Nov., 1958
1455
NOTES
electrode. The half cell contains an electrode, e.g., Ag, the molten salt, e.g., AgN03, and a porous diaphragm.2 It can be shown readily that with appropriate precautions weight changes of 0.5 mg. can be detected. The experiment was attempted first with silver nitrate and silver electrodes using a Pyrex U F gintered glass disk for the membrane. Before current was passed through the cell it was noted that the weight of the half cell changes steadily indicating that leakage of the molten salt through the membrane was occurring due to the hydrostatic head. Since this meant that the membrane is unsuitable for such a measurement a search was instituted for a membrane material which would have no detectable leakage but would also have a sufficiently low resistance (less than 10,000 ohms/cm.2) to permit use in a transference experiment. We report here on the results of this search. Five types of membrane were evaluated by determining their leakage rates and electrical resistances under a hydrostatic head of 1 cm. of the appropriate molten salt. The list includes all of the types of membrane used by researchers in this field.3 The results are summarized in Table I. TABLE I Membrane
Salt
Temp., OC.
Leakage
Resistance, ohms
Pyrex UF 2,000 PbBrz 420 >50 mg./hr. AgCl 485 10 mg./hr. 10,000 Vycor UF PbCla 550 30 4,000 500 5 10,000 (0.9-1.4mp) AgCl No. 7930" PbClz 550 0 > 100,000 510 0 > 100,000 (4 m m d AgCl "Porcelain"* AgNOa 220 20 6,000 (0.15 mp) AgCl 500 Erratic 10,000 Alundum' AgN03 220 Rapid AgCl 500 Rapid 5 A Vycor-like glass which has been leached but not shrunk. Corning Glass Co. b Selas Corp. A consolidated membrane composed of Norton Co. RA 1055 Alundum cement supported on a sintered glass disk. (0.9-1.4 mp)
The following points should be emphasized. All of the membranes leaked substantially except Corning no. 7930 which showed unmistakable signs of reaction and had too large an impedance. In all cases the passage of small currents (1 ma./cm.2) caused a substantial increase in leakage rate, probably because of local heating. This means that estimates of leakage rates which have sometimes been reported in terms of the no-current condition3 are too small. The increase in electrical resistance with decrease in leakage rate reflects the close parallel between viscosity and electrical resistivity in a molten salt. From an inspection of the magnitude of the entries in Table I it seems likely that a further search for a membrane with both low enough resistivity and low enough leakage rate will not be successful. (2) By providing a side pocket to the cell the method has been extended t o liquid electrode materials such as Pb. (3) F. R. Duke and R. W. Laity, THIS JOURNAL, 68, 549 (1955): J . A m . Chem. SOC.,7 6 , 4046 (1955); H.Bloom and N . J. Doull, THIS 60, 620 (1956); G.Jans and M.Lorena, AFOSR-TN 57-240. JOURNAL. A D No. 126537, 1957; F. R. Duke and R. W. Laity, J . Electrochem., 101, 97 (1958).
Fig. I.-Cell for evaluation of membrane leakage.
This work has been assisted by the United States Atomic Energy Commission, Contract AT (30-1) 1938 with New York University. (4) C/. M. R. Lorenz and G. J. Janz, THISJOURNAL,61, 1683 (1957).
INTERPRETATION O F T H E PROPERTIES OF PROTEINS I N CONCENTRATED SALT SOLUTIONS BY J. R. MCPHEE W o o l Textile fiesearch Laboratories, Commonwealth Scienli$c Industrial Research Organization, Geelong, Auslralza Received April 26, 1968
and
The presence of high concentrations of electrolyte considerably modifies the behavior of proteins in aqueous solution affecting such properties as swelling,' solubilit8y,2 denat~ration,~meltingpoint of gelatin gels,4 thermal shrinkage of collageii,b strength6 and supercoiitraction of keratin fibers.' ( 1 ) F. Hofmeister, Arch. Bxp. Pathol. Pharmakol., Naunyn-Schniedeberg's, 24, 247 (1888); 27, 395 (1890); 23, 210 (1891). 43, 513 (2) A. R. Docking and E. Heymann, THISJOURNAL, (1039). (3) H.Chick and C. J. Martin, J . Physiol. (London), 40, 404 (1910); 46, 61 (1912): A. E. Mirsky and M. L. Anson, J . Gen. Physiol., 13, 307 (1938); A. E.Mirsky, ibid., 19, 559, 571 (1936); J. P . Greenstein, J . Biol. Chem., 130, 519 (1930); N. F. Burk, THISJOURNAL, 47, 104 (1943); R. B. Sinipson and W. Kauzmann, J . Am. Chsm. SOC.,71, 5139 (1953). (4) W. Pauli, Aich. ges. Physiol., Pjluger's, 71, 333 (1898); J. Bello. H.C. A. Riese and J. R. Vinograd, THISJOURNAL, 60, 1299 (1956). (5) J. R. Rats and A. Weidinger, Biochem. Z . , 269, 191 (1933); F. G. Lennox, Biochim. et Biophys. Acta, 3, 170 (1949). (6) A. h.1. Sookne and h l . Harris, J . Research Natl. Bur. Standards, 19, 535 (1937); J. B . Speaklnan and E. Whewell, J . Testile Inst.. 41, T329 (1950); P. Alexander, Kolloid-Z., 122, 8 (1951). (7) W. G. Crewther and L. M. D o d i n g , Nature, 178, 544 (1956).
NOTES
1456
The concentrated salt solutions are generally considered to influence proteins either by dehydration, due to certain ions (e.g., SO4-) having a high energy of hydration and therefore removing water from the protein, or by increasing the protein hydration, due t o the possibilities, for example, . that certain ions (e.g., CNS-) may be adsorbed a t the protein-water interface or perhaps be bound by polar groups, and that these ions would carry some water molecules with them although their energy ~~ although specific of hydration is I O W . ~However, sorption of CNS- has been observed in one case,2 this could not be d e t e ~ t e din ~ . other ~ studies. It would appear preferable, therefore, to explain salt effects in other terms until such sorption has been clearlv demonstrated exDerimentallv. Doiking and Heym;nn2 demoGstrated an apparent negative sorption of anions when gelatin was equilibrated with certain salt solutions. Their results for sulfate mean that gelatin is fully hydrated in concentrated S04= solutions and that the sorbed water contains no electrolyte. Therefore, although gelatin swells less in solutions of sodium sulfate than in water, this is not due to a decrease in chemically bound water. McDevit and Longlo relate the magnitude of salt effects on non-polar molecules t o the changes in volume which take place when the salts are dissolved in water. If there is a decrease in total volume on mixing salt and water, due to ionwater interaction, work will then have to be done against ion-water forces to intrpduce a non-polar solute, the only role of this non-polar solute being to occupy its given volume. Such ion-water interaction results, therefore, in the observed “salting-out” of non-electrolytes. When there is a volume increase on mixing salt and water, the semicrystalline water structure is “loosened” and “salting-in” may take place. This would occur with large ions where the electrostatic field strength a t the surface of the ions would be low and when water molecules would be displaced against their own cohesive forces. McDevit and Long also express their theory in terms of the “effective pressure,” P,, exerted by a salt in solution. Gibson“ introduced P e as a parameter in Tait’s equation for the compressibility of water, and Pe can be considered to be the increase in internal pressure of water due to the addition of a solute. This follows Tarnmann’sl2 suggestion that a given amount of water in solution behaves as the same amount of pure water under a constant external pressure, and P e is then the hydrostatic pressure which changes the properties of water to the same extent as a given concentration of solute. The extra work necessary to introduce a non-polar solute into an electrolyte solution is simply the product of P , and the increase in volume of the liquid on introducing the non-polar solute. (8) J. R. Kate and F. W. Musohter, Biochem. Z . , 267, 385 (1933). (9) J. W.Williams, W. M. Saunders and J. 8. Cicirelli, THISJOUR774 (1954). (10) W.F. MoDevit and F. A. Long, J . Am. Chem. floc., 74, 1773 (1952). (11) R.E.Gibson, mbid., 56, 4, 865 (1934); 67, 284 (1935). (12) G. Tammann, Z.onovg. &em. Chem., 164, 25 (1926). NAL, 58,
__
Vol. 62
With polar solutes, other factors, such as the dipole moment of the molecule and possible interaction of ions with polar groups, could be superimposed on the major role of the solute, which is the occupying of a given volume. There seems t o be little doubt that the specific effects of salt solutions must arise mainly from the properties of the solutions themselves, since the relative effects of different anions are usually similar on solutes ranging from simple, non-polar, relatively inert compounds such as hydrogen and benzene which would not be dehydrated by salt solytions or would not bind ions, to complex macromolecules such as proteins. Solutions of SzOchave now been found to have an effect on wool similar to that of SO4- on gelatin and it is the purpose of this note to report this behavior and to indicate that the concept of salt solutions exerting an “effective pressure” is adequate in qualitatively explaining salt effects on proteins. Experimental Wool.-Worsted fabric was Soxhlet extracted for ti hours with light petroleum, followed by Soxhlet extraction for 24 hours with cold ethanol. After rinsing in several changes of distilled water, the wool was allowed to stand in 0.001 N HC1 for two days and then washed in many changes of distilled water daily until the pH of the water did not change overnight. The ash content was 0.07%. Similarly purified Corriedale ioose wool was used for swelling determinations. Sorption Experiments.-Wool samples (about 5 g. dry weight) were wet out in vacuo in 75.0 ml. of salt solution at 25O; samples of solution were withdrawn at intervals and diluted appropriately for volumetric analysis for the anion. Equilibrium was reached in 1-2 days. Sample volumes and dilutions were such that titers were approximately 20 ml. Thiocyanate was titrated with standard silver nitrate solution using ferric ammonium sulfate as indicator, chloride was estimated by Mohr’s method and thiosulfate was determined by adding an excess of standard potassium iodate solution and back-titrating with standard thiosulfate solution. Identical results were found using dry wool or wool conditioned in an atmosphere at 65% R.H. and 20’, when allowance had been made for the moisture content of this conditioned wool. Swelling Measurements.-These were made in a circular polythene cell, about 1in. diameter, which was designed and constructed by Mr. J. Delmenico. The cell contained two diametrically opposite metal rods which could be rotated by hand. The fiber to be studied was attached to the ends of the rods with polystyrene and the cell clamped to the mechanical stage of a suitable microscope containing a micrometer scale in the eyepiece. The diameter of each fiber was measured at four points along the fiber, four measurements being made at each point by turning the fiber through 45, 90 and 135’. Fibers coeditioned at 65% R.H. and 20’ were used. These had a diameter 7% greater than the dry fiber, as measured in anhydrous glycerol. Measurements at the identical points were then taken after equilibration in the salt solution. The swellings given were calculated from the values obtained on conditioned fibers.
Results and Discussion The water sorbed preferentially by wool was calculated from the change in concentration of salt solutions. The data from this differential titration procedure are rather inaccurate and are used only in a qualitative manner. More definite information could be obtained by direct determination of the amount of salt in the wool but this has not yet been done. The concentration of each salt solution increased when wool was added (Table I), except for 2 M NaCNS where it did not change. (This be-
NOTES
Nov., 1958
i457 ,
TABLE I:
SWELLINQ OF
Wo OL
Wt. of wool
Solution
(P.1
AND
PREFERENTIAL SORPTION OF WATER FROM SALT SOLUTIONS AT 25’ ---Concn. Initial
(M)Final
Water sorbed preferentiallyG (% on wt. of d r y wool)
Swellingb
(%)
Effective pressure” (bar)
Water 33d 18 0 NazSz03 (2 M ) 5.003 2.004 2.043 29 12 (1700) Na&03 (satd.) 5.010 3.818 3.898 30 6 (3300) Na2S04 (satd.) 1.9 12 1600 NaCl (2 M ) 4.996 2.012 2.039 20 14 540 NaCl (satd.) 5.008 5.193 5.278 24 13 1400 NaNOj (2 M ) 2.0 16 400 NaN03 (satd.) 7.9 11 1600 NaCNS (2 M ) 5.011 2.015 2.013 0 19 (350) NaCNS (satd.) 4.992 8.917 9.086 28 10 (1600) a Calculated from the increase in concentration of salt solution. b The figure given for each solution is the mean of the measured swellings for five fibers. The standard deviations of the means were in all cases f 1. From ref. 10; the figures for NazSz03and NaCNS were calculated assuming the increase in effective pressures per mole to be the same as for NatSOa and NaI, respectively, although NaCNS almost certainly would give lower pressures than NaI. Accepted value for the water sorbed by wool in an atmosphere at 100% R.H. or in pure water.13
havior is similar to gelatin,2 where, however, the CNS- solution showed a decrease in concentration.) The water sorbed by wool from thiosulfate solutions is approximately equivalent to that taken up from pure water if no salt has been sorbed. Although the wool is therefore fully hydrated in thiosulfate solutions, it swells less than in pure water (analogous to the effect of SO4- on gelatin). Wool swells to approximately the same extent in 2 M NaCNS as in water alone, the solution being sorbed as such so that either the water which can be sorbed preferentially by wool now contains 2 M NaCNS, or, alternatively, NaCNS is sorbed a t specific sites a t the same time as water. It is perhaps unlikely that the combination of these latter two effects would lead to the concentration of the solution remaining constant, but the present results do not allow a distinction to be made between the alternative explanations. Saturated Na2S04 solution (1.9 M ) is similar to 2 M NazSz03whereas 2 M NaCl and 2 M NaN03 are intermediate in behavior between NazS2O3and NaCNS. Thus, in electrolyte solutions where ion-water interaction is high (SzO3-), and soluble non-electrolytes are salted out, proteins such as wool and gelatin are prevented from swelling due to the increased internal pressure of water brought about by ionwater forces. In the 2 M solutions studied, swelling increases with decreasing effective pressure of the solution, until, in 2 Af NaCNS, swelling may be slightly greater than in water alone. A further effect would arise from the protein containing a solution of high ionic strength when the effective pressure is low. This would reduce interaction between polar groups and allow the structure to be more easily disrupted. The effect on the swelling of wool of 2 M CNS- is small but might be expected to show up more clearly in other properties of wool or with soluble proteins. When saturated solutions are compared, rather than a series a t given concentration, the solubility of the salts needs to be considered since, even though the increase in effective pressure per mole may be low, the effective pressure of the saturated solution may be large if the salt is highly soluble. This appears to be the case with NaCNS. As(13) P. Alexander and R. F. Hudson, “Wool. Its Chemistry and Physics,“ Chapinan and Hall Ltd., London, 1954, p. 126.
suming a linear relation between effective pressure and molarity, the saturated salts in Table I should give approximately the same pressure and therefore swelling, except Na2S203where the pressure would be much greater and the swelling consequently lower. This is approximately true. It generally has been considered that the properties of wool in concentrated salt solutions are those of wool in an atmosphere a t the relative humidity above the solutions. The wool would then contain less sorbed water and would be swollen less than in dilute aqueous solutions. However, this would mean that the series of 2 M salt solutions should all have had approximately the same effect on wool. Also, the swelling in saturated thiosulfate solution is less than expected from the decrease in vapor pressure (6% found compared with 9% calculated), while the water sorbed (containing no electrolyte) is greater than expected (30% found compared with 18% calculated). Properties of proteins such as swelling, solubility, denaturation, thermal shrinkage, etc., which are influenced by salt solutions, can be thought of simply as general configurational changes. Certain ions in water, e.g., CNS-, I-, promote such changes since the solutions exert a low effective pressure, or the ions may perhaps ‘Lloosen”the water structure, and the protein then contains solutions of high ionic strength. Other ions in water, e.g., SO4=, prevent or considerably retard the rate of these structural changes since the solutions exert a strong effective pressure and the water sorbed by the protein coiitains no electrolyte. Acknowledgment.-I am indebted to Dr. M. Lipson and Dr. J. H. Bradbury for many most helpful discussions and to Miss L. Boardman for assistance with experimental work.
KINETICS OF HYDROLYSIS OF DECABORANE BY R. W.ATTEBERRY High Energy Fuels Division, Olin Mathieson Chemical Corp., Niagara Palls, N . Y. Receiued M a y B 1 , 1968
Kinetic studies of the alcoholysis of decaborane’ show that reactions with secondary alcohols possess
