Oct., 19511 HAMENCE 563 The Determination of the Availability of Nitrogen in Nitrogenous Fertilisers. Part 11* BY J. HUBERT HAMENCE (Presented at the meeting of the Society on Wednesday, April 4th, 1951) I n 1949 the author described a method for the determination of the available nitrogen in fertilisers based upon a determination of the nitrate produced when the fertiliser is mixed with soil under conditions that give maximum nitrification. The nitrate produced is compared with the nitrate produced from dried blood containing an equivalent amount of nitrogen and treated under similar conditions. The availability of the fertiliser is calculated by taking that of dried blood as 100. This test has now been applied to the majority of the common nitrogenous fertilisers and the results are given in this paper.It is shown that additional valuable information concerning the availability of the nitrogen may be gained by determining the nitrate produced a t the end of three or more different intervals of time and plotting and studying the time - nitrate curve. With very slow-acting fertilisers it is necessary to extend the period of test beyond the usual 21 days. The initial absorption of nitrate nitrogen from the soil, which takes place during the decomposition of some fertilisers containing substantial amounts of carbohydrates, can be demonstrated by this test. IN an earlier paper1 the author described a method for the determination of the relative availability of the nitrogen in nitrogenous fertilisers. This method is based upon a deter- mination of the nitrate nitrogen produced from the fertiliser when it is mixed with soil under such conditions as to give the maximum rate of nitrification. The nitrate nitrogen produced from the fertiliser in 18 days is compared with the quantity of nitrate nitrogen produced from dried blood containing an equivalent amount of nitrogen and treated under identical conditions.The availability of the fertiliser is calculated by taking that of dried blood as 100. The results for the availability, as determined by this method, of some of the more common nitrogenous fertilisers were given in that paper (Zoc. cit.). From the results the opinion was expressed that this test gave a better measure of availability of the nitrogen in nitrogenous fertilisers than the Street permanganate test,2 which has been widely used for this purpose.The same conclusion was arrived at by Clark and G a d d ~ , ~ who compared the behaviour of the water-insoluble nitrogen in a wide range of mixed fertilisers, which are available on the American market, in the neutral and alkaline permanganate tests with the corresponding nitrification values. As the result of this work they report that the lack of correlation between alkaline and neutral permanganate activities and observed nitrification values indicates that the official methods of the A.O.A.C. are inadequate for proper characterisation of the insoluble nitrogen content of many organic materials. A similar conclusion was arrived at by Owen, Rogers and Win~or,~ who were carrying out nitrification studies of nitrogenous fertilisers at the same tifie as the author.They used larger quantities of soil than did the author, although in some of the tests the ratio of fertiliser to soil was substantially the same. The tests differ in that the nitrification was studied over a longer period than the 21 days that was suggested as the standard time in the new technique. Where the results of the two laboratories impinge they are in general agreement. Time - “nitrate produced” graphs are given by Owen, Rogers and Winsol“‘ for a number of the commoner nitrogenous fertilisers. The new test has been applied also to new materials of unknown value and the results of laboratory tests with this technique have been confirmed subsequently by field trials. * Part I appeared in J .Sci. Food Agric., 1950, 1, 92.564 HAMENCE : THE DETERMINATION OF THE AVAILABILITY OF SURVEY OF AVAILABILITY OF NITROGENOUS FERTILISERS [Vol. 76 Since its publication this method has been applied in our laboratory to nearly all common nitrogenous fertilisers, and it is the purpose of t'his communication to describe and discuss the results that have been obtained. In the course of this work the practice of determining the nitrate nitrogen at the end of three fixed intervals of time has been maintained and experience has shown that this technique is valuable in that it gives a more complete picture of the decomposition taking Nitrate Nitrogen, mg Fig. 1 Nitrate Nitrogen, mg Fig. 2 place in the soil than could possibly be obtained by making only one determination of the nitrate nitrogen a t the end of 21 days.This supports the findings of Owen, Rogers and Winsor4; moreover, the curve obtained by plotting the production of nitrate nitrogen against time yields valuable information and the different types of nitrogenous fertilisers have each their own characteristic type of curve. For some slow-acting fertilisers the tests have been continued beyond the normal period of 21 days and the results of these extended tests are given. For the purpose of reporting the results, the fertilisers have been divided into classes; the classification is based partly on composition and partly on the origin of the materials, as follows- 1. Protein by-products. 2. Protein and bone by-products. 3. Treated protein materials. 4. Seed and plant residues.5. Excreta. 6. Synthetic compounds. 7. Other fertilisers. Protein by-products are taken as the first grsup because dried blood, the material used for comparison purposes, falls within it. In the results that foUow each group is treated separately. Besides the results of the availability tests, typical curves for the different materials are given and the results are discussed from the point of view of the composition of the fertiliser. PROTEIN BY-PRODUCTS- divided state, as in this test (see Table I). nature, and Fig. 1 shows the curves for dried blood and shoddy. Protein by-products all have a,similar availability in the soil when in the same finely The nitrate - time curves are also all similar in They indicate a steadyOct., 19511 NITROGEN IN NITROGEKOUS FERTILISERS 565 decomposition of the protein material with the rate of nitrification slowing as decomposition in the soil proceeds and reaches completion.Moreover, although the rate of nitrification slows, the curve does not become parallel to the time axis, as occurs with some fertilisers, TABLE I 1. PROTEIN BY-PRODUCTS Fertiliser Nitrogen, Availability per cent. Dried blood . . .. .. .. 14.00 100 (by definition) Hoof meal . . . . .. .. 15.10 118 Hoof meal . . .. .. . . 14.85 80 Hoof and horn meal . . .. . . 14.30 100 Shoddy . . .. . . .. . . 10.00 91 Gluten* . . .. .. .. .. 14.70 114 * Not a usual fertilising material, but included for comparison purposes. and if the time of the test is extended to 60 days nearly the whole of the organic nitrogen is recovered in the form of nitrate.It is interesting to note that gluten, a protein material, behaves in a similar manner. TABLE I1 2. PROTEIN AND BONE BY-PRODUCTS Fertiliser Nitrogen, Availability per cent. Meat and bone meal . . .. . . 8.57 Fish meal .. . . .. . . 10.67 Bone meal . . . . .. . . 4.19 Bone meal . . . . . . . . 3.75 Steamed bone flour . . .. . . 1-64 104 131 18 14 80 PROTEIN AND BONE BY-PRODUCTS- Fig. 2 shows typical curves for bone meal and for meat and bone meal. These results indicate that those fertilisers of this group in which the predominating material is protein have an availability similar to that of dried blood, and the nitrate - time curves are also similar to the dried blood curve. Bone meal, in contrast, presents a different picture, showing only a comparatively small availability and a correspondingly different nitrate - time curve.In other words the test shows bone meal to be a slow-acting nitrogenous fertiliser compared with dried blood and with meat and bone meal-thus confirming the accepted view of its slow-acting nature. TABLE I11 Table I1 shows the availability of the nitrogen in protein and bone by-products. RESULTS OF EXTENDED TESTS Nitrogen converted to nitrate in A \ Fertiliser 21 days, 39 days, 77 days. % % % Hoof meal . . .. .. .. .. 59 Bone meal .. .. .. . . 11 8 Plastic by-product, 22.2% of nitrogen . . Sewage sludge . . .. .. .. 9 77 19 10 10 In the studies of the availability of a slow-acting material like bone meal, much valuable information is gained by prolonging the tests beyond the normal 18 to 21 days.By extending the test for a further 21 days it is found that an increase in the rate of nitrification and con- sequently of the availability takes place, and this differentiates bone meal from very slow- acting materials, such as certain plastic by-products, which do not exhibit this increased rate of nitrification during the second period of 21 days (see Table 111). The extended period of time immediately shows a difference between bone meal and the other slow-acting materials.566 HAMENCE : THE DETERMINATION OF THE AVAILABILITY OF [Vol. 76 A t the present time the reason for the slow-acting nature of the nitrogen in bone meal is not by any means clear, but two possibilities present themselves; either (i) the protein, being embedded in the phosphate matrix, is protected by the calcium phosphate from the attack of micro-organisms, or (ii) the protein, being in the form of collagen, needs to be hydrolysed to gelatin before it is capable of being attacked by bacteria.Steamed bone meal presents an entirely different picture from bone meal in that it has a high availability, presumably owing to the steaming process rendering the protein more available for decomposition. The high nitrogen availability would appear to support the second possibility as being the more correct, as the steaming process would bring about hydrolysis of the collagen to gelatin; again, if the first hypothesis were the more probable, it would be expected that the nitrogen in steamed bone meal would be less available, as the nitrogen remaining in steamed bone meal is the residue after the more easily extractable protein has been removed.Some interesting results on the availability of the nitrogen in bone meal have been obtained by Long, Owen and Winsor,6 who showed that availability depends upon the age of the bone meal and the extent to which decomposition has occurred. # TABLE I’V 3. TREATED PROTEIN MATERIAL Fertiliser Bark tanned leather Chrome leather . . Treated leather meal Treated leather meal Casein waste . . Comb dust . . Keronikon . . Keronikon . . Nitrogen, Availability per cent. .. .. . . 5.07 . . . . .. 8.38 .. .. .. 6.67 .. .. . . 6-7 1 .. . . .. 13.38 .. .. .. 12.05 .. .. .. 15.03 .. .. .. 13.30 nil 8 46 47 30 53 53 46 TREATED PROTEIN MATERIAL- The results for treated and untreated leather waste have already been discussed in the original paper and are repeated here in Table IV for the sake of completeness.Typical curves for these materials are shown in Fig. 3. The considerable increase in availability as the result of treatment of leather wastes is immediately apparent from Table IV and Fig. 3. The treated products yield a substantial amount of readily available nitrogen in the first 7 days-more than the majority of other fertilisers-but the rate of breakdown soon slows up very considerably. They may thus be regarded as consisting of two parts, one a quick-acting fertiliser and the other slow-acting. TABLE V 4. SEED AND PLANT RESIDUES Fertiliser Nitrogen, Availability per cent. Castor meal . . .. .. .. 4.75 Castor meal . ... .. .. 5-47 Cotton-seed meal . . .. . . 5.62 Cotton-seed meal . . .. . . 5.74 Rape-seed meal .. .. .. 5.16 Cocoa shell . . .. .. .. 2.68 Dried grass meal . . .. .. 1.95 96 108 36 39 69 nil nil SEED AND PLANT RESIDUES- The materials in this group are mainly the residues remaining after the extraction of the oil from oil-bearing seeds. It will be seen from the results in Table V that castor meal has by far the highest avail- ability of the materials in this group. The availability is similar to that of dried blood and this fact probably accounts for the extensive use of castor meal as an organic nitrogenous fertiliser in past years.Oct., 19511 NITROGEN I N NITROGENOUS FERTILISERS 567 Curves for castor meal and cotton-seed meal are shown in Fig.4. This group of materials is interesting in that it shows a phenomenon that has not been referred to before in this work, namely, the non-availability of the nitrogen after 21 days in cocoa-shell meal and the dried-grass materials, which should break down easily in the soil. Nitrate Nitrogen, mg Fig. 3 Nitrate Nitrogen, mg Fig. 4 This result is merely a laboratory demonstration of the well-known phenomenon that often occurs when attempts are made to increase the organic matter in soils by the incorporation of straw or of a green crop. This practice often leads to the utilisation of the natural soil nitrate for the decomposition of the carbohydrate portion of the organic material. The substantial withdrawal of nitrate nitrogen from the soil by cocoa shell and dried grass is readily seen from Fig.5. \ \ \ \ I Cocoa Shel,l - ', Dried Grass - - - - \ I -I Ir 5 -10 Nitrate Nitrogen, mg Fig. 5. Nitrate absorbed from the soil Nitrate Nitrogen, mg Fig. 6568 HAMENCE THE DETERMINATION OF THE AVAILABILITY OF [Vol. 76 Fertiliser TABLE VI 5. EXCRETA Nitrogen, Availability per cent. Urea . . .. .. . . .. 46.60 170 . . 16.8 163 1.46 nil Bird guano . . .. . . Fresh horse-manure . . . . .. Sewage sludge . . .. . . . . 3.04 32’ Sewage sludge . . .. . . .. 2.74 34 Sewage sludge . . . . .. .. 2.14 14 EXCRETA- It will be seen from Table VI that there is a very wide difference in the availability of these different products, which represent excreta i n different forms. The nitrogen in urea is very rapidly broken down, and the figures show urea to be the most readily available of all nitrogenous fertilisers apart from the alkali nitrates.Bird guano similarly breaks down with extreme rapidity and behaves in many respects similarly to urea in that decomposition is so rapid that it appears to stimulate the natural formation TABLE VII 6. SYNTHETIC COMPOUNDS Fertiliser Nitrogen, Availability per cent. Hexamine . . .. .. .. 40.00 Cyanamide . . .. . . .. 16.04 Ammonium nitrate . . .. .. 35.00 Ammonium sulphate . . . . .. 21.00 163 84 120 164 of nitrate from the soil; moreover that this stimulation is only temporary and subsequently loss of nitrate takes place. These observations are in accord with the fact that the nitrogen in bird guano is mainly in the form of urea or ureides. They also afford an explanation of the great value that was placed on guano by the horticulturalists at the beginning of this century. Fresh horse-manure shows a strong initial absorption of soil nitrate by the residual carbohydrate material, so demonstrating one reason for the age-old custom of not using this type of fertiliser in its fresh condition.The curves for some of these materials are shown in Fig. 6. The availability of the nitrogen is very much less than that of urea-containing substances in this group, and reference to the curves in Fig. 6 shows that, although there is a small, fairly rapid, initial formation of nitrate, this in some instances diminishes by the absorption of nitrate presumably required for the TABLE VIII Sewage sludge presents a contrasting picture.7. MI SCELLANE ou s Fertiliser Nitrogen, Availability per cent. Hop manure . . .. .. .. 3.46 128 Peat . . . . .. .. .. 0.90 16 Humic acid* . . .. .. .. 2.00 12 * Contaminated with sand. decomposition of the other organic matter present in the sludge. Experiments in which the time has been extended beyond the normal 21 days show that the subsequent breakdown of the sewage sludge is extremely slow and therefore only a relatively small part of the nitrogen in the sludge is available for immediate plant growth. It seems possible that the very slow- acting nature of a substantial part.of the nitrogen in sewage sludge may account for its unpopularity in some quarters as a nitrogenous fertiliser. Clearly, in the process of sewage purification the readily decomposable and water- soluble nitrogenous compounds are washed out and.removed, leaving behind material having a humic and somewhat inert nature.Oct., 19511 KITROGEX IN NITROGENOUS FERTJLISERS 569 SYNTHETIC COMPOUNDS- ability, and follow the normal type of curve as shown in Fig. 7. employed as a fertiliser, behaves in the soil in a manner similar to urea. from the other members of the group in that it is slow-acting in the initial stages. The ammonium salts in Table VII show, as would be expected, high figures for avail- Hexamine, a compound not Cyanamide is different 0 5 10 15 Nitrate Nitrogen, mg Fig. 7 MISCELLANEOUS FERTILISERS- Most of the materials in this group call for some special attention. Hop manure-Hop manure frequently consists of spent hops suitably reinforced with concentrated fertilisers such as ammonium sulphate. The figures in Table VIII show that the nitrogen in hop manure is very readily available.It may be argued that this would be expected, as the main bulk of the nitrogen is in the form of ammonium salts; on the other hand, the fertiliser contains a substantial amount of organic matter and therefore an initial loss of the readily available nitrogen by decomposition of the organic matter might be expected. Peat and hzmic acid-Peat yields a small amount of available nitrogen and in that respect is similar to humic acid. Clearly, the accumulation of these substances in the soil is only possible if they are relatively inert in nature and decompose only slowly in the soil. ABSORPTION OF SOIL NITRATE DURING DECOMPOSITION This phenomenon is readily shown in this test by studying the rate of nitrate formation from protein-enriched bread and from the major constituents of this bread, namely, wheat starch and gluten.With wheat starch the nitrate nitrogen of the soil is completely removed or utilised by carbo- hydrate decomposition and even after 21 days the soil contains only insignificant traces of nitrates. With the protein gluten, decomposition is slow in starting, but having once started it proceeds rapidly. With bread itself the carbohydrate a t first removes soil nitrate, but once gluten decomposition gains pace the absorption of nitrate by the carbohydrate is more than compensated by the production of nitrate from the gluten, and free nitrate again appears in the soil.If we consider the nitrate balances at the end of 13 and 21 days, from the curves shown in Fig. 8, we obtain the results shown in Table IX. These results show that the calculated balance of nitrate from the individual gluten and starch curves agrees well with the balance of nitrate as determined from the bread. It follows therefore that the two reactions, viz., The nitrate formation with these materials is shown in Fig. 8.570 HAMENCE : THE DETERMINATION OF THE AVAILABILITY OF [Vol. 76 the decomposition of carbohydrate involving the absorption of nitrate and the decomposition of protein with accompanying nitrification proceed simultaneously in the soil. I Nitrate Nitrogen, mg Fig. S TABLE IX NITRATE BALANCES FOR BREAD AND ITS CONSTITUENTS After 13 days, After 21 days, g 0.0078 0.0072 0.0006 0*0005 Nitrate produced from gluten .. .. Nitrate absorbed by starch . . . . .. Free nitrate on balance . . .. . . Free nitrate found from decomposition of .. .. .. .. bread . . g 0.0089 0.0077 0.0012 0.0014 RAPID ROUTINE TEST The principal disadvantage of the method is the time it takes; as the method requires three weeks for completion, it is hardly justifiable suggest its adoption as a routine test in, say, the regulations of the Fertilisers and Feeding Stuffs Act. Clearly there is still room for a more rapid method, and search for such a method :has been made in our laboratory for some RESULTS OF PEPSIN DIGESTIBILITY TEST Fertiliser Dried blood . . .. .. .. Castor meal . . .. .. .. Casein waste . . .. .. .. Sewage ..Sewage .. .. Urea plastic . . .. .. .. Leather meal . . .. .. . . .. . . . . .. . . Total nitrogen, per cent. 14.0 5.47 12.75 1.72 2.74 19-46 6.13 Availability (by soil test) 100 10s 35 30 34 12 54 Nitrogen insoluble in pepsin, per cent. 0.44 0.60 6.00 1-33 2.20 3.00 3.15 Availability in pepsin 97 89 56 23 20 84 49 time, all possible sorting methods being compared with the results of the longer test that has just been described. So far, the most promising rapid test has been the one that is commonlyOct., 19511 NITROGEN IN NITROGENOUS FERTILISERS 571 used for the determination of available protein in feeding stuffs. This is a pepsin digestibility test. Procedwe-Dissolve 1 g of pepsin in 480 ml of water and 10 ml of 126 per cent. v/v hydrochloric acid. Add 1.6g of the finely ground fertiliser, mix thoroughly and incubate at blood heat for 24 hours.At the end of 24 hours add 10 ml of 12.5 per cent. v/v hydro- chloric acid and incubate for a further 24 hours. Filter off the insoluble matter, wash well with water and determine the nitrogen in the insoluble matter remaining on the filter-paper. As will be seen from these results for a number of fertilisers, the pepsin solubility test is a useful guide to the availability of the nitrogen, but unfortunately it breaks down seriously with the urea- formaldehyde type of plastic waste material that is now becoming available for fertilising purposes. It is conducted as follows. Resalts-By this test the results shown in Table X have been obtained. REFERENCES 1. 2. 3. 4. 5 .Hamence, J. H., J . Sci. Food Agric., 1950, 1, 92. “Official Methods of Analysis of the Association of Official Agricultural Chemists,” Association of Clark, K. G., and Gaddy, V. L., J . Ass. Off. Agric. Chew., 1950, 33, 480. Owen, O., Rogers, D. W., and Winsor, G. W., J . Agric. Sci., 1951, 40, 185. Long, M. I. E., Owen, O., and Winsor, G. W., J . Sci. Food Agric., 1951, 2, 125. Official Agricultural Chemists, Washington, 1950. DR. BERNARD DYER AND PARTNERS LONDON, E.C.3 DISCUSSION THE PRESIDENT congratulated Dr. Hamence on producing a most important paper on the availability of fertilisers. DR. 0. OWEN said he considered the technique was valuable in two ways. First it indicated the extent of conversion of the source of nitrogen into nitrate. Secondly, it indicated the rates a t which nitrogenous compounds decomposed, provided that periodic determinations were carried out. For this second purpose the work a t Cheshunt had shown that both nitrate and ammonia nitrogen should be determined, as their sum represented the decomposition in the early stages and ammonia was known to be a precursor of nitrate. Dr.Hamence’s results for cocoa shell suggested that the use of this material as a filler for quick-acting top dressings might defeat the object of such fertilisers in that the correct timing of a nitrogenous fertiliser for a rapidly growing crop was often critical. The results shown for horse manure were unexpected, as this material was considered to be so valuable in intensive horticulture. Similar work had been carried out on a range of urea - formaldehyde condensation products a t Cheshunt and some of the American conclusions had been confirmed. There still remained the problem of the fate of that part of the nitrogen that had not been converted into nitrate a t the end of an experiment.MR. D. D. MOIR enquired whether the author could give any information as to the degree of fineness of the various fertilisers, or whether the commercial articles had been used without any additional grinding. Since the availability would be dependent on the fineness of grinding this would appear to be important. DR. HAMENCE expressed his pleasure that Dr. Owen had been able to come to the meeting, as they had both been working independently on the problem of nitrogen availability for a number of years.In reply to Dr. Owen’s remarks, he said that, under the conditions in which the decomposition of the fertiliser was carried out with his test, it was his experience that no appreciable quantities of free ammonia were produced, and it was for this reason that he had confined himself almost entirely to nitrate determina- tions. Moreover, he had expressed the view in the earlier paper that, if ammonia accumulated in the system, the mechanism of nitrification was not functioning properly. He agreed with Dr. Owen that the results of this work had shown that great care must be taken when selecting an organic filler for a compound fertiliser. If the filler contemplated had the power of absorbing substantial quantities of nitrate from the soil, it was important to ensure that adequate supplies of readily available nitrogen were present to counteract this tendency.In connection with the speaker’s remarks on the subject of horse manure it was important to remember that the sample examined was fresh and that i t was unusual to use the material in this fresh condition in practice. Presumably this was one of the reasons for avoiding the use of the fresh material, and that one of the functions of the preliminary “aging” before use was to enable the readily available carbohydrate fraction to expend its nitrogen absorbing power before it was applied to the crop. In reply to Mr. Moir, Dr. Hamence said that all materials used in the test were in the same finely divided state. The sample, after having been ground as finely as possible in the mill, was weighed and then thoroughly ground with about 10 g of air-dried soil, together with 0.1 g of chalk.In this way i t had been found possible to reduce even very coarse and fibrous materials Indeed, this was part of the technique.572 HAMENCE [Vol. 76 that were otherwise difficult to grind in a laboratory mill to the same very finely divided state. This intimate mixture of soil and fertiliser was then mixed with the bulk of the soil that was used for the test. MR. HASLAM said he gathered that Dr. Hamence was surprised that, on treatment of urea - formaldehyde plastic waste with dilute acid and pepsin, relatively large amounts of nitrogenous substances passed into solution. It would occur in the absence of pepsin with most urea - formaldehyde preparations from the original urea - formaldehyde syrup to the moulding powder used in the preparation of the final moulded article.Simple treatment of most urea -formaldehyde preparations with weak acid would yield a comparatively large amount of soluble nitrogenous matter. DR. HAMENCE thanked Mr. Haslam for these very helpful observations, which provided an immediate answer to a problem that had perplexed him for some time, as he had always regarded urea - formaldehyde plastic as being an inert material and certainly not one that would have a water solubility. DR. K. A. WILLIAMS asked if the theobromine in cocoa shell had any effect on the bacteria of thesoil. DR. HAMENCE said that he doubted it very much, as nitrogenous compounds of the nature of theo- bromine were usually broken down rapidly in the soil.DR. E. C. WOOD asked if it were possible to draw up a balance sheet for soil nitrogen in all forms during the period of denitrification that preceded nitrification when fertilisers containing carbohydrate were added to the soil. Was the nitrogen withdrawn only temporarily, to re-appear a t a later stage as available nitrogen, or was there a net loss of nitrogen in the early stages-as ammonia, for instance? DR. HAMENCE replied that it appeared from the work they had carried out that the course of the decomposition would depend largely on the ratio of nitrogen to carbohydrate in the fertiliser and also .on the form in which the nitrogen was present. It was evident also that both the process of nitrification and of absorption of nitrate by the carbohydrate portion of the fertiliser went on simultaneously and that the proportion of free nitrate available for plant growth depended on the resultant of the two processes. As regards losses from the soil in the form of nitrogen, there was now fairly substantial evidence of such a phenomenon taking place in certain conditions and he thokght that, now a labelled isotope of nitrogen was available, more information on the point should soon be forthcoming. MR. R. F. MILTON asked if the apparent slow availability of certain organic fertilisers was not due to the “locking-up” in micro-organisms of nitrogen that became available later. DR. HAMENCE in reply said that he thought this a re,asonable explanation; a t the present time in his laboratory they were trying out a method that he thought might provide a measure of the nitrogen present in this form. In his view it was the only reasonable explanation of strong residual nitrate action observed some three or four years after the application of unusually heavy dressings of readily available organic nitrogenous fertilisers, and he hoped that the test now being developed would give a definite answer to this problem. That did not surprise him in the least.