In addition to fractional distillation, several methods for the assay of cineol have been recommended. They all depend on the capacity of cineol to form addition products with certain other compounds. The description of the individual method is here given.

1. Distillation method.1) The oil to be examined is fractionated, fractions of two degrees each being collected, placed in a freezing mixture and their temperature reduced to - 15 to - 18°. Attempts are then made to cause them to congeal by shaking or by introducing a cineol crystal. After having stood in the freezing mixture for an hour, the liquid portion is drawn off by means of a pipette drawn out to a fine point. With some experience a well nigh dry mass of crystals can be obtained, from which the last traces of liquid can be removed by shaking the crystals. The liquified cineol of all fractions is united and weighed.

In as much as a portion of the cineol remains dissolved in the terpenes, all of the cineol is not obtained by this method. Hence the method is applicable only then when it is desired to ascertain which of a number of oils is richest in cineol.

2. Hydrogen bromide method. Into a solution of 10 cc. of oil in 40 cc. of low boiling petroleum ether (b. p. 35 to 40°), thoroughly chilled in a freezing mixture, absolutely dry hydrogen bromide is passed so long as a precipitate continues to be formed. The purely white cineol hydrobromide (C10H18OHBr) is quickly separated with the aid of a suction pump and washed with cold petroleum ether. Into the filtrate more hydrogen bromide is passed and, if additional precipitate results, this is collected separately and combined with the bulk of precipitate first obtained and separated.

1) Helbing's Pharmacological Record No. VIII. London 1892.

In order to remove the petroleum ether, the cineol hydro-bromide is left in a vacuum desiccator for a quarter of an hour. It is then transferred to a cassia flask with the aid of a little alcohol and decomposed with water. By the addition of more water the cineol is raised into the neck of the flask and its volume read off by means of the graduated scale. Multiplication with 10 yields the percentage of cineol by volume in the oil.

3. Phosphoric acid method,1) adopted by the U. S. Pharmacopoeia, 8th decennial revision. To a solution of 10 cc. of oil in 50 cc. petroleum ether, which is thoroughly chilled in a freezing mixture, concentrated phosphoric acid is gradually added while stirring until the white precipitate (C10H18OH3PO4) attains a yellowish or reddish tint. The crystalline mass is removed with the aid of a force filter, washed with petroleum ether, dried by pressing between porous plates, and decomposed with water. The regenerated cineol is measured and the volume percentage computed.

Both the hydrobromic acid and phosphoric acid methods suffer form the drawback that their addition products with cineol are readily decomposible thus rendering difficult a quantitative separation. In consequence the results, as shown by Schimmel & Co.2) in connection with trial assays of mixtures of known cineol content, are unreliable. In part they deviate very materially from the true composition. Nevertheless, the hydrogen bromide method may prove useful in cases where the cineol content is low since then all other methods fail.

4. Resorcinol method. Recently a method has been worked out in the laboratory of Schimmel & Co.3) which depends on the property of cineol to form an addition product with resorcinol which is soluble in an excess of concentrated resorcinol solution. Method of procedure: To 10 cc. of oil contained in a 100 cc. cassia flask (fig. 75, p. 583) enough 50 p. c. resorcinol solution is added to fill the flask about four-fifths. For five minutes the mixture is thoroughly shaken, then that portion of the oil which has not gone into solution is driven into the neck with resorcinol solution. Any oily particles adhering to the walls of the flask are caused to rise to the surface by rotating the flask or gently tapping it. After the resorcinol solution has become perfectly clear, which usually requires several hours, the volume of oil remaining is read off, the cineol content ascertained by subtracting this amount from 10 and the resultant multiplied with 10 in order to obtain the percentage by volume. Oils very rich in cineol are advantageously diluted with an equal volume of turpentine oil since the cineolresorcinol occasionally crystallizes from concentrated solutions thus rendering futile the entire process.

1) The method was first suggested by Helbing and Passmore who, however, employed no diluent. Helbing's Pharmacological Record No. XXIV. London 1893. Kebler attempted to introduce an improvement by titrating the phosphoric acid set free from the regenerated cineol after the cineol phosphate had been expressed. Americ. Journ. Pharm. 70 (1898), 492. If one, however, recalls how difficult it is to remove the sirupy phosphoric acid from the tough cineol phosphate cake, this change is not likely to enjoy the confidence of the analyst.

2) Report of Schimmel & Co. October 1907, 47.

3) Report of Schimmel & Co. October 1907, 47; Wiegand and Lehmann, Chem. Ztg. 32 (1908), 109; Report of Schimmel & Co. April 1908, 52.

The method as described above is to be applied only to such oils that contain no appreciable amounts of oxygenated compounds (alcohols, aldehydes) since resorcinol dissolves these also, thus causing the results to be too high. In all other cases the assay is carried out, not with the original oil but with the cineol fraction. For this purpose 100 cc. of oil are distilled from a Ladenburg fractionating flask with three bulbs (fig. 71, p. 566) at such a rate that a drop is collected each second. Fraction 170 to 190°, which contains all of the cineol of the oil, is collected separately and its volume ascertained. The cineol content of this fraction is then determined as described above and the percentage of cineol in the original oil computed. In the examination of mixtures of known cineol content, Wiegand and Lehmann have shown that the error is at most 2°/o.

If only small amounts of oil are available the method can be modified so as to separate the solid resorcinol compound and to decompose the latter. With some practice acceptable results can thus also be obtained though not with the same certainty as with the fractionation method just described. Nevertheless, it is preferable to the phosphoric acid addition method since the additionproduct of cineol with resorcinol is more stable than that with phosphoric acid.

10 cc. of oil are mixed with 20 cc. of a 50 p. c. resorcinol solution. If necessary, a crystal of cineol resorcinol is added and the resulting crystalline mass is stirred to form a uniform mixture. The crystalline mass is separated by means of a force filter, pressed between filter paper to remove the last traces of oil, and transferred to a beaker. Caustic alkali is then added, the mixture gently heated and the liquid transferred to a cassia flask by means of a funnel the tube of which passes to the bottom of the flask. The flask is finally filled to the mark, the amount of separated cineol read off and multiplied by 10 to obtain percentage by volume.

For the regeneration of the resorcinol used for these determinations steam is passed through the solution from which the non-cineol portions of the oil has been previously separated. The cineol distills over and the aqueous solution is evaporated for the recovery of the resorcinol which can be used again for another assay.