Polymeric nanoparticles gain a widespread interest in food and pharmaceutical industries as delivery systems that encapsulate, protect, and release lipophilic compounds such as omega-3 fatty acids, fat-soluble vitamins, carotenoids, carvedilol, cyclosporine, and ketoprofen. In this study, medium-chain-length poly-3-hydroxyalkanoate (mcl-PHA)-incorporated nanoparticle was developed via facile organic solvent-free nanoemulsion templating technique. The water content (W/surfactant-to-oil (S/O)), S/O, and Cremophor EL-to-Span 80 (Cremo/Sp80) ratios were first optimized using response surface methodology (RSM) to obtain nanoemulsion template prior to incorporation of mcl-PHA. Their effects on nanoemulsion formation were investigated. The mcl-PHA-incorporated nanoparticle system showed a good preservation capability of β-carotene and extended storage stability.
C50 carotenoids, as unique bioactive molecules, have many biological properties, including antioxidant, anticancer, and antibacterial activity, and have a wide range of potential uses in the food, cosmetic, and biomedical industries. The majority of C50 carotenoids are produced by the sterile fermentation of halophilic archaea. This study aims to look at more cost-effective and manageable ways of producing C50 carotenoids. The basic medium, carbon source supplementation, and optimal culture conditions for Halorubrum sp. HRM-150 C50 carotenoids production by open fermentation were examined in this work. The results indicated that Halorubrum sp. HRM-150 grown in natural brine medium grew faster than artificial brine medium. The addition of glucose, sucrose, and lactose (10 g/L) enhanced both biomass and carotenoids productivity, with the highest level reaching 4.53 ± 0.32 μg/mL when glucose was added. According to the findings of orthogonal studies based on the OD600 and carotenoids productivity, the best conditions for open fermentation were salinity 20-25%, rotation speed 150-200 rpm, and pH 7.0-8.2. The up-scaled open fermentation was carried out in a 7 L medium under optimum culture conditions. At 96 h, the OD600 and carotenoids productivity were 9.86 ± 0.51 (dry weight 10.40 ± 1.27 g/L) and 7.31 ± 0.65 μg/mL (701.40 ± 21.51 μg/g dry weight, respectively). When amplified with both universal bacterial primer and archaeal primer in the open fermentation, Halorubrum remained the dominating species, indicating that contamination was kept within an acceptable level. To summarize, open fermentation of Halorubrum is a promising method for producing C50 carotenoids.
This research work seeks to evaluate the impact of selected enzyme complexes on the optimised release of phenolics from leaves of Pongamia pinnata. After preliminary solvent extraction, the P. pinnata leaf extract was subjected to enzymatic treatment, using enzyme cocktails such as kemzyme dry-plus, natuzyme, and zympex-014. It was noticed that zympex-014 had a greater extract yield (28.0%) than kemzyme dry-plus (17.0%) and natuzyme (18.0%). Based on the better outcomes, zympex-014-based extract values were subsequently applied to several RSM parameters. The selected model is suggested to be significant by the F value (12.50) and R2 value (0.9669). The applicability of the ANN model was shown by how closely the projected values from the ANN were to the experimental values. In terms of total phenolic contents (18.61 mg GAE/g), total flavonoid contents (12.56 mg CE/g), and DPPH test (IC50) (6.5 g/mL), antioxidant activities also shown significant findings. SEM analysis also revealed that the cell walls were damaged during enzymatic hydrolysis, as opposed to non-hydrolysed material. Using GC-MS, five potent phenolic compounds were identified in P. pinnata extract. According to the findings of this study, the recovery of phenolic bioactives and subsequent increase in the antioxidant capacity of P. pinnata leaf extract were both positively impacted by the optimisation approaches suggested, including the use of zympex-014.
Acetone-butanol-ethanol (ABE) production from renewable resources has been widely reported. In this study, Clostridium butyricum EB6 was employed for ABE fermentation using fermentable sugar derived from treated oil palm empty fruit bunch (OPEFB). A higher amount of ABE (2.61 g/l) was produced in a fermentation using treated OPEFB as the substrate when compared to a glucose based medium that produced 0.24 g/l at pH 5.5. ABE production was increased to 3.47 g/l with a yield of 0.24 g/g at pH 6.0. The fermentation using limited nitrogen concentration of 3 g/l improved the ABE yield by 64%. The study showed that OPEFB has the potential to be applied for renewable ABE production by C. butyricum EB6.
Almost 70% of clinically used antineoplastic drugs are originated from natural products such as plants, marine organism, and microorganisms and some of them are also structurally modified natural products. The naturally occurring drugs may specifically act as inducers of selective cytotoxicity, anti-metastatic, anti-mutagenic, anti-angiogenesis, antioxidant accelerators, apoptosis inducers, autophagy inducers, and cell cycle inhibitors in cancer therapy. Precisely, several reports have demonstrated the involvement of naturally occurring anti-breast cancer drugs in regulating the expression of oncogenic and tumor suppressors associated with carcinogen metabolism and signaling pathways. Anticancer therapies based on nanotechnology have the potential to improve patient outcomes through targeted therapy, improved drug delivery, and combination therapies. This paper has reviewed the current treatment for breast cancer and the potential disadvantages of those therapies, besides the various mechanism used by naturally occurring phytochemicals to induce apoptosis in different types of breast cancer. Along with this, the contribution of nanotechnology in improving the effectiveness of anticancer drugs was also reviewed. With the development of sciences and technologies, phytochemicals derived from natural products are continuously discovered; however, the search for novel natural products as chemoprevention drugs is still ongoing, especially for the advanced stage of breast cancer. Continued research and development in this field hold great promise for advancing cancer care and improving patient outcomes.
Viral diseases are causing mayhem throughout the world. One of the zoonotic viruses that have emerged as a potent threat to community health in the past few decades is Nipah virus. Nipah viral sickness is a zoonotic disease whose main carrier is bat. This disease is caused by Nipah virus (NiV). It belongs to the henipavirous group and of the family paramyxoviridae. Predominantly Pteropus spp. is the carrier of this virus. It was first reported from the Kampung Sungai Nipah town of Malaysia in 1998. Human-to-human transmission can also occur. Several repeated outbreaks were reported from South and Southeast Asia in the recent past. In humans, the disease is responsible for rapid development of acute illness, which can result in severe respiratory illness and serious encephalitis. Therefore, this calls for an urgent need for health authorities to conduct clinical trials to establish possible treatment regimens to prevent any further outbreaks.
Recent studies on biocatalysis in water-organic solvent biphasic systems have shown that many enzymes retain their catalytic activities in the presence of high concentrations of organic solvents. However, not all enzymes are organic solvent tolerant, and most have limited and selective tolerance to particular organic solvents. Protein modification or protein tailoring is an approach to alter the characteristics of enzymes, including solubility in organic solvents. Particular amino acids may play pivotal roles in the catalytic ability of the protein. Attaching soluble modifiers to the protein molecule may alter its conformation and the overall polarity of the molecule. Enzymes, in particular lipases, have been chemically modified by attachment of aldehydes, polyethylene glycols, and imidoesters. These modifications alter the hydrophobicity and conformation of the enzymes, resulting in changes in the microenvironment of the enzymes. By these modifications, newly acquired properties such as enhancement of activity and stability and changes in specificity and solubility in organic solvents are obtained. Modified lipases were found to be more active and stable in organic solvents. The optimum water activity (a(w)) for reaction was also shifted by using modified enzymes. Changes in enantioselective behavior were also observed.
Keratinous wastes have increasingly become a problem and accumulate in the environment mainly in the form of feathers, generated mainly from a large number of poultry industries. As keratins are very difficult to degrade by general proteases, they pose a major environmental problem. Therefore, microorganisms which would effectively degrade keratins are needed for recycling such wastes. A geophilic dermatophyte, Microsporum fulvum IBRL SD3 which was isolated from a soil sample collected from a chicken feather dumping site using a baiting technique, was capable to produce keratinase significantly. The crude keratinase was able to degrade whole chicken feathers effectively. The end product of the degradation was protein that contained essential amino acids and may have potential application in animal feed production. Thus, M. fulvum could be a novel organism to produce keratinase for chicken feathers degradation.
Diabetes mellitus (DM) is considered as a complex metabolic disease because it affects the metabolism of glucose and other metabolites. Although many diabetes studies have been conducted in animal models throughout the years, the pathogenesis of this disease, especially between lean diabetes (ND + STZ) and obese diabetes (OB + STZ), is still not fully understood. In this study, the urine from ND + STZ, OB + STZ, lean/control (ND), and OB + STZ rats were collected and compared by using (1)H NMR metabolomics. The results from multivariate data analysis (MVDA) showed that the diabetic groups (ND + STZ and OB + STZ) have similarities and dissimilarities for a certain level of metabolites. Differences between ND + STZ and OB + STZ were particularly noticeable in the synthesis of ketone bodies, branched-chain amino acid (BCAA), and sensitivity towards the oral T2DM diabetes drug metformin. This finding suggests that the ND + STZ group was more similar to the T1DM model and OB + STZ to the T2DM model. In addition, we also managed to identify several pathways and metabolism aspects shared by obese (OB) and OB + STZ. The results from this study are useful in developing drug target-based research as they can increase understanding regarding the cause and effect of DM.
Evidence pointed towards the benefits of Marantodes pumilum in treating osteoporosis after menopause; however, the detailed mechanisms still have not been explored. Therefore, this study aims to identify the molecular mechanisms underlying M. pumilum's bone-protective effect via the involvement of RANK/RANKL/OPG and Wnt/β-catenin signaling pathways. Ovariectomized adult female rats were given M. pumilum leaf aqueous extract (MPLA) (50 and 100 mg/kg/day) and estrogen (positive control) orally for twenty-eight consecutive days. Following the treatment, rats were sacrificed, and femur bones were harvested. Blood was withdrawn for analysis of serum Ca2+, PO43-, and bone alkaline phosphatase (BALP) levels. The bone microarchitectural changes were observed by H&E and PAS staining and distribution and expression of RANK/RANKL/OPG and Wnt3a/β-catenin and its downstream proteins were determined by immunohistochemistry, immunofluorescence, Western blot, and real-time PCR. MPLA treatment increased serum Ca2+ and PO43- levels and reduced serum BALP levels (p
In silico and experimental investigations were conducted to explore the effects of substituting hydrophobic residues, Val, Met, Leu, Ile, Trp, and Phe into Gln 114 of T1 lipase. The in silico investigations accurately predicted the enzymatic characteristics of the mutants in the experimental studies and provided rationalization for some of the experimental observations. Substitution with Leu successfully improved the conformational stability and enzymatic characteristics of T1 lipase. However, replacement of Gln114 with Trp negatively affected T1 lipase and resulted in the largest disruption of protein stability, diminished lipase activity and inferior enzymatic characteristics. These results suggested that the substitution of a larger residue in a densely packed area of the protein core can have considerable effects on the structure and function of an enzyme. This is especially true when the residue is next to the catalytic serine as demonstrated with the Phe and Trp mutation.
COVID-19 is a disease that puts most of the world on lockdown and the search for therapeutic drugs is still ongoing. Therefore, this study used in silico screening to identify natural bioactive compounds from fruits, herbaceous plants, and marine invertebrates that are able to inhibit protease activity in SARS-CoV-2 (PDB: 6LU7). We have used extensive screening strategies such as drug likeliness, antiviral activity value prediction, molecular docking, ADME, molecular dynamics (MD) simulation, and MM/GBSA. A total of 17 compounds were shortlisted using Lipinski's rule in which 5 compounds showed significant predicted antiviral activity values. Among these 5, only 2 compounds, Macrolactin A and Stachyflin, showed good binding energy of -9.22 and -8.00 kcal/mol, respectively, within the binding pocket of the Mpro catalytic residues (HIS 41 and CYS 145). These two compounds were further analyzed to determine their ADME properties. The ADME evaluation of these 2 compounds suggested that they could be effective in developing therapeutic drugs to be used in clinical trials. MD simulations showed that protein-ligand complexes of Macrolactin A and Stachyflin with the target receptor (6LU7) were stable for 100 nanoseconds. The MM/GBSA calculations of Mpro-Macrolactin A complex indicated higher binding free energy (-42.58 ± 6.35 kcal/mol). Dynamic cross-correlation matrix (DCCM) and principal component analysis (PCA) on the residual movement in the MD trajectories further confirmed the stability of Macrolactin A bound state with 6LU7. In conclusion, this study showed that marine natural compound Macrolactin A could be an effective therapeutic inhibitor against SARS-CoV-2 protease (6LU7). Additional in vitro and in vivo validations are strongly needed to determine the efficacy and therapeutic dose of Macrolactin A in biological systems.
Lipase from Candida rugosa was immobilized by entrapment on poly(N-vinyl- 2-pyrrolidone-co-2-hydroxyethyl methacrylate) (poly[VP-co-HEMA]) hydrogel, and divinylbenzene was the crosslinking agent. The immobilized enzymes were used in the esterification reaction of oleic acid and butanol in hexane. The activities of the immobilized enzymes and the leaching ability of the enzyme from the support with respect to the different compositions of the hydrogels were investigated. The thermal, solvent, and storage stability of the immobilized lipases was also determined. Increasing the percentage of composition of VP from 0 to 90, which corresponds to the increase in the hydrophilicity of the hydrogels, increased the activity of the immobilized enzyme. Lipase immobilized on VP(%):HEMA(%) 90:10 exhibited the highest activity. Lipase immobilized on VP(%):HEMA(%) 50:50 showed the highest thermal, solvent, storage, and operational stability compared to lipase immobilized on other compositions of hydrogels as well as the native lipase.
The majority of pathogenic microorganisms have developed resistance to commercial antibiotics. It causes the risk of illness relapse with current antimicrobial therapy regimens; additional and/or different antibacterial drugs are needed to treat diseases caused by these pathogenic microorganisms. The applied analysis in the present study was purification and characterization of plant peptides isolated from the leaves of Fagonia bruguieri as well as their antibacterial activities against Gram-positive bacteria, Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus, and methicillin-resistant Staphylococcus aureus, in addition to Gram-negative bacteria, Escherichia coli and Pseudomonas aeruginosa. The minimum inhibitory concentration for the isolated peptide ranges from 25 to 62.5 mg/mL. The methanolic solvent was used for the extraction followed by reversed-phase high-performance liquid chromatography for purification of peptides. Eventually, the peptide characterization and identification were also determined by MALDI-TOF/TOF and SEM analysis. This study paves a way to the effective antimicrobials from the plant resources.
Several filamentous fungi are able to concomitantly assimilate both aliphatic and polycyclic aromatic hydrocarbons that are the biogenic by-products of some industrial processes. Cytochrome P450 monooxygenases catalyze the first oxidation reaction for both types of substrate. Among the cytochrome P450 (CYP) genes, the family CYP52 is implicated in the first hydroxylation step in alkane-assimilation processes, while genes belonging to the family CYP53 have been linked with oxidation of aromatic hydrocarbons. Here, we perform a comparative analysis of CYP genes belonging to clans CYP52 and CYP53 in Aspergillus niger, Beauveria bassiana, Metarhizium robertsii (formerly M. anisopliae var. anisopliae), and Penicillium chrysogenum. These species were able to assimilate n-hexadecane, n-octacosane, and phenanthrene, exhibiting a species-dependent modification in pH of the nutrient medium during this process. Modeling of the molecular docking of the hydrocarbons to the cytochrome P450 active site revealed that both phenanthrene and n-octacosane are energetically favored as substrates for the enzymes codified by genes belonging to both CYP52 and CYP53 clans, and thus appear to be involved in this oxidation step. Analyses of gene expression revealed that CYP53 members were significantly induced by phenanthrene in all species studied, but only CYP52X1 and CYP53A11 from B. bassiana were highly induced with n-alkanes. These findings suggest that the set of P450 enzymes involved in hydrocarbon assimilation by fungi is dependent on phylogeny and reveal distinct substrate and expression specificities.
Agitation speed was found to influence the tannase production and fungal growth of Aspergillus niger FETL FT3. The optimal agitation speed was at 200 rpm which produced 1.41 U/ml tannase and 3.75 g/l of fungal growth. Lower or higher agitation speeds than 200 rpm produced lower enzyme production and fungal growth. Based on the SEM and TEM micrograph observation, there was a significant correlation between agitation speed and the morphology of the fungal mycelia. The results revealed an increase of the enzyme production with the change of the fungal growth morphology from filamentous to pelleted growth forms. However, the exposure to higher shear stress with an increasing agitation speed of the shaker also resulted in lower biomass yields as well as enzyme production.
Copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] has been the center of attention in the bio-industrial fields, as it possesses superior mechanical properties compared to poly(3-hydroxybutyrate) [P(3HB)]. The usage of oleic acid and 1-pentanol was exploited as the carbon source for the production of P(3HB-co-3HV) copolymer by using a locally isolated strain Cupriavidus sp. USMAA2-4. In this study, the productivity of polyhydroxyalkanoate (PHA) was improved by varying the frequency of feeding in fed-batch culture. The highest productivity (0.48 g/L/h) that represents 200 % increment was obtained by feeding the carbon source and nitrogen source three times and also by considering the oxygen uptake rate (OUR) and oxygen transfer rate (OTR). A significantly higher P(3HB-co-3HV) concentration of 25.7 g/L and PHA content of 66 wt% were obtained. The 3-hydroxyvalerate (3HV) monomer composition obtained was 24 mol% with the growth of 13.3 g/L. The different frequency of feeding carried out has produced a blend copolymer and has broadened the monomer distribution. In addition, increase in number of granules was also observed as the frequency of feeding increases. In general, the most glaring increment in productivity offer advantage for industrial P(3HB-co-3HV) production, and it is crucial in developing cost-effective processes for commercialization.
B cells and in particular antibodies has always played second fiddle to cellular immunity in regard to tuberculosis (TB). However, recent studies has helped position humoral immunity especially antibodies back into the foray in relation to TB immunity. Therefore, the ability to correlate the natural antibody responses of infected individuals toward TB antigens would help strengthen this concept. Phage display is an intriguing approach that can be utilized to study antibody-mediated responses against a particular infection via harvesting the B cell repertoire from infected individuals. The development of disease-specific antibody libraries or immune libraries is useful to better understand antibody-mediated immune responses against specific disease antigens. This study describes the generation of an immune single-chain variable fragment (scFv) library derived from TB-infected individuals. The immune library with an estimated diversity of 109 independent clones was then applied for the identification of monoclonal antibodies against Mycobacterium tuberculosis α-crystalline as a model antigen. Biopanning of the library isolated three monoclonal antibodies with unique gene usage. This strengthens the role of antibodies in TB immunity in addition to the role played by cellular immunity. The developed library can be applied against other TB antigens and aid antibody-derived TB immunity studies in the future.
Synthesis of layered double hydroxides (LDHs) of Zn/Al-NO3- hydrotalcite (HIZAN) and Zn/Al-diocytyl sodium sulfosuccinate (DSS) nanocomposite (NAZAD) with a molar ratio of Zn/Al of 4:1 were carried out by coprecipitation through continuous agitation. Their structures were determined using X-ray diffractometer spectra, which showed that basal spacing for LDH synthesized by both methods was about 8.89 A. An expansion of layered structure of about 27.9 A was observed to accommodate the surfactant anion between the interlayer. This phenomenon showed that the intercalation process took place between the LDH interlayer. Lipase from Candida rugosa was immobilized onto these materials by physical adsorption method. It was found that the protein loading onto NAZAD is higher than HIZAN. The activity of immobilized lipase was investigated through esterification of oleic acid and 1-butanol in hexane. The effects of pore size, surface area, reaction temperature, thermostability of the immobilized lipases, storage stability in organic solvent, and leaching studies were investigated. Stability was found to be the highest in the nanocomposite NAZAD.
Microbial pigments are gaining intensive attention due to increasing awareness of the toxicity of synthetic colours. In this study, a novel polymer-producing bacterium designated as Cupriavidus sp. USMAHM13 was also found to produce yellow pigment when cultivated in nutrient broth. Various parameters such as temperature, pH and ratio of culture volume to flask volume were found to influence the yellow pigment production. UV-Visible, Fourier transform infrared and (13)C-nuclear magnetic resonance analyses revealed that the crude yellow pigment might probably represent new bioactive compound in the carotenoid family. The crude yellow pigment also exhibited a wide spectrum of antimicrobial activity against Gram-negative and Gram-positive bacteria with their inhibition zones and minimal inhibitory concentrations ranged from 25 to 38 mm and from 0.63 to 2.5 mg/ml, respectively. To the best of our knowledge, this is the first report on the identification and characterization of yellow pigment produced by bacterium belonging to the genus Cupriavidus.