Aug., 195G
ANOMALOUS ADSORPTIVEPROPERTIES OF NITRICOXIDE
1063
THE ANOMALOUS ADSORPTIVE PROPERTIES OF NITRIC OXIDE1 BY R. NELSONSMITH,DAVIDLESNINIAND JOHN Moo1 Pomona College, Claremont, Calif. Received March 1.9, 1966
Nitric oxide reacts readily with carbon surfaces at 0 and -78' to form gaseous nitrogen and carbon-oxygen surface complexes. Thus, adsorption isotherms determined at these temperatures with carbon are fictitious; likewise, previous calorimetric measurements are heats of reactions, not heats of adsorption, for unknown amounts of nitric oxide. Nitric oxide does not react with a carbon surface at -154' and adsorption appears to be normal, though the adsorbed state is probably NzOz. NO also adsorbs normally on both porous and non-porous Si02 a t 0' and -78", though there is some reversible chemisor tion in each case (NO can be removed intact by heating) and it appears that the NO interacts t o some extent at -78' wit{ the water bound in the silica gel.
The original objective of this study was utilization of the free radical properties of NO as a means of characterizing the "free-valence" nature of surfaces. It was thought that the odd electron in NO would pair off with an odd electron in the surface, and that the amount of NO thus chemisorbed would be a measure of this type of surface activity. This phenomenon does not occur, and the reasons therefor are the subject of this paper. Experimental Two types of carbon and two types of silica were used in this study. Graphon.-A partially graphitized carbon black (lot #628) furnished through the courtesy of the Godfrey L. Cabot Go., Boston, Mass. The surface area is 82.8 m.2 per gram and the ash content is about 0.02%. Before each series of measurements the Graphon was given three 15minute treatments with Hz gas a t 1000". The HZ was pumped out between each treatment and after the last treatment the sample was cooled slowly with continuous pumping. About 1.9-g. samples were used for measurements. Su-60.-An activated sugar charcoal of extremely low ash content (less than 0.005a/0) prepared from Confectioners AA sugar furnished by the California .and Hawaiian Sugar Refining Corporation, San Francisco, Calif. The activation method is described elsewhere.2 The BET nitrogen surface area is 1060 m.2 per gram. It was Hz-treated in the same way as for Graphon before each series of measurements. About 0.8-g. samples were used for measurements. XC-54.-A non-porous "hyperfine silica" furnished through the courtesy of Dr. Mark Olson, Pigments Dept., E. I. du Pont de Nemours & Go. Electron micrographs show most of the particles to be discrete spheres of the order of 10-40 millimicrons diameter. The nitrogen surface area is 20.5 m.2 per gram. Before malting adsorption measurements the samples were heated to 850' zn uucuo for about 1 hour, then cooled with continuous pumping. Heating to higher temperature caused partial sintering of the silica. Approximately 0.45-g. samples were used. D-3707.-A commercial silica gel furnished through the courtesy of the Davison Chemical Gorp., Baltimore, Md. The BET ethyl chloride surface area is 838 m.2 per gram. Translucent particles of approximately 0.5-1 .O mm. diameter were used in the measurements. Approximately 0.49-g. samples were used and outgassed for several hours a t 300" before making measurements. Nitric oxide was prepared by the method of Marqueyrol and Florentin.3 Twenty g. of diphenylnitroRoamine (Eastman #759) was put into a 200-ml. round-bottom flask and sealed to the high-vacuum system which included a 2-liter evacuated Pyrex reservoir and a Dry Ice trap between the reservoir and diphenylnitrosoamine. The diphenylnitrosoamine was outgassed with continuous pumping and its temperature raised to about 150" with a Glas-col heater. At this temperature the gas generating system was closed (1) Progress report of work done under contract N8onr54700 with the Office of Naval Research. Reproduction in whole or in part is permitted for any purpose of the United States Government. (2) R. N. Smith and J. Mooi, T w a JOURNAL, 69, 814 (1955). (3) M. Marqueyrol and D. Florentin, Bull. SOC.chim. France, [4] 11, 804 (1912).
off from the pumping system and opened to the %lite; reservoir. The heating was then continued to about 250 until decomposition was complete, as evidenced by no more gentle bubbling of the liquid. The remaining tetraphenylhydrazine and the Dry Ice trap were then closed off from the rest of the system and coded. The gas analytical methods used in this study showed this gas to be pure NO. A conventional volumetric adsorption apparatus was used for determination of the isotherms. The dead space in the system was kept as small as possible; it was determined volumetrically with helium a t each temperature for each sample. The Graphon, Su-60 and XC-54 samples were sealed in quartz tubes and connected to the system with Pyrex-to-quartz graded seals so that these samples could be treated at temperatures up to 1000" directly in the system prior to adsorption measurements. The D-3707 was sealed in a Pyrex tube since it was not possible to heat it above 300'. Provision was made for removal of small amounts of gas by a Toepler pump and for its analysis by the micro methods of Blacet and Leighton. Two types of gas samples were involved; mixtures of CO, COZ and Nz and mixtures of COZ, NO and Nz. Both types were analyzed by the same method. COZ was removed by a KOH bead4; GO or NO was removed by a dry AgzO bead' held to the platinum wire with Kronig cement (NO has not heretofore been determined by this method); Ne was determined by difference. For samples containing a high percentage of GO or NO, it was found expedient to dilute the samples with pure Nz before analysis. To achieve the constant temperatures used for the various adsorption measurements the following materials were used in Dewar flasks: ice and water for 0"; Dry Ice for -78'; and solidliquid mixture of 2-methylpentnne (99 mole Yo ininimun purity, Phillips Petroleum Go.) for -154". 2-Methylpentane is convenient in that it supplies the desired temperature a t its freezing point and yet is easily stored a t room temperature (b.p. SO'). It is inconvenient in that its ice must be made with the adsorption bulb in situ by carefully adding liquid nitrogen and stirring well. A good large Dewar flask, well-insulated from the surroundings and initially filled almost solid with the ice, will keep about 7 to 8 hours before re-preparation of ice is necessary. The temperature obtained with 2-methylpentane mush was probably not controlled very precisely in this manner, but for the purpose of these experiments it was satisfactory, and will be referred to as - 154".
Results Figure 1 shows characteristic adsorption and desorption isotherms for NO on Su-GO. The adsorption a t 0 and -78" is not large, but judging from the unusual hysteresis loop the adsorbed N O is unquestionably chemisorbed. If one then endeavors to desorb this tightly-bound NO by vigorous heating, he finds that the volume of gas thus evolved plus the small bit desorbed before heating is only a little more than half of the gas which was originally introduced to the charcoal. Analysis of the gas desorbed on heating shows it to be al(4) F. E. Blacet and P. A. Leighton, Ind. Eng. Chem., Anal. Ed., 3, 266 (1931). (5) F. E. Blaqet, G. D . McDonald and P. A, Leighton, ibid., 6 , 272 (1933).
R. N. SMITH,D. LESNINIAND J. Moo1
1064
F
I 20
I
I 60
40
I 80
I
I
I
120
100
I 160
140
I I00
Vol. 60
sample at 0" and at a pressure of 50 mm., the "equilibrium gas" was 14.0% Nz and 86.0% NO; a t -78" and GO mm. pressure, the gas was 38% N2 and 62% NO. Another experimental difficulty involved with "adsorption" of NO a t these temperatures is the slow rate of attainment of equilibrium after the initial adsorption. This difficulty is caused by NO having to diffuse through N2 in order t o reach the surface, and it can be eliminated by occasionally withdrawing and mixing the gases with a gas buret and returning them t o the carbon sample. Perreue had the same difficulty with slow attainment of equilibrium in his calorimetric studies. At -154" (in the narrow liquid range of NO, - 151.8 to 163.5") the situation is quite different. Here the adsorption is some 10 times greater than the apparent adsorption observed at -78 and 0", and furthermore analysis shows the equilibrium gas to contain only a trace of N2. One also finds that equilibrium is rapidly attained. I n other words in the temperature range where NO is normally a liquid, true adsorption occurs and not reaction. However, even this true adsorption is complicated for all evidence indicates that liquid nitric oxide is a dimer.' This dimerization also probably accounts for its unreactivity toward the Carbon surface. A B E T Plot of the adsorption of NO on Graphon at -154" gives a v m value of 8-75 x mole of NO Per gram; this in turn corresponds to the reasonable cross-sectional area of 15.7 per m o l ~ u l eof NO (or 31.4 1.'per molecule of N202). ~ ~ 1 c u l a t i o using ns gas analyses and the apparent adsorptions show that a t 0" and 200 mm. pressure 2% of the surface carbon atoms of Su-60 are oxidized by NO (assuming 1 oxygen atom per carbon atom), whereas under the same conditions 0 2 oxidized 1.5%. At -Bo, 1% of the surface is oxidized by NO. In the case of Graphon, similar calculations show 1.1% oxidized a t 0" and 0.3% a t -78%. These results are significant in connection with the heats of adsorption (- AH,d,) which have been measured for nitric oxide on charcoal. These heats have in reality been heats for the reaction
P, mm.
Fig. 1.-Adsorption (0) and desorption ( 0 )isotherms for NO on charcoal, Su-GO: curve 1 for 0 ' ; multiply ordinate for moles/g.; curve 2 for -78'; multiply ordinate by by 4 X for moles/g.; curve 3 for -154"; multiply ordinate by foi, moles/g.; curve 3 is the only true isotherm.
most entirely C 0 2and CO, and in relative amounts typical of the thermal decomposition of carbonoxygen surface complexes (preponderance of C02 a t the lower temperatures; CO a t the higher), One cannot escaDe the fact that the NO has reacted rapidly wit'h the carbon surface to form N2 and carbon-oxygen surface complexes, and that the gas pressure measured for the isotherm is essentially N2, not NO. This is easily verified by analysis, and for an "equilibrium pressure" of about 75 mm. a t 0" one finds a typical analysis to be 92.9% N2, (3.2% NO and 0.9% COz after standing 1 hour, and after 21 hours the analysis becomes 99.0% Nz, 0.5% NO and 0.5% COz. With higher pressures one finds increasing amounts of NO in the gas phase. At -78" ''equilibrium" is also rapidly achieved, and a t a pressure of 113 mm. the gas was found to be 96.2% N2, 3.1% NO and 0.7% COz. With Graphon (Fig, 2) the apparent effect is not so pronounced as with Su-GO simply because of the smaller specific surface area. Thus, with a larger
NO
+C
(CO)
+ '/zNz
(1)
where (CO) is the solid carbon-oxygen surface complex. In these calorimetric measurements the amounts of NO assumed to have been adsorbed are in error, for in correcting for the amount of gas unadsorbed the assumption was made that the residual gas was NO, not Nz. I n reality, then, more moles of NO had reacted with the surface than had been assumed to be adsorbed, and thus the integral heats of reaction (cal./mole) would be lower than those published erroneously as integral heats of adsorption. A plot of integral heats vs. gas pressure is meaningless since the gas (Nz NO) pressures are in reality functions of the dead space in the calorimeter. It is also not reasonable to compare directly the heats of adsorption of NO with those for Oz as has
+
B
16
24
32
40
40
SB
64
71
BO
t
-
I 200
ea
P,mm.
Fig. 2.-Adsorption isotherms for N O on Graphon, a carbon black: curve 1 for 0"; multiply ordinate by 1 0 - 6 for moles/ g.; curve 2 for -78"; multiply ordinate by 10-6 for moles! g.; CwVe 3 for -154'; multiply ordinate, by for moles/g.; curve 3 is the only true isotherm,
(6) J. Perreu, B U Z ZSOC. . chim. France, 15116,919 (1949). (7) A. L. Elniith, W. E. Keller and H. L. Johnston, J . Chem. Phys., 19, 189 (1951).
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Aug., 1956
ANOMALOUS ADSORPTIVE PROPERTIES OF NITRICOXIDE
1065
been done previously. However, one might look a t the "adsorption" of each of these gases as the production of a carbon-oxygen surface complex. For oxygen, the "heat of adsorption" is identical with the "heat of formation" of this surface complex. For NO, the heat of formation of the surface complex may be calculated assuming AH = 0 for the formation of C and N2 and AH = +21.6 kcal./mole for NO. Thus, from (1) A H c o m p ~ e x = AHada
- 21.6
where --Hads = the observed heat of reaction. Using Perreu's6 values for the adsorption of NO a t 0" on an activated coconut charcoal one may calculate that the heat of formation of the complex, -AHco,nplex, for the first amount added is 48.6 kcal. per g. atom of oxygen and that this drops to about 45 kcal. per g. atom of oxygen at a pressure of about 100 mm. If all the unadsorbed gas in the calorimetric measurements was nitrogen, then these values should be cut in half for the number of moles of NO which had reacted would be twice the number believed to have been adsorbed. Actually, the unadsorbed gas will be a mixture of NZ and NO (being almost entirely NO in the leads) and these heats of formation must lie somewhere between 25 and 50 kcnl. per g. atom of oxygen. In another paper, Perreus also determined the heat of adsorption of oxygen (the heat of formation of surface complex) on this charcoal a t 0" and found a value of 78.9 kcal. per mole of Oz;this is equivalent to 39.5 kcal. per g. atom of oxygen, a value of the same order as that obtained with NO. In other ways too (from the composition as a function of temperature of the gases evolved on heating) it appears that the carbon-oxygen complexes formed by NO and O2 are much the same. The mechanism by which these two dissimilar molecules give the same complex is of some interest. Each is paramagnetic and it may be that the first step in each case is the pairing of electrons by oxygen atoms with the surface. Bull and Garnerghave published for an activated charcoal graphs of differential heats of adsorption of NO and O2 vs. the amount absorbed, and these values too must be in error for NO since the curve (of integral heats vs. concentration) from which the differential heats were taken must be in error for the reasons given above. The adsorption studies using NO and Si02 were undertaken to see whether NO would be chemisorbed on this type of surface, and to check on the very unusual adsorption isotherm found a t - 78" for NO on silica gel by Briner and Sguaitamatti.'O The results are summarized in Figs. 3 and 4. The same amount (about 36 pmoles per g.) of NO is very strongly adsorbed on the non-porous silica a t both -78 and 0 " ; a t 0" this is about all of the NO which is adsorbed even a t the higher pressures. There is a great difference between the amount of NO very strongly held by the silica gel at the two temperatures; a t 0" the amount is about 7 pmoles per gram and a t -78" it is about (8) J. Perreu, Compl. rend., 226, 907 (1948). (9) H. E. Bull and W. E. Garner N n l u i e , 134, 409 (1929). (10) E. Briner and B. Sguaitarnatti, Helv. Chim. ilcla, 28, 1216 (1940).
Fig. 3.-Adsorption ( 0 )and desorption ( 0 )isotherms for NO on XC-54, a non-porous silica: curve 1 for -78"; multiply ordinate by 10-5 for molcs/g.; curve 2 for 0"; multiply ordillate by 10-8 for rnoles/g.
S
Fig. 4.-4dsorption ( 0 )and desorption ( 0 )isotherms for NO on D-3707, a silica gel: riirve 1 for 0 " ; multiply for moles/g.; curve 2 for -78'; multiply ordinate by ordinate by 10-4 for rnoles/g.
440 pnzoles per g. For both the non-porous silica and the silica gel it was found that the tightly-held NO could be removed intact by heating and in amounts corresponding to that expected from the hysteresis loop of the isotherm. It is interesting tto note that some NO is "chemisorbed" on these samples of SiO2, but the amount does not seem to be related in any simple way to the nature of the surface. The adsorption isotherms appear to be normal and the one published by Briner and Sguaitamatti showing an enormous increase in adsorption a t a pressure of 300-400 mm. a t -78" seems very improbable. Their isotherm was based on only 4 points determined by a gravimetric method, and no points are given for pressures below 300 mm. It was observed under certain conditions, in agreement with Briner and Sguaitamatti, that silica gel with its adsorbed NO is brick-red in color a t -78". If, starting a t a pressure of GOO mm., the temperature is allowed to rise and the NO to escape, the color weakens, changes to an olive green and then to colorless. If one starts at -78" and a pressure of 1 mm. the color is initially a pale olive green which changes to colorless as the temperature rises. At a pressure of GOO mm. the silica
108F
MYRON1,. WAGNER A N D HAROLD 24.SCHERACT.4
gel contains 1.25 mmoles per gram, while a t a pressure of 1 mm. it contains 0.035 mmole per gram. The non-porous SiOz holds 0.35 mmole per gram a t 500 mm. pressure and is absolutely colorless (white) just as it is a t 1 nim. pressure where it holds 0.01 mmole per gram. Briner and Sguaitamatti ascribe this coloration in the gel to the color of the dimer, but NO when at temperatures above its boiling point (and not adsorbed) is a m0nonier.7 Thus, it seems unlikely that there will be any significant amount of N20z at a low pressure a t -78" (which is 72" higher than the boiling point and 16" above the critical temperature.) I t is also difficult to explain why the NO would be adsorbed as NzOz on the gel and yet only as KO on the non-porous SiO2, with the amounts adsorbed being of the same order of magnitude. It seems more likely that the NO is in some way bound with some of the water which is present in the gel (and which is not present
T'ol. 60
in the non-porous silica) and that it is this waterbound NO which is the cause of the color. After the usual outgassing (300" for 2 hours) a sample of silica gel still retains 3.7% water, as judged by weight loss after further outgassing for one half hour a t 1000°. When exposed to a nitric oxide pressure of 640 mm. a t -78", this sample showed only a faintly reddish color. After the sample had been heated to about 1400" in oucuo it was exposed to a nitric oxide pressure of 550 mm. at -78"; its color was then only an olive green. These qualitative tests do not show conclusively that the red coloration is due to the presence of water, for atJ the same time the water was removed the gel was doubtless sintered to a considerable extent. I t does seem more reasonable, however, to associate this phenomenon with the water than with the presence of pores.
E
GOUY DIFFUSION STUDIES OF BOVINE SERUM ALBUMIN' BY MYRONL. WAGNER AND HAROLD A. SCHERAGh Depurtment of Chemistry, Cornell University, Zthaca, N . Y Received March IS, 1956
In conjunction with studies of the hydrodynamic properties of proteins, diffusion coefficients have been obtained by the Gouy method (making use of Rayleigh interference patterns to determine the total number of fringes). The satisfactor!, performance of the Spinco Model H apparatus for this purpose was established by the excellent precision, and good agreemeit with literature values, obtained in diffusion runs on sucrose a t 1'. The use of the cylindrical lens to obtain a threefold magnification of the Gouy pattern, and the passage of the light beam twice through the diffusion cell, as well as other novel features of the instrument appear to be satisfactory. Accordingly, a study of bovine serum albumin in 0.5 ik? KC1 a t p H 5.14 has been carried out in the mean concentration range of 0.25 to 1.25% as part of a program to relate the hydrodynamic properties of albumin to its reactivity. Extrapolation to zero concentration yielded a value for the diffusion coefficient of 3.261 f 0.004 Fick units (1 Fick unit = 10-7 cm.2/sec.)a t 1" in 0.5 M KCI a t pH 5.14. The effect of heterogeneity on the diffusion coefficient has been discussed. I t is concluded that the presence of 2 to 3% of a second corn onent having a diffusion coefficient 1.5 times smaller than that of albumin is compatible with the diffusion data and also seimentation data on the same samples.
Introduction Recent considerations of the hydrodynamic properties of proteins2 have indicated the need for high precision in order to be able to interpret data obtained from measurements of intrinsic viscosity, sedimentation, diffusion, flow birefringence, etc. Appropriate combinations of pairs of data, such as intrinsic viscosity and translational frictional coefficient provide a basis for computing the size and shape of an equivalent hydrodynamic ellipsoid for the protein molecule. Since good precision is obtainable in viscosity measurements, our attention is centered here on the determination of precise translational frictional coefficients of proteins from diffusion measurements in dilute solution. If the solvent consists of more than one con~ponent,it is preferable to obtain the frictional coefficient from diffusion, rather than from sedimentation measurements, since the former technique does not involve the partial specific volume problem (1) This investigation was supported by grant NSF G-507 from the Nat.iona1 Science Foundation and by Grant H-1662 from the National Heart Institute of t h e National Institutes of Health, Public Health Service. (2) H. A. Scheraga and L. Mandelkern, J. Am. Chem. Soc., 76, 179 (1953).
preseiit in sedimentat,ion, as has been discussed re~ently.~ Bovine serum albumin has been chosen for this study because it appears that, its configuration plays an important role i n determining its reactivity ( e . g . , binding of hydrogen and other small ions). At present, the reactivity of this molecule is known to be anomalous a t low and high pH.4,6 These anomalies could arise either from changes in molecular size and shape,4or from reversible formation and breakage of internal hydrogen bonds16or from a combination of both of these effects. It appeared that tJhisquestion could be resolved by an invest,igation of the effect of pH on the size and shape of the equivalent hydrodynamic ellipsoid of bovine serum albumin. Due to aoniplications from heterogeneity7 at low and high pH, and to ( 3 ) H. .4. Scheraga, W. R. Carroll, L. F. Nims, E. Sutton, J. K. Backus and J. M. Saunders, J . Polumer Sci., 14,427 (1954). (4) C. Tanford, J . G. Buzzell, D. G. Rands and 8. A. Swanson, J . Am. Chem. Soc., 77,6421 (1955). ( 5 ) I. MI. Klotz and J. Ayers, Disc. Faraday SOC.,13, 189 (1953). (6) M. Laskowski, Jr., and H. A. Scheraga. J. A m . Chem. SOC.,76, 6305 (1954). (7) (a) H. A . Saroff, G. I. Loeb and H. A. Scheraga, ibid., 77, 2908 (1955); (b) P. Bro, S. J. Singer and J. RI. Bturtevant,, ibid., 77, 4024 (1955).
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