SYNTHESIS OP GLUCOSE DERIVATIVES

OF SOME BARBITURATES

DISSERTATION

Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University

By

GLENN ARTHUR PORTMANN, B.S., M.Sc

******

The Ohio State University 1957

Approved by;

Adviser College of Pharmacy ACKNOWLEDGMENT

The author acknowledges his adviser Professor Loyd

E. Harris for suggesting this problem and for the assistance he has rendered during the course of my education at The Ohio State University. I would also like to acknowledge the help of my fellow graduate stu­ dents and the financial assistance of The American Founda­ tion for Pharmaceutical Education.

ii TABLE OP CONTENTS

Page

STATEMENT OF P ROBLEM...... 1

INTRODUCTION ...... 2

HISTORICAL ...... 18

EXPERIMENTAL...... 20

I Synthesis of Tetra-0~acetyl-OC.-D-glucopy- ranosylbroraide ...... 20

II Nitrogen Glucosides ...... 21

A. Synthesis of 5,5-DiethyI-l,3-di-^ -D- glucopyranosylbarbituric Acid ...... 22

B. Studies with Different Reaction Media . . . 29

C. Synthesis of 5-AlIyl-5-(l-methylbutyl)-, and 5-Ethyl“5-phenyl-1, 3-di- -D- glucopyranosylbarbituric Acid ...... 30

D. Synthesis of 9-Ethyl-5-isoamy1-1-jS -D- glucopyranosylbarbituric Acid ...... 32

E. Synthesis of 5-Ethyl-5-(l-methylbutyl)-I, 3-di-y0 -D-glucopyranosy 1-2-thiobarbi- turic A c i d ...... 33

III Oxygen Glucosides and Ethers ...... 35

A. Attempted Synthesis of Substituted Dialuric Acid Glucosides and Ethers ...... 35

B. Attempted Synthesis of Enolic Ethers and Glucosides of Substituted Barbituric A c i d s ...... 38

C. Synthesis of 5,5-Diethyl-4-(tetra-0-acetyl-^ -D-glucopyranosyloxy)-2, 6-pyrimidinedione . 4l

IV Pharmacological Testing ...... 43

V Analysis of Infrared Spectra ...... 44

iii iv

Page

DISCUSSION...... » . . . . 53

SUMMARY ...... 57

AUTOBIOGRAPHY ...... 60 LIST OP FIGURES

Figure Page

1. Infrared Spectrum of 5,5-Diethyl-4-(tetra~0~ acetyl-^ -D-glucopyranosyloxy)-2, 6- pyrimldinedione...... , . . 46

2. Infrared Spectrum of 5,5-Dlethylbarblturlc Acid ...... 47

3. Infrared-Spectrum of 5,5-Diethyl-l, 3-di- (tetra-0-acetyl-/S -D-glucopyranosyl) barbituric A c i d ...... 48

4. Infrared Spectrum of 5,5-Diethy1-1, 3-di-Æ -D- glucopyranosylbarbituric Acid ...... 49

5. Infrared Spectrum of 5-Ethyl-5-(l-methylbutyl) -2-thiobarbituric Acid ...... 50

6. Infrared Spectrum of 5“Ethyl-5-(1-methylbutyl)- 1, 3“di-(tetra-0-acetyl-y$f-D-glucopyranosyl)- 2-thiobarbituric Acid ...... 51

7. Infrared Spectrum of 5-Ethyl-5-isoamy1-1- (tetra-0-acetyl-/Gp-D-glucopyranosyl) barbituric A c i d ...... 52

V STATEMENT OF PROBLEM

Many glycosides have very pronounced and widely varied pharmacological effects. The type of pharmacological response depends on the specific structure of the aglycone but the sugar moiety contributes important absorption and distribution characteristics.

Glucose and some of its metabolites will cause a prompt return to anesthesia if injected into various animals immediately after recovery from barbiturate anesthesia.

Also glucose will significantly prolong the sleeping time if injected with the barbiturate. The question arises as to what effect glucose would have if chemically combined with barbiturates.

Further, the only method which has been used, in the past, to prepare water-soluble barbiturates is salt forma­ tion, All of these salts have the undesirable property of forming alkaline aqueous solutions due to hydrolysis.

Glucose derivatives of barbiturates should therefore be desirable because of their stability and water solubility.

The purpose of this study was to prepare glucose derivatives of barbiturates with the expectation of form­ ing a more potent, stable, water-soluble barbiturate. INTRODUCTION

Many investigators have studied the effect of glucose and its metabolic products on barbiturate anesthesia.

Lamson and his co-workers (l) were the first to publish their observations made when glucose was injected into dogs which recovered from pentobarbital anesthesia. They found that the dogs returned to anesthesia and remained in this state for an average time of one hour. Upon subse­ quent injections of glucose, the duration of anesthesia decreased until eventually further anesthesia could not be produced by the injection of glucose solution. This pre­

liminary investigation was extended by Lamson and co­ workers (2,3). They observed that other substances such as products of glycolysis and the Krebs Cycle, vitamins, malonic acid, glycerine, and epinephrine would also potentiate barbiturate anesthesia as indicated by a return

to sleep on intravenous injection. Sodium lactate, pyruvate,

and glutamate definitely affected the rate at which

(1) P. D. Lamson. M. E. Greig, and B. H. Robbins, Science, 110. 690 (1949).

(2 ) P. D. Lamson. et al,, J. Pharmacol. Exptl. Therap., 101, 460 (1951).

(3) P. D. Lamson, et aL, Ibid., I0 6 . 219 (1952).

2 3 barbiturates entered the brain but glucose had little or no influence on the rate of entry. Acetylcholine completely blocked the action of sodium lactate.

Adams and Larson (4) also investigated the effect of sodium succinate, malonate, acetate and citrate on the duration of pentobarbital anesthesia and found all of the ions to be potentiating agents. However, Westfall (5) stated that pyruvates are antagonistic to pentobarbital anesthesia and believed that the antagonism was due to polymers of the acid which form very readily, Bester and

Nelson (6) found that sodium citrate, lactate, acetate,

succinate, and pyruvate produced the symptoms of alkalosis but did not induce sleep in rabbits that had been anesthe­

tized with pentobarbital as determined by response to

stimuli. The discrepancy between their results and those

of Lamson, et al was explained as probably being due to the

use of different criteria in deciding whether the animals

were anesthetized.

(4) J, P* Adams and E, Larson, Fed. Froc, 253 (1950),

(5) B, A, Westfall, Anesthesia and Analgesia, 28, l6l (1949).

(6) J, F, Bester and J, W. Nelson, J, Am, Pharm, A, (Scientific Edition), 42, 421 (1953). 4

Previously, Bester (7) substantiated the potentia­ tion of pentobarbital anesthesia by glucose. He also found a significant increase in sleeping time when glucose was injected with pentobarbital, and that normal blood glucose levels did not have an apparent affect on the duration of anesthesia following pentobarbital administration. A com­ petitive inhibition mechanism was proposed by Bester and

Nelson to explain this potentiation phenomenon. Since glucose metabolism and barbiturate detoxification probably involve the same oxidative enzyme system, an increase in

concentration of glucose could delay oxidation of the barbiturate causing increased blood and brain levels. The depressant action of barbiturates upon the central nervous

system is brought about by inhibition of glucose utiliza­

tion. By injecting glucose, nerve tissue metabolism would be increased requiring the presence of greater amounts of barbiturate to inhibit this increased metabolism and cause

anesthesia. However, if the increase in barbiturate blood

level is great enough, the over-all effect would be a

prolongation of barbiturate anesthesia.

Recent work (8) indicates that barbiturates may

block the Cytochrome Oxidase-Cytochrome-C system. It was

observed that rabbits given lethal doses of pentothal would

(T) J. P, Bester, M, S, Thesis, The Ohio State Univer­ sity (1950), also J. W. Nelson, and J, P, Bester, Can, Pharm. J, 8^, 22 (1952), (8) B. Giovanni, Giorn. ital, chir, IJ^, 15OO (1955), through Chem, Abs, ^0^ 9622 (1956). 5 survive if Cytochrome-C was injected intravenously either mixed with or given immediately after the drug. Pretreat­ ment with Cytochrome-C had no effect.

Ratcliff, et al (9) found that, in insulin treated animals, there was a need for more frequent administration and greater quantities of pentothal to produce deep surgical anesthesia than was required in animals not given insulin.

Whether the fall in blood glucose level caused by insulin is directly related to potentiation by injection of glucose is not known. Many other compounds will prolong the hypnotic effect of barbiturates. Some of them are iodides, bromides

(10), antihistamines (11,12), cholesterol (13,14),

(9) C. M. Ratcliff, et al. Fed, Froc. 8, 326 (1949). (10) J, C, Krantz, Jr, and M, J, Fassel, J. Am. Pharm. A. (Scientific Edition), 511 (1951).

(11) H. Lightstone and J, W. Nelson, Ibid.. 43. 263 (1954), (12) C. A, Winter, J, Pharmacol. Explt. Therap., 24, 7 (1948).

(13) E, Starkenstein and H. Weden, Arch. Explt, Pathol, Pharmakol,, 182. 700 (1936).

(14) C. B. deFarson, C, J, Carr, and J. C. Krantz, Jr., J. Pharmacol, Exptl, Therap,, 82.» 222 (1947). 6 thiamine (15), nitrites (16), and uracil (17). Their mechanism of potentiation is not known.

One of the purposes of the present investigation was to observe what effect glucose would have on the hypnotic properties of barbiturates when chemically combined with them. A survey of the literature was made to determine how the glucose moiety affected the pharmacological properties of other drugs and to determine if any generalizations could be made. The pharmacological properties of the glucosides had been determined in many cases but a comparative study of the aglycone could not be found or its activity had been determined by different methods. The type of pharmacologi­ cal response depends on the specific structure of the agly­ cone but the sugar moiety contributes important absorption and distribution characteristics.

An excellent example of the sugar moiety changing distribution characteristics is the well known Cardiac or

Digitalis Glycosides, Their steroidal lactone aglycones possess a very low fixation power for the heart and there­

fore have only a very transitory action (18), Hence, for (15) B, DeBoer, J, Am, Pharm, A. (Scientific Edi­ tion), 21, 302 (1948). (16) H, A, Wooster, Jr, and P, W, Sunderman, J, Pharmacol, Exptl, Therap,, 9%, 140 (1949).

(17) D, G, Wenzel and M, L. Keplinger, J. Am, Pharm, A, (Scientific Edition), # , 56 (1955).

(18) Arthur Grollman, Pharmacology and Therapeutics, Lea and Febiger, Philadelphia, 1954, p. 333. 7 medicinal purposes, the glycosides which have a special

affinity for cardiac muscle are preferred. All the aglycones

of the cardiac glycosides, so far known, possess a secondary

hydroxyl at position 3 and a tertiary hydroxyl at position

14 and, in all except one, the glycosidic link is at the

C-3 hydroxyl (19). Thirteen different monosaccharides have

been isolated by hydrolysis and the only common ones are

D-glucose, L-rharnnose, and D-fucose.

Both natural and artificial glycosides have been

synthesized from the natural cardiac aglycones. Elderfield

and Uhle (20) synthesized some monosides of natural cardiac

aglycones containing five-membered lactones. These glyco­

sides are not only more potent than their respective

aglycones but also more potent than the natural glycosides

which were either di- or trisaccharides.

Leinzinger (21) investigated the pharmacology of

morphine glycosides. The acetal bond was formed through the

3 position of morphine where there is a phenolic hydroxyl.

The following derivatives of morphine were prepared and

tested; the glucoside, tetra-O-acetylglucoside, galactoside.

(19) C. M. Suter and P. P. Blicke, Medicinal Chem­ istry, Vol. II, John Wiley & Sons, New York, 1956, p. 25 .

(20) P. C. Uhle and R. C. Elderfield, J. Org. Chem. a, 162 (1947). (21) E. Leinzinger, Pharm. Acta. Helv., 22., II6 (1947). 8 tetra-O-acetyIgalactoslde, lactoside, and hepta-O-acatyl- lactoside. The activity of each was found to be much less than that of morphine and their therapeutic indices were close to unity. Casparis (22) also found diminished activity of the C-3 glucoside, but glucoside formation at the 6 position (secondary alcohol) caused an increase in analgesic activity as measured on a molar basis. Also the

C-6 glucoside had less depressant action on respiration and did not decrease motility of intestinal smooth muscle as did morphine. Other monosaccharides and their acetates produced the same effect as glucose, producing very desirable anal­ gesics because of their increased analgesic activity and lessened side effects.

The nitrogen glucoside of sulfathiazole is reported to be much less toxic and to have slightly less tuberculo­ static activity than sulfathiazole (23). Luquet (24) reported that the di-N-glucoside of arsphenamine was one- half as toxic for rabbits as arsphenamine on a weight basis.

B A L (2,3-dimercaptopropanol), an arsenic antidote, was

(22) P. Casparis, Boll Chim. Farm., 8£, 309 (1950); through Chem. Abs*, 255 (1951).

(23) Y. Miura, J. Biochera (Japan), 31, 205 (1950); through Chem. Abs. 10394 (1951)

(24) A. Luquet, Compt. rend. soc. biol. 81, 1020 (1922). 9 converted to a primary alcoholic glucoside which was found to be less active on a molar basis than B A L in the detoxification of raapharsen and phenylarsenoxide (25)* The nitrogen glucoside of procaine has no local anesthetic action (26). Also, when procaine is dissolved in glucose solution and injected, its local anesthetic power is diminished. Glucosides of o-, m-, and p-cresol, guaiacol, and thymol were synthesized and examined for curarimimetic activity (27). They have one-half to two-thirds the activ­ ity of raephenesin (3-o-toloxy-l,2,-propanediol) on a weight basis. No glucoside of mephenesin was reported.

Glucosides of two hormones have been synthesized and pharmacologically tested. The phenolic glucoside of estradiol is only two-thirds as active as estradiol in rats on a weight basis (28). Desoxycorticosterone glucoside maintained life and growth in young adrenalectomized rats and dogs when given daily by subcutaneous or intramuscular injection (29,

(25) J. A. C. Weatherall and M. Weatherall, Brit. J. Pharm., 4, 260 (1949); through Chem. Abs. M , 4l43 (1950).

(26) C. Sannie and M, Vincent, Compt. rend. soc. biol. 142. 493 (1948).

(27) T, Omiya and N. Pujitoke, Folia Pharmacol. Japan, 42, 84 (1953); through Chem. Abs. ^ 10720 (1953).

(28) H. Lehr and A. Schmitz, Arch, intern, phamaco- dynaraie, 432 (1952).

(29) H. Duell, et al.. Endocrinology, 46, 30 (1950). 10

30, 31, 32). It is one-fourth to one-half as active as desoxycorticosterone acetate on a weight basis. Restitution of adrenalectomized dogs is almost instantaneous with desoxy­ corticosterone glucoside in contrast to that brought about less readily by desoxycorticosterone acetate (33). Also, glucosuria resulting from decreased tubular reabsorption of glucose occurs with desoxycorticosterone glucoside but not with the acetate (34,35).

This survey of the literature shows that glucose does not contribute any consistent change in the varied pharma­ cological activities of the compounds studied.

Since it is important to synthesize the enolic or the nitrogen glucosides of 5,5-disubstituted barbituric acids, it is necessary to understand their cheMstry especially their ability to exist in different forms.

A survey of the literature pertaining to tautomeri- zation of barbiturates revealed that very little is known

(30) W. Fischer and C. Meystre, Helv. Chim. Acta., 2 ^ 40 (1942).

(31) R. Meier, H. Gysel, and R. Muller. Schweiz. Med. Wochschr., 93 (1944); through Chem. Abs. 40^ 6612 (1946).

(32) W. W. Swingle, Am. J. Physiol., 1 ^ , 278 (1952).

(33) R. Meier, Helv. Physiol. Pharmacol. Acta., L, 63 (1943); through Chem. Abs. 3â.» 4613 (1944). (34) A. Despopovlos and E. H. Kaufman, Am. J. Physiol. 120, 11 (1952).

(35) D. M Green, et al.. Endocrinology, ^ 338 (1950). 11 about this important field. Tautomerism occurs whenever the methods of preparation or the reactions (of both) of a compound require the assignment of two or more distinct structures which differ in the positions of at least one atomic nucleus (36). The sequence of reactions shown below suggests the presence of tautomerism (37) in the barbiturates.

ikOÇjtV» o

to , cl C - A/" ^ ^ z=.f\l H.s_;c=o + pou,-— -

O Cl

A tautomeric substance usually appears as a single pure substance since ordinarily only one of the tautomers exists to an appreciable degree in the equilibrium mixture,

Even when two or more tautomers are present in significant amounts, their interconversion is relatively rapid.

(36) Gr. W. Wheland, Advanced Organic Chemistry, John Wiley ft Sons, New York, 1949, p. 581.

(37) A. W. Dox, J. Am. Chem. Soc., 51, 1559 (1931). 12

Therefore each component of the tautomeric mixture can undergo the reactions characteristic of any of its tau­ tomers, since it can be changed very rapidly into that tautomer. Some workers have postulated that enolization occurs with barbituric acid and its derivatives on the basis of differences in the ultraviolet absorption spectra in acidic and basic media (38,39). Since barbituric acid, in alkaline media, has a spectrum which is practically identical to that in acid media, it was concluded that barbituric acid exists in the enolized form. A shift in the wavelength of maximum absorption was shown for barbital and phénobarbital in alkaline media as compared to acid media. However, when an aqueous solution of 5 ,5 -dimethyl- barbituric acid was made alkaline, no change in the spectrum was observed. Stuckey assumed that the negative charge remains on the nitrogen, thus no tautomerism takes place.

Fischer and Dilthey (40) also observed that dimethyl- barbituric acid did not react like other barbituric acids with respect to formation of sodium salts. They observed that 5 ,5 -dimethylbarbituric acid was the only one of a

(38) A. K. Macbeth, T. H. Nunan, and D. Traill, J. Chem. Soc., 1248 (1926),

(39) R. E. Stuckey, Quart. J. Pharm. Pharmacol., 11, 312 (1940); 14i 217 (1941); 11, 370 (1942); 11, 378 11942).

(40) E. Fischer and A. Dilthey, Ann., 335, 34l (1904). 13

series of 5,5 -disubstituted barbituric acids that formed a

stable disodium salt even when one equivalent of sodium hydroxide was present. The anomalous behavior of dimethyl- barbituric acid cannot be explained on the basis of dif­

ference in dissociation because the first and second dis­

sociation constants of barbiturates do not differ appreci­

ably. Disilver salts of diraethylbarbituric acid and other barbituric acids have been reported (41,42,43,44). They were made with ammoniacal silver nitrate or sodium hydroxide

plus silver nitrate or by adding sodium carbonate to a

sodium-silver-barbiturate complex.

Two different complexes of phénobarbital with cobalt

have been prepared (45) which differ in color and stability

to heat. It was postulated that the differences in the

complexes were due to the different tautomeric forms of

phénobarbital and that the enol form changed to the keto

form on heating.

(41) L. I, Rapaport, and G. A, Vaisraan, Ukrain. Khim. Zhur. 20, 424 (1954); through Chem. Abs., 50 , 17318 (1956).

(42) L. I. Rapaport, Ibid.. 20, 430 (1954); through Chem. Abs., 51, 669 (1957).

(4 3 ) P. Viebock and K. Fuchs, Pharm. Ber., 10_, 5 (1935); through Chem. Abs. 2^, 8233 (1935). (44) M. Conrad and M. Guthzeit, Ber., 15^, 2844 (1822)

(4 5 ) E. Vascautanu and R. Cernatescu, Bull. Sect. Soi. Acad. Roumaine, 25,, 459 (1943); through Chem. Abs., 32, 3259 (1945). 14

Speziole and co-workers (46) isolated a mixture of enolic and ketonic secobarbital sodium in the solid state.

Isolation of secobarbital sodium from anhydrous butanol or aqueous ethanol gave two products which differed in some physical properties. The material which crystallized from anhydrous butanol appeared to contain 30.75^ impurities whereas that obtained by evaporation of aqueous-ethanol solution contained 9^ impurities when both were examined by the minimum solubility purity test. The two tautomeric forms apparently differ in their solubilities in anhydrous

2-methylbutanol-2 which was used in the purity test. When the mixture, which was crystallized from butanol, was sub­ jected to the aqueous - ethanol method, a purity test

revealed the presence of only 3.1^ impurities. Infrared absorption spectra showed stronger hydroxyl group absorption in the 2.9 x region with the material from anhydrous butanol. They suggested that the displacement of tautomeric

equilibrium of disubstituted barbituric acids from anhydrous

alkaline media may be general.

Alkylation of 5,5-disubstituted barbituric acids with alkyl halides, diazomethane, diazoethane, and dimethyl-

sulfate results in N-alkylation which indicates reaction of

the barbituric acid derivative in the ketonic or lactam

form. It is possible that the enolic form reacts with

subsequent rearrangement to the N-alkyl derivative. Even ' " (46) A. J. Speziole, et al„ J. Am. Pharm. A. (Scientific Edition), 44, 617 (1955). • 15 if rearrangement does not occur, formation of the N-alkyl derivative is not proof that the lactam form predominates.

The possibility exists that the lactam form may be the minor component but reacts much faster than the predominant

lactim forms.

Direct alkylation of the sodium salts of 5 ,5-dialkyl- barbituric acids with alkyl halides was studied by Dox and

Jones (4 7 ). They obtained N-alkyl and N,N-dialkyl deriva­

tives. Proof of nitrogen linkage was done by unequivocal

syntheses using substituted ureas and substituted malonic

esters and the procedure developed by Fischer and Dilthey

(48). Doran and Shonle found that reaction of the sodium

salt of 5 -butyl-2 -thiobarbituric acid with crotyl bromide

resulted in substitution through the sulfur atom (49).

Possibly the reason for the difference in position of

alkylation between thiobarbituric acid and barbituric acid

derivatives is a greater enolization in the former.

Dox obtained quantitative yields of tetraalkyl-

barblturic acids with diazomethane or diazoethane and

(4 7 ) A. W, Dox and E. G. Jones, J. Am. Chem. Soc., 51 . 316 (1929). (48) E. Fischer and A. Dilthey, Ann., 335. 334 (1904).

(49) W, J. Doran and H. A. Shonle, J. Am. Chem, Soc., 60, 2880 (1938). 1 6

5,5~ciisubstituted barbituric acids (50), Biltz and Wittek

(51 ) obtained the same trimethylbarbituric acid by diazo­ methane treatment of barbituric acid, 1-methylbarbituric acid and 1,3-dimethylbarbituric acid which indicated that both nitrogens are subject to méthylation but only one of the methylene hydrogens,

Dimethylsulfate has been used for N-methylation of barbital in.alkaline solution by Bush and Butler (52),

Substituted thiobarbituric acids react differently than substituted barbituric acids with dimethylsulfate, Lee

(53 ) observed that dimethylsulfate reacted with 5-substituted-

2-thiobarbituric acids to produce the 2-methylthioether derivative. This indicated that thiobarbituric acids may exist in the enol form to a greater degree than barbituric acids. This difference in position of alkylation did not occur with thiourea and urea which produce the Ç-CH^ and mainly the 0-CH3 derivatives respectively with dimethyl­ sulfate (54).

" (50) A. W, Dox, J, Am, Chem. Soc,, 5 8 , 1633 (1936),

(51) H. Biltz and H. Wittek, Ber,, ^4, 1035 (1921).

(52) H. Bush and J, Butler, J, Pharmacol,, 63^, 139 (1937). (53) J. Lee, J, Am. Chem. Soc., §0 , 993 (1938).

(54) E. A.Wemer, J, Chem. Soc,, 105. 923 (I9l4). 17 Only one enolic derivative of 5 ,5 ”disubstituted barbituric acids has been prepared. This was made by reac­ tion of diethylmalonyldichloride with methoxyisourea to produce 5,5"diethyl-2-methoxy-4,6-primidinedione (55 )»

Snyder and Link (56) prepared the same compound using diethylmalonyldichloride. Tautomeric substances usually produce mainly the 0 -alkyl derivative in the presence of silver oxide or silver salts and alkyl halides. This would be another method of preparation of enolic derivatives of barbituric acids,

Dox discovered an interesting reaction between

5 ,5"disubstituted barbituric acids and an excess of

Grignard reagent (57). Two carbonyls reacted with ethyl- magnesiumbromide with subsequent loss of one molecule of water probably with the formation of an internal ether.

With a tetraalkylbarbituric acid, three carbonyls reacted with the loss of one molecule of water.

Further study of barbituric acid chemistry with kinetic and isotope methods should reveal more about the exact mechanisms of reaction,

(5 5 ) Baeyer and Co,, German Patent, 249, 907, P 32,009. (56) J, A, Snyder and K, P, Link, J. Am. Chem, Soc,, II, 1881 (1953). (57) A, W, Dox, J, Am, Chem. Soc,, 42., 2275 (1927). HISTORICAL

A survey of the literature pertaining to glucose derivatives of barbiturates revealed that only three barbiturate glucosides have been prepared. Helferich and Kosche (58) prepared 5,5-diethyT-l-Dnglucopyranosyl- barbituric acid in 13.5^ yield by reacting diethylmalonyl­ dichloride with tetra-O-acetylglucose-urea in the presence of pyridine. Bodendorf (59) prepared 5,5-diethy1-1, 3“Ûi-

(tetra-O-acetyl-^-D-glucopyranosyl) barbituric acid in

20,95^ yield by reacting tetra-0-acetyl-(Xj-D-glucopyranosyl- bromide with the potassium salt of 5 ,5-diethylbarbituric acid in aqueous - acetone media. Deacetylation of this compound was accomplished with a catalytic amount of sodium methoxide in absolute methanol resulting in an amorphous solid. No proof of the nitrogen linkage was presented.

Snyder and Link (60) prepared 5,5-diethyl-4-(tetra-0-acetyl

-^-D-glucopyranosyloxy)-2, 6-pyrimidinedione in 2,6^ yield by reacting 5 ,5 -diethylbarbituric acid with tetra-0 -acetyl

(5Ô) B. Helferich and W. Kosche, Ber. 52., 69 (1926).

(59) K, Bodendorf, Arch. Pharm., 287. 78 (1944).

(60) J. A. Snyder and K. P. Link, J. Am, Chem. Soc., Z5.» 1881 (1953).

18 . • 19 - oC-D-glucopyranosylbromide in the presence of silver oxide and a catalytic amount of quinoline. Indirect proof of glucoside formation at C-4 was obtained by methanolysis with barium methoxide and by comparison of ultraviolet spectra.

The ultraviolet spectra of 5,5-diethyl-2-methoxy-4, 6-pyrim- idinedione, which they synthesized, and the methoxypyrimi- dinedione obtained from methanolysis of the enolic acetylated glucoside were not the same. Therefore, it was concluded that the enol-acetal bond mùst be the C-4 position of barbital. EXPERIMENTAL

I Synthesis of Tetra-0-aoety 1 -OC-D -glucopyranosylhromide

Procedures for the preparation of tetra-O-acetyl

- CXrD-glucopyranosylbromide in 80-90# yields are described in Organic Syntheses (6l) and Biochemical Preparations (62),

Low yields were obtained with the latter procedure. A combination of the two methods was found to be satisfactory.

A known procedure was used to prepare penta-O-acetyl

-^-D-glucose in 72# yield fromo<^-D-glucose, sodium acetate and acetic anhydride (63).

Three hundred and fifty ml. of a 30# solution of hydrogen bromide in glacial acetic acid were cooled to 5°

and added to 200 g. of dry ^-D-glucose pentaacetate

(mp. 132-134°). The resulting red solution was allowed to

remain at room temperature for four hours, after which the hydrogen bromide was removed under reduced pressure using a water aspirator and a mineral oil bath which was kept at

40-50°, Dry nitrogen was passed through the capillary. It

(61) C. E. Redemann and C. Niemann, Organic Syntheses, Vol. 22, John Wiley and Sons, New York, 1942, p. 1,

(62) M, E. Krahl and C. P. Cori, Biochemical Prepar­ ations, Vol. 1, John Wiley and Sons, New York, 1949, p. 34.

(63) Ibid.. p. 33.

20 21

required approximately three hours to remove the hydrogen bromide. Acetic acid was then distilled at a bath tempera­

ture of 55~6o° and a pressure of 15 mm. The dark brown

residue was dissolved in 1500 ml. of dry ether (dried over

sodium wire) and kept at 5° overnight. The resulting white

crystals were filtered and washed with cold ether. Petro­

leum ether (200 ml,) was added to the filtrate to cause

crystallization of the remaining product. The yield was

127.3 g. (60$) having a m,p, of 89-90°,* The reported m.p,

was 89-90° and 87-88°, This product will turn to a black

tarry mass if kept in a desiccator, over phosphorus pent-

oxide at room temperature for approximately one week.

Subsequently:,, it was kept at -10° for several months without

decomposition,

II Nitrogen Glucosides

A, Synthesis of 5 ,5-Diethy1-1, 3-di- B -D-glucopyranosyl- barbituric Acid

Duplication of the procedure of Bcdendorf (64) was

at t empted. Tetra-0 -ac etyl-o(.-D-g lue opy rano sy lb romide,

12,5 g. (0.03 mole), and 2,75 g. (0,015 mole) of diethyl-

barbituric acid were dissolved in 50 ml. of acetone. To

(64) Bodendorf, op. cit,. p, 78. * All melting points were taken with a Fisher-Johns Melting Point Apparatus and are uncorrected. 22 this solution was added 15 ml. of 2,0 M. KOH (0,03 mole)*

After four days approximately one-half of the acetone was removed with a water aspirator resulting in a 2 phase system, the bottom layer of which was syrupy. Crystalli­ zation of the syrup began after two hours and was complete after six hours. The light yellow crystals were filtered, washed with absolute ethanol and recrystallized twice from absolute ethanol. The yield was 1,8 g, (l4^) of fine needles, m,p, 173-174®, [ - 28,7° (C 4.17, chloroform)

Analytical Data**

Calculated for

N, 3.31; Pound; 3.29 Deacetylation of this compound was accomplished with a catalytic amount of sodium methoxide. The acetylated compound, 1,69 g. (0,002 mole) was partially dissolved in

loo ml, of absolute methanol*** contained in a tared Erlen- meyer flask and then mixed with 6 ml, of sodium methoxide

*At this point, Bodendorf observed a crystalline precipitate which increased in quantity up to four days.

Elemental analyses were obtained from Galbraith Mioroanalytical Laboratories, Knoxville, Tenn,

***ijhe absolute methanol was prepared by adding 4 g, of sodium to 200 ml, of commercial absolute methanol and then distilling. 23 solution (1 rag, of sodium per ml. of methanol). Methanol

and methyl acetate were distilled using a steam bath. The

remaining traces of the alcoho^. were removed in a vacuum desiccator resulting in a light brown amorphous substance, yield, 1.03 g. (0.002 mole). After decolorization with

activated charcoal and absolute alcohol, a white brittle

amorphous solid was obtained.

Analytical Data

Calculated for C20H32O 13N2

N, 5 .51; Pound; 5.12

Attempts to crystallize the product from many sol­

vents failed. It did not reduce hot Benedicts reagent but

after hydrolysis with hot 10^ hydrochloric acid for 30

minutes and subsequent neutralization with 5^ sodium hydrox­

ide, it reduced the reagent in 4 minutes.

Deacetylation was also accomplished with ammonia in

absolute methanol. The acetylated compound, 1.27 g . (0,0015

mole), was partially dissolved in 60 ml. of absolute

methanol and saturated with dry ammonia gas at 5° (17 g. of

ammonia). The mixture was kept at 0° for twenty-two hours,

after which the ammonia and methanol were removed under

reduced pressure. Ten ml. of chloroform was added to the

white residue in order to remove the ac et amide. After

filtering, the yield was 0.49 g. (0.001 mole) of a white

amorphous material which did not reduce hot benedicts 2k reagent. It is very soluble in water. Attempts to crystal­ lize the product from many solvents failed.

The glucoside bonds in the diglucoside of barbital could be either through both nitrogen atoms or through two oxygen atoms of the di-enolic form of the barbiturate.

Therefore, an unambiguous synthesis of the acetylated di­ nitrogen glucoside was proposed. The synthetic approach which was chosen involves the following steps: a. synthesis of syra-di-(tetra-0-acetyl-^-D-glucopyranosyl) urea, b. con­ densation of the acetylated diglucose-urea derivative with diethylmalonyldichloride. a. Synthesis of 8ym-di - ( t et ra-0 -ac e ty 1 - Ç -D-g lue opy rano sy 1 ) urea

It was found that the only non-protonic solvents in which urea was soluble are dimethylformamide (IMP) and dimethylsulfoxide (DM8).

DMP (redistilled, 151-152°) was found to be a poor reaction media due to the slowness of reaction as determined by color changes. Because black silver oxide reacts to form yellow silver bromide, the extent of reaction can be determined by color changes. With DMP as the solvent, the color was dark brown after two days.

DM8 proved to be a good reaction media. Ten ml. of

DM8 (redistilled, 189°) was used to dissolve 0.60 g. (0.01 mole) of urea (mp. 136-137°). Ten grams of powdered drierite,

2-.3^ g. (0.01 mole) of silver oxide and 8.22 g. (0.02 mole) 25 of tetra-O-acetyl- QX^-D-glucopyranosylbrcmide were added to the urea solution. Immediately the mixture turned light blue and then green. A magnetic stirrer was used. By test­ ing a few drops of clear solution periodically with silver nitrate test solution, it was shown that all of the acetyl­ ated glucosylbromide had reacted within 12 hours. The yellow-green precipitate was filtered and washed with 20 ml. of dry ether. Ether and IMS were removed from the filtrate by distillation, the IMS distilling at 58° (5 mm.). It was not possible to crystalline the opaque syrupy residue

(3.80 g.) from many solvents. It is possible that diffi­

culty of crystallization was due to the presence of anomeric

forms, b. Condensation of the Acetylated Diglueose-Urea Derivative with Diethylmalonyldichloride

Diethyldiethylmalonate was prepared according to the

procedure of Vogel (05) with slight modifications. The

absolute ethanol for this preparation was made by adding

7 g, of sodium to 500 ml. of commercial absolute alcohol

and distilling 300 ml. directly into the reaction flask.

Sodium, 23 g. (1.0 mole), was added to the 300 ml, of

absolute ethanol. It was necessary to warm the mixture on

a steam bath to cause the last traces of sodium to react.

(65) A. I. Vogel, Practical Organic Chemistry, Longmans Greer & Co., New York, 1951» P. 468. 2 6

Diethylraalonate, 80,0 g., (0,5 mole, redistilled 76-78°/5 mm,) was added dropwise over a period of one-half hour while

stirring with a magnetic stirrer. Ethyl bromide, 132 g.

(1,2 mole), was added dropwise over a period of one hour.

Sodium bromide precipitated during the addition. The mixture

was refluxed for four hours. After cooling to room tempera­

ture, the white solid material was filtered and washed with

30 ml, of absolute ethanol. This alcohol filtrate was used

in the next step,

Diethylraalonic acid was then prepared by saponifica­

tion of the ester with sodium hydroxide. One hundred grams

of sodium hydroxide (2,5 mole) were partially dissolved in

200 ml, of absolute ethanol. The alcohol solution of the

ester was added dropwise to the basic alcohol solution at

reflux temperature over a period of one-half hour. After

the addition, the mixture was refluxed for 2 hours. Two

hundred ml, of absolute ethanol were removed by distillation.

After adding 100 ml, of water to the very thick white

residue, an additional 200 ml, of alcohol was distilled.

After cooling to room temperature, the white residue was

adjusted to a pH of 2 with 150 ml, of concentrated hydro­

chloric acid, filtered and washed with 200 ml, of ether.

The ether layer was separated and the aqueous layer was

extracted with five 200 ml, portions of ether. The ether

extracts were combined, concentrated to 300 ml, and extracted

with three 75 ml, portions of aqueous sodium carbonate 27 solution (30^). The alkaline aqueous solution was then adjusted to pH 3 with I30 ml. of concentrated hydrochloric acid and cooled at 5°. A red oily top layer separated.

The clear aqueous bottom layer was extracted with five 200 ml. portions of ether. The ether extracts were added to the red oil and dried over -anhydrous sodium sulfate. After distilling practically all of the ether, the resulting red oil was cooled to 5°. Crystallization was complete after one week. After recrystallization from chloroform the yield was 15.1 g. (0.095 mole, 19^ based on diethylmalonate), mp. 124-125.° The reported m.p. was 125° (66).

Diethylmalonyldichloride was prepared using phos­ phorus pentachloride. One-tenth mole (20,8 g.) of PCI5 was added in five portions to 8.0 g. (O.O5 mole) of diethyl- malonic acid. After refluxing for 2 hours, phosphorus oxychloride was distilled from the dark mixture at 35-40° using a water aspirator. The acid chloride was distilled at 85-89° (14 mm.). The yield was 9.1 g. (0.046 mole,

92%), b.p. 199°. The reported b.p, was 197° (67).

The syrup obtained in section a, (3.5 g. ) and pyridine (1.2 g.) (O.OI5 mole), were dissolved in 50 ml. of

(65) N, A. Lange, Handbook of Chemistry, 6th Edition, Handbook Publishers, Sandusky, Ohio, 1946, p. 446,

(67) M. Heibron, Dictionary of Organic Compounds, Oxford University Press, New York, Vol. 2, 1953, P. 186. 28

dry ether, Diethylmalonyldichloride, 1.37 g. (0,007 mole), was added to the ether solution and caused a tan lumpy

precipitate to deposit immediately followed by a slow

precipitation of a white fluffy substance. After 20 hours

reaction time, the mixture was filtered and the residue

washed with 20 ml. of dry ether. The ether filtrate was then

extracted with two 50 ml, portions of 5^ sodium carbonate

solution and two 50 ml, portions of water. After drying the

ether solution with anhydrous sodium sulfate, the solvent

was evaporated. Five ml, of absolute ethanol was added to

the opaque syrup, but crystallization could not be induced.

Enough water (3 ml.) was added to just cause cloudiness.

After 3 days the solution was seeded with the acetylated

barbital diglucoside which was obtained as described pre­

viously. Crystallization began after 2 days. The crystals

were filtered after 10 days and washed with 10 ml. of water.

Some of the syrup was adhering to the crystals. Recrystal­

lization was accomplished from absolute methanol, resulting

in needles, m.p. 174-175°. A mixed melting point with the

acetylated barbital diglucoside obtained previously showed

no depression. The yield was 0.48 g, (0.00057 mole,

10.9^, based on urea). The structure of the glucoside is

shown on the following page. 29

B, Studies with Different Reaction Media

Studies pertaining to the formation of the acetylated di-nitrogen glucoside of barbital were made with the intent to increase the yield beyond that obtain in section A,

Bodendorf*s method of preparation, as described in section A, was repeated five times but the reaction was stopped at various time intervals. The yield, pH, and time of solvent removal are listed below. The initial pH was 9.

Yields (^) pH Time (hours)

10 6 4 14 6 22 15 5 30 16 4 17 4 96 This experiment shows that despite the increasing acidity

as the reaction progresses, the yield increases. The lew pH

after about four hours reaction time indicated that most of

the barbital was present in the acid form and that the 30 potassium hydroxide had reacted with most of the acetobromo- gluccse. Therefore most of the reaction was between barbital and acetobromoglucose after approximately four hours.

A weaker base was then used instead of potassium hydroxide in order to reduce the side reaction between it and acetobromoglucose. When pyridine was used as the proton acceptor instead of the aqueous solution of potassium hydroxide, the yield was 3.9-4.2 percent at the end of four days. This indicated the necessity of a base which was strong enough to form a salt of barbital. When sodium barbital and pyridine was used in place of barbital and potassium hydroxide, a yield of 7.7^ was obtained. The same acetylated di-nitrogen glucoside was obtained in l4^ yield when 0.1 mole of sodium barbital was reacted with 0.1 mole of acetobromoglucose in aqueous-acetone media for four days.

C. Synthesis of 5-Allyl-5-(l-methylbutyl)-, and 5-Ethyl -5-phenyl-l, 3-di- p D-glucopyranosylbarbituric acid

Both of these glucosides wereprepared by the same method. Fifty ml. of acetone was used to dissolve 12.5 g.

(0.03 mole) of acetobromoglucose and 0,015 mole of the barbiturate. Fifteen ml. of 2.ON. potassium hydroxide was added to the acetone solution. After four days the acetone was removed, under reduced pressure, until a syrup started 31 to separate. Attempts to crystallize the syrup from various solvents were unsuccessful. The acetylated glucoside was removed from any unreacted barbiturate by dissolving the syrup in 50 ml. of benzene and extracting with three 50 ml. portions of 20 percent sodium hydroxide and three 50 ml. portions of water. After drying the benzene solution over anhydrous sodium sulfate, the solvent was removed with a water aspirator at room temperature. The remaining traces of benzene were removed in a vacuum desiccator causing the syrup to foam and then harden to a white glassy product.

The yields for phénobarbital and secobarbital were 7.5 and

15.3 percent respectively. Both compounds did not reduce hot Benedicts reagent. This indicated the absence of

2,3,4,6-tetra-O-acetyl-D-glucose which would be fomed by the reaction between potassium hydroxide and acetobromo­ glucose and which reduces hot Benedicts reagent in one minute. A negative test was obtained with alcoholic silver nitrate test solution indicating the absence of aceto­ bromoglucose.

Analytical Data

Phénobarbital Acetylated Glucoside

Calculated for C4QH4QN2O21

N, 3.14: Pound: 2.93

- 8.4° (c4.05, chloroform)

m.p. 90-96° 32

Secobarbital Acetylated Glucoside

Calculated for C40H54N2O21

C, 53.45; Found; 53.30 H, 6.01; Pound: 6.02

N, 3.12; Pound; 3.27

- 29.3° (c 2 .49, chloroform)

m.p. 77-82°

Deacetylatlons were accomplished with a catalytic quantity of sodium methoxide in absolute methanol as des­ cribed! in section A. The yields were practically 100 percent for both compounds. White amorphous products were produced which were very soluble in water. They did not reduce hot

Benedicts reagent,

D. Synthesis of 5-Ethyl-5 -isoamy 1 -1 - ^ -D-glucopyranosyl­ barbituric Acid

The acetylated nitrogen glucoside of araobarbital was prepared in 7 .5$^ yield by the method described in section C.

Deacetylation of this compound with a catalytic amount of sodium methoxide was accomplished in 96 .5# yield. Both were hard, white, amorphous compounds which did not reduce hot

Benedicts reagent. They gave negative tests with alcoholic silver nitrate test solution. Attempts to crystallize them from many solvents were unsuccessful. 33 Analytical Data

Calculated for

C, 54.25; Found: 53.35

H, 6,51; Pound: 6,68

N, 5.06; Found: 5.35 - 18,60 (c, 4.24, chloroform)

m,p. 47-520

The elemental analyses Indicate that a di-nitrogen

glucoside was not formed. As seen previously, barbital,

secobarbital, and phénobarbital form di-nitrogen glucosides.

The reason why amobarbital should form a mono-nitrogen

glucoside is not readily apparent,

E, Synthesis of 5-Ethyl-5-(1 -methyIbuty1)-1, 3 - d i - ^ D -glucopyranosyl-2-thiobarbituric Acid

Sterile sodium thiopental,* 6.0g, (0.02 mole), was

dissolved in 60 ml, of dry acetone. Tetra-O-acetyl-O^-D

-glucopyranosylbromide, 8.2 g, (0,02 mole), was added to the

acetone solution. Sodium bromide started to precipitate

immediately and the light yellow color of the solution

became darker yellow as the reaction progressed. After 26

hours, the mixture was filtered through a sintered glass

filter and washed with 5 ml, of dry acetone. The residue

weighed 2.48 g. which represented 1.98 g. of sodium bromide.

*Sterile sodium thiopental contains 8.3# sodium carbonate. 34

The presence of the bromide ion was confirmed by the yellow precipitate obtained with silver nitrate test solution and by the brown color imparted to carbon tetrachloride when chlorine water was added to an acid solution of the precipi­ tate. The yield of the glucoside based on sodium bromide was 96.0#. Upon removal of the acetone, a yellow syrup resulted which was dissolved in 30 ml, of benzene and extracted with three 10 ml. portions of 2$ sodium hydroxide solution and then with three 10 ml, portions of water. The yellow benzene solution was dried over anhydrous sodium sulfate. After removing the solvent under reduced pressure,

7.79 g. (0.0087 mole) of a yellow amorphous powder was obtained. The yield was 87-0%. Attempts to crystallize the compound from many solvents were unsuccessful. It gave a negative test with alcoholic silver nitrate test solution and did not reduce hot Benedicts reagent. Its melting point was 62-67°. The optical rotation of this compound could not be determined accurately because the transmitted light was not bright enough to distinguish half shadow changes.

Analytical Data

Calculated for CggH^gNgOgQS

C, 52.24 Found; 52.57

H, 5.36 Pound: 5.67

N, 3.12 Found ; 3.46 35 Deacetylation was accomplished with a catalytic quantity of sodium methoxide in absolute methanol as des­ cribed in section A. A yellow amorphous compound was pro­ duced in 98 percent yield. It is very water soluble and reduces hot Benedicts reagent in 25 minutes.

Ill Oxygen Glucosides and Ethers

A. Attempted Synthesis of Substituted Dialuric Acid Glucosides and Ethers

The synthetic method originally planned for the formation of dialuric acid glucosides involves the reaction between the 5-substituted dialuric acid and acetobromo­ glucose in the presence of silver oxide.

Diethylethylmalonate was synthesized in 56 percent yield by a procedure identical to that described in Organic

Syntheses for the preparation of diethyIbutyImalonate (68).

The malonate derivative was reacted with urea in the presence of sodium ethoxide to form 5-ethylbarbituric acid

in 58 percent yield by a procedure similar to that used by

Vogel (69), m.p. 204-20^. Ethyldialuric acid was prepared by oxidation of 5-ethylbarbituric acid with 3 percent

(68) A, H. Blatt, Organic Syntheses, Collective Volume 1, John Wiley and Sons, New York, 1941, p. 250.

(69) A. 1, Vogel, Practical Organic Chemistry, Longmans Green and Co., New York, 1951 > P. 868. 36 hydrogen peroxide according to the procedure of Marberg and

Stranger (70) in 75 percent yield, m.p. 228-230°.

Ethyldialuric acid, 1.71 g. (0.01 mole), silver oxide, 1.16 g. (0.005 mole), and powdered drierite, 20 g., were mixed with 30 ml. of dry benzene. Acetobromoglucose,

4.11 g. (0.01 mole), was dissolved in 15 ml. of dry benzene and added to the black mixture which was stirred with a magnetic stirrer. A few drops of the clear solution was tested periodically with a saturated solution of silver nitrate in ethanol in order to detect the presence of any unreacted acetobromoglucose. After 26 hours, a positive test was still obtained. The brown reaction mixture was filtered through Celite and washed with 50 ml. of dry benzene. The benzene was removed under reduced pressure without heat. The yellow syrup was taken up in 15 ml, of dry ether and cooled to -10°. Crystallization occurred after one week. After filtration and decolorization of the yellow crystals with charcoal in absolute methanol, the yield of white crystals were 1.4 g. {Q0%), m.p. 228-230°, A mixed melting point with 5-ethyldialuric acid showed no depression.

No crystalline products could be obtained from the yellow syrup which resulted upon evaporation of the ether filtrate.

This syrup gave a positive test with alcoholic silver

(70) 0. M. Marberg and D, W. Stranger, J. Am. Chem. See., 6 1 , 2736 (1939). 37 nitrate test solution and. reduced hot benedicts in 3 min­ utes, This procedure was repeated with acetone and acetonitrile with essentially the same results. Different dialuric acids were also used. Recovery of 5-butyl and

5“isoamyldialuric acid was 60-80 percent. No reaction occurred with dioxane as the solvent even when the mixture was refluxed for 14 hours.

The next method that was tried involved the reaction between 5“bromo-5-ethylbarbituric acid and the sodium salt of © -D-glucose-2,3,4,6-tetraacetate and the corresponding

1,2,3,4-tetraacetate. The 5-bromo-5-ethylbarbituric acid was prepared according to the procedure of Dox and Yoder

(71 ) in 88 percent yield. Procedures given in Organic

Syntheses were used to prepare ^-D-glucose-1,2,3,4-tetra­ acetate in 8.7 percent yield (72) and ^ -D-glucose-2,3,4,6

-tetraacetate in 72 percent yield (73).

Glucose-1,2,3,4-tetraacetate, 1,74 g. (0,005 mole) was dissolved in 50 ml, of dry ether. Finely cut sodium,

120 mg.(0,005 mole), was added to the ether solution.

Bubbles immediately started to form around the sodium.

After 12 1/2 hours, the mixture was dark brown and approxi­ mately one-half of the sodium had reacted. After adding

(71) A. W. Dox and L. Yoder, J, Am. Chem. Soc., 44 . 1578 (1922).

(72) E. C. Horning, Organic Syntheses, Collective Volume III, John Wiley and Sons, New York, 1955, P. 432.

(73) Ibid.. p. 434. 38

1,17 g. (0,005 mole) of 5-bromo-5-ethylbarblturlc acid to

the mixture, a black color developed. The black tarry mixture was filtered after 30 minutes and washed with 25 ml,

of dry ether. Ether was removed from the yellow filtrate

until crystallization began. The mixture was filtered

after 30 minutes and the yellow residue was recrystallized

from absolute methanol. It had a m,p, of 202-204° and the

yield was 0,26 g, A mixed melting point with 5-bromo-5

-ethylbarbituric acid showed no depression. When the

remaining ether was removed from the filtrate, a 210 mg, of

a yellow syrup resulted which reduced Benedicts reagent

in 3 minutes. No crystalline products could be obtained

from this syrup.

Sodium would not react with ^ -D-glucose-2,3,4,6

-tetraacetate dissolved in ether or dioxane, A black tar

resulted when a benzene mixture was refluxed,

B, Attempted Synthesis of Enolic Ethers and Glucosides of Substituted Barbituric Acids

The method that was attempted first involved utili­

zation of the directive influence of silver in promoting

enolization.

Dry silver barbital was prepared by adding an aqueous

solution of silver nitrate to an aqueous solution of sodium

barbital with vigorous stirring. The white precipitate was

filtered, washed with absolute ethanol and dried at 110°, 39 When 5 ml. of dimethylformamide (redistilled, 151-152°) was

added to 2,08 g. (0,007 mole) of silver barbital and 2.88 g. (0,007 mole) of acetobromoglucose, a yellow precipitate of

silver bromide was immediately formed and after 5 minutes,

the mixture was brown. Two drops of the clear solution was

tested periodically with alcoholic silver nitrate test

solution in order to detect the presence of any unreacted

acetobromoglucose. After 26 hours, a slight positive test

was still obtained. The brown suspension was filtered

through Celite and washed with 20 ml, of benzene. Charcoal

was ineffective in removing the light brown color from the

filtrate. A brown syrup was recovered by distillation of

benzene and dimethylformamide under reduced pressure.

Twenty ml. of dry ether was added to approximately one-half

of the syrup resulting in precipitation of a brown amorphous

substance which settled out leaving a yellow solution. A

red syrup was obtained by evaporation of the ether. This

syrup was dissolved in 5 ml. of absolute methanol and

brought to the point of cloudiness by adding 3 ml. of water.

After 1 week at room temperature, crystallization had

occurred. The yield of white needles was 50 mg. m.p. 176-

178°. No depression was obtained when a mixed melting point

was made with the acetylated di-nitrogen glucoside of

barbital. 40

The other half of the syrup was dissolved in 10 ml. of dry ether and extracted with 10 ml. of 5 percent sodium hydroxide solution and 10 ml. of distilled water. After drying the ether solution over anhydrous sodium sulfate and distilling, fine needles were obtained. They were recrystal- lized from absolute methanol; yield 6o mg. m.p. 174-175°.

Other experiments with silver barbital or silver oxide plus barbital with acetone, benzene or DMF were un­ successful. The rate of reaction decreases markedly as the polarity of the solvent decreases.

The next proposed method of synthesis was to react chlorinated barbiturates with glucose tetraacetate.

Chlorination of barbital was attempted with phosphorus oxychloride and phosphorus pentachloride. Baddiley and

Topham (74) prepared, 2,4,6-trichloropyrimidine by reacting phosphorus oxychloride with barbituric acid in dimethyl- aniline. Their procedure of chlorination was repeated with barbital. Ninety percent of the barbital was recovered.

Phosphorus pentachloride, a stronger chlorinating agent, was tried next. Phosphorus pentachloride, 41,7 g. (0,2 mole), was added to a mixture of 18,4 g (0.1 mole) of barbital and 40 ml, of phosphorus oxychloride. It was necessary to reflux the mixture in order to cause evolution

(74) J. Baddiley and A. Topham, J. Chem. Soc., 678 (1944). 41 of hydrogen chloride. After 70 minutes, no more hydrogen chloride was evolved. Phosphorus oxychloride was distilled at 4o° under reduced pressure. Ten ml, of dry ether was added to the black syrup. The mixture was filtered and the tan crystals washed with 5 ml, of ether. After recrystal- lization from absolute methanol, their melting point was

192-194°, The yield was 5,3 g. of barbital, A light yellow liquid was obtained by distilling the black ether filtrate.

The distilling range was 80-90° (15 mm.). A negative test for chlorine was obtained by subjecting the liquid to sodium fusion and testing with silver nitrate test solution.

No pure products could be obtained from the tarry residue,

C, Synthesis of 5,5-Diethyl-4-(tetra-0-acetyl-%&-D-gTuco- pyrano syloxy)-2, 6-pyrimidinedione

This compound was synthesized by the procedure of

Snyder and Link (75). A mixture of 4l.l g. (0,10 mole) of tetra-0-acetyl-h(-D-glucopyranosylbromide, 18,5 g . (0,10 mole) of barbital, 4l g, of drierite, 17,4 g, (0,30 mole) of silver oxide, 0,9 ml, of quinoline and 400 ml, of dry ether was stirred with a magnetic stirrer. The flask was

covered with aluminum foil. At the end of 21 hours, a negative test for bromide ion was obtained with alcoholic

silver nitrate. The suspension was filtered through Celite

(75 ) Snyder and Link, op, cit.. p. 1882, 42 and washed with three 100 ml. portions of chloroform. The solvents were removed under reduced pressure and the re­ sulting yellow syrup was dissolved in 100 ml. of dry ether.

After keeping the ether solution at 5° for 20 hours, a gummy crystalline mass separated which was filtered and washed with two 50 ml. portions of dry ether. The yield was 7,8 g. and the m.p, was 145-152°, This was dissolved in 150 ml, of boiling methanol, decolorized with activated charcoal and 100 ml, of water was added. Fine white needles separated on cooling to 5°. After filtering and washing with 100 ml, of water, the yield was 3.8 g., m,p. 158-163°.

After recrystallizing from 1:1 methanol-water, the yield was 2.75 S. (0.0053 mole), m.p. 178-179°.

Analytical Data

Calculated for OggH^QOj^gNg

C, 51.35; Found: 51.20

H, 5.88; Found: 5.75

N, 5.45; Found: 5.66

[ cx 8.04° (C, 4 .17, chloroform)

Deacetylation was attempted by the procedure described

in section II-A. The result was cleavage of the glucoside bond and deacetylation which was the same as that obtained

by Snyder and Link with barium methoxide. 43 IV Pharmacological Testing

A comparison was made between the barbiturate gluco­ sides and the corresponding sodium barbiturate salts for hypnotic activity in albino rats. The lack of hypnotic activity of the barbiturate glucosides was demonstrated with intraperitoneal injections of 6 to 25 times the hypnotic dose of the corresponding barbiturate salt. Both the enolic tetraacetate and nitrogen type glucosides were inactive. The results are summarized in Table I.

TABLE I

PHARMACOLOGICAL RESULTS

Anesthetic Dose Dose of of Sodium Type of Barbiturate Barbiturate Barbiturate* Glucoside Glucoside* Results

Barbital 200 Di-Nitrogen 1,500 Inactive

Barbital 200 Mono-Enolic Tetraacetate* 1,200 Inactive

Phénobarbital 100 Di-Nitrogen 1,000 Inactive

Secobarbital 40 Di-Nitrogen 1,000 Inactive Painful

Amobarbital 200 Mono-Nitro­ gen 2,000 Inactive

Thiopental 40 Di-Nitrogen 500 Inactive

All were intraperitoneal injections of aqueous solu­ tions except the mono-enolic glucoside tetraacetate of bar­ bital which was in the form of a 1^ acacia aqueous suspension. The doses are in rag. of compound per kg. of rat. 44

V Analysis of Infrared Spectra

The infrared spectra were prepared in the Spectro- graphic Laboratory of the Chemistry Department with a Baird

Associates Infrared Recording Spectrophotometer, Model B, equipped with a NaCl prism. A Nujol mull was used for each compound. The absorption bands for Nujol are marked on the figures with an asterisk (3.41, 6,84, and 7.2% y, ).

The enolic glucoside tetraacetate of barbital which is shown in Figure 1 has a ring -C = N- absorption band at

6.1 ^ , Ring -C = N- bands are from 5.97 to 6.29 W (76).

This band is absent in all the other spectra and therefore is diagnostic for an enolic type barbiturate glucoside. The

-NH band at 3.1 V» in Figures 1 and 7 indicates that the mono

-glucoside was formed. Because this band is absent in

Figures 3 and 6, a di-nitrogen glucoside of barbital and thiopental must have been formed. The assigned values for

-OH and -NH bands are 2.66 - 2.98 ^ and 2.88 - 3.28 ys , respectively (77). The -OH band at 3.0 ^ is very prominent in the spectrum of the di-nitrogen glucoside of barbital

(Figure 4). It is not possible to distinguish between the

-NH and -OH bands. The acetate bands are seen at 8,05

- 815 y\ in Figures 1, 3, and 6 (assigned value 8.03 ^ ).

(76) H, M. Randall, et ai.. Infrared Determination of Organic Structures, D, Van No strand Co., New York, 1949, pp. 26-31.

(77) Ibid.. p. 6. 45

The C-4 and C-6 carbonyls of barbituric acid absorb at

5 .7 1 ^ and the C-2 carbonyl absorbs at 5.89 ^ (78).

']?hese bands are seen at 5.60 - 5.67 |u| and 5.75 - 5.80 ^ respectively in Figures 1-3.

(78) Ibid., pp. 21 and 25 . WAVE NUMBERS IN CM-' WAVE NUMBERS IN CM-' 5000 4000 3000 2500 2000 1500 1400 1300 1200 1100 1000 BOO900 1001- i 'Li - f -

- -i

....

WAVE LENGTH IN MICRONS WAVE LENGTH IN MICRONS

Pig, 1.“ Infrared Spectrum of 5,5-Dlethyl-4-(tetra-0-acetyl- R -D-gluco- pyranosyloxy)-2,6-pyrimidinedione WAVE NUMBERS IN CM ' WAVE NUMBERS IN CM-' 5000 4000 3000 2500 2000 1500 1400 1300 1000 900 800 100

60

5 «

WAVE LENGTH IN MICRONS WAVE LENGTH IN MICRONS

Pig. 2 .- infrared Spectrum of5 , 5 -D iethylbarbituric Acid

-4 WAVE NUMBERS IN CM-' WAVE NUMBERS IN CM " 5000 3000 2500 1500 1400 1300 1200 1100 1000 900 800 IOC

= 60

40

WAVE LENGTH IN MICRONS WAVE LENGTH IN MICRONS

Fig. 3«~ Infrared Spectrum of 5,5-Diethyl-l,3-dl-(tetra-0-acetyl-(3 -B -glucopyranosyl)barbituric Acid

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60

5 «

W A VE LENG TH IN MICR ON SWAVE LENGTH IN MICRONS WAVE LENGTH IN MICRONSWAVE

Pig. 4,“ Infrared Spectrum of 5,5-Dlethyl-l,3-dl-j3?-D glUGopyranosylbarbituric Acid

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60

5 «

W A V E LENG TH IN MICR O NSWAVE LENGTH IN MICRONS WAVE LENGTH IN MICRONSWAVE

Pig. 5.“* Infrared Spectrum of 5“Pthyl“5“(l~niethylbutyl)-2“thio- barbituric Acid

ui o WAVE NUMBERS IN CM-' WAVE NUMBERS IN CM-' 5000 4000 3000 2500 2000 1500 1400 1300 1200 1100 1000 900 800 100

C 60

fc 40

WAVE LENGTH IN MICRONS WAVE LENGTH IN MICRONS

Pig. 6.- Infrared Spectrum of 5-Ethyl-5-(l-methylbutyl)-l,3-dl “tetra-0“acetyl“ ^ “-D“Slucopyranosyl)“2“thiobarblturic Acid

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WAVE LENGTH IN MICRONS WAVE LENGTH IN MICRONS

Fig, 7.“ Infrared Spectrum of 5“Ethyl-5“Isoamyl-l“(tetra“0-acetyl “D“glucopyranosyl)barbituric Acid

\ji ro DISCUSSION

Barbital, secobarbital and phénobarbital produced the dl-nitrogen glucoside by reacting with acetobromoglucose in aqueous-acetone media with two equivalents of potassium hydroxide. Even when one equivalent of base was present, the di-nitrogen glucoside was produced. As the reaction progressed, the media became acid but the yield continued to increase. This indicated that the anion of barbital was not essential for the reaction but lower yields were ob­ tained when pyridine was substituted for potassium hydroxide.

High yields of the di-nitrogen glucoside were obtained when sodium pentothal was reacted with acetobromoglucose in dry acetone solution. The reasons for the increased yield with dry acetone as the solvent instead of an alkaline aqueous-acetone media are; (1) the absence of the side reaction between potassium hydroxide and acetobromoglucose and (2 ) the absence of barbiturate anion hydrolysis,

Amobarbital produced the mono-nitrogen glucoside with

acetobromoglucose in alkaline aqueous-acetone media. The reason for the anomalous behavior of amobarbital is not

readily apparent. The isoamyl group offers some steric hindrance to the approach of the acetobromoglucose molecule.

However, this hindrance is only slightly greater than that

53 54 of the sec~butyl group present in secobarbital and thio­ pental as illustrated by the use of models. Rapaport (79) found that amobarbital did not react like other barbitur­ ates with regard to di-salt formation. The- disilver salt could not be prepared.

In the attempted preparation of glucose derivatives of dialuric acid, the sodium salt of glucose-1,2,3^4-tetra- acetate and the corresponding 2,3,4,6-tetraacetate could not be prepared. It is difficult to understand why only approx­ imately one-half of the sodium reacted because primary alcohols usually react with sodium very rapidly and the hemi-acetal group is more acidic than an alcohol group. The tars which were produced could have been the result of polymerization due to the displacement of acetate by the alkoxide and base catalyzed aldehyde reactions.

In other experiments, attempts were made to synthesize dialuric acid glucosides with acetobromoglucose and silver

oxide. These results were inconclusive due to inability to purify any products. Probably, a mixture of the di-nitrogen

and alcoholic glucosides were produced. It was shown that

some of the acetobromoglucose did not react. The formation

of the alcohol type glucoside would be hlndred due to the

(79) L, I. Rapaport, Ulcrain,Ki-.iim. Zhur., 20_, 430 (1954); through Chera, Abs., 669 (1957). 55 tv/o adjacent carbonyls decreasing the electron density on the alcohol oxygen atom and due to steric hindrance.

The influence of solvent polarity on the rate of reaction was demonstrated with the attempted preparation of enolic glucosides of barbital. As the polarity of the solvent increased, the rate of reaction increased as demon­ strated by the formation of silver bromide from silver oxide. This is in accordance with the unimolecular mechan­ ism of nucleophilic substitution with acylglycosylhalides as postulated by Newth, Phillips and Mattok (80,81).

The chlorination of barbital with phosphorus oxy- chloride and phosphorus pentachloride was unsuccessful.

The procedure by which 2,4,6-trichloropyrimidine was pre­ pared from barbituric acid and phosphorus oxychloride was repeated with barbital. A powerful driving force in the trichloropyrimidine synthesis is the large resonance energy present in the product as compared to the lack of resonance in the reactant. The chloropyrimidone that would be formed from barbital does not have this large resonance energy and hence this reaction would not have this driving force. It is possible that the ring was cleaved. Dox (82) formed the

(80) P. H. Newth and G. 0, Phillips, J. Chem. Soc., 2896 (1953).

(81) G. L. Mattok and G. 0. Phillips, ibid.. 1836 (1956). (82) A. W. Dox, J. Am. Chem. Soc., I559 (1931) 56 tetrachloro derivative of barbital by treating it with a large excess of phosphorus pentachloride at 120° for 6 hours and then at 135° for 4 hours after distilling the phosphorus oxychloride. The yield was low due to ring cleavage forming diethylcyanoacetylchloride.

The lack of hypnotic activity of the glucosides indicated that they were not hydrolyzed. The enolic type glucoside was much easier to. hydrolyze than the nitrogen type glucoside. Therefore the presence of the glucose moiety on barbiturates completely blocked the inhibitory action of barbiturates on glucose oxidation. SUMMARY

1, The following nitrogen glucosides were synthesized by

reacting the 5 ,5-disubstituted barbituric acid with

acetobromoglucose In an alkaline aqueous-acetone media

followed by deacetylation with a catalytic quantity of

sodium methoxide in absolute methanol.

(1) 5,5-Diethyl-l,3~cli-y5’ -D-glucopyranosylbarbituric

acid

(2 ) 5-Allyl“5-(1 -methyIbutyl)-l,3-di- -D-gluco-

pyranosylbarbituric acid (3) 5-Ethyl-5-phenyl-l,3-di-J3 -D-glucopyranosyl- barbituric acid

(4) 5-Ethyl“5-isoamyl-l-^ -D-glucopyranosyl-

barbituric acid

2, The synthesis of 5-ethyl-5-( 1 -methyIbutyl) -1 ^ 3-di-^

-D-glueopyranosy 1 -2-thiobarb 11uric acid was accomplished

by reacting acetobromoglucose and sodium thiopental in

dry acetone media followed by deacetylation with a

catalytic quantity of sodium methoxide in methanol,

3 , All of the glucosides were very water soluble,

4, The glucoside, 5,5-diethyl-4-(tetra-0-acetyl-^ -D-

g lue opy ranosyloxy ) -2 ,6-pyrimidinedione was synthesized

according to the method of Snyder and Link, This was

done bjr using barbital, acetobromoglucose and a 57 58

catalytic quantity of quinoline in an ether suspension

of silver oxide. The yield was 5.3^ as compared to the

reported yield of 2.6^. Attempts to deacetylate the

compound resulted in cleavage of the glucoside bond.

5. Studies pertaining to the synthesis of 5,5~diethyl-l,

3-di-^ -D-glucopyranosylbarbituric acid were made in

different reaction media.

6. The results of the following attempted syntheses of

substituted dialuric acid glucosides and ethers were

inconclusive.

{a) The ^-substituted dialuric acid was reacted with

acetobromoglucose in the presence of silver oxide,

(b) The sodium salt of-D-glucose-l,2,3,4-tetra-

acetate and the corresponding 2,3,4,6-tetraacetate

was reacted with 5-bromo-5 ethylbarbituric acid.

7. Chlorination of 5,5-diethylbarbituric acid with phos­

phorus oxychloride and phosphorus pentachloride was

unsuccessful.

8. The reaction between the silver salt of 5,5-diethy1-

barbituric acid and acetobromoglucose was unsuccessful.

9. Both the nitiogen and enolic type glucosides were

demonstrated to have no hypnotic effect when tested in

rats by intraperitoneal injection of massive doses. 59

10. Analysis of infrared spectrographs showed that the

enolic type glucoside could be differentiated from

the di-nitrogen type glucoside by the presence of a

ring -C = N- absorption band in former type. Also

mono-nitrogen acetylated glucosides could be distin­

guished from the di-nitrogen type by the presence of

the -NH absorption band in the former type. AUTOBIOGRAPHY

I, Glenn Arthur Portmann, was born in Canton, Ohio,

August 13, 1931. My secondary school education was received in Canton, Ohio. The Ohio State University granted me a Bachelor of Science degree in Pharmacy in

1954 and a Master of Science degree in Pharmacy in 1955.

I held an American Foundation for Pharmaceutical Education

Fellowship from 1954 to 1957.

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