This study investigated the reaction kinetics and mechanism of the degradation of 5,5-diphenylhydantoin (DPH) during conventional chlorination and UV/chlorination. DPH is one of the antiepileptic drugs, which has frequently been detected in the aquatic environment. For chlorination, the second-order rate constant for the reaction between DPH and free active chlorine (FAC) was determined at pH 5 to 8. At pH 6 to 8, the efficiency of chlorination in the removal of DPH was found to be dominated by the reaction involving hypochlorous acid (HOCl). The result also showed that anionic species of DPH was more reactive toward FAC as compared with neutral DPH. For UV/chlorination, the effect of FAC dosage and pH on the degradation of DPH was evaluated. UV/chlorination is a more effective method for removing DPH as compared with conventional chlorination and UV irradiation. The DPH degradation rate was found to increase with increasing FAC concentration. On the other hand, the degradation of DPH was found to be more favorable under the acidic condition. Based on the identified transformation by-products, DPH was found to be degraded through the reaction at imidazolidine-2,4-dione moiety of DPH for both chlorination and UV/chlorination. Toxicity study on the chlorination and UV/chlorination-treated DPH solutions suggested that UV/chlorination is a more efficient method for reducing the toxicity of DPH.
This study investigated the reaction kinetics and the transformation by-products of acebutolol during aqueous chlorination. Acebutolol is one of the commonly used β-blockers for the treatment of cardiovascular diseases. It has been frequently detected in the aquatic environment. In the kinetics study, the second-order rate constant for the reaction between acebutolol and chlorine (k app) was determined at 25 ± 0.1 °C. The degradation of acebutolol by free available chlorine was highly pH dependence. When the pH increased from 6 to 8, it was found that the k app for the reaction between acebutolol and free available chlorine was increased from 1.68 to 11.2 M(-1) min(-1). By comparing with the reported k app values, the reactivity of acebutolol toward free available chlorine was found to be higher than atenolol and metoprolol but lower than nadolol and propranolol. Characterization of the transformation by-products formed during the chlorination of acebutolol was carried out using liquid chromatography-quadrupole time-of-flight high-resolution mass spectrometry. Seven major transformation by-products were identified. These transformation by-products were mainly formed through dealkylation, hydroxylation, chlorination, and oxidation reactions.
In this study, the adsorption behavior of azo dye Acid Orange 7 (AO7) from aqueous solution onto macrocomposite (MC) was investigated under various experimental conditions. The adsorbent, MC, which consists of a mixture of zeolite and activated carbon, was found to be effective in removing AO7. The MC were characterized by scanning electron microscopy (SEM), energy dispersive X-ray, point of zero charge, and Brunauer-Emmett-Teller surface area analysis. A series of experiments were performed via batch adsorption technique to examine the effect of the process variables, namely, contact time, initial dye concentration, and solution pH. The dye equilibrium adsorption was investigated, and the equilibrium data were fitted to Langmuir, Freundlich, and Tempkin isotherm models. The Langmuir isotherm model fits the equilibrium data better than the Freundlich isotherm model. For the kinetic study, pseudo-first-order, pseudo-second-order, and intraparticle diffusion model were used to fit the experimental data. The adsorption kinetic was found to be well described by the pseudo-second-order model. Thermodynamic analysis indicated that the adsorption process is a spontaneous and endothermic process. The SEM, Fourier transform infrared spectroscopy, ultraviolet-visible spectral and high performance liquid chromatography analysis were carried out before and after the adsorption process. For the phytotoxicity test, treated AO7 was found to be less toxic. Thus, the study indicated that MC has good potential use as an adsorbent for the removal of azo dye from aqueous solution.
A. niger produced alpha-glucosidase, alpha-amylase and two forms of glucoamylase when grown in a liquid medium containing raw tapioca starch as the carbon source. The glucoamylases, which formed the dominant components of amylolytic activity manifested by the organism, were purified to homogeneity by ammonium sulfate precipitation, ion-exchange and two cycles of gel filtration chromatography. The purified enzymes, designated GA1 and GA2, a raw starch digesting glucoamylase, were found to have molar masses of 74 and 96 kDa and isoelectric points of 3.8 and 3.95, respectively. The enzymes were found to have pH optimum of 4.2 and 4.5 for GA1 and GA2, respectively, and were both stable in a pH range of 3.5-9.0. Both enzymes were thermophilic in nature with temperature optimum of 60 and 65 degrees C, respectively, and were stable for 1 h at temperatures of up to 60 degrees C. The kinetic parameters Km and V showed that with both enzymes the branched substrates, starch and amylopectin, were more efficiently hydrolyzed compared to amylose. GA2, the more active of the two glucoamylases produced, was approximately six to thirteen times more active towards raw starches compared to GA1.
Three novel peptide sequences identified from palm kernel cake (PKC) generated protein hydrolysate including YLLLK, WAFS and GVQEGAGHYALL were used for stability study against angiotensin-converting enzyme (ACE), ACE-inhibition kinetics and molecular docking studies. Results showed that the peptides were degraded at different cleavage degrees of 94%, 67% and 97% for YLLLK, WAFS and GVQEGAGHYALL, respectively, after 3 h of incubation with ACE. YLLLK was found to be the least stable (decreased ACE-inhibitory activity) compared to WAFS and GVQEGAGHYALL (increased ACE-inhibitory activity). YLLLK showed the lowest Ki (1.51 mM) in inhibition kinetics study when compared to WAFS and GVQEGAGHYALL with Ki of 2 mM and 3.18 mM, respectively. In addition, ACE revealed the lowest Kmapp and Vmaxapp and higher catalytic efficiency (CE) in the presence of YLLLK at different concentrations, implying that the enzyme catalysis decreased and hence the inhibition mode increased. Furthermore, YLLLK showed the lowest docking score of -8.224 and seven interactions with tACE, while peptide GVQEGAGHYALL showed the higher docking score of -7.006 and five interactions with tACE.
There has been significant progress in the last few decades in addressing the biomedical applications of polymer hydrogels. Particularly, stimuli responsive hydrogels have been inspected as elegant drug delivery systems capable to deliver at the appropriate site of action within the specific time. The present work describes the synthesis of pH responsive semi-interpenetrating network (semi-IPN) hydrogels of N-succinyl-chitosan (NSC) via Schiff base mechanism using glutaraldehyde as a crosslinking agent and Poly (acrylamide-co-acrylic acid)(Poly (AAm-co-AA)) was embedded within the N-succinyl chitosan network. The physico-chemical interactions were characterized by Fourier transform infrared (FTIR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and field emission scanning electron microscope (FESEM). The synthesized hydrogels constitute porous structure. The swelling ability was analyzed in physiological mediums of pH 7.4 and pH 1.2 at 37°C. Swelling properties of formulations with various amounts of NSC/ Poly (AAm-co-AA) and crosslinking agent at pH 7.4 and pH 1.2 were investigated. Hydrogels showed higher swelling ratios at pH 7.4 while lower at pH 1.2. Swelling kinetics and diffusion parameters were also determined. Drug loading, encapsulation efficiency, and in vitro release of 5-fluorouracil (5-FU) from the synthesized hydrogels were observed. In vitro release profile revealed the significant influence of pH, amount of NSC, Poly (AAm-co-AA), and crosslinking agent on the release of 5-FU. Accordingly, rapid and large release of drug was observed at pH 7.4 than at pH 1.2. The maximum encapsulation efficiency and release of 5-FU from SP2 were found to be 72.45% and 85.99%, respectively. Kinetics of drug release suggested controlled release mechanism of 5-FU is according to trend of non-Fickian. From the above results, it can be concluded that the synthesized hydrogels have capability to adapt their potential exploitation as targeted oral drug delivery carriers.
The synthesis of bacterial polyhydroxyalkanoates (PHA) is very much dependent on the expression and activity of a key enzyme, PHA synthase (PhaC). Many efforts are being pursued to enhance the activity and broaden the substrate specificity of PhaC. Here, we report the identification of a highly active wild-type PhaC belonging to the recently isolated Chromobacterium sp. USM2 (PhaC(Cs)). PhaC(Cs) showed the ability to utilize 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3HV), and 3-hydroxyhexanoate (3HHx) monomers in PHA biosynthesis. An in vitro assay of recombinant PhaC(Cs) expressed in Escherichia coli showed that its polymerization of 3-hydroxybutyryl-coenzyme A activity was nearly 8-fold higher (2,462 ± 80 U/g) than that of the synthase from the model strain C. necator (307 ± 24 U/g). Specific activity using a Strep2-tagged, purified PhaC(Cs) was 238 ± 98 U/mg, almost 5-fold higher than findings of previous studies using purified PhaC from C. necator. Efficient poly(3-hydroxybutyrate) [P(3HB)] accumulation in Escherichia coli expressing PhaC(Cs) of up to 76 ± 2 weight percent was observed within 24 h of cultivation. To date, this is the highest activity reported for a purified PHA synthase. PhaC(Cs) is a naturally occurring, highly active PHA synthase with superior polymerizing ability.
As a remedy for environmental pollution, a versatile synthetic approach has been developed to prepare polyvinyl alcohol (PVA)/nitrogen-doped carbon dots (CDs) composite film (PVA-CDs) for removal of toxic cadmium ions. The CDs were first synthesized using carboxymethylcellulose (CMC) of oil palms empty fruit bunch wastes with the addition of polyethyleneimine (PEI) and then the CDs were embedded with PVA. The PVA-CDs film possess synergistic functionalities through increasing the content of hydrogen bonds for chemisorption compared to the pure CDs. Optical analysis of PVA-CDs film was performed by ultraviolet-visible and fluorescence spectroscopy. Compared to the pure CDs, the solid-state PVA-CDs displayed a bright blue color with a quantum yield (QY) of 47%; they possess excitation-independent emission and a higher Cd2+ removal efficiency of 91.1%. The equilibrium state was achieved within 10 min. It was found that adsorption data fit well with the pseudo-second-order kinetic and Langmuir isotherm models. The maximum adsorption uptake was 113.6 mg g-1 at an optimal pH of 7. Desorption experiments showhe that adsorbent can be reused fruitfully for five adsorption-desorption cycles using 0.1 HCl elution. The film was successfully applied to real water samples with a removal efficiency of 95.34% and 90.9% for tap and drinking water, respectively. The fabricated membrane is biodegradable and its preparation follows an ecofriendly green route.
Previous molecular dynamic simulations have reported elongation of the existing beta-sheet in prion proteins. Detailed examination has shown that these elongations do not extend beyond the proline residues flanking these beta-sheets. In addition, proline has also been suggested to possess a possible structural role in preserving protein interaction sites by preventing invasion of neighboring secondary structures. In this work, we have studied the possible structural role of the flanking proline residues by simulating mutant structures with alternate substitution of the proline residues with valine. Simulations showed a directional inhibition of elongation, with the elongation progressing in the direction of valine including evident inhibition of elongation by existing proline residues. This suggests that the flanking proline residues in prion proteins may have a containment role and would confine the beta-sheet within a specific length.
The main objective of the present study is the physicochemical characterization of naturally available Terminalia catappa gum (Badam gum [BG]) as a novel pharmaceutical excipient and its suitability in the development of gastroretentive floating drug delivery systems (GRFDDS) to retard the drug for 12 h when the dosage form is exposed to gastrointestinal fluids in the gastric environment. As BG was being explored for the first time for its pharmaceutical application, physicochemical, microbiological, rheological, and stability studies were carried out on this gum. In the present investigation, the physicochemical properties, such as micromeritic, rheological, melting point, moisture content, pH, swelling index, water absorption, and volatile acidity, were evaluated. The gum was characterized by scanning electron microscopy, differential scanning calorimetry (DSC), powder X-ray diffraction studies (PXRD), and Fourier transform infrared spectroscopy (FTIR). Gastroretentive floating tablets of BG were prepared with the model drug propranolol HCl by direct compression methods. The prepared tablets were evaluated for all their physicochemical properties, in vitro buoyancy, in vitro drug release, and rate order kinetics. PBG 04 was selected as an optimized formulation based on its 12-h drug release and good buoyancy characteristics. The optimized formulation was characterized with FTIR, DSC, and PXRD studies, and no interaction between the drug and BG was found. Thus, the study confirmed that BG might be used in the gastroretentive drug delivery system as a release-retarding polymer.
Hydroxylated polymethoxyflavones (HPMFs) have been shown to possess various anti-disease effects, including against obesity. This study investigates the anti-obesity effects of HPMFs in further detail, aiming to gain understanding of their mechanism of action in this context. The current study demonstrates that two HPMFs; 3'-hydroxy-5,7,4',5'-tetramethoxyflavone (3'OH-TetMF) and 4'-hydroxy-5,7,3',5'-tetramethoxyflavone (4'OH-TetMF) possess anti-obesity effects. They both significantly reduced pancreatic lipase activity in a competitive manner as demonstrated by molecular docking and kinetic studies. In cell studies, it was revealed that both of the HPMFs suppress differentiation of 3T3-L1 mouse embryonic fibroblast cells during the early stages of adipogenesis. They also reduced expression of key adipogenic and lipogenic marker genes, namely peroxisome proliferator-activated receptor-gamma (PPARγ), CCAAT/enhancer-binding protein α and β (C/EBP α and β), adipocyte binding protein 2 (aP2), fatty acid synthase (FASN), and sterol regulatory element-binding protein 1 (SREBF 1). They also enhanced the expression of cell cycle genes, i.e., cyclin D1 (CCND1) and C-Myc, and reduced cyclin A2 expression. When further investigated, it was also observed that these HPMFs accelerate lipid breakdown (lipolysis) and enhance lipolytic genes expression. Moreover, they also reduced the secretion of proteins (adipokines), including pro-inflammatory cytokines, from mature adipocytes. Taken together, this study concludes that these HPMFs have anti-obesity effects, which are worthy of further investigation.
Extraction of protease from a local ginger rhizome (Zingiber officinale var. Bentong) was carried out. The effect of extraction pH (6.4, 6.8, 7.0, 7.2, 7.6, 8.0, 8.4, and 8.8) and stabilizers (0.2% ascorbic acid, 0.2% ascorbic acid and 5 mM EDTA, or 10 mM cysteine and 5 mM EDTA) on protease activity during extraction was examined. pH 7.0 potassium phosphate buffer and 10 mM cysteine in combination with 5 mM EDTA as stabilizer were found to be the most effective conditions. The extraction procedure yielded 0.73% of Bentong ginger protease (BGP) with a specific activity of 24.8±0.2 U/mg protein. Inhibitory tests with some protease inhibitors classified the enzyme as a cysteine protease. The protease showed optimum activity at 60 °C and pH 6-8, respectively. The enzyme was completely inhibited by heavy metal cations such as Cu2+, and Hg2+. SDS stimulated the activity of enzyme, while emulsifiers (Tween 80 and Tween 20) slightly reduced its activity. The kinetic analysis showed that the protease has Km and Vmax values of 0.21 mg mL-1 and 34.48 mg mL-1 min-1, respectively. The dried enzyme retained its activity for 22 months when stored at -20 °C.
Production cost of enzyme is largely determined by the type of the strain and raw material used to propagate the strain. Hence, selection of the strain and raw materials is crucial in enzyme production. For Glucose oxidase (GOx), previous studies showed Aspergillus terreus UniMAP AA-1 offers a better alternative to the existing sources. Thus, a lower production cost could be logically anticipated by growing the strain in a cheaper complex media such as molasses. In this work, sugar cane molasses, supplemented with urea and carbonate salt and a locally isolated strain Aspergillus terreus UniMAP AA-1 were used to produce a crude GOx enzyme in a small scale. A statistical optimization approach namely Response Surface Methodology (RSM) was used to optimize the media components for highest GOx activity. It was found that the highest GOx activity was achieved using a combination of molasses, carbonate salt and urea at concentration 32.51, 4.58 and 0.93% (w/v), respectively. This study provides an alternative optimized media conditions for GOx production using locally available raw materials.
1. The effect of flavonoids on coumarin 7-hydroxylation, an activity marker of an important human liver cytochrome P450 isoform, cytochrome P450 2A6 (CYP2A6), was investigated in this study. 2. Coumarin 7-hydroxylase activity was measured fluorometrically in reaction mixtures containing cDNA-expressed CYP2A6, nicotinamide adenine dinucleotide phosphate generating system and 10 uM coumarin, at various concentrations of flavonoids. 3. Among the 23 compounds tested, most of the active members were from flavonol group of hydroxylated flavonoids, with myricetin being the most potent inhibitor followed by quercetin, galangin, and kaempferol. 4. Further exploration of the inhibition mechanism of these compounds revealed that myricetin, galangin, and kaempferol exhibited mixed-type of inhibition pattern while quercetin was observed to exhibit competitive mode of inhibition. 5. Structure-function analyses revealed that degree of inhibition was closely related to the number and location of hydroxyl groups, glycosylation of the free hydroxyl groups, degree of saturation of the flavane nucleus as well as the presence of the alkoxylated function.
This study focuses on a novel lipase from Bacillus licheniformis IBRL-CHS2. The lipase gene was cloned into the pGEM-T Easy vector, and its sequences were registered in GenBank (KU984433 and AOT80658). It was identified as a member of the bacterial lipase subfamily 1.4. The pCold I vector and E. coli BL21 (DE3) host were utilized for expression, with the best results obtained by removing the enzyme's signal peptide. Optimal conditions were found to be 15°C for 24 h, using 0.2 mM Isopropyl β-D-1-thiogalactopyranoside (IPTG). The His-tagged lipase was purified 13-fold with a 68% recovery and a specific activity of 331.3 U/mg using affinity purification. The lipase demonstrated optimal activity at 35°C and pH 7. It remained stable after 24 h in 25% (v/v) organic solvents such as isooctane, n-hexane, dimethyl sulfoxide (DMSO), and methanol, which enhanced its activity. Chloroform and diethyl ether inhibited the lipase. The enzyme exhibited the highest affinity for p-nitrophenol laurate (C12:0) with a Km of 0.36 mM and a Vmax of 357 μmol min-1 mg-1. Among natural oils, it performed best with coconut oil and worst with olive oil. The lipase was stable in the presence of 1 mM and 5 mM Ca2⁺, K⁺, Na⁺, Mg2⁺, and Ba2⁺, but its activity decreased with Zn2⁺ and Al3⁺. Non-ionic surfactants like Triton X-100, Nonidet P40, Tween 20, and Tween 40 boosted activity, while Sodium Dodecyl Sulfate (SDS) inhibited it. This lipase's unique properties, particularly its stability in organic solvents, make it suitable for applications in organic synthesis and various industries.
One pot, three-component reaction of 1-acryloyl-3,5-bisarylmethylidenepiperidin-4-ones with isatin and sarcosine in molar ratios of 1:1:1 and 1:2:2 furnished to mono- and bis-spiropyrrolidine heterocyclic hybrids comprising functionalized piperidine, pyrrolidine and oxindole structural motifs. Both mono and bis-spiropyrrolidines displayed good inhibitory activity against acetylcholinesterase (AChE) with IC₅₀ values of 2.36-9.43 μM. For butyrylcholinesterase (BChE), mono-cycloadducts in series 8 with IC₅₀ values of lower than 10 μM displayed better inhibitory activities than their bis-cycloadduct analogs in series 9 with IC₅₀ values of 7.44-19.12 μM. The cycloadducts 9j and 8e were found to be the most potent AChE and BChE inhibitors with IC₅₀ values of 2.35 and 3.21 μM, respectively. Compound 9j was found to be competitive inhibitor of AChE while compound 8e was a mixed-mode inhibitor of BChE with calculated Ki values of 2.01 and 6.76 μM, respectively. Molecular docking on Torpedo californica AChE and human BChE showed good correlation between IC₅₀ values and free binding energy values of the synthesized compounds docked into the active site of the enzymes.
In the study reported here, magnesium/aluminum (Mg/Al)-layered double hydroxide (LDH) was intercalated with an anticancer drug, protocatechuic acid, using ion-exchange and direct coprecipitation methods, with the resultant products labeled according to the method used to produce them: "PANE" (ie, protocatechuic acid-Mg/Al nanocomposite synthesized using the ion-exchange method) and "PAND" (ie, protocatechuic acid-Mg/Al nanocomposite synthesized using the direct method), respectively. Powder X-ray diffraction and Fourier transform infrared spectroscopy confirmed the intercalation of protocatechuic acid into the inter-galleries of Mg/Al-LDH. The protocatechuic acid between the interlayers of PANE and PAND was found to be a monolayer, with an angle from the z-axis of 8° for PANE and 15° for PAND. Thermogravimetric and differential thermogravimetric analysis results revealed that the thermal stability of protocatechuic acid was markedly enhanced upon intercalation. The loading of protocatechuic acid in PANE and PAND was estimated to be about 24.5% and 27.5% (w/w), respectively. The in vitro release study of protocatechuic acid from PANE and PAND in phosphate-buffered saline at pH 7.4, 5.3, and 4.8 revealed that the nanocomposites had a sustained release property. After 72 hours incubation of PANE and PAND with MCF-7 human breast cancer and HeLa human cervical cancer cell lines, it was found that the nanocomposites had suppressed the growth of these cancer cells, with a half maximal inhibitory concentration of 35.6 μg/mL for PANE and 36.0 μg/mL for PAND for MCF-7 cells, and 19.8 μg/mL for PANE and 30.3 μg/mL for PAND for HeLa cells. No half maximal inhibitory concentration for either nanocomposite was found for 3T3 cells.
Cu(II) removal efficacies of alginate-immobilized Trichoderma asperellum using viable and non-viable forms were investigated with respect to time, pH, and initial Cu(II) concentrations. The reusability potential of the biomass was determined based on sorption/desorption tests. Cu(II) biosorption by immobilized heat-inactivated T. asperellum cells was the most efficient, with 134.22mg Cu(II) removed g(-1) adsorbent, compared to immobilized viable cells and plain alginate beads (control) with 105.96 and 94.04mg Cu(II) adsorbed g(-1) adsorbent, respectively. Immobilized non-viable cells achieved equilibrium more rapidly within 4h. For all biosorbents, optimum pH for Cu(II) removal was between pH 4 and 5. Reusability of all biosorbents were similar, with more than 90% Cu(II) desorbed with HCl. These alginate-immobilized cells can be applied to reduce clogging and post-separation process incurred from use of suspended biomass.
The feasibility of preparing activated carbon (JPAC) from jackfruit peel, an industrial residue abundantly available from food manufacturing plants via microwave-assisted NaOH activation was explored. The influences of chemical impregnation ratio, microwave power and radiation time on the properties of activated carbon were investigated. JPAC was examined by pore structural analysis, scanning electron microscopy, Fourier transform infrared spectroscopy, nitrogen adsorption isotherm, elemental analysis, surface acidity/basicity and zeta potential measurements. The adsorptive behavior of JPAC was quantified using methylene blue as model dye compound. The best conditions resulted in JPAC with a monolayer adsorption capacity of 400.06 mg/g and carbon yield of 80.82%. The adsorption data was best fitted to the pseudo-second-order equation, while the adsorption mechanism was well described by the intraparticle diffusion model. The findings revealed the versatility of jackfruit peels as good precursor for preparation of high quality activated carbon.
The feasibility of langsat empty fruit bunch waste for preparation of activated carbon (EFBLAC) by microwave-induced activation was explored. Activation with NaOH at the IR ratio of 1.25, microwave power of 600 W for 6 min produced EFBLAC with a carbon yield of 81.31% and adsorption uptake for MB of 302.48 mg/g. Pore structural analysis, scanning electron microscopy and Fourier transform infrared spectroscopy demonstrated the physical and chemical characteristics of EFBLAC. Equilibrium data were best described by the Langmuir isotherm, with a monolayer adsorption capacity of 402.06 mg/g, and the adsorption kinetics was well fitted to the pseudo-second-order equation. The findings revealed the potential to prepare high quality activated carbon from langsat empty fruit bunch waste by microwave irradiation.