SOIL ANALYSIS:

Soil sample were used for the experiment were subjected to chemical analysis. In accordance to
the methods of Eno, J. U., Ibia, T. O., Ogunwale, J. A., Ano, A. O. and Esu, I. E.
(2009). Mannual of soil, plant and water Analysis. Sibon books Ltd, Lagos. 216p.

Ibitoye, A. A. (2008). Laboratory Manual on Basic Soil Analysis 3rd Edition. Foladave
Nig. limited.

Total Nitrogen determination (using Kjeldahl method)

Digestion

1g of air-dried soil of fine tilth was weighed into a 250ml kjeldahl flask.

One catalyst tablet (comprising of CuSo4, K2SO4 and a pinch of selenium) was added.

The mixture was heated till it becomes clear (light green colour).

It was allowed to cool after it was removed from heat.

About 10ml of water was added and the content was filtered using Whatman 45 filter paper into

100ml volumetric flask, it was made up to mark and shaken together for proper mixing.

Distillation

10ml aliquot was transferred into 500ml kjeldahl flask.

30ml of water was added.

15ml 0f NaOH (excess base) was added.

Heat was applied.

25ml distillate was collected in 5ml boric acid indicator

The NH4-Nitrogen was determined by titrating distillate with 0.01M standard HCl.

NB: Colour changed from green to pink and %N was calculated as follows:

T X M X 14 X V1 X 100

1000 V2 W

Where:

T = Titre value of sample

M = Concentration of HCl used for titration in molarity

1000 = constant

V1 = Final volume

V2 = Volume of aliquot used for distillation

W = Weight of sample used

Available Phosphorus Determination (Bray-1 Method).

5g of soil sample was weighed into extractions cup.

30ml of Bray-1 solution was added

It was stirred in a mechanical shaker for 5mins.

It was then filtered into a reagent bottle.

1ml of extract (aliquot) was pipetted into a 50ml volumetric flask.

6ml of distil water was added.

2ml of colour developing reagent was added and mixed well.

1ml of ascorbic acid solution was added.

It was left for about 10minutes for the colour to develop.

The solution was measured at 650nm in visible-range spectrophotometer

A graph of absorbance versus ppm standard and ppm P interpolation was done

Calculation

Ppm P(µg P/kg of soil) = R x 30

=Rx6

Organic Carbon (using the Walkley-Black wet-oxidation Method)

1g of soil was weighed into 250ml conical flask

10ml of potassium dichromate was added.

20ml of concentrated H2SO4 was added vigorously.

It was allowed to stay for 30minutes.

100ml of water was added.

5 drops of ferrion indicator was added.

The mixture was titrated with 0.5N ferrous sulphate (FeSO4.7H2O).

A blank titre was also taken but without soil sample.

% Organic carbon was calculated as follows:

(B-T) X N X 0.003F X 100

Where:

B = Blank titre value

T = Sample titre value

N = concentration of FeSO4 in normality

F = Correction factor (1.33)

NB: % Organic Carbon was converted to % organic matter by multiplying it by 1.724

and answers were reported in g/kg by multiplying % organic carbon or %organic matter by 10.

Exchangeable Bases. (K, Ca, Mg, Na)

Ammonium Saturation Method

10g of soil was weighed into a 250ml soil shaking bottle.

100ml of 1N Ammoniun Acetate was added and was shaken in a mechanical shaker for 1 hour

The soil was filtered using a Whatman No 45 filter paper into a 100ml volumetric flask and

made up to mark with Ammoniun Acetate solution and stored in a 100ml plastic

Reagent bottle.

K and Na were read in a “ Jennway model” flame photometer and Ca and Mg were read in an

unicam series 969 model Atomic Absorption Spectrophotometer (AAS).

pH Determination (Electronic pH Meter Method)

Soil pH in H2O 1:1

10g of soil was weighed into 50ml beaker.

10ml of distilled water was added.

The mixture was stirred for 30minutes intermittently, and pH was read in a standardized pH

meter.

Confirmation with CaCl2

Add two drops of 1m CaCl2 solution into the solution and stirred for 10 minutes intermitently

and read the pH using a standardized pH meter.

Exchange Acidity (E.A-Titration Method)

5g of air-dried soil was weighed into a soil shaking bottle.

50ml 0f 1M KCl was added and the mixture shaken for 1 hour in a mechanical shaker.

The soil was filtered into a 100ml volumetric flask and was made up to 100ml mark

with 1M KCl solution.

25ml of soil extract was measured into a 250ml conical flask

5drops of phenolphthalein indicator was added.

Mixture was titrated with 0.05N NaOH to pink endpoint.

NB: The exchangeable acidity was calculated as follows:

V X 0.05 X 100 (Mg eqivalent/100g )

Where:

W = weight of sample used

V = Titre value

For Exchangeable Aluminum in soil

5ml of 3N NaF was added to the titrated extract.

The mixture was titrated with 0.05N HCl to end point.

NB: Exchangeable Aluminium was calculated Meq/100g as follows:

V X 0.05 X 100 (Meq/100g)

(mMol/kg) = Mq/100g x 2

Meq/100g x 2 = Mmol/kg = (mol/kg}

Where:

W = weight of sample used

V = Titre value

Particle Size Determination

(Hydrometer Method)

51g air dry soil (<2mm) into 250ml beaker.

50ml of calgon (sodium hexamatoxphosphate + sodium carbonate)

100ml of distilled water was added and stirred vigorously for 1minute using glass rod

and allowed to stand for 30minutes.

The suspension was transferred into a mixer and mixed for 15minutes at a medium speed.

The suspension was transferred into a sedimentation cylinder and make up to 1litre

mark using distilled water.

Measurements

Mix the suspension in the cylinder by several vertical movement of the plunger. Place the

cylinder on a flat surface and then time. The first reading was taking after 40seconds by sliding

the hydrometer slowly into the suspension, and thereafter temperature reading taken. The second

reading was taken 2hours later and was calculated to determine percentage sand silt and clay.

CEC (Cation Exchangeable Capacity) = Ca+Mg+Na+K+H

ECEC = Effective cation exchangable capacity = Ca+Mg+Na+k+EA

% base saturation = Ca+ Mg +Na +k x 100

Ca+Mg+Na+K+EA 1

STATISTICAL ANALYSIS
All analysis was conducted in triplicates, mean scores of the results and their standard deviations

were reported. Data were subjected using analysis of variance and Duncan’s multiple range

(Duncan, 1955) test was used to separate the means .

Eno, J. U., Ibia, T. O., Ogunwale, J. A., Ano, A. O. and Esu, I. E. (2009). Mannual of soil, plant
and water Analysis. Sibon books Ltd, Lagos. 216p.

Ibitoye, A. A. (2008). Laboratory Manual on Basic Soil Analysis 3rd Edition. Foladave Nig.
limited.

Determination of crude protein content: The crude protein of the sample was
determined using the micro – Kjeldahl method described by AOAC (1990). The
principle of this method is based on the transformation of protein and that of the
other nitrogen containing organic compounds, other than nitriles and nitrates into
ammonium sulphate by acid digestion.
Sample nitrogen + H2SO4(aq) Catalyst (NH4)2SO4(aq)
(NH4)2 SO4(aq) + 2NaOH (aq) 2NH3(aq) + 2H2O + Na2SO4(aq)
NH3(aq) + H3BO3 (aq) NH+4 (aq) + H2BO-3 (aq)
H+(aq) + H2BO-3(aq) H3BO3 (aq)
The sample (0.5g) was weighed into a micro – Kjeldahl digestion flask. Of foss
automatic digester block system, It was shaken and allowed to stand for sometime.
One tablet of selenium catalyst with a mixture of 2:1 cupper sulphate and
potassium sulphate was added followed by the addition of 20cm3 concentrated
sulphuric acid. The flask was heated on the digestion block at 4500C for 1 hours
until the digest became clear. The flask was removed from the block and allowed
to cool. The content was transferred into 100cm3 volumetric flask and diluted to
the mark with water.
An aliquot of the digest (10cm3) was transferred into another micro-Kjeldahl flask
along with 20cm3 of distilled water, and placed in the distilling outlet of the micro
– Kjeldahl distillation unit. A conical flask containing 20cm3 of boric acid
indicator was placed under the condenser outlet. Sodium hydroxide solution
(20cm3, 40%) was added to the content in the Kjeldahl flask by opening the funnel
stopcock. The distillation start and the heat supplied was regulated to avoid
sucking back. When all the available distillate was collected in 5cm3 of boric acid
mix indicator, the distillation was stopped. The nitrogen in the distillate was
determined by titrating with 0.1N of HCl; the end point was obtained when the
colour of the distillate changed from green to pink.

Crude protein is a measure of nitrogen in the sample. It was calculated by

multiplying the total nitrogen content by a constant, 6.60. This is based on the
assumption that, proteins contain about 16%N which includes both true protein and
non – protein N and does not make a distinction between available or unavailable
protein (Udo and Ogunwele, 1986). The crude protein was calculated using

% crude protein = %N x 6.25

The nitrogen content of the sample is given by the formula below.

where
Tv = Titre value of acid (cm3)
Na = Concentration or normality of acid (0.1N HCl)
V1 = Volume of distilled water used for distilling the digest (100cm3).
V2 = Volume of aliquot used for distillation (10cm3)
G = Original weight of sample used, g
0.014 = milliequivalent of Nitrogen
100 = percentage

Mineral analysis: The minerals were analyzed by dry ashing the samples at 550°C
to constant weight and dissolving the ash in volumetric flask using distilled, de-
ionized water with and 20 % of 10ml hydrochloric acid were added and then filter
and make up to mark of 100ml. the solution were used in the determination of the
elements using by Atomic Absorption spectrophometer unican 939 series
(Agte et al., 1995), while sodium and potassium were determined using flame
photometry (Chapman and Pralt, 1961) using NaCl and KCl to prepare the
standards.

Micronutrient Determination {Available Nutrient}

(Using DPTA-TEA Extract method)

10g of soil was weighed into a soil shaking bottle, 50ml of DPTA-TEA solution was added and

suspended, it was shaked I for 2hours in mechanical shaker.The soil was filtered using whattman

No 1filter paper into a 100ml volumetric flask ready for micro-nutrient determination ( Fe, Mn,

Zn, Cu) using an Atomic Absorption Spectrophotometer (AAS).

The microbial populations counts were determined by the serial dilution, using

1 x 10 dilutions factor.

A total of 5 dilutions were done per sample and plating was done from the 4th

dilution, and calculated accordingly. The agar (Nutrient Agar, potatoe Dextrose

Agar ) used were incubated after inculation.

The counting were done with the aid of land lens dialing. Counting was done for 5

days before it becomes impossible to count because counting was physically done.

REFERENCES

The McGraw-Hill (2001): Benson: Microbiological Applications Laboratory

Manual Eighth Editions: pp154-169.

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