaluable.