THEPRESENTCARLSBERG LABORATORY, COPENHAGEN The director is S. P. L. Siirensen, best known to students of chemistry as the inventor of the pH concept. This laboratory was opened in 1896.
KJELDAHL and the DETERMINATION of NITROGEN RALPH E. OESPER University of Cincinnati, Cincinnati, Ohio
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OMMEMORATION of the anniversaries of his- the protein content of various animal and vegetable products torical events is a practice which the chemical which are often best assayed through this content. Consequently, there is frequent and imperative necessity for nitrogen world sometimes follows, and as the Kjeldahl determinations in technical research laboratories, in agricultural method of determining nitrogen has just enlered on the experiment stations, in physiological institutes. I n many cases second half-century of its usefulness, this seems an such determinations made regularly would be of valuable aid in appropriate time to pay tribute to this invaluable the control of certain industrial processes utilizing nitrogenous materials. If, however, nitrogen determinations are not analytical aid and to its inventor, to whom its dis- raw made oftener than is now the case, the reasons are not far to covery brought a lasting international reputation. seek. This determination is always a-relatively tedious task, The method was made public for the fist time in a no matter whether the Dumas or the Will-Varrentrapp method lecture delivered to the Chemical Society of Copen- he employed, full advantage being taken of all the impravehagen on March 7, 1883, and shortly thereafter ap- ments and simplifications of the technic which the experience of many years and the co6peration of many workers have brought peared in the Zeitschrift fGr amlytische Chemie (22, t o these methods. A single determination requires several hours' 366) under the title, "A New Method of Determining work, and the constant attention with which the whole course Nitrogen in Organic Materials." The paper opens of the analysis must be followed precludes the carrying out af many analyses simultaneously. Hence such analyses, if a whole with an apposite discussion: The determination of nitrogen occupies a place of special and important significance in the ultimate analysis of organic materials. While the quantitative estimation of carbon and hydrogen is usually of interest only in purely scientific studies-the fixing of the cornnosition of new comnounds. etc.-the determination of nitrogrn is eqwcially important in technical practice siucc it afford9 tlsc only fairly accurate means now known oi c\.aluatmg ~
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series of nitrogen determinations must be made daily, can only be accomplished with great expenditure of labor. Furthermore, the usual ultimate analysis demands a certain degree of proficiency and can therefore be accomplished only by skilled chemists, and also an expensive special piece of apparatus (combustion furnace) is necessary. Therefore i t is clear why these determinations could only he carried out in real chemical laboratories and why in these they have taken up a disproportionate share of the time of the chemist and nhvsiolo~ist.
as there was available another method which is not only as reliable but simpler, quicker, less expensive, more convenient, and in general better suited to the quickened pace of pure and applied chemistry. The Dumas method, in the hands of competent analysts who are cognizant of the errors attending the gasometric determination of nitrogen, is doubtless the most reliable and accurate of all presentday procedures, but this was not always the case. So uncertain was its performance in its youth that though it [Ann. chim. phys., [2],47, 198 (1831)l was ten years the senior of the Will-Varrentrapp method, the latter almost immediately took the center of the stare and indisputably held that position for KJELDAHL I N THE OLD CARLSBERG LABORATORY, COPENHAGEN twenty years. The Dumas method This is a photograph of an oil painting by Haslund which is hung in the is so well known that even an abcouncil room of the Carlsberg Laboratory. Opposite is hung the companion picture of Kjeldahl's colleague, Hansen, reproduced on page 462. stract of the procedure is superfluous here, but most of those who use it have little, if any, knowledge A closer examination of the pre-Kjeldahl methods of the efforts that have gone into bringing it into its will not only emphasize what he himself wrote -but present state of excellence. The history of an anawill give a truer background from which to appraise lytical procedure is often as fascinating and instructive the value of his contribution. The WiU-Varrentrapp as the biography of a scientist, and many teachers method has now practically passed into oblivion; not as well as their students would profit greatly by such many of those practicing chemistry today have used studies. They would then learn the great truth that it, though occasionally it gives reliable results when analytical procedures are not, foimd inscribed on tabother methods fail. Brought out in 1841 (Ann., 39, lets of stone but are products of evolution, and their 257) it was welcomed, for its essential feature was the inherent faults, like those of humans, are eradicated conversion of the nitrogen of the sample into a chemi- largely by the labors of those who work not for their cal compound whose quantitative determination was own advancement but for the making of a better easy and sure as compared with the uncertainties then world. Some of the problems that, had to be solved residing in the gasometric methods. The method in before the Dumas method could become worthy of a brief is as follows: The sample toge+er with soda place in the chemist's annory were the discovery of: lime is placed in a tube drawn out to a point at one end, (1) tractable sources of carbon dioxide; (2) methods the other end being joined to a receiver charged with of completely removing the air from the combustion acid. The contents of the tube are carefully heated, tube; (3) sure means of de-oxidizing nitrogen oxides; the nitrogen is evolved as ammonia (or related bases) (4) impeccable reducing agents for the copper spiral, and after the reaction is over, the tip of the tube is since hydrogen was found to be absorbed and later rebroken off and the residual gases are drawn through leased; (5) simple yet trustworthy amtometers. All the acid by aspiration. Originally the analysis was the defects of the method have not yet been comcompleted by determining the ammonia produced as pletely eliminated and the excellent results it delivers chloroplatinate or platinum, but with the development are probably due to a fortunate compensation of errors. of volumetric analysis it was found more convenient No matter what improvements are attained, the to absorb the ammonia in standard acid. Of the Dumas and the Will-Varrentrapp methods must numerous modifications which have been suggested patently remain tedious, expensive, and delicate operfew have proved of real value and the same may be ations and utterly unsuited to large-scale, routine desaid of the criticisms aimed a t this method. It is, of mands. Furthermore, samples that resist being finely course, not applicable to nitro compounds, etc., but divided obviously cannot be properly mixed with soda used understandingly and skilfully it will do all that lime, copper oxide, etc., and the analysis of liquid its inventors claimed for it and it played an honor- samples also presents difficulties. It is not surprisable part in the development of organic chemistry. ing then that attempts were made to substitute flasks Of necessity it was doomed to abandonment as soon for the fragile tubes and also to discover wet methods
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AUGUST,1934 that, because of their easier regulation, would obviate the constant supervision occasioned by the heating of dry mixtures. These schemes were either of limited applicability or were burdened with "improvements" until they became too complicated to be of general value. Mere oxidation of the samples gave indifferent success and, following the lead of the Will-Varrentrapp method, boiling with caustic solutions or fusion with alkalies gave satisfactory results only. with compounds which are direct derivatives of ammonia. However, these attempts showed tbat complete ammonification can only be expected to follow a preliminary destructive treatment of the organic substance. Alkaline permanganate gives incomplete decomposition but, nevertheless, this formed the basis of Wanklyn's method (1877) of determining the protein content of vegetable material, a method which, though unreliable, was widely used. Grete (1878) and Dreyfus (1883) used concentrated sulfuric acid in the preliminary treatment of wool, born, leather, and fertilizers in order to make these substances, which are bard to subdivide, more susceptible of attack by soda lime. The charred acid residue was mixed with an excess of soda lime or calcium carbonate and the resulting dry mass was transferred to a tube and cariied through the Will-
459 version of the nitrogen of proteins, etc., into ammonia could be assured. Reasoning that there would be a greater tendency toward ammonia formation in the presence of acid, he boiled his samples with solutions containing dilute sulfuric acid and excess permanganate. Alkali was then added, the ammonia distilled out, and though the yield was higher, the conversion was still quite incomplete and the results fluctuated. Encouraged, he continued his efforts along this line and eventually discovered that the total nitrogen content of practically all the materials in which he was then interested could be converted into ammonia by treating the material first with boiling concentrated sulfuric acid until a solution was formed in which the nitrogen was either present as ammonium sulfate or in such combination that oxidation by the addition of excess powdered permanganate completed the ammonification. The firm foundations of the method had been laid, but much tedious labor was consumed before the method could be given to the chemical world. Kjeldahl worked out the numerous details, he checked and tested each step, and when his paper appeared it set forth a really practicable procedure. The relative simplicity of the new method attracted widespread attention; it was promptly tested, particularly in the German agricultural stations, and the verdict was favorable. Experience not only confirmed the excellences but also brought to light possible improvements, for the method possessed the high virtue of not only being better but of being capable of betterment. The improvements and modifications ran chiefly
JEANBAPTISTE ANDRBDUMAS 1800-1884
Varrentrapp procedure. At this point Kjeldahl appeared on the scene, and he bad a very personal stake in the solution of this problem because the progress of his researches was greatly handicapped by the nonexistence of a quick yet reliable method of determining nitrogen in grains, malts, etc. 1-4 Kjeldahl knew tbat the simple apparatus and the easy technic of the Wanklyn method were well suited to his purpose and so he began the search to so modify this admittedly defective method that complete con-
AN OLD FHOTOGR~PH OP KJEDAHL
nol followed by reduction of the nitrophenolsulfonic acid. The relatively short interval between the first announcement of the method and publications of modifications and improvements should not be construed as evidence that the method in its original form left much to be desired but should rather be taken as a testimonial of the intense interest aroused by this useful addition to the analyst's repertoire. The subsequent work on this method has added nothing of fundamental significance, though special mention should be made of Dafert's studies of the applicability of the Kjeldahl method and its various modificaA GROUPIN KJELDAKL'S LA~ORATORY tions. He was able to lay down Reading from left to righl: ( 1 ) Mr. H. Jessen-Hansen, first assistant to Kjeldahl, general rules as to when preliminary and then to S6rensen; (2) Mr. S Orla-Jensen, now professor of biochemistry, Copenhagen; (3) Kjeldahl; (4) Mr. Iwan, now technical director a t a brewery in Oslo. treatment is required, and his classifications include most of the in the following channels: (1) the use of mercury types of nitrogenous materials, which is tantamount and copper compounds, etc., as accelerators in the to saying that only the exceptional cases are not digestion, a field opened up by Wilfarth in 1885; (2) amenable to treatment by the method in its present the inclusion of potassium sulfate in the digestion mix- form. A comparison of current practice with the ture, an advance suggested by Gunning in 1889; (3) original procedure shows, in addition to the foregoing, the development of multiple digestion and distillation variation only in such details as the use of the same equipment, the first example of many being that con- flask for both digestion and distillation, a safer method structed in 1884 by Heffter, Hollrung, and Morgen; (4) of adding the alkali preparatory to the distillation, the multifarious stillheads or spray traps, the earliest the use of paraffin to abate foaming, the omission of design being that of Reitmair and Stutzer, 1885 the permanganate in many cases where accelerators (Kjeldahl himself, in 1888, described a device in which are used, and finally the back-titration of the excess the ammonia-bearing vapors were scrubbed free of sodium hydroxide); (5) extension of the method to materials requiring pre-treatment before digestion with the acid. Kjeldahl knew that his method was not applicable to nitrates, nitro bodies, alkaloids, and various other classes of compounds, but to his surprise he found that part of the non-amide nitrogen of such materials is converted into ammonia by digestion with concentrated sulfuric acid. For example, mixtures of potassium nitrate and sugar gave 60-80% yield of ammonia. Following this lead, Asboth in 1886 obtained fairly good results when he added sugar to a number of these resistant compounds, and even the nitrates were brought more nearly into line by the addition of benzoic acid. Jodlhauer, in this same year, made this modification really workKJELDAHL'S NEW LABORATORY able by substitutingfor the benzoic Reading from left to right: ( 1 ) Mr. H. Jessen-Hansen, (2) Kjeldahl, and (3) Mr. acid the more easily nitrated pheN. Hjelte Claussen, now technical director of the Carlsberg Breweries.
acid with standard alkali in place of the iodometric institution was part of this plan and the Carlsberg titration recommended by Kjeldahl. He certainly Laboratory came into being. Jacobsen requested his would have rejoiced in the further simplification made son to transfer Kjeldahl and Hansen to the new labopossible by the Winkler (1914) method of absorbing ratory and in October, 1876, they began theirwork under the new auspices, in charge of the chemical and physiothe ammonia in boric acid. The life story of Johan G. C. T. Kjeldahl is soon told. logical sections, respectively. The rest of their lives The son of a physician, he was born on August 16, were spent in the service of science in this institution 1849, a t Jaegerspris in the northern part of the Danish whose high purposes are set forth in a tablet placed a t island, Zealand. He received his baccalaureate degree the threshold of the laboratory: "No result achieved from the Gymnasium a t Roskilde in 1867, and then through the activity of the Carlsherg Laboratory and went to Copenhagen where he specialized in chemistry having either theoretical or practical significance is to and physics, attending both the University and the be held secret." The work on the brewing industry was continued, but Polytechnic High School. In 1873 he passed the state examination in "applied science" with distinction, and on a much wider scale, and most of Kjeldahl's researches soon thereafter was appointed assistant to C. T. Bar- were related to the fermentation industries. The chief
F n ~ mVARRENTKAPP August 29, 1815-March 1, 1877 Photograph taken from Berichte, 10, 2291 (1877).
Photograph taken from Berichte, 23, Rep. 852,. 1890.
J . \V G u n ~ l x c 1527-1900
Photograph from Cohen's "Jacobus Hsnrieus van't Hoff, rein Leben und Wirken." p 169.
foed. The latter was well pleased with his work and recommended him highly to Carl Jacobsen, the brewer, who had decided that his brewery could he more successfully operated if daily tests on the beer, yeast, malt, etc., were made by men of scientific training. Kjeldahl entered on this service May 1, 1875, and was entrusted with the fitting up of the works laboratory and putting it into operation. Associated with him, as biolopist, was E. C. Hansen (1842-1909) who later achieved an international reputation as the originator of pure yeast cultures. J. C. Jacobsen, the father of Kjeldahl's employer, was also a brewer and he engaged in business primarily to secure funds for his philanthropic ventures. In 1876 he formed the darlsberg Foundation for the promotion of arts and sciences. A scientific research
fields of his endeavor were enzymatic action, the determination of sugars singly and in mixtures by the copper reduction method, the optical activity of vegetable proteins, and their solubility relations in alcohol of varying strengths. In 1881. while studvine the chanees in rotei in con, " tent of grain during germination and fermentation, he found his progress blocked by the inadequacy of the available methods of determining nitrogen. He resolutely put aside all other work and devoted himself to the quest for this indispensable analytical weapon. His success was his monument. Kjeldahl's constitution was not rugged; part of his mental resources were expended on his enthusiastic pursuit of art and letters; he could not endure long periods of intense laboratory toil. The volume of his ~
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Elan Canrsnm HANSEN, IN THE
OLD CARLSBERG LABORATORY
This painting by 0. Haslund is the companion picture to that on page 458.
work was not large but its quality was always high. He had many of the qualities of a good teacher; he gave generously of his time to young men of his circle who were striving to improve their scientific and technical knowledge. He often regretted that his laboratory duties allowed him no opportunity to give regular
courses of instruction, for he felt this type of endeavor would have been quite beneficial to him personally, particularly when in his last years he suffered from periods of mental fatigue. Kjeldahl was elected to membership in the scientific academies of Denmark and Christiania, and received an honorary doctorate from the University of Copenhagen. On July 18, 1900, while bathing a t Tisvilde, Zealand, he suffered a heart attack and died in the water.
****** The author gratefully acknowledges the courtesy of Prof. S. P. L. Sorensen who provided the pictures of Kjeldahl and the Carlsberg Laboratory.
