Indian Journal of Fertilisers, Vol. 14 (5), pp.32-51 (20 pages)
Soil Tests for Micronutrients : Current Status and Future Thrust S.P. Datta1, Mahesh C. Meena1, M. Barman1, D. Golui1, R. Mishra1 and A.K. Shukla2 1Division of Soil Science and Agricultural Chemistry, ICAR-Indian Agricultural Research Institute, New Delhi 2ICAR-Indian Institute of Soil Science, Bhopal, Madhya Pradesh Abstract The range between deficiency and toxicity of micronutrient elements is very narrow, which underlines the importance of precise estimation. It appears that use of short sequential fractionation scheme for micronutrient cations may be practical way of solid phase speciation. A considerable progress has been made in assessing the fraction of micronutrients except Mo in soil for providing insight into their forms, availability and association with different soil constituents. Over the years, several extractants, like dilute acids and salt solutions have been used to assess available micronutrient status of soil. Although use of dilute acid does not have sound theoretical background, huge database has, however, been generated over the years indicating the close agreement between micronutrient content in soil and plant, particularly in acid soils. On the other hand, salt solutions have been found to be effective over a wide range of soils in respect of reaction (pH). Development of DTPA (diethylene triamine penta acetic acid) soil test brought revolutionary changes in soil testing for micronutrient cations (zinc, copper, iron and manganese), which have sound theoretical background. By and large, general validity of DTPA soil test for zinc and copper is evident, whereas, this soil test does not hold good for manganese in some cases and iron in general. In limited studies, DTPA soil test was found to be more efficient than Sr(NO3)2 and Ca(NO3)2 for assessing nickel deficiency in soil. For boron, salicylic acid and manitol-CaCl2 have shown potential in acid and alkaline soils, respectively. In case of molybdenum, Grigg’s reagent has been used. The tediousness of this method is reflected in the meagre number of soil samples analysed so far across the country. There is an urgent need to develop simple, rapid and reproducible soil test method for analysing the minute quantities of available Mo often encountered in the soil. Commonly used extractants like DTPA and EDTA (ethylene diamine tetra acetic acid) interfere with the estimation of micronutrient cations with ICP-MS (inductively coupled plasma mass spectrometer) in soil extracts. In most cases, scant information is available on critical levels of extractable micronutrients in soils. Future lines of work are suggested on various aspects of soil test methods of micronutrients. Key Words: Micronutrients, soil test methods, advanced techniques, critical limits
Introduction The term ‘micronutrients’ represents some essential nutrients that are required in small quantities for the normal growth and development of plants. These include zinc (Zn), copper (Cu), iron (Fe), manganese (Mn), nickel (Ni), boron (B), molybdenum (Mo) and chlorine (Cl). Importance of micronutrients has been realized during the past four decades when widespread micronutrient deficiencies, particularly of Zn and B, were observed in most of the soils in our country, especially under intensive agriculture (Katyal, 2018). Since the very inception of Green Revolution, introduction of high yielding varieties, extension of irrigated areas and use of high analysis micro-nutrient-free NPK fertilizers particularly under intensive agriculture have increasingly catalysed the depletion of finite reserves of soil micronutrients, leading to the occurrence of widespread and
[email protected]
multi-micronutrient deficiencies. According to the latest reports of All India Coordinated Research Project on Micro- and Secondary Nutrients and Pollutant Elements in Soils and Plants (AICRP-MSPE), 36.5, 23.2, 12.8, 4.2 and 7.91 of soil samples across the country are deficient in Zn, B, Fe, Cu and Mn, respectively (Shukla et al., 2018). Currently, no such information is available on the status of available Mo. The deficiency of micronutrients causes several physiological diseases and disorders, which reduce the yield of crops. Micronutrients are not only important for better crop productivity, but are also essential for sustaining human and animal health. Among micronutrients, Zn and Fe have very important bearing on human health (Rattan, 2017). On an average, Zndeficiency affects one-third of World’s population ranging from 4 to 73% in different countries (Hotz and Brown, 2004). It is reported that over 2 billion people throughout the world are affected Indian Journal of Fertilisers, May 2018 32
by Fe deficiency (Welch and Graham, 1999). A sizable portion of Indian population, particularly the children and women, has also been suffering from the deficiency of these two elements. Positive responses of crops to micronutrient application, particularly on deficient soils, obtained in a huge number of experiments underline the importance of accretion of micronutrients from external sources. First and most important step in management of micronutrients is the correct assessment of their available status in the soil. Because, range between deficiency and toxicity limits of micronutrient in soil is narrow for most of the crops, it is imperative to go in for cautious micronutrient fertilization programme even in the deficient soil. Total micronutrient content in soil is a poor predictor of their availability to plants because it doesn’t take into account of the fact as to how and to what extent the solubility and mobility of micronutrients
are affected by important soil properties. Hence, it is the extractable micronutrient, what we call available pool in soil, is of utmost importance. The reliability of soil test crop response correlation studies is greatly enhanced, if soil test is based on sound theoretical background. Careful consideration of chemistry of micronutrient behaviour in soil including fractionation as a part of the soil test development should further make soil test more efficient and reliable. Against this background, an attempt has been made in this review to collect, collate and present the available information on current status of soil tests for micronutrients with suggestions for future line of work and prospects. Chemical Pools of Micronutrients in Soil Micronutrient Cations Zinc, Cu, Fe, Mn and Ni are called micronutrient cations as they carry positive charge. Fractionation of micronutrients in soil provides an insight into their binding forms, dynamics, plant availability and possible environmental impacts. Each chemical pool of a micronutrient in soil is assumed to have attributes of concentration, size, turnover rate and equilibrium with other pools. According to Viets (1962), micronutrients are associated with mainly five pools in soil namely, i) soil solution, ii) surface adsorbed and exchangeable, iii) associated with organic matter, iv) oxide-bound, and v) present in primary and secondary aluminasilicate minerals. These are conceptual pools of micronutrients, which are sequentially extracted by employing various reagents like water, salt solutions, acids and bases. Practically the portion of total micronutrient, which could be extracted with specific reagents, is known as fraction. Generally reagents, shaking time and temperature are used sequentially from the least to the greatest extremes to extract various fractions of micronutrients in the soil. In other words, initially
most loosely bound micronutrient is extracted and gradually more and the most tightly bound micronutrient fractions are removed. Water soluble pool mainly includes non-adsorbed ions and adsorbed on suspended colloids that are estimated in a displaced soil solution or in an aqueous extract. Exchangeable pool is obtained by extracting the soil with weak cation and anion exchangers. Water soluble pool is usually too small to measure. Hence, exchangeable pool usually includes water soluble pool also and is measured as water soluble plus exchangeable content together. The specifically adsorbed pool is replaced by the mass of cations with similar affinities for the adsorbent. Micronutrients in this pool are adsorbed with great affinity mainly on clay and humus of soil. Oxide bound fractions are extracted using reducing agents, while oxidising agents are used to extract organically bound fractions. The portions of micronutrients associated with amorphous and crystalline Fe oxides as well as Mn oxides are determined separately, if required. Dilute inorganic acids are usually used to extract carbonate bound fraction in soil. To dissolve micronutrients present as a structural component and mineralogical make up, concentrated inorganic acids are used coupled with strong heating. A number of sequential fractionation procedures have been developed based on the Viets’s concept. The first sequential fractionation scheme for copper was developed by McLaren and Crawford (1973) and over a period of time, other researchers modified it (Tessier, 1979; Murthy, 1982; Mandal and Mandal, 1986; Miller et al., 1986; Iwasaki and Yoshikawa, 1990; Phillips and Chapie, 1995; Ma and Uren, 1998; Sanchez-Martin et al., 2007) for micronutrient cations in general (Table 1). McLaren and Crawford (1973) concluded that amount of Cu available to plants was controlled by equilibria involving soil solution plus exchangeable, specifically adsorbed forms and organically bound Indian Journal of Fertilisers, May 2018 33
fraction. For waterlogged soil, Murthy (1982) developed a fractionation procedure at IRRI, Phillipines, where exchangeable plus complexed Zn was extracted with Cu(OAc) 2. Further, Mandal and Mandal (1986) split the Cu(OAc)2 extractable fraction in to water soluble plus organic complexed fraction. Organic complexed fraction plays the most important role in Zn nutrition of wet land rice in acid soils. Total metal in soil was partitioned in to nine fractions by Miller et al. (1986), where, no provision was made to determine carbonate bound fraction. Hence, Ma and Uren (1998) developed the sequential fractionation procedure to assess carbonate bound fraction separately as this fraction may play a major role in soils of arid regions. Adhikari and Rattan (2007) studied the distribution of Zn among the soil fractions employing the procedure of Miller et al. (1986). The study revealed that more than 90% of the total Zn content occurred in the relatively inactive clay lattice and other mineral bound form (residual) and that only a small fraction occurred in the forms of water soluble plus exchangeable (0.31 to 3.15%), organically bound (3.29 to 6.86%), amorphous Feoxide bound (2.43 to 5.33%) and crystalline Fe oxide bound (1.99 to 5.00%) forms. Using the same scheme, Barman et al. (2015) showed that residual Ni was the most dominant fraction in soil constituting 3.19 to 63.6% of total Ni in tropical alluvial soils of divergent physicochemical characteristics. The water soluble plus exchangeable Ni accounted for only 0.70 to 4.04% of total soil Ni. Organically bound Ni varied from 1.60 to 6.85% of total Ni; these values are relatively lower as compared to those reported for temperate soils. Main problems associated with sequential fractionation scheme are selectivity of the reagents for specific fraction and re-adsorption as well as precipitation of the target micronutrients. Such problems are aggravated with the increase in number of fractions. Hence, Sanchez-Martin et al. (2007) as well
Table 1. Fractionation of micronutrient cations in soil Scheme proposed by Miller et al. (1986) modified by Iwasaki and Yoshikawa (1990) Fraction
Reagent
Exchangeable and water soluble Pb-displaceable Acid soluble Mn-oxide bound Organic bound Amorphous Fe-oxide bound Crystalline Fe-oxide bound Residual Scheme proposed by Sanchez-Martin et al.(2007) Exchangeable and water soluble Weak acid extractable Fe–Mn oxide bound Organically bound Residual Scheme proposed by Phillips and Chappie (1995) as Water soluble plus exchangeable Carbonate bound Fe–Mn oxide bound Organically bound Residual
as Phillips and Chappie (1995) proposed short fractionation procedures, which include only five major fractions. Either of these methods can be used for sequential extraction of micronutrients from soil depending upon the availability of reagents and ease of extraction procedure including shaking and temperature. However, use of 1M MgCl 2 (magnesium chloride) in extraction of water soluble plus exchangeable fraction of metals as suggested by Phillips and Chappie (1995), creates problem with ICP-MS. Hence, 0.5M Ca(NO3)2 as used by Miller et al. (1986) was suggested for extraction of water soluble plus exchangeable fractions (Golui et al., 2017). For ICP-MS, reagents used for extraction of organically bound fraction by Phillips and Chappie (1995) should be more suitable than that of scheme proposed by Sanchez-Martin et al. (2007). Employing the procedure of Phillips and Chappie (1995), Ray et al. (2016) reported that more than 50% of Zn in soil is present in residual fraction and as high as 1.98, 4.66, 25.6 and 16.8% of total Zn is constituted by water soluble plus exchangeable, carbonate bound, oxide-bound and organically bound fractions,
Shaking time (h)
0.05M Ca(NO3)2 0.05M Pb(NO3)2 + 0.01M Ca(NO3)2 0.44M CH3COOH + 0.01M Ca(NO3)2 0.01M NH2OH.HCl + 0.1 M HNO3 0.1M K4P2O7 0.175M (NH4)2C2O4 + 0.1 M H2C2O4 0.1M H2C2O4 + 0.1M ascorbic acid Aqua-regia + HF
16 16 8 30 min 24 4 30 min (Boiling)
1M MgCl2 (pH 7.0) 1M NaOAc adjusted to pH 5 with HOAc 0.175 M (NH4)2C2O4 + 0.1 M H2C2O4 0.1 M Na4P2O7 Digestion with aqua-regia modified by Golui et al.(2017) 0.5M Ca(NO3)2 1M NaOAc (pH 5.0) 0.04M NH2OH.HCl in 25% acetic acid (vol/vol) 0.02 M HNO3+ 30% H2O2 (pH 2.0) vol/vol) + 3.2M NH4OAc in 20% HNO3 (vol/vol) Conc. H2SO4 + 48% HF + 60% HClO4
1 5 4 24 -
respectively. Long back, it was established that generally, less than 10% of total soil micronutrients are present in water soluble and exchangeable forms and these make principal contributions to the available pool in soil (Lake et al., 1984). Adhikari and Rattan (2007) also obtained the most consistent positive relationships of plant Zn uptake with relatively more labile Zn fractions in soil. In another study, water soluble plus exchangeable, specifically adsorbed and organically bound contributed positively towards Ni uptake by soybean, while, Mn and Fe-oxide bound as well as residual Ni had negative influence on its content in soybean plant (Barman et al., 2014). Micronutrient Anions Boron, Mo and Cl occur as anions in soil solution and carry negative charge. Jin et al. (1987) developed a separate extraction scheme to determine the distribution of B among different fractions in soils. Hou et al. (1994, 1996) modified this scheme and developed a sequential fractionation method for partitioning total soil B in to different distinct pools. In both of Indian Journal of Fertilisers, May 2018 34
16 6 6 2.5 -
these schemes, B in the extracts of the different fractions is measured by inductively coupled plasma atomic emission spectroscopy (ICP-AES). Because of high initial and maintenance costs, ICP-AES is not available in most of the laboratories, particularly in developing countries. To overcome this problem, a sequential fractionation scheme compatible with colori-metry was developed (Figure 1). On an average, readily soluble, specifically adsorbed, oxide bound and organically bound B constituted only 0.73 to 8.85% of the total B in soils having pH in the range of 3.72 to 8.50. The mean proportion of extractable B was 0.38% as readily soluble form, 0.34% as specifically adsorbed form, 1.45% associated with various oxides and hydroxides, and 0.62% associated with organic matter (Datta et al., 2002). In acid soils of West Bengal (pH 5.4), residual fraction was most dominant (83.4%), followed by organically bound (7.58%), oxide bound (5.86%), specifically adsorbed (1.72%) and readily soluble (1.41%) (Barman et al., 2014). Specifically adsorbed and organically bound B were found to be the main contributors to the
Figure 1. Flow diagram for the fractionation of B in soils
available B (Dey et al., 2017). Speciation of Mo in soil has an important implication in understanding its chemistry and subsequent transfer to plants. About 88 to 94% of total Mo was considered to be unavailable to plants (Shuman, 1991). Unlike other micronutrients, Mo is predominantly (i.e., 20-50% of total) present in amorphous and crystalline Fe-oxide fractions. Besides, fractionation of Mo can provide information on its release, fixation, mobility and enrichment in the environment. Very little information is available on solid phase speciation of Mo in soil. Recently, Liang and Zhu (2016) optimized sequential extraction scheme for Mo, which was compared with Tessier (1979) and Commission of European Community Bureau of Reference (Rauret et al., 1989). Out of these three extraction procedures, the optimized extraction scheme proved to be suitable in black shales. In proposed scheme, exchangeable plus water soluble, organically bound, oxide bound,
sulphide bound and residual Mo are extracted with KH 2 PO 4 + K 2 HPO 4 , NaOH, HCl, H 2 O 2 and HNO3+HF+H2O2, respectively. This fractionation scheme needs to be evaluated and validated for different types of soils. Soil Test Micronutrients
Methods
for
Micronutrient Cations Total elemental analysis of soils necessitates their decomposition into soluble forms by acid digestion (HF and either HClO4 or H 2SO 4) or fusion with various fluxes such as Na 2CO 3. However, total micronutrient content in soil includes large fractions that are unavailable to plants, microorganisms and soil fauna. Hence, various chemical reagents have been tried over the years to extract available pools of micronutrients from different soils. Broadly all the extractants, which have been used for assessing available micronutrients in soil, can be classified into two groups, viz. (i) dilute acids and salt solutions, and (ii) extractants that use chelating Indian Journal of Fertilisers, May 2018 35
agents. In acid soils, dilute acids have been used as extractants for assessing available micronutrients in soil for many years specifically prior to the use of chelating agents, whereas, salt solutions of varying strengths have been tried for both acid and alkaline soils. Although, acidic extractants do not have a sound theoretical basis, a welldeveloped database exists to show the positive relationship between acid extractable micronutrients in soil and crop response. These extractants had been widely used with variable success up to late 1970’s. One of the major advances in micronutrient soil testing has been the development of extracting solution that contains chelating agents, primarily DTPA and EDTA. During the extraction, chelates reduce the activity of free metal ions in solution through the formation of soluble metal-chelate complex. In response to the depletion of metal ions in solution, more metal ions come from solid phases to replenish this depletion. Such depletion in the activity of metal ions in solution closely
Table 2. Important soil test methods for micronutrients (Datta and Meena, 2015) Element
Extractant
Soil : solution ratio
Shaking period (minutes)
Critical level of deficiency (mg kg-1)
Zinc
0.005M DTPA + 0.1M TEA* + 0.01M CaCl2 (pH 7.3) 0.1 N HCl 1 N NH4OAc (pH 4.6) 0.05 N HCl
1:2 1:5 1:10 1:2
120 30 60 5
0.5-1.0 1-5 0.2-0.5 1.0
Copper
0.005 M DTPA + 0.1 M TEA + 0.01 M CaCl2 (pH 7.3) 0.1 N HCl 1N NH4OAc (pH 4.8)
1:2 1:5-10 1:2-4
120 30 60
0.2-0.5 1-3 0.2
1:2 1:10-20
120 60
2.5-5.8 2
0.005 M DTPA+0.1 M TEA + 0.01 M CaCl2 (pH 7.3) 1N NH4OAc (pH 7.0) 3N NH4H2PO4 and 0.1 N H3PO4
1:2 1:10 1:10
120 30 60
2-4 3-4 15-20
1:2
120
0.17
Boiling hot water 0.01M CaCl2 + 0.05 M mannitol 0.1M salicylic acid
5:10 1:2 1:2
5 60 60
0.5-1.0 0.25 0.45
0.175M Ammonium oxalate (pH 3.3)
1:10
360
0.05-0.2
Iron Manganese
Nickel Boron
Molybdenum
0.005 M DTPA + 0.1 M TEA + 0.01 M CaCl2 (pH 7.3) 1 N NH4OAc (pH 4.8)
0.005M DTPA + 0.1 M TEA + 0.01 M CaCl2 (pH 7.3)
TEA = Triethanolamine
mimics the reduction in activity of metal ions in solution due to plant uptake (Figure 2). Thus, quantity of micronutrients extracted by a chelate reflects both initial concentration in the soil solution (intensity factor) and the ability of
soil to maintain this concentration (capacity factor). Thus, chelating agent simulates micronutrient removal by plant roots and subsequent replenishment from labile solid phases in the soil. DTPA soil test [0.005M DTPA + 0.001M
CaCl 2 .2H 2 O + 0.1M tri-ethanol amine (TEA)] of Lindsay and Norvell (1978) is widely used to assess the status of available micronutrient cations (Zn, Cu, Fe, Mn and Ni) in soil (Table 2). Selection of DTPA is based on its
Figure 2. Extraction with DTPA simulates micronutrient removal by plant roots and replenishment from labile solid phases in soil Indian Journal of Fertilisers, May 2018 36
optimum combination of stability constants necessary to simultaneously extract Zn, Cu, Fe and Mn. Since, this extractant was developed for calcareous and alkaline soils, it was specifically designed to avoid excessive dissolution of CaCO3 with release of occluded micronutrients, which are normally not available to plants. This objective was achieved partially by buffering the extractant in a slightly alkaline pH with TEA and partially by including soluble Ca 2+ . Triethanolamine is selected as buffer because of its pKa = 7.8 and it clearly burns during flame atomization in atomic absorption spectrometry. Approximately 2/3rd of the Ca is associated with DTPA. At selected pH of 7.3, approximately 3/4 th of TEA is protonated and present as HTEA+. When TEA is added to soil, additional Ca2+ and Mg2+ enter into solution, largely because of the HTEA+ exchanges with Ca and Mg from soil exchange sites. This exchange generally raises the concentration of Ca2+ by 2 to 3 fold and aids in suppressing the dissolution of CaCO3 in calcareous soils. At pH 7.3, 70-80% of buffering capacity provided by TEA is consumed in alkaline soils. Use of this extractant in acid soils likely results in neutralization of the remaining buffer capacity and pH of DTPA and soil extract becomes unpredictable. This leads to the disruption of metal-chelate equilibria causing shifts in metal chelating tendencies. Nutrientwise brief accounts on soil test methods used over the years are
given below: Zinc Before the advent of chelating agent-based extractants for assessment of available Zn in soil, two soil test methods had been widely used and calibrated against plant data, which include extraction with a two phase system of aqueous 1M NH4OAc and 0.01% dithizone in CCl 4 and 0.1M HCl. Although dithizone method was first used by Shaw and Dean (1952), usefulness of dithizone method had been established as a means of predicting Zn uptake by plants in a large number of studies (Martens et al., 1966; Brown and Krantz, 1961; Brown et al., 1962; Shaw and Dean, 1952; Muthukumararaja and Sriramachandrasekharan, 2012). Dithizone complexes Zn during extraction in solution phase. The concentration of Zn-dithizone complex is kept low in aqueous NH 4OAc phase during extraction due to addition of CCl4 (Shaw and Dean, 1952). In some cases, dithizone was superior to 0.1N HCl in assessing available Zn status of soil (e.g., Martens et al., 1966; Brown et al., 1971). However, inclusion of soil pH with 0.1N HCl extractable Zn was suggested as the most practical approach for routine soil testing. Similarly, dithizone method proved to be more efficient, if soil reaction was taken in to account. For example, critical level of deficiency of dithizone extractable Zn in soil was 0.3 mg kg -1 at pH 5 which was linearly increased to 2.3 mg kg -1 at pH 8 (Shaw and Dean, 1952). In 1969,
0.01M EDTA-1M (NH4)2CO3 soil test was introduced, which compared favourably with dithizone method and was an improvement over 0.1N HCl method (Trierweiler and Lindsay, 1969). Unlike strong acid extractants, this method suppressed the dissolution of carbonate and oxides and thereby avoids extraction of occluded Zn. However, in 80s all the extractants, be these dilute acids or salt solutions, have been almost completely replaced by DTPA soil test of Lindsay and Norvell (1978), surprisingly in all type of soils. Efficacy of DTPA soil test for separating Zn responsive and nonresponsive soils was compared with other extractants. In most cases DTPA was evaluated as the most promising for assessing available Zn in soil as compared to other extractants (e.g., Rajendran and Iyer, 1981; Gajbhiye et al., 1984; Singhal and Rattan, 1999). Lowland rice is one of highly sensitive crops to Zn deficiency with Zn being the most important micronutrient limiting rice yields (Alloway, 2004; Dong et al., 2006; Rattan et al., 2008; Muthukumararaja and Sriramachandrasekharan, 2012). Muthukumararaja and Sriramachandrasekharan (2012) reported that chelating agent based extractants, dithizone and NH 4OAc could successfully assess the available Zn status in Entisols (pH 6.5-8.7) and Vertisols (pH 7.48.8) (Table 3). However, DTPA had most consistent correlation with Bray’s % yield of rice. In some acid soils of Assam (pH 4.9-5.8), soil Zn
Table 3. Relationship of extractable Zn in soil with Bray’s per cent yield and Zn content in rice (Muthukumararaja and Sriramachandrasekharan, 2012) Extractant Bray’s per cent yield Tissue Zn concentration Vertisol Entisol Vertisol Entisol 0.005M DTPA + 0.01M CaCl2 + 0.1M TEA (pH 7.3)
0.623 b
0.833 b
0.779 b
0.802 b
0.01M EDTA + 1M (NH4)2CO3 (pH 8.6)
0.517 a
0.663 b
0.980 b
0.780 b
0.01M EDTA + TEA (pH 6.7)
0.156 a
0.161 a
0.950 b
0.839 b
0.01M EDTA + 1N NH4OAc (pH 7.0)
0.196 a
0.068 a
0.969 b
0.880 b
1N NH4OAc + 0.01% dithizone
0.379 a
0.375 a
0.969 b
0.957 b
0.01M EDTA
0.072 a
0.013 a
0.962 b
0.882 b
1N NH4OAc (pH 7.0)
0.053 a
0.445 a
0.967 b
0.970 b
a, b indicate
that values of r are significant at 0.05 and 0.01 probability levels, respectively. Indian Journal of Fertilisers, May 2018 37
extracted with 0.005M DTPA and 0.01M EDTA-1M (NH 4 ) 2 CO 3 showed more consistent positive relationship with Bray’s per cent yield of rice as compared to 0.1N HCl, 1N ammonium acetate and 2N MgCl 2 (Shukla and Tiwari, 2016). Similarly, for upland crops like mustard and soybean, DTPA, EDTA-(NH 4 )2 CO 3 and 0.05N HCl were equally effective in assessing available Zn status of some alluvial soils (Singhal and Rattan, 1999). Rapidity and capability of soil test methods are becoming increasingly important, particularly in the face of assessing soil health for millions of farmers across the country within a stipulated time. In this context, use of multinutrient extractants for assessing available major and micro nutrients in soil should be quite helpful. Quite a few multinutrient extractants have been devised over the years (Table 4). The first universal extractant was developed by Morgan in as early as 1941 and pH was chosen as 4.8 to simulate the CO2 saturated solution adjacent to root hair. It was envisaged that this extractant would act as a mild solvent for Fe and Al phosphate as well as other minerals that might release nutrients in the soil solution. Wolf modified Morgan’s reagent in 1982 and added DTPA to enhance the extraction efficacy of
micronutrient cations. In 1954, Mehlich used dilute double acid solution as an extractant (Mehlich 1) for multinutrients, which was modified later in 1978 (Mehlich 2), where H 2 SO 4 was replaced, the strength of HCl was reduced and NH 4 F was included to make it suitable for calcareous soil. Further, Mehlich 3 was devised, where mainly EDTA was introduced in Mehlich 2 to enhance the extractability of Cu and other micronutrient cations. The ABDTPA extractable P, K, and micronutrients in alkaline soils showed a positive correlation with Olsen’s P, ammonium acetate K and Lindsay and Norvell’s DTPA-Zn, Cu, Fe and Mn (Soltanpour and Schwab, 1977). However, for acid soil under rice crop, AB-DTPA buffered at pH 5.6 was more suitable for assessing available soil Zn status as compared to the ABDTPA method of Soltanpour and Schwab (1977). Critical levels of deficiency of Zn were worked out as 0.48, 0.80, 0.70 and 2.2 mg kg-1 for DTPA, NH4OAc plus EDTA, AB-EDTA and 0.1M HCl, respectively for some noncalcareous soils of India. In some calcareous soils, DTPA was a useful extractant for predicting response of rice and wheat to applied Zn (Sakal et al., 1981). Unfortunately, these extractants have not been calibrated with plant responses or currently used soil test methods extensively. Under
AICRP-MSPE, AB-DTPA extractable micronutrient cations in alkaline and calcareous soils showed a highly consistent positive relationship with that of DTPA extractable micronutrients (Shukla and Tiwari, 2016). Further evaluation and calibration of this soil test in relation to plant response for all nutrients are in progress. From a practical point of view, at least the suitable quantitative relationship and rating of multinutrient extractable nutrient with standard soil test need to be developed for making multinutrient soil test useful in the recommendation (e.g. Table 5). For this purpose, calibration of these soil tests directly with plant response will be the best option. Whenever one strays from the original design of the soil test, one should be aware of the possible consequences and pass that awareness to others. For example, if the DTPA soil test is applied to acid and metal contaminated soils, the buffering capacity of the soil test solution may be exceeded (O’Connor, 1988). Metal-chelate equilibria disrupt leading to the dramatic shift in individual metal chelating tendencies. Use of the DTPA soil test in acid soil with high organic matter content does not yield enough aliquot for proper estimation even with AAS. Norvell (1984) suggested the recognition of the limitations of chelating
Table 4. Multi-nutrient extractants with their soil adaptability and elements of determination Extractants
Soil : solution Shaking ratio time (min)
Soil type
Element determined
0.72N NaOAc + 0.52N CH3COOH
1:4
15
Acid soil
0.073M NaOAc + 0.52N CH 3COOH + 0.001M DTPA
1:2
5
0.05N HCl + 0.025N H2SO 4
1:4
5
Acid to neutral soils,organic soils Acid soils
0.2N CH3COOH + 0.015N NH4F + 0.2N NH4Cl + 0.012N HCl
1:10
10
Acid to alkaline soils
0.2N CH3COOH + 0.25N NH4NO3 + 0.013N HNO3 + 0.015N NH4F + 0.001M EDTA 0.005M DTPA-1M NH4HCO3 (pH 7.6)
1:10
5
1:2
15
Acid to P, K, Ca, Mg, Na, neutral soils Cu, Mn, Zn Alkaline soils P, K, Na, Fe, Mn, Zn, As, Cd, NO3
Indian Journal of Fertilisers, May 2018 38
Reference
P, K, Ca, Mg, Cu, Fe, Mn, Zn, NO3, NH4, SO4, Al, As, Hg, Pb P, K, Ca, Mg, B, Cu, Fe, Mn, Zn, Al, NO3, NH4 P, K, Ca, Mg, Na, Mn, Zn P, K, Mg, Ca, Na, Mn, Zn
Morgan (1941) Wolf (1982)
Mehlich (1954) Mehlich (1978) Mehlich (1984) Soltanpour and Schwab (19 77)
Table 5. Index values (mg kg-1) used for Zn, Fe, Cu and Mn extracted from soils by the DTPA method of Lindsay and Norvell and their equivalents for the new soil test method for irrigated corn, sorghum, Sudan grass, sorghum × Sudan grass hybrids, beans and potatoes grown in Colorado (Soltanpour and Schwab, 1977) Category
DTPA of Lindsay and Norvell Zn
Low Marginal Adequate
0-0.5 0.6-1.0 >1.0
Fe 0-2.0 2.1-4.0 >4.0
capacity when extracting metal rich soils, especially at low pH. In this context, either concentration of chelating agents can be increased or extractant to soil ratio may be widened. Considering the suitability of extractant in respect of quality of flame of AAS, widening extractant to soil ratio should be a better proposition. Hence, Norvell (1984) suggested to evaluate 0.005M chelating agents [DTPA, HEDTA (hydroxyl ethylene diamine triacetic acid) and NTA (nitrilotriacetic acid)] containing 0.01M CaCl2 buffered at pH 5.3 by a mixture of ammonium acetate and acetic acid (0.01M total acetate) particularly for acid soils. However, this extractant is yet to be evaluated extensively in acid soils. Copper In view of the adequacy of Cu in most of our soils, soil test method for Cu has seldom been evaluated and calibrated with plant response. Most reported cases of Cu deficiency have been confined to organic soils, where the problem is of availability rather than low Cu content. Bioassay technique has been used earlier for assessing Cu supplying capacity of the soil, where the growth of microorganisms was monitored (Cox and Kamprath, 1972). Subsequently, various extractants came into existence over the years. 0.01M EDTA-1N NH4OAc (pH 7.0) was a better extractant for predicting Cu uptake by oats grown on some acid soils of Western Nigeria as compared to that of 0.005M DTPA (Lindsay and Norvell, 1969), 0.1N HCl and 1N HCl (Osiname et al., 1973). However, Grewal et al. (1969) found that response of maize and wheat to Cu was better estimated using 1N
Cu
Mn
0-0.2 >0.2
0-1.0 >1.0
NH 4HCO3-DTPA proposed by the authors Zn
0-0.9 1.0-1.5 >1.5
NH4OAc than a chelating agent or dilute acid. In another study, dilute 0.1N HCl, 0.01M EDTA, and ion exchange resin were equally effective in evaluating the Cu status of some acid soils using Phaseolus mungo as a test crop (Acquaye et al., 1972). Now in India, DTPA soil test (Lindsay and Norvell, 1978) has almost invariably been used for assessing Cu status of soil under AICRPMSPE. This soil test showed reasonably good correlation with AB-DTPA (Soltanpour and Schwab, 1977) in some soils of Tamil Nadu (pH 3.74-9.10) and Bihar (pH 5.908.20). As such delineation of Cu status of soil is not needed until and unless foliar deficiency symptoms appear. Iron A wide variety of extractants including NH4OAc, EDDHA, EDTA, DTPA, H2SO 4, NaNO3, HCl, acetic acid and cation exchange resin (Cox and Kamprath, 1972; Acquaye et al., 1972; Eteng and Asawalam, 2015) have been tested for their efficacy in assessing available Fe status of soils. Olsen and Carlson (1950) reported correlation of 1M NH 4OAc (pH 4.8) extractable Fe status in soil with a chlorotic system of plants. Iron extracted with NH 4 OAc and EDDHA [ethylenediamine-N,N’-bis(2hydroxyphenylacetic acid)] showed a better correlation with plant growth than EDTA and H 2 SO 4 . Acquaye et al. (1972) reported that cation exchange resin method could successfully indicate the availability of Cu, Mn, Zn but was not sensitive enough for the prediction of available Fe. In some acid soils of Nigeria (pH 4.01-5.89), 0.1N HCl and 0.05 M EDTA were capable of predicting Fe availability to maize better as Indian Journal of Fertilisers, May 2018 39
Fe
Cu
Mn
0-2.0 2.1-4.0 >4.0
0-0.5 >0.5
0-1.8 >1.8
compared to 1N NH 4OAc, 0.05M EDTA+1N NH 4 OAc (Eteng and Asawalam, 2015). Shukla and Tiwari (2016) reported close agreement between DTPA extractable Fe with multinutrient extractant AB-DTPA (Soltanpour and Schwab, 1977) in both calcareous and noncalcareous soils. Availability of Fe is affected by soil pH to great extent, whereas its availability also depends on drainage or redox condition of soil (Ponnamperuma et al., 1967; Lindsay, 1979; Sahrawat, 2003; Nayak, 2008). Iron toxicity is unlikely to be a problem where a soil is well aerated and well drained with good structure and porosity (Nayak, 2008). Iron chelates must be stable in soil environments, which aid in the movement of Fe to plant roots (Norvell and Lindsay, 1982). Failure of the soil test for Fe is attributed to such variety of complexities and interactions with the physical and chemical environment of soil. In case of Fe, none of the methods has received wide uses or accepted as a standard so far. Even DTPA soil test (Lindsay and Norvell, 1978) is not universally applicable and reports are there in literature indicating the failure of DTPA soil test in assessing available Fe content in soils. For example, Katyal and Sharma (1984) failed to establish any relationship of DTPA extractable Fe with Bray’s per cent yield and total Fe concentration in rice plants grown on sixteen upland alkaline soils with diverse phyisco-chemical properties. Similarly, DTPA extractable Fe did not show any relationship with the total Fe content in various crop species, including rice grown on farmers’ fields, which had been receiving sewage irrigation
Table 6. Simple correlation coefficient (r) among different soil and plant parameters (Pal et al., 2008) Parameter DTPA-Fe at 60 DAS@ DTPA-Fe at 90 DAS@ 1DTPA-Fe at harvest@ 2NH OAc-Fe at harvest@ 4 Total plant-Fe at 60 DAS$ Total plant-Fe at 90 DAS$ Plant-Fe2+ at 60 DAS$ Plant-Fe2+ at 90 DAS$
Correlation coefficient (r) Plant Fe Total Fe2+ 0.41 0.76 b 0.28 0.75 b 0.41 0.27 Yield Straw Grain 0.46 0.39 0.43 0.44 0.69 b 0.60 a 0.62 b 0.61 a
@Based on all the treatment combinations except foliar
spray (12 observations) the treatment combinations (16 observations) a, b indicate that values of r are significant at 0.05 and 0.01 probability levels, respectively. 1Lindsay and Norvell (1978) 2Olsen and Carlson (1950) $Based on
(Rattan et al., 2005). In a study at Coimbatore, sorghum responded significantly to Fe application on calcareous soils (Alfisol and Vertisol) having up to 20-50 mg kg -1 DTPA extractable Fe (Katyal, 1985). Even inclusion of some important soil properties (pH, organic carbon and clay) in the regression equation along with DTPA extractable Fe failed to improve the prediction of Fe availability to rice (Katyal, 1985). The prediction of micronutrient deficiencies based on total content in plant tissues has been reasonably successful for all micronutrients except Fe because chlorotic plants generally have as much or higher Fe than green ones (Katyal and Sharma, 1980, 1984; Takkar and Kaur, 1984). In this regard, Fe2+ content in rice leaves is more useful than total Fe content as an indicator of its nutritional status in plant. Ferrous iron (Fe2+) content in rice plants proved to be a better index of Fe-nutrition status compared to total plant Fe and chemically extractable soil Fe (Table 6). The Fe2+ concentration of 37 mg kg-1 in plants (on dry weight basis) appeared to be an adequate level at 60 days after sowing for direct seeded rice grown under upland aerobic condition (Pal et al., 2008). Manganese Chemistry of Mn in soil is complex
because of its various oxidation states. Usually, Mn occurs in the soil as insoluble oxides of trivalent and tetravalent Mn as well as exchangeable and water-soluble divalent Mn. Divalent Mn (Mn2+) is usable forms for plants, which is needed in a small amount and uptake is
