UROKO, ROBERT IKECHUKWU (PG/M. Sc/12/61371) Robert.pdf · impact of fresh and fermented palm oil mill effluents on soil physicochemical parameters and enzyme activities in umuaka,

EFFLUENTS ON SOIL PHYSICOCHEMICAL PARAMETERS

AND ENZYME ACTIVITIES IN UMUAKA, NJABA, IMO STATE

Ugwoke Oluchi C.

FACULTY OF BIOLOGICAL SCIENCES

i



OF NIGERIA

Digitally Signed by: Content manager’s

DN : CN = Webmaster’s name

O = University of Nigeria, Nsukka

OU = Innovation Centre

Ugwoke Oluchi C.

UROKO, ROBERT IKECHUKWU

(PG/M. Sc/12/61371)


DEPARTMENT OF BIOCHEMISTRY



: Content manager’s Name

me

a, Nsukka




ii

IMPACT OF FRESH AND FERMENTED PALM OIL MILL


AND ENZYME ACTIVITIES IN UMUAKA, NJABA, IMO

STATE OF NIGERIA

BY


(PG/M. Sc/12/61371)


UNIVERSITY OF NIGERIA

NSUKKA

JULY, 2014

iii

TITLE PAGE

IMPACT OF FRESH AND FERMENTED PALM OIL MILL EFFLUENTS ON SOIL

PHYSICO-CHEMICAL PARAMETERS AND ENZYME ACTIVITIES IN UMUAKA,

NJABA, IMO STATE OF NIGERIA

A RESEARCH PROJECT SUBMITTED IN PARTIAL FULFILLMENT FOR THE

AWARD OF MASTERS OF SCIENCE (M.Sc) IN INDUSTRIAL BIOCHEMISTRY

AND BIOTECHNOLOGY


(PG/M.Sc/12/61371)


UNIVERSITY OF NIGERIA,

NSUKKA

SUPERVISOR:

PROF. OBI U. NJOKU

JULY, 2014

iv

CERTIFICATION

This is to certify that Uroko, Robert Ikechukwu, a postgratuate student with

registration number PG/M.Sc/12/61371 in the Department of Biochemistry, University of

Nigeria, Nsukka, has satisfactorily completed the requirement for course work and research

for the degree of Masters (M.Sc) in Industrial Biochemistry and Biotechnology. The work

embodied in this report is original and has not been submitted in part or in full for any other

diploma or degree of this or other University.

------------------------------------------- ----------------------------------------

-----

PROF. O. U. NJOKU PROF. O.F.C. NWODO

(Supervisor) (Head of Department)

------------------------------------------

EXTERNAL EXAMINER

v

DEDICATION

I dedicate this research work to the Almighty God for the strength, grace and

protection bestowed on me during the course of this research work, Prof. H.C Nzelibe my

academic father, Uroko Ngozi Jacinta,Agu Gladys Nneka, Ocean Helen Ojomachenwu and to

all my friends, most especially Ozioko Chidimma Lilianfor their prayers and supports.

vi

ACKNOWLEDGEMENTS

It is pertinent to acknowledge the great feeling of relief and happiness to know that I

have completed my postgraduate programme. It has been two years of hard work and being

away from family and friends, but the experience of studying and living in Nsukka will be

unforgettable. During this journey, I have had the pleasure to meet great and interesting

people and, to be taught by great lecturers who I never imagined. Thus, this whole experience

enriched my knowledge, my personal development and gave me the chance to realize that I

am capable of doing more than I thought.

In the first place, I would like to express my gratitude to my supervisor, Prof. Obi U.

Njoku, who believed in my capability to carry out this project. He guided and supported me,

not just through this project but during my two years of studies in the University of Nigeria,

Nsukka. His work has been an inspiration for my future development as a biochemist.

A warm “thank you” to all my colleagues from the Department of Biochemistry who

gave me the opportunity to be part of them and have a great experience from the University

of Nigeria, Nsukka. During my studies, I met and worked with wonderful people, who made

my living experience in the University of Nigeria, much happier and busier.

I highly appreciate the immense contributions of Dr. Parker E. Joshua,Dr (Mrs). C.A.

Anosike,Dr. C. S. Ubani, Agu Chidozie Victor, Okonkwo Chukwudi C. and the entire staff of

the Department of Biochemistry, Projects Development Institute (PRODA), Enugu,

Department of Soil Science and Crop Science, University of Nigeria, Nsukka for their

immense contributions in making this work a reality. I am also grateful for the patient and

support of my friends most especially Ozioko, Chidinma Lilian, Ani Chijoke Collins, Aruma,

Onyedika, Ocean Helen Ojomachenwu and Nweje, Anyalowu Paul, whom during these two

years were as my family.

Last but not the least an enormous thanks to my family, for their love, support and

understanding during the years of being at the University of Nigeria, Nsukka. To my mother

who is a great personality, worthy of admiration and through all the years, she has been my

role model. Her strength and guidance has helped me to become better in my personal and

professional life. To my brothers Benjamin, Celestine, Thaddeus, Ignatius and my sisters

Ngozi and Eucharia; who always remind me, that it is important to work hard and enjoy life,

as it is.

vii

ABSTRACT

The impact of palm oil mill effluents (POME) on soil fertility was determined by studying the physico-chemical and heavy metals parameters found in fresh and fermented POME from six palm oil milling sites. Some soil enzymes (like catalase, dehydrogenase and lipase) and physico-chemical parameters of POME polluted soil wereevaluated from six dumpsites and soil from ten-yards distance from dumpsites in relation to the control farmland in the area. The fresh and fermented POME samples were randomly collected aseptically from small scale palm oil milling sites in Umuaka inNjaba Local Government Area ofImo State, Nigeria. The soil samples were collected aseptically into sterile containers from topsoil (0 – 15cm deep), and subsoil (15 – 30cm deep) at dumpsites, ten-yards distance from dumpsites and farmland one kilometre from various dumpsites. The study of both soil and POME samples were carried out using standard analytical procedures. The analysis of variance of the results of physicochemical parameters in POME samples showed that there were significant(p<0.05) differences in all the parameters which include: pH, magnesium ion (Mg2+), calcium (Ca2+), potassium (K+), sodium (Na+), total solid (TS), suspended solids (SS), total volatile solids (TVS), chemical oxygen demand (COD), dissolved oxygen (DO), nitrogen (N) and phosphorus (P) in fermentedPOME when compared tofresh POME.The concentration of heavy metals in the POME samples showed that they were rich in chromium, copper, iron and lacked cadmium. The results of the soil fertility parameters showed that POME polluted soils are rich in soil nutrients such as nitrogen, organic carbon (OC), organic matter (OM), cation exchange capacity (CEC) and most especially exchangeable cations (Mg2+, Ca2+, K+ and Na+) in topsoils and subsoils than in their respective non POME polluted soils. Although the results of the soil fertility analysis showed that POME-polluted soil had high fertility index, it has low phosphorus content which was due to negative effect of the acidic pH of the POME on phosphorus availability.The analysis of the results of activities of soil enzymes in the POME- polluted soil showed that discharge of the fresh and fermented POME caused induction of lipase activity. Catalase activitydecreasedsignificantly (p<0.05)from 1.35±0.02 to 0.33±0.01 mM H2O2/g soil/h for topsoil and 2.08±0.08 to 0.21±0.01mM H2O2/g dry soil/h for subsoil from dumpsites respectively while dehydrogenase activity in topsoil and subsoil ten yards away from dumpsitedecreasedsignificantly (p<0.05)from 1.35±0.02 to 0.18mM H2O2/g soil/h and 2.08±0.08 to 0.21±0.01mM H2O2/g dry soil/h,respectively. Dehydrogenase activitydecreasedsignificantly (p<0.05) from 0.76±0.001 to 0.10 mg/gdrysoil/96h for topsoil and 0.86±0.01 to 0.077±0.01mg formazan/gdrysoil/96h for subsoil from dumpsites while dehydrogenase activity in topsoil and subsoil ten yards away from dumpsitedecreasedsignificantly (p<0.05) from 0.76±0.001 to 0.10±0.03 mgformazan/gdrysoil/96h and 0.86±0.01 to 0.07±0.01 mgformazan/gdrysoil/96h, respectively. Furthermore, the results obtained for the soil pH indicate that the soils are acidic and have high exchangeable acidity. Considering the high fertility potentials of POME, it is necessary to make good effort to maximize these potentials by reducing its high pollution level such as BOD, COD, heavy metals, oil and grease in the fresh and fermented POME.

viii

TABLE OF CONTENTS

Title Page- - - - - - - - - - - -i

Declaration- - - - - - - - - - - -ii

Certification- - - - - - - - - - - -iii

Dedication- - - - - - - - - - - -iv

Acknowledgement- - - - - - - - - - -v

Abstract- - - - - - - - - - - -vi

List of Figures- - - - - - - - - - -

xiii

List of Tables- - - - - - - - - - - -xiv

CHAPTER ONE: INTRODUCTION

1.1 Scientific classification of oil palm- - - - - - - -2

1.2 Palm seeds- - - - - - - - - - -3

1.3 Palm oil production- - - - - - - - - -3

1.4 Properties of POME- - - - - - - - -6

1.4.1 Physicochemical characterisation of POME- - - - - - -7

1.4.1.1 Dissolved oxygen- - - - - - - - - -7

1.4.1.2 Biochemical oxygen demand- - - - - - - -8

1.4.1.3 Chemical oxygen demand- - - - - - - - -8

1.4.1.4 Total solid- - - - - - - - - - -8

1.4.1.5 Suspended solids- - - - - - - - - -9

1.4.1.6 Total volatile solid- - - - - - - - - -9

ix

1.4.1.7 Oil and grease- - - - - - - - - -9

1.5 Heavy metals- - - - - - - - - - -9

1.5.1 Impact of heavy metals on soil enzymes- - - - - - -10

1.6 Socioe-conomic importance of POME- - - - - - - -12

1.6.1 Pre-treatment- - - - - - - - - - -13

1.6.2 Biological treatment technology- - - - - - - -13

1.6.3 Land application system- - - - - - - - -13

1.6.4 Evaporation technology- - - - - - - - -13

1.7 General uses of POME- - - - - - - - - -14

1.7.1 Biogas- - - - - - - - - - - 14

1.7.2 Bioplasticformation- - - - - - - - - -15

1.7.3 Bioethenol formation- - - - - - - - - -15

1.7.4 Compost and Biofertilizer- - - - - - - - -15

1.7.5 Medium for isolation and growth of Micro-organisms- - - - -16

1.7.6 Citric acid production- - - - - - - - - -16

1.7.7 Carotenoid extraction- - - - - - - - - -17

1.7.8 Enzyme production- - - - - - - - - -17

1.8 Soil fertility- - - - - - - - - - -17

1.8.1 Soil pH- - - - - - - - - - - -18

1.8.2 Soil enzymes- - - - - - - - - - -19

1.8.2.1 Soil dehydrogenase- - - - - - - - - -20

1.8.2.2 Soil catalase- - - - - - - - - - -21

1.8.2.3 Soil lipase- - - - - - - - - - -21

x

1.8.2.4 General importance of the soil enzymes- - - - - - -22

1.9 Cation exchange capacity- - - - - - - - -22

1.9.1 Exchangeable potassium- - - - - - - - -23

1.9.2 Exchangeable calcium- - - - - - - - - -23

1.9.3 Exchangeable magnesium- - - - - - - - -23

1.9.4 Exchangeable sodium- - - - - - - - - -23

1.10 Soil organic matter (SOM)- - - - - - - - -24

1.11 Functions of SOM- - - - - - - - - -24

1.12 Soil organic carbon- - - - - - - - - -25

1.13 Aim and objectives of the Study- - - - - - - -26

1.13.1 Aim of the study- - - - - - - - - -26

1.13.2 Specific objectives of the study- - - - - - - -26

CHAPTER TWO: MATERIALS AND METHODS

2.1Materials- - - - - - - - - - - -27

2.1.1 Palm Oil Mill Effluents- - - - - - - - -27

2.1.2Soil samples- - - - - - - - - - -27

2.1.3 Equipment- - - - - - - - - - -27

2.1.4 Chemicals and Reagents- - - - - - - - -28

2.2 Methods- - - - - - - - - - - -29

2.2.1 Collection of the fresh and fermented palm oil mill effluents- - - - -29

2.2.2 Collection of the soil samples- - - - - - - - -29

2.2.3 Preparation of the soil samples- - - - - - - - -29

xi

2.2.4 Preparation of the fresh and fermented POME samples- - - - -29

2.2.5 Preparation of reagents- - - - - - - - - -29

2.2.5.1 Phosphate Buffer (0.05 N, pH 7.4)- - - - - - - -29

2.2.5.2 Concentrated Sulphuric Acid (6 N H2SO4)- - - - - - -29

2.2.5.3 Hydrogen Peroxide Solution (2 mMol H2O2)- - - - - -29

2.2.5.4 Potassium Permanganate Solution (0.1 N KMnO4)- - - - - -29

2.2.5.5 3% TTC (2, 3, 5-triphenyltetrazolium chloride)- - - - - -29

2.2.5.6 Potassium Chloride Solution (1 M KCl)- - - - - - -30

2.2.5.7 Ammonium Metavanadate Reagent- - - - - - - -30

2.2.5.8 Carbon Carbonate Solution- - - - - - - - -30

2.2.5.9 Sodium Chloride Solution (1 M NaCl)- - - - - - -30

2.2.5.10 Magnesium Chloride Solution (1 M MgCl2)- - - - - -30

2.2.5.11 10% (w/v) Potassium Hydroxide Solution (KOH)- - - - - -30

2.2.5.12 EDTA (Ethylenediamine tetra-acetic acid)- - - - - - -30

2.2.5.13 Formalin Solution- - - - - - - - - -30

2.2.5.14 Sodium Hydroxide Solution (0.1 NaOH)- - - - - - -

312.2.5.15 Potassium Dichromate Solution- - - - - - - -31

2.2.5.16 Ferrous Sulphate Solution- - - - - - - - -31

2.2.5.17 Sodium Acetate Solution- - - - - - - - -31

.2.2.5.18 Lauric Acid.- - - - - - - - - -31

2.2.5.19 40% Sodium Hydroxide (40% NaOH)- - - - - - -31

xii

2.2.5.20 Hydrogen Chloride (0.01 M HCl)- - - - - - - -31

2.2.6 Determination of the fresh and fermented POME pH- - - - -31

2.2.7 Determination of the soil pH and exchangeable acidity- - - - -32

2.2.8 Determination of total nitrogen- - - - - - - -33

2.2.9 Determination of available phosphorus- - - - - - -33

2.2.10 Determination of sodium and potassium ions- - - - - -34

2.2.11 Determination of Calcium Ions- - - - - - - -36

2.2.12 Determination of magnesium ions- - - - - - - -36

2.2.13 Determination of cation exchange capacity (CEC)- - - - - -37

2.2.14 Determination of heavy metals in the fresh and fermented POME samples- - -37

2.2.15 Determination of dissolved oxygen in the fresh and fermented POME samples- -37

2.2.16 Determination of the BOD in the fresh and fermented POME samples- - -38

2.2.17 Determination of COD in the fresh and fermented POME samples- - - -39

2.2.18 Determination of total solids, suspended solid and volatile solids in the POME- -40

2.2.18.1 Total solids and volatile solids in fresh and fermented POME- - - -40

2.2.18.2 Total suspended solids in fresh and fermented POME- - - - -40

2.2.19 Determination of soil organic carbon and soil organic matters- - - -41

2.2.20 Determination of oil and grease in POME- - - - - - -41

2.2.21 Determination of the soil dehydrogenase activity- - - - - -42

2.2.22 Determination of the soil catalase activity- - - - - - -43

2.2.23 Determination of the soil lipase activity- - - - - - -43

2.2.24 Statistical Analysis- - - - - - - - - -44

xiii

CHAPTER THREE: RESULTS

3.1 pH of the fresh and fermented POME samples- - - - - - -45

3.2 pH and exchangeable acidity of the soil contaminated with the fresh and fermented

POME - - - - - - - - - 47

3.3 Nitrogen content in the fresh and fermented POME- - - - - -49

3.4 Effect of the fresh and fermented POME on the nitrogen content in soil- - -51

3.5 Effect of thefresh and fermented POME on the phosphorus content in soil- - -53

3.6 Phosphorus content in the fresh and fermented POME- - - - - -55

3.7 Calcium content in the fresh and fermented POME- - - - - 57

3.8 Magnesium content in the fresh and fermented POME- - - - - -59

3.9 Sodium content in the fresh and fermented POME- - - - - -61

3.10 Potassium content in the fresh and fermented POME- - - - - -63

3.11 Calcium content of the soil contaminated with the fresh and fermented POME- -65

3.12 Potassium content in the soil contaminated with the fresh and fermentedPOME- -67

3.13 Sodium content in the soil contaminated with the fresh and fermented POME- -69

3.14 Magnesium content in the soil contaminated with the fresh and fermented POME- -71

3.15 Effect of the fresh and fermented POME on the cation exchange capacity in the soil- 73

3.16 Heavy metal contents in the fresh and fermented POME- - - - -75

3.17 Dissolved (DO) content in the fresh and fermented POME- - - - -77

3.18 Biochemical oxygen demand (BOD) content in the fresh and fermented POME- -79

3.19 Chemical oxyegen demand (COD) content in the fresh and fermented POME- -81

3.20 Total solidscontent in the fresh and fermentedPOME- - - - - -83

3.21 Total volatile solids content in the fresh and fermented POME- - - -85

xiv

3.22 Suspended solids in the fresh and fermentedPOME- - - - - -87

3.23 Organic carbon content in the soil contaminated with the fresh and fermented POME-89

3.24 Organic matter content in the soil contaminated with the fresh and fermented POME-91

3.25 Percentage content of the oil and grease in the fresh and fermented POME- - -93

3.26 Dehydrogenase activity in the soil contaminated with the fresh and fermented POME-95

3.27 Catalase activity in the soil contaminated with the fresh and fermented POME- -97

3.28 Lipase activity in the soil contaminated with the fresh and fermented POME- -99

CHAPTER FOUR: DISCUSSION

4.1 Discussion- - - - - - - - - - 101

4.2 Conclusion- - - - - - - - - - 112

4.3 Suggestions for Further Studies- - - - - - - - 113

References- - - - - - - - - - - 114

Appendices- - - - - - - - - - - 127

xv

LIST OF FIGURES

Fig. 1: Fresh fruit bunch- - - - - - - - - -3

Fig. 2: Palm seeds- - - - - - - - - - -3

Fig. 3: Palm oil mill effluent dumpsite- - - - - - - -6

Fig 4:Effect of the Fresh and Fermented POME on the Nitrogen Content in the Soil- -53

Fig. 5: Effect of the Fresh and Fermented POME on Phosphorus Content in the Soil- -55

Fig. 6:Calcium Content in the Soil Contaminated with the Fresh and Fermented POME- -67

Fig. 7: Potassium Content in the Soil contaminated with the Fresh and Fermented POME- 69

Fig. 8: Sodium Content in the Soil Contaminated with theFresh and Fermented POME- -71

Fig. 9: Magnesium Content in the Soil Contaminated with the Fresh and Fermented POME73

Fig. 10: Effect of the Fresh and Fermented POME on the Cation Exchange Capacity (CEC) in

the Soil- - - - - - - - - - -75

Fig. 11:Organic Carbon Content in the Soil Contaminated with the Fresh and Fermented

POME- - - - - - - - - - 91

Fig. 12:Organic Matter Content in the Soil Contaminated with the Fresh and Fermented

POME- - - - - - - - - -93

Fig. 13:Dehydrogenase activity in the soil contaminated with the fresh and fermented

POME- - - - - - - - - --97

xvi

LIST OF TABLES

Table 1: Summary units of operation in palm oil production- - - - -7

Table 2: Characteristics of POME and its respective standard discharge limit- - -8

Table 3: Classification of soil based on the pH range- - - - - -20

Table 4: The pH of the Fresh and Fermented POME- - - - - -47

Table 5: Effect of the Fresh and Fermented POME on the Soil pH and Soil Exchangeable

Acidity- - - - - - - - - -49

Table 6: Percentage nitrogen (%) in the Fresh and Fermented POME- - - -51

Table 7: Phosphosrus content in the fresh and fermented POME- - - - -57

Table 8: Perentage Calcium (%) Content in the Fresh and Fermented POME- - -59

Table 9: Percentage Magnesium (%) in the Fresh and Fermented POME- - - -61

Table 10: Percentage Sodium (%) in the Fresh and Fermented POME- - - -.63

Table 11: Percentage Potassium (%) in the Fresh and Fermented POME- - - -65

Table 12: Heavy Metal Content inthe Fresh and Fermented POME- - - -77

Table 13: Dissolved Oxygen (DO) Content in the Fresh and Fermented POME- - -79

Table 14: Biochemical Oxygen Demand content in the Fresh and Fermented POME- -81

Table 15: Chemical Oxygen Demand (COD) Content in the Fresh and Fermented POME- 83

Table 16: Total Solids in the Fresh and Fermented POME- - - - - -85

Table 17: Total Volatile Solids Content in the Fresh and Fermented POME- - -87

Table 18: Suspended Solids in the Fresh and Fermented POME- - - - -89

Table 19: Percentage Oil and Grease in the Fresh and Fermented POME- - - -95

1

CHAPTER ONE

INTRODUCTION

Palm oil mill effluent (POME) has been a major environmental concern in countries

producing them. This effluent is a land and aquatic pollutant when discharged fresh and

fermented due to the presence of moderate amount of organic load in it and its phytotoxic

properties (Okwute and Isu, 2007). The uses of wastes such as POME in agriculture and for

land reclamation are a common practice in regions with its abundant supply especially for

irrigation, soil conditioning, amendment and conservation purposes (Navas et al., 1998;

Pascual et al., 2007). Palm oil production requires addition of large quantity of water which is

eventually discharged as waste effluent (Nwoko and Ogunyemi, 2010). POME is a mixture of

water, oil and natural sediments (solid particles and fibres), large quantity of which is

generated annually during crude palm oil production (Salihu et al., 2011b) and is amenable to

microbial degradation (Nwoko and Ogunyemi, 2010; Nwoko et al., 2010). POME includes

dissolved constituents such as high concentration of protein, carbohydrate, nitrogenous

compounds, lipids and minerals, which may be converted into useful materials using

microbial processes. Nevertheless, POME if not discharged properly and treated, may lead to

considerable environmental problems (Singh et al., 2010). In nature, both nitrogen and

phosphorus come from the soil and decaying plants and animals (Logan et al., 1997; Navas et

al., 1998). Fertilizers, fresh and fermented sewage as well as domestic and wild animal

wastes are common sources of plant nutrients.

The input of effluent materials with high organic matter content will help replenish

the soil for sustainable agriculture. POME application to soil can result in increases of some

beneficial soil chemical and physical characteristics such as increases in organic matter,

carbon, major nutrients (such as nitrogen, potassium, calcium, and magnesium), water-

holding capacity and porosity (Logan et al., 1997; Navas et al., 1998). However, it may bring

about undesirable changes such as decrease in pH and increases in salinity (Kathiravale and

Ripin, 2000). These effects mostly occur very slowly and take many years to be significant.

Soil microbiological and biochemical properties have been considered early as sensitive

indicators of soil changes and can be used to predict long - term trends in the quality of soil

(Ros et al., 2003). Soil microbial properties are equally affected by environmental factors.

(Dick and Tabatabai, 1992) reported that high rate of inorganic fertilizer application

suppresses microbial respiration and dehydrogenase activity. Other factors such as increase in

salinity or decrease in water availability may also reduce biological activity (Paredes et al.,

2

2005). POME contains high organic load, substantial amounts of plant nutrients and represent

a low cost source of plant nutrients when fermented (Onyia et al., 2001). It is generally

believed that the toxic effect of POME is due to its possession of phenols and other organic

acids which are responsible for phytotoxicity and antibacterial activity (Capasso et al., 1992;

Piotrowska et al., 2006). However, the polyphenolic fraction is degraded with time and

partially transforms into humic substances (Piotrowska et al., 2006). Little information is

known on the impact of POME on the biochemical and microbial properties of soil. Studies

show that effects of wastes applied to soil occurred mainly in the first weeks after amendment

(Martens et al., 1992; Perucci, 1992; Binder et al., 2002). Indeed, investigating the effects

POME has on soil properties would help farmers mostly in rural areas to improve food

production through expanding their understanding of the importance of POME as well as the

quantity to be added to soil during farming operation prior to planting. Also, knowledge on

the remediating effect of POME on the soil will assist government in its drive to increase

food production by helping farmers to improve soil fertility through adequate harnessing and

processing of POME (Nwoko and Ogunyemi, 2010).

1.1 Scientific Classification of Oil Palm (Elaeis guineensis)

Elaeis guineensis is a member of the family Arecaceae (Reeves and Weihrauch,

1979). It is native to West and Southwest Africa and is vastly cultivated as a source of oil in

Nigeria. It has a lifespan of over 200 years, while the economic life is about 20-25 years. The

nursery period is 11-15 months and first harvest is done 32-38 months after planting (Reeves

and Weihrauch, 1979). The yield is about 45-56% of fresh fruit bunch and the fleshy

mesocarp of the fruit is used as oil source. The yield of oil from the kernel is about 40-50%

(Rupani et al., 2010). The plant is classified as follows:

Kingdom Plantae

Division Magnoliophyta

Class Liliopsida

Order Arecales

Family Arecaceae

Genus Elaeis

Specie Elaeis guineensis.

Source: Reeves and Weihrauch (1979).

While oil palm is recognized for its contribution to economic growth, the rapid

development of palm products has also correspondingly led to environmental pollution.

3

1.2 The Palm Seeds

Palm seeds are reddish, about the size of a large plum and grow in large bunches.

Each fruit is made up of an oily and fleshy outer layer (the pericarp) with a single seed (the

palm kernel) that is also rich in oil. The seeds are used for propagating the plant and are eaten

roasted or boiled. The pulp is pressed to produce palm oil while the kernel is used to produce

palm kernel oil (Rupani et al., 2010).

Fig 1: Fresh fruit bunch (FFB). Fig 2: Palm seeds

One of the seeds was cracked open to show the pulp segment, the kernel shell and the kernel

seed.

1.3 The Palm Oil Production

Palm oil is edible oil derived from the fleshy mesocarp of the fruit of oil palm. It is

one of the most widely consumed plant oil across the world (Rupani et al., 2010). In general,

the palm oil milling process can be categorised into dry and wet (standard) processes. The

wet process of palm oil milling is the most common and typical way of extracting palms

especially in Nigeria (Okwute and Isu, 2007). Despite the fact that the POME can cause

environmental pollution, not much has been done to mitigate this effect. The technology

applied in almost all palm oil mills is based on methods developed in the 1970s and 80s

(Zaini et al., 2010). The major steps in the oil palm processing as reported by Zaini et

al.(2010) are as follows:

4

Threshing: This is the removal of fruits from bunches. The fresh fruit bunches

consists of the fruits that are attached onto the spikelet growing on a main stem. The fruit-

laden spikelet are cut from the bunch stem using axe for manual threshing before separating

the fruits from the spikelets (Zaini et al., 2010).

Sterilisation: Loose fruits are boiled in batches using high temperature wet-heat

treatment. This is carried out in autoclave by steam application at temperature and pressure

ranges of 120-140°C at 3-3.5 bar, for 75 minutes. Boiling prevents fatty acid formation and

assists in fruit stripping as well as prepares the fruit fibre for the next processing step. Boiling

breaks down oil-splitting enzyme and stops hydrolysis and auto-oxidation(Zaini et al., 2010).

Crushing process: In this step the palm fruits are passed through shredder and

pressing machine to separate the oil from the fibre and seeds (Zaini et al., 2010).

Digestion of the Fruit: This process releases the palm oil in the fruit through cracks

in the oil-bearing cells. The digester consists of a steam-heated cylindrical vessel with central

rotating shaft that is filled with several beater arms. The fruit is pounded by the rotary beater

arms at high temperature to reduce the oil viscosity. This destroys the exocarp fruits or the

outer covering and completes the disruption of the oil cell already begun in the sterilisation

process. The digester must be filled to ensure the maximum storage and the effect of the

agitation (Zaini et al., 2010).

Extracting the Palm Oil: There are two distinct methods of extracting oil from the

digested palm fruit: one system uses mechanical presses and is called the “dry” method. The

other called the “wet” method uses hot water to leach out the oil (Zaini et al., 2010).

Kernel Recovery - The residue from the press consists of a mixture of fibres and

palm nuts which are at this stage sorted. The sorted fibres are covered and allowed to be

heated by itsinternal exothermic reactions for about two or three days. The fibres are then

pressed in spindle press to recover second grade (technical) oil that is used normally in soap

making. The nuts are usually dried and sold to other operators who process them into palm

kernel oil (Zaini et al., 2010).

Refining: Refining converts the crude palm oil (CPO) into refined form. The CPO is

processed to segregate fat and obtain refined palm oil (Zaini et al., 2010).

Oil Storage: The palm oil is stored in large steel tanks at 31 to 40°C to keep it in

liquid form during bulk transport. The tank headspace is often flushed with CO2 to prevent

oxidation. Higher temperatures are used during filling and draining of the tanks (Zaini et al.,

2010).

5

Fig 3:Palm oil mill effluent dumpsite

KEY:

a = Fermented POME

b = Fresh POME

a

b

6

Table 1: Summary of Unit Operations in Palm Oil Production (Rupani et al., 2010)

Unit operation Purpose

Fruit fermentation To loosen the fruit base from the spikelets and to allow ripening

processes to abate

Bunch chopping To facilitate manual removal of the fruit

Fruit sorting To remove and sort the fruit from spikelets

Fruit boiling To sterilise the fruit and stop enzymatic spoilage, coagulate protein and

expose microscopic oil cells

Fruit digestion To rupture the oil-bearing cells in order to allow oil flow during

extraction while separating fibres from nuts

Mash pressing To release the fluidal palm oil using applied pressure on ruptured

cellular contents

Oil purification To boil the mixture of oil and water in order to remove the water-

soluble gums and resins in the oil as well as to dry decanted oil by

further heating

Fibre-nut separation To separate the oiled fibres from palm nuts.

Second pressing To recover residual oil for use as soap stock

Nut drying To sun-dry nuts for later cracking

1.4 The Properties of POME

The two main wastes resulting from palm oil production in oil mills are the solid and

liquid wastes (Kathiravale and Ripin, 1997). The solid waste typically consists of palm kernel

shells (PKS), mesocarp fruit fibres (MFF) and empty fruit bunches (EFB). The liquid waste

generated from the extraction of palm oil in wet process comes mainly from the oil room

after the kernel recovery. This liquid waste combined with the wastes from the steriliser

condensate and cooling water is called palm oil mill effluent (POME) (Zaini et al., 2010).

Palm oil production requires input of large quantity of water which is eventually

discharged as waste effluent. POME is an effluent generated from palm oil milling activities

which requires effective treatment before discharge into watercourses (Nwoko and

Ogunyemi, 2010). POME generated from mill operation is thick, brownish,

highlyconcentrated,colloidal and slurry with pH ranging from 4.0 to 4.5 and a temperature of

between 80 and 90°C (Zaini et al., 2010; Alrawi et al., 2012). It contains mainly water (95-

7

96%), suspended solids (2-4%) and oil and grease (0.6-0.7%) (Ahmad et al., 2003). Palm oil

production process does not utilize any chemical and hence POME is considered as a non-

toxic wastewater.

Table 2: Characteristics of POME and its respective standard discharge limit

Parameters Experimental values obtained by some previous

researchers on POME.

Standard

limit (mg/L)

pH 4.7 3.8-4.4 4.24-4.66 5-9

Oil and grease (mg/L) 4,000 4,900-5,700 8,845-10,052 50

Biological oxygen demand

(mg/L)

25000 23,000-26,000 62,500-69,215 100

Chemical oxygen

demand (mg/L)

50,000 42,500-55,700 95,465-112,023 -

Total solids (mg/L) 40,500 - 68,854-75,327 -

Suspended

solids (mg/L)

18,000 16,500-19,500 44,680-47,140 400

Total nitrogen (mg/L) 750 500-700 1,305-1,493 150

Total volatile solids

(mg/L)

34,000 - 4,045-4,335 -

Sources: Department of EnvironmentMalaysian, (1999); Ahmad et al., 2003; Najafpou et al.,

2006; Choorit and Wisarnwan(2007).

1.4.1 Physicochemical Characterisation of POME

1.4.1.1 Dissolved Oxygen (DO)

The dissolved oxygen is a measure of the amount of gaseous oxygen dissolved in an

aqueous solution. Analysis of DO is a key test in water pollution. The DO levels in POME

depend on the physical, chemical and biochemical activities in POME. Adequate DO is

necessary for good quality of water. Oxygen is an essential element to all forms of life. The

DO concentrations ought not to exceed 110% otherwise; it may be harmful to aquatic life. As

DO levels in water drop below 5.0 mg/L, aquatic life is put under stress; the lower the

concentration of DO, the greater the stress. Death usually occurs at concentrations less than 2

mg/L. The World Health Organization (WHO) suggested the standard of DO greater than

5mg/L for river water monitoring (Sehar et al., 2011).

8

1.4.1.2 Biochemical Oxygen Demand (BOD)

The standard, five-day BOD (BOD) value is commonly used to determine the amount

of organic pollution in water and wastewater. Determination of BOD involves measuring the

oxygen demand of both the organic matter and organism in the POME. BOD is the amount of

dissolved oxygen needed by aerobic biological organisms in a body of water to break down

organic material present in a given water sample at certain temperature over a specific period

of time (Okwute and Isu, 2007). It is an empirical test that determines the relative oxygen

requirements of wastewater, effluents and polluted water. BOD tests measure the molecular

oxygen utilised during a specified duration incubation for the biochemical degradation of

organic materials (carbonaceous demand) and the oxygen used to oxidise inorganic material

such as ferrous iron and sulfides (Sehar et al., 2011).

1.4.1.3 Chemical Oxygen demand (COD)

The chemical oxygen demand (COD) is used to measure the oxygen equivalent of the

organic material in wastewater. It is a useful measure of water quality. In most cases

applications of COD determine the amount of organic pollutants found in water (Sehar et al.,

2011). The COD test is commonly used to indirectly measure the amount of organic

compounds in water. The COD is the amount of oxygen required to chemically oxidize

organic water contaminants to inorganic end products (Okwute and Isu, 2007). Most

applications of COD determine the amount of organic pollutants found in surface water. COD

is often measured as a rapid indicator of organic pollution in water (Sehar et al., 2011).

1.4.1.4 Total Dissolved Solids (TS)

The total solids represent all solids in a water sample. They include the total

suspended solids, total dissolved solids, and volatile suspended solids. The range of 37900 -

45 000 mg/L has been reported by Wood et al. (1979); Wong et al. (2009) and MPOB

(2004). TS is a measure of the amount of filterable solids in a water sample (Sehar et al.,

2011).

1.4.1.5 Total Suspended Solids (SS)

These are amounts of filterable solids in a water sample. POME samples are filtered

through a glass fibre filter. The filters are dried and weighed to determine the amount of total

suspended solids in mg/L of sample. Suspended solid of POME were reported to be 18,000

mg/L by Ahmad et al. (2003) and MPOB (2004). The higher suspended solid was 25,800

mg/L and was recorded by Wu et al., 2007

9

1.4.1.6 Volatile Suspended Solid (TVS)

Volatile solids are those solids lost on ignition (heating at 550 0C). They are useful in

application for treatment plant operator because they give a rough approximation of the

amount of organic matter present in the solid fraction of wastewater, activated sludge and

industrial wastes. Wood et al. (1979) and Wong et al. (2009) reported VSS range of 27300

mg/L to 30150 mg/L.

1.4.1.7 Oil and Grease (O and G)

Oil and grease have poor solubility in water. Thus, oil and grease content of industrial

wastes are important consideration in handling and treatment of the material for disposal

(Salihu et al., 2011a). The concentration of oil in effluents from different industrial sources

can be as high as 40,000mg/L (Arcadio and Gregoria, 2003). Unlike free or floating oil

spilled in the sea, lakes or rivers, most of the industrial wastewaters contain oil-in-water

emulsions among other basic contaminants. Emulsified oil in wastewater can lead to severe

problems in different treatment stages. Oil in wastewaters has to be removed in order to: (1)

prevent interfaces in water treatment units (2) reduce fouling in process equipment (3) avoid

problems in biological treatment stages and (4) comply with water discharge requirements

(Arcadio and Gregoria, 2003).

1.5 Heavy Metals

The term heavy metal refers to any metallic chemical element that has a relatively

high density and is toxic or poisonous at low concentrations (Kızılkaya et al., 2004).

Examples of heavy metals include mercury (Hg), cadmium (Cd), arsenic (As), chromium

(Cr), thallium (Tl), and lead (Pb) (Lebedeva et al., 1995). The use of metals by humans was

and is still accompanied by increasing inputs of metals into soils through different types of

wastes (Welp, 1999). The major sources of chromium include the metal finishing industry,

petroleum refining, leather tanning, iron and steel industries, production of inorganic

chemicals, textile manufacturing and pulp production. Because metals persist in soils and

their leaching is a very slow process, they tend to accumulate in the soils (Irha et al., 2003).

Chromium is one of the heavy metals and has oxidation states ranging from chromium (III) to

chromium (VI).Chromium compounds are stable in the trivalent state and occur in nature in

this state in ores such as ferrochromite, while chromium (VI) is usually produced from

anthropogenic sources (Cervantes et al., 2001). Hexavalent chromium compounds have been

used in a wide variety of commercial processes (Turick et al., 1996). Upon the reduction of

10

chromium (VI) to chromium (III), the toxic effects are significantly decreased in humans,

animals and plants because of a decrease in the solubility and bioavailability of chromium

(III) (Turick et al., 1996). The reduction of the highly toxic and mobile Cr (VI) to the less

toxic and less mobile Cr (III) is likely to be useful in the remediation of contaminated waters

and soils. This problem has stimulated interest in microorganisms as alternatives to

conventional methods due to their eco-friendly nature. Cr (III) is transformed to Cr (VI)

mainly inside root cells but also in the aerial part of plant (Cervantes et al., 2001). Roots

accumulate 10-100 times more Cr than shoots and other tissues. As a consequence, inhibition

of growth, photosynthesis and respiration processes in plants and microorganisms are

observed (Cervantes et al., 2001). Reduction of soluble Cr (VI) to insoluble Cr (III) occurs

only within the surface layer of aggregates with higher available organic carbon and higher

microbial respiration (Tokunaga et al., 2003). Thus, spatially resolved chemical and

microbiological measurements are necessary within anaerobic soil aggregates to characterise

and predict the fate of chromium contamination (Tokunaga et al., 2003). Heavy metals can

enter a water supply by industrial and consumer waste or even from acidic rain, breaking

down soils and releasing heavy metals into streams, lakes, rivers and groundwater (Zheng et

al., 1999). The impacts of elevated heavy metal levels on the size and activity of natural soil

microbial communities have been well documented. Field studies of metal -contaminated

soils have shown that elevated metal loadings can result in decreased microbial community

size (Jordan and LeChevalier, 1975; Brookes and McGrath, 1984; Chander and Brookes,

1991; Konopka et al., 1999) in organic matter mineralisation (Chander and Brookes, 1991)

and leaf litter decomposition (Strojan, 1978).

1.5.1 Impact of Heavy Metals on the Soil Enzymes

By taking part and playing an important role in chemical changes of carbon, nitrogen,

phosphorus and sulphur compounds, soil enzymes can serve as a tool for determining the

biochemical soil properties (Dick and Tabatabai, 1992). For this purpose, activity of

dehydrogenases is most commonly assayed as it is usually positively correlated with the

volume of yields which in turn may indirectly indicate, however, that the activity of those

enzymes is related to soil fertility (Dick, 1997). The activity of other soil enzymes such as

catalase, lipase, urease or phosphatase, can also be helpful because they are as sensitive as

dehydrogenases in indicating processes occurring in soil. Soil enzyme activities are

considered to be sensitive to pollution and have been proposed as indicators of soil

degradation (Trasar-Cepedaet al., 2007). Catalase (hydrogen peroxide oxidoreductase, EC

11

1.11.1.6) is an intracellular enzyme found in all aerobic bacteria and most facultative

anaerobes but absent in obligate anaerobes (Trasar-Cepedaet al., 2007 and Shiyinet al.,

2004).

This enzyme can be found in all aerobic microorganisms, plant and animal cells (Alef

and Nannipieri, 1995). Although, it was one of the first isolated and purified enzymes, its

physiological function and regulation are still poorly understood. Catalase activity may be

related to the metabolic activities of aerobic organisms and has been used as an indicator of

soil fertility (Gianfreda and Bollag, 1996; Trasar-Cepedaet al., 2007: Shiyinet al., 2004).

Catalase activity is very stable in soil and shows a significant correlation with the content of

organic carbon decreasing with soil depth (Alef and Nannipieri, 1995). Heavy metals,

however, are regarded as inhibitors of enzymatic and microbiological activity of soil. This is

because if added to soil (whether on purpose or by accident) it causes a quantitative and

qualitative change in the composition of micro flora and enzymatic activities. Chromium (VI)

has a harmful effect on soil microorganisms by depressing their biological activity

(Wyszkowskaet al., 2001).

Contamination with heavy metals, fertilisation, application of plant protection

chemicals and soil cultivation all modify the physicochemical characteristics of the soil and

change its biological activity (Milosevic et al., 1997). Of special importance are the heavy

metals which may stimulate the activity of the soil enzymes if present in small amounts but

will act as inhibitors if found in high concentrations (Landiet al., 2000). The effect of heavy

metals on the biological activity of the soil depends on the physicochemical properties of the

soil particularly on its humus content. On the other hand, it is also dependent on the

concentrations as well as the kinds of pollutants or enzymes involved (Landiet al., 2000).

Cadmium represents a group of heavy metals causing the most severe changes in the

biological properties of soils (Akmalet al., 2005). Many researchers have also suggested

significant negative correlation between the concentration of cadmium in the soil and the

activity of dehydrogenases and phosphatases (Doelman and Haanstra, 1984). Soil enzymatic

activity is one of the factors which allow for the prediction of the volume of yields.

1.6.0 Socio-Economic Importance of POME

Oil palm is a valuable economic crop and provides a major source of employment.

Active palm oil milling activities have led to the release of palm mill pollutants such as

POME in the environment. Land application of palm oil mill effluent (POME) is one of the

disposal alternatives. Discharging the POME on the land results in clogging and water

12

logging of the soil and kills the vegetation in contact with it (Rupani et al., 2010). The direct

discharge of palm oil mill effluent (POME) wastewater causes serious environmental

pollution due to its high chemical oxygen demand (COD) and biochemical oxygen demand

(BOD). BOD of above 25,000mg/L makes it objectionable to aquatic life when introduced in

waterways (Abdurahman et al., 2011). POME is a non-toxic waste, as no chemical is added

during the oil extraction process, but will pose environmental issues due to the large oxygen

depleting capability in aquatic system due to organic and nutrient contents (Zaini et al.,

2010). Increased damage to the environment overtime resulting from increasing milling

activities has incited a need for environmental protection through efficient effluent

monitoring (treatment and discharge), resulting in more environmental-friendly practices. As

reported by (Okwute and Isu, 2007), the raw POME has an extremely high content of

degradable organic matter which in part is due to the presence of unrecovered palm oil. Thus,

POME should be treated before discharge to avoid serious environmental pollution. Raw

POME has BOD values averaging around 25,000 mg/L, making it about 100 times more

polluting than domestic sewage (Okwute and Isu, 2007).

Residual oil is one of the key ingredients of POME which influences the high values

of COD and BOD. The palm oil present in the effluent may float to the surface of the water

body and form a wide-spread film which can efficiently cut-off and avert atmospheric oxygen

from dissolving in its waters. Furthermore, when the organic load far exceeds its waste

assimilation capacity, the available oxygen in the water body is rapidly consumed as a result

of the natural biochemical processes that take place. The water body may become completely

devoid of dissolved oxygen. This will lead to anaerobic conditions in which hydrogen

sulphide and other malodorous gases are generated and released to the environment resulting

in objectionable odours. Additional damaging effects include the decline and eventual

destruction of aquatic life and deterioration in the riverine ecosystem. Hence the challenge of

converting POME into an environmental-friendly waste requires sound and effective

treatment as well as efficient disposal (Sethupathi, 2004).

1.6.1 Pre-treatment

Initial pre-treatment involves the use of flocculation, solvent extraction, adsorption

and membrane separation processes to remove the suspended solids, residual oil and grease

(Abdul Latif et al.,2003a). Abdul Latif et al. (2003b) showed the superiority of membrane

separation using ultrafiltration and reverse osmosis over the use of coagulation and activated

carbon adsorption for POME treatment. The latter had removal efficiencies of 97.9%

13

turbidity, 56% COD and 71% BOD while the former yielded a higher removal efficiencies of

100% turbidity, 98.8% COD and 99.4% BOD. Although, anaerobic and facultative ponding

systems are widely employed for POME treatment (Ma, 1999), however, in order to prevent

the escape of methane gas generated, these systems are coupled with closed digesters so that

the generated methane can be used as an auxiliary fuel in the boilers of palm oil mill

industries (Vijayaraghavan et al., 2007).

1.6.2 Biological Treatment Technology

The biological treatment relies greatly on a mixed population of active

microorganisms which utilises the organic substances polluting the water as nutrients and

finally breakdown of the organic matter into simple end-product gases such as methane,

carbon dioxide, hydrogen sulphide and water (Sethupathi, 2004).

1.6.3 Land Application System

The anaerobically digested POME contains high concentrations of plant nutrients.

The application of the effluent to the cropland, not only provides nutrient and water to the

vegetation, but also serves as an alternative disposal of the effluent. This has resulted in

substantial saving in fertilizer bills and increased income due to higher crop yield

(Sethupathi, 2004).

1.6.4 Evaporation Technology

Evaporation is one of the most widely used unit operation in water regeneration in a

wide range of processing applications. POME is made up of about 95 - 96% water. Thus, by

this evaporation technology, the water could be recovered and the residual solid concentrated

for advanced utilisation (Sethupathi, 2004).

1.7 General Uses of POME

While POME has not quite had a clean slate on environmental friendliness, there is a

world of use to which it could be put. There has been recent research and subsequent

discovery on various uses POME could be put to by subjecting it to specific treatment

(Okwute ana Isu, 2007). POME is a biphasic product which is considered as a waste but at

the same time, utilized as a raw material in other processes. Many processing technologies for

converting POME into value added products have been developed. These products as

reported by Salihu and Alam (2012) include:

14

1.7.1 Biogas (Biohydrogen)

Biohydrogen production attracts attention of researchers as it is less energy-intensive

and can be coupled with wastewater treatment processes using dark-and photo-fermentation

techniques. Thus, hydrogen is characterised with a high energy yield of 122 KJ/g which is

2.75 times higher than the hydrocarbon fuels and the only end-product is water (Chong et

al.,2009). These characteristics make it a promising alternative fuel. The recent biological

approach to producing hydrogen is to convert agro-industrial residues into hydrogen-rich gas

through anaerobic processes by potential bacterial strains (O’Thonget al., 2007). As a result

many researchers have recognized the potential of harnessing hydrogen from POME (Salihu

et al., 2011a).

1.7.2 Bioplastic Formation

Polyhydroxyalkanoates (PHAs) are thermoplastics that have excellent mechanical

properties similar to those of polypropylene (Purushothamanet al., 2001; Mumtazet al.,

2010). They are endowed with other additional advantages such as biodegradability,

biocompatibility and production are mainly from renewable resources (Purushothaman

etal.,2001; Mumtaz et al.,2010). The PHAs are intracellular products generated by some

bacteria species under nutrient limiting conditions especially in a medium rich in carbon with

little or no nitrogen or phosphorus present (Hassan et al., 1996). Based on this, high organic

content of POME makes it a good substrate for bioplastic production. Hassan et al.(1996)

developed a two-stage process for the production of PHA using POME. The initial stage

involves the production of organic acids (acetic and propionic acids) by anaerobic treatment

of POME followed by the conversion of the produced organic acids to PHA using a

phototrophic bacterium, Rhodobactersphaeroides. Thus, renewable agricultural residues

coupled with highly producing microorganisms are the pre-requisites for making the

production of PHA economically viable since the types and compositions of the substrates

determine the final polymer properties of the bioplastics (Mumtaz et al.,2010). For this

reason the use of POME as an inexpensive carbon source to produce PHAs may lead to some

significant economic advantages.This is because substantial amount (40%) of total operating

costs of PHA production could be linked to the raw materials and more than 70% of this cost

is attributed to the carbon source. (Alias and Tan, 2005) isolated a bacteria species from

POME which has the ability to produce poly-(3) hydroxybutyrate. The bacterium was

tentatively identified as Burkholderia cepacia. Generally, wild type microorganisms do not

produce a high yield of PHA to be considered for use at an industrial scale but their ability to

15

synthesise PHA would be very important as it may lead to genetic modification of the

microorganisms (Mumtaz et al.,2010).

1.7.3 Bioethanol production

Large scale production of bioethanol using lignocellulosic biomass and agro-

industrial residues could reduce the reliance on fossil sources and curb some levels of

environmental pollution (Alametal., 2008b). This is because combustion of ethanol does not

contribute to global warming unlike fossil fuel that engenders atmospheric build-up of carbon

dioxide in the atmosphere. Several renewable residues have been used in the production of

ethanol; these include waste paper, domestic refuse and lignocellulosic biomass (corn stover,

crop straw, sugar cane bagasse). The production of bioethanol using POME as the main

substrate in the presence of Saccharomyces cerevisiae under controlled laboratory condition

had an optimum yield of 16% (Cheng et al., 2007). Based on this, the utilisation of POME for

bioethanol production offers a simple and effective treatment of this effluent produced by

palm oil mills (Alam et al., 2008b). Thus, the oil palm-based residues could provide an

impetus for sustainable generation of bioethanol as well as other value added products

(Cheng et al.,2007).

1.7.4 Compost and Biofertilizer

Composting is a process whereby complex organic residues of plant and animal

origins are converted into manure or biofertilizer through the activities of several microbial

systems such as bacteria and fungi (Singh et al.,2010). This process helps in stabilizing

various types of industrial wastes and sludge (sludge from pulp and paper, sugar,

oleochemical, oil mills) and aids in reducing the volume/weight of the produced sludge (Abd-

Rahman et al., 2003). Since POME is of biological origin, its composting can be a good

alternative for its sustainable management though, oil palm empty fruit bunches (EFB) have

been used to produce organic fertilizer and the process led to their bulk volume reduction by

about 50% (Chavalparit et al.,2006). (Baharuddin et al., 2009) studied the physicochemical

changes during co-composting of EFB with partially treated POME on a pilot scale and

reported that EFB could be used with POME for composting process to produce acceptable

quality compost that can be applied to palm oil plantations as biofertilizer and for soil

conditioning. Therefore, the production of compost using POME could serve as a valorisation

strategy that can reduce the volume of this waste with easy land application.

16

1.7.5 Medium for Isolation and Growth of Microorganisms

Numerous microorganisms can invade and grow in POME, breaking down complex

molecules into simple ones. The high organic matter in POME has contributed in the

proliferation of aerobic microorganisms. The growth of Yarrowia lipolytica NCIM 3589 in

raw POME led to a substantial reduction in COD within 48h and the acidic pH of POME

became alkaline due to the utilisation of fatty acids by the organism (Oswal et

al.,2002).Rashid et al. (2009) isolated and purified some filamentous fungi from POME into

three different genera namely: Penicillium, Rhizopus and Basidiomycetes. Also, some

bacterial species isolated from POME ponds at a palm oil processing factory, showed high

potential for PHAs production based on initial staining screening experiments using sudan

black B plates (Redzwan et al., 1997). Rhizopus oryzae and Rhizopus rhizopodiformis were

thermophilic fungi isolated from POME. These fungi showed high extracellular and

intracellular lipase activity at 50°C (Razak et al., 1997).

1.7.6 Citric Acid Production

In an effort to utilise the effluent discharge of the palm oil industries, citric acid was

been produced using POME as a raw material (Jamal et al., 2007). Industrial production of

citric acid depends on fungal fermentation using for example, Aspergillus niger in the

presence of glucose (or sucrose) as the main substrate using liquid and solid state

fermentation techniques (Grewal and Karla, 1995). However, several cheaper substrates have

been used for citric acid production by Aspergillus niger. These include inulin (Drysdale and

McKay, 1995), date fruit syrup (Roukas and Kotzekidou, 1997), sugarcane molasses (Gupta,

1994) among others. All these substrates have lesser yields when compared to POME. (Alam

et al., 2008a) obtained the citric acid yield of 5.2g/L from POME using a seven day

fermentation period. Citric acid is used extensively in various industrial processes especially

as acidulant, stabiliser, flavour enhancer, preservative, antioxidant, emulsifier and chelating

agent (Jamal et al.,2007). Thus, annual production of citric acid is estimated to be about half

a million tonnes based on the fermentation of conventionally used raw materials (glucose and

sucrose). Based on the extensive use of citric acid, several strategies have been put in place

for its efficient production using agricultural residues. Efforts to achieve this goal by using

POME as a new substrate proved effective (Jamal et al.,2007; Alam et al.,2008a).

17

1.7.7 Carotenoid Extraction

Carotene is a provitamin, which is converted in the body to a vitamin. The β-carotene

is a precursor of vitamin A that plays major roles in the maintenance of strong bones, healthy

skin, teeth and hair. It can also be utilised in the production of vitamin E (tocopherol). It is

also known as a potent antioxidant, protecting the cells against effects of free radicals in the

body (Ahmad etal., 2008). The amount of carotene extracted from POME is closely related to

that obtained from the crude palm oil which carotene concentration ranges between 400 and

3500ppm. Carotenoids have been extensively used in food, cosmetic and pharmaceutical

industries and as such their extraction and recovery from POME could be important (Jamal et

al.,2007).

1.7.8 Enzyme Production

Hydrolytic enzymes are among the most important groups of industrial enzymes

because of their potential for biotechnological use. According to technical market research

report, enzymes for industrial applications were expected to increase to over $2.7 billion by

the year 2012 with an average annual growth rate of 4%. The total industrial enzyme market

in 2009 was reported to be $2.4 billion (Hassan et al., 2006).

1.8.0 Soil Fertility

The term, soil fertility refers to the quality of the soil which sustains plants and helps

them to grow well. It is a very significant quality of the soil. It depends on many things such

as soil texture, pH, nutrients, organic matter, water-holding capacity, microorganism,

structure, microclimate, irrigation facility, land fragmentations, erosion, agricultural system

and practices, diseases and insects, consumption of nutrients by crops, conversion of nutrients

into inconsumable by plants and gases and leaching of nutrients (Sojka and Upchurch,1999).

The word soil is derived from a Latin word “solum” meaning floor. Soil is the upper thin

layer of earth’s surface derived from the weathering of rocks and minerals. The term soil is a

complex mixture of eroded rocks, minerals, nutrients, decaying organic matter, water, air and

billions of living organisms which mostly are microscopic decomposers. Although, soil is a

potentially renewable resource, it is produced very slowly by the weathering of rocks,

deposits of sediment by erosion and decomposition of organic matter in dead organisms

(Romiget al., 1996 and Sarrantonioet al., 1996). It is the heart of terrestrial ecological system,

so understanding of the soil system is the key to the success and environmental harmony of

any human use of the land. Soil chemical analysis is made to assess the available amounts of

18

major nutrients, nitrogen, phosphorus and potassium and to assess a few other determinations

which are correlated to soil fertility such as soil texture, soil reaction (pH) and salinity (Doran

and Parkin, 1994).

2.8.1 Soil pH

The soil pH is a basic property of the soil that affects many chemicals and biological

activity of the soil. The degree of acidity and alkalinity of the soil is known as the soil pH.

The soil pH is determined by the hydrogen ion concentration in the soil solution. An acidic

soil has more hydrogen than hydroxyl ions, while basic or alkaline soil has more hydroxyl

than hydrogen ions. It has been reported that when raw POME is discharged, the pH is acidic

and gradually increase to alkaline as biodegradation takes place (Hemming, 1977). Many of

the soils of the world is affected by excess acidity, a problem exacerbated by heavy

fertilisation with certain nutrients and by acid rain (Paul and Clarke, 1989). The soil pH

increases with increasing soil depth (Skyllberg, 1993). The soil acidity is one of the principal

factors affecting nutrient availability to plants. Therefore, the availability of plant nutrients in

the soil is affected by the pH of soil. Most crops grow at a pH between 6.5 and 7.5 (Hajek et.

al., 1990). The major nutrients (nitrogen, potassium and phosphorus) cannot effectively

promote high crop yields if the soil pH is not suitable. At pH values less than 5.5, toxic levels

of these elements may be present in the soil. Soils have a near-neutral pH, unless the soil is

used for growing acid-loving crops (Paul and Clarke, 1989).

Table 3: Classification of soil based on the pH range

pH Ranges Soil Type

<4.0 Very strongly acidic

4.0 – 5.5 Strongly acidic

5.5 – 6.0 Medium acidic

6.0 – 6.5 Slightly acidic

6.5 – 7.0 Very slightly acidic

7.0 – 7.5 Very slightly alkaline

7.5 – 8.0 Slightly alkaline

8.0 – 8.5 Medium alkaline

8.5 – 10.0 Strongly alkaline

> 10 Very strongly alkaline

Source: (Hajek et al., 1990).

19

1.8.2 Soil Enzymes

The biological oxidation of soil organic compounds is generally a dehydrogenation

process carried out by specific dehydrogenases involved in the oxidative energy transfer of

microbial cells (Welp, 1999). This activity is a measure of the microbial metabolism and thus

of the oxidative microbial activity in the soil. A good correlation has been established

between microbial biomass and soil dehydrogenase activity (Chander et al., 1977). The

determination of the soil dehydrogenase activity is generally done by adding alternative

electron acceptors to soil samples. Water-soluble tetrazolium salts are the preferred oxidants

because they form water-insoluble coloured formazans which can be measured

spectrophotometrically (Kandeler et al., 1996). Microorganisms are the main source of

enzymes in the soils and thus the composition of the soil microbial communities strongly

affects the potential of a soil for enzyme-mediated substrate catalysis (Kandeler et al., 1996).

Specifically, the assessment of the activities of hydrolases can provide information on the

status of the key reactions that participate in rate limiting steps of the decomposition of

organic matter and transformation of nutrients in the soil (Chander et al., 1977). Thus, the

knowledge of activities of several soil enzymes can provide information on the soil

degradation potential (Tra´sar-Cepeda et al., 2007). The assessment of the soil enzyme is

simple and requires low labour costs compared to other biochemical analysis (Ndiaye et al.,

2000) and the results are correlated to other soil properties (Klose et al., 1999; Moore et al.,

2000; Ndiaye et al., 2000; Tra´sar-Cepeda et al., 2007). Furthermore, it has been reported that

any change in soil management and land use is reflected in the activities of soil enzyme and

that they can anticipate changes in soil quality before they are detected by other soil analyses

(Ndiaye et al., 2000). The activities of soil enzymes are very sensitive to both natural and

anthropogenic disturbances, and show a quick response to the induced changes (Quilchano

and Maranon, 2002). Therefore, enzyme activities can be considered effective indicators of

soil quality changes resulting from environmental stress or management practices (Quilchano

and Maranon, 2002). The studies of the activities of the enzymes provide information on the

biochemical reactions occurring in the soil (Masciandro et al., 2000). However, the

relationship between an individual biochemical property and the total microbial activity is not

always obvious especially in the case of complex systems like soils, where both

microorganisms and processes involved in the degradation of organic compounds are highly

diverse (Quilchano and Maranon, 2002).

20

1.8.2.1 Soil Dehydrogenases

The dehydrogenases (EC 1.1.1.1) are enzymes which catalyse the removal of

hydrogen atom from different metabolites (Margesin et al., 2000). Active dehydrogenases are

considered to exist in the soil as an integral part of intact cells. They conduct a wide range of

oxidative activities that are responsible for the degradation of the soil organic matter

(Margesin et al., 2000). The soil dehydrogenase activity can reflect changes in the respiratory

activity of a given population size in response to changes in the soil environment (Schinner et

al., 1996). The soil quality and its degradation depend on a large number of physical,

chemical, microbiological and biochemical properties of the soil (Margesin et al., 2000). The

microbiological and biochemical properties are the most sensitive since they respond rapidly

to changes. The microbiological activity of a soil directly influences ecosystem stability and

fertility and it is widely accepted that a good level of microbiological activity is essential for

maintaining the soil quality. The soil microbiological activities play a key role in the soil

nutrient cycling; its activity is essential in both the mineralisation and transformation of

organic matters in the soil ecosystem (Moreno et al., 2002). These soil enzymes play a

fundamental role in establishing biogeochemical cycles and facilitate the development of

plant cover (Dick and Tabatabai, 1992). It is an important aspect of the below-ground

processes and give insight into the relative changes in below-ground system functioning as a

plant community develops over time. The enzyme activity in the soil results from the activity

of the accumulated enzymes and the enzymatic activity of the proliferating microorganisms

(Dick and Tabatabai, 1992). The study of soil enzymes gives information about the release of

nutrients into the soil by means of organic matter degradation and microbial activity as well

as indicators of ecological change (Lenhard, 1956). The analyses of soil enzymes help to

establish correlation with the soil fertilization, microbial activity, biochemical cycling of

various elements in soil, degree of pollution (heavy metals) and assess the succession stage of

an ecosystem (Dick et al., 1996). So, the measurement of the enzyme activity in degraded

soils is useful in examining the impacts of environmental change or management on the soil

enzyme activity. Several works have been reported on the potential use of enzyme activity as

an index of soil productivity or microbial activity (Dick, 1984; Visser and Parkinson, 1992).

One of the general criteria used to determine the microbial activity and biomass in soil is soil

dehydrogenase activity (Waksman, 1992).

21

1.8.2.2 Soil Catalase

The enzyme, catalase (hydrogen peroxide oxidoreductase, EC 1.11.1.6) is an

intracellular enzyme found in all aerobic bacteria and most facultative anaerobes, but absent

in obligate anaerobes (Trasar-Cepeda et al., 2007; Shiyin et al., 2004). It is well known that

the products of oxygen reduction, such as hydrogen peroxide, superoxide radical, and

hydroxyl radical, can be highly toxic for cells and might damage cellular macromolecules.

Catalase can split hydrogen peroxide into molecular oxygen and water and thus prevent cells

from damage by reactive oxygen species (Shiyin et al., 2004). Although it was one of the first

isolated and purified enzymes, its physiological function and regulation are still poorly

understood (Shiyin et al., 2004). Catalase activity may be related to the metabolic activity of

aerobic organisms and has been used as an indicator of soil fertility (Gianfreda and Bollag,

1996; Shiyin et al., 2004; Trasar-Cepeda et al., 2007). Catalase activity is very stable in soil

and shows a significant correlation with the organic carbon content in soil and decrease with

soil depth (Alef and Nannipieri, 1995).

1.8.2.3 Soil Lipase

The lipases (EC 3.1.1.3) are enzymes which have the ability to hydrolyse

triacylglycerols to release free fatty acids and glycerol. Lipases constitute a major group of

biocatalysts that have immense biotechnological applications (Margesin et al., 1999). The

soil lipase activity is a valuable indicator of oil biodegradation in the soils. The soil lipase

activity is a valuable tool to monitor biodegradation of hydrocarbon chains of oily

contaminated soil. The increase in lipase activity with increasing oil and grease demonstrates

the induction of this enzyme activity by the contamination. Lipase degrades lipids derived

from a large variety of microorganisms, animals and plants. The research under taken in this

area is likely to progress the knowledge in the bioremediation of oil spill. Lipases are

ubiquitous enzymes which catalyze the hydrolysis of triacylglycerols to glycerol and free –

fatty acids (Margesin et al., 1999)

1.8.2.4 General Applications and Importance of the Soil Enzymes

The general applications and importance of soil enzymes are as followings:

• Release of nutrients into the soil by means of organic matter degradation

• Identification of the soils

• Identification of the microbial activity

• Serve as sensitive indicators of ecological change

22

• Correlation with the soil fertility

• Correlation with the microbial activity

• Correlation with the biochemical cycling of various elements in the soil (C, N, S)

• Show the degree of pollution (heavy metals, sulphur iv oxide (SO4))

• To assess the succession stages of an ecosystem

• Forensic purposes

• Rapid degradation of pesticides

• Disease studies

Sources: (Margesin et al., 1999).

1.9 Cation Exchange Capacity (CEC)

This is the maximum quantity of total cations of any class that the soil is capable of

holding at a given pH value available for exchange with the soil solution (Thomas, 1982).

The CEC of the soil is based on the percentage (%) and type of clay mineral (smectites,

kaolinites, and sesquioxides) as well as the percentage organic matter. It is used as a measure

of soil fertility, nutrient retention capacity, and the capacity to protect groundwater from

cation contamination (Rhoades, 1982). The negatively charged surfaces of these organic

matters have the cations of calcium, magnesium, potassium, ammonium and sodium adsorbed

onto it. These cations are readily exchanged with similar cations in the soil solution. In most

calcareous soils, the CEC will range between 5-35 milliequivalents per 100 grams of soil

(meq/100g). Example, it takes one milliequivalent of calcium ions (20 mg of calcium) to

displace about one milliequivalent of sodium ions (23mg of sodium). Low percentage organic

matter and coarser texture soils have lower CECs and higher percentage organic matter and

finer texture soils have higher CECs. This electric charge is critical to the supply of nutrients

to the plants because many nutrients exist as cations (Mg, K, Ca). In general, soils with large

quantities of negative charge are more fertile because they retain more cations. The main

cations associated with CEC in the soil are the exchangeable cations: calcium (Ca2+),

magnesium (Mg2+), sodium (Na+) and potassium (K+) (Thomas, 1982)

1.9.1 Exchangeable Potassium (K+)

Potassium deficiency is most commonly associated with sandy soils. The optimal

level will vary with the crop type and yield and the soil type. It is an immobile nutrient like

phosphorus and its levels change slowly. Potassium typically accounts for 2-8 % of the cation

exchange capacity (Brady and Nyle, 1984).

23

1.9.2 Exchangeable Calcium (Ca2+)

Calcium deficiency is rare in most calcareous soil where the soil pH is adequate. The

need for calcium as a nutrient element is directly related with the soil pH. Its level in the soil

can exceed 4,000 ppm without evidence of excess in the plant. Calcium typically accounts for

70-85% of the cation exchange capacity (Havlinet al., 2005).

1.9.3 Exchangeable Magnesium (Mg2+)

Most soils contain high levels of magnesium. Therefore, yield response to magnesium

fertilizer application is uncommon with the exception on sandy soils that have lowcation

exchange capacityand organic matter (OM). Magnesium typically accounts for 10-20% of the

cation exchange capacity (Brady and Nyle, 1984). Magnesium deficiency is more common

than calcium. Some of the total soil magnesium is found in non-exchangeable form. Thus, the

exchangeable magnesium level changes slowly with time because of equilibrium with

minerals (Sumner and Miller, 1996).

1.9.4 Exchangeable Sodium (Na+)

This element is retained in the soil as an easily exchangeable cation which at elevated

levels can detrimentally influence the soil solution pH, nutrient balances, soil infiltration and

permeability, and soil solution salinity levels (Sumner and Miller, 1996). Sodium is not

considered an essential plant nutrient and consequently, a very low level is desired. Soil

amendments such as gypsum and elemental sulfur can be used to reduce the amount of

exchangeable sodium (Brady and Nyle, 1984). Sodium typically accounts for 3-10% of the

CEC (FAO, 1998).

1.9.5 Soil Organic Matter

The soil organic matter (SOM) is the organic component of the soil consisting of three

primary parts including small (fresh) plant residues and small living soil organisms,

decomposing (active) organic matter and stable organic matter (humus) (Palm et al., 2001).

The soil organic matter serves as a reservoir of nutrients for crops, provides soil aggregation,

increases nutrient exchange, retains moisture, reduces compaction, reduces surface crusting

and increases water infiltration into the soil (De Ridder and Van, 1990). Plant residues on the

soil surface such as leaves, manure, or crop residues are not considered as SOM and are

usually removed from soil samples by sieving through a 2 mm wire mesh before analysis.

The soil organic matter content can be estimated in the field and tested in the laboratory to

24

provide estimates for nitrogen, phosphorus and sulfur mineralised available for crop

production and adjust fertilizer recommendations (Vanlauweet al., 2005).

The inherent factors affecting soil organic matter such as climate and soil texture

cannot be changed. Climatic conditions such as rainfall, temperature, moisture and soil

aeration (oxygen levels) affect the rate of organic matter decomposition (Palm et al., 1997).

Organic matter decomposes faster in climates that are warm and humid and slower in cool

and dry climates. Organic matter also decomposes faster when the soil is well aerated (higher

oxygen levels) and much slower on saturated wet soils (Vanlauwe et al., 2005). The soils

formed under forests usually have comparably low organic-matter levels because of two main

factors listed below:

• Trees produce a much smaller root mass per acre than grass plants

• Trees do not die back annually and decompose every year. Instead, much of the

organic material in a forest is tied up in the wood of the trees rather than being

returned to the soil annually (Vanlauwe et al., 2005).

1.9.5.1 Functions of the Soil Organic Matter (SOM)

Nutrient Supply: Upon decomposition, nutrients are released in a plant-available

form while maintaining the current levels. Each percent of SOM in the top 6 inches (15.2 cm)

of the medium textured soil (silt and loam soils with a bulk density of 1.2) releases about 10-

20 pounds of nitrogen, 1 to 2 pounds of phosphorus and 0.4 to 0.8 pound of sulfur per acre

annually (Vanlauwe et al., 2005).

Water-Holding Capacity: Organic matter behaves somewhat like a sponge. It has

the ability to absorb and hold up to 90 percent of its weight in water. Another great advantage

of organic matter is that it releases nearly the whole water it holds for use by the plants. In

contrast, clay holds great quantity of water but much of it is unavailable to the plants

(Vanlauwe et al., 2005).

Soil Aggregation: Organic matter improves soil aggregation, which improves soil

structure. With better soil structure, water infiltration through the soil improves, which

improves soil’s ability to take up and hold water (Vanlauwe et al., 2005).

Erosion Prevention: Erosion is reduced with increased organic matter due to the

increased water infiltration and stable soil aggregates.The term steady state is when the rate

of organic matter addition from crop residues, roots and manure or other organic materials

equals the rate of decomposition (Franzluebbers, 2002). If the rate of organic matter addition

25

is less than the rate of decomposition, SOM will decline and conversely, if the rate of organic

matter addition is greater than the rate of decomposition, SOM will increase (Cambardella

and Elliott,1993). Due to the difficulties in measuring SOM directly, it is substituted with the

measurement of soil organic carbon (Baldock and Skjemstad, 1999). A convenient way to

calculate SOM is by multiplying the percentage of organic carbon by a factor; however,

conversion factors vary between 1.4 and 3.3 (Rasmussen and Collins, 1991; Krull et al.,

2003) and this large range is due to the inherent differences between the soils. Most

commonly, a conversion factor of 1.72 is used (Baldock and Skjemstad, 1999).

1.9.6 The Soil Organic Carbon (SOC)

This is the measure of the carbon (C) stored in the SOM. Organic carbon (OC) enters

the soil through the decomposition of plant and animal residues, root exudates, living and

dead microorganisms and soil biota (Sikora and Stott, 1996). SOC is the main source of

energy and nutrients for the soil microorganisms (Dobermann and Cassman, 2002). SOC

affects many soil characteristics including colour, nutrient-holding capacity (cation and anion

exchange capacity), nutrient turnover and stability which in turn influence water relations,

aeration and workability (Grandy and Robertson, 2007). The ease and speed with which SOC

becomes available is related to the fraction of SOM in which it resides (Franzluebbers, 2002).

SOM contains approximately 58% C; therefore, a factor of 1.72 can be used to convert OC to

SOM. TOC is expressed as %C per 100g of the soil. SOC is important because it is one of the

most important constituents of the soil due to its capacity to affect plant growth as both a

source of energy and the trigger for nutrient availability through mineralization (Cameron et

al., 1996). The direct effect of poor SOC are decreased microbial growth, activity and

nutrient mineralisation due to the shortage of energy sources (Cameron et al., 1996).

1.10 Aim and objectives of the Study

1.10.1 Aim of the study

This work was aimed at evaluating the ecological impact of POME on soil fertility

with respect to the soil enzymes and soil physicochemical properties.

1.10.1Specific objectives of the study

The study is designed to achieve the following specific objectives;

1. To determine the physicochemical properties of the palm oil mill effluents such as

pH, dissolved oxygen (DO), chemical oxygen demand (COD), biochemical oxygen

26

demand (BOD), nitrogen, Potassium, oil and grease, total phosphate, total solids,

suspended solid, total volatile solid, heavy metal such as cadmium, copper, iron and

chromium.

2 To determine the activities of the catalase, lipase and dehydrogenase of a POME

contaminated soil

3 To determine physicochemical properties of a POME contaminated soil such pH,

nitrogen, available phosphorus, cation exchange capacity, organic carbon, organic

matter and exchangeable cations

27

CHAPTER TWO

MATERIALS AND METHODS

2.1 Materials

2.1.1Palm oil mill effluents

2.1.2Soil samples

2.1.3 Equipment

The equipment used was obtained from the Project DevelopmentAgency (PRODA)

Enugu, Departments of Biochemistry, Soil Science and Crop Science, University of Nigeria,

Nsukka. They include:

Equipment Manufacturer

Analytical balance (0-400g) Meter telado B-204-8 Switzerland

Centrifuge Gallenkamp, England

Condenser Pyrex, England

Conical flask Pyrex, England

Desiccator Wheaton, USA

Filter paper Whatman

Flat bottom flask Pyrex, England

Glass rod Pyrex, England

Heating mantle Everest, China

Hot air oven (0-200C) Gallenkamp, England

pH meter Hanna instruments

Photoelectric flame analyser Gallenkamp, England

Refrigerator Thermocool

Soxhlet extractor Pyrex, England

2.1.4 Chemicals and Reagents

The chemicals used for this study were of analytical grade. The chemicals used in this

study include:

Chemicals Manufacturer

Absolute ethanol BDH Analar, England

Alkaline iodide-azide JHD, China

Ammonium acetate BDH Analar, England

28

Ammonium chloride JHD, China

Ammonium fluorate indicator BDH Analar, England

Ammonium molybdate Sigma-Aldrich, USA

Calcine indicator Sigma-Aldrich, USA

Calcium carbonate BDH Analar, England

Concentrated sulphuric acid BDH Analar, England

Erichrome Black-T indicator Sigma-Aldrich, USA

Ferrous sulphate Sigma-Aldrich, USA

Hydrochloric acid Sigma-Aldrich, USA

Hydrogen peroxide Sigma-Aldrich, USA

Lauric acid Sigma-Aldrich, USA

Magnesium chloride Merck, England

Phenol red Sigma-Aldrich, USA

TTC (2, 3, 5-triphenyltetrazolium chloride) BDH Analar, England

2.2 Methods

2.2.1 Collection of the Fresh and Fermented Palm Oil Mill Effluents

The palm oil mill effluents samples were collected from six different palm oil milling

sites in six different villages in Umuaka, in Njaba Local Government Area of Imo State. In

every milling site fermented and fresh effluents were collected separately in different sterile

containers.

2.2.2 Collection of the Soil Samples

The soil samples were collected from six different palm oil milling sites in six

different villages in Umuakain Njaba Local Government Area of Imo State. Sampling was

done three times from the six respective effluent dumpsites, sites about 10 yards away from

the respective effluent dumpsites, and a non-effluent dumpsite about 1km from each

dumpsites which served as the control according to the method described by (Okwute and

Isu, 2007). Soil samples from each of these three sites were collected at depths of 0-15cm and

15-30cm respectively.

2.2.3 Preparation of the Soil Samples

The soil samples were air-dried and sieved using a 2 mm sieve and then stored in

fresh, clean polyethylene bags in the refrigerator at 4˚C prior to laboratory analysis so as to

maintain the stability of samples without significant alteration in their biological properties.

29

2.2.4 Preparation of the Fresh and Fermented POME samples

The POMEsamples collected aseptically were fully certified by their respective mill

operators and stored under refrigeration at 4˚C prior to laboratory analysis.

2.2.5 Preparation of Reagents

2.2.5.1 Phosphate Buffer (0.05N, pH 7.4)

Disodium hydrogen phosphate (2.37g) was dissolved with small volume of distilled

water in one litre standard volumetric flask and made the volume up to one litre with distilled

water. Sodium dihydrogen phosphate (dihydrate) 2.98g was also dissolved with small volume

of distilled water in one litre standard volumetric flask and made the volume up to one litre

with distilled water. The resulting solution of disodium hydrogen phosphate, 720ml was

mixed with 280ml of sodium dihydrogen phosphate (dihydrate) and then standardised at pH

7.4.

2.2.5.2 Concentrated Sulphuric Acid (6N H2SO4)

Concentrated tetraoxosulphate (iv) acid, H2SO4, 98% was prepared by adding

16.30ml of the concentrated acid solution to 83.7ml of distilled water in a 100ml standard

volumetric flask.

2.2.5.3 Hydrogen Peroxide Solution (2mMol H2O2)

Hydrogen peroxide, H2O2, 40% was prepared by diluting 0.2ml of stock solution with

99.8ml of distilled water in a 100ml standard volumetric flask.

2.2.5.4 Potassium Permanganate Solution (0.1N KMnO4)

Potassium permanganate (KMnO4), 1M was prepared by dissolving 15.7g of the

pellets with small amount of distilled water in a standard volumetric flask. The standard

volumetric flask with its content was shook very well and the solution made up to 1litre mark

with distilled water.

2.2.5.5 3% TTC (2, 3, 5-triphenyltetrazolium chloride)

Triphenyltetrazolium chloride (TTC), 1M of it was prepared by dissolving 3g of its

powder in 100ml standard volumetric flask with little distilled water. It was shaken very well

and the resulting solution was made up to 100ml mark of the standard volumetric flask with

distilled water.

30

2.2.5.6 Potassium Chloride Solution (1M KCl)

Potassium chloride, 1M was prepared by dissolving 2.485g of its pellets with little

amount of distilled water in one litre standard volumetric flask. It was shaken very well and

made up to the 1000ml mark of the flask with distilled water.

2.2.5.7 Ammonium Metavanadate Reagent

Ammonium molybdate (1M), 25g and ammonium metavanadate (1M), 1.25g were

added into 300ml of distilled water and then warmed to dissolve. After cooling, the mixture

was made up to 500ml using distilled water. On the other hand, 5M HCL was prepared by

adding 215ml of concentrated HCl (36%) to 500ml of distilled water. The two solutions were

subsequently mixed.

2.2.5.8 Carbon Carbonate Solution

Carbon carbonate (1M), 2.497g was digested with 10ml of 50% HCl and dissolved to

1 litre with distilled water.

2.2.5.9 Sodium Chloride Solution (1M NaCl)

Sodium chloride solution (1M), was prepared by dissolving 2.541g of it with little

amount of distilled water and made the volume up to 1litre with distilled water to form the

stock solution. Dilute solution of the resulting sodium chloride was made by taken 5ml of the

stock solution and diluting with 50ml of distilled water.

2.2.5.10 Magnesium Chloride Solution (1M MgCl2)

Magnesium chloride was prepared by dissolving 2.098g in small volume of distilled

water and made the volume up to1litre with distilled water.

2.2.5.11 10% (w/v) Potassium Hydroxide Solution (KOH)

Potassium hydroxide solution was prepared by dissolving 100g of KOH with little

distilled water and made the volume up to 1litre with distilled water.

2.2.5.12 EDTA (Ethylenediamine tetra-acetic acid)

EDTA (0.01N) was prepared by dissolving 0.974g free acids in distilled water and

making the volume up to 1litre with distilled water.

31

2.2.5.13 Formalin Solution

Formaldehyde (1M) solution was prepared by diluting 75ml of the stock solution with

equal volume of distilled water and standardized with 0.1N sodium hydroxide solution with 2

drops of phenolphthalein indicator.

2.2.5.14 Sodium Hydroxide Solution (0.1 NaOH)

Sodium hydroxide (0.1N) solution was prepared by dissolving 1.33g of the pellets in

small volume of distilled water and made the volume up to 1litre with distilled water.

2.2.5.15 Potassium Dichromate Solution

Potassium dichromate solution (1N) was prepared by dissolving 97.9g in small

distilled water. It was shaken very well and the volume made up to 1litre with distilled water.

2.2.5.16 Ferrous Sulphate Solution

Ferrous sulphate (0.1N) solution was prepared by dissolving 4.5g of Ferrous sulphate

in a small volume of distilled water. It was shaken very well and the volume was made up to

1litre with distilled water.

2.2.5.17 Sodium Acetate Solution

Sodium acetate (0.2M) solution was prepared by dissolving 16.4g in a small volume

of distilled water. It was shaken and the volume was made up to 1litre with distilled water.

2.2.5.18 Lauric Acid

Lauric acid (0.2M) solution was prepared by dissolving 0.2g of lauric acid in small

volume of distilled water. It was shaken very well and the volume was made up to1litre of

distilled water.

2.2.5.19 40% Sodium Hydroxide (40% NaOH)

Sodium hydroxide solution was prepared by dissolving 40g of the pellets with small

volume of distilled water. The solution was shaken very well and the volume was made up to

400ml with distilled water.

2.2.5.20 Hydrogen Chloride (0.01M HCl)

The hydrogen chloride (0.01M) solution was prepared by diluting 0.86ml stock

solution of hydrochloric acid with distilled water to 1litre and subsequently standardized

using sodium carbonate anhydrous.

32

2.2.6 Determination of theFresh and FermentedPOMEpH

The pH of palm oil mill effluents were determined in – situ by the method described

by (Ademoroti, 2006) using pH Meter. About 10g of palm oil mill effluent samples were

weighed into separate beakers and mixed with 25ml of distilled water. The mixtures were

shaken well for 30 minutes and readings were taken by dipping glass electrode of digital pH

meter initially standardized with buffer solution (pH – 7) and (pH – 4.1).

2.2.7 Determination of the Soil pH and Exchangeable Acidity

The pH of soil samples were determined in – situ by the method described by

(Ademoroti, 2006) using pH Meter. Twenty grams soil samples were weighed into different

clean beakers and mixed with 50ml of distilled water. The mixtures were shaken well for 25

minutes and readings were taken by dipping glass electrode of digital pH meter initially

standardized with distilled water (pH – 7) and buffer solution (pH – 4.1). The process was

repeated with 0.1NKCl and pH values observed when distilled water and 0.01KCl were used

respectively were recorded and exchangeable acidity was calculated.

Thus,Exchangeable acidity (meq/100g) = 20.1 – (0.88 x pH in water) – (2.46 x pH in KCl)

2.2.8 Determination of Total Nitrogen

Total Nitrogen in both soils and palm oil mill effluents were determined using the

Kjeldahl method as described by (Ademoroti, 1996).

Principle: Nitrogen is present mostly in organic form, together with small quantities

of ammonium and other forms. This method measures only organic and ammonium forms of

nitrogen. On treatment of sample with potassium sulphate and concentrated Sulphuric acid

and heat, nitrogen, if present in the sample is converted to ammonium sulphate.

Method: One gramme(1g) of sieved soil sample was placed in a round bottom flask

containing 10ml of distilled water and the mixture left to stand for an hour (and for the

POME. One gramme(1 g)of effluent was introduced into a round-bottom flask, 10ml of

distilled water was added and the mixture left to stand for about 30minutes). Then, 20g of

catalyst digestion mixture and 10ml of concentrated H2SO4 were added into the mixture,

refluxed for 90 min and cooled. The mixture was agitated and introduced into a 250 ml

volumetric flask and made up to the 100ml mark using distilled water. From this new

volume, 20 ml was taken and put in a distillation flask, 20ml of NaOH was added and a few

pieces of zinc (to enable smooth boiling of the mixture) and the mixture was gently swirled.

Approximately 50 ml of the sample in the 250 ml volumetric flask was placed in a conical

33

flask and 10ml of boric acid (4%), 2 drops of mixed indicator was added into it. The conical

flask was then placed below the condenser to allow the tip of the condenser dip into the

solution. After condensation, about 150 ml of condensate was collected, and the flask

removed to avoid sucking back. The mixed indicator in the condensate turned blue owing to

the dissolution of ammonia. The condensate was titrated against HCl (0.1N) until a colour

change to light brown (original colour of the indicator) was observed. Total nitrogen was then

calculated.

Calculation:

% �� . ��

�� . � � � ��

2.2.9 Determination of Available Phosphorus

Available phosphorus in the effluent and Soil samples were determined by Vanado-

molybdo-phosporic acid colorimetric method as described by Ademoroti (1996).

Preparation of Phosphorus Stock Solution

Phosphorus stock solution was prepared by dissolving 0.88g of dried phosphorus

dihydrogen orthophosphate (dried at 105˚C for 1hr) with distilled water and 1ml of

concentrated HCl was added. The mixture was diluted to 200ml with distilled water and 2ml

of toluene was subsequently added to obtain 1mgP/ml.

Preparation of Working Standard for the Determination of Phosphorous

The standard was prepared by taking: 2ml, 4ml, 6ml, 8ml and 10ml of the

phosphorous stock solution respectively into different 200ml volumetric flasks and diluting

them to mark with distilled water. About, 5ml of each phosphorus standard solution was

pipette into 50ml graduated flask then 10ml molybdate mixture was added and diluted to

mark with water, allowed to stand for 15 min to allow for colour development. The

absorbance was measured at 490nm against blank and a graph of absorbance against

concentration was plotted to get the standard curve.

Method: One gramme of soil samples (and also for the POME samples, one gram)

was introduced into 500ml conical flasks for the determination of total available phosphate in

soil and POME respectively. Then, 200ml of sulphuric acid (0.002N) was added to each

conical flask respectively. The suspension was agitated for 30mins and filtered to get a clear

solution. Then 50ml of this filtrate was introduced into a clean conical flask and 2ml

34

ammonium molybdate was added, followed by five drops of SnCl2 solution. At this point, a

blue coloration was observed. The absorbance was read at 490nm. The spectrophotometer

was zeroed using a blank (distilled water) containing all the reagents (except the sample). The

concentrations were then extrapolated from the standard curve.

Calculation:

� �!" ��! �� /$� %&!��&�' �� !��"�

(��" � )�� '��

2.2.10 Determination of Exchangeable Cations

2.2.10.1 Determination of Sodium and Potassium

The exchangeable cations determined include calcium (Ca), magnesium (Mg), sodium

(Na), and potassium (K). These were determined using modified methods described by IITA

(1979) and Agbenin (1995). A flame photometer (FP 640) was used in this assay.

Standard for the DeterminationPotassium (K)

Potassium chloride (1.91 g) was dissolved in one litre of distilled water and 10ml

from this preparation was diluted in 100ml of distilled water. Thereafter, 1ml of the resulting

solution was diluted in 50ml of water to give 2ppm of potassium.

Standard for the Determination of Sodium (Na)

A known weight,2.54g of sodium chloride was dissolved in one litre of distilled water

to make a stock solution. Then 10ml of the stock solution was diluted to 100ml with distilled

water; 1ml of the resultant solution was diluted to 50ml with distilled water to give 2ppm of

sodium.

Preparation of Soil Leachate for Elemental Analysis in Soil

Filter papers were placed into different funnels and the funnels channelled into

collecting conical flasks and 5g of each soil samples was placed in each filter paper. Then

100ml of ammonium acetate (pH 7) was used to leach the soil samples, thus producing the

first leachate. After collecting the first leachate, the soil samples were washed using methanol

and the leachate collected separately. The soil samples were left to dry for 24hrs, after which

0.1N of KCl was used to leach the soil. The leachates were collected in different bottles

specific to cation exchange capacity (CEC) experiment.

35

Preparation of Fresh and Fermented POME for Elemental Analysis

A measured volume, 25ml of concentrated HNO3,was added into conical flasks

containing 5g of each palm oil mill effluent samples and the mixtures were bleached using

10ml H2O2. The mixtures were incubated at room temperature for 30minutes before placing

the conical flasks on heating mantles. The colour of the liquids changed to brown colour and

emitted white fumes to evaporate reagents used for digestion, after which the brown colour

returned to the original colourless liquid and heating was stopped. The flasks were brought

down and allowed to cool. Their volumes were made up to 100ml with distilled water and the

suspended fatty substances were filtered off, leaving the clear filtrates.

Method: The flame photometer was setup according to the instruction provided by

the manufacturer. The instrument readout was calibrated using the standard solution and the

meter reading was set at 100%E (emission) while aspirating the top concentration of the

standard. The standard curves were plotted, which were used to calculate the concentration of

sodium and potassium respectively. Using the Gallenkamp flame analyser, initial leachates

from the soil samples were analysed for sodium and potassium ions. A sensitive electrode-

like material was inserted into the leachates of the different soil samples and automatically it

gave the absorbance readings of the sodium or potassium according to the standard used to

standardise the analyser (either sodium or potassium standards). Concentration of the element

in sample solution was read from the standard curve

Calculation:

% * �� ""� � �� )�� '��

��

Where ppm Absorbance of sample

Slope

Dilution factor �DF� Total volume

Volume of sample

2.2.10.2 Determination of Calcium Ions

To 10ml of treated POME (and for soil sample, 10ml of lecheate was used), 25ml of

distilled water was added to the conical flask. About 25ml of potassium hydroxide (10%) was

added followed by 25ml of deionised water. It was titrated against 0.01N EDTA using two

drops of calcine indicator (colour changes from light yellow to light orange). The

concentration of the calcium was calculated.

36

Calculation:

% ��'��

�C�� DE�% �!) � F�� DE�% � ��' G�� '��'��

�� G�� !��"� � ��H�� !)

Where atomic wt of Ca2+ = 40, Molarity of EDTA 0.01

2.2.10.3 Determination of Magnesium Ions

A known volume, 10ml of treated POME (and for soil, 10ml of lecheate was used),

25ml of deionised water was added to the conical flask. Ammonium chloride buffer (25 ml)

solution (pH 7.0) was also added. It was then titrated with 0.01N EDTA using erichrome

black- T indicators that changes colour from black to purple at end point. The concentration

of the magnesium (Mg) was determined.

Calculation:

% F�

�C�� DE�% �!) � F�� DE�% � %��' G�� F� � ��

�� G�� !��"� � ��H�� !)

Where atomic weight of Mg =24, normality = 0.01N

EDTA = volume of ethylenediamine tetra-acetic acid used

2.2.11 Determination of Cation Exchange Capacity (CEC)

Cation Exchange Capacity was determined according to the method described by

IITA (International Institute for Tropical Agriculture, 1979) and Agbenin (1995).

The soil leachate (50 ml) was carefully transferred into a clean dry conical flask and

20ml of 2% formalin was added. Then two drops phenolphthalein indicator was added and

the resulting solution was titrated with 0.1N Sodium hydroxide till a light pink colour is

detected at the end point and the titre value was recorded. The process was repeated three

times to achieve a triplicate determination of the Cation Exchange Capacity. The Cation

Exchange Capacity of the soil sample was calculated according to the formula below:

Calculation:

�D� ��DH/�� !�� I� ��. � ��

�� H��

37

2.2.12 Determination of Heavy Metals in the Fresh and Fermented POME

The heavy metals in POME samples (Cd, Cu, Cr and Fe) were determined after the

digestion of the effluents using Atomic Absorption Spectrophotometer as described by

APHA (American Public Health Association, 1998).

Wet digestion: One gram of POME samples were weighed into clean dried conical

flasks. A measured quantity, 20ml of concentrated Nitric acid (HNO3) and 20ml of hydrogen

peroxide (H2O2) were added to each conical flask. They were digested on a hot plate at 130oC

with the glasses covered with watch glasses. The digestion was continued until the colour

appeared clear. They were cooled and made up to 50ml, mixed and transferred to 250ml

volumetric flasks. The residues were washed severally with deionised water and made up to

the mark. They were mixed very well and filtered into clean dried containers and stored for

determination of minerals.

2.2.13 Determination of Dissolved Oxygen (DO) in the Fresh and Fermented POME

The dissolved oxygen was determined in – situ using DO meter, as described by the

scheme of Ademoroti (1996).

Principle: The azide modification of the iodometric method is the most common

chemical technique to measure dissolved oxygen by addition of manganese sulphate and

alkali-iodide-azide. Mn2+ is oxidised if oxygen is present to give a white precipitate. By

addition of sulphuric acid, free iodine is liberated, which is converted into blue colour, by

adding starch as indicator. By addition of sodium thiosulphate, the blue colour disappears to

give an indication of dissolved oxygen present originally. It is pertinent that microbial

activity must be stopped at the time of sample collection. Copper sulphate was used for this

purpose.

Method: Standard BOD bottles were filled with palm oil mill effluents completely.

MnSO4(2 ml) and 2 ml of alkaline iodide-azide were added onto the surface of the samples in

the BOD bottles and allowed to stand for 5 minutes. A Known quantity, 1ml of concentrated

Sulphuric acid solution was added and the mixture left to stand for another 5 minutes. A

volume, 200ml of these clear solutions were collected in different conical flasks and 2drops

of starch indicators were added to each flask, which gives a deep blue coloration.

Subsequently, the mixtures were titrated with sodium thiosulphate (0.025N) solution till the

colour changed to colourless and the titre values recorded. The DO was calculated using the

formula:

38

EI ��/$� J�� )�� (�)�� !) � K��

C�� (��"� �!)

2.2.14 Determination of Biochemical Oxygen Demand (BOD) in the Fresh and

Fermented POME

The Biological oxygen demand (BOD) was determined using the method described

by Ademoroti (1996).

Principle: Biological oxygen demand is the quantity of oxygen utilised by a mixed

population of micro-organism in oxidation of organic matter at room temperature. The

diluted water containing phosphate buffer, manganese sulphate, calcium chloride and ferric

chloride was saturated with dissolved oxygen. The palm oil mill effluents supply the organic

matter and the diluted water supplies dissolved oxygen. However, the degree of

reproducibility of biological oxygen demand cannot be precisely defined on account of

variations that occur in decomposition of organic matter.

Method: Five litres of distilled water was added in a glass container and compressed

air was bubbled into it for 2 days to attain saturation. About 1ml each of manganese sulphate,

phosphate buffer, ferric chloride and calcium chloride solution were added for each litre of

distilled water. About 2ml of the effluent was added to 1000ml of water and the pH of the

solution was maintained at pH 7. The dilution factor was noted. A dilution of the POME

sample was made, such that about 50% depletion of dissolved oxygen takes place and

residual dissolved oxygen after incubation for 5days.

Six bottles of BOD were prepared; 2 bottles as blank for determination of initial

dissolved oxygen, and another 2 bottles for diluted effluent. The 4 bottles were kept for

incubation at room temperature for 5days. One bottle for blank and one sample bottle were

used to determine dissolved oxygen after adding manganese sulphate, alkaline-iodide-azide

and concentrated sulphuric acid and titrated against 0.025N sodium thiosulphate solution

using starch indicator, the burette reading was noted. After 5 days the final dissolved oxygen

was determined.

Calculation:

BOD (mg/L) = (D0 – D1) – (C0 – C1) x % dilution

Where D0 =dissolved oxygen(DO)of Sample on the 0th day

D1 = dissolved oxygen(DO) of Sample on the 5th day

39

C0 = dissolved oxygen(DO) of Blank on the 0th day

C1 = dissolved oxygen(DO) of Blank on the 5th day

2.2.15 Determination of Chemical Oxygen Demand (COD) in the Fresh and Fermented

POME

The chemical oxygen demand was determined by titrimetric/dichromate oxidation

method as described by Ademoroti (1996).

Principle: chemical oxygen demand is a measure of oxygen equivalent of that portion

of the organic matter in the sample that is susceptible to oxidation by strong chemical

oxidant. Most of the organic matter is destroyed by boiling in chromic and sulphuric acid.

This is done alongside the blank to compensate for any error that may result because of the

presence of extraneous organic matter in the reagents.

Method: A Know weight, 0.4g of H2SO4 and 20ml of the POME sample were mixed

in a reflux flask followed by the addition of 10ml of 0.25N of K2Cr2O7. Some pumice stones

were dropped in the mixture and slowly, 30ml of concentrated H2SO4-AgSO4 reagent was

added. Contents were mixed thoroughly and flask was connected to condenser and refluxed

for 2hrs. The setup was allowed for some time to cool and the mixture made up to 150ml

with distilled water. Approximately 3drops of ferroin indicator was added and titrated against

0.1N of ferrous ammonium sulphate solution, till colour changes from green to wine red and

the end point was noted. Same procedure was performed for blank using distilled water only.

The chemical oxygen demand was calculated.

Calculation:

�IE ��/$� �C� L CM� � �� !��" �� !)

C�� (��"� � ��

Where V1 = Titre value of Sample

V2 = Titre of Blank

8000 = milliequivalent weight of oxygen X 1000 ml/L.

2.2.16 Determination of Total Solids, Suspended Solids and Total Volatile Solids in the

Fresh and Fermented POME

Total solids, suspended solid and total volatile solids were determined by a modified

method as described by Walkley and Black (1934).

40

2.2.16.1 Total Solids and Total Volatile Solids in the Fresh and Fermented POME

Dry weight of the crucible was taken as W1 and 25ml of the effluent sample was

added into the crucible and then heated to dryness and was allowed to cool for 20 minutes. It

was weighed again and recorded as W2; the crucible was subsequently placed in muffle

furnace at a temperature of 650˚C for 30 min for the volatile and organic matter in the solids

to evaporate, leaving the fixed solids. The crucible was allowed to cool then it was weighed

and recorded as W3. The concentration of volatile solids was calculated.

Calculation:

N��) !��) ��/$� �OM L OP�

C�� (��"�Q ��

�� C�� (��) ��C(��/$� �OM L O��

C�� (��"��

Total Volatile Solid (TVS) (mg/L) = Total Solid – Fixed Solid

2.2.16.2 Total Suspended Solids in the Fresh and Fermented POME

A homogenized effluent sample was centrifuged at 2000rpm for 10minutes and the

residue was collected by decantation then heat-dried at 105˚C and allowed to cool. The

weight was measured and recorded as the weight of the total suspended solids.

Calculation:

�� (�!")) (��) ��((� �OM L O��

C�� (��"� � ��

Where W1 = weight of empty crucible

W2 = weight of crucible and heat – dried residue

2.2.17 Determination of Soil Organic Carbon and Organic Matter

Organic carbon measurement was carried out by the method described by Kalembasa

and Jenkinson (1973).

A known weight, 1g of each soil sample was measured into different clean dried

round bottom flasks. The samples were digested using 10ml of (1N) K2Cr2O7 and 20ml of

concentrated H2SO4. Then 200ml of distilled water and a little quantity of ammonium

flourate were added to the solution (to clearly define the end point). The samples were then

titrated against (1N) Ferrous Ammonium Sulphate (FAS) using 1ml of diphynylamine as

indicator till the colour changed to brilliant green at the end point. The process was repeated

41

for blank with same quantity of the chemicals but without soil. The titre values were recorded

and total organic carbon determined.

Calculations:

(a). % ��&� �� L �/(��P. R��/��

(b). % Organic Matter = % Carbon × 1.724

Where, g = weight of soil sample in gram

S = Volume (ml) of FAS used with blank titr|ation

T = Volume (ml) of FAS used with sample titration

The factor 1.724 is based on the assumption that carbon is only 58 % of the organic matter.

2.2.18 Determination of Oil and Grease in the Fresh and Fermented POME (soxhlet

extraction method)

Oil and grease was determined using gravimetric method after soxhlet extraction as

described by Ademoroti (1996).

Principle: The solvent, n-hexane was used to extract the oil and grease from the

samples. The resulting values of oil and grease were determined after evaporating the n-

hexane from the recovered oil.

Method: Raw POME was allowed to naturally settle for 24 – 48 hours to obtain

POME sludge. A portion of the oily POME sediments was treated with n-hexane (C6H14) in a

soxhlet until all the oil has been removed to obtain non-oily POME sediments. A soxhlet

flask (250ml) was oven dried and its dry weight was taken. Cotton wool was used to block

the kink of the thimble and 20g of the POME was added to the base of the thimble. n-hexane

(200 ml) was added in the soxhlet flask, the thimble was fixed into the flask and the

condenser was fixed into the thimble. The set up was mounted on a heating mantle with the

temperature above 700C and oil extraction by n-hexane was allowed to go on for about 3

hours. The process was repeated for the other POME samples. After heating, the n-hexane

was recovered by carefully removing the POME sample and cotton wool from the thimble

and heating the mixture of oil and n-hexane in the round flat bottom flask. After recovering

of the n-hexane, the remaining mixture was oven dried to ensure that all the remaining n-

hexane are totally evaporated. The difference between the empty oven dried flask and the oil

containing flask was determined and the percentage of oil and grease was calculated.

42

Calculation:

% I�� ) ��! O�� O��

O�� (��"� ��

Where W1 (g) = difference between the weight of empty oven dried flask and the oil

containing flask

Weight of the sample = 20g

2.2.19 Determination of Soil Enzyme Activity

2.2.19.1 Determination of the Soil Dehydrogenase Activity

Soil dehydrogenase activity was determined using the method described by Tabatabai

(1982). Principle: Soil dehydrogenases convert 2, 3, 5-triphenyl tetrazolium chloride to

formazan that has optimum absorbance at 485nm wave length.

Method: One gram of sieved soil sample was weighed into test tubes, mixed with

1ml of 3% (w/v) aqueous 2, 3, 5-triphenyl tetrazolium chloride and stirred with a glass rod.

After 96hours of incubation at (27˚C), 10ml of absolute ethanol was added to each test tube

and the suspension was vortexed for 30seconds. The tubes were then incubated for 1hour to

allow suspended soil to settle. The resulting supernatant (5ml) was carefully transferred to

clean test tubes and absorbance was subsequently read spectrophotometrically at 485nm after

having zeroed with distilled water. The concentration of formazan was evaluated using

extinction coefficient of 15433Molcm-1 (Dushoff et al., 1965).

Calculation:

%'�� %&!��&�' � ��

D��'�� '��'�� D� � (��"� ��

Where E = 15433Molcm-1

Path length = 1cm

2.2.19.2 Determination of the Soil Catalase Activity

Catalase activity of the soil samples was assayed using the method described by Rani

et al.(2004).

Principle: Catalasedecomposes hydrogen peroxide, the extent of which was measured

by reacting it with excess potassium tetraoxomanganate (iv) oxide (KMnO4). Residual

KMnO4 is measured spectrophotometrically at 480nm.

43

Method: A known quantity, 100ml of phosphate buffer (0.05N, pH 7.4) was added to

10g of each soil sample contained in test tubes and the mixture was homogenized by shaking.

The mixture was filtered using cheesecloth and the filtrate obtained was centrifuged at 7000 x

g for 5 minutes. The supernatant obtained was decanted and 5ml of the supernatant was

introduced into differently labelled test tubes containing 0.5 ml of 2mM H2O2 and a blank

containing 0.5ml of distilled water. Then 0.5ml of 6N H2SO4 was added to the test tubes one

at a time, immediately followed by the addition of 3.5ml of KMnO4 (0.1N). The solution

was mixed thoroughly and absorbance was subsequently read using a spectrophotometer at

480nm. Spectrophotometer standard was prepared by adding KMnO4 (3.5ml) to a mixture of

phosphate buffer (2.75ml) and H2SO4 (0.5ml). The spectrophotometer was then zeroed with

distilled water before taking absorbance readings.

Calculation:

%'�� %&!��&�' � �� C��

D��'�� '��'�� D� � (��"� ��

Where E = 1206Molcm-1 (from the Standard)

Path length = 1cm

2.2.19.3 Determination of the Soil Lipase Activity

Soil lipase activity was determined using the method described by Margesin et al.

(2002).

Principle: Lipase releases lauric acid on incubation with Tween 20 and toluene at 30

0C for 18 hours under agitation.

Preparation of Standard for the Determination of Soil Lipase:

The standard was prepared by dissolving 0.1g of lauric acid in 500ml of toluene.

Serial dilutions (5ml in 10ml) of the mixture were made with toluene, with other reagents

added as was in the samples. These serial dilutions were then titrated against 0.01M NaOH

and the titre values were used to plot a standard curve of absorbance against concentration.

From this standard curve, the titre values of the samples were extrapolated to generate the

concentrations of lauric acid in soil samples.

Method: For each soil sample, 1g was weighed in triplicate into three test tubes. To

each test tube, 0.2ml of toluene, 0.6ml of Tween 20, 1.15ml of distilled water and 0.2ml of

sodium acetate was added. Each test tube was capped with silicon material and left to

incubate at 30 0C for 18hours with intermittent shaking. After incubation, 8ml of absolute

44

ethanol was added and the mixture was swirled for 10 seconds. Subsequently, each mixture

was centrifuged at 3000 x g for 10 minutes. The supernatants were decanted and to each,

0.375ml of phenol red indicator (0.02g/L in ethanol.) was added. Each of the resulting

solutions was titrated with 0.01M NaOH and the titre value was noted.

Calculation:

%'�� '�� F�� C��

(��" �� !��)��) '�� !�� !��"� �!) � ��

Where t = 18 hours, g of soil sample used = 1g of soil sample

2.2.20 Statistical Analysis

Statistical Product and Service Solutions (SPSS) software (IBM SPSS Statistics 20)

was used to carry out the statistical analysis with the results expressed as mean S standard

deviation. A one way analysis of variance (ANOVA) was carried to check for difference

between means; results were considered significant at P<0.05 and Duncan’s multiple range

test was used to discern the source of observed differences.

45

CHAPTER THREE

RESULTS

3.1 pH of the Fresh and Fermented Palm Oil Mill Effluents (POME)

As shown in Table 4, there was significant increase (p<0.05) in the pH values of the

fermented POME samples from Amurie and Obeakpu dumpsites compared to the pH value of

their respectivefresh POME samples. However, there were significant decreases (p<0.05) in

pH values of fermented POME samples from Amurie, Nnerim, Obeakpu, Olori and Eziiu

dumpsites compared to the pH values of their respective fresh POME.

Table 4: pH of the Fresh and Fermented POME

Each value is expressed as mean ± standard deviation (n = 3).

Mill Sites pH of Fresh POME pH of Fermented

Amurie 4.30±0.2 4.70±0.5

Eziisu 4.44±0.5 4.30±0.3

Nnerim 4.68±0.3 4.30±0.3

Olori 4.65±0.3 4.32±0.2

Umudiokpara 4.54±0.6 4.25±0.4

Obeakpu 4.36±0.4 4.55±0.5

46

3.2 Effect of the Fresh and Fermented POME on the Soil pH and Soil Exchangeable

Acidity

As shown in Table 5, the topsoil from Eziisu POME dumpsite had the highest

exchangeable acidity of 5.33 meq/100g and the topsoil from Amurie POME dumpsite had the

lowest exchangeable acidity of 3.21 meq/100g among the palm oil mills while all the subsoil

had lower exchangeable acidity compared to the control. The topsoil from Eziisu POME

dumpsite had the lowest pH in KCl relative to that of the control and highest exchangeable

acidity but their pH in distilled water were approximately equal. The result obtained for the

soil pH showed that soils in this study area were generally acidic. Though, POME soil and

control soils were acidic, there was a mixed trend observed in their acidity levels. With the

exception of the topsoil ten yards away from Eziisu POME dumpsite and ten yards away

from Obeakpu POME dumpsite that showed equal acidity level with the control soil pH, all

other topsoil showed slight increases in their acidity level compared to the control pH when

their pH were determined in distilled water (H2O). Apart from topsoil ten yards away from

Olori POME dumpsite and topsoil from Eziisu POME dumpsite that showed more acidity

compared to the control topsoil when their pH were determined in potassium chloride

solution (KCl), all other topsoil showed lesser acidity compared to the control. All the subsoil

showed lower acidity compared to the control in KCl. Only the subsoil ten yards away from

Nnerim and Umudiokpara POME dumpsites showed equal acidity with the control while the

rest subsoil ten yards away from other POME dumpsites showed slightly lower acidity

compared to the control subsoil when their soil pH were determined in distilled water.

47

Table 5: Effect of the Fresh and Fermented POME on the Soil pH and Soil

Exchangeable Acidity

Topsoil Subsoil

POME Sites

Soil

Samples

pH(H20) pH(KCl)

Exchangeable

Acidity

(meq/100g)

pH

(H20)

pH

(KCl)

Exchangeable

Acidity

(meq/100g)

Amurie

(10yrds) 5.5 4.9 3.21 5.5 4.5 4.19

Dumpsite 5.4 4.9 3.29 5.6 4.7 3.61

Olori

10yrds 5.2 4.3 4.95 5.7 4.5 4.01

Dumpsite 5.2 4.6 4.21 5.6 4.5 4.10

Ezisu

10yrds 5.6 4.4 4.35 5.4 4.6 4.03

Dumpsite 5.6 4.0 5.33 5.5 4.8 3.45

Nnerin

10yrds 5.2 4.4 4.70 5.3 4.6 4.12

Dumpsite 5.3 4.6 4.12 5.3 4.6 4.12

Obeakpu

10yrds 5.6 4.7 3.61 5.7 4.7 3.52

Dumpsite 5.5 4.6 3.94 5.6 4.6 3.86

Umudiokpara

10yrds 5.4 4.6 4.03 5.6 4.6 3.86

Dumpsite 5.5 4.7 3.70 5.3 4.7 3.87

Control 5.6 4.4 4.35 5.3 4.3 4.86

48

3.3 Nitrogen Content in the Fresh and Fermented POME

The results of the analysis of the nitrogen content in the fresh and fermented POME in

Table 6 show that the fresh POME from Olori and Obeakpu palm oil mills had the highest

and the least nitrogen content respectively among all the fresh and fermented POME tested.

The nitrogen content in the fresh POME from Olori and Umudiokpara palm oil mills were

significantly higher (p<0.05) than the nitrogen content in their respective fermented POME

while the nitrogen content in the fermented POME from Obeakpu palm oil mill was

significantly higher (p<0.05) than the nitrogen content in its corresponding fresh POME.

Also the nitrogen content in the fermented POME from Amurie and Nnerim palm oil mills

were not significantly higher (p>0.05) than the nitrogen content in their respective fresh

POME. However, the nitrogen content of the fresh POME from Eziisu was not

significantlyhigher(p>0.05) than the nitrogen content of its fermented POME.

Table 6: Percentage nitrogen (%) in the Fresh and Fermented POME


Mill Sites Percentage nitrogen (%) in

Fresh POME

Percentage nitrogen (%) in

Fermented POME

Amurie 1.44±0.002 1.57±0.003

Eziisu 1.47±0.003 1.37±0.003

Nnerim 1.33±0.003 1.36±0.004

Olori 2.05±0.003 1.48±0.003

Umudiokpara 1.57±0.003 1.17±0.005

Obeakpu 1.04±0.005 1.40±0.003

49

3.4 Effect of the Fresh and Fermented POME on Nitrogen Content in the Soil

In Fig.4below, there were significant (p<0.05) difference observed between the

nitrogen content in the POME dumpsites and ten yards away from the POME dumpsites as

well as that of theten yards away from the POME dumpsites and control but no significant

(p>0.05) differences was observed between the nitrogen content in the POME dumpsites and

that of the control. The result showed that the POME dumpsites were richest in nitrogen

compared to the soils ten yards away from POME dumpsites and control soil with the subsoil

in POME dumpsites having the highest amount of nitrogen relative to the nitrogen in their

respective topsoil. Also, subsoilten yards away from Amurie, Nnerim, Umudiokpara and

Obeakpu POME dumpsites had higher nitrogen content compared to their respective topsoil

in a similar trend observed in the control soil. Only the subsoil ten yards away from Eziisu

and Olori POME dumpsites had lower nitrogen content compared to their respective subsoil

and this was a deviation from the trend observed in the control subsoil.

Fig 4:Effect of the Fresh and Fermented

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control site

0

0.05

0.1

0.15

0.2

0.25

0.3

0.35

A B

Nit

rog

en c

on

ten

t (m

eq/1

00

g)

50

Fig 4:Effect of the Fresh and Fermented POME on the Nitrogen Content in the Soil

C D E F G

SITES

POME on the Nitrogen Content in the Soil

Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

51

3.5 Effect of the Fresh and Fermented POME on Phosphorus Content in the Soil

As shown in Fig. 5 and appendix 6, there were significant (p<0.05)differences

observed between the phosphorus content in the POME dumpsites and that often yards away

from the POME dumpsites, the POME dumpsites and control as well as that often yards away

from POME dumpsites and the control. Both the topsoil and subsoil from all the POME

dumpsites showed significantly lower (p<0.05) phosphorus content compared to the

phosphorus content in the topsoil and subsoil ten yards away from the POME dumpsites and

controls respectively. Only the subsoil from Umudiokpara and Obeakpu POME dumpsites

showed significant increase (p<0.05) in the nitrogen content compared to the nitrogen content

in their respective topsoil in line with the trend observed in the control. Subsoilten yards

away from the Obeakpu POME dumpsite had the highest phosphorus content compared to

the control and other soil types from other palm oil mill sites tested while topsoil and subsoil

from the Olori POME dumpsite had the least nitrogen content among all the soil types tested.

Fig. 5: Effect of the Fresh and Fermented POME on Phosphorus Content in the Soil

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

5

10

15

20

25

30

35

40

A B

Ph

osp

ho

rus

(pp

m)

52


C D E F G

SITES


Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

53

3.6 Phosphorus Content in the Fresh and Fermented POME

In Table 7 below, there were significant (p<0.05) differences observed between the

phosphorus content in the fresh and fermented POME from all the six sites tested. There was

significant decrease(p<0.05) in the phosphorus content of the fermented POME from Amurie

palm oil mill relative to the phosphorus content in the corresponding fresh POME. Fresh

POME from Amurie, Nnerim and Umudiokpara palm oil mills showed significantly higher

(p<0.05) phosphorus content relative to those of their corresponding fermented POME. In

addition, fermented POME from Eziisu, Olori and Obeakpu palm oil mills showed

significantly higher (p<0.05) phosphorus content compared to the phosphorus content of their

respective fresh POME.

Table 7: Phosphorus content in the fresh and fermented POME


Mill Sites Phosphorus (ppm) content

in Fresh POME

Phosphorus (ppm) content in

Fermented POME

Amurie 5.55±0.005 3.90±0.004

Eziisu 4.43±0.005 4.93±0.003

Nnerim 5.32±0.009 4.24±0.003

Olori 3.89±0.003 5.84±0.003

Umudiokpara 5.72±0.005 5.56±0.003

Obeakpu 5.68±0.005 6.14±0.003

54

3.7 Calcium Content in the Fresh and Fermented POME

Table 8 showsthe calcium content in thefresh and fermented POME. There were

significant (p<0.05) differences observed in the calcium content in the fresh and fermented

POME from Amurie, Nnerim, Olori, Umudiokpara and Obeakpu palm oil mills but no

significance (p>0.05) difference was observed in the calcium content in fresh and fermented

POME from Eziisu. Fresh POME from Nnerim and Olori palm oil millshad significantly

higher (p<0.05) calcium content compared to those of their respective fermented POME and

other fresh POME tested. In addition, fermented POME from Amurie, Umudiokpara and

Obeakpu palm oil mills had significantly higher (p<0.05) calcium content compare to those

of their respective fresh POME.

Table 8: PercentageCalcium (%) Content in the Fresh and Fermented POME


Mill Sites Percentage Calcium (%) in

Fresh POME

Percentage Calcium (%) in

Fermented POME

Amurie 0.07±0.002 0.07±0.004

Eziisu 0.07±0.002 0.06±0.004

Nnerim 0.09±0.002 0.07±0.003

Olori 0.08±0.004 0.07±0.003

Umudiokpara 0.07±0.004 0.09±0.004

Obeakpu 0.06±0.003 0.08±0.004

55

3.8 Magnesium Content in the Fresh and Fermented POME

As shown in Table 9, there was no significant decrease (p>0.05) in the magnesium

content in the fermented POME from Amurie, Eziisu and Obeakpu palm oil mills compared

to those of their respective fresh POME while there was no significant increase in the

magnesium content in the fermented POME from Olori and Umudiokpara palm oil relative to

those of their corresponding fresh POME. It was only the fermented POME from Nnerim

palm oil mill that had significant increase in its magnesium content comparedto magnesium

content of the corresponding fresh POME.

Table 9: Percentage Magnesium (%)in the Fresh and Fermented POME

Mill Sites Percentage Magnesium

(%) Fresh POME

Percentage Magnesium (%)

Fermented POME

Amurie 0.03±0.004 0.03±0.006

Eziisu 0.03±0.002 0.03±0.003

Nnerim 0.03±0.003 0.04±0.003

Olori 0.03±0.003 0.03±0.003

Umudiokpara 0.04±0.004 0.04±0.004

Obeakpu 0.04±0.003 0.03±0.003

56


3.9 The Sodium Content in the Fresh and Fermented POME

InTable 10, there were significant (p<0.05) differences in the sodium content in all the

fresh and fermented POME samples tested. Sodium content in the fresh POME from Nnerim,

Olori and Umudiokpara palm oil mills were significantly higher than the sodium content of

their respective fermented POME. However, there were significant increases (p<0.05) in the

sodium content in the fermented POME from Amurie, Eziisu and Obeakpu palm oil mills

compare to the sodium content of their respective fresh POME.

Table 10: Percentage Sodium in the Fresh and Fermented POME

Mill Sites Percentage Sodium (%) in

Fresh POME

Percentage Sodium (%)

in Fermented POME

Amurie 1.38±0.003 1.46±0.004

Eziisu 1.39±0.005 1.45±0.004

Nnerim 1.64±0.003 1.56±0.004

Olori 1.61±0.004 1.43±0.003

57


3.10 Potassium Content in the Fresh and Fermented POME

In Table 11, there were significant increases (p<0.05) in the potassium content in the

fermented POME samples from Amurie, Olori, Umudiokpara and Obeakpara dumpsites

compared to the potassium content in their respective fresh POME samples.However, there

were no significant decreases (p>0.05) in the potassium content in the fermented POME

samples from Eziisu and Nnerim dumpsites compared to potassium content of their

respective fresh POME samples. The fresh and fermented POME from Amurie palm oil mill

had the least and highest potassium content respectively among all the POME samples tested.

Table 11: Percentage Potassium inthe Fresh and Fermented POME

Umudiokpara 1.45±0.005 1.31±0.004

Obeakpu 1.44±0.004 1.56±0.004

Mill Sites Percentage Potassium (%)

in Fresh POME

Percentage Potassium

(%) in

FermentedPOME

Amurie 1.37±0.003 3.38±0.578

58


3.11 Calcium Content of the Soil Contaminated with the Fresh and Fermented POME

As shown in Fig. 6, there were significant (p<0.05) differences between the calcium

content of thePOME dumpsites and those of the ten yards away from POME dumpsitesand

the controls. Topsoil and subsoil from Amurie had the highest calcium content at the

dumpsite and Ten yardsrespectively relative to the calcium content in topsoil and subsoil

from other palm oil mill and the controls.The subsoil from Eziisu, Nnerim, Olori and

Obeakpu palm oil mills had significantly (p<0.05) higher calcium content relative to the

calcium content of their respective topsoil which in a trend as observed in the calcium content

in the control soils. Topsoil from Amurie and Umudiokpara POME dumpsites showed

significantly (p<0.05) higher calcium content compared to the calcium content in their

respective subsoil and control subsoil. The soils ten yards away from Amurie, Umudiokpara,

Eziisu and Obeakpu POME dumpsites had significantly (p<0.05) higher calcium content

relative to the calcium content in their respective subsoil which was a deviation from the

normal trend observed in the controls. However, subsoil ten yards away from Nnerim and

Eziisu 2.48±0.002 2.38±0.004

Nnerim 2.56±0.003 2.34±0.003

Olori 2.35±0.002 2.52±0.135

Umudiokpara 1.86±0.003 2.44±0.004

Obeakpu 1.94±0.003 2.61±0.003

Olori POME dumpsites had significantly lower (p<0.05) calcium

calcium content in their respective topsoil which was in line with the normal trend observed

in the control.

Fig. 6:Calcium Content of the Soil Contaminated with the Fresh and Fermented POME.

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2

A B

Ca

lciu

m (

meq

/10

0g

)

59

Olori POME dumpsites had significantly lower (p<0.05) calcium content relative to the



C D E F G

SITES

content relative to the



Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

60

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

3.12 Potassium Content of the Soil Contaminated withthe Fresh and Fermented POME

In Fig. 7 below, there were significant increases (p<0.05) in the potassium content in

the various soil types from all the palm oil mills tested.Onlythe topsoil and subsoilten yards

awayfrom Eziisu POME dumpsites as well as thetopsoil ten yards awayfrom Olori and

Obeakpu POME dumpsites showed no significant (p>0.05) difference in their potassium

content from that of the controls. All the topsoil from the six POME dumpsites had

significantly higher potassium content compared to the potassium content in their respective

subsoil, in line with the trend observed in the potassium content in the control. Topsoil from

Umudiokpara POME dumpsite and ten yards away from Umudiokpara POME dumpsite had

the highest and least potassium content respectively among soil types tested.

Fig. 7: Potassium Content in the Soil contaminated with the Fre

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

0

0.005

0.01

0.015

0.02

0.025

0.03

0.035

0.04

0.045

0.05

A B

Po

tass

ium

(m

eq/1

00

g)

61

Fig. 7: Potassium Content in the Soil contaminated with the Fresh and Fermented POME

C D E F G

SITES

sh and Fermented POME

Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

62

F: Obeakpu Site

G: Control

3.13 Sodium Content of the Soil Contaminated with the Fresh and Fermented POME

In Fig. 8 below, there were significant (p<0.05) differences between the sodium

content in the POME dumpsites and ten yards away from POME dumpsites as well as that of

the control soil but there were no significant (p>0.05) differences between the sodium content

in the dumpsites and control soils. Subsoil from Umudiokpara and Obeakpu POME

dumpsites had significant (p<0.05) decreases in their sodium contents compared to the

sodium content in the control soils.The subsoil ten yards away from Obeakpu POME

dumpsite had the highest calcium content compared to all the soil types tested. There was

significant (p<0.05) increase in the sodium content of the topsoil from Amurie, Eziisu,

Nnerim and Olori POME dumpsites compared to the sodium content of the control topsoil

while there were no significant (p>0.05) differences between the sodium content of the

topsoil from Umudiokpara and Obeakpu POME dumpsites compared with the sodium

content of the control topsoil. Only the subsoil from Nnerim POME dumpsite had significant

increase (p<0.05) in the sodium content while there was no significant (p>0.05) increase in

sodium content of the subsoil from Amurie, Olori and Eziisu POME dumpsites. Subsoil from

Umudiokpara and Obeakpara POME dumpsites showed significant (p<0.05) decrease in their

sodium content andhad the least sodium content among all the soil types tested compared to

those of the control subsoil . There were significant (p<0.05) increase in the level of sodium

in the topsoil Ten yards from Eziisu, Nnerim and Obeakpu POME dumpsites compared to the

sodium content in the control topsoil . However, topsoil ten yards away from Amurie and

Umudiokpara dumpsites showed no significant (p>0.05) decrease in their sodium content

compare to the sodium content in the control topsoil. All the subsoil ten yards away from the

six different POME dumpsites had significant (p<0.05) increase in their sodium content

compared to the sodium content in the control subsoil.

Fig. 8: Sodium Content of the Soil Contaminated with theFresh and Fermented POME

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

0.002

0.004

0.006

0.008

0.01

0.012

A B

So

diu

m (

meq

/10

0g

)

63


C D E F G

SITES


Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

64

3.14 Magnesium Content of the Soil Contaminated with the Fresh and Fermented

POME

As shown in Fig. 9 and appendix 6, the subsoil from both Olori POME dumpsite and

ten yards away from Olori POME dumpsite had the lowest magnesium content while the

topsoil from both Umudiokpara POME dumpsite and ten yards away from Umudiokpara

POME dumpsite had the highest magnesium content relative to other sites tested. There were

significant increase (p<0.05) in the magnesium content of the topsoil from Eziisu, Nnerim

and Umudiokpara POME dumpsites relative to the magnesium content of the control topsoil

while no significant increase (p>0.05) in magnesium content were observed in the topsoil

from Amurie, Olori and Obeakpu POME dumpsites compared to the magnesium content of

the control topsoil.. In addition, all the six subsoil ten yards away from their respective

POME dumpsites showed significant increase (p<0.05) in magnesium content compare to the

magnesium content of the control subsoil. This result showed that both the topsoil and subsoil

from Amurie, Eziisu, Nnerim, Umudiokpara and Obeakpu POME dumpsites and subsoil

fromOlori POME dumpsite were richest in magnesium compare to that of the control topsoil

and subsoil .

Fig. 9: Magnesium Content of the Soil Contaminated with the Fresh and Fermented POME

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

Site A Site B Site C

Ma

gn

esiu

m (

meq

/10

0g

)

65


Site C Site D Site E Site F Site G

SITES


Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

66

3.15 Effect of the Fresh and fermented POME on the Cation Exchange Capacity (CEC)

in the Soil

As shown in Fig. 10, there were significant increases(p<0.05) in the cation exchange

capacity observed in both the topsoil and subsoil from the six dumpsites relative to the cation

exchange capacity in the control topsoil and subsoil. Also, all the topsoil and subsoil ten

yards away from their respective POME dumpsites showed significant increase (p<0.05) in

the cation exchange capacity compared to the cation exchange capacity obtained in the

control topsoil and control subsoil. The result showed that thecation exchange capacity in the

topsoil from all the POME dumpsites though significantly higher than the cation exchange

capacity ofthe controltopsoil, had the lowest cation exchange capacity in all the palm oil mills

compared to those of the subsoil.


the Soil

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

5

10

15

20

25

A B

Ca

tio

n E

xch

an

ge

Ca

pa

city

(m

eq/1

00

g)

67


C D E F G

SITES


Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

68

3.16 Heavy Metal Content in the Fresh and Fermented POME

In Table 12 below, there were significantly lower (p<0.05) level of copper in the

fermented POME from Amurie and Obeakpu palm oil mills compared to the level of copper

found in their respective fresh POME. However, significantly higher (p<0.05) level of copper

were observed in the fermented POME relative to copper level in the respective fresh POME

from Eziisu, Nnerim and Olori palm oil mills while no significant increase (p>0.05) in the

copper level in the fermented POME from Umudiokpara palm oil mill with respect to the

corresponding level of copper in the fresh POME was observed. There were no detectable

levels of cadmium in both the fresh and fermented POME from all the six palm oil mills

analysed. There were no detectable levels of chromium in the fresh and fermented POME

from Eziisu, Nnerim, Umudiokpara and Obeakpu palm oil mills while traces of chromium

fromAmurie and Olori palm oil mills were significantly (p<0.05) higher in the fresh POME

than in the corresponding fermented POME. There were significantly higher iron levels in the

fermented POME from Amurie, Olori and Umudiokpara palm oil mills compared to the iron

level in their respective fresh POME. On the contrary, fresh POME from Eziisu, Nnerim and

Obeakpu palm oil mills had significantly higher (p<0.05) iron level compared to the iron

level in their respective fermented POME.

69

Table 12: Heavy Metal Content inthe Fresh and Fermented POME

Mill Sites POME Sample

Copper

(mg/L)

Cadmium

(mg/L)

Chromium

(mg/L)

Iron

(mg/L)

Amurie Fresh 2.30±0.10 ND 0.07±0.0 0.62±0.005

Fermented 1.61±0.01 ND 0.02±0.0 1.23±0.002

Eziisu Fresh 0.20±0.12 ND ND 2.63±0.005

Fermented 2.30±0.03 ND ND 1.24±0.002

Nnerim Fresh 0.60±0.01 ND ND 1.90±0.015

Fermented 1.30±0.01 ND ND 1.32±0.002

Olori Fresh 0.70±0.01 ND 0.19±0.0 0.75±0.005

Fermented 1.60±0.02 ND 0.07±0.0 1.20±0.008

Umudiokpara Fresh 0.60±0.02 ND ND 0.36±0.001

Fermented 0.70±0.01 ND ND 1.34±0.002

Obeakpu Fresh 0.60±0.01 ND ND 2.39±0.010

Fermented 0.40±0.00 ND ND 1.35±0.005

Each value is expressed as mean ± standard deviation (n = 3). ND = Not detected

70

3.17 Dissolved Oxygen (DO) Content in the Fresh and Fermented POME.

In Table 13 below, there were significant increases (p<0.05) observed in the dissolved

oxygen content in the fermented POME from Eziisu and Obeakpu palm oil mills compared to

the dissolved POME in their respective fresh POME. The increase in the dissolved oxygen

content observed in the fermented POME from Nnerim palm oil mill was not significant

(p>0.05) compared to the dissolved oxygen content in its fresh POME. There were significant

decrease (p<0.05) in the dissolved oxygen content in the fermented POME from Amurie and

Umudiokpara palm oil mills compared to the dissolved oxygen in their respective fresh

POME while the decrease observed in the dissolved oxygen content in the fermented POME

from Olori palm oil mill was not significant (p>0.05) compared to the dissolved oxygen

content in its correspondingfresh POME.

71

Table 13: Dissolved Oxygen (DO)Content in the Fresh and Fermented POME

Each value

is expressed as

mean ± standard deviation (n = 3).

Mill Sites Dissolved Oxygen (mg/L) in

Fresh POME

Dissolved Oxygen (mg/L)

in Fermented POME

Amurie 5.77±0.4 4.92±0.4

Eziisu 2.57±0.3 5.74±0.3

Nnerim 3.69±0.3 3.88±0.5

Olori 4.77±0.4 4.65±0.3

Umudiokpara 3.87±0.4 3.16±0.4

Obeakpu 3.17±0.5 4.12±0.4

72

3.18 Biochemical Oxygen Demand (BOD) in the Fresh and Fermented POME

As shown in Table 14, there were significant increases (P<0.05) in the biochemical

oxygen demand in the fermented POME from Olori, Umudiokpara and Obeakpu palm oil

mills compared to the biochemical oxygen demand of their respective fresh POME. However,

there were no significant decreases (p>0.95) in the biochemical oxygen demand in the

fermented POME from Amurie and Eziisu palm oil mills compared to the biochemical

oxygen demand in their respective fresh POME. The fresh and fermented POME from

Amurie palm oil mill had the highest amount of biochemical oxygen demand among all the

POME samples tested from the six different palm oil mills. The fresh POME from

Umudiokpara and fermented POME Nnerim had the least amount of biochemical oxygen

demand among all the POME samples tested from the six different palm oil mills.

73

Table 14: Biochemical Oxygen Demand (BOD) content in the Fresh and Fermented

POME


Mill Sites BOD (mg/L) content in

Fresh POME

BOD (mg/L) content in

Fermented POME

Amurie 365.43±0.0 356.13±7.7

Eziisu 283.03±4.6 280.86±7.4

Nnerim 316.38±5.6 275.22±8.4

Olori 236.82±8.7 312.56±8.9

Umudiokpara 197.86±9.2 289.87±5.8

Obeakpu 255.64±8.4 274.22±9.3

74

3.19 Chemical Oxygen Demand (COD) Content in the Fresh and Fermented POME

As shown in Table 15, there were no significance increases (p>0.05) in the level of

chemical oxygen demand observed in the fermented POME from Amurie and Olori palm oil

mills compared to the chemical oxygen demand in their respective fresh POME. However,

significant increase (p<0.05) in the level of biochemical oxygen demand were observed in the

fermented POME from Nnerim and Umudiokpara palm oil mill compared to the level of the

chemical oxygen demand in their respective fresh POME. Also, there were significant

decreases(p<0.05) in the level of chemical oxygen demand in the fermented POME from

Eziisu and Obeakpu palm oil mills compared to the level of chemical oxygen demand in their

respective fresh POME.

Table 15: Chemical Oxygen Demand (COD) Content in the Fresh and Fermented

POME

Each valu

e is expr

essed as mean ± standard deviation (n = 3).

Mill Sites COD (mg/L) Content in

Fresh POME

COD (mg/L) Content in

Fermented POME

Amurie 3689.54±55 3898.09±49

Eziisu 3466.26±35 2897.36±40

Nnerim 2756.32±36 4569.12±47

Olori 2468.51±46 2664.15±45

Umudiokpara 2422.32±34 3960.07±42

Obeakpu 3783.19±43 2196.18±42

75

3.20 Total Solids in the Fresh and Fermented POME

In Table 16 below, there were significant decreases (p<0.05) in the amount of total

solids in the fermented POME from Eziisu, Nnerim, Umudiokpara and Obeakpu palm oil

mills compared to the amount of total solids in their respective fresh POME. Also, there was

no significant decrease (p>0.05) in the amount of total solids in the fermented POME from

Amurie palm oil mill compared to the total solids in its corresponding fresh POME.

However, it was also observed that there was no significant increase (p>0.05) in the amount

of total solids in the fermented POME from Olori palm oil mill compare to total solids in the

fresh POME.

Table 16:Total Solids in the Fresh and Fermented POME


Mill Sites Total Solids (mg/L) in Fresh

POME

Total Solids (mg/L) in

Fermented POME

Amurie 995.56±6 970.63±8

Eziisu 1110.36±7 962.57±6

Nnerim 987.57±6 898.54±7

Olori 956.15±5 973.38±4

Umudiokpara 1050.35±7 938.33±8

Obeakpu 1050.38±8 880.80±4

76

3.21 Total Volatile Solids Content in the Fresh and Fermented POME

As shown in Table 17, there were no significant increases (p>0.05) in total volatile

solids in the fermented POME from Eziisu, Nnerim, Olori and Umudiokpara palm oil mills

compared to the total volatile solids in their respective fresh POME. However, there were

significant increase (p<0.05) in the total volatile solids in the fermented POME from Amurie

and Obeakpu palm oil mills compared to the total volatile solids in their respective fresh

POME.

Table 17: Total Volatile Solids Content in the Fresh and Fermented POME


Mill Sites Total Volatile Solids

(mg/L)Fresh POME

Total Volatile Solids (mg/L)

Fermented

Amurie 12.25±0.2 14.86±0.3

Eziisu 15.72±0.7 15.81±0.3

Nnerim 15.65±0.6 15.69±0.3

Olori 14.28±0.5 14.39±0.4

Umudiokpara 14.58±0.4 14.93±0.3

Obeakpu 13.38±0.7 15.92±0.5

77

3.22 Suspended Solids in the Fresh and Fermented POME

As shown in Table 18, there were significant increases (p<0.05) in the suspended

solids in the fermented POME from Eziisu and Obeakpu palm oil mills compared to the

suspended solids in their respective fresh POME. There were significant decreases(p<0.05) in

the suspended solids in the fermented POME from Nnerim, olori and Umudiokpara palm oil

mills compared to the suspended solids in their respective fresh POME. Fresh and fermented

POME from Nnerim palm oil mill had the least amount of total suspended solids among all

the fresh and fermented POME from the various mills tested. The fresh POME from Amurie

palm oil mill and fermented POME from Obeakpu palm oil mill had the highest amount of

suspended solids among all the fresh and fermented POME tested.

Table 18: Suspended Solids in the Fresh and Fermented POME


Mill Sites Suspended Solids (mg/L) in

Fresh POME

Suspended Solids (mg/L)

Fermented

Amurie 121.43±3 120.97±3

Eziisu 113.27±3 116.59±3

Nnerim 99.58±3 86.65±3

Olori 116.68±2 106.34±1

Umudiokpara 106.34±5 98.60±3

Obeakpu 103.43±7 125.59±5

78

3.23 Organic Carbon Content in the Soil Contaminated with the Fresh and Fermented

POME

As shown in Fig. 11, there were significant increases (p<0.05) in the organic carbon

content in the soils types fromthe POME dumpsites and those of the soil types ten yards away

from the POME dumpsitescompared to controls in all the palm oil mills except the soil types

from Olori palm oil mill that had no significant (p>0.05) difference fromthat of the controls.

Topsoil from Amurie, Eziisu and Umudiokpara palm oil mills had significant increases

(p<0.05) in their organic carbon content compared to the organic carbon content in their

respective subsoil which was a clear deviation from the trend in the control where subsoil

contained more organic carbon content compared to their respective topsoil . The topsoil and

subsoil from Olori POME dumpsite and ten-yard distance from Olori POME dumpsite had

the least Organic carbon content among all the soil types tested from the six palm oil mills.

Topsoil and subsoil from Umudiokpara POME had the highest organic carbon content among

all the palm oil mills tested.


POME

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

0.5

1

1.5

2

2.5

3

3.5

4

A B

Org

an

ic C

arb

on

(%

)

79


C D E F G

SITES


Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

80

3.24 Organic Matter Content in the Soil Contaminated with the Fresh and Fermented

POME

As shown in Fig. 12 and appendix 6, there were significant increases (p<0.05) in the

organic matter content in the soils types fromthe POME dumpsites and those of the soil types

ten yards away from POME dumpsitescompared to controls in all the palm oil mills except

the soil types from Olori palm oil mill that had no significant (p>0.05) differencefrom that of

the controls. Topsoil from Amurie, Eziisu and Umudiokpara palm oil mills had significant

increases(p<0.05) in their organic matter content compared to the organic matter content in

their respective subsoil which was a clear deviation from the trend in the control where

subsoil contained more organic matter content than their respective topsoil. The topsoil

andsubsoil from Olori POME dumpsite as well as the soil typesten yards away from the Olori

POME dumpsite had the least Organic matter content among all the soil types tested from the

six different palm oil mills. The topsoil and subsoil from Umudiokpara dumpsite palm oil

millhad the highest organic matter content among all the palm oil mills tested.

Fig. 12:Organic Matter Content in the Soil Contaminated with the Fresh and Fermented

POME

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

1

2

3

4

5

6

7

A B

% O

rga

nic

Ma

tter

81

. 12:Organic Matter Content in the Soil Contaminated with the Fresh and Fermented

C D E F G

SITES

. 12:Organic Matter Content in the Soil Contaminated with the Fresh and Fermented

Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

82

3.25 Percentage Content of Oil and Grease in the Fresh and Fermented POME

InTable 19 below, there were significant decreases (p<0.05) in the percentage oil and

grease in the fermented POME compared to the percentage oil and grease of their respective

fresh POME from all the palm oil mills tested. The fresh POME from Amurie, Umuiokpara

and Obeakpu palm oil millshad very high percentage oil and grease compared to those of the

fresh POME from Eziisu, Nnerim and Olori palm oil mills. The fresh POME from Amurie

and Eziisu palm oil mills had the highest and least percentage oil and grease respectively

among all the fresh POME tested. The fermented POME from Umudiokpara and Eziisu palm

oil mills had the highest and least percentage oil and grease respectively among all the

fermented POME tested.

Table 19: Percentage Oil and Grease in the Fresh and Fermented POME


Mill Sites

Percentage Oil and Grease (%)

Fresh POME

Percentage Oil and Grease (%)

Fermented POME

Amurie 12.52±0.62 3.83±0.84

Eziisu 1.09±0.38 0.79±0.10

Nnerim 4.53±0.24 3.79±0.10

Olori 4.59±0.33 3.69±0.97

Umudiokpara 9.68±0.27 6.94±1.03

Obeakpu 9.36±0.78 4.67±0.38

83

3.26 Dehydrogenase activity in the soil contaminated with the fresh and fermented

POME

As shown in Fig. 13, all the soil types from the POME dumpsites as well as soil

typesten yards away from the POME dumpsites showed significant decreases (p<0.05) in the

dehydrogenase activity compared to the dehydrogenase activity of their respective control

soils. The dehydrogenase activities of the microbes at the subsoil were most affected

compared to the dehydrogenase activities of the microbes at the topsoil in all the six different

palm oil mills. Here, topsoil showed higher activities compared to those of their

corresponding subsoil which was a clear deviation from the controls where topsoil had lesser

activity than subsoil. Only the subsoil ten yards away from Amurie and Nnerim palm oil

mills followed the normal trend in the controls.

Fig. 13:Dehydrogenase activity in the soil contaminated with the fresh and fermented POME

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

B: Control

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

A B

Act

ivit

y (

mg

fo

rma

zan

g-1

dry

so

il/9

6h

)

84

ivity in the soil contaminated with the fresh and fermented POME

C D E F GSITES

ivity in the soil contaminated with the fresh and fermented POME

Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

85

3.27 Catalase activity in the soil contaminated with the fresh and fermented POME

As shown in Fig. 14 and appendix 8, there were significant (p<0.05) differences

observed in the catalase activity in the topsoil and subsoil from the dumpsites as well as soil

types ten yardsaway from dumpsites compared to the catalase activities in the control soils.

Topsoil from Amurie, Eziisu, Nnerim and Olori POME dumpsites showed significant

increase (p<0.05) in the catalase activity compare tothe catalase in the controlsoils while

topsoil from Umudiokpara and Obeakpu POME dumpsites showed significant

decreases(p<0.05) in the catalase activity compare tothe catalase activity in the control

topsoil. All the subsoil from the dumpsites and ten yardsaway from the dumpsites showed

significant decreases(p<0.05) in theircatalase activity compared to those of the controlsoils.

However, topsoilten yards away from Eziisu, Nnerim and Umudiokpara POME dumpsites

showed significant increases(p<0.05) in their catalase activity relative to those of the control

topsoil. Topsoil ten yardsfrom Amurie, Olori and Obeakpu POME dumpsites showed

significant decreases(p<0.05) in their catalaseactivity compared to those of the controltopsoil.

Fig. 14:Catalase activity in the soil contaminated with the fresh and fermented POME

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

0.5

1

1.5

2

2.5

A B

Act

ivit

y

(mM

H2O

2/g

dry

so

il /

h)

86


C D E F G

SITES


Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

87

3.28 Lipase activity in the soil contaminated with the fresh and fermented POME

As shown in Fig. 15, there were significant increases (p<0.05) in the lipase activity of

the topsoil from the six dumpsites tested compared to those of the controltopsoil. With the

exception of subsoilfrom OloriPOME dumpsite which had no significant (p>0.05) difference

in its lipase activity compared to the lipase activity of the control subsoil, all the subsoil from

POME dumpsites showed significant decrease (p<0.05) in their lipase activity compared to

those of the controlsubsoil. Lipase activity in the topsoil ten yardsaway from Olori dumpsite

showed no significant (p>0.05) difference compared to the lipase activity of the control

topsoil, while there were significant (p<0.05) difference in the lipase activities in the topsoil

ten yards away from Eziisu, Nnerim, Amurie, Umudiokpara and Obeakpu POME dumpsites.

Only the ten yards away from Nnerim POME dumpsite showed significant (p<0.05) increase

in lipase activity compared to the lipase activity of the control subsoil.

Fig. 15:Lipase activity in the soil contaminated with the fresh and fermented POME.

KEY:

A: Amurie Site

B: Eziisu Site

C: Nnerim Site

D: Olori Site

E: Umudiokpara Site

F: Obeakpu Site

G: Control

0

5

10

15

20

25

30

A B

Lip

ase

act

ivit

y (

mM

/g d

ry s

oil

/18

h)

88


C D E F G

SITES


Dumpsite topsoil

Ten-yards topsoil

Dumpsite subsoil

Ten-yards subsoil

89

CHAPTER FOUR

DISCUSSION

The impact of palm oil mill effluents (POME) on soil fertility was determined by

studying the activities of some soil enzymes and the levels of physicochemical parameters of

POME polluted soil from six dumpsites and soil samples ten yards awayfrom dumpsites in

relation to the control farmland in the area.

The fermented POME from Amurie and Umudiokpara palm oil mills had significantly

higher pH values relative to their respective fresh POME which may be attributed to high

level of free fatty acids in freshPOME as shown by the results of oil and grease

determination. The pH values obtained for POME samples from Eziisu, Nnerim, Obeakpu

and olori palm oil mills showed that fermentation increases the acidity of POME as more

acidic products were produced in the POME during fermentation. The initial pH of POME

ranges from 4.5–5.5(Onyia et al., 2001). This result is similar to the findings of other authors

who reported the pH range of POME to be; 4 – 5 (Rupani et al., 2010), 3.5 – 4.5 (Ma, 1999),

4.0 – 9.0 (Ma, 2000), 6.25 (O’Thong et al., 2007), 5.34 (Awotoye et al., 2011) and 4.7

(Ahmad et al., 2003). The value of pH recorded in this study was however lower i.e. more

acidic than the International Finance Corporation (2007) guideline value (pH 6 – 9) for

effluent from vegetable oil processing. The low pH of the POME indicated that it was acidic

(Hemming, 1977). The acidic nature of POME may have been influenced by organic acids

found in fresh fruit. When the POME was discharged into the soil, it affected nutrient

availability of the nearby plants (Okwute and Isu, 2007), because most plants grow and do

better within a pH range of 6.5 – 7.5 (Hajek et al., 1990).

The pH (KCl) of the control soil showed that the soil within the area studied was

strongly acidic with the pH range of 4.4 for topsoil and 4.3 for subsoil. Subsoil from Eziisu

dumpsite with pH (KCl) 4.0 was the most acidic soil among all the soil samples tested

relative to the control. This low pH may be attributed to the effect of acidic POME recorded

from Eziisu palm oil mill and release of high level of free fatty acids from the degradation of

long chain fatty acids by soil enzymes especially lipase. The higher acidic nature of the soil

samples may not be attributed to effects of POME alone but also tohigher rainfall, as areas of

the world with higher rainfall typically have acidic soil (pH < 4.6), which are too acidic for

most plants.Rainfall affects soil pH because water passing through the soil leaches basic

nutrients such as calcium and magnesium from the soil. They are replaced by acidic elements

such as aluminium and iron. For this reason, soils formed under high rainfall conditions are

90

more acidic than those formed under arid (dry) conditions. The ideal pH range for soil is

from 6.0 to 6.5 because most plant nutrients are in their available state. It is recommended

that when soil test indicates pH below 6.5 as in this case, ground limestone should be applied

to raise the pH level. Thus, soil pH affects the availability of several plant nutrients. A

nutrient must be soluble and remain soluble long enough to successfully travel through the

soil into the roots. The base cations (sodium (Na+), potassium (K+),calcium (Ca+2),and

magnesium (Mg+2) are bound more weakly to the surface soil, especially at low pH.

Therefore, they are less available at low pH. Lower pH generally causes lower cation

exchange capacity (CEC), because the higher concentration of H+ ions in solution will

neutralize the negative charges on organic matter. It also affects the activity of

microorganisms responsible for breaking down organic matter and most chemical

transformations in the soil.

The significant increase (p<0.05) in the total nitrogen content of the fermented POME

from Amurie, Nnerim and Obeakpu may be attributed to non POME sources such as

decaying leaves or debris that on their decomposition may add their nitrogen content to the

fermented POME. However, the significant increase in the nitrogen content of the fresh

POME from Eziisu, Umudiokpara and most especially in Olori could probably be due to the

age and nature of the palm fruits processed. Ohimain et al. (2012) reported the total nitrogen

range of 0.000755 -0.00201%. Other authors have reported values of 0.0064 % (Ma, 2000)

and 0.075 % (Wood et al., 1979). Total nitrogen is consistently found in high concentration

in POME (Ho et al., 1984; Habib et al., 1997 and Muhrizalet al., 2006). International Finance

Corporation (2000), recommended guideline for vegetable oil processing effluent is 0.001 %.

The total nitrogen content of POME soil was found to be significantly higher (p<0.05)

than the non POME soil. Also the higher total nitrogen content in POME soil may be due to

the decomposition of organic constituents and continuous release of nitrogen in the POME

and larger proportion of the nitrogen residing in the subsoil. The higher total nitrogen content

observed in the POME soil in this study correlates with the findings of Wood (1977), Huan

(1987) and Dolmat et al. (1987) who had earlier reported higher nitrogen content in POME

soil.

There were significant decreases (p<0.05) in the phosphorus content observed in both

topsoil and subsoil from various dumpsites which could be attributed to low pH which causes

precipitation reactions with aluminium (Al) and iron (Fe). The tie-up of P in A1-P and Fe-P

minerals under acidic conditions tends to be more permanent than in Ca-P minerals under

normal pH. Thus, low pH has negative effect on nutrient cycling and limits the levels of

91

available phosphorus in acidic soil.However, there were significant increases (p<0.05) in

phosphorus content in soil samples ten yards awayfrom POME dumpsites with subsoil having

significantly higher phosphorus content compared to their corresponding topsoil except in

Amurie where topsoil from Ten yards has the highest amount of phosphorus. The increase in

available phosphorus in POME soil ten yards awayfrom POME dumpsites may have been

borne out of the fact that the discharge of POME into the soil results in a moderate rise in pH,

which may have enhanced the availability of phosphorus (Ayodele and Agbola, 1981; Foth

and Ellis, 1997). Available phosphorus was found to be higher in POME soil than in the non

POME soil. Phosphorus is absorbed by plants in the form of H2PO4- at lower pH and HPO4

2-

at higher pH (Wood, 1977). The soil under the use of POME retains the phosphorus due to

enzymatic actions of the microbial organisms. Here the phosphorus level in the POME soil

was found to be slightly higher than the non POME soil. This agrees with the findings of

Wood (1977) and Huan (1987) which reported that, high organic matter content in POME

contributes to nutrient cycling in POME soil. There is good evidence suggesting that

phosphorus is the dominant element controlling carbon and nitrogen immobilization (Paul

and Clarke, 1989). The increase in the available phosphorus in the POME soil suggests a

possibly high absorption in the soil or a possible precipitation of phosphate (Huan, 1987).

This may be due to the gradual biodegradation of POME, which leads to a delayed effect on

the soil.

The phosphorus content of the fresh POME ranges from 3.90±0.003 – 5.72±0.005

mg/L and in the fermented POME, it ranges from 3.90±0.004 – 6.14±0.003 mg/L. These

concentrationswere far lower than the results of Awotoye et al. (2011) that reported

phosphate value of 165.65mg/L but closer to 5.267 – 8.68 mg/L reported by Ohmain et al.

(2012). In Eziisu, Olori and Obeakpu, the phosphorus content of the fermented POME were

significantly higher than their corresponding fresh POME, but the reverses were the cases in

Amurie, Nnerim and Umudiokpara palm oil mills. There were higher phosphorus contents

recorded in the fresh POME from Amurie, Nnerim and Umudiokpara palm oil mills relative

to their corresponding fermented POME. This may be due to nutrient cycling that occurred in

fermented POME as the fermentation progressed with resultant increase in most nutrients

especially phosphorus.

The calcium levelsin POME was significantly higher (p<0.05) in fresh POME from

Nnerim and Olori palm oil mills than their corresponding fermented POME samples but the

reverse were the cases in the POME samples from Amurie, Umudiokpara and Obeakpu palm

oil mills. The higher calcium levels observed in the fresh POME samples from Nnerim and

92

Olori palm oil mills may be attributed to the nature and sources of water used in the

production process but the increases in the calcium content in the fermented POME from

Amurie, Umudiokpara and Obeakpu palm oil mills may be due to calcium released from

decomposition of organic matter in POME by microbes.

The increase in the magnesium levels observed in the fermented POME from Nnerim

and Umudiokpara palm oil mills may be attributed to the release of magnesium by the

microbial decomposition of organic constituents of POME during fermentation process while

the decreases observed in the magnesium content of fermented POME from Obeakpu palm

oil mill may be due to effect of dilution by addition of more water by rainfall4 and absorption

of some magnesium into the soil.

The significant differences in the levels of sodium in the fresh POME from various

palm oil mills may be due to difference in the ages of palm fruits and nature of the water

used, as salty water could contribute to increase in the sodium level in the POME. On the

other hand the increase in the sodium level in the fermented POME in Amurie, Eziisu and

Obeakpu palm oil mills may be due to the microbial process that occurred during

fermentation. The decreases in the levels of sodium recorded in Nnerim, Olori and

Umudiokpara palm oil mills may be attributed to possible adsorption of particles and

absorption of some fractions by plant roots while fermentation lasted.

The results of potassium content in the fresh and fermented POME from the various

palm oil millswere significantly higher than the potassium content of POME independently

reported by other authors. Wood et al. (1979) and Ohimain et al. (2012) reported the

potassium level in POME as 0.0162 % and 0.0009533 - 0.002914 % respectively which is far

lowerthan 1.37 – 2.56 % and 2.34 – 3.38 % obtained for the fresh and fermented POME

respectively in this study

There were significant increases (p<0.05) in cation exchange capacity (CEC)levelin

all the soil types from various mills with respect to the control. The subsoil from Amurie,

Eziisu and Nnerim POME dumpsites have significantly higher CEC value relative to their

corresponding topsoil as observed in the control which could indicate that enough CEC reside

in subsoil and give subsoil more fertility potential to support plant growth and productivity.

Furthermore, the soil samples ten yardsfrom dumpsites have significant increase (p<0.05) in

CEC than the dumpsites except in Eziisu and Nnerim where subsoil have the highest CEC

content which could be attributed to the higher organic matter in the subsoil most especially

active organic matter that contributed to the soil mineralisation. The increase in CEC in

POME soil is attributed to the addition of organic matter from the effluent (Huan, 1987).

93

However, the high value of CEC in POME soil also agrees with the study by (Oviasogie and

Aghimien, 2002) that attributed the increase in CEC to the increase in the pH as well as the

addition of organic matter from the effluent. The cation exchange capacity (CEC) of a soil is

a measure of the quantity of negatively charged ions on soil surfaces that can retain positively

charged ions (cations) such as calcium, magnesium, sodium and potassium by electrostatic

forces. Cations retained electrostatically are easily exchangeable with cations in the soil

solution so a soil with a higher CEC has a greater capacity to maintain adequate quantities of

Ca2+, Mg2+, Na+ and K+ than a soil with low CEC. A soil with higher CEC may not

necessarily be more fertile because a soil’s CEC can also be occupied by acidic cations such

as hydrogen (H+) and aluminium (Al3+) (Sparks, 1995). However, when combined with other

measures of soil fertility, CEC is a good indicator of soil quality and productivity. A CEC

above 15 meq/100g has a relatively high capacity to hold nutrient cations, which include

Ca+2, Mg+2, K+, NH4+, Cu+2, Fe+2, Mn+2, and Ni+2 (Marschner, 1995).

The exchangeable cations (Ca2+, Mg2+, Na+ and K+) values observed in this study

when considered was significantly higher (P<0.05) in the POME soil than the non-POME

soil. In a similar study (Oviasogie and Aghimien, 2002), had shown that there were an overall

increase in the exchangeable cations of POME soils especially at the area close to the source

of the POME and thus agreed with the observation in this study. The results showed

enrichment of the soils in calcium, magnesium, sodium and potassium due to the application

of POME. Increase in exchangeable cations could be attributed to the increase in the pH

dependent charge as well as the addition of organic matter from the effluent as observed by

Huan, (1987). Lim and P’ng (1983) also recorded increase in potassium, sodium, calcium,

magnesium and organic matter content with the application of POME to soil.

There were significant increases in the calcium content in POME soil compared to

non POME soil. The increase incalcium content in POME soils observed in this study may be

due to the addition of nutrient released from slow decomposition of organic constituent of

POME to soil and it also, may be due to the presence of inorganic salt in the POME.

There were significant increases (p<0.05) in the potassium content in the topsoil from

POME dumpsites relative to the control, with the topsoil from Umudiokpara dumpsite having

the highest increase in potassium content. With exception of ten yards from Eziisu dumpsite

and subsoil from Olori and Obeakpu, there were significant increases in the potassium

content of the soil samples from ten yards compared to the control. These increases in

potassium content of POME soil observed in this study may be attributed to use of hard water

and salty water in the palm oil processing, wearing of engine parts used in the processing

94

palm fruits and also due to potassium being a natural constituent of the POME.Potassium

deficiency is most commonly associated with sandy soils. The optimallevelof potassium will

vary with crop, yield, and soil type. It is an immobile nutrient like phosphorus and levels

change slowly.

There was significant increase(p<0.05) in the sodium content of the topsoil from

Amurie, Eziisu, Nnerim and Olori POME dumpsites relative to control but no significant

(p>0.05) difference in both topsoil and subsoil from Umudiokpara and control and also

between topsoil from Obeakpu and control. There were also significant decreases in the

sodium content in the subsoil from Umudiokpara and Obeakpu POME dumpsites compared

to sodium content in the control which may be due to negative effect of acidic pH on the

absorption of sodium. Topsoil and subsoil from Obeakputen yards away fromPOME

dumpsite have significant amount of sodium against the sodium content in the topsoil and

subsoilfrom its dumpsite which could be as a result of erosion washing away much of the

sodium ions from the dumpsites to nearby areas.Sodium ionsare retained in the soil as an

easily exchangeable cation, which at elevated levels can detrimentally influence the pH of

soil solution, nutrient balances, soil infiltration and permeability, and soil solution salinity

levels. Sodium is not considered an essential plant nutrient and consequently a very low value

or range is desired.

Topsoil from Amurie, Eziisu and Obeakputen yards awayfrom POME dumpsites had

the highest magnesium content while topsoil from Nnerim, Olori and Umudiokpara

dumpsites have very significant increase in the magnesium content compared to the control

which may be due to released nutrient and salts from the POME. The deviation observed in

Amurie, Eziisu and Obeakpu where topsoil Ten yards from dumpsites had highest

magnesium content could probably be due to effect of leaching or washing of nutrients away

by erosion to nearby areas.Some of the total soil magnesium is found in non-exchangeable

form and thus, the exchangeable magnesium level changes slowly with time because of

equilibrium with minerals.

Although Wood et al. (1979) reported 0.001mg/L cadmium in POME, the levels of

copper detected in Amurie, Eziisu, Nnerim and Olori POME samples were higher than

0.09mg/L and 0.89mg/L reported respectively by Wood et al. (1979) and Ma (2000). The

iron and copper levels are significantly (P<0.05) different among the various mills. Wood et

al. (1979) and Ma (2000) reported the level of iron in POME as 11 and 46.6mg/L

respectively. The chromium level is significantly (P<0.05) different between fresh and

fermented POME from Olori while there was no significant (p>0.05) difference between

95

fresh and fermented POME from Amurie. Wood et al. (1979) found from their study that the

level of chromium in POME is 0.01mg/L. The concentration range of the heavy metal

indicates lack of uniform distribution of heavy metals in POME; however same variations of

this magnitude have also been reported by Obodo (2002). The variations observed were

probably due to various factors such as trace metal contents of the crops, contamination from

the engine during digestion process (Abulude et al., 2007). The degree of hardness of the

water used in processing might affect the dissolution of heavy metals (Adeyeye and Ayejuyo,

2002). The results of this study found that the POME had high potential to pollute the natural

waterways and farmlands. Increased heavy metal content negatively affects soil microbial

population which may have direct negative effect on soil fertility (Ahmad et al., 2005).

Environmental pressure resulting from the contamination may reduce the biodiversity of

microorganisms and disturb the ecological balance. However, there are reports that soil

microorganisms may adapt to the increased, even toxic heavy metals and other xenobiotics

concentration in soil (Kozdroj, 1995) by developing various mechanisms to resist heavy

metal contamination (Rathnayake et al., 2010).

The dissolved oxygen (DO) in the POME samples from the various mills were in the

range of 2.57±0.3– 5.77±0.4mg/L in fresh POME and 3.16±0.4 – 5.74±0.3mg/L in fermented

POME, being significantly different (p<0.05) between fresh and fermented POME samples

among the six mills tested. Fresh POME from Amurie, Olori and Umudiokpara mills had

significantly (p<0.05) higher DO content relative to their respective fermented POME,

however the reverse were the cases in the POME from Eziisu, Nnerim and Obeakpu mills.

Awotoye et al. (2011) reported that the dissolved oxygen of raw POME is 1.250mg/L. The

relatively high DO reported in this study may be due to the high temperature and duration of

bright sunlight, which influenced the percentage of soluble gases (O2 and CO2) in the effluent

(Manjare et al., 2010). DO is an important parameter in POME quality assessment and

reflects the physical and biological processes prevailing in the POME, it indicates the degree

of pollution in water bodies (Murhekar, 2011).

The biochemical oxygen demand (BOD) obtained from the mill effluents were in the

range of 197.86±9.2 – 365.43±0.0mg/L in the fresh POME and 275.22±8.4 –

356.13±7.7mg/L in the fermented POME, having no significant (p>0.05) differences between

fresh and fermented POME, except in Nnerim and Obeakpu mills that showed significant

(p<0.05) different between the fresh and fermented POME. Fresh and fermented POME from

Amurie had the highest amount of BOD among all the palm oil mills which probably could

be attributed to POME from Amurie mill having the largest amount of biodegradable organic

96

matter, oil and grease. Prasertsan and Prasertsan (1996), Ma (2000), Wood et al., (1979) and

Awotoye et al. (2011) reported BOD of POME to be 50000, 25000, 2080 and 123.675mg/L

in their independent studies respectively. The BOD values recorded in this study were

significantly lower (p<0.05) than that those of other authors, but higher than that reported by

Awotoye et al. (2011) and the International Finance Corporation (2007) standard value of

50mg/L for vegetable oil processing effluents. Low concentration of BOD shows that the

effluent has less organic matter. This means that the microorganisms will consume less

oxygen to decompose the organic matter in the effluent. This difference in BOD values may

be due to variation of different batches, day and factories, depending on the processing

techniques and age or type of fruits. Some researchers believed it also may be due to

variation in the discharge limit of the factory, climate and condition of palm oil processing

Awotoye et al. (2011).

The chemical oxygen demand (COD) obtained in thefresh and fermented POME

samples were in the range of 2196.18±42– 4569.12±47 mg/Lin the fresh POME and

2422.32±34 – 3783.19±43 mg/Lin the fermented POME. Ma (2000), Wood et al., (1979) and

Awotoye et al. (2011) presented the COD of POME as 50000, 10250 – 43,750, 5790 and

284.875mg/L in their various studies respectively. The values of COD recorded in this study

were close to the results of other authors but several orders higher than the International

Finance Corporation (2007) standard value of 250 mg/L for effluent from vegetable oil

processing. In comparison of the measured BOD value with the COD value, thelarge

difference observed in this study indicates that the organic constituents cannot be easily

broken down. The BOD detects only the destructible proportion of organic substances and as

a general principle is therefore lower than the COD value, which also includes inorganic

materials and those materials which cannot be biologically oxidized.

The present value of total solid ranges from 956.15±5.0 – 1110.36±7.0 mg/Lin fresh

POME and 880.80±4.0 – 973.38±4.0 mg/Lin the fermented POME which were far below the

range of previous study that came up with their findings that the total solid in POME were

between 37900 and 45 000 mg/L by Wood, et al. (1979) Wong, et al. (2009) and MPOB

(2004), respectively. The decreases recorded in the amount of the total solid after

fermentation is a clear indication that fermentation can help in the biodegradation of the

solids present in POME which may lead to increased nutrients in POME after fermentation.

The low range of total solid recorded in this study may be attributed to the net result of the

fusion of traditional and mechanized methods in the production of palm oil from the palm

fruits in the area of this study.

97

The volatile solidsgave a rough approximation of the amount of organic matter

present in the solid fraction of wastewater, activated sludge and industrial wastes. The present

range of volatile solid is 12.25±0.2 – 15.72±0.7 mg/Lin the fresh POME and 14.39±0.4 –

15.92±0.5 mg/L in the fermented POME. The range are far below the recorded data by

Wood, et al. (1979) and Wong, et al. (2009) which was in the range of 27300 mg/L to 30150

mg/L. The increase in volatile solid observed after fermentation may also be due to addition

of volatile solids to the POME during fermentation from external sources and also it may be

due to release of some volatile solids from breakdown of organic constituents of POME.

The present value of suspended solids were in therange of 99.58±3.0 – 121.43±3.0

mg/Lin fresh POME and 86.65±3.0 – 125.59±5.0 mg/Lin fermented POME which were far

below 18000 mg/L value recorded by Ahmad et al., (2003) and MPOB (2004). The higher

suspended solid of 25800 mg/L has been reported by Wu, et al. (2007). There were

significant (p<0.05) decreases in suspended solids in the fermented POME from Nnerim,

Olori and Umudiokpara palm oil mills relative to their respective fresh POME which may be

attributed to the decomposition of some suspended solids by fermentation process and with

time. The increasein the suspended solid in the fermented POME from Obeakpupalm oil mill

may be due to addition of other suspended solids to the POME from non POME sources

during the fermentation process since open fermentation was carried out.

The significant increase (p<0.05) in organic carbon(OC) and organic

matter(OM)observed in the various soil types ten yards awayfrom Amurie, Eziisu and

Nnerim dumpsites could be attributed to leaching or washing away of organic constituents of

POME by erosion while the increase in OC and OM content in topsoil from Amurie, Eziisu

and Obeakpu dumpsites could be attributed to abundant deposit of POME at the topsoil and

slow decomposition of POME constituents to the subsoil as total solids and suspended solids

in the raw and fresh and fermented POME hinder their movement in the subsoil. The higher

OC and OM observed at Umudiokpara dumpsite indicate increased accumulation of OC and

OM with increased dumping of POME at the dumpsite. Organic matter content strongly

affects the soil fertility by increasing the availability of plant nutrients, improving the soil

structure and the water holding capacity and also acting as an accumulation phase for toxic,

heavy metals in the soil environment (Deiana et al., 1990). For this reason, the recycling of

organic wastes through their application to the soil can be an important promising practice for

agricultural activities.The accumulation of OC in POME soil couldbe attributed to the

constituents of fresh and fermented POME which slow down the decomposition of organic

matter in POME under water-saturated conditions and low temperature (Batjes, 1996;

98

Okwute and Isu, 2007). This implies that POME increases the level of organic matter in the

soil, as such if properly treated and applied on farmlands can be a promising agricultural

practice to augment the soil to improve its fertility. The higher OC and increased content of

OM obtained for POME soil in this study agrees with the findings of Dolmat et al. (1987);

Shamshuddin et al. (1995), when they attributed the increase on the organic constituents of

POME.

The higher percentage oil and grease recorded in fresh POME from Amurie,

Umudiokpara, Obeakpu, Olori and Nnerim palm oil mills showed that these palm oil mills

have poor oil recovery techniques compared to Eziisu palm oil mill. Thus, they have higher

pollution potential to contaminate farmlands where their POMEwas discharged without

proper treatment.These results were significantly higher than the 0.6 – 0.7 % reported by Ma

(2000); Onyia et al. (2001); Ahmad et al. (2005). Discharge of fresh and fermented POME

with higher content of oil and grease will probably affect soil micro-organisms negatively

due to reduction of oxygen level in the soil thereby hindering microbial respiration and

decomposition of organic matter. This will also invariably lead to induction of Lipase activity

in the soil as amount of hydrocarbon content of the soil increased with increased deposition

of oil and grease.

The significant decrease in dehydrogenase activity may be attributed to the chronic

exposure of the soil microorganisms to palm oil effluent which exposed the organisms’

dehydrogenase systems to more work, in the oxidation of organic matter contained in the

effluent-polluted soil. The decrease in dehydrogenase activity recorded in the topsoil from

POME dumpsites in: Amurie, Eziisu, Nnerim, Umudiokpara and Obeakpu palm oil mills

compare to their respective subsoil could be probably due to topsoil in dumpsites from these

mills having more viable microorganisms as a result of adaptation to polluted environment,

abundant supply of oxygen and availability of stabilized enzyme at the topsoil compare to

subsoil. This result agrees with the reports of Garcia – Gil et al. (2000), who worked on long

term effects of municipal solid waste compost application on soil enzyme activities and

microbial biomass, and Drucker et al. (1979); Kizilkaya et al. (2004) who also worked on the

effect of heavy metal contamination on soil microbial biomass and activity.The

dehydrogenase systems apparently fulfil a significant role in the oxidation of organic matter,

as they transfer electron from substrates to acceptors. Soil dehydrogenase increases the

reaction rate at which plant residues decompose and release plant available nutrients.

Therefore, internal and external factors that cause decreases in soil dehydrogenase activity

will translate to decrease in available plant nutrients (decrease in soil fertility) and poor

99

productivity. The most widely used substrate for dehydrogenase assay is 2, 3, 5- triphenyl

tetrazolium chloride (TTC), which produces red coloured water insoluble tripheny formazan.

The apparent redox potential of TTC is about -0.08v, which makes it act as an acceptor for

many dehydrogenases. The use of total dehydrogenase assay is recognized as a useful

indicator of the overall measure of microbial metabolism (Tabatabai, 1982).

The results obtained for catalase activity in this study showed that chronic palm oil

effluent pollution altered soil catalase activity.Chronic exposure of soil to palm oil effluent

brings changes in soil conditions such as pH, hypoxia, heavy metals as well as reduction in

the number and activities of soil micro-organisms (Maila and Cloete, 2005). These changes

make the soil which was previously fertile before exposure to the effluents pollution to lose

its productive potentials (Maila and Cloete, 2005). Soil enzyme inhibition depends on the

nature and concentration of heavy metals such as chromium, iron and copper found in this

study. Heavy metals could have inhibited catalase enzymatic activities by denaturing the

enzyme protein architecture and interacting with its active sites (Moreno et al., 2002). Heavy

metals can also influence microbial community, which ultimately lead to changes in soil

catalase enzymatic activities (Kandeler et al., 2000; Moreno et al., 2002). When soil enzyme

activity is negatively affected by effluent discharges, plants, man and other animals that

depend directly and indirectly on it are also adversely affected (Osuji and Nwoye, 2007).

Environmental sustainability depends largely on a sustainable soil ecosystem (Adriano et al.,

1998). When soil is polluted, the physiochemical properties are affected which may decrease

its productive potentials (Osuji and Nwoye, 2007).

The significant increase in lipase activity could be attributed to the chronic exposure

of the soil microorganisms to palm oil mill effluent which exposed the organisms’ lipase

systems to more work, in the biodegradation of hydrocarbon chains contained in the effluent-

polluted soil. The increase in lipase activity with increasing oil and grease mostly in the

various dumpsites demonstrates the induction of lipase activity by the oily contaminants. The

higher lipase activity recorded in both topsoil and subsoil in Nnerim dumpsites and ten-yard

compared to Amurie dumpsite that has highest percentage oil and grease indicates the

presence of other hydrocarbon source in the area serving as inducer to the lipase activity.

With the exception of subsoil from Olori and Obeakpu dumpsites which have higher lipase

activities relative to their respective topsoil, which is an indication of the subsoil having more

hydrocarbon than their topsoil, topsoil from Amurie, Eziisu, Nnerim and Umudiokpara

dumpsites had the highest lipase activity in all the dumpsites tested. This was probably due to

more oil and grease contamination. Lipase activity was found to be the most useful indicator

100

parameter for testing hydrocarbon degradation in soil (Margesin, 1999). Lipases are

ubiquitous enzymes which catalyze the hydrolysis of triacylglycerols (the main component of

natural oil or fat) to glycerol and free fatty acids.

4.2 Conclusion

Although POME has high levels of soil nutrients which could improve soil fertility

and crop yield, its high levels of acidity, BOD, COD, oil and grease negatively affected soil

quality, nutrient availability and solubility.Due to the deleterious effects of POME on land

and water, it is preferable to recycle it rather than discharging directly into the soil.

Consequently, those involved in palm oil processing using oil mills, should be enlightened on

the danger inherent in indiscriminate discharge of the fresh and fermented POME on our

agricultural soil, as it adversely affects soil fertility and productivity. Effective enlightenment

can be achieved through sustained dissemination of information in this regard, using the

media and trained agricultural officers.

The findings of this research work has shown that poor processing and oil recovery

techniques employed in the various palm oil mills constitute large economic waste as much

of the palm oil were lost to the environment. Fermentation process cannot effectively

detoxify POME to make it suitable for discharge on farm lands and water bodies despite

reducing the percentage of oil and grease in the fermented POME.

4.3 Suggestions for Further Studies

The impact of palm oil mill effluents on soil fertility was investigated in this study and it

is therefore suggested that further studies should be done on the following:

• The effect of palm oil mill effluents on soil aerobic population and soil water holding

capacity.

• Types of crops best grown on farmlands where palm oil mill effluents are discharged

and the impact on the nutritional composition of the crops.

• Possibilities of using palm oil mill effluents in the production of organic fertilizer.

• Processing and recycling methods best suitable in reducing the toxicity and pollution

levels in palm oil mill effluents.

• Water samples from the various communities should be tested for the presence of

heavy metals and cations such as sodium, magnesium, iron and calcium.

•

101

REFERENCES

Abd-Rahman, R., Mohd, K.S. and Abu, Z.Y. (2003). Composting palm oil mill sludge -sawdust: Effect of different amount of sawdust. Proceedings of National workshop inconjunction with ARRPET workshop on Wastewater treatment and recycling 2: Removal chloroorganics and heavy metals. ISBN 983-2982-04-9.

Abdurahman, N.H., Rosli, Y.M., Azhari, N.H. and Tam, S.F. (2011).Biomethanation of Palm Oil Mill Effluent (POME) by Membrane Anaerobic System (MAS) using POME as a Substrate.World Academy of Science, Engineering and Technology, 51.

Abdul Latif, A., Suzylawati, A. I., Norliza I. and Subhash, B. (2003a). Removal of suspended solids and residual oil from palm oil mill effluent. Journal of Chemical Technology

and Biotechnology, 78(9):971-978.

Abdul Latif, A., Suzylawati, I., Norliza, I. and Subhash, B. (2003b). Water recycling from palm oil mill effluent using membrane technology. Desalination, 157: 87-95.

Abulude, F.O., Obidiran, G.O. and Orungbemi, S. (2007). Determination of physico-chemical parameter and trace metal contents of drinking water samples in Akure Nigeria. Electronic Journal of Environmental, Agricultural and Food Chemistry, 6(8):2297-2303.

Ademoroti, C.M.A (1996). Standard Methods for Water and Effluent Analysis. Published by Foludex Press, Ibadan. pp. 11-58

Ademoroti, C.M.A. (2006). Standard Method for Water and Effluents Analysis. 1st Edition. Foludex Press Limited, Ibadan. pp. 3-39

Adeyeye, E.I. and Ayejuyo, O.O. (2002). Assessment of the physicochemical status of a textile industry’s effluent and its environment. Pakistan Journal of Scientific

and Industrial Research,45:10-16.

Adriano, O.C., Chopecka, A. and Kaplan, K.I. (1998). Role of soil chemistry in soil remediation and ecosystem conservation. American Journal of Soil Science, 12: 361-386.

Agbenin, J.O. (1995). Laboratory Manual for Soil and Plant Analysis (Selected Methods and Data Analysis). Faculty of Agriculture/Institute of Agricultural Research, A.B.U. Zaria. pp. 7-71.

Ahmad, A.L., Ismail, S. and Bhatia, S. (2003). Water recycling from palm oil mill effluent (POME) using membrane technology. Desalination,157:87-95.

Ahmad, A.L., Sumathi, S. and Hameed, B.H. (2005). Adsorption of residue oil from palm oil mill effluent using powder and flake chitosan: Equilibrium and kinetic studies. Water

Resources,39: 2483-2494.

Ahmad, A.L., Chan, C.Y., AbdShukor, S.R. and Mashitah, M.D. (2008). Recovery of oil and carotenes from palm oil mill effluent (POME). Chemical Engineering Journal, 141: 383-386.

102

Akmal, M., Wang, H.Z. and Wu, J.J. (2005). Changes in enzymes activity, substrate utilization pattern and diversity of soil microbial communities under cadmium pollution.Journal of Environmental Science, 17: 802-807.

Alam, M.Z., Jamal, P. and Nadzir, M.M. (2008a). Bioconversion of palm oil mill effluent for citric acid production: Statistical optimization of fermentation media and time by central composite design. World Journal of Microbiological Biotechnology, 24: 1177-1185.

Alam, M.Z., Kabbashi, N.A. and Zain, K.H.M. (2008b). Optimization of Media Composition for Bioethanol Production by Direct Bioconversion of Palm Oil Mill Effluent, Proceedings of International Conference on Environmental Research and Technology. pp. 952-956.

Alef, K. and Nannipieri, P. (1995). Catalase Activity. In: Methods in Applied Soil Microbiology and Biochemistry. London: Academic Press. pp. 362-363.

Alias, Z. and Tan, I.K.P. (2005). Isolation of palm oil-utilising, polyhydroxyalkanoate producing bacteria by an enrichment technique. Bioresources Technology, 96:1229-1234.

Alrawi, R.A., Ab-Rahman, N.N.N., Ahmad, A., Ismail, N. and Mohd Omar, A.K. (2012). Characterization of oily and non-oily natural sediments in palm oil mill effluent. Journal of Chemistry, 2013:1-12

APHA. (1998). Standard Methods for Examination of water and wastewater. 19th Edition American Public Health Association, American Water Works Association and Water Pollution Control Federation, Washington, D.C. pp. 85-135

Arcadio, P. and Gregoria, A. (2003). Physical and Chemical Treatment of Water and

Wastewater. Sincero, A.P. and Sincero, G.A. U.K. pp. 320: 235.

Awotoye, O.O., Dada, A.C. and Arawomo, G.A.O. (2011). Impact of palm oil processing effluent discharging on the quality of receiving soil and rivers in south western Nigeria. Journal of Applied Sciences Research, 7(2): 111-118.

Ayodele, O.J. and Agbola, A.A. (1981). The relationship between Bray P-1, modified NaHNO3, new Mehlich and NH4F. HF extractants for phosphorus in savannah soils of western Nigeria. Soil Society of American Journal, 45:462-464.

Baharuddin, A.S., Wakisaka, M., Shirai, Y., Abd Aziz, S., Abdul Rahman, N.A. and Hassan, M.A. (2009). Composting of empty fruit bunches and partially treated palm oil mill effluents in pilot scale. InternationalJournal of Agricultural Resources, 4(2): 69-78.

Baldock, J.A. and Skjemstad, J.O. (1999). Soil Organic Carbon/Soil Organic Matter. In 'Soil Analysis: An Interpretation Manual.Peverill, K.I., Sparrow, L.A. and Reuter, D.J. (Eds).pp. 159-170.

Batjes, N.H. (1996). Total carbon and nitrogen in the soils of the world. European Journal of

Soil Science, 47(2): 151-163.

103

Binder, D.L., Dobermann, A., Sander, D.H. and Cassman, K.G. (2002). Biosolids as nitrogensource for irrigated maize and rainfed sorghum. Soil Science Society of

AmericanJournal, 66:531-543.

Brady and Nyle, (1984). The Nature and Properties of Soils (9th Edition). Macmillan Publishing Co. USA. pp. 47-53

Brookes, P.C. and McGrath, S.P. (1984). Effects of metal toxicity on the size of the soil microbial biomass. Journal of Soil Science, 35:341-346.

Cambardella, C.A. and Elliott, E.T. (1993). Methods for physical separation and characterization of soil organic matter fractions. Geoderma,56:449-457.

Cameron, K.C., Rate, A.W., Noonan, M.J., Moore, S., Smith, N.P. and Kerr, L.E. (1996). Lysimeter study of the fate of nutrients following subsurface injection and surface application of dairy pond sludge to pasture. Agriculture, Ecosystems and

Environment, 58:187-197.

Capasso, R., Cristtinzio, G., Evidente, A. and Scognamiglio, F. (1992). Isolation, spectroscopy and selective phytotoxic effects of polyphenols from vegetable wastewaters. Journal ofPhytochemistry, 31:4125-4128.

Cervantes, C., Campos-Garcia, J., Devars, S., Guatierrez-Corrora, F., Loza-Tavera, M. and Torres-Guzman J.C. (2001). Interactions of chromium with microorganisms and plants. FEMS Microbiology Reviews, 25:335-347.

Chander, K. and Brookes, P.C. (1991). Effects of heavy metals from past applications of sewage sludge on microbial biomass and organic matter accumulation in a sandy loam soil and silt loam UK soil. Journal ofSoil Biology and Biochemistry,23:927-932

Chander, K., Goyal, S., Mundra, M.C. and Kapoor, K.K. (1977). Organic matter, microbial biomass and enzyme activity of soils under different crop rotations in the tropics.Journal ofBiology and Fertility of Soils,24: 306-312.

Chavalparit, O., Rulkens, W.H., Mol, A.P.J. and Khaodhair, S. (2006). Options for environmental sustainability of the crude palm oil industry in Thailand through enhancement of industrial ecosystems.Journal ofEnvironmental Development and

Sustainability, 8:271-287.

Cheng, C.K., Hani H.H. and Ismail K.S.K. (2007). Production of bioethanol from oil palm empty fruit bunches. International Conference on Sustainable Materials,1: 69-72.

Chong, M., Abdul-Rahman, N.A., Abdul Rahim, R., Abdul Aziz, S., Shirai, Y. and Hassan, M.A. (2009). Optimization of biohydrogen production by clostridium butyricum EB6 from palm oil mill effluent using response surface methodology. International

Journal of Hydrogen Energy, 34:7475-7482.

Choorit, W. and Wisarnwan, P. (2007). Effect of temperature on the anaerobic digestion of palm oil mill effluents. Electronic Journal of Biotechnology, 10(3): 376-385.

104

De Ridder, N. and Van Keulen, H. (1990). Some aspects of the role of organic matter in sustainable intensified arable farming systems in the West African semi-arid tropics. Journal ofFertilizer Research, 26:299-310.

Deiana, S., Gessa, C., Manunza, B., Rausa, R. and Sebeer, R. (1990). Analytical and spectroscopic characterization of humic acids extracted from sewage sludge, manure and worm compost. Journal of Soil Science, 150(1):419-426.

Department of Environment Malaysia (1999). Industrial Processes and the Environment. Crude Palm Oil Industry Handbook.pp.5-10.

Dick, R.P. (1997). Soil Enzyme Activities as Integrative Indicators of Soil Health. Biological Indicators of Soil Health. Cab International, New York.pp. 121-156.

Dick, W.A. (1984). Influence of long-term tillage and crop rotation combinations on soil enzyme activities. Soil Science Society of American Journal,48(3): 569-575.

Dick, W.A. and Tabatabai, M.A. (1992). Significance and Potential Uses of Soil Enzymes. In: Blain, F.J (Ed), Soil Microbial Ecology Application in Agric and EnvironmentalManagement. Marcel Dekker, New York.pp. 95-127.

Dobermann, A. and Cassman, K.G. (2002). Plant nutrient management for enhanced productivity in intensive grain production systems of the United States and Asia. Journal ofPlant and Soil Science, 247:153-175.

Doelman, P. and Haanstra, L. (1984). Short-term and long-term effects of cadmium, chromium, copper, nickel, lead and zinc on soil microbial respiration in relation to abiotic soil factors. Journal of Plant and Soil Science,79: 317-327.

Dolmat, M.T., Lim, K.H., Zakaria, Z.Z. and Hassan, A.H. (1987). Recent studies on the effects of land application of palm oil mill effluent on oil palm and the environment. In: Proceedings of international oil palm, palm oil conferences. Pakistan Journal of

Science and Industrial Research, 2:596-604.

Doran, J.W. and Parkin, T.B. (1994). Defining soil quality for a sustainable environment. Soil Science Society of America Incorporated and the American Society of Agronomy Incorporated, Special Publication. No. 35, pp.3-21.

Drucker, H., Garland, T.R. and Wildung, R.E. (1979). Metabolic Response of Microbiota to

Chromium and other Metals. Trace metals in health and disease.pp. 1-25.

Drysdale, C.R. and McKay, M.H. (1995). Citric acid production by Aspergillus niger on surface culture on inulin. Letter of Application of Microbiology, 20:252-254.

Dushoff, I.M., Payne, J., Hershey, F.B. and Donaldson, R.C. (1965). Oxygen uptake and tetrazolium reduction during skin cycle of mouse. American Journal of Physiology, 209: 231-235.

Food and Agriculture Organization (FAO). (1998). World reference base for soils. World Soil Research, Report, 84.

105

Foth, H.D. and Ellis, B.G. (1997). Soil Fertility. 2nd Ed. CRC Press. Boca Raton, Florida.pp. 290-294

Franzluebbers, A.J. (2002). Soil organic matter stratification ratio as an indicator of soil quality. Journal ofSoil and Tillage Research,66:95-106.

Garcia-Gil, J.C., Plaza, C., Soler-Rovira, P. and Polo, A. (2000). Long term effects of municipal solid waste compost application on soil enzyme activities and microbial biomass. Journal ofSoil Biology and Biochemistry,32:1907-1913.

Gianfreda, L. and Bollag, J.M. (1996). Influence of natural and anthropogenic factors on enzyme activity in soil. Journal of Soil Biochemistry,9:123-193.

Grandy, A.S. and Robertson, G.P. (2007). Land-use intensity effects on soil organic carbon accumulation rates and mechanisms. Journal ofEcosystems, 10:58-73.

Grewal, H.S. and Karla, A.L. (1995). Fungal production of citric acid. Journal of Advance

Biotechnology, 13: 209-234.

Gupta, S. (1994). Continuous production of citric acid from sugar cane molasses using acombination of submerged immobilized and surface stabilized cultures of AspergillusnigerKCU520. Biotechnology Letter, 16:599-604.

Habib, M.A.B., Yusoff, S.M., Phang, K.J. and Mohamed, S. (1997). Nutritional values of chironomid larvae grown in palm oil mill effluent and algalculture. Journal

ofAquaculture,158: 95-105.

Hajek, B.F., Karlen, D.L., Lowery, B., Power, J.F., Schumaker, T.E., Skidmore, E.L. and Sojka, T. (1990). Erosion and Soil Properties In: W.E. Larson, G.R. Foster, R.R. Allmaras and G.M. Smith (Eds.) Research Issues in Soil Erosion Productivity. pp. 23-40.

Hassan, M.A., Shirai, Y., Kusubayashi, N., Abdulkarim, M.I., Nakanishi,K. and Hashimoto, K. (1996). Effect of organic acid profiles during anaerobic treatment of palm oil mill effluent on the production of polyhydroxyalkanoates by rhodobactersphaeroides. Journal of Fermentation and Bioengineering, 82(2):151-156.

Hassan, F., Shah, A.A. and Hameed, A. (2006). Industrial applications of microbial lipases. Journal of Enzyme Microbial Technology, 39: 235-251.

Havlin, J.L., Kissel, D.E, Maddux, L.D., Claassen, M.M. and Long, J.H. (2005). Crop rotation and tillage effects on soil organic carbon and nitrogen. Soil Science Society of

American Journal,54:448-452.

Hemming, M.L. (1977). A viable solution to the palm oil effluent problem. In: Proceedings of the Malaysian International Symposium on Palm Oil Processing and Marketing.Earp, D.A. and Newall, W. (Eds.) Kuala Lumpur, 17-19th,pp. 79-95.

Ho, C.C., Tan, Y.K. and Wang, C.W. (1984). The distribution of chemical constituents between the soluble and the particulate fractions of palm oil mill effluent and its significance on its utilization/treatment. Journal of Agricultural Wastes, 11: 61-71.

106

Huan, K.L. (1987). Trials on long-term effects of application of POME on soil properties, oil palm nutrition and yields. In: Proceedings of the international oil palm, palm oil conferences. Dr. Abdul Halim,H.B., Hassan, C.P.S., Woond, B.J. and Dr. PushParajah,E.(Eds.) industry’s effluent and its environment. Pakinstan Journal of

Science and Industrial resources, 45:10-16.

International Finance Corporation (IFC). (2007). Environment, Health, and Safety Guidelines for Vegetable Oil Processing. World Bank Group. pp. 7-11.

International Institute for Tropical Agriculture (I.I.T.A). (1979). Selected Methods for Soil

and Plant Analysis, Manual Series. I.I.T.A. Ibadan.pp. 4-12.

Irha, N., Slet, J. and Petersell, V. (2003). Effect of heavy metals and polyhydroxyalkanoate on soil accessed via dehydrogenase assay. Journal ofEnvironment International, 28:779-782.

Jamal, P., Alam, M.Z. and Mohamad, A. (2007). Microbial Bioconversion of Palm Oil Mill Effluent to Citric Acid with Optimum Process Conditions, In: Ibrahim, F., N.A. Abu Osman, J. Usman and N.A. Kadri (Eds.): Biomed 06, IFMBE Proceedings, 15:483-487.

Jordan, M.J. and LeChevalier, M.P. (1975). Effects of zinc-smelter emissions on forest soil microflora. Canadian Journal of Microbiology,21:1855-1865.

Kalembasa, S.J. and Jenkinson, D.S. (1973). Soil Chemical Analysis. Prentice-Hall Inc., New York.pp. 23-25.

Kandeler, E., Kampichler, C. and Horak, O. (1996). Influence of heavy metals on the functional diversity of soil microbial communities. Journal ofBiology and Fertility of

Soils, 23: 299-306.

Kandeler, E., Tscherko, D., Bruce, K.D., Stemmer, M., Hobbs, P.J. and Bardget, R.D. (2000). Structure and function of the soil microbial community in microhabitats of a heavy metal polluted soil. Journal ofBiology and Fertility of Soils, 32: 390-400.

Kathiravale, S. and Ripin, A. (1997). Palm oil mill effluent treatment towards zero discharge. A paper presented at National Science and Technology Conference, Kuala Lampur,Malaysia. Report.3:1-8.

Kızılkaya, R., Aşkın, T., Bayrakl, B. and Sağlam, M. (2004). Microbiological characteristics of soils contaminated with heavy metals. European Journal of Soil Biology, 40:95-102.

Klose, S., Moore, J.M. and Tabatabai, M.A. (1999). Arylsulfatase activity of microbial biomass in soils as affected by cropping systems. Journal ofBiology and Fertility of

Soils, 29:46-54.

Konopka, A., Zakharova, T., Bischoff, M., Oliver, L., Nakatsu, C. and Turco, R.F. (1999). Microbial biomass and activity in lead-contaminated soil. Journal ofApplied and

Environmental Microbiology,65:2256-2259

107

Kozdrój, J. (1995). Microbial responses to single or successive soil contamination with Cd or Cu. Journal ofSoil Biology and Biochemistry, 27: 1459 1465.

Krull, E.S., Baldock, J.A., and Skjemstad, J.O. (2003). Importance of mechanisms and processes of the stabilization of soil organic matter for modelling carbon turnover. Journal ofFunctional Plant Biology,30: 207-222.

Landi, L., Renella, G., Moreno, J.L., Falchini, L. and Nannipieri, P. (2000). Influence of cadmiumon the metabolic quotient, l:-d-glutamic acid respiration ratio and enzyme activity: microbial biomass ratio under laboratory conditions. Journal ofBiology and

Fertility of Soils, 32(1): 8-15

Lebedeva, L.A., Lebedev S.N. and Edemskaya, N.L. (1995). The effect of heavy metals and lime on urease activity in soddy-podzolic soil. Moscow UniversitySoil

SciecityBulletin, 50(2): 53-60

Lenhard, G. (1956). The dehydrogenase activity in soil as a measure of the activity of soil microorganisms. Z Pflanzenernaehr Dueng Bodenkd, 73:1-11.

Lim, C.A. and P’ng, T.C. (1983). Land application of digested POME by sprinkler system. Proceedings of the seminar on land application of oil palm and rubber factory effluent, Serdang.

Logan, T.J., Lindsay, B.J. Goins, L.E. and Ryan, J.A. (1997). Field assessment of sludge metal bioavailability to crops: sludge rate response. Journal of Environmental

Quality, 26:534-550

Ma, A.N. (1999). Treatment of palm oil mill effluent. Oil palm and the environment. A Malaysian perspective. MOPGC, Kuala Lumpur,8:113-125.

Ma, A.N. (2000). Environmental management for the oil palm industry. Journal of Palm Oil

Development, 30:1-10.

Maila, M.P. and Cloete, T.E. (2005). The use of biological activities to monitor the removal of fuel contaminants perspectives to monitoring hydrocarbon contamination. Areview

ofInternational Biodeterioration and Biodegradation,55:1- 8.

Manjare, S. A., Vhanalakar S. A. and Muley, D.V. (2010). Analysis of water quality using physico-chemical Parameters tamdalge tank in kolhapur district, Maharashtra.

InternationalJournal of Advanced Biotechnology and Research,1(2):115-119.

Margesin, R., Feller,G., Hammerle, M., Stegner, U. and Schinner, F. (2002). Colorimetric methodfor determination of lipase activity in soil. Biotechnology Letters, 24: 27-33.

Margesin, R.G., Walder, A. and Schinner, F. (2000). The impact of hydrocarbon remediation on enzyme activity and microbial properties of soil. Acta Biotechnologica, 20:313-333.

Margesin, R., Zimmerbauer, A. and Schinner, F. (1999). Soil lipase activity: A useful indicator of oil biodegradation. Journal ofBiotechnology Techniques,13:859–863.

108

Marschner, H. (1995). Mineral Nutrition of Higher Plants. 2nd edn. Academic Press,London. pp.889-890

Martens, D.A., Johanson, J.B. and Frankenberger, W.T. (1992). Production and persistence of soil enzymes with repeated addition of organic residues. Journal of Soil Science,

153:53-61.

Masciandro, G., Ceccanti, B., Ronchi, V. and Baner, C. (2000). Kinetic parameters of dehydrogenase in the assessment of the response of soil to vermicompost and inorganic fertilisers. Journal ofBiology and Fertility ofSoils, 32:479-483.

Milosevic, N., Govedarica, M., Jarak, M., Petrovic, N., Jevtic, S. and Lazic, B. (1997).The effect of heavy metals on total soil microbiological activity in lettuce.Acta

Horticulturae, 462:133-140.

Moore, J.M., Klose, S. and Tabatabai, M.A. (2000). Soil microbial biomass carbon and nitrogen as affected by cropping systems. Journal ofBiology and Fertility of Soils, 31:200-210.

Moreno, J.L., Garcia, C., Landi, L., Falchini, L., Pietramellara, G. and Nannipieri, P. (2002).The ecological dose value (ed50) for assessing cadmium toxicity on dehydrogenase and urease activities of soil.Journal ofSoil Biology and

Biochemistry,33(4-5):483-495

Malaysian Palm Oil Board(MPOB) Data for Engineers (2004): Palm oil mill effluents.Palm

Oil Engineering Bulletin, 71:34-35

Muhrizal, S., Shamshuddin, J. Fauziah, I. and Husni,M.A.H. (2006). Changes in iron-poor acid sulfate soil upon submergence. Journal ofGeoderma,131:110-122.

Mumtaz, T., Yahaya,N.A., Abd-aziz,S.,Abdul-Rahman,N.P.L., Yee, Y.S. and Hassan, M.A. (2010). Turning waste to wealth biodegradableplastics polyhydroxyalkanoates from palm oil mill effluent – a Malaysian perspective. Journal of Clean Production, 18(14):1393-1402.

Murhekar, G.H. (2011). Determination of Physico-Chemical parameters of Surface Water Samples in and around Akot City. International Journal of Research in Chemistry

and Environment, 1(2):183-187.

Najafpour, G.D., Zinatizadeh, A.A.L., Mohamed, A.R., Isa, M.H. and Nasrollahzadeh,H. (2006). High-rate anaerobic digestion of palm oil mill effluent in an up flow anaerobic sludge- fixed film bioreactor. Journal ofProcess Biochemistry, 41(2):370-379.

Navas, A., Bermudez, F. and Macin, J. (1998). Influence of sewage sludge application on physical and chemical properties of gypsisols. Journal ofGeoderma,87:123-135.

Ndiaye, E.L., Sandeno, J.M., McGrath, D. and Dick, R.P. (2000). Integrative biologicalindicators for detecting change in soil quality. American Journal of

Alternative Agriculture, 15:26-36.

109

Nwoko, C.O. and Ogunyemi, S. (2010). Evaluation of palm oil mill effluent on maize (Zea

mays L) crop: yield, tissue nutrient content and residual soil chemical properties.Australian Journal of Crop Science,4(1): 16-22

Nwoko, C.O., Ogunremi. S., Nkwocha, E.E. and Nnorom, I.C. (2010).Evaluation of phytotoxicity effect of palm oil mill effluent and cassava mill effluent on tomato (Lycopersicumesculentum) after pretreatment option.International Journal of

Environmental Science and Development,1: 67-72

O’Thong, S., Prasertsan, P., Intrasungkha, N., Dhamwichukorn, S. and Birkeland, N. (2007). Optimization of simultaneous thermophilic fermentative hydrogen production and COD reduction from palm oil mill effluent bythermoanaerobicbacterium-rich sludge. International Journal of Hydrogen Energy, 33:1221-1231.

Obodo, G.A. (2002). The bioaccumulation of heavy metals in fish from Anambra river. Journal of Chemical Society of Nigeria, 29:60-62.

Ohimain, E.I., Enetimi, I.S., Izah, S.I., Oghenegueke, E.V. and Perewarebo, T.G. (2012). Some selected physicochemical and heavy metal properties of palm oil mill effluents. Greener Journal of Physical Sciences, 2(4):131-137

Okwute, O.L. and Isu, N.R. (2007). The environmental impact of palm oil mill effluent on some physicochemical parameters and total aerobic bioload of soil at a dump site in Anyigba, KogiState, Nigeria. African Journal of Agricultural Research,2(12):656-662.

Onyia, C.O., Uyub, A.M., Akunna, J.C., Norulaini, N.A. and. Omar A.K.M. (2001). Increasing the fertilizer value of palm oil mill sludge: Bioaugumentation in nitrification. Journal ofWater Science and technology, 44(10):157-162.

Osuji, L.C. and Nwoye, I. (2007). An appraisal of the impact of petroleum hydrocarbons on soil fertility: The Owaza experience. African Journal of AgriculturalResearch, 2(7): 318-324.

Oswal, N., Sarma P.M., Zinjarde, S.S. and Pant, A. (2002). Palm oil mill effluent treatment by a tropical marine yeast. Journal ofBioresources Technology, 85: 35-37.

Oviasogie, P.O. and Aghimien, A.E. (2002). Macro nutrient status and speciation of Cu, Fe, Zn and Pb in soil containing palm oil mill effluent. Global Journal of Pure and

Applied Science, 9(1):71-80.

Palm, C.A., Gachengo,C.N. Delve,R.J. Cadisch, G. and Giller,K.E. (2001). Organic inputs for soil fertility management in tropical agroecosystems: Application of an organic resource database. Agricultural Journal of Ecosystem Environment,83:27-42.

Paredes, C., Cegarra, J. Bernal, M.P. and Roig, A. (2005). Influence of olive mill waste water in composting and impact of the compost on a Swiss chard and soil properties.Journal

ofEnvironmental International, 31:305-312.

Pascual, A.C., Garcia, C., Polo A. and Sanchez-Diaz, M. (2007). Effect of water deficit on microbial characteristics in soil amended with sewage sludge or inorganic fertilizer under laboratory conditions. Journal ofBioresources Technology,98:29-37

110

Paul, E.A. and Clark, F.E. (1989). Soil Microbiology and Biochemistry. Academic Press Incorporated San Diego, California. pp.111-234.

Perucci, P. (1992). Enzymes activity and microbial biomass in a field soil amended with municipal refuse. Journal ofBiology and Fertility of Soils, 14:54-60.

Piotrowska, A., Iamarino, G., Rao, M.A. and Gianfreda, L. (2006). Short-term effects of Oliver mill waste water (OMW) on chemical and biochemical properties of a semiarid Mediterranean soil. Journal ofSoil Biology and Biochemistry, 38:600-610.

Prasertsan, S. and Prasertsan, P. (1996). Biomass residues from palm oil mill in Thailand: an overview on quantity and potential usage. Journal ofBiomass and Bioenergy, 11(5):387-395.

Purushothaman, M., Anderson,R.K.I., Narayana,S. and Jayaraman,V.K. (2001). Industrialbyproducts as cheaper medium components influencing the production of Polyhydroxyalkanoates (PHA) – Biodegradable Plastics. Journal ofBioprocess and

Biosystem Engineering, 24:131-136.

Quilchano, C. and Maranon, T. (2002). Dehydrogenase activity in Mediterranean forest soils. Journal ofBiology and Fertility of Soils, 35:102-107.

Rani, P., Meena, U.K. and Karthikeyan, J. (2004). Evaluation of antioxidant properties of berries. Indian Journal of Clinical Biochemistry, 19(2):103-110.

Rashid, S.S., Alam, M.Z., Abdulkarim, M.A. and Salleh, M.H. (2009). Management of palm oil mill effluent through production of cellulases by filamentous fungi. World Journal

of Microbiology and Biotechnology, 25(12):2219-2226.

Rasmussen P.E. and Collins H.P. (1991) Long-term impacts of tillage, fertilizer, and crop residue on soil organic matter in temperate semiarid regions. Journal of Advance

Agronomy,45:93-134

Rathnayake, I.V.N, Megharaj, M., Bolan, N. and Naidu, R. (2010). Tolerance of heavy metals by Gram positive soil bacteria. International Journal of Civil Environmental

Engineering, 2: 191-195.

Razak, C.N.A., Salleh, A.B., Musani, R., Samad, M.Y. and Basri, M. (1997). Some characteristics of lipases from thermophilic fungi isolated from palm oil mill effluent. Journal of Molecular Catalysis B: Enzyme, 3:153-159

Redzwan, G., Gan, S.N. and Tan, I.K.P. (1997). Short communication: Isolation of polyhydroxyalkanoate producing bacteria from an integrated-farming pond and palm-oil mill effluent ponds. World Journal of Microbiology and Biotechnology, 13:707-709.

Reeves, J.B. andWeihrauch,J.L. (1979) Consumer and Food Economics Institute (1979). Composition of foods: fats and oils. Agriculture handbook 8-4. Washington, D.C. U.S. Department of Agriculture, Science and Education Administration. p. 4.

111

Rhoades, J.D. (1982). Cation exchange capacity. In: A.L. Page, R.H. Miller and D.R. Keeney (Eds.) Methods of Soil Analysis, Part 2, Chemical and Microbiological Properties. Madison, Wisconsin. pp.149-157.

Romig, D.E., M.J. Garlynd, and Harris,R.F. (1996). Farmer-based assessment of soil quality: A soil health score card. Methods for Assessing Soil Quality. In: J.W. Doran and A.J. Jones (Eds.) SSSA Special Publication No. 49, Soil Science Society of America, Incorporated, Madison, WI. pp. 39-60.

Ros, M., Hernandez, M.T. and Garcia, C. (2003). Soil microbial activity after restoration of a semiarid soil by organic amendments. Journal ofSoil Biology and Biochemistry,

35:463-469.

Roukas, T. and Kotzekidou, P. (1997). Pretreatment of date syrup to increase citric acid production. Journal ofEnzyme Microbiology and Technology, 21: 273-276.

Rupani, P.F., Singh, R.P., Ibrahim, H. and Esa, N. (2010). Review of current palm oil mill effluent (POME) treatment methods: vermicomposting as a sustainable practice. World Applied Sciences Journal, 11(1):70-8I.

Salihu, A., Alam, M.Z., Abdulkarim, M.I. and Salleh, H.M. (2011a). Suitability of using Palm oil mill effluent as a medium for lipase production. African Journal of

Biotechnology, 10(11):2044-2052.

Salihu, A., Alam, M.Z., Abdulkarim, M.I. and Salleh, H.M. (2011b). Optimization of lipaseproduction by Candida cylindracea in palm oil mill effluent based medium using statistical experimental design. Journal Molecular Catalysis B: Enzyme, 69: 66-73.

Salihu, A. and Alam, M.Z. (2012). Palm oil mill effluent: a waste or a raw material? Journal of Applied Sciences Research, 8(1): 466-473.

Sarrantonio, M., Doran,J.W. Liebig,M.A. and Halvorson,J.J. (1996). On-farm assessment of soil quality and health: Methods for Assessing Soil Quality. SoilScience Society of

America Journal,49: 83-105.

Schinner, F., Ohlinger, R., Kandeler, E. and Margesin, R.(1996).Methods in Soil Biology. Springer, Heidelberg. pp. 204–207.

Sehar, S., Naz, I., Ali, M.I. and Ahmad, S. (2011). Monitoring of physicochemical and microbiological analysis of underground water samples of district Kallar Syedan, Rawalpindi-Pakistan. Research Journal of ChemicalSciences, 1(8): 24-30.

Sethupathi, S.(2004).Removal of residue oil from palm oil mill effluent treatment.Process Biochemistry, palm oil mill effluent (POME) using chitosan.Journal ofUniversiti,41: 962-964.

Shamshuddin, J., Jamilah, I. and Ogunwale, J.A. (1995). Formation of hydroxyl-sulfates from pyrite in coastal acid sulfate soil environments in Malaysia. Journal ofSoil Science

and Plant Analysis, 26(17-18):2769-2782.

112

Shiyin, L., Lixiao, N., Panying, P., Cheng, S. and Liansheng, W. (2004). Effects of pesticides and their hydrolysates on catalase activity in soil. Bulletin of Environmental

Contaminationand Toxicology, 72: 600–606.

Sikora, L. and Stott, D. (1996). Soil organic carbon and nitrogen: methods for assessing soil quality. Soil Science Society of America Journal,49: 157–167.

Singh, R.P., Ibrahim, M.H., Esa, N. and Iliyana, M.S. (2010). Composting of waste from palm oil mill: A sustainable waste management practice. Reviewof Environmental

Science and Biotechnology, 9: 331-344.

Skyllberg, U. (1993). Acid-Base properties of humus layers in northern coniferous forests. Dissertation, Swedish University of Agricultural Sciences, Department of Forest Ecology. Soil Science section, 54.

Sojka, R.E., and Upchurch, D.R. (1999). Reservations regarding the soil quality concept. Soil

Science Society of American Journal,63:1039-1054.

Sparks, D.L. (1995). Environmental Soil Chemistry. Academic Press. San Diego. p. 267

Strojan, C.L. (1978). Forest leaf litter decomposition in the vicinity of a zinc smelter.Oecologia Journal,32:203-212

Sumner, M.E. and Miller, W.P. (1996). Cation exchange capacity, and exchange coefficients. In: D.L. Sparks (Ed.) Methods of soil analysis. Part 2: Chemical properties (3rd Edition). Soil Science Society of America,Madison, WI. pp. 115-137

Tabatabai, M.A. (1982). Soil enzymes. In: Methods in soil analysis, part 2, Wisconsin. American Society of Agronomy. Journal ofSoil Science, 27: 903-947.

Thomas, G.W. (1982). Exchangeable cations.In:A.L. Page (Ed.). Methods of soil analysis. Part 2: Chemical and microbiological properties (2nd Edition).Journal of

Agronomy,9:159-165.

Tokunaga, T.C., Wan, J., Hazen, T.C., Schwartz, E., and Firestone, M.K. (2003). Distribution of chromium contamination and microbial activity in soil aggregates. Journal of

Environmental Quality,32:541-549.

Trasar-Cepeda, C., Leiros M.C., Seoane, S. and Gil-Sotres, F. (2007). Limitations of soil enzymes as indicators of soil pollution. Journal ofSoil Biology and Biochemistry, 32: 1867-1875.

Turick, C.E., Apel, W.A., and Carmiol, N.S. (1996). Isolation of hexavalent chromium reducing anaerobes from hexavalent chromium contaminated and non contaminated environments. Journal of Microbial Biotechnology, 44:683-688.

Vanlauwe, B., Diels,N. Sanginga, J. and Merckx, R. (2005). Long-term integrated soil fertility management in south-western Nigeria: crop performance and impact on the soil fertility status. Journal of Plants and Soil,273: 337-354.

Vijayaraghavan, K., Ahmad, D. and Abdul Aziz, M.E. (2007). Aerobic treatment of Palm oil mill effluent. Journal of Environmental Management, 82:24-31.

113

Visser, S. and Parkinson, D. (1992). Soil biological criteria as indicators of soilquality: Soil microorganisms. American Journal of Alternative Agriculture, 7:3-37.

Waksman, S.A. (1992). Microbiological analysis of soil as an index of soil fertility. Influence of fertilization upon numbers of microorganisms in soil. Journal ofSoil Science, 14:321-346.

Walkley, A. and Black, I.A. (1934). An examination of degtjareff method for determining Soil organic matter and a proposed modification of the chromic acid titration method.Journal ofSoil Science, 37: 29-37.

Welp, G. (1999). Inhibitory effects of the total water-soluble concentrations of nine different metals on the dehydrogenase activity of a loess soil.Journal ofBiology and Fertility of

Soils, 30(1-2):132-139

Wong, Y.S., Kadir, M.O.A.B. and Teng, T.L. (2009). Biological kinetics evaluation of anaerobic stabilization pond treatment of palm oil mill effluent. Journal

ofBioresources and Technology, 100:4969-4975

Wood, B.J. (1977). A review of current methods for dealing with palm oil mill effluents. Planter,53: 477-495.

Wood, B.J., Pilai, K.R. and Rajaratnam, J.A. (1979). Palm oil mill effluent disposal on land. Journal of Agricultural Wastes, 1:103-127

Wu, T.Y., Mohammad, A.W., Jahim J.M.D. and Anuar, N. (2007). Palm oil mill effluent (POME) treatment and bioresources recovery using ultrafiltration membrane: Effect of pressure on membrane fouling. Biochemical Engineering Journal, 35:309-317

Wyszkowska, J., Kucharski, J., Jastrzebska, E. and Hlasko, A. (2001).The biological properties of the soil as influenced by chromium contamination.PolishJournal of

Environmental Studies, 10(3): 175-180

Zaini, U., Salmiati, S. and Salim, M.R. (2010). Microbial Biopolimerization Production from Palm Oil Mill Effluent (POME), Biopolymers, Magdy Elnashar (Ed.), InTech, Available from:http://www.intechopen.com/books/biopolymers/microbial-biopolimerization-production-from palm-oil-mill-effluent-pome

Zheng, C.R., Tu, C.and Chen, H.M. (1999).Effect of combined heavy metal pollution on nitrogen mineralization potential, urease and phosphatase activities in a typicudicferrisol.Journal ofPedosphere,9(3): 251-258

114

APPENDICES

Appendix 1: Standard Curve for the Determination of Potassium

Appendix 2: Standard Curve for the Determination of Sodium

y = 0.68x

0

5

10

15

20

25

30

0 10 20 30 40 50

Ab

sorb

an

ce

Concentration of sodium (ppm)

y = 0.68x

0

5

10

15

20

25

30

0 10 20 30 40 50

Ab

sorb

an

ce

Concentration of potassium (ppm)

115

Appendix 3: Standard Curve for the Determination of Phosphorus

Appendix 4: Standard Curve for the Determination of the Soil Lipase Activity

y = 0.018x

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

0 20 40 60 80 100

Ab

sorb

an

ce

Concentration (mg/L)

y = 3.405x - 22.38R² = 0.980

-20

0

20

40

60

80

100

120

0 10 20 30 40

Con

cen

trati

on

of

lau

ric

aci

d (

mg/L

)

Titre values (ml)

Conc. (mg/L)

Linear (Conc. (mg/L))

116

Appendix5: Physicochemical Parameters in the Soil Contaminated with the Fresh and

Fermented POME

Mill Sites Sites

Soil types

Nitrogen (meq/100g)

Sodium (meq/100g)

Potassium (meq/100g)

Calcium (meq/100g)

Amurie Dumpsite Topsoil 0.21±0.001 0.0078±0.001 0.0380±0.004 1.84±0.005

Subsoil 0.24±0.001 0.0062±0.001 0.0280±0.005 1.44±0.005

Tenyards Topsoil 0.25±0.001 0.0059±0.001 0.0330±0.003 1.85±0.003

Subsoil 0.23±0.002 0.0065±0.001 0.0360±0.003 1.49±0.004

Control Topsoil 0.23±0.001 0.0060±0.001 0.0377±0.005 1.34±0.005 Subsoil 0.23±0.030 0.0048±0.001 0.0356±0.005 1.47±0.003

Eziisu Dumpsite Topsoil 0.25±0.002 0.0085±0.002 0.0294±0.003 1.58±0.004

Subsoil 0.20±0.001 0.0058±0.001 0.0285±0.004 1.68±0.004

Tenyards Topsoil 0.22±0.001 0.0068±0.001 0.0340±0.004 1.77±0.003

Subsoil 0.20±0.002 0.0067±0.001 0.0218±0.004 1.68±0.004

Control Topsoil 0.22±0.001 0.0060±0.001 0.0377±0.005 1.34±0.005

Subsoil 0.23±0.030 0.0048±0.001 0.0356±0.003 1.47±0.003

Nnerim Dumpsite Topsoil 0.21±0.001 0.0083±0.002 0.0412±0.003 1.32±0.002

Subsoil 0.16±0.001 0.0071±0.001 0.0315±0.005 1.39±0.001

Tenyards Topsoil 0.25±0.001 0.0078±0.001 0.0292±0.005 1.57±0.004 Subsoil 0.21±0.002 0.0077±0.001 0.0284±0.004 1.85±0.030

Control Topsoil 0.23±0.001 0.0060±0.001 0.0377±0.003 1.34±0.005

Subsoil 0.23±0.030 0.0048±0.001 0.0356±0.003 1.47±0.003

Olori Dumpsite Topsoil 0.25±0.002 0.0079±0.001 0.0196±0.002 1.57±0.002

Subsoil 0.24±0.017 0.0050±0.001 0.0274±0.002 1.83±0.004

Ten yards Topsoil 0.23±0.001 0.0066±0.001 0.0274±0.001 1.63±0.003

Subsoil 0.22±0.001 0.0060±0.001 0.0290±0.005 1.74±0.005

Control Topsoil 0.23±0.001 0.0060±0.001 0.0377±0.004 1.34±0.005

Subsoil 0.23±0.030 0.0048±0.001 0.0360±0.004 1.47±0.003

Umudio-kpara Dumpsite Topsoil 0.25±0.001 0.0061±0.001 0.0444±0.003 1.65±0.004

Subsoil 0.27±0.001 0.0011±0.001 0.0286±0.003 1.27±0.003


Subsoil 0.22±0.001 0.0061±0.001 0.0394±0.005 1.51±0.006

Control Topsoil 0.23±0.002 0.0060±0.001 0.0377±0.005 1.34±0.004

Subsoil 0.23±0.030 0.0048±0.001 0.0356±0.004 1.47±0.003

Obeakpu Dumpsite Topsoil 0.22±0.002 0.0061±0.001 0.0016±0.001 1.53±0.004

Subsoil 0.20±0.001 0.0011±0.001 0.0376±0.003 1.63±0.005


Subsoil 0.19±0.001 0.0097±0.002 0.0376±0.002 1.76±0.003

Control Topsoil 0.23±0.001 0.0060±0.001 0.0377±0.003 1.34±0.005

Subsoil 0.23±0.030 0.0048±0.001 0.0356±0.003 1.47±0.003

117

Appendix6: Continuation of the Physicochemical Parameters in the Soil Contaminated with

the Fresh and Fermented POME

Mill Sites Sites

Soil types

Organic carbon (%)

Organic Matter (%)

CEC (meq/100g)

Phosphorous (ppm)

Magnesium (meq/100g)

Amurie Dumpsite Topsoil 2.86±0.004 4.94±0.007 15.52±0.003 21.68±0.002 0.51±0.003

Subsoil 2.67±0.004 4.61±0.006 18.75±0.116 21.59±0.002 0.49±0.004

Tenyards Topsoil 2.99±0.002 5.28±0.006 18.83±0.004 26.63±0.003 0.53±0.004

Subsoil 3.06±0.003 5.15±0.003 16.63±0.005 24.86±0.004 0.50±0.003

Control Topsoil 2.30±0.002 3.96±0.250 15.53±0.004 26.66±0.003 0.63±0.003

Subsoil 2.42±0.002 4.16±0.004 19.63±0.005 30.63±0.007 0.63±0.002

Eziisu Dumpsite Topsoil 2.84±0.003 4.80±0.173 14.93±0.003 22.77±0.005 0.54±0.004

Subsoil 2.70±0.003 4.65±0.005 18.61±6.348 22.77±0.003 0.55±0.004


Subsoil 3.12±0.002 5.37±0.003 16.12±0.004 26.50±0.233 0.52±0.004

Control Topsoil 2.30±0.002 3.96±0.250 15.53±0.040 26.66±0.003 0.63±0.003

Subsoil 2.42±0.002 4.16±0.004 19.63±0.005 30.63±0.007 0.63±0.002

Nnerim Dumpsite Topsoil 2.69±0.003 4.63±0.005 15.69±0.003 24.50±0.132 0.67±0.003

Subsoil 3.14±0.004 5.41±0.006 21.28±0.001 22.64±0.004 0.57±0.015


Subsoil 3.25±0.003 5.60±0.004 20.39±0.003 25.64±0.005 0.53±0.003

Control Topsoil 2.30±0.002 3.96±0.250 15.53±0.004 26.66±0.003 0.63±0.003

Subsoil 2.42±0.002 4.16±0.004 19.63±0.005 30.63±0.007 0.63±0.002

Olori Dumpsite Topsoil 2.51±0.004 4.33±0.007 14.79±0.002 20.66±0.002 0.48±0.002

Subsoil 2.52±0.005 4.35±0.008 14.56±0.005 20.74±0.003 0.36±0.004


Subsoil 2.37±0.002 4.08±0.003 20.84±0.004 29.63±0.004 0.44±0.004

Control Topsoil 2.30±0.002 3.96±0.250 11.53±0.004 26.66±0.003 0.63±0.003

Subsoil 2.42±0.002 4.16±0.004 13.63±0.005 30.63±0.007 0.63±0.002

Umudiokpara Dumpsite Topsoil 3.54±0.004 6.09±0.007 14.43±0.003 22.43±0.003 0.69±0.003

Subsoil 3.61±0.002 6.23±0.004 13.38±0.002 27.64±0.003 0.67±0.004


Subsoil 2.64±0.003 4.56±0.004 17.66±0.004 22.75±0.004 0.67±0.004

Control Topsoil 2.30±0.002 3.96±0.250 11.53±0.004 26.66±0.004 0.63±0.003

Subsoil 2.42±0.002 4.16±0.004 13.63±0.005 30.63±0.007 0.63±0.002

Obeakpu Dumpsite Topsoil 3.17±0.004 5.47±0.006 14.83±0.002 20.76±0.003 0.49±0.002

Subsoil 2.89±0.004 4.99±0.007 14.84±0.005 24.80±0.003 0.44±0.003


Subsoil 2.57±0.005 4.42±0.008 18.74±0.005 32.71±0.003 0.57±0.003

Control Topsoil 2.30±0.002 3.96±0.250 15.53±0.004 26.66±0.003 0.63±0.003

Subsoil 2.42±0.002 4.16±0.004 19.63±0.005 30.63±0.007 0.63±0.002

118

Appendix 7: Dehydrogenase activity in the soil contaminated with the fresh and fermented

POME (mg formazan g-1 dry soil/96h)

Mill Sites

Dumpsite1 Ten yards

Control Topsoil Subsoil Topsoil subsoil Topsoil Subsoil

Amurie 0.1757±0.04 0.0993±0.01 0.0744±0.01 0.1014±0.01 0.758±0.001 0.86±0.01

Eziisu 0.1575±0.03 0.0965±0.02 0.1186±0.03 0.0762±0.01 0.758±0.001 0.86±0.01

Nnerim 0.3208±0.03 0.2732±0.03 0.1112±0.01 0.2302±0.02 0.758±0.001 0.86±0.01

Olori 0.0943±0.03 0.0936±0.01 0.1157±0.01 0.1069±0.01 0.758±0.001 0.86±0.01

Umudiokpara 0.8417±0.08 0.1623±0.05 0.0877±0.01 0.0858±0.01 0.758±0.001 0.86±0.01

Obeakpu 0.1471±0.06 0.0767±0.01 0.1126±0.02 0.0653±0.01 0.758±0.001 0.86±0.01

Appendix 8:Catalase activity in the soil contaminated with the fresh and fermented POME

(mM H2O2/g dry soil/h)

Mill sites

Dumpsites Ten yards

Control Topsoil Subsoil Topsoil subsoil topsoil Subsoil

Amurie 0.822±0.03 0.385±0.01 0.185±0.01 0.442±0.07 1.346±0.02 2.084±0.08

Eziisu 0.445±0.03 1.012±0.07 1.038±0.20 1.012±0.07 1.346±0.02 2.084±0.08

Nnerim 0.391±0.05 0.603±0.01 0.353±0.04 0.977±0.03 1.346±0.02 2.084±0.08

Olori 0.465±0.09 0.205±0.01 0.192±0.01 0.207±0.01 1.346±0.02 2.084±0.08

Umudiokpara 0.332±0.01 0.400±0.02 1.105±0.09 1.420±0.01 1.346±0.02 2.084±0.08

Obeakpu 0.179±0.01 0.498±0.01 0.178±0.02 0.498±0.07 1.346±0.02 2.084±0.08

Appendix 9:Lipase activity in the soil contaminated with the fresh and fermented

POME(mM/g dry soil/18h)

Mill Sites

Dumpsites Ten yards Control

Topsoil Subsoil topsoil Subsoil topsoil Subsoil

Amurie 17.295±1.7 08.695±0.3 08.030±0.5 06.838±0.3 7.937±0.7 13.137±0.5

Eziisu 14.691±0.3 09.575±0.8 09.724±0.7 08.579±0.4 7.937±0.7 13.137±0.5

Nnerim 22.687±0.8 08.950±1.1 10.203±0.3 19.095±2.1 7.937±0.7 13.137±0.5

Olori 09.152±0.9 13.576±1.1 08.178±0.2 07.164±0.3 7.937±0.7 13.137±0.5

Umudiokpara 15.726±0.2 08.627±0.3 11.371±0.2 10.151±2.0 7.937±0.7 13.137±0.5

Obeakpu 08.883±0.3 09.130±2.3 10.057±1.1 08.031±0.3 7.937±0.7 13.137±0.5

Documents

UROKO, ROBERT IKECHUKWU (PG/M. Sc/12/61371) Robert.pdf · impact of fresh and fermented palm oil mill effluents on soil physicochemical parameters and enzyme activities in umuaka,