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Oct., 1947
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The purified product was obtained crystalline Since the yield of 6-tetrahydroxypteriiie by the above procedure was small, our attention was as clusters of yellow needles, and in 0.1 N sodium centered on the more direct synthesis of I11 by the hydroxide solution it shows ultraviolet absorption condensation of I with osones (11). The reaction maxima a t 260, 284 and 370 mp, and minima a t of I with I1 is rapid and yields I11 in good quan- 239, 271 and 333 mp. Anal. Calcd. for CI9H2,tity. The conditions for obtaining the preferred 0&8.2H20: C, 47.9; H, 5.1; N, 23.5. Found: isomer appear to be reversed from that described C, 47.:3; H, 5.15; N, 23.4. Magnesium salt: * ~44.2; H ~ O :H, above-;. e., I and I1 a t p H 5-9 yield the 6-isomer, Calcd. for C I ~ H ~ E O ~ N E M ~C, while the condensation of I-bisulfite and I1 in 4.T; N, 21.7; Mg, 4.7. Found: C, 44.6; H, strongly acidic solution yields a mixture richer in 4.%; N , 21.4; Mg, 4.82. The biological properties have been examined by Dr. B. L. Hutchings the 7-isomer. Although details of work on pure isomers will and Dr. E. L. K. Stokstad of the Lederle Laborabe published later it is deemed worthy to report tories Division, American Cyanamid Company, the synthesis oF the isomeric mixture of I11 and Pearl River, New York. The inhibition ratio for the preparation of the isomeric mixture of formyl- half-maximum inhibition of the growth of Strepfosoc~irsfuecalis R is 1.9, 0.7 and 0.4 a t concenpterine (IV) from I11 by the method outlined. D-Glucosone was heated with an eqnivaleiit trations of pteroylglutamic acid of 0.003, 0.005 amount of “4,3-triainiiio-Ci-hydroxypyrimiditie ~ i i d0.01 microgram per 10 ml., respectively. Details of the synthesis and properties of this bisulfite in 7tjf( acetic acid a t 73” for forty-five minutes. The mixture was cooled and the pre- and related compounds will be the subject of subcipitate collected. The product was exhaustively sequent communications. extracted with hot alcohol aiid dried. Yield of Car co CHEMICAL DIVISION DORISR. SEEGER ~) (169.2 mg. per 100 ~ I E R I C A YC Y A Y a M I D c O M P A \ Y JAMES M. S M I T H , JR. 111 was 6Oz,,[ c x ] ~ ‘ ~-70.9’ JERSEY XARTIS E. HULTQUIST ml. of -IrNaOH). Absorption spectrum in 0.1 -V 1 3 0 ~VI-, BROOK,SEW RECEIVEDSEPTEMBER 19, 1947 KaOH showed maxima a t 232 mp and 360-362 nip with e of 19,000 and 7940, respectively. Anal. Calcd. for C10H13N606: C, 42.39; H, BIOSYNTHESES INVOLVING PANTOTHENIC AClD 1.62; N, 24.71. Fourid: C, 42.1:; H, 4.(12; N Sir : (Kjeldahl), 25.11. In Esdherichitc soli cysteic acid appears to preI11 was oxidized with lead tetraacetate to IV, vent competitively the decarboxylation of aspartic a n isomeric mixture, obtained in S3yGyield. IV acid to @-alaninewhich results in pantothenic acid contained ash which was hard to remove. It ex- becoming a limiting growth factor.’ Under our hibited strong carbonyl activity forming oximes, testing conditions the rate of pantothenic acid hydrazones and Schiff bases readily. IV treated synthesis is determined by the ratio of cysteic to with a slight excess of barium permanganate gives aspartic acid, and exogenous substances allowing Y,identity of which was established by its ultra- growth to occur a t a lower rate of pantothenic violet absorption, titration curve and analysis. acid synthesis produce an increased antibacterial Anal. Calcd. for C7H6N6o2H20: C , 40.2; index. K , 33.5. Found: C, 38.8; N (Kjeldahl), 31.5 Such an effect is obtained with citric, cis(cor. for 4.90% ash). aconitic or a-ketoglutaric acids. The antibac . H. G. PETERINGterial index over a thirty-fold range in aspartic THEUPJOHXCOMPANY KALAMAZOO, MICHIGAN D. I. WEISBLAT acid concentrations was 300 in the medium conRECEIVED AUGUST18, 1947 taining these substances but only 30 in their absence. Oxalacetic and pyruvic acid were inactive alone, but a mixture of both necessitated a slight ANTAGONIST FOR PTEROYLGLUTAMIC ACID increase in the concentration of cysteic acid to obSir: tain the same growth inhibition. Acetate alone \Ve wish to report the synthesis of a potent possessed some activity. Pantoic acid was inacpteroylglutamic acid antagonist, N- [4-{ [ (2,4-d1- tive. The apparent “sparing action” of cisamino - 6 - pteridyl) - methyl] - amino] - benzoyll- aconitic acid on the pantothenic acid requireglutamic acid. In the course of an investigation ment of E . coli is not equaled by its precursors; of analogs of ptieroylglutamic acid, this compound hence, it appears that pantothenic acid deficient was prepared from 2,4,5,6-tetraminopyrimidine cells are unable to convert effectively pyruvate sulfate,’ 2,3-d1bromopropionaldehyde, and p - and oxalacetate to cis-aconitate (or ketoglutarate). aminobenzoylglutamic acid under the conditions This datum explains the previously reported’ described for the synthesis of pteroylglutamic enhanced activity of glutamic over aspartic acid acid.2 Purification of the crude product was ac- in preventing the toxicity of cysteic acid. The complished by a method very similar to that used transamination reaction produces both aspartic for pteroylglutamic acid.3 and a-ketoglutaric acids, the latter having a ( 1 ) Traube Ber ( 2 ) Angler rl ni (31 Vvaller el al
37, 4545 (1904) Sctence 103, 667 (1946) ’IHIS J O U R N A L 69, In Dress (1947)
(1) Ravel a n d Shive, J Bid Chem., 166, 407 (1946). (2) Molar ratio (analog t o metabolite) just necessary for maximum inhibitiuu of growth.
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“sparing action” on the product of the blocked reaction. Previous evidence3 involving pantothenate in the oxidation of pyruvate can also be explained on the basis of pantothenate mediating cis-aconitate synthesis. That the above effect directly involves pantothenic acid was demonstrated by the reversing effect of both pantothenic and a-ketoglutaric acids on a pantothenic acid antagonist, N-a,ydihydroxy - p,p - dimethylvaleryl - p - aminobutyric acid, for E. coli. Further, the pantothenic acid ,requirement of Proteus morganii in a medium of inorganic salts, glucose, nicotinamide and cystine was appreciably decreased by aketoglutaric acid. With Lactobacillus arabinosus, an oleic acid source ( ‘Tween SO”) or sodium glycocholate increased the antibacterial index from approximately 3,000 to 30,000 for the competitive inhibition of N-pantoyl-n-butylamine4 of pantothenic acid functioning. Both substances added simultaneously did not enhance the effect. Since this organism presumably requires acetate for synthesis of sterols and fatty acids, this inhibitor appears to prevent the conversion of acetate to an intermediate common to both sterol and oleic acid synthesis. The reported involvement of pantothenic acid in the conversion of glycine t o t h r e ~ n i n e ,the ~ demonstration by Lipmann, et u L . , ~ of the presence of pantothenic acid in the coenzyme for acetylation of sulfanilamide and choline, and the results of the above inhibition analyses tend to indicate that many of the enzymatic reactions in which pantothenic acid functions involve the hypothetical ‘‘act.ive”acetyl radical.
Vol. 69
enantiomorphs2 of methyl hydrogen @methylglutarate (I) CHs
I
CHjOOC-CH-C-CH~-COOH
I H
(1)
Rearrangement leads in this case to racemization and can be detected simply by pouring the ester acid chloride into water and measuring the rotation of the recovered half ester. In this way i t has been found that no rearrangement takes place when the ester acid chloride is prepared by the action of oxalyl chloride in benzene solution. In case of thionyl chloride the occurrence and extent of rearrangement depend on the purity of the reagent and on the reaction temperature. If pure thionyl chloride (Kahlbaum “reinst, wasserhell”) is used no rearrangement occurs if the reaction takes place a t 30’ and excess reagent is removed under reduced pressure on a water-bath kept at 50’. Use of less pure thionyl chloride (Kahlbaum, “purum”) leads under the conditions just described to rearrangement, the extent of which increases if the reaction temperature is raised. (The ester acid chlorides have not been distilled.) It is well known that anhydrides may be formed during the action of thionyl chlorides on acids and that the yield of acid chloride is lower if impure thionyl chloride is used or if the reaction temperature is too high.3 With dibasic acids of the succinic and glutaric acid series thionyl chloride gives the cyclic anhydrides only. It appears likely that the rearrangement observed in the preparation of the ester acid chlorides occurs via the anhydrides. That rearrangement may be avoided during the (3) Dorkman, el at., J . B i d . Chem., 144, 393 (1942). (4) Snell a n d Shive, ibid., 160, 287 (1945). preparation of the ester acid chloride of (I) is evi(5) Kossi a n d Cennamo, Chem. Abstr., 40, 6543 (1946). dent from the fact that lengthening of the chain ( 6 ) Lipmann, (et ai.,J . Elid.Chem., 167, 869 (1947). of the dextrorotatory enantiomorph of (I) by the LVILLIAM SHIVE BIOCHEMICAL IXSTITUTE AND THE , .4rndt-Eistert synthesis has given (+)-P-methylCHEMISTRY DEPARTMENT OF THE W. W. ACKERMANNadipic acid identical with that derived from UNIVERSITY OF TEXAS,AND THE JOANNE MACOW RAVEL natural products.2 Furthermore, the two enanFOUNDATION FOR RESEARCH CLAYTON tiomorphs of (I) have been used as starting maJUDITH ELIOTT SUTHERLAND AUSTIX,TEXAS terial for the synthesis of d ( + ) - and Z(-)-3RECEIVED SEPTEMBER 19,1947 methyltetracosanoic acids. The intermediate ester acid chloride was i n cnsc o f one enantioinorph prepared by means of oxalyl chloride :tnd in case REARRANGEMENT IN THE PREPARATION OF of the other by means of pure thionyl chloride. ESTER ACID CHLORIDES The enantiomorphic long chain p-methyl subSir: stituted acids both melted sharply a t 65.5’ Cason’ has recently drawn attention to the fact (cor.), and showed numerically equal optical that during the preparation of the ester acid chlo- rotations, [hIIz5D 13.2’ (chloroform, c, 5.78). rides of the isomeric half esters of dibasic acids Mixed in equal proportions the acids gave a racesuch a s a-ethyl-a-butylglutaric acid by means of mic compound melting sharply a t 68.6’ (cor.). thionyl chloride rearrangement may occur, the INSTITUTE OF MEDICAL CHEMISTRY derivatives obtained from the ester acid chlorides UNIVERSITY OF UPPSALA STINA STALLBERG-STENHAGEN being mixtures derived from both isomers. UPPSALA,SWEDEN The writer has studied’the conditions under RECEIVEDSEPTEMBER 11,1947 which this type of rearrangement occurs when S. StLllberfi--Stenhagen, Arkiw Kemi, &fin., Geol.. M A , No. preparing the ester acid chlorides of the two 10(2) (1947). (1)
1. Cason, rTHIS JOURNAL, 69, 1548 (1947).
(3) “Organic Syntheses,’’ Coli. Val. I, 2nd ed., p. 147.
