Procedure

(i) Clear the soil top area where the auger will be punched. Place the PVC sleeve over the site to be sampled and drive it into the ground approximately 5 cm using the wood and the rubber mallet.

(ii) Place a clean auger in the sleeve. Turn the handle to advance the auger until it is filled with soil. Pull the auger up and tap all the soil into a 40 x 50 cm plastic bag. Return the auger to the sleeve and continue turning until the top of the auger is levelled with the soil surface. Again pull the auger up and load all the soil into the same bag. Surface soils tend to be compact, hence, fill the auger two or more times to go from a depth of 0 to 10 cm.

(iii) If deep soil samples are needed, drive the auger as indicated in step 2 but discard the soil extracted until the desired depth is reached. Collect the samples in bags as indicated in step 2 at the desired depth.

(iv) Put on the gloves and hold the top of the plastic bag tightly closed while shaking the soil to mix it thoroughly.

(v) Clean the auger by scrubbing it with a brush in a 5-gallon bucket filled with soapy water. Then, rinse in a 5-gallon bucket filled with clear water. Then, rinse again spraying with double distilled water. Give a final rinse with isopropyl alcohol dispensed from a clean bottle.

(vi) Transport the samples to the laboratory on dry ice and store at -20°C.

(vii) Prevent contamination from the 0 to 10 cm soil, cleaning the PVC sleeve with a gloved hand and scooping it out of the bottom of the hole before taking the second sample.

(viii) Label and transport the samples to the laboratory on dry ice.

3.1 Examples of Soil Collection

Collection of soil rhizosphere from a plant producing allelochemicals (Fujii et al., 2005): Grow the plants from a selected species in plastic pots in controlled conditions for a suitable time (i.e. Medicago sativa in a greenhouse at 25°C for four weeks). Take out the plants from the plastic pot without disturbance. Then, gently shake the plant roots and collect the root-zone soil in plastic bags (Fig. 5A). Then, remove and collect the soil adhering to the surroundings of roots (rhizosphere soil) in plastic bags. Sieve soil samples through 1 mm mesh. Discard root residues of rhizosphere soils. Place in plastic bags and freeze (-20°C) before extraction of soil allelochemicals. These samples should be processed during the following three months after collection.

Collection of soil samples in a walnut-corn cropping system (Jose and Gillespie, 1998): Take the soil cores (0-10 cm depth) at four different distances from the tree rows: 0, 0.9, 2.5 and 4 m from the row (Fig. 5B). Collect and combine 10 soil cores at each plot and collection distance. Place each soil sample in a plastic bag and immediately transport to the laboratory. Refrigerate for no longer than 24 hours before allelochemical extraction. Determine gravimetric soil water for each sample using subsamples. Collect on each sampling date, a composite sample of 10 soil cores from a nearby cornfield, which serves as a control for recovery experiments. Take samples as mentioned at different times during the year to evaluate fluctuations of soil allelochemical content.

A Black walnut ®

A Black walnut ®

Soil sampling distances

Figure 5 (A) Separation between root-zone soil and rhizosphere soil from a plant producing allelochemicals (Source: Fujii et al., 2005). (B) Walnut-corn cropping system (Adapted from Jose and Gillespie, 1998).

Soil sampling distances

Figure 5 (A) Separation between root-zone soil and rhizosphere soil from a plant producing allelochemicals (Source: Fujii et al., 2005). (B) Walnut-corn cropping system (Adapted from Jose and Gillespie, 1998).

Collection of soil samples from crop plots subjected to conventional and no-till cropping systems (Blum et al., 1992): Assuming plots of 30 x 8 m with eight rows subjected to conventional and no-till systems, divide each plot into four sections (15 x 4 m) for sampling. Take two soil cores (5.5 cm in diameter; 0-10 cm depth) per section and combine them. Also take additional soil cores (0 to 2.5 cm depth) adjacent to the previous sampling locations. The 0 to 2.5 cm core samples are taken because allelochemicals released from plant residues are often more concentrated in top soil, making it possible to detect vertical gradients in the soil. Take samples, as mentioned, at several times after crop harvest to evaluate changes in soil allelochemical content.

3.2 Handling Soil Samples

Air-dry composite soil samples by spreading out on paper or any other suitable material in the laboratory. After five or six weeks, store the dry samples in labelled paper bags at room temperature. Several compounds are irreversibly adsorbed to soil particles during the drying process. Hence, these substances will not be available when soil is re-hydrated for chemical analysis. This problem is overcome by freezing the fresh soil samples at -20 °C upto the time of chemical analysis.

3.3 Extraction of Soil Allelochemicals

Experiment 5: Extraction of Phenolic Acids

Soil is extracted with water or ethylenediamine tetraacetic acid (EDTA). Water extraction recovers primarily phenolic acids in soil solution, while the difference between EDTA and water extraction provides an estimate of the reversibly bound phenolic acids weakly sorbed to soil particles (Blum et al., 1992).

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