Most important is to obtain different phosphorous fractions ( Water soluble- P , Fe- P , Al- P , Ca- P , occluded P , adsorbed-P etc ) out of a given soil following a sequential extraction method , conventionally followed and such sequential extractions still stand valid and relate well Q/ I relation of soil P.

Dear Dr Mina Jalali . See the following useful RG link: Article Innovative methods in soil phosphorus research: A review

Kindly see also the following ver good RG link: Article Assessment of a phosphorus fractionation method for soils: p...

Also check please the following useful RG link: Article Phosphorus Fractions in the Sediments of Yuecheng Reservoir, China

The following RG link is also very useful: Article Comparison of analytical procedures for measuring phosphorus...

Procedure i used in my doctoral degree research

Forms of Phosphorus

Total Phosphorus

The total phosphorus was extracted by digesting the soil with nitric acid and perchloric acid until a white residue was left. The residue was filtered and made to a known volume. Total phosphorus was then estimated by vanadomolybdophosphoric yellow colour method (Hesse, 1971)

Organic phosphorus

Organic phosphorus was determined by deducting the sum of total inorganic phosphorus from total phosphorus as suggested by Mehta et al. (1954).

Available phosphorus

The available phosphorus was extracted using Bray’s No.1 extractant for the soils having pH less than 6.5 and Olsen’s extractant for the soils having pH 6.5 and above. The extracted phosphorus was estimated by chlorostannous reduced molybdophosphoric blue colour method (Jackson, 1973).

Forms of inorganic phosphorus

The method outlined by Peterson and Corey (1966) was followed to fractionate soil inorganic phosphorus.

Saloid bound phosphorus (Saloid P)

Exactly 0.5 g of soil was taken in to a 50 mL polyethylene centrifuge tube, 25mL of 1 M NH4Cl solution was added and shaken for 30 minutes. Saloid P was estimated by molybdosulphuric acid reagent, using stannous chloride as reductant after taking the extract from supernatant solution after centrifugation in to an aliquot of 10 mL isobutyl alcohol. Blue colour intensity was measured at 660 nm using spectrophotometer.

Aluminium phosphorus (Al-P)

The soil residue left after saloid P estimation was shaken for one hour with 25 mL of 0.5 M NH4F (pH 8.2). The Al-P in the supernatant centrifuged suspension was determined by chloromolybdic – boric acid reagent and chlorostannous reductant. The intensity of blue colour developed was read in spectrophotometer at 660 nm.

Iron phosphorus (Fe-P)

The soil sediment from Al-P estimation was washed twice with 25 mL portion of saturated NaCl solution by shaking and centrifuging. The soil was then treated with 0.1 M NaOH and shaken for 17 hours and centrifuged. The supernatant solution was then treated with five drops of concentrated sulphuric acid. Phosphorus free activated carbon was used to remove suspended organic matter. The Fe-P content in the filtrate was determined by chloromolybdic – boric acid reagent and chlorostannous reductant. The intensity of blue colour developed was measured using spectrophotometer at 660 nm.

Reductant soluble phosphorus (Red-P)

The soil residue from Fe-P estimation was washed twice with 25 mL of saturated NaCl solution by shaking and centrifuging. Soil was then suspended in 15 mL of 0.3 M sodium citrate solution and shaken for 15 minutes with 0.5 g sodium dithionate. The suspension was heated on a water bath at 80ºC for a few minutes. Clear supernatant solution was decanted into a 50 mL volumetric flask after centrifugation. Soil was then washed twice with saturated NaCl and the washings returned to sodium citrate- dithionate extract which was taken for Red-P estimation. Excess of citrate and dithionate were oxidised by 1.5mL of 0.25 M KMnO4 solution. The Red-P was estimated by molybdatesulphuric acid reagent with stannous chloride as reductant after taking the extract into an aliquot of 10mL isobutyl alcohol. The blue colour intensity developed was diluted with equal quantity of absolute ethyl alcohol and read at 660 nm in spectrophotometer.

Occluded phosphorus (Occl-P)

The soil residue left out in the estimation of Red-P was added with 25 mL of 0.1 M NaOH and shaken for one hour. Supernatant solution after centrifugation was taken for estimation of Occl-P by chloromolybdic-boric acid reagent with chlorostannous reductant.

Calcium phosphorus (Ca-P)

The soil residue after extraction of occluded phosphorus was washed twice with 25 mL of saturated NaCl solution and washings were discarded. Ca-P was extracted by using 0.25 M H2SO4 and shaking for one hour and centrifuging for five minutes. The phosphorus in supernatant solution was estimated by chloromolybdic-boric acid reagent with chlorostannous reductant.

Soil phosphorus fractionation as affected by paper mill biosolids applied to soil of contrasting prop. Front. Env. Sci. https. //doing.org/103389/fenvs 2020.00038 A rapid fractionation method for assessing key soil phosphorus parameters in agro-ecosystem. Geoderma 2020.http//doil.org/1016/j.geoderma 2020.114893

Be very cautious about any suggestion that sequential fractionation procedures can identify solid phase species. The fractions extracted should simply be regarded as, e.g., extracted by ..... under the conditions used. Genuine solid phase speciation requires spectroscopic observations

Paul Milham I agree with you. But the fractions of P obtained with different extractants give an idea of the size of pool/amount of P available to plants from different fractions .Modified Hedley fraction is being followed by different researchers.I am attaching one detailed procedure based Hedley's scheme.

I am attaching the file

Fine Annagi, as long as you simply describe the extracted fractions by the procedure used and do not attempt to attribute them to solid phases, Paul.

Paul Milham True.We can not measure different solid phases P and their constituent amounts through fraction schemes.

In this case, it would be interesting to evaluate the article Fractions of phosphorus in soil fertilized with phosphates in different application modes and cultivated with corn, at the following site: https://www.scielo.br/j/rbcs/a/bnJY9fBQKXsS3hwNpZKtcCC/?lang=pt

The forms in which phosphorus (P) accumulates in soils are dependent on management practices, fertilizer sources, and methods of application, which may promote distinct P solubility and plant uptake how soil tillage and phosphate fertilization strategies affected soil P fractions over years and to identify best management practices for improving labile P fractions? Evidence is provided for microbial activity playing a major role in redistributing P into different forms in the soil. Sequential extraction of both fumigated and unfumigated samples to determine the fate of P of microbial origin is an alternative but the fractions of P obtained with different extractants give an idea of the size of pool/amount of P available to plants from different fractions.

I have come across a new method for measuring the P fractions in diverse media.The authors claim that the method is simple , and large number of samples can be handled easily.

Malachite green method for determining phosphorus concentration in diverse matrices .Rahutomo,S.et al. Commun. Soil Sci. Plant Anal. 50(14) 2019

https://doi.org/10.1080/00103624.2019.1635140

Please refer to the following papers of mine where I have used soil P fractionation procedure. It is clearly explained in the methods section.

1. Bouray, M., Moir, J., Condron, L., and Lehto, N. (2021). Lime-induced pH elevation influences phosphorus biochemical processes and dynamics in the rhizosphere of Lupinus polyphyllus and Lupinus angustifolius. Journal of Soil Science and Plant Nutrition, 1-15. https://doi.org/10.1007/s42729-021-00495-z 3.

2. Bouray, M., Moir, J., Condron, L., and Paramashivam, D., (2021). Early effects of surface liming on soil P biochemistry and dynamics in extensive grassland. Nutrient Cycling in Agroecosystems,1-15. https://doi.org/10.1007/s10705-021-10163-4