2,4-dinitrophenol (2,4-DNP) is utilized in the production of wood preservatives, dyes, and also
as a pesticide. Human acute (short-term) exposure to 2,4-DNP in humans by means of oral
exposure are nausea or vomiting, sweating, headaches, dizziness, and weight reduction. Thus, the
removal of this compound is highly sought. A 2,4-DNP-degrading bacterium (isolate 1) was
isolated from a sample soil from Terengganu. This bacterium (isolate 1) was characterized as a
rod Gram positive, non-sporulated, and non-motile bacterium. The bacterium is oxidase negative
and had catalase positive activity and was able to grow aerobically on 2,4-dinitrophenol as the
sole carbon source. This bacterium showed maximal growth on 2,4-DNP at the temperature
optimum of 30 oC, pH 5.0 and was tolerant to 2,4-DNP concentration of up to 0.5 mM (0.092
g/L). This bacterium prefers to use urea as the nitrogen source in addition to yeast extract for
mineral source and vitamin precursors.
Contamination of organic xenobiotic pollutants and heavy metals in a contaminated site allows
the use of multiple bacterial degraders or bacteria with the ability to detoxify numerous toxicants
at the same time. A previously isolated SDS- degrading bacterium, Acinetobacter baumannii
strain Serdang 1 was shown to reduce molybdenum to molybdenum-blue. The bacterium works
optimally at pH 6.5, the temperature range between 25 and 34°C with glucose serves as the best
electron donor for molybdate reduction. This bacterium required additional concentration of
phosphate at 5.0 mM and molybdate between 15 and 25 mM. The absorption spectrum of the
molybdenum blue obtained is similar to the molybdenum blue from other earlier reported
molybdate reducing bacteria, as it resembles a reduced phosphomolybdate closely. Ag(i), As(v),
Pb(ii) and Cu(ii) inhibited molybdenum reduction by 57.3, 36.8, 27.7 and 10.9%, respectively, at
1 p.p.m. Acrylamide was efficiently shown to support molybdenum reduction at a lower
efficiency than glucose. Phenol, acrylamide and propionamide could support the growth of this
bacterium independently of molybdenum reduction. This bacterium capability to detoxify several
toxicants is an important tool for bioremediation in the tropical region.
The growth of microorganism on substrates, whether toxic or not usually exhibits sigmoidal
pattern. This sigmoidal growth pattern can be modelled using primary models such as Logistic,
modified Gompertz, Richards, Schnute, Baranyi-Roberts, Von Bertalanffy, Buchanan threephase
and Huang. Previously, the modified Gompertz model was chosen to model the growth of
Burkholderia sp. strain Neni-11 on acrylamide, which shows a sigmoidal curve. The modified
Gompertz model relies on the ordinary least squares method, which in turn relies heavily on
several important assumptions, which include that the data does not show autocorrelation. In this
work we perform statistical diagnosis test to test for the presence of autocorrelation using the
Durbin-Watson test and found that the model was adequate and robust as no autocorrelation of
the data was found.
Mass production of fish broodstock with high quality eggs requires the knowledge on the chemical composition and physiochemical properties of vitellogenin (Vtg) during ovulation. Vtg is an egg yolk precursor phospholipoglycoprotein, and has been analysed to evaluate the reproductive conditions and determine the spawning period in captive and wild fish. In this study, Vtg was induced in male H. nemurus through three intramuscular injections of 17-estradiol (E2). The Vtg was purified from the serum using gel filtration chromatography and the purified protein was reduced via SDS-PAGE. One major polypeptide corresponding to 130 kDa was observed. Vtg identification was done using peptide mass fingerprint (PMF) from the trypsin digestion of male H. nemurus Vtg induced with E2. The sequence homology of H. nemurus AYLAGAAADVLEVGVR matched the Vtg of other fish species when analysed using MALDI-TOF. Vtg was confirmed by MASCOT at 95% significant level. The potential protein that controls the reproductive process and oocyte development isolated from this study was discussed to understand the structure and function of Vtg.
Quinolines compounds are toxic pollutants. Their biodegradation by microbes represents a tool
for bioremediation. The growth of Klebsiella penumoniae on 2-methylquinoline shows typical
sigmoidal bacterial growth curves. Since there exists a variety of models for describing the
growth profile of microorganism such as logistic, Gompertz, Richards, Schnute, Baranyi-
Roberts, Von Bertalanffy, Buchanan three-phase and more recently Huang models, the growth
curves exhibit under such conditions would be an excellent study for finding the best model.
The Huang model was chosen as the best model based on statistical tests such as root-meansquare
error (RMSE), adjusted coefficient of determination (R2), bias factor (BF), accuracy
factor (AF) and corrected AICc (Akaike Information Criterion). Novel constants obtained from
the modelling exercise would be used for further secondary modelling.
The trend of global under-five child mortality rate showed a steady decrease at -3.14% per year
approaching the United Nations millennium development goals target (-4.0% reduction per year
from 1990-2015). This rate is usually inversely proportional to the nations GDP. However, the
progress in the recent years (2006-2012) in Malaysia has raised a disturbing concern with an
annual average increment of +0.81% per year. Thus, there is an urgent need to identify the
reasons of such risky ecosystem. By analyzing recorded data on river pollution from 2007-
2012, we found that river pollution has high correlation with the gross domestic products, not at
the same year, but for two years prior that indicates a lag time. We also found that under-five
child mortality has strong correlation with the river pollutions. The development-pollutionhealth
triangular cycle needs to be put in a balanced to ensure the nation prosperity and
sustainability of the nation.
Bioremediation is a new green economic approach in providing solutions for cleaning up
contaminated sites. Phytoremediation uses plants as a tool for remediation purposes. The usage
of plant species offers higher potential solution to remediate heavy metal contaminated sites.
This study aimed on screening potential plant species for phytoremediation of heavy metal
contaminated water. The potential of three aquatic macrophytes species (Eichorrnia crassipes,
Pistia stratiotes and Ipomoea aquatica) for chromium and nickel phytoremediations was tested.
The plants were exposed for 10 days under hydroponic conditions in heavy metal contaminated
water. E. crassipes showed the highest chromium and nickel concentrations in its biomass, 1.60
and 2.40 μg/L respectively. Meanwhile, P. stratiotes had chromium and nickel concentrations
detected at 0.89 and 0.081 μg/L, respectively; chromium and nickel concentrations of I.
aquatica detected were, 0.49 and 0.08 μg/L, respectively. The ability of these plants to
accumulate heavy metals and survived throughout the experiment demonstrates the potential of
these plants to remediate metal-enriched water. Among the three tested aquatic plants, E.
crassipes was proven to be the most suitable plant species that can phytoremediate heavy metal
contaminated water followed by P. stratiotes and I. aquatica.
Protein function depends greatly on its structure. Based on this principle, it is vital to study the
protein structure in order to understand its function. This study attempts to build the predicted
model of lipase gene in Rhodococcus sp. NAM81 using homology modelling method. The
predicted structure was then used to investigate the function of protein through several
bioinformatic tools. The DNA sequence of lipase gene was obtained from the Rhodococcus sp.
NAM81 genome scaffold. Blastx analysis showed 100% identity to the target enzyme andthe
appropriate template for homology modelling was determined using Blastp analysis. The 3D
protein structure was built using two homology modelling software, EsyPred3D and Swiss
Model Server. Both structures built obtained LGScore of greater than 4, which means they are
extremely good models according to ProQ validation criteria. Both structures also satisfied the
Ramachandran plot structure validation analysis. The predicted structures were 100% matched
with each other when superimposed with DaliLite pairwise. This shows that both structure
validation servers agreed on the same model. Structure analysis using ProFunc had found seven
motifs and active sites that indicate similar function of this protein with other known proteins.
Thus, this study has successfully produced a good 3D protein structure for the target enzyme.
Most often than not, microorganism’s growth curve is sigmoidal in characteristics.
The modified Gompertz model via nonlinear regression using the least square method
is one of the most popular methods to describe the growth curve. One of the
assumptions of a good model is that the variance of the data must be homogenous
(homoscedasticity). In this work, two statistical diagnostics; the Bartlett and the
Levene’s tests was performed to a modified Gompertz model utilized to model the
growth of the bacterium Burkholderia sp. strain Neni-11 on acrylamide in order to
satisfy the requirement above and found that data conformed to the requirement
indicating the modified Gompertz model is a robust model for modelling the bacterial
growth process.
The pollution of heavy metals and toxic xenobiotics has become a central issue worldwide.
Bioremediation of these toxicants are being constantly carried out using novel microbes.
Molybdenum reduction to molybdenum blue is a detoxification process and mathematical
modelling of the reduction process can reveal important parameters such as specific reduction
rate, theoretical maximum reduction and whether reduction at high molybdenum concentration
affected the lag period of reduction. The used of linearization method through the use of natural
logarithm transformation, although popular, is inaccurate and can only give an approximate
value for the sole parameter measured; the specific growth rate. In this work, a variety of
models for such as logistic, Gompertz, Richards, Schnute, Baranyi-Roberts, Von Bertalanffy,
Buchanan three-phase and more recently Huang were utilized for the first time to obtain values
for the above parameters or constants. The modified Gompertz model was the best model in
modelling the Mo-blue production curve from Serratia marcescens strain DR.Y10 based on
statistical tests such as root-mean-square error (RMSE), adjusted coefficient of determination
(R2), bias factor (BF), accuracy factor (AF) and corrected AICc (Akaike Information Criterion).
Parameters obtained from the fitting exercise were maximum Mo-blue production rate (μm), lag
time (l) and maximal Mo-blue production (Ymax) of X (h-1), Y (h) and Z (nmole Mo-blue),
respectively. The application of primary population growth models in modelling the Moblue
production rate from this bacterium has become a successful undertaking. The model
may also be used in other heavy metals detoxification processes. The parameters
constants extracted from this work will be a substantial help for the future development
of further secondary models.
The volume of contaminated rivers in Malaysia continues to keep rising through the years. The
cost of instrumental monitoring is uneconomical and prohibits schedule monitoring of
contaminants particularly heavy metals. In this work, a rapid enzyme assay utilizing the
molybdenum-reducing enzyme as an inhibitive assay, prepared in crude form from the
molybdenum-reducing bacterium Serratia sp. strain DRY5 has been developed for monitoring
the heavy metals mercury, silver, copper and chromium in contaminated waters in the Juru
Industrial Estate. The crude enzyme extract transformed soluble molybdenum
(phosphomolybdate) into a deep blue solution, which is inhibited by heavy metals such as
mercury, silver, copper and chromium. The IC50 and Limits of Detection (LOD) values for
mercury, copper, silver and cadmium were 0.245, 0.298, 0.367, 0.326, and 0.124, 0.086, 0.088
and 0.094 mg L-1, respectively. The assay is rapid, and can be carried out in less than 10 minutes.
In addition, the assay can be carried out at ambient temperature. The IC50 values for these heavy
metals are more sensitive than several established assays. Water samples from various locations
in the month of November from the Juru Industrial Estate (Penang) were tested for the presence
of heavy metals using the developed assay. Enzyme activity was nearly inhibited for water
samples from several locations. The presence of heavy metals was confirmed instrumentally
using Atomic Emission Spectrometry and a Flow Injection Mercury System. The assay is rapid
and simple and can be used as a first screening method for large scale monitoring of heavy
metals.
Chemical toxins and organic contaminants such as hydrocarbons and dyes are major global
contaminants with countless tones of those chemicals are created yearly with a significant
amount release to the environment. In this work we screen the ability of a molybdenum-reducing
bacterium isolated from contaminated soil to decolorize various azo and triphenyl methane dyes
independent of molybdenum reduction. Biochemical analysis resulted in a tentative identification
of the bacterium as Enterobacter sp. strain Zeid-6. The bacterium was able to decolorize the azo
dye Orange G. The bacterium reduces molybdate to Mo-blue optimally at pH between 5.5 and
8.0 and temperatures of between 30 and 37 oC. Other requirements include a phosphate
concentration of 5 mM and a molybdate concentration of 20 mM. The absorption spectrum of the
Mo-blue produced was similar to previous Mo-reducing bacterium, and closely resembles a
reduced phosphomolybdate. Molybdenum reduction was inhibited by copper, lead, mercury and
silver which showed 36.8, 16.9, 64.9 and 67.6% inhibition to Mo-reducing activity of
Enterobacter sp. strain Zeid-6, respectively. The resultant molybdenum blue spectrum closely
resembles the spectrum of molybdenum blue from the phosphate determination method. The
ability of this bacterium to detoxify molybdenum and decolorize azo dye makes this bacterium
an important tool for bioremediation.
The 3D structure of the insecticidal protein Cry1Ba4 produced by B. thuringiensis subsp.
Entomocidus HD-9 was determined using homology modelling. From the model built, we have
been able to identify the possible sites for structure modification by site-directed mutagenesis.
The mutation was introduced at the conserved region of -helix 7 by substituting the
hydrophobic motif that comprises alanine 216, leucine 217 and phenylalanine 218 with arginine.
Wild and mutant Cry1Ba4 genes were cloned into pET200/D-TOPO and expressed in the
expression host. The result suggests that mutant Cry1Ba4 protein was less toxic to the larvae
Plutella xylostella compared to the wild-type. In conclusion, alteration in the structure of
Domain I had left an impact on the toxicity of Cry1Ba4 against P. xylostella.
In this work, a temporal monitoring work for heavy metals from an effluent discharge point in
the Juru Industrial Estate was carried out using the protease extracted from garlic (Allium
sativum) as the principal bioassay system. casein-Coomassie-dye binding assay method has
utilized this purpose. The periodic sampling results for one day of a location in the Juru
Industrial Estate showed temporal variation of copper concentration coinciding with garlic
protease inhibition with the highest concentrations of copper occurring between 12.00 and 16.00
hours of between 3 and 3.5 mg/L copper. The crude proteases extracted from Allium sativum
successfully detect temporal variation of copper form this location. In conclusion, this assay
method has the potential to be a rapid, sensitive, and economic inhibitive assay for the largescale
biomonitoring works for the heavy metal copper from this area.
Molybdenum is an emerging pollutant. Bioremediation of this heavy metal is possible by the
mediation of Mo-reducing bacteria. These bacteria contain the Mo-reducing enzymes that can
conver toxic soluble molybdenum into molybdenum blue; a less soluble and less toxic form of the
metal. To date only the enzyme has been purified from only one bacterium. The aim of this study is
to purify the Mo-reducing enzyme from a previously isolated Mo-reducing bacterium Bacillus
pumilus strain Lbna using ammonium sulphate fractionation followed by ion exchange and then
gel filtration. Two clear bands were obtained after the gel filtration step with molecular weights
of 70 and 100 kDa. This indicates that further additional purification methods need to be used
to get a purified fraction. Hence, additional steps of chromatography such as hydroxyapatite or
chromatofocusing techniques can be applied in the future.
Phenolic compounds or phenols are a group of aromatic compounds that comprises a hydroxyl
group (OH) that is directly bonded to an aromatic ring. Phenols are injurious to organisms even
at even low concentrations with many of them are categorized as dangerous pollutants because of
their likely harm to human well-being. This review attempts to discuss the various merits and
demerits of immobilization matrices employed for phenol-degrading microorganisms’
immobilization. One of several key points of cellular immobilization is the capacity to protect
bioremediation agents towards toxic levels of specific toxicants and safeguarding from predatory
microorganisms. However, this shielding course of action should never impede the diffusion of
substrates into the pores of the immobilization structure. In the end the choice of a particular
immobilization method will strongly hinge on aspects of economy, safety and efficacy.
In this study, a novel glyphosate-degrading shows the ability to reduce molybdenum to
molybdenum blue. The enzyme from this bacterium was partially purified and partially
characterized to ascertain whether the Mo-reducing enzyme from this bacterium shows better or
lower efficiency in reducing molybdenum compared to other Mo-reducing bacterium that only
exhibits a single biotransformation activity. The enzyme was partially purified using ammonium
sulphate fractionation. The Vmax for the electron donating substrate or NADH was at 1.905 nmole
Mo blue/min while the Km was 6.146 mM. The regression coefficient was 0.98. Comparative
assessment with the previously characterized Mo-reducing enzyme from various bacteria showed
that the Mo-reducing enzyme from Burkholderia vietnamiensis strain AQ5-12 showed a lower
enzyme activity.
Observations on the effects of copper on the liver proteome of Puntius javanicus based on the
one dimensional PAGE was carried out. The liver was dissected from each fish, which was
separately treated with different concentrations of copper sulfate ranging from 0.1 to 5.0 mg/L.
The livers were extracted and one dimensional PAGE was performed under nonreducing
(native) and reducing (SDS)-PAGE. Several bands were resolved in the native PAGE with
probable candidates for the effect of copper observed showing an increased in the expression
and downregulation strongly associated with increasing copper concentrations. This study
showed that high concentrations of copper significantly alters P. javanicus liver at the proteome
level, and preliminary screening based on one dimensional PAGE is considered rapid and
simple to assess the toxicity effect of copper before more advanced and extensive assesment
with a second dimensional PAGE is carried out.
Investigation on in vivo effects of copper (Cu) on the ultrastructure of P. javanicus liver was
carried out using transmission electron microscopy (TEM). The addition of sublethal
concentration of 5 mg/L of Cu caused abnormalities on the bile canaliculi (BC) including
dilation and elongation compared to control and at lower concentrations of copper with a normal
round shape form. Findings from this study support an alternative histological assessment of the
effects of Cu concentration on P. javanicus liver.
Water contamination by herbicides and chelating agents is increasing mainly due to the
increasing agricultural activities. Water contamination by these compounds has become a
concern due to their adverse effects to the environment and humans. Seven sampling sites of
water sources in Selangor and Johor were chosen for the study. Contamination level of
Mecoprop (MCCP), Nitrilotriacetic acid (NTA) and Ethylenediaminetetraacetic acid (EDTA) in
these water body areas was determined by using Gas Chromatography-Electron Capture
Detector (GC-ECD). Our results indicated that water samples of Sungai Melot in Selangor
showed the highest presence of EDTA. MCCP was detected at a high level at Sungai Sarang
Buaya, Johor while NTA showed similar level of concentration at three different sites, Ladang
10, Ladang Sayur and Mardi, Selangor.