Hypercholesterolemia is the major risk factor that leads to atherosclerosis. Nowadays, alternative treatment using medicinal plants gained much attention since the usage of statins leads to adverse health effects, especially liver and muscle toxicity. This study was designed to investigate the hypocholesterolemic and antiatherosclerotic effects of Basella alba (B. alba) using hypercholesterolemia-induced rabbits. Twenty New Zealand white rabbits were divided into 5 groups and fed with varying diets: normal diet, 2% high cholesterol diet (HCD), 2% HCD + 10 mg/kg simvastatin, 2% HCD + 100 mg/kg B. alba extract, and 2% HCD + 200 mg/kg B. alba extract, respectively. The treatment with B. alba extract significantly lowered the levels of total cholesterol, LDL, and triglycerides and increased HDL and antioxidant enzymes (SOD and GPx) levels. The elevated levels of liver enzymes (AST and ALT) and creatine kinase were noted in hypercholesterolemic and statin treated groups indicating liver and muscle injuries. Treatment with B. alba extract also significantly suppressed the aortic plaque formation and reduced the intima: media ratio as observed in simvastatin-treated group. This is the first in vivo study on B. alba that suggests its potential as an alternative therapeutic agent for hypercholesterolemia and atherosclerosis.
A molybdenum-reducing bacterium from Antarctica has been isolated. The bacterium converts sodium molybdate or Mo⁶⁺ to molybdenum blue (Mo-blue). Electron donors such as glucose, sucrose, fructose, and lactose supported molybdate reduction. Ammonium sulphate was the best nitrogen source for molybdate reduction. Optimal conditions for molybdate reduction were between 30 and 50 mM molybdate, between 15 and 20°C, and initial pH between 6.5 and 7.5. The Mo-blue produced had a unique absorption spectrum with a peak maximum at 865 nm and a shoulder at 710 nm. Respiratory inhibitors such as antimycin A, sodium azide, potassium cyanide, and rotenone failed to inhibit the reducing activity. The Mo-reducing enzyme was partially purified using ion exchange and gel filtration chromatography. The partially purified enzyme showed optimal pH and temperature for activity at 6.0 and 20°C, respectively. Metal ions such as cadmium, chromium, copper, silver, lead, and mercury caused more than 95% inhibition of the molybdenum-reducing activity at 0.1 mM. The isolate was tentatively identified as Pseudomonas sp. strain DRY1 based on partial 16s rDNA molecular phylogenetic assessment and the Biolog microbial identification system. The characteristics of this strain would make it very useful in bioremediation works in the polar and temperate countries.
Heavy metals pollution has become a great threat to the world. Since instrumental methods are expensive and need skilled technician, a simple and fast method is needed to determine the presence of heavy metals in the environment. In this study, an inhibitive enzyme assay for heavy metals has been developed using crude proteases from Coriandrum sativum. In this assay, casein was used as a substrate and Coomassie dye was used to denote the completion of casein hydrolysis. In the absence of inhibitors, casein was hydrolysed and the solution became brown, while in the presence of metal ions such as Hg²⁺ and Zn²⁺, the hydrolysis of casein was inhibited and the solution remained blue. Both Hg²⁺ and Zn²⁺ exhibited one-phase binding curve with IC₅₀ values of 3.217 mg/L and 0.727 mg/L, respectively. The limits of detection (LOD) and limits of quantitation (LOQ) for Hg were 0.241 and 0.802 mg/L, respectively, while the LOD and LOQ for Zn were 0.228 and 0.761 mg/L, respectively. The enzyme exhibited broad pH ranges for activity. The crude proteases extracted from Coriandrum sativum showed good potential for the development of a rapid, sensitive, and economic inhibitive assay for the biomonitoring of Hg²⁺ and Zn²⁺ in the aquatic environments.
Phaleria macrocarpa (Scheff.) Boerl (Thymelaceae) originates from Papua Island, Indonesia and grows in tropical areas. The different parts of the fruit of P. macrocarpa were evaluated for antioxidant, anti-inflammatory, and cytotoxic activities.
Phaleria macrocarpa (Scheff.) Boerl (Thymelaceae) is commonly known as 'Crown of God', 'Mahkota Dewa', and 'Pau'. It originates from Papua Island, Indonesia and it grows in tropical areas. Empirically, it is potent in treating the hypertensive, diabetic, cancer and diuretic patients. It has a long history of ethnopharmacological usage, and the lack of information about its biological activities led us to investigate the possible biological activities by characterisation of flavonoids and antimicrobial activity of various part of P. macrocarpa against pathogenic bacteria and fungi. The results showed that kaempferol, myricetin, naringin, and rutin were the major flavonoids present in the pericarp while naringin and quercetin were found in the mesocarp and seed. Furthermore, the antibacterial activity of different parts of P. macrocarpa fruit showed a weak ability to moderate antibacterial activity against pathogenic tested bacteria (inhibition range: 0.93-2.17 cm) at concentration of 0.3 mg/disc. The anti fungi activity was only found in seed extract against Aspergillus niger (1.87 cm) at concentration of 0.3 mg/well. From the results obtained, P. macrocarpa fruit could be considered as a natural antimicrobial source due to the presence of flavonoid compounds.
An inhibitive assay of insecticides using Acetylcholinesterase (AChE) from the local fish Clarias batrachus is reported. AChE was assayed according to the modified method of Ellman. Screening of insecticide and heavy metals showed that carbofuran and carbaryl strongly inhibited C. batrachus AChE. The inhibition concentration (IC) IC50 values (and the 95% confidence interval) for both carbofuran and carbaryl inhibition on C. batrachus AChE at 6.66 (5.97-7.52) and 130.00 (119.3-142.5) microg l(-1), respectively was within the IC50 range of Electrophorus electricus at 6.20 (6.03-6.39) and 133.01 (122.40-145.50) microg l(-1), respectively and were much lower than bovine AChE at 20.94 (19.53-22.58) and 418.80 (390.60-451.60) microg l(-1), respectively. The results showed that C. batrachus have the potential to be used as a cheaper and more readily available source of AChE than other more commercially available sources.
A stab-culture method was adapted to screen for azo dyes-decolorizing bacteria from soil and water samples. Decolorized azo dye in the lower portion of the solid media indicates the presence of anaerobic azo dyes-decolorizing bacteria, while aerobic decolorizing bacteria decolorizes the surface portion of the solid media. Of twenty soil samples tested, one soil sample shows positive results for the decolourisation of two azo dyes; Biebrich scarlet (BS) and Direct blue 71 (DB) under anaerobic conditions. A gram negative and oxidase negative bacterial isolate was found to be the principal azo dyes degrader The isolate was identified by using the Biolog identification system as Serratia marcescens.
In this work, a subtractive inhibition assay (SIA) based on surface plasmon resonance (SPR) for the rapid detection of Campylobacter jejuni was developed. For this, rabbit polyclonal antibody with specificity to C. jejuni was first mixed with C. jejuni cells and unbound antibody was subsequently separated using a sequential process of centrifugation and then detected using an immobilized goat anti-rabbit IgG polyclonal antibody on the SPR sensor chip. This SIA-SPR method showed excellent sensitivity for C. jejuni with a limit of detection (LOD) of 131 ± 4 CFU mL-1 and a 95% confidence interval from 122 to 140 CFU mL-1. The method has also high specificity. The developed method showed low cross-reactivity to bacterial pathogens such as Salmonella enterica serovar Typhimurium (7.8%), Listeria monocytogenes (3.88%) and Escherichia coli (1.56%). The SIA-SPR method together with the culturing (plating) method was able to detect C. jejuni in the real chicken sample at less than 500 CFU mL-1, the minimum infectious dose for C. jejuni while a commercial ELISA kit was unable to detect the bacterium. Since the currently available detection tools rely on culturing methods, which take more than 48 hours to detect the bacterium, the developed method in this work has the potential to be a rapid and sensitive detection method for C. jejuni.
In this work, we report on the isolation of a phenol-degrading Rhodococcus sp. with a high tolerance towards phenol. The isolate was identified as Rhodococcus sp. strain AQ5NOL 2, based on 16S rDNA analysis. The strain degraded phenol using the meta pathway, a trait shared by many phenol-degraders. In addition to phenol biodegradation, the strain was also capable of degrading diesel. Strain AQ5NOL 2 exhibited a broad optimum temperature for growth on phenol at between 20 °C and 35 °C. The best nitrogen sources were ammonium sulphate, glycine or phenylalanine, followed by proline, nitrate, leucine, and alanine (in decreasing efficiency). Strain AQ5NOL 2 showed a high tolerance and degradation capacity of phenol, for it was able to register growth in the presence of 2000 mg l(-1) phenol. The growth of this strain on phenol as sole carbon and energy source were modeled using Haldane kinetics with a maximal specific growth rate (μ(max)) of 0.1102 hr(-1), a half-saturation constant (K(s) ) of 99.03 mg l(-1) or 1.05 mmol l(-1), and a substrate inhibition constant (K(i)) of 354 mg l(-1) or 3.76 mmol l(-1). Aside from phenol, the strain could utilize diesel, 2,4-dinitrophenol and ρ-cresol as carbon sources for growth. Strain AQ5NOL 2 exhibited inhibition of phenol degradation by Zn(2+), Cu(2+), Cr(6+), Ag(+) and Hg(2+) at 1 mg l(-1).
As well as for chemical and environmental reasons, acrylamide is widely used in many industrial applications. Due to its carcinogenicity and toxicity, its discharge into the environment causes adverse effects on humans and ecology alike. In this study, a novel acrylamide-degrading yeast has been isolated. The isolate was identified as Rhodotorula sp. strain MBH23 using ITS rRNA analysis. The results showed that the best carbon source for growth was glucose at 1.0% (w/v). The optimum acrylamide concentration, being a nitrogen source for cellular growth, was at 500 mg l(-1). The highest tolerable concentration of acrylamide was 1500 mg l(-1) whereas growth was completely inhibited at 2000 mg l(-1). At 500 mg l(-1), the strain MBH completely degraded acrylamide on day 5. Acrylic acid as a metabolite was detected in the media. Strain MBH23 grew well between pH 6.0 and 8.0 and between 27 and 30 °C. Amides such as 2-chloroacetamide, methacrylamide, nicotinamide, acrylamide, acetamide, and propionamide supported growth. Toxic heavy metals such as mercury, chromium, and cadmium inhibited growth on acrylamide.
Extensive use of metals in various industrial applications has caused substantial environmental pollution. Molybdenum-reducing bacteria isolated from soils can be used to remove molybdenum from contaminated environments. In this work we have isolated a local bacterium with the capability to reduce soluble molybdate to the insoluble molybdenum blue. We studied several factors that would optimize molybdate reduction. Electron donor sources such as glucose, sucrose, lactose, maltose and fructose (in decreasing efficiency) supported molybdate reduction after 24 h of incubation with optimum glucose concentration for molybdate reduction at 1.5% (w/v). The optimum pH, phosphate and molybdate concentrations, and temperature for molybdate reduction were pH 6.5, 5.0, 25 to 50 mM and 37 degrees C, respectively. The Mo-blue produced by cellular reduction exhibited a unique absorption spectrum with a maximum peak at 865 nm and a shoulder at 700 nm. Metal ions such as chromium, cadmium, copper, silver and mercury caused approximately 73, 71, 81, 77 and 78% inhibition of the molybdenum-reducing activity, respectively. All of the respiratory inhibitors tested namely rotenone, azide, cyanide and antimycin A did not show any inhibition to the molybdenum-reducing activity suggesting components of the electron transport system are not responsible for the reducing activity. The isolate was tentatively identified as Enterobacter sp. strain Dr.Y13 based on carbon utilization profiles using Biolog GN plates and partial 16S rDNA molecular phylogeny.
The development of glyphosate-resistant genetically modified organisms (GMO) has increased the use of herbicide glyphosate by several magnitudes in recent years. It is now the most commonly used pesticide globally that affects aquatic habitats, especially fish. This study aims to add new knowledge on the effect of technical grade glyphosate on several toxicity parameters and to identify the most effective parameter in predicting technical grade glyphosate chronic toxicity (seven weeks) to fish, especially Malaysia's heavily farmed red tilapia. The results show that a relatively high concentration of technical grade glyphosate is needed to induce significant changes in all tested parameters. However, the results also indicate that the bodyweight index is the most sensitive toxicity parameter in that a reduction in body weight was observed at 25 mg/L of glyphosate. Negative correlations between the glyphosate concentration and toxicity parameters such as specific growth rate (SGR), hepato-somatic index (HIS), and gonado-somatic index (GSI) were observed. The fish condition factor and feed conversion ratio were found not to be affected at the highest glyphosate concentration tested (150 mg/L). To conclude, crossbred red tilapia (O. niloticus × O. mossambicus) is one potential species for evaluating the toxic effects of technical grade glyphosate on fish.
A local molybdenum-reducing bacterium was isolated and tentatively identified as Acinetobacter calcoaceticus strain Dr.Y12 based on carbon utilization profiles using Biolog GN plates and 16S rDNA comparative analysis. Molybdate reduction was optimized under conditions of low dissolved oxygen (37 degrees C and pH 6.5). Of the electron donors tested, glucose, fructose, maltose and sucrose supported molybdate reduction after 1 d of incubation, glucose and fructose supporting the highest Mo-blue production. Optimum Mo-blue production was reached at 20 mmol/L molybdate and 5 mmol/L phosphate; increasing the phosphate concentrations inhibited the production. An increase in an overall absorption profiles, especially at peak maximum at 865 nm and the shoulder at 700 nm, was observed in direct correlation with the increased in Mo-blue amounts. Metal ions, such as chromium, cadmium, copper, mercury and lead (2 mmol/L final concentration) caused approximately 88, 53, 80, 100, and 20 % inhibition, respectively. Respiratory inhibitors, such as antimycin A, rotenone, sodium azide and cyanide showed in this bacterium no inhibition of the Mo-blue production, suggesting that the electron transport system is not a site of molybdate reduction.
Arsenic is a toxic metalloid which is widely distributed in nature. It is normally present as arsenate under oxic conditions while arsenite is predominant under reducing condition. The major discharges of arsenic in the environment are mainly due to natural sources such as aquifers and anthropogenic sources. It is known that arsenite salts are more toxic than arsenate as it binds with vicinal thiols in pyruvate dehydrogenase while arsenate inhibits the oxidative phosphorylation process. The common mechanisms for arsenic detoxification are uptaken by phosphate transporters, aquaglyceroporins, and active extrusion system and reduced by arsenate reductases via dissimilatory reduction mechanism. Some species of autotrophic and heterotrophic microorganisms use arsenic oxyanions for their regeneration of energy. Certain species of microorganisms are able to use arsenate as their nutrient in respiratory process. Detoxification operons are a common form of arsenic resistance in microorganisms. Hence, the use of bioremediation could be an effective and economic way to reduce this pollutant from the environment.
A novel molybdate-reducing bacterium, tentatively identified as Klebsiella sp. strain hkeem and based on partial 16s rDNA gene sequencing and phylogenetic analysis, has been isolated. Strain hkeem produced 3 times more molybdenum blue than Serratia sp. strain Dr.Y8; the most potent Mo-reducing bacterium isolated to date. Molybdate was optimally reduced to molybdenum blue using 4.5 mM phosphate, 80 mM molybdate and using 1% (w/v) fructose as a carbon source. Molybdate reduction was optimum at 30 °C and at pH 7.3. The molybdenum blue produced from cellular reduction exhibited absorption spectrum with a maximum peak at 865 nm and a shoulder at 700 nm. Inhibitors of electron transport system such as antimycin A, rotenone, sodium azide, and potassium cyanide did not inhibit the molybdenum-reducing enzyme. Mercury, silver, and copper at 1 ppm inhibited molybdenum blue formation in whole cells of strain hkeem.
The main strategy for lowering blood cholesterol levels is through the inhibition of the NADPH-dependent HMG-CoA reductase (3-hydroxy-3-methyl-glutaryl-CoA reductase). The enzyme catalyses the reduction of HMG-CoA to mevalonate and this process is inhibited by statins that form the bulk of the therapeutic agents to treat high cholesterol since the 1970s. Newer drugs that are safer than statins are constantly being developed. The inhibition of candidate drugs to HMG-CoA reductase remains the mainstay of drug development research. The determination of the enzyme activity is important for the correct assessment of potency of the enzyme as well as determining the inhibition of potential therapeutic agents from the plant and microbial extracts. Also, this chapter covers the use of the popular four-parameter logistics model that can yield accurate estimation of the IC50 values of therapeutic agents and their 95% confidence intervals.