Therefore, a volume of nitrogen which occupies a space of thirty-five cubic centimeters at a temperature of 22°, and at a barometric pressure of 740 millimeters, becomes 31.33 cubic centimeters at a temperature of 0° and a pressure of 760 millimeters.

One liter of nitrogen at 0° and 760 millimeters pressure weighs 1.25456 grams; and one cubic centimeter therefore 0.00125456 gram. To find the weight of gas obtained in the above supposed analysis, it will only be necessary to multiply this number by the volume of nitrogen expressed in cubic centimeters under the standard conditions; viz., 0.0125456 × 31.33 = 0.039305 gram. If the sample taken for analysis weighed half a gram, the percentage of nitrogen found would be 7.85.

164. Tension of the Aqueous Vapor.—It has been shown by experience that when a gas is collected over a potash solution containing fifty per cent of potassium hydroxid, the tension of the aqueous vapor is so far diminished as to be of no perceptible influence on the final result. To correct the volume of a gas, therefore, so collected for this tension, would involve an unnecessary calculation for practical purposes. If a gas thus collected should be transferred to a burette over mercury, on which some water floats, then the correction should be made.

At 0° the tension of aqueous vapor will support a column of mercury 4.525 millimeters high, and at 40° one of 54.969 millimeters.

The following table gives the tension of aqueous vapors in millimeters of a mercurial column for each degree of temperature from zero to forty.

When a gas is in contact with water the aqueous vapor is diffused throughout the mass, and the pressure to which the mixture is subjected, is partly neutralized by the tension of the water vapor. The real pressure to which the gas, whose volume is to be determined is subjected, is therefore diminished by that tension. If for instance a gas in contact with water show a volume of thirty-five cubic centimeters at 22° and 740 millimeters barometric pressure its volume is really greater than if it were perfectly dry. How much greater can be determined by inspecting the table, for at 22° the tension of water vapor is 19.675 millimeters of mercury. The real pressure to which the volume of gas is subjected is therefore 740 - 19.675 = 720.325 millimeters.

If, therefore, in the example given, the nitrogen were in contact with water, the calculation would proceed as follows:

and 30.5 × 1.25456 = 38.26.

Hence, 38.26 milligrams of nitrogen correspond to 7.65 per cent, when half a gram of substance is taken for the combustion.

165. Aqueous Tension in Solutions of Potassium Hydroxid.—Even in strong solutions of potassium hydroxid the tension of aqueous vapor is not destroyed, but is reduced to a minimum, which is negligible in the calculation of the percentage by weight of the nitrogen in a sample of fertilizer. When dilute solutions of a caustic alkali are used however, the neglect of the tension of the aqueous vapor may cause an error of some magnitude. In such cases the strength of the solution should be known and correction made according to the following table:[137]

166. Use of Volumetric Method.—For practical purposes it may be said that the volumetric determination of nitrogen in fertilizer analysis has gone entirely out of use. For control and comparison it is still occasionally practiced but it has had to give way to the more speedy and fully as accurate processes of moist combustion with sulfuric acid which have come into general use in the last decade. The student and analyst however should not fail to master its details and become skilled in its use. There are certain nitrogenous substances such as the alkaloids which are quite refractory when subjected to moist combustion. While such bodies may not occur in fertilizers it is well to have at hand a means of accurately determining their nitrogen content.

167. Tables for Calculating Results.—Where many analyses are to be made by the copper oxid process it has proved convenient to shorten the work of calculating analyses by taking the data given in computation tables.[138] Before using these tables it must be known whether they are calculated on the supposition that the gas is measured in a moist state, partly moist, or wholly dry. Where the nitrogen is collected over water a table must be used in which allowance has been made for the tension of aqueous vapor. In case a saturated solution of a caustic alkali be used in the azotometer it is customary to take no account of the tension and the table employed must be constructed on this supposition. In point of fact even in the strongest alkali solution there is a certain amount of tension but this is so slight as only to affect the results in the second place of percentage decimals. Since, as a rule, only a few analyses are made by this method it will be found safer to use a caustic alkali solution of given strength and to calculate the results from the tables of aqueous tensions given above.

168. The Soda-Lime Process.—This process originally perfected by Varrentrap and Will, and improved by Peligot, was used almost exclusively by analysts until within the last decade for the determination of nitrogen not existing in the nitric form. It is based on the principle that when nitrogen exists as a salt of ammonia, or as an amid, or as proteid matter, it is converted into gaseous ammonia by combustion with an alkali. This ammonia can be carried into a set solution of acid by a stream of gas free of ammonia and the excess of acid remaining after the combustion is complete can be determined by titration against a standard alkali solution. The results under proper conditions are accurate even when a small quantity of nitric nitrogen is present. When, however, there is any considerable quantity of this compound in the sample the method becomes inapplicable by reason of non-reduction of some of the nitrogen oxids produced by the combustion.

In bodies very rich in nitrogen such as urea all the nitrogen is not transformed directly into ammonia at the commencement of the combustion. A portion of it may unite with a part of the carbon to form cyanogen, which may unite with the soda to form sodium cyanid. With an excess of alkali, however, and prolonged combustion this product will be finally decomposed and all the nitrogen be secured as ammonia.

The nascent hydrogen which unites with the nascent nitrogen during the combustion is also derived from the organic matter which always contains enough carbon to decompose the water formed in order to be oxidized to carbon dioxid. While at first, therefore, during combustion, the hydrogen may unite with the oxygen, it becomes again free by the oxidation of the carbon and in this condition unites with the nascent nitrogen to form ammonia. In addition to carbon dioxid, ammonia, and free hydrogen there may also be found among the products of combustion marsh and olefiant gases and other hydrocarbon compounds which dilute, to a greater or less extent, the ammonia formed and help to carry it out of the combustion tube and into the standard acid.

169. The Official Method.—Reagents and Apparatus.—(1) Standard solutions and indicator the same as for the kjeldahl method:

(2) Dry granulated soda-lime, fine enough to pass a 2.5 millimeter sieve:

(3) Soda-lime, fine enough to pass a 1.25 millimeter sieve.[139]

Soda-lime may be easily and cheaply prepared by slaking two and one-half parts of quicklime with a strong solution of one part of commercial caustic soda, care being taken that there is enough water in the solution to slake the lime. The mixture is then dried and heated in an iron pot to incipient fusion, and, when cold, ground and sifted as above.

Instead of soda-lime Johnson’s mixture of sodium and calcium carbonate, or slaked lime may be used. Slaked lime may be granulated by mixing it with a little water to form a thick mass, which is dried in the water-oven until hard and brittle. It is then ground and sifted as above. Slaked lime is much easier to work with than soda-lime, and gives excellent results, though it is probable that more of it should be used in proportion to the substance to be analyzed than is the case with soda-lime.

(4) Asbestos, which has been ignited and kept in a glass-stoppered bottle.

(5) Combustion tubes about forty centimeters long and twelve millimeters internal diameter, drawn out to a closed point at one end.

(6) Large-bulbed U tubes with glass stop-cock, or Will’s tubes with four bulbs.

Manipulation.—The substance to be analyzed should be powdered finely enough to pass through a sieve of one millimeter mesh; from seven-tenths to one and four-tenths grams, according to the amount of nitrogen present, are taken for the determination. Into the closed end of the combustion tube, put a small loose plug of asbestos, and upon it about four centimeters of fine soda-lime. In a porcelain dish or mortar, mix the substance to be analyzed, thoroughly but quickly, with enough fine soda-lime to fill about sixteen centimeters of the tube, or about forty times as much soda-lime as substance, and put the mixture into the combustion tube as quickly as possible by means of a wide-necked funnel, rinsing out the dish and funnel with a little more fine soda-lime, which is to be put in on top of the mixture. Fill the rest of the tube to within about five centimeters of the end with granulated soda-lime, making it as compact as possible by tapping the tube gently while held in a nearly upright position during the filling. The layer of granulated soda-lime should not be less than twelve centimeters long. Lastly, put in a plug of asbestos about two centimeters long, pressed rather tightly, and wipe out the end of the tube to free it from adhering soda-lime.

Connect the tube by means of a well-fitting rubber stopper or cork with the U tube or Will’s bulbs, containing ten cubic centimeters of standard acid, and adjust it in the combustion furnace so that the end of the tube projects about four centimeters from the furnace, supporting the U tube or Will’s bulb suitably. Heat the portion of the tube containing the granulated soda-lime to a moderate redness, and when this is attained extend the heat gradually through the portion containing the substance, so as to keep up a moderate and regular flow of gases through the bulbs, maintaining the heat of the first part until the whole tube is heated uniformly to the same degree. Continue the combustion until gases have ceased bubbling through the acid in the bulbs, and the mixture of substance and soda-lime has become white, or nearly so, which shows that the combustion is finished. The process should occupy about three-quarters of an hour, or not more than one hour. Remove the heat, and when the tube has cooled below redness break off the closed tip and aspirate air slowly through the apparatus for two or three minutes to bring all the ammonia into the acid. Disconnect the tube, wash the acid into a beaker or flask, and titrate with the standard alkali.

During the combustion the end of the tube projecting from the furnace must be kept heated sufficiently to prevent the condensation of moisture, yet not enough to char the stopper. The heat may be regulated by a shield of tin slipped over the projecting end of the combustion tube.

It is found very advantageous to attach a bunsen valve to the exit tube, allowing the evolved gases to pass out freely, but preventing a violent sucking back in case of a sudden condensation of steam in the bulbs.

170. The Official French Method.—The French chemists prefer to drive out the traces of ammonia remaining in the combustion tube by means of the gases arising from the decomposition of oxalic acid.[140] The operation is conducted by mixing about one gram of oxalic acid with enough of dry granular soda-lime to form a layer of four centimeters in length at the bottom of the tube. The rest of the tube is then charged substantially as directed above. At the end of the combustion the oxalic acid is decomposed by heat furnishing sufficient hydrogen to remove from the tube all traces of ammonia which it may contain. The French chemists employ, for titration, either normal acids and alkalies or some decimal thereof or else an acid of such strength as to have each cubic centimeter thereof correspond to ten milligrams of nitrogen, thus making the computation of results exceedingly simple. Such an acid is secured when one liter thereof contains thirty-five grams of pure monohydric sulfuric acid or forty-five grams of pure crystallized oxalic acid. The corresponding alkaline reagent should contain, in each liter, forty grams of pure potassium hydroxid.

171. The Hydrogen Method.—Thibault and Wagner recommend that the combustion with soda-lime be conducted in an atmosphere of hydrogen,[141] and Loges replaces this by common illuminating gas freed from ammonia by conducting it through a tube filled with glass balls moistened with dilute sulfuric acid.[142]

In these cases the combustion tube is left open at both ends and the materials under the tube confined to the proper position by asbestos plugs. The gases used act in a merely mechanical manner and their use affords so few advantages over the method of aspirating air at the end of the combustion as to render it unadvisable.

172. Coloration of the Product.—It often happens, especially in the combustion of animal products, such as tankage and fish scrap, that the acid securing the ammonia is deeply colored by the condensation of some of the other products of combustion. This coloration interferes in a very serious way with the delicacy of the indicator used to determine the end of the reaction. In this case the liquid may be mixed with an alkali and distilled and the ammonia secured in a fresh portion of the standard acid as in the moist combustion process to be hereafter described.

173. General Considerations.—(1) Preparation of Sample.—In the soda-lime method it is of great importance that the organic substances be in a fine state of subdivision so as to admit of intimate mixture with the alkali. In cases where fragments of hoof, horn, hair, or similar substances are to be prepared for combustion it is advisable to first decompose them by heating with a small quantity of sulfuric acid. The excess of acid may be neutralized with marble dust and the resulting mixture dried, rubbed to a fine powder, and mixed with the soda-lime in the usual way. Care must be taken not to lose any of the ammonia from the sulfate which may be formed in mixing with the soda-lime in filling the tube.

(2) Purity of Soda-Lime.—The soda-lime employed must be entirely free of nitrogenous compounds and some blank combustions should be made in proof of its purity.

(3) Temperature.—The temperature of the combustion should not be allowed to exceed low redness. At very high temperatures there would be danger of decomposing the ammonia.

(4) Aspiration of Air.—Before aspiring a current of air through the tube to remove the last traces of ammonia the gas should be put out under the furnace and the tube be allowed to cool below redness to avoid any danger of acting on the nitrogen in the air.

174. The Ruffle Soda-Lime Method.—Many attempts have been made to adapt the soda-lime method to the determination of nitric nitrogen. Of these the process devised by Ruffle is the only one which has proved successful.[143] The method is founded on the action of sulfurous vapors on the nitrogen oxids produced during the combustion whereby sulfuric acid is formed and the nascent nitrogen is joined with hydrogen to form ammonia. By this process all the nitrogen contained in the sample, even if in the nitric form, is finally obtained as ammonia. In the original method the reagents employed were a mixture of sodium thiosulfate and soda-lime and a mixture of charcoal, sulfur, and granulated soda-lime. Subsequently the official chemists substituted sugar for the charcoal.[144] The method was used for a long time by the official chemists and came into general favor until displaced by the simpler and cheaper processes of the moist combustion method adapted to nitric nitrogen. As finally modified and used by the official chemists the process was conducted as described below.

175. The Official Ruffle Method.—[145]Reagents.—(1) Standard solutions and indicator the same as for the kjeldahl method.

(2) A mixture of equal parts by weight of fine-slaked lime and finely powdered sodium thiosulfate dried at 100°:

(3) A mixture of equal parts of weight of finely powdered granulated sugar and flowers of sulfur:

(4) Granulated soda-lime, as described under the soda-lime method:

(5) Combustion tubes of hard Bohemian glass seventy centimeters long and one and three-tenths centimeters in diameter:

(6) Bulbed U tubes or Will’s bulbs, as described under the soda-lime method:

Manipulation.—(a) Clean the U tube and introduce ten cubic centimeters of standard acid.

(b) Fit the cork and glass connecting tube. Fill the tube as follows: (1) A loosely fitting plug of asbestos, previously ignited, and then two and five-tenths to three and five-tenths centimeters of the thiosulfate mixture: (2) The weighed portion of the substance to be analyzed is intimately mixed with from five to ten grams of the sugar and sulfur mixture: (3) Pour on a piece of glazed paper or in a porcelain mortar a sufficient quantity of thiosulfate mixture to fill about twenty-five centimeters of the tube; then add the substance to be analyzed, as previously prepared, mix carefully, and pour into the tube; shake down the contents of the tube; rinse off the paper or mortar with a small quantity of the thiosulfate mixture and pour into the tube; then fill up with soda-lime to within five centimeters of the end of the tube: (4) Place another plug of ignited asbestos at the end of the tube and close with a cork: (5) Hold the tube in a horizontal position and tap on the table until there is a gas-channel along the top of the tube: (6) Make connection with the U tube containing the acid; aspirate and see that the apparatus is tight.

The Combustion.—Place the prepared combustion tube in the furnace, letting the open end project a little, so as not to burn the cork. Commence by heating the soda-lime portion until it is brought to a full red heat. Then turn on slowly jet after jet toward the outer end of the tube, so that the bubbles come off two or three a second. When the whole tube is red hot and the evolution of the gas has ceased and the liquid in the U tube begins to recede toward the furnace, attach the aspirator to the other limb of the U tube, break off the end of the tube, and draw a current of air through for a few minutes. Detach the U tube and wash the contents into a beaker or porcelain dish; add a few drops of the cochineal solution, and titrate.

176. Observations.—In our experience we have found it much more satisfactory to adhere to the earlier directions for preparing the mixture of thiosulfate and alkali. We much prefer to make the mixture with soda-lime and without the previous drying of the sodium salt. Ruffle himself says that the sodium thiosulfate should be dry but not deprived of its water of crystallization.[146] The best method to dry the crystal powder without depriving it of its crystal water is to press it between blotting papers. The official method also contains a typographical error in prescribing that the combustion tube should have a length of thirty centimeters where evidently thirty inches were meant. Ruffle’s original tube was twenty-two inches in length.

As is seen from the above description the method is essentially a reduction process by the action of a powerful deoxidizer in the presence of an alkali. The crystals of the thiosulfate salt cannot be brought into direct contact with a pure alkali, like soda or potash, without forming at once a wet mass which would tend to cake and obstruct the tube. The soda-lime is therefore a mechanical device to prevent this fusion. Where many analyses are to be made an iron tube, for economical reasons, may be substituted for the glass; but the glass tube permits a more intelligent observation of the progress of the analysis.

Since charcoal has very high absorbent powers it will be found always to contain a little nitrogen which may be in a form to generate ammonia during the combustion. The charcoal used should therefore be previously boiled with caustic soda or potash solution, dried, powdered, and preserved in well-stoppered bottles. Although pure sugar is practically free of nitrogen, even when it is used, it is advisable to occasionally make a blank determination and thus ascertain the correction to be made for possible contamination.

177. Boyer’s Modification of Ruffle’s Method.—The principle of the method rests on the observation that if nitrates be heated in a combustion tube with calcium oxalate and soda-lime, not more than two-thirds of the total nitrogen appear as ammonia; but if a certain proportion of sulfur be added the whole of the nitrogen is recovered.[23] The process may be divided into two reactions; viz.:

(1) Action of the calcium oxalate upon the sodium nitrate in presence of soda-lime:

(2) The action of sulfurous acid and of calcium oxalate upon the sodium nitrate in presence of soda-lime.

The analysis is conducted as follows: Dry and pulverize one-half gram of nitrate (Na or K) and mix it intimately with fifty grams of the reducing compound containing approximately ten per cent sulfur, 22.5 per cent neutral calcium oxalate, and 67.5 per cent soda-lime. The combustion tube is charged as follows:

The tube is heated gradually from the front backwards, the calcium oxalate furnishing finally the gas necessary to drive out the last traces of ammonia. The process is equally applicable to the determination of nitrogen in all its forms or to mixtures thereof.

The method has also been applied to the mixture of ammoniacal and organic nitrogen and to the mixture of ammoniacal, nitric, and organic nitrogen, the combustions having been made both in an iron and a glass tube. The amounts of material to be used vary from one-half gram to a gram, according to its richness in nitrogen.

The combustion should be terminated in forty minutes.

When a combustion is terminated, the acid containing the ammonia is placed in a beaker and boiled for two or three minutes to drive off the sulfurous and carbonic acids. The titration is then conducted in the usual manner.

The combustion can be carried on just as well in an iron tube as in a glass one. The reagents employed, especially soda-lime, being hygroscopic, a little water is disengaged in heating, which is condensed at the cold extremity of the tube, and which may absorb a little ammonia, unless special precautions are taken to have the materials dry.

THE MOIST COMBUSTION PROCESS.

178. Historical.—As long ago as 1868 Wanklyn proposed to conduct the combustion of organic bodies in a wet way, using potassium permanganate as the oxidizing body.[147] About ten years after this he attempted to extend the method so as to estimate the quantity of proteid matter in a sample by treatment with an alkaline solution in presence of the permanganate salt. One gram of the finely pulverized sample was treated in a liter flask with one-tenth normal potash lye. After digestion for some time, from ten to twenty cubic centimeters were taken for the determination. According to the supposition of Wanklyn, pure albuminoid matters thus treated yielded one-tenth of their weight of ammonia, or about fifty per cent of the total nitrogen appeared as ammonia. The ammonia content of the sample was determined by the colorimetric process devised by Nessler. It is needless to add that the process of Wanklyn proved to be of no practical use whatever, acting differently on different albuminoid matters, and even on the same substance. No other attempt was made to perfect the moist combustion process until Kjeldahl[148] introduced the sulfuric acid method in 1883. The simplicity, economy, and adaptability of this method have brought it into general use. At first the process was only applied to organic nitrogenous compounds in the absence of nitrates, but especially by the modifications proposed by Asboth, Jodlbaur, and Scovell, it has been made applicable to all cases, with the possible exception of a few alkaloidal and allied bodies. The moist combustion process for determining nitrogen is now generally employed by chemists in all countries, not only for fertilizer control, but also for general work.

179. The Method of Kjeldahl.—The process originally proposed by Kjeldahl is applicable only to nitrogenous bodies free of nitric nitrogen. The principle of the process is based on the action of concentrated sulfuric acid at the boiling-point in decomposing nitrogenous compounds without producing volatile combinations and the subsequent completion of the oxidation by means of potassium permanganate. The original process has been modified by many analysts but the basic principle of it has remained unchanged. It will therefore prove useful here to describe the process as originally given.[149]

The weighed substance is placed in a small flask. With solid bodies this is a very simple operation, but with liquids more difficult. Liquids which are not decomposed, on heating, should be evaporated in a thin glass dish which can be ground up and placed in the digestion flask with the desiccated sample. The strongest sulfuric acid is added in sufficient quantity to secure complete decomposition, not less than ten cubic centimeters in any case. The acid must be free of ammonia and be kept in such a way as not to absorb ammonia from the atmosphere of the laboratory. To guard against danger of error from such an impurity frequent control determinations should be made. In control experiments one or two grams of pure sugar should be used as the organic matter. If the acid employed contain traces of ammonia the necessary corrections should be made in each analysis. The flask having been charged is placed on a wire gauze over a small flame. The organic matter becomes black and tar-like and soon there is a rapid decomposition attended with the evolution of gaseous products among which sulfur dioxid is found. To avoid danger from spurting, the digestion flask should be placed in an oblique position. The flask should have, at least, a capacity of 100 cubic centimeters and a long neck and be made of a kind of glass capable of withstanding the action of the boiling acid. Particles of the carbonized organic matter left on the sides of the flask by the foaming of the mass at first are gradually dissolved by the vapors of the boiling acid as the digestion proceeds. The action of the sulfuric acid is not entirely finished when gases cease to be given off but the digestion should be continued until the liquid in the flask is clear and colorless or nearly so. Usually about two hours are required to secure this result. When aided by the means mentioned below the time of digestion can be very materially shortened. By adding some fuming sulfuric acid, or glacial phosphoric acid, the dilution caused by the formation of water in the combustion of the organic matter, can be avoided. For albuminoid bodies it is hardly necessary to continue the combustion until all carbonaceous matter is destroyed. The full complement of ammonia is usually obtained after an hour’s combustion even if the liquid be still black or brown, but with other nitrogenous bodies the case is different so that upon the whole it is safest to secure complete decoloration.

The temperature must be maintained at the boiling-point of the acid or near thereto since at a lower temperature, for instance from 100° to 150°, the formation of ammonia is incomplete. Since all organic substances of whatever kind are dissolved by the boiling acid the previous pulverization of the material need be carried only far enough to secure a fair sample. Many substances give up practically all their nitrogen as ammonium sulfate when heated with sulfuric acid as, for instance, urea, asparagin, and the glutens. In most of the other organic bodies fully ninety per cent of the nitrogen are likewise secured as the ammonium salt. In the aromatic compounds, or even in the form of amid in anilin salts, the nitrogen is more resistant to the action of sulfuric acid. In the alkaloids where the nitrogen is probably a real component of the benzol skeleton the formation of ammonia is very incomplete. But even in the cases where the conversion of the nitrogen into ammonia is practically perfect it is advisable to finish the process by completing the oxidation with potassium permanganate. The permanganate should be used in a dry powdered form and added little by little to the hot contents of the digestion flask, the latter being held in an upright position and removed meanwhile from the lamp. When carefully performed there is no danger of loss of ammonia although the oxidation is, at times, so vigorous as to be attended with evolution of light. The permanganate must always be added in excess and until a permanent green color is produced. The flask is then gently heated for from five to ten minutes over a small flame, but this is not important. The heating must not be too strong or else a strong evolution of oxygen will take place with a consequent reduction of the manganese compound. When this happens the liquid again becomes clear and there is a loss of ammonia.

After cooling, the contents of the flask are diluted with water, the green color giving place to a brown with a rise of temperature. After cooling a second time the whole is brought into a distillation flask of about three-quarters of a liter capacity and attached to a condenser which ends in a vessel containing titrated sulfuric acid. About forty cubic centimeters of sodium hydroxid solution of one and three-tenths specific gravity are added and the stopper at once inserted to prevent any loss of ammonia. To prevent bumping during the distillation some zinc dust is added securing an evolution of hydrogen during the progress of the distillation. In this case the bumping is prevented until near the end of the operation when it begins anew, probably by reason of the separation of solid sodium sulfate. After the end of the distillation, the excess of acid remaining in the receiver is determined by a set alkali solution and thus the quantity of ammonia obtained easily calculated. Kjeldahl, however, preferred to titrate the solution after adding potassium iodate and iodid, a mixture which in the presence of a strong acid sets free a quantity of iodin equivalent to the free acid present. The iodin thus set free is titrated by a set solution of sodium thiosulfate using starch as an indicator. The merits of this method are sharpness of the end reaction and the possibility of using only a small quantity of the nitrogenous body for the combustion. The sulfuric acid used in the receiver is made of the same strength as the thiosulfate solution; viz., about one-twentieth normal. Thirty cubic centimeters of this were found to be the proper amount for use with substances taken in such quantities as to produce ammonia enough to neutralize about half of it. The titration is carried on as follows: A few crystals of potassium iodid are dissolved in the acid mixture obtained after the distillation is completed, then a few drops of the starch-paste and finally a few drops of a four per cent solution of potassium iodate. The iodin set free is then oxidized by the addition of the one-twentieth normal sodium thiosulfate solution until the blue color disappears.

In the computation it is more simple to multiply the corresponding number of cubic centimeters of thiosulfate by seven, half the atomic weight of nitrogen, and divide the product by the weight of the substance taken, which will give the per cent of nitrogen therein.

A more detailed description of the method of making the titration follows: After the distillation is finished the condensing-tube is rinsed with a little water, after which the sulfuric acid unneutralized in the receiver is determined. It is advisable first to test the reaction of the distillate with litmus paper before going any further; for if at any time all the acid should be found neutralized it will be necessary to add a sufficient quantity of one-twentieth normal sulfuric acid before adding the potassium iodid, etc., otherwise the determination will be irreparably lost. Add to the contents of the flask ten cubic centimeters of the potassium iodid and two cubic centimeters of the potassium iodate solutions, described further on, and the sodium thiosulfate is then run in from a burette till the fluid, which is constantly kept agitated by shaking the flask, shows only a bare trace of yellow coloration from the iodin still present. Starch solution is then added, and the blue color obtained is at once removed by additional thiosulfate solution. When some experience has been gained, the eye is able to discern, with great certainty, even the slight coloration caused by only a small trace of free iodin.

In regard to the sensitiveness of the end reaction and the accuracy of the result, this method of titration leaves nothing to be wished for. The strength of the thiosulfate solution is determined in exactly the same manner, and with starch as an indicator. For this purpose, measure ten cubic centimeters of one-twentieth normal sulfuric acid into an erlenmeyer, add 120 cubic centimeters of ammonia-free water, ten cubic centimeters of potassium iodid solution, and two cubic centimeters of iodate solution; add thiosulfate solution till the fluid shows only the above-mentioned light yellow tint, then add starch, and finally thiosulfate. In this way the strength of the thiosulfate is ascertained, which of course must be occasionally re-determined, under exactly the same conditions as with the nitrogen determinations themselves, and every possible error is thereby excluded. That the solution once decolorized within a short time again assumes a deep blue color, is a matter of no concern, inasmuch as both solutions are added in such a manner that the end reaction lies exactly at the point when the starch iodid reaction distinctly disappears.

180. Theory of the Reactions.—As has been seen above the final product of heating a nitrogenous organic compound with sulfuric acid and an oxidizing body is ammonium sulfate. The various steps by which this is obtained have been traced by Dafert:[150]

(1) The sulfuric acid abstracts from the organic matter the elements of water:

(2) The sulfur dioxid produced by the action of the residual carbon on sulfuric acid exercises a reducing effect on the nitrogenous bodies present:

(3) From the nitrogenous bodies produced by the above reduction ammonia is formed by the action of an oxidizing body:

(4) The ammonia formed is at once fixed by the acid as ammonium sulfate. According to the theory of Asboth the hydrogen which is formed during the action of sulfuric acid on organic matter, when in a nascent state, also aids greatly in the production of ammonia. This idea is based on the fact that with those bodies which afford a deficit of hydrogen the formation of ammonia is imperfect.[151]

181. Preparation of Reagents.—(1) Pure Sulfuric Acid.—As is well known the so-called pure sulfuric acid in the market usually contains ammonia, a fact which compelled Kjeldahl to determine the quantity of nitrogen in the acid in every instance, and to make correction for the same in the analysis. An acid absolutely free from this impurity may, however, easily be prepared by the distillation of the commercial article in a small glass retort holding easily about 400 cubic centimeters. To conduct this operation without danger it is only necessary to arrange the apparatus so that the heavy fluid is heated to boiling, not from the bottom of the retort, but from its sides, and that the upper portion of the body and neck is kept sufficiently warm so that the sulfuric acid fumes are not allowed to condense and flow back into the retort. Both these ends are attained simply by surrounding the retort with a piece of sheet iron, cylinder-shaped beneath, and with an oval upper part, having an opening of about one centimeter in diameter for the neck of the retort. To conduct the distillation a burner is used with an arrangement for spreading the flame. To avoid with certainty all bumping of the sulfuric acid and the resulting danger therefrom, the lamp is so arranged that only the products of combustion go up between the retort and its iron hood, without allowing the flame itself to come into contact with the glass vessel. The retort should be filled about half full, or with 200 cubic centimeters of acid. By this device, without any danger whatever, about one liter of sulfuric acid may be distilled in a day. The retort will stand numerous distillations. Once begun, the distillation takes care of itself; it is necessary to discontinue it when only the bottom of the retort is covered with sulfuric acid, and to fill fresh acid through a funnel when the retort has cooled off. The first twenty cubic centimeters of the distillate going over are collected by themselves and rejected. What comes over later is, as shown by experience, absolutely ammonia-free, and can be used without any correction, for the nitrogen determinations according to Kjeldahl. The acid is kept in a stoppered bottle in a place not reached by ammonia fumes. The ten cubic centimeter pipette used for measuring the quantity of sulfuric acid required for each determination, is fastened in the perforated rubber stopper with which the bottle is kept closed, and is itself closed above by a small rubber tube with a plug of glass wool in it.

(2) Potassium Permanganate.—Crystals of this salt are crushed (not pulverized) with a pestle into small pieces of about one-half millimeter size, which are kept in a long glass tube of about ten millimeters diameter, closed with a stopper.

(3) Ammonia-free Water.—Common distilled water cannot be used in the determination of nitrogen according to Kjeldahl, since it contains ammonia. It may be obtained free from the same by redistillation in a large glass retort with the addition of a few drops of sulfuric acid. All vessels used in the determination are rinsed out beforehand with this water.

(4) Ammonia-free Soda-lye is most conveniently prepared by adding 270 grams of common sodium hydroxid in sticks, little by little, to one liter of distilled water which is kept continually boiling, by means of a small flame, in a good-sized silver dish. The dish is kept covered with a glass plate. Care has to be exercised not to add the alkali too rapidly, nor in too large quantities at a time for in this case the fluid will boil too violently at every addition of the alkali. After the operation is finished the lye is at once siphoned into a glass flask, and when cold, is poured into a glass-stoppered bottle.

(5) One-twentieth Normal Sulfuric Acid is prepared from sulfuric acid and water both absolutely ammonia-free, and is kept in a place where no fumes of ammonia can reach it, in a well-stoppered glass bottle, the stopper being smeared with vaseline.

(6) Sodium Thiosulfate Solution.—This should be of the same strength as the one-twentieth normal sulfuric acid. It is prepared by dissolving the salt in ammonia-free water, and is compared with the acid, to which has been added potassium iodid and iodate, using starch as an indicator, in the manner described above. The solution is kept in a well-stoppered bottle, in the dark. When the salt and water used are perfectly pure, it will keep unchanged for a long time.

(7) Potassium Iodid.—Dissolve five grams of chemically pure potassium iodid in ammonia-free water and make the volume 100 cubic centimeters. Ten cubic centimeters of this solution are used for each determination; keep the solution in the dark and in a well-stoppered bottle.

(8) Potassium Iodate.—Dissolve four grams of chemically pure potassium iodate in ammonia-free water and make the volume 100 cubic centimeters. Use two cubic centimeters of this solution for each determination.

(9) Starch Solution.—Digest pure starch for about a week with dilute hydrochloric acid, wash perfectly free from chlorin by decantation, and finally dry it between filter-paper. The starch is then dissolved in water with the aid of heat. Such a solution will keep for an indefinite time, if it be saturated with common salt. Ten grams of this starch are dissolved in 1,000 cubic centimeters of ammonia-free water; use one or two cubic centimeters for each determination.

182. Kjeldahl Method as Practiced by the Holland Royal Experiment Station.—Necessary Reagents: 1. Phosphosulfuric acid, made by mixing a liter of sulfuric acid of specific gravity 1.84 with 200 grams of phosphoric anhydrid:

2. Alkaline sodium sulfid solution, made by dissolving 500 grams of sodium hydroxid and six grams of sodium sulfid or eight and one-half grams of potassium sulfid, in a liter of water:

3. Mercury:

4. Paraffin in small pieces:

5. Dilute sulfuric acid and dilute potash solution, both of known strength:

6. Pieces of previously ignited pumice stone or of granulated zinc:

7. Neutral solution of rosolic acid or litmus.

Apparatus: The apparatus necessary consists of oxidation flasks of about 200 cubic centimeters capacity and distillation flasks of about 500 cubic centimeters capacity, both of bohemian glass. Copper may be used for the distillation flasks.[152]

The Process: A gram of the sample to be analyzed is placed in an oxidation flask together with twenty cubic centimeters of phosphosulfuric acid and a drop of mercury, about 600 milligrams, and heated till the fluid becomes colorless. After cooling, dilute and wash the contents of the flask into a distillation flask. The resulting volume should be about 300 cubic centimeters. Add 100 cubic centimeters of the alkaline sodium sulfid solution and some pieces of ignited pumice stone or granulated zinc. Distill the ammonia, receiving the distillate in a flask containing a known volume of the standard sulfuric acid. Titrate with tenth-normal potash, using litmus or rosolic acid as indicator.

183. The Kjeldahl Method as Practiced at the Halle Station.—The method at present in vogue in the German stations of conducting the moist combustion process is well illustrated by the method of procedure followed at Halle.[29] From seven-tenths to one and five-tenths grams of the sample are taken for analysis according to its richness in nitrogen. Because of the fact that so small a quantity of the sample is taken it is of the highest importance that it be perfectly homogeneous throughout its entire mass. Otherwise, grave errors may arise. From the sample, as sent to the laboratory, the analyst should take a subsample and this should be rubbed to a fine powder and the part used for analysis carefully taken therefrom. If the sample be moist it may be rubbed up with an equal weight of gypsum, in which case a double quantity is taken for the determination. Substances like bone-meal, which do not keep well mixed, especially when occasionally shaken, should be intimately mixed before each weighing. The sample taken for analysis is placed in a glass flask of about 150 cubic centimeters capacity. The flasks should be made of a special glass to withstand the tension of the combustion. Those made by Kavalier at Sazava, in Bohemia, have proved to be the most lasting. A globule of mercury weighing a little less than one gram is placed in the flask and also twenty cubic centimeters of pure sulfuric acid of 1.845 specific gravity. The mercury is conveniently measured by an apparatus suggested by Wrampelmayer. It consists of an iron tube holding mercury, and is conveniently filled, from time to time, from a supply vessel placed in a higher position and joined by means of a heavy glass tube and rubber tube connections. The lower end of the iron tube is provided with a movable iron stopper having a pocket just large enough to hold a globule of mercury, weighing a little less than a gram. On turning the stopper the pocket is brought opposite a discharge orifice and the measured globule of mercury is discharged. With substances which tend to produce a strong foaming a little paraffin is used. The flasks after they are charged are placed on circular digesting ovens under a hood as shown in figure 11.