INDUSTRIAL AND ENGINEERING CHEMISTRY
146
orange to a light green. Solutions having a high temperature cause the indicator to fade (9).
Results The volumetric ceric sulfate method compared with the volumetric permanganate ( I ) , Munson-Walker (1, 6), and Shaffer-Hartman (6) methods gave essentially the same results on a known dextrose solution, an aqueous extract of tung leaves, and a mixture of the extract and a known dextrose solution (Table I). It is evident that ceric sulfate can be used successfully in this procedure.
Literature Cited (1) Assoc. 0 5 c i a l Agr. Chem.. Official and Tentative Analysis, 5th ed., pp. 125, 138-9, 500 (1940).
Methods of
Vol. 14, No. 2
Furman, H., J . Am. Chem. Soc., 50, 755-64 (1928). Hassid. W. Z.. IND.ENG.CHEM..ANAL.ED..9. 228-9 (1937). (4) Kraybill, H. R., Youden, W. J., and Sullivan; J. T., As'soc.
(2) (3)
(5) (6) (7) (8) (9) (10) (11)
>.
Oficial A p . Chem., 19, 125 (1936). Munson, L. S., and Walker, P. H., J. Am. Chem. Soc., 28,663-86 (1906). Shaffer, P. A,, and Hartman, A. F., J . Biol. Chem., 45, 349-64 (1921). Smith, G. F., "Ceric Sulfate", Vol. 1, 4th ed., pp. 31, 52, Columbus, Ohio, G. Frederick Smith Chemical co., 1940. Stegman, R. A., and Englis, D. T., Trans. Illinois State Acad. SCi., 27, 75-6 (1934). Sullivan, J. T., J . Assoc. Oficial Agr. Chem., 18, 382-6 (1935). Vanossi, R., and Ferramola, R., Anales soc. cient. Argentina, 121, 59-73 (1936). Walden, G. H., Hammett, L.P., and Chapman, R . P., J . Am. Chem. Soc., 55,2649-54 (1933).
Content of LPirnaric Acid in Pine Oleoresin Improved Methods for Its Determination ELMER E. FLECK AND SAMUEL PALKIN Bureau of Agricultural Chemistry and Engineering, United States Department of Agriculture, Washington, D. C.
T
HE I-pimaric acid content of pine oleoresin has been a matter of considerable speculation for a long time.
Dupont (4) advanced a theoretical basis for predicting the amount of individual resin acids present in pine oleoresin. He estimated the amounts of these acids present by observation of the optical rotation of mixtures ( 2 ) and by isomerization of the I-pimaric acid present with mineral acid (3). This type of analysis was further developed by Sandermann (6) for I-pimaric acid present in oleoresin by change in optical rotation when this ~ is isomerized with mineral acid to I-abietic acid of [ a ]-280" -104". The method is subject t o the objection that, acid DI.[ in addition to I-pimaric acid, other unidentified acidic constituents are also isomerized, under the condition of this determination, to l-abietic acid. The recent discovery that I-pimaric acid readily forms a DielsAlder addition compound in the cold with maleic anhydride ( 1 , . 7 ) has provided a new basis for analytical determinations of l-pimanc acid and possibly other conjugated double bond acids that might be present. The mild conditions of this reaction and the technique for quantitative removal of the addition product, without altering the other unstable acids present in the complex, make possible a new approach to the problem of the "sapinic acids". Sandermann ( 5 ) used this reaction with maleic anhydride t o replace the isomerization of Gpimaric acid with mineral acid. The change in optical rotation was observed and used as a basis for calculating the amount of I-pimaric acid present. Another method developed by Sandermann (5, 6) also made use of the addition product of maleic anhydride with I-pimaric acid, but depended upon acidimetric titration and not rotation measurements. In this method a known weight of resin acids was reacted with an excess of maleic anhydride, the excess maleic anhydride was then extracted, and the resin acids plus the 1pimaric acid addition product was titrated with alkali. The per cent of E-pimaric acid present was calculated from the increased alkali needed to neutralize the reaction product over that needed for the original acids. I n the present work this maleic anhydride procedure was modified so that a known amount of maleic anhydride was added t o a known amount of resin acid mixture in n-pentane solution. After reaction had taken place the excess maleic anhydride was extracted quantitatively and the aqueous extract titrated with alkali. Since the addition compound of I-pimaric acid is not soluble in water, this makes possible the determination of the amount of maleic anhydride reacted and from that the per cent of l-pimaric acid can be calculated. While no reaction between maleic anhydride and gum turpentine constituents is to be expected a t room temperatures,
nevertheless to eliminate the possibility of abnormal behavior blank determinations with turpentine were made. These showed that the reaction of maleic anhydride with turpentine at a temperature around 20" C. for a period of 4 hours was smaller than the limits of experimental error. This makes possible the determination of l-pimaric acid present in pine oleoresin without the necessity of first extracting the acidic portion of the oleoresin from the turpentine. I n this method any unknown compound, present in the oleoresin, that reacts with maleic anhydride at room temperature will give too high a value for the I-piniaric acid content of oleoresin. A new method for the determination of I-pimaric acid makes use of the insolubility of the I-pimaric acid addition product of maleic anhydride in n-pentane. A t 20" C. the solubility of this addition product was found to be 0.03 gram in 100 cc. of n-pentane. For analysis the pine oleoresin was dissolved in n-pentane and filtered free from dirt, chips, and water. The concentration of the acidic constituents was determined by titration with alkali and then a known volume of n-pentane solution xas reacted with an excess of maleic anhydride in acetone solution. The addition product crystallized from solution and was filtered off, washed, dried, and weighed. This addition product was found to be of high purity. All samples melted above 220" C. The purified material melts a t 226-229" C. ( 7 ) . The completeness of the removal of this addition compound from n-pentane solution is illustrated by the reaction of pure I-pimaric acid with maleic anhydride. Thus, of a 3-gram sample of i-pimaric acid, less than 0.05 gram of noncrystalline material was removed from the mother liquor after the addition compound had been removed by filtration. The quantitative nature of this reaction a t room temperature was not disturbed by the presence of other nonreacting resin acids and turpentine. This was shown by addition of pure I-pimaric acid to an n-pentane solution of oleoresin to increase the I-pimaric acid content from 32.7 to 35.9 per cent. I-Pimaric acid found in the fortified oleoresin was 36.2 per cent. The solubility of the addition product is increased somewhat by the small amount of acetone added and also b y the resin acids of unknown identity that are present in the oleoresin.
ANALYTICAL EDITION
February 15, 1942
These factors would produce too low a value for I-pimaric acid of pine oleoresin. These two methods for the analysis of pine oleoresin were applied t o a series of longleaf oleoresin samples collected during the season of 1940 near Olustee, Fla. The samples were stored at -5" C. and all determinations were made at the end of the turpentining season. These results (Table I) are in fair agreement with those reported b y Sandermann (5) and show that I-pimaric acid does not account for the major portion of the primary acids present in pine oleoresin. The straight-line decrease of Z-pimaric acid content of longleaf oleoresin with the season indicates that the controlling factor in the decrease of Ipimaric acid is the length of the face on the tree over which the oleoresin flows, exposed to the air, to reach the cup. Evidently I-pimaric acid is not appreciably isomerized b y rise in temperature during the hot part of the season, as is indicated b y Sandermann ( 5 ) . If temperature were a governing factor, the I-pimaric acid content of oleoresin should reach a minimum at midsummer and rise again with the cooler weather of fall.
TABLE I.
l-PIMARIC
Month, 1940 April June August
November
ACID PRESENT I N RESINACIDFR.4CTION PINEOLEORESIN
OF
I-Pimaric Acid Volumetric Gravimetric method, % method, % 38.4 36.0 38.7 36.3 34.2 36.6 33.7 35.4 32.4 35.1 31.7 33.2 31.3 33.7 30.5
Quantitative Reaction of I-Pimaric Acid with Maleic Anhydride A solution of 3.00 grams of I-pimaric acid, [0]9-274O (2 per
cent absolute alcohol), in 150 cc. of n-pentane was stirred while a solution of 1.3 grams of maleic anhydride in 10 cc. of absolute ether was added. Stirring was continued for 2 hours, during which time the addition compound crystallized from solution. It was removed by filtration and the filtrate was extracted with water until the aqueous extracts reacted neutral to Congo red. The n-pentane was then distilled and the noncrystalline residue weighed less than 0.05 gram.
Description of Methods Approximately 100 grams of well-mixed pine gum were dissolved in 200 cc. of n-pentane, and the mixture was filtered through a fluted filter paper into a glass-stoppered flask. This filtration removed dirt, chips, and water found in the pine oleoresin. To determine the total resin acid content of this solution 10 cc. were pipetted into 25 cc. of an alcohol-benzene mixture (500 cc. of alcohol, 500 cc. of benzene, and 0.2 gram of phenolphthalein). This solution was then titrated with 0.5 N alcoholic potassium hydroxide. VOLUMETRIC DETERMIXATION O F I-PIMARIC ACID. Twentyfive cubic centimeters of the above standardized n-pentane solution of pine oleoresin were placed in an iodine flask, and to it were added 20 cc. of standardized 10 per cent maleic anhydride in acetone solution. The flask was shaken and then allowed to stand a t 20" C. for 4 hours. At the end of this time the contents of the flask were transferred into a separatory funnel with the aid of 50 cc. of benzene. The maleic anhydride was then extracted as maleic acid with three 100-cc. portions of water. These aqueous extracts were united in a second separatory funnel and then extracted with 25 cc. of benzene. The aqueous layer was drawn off into the titration beaker and the benzene extract was washed by shaking with 15 cc. of additional water. This wash was also added to the titration beaker. The amount of maleic acid in the aqueous extracts was determined by titration with aqueous 0.5 N sodium hydroxide, using phenolphthalein as an indicator. The dif-
147
ference between the amount of maleic anhydride originally added and that accounted for in the aqueous extracts represents the amount of maleic anhydride reacted with I-pimaric acid. The per cent of I-pimaric acid present in the acidic portion of the pine oleoresin was calculated by the formula:
a X 3.082 X 100 b where a equals the weight in grams of maleic anhydride reacted with I-pimaric acid, and b equals the weight in grams of resin acids present in the 25 cc. of n-pentane solution used. GRAVIMETRIC DETERMINATION O F l-PIMARIC ACID. TWO hundred cubic centimeters of the standardized n-pentane solution of pine oleoresin were placed in a three-necked distilling flask fitted with a thermometer and a stirrer. The stirrer was connected into the stoppered reaction flask through a bearing which was not gas-tight. The n-pentane solution was stirred and a solution of 7 grams of maleic anhydride in 4 cc. of acetone was added rapidly. Stirring was continued for 2.5 hours. During the first half hour the temperature of the reaction mixture increased and cooling with water was necessary to hold the temperature at 30" C. At the end of the first hour crystallization had taken place and at the end of 2 hours the reaction was packed in ice and stirring was continued for a half hour longer. The crystalline material was then filtered into a Bdchner funnel using mild suction. A portion of the filtrate was used to make the transfer as quantitative as possible. Suction was then cut off entirely, 25 cc. of n-pentane were added to the filter cake, and the cake was puddled to ensure efficient washing. The n-pentane wash was drawn off slowly with very mild suction. The crystalline material was transferred to a weighed crystallizing dish and dried in vacuum at 100" C. for 2 hours and then weighed. The per cent of I-pimaric acid present in the acids of pine oleoresin was calculated by the formula:
% ' I-pimaric acid
=
yo I-pimaric acid
=
c X 0.755 X
100
d
where c equals the weight in grams of the dried addition product of I-pimaric acid and maleic anhydride and d equals the weight in grams of the resin acids present in the 200 cc. of n-pentane solution used. The amount of maleic anhydride added was adjusted so as to have approximately 1 gram excess a t the end of the reaction. Larger excess was avoided in order to keep to a minimum the amount of acetone added. A large excess also causes maleic anhydride to crystallize from the reaction mixture. DETERJIIXATION OF I-PIMARIC ACID ADDED TO PIXE OLEORESIX. A solution of 200 grams of longleaf pine oleoresin in 400 cc. of n-pentane was filtered as outlined above. This solution was titrated with alcoholic potassium hydroxide t o determine the concentration of resin acids present. A solution of 6 grams of maleic anhydride in 3 cc. of acetone vas then reacted with 200 cc. of the n-pentane solution which contained 44.10 grams of resin acids. The dried addition product rreighed 19.10 grams, which corresponds to 32.7 per cent of I-pimaric acid present in the acidic portion of the oleoresin. To a second 200-cc. portion of the standardized resin acid solution were added 2.20 grams of pure I-pimaric acid. A solution of 7 grams of maleic anhydride in 4 cc. of acetone was added as outlined above. The dried addition product weighed 22.20 grams, which corresponds t o 36.2 per cent of I-pimaric acid. Based on the control run and the amount of pure I-pimaric acid added, the calculated value should be 35.9 per cent.
Summary Determination of the l-pimaric acid content of longleaf pine oleoresin throughout the turpentining season showed a constant decrease in I-pimaric acid content with the progress of the season. The progressive change in I-pimaric acid content was followed by means of a volumetric and a gravimetric method based on the Diels-Alder reaction with maleic nnhydride.
Literature Cited (1) Bacon and Rusicka, Chemistry & Industry, 1936, 546 (2) Dupont, Bull. chim. we., ( 3 ) 29, 726 (1921). (3) Ibid., p. 734. (4) Dupont, Cornpt. rend., 178, 1560 (1924). (5) Sandermann, Be?., 71, 2005 (1938). (6) Sandermann, BUZZ. inst. p i n , (3) 31, 137 (1937). (7) Wienhaus and Sandermann, Ber., 69, 2202 (1936).
