A quantitative assessment of phosphorus forms in some Australian soils.
Australian soils are generally characterised by low phosphorus (P) concentrations by world standards, primarily because they are generally highly weathered and many are derived from sedimentary rocks, including sandstone, which are generally low in P (Beadle 1962; Holford 1997). Plant-available P in soil is an important determinant of soil fertility (Norrish and Rosser 1983); consequently, application of P fertilisers is usually required to ensure adequate crop production in Australian soils (Cornish 2009a). The long-term application of P fertilisers in southern Australia has exceeded P removal such that substantial quantities of P have accumulated in some soils (McLaughlin et al. 1991). This not only represents an inefficient and unsustainable use of P fertilisers but also increases the potential for nutrient leaching to the off-farm environment and can cause eutrophication of waterways (Sharpley et al. 1994).
Ineffectiveness of P fertilisers is due to the fixation of P by sorption, complexation, and precipitation reactions with soil constituents. Generally, 10-30% of applied fertiliser P is recovered by crops in the year following application. The remainder of P necessary for plant growth is obtained from residual inorganic P and recalcitrant forms of organic P or 'unavailable' P forms (Bolland and Gilkes 1998; Bunemann et al. 2006). Recent research has examined ways to reduce the input of soluble P fertilisers and to better use existing soil P (e.g. Conyers and Moody 2009; Cornish 2009b; Evans and Condon 2009; Guppy and McLaughlin 2009; Richardson et al. 2009). Potential strategies to increase plant-available P include the selection or modification of plants with favourable root architecture (fine roots, lots of root hairs) and effective root exploration (Richardson et al. 2009), the mutualistic association of plant roots with mycorrhizal fungi (Smith et al. 2003), and the direct application of manufactured extracellular enzymes (Conyers and Moody 2009; Evans and Condon 2009). However, the success of any technique that aims to increase the availability of soil P and implement a more sustainable approach to P management in Australian soils will rely on an understanding of the P forms (including organic P forms) that exist within the soil.
Early work on the characterisation of soil organic P used extraction and chromatographic techniques. These techniques indicated myo-inositol hexakisphosphate (phytate), a P-rich compound derived predominantly from plant seeds (Reddy et al. 1989), to be a dominant form of soil organic P. Initial investigations of soil 'phytate fractions' in three Australian soils were undertaken by Cosgrove (1962, 1963) and Cosgrove and Tate (1963), where they were able to show the presence of myo- + DL, neo-, and scyllo-inositol hexaphosphates. Cosgrove (1963) provided approximate concentrations of some inositol phosphate components, with estimates of myo-+DL-inositol hexaphosphate equating to 10-13% of total organic phosphate and scyllo-inositol hexaphosphate ranging from 2 to 3%. Williams and Anderson (1968) undertook the first comprehensive characterisation of Australian soils and focused on identifying the inositol-phosphate concentrations of 47 surface soils from eastern Australia. The concentrations varied widely (0.4-38% of total organic P) but the authors did not identify other forms of organic P. Irving and Cosgrove (1982) detected chiro-, neo-, myo-, and scyllo-inositol pentakis-and hexakisphosphates in four soils from New South Wales, the two most abundant species being the myo- and scyllo-hexakisphosphates with concentrations ranging from 12 to 167 mg P/kg. Similarly, Steward and Tate (1971) reported that inositol polyphosphates comprised 2 31% of soil organic P in four South Australian (Urrbrae) and four eastern Australian soils.
More recently, solution 3,p nuclear magnetic resonance (NMR) spectroscopy has become the most widely used technique for the speciation of soil P (McDowell et al. 2005; Turner et al. 2005; Murphy et al. 2009). However, the application of solution 31p NMR spectroscopy to Australian soils and therefore a more detailed characterisation of P forms is limited to a few studies (Smernik and Dougherty 2007; Bunemann et al. 2008b; Doolette et al. 2009), one of which suggested that phytate concentrations can be overestimated in 31p NMR analyses through mis-assignment of phytate peaks (Smernik and Dougherty 2007). This is consistent with the earlier findings of McLaughlin et al. (1988), who suggested that a major portion of organic P that accumulated in cropping soils was of microbial, rather than plant, origin.
In this study we apply solution [sup.31]P NMR spectroscopy to a diverse range of Australian soils with varying chemical and physical properties to provide a detailed characterisation and quantitative assessment of the forms of sodium hydroxide-ethylenediaminetetra-acetic acid (NaOH-EDTA) extractable P.
Materials and methods
Soil chemical and physical analysis
Eighteen topsoils were collected from New South Wales, South Australia, and Tasmania. Eight of these soils (Nowra, Casino, Collector, Coonabarabran, Wollongbar, Tocal, and Somersby A & B) were included in a previous study (Doolette et al. 2009). Of the extra 10 soils included in this study, two are from South Australia and the remaining eight comprise four pairs of soils with contrasting P fertiliser histories from Tasmania. All sites had been under their current management regime for at least 4 years. All soils were sampled, after the removal of the litter layer, to a depth of 0.10 m and air-dried.
Total soil P and inorganic and organic P contents were determined using the acid extraction and ignition methods of Saunders and Williams (1955). Soil pH was determined on 1 : 5 soil : water extracts. Organic C was determined using the method of Walkley (1947). Soils were classified according to the Australian Soil Classification (ASC) (Isbell 1996).
Samples were ground to pass through a 2-mm sieve before extraction. Soils were extracted in triplicate using methods based on those of Cade-Menun and Preston (1996). For all but three soils, 2.0 g of soil was shaken with 40 mL of 0.25 M NaOH and 0.05 M Na2EDTA for 16 h. For three low-P soils (Somersby A & B and Coonabarabran), 6.0 g rather than 2.0 g of soil was used in the extraction to provide acceptable signal-to-noise ratios in the NMR spectra. All extracts were centrifuged (1400G) for 10 min and filtered using Whatman no. 42 filter paper. A 15-mL aliquot was immediately frozen and freeze-dried for NMR analysis. Triplicate subsamples of the supernatant were also taken to determine the total P concentrations using nitric acid digestion and subsequent analysis by inductively coupled plasma-atomic emission spectroscopy (ICP-AES). The determination of inorganic P in the NaOH-EDTA soil extracts was based on colourimetry using the molybdenum blue method of Murphy and Riley (1962). Inorganic P in the NaOH-EDTA extracts was also quantified using deconvolution of 31p NMR spectra (detailed below).
NMR analysis of NaOH-EDTA extracts
Triplicate freeze-dried NaOH-EDTA extracts for each soil were combined for NMR analysis. A 500mg subsample of each composite extract was ground, re-dissolved in 5 mL of deionised water, and centrifuged (1400G) for 20 min. The supernatant solution (3.5mL) and deuterium oxide ([D.sub.2]O, 0.3mL) were placed in a 10-mm-diameter NMR tube. Solution [sup.31]P NMR spectra were acquired at 24[degrees]C on a Varian INOVA400 NMR spectrometer (Varian, Palo Alto, CA) at a [sup.31]P frequency of 161.9 MHz. Recovery delays ranged from 10 to 30s and were set to at least five times the [T.sub.1] value of the orthophosphate resonance determined in preliminary inversion-recovery experiments for each extract (data not presented). We used a 90[degrees] pulse of 32-45 [mu]s, an acquisition time of 1.0 s, and broadband [sup.1]H decoupling. Between 3224 and 49 000 scans were acquired for each sample, depending on the P concentration of the freeze-dried extract. Chemical shifts were referenced to [beta]-glycerophosphate at 4.63 ppm according to Doolette et al. (2009). The spectra presented have a line broadening of 2 Hz.
Quantification of P species from [sup.31]P NMR spectra
The relative concentrations of P species in the NaOH-EDTA extracts were determined from [sup.31]P NMR spectra using a combination of integration and deconvolution. Orthophosphate diester (2.0 to 1.0ppm) and pyrophosphate (-4.5 to -5.5ppm) concentrations were determined using integration alone. Integration was used to determine the combined concentration of inorganic orthophosphate and orthophosphate monoester P (6.5-2.6ppm). The relative concentrations of P species giving rise to the numerous individual peaks in this region of the spectrum were quantified by spectral deconvolution, using a method similar to that of Bunemann et al. (2008b). Each spectrum was fitted with up to 15 sharp peaks as well as a broad signal that spanned most of the monoester region. The sharp peaks included those of orthophosphate, [alpha]- and [beta]-glycerophosphate, phytate, adenosine-5'-monosphosphate (AMP), scyllo-inositol phosphate, five unknown peaks in the monoester region, and two unknown peaks downfield (higher ppm values) of orthophosphate. Each peak was defined by three parameters: the chemical shift (frequency), intensity, and line width. The degrees of freedom for the fit were limited by fixing the line widths of the sharp peaks at 10 Hz (0.06 ppm), and the broad resonance at 200Hz (1.24ppm). The line width of the well-defined inorganic orthophosphate resonance was allowed to vary. The absolute concentration of each P species (including those determined using integration alone and those determined using integration and deconvolution combined) was calculated by multiplying its relative contribution to total NMR signal by the total NaOH-EDTA extractable P concentration determined by ICP-AES.
A minor correction was used in the determination of phytate and orthophosphate concentrations due to the overlap of the phytate C-2 peak with the much larger orthophosphate peak. Total phytate was calculated as 6/5 times the total concentration of the three observable resonances, and 1/5 of this value was subtracted from the total orthophosphate concentration (Doolette et al. 2009).
Spiking experiments were carried out on two soil extracts to confirm the accuracy of the spectral deconvolution technique used for quantifying P species. This enabled an independent measure of the concentration of one of the species ([beta]-glycerophosphate) using the method of Smemik and Dougherty (2007). An aqueous solution (0.1 mL) of [beta]-glycerophosphate was spiked into the Parkham +P (0.74 g/L) and Elizabeth Town control (1.4 g/L) redissolved NaOH-EDTA soil extracts (3.5 mL and 0.3 mL [D.sub.2]O). The [sup.31]P NMR spectrum of the unspiked NaOH-EDTA extract was subtracted from the [beta]-glycerophosphate-spiked NaOH-EDTA spectrum, resulting in a spectrum of the [beta]glycerophosphate spike alone. Increasing proportions of the [beta]-glycerophosphate spectrum were then subtracted from the unspiked NaOH-EDTA spectrum until the [beta]-glycerophosphate resonance was null. The lower limit of [beta]-glycerophosphate was defined by the maximum proportion of the [beta]-glycerophosphate spectrum that resulted in the appearance of a positive [beta]-glycerophosphate resonance in the unspiked NaOH-EDTA spectrum. The upper limit was defined by the minimum proportion of the [beta]-glycerophosphate spectrum that resulted in the appearance of a negative [beta]-glycerophosphate resonance in the unspiked NaOH-EDTA spectrum.
Results and discussion
The 18 Australian soils selected for this study span a wide range of soil management, climate, chemical and physical properties, and soil orders (Table 1). Mean annual temperature ranged from 15 to 23[degrees]C and mean annual precipitation from 387 to 1318 mm (www.bom.gov.au). Soil pH([H.sub.2]O) varied from moderately acidic (4.5) to moderately alkaline (8.3). Soil organic carbon ranged from 10 to 95 g/kg. Soils were used for a range of agricultural practices including pasture production (dairy and sheep), horticulture (conventional and organic), cropping, and a cropping/grazing rotation. Four pairs of soils differed in P fertilisation history, with each pair having one unfertilised soil and one with a known history of inorganic P fertilisation (Togari, Parkham, Elliot Research Station, and Elizabeth Town).
Table 2 shows the variation in total P and organic P concentrations of the whole soils. Total soil P concentration across all soils ranged from 239 to 15111mg/kg, of which inorganic P comprised 25-90% (84-12 559 mg/kg) and organic P 10-75% (72-2552mg/kg). Comparisons of the total, inorganic, and organic P fractions of each soil are shown in Fig. 1a.
In all cases, the P-treated soils (+P) had a higher total P concentration, of which the majority of the extra P was inorganic P, except Togari, where the +P treatment was higher in organic P. This is consistent with the results of McLaughlin et al. (1988), which showed, using isotope tracer experiments, that fertiliser was not quickly incorporated into organic P forms.
Extractability of inorganic and organic P
Total NaOH-EDTA extractable P ranged from 149 to 6222 mg/kg (Table 2) representing an extraction efficiency of 21 89%, with an average of 64% (Fig. 1b). Comparable extraction efficiencies have been reported for other [sup.31]P NMR studies. Dou et al. (2009) extracted 67-97% of total P from soils manured for 8-10 years, while Turner et al. (2003a) reported considerably lower extraction efficiencies (12-45%) for a range of cropped soils. Plots of whole soil and NaOH-EDTA extractable inorganic and organic P are shown in Fig. 2a, b. These exclude data for the Nowra sample (total P, 15 111 mg/kg; total extractable P, 6222 mg/kg) as they were several times higher than for any of the other soils presented in Fig. 2a.
The 1 : 1 line shown in Fig. 2a, b indicates the potential maximum (100%) extraction efficiency. It is clear that the average extraction efficiency of inorganic P was greater (82%) than for organic P (47%). This implies that, on average, around half of the organic P in these soils is not extractable and therefore cannot be identified by solution 3~p NMR analysis. However, it is likely that hydrolysis of organic P in the alkaline extract results in an overestimation of the extraction efficiency of inorganic P, consequently underestimating the extraction efficiency of organic P. Indeed, for four soils (Tocal, Wollongbar, Coonabarabran, and Collector) the apparent extraction efficiency of inorganic P was >100%. It should also be noted that the ashing technique used to determine the organic P content of the whole soils can be unreliable (Condron et al. 1990b), and this represents a second possible source of bias in the reported inorganic and organic P extraction efficiencies.
The Kadina soil had a total P extraction efficiency of only 21% (Fig. 1b), with the extraction efficiency of inorganic P particularly low (16%) compared with the other soils. The low extractability may be due to the calcareous nature of the Kadina soil, as high soil pH and the presence of calcium minerals has potential to complex P, rendering it hard to extract from soil (Cole et al. 1953; Celi et al. 2000). Across all soils, there was no noticeable relationship between P extractability and soil pH. The only other sample with a high soil pH was Nowra, and it too had a comparatively low P extraction efficiency (41%). Differences in extraction efficiency are also likely to have affected inorganic and organic P concentrations of the P-treated and P-untreated paired soil samples. The extraction efficiency of organic P tended to be greater in the treated soils than the untreated soils.
Assessment of the reliability of spectral deconvolution
Overlap of resonances in the 2-7ppm region of [sup.31]P NMR spectra of soil extracts, the region that encompasses the orthophosphate peak and numerous monoester peaks, hampers quantitative assessment of these P types. This problem can be overcome by using spectral deconvolution, which involves a numerical least-squares fit of the spectrum as the sum of multiple peaks of standard shape. Spectral deconvolution has been used in several [sup.31]P NMR soil studies (Turner et al. 2003d; McDowell and Stewart 2005; McDowell et al. 2005). Here, we use a modified version (see Materials and methods for description) that has been used only once before (Bunemann et al. 2008b). The main modification is the inclusion of a broad signal (3.95-5.45 ppm) in the fit. In a study that compared extracts of 'real' soils and 'model' soils (mixtures of pure sand and clay that were incubated with pure organic substrates, e.g. cellulose, starch, and glucose), Bunemann et al. (2008b) observed that the extracts of the model soils contained only sharp resonances. These resonances could be assigned to specific, simple organic P molecules. In contrast, the spectra of the real soils contained a broad signal in addition to the sharp resonances. This modified technique is yet to be applied to a variety of soils with considerably different inorganic and organic P concentrations.
The reliability of our methods for quantifying P species was tested by: (i) comparing orthophosphate concentrations determined using NMR with those determined colourimetrically, and (ii) using the spiking method of Smernik and Dougherty (2007) to provide an independent measure of the concentration of the monoester compound [beta]-glycerophosphate in two soil extracts.
Figure 3 shows the close relationship between NaOH-EDTA extractable orthophosphate concentrations measured using spectral deconvolution and those measured using molybdate colourimetry. Both methods gave very similar values (i.e. the line of regression is close to the 1 : 1 line in Fig. 3), but concentrations determined by colourimetry were generally slightly higher (on average 8%) than those determined using spectral deconvolution.
[FIGURE 1 OMITTED]
[FIGURE 2 OMITTED]
Spiking experiments were carried out on Elizabeth Town control and Parkham +P extracts. We calculated the [beta]-glycerophosphate concentration in the Elizabeth Town control sample to be 22 [+ or -] 1.9 mg P/kg using the spiking method and 21 mg P/kg using spectral deconvolution. Spiking the Parkham +P NaOH-EDTA soil extract revealed a [beta]-glycerophosphate concentration of 5 [+ or -] 1.8 mg P/kg, which was slightly less than that determined using deconvolution (10 mg P/kg). This suggests that deconvolution may overestimate the concentration of species with low abundances; however, the practical implication of such errors is relatively small.
Quantitative analysis and interpretation of [sup.31]P NMR spectra of NaOH-EDTA soil extracts
Spectra of eight of the soil extracts are not presented here as they have been reported elsewhere (Doolette et al. 2009) but have not been quantitatively analysed. For two of the soils (Kadina and Crystal Brook), we were unable to obtain an adequate [sup.31]P NMR spectrum. As discussed, the extraction efficiency was very low for the Kadina soil and this resulted in a low concentration of P in the NaOH-EDTA extract. This was also the case, but to a lesser extent, for the Crystal Brook soil, which suffered from the combination of a relatively low P concentration and relatively low extraction efficiency. However, it should be noted that we were able to obtain adequate NMR spectra on other soil extracts with similar, or even lower, P contents (e.g. Togari and Collector). This shows that the difficulties encountered in the analysis of these two soil extracts are not entirely explained by their low P content. The Kadina and Crystal Brook soils were also the only alkaline soils analysed, other than the Nowra soil, which had an extremely high P content (Tables 1 and 2). The potential link between difficulties in NMR analysis and high soil pH requires further investigation.
[FIGURE 3 OMITTED]
The [sup.31]P NMR spectra of the other NaOH-EDTA soil extracts, which consist of four pairs of control and P-treated soils, contain signals in four diagnostic chemical shift regions, orthophosphate (6.5-5.3 ppm), orthophosphate monesters (5.3-2.6 ppm), orthophosphate diesters (2.0 to -1.0 ppm), and pyrophosphate (-4.5 to -5.5 ppm) (Fig. 4). An expansion of the orthophosphate and orthophosphate monoester regions is shown in Fig. 5. Equivalent spectra for the other soils can be found in Doolette et al. (2009). The two regions that comprised the majority of signal across all soils were orthophosphate and orthophosphate monoester, together equating to 94-98% of total NaOH-EDTA extractable P. Orthophosphate diesters and pyrophosphate were minor components, comprising only 2-6% of total NaOH-EDTA extractable P. When comparing the control and treated soils, the only noticeable differences were in the orthophosphate region. The differences between the control and treated soils in the orthophosphate monoester, orthophosphate diester, and pyrophosphate chemical shift regions were relatively small. However, it might be expected that with continued P fertiliser application, soil organic P could increase (Condron et al. 1985; McLaughlin et al. 1988).
[FIGURE 4 OMITTED]
Quantitative distributions of P forms determined for all soil extracts are presented in Table 3. The percentage distribution of each P species, excluding orthophosphate in relation to total non-orthophosphate P, is presented in Fig. 1c.
Orthophosphate (5.69-5.78 ppm) produced the most intense resonance in each spectrum and is the most abundant P species detected in every soil extract. Orthophosphate concentrations ranged from 86 to 5622 mg/kg, equivalent to 37-90% of total NaOH-EDTA extractable P (Table 3).
Orthophosphate monoesters represented the next-largest P class and accounted for 7 55% of total NaOH-EDTA extractable P. The majority of the orthophosphate monoester was assigned to a broad signal, centred at 3.95-5.45 ppm. This signal is present in all spectra but is more easily discerned in the spectra that contained less-intense sharp resonances, e.g. Parkham, Elliot, and Elizabeth Town (Fig. 5). The broad monoester signal represented 31-355mgP/kg (equivalent to 2-39% of total extractable P or 32-86% of total orthophosphate monoesters) (Table 3). The absolute P concentration and proportion of organic or total extractable P of the broad resonance did not appear to be associated with any of the climatic, chemical, or physical properties measured, but more soils need to be studied to further examine this possibility.
[FIGURE 5 OMITTED]
Most previous attempts at quantifying monoester P species using deconvolution analysis of [sup.31]P NMR spectra of soil extracts have not included a broad peak in the fit, and consequently, signal from this broad peak is assigned to species that give rise to
the sharper peaks in the spectrum. More specifically, we have previously tested this method of deconvolution and shown that, by not accounting for the broad peak, phytate concentrations were overestimated by 54% (Doolette et al. 2010). Dougherty et al. (2007) suggested that this broad signal is due to large, complex, humic molecules, as opposed to smaller, specific monoester P compounds that produce sharp resonances. This explanation is supported by Bunemann et al. (2008b), who found this broad signal to be absent in model soils that have had insufficient time for extensive humification to have occurred. Earlier evidence for the dominance of humic P can be seen in the work of Anderson and Hance (1963), who found evidence of phytate and other organic phosphate molecules bound in a complex or compound that contained appreciable amounts of carbohydrates and proteins. Omotoso and Wild (1970) also considered that phytate existed in an organic complex combined with other organic P compounds.
The concentrations of humic organic P reported here are consistent with those determined by different methods in previous studies of volcanic and alpine soils. The humic fractionation of volcanic soils showed that P associated with humic and fulvic acids accounted for 32-75% and 51-68% of organic P, respectively (Borie et al. 1989; Escudey et al. 2001; Borie and Rubio 2003). In alpine soils, 52-90% of alkali-extractable organic P was associated with humic acids with phosphate monoesters the dominant P species (Makarov et al. 1997). Furthermore, using [sup.3t]P NMR, He et al. (2006) examined the spectral and chemical characteristics of humic substances and also found phosphate monoesters to be the dominant P species. More importantly, they were unable to identify individual monoester P compounds, as the spectra had no sharp resonances but instead a broad signal in the monoester region. We conclude that failing to include the broad signal in the deconvolution procedure would result in the overestimation of the concentration of species that give rise to the sharp monoester resonances.
The remainder of orthophosphate monoester P was assigned to the [sup.3t]P NMR signal contained within the sharp resonances. Phytate (4.17-4.28, 4.35-4.40, 4.71-4.82ppm), [alpha]-glycerophosphate (4.95-4.98 ppm), and [beta]-glycerophosphate (4.59-4.64ppm) were the most prominent sharp resonances. Phytate was present in all soil extracts except for the Elliot control and +P samples. Concentrations in the other soils ranged from 5 to 86 mg P/kg (equivalent to up to 9% of total NaOH-EDTA extractable P) (Table 3). Phytate concentrations averaged just 3% of total NaOH-EDTA extractable P and 13% of NaOH-EDTA extractable organic P. Both or- and [beta]-glycerophosphate were present in all [sup.3t]P NMR spectra but were only a very minor component in some extracts; [beta]-glycerophosphate comprised <1% of total NaOH-EDTA extractable P in the Wollongbar extract, [alpha]-glycerophosphate comprised <1% in the Coonabarabran extracts, and combined, [alpha]- and [beta]-glycerophosphate comprised <1% in both the Somersby A and B samples. In the remaining spectra, [alpha]- and [beta]-glycerophosphate resonances were more prominent; [alpha]-glycerophopshate concentrations ranged from 3 to 80mgP/kg (1-5% total NaOH-EDTA extracable P) and [beta]-glycerophosphate ranged from 7 to 80 mg P/kg (1-5% total NaOH-EDTA extractable P) (Table 3). In total, [alpha]- and [beta]-glycerophosphate comprised up to a maximum of 10% of extractable P.
Phytate in soils primarily originates from the decay of crop seeds and fruits (Lott et al. 2000). Relatively high phytate concentrations in soils can result from the addition of animal manures, e.g. poultry and swine manure, as phytate passes though the digestive tracts of these animals since they lack the ability to break it down (Selinger et al. 1996). This may explain the relatively high concentration of phytate in the Tocal soil extract, which had received poultry manure.
The remainder of the soils are predominantly dairy pastures where the strongest monoester resonances have been assigned to [alpha]- and [beta]-glycerophosphate, which are most likely products of the alkaline hydrolysis of phospholipids (Doolette et al. 2009) rather than being actual soil constituents themselves. They are likely to be derived mainly from microbial biomass, as Bunemann et al. (2008a) showed that [alpha]- and [beta]-glycerophosphate were the major forms of monoester P identified in NaOH-EDTA extracts of bacterial and fungal cultures. Microbial biomass is often quite high in dairying soils due to high organic carbon and moisture regimes (Burkitt et al. 2007).
Other orthophosphate monoester compounds present in the soil extracts were detected in much smaller quantities. For example, scyllo-inositol hexakisphosphate (3.85-3.91 ppm), a common stereoisomer of phytate, was present in all soil extracts at concentrations ranging from 2 to 20 mg P/kg or <1-4% of total extractable P (Table 3). Despite the widespread occurrence of scyllo-inositol hexakisphosphate in soils (McKercher and Anderson 1968; Turner et al. 2005; Turner 2007; Murphy et al. 2009) its origins are still unknown. Furthermore, there are conflicting reports regarding the bioavailability or susceptibility of scyllo-inositol hexakisphosphate to hydrolytic attack by phytase (Cosgrove 1966, 1970; Turner et al. 2005).
Trace amounts of AMP ([less than or equal to] 2% extractable P; 4.47-4.52 ppm) were present in all soil extracts except Somersby B. This resonance is also likely to include other ribonucleotides (RNA monomers) and deoxyribonucleotides (DNA monomers), as they resonate at very similar chemical shifts (Turner et al. 2003b; Bunemann et al. 2008a; Doolette et al. 2009). However, due to the instability of RNA it is not possible to conclude whether the nucleotides are present in the soil or are products of alkaline hydrolysis during the sample extraction/preparation stages.
Five other well-resolved orthophosphate monoester P resonances (5.23-5.24, 5.10-5.11, 4.064.17, 3.04ppm) were identified and are labelled as unknown compounds a-e in Table 3. In total, these unknown compounds accounted for [less than or equal to] 3% of total extractable P, but in most cases <1%. Two further unidentified resonances between 6.34 and 6.58ppm (comprising <1% of total extractable P) have previously been assigned as aromatic orthophosphate diesters in the NaOH extracts of Scottish mineral soils by Bedrock et al. (1994). Aromatic orthophosphate diesters are generally thought to be associated with cold and wet environments. They have been identified in grassland soils from Russia and New Zealand (Amelung et al. 2001; McDowell 2003) and pasture soils from New Zealand, England, and Wales (Turner et al. 2003c; McDowell et al. 2005). However, little is known about the possible structures or origins of these compounds, and the chemical shifts of aromatic orthophosphate diesters can vary widely (Turner et al. 2003b; Makarov et al. 2004). It has been proposed that it is more likely that these signals actually represent stereo-isomers of inositol phosphates (Turner et al. 2005).
Orthophosphate diesters represent the next largest class of P compounds and were present in all soil extracts, with concentrations ranging from 4 to 87 mg/kg or 1-5% of total [sup.31]P NMR signal (Table 3). Orthophosphate diesters comprised only DNA with other potential diester P forms such as phospholipids and RNA absent from our [sup.31]P NMR spectra. Phospholipids and RNA are rarely detected in soils (Makarov et al. 2002; Turner et al. 2007), despite their rate of addition to soils being much greater than that of other organic P forms (Anderson 1980). This is likely governed by the low stability of these compounds in the soil and in alkaline solutions (Makarov et al. 2002; Turner et al. 2003b) and the low aqueous solubility of phospholipids' (Doolette et al. 2009).
Low concentrations of diester P reported in some soils have been attributed to the preferential mineralisation of diester P compared with other more stable organic P forms (Hawkes et al. 1984; Condron et al. 1985; Condron et al. 1990a) and are considered to be linked to nutrient turnover, with diester P representing a labile pool of soil organic P (Turner et al. 2003c). In particular, cell-free RNA and DNA are highly susceptible to degradation due to the presence of active nucleases in soils (Paul and Cark 1996). However, extracellular DNA can strongly sorb to sand, clay minerals, and humic substances, providing protection from degradation by nucleases compared with free DNA in solution (Pietramellara et al. 2009). This may be one of the factors contributing to the presence of DNA in preference to other diester P forms.
It should be noted that the concentrations of diester P in this study (as in other 31p NMR studies) are likely to be underestimated due to its degradation in alkaline solution during extraction. Turner et al. (2003b) showed RNA to be highly unstable in NaOH-EDTA, completely degrading to orthophosphate monoesters within 24 h, and this is consistent with the findings of Makarov et al. (2002).
Phospholipids are also susceptible to hydrolysis, although the rate of degradation varies between phospholipids. One of the most common phospholipids, phosphatidyl choline (lecithin), completely degrades in NaOH-EDTA within 24 h (Turner et al. 2003b) to [alpha]- and [beta]-glycerophosphate (Doolette et al. 2009). In contrast, DNA is more stable (Turner et al. 2003b) and can remain intact in alkaline solutions for at least 24 h (Makarov et al. 2002). Low concentrations of diester P may also be due to a combination of the low aqueous solubility of phospholipids in NaOH-EDTA (Doolette et al. 2009) and low organic P extractability. On average only 47% of total organic P was extracted, which suggests some forms of diester P that are not extracted from the soil.
Pyrophosphate was present in all soil extracts at concentrations ranging from 1 to 62 mg/kg (up to 5% of total NaOH-EDTA extractable P). No higher polyphosphates, which produce resonances in the range -18 to -22 ppm, were detected in any of the soil extracts. Pyrophosphate concentrations reported here are similar to those reported elsewhere: 1-7% of total extractable P in pasture soils (Turner et al. 2003c), 0.5-4.3% in cropping soils (Turner et al. 2003a), <1-4.85% in manure-treated soils (Dou et al. 2009), and on average 3% in grassland soils (Murphy et al. 2009). Inorganic polyphosphates, of which pyrophosphate is the simplest form, are believed to accumulate in microbial biomass as a P storage form. The factors controlling their abundance remain unclear (Blanchar and Hossner 1969; Turner et al. 2003a).
The results presented here suggest that the largest portion of organic P in some Australian soils is bound in complex humic P molecules and not in smaller specific compounds such as phytate. This not only challenges our understanding of soil organic P cycling but has implications for the way in which methods are developed to increase P availability in the soil profile. In terms of sustainable P management, it is likely that a slightly new approach is required, especially in low-input and organic agricultural systems where recalcitrant or poorly available forms of P such as phytate are seen as potentially available forms of organic P (Guppy and McLaughlin 2009). There is likely to be great promise for plant-based approaches to increase P efficiency in agricultural systems (Richardson et al. 2009). However, some of the most effective strategies for increased P availability may not be through techniques that target specific soil organic P compounds, such as the genetic modification of plants with increased phytase activity (George et al. 2004, 2005) or the direct application of manufactured extracellular enzymes to increase mineralisation of inositol P (Conyers and Moody 2009). Instead, greater success could come from techniques isolating genes in plants that aid or alter root development (Grierson et al. 2001; Williamson et al. 2001), thereby increasing the chances of P uptake, or focusing on the exudation of organic anions with a known capability of mobilising P bound in humic-metal complexes (Gerke 1993). It is doubtful that a single strategy that increases the availability of organic P would be applicable to all soils. However, we suggest that the development of strategies to increase uptake of P by plants and crops should not be focused only on increasing the availability of P contained in small, supposedly recalcitrant compounds, such as phytate. Consideration should also be given to the availability of P in larger, complex molecules.
We have used an improved method for assessing the speciation of soil P using solution [sup.31]P NMR and spectral deconvolution. This method not only provided a more detailed characterisation of organic P forms in Australian soils than has hitherto been undertaken, but also provided a more accurate estimation of the quantity of these forms by minimising the potential for overestimation of the P concentrations of simple orthophosphate monoesters. The [sup.31]P NMR spectra of 18 NaOH-EDTA soil extracts revealed that a large proportion of organic P is present in large, complex molecules such as humic acids. Phytate was the most abundant identifiable organic P compound, but only comprised a maximum of 10% of total extractable P. In most cases, humic organic P represented more P than all of the small discrete P molecules combined. These findings have implications for attempts to increase the availability of stabilised forms of organic P. Consideration should be given to further investigation of the presence and availability of P in large, complex molecules, rather than solely developing methods to increase the availability of individual P compounds (such as phytate) that are present in the soils at much lower concentrations.
Amelung W, Rodionov A, Urusevskaja IS, Haumaier L, Zech W (2001) Forms of organic phosphorus in zonal steppe soils of Russia assessed by [sup.31]P NMR. Geoderma 103, 335 350. doi:10.1016/S0016-7061(01) 00047-7
Anderson G (1980) Assessing organic phosphorus in soils. In 'Role of phosphorus in agriculture'. (Eds FE Khasawneh, EC Sample, EJ Kamprath) pp. 411-428. (ASA: Madison, WI)
Anderson G, Hance RJ (1963) Investigation of an organic phosphorus component of fulvic acid. Plant and Soil 19, 296 303. doi:10.1007/ BF01379483
Beadle NCW (1962) An alternative hypothesis to account for the generally low phosphate content of Australian soils. Australian Journal of Agricultural Research 13, 434-442. doi: 10.1071/AR9620434
Bedrock CN, Cheshire MV, Chudek JA, Goodman BA, Shand CA (1994) Use of [sup.31]P-NMR to study the forms of phosphorus in peat soils. The Science of the Total Environment 152, 1-8. doi:10.1016/0048-9697(94) 90545-2
Blanchar RW, Hossner LR (1969) Hydrolysis and sorption of ortho-, pryo-, tripoly-, and trimetaphosphate in 32 midwestern soils. Soil Science Society of America Journal 33, 622-625. doi:10.2136/sssaj1969. 03615995003300040037x
Bolland MDA, Gilkes RJ (1998) The chemistry and agronomic effectiveness of phosphate fertilizers. Journal of Crop Production 1, 139-163. doi: 10.1300/J144v01n02_07
Borie F, Rubio R (2003) Total and organic phosphorus in Chilean volcanic soils. Gayana Botany 60, 69-78.
Borie F, Zunino H, Martanez L (1989) Macromolecule P associations and inositol phosphates in some Chilean volcanic soils of temperate regions. Communications in Soil Science and Plant Analysis 20, 1881-1894. doi: 10.1080/00103628909368190
Bunemann EK, Heenan DP, Marschner P, McNeill A (2006) Long-term effects of crop rotation, stubble management and tillage on soil phosphorus dynamics. Australian Journal of Soil Research 44, 611-618. doi:10.1071/SR05188
Bunemann EK, Smeruik R J, Doolette AL, Marschner P, Stonor R, Wakelin SA, McNeill AM (2008a) Forms of phosphorus in bacteria and fungi isolated from two Australian soils. Soil Biology & Biochemistry 40, 1908-1915. doi:10.1016/j.soilbio.2008.03.017
Bunemann EK, Smernik RJ, Marschner P, McNeill AM (2008b) Microbial synthesis of organic and condensed forms of phosphorus in acid and calcareous soils. Soil Biology & Biochemistry 40, 932-946. doi:10.1016/ j.soilbio.2007.11.012
Burkitt LL, Small DR, McDonald JW, Wales W J, Jenkin ML (2007) Comparing irrigated biodynamic and conventionally managed dairy farms. I. Soil and pasture properties. Australian Journal of Experimental Agriculture 47, 479488. doi:10.1071/EA05196
Cade-Menun BJ, Preston CM (1996) A comparison of soil extraction procedures for [sup.31]P NMR spectroscopy. Soil Science 161, 770-785. doi: 10.1097/00010694-199611000-00006
Celi L, Lamacchia S, Barberis E (2000) Interaction of inositol phosphate with calcite. Nutrient Cycling in Agroecosystems 57, 271-277. doi: 10.1023/A: 1009805501082
Cole CV, Olsen SR, Scott CO (1953) The nature of phosphate sorption by calcium carbonate. Soil Science Society of America Journal 17, 352-356. doi: 10.2136/sssaj1953.03615995001700040013x
Condron LM, Goh KM, Newman RH (1985) Nature and distribution of soil phosphorus as revealed by a sequential extraction method followed by [sup.31]P nuclear magnetic resonance analysis. Journal of Soil Science 36, 199-207. doi:10.111 l/j.1365-2389.1985.tb00324.x
Condron LM, Frossard E, Tiessen H, Newman RH, Stewart JWB (1990a) Chemical nature of organic phosphorus in cultivated and uncultivated soils under different environmental conditions. Journal of Soil Science 41, 41-50. doi:10.1111/j.1365-2389.1990.tb00043.x
Condron LM, Moir JO, Tiessen H, Stewart JWB (1990b) Critical evaluation of methods for determining total organic phosphorus in tropical soils. Soil Science Society of America Journal 54, 1261-1266. doi:10.2136/ sssaj1990.03615995005400050010x
Conyers MK, Moody PW (2009) A conceptual framework for improving the P efficiency of organic farming without inputs of soluble P fertiliser. Crop & Pasture Science 60, 100-104. doi:10.1071/CP06327
Cornish PS (2009a) Phosphorus management on extensive organic and low-input farms. Crop & Pasture Science 60, 105-115. doi:10.1071/ CP07134
Cornish PS (2009b) Research direction: Improving plant uptake of soil phosphorus, and reducing dependency on input of phosphorus fertiliser. Crop & Pasture Science 60, 190-196. doi:10.1071/CP08920
Cosgrove DJ (1962) Forms of inositol hexaphosphate in soils. Nature 194, 1265- 1266. doi:10.1038/1941265a0
Cosgrove DJ (1963) The chemical nature of soil organic phosphorus. Australian Journal of Soil Research 1, 203-214. doi:10.1071/ SR9630203
Cosgrove DJ (1966) Synthesis of the hexaphosphates of myo-, scyllo-, neo-, and D-inositol. Journal of the Science of Food and Agriculture 17, 550-554. doi: 10.1002/jsfa.2740171206
Cosgrove DJ (1970) Inositol phosphate phosphatases of microbial origin. Inositol phosphate intermediates in the dephosphorylation of the hexaphosphate of myo-inostiol, scyllo-inositol and D-chiro-inositol by bacterial (Pseudomonas sp.) phytase. Australian Journal of Biological Sciences 23, 1207-1220.
Cosgrove DJ, Tate ME (1963) Occurrence of neo-inositol hexaphosphate in soil. Nature 200, 568-569. doi:10.1038/200568b0
Doolette AL, Smernik RJ, Dougherty WJ (2009) Spiking improved solution phosphorus-31 nuclear magnetic resonance identification of soil phosphorus compounds. Soil Science Society of America Journal 73, 919-927. doi:10.2136/sssaj2008.0192
Doolette AL, Smernik RJ, Dougherty WJ (2010) Rapid decomposition of phytate applied to a calcareous soil demonstrated by a solution [sup.31]P NMR study. European Journal of Soil Science 61, 563-575. doi:10.1111/ j.1365-2389.2010.01259.x
Dou Z, Ramberg CF, Toth JD, Wang Y, Sharpley AN, Boyd SE, Chen CR, Williams D, Xu ZH (2009) Phosphorus speciation and sorption-desorption characteristics in heavily manured soils. Soil Science Society of America Journal 73, 93-101. doi:10.2136/sssaj2007.0416
Dougherty WJ, Smernik RJ, Bunemann EK, Chittleborough DJ (2007) On the use of HF pre-treatment of soils for [sup.31]P NMR analyses. Soil Science Society of America Journal 71, 1111-1118. doi:10.2136/ sssaj2006.0300
Escudey M, Galindo G, Forster JE, Briceno M, Diaz P, Chang A (2001) Chemical forms of phosphorus of volcanic ash-derived soils in Chile. Communications in Soil Science and Plant Analysis 32, 601-616. doi:10.1081/CSS-100103895
Evans J, Condon J (2009) New fertiliser options for managing phosphorus for organic and low-input farming systems. Crop & Pasture Science 60, 152-162. doi:10.1071/CP07153
George TS, Richardson AE, Hadobas PA, Simpson RJ (2004) Characterization of transgenic Trifolium subterraneum L. which expresses phyA and releases extracellular phytase: growth and P nutrition in laboratory media and soil. Plant, Cell & Environment 27, 1351-1361. doi:10.1111/j.1365-3040.2004.01225.x
George TS, Simpson RJ, Hadobas PA, Richardson AE (2005) Expression of a fungal phytase gene in Nicotiana tabacum improves phosphorus nutrition of plants grown in amended soils. Plant Biotechnology Journal 3, 129-140. doi: 10.1111/j.1467-7652.2004.00116.x
Gerke J (1993) Solubilization of Fe (III) from humic-Fe complexes, humic/ Fe-oxide mixtures and from poorly ordered Fe-oxide by organic acids consequences for P adsorption. Zeitschrift fur Pflanzenernahrung und Bodenkunde 156, 253 257. doi: 10.1002/jpln.19931560311
Grierson CS, Parker JS, Kemp AC (2001) Arabidopsis genes with roles in root hair development. Journal of Plant Nutrition and Soil Science 164, 131-140. doi:10.1002/1522-2624(200104)164:2<131::AID-JPLN131> 3.0.CO;2-2
Guppy C, McLaughlin MJ (2009) Options for increasing the biological cycling of phosphorus in low-input and organic agricultural systems. Crop and Pasture Science 60, 116-123. doi:10.1071/CP07157
Hawkes GE, Powlson DS, Randall EW, Tate KR (1984) A P-31 nuclear magnetic-resonance study of the phosphorus species in alkali extracts of soils from long-term field experiments. Journal of Soil Science 35, 35-45. doi:10.1111/j.1365-2389.1984.tb00257.x
He Z, Ohno T, Cade-Menun BJ, Erich MS, Honeycutt CW (2006) Spectral and chemical characterization of phosphates associated with humic substances. Soil Science Society of America Journal 70, 1741-1751. doi:10.2136/sssaj2006.0030
Holford ICR (1997) Soil phosphorus: its measurement, and its uptake by plants. Australian Journal of Soil Research 35, 227-240. doi: 10.1071/ S96047
Irving GCJ, Cosgrove DJ (1982) The use of gas-liquid chromatography to determine the proportions of inositol isomers present as pentakis- and hexakisphosphates in alkaline extracts of soils. Communications in Soil Science and Plant Analysis 13, 957-967. doi:10.1080/00103628 209367324
Isbell RF (1996) 'The Australian Soil Classification.' (CSIRO Publishing: Melbourne)
Lott JNA, Ockenden I, Raboy V, Batten GD (2000) Phytic acid and phosphorus in crop seeds and fruits: a global estimate. Seed Science Research 10, 11-33.
Makarov MI, Malysheva TI, Haumaier L, Alt HG, Zech W (1997) The forms of phosphorus in humic and fulvic acids of a toposequence of alpine soils in the northern Caucasus. Geoderma 80, 61 73. doi: 10.1016/ S0016-7061(97)00049-9
Makarov MI, Haumaier L, Zech W (2002) Nature of soil organic phosphorus: an assessment of peak assignments in the diester region of [sup.31]P NMR spectra. Soil Biology & Biochemistry 34, 1467-1477. doi:10.1016/S0038-0717(02)00091-3
Makarov MI, Haumaier L, Zech W, Malysheva TI (2004) Organic phosphorus compounds in particle-size fractions of mountain soils in the northwestern Caucasus. Geoderma 118, 101-114. doi:10.1016/ S0016-7061(03)00187-3
McDowell R (2003) Identification of phosphorus species in extracts of soils with contrasting management histories. Communications in Soil Science and Plant Analysis 34, 1083-1095. doi:10.1081/CSS-120019111
McDowell RW, Stewart I (2005) An improved technique for the determination of organic phosphorus in sediments and soils by [sup.31]P nuclear magnetic resonance spectroscopy. Chemistry and Ecology 21, 11-22. doi: 10.1080/02757540512331334942
McDowell RW, Condron LM, Stewart l, Cave V (2005) Chemical nature and diversity of phosphorus in New Zealand pasture soils using [sup.31]P nuclear magnetic resonance spectroscopy and sequential fractionation. Nutrient Cycling in Agroecosystems 72, 241-254. doi: 10.1007/s10705-005-2921-8
McKercher RB, Anderson G (1968) Characterization of inositol penta- and hexaphosphate fractions of a number of Canadian and Scottish soils. Journal of Soil Science 19, 302-310. doi:10.1111/j.1365-2389.1968. tb01542.x
McLaughlin MJ, Alston AM, Martin JK (1988) Phosphorus cycling in wheat pasture rotations. Ill. Organic phosphorus turnover and phosphorus cycling. Australian Journal of Soil Research 26, 343-353. doi:10.1071/SR9880343
McLaughlin MJ, Fillery IR, Till AR (1991) Operation of the phosphorus, sulphur and nitrogen cycles. In "Australia's renewable resources: sustainability and global change'. (Eds RM Gifford, MM Barson) pp. 67-116. (Australian Government Printing Services: Canberra, ACT)
Murphy J, Riley JP (1962) A modified single solute method for the determination of phosphate in natural waters. Analytica Chimica Acta 27, 31-36. doi: 10.1016/S0003-2670(00)88444-5
Murphy PNC, Bell A, Turner BL (2009) Phosphorus speciation in temperate basaltic grassland soils by solution [sup.31]P NMR spectroscopy. European Journal of Soil Science 60, 638-651. doi: 10.1111/j.1365-2389.2009. 01148.x
Norrish K, Rosser H (1983) Mineral phosphate. In 'Soils: an Australian viewpoint', pp. 335-361. (CSIRO: Melbourne/Academic Press: London)
Omotoso TI, Wild A (1970) Occurrence of inositol phosphates and other organic phosphate components in an organic complex. Journal of Soil Science 21, 224-232. doi:10.1111/j.1365-2389.1970.tb01171.x
Paul EA, Cark FE (1996) 'Soil microbiology and biochemistry.' (Academic Press, Inc.: San Diego, CA)
Pietramellara G, Ascher J, Borgogni F, Ceccherini MT, Guerri G, Nannipieri P (2009) Extracellular DNA in soil and sediment: fate and ecological significance. Biology and Fertility of Soils 45, 219-235. doi:10.1007/ s00374-008-0345-8
Reddy NR, Pierson MD, Sathe SK, Salunkhe DK (1989) 'Phytates in cereals and legumes.' (CRC Press: Boca Raton, FL)
Richardson AE, Hocking P J, Simpson R J, George TS (2009) Plant mechanisms to optimise access to soil phosphorus. Crop & Pasture Science 60, 124-143. doi:10.1071/CP07125
Saunders WM, Williams EG (1955) Observations on the determination of organic phosphorus in soils. Journal of Soil Science 6, 254-267. doi: 10.1111/j.1365-2389.1955.tb00849.x
Selinger LB, Forsberg CW, Cheng KJ (1996) The rumen: a unique source of enzymes for enhancing livestock production. Anaerobe 2, 263-284. doi:10.1006/anae.1996.0036
Sharpley AN, Chapra SC, Wedepohl R, Sims JT, Daniel TC, Reddy KR (1994) Managing agricultural phosphorus for protection of surface waters: issues and options. Journal of Environmental Quality 23, 437-451. doi: 10.2134/jeq 1994.00472425002300030006x
Smernik R J, Dougherty WJ (2007) Identification of phytate in phosphorus--31 nuclear magnetic resonance spectra the need for spiking. Soil Science Society of America Journal 71, 1045-1050. doi:10.2136/ sssaj2006.0295
Smith SE, Smith FA, Jakobsen I (2003) Mycorrhizal fungi can dominate phosphate supply to plants irrespective of growth responses. Plant Physiology 133, 16-20. doi:10.1104/pp.103.024380
Steward JH, Tate ME (1971) Gel chromatography of soil organic phosphorus. Journal of Chromatography A 60, 75-82. doi:10.1016/ S0021-9673(00)95531-6
Turner BL (2007) Inositol phosphates in soil: amounts, forms and significance of the phosphorylated inositol stereoisomers. In 'Inositol phosphates: linking agriculture and the environment'. (Eds BL Turner, AE Richardson, EJ Mullaney) pp. 186-206. (CABI: Wallingford, UK)
Turner BL, Cade-Menun BJ, Westermanm DT (2003a) Organic phosphorus composition and potential bioavailability in semi-arid arable soils of the western United States. Soil Science Society of America Journal 67, 1168-1179. doi:10.2136/sssaj2003.1168
Turner BL, Mahieu N, Condron LM (2003b) Phosphorus-31 nuclear magnetic resonance spectral assignments of phosphorus compounds in soil NaOH-EDTA extracts. Soil Science Society of America Journal 67, 497-510. doi:10.2136/sssaj2003.0497
Turner BL, Mahieu N, Condron LM (2003c) The phosphorus composition of temperate pasture soils determined by NaOH-EDTA extraction and solution [sup.31]P NMR spectroscopy. Organic Geochemistry 34, 1199-1210. doi:10.1016/S0146-6380(03)00061-5
Turner BL, Mahieu N, Condron LM (2003d) Quantification of myo-inositol hexakisphosphate in alkaline soil extracts by solution [sup.31]P NMR spectroscopy and spectral deconvolution. Soil Science 168, 469-478. doi:10.1097/00010694-200307000-00002
Turner BL, Mahieu N, Condron LM, Chen CR (2005) Quantification and bioavailability of scyllo-inositol hexakisphosphate in pasture soils. Soil Biology & Biochemistry 37, 2155-2158. doi:10.1016/j.soilbio. 2005.03.005
Turner BL, Condron L, Richardson S, Peltzer D, Allison V (2007) Soil organic phosphorus transformations during pedogenesis. Ecosystems 10, 1166-1181. doi:10.1007/s10021-007-9086-z
Walkley A (1947) A critical examination of a rapid method for determining organic carbon in soil-effect of variations in digestion concentration and of inorganic soil constituents. Soil Science 63, 251-264. doi:10.1097/ 00010694-194704000-00001
Williams CH, Anderson G (1968) Inositol phosphates in some Australian soils. Australian Journal of Soil Research 6, 121-130. doi:10.1071/ SR9680121
Williamson LC, Ribrioux SPCP, Fitter AH, Leyser HMO (2001) Phosphate availability regulates root system architecture in Arabidopsis. Plant Physiology 126, 875-882. doi:10.1104/pp.126.2.875
Manuscript received 21 April 2010, accepted 2 September 2010
A. L. Doolette (A,C), R.J. Smernik (A), and W. J. Dougherty (B)
(A) School of Agriculture, Food and Wine, University of Adelaide, Waite Campus, Urrbrae, SA 5064, Australia. (B) Department of Primary Industries, Industry and Investment New South Wales, Locked Bag 4, Richmond, NSW 2753, Australia. (C) Corresponding author. Email: email@example.com
Table 1. Selected physical, climatic, and chemical properties of the 18 Australian soils used in this study MAT, Mean annual temperature; MAP, mean annual precipitation; OC, organic carbon, pH in water (1:5 soil/water ratio) Soil Land use/treatment Nowra (A) Dairy pasture + effluent Casino (A) Dairy pasture Collector (A) Sheep pasture Coonabarabran (A) Horticulture Wollongbar (A) Dairy pasture Tocal (A) Dairy pasture + poultry litter Somersby A (A) Organic vegetable Somersby B (A) Conventional vegetable Kadina Cropping Crystal Brook Cropping/grazing rotation Togari control Dairy pasture Togari +P Dairy pasture Parkham control Dairy pasture Parkham +P Dairy pasture Elliot control Dairy pasture Elliot +P Dairy pasture Elizabeth Town control Dairy pasture Elizabeth Town +P Dairy pasture Soil MAT ([degrees]C) MAP (mm) pH OC (g/kg) Nowra (A) 16 1110 8.1 38 Casino (A) 20 1096 6.5 42 Collector (A) 14 644 5.0 27 Coonabarabran (A) 16 748 4.5 16 Wollongbar (A) 19 1793 5.2 38 Tocal (A) 18 922 6.1 43 Somersby A (A) 17 1318 6.7 11 Somersby B (A) 17 1318 5.9 10 Kadina 23 387 8.3 19 Crystal Brook 23 471 7.9 17 Togari control 15 1243 5.5 87 Togari +P 15 1243 5.3 87 Parkham control 16 947 5.7 28 Parkham +P 16 947 5.9 28 Elliot control 16 1188 5.2 95 Elliot +P 16 1188 5.3 95 Elizabeth Town control 16 947 5.4 60 Elizabeth Town +P 16 947 5.3 60 Soil Soil texture Soil Classif. (B) Nowra (A) Clay loam Chromosol Casino (A) Silty clay Vertosol Collector (A) Loam Chromosol Coonabarabran (A) Loamy sand Podosol Wollongbar (A) Clay Ferrosol Tocal (A) Loamy sand Chromosol Somersby A (A) Sandy loam Tenosol Somersby B (A) Loamy sand Tenosol Kadina Loamy sand Calcarosol Crystal Brook Sandy loam Calcarosol Togari control Loamy sand Hydrosol Togari +P Loamy sand Hydrosol Parkham control Clay loam/loam Vertosol Parkham +P Clay loam/loam Vertosol Elliot control Silty loam Ferrosol Elliot +P Silty loam Ferrosol Elizabeth Town control Silty clay loam Ferrosol Elizabeth Town +P Silty clay loam Ferrosol (A) Data reproduced from Doolette et al. (2009). (B) Australian Soil Classification (Isbell 1996). Table 2. Phosphorus characteristics (mg/kg) of the whole soil and NaOH-EDTA extractable P fractions Values in parentheses are percentages of total NaOH-EDTA extractable P Soil Whole soil P NaOH-EDTA Total Organic extractable P Nowra (A) 15111 2552 6222 (41) Casino (A) 1576 423 1184 (75) Collector (A) 257 172 166 (64) Coonabarabran (A) 239 144 149 (63) Wollongbar (A) 1113 650 882 (79) Tocal (A) 1214 564 887 (73) Somersby A (A) 743 90 542 (73) Somersby B (A) 702 72 627 (89) Kadina 844 200 178 (21) Crystal Brook 571 214 335 (59) Togari control 465 220 340 (73) Togari +P 665 490 336 (51) Parkham control 627 470 355 (60) Parkham +P 1386 465 950 (69) Elliot control 1106 816 677 (61) Elliot +P 2526 776 1912 (76) Elizabeth Town control 1910 1094 1066 (56) Elizabeth Town + P 2293 963 1443 (63) (A) Data reproduced from Doolette et al. (2009). Table 3. Concentrations (mg-kg) of phosphorus forms and their percentages (in parentheses) of NaOH-EDTA extractable total P as detected by 31P NMR and quantified using spectral deconvolution Ortho-P, Orthophosphate; GP, glycerophosphate; AMP, adenosine-5'-monophosphate; s-IHP, scyllo-inositol hexakisphosphate; unknown monoesters (a-e) are the sum of five individual resonances with the following chemical shift ranges (ppm): (a) 5.23-5.24, (b) 5.10-5.11, (c) 4.06-4.17, (d) 3.04, (e) 2.89; Pyro-P, pyrophosphate; detected n.d., not Soil Unknown P Ortho-P [alpha]-GP Chemical shift 6.53-6.58 5.69-5.78 4.95-4.98 ranges (ppm): 6.34-6.36 Nowra 0 (0) 5622 (90) 80 (1) Casino 0 (0) 904 (76) 22 (2) Collector 0 (0) 86 (52) 8 (5) Coonabarabran 1 (<1) 94 (63) 3 (2) Wollongbar 0 (0) 455 (52) 6 (<1) Tocal 2 (<1) 595 (67) 17 (2) Somersby A 0 (0) 460 (85) 2 (<1) Somersby B 0 (0) 509 (81) 1 (<1) Kadina n.d. 104 (58) n.d. Crystal Brook n.d. 249 (74) n.d. Togari control 3 (<1) 134 (39) 16 (5) Togari +P 0 (0) 176 (52) 13 (4) Parkham control 4 (1) 132 (37) 12 (4) Parkham +P 4 (<1) 739 (78) 14 (2) Elliot control 6 (<1) 310 (46) 17 (3) Elliot +P 4 (<1) 1574 (82) 18 (1) Elizabeth Town control 9 (1) 520 (49) 37 (4) Elizabeth Town +P 6 (<1) 939 (65) 34 (2) Soil Humic P Phytate [beta]-GP AMP Chemical shift 3.95-5.45 4.17-4.28 4.59-4.64 4.47-4.52 ranges (ppm): 4.35-4.40 4.71-4.82 Nowra 143 (2) 86 (1) 80 (1) 41 (Q) Casino 143 (12) 26 (2) 23 (2) 15 (1) Collector 31 (19) 14 (8) 8 (5) 3 (2) Coonabarabran 37 (24) 5 (3) 1 (<1) 1 (<1) Wollongbar 274 (31) 55 (6) 10 (1) 7 (<1) Tocal 131 (15) 65 (7) 15 (2) 11 (1) Somersby A 43 (8) 15 (3) 3 (<1) 2 (<1) Somersby B 77 (12) 19 (3) 4 (<1) 0 (0) Kadina n.d. n.d. n.d. n.d. Crystal Brook n.d. n.d. n.d. n.d. Togari control 108 (32) 29 (9) 12 (4) 8 (2) Togari +P 90 (27) 13 (4) 12 (4) 4 (1) Parkham control 135 (38) 10 (3) 7 (2) 7 (2) Parkham +P 130 (14) 14 (1) 10 (1) 10 (1) Elliot control 263 (39) 0 (0) 10 (2) 10 (2) Elliot +P 210 (11) 0 (0) 18 (1) 19 (1) Elizabeth Town control 355 (33) 23 (2) 21 (2) 19 (2) Elizabeth Town +P 323 (22) 24 (2) 17 (1) 13 (1) Soil s-IHP Unknown Diester Chemical shift 3.85-3.91 monoesters 2 to -1 ranges (ppm): a-e Nowra 5 (<1) 11 (<1) 87 (1) Casino 11 (1) 6 (<1) 14 (l) Collector 5 (3) 2 (1) 8 (1) Coonabarabran 2 (1) 0.4 (<1) 5 (1) Wollongbar 14 (2) 6 (<1) 39 (2) Tocal 20 (2) 6 (<1) 14 (1) Somersby A 4 (<1) 0 (0) 8 (1) Somersby B 5 (<1) 0 (0) 8 (1) Kadina n.d. n.d. n.d. Crystal Brook n.d. n.d. n.d. Togari control 12(4) 0 (0) 6 (3) Togari +P 7 (2) 0 (0) 4 (5) Parkham control 8 (2) 9 (3) 16 (4) Parkham +P 9 (1) 2 (<1) 5 (1) Elliot control 5 (<1) 0 (0) 31 (4) Elliot +P 9 (<1) 0 (0) 28 (2) Elizabeth Town control 14 (1) 27 (3) 19 (2) Elizabeth Town +P 13 (1) 22 (2) 28 (2) Soil Pyro-P Chemical shift -4.5 to -5.5 ranges (ppm): Nowra 62 (1) Casino 11 (1) Collector 1 (1) Coonabarabran 2 (1) Wollongbar 14 (2) Tocal 10 (1) Somersby A 5 (1) Somersby B 5 (1) Kadina n.d. Crystal Brook n.d. Togari control 12 (3) Togari +P 17 (5) Parkham control 16 (4) Parkham +P 13 (1) Elliot control 24 (4) Elliot +P 33 (2) Elizabeth Town control 20 (2) Elizabeth Town +P 25 (2)
|Printer friendly Cite/link Email Feedback|
|Author:||Doolette, A.L.; Smernik, R.J.; Dougherty, W.J.|
|Date:||Mar 1, 2011|
|Previous Article:||Assessment of topsoil properties in integrated crop-livestock and continuous cropping systems under zero tillage.|
|Next Article:||Nitrogen, phosphorus, and potassium prediction in soils, using infrared spectroscopy.|