Interlaboratory validation of the Mehlich 3 method as a universal extractant for plant nutrients.

The Mehlich 3 (M3) method is widely used to extract plant-available phosphorus from soil over a wide range of pH values. The method is also used by many laboratories to determine multiple plant-available nutrients simultaneously. However, except for P, this method has not been statistically validated within and among laboratories. The objective of this study was to determine the repeatability (within-laboratory performance) and reproducibility (among-laboratories performance) of the M3 method for several different nutrients by using a wide variety of soils. An in-house homogeneity test was conducted for 11 soils. Three replicates of each of the 11 soils were sent to 23 domestic and international laboratories for analyses primarily for K, Ca, Mg, Zn, Mn, Fe, and Cu. Samples were scooped, weighed, or both scooped and weighed for extraction. The various nutrients in the extracts were quantified by the participating laboratories by using inductively coupled plasma-atomic emission spectrometry. Ranges (in parentheses) of the repeatability relative standard deviation (RSDr) values for the scooped samples were K (3.88-6.14%), Ca (2.19-10.6%), Mg (2.27-5.73%), Zn (4.11-42.7%), Mn (3.15-8.53%), Fe (2.32-7.74%), and Cu (3.65-11.2%). For the weighed samples, the ranges (in parentheses) of the RSDr values were K (1.65-4.65%), Ca (1.43-16.9%), Mg (1.37-9.83%), Zn (2.60-33.0%), Mn (1.61-4.90%), Fe (1.56-4.47%), and Cu (2.95-17.9%). Ranges (in parentheses) of the reproducibility relative standard deviation (RSDR) values for the scooped samples were K (7.44-20.0%), Ca (7.10-33.5%), Mg (7.63-26.4%), Zn (11.6-48.7%), Mn (10.5-19.7%), Fe (12.4-22.0%), and Cu (8.86-45.3%). Ranges (in parentheses) of the RSDR values for the weighed samples were K (3.47-12.7%), Ca (7.61-34.6%), Mg (7.53-29.0%), Zn (10.6-49.2%), Mn (6.54-20.7%), Fe (11.0-21.5%), and Cu (9.73-43.0%). The Horwitz ratios (HorRat) were also used to evaluate the repeatability, HorRatr, and the reproducibility, HorRatR. Overall, the M3 method appears to be both repeatable and reproducible across the two categories for K, Mg, Zn, Mn, and Cu, and the vast majority of the HorRat values for both repeatability and reproducibility were within the acceptable range. However, a large number of soils in both the weighed and scooped categories displayed unacceptable HorRat values for the reproducibility of Ca and Fe, indicating that the M3 is not suitable for the extraction and quantitation of Ca and Fe in soils. The results of this study indicate that the M3 method for the determination of K, Mg, Zn, Mn, and Cu in soils is both accurate and precise when standardized procedures are used. The method has been proven to be suitable for use as a reference method for testing soil materials for extractable P, K, Mg, Zn, Mn, and Cu. Further study may be needed to confirm the suitability of the M3 method for Ca and Fe.

Interlaboratory validation of the Mehlich 3 method for extraction of plant-available phosphorus.

The Mehlich 3 (M3) method is widely used for extraction of plant-available phosphorus (P) from soil over a wide range of pH values. The method is also used by many laboratories to determine multiple plant-available nutrients simultaneously. However, this method has not been statistically validated within and among laboratories. The objective of this study was to determine the repeatability (within-laboratory performance) and reproducibility (among-laboratories performance) of the M3 method by using a wide variety of soils. An in-house homogeneity test was conducted for 10 soils. Three replicates of each of the 10 soils were sent to 26 domestic and international laboratories primarily for P analysis. Samples were scooped, weighed, or both scooped and weighed for extraction. The P in extracts was quantified by the participating laboratories by using inductively coupled plasma-atomic emission spectrometry (ICP-AES) or colorimetrically. For the scooped samples analyzed colorimetrically, the repeatability relative standard deviation (RSDr) ranged from 2.07 to 12.1%; the RSDr ranged from 2.2 to 21.4% for the scooped samples analyzed by ICP-AES. For the weighed samples analyzed colormetrically, the RSDr values were 1.09-9.34%, and for the weighed samples analyzed by ICP-AES, they were 1.70-5.76%. For the reproducibility data, the RSDR values ranged from 6.85 to 50.8% for the scooped-colorimetry category, from 6.95 to 73.9% for the scooped-ICP-AES category, from 7.19 to 42.6% for the weighed-colorimetry category, and from 5.29 to 35.9% for the weighed-ICP-AES category. The greatest RSD values were associated with the Susitna soil, which had the smallest concentration of extractable P. Because of the relatively small concentration of P in this soil, the laboratories were attempting to measure solution concentrations that were close to the detection limits. The Horwitz ratios (HorRat) were also used to evaluate the repeatability, HorRat(r), and reproducibility, HorRat(R). Overall, the M3 P method appears to be both repeatable and reproducible across the 4 categories, and the vast majority of the HorRat values for both repeatability and reproducibility were within the acceptable range. The results of this study indicate that the M3 P method for the determination of plant-available P in soil is both accurate and precise when standardized procedures are used. The method has been shown to be suitable for use as a reference method for testing soil materials for extractable P.

Evaluating the contribution of soil properties to modifying lead phytoavailability and phytotoxicity.

Soil properties affect Pb bioavailability to human and ecological receptors and should be considered during ecological risk assessment of contaminated soil. We used path analysis (PA) to determine the relative contribution of soil properties (pH, organic C [OC], amorphous Fe and Al oxides [FEAL], and cation-exchange capacity [CEC]) in modifying Pb bioavailability. The response of biological endpoints (bioaccumulation and dry matter growth [DMG]) of lettuce (Lactuca sativa) grown on 21 Pb-spiked (2,000 mg/kg) soils were determined. Lettuce tissue Pb ranged from 3.22 to 233 mg/kg, and relative DMG ranged from 2.5 to 88.5% of their respective controls. Simple correlation showed strong relationships between CEC and OC (p < 0.01) and weaker relationships between pH and FEAL (p < 0.05) and Pb bioaccumulation. Results of PA suggest that soil pH increased the negative surface charge of organic matter and clay, thereby increasing CEC and decreasing Pb bioaccumulation. Also, the direct effect of OC on tissue Pb can be attributed to formation of surface Pb complexes by organic matter functional group ligands. Increased OC and/or CEC reduced Pb solubility and bioavailability in the 21 soils in the present study. The relative importance of soil properties likely will vary between studies employing different soils. Soil properties should be considered during the ecological risk assessment of metal in contaminated soils. Path analysis is useful for ecological studies involving soils with a wide range of physicochemical properties and can assist in site risk assessment of metals and remediation decisions on contaminated sites.

Effect of soil properties on lead bioavailability and toxicity to earthworms.

Soil properties are important factors modifying metal bioavailability to ecological receptors. Twenty-one soils with a wide range of soil properties (USA; http://soils.usda.gov/technical/classification/taxonomy/) were amended with a single concentration of Pb (2,000 mg/kg) to determine the effects of soil properties on Pb bioavailability and toxicity to earthworms. Earthworm mortality ranged from 0 to 100% acute mortality following exposure to the same total concentration of Pb (2,000 mg/kg) in amended field soils. Internal Pb concentrations in earthworms ranged from 28.7 to 782 mg/kg, with a mean of 271 mg/kg. Path analysis was used to partition correlations in an attempt to discern the relative contribution of each soil property. Results of path analysis indicated that pH was the most important soil property affecting earthworm mortality (p < 0.01) and internal Pb (p < 0.05). Soil pH was related inversely to mortality and internal Pb, soil solution Pb, and Pb bioavailability. The most important soil property modifying reproduction was amorphous iron and aluminum oxides (FEAL). Because FEAL is rich in pH-dependent cation-exchange sites, several soil properties, including pH, FEAL, and cation-exchange capacity, have a causal effect on Pb adsorption and soluble Pb. Path analysis is useful for assessing contaminated soils with a wide range of soil properties and can assist in ecological risk assessment and remediation decisions for contaminated sites. Soil properties are important factors modifying metal bioavailability and toxicity and should be considered during the ecological risk assessment of metals in contaminated soils.

Assessing availability, phytotoxicity and bioaccumulation of lead to ryegrass and millet based on 0.1 mol/L ca(NO3)2 extraction.

This study was conducted to assess availability, phytotoxicity and bioaccumulation of lead (Pb) to ryegrass (Lolium perenne L.) and millet (Echinochloa crusgalli) based on the 0.1 mol/L Ca(NO3), extraction. Effect of soil properties on availability, phytotoxicity and bioaccumulation of Pb to the two plants was also evaluated. Five soils with pH values varying from 3.8 to 7.3, organic carbon (OC) contents from 0.7% to 2.4%, and clay contents from 11.6% to 35.6% were selected. Soils were spiked with Pb to achieve a range of concentrations: 250, 500, 1000, 3000 and 5000 mg/kg. Pb availability in the spiked soils was estimated by extracting soil with 0.1 mol/L Ca(NO3)2. The results indicate that plants yield decreased with decreasing soil pH and increased with increasing soil clay and OC content. Negative relationship between available Pb and the relative dry matter growth (RDMG) of the two plants were significantly related. Available Pb used to assess EC20 (20% effective concentration) and EC50 (50% effective concentration) of millet was 119 and 300 mg/kg, respectively. Available Pb used to assess EC20 and ECs, of ryegrass was 63 and 157 mg/kg, respectively. Bioaccumulation, expressed as bioconcentration factors of Pb, was inversely related to soil pH, soil OC and clay content. Strong relationships were found between available lead and uptake by the two plants (i was 0.92 and 0.95 respectively). In general, 0.1 mol/L Ca(NO3)2 available Pb may be used to assess the availability, phytotoxicity and bioaccumulation of lead to the two plants tested.

Soil characteristics and phosphorus level effect on phosphorus loss in runoff.

The loss of phosphorus (P) in runoff from agricultural soils may accelerate eutrophication in lakes and streams as well as degrade surface water quality. Limited soil specific data exist on the relationship between runoff P and soil P. This study investigated the relationship between runoff dissolved reactive phosphorus (DRP) and soil P for three Oklahoma benchmark soils: Richfield (fine, smectitic, mesic Aridic Argiustoll), Dennis (fine, mixed, active, thermic Aquic Argiudoll), and Kirkland (fine, mixed, superactive, thermic Udertic Paleustoll) series. These soils were selected to represent the most important agricultural soils in Oklahoma across three major land resource areas. Surface soil (0-15 cm) was collected from three designated locations, treated with diammonium phosphate (18-46-0) to establish a wide range of water-soluble phosphorus (WSP) (3.15-230 mg kg(-1)) and Mehlich-3 phosphorus (M3P) (27.8-925 mg kg(-1)). Amended soils were allowed to reach a steady state 210 d before simulated rainfall (75 mm h(-1)). Runoff was collected for 30 min from bare soil boxes (1.0 x 0.42 m and 5% slope) and analyzed for DRP and total P. Soil samples collected immediately before rainfall simulation were analyzed for the following: M3P, WSP, ammonium oxalate P saturation index (PSI(ox)), water-soluble phosphorus saturation index (PSI(WSP)), and phosphorus saturation index calculated from M3P and phosphorus sorption maxima (P(sat)). The DRP in runoff was highly related (p < 0.001) to M3P for individual soil series (r2 > 0.92). Highly significant relationships (p < 0.001) were found between runoff DRP and soil WSP for the individual soil series (r2 > 0.88). Highly significant relationships (p < 0.001) existed between DRP and different P saturation indexes. Significant differences (p < 0.05) among the slopes of the regressions for the DRP-M3P, DRP-WSP, DRP-PSI(ox), DRP-PSI(WSP), and DRP-P(sat) relationships indicate that the relationships are soil specific and phosphorus management decisions should consider soil characteristics.

Ecotoxicological risks associated with land treatment of petrochemical wastes. I. Residual soil contamination and bioaccumulation by cotton rats (Sigmodon hispidus).

Petrochemical waste contains both organic and inorganic contaminants that can pollute soil and may pose significant ecological risks to wildlife. Petrochemical waste typically is disposed of in land treatment units, which are widespread throughout Oklahoma and the United States. Few studies have been conducted evaluating possible toxicity risks to terrestrial organisms residing on these units. In this study, the extent of soil contamination with fluoride (F), metals, and organic hydrocarbons, the bioaccumulation of F and metals in cotton rats (Sigmodon hispidus), the relationship between contaminants in soil and in tissues of cotton rats, and the level of potentially toxic polycyclic aromatic hydrocarbons (PAHs) in soil were determined on land treatment units. Over a 2-yr period, cotton rats and soils were collected and analyzed from 5 land treatment and matched reference units. The number of land treatment units with soil metal contamination (in parentheses) included: Cr, Cu, Pb (5). Al, As, Ni, Sr, Zn (4). Ba (3). and Cd, V (2). The number of land treatment units with soil PAH contamination (in parentheses) were naphthalene, phenanthrene, benzo[g,h,i]perylene (3). acenaphthene, anthracene, pyrene, benz[a]anthracene, chrysene, benzo[b]fluoranthene, benzo[a]pyrene, indeno[1,2,3-c,d]pyrene, dibenz[a,h]anthracene (2). and acenaphthylene, fluorene, fluoranthene, benzo[k]fluoranthene (1). Total PAH and total petroleum hydrocarbons (TPH) were elevated at all five land treatment units. Mean sums of benzo[a]pyrene (BaP) equivalents (BaPequiv ) were not affected on

Ecotoxicological risks associated with land treatment of petrochemical wastes. II. Effects on hepatic phase I and phase II detoxification enzymes in cotton rats.

The purpose of this study was to evaluate possible exposure and resultant hepatic effects of petrochemical waste on cotton rats (Sigmodon hispidus) living on landfarmed sites. Male and female cotton rats were collected in summer, fall, and winter from four landfarm sites and four ecologically similar reference sites. Hepatic methoxyresorufin O-deethylase (MROD) activity was significantly induced in male and female rats collected from landfarms compared to rats collected from reference sites. In contrast, changes in ethoxyresorufin O-deethylase (EROD) activity were inconsistent due to season, sex, and treatment variation. A significant decrease in EROD and MROD activity was found in cotton rats held for 48 h prior to sacrifice compared to rats euthanized on the day of capture. These results indicate that when using hepatic EROD and MROD activities as biochemical markers of exposure to aryl hydrocarbon receptor agonists, animals should be euthanized as quickly as possible after capture. The cotton rats collected from one landfarm unit exhibited a pattern of consistent elevation of EROD, MROD, and pent-oxyresorufin O-deethylase (PROD) activity. This unit also had a pattern of elevated CYP1A2 protein expression determined by Western blotting. There were no consistent alterations from contaminant exposure on hepatic glutathione S-transferase (GST) activity, glutathione levels, or CYP1A1 protein. Hepatic EROD and MROD activities varied considerably between seasons and sex of rats. In conclusion, consistent induction of hepatic EROD and MROD activities in cotton rats was found in three out of four sampled landfarm sites compared to the rats collected from the reference sites, indicating exposure to contaminants-likely polyaromatic hydrocarbons.

Ecotoxicological risks associated with land treatment of petrochemical wastes. III. Immune function and hematology of cotton rats.

Landfarming is a widely used method of treating petrochemical waste through microbial bio-degradation. The effects of residual petrochemical contamination on wildlife, especially terrestrial mammals, are poorly understood. The effects of contaminants on the immune system and hematology of cotton rats (Sigmodon hispidus) living on five abandoned petrochemical landfarms (units 1-5) in Oklahoma were studied. Cotton rats were sampled seasonally (summer and winter) from each landfarm and from five ecologically matched reference sites for 2 yr (1998-2000) and returned to the laboratory for immunological and hematological assays. Overall analysis indicated that rats inhabiting landfarms exhibited decreased relative spleen size compared to rats collected from reference sites, with the landfarm at unit 1 showing the greatest reduction. Cotton rats collected from landfarms also had increased hemoglobin, hematocrit, and platelet levels and decreased blood leukocytes during summer. During winter, an increase in the number of popliteal node white blood cells was observed from rats collected on landfarms. No marked difference was detected for lymphocyte proliferation in response to concanavalin A, pokeweed, or interleukin-2. Lymphokine-activated killer cell lytic ability showed a seasonal pattern, but no treatment differences. No differences between landfarm and reference sites were detected in the hypersensitivity reaction of rats given an intradermal injection of phytohemagluttinin (PHA-P). Comparisons within individual sites indicated that two sites (units 1 and 3) had the greatest effects on immune function and hematology of cotton rats. The results of this study suggest that residual petrochemical waste affects the immune system and hematology of cotton rats living on abandoned landfarms during summer and is complicated by variation in the contaminants found on individual petroleum sites.