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INDIANA UNIVERSITY
SO TRS
Contributions to Knowledge Made by In- ~ structors and Advanced Students of the University
VoLUME I
Nos: I-18a-— Jury, £910—-Jury, 1913
nial Mstitge: a F
SA? aS
Nov 10 923
BLOOMINGTON, INDIANA Published by the CUnibersityp
£913
The ‘University Studies’ constitute a sub-series of the INDIANA UNIVERSITY BULLETIN in which from time to time are published some of the contributions to knowledge made by instructors and advanced students of the University. At present not more than two or three such numbers are issued a year.
In the present volume continuous pagination ceases with page 219; hence in the following table of contents no page reference is given for the last four contributions. They are listed, however, in the order of ther publication, and of their appearance in this volume.
-_ .
(9)
10.
Hat
14.
TABLE OF CONTENTS
A Note on the Pharmacalogical Action of Uranium. By DENNIS
DRIENGINSON: AIG EORANIX © MDAININ. .0c.5-0e0 ccd esl 'b soos Canelnecete pete 6
The effect of Starvation for Five Generations on the Sex-Ratio of Drosophila Ampelophila. By CLAauDE DUVALL HolMEs.......
Studies on Perchloric Acid (1): The Preparation of Perchloric Acid from Sodium Perchlorate. By FRANK C. MATHERS......
Studies on Perchloric Acid (II): Electrodeposition of Lead from Rerchlioricsbaths, « By Weank: ©C..MATHERS 925. oe. 3 i. Se
Studies on Perchloric Acid (III): Mercurous Perchlorate Volta- meter. By FrRaNK G. MATHERS and A. F. O. GERMANN.......
The Preparation of Ammonium-Selenate: A New Method. By HeNKee eS NTATEERS and ROW SN. -BONSIB: sce o.oo sw be 2 tse
A Centinuous Function Having Nowhere a Derivative. By Rarn- SRY « LESIBINFITCOUN eed 00) 553) BS Persea aaeen ye eR as rice es are nee ar
The Derivation of Poisscn’s Equation by Means of Gauss’s The- orem of the Arithmetic Mean. By KENNETH P. WILLIAMS...
The Odlitic Limestone Industry in Indiana. By OLIVER C. Lock- PSUR TRIP “eeargithein cot ce Eee Re a nee ceo NE er ee
An Investigation of Housing and Living Conditions in Three Dis- tricts of Indianapolis. By lL. M. CAMPBELE ADAMS...........
An Example of Plagiarism Among Elizabethan Pamphleteers: Samuel Rowland’s ‘Greenes Ghost Haunting Conie-Catchers.’ Ee DNA D 1D SVC ON AUD es ctocwherccocuen soatarscs bane one Dears Sree:
Studies on Perchloric Acid (IV): Distillation of Potassium with SU ple ANGCIds = Biya RANK “(Ge IMUATETERS vances saucrone Sand cre us «cs
Qualitative Separation and Detection of Potassium and Sodium
with Perchloric Acid and Hydrofluosilicic Acid. By FRANK (OF NV ANTES 11ers OG a) By ed BD esl a DE a a tress SN Te
(@)
16
BU
aye)
111
145
173
15. State Banking in Indiana, 1814-1873. By LoGcaw ESABRY........-.
16. Some Results from an Ichthyological Reconnaissance of Colom- bia, South America. Part Il. By Cart H HIcgENMANN.
17. Materials, Methods, and Administration of History Study in the Elementary Schools of the United States. By RoLia MILtTon
TRYON.
Some Results from an Ichthyological Reconnaissance of Colombia, South America. Part I]. By CARL H HIGENMANN.
= D
18a. Wordsworth’s Mind. By RicHarp RICcE, JR.
INDIANA UNIVERSITY BULLETIN July, 1910
Vou. vill, No. 7
“INDIANA UNIVERSITY oe STUDIES
CONTENTS
1. A Nore on THE PHARMACOLOGICAL Action oF Uranium. By _ Dennis E. Jackson and Frank C. Mann. } “Tue Errect of SrarVATION FOR Five GENERATIONS ON THE SEx-
Bere Ratio or Drosopnita AmpetopHita. By Claude DuVall _ Holmes.
- 3. Srupres on PercHLoric AcID: THE PREPARATION OF PERCHLORIC Bae Acip FROM SopIuM PERCHLORATE. By Frank C. Mathers.
4 Srupies on PercHLoric AciD: ELECTRODEPOSITION OF LEAD FROM . PERCHLORATE BaTHs. By Frank C. Mathers.
5. Srupres on Percuioric AciD: Mercurous PERCHLORATE VOLT- ameter. By Frank C. Mathers and A. F. O. Germann.
26; Tue PREPARATION OF AMMONIUM-SELENATE: A New MerHop. By Frank C. Mathers and Roy S. Bonsib. _ :
7. A Continuous Function Havinc NowHERE A Derivative. By Rainard Benton Robbins.
8. Tue DerivaTIon oF Poisson’s Equation py Means or Gauss's THEOREM OF THE Arrrametic Mean. By Kenneth P. Williams. — | | eS
att
th
INDIANA UNIVERSITY STUDIES
Nos. 1 to 8 BLOOMINGTON, INDIANA Juty, 1910
Prefatory Note
The main purpose of the INDIANA UNIversity BULLETIN is the publication of the official statements of the various subdivisions of the University. including their announcements of courses. From time to time this publication has further been used to present ad- dresses given at the University, educational studies which are of value to teachers of the State, and other matters which may inter- est the University alumni and its patrons.
Hitherto no attempt has been made to include in the BULLETIN papers embodying researches made in the various laboratories and departments of the University. These have been left to find their way into print through other channels than the official University publication. With this issue of the BULLETIN a sub-series is estab- lished, to be known as the ‘University Studies,’ in which will be published from time to time some few of the contributions to knowledge made by instructors and advanced students of the University.
At present it is not intended to issue more than two or three such numbers a year, but it is hoped ultimately to be able to enlarge the scope of the series. The ‘Studies’ will be continuously num- bered, and from time to time a title-page and table of contents will be issued, for binding them up in vclumes. The size page of the BULLETIN is changed with this issue to suit better the character of the new publications.
For information concerning the ‘University Studies,’ address
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2 ES DAVIN, A UNE IBS RSME GE NY >
1. A Nore on tHE PHARMACOLOGICAL ACTION OF URANIUM.
By DENNIS IiMERSON JACKSON, Assistant Professor of Physiology, in col- laboration with FRANK CHARLES MANN.
Introduction. The various compounds of uranium have never occupied a prominent place in the field of practical therapeutics. The element possesses marked toxic properties, and but few scien- tific suggestions have ever been made econeerning the utilization of nny of its pharmacological actions in the treatment of disease. |
A brief survey of the literature is sufficient to convinee one that the physiological actions of this substance have by no means been completely determined. Perhaps the most complete and satis- factory study of the physiologica! action of uranium which has yet appeared was published in 1890 by Woroschilsky'. Valuable work was also done by Chittenden? and his collaborators in 1885-6. Recently some attention has been given to uranium as a convenient agent for the production of certain forms of experimental edema’.
This article will deal with only three general phases of *he pharmacological action of this metal, viz., first, the action of the substance on the secretion of lymph; second, its action on the eir- eulation and respiration; and third, its action on the blood.
If sodium uranate be added to a strong aqueous solution of tartaric acid a soluble compound of uranium will be formed which may be used for intravenous injections without precipitation of the blood. It is usually necessary to add a small amount of sodium hydrate in order to neuiralize completely the tartaric acid after the desired amount of sodium uranate has been added. It is, how- ever, probably shehtly more convenient to proceed as did Woro- schilsky in making the double salt. Nearly all of the following ex- periments have been performed with solutions made up after this method. It consists in heating uranium nitrate in a crucible until all the acid is driven off and the reddish-brown oxide (UO,) has been formed. ‘The oxide is then carefully weighed and from this weight the percentage strength of the solution is caleulated. The oxide is placed in a small amount of distilled water, heated. and small quantities of tartaric acid are added from time to time until the oxide is dissolved. The slight excess of tartaric acid which is usually found to be present is neutralized with sodium hydrate, using phenolphthalein as an indicator. The solution is then diluted
1 Woroschilsky, Arbeiten des Pharmakologischen Instituts zu Dorpat, V, (1890), 1.
* Chittenden, Studies from the Sheffield Scientific School, 1, II, III, 1885-6.
3 Pearce, The Archives of Internal Medicine, Jone, 1909.
PHARMACOLOGICAL ACTION OF URANIUM 3
with distilled water until the desired percentage is obtained. The neutralized solution is of a bright golden-yellow color. I have used im nearly all cases a two per cent solution of the red oxide (UO,), the percentage being: based on the relation of the weight of the oxide used to the amount of the solution when changed into the double salt (sodium-uranium-tartrate) and completely diluted. <A solution made up in this manner gives no visible precipitate of egg albumen, blood, or serum when added to any of these proteid solutions in a test.tube. It possesses moderate antiseptic power, but some fungi grow readily in two per cent solutions of it. If the solution be made slightly acid it gains greatly in antiseptic power, and will also precipitate proteids. Addition of a little so- dium hydrate will again dissolve the proteid precipitate, which is also soluble in an excess of the albumen.
The administration of sufficient quantities of uranium to an animal is usually followed within a few days by albuminuria, glyco- surla, parenchymatous degenerations, gastro-intestinal disturbances, paralyses; and, according to most observers, certain marked patho- logical changes are found in the blood vessels.
Action on Lymph Flow. The effect of uranium on lymph flow may best be described by the following protocol of one of my ex- periments:
“Feb. 18, 1910. Dog, male, yellow, weight 18 kilos, in good con- dition. Twelve hours previously the animal had eaten some meat, no: other solid food taken before the experiment. Etherized and placed upon the operating table. Arrangements made for artificial respiration when needed. Blood pressure taken from right carotid artery, injections made into the left femoral vein. Chest opened a little at the apex and a cannula placed in the thoracie duct. Normal lymph flow was 37 drops in 15 minutes, 1.e., 124 drops in each five minutes. Normal time required for clotting of the lymph was 44 minutes. - Lymph was thin and somewhat opalescent, 1.e.. fat- was present but not abundant.
. 11:22. Injected,10 -cé.,of .a two per cent (UO,) solution of sodium uranium tartrate. In the next five minutes 12 drops of lymph were secreted.
11:28.~ Injected 10 cc. of drug. In the next five minutes 16 drops of lymph were seereted.
71:34. Injected 10 ec. of drug. Lymph flow 13 drops in five minutes.
Lymph clotted readily after 30 ec. of drug had been injected. Hence not more than traces of the uranium could be present in an
2—23929
4 INDIANA UNIVERSITY
active form in the secreted lymph, for the addition of a very small amount of the drug to the lymph outside the body prevents clotting.
11:42. Blood pressure fallen about one-half. Muscular tre- mors well marked but weaker than with eserine. Lymph remains clear and opalescent. No noticeable increase in the rate of flow.
11:56. Lymph still clear, no traces whatever of any blood streaks.
11:57. Injected 80 cc. of drug to kill the animal. Urinated, convulsions. tremors. Slight inerease and then a decrease in rate of lymph flow.
12:00. Artificial respiration stopped and animal died of as- phyxia. Was still in fair condition just before death.
In this case observations upon the lymph flow were continued for only about three-quarters of an hour, but the results are per- feetly typical of those obtained in other cases in which observa- tions were carried on for a much longer time. In some instances a very slight increase over the normal is obtained. I believe this to be due not to any specifie action of the drug on the formation of lymph, but rather to the muscular movements of the visceral organs; and in ease of convulsive contractions of the skeletal mus- cles in general these probably also help to force a few extra drops of lymph out of the thoracic duct or its immediate branches. The increased muscular movements are probably partly due to a slight stimulation of the motor endings in striated muscle. There seems also to be a clearly defined but transitory stimulation of certain parts of the central nervous system. This is later followed by marked depression and paralysis. In a few instances slight traces of blood were seen in the lymph as it flowed from the duct. In at least one case the quantity of blood thus observed seemed to in- erease progressively in amount as the intoxication progressed. At no time, however, did the lymph acquire more than a slightly red- dish tinge and the rate of lymph flow was not noticeably increased. Pearce* produced edema in rabbits suffering from uranium nephri- tis when he gave them 100 ee. of water by stomach daily for two or three days. He assumed that the uranium had injured the capil- lary epithelium and concluded that the combination of these three factors, nephritis, injury to the capillary walls, and the adminis- tration of water was sufficient to produce a well-marked edema in rabbits. His experiments extended over a number of days, and it is quite probable that in that interval pathological conditions
4 Pearce, loc. cit.
e
PHARMACOLOGICAL ACTION OF URANIUM »)
might develop which would be entirely absent in an animal which was killed by uranium in a relatively short period of time. In the present instance I have not attempted to do more than to study the immediate action of the drug. It may be said, however, that im animals which die within an hour or so after the first injection of uranium numerous small ecchymotic patches may be seen in the intestinal mucosa. It is probable that in some eases such an action as this might lead to the formation of lymph containing slight traces of blood. If the capillary epithelium is weakened sufficiently to allow the escape of a considerable amount of fluid from the blood then this fluid is not, at least within the first two or three hours, passed out of the thoracic duct. The significance which this may have in relation to the production of edema when water is administered to an animal after two or three days’ treatment with uranium I do not eare to discuss.
Action on the Circulation. Woroschilsky® observed a rise in blood-pressure which he attributed to stimulation of the vaso-con- strictor center. I have corroborated his results and find that with moderate injections (6 to 10 ce. of two per cent solution) the rise is at first only of a transient character. With repeated admin- istrations, however, there is gradually developed a tendency for the pressure to maintain an elevation slightly above the normal (Fig. 1). This seems to be at least partly due to a slight, con- tinuous, but gradually increasing stimulation of the vaso-constrictor center in the medulla. This stimulation is probably due in part to a sort of compensatory (asphyxial) action by which the circula- tion tries to counterbalance the gradually progressing depression of the respiratory center. The increased volume of blood caused by the rapid introduction of eight or ten cubic centimeters of fluid into the circulation usually produces a slight rise of blood- pressure lasting about as long as the injection continues. I have observed that this elevation is often followed by a slight fall which is also transient and may amount to twenty or twenty-five milli- meters (Hg manometers) (Fig. 1). I believe this fall is mainly due to a direct depressant action on the heart, for it also occurs after section of the vagi, and sometimes I have noted after large injections that the heart may weaken and miss one or two beats and then gradually regain its normal strength. This may also occur after section of the vagi. Seven or eight cubic centimeters of a two per cent solution (UO, changed tc the double salt) gen- erally produce a rise of about fifty millimeters of mereury in a ae Woroschilsky, loc. cit., p. 28.
b INDIANA UNIVERSITY
medium sized dog. Electrical stimulation of a vagus nerve stops the heart after repeated large injections of the substance. Inci- dentally it may be mentioned that the secretory endings..of the ehorda tympani in the submawillary gland; the endings ~of the phrenies and the motor endings of the sciatic remain active through- out the intoxication. Injection of a two per cent solution of ura- nium nitrate causes a marked transient fall in blood-pressure which ean scarcely be distinguished from the action of-amyl nitrite. Ura-
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FIGURE 1. Blood pressure and respiratory tracings from a dog. Injection of 8 ce. of 2 per cent. (UO;) sodium-uranium-tartrate solution. (See text for description.)
nium acetate also causes a lowering of pressure. Disodium tartrate causes no effect upon the circulation aside from the increased vol- ume of fluid in the vessels. Bish.
Action on the Respiration. Injection of moderate amounts 7 to 10 ce.) of two per cent solution of. the double salt caused a very shght increase in both rate and depth of respiration (Fig. ale “The immediate cause of death was respiratory paralysis. U ranium ‘has long been supposed to possess an action somewhat resembling that
PHARMACOLOGICAL ACTION. QF URANIUM (
of hydroeyanie acid’. I find its stimulating effect upon the respira- tion to be much smaller than that of the cyanides. Early in its action the depression of the respiratory center becomes evident. After a time the respiration ceases, but the circulation remains in good condition and if artificial respiration be given the animal may be kept alive for a long time. Normal respiration does not return until after ten to thirty minutes, if it is reinstated at all. The cessation of the breathing ?s usually rather rapid and there is but little tendency toward Cheyne-Stokes respiration, as is often seen with the cyanides. I have repeatedly noticed when the res- piration had become greatly depressed that the injection of another dose of the substance would tend to revive the animal, apparently very much in the same way as the evanides. This is probably due mainly to a slight direct stimulation of the central nervous system, for the effect comes on immediately after myjection of the drug. The remarkable specific depression which this substance exercises on the respiratory center reminds one of morphine. I have regu- larly observed that, after a certain amount of the drug had been administered, the animal would pass into a sort of comatose condi- tion, in which but litle ether would be required to maintain the anzsthesia‘. This may be due to the gradually progressing central paralysis, but it may also be contributed to, in part at least, by some specific action on the blood. The view has been generally held of late years that uranium retards the reduction of the oxyhaemo- globin in the tissues. I shall refer to this point again.
Action on the Blood. Uranium and the cyanides both prevent coagulation when added in sufficient quantity to drawn blood. Is this action the same in each case? I[ have attempted to throw some heht on this question by trying the action of the drugs on certain common ferments.
Tf an aqueous extract of potato peelings be treated with a small amount of tincture of guaiac, an oxidizing ferment from the potato will at once oxidize the guaiae over to a blue compound. If a uranium solution (either the acetate. nitrate, or double salt) be added to the aqueous extract before the tincture of guaiae is poured m, there will be no hindrance whatever to the normal change over to the blue compound. Apparently uranium exercises no inhibitory power at all upon this ferment. If a solution of potassium cyanide
® Kobert, Ueber Cyanmethaemoglobin und den Nachweis der Blausdure, Stutt- gart, 1891. (Lehrbuch der Intoricationen, 2d ed., pp. 94-99.) Geppert, Zeitschrift fiir klinische Medicin, XV, 208, 30T. 7 Meitzer, American Journal of Physiology, XXIII, (1909), 141.
8 INDIANA UNIVERSITY
be added to the aqueous extract before the tincture of guaiac is added, then no blue color will appear at all. Evidently hydro- eyanic acid acts differently from uranium upon this ferment.
When normal blood is treated with hydrogen peroxide a marked evolution of gas is at once produced. If a solution of uranium (either the acetate, nitrate, or double salt) be added to the blood before the hydrogen peroxide is added, the formation of gas is in no wise hindered. Apparently the reaction takes place exactly the same as in normal blood. When blood is treated with a cyanide and then hydrogen peroxide is added, it is generally stated that no gas at all is produced. As a matter of fact, a very little gas is usually formed, but the reaction is entirely different from that produced by the hydrogen peroxide on normal blood or in the presence of uranium. Evidently uranium and hydrocyanie acid manifest different properties so far as this reaction is concerned.
If normal blood be treated with a little tincture of guaiac, a yellowish mixture is obtained, but no blue compound is produced. Addition of a little hydrogen peroxide to the mixture at once causes a great evolution of gas and the development of a deep blue color. The presence of some of the earlier formed yellowish substance gives the whole mixture a deep greenish-blue appearanee. If the experiment be repeated in the same manner, but with the addition of a little uranium solution to the bloed before the hydrogen perox- ide is added, a profuse evolution of gas and the formation of a deep ereenish-blue color is at once observed. The reaction appears to take place exactly as it would if no uranium had been added. When a cyanide is substituted in place of the uranium in the above experiment no gas is formed but the deep greenish-blue color at once makes its appearance. Evidently in this case, so far as the development of the blue compound is concerned, neither uranium nor the cyanides exercises any noticeable inhibitory activity. The formation of the blue compound in this case seems to be separate and independent from the formation of the methemoglobin. In- cidentally these experiments show that the ferment in potato peel- ings which causes the guaiae to be oxidized to the blue compound is by no means so resistant as the corresponding ferment in blood, for cyanides stop the action of the former and not that of the latter.
The general results of these experiments may be summed up in a few words. Cyanides and uranium act differently on some fer- ments, but apparently act similarly (or possibly exert almost no action) on others. Unfortunately these conclusions cannot throw
PHARMACOLOGICAL ACTION OF URANIUM © 9
any light upon the question of the similarity or the difference in the methods by whick cyanides and uranium prevent the coagula- tion of blvod. It is to be noted, however, that uranium does not inhibit the action of these ferments*.
It was shown by Collingwood? that if a two per cent solution of disodium-hydrogen-phosphate (Na,HP,) be added to blood (in any amount up to an equal volume) and the free calcium ions be thus precipitated out, the blood would still clot, but a somewhat longer time (one-half hour) was required for the completion of the process than was necessary in the case of normal blood. The usual test for uranium is the formation of a precipitate when a phos- phate solution is added to a solution ccntaining uranium. It seemed that this coincidence might furnish an opportunity to determine something concerning the action of uranium. Since sodium-urani- um-tartrate does not precipitate dilute calcium chloride solutions, we should expect some sert of proteid combination to be formed by the uranium, or, the presence of the metal may in some way check the action of the ferments. At any rate, if it should be pos- sible to add sufficient phosphate to precipitate all the free calcium in the blood (and still leave an excess of phosphate ions free (?) in the plasma), then the addition of an equal volume of a two per cent solution of disodium-hydrogen-phosphate to blood which has been prevented from clotting by the previous addition of the small- est effective quantity of uranium might cause clotting to take place, this, however, being dependent upon the nature of the uranium action. If it were merely inhibiting the ferments by its presence in solution, clotting ought to oceur. If it were loosely bound to some proteid element, clotting would probably occur. If it should be firmly combined with some proteid absolutely essential to clot formation, clotting would probably not occur.
A series of experiments were carried out to test this point. Perfectly fresh blood was obtained from etherized dogs and was used immediately after withdrawal from the right femoral artery. Tt was found that if a small amount (one-tenth volume of two per eent solution) of sodium-uranium-tartrate be added to fresh blood and clot formation be thereby prevented, then the addition of a two per cent solution of disodium-phosphate (Na,HPO,) in any amount up to one-half of the total volume would not cause clot-
8 Chittenden (loc. cit.) found in general that an extremely small amount of uranium increased, while more decreased the activity of the digestive ferments, the reactions varying somewhat with the different uranium salts used.
? Collingwood, Journal of Physiology, XXXVIII, (1909), Proceedings of the Physiological Society, \xxix.
10 INDIANA UNIVERSITY
ting. Nor would the addition of calcium ¢hloride solution cause the blood to clot, either before or after the addition of disodium phosphate. This would indicate that either the ferments or some of the proteid elements of the blood had been affected by the uranium and that its action is different from that of the oxalates, for the careful addition of a slight excess of calcium chloride causes oxalated blood to clot. Again the action of uranium in preventing clot formation differs from that of the fluorides in at least two particulars. The addition of a small amount of throm- bokinase derived by aqueous extraction from minced spleen and lymphatic glands causes blood treated with potassium fluoride to clot in almost the normal time. ‘The addition of thrombokinase is entirely without effect upon blood treated with uranium. Also, if a small excess of caleium chloride be added cautiously to blood treated with a fluoride solution clotting will occur. As stated above, addition of caleium does not cause clotting in blood containing uranium. It was supposed by Rettger’® that the fluorides act by combining with the calcium to form caleium fluoride which in turn is bound in some way to a portion of the protein. It may be that uranium acts in some such manner as this, but it is to be noted that blood treated with uranium behaves itself in an entirely differ- ent manner toward calcium and thrombokinase from blood treated with fluorides. It may be further mentioned that dialysis causes fluoride blood to clot. I was unable to detect the formation of any true clot in uranium blood after twenty-four hours of dialyzing in running (tap) water. Uranium acts differently from magnesium sulphate, inasmuch as the addition of thrombokinase causes clotting in blood treated with that salt. And further, magnesium sulphate blood may be made to clot by sufficient dilution with salt solution. No amount of dilution will cause clotting in blood treated with uranium.
It might be questioned whether or not disodium phosphate would precipitate out all the uranium when added to blood which had been treated with that metal. If to a Ringer’s or similar salt solution to which a small amount (one-tenth volume of two per cent solution) of sodium uranium tartrate has been added there be further added a slightly larger quantity (one-quarter volume of two per cent solution) of disodium hydrogen phosphate, a marked precipitate will soon be produced. Mainly calcium phosphate comes down at first, but after a little time it appears that the uranium also is precipitated, for if blood be added to the clear filtrate, clot-
1° Rettger, American Journal of Physiology, (1909) XXIV, 406.
PHARMACOLOGICAL ACTION OF URANIUM 1]
ting will occur in approximately the normal time. And if the precipitated calcium and uranium phosphate be washed with a little distilled water, dried and then, after comminution with a little normal salt solution, be added to blood, clotting will occur in about the normal time. It would appear, then, that pure uranium phosphate is sufficiently insoluble in blood to be unable to prevent clot formation. (This also holds good for sodium uranate which is insoluble.) It should be stated that when a solution of disodium hydrogen phosphate is added to a solution of sodium uranium tar- trate the uranium phosphate is not precipitated immediately, but some ten to twenty minutes are required for complete precipita- tion. Consequently, when disodium hydrogen phosphate is added to blood previously treated with uranium it is probable that com- plete precipitation would not cecur for some twenty minutes or longer. But if after a period of twenty or thirty minutes either ealeium chloride, thrombokinase, fibrin ferment (serum from blood clot) or any two (or all three of these) be added, no clot will be formed. The addition of fibrin ferment would seem to indicate that the fibrinogen had been affected, for if all of the free soluble uralium has been precipitated, then the addition of fibrin ferment should produce clotting unless the fibrinogen has been affected. It seems probable, however, that the uranium when first added to the blood rapidly forms combinations with practically all of the pro- teid elements at hand and that even if the excess of free uranium (provided any such quantity should be left over) is later precipi- tated out by disodium hydrogen phosphate, then the uranium-pro- teid compounds are not broken up but still maintain practically their original condition. These compounds are apparently also soluble in excess of proteid, for if a small amount (2 ee.) of the blood treated with phosphate, after uranium, be added to a larger quantity (6 ec.) of fresh blood and the mixture well shaken, clot- ting will be prevented.
It may be objected that in the presence of soluble proteids and dilute salt solutions, such as constitute bleod, the disodium hydrogen phosphate would not precipitate out the uranium in the usual man- ner. But if uranium be added to dilute salt solutions approximat- ing as nearly as possible the quantity and quality of those found in the blood, then the disodium hydrogen phosphate will precipitate the uranium so completely that tne clear filtrate exercises no notice- able inhibitory action upon clot formaticn in fresh blood. If such precipitate is not formed when uranium, and then phosphate, are added to blood, it would appear to be because the uranium has
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12 INDIANA UNIVERSITY
already entered into some sort of combination with the proteids themselves. This is what I believe actually occurs. In this con- nection may be recalied the action of oxalates. which precipitate the free calcium of the blood, but apparently do not form proteid combinations, for addition of a slight excess of calcium chloride again establishes the conditions requisite for coagulation of the blood. It must, of course, not be forgotten that the addition of a two per cent solution of the disodim~n hydrogen phosphate (in any amount up to an equal volume) to normal blood does not prevent coagulation.
It is generally assumed that the cyanides prevent coagulation by inhibiting the action of the ferments concerned in clot forma- tion. If this be the whole truth (which I am inclined to doubt), then the above results would seem te indicate that the action of uranium is much more extensive than that of the cyanides so far as the inhibition of clot formation is concerned. And uranium certainly acts differently from several of the other inorganic com- pounds which are ordinarily used to prevent clot formation.
When a cyanide is added to drawn blood outside the body, a bright red color is at once produced. This is usually described as being the same color as that possesssed by pure arterial blood. In a similar manner if a small quantity (one-twentieth volume of two per cent) of sodium uranium tartrate solution be added in the presence of oxygen or air tu freshly drawn blood, this blood will also rapidly take on a bright red color resembling that of ar- terial blood. To the naked eye cyanide blood and uranium blood look very much alike. It is to remembered, however, that retinal fatigue, and the constant color variations which are generally to be readily observed in freshly drawn blood when exposed to the alr (and particularly if shaken or stirred), render close color dis- tinctions in such eases as this very difficult to make. It would appear that the presence of a certain amount of available oxygen is absolutely necessary for this color reaction to take place when the blood has been treated with uranium. In order to test this point I tried the following experiment. In an etherized dog the external jugular vein was dissected out and clamped with a ser- resfin. The venous blood (dark) soon accumulated in the vein above the serresfin. A small glass-barreled hypodermic syringe was then partly filled with uranium solution, care beimg taken to exclude all air bubbles. The point of the syringe was then passed into the swollen vein and blood was drawn directly into the syringe until the barrel was three-fourths full. The point was then with-
PHARMACOLOGICAL ACTION OF URANIUM ikS
drawn and a little of the uranium solution was again drawn into the syringe. Jn this manner but very little opportunity was af- forded for air to enter the syringe, which was filled with dark venous blood diluted with uranium solution. It was found that, so long as no air entered the syringe, the blood remained dark and very venous in appearance. But so soon as the contents of the syringe were emptied out into an open test tube and well aerated, the bright red arterial color at once appeared. This ex- periment is significant, inasmuch as it would appear to indicate that oxygen can readily pass through the walls of the red cor- puscles and form a combination with the hemoglobin within. Since it is not possible to detect any difference spectroscopically between normal blood and biood to which uranium has been added, it there- fore becomes difficult to see how uranium could retard the reduc- tion of oxyhxmoglobin so far as the blood itself is concerned. For if oxygen can readily pass into the corpuscles, and no special com- bination between the uranium and the hemoglobin is formed, then it would seem that the oxygen might also readily pass out of the corpuscles again.
The bright red color which blood treated with uranium as- sumes in the air is not appreciably affected by the addition of disodium hydrogen phosphate. It seems probable that the reten- tion of this color is, partly at least, due to changes in the proteids of the red cells. For the color of iaked blood is not materially influenced by the addition of uranium.
If a sample of fresh blood be treated with uranium and diluted to a one or two per cent solution, and left standing in a stoppered bottle, it will become dark and show decomposition within one or two days. If a similar sample of blood be treated with a solution of potassium cyanide of corresponding strength and then be sim- arly diluted and left standing, it will retain its bright red color for several weeks.
It seems that no chemical combination whatever is formed be- tween uranium and the hemogiobin of the blood. In order to test this I made a long series of observations both with the spectroscope and with the diffraction grating. In the latter case photographic records were made'!. I was unable to detect by either method any difference between normal blood diluted to one per cent or one-half per cent and blood which had been first treated with uranium and then diluted to one or one-half per cent. A comparison was also
11] am greatly indebted to Prof. R. R. Ramsey of the Department of Physics of Indiana University for much valuable assistance in making the photographic observations.
14 INDIANA “UNIVERSITY
made between blood treated with uranium and blood to which potassium cyanide had been added.
The spectrum of cvanhemoglobin differs but little from that of oxvhemoglobin, but a longer time is required for its redue- tion by ammonium sulphide. Perhaps the reduction of blood treated with uranium may be sheghtly retarded, but considering the difficulty attendant upon the determination of the completion of the reduction process one would seareely be justified in saying that the time was prolonged.
If a sample of methemoglobin be made pe the addition of either iodine or potassium ferricyanide to fresh blood and then the mix- ture be diluted to a one per cent solution, the addition of potassium eyanide will at once cause the formation of eyanmethaemoglobin”’, the spectrum of which very closely resembles that of reduced hzemo- globin. But if to a dilute solution of methemoglobin, which has been made by the addition of either iodine or potassium ferricy- anide to fresh blood, there be added a solution of sodium uranium tartrate, no change whatever can be made out in the spectrum of the methemoglobin.
This seems to show that uranium does not form any combina- tion with methemoglobin, and it certainly indicates that the ac- tion of the metal is different from that of the ecyanides so far as the hemoglobin of the blood is concerned. The addition of diso- dium tartrate does not produce any change in the spectrum of normal blood.
Conclusions. (1) The intravenous injection of a solution of sodium uranium tartrate in any quantity up to the lethal dose does not produce a noticeable increase in the rate of lymph flow from the thoracic duct in the dog.
(2) The rise in blood pressure produced by the intravenous injection of a solution of sodium uranium tartrate into a dog is of a much more proncunced and prolonged character than the rise produced by injection of a corresponding quantity of a cyanide.
(3) The stimulating action which uranium exercises upon the respiration is vastly less vigorous than that manifested by the eyanides. :
(4) The method by which uranium prevents the coagulation of blood appears to be different from that exercised by most other substances and probably consists in the formation of a close direct combination between the metal and some one or more of the proteid
12 Kobert, Maly’s Jahresbericht, 1891, 443;