Polarographically Active Substances

Values elevated above 21 were found sporadically in some cases of brain tumor. It is evidently a decisive factor whether polarographically active substances do or do not penetrate into the cerebrospinal fluid. It can be said that values above 21 always portend a rather serious prognosis. 

Gastric juice was examined (237) by the Brdidka filtrate reaction, and a polarographically active substance was found only in gastric mucus but not in the proper gastric juice. For examination, the mucus was separated from the juice by centrifugation, dried in a desiccator, and weighed in 20-mg portions to obtain standard conditions. The curves were, however, not always typical the author explmned the deformation by alterations of the mucus due to the drying process, and did not recommend his procedure. 

The potential, at which the current reaches the half value of the limiting current, is called the half-wave potential in Fig. 2). This value of the oxidation or reduction potential is a physical constant, in most instances practically independent of the concentration of the particular compound and characterizing qualitatively the electroactive compound. The half-wave potential ( j) is dependent on the nature of the electroactive species, and on the composition of the supporting electrolyte. Only in some instances, and to a comparatively small degree, is the half-wave potential dependent on the concentration of the polarographically active substances, temperature and the characteristics of the capillary used. 

Some forms of polarographically active substances are adsorbed at the mercury dropping electrode. In such instances two waves may be observed on polarographic curves, one of which corresponds to the reduction (or oxidation) of the free form and the other of the substance in the adsorbed state. The adsorption current increases with the increasing concentration of the electroactive species, but only to a certain value, which is a function of the surface of the electrode. No further increase of current is observed with further increases of concentration, and we suppose that the surface of the electrode under such conditions is covered by a layer of the adsorbed substance. The adsorption current (fo) is given by the equation< >... 

The procedure known as quotient of two waves is independent of temperature. In this method a known quantity of a certain polarographically active substance is added to each solution in which another component is to be determined. It is assumed that the ratio of the two waves (for the unknown and for the added standard or pilot) is independent of temperature, viscosity, rate of flow of mercury etc. The calibration curves are constructed by plotting the ratio of the two waves against the concentration of the substance to be determined, keeping the concentration of the pilot constant. These calibration curves are used in the same way as the ordinary curves. 

This enables traces as low as 10 M to be determined. With the square-wave polarograph, concentrations one order lower can be analysed. The most sensitive method as yet seems to be the method of anodic stripping . In this method reduction products can be formed at the surface of the electrode and then stripped anodically. The anodic current corresponding to the dissolution process is either followed polarographically or with an oscilloscope. Similarly oxidizable substances can be stripped cathodically. These methods enable polarographically active substances to be detected in 10 or even 10 M solutions. )... 

The effect of pH, concentration of the polarographically active substance and temperature on half-wave potentials. 

In indirect methods, as mentioned in Chapter IV, polarographic-ally inactive substances are transformed into compounds showing waves on polarographic curves, or concentration changes of a polarographically active substance which reacts with the electro-inactive compound to be determined, are measured. Finally, polarometric (amperometric) titrations can be included in this group of analytical methods. 

Together with addition methods mentioned here, the reaction of unsaturated bonds with maleic anhydride is discussed in the section devoted to the decrease of polarographically active substances (p. 143). 

Here Ee is the standard potential of the reaction against the reference electrode used to measure the potential of the dropping electrode, and the potential E refers to the average value during the life of a mercury drop. Before the commencement of the polarographic wave only a small residual current flows, and the concentration of any electro-active substance must be the same at the electrode interface as in the bulk of the solution. As soon as the decomposition potential is exceeded, some of the reducible substance (oxidant) at the interface is reduced, and must be replenished from the body of the solution by means of diffusion. The reduction product (reductant) does not accumulate at the interface, but diffuses away from it into the solution or into the electrode material. If the applied potential is increased to a value at which all the oxidant reaching the interface is reduced, only the newly formed reductant will be present the current then flowing will be the diffusion current. The current / at any point... 

General considerations. Conventional d.c. polarographic analysis is most conveniently carried out if the concentration of the electro-active substance is 10-4 — 10 3 molar and the volume of the solution is between 2 and 25 mL. It is, however, possible to deal with concentrations as high as 10 2 molar or as low as 10 5 molar and to employ volumes appreciably less than 1 mL. 

Simultaneously the wave in increases, and in the final product, corresponding to a mixture of benzaldehyde and acetophenone, only waves 3 and 4 persist. The wave at intermediate stages of the reaction therefore corresponds to the reduction both of the ketol VII and of acetophenone, which is reasonable since the polarographically active benzoyl grouping is the same in both substances. To determine the ketol concentration it is necessary to subtract from wave in the current corresponding to acetophenone reduction. Since acetophenone is formed at the same concentration as benzaldehyde and because for equimolar solutions of benzaldehyde and acetophenone the ratio of wave heights iz/ii is practically equal to unity, it is possible to determine the ketol concentration simply from the difference 4 — t3. 

Sulphydryl compounds belong to the most important among biologically active substances. The polarographic behaviour of sulphydryl compounds... 

Polarography offers some possibilities for the study of reaction kinetics and mechanisms of homogeneous organic reactions. The main advantages are a rather simple and easily accessible experimental technique, the possibility to work in dilute solutions and limited requirements on the amount of substances studied. The main limitation is that some of the components of the reaction mixture must be polarographically active. But this limitation is not so restrictive as it would appear, because most substances that can be studied spectro photometrically are electro-active as well. For rapid reactions polarography seems to be most useful for a range of second-order rate constants between about 10 -10 sec M, whereas for faster reaetions the specific properties of the electrode, in particular its electrical field and adsorption, can play a role. A certain limitation is that for most systems the equilibrium constant has to be known from independent measurements. 

Maxima are least likely to be encountered if one uses a low rate of mercury flow (jn < 0.5 mg/sec), a concentrated supporting electrolyte, and a low concentration of the electroactive species ( < 10 Af). Maxima can sometimes be avoided by altering the chemical nature of the supporting electrolyte. If need be, the maxima can usually be suppressed by adding to the solution a small amount of a surface-active substance, such as Triton X-100. One should add just enough to eliminate the maximum (determining the amount by trial and error), since even a moderate excess can distort the polarographic wave. 

PUot-Ion Method. Another method, known as the quotient of two waves or pilot-ion method, is less frequently used, but offers some advantages. (This is known more generally as an internal standard method.) A known quantity of a standard pilot substance is added to each investigated solution. This pilot substance must be polarographically active and give a wave or waves in a potential region (preferably at more positive potentials) sufficiently different from that of the compound to be determined. 

Finally, when amperometric measurements are carried out the decision must be taken automatically which set of the currents samples represents the diffusion current only. If the current samples are going to be used for a polarographic record the same principle is employed. Current samples for which the exponent value suggests any electrochemical complication are rejected. In the past much attention was given to studies of the i-t curves on a single drop as well as how the surface active substances influence the shapes of the curves. Worth noticing in this respect, according to the authors opinion, are a number of studies which sum up the more important developments. ... 

Earlier examinations of cerebrospinal fluid were performed on the basis of suppression of polarographic maxima (214). The estimation of total protein in 0.1 ml of cerebrospinal fluid, calculated from the percentile depression of the maximum, is in fact an estimation of surface-active substances although protein is much active in this respect, the method represents no specific determination of protein. The same author (214) and also further authors (215) also made use of the pre-sodium wave and of the protein wave in buffered solutions of Co salts, but here also no parallelity was found between the wave height and the protein content, since the number of polarographically active groups present was variable. 

When the polarographic wave is preceded by another wave, the limiting current is the difference between the parallel currents in the region where they are independent of potential. The limiting current is usually proportional to the concentration of the polaro-graphically active substance. 

Whereas two coincident waves, corresponding to two different reducible groups, are not too often encountered in practice, a current problem is in distinguishing two structurally similar substances with the same polarographically active group. The half-wave potentials of two closely related substances only occasionally differ sufficiently to allow two separate waves to be measured. 

When maxima are observed on polarographic curves, gelatin or other surface active substances are sometimes added to the solution. The concentration of surface active substance should be as low as possible, because a higher concentration of gelatin, deforms the polarographic curve. The surface active agent damps the streaming of electrolyte around the polarized mercury drop. 

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