Determination of ammonium,amides, and soluble carboxylates in plant tissues

Methods for extraction and determination of ammonium, amides and soluble carboxylates in plant tissues are compared and discussed. The procedure recommended involves extraction of finely ground plant tissues with 0.2 M formic acid, determination of ammonium and amides in the extract by overnight alkaline hydrolysis and distillation in Conway microdiffusion dishes, measurement of extracted ammonium using Carlson's rapid diffusion‐conductivity instrument, and estimation of soluble organic acids by titration after separation and purification with ion exchange resins. The method described is reasonably rapid, and gives complete recovery and highly reproducible results. The final solution of organic acids is suitable for direct chromatographic analysis.     

Use of an AutoAnalyzer for total nitrogen determination in plants

Abstract

An AutoAnalyzer was evaluated as a means of determining ammonium concentration in semi‐micro Kjeldahl digests of plant tissue. Results of the AutoAnalyzer determination agree closely with distillation‐titration results. Advantages of the AutoAnalyzer method include speed, precision, and release of labor. Only a small aliquot of each digest is utilized.     

Use of ammonia electrode for total nitrogen determination in plants

Abstract

An ammonia electrode was evaluated as a means of determining ammonium concentration in semi‐micro Kjeldahl digests of plant samples. Results of the ammonia electrode determination agreed closely with distillation and titration results. Advantages of the electrode method include speed, precision, increased safety, simplicity and the fact that only a small aliquot of the digest is used.     

A simple method for determination of ammonium in semimicro‐Kjeldahl analysis of soils and plant materials using a block digester

Abstract

A simple method for determination of ammonium in semimicro‐Kjeldahl analysis of soils and plant materials using a Tecator or Technicon 40‐tube block digester is described. It involves use of an inexpensive steam distillation apparatus that permits direct distillation of ammonium from the tubes used for Kjeldahl digestion in 40‐tube block digesters. The method is rapid and precise, and it gives results that agree closely with those obtained by the customary method of ammonium N analysis involving transfer of the Kjeldahl digest before distillation.     

Amino acid interference with ammonium determination in soil extracts using the automated indophenol method

Abstract

The formation of a colored indophenol complex is commonly used as a quantitative measure of the ammonium content of soil extracts. The potential interference with ammonium determination from co‐extracted amino acids was examined. The extent of color development was examined for 22 amino acids by subjecting pure solutions to ammonium determination by both the indophenol method and steam distillation. Apparent detection of amino acid as ammonium ranged from 0 to 94 % of total nitrogen for the indophenol procedure, whereas steam distillation resulted in little apparent ammonium recovery. With the exception of threonine, the extent of color development was inversely related to amino acid molecular weight. The range in recoveries for the indophenol procedure suggests both size and composition of the co‐extracted amino acid pool is important in determining the extent of interference.

Significantly (p=0.001) greater estimates, averaging 0.4 μg mL^‐1, were found in indophenol estimates of mineral‐N content of moist, fresh soil samples. Air drying, oven drying or chloroform fumigation significantly increased the difference (0.3 ‐ 0.7 μg mL^‐1) in estimates of ammonium content. At 10: 1 extract: soil ratios this could cause ? to be overestimated by 3–7 μg g^‐1soil. The increased interference was attributed to a release of amino acid as a result of pretreatment. The difference between distillation and indophenol estimates of ammonium content of 0.5 M K₂SO₄was found to be dependent upon ammonium content. The use of procedures employing a distillation step (manual or automated) is recommended to avoid amino acid interference when precise NH₄+‐N determinations are needed on dried or fumigated samples.     

Comparison of chemical methods of assessing potentially available organic nitrogen in soil

Abstract

We recently developed two rapid and precise chemical methods of assessing potentially available organic N in soils. One method involves determination of the ammonia‐N produced by steam distillation of the soil sample with pH 11.2 phosphate‐borate buffer solution for 8 min. The other involves determination of the ammonium‐N produced by treatment of the soil sample with 2M KCl solution at 100°C for 4 hours. Studies using 33 Brazilian soils showed that the results obtained by these methods were highly correlated with those obtained by anaerobic and aerobic incubation methods of assessing potentially available organic N in soil.

The two methods were further evaluated by applying them to 30 Iowa soils and by comparing their results and those obtained by other chemical methods with the results of the incubation methods considered to be the best laboratory methods currently available for assessment of potentially available organic N in soil. The chemical methods used included the acid KMnO₄ method, the alkaline KMnO₄ method, the CaCl₂‐autoclave method, and the NaHCO₃ UV method. The incubation methods used involved determination of the ammonium‐N produced by incubation of the soil sample under anaerobic conditions for 1 week or determination of the (ammonium + nitrate + nitrite)‐N produced by incubation of the sample under aerobic conditions for 2 and 12 weeks. The data obtained showed that the results of the two chemical methods evaluated were highly correlated with those obtained by the incubation techniques used for comparison and that the correlations observed with these two methods were higher than those observed with the previously proposed chemical methods. It is concluded that these two rapid and simple methods are the best chemical methods thus far developed for laboratory assessment of potentially available organic N in soil.     

A rapid and precise method for routine determination of organic carbon in soil

Abstract

A simple method for routine determination of organic carbon in soil by a modified Mebius procedure is described. It involves (a) digestion of the soil sample with an acidified dichromate (K₂Cr₂O₇‐H₂SO₄) solution for 30 minutes in a Pyrex digestion tube in a 40‐tube block digester preheated to 170°C and (b) estimation of the unreacted dichromate by titration of the cooled digest with an acidified solution of ferrous ammonium sulfate with use ofN‐phenylanthranilic acid as an indicator. The method is more rapid and precise than the Mebius procedure commonly used for routine analysis of soils for organic carbon, and the only equipment required for its use is equipment now commonly used for routine Kjeldahl analysis of soils for total nitrogen.     

A manual colorimetric procedure for measuring ammonium nitrogen in soil and plant Kjeldahl digests

Abstract

The measurement of NH4⁺‐N in soil, and plant digests is one of the greatest needs in laboratories conducting agricultural and environmental research. Many laboratories do not have access to automated equipment for colorimetric analysis of soil and plant digests. The objective of this research was to modify an automated colorimetric analysis procedure for determining NH₄+‐N in soil and plant digests for manual use, and compare the proposed technique with the standard distillation‐titration technique. The modified procedure is based on the color reaction between NH₄ ⁺‐ and a weakly alkaline mixture of Na salicylate and a chlorine source in the presence of Na nitroprusside. Wavelength scans indicated a very well defined peak for determinations at 650 nm. Time scans showed that color development in the manual procedure was rapid, 12 to 40 minutes depending on temperature, and that the color development remained stable for at least 120 minutes. Regression analysis of the results from 18 soil and 20 plant tissue sample determinations by distillation‐titration and the proposed method indicated NH₄ ⁺ ‐N recoveries of 99% or higher. The results obtained using the colorimetric procedure were very similar to the values obtained by distil ling and titrating the digests for both soil and plant samples as indicated by the large coefficients of determination (R² = 0.99).     

A rapid steam distillation method of assessing potentially available organic nitrogen in soil

Abstract

A rapid steam distillation of assessing potentially available organic nitrogen in soil is described. It involves determination of the ammonia‐N produced by steam distillation of the soil sample with pH 11.2 phosphate‐borate buffer solution for 8 min. The method is simple and precise, and its results are not significantly affected by air‐drying or air‐dry storage of the soil sample before analysis. It is well suited for use in soil testing laboratories because it does not require extraction, filtration or transfer steps. Studies using 33 Brazilian soils showed that the results obtained by this method were highly correlated with those obtained by aerobic and anaerobic incubation methods of assessing potentially available organic nitrogen in soil.     

Phosphate interference in the Devarda's alloy reduction method for nitrate

Abstract

Phosphate interfered markedly with a Devarda's alloy distillation method for the determination of nitrate. Only 9% of added nitrate was recovered by this method when 200 mg P was present. This interference can be overcome by the addition of calcium ions, to precipitate the phosphate, and an extended distillation time.     

Determination of ammonium n and nitrate n in acid permanganate solution used to absorb ammonia,nitric oxide,and nitrogen dioxide evolved from soils

Abstract

Simple steam distillation methods are described for determination of ammonium N and nitrate N in acid KMnO₄ solution used to absorb NH₃, NO and NO₂ evolved from soils. They involve use of MgO for distillation of ammonia and of FeSO₄, Ag₂SO₄, and MgO for reduction of nitrate to ammonia. The methods are rapid and precise, and they permit nitrogen‐15 analysis of NH₃‐N and (NO + NO₂)‐N evolved from soils.     

Oxidation and fate of chromium in soils

Abstract

The ammonia electrode method was used to characterize soil organic nitrogen. Amide-N, α-amino-N and hexosamine-N were estimated by measuring the ammonium-N derived from each organic-N with an ammonia electrode after hydrolysis of standard compounds followed by certain treatments. Each organic-N was recovered quantitatively from each standard compound in the presence and absence of soil. Twenty soil samples were analyzed for the above forms of organic nitrogen by both the distillation and electrode methods. The results obtained by the ammonia electrode method for each form of organic nitrogen agreed closely with those obtained by the distillation method. The electrode method was preferred for subsequent soil studies since it is simple and sensitive.     

Determination of organic acids and phosphate in soil aqueous extracts by capillary zone electrophoresis

Abstract

Organic acids and phosphate were determined in soil aqueous extracts by capillary zone electrophoresis using indirect UV detection. The electrolyte system was 10 mM sodium benzoate with 0.5 mM tetradecyltrimetyl‐ammonium bromide as flow modifier at pH 4.0, pH 4.5, or pH 5.0. This methodology was adequate to determine organic acids and phosphate in soil samples, but the major inorganic anions interfered in the determination. In all the samples of soil extracts, the presence of phosphate was detected. Acetate was found in most of the samples and lactate and formate in some of them.     

Technique of soil sample pretreatment for analysis of 15N:14N isotope ratio

Abstract

The technique of simultaneous quantitative determination of mineral N soil forms (nitrates, exchangeable and non‐exchangeable ammonium, and total amount of these compounds) and sample pretreatment for the analysis of ¹⁵N:¹⁴N ratio is suggested. The technique is based on the selective association of NH₄ ⁺‐ions into indophenol complex and subsequent ethyl‐acetate extraction of this complex from solution. The mineralization of indophenol is carried out in alkaline medium with simultaneous NH₃ distillation into H₂SO₄ titrant. The application of given technique allows us to shorten significantly the time required for analysis and to increase the accuracy of analytical determination.     

Preparation of Ammonium‐15N and Nitrate‐15N Samples by Microdiffusion for Isotope Ratio Analysis by Optical Emission Spectrometry

Abstract

A diffusion method for the preparation and measurement of ¹⁵N abundance of ammonium and nitrate in KCl extracts of soil using optical emission spectrometry (OES) was compared with conventional continuous flow isotope ratio mass spectrometry (IRMS). There were highly significant correlations between the values obtained by using OES and IRMS. The 99% confidence interval of the intercept included the value 0 and the 99% confidence interval of the slope included the value 1 for both nitrate and ammonium measurements, suggesting that the results from the two methods did not differ significantly. In another experiment, ¹⁵N values of nitrate and ammonium from soil extracts prepared by using the standard distillation procedures for OES were compared against the microdiffusion preparation method. Again, there was a highly significant correlation between the values: the 95% confidence interval of the intercept included the value 0, and the 95% confidence interval of the slope included the value 1, again suggesting that the two methods did not differ significantly. It was concluded that the diffusion technique is an appropriate and simple method of sample preparation for inorganic N analysis of KCl extracts using OES.     

Mechanisms behind soil N dynamics following cover restoration in degraded land in subtropical China

Purpose

Nitrogen (N) is an important nutrient for re-vegetation during ecosystem restoration, but the effects of cover restoration on soil N transformations are not fully understood. This study was conducted to investigate N transformations in soils with different cover restoration ages in Eastern China.

Materials and methods

Soil samples were collected from four degraded and subsequently restored lands with restoration ages of 7, 17, 23, and 35 years along with an adjacent control of degraded land. A ¹⁵N tracing technique was used to quantify gross N transformation rates.

Results and discussion

Compared with degraded land, soil organic carbon (SOC) and total N (TN) increased by 1.60–3.97 and 2.49–5.36 times in restoration land. Cover restoration increased ammonium and nitrate immobilization, and dissimilatory nitrate reduction to ammonium (DNRA) by 0.56–0.96, 0.34–2.10, and 0.79–3.45 times, respectively, indicating that restoration was beneficial for N retention. There were positive correlations between SOC content and ammonium and nitrate immobilization and DNRA, indicating that the increase in soil N retention capacity may be ascribed to increasing SOC concentrations. The stimulating effect of SOC on ammonium immobilization was greater than its effect on organic N mineralization, so while SOC and TN increased, inorganic N supply did not increase. Autotrophic and heterotrophic nitrification increased with increasing SOC and TN concentrations. Notably, heterotrophic nitrification was an important source of NO₃^??N production, accounting for 47–67% of NO₃^??N production among all restoration ages.

Conclusions

The capacity of N retention was improved by cover restoration, leading to an increase in soil organic carbon and total N over time, but inorganic N supply capacity did not change with cover restoration age.

     

Automated determination of trace amounts of phosphate in soil extracts using malachite green

Abstract

A sensitive and simple automatic continuous‐flow determination of orthophosphate at trace levels has been worked out. The detection limit was lower than 2 μg/L. The interferences due to silicate and dissolved organic matter in soil extracts have been eliminated.     

Determination of (Nitrate + Nitrite )‐N in alkaline permanganate solutions

Abstract

The quantitative reduction of nitrate in an acid medium with reduced Fe was applied to the alkaline permanganate solution used to absorb NO and NO₂ evolved from soils during denitrification reactions. The method involves addition of H₂SO₄ to acidify the solution and ensure oxidation of nitrite to nitrate, and treatment with reduced Fe at 100°C to reduce nitrate to ammonium. The solution is made alkaline with NaOH and ammonium determined by standard distillation procedures. It is simple and precise, and applicable to nitrogen isotope ratio analysis of NO and NO₂ evolved from soils.     

Arylamidase Activity as an Index of Nitrogen Mineralization in Soils

Abstract

The enzyme arylamidase [EC 3.4.11.2] catalyzes the hydrolysis of N‐terminal amino acids from arylamides. Because it has been proposed that this enzyme may play a major role in nitrogen (N) mineralization in soils, studies were carried out using short‐term laboratory incubations under aerobic and anaerobic conditions and chemical hydrolysis of soil organic N to assess the N mineralization in a range of 51 soils from six agroecological zones of the North Central region of the United States. The enzyme activity was assayed at its optimal pH value. With the exception of the values obtained for field‐moist soils incubated under anaerobic conditions, the amounts of N mineralized by all the biological and chemical methods studied were significantly correlated with arylamidase activity, with r values of 0.54*** for the amounts of inorganic N produced under aerobic incubation, of 0.44** for anaerobic incubation of air‐dried soils, of 0.53*** and 0.55*** for the amounts of ammonium (NH₄ ⁺)‐N released by steam distillation with PO₄‐B₄O₇ for 4 and 8 min, respectively; and of 0.49*** and 0.53*** for the amounts of NH₄ ⁺‐N released by steam distillation with disodium tetraborate (Na₂B₄O₇) for 4 min or 8 min, respectively. The amounts of N extractable with hot potassium chloride (KCl) were most significantly correlated with arylamidase activity (r=0.56***). Arylamidase activity was significantly correlated with organic carbon (C) (r=0.49***), organic N (r=0.55***), and fixed ammonium (NH₄ ⁺)‐N (r =0.42**).     

Determination of Bicarbonate in Substrate Extracts Using Back‐Titration and the Gran Plot Approach

Abstract

The back‐titration procedure, associated with the Gran Plot method to detect the process end point, was compared with other volumetric procedures for the potentiometric acid‐base titration determination of bicarbonate and was explored as a tool for the determination of the bicarbonate content in substrates used as growing media. This procedure was compared with the second derivative approach as well as with continuous titration with standard HCl on the extract solution (“direct measurement”), until reaching pH 4.0. By using known standard samples, it was noted that the results obtained with the second derivative method were both lower and inconsistent, because of the high dilution of both the sample solution and the titrant. On the other hand, the continuous direct titration (until pH 4.0) presents positive errors due to the presence of dissolved CO₂ in the titrated solution. Although comparable in precision, the best performance and accuracy were observed for the back‐titration with the Gran Plot approach. These aspects, associated with its low cost and inherent operational simplicity, permit us to suggest the Gran Plot method as a reliable option for routine determination of bicarbonate in substrate extracts and related materials.