Purification and physicochemical characterisation of Aspergillus niger USM F4 β-mannanase

Aims@#This present study focused on purification of fungal β-mannanase produced by Aspergillus niger USM F4 and also physicochemical characterisation of the purified enzyme.@*Methodology and results@#The purified β-mannanase with a molecular mass of ~47.4 kDa was demonstrated on SDSPAGE gel. The enzyme signified a purification degree of 4-fold, with final specific activity of 196.42 U/mg. It reached an optimum catalytic activity at pH 4.0 and 60 °C. The thermal stability of the enzyme was up to 70 °C and maintained the 50% activity after 30 min at 80 °C. Meanwhile, the pH stability was in the range of pH 3.0-9.0 and a 30 min half-life at pH 10.0. All chemical substances manifested an inhibitory effect on purified β-mannanase, with SDS (28.16 ± 0.05% residual activity) as the strongest inhibitor, followed by cupric ion (Cu2+) (49.51 ± 0.09% residual activity). As a whole, the enzyme displayed a substrate specificity in the order of locust bean gum (LBG) > carboxymethylcellulose > soluble starch > xylan from oat spelt > α-cellulose. Its preference for LBG has generated the Km and Vmax values of 0.20 mg/mL and 9.82 U/mL, respectively.@*Conclusion, significance and impact of study@#The outcomes of our study offer potential for use at industrial scales, particularly in the oligosaccharides production that involve acid-related activity, wide-ranging temperature and pH stability.

In vitro antimicrobial activities of methanolic extract from marine alga Enteromorpha intestinalis

To extract the bioactive compound from Enteromorpha intestinalis (E. intestinalis) and determine its in vitro antimicrobial activity. Methods: E. intestinalis was extracted by methanol and subjected to antimicrobial screening. The antimicrobial activity was studied by using disc diffusion and broth dilution method. The effect of the extract on the growth profile of the bacterial was also examined via time-kill assay. Microscopy observations using SEM was done to determine the major alterations in the microstructure of methicillin-resistant Staphylococcus aureus (MRSA). Results: The results showed methanolic extract of E. intestinalis exhibited a favourable antimicrobial activity against tested bacteria with produced inhibition zone ranging from 8.0-19.0 mm. However, all the tested fungi and yeast were resistant to the extract treatment. Time kill assay suggested that methanolic extract of E. intestinalis had completely inhibited MRSA growth and also exhibited prolonged antibacterial activity. The main abnormalities noted from the microscopic observations were the structural deterioration in the normal morphology and complete collapsed of the bacteria cells after 36 h of treatment. Conclusions: The significant antibacterial activity shown by crude extract suggested its potential against MRSA infection. The extract may have potential to develop as antibacterial agent in pharmaceutical use.

Prodigiosin - an antibacterial red pigment produced by Serratia marcescens IBRL USM 84 associated with a marine sponge Xestospongia testudinaria.

The objectives of present study are to extract the antibiotic compound from marine isolate and to determine its in vitro antimicrobial activity against bacteria. A marine bacterial isolate Serratia marcescens IBRL USM 84 was isolated from the surface of a marine sponge Xestospongia testudinaria. This species of bacteria produced red pigment with antibacterial activity. The red antibacterial pigment was produced intracellularly and inhibited 13 out of 18 tested bacteria, with Gram positive was more susceptible than the Gram negative bacteria. The growth and antibacterial red pigment production profiles demonstrated the highest antibacterial red pigment production was achieved at the 48 hours of cultivation (14.08 U/ml) time in marine broth when incubated at 25 °C with 150 rpm agitation. The antibacterial red pigment was extracted, purified and confirmed as prodigiosin.

In vitro activity of methanolic extract from Lagerstroemia speciosa (Linn. ex. Murray) bark against pathogenic bacteria.

Crude methanolic extract of Lagerstroemia speciosa barks (Linn. ex. Murray) was subjected to antimicrobial screening including six Gram-positive and eight Gram-negative bacteria. The extract demonstrated significance antibacterial activities on 5 tested bacteria with the inhibition zone ranging from 10-15 mm. The minimum inhibitory concentration (MIC) of the extract was assessed by microdilution method. MIC value of Bacillus spizizenii ATCC6633, Bacillus cereus and Streptococcus coagulase-negative (SCN) was 0.25 mg/ml whilst Bacillus licheniformis and Acinetobacter anitratus showed MIC value of 0.50 mg/ml. However, only B. spizizenii ATCC6633 and B. licheniformis showed the minimum bactericidal concentration (MBC) value of 2.00 mg/ml and 1.00 mg/ml, respectively. The morphological changes of B.spizizenii ATCC6633 and A. anitratus with the treatment of the extract were observed under Scanning Electron Microscope (SEM). The results suggested that the L. speciosa methanolic extract had caused membrane structural degenerations of the cells and finally leading to cell-lysis.

Efficacy of pyroligneous acid from Rhizophora apiculata on pathogenic Candida albicans.

Rhizophora apiculata pyroligneous acid which is a crude condensate produced from the distillation of smoke generated in the process of charcoal making has the potential to be used as antifungal agent especially to treat candidal infections. In this study, pyroligneous acid (PA), concentrated pyroligneous acid (CPA), Dichloromethane extracts of CPA namely DCM A and B were tested against four pathogenic strains of Candida albicans. The results exhibited significant inhibition zones within the range of 7.00 -8.00 mm for PA, 16.00- 17.00 mm for CPA, 16.00-18.00 mm for DCM A and 19.00-22.00 mm for DCM B. The results also revealed that extract DCM B of CPA was the most potential to be used as anticandidal agent with the minimum inhibitory concentration values between 3.13-6.25 mg/mL. Scanning electron micrographs of DCM B treated C. albicans cells confirmed the damaged cells caused by the extract.

Effect of Cultural Conditions on Lovastatin Production by Aspergillus Niger Sar I Using Combination of Rice Bran and Brown Rice as Substrate.

A local fungal isolate Aspergillus niger SAR I produced high level of lovastatin activity when cultivated under solid substrate fermentation using a combination of rice bran and brown rice as substrate. The highest lovastatin production of 305.08±14.65 ug/g dry solid and 12±0.01 mg glucosamine/g substrate of fungal growth were achieved at the 10th day of cultivation after using all the optimized parameters (5 gram of unground rice bran and brown rice at the ratio of 1:1, water content of 70% (v/w) using sterile distilled water, inoculum size at 1x105 spores/mL and cultivation temperature of room temperature of 30±2 ºC). There was about 90.64% increment in lovastatin production after the optimization of cultural conditions compared with before optimization condition.

Screening antimicrobial activity of various extracts of Urtica dioica

Urtica dioica or stinging nettle is traditionally used as an herbal medicine in Western Asia. The current study represents the investigation of antimicrobial activity of U. dioica from nine crude extracts that were prepared using different organic solvents, obtained from two extraction methods: the Soxhlet extractor (Method I), which included the use of four solvents with ethyl acetate and hexane, or the sequential partitions (Method II) with a five solvent system (butanol). The antibacterial and antifungal activities of crude extracts were tested against 28 bacteria, three yeast strains and seven fungal isolates by the disc diffusion and broth dilution methods. Amoxicillin was used as positive control for bacteria strains, vancomycin for Streptococcus sp., miconazole nitrate (30µg/mL) as positive control for fungi and yeast, and pure methanol (v/v) as negative control. The disc diffusion assay was used to determine the sensitivity of the samples, whilst the broth dilution method was used for the determination of the minimal inhibition concentration (MIC). The ethyl acetate and hexane extract from extraction method I (EA I and HE I) exhibited highest inhibition against some pathogenic bacteria such as Bacillus cereus, MRSA and Vibrio parahaemolyticus. A selection of extracts that showed some activity was further tested for the MIC and minimal bactericidal concentrations (MBC). MIC values of Bacillus subtilis and Methicillin-resistant Staphylococcus aureus (MRSA) using butanol extract of extraction method II (BE II) were 8.33 and 16.33mg/mL, respectively; while the MIC value using ethyl acetate extract of extraction method II (EAE II) for Vibrio parahaemolyticus was 0.13mg/mL. Our study showed that 47.06% of extracts inhibited Gram-negative (8 out of 17), and 63.63% of extracts also inhibited Gram-positive bacteria (7 out of 11); besides, statistically the frequency of antimicrobial activity was 13.45% (35 out of 342) which in this among 21.71% belongs to antimicrobial activity extracts from extraction method I (33 out of 152 of crude extracts) and 6.82% from extraction method II (13 out of 190 of crude extracts). However, crude extracts from method I exhibited better antimicrobial activity against the Gram-positive bacteria than the Gram-negative bacteria. The positive results on medicinal plants screening for antibacterial activity constitutes primary information for further phytochemical and pharmacological studies. Therefore, the extracts could be suitable as antimicrobial agents in pharmaceutical and food industry.

Urtica dioica u ortiga se utiliza tradicionalmente como medicina herbaria en el oeste de Asia. En esta investigación se estudia la actividad antimicrobiana de nueve extractos crudos de U. dioica, los cuales fueron preparados utilizando diferentes disolventes orgánicos y obtenidos a partir de dos métodos de extracción: el extractor Soxhlet (Método I), que incluía el uso de cuatro disolventes con acetato de etilo y hexano, y las particiones secuenciales (Método II) con un sistema de cinco disolventes (butanol). Las actividades antibacterianas y antifúngicas de extractos crudos fueron ensayados contra 28 bacterias, tres cepas de levadura y siete cepas fúngicas por la difusión en disco y el método de dilución en caldo. La amoxicilina se utilizó como control positivo para cepas de bacterias, vancomicina para Streptococcus sp., nitrato de miconazol (30μg/mL) como control positivo para los hongos y levaduras, y el metanol puro (v / v) como control negativo. El ensayo de difusión en disco se utilizó para determinar la sensibilidad de las muestras, mientras que el método de dilución en caldo se utilizó para la determinación de la concentración de inhibición mínima (CIM). El acetato de etilo y el extracto de hexano del método de extracción I (AE I y EH I) mostraron mayor inhibición contra algunas bacterias patógenas tales como Bacillus cereus, MRSA y Vibrio parahaemolyticus. Una selección de extractos que mostraron algún tipo de actividad se probó para el CIM y las concentraciones mínimas bactericidas (CMB). Los valores de CIM de Bacillus subtilis y de Staphylococcus aureus resistentes a la meticilina (MRSA) usando extracto de butanol mediante el método de extracción II (EB II) fueron: 8.33 y 16.33mg/ mL, respectivamente; mientras que el valor de MIC con el uso del extracto de acetato de etilo por el Método de extracción II (EAE II) para Vibrio parahaemolyticus fue 0.13mg/mL. Nuestro estudio mostró que el 47.06% de los extractos inhibieron bacterias Gram-negativas (8 de 17), y el 63,63% de los extractos también inhibieron bacterias Gram-positivas (7 de 11), además que estadísticamente la frecuencia de la actividad antimicrobiana fue de 13.45% (35 de 342), que de este porcentaje un 21.71% pertenece alos extractos de actividad antimicrobiana con el método de extracción I (33 de 152 de los extractos crudos) y un 6.82% del método de extracción II (13 de 190 de los extractos crudos). Sin embargo, los extractos crudos del método I exhibieron una mejor actividad antimicrobiana contra las bacterias Gram-positivas que las Gram-negativas. Los resultados positivos en la detección de plantas medicinales para la actividad antibacteriana constituye información primaria para la realización de nuevos estudios fitoquímicos y farmacológicos. Por lo tanto, los extractos podrían ser adecuados como agentes antimicrobianos en la industria farmacéutica y de alimentos.

Characterization of Solid State Fermentation Culture Conditions for Growth and Mananase Production by Aspergillus niger USM F4 on Rice Husk in Tray System.

Aim: The study evaluated various fermentation conditions for the production of mannanase. Place and Duration of Study: Industrial Biotechnology Research Laboratory (IBRL), School of Biological Sciences, Universiti Sains Malaysia, 11800 Penang, Malaysia between May 2009 and September 2010. Methodology: Solid substrate fermentation was carried out in a shallow aluminum tray system (16 cm x 16 cm x 5 cm) for maximum mannanase production by Aspergillus niger USM F4 using rice husk as a substrate. Results: The maximum mannanase activity of 119.91 U/g substrate was achieved on the 6 days of cultivation when the optimized physical parameters were used (substrate thickness of 1.6 cm or equivalent to 80 g of 0.75 mm rice husk, moisture content to substrate ratio of 1:1 (w/v), cultivation temperature at room temperature (28±2ºC), inoculum size of 6x106 spores/ml and in static condition (no mixing during the fermentation process). The results showed an increment of about 30.79% of mannanase activity after the optimization (119.91 U/g substrate) compared to before optimization (91.68 U/g substrate). Conclusion: The results obtained from this study revealed that rice husk can be used as a substrate for mannanase production in solid state fermentation process.

Pharmacological screening of methanolic extract of Ixora species

<p><b>OBJECTIVE</b>To investigate the antimicrobial activity of methanolic extracts of different parts of Ixora species.</p><p><b>METHODS</b>Antimicrobial activity was carried out using disc diffusion assay against fungi, gram-positive and gram-negative bacteria.</p><p><b>RESULTS</b>All methanolic extracts of different parts of Ixora species showed a broad-spectrum of antibacterial and antiyeast activities, which inhibited the growth of at least one bacterium or yeast. There was no remarkable difference between different Ixora species observed in this study.</p><p><b>CONCLUSIONS</b>The significant antimicrobial activity shown by this Ixora species suggests its potential against infections caused by pathogens. The extract may be developed as an antimicrobial agent.</p>

Gallic acid: An anticandidal compound in hydrolysable tannin extracted from the barks of Rhizophora apiculata Blume.

A research was conducted to study the anticandidal compound of tannin extracted from the barks of a mangrove tree, Rhizophora apiculata Blume. Tannin obtained from the barks of Rhizophora apiculata Blume was further separated into condensed and hydrolysable tannins. A strong anticandidal activity was detected in the hydrolysable tannin, which exhibited minimal inhibitory concentration (MIC) of 6.25 mg/ml, and it was found to have yeastostatic activity at lower concentration (below MIC value) and yeastocidal activity at higher concentration (more than the MIC value). Furthermore, the isolation of the bioactive compound in hydrolysable tannin that responsible for the anticandidal activity was also determined using thin layer chromatography and high-performance liquid chromatography (HPLC). The results obtained confirmed that gallic acid was the bioactive compound that plays role in inhibiting and killing the Candida albicans cells.