EFFECT OF NITROHUMIC ACID DERIVED FROM LOW-
RANK COAL OF SARAWAK ON GROWTH OF BRASSICA
OLERACEA SP.
ZULFAQAR BIN SA’ADI
Final report submitted as partial fulfillment of the
requirements for the degree of
Bachelor of Science with Honors in
Resources Chemistry
Faculty of Resource science and technology
UNIVERSITI MALAYSIA SARAWAK
2008
i
DECLARATION
No portion of the work referred to in this description has been
submitted in support of an application for another degree of
qualification of this and any other university or institution of higher
learning.
-------------------------
Zulfaqar bin Sa’adi
Resources Chemistry
Faculty of Resource science and technology
Universiti Malaysia Sarawak
ii
APPROVAL SHEET
Name of candidate: Zulfaqar bin Sa’adi
Title of dissertation: Effect of Nitrohumic acid derived from Low-
rank coal of Sarawak on growth of Brassica Oleracea sp.
------------------ ………………
Mdm Rafeah Wahi Dr Petrus Bulan
Supervisor Co-supervisor
------------------
Head of Chemistry Resources
Faculty of Resource science and technology
iii
ACKNOWLEDGMENTS
Firstly, I would like to thank Allah for His blessing upon me to
complete my project. Besides, special thanks go to Mdm. Rafeah
Wahi as Supervisor for her full encouragement, guidance,
supervision and professionalism. Furthermore, Thanks also to our
lab assistants, for their materials provided, patience and helps
concerning the lab. Thanks also to PTPTN for providing me
financial supports to complete my final year project. Lastly, to
fellow friends and people that involved direct or indirectly in
finishing this project. The author also wishes to thank University
Malaysia Sarawak for financial support.
iv
TABLE OF CONTENT
Content Page
LIST OF FIGURES……………………………..........................vii
LIST OF TABLES……………………………………………...viii
ABSTRACT……………………..………………………..….……x
ABSTRAK………………………..…………………………….….xi
CHAPTER 1: INTRODUCTION ……………………..….……..1
CHAPTER 2: LITERATURE REVIEW
2.1 Definition of humic acids……………..…………..……..…3
2.2 Important properties of humic acids as soil conditioner…...4
2.3 Extraction of humic acids…………………………....….....6
2.4 Sandy soil……………………………………………..…....7
2.5 Peat soil……………………………………………..…..….7
2.6 Laterite soil……………………………………….…..……8
2.7 Spectroscopic studies on the characteristic of nitrohumic
acids…………………………………………………..……9
2.7.1 Infrared spectroscopy……………………….....…...9
2.7.2 UV-Vis spectroscopy………………………..…....11
v
2.7.3 CHN Analyzer……………...…………………......11
2.8 Aromaticity……………….……...………………….……12
CHAPTER 3: METHODOLOGY
3.1 Material…………………………...………………….…...13
3.2 Nitric acid oxidation………………..………………..……13
3.3 Extraction of humic acid using KOH and NaOH...............14
3.4 Yield of nitrohumic acids…………...………………..…...14
3.5 Characterization of nitrohumic acids derived from coal….14
3.5.1 Moisture content……..………………………..….15
3.5.2 Ash content…………………………………....….15
3.6 Spectroscopic characterization………..………………......15
3.6.1 Fourier-Transform Infrared (FTIR) spectroscopy...15
3.6.2 UV/VIS spectrophotometer………………….........16
3.6.3 CHN Analyzer……………………………..……...16
3.7 Pot and laboratory experiment: Effect of nitrohumic acids on
growth of Brassica Oleracea sp. and soil analysis….....................16
3.7.1 Preparation of soil nitrohumic acids…………...…17
3.7.2 Growth of Brassica Oleracea sp………………………….17
vi
3.7.3 Soil organic matter content, pH values and Total soil water
content……………….........................................................18
3.7.4 Soil density, particle density and porosity..........................20
3.7.5 CHN Analyzer…………………..…………………..…….22
CHAPTER 4: RESULTS AND DISCUSSION
4.1 Characteristics of nitrohumic acids…….........................…23
4.2 FTIR……………………….……..………………..……...24
4.3 UV/VIS…………………….………………...…….……..26
4.4 Ultimate analysis …............................................................27
4.5 Yield of nitrohumic acids………….………………...…....29
4.6 Pot and laboratory experiment: Effect of nitrohumic acids
on growth of Brassica Oleracea sp. and soil analysis……31
4.6.1 Growth of Brassica Oleracea sp………………….31
4.6.2 Soil organic matter content and pH value………...33
4.6.3 Soil density, particle density and porosity…..........36
4.6.4 Total soil water content. …………………....….…41
4.6.5 CHN Analyzer ………………………………........43
vii
CHAPTER 5: CONCLUSIONS.………..………………...……46
REFERENCES…………………………………………………..48
APPENDIX………………………………………….……...……52
LIST OF FIGURES
Figures Pages
Figure 1 Model structure of humic acid (Stevenson, 1982); R
can be alkyl, aryl or arakyl……………………….4
Figure 2 FTIR spectra of coal, nitrated coal and nitrohumic
acid……………………………………………………..24
Figure 3 UV/Vis spectra of nitrohumic acid……………….26
Figure 4 Percentage dry weight of Brassica Oleracea sp.
compare to control………………………………..31
Figure 5 Soil organic matter of soils……………………….33
Figure 6 pH value of soils………………………………….35
Figure 7 Soils Bulk density………………………………...36
Figure 8 Soil particle density……………………………….37
Figure 9 Soil porosity………………………………………39
Figure 10 Soil total water content in volume percentage of
H2O……………………………….……………….41
viii
Figure 11: C/N ratio of soil treat with different level of
nitrohumic acids......................................................43
Figure 12 Chemical properties of humic substances
(Stevenson, 1982)…………………………………52
LIST OF TABLES
Tables Pages
Table 1 Important absorption bands in the IR spectra of
Humus substances (Orlov, 1992)…………………10
Table 2 Proximate analysis of ccoal and nitrohumic acids..23
Table 3 CHN Analyzer result for coal, nitrated coal and
nitrohumic acid…………………………………...27
Table 4 Yield of Nitrohumic acid…………………………29
Table 5 Soil moisture content……………………………..41
Table 6 pH value of soils………………………………….52
Table 7 Soil organic matter of soils……………………….53
Table 8 Soils Bulk density………………………………...53
Table 9 Soil particle density………………………………53
Table 10 Soil porosity………………………………………54
ix
Table 11 Soil total water content……………………..…….54
Table 12 CHN Analyzer results for soil…………….………55
Table 13 C/N ratio of soil………………………………..…..56
x
Effect of Nitrohumic acid derived from Low-rank coal of
Sarawak on growth of Brassica Oleracea sp.
Zulfaqar bin Sa’adi
Chemistry Resources
Faculty of Resource science and technology
Universiti Malaysia Sarawak
ABSTRACT
Humic acid derived from low rank coal of Mukah prepared using alkaline
extraction method was referred as nitrohumic acid. Nitrohumic acid was
characterized for moisture content, ash content, nitrogen content, total acidity,
carboxyl groups and phenolic group. Spectroscopy characterization was
conducted using FTIR spectroscopy, UV/Vis spectroscopy and CHN Analyzer.
Optimization study conducted to investigate the role of the nitrohumic acids to
improve leafy vegetable growth of Brassica Oleracea sp. by using different level
of nitrohumic acids dosage and three types of soil was used namely peat soil,
sandy soil and laterite soil. Growth of Brassica Oleracea sp. had been monitored
for 51 days. The effects of nitrohumic acids on the aggregate stability are
dependent on the soil type. Growth of Brassica Oleracea sp. was improved by
24% and 39% by addition 50g and 100g of nitrohumic acids for peat soil and
23% to 30% for sandy soil, while laterit soil was not favorable for the growth of
Brassica Oleracea sp. Water holding capacity for sandy soil were 4% to 9%,
25% to 36% for peat soil and 20% to 28% for laterite soil. Soil organic matter
content was improved by 7 to 17% with the application of different levels of
nitrohumic acids as compared to control in peat soil, 8% to 188% in sandy soil
and 0.8% to 9% in laterite soil.
xi
ABSTRAK
Asid humik daripada arang batu yang berasal dari Mukah disediakan dengan
menggunakan pengekstrakan beralkal juga dikenali sebagai asid humik nitro.
Ciri-ciri asid humik nitro kemudian dilakukan dengan mengukur kandungan
kelembapan, kandungan abu, kandungan Nitrogen, jumlah keasidan, kumpulan
karboksil dan kumpulan fenolik. Kajian optimisasi dilakukan untuk menyelidiki
kegunaan asid humik nitro dalam meningkatkan pertumbuhan tumbuhan dedaun,
Brassica Oleracea sp. dengan menggunakan humik asid nitro yang pelbagai
peringkat dan tiga jenis tanah iaitu tanah paya, tanah berpasir dan tanah laterit.
Pertumbuhan Brassica Oleracea sp.diperhatikan dan diselidiki untuk setiap
replikasi selama 51 hari. Analisis tumbuhan dan tanah dilakukan kemudian. Asid
humik nitro boleh digunakan sebagai baja dengan kos yang rendah. Secara
keseluruhan, efek asid humik nitro kepada kestabilan tanah bergantung kepada
jenis tanah. Tanah berpasir dapat meningkatkan pertumbuhan Brassica
Oleracea sp. 24%-39%, dengan pertambahan asid humik nitro pada kadar 50g
dan 100g untuk tanah paya, 23% - 30% untuk tanah berpasir manakala tanah
laterite tidak sesuai untuk pertumbuhan Brassica Oleracea sp. . Setiap jenis
tanah memiliki kapasiti air yang berbeza di mana tanah berpasir 4% -9%, tanah
berpaya 25% to 36% dan tanah laterite 20% to 28%. Bahan organic tanah
ditingkatkan pada kadar 7-17% dengan aplikasi asid humik nitro yang berbeza,
8%-188% bagi tanah berpasir dan 0.8% to 9% bagi tanah laterite. Kadar
bahan organic adalah tidak sekata bagi setiap peringkat asid humik nitro dalam
tanah berpaya dan tanah berpasir tetapi bertambah bagi tanah laterit.
1
CHAPTER 1
INTRODUCTION
Soil structure is one of the factors in determining soil productivity.
Organic residues can acted as soil conditioner to improve the
organic matter status. Some synthetic polymers such as
polyacrylamides and polyvinyl alcohols were found to function
similarly (Gabriels 1990; Bryan 1992; Sojka and Lentz 1994).
Nevertheless, these synthetic conditioners were easily degraded by
microorganism. Humic substances had been well established as a
potential soil conditioner, better than synthetic conditioners. Other
advantages of humic substances are its ability to improve and
prolong aggregate stability at low application rate. In addition, they
were free from pollutants and highly reactive towards soil
components due to the presence of acidic functional groups and the
polycondense aromatic structure renders them more resistant to
microbial attack.
Today, humic acids have become commercially available in the
form of inexpensive soluble salts, referred to as sodium or
potassium humates. In Malaysia, they were less commonly used as
these products were imported; therefore they were relatively higher
in selling price than other soil conditioners. However, the
possibility to produce the humic acids from the indigenous source
would make the products better known. In Sarawak, abundant of
low rank coal was discovered in Mukah serving it as the potential
source of humic acids. Research on the effect of the locally
produced humic acids to improve the aggregate stability is limited.
This paper was reported on the potential of the nitrohumic acids
2
derived from the low rank coal of Mukah in improving growth of
plants and soil analysis. Previous study on effect of different levels
of coal derived humic acid on growth of plants was concluded that
the addition of humic acid even at low levels increased shoot and
root yield.
No detailed research studies had been carried out on the agricultural
aspects of humic acids and information in this regard is very
limited. This study was conducted to investigate the effect of the
addition of different levels of humic acids on the growth and
nutrient accumulation by Brassica Oleracea sp. and on soil
microbial activity and population, water-holding capacity, porosity,
pH and other parameters.
The objectives of the study:
a) To Extract and characterize nitrohumic acids derived from low-rank
coal of Mukah
b) To assess the suitability of nitrohumic acid derived from Mukah as
soil conditioner on the growth of Brassica Oleracea sp.
c) To assess the effect of nitrohumic acid derived from Mukah on peat
soil, sandy soil and laterite soil.
3
CHAPTER 2
LITERATURE REVIEW
2.1 Definition of humic acids
Coal humic acids were dark colored substances derived from coals
(Magdaleno and Coichev, 2005). They was occurred naturally in
some lignites and brown coals (Lawson and Stewart, 1989). It was
produced by the decay of organic materials and is found in soil, peat
and lignites (Lawson and Stewart., 1989) Humic acid was an alkali
soluble with aromatic structure substituted by carboxyl, phenolic,
hydroxyl, and alkyl groups linked together through either linkage
(Gaines et al., 1983). The hypothetical structure for humic acid
contains free and bound phenolic OH groups, quinone structures,
nitrogen and oxygen as bridge units and COOH groups variously
placed on aromatic rings.
Molecular weight of humic acids was greater than fulvic acids,
besides, they also less highly charged, less polar and more
aromatics (Hayes et al., 1989). Humic acids were fraction of soil
organic matter which could be precipitated at pH 2 from aqueous
alkaline extracts of soils. Humic acids constitute the higher
4
molecular weight fraction from 1500 to 50,000 to 500,000 Da in
streams and soils.
Figure 1: Model structure of humic acid (Stevenson, 1982); R
could be alkyl, aryl or arakyl
2.2 Important properties of humic acids as soil conditioner
Humic matter influences plant growth and equilibriums in
ecosystems through its effect on the physical, chemical and
biological properties of soils (Stevenson, 1994; Piccolo, 1996). It
was a naturally occurring polymeric organic compound and was
designated by nature to perform a wide variety of functions
(Schnitzer and Khan., 1972).
The properties of Base Exchange capacity and complexing ability
of humic acid were important in soil stability, transport of metal
ions in the soil through plant tissues and stabilization of soil organic
5
matter against microbiological attack (Vaughan and MacDonald,
1976). Humic acids contain many trace elements in its structure.
Taking advantage of the complexing properties, various
micronutrients were further complexed with humic acids to form
chelates (Barron and Wilson, 1981). These chelates were utilized to
overcome a specific deficiency in the soil and were used wherever
required in growth regulation of plants. Humic acids had been
complexed with sodium, potassium, manganese, zinc, calcium, iron,
copper, and with various other elements to overcome a particular
element deficiency in soil (Yingei, 1988).
Humic acid containing 51%–57%, 4%–6%, and 0.2%-1% organic
carbon, Nitrogen, and Phosphorus could improved crops yield due
to its capability of supplying N and P to the plants together with the
improvement in the physicochemical and biological environment of
the soils (Brannon and Sommers, 1985). Humic acids contain very
stable N content, which serves as effective slow releasing N
fertilizer (Nisar and Mir, 1989).
Humic acids serves as a catalyst in promoting the activity of
microorganisms in soil (Bhardwaj and Gaur, 1970). Addition of
humic acids to soil increases the rate of absorption of ions on root
surfaces and their penetration into the cells of the plant tissue.
6
Plants show more active metabolism and increased respiratory
activity, which were attributed to the intervention of the quinone
groups of humic acids (Petronio et al., 1982). There were evidences
that the effect of humic acids on plant growth were longer-lived
than other inorganic sources (Sibanda and Young, 1989). The
integrated uses of organic and inorganic fertilizers not only increase
each other efficiency, but could help in the substitution of chemical
fertilizers (Hussain et al., 1988). The use of humic acids was a
promising natural resource to be utilized as an alternative for
increased crop production (Nisar and Mir, 1989).
2.3 Extraction of humic acids
The most complete information on the content and composition of
the aromatic fragments of humus substances was obtained by the
method of oxidation (Orlov., 1992). The extraction of humic acids
was usually done by using alkaline extraction. Humic acids was
dissolved in the alkali solution and were latter separated from the
insoluble residue by filtration or centrifugation. Humic acids were
extracted from coal with dilute sodium or potassium hydroxide
solutions at concentration of 0.5 to over 10 percent. Weaker base as
sodium carbonate and ammonia were less effective. Air may be
7
excluded to avoid oxidation and the temperature employs were
usually moderate (Sakagami and Shimizugawa, 1962).
2.4 Sandy soil
Sandy Soils had gritty texture and were formed from weathered
rocks such as limestone, quartz, granite, and shale. Sandy soils had
relatively large spaces between particles, which provide for rapid
downward water movement. Substances, dissolved in leaching
waters, were readily transported deep into the soil. Coarse-textured
soils dry out quickly, which tends to increase wind erodibility. If
sandy soil contains enough organic matter it was easy to cultivate,
however it was prone to over-draining and summer dehydration,
and in wet weather it had problems retaining moisture and nutrients.
Bare surface of sandy soil was unable to hold nutrients. By coating
of humates, it provides charged surfaces for nutrient retention.
Treatment of sandy soil was required to reduce soil erosion and
improve texture.
2.5 Peat soil
Approximately 60% of the world's wetlands are peat. The high fertility
and proximity to water made peat soils the most desirable for vegetable
8
production. Peat was soft and easily compressed. Under pressure,
water in the peat is forced out. Peat was also dug into soil to increase
the soil's capacity to retain moisture and add nutrients. However, these
soils were also subject to quick degradation during agricultural usage.
They were not too acid and had effective sub drainage; these were
probably the best natural soils available because they were rich in plant
foods.
2.6 Laterite soil
Laterite soils result from laterisation, the process by which silicate
was removed leaving the soil enriched with oxides of iron and
alumina. The percolating rain water causes dissolution of primary
rock minerals and decrease of easily soluble elements as sodium,
potassium, calcium, magnesium and silicon. This gave rise to a
residual concentration of more insoluble elements predominantly
iron and aluminium. Laterites consist mainly of the minerals
kaolinite, goethite, hematite and gibbsite which form in the course
of weathering. Moreover, many laterites contain quartz as
relatively stable relic mineral from the parent rock. The iron oxides
goethite and hematite cause the red-brown color of laterites.
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9
Lateritic soils form the uppermost part of the laterite cover; in soil
science specific names (oxisol, latosol, ferallitic soil) are given for
them. Laterites can be as well soft and friable as firm and
physically resistant. The texture also varies widely from loamy to
clayey. However the soils were generally well-drained. The soils
show acidic to neutral reaction. Due to dominance of kandite clay
mineralogy, they possess low cation exchange capacity, low base
and nutrient status and poor organic matter but had higher amount
of iron and alumina.
2.7 Spectroscopic studies on the characteristic of humic
acids
2.7.1 Infrared spectroscopy
Infrared spectroscopy was the most useful spectroscopic procedures
for determining functional groups in humic substances. The
identification by infrared spectroscopy of oxygen containing
functional groups in humic substances is by derivation, which give
rise to a shift in the spectral band. By using derivation techniques,
carboxyl, esters, ethers, ketones and hydroxyl functional group can
be identified (Hayes et al., 1989). Table 1 summarizes the FTIR
absorption bands of the functional groups that are commonly found
in humic acids.
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10
Table 1: Important absorption bands in the IR spectra of
Humus substances (Orlov, 1992)
Wavelength (cm-1) Functional group present
3600 Free OH-group : Stretching
3500-3300 OH- group bound with intermolecular hydrogen bonds, partly with NH; stretching
2900 and 2800 CH2, CH3, stretching
1725-1700 C=O in COOH, partly other C=O and complex ethers and stretching
1650 ‘amide I’
1610-1600 C=C (aromatic), participation of carbonyls possible
1590-1580, 1400-1390
-(COO)-
1540 ‘amide II’
1510-1500 C=C (aromatic)
1460-1440 CH in CH2 (or CH3); deformation
1260-1200 Carboxyl group (C-O, partly OH)
1150-1050 Tertiary, secondary and primary alcohols
1080-1050 Polysaccharides
860-730 CH (aromatic) with two or more unsubstitued H
730-720 -(CH2)n- for n ≥4
11
2.7.2 UV-Vis spectroscopy
UV-Visible spectroscopy was a technique in studying the
absorption bands in the ultraviolet and visible spectra of humic
substances. The absorptivity increases at shorter wavelength. The
variation of the ultra-violet visible spectra as function of pH is
consistent with the occurrence of aromatic carboxylic acids and
phenols in humic substances. The absorbance ratio at 465 nm and
665 nm was often used to characterize humic acids. The absorbance
ratio which was referred to as E4/E6 is empirically correlated with
their degrees of aromaticity. The E4/E6 coefficient of humic aicds
was usually less than 5 and the low ratio indicates high degree of
condensation of aromatic constituents (Hayes et al., 1989)
2.7.3 CHN Analyzer
A CHN Analyzer was a scientific instrument which can verify elemental
composition of a sample. The name derives from the three primary
elements measured by the device: carbon (C), hydrogen (H) and nitrogen
(N). Sulfur (S) and oxygen (O) can also be measured. The analyzer uses a
combustion process to fracture down substances into simple compounds
which were then calculated. By sorting out inorganic carbon by means of
a solvent, organic carbon in a sample can be measured using this device as
well
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12
2.8 Aromaticity
The ratio of optical absorbances of humic acids solution at 465 nm
and 665 nm (E4/E6 ratio) is used to characterize the aromaticity. The
E4/E6 ratio is governed primarily by its molecular sizes and does not
reflect condensed aromatic ring (Chen, 1977). There were
relationship between E4/E6 ratio and spin content for fulvic acids
fractions on Sephadax gel. According to Stuermer et al. (1978),
ESR spectroscopy was used to measure free radical concentration
and aromatic character.
