144
INDUSTRIAL AND ENGINEERING CHEMISTRY
the data of Tables I1 and 111) as 10.9 X 10-4. The fact that this deviation is smaller than the above-mentioned deviation of *1.4 X lo-‘ from the pycnometer value, indicates that successive replicates agree better than replicates carried out over a longer period. Errors may be caused if both sides of the aPParatus are exposed to different temperatures as by the action of drafts of air, and the poorer precision may perhaps be explained by the introduction of such variableg over a period of time. ACKNOWLEDGMENT. Grateful acknowledgment is made to E. I. Stearns for his assistance in the preparation of the dim^+ sion of accuracy and precision.
Vol. 18, No. 2
LITERATURE CITED
(1) Ciochina, J., 2.anal. Chem., 107, 108 (1936). (2) Davidson, D., and Popowsky, M., Paper 33, Division of Analytical and Micro Chemistry, 102nd Meeting of the AMERICAN CHEMICAL SOCIETY, Atlantic City, N. J., September, 1941. (3) Davidson, D., Popowsky, M., and Rosenblatt, P., U. S. Patent 2,328,787 (Sept. 7, 1944). (4) Fisher, R. A., “Statistical Methods for Research Workers”, 6th ed., p. 48,London, Oliver and Boyd, 1936. (5) Frivold, 0. E., Phys. Z., 21, 529 (1920): 24,86 (1923). pREsENTED before the Meeting-in-Miniature of the Xew York Section of t h e AMERICAN CHEMICAL SOCIETY, June 8. 1945.
Microdetermination of the Saponification Number of Fats and
Oils
Decigram, Centigram, and Milligram Procedures KALMAN MARCALI AND WILLIAM RIEMAN 111 N. J.
School of Chemistry, Rutgers University, N e w Brunswick,
T
HE literature contains no satisfactory
method for the microdetermination of the saponification number of fats and oils. A semimicromethod was devised by Chargaff (8) but was applied t o only three oils. It has been pointed out that the double-indicator method (9)is more easily adaptable to microprocedures than the standard method (1).
Hydrochloric acid, approximately N , was standardized with pure sodium tetraborate decahydrate and used in all titrations. The acid used for the centigram procedure was standardized by weighing accurately 400 to 600 mg. of a carefully prepared, approximately 0.5 weight-normal solution of borax into a 13 X 100 mm. Pyrex test tube, adding one drop of 0.001 M methyl red, and 0.5
Methods are described for determination of the saponification number of fats and oils with samples of about 500,50, and 5 mg.
APPARATUS
For the decigram procedure, a 10-ml. calibrated buret with 0.05-ml. graduations was used. The tip was constricted so that drops of about 0.01 ml. were delivered. The buret for the centigram procedure is illustrated in Figure 1. A ground-glass rod, A , 5.2 mm. in diameter, fits snugly inside a ground-glass tube, B. The upper end of B is sealed to capillary, C , which is bent twice a t right angles and constricted a t the tip as indicated. A 25-mm. micrometer, D, the head of which has been removed, serves to measure accurately the position of the piston A . Above the piston is a layer of mercury, E , which makes a leakproof seal. Above the mercury is the 0.5 N hydrochloric acid. Castalloy clamps F hold the micrometer and glass tube rigidly in fixed positions. The titration vessel, G , is simply a 13 X 100 mm. Pyrex test tube. A movable arm, H,taken from a Rehberg buret made by Microchemical Service supports the titration vessel. A finely drawn capillary, I, admits a stream of air, free from carbon dioxide, into the titration vessel. The assembly for the titration in the milligram procedure was essentially the same except that a smaller test tube (10 X 75 mm.) and ground-glass rod and tube (3.1 mm.) were used. I n order to refill these micrometer burets the tip is wiped with Kleenex tissue and immersed in the 0 . 5 h hydrochloric acid. The micrometer spindle is turned down, and the piston is slowly pushed down with the fingers tillit makes contact with the spindle. A 4 X 3 X 2 mm. platinum boat was made from foil 0.06 mm. thick. A fluorescent titration illuminator was used for all titrations. REAGENTS
The alcoholic potassium hydroxide was prepared as previously described (5). Alcoholic 0.001 M bromophenol blue was prepared by dissolving 65 mg. of the indicator in 1.0 ml. of 0.10 N sodium hydroxide and adding a mixture of 60 ml. of purified 95% alcohol and 40 ml. of benzene. The alcohol-benzene mixture consisted of 60 ml. of reagentgrade benzene and 40 ml. of purified 95% alcohol.
results were expressed as milliequivalents of acid per millimeter. The acid used for the milligram procedure was standardized analogously. About 10 mg. of pure borax were weighed on a microchemical balance and transferred to a 10 X 75 mm. Pyrex test tube. Four drops of water and one drop of 0.00008 M methyl red were added before the titration. PROCEDURES
DECIGRAM PROCEDURE. Weigh accurately about 500 mg. of sample into a 50-ml. Pyrex Erlenmeyer flask. Add about 5 ml. of the alcoholic potassihm hydroxide, and immediately connect a reflux condenser, the upper end of which is protected by an Ascarite tube. Boil gently for 30 minutes. Add 2 drops of 1% alcoholic phenolphthalein, and titrate with 0.5 N hydrochloric acid to the disappearance of the pink color. Add 3 drops (0.18 ml.) of aqueous 0.001 M bromophenol blue and 1 ml. of benzene. Continue the titration to a green color that does not turn blue on further agitation. CENTIGRAM PROCEDURE. Weigh with an accuracy of 0.02 mg. about 50 mg. of sample into a tared Pyrex test tube, add about 21 drops (0.5ml.) of alcoholic otassium hydroxide, and immediately attach an Ascarite tube. ifupport the tube vertically, so that the bottom rests on t k flat surface of a micro drying block which is kept a t 145’ * 5 . The test tube serves as both reaction vessel and reflux condenser. The saponification is complete in 30 minutes. Add 1 drop (0.03,ml.) of 0.2% alcoholic phenolphthalein. Insert the tip of the micrometer buret (previously wiped with Kleenex tissue) and the tip of the air-delivery tube, and supp0l.t the tube in the titration apparatus (Figure 1). Adjust the air stream to a rate of about 80 bubbles per minute. Add the 0.5 N hydrochloric acid by carefully turning the micrometer spindle until the color just disappears. Read themicrometer a t this point. Without removing either capillary from the test tube, add 1 drop (0.06 ml.) of aqueous 0.001 M bromophenol blue and 7 drops (0.12 ml.) of benzene. Decrease the air stream to about 50 bubbles per minute, and continue the titration till the solution has a milky appearance; then add 7 more drops of benzene, and increase the air stream to about 100 bubbles pertminute. Continue the titration to a green end point. The hydrochloric acid
ANALYTICAL EDITION
February, 1946
used between the two end points is equivalent to the potassium hydroxide that was required for the saponification. Weigh the platinum boat (4 X 3 X MILLIQRAM PROCEDURE. 2 mm.) on a microchemical balance, then place it on a clean surface. Dip the tip of a finely drawn glass rod into the sample, let most of the oil drain off the rod, and touch the center of the boat with the tip of the rod. Transfer the boat to the balance pan, and weigh i t again. The sample should be between 3 and 8 mg. With platinum-tipped forceps, place the boat just inside the open end of a horizontally held 10 X 75 mm. Pyrex test tube. Then tilt the test tube so that the boat slides t o the bottom of the tube. If any oil sticks to the side of the tube, the determination must be started anew. Let 2 drops of the alcoholic potassium hydroxide fall directly from the siphon of the storage bottle into the bottom of this test tube. Immediately connect an Ascarite tube t o the test tube. Then support it with a clamp and ring stand so that the bottom of t h e tube r;sts on the flat surface of a micro drying block heated to 130 * 5
145
holic potassium hydroxide into the Pyrex test tube and titrate it as described above, noting the initial micrometer reading, x, the reading a t the phenolphthalein end point, y, and the reading at the end point of bromophenol blue, z. Then calculate the saponification number with the equation
( u - ~)]56.11F s = [ ( b - C) - B W where S = saponification number
B -
y--z
2-Y F = concentration of hydrochloric acid, milliequivalentu per mm. W = weight of sample, grams
.
RESULTS
The saponification numbers of eleven oils were determined by the standard method ( I ) and by each of the methods described in this paper. Each entry in Table I denotes a single determination except as indicated in the footnotes. No values were omitted except those obtained before the methods had been perfected. DISCUSSION
! Illi
r H
Excellent results are obtained by the decigram and centigram methods, while the milligram method yields very mtisfactory results. In order t o study the precision of the methods, one oil was run several times by the same method. The mean deviation6 were 0.3, 0.1, and 0.5 unit by the decigram, centigram, and milligram methods, respectively.
-G
Table
W
0
Comparison of Results by Various Methods Saponification
Saponification
NO.
NO.
Saponification
No.
Stand- DeciCentiMilliard gram Differ- gram Differ- gram DifferOil method method ence method ence method ence 181.5 181.5 0.0 182.0 +0.5 182.4 +0.9 Castor oil 194.6 194.5 -0.1 195.0 +0.4 195.4 4-0.8 Cocoa butter Coconut oil 258.6 258.9 +0.3 258.5 -0.1 259.2 +0.6 Cod liver oil 188.0 188.6 1 0 . 6 187.9 -0.1 188.2' +0.2 0 0 191.6' 0.0 192.1 + 0 . 5 Corn oil 191.6 191.6b +O.l 195,7 +0.2 196.2 1-0.7 Lard oil 195.5 195.6 187.0 187.1 +0.1 187.4 +0.4 187.4 + 0 . 4 Linseed oil 186.7 186,7 0 0 186.4 -0.3 187.4 f0.7 Neat's-foot oil 191.5 191.8 + 0 . 3 192.0 $0.5 190.7 -0.8 Olive oil 174.3 174.0 -0.3 174.6 +0.3 175.2 +0.9 Rapeseed oil 194.7 194,5 -0.2 194,4 -0.3 195.0 +0.3 Tung oil Mean (signs disregarded) *o 2 *0.3 *O.d a
b
Figure 1.
1.
Mean of 4 determinations. Mean of 10 determinations.
Micrometer Buret
After 30 minutes, remove the Ascarite tube and add 2 drops (0.06 ml.) of 0.02yo alcoholic phenolphthalein. Immediately insert the tip (previously wiped with Kleenex tissue) of the micrometer buret and the tip of the air-delivery tube, and support the tube in the titration apparatus (Figure 1). Take care that the boat does not interfere with the delivery of acid from the buret. Adjust the air stream to about 60 bubbles per minute. Kote the reading, a, of the micrometer before any hydrochloric acid is added to the titration vessel. Now titrate to the disappearance of the pink color of the phenolphthalein. Rinse the inside of the test tube with 0.10 to 0.20 ml. of the alcohol-benzene mixture by means of a measuring pipet. If the pink color reappears, add more hydrochloric acid till it disappears again. Note this reading, b, of the micrometer. Add 2 drops (0.06 ml.) of the alcoholic bromophenol blue. Continue the titration. Just before the end point is reached, rinse the inside of the test tube again with 0.10 t o 0.20 ml. of the alcohol-benzene mixture. Note the micrometer reading, c , when the green end point is reached. Contamination of the alcoholic potassium hydroxide by carbon dioxide of the air could not be avoided in the milligram procedure, although no difficulty was encountered in this respect in the other procedures. Therefore a blank correction must be applied in the milligram procedure. To run the blank, put 2 drops of the alco-
The decigram method is more convenient than the gram method chiefly because the smaller solution cools almost instantaneously after saponification and reduces the waiting period. The centigram procedure is probably a little more troublesome because of the special buret. The milligram procedure is still more troublesome and is to be chosen only when the quantity of sample is very limited. None of these methods is applicable to acetylated oils (9). ACKNOWLEDGMENT
The authors are grateful to the Research Council of Rutgers University for financial aid in this investigation and to Evan Teunon for suggestions regarding the micrometer buret. LITERATURE CITED
(1) Assoc. Official Agr. Chem., Official and Tentative Methods of A4nalysis,5th ed., p. 433, 1940. (2) Chargaff, E., 2. physiol. Chem., 199,221 (1931). (3) Rieman, William, 111, IND.ENQ.CHEM.,ANAL.ED., 15, 326 (1943).
