Asian Soil Laboratory Network: 3rd meeting. Manila, Philippines, 23 - 27 September 2019. Ms. Gina Nilo, Philippines

1. Available Phosphorus Bray I
GLOSOLAN-SOP-BRAY I/01
Gina P. Nilo, Ph.D.
Philippines

2. What is extraction of P by Bray I?
• The extraction of P by the Bray P1 (Bray and Kurtz, 1945) method is
based upon the solubilization effect of the H+ ions on soil P and the
ability of the F– ion to lower the activity of Al3+ and, to a lesser
extent, that of Ca2+ and Fe3+ ions in the extraction system.
• Consequently, the method should normally be limited to soils with
water pH values less than 6.8 when the texture is silty clay loam or
finer.
Source: B. Jones (2001)
Brief Background

3. Common Procedures/Steps
among SEALNET member
countries

4. Particle Size
Country Particle Size
Bangladesh
≤ 2 mm
Bhutan
China
India
Indonesia
Myanmar
Nepal (Nepal Agricultural
Research Council)
Nepal (SMD)
Philippines
Vietnam
Malaysia 0.250 mm
SOP: ≤ 2 mm

5. Sample/Extractant Ratio
Country
Sample
Weight, g
Volume of
Extracting
Solution, mL
Ratio of Soil to
Extracting Solution
Malaysia
2 g
20 mL 1:10
Myanmar 10 mL 1:5
Nepal (Nepal Agricultural
Research Council)
20 mL 1:10
Philippines 14 mL 1:7
Vietnam 14 mL 1:7
Bhutan
5 g
35 mL 1:7
India 50 mL 1:10
China 50 mL 1:10
Bangladesh
2.5 g 18 mL 1:7.2
Indonesia 25 mL 1:10
Nepal (SMD) 2.85 g 20 mL 1:7.02
Sample Weight:
2 grams
Extracting Solution:
14 or 20 mL
Sample Extractant Ratio:
1:7 or 1:10
Note:
1. Original soil/extractant ratio is 1:7 (1g to 7mL.
(ISRIC, 2002; Kurtz, 1945 & Jones, 1998)
2. 1:10 (2g & 20mL) ratio is adjusted only as per
procedure in B. Jones, 2001
Ratios:
1:7 – 3 labs
1:10 – 5 labs

6. Shaking Time and Shaking Method
Country Shaking Time, min
shaking method
(hand, shaker)
Bhutan
1 min
By Hand
Malaysia Shaker
Myanmar By Hand
Nepal (SMD) Shaker
Philippines Shaker
Bangladesh
5 mins
Shaker
India Shaker
Indonesia Shaker
Nepal (Nepal Agricultural
Research Council) Shaker
China 30 mins (25±1℃) Shaker
Vietnam 0.6 min By Hand
Shaking Time:
1 minute
5 labs
Shaking method:
Shaker (reciprocating)

7. Filter Paper No. and Filtration Time
Country Filter Paper No. Filtration time, min
Bangladesh
Whatman No. 42
10 minutes
India 10-15 min
Indonesia
Not to exceed
5 minutes
Malaysia Not exceed to 10 minutes
Myanmar Not exceed to 10 minutes
Nepal (SMD) -
Nepal Nepal Agricultural
Research Council
5 minutes
Philippines Not exceed to 10 minutes
Vietnam Not exceed to 10 minutes
China
No.102 (max pore
size 15-20 μm Filter Now
Bhutan
Whatman No. 5 or
Equivalent
10-15 minutes depending on
nature of sample
Filtration Time:
Not to exceed 10 mins
Filter Paper No.:
Whatman No. 42

8. COMPOSITION OF MIXED REAGENT
Country Composition of Mixed Reagent
Bangladesh
Ascorbic Acid Method
Deionized Water, Sulfuric acid, Ammonium molybdate
solution, Ascorbic acid solution, Potassium antimony
tartrate solution
China
Indonesia
Malaysia
Nepal (Nepal Agricultural
Research Council)
Philiipines
Vietnam
India
Malaysia
Bhutan
Nepal (SMD)
Stannous Chloride Method
Chloromolybdic acid, Ammonium molydbate (NH4)6Mo7O24.4H2O,
HCl, Stannous Chloride stock solution, SnCl2.2H2O Stannous Chloride
working solution, Stannous Chloride stock, Distilled water
Composition of Mixed
Reagent:
Ascorbic Acid Method

9. Standard Series*, mg/L used, Volume of Standard Series, mL, Method
Blank, mL, and Sample Extracts, mL
Country Standard Series*, mg/L
Volume of
Standard Series,
mL
Volume of
Method Blank,
mL
Volume of Sample
Extracts, mL
Bhutan 0, 0.5, 1.0, 2.0, 3.0, 4.0
*Volume to mark with
deionized water in 100 mL
volumetric flask
0.32 mL 0.32 mL 0.32 mL
Malaysia 5 mL 5 mL 5 mL
Bangladesh
0.0, 0.1, 0.2, 0.3, 0.4
*Volume to mark with
distilled water in 50 mL
volumetric flask
50 mL 50 mL 5 mL
China
0.00, 0.10, 0.20, 0.40, 0.60,
0.80, 1.00
*Volume to mark with
deionized water in 50 mL
volumetric flask
10 mL 10 mL 10 mL
India
0.5, 1, 1.5, 2, 2.5 ppm
*Volume to mark with
distilled water in 100 mL
volumetric flask
100 mL 5 mL 5 mL
Standard Series, mg/L:
Varies per
reference

10. Continuation of Standard Series*, mg/L used
Indonesia
0 PO4, 2, 4, 8, 12, 16, 20
*Volume to mark with deionized water
in 100 mL volumetric flask
2 mL 2 mL 2 mL
Myanmar
0.2, 0.4, 0.6, 0.8, 1.0
*Volume to mark with deionized water
in 50 mL volumetric flask
1 mL 1 mL 1 mL
Nepal (Nepal
Agricultural Research
Council)
0.2, 0.4, 0.8, 1,2 25 mL 25 mL 5 mL
Nepal (SMD)
0, 0.04, 0.08, 0.16, 0.24, 0.32, 0.4
*Volume to mark with distilled water in
50 mL volumetric flask
50 mL 50 mL 5 mL
Philippines
0, 1.2, 2.4, 3.6, 4.8, 6.0
*Volume to mark with deionized water
in 100 mL volumetric flask
1 mL 1 mL 1 mL
Vietnam
0, 0.2, 0.4, 0.6, 0.8, 1.0, 1.2, 1.4, 1.6, 2.0
*Volume to mark with the extracting
solution in 100 mL volumetric flask
Volume of Standard
Series, Blank &
Sample Extract mL:
Varies per
reference
Country Standard Series*, mg/L
Volume of
Standard
Series, mL
Volume of
Method
Blank, mL
Volume of
Sample
Extracts, mL

11. Volume of 1% Boric Acid
Volume 1% Boric
Acid, mL:
2 mL
Country
Volume of 1% Boric
Acid, mL
China
2 mLMyanmar
Philippines
Malaysia 1 mL
Nepal (Nepal Agricultural Research
Council)
5% boric acid (7.5 ml in
each )
Nepal (SMD) 15 mL 5% Boric Acid
Bhutan N/A
India N/A
Indonesia N/A
Vietnam N/A
Note:
It will vary depending on the reference

12. Volume of Mixed Reagent, mL
Volume Mixed
Reagent, mL:
Varies with
references
Country
Volume of the Mixed
Reagent, mL
Bangladesh
10 mLIndonesia
Nepal (SMD)
Bhutan 0.42 mL
China 5 mL
India 4 mL
Malaysia 15 mL
Myanmar 3 mL
Nepal (Nepal Agricultural
Research Council)
4 mL
Philippines 3 mL
Vietnam N/A
Note:
ISRIC has a very good description of the steps from
preparing calibration standards, mixed reagent,
etc.

13. Standing time for color development, min
Standing Time:
*References suggests 30
mins and 60 mins to
develop its maximum
Country Standing time for color development, min
Indonesia
At least 30 mins to develop its maximum
Vietnam
Philippines At least 60 mins to develop its maximum
Myanmar 60 mins
Bangladesh 15 mins
Malaysia At least 120 mins to develop its maximum
Nepal (Nepal Agricultural
Research Council)
10 minutes
Nepal (SMD) At least 5 mins to 15 mins develop its maximum
Bhutan -
India 10 mins to develop the colour
China 30 mins

14. Wavelength, nm
Wavelength, nm:
882 nm
Country Wavelength, nm
Bhutan
882 nm
Myanmar
Nepal (Nepal Agricultural
Research Council)
Philippines
Vietnam
Bangladesh 890 nm
China 700 nm
India 880 nm
Indonesia 889 nm
Malaysia
660 nm
Nepal (SMD)

15. Reference Materials Used
Reference
Materials:
NIST Traceable
Phosphate
Standard Solution
(1000 mg/L),
Certified
Reference
Material for Soil,
Check Sample
(Internal Sample)
Country Reference Materials
Malaysia
NIST Traceable Phosphate Standard Solution (1000 mg/L), Certified
Reference Material for Soil, Check Sample (Internal Sample)Philippines
Vietnam
Bangladesh Check Sample (Internal Sample)
Myanmar KH2PO4 solution (100 mg P/L), Check Sample (Internal Sample)
Nepal (Nepal Agricultural
Research Council)
NIST Traceable Phosphate Standard Solution (1000 mg/L) 2. Local
Control (Master) Sample
Nepal (SMD)
Standard Solution (AR KH2PO4 ), Certified Reference Material for
Soil, Check Sample (Internal Sample)
Bhutan Internal Master sample, and standard as Drift for quality control.
India
Check sample (Internal sample) already tested with NIST and WAS
international reference standard material
Indonesia Check Sample (Internal Sample)
China
Phosphate Standard Solution (1000 mg/L), Certified Reference
Material for Soil, Check Sample (Internal Sample)
Note:
Preparation of Check Sample
should follow ISO Guide 80

16. Quality Control Measures
Quality Control
Measure:
1. Duplicate analysis
in every 10 samples
and employing
Relative Standard
Deviation, %
2. Quality Control
Chart + Grubb's
Test and Shapiro-
Wilk Test for
Outliers
3. Method Blank,
Method Detection
Limit
4. Participation in
Proficiency Testing
Programs
Country Quality Control Measures
Indonesia Duplicate analysis in every 10 samples and employing Relative Standard Deviation,
% Quality Control Chart + Grubb's Test and Shapiro-Wilk Test for Outliers, Method
Blank, Method Detection Limit, Participation in Proficiency Testing ProgramsMalaysia
Philippines
Bangladesh Duplicate analysis in every 10 samples, Method Blank, Method
Myanmar
10% duplication (2 samples are duplicate in one batch-20 samples) - Quality Control
Chart
Nepal (Nepal
Agricultural Research
Council)
Setting spectrophotometer for measuring a sample in 2 times 2. Using 2 Local
Control (Master Sample) in each lot of samples 3. Trying to make lab temperature 25
degree as much as possible 4. Including 2 blank samples in each lot of measurement
Nepal (SMD)
Duplicate analysis in every 10 samples and employing , Method Blank, Method
Detection Limit
Bhutan
Internal Master sample, standard as Drift for quality control, and control chart based
on Internal Master sample values.
India
Duplicate analysis, keeping internal check sample and check standard in every 10
samples
China
Duplicate analysis in every 10 samples , Method Blank, Method Detection Limit,
Participation in Proficiency Testing Programs
Vietnam -

17. Reported Unit and Number of Decimal Places
SOP for Unit, nm:
mg/Kg
Country Unit
Decimal
Places
Bangladesh
mg/Kg
2
Bhutan 3
China 1
India 2
Indonesia 2
Myanmar 1
Nepal ( Nepal Agricultural
Research Council)
2
Philippines 2
Vietnam 2
Malaysia mg/L 2
Nepal (SMD) P2O5 Kg/ha 2
SOP for Decimal
Places:
2 Decimal Places

18. Report base on air dry or oven dry
Report base on air
dry or oven dry:
Oven Dry Basis
Country Oven Dry or Air Dry
Bhutan
Oven Dry Basis
India
Indonesia
Myanmar
Nepal ( Nepal Agricultural
Research Council)
Philippines
Vietnam
Malaysia
Air Dry Basis
Bangladesh
China
Nepal (SMD)

19. Factors Affecting Analysis of Available P – Bray I
1. Sample particle size
2. Extraction time
3. Composition of mixed reagent
4. Wavelength

20. Effect of Particle Size, mm
• Sieving through a 10-mesh (2-mm) screen removes stones and other
extraneous substances, yielding a uniform sample that can be easily
handled in the laboratory and stored indefinitely (J.Benton Jones).
• The soil is screened through a 2-mm sieve. The practice of passing
only a portion of the ground sample through the sieve and discarding
the remainder is erroneous. This introduces a positive bias in the
sample as the rejected part may include soil elements with
differential fertility. Therefore, the entire sample should be passed
through the sieve except for concretions and pebbles of more than
2 mm (Motsara, M.R, Roy, R.N)
Conclusion: The size of the sample to be use must be uniformed.

21. Extraction Time
Extraction time is based on time necessary to reach
equilibrium. For most acid extraction reagents, equilibrium is
reach quickly, and the time of the extraction is based more on
ease of mechanical handling. For Bray I, equilibrium is
reached in 60 s or 1 min. Long periods of contact between soil
and extraction reagent, by overextending the extraction time
or delaying separation by either filtration or centrifugation,
allow P reabsorption to take place.
Source: B. Jones (2001) - Laboratory Guide for Conducting Soil Tests and Plant Analysis.

22. COMPOSITION OF MIXED REAGENT
• The Mo-ascorbic blue methods are the most sensitive and, as a result, are widely
used for soil extract containing small amounts of P as well as for total P in soils. The
intensity of the blue color varies with the P concentration but is affected also by other
factors such as acidity, arsenates, silicates, and substances that influence the oxidation-
reduction condition of system. (R.H Miller, Methods of Soil Analysis Part 2)
• The advantages of the ascorbic acid method (Murphy & Riley, 1962) over the
stannous chloride method are the longer stability of the molybdenum blue color and
tolerance to high salt and Fe3+ (up to 2.5 mgL-1) concentrations.
• With the acid molybdate solution phosphate forms phospho-molybdenic acid which is
reduced to phospho-molybdenic with Ascorbic Acid. The antimony accelerates the
development of the blue colour and stabilizes this for up to 24 hrs. With this method
interference of Si is not expected. (ISRIC, 2002).

23. Wavelength, nm
• When the method was developed it was recommended that the
determination be made at 660 nm using a spectrophotometer.
• However, subsequent research has shown that the 882 nm wavelength is
superior and does not have a soil soluble organic interference that is noted
for the 660 nm wavelength.
• The absorption maximum of the phosphomolybdenum blue formed in the
presence of antimony is at 882 nm
Source:
Murphy, J., & Riley, J. P. (1962). A modified single solution method for the determination of phosphate in
natural waters. Analytica Chimica Acta, 27, 31–36

24. Wavelength, nm
Source: Murphy, J., & Riley, J. P. (1962). A modified single solution method for the determination
of phosphate in natural waters. Analytica Chimica Acta, 27, 31–36

25. Thank you for your attention.