The performance of gas chromatography (GC) combined with a hybrid quadrupole time-of-flight (QTOF) mass spectrometry (MS) system for the determination of volatile and semi-volatile compounds in honey samples is evaluated. After headspace (HS) solid-phase microextraction (SPME) of samples, the accurate mass capabilities of the above system were evaluated for compounds identification. Accurate scan electron impact (EI) MS spectra allowed discriminating compounds displaying the same nominal masses, but having different empirical formulae. Moreover, the use of a mass window with a width of 0.005 Da provided highly specific chromatograms for selected ions, avoiding the contribution of interferences to their peak areas. Additional information derived from positive chemical ionization (PCI) MS spectra and ion product scan MS/MS spectra permitted confirming the identity of novel compounds. The above possibilities are illustrated with examples of honey aroma compounds, belonging to different chemical classes and containing different elements in their molecules. Examples of compounds whose structures could not be described are also provided. Overall, 84 compounds, from a total of 89 species, could be identified in 19 honey samples from 3 different geographic areas in the world. The suitability of responses measured for selected ions, corresponding to above species, for authentication purposes is assessed through principal components analysis.
In this present study, isolation, characterization and protective effect of sulfated polysaccharide (SP) isolated from the brown algae Padina gymnospora was investigated. SP was isolated and characterized through FT-IR, 1H NMR, TGA, GC-MS and CHN analysis. The molecular weight of SP was found to be 16 kDa. The isolated SP contains 29.4 ± 0.35% of sulfate, 27 ± 0.11% of fucose, 0.05 ± 0.12% of protein, respectively. Furthermore, SP exhibits its excellent radical scavenging effects were evaluated by DPPH, ABTS radical scavenging and reducing power assays. Moreover, pretreatment with SP significantly mitigates H2O2 induced cytotoxicity in L-929 cells in a dose dependent manner. Furthermore, SP pretreatment ameliorates oxidative stress induced apoptosis and DNA damage, alleviates the generation of intracellular reactive oxygen species (ROS) and restores mitochondrial membrane potential (MMP) in L-929 cells through its antioxidant potential. Together, these results suggest that SP can be exploited as a natural antioxidant in the food and pharmaceutical industries.
The potential of Averrhoa bilimbi pectin (ABP) as a source of biopolymer for edible film (EF) production was explored, and deep eutectic solvent (DES) (1% w/w) containing choline chloride-citric acid monohydrate at a molar ratio of 1:1 was used as the plasticizer. The EF-ABP3:1, which was produced from ABP with large branch size, showed a higher value of melting temperature (175.30 °C), tensile stress (7.32 MPa) and modulus (33.64 MPa). The EF-ABP3:1 also showed better barrier properties by obtaining the lowest water vapor transmission rates (1.10-1.18 mg/m2.s) and moisture absorption values (2.61-32.13%) depending on the relative humidity compared to other EF-ABPs (1.39-1.83 mg/m2.s and 3.48-51.50%, respectively) that have linear structure with smaller branch size. From these results, it was suggested that the galacturonic acid content, molecular weight, degree of esterification and pectin structure of ABP significantly influenced the properties of EFs. The interaction of highly branched pectin chains was stronger than the linear chains, thus reduced the effect of plasticizer and produced a mechanically stronger EF with better barrier properties. Hence, it was suggested that these EFs could be used as alternative degradable packaging/coating materials.
Stillbirth is a devastating event to the parents, relatives, friends, and families. The role of anticoagulants in the prevention of unexplained stillbirths is uncertain. An open-label interventional prospective cohort study was conducted on 144 women with a history of unexplained stillbirths. The intervention group had a high umbilical artery resistance index (RI) and received bemiparin. The nonintervention group had a normal RI and did not receive any intervention. We measured the adjusted odds ratio (OR) and 95% confidence interval (CI) of the main outcome for these variables using logistic regression analysis. Fresh stillbirth and early neonatal death rates were lower (P = .005, OR = 11.949 and 95% CI = 2.099-68.014) and newborn weight was higher (P = .015, OR = 0.048, 95% CI = 0.004-0.549) in the group that received bemiparin. Bemiparin is effective in decreasing the rate of stillbirth in women with a history of previous unexplained stillbirths.
The binary toxin from Lysinibacillus sphaericus has been successfully used for controlling mosquito-transmitted diseases. An activation step shortens both subunits BinA and BinB before their interaction with membranes and internalization in midgut cells, but the precise role of this activation step is unknown. Herein, we show conclusively using three orthogonal biophysical techniques that protoxin subunits form only monomers in aqueous solution. However, in vitro activated toxins readily form heterodimers. This oligomeric state did not change after incubation of these heterodimers with detergent. These results are consistent with the evidence that maximal toxicity in mosquito larvae is achieved when the two subunits, BinA and BinB, are in a 1:1 molar ratio, and directly link proteolytic activation to heterodimerization. Formation of a heterodimer must thus be necessary for subsequent steps, e.g., interaction with membranes, or with a suitable receptor in susceptible mosquito species. Lastly, despite existing similarities between BinB C-terminal domain with domains 3 and 4 of pore-forming aerolysin, no aerolysin-like SDS-resistant heptameric oligomers were observed when the activated Bin subunits were incubated in the presence of detergents or lipidic membranes.
The molecular mass distribution, amino acid composition and radical-scavenging activity of collagen hydrolysates prepared from collagen isolated from the sea cucumber Stichopus vastus were investigated. β and α1 chains of the collagen were successfully hydrolysed by trypsin. The molecular mass distribution of the hydrolysates ranged from 5 to 25 kDa, and they were rich in glycine, alanine, glutamate, proline and hydroxyproline residues. The hydrolysates exhibited excellent radical-scavenging activity. These results indicate that collagen hydrolysates from S. vastus can be used as a functional ingredient in food and nutraceutical products.
Texture and biochemical changes, specifically in moisture, protein content and amino acid profile were studied throughout the molt cycle of Panaeus monodon. This study was initiated to investigate changes that may occur in mass and tissue composition during different molting stages. Molting of Penaeus monodon are classified into 3 main stages; postmolt, intermolt and premolt. Result of biochemical analysis, shows that moisture varies from 78.02-80.88%, indicating a maximum value during postmolt and minimum value during premolt. Total protein content was found to be higher during intermolt (23.48%) and postmolt (22.27%) compared to premolt (23.10%). The result of SDS-PAGE electrophoresis shows a higher intensity of protein band during intermolt which corresponds to higher protein content. Thus, this electrophoresis method is useful as an alternative to determine different stages of shrimp molting. This is based on intensity of the protein band which is referring to protein content of the shrimp at its different stages. The amino acid profile showed that arginine, alanine, glutamic acid, glycine, lycine, and thronine increased during premolt whilst isoleucine and phenylalanine higher during postmolt. Aspartic acid and histidine was higher in intermolt than in premolt and postmolt. Texture analysis carried out give an elastic modulus value that is high during premolt compared to postmolt and intermolt. There are significant changes in texture and biochemical composition through elastic modules value and protein composition of Penaeus monodon during each stages of moltcycle.
Human skin contains numerous antigen-presenting cells that are a potential target for several immune-based therapies, including vaccination and cancer immunotherapy. However, the outermost layer of the skin-the stratum corneum-acts as a major physical barrier against the permeation of antigens that have a molecular weight > 500 Da. In this study, an ionic liquid-assisted delivery system (ILDS) was developed, which enabled the successful transdermal delivery of an antigenic protein, ovalbumin (OVA), with a toll-like receptor agonist, imiquimod, as an adjuvant, to stimulate a specific immune response. Both the ionic liquids and ILDS were completely biocompatible for topical or transdermal application for therapeutic purposes. The skin permeation of the antigenic protein and adjuvant was found to be significantly enhanced because of the incorporation of a surface-active ionic liquid in the ILDS. An in vivo immunization study showed that there was a high level of OVA-specific IgG antibody production because of the enhanced permeation of the antigen and adjuvant across and into the skin. In a preclusive anticancer study, vaccination through ILDS showed stronger tumor-growth inhibition compared to control group. These results indicated that the ILDS could be a promising strategy for transdermal immunization as future therapeutics.
This study aimed at purification of phycocyanin (PC) from Phormidium tergestinum using an aqueous two-phase system (ATPS) comprised of polyethylene glycol (PEG) and salts. The partitioning efficiency of PC in ATPS and the effect of phase composition, pH, crude loading, and neutral salts on purification factor and yield were investigated. Results showed that PC was selectively partitioned toward bottom phase of the system containing potassium phosphate. Under optimum conditions of 20% (w/w) PEG 4000, 10% (w/w) potassium phosphate, 20% (v/v) crude load at pH 7, with addition of 0.5% (w/w) NaCl, PC from P. tergestinum was partially purified up to 5.34-fold with a yield of 87.8%. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the molecular weight of PC was ∼19 kDa. Results from this study demonstrated ATPS could be used as a potential approach for the purification of PC from P. tergestinum.
Poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-4-hydroxybutyrate) [P(3HB-co-3HV-co-4HB)] terpolymer was produced using Cupriavidus sp. USMAA2-4 via one-step cultivation process through combination of various carbon sources such as 1,4-butanediol or γ-butyrolactone with either 1-pentanol, valeric acid, or 1-propanol. Oleic acid was added to increase the biomass production. The composition of 3HV and 4HB monomers were greatly affected by the concentration of 1,4-butanediol and 1-pentanol. Terpolymers with 3HV and 4HB molar fractions ranging from 2 to 41 mol.% and 5 to 31 mol.%, respectively, were produced by varying the concentration of carbon precursors. The thermal and mechanical properties of the terpolymers containing different proportions of the constituent monomers were characterized using gel permeation chromatography (GPC), DSC, and tensile machine. GPC analysis showed that the molecular weights (M (w)) of the terpolymer produced were within the range of 346 to 1,710 kDa. The monomer compositions of 3HV and 4HB were also found to have great influences on the thermal and mechanical properties of the terpolymer P(3HB-co-3HV-co-4HB) produced.
Palm kernel oil, palm olein, crude palm oil and palm acid oil were used for the synthesis of poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] by a mutant strain of Wautersia eutropha (formerly Ralstonia eutropha) harboring the Aeromonas caviae polyhydroxyalkanoate (PHA) synthase gene. Palm kernel oil was an excellent carbon source for the production of cell biomass and P(3HB-co-3HHx). About 87% (w/w) of the cell dry weight as P(3HB-co-3HHx) was obtained using 5 g palm kernel oil/l. Gravimetric and microscopic analyses further confirmed the high PHA content in the recombinant cells. The molar fraction of 3HHx remained constant at 5 mol % regardless of the type and concentration of palm oil products used. The small amount of 3HHx units was confirmed by 13C NMR analysis. The number average molecular weight (M(n)) of the PHA copolymer produced from the various palm oil products ranged from 27 0000 to 46 0000 Da. The polydispersity was in the range of 2.6-3.9.
The biosynthesis and characterization of medium chain length poly-3-hydroxyalkanoates (mcl-PHA) produced by Pseudomonas putida Bet001 isolated from palm oil mill effluent was studied. The biosynthesis of mcl-PHA in this newly isolated microorganism follows a growth-associated trend. Mcl-PHA accumulation ranging from 49.7 to 68.9% on cell dry weight (CDW) basis were observed when fatty acids ranging from octanoic acid (C(8:0)) to oleic acid (C(18:1)) were used as sole carbon and energy source. Molecular weight of the polymer was found to be ranging from 55.7 to 77.7 kDa. Depending on the type of fatty acid used, the (1)H NMR and GCMSMS analyses of the chiral polymer showed a composition of even and odd carbon atom chain with monomer length of C4 to C14 with C8 and C10 as the principal monomers. No unsaturated monomer was detected. Thermo-chemical analyses showed the accumulated PHA to be semi-crystalline polymer with good thermal stability, having a thermal degradation temperature (T(d)) of 264.6 to 318.8 (± 0.2) (o)C, melting temperature (T(m)) of 43. (± 0.2) (o)C, glass transition temperature (T(g)) of -1.0 (± 0.2) (o)C and apparent melting enthalpy of fusion (ΔH(f)) of 100.9 (± 0.1) J g(-1).
A combination of palm olein (POl) and fructose was used as carbon source for the biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] by recombinant Cupriavidus necator Re2058/pCB113. Cultures grown using 5g/L PO alone as carbon source produced cell dry weight (CDW) of 5.13g/L, 67% PHA/CDW and accumulated a copolymer containing 27mol% 3HHx in shake flask cultures. When cultures were grown in 5g/L fructose alone as the carbon source they produced CDW of 2.32g/L, 11% PHA/CDW and accumulated only poly(3-hydroxybutyrate) [P(3HB)] homopolymer. When the cells were cultured in 5g/L POl in combination with 7g/L fructose, CDW of 7.41g/L and 80% PHA/CDW was obtained with 17mol% 3HHx monomer fraction. Biosynthesis was carried out using a 13L fermenter to study the accumulation of 3HHx monomer fraction in the bacterial cells at different time point. The molecular weights of P(3HB-co-3HHx) with 4-15mol% 3HHx monomer were in the range between 5.47-6.85×105Da, which were at least two fold higher than previously reported values. Interestingly, the increase in Mwof the copolymer along with the increase in 3HHx molar fraction was observed. The viscoelastic property of the copolymer further confirmed the increase in Mw.
The potential use of n-dodecane and n-hexadecane as oxygen vectors for enhancing hyaluronic acid (HA) biosynthesis by Streptococcus zooepidemicus ATCC 39920 was investigated using a 2-L stirred-tank bioreactor equipped with helical ribbon or Rushton turbine impellers. The volumetric fraction of the oxygen vector influenced the gas-liquid volumetric oxygen transfer coefficient (K(L)a) positively. Batch HA fermentation with 1% (v/v) n-dodecane or 0.5% (v/v) n-hexadecane addition was carried out at different impeller tip speeds. Even though cell growth was lower in the fermentation with oxygen vector addition, the HA productivity and molecular weight were higher when compared to the fermentation without oxygen vector at low impeller tip speed. The highest HA concentration (4.25 gHA/l) and molecular weight (1.54 × 10(7) Da) were obtained when 0.5% (v/v) n-hexadecane and 0.785 m/s impeller tip speed of helical ribbon were used.
A halophilic bacterium isolated from mangrove soil sample in Northern Vietnam, Yangia sp. ND199 was found capable of producing homopolymer poly(3-hydroxybutyrate) [P(3HB)], copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)], and copolymer poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)] from different carbon sources. The presence of 3HB, 3HV, and 4HB monomers were confirmed by gas chromatography and nuclear magnetic resonance analysis. Only P(3HB) was produced using carbon sources such as fructose or by a combination of fructose with 1,5-pentanediol, 1,6-hexanediol, sodium hexanoate, or sodium octanoate. The biosynthesis of P(3HB-co-3HV) was achieved by adding cosubstrates such as sodium valerate and sodium heptanoate. When 1,4-butanediol, γ-butyrolactone or sodium 4-hydroxybutyrate was added to the culture medium, P(3HB-co-4HB) containing 4.0-7.1mol% 4HB fraction was accumulated. The molecular weights and thermal properties of polyesters were determined by gel permeation chromatography and differential scanning calorimeter, respectively. The results showed that Yangia sp. ND199 is able to produce polyester with high weight average molecular weight ranging from 1.3×10(6) to 2.2×10(6) Dalton and number average molecular weight ranging from 4.2×10(5) to 6.9×10(5) Dalton. The molecular weights, glass transition temperature as well as melting temperature of homopolymer P(3HB) are higher than those of copolymer P(3HB-co-3HV) or P(3HB-co-4HB).
Two-stage fermentation was normally employed to achieve a high poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)] productivity with higher 4HB molar fraction. Here, we demonstrated single-stage fermentation method which is more industrial feasible by implementing mixed-substrate cultivation strategy. Studies on bioreactor scale show a remarkably high PHA accumulation of 73 wt%, contributing to a high PHA concentration and product yield of 8.6 g/L and 2.7 g/g, respectively. This fermentation strategy has resulted in copolymers with wider range of 4HB monomer composition, which ranges from 12 to 55 mol%. These copolymers show a broad range of weight average molecular weight (M w ) from 119.5 to 407.0 kDa. The copolymer characteristics were found to be predominantly affected by the nature of the substrates and the mixture strategies, regardless of the 4HB monomer compositions. This was supported by the determination of copolymer randomness using (13)C-NMR analysis. The study warrants significantly in the copolymer scale-up and modeling at industrial level.
Green mussel (Perna viridis) was hydrolysed with alcalase under two different conditions consisting of pH7, E/S5% or pH 9, E/S 3% at 60°C for two hours. Hydrolysis at pH 9, E/S3% resulted in a higher degree of hydrolysis (DH) than pH7, E/S5% with degree of hydrolysis of 37.00% and 28.33%, respectively. The green mussel hydrolysates were characterized by molecular weight of
Low-molecular-weight heparins (LMWHs) are antithrombotic agents utilised in the treatment of acute coronary syndromes. They have been shown to be more effective than unfractionated heparins (UFHs) in reducing ischeamic events, which include death, myocardial infarction (MI) and urgent revascularisation. Enoxaparin is one of the products of LMWHs. Its safety and efficacy has been proven in the ESSENCE and TIMI IIB studies. This study was carried out to identify risk factors that may affect bleeding complications associated with the use of enoxaparin for non-ST-elevation MI (NSTEMI) or unstable angina (UA) in Universiti Kebangsaan Malaysia Hospital (HUKM). This observational, longitudinal study was conducted on patients who were admitted to the Coronary Care Unit (CCU), Coronary Rehabilitation Ward (CRW), Medical 1 and Medical 2 wards at HUKM and initiated on enoxaparin for NSTEMI/UA from 22nd of March until 22nd of April 2004. A total of 40 patients were included in the study with median age of 65 years, male to female ratio of 3:1, diagnosed with NSTEMI (55%) and UA (45%). 45% of patients developed an episode of bleeding and among them 83.3% (15 patients) characterised by haematuria. Higher percentages of women (80%) and those with creatinine clearance of < 30ml/min (100%) had incidence of bleeding as compared to men (50%) and those with creatinine clearance = 30 ml/min, respectively (p < 0.05 for both parameters). Age, enoxaparin dose and duration of therapy, smoking and concomitant aspirin/ticlopidine therapy did not significantly affect the incidence of bleeding. In conclusion, renal impairment and gender were associated with bleeding in relation with the use of enoxaparin that may require dose adjustments.
Host induced control of pathogens involves, but not limited to, withholding of essential transition metals as well as releasing the metals at a toxic level. Zinc is one of these transition elements that plays critical role in controlling the pathogens in that manner – a key mediator in nutritional immunity. A number of subcellular and molecular mecha-nisms such as transport and storage proteins are known to maintain Zn homeostasis and scuffle with the pathogens. Pathogenic bacteria also use a number of mechanisms to combat the scuffle and fight for the right amount of Zn for their survival and growth. From the host perspective, a “delicate” balance of Zn must be maintained for immune surveillance while making the level of Zn either to starve or to intoxicate the pathogens. Metallothionein (MT), a group of low molecular weight proteins, is well known for its Zn transport and storage ability and is expected to play an important role in that nutritional immunity. Zn homeostasis by MT to fight oral pathogens is not unexpected too. Periodontitis and dental caries are two most common oral diseases which are linked to the pathogenic carnival of opportunistic bacteria. Can those culprits be exterminated through nutritional immunity using MT? Or could it be - those human hosts who become the easy prey of those pathogens lack inducible expression of MT in their oral tissues? The synthesis or degradation of MT in response to invading pathogens in oral tissues, the human-MT medi-ated Zn homeostasis in response to infectious insult in oral tissues are evident. Nonetheless, the cross talk between MT and Zn in oral nutritional immunity is largely unknown.
Infection control and wound healing profiles of sodium carboxymethylcellulose (SCMC) films were investigated as a function of their anti-bacterial action, physical structures, polymer molecular weights and carboxymethyl substitution degrees. The films were prepared with in vitro polymer/film and in vivo microbe-colonized wound healing/systemic infection profiles examined. Adhesive high carboxymethyl substituted SCMC films aided healing via attaching to microbes and removing them from wound. Pseudomonas aeruginosa was removed via encapsulating in gelling low molecular weight SCMC film, whereas Staphylococcus aureus was trapped in tight folds of high molecular weight SCMC film. Incomplete microbe removal from wound did not necessary translate to inability to heal as microbe remnant at wound induced fibroblast migration and aided tissue reconstruction. Using no film nonetheless will cause systemic blood infection. SCMC films negate infection and promote wound healing via specific polymer-microbe adhesion, and removal of S. aureus and P. aeruginosa requires films of different polymer characteristics.