For analytical purposes, the extraction of lecithin from vegetable substances is conducted in this laboratory as follows:[362] The fine-ground pea or bean meal is placed in an extraction apparatus and treated continuously with anhydrous ether for fifteen hours. The ether in the apparatus is replaced with absolute alcohol and the extraction continued for six hours longer. The alcoholic extract is evaporated to dryness and treated with ether. The part of the lecithin at first insoluble in ether becomes soluble therein after it has been removed from the vegetable tissues by alcohol. Moreover, any trace of inorganic phosphorus which may have been removed by the alcohol, is left undissolved on subsequent treatment with ether. The ether extract from the alcohol residue is added to that obtained directly, the ether removed by evaporation, and the total lecithin oxidized and the residue used for the estimation of phosphorus as already described.
In determining the lecithin in eggs, the procedure employed for vegetable tissues is slightly changed.[363] The whole egg, excluding the shell, is placed in a flask with a reflux condenser and boiled for six hours with absolute alcohol. The alcohol is then removed from the flask by evaporation and the residue treated in like manner with ether for ten hours. The ether is removed and the dry residue rubbed to a fine powder, placed in an extractor and treated with pure ether for ten hours. The extract thus secured is oxidized after the removal of the ether by fusion with mixed alkaline carbonates and the phosphorus determined in the usual way.
390. Factor for Calculating Results.—The percentage of lecithin is calculated from the weight of magnesium pyrophosphate obtained by multiplying it by the factor, 7.2703.[364] This factor is calculated from the second formula for lecithin given above, in which the percentage of phosphorus pentoxid, P₂O₅, is 8.789.
Example.—In fifty-four grams of egg, exclusive of the shell, is found an amount of organic phosphorus yielding 0.0848 gram of magnesium pyrophosphate. Then 0.0848 × 7.2703 = 0.61652 and 0.61652 × 100 ÷ 54 = 1.14. Therefore the percentage of lecithin in the egg is 1.14.
391. Estimation of Alkaloidal Nitrogen.—The alkaloids contain nitrogen in a form more difficult of oxidation than that contained in proteid or albuminoid forms. It is doubtful whether any of the nitrogen in alkaloids becomes available for plant nutrition by any of the usual processes of fermentation and decay to which nitrogenous bodies are submitted in the soil. Likewise, it is true that it is not attacked by the digestive processes in any way preparatory to its assimilation as food by the animal tissues. Alkaloidal nitrogen is therefore not to be regarded as a food either for the animal or plant.
For the general methods of estimating alkaloids the reader is referred to standard works on plant chemistry and toxicology. The alkaloids of interest in this manual are those which are found in tobacco, tea, coffee and a few other products of agricultural importance. The best methods of isolating and estimating these bodies will be given in the part of the volume devoted to the special consideration of the articles mentioned.
SEPARATION OF PROTEID BODIES
IN VEGETABLE PRODUCTS.
392. Preliminary Treatment.—The chief disturbing components of vegetable tissues, in respect of their influence on the separation and estimation of the proteid constituents, are fats and oils and coloring matters. In many cases these bodies are present in such small quantities as to be negligible, as, for instance, in rice. In other cases they exist in such large proportions as to present almost insuperable difficulties to analytical operations, as is the case with oily seeds. In all instances, however, it is best to remove these bodies, even when present in small proportions, provided it can be done without altering the character of the proteid bodies. This is secured by extracting the fine-ground vegetable material first with petroleum ether, and afterwards with strong alcohol and ether. Practically, all of the fatty bodies and the greater part of the most objectionable coloring matters are removed by this treatment. The extraction should in all cases be made at low temperatures, not exceeding 35°, to avoid the coagulating effect of higher temperatures upon the albuminous bodies which may be present.
In this laboratory, fatty seeds, as for instance peanuts, are first ground into coarse meal, then extracted with petroleum ether, ground to a fine meal and the fat extraction completed with petroleum ether, ninety-five per cent alcohol and pure sulfuric ether. The residue of the last solvent may be removed by aspirating air through the extracted meal. In some cases, it is advisable to extract with ethyl ether before as well as after the alcoholic extraction. This treatment removes at least a part of the water and prevents the dilution of the first part of alcohol added to such an extent as to make it dissolve some of the proteid matters. In each case, a portion of the alcoholic extract should be tested qualitively for proteid matter. If any be found, stronger alcohol should be used for, at least, the first extraction. A portion of the meal, prepared as above directed, is extracted with a ten per cent solution of sodium chlorid, as described further on, and a measured portion of the filtered extract diluted with water until the proteid matter in solution begins to be precipitated. By this treatment the proper strength of the salt solution, to be used for the subsequent extraction, is determined. To save time in dialyzing, the solution of salt employed as a solvent should be as dilute as possible.
The mixture of meal and solvent sometimes filters with difficulty. In these cases, it is advisable to first pour it into a linen bag from which the liquid portion can be removed by gentle pressure and subsequently filtered through paper. As a last resort, the liquid secured from the linen filter can be saturated with ammonium, zinc or magnesium sulfate, whereby all the proteid matters are thrown out. After filtering, the residue is again dissolved in salt solution and can then be readily filtered through paper.