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Research Article

Influence of Nitrification Inhibitor on Transformations and Recovery of N in Grassland Soil

M. K. Abbasi and W. A. Adams
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The recovery of both mineralized and applied N in soil-plant system is low throughout the world. Leaching, denitrification and NH3 volatilization are thought to be the major N loss mechanisms responsible for low N recovery. The aim of this study was to evaluate the effect of nitrification inhibitor nitrapyrin on N transformations, recovery and to prevent formation of -N which could be lost by leaching and denitrification. Intact soil cores were collected from a grassland field, incubated under aerobic conditions at 20°C with 70 percent WFPS. Amendments used were plus or minus and plus or minus nitrapyrin. Nitrapyrin was used as a nitrification inhibitor. The fate of mineralized and added , accumulation of -N and apparent recovery of applied N were examined during the study. In the absence of nitrpyrin, more than 60 percent of the -N initially present was lost within 42 days period. The apparent recovery of applied N decreased significantly and more than 50 percent of applied N being lost from mineral pool. Accumulation of 60 mg -N kg–1 soil in the surface 0-2.5 cm layer indicated that nitrification was an active process in the system. Addition of nitrapyrin decreased and delayed the extent of disappearance, inhibited nitrification by approximately 73 percent and improved N recovery by 36 percent.

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M. K. Abbasi and W. A. Adams, 2000. Influence of Nitrification Inhibitor on Transformations and Recovery of N in Grassland Soil. Pakistan Journal of Biological Sciences, 3: 812-815.

DOI: 10.3923/pjbs.2000.812.815



The yield potential of crops and productivity of forage species are often constrained by the supply of nitrogen (N). To overcome this limitation, N fertilizers are commonly applied to grasses, important food and cash crops. It has been suggested (Garret, 1993) that at least half the productivity of modern temperate grassland systems is due to the fertilizer N input. Supply of fertilizer N not only increases primary plant and subsequent animal production, but also allowed farmers a good deal of flexibility in the management of their enterprises. According to Munro and Davies (1973), in all temperate grassland regions N supply and pasture production are closely linked and yield is generally proportional to the supply of fertilizer N.

Nitrogen fertilizer is an expensive input and in many trials in Australia less than 50 percent of applied N is recovered in the crop and soil with the remainder being lost (Freney et al., 1993; Rochester et al., 1996). The apparent recovery of N applied to grass grown in the United Kingdom varies widely, but a value in the range of 50-70 percent is frequently observed (Morrison et al., 1980). A recovery of less than 45 percent from a single application of N fertilizer was reported by White (1985) due to abnormal weather conditions causing a large leaching or gaseous loss of N. Under wet conditions very low recoveries of applied N under 40 percent were reported by Van der Meer (1982). Fertilizer N recovery in plants and soil under upland conditions in India rarely exceeds 50 percent (Patra et al., 1992). Increasing the amount of N applied does not increase the amount of N available to the crops, because the fertilizer N is lost before it can be assimilated by the crop (Freney et al., 1992).

Most of the fertilizer N applied to soils is largely in the form of NH4 or NH4‾ producing compounds such as urea and usually oxidized quite rapidly to nitrate. The NO3 formed through nitrification of added N by soil microorganisms is susceptible to loss by leaching and denitrification and may contribute to environmental and surface and ground waters pollution. Thus nitrification acts as a key process in determining fertilizer use efficiency by crops as well as N losses from soils. The adverse effects of oxidation of to by the nitrifying microorganisms in soil have stimulated research to find compounds that can be used to retard nitrification of fertilizer N. Nitrification inhibitors have been used in agriculture to improve fertilizer efficiency and crop yields and minimize denitrification and/or leaching losses of , by delaying the biological oxidation of to NO2 and thus reduces the availability of and maintaining applied fertilizer N in the soil as -N (McTaggart et al., 1997). A number of compounds have been shown to inhibit the nitrification process in soil and the most commonly used is nitrapyrin (2-chloro-6-) trichloromethyl) pyridine.

The aim of this investigation was to determine the effect of nitrification inhibitor nitrapyrin on N transformations and its impact on recovery of applied N in grassland soil under laboratory conditions.

Materials and Methods

The site, which was almost level and in permanent pasture, situated adjacent to Blaendolau playing fields about 1 km from Soil Science Unit, University of Wales Aberystwyth, UK. The soil was a Dystric Eutrochrept and classified as Conway series (Grid Ref. SN 597804) as described in detail in an earlier study (Abbasi and Adams, 1998).

Fig. 1:
Effect of nitrification inhibitor nitrapyrin on the recovery of N applied to intact soil cores during 42 days incubation. Vatical bars indicate least significant difference (LSD) between with and without nitrapyrin treatments at different time intervals.

Some physical and chemical characteristics of the soil used are presented in Table 1. Seventy intact soil cores (7.0 cm dia. x 7.5 cm deep) were collected according to the method described in Abbasi and Adams (1998).

Table 1:Some chemical and physical properties of the soil used during the study Depth (cm)

Table 2:Effect of nitrification inhibitor nitrapyrin on the concentration of -N and -N in control soil without added N at different depth over a period of 42 days
*LSD represent the least significant difference (p≤0.051 appropriate to comparisons between the values within each column and rows

Table 3:Effect of nitrification inhibitor nitrapyrin on the concentration of -N and -N in added soil at different depth over aperiod of 42 days
*LSD represent the least significant difference (p<0.05) appropriate to comparisons between the values within each column and rows

The moisture content of soil cores held within brass sampling rings, was adjusted at 70 percent WFPS by saturating and then equilibrating at -5 kPa over 3 days on a tension table. The soil cores were arranged into four treatments i.e. T1 = control, T2 = control with nitrapyrin, T3 = amended cores without nitrapyrin and T5 = NH4+ amended cores with nitrapyrin. Five ml solution of [NH4)2HPO4)] at a rate equivalent to 200 mg N kg–1 (on moist soil basis) and 2 ml solution of 2-chloro-6- (trichlorornethyl) pyridine known commonly as nitrapyrin at a rate of 100 mg kg–1 soil were uniformly applied on cores surface. Cores were placed in an incubator at 20°C in the dark for 42 days. Small amount of distilled water was added after every two days if necessary to restore its initial weight.

Triplicate cores from each treatment were removed from the incubator at the beginning and after 7, 14, 28 and 42 days of incubation and sectioned into 0-2.5, 2.5-5.0 and 5.0-7.5 cm layers. Each section was broken down separately and mixed thoroughly. The concentration of total mineral N and -N in soil samples were determined by shaking 40 g sub-samples of fresh soil for 1 h with 200 ml of 2 M KCl followed by steam distillation with Mg0 and Oevarda's alloy and titration (Keeney and Nelson, 1982). -N was determined by subtracting -N from total mineral N. Total N in the soil was determined following the Kjeldahl method of Bremner and Mulvaney (1982). Organic matter was estimated as weight loss on ignition at 400°C (Ball, 1964). Soil pH was measured in a water suspension (1:1 v/v). Particle size distribution was determined by the pipette method described by Avery and Bascomb (1974). Soil moisture content was determined gravimetrically after drying of sub-samples at 105°C for 24 h. Water-filled pore space (WFPS), synonymous with relative moisture content, was calculated following the method of Aulakh and Bijay-Singh (1996). WFPS = [(gravimetric water content x soil bulk density/ total soil porosity)], where total soil porosity --- [1-(soil bulk density/2.65)1 and 2.65 was the assumed particle density of soil. All data in Table 2, 3 and Fig. 1 were statistically analysed by multifactorial analysis of variance (ANOVA) using the software package StatGraphics Manugistics Inc (1992).

Results and Discussion

Effect of inhibitor on mineral N of the control soil: Initially at the start of the experiment the concentration of in the control soil without nitrapyrin was in the range of 8-24 mg kg–1 soil with greater concentration in the surface 0-2.5 cm layer. Table 2 indicated that with one exception at day 42, did not show significant changes (p<0.05) either with time or depth. Thus changes in -N did not show any consistent pattern in control soil throughout the incubation. It was, however, clear that the surface 0-2.5 cm contained 2-3 times more -N than the other two layers, The higher level of -N in the surface layer attributed the variation in mineralization potential within depth which may be due to more labile organic N compounds, a higher level of microbial biomass and their activity and decrease organic C with depth which would limit the microbial populations from mineralizing organic N (Bolton et al., 1990). In the soil to which nitrapyrin was added, a significant (p≤0.05) increase in NH4+ concentration was observed both with time and depth (Table 2). Values were generally higher in the 0-2.5 cm layer compared to the 2.5-5.7 cm layers. Over 42 days the concentration of -N in the 0-2.5 cm layer of nitrapyrin added soil was 2 fold greater than the soil without nitrapyrin. This showed that in the soil without nitrapyrin a substantial amount of -N resulted from the mineralization of organic N disappeared/lost from the system as observed in our previous study (Abbasi and Adams, 1998). The pattern of changes in -N in the control soil with and without nitrapyrin was similar to that of -N (Table 2). Results indicated that the changes in -N during incubation were dependent mainly on the presence and availability of . A maximum of 15 mg -N kg–1 was accumulated in the surface 0-2.5 cm of soil without nitrapyrin indicating that the soil at 70 percent WFPS exhibited a substantial nitrification potential. However, even within this moisture range, chances of loss of accumulated -N through denitrification or leaching are there so, determination of exact rate of nitrification under such conditions became difficult. In the soil that received the inhibitor, -N was never exceeded 2 mg kg–1. The lower concentration in T2 compared to the T1, treatment showed that inhibitor was effective in inhibiting the oxidation of which increased the level of in the soil.

Effect of inhibitor on mineral N of N added soil: In the soil without nitrapyrin, -N was lost continuously from the surface 0-2.5 cm layer from the time of application till the end (Table 3). About 60 percent of -N initially present was lost within 42 days of incubation. In lower layers concentration of -N increased to some extent but the difference within incubation periods was not significant. Similar results were also found in our previous study (Abbasi and Adams, 1998). However, the extent of loss of -N in this study was less because of the differences in moisture level between the two experiments. Results revealed that when moisture contents were above 80 percent WFPS, 84 percent of initially present was lost from the mineral N pool compared with 60 percent loss in the present study having 70 percent WFPS. This showed that moisture is an important factor in N recovery and loss. In the soil cores to which nitrapyrin was added, the concentration of in the 0-2.5 cm layer decreased slowly with time but the rate and amount of disappearance was much smaller and a substantial amount was left till the end of incubation. Most of the disappeared from the surface layer moved down to the lower layers as a significant (p>0.005) increase in -N was found in 2.5-7.5 cm layers. Comparison between the two treatments revealed that nitrapyrin had a significant effect on the persistence and availability of , concentration of the inhibitor treatment was significantly (p<0.005) greater than that in the treatment without nitrapyrin. In the soil without nitrapyrin, after taking into account the -N in control soil, 55 percent of the added -N was unaccounted for at the end of study relative to 10 percent of the nitrapyrin added soil. Results showed that nitrapyrin proved to be very effective in retarding oxidation of and thus allowed N to remain in the soil for longer period for crop use. These results were in agreement with previous findings of Freney et al. (1992); Chen et al. (1994) and Abbasi and Adams (1998).

The concentration of -N in the soil without nitrapyrin increased slowly during the first 14 days of incubation (Table 2). Thereafter, concentration increased significantly (p≤0.005) and a maximum of 60 mg -N kg–1 was accumulated at day 28 in the surface 0-2.5 cm. The increase in -N in surface layer of N added soil compared with the control one indicated that nitrifibation was occurring. The pattern of increase in -N in lower layers was similar but the rate and amount of increase was significantly less compared with the surface 0-2.5 cm. The effect of depth on nitrification can be anticipated, since the supply of from organic N mineralization and from added N decreased with depth and depth may also affect nitrifiers population (Hadas at al., 1986). The accumulation of -N in the soil of the inhibitor treatment was significantly (p≤0.05) lower than that in the soil without nitrapyrin and a maximum of 13 mg -N kg was found in the 0-2.5 cm at day 28. On an average the NO3, concentration of the added soil without nitrapyrin was greater at least 5-fold relative to soil with nitrapyrin. It revealed that nitrapyrin became effective in blocking biological oxidation of to NO‾2 and thus reduces the availability of -N and maintaining applied fertilizer N in the soil as -N.

Effect of inhibitor on recovery of applied N: Data regarding the recovery of applied N (Fig. 1) showed that in the soil cores without nitrapyrin added N was lost continuously after day 7 till the end of incubation. By that time, 53 percent of the N initially present was lost from the system. The rate of N loss was greatest at day 14 and 42, equivalent to more than 1 kg–1 day–1. By contrast, 17 percent of the applied N was unaccounted for from the soil cores treated with nitrapyrin. Taking these losses into account, over 42 days about 83 percent of the added N was recovered in the nitrapyrin treatment compared with 47 percent recovery in soil cores without nitrapyrin. Nitrapyrin increased the apparent recovery of applied N and of the total amount of N applied 36 percent more N was recovered relative to soil without nitrapyrin. Previous work had shown that ammonia volatilization and leaching were not important pathways for N loss (Abbasi et al., 1997). Consequently it was concluded that major losses were due to denitrification of accumulated -N after nitrification of added or mineralized and that the use of nitrification inhibitor reduced production and therefore denitrification.

Nitrogen is usually the plant nutrient most limiting in arid and semiarid ecosystems that have a limited input of N and must rely on a controlled and efficient N cycle for continued plant growth and ecosystem function (Bolton et al., 1990). The fact that nitrapyrin conserved for a considerable time and improved the recovery of applied N showed that it can be used and practiced in Pakistan agriculture as welt. A substantial area of Pakistan is under rice where denitrification and movement and leaching might be major mechanism of N loss. Similarly, under normal agriculture, use of heavy irrigation after fertilizer application created conditions conducive to denitrification. Under such circumstances use of nitrification inhibitors can be proved very effective to increase the fertilizer use efficiency as well as improved the apparent recovery of applied N both in crops and soil. However, in Pakistan because of high soil pH and temperature, reduced nitrification could also increase N losses via NH3 volatilization. To obtain full benefits from inhibitors, techniques are needed to predict the optimum time and methods for their application.


The authors would like to thanks the Government of Pakistan for financial support, the University of Jammu & Kashmir Muzafferabad for nominating the senior author for post graduate scholarship.

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