Part 66

The process of analysis is conducted as follows: About three grams of the oil or fat are placed in a flask, with fifty cubic centimeters of alcoholic potash lye, containing enough potassium hydroxid to ensure complete saponification. The flask is closed and heated on a water-bath until saponification is complete. The pressure flask to be described hereafter may be conveniently used. After cooling, the excess of alkali is neutralized with acetic acid in presence of phenolphthalien and then alcoholic potash added until a faint pink color is produced. In a large porcelain dish place 200 cubic centimeters of water and thirty of a ten per cent solution of lead acetate and boil. Pour slowly, with constant stirring, into the boiling liquid the soap solution prepared as above described, and allow to cool, meanwhile continuing the stirring. At the end, the liquid remaining is poured off and the solid residue washed with hot water by decantation.

The precipitate of lead salts is finally removed from the dish into a stoppered bottle, the dish washed with pure ether, the washings added to the bottle together with enough ether to make the total volume thereof 120 cubic centimeters. The closed bottle is allowed to stand for twelve hours with occasional shaking, by which time the lead oleate will have been completely dissolved. The insoluble lead salts are next separated by filtration, and the filtrate collected in the olein tube. The washing is accomplished by ether and, to avoid loss, the funnel is covered with a glass plate. The ethereal solution of lead oleate is decomposed by dilute hydrochloric acid, using about forty cubic centimeters of a mixture containing one part of strong acid to four of water. The olein tube is closed and shaken until the decomposition is complete, which will be indicated by the clearing of the ethereal solution. The tube is allowed to remain at rest until the liquids separate and the aqueous solution is run out from the pinch-cock at the lower end. The residue is washed with water by shaking, the water drawn off as just described, and the process continued until all acidity is removed.

Water is then added until the separating plane between the two liquids is at the zero of the graduation, and enough ether added to make the ethereal solution of a desired volume, say 200 cubic centimeters. After well mixing, the ethereal solution or an aliquot part thereof, e.g., fifty cubic centimeters, is removed by the side tubulure and nearly the whole of the ether removed from the portion by distillation. To the residue are added fifty cubic centimeters of pure alcohol and the solution is titrated for oleic acid with decinormal sodium hydroxid solution. Each cubic centimeter of the hydroxid solution used is equivalent to 0.0282 gram of oleic acid. The total quantity of oleic acid contained in the amount of fat used is readily calculated from the data obtained.

To determine the iodin absorption of the free acid another measured quantity of the ethereal solution containing as nearly as possible half a gram of oleic acid, is withdrawn from the olein tube, and the ether removed in an atmosphere of pure carbon dioxid. To the residue, without allowing it to come in contact with the air, fifty cubic centimeters of Hübl’s reagent are added and the flask put aside in the dark for twelve hours. At the end of this time thirty-five cubic centimeters of a ten per cent solution of potassium iodid are added, the contents of the flask made up to a quarter of a liter with water, fifteen cubic centimeters of chloroform added, and the excess of iodin titrated in the way already described. The percentage of iodin absorbed is calculated as already indicated.

Lane has proposed a more rapid process for the above determination. The lead soaps are precipitated in a large erlenmeyer and cooled rapidly in water, giving the flask meanwhile a circular motion which causes the soaps to adhere to its walls. Wash with hot water, rinsing once with alcohol, add 120 cubic centimeters of ether, attach a reflux condenser, and boil until the lead oleate is dissolved, cool slowly, to allow any lead stearate which has passed into solution to separate, and filter into the olein tube. The rest of the operation is conducted as described above. The percentage of oleic acid and its iodin absorption in the following glycerids are given in the table below:

Cottonseed oil. Lard. Peanut oil. Per cent oleic acid 75.16 64.15 79.84 Per cent iodin absorbed 141.96 99.48 114.00

=340. Saponification.=—In many of the analytical operations which are conducted on the glycerids it is necessary to decompose them. When this is accomplished by the action of a base which displaces the glycerol from its combination with the fat acids, the resulting salts are known as soaps and the process is named saponification. In general use the term saponification is applied, not only strictly, as above defined, but also broadly, including the setting free of the glycerol either by the action of strong acids or by the application of superheated steam. In chemical processes the saponification of a glycerid is almost always accomplished by means of soda or potash lye. This may be in aqueous or alcoholic solution and the process is accomplished either hot or cold, in open vessels or under pressure. It is only important that the alkali and glycerid be brought into intimate contact. The rate of saponification is a function of the intimacy of contact, the nature of the solvent and the temperature. For chemical purposes, it is best that the decomposition of the glycerid be accomplished at a low temperature and for most samples this is secured by dissolving the alkali in alcohol.

In respect of solvents, that one would be most desirable, from theoretical considerations, which acts on both the glycerids and alkalies. In the next rank would be those which dissolve one or the other of the materials and are easily miscible, as, for instance, carbon tetrachlorid for the glycerid and alcohol for the alkali. As a rule, the glycerid is not brought into solution before the saponification process is commenced. Instead of using an alcoholic solution of sodium or potassium hydroxid the sodium or potassium alcoholate may be employed, made by dissolving metallic sodium or potassium in alcohol. It is probable, however, that a little water is always necessary to complete the process.

If a fat be dissolved in ether and treated with sodium alcoholate, a granular deposit of soap is soon formed and the saponification is completed in twenty-four hours. As much as 150 grams of fat can be saponified with ten grams of metallic sodium dissolved in 250 cubic centimeters of absolute alcohol. For practical purposes the alcoholic solution of the hydroxid is sufficient.

The chemical changes which fats undergo on saponification are of a simple kind. When the process is accomplished by means of alkalies, the alkaline base takes the place of the glycerol as indicated in the following equation:

Triolein 884. Potassium hydroxid 168. C₃H₅(O.C₁₈H₃₃O)₃ + 3KOH =

Potassium oleate 960. Glycerol 92. (KO.C₁₈H₃₃O)₃ + C₃H₅(OH)₃.

The actual changes which take place in ordinary saponification are not so simple, however, since natural glycerids are mixtures of several widely differing fats, each of which has its own rate of decomposition. Palmitin and stearin, for instance, are saponified more readily than olein and some of the saponifiable constituents of resins and waxes are extremely resistant to the action of alkalies. The above equation may be regarded as typical for saponification in aqueous or alcoholic solutions in open dishes or under pressure. If the alkali used be prepared by dissolving metallic sodium or potassium in absolute alcohol (sodium alcoholate or ethoxid) the reaction which takes place is probably represented by the equation given below:

C₃H₅(O.C₁₈H₃₃O)₃ + 3C₂H₅.ONa = C₃H₅(ONa)₃ + 3C₁₈H₃₃O.O.C₂H₅,

in which it is seen that complete saponification cannot occur without the absorption of some water, by which the sodium glyceroxid is converted into glycerol and sodium hydroxid, the latter compound eventually uniting with the ethyl ether of the fat acid.

Glycerids are decomposed when heated with water under a pressure of about sixteen atmospheres or when subjected to a current of superheated steam at 200°. The reaction consists in the addition of the elements of water, whereby the glyceryl radicle is converted into free glycerol and the fat acid is set free. The chemical change which ensues is shown below:

C₃H₅(O.C₁₈H₃₃O)₃ + 3H₂O = 3C₁₈H₃₄O₂ + C₃H₅(OH)₃.

The details of saponification with sulfuric acid are of no interest from an analytical point of view.

=341. Saponification in an Open Dish.=—The simplest method of saponifying fats is to treat them with the alkaline reagent in an open dish. In all cases the process is accelerated by the application of heat. Vigorous stirring also aids the process by securing a more intimate mixture of the ingredients. This method of decomposing glycerids, however, is not applicable in cases where volatile ethers may be developed. These ethers may escape saponification and thus prevent the formation of the maximum quantity of soap. While not suited to exact quantitive work, the method is convenient in the preparation of fat acids which are to be the basis of subsequent analytical operations, as, for instance, in the preparation of fat acids for testing with silver nitrate. Large porcelain dishes are conveniently used and the heat is applied in any usual way, with care to avoid scorching the fat.

=342. Saponification under Pressure.=—The method of saponification which has given the best satisfaction in my work and which has been adopted by the Association of Official Agricultural Chemists is described below.

Reagents.—The reagents employed are a solution of pure potash containing 100 grams of the hydroxid dissolved in fifty-eight grams of recently boiled distilled water, alcohol of approximately ninety-five per cent strength redistilled over caustic soda, and sodium hydroxid solution prepared as follows:

One hundred grams of sodium hydroxid are dissolved in 100 cubic centimeters of distilled water. The caustic soda should be as free as possible from carbonates, and be preserved from contact with the air.

Apparatus.—A saponification flask; it has a round bottom and a ring near the top, by means of which the stopper can be tied down. The flask is arranged for heating as shown in Fig. 101. A pipette graduated to deliver forty cubic centimeters is recommended as being more convenient than a burette for measuring the solutions: A pipette with a long stem graduated to deliver 5.75 cubic centimeters at 50°.

Manipulation.—The fat to be examined should be melted and kept in a dry warm place at about 60° for two or three hours, until the water has entirely separated. The clear supernatant fat is poured off and filtered through a dry filter paper in a jacket funnel containing boiling water. Should the filtered fat, in a fused state, not be perfectly clear, it must be filtered a second time. The final drying is accomplished at 100° in a thin layer in a flat bottom dish, in partial vacuum or an atmosphere of inert gas.

The saponification flasks are prepared by thoroughly washing with water, alcohol, and ether, wiping perfectly dry on the outside, and heating for one hour at the temperature of boiling water. The hard flasks used in moist combustions with sulfuric acid for the determination of nitrogen are well suited for this work. The flasks should be placed in a tray by the side of the balance and covered with a silk handkerchief until they are perfectly cool. They must not be wiped with a silk handkerchief within fifteen or twenty minutes of the time they are weighed or else the electricity developed will interfere with weighing. The weight of the flasks having been accurately determined, they are charged with the melted fat in the following way:

The pipette with a long stem, marked to deliver 5.75 cubic centimeters, is warmed to a temperature of about 50°. The fat, having been poured back and forth once or twice into a dry beaker in order to thoroughly mix it, is taken up in the pipette, the nozzle of the pipette having been previously wiped to remove any externally adhering fat, is carried to near the bottom of the flask and 5.75 cubic centimeters of fat allowed to flow into the flask. After the flasks have been charged in this way they should be re-covered with the silk handkerchief and allowed to stand for fifteen or twenty minutes, when they are again weighed.

=343. Methods of Saponification.=—In the Presence of Alcohol.—Ten cubic centimeters of ninety-five per cent alcohol are added to the fat in the flask, and then two cubic centimeters of the sodium hydroxid solution. A soft cork stopper is inserted and tied down with a piece of twine. The saponification is completed by placing the flask upon the water or steam-bath. The flask during the saponification, which should last one hour, should be gently rotated from time to time, being careful not to project the soap for any distance up its sides. At the end of an hour the flask, after having been cooled to near the room temperature, is opened.

Without the Use of Alcohol.—To avoid the danger of loss from the formation of ethers, and the trouble of removing the alcohol after saponification, the fat may be saponified with a solution of caustic potash in a closed flask without using alcohol. The operation is carried on exactly as indicated above for saponification in the presence of alcohol, using potassium instead of sodium hydroxid solution. For the saponification, use two cubic centimeters of the potassium hydroxid solution which are poured on the fat after it has solidified in the flask. Great care must be taken that none of the fat be allowed to rise on the sides of the saponifying flask to a point where it cannot be reached by the alkali. During the process of saponification the flask can only be very gently rotated in order to avoid the difficulty mentioned. This process is not recommended with any apparatus except a closed flask with round bottom. Potash is used instead of soda so as to form a softer soap and thus allow a more perfect saponification.