Definite volumes of the aqueous solution of the caffein are acidulated with sulfuric and the alkaloid precipitated by an excess of a set solution of iodin in potassium iodid. After filtering, the excess of iodin in an aliquot part of the filtrate is determined by titration with a tenth normal solution of sodium thiosulfate. The filtration of the iodin liquor is accomplished on glass wool or asbestos. The results of the analyses are calculated from the composition of the precipitated caffein periodid; viz., C₈H₁₀N₄O₂.HI.I₄. The weight of the alkaloid is calculated from the amount of iodin required for the precipitation by the equation 4I: C₈H₁₀N₄O₂ = 508: 194. From this equation it is shown that one part of iodin is equivalent to 0.3819 part of caffein, or one cubic centimeter of tenth normal iodin solution is equal to 0.00485 gram of iodin.
In practice, it is recommended to divide the aqueous extract of the alkaloid, prepared as directed above, into two portions, one of which is treated with the iodin reagent without further preparation, and the other after acidulation with sulfuric. After ten minutes, the residual iodin is estimated in each of the solutions as indicated above. The one portion, containing only the acetic acid resulting from the decomposition of the lead acetate, serves to indicate whether the aqueous solution of the caffein contains other bodies than that alkaloid capable of forming a precipitate with the reagent, since the caffein itself is not precipitated even in presence of strong acetic acid.
In the solution acidulated with sulfuric, the caffein, together with the other bodies capable of combining with iodin, is precipitated. The residual iodin is determined in each case, and thus the quantity which is united with the caffein is easily ascertained. The weight of iodin which has entered into the precipitated caffein periodid multiplied by 0.3819 gives the weight of the caffein in the solution.
Gomberg’s method has been subjected to a careful comparative study by Spencer and has been much improved by him in important particulars.[599]
It is especially necessary to secure the complete expulsion of the hydrogen sulfid and to observe certain precautions in the addition of the iodin reagent. The precipitation should be made in a glass-stoppered flask, shaking thoroughly after the addition of the iodin and collecting the precipitate on a gooch. As thus modified, the iodin process gives results comparable with those obtained by Spencer’s method, and it can also be used to advantage in estimating caffein in headache tablets in the presence of acetanilid.
582. Freeing Caffein of Chlorophyll.—Any chlorophyll which may pass into solution and be found in the caffein may be removed by dissolving the caffein in ten per cent sulfuric acid, filtering, neutralizing with ammonia and evaporating to dryness. The residue is taken up with chloroform, the chloroform removed at a low temperature and the pure caffein thus obtained.[600]
583. Proteid Nitrogen.—The proteid nitrogen in tea and coffee may be determined in the residue after extraction of the alkaloid by boiling water as described above. More easily it is secured by determining the total nitrogen in the sample and deducting therefrom the nitrogen present as caffein. The remainder, multiplied by 6.25, will give the quantity of proteid matter.
584. Carbohydrates of the Coffee Bean.—The carbohydrates of the coffee bean include those common to vegetable substances; viz., cellulose, pentosan bodies (xylan, araban), fiber etc., together with certain sugars, of which sucrose is pointed out by Ewell as the chief.[601] In smaller quantities are found a galactose yielding body (galactan), as pointed out by Maxwell, a dextrinoid and a trace of a sugar reducing alkaline copper solution.
The sucrose may be separated from the coffee bean by the following process:[602] The finely ground flour is extracted with seventy per cent alcohol, the extract clarified with lead acetate, filtered, the lead removed from the filtrate with hydrogen sulfid, the excess of the gas removed by boiling, the filtrate evaporated in a partial vacuum to a sirup and the sucrose crystallized from a solution of the sirup in alcohol.
For a quantitive determination, ten grams of the coffee flour are extracted with ether and the residue with seventy-five per cent alcohol. This process, conducted in a continuous extraction apparatus, should be continued for at least twenty-four hours. The alcohol is removed by evaporation, the residue dissolved in water, clarified with basic lead acetate, filtered, the precipitate washed, the lead removed, again filtered, the filtrate washed and wash water and filtrate made to a definite volume. In an aliquot part of this solution the sugars are determined by the alkaline copper method, both before and after inversion. From the data obtained the percentage of sucrose is calculated.