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Phosphorus Determination after Mehlich3 Extraction and Anion Exchange Resinin an Agricultural Soil of NorthwesternSpainJorge Paz-Ferreiro a , Eva Vidal Vázquez b & Cleide Aparecida deAbreu ca Centro de Investigaciones Agrarias de Mabegondo (CIAM) , LaCoruña , Spainb Facultad de Ciencias , Universidade da Coruña , La Coruña , Spainc Instituto Agronômico de Campinas , Campinas , BrazilPublished online: 30 Jan 2012.

To cite this article: Jorge Paz-Ferreiro , Eva Vidal Vázquez & Cleide Aparecida de Abreu (2012)Phosphorus Determination after Mehlich 3 Extraction and Anion Exchange Resin in an AgriculturalSoil of Northwestern Spain, Communications in Soil Science and Plant Analysis, 43:1-2, 102-111, DOI:10.1080/00103624.2012.634688

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Communications in Soil Science and Plant Analysis, 43:102–111, 2012Copyright © Taylor & Francis Group, LLCISSN: 0010-3624 print / 1532-2416 onlineDOI: 10.1080/00103624.2012.634688

Phosphorus Determination after Mehlich3 Extraction and Anion Exchange Resin in an

Agricultural Soil of Northwestern Spain

JORGE PAZ-FERREIRO,1 EVA VIDAL VÁZQUEZ,2 ANDCLEIDE APARECIDA DE ABREU3

1Centro de Investigaciones Agrarias de Mabegondo (CIAM), La Coruña, Spain2Facultad de Ciencias, Universidade da Coruña, La Coruña, Spain3Instituto Agronômico de Campinas, Campinas, Brazil

Differences in soil phosphorus (P) contents measured by various techniques may haveimplications for agronomic and environmental testing. Reduced-tillage systems com-bined with surface manure application increase the potential risk of nutrient lossesby surface runoff. A field trial was conducted to evaluate the effect of livestock slurryon nutrient accumulation at the surface layer of an acidic soil rich in organic matterwith excess P levels and loamy texture. Two reduced-tillage systems, no tillage (NT)and minimum tillage (MT), and four different fertilizer treatments were assessed. Theamounts of P extracted by anion-exchange resin (AER) and by Mehlich 3 (M3) werecompared; in addition, differences between two determination methods of P con-tents extracted by Mehlich 3, namely molybdic acid colorimetric standard procedure(M3-COL) and inductively coupled plasma–mass spectroscopy (M3-ICP), were eval-uated. Ninety-six soil samples were taken from the 0- to 5-cm surface layer in threesuccessive dates after increasing manure addition. Colorimetric Mehlich 3 P rangedfrom 49 to 431 mg dm–3. The ranks of mean extractable soil P concentrations wereAER < M3-COL < M3-ICP. The linear correlation coefficient between M3-COL andM3-ICP was highly significant (R2 = 0.89; P < 0.01), but a two-straight-lines modelor a quadratic relationship were more adequate for describing the dependence betweenthe two determination procedures after M3 extraction. Relative and absolute differencesbetween M3-COL and M3-ICP showed a tendency to increase as organic carbon con-tent increased. Phosphorus content extracted by AER and M3-COL or M3-ICP reporteda significant but much less predictable relationship with R2 values of 0.27 and 0.21(n = 96), respectively. The P in the surface layer accumulated more under NT thanunder MT.

Keywords Colorimetric analysis, ICP, ion exchange resin, livestock manure,Mehlich 3, phosphorus

Introduction

Phosphorus (P) is an essential element for crop growth, and it is necessary to maintainprofitable agriculture. However, in agricultural areas with high cattle manure and/or slurryinputs, soil P has accumulated to levels in excess of crop needs and has the potential toenrich surface runoff. Thus, P export in watershed runoff can trigger the eutrophication

Address correspondence to Jorge Paz-Ferreiro, Centro de Investigaciones Agrarias deMabegondo (CIAM), Apartado 10, 15080, La Coruña, Spain. E-mail: jpaz@udc.es

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Phosphorus Detection Methods 103

of surface water bodies (Gartley and Sims 1994; Sims et al. 2000, 2002; Kronvang et al.2005). The accelerated nutrient enrichment or eutrophication of surface waters has becomea significant environment problem in many developed countries, and agriculture has beenidentified as a significant P source (Gartley et al. 2002; Kronvang et al. 2005). This is alsothe case in the Atlantic temperate humid region of Galicia, northwestern Spain (SandeFouz 2005).

No-tillage (NT) systems are efficient in reducing soil losses caused by rainfall erosion,when compared with conventional tillage. However, these effects are generally less onmineral and organic nutrient losses. In the case of P, Bertol et al. (2007) found greaterconcentrations in the superficial 0- to 0.025-m soil layer of NT than in conventionallytilled erosion plots. The marked accumulation of P in the superficial soil layer in systemswith reduced tillage was responsible for the greater concentration of this element in runoffand sediment from NT plots when compared with conventional tillage.

Soil testing is a useful diagnostic tool to assess plant P availability. Most soil teststypically involve extracting the soil with a specific solution for a given amount of time. TheMehlich 3 (M3) universal extractant (Mehlich 1984) is suitable for acidic to neutral soils,which include the majority of soil types used for cropping in our study area, northwesternSpain.

Soil tests for M3 soil P use a colorimetric analysis method to measure the P extractedfrom the soil (Mehlich 1984; Jones 1998; Zbiral and Nemec 2000; Caridad Cancela 2002).However, increasing soil-test interpretations for a new version of the M3 P test have beenprovided (Eckert and Watson 1996; Jones 1998; Eralshidi and Mays 2003). This new ver-sion uses inductively coupled plasma (ICP) techniques to measure the P extracted from thesoil. The new method was adopted rapidly because the ICP instrument can be used to mea-sure amounts of other nutrients in the same soil extract. This notwithstanding, the P valuewith ICP is not always comparable with the colorimetric P value that has been previouslyused to set up fertilizer P recommendations. Using the ICP method may result in greater Ptest values (Eckert and Watson 1996; Eliason, Lamb, and Rehm 2001; Nathan et al. 2002;Mallarino 2003; Pittman et al. 2005; Sikora et al. 2005). The reason is that soil extracts forall P tests have the orthophosphate P form and also small amounts of other P forms (suchas other inorganic and simple organic forms). The ICP instrument is thought to measure allP forms in the sample (Mallarino 2003).

Instead of using extractants for soil P analysis, chemical sink–based tests that relyon P sorption–desorption reactions have been proposed. One test is based on ion-exchange resins, either resin beads or plastic membranes (Olsen and Sommers 1982; Olsen,Watanabe, and Bowman 1983). Recent research suggests that anion-exchange resin (AER)may be as effective as commonly used routine P tests based on a chemical extraction atpredicting crop response to P fertilization. Therefore, it would be a useful tool for bothagronomic and environmental soil P testing.

Universal extractants and ion-exchange resins are capable of simultaneously determin-ing a multiple number of elements in a single step. The adoption of cheaper soil-testingmethods can lead to more precise fertilizer application, thereby reducing environmentalimpact of excess levels. However, differences in P contents measured by various techniquesmay have implications for agronomic and environmental testing.

The objectives of this research were (1) to compare P concentrations as extracted byM3, determined both by colorimetry and ICP, and obtained by ion-resin exchange and(2) to differentiate between the effects of two reduced-tillage systems on P accumulationat the soil surface layer.

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104 J. Paz-Ferreiro, E. V. Vázquez, and C. A. de Abreu

Material and Methods

The experimental site was located near Pastoriza, Lugo Province, Galicia, northwesternSpain. A field trial took place from 2004 to 2006 for testing four fertilizer managementconditions (mineral fertilizer as a control and livestock manure applied at rates of 30, 60,and 90 Mg ha–1), which were factorially combined with two reduced-tillage systems: min-imum tillage (MT), consisting of chisel plowing at 60 cm deep followed by direct drilling,and no tillage (NT). The experimental design was a split plot with four replications,given 32 individual plots. Before to our trial, the experimental field was conventionallytilled.

The crop rotation was maize and winter cereal. Mineral fertilizer and livestock manurewere applied on the soil surface in March or April (spring) or in October or November(winter). Samples were collected at three different dates. Table 1 lists the cumulative min-eral or organic liquid manure on the four different treatments before each of the threesampling dates.

Soil was sampled at 0–5 cm deep, because this depth is more relevant to the potentialfor erosive movement of nutrients dissolved or suspended in runoff. Samples were air dried,crushed, and sieved with a 2-mm sieve. Soil general properties were determined by routineanalysis as follows: texture by the pipette method (Guitian and Carballas 1976) and pH(H2O) and organic-matter content (wet oxidation) according to procedures described indetail in van Raij et al. (2001). On average, sand, silt, and clay contents were 29.1%,46.7%, and 24.2%, respectively; gravel content was 39.8%. Table 2 summarizes results fororganic-matter content and pH during the three successive sampling dates. Mean organic-matter content showed a trend to increase from the first to the second sampling date, withvalues of 64.2 and 69.7 g kg–1 respectively, but these differences were not significantlydifferent (P < 0.01). Minimum organic-matter contents steadily increased with values of42.56 and 59 g kg–1 during the first, second, and third sampling dates, respectively. Valuesof pH were rather stable along the successive sampling dates with mean values of 5.7, 5.8,and 5.7.

Soil P was extracted with the M3 test (Mehlich 1984), as briefly described: 5 cm3

of soil and 50 mL of the extractant solution [0.2 mol L–1 acetic acid (CH3COOH),0.25 mol L–1 ammonium nitrate (NH4NO3), 0.015 mol L–1 ammonium fluoride (NH4F),0.013 mol L–1 nitric acid (HNO3), and 0.001 mol L–1 ethylenediaminetetraacetic acid(EDTA), adjusted at pH 2.5] were taken. The suspensions were placed in polyethyleneflasks covered with plastic stoppers and shaken by horizontal and circular movements at

Table 1Cumulative amounts of mineral or organic fertilizers applied during

three successive sampling dates

Sampling date

Treatment 1[mineral(P2O5),kg ha−1]

Treatment 2(livestockmanure,

30 Mg ha−1)

Treatment 3(livestockmanure,

60 Mg ha−1)

Treatment 4(livestockmanure,

90 Mg ha−1)

1 300 120 180 2702 400 150 240 3603 500 180 300 450

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Phosphorus Detection Methods 105

Table 2Summary statistics for organic-matter content and pH during three

successive sampling dates

Organic-matter content (g dm−3) pH

Sampling date Mean Maximum Minimum Mean Maximum Minimum

1 64.2 80 42 5.7 6.1 5.22 69.7 102 56 5.8 6.2 5.33 68.8 85 59 5.7 6.3 5.1

240 oscillations per minute for 5 min. After filtration, the P content was determined bothby colorimetry and ICP.

Phosphorus analysis by ion-exchange resin was performed as outlined by van Raijet al. (2001): 2.5 cm3 of soil and 25 mL water were taken and the soil suspension wasshaken for 15 mins. Then 2.5 cm3 resin were added, and the suspension of soil plus resinwas shaken for 16 h. The suspension is sieved with a 0.4-mm sieve. Resin was transferredin polyethylene flasks using 50 mL ammonium chloride (NH4Cl) 0.8 mol L–1 solution inhydrochloric acid (HCl) 0.2 mol L–1.

Soil P desorbed by the resin and extracted by M3 was determined by the ascorbic acid–ammonium molybdate colorimetric method based on the Murphy and Riley (1962) methodfollowing procedures described in van Raij et al. (2001). For the ICP P determination,M3 extracts were analyzed with a Jobin Yvon JY 50-P ICP emission spectrophotometer(Jobin Yvon Emission Instruments, Longjumeau, France) (van Raij et al. 2001).

Results and Discussion

Comparison of Phosphorus Amounts Extracted by Soil Tests

Averaged over all three sampling data, the amounts of P measured by AER, M3-COL, andM3-ICP were 182.0, 267.5, and 296.8 mg dm–3, respectively. The range of measured Pvalues was from 62 to 364 mg dm–3 for resin, from 49 to 431 mg dm–3 for M3 colorimetricmethod, and from 45.6 to 698.8 mg dm–3 for M3-ICP.

The relationship between ICP and colorimetric P determination in M3 extracts of the96 soil samples in this study is shown in Figure 1. During the first sampling date, whenM3 extractable P was relatively low, P concentrations determined by ICP were generallyless than colorimetric values, except for one out of 32 samples. However, during the sec-ond and the third sampling dates, P concentrations determined by ICP were greater thancolorimetric values, except for one out of 64 samples.

A significant correlation was found between the two methods of M3 determination(R2 = 0.898) by linear regression. However, the intercepts of the line shown in Figure 1equals 90.3 mg dm–3, meaning that when the extractable P in the soil is low a single linearmodel would predict even greater colorimetric values than those experimentally measured.Therefore, models different from the lineal model have been tested.

A quadratic relationship exhibits a greater determination coefficient (R2 = 0.939). Thefitted equation was ICP P = 0.0008 (COL P) 2 + 1.131 (COL P) + 17.086. Also, a two-straight-line model with a break at 300 mg dm–3 was superior to a single lineal model

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106 J. Paz-Ferreiro, E. V. Vázquez, and C. A. de Abreu

y = 0.596x + 90.3R2

= 0.898

0

100

200

300

400

500

0 100 200 300 400 500 600 700

ICP-P (mg.dm−3)

Co

lori

met

ric-

P (

mg

.dm

−3)

Figure 1. Relationship between P contents measured by extraction with Mehlich 3 and two differentdetermination methods, based on inductively coupled plasma emission spectroscopy (ICP-P) andcolorimery (colorimetric P, COL-P) (color figure available online).

(Figure 1). We are not aware of any published study that has specifically evaluated thisbreak in the linear relationship between ICP P and COL P extracted by M3 solution.

In our study, colorimetric P was on average 90% of the P determined by ICP, butas previously stated, values in the lower range for COL P were greater than their corre-sponding ICP P values. Mallarino (2003) and Pittman et al. (2005) found M3 results to begreater when P was measured with ICP as compared to a colorimetric approach for lowranges of extractable P, where P fertilizer would be recommended, that is, values generallyless than 60 mg kg–1. Sikora et al. (2005) found the slopes of correlations between ICPand colorimetric P determination in M3 extracts were much closer to 1 (1.02 and 0.968 fortwo data sets) as compared to the results of Pittman et al. (2005) and Mallarino (2003).However, when M3-extractable P was low and P fertilizer would be recommended, P con-centrations determined by ICP were greater than colorimetric values more frequently thanthe reverse. Jacoby (2005) evaluated 23 fields where manure had been applied either via amanure spreader, grazing cattle, or both on feed yards and dairies. The range in colorimetricP was from 31 to 472 mg kg–1 and ICP P ranged from 40 to 772 mg kg–1.

Determination coefficient between M3 P by ICP P and the colorimetric method wasR2 = 0.76 with a 47 mg kg–1 P intercept. The colorimetric P was 55% of the P determinedby ICP.

The relative difference (ratio) ICP P / COL P was between 0.62 and 1.6. The absolutedifferences, ICP P minus COL P, were between –37.8 and 60 mg dm–3. Figure 2 shows therelation between the ratio ICP P / COL P and the absolute differences ICP P minus COL Pversus colorimetric P. A positive and significant relationship (P < 0.01) was found in bothcases. In the range of M3-COL P < 80 mg kg–1, Mallarino (2003) found a exponentialnegative relationship between the ratio ICP P / COL P and colorimetric P; at low P con-tents, relative differences between both P determination methods could be as high as 3.5.Therefore, soil P content plays a role in the relationship between the two determinationmethods but does not explain differences between them.

Weak positive relationships were found between the ratio ICP P / COL P versusorganic carbon (R2 = 0.165) and also between the absolute difference ICP P minus COL

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Phosphorus Detection Methods 107

y = 0.7181e0.0015x

R2 = 0.583

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

0 100 200 300 400 500

M3-COL P (mg dm−3)

M3-

ICP

/M3-

CO

L P

rat

io

y = 0.1591x − 33.991R2

= 0.614

−60

−40

−20

0

20

40

60

80

0 100 200 300 400 500

M3-COL P (mg dm−3)

(M3-

ICP

) - (M

3-C

OL

) P

(mg

dm

−3)

Figure 2. Relationship between soil P measured by extraction with Mehlich 3 and withdetermination based on colorimetry and the relative (M3-COL/M3-ICP) or absolute (M3-COLminus M3-ICP) differences from soil P determined by ICP and colorimetry.

P and organic carbon (R2 = 0.141). This result suggests that organic carbon changes alongtime are responsible for the differences between ICP and colorimetric procedure.

A possible explanation for the greater values obtained by colorimetric over ICP inthe first sampling date includes the presence of organic by-products. Such substances mayabsorb light at the specific wavelength corresponding to the colored molybdic complex,thereby increasing the colorimetric P reading. On the other hand, several authors indicatethat the commonly held belief is that ICP would measure greater P concentrations thancolorimetric procedures because the high-temperature environment of the plasma wouldallow the measurement of organic P compounds or other soluble P complexes that wouldnot be measured colorimetrically (Mallarino 2003; Pittman et al. 2005).

It is likely that organic P species plays a role in the discrepancy between ICPand colorimetric P analyses when ICP results are greater; other as yet unidentified fac-tors may also play roles. However, currently there is little insight into what may causecolorimetric P analyses to be greater than ICP results for water extracts of organicby-product materials.

Phosphorus content extracted by AER when compared to M3-COL or M3-ICPreported a significant but much less predictable relationship with R2 values of 0.27 and

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108 J. Paz-Ferreiro, E. V. Vázquez, and C. A. de Abreu

0

100

200

300

400

0 100 200 300 400 500 600 700 800

ICP-P (mg dm–3)

Res

in-P

(m

g d

m–3

)

1st sampling 2nd sampling 3rd sampling

Figure 3. Relationship between resin P and P extracted by Mehlich 3 with determination by ICP(color figure available online).

0.21 (n = 96), respectively. The regression equations were as follows: AER P = 0.362 M3-COL+ 84.97 and AER P = 0.204 M3-ICP + 121.4.

Figure 3 shows the relationship between AER P versus P extracted by M3-ICP forthe three successive sampling dates in this study. The relationship between both variablesis stronger when considering each of the sampling date. The respective regression equa-tions and coefficients of determination are as follows: AER P = 0.791 M3-ICP + 17.08(R2 = 0.626), AER P = 0.320 M3-ICP + 110.9 (R2 = 0.265), and AER P = 0.419 M3-ICP + 0.064 for the first, second, and third sampling dates, respectively. From the slopeand intercept of the regression lines, it is apparent that the ratio AER P / ICP P decreasesas the fertilizer addition and thus the soil surface layer P content increases.

Effect of Tillage and Management Systems

Table 3 lists mean P concentrations as extracted by resin and determined by thecolorimetric method (AER P) and as extracted by M3 solution and determined by ICPand colorimetry. The P levels were in excess of levels required for maximum crop yield.Manure applications to these soils should be restricted because of risk of loss of P to surfacewaters.

Mineral fertilizer in the control treatment and liquid manure applications in the threeremaining treatments significantly increased soil-test P levels, and these effects weredetectable with all the analytical methods. However, the differences in the amounts of Pbetween the second and the third sampling dates were not detected by the resin extractionmethod. The widest differences between mean P contents during the successive samplingdates were those extracted with M3 and determined by ICP.

Differences between liquid organic fertilizer treatments were not significantly differ-ent along the successive sampling dates. Because of considerable spatial variability in theexperimental field, there was considerable scatter between individual plots, which maskedclear evidence of P differences between fertilizer treatments.

Phosphorus contents extracted by M3 and determined both by ICP and colorimetryfrom the 0- to 5-cm surface layer were significantly greater (P < 0.01) in the no-tillagethan in the minimum-tillage treatments for all the different mineral or organic manuredoses and during all three study sampling dates. In general, AER P concentrations werealso greater in NT than in MT treatments, but there were some exceptions. Thus, no tillage

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Phosphorus Detection Methods 109

Table 3Mean soil P contents resulting from three different methods for two management systems

and four fertilizer treatments (mg dm–3)

Minimum tillage No tillage

Treatment Resin M3-COL M3-ICP Resin M3-COL M3-ICP

First date1 153.3 203.5 165.4 158.8 213.8 178.62 125.0 172.5 160.2 192.0 240.8 222.13 147.0 200.5 170.7 200.5 240.3 209.84 158.0 203.3 186.7 216.8 253.3 240.6Mean 145.8 195.0 170.7 192.0 237.1 212.8Second date1 239.0 219.8 243.0 204.5 260.0 286.52 153.0 238.8 254.2 211.5 284.3 332.53 194.5 261.5 280.3 203.8 256.8 268.54 201.8 244.0 258.1 178.5 243.0 260.6Mean 197.1 241.0 258.9 199.6 261.0 287.0Third date1 149.5 318.3 384.5 172.8 334.5 415.42 134.5 318.3 350.7 194.8 326.7 474.73 182.0 285.5 434.6 226.8 334.5 464.64 170.8 328.0 406.5 198.8 368.5 474.7Mean 159.2 312.5 394.1 198.3 341.1 457.3

was most efficient in accumulation of mineral fertilizer and manure at the uppermost soilsurface layer, whereas minimum tillage facilitated incorporation into deeper soil layersthan no tillage.

Conclusions

This study confirms that M3 soil tests are an effective means for evaluating the P status ofagricultural soils with excess levels and high inputs of organic or mineral fertilizers.

Mehlich 3 P determined by ICP was greater than colorimetric P at high values ofextractable P, whereas the reverse was true at low values of extractable P. Both determina-tion methods were capable of evaluating excessive soil surface layer P due to mineral andliquid organic fertilization.

The absolute and the relative differences between the M3-ICP and M3-COL tests werecorrelated with the soil P level in the M3-COL P range from 49 to 431 mg dm–3.

Phosphorus contents extracted by M3 and determined both by ICP and colorimetryfrom the 0- to 5-cm surface layer were significantly greater (P < 0.01) in the no-tillagethan in the minimum-tillage treatments.

Acknowledgments

This study was supported by Spanish Education and Science Ministry (MEC) under ProjectAGL2003-09284-C02-01.

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