[Advances in Carbohydrate Chemistry] Volume 6 || Aconitic Acid, A by-Product in the Manufacture of...

ACONITIC ACID, A BY-PRODUCT IN THE MANUFACTURE OF SUGAR

BY ROBERT ELLSWORTH MILLER AND SIDNEY M. CANTOR

Research and Development Division, American Sugar Refining Company, Philadelphia, Pennsylvania

CONTENTS I. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 231

11. Physical Properties of Aconitic Acid.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 234 236

IV. The Recovery of Aconitic Acid in the Manufacture of Sugar.. . . . . . . . . . . . . 239 V. Chemistry and Uses of Aconitic Acid., . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 244

111. Analytical Estimation of Aconitic Acid.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

I. INTRODUCTION

For many years, aconitic acid (I), an unsaturated tribasic aliphatic

H2C-COOH

A-COOH II

HC-COOH

I

acid, l12,3-propenetricarboxylic acid, has been recognized as a constituent of extracts from sugar-containing plants. Only in the past ten years, however, has it become apparent that the recovery of aconitic acid as a by-product in the manufacture of sugar was feasible. Recently this recovery has assumed greater importance with the realization that aconitic acid, its salts, and its esters are important chemical intermediates in the preparation of plasticizers for various types of resins and also in the preparation of surface-active agents.

Aconitic acid was first isolated in 1820 by Peschier' and during the early years after its discovery a variety of names, e.g., aconitic acid,' equisetic acid, 2,a achillea acid14 " Brenzcitronsaure1116 and " Citridin-

(1) Anon., Ann., 28, 243-46 (1838). (2) H. Braconnot, Ann. chim. phys., 58, 5-24 (1828). (3) V. Regnault, Ann., 19, 145-54 (1836); Ann. chim. phys., 62, 208-17 (1836). (4) B. Zanon, Ann., 68, 21-36 (1846). (5) G. L. Crasso, Ann., 54, 53-84 (1840).

23 1

232 ROBERT ELLSWORTH MILLER AND SIDNEY M. CANTOR

were assigned to the acid. It had been named originally as aconitic acid by Peschier due to his isolation of the acid from Aconitum napellus and paniculatum and this name became the adopted common name for the acid.

Very little quantitative data have been reported in relation to the concentration of aconitic acid in various plant juices other than from cane sorghum and from sugar cane varieties although the acid has been identified in the juices from many other species of plants. It has been isolated from a variety of species of A ~ o n i t u m , ' , ~ - ~ ~ D e l p h i n i ~ m , ~ ~ ~ ~ and E q ~ i s e t u r n , ~ ~ ~ ~ ~ ~ ~ and also from Achillea rnillef~lium,~~'~ Adonis vernalis, 1**16

Helinus ovatus, l6 Sansevieria zeylancia, l7 wheat , l a and from barley, maize, oat and rye plants.1° It can indeed be said that aconitic acid, at least qualitatively, is a commonly occurring constituent in the plant kingdom.

BehrP2O in 1877, identified aconitic acid as a constituent of sugar cane molasses while its occurrence in sugar beet products was demonstrated in 1879 by von Lippmann.21 Numerous other investigators since then have isolated aconitic acid from sugar cane sugar cane molas-

(6) S. Baup, Ann. chim. phys., [3] 30, 312-24 (1850); Ann., 77, 293-305 (1851). (7) Anon., Wyoming Agr. Expt. Sta. Repts., 131 (1920); Ezpt. Sta. Record, 46, 410

(8) D. Wasowicz, Arch. Pharm., 214, 193 (1879). (9) 0. A. Beath, J . Am. Chem. SOC., 48,2155-58 (1926). (10) A. Jermstad, Medd. Norsk. farm. Selsk., 6, 81-86 (1944) (Chem. Zentr.,

(11) H. A. D. Jowett, J . Chem. SOC., 69, 1518-26 (1896). (12) W. Wiche, Ann., 90, 98-99 (1854). (124 W. Wohlbier and S. Beckmann, Ber., 83, 310-14 (1950). (13) H. Hlasiwetz, Jahresber. Fortschritte der Chem., 331 (1857); J . prakt. Chem.,

(14) F. Linderos, Ann., 182, 365-66 (1876); F. W. Heyl, M. C. Hart and J. M. Schmidt, J . Am. Chem. Soc., 40, 436-53 (1918) report that they were unable to find any aconitic acid in Adonis vernalis.

(16) N. Orlow, Pharm. 2. f. Russland, 33, 771 (Chem. Zentr., 1895, I, 202). (16) J. A. Goodson, J . Chem. SOC., 117, 140-44 (1920). (17) S. Scheindlin and A, A. Dodge, Am. J . Pharm., 119, 232-53 (1947). (18) E. K. Nelson and H. Hasselbring, J. Am. Chem. SOC., 63, 104043 (1931). (19) E. K. Nelson and H. H. Mottern, J . Am. Chem. SOC., 63, 3046-48 (1931). (20) A. Behr., Ber., 10, 361-65 (1877). (21) E. 0. von Lippmann, Ber., 12, 1649-61 (1879). (22) P. A. Yoder, J . Znd. Eng. Chem., 3, 640-46 (1911). (23) C. 8. Taylor, J . Chem. SOC., 116, 886-89 (1919). (24) F. W. Zerban, J . Ind. Eng. Chem., 11, 103436 (1919).

(Chem. Abstracts, 16, 3500 (1922)).

1944, 11, 1198).

72, 429-31 (1867).

ACONITIC ACID I N T H E MANUFACTURE OF SUGAR 233

ses,25--28 and from sorghum product^^^-^^^ and it has been reported that the average aconitic acid content of sugar cane is about 25 to 40 percent of the amount in sorgho.82 Several workers have observed the deposition of alkaline earth aconitates as a sediment in molasses, as scale in sugarhouse evaporators and vacuum pans, and in some cases as a precipitate in concentrated sugar liquors during the crystallization procedure^.^^,^^-^^ It was not until the comparatively recent work of McCalip and Seibert,34 and of Balch, Broeg and Ambler,37 however, that quantitative data on the amount of aconitic acid in various sugar cane products were available. These workers found that the amount of aconitic acid in the sugar cane juice was only a few tenths of a percent of the total weight of the juice. The acid concentration, however, became higher as the sugar liquors were processed and various molasses samples were found to contain from 1.8 to 6 percent aconitic acid based on the Brix solids of the molasses.

The actual function of aconitic acid in the plant physiology is not clearly understood. Cis-aconitic acid undoubtedly is present t o a certain extent in the plant because of its role in the citric acid cycle but the factors which cause the accumulation of comparatively large quantities of aconitic acid have not been clearly defined. Whether the concentration of aconitic acid which is formed in the plant is the cis-form exclu- sively or is a mixture of the cis- and the trans- acids is also unknown. Balch, Broeg and Ambler,37 in their investigations, reported considerable variation in the aconitic acid content of various crusher juices of Louisiana

(25) E. K. Nelson, J . Am. Chem. SOC., 61, 280&10 (1929). (26) E. K. Nelson and C. A. Greenleaf, Ind. Eng. Chem., 21, 857-59 (1929). (27) H. C. Prinsen-Geerligs, Arch. Suikerind., 41, 720-21 (1933). (28) K. Miti, Bull. Inst. Phys. Chem. Research (Japan), 22, 671-73 (1943). (29) H. B. Parsons, Amer. Chem. J . , 4, 39-42 (1882-3). (30) H. W. Wiley and W. Maxwell, Amer. Chem. J., 12, 216 (1890). (31) J. J. Willaman, R. M. West and G. E. Holm, J . Agr. Research, 18, 1-33

(31a) R. H. Cotton, L. W. Norman, G. Rorabaugh and H. F. Haney, Znd. Eng.

(32) Anon., Louisiana State Univ. Eng. Ezpt. Station News, 1, No. 2, 9 (1945). (33) E. K. Ventre, Sug. Jour., 8, No. 7, 23-30 (1940). (34) M. A. McCalip and A. H. Seibert, Ind. Eng. Chem., 38, 637-40 (1941). (35) W. L. McCleery, Repts. of 62nd meeting of Hawaiian Sugar Planters Assoc.,

(36) H. A. Cook, Hawaiian Planter's Record, 47, No. 2, 71-73 (1943). (37) R. T. Balch, C. B. Broeg and J. A. Ambler, Sugar, 40, No. 10, 32-35 (1945);

(38) Anon., Intern. Sugar J. , 47, 112 (1945). (39) J. McGlashan, Proc. 16th Conv. Sugar Tech. Assoc. India, 1,83-93; as reported

(1919) [Chem. Abstracts, 14, 961 (1920)l.

Chem., 48, 62l3-35 (1951).

pp. 83-84; as reported in Intern. Sugar J . , 46, 244 (1943).

41, No. 1, 46 (1946).

in Intern. Sugar J . , 61, 31-32 (1949).


sugar canes. This fact led these authors to suggest that an important factor which may affect the aconitic acid concentration in the plant is the composition of the soil and that the acid may play an important part in stabilizing the reactions of the plant ~ a p . 4 ~ In their opinion the function of the aconitic acid may be to neutralize the basic elements which are absorbed from the soil-in order to maintain the normally acid condition of the juice (pH value about 5.4). There also may be a relationship between the aconitic acid content and the alkaloid content of the plant. These authors also observed that the juice from the immature growing portions of the sugar cane stalk-the tops which are normally discarded-contained from three to five times as much aconitic acid as the juice from the mature and millable portions of the cane. On the basis of the above findings the growing of sugar cane under carefully controlled conditions (e.g. soil composition and maturity of the cane) primarily for the production of aconitic acid is a distinct possibilitya’ although economic considerations would point away from this.

11. PHYSICAL PROPERTIES OF ACONITIC ACID

Aconitic acid, a white crystalline solid, exists as either the trans- or the cis-form with the trans-aconitic acid being the more stable form41 while the cis-acid is the stronger acid. The trans-aconitic acid exhibits a decomposition point rather than a true melting point. The values reported in the literature vary over a range from 185’ to 208’ and the observed decomposition point is a function not only of the purity of the acid but also of the temperature of the melting point bath when the sample is introduced and of the rate of heating.a4*42

The trans-aconitic acid is easily soluble in water and the solubility increases as the temperature rises. The acid is quite soluble in methanol, ethanol, aqueous ethanol, dioxane, slightly soluble in diethyl ether and can be recrystallized from water, ether or concentrated hydrochloric acid.

(40) Analyses of numerous molasses samples from various geographical areas as carried out in the research laboratories of The American Sugar Refining Co. also have revealed a considerable fluctuation of the aconitic acid concentration. A comparison of the aconitic acid concentration in these blackstrap molasses should serve as an indication of the variation in the aconitic acid concentration of the sugar cane juices from which the respective molasses were derived. Louisiana molasses in general contain more aconitic acid than Cuban blackstraps. Thus in a series of ten Louisiana samples the aconitic acid content ranged from 2.10 to 7.39 percent with an average value of 4.83 percent (based on dry solids). In a series of forty-four samples of Cuban molasses the aconitic acid content ranged from 1.64 to 4.37 percent with an average value of 3.20 percent (based on dry solids).

(41) For a discussion of this cis-trans relationship see page 244. (42) W. F. Bruce in “Organic Syntheses” (H. A. Blatt, editor), John Wiley and

Sons, Inc., New York, Coll. Vol. 11, 12-14 (1943).

ACONITIC ACID IN THE MANUFACTURE O F SUGAR 235

The physical properties of aconitic acid are listed in Table I.

TABLE I Physical Properties of Aconitic Acid

Melting Point

Solubility in Water

Ionization Constants ki

k i

Molecular Heat of Com- bustion

Refractive Indices of Crystals

trans-Aconitic Acid

Reported in range from 180' to

Refs. 11, 16, 17, 21, 34, 39, 42-48 208" (dec.)

18.6 g./100 ml. at 13" 5o

26.4 g./100 ml. a t 25" 5 L

110.7 g./100 ml. at 90" 5 1

1.31 X at 20" 62

1.58 X 10-8 a t 25" 53 1.36 x 10-846

3 .5 X a t 25" I4

1 . 1 X a t 15" 6 5

481.3 kcal. (constant volume)5' 476.3 kcal. (constant volume)6* 475.4 kcal. (constant pressure)&*

n, 1.4906g; 1.47534 np indeterminate ny 1.610L8; 1.64234

cis-Aconitic Acid

125" 49

1.19 X 10-3 at 20" 1.13 X lo-' 66

(43) M. Conrad, Ber., 32, 1007 (1899). (44) W. Hentschel, J. prakt. Chem., [2] 36, 205-6 (1887). (45) L. Claisen and E. Hori, Ber., 24, 12CL27 (1891). (46) T. H. Easterfield and W. J. Sell, J. Chem. SOC., 61, 1003-12 (1892). (47) H. 0. L. Fischer and Gerda Dangschat, Helu. Chim. Acta, 18, 1204-6 (1935). (48) P. Walden, Z. physik. Chem., 10, 563-79 (1892). (49) R. Malachowski and M. Maslowski, Ber., 61, 2521-25 (1928). (50) V. Dessaignes, Ann., Suppl., 2, 189 (1862). (51) Taken from a catalogue published by Chas. Pfieer and Co. (52) R. Malachowski, Bull. intern acad. polon., 1931A, 369-82. (53) J. Walker, J. Chem. Soc., 61, 696-717 (1892). (54) R. Wegscheider, Monats., 23, 599-668 (1902). (55) I. M. Kolthoff, "The Use of Color Indicators," 2nd Ed. (Berlin 1923) p. 166

(56) G. Semerano and L. Sartori, Gazz. chim. ital., 68, 167-73 (1938). (57) W. Louguinine, Ann. chim. phys. [6], 23, 206 (1891). (58) F. Stohmann and C. Kleker, Z. physik. chem., 10, 417 (1892). (59) E. K. Nelson, J. Am. Chem. SOC., 47, 568-72 (1925).

(Bedstein, Z I , 2nd suppl., p. 693).


111. ANALYTICAL ESTIMATION OF ACONITIC ACID

Three separate and distinct methods have been utilised for the determination of aconitic acid in sugar cane and/or sorghum products. One involves the extraction of aconitic acid from the sample with an organic solvent, the second is based upon the decarboxylation of aconitic acid while the third employs the polarographic techniq~e .~~"

The first of these, utilized by YoderJa5 McCalip and SeibertJa4 and by Balch, Broeg and Ambler,a7 provides for the extraction of the aconitic acid from the sample being investigated, usually with diethyl ether, and the subsequent isolation of the acid from the solvent. In dealing with solid samples, e.g. alkaline earth aconitates, evaporator scale, etc., the prescribed procedure is to dissolve the material in aqueous mineral acid and to extract the acid solution exhaustively with ether. The ether extract is then evaporated under reduced pressure, the dried residue titrated with standard alkali and the titratable acid calculated as aconitic acid. In dealing with such solid samples it is often necessary to make an additional determination for oxalic acid which otherwise would be assumed to be aconitic The aconitic acid in liquid samples is usually precipitated as the insoluble lead salt which is separated and treated as any other solid sample. In some cases this procedure is unnecessary and the liquid samples are merely acidified with a mineral acid and then extracted with ether.a7 This method for the determination of aconitic acid, however, requires a considerable amount of time and is further complicated by the interference of ether-soluble waxes and non-volatile acids.

The second method, developed by Ambler and Robert~,~@-~~ involves the decarboxylation of aconitic acid. These workers found that aconitic

(59a) A fourth method has recently been reported by K. Lauer and S. M. Makar (Anal. Chem., 23, 587-89 (1951)). These authors have reported an analytical procedure for the determination of aconitic acid which involves titration of an acidified (sulfuric acid) aqueous solution of aconitic acid a t the boiling point with a standard potassium permanganate solution. Since the oxygen consumption was found to vary with the concentration of the potassium permanganate solution this latter solution must be standardized against a pure known sample of aconitic acid. Procedures are described for the determination of aconitic acid in pure solutions, in technical aconitates and in molasses. While itaconic acid and its salts constitute an interference the authors outline a procedure in which the aconitic acid is separated from the itaconic acid as its mercurous salt before the aconitic acid determination is carried out.

(60) E. J. Roberts and J. A. Ambler, Anal. Chem., 19, 118-20 (1947). (61) J. A. Ambler and E. J. Roberts, Anal. Chem., 19, 877-78 (1947). (62) J. A. Ambler and E. J. Roberts, Anal. Chem., 19, 879-80 (1947). (63) J. A. Ambler and E. J. Roberts, Anal. Chem., 20, 880 (1948).

ACONITIC ACID IN THE MANUFACTURE OF SUGAR 237

acid and certain aconitates undergo a rapid decarboxylation when refluxed with anhydrous acetic acid-potassium acetate mixtures and one mole of carbon dioxide is liberated per mole of original aconitic acid. The carbon dioxide is collected and determined by any standard method. With liquid samples the aconitic acid is isolated as the lead salt which is carefully dried and then added to the acetic acid reagent.

A number of materialslsO water, certain organic acids, the most important of which is citric acid, and certain inorganic salts, interfere with the determination. The decarboxylation cannot be conducted in the presence of nitrates and when carbonates are present two separate determinations of carbon dioxide are necessary. The procedure is also inapplicable when oxidizing compounds which are soluble in the hot reagent are present. Another interfering substance, sulfur dioxidelB1 can be eliminated by the use of a saturated, acidified (sulfuric acid) solution of potassium dichromate to wash the gases evolved by the decarboxylation procedure.

Ambler and Robertslsz on further examination of the interference of citric acid, found that the addition of boric acid to the acetic acid- potassium acetate reagent prevented the decarboxylation of the citric acid. When boric acid was used in the reagent, however, i t was found that oxalic acid, galacturonic acid and mucic acid were slowly decarboxylated. Polyuronic acids were insoluble in the acetic acid reagent and were not decarboxylated by the procedure.63 In addition, when the decarboxylation procedure was applied to methylgalacturonide dihydrate it was found that the compound was easily soluble in the reagent but gave no carbon dioxide even after refluxing for three hours. The evidence indicated, therefore, that only the free galacturonic acid was decarboxylated in the aconitic acid method and that the potassium acetate- acetic acid reagent, being practically anhydrous, is unable to hydrolyze soluble uronides to the free uronic acids.

Although polarographic studies of aconitic acid in pure solutions had been made by several investigators, the application of such tech- niques to the determination of aconitic acid in sugarhouse products had been n e g l e ~ t e d . ~ ~ - ~ ~ Recently, however, a procedure has been developed

(64) L. Schwaer, Chem. Listy, 26, 485-89 (1932). (65) K. Shoji, Bull. Inst. Phys. Chem. Research (Tokyo), 9, 69-78 (1930); Absts.

(66) L. Schwaer, Coll. Czechoslou. Chem. Commun., 7 , 326-35 (1935). (67) G. Semerano and L. Sartori, Mikrochemie, 24, 130-33 (1938). (68) H. Siebert, 2. Elektrochem., 44, 768-69 (1938). (69) A. Miolati and G. Semerano, 2. Electrochem., 46, 226-28 (1939).

9-11 pub. with Sci. Papers Inst. Phys. Chem. Research (Tokyo), 12, No. 221-27.


whereby the amount of aconitic acid in sugar products can be quantitatively determined polar~graphically.~~

The determination of aconitic acid in comparatively pure solutions is not difficult and a well defined wave having a half-wave potential of -0.55 volt (referred to a saturated calomel electrode) is obtained when normal hydrochloric acid is used as the supporting electrolyte. 'For using most automatic recording polarographs with pure solutions a range in aconitic acid concentration of two to ten milligrams of acid per 100ml. is chosen as the most optimum conditions from the standpoint of sensitivity range and in order to obtain wave heights which are sufficiently large to minimize the errors due to measurement. With pure solutions a simple dilution with normal hydrochloric acid to the desired concentration is satisfactory.

In applying the procedure to blackstrap molasses a comparative method is employed in which the concentration of the unknown solution is determined by comparison of the wave height with that from a known concentration. Interfering substances are present in blackstrap molasses, however, which prevent the formation of a well defined current- voltage relationship and special treatment of such samples is therefore required. The removal of the interfering substances can be accomplished by treating the molasses sample with activated carbon. The conditions under which the samples are prepared must be rigidly standardized in order to obtain reproducible results and the following procedure is recommended :

A sample of the blackstrap molasses (25 9.) is transferred quantitatively to a 500 ml. Kohlrausch flask using only sufficient water to make the transfer; concentrated hydrochloric acid (20 ml.) is added, the contents of the flask are thoroughly mixed and then made up to volume with distilled water. An aliquot (20 ml.) of this solution is adjusted to a pH value of 10 f 0.2 with six normal sodium hydroxide and then diluted to 50 ml. with distilled water. The entire 50 ml. is transferred to a 125 ml. Erlenmeyer flask to which is added a carbon-filter aid mixture (approximately 2 9.) composed of 40% Darco KB and 60% filter aid. (The type of filter aid used is not critical.) The flask is placed upon a hot plate (at approximately 150°), allowed to remain for exactly four minutes and then immediately filtered, using a very rapid filter paper. An aliquot (20 ml.) of the filtrate is made up to 50 ml. with normal hydrochloric acid and the solution is then ready for the electrolysis cell. The sample is placed in the cell, a current of nitrogen is bubbled through it for ten minutes, the bridge is placed in operation and the polarogram is obtained. The wave height of the polarogram is measured, corrected for the sensitivity employed and then by comparison with the standard solutions the aconitio acid concentration is calculated. The usual precautionary measures apply which are necessary for all polarographic procedures (e.g. temperature control, drop rate, protection from vibration, etc.).

(70) R. W. Liggett, J. A. Devlin and S. M. Cantor, Unpublished work from laboratories of American Sugar Refining Co.

ACONITIC ACID I N THE MANUFACTURE O F SUGAR 239

Numerous other methods, many of which are useful only in special cases, have been utilized to determine aconitic acid in various solutions. The color reaction of aconitic acid with acetic anhydride has been employed to a very minor extent in sugar t e c h n o l 0 g y . ~ 3 ~ ~ ~ ~ ~ ~ Malachow- skilS2 in analyzing solutions of cis- and trans-aconitic acid, made use of the difference in the specific conductance of the two compounds to determine the composition of the solutions. Krebs and Eggle~ton,?~ seeking a method for the determination of cis-aconitic acid, used aconitase to convert the cis-aconitic acid to citric acid which was then analyzed according to the method of Pucher, Vickery and Lea~enworth.?~ Ksebs and Eggleston also utilized this procedure for studies of the equilibrium between the cis- and trans-aconitic acid in aqueous solutions. Ambler and Roberts,7s who were studying the stability of cis-aconitic acid in aqueous solutions, made use of the difference in the solubilities of the st,rontium salts of the cis- and trans-acids to determine the composition of the equilibrium mixtures obtained. The latter procedures, however, are inapplicable to the determination of aconitic acid in sugarhouse products.

IV. THE RECOVERY OF ACONITIC ACID IN THE MANUFACTURE OF

SUGAR

During the last ten years considerable interest has developed in connection with the commercial recovery of aconitic acid from sugar- containing liquors such as sorghum juice or sugar cane juice. The isolation of aconitic acid from raw sorghum juice, which contains a considerable amount of aconitic acid, can be carried out before the sugar is crystallized. In sugar cane juice, however, the amount of aconitic acid is much lower and usual starting points for the isolation of aconitic acid from this source are the “B” molasses which is obtained after two crops of sugar have been crystallized from the juice, and the final blackstrap molasses.

The first indication that the recovery of insoluble aconitates from these sugar-containing juices was feasible was the initial work by Ventr433*76 and Ventre and Paine.?? These workers found that during the evaporation of sorghum juice an insoluble aconitate salt separated in amounts

(71) 0. Furth and H. Hermann, Biochem. Z., 280, 448-57 (1935). (72) E. K. Ventre, J. A. Ambler, H. C. Henry, S. Byall and H. S. Paine, I d . Eng.

(73) H. A. Krebs and L. V. Eggleston, Biochem. J . , 38, 426-37 (1944). (74) G. W. Pucher, H. B. Vickery and C. S. Leavenworth, Ind. Eng. Chem., Anal.

(75) J. A. Ambler and E. J. Roberts, J . Org. Chem., 13, 399-402 (1948). (76) E. K. Ventre, Sugar, 36, No. 1, 36-37 (1941). (77) E. K. Ventre and H. S. Paine, U. S. Pat. 2,280,085 (Apr. 21, 1942).

Chem., 38, 201-4 (1946).

Ed., 6, 190 (1934).


which .made it practicable to recover this salt as a by-product. The only chemical addition involved in this early work was that of an alkaline earth clarifying agent which was employed to adjust the pH value of the juice to within the range 6.8 to 7.2. Since the alkaline earth aconitates are more insoluble at higher temperatures, the temperatures employed subsequently in the evaporation of the sorghum juice, were of the proper range to cause the formation of the insoluble aconitates.

Ventre and c o - w o r k e r ~ , ~ ~ ~ ~ ~ in later work, found that the addition of calcium chloride greatly increased the amount of aconitates thus recovered. The procedure used was as follows: lime was added to a quantity of sorghum juice to adjust the pH value to approximately 6.9. The amount of lime required for this treatment was calculated as equivalents of aconitic acid and this portion of the aconitic acid in the original sample of juice was assumed to be free aconitic acid. The remaining aconitic acid was assumed to be bound in the form of a soluble aconitate. An amount of calcium chloride equivalent to the bound aconitate was then added to the juice for the precipitation. The insoluble aconitates which precipitated during the subsequent evaporation of the juices were removed by some appropriate means at a later point in the sugar recovery system.

Following this initial work on the isolation of aconitates from sorghum juice the same process was applied to sugar cane "B" molasses.82~37~88~Te Ambler, Turer and Keenan,80 in 1945 investigated various salts of aconitic acid and reported that the dicalcium magnesium aconitate hexahydrate and aconitates containing lesser amounts of magnesium were less soluble than either of the hydrates of tricalcium aconitate. Thus Ambler, Roberts and Weissborn, Jr.,81 and Ambler and Roberts,82 soon reported an improved process for the recovery of aconitates from molasses in which both calcium chloride and magnesium chloride were added to the molasses. According to these workers the optimum conditions for aconitate recovery were fivefold:

0 a. The molasses was diluted to 50" to 55" Brix. b. Calcium oxide or hydroxide were the best agents for adjustment

of the pH value. The optimum pH values were found to be within the range of 6.5 to 6.8.

(78) E. K. Ventre, J. A. Ambler, S. Byall and H. C. Henry, U. S. Pat. 2,359,537

(79) Anon., Sugar Bull., 23, No. 19, 173-74 (1945). (80) J. A. Ambler, J. Turer and G. L. Keenan, J . Am. Chem. SOC., 67, 1-4 (1945). (81) J. A. Ambler, E. J. Roberts and F. W. Weissborn, Jr., U. S. Bur. Agr. Id.

(82) J. A. Ambler and E. J. Roberts, U. 5. Pat. 2,481,557 (Sept. 13, 1949).

(Oct. 3, 1944).

Chem., Mimeographed Circ. Ser. AIC 196 (1946).

.4CONITIC ACID I N THE MANUFACTURE OF SUGAR 24 1

c. Calcium chloride and magnesium chloride were used to increase the calcium and magnesium content of the molasses sample. The most satisfactory results were obtained by adding a solution containing three parts of anhydrous calcium chloride and one part of anhydrous magnesium chloride for every five parts of aconitic acid in the solution. These were the least amounts required. Larger quantities had no affect upon the aconitate recovery but increased unnecessarily the amount of ash and particularly chlorides in the molasses returned to the sugarhouse.

d. Best results were obtained if the precipitation were carried out at temperatures between 93" and 99".

e. The reaction temperature was maintained for at least forty-five minutes. Extension of the reaction time beyond one hour was found to have very little affect upon the recovery.

The findings of G o d ~ h a u x ~ 3 9 ~ ~ agreed well in most respects with the above optimum conditions but differed in respect of the chemical addition required. The latter worker and his associates indicated that magnesium chloride was not necessary for the precipitation of the aconitic acid but actually was detrimental to sugar recovery. More important was the realization that a definite chemical addition could not be used for all molasses samples. Certain molasses when limed and heated precipitated a large percentage of aconitic acid as insoluble alkaline earth aconitates whereas other molasses required excessive amounts of chemical addition. This inability t o designate a definite chemical addition for all molasses was also discussed by Ventre;85 the suggested resolution for this problem was the use of preliminary laboratory experiments to determine the optimum conditions for each molasses.

A more recent investigation86 has also indicated that it is impossible to state a specific procedure for the recovery of aconitates which is appli- cable to all molasses but that each type of molasses must be investigated and handled as a separate entity. The samples of Cuban b1ackstra.p molasses which were studied gave good recoveries of insoluble aconitates by simple dilution and heating, and the addition of soluble calcium salts in the usual manner to these molasses did not materially improve the aconitate recoveries. Louisiana blackstrap molasses in general, however, were found to require the addition of large amounts of chemicals

(83) R. J. Fume and L. Godchaux, 11, Sugar Research Foundation (New York),

(84) Anon., DeLauaZ Centrzfugul Rev., 16, No. 1, pp. 3, 4, 11 (1949). (85) E. K. Ventre, U. S. Pat. 2,469,090 (May 3, 1949). (86) R. E. Miller, R. Netsch and R. W. Liggett, unpublished data.

BUZZ., 4, NO. 20, 78-81 (1948).


Louisiana Molasses Samples 1 2 3 4 5 6 7 8 9

10 Cuban Molasses Samples

13 14 15 16

before satisfactory yields of aconitate could be recovered. Typical results in relation to the precipitation of aconitates from Cuban and Louisiana blackstrap molasses are presented in Table 11.

TABLE I1 Precipitation of Aconitates from Cuban and Louisiana Blackstrap Molasses

41 66 27 67 6 61

19 67 0 66

32 74 42 48 2 61 4 57

12 70

68 74 68 75 55 65 57 64

Percent of Total Aconitic Acid Precipitated as Insoluble Aconitates

Procedure 10 1 Procedure IP

Molasses Sample

0 Procedure I: The molasses was diluted to 53" Brix and heated for one hour a t 90' with mechanical stirring. A portion of the reaction mixture was then centrifuged and the exhausted molasses (supernatant liquor) was analyzed for aconitic acid.

* Procedure 11: The molasses was diluted to 53" Brix and a CaO slurry (10% by weight) was added to a pH value of 7.0. The desired amount of 3M CaCll solution was added (enough to bring the total equivalents of calcium ion up to 1.5 times the number of equivalents of aconitic acid present), and the reaction mixture was heated at 90" for several hours with mechanical stirring. The reaction mixture was then centrifuged and the exhausted molasses was analyzed for aconitic acid.

The insoluble aconitate salts isolated by the aforementioned procedures usually crystallize as hexahydrates and generally contain both calcium and magnesium even when magnesium salts have not been added to the molasses prior to the aconitate precipitation. It is reportedso that the salts have the optical-crystallographic characteristics of dicalcium magnesium aconitate hexahydrate, CazMgAcon2.6H20, although their magnesium content is usually less than that of this salt. Ambler, Turer and Keenanso made several preparations of this substance which contained magnesium in quantities ranging from almost theoretical down to

ACONITIC ACID I N T H E MANUFACTURE OF SUGAR 243

approximately 25% of theory, and which, regardless of the magnesium content within these limits, were homogeneous and showed identi- cal optical-crystallographic properties. These crystalline hexahydrates obtained from molasses show no indication of being physical mixtures of insoluble calcium aconitate with soluble magnesium aconitate since it is impossible to separate magnesium aconitate from the salts by leaching with hot water.72 The salts, therefore, are believed to be members of a series of mixed crystals or solid solutions of dicalcium magnesium aconitate hexahydrate and tricalcium aconitate hexahydrate.s0ss2

D. W. C0llier,~7 in a recent patent, reports that the use of a small amount of barium and/or strontium salts in conjunction with the calcium and/or magnesium salts ordinarily used in the precipitation of the aconitic acid from molasses gives much higher aconitate recoveries than those previously reported. The use of a small amount of either barium or strontium salts causes much greater precipitation than an equivalent amount of calcium ion. Thus the use of a mixture of a calcium salt and a barium salt which contained cations equivalent to the aconitic acid (90% equivalence of calcium ion and 10% equivalence of barium ion) in a trisodium aconitate solution was found to give a residual aconitate solubility (3.1 g./liter) much lower than that (12.6 g./liter) obtained when an equivalent amount of calcium ion was used alone.

The procedure as outlined by Collier is very similar to that previously described. The molasses is neutralized with lime to a pH value of 6.8, heated, and the desired amount of an aqueous calcium chloride solution is added. This mixture is subjected to the usual precipitation conditions (90-95', mechanical agitation) for approximately forty-five minutes and the soluble barium and/or strontium salts are then added. The heating is continued for another forty-five minutes and the insoluble aconitates are recovered from the molasses. The salts obtained from this latter process thus contain barium and/or strontium ions in addition to the calcium and magnesium cations usually present.

From a consideration of these data, it would appear that precipitation of insoluble aconitates from molasses is an extremely complicated reaction and is a function not only of the relative amounts of acid and metallic ions present but also the status of the acid in solution. In this latter respect, it would seem that aconitic acid exists in molasses not only as free ionized acid but also as soluble complexes and the conversion of these soluble complexes to insoluble salts is an important yield-governing consideration.

One phase of the aconitate precipitation process which has not been previously mentioned but is extremely important is the disposal of the

(87) D. W. Collier, U. S. Pat. 2,513,287 (July 4, 1950).


molasses after the insoluble aconitates have been removed. With the present demand for blackstrap molasses for. fermentations and livestock feed and the comparatively high price level of molasses it is quite evident that a careful examination of the effect of the various cations employed for the precipitation upon the final use of the molasses is necessary.

In large scale operations the aconitic acid is usually recovered from the crude calcium magnesium aconitates by acidification with a mineral acid followed by the crystallization of the aconitic acid from the liquors obtained. Thus Ventre, Henry and Gayle,88 acidified the crude salts with dilute sulfuric acid. The insoluble calcium sulfate was removed by filtration and the aconitic acid was separated from the magnesium sulfate by fractional crystallization.

Ambler and R o b e r t ~ , ~ ~ , ~ ~ in subsequent work, found that if the calcium magnesium salts were heated to remove a portion of the water of hydra- tion the magnesium content could be replaced by calcium by treating the dried salts with a hot concentrated calcium chloride solution. Thus the aconitates could be converted into tricalcium aconitate and subsequent treatment of this salt with sulfuric acid enabled the removal of the cations as insoluble calcium sulfate. The crystallization of aconitic acid from such a filtrate was therefore not complicated by the necessity of a fractional crystallization to separate the aconitic acid and the magnesium sulfate formerly obtained.

Hydrochloric acid has also been used for the acidifi~ation.~~ In this case the aconitic acid is then crystallized from the solution of aconitic acid and alkaline earth chlorides.

V. CHEMISTRY AND USES OF ACONITIC ACID

Because of the polyfunctional character of aconitic acid, it being both an unsaturated acid and a polybasic acid, the compound can undergo a variety of chemical reactions. Application of derivatives obtained by such reactions has aroused considerable interest in the plasticizer, wetting agent and resin manufacturing fields.

Much of the chemical behavior of aconitic acid is closely related to the chemistry of maleic and fumaric acids. As mentioned earlier, aconitic acid can exist as either the trans- oqthe cis-isomer. It was not until 1928, over one hundred years after its initiaI discovery, that the trans-aconitic

(88) E. K. Ventre, H. C. Henry and F. L. Gayle, U. S. Pat. 2,345,079 (Mar. 28,

(89) J. A. Ambler and E. J. Roberts, U. S. Pat. 2,432,223 (Dec. 9, 1947). (90) H. F. Reeves, Jr., U. S. Pat. 2,614,010 (July 14, 1950).

1944).

ACONITIC ACID I N THE MANUFACTURE OF SUGAR 245

acid was identified as the more stable form,46~48~49~91-g7 the so-called “ordinary” aconitic acid. Only a few investigations have been made, however, in connection with the cis-trans equilibrium and the factors which affect this equilibrium. Malachowskils2 working with aqueous solutions of cis- and trans-aconitic acid, has reported that the equilibrated solutions contain approximately 85 per cent trans-aconitic acid. The amount of cis-aconitic acid increases slightly with increasing temperatures and with dilution of the solution. Krebs and Eggle~ton’~ found that in neutral solution sodium cis-aconitate is quite stable and that the most rapid and extensive conversion of the cis- to the trans-acid takes place in ‘strongly acidic or strongly alkaline solutions. Ambler and R ~ b e r t s , ’ ~ in a further investigation of the affect of the pH value of the solution upon the stability of cis-aconitic acid, found that cis-aconitates are stable in neutral and slightly alkaline solution, but unstable at high alkalinities, especially if the solutions are heated. In acid solutions the stability decreases as the acidity of the solution increases.

As a result of the cis-trans isomerization two isomeric aconitic anhy- drides are known. Treatment of the trans-aconitic acid with acetyl ~hloride,46JJ6*~6 or with acetic anhydridelg60g7 leads to the formation of both the cis-aconitic anhydride (11) and the trans-aconitic anhydride (111). More recently i t has been reported that good yields of the cis-anhydride

HC-C=O

/ I ‘0

HC-C=O €IOOC--c ‘1, ‘()

I1 / I121:-C=0 / c-c=o Hz -COOH

I1

A I11

can be obtained by refluxing a mixture of trans-aconitic acid in xylene with catalytic amounts of p-toluenesulfonic The cis-aconitic acid can be prepared by the hydrolysis of the cis-anhydride under carefully controlled conditions49 while the trans-acid is prepared in the laboratory by the sulfuric acid dehydration of citric

Aconitic acid is easily esterified by conventional methods and physical properties of some of the trialkyl aconitates are presented in Table 111.

(91) A. Michael, Ber., 19, 1381-86 (1886); Amer. Chem. J. , 9, 193 (1887). (92) S. Ruhemann and K. J. 1’. Orton, Ber., 27, 3449-57 (1894). (93) R. Anschtitz and W. Bertram, Ber., 37, 3967-70 (1904). (94) H. Rogerson and Jocelyn F. Thorpe, J . Chem. SOC., 89, 631-52 (1906). (95) N. Bland and Jocelyn F. Thorpe, J . Chem. SOC., 101, 1490-98 (1912). (96) P. E. Verkade, Rec. trau. chim., 40, 381-86 (1921). (97) R. Malachowski, M. Giedroyc and Z. Jerznianowska, Ber. 61,2525-38 (1928). (98) W. P. Ericks and E. R. Meincke, U. S. Pat. 2,345,041 (Mar. 28, 1944).


Certain of these trialkyl aconitates are reported to be effective for con- trolling soft-bodied and sucking insects.eQ The six isomeric monomethyl cis- and trans-aconitates were prepared by Malachowski, Giedroyc and Jerzmanowskas7 and were obtained as crystalline solids melting sharply within the range of 100" to 150". The monomethyl anhydro-cis-aconitate (IV) also prepared by these workers was a low melting solid (37-38"). Ericks and Meinckes8 have reported the use of cis-aconitic anhydride to prepare monoalkyl aconitates and also dialkyl aconitates although the resulting esters were not described fully nor were the physical properties reported.

HC-C=O

I/ >O c-c=o I

H~CI-COOCH~ IV

TABLE I11 Physical Properties of Trialkyl Aconitates

Trialkyl Aconitate

Trimet h y P Triethylgga Tri-n-propyl99

Tri-act .-amy199

Tri-2-ethylhexylg'J Trilaurylgg Tristearyl'J'J

M. P. I B.P.

Liquid Liquid Liquid

Liquid

Liquid 10"

5 5 . 5 O

160' at 20 mm. 172' at 18 mm. 157"-162" a t 2

193-197" at 3 mm.

mm. - -

Refractive Index at 26'

- -

1.4521

1.4540

1 .4600 1.4578

Early investigators had reported the preparation of aconityl chloride in low yields by the use of phosphorus oxychloride and phosphorus pentachloride. loo~lol Froschl and Maier,Io2 however, tried unsuccessfully to repeat this work and reported that the use of thionyl chloride was also without success.

(99) E. R. Meincke, U. S. Pat. 2,475,629 (July 12, 1949). (99a) C. K. Ingold, J. H. Oliver and Jocelyn F. Thorpe, J . Chem. Soc., 126,

(100) Klimenko and Buchstab, J. Russ. Phys.-Chem. SOC., 22, 99 (1880) (Beilstein,

(101) A. Michael and G. Tissot, J . prakt. Chem., NF [2], 62, 33143 (1895). (102) N. Froschl and A. Maier, Monatsh., 69, 274 (1932).

2128-36 (1924).

11, 852).

ACONITIC ACID I N THE MANUFACTURE OF SUGAR 247

The decarboxylation of aconitic acid to itaconic acid (V) and to citraconic acid (VI) proceeds easily and when carried out under controlled

XrCOOH II HC-COOH

V VI

conditions leads primarily to itaconic acid. Early workers had found that heating aconitic acid above its melting point or in aqueous solutions under pressure led to the formation of itaconic a ~ i d . ~ J ~ ~ J ~ ~ Ambler and coworkers,82-'06J06 after finding that a small amount of an inorganic aconitate catalyzed the decomposition of aconitic acid in aqueous solution to itaconic acid, utilized the crude calcium magnesium aconitates obtained from molasses as the starting materials. Enough sulfuric acid was added t o the alkaline earth aconitates to convert a portion of them to the free acid while a portion remained as the aconitates t o catalyze the decomposition.

Tricarballylic acid, 1,2,3-propanetricarboxylic acid, is produced by the reduction of aconitic acid by catalytic method^,^^^-^^' electrolyti- ~ally,1~~-1~4 or by sodium amalgam.60s116 Sulfotricarballylic acid (VII), its salts, and its esters have become of interest recently due to their

H&-COOH H2C-COOR

Ha -COOH

M03s-t-c00R Hz -COOR

VIII

HoQ-fCooH VII

(103) L. Pebal, Ann., 98, 67-98 (1856). (104) T. Swarts, Jahresber. Fortschritte Chem., 579 (1873); Bull. acad. roy. Belg.

(105) J . A. Ambler and A. L. Curl, U. S. Pat. 2,448,506 (Sept. 7, 1948). (106) E. J. Roberts, J. A. Ambler and A. L. Curl, U. S. Pat. 2,448,831 (Sept. 7,

(107) S. Fokin, J . Russ. Phys.-Chem. SOC., 40, 316 (Chem. Zen&., 1908, ZZ, 1996). (108) S. Fokin, Z . Angew. Chem., 22, 1492-1502 (1909). (109) J. Boeseken, B. Van Der Weide and C. P. Mom, Rec. trau. chim., 36,26&87

(110) B. B. Allen, B. W. Wyatt and H. R. Henae, J . Am. Chem. SOC., 61, 843-46

(111) R. Malachowski, Bull. intern. acad. polon. sci., 1919A, 265-73. (112) C. Marie, Compt. rend., 136, 1331-32 (1903). (113) U. Pomilio, 2. Elektrochem., 21, 444-48 (1915). (114) V. V. Levchenko, J . Gen. Chem. (U.S.S.R.), 18, 1237-44 (1948). (115) H. Wichelhaus, Ann., 132, 61-66 (1864).

121, 36, 7 (1873).

1948).

(1916).

(1939).


surface active p r o p e r t i e ~ . ~ ~ ~ ~ ~ ~ J ~ ~ Salts of this acid are usually prepared by treating aconitic acid in neutral or slightly acidic solutions with sodium bisulfite. The sulfotricarballylic acid may be liberated by treatment with mineral acids and is easily esterified by conventional methods to yield salts of trialkyl sulfotricarballylates (VIII).

A series of sulfotricarballylic acid derivatives of potential use for the resolution of crude oil field emulsions of the water-in-oil type and as detergents has been described by Ericks and M e i n ~ k e . ~ ~ The preparation of these derivatives, salts of monoalkyl sulfotricarballylates and of dialkyl sulfotricarballylates, is illustrated by the following series of reactions.

HC-C=O HC-COOR HC-COOR HC-COOR - HnO R'OH ( 1 >O (!!-GOOH --+ L = O - &-COORr

c-c=o Hz c! -COOH HA-COOH H2A-cOOH HzC-C=O I ?

I1 IX 1

X XI 1

HzC-COOR H&-COOR HzC-COOR

A R'OH b - HsO HO&d-COOH - H O a S -C=O - HOaS- -GOOR'

Ht b -GOOH Hz b -GOOH

HpC-C=O

XI1 XI11 XIV

Numerous other uses of aconitic acid or its derivatives have been Many of these relate to their described in the patent literat~re.~~s-l42

(116) National Oil Products Go., Brit. Pat. 551,246 (Feb. 15, 1943). (117) P. Nawiasky and G. E. Sprenger, U. S. Pat. 2,315,375 (Mar. 30, 1943). (118) N. Oelwerke and G. van der Lane, Brit. Pat. 530,916 (Dec. 24, 1940). (119) T. Curten, Ger. Pat. 722,356 (May 21, 1942). (120) T. Habu, Jap. Pat. 93,028 (Sept. 29,1931) (Chem. Abstracts, 26,4488 (1932));

(121) T. Habu and S. Ogura, Jap. Pat. 111,256 (June 21, 1935) (Chem. Abstracts,

(122) E. F. Isard, U. S. Pat. 1,993,552 (Mar. 5, 1935). (123) H. Kraikalla and W. Wolff, U. S. Pat. 2,039,243 (Apr. 28, 1936). (124) H. M. Kvalnes, U. S. Pat. 2,091,241 (Aug. 24, 1937). (125) C. N. Anderson, U. S. Pat. 2,118,033 (May 24, 1938). (126) E. T. Clocker, U. S. Pat. 2,188,883 (Jan. 30, 1940); 2,188,884 (Jan. 30,

1940); 2,188,885 (Jan. 30, 1940); 2,188,886 (Jan. 30, 1940); 2,188,888 (Jan. 30, 1940); 2,188,889 (Jan. 30, 1940); 2,188,890 (Jan. 30, 1940); 2,275,843 (Mar. 10, 1942).

(127) M. W. Perrin, E. W. Fawcett, J. G. Paton and E. G. Williams, U. S. Pat. 2,200,429 (May 14, 1940).

ibid., 110, 730 (May 13, 1935) (Chem. Abstracts, 30, 2283 (1936)).

SO, 2284 (1936)); ibid., 111,259 (June 21, 1935) (Chem. Abstracts, 30, 2284 (1936)).

ACONITIC ACID I N T H E MANUFACTURE O F SUGAR 249

incorporation in the preparation of various polymers such as copolymers of alkyl aconitates and vinyl chloride,13’ high molecular weight polyesters prepared from mixtures of ethylene glycol, isopropylene glycol, sebacic acid and aconitic and as plasticizers in the preparation of stabi- lized vinylidene chloride ~ompositions.13~

(128) H. S. Rothrock, U. S. Pat. 2,221,662 (Nov. 12, 1940); 2,221,663 (Nov. 12,

(129) A. Hill, U. S. Pat. 2,230,351 (Feb. 4, 1941). (130) G. F. D’Alelio, U. S. Pat. 2,260,005 (Oct. 21, 1941); 2,288,315 (June 30,

1942); 2,308,494 (Jan. 19, 1943); 2,308,495 (Jan. 19, 1943); 2,319,798 (May 25, 1943); 2,319,799 (May 25, 1943); 2,323,706 (July 6, 1943); 2,232,898 (Oct. 26, 1943); 2,337,- 873 (Dec. 28, 1943); 2,337,874 (Dec. 28, 1943); 2,340,109 (Jan. 25, 1944).

(131) A. W. Hanson and W. C. Goggins, U. S. Pat. 2,273,262 (Feb. 17, 1942). (132) M. C. Agens, U. S. Pat. 2,319,576 (May 18, 1943). (133) C. M. Blair, Jr., U. S. Pat. 2,375,516 (May 8, 1945); 2,384,595 (Sept. 11,

(134) F. J. Kaszuba, U. S. Pat. 2,380,896 (July 31, 1945). (135) M. C. Agens and B. W. Nordlander, U. S. Pat. 2,404,204 (July 16, 1946). (136) E. L. Kropa, U. S. Pat. 2,409,633 (Oct. 22, 1946); 2,443,741 (June 22, 1948). (137) F. W. Cox, U. S. Pat. 2,419,122 (Apr. 15, 1947). (138) C. J. Fro~ch, U. S. Pat. 2,423,093 (July 1, 1947). (139) D. E. Adelson and H. F. Gray, Jr., U. S. Pat. 2,426,913 (Sept. 2, 1947). (140) T. W. Evans and D. E. Adelson, U. S. Pat. 2,435,429 (Feb. 3, 1948). (141) C. Struyk and S. C. Dollman, U. S. Pat. 2,523,160 (Sept. 19, 1950); 2,523,

(142) P. 0. Tawney, U.S. Pat. 2,553,430 (May 15, 1951); 2,553,431 (May 15,

1940); 2,321,942 (June 15, 1943).

1945).

161 (Sept. 19, 1950).

1951).

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