Feb. 5, 1955
NOTES
metal was lost to the walls of the distillation tube, and some remained in the samarium lean fractions which were not redistilled. About 30 g. of high purity samarium metal has been prepared by the method outlined here. Undoubtedly calcium could be used for partial separation of the rare earths by fractional distillation of the metals from a calcium-rare earth oxide mixture. Spedding and Daane6 have shown that lanthanum is quite effective for this type of separation.
the chloride form, w;ts treated with ca. 0.25 M EDTA solution of p H ca. 11. At this pH, H4A may be considered to be largely dissociated to A-4 and thus the resin treated with 0.25 M EDTA of p H 11 might be expected to be principally in an A-4 - HA+ form with A-4 possibly predominating. After the resin had been treated with several volumes of alkaline EDTA solution, i t was washed with water to remove excess EDTA. A portion of the resin was then analyzed for chloride. It was found that the resin retained significant amounts of chloride which could be removed only slowly by further washing with EDTA solution.
(5) F.H.Spedding and A. H. Daane, “Methods of Producing Rare Earth Metals as Developed at Iowa State College,” Paper presented at the Annual Meeting of the AIME (Feb. 1954).
813
In a typical experiment designed to demonstrate separation of alkali metals, an 0.41 X 23 cm. column of this resin was pretreated with ca. 2 column volumes of 2.5 X lo-’ M EDTA solution a t PH 10.9. A 1.0-ml. aliquot of a solution containing 0.12 M LiOH, 0.015 M NaOH (Na2a), CslS7 and 0.13 M EDTA a t PH 10 was then added to the column and elution was carried out with the same dilute EDTA Anion-exchange Studies. XIV. The Alkali Metals solution which was used to pretreat the column. The in Ethylenediaminetetraacetic Acid Solutions1B2 effluent was analyzed radiometrically for CS’~’and NaZ2 and spectrographically or flame spectrophotometrically for BY FREDERICK NELSON Li .T RECEIVED JULY 15, 1954 As shown in Fig. 1, cesium appeared in the ef-
Los ALAMOS SCIENTIFIC LABORATORY UNIVERSITY OF CALIFORNIA Los ALAMOS,NEWMEXICO
fluent first, essentially unadsorbed, and was satisfactorily separated from sodium which was slightly retained by the column. It is predicted that rubidium and potassium ions in this medium would also have shown negligible adsorption. The cesium and M + A-4 MA-’ (1) sodium fractions were analyzed spectrographically where M + is the alkali ion and A-4 is ethylenedi- and found to be free of lithium within the limits of aminetetraacetate (EDTA). The concentration the spectrographic method. Satisfactory elution of quotients kmol = m M X - ” m M + m A - I were estimated lithium was achieved with 0.25 M EDTA solution to be kmol = 620 for Lif and kmol = 46 for Na+ a t at p H 4.2. Effluent solutions were analyzed by 20’ in 0.1 M KCl. Under the same conditions neg- flame spectrophotometry.~ Appearance of lithium ligible complexing of potassium ions was assumed. in the effluent was paralleled by a decrease in p H One might also expect negligible complexing of ce- as shown by the dashed line in Fig. 1. This decrease sium and rubidium ions by EDTA. The apparent in PH was delayed as shown in Fig. 1, suggesting formation of negatively charged complexes of lith- that acid ions of EDTA (e.g., H2A’) are adsorbed ium and sodium in alkaline EDTA solutions sug- by the resin as expected on the basis of earlier work gested that their separation from each other and on acid adsorption by “strong base” resins confrom other alkali metals might be accomplished by taining polyvalent anions.8 Adsorption of base can also be demonstrated anion exchange in a manner similar to the separation of the alkaline earths in citrate solutions which with this “strong base” resin in the EDTA form. was described earlier.2 I n this connection it might Thus when the resin was pretreated with a 0.25 M be mentioned that the effect of another chelating EDTA solution near PH 11, reasonably good (total) agent, uramil-diacetic acid4 in the cation-exchange adsorption of LiOH occurred, e.g., ca. 3 4 column separation of alkali metals was recently investi- volumes of a 0.08 M LiOH solution could be passed gated.6 Although uramil-diacetate forms better into the columns before breakthrough of lithium complexes with lithium and sodium than EDTA occurred. When the resin was pretreated with ca. (compare references 3 and 4),the latter was chosen 0.25 M EDTA solution near p H 4.2 adsorption of for the anion exchange experiments because of its lithium hydroxide also occurred, although some lithium appeared in the effluent almost immedieasier availability. The experiments were carried out with a strong ately. These experiments suggest that adsorption base quaternary amine polystyrene divinylbenzene of base may occur according to reactions of the type resin (Dowex-1) of relatively low cross-linking Li+ OH- +(Li A-s), (OH-), (2) (4%) and mesh size 50-100. EDTA solutions were or prepared by dissolving ethylenediaminetetraacetic (HA-’), Li+ OH- +(Li A-*)r + H20 (3) acid ( H d ) in KOH solutions. The resin, initially in
The alkali metals, lithium and sodium, have been shown to form reasonably stable complexes with the chelating agent, ethylenediaminetetraacetic acid, a t high pHs according to the equilibrium
+
+
+
+
(1) This document is based on work performed for the U. S. Atomic Energy Commission at the Oak Ridge National Laboratory. (2) Previous paper: F.Nelson and K. A. Kraus, TITISJOURNAL, 76,
801 (1954). (3) G. Schwartzenbach and H. Ackermann, Relo. Ckim. A d a , 80, 1798 (1947). (4) G.Schwartzenbach, E.Kampitsch and R. Steiner, ibid., 99, 364 (1946).
(5) W.Buser, ibid., 9.4, 1635 (1951).
+
+
-
-
(6) p K values for the ionization constants of HIA in 0.1N KC1 (20’) are reported as PKi = 1.996,PKa 2.672,PKr 6.161 and p K c 10.262(ref. 3). See also F.F. Carini and A. E. Martell, THISJOURNAL, 7 4 , 5745 (1952). (7) W e are indebted to Mr. M. Murray of the ORNL Analytical Division for the spectrographic analyses and to Mr W. R. Laing of the same Division for the flame spectrophotometric analyses. (8) (a) K.A. %am, F. Nelson and J. F. Baxter, THIS JOURNAL, 76, 2768 (1953); (b) F. Nelson and K. A. Kraus, ibid., 76, 329 (1954).
8 14
VOl. 77
COMMUNICATIONS TO THE EDITOR
-
2.5 x
4c3 M
. I‘-
EDTA
0 . 2 5 M EDTA pH -4.2
p H = 40.9
cs
-
Li
4 10
9g
\\
8 7
6
5 25
30
35
40
VOLUME OF E F F L U E N T (ml.).
Fig. 1.-Separation
of alkali metals by anion exchange.
which are analogous to the acid uptake described earlier .8 Acknowledgment.-The author is indebted to Dr. K. A. Kraus for helpful criticisms and advice
and t o Mrs. L. W. Magnusson, Jr., for valuable technical assistance.
oAgR~~~~N~~~~~~~L~~~~~~~~~ OAKRIDGE,TENNESSEE
COMMUNICATIONS T O T H E E D I T O R THE ACTIVATION OF CHYMOTRYPSINOGEN
10-30 fold molar excess with respect to active diisopropylphosphofluoridate chymotrypsin of (DFP). Free peptides were isolated by applying tion2 of chymotrypsinogen is accompanied by the to an ion exchange column (0.9 X 15 cm.) of XE-64, formation of several peptides, probably arising 150-400 mesh, Na+ form’ either the activation from a common precursor, containing 8 or 9 mixture in the presence of DFP, or the fraction different amino acid residues. We now wish to soluble in 10% trichloroacetic acid (TCX). Elureport the isolation and structure of a dipeptide tion with 0.3M sodium citrate buffer, PH 5.3, which is probably the only peptide liberated in yielded a sharp peptide peak which emerged with stoichiometrically significant amounts during the the same effluent volume as arginine. The yield of peptide was approximately 0.8 mole per mole of activation of chymotrypsinogen. Electrophoretic analysis, end group a n a l y ~ i s , ~chymotrypsinogen activated (molecular weight specificity requirements of the activating enzymes 23,000). Acid hydrolysis of the peptide yielded and other considerations1 suggested that a basic approximately equal amounts of serine and arginpeptide was liberated during the conversion of ine. Analysis of the acid hydrolyzate of the entire n--chymotrypsin to the 6-form. Hence activation TCA-soluble fraction yielded the same two amino mixtures were prepared under conditions yielding acids in stoichiometrically significant amounts, addition to much smaller amounts of numerous preponderantly 6 - c h y m o t r y p ~ i n . ~The ~ ~ products in amino acids. When the peptide was converted of activation were characterized by activity to the dinitrophenyl derivatives and subsequently measurements (esterase6) and by electrophoresis in hydrolyzed, only DXP-serine was obtainedg on acetate buffer, pH 4.97, usually in the presence of a paper chromatograms, indicating that the peptide (1) H . Keurath, J. A. Gladner and E. W. Davie, in W. D . McElroy had the structure seryl-arginine. The same pepand B. Glass, “The Mechanism of Enzyme Action,” John Hopkins tide was observed by paper chromatography as Press, Baltimore, Md., 1954. product of the slow activation of chymotrypsino( 2 ) M. Kunitz and J. H. riorthrop, J . Gen. Physiol.. 18, 433 (1935). gen, whereas it was absent under conditions yielding (3) R D . Wade and W. J . Dreyer, unpublished experiments. (4) F. R. Bettelheim and H. Xeurath, J . Bid. Chem., 2 1 2 , 241 predominantly a-chymotrypsin, thus indicating (19.56). that i t could only have arisen during the n--6
Sir : It was previously reported’ that the slow activa-
3v4
( 5 ) C. F. Jacohsen, Compt. rend. Irav. Lab. Carisberg, Serie chim., 26, 325 (1Y47). (6) S. Kaufman, H. Iieurath :%nd G. W. Schwert, J . B i d . Chem., 177, 493 (1949).
(7) C.H W. Him, S Moore and W H Stein, rbrd , 200, 493 (1953) (8) F Sauger, Bmchem J , 39, 607 (1945), 46, 126, 563 (1949) (9) S. Blackburn and A F Lowther, rbrd , 48, 126 (1951)
