Manufacturing polymethyl methacrylate nanofibers as a support for enzyme immobilization

Nanofibers have a great potential for enzyme immobilization application due to their large surface area to volume ratio besides their porous structure. In this work, we produce polymethyl methacrylate (PMMA) nanofibers via electrospinning method in dimethylformamide (DMF) as solvent. Thereafter, we employ a chemical method on final PMMA nanofiberous web to covalently immobilize acetylcholinesterase (AChE) enzyme on membrane surface. Morphology and tensile properties of nanofibers are studied as first steps of characterization to make sure of obtaining a properly stable membrane for enzyme carrying application. Thereafter, the stability and activity of immobilized enzymes as two main characteristic parameters are tested and reported for different applications such as biosensor manufacturing.     

Evaluating the potential of wine-making residues and corn cobs as support materials for cell immobilization for ethanol production

Three wine-making residues (grape seeds, skins and stems), and corn cobs were evaluated as support material for immobilization of Saccharomyces cerevisiae and the ethanol production by the immobilized cells was assessed. The main objective of this study was to find an abundant and low cost material suitable for the cells immobilization and able to be used in a next step of wine production by immobilized yeast cells. The four natural materials were used as support in two different forms: untreated, and treated by a sequence of acid and basic reactions. Untreated grape skin and corn cobs provided the highest cell immobilization results (25.1 and 22.2 mg cells/g support, respectively). The maximum ethanol production yield (about 0.50 g/g) was also obtained when the cells were immobilized in these untreated materials. It was also found that the support materials released nutrients to the medium, which favored the yeast development and the ethanol production. The use of immobilized cells systems under agitated conditions gave ethanol yields similar to those obtained by the static fermentations, but the immobilized cell concentration was significantly lower. In brief, static fermentation with cells immobilized on grape skins or corn cobs appear to be an interesting alternative for use on wine-making. The use of grape skins, particularly, which is a by-product of the wine elaboration, could be of larger interest to obtain an integrated wine production process with by-product reuse.     

Covalent immobilization of urease to modified ethyl cellulose

Graft polymerization technology is a good way to modify polymers. New functional groups are added to polymer structure using graft polymerization. Enzyme immobilization could be done from these added functional groups. Ethyl cellulose was selected as a support for enzyme immobilization and no many studies has been conducted about it. It is insoluble in water and suitable for reuse. In this study, methacrylic acid was graft polymerized to ethyl cellulose using benzophenone. In graft polymerization studies, optimum amounts of methacrylic acid and benzophenone were determined as 60 mmol and 0.6 g, respectively. Percentage of graft polymerization was maximum in toluene as solvent and optimum graft polymerization time was found as 3 hours. The graft polymerization percentage was 225.7 % at optimum conditions. This value was very good for UV-induced graft polymerization technique. FT-IR spectra of ethyl cellulose and methacrylic acid graft polymerized ethyl cellulose showed that graft polymerization was carried out successfully. -COOH groups were added to ethyl cellulose structure after graft polymerization. Then, urease was immobilized to methacrylic acid graft polymerized ethyl cellulose using 1-ethyl-3-(3-dimetylaminopropyl)-carbodiimide as the condensing agent which accelerates the reaction between -COOH from methacrylic acid graft polymerized ethyl cellulose and -NH2 from urease. Optimization studies were also performed for the immobilization of urease. Optimized values for urease immobilization; optimum amount of 1-ethyl-3-(3-dimetylaminopropyl)-carbodiimide was found as 5 mg, temperature was determined as 37 °C, 2 hours were selected as optimum time, pH and amount of urease were found to be pH 7 and 10 mg, respectively. Remained activity of immobilized urease was 1.74 % before optimization studies. After optimization of immobilization studies, this ratio has increased to 29.85 %. The immobilized urease activity was increased 17.2 times.     

Nano immobilization of enzyme to improvement of biofuel cell electrode's function

This study centers on development of the enzyme presentation as a cathode catalyst. Laccase enzyme is immobilized at nano meter scale in quaternary ammonium bromide salt treated Nafion polymers and assorted with high surface area carbon powders that are then deposited onto a solid support to create the cathode electrode. Optimization of the synthesis circumstances resulted in notably enhanced performance toward oxygen reduction reaction.     

Physico-chemical behaviour of the fish lipid from Lepidocephalus guntea (Hamilton) and variation of lipid profile with size

The fish lipid was extracted from the body muscle (edible portion) of fresh Lepidocephalus guntea (Ham.) fish and the lipid content of the fish in size-1 (below 8.5 cm), size-2 (8.5-9.5 cm) and size-3 (above 9.5 cm) were found to be 2.515, 3.013 and 3.455%, respectively. The specific gravity and refractive index of the lipid were found to be 0.93 at 30 degrees C and 1.467 at 31.5 degrees C, respectively. The saponification value, saponification equivalent, iodine value, peroxide value and acetyl value of the lipid were found to be 220.325, 254.624, 96.05, 1.993 and 11.32, respectively. The acid value, percentage of free fatty acid as oleic and unsaponifiable matter present in the lipid were found to be 2.005, 1.008 and 0.593, respectively. The fatty acid composition of the lipid was determined qualitatively and quantitatively by TLC and GLC. The analysis revealed that the fatty acid composition of the lipid lies between C14 to C20. The fish lipid of Lepidocephalus guntea (Ham.) was found to contain (average value) myristic acid (3.17%), palmitoleic acid (7.45%), palmitic acid (29.16%), linolenic acid (7.13%), linoleic acid (5.57%), oleic acid (22.93%), stearic acid (17.42%), arachidonic acid (7.17%) and arachidic acid in trace amount.     

Application of electrospun silk fibroin nanofibers as an immobilization support of enzyme

Silk fibroin (SF) nanofibers were prepared by electrospinning and their application as an enzyme immobilization support was attempted. By varying the concentration of SF dope solution the diameter of SF nanofiber was controlled. The SF nanofiber web had high capacity of enzyme loading, which reached to 5.6 wt%. The activity of immobilizedα-chymotrypsin (CT) on SF nanofiber was 8 times higher than that on silk fiber and it increased as the fiber diameter decreased. Sample SF8 (ca. 205 nm fiber diameter) has excellent stability at 25°C by retaining more than 90 % of initial activity after 24 hours, while sample SF11 (ca. 320 nm fiber diameter) shows higher stability in ethanol, retaining more than 45% of initial activity. The formation of multipoint attachment between enzyme and support might increase the stability of enzyme. From these results, it is expected that the electrospun SF nanofibers can be used as an excellent support for enzyme immobilization.     

Genetic diversity and population structure of Pisum sativum accessions for marker-trait association of lipid content

Field pea(Pisum sativum L.) is an important protein-rich pulse crop produced globally. Increasing the lipid content of Pisum seeds through conventional and contemporary molecular breeding tools may bring added value to the crop. However, knowledge about genetic diversity and lipid content in field pea is limited. An understanding of genetic diversity and population structure in diverse germplasm is important and a prerequisite for genetic dissection of complex characteristics and marker-trait associations. Fifty polymorphic microsatellite markers detecting a total of 207 alleles were used to obtain information on genetic diversity, population structure and marker-trait associations. Cluster analysis was performed using UPGMA to construct a dendrogram from a pairwise similarity matrix. Pea genotypes were divided into five major clusters. A model-based population structure analysis divided the pea accessions into four groups. Percentage lipid content in 35 diverse pea accessions was used to find potential associations with the SSR markers. Markers AD73, D21, and AA5 were significantly associated with lipid content using a mixed linear model(MLM) taking population structure(Q) and relative kinship(K) into account. The results of this preliminary study suggested that the population could be used for marker-trait association mapping studies.     

Plasma lipid lowering effects of wheat germ in hypercholesterolemic subjects

The present study was performed to investigate the possible effects of wheat germ supplementation on lipid metabolism in humans. Ten free-living adult subjects participated in the study. None was obese or diabetic. They all presented an, hypercholesterolemia (from 6.58 to 9.50 mM), associated in 6 over 10 cases to an hypertriglyceridemia (from 1.70 to 5.00 mM). The subjects were studied in three consecutive periods, during which they first were on their usual diet (first week), they then ingested a daily supplement of 30 g wheat germ (4 weeks) and then they returned to their usual basal diet (4 weeks follow-up). Dietary records were obtained for 7 and 3 consecutive days before and during wheat germ supplementation, respectively. Fasting blood samples were taken at the end of each period. After 4 weeks of wheat, germ intake, glycemia did not change while total plasma cholesterol significantly decreased (paired Student'st test,p0.05) from 7.80 to 7.15 mM. LDL and HDL cholesterol values did not show marked changes, but VLDL cholesterol significantly dropped by 40.6%. Thus, the plasma/HDL total cholesterol ratio was significantly lower. Apoprotein B and A1 decreased. In the hypertriglyceridemic subjects, this was accompanied by a significant reduction of plasma triglycerides (1.64 vs. 2.68 mM) and a marked drop of VLDL triglycerides (–51%). Taken as a whole, the present results obtained in humans are very close to those previously obtained in the rat [40] and point out that wheat germ may play a beneficial role in the dietary management of hyperlipidemia.Part of the data communicated in this paper was presented at the Fibre 90 Conference, 17–20 April 1990, Norwich, UK.     

Variation of wheat grain lipid fraction and its antioxidative status under the impact of delayed sowing

This study determined the profile of hydrophobic phytochemicals (steroid-like components and lipophilic phenolics) and antioxidant potential (DPPH and Rancimat tests) of wheat grain in relation to sowing date (standard vs. delayed) using six cultivars of Triticum aestivum wheat grown in Poland. The study found that “sowing time” generally had a low impact on sterols, carotenoids, squalene and total lipophilic phenolics in wheat grain. The highest impact of “sowing time” was noted for α-tocopherol, C19:0 alkylresorcinol and campesterol. Delayed sowing reduced their content by up to 9%. The antioxidant potential of grain extracts obtained by the use of water-saturated butanol (WSB) was mostly cultivar-dependent (depending on assay: DPPH 56.3% and Rancimat 75.1% of explained variation, respectively). Wheat grain WSB extracts increased rapeseed oil induction time by up to 21% and their antioxidant capacity was up to 1.24 μM TEg⁻¹.     

Puroindolines co-localize to the starch granule surface and increase seed bound polar lipid content

Endosperm texture in wheat is controlled by the Pina and Pinb genes that comprise the Hardness (Ha) locus. Studies have shown that soft and hard varieties differ in the amount of starch bound polar lipids but have not addressed whether PINs are directly involved and whether the presence of one particular PIN affects seed polar lipid levels and cellular localization. Here, we determined the effect of overexpressing PINA or PINB on seed bound polar lipids and PIN localization. F₃ recombinants homozygous for either a Pina or Pinb null Ha locus with or lacking a transgenically added Pina or Pinb were analyzed for grain hardness, PIN abundance, and seed bound polar lipid levels. Overexpressed PINs resulted in reduced hardness, increased starch bound PINs, and increased seed bound polar lipids. Addition of PINA to the PINA nulls or PINB to the PINB nulls resulted in higher bound polar lipid levels than the addition of the alternative PIN. Both PINs localized to the starch surface in the presence or absence of the other protein. Our results indicate that PIN overexpression results in reduced endosperm texture and increased seed bound polar lipids and that PINs independently localize to the surface of starch granules.     

Antioxidant characteristics of phenolipids (quercetin-enriched lecithin) in lipid matrices

A novel phenolipids formulation containing a complex of quercetin and soy lecithin was developed. Antioxidant and antiradical characteristics of native soy lecithin and phenolipids’ formulations (complexes of quercetin and lecithin, 1:99 and 3:97, w/w) in the protection of triolein and sunflower oil (SFO) models stored under accelerated oxidative conditions for 15 day at 60 °C were studied. Progress of oxidation was monitored by recording the ultraviolet absorptivity and measuring the formation of oxidative products (peroxide value). The antiradical action of different lipid matrices against DPPH· and galvinoxyl radicals was screened during Shaal oven test. The factors influencing the oxidative stability (OS) of different models were also discussed. Inverse relationships were noted between peroxide values and oxidative stabilities at termination of the storage. Absorptivity at 232 nm and 270 nm in models containing lecithin increased gradually with the increase in time, due to the formation of conjugated dienes and polyenes. In general, oxidative stabilities of models enriched with phenolipids’ formulations were better than in models containing lecithin or quercetin alone, most likely as a consequence of synergism between polar lipids and quercetin as well as tocopherols. Moreover, increases in concentration of quercetin in phenolipid formulations resulted in an increase in its antioxidative potential. These results will be useful for improving the antioxidative activity, health impact and functionality of polar lipids and phenolic compounds in different food and pharmaceutical applications.     

Characterization and stability of nanostructured lipid carriers as drug delivery system

Recently more focus has been put to the development of innovative drug-delivery systems that includes polymer nanoparticles, emulsions and liposomes and solid lipid nanoparticles (SLNs). The SLNs have been proposed to be an alternative colloidal drug delivery system. The aim of this study was preparation and characterization of solid lipid nanoparticle (SLN) using varieties of emulsifier for encapsulation of the drug with poor water solubility. In these study four types of solid lipid nanoparticles were prepared based on different compositions of palm oil (S154) and lecithin (Lipoid 100) using the high pressure homogenization method. The SLN formulation had the following (palm oil+lecithin) compositions: SLN-01 (90 + 10%, respectively), SLN-02 (80 + 20%, respectively), SLN-03 (70 + 30%, respectively) and SLN-04 (60 + 40%, respectively). The SLNs were characterized and the optimum stability factors for one year storage determined. The parameters used to characterize the SLNs were particle size and polydispersity index (particle sizer), zeta potential (zetasizer), crystallinity (differential scanning calorimetry and wide angle X-ray diffraction), ultrastructure (transmission electron microscopy). Varying the palm oil and lecithin compositions resulted in SLNs of variable sizes and zeta potentials. The particle sizes of SLN-01, SLN-02, SLN-03 and SLN-04 were 298.40 +/- 11.80, 255.40 +/- 3.20, 145.00 +/- 3.39 and 273.00 +/- 86.50 nm, respectively, while the zeta potentials were -19.44 +/- 60.00, -19.50 +/- 1.80, -17.83 +/- 10.00 and -13.33 +/- 2.30 mV, respectively. Thermoanalysis and X-ray diffraction analysis showed that the SLNs had lower crystallinity than bulk lipid. The SLNs were generally round and uniform in shape under transmission electron microscopy. The SLN dimensional data suggested they had high quality physicochemical characteristics, which are conducive for the loading of poor water solubility drugs.     

Synchrotron X-ray scattering analysis of the interaction between corn starch and an exogenous lipid during hydrothermal treatment

Lipids have an important effect on starch physicochemical properties. There exist few reports about the effect of exogenous lipids on native corn starch structural properties. In this work, a study of the morphological, structural and thermal properties of native corn starch with L-alpha-lysophosphatidylcholine (LPC, the main phospholipid in corn) was performed under an excess of water. Synchrotron radiation, in the form of real-time small and wide-angle X-ray scattering (SAXS/WAXS), was used in order to track structural changes in corn starch, in the presence of LPC during a heating process from 30 to 85 °C. When adding LCP, water absorption decreased within starch granule amorphous regions during gelatinization. This is explained by crystallization of the amylose-LPC inclusion complex during gelatinization, which promotes starch granule thermal stability at up to 95 °C. Finally, a conceptual model is proposed for explaining the formation mechanism of the starch-LPC complex.     

Effect of lipid incorporation on functional properties of wheat gluten based edible films

Wheat gluten is an inexpensive protein derived from mill industries with good film forming properties, which allows producing semipermeable membranes able to slow down water migration in foods.The first objective of this study was to evaluate the effects of the incorporation of a lipid phase (25 wt%, dry basis) in wheat gluten on the functional properties of the film, such water sorption, surface hydrophilicity, water barrier properties, mechanical properties and thermal properties. The second one was to asses if such incorporation was able to reduce the water sensitivity of film mechanical properties.Findings clearly showed that the incorporation of a lipid phase was able to decrease the water sorption, water affinity (hydrophilicity) and water transfer (≈2 times) of wheat gluten films. Moreover, mechanical properties are also affected by the lipid addition with a decrease in rigidity and, at high a_w, an increase in extensibility. However, the sensitivity of the mechanical properties to water was not modified. Lastly, DSC (Differential Scanning Calorimetry) analysis proved that changes in mechanical properties of films as a function of hydration state were the consequence of glass transition depletion, which allowed them to turn from a glassy-like behavior to a rubber-like behavior.     

Characterization of supported lipid bilayers incorporating the phosphoinositides phosphatidylinositol 4,5-biphosphate and phosphoinositol-3,4,5-triphosphate by complementary techniques

Phosphoinositides are involved in a large number of processes in cells and it is very demanding to study individual protein-lipid interactions in vivo due to their rapid turnover and involvement in simultaneous events. Supported lipid bilayers (SLBs) containing controlled amounts of phosphoinositides provide a defined model system where important specific recognition events involving phosphoinositides can be systematically investigated using surface sensitive analytical techniques. The authors have demonstrated the formation and characterized the assembly kinetics of SLBs incorporating phosphatidylinositol 4,5-biphosphate (PIP(2); 1, 5, and 10 wt?%) and phosphoinositol-3,4,5-triphosphate (1 wt?%) using the quartz crystal microbalance with dissipation monitoring and fluorescence recovery after photobleaching. An increased fraction of phosphoinositides led to a higher barrier to liposome fusion, but full fluidity for the phosphatidylcholine lipids in the formed SLB. Significantly, the majority of phosphoinositides were shown to be immobile. X-ray photoelectron spectroscopy was used for the first time to verify that the PIP(2) fraction of lipids in the SLB scales linearly with the amount mixed in from stock solutions.     

Extension of green bell pepper shelf life using oilseed-derived lipid films from soapstock

Edible films have been used for decades on fresh produce to create a semipermeable membrane on the surface to suppress respiration, control moisture loss, and more recently to provide a delivery mechanism for the inclusion of functional components. Scientists at the Southern Regional Research Center (SRRC) have previously demonstrated that a thin biodegradable film can be produced from soapstock, an underused byproduct from the vegetable oil industry. After physical and chemical treatments, a thin film was produced from various soapstocks (cottonseed and safflower). Different hydration ratios were tested since the initial soapstock solutions were rather viscous. To examine the potential use of an oilseed-derived lipid film for the extension of shelf life, different types of the oilseed-derived soapstocks were utilized to produce lipid films with different hydration ratios, and containing 0, 5, and 10% of paraffin wax for application on ‘Camelot’ bell peppers. Control bell peppers lost almost 25% weight per unit surface area (SA) in 78 h when stored under ambient conditions. Cottonseed film-coated peppers, hydrated at 1:4, lost only about 5% moisture per unit SA after 78 h and minimized weight loss by up to 79% compared to the control. However, since a 1:4 hydration ratio remained rather viscous, 1:8 was preferred and these cottonseed films reduced weight loss per unit SA by up to 48% during storage. Safflower-derived soapstock film resulted in the least effective water retention of the films and ratios tested, with roughly 21–25% reduction in weight loss per SA compared to controls. Safflower-derived soapstock was higher in unsaturated fatty acids, which are less efficient to control moisture migration because they are more polar than saturated lipid materials, as contained in cottonseed-derived materials. Addition of wax to the cottonseed-derived films decreased water loss slightly, similar to previous reports in the literature. An ANOVA supported the conclusion that the oilseed-derived lipid films significantly reduced moisture loss across the produce epidermis. To avoid potential allergenicity concerns in cottonseed soapstock, additional cleanup steps and tests with commonly used edible coating additives would be required before attaining food grade status.     

Characterization of the effect of high salinity on roots of Chloris gayana Kunth: carbohydrate and lipid accumulation and growth

Branched nodal roots comprise the largest portion of the root system mass in Chloris gayana Kunth and the effects of high salinity on nodal root appearance and elongation rates were analysed in cv. Boma in greenhouse experiments. Roots from salt-treated plants (0·2 mol l^–1 NaCl) were smaller than controls, and accumulated higher concentrations of soluble sugars and reserve lipids. The number of nodal roots was reduced by the saline treatment. Leaf, tiller and nodal root appearance were delayed by salinity but the correlation among these processes was maintained, indicating that the developmental pattern was not altered by this level of salinity, only its rate. Initial nodal root growth rates varied as a function of plant size and were decreased by salinity only after 2 weeks of treatment. When shoots of non-salinized plants were cut, a drastic reduction in nodal root appearance was observed, suggesting emerging leaves were stronger sinks than roots for available reserves. However, when the shoots of salt-treated plants were cut, the already depressed root appearance rate was not further reduced. This suggests that, under salinity, alterations in root ability to metabolize reserves could have been more significant than reserve availability for controlling elongation.